Biotechnol. Prog. 2005, 21, 860−867
860
Adsorption-Desorption Process Using Wood-Based Activated
Carbon for Recovery of Biosurfactant from Fermented Distillery
Wastewater
Kirti V. Dubey, Asha A. Juwarkar,* and S. K. Singh
Environmental Biotechnology Division, National Environmental Engineering Research Institute (NEERI),
Nehru Marg, Nagpur 440 020, India
Methods used for biosurfactant recovery include solvent extraction, precipitation,
crystallization, centrifugation and foam fractionation. These methods cannot be used
when distillery wastewater (DW) is used as the nutrient medium for biosurfactant
production by Pseudomonas aeruginosa strain BS2, because recovery of biosurfactant
by any of these methods imparts color to the biosurfactant. The biosurfactant has a
nonaesthetic appearance with lowered surface active properties. These methods cannot
be used for continuous recovery of biosurfactant during cultivation. Hence, a new
downstream technique for biosurfactant recovery from fermented DW comprised of
adsorption-desorption processes using wood-based activated carbon (WAC) was
developed. This study involves batch experiments to standardize the factors affecting
the rate of biosurfactant adsorption onto WAC. WAC was the most efficient adsorbent
among various ones tested (i.e., silica gel, activated alumina and zeolite). The WAC
(1% w v-1), equilibrium time (90 min), pH range of 5-10 and temperature of 40 °C
were optimum to achieve 99.5% adsorption efficiency. Adsorption kinetics and
intraparticle diffusion studies revealed the involvement of both boundary layer
diffusion and intraparticle diffusion. The Langmuir adsorption isotherm of WAC
indicated the formation of a monolayer coverage of the biosurfactant over a homogeneous carbon surface, while the Freundlich isotherm showed high adsorption at strong
solute concentrations and low adsorption at dilute solute concentrations. WAC
concentration of 4% w v-1 facilitated complete removal of the biosurfactant from
collapsed foam (contained 5-fold higher concentration of biosurfactant than was present
in fermented DW). Biosurfactant adsorption was of chemisorption type. Acetone (polar
solvent) was a specific viable eluant screened among various ones tested because it
selectively facilitated maximum recovery, i.e., 89% biosurfactant from WAC. By acetone
treatment, complete regeneration of WAC was feasible and WAC can be reused for
biosurfactant recovery up to 3 cycles. The recovered biosurfactant showed improved
surface-active property (i.e., much lower critical micelle concentration value of 0.013
verses 0.028 mg mL-1 for biosurfactant recovered by classical methods). The reuse
potential of WAC was assessed and results suggest that the carbon can be reused for
three consecutive cycles for biosurfactant adsorption from fermented wastewater
without any decrease in adsorption efficiency. Thus, this process forms a basis for
continuous recovery of biosurfactant from fermented DW and concentrated foam. This
process reduces the use of high cost solvent, avoids end product inhibition and
minimizes product degradation.
Introduction
Biosurfactants are beginning to be accepted as potential performance-effective molecules that are ecofriendly
alternatives to synthetic surfactants. Potential commercial applications of biosurfactants have been widely
reviewed (1-4). Process development for large-scale
production of biosurfactant is necessary to minimize the
cost of raw material and processing. Economic strategies,
which emphasize the utilization of waste streams as nocost substrate and low cost in situ methods for recovering
biosurfactant are essential for developing large-scale
biosurfactant production technology.
* To whom correspondence should be addressed. Ph: 91-07122249764. E-mail: aajuwarkar@rediffmail.com.
10.1021/bp040012e CCC: $30.25
The search for cheaper raw materials for biosurfactant
production has led to industrial wastes utilization (5).
We have reported that biosurfactant production from
distillery and whey wastewaters and synthetic medium
was comparable (6-8). Kinetic studies of biosurfactant
production from distillery and whey wastes and nutrient
depletion studies have shown that distillery and whey
wastewaters could be used as complete nutrient medium
for biosurfactant production by Pseudomonas aeruginosa
strain BS2.
Most fermentation products are released into dilute
aqueous solutions and in many cases, the downstream
process comprises approximately 60% of the total cost of
the product (9). The methods used for biosurfactant
recovery include solvent extraction, precipitation, crystal-
© 2005 American Chemical Society and American Institute of Chemical Engineers
Published on Web 02/22/2005
Biotechnol. Prog., 2005, Vol. 21, No. 3
lization, centrifugation and foam fractionation, which
cannot be used for continuous product recovery during
cultivation (5). Besides these methods, there are “in situ
recovery” methods for biosurfactant recovery where
products are continuously removed from the culture broth
during cultivation. In situ methods for product recovery
can avoid end-product inhibition during fermentation
(10). Some examples of in situ biosurfactant recovery are
adsorption of the surfactant onto ion-exchange resins or
other suitable adsorbents as reported for lipopeptide
surfactant produced by Candida petrophilum (11) or
rhamnolipids produced by Pseudomonas sp. DSM 2874
(12). Due to their lipophilic ability, rhamnolipids could
be adsorbed onto a support (XAD-2) and recovered by
aqueous buffer systems (13). However, rhamnolipid
recovery from fermented DW cannot use an ion exchanger for recovery because the aqueous buffer system
removes water-soluble impurities (i.e., color and other
metabolites) present in fermented DW along with the
biosurfactant. The black color of DW imparts color to the
biosurfactant and lowers its surface-active efficiency of
biosurfactant due to an increase in critical micelle
concentration (CMC) of biosurfactant. CMC of biosurfactant extracted from fermented DW was higher, i.e., 0.028
mg mL-1 as compared to CMC of biosurfactant extracted
from fermented minimal salts medium, i.e., 0.021 mg
mL-1 using diethyl ether extraction method (7, 14). It is
reported that surface active efficiency of synthetic surfactants and biosurfactants is inversely proportional to
their CMC values (5). Color impurities make the biosurfactant impure, which is not desirable in industries (i.e.,
cosmetics, food and pharmaceutical) where high purity
specifications exist.
This study investigated the recovery of color-free
biosurfactant produced by P. aeruginosa strain BS2 in
batch experiments mediated by an adsorption and desorption process. This process involved adsorption of the
biosurfactant from fermented DW onto WAC and the
selective desorption of biosurfactant by acetone. Various
parameters affecting the adsorption onto WAC were
investigated. The usefulness of WAC for adsorption of
biosurfactant from DW was demonstrated with Langmuir
and Freundlich isotherms. Feasibility of biosurfactant
removal from collapsed foam was demonstrated. The
suitability of 3 times recycled WAC was also assessed
for efficient recovery of biosurfactant using a combination
of adsorption-desorption processes.
Materials and Methods
Materials and Reagents. Distillery spent wash used
as a complete nutrient medium for biosurfactant production was collected from a distillery unit (M/s Jubilant
Organosys Limited, Noida, New Delhi). Silica gel, activated alumina, wood-based activated carbon (WAC) and
zeolite were purchased from Sisco Research Laboratory
(SRL) Private Limited, Mumbai, India. Analytical grade
reagents and chemicals used in various analyses were
purchased from Qualigens Fine Chemicals Private Limited (Mumbai, India).
Cultivation of P. aeruginosa Strain BS2 for Production of Biosurfactant from Distillery Wastewater. Pseudomonas aeruginosa strain BS2 is a potent
biosurfactant-producer isolated at National Environmental Engineering Research Institute (NEERI), in Nagpur,
India (7, 14). Batch fermentation was carried out in New
Brunswick BIO FLO IIC fermenter using distillery spent
wash diluted with tap water (1:3) as nutrient medium
for bacterial growth and biosurfactant production at 37
°C, agitation at 400 rpm, 96 h incubation, aeration at 2
861
SLPM (standard liquid pressure per minute) with 1.5
VVP (volume of air per volume liquid). After 96 h, the
fermented DW was centrifuged at 8,000 × g and filtered
through 0.45 µm membrane filter and 0.2% (v v-1)
formaldehyde was added to prevent microbial growth.
The fermented DW used was characterized for pH,
surface tension (ST), Fcmc, i.e., the factor of diluting the
fermented cell free DW necessary to reach CMC and
biosurfactant yield. In another set of batch fermentations,
biosurfactant generated in the form of foam was removed
by foam fractionation (15) processed and characterized
the same as fermented DW. Adsorption studies were
conducted for both fermented DW and foam-fractionated
biosurfactant.
Screening of Adsorbents. Silica gel, activated alumina, wood-based activated carbon (WAC) and zeolite at
1% w v-1 concentration were added individually to flasks
containing 100 mL of cell-free fermented DW. Flasks
were shaken at 150 rpm and 30 °C for 24 h. The contents
were filtered through Whatman filter paper No. 42 and
the filtrates were analyzed for ST and Fcmc. The percent
adsorption of biosurfactant onto each adsorbent was
determined from the standard graph of Fcmc versus
biosurfactant concentration (14). Based on the highest
adsorption capacity, WAC was selected for further batch
studies.
Batch Studies. The effect of contact time (0-90 min),
carbon concentration (0-4% w v-1), temperature (30, 40
and 50 °C) and pH (3-10) on biosurfactant adsorption
was determined under the experimental conditions described in Figures 1-5.
A suitable concentration of the carbon for complete
biosurfactant removal from the collapsed foam was
determined using WAC in the range of 1-6% w v-1 (see
experimental conditions described in Figure 6).
Adsorption Isotherm. The adsorption isotherm studies were conducted with biosurfactant ranging from
0.001% to 0.04% w v-1, while maintaining 1% w v-1 WAC.
After an equilibrium time of 90 min, the WAC was
separated from the wastewater by filtration and the
filtrate was analyzed for residual biosurfactant. The
biosurfactant adsorptive capacity of the WAC was evaluated from the plots of Langmuir and Freundlich isotherms.
Desorption of Biosurfactant from WAC. Different
concentrations of biosurfactant (ranging from 0.001% to
0.04% w v-1) were added to flasks, adsorbed on WAC (1%
w v-1), the WAC was separated by filtration and washed
with distilled water to remove unadsorbed biosurfactant.
Each of WAC samples was suspended in stoppered flasks
containing 20 mL of acetone and agitated at 100 rpm for
6 h at 30 °C. The acetone extract was separated from
the adsorbent, followed by washing of the adsorbent
several times with acetone. Acetone from each extracted
sample, which contained the biosurfactant, was removed
under reduced pressure. The obtained residue was suspended in 100 mL distilled water (adjusted to pH 8.0 with
NaOH) and analyzed for Fcmc value. The amount of
biosurfactant desorbed by acetone at each concentration
of biosurfactant was computed from the standard graph
of Fcmc versus biosurfactant concentration (14).
Reuse Potential Assessment of WAC for Biosurfactant Recovery from Fermented DW. Shake flask
experiments were conducted to assess the reuse potential
of WAC for biosurfactant recovery from fermented DW
using WAC. Optimized sets of conditions were maintained for biosurfactant adsorption (i.e., WAC concentration of 1% w v-1 and equilibrium time of 90 min) and
desorption using 20 mL acetone. The WAC regenerated
Biotechnol. Prog., 2005, Vol. 21, No. 3
862
Table 1. Screening of Different Adsorbents for Adsorption of Biosurfactant from Fermented DW
ST
(mN m-1)
Fcmc
adsorbent (1% w v-1)
biosurfactant adsorbed
(mg g-1)
biosurfactant not adsorbed
(% w v-1)
%
adsorption
nonea
silica gel
wood-based activated carbon (WAC)
zeolite
activated alumina
27.5
27.5
54.0
29.5
28.0
25.0
25.0
0.0
17.5
23.75
0
0
96.0
29.0
5.7
0.096
0.096
0.0
0.067
0.090
0
100.0
30.20
5.90
a
Data represent characteristics of cell - free fermented DW before treatment.
after desorption of biosurfactant by acetone was reused
to recover biosurfactant from fermented DW by using the
above-mentioned adsorption-desorption processes for
three consecutive cycles. Feasibility of WAC regeneration
after each cycle was assessed by dissolving the eluted
biosurfactant in distilled water of pH 8 (pH raised by
addition of 1 N NaOH) and by using ST and Fcmc
measurements of biosurfactant solution (14).
Analytical Methods. Methods for surface tension (ST)
measurement, Fcmc and estimation of biosurfactant concentration in the cell-free fermented wastewater are
described elsewhere (7). The Branauer, Emmett and
Teller (BET) surface area of the WAC was determined
using Micromeritics ASAP-200 Specific Surface Area
Analyzer. The bulk density and average particle pore
diameter of the WAC was determined by using pore
sizers 9320 V2.05 Mercury Intrusion Porosimeter. The
proximate analysis of the WAC with respect to the total
ash, moisture, volatile matter and fixed carbon contents
was determined using the Indian Standard Methods of
test for coal and coke (16). The decolorizing power and
phenol number of the WAC were analyzed by EPA
method 625/1-71-002a (17). All experiments were performed in triplicate and the results were reproducible
within (5% standard deviation.
Results and Discussion
Screening of Adsorbents. Different adsorbents were
screened for their potential to adsorb biosurfactant from
fermented DW, which has high surface active properties
attributable to low ST (27 mN m-1), high Fcmc (value of
25) and biosurfactant yield (960 mg l-1). Table 1 shows
that silica gel, zeolite and activated alumina were not
suitable adsorbents (i.e., marginal increase in ST of
wastewater from initial value of 27 mN m-1 and negligible decrease in Fcmc value despite being given 24 h of
adsorption). Nevertheless, adsorption on zeolite reduced
the Fcmc of DW from 25 to 17.5 and its adsorptive capacity
was 30.20%. In comparison to these adsorbents, WAC
showed high adsorption capacity. After using WAC
treatment, the ST of the wastewater increased to 54.0
mN m-1, which was similar to the ST of sterile nonfermented DW indicating complete adsorption of the biosurfactant on WAC. There was a significant reduction of
Fcmc value from 25 to 0 and there was no residual
concentration of biosurfactant in the waste.
Physicochemical analysis of WAC showed a mesh size
of 1.0 mm and bulk density of 1.02 g mL-1, while the
surface area and particle pore diameter was 423.45 m2
g-1 and 0.0345 µm, respectively. Carbon content was
72.2627%. WAC had high decolorizing power (82%) and
low phenol number (22.0), which imply that WAC is
suitable for organic adsorption (18).
Batch Studies. Effects of various parameters such as
equilibrium time (to determine the adsorption kinetics),
carbon concentration, pH and the effect of temperature
on adsorption of biosurfactant from fermented DW were
studied to determine the suitability of WAC for adsorption of biosurfactant from DW.
(a) Standardization of Equilibrium Time. Variations in percent adsorption of biosurfactant on WAC from
waste, ST and Fcmc values of waste over time are
presented in Figure 1. On using WAC (1% w v-1), ST of
the wastewater increased gradually from 27.5 to 37 mN
m-1 and Fcmc value decreased from 25 to 0 over 90 min.
Equilibrium time of 90 min resulted in 99.5% adsorption
of biosurfactant from the fermented DW.
(b) Adsorption kinetics. To establish the dynamics
of the sorption reaction in terms of the order of the
reaction and the rate constant of the reaction, equilibrium
time experiments were performed, which describe the
rate of uptake of the adsorbate by the adsorbent. The rate
constant for adsorption of the biosurfactant on WAC from
fermented DW was obtained by assuming a first-order
reaction:
log
C
K
t
) log C Ce
2.303
(1)
where C is initial concentration of the biosurfactant (mg
L-1), Ce is equilibrium concentration of the biosurfactant
(mg L-1), t is time (min), and K is rate constant (min-1).
A linear line plot of log(C/Ce) versus t for WAC indicated
a first-order reaction for the sorption process (inset
Figure 2). Value of the rate constant obtained from the
slope of the straight line was 4.582 × 10-2 min-1 for
biosurfactant sorption on WAC. The observed rate constant for biosurfactant adsorption was comparable to
values reported for different types of adsorbates such as
various heavy metals, dyes and recalcitrant compounds
(18).
(c) Intraparticle Diffusion Study. The sorption
reaction may be accomplished either by film diffusion,
pore diffusion or both, and either of these may be rate
limiting depending upon the system conditions. Intraparticle diffusion (pore diffusion) may be a rate limiting
step even in rapidly stirred batch processes. This situation was tested by plotting time (t0.5) of uptake versus
amount of adsorbed biosurfactant. Time-dependent uptake of the biosurfactant by WAC is shown in Figure 2
the initial curved portion is attributed to boundary layer
diffusion effects (19), while the final linear portion is due
to intraparticle diffusion effects (20). An extrapolation
of the axis provides the intercept, which is proportional
to the extent of the boundary layer thickness. The
amount of biosurfactant adsorbed per gram of WAC per
square root of time was 1.50 mg g-1 min-0.5 as obtained
from initial slope of the curve. It is apparent from the
kinetic and intraparticle diffusion studies that the sorption of biosurfactant onto WAC from fermented DW was
complex, involving both boundary layer diffusion and
intraparticle diffusion, which was the rate-controlling
step.
(d) Effect of WAC Concentration on Adsorption.
Carbon concentration required for complete adsorption
of biosurfactant from a known volume of fermented
distillery wastewater was determined. Figure 3A shows
that, at carbon concentration of 1% w v-1, 99.5% removal
Biotechnol. Prog., 2005, Vol. 21, No. 3
863
Figure 1. Standardization of equilibrium time for adsorption of biosurfactant on WAC from fermented DW.
Figure 2. Plot for intraparticle diffusion of biosurfactant for adsorption on WAC.
of biosurfactant from the fermented wastewater was
observed due to lowered value of Fcmc from 25 to 0 and
rise of ST from 27 to 35 mN m-1, which indicated that
WAC was suitable for biosurfactant adsorption from
waste at low carbon concentration. The remaining leftover biosurfactant in traces can be removed by further
increasing either the contact time or the concentration
of WAC, which were avoided to reduce the time and the
cost of biosurfactant recovery process. Hence, carbon
concentration of 1% w v-1 was selected for the pH,
temperature and contact time optimization studies.
However, it was noticed that, although, the Fcmc of
fermented DW has reached to zero, the ST of DW was
still low at 35 mN m-1 at 1% w v-1 concentration of WAC.
This is due to the presence of very small concentration
of biosurfactant left in traces i.e., concentration even
lower than its CMC (0.028 mg L-1) after biosurfactant
adsorption on WAC from fermented DW. At CMC,
rhamnolipid produced by P. aeruginosa strain BS2 gives
minimum ST of 27 mN m-1 (7, 8, 14). It is known that
Fcmc is the indirect measure of biosurfactant concentration present in the liquid medium and is based on the
fact that the surface activity is dependent in the concentration of the surface-active compound. When concentration is below certain level (i.e., CMC) the surface activity
is lost, which is expressed by increasing in surface
tension. This property was used for the estimation of the
concentration of the surface-active compound. Increasing
Fcmc values indicate the increasing concentration of
biosurfactant (21).
On increasing the concentration of WAC further,
biosurfactant was completely removed and consequently
the ST of fermented DW reached to 54 mN m-1 and
equalized to that of sterile DW (nonfermented DW). High
efficiency of WAC was shown in the breakthrough curve
(Figure 3B). Complete removal of biosurfactant from
approximately 500 mL wastewater was observed with 5
g of WAC. However, concentration of the unadsorbed
biosurfactant increased to 60 mg as the volume of
wastewater increased to 1 L.
(e) Effect of Temperature and pH on Adsorption.
Temperature and pH are two factors that govern the rate
of biosurfactant adsorption on WAC. Results presented
in Table 2 show that biosurfactant adsorption on WAC
was not a physical adsorption because there was no
decrease in amount of biosurfactant adsorbed per gram
of carbon (x/m value) when the temperature was increased from 30 to 40 °C. A chemisorption was indicated
by the rise in x/m value as temperature increased from
30 to 50 °C. Chemisorption needs activation energy (22)
and improvement in rates of adsorption on carbon with
increase of temperature may be attributed to increased
molecular movement at higher temperatures (23). This
could allow biosurfactant to enter the carbon pores more
easily and facilitate biosurfactant adsorption at a faster
rate as temperature increases.
Biotechnol. Prog., 2005, Vol. 21, No. 3
864
Figure 3. Standardization of WAC concentration (A) and breakthrough curve (B) for adsorption of biosurfactant from fermented
DW.
Table 2. Biosurfactant Adsorption on WAC from
Fermented DW at Different Temperatures
temp
(°C)
contact time
(min)
biosurfactant adsorbed
(mg g-1)
30
45
90
45
90
45
90
88
92
94
96
96
96
40
50
Figure 4 shows that as pH increased from 3 to 5,
percent adsorption of biosurfactant on WAC from wastewater increased; 99.5% adsorption of biosurfactant occurred over a wide pH range of 5 to 10. Biosurfactant
adsorption from wastewater was observed by a rise in
ST from 27 to 35 mN m-1 and a decrease in Fcmc from 25
to 0. During optimum biosurfactant production, the pH
of DW is 8-9, which allows fermented DW to be directly
subjected to biosurfactant recovery without modifying the
pH of the adsorption medium.
(f) Studies on Adsorption Isotherms of the Biosurfactant. Adsorption isotherm studies were done to
understand the extent and degree of biosurfactant removal by adsorption alone. The Langmuir equation
applied for adsorption equilibrium for WAC is given by
Ce
Ce
1
+
)
qe Qob Qo
(2)
where Ce is the concentration of biosurfactant in fermented DW after equilibrium (mg L-1), qe is the concentration of biosurfactant adsorbed per gram of carbon after
equilibrium (mg g-1), and Qo and b are Langmuir
constants for adsorption in the Langmuir model for WAC
(24). The linear plot of Ce/qe versus Ce indicated that
biosurfactant adsorption follows the Langmuir model for
WAC. Conformation of the experimental data by linear
plotting of Ce/qe versus Ce showed the formation of
monolayer coverage of adsorption of biosurfactant molecules on the surface of WAC (unpublished data). The
values of Qo and b were calculated from the slope and
intercept of the plot (35.9712 mg g-1 and 3.6579 L mg-1,
respectively).
The essential characteristics of the Langmuir isotherm
can be expressed in terms of a dimensionless constant
separation factor or an equilibrium parameter RL, which
is defined as RL ) 1/(1+ bCo), where b is the Langmuir
constant and Co is the initial concentration (25). RL value
for adsorption of biosurfactant on WAC was 2.8469 ×
10-4. The value of RL was found to be 0 < R < 1, which
showed that biosurfactant adsorption was favorable on
WAC at 30 °C and a concentration of 960 mg L-1.
Adsorption of biosurfactant on WAC was found to obey
Freundlich adsorption isotherm, which is represented by
the following equation
( mx ) ) log K + ( n1 ) log C
log
e
(3)
where, Ce is the equilibrium concentration (mg L-1), x/m
is the amount of biosurfactant adsorbed per unit weight
of carbon (mg g-1), and K and n are Freundlich constants,
which indicate the adsorption capacity and intensity,
respectively. Plot of log x/m versus log Ce for WAC was
linear which indicated that the adsorption follows the
Biotechnol. Prog., 2005, Vol. 21, No. 3
865
Figure 4. Effect of pH on adsorption of biosurfactant on WAC from fermented DW.
Figure 5. Standardization of WAC concentration for adsorption of biosurfactant from collapsed foam.
Freundlich isotherm for biosurfactant (unpublished data).
The constants K and n were 9.358 and 1.229, respectively
for WAC. The values of n in order of 1 < n < 10 show
favorable adsorption of biosurfactant on WAC. A low n
value or steep slope with WAC indicated high adsorption
at strong solute concentrations and low adsorption at
dilute solute concentrations (26).
Standardization of WAC Concentration for Adsorption of the Biosurfactant from Collapsed Foam.
Since the Freundlich isotherm study with WAC indicated
a high adsorption at strong solute concentration and low
adsorption at dilute solute concentration, attempts were
made to recover the biosurfactant from collapsed foam
where biosurfactant was concentrated 5-fold by using
WAC. One liter of fermented DW containing biosurfactant (960 mg L-1) generated nearly 200 mL of collapsed
foam in which approximately 960 mg of biosurfactant was
concentrated (Fcmc value of 125). Depending upon the
concentration of biosurfactant, carbon concentration
required for complete adsorption of biosurfactant from
collapsed foam could vary. The percent adsorption of
biosurfactant from collapsed foam with increasing concentration of WAC ranging from 1% to 6% w v-1 was
investigated. Results shown in Figure 5 reveal a linear
increase in percent adsorption of biosurfactant with
increase in WAC dose from 1% to 3% w v-1 and 99.5%
adsorption of biosurfactant at a carbon concentration of
4% w v-1 (evident from the drastic reduction of Fcmc value
from 125 to 0). Thus, 100 mL collapsed foam required 4
g of WAC for efficient removal of biosurfactant.
Table 3. Desorption of Biosurfactant from Wood-Based
Activated Carbon (WAC) Using Acetone
concn of
biosurfactant
adsorbed (mg g-1)
Fcmc
20
40
60
80
100
7.5
12.5
16.25
20
25
concn of
biosurfactant
%
desorbed (% w v-1) Fcmc desorption
0.0178
0.0358
0.0537
0.0716
0.0896
5
10
12.5
17.5
22.5
89.45
89.60
89.61
89.58
89.62
Desorption of Biosurfactant from WAC. After adsorption of biosurfactant onto WAC from fermented DW,
biosurfactant desorption from WAC was studied using
various organic solvents (i.e., ethanol, methanol, acetone
and diethyl ether) and buffered systems at various pH
values ranging from 3 to 12. Acetone selectively removed
biosurfactant from the surface of WAC. Acetone facilitated nearly 89% desorption of biosurfactant for all tested
concentrations of adsorbed biosurfactant (Table 3). The
varying amounts of biosurfactant desorbed from WAC
when dissolved in 100 mL water resulted in lower Fcmc
values as compared to the control because desorption did
not reach 100%. The recovered biosurfactant was free
from the colored impurities of DW, which were initially
encountered with acid precipitation and solvent extraction methods for biosurfactant recovery. The rhamnolipid
biosurfactant (14) was desorbed by acetone, leaving
colored impurities of DW adsorbed to WAC. Biosurfactant
recovered by this adsorption-desorption process showed
an improved surface-active property (i.e., lower CMC
value of 0.013 mg/mL compared to 0.028 mg mL-1 of
Biotechnol. Prog., 2005, Vol. 21, No. 3
866
Figure 6. Schematic representation of the protocol for recovery of biosurfactant from fermented DW.
Table 4. Reuse Efficiency of WAC (1% w v-1) for Biosurfactant Recovery from Fermented DWa
no. of
cycles
ST
(mN m-1)
Fcmc
concn of biosurfactant
adsorbed (mg g-1)
biosurfactant
adsorbed (%)
concn of biosurfactant
desorbed (% wv-1)
% desorption
1
2
3
38
38
38
0
0
0
95.52
95.52
95.52
99.5
99.5
99.5
0.0089
0.0089
0.0089
89
89
89
a
Biosurfactant concentration in the fermented DW was 960 mg L-1.
biosurfactant recovered by diethyl ether extraction
method). Protocol for recovery of color-free biosurfactant
from fermented distillery wastewater through this adsorption-desorption process is shown in Figure 6.
Reuse Potential of WAC for Biosurfactant Recovery from Fermented DW. Results presented in the
Table 4 show that WAC can be reused for biosurfactant
recovery from fermented DW by adsorption-desorption
process up to three consecutive cycles. After each cycle
of biosurfactant adsorption, the ST of fermented DW
increased from 27 mN m-1 to 38 mN m-1 and Fcmc
decreased from 25 to 0. There was 99.5% removal of
biosurfactant from fermented DW which remained constant at 95.52 mg g-1. The percentage desorption of
biosurfactant from reused WAC also remained constant
to 89%. This study demonstrated that acetone treatment
can regenerate WAC, and reuse of same WAC as fresh
adsorbent for biosurfactant recovery was feasible up to
three consecutive cycles without any decrease in biosurfactant adsorption efficiency.
Biotechnol. Prog., 2005, Vol. 21, No. 3
Conclusions
A new downstream technique comprising adsorptiondesorption processes using wood-based activated carbon
(WAC) was developed for the removal of color-free biosurfactant produced from distillery wastewater (a no-cost
complete nutrient medium). Batch mode studies were
conducted to standardize the factors affecting the rate
of biosurfactant adsorption on WAC. The amount of WAC
and equilibrium time was optimized at 1% w v-1 and 90
min, respectively. A pH range of 5-10 and temperature
of 40 °C were optimum for biosurfactant adsorption.
Effective adsorption of biosurfactant took place at a wide
pH range of 5-10, which implies that the fermented DW
(without any pH modification) can directly be used for
biosurfactant recovery. Freundlich isotherm experiments
showed that WAC had high adsorption capacity at strong
solute concentrations and low adsorption at dilute solute
concentrations. A preliminary feasibility study for using
WAC for biosurfactant recovery from cell-free collapsed
foam (biosurfactant was concentrated 5-fold) was done.
Biosurfactant adsorption on WAC was chemisorption in
contrast to a physical interaction, which indicated that
suitable solvent could selectively be used as an eluant
for biosurfactant recovery from WAC instead of an
aqueous buffer system that would also remove watersoluble impurities (i.e., color and other metabolites) along
with biosurfactant. The desorption study showed 89%
recovery of biosurfactant by acetone, which was free of
the colored impurities from DW indicating that acetone
was a suitable eluant for biosurfactant recovery. The
recovered biosurfactant showed improved surface-active
property (i.e., lower critical micelle concentration (CMC
value) of 0.013 mg mL-1 compared to 0.028 mg mL-1 for
biosurfactant recovered by classical method. Reuse potential of WAC was assessed and results suggested that
the same carbon could be reused for three consecutive
cycles for biosurfactant adsorption from fermented DW.
This process forms a basis for continuous recovery of
biosurfactant from fermented DW, as well as from
concentrated foam, through an in situ method that avoids
end product inhibition, reduces the use of high cost
solvent, and minimizes product degradation and minimizes the pollution load of DW.
Acknowledgment
The authors acknowledge the kind support and encouragement extended by Dr. Sukumar Devotta (Director, NEERI, Nagpur). Authors also acknowledge the kind
help of Mr. T. V. Subbarao (Senior Scientist, NEERI,
Nagpur) for editing and reviewing the manuscript.
Instrumental facilities from Jawaharlal Nehru Aluminum Research and Development Center, Nagpur for pore
size, surface area and SEM analyses of activated carbon
is also acknowledged.
Note Added after ASAP Publication. An author
was omitted in the version published ASAP February 22,
2005. The version published June 3, 2005 is correct.
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Accepted for publication November 23, 2004.
BP040012E