【作者】 孙宗保；

【导师】 赵杰文；

【作者基本信息】 江苏大学 ， 农产品加工及贮藏工程， 2014， 博士

【摘要】 镇江香醋是我国典型的传统发酵食品,以“色、香、酸、醇、浓”的特点,享誉国内外。香是镇江香醋最重要的的感官品质之一,是醋中多种挥发性成分共同作用的结果,但目前对镇江香醋挥发性成分的研究还有待进一步深入,如它们在生产和陈酿过程中有何变化规律,哪些具有香气活性,它们与香醋质量或醋龄的关系等。本研究以固相微萃取(SPME)、气相色谱-质谱联用(GC-MS)技术和气相色谱嗅觉测量技术(GC-O)从生产过程到陈酿过程,从整体挥发性成分到对香醋香气起直接贡献的香气活性成分,全面解析镇江香醋香气的产生和演变以及醋龄对其的影响。此外,目前市场上镇江香醋醋龄标注混乱,亟待需要科学准确的方法进行规范和识别。本研究利用不同醋龄镇江香醋中的挥发性成分、香气活性成分及其香气活性值(OAV)以及电子感官分析技术(电子鼻和电子舌技术及其融合技术)对不同醋龄的镇江香醋进行分析,建立相应的醋龄识别方法,以实现镇江香醋醋龄的准确区分和快速判别。本论文的主要研究内容如下：1.镇江香醋挥发性成分生产过程中的变化研究。通过SPME-GC-MS分析了镇江香醋各生产阶段(含辅料)的挥发性成分。共检测到108种挥发性成分,酯类化合物种类最多,其次是杂环化合物。由于固态醋醅的环境极利于产酯酵母的增殖和发酵,大量酯类物质在醋酸发酵中期的醋醅中含量达到最大值；除糠醛外,镇江香醋生产阶段产生的杂环化合物大都含量较低,主要来自原料的蒸煮、炒米色和煎醋阶段。乙酸、3-甲基丁酸、2-甲基丁酸和己酸以及乙醇、苯乙醇、3-甲基丁醇和2-甲基丁醇是镇江香醋生产过程中主要的酸和醇类物质,分别主要产生于醋酸发酵和酒精发酵阶段。短链脂肪醛主要产生于原料的蒸煮(液化和糖化),镇江香醋糖化以及炒米色和煎醋均需加热处理,有利苯甲醛和苯乙醛两种芳香醛的生成。2,3-丁二酮和3-羟基-2-丁酮是镇江香醋生产过程中产生的两种最主要的酮类物质,是四甲基吡嗪合成的前体物质。醋酸发酵阶段尤其是醋酸发酵后期是3-羟基-2-丁酮生成的主要阶段,固态醋酸发酵有利于3-羟基-2-丁酮的生成。2.不同醋龄镇江香醋挥发性成分的变化研究。采用SPME-GC-MS分析了新醋以及醋龄为6、12、18、48和78个月的6种醋龄镇江香醋中挥发性成分含量的变化。①88种挥发性成分被确定,杂环类化合物和酯类化合物种类最多,分别达到27种和18种。随着醋龄的增加,镇江香醋挥发性成分个数呈现不断增加的趋势。陈酿对镇江香醋挥发性成分中的酸类化合物总体影响较少,大多数醇类化合物含量在新醋中明显高于经过陈酿的镇江香醋,陈酿有利于乙酸乙酯等多种酯类物质含量的增加。苯乙醇、苯甲醛、3-羟基-2-丁酮、2,3-丁二酮、乙酸苯乙酯和乙酸乙酯分别是镇江香醋中含量最高的醇、醛、酮和酯类物质。②镇江香醋中2-甲基丁醇/3-甲基丁醇含量比值具有较好的稳定性,可以作为镇江香醋真伪鉴别的参考。③杂环化合物在镇江香醋陈酿过程中变化最为显著,随醋龄的增加杂环化合物个数明显增加,这种变化主要由吡嗪和恶唑类化合物引起。杂环化合物中吡嗪种类最多,大都随醋龄增加含量明显增加。2-乙基-4,5-二甲基恶唑等四种恶唑类化合物在镇江香醋中首次发现,恶唑类化合物随镇江香醋醋龄增加含量显著增加。杂环化合物种类丰富、含量较高是形成镇江香醋特别是醋龄较长的镇江香醋特有风味的重要原因。④苯甲醛、三甲基吡嗪、2,6-二乙基吡嗪、四甲基吡嗪、三甲基恶唑能较好的表征镇江香醋醋龄的变化,它们可以作为镇江香醋醋龄变化的重要标志物。3.不同醋龄镇江香醋香气活性成分的研究。①利用DB-WAX和DB-5MS两种不同极性色谱柱通过香气提取稀释分析法(AEDA)对新醋、醋龄12、48和78个月四种醋龄镇江香醋香气活性成分进行分析。首次发现了两种香气活性成分,其中一种为3-甲硫基丙醛,虽含量极少(低于GC-MS仪器检测限),但对镇江香醋特有香气具有极其重要的贡献(香气稀释因子(FD因子)≥2048)；另一种为2-乙基-4,5-二甲基恶唑,镇江香醋醋龄对其有极其显著的影响,随醋龄增加其FD因子有明显的增加。②将AEDA和直接强度法两种嗅闻分析方法结合分析了四种不同醋龄镇江香醋的香气活性成分,发现香气活性成分中3-甲基丁醛、2,3-丁二酮、3-甲基丁酸在四种醋龄中香气强度≥3,同时FD≥256,它们是镇江香醋香气最基本的组成成分。③基于GC-O的嗅闻结果表明,新醋和经过陈酿的镇江香醋香气轮廓差异非常明显,杂环化合物(大都具有杏仁味和药味,且气味强烈)和酯类(大都具有果香和甜味,气味较柔和)的差异尤其是杂环化合物的差异是不同醋龄镇江香醋特征香气差异最主要的原因之一。4.基于挥发性成分和香气活性成分的不同醋龄镇江香醋的识别研究。①对不同醋龄镇江香醋中挥发性成分含量和香气活性成分OAV值的数据进行主成分分析(PCA)和聚类分析(CA),结果表明新醋和醋龄较长的镇江香醋区分明显,对陈酿时间较短醋龄较接近的镇江香醋区分不十分理想。此外,比较两种方法的PCA和CA结果可以看出,镇江香醋短时陈酿的过程中,挥发性成分的变化主要是一些对其特征香气贡献较小的组分。②基于挥发性成分和基于香气活性成分的线性判别分析(LDA)均能有效将6种不同醋龄镇江香醋区分开,其训练集和测试集总体识别率均达到100%。但基于香气活性成分的LDA能更清晰地反映不同醋龄镇江香醋之间的变化,更准确的表征不同醋龄镇江香醋之间的内在联系。5.基于电子感官分析技术的不同醋龄镇江香醋的识别研究。利用电子鼻和电子舌及两者的融合技术对6种不同醋龄镇江香醋进行识别。①在基于电子鼻的醋龄识别研究中,分别用LDA、支持向量机(SVM)和反向传播神经网络(BPNN)三种模式识别方法进行醋龄识别建模,结果表明,BPNN要略优于LDA和SVM方法,其训练集和测试集识别正确率分别为98.75%和97.5%。②基于电子舌的不同醋龄镇江香醋LDA识别模型和基于电子鼻和电子舌信息融合后的LDA识别模型均能有效区分6种不同醋龄的镇江香醋,其训练集和测试集的正确率均达到100%,但电子鼻和电子舌信息融合后的LDA识别模型中相同醋龄样本的集中度显著提高,不同醋龄镇江香醋之间界限更为明显。更多还原

【Abstract】 Zhenjiang aromatic vinegar is Chinese typical fermentative food. It is famous for its color, aroma, acid, mellowness and denseness. Aroma is one of the most important sensory qualities of Zhenjiang aromatic vinegar, which is the result of kinds of volatile components in vinegar. But the present research on the he volatile compents of Zhenjiang aroimatic vinegar remains to be further in-depth,such as how do they change in the production and aging process,which have the odour activity,and what they have relations to the quality of Zhenjiang aromatic vinegar and vinegar ages and so on.This study had an comprehensive analysis on the generation and evolution of aroma of Zhenjiang aromatic vinegar and the influence of vinegar ages to the aroma, using Solid phase micro extraction (SPME), Gas chromatography-mass spectrometry (GC-MS) and GC-Olfactometry(GC-O),from production process to aging process,from the overall volatile compounds to the odour active compounds which has direct effect on the aroma. In addition, the ages of vinegar were marked confusedly currently on the market, so, we urgently need scientific ways to correct and recognize the ages of vinegar. We used volatile components of different ages of vinegar, odour active compounds and their odour activity value (OAV) and electronic sensory technology(electronic nose,electronic tongue and the fusion of two techniques) to tell differences from different ages of vinegar and to build up some corresponding methods to identify vinegar age.So wo can discriminate the ages of Zhenjiang aromatic vinegar correctly and rapidly.The main contents of this paper were as follows:1. Study on the volatile components in Zhenjiang vinegar changes in the process of production. By using SPME-GC-MS, we analyzed volatile compositions of Zhenjiang aromatic vinegar during each production stage (including accessories). Totally108kinds of volatile components were detected, in which, esters were most, and followed by heterocyclic compounds. Because the environment of solid-state vinegar is very conducive to the proliferation and fermentation of yeasts, the number of esters reached the maximum in the fermentation medium term of the acetic acid fermentation; Besides furfural, the contents of most heterocyclic compounds were very low in production stage of Zhenjiang vinegar, mainly from the cooking of raw material,"Chaomi" color and the fried phase of vinegar. Acetic acid,3-methyl butanoic acid,2-methyl butanoic acid and hexanoic acid and ethanol, Phenylethanol,3-methyl-1-butanol and2-methyl-1-butanol were the main acid and alcohol compounds in the production of Zhenjiang aromatic vinegar, respectively mainly produced in the alcohol fermentation and the acetic acid fermentation stage. Short chain fatty aldehyde was mainly produced in the raw material of cooking (liquefaction and saccharification). Zhenjiang vinegar’s saccharification and "Chaomi" color and the fried phase of vinegar all requried the heat treatment, which is good for the generation of two kinds of aromatic aldehyde namely benzaldehyde and phenylacetaldehyde..2,3-butanedione and3-hydroxy-2-butanone were the main two kinds of material of ketones in the production process of Zhenjiang vinegar, and the precursor compounds from the synthesis of tetramethyl pyrazine. Acetic acid fermentation stage especially the late stage were the main stage of generation of3-hydroxy-2-butanone. The solid-state acetic acid fermentation avails the production of3-hydroxy-2-butanone.2. Study on the changes of volatile composition of different ages of Zhenjiang vinegar.By using SPME-GC-MS,we analyzed the changes of contents of volatile components of Zhenjiang aromatic vinegar in6kinds of different vinegar ages of the new vinegar and the age of6,12,18,48and78months.①88kinds of volatile components were identified and heterocyclic compounds and ester compounds were most which respectively reached27and18kinds.With the increase of ages of vinegar, the number of volatile component of Zhenjiang vinegar increased. Overall aging of Zhenjiang aromatic vinegar had less influences on the acid compounds of volatile components, most of the alcohol content were significantly higher in the new vinegar than in the aging of Zhenjiang aromatic vinegar, Aging is conducive to the increase of content of ethyl acetate and some other ester. Phenylethanol, benzaldehyde,3-hydroxy-2-butanone,2,3-butanedione, phenyl ethyl acetate and ethyl acetate were the highest content of alcohols,aldehydes,ketones and esters of Zhenjiang aromatic vinegar respectively.②2-methyl-l-butanol/3-methyl-1-butanol contents’ratio in Zhenjiang vinegar had good stability, which could as a reference to identify the authenticity of Zhenjiang vinegar’s.③The change of heterocyclic compounds was the most significant in the aging process of Zhenjiang aromatic vinegar. With the increase of age of Zhenjiang aromatic vinegar, the number of heterocyclic compounds increased significantly. The change was mainly caused by pyrazine and oxazole compounds. Pyrazines are the most substance in heterocyclic compounds, most of their contents increased obviously with the increase of age of vinegar.2-ethyl-4,5-dimethyl oxazole and other three kinds of oxazole compounds were firstly discovered in Zhenjiang vinegar in this work. The content of oxazole compounds increased significantly with age. The various varieties and high content of heterocyclic compounds were the important reasons for the characteristic flavor and odor of Zhenjiang vinegar especially the vinegar with larger age.④Benzaldehyde, trimethyl pyrazine,,2,6-diethyl pyrazine, tetramethylpyrazine, trimethyl oxazole can better characterize the change of the age of Zhenjiang aromatic vinegar, which can serve as important marker of the changes of age of Zhenjiang vinegar.3. Study on the odour active compounds of Zhenjiang vinegar with different ages.①The DB-WAX and DB-5MS as two different polarity columns, combined with AEDA, were simultaneously used for the analysis of odour active compounds in Zhenjiang vinegar with four different ages of the new vinegar and the age of12,48and78months.Two odour active compounds were firstly discovered in Zhenjiang vinegar,one was3-(methylthio)-propanal whose content was rare (less than GC-MS instrument detection limit) but extremely important to the unique aroma of Zhenjiang vinegar (aroma dilution factor (FD)>2048).Another was2-ethyl-4,5-dimethyl oxazole, whose FD factor was obviously increased with age of Zhenjiang aromatic vinegar.②AEDA and direct strength method, as two sniffing analysis methods, were used and combined to analyze the odour active compounds in Zhenjiang aromatic vinegar with four different ages. It was found that3-methyl-butanal,2,3-butanedione and3-methyl butanoic acid in the four different ages of vinegar, whose aroma intensity≥3and FD>256, were the basic components in Zhenjiang aromatic vinegar’s aroma.③The results of GC-O showed that differences of the flavor profile between the new vinegar and the aging vinegar were very apparent.The differences of heterocyclic compounds (mostly with almond flavor and taste, and strong smell) and esters with softer (mostly with fruity and sweet, and softer smell), especially the heterocyclic compounds, which were one of the main reasons for the characteristic differences of Zhenjiang aromatic vinegar’s flavors with different ages.4. Study on the recognition of different ages of Zhenjiang aromatic vinegar based on the volatile constituents and odour active compounds.①The contents of volatile components and the OAV of the odour active compounds in Zhenjiang aromatic vinegars with different ages were analyzed by principal component analysis (PCA) and cluster analysis (CA),the results showed that the new vinegar and the long aged ones could be distinguished from other ages obviously, but the result was not very ideal for identifying the vinegars with short aging time and close ages.In addition, compared results of PC A and CA, it can be seen that the change of volatile components is mainly related with some components which make small contributions to the characteristic aroma in the short aging process of vinegar.②According to volatile components and the odour active compounds, LDA on can effectively separate the Zhenjiang aromatic vinegars with6different ages. The recognition rate in the training set and testing set reach was achieved with100%respectively. But the LDA result based on odour active compounds could more clearly reflect the different changes between Zhenjiang vinegar of different ages and show more accurate characterizations of the internal relations between them.5. Study on the recognition of Zhenjiang aromatic vinegar with different ages using the electronic sensory technology. Electronic nose and electronic tongue were combined to analyze the Zhenjiang aromatic vinegar with6different ages.①In the recognition of different ages of Zhenjiang aromatic vinegar using electronic nose technique, LDA, support vector machine (SVM) and back-propagation neural network (BPNN) were used to build up models to recognize vinegar age.The result showed that the performance of BPNN was better than that of LDA and SVM,whose recognition accuracy rate in the training set and test set achieved98.75%和97.5%respectively.②Electronic tongue with LDA model and multi-sensor fusion (electronic nose and electronic tongue) with LDA model could distinguish the6different ages of Zhenjiang vinegar. The identification rate in the training set and testing set of these models all achieved100%; however, samples from the same age gathered closer in multi-sensor fusion (electronic nose and electronic tongue) with LDA model, and the boundaries between different groups were more obvious.更多还原