少动鞘氨醇单胞菌产高水溶性结冷胶的分子特征与制备平台

Molecular characteristics and preparation platform for highly water-soluble gellan produced by Sphingomonas paucimobilis

探讨少动鞘氨醇单胞菌(Sphingomonas paucimobilis)ATCC 31461 GL-1变异株发酵生产胞外多糖(exopolysaccharides,EPS)(结冷胶)的单糖组成、分子参数及热水溶解速率受豆粕粉氮源与沉淀方式的影响,确定制备高水溶性结冷胶的流程。该菌以豆粕粉(和蔗糖)为培养基,发酵所得胞外多糖(EPS-Sw)产量为29.1 g/L、可溶性总糖含量为80%,乙酰基含量为4.0%,皆显著高于以酵母膏和蛋白胨发酵并醇沉收集的样品(分別为EPS-Ye和EPS-Pe)。EPS-Sw、EPS-Ye、EPS-Pe与结冷胶商品Gellan-F(参考组)共同含有4种主要单糖:鼠李糖(rhamnose,Rha)、葡萄糖(glucose,Glc)、葡萄糖醛酸(glucuronic acid,GlcA)及半乳糖醛酸(galacturonic acid,GalA)。EPS-Sw的糖摩尔比为M(Rha)∶M(Glc)∶M(GlcA)∶M(GalA)=1∶0.9∶2.3∶1.7,与EPS-Ye和EPS-Pe的糖比例相近。EPS-Sw的结冷胶相对纯度(共同糖组成总和)为62.8%。经微过滤后,上述醇沉的EPS样品和Gellan-F皆呈三分子尖峰分布。针对豆粕粉发酵得的上清液,添加5 mmol/L Ca^(2+)复合乙醇的沉淀处理可提高制备物(EPS-CaEt)的产量、可溶性总糖含量、乙酰基含量、结冷胶相对纯度、热水既溶性以及溶解速率常数(k_(s1))。超大分子组分(F1)为重要的分子因素,F1含量与结冷胶相关的糖组成含量呈高度正相关,与热水既溶性和速率常数k_(s1)呈高度负相关。该研究阐明了少动鞘氨醇单胞菌以豆粕粉氮源发酵所产结冷胶的关键糖组成与分子参数,并建立了一个有效富集具有高结冷胶纯度与高水溶性的EPS的Ca^(2+)-乙醇复合沉淀的制备平台。

This study aimed to investigate the effects of soybean cake powder as nitrogen source and precipitation method on the monosaccharide composition,molecular parameter,and hot-water solubilization rate of the exopolysaccharide(EPS),gellan,from the mutant strain GL-1 of Sphingomonas paucimobilis ATCC 31461,so as to determine a proper precipitation treatment for preparing highly water-soluble gellan.S.paucimobilis was cultivated with the medium containing soybean cake powder(and sucrose),the EPS(EPS-Sw)yield was 29.1 g/L,with the total soluble carbohydrate content of 80%and acyl content of 4.0%,significantly higher than those cultivated with yeast extract and peptone,and collected by ethanol precipitation(EPS-Ye and EPS-Pe,respectively).EPS-Sw,EPS-Ye,EPS-Pe,and commercial gellan Gellan-F(reference)were commonly composed of four major monosaccharides:rhamnose(Rha),glucose(Glc),glucuronic acid(GlcA),and galacturonic acid(GalA).The molar ratio Rha∶Glc∶GlcA∶GalA of EPS-Sw was 1∶0.9∶2.3∶1.7,close to those of EPS-Ye and EPS-Pe.The gellan relative purity(total amount of the common sugars)was 62.8%for EPS-Sw.After microfiltration,above-mentioned ethanol-precipitated EPS and Gellan-F exhibited triple molecular distributions.For the supernatant of mash fermented by soybean cake powder,adding 5 mmol/L Ca^(2+)coupled with ethanol precipitation led to the EPS preparation(EPS-CaEt)of the increments in yield,total soluble carbohydrate content,acyl content,gellan relative purity,hot-water instant solubility,and solubilizing rate constant(k_(s1)).Super large molecular fraction(F1)was a crucial factor and highly correlated positively with gellan-related sugar compositions and negatively with hot-water instant solubility and k_(s1).Conclusively,this study elucidates the key monosaccharide composition and molecular parameters of S.paucimobilis EPS produced by soybean cake powder as nitrogen source.A cost-effective preparation method for the EPS of improved gellan relative purity and high water-solubility was established.The results provide a theoretical basis for gellan-producing industry.