威兰胶发酵及其流变性研究
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摘要
威兰胶是鞘氨醇单胞菌Sphingomonas sp. ZW-3分泌的高分子粘性多糖,与众不同的分子结构赋予了它卓越的流变学性质,因此,在石油开采、建筑混凝土等领域极具开发潜力。
本文旨在以各种低成本原料进行威兰胶发酵研究,实现威兰胶的低成本生产,并探索与其分子量相关的流变学性质。首先,研究不同营养条件下Sphingomonas sp. ZW-3分批发酵及补料分批过程;其次,分析不同营养条件下威兰胶流变学性质,对其平均分子量及其分布、假塑性及粘性、储能模量及损耗模量进行比较,并对样品的抗盐耐温性进行初步探索。论文主要研究成果如下:
首先,在已优化培养条件的基础上利用低成本碳源如甘蔗糖蜜、甘油、果葡糖浆及木薯淀粉开展威兰胶发酵研究。结果表明:前三种碳源的威兰胶产量不够理想,仍需深入研究;而基于木薯淀粉的高温α-淀粉酶、葡萄糖淀粉酶水解规律的分析,不同DE值木薯淀粉水解液为碳源的分批发酵结果表明,DE=87%时威兰胶产量为21.2 g/L,稍高于葡萄糖为碳源时的威兰胶产量(20.2 g/L)。
其次,考察了不同氮源豆粕、玉米浆干粉及复合氮源(硝酸钠和大豆蛋白胨混合物)及碳源(葡萄糖、蔗糖)对威兰胶分批发酵影响。结果表明:与所考察碳源相比,氮源对威兰胶产量影响显著,豆粕为氮源时威兰胶产量最高(约20.0 g/L),玉米浆干粉为氮源时次之(17.0-18.0 g/L),复合氮源为氮源时威兰胶产量约15.0 g/L。补料分批发酵结果表明:初糖浓度10.0 g/L比20.0 g/L对威兰胶合成更有利,发酵72 h的威兰胶产量达到24.8 g/L。
再次,基于不同氮源及碳源的威兰胶分批发酵分析,对不同威兰胶样品的高分子流变学性质进行比较。对各种威兰胶样品溶液进行零剪切粘度测试、稳态流变学测试及动态粘弹性测试,并对所得测定数据进行Carreau模型、Power Law模型拟合及复数粘度(logη*)对(logω)的线性拟合。结果表明:与所考察碳源相比,氮源对威兰胶平均分子量及分子量分布影响显著;产自复合氮源的威兰胶平均分子量最高,分子量分布最窄,储能模量最高;产自氮源豆粕的威兰胶次之,而产自玉米浆干粉的威兰胶平均分子最低,分子量分布最宽,储能模量最低,损耗模量最高。
最后,考察NaCl添加对威兰胶海水溶液表观粘度的影响,并对其在NaCl质量分数10%时的耐高温性能进行研究。结果表明:低浓度NaCl有利于提高威兰胶海水溶液表观粘度,而高浓度NaCl有降低该体系粘度的性能。标准海水浓度下葡萄糖豆粕来源的威兰胶溶液的表观粘度较高,且在100℃下稳定,而在120℃下商品威兰胶溶液粘度保留能力较强,但粘度明显降低。
Welan gum, a kind of highly vicious extracellular polysaccharide produced by Sphingomonas sp. ZW-3, has found great potential application in oil recovery, concrete construction industry for its unique rheological properties.
The proposal of this thesis is to produce welan gum by optimizing various low-cost raw materials, and study the molecular weight characterization with rheological properties. At first, welan batch fermentation under different nutrition conditions with Sphingomonas sp. ZW-3 was explored, and different modes of fed-batch fermentation were tested. Then, the rheological properties of welan solutions were analyzed, and their average molecular weight and molecular weight distribution, pseudoplasticity and viscosity, storage modulus and loss modulus were compared. In addition, preliminary exploration on welan samples’temperature resistance and salt tolerance was carried out. Outlines of this thesis are as follows:
At first, a series of low-cost carbon sources such as sugar molasses, glycerol, fructose syrup and cassava starch were used for welan fermentation. The results indicated that the first three carbon sources could not achieve high welan yield, which needs further research. According to the principles of cassava starch hydrolysis with highly thermostableα-amylase and glucoamylase treatment, welan batch fermentation process with cassava starch hydrolysate of different DE value as carbon sources showed that welan yield from batch fermentation with DE=87% cassava starch hydrolysate reached 21.2 g/L, which was a little higher than that of batch fermentation with glucose as carbon sources.
Second, influences of different nitrogen sources (soybean flour, dried corn steep liquor powder and combined nitrogen sources (the mixture of NaNO3 and soybean peptone) and different carbon sources (glucose and sucrose) on welan batch fermentation were investigated. The results showed nitrogen sources had greater impact on welan yield in comparison with tested carbon sources. The highest welan yield of 20.0 g/L was obtained with soybean flour as nitrogen sources. The welan yield of 17.0-18.0 g/L was achieved with dried corn steep liquor powder as nitrogen source. The lowest welan yield of 15.0 g/L was from combined nitrogen sources. The results of fed-batch fermentation suggested that the starting glucose concentration 10.0 g/L was more favorable for welan production as compared to the initial 20.0 g/L glucose, and the final welan yield was 24.8 g/L.
Then, according to welan batch fermentation under different carbon and nitrogen sources, polymer rheological properties of the produced welan solutions were analyzed by using zero shear viscosity measurement, steady flow measurement and dynamic viscoelasticity measurement. The resultant data was applied to Carreau model, Power Law model and the linear simulation of complex viscosity log (η*) to log (ω). The analysis indicated that welan average molecular weight and molecular weight distribution were greatly affected by nitrogen sources in comparison with tested carbon sources. Welan samples derived from combined nitrogen media exhibited highest average molecular weight, narrowest molecular weight distribution and highest storage modulus. Welan sample from soybean flour nitrogen sources media was modest, while welan sample produced from dried corn steep liquor powder media exhibited lowest average molecular weight, widest molecular weight distribution, lowest storage modulus but highest loss modulus.
Finally, influences of NaCl on viscosity of welan standard sea-water solutions was tested, and their high temperature resistance at NaCl 10%(w/w)was also studied. The results suggested that low NaCl concentration was favorable for improving the viscosity of welan standard sea-water solutions, and high NaCl concentration would decrease the viscosity. The viscosity of welan standard sea-water solutions was higher, and stable at 100℃. But when the temperature was increased to 120℃, the viscosity retention ability of commercial welan sea-water solutions was stronger with viscosity also apparently reducing.
本文旨在以各种低成本原料进行威兰胶发酵研究,实现威兰胶的低成本生产,并探索与其分子量相关的流变学性质。首先,研究不同营养条件下Sphingomonas sp. ZW-3分批发酵及补料分批过程;其次,分析不同营养条件下威兰胶流变学性质,对其平均分子量及其分布、假塑性及粘性、储能模量及损耗模量进行比较,并对样品的抗盐耐温性进行初步探索。论文主要研究成果如下:
首先,在已优化培养条件的基础上利用低成本碳源如甘蔗糖蜜、甘油、果葡糖浆及木薯淀粉开展威兰胶发酵研究。结果表明:前三种碳源的威兰胶产量不够理想,仍需深入研究;而基于木薯淀粉的高温α-淀粉酶、葡萄糖淀粉酶水解规律的分析,不同DE值木薯淀粉水解液为碳源的分批发酵结果表明,DE=87%时威兰胶产量为21.2 g/L,稍高于葡萄糖为碳源时的威兰胶产量(20.2 g/L)。
其次,考察了不同氮源豆粕、玉米浆干粉及复合氮源(硝酸钠和大豆蛋白胨混合物)及碳源(葡萄糖、蔗糖)对威兰胶分批发酵影响。结果表明:与所考察碳源相比,氮源对威兰胶产量影响显著,豆粕为氮源时威兰胶产量最高(约20.0 g/L),玉米浆干粉为氮源时次之(17.0-18.0 g/L),复合氮源为氮源时威兰胶产量约15.0 g/L。补料分批发酵结果表明:初糖浓度10.0 g/L比20.0 g/L对威兰胶合成更有利,发酵72 h的威兰胶产量达到24.8 g/L。
再次,基于不同氮源及碳源的威兰胶分批发酵分析,对不同威兰胶样品的高分子流变学性质进行比较。对各种威兰胶样品溶液进行零剪切粘度测试、稳态流变学测试及动态粘弹性测试,并对所得测定数据进行Carreau模型、Power Law模型拟合及复数粘度(logη*)对(logω)的线性拟合。结果表明:与所考察碳源相比,氮源对威兰胶平均分子量及分子量分布影响显著;产自复合氮源的威兰胶平均分子量最高,分子量分布最窄,储能模量最高;产自氮源豆粕的威兰胶次之,而产自玉米浆干粉的威兰胶平均分子最低,分子量分布最宽,储能模量最低,损耗模量最高。
最后,考察NaCl添加对威兰胶海水溶液表观粘度的影响,并对其在NaCl质量分数10%时的耐高温性能进行研究。结果表明:低浓度NaCl有利于提高威兰胶海水溶液表观粘度,而高浓度NaCl有降低该体系粘度的性能。标准海水浓度下葡萄糖豆粕来源的威兰胶溶液的表观粘度较高,且在100℃下稳定,而在120℃下商品威兰胶溶液粘度保留能力较强,但粘度明显降低。
Welan gum, a kind of highly vicious extracellular polysaccharide produced by Sphingomonas sp. ZW-3, has found great potential application in oil recovery, concrete construction industry for its unique rheological properties.
The proposal of this thesis is to produce welan gum by optimizing various low-cost raw materials, and study the molecular weight characterization with rheological properties. At first, welan batch fermentation under different nutrition conditions with Sphingomonas sp. ZW-3 was explored, and different modes of fed-batch fermentation were tested. Then, the rheological properties of welan solutions were analyzed, and their average molecular weight and molecular weight distribution, pseudoplasticity and viscosity, storage modulus and loss modulus were compared. In addition, preliminary exploration on welan samples’temperature resistance and salt tolerance was carried out. Outlines of this thesis are as follows:
At first, a series of low-cost carbon sources such as sugar molasses, glycerol, fructose syrup and cassava starch were used for welan fermentation. The results indicated that the first three carbon sources could not achieve high welan yield, which needs further research. According to the principles of cassava starch hydrolysis with highly thermostableα-amylase and glucoamylase treatment, welan batch fermentation process with cassava starch hydrolysate of different DE value as carbon sources showed that welan yield from batch fermentation with DE=87% cassava starch hydrolysate reached 21.2 g/L, which was a little higher than that of batch fermentation with glucose as carbon sources.
Second, influences of different nitrogen sources (soybean flour, dried corn steep liquor powder and combined nitrogen sources (the mixture of NaNO3 and soybean peptone) and different carbon sources (glucose and sucrose) on welan batch fermentation were investigated. The results showed nitrogen sources had greater impact on welan yield in comparison with tested carbon sources. The highest welan yield of 20.0 g/L was obtained with soybean flour as nitrogen sources. The welan yield of 17.0-18.0 g/L was achieved with dried corn steep liquor powder as nitrogen source. The lowest welan yield of 15.0 g/L was from combined nitrogen sources. The results of fed-batch fermentation suggested that the starting glucose concentration 10.0 g/L was more favorable for welan production as compared to the initial 20.0 g/L glucose, and the final welan yield was 24.8 g/L.
Then, according to welan batch fermentation under different carbon and nitrogen sources, polymer rheological properties of the produced welan solutions were analyzed by using zero shear viscosity measurement, steady flow measurement and dynamic viscoelasticity measurement. The resultant data was applied to Carreau model, Power Law model and the linear simulation of complex viscosity log (η*) to log (ω). The analysis indicated that welan average molecular weight and molecular weight distribution were greatly affected by nitrogen sources in comparison with tested carbon sources. Welan samples derived from combined nitrogen media exhibited highest average molecular weight, narrowest molecular weight distribution and highest storage modulus. Welan sample from soybean flour nitrogen sources media was modest, while welan sample produced from dried corn steep liquor powder media exhibited lowest average molecular weight, widest molecular weight distribution, lowest storage modulus but highest loss modulus.
Finally, influences of NaCl on viscosity of welan standard sea-water solutions was tested, and their high temperature resistance at NaCl 10%(w/w)was also studied. The results suggested that low NaCl concentration was favorable for improving the viscosity of welan standard sea-water solutions, and high NaCl concentration would decrease the viscosity. The viscosity of welan standard sea-water solutions was higher, and stable at 100℃. But when the temperature was increased to 120℃, the viscosity retention ability of commercial welan sea-water solutions was stronger with viscosity also apparently reducing.
引文
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