羊毛生物整理复合酶高产菌的激光选育

采用激光对宇佐美曲霉棕色突变株W25进行诱变处理。选衣到一株产酸性蛋白酶、纤维素CMC酶的复合酶生产菌L86,与出发菌株相比发酵单位酸性蛋白酶提高了42．7％、纤维素CMC酶提高了40．9％。所选衣的L86经多次传代,遗传性状非常稳定。生物整理试验表明,L86复合酶比较适合用作羊毛织物生物整理用酶。     

羊毛生物整理复合酶L86发酵工艺优化研究

采用L9（34）正交试验对L86复合酶生产菌的发酵培养基进行优化、并通过溶氧浓度调节和中期补加蛋白水解液对发酵过程进行调控。结果表明较优的培养基组成为（g/100ml）：黄豆粉4.0、玉米粉1.0、鱼粉0.6、蚕蛹粉0.6、CaCl20.5、NH4Cl1.0、Na2HPO4·2H2O 0.4;通气量控制30h前1:0.5、30-50h1:1.0、50h后1:1.2 v/v/m。在上述条件下摇瓶,酸性蛋白酶酶活10000u/ml、纤维素CMC酶3600u/ml;在0.5m3搅拌罐中扩大中试平均发酵单位酸性蛋白酶5500u/ml、纤维素CMC酶2200u/ml。     

高产复合酶菌株HD-1选育的研究

从产果胶酶的黑曲霉(Aspergillus niger)菌中分离出一支产酸性蛋白酶等多种水解酶的菌株No．3号。经铜蒸汽激光诱变,选育出一支高产复合酶的菌株FID-1,该菌在简单的固体发酵培养基上,28℃培养48hr,每克鲜曲的酶活力(U)为：酸性蛋白酶7500u、纤维素Cx 酶 9061U、纤维素CL酶3581U、果胶酶5471U、糖化酶5582U。酶活平均提高率为38.86％,最高幅度是78.6％。该菌经多次连续传代和贮存一年后,产酶性状稳定。该菌株是目前国内饲料用酶制帮生产株中,酶系最齐全,产酶水平位于前列的优良菌株。     

羊毛生物整理复合酶L86发酵工艺优化研究

采用L9（9^4)正交试验对L86复合酶生产菌的发酵培养基进行优化、并通过溶氧浓度调节和中期补加蛋白水解液对发酵过程进行调控,结果表明罪状优的培养基组成为（g/100ml);黄豆粉4.0、玉米粉1.0、鱼粉0.6、蚕蛹粉0.6、CaCl20.5、NH4Cl1.0、Na2HPO4.2H2O0.4;通气量控制30h前1：0.5、30-50h1:1.0、50h,后1：1.2v/v/m。在上述条件下摇瓶,酸性蛋白酶酶活10000u/ml、纤维素CMC3600u/ml;在0.5m^3搅拌罐中扩大中试平均发酵单位酸性蛋白酶5500u/ml、纤维素CMC酶2200u/ml。     

利用柑橘皮固体发酵生产复合酶菌株的选育

本试验对14个菌株以柑橘皮为主要原料固体发酵生产复合酶的生产性能进行了比较, 发现宇佐美曲霉Aspergillus usaanii具有较好的复合酶产率,为进一步提高该菌株的产酶能力, 我们利用γ-射线辐射对其进行了诱变育种,选育得到一株纤维素酶活提高26%、酸性蛋白酶提高28%、木聚糖酶提高24.5%的突变菌株AU—C33。采用正交试验方法对该菌株的基础培养基进行了优化,结果表明豆粕和尿素含量对各酶活具有极显著影响。以柑橘粉计,当添加23% 豆粕、8%麸皮、3%尿素,水分含量在60%,28℃培养60h后,CMCase、FPA、β-葡萄糖苷酶、酸性蛋白酶和木聚糖酶分别达到了20.8U/g、7.25U/g、74.7U/g、7248.4U/g和3222.6U/g。     

中温（37℃）纤维素分解菌的筛选及混合培养研究

目的:筛选纤维素分解菌,构建复合微生物菌系进行混合培养,获得降解纤维素能力强的复合菌系.方法:从高温阶段堆肥样品、腐熟肥料、牛粪和土壤中筛选出能较好降解纤维素的菌株,进行单独与混合发酵培养,测其酶活.结果:获得降解纤维素能力较强的细菌3株、霉菌4株、放线菌2株.按不同的接种比例构建了4组复合微生物菌系,4个组合的滤纸失重率分别达到41.52%、44.94%、41.82%、37.11%,液体发酵的平均CMC酶活分别为624U/g、988U/g、769U/g、1041U/g,固体发酵的平均CMC酶活分别为4240U/g、5289U/g、4807U/g、5344U/g.结论:综合分析复合菌系滤纸条降解能力、CMC酶活、FAP酶活表明,组合二分解纤维素能力明显强于其他几个组合.     

纤维素分解菌的选育及酶活测定

纤维素是地球上最丰富的有机物质,这些丰富的宝贵资源大部分被浪费了,而且由于部分地区焚烧秸杆造成了严重的环境污染。为了充分利用纤维素,纤维素分解菌的筛选研究逐步展开。通过新华滤纸为唯一碳源的杜氏培养基和刚果红纤维素培养基,从堆肥、污泥、马粪和土壤中分离得到7株纤维素分解菌。以5号菌株为出发菌株,经过紫外线诱变,用刚果红纤维素平板透明圈选育法得到8号菌株。为了评价筛选工作,对8株纤维素分解菌进行酶活测定。结果表明,8号菌株具有最高的CMC酶活和FPA酶活。     

复合水解酶黑曲霉HD—1固体发酵工艺研究

黑曲霉ＨＤ－１是一株分泌高单位酸性蛋白酶、纤维素酶、果胶酶、糖化酶等多种水解酶的生产菌,本研究采用单因素搜索和正交试验对其固体发酵工艺进行优化,并采用中心组合设计进行扩大生产试验,结果表明酶活可达;酸性蛋白酶１．１万Ｕ左右,果胶酶９千Ｕ以上,纤维素ＣＸ酶１．１万Ｕ左右,纤维素ＣＬ酶４千Ｕ以上,糖化酶７千Ｕ左右。     

重离子诱变枯草芽孢杆菌高产复合酶菌种的筛选

重离子诱变作为一种新型高效的辐照诱变方式,具有突变率高、突变谱广、突变体稳定等特点。枯草芽孢杆菌(Bacillus subtilis)因具有多种水解酶活力而广泛应用于工业生产,虽然经过多次诱变和改造,其酶活力不断提高,但其复合酶活力仍有提升空间。同时,多次同种诱变剂的使用容易导致菌种产生"抗性",酶活力提高不大。为此,本研究首次利用不同剂量¹²C⁶⁺重离子束对枯草芽孢杆菌20076进行辐照诱变。诱变后通过透明圈初筛和酶活力验证复筛方式选育出一株复合酶活力较高的菌株B. subtilis KC-1。与原始菌株相比,其发酵液滤纸酶活力、内切葡聚糖酶活力、β-葡萄糖苷酶活力、外切葡聚糖酶活力、蛋白酶活力和α-淀粉酶活力分别提高了20. 2%、79. 6%、98. 2%、7. 4%、10. 4%和27. 4%。11代遗传稳定性实验结果表明该突变株具有良好的稳定性。     

利用黑曲霉固态发酵啤酒糟生产饲料复合酶的研究

以啤酒糟为主要基质,利用黑曲霉固态发酵生产酸性蛋白酶、木聚糖酶和纤维素酶等多种饲料复合酶,研究了黑曲霉固态发酵培养基组成对复合酶酶活的影响,确定最优培养基配方为:啤酒糟75%,麸皮25%,硫酸铵1%,KH_2PO_4 0.2%,MnSO_4 0.1%、ZnSO_4 0.2%,料水比1:2。在适宜的发酵条件下,经30℃发酵5 d,烘干后得到的复合酶制剂中,具有多种酶活性(以干基计)。其中酸性蛋白酶活力3 800 U/g,木聚糖酶活力12 00 U/g和纤维素酶活力18 U/g。     

小单孢菌叠加诱变与发酵过程的研究

采用庆大霉素产生菌—绛红小单孢菌 (M. purpurea) 在培养36h时添加15μg/ml的青霉素,对培养至48h对数生长期的菌体进行紫外线照射+亚硝基胍复合处理,置培养箱中37℃培养5~8d待长出菌落后再用紫外线照射叠加处理,获得一高产菌株N-14。试管发酵生测效价1913u/ml(对照625u/ml);摇瓶发酵培养120h达2178(u/ml),比对照(1243u/ml)提高75.2%;并进行了5L发酵罐发酵培养,发酵过程中测定了发酵液还原糖、总糖、氨基氮、pH、菌体浓度等代谢参数,探索了适合N-14的培养条件,连续5批次生测效价平均1967u/ml,较对照1171u/ml提高了68%, 效果明显。     

Restricting detergent protease action to surface of protein fibres by chemical modification

Due to their excellent properties, such as thermostability, activity over a broad range of pH and efficient stain removal, proteases from Bacillus sp. are commonly used in the textile industry including industrial processes and laundry and represent one of the most important groups of enzymes. However, due to the action of proteases, severe damage on natural protein fibres such as silk and wool result after washing with detergents containing proteases. To include the benefits of proteases in a wool fibre friendly detergent formulation, the soluble polymer polyethylene glycol (PEG) was covalently attached to a protease from Bacillus licheniformis. In contrast to activation of PEG with cyanuric chloride (50%) activation with 1,1′-carbonyldiimidazole (CDI) lead to activity recovery above 90%. With these modified enzymes, hydrolytic attack on wool fibres could be successfully prevented up to 95% compared to the native enzymes. Colour difference (ΔE) measured in the three dimensional colour space showed good stain removal properties for the modified enzymes. Furthermore, half-life of the modified enzymes in buffers and commercial detergents solutions was nearly twice as high as those of the non-modified enzymes with values of up to 63 min. Out of the different modified proteases especially the B. licheniformis protease with the 2.0-kDa polymer attached both retained stain removal properties and did not hydrolyse/damage wool fibres.     

Abatement of microfibre pollution and detoxification of textile dye – Indigo by engineered plant enzymes

Microfibres (diameter <5 mm) and textile dyes released from textile industries are ubiquitous, cause environmental pollution, and harm aquatic flora, fauna, animals and human life. Therefore, enzymatic abatement of microfibre pollution and textile dye detoxification is essential. Microbial enzymes for such application present major challenges of scale and affordability to clean up large scale pollution. Therefore, enzymes required for the biodegradation of microfibres and indigo dye were expressed in transplastomic tobacco plants through chloroplast genetic engineering. Integration of laccase and lignin peroxidase genes into the tobacco chloroplast genomes and homoplasmy was confirmed by Southern blots. Decolorization (up to 86%) of samples containing indigo dye (100 mg/L) was obtained using cp-laccase (0.5% plant enzyme powder). Significant (8-fold) reduction in commercial microbial cellulase cocktail was achieved in pretreated cotton fibre hydrolysis by supplementing cost effective cellulases (endoglucanases, ß-glucosidases) and accessory enzymes (swollenin, xylanase, lipase) and ligninases (laccase lignin peroxidase) expressed in chloroplasts. Microfibre hydrolysis using cocktail of Cp-cellulases and Cp-accessory enzymes along with minimal dose (0.25% and 0.5%) of commercial cellulase blend (Ctec2) showed 88%–89% of sugar release from pretreated cotton and microfibres. Cp-ligninases, Cp-cellulases and Cp-accessory enzymes were stable in freeze dried leaves up to 15 and 36 months respectively at room temperature, when protected from light. Use of plant powder for decolorization or hydrolysis eliminated the need for preservatives, purification or concentration or cold chain. Evidently, abatement of microfibre pollution and textile dye detoxification using Cp-enzymes is a novel and cost-effective approach to prevent their environmental pollution.     

微生物法生产普鲁兰酶的研究

对产普鲁兰酶的出发菌株进行筛选和诱变,使酶活从22．10u／ml提高到了40．77u／ml,酶活提高了84．4％。随后对菌体产酶培养基进行了确定和优化,得到最佳发酵培养基,在此培养基下,菌体产酶达46．76u／ml,为原酶活的114．7％。     

Application of immobilized enzyme technologies for the textile industry: a review

Abstract

A number of enzymes have applications in the textile sector, as several natural fibres (e.g. cotton, wool and flax) can be subjected to processing using such natural biocatalysts. In the latest two decades, demand for industrial enzymes has increased considerably; however, the textile industry requires highly stable enzymes, and good performance at extreme values of pH and temperature. New strategies are continuously emerging for the immobilization of enzymes with superior efficiency and usage. Enzymatic immobilization can stabilize enzymes and extend their useful life. Additionally, reduction in effluent treatment costs and improvement of efficiency is possible. This paper briefly reviews the most recent research efforts pertaining to immobilization of enzymes with potential application in the textile industry.     

产适冷木聚糖酶的海洋产青霉的筛选和诱变

从黄海深层海底泥样中分离到1株产低温木聚糖酶的青霉,经EMS诱变得到11株酶活性提高的菌株,对其中1株产低温木聚糖酶活力最高的菌株产酶性质进行了初步研究。所产木聚糖酶在pH4.6,45℃时酶活可达25.8u/ml,比出发菌株提高126%。诱变后菌株所产木聚糖酶在0℃仍有显著酶活性,达8.2u/ml。     

角质酶/角蛋白酶一浴法处理对羊毛性能的影响

采用T.fusca产角质酶以及Bacillus subtilis产角蛋白酶一浴法的方式处理羊毛,通过毡缩率、断裂强力、碱溶解度、上染速率、K/S值和接触角等指标考察了该处理对羊毛的改性效果,并运用XPS、氨基酸分析和SEM考察了其对羊毛结构与性质的影响。实验结果表明：经一浴法处理后,羊毛织物的毡缩率下降明显,达到机可洗要求;断裂强力下降较少,碱溶解度增加较少,上染速率提高,K/S值增加;XPS分析表明,经处理后羊毛纤维表面的元素含量变化较大;氨基酸分析表明,经处理后羊毛纤维中的胱氨酸质量分数有所降低;SEM显示,羊毛鳞片层大部分被剥除,综上可以说明角质酶/角蛋白酶的一浴法处理对羊毛具有明显的改性作用。     

C．shehatae TZ8耐乙醇菌株的筛选及发酵性能研究

以C.shehataeTZ8为出发茵株,利用1％溶壁酶和1％蜗牛酶酶解1．5h,制备成C.shehataeTZ8原生质体,并对原生质体进行紫外诱变,以含不同浓度乙醇的木糖液体培养基培养进行初筛和复筛,获得一株遗传性能稳定、耐乙醇能力达5．5％（v／v）的蕾株C．shehataeTZ8-4,比初始菌株耐乙醇能力提高了2％。对突变株C．shehataeTZ8-4发酵性能的研究结果表明：C．shehataeTZ8-4发酵糖能力从80g/L（葡糖糖和木糖比为2：1）提高到120g／L,最大乙醇产量从27．41g／L提高到43．12g／L。     

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.