气相色谱-嗅闻/质谱联用分析酵母菌发酵酱油中香气物质

酱油香气物质的形成与酵母菌的代谢息息相关,本文旨在研究酵母菌对发酵酱油中香气物质形成与含量的影响。采用固相微萃取(SPME)结合气相色谱-嗅闻-质谱(GC-O/MS)联用技术分别对不加酵母菌、加AY酵母菌以及加入T酵母菌发酵制得酱油中的香气物质进行检测分析。结果显示:加入酵母菌的酱油中形成的各类香气物质更加丰富,其中AY型鲁氏酵母菌能够产生更多种类的酯类(11种)及醇类(12种)物质;T型鲁氏酵母菌有助于醛类(7种)物质的形成。且AY型鲁氏酵母菌在酱油酿造过程中产生的酯类(155.19 μg/L)、呋喃酮类(68.53 μg/L)、醛类(86.46 μg/L)等物质的含量更高,T型鲁氏酵母菌的产醇类(1260.64 μg/L)物质能力则更加优越。通过对酱油样品中OAV的计算,发现,AY酵母菌酿造酱油中的4-羟基-2-甲基-5-乙基-3(2H)-呋喃酮(HEMF),2/3-甲基丁醛、及苯乙醛的OAV高于其他样品的OAV值,T型鲁氏酵母菌的1-辛烯-3-醇及3-甲硫基丙醛的OAV值高于其他样品的OAV值。将OAV值与PCA结合分析,可以看出,AY型鲁氏酵母菌发酵酱油的香气优势明显,并且T型鲁氏酵母菌发酵酱油以熟土豆气味为主,并伴有蘑菇香气;AY型鲁氏酵母菌发酵酱油以甜香香气为主,并伴有麦芽香气。综上,AY型鲁氏酵母菌的整体产香优于T型鲁氏酵母菌的产香效果。     

高效液相色谱串联质谱法测定酱油中水解氨基酸

建立了高效液相色谱串联质谱(HPLC-MS/MS)法测定天然酿造酱油原油(FSS)和酸水解植物蛋白调味液(HVP)中多肽的水解氨基酸的组成的方法。FSS多肽总量约是HVP的10倍,其中FSS中多肽的总量为2378.76mg/L,HVP中多肽的总量为247.57mg/L。天然发酵酱油原油中赖氨酸在多肽中的含量21倍高于酸水解植物蛋白液。FSS多肽中赖氨酸、组氨酸、酪氨酸相对百分含量约是HVP的2倍,HVP中丙氨酸、脯氨酸相对百分含量约是FSS的2倍。该研究为鉴别天然酿造酱油与酸水解酱油提供依据。     

酱油渣蛋白质的分离、鉴定和氨基酸组成特征研究

本文以酱油上清液为对照,以过滤-冷冻真空干燥法制备了酱油渣。首先,对比研究了酱油渣的常规理化指标。其次,以SDS-PAGE、MALDI-TOF/TOF MS和HPLC法对酱油渣和酱油上清中蛋白质分子量分布、来源与种类及氨基酸组成特征进行了研究。结果显示酱油渣（湿渣）主要由15.98%的碳水化合物、10.19%的蛋白质、10.10%的食盐和59.50%的水分组成。SDS-PAGE和MALDI-TOF/TOF MS分析结果显示酱油渣蛋白质主要由分子量约为19.3 kDa（a, 59.40%）、31.8 kDa（b, 12.91%）和12.3 kDa（c, 27.69%）的蛋白质亚基组成,经质谱鉴定分子量为19.3 kDa的多肽（a）为Glycinin G4亚基碱性端B3。酱油渣蛋白质氨基酸组成分析表明,与酱油上清中蛋白质相比,酱油渣蛋白质中疏水性氨基酸含量和平均疏水值明显高于酱油上清中蛋白质。上述差异是酱油渣蛋白质氨基酸组成的主要特征,也是酱油渣蛋白质水溶性差和形成的主要原因。     

A Novel Method for the Determination of the Ethanol Content in Soy Sauce by Full Evaporation Headspace Gas Chromatography

A full evaporation headspace gas chromatographic (HSGC) method is described for rapid determination of ethanol in soy sauce. The results demonstrated that a near-complete mass transfer of ethanol in the soy sauce sample to the vapor phase (headspace) was achieved within 2 min at 105 °C when a very small (≤40 μL) sample was added to a 20-mL headspace sample vial. The ethanol in the vapor phase was then determined by HSGC using a flame ionization detector. The results showed that the method has an excellent measurement precision (relative standard deviation <0.41 %) and accuracy (recovery?=?99.85?±?1.85 %) for ethanol quantification in soy sauce samples. The method requires no sample pretreatment, so it is a simple, rapid, and sensitive method for the accurate quantification of ethanol content in soy sauce and suitable for applications in food industry.     

酱油中挥发性风味物质的气相色谱-离子迁移谱分析

采用气相色谱-离子迁移谱（GC-IMS）对不同酱油样品中的风味物质进行检测,根据挥发性成分指纹图谱,利用多元统计法分析散装酱油与品牌酱油的差异和散装酱油与不同等级品牌酱油的差异。结果表明,酱油中主要的挥发性风味成分包括苯乙醛、丙醛、5-甲基-2-呋喃甲醇、2,3-丁二酮、5-甲基糠醛、乙醇、乙酸乙酯等11种化合物。散装酱油和品牌酱油的风味物质存在明显差异,正交偏最小二乘法判别分析（OPLS-DA）结果显示,大部分风味物质在散装酱油中的含量低于品牌酱油,与风味指纹图谱结果基本一致;散装酱油与不同等级的品牌酱油也能区别开。该研究结果表明气相-离子迁移谱技术为酱油风味检测技术的开发和研究提供新思路。     

蒸汽爆破花生壳对花生酱油风味的影响

为促进花生壳再利用,将其经蒸汽爆破后代替部分面粉酿造花生酱油,并采用电子鼻检测及顶空固相微萃取-气相色谱质谱联用技术（HS-SPME-GC-MS）法对5种酱油样品的风味进行分析。电子鼻结果表明,添加花生壳或蒸汽爆破花生壳会影响酱油的整体风味;GC-MS结果表明,不添加花生壳酿造的酱油（HS0）风味物质主要由酯类构成,其相对含量高达85.35%;添加花生壳的酱油（HS1、HS2）风味物质总含量只有HS0（128.40 mg/kg）的21.79%～33.59%,且酯类相对含量下降至21.95%～38.47%;添加蒸汽爆破花生壳的酱油（BS3、BS4）风味物质组成结构（酯类相对含量为46.28%～86.51%）与HS0具有一定相似性,风味物质总含量大幅度提升（分别为514.89 mg/kg、451.74 mg/kg）,远高于HS0,其中蘑菇醇、十六酸乙酯、亚油酸乙酯等香气活性物质更突出。结果表明蒸汽爆破花生壳代替部分原料可明显提升酱油风味。     

一株耐盐产香鲁氏接合酵母FA-1的鉴定及其在酱油酿造中的应用

为提高酱油风味与品质,从酱油醪液中筛选耐盐产香酵母菌株并应用于酱油酿造。对筛选菌株进行鉴定,并采用顶空固相微萃取-气质联用法（HS-SPME-GC-MS）分别检测其在不同NaCl浓度的发酵液及酱油成品中的挥发性香气成分。结果表明,筛选出1株耐盐产香菌株FA-1,经鉴定为鲁氏接合酵母（Zygosaccharomyces rouxii）,可耐受22%的食盐浓度。在不同NaCl浓度的酱油发酵液中共检出56种挥发性风味物质,且当NaCl质量分数为18%时,更有利于醇类化合物的富集。在酱油生产试验中,添加鲁氏接合酵母FA-1的实验组中醇类化合物占比最大（72.03%）,其中苯乙醇、3-甲基-1-丁醇、乙醇的含量较高,且醇类和酯类化合物的含量分别较添加酵母As 1.039的对照组提高131.05%和189.65%。因此,菌株FA-1具有良好的耐盐产香性能,可作为潜在的酱油发酵菌株。     

替代法低盐酿造酱油品质研究

目的 为降低酱油中氯化钠的含量,采用氯化钾(K组)与乙醇(乙组)替代酱油中的氯化钠,对比研究两种低盐酱油的品质和安全性。方法 通过高盐稀态一贯型发酵酿造高品质低盐酱油,测定了不同氯化钾(K组)与乙醇(乙组)低盐酱油的氨基酸态氮,总酸,食盐,呈香物质等指标。结果 与CK组相比,氯化钾与氯化钠比例为4∶6,5∶5和6∶4时能显著增加酱油中氨基酸态氮的含量;乙醇和氯化钠的浓度比例分别为3∶20,4∶19,5∶18和6∶17时会降低酱油中总酸的含量。K组和乙组成品酱油中的食盐含量均随替代比例的增大而显著降低,且乙组降低更为显著。另外,氯化钾与氯化钠比例为4∶6的酱油成品挥发性香气物质的种类最多;乙醇和氯化钠比例为5∶18的酱油成品挥发性香气物质的种类最多且含量最高。结论 适量的氯化钾和乙醇替代氯化钠进行高盐稀态发酵可以获得高品质的低盐酱油,从安全性角度则是适量氯化钾优于乙醇,本研究为低盐酱油的酿造提供了一定理论基础。     

An Improved HPLC-FLD for Fast and Simple Detection of Ethyl Carbamate in Soy Sauce and Prediction of Precursors

A high-performance liquid chromatography with fluorescence detection (HPLC-FLD) method for ethyl carbamate (EC) determination in Chinese soy sauce was developed and used to predict EC precursors, and extraction, derivatization, and chromatographic conditions were optimized systematically. Under optimal conditions, the limit of detection, linear range, and recoveries were 3.91 μg/L, 12.87–274.87 μg/L, and 81.5–95.4%, respectively. The method precision was less than 8.9% (RSD) and no significant difference was found between EC determinations measured by HPLC-FLD and GC/MS. Using the proposed HPLC-FLD method, the EC contents in Chinese soy sauces ranged from not detected to 64.88 μg/L (n = 19). Except for one soy sauce brand, the EC contents in low-salt solid-state fermented (LSF) soy sauces were lower than those in high-salt liquid fermented (HLF) soy sauces and were mainly affected by alcohol content. Moreover, the free ornithine and total arginine contents were significantly correlated with EC content in the soy sauce products. The results of this work provide a foundation for further study of EC formation and inhibition in Chinese soy sauces.     

13种市售原酿本味酱油品质分析

采用电子鼻、气相色谱-质谱法（GC-MS）、氨基酸分析仪对市售的13种原酿本味酱油的挥发性物质、氨基酸组成进行解析,同时结合感官评价,对应分析评判不同产品风味物质组成及差异性。结果表明:受原料和发酵工艺的影响,我国酱油产品品质差异较大。由理化指标分析可知,不同产品乙醇和总糖含量变化最为明显。乙醇、氮氧化合物及无机硫化物是造成酱油风味差异的主要物质基础。酱油游离氨基酸组成不仅对酱油鲜味特征有显著影响,同时直接或间接与糖、酸、全氮等共同影响酱油鲜、咸、甜、酸等滋味特征。综合比较认为:当样品的氨基酸态氮≥1.00 g/100 mL,全氮≥1.80 g/100 mL,糖含量≥6.0 g/100 mL,糖/酸比值在3.40~5.80,游离氨基酸含量≥60 mg/mL时,酱油风味品质最佳。本文研究结果有助于补充和完善酱油风味物质图谱、指导建立酱油品质优劣评判标准,还可以用于追溯评估生产工艺技术条件及指导实际生产。     

大豆高压短时蒸煮显著提高酱油二次沉淀蛋白降解率

酱油二次沉淀严重影响其外观质量和商品价值。本研究以大豆低压长时(0.24 MPa/18 min)蒸煮工艺为对照,采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和基质辅助激光解吸附电离串联飞行时间质谱(MALDI-TOF/TOF MS)等方法研究了大豆高压短时(0.54MPa/14min)蒸煮工艺对酱油二次沉淀蛋白和二次沉淀生成量的影响。实验结果表明大豆球蛋白G4中的B3亚基(20 ku)和G1中的A1a亚基(35 ku)是酱油二次沉淀蛋白的主要成分,采用大豆高压短时蒸煮工艺制备的酱油(样品)比其对照酱油中B3亚基减少65.38%,样品中A1a亚基被完全降解;样品中二次沉淀含量比其对照减少72.69%。同时,采用大豆高压短时蒸煮工艺可提高酱油原料蛋白质利用率9.37%、氨基酸态氮14.14%、总氮10.74%、总糖22.36%、还原糖27.87%和无盐固形物12.38%(p0.05)。因此,采用大豆高压短时蒸煮工艺可显著提高酱油二次沉淀蛋白降解率、减少酱油二次沉淀生成量和提高酱油滋味物质含量,同时改善酱油外观质量和滋味。     

Characterisation of sugars as the typical taste compounds in soy sauce by silane derivatisation coupled with gas chromatography–mass spectrometry and electronic tongue

A total of 214 taste compounds were identified in soy sauce by silane derivatisation coupled with gas chromatography–mass spectrometry (SD-GC-MS), including 74 kinds of sugars. The proportion of total sugars is highest from 37.24–77.24% among all taste compounds. In particular, rare sugars were detected and identified as the special compounds in soy sauce which come from the sugar metabolism. Rare sugars identified in soy sauce by traditional fermented technology were prominent than Japanese technology. Principal component analysis (PCA) investigation of these results showed that samples can be distinguished by sugars as the first principal component and the general evaluation index (GEI) of samples was consistent with the result of sensory evaluation. Meanwhile, sweetness had the maximum range (from −9.89 to 14.10) in all taste indexes by electronic tongue (E-tongue) analysis.     

Quantification of ethyl carbamate in soy sauce consumed in Korea and estimated daily intakes by age

To estimate the daily intake of ethyl carbamate (EC, a possible human carcinogen) from soy sauce consumed in Korea, 136 soy sauce samples were collected from various regions and analysed by gas chromatography/selected ion mode mass spectrometry (GC/SIM‐MS). The distribution of EC varied significantly among samples, ranging from not detected to 128.9 µg kg^?1, with the highest level in Japanese‐style soy sauce. Based on individual intakes in the Complementary Report on 1998 National Health and Nutrition Survey and EC contents analysed in the present study, the contribution of soy sauce to EC intake was estimated by multiplying the mean concentration of EC by individual soy sauce intake data. Daily intakes of EC kg^?1 body weight by high consumers were particularly high in age groups 1–2 years (78.59 ng), 3–6 years (86.37 ng), 50–64 years (86.24 ng) and ≥ 65 years (76.86 ng). Based on a benchmark dose confidence limit (BMDL) of 0.3 mg EC kg^?1 body weight day^?1, the margin of exposure (MOE) calculated from the daily intake of EC in soy sauce for high consumers ranged from 3488 to 7317, which is of concern. Daily EC intakes are likely to be exceeded in groups who consume other fermented foods as well as alcoholic beverages. Copyright © 2006 Society of Chemical Industry     

Reducing the Influence of the Thermally Induced Reactions on the Determination of Aroma-Active Compounds in Soy Sauce Using SDE and GC-MS/O

A method based on simultaneous distillation extraction (SDE) coupled with gas chromatography–mass spectrometry/olfactometry (GC-MS/O) was developed for the analysis of volatile profiles in soy sauce. Its optimum operating conditions were as follows: 100 mL soy sauce plus 100 mL saturated brine; 2 h extraction using 50 mL dichloromethane. Eighty-eight volatile compounds were identified. Of these, 26 aroma-active compounds were detected by GC-O including 3-(methylthio)propanal, 2-methoxy-4-vinylphenol, 2-methoxyphenol, 2-methylbutanoic acid, 2,3-butanedione, phenylacetaldehyde, 2-ethyl-6-methylpyrazine, 3-methylbutanal, 2-methylbutanal, 3-methyl-1-butanol, and 2-methyl-1-butanol (which occurred at a high odor intensity in both high-salt liquid-state fermentation soy sauce (HLFSS) and low-salt solid-state fermentation soy sauce (LSFSS). SDE analysis of the odorless soy sauce indicated 12 volatiles (mainly Strecker aldehydes and α-dicarbonyl compounds). Thus, the proportions of these compounds (1.94–54.07 %) shown in previous GC-MS results could result from thermal degradation of non-volatile compounds in soy sauce. Especially, the contribution of 3-methylbutanal, 2-methylbutanal, phenylacetaldehyde, 3-(methylthio)propanal, and 2,3-butanedione to the aroma of soy sauce could be overestimated by conventional SDE-GC-O. With a full consideration of such limitations, this study will improve the application of SDE on aroma analysis of complex food matrix.     

挥发性成分聚类分析在酱油鉴别中的应用

应用同时蒸馏萃取法和气质联用技术对对酿造酱油、配制酱油和酸世界植物蛋白调味液共17个样品的挥发性成分进行分析,筛选其特征挥发性组分进行比较。发现酿造酱油特征组分以醇类、醛类、酸类为主,酸水解植物蛋白调味液特征组分以醛类、酚类、杂环化合物为主,而配制酱油挥发性成分则兼有前两者的特征组分。以17个样品的挥发性成分及其含量进行聚类分析,建立了区分酿造酱油、配制酱油和酸水解植物蛋白调味液的识别模型,为建立酿造酱油和配制酱油鉴别技术打下较好的基础。     

3-Monochloro-1,2-propandiol (3-MCPD) in soy sauce from the Bulgarian market

The 3-monochloro-1,2-propandiol (3-MCPD) levels in soy sauces which contained hydrolysed vegetable protein were evaluated for the Bulgarian market. For analysis of 3-MCPD, a gas chromatography–mass spectrometry (GC-MS) method was applied with a linear range of 0.03–2.00 μg mL^?1 and a limit of detection (LOD) of 2.3 μg kg^?1 and a limit of quantification (LOQ) of 3.4 μg kg^?1. At these levels, the standard deviation was 5.1%, with recoveries between 81% and 102%. The method was applied to the analysis of 21 samples of soy sauce from the Bulgarian market. Results ranged from 3.7 to 185.6 μg kg^?1. Soy sauces produced from hydrolysed soy protein contained higher levels of 3-MCPD than naturally fermented sauces. In 38.4% of samples of Bulgarian origin, the 3-MCPD content was above the EU limit of 20 μg kg^?1. In all analysed samples, 33.3% had a 3-MCPD content above the EU limit.     

液体发酵中以乙醇代盐生产低盐酱油的研究

用乙醇代替部分盐发酵55d生产的低盐酱油,其中20%盐、1%乙醇溶液泡曲发酵的原酱油中氨基酸态氮为0.865g/100mL,15%～17%乙醇溶液泡曲发酵生产的无盐原酱油中氨基酸态氮约为1.020g/100mL,提高了17%;在5%盐条件下,12%乙醇溶液泡曲得到低盐酱油的氨基酸态氮为0.992g/100mL,提高了14%;10%盐条件下,5%乙醇溶液泡曲得到低盐酱油的氨基酸态氮也达到0.990g/100mL,提高了大约14%。而含有10%盐、5%～7%乙醇的酱油样品风味最好。     

酱油增香用耐盐酵母的分离及生长特性分析

从酱醪中分离筛选出2株耐盐产香酵母菌C5、C6,并对其耐盐性及牛长特性进行研究。试验结果表明,2株酵母菌均能耐受24％（w／w）NaCl浓度,其在低盐稀态发酵生酱油的最适生长条件为温度28℃～32℃,pH4．5～5．5,酒精含量O～1．2％vol,葡萄糖或果糖添加量2％-4％（w／w）。此外,结果显示,菌株C5的还原糖消耗能力、酒精耐受性和产酒精能力均强于C6。     

Determination of HMF in Vinegar and Soy Sauce Using Two-Step Ultrasonic Assisted Liquid–Liquid Micro-Extraction Coupled with Capillary Electrophoresis-Ultraviolet Detection

A new, rapid, and inexpensive method using two-step ultrasonic assisted liquid–liquid micro-extraction (UALLME) coupled with capillary electrophoresis-ultraviolet (CE-UV) was developed for 5-hydroxymethylfurfural (HMF) analysis in high ion strength samples (like vinegar and soy sauce). The factors affecting the extraction efficiency were optimized such as the extraction volume, the ultrasonic time, and the power density of ultrasonic. Under the optimum conditions, the limit of detection (LOD) and the limit of quantification (LOQ) for HMF were 0.03 and 0.10 mg/L, respectively. The relative standard deviations (RSD %) for HMF were ranging from 0.53 to 3.17%, and the recoveries of HMF were ranging from 91.24 to 109.39%. The results turned out that two-step UALLME-CE-UV was applicable to analyze HMF in vinegar and soy sauce. Eleven brands of vinegar and soy sauce were tested by two-step UALLME-CE-UV, and the results showed that the method had a potential application in analysis of foodstuffs. The two step UALLME method was effective to improve the selectivity and sensitivity of CE-UV method for HMF analysis in vinegar and soy sauce.     

不同原料酱油抗氧化活性生物测试及风味分析

采用斑马鱼体内抗氧化实验比较了三种不同原料酿造酱油的抗氧化活性,同时采用电子鼻（E-nose）、气相色谱-质谱（GC-MS）结合主成分分析（PCA）分析比较了三种酿造酱油风味特征及风味物质组成。结果表明,黑豆酱油体内抗氧化活性显著高于豆粕和黄豆酱油（P＜0.05）,且其风味特征及挥发性物质组成与黄豆酱油、豆粕酱油均存在较大差异。风味特征差异主要表现为：三者相比氮氧化合物是黄豆酱油特征风味,无机硫化物、W2S醇类、W1S甲基类是豆粕酱油的特征风味,而黑豆酱油特征风味物质主要为杂环类化合物。挥发性风味物质组成差异表现为：黑豆酱油中2-乙酰基吡咯含量较高,豆粕酱油中乙醇和苯乙醛含量相对较高,而黄豆酱油中苯乙醇、4-乙基愈创木酚等重要风味物质相对含量较高。