Environmentally Benign Preparatory Processes – Introducing a Closed-Loop System
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C99-A07
Annual Report, September 1999
Team members:
G. Buschle-Diller, AU (leader)
R. Radhakrishnaiah, GA Tech
H. Freeman, NCSU
S.H Zeronian, UC Davis
Goal Statement
This project focuses on the development of new strategies for textile wet processing using only environmentally friendly compounds. The ultimate goal is to combine desizing/bioscouring and/or desizing/biobleaching or all three processes to produce an effective closed-loop procedure, reusing the entire treatment bath of all three steps. Fundamental questions pertaining to enzyme specificity, the underlying mechanisms and modes of action are addressed. Approaches to anchor the enzymes to the fiber surface will be explored, since this should considerably enhance enzyme efficiency.
Abstract
For bioscouring, combinations of enzymes are applied that decompose the noncellulosic impurities in raw cotton. The enzymes are selected based on their pH and temperature compatibility. The most effective conditions for this process are established in regard to the properties of the scoured goods. Desizing is performed with amyloglucosidase, an enzyme capable of breaking down starches into mostly glucose. For biobleaching, peroxidases, laccase/mediator systems and glucose oxidases are explored and evaluated in terms of whiteness level. Physical and fine structural properties of the textile material will be investigated after each processing step. Glucose oxidases for biobleaching is likely the most promising compound since in this case the treatment baths of all processing steps can be reused.
Background
Enzymatic Desizing
Desizing processes that are considered environmentally friendly usually follow either one of two approaches - natural biodegradable sizing chemicals are applied, such as starch and starch derivatives, or synthetic recyclable formulations are employed. A well-known example for the latter category are PVA-based sizes which can be removed with hot water, purified by ultrafiltration and reused. The application of starch-based sizes is a well-established process. Starches cannot be recycled. These compounds though can be easily decomposed with amylase enzyme formulations. A whole amylase complex usually contains various types of exo- and endo-enzymes, glucoamylases and debranching enzymes with different modes of action. The end-products of the degradation process are sugars which are nontoxic, however, affect the BOD of the wastewater. For the process of enzymatic desizing, the problem of the sugar-containing effluent has to be addressed.
Bioscouring
The water absorbency and whiteness of raw cotton is very limited. Noncellulosic impurities, such as fats, waxes, proteins, pectins, natural colorants, minerals and water-soluble compounds, are found to a large extent in the primary wall and to a lesser extent in the secondary wall (see Table 1). Quantity and composition varies with growing conditions, climatic factors and cotton variety. Conventional scouring is performed with 3-6% aqueous sodium hydroxide solution at the boil. Although very effective, the process affords huge amounts of rinse water to remove the alkaline treatment solution once the process is complete. In times of increasingly stringent environmental regulations excessive usage of water and water contamination are unacceptable and an alternative way of scouring needs to be found.
Compound / Amount (%) / (%) in primary wallCellulose / 86-93 / 52
Pectins, proteins, natural colorants / 5-6 / 24
Oils, fats, waxes / 0.1-1 / 7
Minerals / 1
Moisture / 8
Table 1. Average composition of raw cotton [1, 2].
Bioscouring is a novel process based on the idea of particularly targeting the noncellulosic impurities with specific enzymes. For example, pectinases could be used for the decomposition of pectinic substances, proteases for proteins, lipases for fats. Part of the natural pigments are associated with the non-cellulosic compounds and will be lifted off the fiber during bioscouring. Enzymes used for bioscouring generally do not affect the cellulose backbone and fiber damage is expected to be limited.
Biobleaching
Currently, the most common industrial bleaching agent is hydrogen peroxide, which, when applied at pH 10.5-11 and temperatures close to the boil, produces the desired level of whiteness of the textile goods. Hydrogen peroxide decomposes into environmentally benign compounds (water and oxygen). The greatest problem occurring during bleaching with peroxide are radical reactions of the bleaching compounds with the fiber. These reactions can lead to a decrease in the degree of polymerization and eventually to a drop in tensile strength, especially in presence of metal ions which act as activators for hydrogen peroxide [3].
Enzymatic bleaching is still in the developing stage. Three different approaches have been discussed. First, laccase/mediator systems have been used successfully for bleaching of wood pulp and it could be expected that they are applicable to bleaching of cotton as well. However, suitable mediator compounds have to be found. Currently available mediators raise questions about efficiency and toxicity. Mediators are "quasi’- catalysts which act as electron transfer components to support the action of the fairly non-specific laccases. They are consumed during the reaction and as such not considered true catalysts.
The second possibility for enzymatic bleaching are peroxidases. This class of enzymes is capable of activating various oxidizing agents, one of which is hydrogen peroxide. A fully satisfactory bleaching effect, however, has not yet been established with these enzymes. It is possible that these enzymes are deactivated too rapidly during the bleaching process and thus become ineffective fairly quickly.
The third and most promising approach is concerned with glucose oxidases. These are enzymes that generate hydrogen peroxide and gluconic acid from glucose and oxygen. This type of enzymes seems the most suitable of all options since it makes it possible to reuse sugar-contaminated effluent from other wet processing steps.
Project Approach – Creating a Closed-Loop
Since one of the main goals of this project is to develop a closed-loop system, which includes the reuse of the treatment effluent, we are primarily focusing on glucose oxidases for bleaching since glucose is generated at any event during desizing and scouring in sufficient amounts. However, the other bleaching systems mentioned above are also considered. For desizing, enzymes are selected that predominantly generate glucose as degradation product of starch-based sizes. Bioscouring is performed with combinations of enzymes that are promising in regard to both goals – to significantly increase fiber absorbency and to yield sufficient glucose from the decomposition of the non-cellulosic impurities. The addition of cellulase would definitely help the process, but might cause the typical fiber damage as observed with cellulase finishing procedures (biopolishing, [3]).
Results
Desizing and Bioscouring
The desizing process can be performed with any type of amylase. These enzymes are commercially available with flexible pH and temperature ranges. Nowadays, even thermostable amylases can be obtained. For this project, amyloglucosidases were applied since these enzymes produce primarily glucose as the decomposition product, which will be useful for the bleaching step.
Pectinases applied alone did not seem to be effective enough in regard to improving the water pick-up of the scoured samples. As the data in Table 2 show, the admixture of lipases clearly increased the water absorbency of the fiber. Cellulases in combination with pectinases produced a surprising softness of the scoured goods which is an asset at this early processing stage. However, in terms of water absorbency cellulases do not seem to be very effective. Also, as mentioned above, care has to be taken to avoid excessive fiber damage. Xylanases, enzymes advantageous during biopolishing of cellulosic fibers other than cotton especially, showed a moderate effect in this case. When all enzymes were applied in combination, the highest weight loss was observed and the water absorbency came close to the value of the commercially treated sample. This effect can most likely be attributed to the action of the lipases in the blend, probably in combination with some synergistic reactions of the other involved enzymes. It has been found that proteins in raw cotton have a very minor influence on the properties of the unscoured fiber and do not need to be specifically targeted.
The treatment conditions are energy efficient because all the enzymes work well at temperatures of 50 to 60°C or even lower. Since enzymes are biocatalysts only minute amounts are necessary for the desired effects. The deactivation of the enzymes is achieved by simply raising the temperature or the pH. This step can easily be incorporated in the following processing stage.
Enzyme system / Weight loss (%) / Rel. absorbency (%)Control* / - / 100
Pectinase / 11.5 / 18
Pectinases, cellulase / 13.3 / 26
Pectinase, lipase / 11.5 / 64
Pectinase, xylanase / 11.5 / 45
Combination of all enzymes / 13.9 / 87
* For reasons of comparison a conventionally scoured and bleaced cotton fabric was used as a control sample.
Table 2. Weight loss and relative water absorbency recorded after a 1-h enzymatic scouring process at 50°C (tumbling speed 42 rpm, 5 steel balls/g fabric).
Combinations of pectinases and lipases not only yielded satisfying results concerning water absorbency. The tensile properties of the in this manner scoured samples were basically unchanged from the untreated control (2-3% strength loss on the average, based on the untreated greige sample). If, however, cellulase was present during the scouring process, tensile strength decreased by approximately 20%, suggesting the usual fiber damage observed during extensive biopolishing. Scanning electron micrographs showed occasional cracks and some surface peeling as expected.
In the process of bioscouring with pectinases, galacturonic acid, glucose and other sugars are generated. With mainly glucose as the degradation product it should be possible to combine and reuse the desizing and scouring treatment baths for bleaching with glucose oxidases in a closed loop. We are presently investigating whether sugars other than glucose inhibit this process.
A current problem encountered with reusing the treatment bath is the increasing contamination, especially if the scouring and the desizing process are combined. These contaminants could redeposit on the cotton material. It is also possible that some of the hydrogen peroxide is working on these contaminants and therefore remain unavailable for bleaching. We are at present trying filtration and centrifugation methods for purification.
It is known that scouring and especially bleaching, if performed in the conventional manner, causes a reduction in degree of polymerization. If hydrogen peroxide is used as the bleaching agent, radical reactions can occur that affect the cellulose backbone. We measured the intrinsic viscosity of some commercially treated samples, calculated the degree of polymerization (DP). The results are summarized in Table 3. The first three samples (Testfabrics, Inc., NJ) in Table 3 are of the same origin, yarn count and fabric construction. The following samples were treated in our laboratory according to industrial procedures. The last sample was bioscoured with pectinase. It is quite obvious that the DP is strongly influenced by the preparation process conditions. The drop in DP might not manifest itself in a simultaneous decrease in tensile strength at this stage, however, might lead to problems at a later stage.
Sample / Degree of polymerizationGreige cotton (Testfabrics) / 3850
Desized coton (comercial sample) / 3700
Desized, scoured and bleached (commercial sample) / 1450
Alkaline scouring (lab) / 2800
Alkaline scouring and hydrogen peroxide bleaching (lab) / 1100
Scouring with pectinase / 3750
Table 3. Degree of polymerization of various cotton samples.
The DP of the bioscoured sample is basically unchanged from the untreated cotton. This is expected since pectinase does not affect cellulose. We are currently in the process of determining the DP values of our other bioscoured samples.
Biobleaching
For biobleaching we started with experiments involving laccases and mediators. Two mediators, commonly used for wood pulp bleaching, have been applied so far – violuric acid and HBT (N-hydroxybenzotriazole). So far we have varied the concentration of laccases and mediators as well as the temperature (from 35 to 60° C). The degree of whiteness has been improved by only 14-18%, which is, of course, not satisfactory and more experiments are necessary.
Experiments using glucose oxidase as the bleaching agent are in progress. Conditions have to be found that allow the enzyme to be most active, but also the hydrogen peroxide to be effective in regard to the bleaching reaction.
Dyes for Structural Investigations
Synthesis
The dyes we synthesized and the synthetic routines employed are shown in Figure 1. Both of the direct dyes utilized a nongenotoxic benzidine (5) as the precursor. The two diamines were prepared in 4 steps, starting with the alkylation of the appropriate 2-nitrophenol (1). The alkylation step utilized a hot mixture of 1-bromopropane, potassium carbonate and 2-methoxyethanol. The corresponding nitroaryl ethers (2) were subjected to alkaline reduction to give the air sensitive hydrazines 3. The benzidine rearrangement step (3 to 4) was accomplished by adding 15% HCl aqueous solution, and was followed by treatment of 4 with 10% NaOH solution to make the free bases 5 (R = H, CH3).
To synthesize type 8 dyes, benzidines 5 were tetrazotized at 0-5° C in the presence of HCl and NaNO2 and then coupled with J-acid (7) at 0-5° C and pH 8-9. The progress of the reactions was followed by TLC (BuOH:EtOH:NH4OH:pyridine/4:1:3:2, BuOH:EtOH:NH4OH:H2O/ 3:1:0.5:1.5), and the dyes were isolated by adding NaCl to effect precipitation. The precipitated dyes were collected by filtration and dried, to give reaction yield was >85%.
Dye structures were confirmed by mass spectrometry, the results of which are summarized in Table 4. In both cases, the expected supporting data were produced, with each dye giving an m/2 peak consistent with the parent molecular weight.
Absorption spectra of the two dyes show l max = 506 (red) and 553 (purple) for R=CH3 and R=H, respectively. The significant difference in color arises from the loss of planarity around the biphenyl linkage that is imposed by the methyl group.
Table 4. Electrospray ionization (ES) mass spectral data for dyes 8.
DYE / MW / Peaks observed and assignments8 (R = CH3) / 872.87 / 413.3 = (M-2Na)2-/2
8 (R = H) / 844.82 / 399.1 = (M-2Na)2-/2
Fig. 1 Synthesis of direct dyes 8 (R = H, CH3).
Future Experiments
Besides working on establishing effective processing conditions for the biopreparation processes, we are planning to assess the scoured and bleached samples concerning their fine structural properties to better understand how the various enzymes function. These fine structural characterizations will be complemented with molecular mechanics calculations. The hand properties will be evaluated using the Kawabata KES-F system.