JP7159492B2

JP7159492B2 - Binary compound fermentation agent and its use

Info

Publication number: JP7159492B2
Application number: JP2021570889A
Authority: JP
Inventors: 信李; 俊紅張; 鵬景崔; 勝虎朱; 玲玲呉; 鋒熊; ▲ウェン▼ 陳; 芸王; 栄松陸; ▲パン▼▲パン▼ 蔡
Original assignee: Jiangsu Hengshun Vinegar Industry Co Ltd
Current assignee: Jiangsu Hengshun Vinegar Industry Co Ltd
Priority date: 2019-12-16
Filing date: 2020-07-22
Publication date: 2022-10-24
Anticipated expiration: 2040-07-22
Also published as: WO2021120618A1; CN110819576B; JP2022526463A; CN110819576A

Description

CGMCC CGMCC 1222512225 CGMCC CGMCC 1222612226 CGMCC CGMCC 1693816938 CGMCC CGMCC 1582415824 CGMCC CGMCC 1780117801 CGMCC CGMCC 1780217802 CGMCC CGMCC 1634516345

本発明は、二元複合発酵剤に関し、さらに食酢の醸造における前記二元複合発酵剤の使用に関する。 The present invention relates to a binary compound fermenting agent, and further to the use of the binary compound fermenting agent in vinegar brewing.

食酢は、デンプンや糖を含有する各種材料、食用アルコールを単独又は混合して、微生物で発酵させた液体酸性調味料である。欧米諸国では、一般的に単一菌種発酵技術を用いて食酢を製造しており、中国や東南アジア諸国では、主に多菌発酵技術を用いて食酢を製造している。単一菌種発酵は主に純粋なアセトバクターを主とし、風味が単一で口当たりが悪い。伝統的な多菌種発酵プロセスは天然多菌種開放発酵を採用することが多く、発酵菌種の由来は前世代から種醪（又は種子液）の形で混入することが多く、すなわち、前世代の酢醪又は酢酸発酵液を一定量で次世代に接種し、このように世代ずつ持続する。伝統的な接種方式は、コア菌種の衰退、微生物叢構造の変化が現れやすく、変化後の菌叢構造の制御と回復が難しく、製品の品質と安定性に巨大な挑戦をもたらす。また、種の量が多く、貯蔵や輸送が容易ではなく、使用条件では多方面の制限があり、それにより、食酢の醸造技術の発展が大きく制限されている。 Vinegar is a liquid acidic seasoning made by fermenting various materials containing starch or sugar and edible alcohol alone or in combination with microorganisms. In Western countries, vinegar is generally produced using a single bacterial species fermentation technology, and in China and Southeast Asian countries, vinegar is mainly produced using a multibacterial fermentation technology. Single-species fermentation mainly consists of pure Acetobacter, which has a single flavor and is not palatable. The traditional multi-seed fermentation process often adopts natural multi-seed open fermentation, and the origin of the fermenting fungi seeds is often mixed in the form of seed mash (or seed liquid) from the previous generation. A certain amount of the vinegar mash or acetic acid fermentation liquid of the generation is inoculated to the next generation, and thus continues generation by generation. The traditional inoculation method is prone to the decline of the core species and changes in the microbiome structure, and the post-change microbiota structure is difficult to control and recover, posing great challenges to the quality and stability of the product. In addition, it contains a large amount of seeds, is not easy to store and transport, and has many limitations in terms of usage conditions, which greatly restricts the development of vinegar brewing technology.

従来技術には、直投式発酵剤及びその使用があり、例えば、直投式発酵剤は、ヨーグルト、生地、キムチなどの発酵食品の製造においてすでに工業化の応用段階に入っているが、伝統的な食酢の醸造に固有の環境及び要件（高酸、高アルコール、低栄養、好気性微生物と嫌気性微生物の共存、上下層の協同発酵など）に適する発酵剤の研究は極めて少ない。 The prior art includes direct-injection fermenting agents and their use, for example, direct-injecting fermenting agents have already entered the application stage of industrialization in the production of fermented foods such as yogurt, dough, and kimchi, but traditional There is very little research on fermenting agents suitable for the specific environment and requirements of vinegar brewing (high acid, high alcohol, low nutrition, coexistence of aerobic and anaerobic microorganisms, cooperative fermentation of upper and lower layers, etc.).

本発明は、使用量が少なく、発酵効率が高く、製品の収率が高く、製品の品質が安定している二元複合発酵剤を提供することを目的とし、食酢の醸造における前記二元複合発酵剤の使用を提供することを別の目的とする。 An object of the present invention is to provide a binary compound fermenting agent that is used in a small amount, has a high fermentation efficiency, a high product yield, and a stable product quality. Another object is to provide the use of leavening agents.

本発明の二元複合発酵剤は、複合菌剤Ｉと複合菌剤ＩＩとを含み、
複合菌剤Ｉは、ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８、バチルス・ソノレンシＣＧＭＣＣ１５８２４、バチルス・コアグランスＣＧＭＣＣ１７８０１、耐酸性中温α－アミラーゼ、プルラナーゼ、グルコアミラーゼ、セルラーゼ、及び酸性プロテアーゼを含み、
複合菌剤ＩＩは、アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５、及びグルコースを含む、を含むことを特徴とする。 The binary composite fermenting agent of the present invention comprises a composite fungicide I and a composite fungicide II,
The compound fungicide I includes Lactobacillus helvetics CGMCC12225, Lactobacillus fermentum CGMCC12226, Lactobacillus acetotolerance CGMCC16938, Bacillus sonorensi CGMCC15824, Bacillus coagulans CGMCC17801, acid-stable medium-temperature α-amylase, pullulanase, glucoamylase, including cellulase and acid protease,
Combined fungal agent II is characterized by comprising Acetobacter pasteurianus CGMCC17802, Gluconacetobacter europeus CGMCC16345, and glucose.

前記ラクトバチルス・ヘルヴェティクス（Ｌａｃｔｏｂａｃｉｌｌｕｓｈｅｌｖｅｔｉｃｕｓ）ＣＧＭＣＣ１２２２５、ラクトバチルス・ファーメンタム（Ｌａｃｔｏｂａｃｉｌｌｕｓｆｅｒｍｅｎｔｕｍ）ＣＧＭＣＣ１２２２６、ラクトバチルス・アセトトレランス（Ｌａｃｔｏｂａｃｉｌｌｕｓａｃｅｔｏｔｏｌｅｒａｎｓ）ＣＧＭＣＣ１６９３８、バチルス・ソノレンシ（Ｂａｃｉｌｌｕｓｓｏｎｏｒｅｎｓｉｓ）ＣＧＭＣＣ１５８２４、バチルス・コアグランス（Ｂａｃｉｌｌｕｓｃｏａｇｕｌａｎｓ）ＣＧＭＣＣ１７８０１、アセトバクター・パスツリアヌス（Ａｃｅｔｏｂａｃｔｅｒｐａｓｔｅｕｒｉａｎｕｓ）ＣＧＭＣＣ１７８０２、グルコナセトバクター・ユーロペウス（Ｋｏｍａｇａｔａｅｉｂａｃｔｅｒｅｕｒｏｐａｅｕｓ）ＣＧＭＣＣ１６３４５は、中国微生物菌種寄託管理委員会普通微生物センターにブタペスト条約に基づく国際寄託がなされ、、寄託場所は北京市朝陽区北辰西路１号中国科学院微生物研究所であり、寄託登録番号はそれぞれＣＧＭＣＣＮｏ．１２２２５、ＣＧＭＣＣＮｏ．１２２２６、ＣＧＭＣＣＮｏ．１６９３８、ＣＧＭＣＣＮｏ．１５８２４、ＣＧＭＣＣＮｏ．１７８０１、ＣＧＭＣＣＮｏ．１７８０２、ＣＧＭＣＣＮｏ．１６３４５であり、寄託日はそれぞれ２０１６年０３月１８日、２０１６年０３月１８日、２０１８年１２月１３日、２０１８年０５月３０日、２０１９年０５月１３日、２０１９年０５月１３日、２０１８年０５月１３日、２０１８年０８月２７日である。前記ラクトバチルス・ヘルヴェティクス（Ｌａｃｔｏｂａｃｉｌｌｕｓｈｅｌｖｅｔｉｃｕｓ）ＣＧＭＣＣ１２２２５、ラクトバチルス・ファーメンタム（Ｌａｃｔｏｂａｃｉｌｌｕｓｆｅｒｍｅｎｔｕｍ）ＣＧＭＣＣ１２２２６、ラクトバチルス・アセトトレランス（Ｌａｃｔｏｂａｃｉｌｌｕｓａｃｅｔｏｔｏｌｅｒａｎｓ）ＣＧＭＣＣ１６９３８、バチルス・ソノレンシ（Ｂａｃｉｌｌｕｓｓｏｎｏｒｅｎｓｉｓ）ＣＧＭＣＣ１５８２４、バチルス・コアグランス（Ｂａｃｉｌｌｕｓｃｏａｇｕｌａｎｓ ) CGMCC17801, Acetobacter pasteurianus CGMCC17802, Gluconacetobacter europaeus (Komagataeibacter europaeus) CGMCC16345 are international deposits based on the Budapest Treaty at the General Microorganism Center of the China Microbial Species Deposit Management Committee. are the Institute of Microbiology, Chinese Academy of Sciences, No. 1 Beichen West Road, Chaoyang District, Beijing, and the deposit registration numbers are CGMCC No. 12225, CGMCC No. 12226, CGMCC No. 16938, CGMCC No. 15824, CGMCC No. 17801, CGMCC No. 17802, CGMCC No. 16345, deposited on March 18, 2016, March 18, 2016, December 13, 2018, May 30, 2018, May 13, 2019, May 13, 2019, respectively; May 13, 2018 and August 27, 2018.

好ましくは、前記複合菌剤Ｉは、ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５１０～２５重量部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６５～１５重量部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８５～２０重量部、バチルス・ソノレンシＣＧＭＣＣ１５８２４３～１５重量部、バチルス・コアグランスＣＧＭＣＣ１７８０１２～１０重量部、耐酸性中温α－アミラーゼ５～１３重量部、プルラナーゼ１～３重量部、グルコアミラーゼ２～５重量部、セルラーゼ３～９重量部、及び酸性プロテアーゼ１～５重量部を含み、
複合菌剤ＩＩは、アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２３５～５５重量部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５２～１０重量部、グルコース４０～６０重量部を含む。 Preferably, the compound fungicide I comprises 10 to 25 parts by weight of Lactobacillus helvetics CGMCC12225, 5 to 15 parts by weight of Lactobacillus fermentum CGMCC12226, 5 to 20 parts by weight of Lactobacillus acetotolerance CGMCC16938, and Bacillus sonorensii. CGMCC15824 3-15 parts by weight, Bacillus coagulans CGMCC17801 2-10 parts by weight, acid-resistant medium-temperature α-amylase 5-13 parts by weight, pullulanase 1-3 parts by weight, glucoamylase 2-5 parts by weight, cellulase 3-9 parts by weight , and 1 to 5 parts by weight of an acid protease,
Compound fungicide II contains 35-55 parts by weight of Acetobacter pasteurianus CGMCC17802, 2-10 parts by weight of Gluconacetobacter europeus CGMCC16345, and 40-60 parts by weight of glucose.

好ましくは、前記複合菌剤Ｉ中、ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８、バチルス・ソノレンシＣＧＭＣＣ１５８２４、及びバチルス・コアグランスＣＧＭＣＣ１７８０１の生菌数がいずれも１．０×１０^８～１．０×１０^１０ＣＦＵ／ｇであり、耐酸性中温α－アミラーゼ、プルラナーゼ、グルコアミラーゼ、セルラーゼ、酸性プロテアーゼの酵素活性がいずれも２～１０万Ｕ／ｇである。 Preferably, the number of viable bacteria of Lactobacillus helvetics CGMCC12225, Lactobacillus fermentum CGMCC12226, Lactobacillus acetotolerance CGMCC16938, Bacillus sonorensi CGMCC15824, and Bacillus coagulans CGMCC17801 in the composite antibacterial agent I is 1. 0×10 ⁸ to 1.0×10 ¹⁰ CFU/g, and the enzymatic activities of acid-resistant mesophilic α-amylase, pullulanase, glucoamylase, cellulase, and acid protease are all 20,000 to 100,000 U/g.

好ましくは、前記複合菌剤ＩＩにおけるアセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２及びグルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５の生菌数が１．０×１０^７～１．０×１０^９ＣＦＵ／ｇである。 Preferably, the viable cell count of Acetobacter pasteurianus CGMCC17802 and Gluconacetobacter europeus CGMCC16345 in the compound antibacterial agent II is 1.0×10 ⁷ to 1.0×10 ⁹ CFU/g.

食酢の醸造における本発明の二元複合発酵剤の使用。 Use of the dual compound fermenting agent of the present invention in vinegar brewing.

好ましくは、前記複合菌剤Ｉは酢酸発酵前に添加され、前記複合菌剤ＩＩは酢酸発酵段階で添加される。 Preferably, the compound fungicide I is added before the acetic acid fermentation, and the compound fungicide II is added during the acetic acid fermentation stage.

好ましくは、前記二元複合発酵剤は種醪と併用されてもよい。 Preferably, the binary compound fermenting agent may be used in combination with seed mash.

好ましくは、前記食酢の醸造は、固体食酢発酵又は液体食酢発酵である。 Preferably, the vinegar brewing is solid vinegar fermentation or liquid vinegar fermentation.

好ましくは、前記二元複合発酵剤は、直投方式で添加される。 Preferably, the binary compound fermenting agent is added by a direct injection method.

好ましくは、前記食酢は、鎮江香酢、山西老陳酢、米酢又はリンゴ酢である。 Preferably, the vinegar is Zhenjiang fragrant vinegar, Shanxi Laochen vinegar, rice vinegar or apple cider vinegar.

研究によると、微生物の多様性は、醸造食酢の風味が豊富で、口当たりがリッチで柔らかいことを実現する重要な基礎となっている。本発明の二元複合発酵剤は、風味の豊富さ、口当たりのリッチさや柔らかさを維持、向上しながら、単一発酵剤で製造された食酢製品の風味の単一性、口当たりの刺激、リッチさや柔らかさの欠如、及び低収率の問題を解決し、菌種混入過程で出現しやすいコア菌種の衰退、微生物叢構造の変化、変化後の菌叢構造の調整と回復が難しく、製品の品質と安定性に悪影響をもたらすなどの従来の問題を回避し、さらに、種の量が多く、貯蔵と輸送が容易ではないという従来の問題を解決する。 Studies have shown that microbial diversity is an important basis for making brewed vinegars rich in flavor and rich and soft in the mouth. The dual compound fermenting agent of the present invention maintains and improves the richness of flavor, the richness and softness of the mouthfeel, while maintaining and improving the uniformity of the flavor of vinegar products produced with a single fermenting agent, and the stimulation and richness of the mouthfeel. Solving the problem of lack of pod softness and low yield, the decline of core fungal species that are likely to appear during the process of contamination, changes in the microflora structure, and difficult adjustment and recovery of the microflora structure after the change. It avoids the conventional problems such as adversely affecting the quality and stability of the seed, and also solves the conventional problem of large amount of seeds and not easy storage and transportation.

本発明の二元複合発酵剤は、食酢の醸造の特殊な環境に適しており、効果が高く、応用場面が広く、従来の種子に混入せずに使用することも、従来の種子に混入して強化発酵剤として使用することもできる。 The dual compound fermenting agent of the present invention is suitable for the special environment of vinegar brewing, is highly effective, and has a wide range of applications. It can also be used as a fortified leavening agent.

本発明の二元発酵剤は、従来のシートに混入せずに使用された、又は従来のように種子に混入して強化発酵剤として使用された場合、発酵効率が速く、収率が高く、品質が安定しており、不揮発性酸、アミノ酸態窒素、エステル類などの風味物質の含有量を著しく高めることができ、製品は、風味が豊富で、口当たりがリッチで柔らかく、香りが際立っており、単一や普通の配合発酵剤は比べ物にならない。 The dual fermenting agent of the present invention has a fast fermentation efficiency and a high yield when used without being mixed with conventional sheets or when mixed with seeds in the conventional manner and used as a fortified fermenting agent. The quality is stable, and the content of non-volatile acid, amino acid nitrogen, esters and other flavor substances can be significantly increased. , Single or ordinary compound fermenters are incomparable.

従来技術と比較して、本発明には次の顕著な利点がある。
（１）本発明の二元複合発酵剤は、固体食酢発酵剤又は液体食酢発酵剤として、単独で又は種醪と組み合わせて使用することができる。
（２）本発明の二元複合発酵剤は、固体食酢発酵に適用すると、不揮発性酸、アミノ酸態窒素、エステル類などの風味物質の含有量を著しく高めることができ、液体食酢発酵に適用すると、不揮発性酸やエステル類風味物質の含有量を著しく高めることができる。 Compared with the prior art, the present invention has the following significant advantages.
(1) The dual compound fermenting agent of the present invention can be used alone or in combination with seed mash as a solid vinegar fermenting agent or a liquid vinegar fermenting agent.
(2) When the dual compound fermenting agent of the present invention is applied to solid vinegar fermentation, it can significantly increase the content of flavor substances such as non-volatile acids, amino acid nitrogen, and esters, and when applied to liquid vinegar fermentation. , the content of non-volatile acid and ester flavor substances can be significantly increased.

本発明のアセトバクター・パスツリアヌスのコロニー形態である。It is the colony morphology of Acetobacter pasteurianus of the present invention.

以下、実施例を参照して本発明の技術案をさらに説明する。 Hereinafter, the technical solution of the present invention will be further described with reference to examples.

本発明の複合菌剤Ｉ及び複合菌剤ＩＩの製造方法は、以下の通りである。各菌をそれぞれ対応する培地で三段階に拡張培養し、発酵させ、発酵終了後、中空繊維膜を用いて発酵液を元の発酵液量の１／５まで濃縮し、その後、無菌脱脂粉乳２０ｇ／１００ｍｌ、グルタミン酸ナトリウム１２ｇ／１００ｍｌ、ソルビトール４ｇ／１００ｍｌを濃縮発酵液と均一に混合し、－８０℃の超低温冷蔵庫に入れて２～５ｈ予備凍結し、最後にサンプルディスクを凍結乾燥機にセットして２４～４８ｈの凍結乾燥処理を行い、平板計数法を用いて生菌数が要件（生菌数はいずれも１．０×１０^８～１．０×１０^１０ＣＦＵ／ｇ）を満たすと検出すると、使用に備えた。活性単位が要件（酵素活性２～１０万Ｕ／ｇ）を満たす市販の関連酵素製剤と食品級グルコースを購入し、クリーンルームで本発明の二元複合発酵剤の配合比で均一に混合し、最後に、真空包装袋に分包し、分包後の製品を４℃で保管した。 The method for producing the compound fungal agent I and the compound fungal agent II of the present invention is as follows. Each bacterium is expanded and cultured in three stages in a corresponding medium, fermented, and after fermentation is completed, the fermented liquid is concentrated to 1/5 of the original volume using a hollow fiber membrane, and then 20 g of sterile skim milk powder. /100ml, sodium glutamate 12g/100ml, sorbitol 4g/100ml and the concentrated fermented liquid are uniformly mixed, placed in an ultra-low temperature refrigerator at -80°C and pre-frozen for 2-5 hours, and finally the sample disk is placed in the freeze dryer. Freeze-dry for 24 to 48 hours, and use the plate count method to detect if the viable count satisfies the requirements (the viable count is 1.0×10 ⁸ to 1.0×10 ¹⁰ CFU/g in any case). Then it was ready for use. Purchasing a commercially available related enzyme preparation and food-grade glucose whose activity unit meets the requirements (enzyme activity 20,000 to 100,000 U/g), uniformly mixing in a clean room at the blending ratio of the binary compound fermenting agent of the present invention, and finally Then, it was divided into vacuum packaging bags, and the divided and packaged product was stored at 4°C.

実施例では、各指標の測定方法は次の通りである。総酸と不揮発酸は「ＧＢ１８１８７－２０００」の方法を参照して測定し、総酸は酢酸として、不揮発酸は乳酸として測定し、アミノ酸態窒素は「ＧＢ１８１８６－２０００」方法を参照して測定し、総エステルは「ＧＢ／Ｔ１９７７７－２０１３」の方法を参照して測定した。 In the examples, the method for measuring each index is as follows. Total acid and nonvolatile acid are measured with reference to the method of "GB18187-2000", total acid is measured as acetic acid, nonvolatile acid is measured as lactic acid, and amino acid nitrogen is measured with reference to the "GB18186-2000" method. , Total ester was measured by referring to the method of "GB/T19777-2013".

本発明の二元複合発酵剤は、下記の食酢の醸造だけでなく、他の固体及び液体発酵食酢の醸造にも適用することができる。 The dual compound fermenting agent of the present invention can be applied not only to the following vinegar brewing but also to the brewing of other solid and liquid fermented vinegars.

実施例１：アセトバクター・パスツリアヌス菌株の分離及び同定
本実施例において、炭酸カルシウム、グルコース、寒天粉、無水エタノール、酢酸及び水酸化ナトリウムはいずれも国薬集団化学試薬有限公司から購入し、酵母エキスは英国ＯＸＯＩＤ社から購入した。 Example 1: Isolation and identification of Acetobacter pasteurianus strain was purchased from OXOID, UK.

１、菌株分離
鎮江香酢の醪サンプル１０ｇを採取し、滅菌された生理食塩水９０ｍｌに加え、シェーカーで均一に振った後、１００μｌのサンプルを採取し、９００μｌの生理食塩水に加え、ボルテック振とう器で均一に混合し、その後、勾配希釈を行った。均一に混合して、炭酸カルシウム２０ｇを添加した固体培地（１Ｌ当たりグルコース２０ｇ、酵母エキス１０ｇ、寒天粉末１５ｇを含有、１２１℃で２０min殺菌後冷却し、３％エタノールを添加）に順次塗布し、３０℃で３日間培養した。平板上に透明ループがあるかどうかを観察し、該当する菌株を選別した。 1. Strain isolation Collect 10 g of mash sample of Zhenjiang fragrant vinegar, add to 90 ml of sterilized physiological saline, shake evenly with a shaker, then collect 100 μl of sample, add to 900 μl of physiological saline, and vortex shake. Mix uniformly with a toaster, followed by a gradient dilution. Uniformly mixed, solid medium containing 20 g of calcium carbonate (20 g of glucose, 10 g of yeast extract, 15 g of agar powder per 1 L, sterilized at 121 ° C. for 20 minutes, cooled, added with 3% ethanol), Cultured at 30° C. for 3 days. The presence of transparent loops on the plate was observed to select the relevant strains.

２、菌株の二次スクリーニング
一次スクリーニングした菌株を、二次スクリーニング用固体平板（グルコース２０ｇ、酵母エキス１０ｇ、酢酸３０ｍｌ、及びエタノール３０ｍｌ、寒天粉末１５ｇ／Ｌ添加）に接種し、３０℃で３日間培養した後、透明ループが最大となる菌株を選別した。 2. Secondary screening of strains The primary screened strains were inoculated on a solid plate for secondary screening (glucose 20 g, yeast extract 10 g, acetic acid 30 ml, ethanol 30 ml, agar powder 15 g / L added), and kept at 30 ° C. for 3 days. After culturing, the strain with the largest clear loop was selected.

スクリーニングした菌株を、二次スクリーニング用液体培地（グルコース２０ｇ、酵母エキス１０ｇ、酢酸３０ｍｌ、エタノール５０ｍｌ、及び蒸留水１Ｌ）に接種し、３０℃、２００ｒｐｍの回転数で２０ｈ培養し、水酸化ナトリウムにより総酸（酢酸換算）量を滴定し、２４ｈおきに測定し、酸産生の変化を記録した。 The screened strains were inoculated into a liquid medium for secondary screening (20 g of glucose, 10 g of yeast extract, 30 ml of acetic acid, 50 ml of ethanol, and 1 L of distilled water), cultured at 30° C. and 200 rpm for 20 h, and added with sodium hydroxide. Total acid (in terms of acetic acid) was titrated and measured every 24 h to record changes in acid production.

２回の二次スクリーニングを経たところ、最終的に酸産生性の優れたアセトバクター・パスツリアヌスＨＳＣＹ１０８５菌株が得られ、そのコロニーの形態は図１に示される。 After two rounds of secondary screening, an Acetobacter pasteurianus HSCY1085 strain with excellent acid production was finally obtained, and its colony morphology is shown in FIG.

３、菌株の同定
測定された１６ＳｒＤＮＡ配列をＮＣＢＩデータベースで比較分析し、生理学的・生化学的特徴を組み合わせて、本発明の菌株をアセトバクター・パスツリアヌスＨＳＣＹ１０８５（Ａｃｅｔｏｂａｃｔｅｒｐａｓｔｅｕｒｉａｎｕｓ）と命名し、１６ＳｒＤＮＡ配列をＳＥＱＩＤＮｏ．１に示す。 3, Identification of the strain Comparative analysis of the measured 16S rDNA sequences in the NCBI database, combined physiological and biochemical characteristics, named the strain of the present invention Acetobacter pasteurianus HSCY1085 (Acetobacter pasteurianus), the 16S rDNA sequence SEQ ID No. 1.

この菌株は中国微生物菌種寄託管理委員会普通微生物センターに寄託され、寄託場所は北京市朝陽区北辰西路１号院３号中国科学院微生物研究所であり、寄託日は２０１９年５月１３日であり、寄託番号はＣＧＭＣＣＮｏ．１７８０２であり、その分類命名はＡｃｅｔｏｂａｃｔｅｒｐａｓｔｅｕｒｉａｎｕｓである。 This strain was deposited at the General Microbiology Center of the Chinese Microbial Species Deposit Management Committee, at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, No. 1, Beichen West Road, Chaoyang District, Beijing, and the date of deposit was May 13, 2019. and the deposit number is CGMCC No. 17802 and its taxonomic nomenclature is Acetobacter pasteurianus.

実施例２：鎮江香酢の醸造における使用
本実施例は、従来の種醪及び接種方式を用いずに、本発明の二元複合発酵剤のみを鎮江香酢の醸造に適用することを提供する。 Example 2: Use in brewing Zhenjiang fragrant vinegar This example provides that only the dual compound fermenting agent of the present invention is applied to brewing Zhenjiang fragrant vinegar without using conventional seed mash and inoculation method. .

１、試験群
（１）４００ｋｇのかめを９個準備し、もち米５０ｋｇに水を加えて一晩浸した。もち米を蒸気で蒸し、約４０℃の温度になるまで冷水をかけて酒薬０．３ｋｇを加え、よく混ぜた後、かめに入れてラッパ状の穴を作った。 1. Test Group (1) Nine jars of 400 kg were prepared, and 50 kg of glutinous rice was added with water and soaked overnight. The glutinous rice was steamed and then poured with cold water until it reached a temperature of about 40°C.

（２）穴の中に一定の酒液が出てきたら、かめごとに麦麹２．５ｋｇを加え、次に水１５０ｋｇを加えてよく混ぜた。 (2) When a certain amount of sake liquid appeared in the hole, 2.5 kg of barley koji was added to each pot, and then 150 kg of water was added and mixed well.

（３）アルコール発酵過程において、定期的に撹拌し、温度を３０℃程度に制御し、約５～７ｄ程度発酵させると終了した。 (3) During the alcohol fermentation process, the mixture was stirred periodically, the temperature was controlled at about 30°C, and fermentation was completed for about 5-7 days.

（４）上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、バチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温した。 (4) 200 kg of sake mash was added to each of the three jars of 400 kg in step (3) above, and the compound fungicide agent I of the present invention (25 parts of Lactobacillus helvetics CGMCC12225, 15 parts of Lactobacillus fermentum CGMCC12226) was added. , Lactobacillus acetotolerance CGMCC 16938 20 parts, Bacillus sonorensii CGMCC 15824 15 parts, Bacillus coagulans CGMCC 17801 10 parts, acid-stable medium-temperature α-amylase 13 parts, pullulanase 3 parts, glucoamylase 5 parts, cellulase 9 parts, and acid protease 5 parts Part) was added in an amount of 0.15‰ (w / w) of the mass of sake mash, mixed well, then 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar). . The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash. Add about 1000g of sake moromi (30°C) to the small tub and mix well. Sprinkle over the top of the vinegared moromi in each pot. Put your hand about 10 cm deep from the surface and mix well. Finally, add rice bran to the top. to keep warm.

（５）鎮江香酢の醸造プロセスに従って層ごとに醪を切り返し、総酸がこれ以上増加しなくなるまで酢醪を発酵させると、発酵を終了した。塩を加えて醪を１５ｄ密閉し、次に、炒めた米を加えて酢を抽出した。酢を煎じ、熟成し、殺菌して充填すれば、完成品が得られた。 (5) According to the brewing process of Zhenjiang fragrant vinegar, the mash was cut layer by layer, and the vinegar mash was fermented until the total acidity did not increase any more, and the fermentation was finished. Salt was added to seal the mash for 15d, then the fried rice was added to extract the vinegar. Once the vinegar was decoctioned, aged, pasteurized and filled, the finished product was obtained.

２、比較試験
（１）対照群１
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇ、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。かめごとに市販の光明牌醸造酢酢酸菌剤を、酒もろみの質量の０．５‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温し、次のステップは上記のステップ（５）と同じであった。 2, Comparative test (1) Control group 1
200 kg of sake mash, 80 kg of wheat bran and 45 kg of rice bran were added to each of three of the 400 kg jars of step (3), and the sake mash and grains were uniformly mixed (vinegar mash). Commercially available Komyohai brewed vinegar acetic acid bacterium agent was added to each tortoise in an amount of 0.5‰ (w/w) of the mass of sake moromi, and about 1000 g of sake moromi at 30 ° C was added to a small bucket and mixed well. After that, sprinkle it on the top of the vinegar moromi, put your hand about 10 cm deep from the surface and mix well, finally cover the top with rice bran and keep it warm, and the next step was the same as step (5) above.

（２）対照群２
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇ、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。伝統的な種醪２６ｋｇを酢醪の上部に取り、米ぬかを被せて保温し、次のステップは上記のステップ（５）と同じであった。 (2) Control group 2
200 kg of sake mash, 80 kg of wheat bran and 45 kg of rice bran were added to each of three of the 400 kg jars of step (3), and the sake mash and grains were uniformly mixed (vinegar mash). 26 kg of traditional seed mash was taken on top of the vinegar mash, covered with rice bran and kept warm, the next step was the same as step (5) above.

対照群１と比較して、本発明の二元複合発酵剤を使用した試験群は、昇温速度が速く（初回温度が４０℃以上に達するための時間が３８ｈから２０ｈに１８ｈ短縮された）、発酵時間は２２ｄから１５ｄに７ｄ短縮され、醪密閉が終了した後、基材醪中、総酸含有量は１４．７３％、不揮発酸含有量は８２．８４％、アミノ酸態窒素含有量は１０５．５６％、総エステル含有量は１２６．０５％向上し、製品の収率は１７．０１％向上し、得られた製品は、香りがはっきりし、風味が豊かで、口当たりが柔らかくリッチであり、官能スコアが高かった。 Compared to the control group 1, the test group using the dual compound fermenting agent of the present invention has a faster temperature rise rate (the time for the initial temperature to reach 40 ° C. or higher was shortened from 38 hours to 20 hours by 18 hours). , the fermentation time was shortened from 22d to 15d by 7d, and after sealing the mash, the total acid content was 14.73%, the non-volatile acid content was 82.84%, and the amino acid nitrogen content was 105.56%, the total ester content was improved by 126.05%, the product yield was improved by 17.01%, and the resulting product was clear in aroma, rich in flavor, soft and rich in mouthfeel. Yes, and the sensory score was high.

対照群２と比較して、本発明の二元複合発酵剤を使用した試験群は、発酵時間が１９ｄから１５ｄに４ｄ短縮され、醪密閉が終了した後、基材醪中、総酸含有量は１０．２２％、不揮発性酸含有量は２１．８９％、アミノ酸態窒素含有量は７６．１９％、総エステル含有量は８４．７９％向上し、製品の収率は１１．９５％向上し、得られた製品は、全体として風味や口当たりについて鎮江香酢の特徴を有し、香りがよりはっきりし、口当たりがより柔らかく、より豊かで、よりリッチであった。 Compared to the control group 2, the test group using the dual compound fermenting agent of the present invention shortened the fermentation time from 19d to 15d by 4d, and after the mash was sealed, the total acid content in the base mash was is 10.22%, the non-volatile acid content is 21.89%, the amino acid nitrogen content is 76.19%, the total ester content is 84.79%, and the product yield is 11.95%. However, the resulting product as a whole has the characteristics of Zhenjiang fragrant vinegar in terms of flavor and mouthfeel, with a clearer aroma and a softer, richer and richer mouthfeel.

実施例３：鎮江香酢の醸造における強化使用
本実施例は、伝統的な種醪及び接種方式を保持しながら、本発明の二元複合発酵剤を強化のために鎮江香酢の醸造に使用することを提供する。 Example 3: Enhanced use in brewing Zhenjiang fragrant vinegar This example uses the dual compound fermenting agent of the present invention to enhance Zhenjiang fragrant vinegar while retaining the traditional seed mash and inoculation method offer to do.

（３）アルコール発酵過程において、定期的に撹拌し、温度を３０℃程度に制御し、約５～７ｄ発酵させると終了した。 (3) During the alcohol fermentation process, the mixture was stirred periodically, the temperature was controlled at about 30°C, and fermentation was completed for about 5-7 days.

（４）上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５１０部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６５部、ラクトバチルス・アセトトレランス菌ＣＧＭＣＣ１６９３８５部、バチルス・ソノレンシ菌ＣＧＭＣＣ１５８２４３部、バチルス・コアグランスＣＧＭＣＣ１７８０１２部、耐酸性中温α－アミラーゼ５部、プルラナーゼ１部、グルコアミラーゼ２部、セルラーゼ３部、及び酸性プロテアーゼ１部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。種醪２６ｋｇを酢醪の上部に取り、本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２３５部、かめごとにグルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５２部、及びグルコース４０部）を、酒もろみの質量の０．０５‰（ｗ／ｗ）の添加量で加え、種醪に均一に混ぜて米ぬかをかけて保温した。 (4) 200 kg of sake mash was added to each of the three jars of 400 kg in step (3) above, and the compound fungicide I of the present invention (10 parts of Lactobacillus helvetics CGMCC12225, 5 parts of Lactobacillus fermentum CGMCC12226) , Lactobacillus acetotolerance CGMCC16938 5 parts, Bacillus sonorensi CGMCC15824 3 parts, Bacillus coagulans CGMCC17801 2 parts, acid-stable medium-temperature α-amylase 5 parts, pullulanase 1 part, glucoamylase 2 parts, cellulase 3 parts, and acidic Protease 1 part) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash, and after mixing well, 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar Moromi). Take 26 kg of the seed mash on the top of the vinegar mash, add the compound fungicide II of the present invention (35 parts of Acetobacter pasteurianus CGMCC17802, 2 parts of Gluconacetobacter europeus CGMCC16345 and 40 parts of glucose per jar) to the mass of sake mash. was added in an amount of 0.05‰ (w/w), mixed uniformly with the seed mash, covered with rice bran and kept warm.

（５）鎮江香酢の醸造プロセスに従って層ごとに切り返し、総酸がこれ以上増加しなくなるまで発酵させると、発酵を終了した。塩を加えて醪を１５ｄ密閉し、炒めた米を加えて酢を抽出した。酢を煎じ、熟成し、殺菌して充填すれば、完成品が得られた。 (5) According to the brewing process of Zhenjiang fragrant vinegar, cut layer by layer, fermented until the total acid no longer increased, and finished the fermentation. Salt was added and the mash was sealed for 15d, and the fried rice was added to extract the vinegar. Once the vinegar was decoctioned, aged, pasteurized and filled, the finished product was obtained.

２、比較試験
（１）対照群１
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇ、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。種醪２６ｋｇを酢醪の上部に取り、米ぬかを被せて保温し、次のステップは上記のステップ（５）と同じであった。 2, Comparative test (1) Control group 1
200 kg of sake mash, 80 kg of wheat bran and 45 kg of rice bran were added to each of three of the 400 kg jars of step (3), and the sake mash and grains were uniformly mixed (vinegar mash). 26 kg of the seed mash was taken on top of the vinegar mash, covered with rice bran and kept warm, the next step was the same as step (5) above.

（２）対照群２
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇ、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。種醪２６ｋｇを酢醪の上部に取り、かめごとに市販の光明牌醸造酢酢酸菌剤を種醪に均一に混ぜた後、米ぬかを被せて保温し、添加量は酒もろみの質量の０．３‰（ｗ／ｗ）とし、次のステップは上記のステップ（５）と同じであった。 (2) Control group 2
200 kg of sake mash, 80 kg of wheat bran and 45 kg of rice bran were added to each of three of the 400 kg jars of step (3), and the sake mash and grains were uniformly mixed (vinegar mash). Place 26 kg of the seed mash on top of the vinegar mash, mix the commercially available Komyokai brewing vinegar acetic acid bacteria agent with the seed mash evenly for each pot, cover with rice bran and keep warm. 3‰ (w/w) and the next step was the same as step (5) above.

対照群１と比較して、本発明の二元複合発酵剤を用いて強化した試験群は、発酵時間が１９ｄから１３ｄに６ｄ短縮され、醪密閉が終了した後、基材醪中、総酸含有量は１１．８０％、不揮発性酸含有量は２７．３６％、アミノ酸態窒素含有量は８５．７１％、総エステル含有量は９１．２５％向上し、製品の収率は１４．１５％向上し、得られた製品は、香りがよりはっきりし、口当たりの柔らかさがより強く、感官スコアがより高かった。 Compared to the control group 1, the test group reinforced using the dual compound fermenting agent of the present invention shortened the fermentation time from 19d to 13d by 6d, and after the mash was sealed, the total acid in the base mash was The content is 11.80%, the non-volatile acid content is 27.36%, the amino acid nitrogen content is 85.71%, the total ester content is 91.25%, and the product yield is 14.15%. % improvement and the resulting product had a more pronounced aroma, a stronger mouthfeel and a higher sensory score.

対照群２と比較して、本発明の二元複合発酵剤を用いて強化した試験群は、発酵時間が１８ｄから１３ｄに５ｄ短縮され、醪密閉が終了した後、基材醪中、総酸含有量は８．５２％、不揮発性酸含有量は１８．５２％、アミノ酸態窒素含有量は６９．５７％、総エステル含有量は８４．２５％向上し、製品の収率は１０．０３％向上し、得られた製品は、香りがはっきりし、口当たりが柔らかで、感官スコアが高かった。 Compared to the control group 2, the test group reinforced with the dual compound fermenting agent of the present invention shortened the fermentation time by 5d from 18d to 13d, and after the mash was sealed, the total acid in the base mash was The content is 8.52%, the non-volatile acid content is 18.52%, the amino acid nitrogen content is 69.57%, the total ester content is improved by 84.25%, and the product yield is 10.03%. % improvement and the resulting product had a pronounced aroma, a soft mouthfeel and a high sensory score.

実施例４：山西老陳酢の醸造における使用
本実施例は、従来の種子醪及び播種方式を用いずに、本発明の複合発酵剤のみを山西老陳酢の醸造に適用することを提供する。 Example 4: Use in Brewing Shanxi Laochen Vinegar This example provides the application of only the compound fermenting agent of the present invention in brewing Shanxi Laochen Vinegar without the traditional seed mash and seeding method. .

１、試験群
（１）コウリャンを５～１０部分に粉砕した後、ぬるま湯を加え、１～２ｈ煮込んだ後、コウリャン１００ｋｇ当たり６２．５ｋｇの大麹粉を加えて均一に混合した。 1. Test group (1) After crushing sorghum into 5-10 pieces, lukewarm water was added and the mixture was boiled for 1-2 hours.

（２）上記原料を６つのかめに入れてアルコール発酵を行い、かめに入れた初期温度を約２５℃とし、後期では１８～２５℃に保持した。最初の４日間は開放発酵し、後の８日間は密閉発酵し、約１２～１５日間で終了した。 (2) The above raw materials were put into six jars and subjected to alcoholic fermentation. The first four days were open fermentation, the last eight days were closed fermentation, and were completed in about 12-15 days.

（３）上記ステップ（２）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２０部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６８部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１６９３８１５部、ラクトバチルス・ソノレンシＣＧＭＣＣ１５８２４７部、ラクトバチルス・コアグランスＣＧＭＣＣ１７８０１６部、耐酸性中温α－アミラーゼ１０部、プルラナーゼ１．５部、グルコアミラーゼ３部、セルラーゼ６部、及び酸性プロテアーゼ２部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま９０ｋｇ、穀糠１００ｋｇを加え、酒もろみと穀物を均一に混合した。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２４０部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５８部、及びグルコース４５部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約２０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布してよく混ぜ、山西老陳酢のプロセスに従って醪を切り返した。 (3) 200 kg of sake mash was added to each of the three jars of 400 kg in step (2) above, and the compound fungicide I of the present invention (20 parts of Lactobacillus helvetics CGMCC12225, 8 parts of Lactobacillus fermentum CGMCC12226) was added. , Lactobacillus fermentum CGMCC16938 15 parts, Lactobacillus sonorensi CGMCC15824 7 parts, Lactobacillus coagulans CGMCC17801 6 parts, acid-stable medium temperature α-amylase 10 parts, pullulanase 1.5 parts, glucoamylase 3 parts, cellulase 6 parts, and 2 parts of acidic protease) are added in an amount of 0.15‰ (w/w) of the mass of sake mash, mixed well, then 90 kg of bran and 100 kg of bran are added, and the sake mash and grains are uniformly mixed. did. The compound fungicide II of the present invention (40 parts of Acetobacter pasteurianus CGMCC17802, 8 parts of Gluconacetobacter europeus CGMCC16345, and 45 parts of glucose) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash. In addition, about 2000 g of sake moromi at 30° C. was added to the small tub and mixed well, then sprinkled on the top of the vinegar mash in each pot and mixed well, and the mash was cut back according to the process of Shanxi Laozhen vinegar.

（４）総酸がこれ以上増加しなくなるまで酢醪を発酵させると、発酵を終了した。塩を加えて醪を１０ｄ密閉し、その後、燻製、酢抽出をして、最後に酢を煎じ、熟成し、殺菌して充填すれば、完成品が得られた。 (4) Fermentation was terminated when the vinegar mash was fermented until the total acidity did not increase any more. Salt is added and the mash is sealed for 10d, then smoked, vinegar extracted, finally vinegared, aged, sterilized and filled to obtain the finished product.

２、比較試験
（１）対照群
上記ステップ（２）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇ、ふすま９０ｋｇ、穀糠１００ｋｇを加え、酒もろみと穀粒を均一に混合した。市販の光明牌醸造酢酢酸菌剤４０ｇを、酒もろみの質量の０．５‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約２０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布してよく混ぜ、山西老陳酢のプロセスに従って切り返しを行い、次のステップは上記のステップ（４）と同じであった。 2. Comparative test (1) Control group 200 kg of sake mash, 90 kg of bran and 100 kg of rice bran were added to each of 3 of the 400 kg jars of step (2) above, and the sake mash and grains were uniformly mixed. Add 40 g of commercially available Komyochai brewing vinegar acetic acid bacteria agent in an amount of 0.5 ‰ (w / w) of the mass of sake moromi, add about 2000 g of sake moromi at 30 ° C to a small bucket and mix well, Sprinkle on the top of the vinegar mash in each pot and mix well, then cut back according to the process of Shanxi Lao Chen Vinegar, the next step is the same as step (4) above.

対照群と比較して、本発明の二元複合発酵剤を用いて強化した試験群は、発酵時間が９ｄから７ｄに２ｄ短縮され、燻製終了後、酢醪中、総酸含有量は１８．２６％、不揮発酸含有量は５９．１８％、アミノ酸態窒素含有量は９４．４４％、総エステル含有量は４５．４８％向上し、製品の収率は１９．３５％向上し、得られた製品は、風味が豊富で、口当たりが柔らかでリッチであり、官能スコアが高かった。 Compared with the control group, the test group reinforced with the dual compound fermenting agent of the present invention had a fermentation time shortened from 9d to 7d by 2d, and the total acid content in the vinegar mash after smoking was 18.5d. 26%, the non-volatile acid content is 59.18%, the amino acid nitrogen content is 94.44%, the total ester content is 45.48%, the yield of the product is 19.35%, the obtained The products were rich in flavor, soft and rich in mouthfeel, and had high sensory scores.

実施例５：米酢の醸造における使用
本実施例は、米酢の醸造における本発明の二元複合発酵剤の使用を提供する。 Example 5 Use in Brewing Rice Vinegar This example provides the use of the dual compound fermenting agent of the present invention in brewing rice vinegar.

１、試験群
（１）粒がリッチでいて、かびのないもち米を選んで、ほこりを除去した後、粉砕機でもち米を粉砕して７０～８０メッシュの微粉末にした。粉砕したもち米粉と水とを、もち米粉：水＝１：５（ｗ／ｗ）の割合で糊化槽に入れた。高温α－アミラーゼ２．５万Ｕ／ｍｌをもち米１トン当たり０．４～０．５Ｌ加えた。同時に９０～９５℃に昇温し、３０～４０min保温した。 1. Test group (1) Select glutinous rice with rich grains and no mold, remove dust, and grind the glutinous rice into fine powder of 70 to 80 mesh with a grinder. The pulverized glutinous rice flour and water were placed in a gelatinization tank at a ratio of glutinous rice flour:water=1:5 (w/w). 0.4 to 0.5 L of hot α-amylase 25,000 U/ml was added per ton of glutinous rice. At the same time, the temperature was raised to 90 to 95°C and maintained for 30 to 40 minutes.

（２）もろみをスクリュー冷却器で５５～６０℃に降温しながら糖化槽に移した。グルコアミラーゼ１０万Ｕ／ｇを、もち米１トン当たり４０～５０ｇで加えた。温度を５５～６０℃に制御し、３０～６０min保温した。保温終了後、スクリュー式冷却器を用いて２５～３５℃に降温しながら、アルコール発酵槽に移した。 (2) The mash was transferred to a saccharification tank while the temperature was lowered to 55-60°C by a screw cooler. Glucoamylase 100,000 U/g was added at 40-50 g per ton of glutinous rice. The temperature was controlled at 55-60°C and maintained for 30-60 minutes. After the end of the heat retention, the mixture was transferred to an alcohol fermentation tank while the temperature was lowered to 25 to 35°C using a screw cooler.

（３）サッカロミセス・セレビシエ（ＣＩＣＣから購入、菌株番号ＣＩＣＣ１００１）を５％（ｖ／ｖ）接種量で添加した。温度を２５～３５℃に制御し、アルコール度数５％ｖｏｌ程度に発酵させた。次に、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２０部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１０部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８９部、ラクトバチルス・ソノレンシＣＧＭＣＣ１５８２４５部、ラクトバチルス・コアグランスＣＧＭＣＣ１７８０１６部、耐酸性中温α－アミラーゼ８部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ６部、及び酸性プロテアーゼ２部）を、酒もろみ量の０．２‰（ｗ／ｗ）の添加量で加え、アルコール度８％ｖｏｌまで発酵を続けた。 (3) Saccharomyces cerevisiae (purchased from CICC, strain number CICC1001) was added at 5% (v/v) inoculum. The temperature was controlled at 25 to 35° C., and fermentation was carried out to about 5% vol of alcohol. Next, the compound fungicide I of the present invention (20 parts of Lactobacillus helvetics CGMCC12225, 10 parts of Lactobacillus fermentum CGMCC12226, 9 parts of Lactobacillus acetotolerance CGMCC16938, 5 parts of Lactobacillus sonorensi CGMCC15824, Lactobacillus sonorensi CGMCC15824, 6 parts of coagulans CGMCC 17801, 8 parts of acid-resistant mesophilic α-amylase, 3 parts of pullulanase, 5 parts of glucoamylase, 6 parts of cellulase, and 2 parts of acid protease) were added in an amount of 0.2‰ (w/w) of sake mash. The fermentation was continued until the alcohol content was 8% vol.

（４）上記ステップ（３）の５００Ｌ発酵槽を１つ選び、フィルタープレスで濾過した清澄酒液２００Ｌを加え、本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＰＧＭＣＣ１７８０２３５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース５５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、通気量０．３～０．４ｖｖｍ、撹拌速度２５０ｒ／ｍｉｎに調整して酢酸発酵を行った。 (4) Select one of the 500 L fermenters in step (3) above, add 200 L of clarified liquor filtered by a filter press, and add 35 parts of the compound fungicide II of the present invention (Acetobacter pasteurianus PGMCC17802, Gluconacetobacter europeus) 10 parts of CGMCC16345 and 55 parts of glucose) was added in an amount of 0.15‰ (w/w) of the mass of sake mash, and the aeration rate was adjusted to 0.3 to 0.4 vvm and the stirring speed was adjusted to 250 r/min. acetic acid fermentation was performed.

（５）発酵が終了した酢液を０．０１μｍセラミック膜でろ過した後、酸度を調整し、充填すれば、完成品が得られた。 (5) After the fermentation was completed, the vinegar solution was filtered through a 0.01 μm ceramic membrane, adjusted for acidity, and filled to obtain a finished product.

２、比較試験
（１）対照群
上記ステップ（３）の５００Ｌ発酵槽を１つ選び、フィルタープレスで濾過した清澄酒液２００Ｌを加え、市販の光明板醸造酢酢酸菌剤を、酒もろみの質量の０．７‰（ｗ／ｗ）の添加量で加え、通気量０．３～０．４ｖｖｍ、撹拌速度２５０ｒ／ｍｉｎに調整して酢酸発酵を行い、次のステップは上記ステップ（５）と同じであった。 2. Comparative test (1) Control group Select one of the 500L fermenters in step (3) above, add 200L of clarified sake filtered by a filter press, add a commercially available light plate brewing vinegar acetic acid bacteria agent, and add the mass of sake mash. 0.7 ‰ (w / w) of the acetic acid fermentation is performed by adjusting the aeration rate to 0.3 to 0.4 vvm and the stirring speed to 250 r / min, and the next step is the above step (5). was the same.

対照群と比較して、試験群は、開始速度が速く、発酵周期が２３．７５％短縮され、総酸含有量は１１．５２％、不揮発酸含有量は１２．５倍以上、総エステル含有量は１４倍以上向上した。試験群で製造したリンゴ酢は、刺激性が著しく低下し、口当たりが柔らかく、顕著な総合的な香りがあり、品質が著しく向上した。 Compared with the control group, the test group has a faster initiation rate, a 23.75% shorter fermentation cycle, a total acid content of 11.52%, a non-volatile acid content of more than 12.5 times, and a total ester content. Volume improved more than 14 times. The apple cider vinegar produced in the test group was significantly less pungent, had a softer mouthfeel, a pronounced overall aroma, and was significantly improved in quality.

実施例６：リンゴ酢の醸造における使用
本実施例は、リンゴ酢の醸造における本発明の複合発酵剤の使用を提供する。 Example 6 Use in Brewing Apple Cider Vinegar This example provides the use of the compound fermenting agent of the present invention in brewing apple cider vinegar.

１、試験群
（１）２つの５００Ｌ発酵槽に、糖含有量１６～１８％（ｗ／ｗ）、ｐＨ４．５～５．０に調整したリンゴジュースをそれぞれ２００Ｌ添加し、酵母菌（ＣＩＣＣから購入、菌株寄託番号ＣＩＣＣ１００１）を５％（ｖ／ｖ）で接種し、２８～３０℃でアルコール度約４％ｖｏｌまで発酵させた。 1. Test group (1) Add 200 L each of apple juice adjusted to a sugar content of 16-18% (w/w) and a pH of 4.5-5.0 to two 500 L fermenters, yeast (from CICC purchased, strain deposit number CICC1001) was inoculated at 5% (v/v) and fermented at 28-30°C to about 4% vol of alcohol.

（２）上記ステップ（１）の発酵槽を１つ選び、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５１５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８１０部、ラクトバチルス・ソノレンシＣＧＭＣＣ１５８２４３部、ラクトバチルス・コアグランスＣＧＭＣＣ１７８０１２部、耐酸性中温α－アミラーゼ５部、プルラナーゼ１部、グルコアミラーゼ２部、セルラーゼ８部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、発酵槽温度を３０℃、圧力を０．０５ＭＰａ、撹拌速度を６０ｒ／ｍｉｎとし、アルコール度約７％ｖｏｌまで発酵させた。 (2) Select one of the fermenters of step (1) above, and use the compound fungicide I of the present invention (Lactobacillus helvetics CGMCC12225 15 parts, Lactobacillus fermentum CGMCC12226 15 parts, Lactobacillus acetotolerance CGMCC16938 10 part, Lactobacillus sonorensi CGMCC15824 3 parts, Lactobacillus coagulans CGMCC17801 2 parts, acid-stable medium-temperature α-amylase 5 parts, pullulanase 1 part, glucoamylase 2 parts, cellulase 8 parts, and acid protease 5 parts), sake mash was added in an amount of 0.15‰ (w/w) of the mass of , and fermentation was performed with a fermenter temperature of 30°C, a pressure of 0.05 MPa, and a stirring speed of 60 r/min until the alcohol content reached about 7% vol.

（３）発酵槽に本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５０部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５８部、及びグルコース４５部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加えた。 (3) The compound fungicide II of the present invention (50 parts of Acetobacter pasteurianus CGMCC17802, 8 parts of Gluconacetobacter europeus CGMCC16345, and 45 parts of glucose) was added to the fermenter at 0.1‰ (w/ w) was added in the amount to be added.

（４）発酵が終了した酢液をセラミック膜でろ過した後、酸度を調整し、充填すれば、完成品が得られた。 (4) The vinegar solution after fermentation was filtered through a ceramic membrane, the acidity was adjusted, and the product was filled to obtain a finished product.

２、比較試験
（１）対照群
上記ステップ（１）の発酵槽を１つ選び、発酵槽の温度を３０℃、圧力を０．０５Ｍｐａ、撹拌速度を６０ｒ／ｍｉｎに維持し、アルコール度約７％ｖｏｌまで発酵させ、次に、市販の光明牌醸造酢酢酸菌剤を、酒もろみの質量の０．５‰（ｗ／ｗ）の添加量で加え、通気量を０．３～０．４ｖｖｍ、撹拌速度を２５０ｒ／ｍｉｎに調整して酢酸発酵を行い、次のステップは上記ステップ（４）と同じであった。 2. Comparative test (1) Control group Select one fermenter from step (1) above, maintain the temperature of the fermenter at 30°C, the pressure at 0.05Mpa, the stirring speed at 60r/min, and the alcohol content at about 7. % vol, then add the commercially available Komyohui brewing vinegar acetic acid agent in an amount of 0.5 ‰ (w / w) of the mass of sake mash, and increase the aeration rate to 0.3 to 0.4 vvm. , and the stirring speed was adjusted to 250 r/min to carry out acetic acid fermentation, and the next step was the same as step (4) above.

対照群と比較して、試験群は、開始速度が速く、発酵周期が１８．８９％短縮され、総酸含有量は１４．８２％、不揮発酸含有量は１０．５倍以上、総エステル含有量は１９倍以上向上した。試験群で製造されたリンゴ酢は、刺激性が著しく低下し、口当たりがより柔らかく、顕著な総合的な香りがあり、品質が著しく向上した。 Compared with the control group, the test group has a faster initiation rate, a 18.89% shorter fermentation cycle, a total acid content of 14.82%, a non-volatile acid content of more than 10.5 times, and a total ester content. The amount improved more than 19 times. The apple cider vinegar produced in the test group was significantly less pungent, had a softer mouthfeel, a pronounced overall aroma, and was of significantly improved quality.

実施例７：二元複合発酵剤の処方及び添加方法の比較
本実施例は、本発明の二元複合発酵剤の処方及び添加方法の比較を提供し、さらに、本発明の二元複合発酵剤の処方組成及びその添加方法の使用効果の重要性を説明する。 Example 7: Comparison of formulation and addition method of binary composite fermenting agent This example provides a comparison of formulation and addition method of the binary composite fermenting agent of the present invention, furthermore I will explain the importance of the use effect of the formulation composition and its addition method.

１、試験群
（１）４００ｋｇのかめを２４個準備し、もち米５０ｋｇに水を加えて一晩浸した。もち米を蒸気で蒸し、約４０℃になるまで冷水で冷却し、酒薬０．３ｋｇを加え、よく混ぜた後、かめに入れてラッパ状の穴を作った。 1. Test Group (1) Twenty-four jars of 400 kg were prepared, and 50 kg of glutinous rice was added with water and soaked overnight. The glutinous rice was steamed and cooled with cold water until it reached about 40°C.

（２）穴の中に一定の酒液が出たら、かめごとに麦麹２．５ｋｇを加え、次に水１５０ｋｇを加え、よく混ぜた。 (2) When a certain amount of sake liquid appeared in the hole, 2.5 kg of barley koji was added to each jar, then 150 kg of water was added and mixed well.

（３）アルコール発酵過程において、定期的に撹拌し、温度を３０℃程度に制御し、５～７ｄ程度発酵させると終了した。 (3) During the alcohol fermentation process, the mixture was stirred periodically, the temperature was controlled at about 30°C, and fermentation was completed for about 5-7 days.

（４）上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、バチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコースを６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみを約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温した。 (4) 200 kg of sake mash was added to each of the three jars of 400 kg in step (3) above, and the compound fungicide agent I of the present invention (25 parts of Lactobacillus helvetics CGMCC12225, 15 parts of Lactobacillus fermentum CGMCC12226) was added. , Lactobacillus acetotolerance CGMCC 16938 20 parts, Bacillus sonorensii CGMCC 15824 15 parts, Bacillus coagulans CGMCC 17801 10 parts, acid-stable medium-temperature α-amylase 13 parts, pullulanase 3 parts, glucoamylase 5 parts, cellulase 9 parts, and acid protease 5 parts Part) was added in an amount of 0.15‰ (w / w) of the mass of sake mash, mixed well, then 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar). . The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added in an amount of 0.1‰ (w/w) of the mass of sake mash. Add about 1000g of sake mash at 30°C to a small bucket and mix well, then sprinkle on top of the vinegared moromi in each pot. It was kept warm by covering it with rice bran.

（５）鎮江香酢の醸造プロセスに従って層ごとに醪を切り返し、総酸がこれ以上増加しなくなるまで酢醪を発酵させると、発酵が終了した。塩を加えて醪を１５ｄ密閉し、次に炒めた米を加えて酢を抽出した。酢を煎じ、熟成し、殺菌して充填すれば、完成品が得られた。 (5) Cut the mash layer by layer according to the brewing process of Zhenjiang fragrant vinegar, let the vinegar mash ferment until the total acid no longer increases, and the fermentation is finished. Salt was added and the mash was sealed for 15d, then the fried rice was added to extract the vinegar. Once the vinegar was decoctioned, aged, pasteurized and filled, the finished product was obtained.

２、比較試験
（１）対照群Ａ
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、、ラクトバチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温し、次のステップは上記のステップ（５）と同じであった。 2, Comparative test (1) Control group A
200 kg of sake mash was added to each of the three jars of 400 kg in step (3) above, and the compound fungicide agent I of the present invention (15 parts of Lactobacillus fermentum CGMCC12226, 20 parts of Lactobacillus acetotolerance CGMCC16938, Lactobacillus 10 parts of coagulans CGMCC17801, 13 parts of acid-stable medium-temperature α-amylase, 3 parts of pullulanase, 5 parts of glucoamylase, 9 parts of cellulase, and 5 parts of acid protease) were added to 0.15‰ (w/w) of the mass of sake mash. After mixing well, 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar mash). The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash. Add about 1000g of sake moromi (30°C) to the small tub and mix well. Sprinkle over the top of the vinegared moromi in each pot. Put your hand about 10 cm deep from the surface and mix well. Finally, add rice bran to the top. was covered and kept warm, the next step was the same as step (5) above.

（２）対照群Ｂ
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温した。 (2) Control group B
200 kg of sake mash was added to each of the three jars of 400 kg in step (3) above, and the compound fungicide I of the present invention (25 parts of Lactobacillus helvetics CGMCC122, 15 parts of Lactobacillus fermentum CGMCC12226, Bacillus 15 parts of Sonorenshi CGMCC15824, 13 parts of acid-resistant mesophilic α-amylase, 3 parts of pullulanase, 5 parts of glucoamylase, 9 parts of cellulase, and 5 parts of acid protease) were added to 0.15‰ (w/w) of the mass of sake mash. After adding and mixing well, 80 kg of wheat bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar mash). The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash. Add about 1000g of sake moromi (30°C) to the small tub and mix well. Sprinkle over the top of the vinegared moromi in each pot. Put your hand about 10 cm deep from the surface and mix well. Finally, add rice bran to the top. to keep warm.

（３）対照群Ｃ
上記ステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、ラクトバチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、ラクトバチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＰＧＭＣＣ１７８０２５５部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に米ぬかを被せて保温した。次のステップは上記のステップ（５）と同じであった。 (3) Control group C
200 kg of sake mash was added to each of the three jars of 400 kg in step (3) above, and the compound fungicide I of the present invention (25 parts of Lactobacillus helvetics CGMCC12225, 15 parts of Lactobacillus fermentum CGMCC12226, Lactobacillus) was added.・20 parts of acetotolerance CGMCC16938, 15 parts of Lactobacillus sonorensi CGMCC15824, 10 parts of Lactobacillus coagulans CGMCC17801, 13 parts of acid-stable medium-temperature α-amylase, 3 parts of pullulanase, 5 parts of glucoamylase, 9 parts of cellulase, and 5 parts of acid protease ) was added in an amount of 0.15‰ (w/w) of the mass of the sake mash and mixed well, then 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar mash). The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus PGMCC17802 and 60 parts of glucose) was added in an amount of 0.1‰ (w/w) of the mass of sake mash, and about 1000 g of sake mash at 30 ° C. was added to the small tub and mixed well, then sprinkled on top of the vinegared moromi in each container, and the hand was placed about 10cm deep from the surface to mix well, and finally the mixture was covered with rice bran to keep it warm. The next step was the same as step (5) above.

（４）対照群Ｄ
上記のステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、バチルス・コアグランスＣＧＭＣＣ１７８０１１０部、プルラナーゼ３部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温し、次のステップは上記ステップ（５）と同じであった。 (4) Control group D
200 kg of sake mash was added to each of the three 400 kg jars of step (3) above, and the compound fungicide I of the present invention (25 parts of Lactobacillus helvetics CGMCC12225, 15 parts of Lactobacillus fermentum CGMCC12226, Lacto 20 parts of Bacillus acetotolerance CGMCC16938, 15 parts of Bacillus sonorensi CGMCC15824, 10 parts of Bacillus coagulans CGMCC17801, 3 parts of pullulanase, 9 parts of cellulase, and 5 parts of acid protease) were added to the mass of sake mash at 0.15‰ (w/ w) was added, and after mixing well, 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar mash). The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash. Add about 1000g of sake moromi (30°C) to the small tub and mix well. Sprinkle over the top of the vinegared moromi in each pot. Put your hand about 10 cm deep from the surface and mix well. Finally, add rice bran to the top. and kept warm, the next step was the same as step (5) above.

（５）対照群Ｅ
上記のステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８１５部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、バチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、よく混ぜた後、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れて混ぜ、最後に上部に米ぬかを被せて保温し、次のステップは上記ステップ（５）と同じであった。 (5) Control group E
200 kg of sake mash was added to each of the three 400 kg jars of step (3) above, and the compound fungicide I of the present invention (25 parts of Lactobacillus helvetics CGMCC12225, 15 parts of Lactobacillus fermentum CGMCC12226, Lacto Bacillus acetotolerance CGMCC16938 15 parts, Bacillus sonorensii CGMCC15824 15 parts, Bacillus coagulans CGMCC17801 10 parts, acid-stable medium-temperature α-amylase 13 parts, pullulanase 3 parts, glucoamylase 5 parts, and acid protease 5 parts) were mixed with sake mash. After mixing well, 80 kg of bran and 45 kg of rice bran were added, and the sake mash and grains were uniformly mixed (vinegar mash). The compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added in an amount of 0.1 ‰ (w / w) of the mass of sake mash. Add about 1000g of sake mash (30°C) to the small tub and mix well. Sprinkle on top of the vinegared moromi in each pot. Put your hands about 10cm deep from the surface to mix. Finally, add rice bran to the top. Covered and kept warm, the next step was the same as step (5) above.

（６）対照群１
上記のステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇを加え、本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、バチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合し（酢醪）、最後に上部に米ぬかを被せて保温し、次のステップは上記ステップ（５）と同じであった。 (6) Control group 1
200 kg of sake mash was added to each of the three 400 kg jars of step (3) above, and the compound fungicide I of the present invention (25 parts of Lactobacillus helvetics CGMCC12225, 15 parts of Lactobacillus fermentum CGMCC12226, Lacto Bacillus acetotolerance CGMCC16938 20 parts, Bacillus sonorensii CGMCC15824 15 parts, Bacillus coagulans CGMCC17801 10 parts, acid-stable medium-temperature α-amylase 13 parts, pullulanase 3 parts, glucoamylase 5 parts, cellulase 9 parts, and acid protease 5 parts) was added in an amount of 0.15‰ (w/w) of the mass of sake mash, and the compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus GMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose Part) is added in an amount of 0.1‰ (w / w) of the mass of sake moromi, 80 kg of bran and 45 kg of rice bran are added, the sake moromi and grains are uniformly mixed (vinegar mash), and finally Covered with rice bran and kept warm, the next step was the same as step (5) above.

（７）対照群２
上記のステップ（３）の４００ｋｇのかめのうち３個のそれぞれに酒もろみ２００ｋｇ、ふすま８０ｋｇ、米ぬか４５ｋｇを加え、酒もろみと穀物を均一に混合した（酢醪）。本発明の複合菌剤Ｉ（ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５２５部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６１５部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８２０部、バチルス・ソノレンシＣＧＭＣＣ１５８２４１５部、バチルス・コアグランスＣＧＭＣＣ１７８０１１０部、耐酸性中温α－アミラーゼ１３部、プルラナーゼ３部、グルコアミラーゼ５部、セルラーゼ９部、及び酸性プロテアーゼ５部）を、酒もろみの質量の０．１５‰（ｗ／ｗ）の添加量で加え、また、本発明の複合菌剤ＩＩ（アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２５５部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５１０部、及びグルコース６０部）を、酒もろみの質量の０．１‰（ｗ／ｗ）の添加量で加え、３０℃の酒もろみ約１０００ｇを小桶に加えてよく混ぜた後、それぞれのかめ内の酢醪の上部に散布し、手を表面から約１０ｃｍ深く入れてよく混ぜ、最後に上部に米ぬかを被せて保温した。次のステップは上記のステップ（５）と同じであった。 (7) Control group 2
200 kg of sake mash, 80 kg of bran and 45 kg of rice bran were added to each of three of the 400 kg jars of step (3) above, and the sake mash and grains were uniformly mixed (vinegar mash). Combined fungal agent I of the present invention (Lactobacillus helvetics CGMCC12225 25 parts, Lactobacillus fermentum CGMCC12226 15 parts, Lactobacillus acetotolerance CGMCC16938 20 parts, Bacillus sonorensi CGMCC15824 15 parts, Bacillus coagulans CGMCC17801 10 parts, 13 parts of acid-stable medium-temperature α-amylase, 3 parts of pullulanase, 5 parts of glucoamylase, 9 parts of cellulase, and 5 parts of acid protease) are added in an amount of 0.15‰ (w/w) of the mass of sake mash, In addition, the compound fungicide II of the present invention (55 parts of Acetobacter pasteurianus CGMCC17802, 10 parts of Gluconacetobacter europeus CGMCC16345, and 60 parts of glucose) was added at 0.1‰ (w/w) of the mass of sake mash. After adding about 1000g of sake moromi at 30°C to the small tub and mixing well, sprinkle it on the top of the vinegar moromi in each pot, put your hand about 10 cm deep from the surface and mix well, and finally top the top. It was kept warm by covering it with rice bran. The next step was the same as step (5) above.

試験群と比較して、対照群Ａ～Ｅ中、総酸、不揮発酸、アミノ酸態窒素及び総エステル指標はいずれも対照群の効果に達しておらず、本発明の二元複合発酵剤処方は良好な効果を確保する上で重要な役割を果たしていることを示している。 Compared with the test group, the total acid, non-volatile acid, amino acid nitrogen and total ester indexes in the control groups A to E did not reach the effect of the control group, and the dual compound fermentation agent formulation of the present invention It shows that it plays an important role in ensuring a good effect.

試験群と比較して、対照群１は、昇温速度が遅く、初めて温度が４０℃以上に達する時間は２０ｈから１０３ｈまで８３ｈ延長し、発酵時間は１５ｄから２１ｄに６ｄ延長した。対照群２では、不揮発性酸含有量は著しく低下し、総エステル含有量は著しく低下した。 Compared to the test group, the control group 1 had a slower rate of temperature rise, the time when the temperature first reached 40°C or higher was extended by 83 hours from 20 hours to 103 hours, and the fermentation time was extended by 6 days from 15 hours to 21 hours. In control group 2, the non-volatile acid content was significantly reduced and the total ester content was significantly reduced.

本発明の二元複合発酵剤の配合組成及びその添加方法は、顕著な使用効果を達成させるのに重要な影響を与える。 The composition of the dual compound fermenting agent of the present invention and the method of adding it have an important effect on achieving remarkable effects.

Claims

複合菌剤Ｉと複合菌剤ＩＩとを含む二元複合発酵剤であって、
複合菌剤Ｉは、ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８、バチルス・ソノレンシＣＧＭＣＣ１５８２４、バチルス・コアグランスＣＧＭＣＣ１７８０１、耐酸性中温α－アミラーゼ、プルラナーゼ、グルコアミラーゼ、セルラーゼ、及び酸性プロテアーゼを含み、
複合菌剤ＩＩは、アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５、及びグルコースを含む、を含むことを特徴とする二元複合発酵剤。 A binary composite fermenting agent comprising a composite fungicide I and a composite fungicide II,
The compound fungicide I includes Lactobacillus helvetics CGMCC12225, Lactobacillus fermentum CGMCC12226, Lactobacillus acetotolerance CGMCC16938, Bacillus sonorensi CGMCC15824, Bacillus coagulans CGMCC17801, acid-stable medium-temperature α-amylase, pullulanase, glucoamylase, including cellulase and acid protease,
A dual compound fermenting agent, wherein the compound fungicide II comprises Acetobacter pasteurianus CGMCC17802, Gluconacetobacter europeus CGMCC16345, and glucose.

前記複合菌剤Ｉは、ラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５１０～２５重量部、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６５～１５重量部、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８５～２０重量部、バチルス・ソノレンシＣＧＭＣＣ１５８２４３～１５重量部、バチルス・コアグランスＣＧＭＣＣ１７８０１２～１０重量部、耐酸性中温α－アミラーゼ５～１３重量部、プルラナーゼ１～３重量部、グルコアミラーゼ２～５重量部、セルラーゼ３～９重量部、及び酸性プロテアーゼ１～５重量部を含み、
複合菌剤ＩＩは、アセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２３５～５５重量部、グルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５２～１０重量部、及びグルコース４０～６０重量部を含む、ことを特徴とする請求項１に記載の二元複合発酵剤。 The compound fungicide I includes 10 to 25 parts by weight of Lactobacillus helvetics CGMCC12225, 5 to 15 parts by weight of Lactobacillus fermentum CGMCC12226, 5 to 20 parts by weight of Lactobacillus acetotolerance CGMCC16938, and 3 to 20 parts by weight of Bacillus sonorensi CGMCC15824. 15 parts by weight, 2-10 parts by weight of Bacillus coagulans CGMCC17801, 5-13 parts by weight of acid-resistant medium-temperature α-amylase, 1-3 parts by weight of pullulanase, 2-5 parts by weight of glucoamylase, 3-9 parts by weight of cellulase, and acidity Contains 1 to 5 parts by weight of protease,
2. The compound fungicide II according to claim 1, comprising 35 to 55 parts by weight of Acetobacter pasteurianus CGMCC17802, 2 to 10 parts by weight of Gluconacetobacter europeus CGMCC16345, and 40 to 60 parts by weight of glucose. Binary compound fermentation agent.

前記複合菌剤Ｉにおけるラクトバチルス・ヘルヴェティクスＣＧＭＣＣ１２２２５、ラクトバチルス・ファーメンタムＣＧＭＣＣ１２２２６、ラクトバチルス・アセトトレランスＣＧＭＣＣ１６９３８、バチルス・ソノレンシＣＧＭＣＣ１５８２４、及びバチルス・コアグランスＣＧＭＣＣ１７８０１の生菌数がいずれも１．０×１０^８～１．０×１０^１０ＣＦＵ／ｇであり、耐酸性中温α－アミラーゼ、プルラナーゼ、グルコアミラーゼ、セルラーゼ、及び酸性プロテアーゼの酵素活性がいずれも２～１０万Ｕ／ｇである、ことを特徴とする請求項１に記載の二元複合発酵剤。 The number of viable bacteria of Lactobacillus helvetics CGMCC12225, Lactobacillus fermentum CGMCC12226, Lactobacillus acetotolerance CGMCC16938, Bacillus sonorensi CGMCC15824, and Bacillus coagulans CGMCC17801 in the compound fungicide I is all 1.0×10. ⁸ to 1.0×10 ¹⁰ CFU/g, and the enzymatic activities of acid-stable medium-temperature α-amylase, pullulanase, glucoamylase, cellulase, and acid protease are all 20,000 to 100,000 U/g. The binary composite fermenting agent according to claim 1.

前記複合菌剤ＩＩにおけるアセトバクター・パスツリアヌスＣＧＭＣＣ１７８０２及びグルコナセトバクター・ユーロペウスＣＧＭＣＣ１６３４５の生菌数が１．０×１０^７～１．０×１０^９ＣＦＵ／ｇである、ことを特徴とする請求項１に記載の二元複合発酵剤。 The number of viable cells of Acetobacter pasteurianus CGMCC17802 and Gluconacetobacter europeus CGMCC16345 in the compound antibacterial agent II is 1.0×10 ⁷ to 1.0×10 ⁹ CFU/g. Binary composite fermentation agent according to.

食酢の醸造における請求項１～４のいずれかに記載の二元複合発酵剤の使用。 Use of the binary compound fermenting agent according to any one of claims 1 to 4 in vinegar brewing.

前記複合菌剤Ｉは酢酸発酵前に添加され、前記複合菌剤ＩＩは酢酸発酵段階で添加される、ことを特徴とする請求項５に記載の使用。 The use according to claim 5, characterized in that the compound fungicide I is added before the acetic acid fermentation and the compound fungicide II is added during the acetic acid fermentation stage.

前記二元複合発酵剤は種醪と併用される、ことを特徴とする請求項５に記載の使用。 The use according to claim 5, characterized in that said binary complex fermenting agent is used in combination with seed mash.

前記食酢の醸造は、固体食酢発酵又は液体食酢発酵である、ことを特徴とする請求項５に記載の使用。 6. Use according to claim 5, characterized in that the vinegar brewing is solid vinegar fermentation or liquid vinegar fermentation.

前記二元複合発酵剤は、直投方式で添加される、ことを特徴とする請求項５に記載の使用。 The use according to claim 5, characterized in that said binary compound fermenting agent is added by direct injection method.

前記食酢は、鎮江香酢、山西老陳酢、米酢又はリンゴ酢である、ことを特徴とする請求項５に記載の使用。 The use according to claim 5, characterized in that the vinegar is Zhenjiang fragrant vinegar, Shanxi Laochen vinegar, rice vinegar or apple cider vinegar.