Enzymatic treatment of wool pre-treated with cetyltrimethylammonium bromide to achieve machine washability

Knitted wool fabric was pre-treated with cetyltrimethylammonium bromide in alkaline conditions in order to remove surface lipid to make the fibre more hydrophilic to enable more efficient subsequent treatment with the enzyme protease. As residual cetyltrimethylammonium bromide on the fibre surface causes an inhibitory effect towards protease, cetyltrimethylammonium bromide has to be removed from the wool. The efficacy of three different anionic surfactants in acid conditions was examined for removing residual cetyltrimethylammonium bromide from the treated wool. If the cetyltrimethylammonium bromide-treated knitted wool was washed with the anionic surfactant, Libraphos HC2A, in acidic conditions, followed by treatment with a chemically modified protease, machine-washable standards could be achieved. The possibility of whether native protease could be used instead of chemically modified protease in achieving shrink-resistance without excessive fibre damage was also studied, exploiting the tendency of residual cetyltrimethylammonium bromide to decrease the activity of the enzyme. It was found that if native protease or native protease in the presence of an enteric polymer was used to treat cetyltrimethylammonium bromide pre-treated wool, an improvement in shrink-resistance without excessive fibre degradation was observed. Machine washability could be achieved by optimizing the proportion of enteric polymer to native protease used to treat cetyltrimethylammonium bromide pre-treated wool. Up-scaling this process showed similar results. The dyeability of the wool with reactive dyes was unaffected by the treatment.     

Modification of Esperase® by covalent bonding to Eudragit® polymers L 100 and S 100 for wool fibre surface treatment

The protease Esperase® was modified by covalent bonding with two grades of a reversible soluble-insoluble co-polymer of methacrylic acid and methyl-methacrylate, namely Eudragit® L 100 and Eudragit® S 100. The optimum reaction conditions and washing protocol were investigated and it was found that Esperase® modified with Eudragit® L 100 showed greater activity than if modified with Eudragit® S 100. This should be expected as there is a greater quantity of active sites, namely carboxyl groups, per mass of Eudragit® L 100 in comparison with Eudragit® S 100 to interact with the enzyme. Gel filtration confirmed that Eudragit® L 100 covalently bonded to Esperase®. Treatment of the modified Esperase® on wool showed that the enzyme modified with Eudragit® L 100 had greater activity towards the wool and appeared more effective in shrink resistant finishing.     

An ethoxylated alkyl phosphate (anionic surfactant) for the promotion of activities of proteases and its potential use in the enzymatic processing of wool

Pretreatments of wool fabrics with cationic, anionic or non-ionic surfactants were investigated to reduce surface tension and improve the wettability of the fibres in order to promote protease activity on the fibres in subsequent processes. Results showed that an ethoxylated alkyl phosphate (specific anionic surfactant) as well as the widely used non-ionic surfactant was compatible with proteases in the enzymatic treatment of wool. There is therefore a potential for using specific anionic surfactants to achieve efficient enzymatic scouring processes.     

Chemical modification of proteases for wool cuticle scale removal

Over the last few decades several enzymatic processes to improve properties of wool fabrics like felting tendency, shrink resistance, dyeing ability and handling characteristics have been described. Previous investigations into the use of proteases to hydrolyse the cuticles at the surface of wool fibres, resulted in high strength and weight losses. Therefore restriction of the enzyme activity to the wool surface or control of enzyme diffusion to the cortex cells is required.

To change the diffusion behaviour of proteases in wool fibres, the soluble polymer PEG was covalently attached to a protease from Bacillus lentus. Modified enzymes with different molecular weights were compared. These modified enzymes retained up to 80% of their activity in the standard assay while hydrolysis of wool fibres was successfully restricted to cuticles, resulting in a 90% decrease in weight losses compared to non-modified enzymes.     

Chemical modification of proteases for wool cuticle scale removal

Over the last few decades several enzymatic processes to improve properties of wool fabrics like felting tendency, shrink resistance, dyeing ability and handling characteristics have been described. Previous investigations into the use of proteases to hydrolyse the cuticles at the surface of wool fibres, resulted in high strength and weight losses. Therefore restriction of the enzyme activity to the wool surface or control of enzyme diffusion to the cortex cells is required.

To change the diffusion behaviour of proteases in wool fibres, the soluble polymer PEG was covalently attached to a protease from Bacillus lentus. Modified enzymes with different molecular weights were compared. These modified enzymes retained up to 80% of their activity in the standard assay while hydrolysis of wool fibres was successfully restricted to cuticles, resulting in a 90% decrease in weight losses compared to non-modified enzymes.     

Treatment of wool fibres with subtilisin and subtilisin-PEG

In this work the diffusion of serine proteases into wool fabrics and yarns was studied. The proteases used were free subtilisin and subtilisin-PEG (the same enzyme that was covalently cross linked to polyethylene glycol). It is shown that the adsorption and diffusion is facilitated by the pre-treatment performed, being the alkaline surfactant washing and bleaching the most effective in what concerns enzyme adsorption. Furthermore, this study suggests that the diffusion of proteases into wool is dependent on the size of the protease. The free enzyme penetrates into wool fibre cortex while the modified bigger enzyme is retained only at the surface, in the cuticle layer. Also, proteins without proteolytic activity do not adsorb considerably on wool due to its hydrophobic nature, therefore the diffusion is facilitated by hydrolytic action.

These results have important practical implications for the establishment of enzymatic wool finishing processes, since they allow for control of the enzyme hydrolysis, which was the major drawback of this environmental friendly option to the conventional chlorine treatments.     

Simultaneous protease and transglutaminase treatment for shrink resistance of wool

A bioprocess for machine washable wool, combining the advantages of both protease and transglutaminase in a simultaneous enzymatic treatment has been developed. This process reduced the felting tendency of woven wool fabrics by 9% at the expense of only 2% weight and tensile strength loss. In contrast to previously described protease-based processes for shrink resistant wool, the anti-felting properties achieved in the simultaneous enzymatic treatment produced insignificant fibre damage, confirmed also by scanning electron images of the fabrics.     

Industrial production of enzyme-modified wool fibers for machine-washable bed coverings

Enzyme technology is explored on wool fibers to prevent shrinkage and consolidation behavior during washing of woolen bed coverings using normal household machine conditions. Enzyme modification of wool fibers after two different pretreatments has been realized on industrial scale. Enlarged proteolytic enzyme by chemical modification was applied successfully to prevent substantial fiber strength loss. Felt-ball analysis of the fibers as obtained from this industrial process showed substantial improvement in felting resistance. Further processing of these enzyme-modified fibers and finally integration in bed covering quilts have been executed successfully on industrial production lines. The observed fiber losses during processing were in the range of 4.5–6% which is comparable with that of nonmodified fibers. The machine-washability of these produced bed covering quilts was tested in a household washing machine using both wool and normal wash programs applied at different temperatures. It appeared that, contrary to the good washing results in terms of shrinkage and consolidation resistance using the wool program at moderate temperatures, this resistance is marginal when washed with the normal washing program with higher mechanical agitation level or with the wool program at elevated temperature. This result was different from that obtained with woolen fabrics and explained by the less-structured organisation of fibers within a fleece.     

AN INVESTIGATION ON KERATIN EXTRACTION FROM WOOL AND FEATHER WASTE BY ENZYMATIC HYDROLYSIS

In this study, the possibility of keratin extraction from wool and feather by an enzymatic treatment along with a reducing agent has been investigated. The effects of different parameters, that is, enzyme loading, type of substrate and surfactant, hydrolysis time, and reducing agent concentration, have been examined in order to optimize the enzymatic hydrolysis. The optimal condition for maximum keratin extraction was attained by making use of 1 g/L sodium dodecyl sulfate (an anionic surfactant) and 2.6% (v/v) protease (Savinase), along with 8.6 and 6.4 g/L sodium hydrogen sulfite (a reducing agent) for wool and feathers, respectively, at liquor to fiber ratio of 25 mL/g for 4 hr. The obtained results indicated higher degradation of wool fiber in comparison with feathers, which might be due to the higher hydrophilic nature of the former. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) patterns revealed that the molecular weights of the extracted proteins from wool and feather were lower than those for the untreated fibers. Scanning electron micrographs showed fibers fibrillation and degradation upon enzymatic treatment. Besides, Fourier-transform infrared (FTIR) spectra indicated no evident changes in the chemical structure of the hydrolyzed fibers. However, wool and feather remainders were mostly composed of α-helix and β-sheets conformations, respectively.     

Promotional effect of 1‐butyl‐3‐methylimidazolium chloride ionic liquid on the enzymatic finishing of wool

To improve the effects of protease finishing on wool, 1‐butyl‐3‐methylimidazolium chloride ionic liquid was employed as a pretreatment reagent. It was found that ionic liquid pretreatment significantly changed the wool surface characteristics. The Allwördern reaction showed that the epicuticle layer was damaged by the ionic liquid, and X‐ray photoelectron spectroscopy analysis further demonstrated that the surface elemental composition was significantly changed. Ionic liquid pretreatment remarkably improved the accessibility of protease to the wool and thus accelerated the hydrolysis rate of keratin. The properties of wool fabric after combined processing were also changed. Dyeability results showed that the color depth was increased but the wet rubbing and washing fastness of wool fabrics showed a decreased half grade. The wettability results demonstrate that the contact angle was further reduced after the comprehensive treatment because of the exposure of more proteins under the fatty‐acid layer. In addition, the shrink proofing of wool fabric was also enhanced after combined processing. In summary, ionic liquid modification presents a promising pretreatment method for protease processing of wool.     

Oxidant and SDS-stable alkaline protease from Bacillus clausii I-52: production and some properties

AIMS: An investigation was carried out on an oxidative and SDS-stable alkaline protease secreted by Bacillus clausii of industrial significance. METHODS AND RESULTS: Maximum enzyme activity was produced when the bacterium was grown in the medium containing (g l-1): soyabean meal, 15; wheat flour, 10; liquid maltose, 25; K2HPO4, 4; Na2HPO4, 1; MgSO4.7H2O, 0.1; Na2CO3, 6. The enzyme has an optimum pH of around 11 and optimum temperature of 60 degrees C. The alkaline protease showed extreme stability towards SDS and oxidizing agents, which retained its activity above 75 and 110% on treatment for 72 h with 5% SDS and 10% H2O2, respectively. Inhibition profile exhibited by phenylmethylsulphonyl fluoride suggested that the protease from B. clausii belongs to the family of serine proteases. CONCLUSIONS: Bacillus clausii produced high levels of an extracellular protease having high stability towards SDS and H2O2. SIGNIFICANCE AND IMPACT OF THE STUDY: The alkaline protease from B. clausii I-52 is significant for an industrial perspective because of its ability to function in broad pH and temperature ranges in addition to its tolerance and stability in presence of an anionic surfactant, like SDS and oxidants like peroxides and perborates. The enzymatic properties of the protease also suggest its suitable application as additive in detergent formulations.     

海南近海水域产碱性蛋白酶菌株的分离筛选及发酵条件优化

【背景】微生物蛋白酶在工业生物技术上具有广阔的应用前景。在微生物蛋白酶中,碱性蛋白酶占全球酶总产量的50%以上,获取产碱性蛋白酶的新微生物资源意义重要。【目的】在海南近海贝类养殖基地海泥中筛选获得高产碱性蛋白酶的菌株,对其生长特性进行探究并优化菌株产酶条件,获得新的蛋白酶生产资源。【方法】以酪素培养基为筛选培养基,采用形态学结合系统发育分析鉴定菌株,通过响应面实验优化菌株的产酶条件。【结果】筛选获得一株高产碱性蛋白酶的菌株F3,鉴定为粘质沙雷氏菌（Serratia marcescens）。菌株在最优产酶条件下发酵酶活达到（339.36±4.30） U/mL。【结论】筛选获得的菌株粘质沙雷氏菌F3有较好的产碱性蛋白酶的能力。     

Bio‐antifelting of wool based on mild methanolic potassium hydroxide pretreatment

Covalently bound lipids cover the wool surface and make enzymatic degradation of wool scales very difficult. In this paper, methanolic potassium hydroxide (MPH) pretreatment was used prior to enzymatic treatment of wool with protease, aiming at hydrolyzing the outmost lipids on the wool surface and promoting the subsequent proteolytic reaction. The efficacy of lipid removal from the fiber surface and the properties of the protease‐treated wool were evaluated. The results indicated that mild MPH pretreatment with 0.10 mol/L MPH for 10 min improved the wettability of the wool without adverse impacts on its mechanical properties. The wetting time and area shrinkage of the wool fabric reached 0.5 s and 5.6%, respectively, and the strength loss was within the acceptable range. Pretreatment with high concentrations of MPH for longer times led to significant damage to the wool fibers and caused heavy strength loss, without improving the antifelting properties after protease treatment. Thus, the combination of mild MPH and protease treatments endowed the wool with desirable properties in contrast to the treatment with protease alone.     

Alkaline protease production by Basidiobolus (N.C.L. 97.1.1): effect of 'darmform' morphogenesis and cultural conditions on enzyme production and preliminary enzyme characterization

A strain of Basidiobolus (N.C.L. 97.1.1) was isolated from plant detritus which secreted alkaline protease optimally active at pH 10.0. It is the first report of a protease from Basidiobolus, which is stable to and active under high alkaline conditions. When incubated under stationary conditions in broth cultures containing salts such as ammonium chloride, 'darmform' morphogenesis was readily induced through enlargement and internal division of the hyphal segments. Secretion of high activity alkaline protease was obtained in cultures initiated with darmform morphogenesis whereas cultures initiated from mycelial inocula grew as large pellets in submerged cultures, with little or no protease secretion. Cultural conditions favoring alkaline protease secretion have been optimized and a preliminary characterization of the enzyme is presented. Compatibility of the alkaline protease with commercial detergents as well as its potential application in recovering silver from spent photographic films have also been investigated.     

Extremozymes for improving wool properties

The project ‘EXTRETEX’ funded by the German Federal Foundation Environment (DBU, Osnabrück, Germany) aims at the improvement of wool properties dyeability, handle, felting behaviour and degree of whiteness by means of enzymes derived from extremophilic micro-organisms. In this paper the effects of a commercial thermo- and alkalistable protease on wool with regard to the degree of whiteness, the dyeability and the felting behaviour are presented. A method to treat wool top and wool fabric was developed on a laboratory scale in which the protease was integrated into the pre-washing step of a dyeing process. This treatment method was than scaled up and tested on an industrial winch beck for fabric. With this method—the addition of enzyme in the pre-washing step—the degree of whiteness is generally enhanced. Dyeing untreated and the enzyme-treated wool with Lanasol Blue 8G leads to an improved dyestuff uptake and a distinctive difference in the colour shade for the latter. Microscopy pictures of fibre cross-sections of these samples display a more even distribution of the dyestuff and a better penetration in the enzyme-treated wool fibres but the colour fastness of the enzyme-treated wool is decreased. Though the felting behaviour of the protease treated wool is significantly improved the felting tendency is still too high for an antifelting finish. An increased damage of the enzyme-treated wool in comparison with the untreated one was not observed.     

Development and industrialisation of enzymatic shrink-resist process based on modified proteases for wool machine washability

There is currently considerable interest in the use of enzymes to achieve a variety of finishing effects on wool, but it is apparent that the extent of fibre degradation by enzymes is of major concern during their commercial application. Proteolytic enzymes are known to penetrate and degrade the internal wool structure during processing, causing fibre damage, rather than limiting the degradation to the cuticle cells. The ability to be able to control the exact location of proteolytic attack on wool protein structures will lead to the successful development of enzymatic treatments for achieving a variety of finishing effects for wool-containing products. This present work describes the modification of proteases so that enzymatic modification of wool fibres is restricted to the cuticle scales of the fibres.

Bulk trials have demonstrated that novel modifications of the enzyme enable the reaction of the enzyme with wool to be controlled, so that less degradation of the wool occurs than in similar treatments with the native protease. An anti-felting effect has been achieved without any significant weight loss being caused by the modified protease during the treatment. This novel enzymatic process leads to environmentally friendly production of machine washable wool.     

Development of an eco-friendly antibacterial textile: lysozyme immobilization on wool fabric

Lysozyme, a type of natural enzyme, has been widely used for bacteriostatic functionalization of various materials due to its efficient and selective antibacterial properties. Herein, we report the preparation and characterization of an eco-friendly antibacterial textile based on the immobilization of lysozyme from chicken egg white onto wool fibers. Tris(hydroxymethyl)phosphine (THP) was employed as the cross-linker for the immobilization of lysozyme on the surface of wool fiber. The mechanism of THP cross-linking was investigated via phosphorus test, energy-dispersive spectroscopy (EDX) and Fourier transform infrared spectroscopy (FT-IR). The surface staining, optimization of immobilization parameters, morphology, antibacterial properties, and durability of wool fibers with immobilized lysozyme were also assessed. The results show that hydroxymethyl groups of THP reacted with amino groups of wool fiber and lysozyme through Mannich reaction, which successfully immobilized lysozyme on the wool fiber. The wool fibers incorporated with lysozyme had better antibacterial properties and durability compared with the untreated wool fabric. This facile immobilization approach of lysozyme provides an effective strategy for environmentally benign modification and functionalization of keratin and keratin-containing materials.

     

Use of protease produced byBacillus sp. SJ-121 for improvement of dyeing quality in wool and silk

In this study, a microorganism-produced protease was used to improve the quality of fabrics. First, the protease-producing bacteria were isolated from soils, and one of them was selected and identified asBacillus sp. SJ-121. The optimal medium composition for its growth and protease production was determined to be as follows: glucose 1 g/L, soybean meal 0.5 g/L, soy peptone 0.5, K₂HPO₄ 0.2, MgSO₄·7H₂O 0.002, Nacl 0.002, and Na₂CO₃ g/L. Also, the optimal temperature for the production of the protease byBacillus sp. SJ-121 was about 40°C at pH 7. The wool and silk were treated with the protease fromBacillus sp. SJ-121. Follwoing the protease treatment, changes in the surface of a single yarm of the fabrics were observed by both an optical microscope and a scanning electron microscope (SEM). Changes in the K/S value of the wool and silk were measured by spectrophotometric analysis, in order to determine the amount of dye uptake in the fabrics. We also performed a tensile strength examination in order to determine the degree and nature of mechanical changes in single yarns of the wool and silk fabrics. By increasing the protease treatment time to 48h, the dyeing characteritics of the fabrics were enhanced, and the surfaces of the single yarns of the fabrics became smoother, due to the removal of soil and scale in them. However, no mechanical changes were detected in the fabrics. Therefore, we suggest that proper treatment of the protease produced byBacillus sp. can improve the quality of silk and wool.     

Effects of cutinase on the enzymatic shrink-resist finishing of wool fabrics

A novel microbial cutinase from Thermobifida fusca WSH04 was applied in the pretreatment of wool fabrics followed by protease treatment, aiming at improving the wettability of the samples by hydrolyzing the outmost bound lipids in the wool surface. Cutinase pretreatment could increase the efficacy of the subsequent protease treatment by improving the wettability, dyeability, and shrink-resistance of the wool fabrics. The data obtained by the XPS method showed the changes of elemental concentration in the wool surface after cutinase pretreatment. Compared with the fabrics treated with hydrogen peroxide and protease, the combination of cutinase and protease treatments produced better results in terms of wettability and shrink-resistance with less strength loss. The anti-felting property of the fabrics treated with the enzymatic resist-shrink technique is very promising to meet the commercial standard.     

Chitosan contribution on wool treatments with enzyme

In a previous research work, it was observed that the application of biopolymer chitosan (CHT) on wool fabrics before the enzymatic treatment promotes an increase of the weight loss. In order to deep on the role played by CHT, several experimental conditions have been selected according to a hybrid experimental design and different parameters, such as weight loss and shrink-resist properties, have been controlled. To enhance the CHT sorption on the wool fibre surface, wool was submitted previously to a water-vapour low-temperature plasma treatment. The weight loss results reveal that the enzyme effect increases by increasing the CHT concentration applied to untreated wool. However, CHT concentration does not have any influence when wool has been previously treated with plasma. It is deduced that the surface free energy of wool fibres plays an important role on the enzyme activity. Therefore, the results obtained reveal that the main contribution of CHT on hydrophobic surface of untreated wool fibres is to confer hydrophilicity to wool. Furthermore, CHT tends to coat the wool fibres by film formation reducing apparently the fibre damage promoted by enzyme treatment and also reducing the wool shrinkage.