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Enzyme processing of textiles in reverse micellar solution
Journal of Biotechnology 89 (2001) 263– 269 www.elsevier.com/locate/jbiotec
Enzyme processing of textiles in reverse micellar solution Kazuya Sawada a,*, Mitsuo Ueda b a
Japan Society for the Promotion of Science, Faculty of Engineering and Design, Kyoto Institute of Technology, Matsugasaki, Sakyo-Ku, 606 -8585, Japan b Faculty of Textile Science, Kyoto Institute of Technology, Matsugasaki, Sakyo-Ku, 606 -8585, Japan Received 26 June 2000; received in revised form 9 January 2001; accepted 15 January 2001
Abstract Scouring of cotton using pectinase enzyme, bioscouring, in reverse micellar system was studied. The effectiveness of bioscouring was evaluated by measuring weight loss of cotton, analyzing pectin and cotton wax remaining and by wetness testing. Pectinase enzyme showed excellent activity even in organic media, and the effectiveness of scouring was equivalent or better than that achieved by conventional alkaline process or bioscouring in aqueous media. Enzymatic modification of wool using protease enzyme in the same system was also studied. It has found that felting property and tensile strength of wool fabrics treated by protease in reverse micellar system were superior to those in aqueous media. Possibilities of utilization of the same system for the subsequent textile dyeing process were also investigated. It was found that cotton and polyester fabrics were dyed satisfactorily by reverse micellar system compared to conventional aqueous system. © 2001 Elsevier Science B.V. All rights reserved. Keywords: Reverse micelle; Bioscouring; Pectinase; Protease; Dyeing
1. Introduction For the past few decades, various kinds of dyeing and textile processing technologies have been developed and the processes have become performed effectively. However, these technologies are not necessarily ideal from the points of view of energy consumption, environmental impacts and safety in working circumstances. In * Corresponding author. Tel.: + 81-75-724-7567. E-mail address: [email protected] (K. Sawada).
recent years, utilization of enzyme for textile processing is focused to improve these problems (Minagawa, 1985; Tanida, 1994; Hartzell and Hsieh, 1997). For example, investigation of the scouring of cotton using pectinase enzyme, ‘Bioscouring’, is one of the interesting projects (Heine and Hocker, 1995). Effective removal of pectin and wax from raw cotton substrate with the enzyme under the mild conditions will provide high quality products for the subsequent dyeing and finishing processes with less energy consumed under safer conditions. In addition, modification of wool by a enzymatic
0168-1656/01/$ - see front matter © 2001 Elsevier Science B.V. All rights reserved. PII: S 0 1 6 8 - 1 6 5 6 ( 0 1 ) 0 0 3 1 0 - 8
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process is also a favorable process (Heine and Hocker, 1995). Wool fabrics have a disadvantage to felt by rubbing under wet conditions. It is generally well known that felting of wool fabrics is caused by the tangle between the cuticles at external surface of wool fiber. Moderate removal of external surface region, therefore, will prevent such shortcomings. Based on these concepts, many investigations for the wool modification using enzymes have been carried out. However, the new technology for the wool modification using enzymes has not been established as effective to replace the conventional process. In our previous study, we have reported that pectinase enzyme in aqueous solution has excellent ability to scour cotton fabrics when small amount of organic solvent and mixed surfactants are present in the system (Sawada et al., 1998). However, some problems such as the necessities of prolonged treatment and of high concentration of enzyme have still been unsolved. It is well known that some kinds of surfactants form reverse micelle in non-polar solvent. The reverse micellar system has excellent properties to solubilize relatively large amounts of water at the interior of reverse micelle and to provide stable aqueous microenvironment, so-called water-pool in the non-aqueous media. Hydrophilic substances such as enzymes and dyes can also be solubilized in the water-pool. Many investigators have reported that the enzymes solubilized in the interior of reverse micelles showed their activities even in such a complicated system (Martinek et al., 1986; Shield et al., 1986; Luisi et al., 1979; Menger and Yamada, 1979). If pectinase and other enzymes that are suitably designed for textile processing display their activities in such reverse micellar solution, it can be expected that this enzyme system may have high potential for replacement of conventional practical textile processes. In this paper, we report the results of investigation with respect to the bioscouring of cotton and the modification of wool using enzymes in the reverse micellar system. In addition, we report the results of investigation of the possibilities of utilization of the same system for the subsequent textile dyeing process.
2. Materials and methods
2.1. Chemicals Surfactant used in this study was sodium bis-2ethylhexylsulpho-succinate (Aerosol-OT, AOT). AOT was obtained from Nacalai Tesque Co., Ltd and was used without further purification. Initial water content in AOT was found to be 0.7% (w/w) through Karl Fisher titration. The quantity of solubilized water in the reverse micellar solution was shown by the molar ratio of injected water to surfactant, that is, w0 = [H2O]/[surfactant]. Maximal w0 value without phase separation of AOT reverse micellar system attained in this study is ca. 60. Isooctane (Nacalai Tesque Co., Ltd.) and all other chemicals used in this study were of reagent grade. Cotton fabrics used in this study for the bioscouring were gray goods (Shikisen-sha Co., Ltd., Japan). All cotton fabrics were pretreated in boiling water for 1 h before scouring. Wool muslin used in this study was obtained from Shikisen-sha Co., Ltd. and was used without pretreatment. Pectinase (Aspergillus niger EC 3.2.1.15) used for the bioscouring of cotton was obtained from Tokyo Chemical Industry Co., Ltd. Protease, ‘Bioprase APL-30’ (Bacillus subtilis) used for the wool modification was obtained from Nagase Biochemical Co., Ltd. These enzymes were used without further purification. Water-pool in the reverse micelle were buffered with 0.1 M CH3COOH/ CH3COONa (pH 4) and 0.1 M KH2PO4/NaOH (pH 7) for pectinase and protease, respectively.
2.2. Bioscouring of cotton with pectinase enzyme AOT reverse micellar solutions were prepared by an injection of prescribed volumes of aqueous stock solution either in buffer alone or enzyme in buffer to 0.05 M AOT/isooctane solutions. After an injection of aqueous stock solution, reverse micellar solutions were gently stirred for a few minutes until the solutions became transparent. After preparation of AOT reverse micellar solution, cotton fabric specimens were then put in the solution. The experiment of bioscouring was car-
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ried out at 40 °C for 0.5– 24 h, liquor ratio 50:1. Alkali scouring was carried out with 20% o.w.f. (on the weight fiber) sodium hydroxide solution containing 2 g l − 1 surfactant (Triton X-100) at 100 °C, liquor ratio 20:1. Weight loss was evaluated by measuring absolute dry weight of the fabric before and after treatment with a high-temperature electric balance (Chyo Balance Co., MC30MB). Residual cotton wax was evaluated by extraction method (JISL 1096) with carbon tetrachloride solvent using Soxhlet apparatus. Removal of pectic substances was estimated from dye uptake of Methylene Blue, measuring dye concentration with a Shimadzu UV-200S doublebeam spectrophotometer.
2.3. Modification of wool with pectinase enzyme Preparations of AOT reverse micellar solution and evaluations of weight loss were the same as described above. The experiment was carried out at 40 °C for 0.5–5 h, liquor ratio 100:1. Tear strength test of wool fabric was carried out following venjuram method with an Elmendorf Tearing Tester. Felting property of wool fabric was evaluated by measuring the length of wool fabrics before and after heavy agitation in hot water (70 °C) for 0.5 h.
2.4. Textile dyeing in re6erse micellar solution Dye liquors were prepared by an injection of prescribed volumes of aqueous dye solution to 0.05 M AOT/isooctane solutions. Dyeing of cotton fabrics were carried out with direct dye (CI Direct Red 28, Tokyo Chemical Industry Co., Ltd.) under the conditions at 40 °C for 24 h. Dyeing of polyester fabrics were carried out with disperse dye (C.I. Disperse Violet 1, Tokyo Chemical Industry Co., Ltd.) under the condition at 130 °C for 1 h. Dyeing of cotton– polyester fabrics with reactive dye (C.I. Reactive Red 2, Nippon Kayaku Co., Ltd.) and disperse dye (C.I. Disperse Violet 1) were carried out in one-bath dyeing process. Dye liquor that contains both reactive dye and disperse dye was kept at 40 °C for 2 h at first for the dyeing of cotton fabrics. After the process of dyeing of cotton, temperature of dye
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liquor was raised to 130 °C for the dyeing of polyester. In this process, fixation of reactive dye to cotton fabric can be completed.
3. Results and discussions
3.1. Bioscouring of cotton with pectinase enzyme Fig. 1 compares the weight loss of fabrics with treatment time treated by bioscouring in reverse micellar system and by conventional alkaline scouring. In a preliminary experiment, we have found that weight of fabrics decreased by ca. 1% when merely shaken in water, mainly due to falling off of yarn waste at the edge of fabrics. Weight loss of cotton fabrics was also observed in the treatments of reverse micellar system with buffer solution rather than using isooctane solvent alone, indicating that fuzz, yarn waste and cotton wax on the cotton fabrics may be removed by those solvent treatments. In the process of reverse micellar system with pectinase enzyme, on the other hand, weight of fabrics gradually decreases with increasing treating time. Pectic substances on the cotton fabrics seem to be removed by the enzymatic hydrolysis. The weight of the bioscoured fabrics falls to an almost constant 97.5–98.5% of the weight of raw cotton fabric. It should be noted here that scouring effect could be observed obviously even in very low enzyme con-
Fig. 1. Weight losses of cotton fabrics by scouring.
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Fig. 2. Reduction of wax from cotton fabric by scouring.
centration as 0.1 g l − 1. As reported in the previous paper (Sawada et al., 1998), it required at least 10 g l − 1 of pectinase in order to bring about similar performance in aqueous system. Pectinase in reverse micellar system seems to have superior activity compared to aqueous system. Great weight loss observed by conventional alkaline scouring process can be attributed to the excess removal of cellulosic backbone by strong alkali process, which simultaneously occurred with the removal of pectic substances by strong alkaline solution. Fig. 2 shows residual cotton wax on the fabrics through the treatment against the treating time. As Fig. 2 shows, the amount of residual cotton wax on the fabrics treated by aqueous buffer solution (aqueous process without enzyme) are nearly constant at 0.4%, indicating that cotton wax are not removed by the process in aqueous solution without enzyme. In contrast, in the processes of conventional alkaline scouring and of reverse micellar system with enzyme, as well as even without enzyme, cotton wax is almost completely removed. Treatment with isooctane alone was less successful, although the differences between this and the reverse micellar system or conventional alkaline scouring decreased greatly with prolonged treatment. It seems that reverse micellar system is as effective as the conventional alkaline process in removing cotton wax. Since the effect of the treatments in the reverse micellar system with and without pectinase enzyme is similar, the enzyme seems not to be vital for the removal of cotton wax in this system. Regarding the total effectiveness of the scouring process in reverse micellar system, there is concern about the
effect of the process on the removal of pectic substances from fabrics. Pectic substances are known to inhibit dye adsorption and to be one of the factors of irregular dyeing in the practical dyeing processes. Removal of pectic substances, therefore, is also one of the important purposes of the scouring of cotton. Fig. 3 compares residual pectic substance on the cotton fabrics after the scouring process. Residual pectic substance is expressed as MB value, which is evaluated in an arbitrary unit by measuring the equilibrium adsorption of Methylene Blue (C.I. Basic Blue 9) on the scoured cotton fabrics from aqueous solution (10 − 3 mol l − 1) as described in the previous paper (Sawada et al., 1998). This evaluation method is based on the stoichiometric interaction between dye cation and carboxylate anion of pectin. Therefore, removal of pectic substance from cotton fabrics will bring about the diminution of dye uptake of Methylene Blue, i.e. diminution of MB value of fabrics. As Fig. 3 shows, MB-values of fabrics treated by reverse micellar system with enzyme are remarkably decreased compared to those of others. In contrast, effect of the treatments in reverse micellar system without enzyme is considerably poor compared with those with enzyme while the process also shows the positive effect. The difference between them obviously indicates that the pectinase is active in reverse micellar system. Effectiveness of scouring process in reverse micellar system with pectinase is equivalent or better than that of conventional alkaline scouring. Satisfactory scouring effect could be achieved with remarkably low enzyme concentration as 0.1 g l − 1, which is 100 times lower than
Fig. 3. MB value of scoured cotton fabric.
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Fig. 4. Weight losses of wool fabrics by protease treatment.
the case of the bioscouring process in aqueous system (Sawada et al., 1998). From the results mentioned above, total effectiveness of the bioscouring of cotton with pectinase enzyme could be remarkably improved by the use of the reverse micellar system. If other enzymes in the reverse micellar system also perform an excellent activity as pectinase, this system may have possibility as a new treatment media for various textile processes.
3.2. Modification of wool with protease enzyme Fig. 4 compares weight loss of wool fabrics treated with protease enzyme (‘bioprase’) in aqueous and in reverse micellar system. In both systems, similar weight losses can be observed at the initial stage of the treatment. A part of keratin that is the main component of wool fabric would be removed by the hydrolysis of protease catalytic reaction. However, weight losses of wool fabrics treated in reverse micellar solution become constant after 1 h treatment. These apparent less activities of the protease in reverse micellar solution may be caused by a product inhibition of hydrolyzates, i.e. amino acids and their oligomers, in the reverse micellar solution. Dissolution of the hydrolyzates into the reverse micellar solution would be limited because these substances must be located in the water-pool of the micelle. Waterpool may be saturated with the hydrolyzates after 1 h treatment. Fig. 5 compares shrinkage percentages of treated wool fabrics. Remarkable decrease of shrinkage percentages can be observed even in
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Fig. 5. Felting property of enzyme treated wool.
0.5% weight loss. These results suggest that protease enzyme hydrolyzes cuticles at the external surface of wool fiber. In both aqueous and reverse micellar system, effects seem to be similar. In order to investigate the action of protease treatment on the fabrics in detail, evaluation of tear strength of the treated fabrics has been carried out and shown in Fig. 6. Increasing the weight loss decreases tear strength. If protease hydrolyzes only the external surface of wool fiber, tear strength of the treated fabrics should not depend on the degree of the weight loss. However, from the results shown in Fig. 6, it suggests that protease hydrolyzes mainly the inside of the fiber rather than cuticle. Those remarkable declines of the tear strength by the treatment would be the important subjects to be solved with further studies. Similarly in the case of shrinkage percentages, the effects on the tear strength are nearly equal in both aqueous and reverse micellar systems.
Fig. 6. Tear strength of enzyme treated wool.
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Fig. 7. Adsorption isotherm of CI Direct Red 28 on cotton in aqueous and reverse micellar system at 40 °C.
From these results, it can be concluded that total effectiveness of protease for the modification of wool in reverse micellar system is not necessarily excellent compared to those in aqueous system. However, it is very interesting that protease maintains its activities even in non-aqueous media. Further detailed investigations for the selective hydrolysis of cuticle and the selection of the treatment condition where protease shows excellent activity must be necessary.
3.3. Dyeing of textiles in re6erse micellar system Fig. 7 shows adsorption isotherm of direct dye (C.I. Direct Red 28) onto cotton fabrics in reverse micellar solution and in conventional aqueous solution without electrolyte. As can be seen in Fig. 7, adsorption of direct dye onto cotton fabrics is much higher than that in normal aqueous solution. This result indicates that reverse micellar solution can be used as a good dyeing media, not only for the enzymatic textile processing. High exhaustions of dye in reverse micellar system may be attributed to the very low bath-ratio (waterfabric ratio) compared to conventional aqueous dyeing system. Dye exhaustion in reverse micellar solution slightly depends on w0 value in the system. The system that is higher w0 value (higher amount of water) seems to be suitable for the dyeing process. The swelling of cotton by the solubilized water may influence the dye penetration into the fabrics. In the case of dyeing in reverse micellar solution, direct dye would be solubilized in water-
pool. Therefore, similar dyeing processes can be estimated in analogy if hydrophilic fabrics are dyed with other water-soluble dyes, such as acid dyes and reactive dyes. In contrast, water-insoluble dyes such as disperse dyes that is used for dyeing polyester fabrics can be estimated that they are not solubilized in the water-pool but solubilized outside of the pool. In order to evaluate the possibility of dyeing polyester fabrics with such water-insoluble dye in the reverse micellar system, investigations have been carried out using a disperse dye as a dye and polyester fabrics as a substrate. Fig. 8 compares adsorption of disperse dye (C.I. Disperse Violet 1) onto polyester fabrics in aqueous and reverse micellar solution. Dyeabilities of polyester fabrics with disperse dye were evaluated as color depth, i.e. K/S value, calculated through Kubelka–Munk equation. As Fig. 8 shows, disperse dye in non-aqueous media have enough ability to dye polyester fabrics. Disperse dye dispersed in organic solvent (in isooctane, not in water-pool) may exhibit a similar behavior to the aqueous disperse dyeing system. From the results described above, it can be expected that cotton– polyester blends or composites can be dyed simultaneously both with reactive and disperse dyes in the reverse micellar solution at the same time. Fig. 9 shows color depth of polyester and cotton fabrics dyed with disperse (C.I. Disperse Violet 1) and reactive dye (C.I. Reactive Red 2), respectively. Both fabrics show satisfacto-
Fig. 8. Adsorption of CI Disperse Violet 1 on polyester from water, isooctane and reverse micellar system at 130 °C.
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and dyeing can further be combined at one-step. Since the construction of continuous one-step process of textile processing and dyeing process is the ultimate subject for the textile industry, reverse micellar system would be one of the most expected candidate system for that purpose. Further accumulation of advanced knowledge would be necessary for extend the possibilities of the practical application of reverse micellar system in textile industry.
References Fig. 9. Dyeing of cotton-polyester with reactive and disperse dyes from reverse micellar system at 40 – 130 °C.
rily deep color shade, indicating that one-bath dyeing of cotton– polyester blends can be achieved if reverse micellar solution is used. Reactive dye adsorbs on cotton from the inside of the reverse micelle and disperse dye does on polyester fabrics from non-aqueous organic solvent (isooctane, outside of reverse micelle) at the same time. Fixation of the reactive dye on cotton can be completed at the process of dyeing of polyester fabrics under high temperature condition. Stable coexistences of both reactive and disperse dye at the inside and outside of micelle may bring about possible one-bath dyeing of cotton– polyester blends. From the results of the study, reverse micellar system has high potential for the application not only to textile processing but also to dyeing. If enzymes in water-pool maintain their activities under the coexistence of dye, the textile processing
.
Hartzell, M.M., Hsieh, Y-L., 1997. Book of Abstracts, 213th ACS National Meeting, CELL-026. Heine, E., Hocker, H., 1995. Enzyme treatments for wool and cotton. Rev. Prog. Color. 25, 57 – 63. Luisi, P.L., Bonner, F.J., Pellegrini, A., Wiget, P., Wolf, R., 1979. Micellar solubilization of proteins in aprotic solvents and their spectroscopic characterization. Helv. Chim. Acta 62, 740 – 753. Martinek, K., Levashov, A.V., Klyachko, N., Khmelnitski, Y.L., Berezin, I.V., 1986. Micellar enzymology. Eur. J. Biochem. 155, 453 – 468. Menger, F.M., Yamada, L., 1979. Enzyme catalysis in water pools. J. Am. Chem. Soc. 101, 6731 – 6734. Minagawa, M., 1985. Study on the detergency of protein stains. J. Jpn. Res. Assn. Text. End-Use 26, 322 – 326. Sawada, K., Tokino, S., Ueda, M., 1998. Bioscouring of cotton with pectinase enzyme. J. Soc. Dye. Col. 114, 333 – 335. Shield, J.W., Ferguson, H.D., Bommarius, A.S., Hatton, T.A., 1986. Enzymes in reversed micelles as catalysts for organicphase synthesis reactions. Ind. Eng. Chem. Fundam. 25, 603 – 612. Tanida, O., 1994. Enzyme treatment on cellulose fiber for weight reduction and modification. Sen’i Gakkaishi 50, 75 – 79.
Enzyme processing of textiles in reverse micellar solution Kazuya Sawada a,*, Mitsuo Ueda b a
Japan Society for the Promotion of Science, Faculty of Engineering and Design, Kyoto Institute of Technology, Matsugasaki, Sakyo-Ku, 606 -8585, Japan b Faculty of Textile Science, Kyoto Institute of Technology, Matsugasaki, Sakyo-Ku, 606 -8585, Japan Received 26 June 2000; received in revised form 9 January 2001; accepted 15 January 2001
Abstract Scouring of cotton using pectinase enzyme, bioscouring, in reverse micellar system was studied. The effectiveness of bioscouring was evaluated by measuring weight loss of cotton, analyzing pectin and cotton wax remaining and by wetness testing. Pectinase enzyme showed excellent activity even in organic media, and the effectiveness of scouring was equivalent or better than that achieved by conventional alkaline process or bioscouring in aqueous media. Enzymatic modification of wool using protease enzyme in the same system was also studied. It has found that felting property and tensile strength of wool fabrics treated by protease in reverse micellar system were superior to those in aqueous media. Possibilities of utilization of the same system for the subsequent textile dyeing process were also investigated. It was found that cotton and polyester fabrics were dyed satisfactorily by reverse micellar system compared to conventional aqueous system. © 2001 Elsevier Science B.V. All rights reserved. Keywords: Reverse micelle; Bioscouring; Pectinase; Protease; Dyeing
1. Introduction For the past few decades, various kinds of dyeing and textile processing technologies have been developed and the processes have become performed effectively. However, these technologies are not necessarily ideal from the points of view of energy consumption, environmental impacts and safety in working circumstances. In * Corresponding author. Tel.: + 81-75-724-7567. E-mail address: [email protected] (K. Sawada).
recent years, utilization of enzyme for textile processing is focused to improve these problems (Minagawa, 1985; Tanida, 1994; Hartzell and Hsieh, 1997). For example, investigation of the scouring of cotton using pectinase enzyme, ‘Bioscouring’, is one of the interesting projects (Heine and Hocker, 1995). Effective removal of pectin and wax from raw cotton substrate with the enzyme under the mild conditions will provide high quality products for the subsequent dyeing and finishing processes with less energy consumed under safer conditions. In addition, modification of wool by a enzymatic
0168-1656/01/$ - see front matter © 2001 Elsevier Science B.V. All rights reserved. PII: S 0 1 6 8 - 1 6 5 6 ( 0 1 ) 0 0 3 1 0 - 8
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process is also a favorable process (Heine and Hocker, 1995). Wool fabrics have a disadvantage to felt by rubbing under wet conditions. It is generally well known that felting of wool fabrics is caused by the tangle between the cuticles at external surface of wool fiber. Moderate removal of external surface region, therefore, will prevent such shortcomings. Based on these concepts, many investigations for the wool modification using enzymes have been carried out. However, the new technology for the wool modification using enzymes has not been established as effective to replace the conventional process. In our previous study, we have reported that pectinase enzyme in aqueous solution has excellent ability to scour cotton fabrics when small amount of organic solvent and mixed surfactants are present in the system (Sawada et al., 1998). However, some problems such as the necessities of prolonged treatment and of high concentration of enzyme have still been unsolved. It is well known that some kinds of surfactants form reverse micelle in non-polar solvent. The reverse micellar system has excellent properties to solubilize relatively large amounts of water at the interior of reverse micelle and to provide stable aqueous microenvironment, so-called water-pool in the non-aqueous media. Hydrophilic substances such as enzymes and dyes can also be solubilized in the water-pool. Many investigators have reported that the enzymes solubilized in the interior of reverse micelles showed their activities even in such a complicated system (Martinek et al., 1986; Shield et al., 1986; Luisi et al., 1979; Menger and Yamada, 1979). If pectinase and other enzymes that are suitably designed for textile processing display their activities in such reverse micellar solution, it can be expected that this enzyme system may have high potential for replacement of conventional practical textile processes. In this paper, we report the results of investigation with respect to the bioscouring of cotton and the modification of wool using enzymes in the reverse micellar system. In addition, we report the results of investigation of the possibilities of utilization of the same system for the subsequent textile dyeing process.
2. Materials and methods
2.1. Chemicals Surfactant used in this study was sodium bis-2ethylhexylsulpho-succinate (Aerosol-OT, AOT). AOT was obtained from Nacalai Tesque Co., Ltd and was used without further purification. Initial water content in AOT was found to be 0.7% (w/w) through Karl Fisher titration. The quantity of solubilized water in the reverse micellar solution was shown by the molar ratio of injected water to surfactant, that is, w0 = [H2O]/[surfactant]. Maximal w0 value without phase separation of AOT reverse micellar system attained in this study is ca. 60. Isooctane (Nacalai Tesque Co., Ltd.) and all other chemicals used in this study were of reagent grade. Cotton fabrics used in this study for the bioscouring were gray goods (Shikisen-sha Co., Ltd., Japan). All cotton fabrics were pretreated in boiling water for 1 h before scouring. Wool muslin used in this study was obtained from Shikisen-sha Co., Ltd. and was used without pretreatment. Pectinase (Aspergillus niger EC 3.2.1.15) used for the bioscouring of cotton was obtained from Tokyo Chemical Industry Co., Ltd. Protease, ‘Bioprase APL-30’ (Bacillus subtilis) used for the wool modification was obtained from Nagase Biochemical Co., Ltd. These enzymes were used without further purification. Water-pool in the reverse micelle were buffered with 0.1 M CH3COOH/ CH3COONa (pH 4) and 0.1 M KH2PO4/NaOH (pH 7) for pectinase and protease, respectively.
2.2. Bioscouring of cotton with pectinase enzyme AOT reverse micellar solutions were prepared by an injection of prescribed volumes of aqueous stock solution either in buffer alone or enzyme in buffer to 0.05 M AOT/isooctane solutions. After an injection of aqueous stock solution, reverse micellar solutions were gently stirred for a few minutes until the solutions became transparent. After preparation of AOT reverse micellar solution, cotton fabric specimens were then put in the solution. The experiment of bioscouring was car-
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ried out at 40 °C for 0.5– 24 h, liquor ratio 50:1. Alkali scouring was carried out with 20% o.w.f. (on the weight fiber) sodium hydroxide solution containing 2 g l − 1 surfactant (Triton X-100) at 100 °C, liquor ratio 20:1. Weight loss was evaluated by measuring absolute dry weight of the fabric before and after treatment with a high-temperature electric balance (Chyo Balance Co., MC30MB). Residual cotton wax was evaluated by extraction method (JISL 1096) with carbon tetrachloride solvent using Soxhlet apparatus. Removal of pectic substances was estimated from dye uptake of Methylene Blue, measuring dye concentration with a Shimadzu UV-200S doublebeam spectrophotometer.
2.3. Modification of wool with pectinase enzyme Preparations of AOT reverse micellar solution and evaluations of weight loss were the same as described above. The experiment was carried out at 40 °C for 0.5–5 h, liquor ratio 100:1. Tear strength test of wool fabric was carried out following venjuram method with an Elmendorf Tearing Tester. Felting property of wool fabric was evaluated by measuring the length of wool fabrics before and after heavy agitation in hot water (70 °C) for 0.5 h.
2.4. Textile dyeing in re6erse micellar solution Dye liquors were prepared by an injection of prescribed volumes of aqueous dye solution to 0.05 M AOT/isooctane solutions. Dyeing of cotton fabrics were carried out with direct dye (CI Direct Red 28, Tokyo Chemical Industry Co., Ltd.) under the conditions at 40 °C for 24 h. Dyeing of polyester fabrics were carried out with disperse dye (C.I. Disperse Violet 1, Tokyo Chemical Industry Co., Ltd.) under the condition at 130 °C for 1 h. Dyeing of cotton– polyester fabrics with reactive dye (C.I. Reactive Red 2, Nippon Kayaku Co., Ltd.) and disperse dye (C.I. Disperse Violet 1) were carried out in one-bath dyeing process. Dye liquor that contains both reactive dye and disperse dye was kept at 40 °C for 2 h at first for the dyeing of cotton fabrics. After the process of dyeing of cotton, temperature of dye
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liquor was raised to 130 °C for the dyeing of polyester. In this process, fixation of reactive dye to cotton fabric can be completed.
3. Results and discussions
3.1. Bioscouring of cotton with pectinase enzyme Fig. 1 compares the weight loss of fabrics with treatment time treated by bioscouring in reverse micellar system and by conventional alkaline scouring. In a preliminary experiment, we have found that weight of fabrics decreased by ca. 1% when merely shaken in water, mainly due to falling off of yarn waste at the edge of fabrics. Weight loss of cotton fabrics was also observed in the treatments of reverse micellar system with buffer solution rather than using isooctane solvent alone, indicating that fuzz, yarn waste and cotton wax on the cotton fabrics may be removed by those solvent treatments. In the process of reverse micellar system with pectinase enzyme, on the other hand, weight of fabrics gradually decreases with increasing treating time. Pectic substances on the cotton fabrics seem to be removed by the enzymatic hydrolysis. The weight of the bioscoured fabrics falls to an almost constant 97.5–98.5% of the weight of raw cotton fabric. It should be noted here that scouring effect could be observed obviously even in very low enzyme con-
Fig. 1. Weight losses of cotton fabrics by scouring.
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Fig. 2. Reduction of wax from cotton fabric by scouring.
centration as 0.1 g l − 1. As reported in the previous paper (Sawada et al., 1998), it required at least 10 g l − 1 of pectinase in order to bring about similar performance in aqueous system. Pectinase in reverse micellar system seems to have superior activity compared to aqueous system. Great weight loss observed by conventional alkaline scouring process can be attributed to the excess removal of cellulosic backbone by strong alkali process, which simultaneously occurred with the removal of pectic substances by strong alkaline solution. Fig. 2 shows residual cotton wax on the fabrics through the treatment against the treating time. As Fig. 2 shows, the amount of residual cotton wax on the fabrics treated by aqueous buffer solution (aqueous process without enzyme) are nearly constant at 0.4%, indicating that cotton wax are not removed by the process in aqueous solution without enzyme. In contrast, in the processes of conventional alkaline scouring and of reverse micellar system with enzyme, as well as even without enzyme, cotton wax is almost completely removed. Treatment with isooctane alone was less successful, although the differences between this and the reverse micellar system or conventional alkaline scouring decreased greatly with prolonged treatment. It seems that reverse micellar system is as effective as the conventional alkaline process in removing cotton wax. Since the effect of the treatments in the reverse micellar system with and without pectinase enzyme is similar, the enzyme seems not to be vital for the removal of cotton wax in this system. Regarding the total effectiveness of the scouring process in reverse micellar system, there is concern about the
effect of the process on the removal of pectic substances from fabrics. Pectic substances are known to inhibit dye adsorption and to be one of the factors of irregular dyeing in the practical dyeing processes. Removal of pectic substances, therefore, is also one of the important purposes of the scouring of cotton. Fig. 3 compares residual pectic substance on the cotton fabrics after the scouring process. Residual pectic substance is expressed as MB value, which is evaluated in an arbitrary unit by measuring the equilibrium adsorption of Methylene Blue (C.I. Basic Blue 9) on the scoured cotton fabrics from aqueous solution (10 − 3 mol l − 1) as described in the previous paper (Sawada et al., 1998). This evaluation method is based on the stoichiometric interaction between dye cation and carboxylate anion of pectin. Therefore, removal of pectic substance from cotton fabrics will bring about the diminution of dye uptake of Methylene Blue, i.e. diminution of MB value of fabrics. As Fig. 3 shows, MB-values of fabrics treated by reverse micellar system with enzyme are remarkably decreased compared to those of others. In contrast, effect of the treatments in reverse micellar system without enzyme is considerably poor compared with those with enzyme while the process also shows the positive effect. The difference between them obviously indicates that the pectinase is active in reverse micellar system. Effectiveness of scouring process in reverse micellar system with pectinase is equivalent or better than that of conventional alkaline scouring. Satisfactory scouring effect could be achieved with remarkably low enzyme concentration as 0.1 g l − 1, which is 100 times lower than
Fig. 3. MB value of scoured cotton fabric.
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Fig. 4. Weight losses of wool fabrics by protease treatment.
the case of the bioscouring process in aqueous system (Sawada et al., 1998). From the results mentioned above, total effectiveness of the bioscouring of cotton with pectinase enzyme could be remarkably improved by the use of the reverse micellar system. If other enzymes in the reverse micellar system also perform an excellent activity as pectinase, this system may have possibility as a new treatment media for various textile processes.
3.2. Modification of wool with protease enzyme Fig. 4 compares weight loss of wool fabrics treated with protease enzyme (‘bioprase’) in aqueous and in reverse micellar system. In both systems, similar weight losses can be observed at the initial stage of the treatment. A part of keratin that is the main component of wool fabric would be removed by the hydrolysis of protease catalytic reaction. However, weight losses of wool fabrics treated in reverse micellar solution become constant after 1 h treatment. These apparent less activities of the protease in reverse micellar solution may be caused by a product inhibition of hydrolyzates, i.e. amino acids and their oligomers, in the reverse micellar solution. Dissolution of the hydrolyzates into the reverse micellar solution would be limited because these substances must be located in the water-pool of the micelle. Waterpool may be saturated with the hydrolyzates after 1 h treatment. Fig. 5 compares shrinkage percentages of treated wool fabrics. Remarkable decrease of shrinkage percentages can be observed even in
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Fig. 5. Felting property of enzyme treated wool.
0.5% weight loss. These results suggest that protease enzyme hydrolyzes cuticles at the external surface of wool fiber. In both aqueous and reverse micellar system, effects seem to be similar. In order to investigate the action of protease treatment on the fabrics in detail, evaluation of tear strength of the treated fabrics has been carried out and shown in Fig. 6. Increasing the weight loss decreases tear strength. If protease hydrolyzes only the external surface of wool fiber, tear strength of the treated fabrics should not depend on the degree of the weight loss. However, from the results shown in Fig. 6, it suggests that protease hydrolyzes mainly the inside of the fiber rather than cuticle. Those remarkable declines of the tear strength by the treatment would be the important subjects to be solved with further studies. Similarly in the case of shrinkage percentages, the effects on the tear strength are nearly equal in both aqueous and reverse micellar systems.
Fig. 6. Tear strength of enzyme treated wool.
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Fig. 7. Adsorption isotherm of CI Direct Red 28 on cotton in aqueous and reverse micellar system at 40 °C.
From these results, it can be concluded that total effectiveness of protease for the modification of wool in reverse micellar system is not necessarily excellent compared to those in aqueous system. However, it is very interesting that protease maintains its activities even in non-aqueous media. Further detailed investigations for the selective hydrolysis of cuticle and the selection of the treatment condition where protease shows excellent activity must be necessary.
3.3. Dyeing of textiles in re6erse micellar system Fig. 7 shows adsorption isotherm of direct dye (C.I. Direct Red 28) onto cotton fabrics in reverse micellar solution and in conventional aqueous solution without electrolyte. As can be seen in Fig. 7, adsorption of direct dye onto cotton fabrics is much higher than that in normal aqueous solution. This result indicates that reverse micellar solution can be used as a good dyeing media, not only for the enzymatic textile processing. High exhaustions of dye in reverse micellar system may be attributed to the very low bath-ratio (waterfabric ratio) compared to conventional aqueous dyeing system. Dye exhaustion in reverse micellar solution slightly depends on w0 value in the system. The system that is higher w0 value (higher amount of water) seems to be suitable for the dyeing process. The swelling of cotton by the solubilized water may influence the dye penetration into the fabrics. In the case of dyeing in reverse micellar solution, direct dye would be solubilized in water-
pool. Therefore, similar dyeing processes can be estimated in analogy if hydrophilic fabrics are dyed with other water-soluble dyes, such as acid dyes and reactive dyes. In contrast, water-insoluble dyes such as disperse dyes that is used for dyeing polyester fabrics can be estimated that they are not solubilized in the water-pool but solubilized outside of the pool. In order to evaluate the possibility of dyeing polyester fabrics with such water-insoluble dye in the reverse micellar system, investigations have been carried out using a disperse dye as a dye and polyester fabrics as a substrate. Fig. 8 compares adsorption of disperse dye (C.I. Disperse Violet 1) onto polyester fabrics in aqueous and reverse micellar solution. Dyeabilities of polyester fabrics with disperse dye were evaluated as color depth, i.e. K/S value, calculated through Kubelka–Munk equation. As Fig. 8 shows, disperse dye in non-aqueous media have enough ability to dye polyester fabrics. Disperse dye dispersed in organic solvent (in isooctane, not in water-pool) may exhibit a similar behavior to the aqueous disperse dyeing system. From the results described above, it can be expected that cotton– polyester blends or composites can be dyed simultaneously both with reactive and disperse dyes in the reverse micellar solution at the same time. Fig. 9 shows color depth of polyester and cotton fabrics dyed with disperse (C.I. Disperse Violet 1) and reactive dye (C.I. Reactive Red 2), respectively. Both fabrics show satisfacto-
Fig. 8. Adsorption of CI Disperse Violet 1 on polyester from water, isooctane and reverse micellar system at 130 °C.
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and dyeing can further be combined at one-step. Since the construction of continuous one-step process of textile processing and dyeing process is the ultimate subject for the textile industry, reverse micellar system would be one of the most expected candidate system for that purpose. Further accumulation of advanced knowledge would be necessary for extend the possibilities of the practical application of reverse micellar system in textile industry.
References Fig. 9. Dyeing of cotton-polyester with reactive and disperse dyes from reverse micellar system at 40 – 130 °C.
rily deep color shade, indicating that one-bath dyeing of cotton– polyester blends can be achieved if reverse micellar solution is used. Reactive dye adsorbs on cotton from the inside of the reverse micelle and disperse dye does on polyester fabrics from non-aqueous organic solvent (isooctane, outside of reverse micelle) at the same time. Fixation of the reactive dye on cotton can be completed at the process of dyeing of polyester fabrics under high temperature condition. Stable coexistences of both reactive and disperse dye at the inside and outside of micelle may bring about possible one-bath dyeing of cotton– polyester blends. From the results of the study, reverse micellar system has high potential for the application not only to textile processing but also to dyeing. If enzymes in water-pool maintain their activities under the coexistence of dye, the textile processing
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