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452 Asian Pac J Trop Dis 2014; 4(6): 452-456
Document heading doi:10.1016/S2222-1808(14)60605-8 2014 by the Asian Pacific Journal of Tropical Disease. All rights reserved.
Peer reviewer Objective: To quantify the microorganisms contaminating the common preservatives used in food
S usheel K umar S ingh, P h D , C entre as well as to detect their in vitro anti-bacterial traits.
for Medical Parasitology, Department Methods: A total of 9 preservatives were subjected to conventional cultural and biochemical
of International Health, Immunology methods for microbial enumeration. Anti-bacterial activities were demonstrated through the agar
and Microbiology, Faculty of Health well diffusion method.
Sciences, University of Copenhagen Results: All samples were found to be contaminated with bacteria up to 105 CFU/g and with
CSS , ster F arimagsgade 5 , 1014 the fungal flora within a range of 101-102 CFU/g. Escherichia coli, Pseudomonas spp. and
Copenhagen K, Denmark. Staphylococcus spp. were demonstrated in most of the samples. Sodium sulfite and citric acid
Tel: +45-52696790 possessed the strongest anti-bacterial trait against all of the test bacteria. Acetic acid exhibited
E-mail: susi@sund.ku.dk, activity against 6 out of 8 test bacteria while vinegar exhibited the activity against 4 bacteria.
Activity of salt was demonstrated only against Listeria spp. and Bacillus spp., while activity of
Comments sugar and honey was found only against Escherichia coli and Klebsiella spp., respectively.
T his is a good study in which the Conclusions: According to the current investigation, sodium sulfite and citric acid samples were
authors evaluated the distribution found to be satisfactory preservatives both in terms of microbiological criteria and their anti-
of microbial loads in common bacterial traits.
food preservatives. T he results are
interesting and suggested that sodium
bisulfate and citric acid samples were
satisfactory preservatives both in terms
of microbiological criteria and their KEYWORDS
anti-bacterial traits.
Preservatives, Microorganisms, Anti-bacterial activity, Food safety, Public health
Details on Page 455
arose through canning, pasteurization, irradiation, filtration, was performed by using agar well diffusion method as
addition of natural or synthetic preservatives[13-16]. described previously[21]. Lawns of bacterial pathogens (E.
While the microbiological spoilage in food, pharmaceutical coli, Klebsiella spp., Pseudomonas spp., Salmonella spp.,
and cosmetics items are globally common, the use of Staphylococcus spp., Vibrio spp., Listeria spp. and Bacillus
preservatives used in those products against microbial spp.) were prepared over the Mueller Hinton agar plates and
contamination is also expected [17-20] . I n context of holes were made in the Mueller Hinton agar by cork borer.
Bangladesh the microbial prevalence in food and consumer Each of the homogenized preservative blends (around 10 g/
items is too frequent, resulting in disease outbreaks[21-27]. mL) was then introduced separately in the specified hole
However, the knowledge on the extent of microbiological with a positive control (streptomycin, 10 g/mL) and negative
contamination of the associated preservatives have not control (normal saline). Presence of clear zone around the
been provided. Moreover, the microbial content in the sample suspension indicated the presence of anti-bacterial
preservatives as well as the demonstration of antimicrobial activity.
activity would imply the efficiency of the preservatives.
Based on these facts, current study attempted to isolate and
enumerate the microorganisms accessing the preservatives 3. Results
and to detect their anti-bacterial traits.
All samples studied were found to be populated with
bacteria within a range of 102-105 CFU/g with the presence
2. Materials and methods of specific pathogenic microorganisms, i.e., E. coli,
Pseudomonas spp. and Staphylococuus spp. in most of the
2.1. Sampling, sample processing and microbiological samples (Tables 1 and 2). Sodium benzoate, vinegar and
analysis honey samples were found to harbor the highest number of
spoiling bacteria (~105 CFU/g). Next prevalence was noticed
A total of 9 samples of different categories of natural in case of sugar and turmeric samples (104 CFU/g) while salt
and synthetic preservatives ( with appropriate dates of and acetic acid samples were populated by a lesser extent
manufacturing and expiry on the packs) were collected of bacteria (103 CFU/g). The least microbial spoilage was
from different super shops in Dhaka city during September in the citric acid and sodium sulfite samples (102 CFU/g).
2013 to December 2013. Samples included salts, sugars, Except sugar and vinegar, the other 7 samples exhibited the
sodium sulfite, sodium benzoate, acetic acid, citric acid, proliferation of fungal population.
vinegar, honey and turmeric preservatives. All samples Table1
were transported to the Microbiology Laboratory in order Microbial analysis of different preservative used in food.
to assess their microbiological quality. A total of 10 g of Total aerobic Fungi E. coli Pseudomonas Staphylococcus
Sample
samples were homogeneously mixed with 90 mL of buffer bacteria (CFU/g) (CFU/g) (CFU/g) spp. (CFU/g) spp. (CFU/g)
peptone water, and serial dilutions were prepared up to 10-4 Sugar (n=5) 4.110 7.010 1.910
4 1 2
0 0
Salt (n=5) 3.210 3.710 8.010
following the standard protocols[5-6]. An aliquot of 0.1 mL of
3 2 1
0 0
Acetic acid (n=5) 3.610 4.910 1.210
3 2 1
0 0
each suspension from the dilution 10-2 and 10-4 was spread Citric acid (n=5) 2.910
2
2.010
2
0 0 0
onto nutrient agar plate to enumerate the total bacteria and Sodium benzoate (n=5) 5.810
5
4.510
2
1.610
1
0 3.810
2
on Sabouraud dextrose agar plate for the estimation of fungal Vinegar (n=5) 6.410
5
0 2.210
1
3.610
2
4.610
2
load. Then the nutrient agar plate and Sabouraud dextrose Sodium sulfite (n=5) 2.810 2.110
2 2
0 0 0
Honey (n=5) 3.910 3.310 1.810 3.310
agar plates were incubated at 37 C for 18 to 24 h and at 25 C
5 2 1 2 2
110
Turmeric (n=5) 4.310 3.910 1.610 5.910
4 1 1 2
0
for 48 to 72 h, respectively. Fecal coliforms were absent in all cases.
For the enumeration of specific pathogens, 0.1 mL from Acceptable microbial limits[48]: Total aerobic bacteria: 10 CFU/g; total fungal
3
the dilution of 10-2 and 10-4 of each sample was spread onto load: 10 CFU/g; absence of fecal coliforms, E. coli, S. aureus and Pseudomonas
2
membrane fecal coliform, MacConkey agar, mannitol salt spp. per 1 g of the preservative; absence of Salmonella spp. per 10 g of the
agar, and cetrimide agar for the enumeration of total fecal preservative.
coliform, Escherichia coli (E. coli), Staphylococcus spp.,
Table 2
and Pseudomonas spp., consecutively. All the plates were
C onfirmative biochemical tests for the different isolates.
incubated at 37 C for 24 h except membrane fecal coliform
H2S Indole MR VP Citrate Motility Oxidase
agar which was incubated at 44.5 C for 18-24 h. Presence Assumed TSI
Slant Butt Gas reaction test test test Test test
of E. coli was further confirmed by the appearance of organism
TSI: Triple sugar iron test; Y: yellow (acid); R: red (alkaline); MR:
methyl red; VP: Voges-Proskauer.
2.2. Determination of anti-bacterial activity of the
preservatives Sodium sulfite and citric acid were found to be free from
The anti-bacterial activity of the preservative samples any contaminating specific pathogenic bacteria while
454 Tohora Sultana et al./Asian Pac J Trop Dis 2014; 4(6): 452-456
vinegar and honey samples were found to be contaminated tested in our study might be due to the relatively low
with E. coli, Pseudomonas spp. and Staphylococcus spp. Salt concentrations of these food grade products as well as
and acetic acid samples harbored only Pseudomonas spp. due to the common cause of unhygienic preparation of the
(~10 CFU/g), sodium benzoate was found to be contaminated preservatives[7,9,11,14,18]. Nevertheless, to our knowledge, the
2
with E. coli and Staphylococcus spp., and the turmeric abundance of microorganisms in common preservatives used
samples were contaminated with Pseudomonas and in Bangladesh has been first time reported in our study.
Staphylococcus spp. Additionally, the microbial load among the preservatives
tested further poses the extended spoilage of the intended
3.2. Anti-bacterial traits of the preservative samples food or pharmaceutical products to be used and hence raises
the application risk.
Among the samples studied in our study, sodium sulfite It is indeed well reported that the natural substances
showed the highest activity against all of the test bacteria including salt, sugar and vinegar are also used as traditional
(Table 3). Acetic acid was found to exhibit the anti-bacterial preservatives[29]. Besides non-toxicity with acceptability
activity against 6 out of 8 test bacteria while vinegar in taste and odour, a preservative should be effective
exhibited the activity against 4 bacteria. Anti-bacterial against a wide spectrum of microorganisms. Almost all of
activity of both of these preservatives was most prominent the preservatives, either natural or synthetic, act as either
against Pseudomonas spp. (Table 3). Another organic acid, antimicrobials or antioxidants or both and hence are known
the citric acid in our study also exhibited the activity against to prevent the growth of molds, yeasts and bacteria[13,29-
all bacteria; however, to a lesser extent compared to that 33]. The history of sodium sulfites anti-bacterial trait has
of sodium sulfite. Compared to sodium sulfite, acetic acid, long been known and this preservative has been shown to
citric acid and vinegar, other samples in our study were pose the therapeutic efficacy[7,34-36]. Consistently, our study
found to pose the anti-bacterial activity to a lesser extent. also demonstrated the highest anti-bacterial activity of this
preservative, which is interestingly in consistent to the least
Table 3 microbial prevalence as observed through microbiological
Anti-bacterial activity of different preservatives (mm).
enumeration assay.
Organic acids are popular preservatives with marked
Klebsiella Pseudomonas Salmonella Staphylococcus Vibrio Listeria Bacillus
Sample E. coli
anti-bacterial traits[37-40]. In our study, both acetic acid
spp. spp. spp. spp. spp. spp. spp.
Sugar
and vinegar exhibited the anti-bacterial activity against
18 0 0 0 0 0 0 0
Salt 0 0 0 0 0 0 10 20
Antimicrobial effect of natural food preservatives in fresh basil- Moringa oleifera plant extracts in fish from Lake Victoria Basin
based pesto spreads. J Food Process Preserv 2013; doi: 10.1111/ of Kenya. Afr J Food Agric Nutr Dev 2013; 13(3): 7772-7788.
jfpp.12091. [32] Shahnawaz M, Sheikh SA, Minhas S. Role of sodium benzoate
[14] O regon S tate U niversity 2013 . Food safety & preservation: as a chemical preservative in extending the shelf life of orange
preserving for special diets. Corvallis: Oregon State University juice. Global Adv Res J Food Sci Technol 2013; 2(1): 7-18.
Extension Service; 1980. [33] Chatkaew A, Suudi M, Lim S, Hwang BH, Kim J. Pectobacterium
[15] Bahtiti NH. Study of preservative effect of propolis on the carotovorum inhibition by preservative agents in sprouting radish
storage quality of mashed potatoes. Food Sci Technol 2013; 1(2): seeds. Int Res J Biol Sci 2013; 2(9): 22-28.
17-20. [34] Kropec A, Huebner J, Frank U, Lemmen S, Hirt U, Daschner
[16] Pawar HA, Shenoy AV, Narawade PD, Soni PY, Shanbhag PP, FD. In vitro activity of sodium bisulfite and heparin against
Rajal VA. Preservatives from nature: a review. Int J Pharm staphylococci: new strategies in the treatment of catheter-related
Phytopharmacol Res 2011; 1: 78-88. infection. J Infect Dis 1993; 168(1): 235-237.
[17] Baydar H, Kuleasan H, Kara N, Secilmis-Canbay H, Kineci S. [35] Y amazaki H , T oda M , M iyaji K , Y amaguchi T , F ukutome I .
The effects of pasteurization, ultraviolet radiation and chemical [Anti-bacterial activity of sodium sulfite on the mucoid variant
preservatives on microbial spoilage and scent composition of of enteric bacteria]. Showa Igakkai Zasshi 1966; 26(2): 93-101.
rose water. J Essent Oil Bear Plants 2013; 16: 151-160. Japanese.
[18] Stratford M, Steels H, Nebe-von-Caron G, Novodvorska M, Hayer [36] In YW, Kimi JJ, Kim HJ, Oh SW. Antimicrobial activities of acetic
K, Archer DB. Extreme resistance to weak-acid preservatives acid, citric acid and lactic acid against Shigella species. J Food
in the spoilage yeast Zygosaccharomyces bailii. Int J Food Saf 2013; 33: 7985.
Microbiol 2013; 166(1): 126134. [37] Mani-Lpez E, Garca HS, Lpez-Malo A. Organic acids as
[19] Sharma N, Garcha S, Singh S. Potential of Lactococcus lactis antimicrobials to control Salmonella in meat and poultry
subsp. lactis MTCC 3041 as a biopreservative. J Microbiol products. Food Res Int 2012; 45: 713-721.
Biotechnol Food Sci 2013; 3(2): 168-171. [38] Park SH, Choi MR, Park JW, Park KH, Chung MS, Ryu S, et
[20] Siegert W. Comparison of microbial challenge testing methods for al. Use of organic acids to inactivate Escherchia coli O157:H7,
cosmetics. Household and Personal Care Today 2013; 8: 32-39. Salmonella Typhimurium, and Listeria monocytogenes on organic
[21] Ahmed T, Baidya S, Sharma BC, Malek M, Das KK, Acharjee M, et fresh apples and lettuce. J Food Sci 2011; 76: M293M298.
al. Identification of drug-resistant bacteria among export quality [39] Jang JS, Lee HJ, Oh BY, Lee JM, Go JM, Kim YH. Inactivation of
shrimp samples in Bangladesh. Asian J Microbiol Biotechnol Escherichia coli O157:H7, Salmonella and Listeria monocytogenes
Environ Sci 2013; 15(4): 655-660. by organic acid. Korean J Environ Health Sci 2007; 33: 403407.
[22] Feroz F, Senjuti JD, Noor R. Determination of microbial growth [40] Nwachukwu E, Ezeigbo CG. Changes in the microbial population
and survival in salad vegetables through in vitro challenge test. of pasteurized soursop juice treated with benzoate and lime
International J Nutr Food Sci 2013; 2(6): 312-319. during storage. Afr J Microbiol Res 2013; 7(31): 3992-3995.
[23] Noor R, Acharjee M, Ahmed T, Das KK, Paul L, Munshi SK, et [41] Bellagha S, Sahli A, Farhat A, Kechaou N, Glenza A. Studies on
al. Microbiological analysis of major sea fish availavle in local salting and drying of sardine (Sardinella aurita): experimental
markets of Dhaka city, Bangladesh. J Microbiol Biotechnol Food kinetics and modeling. J Food Eng 2007; 78: 947-952.
Sci 2013; 2(4): 2420-2430. [42] K han MAA , K han YSA . I nsect infestation and preventive
[24] Noor R, Uddin MA, Haq MA, Munshi SK, Acharjee M, Rahman measures in dry fish storage of Chittagong, Bangladesh. J Biol
MM. Microbiological study of vendor and packed fruit juices Sci 2001; 1(10): 963-965.
locally available in Dhaka city, Bangladesh. Int Food Res J 2013; [43] L eroi F , J offraud JJ . S alt and smoke simultaneously affect
20(2): 1011-1015. chemical and sensory quality of cold-smoked salmon during 5
[25] Hassan MR, Acharjee M, Das E, Das KK, Ahmed T, Akond MA, et degrees C storage predicted using factorial design. J Food Prot
al. Microbiological study of sea fish samples collected from local 2000; 63(9): 1222-1227.
markets in Dhaka city. Int Food Res J 2013; 2(3): 1491-1495. [44] Leroi F, Joffraud JJ, Chevalier F. Effect of salt and smoke on the
[26] Rabbi FA, Rabbi F, Runun TA, Zaman K, Rahman MM, Noor microbiological quality of cold-smoked salmon during storage at
R. Microbiological quality assessments of foods collected from 5 degrees C as estimated by the factorial design method. J Food
different hospitals within Dhaka city. Stamford J Microbiol 2011; Prot 2000; 63(4): 502-508.
1(1): 31-36. [45] Wei CI, Huang TS, Kim JM, Lin WF, Tamplin ML, Bartz JA.
[27] Rahman T, Hasan S, Noor R. An assessment of microbiological Growing and survival of Salmonella Montevideo on tomatoes and
quality of some commercially packed and fresh fruit juice disinfection with chlorinated water. J Food Prot 1996; 58(8): 829-
available in D haka city: a comparative study. Stamford J 836.
Microbiol 2011; 1(1): 13-18. [46] P ark DL , R ua SM J r., A cker RF . D irect application of new
[28] Cappuccino J, Sherman N. Microbiology-a laboratory manual. hypochlorite sanitizer for reducing bacterial contamination on
4th ed. Menlo Park, California: Benjamin/Cummings Publishing foods. J Food Prot 1991; 54: 960-964.
Co., Inc.; 1996, p. 13-182. [47] F o o d a n d D r u g A d m i n i s t r a t i o n . S p e c i f i c a t i o n s : t e s t
[29] Shee AK, Raja RB, Sethi D, Kunhambu A, Arunachalam KD. procedures and acceptance criteria for new drug substances
Studies on the anti-bacterial activity potential of commonly used and new products: chemical substances. S ilver S pring:
food preservatives. Int J Eng Sci Technol 2010; 2(3): 264-269. F ood and D rug A dministration; 2006 . [ O nline] A vailable
[30] Chand B. Anti-bacterial effect of garlic (Allium sativum) and from: http://www.fda.gov/downloads/ A nimal V eterinary/
ginger ( Zingiber officinale ) against Staphylococcus aureus, G uidance C ompliance E nforcement/ G uidancefor I ndustry/
Salmonella typhi, Escherichia coli and Bacillus cereus. J UCM052487.pdf. [Accessed on 14th January 2014]
Microbiol Biotechnol Food Sci 2013; 2(4): 2481-2491. [48] European Pharmacopoeia Commission. European Pharmacopeia,
[31] Onyuka JHO, Kakai R, Arama PF, Ofulla AVO. Comparison of Vol 5.6 supplement. 5th ed. Illinois, USA: Balogh Scientific Books;
antimicrobial activities of brine salting, chlorinated solution and 2006, p. 4451-4452.