Determining Fiber Loss in Biofinishing
of Cotton and CottonNvool Blended Fabrics
~~~
By Shridhar V. Chikkodi, International Textile Center, Texas Tech University, Lubbock
B iofinishing is a relatively new con-
cept of treating the fabrics with en-
zymes. Recent advances in biofin-
ishing of cellulosic fabrics have led to
multiple improvements of surface
properties. The main objective of
biofinishing is to upgrade the fabric by
removing the protruding fibers. The
conventional methods of removing the
protruding fibers employ a burning-off
process (singeing) or a chemical treat-
ment. The conventional methods are
temporary, potentially toxic, and fibers
return to the surface after a few
washings and form fuzz. The fuzz on
the surface of the fabrics constitutes the
major reason for customer dissatisfac-
tion. However, by using enzymes in the
finishing process, the protruding fibers
can be permanently removed from the
fabric thus eliminating the fuzz. The
enzyme treatment not only keeps the
fabric looking new after repeated
washings, but enhances feel, color,
softness and drapeability which trans-
lates into a higher quality textile or
apparel product.*P2
ABSTRACT
In recent years, biofinishing, an
application technique using enzymes
such as cellulases and proteases to
digest the protruding fibers, has
gained considerable attention.
Several studies have reported that a
certain weight loss is desirable as a
result of an effective enzyme
treatment. In this study the
relationship between weight loss and
the amount of glucose and amino
acids released was examined. It
appeared that weight loss conformed
with the glucose and amino acids
released, thus confirming the
existence of a relation between
weight loss and the activity of
specific enzymes.
KEY TERMS
Biofinishing
Cellulases
Enzymes
Proteases
28 Textile Chemist and Colorist
In the p r e s e k study, cotton and
cottonlwool blended fabrics were
treated with cellulase and cellulase or
protease enzymes, respectively. Cellu-
lases degrade cellulose by breaking the
1,4 D-glucosidic bonds, whereas pro-
teases hydrolyze the peptide bonds of
rotei ins.^^^ The hydrolysis of gluco-
sidic or peptide bonds leads to the
degradation of the molecules into
smaller units, which are amenable to
further degradation into glucose and
amino acids, respectively. An effective
enzymatic treatment results in suc-
cessful removal of protruding fibers,
hence a certain weight loss is ex-
pected. The main objective of this
study was to determine the relation-
ship between fabric weight loss and
free glucose and amino acids found in
enzyme solutions after biofinishing
cotton and cottonlwool blended fab-
rics. In the process, the assay meth-
ods were also optimized to measure
the free glucose and amino acids re-
leased from the enzymatic action.
Material and Methods
Plain woven fabrics of 37 tex ring-spun
yarn were used. The fiber contents of
the fabrics included 100% cotton and
80120 and 60140 cottonlwool blends.
Prior to enzymatic treatment, fabrics
were desized in a jig using 1:10 liquor
ratio, 1% nonionic detergent, 0.5%
hydrogen peroxide and 0.25% acetic
acid at 9OC for 30 minutes, followed
by a hot rinse (9oC) for 20 minutes and
a final cold rinse to thoroughly remove
the PVA added to the warp during
slashing. Cellulase and protease en-
zymes used in this study came from
Royal Gist-Brocades (Charlotte, N.C.).
Cellulase enzymes used were of indus-
trial grade with an activity of 100 CCUI
gram and the protease enzymes had the
proteolytic activity of 400,000 Delft
Unitslgram.
Cellulase was applied on 100%
cotton and on cotton/wool blends to
digest the protruding cotton fibers.
Protease was used on cottonlwool
blends to digest the protruding wool
fibers.
Cellulase Digestion
The experimental fabric, three meters
in length, was weighed, loaded on a
laboratory jig, and run dry four times
around doctor blades to raise the loose
fibers to the surface. The doctor blades
also assured a thorough mechanical
action throughout the enzyme applica-
tion. The aqueous bath had a liquor
ratio of 1:10, pH of 4.5 (using sodium
acetate buffer), temperature of 60C and
cellulase enzyme added at 3 or 5% on
the weight of fabric (owf), equivalent
to cellulase activity of 3 or 5 CCU/gram
of fabric. The pH was periodically
checked during the one-hour enzy-
matic treatment and adjusted with ace-
tic acid when necessary. After the treat-
ment, the fabric was washed in hot
deionized water at 9OC to terminate the
enzyme action, and then dried using a
drum dryer.4
Protease Digestion
Proteases were applied in a second in-
dependent treatment to the cottonlwool
blend using the same conditions as in
cellulase digestion changing only the
bath pH to 8.5 using boric acid buffer
and sodium hydroxide. Enzyme activ-
ity of 12,000 or 20,000 Delf Units/gram
of fabric were equivalent to 3 or 5% owf.
Analysis of Enzymatic Digestion
At the same time the fabric was re-
moved, ten liquid samples of 10 mL
each were collected from the well-
mixed bath liquor and frozen imme-
diately (-20C) for the glucose and
amino acid assay. To determine
whether the effect of treatment was
truly due to enzymatic action, samples
of fabrics treated in the same manner
but without the addition of enzymes
were collected.
Glucose Assay
In order to quantify glucose, it was nec-
essary to digest all the remaining cel-
lulose fragments. After thawing the fro-
zen samples with hot water, four 2 mL
aliquots were drawn from each 10 mL
liquid sample. To each of these test
samples, 0.158 mL of 1M sodium ac-
Vol. 28, No. 3
 etate buffer (pH 4.5) and 1 mL of cel- zine methosulfate). To 6.7 pL of each 1. Visual examination was performed
lulase enzyme (100 CCU) were added. aliquot, 0.33 mL of assay reagent was with an Olympus BH-2 microscope
To aid complete digestion the samples added and the sample was mixed by after staining with 0.1% Aniline
were incubated in a sealed condition gentle inversion and allowed to stand Blue to detect protein.
at 60C for 24 to 96 hours with constant for 10 minutes. Then 3.33 mL of 0.1N 2. Amino acid assay was performed at
stirring in a Reactitherm (Pierce Chem HC1 was added and the optical density 24, 48, 72 and 96 hours of protease
Co., Rockford, Ill.). Complete digestion was measured at 520 nm using a treatment.
was ensured by two methods: Bausch & Lomb Spectronic 2 1 Spectro- Once the time required for conrplete
1. Visual examination was performed photometer against a reagent blank of digestion was established, triplicate
with an Olympus BH-2 microscope 6.7 pL of nanopure water and 0.33 mL aliquots of each experimental sample
equipped for polarization analysis to of assay agent. To quantify the glucose were assayed as follows. The amino
detect cellulose birefringence and for concentration in the test samples, a acid assay was carried out using the
fluorescence analysis to detect cel- standard curve was prepared using 1:1, Ninhydrin reaction. Ninhydrin reagent
lulose after staining with the fluores- 1 2 , 1:4 and 1:9 dilutions of stock so- was obtained from Sigma Diagnostics.
cent brightener Tinopal LPW (Ciba lution consisting of 5.56 mmoles/L glu- The 10 mmoles/L amino acid standard
Geigy, Greensboro, N.C.). cose in saturated benzoic acid. was prepared by combining 17.12 g of
2. Glucose assay was performed at 24, aspartic acid,-13.12 g of leucine and
48,72 and 96 hours of cellulase treat- Protein Assay 17.42 g of arginine per 100 mL of wa-
ment. To quantify amino acids, it was neces- . ~ 2 mL of each
ter as per P l ~ m m e rTo
Once the time required for complete sary to digest any remaining wool frag- sample, 2 mL of Ninhydrin reagent was
digestion was established, triplicate ments. After thawing the frozen added. The samples were heated for 15
aliquots of each experimental sample samples with hot water, four 2 mL minutes in boiling water and cooled to ’
were assayed as described below. aliquots were taken from each 10 mL room temperature. Then 3 mL of 50%
Glucose was analyzed using a diag- liquid samples for the assay. To each ethanol was added and the optical den-
nostic kit based on hexokinase (Sigma of these test samples, 0.158 mL of 1M sity was measured at 570 nm using a
Chemical Co., Catalog #115-A). The Boric Acid buffer (pH 8.5) and 1mL of Bausch & Lomb Spectronic 2 1 Spectro-
glucose assay reagent was prepared by protease enzyme equivalent to 400,000 photometer against a reagent blank of
mixing one vial of glucose enzyme re- Delf Units were added. The samples 2 mL of nanopure water, 2 mL Ninhy-
agent with 17.0 mL of deionized water were incubated at 60C for 24 to 96 drin reagent and 3 mL of 50% ethanol.
and 4 mL of glucose color reagent (3.95 hours with constant stirring to ensure In order to quantify the amino acid
mmoles/L of iodonitrotetrazolium complete digestion. Complete diges- concentration in the test samples, a
chloride and 1.63 mmoles/L of phena- tion was ensured by two methods: standard curve was prepared using i:i,
1:2, 1:4 and 1:9 dilutions of 0.1
mmoles/L amino acid solution.
Table 1. Effects of Cellula and Protease Treatment Results and Discussion
on Weight Loss ( Merent Fabrics
In the presence of buffer, 24 hours of
Weight Before Weight After Weight Weight Loss Due cellulase or protease treatment brought
Treatment Treatment Loss to Enzymes about complete digestion. Aftor this
Test Sample Enzyme (grams) (grams) (“N (“4 amount of time, there was no further
100% Cotton decrease in the number and sizo of fi-
Control 207.9 205.1 1.35
3% Cellulase 212.0 203.9 3.82 2.47 bers and no further increase in glucose
5% Cellulase 218.3 205.4 5.91 4.56 or amino acids. It should be noted that
00120 cottomooi these results indicate that the addition
Control 232.0 229.2 1.21 of buffer greatly enhances rate of ac-
3% Cellulase 230.4 222.8 3.25 2.04
5% Cellulase 234.1 221.1 5.55 4.34 tion of these enzymes, an observation
3% Protease 206.2 201.1 2.47 1.26 which has bearing on design of Fabric
5% Protease 209.0 201.3 3.68 2.47 enzymatic treatment processes.
60140 c o t t o m o o i The weights of the experimental
Control 189.3 187.4 1.01
3% Cellulase 196.5 191.1 2.75 1.74 fabrics were measured before and af-
5% Cellulase 190.8 182.8 4.19 3.18 ter enzyme treatments and reported to
3% Protease 193.0 185.4 3.94 2.93 the nearest gram. Table I shows the
5% Protease 190.0 179.7 5.42 4.41 weight loss after enzyme treatments for
all experimental fabrics at both enzyme
concentrations. As indicated, fabrics
Table 11. Released Glucose treated with 5% enzymes lost about 4-
Adjusted
6% weight, whereas those with 3%
Cellulase Glucose Released Glucose Released Standard treatment lost 2-4%. Effective removal
Test Sample (%I (mmoles/L) (mmoledL) Error of protruding loose fibers contributed
100% Cotton to most of the weight loss.6 However,
Control 6.45 f 0.25 a small weight loss of about 1-2% was
3% Cellulase 13.12 6.67 f 0.25 observed for control fabrics, which
5% Cellulase 14.84 8.39 f 0.00 may be due to the mechanical action
~mo cottomooi
that took place during the treatment.
Control 5.20 t 0.00
3% Cellulase 11.12 6.00 f 0.00 In the case of 100% cotton fabrics
5% Cellulase 11.36 6.16 f 0.25 treated with 5% cellulase, the weight
60140 Cotton/Wool
Control 4.96 _+ 0.23
loss was found to be more than that of
3% Cellulase 9.12 4.16 f 0.25 blended fabrics, probably because
5% Cellulase 10.36 5.40 f 0.00 more protruding cellulose fibers were

March 1996 Textile Chemist and Colorist 29

 Table 111. Released Amino Acids Table I11 shows the amount of amino
acid released and the corresponding
Amino Adjusted standard error. Fabric containing
Protease Acids Released Amino Acids Standard
Test Sample (“A) (mmoledL) Released (mmoles/L) Error higher wool content and fabric treated
with 5% protease enzymes released
80/20 ColtonMlool more amino acids as compared to those
Control 0.023 ? 0.002
3% Protease 0.059 0.036 * 0.002 treated with 3% protease enzymes. Fig.
5% Protease 0.068 0.045 i 0.000 2 illustrates the amount of amino ac-
60/40 CottonMlool ids released in mmoles.
Control 0.038 *f.0.005 The relationship between weight
3% Protease 0.077 0.039 0.002
5% Protease 0.085 0.047 f.0.000 loss and the amount of glucose released
for the fabrics treated with cellulase
enzymes is shown in Figs. 3-5. Linear
regression was used to examine this
16
14.84 3% relationship. The R2 values (0.919 and
Control
0.924) indicate the significant relation-
14 ta 3% cellulase treatment ships between the fabric weight loss
I5% cellulase treatmenl
and the amount of glucose molecules
12
released for 100% cotton and 60140
cottonlwool blend. However, the 801
10.36
20 cottonlwool blend exhibited a mod-
-E
E
U
10
erate R2 value (0.747). Similarly the
% data in Figs. 6 and 7 shows the corre-
-
m
a
E a lation between the weight loss and the
8 amount of amino acids released for the
8 fabrics treated with protease enzymes.
2 6
(3 High R2 (0.88 and 0.94) values were
observed for blended fabrics. In sum-
4 mary, there is a high correlation be-
tween weight loss and glucose and
2 amino acid concentrations.
Conclusions
0
100% cotton 80/20 cotton/wool j0/40 cotton/wool A method to indirectly assess the
amount of fiber loss in biofinishing of
Fig. 1. Effect of cellulase on the amount of glucose released. cotton and cottonlwool blended fabrics
was examined. Effective enzyme treat-
ment, which depended on fiber content
available for the cellulase enzyme to standard error. With the decrease in the and treatment level, resulted in pro-
attack. Similarly more weight loss was amount of cellulose content in the test gressive weight loss. Assays were used
observed in the case of 60140 cotton/ fabrics, a decrease in the amount of glu- to determine the amount of cellulose
wool blended fabrics treated with 5% cose released was observed. The glu- and protein fiber loss. This study
protease than in blends with lower cose released also depended upon the showed that the reported weight loss
percent wool, because more protrud- treatment level. Five percent cellulase- was proportional to the release of glu-
ing wool fibers were available for the treated fabrics released more glucose cose and amino acids. It confirmed a
protease enzyme to attack. than three percent treated fabrics. Fig. relation between weight loss and the
Table I1 shows the amount of glu- 1illustrates the amount of glucose re- amount of glucose and amino acid re-
cose released and the corresponding leased in mmoles. leased by specific enzymes.

-H
0.09

0.08

0.07
0.085
Control
3% Drotease treatment
5% brotease treatment

0.068 -
P
16 7
A /!
E
v E
1
0.06 V
ul
a,
m
0.05 -
a,
a,
8
=m
0 0.04 0
L

6
0
.-C 0.03 4 A 100% Cotton
E -Linear (100% Cotton: j
a 0.02 2

0.01 0 ............................................................................................................. j

0
60140 80/20
cottonlwool cottonlwool
Fig. 2. Effect of protease on the amount of amino acid released.

30 Textile Chemist and Colorist COi, Vol. 28, No. 3

 14 :

A
A a0120 cottonlwool
-Linear (80120
cottonlwool)

0 1 2 3 4 5 6
Weight loss in %
Fig. 4. Relationship between weight loss and glucose released for cel- Fig. 5. Relationship between
lulase-treated fabrics. cellulase-treated fabrics.
0.08 0.09

0.08
-$ 0.07

0.06
sE 0.07
v Y

U p 0.06
2 0.05 In
-2mal 0.04 -$ 0.05
E
0 0.04
0
2 0.03 A
m
2 0.03 A
0
.-K A A 80120 cottonlwool .- A 60140 cotton wool
E 0.02 E 0.02
<
< -Linear (80120 -Linear (60140 cotton
0.01 cottonlwool) 0.01
wool)

0 0
0 1 2 3 4 0 1 2 3 4 5 6
Weight loss in % Weight loss in %

Fig. 6. Relationship between weight loss and amino acid released for Fig. 7.Relationship between weight loss and amino acid released for
protease-treated fabric. protease-treated fabric.

Plans for future research include the
use of other commercially available
cellulase and protease preparations of
various activities from different manu-
facturers to examine the effectiveness of
other enzyme preparations. It should be
noted that names of commercial prod-
ucts used in this study are solely pro-
vided for the purpose of providing
specific information and their mention
does not imply recommendation.
Acknowledgements
The author thanks Candace Haigler,
Department of Biology at Texas Tech
University, for her supervision during
the course of this work and technicaI
guidance during the preparation of this
manuscript. Appreciation is extended
to B. G. Wyatt, International Textile
Center at Texas Tech University, for his
comments in reviewing this paper. The
Department of Biology and Interna-
tional Textile Center at Texas Tech
University, is also acknowledged for
providing resources to undertake this
work.
References
1. Hemmpel, W. H., International Textile Bulle-
tin Dyeing /Printing/Finishing, Vol. 3 7 , No. 3,
March 1991, p5.
2. Schubel, P.. Textile Industries Dyegest-SA,
Vol. 9, No. 11. November 1990, p4.
3. Enzymes in Industry: Production and Appli-
cations, Edited by W. Gerhartz, Weinheim
(FRG): VCH Verlagsgesellschaft. 1990.
4. Chikkodi. S., Thesis:EffecfsofBiofinishingon
Cotton and Catton/Wool Blended Fabrics,
Texas Tech University, Lubbock, 1994.
5 . Plummer, D. T., An Introduction to Practical
Biochemistry, McGraw-Hill Book Co., London,
1989.
6. Tyndall, R. M.. Textile Chemist and Colorist,
Vol. 24, No. 6, June 1992, p23.
Author's Address
Shridhar V. Chikkodi, International
Textile Center, Texas Tech University,
Lubbock, Texas 79408.

March 1996 033 Textile Chemist and Colorist 31