米糠不溶性膳食纤维结合酚结构特性及其对肠道菌群的影响

施宇萌; 梁富强; 郭锐林; 段姗姗; 濮欣然; 石嘉怿

doi:10.13386/j.issn1002-0306.2022120059

米糠不溶性膳食纤维结合酚结构特性及其对肠道菌群的影响

doi: 10.13386/j.issn1002-0306.2022120059

南京财经大学食品科学与工程学院，江苏南京 210023

基金项目: 国家自然科学基金（32001702）

详细信息

作者简介:
施宇萌（1998−），女，硕士，研究方向：天然产物与功能性食品，E-mail：shiyumengi@163.com

通讯作者:
梁富强（1991−），男，博士，讲师，研究方向：天然产物与功能性食品，E-mail：fuqiangliang@nufe.edu.cn

中图分类号: TS201.1
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- 被引次数: 0
出版历程
- 收稿日期: 2022-12-07
- 网络出版日期: 2023-05-19
- 刊出日期: 2023-05-15

Structural Characteristics of Rice Bran Insoluble Dietary Fiber Bound Phenolic and Its Effect on Gut Microbiota

College of Food Science and Engineering, Nanjing University of Finance and Economics, Nanjing 210023, China

摘要

摘要: 米糠是谷物酚类物质的重要来源，其中多酚主要与膳食纤维结合，以结合态形式存在。本研究通过顺序酶法制备米糠不溶性膳食纤维结合酚（Insoluble Dietary Fiber-Bound Phenolic，IDF-BP），采用激光共聚焦显微镜、红外光谱、扫描电镜和X-射线衍射来表征IDF-BP结构变化，并对IDF-BP进行体外模拟胃肠消化和结肠发酵实验，随后对发酵物进行16S rRNA高通量测序的短链脂肪酸测定。结果表明，从米糠中制备得米糠IDF-BP得率为23.73%±0.008%，结合酚（Bound Phenolic，BP）含量为6.11±0.085 mg GAE/g DW，其中以阿魏酸含量最高为1.41±0.013 mg/g DW，其是典型的纤维素晶型，具有纤维素和半纤维素特征基团、酯键特征峰，以酯键形式与膳食纤维结合。肠道菌群多样性和菌群丰度结果显示，IDF-BP可通过降低肠道菌群多样性，下调肠道有害菌如拟杆菌属、考拉菌属，上调肠道有益菌如双歧杆菌属和艾克曼菌属，从而达到调节肠道菌群平衡的目的。同时也能促进短链脂肪酸（主要是乙酸、丙酸和丁酸）的产生以调节肠道pH。米糠IDF-BP兼具膳食纤维和多酚的特性，二者对肠道微生物和短链脂肪酸有较好的协同作用，有助于改善机体肠道环境，促进机体健康。
- 米糠 /
- 不溶性膳食纤维结合酚 /
- 结构性质 /
- 结肠发酵 /
- 肠道菌群
Abstract: Rice bran is a major source of phenolic substances in cereals, in which phenolics mainly bound with dietary fiber in the form of conjugated state. In this study, rice bran insoluble dietary fiber-bound phenolic (IDF-BP) was prepared by sequential enzymatic method. The structure of IDF-BP was characterized by confocal laser scanning microscope (CLSM), Fourier transform infrared spectrometer (FT-IR), scanning electron microscope (SEM) and X-ray diffraction. The IDF-BP was subjected to simulated gastrointestinal digestion and colon fermentation in vitro, followed by 16S rRNA high-throughput sequencing of short-chain fatty acids. The results showed that the yield of rice bran IDF-BP was 23.73%±0.008%, the content of BP was 6.11±0.085 mg GAE/g DW, and the content of ferulic acid was the highest (1.41±0.013 mg/g DW). The IDF-BP was a typical cellulose crystal form of cellulose, which had both cellulose and hemicellulose characteristic groups and ester bond characteristic peaks, indicating that bound phenolic were bound to dietary fiber in the form of ester bond. The results of intestinal flora diversity and flora abundance showed that IDF-BP could regulate the balance of intestinal flora by reducing the diversity of intestinal flora, down-regulating intestinal harmful bacteria such as Bacteroides and Phascolarctobacterium, and increase intestinal beneficial bacteria such as Bifidobacterium and Akkermansia to achieve the goal of regulating the balance of gut microbiota. Additionally, it also had the ability to boost the production of short chain fatty acids (mainly acetic acid, propionic acid and butyric acid) to adjust intestinal pH. Rice bran IDF-BP had both the characteristics of dietary fiber and phenolics, and the two had a good synergistic effect on intestinal flora and short chain fatty acids, which would indicate that rice bran IDF-BP has the potential to improve the intestinal environment, thus achieving health benefits.
- rice bran /
- insoluble dietary fiber bound phenolic /
- structural properties /
- colonic fermentation /
- gut microbiota

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图 1 米糠结合酚和酚酸标准品液相图

注：a：米糠结合酚液相图，图中1为丁香酸，2为对香豆酸，4为阿魏酸；b：阿魏酸标准品液相图；c：丁香酸标准品液相图；d：对香豆酸液相图。

Figure 1. HPLC-DAD diagram of rice bran bound phenol and phenolic acid standard

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图 2 IDF-BP（a）和JIDF（b）的CLSM观察图

Figure 2. CLSM observations of IDF-BP (a) and JIDF (b)

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图 3 IDF-BP和JIDF的FT-IR图

Figure 3. FT-IR spectra of IDF-BP and JIDF

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图 4 IDF-BP（a）和JIDF（b）扫描电镜图

Figure 4. Scanning electron microscopy images of IDF-BP (a) and JIDF (b)

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图 5 IDF-BP和JIDF的X-射线衍射图

Figure 5. X-ray diffraction patterns of IDF-BP and JDF

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图 6 肠道微生物多样性

注：a：韦恩图；b：Chao指数图；c：ACE指数图；d：Shannon指数图；e：Simpson指数图；Blank组为空白组，JIDF组为单独膳食纤维组，FA组为单独阿魏酸组，IDF-BP为米糠不溶性膳食纤维结合酚组；图中不同字母表示有显著性差异，P<0.05。

Figure 6. Intestinal microbial diversity

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图 7 肠道菌群相对丰度图和热图

注：a：门水平肠道菌群相对丰度图；b：属水平肠道菌群相对丰度图；c：属水平肠道菌群热图。

Figure 7. Figure of relative abundance of intestinal flora and heatmap

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图 8 发酵过程中SCFAs含量变化

注：a：乙酸含量图；b：丙酸含量图；c：丁酸含量图；不同小写字母表示组内差异显著（P<0.05），不同大写字母表示组间差异显著（P<0.05）。

Figure 8. Changes of SCFAs content during fermentation

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表 1 模拟胃肠消化液配制

Table 1. Preparation of stock solutions of simulated digestion fluids

试剂	SSF（pH7）	SGF（pH3）	SIF（pH7）
KCl（mmol/L）	15.1	6.9	6.8
KH₂PO₄（mmol/L）	3.7	0.9	0.8
NaHCO₃（mmol/L）	13.6	25	85
NaCl（mmol/L）	−	47.2	38.4
MgCl₂（H₂O）（mmol/L）	0.15	0.1	0.33
（NH₄）₂CO₃（mmol/L）	0.06	0.5	−

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表 2 酚酸标准曲线

Table 2. Standard curves of phenolic acid

标准品名称	出峰时间（min）	标准曲线方程
丁香酸	18.509	y=64462x+46.473，R²=0.999
对香豆酸	24.051	y=67089x−45.209，R²=0.999
阿魏酸	27.437	y=16719x−120.76，R²=0.999

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表 3 结肠发酵过程中pH变化

Table 3. pH value during the fermentation

样品	0 h	6 h	12 h	24 h	30 h	48 h
Blank	6.85±0.035^Ba	5.13±0.005^Ae	5.47±0.007^Ad	5.79±0.042^Ac	5.86±0.014^Ab	5.92±0.035^Ab
IDF-BP	6.90±0.042^Ba	4.70±0.071^Be	4.82±0.042^Bd	4.86±0.035^Cd	5.08±0.049^Cc	5.47±0.014^Cb
JIDF	7.02±0.042^Aa	4.70±0.021^Bc	4.70±0.021^Cc	4.75±0.035^Dc	4.76±0.035^Dc	4.85±0.028^Db
FA	6.45±0.014^Ca	4.76±0.071^Be	4.76±0.071^BCe	5.34±0.0141^Bd	5.56±0.021^Bc	5.79±0.028^Bb
注：不同小写字母表示组内差异显著（P<0.05），不同大写字母表示组间差异显著（P<0.05）。

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参考文献(38)

[1]	陈甜妹. 米糠固态发酵工艺优化及其对米糠营养功能特性的影响[D]. 佛山: 佛山科学技术学院, 2020 CHEN T M. Technology optimization for rice bran solid state fermentation and its effects on the nutritional and functional properties of rice bran[D]. Foshan: Foshan University, 2020.
[2]	BUTSAT S, SIRIAMORNPUN S. Antioxidant capacities and phenolic compounds of the husk, bran and endosperm of Thai rice[J]. Food Chemistry,2010,119(2):606−613. doi: 10.1016/j.foodchem.2009.07.001
[3]	DE MIRA N V M, MASSARETTO I L, PASCUAL C S C I, et al. Comparative study of phenolic compounds in different Brazilian rice (Oryza sativa L.) genotypes[J]. Journal of Food Composition and Analysis,2009,22(5):405−409. doi: 10.1016/j.jfca.2008.06.012
[4]	ACOSTA-ESTRADA B A, GUTIÉRREZ-URIBE J A, SERNA-SALDÍVAR S O. Bound phenolics in foods, a review[J]. Food Chemistry,2014,152:46−55. doi: 10.1016/j.foodchem.2013.11.093
[5]	ZHANG X, ZHANG M, DONG L, et al. Phytochemical profile, bioactivity, and prebiotic potential of bound phenolics released from rice bran dietary fiber during in vitro gastrointestinal digestion and colonic fermentation[J]. Journal of Agricultural and Food Chemistry,2019,67(46):12796−12805. doi: 10.1021/acs.jafc.9b06477
[6]	PALAFOX-CARLOS H, AYALA-ZAVALA J F, GONZÁLEZ-AGUILAR G A. The role of dietary fiber in the bioaccessibility and bioavailability of fruit and vegetable antioxidants[J]. Journal of Food Science,2011,76(1):R6−R15. doi: 10.1111/j.1750-3841.2010.01957.x
[7]	武锦春. 正常人肠道菌群对酸枣仁总黄酮的体外代谢研究[D]. 太原: 山西大学, 2021 WU J C. Study on the in vitro metabolism of total flavonoids in Ziziphi Spinosae Semen by normal human intestinal flora[D]. Taiyuan: Shanxi University, 2021.
[8]	DONG R, YU Q, LIAO W, et al. Composition of bound polyphenols from carrot dietary fiber and its in vivo and in vitro antioxidant activity[J]. Food Chemistry,2021,339:127879. doi: 10.1016/j.foodchem.2020.127879
[9]	李文婷. 六种豆类中不同结合态多酚组成、胃肠道生物可接受率及其抗氧化活性研究[D]. 南昌: 南昌大学, 2018 LI W T. The composition of free, conjugated and bound phenolics in six legumes, and their bioacessibility of gastrointestinal tract and antioxidant activity[D]. Nanchang: Nanchang University, 2018.
[10]	徐灼辉. 荔枝干果肉膳食纤维—结合酚加合物结构表征及其润肠通便作用机制[D]. 广州: 广州大学, 2020 XU Z H. Structural characterization of dietary fiber-bound phenolic adducts in dried lychee pulp and its mechanism of moistening intestine and laxative effect[D]. Guangzhou: Guangzhou University, 2020.
[11]	LI Y, LI M, WANG L, et al. Effect of particle size on the release behavior and functional properties of wheat bran phenolic compounds during in vitro gastrointestinal digestion[J]. Food Chemistry,2022,367:130751. doi: 10.1016/j.foodchem.2021.130751
[12]	任明非. 米糠不溶性膳食纤维与Cd²⁺结合特性的研究[D]. 无锡: 江南大学, 2020 REN M F. Study on the binding characteristics of rice bran insoluble dietary fiber with Cd²⁺[D]. Wuxi: Jiangnan University, 2020.
[13]	王洪欣. 铜藻膳食纤维的提取、性质及可食用膜的制备[D]. 杭州: 浙江工商大学, 2020 WANG H X. Extraction, properties and preparation of edible film of dietary fiber from Sargassum horneri[D]. Hangzhou: Zhejiang Gongshang University, 2020.
[14]	MINEKUS M, ALMINGER M, ALVITO P, et al. A standardised static in vitro digestion method suitable for food-an international consensus[J]. Food & Function,2014,5(6):1113−1124.
[15]	顿倩. 黑豆膳食纤维结合的多酚构成和微生物代谢规律的研究[D]. 南昌: 南昌大学, 2019 DUN Q. Study on composition and microbial metabolism of bound phenolics binding to dietary fiber of black bean[D]. Nanchang: Nanchang University, 2019.
[16]	程玉鑫. 干酪乳杆菌发酵蓝莓果渣对高脂膳食小鼠肠道屏障功能的调节作用及机制[D]. 武汉: 华中农业大学, 2020 CHENG Y X. Lactobacillus casei fermented blueberry pomace regulates the intestinal barrier function of high fat diet mice and the underlying mechanisms[D]. Wuhan: Huazhong Agricultural University, 2020.
[17]	赵珊, 仲伶俐, 周虹, 等. 超高效液相色谱-串联质谱法鉴定和分析稻米中酚酸类化合物的组成及分布[J]. 中国农业科学,2020,53(3):612−631. [ZHAO S, ZHONG L L, ZHOU H, et al. Identification and analysis of phenolic acids in rice using ultra-high performance liquid chromatography-tandem mass spectrometry[J]. Scientia Agricultura Sinica,2020,53(3):612−631.
[18]	XU Z, XIONG X, ZENG Q, et al. Alterations in structural and functional properties of insoluble dietary fibers-bound phenolic complexes derived from lychee pulp by alkaline hydrolysis treatment[J]. LWT,2020,127:109335. doi: 10.1016/j.lwt.2020.109335
[19]	刘帅. 基于微生物发酵研究胡萝卜膳食纤维与结合多酚之间酵解特性的相互影响[D]. 南昌: 南昌大学, 2020 LIU S. Study on the interaction between carrot dietary fibers and bound polyphenols based on microbial fermentation[D]. Nanchang: Nanchang University, 2020.
[20]	黄冬云. 米糠膳食纤维的酶法改性及功能性质研究[D]. 无锡: 江南大学, 2014 HUANG D Y. Enzymatic modification and functional properties of rice bran dietary fiber[D]. Wuxi: Jiangnan University, 2014.
[21]	郭天时. 米糖膳食纤维的微粉—酶法改性及其理化功能性质的变化研究[D]. 哈尔滨: 哈尔滨商业大学, 2017 GUO T S. Micronized-enzymatic modification of rice bran dietary fiber and changes of its physicochemical and functional properties[D]. Harbin: Harbin University of Commerce, 2017.
[22]	祁静. 高吸附性米糠纤维的制备及其吸附特性的研究[D]. 无锡: 江南大学, 2016 QI J. The research on the preparation and adsorption properties of rice bran fiber with high adsorption capacities[D]. Wuxi: Jiangnan University, 2016.
[23]	曹龙奎, 康丽君, 寇芳, 等. 改性前后小米糠膳食纤维结构分析及体外抑制α-葡萄糖苷酶活性[J]. 食品科学,2018,39(11):46−52. [CAO L K, KANG L J, KOU F, et al. Structural analysis and in vitro inhibitory effect on α-glucosidase activity of millet bran dietary fiber before and after modification[J]. Food Science,2018,39(11):46−52. doi: 10.7506/spkx1002-6630-201811008
[24]	HUANG H, CHEN J, HU X, et al. Elucidation of the interaction effect between dietary fiber and bound polyphenol components on the anti-hyperglycemic activity of tea residue dietary fiber[J]. Food & Function,2022,13(5):2710−2728.
[25]	刘倩, 范誉川, 刘素诗, 等. 米糠粕不溶性膳食纤维理化性质及对高脂大鼠肠道菌群的影响[J]. 食品科学,2021,42(15):174−179. [LIU Q, FAN Y C, LIU S S, et al. Physicochemical properties of insoluble dietary fiber from defatted rice bran and its effect on intestinal microbiota of high-fat diet fed rats[J]. Food Science,2021,42(15):174−179. doi: 10.7506/spkx1002-6630-20200730-395
[26]	LIU S, YU Q, HUANG H, et al. The effect of bound polyphenols on the fermentation and antioxidant properties of carrot dietary fiber in vivo and in vitro[J]. Food & Function,2020,11(1):748−758.
[27]	董瑞红. 胡萝卜膳食纤维中结合多酚的体外消化酵解规律和营养功能特性及其在酸奶中的应用[D]. 南昌: 南昌大学, 2021 DONG R H. The release and nutritional functional properties of bound polyphenols from carrot dietary fiber during in vitro digestion and colonic fermentation and its application in yogurt[D]. Nanchang: Nanchang University, 2021.
[28]	RASTMANESH R. High polyphenol, low probiotic diet for weight loss because of intestinal microbiota interaction[J]. Chemico-Biological Interactions,2011,189(1−2):1−8. doi: 10.1016/j.cbi.2010.10.002
[29]	AURA A M. Microbial metabolism of dietary phenolic compounds in the colon[J]. Phytochemistry Reviews,2008,7(3):407−429. doi: 10.1007/s11101-008-9095-3
[30]	韩飞, 韩阳阳, 赵建新, 等. 4种杂粮对大鼠肠道菌群组成及短链脂肪酸的影响[J]. 食品科学,2022,43(13):101−108. [HAN F, HANY Y, ZHAO J X, et al. Effects of four cereals on composition of gut microbiota and short-chain fatty acids in rats[J]. Food Science,2022,43(13):101−108. doi: 10.7506/spkx1002-6630-20210430-422
[31]	张文易. 绣球菌多糖结构表征及其对阿尔茨海默症小鼠的干预作用[D]. 无锡: 江南大学, 2022 ZHANG W Y. Structural characterization and the intervention of Sparassis crispa polysaccharides on the Alzheimer's disease mice[D]. Wuxi: Jiangnan University, 2022.
[32]	DONG R, LIU S, XIE J, et al. The recovery, catabolism and potential bioactivity of polyphenols from carrot subjected to in vitro simulated digestion and colonic fermentation[J]. Food Research International,2021,143:110263. doi: 10.1016/j.foodres.2021.110263
[33]	谢甜, 孙琳, 范伟, 等. 番石榴结合态多酚对小鼠肠道菌群结构及多样性的影响研究[J]. 核农学报,2021,35(11):2589−2597. [XIE T, SUN L, FAN W, et al. Effects of bound polyphenols in psidium guajava on mice intestinal flora structure and diversity[J]. Journal of Nuclear Agriclutural Sciences,2021,35(11):2589−2597. doi: 10.11869/j.issn.100-8551.2021.11.2589
[34]	单琴. 油茶籽粕多糖的降血脂功效及其对肠道菌群的影响[D]. 合肥: 合肥工业大学, 2021 SHAN Q. The hypolipidemic effect of Camellia seed meal polysaccharide and its influence on intestinal flora[D]. Hefei: Hefei University of Technology, 2021.
[35]	罗珍. 核桃壳多糖的分离鉴定及消化代谢特性研究[D]. 秦皇岛: 燕山大学, 2021 LUO Z. Isolation and identification of poly saccharide from juglans regial and its digestion and metabolism characteristics[D]. Qinhuangdao: Yanshan University, 2021.
[36]	ZHENG S, ZHANG H, LIU R, et al. Do short chain fatty acids and phenolic metabolites of the gut have synergistic anti-inflammatory effects?–New insights from a TNF-α-induced Caco-2 cell model[J]. Food Research International,2021,139:109833. doi: 10.1016/j.foodres.2020.109833
[37]	张瑞麟. 英国红芸豆多糖提取、结构鉴定及对T2DM小鼠改善作用研究[D]. 大庆: 黑龙江八一农垦大学, 2022 ZHANG R L. Study on extraction, structure identification and improvement effect of British red kidney bean polysaccharides on T2DM mice[D]. Daqing: Heilongjiang Bayi Agricultural University, 2022.
[38]	鲁朝凤, 黄佳琦, 黄勇桦, 等. 青稞膳食纤维和多酚对肠道微生物的协同调节作用[J]. 食品与发酵工业,2021,47(8):6−13. [LU C F, HUANG J Q, HUANG Y H, et al. Synergetic regulation of gut microbiota by dietary fiber and phenolic compounds in hulless barley[J]. Food and Fermentation Industries,2021,47(8):6−13.