酶解辅助提取对铁皮石斛多糖结构和菌群调节功能的影响

doi:10.3969/j.issn.1004-1524.20231114

浙江农业学报 ›› 2024, Vol. 36 ›› Issue (9): 2099-2109.DOI: 10.3969/j.issn.1004-1524.20231114

酶解辅助提取对铁皮石斛多糖结构和菌群调节功能的影响

张妮¹(), 陶文扬², 罗梦帆², 周万怡², 郑晓杰³, 李彦坡³, 金火喜¹^,^*(), 杨颖²^,^*()

1.浙江海洋大学食品与药学学院,浙江舟山 316002
2.浙江省农业科学院食品科学研究所,浙江杭州 310021
3.温州科技职业学院(温州市农业科学研究院),浙江温州 325000

收稿日期:2023-09-21 出版日期:2024-09-25 发布日期:2024-09-30
作者简介:杨颖,E-mail:ying_yang4535F@sina.com
*金火喜,E-mail: jinhuoxi@163.com;
张妮(1997—),女,山东潍坊人,硕士研究生,主要从事铁皮石斛多糖功效研究。E-mail:17863129235@163.com
通讯作者: *金火喜,E-mail: jinhuoxi@163.com; 杨颖,E-mail:ying_yang4535F@sina.com
基金资助:
浙江省市级农科院联盟区域示范性项目(2022SJLM06)

Effects of enzyme-assisted extraction on composition and gut microbiota regulation function of Dendrobium officinale polysaccharide

ZHANG Ni¹(), TAO Wenyang², LUO Mengfan², ZHOU Wanyi², ZHENG Xiaojie³, LI Yanpo³, JIN Huoxi¹^,^*(), YANG Ying²^,^*()

1. College of Food and Medicine, Zhejiang Ocean University, Zhoushan 316002, Zhejiang, China
2. Institute of Food Science, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
3. Wenzhou Vocational College of Science and Technology (Wenzhou Academy of Agricultural Sciences), Wenzhou 325000, Zhejiang, China

Received:2023-09-21 Online:2024-09-25 Published:2024-09-30

摘要/Abstract

摘要：

酶解或许会改变多糖分子内某些基团的数量或结构,从而改变其性能。为明确酶解对铁皮石斛多糖结构与功能的影响,本研究选择同一品种石斛鲜茎为原料,分别用中性蛋白酶、麦芽糖淀粉酶、纤维素酶辅助提取,获得同一基质多糖的不同结构形式,分析其结构区别,并利用体外模拟发酵系统研究铁皮石斛多糖菌群调节功能的变化。结果表明,与未酶解样品相比,中性蛋白酶辅助提取可以增加多糖的聚合度,而纤维素酶、麦芽糖淀粉酶介入则可以显著降低多糖聚合度并改变单糖组成;中性蛋白酶组的CH₄与H₂S生成量显著增加,纤维素酶组的丁酸与戊酸产量显著增加。多糖能改善肠道菌群的组成,其中纤维素酶组改变程度较大,双歧杆菌属、乳杆菌属、粪杆菌属、罗氏菌属等肠道基石菌与有益菌的种类与相对丰度显著增加,戴阿利斯特杆菌属、多尔氏菌属等条件致病菌的丰度显著下降,与丁酸、戊酸显著正相关的9个菌属在该组富集了8个(巨球型菌属、链球菌属、双歧杆菌属、欧陆森氏菌属、罗氏菌属、魏斯氏菌属、乳杆菌属、普拉梭菌属)。酶辅助提取可以改变石斛多糖的结构,并改变其菌群调节功能,该研究为不同功效多糖的定向生产提供了参考。

关键词: 铁皮石斛, 多糖, 酶解, 肠道菌群, 体外发酵

Abstract:

Enzymatic hydrolysis may alter the number or structure of certain groups within the polysaccharide molecule, thus altering its properties. To determine the effect of enzymatic hydrolysis on the structure and function of Dendrobium officinale polysaccharide, this study selected the same species of D. officinale fresh stems as raw materials, and obtained different structures of the same matrix polysaccharide through neutral protease, maltose amylase, and cellulase-assisted extraction. Then, analyzing structural differences between them, and studying their various regulatory functions to intestinal flora using an in vitro simulated fermentation system. The results showed that compared with unenzymatically hydrolyzed samples, neutral protease-assisted extraction increased the degree of polymerization of polysaccharides, while the intervention of cellulase and maltose amylase significantly reduced the degree of polysaccharide polymerization and changed their monosaccharide composition. The production of CH₄ and H₂S in the neutral protease-treated group increased significantly, while the production of butyric acid and valeric acid in the cellulase-treated group increased significantly. Polysaccharides improved the composition of intestinal flora, especially the cellulase-treated group. The types and relative abundance of intestinal cornerstone bacteria as well as beneficial bacteria, such as Bifidobacterium, Lactobacillus, Faecalibacterium and Roseburia, increased significantly, while the relative abundance of opportunistic pathogenic bacteria, such as Dialister and Dorea, decreased significantly. There were nine bacterial genera that were significantly positively correlated with butyric acid and valeric acid levels, and eight of them were enriched in the cellulase-treated group. Enzymatic hydrolysis-assisted extraction changes the structure of D. officinale polysaccharides, thereby changing its regulatory functions to intestinal flora. This study provides a reference for the targeted production of polysaccharides with different functions.

Key words: Dendrobium officinale, polysaccharides, enzymatic hydrolysis, intestinal flora, in vitro fermentation

中图分类号:

R285

张妮, 陶文扬, 罗梦帆, 周万怡, 郑晓杰, 李彦坡, 金火喜, 杨颖. 酶解辅助提取对铁皮石斛多糖结构和菌群调节功能的影响[J]. 浙江农业学报, 2024, 36(9): 2099-2109.

ZHANG Ni, TAO Wenyang, LUO Mengfan, ZHOU Wanyi, ZHENG Xiaojie, LI Yanpo, JIN Huoxi, YANG Ying. Effects of enzyme-assisted extraction on composition and gut microbiota regulation function of Dendrobium officinale polysaccharide[J]. Acta Agriculturae Zhejiangensis, 2024, 36(9): 2099-2109.

图/表 8

表1 提取方式对石斛多糖峰值分子量的影响

Table 1 Effect of extraction method on peak molecular weight of Dendrobium officinale polysaccharides u

提取方式 Extraction mode	峰值分子量Peak molecular weight
提取方式 Extraction mode	组分1 Component 1	组分2 Component 2
未酶解No enzymolysis	568 547	2 028
中性蛋白酶Neutral protease	892 441	2 228
麦芽糖淀粉酶Maltose amylase	248 758	1 745
纤维素酶Cellulase	22 044	1 356

表2 提取方式对铁皮石斛单糖组成的影响

Table 2 Effect of extraction method on monosaccharide composition from Dendrobium officinale

提取方式 Extraction mode	摩尔比Molar ratio
提取方式 Extraction mode	阿拉伯糖Arabinose	半乳糖Galactose	葡萄糖Glucose	甘露糖Mannose
未酶解No enzymolysis	0.003	0.005	1	1.83
中性蛋白酶Neutral protease	0.001	0.002	1	1.65
麦芽糖淀粉酶Maltose amylase	0	0	1	1.67
纤维素酶Cellulase	0.001	0.002	1	0.84

图1 铁皮石斛多糖对肠道微生物发酵气体的影响 Bc,空白对照组;Nc,未酶解对照组;Mal,麦芽糖淀粉酶组;Cel,纤维素酶组;Neu,中性蛋白酶组。散点图上无相同小写字母的表示各处理间差异显著(P<0.05)。下同。

Fig.1 Effect of Dendrobium officinale polysaccharides on intestinal microbial fermentation gas Bc, Blank control; Nc, No enzymolysis control; Mal, Maltose amylase group; Cel, Cellulase group; Neu, Neutral protease group. A scatterplot without the same lowercase letter indicates significant differences between treatments (P<0.05). The same as below.

图2 铁皮石斛多糖对短链脂肪酸含量的影响

Fig.2 Effect of Dendrobium officinale polysaccharides on the content of short-chain fatty acids

图3 铁皮石斛多糖对肠道菌群α多样性的影响

Fig.3 Effect of polysaccharides from Dendrobium officinale on α diversity of intestinal flora

图4 铁皮石斛多糖对肠道菌群β多样性的影响 A,主坐标(PCoA)分析;B,非度量多维尺度(NMDS)分析。

Fig.4 Effect of polysaccharides from Dendrobium officinale on β diversity of intestinal flora A, Principal co-ordinates analysis; B, Non-metric multi-dimensional scaling.

图5 铁皮石斛多糖对肠道菌群群落组成的影响 A,门水平组成差异;B,属水平热图。

Fig.5 Effect of polysaccharides from Dendrobium officinale on intestinal flora community composition A, Composition difference at phylum level; B, Multi-group comparison difference at genus level.

图6 铁皮石斛多糖对肠道菌群物种组成差异与相关性的影响 A,空白对照、未酶解对照与麦芽糖淀粉酶组间显著差异LDA值分布柱状图;B,空白对照、未酶解对照与纤维素酶组间显著差异LDA值分布柱状图;C,空白对照、未酶解对照与中性蛋白酶组间显著差异LDA值分布柱状图;D,属水平肠道菌群间相关性分析图,*表示0.01<P≤0.05,**表示0.001<P≤0.01,***表示P≤0.001。

Fig.6 Effect of polysaccharides from Dendrobium officinale on on species composition difference and correlation of intestinal flora A, Histogram of distribution of significantly different LDA values among blank control, no enzymolysis control and maltose amylase groups; B, Histogram of distribution of significantly different LDA values among blank control, no enzymolysis control and cellulase groups; C, Histogram of distribution of significantly different LDA values among blank control, no enzymolysis control and neutral protease groups; D, Correlation analysis diagram of intestinal flora at genus level, * represented 0.01<P≤0.05, ** represented 0.001<P≤0.01, *** represented P≤0.001.

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酶解辅助提取对铁皮石斛多糖结构和菌群调节功能的影响

Effects of enzyme-assisted extraction on composition and gut microbiota regulation function of Dendrobium officinale polysaccharide

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