Soy sauce is a traditional Chinese fermented condiments, and its distinctive flavor and high nutritional content have made it a vital ingredient in the diets of East Asians, and even Europeans, and Americans. 4-Ethylguaiacol is an essential aroma compound in soy sauce, and raising its concentration is advantageous for soy sauce′s quality. In this study, high-salt fermentation samples (sauce mash, fish sauce mash, etc.) from 10 high-salt fermentation condiment companies in four regions (East China, South China, Central China, and North China) were used to screen salt-tolerant lactic acid bacteria. After the initial screening and secondary examination, a salt-tolerant lactic acid bacterium LBM 20027 which produces 4-ethylguaiacol at high levels was isolated from soy sauce mash and been identified as Tetragenococcus halophilus. T. halophilus is the predominant microbe in the spontaneous soy sauce fermentation process and it can generate a variety of flavor components or flavor precursors. As we all know, one of the most important elements in assessing whether a strain can successfully colonize and enhance the flavor of a food product during fermentation is its fermentation performance, which includes but not limited to growth rate and acid tolerance. In this study, we conducted a comprehensive analysis of the physiological properties and fermentation performance of T. halophilus LBM 20027. T. halophilus LBM 20027 can grow under pH 5.5 or 20% NaCl concentration, and metabolizes glucose to produce lactate and acetate as its primary metabolites. In addition to producing a large amount of 4-ethylguaiacol, T. halophilus LBM 20027 can efficiently convert aspartic acid to sweet alanine effectively. We have also determined that this strain does not generate biogenic amine. We performed a simulated soy sauce fermentation using this strain as a starter. Improvements in acid production, amino acid concentration, and 4-ethylguaiacol production were seen when the strain was introduced. Using T. halophilus LBM 20027 as the starter, the 4-ethylguaiacol concentration increased significantly over the first 20 days of fermentation, and was 1.44 and 3.34 times greater than the control strain and the blank group, respectively. It seems that T. halophilus LBM 20027 showed a high efficiency in the synthesis of 4-ethylguaiacol in both single fermentation and simulated soy sauce fermentation. Through the comprehensive study on the fermentation and aromatic capacity of the strain, it was found that the excellent qualities of T. halophilus LBM 20027 will assist its continued use in soy sauce production.
[1] 杨兰. 高盐稀态酱油发酵过程工艺优化及作用机理的研究[D].广州:华南理工大学, 2010.
YANG L.The study on the optimization and mechanism during high-salt soy sauce fermentation with liquid state[D].Guangzhou:South China University of Technology, 2010.
[2] HU Y Y, ZHANG L, WEN R X, et al.Role of lactic acid bacteria in flavor development in traditional Chinese fermented foods:A review[J].Critical Reviews in Food Science and Nutrition, 2022, 62(10):2741-2755.
[3] ZHANG L J, ZHANG L, XU Y.Effects of Tetragenococcus halophilus and Candida versatilis on the production of aroma-active and umami-taste compounds during soy sauce fermentation[J].Journal of the Science of Food and Agriculture, 2020, 100(6):2782-2790.
[4] RZEPKOWSKA A, ZIELIŃSKA D, OŁDAK A, et al.Safety assessment and antimicrobial properties of the lactic acid bacteria strains isolated from polish raw fermented meat products[J].International Journal of Food Properties, 2017, 20(11):2736-2747.
[5] QI Q, HUANG J, ZHOU R Q, et al.Exploring a degradation strategy for biogenic amines based on the Cantonese soy sauce production method[J].Food Control, 2021, 130:108281.
[6] LI J L, HUANG J, JIN Y, et al.Aflatoxin B1 degradation by salt tolerant Tetragenococcus halophilus CGMCC 3792[J].Food and Chemical Toxicology, 2018, 121:430-436.
[7] YAO S J, ZHOU R Q, JIN Y, et al.Co-culture with Tetragenococcus halophilus changed the response of Zygosaccharomyces rouxii to salt stress[J].Process Biochemistry, 2020, 95:279-287.
[8] SINGRACHA P, NIAMSIRI N, VISESSANGUAN W, et al.Application of lactic acid bacteria and yeasts as starter cultures for reduced-salt soy sauce (moromi) fermentation[J].LWT, 2017, 78:181-188.
[9] HARADA R, YUZUKI M, ITO K, et al.Microbe participation in aroma production during soy sauce fermentation[J].Journal of Bioscience and Bioengineering, 2018, 125(6):688-694.
[10] UDOMSIL N, CHEN S, RODTONG S, et al.Improvement of fish sauce quality by combined inoculation of Tetragenococcus halophilus MS33 and Virgibacillus sp. SK37[J].Food Control, 2017, 73:930-938.
[11] 房峻, 刘佳乐, 彭玉慧, 等.强化嗜盐四联球菌对模拟低盐酱油发酵的影响[J].中国酿造, 2020, 39(12):146-152.
FANG J, LIU J L, PENG Y H, et al.Effect of enrichment of Tetragenococcus halophilus on simulated fermentation of low-salt soy sauce[J].China Brewing, 2020, 39(12):146-152.
[12] 王淑梅, 宋倩倩, 王晓磊, 等.传统发酵乳制品中乳酸菌的分离及鉴定[J].粮食与油脂, 2021, 34(2):5-7;17.
WANG S M, SONG Q Q, WANG X L, et al.Isolation and identification of lactic acid bacteria in traditional fermented dairy products[J].Cereals & Oils, 2021, 34(2):5-7;17.
[13] XU L L, ZHANG H M, CUI Y Q, et al.Increasing the level of 4-vinylguaiacol in top-fermented wheat beer by secretory expression of ferulic acid decarboxylase from Bacillus pumilus in brewer’s yeast[J].Biotechnology Letters, 2020, 42(12):2711-2720.
[14] 郭明威,耿予欢.酱油发酵过程中一株4-乙基愈创木酚转化菌的筛选及鉴定[J].食品与发酵工业, 2021, 47(21):18-23.
GUO M W, GENG Y H.Screening and identification of a 4-ethyl guaiacol producing strain in soy sauce fermentation[J].Food and Fermentation Industries, 2021, 47(21):18-23.
[15] 马瑞, 欧阳嘉, 李鑫, 等.高效液相色谱法同时测定生物质乳酸发酵液中有机酸及糖类化合物[J].色谱, 2012, 30(1):62-66.
MA R, OU Y J, LI X, et al.Simul aneous determination of orgamc acids and saccharides in lactic acid fermentation broth from biomass using high performance liquid chromatography[J].Chinese Journal of Chromatography, 2012, 30(1):62-66.
[16] 亢乐, 张丽杰, 徐岩.优质米曲霉筛选及复合菌剂构建提升郫县豆瓣蚕豆曲品质[J].食品与发酵工业, 2022, 48(22):18-25.
KANG L, ZHANG L J, XU Y.Generation of high-quality Aspergillus oryzae and thus mixed strains to improve the character of broad bean meju of Pixian Douban[J].Food and Fermentation Industries, 2022, 48(22):18-25.
[17] 鲍奕达. 郫县豆瓣酿造微生物来源分析及多肽降解功能微生物解析[D].无锡:江南大学, 2020.
BAO Y D.Source tracking of microbial community and functional microbiota analysis for polypeptide-degrading in Pixian broad bean paste[J].Wuxi:Jiangnan University, 2020.
[18] 郑鹏飞, 张丽杰, 王栋, 等.一种自下而上的合成微生物组理性构建策略, 用于郫县豆瓣发酵剂设计[J].微生物学报, 2022, 62(10):3913-3931.
ZHENG P F, ZHANG L J, WANG D, et al.A bottom-up strategy for constructing a synthetic microbiome and its application to the design of Pixian Douban starter[J].Acta Microbiologica Sinica, 2022, 62(10):3913-3931.
[19] 王博, 陈坚, 堵国成, 等.酱醪嗜盐四联球菌的分类及特性研究[J].微生物学报, 2018, 58(10):1826-1838.
WANG B, CHEN J, DU G C, et al.Classification and characteristics of Tetragenococcus halophilus derived from moromi[J].Acta Microbiologica Sinica, 2018, 58(10):1826-1838.
[20] 王少磊, 曹荣升, 沈芳, 等.浓香型大曲中4-乙基愈创木酚产生菌的筛选及其鉴定[J].酿酒科技, 2018(5):48-52.
WANG S L, CAO R S, SHEN F, et al.Screening and identification of a 4-ethyl guaiacol-producing strain from Nongxiang Daqu[J].Liquor-Making Science & Technology, 2018(5):48-52.
[21] 邹谋勇, 何理琴, 孙启星, 等.产4-乙基愈创木酚酵母的鉴定及其在酱油中的应用[J].食品科学, 2021, 42(12):138-144.
ZOU M Y, HE L Q, SUN Q X, et al.Identification of 4-ethylguaiacol producing yeast and its application in soy sauce brewing[J].Food Science, 2021, 42(12):138-144.
