Article(id=1320880575842468155, tenantId=1146029695717560320, journalId=1260987677001138203, issueId=1320880262012056083, articleNumber=null, orderNo=null, doi=10.13386/j.issn1002-0306.2025110022, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1762272000000, receivedDateStr=2025-11-05, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1792852070055, onlineDateStr=2026-10-24, pubDate=1789401600000, pubDateStr=2026-09-15, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1792852070055, onlineIssueDateStr=2026-10-24, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1792852070055, creator=13701087609, updateTime=1792852070055, updator=13701087609, issue=Issue{id=1320880262012056083, tenantId=1146029695717560320, journalId=1260987677001138203, year='2026', volume='47', issue='18', pageStart='1', pageEnd='613', issueExtLink='null', onlineDate='null', pubDate='1789401600000', pubDateStr='2026-09-15', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=1, specialIssue=null, createTime=1792851995232, creator='13701087609', updateTime=1792921655980, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1321172440479290195, tenantId=1146029695717560320, journalId=1260987677001138203, issueId=1320880262012056083, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1321172440479290196, tenantId=1146029695717560320, journalId=1260987677001138203, issueId=1320880262012056083, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=14, endPage=23, ext={EN=ArticleExt(id=1320880576073154876, articleId=1320880575842468155, tenantId=1146029695717560320, journalId=1260987677001138203, language=EN, title=Isolation and Identification of Bacillus licheniformis in High-temperature Daqu and Analysis of Its Aroma-producing Characteristics, columnId=1320880408649118389, journalTitle=Science and Technology of Food Industry, columnName=Guest Editor Column—Frontiers and Applications of Food Flavor Perception‌ (Guest Editors: LIU Yuan, SUN Jinyuan, KOU Xingran, CHENG Huan; Industry Advisor: CHEN Li), runingTitle=null, highlight=null, articleAbstract=

To clarify the aroma-producing characteristics of Bacillus licheniformis in high-temperature Daqu of sauce-flavor Baijiu and its contribution to Baijiu flavor formation, this study isolated and purified 16 strains of B. licheniformis from high-temperature Daqu. Using programmed temperature solid-state fermentation combined with artificial sensory evaluation, as well as headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography–mass spectrometry (GC-MS), the relationship between metabolic profiles and sensory attributes of different strains of the same species was systematically analyzed. The results showed that the flavor characteristics of solid-state fermentation products from different strains could be classified into five categories: Roasted/nutty-fatty aroma, douchi aroma, sulfurous notes, soy sauce/fermented paste aroma, and fruity-acidic aroma. A total of 198 volatile compounds were identified through volatile component analysis. Pyrazines were dominant in all strain types, while the composition of other volatile compounds showed distinct differences: Aldehydes, ketones, and alcohols were more abundant in strains with roasted/nutty-fatty aroma, douchi aroma, sulfurous notes; Aldehydes, acids, and alcohols were higher in soy sauce/fermented paste aroma strains; Acids and ketones were predominant in fruity-acidic aroma strains. Key differential aroma components were further screened by orthogonal partial least squares-discriminant analysis (OPLS-DA) with a variable importance in projection (VIP) value>1.0, and combined with a relative odor activity value (ROAV)>1.0. Five substances were ultimately identified as crucial for discriminating the aroma profiles of different sensory categories of B. licheniformis: 4-methoxyphenol (a key common component of roasted/nutty-fatty aroma, douchi aroma, and soy sauce/fermented paste aroma strains), isovaleric acid (a key common component of douchi aroma and fruity-acidic aroma strains), 2-octanone (a key component of fruity-acidic aroma strains), 2,3,5-trimethylpyrazine (a key component of roasted/nutty-fatty aroma, douchi aroma, and sulfurous notes strains), and 2,3,5,6-tetramethylpyrazine (a key component of soy sauce/fermented paste aroma strains).

, authors=Jiali WANG1, Zhuo HE1, Songtao WANG2, Hongmei MING1, Yang YANG2, Mansi NIU2, Qiuwei ZHAO3, Zhengjie ZHANG1, Shu LI2, *, Zonghua AO1, 2, *, authorsList=Jiali WANG, Zhuo HE, Songtao WANG, Hongmei MING, Yang YANG, Mansi NIU, Qiuwei ZHAO, Zhengjie ZHANG, Shu LI, Zonghua AO, authorCompany=null, correspAuthors=Shu LI, Zonghua AO, authorNote=null, correspAuthorsNote=null, copyrightStatement=Copyright © 2026 Science and Technology of Food Industry. All rights reserved., copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1320880579361489229, articleId=1320880575842468155, tenantId=1146029695717560320, journalId=1260987677001138203, language=CN, title=高温大曲地衣芽孢杆菌的分离鉴定及其产香特性解析, columnId=1320880408842056375, journalTitle=食品工业科技, columnName=特邀主编专栏—食品风味感知前沿与应用(客座主编:刘源、孙金沅、寇兴然、程焕;产业顾问:陈丽), runingTitle=null, highlight=null, articleAbstract=

为明确酱香型白酒高温大曲中地衣芽孢杆菌(Bacillus licheniformis)的产香特性及其对白酒风味形成的贡献,本研究从高温大曲中分离纯化得到16株B. licheniformis,采用程序升温固态发酵结合人工感官评定,并利用顶空固相微萃取(Headspace Solid-Phase Microextraction,HS-SPME)和气相色谱-质谱(Gas Chromatography-Mass Spectrometry,GC-MS)联用技术,系统分析同种不同株的代谢谱与其感官属性之间的关联。结果表明,不同菌株固态发酵产物的风味特征可分为五类:烘焙香-坚果油脂香、豆豉香、硫味、酱香-酱油香及果酸香。挥发性组分分析共鉴定出198种化合物,其中吡嗪类物质在各类菌株中均占主导,其他挥发性成分组成存在明显差异:烘焙香-坚果油脂香、豆豉香及硫味菌株中醛类、酮类和醇类较多;酱香-酱油香菌株中醛类、酸类和醇类较多;果酸香菌株则为酸类和酮类较多。进一步通过正交偏最小二乘判别分析(Orthogonal Partial Least Squares-Discriminant Analysis,OPLS-DA)与VIP值(Variable Importance in Projection,VIP)大于1.0筛选出关键差异香气成分,并结合相对气味活度值(Relative Odor Activity Value,ROAV)大于1.0,最终确定这5种物质对不同感官类别B. licheniformis的香气特征具有重要判别作用:4-甲氧基苯酚(为烘焙香-坚果油脂香、豆豉香及酱香-酱油香菌株的共同关键成分)、异戊酸(豆豉香与果酸香菌株共同的共同关键成分)、2-辛酮(果酸香菌株的关键成分)、2,3,5-三甲基吡嗪(烘焙香-坚果油脂香、豆豉香及硫味菌株的关键成分)、2,3,5,6-四甲基吡嗪(酱香-酱油香菌株的关键成分)。

, authors=王嘉丽1, 何卓1, 王松涛2, 明红梅1, 杨阳2, 牛曼思2, 赵秋伟3, 张正杰1, 李姝2, *, 敖宗华1, 2, *, authorsList=王嘉丽, 何卓, 王松涛, 明红梅, 杨阳, 牛曼思, 赵秋伟, 张正杰, 李姝, 敖宗华, authorCompany=null, correspAuthors=李姝, 敖宗华, authorNote=

王嘉丽(2000−),女,硕士研究生,研究方向:微生物发酵代谢调控,E-mail:

, correspAuthorsNote=
李姝(1991−),女,博士,工程师,研究方向:白酒生产技术、酿造微生物、天然产物与功效评价,E-mail:。
敖宗华(1971−),男,博士,教授级高级工程师,研究方向:发酵产品风味物质研究和功能因子分析,E-mail:。
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Science and Technology of Food Industry, 2025, 46(11): 263−272., articleTitle=null, refAbstract=null), Reference(id=1320880593638900181, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, doi=null, pmid=null, pmcid=null, year=null, volume=null, issue=null, pageStart=null, pageEnd=null, url=null, language=null, rfNumber=32, rfOrder=43, authorNames=null, journalName=null, refType=null, unstructuredReference=DENG Jie, ZHENG Jia, LI Xing, et al. Characterization of Qu-aroma of medium–high temperature Daqu from different production areas using sensory evaluation, E-nose, and GC–MS/O analysis[J]. 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tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, xref=2, ext=[AuthorCompanyExt(id=1320880579676062034, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, companyId=1320880579663479121, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2National Engineering Research Center of Solid-State Brewing, Luzhou Laojiao Co., Ltd., Luzhou 646000, China), AuthorCompanyExt(id=1320880579684450643, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, companyId=1320880579663479121, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2泸州老窖股份有限公司,国家固态酿造工程技术研究中心,四川泸州 646000)]), AuthorCompany(id=1320880579755753812, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, xref=3, ext=[AuthorCompanyExt(id=1320880579764142421, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, companyId=1320880579755753812, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3Department of Microbial Physiological & Metabolic Engineering, State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China), AuthorCompanyExt(id=1320880579776725334, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, companyId=1320880579755753812, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3中国科学院微生物研究所,微生物资源前期开发国家重点实验室,微生物生理与代谢工程研究室,北京 100101)])], figs=[ArticleFig(id=1320880585174794644, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.1, caption=Schematic diagram of the programmed temperature, figureFileSmall=YLyqPA4ilrfkSgPJ19Dnwg==, figureFileBig=GuiDvxJ9IGfPkufKnF52Bg==, tableContent=null), ArticleFig(id=1320880585288040853, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图1, caption=程序升温示意图, figureFileSmall=YLyqPA4ilrfkSgPJ19Dnwg==, figureFileBig=GuiDvxJ9IGfPkufKnF52Bg==, tableContent=null), ArticleFig(id=1320880587041259926, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.2, caption=Fermentation product of aroma-producing functional bacteria for fragrance evaluation and flavor sunburst chart, figureFileSmall=zH4Ywrq3hfZeQKmPunqifw==, figureFileBig=Tf/f1jGxtsouPuH+29gfIg==, tableContent=null), ArticleFig(id=1320880587116757399, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图2, caption=产香功能菌发酵物闻香评定风味旭日图, figureFileSmall=zH4Ywrq3hfZeQKmPunqifw==, figureFileBig=Tf/f1jGxtsouPuH+29gfIg==, tableContent=null), ArticleFig(id=1320880587183866264, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.3, caption=Colonial morphology and strain designations of B. licheniformis, figureFileSmall=r/svLQ2T9XzBAYM78J0o8g==, figureFileBig=k/MDPJKf1NfZxcwGItIFlg==, tableContent=null), ArticleFig(id=1320880587267752345, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图3, caption=B. licheniformis菌落形状及其编号, figureFileSmall=r/svLQ2T9XzBAYM78J0o8g==, figureFileBig=k/MDPJKf1NfZxcwGItIFlg==, tableContent=null), ArticleFig(id=1320880587355832730, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.4, caption=Phylogenetic tree of the 16 strains based on 16S rRNA gene sequences, figureFileSmall=XP6Fh0FOq88oCeMxaJLQyw==, figureFileBig=MYCFZryaYBwgVkv5mmlOkQ==, tableContent=null), ArticleFig(id=1320880587439718811, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图4, caption=基于16株菌株16S rRNA基因序列构建的系统进化树, figureFileSmall=XP6Fh0FOq88oCeMxaJLQyw==, figureFileBig=MYCFZryaYBwgVkv5mmlOkQ==, tableContent=null), ArticleFig(id=1320880587540382108, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.5, caption=Sensory radar chart of solid-state fermentation products from 16 B. licheniformis, figureFileSmall=W4zmOXqxG/dHZ9FPBZMvkg==, figureFileBig=evyOB+bOODN62F9pWH1T/w==, tableContent=null), ArticleFig(id=1320880587620073885, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图5, caption=16株B. licheniformis固态发酵物感官雷达图

注:A为烘焙香-坚果油脂香菌株;B为豆豉香菌株;C为硫味菌株;D为酱香-酱油香菌株;E和F为果酸香菌株。

, figureFileSmall=W4zmOXqxG/dHZ9FPBZMvkg==, figureFileBig=evyOB+bOODN62F9pWH1T/w==, tableContent=null), ArticleFig(id=1320880587678794142, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.6, caption=Stacked bar chart of volatile flavor compound varieties produced by fermentation of 16 B. licheniformis strains (A), stacked bar chart of volatile flavor compound contents (B), and Upset plot of flavor categories (C), figureFileSmall=BP9tH6PcNJfDqni0KvZMmg==, figureFileBig=Vqdrt0zZ9TXZfPRqaZJWGg==, tableContent=null), ArticleFig(id=1320880587758485919, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图6, caption=16种B. licheniformis发酵产生的挥发性风味物质种类堆积柱状图(A)、挥发性风味物质含量堆积柱状图(B)、风味种类Upset图(C)

注:a. 烘焙香-坚果油脂香;b. 豆豉香;c. 硫味;d. 酱香-酱油香;e. 果酸香。

, figureFileSmall=BP9tH6PcNJfDqni0KvZMmg==, figureFileBig=Vqdrt0zZ9TXZfPRqaZJWGg==, tableContent=null), ArticleFig(id=1320880587838177696, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.7, caption=Grouping of B. licheniformis based on sensory evaluation, figureFileSmall=cf0+emwneZG47FAhyTw2eQ==, figureFileBig=0dqw+/zaV24s4SD5nswB4w==, tableContent=null), ArticleFig(id=1320880587905286561, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图7, caption=不同感官评定B. licheniformis风味的聚类分析

注:A. PCA图;B. OPLS-DA图;C. 模型交叉验证结果。

, figureFileSmall=cf0+emwneZG47FAhyTw2eQ==, figureFileBig=0dqw+/zaV24s4SD5nswB4w==, tableContent=null), ArticleFig(id=1320880587976589730, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Fig.8, caption=Clustered heatmap of differential aroma compounds in sensorily distinct groups of B. licheniformis, figureFileSmall=iL9sQcCrj0woKc/S97mnng==, figureFileBig=b0Bkam1TyWMsVWUi0dT1Lg==, tableContent=null), ArticleFig(id=1320880588060475811, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=图8, caption=不同感官评定B. licheniformis的差异香气成分聚类热图

注:a:烘焙香-坚果油脂香、b:豆豉香、c:硫味、d:酱香-酱油香、e:果酸香、f:生麦香(KB)。

, figureFileSmall=iL9sQcCrj0woKc/S97mnng==, figureFileBig=b0Bkam1TyWMsVWUi0dT1Lg==, tableContent=null), ArticleFig(id=1320880588144361892, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Table 1, caption=

Isolation and colonial morphology of B. licheniformis

, figureFileSmall=null, figureFileBig=null, tableContent=
序号菌株编号分离情况菌落形态
分离温度(℃)分离培养基形态边缘表面性状隆起度正面颜色透明度
1TSA-37-337TSA不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
2TSA-50-2650TSA不规则锯齿状整体光滑整体隆起褶皱灰白色不透明
3TSA-50-2750TSA不规则锯齿状整体光滑整体隆起黄色不透明
4TSA-37-2837TSA近圆形锯齿状整体粗糙中央隆起褶皱黄白色不透明
5ISP2-55-3355ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
6ISP2-55-3455ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
7ISP2-55-3555ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
8ISP2-55-3655ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起白色半透明
9ISP2-55-3755ISP2不规则锯齿状整体光滑整体隆起黄色不透明
10ISP2-55-3955ISP2不规则锯齿状边缘光滑,中间粗糙中央隆起褶皱黄白色不透明
11ISP2-55-4055ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
12ISP2-55-4155ISP2近圆形波浪状整体光滑整体轻微隆起黄白色不透明
13G2-55-4355G2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
14TSA-50-4750TSA不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
15TSA-37-4837TSA不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
16NA-55-5155NA近圆形锯齿状整体粗糙整体轻微褶皱白色不透明
), ArticleFig(id=1320880588232442277, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=表1, caption=

B. licheniformis分离情况以及菌落形状描述

, figureFileSmall=null, figureFileBig=null, tableContent=
序号菌株编号分离情况菌落形态
分离温度(℃)分离培养基形态边缘表面性状隆起度正面颜色透明度
1TSA-37-337TSA不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
2TSA-50-2650TSA不规则锯齿状整体光滑整体隆起褶皱灰白色不透明
3TSA-50-2750TSA不规则锯齿状整体光滑整体隆起黄色不透明
4TSA-37-2837TSA近圆形锯齿状整体粗糙中央隆起褶皱黄白色不透明
5ISP2-55-3355ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
6ISP2-55-3455ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
7ISP2-55-3555ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
8ISP2-55-3655ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起白色半透明
9ISP2-55-3755ISP2不规则锯齿状整体光滑整体隆起黄色不透明
10ISP2-55-3955ISP2不规则锯齿状边缘光滑,中间粗糙中央隆起褶皱黄白色不透明
11ISP2-55-4055ISP2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
12ISP2-55-4155ISP2近圆形波浪状整体光滑整体轻微隆起黄白色不透明
13G2-55-4355G2不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
14TSA-50-4750TSA不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
15TSA-37-4837TSA不规则波浪状边缘光滑,中间粗糙中央轻微隆起黄白色半透明
16NA-55-5155NA近圆形锯齿状整体粗糙整体轻微褶皱白色不透明
), ArticleFig(id=1320880588312134054, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Table 2, caption=

Aroma type classification and strain designations of solid-state fermentates from B. licheniformis

, figureFileSmall=null, figureFileBig=null, tableContent=
菌株编号发酵醅主体风味
TSA-50-26烘焙香-坚果油脂香
TSA-50-27烘焙香-坚果油脂香
NA-55-51豆豉香
TSA-37-48豆豉香
TSA-37-28硫味
TSA-50-47硫味
ISP2-55-41酱香-酱油香
G2-55-43酱香-酱油香
TSA-37-3果酸香
ISP2-55-33果酸香
ISP2-55-34果酸香
ISP2-55-35果酸香
ISP2-55-36果酸香
ISP2-55-37果酸香
ISP2-55-39果酸香
ISP2-55-40果酸香
), ArticleFig(id=1320880588400214439, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=表2, caption=

B. licheniformis固态发酵产物香型分类及菌株编号

, figureFileSmall=null, figureFileBig=null, tableContent=
菌株编号发酵醅主体风味
TSA-50-26烘焙香-坚果油脂香
TSA-50-27烘焙香-坚果油脂香
NA-55-51豆豉香
TSA-37-48豆豉香
TSA-37-28硫味
TSA-50-47硫味
ISP2-55-41酱香-酱油香
G2-55-43酱香-酱油香
TSA-37-3果酸香
ISP2-55-33果酸香
ISP2-55-34果酸香
ISP2-55-35果酸香
ISP2-55-36果酸香
ISP2-55-37果酸香
ISP2-55-39果酸香
ISP2-55-40果酸香
), ArticleFig(id=1320880588488294824, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=EN, label=Table 3, caption=

Analysis of differential aroma compounds in B. licheniformis with distinct sensory profiles

, figureFileSmall=null, figureFileBig=null, tableContent=
CAS香气成分VIP值阈值
(μg/g)
16株菌株香气描述
物质浓度(μg/g)OAVROAV
1124-11-42,3,5,6-四甲基吡嗪2.7796612.55~16.942.55~16.941.32~1.02坚果、泥土、烘烤
14667-55-12,3,5-三甲基吡嗪2.352790.351.14~10.913.24~31.171.68~1.87坚果、青草、可可
100-52-7苯甲醛2.297840.1880.09~5.490.46~29.190.24~1.75苦杏仁、樱桃及坚果香
123-32-02,5-二甲基吡嗪2.240021.820.23~8.820.13~4.850.07~0.29烘烤坚果
503-74-2异戊酸2.054540.0120.4~5.5733.37~464.2117.31~27.87酸味、湿奶酪
123-51-3异戊醇1.635110.20.23~6.451.13~32.240.59~1.94威士忌、香熏、果味
100-51-6苯甲醇1.536130.620.09~5.490.14~8.850.07~0.53蜂蜜、花香、玫瑰、甜香
150-76-54-甲氧基苯酚1.506690.00270.52~4.44193.2~1667.5100动物皮毛
98-86-2苯乙酮1.15115650.03~1.850~0.030甜香、樱桃香气、香草
60-12-8苯乙醇1.113710.560.37~2.060.66~3.680.34~0.22玫瑰香气、花香
111-13-72-辛酮1.008450.040.13~1.923.17~45.811.64~2.75花香、草香
), ArticleFig(id=1320880588597346729, tenantId=1146029695717560320, journalId=1260987677001138203, articleId=1320880575842468155, language=CN, label=表3, caption=

不同感官评定B. licheniformis差异香气成分分析表

, figureFileSmall=null, figureFileBig=null, tableContent=
CAS香气成分VIP值阈值
(μg/g)
16株菌株香气描述
物质浓度(μg/g)OAVROAV
1124-11-42,3,5,6-四甲基吡嗪2.7796612.55~16.942.55~16.941.32~1.02坚果、泥土、烘烤
14667-55-12,3,5-三甲基吡嗪2.352790.351.14~10.913.24~31.171.68~1.87坚果、青草、可可
100-52-7苯甲醛2.297840.1880.09~5.490.46~29.190.24~1.75苦杏仁、樱桃及坚果香
123-32-02,5-二甲基吡嗪2.240021.820.23~8.820.13~4.850.07~0.29烘烤坚果
503-74-2异戊酸2.054540.0120.4~5.5733.37~464.2117.31~27.87酸味、湿奶酪
123-51-3异戊醇1.635110.20.23~6.451.13~32.240.59~1.94威士忌、香熏、果味
100-51-6苯甲醇1.536130.620.09~5.490.14~8.850.07~0.53蜂蜜、花香、玫瑰、甜香
150-76-54-甲氧基苯酚1.506690.00270.52~4.44193.2~1667.5100动物皮毛
98-86-2苯乙酮1.15115650.03~1.850~0.030甜香、樱桃香气、香草
60-12-8苯乙醇1.113710.560.37~2.060.66~3.680.34~0.22玫瑰香气、花香
111-13-72-辛酮1.008450.040.13~1.923.17~45.811.64~2.75花香、草香
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高温大曲地衣芽孢杆菌的分离鉴定及其产香特性解析
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王嘉丽 1 , 何卓 1 , 王松涛 2 , 明红梅 1 , 杨阳 2 , 牛曼思 2 , 赵秋伟 3 , 张正杰 1 , 李姝 2, * , 敖宗华 1, 2, *
食品工业科技 | 特邀主编专栏—食品风味感知前沿与应用(客座主编:刘源、孙金沅、寇兴然、程焕;产业顾问:陈丽) 2026,47(18): 14-23
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食品工业科技 |特邀主编专栏—食品风味感知前沿与应用(客座主编:刘源、孙金沅、寇兴然、程焕;产业顾问:陈丽) 2026 , 47 (18) : 14 -23
高温大曲地衣芽孢杆菌的分离鉴定及其产香特性解析
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王嘉丽1 , 何卓1, 王松涛2, 明红梅1, 杨阳2, 牛曼思2, 赵秋伟3, 张正杰1, 李姝2, * , 敖宗华1, 2, *
作者信息
  • 1四川轻化工大学食品与酿酒工程学院,四川宜宾 644005
  • 2泸州老窖股份有限公司,国家固态酿造工程技术研究中心,四川泸州 646000
  • 3中国科学院微生物研究所,微生物资源前期开发国家重点实验室,微生物生理与代谢工程研究室,北京 100101
通讯作者:
李姝(1991−),女,博士,工程师,研究方向:白酒生产技术、酿造微生物、天然产物与功效评价,E-mail:。
敖宗华(1971−),男,博士,教授级高级工程师,研究方向:发酵产品风味物质研究和功能因子分析,E-mail:。
作者简介:

王嘉丽(2000−),女,硕士研究生,研究方向:微生物发酵代谢调控,E-mail:

Isolation and Identification of Bacillus licheniformis in High-temperature Daqu and Analysis of Its Aroma-producing Characteristics
Jiali WANG1 , Zhuo HE1, Songtao WANG2, Hongmei MING1, Yang YANG2, Mansi NIU2, Qiuwei ZHAO3, Zhengjie ZHANG1, Shu LI2, * , Zonghua AO1, 2, *
Affiliations
  • 1School of Food and Liquor Engineering, Sichuan University of Science and Engineering, Yibin 644005, China
  • 2National Engineering Research Center of Solid-State Brewing, Luzhou Laojiao Co., Ltd., Luzhou 646000, China
  • 3Department of Microbial Physiological & Metabolic Engineering, State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China
出版时间: 2026-09-15 doi: 10.13386/j.issn1002-0306.2025110022
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为明确酱香型白酒高温大曲中地衣芽孢杆菌(Bacillus licheniformis)的产香特性及其对白酒风味形成的贡献,本研究从高温大曲中分离纯化得到16株B. licheniformis,采用程序升温固态发酵结合人工感官评定,并利用顶空固相微萃取(Headspace Solid-Phase Microextraction,HS-SPME)和气相色谱-质谱(Gas Chromatography-Mass Spectrometry,GC-MS)联用技术,系统分析同种不同株的代谢谱与其感官属性之间的关联。结果表明,不同菌株固态发酵产物的风味特征可分为五类:烘焙香-坚果油脂香、豆豉香、硫味、酱香-酱油香及果酸香。挥发性组分分析共鉴定出198种化合物,其中吡嗪类物质在各类菌株中均占主导,其他挥发性成分组成存在明显差异:烘焙香-坚果油脂香、豆豉香及硫味菌株中醛类、酮类和醇类较多;酱香-酱油香菌株中醛类、酸类和醇类较多;果酸香菌株则为酸类和酮类较多。进一步通过正交偏最小二乘判别分析(Orthogonal Partial Least Squares-Discriminant Analysis,OPLS-DA)与VIP值(Variable Importance in Projection,VIP)大于1.0筛选出关键差异香气成分,并结合相对气味活度值(Relative Odor Activity Value,ROAV)大于1.0,最终确定这5种物质对不同感官类别B. licheniformis的香气特征具有重要判别作用:4-甲氧基苯酚(为烘焙香-坚果油脂香、豆豉香及酱香-酱油香菌株的共同关键成分)、异戊酸(豆豉香与果酸香菌株共同的共同关键成分)、2-辛酮(果酸香菌株的关键成分)、2,3,5-三甲基吡嗪(烘焙香-坚果油脂香、豆豉香及硫味菌株的关键成分)、2,3,5,6-四甲基吡嗪(酱香-酱油香菌株的关键成分)。

高温大曲  /  地衣芽孢杆菌  /  风味  /  HS-SPME-GC-MS  /  OPLS-DA

To clarify the aroma-producing characteristics of Bacillus licheniformis in high-temperature Daqu of sauce-flavor Baijiu and its contribution to Baijiu flavor formation, this study isolated and purified 16 strains of B. licheniformis from high-temperature Daqu. Using programmed temperature solid-state fermentation combined with artificial sensory evaluation, as well as headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography–mass spectrometry (GC-MS), the relationship between metabolic profiles and sensory attributes of different strains of the same species was systematically analyzed. The results showed that the flavor characteristics of solid-state fermentation products from different strains could be classified into five categories: Roasted/nutty-fatty aroma, douchi aroma, sulfurous notes, soy sauce/fermented paste aroma, and fruity-acidic aroma. A total of 198 volatile compounds were identified through volatile component analysis. Pyrazines were dominant in all strain types, while the composition of other volatile compounds showed distinct differences: Aldehydes, ketones, and alcohols were more abundant in strains with roasted/nutty-fatty aroma, douchi aroma, sulfurous notes; Aldehydes, acids, and alcohols were higher in soy sauce/fermented paste aroma strains; Acids and ketones were predominant in fruity-acidic aroma strains. Key differential aroma components were further screened by orthogonal partial least squares-discriminant analysis (OPLS-DA) with a variable importance in projection (VIP) value>1.0, and combined with a relative odor activity value (ROAV)>1.0. Five substances were ultimately identified as crucial for discriminating the aroma profiles of different sensory categories of B. licheniformis: 4-methoxyphenol (a key common component of roasted/nutty-fatty aroma, douchi aroma, and soy sauce/fermented paste aroma strains), isovaleric acid (a key common component of douchi aroma and fruity-acidic aroma strains), 2-octanone (a key component of fruity-acidic aroma strains), 2,3,5-trimethylpyrazine (a key component of roasted/nutty-fatty aroma, douchi aroma, and sulfurous notes strains), and 2,3,5,6-tetramethylpyrazine (a key component of soy sauce/fermented paste aroma strains).

high-temperature Daqu  /  Bacillus licheniformis  /  flavor  /  HS-SPME-GC-MS  /  OPLS-DA
王嘉丽, 何卓, 王松涛, 明红梅, 杨阳, 牛曼思, 赵秋伟, 张正杰, 李姝, 敖宗华. 高温大曲地衣芽孢杆菌的分离鉴定及其产香特性解析. 食品工业科技, 2026 , 47 (18) : 14 -23 . DOI: 10.13386/j.issn1002-0306.2025110022
Jiali WANG, Zhuo HE, Songtao WANG, Hongmei MING, Yang YANG, Mansi NIU, Qiuwei ZHAO, Zhengjie ZHANG, Shu LI, Zonghua AO. Isolation and Identification of Bacillus licheniformis in High-temperature Daqu and Analysis of Its Aroma-producing Characteristics[J]. Science and Technology of Food Industry, 2026 , 47 (18) : 14 -23 . DOI: 10.13386/j.issn1002-0306.2025110022
高温大曲(亦称酱香型大曲)以小麦为原料,经粉碎、调湿、成型、高温发酵、熟化等工序制成,是酿造酱香型白酒的重要糖化发酵剂[1]。在酱香型白酒酿造中,高温大曲与高粱的比例约为1.1:1,这一高比例的大曲使用量被认为是塑造酱香型白酒独特风味的重要因素[2]。尽管目前酱香主体香成分的确认仍存在争议,但普遍认为其香型的形成与高温大曲中微生物的代谢密切相关[3−4]。
研究表明,芽孢杆菌属(Bacillus)在高温大曲的微生态结构中占据主导地位[5]。其中,地衣芽孢杆菌(Bacillus licheniformis)作为一种革兰氏阳性兼性厌氧菌[6],其生态优势主要源于其卓越的耐热特性与广泛的代谢能力。已有研究指出,该菌在固态发酵过程中能够产生超过70种代谢产物,包括芳香族化合物、C4类物质、吡嗪类以及有机酸等。此外,B. licheniformis还可合成多种酶类,如蛋白酶、脂肪酶、热稳定性α-淀粉酶、纤维素酶和半纤维素酶,这些酶协同促进原料中高分子物质的降解及褐变产物的形成,因此其被视为形成酱香风味的关键功能菌属之一[7−8]。
黄晓宁等[9]从不同香型大曲中筛选出一株具有优秀的产淀粉酶和蛋白酶能力的B. licheniformis,实际应用到白酒生产中,检测到酒醅中挥发性风味物质的含量显著提高。张颖等[10]从酱香大曲中筛选出的具有较强产香能力的B. licheniformis,其能够代谢愈创木酚、吡嗪类、呋喃类等风味物质。TANG等[11]从太空大曲中筛选出突变的B. licheniformis,将其应用于强化大曲中,增加了吡嗪、2,3-丁二醇等特征风味的含量。以上研究表明B. licheniformis是一种功能多样且应用潜力较大的菌种。然而,目前行业针对高温大曲Bacillus中各成员的风味成分及其感官特征研究仍相对有限,特别是关于B. licheniformis的关键风味物质及其对酒体风味形成的影响机制,尚缺乏系统深入的探讨。
本研究通过平板划线法分离纯化高温大曲中的Bacillus菌株,通过形态学观察、16S rDNA序列分析对菌株进行鉴定。重点对其中的B. licheniformis菌株进行程序升温模拟发酵,结合人工感官评分与HS-SPME-GC-MS技术对其香气特征进行测定分析,并建立高温大曲来源B. licheniformis判别模型,旨在为B. licheniformis的发酵机制以及代谢差异提供参考依据。
高温大曲 采集自泸州市江阳区某酒厂;所有大曲样品都经过了完整的发酵和储存过程。从大曲堆的顶部、中部和底部随机选择六块大曲,随后将曲块粉碎并充分混合。重复取样三次(每个样品500 g)。大曲粉被转移至无菌包装袋中,立即运输至实验室,4 ℃保存,并立即开展可培养菌株筛选实验;胰蛋白胨大豆琼脂培养基(TSA):胰蛋白胨15.0 g,氯化钠5.0 g,大豆胨5.0 g,琼脂15 g,加水至1000 mL,pH调至7.3±0.2,于121 ℃湿热灭菌15 min;酵母-麦芽琼脂培养基(ISP2):酵母浸粉4.0 g,麦芽浸粉10.0 g,葡萄糖4.0 g,琼脂20.0 g,pH调至7.2±0.2,于121 ℃湿热灭菌15 min;营养琼脂培养基(NA):蛋白胨10.0 g,牛肉浸粉3.0 g,氯化钠5.0 g,琼脂15.0 g,pH调至7.3±0.2,于121 ℃湿热灭菌15 min;改良高氏2号(G2):胰蛋白胨10.0 g,牛心浸粉17.5 g,氯化钠5.0 g,葡萄糖2.0 g,磷酸氢二钠2.5 g,琼脂15.0 g,pH调至7.4±0.2,于121 ℃湿热灭菌15 min;胰蛋白胨大豆肉汤培养基(TSB):胰蛋白胨17.0 g,大豆蛋白胨3.0 g,氯化钠5.0 g,磷酸氢二钾2.5 g,葡萄糖2.5 g,pH调至7.3±0.2,于121 ℃湿热灭菌15 min;小麦固态发酵培养基:取粉碎后的小麦(过20目筛),按料水比1:0.8加水混合均匀,润湿2 h,分装于150 mL组培瓶中,每瓶装料40 g湿润小麦碎,121 ℃灭菌30 min,冷却备用;氯化钠 分析纯,国药集团化学试剂有限公司;2-辛醇 色谱级,上海麦克林生化科技股份有限公司;细菌基因组DNA提取试剂盒 南京诺唯赞生物科技股份有限公司。
QP2020NXnci SYSTEM气相色谱-质谱联用仪 日本岛津有限公司;SAAB57328U 50/30 μm DVB/CAR/PDMS萃取头 广州智达实验室科技有限公司;HP-INNOWAX(60 m×0.25 mm×0.25 μm)毛细管色谱柱 美国安捷伦公司;IFA-240-8强制对流型培养箱 新加坡艺思高科技有限公司;HCB-1300V超净工作台 青岛海尔生物医疗股份有限公司。
分离与纯化:在洁净环境下称取10 g大曲溶解于90 mL无菌带玻璃球的生理盐水中,28 ℃、180 r/min振荡30 min,再转入85 ℃恒温水浴锅中恒温10 min,边加热边振荡。按10倍系列梯度稀释至10−6,吸取10−2~10−6梯度稀释液100 μL均匀涂布于不同碳氮源的固体培养基上(TSA、ISP2、NA、G2),在不同温度(37、45、50、55 ℃)下的生化培养箱倒置培养36~48 h。重复三次实验后,整体挑取形态特征明显的单菌落划线于固体培养基中,反复划线纯化至菌落一致。
保藏:挑取单菌落于TSB培养基中,37 ℃、180 r/min条件下培养12 h,并与50%甘油等体积混合,保存于−20 ℃冰箱,备用。
形态鉴定:依据不同的形态、生理生化特征,参照《伯杰细菌鉴定手册》进行鉴定。
分子鉴定:利用细菌基因组DNA提取试剂盒提取菌株DNA,采用细菌通用引物(27F:5'-AGAGTTTGATCCTGGCTCAG-3'和1492R:5'-TACGGCTACCTTGTTACGACTT-3')进行PCR扩增。扩增体系(20 µL):DNA模板1.5 µL,引物上下游各1 µL,2×Taq Master Mix (南京诺唯赞生物科技股份有限公司)10 µL,无菌ddH2O 6.5 µL。反应条件:95 ℃预变性4 min;94 ℃变性20 s,55 ℃退火20 s,72 ℃延伸1 min 30 s(35次循环);72 ℃延伸10 min。将扩增产物送至上海生工生物有限公司检测,将最终检测的16S rRNA基因序列与NCBI数据库中的序列进行比对。
将保藏的B. licheniformis菌株经TSB液体培养基两级活化(1 mL→5 mL)制备种子液。随后,以10%(v/w)的接种量将种子液接入小麦固态发酵培养基,并添加20%(v/w)无菌水调节湿度,混匀密封后,置于强制对流箱中模拟高温大曲的升温流程培养7 d(图1)。设置接种无菌TSB培养基的空白对照。发酵产物最终通过感官评定与挥发性组分分析进行评估。
根据文献[12−13]提出的研究思路,并借鉴酱香型大曲感官风味评价体系的构建方法及相关酱香功能菌风味鉴定的研究成果[14−15],本研究建立了系统的闻香评价参照体系。该体系以酱香型白酒标样为主要参照,同时涵盖酱香型酒醅、酱香型大曲、商品酱油、商品豆豉、坚果、太妃糖以及多种果蔬作为嗅觉参比实物样,以未接菌的碎小麦作为空白对照。基于上述参照体系,绘制了如图2所示的产香功能菌闻香风味旭日图,作为菌种发酵产物感官评定中的风味标度,用以有效区分不同产香微生物的主体香气特征。每种感官香气模块以5(±0.5)分计,0分为未闻到,5分为闻到的气味最强,按照排名前10的风味标度,并取平均值作风味雷达图。
按照国标GB/T 13868-2009《感官分析建立感官分析实验室的一般导则》来准备闻香环境,根据文献[16]建立品评小组,共10人(5男5女,22~35岁),均经过系统的白酒品评专业培训且具有2年以上白酒品评经验,其中5名均为省级白酒评委,具备国家品酒师资格。
参照文献[17],使用HS-SPME-GC-MS技术测定B. licheniformis发酵醅。
样品前处理:取0.5 g样品加入顶空瓶中,加入2 g氯化钠,5 mL纯水,10 μL 2-辛醇(0.264 μg/μL)为内标物,将制备好的样品于55 ℃保温平衡5 min后,用50/30 μm DVB/CAR/PDMS萃取头在55 ℃萃取45 min,萃取结束后在GC进样口250 ℃解吸5 min。
GC条件:HP-INNOWAX色谱柱(60 m×0.25 mm×0.25 μm);升温程序:起始温度40 ℃,保持1 min,以2.5 ℃/min升到75 ℃,以3 ℃/min升至162 ℃,以6 ℃/min升至230 ℃,保持5 min,载气为高纯氦气(1.0 mL/min);进样口温度250 ℃,不分流。
MS条件:电子电离(Electronic Ionization,EI)源,电子能量70 eV;电子倍增器电压350 V;离子源温度230 ℃;传输线温度250 ℃;扫描范围m/z 40.00~450.00。
定性及半定量分析:以保留时间及保留指数为定性依据,通过美国国家标准技术研究所NIST(National Institute of Standards and Technology)20谱库检索,筛选并去除相似度<80%的峰和硅氧烷类杂质。利用各化合物峰面积与内标物质峰面积之比,计算出各挥发性风味物质含量,公式按式(1)计算。
$ \begin{split} & 待测香气物质含量(\text{μg/g})= \\ &\frac{待测香气物质峰面积\times 内标物含量(\text{μg/g})}{内标物峰面积}\end{split} $
气味活度值(Odor Activity Value,OAV)与相对气味活度值(Relative Odor Activity Value,ROAV)的计算参照文献[18]所述方法。OAV定义为香气成分在样品中的含量与其嗅觉阈值的比值,用于评估各组分对发酵醅整体香气的贡献程度。通常认为,当OAV>1时,该成分对发酵醅呈香具有重要贡献;当OAV>10时,其对香气的贡献尤为显著。为进一步量化不同挥发性成分对发酵醅整体香气的相对贡献程度,引入ROAV进行分析。一般认为,ROAV≥1.00的组分为关键呈香物质,其值越大,香气贡献越大;0.10≤ROAV<1.00的组分则对整体香气具有修饰作用。ROAV按公式(2)计算:
$ \mathrm{ROAV}=\frac{\text{OAV}}{对发酵醅挥发性成分贡献最大的\text{OAV}}\times 100 $
差异香气成分筛选:通过正交偏最小二乘判别分析(Orthogonal Partial Least Squares-Discriminant Analysis,OPLS-DA)进行筛选。基于该模型计算各挥发性成分的变量投影重要性(Variable Importance in Projection,VIP)值,以VIP>1.0作为筛选标准,识别出发酵醅中的差异香气成分。
使用GraphPad Prism 10.1.2和Origin 2025软件制作折线图、堆叠柱状图、雷达图、upset图、旭日图;MEGA11软件构建系统进化树;对每个挥发性化合物分别采用Z-score方法标准化,ChiPlot(https://www.chiplot.online/)绘制了相关热图;SIMCA 14.1软件进行主成分分析法(Principal Component Analysis,PCA)和正交偏最小二乘判别分析(OPLS-DA),计算预测变量重要性投影(VIP)。
本研究采用热处理法,在不同分离培养基及温度条件下,从高温大曲中分离并筛选得到74株Bacillus菌株。经形态学初步鉴定,有16株为潜在的B. licheniformis,占Bacillus菌株总数的21.6%。将16株菌株接种于TSA培养基上,于37 ℃条件下培养,并参照《伯杰细菌鉴定手册》对其菌落形态进行系统观察与分析。结果如图3、表1所示,通过观察菌落形态可以发现虽然这些菌株可能为同种,但不同菌株的菌落形态仍表现出一定差异。如TSA-37-28以及NA-55-51均为表面整体粗糙、有褶皱,ISP2-55-41整体隆起且光滑,与其他菌株差异较大,这说明在B. licheniformis种内仍存在较大多样性,有必要进一步开展不同菌株间发酵特性的比较分析,以揭示其功能差异。
采用MEGA软件,基于邻接法构建系统进化树,并以贝莱斯芽孢杆菌(Bacillus velezensis)Yb-1的16S rRNA基因序列作为外群。系统发育分析结果(图4)表明,所有测试菌株均与B. licheniformis的不同模式菌株聚为高支持率的进化支。具体而言,菌株TSA-37-28、TSA-37-48、TSA-50-47和TSA-50-26与模式菌株B. licheniformis CICC 10087的16S rRNA基因序列相似性最高,分别为99.93%、100.00%、100.00%和100.00%;TSA-50-27和G2-55-43与模式菌株B. licheniformis CPO C11的相似性最高,分别为99.93%和99.79%;NA-55-51与模式菌株B. licheniformis HBUAS67301的相似性为99.92%;其余菌株(TSA-37-3、ISP2-55-33至ISP2-55-43)与模式菌株B. licheniformis GZ-1的序列相似性均高于99.50%。基于上述结果(所有菌株与相应最近模式菌株的16S rRNA基因序列相似性均不低于99.50%),本研究将全部16株菌鉴定为B. licheniformis。
在长时间的高温阶段中,嗜热菌属能够分泌多种耐热酶,这些酶有助于促进美拉德反应的进行,并与酱香型风味物质的生成密切相关。美拉德反应对高温大曲最终风味的形成具有决定性作用[19]。对前期筛选与分子鉴定所得的16株B. licheniformis分别进行小麦固态发酵,在7 d程序升温培养后,对其进行感官评定。结果如图5所示,各菌株发酵产物的风味特征具有明显差异,具体描述如下:菌株TSA-50-26与TSA-50-27(图5A)以坚果油脂香和烘焙香为主导,伴随轻微的焦糊香与酱油香;菌株NA-55-51与TSA-37-48(图5B)主体呈豆豉香,并带有烘焙香、生麦香及黄瓜香气;菌株TSA-37-28与TSA-50-47(图5C)以硫香为主,辅以淡弱豆豉香与轻微坚果油脂及烘焙香气;菌株ISP2-55-41与G2-55-43(图5D)呈现典型的酱香与酱油香,风格偏向焦甜香,同时具备豆豉香与烘烤香,并略带氨味;其余菌株(图5E、图5F)整体表现为果酸香,具有明显发酵酸类气息,并伴随水果香、生麦味及青椒、青草等气息。综合感官评分雷达图与描述语分析,将上述16株B. licheniformis固态发酵产物的主体风味归纳为5大类。菌株编号及其对应风味类型归纳如表2所示。
为解析高温大曲中B. licheniformis的产香特性,本研究采用HS-SPME-GC-MS技术对16株B. licheniformis在单菌固态发酵体系中所产生的挥发性成分进行定性分析。结果表明,B. licheniformis发酵醅中共鉴定出198种挥发性化合物,涵盖酮类(47种)、醇类(35种)、吡嗪类(21种)、酯类(20种)、酸类(19种)、醛类(13种)、酚类(10种)及其他类别(33种),并根据发酵醅的主体风味特征,将菌株划分为不同风味类别(图6);在风味物质种类方面(图6A),呈现烘焙香-坚果油脂香、豆豉香、硫味以及酱香-酱油香风味的菌株及其空白小麦培养基中,共鉴定出77~94种风味物质;而具有果酸味特征的菌株所产风味物质种类相对较少,仅鉴定出42~57种;在风味物质含量方面(图6B),空白组中挥发性风味物质总含量显著低于B. licheniformis发酵组,空白培养基的风味组成以醇类、酯类和醛类为主,辅以少量酸类与酮类化合物;相比之下,B. licheniformis发酵产物中则以吡嗪类、醛类为主,同时检测到含量较高的酚类和醇类,以及较低含量的酸类、酮类及酯类等组分。已有研究指出,酚类、醛类与吡嗪类化合物是构成酱香型白酒典型风味结构的关键组分[20−21],由此可见,B. licheniformis可能是这几类关键香气物质在高温大曲发酵过程中的潜在重要微生物来源。
进一步分析不同菌株风味含量情况(图6B),呈现烘焙香-坚果油脂香的菌株TSA-50-26风味物质总含量最高,达67.29±2.23 μg/g;而呈现果酸香的菌株ISP2-55-39总含量最低,仅为14.15±1.86 μg/g。在不同风味类别的菌株中,各类挥发性物质的含量存在显著差异。在烘焙香-坚果油脂香类别中,菌株TSA-50-26的醇类物质含量最高(17.24±0.24 μg/g),而菌株TSA-50-27则在酯类(0.78±0.01 μg/g)和醛类(16.63±0.13 μg/g)物质含量上最高。在豆豉香类别中,菌株NA-55-51的吡嗪类物质含量最高(32.42±2.07 μg/g),菌株TSA-37-48则在酮类(5.30±0.09 μg/g)和酚类(9.03±0.07 μg/g)物质含量上表现最优。本研究结果表明,B. licheniformis能够产生多样且具特征性的气味谱,其中具有烘焙香-坚果油脂香、豆豉香及酱香-酱油香特征的菌株,在白酒大曲的风味定向强化方面展现出良好的应用前景。
基于前期分析,进一步采用Upset图(图6C)展示了不同感官评定发酵醅中特有挥发性化合物的种类组成。结果显示,果酸香型发酵醅中特有化合物种类最多,共鉴定出17种,涵盖7种酸类、2种吡嗪类、2种醇类、2种酮类、1种酯类及3种其他类化合物,代表性成分包括糠醇、环己烷羧酸、苯并噻唑和7-甲基辛酸等。烘焙香-坚果油脂香组中检出11种特有成分,主要包括3种酯类、2种醇类、1种酸类、1种酮类及4种其他类化合物,如己酸、2-苯基-1-丙醇和2-十三烷酮等。豆豉香组中共鉴定出9种特有化合物,分别为2种吡嗪类、2种酮类、1种酚类及4种其他类,代表性物质包括2-甲基-6-乙烯基吡嗪、愈创木酚和2-哌啶酮等。硫味组中检出5种特有成分,包括2种酮类、1种醇类及2种其他类,如异丙基苯基酮和3,5-辛二烯-2-酮等。酱香-酱油香组中仅检出1种特有挥发性物质,即乙偶姻。
为明确高温大曲中B. licheniformis的特征香气组分,本研究对16株菌株产生的香气化合物及其相对含量进行了聚类分析。从中鉴定出44种共有香气成分(定义为在不少于8株菌中均检出的物质[22]),包括10种酸类、8种醇类、6种吡嗪类、6种酮类、5种酚类、4种醛类、1种酯类以及4种其他类型化合物。以这44个共有香气组分为因变量,不同感官评定结果为自变量,采用主成分分析(PCA)和正交偏最小二乘判别分析(OPLS-DA)进行分析。
PCA结果显示(图7A),前两个主成分累计贡献率为80.8%,可以看出能够很好区分酱香-酱油香菌株以及果酸香菌株,其他组别区分效果不明显。进一步进行OPLS-DA统计分析(图7B),可以实现5类感官评定的B. licheniformis有效区分,本次分析中的自变量拟合指数(R2x)为0.975,因变量拟合指数(R2y)为0.915,模型预测指数(Q2)为0.69,R2和Q2超过0.5表示模型拟合结果可接受[23]。经过200次置换检验,如图7C所示,Q2回归线与纵轴的相交点小于0,说明模型不存在过拟合,模型验证有效,认为该结果可用于B. licheniformis发酵香气的感官表现鉴别分析。
为了进一步分析不同香气成分对区分5类感官评定的B. licheniformis发酵醅的贡献率,基于有效判别模型,采用变量重要性投影(VIP)来评定各组样本分类判别的影响,通常以VIP>1.0为条件筛选具有显著差异的香气物质[24]。如图8所示,本研究共筛选出11种差异香气物质,其中吡嗪类3种、醇类3种、酮类2种、醛类1种、酸类1种、酚类1种。与空白对照组相比,各产香菌株均具有其独特的特征性风味化合物。
单个香气成分含量的高低并不能作为判定酱酒香气特征的依据,通常赋予酱酒香气特征的是具有较高OAV的香气成分[25]。同时ROAV可用于综合评价挥发性成分对基酒香味的贡献程度[26]。结果如表3所示,4-甲氧基苯酚在物质浓度最高菌株和最低菌株中OAV最高,为对发酵醅挥发性成分贡献最大的OAV,进一步将其用于计算ROAV以量化比较各挥发性成分对发酵醅整体呈香的贡献度。
综合图8与表3数据可知,在物质浓度最低与最高的菌株中,共有5种化合物的ROAV值均大于1。其中,4-甲氧基苯酚(ROAV=100~100)为香气贡献最突出的挥发性成分,能够有效区分烘焙香-坚果油脂香、豆豉香及酱香-酱油香菌株与其他类型;文献报道该化合物在某些白酒中可能呈现类似动物毛皮的气味[27]。异戊酸(ROAV=17.31~27.87)在豆豉香与果酸香菌株中含量较高,呈现酸味与湿奶酪样气息,已有研究确认其为豆豉的关键香气成分之一[28]。仲辛酮(2-辛酮,ROAV=1.64~2.75)在果酸香菌株中含量偏高(1.21~1.92 μg/g),可作为该类菌株的鉴别指标,其香气特征表现为花香与草香[29]。2,3,5-三甲基吡嗪(ROAV=1.68~1.87)能够区分烘焙香-坚果油脂香、豆豉香及硫味菌株,文献中其风味通常与坚果、青草及可可等相关[30]。2,3,5,6-四甲基吡嗪(ROAV=1.02~1.32)为酱香-酱油香菌株的特征性化合物,在该类菌株中含量最高(16.36~16.94 μg/g),且在酱香型白酒中占比突出,是极为重要的风味物质[31]。表明这5种差异香气物质可能对判别不同感官判定B. licheniformis的香气特征具有重要作用。
本研究成功从高温大曲中分离并鉴定出16株B. licheniformis,通过人工感官评定将其固态发酵产物划分为五类主体风味,包括烘焙香-坚果油脂香、豆豉香、硫味、酱香-酱油香及果酸香。采用HS-SPME-GC-MS技术共鉴定出198种挥发性化合物,B. licheniformis组风味总含量显著高于空白对照组,并以吡嗪类、醛类为主,同时检测到含量较高的酚类和醇类,以及较低含量的酸类、酮类及酯类等组分。PCA分析与OPLS-DA分析有效区分了不同感官评定的菌株,模型拟合良好且预测能力可靠。通过变量重要性投影(VIP>1.0)筛选出11种关键差异香气成分,其中5种成分(包括4-甲氧基苯酚、异戊酸、仲辛酮、2,3,5-三甲基吡嗪、2,3,5,6-四甲基吡嗪)在各类菌株中相对气味活度值(ROAV)均大于1,表明其对B. licheniformis固态发酵的总体风味具有重要贡献。
本研究首先系统解析了酱香型高温大曲中B. licheniformis的产香特性与代谢多样性,并建立了不同菌株代谢谱与其感官属性之间的关联。其次,研究发现在相同酿造环境下,同一物种的不同菌株会表现出截然不同的感官特性,这直接构成了白酒风味多样性的微生物学基础。鉴于此,本研究建立的判别模型与关键香气指标体系,为后续开展风味导向的优良菌种选育及强化大曲开发提供了坚实的理论依据与数据支持。未来研究可聚焦于关键菌株的功能基因挖掘及代谢调控网络解析。

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2026年第47卷第18期
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doi: 10.13386/j.issn1002-0306.2025110022
  • 接收时间:2025-11-05
  • 首发时间:2026-10-24
  • 出版时间:2026-09-15
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  • 收稿日期:2025-11-05
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    1四川轻化工大学食品与酿酒工程学院,四川宜宾 644005
    2泸州老窖股份有限公司,国家固态酿造工程技术研究中心,四川泸州 646000
    3中国科学院微生物研究所,微生物资源前期开发国家重点实验室,微生物生理与代谢工程研究室,北京 100101

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李姝(1991−),女,博士,工程师,研究方向:白酒生产技术、酿造微生物、天然产物与功效评价,E-mail:。
敖宗华(1971−),男,博士,教授级高级工程师,研究方向:发酵产品风味物质研究和功能因子分析,E-mail:。
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2种不同金属材料的力学参数
科
Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)
属
Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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