工程菌毕赤酵母发酵液制备β-葡寡糖的结构表征及其透皮性
Enzymatic Preparation of β-Glucan Oligosaccharides by Pichia Pastoris and Its Etransdermal Properties
王珂鑫 1Inki Park 2许海燕 2管世敏 1费一弘 3黄煜玲 3荣绍丰1
作者信息
- 1. 上海应用技术大学香料香精化妆品学部,上海 201418
- 2. 科玛化妆品(无锡)有限公司,江苏无锡 214111
- 3. 上海侬本雅生物科技有限公司,上海 201418
- 折叠
摘要
为实现β-葡聚糖的精准小分子化,试验构建了可异源表达β-1,3-葡聚糖内切酶的基因重组毕赤酵母菌(Pichia pastoris MF060.RG.Glu-13),经发酵培养获得的发酵液中β-1,3-葡聚糖内切酶酶活达到 982 U/mL.利用发酵液酶解β-葡聚糖制备分子质量在1～2 kDa的β-葡寡糖.采用红外光谱及核磁共振波谱对β-葡寡糖进行初步结构鉴定,采用Franz扩散池对β-葡寡糖进行透皮性能测试.结果表明,产物β-葡寡糖主链由 5 个葡萄糖糖苷通过β-1,3-糖苷键连接,在第二、第三个葡萄糖糖苷上,各有1个由β-1,6-糖苷键连接的D-葡萄糖分支.酶解产物β-葡寡糖(1～2 kDa)和酶解底物大分子β-葡聚糖(800～1 000 kDa)累计透皮4 h后透皮吸收率分别为2.13%和0.04%,经酶解,小分子产物的透皮率提升至大分子底物的53.25倍.
Abstract
In order to accurately convert β-glucan into small molecules,recombinant Pichia pastoris MF060.RG.Glu-13 was constructed to express β-1,3-glucan endonucliase.The activity of β-1,3-glucan endonucliase reached 982 U/mL in the fermentation broth obtained by fermentation.The β-glucan oligosaccharides with molecular weight of 1-2 kDa were prepared by enzymatic hydrolysis of β-glucan in fermentation broth.IR and NMR were used to preliminarily identify the structure of β-glucoseoligosaccharides,and Franz diffusion cell was used to test the ir transdermal property.The results showed that the main chain of the product β-glucooligosaccharide was connected by five glucosides by β-1,3-glycosidic bonds,and there was a D-glucose branch connected by β-1,6-glycosidic bonds on the second and third glucosides.The transdermal absorption rate of the product β-glucose-oligosaccharide(1-2 kDa)was 2.13%,which was 53.25 times higher than that of the enzymatic substrate,macromolecule β-glucan(800-1 000 kDa),of 0.04%.
关键词
β-葡聚糖/β-葡寡糖/毕赤酵母Key words
β-glucan/β-glucan oligosaccharides/Pichia pastoris引用本文复制引用
出版年
2024
NETL
NSTL
万方数据