Journal of Hazardous Materials 165 (2009) 804–811

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Journal of Hazardous Materials

journal homepage: www.elsevier.com/locate/jhazmat

Color removal from distillery spent wash through coagulation

using Moringa oleifera seeds: Use of optimum response surface methodology
R. Krishna Prasad ∗
School of Chemical & Biotechnology, SASTRA University, Thirumalaisamudram, Thanjavur 613 402, Tamil Nadu, India

a r t i c l e i n f o a b s t r a c t

Article history: The effects of dosage, pH and concentration of salts were investigated for an optimized condition of color
Received 19 July 2008 removal from the distillery spent wash. The optimization process was analyzed using custom response
Received in revised form 16 October 2008 surface methodology (RSM). The design was employed to derive a statistical model for the effect of param-
Accepted 16 October 2008
eters studied on removal of color using Moringa oleifera coagulant (MOC). The dosage (20 and 60 ml), pH
Available online 1 November 2008
(7 and 8.5) and concentration of 0.25 M had been found to be the optimum conditions for maximum 56%
and 67% color removal using sodium chloride (NaCl) and potassium chloride (KCl) salts respectively. The
Keywords:
actual color removal at optimal conditions was found to be 53% and 64% respectively for NaCl and KCl
Response surface design
Moringa oleifera
salts which confirms close to RSM results. The effects of storage duration and temperature on MOC studied
Coagulation reveal that coagulation efficiency of MOC kept at room temperature was effective for 3 days and at 4 ◦ C it
performed coagulation up to 5 days.
© 2008 Elsevier B.V. All rights reserved.

1. Introduction
Distilleries are amongst the most highly polluting industries
with reference to water pollution. The quantity of wastewater gen-
erated from distilleries is large and is characterized by a high
pollution load. Molasses spent wash (MSW) from distillation still
contains nearly 2% of a dark brown recalcitrant pigment called
melanoidin formed due to Maillard amino-carbonyl reaction. The
empirical formula of melanoidin is C17–18 H26–27 O10 N. It is a product
of non-enzymatic reaction between sugars and amino compounds.
The molecular weight distribution is between 5000 and 40,000. It is
acidic, polymeric and composed of highly dispersed colloids, which
are negatively charged due to the dissociation of carboxylic acids
and phenolic groups [1–3]. The dark color imparted by melanoidin
and anaerobic microbial mass in MSW leads to reduction of sunlight
penetration in rivers, lakes and thereby reducing the photosyn-
thetic activity causing harm to aquatic life. Disposal on land cause
reduction in soil alkalinity, manganese availability and inhibits seed
germination [4–6].
Several studies have been carried out concerning the decol-
orization of waste water using Cyanobacterium [7], fungi such as
Aspergillus fumigatus [8], Coriolus [9] and Phanerochaete chrysospo-
rium [10] have shown to degrade melanoidin and anaerobic mass
imparting color to spent wash. Basically being heterotrophs, these
∗ Tel.: +94434 60815 (M); fax: +91 4362 264120.
E-mail address: rkprasad cbe@rediffmail.com.
organisms tend to deplete oxygen in the effluent and further, higher
fungi are not easily adopted for aquatic habitats.
The application of electrochemical methods is another way to
treat wastewaters. This method guarantees high treatment effi-
ciency, but its effectiveness depends on the type of electrodes, the
construction of electrocoagulators and the condition under which
the process is run [11,12].
Coagulation has remained the most widely practiced method
of removing particulate and organic matter in wastewater treat-
ment. Conventional coagulants in waste water treatment are alum
(Al2 (SO4 )3 ·14H2 O), ferric chloride (FeCl3 ·6H2 O), sodium aluminate,
aluminum chloride and ferric sulfate [13–17]. Conventional coag-
ulants are basically salts of a strong acid (e.g., HCl or H2 SO4 ) and
a weak base (Al2 (OH)3 or Fe(OH)3 ); thus they are a mixture of a
cation (from a base) and an anion (from an acid). However recent
studies have pointed out several serious drawbacks of using alu-
minum salts, such as Alzheimer’s disease [18]. There is also the
problem of reaction of alum with natural alkalinity present in the
water leading to a reduction of pH [19].
Currently more effective trivalent aluminum coagulants, such
as poly aluminum chloride and poly aluminum silico sulphate
have been developed [20]. Although these new coagulants have
improved the coagulation process considerably; they have not cor-
rected all the drawbacks mentioned earlier. Therefore it is desirable
that other cost effective and more environmentally acceptable
alternative coagulants be developed to present a viable alternative.
Natural coagulants of vegetable and mineral origin like Moringa
oleifera, Prosopis juliflora and Cactus latifaria were in use in water

0304-3894/$ – see front matter © 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.jhazmat.2008.10.068
 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811 805

treatment before the advent of chemical salts, but have succumbed Table 1
Characteristics of Moringa oleifera coagulant.
progressively under modernization and survived only in remote
areas of some developing countries [21–24]. Sanskrit writings from Parameter NaCl extracted KCl extracted
India reported that the seeds of the nirmali tree (Strychnos potato- pH 6.8 6.8
rum) were used to clarify turbid surface water over 4000 years ago Conductivity (␮mho cm−1 ) 1500 1490
[25]. Chloride (mg l−1 ) 19.2 19.4
M. oleifera is now grown widely throughout the tropics. It prefers Sulphate (mg l−1 ) 9 9

hot, semi-arid regions (annual rainfall 250–1500 mm). It is suitable

for lowland cultivation at altitudes less than 600 m.The tree is tol-
erant of light frosts and can be established in slightly alkaline soils
up to pH 9. M. oleifera seeds possess effective coagulation proper-
ties, and are quite efficient in reducing turbidity, microorganisms
from wastewaters and also in sludge conditioning [26]. The active
agents of coagulation are dimeric cationic proteins of molecular
weight of approximately 13 kilodaltons (kDa) having an isoelectric
point between 10 and 11 [27].
The coagulation activity of M. oleifera coagulant (MOC) increases
with increase of ionic strength of extracting solution. The coagula-
tion capacity of MOC extracted by 1 M NaCl solutions was 7.4 times
as high as that of MOC extracted by distilled water [28,29]. The
improvement in extraction efficiency by salt was ascribed to the
salting-in mechanism. The increase in ionic strength by salt caused
the increase in solubility of active components.
M. oleifera seeds have antimicrobial activity and are utilized for
wastewater treatment in Sudan by rural women to treat highly
turbid Nile water [30]. The seeds of M. oleifera are considered to
be antipyretic, acrid and bitter. Moringa seed kernels contain oil
that is commercially known as “Ben oil” or “Behen oil” which is
used by watchmakers for illumination and lubrication of delicate
mechanisms [31].
Toxicological studies conducted so far reported that M. olifera
seeds do not constitute a serious health hazard [32,33]. A very active
antimicrobial agent (4 ␮m-4-rhamnosyloxy-benzyl-isothiocynate)
obtained from M. oleifera seeds is readily soluble in water [34].
Coagulation active component in MOC is supposed to remove
the suspended solids in wastewater by sweep coagulation mech-
anism. The cations might be electrically adsorbed to the active
component with negative charge at coagulation pH. The active com-
ponent may bind with other components by cations, which forms
the enmeshment of suspended solids by the net like structure.
Sweep coagulation gives considerably improved particle removal
than when particles are destabilized just by charge neutralization.
At least part of the reason is the greatly improved rate of aggrega-
tion, because of the increased solids concentration. Other possible
coagulation mechanism may be due to adsorption between sus-
pended particles and the coagulation active component in MOC to
permit interparticle bridging. This mechanism normally functions
by a material with high molecular weight like 1000 kDa. It is not
likely for the active component in MOC with low molecular weight
(13 kDa) to induce interparticle bridging [35].
2. Materials and methods
2.1. Preparation of coagulant
The husk covering the M. oleifera seeds were manually removed,
good quality seeds were selected, and the kernel was ground to a
fine powder using an ordinary electric blender. The active compo-
nent from coagulant was extracted using sodium chloride (NaCl)
or potassium chloride (KCl) salt solution. A concentration of 4%
(4 g of powder in 100 ml salt solution) was used throughout the
study after several trials varying from 0.5% to 8%. The whole mix-
ture was stirred for 30 min at room temperature using a magnetic
stirrer. The suspension was filtered using Whatman no. 42 filter
paper. The resultant filtrate solution was used as a coagulant. In
order to prevent any aging effects, such as change in pH, viscos-
ity and coagulation activity due to microbial decomposition of
organic compounds during storage a fresh solution was prepared
for each sequence of experiments [36]. The few quality parameters
of MOC coagulant were determined using standard methods [37].
Table 1 shows a typical analysis of MOC. The aluminum sulphate
[Al2 (SO4 )3 ·18H2 O] used in the present study was of laboratory
grade. The coagulation experiments with alum were performed
only for comparison purposes. A 4% solution of alum in deionized
water was used (4 g of alum in 100 ml water). The alum powder was
totally soluble in the water. A fresh solution was prepared every day
for reliable results.
2.2. Coagulation studies
The effluent sample collected from a distillery unit near Tiruchi
city was diluted to 25% of original concentration using deionized
water for further studies. The adjustment of pH was carried using
0.1N H2 SO4 or NaOH solutions as required.
The coagulation studies were performed using Jar test appara-
tus which allowed for six 1 L beakers to be agitated simultaneously
and rotational speed could be varied between 0 and 100 rota-
tions per minute (RPM). The beakers were filled with 500 ml spent
wash sample. During rapid mixing at 100 RPM for 2 min coagulant
dosage was added into each beaker and was followed by slow mix-
ing at 40 RPM for 30 min. The duration of sedimentation was kept
constant at 30 min [38]. The supernatant after sedimentation was
filtered using Whatman no. 42 filter paper. The filtrate was ana-
lyzed for absorbance using UV–vis double beam spectrophotometer
(Systronics 2201) at a maximum wavelength 475 nm [39]. Color
removal efficiency was measured as a decrease in optical density
measurement at 475 nm. The readings were taken in triplicate for
each individual solution to check repeatability. The coefficient of
variation obtained was not more than 4% for the three determina-
tions. The characteristics of raw distillery spent wash are given in
Table 2.
2.3. Response surface methodology and optimization of
parameters
Response surface methodology (RSM) is an empirical quadratic
modeling technique that provides the relationship between a set of
controllable experimental factors and observed results. This tech-
Table 2
Physico-chemical characteristics of raw distillery spent wash (10% diluted).
Parameters Magnitude
pH 4.2–4.3
Temperature (◦ C) 30
Color Dark brown
Odor Burnt sugar
Chemical oxygen demand (mg l−1 ) 10,000–11,000
Biochemical oxygen demand (mg l−1 ) 7000–7500
Total dissolved solids (mg l−1 ) 5500–5700
Chloride (mg l−1 ) 500–600
Potassium (mg l−1 ) 1000–1300
Calcium (mg l−1 ) 210–300
806 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811

Fig. 2. pH variation of distillery spent wash treated with MOC (initial pH 7,

Fig. 1. Effect of storage duration and temperature of MOC stock solution extracted
dosage = 60 ml, concentration = 0.25 M).
with 0.25 M NaCl and KCl salts on its coagulation efficiency (initial pH 7,
dosage = 60 ml).

nique is employed to study the interactions among various factors

and their corresponding responses. It is also used to optimize all
affecting parameters collectively by statistical experimental design
[40,41]. Dosage (X1 ), pH (X2 ) and concentration of salt (X3 ) were
selected as independent variable and efficiency of color removal
(Y) as response variable. The behavior of the system is explained by
following quadratic equation [42,43].
  
Y = b0 + bi Xi + bii Xi2 + bij Xi Xj

The results of experimental design were studied and interpreted by Fig. 3. Effect of MOC extracted using various salts (dosage = 60 ml, pH 7, concentra-
MINITAB 14 statistical software to estimate the response of depen- tion = 0.25 M).
dent variable.
3.3. Effect of salt solution on coagulation activity

3. Results and discussion
3.1. Effect of storage duration and temperature of MOC stock
solution on its coagulation efficiency
The MOC stock solution were divided into two groups and stored
at two different temperatures namely; room temperature (30 ◦ C)
and under refrigeration (4 ◦ C). The effects of storage duration on
MOC stock solution were investigated for 1, 2, 3, 4, 5 and 6 days. Fig. 1
shows the results of color removal using M. oleifera stock solutions
kept at different temperatures for different durations as mentioned
above. Coagulation with M. oleifera stock solutions, which were kept
at room temperature for duration of up to 3 days, is most effective on
color removal of distillery spent wash and beyond 3 days the coag-
ulation efficiency of MOC decreased drastically. At 4 ◦ C, the active
agent in the coagulation with M. oleifera stock solution is effective
up to 5 days and beyond it coagulation efficiency dropped.
The active components in M. oleifera seeds were extracted using
various salt solutions of 0.25 M strength to analyze its coagula-
tion activity. The studies were done using fresh MOC stock solution
extracted at room temperature. Fig. 3 shows the effect of NaCl, KCl,
NH4 Cl, NaNO3 and KNO3 salt solutions extracted MOC on color
removal of spent wash. The bar chart indicates NaCl and KCl salt
solutions extracted MOC is more effective than other salt solutions.
Hence the further studies were done for the two salts to get optimal
solution for color removal. Fig. 4 shows the effect of NaCl and KCl salt
solutions (0.25 M) extracted MOC on color removal of spent wash at
different pH. The color removal increased as dosage increased due
to increased availability of active components for coagulation pro-
cess. The maximum color removal of 75% and 71% were obtained
for 0.25 M NaCl and KCl salt solutions at pH 7 respectively. The
comparative results of color removal with alum are shown in Fig. 5.

3.2. Effect of pH during coagulation

M. olifera did not significantly affect the pH of treated spent wash

samples. The variation of pH values of spent wash during the pro-
cess of coagulation was less significant. Fig. 2 shows the variation
of pH after coagulation for MOC stock solution stored for different
days at room temperature (30 ◦ C) and under refrigeration (4 ◦ C). A
slight decrease in pH of wastewater sample after coagulation may
be due to hydrogen ions of weak acidity of MOC stock solution that
balanced the hydroxide ions in the wastewater. This result is in
agreement that M. oleifera coagulants do not change the pH of the
water being treated [44]. Fig. 4. Effect of 0.25 M NaCl and 0.25 M KCl salt solutions extracted MOC.
 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811 807

Table 3
Experimental range and levels of independent process variables.

Independent variables Low level, −1 High level, +1

pH 4 9
Dosage (ml) 20 60
Concentration of salt (M) 0.25 1

extract active component of MOC. In order to study the com-

Fig. 5. Effect of alum on color removal of distillery spent wash.
3.4. Main effect plots and contour plots for color removal
The most important parameters that affect the coagulation pro-
cess are dosage, pH and concentration of salt solution used to
bined effect of these factors experiments were conducted at
different combination of physical parameters. The range of the
variables studied was given in Table 3.The main effects plot
for NaCl and KCl are shown in Figs. 6 and 7 respectively. It
was observed from the figure that as dosage increases color
removal efficiency increases, which indicate that increase in active
component in dosage, correspondingly increased the coagulation
efficiency.
The contour plots for NaCl and KCl are shown in Figs. 8 and 9
respectively indicate efficiency of color removal as a function of
various variables. The maximum color removal of above 70% is

Fig. 6. Main effects plot for % color removal using NaCl.

Fig. 7. Main effects plot for % color removal using KCl.

808 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811

Fig. 8. Contour plots of % color removal using NaCl. Fig. 9. Contour plots of % color removal using KCl.

3.5. Interaction plots for color removal

obtained in region of maximum dosage of 60 ml and pH of 7 as
shown in pH and dosage plot for both the salts. At higher dosage
above 50 ml, concentration has constant effect on color removal
over the entire region from 0.25 to 1 M for NaCl. This result sig-
nificantly ruled out any further significant rise in color removal by
increasing the concentration. It may be due to salting-out effect at
higher concentration of salt solution.
Apart from linear effect RSM is also useful to study the interac-
tion effects among various parameters, this analysis was performed
using Fischer’s ‘F’ test and Students ‘t’ test. The Students ‘t’ test
was used to determine the regression coefficients of the param-
eters. The ‘P’ values are used as a tool to check the interaction
among various variables [45,46]. The uncoded values of experi-
ment and the corresponding color removal along with fits and

Table 4
Experimental data, fits and residuals.

Run Dosage (ml) (X1 ) pH (X2 ) Concentration % Color removal % Color removal Fit for Residuals Fit for Residuals
(M) (X3 ) NaCl (Y) KCl (Y) NaCl for NaCl KCl for KCl

1 20 4 1 40.452 15.121 45.013 −5.013 14.379 0.621

2 30 4 1 48.262 25.358 50.562 −2.562 21.240 3.760
3 40 4 1 56.428 28.670 54.134 1.866 27.888 0.112
4 50 4 1 57.192 38.385 55.730 1.270 34.321 3.679
5 60 4 1 61.165 42.459 55.350 5.650 40.540 1.460
6 20 7 1 61.174 46.440 53.600 7.400 47.037 −1.037
7 30 7 1 62.440 48.193 60.779 1.221 51.940 −3.940
8 40 7 1 62.385 51.165 65.981 −3.981 56.630 −5.630
9 50 7 1 65.541 56.248 69.208 −4.206 61.105 −5.105
10 60 7 1 67.312 57.422 70.458 −3.458 65.366 −8.366
11 20 9 1 39.220 34.239 34.658 4.342 35.142 −1.142
12 30 9 1 39.322 39.572 42.924 −3.924 3.740 0.260
13 40 9 1 48.475 49.606 49.213 −1.213 42.125 6.875
14 50 9 1 52.321 50.422 53.526 −1.526 45.295 4.705
15 60 9 1 60.431 52.477 55.863 4.137 48.251 3.749
16 20 4 0.25 19.220 10.324 18.097 0.903 11.898 −1.898
17 30 4 0.25 37.229 13.425 30.346 6.654 17.926 −4.926
18 40 4 0.25 37.358 21.285 40.618 −3.613 23.741 −2.741
19 50 4 0.25 46.257 27.544 48.914 −2.914 29.340 −2.340
20 60 4 0.25 53.477 37.582 55.234 −2.234 34.726 2.274
21 20 7 0.25 36.101 65.223 33.790 2.210 57.472 7.528
22 30 7 0.25 37.184 68.073 47.668 −10.668 61.542 6.458
23 40 7 0.25 71.606 71.321 59.571 11.429 65.398 5.602
24 50 7 0.25 72.459 72.138 69.497 2.503 69.040 2.960
25 60 7 0.25 75.380 74.165 77.447 −2.447 72.468 1.532
26 20 9 0.25 10.220 51.229 19.584 −9.584 54.187 −3.187
27 30 9 0.25 43.459 53.223 34.550 8.450 56.953 −3.953
28 40 9 0.25 44.138 57.587 47.539 −3.539 59.504 −2.504
29 50 9 0.25 63.697 58.371 58.553 4.447 61.840 −3.840
30 60 9 0.25 66.138 63.220 67.590 −1.590 63.963 −0.963
 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811 809

Fig. 10. Interaction plot of % color removal for NaCl.

Table 5 Table 6
Coefficients, t, P and standard deviation for NaCl. Coefficients, t, P and standard deviation for KCl.

Term Coefficient Standard t P Term Coefficient Standard t P

deviation deviation

Constant 63.2929 2.389 26.489 0 Constant 59.356 1.9303 30.75 0

Dosage (ml) 14.5855 1.555 9.378 0 Dosage (ml) 8.9842 1.2564 7.151 0
pH 0.5 1.343 0.372 0.713 pH 12.5 1.0845 11.526 0
Normality (N) 3.7974 1.1 3.453 0.002 Normality (N) −3.3079 0.8884 −3.723 0.001
Dosage × dosage −3.9524 2.62 −1.508 0.146 Dosage × dosage −0.4286 2.1168 −0.202 0.842
pH × pH −15.4167 2.438 −6.323 0 pH × pH −21.0417 1.9698 −10.682 0
Dosage × pH 2.7171 1.886 1.441 0.164 Dosage × pH −3.2632 1.5236 −2.142 0.044
Dosage × normality −6.7 1.55 −4.322 0 Dosage × normality 0.8333 1.2523 0.665 0.513
pH × normality −2.9605 1.334 −2.22 0.038 pH × normality −5.3816 1.0774 −4.995 0

R-Sq = 0.89. R-Sq = 0.947.

Fig. 11. Interaction plot of % color removal for KCl.

810 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811

Table 7
ANOVA for color removal using NaCl.

Source Degree of freedom Sequential sum of squares Adjusted sum of squares Adjusted mean of squares F P

Regression 8 6152.49 6152.49 769.06 21.34 0

Linear 3 3703.63 3604.98 1201.66 33.34 0
Square 2 1523.06 1523.06 761.53 21.13 0
Interaction 3 925.79 925.79 308.6 8.56 0.001
Residual error 21 756.96 756.98 36.05

Total 29 6909.47

Table 8
ANOVA for color removal using KCl.

Source Degree of freedom Sequential sum of squares Adjusted sum of squares Adjusted sum of squares F P

Regression 8 8822.65 8822.65 1102.83 46.88 0

Linear 3 5431.94 4653.95 1551.32 65.94 0
Square 2 2685.44 2685.44 1342.72 57.08 0
Interaction 3 705.27 705.27 235.09 9.99 0
Residual error 21 494.02 494.02 23.52

Total 29 9316.67

residuals predicted are shown in Table 4. The regression coef-
ficients, ‘t’ values and ‘P’ values for the analysis is given in
Tables 5 and 6.
Regression models for efficiency of color removal in spent wash
based on RSM for NaCl and KCl extracted coagulant given in follow-
ing equations:
YNaCl = (63.2929) + (14.5855X1 ) + (0.5X2 ) + (3.7974 X3 )
color removal using NaCl and KCl salts respectively. The optimal
experiments performed in triplicate at above obtained optimal con-
ditions provided 53% and 64% color removal for MOC extracted
using NaCl and KCl salt solutions respectively. This is close to the
predicted response at optimal conditions.
4. Conclusion

− (3.9524X12 ) − (15.4167X22 ) + (2.7171X1 X2 )
− (6.7X1 X3 ) − (2.9605X2 X3 )
YKCl = (59.356) + (8.9842X1 ) + (12.5X2 ) − (3.3079X3 )
− (0.4286X12 ) − (21.0417X22 ) − (3.2632X1 X2 )
+ (0.8333X1 X3 ) − (5.3816X2 X3 )
The interaction plot for NaCl and KCl for all variables is shown in
Figs. 10 and 11 respectively. The concentration of salt shows pos-
itive effect in NaCl (+3.7974) and negative effect in KCl (−3.3079).
This contrast result may be due to salting-out mechanism being too
strong with KCl in comparison to NaCl. Hence color removal reaches
maximum at 0.25 M KCl and gradually decreases as concentration
of salt increases to 1 M KCl. The decolorization at different pH pro-
vides that the removal is maximum at pH 7 in comparison to pH 4
and 9 as shown in Figs. 10 and 11. The P values of pH–dosage inter-
actions were 0.044, which shows the consistency of the result. The
decolorization increases with dosage at all pH ranges studied.
3.6. Analysis of variance (ANOVA) test
The statistical significance of ratio of mean square due to regres-
sion and mean square residual error was tested. ANOVA is a
statistical technique that subdivides the total variation in a set of
data into component parts associated with specific sources of vari-
ation for the purpose of testing hypotheses on the parameter of
the model [47]. The ‘P’ values obtained are less indicate the factors
play a significant role and the model is statistically significant [48].
The ANOVA results for NaCl and KCl is shown in Tables 7 and 8
respectively.
Optimization of color removal was done for target value of 80%
color removal using response optimization process. The dosage (20
and 60 ml), pH (7 and 8.5) and concentration (1 and 0.25 M) had
been found to be optimum conditions for maximum 56% and 67%
The present study clearly demonstrates the existence of good
scope for MOC in decolorization of distillery spent wash. The color
removal of distillery spent wash is complex and requires sequential
treatment using various techniques like adsorption, electrocoagu-
lation and ozonation. In this sequence MOC can be introduced as
natural coagulant to remove part of color from spent wash. This
study clearly shows that response surface methodology was one of
the suitable methods to optimize the best operating conditions for
target value of color removal. Satisfactory predicted equations were
developed for both the salts using RSM to optimize the parameters.
References
[1] B.L. Wedzicha, M.T. Kaputo, Melanoidins from glucose and glycine: composi-
tion, characteristics and reactivity towards sulphite ion, Food Chem. 43 (1992)
359–367.
[2] V. Kumar, L. Wati, P. Nigam, I.M. Banat, B.S. Yadav, D. Singh, R. Marchant, Decol-
orization and biodegradation of anaerobically digested sugarcane molasses
spent was effluent from biomethanation plants by white rot fungi, Process
Biochem. 33 (1998) 83–88.
[3] P. Manisankar, C. Rani, S. Viswanathan, Effects of halides in the electro-
chemical treatment of distillery effluent, Chemosphere 57 (8) (2004) 961–
966.
[4] B. Kannabiran, A. Prakasam, Effect of distillery effluent on seed germination,
seedling growth and pigment content of Bigna Mungo Hepper (C.V.T.9), Geobios
20 (1993) 108–112.
[5] C.S. Agarwal, G.S. Pandey, Soil pollution by spent wash discharge: depletion
of manganese (II) and impairment of its oxidation, J. Environ. Biol. 15 (1994)
49–53.
[6] F.J. FitzGibbon, P. Nigam, D. Singh, R. Marchant, Biological treatment of distillery
waste for pollution remediation, J. Basic Microbiol. 35 (1995) 293–301.
[7] D. Francisca Kalavathi, L. Uma, G. Subramanian, Degradation and metab-
olization of the pigment-melanoidin in distillery effluent by the marine
cyanobacterium Oscillatoria boryana BDU 92181, Enzyme Microb. Technol. 29
(2001) 246–251.
[8] S. Ohmomo, Y. Kancko, S. Sirianuntapiboon, P. Somchai, P. Atthasampunna, I.
Nakamura, Decolorization of molasses waste water by a thermophilic strain,
Aspergillus fumigatus G-2-6, Agric. Biol. Chem. 51 (1987) 3339–3346.
[9] V. Kumar, L. Wait, F. Fitzgibbon, P. Nigam, I.M. Bansat, D. Singh, R. Marchant,
Bioremediation and decolorization of anaerobically digested distillery spent
wash, Biotechnol. Lett. 19 (1997) 285–289.
 R.K. Prasad / Journal of Hazardous Materials 165 (2009) 804–811
[10] A.P. Thakkar, V.S. Dhamankar, B.P. Kapadnis, Biocatalytic decolourisation
of molasses by Phanerochaete chrysosporium, Bioresour. Technol. 97 (2006)
1377–1381.
[11] G. Christoskovas, L.D. Lazarov, Electrochemical method for purification and dis-
coloration of cellulose paper industry wastewaters, Environ. Prot. Eng. 14 (3–4)
(1988) 69–76.
[12] P. Manisankar, S. Viswanathan, C. Rani, Electrochemical treatment of distillery
effluent using catalytic anodes, Green Chem. 5 (2003) 270–274.
[13] P.K. Chaudhari, I.M. Mishra, S. Chand, Decolorization and removal of chemical
oxygen demand (COD) with energy recovery: treatment of biodigester effluent
of a molasses-based alcohol distillery using inorganic coagulants, Colloids Surf.
A: Physicochem. Eng. Aspects 296 (2007) 238–247.
[14] V.P. Migo, M. Matsumure, E.J.D. Rosarto, H. Kataka, Decolouration of molasses
wastewater using an inorganic coagulates, J. Ferment. Bioeng. 75 (6) (1993)
438–442.
[15] S. Sinha, Y. Yoon, G. Amy, J. Yoon, Determining the effectiveness of conven-
tional and alternative coagulants through effective characterization schemes,
Chemosphere 57 (2004) 1115–1122.
[16] J.E. Van Benschoten, J.K. Edzwald, Measuring aluminum during water treat-
ments, J. Am. Water Works Assoc. 82 (1990) 71–78.
[17] Z. Song, C.J. Williams, R.G.J. Edyvean, Treatment of tannery wastewater by
chemical coagulation, Desalination 164 (2004) 249–259.
[18] R.G. Miller, F.C. Kopfler, K.C. Kelty, J.A. Stober, N.S. Ulmer, The occurrence of
aluminum in drinking water, J. Am. Water Works Assoc. 76 (1984) 84–91.
[19] F.B. Dilek, S. Bese, Treatment of pulping effluents by using alum and clay—color
removal and sludge characteristics, Water S.A. 27 (3.) (2001).
[20] E. Klimiuk, U. Filipkowska, A. Korzeniowska, Effects of pH and coagulant dosage
on effectiveness of coagulation of reactive dyes from model wastewater by poly
aluminum chloride, Pol. J. Environ. Stud. 8 (2) (1999) 73–79.
[21] S.A.A. Jahn, Traditional water purification in developing countries: exist-
ing methods and potential application, Deutsche Gesellschaft Filr Technische
Zusammenerabeit (GTZ) Manual, vol. 117, 1981, Eschborn.
[22] S.A.A. Jahn, D. Hamid, Studies on natural water coagulant in Sudan with special
reference to Moringa oleifera seeds, Water S.A. 5 (1979) 90–97.
[23] T. okuda, A.U. Baes, W. nishijima, M. okada, Isolation and characterization of
coagulant extracted from Moringa olifera seed by salt solution, Water Res. 35
(2) (2001) 405–410.
[24] A. Diaz, N. Rincon, A. Escorihuela, N. Fernandez, E. Chachin, C.F. Forster, A pre-
liminary evaluation of turbidity removal by natural coagulants indigenous to
Venezuela, Process Biochem. 35 (1999) 391–395.
[25] C.R. Shultz, D.A. Okun, Treating surface waters for communities in developing
countries, J Am. Water Works Assoc. 75 (1983) 212–219.
[26] A. Ademiluyi, Sludge conditioning with Moringa seeds, Environ. Int. 14 (1988)
59–63.
[27] A. Ndabigengesere, K.S. Narasiah, B.G. Talbot, Active agents and mechanism
of coagulation of turbid waters using Moringa oleifera, Water Res. 29 (1995)
703–710.
[28] A. Ndabigengesere, S. Narasiah, Quality of water treated by coagulation using
Moringa olifera seeds, Water Res. 32 (3) (1998) 781–791.
[29] T. Okuda, A.U. Baes, W.T. Nishijima, M. Okada, Coagulation mechanism of salt
solution—extracted active component in Moringa olifera seeds, Water Res. 35
(3) (2001) 830–834.
[30] S.A. Muyibi, L.M. Evison, Moringa oleifera seeds for softening hard water, Water
Res. 29 (4) (1994) 1099–1105.
811
[31] J.P. Sutherland, G.K. Folkard, M.A. Mtawali, W.D. Grant, Moringa oleifera as
natural coagulant, in: Pickford, et al. (Eds.), Affordable Water Supply & San-
itation: Proceedings of the 20th WEDC Conference, Colombo, Sri Lanka,
August 22–26, Intermediate Technology Publications, 1994, ISBN 0906055423,
pp. 297–299.
[32] W.O.K. Grabow, J.L. Slabert, W.S.G. Morgan, S.A.A. Jahn, Toxicity and mutagenic-
ity evaluation of water coagulated with Moringa oleifera seed preparations
using fish, protozoan, bacterial, enzyme and Ames Salmonella assays, Water
S.A. 11 (1985) 9–14.
[33] M.R. Berger, M. Habs, S.A.A. Jahn, D. Schmahl, Toxicological assessment of seeds
from Moringa oleifera and Moringa stenopetala, two highly efficient primary
coagulants for domestic water treatment of tropical waters, East Afr. Med. J. 61
(1984) 712–717.
[34] I.M. Villasenor, C.Y. Lim-Sylianco, F. Dayrit, Mutagens from roasted seeds of
Moringa oleifera, Mutat. Res. 224 (1989) 209–212.
[35] R.F. Packham, Some studies of the coagulation of dispersed clays with hydrolyz-
ing salts, J. Colloid Interface Sci. 20 (1965) 81.
[36] S.A.A. Jahn, Using Moringa seeds as coagulants in developing countries, J. Am.
Water Works Assoc. 90 (1988) 43–50.
[37] APHA, Standard Methods for the Examination of Water and Wastewater, 20th
ed., American Public Health Association, American Water Works Association
and Water Pollution Control Federation, Washington, DC, 1998.
[38] S. Kawamura, Effectiveness of natural polyelectrolyte in water treatment, J. Am.
Water Works Assoc. 83 (1991) 88–91.
[39] C. Raghukumar, G. Rivonkar, Decolorization of molasses spent wash by the
white rot fungus Flavodon flavus, isolated from a marine habitat, Appl. Micro-
biol. Biotechnol. 55 (2001) 510–514.
[40] M. Elibol, Response surface methodological approach for inclusion of perfluro-
carbon in actinohordin fermentation medium, Process Biochem. 38 (2002)
667–773.
[41] K. Ravikumar, S. Krishnan, S. Ramalingam, K. Balu, Optimization of process vari-
ables by application of response surface methodology for dye removal using a
novel adsorbent, Dyes Pigments 72 (1) (2007) 66–74.
[42] G. Annadurai, R.Y. Sheeja, T. Mathalai Balan, V.R. Murugesan, Srinivasamoorthy,
Factorial design of experiments in the determination of adsorption equilibrium
constants for basic methylene blue using biopolymer, Bioprocess Eng. 20 (1999)
37–43.
[43] G. Annadurai, Design of optimum response surface experiments for adsorption
of direct dye on chitosan, Bioprocess Eng. 23 (2000) 451–455.
[44] A. Olsen, Low technology water purification by bentonite clay and Moringa
oleifera seed flocculation as performed in Sudanese village: effects on Schisto-
soma mansoni cercariae, Water Res. 21 (1987) 517–522.
[45] Lawson, Erjavec, Modern Statistics for Engineering and Quality Improvement,
1st ed., Thomson Duxbury, Singapore, 2002.
[46] M.M.D. Zulkali, A.L. Ahmad, N.H. Norulakmal, Oryza sativa L. husk as heavy
metal adsorbent: optimization with lead as model solution, Bioresour. Technol.
97 (2006) 21–25.
[47] K. Ravikumar, S. Ramalingam, S. Krishnan, K. Balu, Application of response
surface methodology to optimize the process variables for reactive red and
acid brown dye removal using a novel adsorbent, Dyes Pigments 70 (1) (2006)
18–26.
[48] H.M. Kim, J.G. Kim, J.D. Cho, J.W. Hong, Optimization and characterization of UV-
curable adhesives for optical communication by response surface methodology,
Polym. Test. 22 (2003) 899–906.