本文主要研究内容
作者方程（2019）在《基于组学的白酒肝损伤和肠道菌群干预调节机制研究》一文中研究指出：研究背景白酒是中国传统发酵酒精饮料,承载着重要的文化内涵和经济价值。在近千年的发展历程中,白酒形成了许多与健康有关的理论体系。然而过去由于对白酒成分认知的不足,未能从现代医学的角度很好的解释其中的机理。近年来的研究发现,白酒中除了含有水和乙醇,还包含上千种化合物。然而,这些研究大部分仅关注具有风味贡献的挥发性成分,而对口感具有贡献的难挥发性成分鲜有文献报道。并且,混菌固态发酵的中国白酒与单菌液态发酵的西方蒸馏酒之间在成分上的异同也不清楚。此外,文献报道白酒中由于含有多种具有潜在生物活性的物质,对哺乳动物的健康有重要影响,但是缺乏有效的体内实验支撑这些物质的生理功能。最近关于酒精性肝脏疾病(Alcoholic liver disease,ALD)的研究表明,肠道微生物可以影响肠屏障功能并在ALD的发展中起重要作用。此外,由于肠道菌还扮演重要的代谢功能,我们猜测白酒中潜在的活性物质可能可以通过调节肠道菌群结构及其代谢改善乙醇诱导的肝损伤。研究目的(1)利用代谢组学的研究手段全面分析白酒的化学组成(挥发性成分与难挥发性成分),并通过识别不同蒸馏酒中的特征性成分,系统比较中西方蒸馏酒之间成分上的异同;(2)通过动物模型验证中国白酒中的成分是否可以减轻乙醇诱导的肝损伤,并从肠道菌群及其代谢的角度阐明潜在的机制。研究方法基于非靶向代谢组学的研究思路,分别利用顶空(Headspace,HS)固相微萃取技术(Solid-phase microextraction,SPME)和N,O-双(三甲基硅烷基)三氟乙酰胺(BSTFA)衍生化对国内外24种蒸馏酒(包括6种典型中国白酒和18种典型西方蒸馏酒)的挥发性成分和难挥发性成分进行前处理,再结合气相色谱-飞行时间质谱(Gas chromatography-time-of-flight mass spectrometers,GC-TOFMS)分别对挥发性成分和难挥发性成分进行检测,然后通过网络分析对不同类型蒸馏酒中的特征性成分进行识别,最终系统比较中西方蒸馏酒之间成分上的异同。通过建立酱香型白酒(CB)和食用酒精(EtOH)干预的小鼠ALD模型,从表型、生化以及肝脏病理的角度比较CB与EtOH干预之间动物的肝损伤差异,并分别利用高通量测序和代谢组学的研究手段对造模动物的肠道菌群结构及其代谢物进行分析。此外,通过组织免疫荧光和荧光定量PCR对动物的肠屏障功能进行了分析。最后,基于白酒全谱成分的认知,建立低酒精剂量的CB干预小鼠模型(低剂量模型)和调配酒(BB)干预的小鼠模型(调配酒模型),对结果进行验证。研究结果(1)在24种蒸馏酒中一共检测到879个挥发性物质的峰,并定性出661种物质,主要包括芳香族(166种)、酯类(159种)、醛酮类(105种)和醇类(92种),其中酱香型白酒中的挥发性成分最为复杂;检测到268个难挥发性物质的峰,并定性出160种物质,主要包括有机酸(46种)、糖类(43种)、芳香族(30种)和醇类(27种),其中威士忌中的难挥发性成分最为复杂。通过比较中国白酒与西方蒸馏酒成分上的异同发现,乙酯类物质、部分芳香族化合物和醇类是大部分蒸馏酒共有,而糖类物质和脂肪醇形成的酯类在中西方蒸馏酒之间存在明显差异。(2)通过对动物模型的表型分析发现CB与EtOH干预都导致了小鼠进食量和体重的下降以及肝重体重比的上升,但是CB干预组小鼠的肝重体重比低于EtOH干预组。进一步的生化分析发现,CB和EtOH干预都导致了血清谷丙转氨酶(Alanine aminotransferase,ALT)和谷草转氨酶(Aspartate aminotransferase,AST)以及肝脏甘油三酯(Triglyceride,TG)和硫代巴比妥酸反应物(Thiobarbituric acid reactive substance,TBARS)水平的升高,但是EtOH干预小鼠的生化改变在整个造模期间都比CB干预组明显。肝脏病理结果与生化分析一致,CB和EtOH干预都导致了小鼠肝脏的脂肪变性与脂滴累积,并且EtOH干预比CB干预更加明显。(3)针对盲肠菌群的16S rRNA基因扩增子测序表明,CB与EtOH干预都导致小鼠盲肠菌群结构发生了改变。同时还发现CB干预与EtOH干预之间也存在明显不同,并且随干预时间的延长,肠道菌群差异有扩大的趋势。进一步通过冗余分析(Redundancy analysis,RDA)找出了CB干预与EtOH干预之间有28个关键OTU存在明显差异,其中CB组中有9个OTU(主要为Akkermansia、Ruminococcus、Pseudoflavonifractor和部分未鉴定的S24-7等微生物属)的丰度显著高于乙醇组;19个OTU(主要为Prevotella、Bacteroides、Helicobacter和部分未鉴定的S24-7等微生物属)的丰度则低于乙醇组。(4)肠屏障功能分析发现CB干预组小鼠回肠组织的紧密连接蛋白Occludin及其mRNA的表达水平高于乙醇干预组,而Claudin-2及其mRNA的表达则低于EtOH干预组。进一步通过分析血清和肝脏中的革兰氏阴性菌细胞壁成分内毒素(Lipopolysaccharide,LPS)的含量发现,CB干预组小鼠血清和肝脏的LPS水平均低于EtOH组,说明CB干预组小鼠的肠屏障功能比EtOH组更加完整。(5)全谱代谢分析发现CB干预与EtOH干预还导致了盲肠内容物的代谢轮廓发生分化。通过差异代谢物分析发现CB组小鼠盲肠内容物中Arachidic acid、Behenic acid、Heptadecanoic acid和Stearic acid等饱和长链脂肪酸(Long-chain fatty acid,LCFA)的含量高于EtOH组,以及Myoinositol、D-galactose、L-sorbose、Rhamnose和Sucrose等多种可发酵性糖的含量低于EtOH组。进一步通过靶向分析定量检测短链脂肪酸(Shortchain fatty acid,SCFA)的含量发现,CB组肠道内容物中的SCFA含量明显高于EtOH组,尤其是乙酸和丁酸。结果暗示CB干预组肠道中较低含量的可发酵性糖类可能被肠道菌代谢成为了SCFA。(6)验证模型中,通过对低剂量模型小鼠的表型、生化和肝脏病理分析发现,在干预12周后,EtOH组小鼠出现明显的肝损伤,而CB组小鼠未观察到明显肝损伤;通过对调配酒模型小鼠的表型、生化和肝脏病理进行分析,结果显示调配酒干预与CB干预小鼠的肝损伤水平基本一致,且均低于EtOH组。研究结论基于BSTFA衍生化结合GC-TOFMS检测蒸馏酒中的难挥发性成分的方法可以有效弥补基于HS-SPME-GC-TOFMS的挥发性成分检测方法的不足,两种方法的结合可以被广泛应用于各种酒精饮料的全谱成分分析。中国白酒与西方蒸馏酒之间由于原料、生产工艺和微生物以及存储容器的不同导致在成分上存在明显的区别。动物模型分析表明CB中的挥发性物质可以减轻乙醇诱导的肝损伤。潜在的机制可能与这些挥发性物质的抗菌性和抗氧化性有关。一方面,这些挥发性物质通过调节肠道菌群,保护乙醇诱导的肠道中A.muciniphila的消耗,从而保护肠屏障的功能,这样从肠道中泄漏的促炎性的LPS减少,减轻了乙醇诱导的肝损伤。另一方面,这些挥发性物质通过调节肠道菌群结构,改变了肠道微生物的代谢功能,诱导产生了较高含量的功能性饱和LCFAs和SCFAs。
Abstract
yan jiu bei jing bai jiu shi zhong guo chuan tong fa jiao jiu jing yin liao ,cheng zai zhao chong yao de wen hua nei han he jing ji jia zhi 。zai jin qian nian de fa zhan li cheng zhong ,bai jiu xing cheng le hu duo yu jian kang you guan de li lun ti ji 。ran er guo qu you yu dui bai jiu cheng fen ren zhi de bu zu ,wei neng cong xian dai yi xue de jiao du hen hao de jie shi ji zhong de ji li 。jin nian lai de yan jiu fa xian ,bai jiu zhong chu le han you shui he yi chun ,hai bao han shang qian chong hua ge wu 。ran er ,zhe xie yan jiu da bu fen jin guan zhu ju you feng wei gong suo de hui fa xing cheng fen ,er dui kou gan ju you gong suo de nan hui fa xing cheng fen xian you wen suo bao dao 。bing ju ,hun jun gu tai fa jiao de zhong guo bai jiu yu chan jun ye tai fa jiao de xi fang zheng liu jiu zhi jian zai cheng fen shang de yi tong ye bu qing chu 。ci wai ,wen suo bao dao bai jiu zhong you yu han you duo chong ju you qian zai sheng wu huo xing de wu zhi ,dui bu ru dong wu de jian kang you chong yao ying xiang ,dan shi que fa you xiao de ti nei shi yan zhi cheng zhe xie wu zhi de sheng li gong neng 。zui jin guan yu jiu jing xing gan zang ji bing (Alcoholic liver disease,ALD)de yan jiu biao ming ,chang dao wei sheng wu ke yi ying xiang chang bing zhang gong neng bing zai ALDde fa zhan zhong qi chong yao zuo yong 。ci wai ,you yu chang dao jun hai ban yan chong yao de dai xie gong neng ,wo men cai ce bai jiu zhong qian zai de huo xing wu zhi ke neng ke yi tong guo diao jie chang dao jun qun jie gou ji ji dai xie gai shan yi chun you dao de gan sun shang 。yan jiu mu de (1)li yong dai xie zu xue de yan jiu shou duan quan mian fen xi bai jiu de hua xue zu cheng (hui fa xing cheng fen yu nan hui fa xing cheng fen ),bing tong guo shi bie bu tong zheng liu jiu zhong de te zheng xing cheng fen ,ji tong bi jiao zhong xi fang zheng liu jiu zhi jian cheng fen shang de yi tong ;(2)tong guo dong wu mo xing yan zheng zhong guo bai jiu zhong de cheng fen shi fou ke yi jian qing yi chun you dao de gan sun shang ,bing cong chang dao jun qun ji ji dai xie de jiao du chan ming qian zai de ji zhi 。yan jiu fang fa ji yu fei ba xiang dai xie zu xue de yan jiu sai lu ,fen bie li yong ding kong (Headspace,HS)gu xiang wei cui qu ji shu (Solid-phase microextraction,SPME)he N,O-shuang (san jia ji gui wan ji )san fu yi xian an (BSTFA)yan sheng hua dui guo nei wai 24chong zheng liu jiu (bao gua 6chong dian xing zhong guo bai jiu he 18chong dian xing xi fang zheng liu jiu )de hui fa xing cheng fen he nan hui fa xing cheng fen jin hang qian chu li ,zai jie ge qi xiang se pu -fei hang shi jian zhi pu (Gas chromatography-time-of-flight mass spectrometers,GC-TOFMS)fen bie dui hui fa xing cheng fen he nan hui fa xing cheng fen jin hang jian ce ,ran hou tong guo wang lao fen xi dui bu tong lei xing zheng liu jiu zhong de te zheng xing cheng fen jin hang shi bie ,zui zhong ji tong bi jiao zhong xi fang zheng liu jiu zhi jian cheng fen shang de yi tong 。tong guo jian li jiang xiang xing bai jiu (CB)he shi yong jiu jing (EtOH)gan yu de xiao shu ALDmo xing ,cong biao xing 、sheng hua yi ji gan zang bing li de jiao du bi jiao CByu EtOHgan yu zhi jian dong wu de gan sun shang cha yi ,bing fen bie li yong gao tong liang ce xu he dai xie zu xue de yan jiu shou duan dui zao mo dong wu de chang dao jun qun jie gou ji ji dai xie wu jin hang fen xi 。ci wai ,tong guo zu zhi mian yi ying guang he ying guang ding liang PCRdui dong wu de chang bing zhang gong neng jin hang le fen xi 。zui hou ,ji yu bai jiu quan pu cheng fen de ren zhi ,jian li di jiu jing ji liang de CBgan yu xiao shu mo xing (di ji liang mo xing )he diao pei jiu (BB)gan yu de xiao shu mo xing (diao pei jiu mo xing ),dui jie guo jin hang yan zheng 。yan jiu jie guo (1)zai 24chong zheng liu jiu zhong yi gong jian ce dao 879ge hui fa xing wu zhi de feng ,bing ding xing chu 661chong wu zhi ,zhu yao bao gua fang xiang zu (166chong )、zhi lei (159chong )、quan tong lei (105chong )he chun lei (92chong ),ji zhong jiang xiang xing bai jiu zhong de hui fa xing cheng fen zui wei fu za ;jian ce dao 268ge nan hui fa xing wu zhi de feng ,bing ding xing chu 160chong wu zhi ,zhu yao bao gua you ji suan (46chong )、tang lei (43chong )、fang xiang zu (30chong )he chun lei (27chong ),ji zhong wei shi ji zhong de nan hui fa xing cheng fen zui wei fu za 。tong guo bi jiao zhong guo bai jiu yu xi fang zheng liu jiu cheng fen shang de yi tong fa xian ,yi zhi lei wu zhi 、bu fen fang xiang zu hua ge wu he chun lei shi da bu fen zheng liu jiu gong you ,er tang lei wu zhi he zhi fang chun xing cheng de zhi lei zai zhong xi fang zheng liu jiu zhi jian cun zai ming xian cha yi 。(2)tong guo dui dong wu mo xing de biao xing fen xi fa xian CByu EtOHgan yu dou dao zhi le xiao shu jin shi liang he ti chong de xia jiang yi ji gan chong ti chong bi de shang sheng ,dan shi CBgan yu zu xiao shu de gan chong ti chong bi di yu EtOHgan yu zu 。jin yi bu de sheng hua fen xi fa xian ,CBhe EtOHgan yu dou dao zhi le xie qing gu bing zhuai an mei (Alanine aminotransferase,ALT)he gu cao zhuai an mei (Aspartate aminotransferase,AST)yi ji gan zang gan you san zhi (Triglyceride,TG)he liu dai ba bi tuo suan fan ying wu (Thiobarbituric acid reactive substance,TBARS)shui ping de sheng gao ,dan shi EtOHgan yu xiao shu de sheng hua gai bian zai zheng ge zao mo ji jian dou bi CBgan yu zu ming xian 。gan zang bing li jie guo yu sheng hua fen xi yi zhi ,CBhe EtOHgan yu dou dao zhi le xiao shu gan zang de zhi fang bian xing yu zhi di lei ji ,bing ju EtOHgan yu bi CBgan yu geng jia ming xian 。(3)zhen dui mang chang jun qun de 16S rRNAji yin kuo zeng zi ce xu biao ming ,CByu EtOHgan yu dou dao zhi xiao shu mang chang jun qun jie gou fa sheng le gai bian 。tong shi hai fa xian CBgan yu yu EtOHgan yu zhi jian ye cun zai ming xian bu tong ,bing ju sui gan yu shi jian de yan chang ,chang dao jun qun cha yi you kuo da de qu shi 。jin yi bu tong guo rong yu fen xi (Redundancy analysis,RDA)zhao chu le CBgan yu yu EtOHgan yu zhi jian you 28ge guan jian OTUcun zai ming xian cha yi ,ji zhong CBzu zhong you 9ge OTU(zhu yao wei Akkermansia、Ruminococcus、Pseudoflavonifractorhe bu fen wei jian ding de S24-7deng wei sheng wu shu )de feng du xian zhe gao yu yi chun zu ;19ge OTU(zhu yao wei Prevotella、Bacteroides、Helicobacterhe bu fen wei jian ding de S24-7deng wei sheng wu shu )de feng du ze di yu yi chun zu 。(4)chang bing zhang gong neng fen xi fa xian CBgan yu zu xiao shu hui chang zu zhi de jin mi lian jie dan bai Occludinji ji mRNAde biao da shui ping gao yu yi chun gan yu zu ,er Claudin-2ji ji mRNAde biao da ze di yu EtOHgan yu zu 。jin yi bu tong guo fen xi xie qing he gan zang zhong de ge lan shi yin xing jun xi bao bi cheng fen nei du su (Lipopolysaccharide,LPS)de han liang fa xian ,CBgan yu zu xiao shu xie qing he gan zang de LPSshui ping jun di yu EtOHzu ,shui ming CBgan yu zu xiao shu de chang bing zhang gong neng bi EtOHzu geng jia wan zheng 。(5)quan pu dai xie fen xi fa xian CBgan yu yu EtOHgan yu hai dao zhi le mang chang nei rong wu de dai xie lun kuo fa sheng fen hua 。tong guo cha yi dai xie wu fen xi fa xian CBzu xiao shu mang chang nei rong wu zhong Arachidic acid、Behenic acid、Heptadecanoic acidhe Stearic aciddeng bao he chang lian zhi fang suan (Long-chain fatty acid,LCFA)de han liang gao yu EtOHzu ,yi ji Myoinositol、D-galactose、L-sorbose、Rhamnosehe Sucrosedeng duo chong ke fa jiao xing tang de han liang di yu EtOHzu 。jin yi bu tong guo ba xiang fen xi ding liang jian ce duan lian zhi fang suan (Shortchain fatty acid,SCFA)de han liang fa xian ,CBzu chang dao nei rong wu zhong de SCFAhan liang ming xian gao yu EtOHzu ,you ji shi yi suan he ding suan 。jie guo an shi CBgan yu zu chang dao zhong jiao di han liang de ke fa jiao xing tang lei ke neng bei chang dao jun dai xie cheng wei le SCFA。(6)yan zheng mo xing zhong ,tong guo dui di ji liang mo xing xiao shu de biao xing 、sheng hua he gan zang bing li fen xi fa xian ,zai gan yu 12zhou hou ,EtOHzu xiao shu chu xian ming xian de gan sun shang ,er CBzu xiao shu wei guan cha dao ming xian gan sun shang ;tong guo dui diao pei jiu mo xing xiao shu de biao xing 、sheng hua he gan zang bing li jin hang fen xi ,jie guo xian shi diao pei jiu gan yu yu CBgan yu xiao shu de gan sun shang shui ping ji ben yi zhi ,ju jun di yu EtOHzu 。yan jiu jie lun ji yu BSTFAyan sheng hua jie ge GC-TOFMSjian ce zheng liu jiu zhong de nan hui fa xing cheng fen de fang fa ke yi you xiao mi bu ji yu HS-SPME-GC-TOFMSde hui fa xing cheng fen jian ce fang fa de bu zu ,liang chong fang fa de jie ge ke yi bei an fan ying yong yu ge chong jiu jing yin liao de quan pu cheng fen fen xi 。zhong guo bai jiu yu xi fang zheng liu jiu zhi jian you yu yuan liao 、sheng chan gong yi he wei sheng wu yi ji cun chu rong qi de bu tong dao zhi zai cheng fen shang cun zai ming xian de ou bie 。dong wu mo xing fen xi biao ming CBzhong de hui fa xing wu zhi ke yi jian qing yi chun you dao de gan sun shang 。qian zai de ji zhi ke neng yu zhe xie hui fa xing wu zhi de kang jun xing he kang yang hua xing you guan 。yi fang mian ,zhe xie hui fa xing wu zhi tong guo diao jie chang dao jun qun ,bao hu yi chun you dao de chang dao zhong A.muciniphilade xiao hao ,cong er bao hu chang bing zhang de gong neng ,zhe yang cong chang dao zhong xie lou de cu yan xing de LPSjian shao ,jian qing le yi chun you dao de gan sun shang 。ling yi fang mian ,zhe xie hui fa xing wu zhi tong guo diao jie chang dao jun qun jie gou ,gai bian le chang dao wei sheng wu de dai xie gong neng ,you dao chan sheng le jiao gao han liang de gong neng xing bao he LCFAshe SCFAs。
论文参考文献
论文详细介绍
论文作者分别是来自江南大学的方程，发表于刊物江南大学2019-10-21论文，是一篇关于白酒论文,代谢组学论文,挥发性成分论文,难挥发性成分论文,肠道菌群论文,肠屏障功能论文,短链脂肪酸论文,江南大学2019-10-21论文的文章。本文可供学术参考使用，各位学者可以免费参考阅读下载，文章观点不代表本站观点，资料来自江南大学2019-10-21论文网站，若本站收录的文献无意侵犯了您的著作版权，请联系我们删除。
标签：白酒论文; 代谢组学论文; 挥发性成分论文; 难挥发性成分论文; 肠道菌群论文; 肠屏障功能论文; 短链脂肪酸论文; 江南大学2019-10-21论文;