【作者】 丁若垚；

【导师】 郁崇文；

【作者基本信息】 东华大学 ， 纺织工程， 2013， 博士

【摘要】 亚麻(拉丁学名：Linum usitatissimum),数千年以来一直是人类优质纤维的来源,如今不仅能用于纺纱织造,也被广泛应用于高性能复合材料等前沿领域。在亚麻纤维的纺纱过程中,存在着一道工序：亚麻粗纱湿纺细纱过程前,需要对粗纱进行脱胶。传统上,亚麻粗纱脱胶使用一种高温碱煮的方法。这种方法,不仅消耗了大量热能,更产生了化学污染的排放。本研究基于现代微生物工程理论,考虑利用生物菌株发酵液或提纯酶液进行亚麻粗纱脱胶的微生物方法,为替代传统化学方法,提供了一种可能。目前苎麻的生物脱胶工艺已经比较成熟而亚麻的生物脱胶手段包括细菌和酶脱胶的研究都相对不够成熟。本研究在前人的基础上,构建技术路线涉及微生物学研究和脱胶工艺研究两部分。传统的微生物培养法,通过稀释梯度涂布和划线分离法,可以获得部分脱胶优势菌种,利用分子生物学手段和生理生化测试对其进行鉴定,同时利用酶学性质进行评价。脱胶工艺研究则揭示了在菌株发酵液深层发酵状况下的脱胶液生理生化变化,为最终工业级发酵水平提供理论基础。首先以亚麻粉为唯一碳源,东海腐烂海草作为微生物来源从富集培养液中分离纯化得10株海洋细菌,10种细菌菌种纤维素酶活较低,木聚糖酶活及果胶酶活较高,都比较适用于亚麻生物煮漂；利用生理生化分析和分子生物学手段,鉴定出10种细菌菌种所属种系。进一步分析表明10株菌株中D3、D4、D8、D10用于亚麻粗纱脱胶,残胶率比较低,并且这几株菌种的脱胶能力都好于现有的商品果胶酶,有进一步研究的潜力。另分离出4株海洋真菌,利用显微镜镜检结果和分子生物学手段,鉴定出4种真菌菌种所属种种系。初步测定真菌发酵液酶活,分析其具有脱胶能力,有待进一步研究。在筛选菌种的基础上构建并优化了亚麻粗纱微生物脱胶的工艺。在摇瓶水平下构建亚麻粗纱的细菌脱胶工艺为：细菌种子→摇瓶活化细菌种子→细菌脱胶液发酵→发酵液粗纱脱胶→湿热灭活后处理。在此基础上聚焦发酵条件和脱胶条件的优化过程。对从海洋环境中筛选到的D3和D4两株菌种的细菌发酵液进行了发酵条件优化。同时对D3和D4两株菌种在得到最优纤纤维性能的发酵条件下制成发酵液的脱胶粗纱进行了纤维性能分析。结果表明生物脱胶效果明显并且柔和。对D3菌种发酵液的脱胶条件进行了优化。在脱胶过程中使得酶活较高的优化条件与使纤维性能较优的优化条件并不相同。要获得较为优化的脱胶后纤维性能,在适当的脱胶基础上,需要一定的高碱性环境和高温环境。细菌脱胶液的酶学性质也在本文中被着重分析。首先对不同发酵水平下D3菌种发酵过程中的生长曲线和酶活曲线进行了测定,发现菌种生长曲线符合微生物典型生长曲线的四期模型,酶活在此过程中存在多个顶峰。进一步分析发现摇瓶水平的菌种浓度和酶活要比发酵水平提前到达顶峰,说明在摇瓶水平条件下发酵液产酶活速率较快。酶活曲线结果说明菌种发酵液的酶活存在好几个顶峰,说明菌种发酵液在脱胶过程中可以被反复利用。进一步研究表明,亚麻粗纱脱胶效果与发酵液酶活力成正相关,与发酵液所含总酶含量也成正相关。相同发酵条件下,使用粗提冻干混合酶与使用发酵脱胶的效果相当。酶量加倍脱胶效果有所提高,但各项物理参数非成倍提高,效果不显著。对于亚麻粗纱生物脱胶的总体效果来说,从发酵液中提取的果胶与木聚糖混合酶,其应用效果要比单一酶效果要好,接近并达到化学脱胶的效果。但如果考虑生产成本,则单纯细菌发酵液也已经能满足生产要求。在实验室的研究基础上,进行了工厂放大实验的研究。放大实验选择了两家工厂进行实验并重复验证实验结果。在浙江金鹰亚麻集团有限公司进行的生物脱胶方案工厂试验,方案在总脱胶时间上优于传统化学脱胶,同时还具备了化学品损耗小、化学污染小和能耗小等优点。从细纱成纱质量看,生物脱胶的纱线质量也不亚于化学脱胶。这说明生物脱胶是完全可以替代传统化学脱胶方法的。在湖北精华纺织集团有限公司实验的结果表明生物脱胶方案在这次试验中也能够用来替代传统化学脱胶,同时生物脱胶在化学灭活前的生物脱胶工艺路线是未来较为适合的生物脱胶放大设计工艺方案。酶与生物发酵液效果差异不大,可以考虑在未来生产中在不适合发酵液的生产工艺中使用生物粗酶。此外在增加化学助剂的基础上,脱胶时间3小时就能满足生产要求。最后总结本研究的结果,展望今后的研究方向。如果能在目前的工作基础上,筛选或通过基因工程手段构建到具有分泌更高活性脱胶酶的菌株,选择到更有利于生物脱胶的脱胶助剂并进一步优化工艺参数,则亚麻粗纱的生物脱胶方法必将得到长足的进步,最终用于工厂生产。更多还原

【Abstract】 Flax(binomial name:Linum usitatissimum) has been a source of high quality fiber for thousands of years. It can be widely used for spinning, weaving, high-performance composite materials and other fields. In processing of flax yarn, traditionally, a high-temperature alkali method was used in flax roving degumming and bleaching. This method both consumes a lot of energy and produces chemical pollution emissions. Based on the theory of modem microbial engineering, the use of microorganism fermentation or purified enzyme is used in degumming of flax roving as an alternative to traditional chemical method.Currently, degumming of ramie has been used in an industrial scale and any other bio-methods used in bast fiber are relatively immature. In this study, microbiology and degumming methods were both researched. For microbiology, microorganism were selected using molecular biology and physiological and biochemical tests. Physiological and biochemical changes in the degumming methods were studied in the deep liquid fermentation conditions. Both microbiology and degumming method research were helpful to build efficient degumming system.Ten strains of marine bacteria were isolated from the enrichment and purified.Since polygalacturonase and xylanasc arc useful in scouring and high cellulase activities damage fiber quality, high polygalacturonase and xylanase and low cellulase activities were chosen as the standard of bacteria scouring. Physiological and biochemical analysis and molecular biology methods were used to identify. Further analysis showed that D3, D4, D8and D10for retting flax roving were better than product pectinase. For further studies, another four strains of marine fungus were also and identified. Determination of fungal fermentation enzyme activity pended further study.Flax roving degumming process was shown:bacterial seed→seed fermentation→bacterial fermentation→degumming→post-processing.D3and D4two strains screened from the marine environment were analyzed. Polyglaeturonase were taken as the enzyme activities. Tenacity and fineness were both considered as properties of fibers(flax bundles) and they were normalized to standardize then given the same weight to define a textile evaluation value as a nondimensional value to measure property of fibers. High temperature(35-40℃) and middle timc(12hours) made good production of polygalcturonase which is the key in scouring and keeping good fiber quality. But it still needs more alkaline for further scouring considering traditionally way is alkaline scouring. Goal programming was used to compute the estimated goal of maximum textile evaluation value. The results of computing indicates that maximum textile evaluation value was at coded factor(01.6820), which is initial pH of9, temperature of40℃, scouring time of12hours. The adequacy of the predicted model was examined by additional independent experiments at the suggested optimal synthesis conditions that flax rovings were scoured to verification that tenacity and fineness was22.78cN/tex and1165Nm respectively and SEM figure also supported the point.Enzyme in degumming was explored. Further analysis revealed that the level of bacteria flask concentration and enzyme activity peaked earlier than the level of fermentation, under the conditions described in the sample fermented liquid level reaches the level unglued faster rate. Activity curve results illustrate the activity of bacteria fermentation broth presence of several peaks, indicating that bacteria in the fermentation broth degumming process can be repeatedly used In the optimal fermentation conditions, the use of ammonium sulfate salting out method of fractional precipitation mainly flax degumming of fermented liquid enzyme, determine the relative enzyme activity get various enzymes ammonium sulfate saturation range, a large number of salting out, and in accordance with the commercial enzyme.Enlarge experiment were carried on on the bias. Test in Zhejiang Golden Eagle Group Co. Showed that the total program time was shorter than traditional chemical degumming and also chemical pollution and energy consumption was low. This shows degumming is completely replace traditional chemical degumming methods. Results of test in Textile Group Co., Ltd. in Hubei showed that the biological degumming solution can also be used to replace traditional chemical degumming, while the chemical degumming before inactivation of biological degumming process route is more appropriate for the future design of biological degumming enlarge technology program. Chemical supplement was favorable for biological degumming and3hours was enough to meet production requirements.In conclusion,based on the screening or through genetic engineering to build higher activity with the secretion of enzyme degumming strains selected to be more conducive degumming degumming auxiliaries and further optimize the process parameters, flax roving biological degumming approach is bound to be great progress, and ultimately used in the factory.更多还原