Hippocratic Jan-Mar 2015 Journal of Unani Medicine Volume 10 1 Jan-Mar 2015

HIPPOCRATIC
JOURNAL OF
UNANI MEDICINE
Volume 10, Number 1, January - March 2015

Hippocratic J. Unani Med. 10(1): 1 - 136, 2015
CENTRAL COUNCIL FOR RESEARCH IN UNANI MEDICINE

Ministry of Ayurveda, Yoga & Naturopathy, Unani, Siddha and Homoeopathy (AYUSH)
Government of India
Hippocratic Journal of Unani Medicine
Chief Patron
Minister of AYUSH, Government of India
Patron
Secretary, Ministry of AYUSH, Government of India
International Advisory Board

Prof. G.N. Qazi, New Delhi, INDIA Prof. Talat Ahmad, New Delhi, INDIA
Prof. Ranjit Roy Chaudhury, Delhi, INDIA Hakim Syed Khaleefathullah, Chennai, INDIA
Dr. Fabrizio Speziale, Paris, France Dr. Suraiya H. Hussein, Kuala Lumpur, MALAYSIA
Mrs. Sadia Rashid, Karachi, PAKISTAN Prof. Allauddin Ahmad, Patna, INDIA
Prof. Ikhlas A. Khan, USA Dr. Maarten Bode, Amsterdam, THE NETHERLANDS
Prof. Abdul Hannan, Karachi, PAKISTAN Prof. Usmanghani Khan, Karachi, PAKISTAN
Prof. Rashid Bhikha, Industria, SOUTH AFRICA Dr. S.S. Handa, Haryana, INDIA
Prof. Ram Vishwakarma, Jammu, INDIA Prof. Irfan Ali Khan, Hyderabad, INDIA
Editorial Board
Prof. Wazahat Husain, Aligarh Prof. V.H. Talib, Dehradun
Dr. (Mrs.) Nandini Kumar, New Delhi Prof. K.M.Y. Amin, Aligarh
Dr. O.P. Agarawal, Delhi, INDIA Dr. A.B. Khan, Aligarh
Prof. Y.K. Gupta, New Delhi Dr. (Mrs.) Neena Khanna, New Delhi
Prof. A. Ray, Delhi, INDIA Dr. (Mrs.) Yasmeen Shamsi, New Delhi
Dr. S. Asad Pasha, New Delhi Dr. Mohammad Khalid Siddiqui, Faridabad
Prof. S. Shakir Jamil, New Dlehi Dr. Ghufran Ahmed, Aligarh
Prof. Mansoor Ahmad Siddiqui, Bengaluru Dr. M.A. Waheed, Hyderabad
Editor-in-Chief
Prof. Rais-ur-Rahman
Director General
Central Council for Research in Unani Medicine (CCRUM)
Associate Editor
Dr. Khalid M. Siddiqui, Deputy Director General, CCRUM
Assistant Editors
Dr. Wasim Ahmed Azmi, Deputy Director, CRIUM, Lucknow Dr. Munawwar Hussain Kazmi, Deputy Director, CRIUM, Hyderabad
Dr. Shariq Ali Khan, Research Officer, RRIUM, Aligarh Mr. Aminuddin, Research Officer (Botany), CCRUM
Mr. Shamsul Arfin, Research Officer (Chemistry), CCRUM Mr. Mohammad Niyaz Ahmad, Research Officer (Publication), CCRUM
Managing Editor
Dr. V.K. Singh, Consultant (Botany), CCRUM
Editorial Office
CENTRAL COUNCIL FOR RESEARCH IN UNANI MEDICINE
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On behalf of Central Council for Research in Unani Medicine (CCRUM) published and printed by Prof. Rais-ur-Rehman
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Contents
1. Therapeutic Evaluation of Unani Coded Drug UNIM-104 in Cases of Non-Alcoholic Fatty .................... 1
Liver Disease (NAFLD) A Preliminary Clinical Trial
Hafiz C. Mohammad Aslam, Zaheer Ahmed, Shahida Begum, Aijaz Ahmad, K. Kabiruddin,
Shaikh Nikhat, Javed Ali, Abdul Kareem and Syed Jameeluddin
2. Exploration of Mechanism of Antiinflammatory Action of Habbe-Gule-Aakh ............................................. 9

Musarrat Nafees, N.A. Khan, K.M.Y. Amin and Ghufran Ahmad
3. Toxicity Study of Qurs-e-Hudar in Experimental Animals ......................................................................... 15

Mohd. Nadeem, Mohd. Urooj, Habibur Rahman and Shariq Ali Khan
4. Comparative Analytical Study of the Efficacy of Unani Formulations with Reference to Oligospermia 23
M. Anas and M.M.H. Siddiqui
5. Re-designing of the Unani Preparation From Powder Form into Tablet and its Standardization .......... 31
Along with HPLC Profile
Aziz ur Rahman and Tajuddin
6. Development of Standard Operative Procedure (SOP) of a Toothpaste Prepared with the .................. 43
Ingredients of Sunun Poste Mughilan
Mohammad Rashid, Shariq Shamsi and Roohi Zaman
7. A Review on Phytochemistry, Pharmacological Properties and Biotechnological Studies in .................. 53

Valeriana officinalis L., An Important Medicinal Herb
Anwar Shahzad and Taiba Saeed
8. Some Depletive Plants of Kumaon Himalaya Used in Unani Medicine for Treating ............................. 73
Non-Communicable Diseases and Their Conservation Strategies
Zaheer Anwar Ali, Sarfraz Ahmad and Shariq Ali Khan
9. Quality Evaluation of Jawarish-e-Ood Kibreet ........................................................................................... 85

Rampratap Meena, D. Ramasamy, S. Mageswari, P. Meera Devi Sri, Shamsul Arfin and Aminuddin
10. Standardization of Unani Drug Post-e-Anar (Fruit rind of Punica granatum L.) ..................................... 95
Nazish Siddiqui, M. Masihuzzaman Ansari, Alvia Khan, Mohd. Bilal Tafseer,
Saba Viquar and Abdul Haleem
11. A Study on Diversity of Unani Medicinal Plants Used for Non-Communicable Diseases in ............... 105
Southern Western Ghats of Tamil Nadu
R. Murugeswaran, K. Venkatesan, Aijaz Ahmed and Aminuddin
12. Pharmacognostical Standardization of Flowers of Althaea officinalis Linn. (Gul Khatami) ................... 117
Nitin Rai, Rampratap Meena, S. Mageswari, Shamsul Arfin, Aminuddin and Rajeev Kr. Sharma
13. Development of Standards of Sapistan (Cordia dichotoma Forst. f.) .................................................... 129
Abdul Haleem, Abdul Latif, Abdur Rauf, Nazish Siddiqui and Sumbul Rehman
Instructions to Contributors
Editorial
The world-wide interest in traditional medicine has led to renewed efforts to study their medical efficacy and
safety on scientific lines. This is particularly true since the dangers and shortcomings of modern medicines have
started getting more apparent. Currently there had been many advances in the strategies for discovery and
evaluation of drugs, particularly plant based. The input of bio-chemistry to pharmacology followed by clinical
trials puts more emphasis on the mode of action of drugs, albeit it becomes clear that the activities of most
drugs are not confined to one single mode of action. This has led to generating lot of new researches and there
is, therefore, an enormous need for exchange of this vital information for wider use by scientific community
across the world engaged in the search of new drugs of plant origin.
Although the Unani medicines have been in use for centuries and are known for their therapeutic efficacies,
there is a need to scientifically establish their efficacy and safety in order to achieve global acceptance.
Organized research work in this system is, therefore, a need of the hour. In this direction, Central Council for
Research in Unani Medicine, through its clinical, drug research, literary research, survey & cultivation of
medicinal plants programme is contributing significantly for last three decades. Vitiligo, sinusitis, filariasis,
eczema, malaria, infective hepatitis, asthma are some of the conditions where Unani therapies have earned
recognition after scientific validation.
The Council has been publishing the peer reviewed Hippocratic Journal of Unani Medicine (HJUM), mainly to
bring out fundamental and applied aspects of Unani Medicine. The journal also publishes recent advances in
other related sciences and traditional medicines as well as different streams of medical sciences, which have
bearing on validation and scientific interpretation of various concpts and strengths of Unani medicine.
In view of an overwhelming response, the journal earlier published twice a year, its periodicity had been
changed to quarterly w.e.f. January 2008 to accommodate more articles for quick dissemination of research
data among scientific community. The journal has sufficient room for invited articles from luminaries of modern
medicine and sciences as well as scholars of Unani medicine. The broad areas being covered include clinical
research on single and compound Unani drugs, validation of regimental therapy, Clinical and experimental
pharmacological studies, standardization of single and compound drugs, development of standard operating
procedures, ethnobotanical studies, experimental studies on medicinal plants and development of agro-techniques
thereof, and literary research on classics of Unani medicine. The journal is also open for studies on safety
evaluation of Unani and other herbo-mineral drugs, nutraceuticals, cosmotherapeutics, aromatics, oral health,
life style disorders, sports medicine etc. and such other newer areas which are the outcome of modern day
living.
The current issue of this journal provides 13 original and review papers in the areas of clinical research, drug
standardization, pharmacology, ethnobotanical surveys and allied disciplines contributed by eminent scholars in
their respective fields. It is hoped that data presented will contribute significantly in R&D sector of traditional
drugs and prove to be an excellent exposition of current research efforts of scientists in this direction. Council
acknowledges the authors for their contributions included in this issue and hope for their continued support in
this endeavor. We wish to ensure the readers to bring out the future issues of the journal on time.
We at the CCRUM have been constantly striving to reach to higher standards and make HJUM the leading
journal of Unani medicine and related sciences. In this context, we thank our learned reviewers for their
invaluable inputs in improving the manuscripts. We sincerely hope and trust that the mission can be accomplished
with active partnership of quality-conscious individuals and institutions. Through these lines we seek your
cooperation and support in materializing our dreams about the HJUM. In this regard, we request you for your
as well as your colleagues contributions for publication in and subscription to the journal. Further, we will
appreciate if the journal is introduced far and wide. We would also welcome esteemed suggestions for achieving
the highest standards of quality for the journal.
April 22, 2015
(Prof. Rais-ur-Rahman)
Editor-in-Chief
Therapeutic Abstract

Evaluation of atty liver disease is caused by an accumulation of fat in the

Unani Coded
Drug UNIM-104
in Cases of
F

hepatocytes. Non Alcoholic Fatty Liver Disease (NAFLD) has been reported
worldwide, estimated to be around 1024% in various populations. There is no
standard medical treatment specifically for Non-alcoholic, non-Diabetic and non-
obese Fatty liver Disease, hence a preliminary clinical study has been conducted
at RRIUM Chennai, to evaluate the safety and efficacy of the Unani coded drug
Non-Alcoholic

UNIM-104 in the treatment of NAFLD. It was observed that there was reduction in

Fatty Liver the sign and symptoms at the end of trial with statistical improvement (p< 0.05) in

Hounz field (HF) Units value on CT scan of liver.

Disease

(NAFLD) A

Keywords: Non-Alcoholic Fatty Liver Disease (NAFLD), Fatty liver, Tashhamul

kabid, Unani Medicine, UNIM 104 coded drug.

Preliminary

Clinical Trial

Introduction

*Hafiz C. Mohammad Aslam,

Non-alcoholic fatty liver disease (NAFLD) is a common clinic-pathological condition

Zaheer Ahmed,

characterized by significant lipid deposition in the hepatocytes of the liver

Shahida Begum,

Aijaz Ahmad,
parenchyma. The pathological picture bears a striking resemblance to that of

K. Kabiruddin, alcohol-induced liver injury, but it occurs in individuals who deny a significant

Shaikh Nikhat, history of alcohol ingestion. It comprises of a wide spectrum of liver damage,

Javed Ali, ranging from simple macrovesicular steatosis to steatohepatitis, advanced fibrosis,

Abdul Kareem
and cirrhosis (Angulo, 2002; Angulo et al., 2003). Subsets of NAFLD which

and
progress to cirrhosis are being increasingly recognized as a major cause of liver-

Syed Jameeluddin

related morbidity and mortality with the potential to progress to liver failure (David,

2005). The term non-alcoholic steatohepatitis, (NASH) and several other terms

Regional Research Institute of

Unani Medicine,
have been used to refer to this entity, including pseudo alcoholic liver disease,

1 West Mada, Church Street,

alcohol-like hepatitis, diabetic hepatitis, non-alcoholic Laennecs disease, and

Royapuram, Chennai-600013

steatonecrosis (Ludwig et al., 1980). However, seeing that the disease represents

a spectrum of pathology, the umbrella term NAFLD, was first introduced in 1986,

became the preferred one (Sheth et al., 1997). The spectrum ranges from simple

benign fatty liver (hepatic steatosis) to NASH, characterized by fatty change with

lobular inflammation, hepatocellular injury, and Mallory hyaline, progressive

fibrosis, and cirrhosis, and has been defined both histologically and clinically

(Tominaga et al., 1995).

NAFLD has been reported worldwide but geographic variations in prevalence are

evident. Although the worldwide prevalence has not yet been determined, it has

been quoted as 1024% in various populations (Angulo, 2002; Angulo et al., 2003)

and it is estimated to be the most common liver disease in the western world, and

*Author for correspondence

Hippocratic Journal of Unani Medicine 1

January - March 2015, Vol. 10 No. 1, Pages 1-7
its prevalence is likely to increase (Tominaga et al., 1995). It affects all racial and

ethnic groups and has no age or sex predilection.

According to Unani system of medicine, the Kabid (liver) is the largest glandular

organ in the body, It breakdown the nutrient materials, participate in the growth

and development of human body and repair the wear and tear of the body (Badal-

e-Ma-Yatahallal). According to Ibn-e-Sina, Kabid (liver) is considered the seat or

centre of Quwa-Al-Tabiyah (Natural Faculty) (Ibn-e-Sina 1906). It is responsible

for the absorption, digestion, retention, metabolism and assimilation of food and

excretion of Fudhlat (waste products) (Abusahal Masihi 810-895 A.D). In Unani

classical text, Su-e-Mizaj-e-Kabid (alteration in normal temperament of liver)

particularly due to kasrate buroodat (excessive coldness) somewhat resembles

to NAFLD, in which liver is loaded with Fudhlat (waste products) and toxins which

ultimately help in deposition of Raddi Shaham (abnormal excessive fats) (Ibn-e-

Rushed, 1126-1198).

Till now there is no standard medical treatment for fatty liver diseases although

several possible treatments are under investigation including Vitamin E and C,

Ursodiol and other medications.9 In contrast Unani literature is replete with drugs

administered for fatty liver like conditions which has the potential to detoxify the

liver. However, considering necessity of novel medical intervention, the Central

Council for Research in Unani Medicine, Ministry of Ayush, Govt. of India, had

initiated a preliminary clinical study to test the safety and efficacy of the Unani

coded drug UNIM-104 in the treatment of non-alcoholic, non-obese and non-

diabetic fatty liver disease.

Materials and Methods

The study was a single blind, prospective clinical trial approved by the Institutional

Ethical Committee of the Institute. Altogether 35 cases from GOPD of RRIUM,

Chennai were enrolled in the study during the period 2010-2012 after getting

voluntary informed consent from the participants. However, 5 patients were lost

to follow up due to personal reasons. Subjects of either gender in the age group

of 40-60 years, non-alcoholic, non-diabetic & non-obese, with or without sign &

symptoms like fatigue, loss of appetite, malaise, dull ache in upper right abdomen,

nausea, hepatomegaly and BMI 29.9 were included in the study. Diagnosis and

grading of fatty liver was done on the basis of Ultra sonogram and CT Scan of

upper abdomen. Subjects with pregnancy or lactation, those unwilling to give

consent to abide by the protocol, systemic illness (cardiac, gastric, hepatic, renal,

neurological disorders etc) were excluded from the study.

Clinical assessment and laboratory investigations were carried out at baseline

and at 30 days interval and CT scan of upper abdomen for measuring liver Hounz


field (HF) unit at base line and end of the study to monitor the prognosis. The

Unani coded drug UNIM-104 was given in form of Majoon in a dosage of 5 gm

twice a day with water before food for a period of 90 days. Patients were advised

strictly to refrain from consumption of alcohol, high carbohydrate and high fat

content diet.

Following investigations were carried out using standard laboratory techniques.

Blood: Heamogram with ESR, Fasting Blood Sugar, S. Bilirubin, SGOT, SGPT,

S.Alkaline Phosphatase, S. Urea, S. Creatinine, S.Cholestrol, S.Triglycerides,

HDL, LDL.

Urine: Routine and Microscopical examination.

Statistical analysis of the data was carried out using Student t test (paired). p

value < 0.05 was considered to be significant.

Results and Discussion

NAFLD is an increasingly important chronic liver disease with a wide spectrum of

histopathology, ranging from bland steatosis to cirrhosis. It is often asymptomatic

and discovered incidentally on routine laboratory screening. In the absence of

established therapies, treatment is generally directed at optimizing body weight

and controlling risk factors.

In the present study it was observed that the prevalence is more in low socio

economical class in age group between 40-45 years which might be due to patients

attending RRIUM, Chennai from lower strata of society. It was also observed that

53.33% patients were of Balghami (phlegmatic temperament) and 36.66% were

of Damavi Mizaj (sanguine temperament) suggesting that those with colder

temperament and robust built are more susceptible to fatty liver thereby

substantiating the claim made by Unani physicians (Ibn-e-Sina -19.6), Ibn-e-

Rushad, 1980). Majority of the patients were overweight (BMI 25), demonstrating

that obesity is a predisposing risk factor for the development of NAFLD (Table-1).

Though majority of the trial participants were asymptomatic which corroborate

various study findings, however dull ache in right hypochondrium was present in

69% of the cases Improvement in most of the sign and symptoms like nausea,

anorexia and reduction in dull ache in right hypochondrium was recorded at the

end of trial (Table 2) indicating the Mushtahi, Muhallil, Musakhkhin, and Muqawwi

kabid properties of polyherbal coded research formulation (UNIM 104).

No any appreciable improvement was observed in liver function tests and lipid

profile as these parameters were within normal range during the course of trial.

(Table 3 & 4) There might be no biochemical evidence of liver disease as stated


Table 1: Demographic data

S.No. Demographic data Unit Mean SD P value

1 Age Mean SD 44.16 7.29

2 Gender Male 16 (53.33%)

Female 14 (46.67%)

3 Duration of disease Mean SD 9.66 10.57

(months)

4 BMI (Kg/m2) Base line 28.05 10.17

After treatment 27.1 1.68 > 0.028

5. Body weight (kg) Base line 73.8 10.17

After treatment 71.26 10.11 > 0.42

6. Mizaj Damavi (sanguine) 11 (36.66%)

Balghami (phlegmatic) 16 (53.33%)

Saudavi (melancholic) 2 (6.66%)

Safravi (choleric) 1 (3.33%)

Table 2: Improvement in Sign & Symptom in NAFLD cases treated with UNIM-104

Status No. of Fatigue Malaise Nausea Loss of Hepato- Dull ache

Cases Appetite megaly in RUQ

Baseline 30 12 9 6 10 7 21

(100%) (40%) (30%) (20%) (33.3%) (23.3%) (70%)

After 30 2 Nil Nil Nil Nil 6

Treatment (100%) (6.6%) (0%) (0%) (0%) (0%) (20%)

Table 3: Biochemical parameters in NAFLD cases treated with UNIM-104

S.No. Parameters Base Line After 90 days P-Value

1 AST (U/Lt) 20.72 2.58 19.93 2.29 >0.21

2 ALT (U/Lt) 24.41 2.88 24.62 3.29 > 0.79

3 Alkaline phosphatise 6.27 1.06 7.08 1.53 > 0.20

(K.A Units)

4 Urea (mg/dl) 19.75 5 21.85 4.73 > 0.10

5 Creatinine (mg/dl) 0.76 0.07 0.78 0.05 > 0.20

by Kumar et al. (2005). Laboratory values may be normal in up to 80% of persons

with fatty liver according to Tierney et al. (2010).


Table 4: Lipid profile in NAFLD cases treated with UNIM-104

S. Parameters Base After After After P

No. Line 30 days 60 days 90 days value

1 Serum 221.82 211.93 205.89 211.41 > 0.34

Cholesterol

(mg/dl) 45.83 40.73 37.80 38.21
2 Serum 136.79 127.58 125.20 131.96 > 0.47

Triglycerides

(mg/dl) 43.46 37.42 49.21 43.72

3 HDL (mg/dl) 42.55 - - 42.17 > 0.47

1.86 2.21

4 LDL (mg/dl) 152.17 - - 142.93 > 0.31

38.31 32.74

Computed tomography is the most reliable imaging modality in the diagnosis of

fatty liver because there is a close correlation between the CT attenuation value

and the degree of fat deposition in the liver parenchyma. Non-contrast CT

attenuation value of the normal liver are on average 8 Hounsfield units (HU) higher

than that of the spleen (Lavine et al., 2000). Normally the attenuation of the hepatic

parenchyma is 50 to 75 HU in a non-contrast CT scan and declines by 1.6 HU for

each milligram of triglyceride accumulated per gram of liver tissue (Subbarao et

al., 2003, Crowley et al., 2007 & Goldman et al., 2008). The study revealed

significant improvement (p< 0.01) in terms of H.U. unit values which enhanced

from 36.69 to 46.95 (Mean) indicating that test drug (UNIM 104) significantly

decreased the accumulated fat from liver due to its Muhallil (Anti-inflammatory),

Musakhkhin (calorifaecient) and Muqawwi kabid (liver tonic) properties. The haar

temperament of UNIM 104 may be responsible in mobilization and redistribution

of Radi Shaham, (morbid fat) which eventually decreases the deposition (Fudhlat

or waste products like alcohol, fats, toxins) of liver (Table 5 & Figure 1).

Table 5: Improvement in C.T Scan H.F Unit values in NAFLD cases treated with

UNIM-104

CT Scan finding Base Line After 90 days P-Value

Liver Hounz field 36.69 10.33 46.95 9.58 <0.01**

(HF) unit


Figure 1: Improvement in C.T Scan H.F Unit values in NAFLD cases treated with

UNIM-104

No adverse effects were observed during and after the trial with the test drug. The

maintenance of laboratory investigations throughout the course of study attests

that the test drug is safe for the patients (Table 3).

Conclusion

It may be concluded that coded Unani formulation UNIM-104 is safe and effective

in the management of Non Alcoholic Fatty liver. It has encouraging potential for

mobilization and redistribution of deposited Raddi Shaham (abnormal excessive

fats) from the liver with no major adverse effects. However, large, long-term,

controlled studies may throw more information about its efficacy.

Acknowledgment

The authors are thankful to the Director General and Dy. Director General, Central

Council for Research in Unani Medicine, New Delhi, for their valuable suggestions,

timely guidance and providing technical facilities in conducting the trial at RRIUM,

Chennai.

References

Angulo, P., 2002. Non-alcoholic fatty liver disease. N. Engl. J. Med. 346 (16):1221

1231.

Angulo, P., Lindor, K.D., 2002. Non-alcoholic fatty liver disease. J. Gastroenterol.

Hepatol, 17 (Suppl):S186S190.

Crowley, L.V., 2007. An introduction to human disease pathology and

pathophysiology correlations. 7th ed. Jones and Bartlett Publishers, U.S.A.,

pp. 569-578.


David, A. Sass, 2005. Non-alcoholic Fatty Liver Disease: A Clinical Review.

Digestive Diseases and Sciences 50(1):171180.

Goldman, L., Ausiello, D., 2008. Cecil medicine. 24th ed. Saunders Elsevier,

Philadelphia, pp. 996-998.

Ibn-e-Rushd Abdul Walee Mohammad (1126-1198), 1980. Kitab-ul-Kulliyat.

Published and Translated by the CCRUM, New Delhi, pp 182-289.
Ibn-e-Sina, 1906. Al-Qanoon Fit-Tib (Urdu Translation by G.H. Kantoori), Vol. 2.

Nawal Kishore Press, Lucknow, p. 20.

Kumar, V., Abbas, A.K., Fausto, N., 2005. Robbins and Cotran Pathologic basis

of disease, 7thed. Saunders Elsevier, New Delhi, pp. 35-36, 904-907.

Lavine, J.E., 2000. Vitamin E treatment of non-alcoholic steatohepatitis in children:

a pilot study. J. Pediatr 136(6):734738.

Ludwig, J., Viggiano, T.R., McGill, D.B., Oh, B.J., 1980. Non-alcoholic

steatohepatitis : Mayo Clinic experiences with a hitherto unnamed disease.

Mayo Clinic Proc. 55(7):434-438.

Sheth, S.G., Gordon, F.D., Chopra, S., 1997. Non-alcoholic steatohepatitis. Ann.

Intern. Med. 126(2):137145.

Subbarao, K., Banerjee, S., Aggarwal, S.K., Bhargava, S.K., 2003. Diagnostic

radiology and Imaging. 2nded. Jaypee brothers Medical publishers Ltd, New

Delhi, pp. 486-495.

Tierney, L.M., McPhee, S.J., Maxine, A., 2010. Current Medical Diagnosis &

Treatment. 49th ed., McGraw Hill, U.S.A., pp. 614-617.

Tominaga, K., Kurata, J.H., Chen, Y.K., 1995. Prevalence of fatty liver in Japanese

children and relationship to obesity: An epidemiological ultrasonographic

survey. Dig. Dis. Sci. 40(9):20022009.



8
Exploration of Abstract

Mechanism of abbe-Gul-e-Aakh (HGA) is an important pharmacopoeal

Anti-
inflammatory
Action of
H

drug of Unani medicine commonly used in a number of inflammatory diseases
such as arthritis and gout etc. It contains four ingredients of plant origin drugs. All
the ingredients have been evaluated individually and shown to possess analgesic,
anti-inflammatory and related effects. HGA as such has also been studied on
experimental models of acute, sub-acute and chronic inflammation and reported
Habbe-Gule-

to be safe and effective anti-inflammatory agent. However, its mechanism of anti-

Aakh inflammatory effect has not been studied so far. Therefore, in the present study

HGA was evaluated experimentally to detect its possible mechanism of action.

1*Musarrat Nafees,

2N.A. Khan, The study was carried out by inoculating the potent chemical mediators like

2K.M.Y. Amin Serotonin (5-HT), Bradykinin, Histamine and PGE1, directly in albino rats by

and subplantar injection to cause edema. The animals were treated with 1000 mg/kg

2Ghufran Ahmad

of 50% hydroalcoholic extract of HGA one hour before injecting the edemagen.

HGA produced 28.0% (p<0.02) inhibition against 5-HT and 41.01% (P=0.001)

1Dept.
of Ilmul Advia,

Z.V.M. Unani Medical College & against PGE1. Although HGA inhibit the Histamine and Bradykinin induced

Hospital, K.B.Hidayatullah Road, inflammation also but the inhibition was found non-significant (16.30% and 3.30%

Azam Campus, Camp,

respectively). It may be concluded therefore that HGA mainly acts by its anti-5HT

Pune-411001

and anti-PGE1 activity.

2Dept.

of Ilmul Advia,

Ajmal Khan Tibbiya College,

Keywords: Unani system of medicine, Habb-e-Gul-e-Aakh, Anti-inflammatory,

Aligarh Muslim University,

Aligarh-202002 Mechanism of action

Introduction

Habbe-Gule-Aakh (HGA) is an anti-arthritic polyherbal formulation of Unani

medicine. It is commonly used in the treatment of inflammatory diseases, mainly

in arthritis and other diseases of joints (Lubhaya, 1979; Khan, ynm).

It is a solid dosage preparation available in a pill form of a gram size containing

four ingredients wiz. Gule-Aakh (Calotropis procera), Zanjabil (Zingiber officinale),

Filfil siyah (Piper nigrum) and Barg-e-Bans (Bambusa arundinacea Retz.) in equal

proportion (Lubhaya, 1979, Khan, ynm). All the four ingredients of HGA are

described to possess anti inflammatory activity, in Unani literature (Attar, 1988;

Ibn Baitar, 1999; Ibn Sina, 1887). Their anti inflammatory activity has also been

proved in various scientific studies (Mascolo et al., 1988; Vendruscolo et al., 2006;

Bang et al., 2009; Muniappan et al., 2003).

Inflammation is a series of pathological changes associated with local vascular

reaction and cellular response of the living tissue to a sub-lethal injury. All



inflammatory reactions are constructed from the same few elements but the

mediators that affect the elements are different. A number of chemicals play major

roles to carry out the process of inflammation (Harsh Mohan, 2000). As chemical

mediators influence inflammatory response, in the very same manner drugs play

roles to influence different mediators in order to show their effects on inflammation.

This influence of drug on chemical mediators is referred as the mechanism of

action of that particular drug. HGA has been described in Unani literature vividly
and is widely prescribed by the physicians in the management of chronic

inflammatory diseases such as arthritis, gout and lumbago etc. Further it has also

been studied scientifically for anti-inflammatory and antiarthritic effect (Nafees et

al., 2012) and shown to possess significant ameliorating property but its

mechanism has still not been elucidated. Therefore present study was undertaken

to explore its mechanism of antiinflammatory action in experimental animals. The

efficacy of HGA was determined against four important inflammatory mediators

viz. Serotonin (5-HT), Bradykinin, Histamine and PGE1. Since these mediators

intervene at different stages of inflammation therefore the exploration of

mechanism will also indicate appropriate therapeutic application of HGA in different

acute, sub acute and chronic forms of inflammation.


Barge-Bans (Bambusa arundinacea Retz.) was supplied by Forest Research

Institute (FRI), Dehradun while the remaining three drugs were purchased from

Dawakhana Tibbiya College, Aligarh Muslim University, Aligarh, which has valid

drug license issued by Govt. of India. Botanists at department of Ilmul Advvia and

department of Botany, AMU, Aligarh identified the drug samples. A voucher

specimen has been deposited in the department of Ilmul Advia, AMU, Aligarh, for

future reference.

Preparation of test drug

All the ingredients were dried under shade and the powder was made from each

drug separately. Then the equiproportional mixture of the four powders was

extracted in 50% alcohol with the help of Soxhlets apparatus for a period of

6 hours. The extract was filtered and evaporated on a hot plate till it dried. The

yield percentage of the extract was calculated with reference to the crude drug

and it was found to be 40.23%. The dried extract however was reconstituted at

the time of administration to animals in to a fresh suspension by adding distilled

water to it. The suspension was administered orally with the help of a gastric

cannula.


Test for mechanism of anti-inflammatory activity

The method described by Parmar et al. (1978) was employed to detect the possible

mechanism of antiinflammatory effect of HGA. Albino rats of either sex, weighing

100-150 gm were divided into 8 groups of 6 animals each. Four groups served as

control and remaining four were used as test groups for each edemagen. Right

hind paw edema was induced by subplantar injection of 0.1 ml. of chemical
mediators viz. 5-HT (1gm/ml) to Group I and II, Histamine (1mg/ml) to Group III

and IV, Bradykinin (20g/ml) to Group V and VI, PGE1 (1g/ml) to Group VII and

VIII one hour after the HGA treatment. The volume of the hind paw was measured

by plethysmometer (UGO Basile7041) at 0 hour and at predetermined intervals

of 30 minutes after administration of 5-HT, Bradykinin and PGE1 and 1 hour after

histamine injection. Each group received 1000 mg/kg of 50% ethanolic extract of

HGA while the animals in Control Group received the vehicle 1 hour before the

edemagen administration. Results were statistically analysed using Students t-

test. A P-value less than 0.05 was considered significant.

Results and Observation

The mean increase in the paw volume produced by 5-HT was found to be 0.308

ml in control group and 0.218 ml. in test group. PGE1 produced edema of 0.206

ml. in control group and 0.11 ml. in test group. Histamine produced a mean increase

of 0.235 ml. in control group and 0.195 ml. in test group while Bradykinin produced

edema of 0.305 ml. and 0.300 ml. in the control and test group, respectively

(Table 1, Fig 1).

Table 1: Effect of HGA on inflammation induced by different endemagen

Groups Paw volume (ml.) Mean Percentage P

mean SE increase value

in Paw of

After treat- volume inhibition

ment (ml.)

5-HT Control 1.520.15 1.830.17 0.308 28.0 <0.02

Treated 1.860.13 2.080.13 0.218

PGE1 Control 1.890.11 2.100.10 0.207 41.01 0.001

Treated 1.620.13 1.740.12 0.110

Histamine Control 1.740.12 1.970.03 0.235 16.30 N.S.

Treated 2.000.04 2.150.02 0.195

Bradykinin Control 1.840.22 2.140.24 0.305 3.30 N.S.

Treated 1.510.06 1.810.12 0.300


Fig. 1: Effect of HGA on inflammation induced by different endemogen

Thus, HGA produced 28.0% (p<0.02) inhibition against 5-HT and 41.01%

(P=0.001) against PGE1. Although HGA inhibit the Histamine and Bradykinin

induced inflammation also but the inhibition (16.30% and 3.30%, respectively)

was non-significant.

Discussion

HGA produced significant effect against 5-HT and PGE 1 induced edema

suggesting that its action is mediated through the inhibition of these two

edemagens. As far as the remaining two chemical are concerned HGA produced

little response that did not amount to be significant statistically. The inflammatory

process is mediated by many chemicals. Vasodilatation is caused by Histamine,

Kinins and particularly by Prostaglandin E and Prostacyclin. Many factors that

are able to increase vascular permeability have also been identified. There are

two or more phases of increased permeability governed by different mediators.

An initial phase begins within the few minutes of injury and lasts for half an hour.

It is usually caused by vasoactive amines like Histamine and Serotonin (Rang et

al, 2003, Ritchie, 1990). The delayed phase which reaches a maximum in few

hours and persists as long as the inflammation lasts is mediated by prostaglandin

E. In the present study HGA was observed to produce significant antiinflammatory

effect against PGE1 and 5-HT induced inflammation. The maximum effect however

was found against PGE1 induced inflammation while the effect against 5-HT

induced inflammation was although significant statistically but was comparatively

of lesser degree. Therefore, it can be inferred that HGA is less effective in early

phase of inflammation. This is so, because 5-HT, Bradykinin and Histamine are


the chemicals which mediate the inflammatory reactions in early phase and the

test drug found to be less effective against these mediators. On the other hand

HGA was able to reduce the inflammation mediated through PGE1 enormously

therefore it may be deduced that the test drug is more effective in delayed phase

of inflammation and may be therapeutically useful in chronic inflammatory

conditions. This finding is in conformity with our previous report where it has been

shown that HGA has very striking antiarthritic effect (Nafees et al., 2012). A drug
specially that having antiinflammatory activity such as HGA if used over a long

period of time, is liable to induce side effects particularly the gastritis and gastric

ulcer etc. However, one of the ingredients of HGA viz. Zanjabil (Zingiber officinale)

possesses comparatively weak anti-inflammatory effect but has been reported to

protect the gastric mucosa significantly (Nanjundaiah et al., 2011). Inclusion of

Zanjabil in the compound is probably meant to minimize the side effect of HGA

which is likely to be used for long period of time in chronic inflammatory condition.

It indicates that HGA is effective and safe PGE1 inhibitor.

Conclusion

It can be concluded therefore that HGA produces anti-inflammatory activity mainly

through inhibiting PGE1. Since this mediator is responsible for delayed phase of

increased permeability and inflammatory response therefore it is likely that the

test drug will be more useful in the management of chronic inflammatory conditions.

References

Attar, H.Z., 1988. Ikhtiyarat-e-Badiyee (Kanpur). Matba Munshi Naval Kishor,

Lucknow, p. 393.

Bang, J.S., Oh, D.H., Choi, H.M., Sur, B., Lim, S., Kim, J.Y., et.al., 2009. Anti-

inflammatory and antiarthritic effects of piperine in human interleukin

1-stimulated fibroblast-like synoviocytes and in rat arthritis models. Arthritis

Res Ther. 11(2): 49.

Harsh, Mohan, 2000. Text book of pathology, 4th ed. Jaypee Brothers Medical

Publishers (P) Ltd., New Delhi, pp. 114,121-122.

Ibn-e-Baitar, 1999. Al-Jamiul Mufradat-al-Advia Wal Aghzia, Vol III & IV. Central

Council for Research in Unani Medicine, New Delhi, pp. 274, 276, 377, 378,

380.

Ibn-e-Sina, 1887. Tarjuma-e-Qanoon. (Translated by Moulvi H. S. Ghulam

Husnain), Vol II. Matba Munshi Naval Kishor, Kanpur, pp. 93,184,186,187,197.

Khan, A.G., ynm. Maseehul Mulk ke Murakkabat. Vol.II. Urdu Dawakhana, pp.

77-78.


Labhaya, R., 1979. Delhi ke Muntakhab Murakkabat, Goswami Kutubkhana, Delhi,

p. 89.

Mascolo, N., Sharma, R., Jain, S.C. and Capasso, F., 1988. Ethnopharmacologic

investigation of Calotropis procera flowers. J. Ethnopharmacol. 22(2): 211-221.

Muniappan, M. and Sundraraj, T., 2003. Anti-inflammatory and anti-ulcer activities

of Bambusa arundinacea. J. Ethnopharmacol. 88(2-3): 161-167.
Nafees, M., Khan, N.A., Amin, K.M.Y., Ahmad, G., 2012. Evaluation of Anti-

inflammatory activity of Habbe-Gule-Aakh in albino Rats. Hippocratic Journal

of Unani Medicine 7(4): 1-9.

Parmar, N.S., Ghosh, ,M.N., 1978. Anti-inflammatory activity of gossypin isolated

from HVLinn. Indian Journal of Pharmacology 10:277-93.

Rang, H.P., Dale, M.M., Ritter, J.M., Moore, P.K., 2003. Pharmacology, 5th ed.,

Churchill Livingstone, New York, p. 219.

Ritchie, A.C., 1990. Boyds Text Book of Pathology, Vol.I, 9thed., Lea and Fabiger,

Chesterfield, pp. 61,62,73,74.

Siddaraju, M., Nanjundaiah, Harish Nayaka, Mysore, Annaiah, and Shylaja M.,

Dharmesh, 2011. Gastroprotective Effect of Ginger Rhizome (Zingiber

officinale) Extract: Role of Gallic Acid and Cinnamic Acid in H+, K+-ATPase/

H. pylori Inhibition and Anti-Oxidative Mechanism. Evidence-Based

Complementary and Alternative Medicine. Article ID 249487.

Vendruscolo, A., Takaki, I., Bersani-Amado, L.E., Dantas, J.A., Bersani-

Amado, C.A., Cuman, R.K.N., 2006. Anti-inflammatory and antinociceptive

activities of Zingiber officinale Rosc. essential oil in experimental animal

models. Indian J. Pharmacol. 38(1): 58-59.


Toxicity Study Abstract

of Qurs-e-Hudar he present study was carried out to investigate the safety of Qurs-

in Experimental
Animals
*Mohd Nadeem,
T

e-Hudar by conducting acute and sub-acute toxicity in Swiss albino mice & rats
respectively. Acute toxicity was determined by administering aqueous extract of
Qurs-e-Hudar orally to two groups of mice of six each at a dose 1gm/kg and 2gm/
kg body weight. The animals were observed for gross behaviour and mortality for
24 hours after drug administration. The formulation was well tolerated by the
Mohd Urooj,

animals and no abnormality was observed in the general behaviour of the animals

Habibur Rahman

and no overnight mortality was recorded. Similarly, Sub-acute toxicity was

and

Shariq Ali Khan determined in albino rats by orally administration of aqueous extract to three groups

of seven animals each at a dose ranges from 1 gm/kg and 3 gm/kg body weight

Pharmacology Research Unit

for 28 days. The results of haematology and biochemistry profile done on 29th


day were found to be normal and no changes were observed in organ to body wt

Unani Medicine,

Post Box 70, Aligarh-202002 ratio of liver, heart, kidney and spleen.

Keywords: Acute toxicity, Sub acute toxicity, Qurs-e-Hudar, Zingiber officinalis,

Strychnos nux-vomica, Detoxification

Introduction

Some drugs of the plant, animal and mineral origin used in Unani system of

medicine have components that are naturally toxic in their properties. Such drugs,

are therefore, detoxified in order to reduce/ overcome their toxicity. In Oriental

medicine Strychnos nux-vomica L. (Loganiaceae) seeds have been used in

activating the channels, alleviating pain and reducing swelling (Bensky and

Gamble, 1989). The pharmacological effects of this plant have also been known

to increase spinal reflexes and stimulate respiratory and sensory centres of the

cerebral cortex (Chung and Shin, 1989). Azaraqi is a deadly poison and is detoxified

before its use for the preparation of drug. Its main constituent is alkaloid strychnine

with human LD50 as 0.7-2.1 mg/kg (Jackson and Marsh, 1997). In the present

study the drug has been evaluated for its toxicity in Swiss albino mice and rats of

either sex.

Determination of efficacy and safety of herbal remedies is necessary because

many people using these agents as self medication. Since there is limited data

available about the safety of the commonly used herbal remedies, therefore, effort

to elucidate health benefits and risk of herbal medicine should be intensified

(Anonymous, 2011). Toxicity testing in animal is typically the initial steps to

determine the effect profile of test substance and potential hazards which occurs

due to short term exposure of test substance. A toxicity study provides information



on the hazardous properties and allows the substance to be ranked and classified

according to the Globally Harmonized System (GHS) for the classification of

chemicals which cause acute toxicity (OECD 2000).

Formulation

Qurs-e-Hudar is a poly herbal formulation containing below mentioned constituents
in equal parts

S.No. Common Name Botanical Name

1 Azaraqi Strychnos nux-vomica

2 Zanjabeel Khushk Zingiber officinalis

3 Filfil Siyah Piper nigrum

Methodology

This study was carried out in Pharmacology Research Unit, Regional Research

Institute of Unani Medicine, Aligarh, and conducted in accordance with the protocol

approved by Institutional Ethics Committee of RRIUM Aligarh.

Procurement of Drug:

The Qurs-e-Hudar formulation was procured in the form of Tablet from Central

Research Institute of Unani Medicine, Hyderabad.

Detoxification of Azaraqi (Nux Vomica) Seeds:

Azaraqi was detoxified by pharmacy section of Central Research Institute of Unani

Medicine, Hyderabad, as per the method given in National Formulary of Unani

Medicine (Anonymous, 1981b). 70 grams of Azaraqi seeds were buried in Peeli

Matti (yellow clay) for 10 days. The clay was kept moist throughout. The seeds

were then removed and washed. The seed coat was peeled off with the help of a

sharp knife and the cotyledons were separated after removing the embryo. The

healthy cotyledons were then washed with hot water. The cotyledons in a cloth

bag were boiled in 2 lit of milk till it evaporated (the care was taken that the cloth

bag should not touch the bottom of the container).The water washed cotyledons

were then used subsequently.

Animals

The study was carried out in Swiss albino mice (20-25 g) and rats (100-150g) of

either sex, for acute & sub-acute toxicity determination respectively. The animals


were procured from M/S Rahat Hussain Enterprises Biological Suppliers, Babri

Mandi, Aligarh. They were acclimatized to the conditions for one week before

experimental study. The animals were maintained in a standard environmental

condition at a room temperature of (252 degree Celsius) with 12 Hrs light/Dark

cycles, humidity (50-55%), and had free access to food pellets. The study was

conducted after approval of protocol from Institutional Ethics Committee of RRIUM,

Aligarh.
Preparation of Drug Extract

The tablets of the drugs were crushed into fine powder and a weighed quantity

was steeped in acidulated distilled water. The water soaked mass of the drug was

warmed over water bath and kept for 24 hours at room temperature. During this

period it was occasionally stirred. After 24 hours it was filtered through a filter

paper and filtrate was dried over water bath. The aqueous extracts of the drug

thus obtained was used in different doses selected according to OECD guidelines

for safety evaluations.

Acute Toxicity Study

The current study was carried out in accordance to Organization of Economic

Co-operation and Development (OECD) guideline for testing of chemicals. Swiss

Albino mice of either sex weighing 20-25 g were randomly selected and divide

into two groups of six mice each. Mice were kept fasted overnight (12hrs) with

free access to water prior to administration of dose ranging 1gm/kg body weight

and 2 gm/kg body weight as per limit test of OECD guideline. The aqueous extract

of the drugs was administered orally. The animals were kept in polypropylene

cages after drug administration and were observed for behaviour pattern

(salivation, lacrymation, lethargy, sleep, clonic convulsions, vigorous rolling and

head tilt) & mortality at 1 hour, 2 hour, 3 hour, 4 hour, 5 hour, 6 hour, 12 hour, 24

hour and thereafter once every day up to 14 days after drug administration.

Sub-Acute Toxicity Study

Swiss Albino rats of either sex weighing between 100-150 g were randomly

selected and divided into three groups of seven animals each. Rats were kept

fasted overnight (12hrs) with free access to water prior to administration of dose

ranging 1gm/kg and 3gm/kg body weight for 28 days as per limit test of OECD

guideline. Group I was kept as normal control which received distilled water for

28 days, while in the IInd and IIIrd groups aqueous extract of the drug was

administered orally at a dose of 1gm/kg and 3gm/kg body weight for 28 days. The

animals were kept in polypropylene cages after drug administration and were


observed for Gross behavior (salivation, lacrymation, lethargy, sleep, clonic

convulsions, vigorous rolling and head tilt) & mortality at 1 hour, 2 hour, 3 hour, 4

hour, 5 hour, 6 hour, 12 hour, 24 hour and thereafter once every day up to 28 days

after drug administration. On 29th day blood was collected of all the three groups

of rats through retro-orbital plexus for estimation of SGOT,SGPT and Serum

alkaline Phosphatase, Serum urea and Serum Creatinine, Serum cholesterol,

Serum triglyceride and Serum HDL, Percentage hemoglobin, ESR, Total leukocyte
count and Differential leukocyte count. After collection of blood the animals in all

the three groups were sacrificed and liver, heart, kidney and spleen were excised

out for determination of organ to body weight ratio. SGOT, SGPT were estimated

by Reitmans and Frankel, 1957 method (Reitman et al., 1957). Alkaline

Phosphatase was estimated by Bessey and Brock, 1946 method (Bessay et al.,

1946). Serum urea was estimated by GLDH, Ureas method given by (Tiffany et

al., 1972), while Serum Creatinine was estimated by Jaffes method given by

(Bowers et al., 1980). Serum HDL was estimated by Phosphotungstic Acid method

given by (Burstein et al.,. 1970), while Serum cholesterol and Triglyceride were

estimated by CHOD-PAP method given by (Roeschlau et al., 1974) and GPO-

Trindev method given by (Mcgowan et al., 1983). ESR and DLC were estimated

by Westergreen and Leishman stain methods given in Medical Laboratory

Technology (Mukherjee, 1990). TLC was estimated by Hemoaltometry method

given by (Plum, 1936). Percent hemoglobin was estimated by Sahlis Acid

Haematin method given by (Newcomer, 1919).

Statistical Analysis

Statistical analysis was performed by using unpaired t test calculating p value at

5 % level. All values are expressed as MeanSEM (standard error of mean). p

value less than 0.05 found to be considered statistically significant


Acute Toxicity Study

The effect of oral administration of single dose of aqueous extract of Qurs-e-

Hudar in Swiss albino mice shows that the formulation was well tolerated by the

animals and no abnormality was observed in the general behaviour of the animals

and no overnight mortality was recorded. Herbs and supplements can be toxic

when used for inappropriate indication, or prepared inappropriately, or used in

large excessive dosages or for a prolonged duration of time. Since it is polyherbal

formulation other ingredients present in formulation helps in reducing the toxic

effect of active component.


Sub-Acute Toxicity Study

The values of all the parameters including liver functions, renal functions

hematology and organ body weight ratio found to be normal as compared to

control group. The effects of the studied drug on organ body weight ratio in control

and treated animals are presented in Figure 5. There were no significant changes

observed in organ body weight ratio of the control and the animals treated with
various doses. Figure 1, 2, and 3 is graphical representation for the effects of

Fig. 1: Effect of Qurs-e-Hudar on Liver Function Test

Fig. 2: Effect of Qurs-e-Hudar on Renal Function Test


drug on the biochemical parameters. There were no significant changes in AST

and ALT levels in all the treated animals compared with the control. Similarly lipid

profile of treated animal was found to be normal as compared to control group as

shown in Figure 3. The values for hematological parameters of treated group as

compared to control showed no significant changes in Hb, TLC, ESR, and %

lymphocyte as shown in figure 4. It can be concluded on the basis of above

observation that drug is quite safe.
Fig. 3: Effect of Qurs-e-Hudar on Lipid Profile

Fig. 4: Effect of Qurs-e-Hudar on Hematological Parameters


Fig. 5: Effect of Qurs-e-Hudar on Organ to the Body Weight Ratio

Conclusion

The formulation Qurs-e-Hudar was well tolerated by the animals and no

abnormality was observed in the general behavior of the animals and no overnight

mortality was recorded. There were no findings of any organ toxicity and

hematological changes as laboratory findings were normal. It can be concluded

on the basis of above observations that drug studied is quite safe in animals.

Acknowledgement

The authors would like to record their gratitude to Director General, Central Council

for Research in Unani Medicine, New Delhi, for providing an academic and

research environment at the institute to work with excellence and meticulousness.

The authors would also like to thank Dr. R.S Verma, Research Officer

(Biochemistry) for his technical support and co-operation in carrying out the

biochemical investigations in biochemistry laboratory of RRIUM, Aligarh.

References

Anonymous, 1981b. (reprinted in 2006). National Formulary of Unani Medicine,

Ist edition. Part 1, GOI, Ministry of H&FW, New Delhi. p. 319.

Anonymous, 2011. Toxicity testing overview. Toxicity Testing Resource Centre

(TTRC), pp 2-25.

Bensky, D. and Gamble, A., 1986. Chinese Herbal Medicine, Eastland press,

Sealtle, p. 646.


Bessay, O.A., Lowry, O.H., Brock, M.J., 1946. A method for the rapid determination

of alkaline phosphates with five cubic millimeters of serum. Journal of Biological

Chemistry 164:321-329.

Bowers, L.D., Wong, E.T., 1980. Kinetic serum creatinine assay I. The role of

various factors in determining specificity. Clin.Chem 26 (5):551-554.

Burstain, M., Scholnic, H.R., Morphin, R., 1970. Rapid method for the isolation of
lipoprotein from human serum by precipitation with polyanions J. Lipid. Res

11:583-595.

Chung, B. and Shin, M.K., 1989. Dictionary of Folk Medicine. Young Lin Press,

Seoul. p. 972.

Jackson, T.A. and Marsh, F.P., 1997. Test methods for vertebrate pest control and

management material.American Society for testing and materials,

Philadelphia, pp. 1-4.

Macgowan, M.W., Artiss, J.D., Strangberg, D.R., Zak, B., 1983. A peroxidase-

coupled method for the colorimetric determination of serum triglycerides. Clin

Chem. 29 (3):538-542.

Mukherjee, K.L., 1990. Medical Laboratory Technology, Vol. 1, 3rd Edition. Tata,

McGraw-Hill Publishing Company Limited, New Delhi, pp. 228-30.

Newcomer, H.S., 1919. Absorption Spectra of Acid Hematin, Oxyhemoglobin and

carbon monoxide hemoglobin. A new hemoglobinometer J .Biol.chem. 37:465-

496.

OECD, 2000. Guidance Document on the Recognition, Assessment and Use of

Clinical Signs as Humane Endpoints for Experimental Animals Used in Safety

Evaluation. Environmental Health and Safety Monograph Series on Testing

and Assesment No 19.

Plum, P., 1936. Accuracy of hematological counting method. Acta. Med. Scandinav

90: 342-364.

Reitman, S., and Frankel, S., 1957. A colorimetric method for determination of

serum glutamic oxaloacetic and glutamic pyruvic

transaminases.Am.J.Clin.Pathol.28:56-63

Roeschlau, P., Bernt, E., Gruber, W., 1974. Enzymatic determination of total

cholesterol in serum. Z Klin. Chem, Klin Biochem. 12 (5):226.

Tiffany, T.O., Jansen, J., Burtis, C.A., Overton, J.B., Scott, C.D., 1972. Enzymatic

kinetic rate and end- point analyses of substrate by use of a GeMSAEC fast

analyzer. Clin. Chem. 18: 829-840.


Comparative Abstract

Analytical retrospective study was conducted to elaborate the

Study of the
Efficacy of
Unani
A

successful treatment for oligospermia. For comparison, four studies with six
common parameters were taken into account involving one hundred twenty six
diagnosed patients with idiopathic oligospermia. The study was divided into four
groups namely A, B, C and D and it was observed that among the six common
parameters, the semen volume was improved in C and B group, semen liquefaction
Formulations

time was raised in A, B and C groups, sperm count was maximally raised in D

with Reference group, sperm motility was improved in A group, and sperm morphology was almost

equal in all groups and pH of semen has shown no change in all groups.

to Oligospermia

Keywords: Oligospermia, Unani treatment, Herbal Medicine, Qillat-e-Huwaniya.

*M. Anas

and
Introduction

M.M.H. Siddiqui

As per the WHO guidelines, a sperm count of <20 millions/ml is considered as

Department of Ilaj-bit-Tadbeer,
oligospermia (Dhaliwal et al., 2011). It is one of the main factors of infertility in

A.K. Tibbiya College,

60% of couples. Among them male infertility is involved in 40% while 20% of the


Aligarh-202002 cases there is a combination of both male and female factors. The factors

responsible for male infertility are low sperm count, abnormal sperm morphology

and impaired sperm transport. As a remedy, a number of nutritional supplements

are used to improve sperm count and sperm motility including Amino acids like

carnitine, arginine, minerals like zinc, selenium, numerous vitamins and

antioxidants like vitamin B12, vitamin C, vitamin E and glutathione and co-enzymes

Q10 in treating male infertility (Mukherjee et al., 2003). Male infertility also involves

harmful environmental and occupational risk factors along with underlying

nutritional imbalances therefore; to overcome these factors including correction

of nutritional imbalances to produce optimum sperm count and their functions, a

multifaceted therapeutic approach can be a management tool to improve male

infertility (Prdani, 1976; Solepure et al., 1979; Sengupta, 1982).

The Unani medical literature regarding male sexual disorders is enriched with its

contents and therapeutics but the terminologies of various disorders are ill defined

that may be due to primitive knowledge of Physiology of the ancient and traditional

Unani scholars. But the contents of therapeutics of ancient Unani physicians clearly

indicate that they were well aware about oligospermia. Unani Medical literature

has several authentic manuscripts that are rich and full of knowledge of sexual

disorders. To explore the things on scientific parameters to validate the ancient

knowledge before the present medical fraternity and public domain has

encouraged us to analyzed oligozoospermia which is a major challenge before

sexual internist and general physician as well.



Purpose of the study

Extensively and critically reviewing and analyzing already carried out four studies

which have similar methodology using different drugs so as to put forth the most

promising drug/drug combination that can be reassessed for its efficacy by using

more comprehensive and standard protocols.

Material and Methods

This retrospective study was carried out to elaborate the promising drug

combination studied in the past for the treatment of Qillat-e-huwaniya-e-maniyat

(Oligospermia) on one hundred Twenty Six patients pre-diagnosed with idiopathic

oligozoospermia. The study was divided into four groups namely A, B, C and D. In

Group A the test drug Safoof-e-Saalab (a combination of Moosli Sainbhal

(Bombax malabaricum DC), Saalab Panja (Orchis latifolia Linn) and Satawar

(Asparagus racemosus Wild), in Group B the test drug was a powdered

combination of Saalab Misri (Orchis latifolia Linn), Khoolanjan (Alpinia galanga

Wild) and Koonch (Mucuna pruriens Bak), in Group C the test drug was Laboob-

e-Kabir Khaas (a Unani polyherbal pharmacopieal preparation) and in Group D

the test drug was a powdered combination of Saalab Gatta (Orchis laxiflora Linn),

Salaajeet (Styrax officinale Linn), Moosli Safaid (Asparagus adscendens Roxb),

Moosli Sainbhal (Bombax malabaricum Linn) and Sataawar (Asparagus

racemosus Wild). All the drugs have six gram dose in each case orally twice a

day for a period of sixty days.


In the study the average semen volume before treatment was observed 3.5ml,

2.1 ml, 2.9ml, and 3.4ml in groups A, B, C and D respectively. After treatment it

was improved to 4.7ml, 3.8ml, 4.6ml and 4.4ml in the same sequence. The average

improvement in semen volume was observed as 1.2ml, 1.7ml, 1.7ml and 1.0 ml

in groups A, B, C, and D respectively. The maximum improvement was noted in

group B and C cases, this improvement of the semen volume was may be due to

drugs such as Orchis latifolia and Asparagus racemosus which possess tonic

and spermatagogue activities (Alok et al., 2013)

In the study, the sperm count improvement was observed as 2.8 million sperm

count in group A, 6.5 million sperm count in group B, 6.4 million sperm count in

group C and 8.2 million sperm count in Group D. This effect of the formulations

may be due to the drugs like Asparagus adscendens, Bombax malabaricum, which

enhances the production of sperm (Sinha and Jagdale, 2013) and the drug like

Orchis latifolia which correct the sluggishness of sperms and helps in better


Table 1: Effect of test samples on volume of semen

Before treatment After treatment

Group Volume of No.of %age Average No.of %age Average Improve-

semen patients volume patients volume ment

in ml in ml in ml in ml

A 2.0-3.9 14 66.7 3.5 06 28.5 4.7 1.2
4.0-7.0 07 33.3 15 71.5

B 2.0-3.9 25 83.3 2.1 25 83.3 3.8 1.7

4.0-7.0 05 16.7 05 16.7

C 2.0-3.9 17 68 2.9 01 04.0 4.6 1.7

4.0-7.0 08 03 24 96.0

D 2.0-3.9 39 78 3.4 16 32.0 4.4 1.0

4.0-7.0 11 22 34 68.0

(Siddiqui, 1991; Qasmi, 1994; Khan, 1996; Ahmad, 2011)

penetration. The formulation ingredients like Styrax officinale is suppose to be a

Muqawwi-e-Aza-e-Raisa wa Umoomi (General tonic) and Muqawwi-e-Baah

(Aphrodiasic) leads to increase the capacity of testes to produce more sperms.

Table 2: Effect of test samples on mean sperm count in millions

Group Mean sperm count Mean sperm count Mean improvement

before treatment after treatment in sperm count

A 9.9 12.7 2.8

B 14.3 20.8 6.5

C 13.3 19.7 6.4

D 14.6 22.8 8.2


Table 3: Effect of test samples on sperm motility in percent

Group Mean sperm motility Mean sperm motility Mean Improvement

before treatment after treatment in sperm motility

A 35.0 50.0 15.0

B 42.6 53.9 11.3

C 41.4 54.5 13.1

D 37.0 47.0 10.0



The increase in sperm count may also be attribute to the drugs like Asparagus

adscendens and Orchis latifolia which contain Protein Saponins, micronutrients

like Iron, Selenium and Zinc (Ainslie, 1984; Kirtikar and Basu, 1996)

Similar to above, it was observed that group A has the maximum mean

improvement in sperm motility followed by group C, group B and lastly group D.
Table 4: Effect of test samples on sperm morphology

Group Mean normal sperm Mean normal sperm Mean

before treatment after treatment Improvement

A 37.0 47.0 10.0

B 39.4 50.0 10.6

C 42.0 52.8 10.8

D 37.0 44.8 07.8


Table 5: Effect of test samples on Liquefaction time


Group Lique- No.of % Avg. No.of % Avg. Improve-

faction patients in pts in ment

time min min in min

(in min)

A 06-10 14 66.7 11 00 00 18 07

11-15 04 19.0 13 62.0

16-20 03 14.3 08 38.0

B 06-10 19 63.3 10 01 3.3 15 05

11-15 08 26.6 16 53.3

16-20 03 10.1 13 43.3

C 06-10 15 60 9.4 01 04 14.9 5.2

11-15 09 36 13 52

16-20 01 04 11 44

D 25 05 10 25 12 24 17 -08

25-30 15 30 10 20

30-35 20 40 16 32

35-40 10 20 12 24



The motility of the sperms increased may due to the combined and stimulant

effect of the herbs. This activity is attributed to the drugs like Orchis latifolia, and

Styrax officinale that improve the strength by virtue of general tonic as well as

aphrodisiac properties (Ainslie,1984; Khan, 1885; Ghani, ynm; Majoosi, 1993)

In the study the effect of test samples on sperm morphology showed that trialed

combinations have restored normal morphology of the sperms to a significant

level. All the groups showed equal mean improvement but with a little difference

i.e 10.8 and 10.6 in the Group A and B followed by 10.0 in Group C and 7.8 in

Group D. The improvement in sperm morphology may possible due to actions of

the drugs like Asparagus adscendens, Bombax malabaricum, and Asparagus

racemosus (Kirtikar and Basu, 1996; Ghani, ynm; Majoosi,1993) The possible

explanation to improve the morphology of the sperm may due to the strengthening

of the QuwwateTanasuliya (reproductive power) of the body which in turn

strengthens its QuwwateMusawwira (Formative power) that is responsible for

the formation and maintenance of the shape of sperm.

In the parameter liquefaction time of semen, it was observed that in group A, B

and C liquefaction time was increased by 07, 05, and 5.2 minutes respectively

and this effect was attributed to the action of drugs like Asparagus adscendens,

Bombax malabaricum, and Asparagus racemosus which possess Mughalliz and

Moallid Mani effect claimed by various writers (Khan, 1885; Ibne sina, 1906;

Majoosi, 1933;and Rabban, 1996)

Table 6: Effect of test samples on pH of semen


Group pH of semen No. of pts Percentage No. of pts Percentage

A 7.0-7.9 05 23.8 05 23.8

8 16 67.2 16 67.2

B 7.0-7.9 10 33.3 10 33.3

8 20 66.6 20 66.6

C 7.0-7.9 06 24.0 06 24.0

8 19 76.0 19 76.0

D 7.0-7.9 20 40.0 20 40.0

8 30 60.0 30 60.0


Finally, it was observed that there was no change in the pH of semen before and

after the treatment in all the groups. This means that all the formulations

experimented do not interfere with pH of the semen.


Conclusion

Over all, it was concluded that all the four combinations studied possess

aphrodisiac/ rejuvenative, spermetogenesis, spermetagogue, antidepressive an

anxiolytic activity. They increase the level of testosterone, libido. These action

made the formulations therapeutically active to treat male oligospermia.

Although all the four groups have nearly similar therapeutic uses and are trialed
for the same conditions but as per observations, semen volume was increased

more in group C and B cases, semen liquefaction time is increased in group A, B,

and C cases, sperm count was increased more in group D cases as compared to

other groups, sperm motility is increased in group A as compared to rest of the

groups, as far as sperm morphology is concerned it increases in all groups.

Therefore, to manage oligospermia the formulation of all four groups may be

specifically used as per the individual case requirement in respect to the

parameters taken into account.

References

Ahmad, B., 2011, Qillat Haiwaniyat-e-Manviyah (Oligospermia) ka Tahqeeqi

mutala aur uskey Ilaj me Salab Gattah, Salajeet, Moosli Safaid, Moosli

Sainbhal, Satawar ki Ifadiat ka Jaizah, M.D (Moalejat) Thesis, Faculty of

Unani Medicine, A. M. U. Aligarh.

Ainslie, W., 1984. Materia Medica, Neeraj Publication House, Delhi, pp. 24-25

and 368-369

Alok, S., Jain, S.K., Verma, A., Kumar, M., Mahor, A., and Sabharwal, M., 2013.

Plant profile, phytochemistry and pharmacology of Asparagus racemosus

(Shatavari): A review. Asian Pacific Journal of Tropical Disease 3(3): 242

251.

Dhaliwal, L.K., Gupta, K.R. and Majumdar, S., 2011. Treatment of oligospermia

with Speman: A formulation of plant origin. Indian Medical Gazette, pp. 375-

379.

Ghani, N., ynm. Khazeenul Advia, , Idara Kitab ul Shifa, Delhi, pp. 542,543,788-

789,820,821,1271,1272

Ibne Sina, Abdullah Ibne Hassan, 1906. Al Qanoon Fit Tib, Vol 1st and 2nd . Munshi

Naval Kishore Press, Lucknow, pp. 67-68

Jagdale, S.P., Shimpi, S., Chachad, D., 2009. Pharmacological Studies Of Salep.

Journal of Herbal Medicine and Toxicology 3 (1) 153-156

Khan, A., 1885. Moheet-e- Azam, Vol 2 and 4. Matba Nizami, Kanpur, pp. 11,47

and 113-114


Khan, B., 1996. Qillat haiwaniyat (Oligospermia) per Laboob Kabeer Khas ki Ifadiat

ka Jaizah, M.D (Moalejat) Thesis, Faculty of Unani Medicine, A. M. U. Aligarh.

Kirtikar, K.R. and Basu, 1996. Indian Medicinal Plants, Vol 4, 2nd edition.

International book distributors, Rajpur, Dehradun, pp. 2412, 2413, 2499-2501

Majoosi, A., 1933. Kamil-us-Sana, Umada-al-Matab , Lucknow, pp. 39-40

Mukherjee, K., Tripathi, A. and Kulkarni, K.S., 2003. Evaluation of the efficacy of
Speman in the management of male subfertility. Indian J. Clinical Practice

13(11): 29-31.

Pardanani, D.S., Delima, R.J., Rao, R.V., Vaze, A.Y., Sheth, A.R. and Jayatilak,

P.G., 1976. Effect of an indigenous drug (Speman) on human accessory

reproductive function. Indian J. Surgery 38(1): 34-39.

Qasmi, M.A., 1994. Qillat wa Zoaf Haiwaniyat Manviyah (Oligospermia) wa Zoaf

Baah (Sexual Weakness) me Salab Misri, Khoolanjan aur Tukhm Koonch ki

Ifadiat ka Jaizah. M.D (Moalejat) Thesis, Faculty of Unani Medicine, A.M.U.,

Aligarh.

Rabban, Tabri, 1996. Firdaus Al Hikma Fit Tib. Hamdard Foundation Press,

Karachi, pp. 706-715

Sengupta, S. 1982, A Clinico-pathological study on the effect of Speman on

spermatogenesis in cases of oligospermia. Probe XXI (4): 275-276

Siddiqui, M.S.H., 1991. Qillat wa Zoaf-e-Haiwaniyat Manviyah (Oligospermia) ka

Tahqeeqi mutala aur uske ilaj me Safoof Salab ki Ifadiat ka Jaizah. M.D

(Moalejat) Thesis, Faculty of Unani Medicine, A.M.U., Aligarh.

Sinha, B.N., Pandey A., Diwan A., Saini, 2013. Bombax malabaricum DC: A

Salutary Boon. International Journal Pharmaceutical Innovations 3(2):17-19.

Solepure, A.B., Nirmala M. Joshi, Deshkar B.V. and Muzumdar, S.R., 1979. Effect

of Speman on quality of semen in relation to magnesium concentration. The

Indian Practitioner, pp. 663-665.



30
Redesigning of Abstract

the Unani ablet is one of the most suitable and preferred solid dosage

Preparation
from Powder
Form into
T

form used all over the world. Almost all drug molecules can be formulated as
a tablet and the process of manufacturing of tablets is very simple, and flexible.
One can administer 0.01 mg to 1 gm of a drug as a tablet by the oral route.
Therefore, in the present study an anti-arthritic pharmacopoeial preparation in
powder form having ingredients Suranjan Talkh(Colchicum luteum), Zanjabeel
Tablet and its

Khushk (Zingiber officinale) and Elwa (Aloe vera) were re-designed and modified

Standardization for use in the form of Tablet (Qurs) and standardized. Organoleptic characters of

tablet, tests for weight variation, uniformity of diameter and thickness, hardness,

Along with

disintegration time and friability of tablets were conducted for standardization and

HPLC Profile

the values obtained indicated the compliance with the pharmaceutical standards.

HPLC profile of tablet and qualitative analysis of chemical constituents present in

*Aziz ur Rahman the tablet were also determined. Furthermore, tablets were also tested for the

and

presence of pesticidal residue by comparing HPLC profile of pesticides and tablet

Tajuddin

in identical conditions and the result shown absence of pesticides in the formulation.

Department of Saidla, These tablets can be used as an alternative of powder form of the given formulation

A.K. Tibbiya College, and the findings will help to set the standards for future reference.


Aligarh-202002

Keywords: Re-design, Colchicum luteum, Zingiber officinale, Aloe vera, Tablet,

and HPLC

Introduction

The Unani System of Medicine and its pharmaceutics are most suitable to the

people of the Indian subcontinent as it was not only the part of culture and heritage

but has a long cherished history during the ages of different dynasties and reigns.

But in this scientific era fast paced life style emerged with exploration of various

scientific inventions in the biomedical fields and pharmaceutical sciences in

accordance with the need and requirement of the modern age. The pharmaceutical

industries of traditional and herbal medicine came up with the invention of

convenient dosage form development in accordance with the demand of

biomedical requirements as well as to provide better compliance to the patient.

Tablet is one of the most convenient and appropriate solid dosage form provided

an accurately measured dose in a convenient portable package; and can be

designed to protect unstable medications or disguise unpalatable ingredients.

Tablets are most stable of all oral dosage forms and easy to swallow. They are

very economical also (http://tabletsdosageform).



Therefore, in view of above and in order to achieve global acceptance, a

modification has been made to pharmacopoeial Unani formulation mentioned in

Ilaj al Amaraz (Khan, 1870) i.e. from powder form in to the tablet as most of the

dosage forms falls in this category, by adding Gum Acacia (S. d. Fine Chemical

Ltd.) as an excipient along with its standardization which can also provide the

better compliance to the patient. Further, one of the ingredients of the formulation

that is Aloe is extremely bitter in taste and the consumption of the drug per oral in
powder form would be more distasteful to the patient which can be easily used in

the form of tablet. Parameters for tablet standerdisation include organoleptic

characters, physical parameters, qualitative analysis and HPLC profile of tablet.

In the form of tablet the formulation seems convenient to take and easy to handle

by the practitioners and patients. The dosage of the formulation becomes more

specific and accurate in the form of the tablet.

Ingredients of formulation

(1) Sonth (Zingiber officinale Linn. Dried Rhizome) 3.5 g

(2) Suranjan talkh (Colchicum luteum Baker Dried Corm) 3.5 g

(3) Elwa (Aloe vera Linn. Dried Exudate) 7.0 g


Crude drug collection and authentication

The raw materials were procured from the local market of Aligarh and the samples

were authenticated in pharmacognosy section of the department of Ilmul Advia,

Faculty of Unani Medicine by Prof. S. H. Afaq and found within range of standards

as mentioned in Ayurvedic and Unani Pharmacopoeia of India (Anonymous, 1999,

2001 and 2007).

Preparation of tablet

The two ingredients of formulation Sonth and Suranjan talkh were powdered in

an electric grinder and Elwa was made Mushawwa (broiled or roasted) by keeping

in an Apple, covered it by the process of Kapoorti, and heated in an oven, till it

was brown. Now, Elwa was taken out of apple and dissolved in distilled water as

required, after few minutes it was filtered and then concentrated. All the three

ingredients were mixed together in order to make Lubdi (dough). The mixture

was dried in shed and then powdered in a mortar. This powder was mixed with a

suitable inert substance viz. powder of Gum Acacia (S.d. Fine Chemical Ltd.) as

excipient. The material in the requisite degree of fineness mixed and damped

with a moistening agent (purified water). The moistened material was made into

granules by passing through a sieve. The granules were dried in shed and again


passed through a sieve for uniformity of size. Tablets of 500 mg were made by

Automatic tablet making machine in Dawakhana Tibbiya College, AMU, Aligarh

(Anonymous, 1968; 1970).

Standardization of the tablet dosage form

The tablets were evaluated for various quality control parameters like organoleptic

characters (appearance, color, smell, taste and texture), weight variation, diameter,

thickness, hardness, friability and disintegration time. Qualitative analysis and

HPLC profile of tablet was also done. Tablets were also tested for the presence of

pesticides by comparing the HPLC profile of tablet with the HPLC of standard

samples of pesticides.

Organoleptic characters

Twenty tablets were taken to check for appearance and any discoloration or

degradation, if any, by visual method (Patel et al., 2010). Tablets were also tested

for their smell, taste and texture. All the data obtained were based on random

selection and multiple observations.

Weight variation of tablet

Ten tablets were selected randomly and weighed individually by an Electronic

weighing machine to check for weight variation and was expressed in mg. Weight

5% deviation is acceptable for the tablets / pills having average weight 250mg

or more, hence considered as appropriate (Subrahmanyam et al., 2006; Dandagi

et al., 2006).

Diameter of tablet

Uniformity of Diameter was assessed by picking three tablets randomly and the

diameter was measured individually by using a Digital Vernier Caliper and

expressed in mm. (Subrahmanyam et al., 2006; Dandagi et al., 2006).

Thickness of tablet

Uniformity of Thickness was assessed by picking three tablets randomly and the

Thickness was measured individually by using a Digital Vernier Caliper and

expressed in mm. (Subrahmanyam et al., 2006; Dandagi et al., 2006).

Hardness of tablet

Hardness or Tablet crushing strength (the force required to break a tablet in a

diametric compression) was measured using Monsanto Tablet Hardness Tester

and expressed in kg/cm2 (Subrahmanyam et al., 2006; Vijaya and Mishra, 2006).


Determination of disintegrating time

The rate of disintegration was measured by a Disintegration-Testing Apparatus

using the two media, the aqueous and the acidic and expressed in minutes. For

measurement in aqueous medium double distilled water was taken and for

measuring in acidic medium simulated gastric fluid (pH about 1.2) was prepared

without enzyme by dissolving 1gm of NaCl in 500 ml of distilled water, adding 7
ml of concentrated HCl, and diluting the volume to 1000 ml with water

(Subrahmanyam et al., 2006; Anonymous, 1989) and modified by Afaq et al. (1994).

Friability test

Friability is the loss of weight of tablet in the container / package, due to removal

of fine particles from the surface. Friability of tablets was determined by using

Roche Friabilator and expressed in percentage (%). This device subjects the

tablets to the combined effect of abrasions and shock in a plastic chamber revolving

at 25 rpm and dropping the tablet at a height of 6 inches in each revolution. Pre

weighed sample of tablets were placed in the friabilator and were subjected to

100 revolutions. Tablets were de-dusted using a soft muslin cloth and re-weighed.

The friability (f) is calculated by the following formula.

W0

f = 1 x100

Where, W0 is the weight of the tablets after the test and W is the weight of the

tablets before the test (Subrahmanyam et al., 2006; Vijaya and Mishra, 2006).

Qualitative analysis of chemical constituents

The qualitative analysis of different chemical constituents, present in the tablet

was carried out according to the scheme proposed by Bhattacharjee and Das

(1969). This scheme has been given in the form of Flow Chart (Fig. 1).

The powder of compound formulation was extracted with petroleum ether. The

petroleum ether extract (I) was tested for free phenols, alkaloids and sterols/

terpenes. A part of this extract was saponified and sap portion (II) was tested for

fatty acids, whereas, unsaponified portion (III) was tested again for alkaloids,

phenols, and sterols/terpenes for confirmation. The defatted marc was divided

into two portions. One portion was extracted with hot water and the other with

ethanol (70%). The aqueous (IV) and ethanolic (V) extracts were tested for

alkaloids, flavonoids, saponins, sugars and tannins. Aqueous extract was extracted

with ether and ether soluble portion (VI) was tested again for alkaloids, sterols/

terpenes, whereas, water-soluble portion (VII) was tested for glycosides. The water-

soluble portion was again hydrolyzed with 5% hydrochloric acid and extracted


Fig. 1: Scheme for qualitative analysis of chemical constituents in the tablet

with chloroform. The aglycone portion (VIII) was tested for insoluble hydrochloride

of alkaloid. Chloroform soluble portion (IX) was tested for alkaloids and sterols/

terpenes, whereas; water-soluble fraction (X) was tested for alkaloids. One part

of this water-soluble portion was basified with any alkali (ammonia) and extracted

with immiscible solvent (ether). The solvent soluble part (XI) was again tested for

alkaloids.


Qualitative chemical analysis of the tablet for above mentioned compounds were

carried out by different tests and methods for the determination of the presence

of active compounds (Overtone, 1963; Harborne, 1973 and Afaq et al., 1994).

HPLC profile of the tablet

General HPLC profile of the hydroalcoholic extract of the tablet was done by a
Cyber labs HPLC system equipped with a single pump and porous silica with 5

m diameter, C 18 4.6250 mm column and software driven peaks were obtained.

The pressure, temperature and flow rate was 7 Pa, 250C and 1.0 ml/min,

respectively. Detector for HPLC was UV and the wave length was 254 nm. Mobile

phase for HPLC profile of extract consisted of acetonitrile: water (55:45). The

hydroalcoholic extract of coarsely powdered drug was determined with the help

of Soxhlets apparatus for 6 hours, extract was filtered and allowed to evaporate

on water bath. 20mg of hydroalcoholic extract was dissolved in 1ml of HPLC

grade distilled water and used for study.

Determination of Pesticidal residue

Common pesticides (Chloropyriphos, DDT, Parathion, Malathion and

Endosulphan) were obtained from Sigma-Aldrich, mixed properly and dissolved

in Acitonitrile. This standard was injected in the C18 column of HPLC instrument

(Cyber lab, USA) considering the retention time in the identical conditions as in

the HPLC profile of the test drug and software driven peaks were obtained. These

peaks were compared with the peaks of tablet.


Organoleptic characters are the first step to identify the drugs and also indicate

the status and condition of the drug. If any discoloration or black spots appears, it

shows the degradation or decomposition of the in the tablet (Patel et al, 2010).

Organoleptic properties like colour, taste, smell, appearance and texture were

noted and depicted in Table 1.

Table 1: Organoleptic description of formulation tablet

Appearance Tablet

Colour Dark Brown

Smell Agreeable

Taste Pungent

Texture Hard


A good quality tablet should be accurate and uniform in weight, diameter and

thickness. But diameter and thickness of a tablet can vary without any change in

its weight. Tablets diameter and thickness are controlled by the machine tooling,

includes a die and punch. This is called a station of tooling. The thickness is

determined by the amount of tablet material and the position of the punches in

relation to each other during compression (http://en.wikipedia.org/wiki/Tablet). It

should be controlled within a 5% variation of standard value (http://

www.pharmainfo.net/namanm/evaluation-tablets). The uniformity of weight,

diameter and thickness was found to be within the prescribed limit and mean

values are shown in Table 2.

Tablet requires a certain amount of strength or hardness and resistance to friability

to withstand mechanical shakes during manufacture, packaging, shipping and

handling by the pharmacist and patient. The resistance of the tablet to chipping,

abrasion or breakage under above mentioned condition before usage depends

on its hardness and friability. Hardness and Friability loss indicate that tablets

have good mechanical resistance and have ability to with stand abrasions. A loss

less than 1% is considered acceptable by industrial standard and all the tablets

were found well within approved range to give good handling properties without

breakage or excessive friability problems.

Tablet should be hard enough to withstand manufacturing, packaging, and

transportation process. However they cannot be too hard since that may alter

disintegration or release of the drug product. Disintegration roughly indicates the

possible pattern of dissolution of active substance. Hence, the experimental

conditions closely mimic the situations that a tablet encounters in gastrointestinal

tract, in terms of temperature, pH and mechanics. Physical parameters such as

area, hardness, surface characteristics and size can significantly affect the rate

of disintegration of drugs contained in a complex system. After administration,

Table 2 : Parameters for standardization of tablet

S.No. Parameters Result (MeanSE)

1. Weight variation of Formulation Tablet (g) 0.525 0.01

2. Uniformity of Diameter (mm) 13.54 0.00

3. Uniformity of Thickness (mm) 4.77 0.01

4. Hardness (kg/cm2) 2.67 0.17

5. Friability (%) 0.47 0.01

6. Disintegration time in the water (minutes) 12.17 0.39

7. Disintegration time in the simulated gastric 10.14 0.52

fluid (minutes)


the tablet should disintegrate readily for quick absorption in gastro-intestinal tract;

therefore, the tablets were also subjected for the evaluation of Disintegration time.

According to the test the tablet must disintegrate and all particles must pass through

the 10 mesh screen in the time specified. If any residue remains, it must have a

soft mass. It is mentioned that uncoated tablets must disintegrate in not more

than 45 minutes (Anonymous, 1989). Results for hardness, friability loss and

disintegration time illustrated in Table 2.
The therapeutic properties of the crude drugs are mainly due to presence of

physiologically active chemical constituents present in the drugs. The presence

and absence of active compounds are shown in Table 3.

HPLC Analysis

The HPLC profile of the formulation was recorded as the obtained graph can be

compared with the batches in future. The HPLC pattern shows 15 peaks and the

peak no. 1 is major peak having concentration 84.632% and retention time was

found to be 1.98 min. The details are depicted in Fig. 2 and Table 4. HPLC is an

important qualitative and quantitative analytical procedure. By doing HPLC we

can also determine the impurities in the formulation. If there is any change in no.

of peaks or retention time or area of peaks from standard it indicates adulteration

or deterioration in the drug. The HPLC profile of the mixture of different pesticides

Table 3: Qualitative analysis of chemical constituents

S.No. Tested for Result

1. Alkaloids + ve

2. Amino acids - ve

3. Fixed oil / Volatile Oil + ve

4. Flavonoids + ve

5. Glycosides + ve

6. Phenols + ve

7. Proteins - ve

8. Resins - ve

9. Saponins + ve

10. Sterols / Terpenes + ve

11. Sugar (Reducing) - ve

12. Sugars (Non-reducing) - ve

13. Tannins + ve


Fig 2: HPLC Profile of the Tablet

Table 4: HPLC Profile of the Tablet

ID Name Retain.T Height Area Conc Tail. Theo.

Factor Plate

1 1.908 22622 185305.8 84,632 1.43 1165

2 2.509 678 3404.8 2.536 1.00 4974

3 2.600 782 4709.3 2.926 2.00 3716

4 2.758 334 2141.5 1.250 1.89 3689

5 2.950 679 4883.2 2.540 1.00 3354

6 3.092 332 5293.5 1.242 4.54 749

7 3.512 167 2262.3 0.625 1.50 1339

8 3,853 145 1907.9 0.542 1.62 1709

9 4.236 245 3758.5 0.917 0.90 1520

10 4.473 138 1264.5 0.516 1.38 4750

11 4,773 177 3669.7 0.662 1.28 1056

12 5,407 234 5956.6 0.875 1.64 890

13 6,302 14 127.0 0.052 1.35 9616

14 7,472 171 1821.3 0.640 1.41 9808

15 8.025 12 93.6 0.045 1.07 21100

Sum: 26730 226599.8 100.0000


in Acetonitrile depicted in Fig. 3 and table 5, shows there is no any peak

corresponds to the HPLC profile of tablet indicating absence of pesticides in the

test drug. A comparison of both of them is also shown in Fig. 4.
Fig 3: HPLC Profile of the mixed pesticides

Table 5: HPLC Profile of the Tablet

ID Name Retain.T Height Area Conc Tail. Theo.

Factor Plate

1 1.092 23 82.9 1.197 1.19 1830

2 1,768 261 6405.7 13.580 1.24 103

3 2.268 54 378.2 2.810 2.25 2091

4 2.912 210 2042.2 10.926 1.12 1787

5 3,203 1009 11936.6 52.497 2.02 1461

6 4.030 21 294.6 1.093 1.22 1645

7 5.665 199 2523.1 10.354 1.27 3979

8 6.058 145 1701.5 7.544 1.73 5313

Sum: 1922 25364.7 100.0000


Fig 4: Comparison of HPLC profile of tablet and mixture of pesticides (Peaks

with sky blue and red colour indicates HPLC profile of tablet and pesticides

respectively)

Conclusion

This study assumes great significance as it will provide a key of diagnostic

characters specially HPLC profile which serves as an important tool in laying

down the standards for quality assurance. The parameters applied for

standardization of formulation viz. organoleptic characters, friability and HPLC

etc. may be taken as standard parameters for future reference.

The study also offers an improvement in Unani health care system by redesigning

of powder (Safoof) form which is less convenient to use into the more convenient

tablet form.

Acknowledgment

The authors are grateful to the, Dept. of Ilmul Advia, F/o Unani Medicine, A.K.

Tibbiya College, Aligarh Muslim University, Aligarh, for providing significant

assistance to carry out this work.

References

Afaq, S.H., Tajuddin., Siddiqui, M.M.H., 1994. Standardization of Herbal Drugs.

Publication Division, AMU, Aligarh, pp. 33-34, 41-42, 100, 143-146.


Anonymous, 1968. British Pharmacopoea, General Medicine Council.

Pharmaceutical Press, Bloomsbury square, London, pp. 27-28(A.V.), 1276-

1277, 1286-1288, 982-985 (Tablet).

Anonymous, 1970. Pharmacopoeia of India, 2nd ed. Ministry of Health & F.W.,

Manager of Publications, Delhi 238-239(A.A.), pp. 496-497.

Anonymous, 1989. Food and Drug Regulations. Ministry of Health, USA, Section

C. 01.015.

Anonymous, 1999. The Ayurvedic Pharmacopoeia of India (Edn. 1st) Ministry of

Health and Family Welfare, Govt. of India, New Delhi, Part-I, Vol.2; 12-14(Z.O.)

Anonymous, 2001. The Ayurvedic Pharmacopoeia of India, (Edn.1st), Ministry of

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pp. 103-104(Z.O.).

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83(A.V.), 88-89(Z.O.).

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8(Z.O.).

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Indian Drugs 43(2): 117-121.


Development of Abstract

Standard he present study was designed to convert a classical Unani

Operative
Procedure
(SOP) of a
T

pharmacopoeial preparation Sunun Poste Mughilan (a tooth powder) into
toothpaste form with the same ingredients as contained in powder form. This
work was undertaken with the objective of developing the Standard Operating
Procedure (SOP) for manufacturing process of toothpaste. The legitimacy of SOP
was determined by assessing the quality of product in three different batches.
Toothpaste

Each batch was assessed three times for spreadability, foam formation and

Prepared with dispersion time in water. The group which was in range with standard limits was

selected as standard batch. The procedures adopted for the preparation of

the Ingredients

standard batch was taken as the SOP.

of Sunun Poste

Mughilan Keywords: SOP, Sunun, Toothpaste, Spreadability, Tooth powder.

*Mohammad Rashid,
Introduction

Shariq Shamsi

and In Unani system of medicine, toothpowders are commonly known as Sunun which

Roohi Zaman
contain finely powdered drugs. Toothpowders are prepared both with herbal and

mineral drugs but more often by combining the two. These powders usually contain

Department of Ilmul Saidla,

National Institute of Unani Medicine, more than one ingredient. Apart from the maintenance of oro-dental hygiene

Kottigepalaya, Magadi Main Road,

toothpowders are also described in Unani medicine to be used in various

Bangalore-560091

pathological conditions of tooth, gum, throat and buccal cavity. Although

toothpowders are in use since ancient periods but due to rapid changes in modern

cult, use of toothpowder is declining day by day even in rural population. Further

toothpastes also have definite advantages over toothpowders which also pave

the way for its wider acceptability. Toothpowders bear less spreadibility than

toothpaste and usually contain coarse particles, hence chances of abrasions,

friction to enamel, dentin and soft tissue ablations are much high, leading to various

periodontal diseases like dental caries, hypersensitivity of teeth, pyorrhoea,

toothache, bleeding gums, gingivitis and recession of gums etc. Secondly, it is

very difficult to protect powders usually containing hygroscopic, deliquescent

(tending to melt or dissolve in humid environment) and aromatic materials from

decomposition. Thirdly, the toothpaste has largest share of dental cleansing and

care preparations. Further, chances of least wastage at use, attractive consistency,

and easy distribution in mouth, and over all convenient and soft use further add to

their acceptability.

In recent years acceptability and demand for natural products has increased many

fold mainly because of the safety concern which is almost unavoidable with most



of the products prepared with synthetic raw materials. In Unani system of medicine

a number of formulations intended to be used to maintain the oral health and

hygiene and to ameliorate various diseases of tooth, gum, mucus membrane and

throat etc are available but most of them are available in powder form such as

Sunun, Zaroor, Qula and Mazmaza etc. The age-old Pharmacopoeial formulation

Sunun Poste Mughilan (SPM) is used to strengthen the dental roots and impart

the sparkle to the teeth (Said, 1997; Kabiruddin, ynm) and used commonly by the
patients and healthy person alike. However, the change of its form is desired in

view of the advantage of paste over powder and also the convenience of use of

paste form. Therefore an attempt has been made to convert SPM into toothpaste.

The Standardization of its procedure and product has also been undertaken to

set the standard of a genuine sample and establish the SOP. Three variables i.e.

spreadibility, foaming power and dispersion time were taken as the attributes for

its standardization and quality determination. During the conversion of SPM into

the toothpaste, different batches (F-1, F-2, and F-3) were prepared and assessed

on three variables as mentioned above.


Procurement of Raw drugs

All the ingredients of SPM were procured from the raw drug dealer, except the

bark of Acacia arabica which was collected from the campus of National Institute

of Unani Medicine (NIUM), Bangalore. Dr. R. Sumathi, Botanist at RMR, FRLHT-

IAM, Bangalore authenticated the plant drugs. The voucher specimens (Areca

catechu-2741, Acacia catechu-2742, Zingiber officinalis-2743, Piper nigrum-2744,

Acacia arabica-2745) have been deposited in the museum of the Institute of

Ayurveda and Integrative Medicine, Bangalore. Other constituents needed to

prepare the toothpaste were procured from the Srinivasa Products, 5th Block,

Rajaji Nagar, Bangalore.

Preparation of extracts

The crude drugs (Table 1) were dried in oven at 350C and coarse powder was

prepared after grinding each drug separately. Then each powder was extracted

separately by continuous hot extraction (Soxhlet extractor) with double distilled

water at 1000C. The extracts were dried on water bath.

All the ingredients of SPM were used in the ratio of 4:1:1:1:0.1:0.1 as mentioned

in Hamdard Pharmacopoeia (Table 2). Toothpaste was prepared by taking extract

of all the plant drugs while the mineral drug (Silicate of magnesia) was taken as

such and mixed with the extracts. Ten percent of total amount of extract of all the


Table 1 : Ingredients of Sunun Poste Mughilan

Unani Name Scientific Name Part Used

1 Post Kikar Acacia arabica Bark 400gms

2 Burnt Supari Areca catechu Nut 100gms

3 Sange Jarahat Silicate of magnesia Stone 100gms
4 Kath Safaid Acacia catechu Extract 100gms

5 Zanjabeel Zingiber officinalis Dried Rhizome 10gms

6 Filfil Siyah Piper nigrum Seed 10gms

Table 2 : Composition of extracts of Toothpaste

Sl.No. Ingredients Gm

1 Acacia Arabica 5.56

2 Areca catechu 1.39

3 Silicate of magnesia 1.39

4 Acacia catechu 1.39

5 Zingiber officinalis 0.14

6 Piper nigrum 0.14

Total 10

plant drugs and Sange Jarahat was taken for the formulation of toothpaste. Other

ingredients required to give the form of paste were also used (Table 3).

Preparation of Toothpaste

Heated Liquid-Phase Process

Ten gm of extract of all plant drugs and Sange Jarahat was weighed first in the

ratio of 4:1:1:1:0.1:0.1. All the ingredients were mixed with Sorbitol, Water and

Saccharin sodium dehydrate in container No. 1. The solution was then heated

with continuous stirring for 15 minutes at 40 0C.

In another container (No. 2), Calcium Carbonate, Carboxy Methyl Cellulose and

Preservatives (Methyl Paraben and Propyl Paraben) were taken and mixed

properly. A hot solution of humectants, water, extracts and sweetener which was

prepared after proper heating and stirring was then, slowly added with mixing to

the powder which was prepared in No. 2 container. The resulting mass was mixed


Table 3 : Composition of Toothpaste

Sl. Ingredients Formulations Property

No. Qty. Used (%) (% w/w)

F1 F2 F3

1. Extracts 10 10 10 Active Ingredient

2. Calcium carbonate 35 45 25 Abrasive

3. Sodium lauryl sulphate 1.5 2.5 1.5 Surfactant

4. Sorbitol 30 20 20 Humectant

5. Sodium Carboxy 1 1 1 Binding agent

Methyl Cellulose (CMC)

6. Sodium saccharine 0.3 0.3 0.3 Sweetener

7. Methyl paraben 0.1 0.1 0.1 Preservative

8. Propyl paraben 0.02 0.02 0.02 Preservative

9. Titanium dioxide 0.5 0.5 0.5 Opacifier

10. Peppermint oil 1 1 1 Flavouring agent

11. DD Water 20.58 19.75 40.58

well to get thick paste. Finally the Sodium Lauryl Sulphate, peppermint oil, Titanium

dioxide were added and mixed properly.

Determination of spreadibility, foaming and dispersion time

Three different batches of toothpaste were prepared with different percentage of

ingredients. Each batch was assessed three times for spreadability, foam

formation, and dispersion time in water.

Determination of Spreadibility

One gm of paste was placed at the centre of the glass plate (10X10 cm) and

another glass plate was placed over it carefully. Above the glass plates 2 kilogram

weight was placed at the centre of the plate. Sliding of the plate was avoided. The

area (cm) of spread of paste was measured. The experiment was repeated three

times and the average was recorded (Akelesh et al., 2010; Pael and Kamani,

2009).

Determination of Foaming Power

Five gram of paste was placed in a 100 ml glass beaker. To this 10 ml of water

was added and the beaker was covered with a watch glass and allowed to stand


for 30 minutes, this operation was carried out to disperse the toothpaste in water.

The contents of the beaker were stirred with a glass rod and the slurry was

transferred to a 250 ml graduated measuring cylinder. During this transfer it was

ensured that no foam was produced and no lump paste went into the measuring

cylinder. The residue left in the beaker was transferred with further portion of 5-6

ml of water to the cylinder. The content of cylinder was adjusted to 50 ml by

adding sufficient water and the content was maintained at 30C. The contents of
the cylinder were stirred with a glass rod to ensure a uniform suspension. Stirring

was stopped when the temperature of the content reached to 30 0C and then 12

complete shakes were given by hand. The cylinder was then allowed to stand for

5 minutes and the volume of foam with water and the volume of water only was

noted.

Foaming power was determined by the following formula (Akelesh et al., 2010):

Foaming power = V1 V2

V1 - Volume in ml of foam with water

V2 - Volume in ml of water only

Determination of Dispersion Time in Water

The USP tablet disintegration test unit was used. 150 ml of distilled water at

370C+1 was placed in the cylinders of the apparatus. One gram of the toothpaste

was placed, cautiously, onto the gauze of the basket. The movement of the basket

was maintained at the normal speed of the apparatus and the temperature was

kept at 370C+1. The time elapsed till no paste was present on the gauze was

determined and was taken as a measure of dispensability of the paste in water. A

time ranging from 10-30 minutes is considered satisfactory (Pander, 1993).

Results

The quality of the toothpaste was determined by assessing each batch three

times for spreadability, foam formation and dispersion time in water and the mean

was regarded as standard parameter value with an aim that if the paste will qualify

the criteria of quality then it will be assumed that the procedure adopted to prepare

the paste is also standard. The results of all batches are given in Table 4 and Fig.

1. The batch F3 with abrasive and humectant shows spreadability, foam formation

and dispersion time in water as 7.47 0.09 cm, 122.33 1.20 ml and 26 0.58

min, respectively. Most appropriate result among three batches was found in this

group. The values of all the above parameters were compared with the standard

values given by Bureau of Indian Standards and with other published work (Table

5) (Fig 2). The findings on comparison were found to be within the normal range.


Table 4 : Results of Formulations Batches

Sl. Formulations Spreadability Foam formation Dispersion time

No. (cm) (ml) (min)

1 F1 6.00 0.12 97 1.2 45 0.63

2 F2 4.2 0.12 11 0.58 55.89 0.35

3 F3 7.47 0.09 122.33 1.20 26 0.58

Fig. 1

Table 5: Comparison of F3 with the optimum values mentioned in Bureau of

Indian Standard

Values Spreadability Foam Formation Dispersion time

(cm) (ml) in water

Toothpaste value 7.47 122 26

Standard value (Max.) 8.5 Min 50 10-30 min


Fig. 2

Discussion

An important Unani powdery dentifrice SPM was successfully converted into paste

form using the standard procedures and approved excipents while retaining the

ingredients of the formulation. The quality standards of the newly converted

formulation were tested on some important variables and it was found that the

formulation qualify the criteria necessary to establish its standard and thereby the

genuineness. The study demonstrated that the paste attained almost same

standard as that mentioned in Bureau of Indian Standard suggesting that the

procedure adopted to prepare the paste was also genuine and standard and may

be taken as the standard operating procedure.

One of the criteria for paste to meet quality standard is that it should possess

good spreadability. The spreadability is used to denote the extent of area to which

a paste readily spreads on application to the skin or affected part, as the therapeutic

efficacy of a formulation also depends upon its spreading ability (Patel, 2012).

Spreadability is imperative characteristic of toothpaste and shows the consistency

of paste. The toothpastes are homogenous in nature and it should not separate

into liquid and solid ingredients. The large spread area as seen in case of the

paste of SPM demonstrated that it possesses good consistency (Das et al., 2013).

Thickness or the consistency felt in the mouth upon using the toothpaste is

important because it is directly related to the usage of the dentifrice. If toothpaste


is too thick, it may be difficult to spread around the teeth and it may not be used

properly. If toothpaste is too thin, it may not be too effective in cleaning. Additionally,

an adequate amount of thickness in paste is required to keep the abrasives properly

dispersed throughout the toothpaste (Mason Stephen, 2000). This depends heavily

on the amounts of gelling agents in the toothpaste, as well as on the fraction of

the toothpaste which is solid. Hence, determination of spreadability is very

important in evaluating paste characteristics. The mean value of spreadability of

toothpaste was found to be 7.47 0.09 cm which is almost equal to that of the

standard value of 8.5.

A controlled level of the foam in toothpaste is necessary for suspending and

removing the debris, rinsing the mouth and giving an intense perception of cleaning

action. This is due to the surface tension reducing properties of detergents (Nanda

et al., 2009). The quality of the foam, i.e. volume and stability or lifetime, depends

upon many factors such as, the amounts and properties of humectants, polymers,

and abrasives used (Pader Morton, 1997). The foam volume depends mainly on

the amount of surfactant. The ratio of amount of surfactant and foam volume can

be best fit line for experimental data. The foaming character was studied for nature,

stability and washability of toothpaste (Mithal and Saha, 2010). The mean value

of foaming power of toothpaste was found to be 122.33 1.20 ml which is

significantly higher than the minimum required value of 50.

Dispersion time is used to measure dispersion power of toothpaste in water. A

time ranging from 10-30 minutes is considered satisfactory. The mean value of

dispersion time in water was found to be 26 0.58 min which was found to be

within the normal range of 10-30. Thus the all three parameters the paste form of

the preparation was tested for were found to be commensurate with the standard

values.

Conclusion

The study demonstrated that the conversion of Sunun Poste Mughilon (SPM)

into paste form was successful as the finished product was found to qualify all the

criteria laid down for toothpastes of standard quality. It was concluded therefore

that the procedure used to prepare the toothpaste is also standard and it may be

used as the SOP for preparation of other herbal and herbo-mineral toothpastes.

Acknowledgement

The authors are grateful to Prof. Mansoor Ahmad Siddiqui, Director, National

Institute of Unani Medicine, Bangalore, for providing the financial and logistic

support.


References

Akelesh,T., Kumar, R.S., Jothi, R.P.V.R., Raj, A., Venkatnarayan, R., 2010.

Evaluation of Standards of Some Selected Cosmetic Preparations. JPRHC

2(4): 302-306.

Das, Ishita, Suki Roy, Shreta Chandni, Karthik, L., Gaurav Kumar and Bhaskar

Rao, 2013. Biosurfactant from marine actinobacteria and its application in

cosmetic formulation of toothpaste. Der Pharmacia Lettre 5 (5):1-6.

Kabiruddin, M., Al-Qarabadin Faisalabad, ynm. Kashmir Book Deport, Faisalabad,

p. 568.

Mason, Stephan, 2000. Dental Hygiene, Pouchers Perfumes, Cosmetics and

Soaps, pp. 217-253.

Mithal, B.M., Saha, R.N., 2010. A Handbook of Cosmetics. Vallabh Prakashan,

Delhi.

Nanda, S., Nanda, A., Khar, R.K., 2009. Cosmetic Technology. Birla Publication

Pvt. Ltd.

Pader, Morton, 1997. Surfactants in Dental Products. Surfactants in Cosmetics.

(Eds. Martin M. Rieger and Linda D. Rhein). CRC Press.

Pader, M., 1993. Dentrifice Rheology. In: Rheological properties of cosmetics

and toiletries. Marcel Dekker Inc., New York.

Patel, R., Kamani, R., 2009. Formulation Optimization and Evaluation of

Mometazone Furate Cream. Journal of Pharmacy Research 2(10:1565-1569.

Patel, H., Panchal, M.S., Shah, S., Vadalia, K.R., 2012. Formulation and Evaluation

of Transdermal Gel of Sildenafil Citrate. IJPRAS 1(3):103-118.

Rashid, M., 2013. Formulation and Evaluation of Unani Toothpaste Composed of

Ingredients of Sunun Post Mughilan: A Unani Tooth Powder. Journal of

Research in Unani Medicine 2(2): 1-9.

Said, H.M., 1997. Hamdard Pharmacopeia of Eastern Medicine. Sri Satguru

Publication, A Division of Indian Books Centre, Delhi, pp. 166-167.



52
A Review on Abstract

Phytochemistry, edicinal plants are serving mankind to cure ailments

Pharmacological
Properties and
Biotechnological

Msince time immemorial and today they are equally important even with a higher
potential, because of the understanding of the mechanism of action with the advent
of modern technique and extensive research. The present review looks into the
medicinal potentials of Valeriana officinalis. To encompass the major activities of
this plant, information is collected pertaining to phytochemistry, pharmacological
Studies in

properties and biotechnological investigation carried out in various laboratories

Valeriana world-wide. Valeriana officinalis, known as Sumbu-ul-teeb in Unani system of

medicine, is an important source of various bioactive compound viz. valerenic

officinalis L.,

acid, valepotriates, alkaloids, flavonoids, lignans etc. Pharmaceutical application

An Important

of valerian is due to its sedative, anticonvulsant, antidepressant, antihypertensive,

hypnotic, antispasmodic, antidysmenorrheal and anxiolytic activity. The literature

Medicinal Herb

reviewed on biotechnological approaches include in vitro protocols for large scale

production of valerian, genetic transformation and elicitation studies undertaken

*Anwar Shahzad

and for enhanced production of valerenic acid.

Taiba Saeed

Keywords: Valeriana officinalis L., Tagar, Valerenic Acid, Valepotriates, Sedative

Plant Biotechnology Laboratory,

Department of Botany,


Introduction

Aligarh-202002

In recent times an upsurge is witnessed worldwide towards the use of medicinal

plants as a whole or in the form of crude drug in traditional systems of medicine

such as Ayurveda, Sidha and Unani, which draw the attention of modern science

researchers to understand the isolation, application of bioactive compounds and

mechanism of action to cure diseases in a more precise way. Now a huge amount

of research outcomes are available to be utilised in a judicious way. The present

review deals with the application of Valeriana officinalis an important medicinal

plant of traditional system of medicine to cure various diseases as well as the use

of biotechnological tools to conserve its fast depleting population because of

indiscriminate collection, and the production of bioactive compounds through hairy

root culture, elicitation, and genetic transformation for the overexpression of gene

related to desired metabolites.

Botanical Name : Valeriana officinalis L.

Common name : All-heal, Garden heliotrope, Great wild Valerian, Amantilla,

Phu, Nard, Theriacaria, Marinella, Genicularis and Terdina.

Unani name : Tagar



The genus Valeriana, with about 200 species, belongs to the Valerianaceae family

and grows throughout the world (Piccinelli et al., 2004). Valeriana officinalis L.

one of the medicinally important species of Valeriana, is a perennial flowering

herb commonly known by various names such as all-heal, garden heliotrope,

great wild Valerian etc (Cunha et al., 2003). Two other ancient names are nard

and phu. Nard is derived from a Sanskrit word meaning strong smell and

phu or fu refers to the usual exclamation of disgust that attends the experience
of smelling the dried root. The word valeriana might have been derived from the

Roman province of Valeria, or from Valerianus, a Roman emperor, or from a

certain Valerius who first used the herb as medicine, while other writers believe it

came from the Latin word valere (to be in health). The plant is native to Europe

and Asia and has naturalized in eastern North America. It appears in moist places

with mild climates, mainly in forests and river margins. It is cultivated in low lying,

damp sandy humus with lime fertilizer (Cunha, 2005). Other important species of

Valeriana includes V. fauriei used in Traditional Chinese Medicine and Japanese

Medicine, V. capensis used in African traditional medicine, V. edulis used in Mexico

and V. wallichii is used in India (Hikino et al., 1975; Iwu, 1993; Huang, 1999). The

collective species V. officinalis L. is very polymorphic, with a number of naturally

occurring subspecies that differ from each other by their degree of ploidy (Bos et

al., 1997). This plant grows from a short rhizome to approximately 1.5 m in height,

it flowers, and then dies back again in the winter. The rhizome is light greyish

brown, about the size of a finger joint, bearing many rootlets. V. officinalis L. has

pinnately-divided leaves, typically with 6 -10 pairs of lanceshaped leaflets, and

bears many small white or pink hermaphroditic flowers in a dense head of several

stalked clusters. These heads bear small (5 mm) tapered seeds (Andrews and

Basu, 2005). The fresh root has no odor, while the dried root smells distinctly

unpleasant, akin to old gym socks, due to isovaleric acid (Bisset, 1994; Flemming,

1998).

V. officinalis (Tagar) is one of the important constituents of many Unani formulations

(Anonymous, 1969) which have been reported to possess some interesting

pharmacological effects (Ahmad et al., 2012).

Phytochemistry

This plant contains over 150-200 chemical constituents, including the essential

oils and their sesquiterpenoid derivatives (valerenic acids), epoxy iridoid esters

(valepotriates), alkaloids, flavonoids and lignans which are mainly produced in

the root and rhizome (Cramer et al., 2006). Among these constituents, valerenic

acids and valepotriates are often regarded as active principles in valerian

commercial and medicinal preparations and its crude extract is widely used in

many countries (Houghton, 1999).


Essential Oil

The essential oil contains about 150 constituents which includes monoterpenes,

sesquiterpenes, less volatile sesquiterpenic carboxylic acids. Borneol (Carretero,

2000; Cunha, 2005) mainly in the form of ester viz. bornyl acetate and bornyl

isovalerate , camphene, and -pinene are the major monoterpenes found in

essential oils of V. officinalis (Carretero, 2000; Barnes et al., 2002; Heinrich et al.,
2004). Sesquiterpenes include valeranone (Dewick, 2002; Heinrich et al., 2004),

valerenal (Carretero, 2000; Dewick, 2002; Heinrich et al., 2004), valerenol (Barnes

et al., 2002; Heinrich et al., 2004), valerianol (Carretero, 2000; Heinrich et al.,

2004), bisabolene, caryophyllene, (Heinrich et al., 2004) pacifigorgiol and ledol

(Barnes et al., 2002). Valerenic acids, acetoxyvalerenic, hydroxyvalerenic are

less volatile sesquiterpenic carboxylic acids present in essential oil (Carretero,

2000; Dewick, 2002; Heinrich et al., 2004; Cunha, 2005). Isovaleric and

hydroxyisovaleric acids are liberated when drying and storing the plant which

gives the valerian its characteristic smell (an unpleasant aroma) which is not

noted in the fresh plant (Weiss, 1988; Dewick, 2002).

Valepotriates

The valepotriates are epoxy iridoid esters (Dewick, 2002) that contain valtrate

(Carretero, 2000; Heinrich et al., 2004; Cunha, 2005) which is about 80% of the

valepotriates, acevaltrate (Cunha, 2005), dihydrovaltrate (Carretero, 2000; Heinrich

et al., 2004; Cunha, 2005) isovaltrate (Carretero, 2000; Dewick, 2002; Heinrich et

al., 2004) isovaleroxy-hydroxydihydrovaltrate (Caron and Reidlinger, 1999;

Carretero, 2000). The valepotriates are very unstable molecules as they can be

easily decomposed by heat, humidity or even pH variations

(www.naturaldatabase.com). Therefore, they could only be present in fresh plants.

However, small quantities of these compounds exist when the plant material is

made to dry at a temperature lower than 40C (Carretero 2000). Being insoluble

in water valepotriates could be obtained only in non-aqueous preparations.

Although valepotriates were once thought to be the active ingredients, these

compounds being chemically unstable degrade readily, poorly absorbed and are

not found in infusions and tinctures (Foerster et al., 1984; Houghton, 1988; Lin et

al., 1991; Reichert, 1998). Instead, their degradation products, baldrinals which

are unsaturated aldehydes, are found in such preparations, and may account for

much of valerians sedative effect. The baldrinal and homobaldrinal are the

degradation products of valtrate and isovaltrate respectively (von der Hude, 1986;

Barnes, 2002; Heinrich, 2004).

Other Constituents

Apart from essential oils and valepotriates, the plant contains other bioactive

compounds including amino acids, alkaloids, flavonoids, lignans etc. Arginine,


glutamine, tyrosine, -aminobutyric acid (GABA) are the important amino acid

constituents. The major flavonoids present in V. officinalis are linarin (Fernandez

et al., 2004), methylapigenin, hesperidin (Marder et al., 2003; Fernandez et al.,

2004). Alkaloidal constituents include chantinine, valerine, valerianine, actinidine,

methyl-2-pyrrole ketone (Torssell and Wahlberg, 1967; Franck et al., 1970; Janot

et al., 1979; Duke, 1985). Hydroxypinoresinol is an important lignan found in this

plant (Houghton, 1999). Other valerian constituents are caffeic and clorogenic
acids, -sitosterol, tannins, choline etc (Barnes et al., 2002)

Pharmacological Properties

Valerian has been used as a medicinal herb since at least the time of ancient

Greece and Rome. Hippocrates described its properties, and Galen later

prescribed it as a remedy for insomnia. Pharmaceutical application of valerian is

due to its sedative, anticonvulsant, antidepressant, antihypertensive, hypnotic

effects, antispasmodic and anxiolytic activity. The pharmacological effects of

valerian have primarily been attributed to the valepotriates (iridoid esters), volatile

oils, monoterpenes, and sesquiterpenes constituents (Wang et al., 2010; Ansari

et al., 2013). Lately, the potential cytoprotective effect of aqueous extract of V.

officinalis on human neuroblastoma cells has also been demonstrated (Madureira

de Oliveria et al., 2009). In 1998, valerian was the tenth most popular herbal cure

sold in the United States. Now, valerian extracts are available as dietary

supplements, which are primarily composed of dried root or extracts from the

root, formulated into tablets or soft gelatin capsules are extensively used with an

estimated 210 million doses sold annually in the United States and 125 million

doses sold annually in Europe (Patocka and Jakl, 2010).

Cardiovascular Activity

The coronary dilating and antiarrhythmic effects of valerian extract has been

demonstrated in rabbits, mice and cats. Petkov (1979) reported that valepotriates

an important component of Valerian, prevented the appearance of acute coronary

insufficiency as well as vasopressin-induced arrhythmia, provoked a short-lived

increase of coronary blood flow, and had moderate positive inotropic and negative

chronotropic effects. In mice, valeranone another bioactive compound found in

small quantities in valerian and in larger amounts in its relative, Nardostachys

jatamansii, exerted weak hypotensive effects (Morazzoni and Bombardelli, 1995).

A significant increase in coronary blood flow, a transient fall in blood pressure and

a decrease in heart rate was noticed when cats were intravenously injected with

valerian extracts (Zhang et al., 1982). Valerian is included in a German heart

tonic to maintain neuro-cardiac stability (Mowrey, 1986). In an open, multicenter

trial of 2,243 patients with a variety of functional cardiac disorders, an herbal


combination (valerian, hawthorn, cereus and camphor) was associated with

improvement in 84% of patients (Busanny-Caspari 1986). Circosta et al. (2007)

reported significant anticoronaryspastic, antibronchospastic, antihypertensive

activities from ethanolic and aqueous extracts of V. officinalis L. roots in

anaesthetized guinea-pigs and were found similar to those exhibited by nifedipine.
Anxiolytic Activity
Hattesohl et al. (2008) evaluated the pharmacological profile of different extracts

derived from V. officinalis L. including two commercially available extracts and

the newly developed preparations VAL SE 35E and phytofin Valerian 368.

Therefore tests for sedative, anxiolytic, antidepressant as well as for myorelaxant

properties were conducted in rodents. The results revealed that none of the valerian

extracts displayed sedative or myorelaxant effects when used up to maximum

dosages of 500 or 1000 mg/kg body weight. However pronounced anxiolytic of

the 45% methanolic and 35% ethanolic extract as well as of phyotofin Valerian

368 was noticed in the elevated plus maze test in a dose range of 100500 mg/kg

body weight. Additionally and different from its primary extract (35% ethanolic

extract) phytofin Valerian 368 showed antidepressant activity in the forced

swimming test after subacute treatment. They concluded that anxiolytic and

antidepressant activity may contribute to the sleep-enhancing properties of

valerian. Further, Murphy et al. (2010) reported primary anxiolytic activity of valerian

due to the presence of valerenic acid and the enhanced anxiolytic effect of valerenic

acid in the presence of GABA. During their experiment rats were administered

either ethanol (1ml/kg), diazepam (1mg/kg), valerian root extract (3ml/kg), valerenic

acid (3mg/kg), oral solution of valerenic acid and exogenous GABA (75 mg/kg

and 3.6 mg/kg, respectively) and assessed for the number of entries and time

spent on the open arms of an elevated plus maze. Results showed that there was

a significant reduction in anxious behaviour when valerian extract or valerenic

acid exposed subjects were compared to the ethanol control group.

Sedative/anticonvulsant Activity

Rosecrans et al. (1961) evaluated the biological activity of the different root extracts

of Valeriana officinalis wherein two fractions (V103 and VI15) were found to

potentiate the pentobarbital sleeping time in rats. In mice, intraperitoneal injections

of valerenic acid, valerenal and whole herb extracts produced significant sedation,

ataxia and anticonvulsant effects (Veith et al., 1986). Intraperitoneal injections of

100 mg/kg had sedative effects as strong as barbiturates; doses of 400 mg/kg led

to death. In comparison with diazepam and chlorpromazine, valerian extract had

weak anticonvulsive properties. Valerian root extract (Valdispert) reduced motility

and increased thiopental and pentobarbital-induced sleeping time. Even the aroma


of valerian root exerted sedative effects in mice (Buchbauer et al., 1992). Marder

et al. (2003) reported the sedative and sleep enhancing property of hesperidin, a

compound isolated from V. officinalis. They also demonstrated the ability of 6-

Methylepigenin, another compound from V. officinalis, to potentiate the sleep

enhancing property of hesperidin. Fernandez et al. (2004) further reported the

presence of the flavone glycoside linarin and its sedative and sleep-enhancing

properties that are potentiated by simultaneous administration of valerenic acid.

An intraperitoneal coadministration of linarin (4 mg/kg) and valerenic acid (5 mg/

kg) had sedative and sleep-enhancing effects as evidenced by the remarkable

reduction in the exploration of holes, the time mice spent head dipping and the

number of their rearings as assayed in the holeboard test and also produced a

striking increase in the sleeping time induced by sodium thiopental whereas the

administration of linarin and valerenic acid independently at the above mentioned

doses did not increase the sleeping time induced by sodium thiopental.

Gastrointestinal Activity

Valerian is traditionally used in the treatment of intestinal spasms, colic, and

nervous stomach. Valerian has a bitter flavor, and bitters have historically been

used to enhance appetite and digestion. Valerenic acid, valtrate and valeranone

exert spasmolytic effects in guinea pig ileum through direct effects on smooth

muscle (Hazelhoff et al., 1982; Busanny-Caspari, 1986).

Antidysmenorrheal activity

Valerian seems to be an effective treatment for dysmenorrhea, probably because

of its antispasmodic effects. In a double-blind, randomized, placebo-controlled

trial, 100 students were randomly assigned to receive valerian (dose 255 mg) 3

times daily for 3 days beginning at the onset of menstruation, for 2 consecutive

menstrual cycles. At baseline and during the intervention cycles, the pain severity

was evaluated with a visual analog scale and the systemic manifestations were

assessed using a multidimensional verbal scale. The pain severity at baseline

did not differ significantly between the groups. After the intervention, the pain

severity was significantly reduced in both groups, but the extent of the reduction

was larger in the valerian group, with the difference between the 2 groups being

statistically significant. The total scores of the systemic manifestations associated

with dysmenorrhea decreased after the intervention, but there was no significant

difference between the groups, with the exception for syncope (Mirabi et al., 2011).

Attention enhancing activity

In Germany valerian is sometimes used to treat attention deficit hyperactivity

disorder (ADHD) in children (Cavazzuti, 1969). German studies during 1960s


reported that valerian could antagonize the hypnotic effects of alcohol, enhancing

concentration and coordination (Mowrey, 1986). In a randomized, placebo-

controlled, double-blind study, valepotriates demonstrated a dose-dependent

increase in concentration abilities in 24 healthy volunteers. Also, valeropotriates

when given in combination with alcohol did not affect blood alcohol levels, sedative

effects or effects on driving performance (Mayer and Springer, 1974). There are

no controlled trials evaluating valerians use in treating attention deficit hyperactivity

disorder (ADHD) (Mowrey, 1986).

Other Neurological Activity

Unlike diazepam, valerian did not affect spontaneous ambulation and rearing or

approach-avoidance conflict in mice in a water-lick conflict test. On the other

hand, valerian and imipramine significantly inhibited immobility induced by a forced

swimming test in rats and significantly reversed reserpine-induced hypothermia

in mice, leading researchers to conclude that valerian may be a useful

antidepressant (Morazzoni and Bombardelli, 1995). Among 80 hospitalized

geriatric patients enrolled in a placebo controlled trial for 14 days, those assigned

to an aqueous valerian extract had significant improvements in mood and

behavioral disturbances as well as sleep (Kam-kohl et al., 1984). Among 121

patients with sleep disturbances enrolled in a controlled trial, those assigned to

an alcoholic extract of valerian (600 mg daily for 28 days) had a significant

improvement in depression, mood and global functioning as well as sleep; no

significant side effects were reported (Vorbach et al., 1996).

Propagation

Conventional Methods

The plant is conventionally propagated through seeds as well as from root cuttings.

Although Valerian can grow in any ordinary soil, but prefer moist humus rich loam

having pH of 6-7. It is often found to flourish in damp and shady places, though

some drought resistant forms also occur in chalk and limestone hills. The plant

may easily be propagated by portions of the old rootstocks either in the autumn

months or in spring. The divisions of the rootstock are spaced 30 cm 60-90 cm.

A good crop of the drug may be harvested in the autumn season if the rootstock

divisions are planted very early in autumn in time to become well established

before the onset of frost. The summer cultivation requires the weeding of beds

and cutting off all floral stalks to avoid exhaustion and promote the formation of

larger rootstocks. Soil should be rich in phosphorous (or supply a fertilizer) as the

leaves turn purple when the plant is grown in phosphorous-deficit soil. Seeds are

directly sown by simply pushing them into the soil during spring season. Avoid


covering them as they require light and a temperature of 68 to 70F to germinate.

Their germination takes one to two weeks. The nursery raised seedlings need

consistent moisture, grow slowly, need protection from faster growing weed and

are first transplanted at a spacing of 18-20 cm in rows 30 cm apart. The seedlings

are then re-transplanted to their permanent sites at the same spacing as used for

root stock divisions. Application of farmyard manure with proper mixing into the

soil before the transplanting of seedlings favours the good growth of the plant.
The formation of large root stock is promoted by raising a low ridge of soil along

the bases of plant. For the better development of the rhizome flowering tops must

be cut off as they appear. Many of the young plants do not flower in the first year,

but produce a luxuriant crop of leaves, and yield rhizome of good quality in the

autumn. The rhizomes and roots of the plants, propagated from the divisions of

the rootstock may be harvested in the autumn of the first years growth, though

the yield is generally low. The bud-raised plants do not attain a size suitable for

harvesting before the end of second growing season (Anonymous, 2003).

In Vitro Methods

Abdi and Khosh-khui (2007) established a highly reproducible micropropagation

protocol through adventitious shoot bud regeneration from leaf explants. They

reported influential role of size as well as orientation of leaves, type of plant growth

regulators on shoot bud induction. Maximum shoot regeneration (74 %) was

observed from abaxial side of leaves with the production of about 7.4 mean number

of shoot on MS (Murashige and Skoog, 1962) supplemented with 1 mg/L 6-Benzyl

adenine (BA). Multiplication of shoots was further achieved by transferring shoots

regenerated from leaves on MS medium supplemented with Isopentenyl Adenine

(2-ip) and indole-3-acetic acid (IAA) alone or in combination. The highest

multiplication rate (93.3 %) was observed on combination of 2-iP (6 mg/L) and

IAA (0.3 mg/L) which induce the formation of about 13.7 mean shoot number. -

naphthalene acetic acid (NAA) at 1 mg/L was the optimal treatment for in vitro

root induction in their study.

Abdi et al. (2008) further evaluated the potential of silver nanoparticle (35 nm

average size) for removal of bacterial contaminants from in vitro cultures of V.

officinalis. Their experiment involved the treatment of explants with different

concentration (25, 50 and 100 mg/L) of silver nanoparticle before and after surface

sterilization for three different time periods (30, 60 and 180 min). Explants were

cultured on MS medium supplemented with 5 mg/L KN and 0.1 mg/L NAA. Their

results revealed the highest percentage (89 %) of disinfected explants by using

100 mg/L of silver nanoparticle solution for 180 min after surface sterilization

while same amount of silver nanoparticle solution used for the similar duration

resulted in only 32 % disinfected explants when the nano silver solution was used


Figure 1: Steps involved in micropropagation protocol of Valeriana officinalis L.

Step A. In vitro induction of shoots from nodal explant in culture media

in V. officinalis; Step B. Multiplication and proliferation of cultures for

enhanced production of V. officinalis; Step C. In vitro rooting in tissue

culture raised plantlets; Step D. Acclimatised in vitro raised plantlet

ready to transfer in field


before surface sterilization. However, no affect on contamination was observed

when silver nanoparticle solution was used without surface sterilization of explants.

However, the authors did not notice any affect of nano silver solution on the

characters measured. On the basis the above mentioned experiment, it was

concluded that NS had a good potential for removing of the bacterial contaminants

in plant tissue culture procedures.

Muntean (2008) reported somaclonal variation among the valerian (V. officinalis),

micropropagated plantlets. During their experiment optimum callus induction was

observed on Murashige and Skoog (MS) medium supplemented with 10 mg/L

2,4-dichlorophenoxyacetic acid (2,4-D) while maximum regeneration was achieved

upon transferring the callus to MS medium containing combination of 2.5 mg/L

BA and 0.5 mg/L IAA. In vitro rooting of regenerated shoots was obtained on MS

medium containing 1 mg/L indole-3-butyric acid (IBA). About 19 plantlets

regenerated from the callus were successfully hardened followed by their transfer

to the greenhouse. The genetic analysis through RAPD marker was further

performed during their study in order to investigate any detectable variation among

the parent plant and the regenerants. They found four somaclonal variants out of

19 regenerants and the obtained somaclones were different from the parental

plants by at least one polymorphic amplification product. However, the remaining

regenerants were found to be genetically stable as compared to the elite donor

plants. The seven primers generated 13 new RAPD markers in the somaclonal

variants that were not found in the donor plant fingerprints. Moreover, two out of

four somaclonal variants were polymorphic, compared with the donor plant, with

two or more primers.

Reza et al. (2009) reported indirect shoot regeneration from leaf and petiole derived

callus. In their study callus was developed on MS medium supplemented with

different combinations of different auxins (2,4-D, NAA, picloram) and N 6-

furfuryladenine (KN) from petiole and leaf segments wherein highest callus

induction frequency was obtained on 1.0 mg/L 2,4-D for petiole (98 %) and leaf

(100 %) explants. For shoot regeneration the calli were transferred to a medium

supplemented with various concentrations of BA or KN and they observed

maximum shoot regeneration frequency on 0.2 mg/L BA or KN with the production

of about 13 adventitious shoots per callus segments derived from leaves. MS

medium supplemented with 1 mg/L NAA was reported to be the optimal rooting

media during their investigation on which 100 % regenerated shoots developed

roots with an average of 10.7 roots per shoot within 3 weeks. The in vitro raised

plantlets were further successfully hardened and transferred to greenhouse with

80 % survival rate.

Zebarjadi et al. (2011) developed in vitro regeneration protocol via indirect

organogenesis from leaves, stems and roots explants excised from 35 day old in


vitro germinated seedlings. The highest callus formation frequency was achieved

on MS medium augmented with 0.1 mg/L KN and 2 mg/L 2,4-D for leaf and root

explants (100 %) while MS medium containing 0.1 mg/L KN and 10 mg/L 2,4-D

induce maximum callus (100 %) from stem explants. Their results further revealed

maximum shoot regeneration frequency of 60 % on MS medium containing 0.5

mg/L NAA and 2 mg/L BA in leaf-derived calli. Regenerated shoots were

subsequently transferred to medium lacking growth regulators and then MS

medium supplemented with 2 mg/L IBA for in vitro rooting.

Tansaz et al. (2014) investigated the effects of three different culture media

formulations viz. MS, B5 (Gamborg et al. 1968) and MB (new combined medium

containing half MS and B5 salts and vitamins, 10 ml/L Fe EDTA) and various

combinations of plant growth regulators (BA and NAA) on in vitro regeneration

and growth parameters of V. officinalis. Excised crown with 2-3 segments having

one bud was used as explants. Among the different media tried during their study,

MB medium exhibited highest regeneration frequency alongwith maximum

induction of mean number of microshoots. However, highest percentage of

regenerated plantlet (80 %) with maximum mean shoot number (9.43) and mean

shoot length (7.04 cm) was obtained on MB medium supplemented with BAP (0.5

mg/L) in combination with NAA (0.25 mg/L). B5 medium without any PGR exhibited

the lowest regeneration frequency of 40 % which increased to 75 % on

supplementation with 1.5 mg/L BA and 0.25 mg/L NAA.

Ghadheri and Jafari (2014) for the first time reported efficient and rapid propagation

protocol of V. officinalis for large-scale production of uniform raw materials for

future pharmaceutical compound extraction as well as the quantification of the

two industrially important bioactive compounds (i.e. valerenic acid and valtrate)

accumulated in tissue culture-raised plants. Petiole and leaf explants were used

for the callus induction and the three types of callus could be recognized by color

(creamy green, light or dark green) and texture (compact, nodular and friable)

depending on the PGR concentrations/combinations used. Compact, nodular,

yellowish green callus usually formed through leaf and petiole explants on medium

containing 4.65 or 9.30 M KN in combination of 2.26 M 2,4-D showed the highest

regenerative capacity with the formation of maximum shoot buds in both types of

explants. IBA at a concentration of 2.46 M was found to be the best rooting

treatment, which promoted the highest frequency of rooting (98 %). Successful

acclimatization of rooted plantlets was achieved in potting medium containing 5:1

mixture of peat and perlite with 95.34 % survival rate. The contents of valtrate and

valerenic acid in biomass extracts from petiole-derived plants were significantly

higher than leaf-derived plants as well as the seed-raised field-grown plants.

Maximum valtrate (6.98 mg g1 DW) and valerenic acids (3.02 mg g1 DW)

contents were quantified in root tissue of petiole derived plants raised on medium


with higher concentration of Kn (9.30 M) in combination with 2.26 M 2,4-D. Further

they also conducted RAPD analysis to screen tissue culture induced genetic

variations in high level valerenic acid-accumulated plants using 15 RAPD primers

(OPA 1-15) wherein only 4 primers viz. OPA-3, 5, 8, and 12 produced reproducible

and scorable bands ranging from 3 to 8, with the generation of 104 amplification

products of 200 to 1400 bp. Their results showed no differences between the

plants raised from in vitro cultures but a slight variance in only one electrophoresis
band, produced by OPA-8 primer, with seed-raised field grown plant was reported

by them which demonstrates significant difference of these plants with seed raised

plants from the metabolite content point of view. They lastly concluded that tissue

culture-raised plants had higher genetic stability than did the seed-propagated

plants.

Bhat and Sharma (2015) evaluated the effect of light and low temperatures as

well as pretreatment of seeds with GA3 on seed germination along with the

development of in vitro propagation protocol through shoot tip explants. Their

results revealed increased germination rate (48 %) in unchilled seeds pretreated

with GA3 (200ppm) for 24 hr in dark as compared to non pretreated seeds with

only 10 % germination under dark conditions. Further maximum germination rate

of 60 % was obtained through pre-chilling treatment of seeds for 72 hr and keeping

them in light conditions during their study. MS medium supplemented with BA

(1mg/L) and IAA (0.5 mg/L) was found to be the optimal treatment for shoot

multiplication and elongation with 50 % response.

Genetic transformation and Elicitation studies for enhanced

metabolites production

Grnicher et al. (1992) established hairy root cultures in V. officinalis

var. sambucifolia by infection of sterile plantlets with Agrobacterium

rhizogenes strain R1601. Different hormone-free liquid media were used for the

growth of transformed roots and the yield of different valepotriates viz. isovaltrate,

valtrate, didrovaltrate, isovaleroxyhydroxydidrovaltrate content was quantified

through HPLC. The highest overall valepotriate content (10.3 % dry wt) which

was about 4 times the amount found in the roots of 9-month-old nontransformed

plants, was observed in half strength Gamborg B5 medium supplemented with 2

% sucrose. They further demonstrated the production of about 44 mg/g dry wt

valepotriates from hairy roots cultured in MS liquid medium supplemented with 2

% sucrose for 50 days.

Zebarjadi et al. (2011) carried out genetic transformation studies in V. officinalis

under in vitro conditions by co-cultivating leaf, stem and root derived callus

with two different strains of A. rhizogenes viz. AR15834 and LBA 9402 for the


induction of hairy root cultures. During their experiment, leaf, stem segments

(1 cm2) and calli were immersed in a suspension of A. rhizogenes (OD600 =

0.6-1) for 5 to 7 min followed by subsequent co-cultivation on solidified MS

medium without any hormone and MS medium containing (0.1 mg/L NAA+1

mg/L BAP), (0.5 mg/L BAP+0.5 mg/L NAA+0.5 mg/L 2,4-D) for 2 days in

darkness. Explants were then transferred onto selective medium containing

250 mg/L cefotaxime and 20 mg/L rifampicin, and further subcultured onto
fresh selective medium every 15 days. Un-inoculated explants were used as

a control and after 28 days of culture, the number of produced roots on

selective medium to the number of cultured explants was determined. The

results obtained demonstrated LBA9402 strain to be better than AR15834

strain in terms of both mean number of hairy root formation (68.62%) and

production of the respective hairy root (90%) while leaf and root calli were

found to be the best explants for transformation with both the strains. The

optimal growth of the hairy roots occurred on selective MS medium containing

20 mg/L rifampicin and 250 mg/L cefotaxime without any plant growth

regulators. Genomic DNA of putative transgenic and non-transgenic (control)

regenerated hairy roots were analyzed by PCR for presence of rol B gene,

using specific primers.

Torkamani and Samadi (2014) studied the influence of four different levels of

Calcium and Potassium (half, full, two and four fold concentrations) on enhanced

accumulation of valerenic acid in hairy root cultures of V. officinalis. Hairy root

cultures were established via A. rhizogenes A13 strain using various explants

derived from 42-day-old sterile seedlings and the transgenic status of the roots

were confirmed by the presence of rol B and rol D amplified products during

Polymerase chain reaction (PCR) analysis. The induced hairy root cultures of V.

officinalis were maintained in the media and after 35 days were used for

investigations of biomass accumulation and valerenic acid production through

high-performance liquid chromatography (HPLC). The results showed highest

valerenic acid accumulation of about 0.690.03 mg/g DW from hairy roots cultured

on media with two-fold calcium, which was 1.92 times higher than normal culture

(0.36 0.01 mg/g DW). The study also demonstrated the positive effect of double

concentration (880 mg/L) of calcium in media on growth of transformed hairy

roots. However, they further reported decrease biomass accumulation but 1.2

fold increase in valerenic acid production than control roots on the media with

3.32 mg/l (four times of control concentration) potassium concentration. Their

results revealed the positive role of calcium in intensification of the valerenic acid

production in V. officinalis hairy roots cultures.

Tokramani et al. (2014a) also studied the effect of exposure time (3 and 7 days)

of different calcium and magnesium concentrations (2 to 6 fold than basal MS


medium) on the growth of hairy roots and valerinic acid production in hairy root

cultures. They used hairy root cultures grown in MS liquid medium without elicitors

for 8 weeks for the quantification of valerinic acid through HPLC. Their results

revealed highest production of valerenic acid (1.83 0.06 mg/g DW) in hairy root

cultures exposed to 6-fold calcium for 7 days which was 7.9 times higher than

that of control culture (0.23 0.01 mg/g DW) while (6-fold) of magnesium at

exposure time of 7 days resulted in the accumulation of valerenic acid at (1.11

0.03 mg/g) DW which is 4.2-fold higher than that of non-elicited control.

Torkamani et al. (2014b) further increased valerenic acid production in the selected

hairy root line LeVa-C4 through the use of three different elicitors viz. Fusarium

graminearum extract (FE), methyl jasmonate (MJ) and salicylic acid (SA). They

use wild-type strain A13 of A. rhizogenes for the induction of hairy roots and 23-

day-old hairy root cultures were further inoculated into 25 ml MS liquid medium

supplemented with various concentrations of MJ (50, 100, 200 M), SA (50, 100,

150 M) or FE (0.5, 1, 2% v/v) for different exposure times (3 and 7 days). They

found 1% FE and 100 M L-1 MJ highly efficient for increased levels of valerenic

acid after 7 days of elicitation which were 12.31 (3.02 mg/g DW) and 6 times

(2.296 mg/g DW) higher than that of non-elicited control (0.24 mg/g DW)

respectively. They further demonstrated that FE did not exert any negative effects

on biomass yield of hairy root and no significant increase in valerenic acid content

was noticed through the use of salicylic acid as elicitor.

Parizi et al. (2014) established hairy root cultures through A. rhizogenes (strain

ATCC 15834) mediated genetic transformation and further determine the effect

of different media viz. MS, B5 media (full and half strength), N6 medium and a

modified MS medium without phytohormones, on the growth parameter of hairy

roots. They also studied the effect of different NH4+/NO3- ratios (00:20, 10:20,

20:20, 20:10, 20:00, 20:40) in MS medium on hairy root growth and the effect of

these treatments were evaluated after 21 days of culture in relation to hairy root

growth. Their results revealed B5 and B5 media to be the best basal medium

with the production of maximum dry weight of hairy roots i.e. 3.85 and 3.67 g/L

respectively while N6 medium exhibited lowest hairy root growth with the production

of only 0.86 g/L dry weight after 21 days. Further among the different NH4+/NO3-

ratios in MS media tested, MS medium supplemented with a 20:20 mM and 20:40

mM ratio of NH4+ to NO3- produced the maximum biomass of 1.80 and 1.62 g/L

respectively after 21 days.

Acknowledgement

Ms. Taiba Saeed acknowledges UGC, for providing financial assistance under

the scheme SRF-Maulana Azad National Fellowship (Award No. MUS-UTT-2624).


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72
Some Depletive Abstract

Plants of he Kumaon Himalaya of Uttarakhand is rich in floristic

Kumaon
Himalaya Used
in Unani
T

composition. It is a land of diverse cultures and ethnic groups. Traditionally the
folk people utilize the vegetation of their ambient environment in the form of different
products as food, fuel wood, fibre, fodder, timber, medicine, etc. During
ethnobotanical explorations of different forest divisions in this region, the authors
collected a large number of information on local plants which are used as folk
Medicine for

drugs for treatment of a wide range of health related problems. This study has

Treating Non- also yielded information on some medicinal plants that are becoming scarce in

the area due to overexploitation, unsustainable harvesting practices and loss of

Communicable

habitat. A review of literature revealed that most of these plant species find use in

Diseases and

Unani medicine for treating various non- communicable diseases (NCDs). Hence,

an attempt has been made to highlight such species in this report. For each plant
their

species are given the correct botanical and vernacular names, habit and habitat,

Conservation

the part used, medicinal use(s) and other observations. The threat of extinction of

Strategies# these useful and commercially viable medicinal plants has been discussed and

also the strategies for their conservation have been suggested.

*Zaheer Anwar Ali,

Sarfraz Ahmad Keywords: Ethnobotanical explorations, Endangered medicinal plants,

and Conservation, NCDs, Kumaon.

Shariq Ali Khan

Introduction

Survey of Medicinal Plants Unit,


Kumaon region of Uttarakhand is situated between 28o 43 55"- 30o 20 12" N

Unani Medicine (CCRUM),

Post Box 70, Aligarh - 202001 (U.P.)

latitude and 78o 44 30"- 80o 18 45" E longitude in Western Himalayas. There is

wide range of natural habitats which provide varied plant life including medicinal

and aromatic plants. It is also the land of diverse culture and ethnic groups. This

Himalayan region has always been reputed as a steady supplier of a good number

of potent medicinal herbs and also one of the leading regions in the use of herbal

drugs. A number of reports on the use of native floras in traditional medicine of

many cultures of this region have been published (Agnihotri et al., 2003; Arya and

Prakash, 1999; Arya et al., 1999; Aswal, 1992; Bhatt and Gaur, 1992; Bhatt et al.,

1987-88; Datt and Lal, 1993; Garbyal et al., 2005; Gupta, 1960; Joshi, 1993;

Joshi et al., 1993; Kalakoti and Pangtey, 1988; Pandey and Pande, 1990; Pandey

et al., 1989,1995; Pangtey et al., 1989; Pant and Pandey, 1998; Rawat and

Pangtey, 1987; Shah and Gupta, 1976; Shah and Jain, 1988; Shah and Joshi,

1971; Singh and Ali, 1997, 1998; Singh, et al., 1980, 1987; Singh and Maheshwari,

#This paper is based on the data presented by the first author in National Seminar on The role of

Unani medicine in non-communicable diseases, organized by Central Council for Research in

Unani Medicine, New Delhi at New Delhi on 14-15 January 2015.

* Author for correspondence


1990, 1993, 1994). Hence this area was selected for an extensive ethnobotanical

study. The main objective of this field study, besides collecting folk medicinal

claims prevalent among the indigenous communities, was to prepare an inventory

of existing Unani medicinal plants and to record the current status as well as

distribution of highly exploited medicinal taxa. Ethnomedicinal uses of plants

collected during the present survey have been published by us (Ali et al., 2008,

2010a; 2010b, 2013a, 2013b, 2013c, 2014a, 2014b; Ali and Ahmad, 2008, 2010).
In this communication an enumeration of some depletive medicinal plants of

Kumaon is presented.

Fieldwork was carried out during the period 1999-2008. The study area includes

different forest divisions of Bageshwar, Champawat, Nainital, Pithoragarh, and

Udham Singh Nagar districts. In the course of this investigation, it was found that

wild plants are still predominantly in use by the natives for their health purposes

and all the basic raw drugs are collected from the forest without replenishing the

growing stocks. Moreover, the natural habitats are being disturbed due to a variety

of factors such as diversion of land for expansion of agriculture, dwellings and

other developmental activities; recurring forest fire; soil erosion; invasion of some

alien weed species; etc. Consequent to this, populations of some commonly used

drug yielding plants have become reduced in this area. A literature survey was

carried out. The Unani Pharmacopoeia of India, Part-I (2007-2009), National

Formulary of Unani Medicine, Part-I (Anonymous, 1981) and other monographs

published by the CCRUM (Standardization of Single Drugs of Unani Medicine,

Part-I (1987), Part-II (1992) and Part-III (1997) were mainly consulted for general

information as well as therapeutic uses and it was found that most of these plant

species are mentioned for treating various non-communicable diseases. The

present communication highlights such species along with pertinent information.

Earlier authors have reported many threatened medicinal and aromatic plants

that were extracted for commercial purposes from this region and suggested their

conservation (Pangtey and Samant, 1988; Shah, 1983; Singh, 1993; Shah and

Kapoor, 1978; Sinha, 1975). This contribution is an addition to the above reports.

The information presented herein is mainly based on our field observations and

enquiries made with knowledgeable village elders belonging to various indigenous

communities and officials of the forest department. The study might be useful for

planning strategies on conservation, and cultivation of medicinal flora particularly

threatened taxa of the region. In the following enumeration the plants are listed in

alphabetic order by their botanical names with respective family, vernacular names,

habit and habitat, medicinal uses and locality. This is followed by a remark on

availability and threat categories. All voucher specimens were prepared and

preserved in the herbarium of Survey of Medicinal Plants Unit, Regional Research

Institute of Unani Medicine, Aligarh (U.P.), India.


Enumeration

Acorus calamus L. (Araceae)

Vernacular names : Bach, Boj, Ghiroch, Ghurvach (L); Waj-e-Turki (U)

Habit & habitat : A semiaquatic, perennial herb with aromatic rhizome.

Growing in wet places like edges of pond and streams

near villages.

Medicinal uses : Rhizome is anthelmintic and also given for hoarse-ness

of voice, dog bite and stomachache.

Locality : Champawat, Nainital, Pithoragarh, Udham Singh Nagar

Remark : Not observed in the forest but now, cultivated by the forest

department.

Berberis aristata DC. (Berberidaceae)

Vernacular names : Kingor (L), Darhald (U)

Habit & habitat : An evergreen shrub. Found in association with other

species of Berberis along roadsides.

Medicinal uses : Root is commonly used for controlling diabetes. Root

extract is instilled in eye for redness.

Locality : Nainital, Pithoragarh

Remark : Overexploited in past for trade. It is an endangered

taxon.

Bergenia ciliata (Haw.) Sternb. (Saxifragaceae)

Vernacular names : Silphori, Patharphori, Pashanbhed (L), Pakhanbhed (U)

Habit & habitat : A perennial herb with thick rootstock, found in forest,

especially on moist rocky slopes.

Medicinal uses : The root of this plant has from very early times been in

much repute for its medicinal properties, particularly for

kidney stones. It is also used for diabetes, anuria and

furunculosis.

Locality : Bageshwar, Champawat, Nainital, Pithroragarh

Remark : Presently common, but the population of this taxon is on

decline due to continuous over collection by the

unauthorized collectors.


Cinnamomum tamala Nees (Lauraceae)

Vernacular names : Kakaria (L), Sazaj Hindi (U)

Habit & habitat : An evergreen small tree, found in shady places in forest.

Medicinal uses : Leaves are used for cough and cold.

Locality : Nainital, Champawat, Pithoragarh
Remark : Leaves and stem bark are heavily extracted which have

much value for condiments. Low-Risk Near Threatened.

Now, it is cultivated.

Curculigo orchioides Gaertn. (Hypoxidaceae)

Vernacular names : Kali musli (L), Musli Siyah (U)

Habit & habitat : Herb with tuberous root stock. This species is

characteristics in Sal trees in the Tarai-Bhabar belts.

Medicinal uses : Root is commonly used for sexual weakness and

leucorrhoea.

Locality : Udham Singh Nagar

Remark : The tuberous root is heavily exploited and also greedily

eaten by wild animals. It is a vulnerable taxon.

Dactylorhiza hatagirea (D. Don) Soo (Orchidaceae)

Vernacular names : Hathajari (L), Salab Panja (U)

Habit & habitat : A terrestrial orchid with tuberous roots which are digitate

or finger shape. Rarely found in damp and shady places

in Oak forest.

Medicinal uses : Tubers are commonly used as general tonic and also

believed as an aphrodisiac agent.

Locality : Bageshwar, Pithoragarh

Remark : It is a critically endangered taxon. Export already banned

(Jain and Sastry, 1980). It is cultivated in Nandadevi

Biosphere Reserve by the tribal (Maikhuri et al., 2002).

Drimia indica (Roxb.) Jessop (Liliaceae)

Vernacular names : Jangli Piyaz, Koli Kanda(L), Isqeel (U)

Habit & habitat : A scape-bearing herb rarely found in forests of Tarai.


Medicinal uses : The bulb is commonly used for burning micturition.


Remark : A vulnerable taxon.

Eulophia herbacea Lindl. (Orchidaceae)

Vernacular names : Salab Misri (L, U)

Habit & habitat : Herb with tuberous roots, rarely found in forest.

Medicinal uses : Root is commonly used for leucorrhoea and as a general

tonic.

Locality : Nainital

Remark : An endangered taxon. Export already banned (Jain and

Sastry, 1980).

Gloriosa superba L. (Liliaceae)

Vernacular names : Kalihari (L), Muleem (U)

Habit & habitat : A climbing herb with tuberous rootstock; climbing by means

of leaves. It is readily recognized by its beautiful flowers.

Rarely found in outskirts of villages in Sub-Himalayan

forest tracts.

Medicinal uses : Tuber is used for joint pain.


Remark : Due to removal of tubers, the whole plant is destroyed as

such wild populations have been reduced very much. It is

an endangered taxon.

Hedychium spicatum Buch.-Ham. ex J.E. Sm. (Zingiberaceae)

Vernacular names : Kapoor kachri, Jangli Haldi (L), Kapura Kachri (U)

Habit & habitat : A robust herb with horizontal root, found in moist and shady

places in forest.

Medicinal uses : Root is regarded by the natives as a remedy for

inflammation and also used for urticaria and kidney stones.

Locality : Champawat, Nainital, Pithoragarh, Udaham Singh Nagar

Remark : It has become vulnerable.


Picrorhiza kurrooa Royle ex Benth. (Scrophulariaceae)

Vernacular names : Kutki (L, U)

Habit & habitat : A perennial herb found in the forest between altitudes of

3300-4300m.

Medicinal uses : Root is given for leucorrhoea.
Locality : Pithoragarh

Remark : An endangered taxon.

Rauwolfia serpentina (L.) Benth. ex Kurz (Apocynaceae)

Vernacular names : Sarpgandha, Swaitbarua (L), Asrol (U)

Habit & habitat : A perennial under shrub. It is seen growing wild in shady

situations in Sal forest of Sub-Himalayan forest tracts.

Medicinal uses : The root is believed to be an antidote to snake poison and

also considered effective for fever and abdominal pain.


Remark : Ramnagar forest is the natural habitat of this plant where

it is still occurs in good quantity. This species has been

depleted in other forest areas due to excessive exploitation

for trade in past. It has poor regeneration. Now, it is under

cultivation.

Swertia angustifolia Buch.-Ham. ex D. Don (Gentianaceae)

Vernacular names : Chiraita(L), Chiraeta Shirin (U)

Habit & habitat : An erect herb, found in the forest up to 1800m.

Medicinal uses : Leaves are used for fever in pneumonia.


Remark : It is highly threatened due to trade and destruction of

natural habitat. Now, it is under cultivation.

Taxus baccata L. sbsp. Wallichiana (Zucc.) Pilger (Taxaceae)

Vernacular names : Thuner, Laventa (L), Talispatr (U)

Habit & habitat : An evergreen tree. Found in the evergreen forest at higher

altitudes associated with other species of gymnosperms.


Medicinal uses : Leaves and stem bark are commonly used to prevent cold.


Remark : The plant yields taxol, a remedy for cancer. It is threatened

to illegal trade and destruction of habitat. Now, it is

cultivated by the forest department.

Valeriana hardwickii Wall. (Valerianaceae)

Vernacular names : Samoy (L), Taggar (U)

Habit & habitat : Herb with white flowers. Found in forest, but also grows

along roadsides near villages.

Medicinal uses : Root is used for headache. It is also used as an insect

repellent by the natives.

Locality : Bageshwar, Nainital, Pithoragarh

Remark : Often exploited commercially, in the past. It is vulnerable

taxon.

L = Local name; U = Unani name

Discussion

This communication has brought to light 15 medicinal taxa which are dwindling

in the forests of the present study area due to various causes. Among other

factors, continued exploitation, non-sustainable harvesting practices and

destruction of natural habitats are largely responsible for their depletion. It is

predicted that some more plants of the area will be decreased in number

soon if destruction of forests and their over-exploitation continue to occur.

Medicinal plants wealth of a region is one of the vital resources having

important bearing on human health, regions economy and environment.

Therefore, preservation of this heritage along with original habitats must be

protected on priority basis. Such observations are of special significance for

the area where there is a threat to the natural habitats and vegetation owing

to increasing human interference. In the situation prevailing here some

measures for conserving diversity of medicinal plants are listed below:

1. Large scale cultivation and domestication of wild medicinal species used

in pharmaceutical industries and local medicine should be encouraged

by the local inhabitants who may reduce the pressure on existing wild

population of the plants.


2. Public awareness programmes on conservation of wild medicinal plants

should be intensified.

3. Promoting the rationale and sustainable utilization of medicinal plants.

4. Standard methods of cultivation, i.e., agro-technology and preservation

of high demand endangered medicinal plants of the area should be

developed.
5. Sensitive habitats of the threatened flora should be protected on priority

basis.

6. Illegal extraction of medicinal plants from the wild should be curbed.

7. Social forestry operation of fuel, fodder and fibres species should be

encouraged.

8. In order to protect and propagate the threatened species, botanical gardens

should be established at different agro-climatic zones.

9. Germplasm of rare species should be collected and maintained under

protected conditions in the field, plantation, botanical gardens, clonal

repositories, gene bank or in plant tissue culture repositories.

10. Endangered medicinal plants which have poor regeneration their breeding

system should be studied. Through various tissue culture and micro

propagation techniques plants can be regenerated.

11. Local medicine men should be involved in the conservation efforts since

they use plant remedies in their homes and are generally respected by

the villagers.

12. Harvesting or collection has very much importance. It should be done

through trained and experienced plant collectors who could identify the

medicinal plants properly and aware of the methodology of preservation

of plants and their parts used in medicine.

13. Training should be provided regularly to the persons engaged in the

collection of crude drugs. They should be trained for proper and scientific

methods of collection, right time of medicinally important plant parts without

damaging whole plant. For the collection of different plant parts following

points should be taken into consideration:

a. Seeds-should be collected after attaining full maturity; b. Leaves-after

flowering stage; c. Stems-after leaf fall or fruiting stage; d. Flowers-during

full flowering stage; e. Fruits- after full ripening; f. Bark-after raining season;

g. Root or root bark- should be collected from the damaged tree or after

attaining full growth and the plant should be replenished in nature.


Acknowledgements

We are highly grateful to the Director General, Central Council for Research in

Unani Medicine, New Delhi, for providing necessary facilities for carrying out this

investigation. We would also record our gratitude to all the informants who

cooperated in the collection of information presented herein.

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Quality Abstract

Evaluation of erbal products have become a major part in curing the

Jawarish-e-Ood
Kibreet
1*Rampratap Meena,
H

different kinds of human ailments. In India, the Unani system of medicine consists
of large number of herbal products to cure the different type of diseases. Jawarish-
e-Ood Kibreet is a Unani herbal formulation prepared in combination of ingredients
like Ood, Dana-e-Heel Khurd, Dana Heel Kalan, Sazaj Hindi, Post-e-Turanj,
Tabasheer, Nana Khushk, Darchini, Gul-e-Surkh, Zarambad, Qaranful, Jauzbuwa,
2D. Ramasamy,

Bisbasa, Nabat Safaid and Arq-e-Kibreet. It is prescribed in the treatment of Zof-

2S.
Mageswari,

e-Meda (weakness of the stomach) and Zof-e-Ishteha (anorexia) disorders.

2P. Meera Devi Sri,

3Shamsul Arfin Evaluation of Unani formulations has become a fundamental requirement of the

and research organizations. Based on the available sources, an attempt has been

3Aminuddin
made to evaluate the drug Jawarish-e-Ood Kibreet. To evaluate the drug the

parameters like macroscopic, microscopic, determination of moisture content,

1Drug Standardisation Research

Institute, PLIM Campus, ash values, bulk density, pH values, extractive values, TLC and analysis of quality

Kamla Nehru Nagar, control parameters viz. heavy metals, microbial content, aflatoxins and pesticide

Ghaziabad-201002

residues were performed. The evaluated data shall help to lay down

2Regional Research Institute of pharmacopoeial standards for the drug Jawarish-e-Ood Kibreet and also in

Unani Medicine, producing quality and efficacious products having batch to batch consistency.

1 West Mada Church Street,

Chennai-600013

Keywords: Jawarish-e-Ood Kibreet, Powder microscopy, Physico-chemical, TLC

3Central
Council for Research and WHO parameters

in Unani Medicine,

61-65 Institutional Area,

Janakpuri, New Delhi-110058 Introduction

Jawarish-e-Ood Kibreet (Anonymous, 2006) is one of the ancient commonly used

Unani formulations. The poly herbal formulation is prepared using 15 ingredients

(Table 1). The drug is prescribed for the treatment of Zof-e-Meda (Weakness of

the stomach) and Zof-e-Ishteha (Anorexia) disorders. Evaluation of Unani

medicines with the perspective of safety, efficacy and quality will not only preserve

the traditional heritage but also rationalize their uses of Unani medicines in the

health care.

Due to lack of standards and quality control methods there are batch to batch

variations in the similar formulations. The main requirement of a standardisation

is to establish the presence of each ingredient in the formulations (Bandaranayake,

2006; Myers and Cheras, 2004). The present study is an attempt to evaluate the

drug by applying modern parameters such as microscopical, physico-chemical,

thin layer chromatography and WHO parameters viz., microbial load, aflatoxin,

heavy metals and pesticide residue.



Table 1: List of the raw drugs of Jawarish-e-Ood Kibreet Formulation

S. Unani name Botanical/ Part used Quantity

No. English name taken

for SOP

1. Ood Aquilaria agallocha Roxb. Heart wood 20 g.

2. Dana-e-Heel Elettaria cardamomum Seed 6 g.

Khurd (L) Maton.

3. Dana Heel Amomum subulatum Seed 6 g.

Kalan Roxb.

4. Sazaj Hindi Cinnamomum tamala Leaves 6 g.

(Buch. Ham.)

Nees & Eberm.

5. Post-e-Turanj Citrus medica Linn. Pericarp 6 g.

6. Tabasheer Bambusa bambos Druce. Bamboo Manna 6 g.

7. Nana Mentha viridis Linn. Aerial part 6 g.

8. Darchini Cinnamomum zeylanicum Inner stem bark 6 g.

Blume.

9. Gul-e-Surkh Rosa damascena Mill. Flower 10 g.

10. Zarambad Curcuma zeodaria Li Rhizome 4 g.

11. Qaranful Syzygium aromaticum 4 g.

(L.) Merr. L M Perry.

12. Jauzbuwa Myristica fragrans Houtt. Endosperm 4 g.

13. Bisbasa Myristica fragrans Houtt. Aril 4 g.

14. Nabat Safaid Sugar Crystals 350 g.

15. Arq-e-Kibreet Sulphur extract Extract 600 ml.


Collection of the raw drugs

Genuine raw drugs namely Ood, Dana Heel Khurd, Dana Heel Kalan, Sazaj Hindi,

Post-e-Turanj, Tabasheer, Nana, Darchini, Gul-e-Surkh, Zarambad, Qaranful,

Jauzbuwa, Bisbasa, Nabat Safaid and Arq-e-Kibreet of the formulation were

procured from raw drugs dealers of Chennai. The raw drugs were identified using

pharmacognostical methods and evaluated their pharmacopoeial standards.


Preparation of the drug

The ploy herbal semisolid drug was prepared in different batches at Laboratory

scale as per the ingredients composition and guidelines of NFUM, PartIV

(Table I).
Powder microscopy
Microscopical examination allows more detail of a drug and can be used to identify

the organised drugs by their known histological characters viz., cell walls, cell

contents, starch grains, calcium oxalate crystals, trichomes, fibres and vessels

(Kokate et al., 2000).

The drug sample (5g) was weighed and mixed with 50ml of water in a beaker with

gentle warming, till the sample completely dispersed in water. The mixture was

centrifuged and decanted the supernatant. The sediment was washed several

times with distilled water, centrifuged again and decanted the supernatant. A few

mg of the sediment was taken and mounted in glycerine. Then few mg was taken

in watch glass and added few drops of phloroglucinol and concentrated

hydrochloric acid, mounted in glycerine. The microscopic salient features of the

drug were observed in different mounts (Wallis, 1997 & Johansen, 1940).

Physico-chemical analysis

The physico-chemical methods viz., moisture content, ash values, solubility in

different solvents, pH values, bulk density and sugar content etc., are useful tools

in standardisation of a herbal product for maintaining batch to batch consistency.

The drug samples were subjected for the standardisation of physico-chemical

parameters and analysed as per the standards method (Anonymous, 1987).

Thin layer chromatography

Thin Layer Chromatography is a physical method of separation in which the

components to be separated are distributed between two phases; one of these is

a stationary phase bed and other is a mobile phase which percolates through this

bed. It offers the best method for recording the finger prints which can be

reproduced anywhere at the same laboratory condition of a particular product.

The samples of the drug (2g) were soaked in chloroform and alcohol separately

for 18 hours, refluxed for ten minutes on water bath and filtered. The filtrates were

concentrated on water bath and made up to 5ml in a standard flask separately

and carried out the TLC studies (Wagner et al., 1984).


Quality control parameters

The usage of herbal products along with higher safety margins, WHO has taken

necessary step to ensure quality control parameters with the modern techniques

and application of suitable standards. The parameters such as microbial load

and heavy metal were carried out as per the WHO guidelines (Anonymous, 1998).

Aflatoxin and pesticide residues were carried out by standard methods
(Anonymous, 2000).


Jawarish-e-Ood Kibreet is a dark brown semi-solid product with sweetish bitter in

taste.

Microscopical observation

The salient features of the raw drugs used in the preparation of Jawarish-e-Ood

Kibreet were evaluated using powder microscopic studies and the photographs

are shown in Fig. 1. Pitted vessels with simple perforation plate upto 200, xylem

parenchyma lignified with pitted walls, xylem ray parenchyma cells along with

fibres, septate fibres (Ood); perisperm cells isolated or in groups with bulbous

projections filled with starch grains and tiny prismatic crystal of calcium oxalate,

elongated thin walled parenchyma cells from aril tissue, thick walled sclerenchyma

cells in surface view (Dana Heel Khurd / Dana Heel Kalan); epidermal cells in

surface view with sunken stomata (paracytic stomata) subsidiary cells not clear

(Sazaj Hindi); epidermal cells in surface view with circular stomata and

schizolysigenous oil glands (Post-e-Turanj); numerous, angular, sharp edged

siliceous sandy particles (Tabasheer); epidermal cells (bigger cells) in surface

view with wavy margin, diacytic stomata, prominent capitate glandular trichomes

upto 80 in length with single basal cell and single head cell, labiaceous glandular

trichomes with single basal cell and a head of 8 cells upto 80 in diameter (Nana

Khushk); fibres thick walled lignified with striated walls and narrow lumen of length

upto 1000 and breadth not over 30, stone cells with horse shoe shaped

thickenings upto 100 (Darchini); long simple unicellular covering trichome, pollen

grains round to oval upto 35 with three distinct germ pores (Gul-e-Surkh);

numerous starch grains of various shapes and size upto 50 (Zarambad); pollen

grains tetrahedral spherical biconvex measuring upto 20 in diameter, spindle

shaped fibres, parenchyma cells with schizolysigenous oil glands, fragments of

anther wall in surface view (Qaranfal); endosperm cells in surface view with

numerous starch grains and crystalloid proteins, each crystalloid proteins upto

40, perisperm cells in surface view filled with reddish brown content (Jouzbuwa);

thick walled elongated parenchyma cells in surface view upto 50 wide (Bisbasa).


Fig. 1: Powder microscopy of Jawarish-e-Ood Kibreet


Fig. 1: Continued...

Chemical analysis

The physico-chemical data, moisture content was obtained in the drug 18.58%.

The alcohol soluble extractive (58.50%) might be due to the extraction of polar

chemicals constituents and the water soluble extractives (63.49%) indicate the

presence of inorganic constituents. The obtained data are shown in Table - 2.

Thin Layer Chromatography analysis

The chloroform and alcohol extract of all the three batch samples showed identical

spots at UV 254nm and 366nm ranges and the Rf values of both the extracts


Table 2: Physico-chemical parameters

Parameters Analyzed Batch Number (n=3)

I II III

Extractives 58.52% 58.24% 58.76%

Alcohol soluble matter 63.16% 63.48% 63.84%

Water soluble matter

Ash 1.82% 1.79% 1.86%

Total ash 0.95% 0.89% 0.98%

Acid insoluble ash

pH values 5.59 5.74 5.41

1% Aqueous solution 4.34 4.41 4.42

10% Aqueous solution

Sugar estimation 35.83% 35.61% 35.84%

Reducing sugar 7.55% 7.74% 7.53%

Non-reducing sugar

Moisture 18.81% 18.26% 18.68%

Bulk Density 1.3997 1.4087 1.4009

Table 3: Rf Values of chloroform extract

Solvent system Rf Values

Toluene: Ethyl acetate (9 : 1)

UV 254nm UV 366nm V. S. Reagent

0.93 Pink 0.90 Light blue 0.94 Blue

0.71 Light pink 0.72 Brown 0.82 Red

0.64 Pink 0.55 Blue 0.77 Grey

0.56 Light pink 0.51 Red 0.67 Grey

0.52 Pink 0.42 Fluorescent blue 0.59 Brown

0.46 Pink 0.34 Red 0.50 Brown

0.32 Yellowish green 0.14 Blue 0.42 Grey

0.14 Light pink 0.32 Violet

0.22 Light grey

B1 B2 B3 0.19 Violet

Fig. 2 V. S. Reagent

are shown in Table 3 and 4. The plates were developed using vanillin-sulphuric

acid and heated at 105C till appeared coloured spots shown in Fig. 2 and 3.


Table 4: Rf Values of alcohol extract

Solvent system Rf Values

Toluene: Ethyl acetate (6 : 4)

UV 254nm UV 366nm V. S. Reagent

0.93 Yellowish green 0.93 Red 0.90 Brown

0.81 Pink 0.81 Blue 0.81 Brown

0.73 Light pink 0.78 Violet 0.69 Violet

0.60 Pink 0.56 Yellow 0.59 Green

0.57 Yellowish green 0.52 Blue 0.53 Light grey

0.47 Light pink 0.47 Brown 0.39 Blue

0.39 Light pink 0.40 Blue 0.26 Grey

0.27 Yellow 0.32 Fluorescent blue 0.14 Grey

0.15 Yellowish green 0.28 Red

B1 B2 B3 0.17 Red

Fig. 2 V. S. Reagent

Quality control parameters

The evaluated quality control parameters such as microbial load and heavy metals

were found within the permissible limit in the drug shown in Table 5 and 6. The

other parameters like aflatoxins B1, B2, G1 and G2 and pesticide residues - organo

chlorine group, organo phosphorus group, acephate, chlordane, dimethoate,

endosulphan, endosulfan, endosulfon, ethion, endosufon sulphate, fenthion,

heptachlor, lindane, methoxychlor, phorate sulfoxide and phorate sulfone were

not detected from the drug samples shown in Table 7 and 8.

Table 5: Analysis of Microbial load

S.No. Parameter Analyzed Results WHO Limits

1 Total Bacterial Count 2,000 CFU / gm 105 CFU / gm

2 Total Fungal Count < 10 CFU/ gm 103 CFU / gm

3 Enterobacteriaceae Absent / gm 103 CFU / gm

4 Salmonella Absent / gm Nil

5 Staphylococcus aureus Absent / gm Nil

Table 6: Estimation of Heavy Metals

S.No. Parameter Analyzed Results WHO & FDA Limits

1 Arsenic Not detected 3 ppm

2 Cadmium Not detected 0.3 ppm

3 Lead 0.0023 10 ppm

4 Mercury Not detected 1.0 ppm


Table 7: Estimation of Aflatoxins

S.No. Aflatoxins Results WHO Limits

1 B1 ND 0. 5ppb

2 B2 ND 0.1ppb

3 G1 ND 0. 5ppb

4 G2 ND 0.1ppb

ND = Not Detected

Table 8: Analysis of Pesticide Residues

S.No. Pesticide Residues Results Limits

1 Organo Chlorine group ND (DL 0.005mg/Kg)

2 Organo Phosphorus group ND (DL 0.005mg/Kg)

3 Acephate ND (DL 0.005mg/Kg)

4 Chlordane ND (DL 0.005mg/Kg)

5 Dimethoate ND (DL 0.005mg/Kg)

6 Endosulphan ND (DL 0.005mg/Kg)

7 Endosulfan ND (DL 0.005mg/Kg)

8 Endosulfon ND (DL 0.005mg/Kg)

9 Ethion ND (DL 0.005mg/Kg)

10 Endosufon sulphate ND (DL 0.005mg/Kg)

11 Fenthion ND (DL 0.005mg/Kg)

12 Heptachlor ND (DL 0.005mg/Kg)

13 Lindane ND (DL 0.005mg/Kg)

14 Methoxychlor ND (DL 0.005mg/Kg)

15 Phorate sulfoxide ND (DL 0.005mg/Kg)

16 Phorate sulfone ND (DL 0.005mg/Kg)

ND Not detected

Conclusion

The evaluated data such as powder microscopy, physico-chemical, TLC and quality

control shows that the genuine raw drugs were added in the formulation and

there is no variation in the batch to batch consistency of the drug.


Acknowledgement

The authors are thankful to the Director General, CCRUM, New Delhi, for his

valuable guidance, encouragement and providing necessary research facilities

to carry out the studies.
Reference
Anonymous, 1987. Physico-chemical standards of Unani Formulations, Part II.

CCRUM, Min. of Health & Family Welfare, New Delhi, pp. 300 - 317.

Anonymous, 1998. Quality Control Methods for Medicinal Plant Materials. World

Health Organization, Geneva, pp. 25 - 28.

Anonymous, 2000. Association of Official Analytical Chemists (AOAC), 17th Edition.

Anonymous, 2006. National Formulary of Unani Medicine, Part IV. Min. of Health

& Family Welfare, New Delhi, pp. 57 58.

Johansen, D.A., 1940. Plant Microtechnique. Mc. Graw Hill Book Company Inc.,

New York and London, pp. 181-186.

Kokate, C.K., Purohit, A.P. and Gokhale, S.B., 2000. Pharmacognosy, Nirali

Prakashans, Pune, pp. 98 99.

Wagner, H., Bladt, S. and E.M., Zgainski, 1984. Plant Drug Analysis, A Thin Layer

Chromatography Atlas (2nd Edition). Springer-Verlag, Germany.

Wallis, R.E., 1997. Text Book of Pharmacognosy, 5th Edition. CBS Publishers &

Distributors, Delhi, pp. 494 496.

Bandaranayake, W.M., 2006. Quality control, screening, toxicity and regulation

of herbal drugs. Modern Phytomedicines, pp. 25 - 57.

Myers, S.P., Cheras, P.A., 2004. The other side of the coin: safety of complementary

and alternative medicine. Medical J. Australia (181): 222 - 225.


Standardization Abstract

of Unani Drug he peel of fruit or fruit rind of Punica granatum L. is used as

Post-e-Anar
(Fruit rind of
Punica
T

herbal drug. In Unani medicine it has been used to cure many ailments and known
as Post-e-Anar. It is anti-oxidant, anti-cancerous, anti inflammatory and anti-
allergic. But its standardization has not been done so far, so keeping in mind the
medicinal properties of Post-e-Anar, its diagnostic characters has been identified
by following the modern scientific quality control procedures. The study provides
granatum L.)

organoleptic, physico-chemical and thin layer chromatographic profile which will

be helpful in laying down the standards and pharmacopoeial parameters. The

*Nazish Siddiqui,
qualitative screening for phytochemicals was carried out to facilitate desirable

M. Masihuzzaman Ansari,

therapeutic efficacy. Besides this, novel Infra-red (IR) spectral study of rind extract

Alvia Khan,

has also been conducted by recording the IR spectrum of the extract, which can

Mohd. Bilal Tafseer,

Saba Viquar be used as fingerprints for identity of drug Post-e-Anar.

and

Abdul Haleem Keywords: Standardization , Post-e-Anar, Physico-chemical, IR Spectrum, Fruit

rind, Phytochemicals.

Department of Ilmul Advia,

A.K. Tibbiya College,


Introduction

Aligarh 202002

The plant Punica granatum L. is a large deciduous shrub or a small tree

(Anonymous, 2009). It belongs to the family Punicaceae. It is commonly known

as Anar and considered to be native of Iran, Afghanistan and Baluchistan. It is

also found growing wild in the Himalayas between 900 to 1800 m and also

cultivated throughout India. Flowers occur during April to July, while fruiting takes

place during July to September (Anonymous, 2007). Various parts of plants are

used as drug. Fruit juice is excellent for curing diarrhoea, dysentary and other

ailments such as colitis, anaemia, jaundice, high blood pressure, piles and arthritis

(Anonymous, 2009; Dipak et al., 2012), decreases cholesterol absorption ( Julie

Jurenka, 2004). The leaf, fresh seeds and root bark of Punica granatum have

been studied and standardized. (Anonymous, 2009; Anonymous, 2007;

Anonymous, 1997). The external leathery cover of fruit of Punica granatum L. is

known as rind and in Unani medicine it is known as Post-e-Anar. The rind or peel

of fruit contains allergic acid as one of the main constituents which interrupts free

radicals induced damage and acts as anti-oxidant (Yunfeng et al., 2006; Amir et

al., 2011 ). It also acts as anti-cancerous inhibiting the growth of tumours and is

useful in cancer of colon, breast, prostate, skin and pancreas (Julie Jurenka,

2004; Amir et al., 2011; Negi et al., 2011). Besides this rind extract has been

found to possess anti-allergic and anti-inflammatory activities also

(Panichayupakaranant et al., 2010). The world is heavily polluted with bacteria,



viruses and fungi infections and these are the major cause of diseases in human

beings. Further, due to indiscriminate use of synthetic anti-microbial drugs, most

infective microorganisms have developed resistance to many antibiotics. Thus,

there is an urgent need to discover novel herbal anti-microbials, as they have

less or no side effects. The fruit rind extract of pomegranate also shows significant

antibacterial and antifungal activities (Panichayupakaranant et al., 2010; Negi et

al., 2003; Adollahzaden et al., 2011; Dahham et al., 2010), Apart from this, Post-
e-Anar extract was found to exhibit antimalarial activity (Agli et al., 2010), It is also

useful for prevention of cardio-vascular diseases, diabetes, alzheimer,s disease,

arthritis, obesity, dental problem (Mohammad & Kashani, 2012; Negi et al., 2003)

and possesses antidiarrhoeal activity (Akter et al., 2013) . Thus, keeping in mind

the medicinal importance of drug, various physico-chemical and phytochemical

studies on the fruit rind of Punica granatum (Post-e-Anar) were carried out with a

view to standardize it and lay down standards for its purity, quality control & quality

assurance, which has not been taken earlier.

Methodology

Collection of Plant Material

Red pomegranate fruits were collected . Longitudinal cuts were given to the peel

of the fruit and peels were gently removed with the help of hand and knife. The

seeds were removed and peel was cut into pieces of 1 inch size. It was then

placed in a sunny spot during day light hours every day till the peel gets hard and

loses all the moisture. Dried peels were powdered to get 60-mesh size using

mixer grinder and placed in air tight container as Post-e-Anar for study.

(i) Organoleptic Parameters: The colour, taste, odour, solubility were noted

which provide first hand information.

(ii) Physico-chemical Parameters: Various physico-chemical studies like total

ash, acid insoluble ash, water soluble ash, successive extractive values

using soxhlet extraction method, water and alcohol soluble matter, pH,

bulk density, moisture content were carried out as per guidelines of WHO

and others (Anonymous, 1998; Anonymous, 1963),

(iii) Qualitative Analysis of Post-e-Anar: was conducted to identify the organic

chemical constituents present in the drug (Overtone, 1963).

(iv) IR Spectroscopic Study: For this alcoholic extract of the drug was obtained

by refluxing powdered drug (5.0 g) with absolute alcohol (50 ml) for 5

hrs and removing the solvent under reduced pressure. The IR spectrum

of alcoholic extract was determined in KBr pellets with Perkin Elmer

1600 FTIR spectrometer.


(v) Thin layer chromatographic analysis: For this ethyl acetate and ethanolic

extract was conducted20,21 in percolated silica gel 60F254 TLC plates.

The plates were visualized in day light and UV short wavelength

respectively (Harborne, 1973; Stahl, 1969).

Observations and Results

Organoleptic characters: The powder of the fruit rind of pomegranate was brownish

yellow with characteristic odour and taste, showed good solubility in water (Table 1).

Physico-chemical constants: Different physico-chemical constants were

determined three times and their average values are depicted (Table 2).

Table 1 : Organoleptic characters of fruit rind of Punica granatum L.

S.No. Parameters Observations

1 Colour Brownish yellow

2 Odour Characteristic

3 Taste Characteristic

4 Solubility Good in water

Table 2 : Physico-chemical Constants of fruit rind of Punica granatum L.

S.No. Parameters Analytical values (%)*

1 Ash Value

Total Ash (w/w) 4.21

Acid Insoluble Ash (w/w) 2.21

Water Soluble Ash (w/w) 2.9

2 Solubility

Alcoholic Soluble Matter (w/w) 7.0

Water Soluble Matter (w/w) 8.8

3 Successive Extractives

Petroleum ether 0.53

Diethyl ether 5.0

Chloroform 0.49

Ethyl acetate 1.73

Absolute alcohol 32.62

Distilled water 25.88

4 pH Values

1% Aqueous solution 3.73

10% Aqueous solution 3.40

5 Moisture content (v/w) 12

6 Bulk density 0.49

*Values are average of three experiments


Qualitative analysis of Post-e-Anar: The phytochemicals present in the drug were

qualitatively analysed by different chemical tests and results are given (Table 3).

IR spectral study of the drug: Novel IR spectral study of the alcoholic extract of

the drug was done by running the alcoholic extract in the IR range (4000-667cm-1)

of the electro-magnetic spectra and major characteristic peaks were noted

(Fig. 1, Table 4).

Thin layer chromatographic profile: Thin layer chromatographic analysis of the

ethanolic and ethyl acetate extract of Post-e-Anar was carried out using Benzene

: methanol : acetic acid (45 : 8 : 4) and ethyl acetate : acetic acid (20:4) respectively

as solvent system. Rf values were calculated after the development of

chromatogram. The Rf values in the given solvent are used to characterize the

drug,s identity and purity. The results obtained are given in table-5.

Table 3: Qualitative Analysis of Phytochemicals of fruit rind of Punica

granatum L.

S.No. Chemical Constituents Test / Reagents Inference

1. Alkaloids Dragendorff,s reagent +ve

Wagner,s reagent +ve

Mayer,s reagent +ve

2. Carbohydrates Molish Test +ve

Fehling Test +ve

Benedicts Test +ve

3. Flavonoids Mg ribbon and dil Hcl +ve

4. Glycosides NaOH Test +ve

5. Tannins / Phenols Ferric chloride Test +ve

6. Protein Xanthoproteic Test +ve

Biuret Test +ve

7. Steroids / Terpenes Salkowski Reaction +ve

8. Amino acids Ninhydrin Solution -ve

Table 4: IR Spectral Details of alcoholic extract of fruit rind of Punica granatum L.

Region of electromagnetic spectra Characteristic peaks

IR, (cm-1) 3412, 3329, 1706, 1615, 1512, 1448,

1345, 1222, 1190, 1048



99
Figure 1
Table 5 : TLC Profile of alcoholic extract of fruit rind of Punica granatum L.

Extract Solvent Visualizing No. of Rf Value

System Agent Spots

Ethyl Ethyl Acetate : Day Light 3 0.41, 0.65, 0.76

Acetate Acetic Acid

(20:4)
Ethanol Benzene : UV Short wave 3 0.47, 0.59, 0.88

Methanol :

Acetic Acid

(45:8:4)

Discussion

The interest in evaluating therapeutic effects of plants have increased as 80% of

worlds population depends on complementary and alternative medicine for their

health care needs (Magee, 2005; Duraipandiyan, 2006). The quality of herbal

drugs is sum of all the factors which contribute directly or indirectly to its safety,

effectiveness and acceptability of the product. Standardization of drugs means

confirmation of its identity, determination of its quality and purity (Ekka, 2008).

Initially organoleptic studies were used to authenticate the drugs but some drugs

require other physical and chemical studies too. So standardization of Post-e-

Anar was carried out as it has an important role in folk medicine. The present

standardization study has brought out many diagnostic characters of herbal drug

Post-e-Anar (fruit rind of Punica granatum). Organoleptic characters provide

preliminary information regarding colour, taste, odour and solubility which is

necessary for authentification. Physico-chemical parameters were determined to

ascertain the identity and purity. Ash values were found as total ash 4.21%, acid

insoluble ash 2.21% and water soluble ash 2.9%, if the drug is adulterated with

siliceous or earthy matter ash values may change. Moisture content has been

determined by toluene distillation method as 12%. If its value increases that may

provide an evidence that the drug is not dried and stored properly. The pH value

of 1% and 10% aq. solution was 3.73 and 3.40 respectively, which helps in knowing

the drug receptor site interactions and also stability and therapeutic activity of the

drug. The amount of extract that a drug yields to a particular solvent is an

approximate measure of the amount of certain constituents or group of related

constituents that the drug contains (Jenkins et al., 1967). Thus extractive values

of Post-e-Anar in six organic solvents were calculated as an index of its purity,

and these are shown in Table 2. Phytochemical screening of drug was done to

know the chemical constituents present in Post-e-Anar (Table 3).


Another important technique taken here to confirm the identity was Infra red (IR)

spectroscopy. The IR spectrum of alcoholic extract of fruit rind of Punica granatum

(as Post-e-Anar) was recorded in fig-1, table-4. Initially the IR spectroscopic method

was restricted only for structural elucidation of isolated compounds from the herbal

matrix but now it is also useful in phytochemical studies as a fingerprinting device

for comparing a natural sample with synthetic sample. In IR spectrum, region

1430-910 cm-1 is known as finger print region, which can be compared with the
finger prints of human beings, which differs from person to person. In the same

manner this region would differ from drug to drug. In addition to this IR spectrum

also provides information about functional groups present in the chemical

constituents of the drug. To check the purity of drug IR spectrum may be compared

with the authentic sample. If the characteristic bands are similar and identical, the

test drug would be genuine. Apart from this thin layer chromatographic (TLC)

study was also carried out. Now a days other instrumental chromatographic

methods are also used like GC & HPLC but TLC provide first characteristic

fingerprints of herbs and various pharmacopoeias still use TLC (Jain and Sharma,

2005). TLC was made here to construct the fingerprint of Post-e-Anar, as it is

simple, versatile, sensitive and economic. Results are summarized in table-5.

This study assumes great significance as it will provide a key of diagnostic

characters which will serve as an important tool in laying down the standards for

quality assurance.

Acknowledgement

The authors are grateful to the Chairman, Department of Ilmul Advia, AMU, Aligarh,

for providing necessary research facilities. Facilities provided by SAP (DRS-1) by

way of generous research support are duly acknowledged.

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104
A Study on Abstract

Diversity of series of medicinal plants collection trips to various forest

Unani Medicinal
Plants Used for
Non-
A

regions of South Western Ghats of Tamil Nadu, have brought to light 53 plants
species widely used in the study area to treat certain non-communicable diseases
as per Unani text. All species collected have been analyzed in respect of their
diversity, life form and taxonomic category. The study reveals that many such
species are either rare or endangered and needs immediate attention for their
Communicable

conservation and protection, through germiplasm collection initiatives before these

Diseases in are lost for ever. All species have been listed providing information on their botanical

name, family, Unani name, therapeutic uses as per Unani text. Further scientific

Southern

studies are suggested with a view to discover new drugs of plant origin to combat

Western Ghats

many such non-communicable diseases, hitherto incurable, in modern medicine.

of Tamil Nadu

Keywords: Medicinal Plants, Western Ghats, Non-Communicable disease, Unani

System of Medicine

1*R. Murugeswaran,

1K. Venkatesan,

1Aijaz Ahmed

Introduction

and

2Aminuddin

The greatest public health challenge of 21st century is chronic non-communicable

1Regional
diseases (Anderson and Chu, 2007). Approximately 36 million deaths were

Research Institute

of Unani Medicine, attributable to NCDs in the year 2008 due to diabetes and cardiovascular diseases

alone (Alwan et al., 2011). India is endowed with rich wealth of medicinal plants

1, West Mada Church Road,

Royapuram, Chennai-600013
which are widely used by all section of peoples either directly or as folk remedies

in different indigenous systems of medicine or indirectly in the pharmaceutical

2CentralCouncil for Research

preparations of modern medicines (Alagesaboopathy, 2011). Medicinal plants are

in Unani Medicine,

61-65, Institutional Area,

distributed across diverse habitats and land scape elements. Around 70% of Indias

Janakpuri, New Delhi - 110058

medicinal plants are found in to tropical areas mostly in the four types spread

across the Western Ghats and Eastern Ghats, Vindiyas, Chotta Nagpur Plateau,

Aravalis & Himalayas. (Sankar Murthy and Kiran, 2012).

India is experiencing great pressure on its resources due to growing demand of

the herbal products in the domestic and global markets. During the past two

decades the human activities on commercialization of plant based drugs and

demand from the pharmaceutical industry for domestic needs and the export of

herbal drugs have led scarcity of medicinal plants in forests and plains. The natural

forests should be conserved seriously to protect many of the economically

important medicinal plants. The utility and need of botanical exploration in the

country is to identify and search the economically important medicinal plants which

has to be propagated and conserved for future generation (Balakrishnan et al.,



2009). In the present study, a large number of medicinal plants were collected

and identified from the southern region in recent years.

The Survey of Medicinal Plants Unit of the Regional Research Institute of Unani

Medicine, Chennai has been extensively involved in medicinal plants survey and

collection trips in different parts of Tamil Nadu for over three decades. The present

report provides information on some 53 important medicinal species as per Unani

text and widely used for treating non-communicable and other diseases in the

study area. Study also analysed the diversity of Unani medicinal plants flora in

respect of their diversity and the predominant families for the study area.

The Study Area

Southern Western Ghats of Tamil Nadu are one of the important medicinal plants

biodiversity hotspots in the country and spread over in the districts of Coimbatore,

Nilgiris, Theni, Tirunelveli and Kannyakumari. The vegetation of this area comprise

evergreen, deciduous, scrub jungles and shola forests. Many of the forest areas

are rich in tribal populations. The Western Ghats mountain range runs parallel to

the West coast of Peninsular India for about 1600km. It has very rich floristic

diversity; about 4000 indigenous angiosperm species which includes 1500

endemic plant species.

Methodology

A series of medicinal plants surveys were conducted in different seasons at

Southern Western Ghats forest region of Tamil Nadu. 53 species of wild medicinal

plants species which are used in the Unani system of medicine are reported in

the present paper with their therapeutic uses. The medicinal species have been

identified through modern floras (Gamble, 1928; Kirtikar & Basu, 1933; Nair &

Henry, 1983; Mathew, 1983) and confirmed at Botanical survey of India,

Coimbatore. The plants are arranged alphabetically according to their botanical

names with collection number, followed by family, Unani name, and their uses in

the Unani system of medicine (Table-I).


In the present study 53 Unani medicinal plants used for non-communicable disease

as per classical text have been collected (Fig. 4-9) from Western Ghats region of

Tamil Nadu (Table-1) and analyzed for their diversity status. Of these, 32 species

are common, 13 falls in sporadic distribution which is very much restricted to

particular area, 8 vulnerable, 5 rare and 1 species each is in endangered category,

respectively (Fig.1). In the life form analysis 21 species of herbs, 18 species of


trees, 9 species of climbers, twiners and creepers and 5 species of shrubs were

collected and identified (Fig:2).

According to the systematic classification, the taxonomic hierarchy has also been

analyzed in which 53 species, belonging to 51 genus and 32 family are recorded

for the study area (Fig: 3). The study also revealed that largest families of the

study area are Apocynaceae, Caesalpiniaceae and Euphorbiaceae with 4 species

each, followed by Amaranthaceae, Solanaceae with each 3 species, similarly

Asclepiadeceae, Combretaceae, Fabaceae, Liliaceae, Loganiaceae, Malvaceae

and Rubiaceae possess 2 species each and other families possess 1 species

each respectively.

Fig. 1: Analysis of diversity of Unani medicinal plants with respect to no. of species

in the study area

Fig. 2: Analysis of Unani medicinal plants life forms with respect to no. of species

in the study


Fig. 3: Analysis of taxonomical ranks of Unani medicinal plants with respect to

number of species in the study

The medicinal plants are used in Unani system of medicine for various ailments

like anthelmintic, arthritis, astringent, aphrodisiac, bleeding hemorrhoids,

diarrhoea, dysentery, gastric ulcer, headache, inflammation and stomach disorders

are among them. Some of the important plant species used are Amaltas (Cassia

fistula L.), Amla (Phyllanthus embilica L.), Bijasar (Pterocarpus marsupium L.),

Bhuiamla (Phyllanthus amarus L.), Chalmugrah (Hidnocarpus laurifolia (Dennst.)

Sleumer.,Chungchi (Abrus precatorius L.), Dudhi (Euphorabia hirta L.), Gilo

(Tinospora cordifolia (Willd.) Miers), Gul-e-abbas (Mirabilis jalapa L.), Gul-e-dahwa

(Anogeissus latifolius (Roxb.ex DC.) Bedd.), Halela (Terminalia chebula L.), Inderjo

shirin (Wrightia tinctorea R.Br.), Kamila (Mallotus philippensis (Lam.) Mull.-Arg.),

Kasoos (Cuscuta reflexa Roxb.).Katai (Solanum virginianum L.), Lajwanti (Mimosa

pudica L.), Mainphal (Catunaregam spinosa (Thumb.) Tirveng., Marorphali

(Helicteres isora L.) Muleem (Gloriosa superba L.), Neem (Azadirachta indica

(L.) A. Juss), Panwar (Cassia tora L.), Patharphodi (Aerva lanata (L.) Jes, Qil qil

(Cardiospermum halicacabum L.)Sadabhagar (Vinca rosea L.),Sambhalu (Vitex

negundo L.), Santhal safaid (Santalum album L.), Sheetraj Hindi (Plumbago

zeylanica L.), Talmakhana (Hygrophila auriculata (Schum.) Heine, Tarwar (Cassisa

auriculata L.), Zarawand (Aristolochia indica L.) etc., were exemplified and have

been discussed for their diversity status.

It has been observed that the plant species like Asgand (Withania somnifera Dunal),

Chalmugrah (Hidnocarpus laurifolia (Dennst.) Sleumer.) Darchini (Cinnamomum

verum Presl.), Ghungchi (Abrus precatorius L.), Kashim (Alstonia scholaris R.Br.),

Gul-e-Dhawa (Anogeissus latifolius (Roxb. ex DC.) Wall. ex Guill. & Perr.), Chironji


Table 1: List of Unani Medicinal Plants Used for Non-Communicable Diseases

in Southern Western Ghats of Tamil Nadu, India

Sl. Botanical Name Family Unani Uses in the Unani Sta-

No. & Collection No. Name System of Medicine tus

1 Abrus precatorius L. Fabaceae Ghungchi Seeds are used as anti- V

SMPU, CH- 8611 inflammatory, and stimulant.

Used in nerve disorders

and ulcer.

2 Abutilon indicum (L.) Malvaceae Kanghi Leaves are used in bleeding C

Sweet piles, constipation and used

SMPU, CH-8511 as aphrodisiac and

analgesic.

3 Achyranthes aspera L. Amaran- Chirchita Whole plant used in dropsy. C

SMPU, CH 8491 thaceae

4 Aerva lanata (L.) Amaran- Pathar- Whole plant used as C

A. Juss. Ex Schult. thaceae phodi diuretic and demulcent.

SMPU, CH-8455

5 Alstonia scholaris Apocyna- Kashim Root used as carminative, S

R.Br. SMPU, ceae digestive, stomachic, used

CH-8590 to expel intestinal worms,

dropsy and hemiplegia.

6 Amaranthus spinosus Amaran- Kateeli Root used as appetizer, C

L. - SMPU, CH-8572 thaceae chauli carminative and anti-

inflammatory.

7 Anogeissus latifolius Combreta- Gul-e- Flowers are used in . S

(Roxb. ex DC.) ceae Dhawa constipation, diarrhea,

Wall. ex Guill. & Perr. leucorrhoea, and intestinal

SMPU, CH-8474 worms

8 Argemone mexicana Papavara- Satyanasi Plant used as diuretic. Juice C

L. SMPU, CH-8561 ceae of shoot used as purgative,

and used as blood purifier.

Seeds used in constipation,

also used as antiseptic. Oil

of seeds used as purgative

and root used as tonic.

9 Aristolochia indica L. Aristolo- Zarawand Seeds and roots are used in R

SMPU, CH-8689 chiaceae general debility, amenorrhea

and wounds, Used as

antidote for poison.

10 Asparagus recemosus Liliaceae Satawar Root powder used as R

Willd. - SMPU, diuretic, aphrodisiac,

CH-8682 antidiarrheal, nervine tonic

and used in rheumatic

complaints.


Table 1: (Continued)



11 Buchanania lanzan Anacardia- Chironji Kernel used as aphrodisiac S

Spreng. - SMPU, ceae and in sexual debility and

CH-8542 body weakness.

12 Calotropis gigantea Asclepia- Madar Root bark used as C

(L.) R.Br. ex Ait. deceae analgesic, digestive, anti-

SMPU, CH-8592 dysenteric, expectorant,

arthritis. Used in asthma,

general debility, Flowers

used as analgesic,

stomachic and expectorant.

Latex used in piles.

13 Cardiospermum Sapinda- Qil Qil Seeds used as aphrodisiac. C

halicacabum L. ceae

SMPU, CH-8444

14 Cassia auriculata L. Caesal- Tarwar Leaves, flowers and roots C

SMPU, CH-8565 piniaceae are used in diabeties,

diarrhea and cardiac tonic.

15 Cassia fistula L. - Caesal- Amaltas Fruit pulp, rind of the fruit C

SMPU, CH-8454 piniaceae and leaves are used in

constipation, asthma,

rheumatism.

16 Cassia tora L. Caesal- Panwar Seeds used as blood C

SMPU, CH-8589 piniaceae purifier. Used in asthma.

17 Catharanthus roseus Apocyna- Sadabahar Leaves and roots are used C

(L.) G. Don. SMPU, ceae in haemorrhage and

CH-8496 constipation.

18 Catunaregam spinosa Rubiaceae Mainphal Fruits used as purgative, C

(Thumb.) Tiruveng. carminative and inflamma-

SMPU, CH-8494 tion. Used in paralysis.

19 Cinnamomum verum Lauraceae Darchini Bark and oil are used as V

Presl. SMPU, liver tonic, carminative,

CH-8802 expectorant, used in piles

and inflammations.

20 Citrus limon (L.) Rutaceae Neembu Fruit used in brain tonic, C

Burm.f. SMPU, constipation, cardiac tonic,

CH-8527 exhilarant, carminative and

digestive.

21 Curcuma longa L. Zingibera- Zard Chob Rhizome used as anti- C/C

SMPU, CH-8609 ceae Haldi inflammatory, expectorant,

used in ulcers, asthma.





22 Cuscuta reflexa Roxb. Convol- Kasoos Seeds used as purgative, C

SMPU, CH-8451 vulaceae carminative, diuretic, anti-

inflammatory, blood purifier.

Used in brain diseases,

epilepsy, paralysis, arthritis.

23 Datura metel L. Solanaceae Jaoz Masil, Leaves used as analgesic, C

SMPU, CH-8570 Dhatura and anesthetic, used in

joint pain.

24 Euphorbia hirta L. Euphorbia- Dudhi Whole plant used as C

SMPU, CH 8524 ceae demulcent, constipation,

expectorant, and

aphrodisiac.

25 Gloriosa superba L. Liliaceae Muleem Root used as astringent, S

SMPU, CH-8622 expectorant. Used in

bleeding piles.

26 Gymnema sylvestre Asclepia- Gurmarbuti Leaves used in polyuria, R

R. Br. SMPU, daceae heart diseases and

CH-8529 diabetes.

27 Helicteres isora L. Sterculia- Marorphali Root and bark are used in C

SMPU, CH-8493 ceae stomach ache.

28 Hibiscus rosa- Malvaceae Gudhal Flowers, leaves, seeds and C

sinensis L. SMPU, roots are used as exhilarant,

CH- 8442 cardio tonic, demulcent and

aphrodisiac. Used in

dysuria, palpitation,

weakness of spleen.

29 Hydnocarpus laurifolia Flacourtia- Chalmu- Seeds, oil used in wounds. V

(Dennst.) Sleumer ceae grah

SMPU, CH-8791

30 Hygrophylla auriculata Acantha- Talma- Seeds, roots are used as V

(Schum.) Heine ceae khana sedative, diuretic, arthritis,

SMPU, CH- 8568 gonorrhea, renal calculus,

and aphrodisiac.

31 Leonotis nepetiifolia Lamiaceae Dipmal Flowering heads are used S

(L.) R.Br. - SMPU, as laxative, emmenagogue

CH-8668

32 Limonia acidissima L. Rutaceae Kaith Fruit and leaves are used V

SMPU, CH-8564 as cardio tonic, liver tonic,

refrigerant, astringent,

strengthening the gum and

diuretic.





33 Mallotus phillipensis Euphorbia- Kamila Tree, fruit capsule, red- C

(Lam.) Muell.-Arg. ceae glandular, common; fruits

SMPU, CH- 8441 are used as antiseptic and

in otorrhoea.
34 Mimosa pudica L. Mimosa- Lajwanti Leaves used as analgesic, C

SMPU, CH-8456 ceae and in intestinal worms.

35 Mimosops elangi L. Sapotaceae Mulsari Root used as aphrodisiac, R

SMPU, CH-8651 diuretic and astringent.

36 Mirabilis jalapa L. Nyctangi- Gul-e- Leaves used as anti- C

SMPU, CH-8458 naceae Abbas inflammation, in dropsy and

jaundice. Flowers used in

piles. Root used in joint

pain, and sexual debility.

Seeds used in leucorrhoea.

37 Momodica charantia Cucurbita- Karaila Fruit used as nervine tonic, C/C

L. SMPU, CH-8552 ceae aphrodisiac. Used in joint

pain, gout, dropsy, intestinal

worms Leaves used as

anti-bilious.

38 Nerium oleander L. Apocyna- Kaner Root used as aphrodisiac C

SMPU, CH-8539 ceae and blood purifier. Used in

sexual debility. Leaves used

as blood purifier. Used in

paralysis.

39 Phyllanthus emblica L. Euphorbia- Amla Fruits used in weakness of C/C

SMPU, CH-8873 ceae brain, amnesia, head ache,

gastric ulcer, and diarrhoea.

40 Piper nigrum L. Piperaceae Filfil Siyah Fruits used as expectorant, C/C

SMPU, CH-8452 stomach tonic, aphrodisiac,

appetizer, carminative.

Root used as digestive,

carminative. Used in

weakness of stomach.

41 Plumbago zeylanica L. Plumbagi- Sheetraj Roots and leaves are used S

SMPU, CH-8457 naceae Hindi in arthritis, and facial

paralysis.

42 Pterocarpus Fabaceae Bijasar Gum used as anthelmintic V

marsupium. Roxb. - and opthalmia.

SMPU, CH-8755





43 Ricinus communis L. Euphorbia- Arand, Seeds used as anti- C

SMPU, CH-8563 ceae Bedanjeer inflammatory, purgative.

Leaves used as purgative.

Oil of seed used as

purgative and anti-

inflammatory. Used in colic,

constipation, worm

infestations and joint pain.

44 Rubia cordifolia L. Rubiaceae Majeed Roots used in retention of C

SMPU, CH-8447 urine, bleeding hemorrhoids

and emmenagogue.

45 Santalum album L. Santala- Sandal Wood used as exhilarant, E

SMPU, CH-8652 ceae Safaid antiseptic, expectorant,

cardiac tonic and astringent.

Oil of the wood used in

gonorrhea.

46 Solanum virginianum Solana- Katai khurd Fruits and roots are used C

L. SMPU, CH-8790 ceae as laxative, inflammation

and anthelmintic.

47 Strychnos nux-vomica Logania- Kuchla Fruits are used as tonic, V

L. SMPU, CH-8566 ceae aphrodisiac, diuretic and

emmenagogue.

48 Strychnos potatorum Logania- Nirmali Seeds used as astringent, V

L. SMPU, CH-8532 ceae aphrodisiac and diuretic.

49 Tamarindus indica L. Caesal- Tamar Fruits used as cardiac tonic C/C

SMPU, CH-8445 piniaceae Hindi, Imli and sedative. Used in

stomatitis. Kernal of seeds

used in sexual disorders.

Stem bark used in anorexia.

50 Terminalia chebula Combreta- Halela Fruit used in constipation, S

(Gaertn.) Retz. ceae expectorant, brain tonic and

SMPU, CH-8453 appetizer. Used in diarrhea,

asthma and headache.

51 Tinospora cordifolia Menisper- Gilo Stem used as blood purifier, C

(Willd.) Miers ex maceae anti-inflammatory, diuretic

Hook.f. & Thoms. - and constipation. Used in

SMPU, CH-8602 joint pain.

52 Wrightia tinctoria L. Apocyna- Inderjo Bark and seeds used as C

SMPU, CH-8465 ceae shirin aphrodisiac.





53 Withania somnifera Solanaceae Asgand Root used as aphrodisiac, R

Dunal SMPU, diuretic, rheumatism, ulcer;

CH-8427 leaves used for painful

swelling sore eye; seed

used for diuretic and

coagulating milk.

C-Common, C/C- Common & Cultivated, S-Sporadic, V-Vulnerable, R- Rare, E- Endangered

Unani Medicinal Plants used in non-communicable diseases in the study area

Fig. 4: Bijasar (Pterocarpus marsupium Fig. 5: Sheetraj Hindi (Plumbago

Roxb.) zeylanica L.)

Fig. 6: Kataikhurd (Solanum Fig. 7: Halela (Terminalia chebula

virginianum L. (Gaertn.) Retz.)

Fig. 8: Ghunghchi (Abrus precatorius L.) Fig. 9: Kuchla (Strychnonux-vomica L.)


(Buchanania lanzan Spreng.), Muleem (Gloriosa superba L.), Gurmarbuti

(Gymnema sylvestre R. Br.), Dipmal (Leonotis nepetiifolia (L.) R.Br), Kaith (Limonia

acidissima L.), Mulsari (Mimusops elangi L.), Talmakhana (Hygrophylla auriculata

(Schum.) Heine), Satawar (Asparagus racemosus Willd.) Sandal Safaid (Santalum

album L.) Kuchla (Strychnos nux-vomica L),. Kuchla (Strychnos potatorum L.)

and Zarawand (Aristolochia indica L.), are very much restricted in their distribution

and such plant species are recommended for cultivation and propagation.
The medicinal plants which are available in the natural sources need to be

conserved and propagated seriously because many species are under threat in

different bio-geographic regions of the country due to various external factors

and many of the valuable medicinal plants species are under threat to become

rare, endangered and some are on the verge of extinction. To avoid the depletion

of such valuable medicinal plants, the only alternative way is to develop many

herbal gardens in the suitable areas for large scale cultivation of important

medicinal plants through modern agronomical techniques.

Acknowledgement

The authors thank the Director General, CCRUM, New Delhi, for providing

necessary facilities, encouragement and financial sanctions to conduct this study.

We also thank the District Forest Officers, for permission to conduct the study

and concerned Forest Range Officers and staff of the study area for providing

necessary field assistance during the study.

References

Alagesaboopathy, C., 2011. Ethno-medicinal plants used as medicine by the

Kurumba tribals in Pennagaram Region, Dharmapuri district of Tamil Nadu,

India. Asian Journal of Exp. Biology Science 2 (1):140-142.

Alwan, A. Maclen D.R., Riley, L.M., dEspaignet, E., Mathers, C.D., Stevens, G.A.

et al., 2010. Monitoring and surveillance of chronic non-communicable

diseases: progress and capacity in high-burden countries. Lancet 376(9755):

1861-8

Anderson, G.F., Chu, E., 2007. Expending priorities: Confronting Chronic Diseases

in countries with low income. N. Engl. J. Med. 356(3): 309-11.

Balakrishnan, V., Prema, P., Ravindran, K.C., Philip Robinson, 2009. Ethno-

botanical Studies among Villagers from Dharapuram Taluk, Tamil Nadu, India.

Global Journal of Pharmacology 3 (1): 08-14.

Gamble, J.S. & C.E.C., Fisher, 1928. Flora of the Presidency of Madras. Adlard &

Son Ltd.,


Kirtikar, K.R. & Basu, B.D., 1933. Indian Medicinal Plants, Vol.1-4. Allahabad,

India.

Mathew, K.M., 1983. Flora of Tamil Nadu Carnatic, Vol.1-3. Botanical survey of

India, Howrah.

Nair, N.C., Henry, 1983. Flora of Tamil Nadu, Vol. 1-3. Botanical Survey of India,

Coimbatore.
Sankar Murthy, K. and Kiran, B.R., 2012. Medicinal plants used as anti-diabetic

drug in pharmaceutical industry and their conservation; an overview.

International Research Journal of Pharmacy 3(10): 65-71.


Pharmacog- Abstract

nostical lthaea officinalis Linn. is equated as botanical source of

Standardization
of Flowers of
Althaea
A

Unani drug Gul Khatmi. This plant species is attributed for various medicinal
properties and its every morphological part yields mucilage, asparagin, betaine,
starch and sugar. The present study is carried out on the pharmacognostical
standardization of Althaea officinalis Linn. (Flower) including Thin- Layer
Chromatographic profile which leads to develop quality standards for the drug.
officinalis Linn.

(Gul Khatami) Keywords: Althaea officinalis Linn., Pharmacognosy, Drug standardization.

1*Nitin Rai, Introduction

2Rampratap Meena,

Althaea officinalis Linn. (Family - Malvaceae) in trade is known as Gul Khatmi or

3S. Mageswari,

4Shamsul Arfin Marsh Mallow. The drug referred in common parlance as Khatmi (root and seed)

and in Ayurveda, Baikh khatmi (root) and Tukhme Khatmi (seed) in Unani and Althea

5Rajeev Kr. Sharma

officinalis (root) in Homoeopathy. The generic name of drug plant species (Althea-

Greek-to heal) also reflects its medicinal utility. It is anti-inflammatory, alterative,

1Pharmacopoeial Laboratory for

Indian Medicine, antacid, anti-tussive, aphrodisiac, antidiarrhoeal, demulcent, diuretic, emollient,

Ministry of Ayush, Govt. of India,

expectorant, laxative, mucilaginous, nutritive, rejuvenative and soothing. The drug

Kamla Nehru Nagar,

is traditionally used in the treatment for the irritation of mucous membranes, as a

Ghaziabad-201002

gargle for mouth and throat ulcers, gastric ulcers, relief to bronchial asthma, in

2Drug Standardization Research

intestinal disorder like colitis, diverticulitis. It also stimulates the immune system

Institute (CCRUM),

and production of white blood cells.

PLIM Campus,

Ghaziabad-201002

Methodology

3RegionalResearch Institute

of Unani Medicine,
Botanically identified genuine sample of Unani drug Gule-e-Khatmi was subjected

1 West Mada Church Street,

to macroscopic and microscopic study, powder analysis and Thin-layer-

Royapuram,Chennai-600013

Chromatography. Hand sections were stained and mounted in canada balsam

4Central
Council for Research
for anatomical studies. Lignifications on smoothed cross-surface were studied

in Unani Medicine,

with phloroglucinol-HCl. For studying powder, Jackson and Snowdon (1992) was

61-65 Institutional Area,

Janakpuri, New Delhi-110058 followed. To determine physico-chemical constants, Indian Pharmacopoeia

(Anonymous, 1955 & 1966) was followed. Standard prescribed procedures for

5Pharmacopoeia Commission for

histochemical studies (Johanson, 1940; Youngken, 1951; Cromwell, 1955; Trease

Indian Medicine and Homeopathy,

PLIM Campus, and Evans, 1978), Chromatography (Shellard, 1968; Stahl, 1969; Smith and

Ghaziabad-201002
Feinberg, 1972) were adopted. The informatics has been compiled by reviewing

the available literature.

Informatics

Drug Specification: The drug consists of dried flowers Althaea officinalis Linn.



Systematics

Family: Malvaceae.

Genus: Althaea

Synonyms: Althaea sublobata Stokes, Althaea taurinensis DC, Althaea vulgaris

Alef., Malva officinalis (l.) Schimp. & Spenn. ex Schimp. & Spenn.

A perennial herb that dies in the winter. Stem erect, 60-90 cm; leaves thick, three

to five lobed with serrated edges and deep veins and covered with a fine down

giving them a silvery shin and softness; stipules linear-subulate; Flowers

peduncled, in axillary clusters, 2.25-5 diam, five separate petals, rosy to off white

with purple lilac stamens; anther sub-globose; ovary many celled; ovules one in

each cell, carpels numerous (Kirtikar and Basu, 1975) (Figure 1. A & B).

Fig 1. Althaea officinalis Linn. (A. Plant in natural habitat and; B. Inflorescence)


Flowering and Fruiting: July- September, Fruiting: August-October.

Distribution: The plant species is distributed in temperate regions of the world; it

grows in salt marshes, damp meadows, by the sides of ditches, sea and on the

banks of tidal rivers. In India, it occurs in Punjab and Kashmir and often it is

cultivated
Observations & Results

I. Macroscopic Characteristics

A. Drug: Dried flowers are cream in colour, ebractetae, pedicellate, hairy, complete,

regular, actinomorphic, bisexual, hypogynous, mucilaginous and cyclic flowers.

Epicalyx 6 to 9 fused, green and hairy; Calyx 5 gamosepalous, companulate 5-

fid, hairy and green; Corolla 5 petals, obovate with sinuate apex, mucilaginous

and twisted; Androecium monoadelphous, epipetalous, staminal tube completely

encircled the gynoecium, the staminal tube gives many free filaments that bear

anthers at their tips, anthers monothecous, extrose; pollen grains large, multiporate

and spinous; Gynoecium polycarpellary, syncarpous, superior, multilocular,

placentation axile, style fused free above, style as many as carpels, stigma linear

(Fig. 2. A & B).

B. Powdered Drug: The powdered drug is light or pale cream.

II. Micro-Morphological Characteristics

A. Drug:

(i) Pedicel: T.S. of pedicel shows circular in outline; epidermis consisting of single

layer of parenchyma cells covered with thick cuticle; numerous trichomes present

on the surface of the epidermis, simple unicellular covering trichomes, stellate

trichomes and glandular trichomes present; collenchyma consisting of 8 to 10

layers of cells followed by parenchyma cells; vascular cylinder consisting of 20 to

25 vascular strands; each vascular strand consisting of 2 to 6 rows of vessels

Fig. 2 A. Flower Fig. 2 B. L. S. of Flower


with phloem on the outer side; pith consisting of large parenchyma cells in the

centre; numerous druses of calcium oxalate crystals present in almost all regions

of the pedicel (Fig. 3. A & B).

(ii) Epicalyx: T.S. of epicalyx shows epidermis consisting of single layer of

parenchyma cells covered with thick cuticle; simple unicellular covering trichomes,

stellate trichomes and glandular trichomes present; mesophyll region consisting

of only parenchyma cells filled with chloroplast; vascular strand consisting of 2 to

Fig. 3 A. T. S. of Pedicel (Diagrammatic)

Fig. 3 B. T. S. of Pedicel (Cellular)


4 rows of vessels with phloem on the lower side; numerous druses of calcium

oxalate crystals present in almost all regions (Fig. 4. A & B).

(iii) Calyx: T.S. of calyx shows epidermis consisting of single layer of parenchyma

cells covered with thick cuticle; simple unicellular covering trichomes, glandular

trichomes and numerous stellate trichomes present only on the lower side;

mesophyll region consisting of only parenchyma cells filled with chloroplast;

vascular strand consisting of 2 to 5 rows of vessels with phloem on the lower side;

numerous druses of calcium oxalate crystals present in almost all regions (Fig. 5.

A & B).

Fig. 4 A. T. S. of Epicalyx (Diagrammatic)

Fig. 4 B. T. S. of Epicalyx (Cellular)

Fig. 5 A. T. S. of Calyx (Diagrammatic)


Fig. 5 B. T. S. of Calyx (Cellular)

(iv) Corolla (lower side): T.S. of corolla shows epidermis consisting of single layer

of parenchyma cells covered with thick cuticle; vascular cylinder consisting of 10

to 15 vascular strands; each vascular strand consisting of 2 to 4 rows of vessels

with phloem on the lower side; rest of the mesophyll region consisting of

parenchyma cells with air spaces; numerous druses of calcium oxalate crystals

present; unicellular covering trichomes present only at corners of the corolla (Fig.

6. A & B).

(v) Corolla: T.S. of corolla shows epidermis consisting of single layer of parenchyma

cells covered with thick cuticle; vascular strand consisting of 2 to 4 rows of vessels

with phloem on the lower side; very few parenchyma cells present with large air

spaces in the mesophyll region (Fig. 6. C & D).

Fig. 6 A. T. S. of Corolla towards lower end (Diagrammatic)


Fig. 6 B. T. S. of Corolla towards lower end (Cellular)

Fig. 6 C. T. S. of Corolla (Diagrammatic)

Fig. 6 D. T. S. of Corolla (Cellular)


B. Powdered Drug: Powdered drug consist of parenchyma cells, spiral vessels

upto 20, simple unicellular covering trichomes, stellate trichomes anther wall in

surface view, pollen grains large, multiporate and spinous upto 150 and druses

of calcium oxalate crystals upto 25 (Fig. 7).
Fig. 7. Cellular components of powdered drug


III. Physico-Chemical Constants

The analytical values in respect of physico-chemical constant of drug were

established and results are reported in Table 1.

IV. Thin-Layer Chromatography

A. 2 g of drug sample extracted with 20ml of chloroform and separated alcohol
and refluxed on a water bath for 30 min. Filtered and concentrated to 5ml to carry

out the Thin- Layer chromatography. Chloroform extract applied on TLC plate.

The plate was developed by using Toluene: Ethyl acetate (9: 1) as mobile phase.

After development the plates were allow drying in air and examined under UV

(254nm) to record spots and Rf values. The plates were dip in vanilline sulphuric

acid reagent followed by heating at 1100 for 5 min and observed under visible

light and its shows different spots. Inferences are tabulated below (Table 2).

Table 1: Analytical Values of Physico-chemical Constants

Sl.No. Physico-Chemical Constants Analytical values

1. Moisture content, % w/w 14.91%

4. Total Ash, % w/w 12.25%

5. Acid insoluble ash, % w/w 5.65%

6. Alcohol soluble extractive % w/w 4.59%

7. Water soluble extractive % w/w 34.81%

Table 2: TLC fingerprinting data

Drug Mobile Derivatizing Visualiza- No. Rf Values

Phase/ Reagents tions of of bands

Solvent Spots

System

Althaea Toluene : 1% vanillin- Under UV 5 0.65 (Pink), 0.55,

officinalis Ethyl sulphuric 254nm 0.34 (Light pink),

Linn. acetate acid 0.21 (Green) and

(9 : 1) reagent 0.10 (Pink)

Under UV 5 0.65 (Red), 0.56

366 nm (Fluorescent green),

0.51, 0.20(Pale blue) and

0.10 (Fluorescent green)

After 9 0.94(Grey), 0.87(Violet),

derivatization 0.75(Grey), 0.63 (Blue),

0.54 (Yellow), 0.44

(Violet), 0.21(Grey), 0.13

(Green) and 0.10 (Violet).


B. The alcohol extract applied on TLC plate and developed by using Toluene:

Ethyl acetate (1: 1) as mobile phase. After development, the plate was allowed to

dry in air and examined under UV (254nm).

Table 3: TLC fingerprinting data

Drug Mobile Derivatizing Visualiza- No. Rf Values of
Phase/ Reagents tions of bands

Solvent Spots

System

Althaea Toluene : 1% vanillin- Under UV 5 0.86 (Light pink),

officinalis Ethyl sulphuric 254nm 0.62 (Yellowish green),

Linn acetate acid reagent 0.58, 0.42 (Pink) and

(1 : 1) 0.13 (Light pink)

Under UV 5 0.90 (Blue), 0.62

366 nm (Yellow), 0.56 (Violet),

0.51 and 0.14 (Blue).

After 8 0.90 (Grey), 0.84,

derivatization 0.74 (Violet), 0.60 (Grey),

0.56 (Yellowish brown),

0.46 (Grey), 0.31(Violet)

and 0.20 (Blue).

Discussion

The present pharmacognostical studies on Althaea officinalis Linn. highlight macro

and microscopical characters which can be employed for authentication of

drug.Physico-chemical data are useful in the assessment of purity and strength

of drug. Thin-layer chromatographic profile is important parameters to identify

and assessing the presence of active and other phyto constituents in the drug.

The standards developed are easy, reliable and cost effective tool for proper

identification and detection of adulteration in the drug material claimed to be

resourced from Althaea officinalis Linn.

References

Anonymous, 1955. Pharmacopoeia of India. Manager of Publications, Govt. of

India, New Delhi.

Anonymous, 1966. Pharmacopoeia of India. Manager of Publications, Govt. of

India, New Delhi.


Cromwell, B.T., 1955. The Alkaloids. In: Modern Methods of Plant Analysis (Eds.

Paeck, K. and M.V. Tracy) Vol. 4. Springer Verlag, Heidelberg.

Jackson, B.P. and Snowdon, D.W., 1992. Powdered Vegetable Drugs. Churchill

Ltd., London.

Johansen, D.A., 1940. Plant Microtechnique. Mc Graw Hill Book Co., New York.

Kirtikar, K.R. and Basu, B.D., 1975. Indian Medicinal Plants, Vol. 1, 3rd ed. L.M.

Basu, Allahabad.

Shellared, E.J., 1968. Quantitative paper and thin-layer chromatography. Academic

Press, London.

Smith, I. and Feinberg, J.G., 1972. Paper chromatography, Thin layer

chromatography and Electrophoresis. Longmans, London.

Stahl, E., 1969. Thin-layer Chromatography. A Laboratory Hand book. (Translated

by M.R.F. Ashworth). Allen and Unwin, London.

Trease, G.E. and Evans, W.C., 1978. Pharmacognosy, 11th edn. Bailliere Tindel,

London.

Youngken, H.W., 1951. Pharmaceutical Botany, 7th ed., The Blackistan Company,

Toronto.



128
Development of Abstract

Standards of hysicochemical and Phytochemical standardization is

Sapistan
(Cordia
dichotoma
P

considered a pre-requisite for the assessment of biological activity or determination
of biological standards of the plant material. It provides the analytical characteristics
which may prove to be useful in fixing the physicochemical standard for the Unani
herbal drugs.
Forst.f.) Sapistan (Cordia dichotoma) belongs to the family Ehretiaceae; its fruits and leaves

are used in respiratory diseases such as asthma, cough, diphtheria, pneumonia,

*Abdul Haleem, sore-throat, coryza. An effort has been made to carry out the physicochemical

Abdul Latif, and phytochemical studies of plant. Physicochemical parameters as Extractive

Abdur Rauf, Values: Petroleum Ether (1.9%), Di-ethyl ether (0.85%), Chloroform (0.59%),

Nazish Siddiqui Acetone (0.12%), Alcoholic (6.13%), Aqueous (10.44%); Solubility: Water (9.44

and

%) & Alcohol (1.16 %); Moisture contents (3.45 %), Total Ash values (7.188%), pH

Sumbul Rehman

of 1% (6.76) & 10% solution (6.16) and loss on drying (5.3%). Phytochemical

Department of Ilmul Advia, analysis: These revealed the presence of almost all the phyto-constituents in the

A.K. Tibbiya College, test drug sample i.e. alkaloid, flavonoids, glycoside, carbohydrate, tannin, protein,


amino acids, starch and resins.

Aligarh-202002

Keywords: Sapistan, Physicochemical, Phytochemical, Standardization, Cordia

dichotoma Forst.f.

Introduction

Sapistan (Cordia dichotoma Forst.f.) belongs to the family Ehretiaceae

(Anonymous, 2006). Use of the drug Sapistan in Unani system of medicine dates

back about a couple of thousands of years. It is mentioned by Theophrastus and

was not disregarded through the Arabic, Persian and Urdu authors in their books.

It was particularly mentioned by Razi (926 A.D.) Ibne Sina (1037 A.D.) Al- Harwi

(10th C.A.D). Sebestan or Sapistan is an abbreviation of sag pistan, derived from

sag, adog and pistan dugs. The fruit resembles bitches dugs in shape. Sapistan

is used for cure of the respiratory diseases (Khory and Katrak, 1984).

Sapistan is a fruit of tree. The tree is of two types, one type of tree have a large

fruits known as Rai Gond. And the other tree having small fruit called Kath Gond.

This tree consists of many branches which are pale red in colour. Flowers are

found in clusters form. Fruit get ripened in the month of May to July (Ghani, 2011).

Medicinally, the dried fruit is valued on account of its mucilaginous nature and

demulcent properties; it is much used in coughs and chest affections, also in



irritation of the urinary passages; in larger quantities it is given in bilious affections

as a laxative. Mahometan writers describe two kinds of Sapistan; the greater, the

pulp of which is separable from the stone, and the lesser, the pulp of which is

adherent.

Botany- brief description:

A middle-sized deciduous tree and usually with a crooked trunk. Bark grey, rough

with shallow longitudinal furrows. Leaves thinly coriaceous, variable in shape and

size, orbicular, ovate-elliptic. Flowers white, fragrant. Drupes ovoid, apiculate,

pink or orange to reddish when ripe, filled with a viscid pulp; stone usually 1.

Common in deciduous forests, flowers March and April; fruits May to July.

Therapeutic action and uses :

Fruits-astringent, anthelmintic, diuretic, demulcent, expectorant. Used in affection

of urinary passages, diseases of the lung and spleen. Bark used in dyspepsia

and fever. Pulp used in ringworm. Leaves used in ulcer, headache (Ambasta,

1986; Chopra et al., 1956; Kirtikar and Basu, 1996; Nadkarni, 1989).

No work appears to have been reported regarding standardization of this drug so

far. Keeping in mind the medicinal importance of this plant in Indian Systems of

Medicine, a physico-chemical and phytochemical study of this drug was carried

out in order to fix-up its pharmacopoeial standards.

Material and Method

Collection of plant material: The drug samples of Sapistan (fruits) were collected

from Bara Duwari market of Aligarh city and were identified. Voucher specimens

were preserved in the herbarium of Medicinal Plants Lab in the Department of

Ilmul Advia, F/O Unani Medicine, Aligarh Muslim University, Aligarh (Voucher No.

SC-0141/14).

Chemical parameters: First, the organoleptic characters were studied. The dried

powder of the fruits of Sapistan was used for chemical analysis. Various physico-

chemical studies like total ash, acid insoluble ash, water soluble ash, alcohol and

water soluble matter, moisture content, successive extractive values using soxhlet

extraction method, bulk density and pH studies were carried out as per guidelines

of WHO (Anonymous, 1998, 2008). Qualitative analysis of the drug was conducted

to identify the organic chemical constituents present in the drug (Overtone, 1963;

Harborne, 1973).

The thin layer chromatographic analysis was conducted following Stahl (1969)

and Harborne (1973) on precoated silica gel 60F254 TLC plates. The plates were


visualized in day light, in short UV and Long UV. They were also derivatised using

iodine vapors.

Observations

(a) Organoleptic characters: The powder of the fruits of Sapistan was brown with

fruity smell any characteristic odour (Table 1).
(b) Physico-chemical constants: The analytical values of different physico-

chemical constants were determined (Table 2).

Table 1: Organoleptic Characters of Cordia dichotoma Forst.f.

S.No. Organoleptic parameters Observations

1. Colour Brwon

2. Smell Fruity

3. Taste Disagreeable

Table 2: Physicochemical study of Powder of Sapistan

S.No. Parameters Percentage (w/w)*

1 Ash value

Total ash 7.188

Acid insoluble ash 1.22

Water soluble ash 5.92

2 Soluble Part

Ethanol soluble 1.66

Aqueous soluble 9.44

3 Successive Extractive Values

Pet. Ether 1.72

Di-ethyl ether 0.85

Chloroform 0.59

Acetone 0.12

Alcohol 6.13

Aqueous 10.44

4 Moisture content 3.45

5 Loss on Drying 5.3

6 pH values

1% water solution 6.76

10% water solution 6.16

7 Bulk density 0.67

*Note: Values are average of three experiments.


(c) Qualitative analysis of organic chemical constituents of drug: The

phytochemicals present in the drug were identified on the basis of different

chemical tests given for various plant constituents (Table 3).

(d) FTAR Analysis: Fluorescence analysis of the successive extract was studied

under day light as well as Ultra Violet (short and long wave length) light, results

have been summarized in Table-4. FTAR Analysis was also done of the

powdered drug after reacting them with various chemical reagents (Table 4).

Table 3: Preliminary Screening of major Phyto-chemicals

S.No. Chemical Constituent Tests/Reagent Inference

1 Alkaloids Dragendorffs reagent +

Wagners reagent +

Mayers reagent +

2 Carbohydrate Molischs Test +

Fehlings Test +

Benedict Test +

3 Flavonoids Mg ribbon and Dil.Hcl +

4 Glycosides NaOH Test +

5 Tannins/Phenols Ferric Chloride Test +

Liebermanns Test +

Lead Acetate Test +

6 Proteins Xanthoproteic Test +

Biuret Test +

7 Starch Iodine Test +

8 Saponins Frothing with NaHCO3 +

9 Steroid/Terpenes Salkowski Reaction +

10 Amino Acids Ninhydrin Solution

11 Resin Acetic Anhydride test +

Indications: Absence and + presence of constituent.

Table 4: FTAR Analysis of Sapistan

S.No. Extract Day Light UV Long UV Short

1. Pet. Ether Brown Black Greenish

2. Di-ethyl ether Greenish Bluish Greenish

3. Chloroform Light Green Black Light Green

4. Acetone Brown Bluish Greenish

5. Alcohol Brown Black Dark Brown

6. Aqueous Brown Black Black


(e) Thin layer chromatographic profile: Thin layer chromatographic analysis of

successive extracts was carried out using different solvent systems and

visualizing agents and Rf values were calculated to standardize the drug for

its identity and purity (Table 5).

Table 5: Fluorescence Analysis of Sapistan with different chemical reagents

S. Powdered drug + Day light UV short UV long

No. Chemical Reagent

1. Powdered drug + Conc. HNO3 Golden Light Green Green

2. Powdered drug + Conc. Hcl Brown Light Green Dark Green

3. Powdered drug + Conc. Dark Brown Dark Green Black

H2SO4

4. Powdered drug + 2% Iodine Brown Green Black

solution

5. Powdered drug + Glacial Light Brown Light Green Dark Green

Acetic acid + HNO3

6. Powdered drug + Glacial Pale Light Green Black

acetic acid

7. Powdered drug + NaOH(10%) Brown Light Green Dark Green

8. Powdered drug + Dil. HNO3 Brown Green Green

9. Powdered drug + Dil. H2SO4 Brown Green Dark Green

10. Powdered drug +Dil. Hcl Light Brown Light Green Black

11. Powdered drug + Dark Brown Bright Green Green

Dragendorffs

12. Powdered drug + Wagners Brown Green Dark Green

Reagent

13 Powdered drug + Benedicts Blackish Light Green Black

reagent

14 Powdered drug + Fehling Brown Light Green Green

reagent

15 Powdered drug + KOH (10%) Brown Green Black

Methanolic

16 Powdered drug + CuSO4 (5%) Brown Light Green Black

17 Powdered drug + Ninhydrin Brown Green Black

(2%) in Acetone

18 Powdered drug + Picric Acid Yellow Bright Green Black

19 Powdered drug + Lead Brown Light Green Black

Acetate (5%)


Table 6: Thin Layer Chromatography

Treatment Mobile phase: No of Rf value and colour of

spots spots

Petroleum Ether Extract

Day Light Petroleum ether : 1 0.42 (Brown)

UV Short Di-ethyl ether (4:1) 1 0.42 (Bluish)
Iodine Vapour 1 0.42 (Yellowish)

Aqueous Extract

Day Light Butanol: Acetic 1 0.75 (Brown)

UV Short acid: Water (5:1:4) 3 0.64 (Bluish), 0.78 (White),

0.85 (White)

UV Long 4 0.21 (white), 0.42 (white),

0.78 (white), 0.85 (white)

Iodine Vapour 1 0.75 (Pale Yellow)

Alcoholic Extract

Day Light Chloroform: 1 0.62 (Pale yellow)

UV Short Methanol (9:1) 2 0.62 (Purple) , 0.75 (Purple)

UV Long 2 0.62 (white), 0.75 (White)

Iodine Vapour 1 0.62 (Pale Yellow)


Physico-chemical standardization is of prime importance in quality control of Unani

drugs. As the efficacy of many drugs mainly depends upon its physical and

chemical properties, therefore, the determination of physico-chemical characters

for the authenticity of a drug is necessary before studying any medicinal property.

Phyto-chemical constituents present in the drug vary, not only from plant to plant

but also among different samples of same species, depending upon various

atmospheric factors, storage and drying conditions.

The present study is an attempt to fix-up pharmacopoeial standards of drug

Sapistan to ascertain its quality, identity, purity and strength. Powder of the drug

has been used for study to bring out several standards like ash, solubility in alcohol

and water, successive, extractive values and quality screening of

physicochemicals, total alkaloids, total flavonoids, phenol, nitrogen, fatty matter,

sterol/ terpenes, protein and carbohydrates.

Based on the various physico-chemical and phytochemical parameters, the drug

Sapistan has been standardized to ensure its use in manufacturing quality and

genuine herbal preparations.


Sapistan Fruits (Cordia dichotoma Forst.f.)

TLC of Alcoholic Extract of Sapistan TLC of Aqueous Extract of Sapistan

References

Afaq, S.H., Tajuddin, Siddiqui, M.M.H., 1994. Standardization of Herbal Drugs.

Publication Division, AMU (Aligarh), pp. 33-34, 41-42, 100, 143-146.

Ambasta, S.P., 1986. The Useful Plants of India. PID, CSIR, New Delhi.

Anonymous, 1997. Standardization of Single Drugs of Unani Medicine, Part III.

CCRUM New Delhi, pp. 124-128.

Anonymous, 1998. Quality control methods for medicinal plant materials. World

Health Organization, Geneva, pp. 25 -28.


Anonymous, 2006. National Formulary of Unani Medicine, Part I. CCRUM, Dept.

of AYUSH, New Delhi, p. 272.

Anonymous, 2008. Quality control manual for Ayurveda, Siddha and Unani

medicine. Govt. of India, Dept. of AYUSH, New Delhi, pp. 21 29.

Anonymous, 2008. Unani Medicinal Plants of Tarai Forests in Kumaon Region of

Uttarakhand. Central Council for Research in Unani Medicine, p. 60.
Baitar I., 1999. Al-jamiul-mufridat-ul-advia-wa-ul-aghzia. Urdu Translation. Part.III.

CCRUM, New Delhi, pp. 23-24

Chopra, R.N., Nayar, S.L. and Chopra, I.C., 1956. Glossary of Indian Medicinal

Plants, CSIR, New Delhi.

Dymock, 1891. Pharmacographica Indica, Part II. Published by Institute of Health

and Tibbi Research, Hamdard National Foundation, Pakistan, pp. 518-519.

Ghani Najmul, 2011. Khazainul Advia. Idara Kitabul Shifa, New Delhi, pp. 787-

788.

Harborne, J. B., 1973. Phytochemical methods. Chapman and Hall. London, p.

70.

Ibn-e-Sina, 2007. Al-Qanoon-fil-tib, Vol. II. Urdu translation by Ghulam Hussain

Kantoori. Idara Kitabul Shifa, New Delhi, p. 167.

Jenkins, G.L., Knevel, A.M. and Digangi, F.E., 1967. Quantitative Pharmaceutical

Chemistry. 6th edition. The Blackiston Division. McGraw Hill Book Company,

U.S.A., pp. 225, 235, 379, 425, 463, 492.

Khory, R.N. and Katrak, N.N., 1984. Materia Medica of India and their Therapeutics.

Neeraj Publishing House, Delhi, p. 421.

Kiritikar, K.R. and Basu, B.D., 1996. Indian Medicinal Plants, Vol. III. International

Book Distributers (Dehradun), pp. 1674-1679.

Nadkarni, K.M., 1989. The Indian Materia Medica, Vol. II. Bombay Prakashan

Pvt. Ltd., Bombay, pp. 379-380.

Overtone, K.H., 1963. Isolation, Purification and preliminary observation in

elucidation of structures by physical and chemical methods. Bentley

Interscience Pub., New York, p. 34.

Stahl, 1969. Thin Layer Chromatography: A laboratory handbook, Springer Verlag

student edition. Springer Verlag, Berlin, pp. 52 86, 127 128, 900.


HIPPOCRATIC JOURNAL OF UNANI MEDICINE

Instructions to contributors

1. The paper(s) should be submitted in duplicate. Submission of a paper will

be taken to imply that it is unpublished and is not being considered for

publication elsewhere.

2. Papers should be written in English language and typed with double spacing

on one side of A-4 size paper leaving top and left hand margin at least 1"

(One inch) wide. Length of the paper should normally not exceed 12

pages.

3. Papers should be headed by a title, the initial(s) and surname(s) of

author(s) followed by address.

4. Each paper should bear abstract, 2 to 5 keywords, introduction,

methodology, observations, results and discussion followed by

acknowledgements and references.

5. In all studies of plants or animals proper identification should be made as

to the materials used.

6. While submitting the paper(s) for publication, Author(s) should decode the

drugs specially in case of clinical studies.

7. Bibliographical references should be listed in alphabetical order of the

author at the end of the paper. Authors should be cited in the text only by

their surname(s) but their initial(s) should be shown in the bibliography.

8. References to periodicals should include the name(s) and initial(s) of

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9. Reference should be cited in the text in parentheses by the name(s) of

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when the authors name is part of the sentence, e.g. Jain (1991) has

reported that. If there are more than two authors it is in order to put et

al. after the first name, e.g., Khan et al., 1981.


10. Each table should be typed on a separate sheet of paper. Tables should

be numbered consequently in Arabic numerals e.g. Table 1, Table 2 etc.,

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and kept as simple as possible and should be referred to in the text as

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11. Figures (including photographic prints, line drawings on strong white or
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HIPPOCRATIC

Volume 10, Number 1, January - March 2015

CENTRAL COUNCIL FOR RESEARCH IN UNANI MEDICINE

International Advisory Board

2. Exploration of Mechanism of Antiinflammatory Action of Habbe-Gule-Aakh ............................................. 9

3. Toxicity Study of Qurs-e-Hudar in Experimental Animals ......................................................................... 15

7. A Review on Phytochemistry, Pharmacological Properties and Biotechnological Studies in .................. 53

9. Quality Evaluation of Jawarish-e-Ood Kibreet ........................................................................................... 85

April 22, 2015

Hounz field (HF) Units value on CT scan of liver.

Keywords: Non-Alcoholic Fatty Liver Disease (NAFLD), Fatty liver, Tashhamul

kabid, Unani Medicine, UNIM 104 coded drug.

*Hafiz C. Mohammad Aslam,

Non-alcoholic fatty liver disease (NAFLD) is a common clinic-pathological condition

characterized by significant lipid deposition in the hepatocytes of the liver

Regional Research Institute of

1 West Mada, Church Street,

lobular inflammation, hepatocellular injury, and Mallory hyaline, progressive

(Tominaga et al., 1995).

*Author for correspondence

Hippocratic Journal of Unani Medicine 1

centre of Quwa-Al-Tabiyah (Natural Faculty) (Ibn-e-Sina 1906). It is responsible

excretion of Fudhlat (waste products) (Abusahal Masihi 810-895 A.D). In Unani

classical text, Su-e-Mizaj-e-Kabid (alteration in normal temperament of liver)

particularly due to kasrate buroodat (excessive coldness) somewhat resembles

ultimately help in deposition of Raddi Shaham (abnormal excessive fats) (Ibn-e-

several possible treatments are under investigation including Vitamin E and C,

liver. However, considering necessity of novel medical intervention, the Central

coded drug UNIM-104 in the treatment of non-alcoholic, non-obese and non-

diabetic fatty liver disease.

Materials and Methods

Ethical Committee of the Institute. Altogether 35 cases from GOPD of RRIUM,

upper abdomen. Subjects with pregnancy or lactation, those unwilling to give

neurological disorders etc) were excluded from the study.

Clinical assessment and laboratory investigations were carried out at baseline

Hippocratic Journal of Unani Medicine 2

S.Alkaline Phosphatase, S. Urea, S. Creatinine, S.Cholestrol, S.Triglycerides,

Urine: Routine and Microscopical examination.

value < 0.05 was considered to be significant.

Results and Discussion

NAFLD is an increasingly important chronic liver disease with a wide spectrum of

histopathology, ranging from bland steatosis to cirrhosis. It is often asymptomatic

and discovered incidentally on routine laboratory screening. In the absence of

established therapies, treatment is generally directed at optimizing body weight

and controlling risk factors.

53.33% patients were of Balghami (phlegmatic temperament) and 36.66% were

of Damavi Mizaj (sanguine temperament) suggesting that those with colder

substantiating the claim made by Unani physicians (Ibn-e-Sina -19.6), Ibn-e-

Though majority of the trial participants were asymptomatic which corroborate

kabid properties of polyherbal coded research formulation (UNIM 104).

(Table 3 & 4) There might be no biochemical evidence of liver disease as stated

Hippocratic Journal of Unani Medicine 3

3 Duration of disease Mean SD 9.66 10.57

4 BMI (Kg/m2) Base line 28.05 10.17

After treatment 27.1 1.68 > 0.028

5. Body weight (kg) Base line 73.8 10.17

After treatment 71.26 10.11 > 0.42

6. Mizaj Damavi (sanguine) 11 (36.66%)

Balghami (phlegmatic) 16 (53.33%)

Saudavi (melancholic) 2 (6.66%)

Safravi (choleric) 1 (3.33%)

Status No. of Fatigue Malaise Nausea Loss of Hepato- Dull ache

Cases Appetite megaly in RUQ