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Bioresource Technology 99 (2008) 63606364

Eect of heat on the adsorption capacity of an activated carbon

for decolorizing/deodorizing yellow zein
D.J. Sessa a,*, D.E. Palmquist b
a
Plant Polymer Research, National Center for Agricultural Utilization Research, US Department of Agriculture,
Agricultural Research Service, 1815 N. University Street, Peoria, IL 61604-3902, USA
b
Biometrical Services, National Center for Agricultural Utilization Research, US Department of Agriculture, Agricultural Research Service,
1815 N. University Street, Peoria, IL 61604-3902, USA

Received 21 June 2007; received in revised form 29 November 2007; accepted 29 November 2007
Available online 29 January 2008

Abstract

The Freundlich model was evaluated for use to assess the eect of heat on the adsorption capacity of an activated carbon for decol-
orizing/deodorizing corn zein. Because zein protein and its color/odor components are all adsorbed by activated carbon, a method to
monitor their removal was needed. Yellow color is due to xanthophylls; a contributor to o-odor is diferuloylputrescine. The o-odor
component absorbs ultraviolet (UV) light at about 325 nm and its removal coincides with removal of yellow color. A spectrophotometric
method based on UV absorbances 280 nm for protein and 325 nm for the o-odor component was used to monitor their adsorptions
onto activated carbon. Equilibrium studies were performed over temperature range from 25 to 60 C for zein dissolved in 70% aqueous
ethanol. Runs made at 55 C adsorbed signicantly more of the color/odor components than the protein.
Published by Elsevier Ltd.

Keywords: Activated carbon; Adsorption; Freundlich isotherm; Thermal eects; Zein decolorization/deodorization

1. Introduction
Zein, a class of proteins known as prolamins, is the
major group of proteins in corn as well as corn gluten meal,
a co-product of the ethanol industry generated by wet-mill-
ing corn. Its use in the medical, pharmaceutical, and cos-
metic elds and also in applications as a paper coating,
packaging material and biodegradable chewing gum base
is limited by its inherent yellow color and o-odor. Sessa
et al. (2003) devised a colorimetric assay to quantify resid-
ual color in decolorized zein products and performed a sta-
tistical assessment for color removal by a variety of
processing methods. Color removal with an activated car-
bon was the best decolorization method.
Activated carbons have been used in the past to decolor-
ize zein (Mason and Palmer, 1934; Swallen, 1938; Pearce,
*
Corresponding author. Tel.: +1 309 681 6351; fax: +1 309 681 6686.
E-mail address: David.Sessa@ARS.USDA.GOV (D.J. Sessa).
1941; Starling et al., 1951; McInnis and Tang, 2003). Acti-
vated carbon is a non-specic adsorbent that not only
binds the color components but also the protein
components.
The objective of this investigation is to determine if a
change in temperature can be used to enhance the binding
capacity of color/odor components onto activated carbon
while limiting that of protein adsorption. A means of mon-
itoring protein versus color/odor components is required to
measure the eect of heat on the kinetics of adsorption of
those components onto activated carbon. In a column
chromatographic scheme for decolorizing zein in aqueous
ethanol (Sessa et al., 2003) column eluates were monitored
by ultraviolet (UV) spectroscopy at wavelengths 280 and
325 nm. Those researchers demonstrated that pooled
eluates with low UV absorbance at 325 nm possessed
slight yellow coloration, whereas, pooled eluates with
increased absorbance at 325 nm were intensely yellow.
The colored components coelute with a polyamine

0960-8524/$ - see front matter Published by Elsevier Ltd.

doi:10.1016/j.biortech.2007.11.076
 D.J. Sessa, D.E. Palmquist / Bioresource Technology 99 (2008) 63606364
conjugate characterized as diferuloylputrescine, a com-
pound found in corn oil by extracting ground corn with
ethanol (Moreau et al., 2001; Moreau and Hicks, 2005).
Yellow zein extracts processed with activated carbon (Sessa
et al., 2003) yielded a white zein product with trace UV
absorbance at wavelength 325 nm. Odor evaluations by
two international companies interested in a decolorized/
deodorized zein declared that the white zein produced
was odor free (unpublished data). Therefore, monitoring
adsorption of yellow aqueous alcohol solutions of zein
onto an activated carbon at wavelength 280 nm for tyro-
sine in zein protein and 325 nm for diferuloylputrescine
provided the means to evaluate the kinetics of adsorption
at several temperatures. We applied the Freundlich iso-
therm model (Crittenden et al., 1985) for this evaluation.
2. Experimental
2.1. Materials
Commercial yellow zein (FC4000) with proximate anal-
ysis, dry-basis: 93.09% crude protein (Dumas N  6.25),
5.26% crude fat, 0.04% crude ber, 0.05% ash, was pur-
chased from Freeman Industries, Tuckahoe, NY. A granu-
lar activated carbon from coal, type CPG-LF 12  40
(acid-washed) was provided by Calgon Carbon Corp.,
Pittsburgh, PA. Ethanol, ACS reagent, 200 proof,
P99.5% was obtained from SigmaAldrich Inc., St. Louis,
MO. The aqueous ethanol concentrations used in this
investigation were measured on a weight/volume basis.
2.2. Preparation of colorless zein product
A highly puried zein product was essential in order to
develop the analytical procedure for our model system. To
accomplish that task yellow zein was processed using a
combination of the activated carbon treatment and ultral-
tration/dialtration on a tangential ow system (Sessa
et al., 2003). An activated carbon lter device with pleated
HEPA (i.e. high eciency particulate air) lter, Carbon-
Cap 150 with area equivalent 82,000 m2, Whatman Inc.,
Clifton, NJ, was hooked in tandem with a Pall Centramate
equipped with a 5000 Da MWCO OMEGA membrane. A
yellow, 10% zein solution in 70% ethanol (350 mL) was dia-
ltered at room temperature (i.e. 25 C) with 12 1-L
batches of 70% aqueous ethanol at a rate of 800 mL/h by
use of a feed pressure of 220 kPa and retentate pressure
of 110 kPa. Solvent was stripped from the retentate on a
rotary evaporator until zein started to precipitate. Remain-
ing alcohol was removed by dialyzing the milky zein disper-
sion contained in 1000 Da MWCO dialysis casing against
water; the precipitated zein was freeze-dried.
2.3. Sample protocol
The activated carbon granules were sieved through a
number 30 screen to remove ne carbon powder. That por-
6361
tion retained on the screen was washed three times with an
excess volume of 70% aqueous ethanol to further remove
powdered carbon adhering to the surface of the carbon
granules. The drained, washed granules, after evaporation
of ethanol, were dried overnight in a hot air oven at 105 C.
The dried carbon granules were measured into eleven
250 mL Erlenmeyer asks as follows: 0, 0.1, 0.2, 0.3, 0.4,
0.5, 1.0, 2.0, 4.0, 7.0, and 10.0 g. A stock solution of
0.4% commercial zein, 100 mL each ask, was added and
the asks then sealed with a rubber stopper. The asks
were gently shaken at 50 rpm on an Innova 4000 Incubator
Shaker, New Brunswick Scientic Co. Inc., Edison, NJ,
where equilibrium conditions were established at 24 h.
Multiple adsorption experiments were run at 25, 40, 45,
50, 55, 60 C with shaking. The volume of the liquid por-
tion from each ask was measured to ensure accurate con-
centration calculation, then decanted into separate tared
evaporating dishes. The respective carbon solids from each
ask were likewise transferred to tared evaporating dishes.
The ultraviolet absorbances of each liquid fraction were
measured at wavelengths 280 and 325 nm. Solvents from
the zein decantates as well as residual solvents remaining
on the carbon solids were each evaporated by air drying
followed by completely drying in a hot air oven at 105 C
for 3 h. The mass of zein as well as carbon solids in each
of the evaporating dishes were measured.
2.4. Analyses
The test adsorptions were designed to permit a general
linear model regression consisting of three replicate trials
measuring log (qe) as a function of log (Ce) at two wave-
lengths and six temperatures.
The dependent variable qe (equilibrium solid phase con-
centration from the Freundlich isotherm qe = KfCe1/n) was
examined as a function of Ce (liquid phase solute concen-
tration for weights of activated carbon) for zein at two
wavelengths (A280 nm and A325 nm) and six temperatures
(25, 40, 45, 50, 55, 60 C). Data from three replicate trials
were averaged and log (qe) and log (Ce) values were used
for developing simple linear regression equations (SLR).
Protein (A280 nm) and contaminants (i.e. color/odor
assessed at A325 nm) equations were compared at each tem-
perature using full and reduced model regression tech-
niques (Neter et al., 1990).
The original Freundlich isotherm equation qe = KfCe1/n
was linearized with the resulting SLR form: log (qe) =
log (Kf) + (1/n)log (Ce). Adsorption capacity estimates (Kf)
were calculated as the antilog of the intercept from the lin-
earized SLRs. Adsorption bond strength estimates (1/n)
were obtained directly as the slope from the linearized
SLRs. Values, where Ce = 0, were deleted from the analy-
ses because the log transformation is undened at that
point.
For each wavelength, intercepts and slopes of the linear-
ized equations for all six temperatures were compared to
determine which temperature had the largest adsorption
6362 D.J. Sessa, D.E. Palmquist / Bioresource Technology 99 (2008) 63606364

capacity and bond strength. When the 95% condence
intervals of the regression parameters overlap, the inter-
cepts or slopes for each temperature are not signicantly
dierent from one another.
3. Results and discussion
3.1. Yellow and white zein standards
Processing yellow zein using a combination of activated
carbon treatment and ultraltration/dialtration on a tan-
gential ow system yielded a white zein product with an
average Dumas nitrogen of 15.31, which when adjusted
for 4.90% moisture, gave 100% protein. Spectral analysis
of that product (Fig. 1), dissolved in 70% aqueous ethanol
from 250 to 400 nm showed one major absorbing peak at
wavelength 280 nm and no absorbance at wavelength
325 nm. The UV absorbing peak at about 325 nm found
in yellow zein solutions (see Plot #1, Fig. 1) is caused by
a polyamine conjugate which has been isolated and identi-
ed as diferuloylputrescine by UV and mass spectroscopy
(Sessa, unpublished data). The UV absorption spectrum
of diferuloylputrescine gave a dual peak with maximum
absorbance at 317 nm and a smaller peak at 293 nm (data
not shown) which UV absorbances are similar to that
reported in the literature (Moreau et al., 2001). The spec-
trum of yellow zein is a convolution of all the components,
whereas, the spectrum of pure zein (see Plot #2 in Fig. 1)
lacks the impurities found in yellow zein. Because the UV
spectrum of diferuloylputrescine overlaps with the protein
component, we monitored its contribution at wavelength
325 nm, where the zein absorbance contribution is essen-
tially zero. Standard curves were developed using pure zein
and yellow zein at a variety of weight percents from 0.4%
to 0.05%. With our pure zein as a reference, subtraction
of the impure yellow zein was used to quantify the amount
of impurities present in zein solutions.
1.8
#1
1.5
#2
1.3
Absorbance
Based on these ndings the spectral trend line for puri-
ed zein was used as the default calculation concerning
the A280 nm and A325 nm values. Hence, the A280 nm used in
this investigation for zein protein standard is dened by a
straight line y = 4.7466x + 0.0281 with an R2 = 0.9994
and the A325 nm contaminant is dened by y =
3.0384x + 0.0032 with an R2 = 0.9987, where x represents
the concentration. Removal of the xanthophylls, that cause
the yellow color in zein (Sessa et al., 2003), accompanies
the removal of diferuloylputrescine. Therefore, x or Ce in
our linearized Freundlich isotherm model for the protein
components is dened by Eq. (1) and the Ce for the
color/odor components is dened by Eq. (2):
y  0:0281
Ce at k280 nm 1
4:7466
y  0:0032
Ce at k325 nm 2
3:0384
3.2. Adsorption isotherms
Kinetics of adsorptions of protein and color/odor com-
ponents onto activated carbon at two temperatures, 25 and
60 C, were performed over a 27 h period to establish the
equilibration times needed to maximize their adsorptions.
At either temperature, maximum adsorption of protein
measured by A280 nm and odor components represented
by A325 nm occurred at 24 h and thereafter. With that equil-
ibration time at 25 C, the A280 nm diminished from 3.34 to
1.41 while A325 nm dropped from 1.78 to 0.17; at 60 C, the
A280 nm changed from 3.29 to 1.07 and A325 nm decreased
from 1.75 to 0.07.
Table 1 data demonstrate the adsorption capacity (Kf)
comparisons and bond strength (1/n) comparisons between
temperatures for each wavelength (k). The adsorption
capacity for zein protein did not change signicantly over
the entire temperature range from 25 to 60 C. However,
adsorption capacities for the color/odor components did
increase signicantly at 55, where the Kf value is well
above all other values while at all other temperatures eval-
uated only slight increases were observed.

1.0 Table 1
Adsorption capacity (KF) comparisons and bond strength (1/n) compar-
0.8 isons between temperatures for each wavelength (k)
Temp. (C) A280 k (nm) A325 k (nm) A280 k (nm) A325 k (nm)
0.5
1 1
KF KF /n /n
0.3 A A B
25 166.5a 155.3d 1.4791a 0.7137aB
40 170.8a 177.5cd 1.0850ab 0.5335bc
0.0
45 190.9a 224.1bc 1.1477ab 0.6659ab
250 275 300 325 350 375 400
50 182.6a 196.4bcd 0.8878b 0.4681c
Wavelength
55 218.2a 291.2a 1.1079ab 0.6998a
60 193.8a 233.1b 0.7550b 0.4362c
Fig. 1. Spectral analysis of yellow zein (#1) at 20 mg/ml and colorless/
A
odorless zein (#2) at 24 mg/ml each in 70% aqueous ethanol over the KF values within a column (k) followed by the same letter are not
wavelength region 250 to 400 nm. (For interpretation of the references in signicantly dierent based on overlap of the 95% condence intervals.
B
colour in this gure legend, the reader is referred to the web version of this Slopes (1/n) within a column (k) followed by the same letter are not
article.) signicantly dierent based on overlap of the 95% condence intervals.
 D.J. Sessa, D.E. Palmquist / Bioresource Technology 99 (2008) 63606364 6363

Table 2 carbon especially as it saturates. The viscosity of zein solu-

Simple linear regression equations from linearized Freundlich isotherms tions in aqueous ethanol decreases with increased tempera-
for zein protein (A280 nm) and color/odor components (A325 nm) adsorbed
to activated carbon at dierent temperatures
tures (Fu and Weller, 1999). Diminished viscosity of the
aqueous ethanol solution of zein should allow transport
Temp. (C) k (nm) Equationa Adj. R2
of the low molecular weight color/odor components into
25 280 Log (qe) = 2.2215 + 1.4791 Log (Ce) 0.94 the interior of the carbon micropores. Bond strength gov-
25 325 Log (qe) = 2.1911 + 0.7137 Log (Ce) 0.98
40 280 Log (qe) = 2.2325 + 1.085 Log (Ce) 0.94
erned by the slope (1/n) for protein showed considerable
40 325 Log (qe) = 2.2491 + 0.5335 Log (Ce) 0.98 overlap of values with temperatures, where bond strengths
45 280 Log (qe) = 2.2808 + 1.1477 Log (Ce) 0.93 for proteins were highest at 25 C and lowest at 50 and
45 325 Log (qe) = 2.3504 + 0.6659 Log (Ce) 0.98 60 C. Bond strength for the odor component at wave-
50 280 Log (qe) = 2.2614 + 0.8878 Log (Ce) 0.90 length 325 nm was signicantly higher at 25 and 55 C than
50 325 Log (qe) = 2.2932 + 0.4681 Log (Ce) 0.98
55 280 Log (qe) = 2.2338 + 1.1079 Log (Ce) 0.94
all other temperatures evaluated. The specic adsorption
55 325 Log (qe) = 2.4642 + 0.6998 Log (Ce) 0.99 temperature of 55 C for the odor component absorbing
60 280 Log (qe) = 2.2874 + 0.7550 Log (Ce) 0.91 at wavelength 325 nm is unusual and may indicate a change
60 325 Log (qe) = 2.3675 + 0.4362 Log (Ce) 0.99 in surface phenomenon attributed to the zein molecule.
a
Within each temperature level there were signicant dierences Selling et al. (2007) demonstrated with circular dichroism
(p 6 .01) between equations for protein (A280 nm) and color/odor (A325 nm). that changes in the secondary structure of zeins primary
structure occurred upon heating 80% aqueous ethanol
solution to 70 C because mean residue ellipticity decreased
SLR equations from linearized Freundlich isotherm 20% at wavelengths 208 and 222 nm. Those changes were
model are given in Table 2. Within each temperature level reversed upon cooling. The two negative peaks at 208
there were signicant dierence (p 6 0.01) between equa- and 222 nm are indicative of the amount of a-helix present
tions for protein (A280 nm) and color/odor (A325 nm). in the protein (Sreerama and Woody, 1994).
Figs. 2 and 3 are visual depictions of trend lines from the A decrease in mean residue ellipticity with heating
tabulated data in Tables 1 and 2 for each temperature and denotes protein denaturation accompanied by a loss of a-
their respective correlation coecients at each wavelength helix character. Momany et al. (2006) proposed a model
280 and 325 nm. for zein based on molecular dynamics simulations where
An increase in adsorption capacity with increased solu- three lutein molecules t into the core of the three triad
tion temperature denotes an endothermic process. Because helical segments helping to stabilize the conguration.
diusion is an endothermic process we can conclude that Conceivably, the loss of a-helix character upon heating
the increased adsorptions observed for the color/odor com- would release those xanthophylls, thereby, making them
ponents of zein with increased heat may be governed by available for adsorption on the activated carbon. Because
either pore diusion (Anoop Krishnan and Anirudhan, the binding of the zein color/odor components dened as
2002) or the energy required to adsorb onto a nonporous 1/n is signicantly higher at 55 C than those in all other

2.5
Linear (60280) y = 0.755x + 2.2874 R2 = 0.9075
Linear (55280) y = 1.1079x + 2.3388 R2 = 0.9355
Linear (50280) y = 0.8878x + 2.2614 R2 = 0.8976
Linear (45280) y = 1.1477x + 2.2808 R2 = 0.9315
Linear (40280) y = 1.085x + 2.2325 R2 = 0.9404
Linear (25280) y = 1.4791x + 2.2215 R2 = 0.9348
2.0
Log Q

1.5

1.0

-1.2 -1.0 -0.8 -0.6 -0.4 -0.2 0.0

Log Ce

Fig. 2. Eect of temperatures on the adsorption of zein protein at k280 nm by a coal-based activated carbon with linearized trend lines from Freundlich
isotherms.
6364 D.J. Sessa, D.E. Palmquist / Bioresource Technology 99 (2008) 63606364

2.5
Linear (60325) y = 0.4362x + 2.3675 R2 = 0.9905
Linear (55325) y = 0.6998x + 2.4642 R2 = 0.9942
Linear (50325) y = 0.4681x + 2.2932 R2 = 0.984
Linear (45325) y = 0.6659x + 2.3504 R2 = 0.9771
Linear (40325) y = 0.5335x + 2.2491 R2 = 0.9776
Linear (25325) y = 0.7137x + 2.1911 R2 = 0.98
2.0
Log Q

1.5

1.0

-2.0 -1.8 -1.6 -1.4 -1.2 -1.0 -0.8 -0.6 -0.4 -0.2 0.0
Log Ce

Fig. 3. Eect of temperature on the adsorption of color/odor components of zein at k325 nm by a coal-based activated carbon with linearized trend lines
from Freundlich isotherms.

zein aqueous ethanol solutions heated above 25 C, the References

unusual adsorption behavior at 55 C may result from a
combination of events involving solvent environment, vis-
cosity changes as well as changes in structure of the zein
molecule. Further investigations are needed to prove this
hypothesis.
4. Conclusions
The Freundlich isotherm model for adsorptions of zein
protein and its color/odor components onto an activated
carbon proved to be a satisfactory model based on statisti-
cal analyses. This model can now be used to investigate the
adsorption phenomena of other adsorbents. Application of
this model to the zein decolorization/deodorization process
to examine the impact of heat on the adsorption capacity
of an activated carbon medium demonstrated that process-
ing zein aqueous ethanol solutions at 55 C signicantly
enhanced the adsorptions of the color/odor components
relative to the adsorption of protein.
Disclaimer
Names are necessary to report factually on available
data; however, the USDA neither guarantees nor warrants
the standard of the product, and the use of the name by
USDA implies no approval of the product to the exclusion
of others that may also be suitable.
Acknowledgements
We thank Mardell L. Schaer and David Baudo for their
excellent technical assistance.
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