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Research Progress on the effect of Bacillus on flavor substances of Maotai flavor Baijiu

Abstract

The brewing of Chinese Baijiu is a microbial community fermentation system, in which there is a close and complex relationship between microorganisms and flavor quality, but the specific deep-seated mechanism still needs to be explored and interpreted continuously so far. In this review, Maotai flavor Baijiu was briefly introduced, and the microorganisms mainly affected by flavor substances, the main components of flavor substances, the factors affecting flavor formation and the application ways of functional microorganisms in Maotai flavor Baijiu were summarized. It provides a deeper understanding for the research of others flavor Baijiu.

Keywords:
Maotai Flavor Baijiu; bacillus; functional components; bitterness; mechanism; influence factor

The six major distilled spirits in the world include whisky, brandy, vodka, rum and gin, as well as China's Baijiu (Shen, 1998Shen, Y. (1998). Complete book of Baijiu production technology. Beijing: China Light Industry Press.). China's traditional fermented distilled spirits have a history of nearly one thousand years. Among the different flavor types of Baijiu in China, Maotai flavor Baijiu is the most mysterious, with the most complex and abundant chemical component sand flavors (Jin et al., 2017Jin, G., Zhu, Y., & Xu, Y. (2017). Mystery behind Chinese liquor fermentation. Trends in Food Science & Technology, 63, 18-28. http://dx.doi.org/10.1016/j.tifs.2017.02.016.
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; Liu et al., 2018Liu, Y., Song, H., & Luo, H. (2018). Correlation between the key aroma compounds and gDNA copies of Bacillus during fermentation and maturation of natto. Food Research International, 112, 175-183. http://dx.doi.org/10.1016/j.foodres.2018.06.033. PMid:30131126.
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; Wang et al., 2019Wang, M. Y., Yang, J. G., Zhao, Q. S., Zhang, K. Z., & Su, C. (2019). Research progress on flavor compounds and microorganisms of Maotai flavor Baijiu. Journal of Food Science, 84(1), 6-18. PMid:30548499.). The typical aroma characteristics of Maotai flavor Baijiu, such as Maotai flavor, caramel flavor, paste flavor, mellow flavor and fruit flavor, as well as the mellow, delicate and long aftertaste, are deeply loved by consumers all over the world (Shen, 1998Shen, Y. (1998). Complete book of Baijiu production technology. Beijing: China Light Industry Press.; Xu et al., 2012bXu, Y., Ji, K. & Piggott J. (2012b). Moutai (maotai): Production and sensory properties. Alcoholic Beverages, 25(S60), 315-330. http://dx.doi.org/10.1533/9780857095176.3.315.
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). In the process of tasting Maotai flavor Baijiu, it mainly includes two aspects: flavor and taste. The former is mainly soy sauce flavor, and the characteristic substances are mainly furan compounds, aromatic compounds and pyrazine compounds (Cang, 2013Cang, Y. (2013). A brief talk on the source of heterogenous flavor in fragrance type Baijiu. Chinese Brewing, 32(5), 134-136.); The latter is mainly "aftertaste" bitter and astringent, that is, Baijiu remains on the root of the tongue or the back of the root of the tongue, which gives people an unpleasant feeling, affects the overall style of the liquor, and directly affects the sensory quality and grade of Baijiu (Xiao, 2014Xiao, L. (2014). Study on formation mechanism and control measures of bitter substances in rice flavor Baijiu. Guangzhou: Zhong kai College of Agricultural Engineering.). Therefore, bitter and astringent taste is listed as the most common peculiar smell in Baijiu (Xiao et al., 2011Xiao, D., Zhao, S., & Chen, Y. (2011). Baijiu production technology (pp. 210-232). Beijing: Chemical Industry Press.). Baijiu in China has bitter and astringent taste to varying degrees (Cang, 2013Cang, Y. (2013). A brief talk on the source of heterogenous flavor in fragrance type Baijiu. Chinese Brewing, 32(5), 134-136.; Xiao et al., 2014Xiao, Z., Hou, X., Lyu, X., Xi, L., & Zhao, J. (2014). Accelerated green process of tetramethylpyrazine production from glucose and diammonium phosphate. Biotechnology for Biofuels, 7(1), 106. http://dx.doi.org/10.1186/1754-6834-7-106.
http://dx.doi.org/10.1186/1754-6834-7-10...
; Wu, 2005Wu, Z. (2005). Bitterness in Luzhou flavor Baijiu. Winemaking Technology, 134(8), 121-123.). The precursors of flavor substances are mostly macromolecular substances, and the degradation of macromolecular substances cannot be separated from the role of microorganisms. Most wine industry peers believe that high-temperature Daqu is the main source of soy sauce flavor substances. Under high temperature, only heat-resistant bacteria are left.

Bacillus is a strain with high biosafety. Because of its fast growth, strong resistance, inhibition of pathogens and other characteristics, it has become a dominant strain in soil and plant ecological environment (Chen et al., 2003Chen, Z., Jie, Z., & Huang, D. (2003). Study on antibacterial mechanism and genetic improvement of Bacillus spp. for biological control of plant diseases. Journal of Plant Pathology, 33(2), 97-103., 2012Chen, Z., Liu, Y., Liu, Y., & Zhang, R. (2012). Research progress of biocontrol Bacillus against plant diseases. Journal of Jiangsu Agriculture, 28(5), 999-1006. , 2018b Chen, Y., Yang, Y., Sang, J., Cai, J., Xu, C. & Huang, G. (2018b). Isolation and identification of antagonistic endophytic bacillus BEB17 and analysis of antibacterial activity of its volatile substances. Journal of Plant Pathology, 48(4), 537-546.; Jing et al., 2018Jing, W., Cao, J., Chen, D., Qiu, J., Wang, X., Feng, C. & Wang, W. (2018). Antibacterial activity and component analysis of volatile organic compounds from Bacillus pupils AR03. China Agricultural Sciences, 51(10), 1908-1919.). Bacillus the Thermophilic bacteria, Bacillus subtilis and Bacillus licheniformis all have strong ability to produce amylase and proteinase (Pylypenko et al., 2021Pylypenko, L., Yegorova, A., Volovyk, T., Konovka, A., & Oliinyk, L. (2021). Rapid sanitary and technological control of culturing the producers of amylolytic enzymes. Food Science and Technology (Campinas), 15(3), 23-29. https://doi.org/10.15673/fst.v15i3.2112.
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). Some amino acids and polypeptides can make the wine taste bitter and astringent, so they are important fungi for flavor generation and style formation of Maotai flavor Baijiu (Gao, 2010Gao, Y. (2010). Polymerase chain reaction denaturing gradient electrophoresis (PCR-DGGE) was used to study the microbial community structure in Daqu of Chinese Baijiu. Wuxi: Jiangnan University.). Studies have confirmed that the characteristic components of Maotai flavor Baijiu are mainly furan compounds, aromatic compounds and pyrazine compounds (Zhuang, 2005Zhuang, M. Y. (2005). Further discussion on Maillard reaction products and the aroma and taste of Chinese Baijiu. Brewing Technology, (5), 34-38.), while thermophilic spores can produce acetoin (Xiao et al., 2009Xiao, Z., Ma, C., Xu, P., & Lu, J. R. (2009). Acetoin catabolism and acetylbutanediol formation by Bacillus pumilus in a chemically defined medium. PLoS One, 4(5), e5627. http://dx.doi.org/10.1371/journal.pone.0005627. PMid:19461961.
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; Nicholson, 2008Nicholson, W. L. (2008). The Bacillus subtilis ydjL (bdhA) gene encodes acetoin reductase/2,3-butanediol dehydrogenase. Applied and Environmental Microbiology, 74(22), 6832-6838. http://dx.doi.org/10.1128/AEM.00881-08. PMid:18820069.
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), tetramethylpyrazine (Adachi et al., 1964Adachi, T., Kamiya, H., & Kosuge, T. (1964). Studies on the metabolic products of bacillus subtilis. 3. relation between amino acids and tetramethylpyrazine production. Yakugaku Zasshi-journal of the Pharmaceutical Society of Japan, 84, 543-545. http://dx.doi.org/10.1248/yakushi1947.84.6_543. PMid:14207111.
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) and furanyl.

At present, Bacillus subtilis has been widely used in scientific research (Kang et al., 2021Kang, Q., Xiang, M., & Zhang, D. (2021). Research progress and industrial application of Bacillus subtilis in systematic and synthetic biotechnology. Sheng Wu Gong Cheng Xue Bao = Chinese Journal of Biotechnology, 37(3), 923-938. PMid:33783158.; Wang et al., 2019Wang, M. Y., Yang, J. G., Zhao, Q. S., Zhang, K. Z., & Su, C. (2019). Research progress on flavor compounds and microorganisms of Maotai flavor Baijiu. Journal of Food Science, 84(1), 6-18. PMid:30548499.), food (Syahbanu et al., 2020Syahbanu, F., Kezia, E., Puera, N., Giriwono, P. E., Tjandrawinata, R. R., & Suhartono, M. T. (2020). Fibrinolytic bacteria of Indonesian fermented soybean: preliminary study on enzyme activity and protein profile. Food Science and Technology (Campinas), 40(suppl 2), 458-465. http://dx.doi.org/10.1590/fst.23919.
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; Alashbayeva et al., 2021Alashbayeva, L., Shansharova, D., Mynbayeva, A., Borankulova, A., & Soltybayeva, B. (2021). Development of technology for bakery products. Food Science and Technology (Campinas), 41(3), 775-781. http://dx.doi.org/10.1590/fst.61120.
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), medicine and other fields. Among them, Bacillus subtilis is the most prominent strain, which is recognized as safe (GRAS) by the US Food and Drug Administration (FDA) because of its strong strain safety. Natto (Japan), tempeh, soy sauce (China), pickles, fermented bean curd and Baijiu are all used. gather γ- Glutamic acid(γ- PGA) is water-soluble, biodegradable and edible. Because of its strong viscosity, it can be used as thickener or flocculant. Research findings γ- PGA is made from soybeans fermented by Bacillus spp. and is also an ingredient of Japanese traditional food natto (Bang et al., 2011Bang, B.-H., Jeong, E.-J., Rhee, M.-S., Kim, Y.-M., & Yi, D.-H. (2011). Isolation of Bacillus subtilis GS-2 Producing γ-PGA from Ghungkukjang Bean Paste and Identification of γ-PGA. Journal of Applied Biological Chemistry, 54(1), 1-6. http://dx.doi.org/10.3839/jabc.2011.001.
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). Douchi is one of the most popular foods in China, because it has a strong Chifalavour (soybean flavor (Sun et al., 2022Sun, X., Du, J., Chen, Y., Zhong, R., Fan, Y., Zhou, X., & Wang, Y. (2022). Research progress on antioxidation of fermented soybean. Modern Chinese Medicine, 1-8.), a special ammonia odor different from natto). Bacillus subtilis is one of the most important microorganisms in the fermentation process of Douchi. Kosuge & Kamiya (1962)Kosuge, T., & Kamiya, H. (1962). Tetramethylpyrazine: a Bacillus subtilis growth factor. Nature, 194(4833), 1099. http://dx.doi.org/10.1038/1941099a0. PMid:14458749.
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first proposed that Bacillus subtilis can synthesize tetramethylpyrazine TTMP more than half a century ago. Tetramethylpyrazine (TTMP) is a nitrogen-containing heterocyclic compound, which contributes to the formation of many aromatic and flavor compounds (Xiao et al., 2014Xiao, Z., Hou, X., Lyu, X., Xi, L., & Zhao, J. (2014). Accelerated green process of tetramethylpyrazine production from glucose and diammonium phosphate. Biotechnology for Biofuels, 7(1), 106. http://dx.doi.org/10.1186/1754-6834-7-106.
http://dx.doi.org/10.1186/1754-6834-7-10...
; Liu et al., 2018Liu, Y., Song, H., & Luo, H. (2018). Correlation between the key aroma compounds and gDNA copies of Bacillus during fermentation and maturation of natto. Food Research International, 112, 175-183. http://dx.doi.org/10.1016/j.foodres.2018.06.033. PMid:30131126.
http://dx.doi.org/10.1016/j.foodres.2018...
; Gao et al., 2017Gao, L., Liu, T., An, X., Zhang, J., Ma, X., & Cui, J. (2017). Analysis of volatile flavor compounds influencing Chinese-type soy sauces using GC-MS combined with HSSPME and discrimination with electronic nose. Journal of Food Science and Technology, 54(1), 130-143. http://dx.doi.org/10.1007/s13197-016-2444-0. PMid:28242911.
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; Wang et al., 2022Wang, J. D., Liu, D. T., Wang, X. Y. (2022). The formation and stabiility of tetramethylpyrazi ne in the brewing process of Shanxi aged vinegar. Shipin Yu Fajiao Gongye, 48(15), 62-68.; Fan et al., 2007aFan, W., Xu, Y., & Zhang, Y. (2007a). Characterization of pyrazines in some Chinese liquors and their approximate concentrations. Journal of Agricultural and Food Chemistry, 55(24), 9956-9962. https://doi.org/10.1021/jf071357q.
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). TTMP belongs to pyrazine substances detected in fermented foods (Hou et al., 2016Hou, X., Gu, R., Liang, W., & Xiao, Z. (2016). Research progress in the production of tetramethylpyrazine by fermentation. Shengwu Jishu Tongbao, (1), 58-64.). A recent study proved that TTmp in Chinese Baijiu is mainly produced by Bacillus subtilis under micro oxygen conditions (Zhang et al., 2019Zhang, W., Si, G., Li, J., Ye, M., Feng, S., Mei, J., Yang, Q., Wang, J., & Zhou, P. (2019). Tetramethylpyrazine in Chinese sesame flavour liquor and changes during the production process. Journal of Brewing Research Institute, 125(1), 155-161. http://dx.doi.org/10.1002/jib.527.
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). Bacillus subtilis used in Chinese sufu can improve the antioxidant activity, fibrinolytic function and sensory quality of sufu (Wang, 2022Wang, H. (2022). Study on Fermentation Process Optimization and Flavor Quality Formation of Bacterial Douchi. USA: Southwest University.). In addition, Bacillus is often used as an excellent chassis cell and transformed into a microbial cell factory for the production of industrial enzymes, vitamins, functional sugars, health products, drug precursors and other target products, with strong industrial production and application capacity (Gu et al., 2018Gu, Y., Xu, X. H., Wu, Y. K., Niu, T., Liu, Y., Li, J., Du, G., & Liu, L. (2018). Advances and prospects of Bacillus subtilis cellular factories: from rational design to industrial applications. Metabolic Engineering, 50, 109-121. http://dx.doi.org/10.1016/j.ymben.2018.05.006. PMid:29775652.
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; Ÿztürk et al., 2016Ÿztürk, S., Ÿalik, P., & Ÿzdamar, T. H. (2016). Fed-batch biomolecule production by Bacillus subtilis: a state of the art review. Trends in Biotechnology, 34(4), 329-345. http://dx.doi.org/10.1016/j.tibtech.2015.12.008. PMid:26775901.
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; Zweers et al., 2008Zweers, J. C., Barák, I., Becher, D., Driessen, A. J., Hecker, M., Kontinen, V. P., Saller, M. J., Vavrová, L., & van Dijl, J. M. (2008). Towards the development of Bacillus subtilis as a cell factory for membrane proteins and protein complexes. Microbial Cell Factories, 7(1), 10. http://dx.doi.org/10.1186/1475-2859-7-10. PMid:18394159.
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).

Maotai flavor Baijiu is loved by consumers for its mellow Maotai flavor and long aftertaste. In this paper, the source of microorganisms, the main components and formation mechanism of flavor substances, and the factors affecting flavor formation in Maotai flavor Baijiu were reviewed. This review will provide inspiration for the follow-up study of Baijiu flavor, and provide a deeper understanding of the bitter and astringent taste of other foods.

1 Overview of Maotai flavor Baijiu

1.1 Research status of Chinese Baijiu

In order to sum up the traditional experience, Chinese Baijiu carried out pilot studies in Maotai, Fenjiu and Luzhou Laojiao in the 1950s and 1960s; Mechanization research in 1970s; Research on tick off technology from 1980s to 1990s; In 2007, universities and enterprises cooperated in the development (Shen, 1998Shen, Y. (1998). Complete book of Baijiu production technology. Beijing: China Light Industry Press.; Shen & Tong, 2007Shen, Y., & Tong, Z. (2007). Understanding and academic discussion on the flavor type of Baijiu. Winemaking, 34(1), 3-4.; Yan, 2015Yan, X. (2015). Study on microorganism and metabolic regulation of Chinese Baijiu based on flavor oriented technology. Brewing Technology, (2), 1-11.). Now the research mainly focuses on the establishment of microorganisms, flavor substances, production technology, intelligent equipment system, etc. Baijiu is divided into 12 flavor types: thick, clear, soy sauce, rice, Phoenix, Jianhe, Dong, special, soy sauce, sesame, Lao baigan and Fuyu (Fan & Qian, 2010Fan, W., & Qian, M. C. (2010). Identification of aroma compounds in Chinese 'Yanghe Daqu" liquor by normal phase chromatography fractionation followed by gas chromatography olfactometry. Flavour and Fragrance Journal, 21(2), 333-342. https://doi.org/10.1002/ffj.1621.
https://doi.org/10.1002/ffj.1621...
). As the representative of Maotai liquor in China, Maotai liquor attracts the attention of researchers at home and abroad due to its unique flavor and complex production technology. However, at present, most scholars study more Luzhou flavor Baijiu, while the typical Maotai flavor Baijiu is relatively less. The production process is the key to Maotai flavor. At present, the fermentation process of most Maotai flavor Baijiu is "four high and two long". In addition to the fermentation process, the fermentation metabolism mechanism of microorganisms is also the core technology. There is a saying about the fermentation mechanism that "fermentation depends on yeast, aroma production depends on bacteria, and saccharification depends on mold". In the process of bacterial metabolism, abundant and complex enzymes are produced, which participate in the formation of flavor substances, and thus determine the flavor and quality of Baijiu. Therefore, in recent years, the research on flavor producing functional bacteria using molecular biology technology and traditional microbial culture methods has gradually increased. Due to the open production environment of Maotai flavor Baijiu, the microorganisms in the fermentation process mainly include environmental entry and Daqu addition. The formation process of Maotai flavor in Maotai flavor Baijiu is precisely the reflection of mutual regulation and mutual growth of functional microorganisms in these bacteria. Now, the microorganisms in the fermentation process of Maotai flavor Baijiu (Wang et al., 2008aWang, C. L., Shi D. J & Gong, G. L. (2008a). Microorganisms in Daqu: a starter culture of Chinese Maotai-flavor liquor. World Journal of Microbiology & Biotechnology, 24(10), 2183-2190. http://dx.doi.org/10.1007/s11274-008-9728-0.
http://dx.doi.org/10.1007/s11274-008-972...
, 2014Wang, X., Fan, W., & Xu, Y. (2014). Comparison on aroma compounds in Chinese soy sauce and strong aroma type liquors by gas chromatography-olfactometry, chemical quantitative and odor activity values analysis. European Food Research and Technology, 239(5), 813-825. http://dx.doi.org/10.1007/s00217-014-2275-z.
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, 2015aWang, J., Wang, X. D., Qiu, S. Y., & Hu, P. G. (2015a). Advances in research on the formation of Maotai-flavor liquor brewing system features. Liquor-Making, 4, 99-104. , 2016aWang, L., Hu, G., Lei, L., Lin, L., Wang, D., & Wu, J. (2016a). Identification and aroma impact of volatile terpenes in Moutai liquor. International Journal of Food Properties, 4(6), 1335-1352. http://dx.doi.org/10.1080/10942912.2015.1064442.
http://dx.doi.org/10.1080/10942912.2015....
; Wu et al., 2012Wu, Q., Xu, Y., & Chen, L. (2012). Diversity of yeast species during fermentative process contributing to Chinese Maotai-flavor liquor making. Letters in Applied Microbiology, 55(4), 301-307. http://dx.doi.org/10.1111/j.1472-765X.2012.03294.x. PMid:22862564.
http://dx.doi.org/10.1111/j.1472-765X.20...
, 2013Wu, Q., Chen, L., & Xu, Y. (2013). Yeast community associated with the solid state fermentation of traditional Chinese Maotai-flavor liquor. International Journal of Food Microbiology, 166(2), 323-330. http://dx.doi.org/10.1016/j.ijfoodmicro.2013.07.003. PMid:23978339.
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, 2015Wu, Q., Chen, B., & Xu, Y. (2015). Regulating yeast flavor metabolism by controlling saccharification reaction rate in simultaneous saccharification and fermentation of Chinese Maotai-flavor liquor. International Journal of Food Microbiology, 200, 39-46. http://dx.doi.org/10.1016/j.ijfoodmicro.2015.01.012. PMid:25676241.
http://dx.doi.org/10.1016/j.ijfoodmicro....
; Xiong, 2005Xiong, Z. S. (2005). Research on three flavor type Baijiu in China (ii) introduction to Maotai-flavor liquor. Niangjiu Ke-Ji, 4, 25-30.; Xiu et al., 2012Xiu, L., Kunliang, G., & Hongxun, Z. (2012). Determination of microbial diversity in Daqu, a fermentation starter culture of Maotai liquor, using nested PCR-denaturing gradient gel electrophoresis. World Journal of Microbiology & Biotechnology, 28(6), 2375-2381. http://dx.doi.org/10.1007/s11274-012-1045-y. PMid:22806111.
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; Xu & Ji, 2012a; Xu et al., 2017Xu, Y., Sun, B., Fan, G., Teng, C., Xiong, K., Zhu, Y., Li, X., & Li, X. (2017). The brewing process and microbial diversity of strong flavour Chinese spirits: a review. Journal of the Institute of Brewing, 123(1), 5-12. http://dx.doi.org/10.1002/jib.404.
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; Yang et al., 2007bYang, D., Fan, G., Wang, D., & Lv, Y. (2007b). Study on microorganism in high temperature Daqu. Winemaking Technology, (5), 37-38.; Zhang et al., 2013Zhang, R., Wu, Q., & Xu, Y. (2013). Aroma characteristics of Moutai-flavor liquor produced with Bacillus licheniformis by solid-state fermentation. Letters in Applied Microbiology, 57(1), 11-18. http://dx.doi.org/10.1111/lam.12087. PMid:23594087.
http://dx.doi.org/10.1111/lam.12087...
; Zheng et al., 2016Zheng, Y., Sun, B., Zhao, M., Zheng, F., Huang, M., Sun, J., Sun, X., & Li, H. (2016). Characterization of the key odorants in Chinese zhima aroma-type Baijiu by gas chromatography-olfactometry, quantitative measurements, aroma recombination, and omission studies. Journal of Agricultural and Food Chemistry, 64(26), 5367-5374. http://dx.doi.org/10.1021/acs.jafc.6b01390. PMid:27263543.
http://dx.doi.org/10.1021/acs.jafc.6b013...
) are summarized in Table 1. At the present stage, the production of Baijiu still continues the traditional production process, which has strong experience and uncertainty. However, with the continuous deepening of the research on Baijiu, in order to improve the national liquor making level, we should innovate and develop on the basis of the traditional process. At present, the urgent research includes the analysis of flavor substances in Baijiu, the development of compound Baijiu, and the establishment of digital system. The healthy development of Baijiu industry is an important development direction of the brewing industry.

Table 1
Microorganisms in the fermentation of Maotai flavor Baijiu.

1.2 Traditional production process of Maotai flavor Baijiu

A sone of the six major distilled spirits in the world, Baijiu with Maotai flavor is loved by consumers for its unique flavor of "prominent Maotai flavor, elegant and delicate, soft and mellow, and long aftertaste". Maotai, the representative of Maotai flavor Baijiu, is well-known at home and abroad (Shen, 1998Shen, Y. (1998). Complete book of Baijiu production technology. Beijing: China Light Industry Press.; Li, 1999Li, D. (1999). Great achievements of Baijiu production technology in the past 50 years since the founding of the people's Republic of China (VI). Winemaking, (6), 19-31.). Through research, it is believed that taste is closely related to brewing process. Maotai flavor Baijiu adopts the brewing process of "four highs and two longs, one big and one big". The "four highs" refer to high-temperature koji making, high-temperature stacking, high-temperature fermentation and high-temperature distillation, "two longs" refer to production cycle and storage time, "one big" refers to large koji consumption, and "many times" refers to multiple rounds (fermentation) of liquor taking. The production cycle is linked in a year (Wu et al., 2012Wu, Q., Xu, Y., & Chen, L. (2012). Diversity of yeast species during fermentative process contributing to Chinese Maotai-flavor liquor making. Letters in Applied Microbiology, 55(4), 301-307. http://dx.doi.org/10.1111/j.1472-765X.2012.03294.x. PMid:22862564.
http://dx.doi.org/10.1111/j.1472-765X.20...
; Xu & Ji, 2012aXu, Y., & Ji, K. (2012a). Moutai (Maotai): production and sensory properties. In J. Piggott (Ed.), Alcoholic beverages: sensory evaluation and consumer research (pp. 315-330). Sawston, Reino Unido: Woodhead Publishing.; Li, 1999Li, D. (1999). Great achievements of Baijiu production technology in the past 50 years since the founding of the people's Republic of China (VI). Winemaking, (6), 19-31.; Cui, 2007aCui Li. (2007a). The key process to form the style and quality of Maotai flavor liquor is "four high and two strong, one big and one many". Wine Making, 34 (3): 24-35.).

The quality of high-temperature Daqu determines the style and quality of Maotai flavor Baijiu. An abstract term is "Qu is the bone of liquor". The raw materials of Daqu are mostly sorghum or wheat, which are naturally fermented. During this period, a large number of microorganisms and flavor precursors will be gradually enriched, and then pressed into starter bricks, and then mature starter will be prepared. This process requires sufficient water, oxygen and high temperature. Under the condition of sufficient water and high temperature, high-temperature resistant microorganisms grow, reproduce and metabolize, decompose various enzymes to produce soy sauce flavor substances. On this basis, create an anoxic environment, so that high-temperature resistant, anaerobic and facultative anaerobic bacteria can multiply in large numbers to obtain the required flavor substances (Cui, 2007bCui Li. (2007b). The key process to form the style and quality of Maotai flavor liquor is "four high and two strong, one big and one many". Wine Making, 34 (3): 24-35.; Ming et al., 2010Ming, H., Liu, Y., Zhuo, Y., Cheng, W. & Yang, W. (2010). Influence of koji making temperature on the quality of Maotai flavor Daqu. Chinese Brewing, 7, 157-160.; Hu et al., 2010Hu, P., Qiu, S., & Li, J. (2010). The relationship between the key links of Maotai flavor Daqu liquor production technology and its style and quality. Winemaking Technology, (8), 36-39.).

The brewing stage of Maotai flavor Baijiu includes two parts: high-temperature stacking koji making and high-temperature stacking liquor making. During the preparation of cooked koji, a large number of yeasts died. In the process of stacking fermentation, open fermentation enable microorganism in the air environment to participate in the fermentation, transforming starch, protein and other macromolecules in raw materials into amino acids, oligosaccharides, fatty acids and other small molecules. This process is equivalent to "secondary koji making" (Peng et al., 2004Peng, J., Jiang, H., Wang, H., Zhang, Z. (2004). Formation mechanism of "waist line" in the accumulation fermentation process of Maotai flavor Baijiu. Brewing Technology, (6), 43-44.; Xu, 2019Xu, L. (2019). Research on technology of Guojing mianya Maotai flavor Baijiu. China: Qilu University of Technology.; Sun et al., 2013Sun, J., Liu, W., Wei, Z., Shen, G., & Ping, W. (2013). Composition of mold community and functional enzyme activity in fermented grains of Maotai flavor Baijiu. Chinese Journal of Food, 13(8), 239-247.). Bacteria, molds and their enzymes can decompose raw materials, which belongs to the role of high temperature accumulation in wine production, aroma generation and saccharification fermentation. In this process, there is a unique process that the fermented grains are piled into the pit, so that the ester producing yeast can ensure the need for subsequent fermentation and wine production. In the fermentation stage of the pit, the growth of some harmful microorganisms is inhibited because the temperature rises to generate acid. Acid can prevent the growth of certain rancidity bacteria, so that the koji is not sour and smelly, thus forming a favorable environment for the generation of soy sauce flavor.

Maotai flavor Baijiu needs to undergo eight rounds of fermentation, multiple rounds of fermentation and multiple rounds of liquor taking. The specific operation steps are shown in (Dai, 2019Dai Y. (2019) Microbial community composition in the brewing process of Maotai flavor Baijiu and its relationship with liquor quality, Hunan Agricultural University.) Figure 1. However, the beneficial microorganisms in the fermentation process are more pure, the compound Maotai flavor in the fermented grains is more intense, the impurities of Maotai flavor precursors are less, and the wine style is better. In addition to multiple rounds, liquor recycling fermentation is also a major feature of Maotai flavor Baijiu fermentation. Liquor recycling fermentation process can nourish distillers' grains with liquor, increase aroma components with high boiling point, and then promote liquor body to be fragrant. The storage time of Baijiu determines the quality of liquor. The storage time of Maotai flavor Daqu liquor is not the same according to different koji. The storage time of Maotai flavor Daqu liquor is more than three years, while the storage time of Maotai flavor bran koji is more than one and a half years. The complex production technology and storage means are the reasons why Maotai flavor Baijiu is different from others flavor Baijiu.

Figure 1
Main production process of Maotai flavor wine. 1. Sorghum 2. Steamed grain 3. Airing 4. Heap fermentation 5. Pit fermentation 6. Steamed wine 7. Take wine 8. Storage 9. Tick.

2. Research status of Bacillus in Maotai flavor Baijiu

The main power source of Maotai flavor Baijiu fermentation has always been considered to be the bacterial group (Fan & Qian, 2010Fan, W., & Qian, M. C. (2010). Identification of aroma compounds in Chinese 'Yanghe Daqu" liquor by normal phase chromatography fractionation followed by gas chromatography olfactometry. Flavour and Fragrance Journal, 21(2), 333-342. https://doi.org/10.1002/ffj.1621.
https://doi.org/10.1002/ffj.1621...
), but because Maotai flavor Baijiu is an open stacking fermentation, the bacterial source should include two parts: environment and Daqu. High temperature accumulation is an important link commonly used in Maotai flavor Baijiu and thick sauce combined Baijiu. Its purpose is to enzymatically hydrolyze starch and protein in fermented grains into sugars and amino acids, and react to generate flavor substances (Zeng, 2012Zeng, C. (2012). Enzymatic hydrolysis of corn brant prepare ferry oligosaccharides and its antioxidant properties. Qiqihar: Qiqihar University; Xiao et al., 2005Xiao, P., Jing, W., & Hui, Y. (2005). Feruloyl Oligosaccharide stimulation growth of Bifidobacterium bifidum. Anaerobe, 11(4), 225-229. http://dx.doi.org/10.1016/j.anaerobe.2005.02.002. PMid:16701572.
http://dx.doi.org/10.1016/j.anaerobe.200...
). The environment of high-temperature brewing is very suitable for the growth of Bacillus groups, which leads to a large number of enrichments of Bacillus groups, making Bacillus the most abundant and abundant bacterial group in the brewing environment of Maotai flavor Baijiu (Wang et al., 2015aWang, J., Wang, X. D., Qiu, S. Y., & Hu, P. G. (2015a). Advances in research on the formation of Maotai-flavor liquor brewing system features. Liquor-Making, 4, 99-104.).

Flavor substances in Baijiu are mainly produced by the decomposition of raw materials by microorganisms (environment + koji) (Hai, 2013 Hai, D. (2013). Influence mechanism and monitoring control of native taste producing bacteria on Baijiu brewing. Wuxi: Jiangnan University.; Hu, 2015Hu, X. (2015). Study on the correlation between the diversity of Clostridium community in pit mud of Luzhou flavor Baijiu and the quality of pit mud. Wuxi: Jiangnan University.). Studies have found that molds decompose starch and protein to produce glucose and amino acids, and produce saccharifying enzymes and amylases through reproduction and metabolism; The bacteria metabolize glucose and amino acid to produce flavor components; Yeast metabolizes glucose to produce ethanol (Wang, 2014Wang, H. (2014). Study on the Evolutionary Law of Microbial Community Structure of Fen Liquor with Fragrance by PCR-DGGE. Wuxi: Jiangnan University.). As for environmental microorganisms playing a very important role in Baijiu brewing, it is often said in the industry that "if you leave Maotai Town, you can't brew Maotai liquor". However, at present, most of the studies on the micro ecology of Baijiu brewing are only focused on the succession of microbial communities during koji making or fermentation, and the studies on the microorganisms in the brewing environment are also limited to the study of air microorganisms in the environment. There is no in-depth study on other environmental microorganisms such as soil in the brewing environment. Moreover, due to the high diversity of wine making environment microecology, conventional culturable or PCR-DGGE technologies in the past could not have a good understanding of wine making environment microecology, and the role of environmental microorganisms in wine making lacked quantitative analysis. Therefore, it is of great significance to analyze the microbial structure of Baijiu fermentation environment, clarify the source of Baijiu brewing microorganisms, analyze the impact of environmental microorganisms on Baijiu fermentation, and analyze the formation mechanism of Daqu microecology to improve the controllability of Baijiu brewing and improve the quality of Baijiu.

2.1 research status of micro ecology of Baijiu fermentation

In recent years, some studies have shown that the formation of Maotai flavor of Baijiu is related to the role of bacteria, and functional microorganisms for Baijiu production have been isolated from Baijiu koji and fermented grains everywhere, including various functional bacteria for Maotai flavor (Peris & Escuder-Gilabert, 2009Peris, M., & Escuder-Gilabert, L. (2009). A 21st century technique for food control: electronic noses. Analytica Chimica Acta, 638(1), 1-15. http://dx.doi.org/10.1016/j.aca.2009.02.009. PMid:19298873.
http://dx.doi.org/10.1016/j.aca.2009.02....
). Cheng Caiying (Cheng et al., 2014Cheng, C., Liu, X., Yuan, Y., Yan Z, Liao Y & Fu S. (2014). Bacterial community structure of Maotai flavor Baijiu koji and seven consecutive rounds of accumulated fermented grains. Journal of Applied and Environmental Biology, 20(05), 825-831.) and others conducted a detailed analysis of the diversity of bacterial flora in Maotai flavor Baijiu koji and fermented grains of different rounds. Through research, they found that there were common dominant bacterial flora in koji and fermented grains of different rounds: Bacillus licheniformis, the Actinomyces, B. subtilis、B. cereus、Lentibacillus. The unique brewing technology of Maotai flavor liquor forms a special microbial flora in its Daqu and fermented grains (Chen et al., 2013Chen, L., Qun, W., & Yan, X. (2013). Study on the characteristics of Schizosaccharomyces cerevisiae with low yield of fusel oil in Maotai flavor Baijiu. International Microbiology, 43(3), 1-6.). Chang lu Wang et al. (2008bWang, C. L., Shi D. J & Gong, G. L. (2008b). Microorganisms in Daqu: a starter culture of Chinese Maotai-flavor liquor. World Journal of Microbiology & Biotechnology, 24(10), 2183-2190. http://dx.doi.org/10.1007/s11274-008-9728-0.
http://dx.doi.org/10.1007/s11274-008-972...
) used the culturable method to analyze the microbial population of Maotai Maotai Maotai Daqu at different maturity stages, and isolated more than 200 strains, mainly Bacillus. High temperature Daqu used in the production of Maotai flavor Baijiu is one of the main sources of Maotai flavor of Baijiu (Shen, 1998Shen, Y. (1998). Complete book of Baijiu production technology. Beijing: China Light Industry Press.). Yang Daiyong (Yang et al., 2007aYang, D. Y., Fan, G. X., Wang, D. Q., & Lu, Y. H. (2007a). Microbes in high temperature starter. Niangjiu Ke-Ji, 5(155), 37-38.) and others preliminarily isolated 98 kinds of microorganisms from Maotai Daqu, including 41 kinds of bacteria, mostly Bacillus, 6 kinds of yeasts, 51 kinds of molds, including Aspergillus, Mucor, etc. Zhang Rong (Rong et al., 2010Rong, Z., Yan, X., Fan, W., & Wang, H. (2010). Analysis of Bacillus licheniformis and its characteristic flavor metabolites in Maotai flavor Daqu. International Microbiology, 40(3), 7-12.) and others screened three strains of Bacillus licheniformis with the function of producing Maotai flavor from Maotai flavor Baijiu. The results showed that acetoin, tetramethylpyrazine and furan twist may be related to the formation of Maotai flavor of Baijiu, but this result still needs further exploration and research. Yu Tingting (Yu et al., 2013Yu, T., Lai, S., Cao, W., Xu, C., & Liu, T. (2013). Screening and identification of high-temperature resistant bacteria producing soy sauce flavor in high-temperature Daqu. Guizhou Agricultural Sciences, 41(11), 109-112.) and others screened a functional Bacillus gzub-2 producing Maotai flavor from a Maotai flavor Baijiu and identified it as Bacillus licheniformis. Zhao Xingxiu (Zhao et al., 2016Zhao, X., He, Y., Zhao, C., Fang, C., Zhang, J., & Zou, W. (2016). Screening of functional bacteria producing soy sauce flavor and study on its flavor substances. Science and Technology of Food Industry, 37(6), 196-200.) and others analyzed the soy sauce ingredients in the soy sauce fermentation experiment through HS-SPME and GC-MS methods from three strains of soy sauce producing functional bacteria Bacillus subtilis in the high-temperature Daqu of a soy sauce distillery. The results showed that the contents of pyrazine and ketone in the soy sauce ingredients of the three strains were high, and the analysis of pyrazine and ketone may have a greater impact on the flavor of soy sauce.

2.2 Research status of microenvironment of Baijiu fermentation environment

The microenvironment of Baijiu fermentation environment is an important component of the microenvironment of Baijiu fermentation and an important factor affecting the quality, safety and flavor characteristics of Baijiu. Microecology of fermentation environment shall include all the environments directly or indirectly contacted by grain raw materials during operation, including but not limited to workshop floor, operating tools, air, workers and other factors.

Due to the close relationship between environmental microecology and Baijiu production, the research on brewing environmental microecology is also increasing. Wang Xingchu (Wang & Guo, 2011Wang, X., & Guo, K. (2011). Study on airborne bacteria flora in different production environments of Maotai liquor. Winemaking Technology, (9), 29-31.) used molecular biology and bioinformatics technology to study the air bacterial flora in the production environment of Maotai Liquor. The results showed that in the production environment of Maotai Distillery, bacteria in the air were selectively enriched, forming a bacterial flora with large cell density and complex structure. Wang Yanhua (Wang, 2016Wang, Y. (2016). Analysis on the microflora structure of the fermentation environment of Fumao cellar wine. China: Fujian Normal University.) used the second generation high-throughput sequencing technology to study the air of the new and old workshops of Fumao cellar liquor and the microbial flora structure in the accumulated and fermented fermented grains. The results showed that during the process from accumulation fermentation to the end of fermentation in the cellar, the bacterial flora structure changed greatly, while the fungal flora structure changed slightly, and the fungal groups in the air of the brewing workshop were closely related to the brewing process of Fumao cellar liquor. Zhang Yali (Zhang, 2014Zhang, Y. (2014). Identification and Analysis of Air Microbes in Maotai, Renhuai Prefecture, Guizhou Province. Beijing: Beijing University of Chemical Technology.) took the air microorganisms in Maotai ancient town as the research object, and carried out regular collection, culture, separation, purification, identification, total extraction, sequencing and microbial community analysis (divided by obvious seasons) four times a year. The research results showed that a large part of microorganisms in the brewing process of Maotai flavor Baijiu came from the air environment. Zhang et al. (2008) Zhang, L., Shen, C., Zhang, S., & Xu, D. (2008). Analyze the regulation of metabolic capacity of functional bacteria in pit mud. Brewing Technology, 61(1), 57-58. compared the composition of bacteria in the air inside and outside Luzhou Laojiao Workshop and believed that Bacillus is the main bacterial population in the air.

3 Research progress on flavor substances of Maotai flavor Baijiu

3.1 Research status of main aroma components

So far, it has been clear that the main aroma component of Luzhou flavor liquor is ethyl caproate; Fen flavor liquor is ethyl acetate; Rice flavored liquor may be ethyl lactate, ethyl acetate and p-phenylethanol. Some characteristic components were also found in other aromatic wines, but the main aroma and flavor components needed to be further analyzed. At present, there are many inferences about the characteristic flavor components of Maotai flavor liquor, but what its essential characteristics are still seems to be a mystery. The studied characteristic substances of Maotai flavor are summarized in Table 2.

Table 2
special flavor substances of Maotai flavor Baijiu.

4-Ethylguaiacol (4-EG)

In 1964, Maotai liquor pilot project (Li et al., 2021a Li, Q., Wang, J., & Zhang, X., Fu C., Wang C., Xu, N. & Lu, W. (2021a). Research progress of 4-vinyl guaiacol, a flavor substance. Chinese Brewing, (4), 15-19.), according to the conditions at that time, proposed for the first time that the main aroma component of Maotai flavor liquor was 4-ethyl guaiacol. 4-ethyl guaiacol is a typical representative of high boiling point compounds in soy sauce aroma, which has the characteristic aroma of soy sauce (Li et al., 1990aLi, C., Peng, Z., & Yang, D. (1990a). What are the main aroma components of Maotai flavor liquor? Some opinions on the main flavor components of Maotai flavor liquor. Winemaking, 56(1), 13-15.). In 1976 (Xiong, 2000Xiong, Z. (2000). Guizhou maotai liquor survey review. Winemaking Technology, (4), 26-29., 2005aXiong, Z. (2005a). Study onthe three major flavor types of Baijiu in China (II) Maotai. Winemaking Technology, (4), 25-30.), a study was conducted by combining gas chromatography analysis with sensory taste, and it was found that 4-ethyl guaiacol had the smell of Beijing "fumigation". It was mellow, sweet, and had nothing to do with the soy sauce flavor, so it was not the main flavor of Maotai liquor.

Pyrazine

Some scholars believe that the main way of formation of soy sauce flavor substances is the thermal decomposition of amino acids and proteins; Sugar involved in the related reactions and some microbial metabolites. Cao Shushun also proposed that pyrazine compounds may be related to the formation of soy sauce flavor (Zhang et al., 2022bZhang, M., Chen, Z., Tian, D., Pan, L. & Liu, W. (2022b). Research progress on health functional components of Luzhou flavor Baijiu. Food Safety Guide, (15), 110-113.): high-temperature Daqu has a flavor similar to soy sauce flavor more than medium temperature Daqu, and its aromatic components may be related to the volatile compounds (such as aldehydes and pyrazines) generated by the reaction. In addition, factors such as large amount of koji, large concentration of matrix, high reaction temperature, etc. in the production process will inevitably produce or accelerate the Maillard reaction rate, thus increasing the production of pyrazine compounds. During distillation, pyrazine compounds volatilize with steam and condense into liquor due to the effect of vapor belt, creating Maotai flavor style. In the later study (Xiong, 1994Xiong, Z. (1994). Brewing of Maotai flavor Baijiu. Beijing: China Light Industry Press.), a researcher conducted sensory identification on tetramethylpyrazine, and found that its smell was not obvious, like the smell of soaking bean water, sweet and strong.

Pyrazine compounds have the aroma and baking smell of green pepper, which are found in large quantities in baked food and can make food produce flavors similar to popcorn, nuts, coffee, spinach and chocolate. However, compounds similar to soy sauce, soy sauce and wheat soy sauce have no aromatic effect except tetramethylpyrazine (Li et al., 1990bLi, C., Peng, Z., & Yang, D. (1990b). What are the main aroma components of Maotai flavor liquor? Some opinions on the main flavor components of Maotai flavor liquor. Winemaking, 56(1), 13-15.; Zhang et al., 2022cZhang, M., Chen, Z., Tian, D., Pan, L., & Liu, W. (2022c). Research progress on health functional components of Luzhou flavor Baijiu. Food Safety Guide, (15), 110-113.; Shen, 2010Shen, H. (2010). Study on aroma substances of Maotai flavor Baijiu (pp. 1-8). Wuxi: Jiangnan University.), so it is believed that pyrazine compounds have limited aromatic effect. Some of these compounds have very low thresholds, while others have very high thresholds. For example, the threshold of pyrazine can reach 300 mg/L (in water), the threshold of 2-methylpyrazine can reach 30 mg/L (in water) (Zhu et al., 2022Zhu, Y., Chen, H., Deng, G., & Fu, H. (2022). Research progress on the detection, source, aroma and functional activity of pyrazines in high-temperature Daqu Baijiu. Science and Technology of Food Industry, 1-12.), and the threshold of 2-methoxy-3-isobutylpyrazine is only 0.002 μ G/L (in water) (Ding et al., 2022Ding, H., Shen, X., Song, C., Xiong, Y., Deng, B., Ao, L., Zeng, S., Li, J., & Yu, J. (2022). Research progress of pyrazine compounds as health functional components of Chinese Baijiu. Chinese Brewing, (9), 8-12.), its threshold value is very different, and does not show soy sauce flavor. The addition test conducted by some researchers proved that (Fan & Yan, 2012Fan, W., & Yan, X. (2012). Review and prospect of research on maotai flavor substances in Maotai flavor Baijiu. Winemaking, 39(3), 8-16.), the addition of pyrazine, especially tetramethylpyrazine with high content, could not produce subtle soy sauce aroma. When pyrazine compounds were added to Luzhou flavor Baijiu, no Maotai flavor was found in Luzhou flavor Baijiu; At the same time, when pyrazines were added to the atypical Maotai flavor Baijiu, it was not found that the Maotai flavor Baijiu became more typical. Therefore, pyrazine compounds are not the key aroma components or main aroma of Maotai flavor Baijiu, but they are the characteristic components of Maotai flavor Baijiu.

Furan and pyran derivatives

In 1983, some studies speculated that furans and pyranoids were important aroma substances and characteristic compounds of Maotai flavor Baijiu. The aroma of Maotai flavor Baijiu (Zhou, 1983Zhou, L. (1983). Discussion on the main aroma components in Maotai flavor Baijiu. Wine Making, (1), 20-22.) is mainly a compound aroma composed of Maotai flavor, burnt flavor, ester flavor and alcohol flavor. The important components of Maotai flavor Baijiu are Maotai flavor and burnt flavor. These two substances are derived from the fat and amino acid components in raw materials and the substances produced by the metabolism of yeast. In the production process, they are affected by raw materials, brewing process, koji, water quality and aging time. These furans and pyran compounds may include HEMF, HDMF, and HEMF has cake and biscuit aroma, which is obviously enhanced when added to soy sauce; When added to Baijiu, the style did not improve significantly. It indicates that the Maotai flavor of soy sauce or sauce is not the same as that of Maotai flavor Baijiu (Dong et al., 2016Dong, L., Xu, C., & Yan, Z. & Li, K. (2016). Discussion on flavor composition and style of Sichuan soy sauce wine. Wine Making, (1), 39-42.).

The furfural content of Maotai flavor Baijiu is higher than that of other flavor Baijiu, which has become a marker component of such flavor Baijiu. Its unique Maotai flavor may have some internal and inevitable relationship with the high content of furfural (Shen, 2010Shen, H. (2010). Study on aroma substances of Maotai flavor Baijiu (pp. 1-8). Wuxi: Jiangnan University.; Xu, 2002Xu, H. (2002). Study on the relationship between furfural content and flavor type in Baijiu. Winemaking, 29(5), 29-30.). Some studies reported that Wuliangye and Jiannanchun Liquor were studied by liquid-liquid extraction GC-MS, and seven furans were detected (Fan & Qian, 2006Fan, W., & Qian, M. C. (2006). Characterization of aroma compounds of Chinese “Wuliangye” and “Jiannanchun” liquors by aroma extract dilution analysis. Journal of Agricultural and Food Chemistry, 54(7), 2695-2703. https://doi.org/10.1021/jf052635t.
https://doi.org/10.1021/jf052635t...
). In addition, hsspme technology was used to study the aroma substances in high-temperature Daqu, and nine furans were found (Fan et al., 2007bFan, W., Zhang, Y., & Xu, Y. (2007b). HS-SPME and GC-MS analysis of trace volatile components in Baijiu Daqu. Winemaking Technology, (12), 74-78.), which is the report with the highest content of furans found in the research on Chinese Baijiu Daqu (Zhao et al., 2008Zhao, S., Fan, W., & Yan, X. (2008). Study on furans in fermented grains of Maotai flavor Baijiu production. Brewing in China, 21, 10-13.). Therefore, some researchers speculate that the main aroma components of Maotai flavor liquor may be furan and pyran derivatives.

3.2 Research status of auxiliary flavor

Sour, sweet, bitter, salty and fresh are the five basic tastes widely accepted by people (Zhang et al., 2017Zhang, Y., Venkitasamy, Z., Pan, Z., Liu, W., & Zhao, L. (2017). Novel umami ingredients: Umami peptides and their taste. Journal of Food Science, 82(1), 16-23. http://dx.doi.org/10.1111/1750-3841.13576. PMid:27926796.
http://dx.doi.org/10.1111/1750-3841.1357...
). In Baijiu, the taste of Baijiu mainly includes sour taste, sweet taste and bitter taste. Although an appropriate amount of bitter taste can enrich its taste (Zhang, 2011Zhang, Y. (2011). Discussion on “bitterness” in Baijiu. Winemaking, 38(1), 61-62.), in most Baijiu, bitter taste will be regarded as peculiar smell because of its high intensity. All kinds of Baijiu have some bitterness problems. Although there are a lot of studies on bitter substances in food (Wiener et al., 2012Wiener, A., Shudler, M., Levit, A., & Niv, M. Y. (2012). BitterDB: a database of bitter compounds. Nucleic Acids Research, 40, D413-D419. http://dx.doi.org/10.1093/nar/gkr755. PMid:21940398.
http://dx.doi.org/10.1093/nar/gkr755...
) internationally, the formation reason and chemical nature of bitter taste in Baijiu are still unclear. Recently, Wang Yinye et al. Took sesame flavor Baijiu as the research object, analyzed the volatile bitter substances in Baijiu, and identified volatile bitter compounds such as furfural, 2-methylpropanol, 3-methylbutanol, 1-butanol and n-propanol (Wang, 2018Wang, Y. (2018). Study on volatile bitter and / or astringent substances in Baijiu. Wuxi: Jiangnan University.).

However, under normal circumstances, flavoring substances are often difficult to volatilize (Farmer, 1994Farmer, L. J. (1994). The role of nutrients in meat flavor formation. The Proceedings of the Nutrition Society, 53(2), 327-333. http://dx.doi.org/10.1079/PNS19940038. PMid:7972147.
http://dx.doi.org/10.1079/PNS19940038...
). In China's Baijiu, the special solid-state distillation process makes Baijiu also contain some nonvolatile components (Zhang et al., 2014aZhang, R., Wu, Q., Xu, Y., & Qian, M. C. (2014a). Isolation, identification, and quantification of lichenysin, a novel nonvolatile compound in Chinese distilled spirits. Journal of Food Science, 79(10), C1907-C1915. http://dx.doi.org/10.1111/1750-3841.12650. PMid:25250990.
http://dx.doi.org/10.1111/1750-3841.1265...
), and studies show that compared with other flavor Baijiu, Maotai flavor Baijiu has the highest content of nonvolatile components and the most complex components (Wei, 2007Wei, Z. (2007). Study on nonvolatile components of Baijiu. Nanjing: Nanjing University of Technology.). Therefore, we speculate that there are still some hard to volatile bitter substances in Maotai flavor Baijiu, which may contribute to the bitterness of Maotai flavor Baijiu to a certain extent, and the research on these taste substances needs to be in-depth.

Furfural

Furfural is the main component of furan system in Baijiu. It is produced by pentosan in rice husk and raw material skin under the action of microorganisms. Furfural is one of the important components of "grain aroma" or "burnt aroma" in wine, but excessive furfural will make wine scorched bitter, and acrolein bitter (Hu, 2006Hu, Y. (2006). “Bitterness” of Baijiu and its solution. Winemaking Technology, 143(5), 67-69.). However, at present, the research on furfural in Baijiu is only qualitative, quantitative and flavor analysis, and there is no report on its taste. The research on acrolein is only limited to the detection and quantification of acrolein in Baijiu for the first time (Zhu et al., 2016Zhu, M., Fan, W., Xu, Y., & Zhou, Q. (2016). 1,1-Diethoxymethane and methanethiol as age markers in Chinese roasted-sesame-like aroma and flavor type liquor. European Food Research and Technology, 242(11), 1985-1992. http://dx.doi.org/10.1007/s00217-016-2697-x.
http://dx.doi.org/10.1007/s00217-016-269...
). Acrolein is easy to be produced in abnormal fermentation due to poor health management of Baijiu fermentation. It is produced by yeast and lactic acid bacteria acting on glycerol during Baijiu fermentation, so it is also known as glyceraldehyde. Acrolein not only has a pungent odor, but also has a great persistent bitter taste (Song et al., 2021Song, K., Wang, D., & Chen, C. (2021). Analysis of bitter taste of Baijiu and Study on its prevention measures. Winemaking Technology, (12), 56-60.).

Higher alcohols

Higher alcohols are alcohol soluble substances with high boiling points, which refer to univalent alcohols with more than three carbons. Liquor mainly containsn-butanol, isobutanol, n-propanol, isoamyl alcohol, etc. The mixture of these higher alcohols is what we usually called fusels. Fusels in Baijiu are not only the chromatographic skeleton components of Baijiu, but also an important component of Baijiu. It is generally believed that tyrosol in fusel is extremely bitter and persistent, isobutanol is extremely bitter, n-propanol is bitter, n-butanol is bitter, and isoamyl alcohol is slightly sweet and bitter (Wang, 2007Wang, H. (2007). Bitterness of Baijiu. Winemaking Technology, 158(8), 165-167.). However, the reported studies on these bitter substances are only qualitative and quantitative (Wang et al., 2014Wang, X., Fan, W., & Xu, Y. (2014). Comparison on aroma compounds in Chinese soy sauce and strong aroma type liquors by gas chromatography-olfactometry, chemical quantitative and odor activity values analysis. European Food Research and Technology, 239(5), 813-825. http://dx.doi.org/10.1007/s00217-014-2275-z.
http://dx.doi.org/10.1007/s00217-014-227...
), and there is no relevant literature report on the threshold measurement and taste analysis of their concentration in the wine body. The main way of producing higher alcohols in wine is that during fermentation, the protein in raw materials is hydrolyzed into amino acids, and the amino acids are deamination and carbon dioxide are used to generate higher alcohols one carbon less than amino acids. Tyrosol, also known as p-hydroxyphenylethanol, is one of the main diols in Baijiu. Although its aroma is very soft, its bitter taste is heavy and long. Generally, when Baijiu contains 0.5/10000 tyrosol, it will have a bitter taste in tasting. The formation of tyrosol is mainly caused by excessive yeast acting on tyrosine. Therefore, excessive use of koji will lead to excessive tyrosol, which makes the wine bitter. This is also the root cause of the bitter of Daqu (Song et al., 2021Song, K., Wang, D., & Chen, C. (2021). Analysis of bitter taste of Baijiu and Study on its prevention measures. Winemaking Technology, (12), 56-60.).

Other substances

There are many bitter components in Baijiu. The raw materials and auxiliary materials for liquor making contain more tannins, which can be decomposed into catechol, pyrophenol, phloroglucinol and other monomers, all of which have bitter or astringent taste (Liu & Sun, 2018 Liu, H., & Sun, B. (2018). Effect of fermentation processing on the flavor of Baijiu. Journal of Agricultural and Food Chemistry, 66(22), 5425-5432. http://dx.doi.org/10.1021/acs.jafc.8b00692. PMid:29751730.
http://dx.doi.org/10.1021/acs.jafc.8b006...
). It is generally believed that sulfide, disulfide (containing-S-S -) and sulfhydryl (- SH) compounds have bitter taste (Lai, 2009Lai, D. (2009). Strengthening the safety and quality control of alcoholic food -- Analysis of the causes of abnormal taste in Baijiu and solutions. Wine Making, (5), 3-5.). At present, qualitative, quantitative and flavor studies have been carried out on sulfur-containing compounds in Baijiu (Sha et al., 2017Sha, S., Chen, S., Qian, M., Wang, C., & Xu, Y. (2017). Characterization of the typical potent odorants in Chinese roasted sesame-like flavor type liquor by headspace solid phase microextraction-aroma extract dilution analysis, with special emphasis on sulfur-containing odorants. Journal of Agricultural and Food Chemistry, 65(1), 123-131. http://dx.doi.org/10.1021/acs.jafc.6b04242. PMid:27989125.
http://dx.doi.org/10.1021/acs.jafc.6b042...
). As for alkaloids, there are no reports about alkaloids in Baijiu. Most peptides are bitter peptides. There are peptides in Baijiu, so it is inevitable to have bitter taste (Wang et al., 2007Wang, H., Zheng, L., & Han, X. (2007). Talk about the “bitterness” of Baijiu. Winemaking, 34(4), 43-44.). However, the current research on polypeptides in Baijiu is only limited to the discovery of polypeptides in Baijiu (Yan et al., 2014Yan, X., Rong, Z., Qun, W., & Zhi, Y. (2014). Identification of bioactive lipopeptide compounds in Baijiu and Study on their functions. Winemaking Technology, 246(12), 1-7.), and the quantification of polypeptides in Baijiu (Zhi et al., 2017Zhi, Y., Qun, W., & Yan, X. (2017). Lipopeptide compounds as bioactive substances in traditional Baijiu and their production mechanism. Liquor-Making Science & Technology, 280 (10), 17-23.). Amino acids exist in Baijiu (Zhang, 2011Zhang, Y. (2011). Discussion on “bitterness” in Baijiu. Winemaking, 38(1), 61-62.). The internationally reported bitter amino acids include L-histidine, L-valine, L-isoleucine, L-leucine, L-lysine, L-phenylalanine, L-tyrosine and L-arginine (Hufnagel & Hofmann, 2008Hufnagel, J. C., & Hofmann, T. (2008). Quantitative reconstruction of the nonvolatile sensometabolome of a red wine. Journal of Agricultural and Food Chemistry, 56(19), 9190-9199. http://dx.doi.org/10.1021/jf801742w. PMid:18710249.
http://dx.doi.org/10.1021/jf801742w...
). At present, 18 kinds of amino acids have been detected in Baijiu (Zhang et al., 2014bZhang, Z., Fan, W., & Yan, X. (2014b). Comparative analysis of free amino acids in different flavor Baijiu. Science and Technology of Food Industry, 35(17), 280-284.). Some soluble inorganic ions, such as magnesium ions and sulfate ions, will bring certain bitterness to the wine, such as MgSO4 and MgCl2, which will lead to bitter taste of Baijiu after blending (Wang, 2004Wang, Y. (2004). The causes and prevention measures of abnormal taste in Baijiu. Journal of Jilin Specialty College, 13(2), 16-17.).

4 Factors affecting the formation of flavor substances

4.1 Impact on brewing technology

Baijiu brewing is an alcoholic beverage made by cooking, saccharification, fermentation, distillation, storage, blending and other processes with sorghum, corn and wheat as raw materials and Daqu saccharifying starter (Zhao et al., 2020Zhao, Y., Tang, J., & Lu, J. (2020). Overview of the influence of brewing technology on Maotai flavor Daqu liquor. Modern Food, (5), 106-111.). Baijiu can be divided into 12 types according to flavor. The brewing process and style characteristics of Maotai flavor, rice flavor, Luzhou flavor and light flavor are shown in Table 3. The brewing process is closely related to the formation of Baijiu style. The most complicated brewing process is the Maotai flavor Baijiu process, which takes sorghum as raw material and adopts stone cellar fermentation. It has the characteristics of "four highs and two longs", that is, the "four highs" are high-temperature koji making, high-temperature fermentation, high-temperature cooking and high-temperature distillation; "Two long" means long production cycle and long storage time of base liquor.

Table 3
Brewing and style characteristics of four kinds of Baijiu.

High temperature is a crucial step in the process of Maotai flavor Baijiu (Yang et al., 2021Yang, G., Cao, L., Pan, C., Xu, X., & Qian, S. (2021). Study on the accumulation process of Zhilan flavor Baijiu. Fermentation Science and Technology Communication, 50(2), 82-85.), which can promote the decomposition of protein and amino acid, the reaction between sugar and protein, ammonia, browning reaction and microbial metabolism in Daqu, so as to form aroma components, such as ketones, aldehydes, alcohols, furans, Pyrans, thiophenols and pyrazines through Maillard reaction (Hai, 2016Hai, C. (2016). Brief analysis of Maillard reaction and the formation of elegant Baijiu style with Yi lite thick sauce. Winemaking, 43(6), 61-65.). The accumulation process can not only enrich many microorganisms, but also produce many biological and chemical reactions, which can form flavor substances, which is very important to enhance the Maotai flavor and improve the quality of Maotai flavor Baijiu. Liu Minwan et al. (Liu et al., 2019Liu, M., Pan, X., Yin, F., & Ge, R. (2019). Study on the mechanism of high-temperature accumulation process of Maotai flavor Baijiu and the analysis of round liquor production. Wine Making, 46(2), 60-64.) found that the flavor compounds detected in the late stage of the accumulated fermented grains were more volatile than those in the early stage. The consistency between flavor substances and aroma components of Maotai flavor finished liquor was high. Yang ManJiang (Yang, 2011Yang, M. (2011). Effect of stacking fermentation on the formation of flavor substances in Maotai flavor Baijiu. Winemaking Technology, (7), 72-73.) studied the effect of stacking fermentation on flavor substances of Maotai flavor Baijiu. The research showed that Maillard reaction occurred during high-temperature stacking fermentation, and furans, pyrazines and aromatic compounds could be generated. The storage of base liquor also affects the style of Baijiu.

The base liquor has strong entrance irritation and poor taste, and the storage period of Maotai flavor Baijiu base liquor is generally not less than 3 years. Qin Dan et al. (Dan et al., 2021Dan, Q., Duan, J., Li, Y., He, F., Li, H., & Sun, B. (2021). Changes of flavor components during aging of Baijiu and research progress of artificial aging technology. Shipin Kexue, 42(23), 260-267.) showed that with the increase of storage time, the mass concentration of total acid and total alcohol in Maotai flavor Baijiu decreased, while phenols, sulfur-containing and nitrogen-containing compounds increased. Jiang Jinjin et al. (Jiang et al., 2019Jiang, J., Li, Y., Li, G., Peng, S., & Chen, S. (2019). Change analysis of flavor components of a sauce flavor Baijiu stored for a long time. Journal of Guangzhou City Vocational College, 13(1), 28-33.) studied the volatile flavor substances of Maotai flavor Baijiu aged 3 months and 20 years, and found that besides the Maotai flavor compound furfural, there was also acetoin (the precursor of 4-methylpyrazine) in 20-year-old Baijiu. Zhang Jiaojiao et al. (Zhang et al., 2022aZhang, J., Liang, M., Lei, X., Xue, X., Chen, Y., Shen, S., He, M., Hu, J., Xie, S., Zhang, M., & Han, X. (2022a). Correlation analysis between sensory characteristics and characteristic flavor of mianrou Maotai flavor Baijiu of Yingbin liquor. Chinese Brewing, 41(2), 69-75.) analyzed the Maotai flavor liquor stored for 3 years, founding that its esters and acids accounted for a high proportion, while alcohols accounted for less.

4.2 Impact on brewing materials

Raw materials for making Baijiu include sorghum, corn, wheat and rice. Liquor making raw materials are not only required for the growth of liquor making microorganisms, but also can be decomposed into alcohol, acid, ester, aldehyde, ketone and other Baijiu flavor substances or precursor substances of flavor substances. The flavor substances produced by different Baijiu making raw materials are not the same. Therefore, liquor making raw materials are one of the important factors affecting the style, taste and quality of Baijiu (Zhong & Zhang, 2022Zhong, M., & Zhang, J. (2022). Research status of the influence of waxy sorghum on the quality of Maotai flavor Baijiu. Chinese Brewing, 41(1), 32-36.). Glutinous sorghum and wheat are used to brew Maotai flavor Baijiu, among which sorghum contains starch, protein, tannin, fat and other components. The mass fraction of amylopectin in glutinous sorghum is not less than 80%, which is suitable for multiple rounds of fermentation of Maotai flavor Baijiu (Degree et al., 2022Degree, C. J., Wang, K., Li, D., Luo, X., & Chen, J. (2022). Research Progress on the effect of sorghum on the quality of Maotai flavor Baijiu. Shipin Kexue, 43(7), 356-364.). Phenolic flavor substances can be formed by fermentation of proper amount of tannin and protein in waxy sorghum and ferulic acid in wheat. During fermentation, proteins in sorghum react with sugars to form furan, pyrazine, pyrrole and other flavor substances (Ding et al., 2019Ding, Y., Zhou, L., Wang, C., Cao, N., Cheng, B., Gao, X., Peng, Q., & Shao, M. (2019). Research progress of glutinous sorghum for Maotai flavor liquor. Shengwu Jishu Tongbao, 35(5), 28-34. http://dx.doi.org/10.13560/j.cnki.biotech.bull.1985.2019-0080.
http://dx.doi.org/10.13560/j.cnki.biotec...
). Jiao Shaojie et al. (Jiao et al., 2015Jiao, S., Wang, L., Jiang, Y., Yan, H., Su, D., & Sun, G. (2015). Research review on the relationship between sorghum and solid Baijiu. Wine making, 42(1), 13-16.) showed that during the fermentation process, the fat in sorghum reacts to produce low molecular aldehyde or ketone, which causes the wine body to sour. Therefore, a small amount of fat in sorghum is beneficial to the wine body quality.

4.3 Microbial impact

In the brewing process of Maotai flavor Baijiu, a large number of microbial flavor, including bacteria, yeast and mold, are enriched through high-temperature koji making, high-temperature accumulation and fermented grains fermentation (Wei et al., 2020Wei, H., Cheng, P., Jian, Z., Li, L., & Hu, F. (2020). Research progress on influencing factors and main flavor components of Maotai flavor Baijiu. Brewing Technology, (4), 85-93.). When microorganisms grow and metabolize, macromolecular substances in raw materials are decomposed into small molecular substances by Secretase to provide rich flavor precursor substances; Some microorganisms produce flavor substances such as vitamins, auxin and amino acids through metabolism (Xu et al., 2016Xu, B., Cui, S., Wang, J., & Liu, L. (2016). HS-SPME/GC-MS analysis of volatile aroma components in four types of famous wines. Journal of Shenzhen Vocational and Technical College, 15(3), 53-58.). Microorganism is an important source that affects the flavor of Maotai flavor Baijiu. In the high-temperature Daqu of Maotai flavor Baijiu brewing, bacteria are a very important group of microorganisms (Xin et al., 2021Xin, S., Hui, M., Tian, Q., & Huang, J. (2021). Application and research progress of esterase. Fermentation Science and Technology Communication, 50(4), 228-232.), abundant in quantity and the power source of flavor production. Flavor substances produced through fermentation and metabolism play an important role in the flavor formation of Maotai flavor Baijiu. Mo Zhenni et al. (Mo et al., 2022Mo, Z., Qiu, S., Zeng, X., & Dai, Y. (2022). Based on Illumina sequence high-throughput sequencing, the bacterial community difference between Daqu peel and Daqu core of Maotai flavor Baijiu was analyzed. Chinese Brewing, 41(2), 48-52.) analyzed the colony structure of Daqu of Maotai flavor Baijiu through high-throughput sequencing technology, and found that there were 55 bacterial genera, including lactobacillus, bacillus, polysaccharide polysaccharide, Cladosporium and Streptomyces. Li et al. (2014 Li, H., Lian, B., Ding, Y., Nie, C., & Zhang, Q. (2014). Bacterial diversity in the central black component of Maotai Daqu and its flavor analysis. Annual Review of Microbiology, 64(4), 1659-1669. http://dx.doi.org/10.1007/s13213-014-0809-z.
http://dx.doi.org/10.1007/s13213-014-080...
) screened three aroma producing strains, namely Bacillus subtilis, Bacillus licheniformis and Bacillus amyloliquefaciens. The fermentation products include propionic acid, 1,3-butanediol, acetic acid and methyl ester, which are similar to the flavor of soy sauce. Bacillus is a kind of soy sauce producing bacteria with strong ability to secrete enzymes (Min et al., 2017Min, G., Huang, Y., Qiu, S., Hu, J., Hu, F., & Zhong, F. (2017). Application of high-throughput sequencing in the study of micro ecological diversity of Maotai flavor Baijiu. Chinese Brewing, 36(5), 146-151.), which can metabolize into alcohols, pyrazine, esters and other flavor substances. Chen et al. (2018a)Chen, M., Liu, X., Ping, W., Chen, J., Cai, X., & Luo, A. (2018a). Effect of ester producing functional bacteria on soy sauce flavor fermented grains. Shipin Kexue, 39(10), 199-205. added Bacillus subtilis with ester producing aroma function to the fermented grains. Chromatographic analysis showed that the mass concentration of esters increased while that of alcohols, acids and ketones and aldehydes decreased during accumulation; During fermentation, the mass concentration of esters, alcohols, acids and ketones and aldehydes increased.

4.4 Environmental impact

China is a vast country with abundant resources. Different geographical environments will breed different kinds of microorganisms, which will produce wines with different qualities and styles. Maotai Town in Renhuai, Guizhou Province is the most famous Maotai Baijiu with Maotai flavor. Maotai town is located on the Bank of Chishui River. It is warm in winter, hot in summer, and has less wind and rain. This climate is conducive to the survival of microorganisms. Many microorganisms in Maotai flavor Baijiu come from the air environment. Zhang Yali (Zhang, 2014Zhang, Y. (2014). Identification and Analysis of Air Microbes in Maotai, Renhuai Prefecture, Guizhou Province. Beijing: Beijing University of Chemical Technology.) isolated and cultured the air microorganisms in Maotai Town, and more than 300 kinds of microorganisms were isolated. Wang Li et al. (2021b)Li, W., Jun, L., & Shan, Q., Tang, P., Bi, R., Wang, F., & Li, C. (2021b). Mineral element characteristics analysis and origin identification of Maotai flavor Baijiu in different areas of Chishui River Basin. Chinese Brewing, 40(9), 58-62. found that the types and quantities of mineral elements contained in Maotai flavor Baijiu in different regions (Chishui River Basin) were different.

5 Summary and perspectives

Maotai flavor Baijiu is the most complex Baijiu category among all mainstream flavor Baijiu in China. After years of research and exploration, researchers have obtained many results on the main flavor substance with sensory characteristics of "Maotai flavor", but they have never been verified. Therefore, in the future, we should further enrich the research methods and tools of flavor omics, but we should not unilaterally pursue the results of more detected substances and rely on limited instrument results; At present, the pre-treatment process of flavor substances detection is complex and time-consuming, and whether new physicochemical reactions are initiated remains to be verified. Therefore, the improvement of pretreatment means that the flavor substances in Baijiu are continuously mined through the improvement of the hardware or software of analytical instruments, the analytical data of instruments (flavor compounds) and sensory data (sensory characteristics) are well combined, and the correlation between flavor compounds and sensory characteristics is found by means of big data analysis. While paying attention to the volatile substances in Baijiu, we should also pay attention to the non-volatile substances in Baijiu, such as the contribution of lipopeptide substances to the flavor of Baijiu. In addition, the impact of nonvolatile substances on volatile substances, such as the size of aroma release and the difference in aroma perception, should be the focus of the future research on flavor chemistry of Baijiu. Continue to explore for the realization of "quality clarity" of Maotai flavor Baijiu.

In addition, the sensory flavor characteristics of Baijiu are closely related to the environmental microecology, brewing microecology and flavor metabolites. A variety of different flavor substances exist in the liquor in a specific proportion, which determines the unique flavor characteristics of Baijiu. In the future, we should consider breaking through the research of flavor chemistry of Baijiu from different directions such as environmental microecology, brewing microecology and flavor metabolites. Metagenomics, meta transcriptomics and metabolomics can be used to deeply study the dynamic changes, interactions and metabolites of microorganisms in the brewing environment and fermentation process, so as to explore the metabolic pathways of various microorganisms in the fermentation process, reveal their specific functions and contributions to flavor, and gradually analyze the causes of complex flavor and quality of Baijiu.

Acknowledgements

The author sincerely thanks the Science and Technology Innovation Project of the Chinese Academy of Agricultural Sciences (ASTIP-TRIC07), the Science and Technology Project of Chenzhou Tobacco Company (CZYC2021JS02), the Science and Technology Project of Nanping Tobacco Company (2021350700240083), and the Science and Technology Project of Anhui Wannan Tobacco Leaf Co., Ltd. (2022341800740280) for supporting this research.

  • Practical Application: This review will further reveal the flavor components of Maotai flavor Baijiu, provide inspiration for subsequent research, and provide more in-depth understanding for the study of bitter taste of other foods.

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Publication Dates

  • Publication in this collection
    06 Jan 2023
  • Date of issue
    2023

History

  • Received
    01 Sept 2022
  • Accepted
    23 Oct 2022
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