WO2021227453A1 - 一种利用小麦b淀粉生产3-羟基丁酮的工艺方法 - Google Patents

一种利用小麦b淀粉生产3-羟基丁酮的工艺方法 Download PDF

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WO2021227453A1
WO2021227453A1 PCT/CN2020/133657 CN2020133657W WO2021227453A1 WO 2021227453 A1 WO2021227453 A1 WO 2021227453A1 CN 2020133657 W CN2020133657 W CN 2020133657W WO 2021227453 A1 WO2021227453 A1 WO 2021227453A1
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starch
hydroxybutanone
wheat
fermentation
process method
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French (fr)
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张家祥
赵祥颖
刘丽萍
韩墨
杨丽萍
姚明静
刘建军
田延军
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山东省食品发酵工业研究设计院
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/24—Preparation of oxygen-containing organic compounds containing a carbonyl group
    • C12P7/26—Ketones
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C45/00—Preparation of compounds having >C = O groups bound only to carbon or hydrogen atoms; Preparation of chelates of such compounds
    • C07C45/78—Separation; Purification; Stabilisation; Use of additives
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C45/00—Preparation of compounds having >C = O groups bound only to carbon or hydrogen atoms; Preparation of chelates of such compounds
    • C07C45/78—Separation; Purification; Stabilisation; Use of additives
    • C07C45/81—Separation; Purification; Stabilisation; Use of additives by change in the physical state, e.g. crystallisation
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C45/00—Preparation of compounds having >C = O groups bound only to carbon or hydrogen atoms; Preparation of chelates of such compounds
    • C07C45/78—Separation; Purification; Stabilisation; Use of additives
    • C07C45/81—Separation; Purification; Stabilisation; Use of additives by change in the physical state, e.g. crystallisation
    • C07C45/82—Separation; Purification; Stabilisation; Use of additives by change in the physical state, e.g. crystallisation by distillation
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C45/00—Preparation of compounds having >C = O groups bound only to carbon or hydrogen atoms; Preparation of chelates of such compounds
    • C07C45/78—Separation; Purification; Stabilisation; Use of additives
    • C07C45/81—Separation; Purification; Stabilisation; Use of additives by change in the physical state, e.g. crystallisation
    • C07C45/82—Separation; Purification; Stabilisation; Use of additives by change in the physical state, e.g. crystallisation by distillation
    • C07C45/83—Separation; Purification; Stabilisation; Use of additives by change in the physical state, e.g. crystallisation by distillation by extractive distillation
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C49/00—Ketones; Ketenes; Dimeric ketenes; Ketonic chelates
    • C07C49/04—Saturated compounds containing keto groups bound to acyclic carbon atoms
    • C07C49/17—Saturated compounds containing keto groups bound to acyclic carbon atoms containing hydroxy groups
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor

Definitions

  • the invention belongs to the technical field of deep processing of agricultural products, and specifically relates to a process method for producing 3-hydroxybutanone by using wheat B starch.
  • 3-Hydroxybutanone also known as acetoin and acetomethylmethanol
  • acetoin is a widely used flavoring and important C4 platform compound, and has a wide range of applications in food, medicine, chemical industry, tobacco and other fields.
  • the 3-hydroxybutanone products on the market are mainly produced by chemical synthesis, and the price is about 100,000 to 150,000 yuan/ton.
  • the chemical method has limited sources of raw materials, many by-products, and products that do not conform to the natural, green, and safe consumption concept, which restricts its application and development in the food industry.
  • the purity of 3-hydroxybutanone produced by chemical methods is low, which limits its application and promotion as a platform compound in the pharmaceutical, chemical and other industries.
  • 3-Hydroxybutanone is a metabolite of a variety of microorganisms, which can be produced through microbial transformation using glucose as a raw material.
  • 3-Hydroxybutanone produced by biotransformation has the advantages of rich sources of raw materials, high product purity, and high product safety.
  • the market price of the product is twice that of chemical products (250,000-300,000 yuan/ton), and the added value of the product is higher. .
  • the inventor has carried out biotechnology to produce 3-hydroxybutanone for many years, and the screened Bacillus subtilis has a high yield of 3-hydroxybutanone and has good industrial application potential (Zhao Xiangying et al., a subtilis that produces high-purity 3-hydroxybutanone Bacillus, Chinese patent, ZL2007100134025, CN101016530B, 2007; Zhao Xiangying et al., a 3-hydroxybutanone-producing bacillus and its application, Chinese patent ZL201310289934.7, 2013; Fan Yixiao et al., screening of 3-hydroxybutanone-producing strains And product analysis, Food Fermentation Industry, 2012,38(11):42-46.).
  • the currently reported biotransformation technology for the production of 3-hydroxybutanone basically uses glucose as a raw material.
  • organic nitrogen sources such as yeast extract and corn steep liquor
  • inorganic nitrogen sources such as ammonium sulfate and urea need to be added to the fermentation medium.
  • the raw material cost accounts for about 60-70% of the total production cost.
  • the production of gluten is one of the main deep processing methods of wheat.
  • the protein content in wheat is about 12-13% and the starch content is 65-70%. Therefore, a large amount of wheat starch will be co-produced in the process of producing gluten.
  • Wheat starch is divided into A starch and B starch according to the different particle size.
  • a starch has larger granules and is relatively easy to separate by centrifugation, and can be further refined and sold as commercial starch.
  • Starch B is a starch with smaller granules.
  • During production it forms a three-phase centrifuge to separate non-starch components such as pentosan and residual protein in wheat starch slurry.
  • the dry content of this component accounts for about 80% of the starch content. ,
  • the viscosity is large, the water holding capacity is strong, and the value-added disposal is difficult.
  • the inventor found that the current production of B starch is mainly used for alcohol fermentation or sold as feed after drying, and the added value of
  • the present invention provides a process method for producing 3-hydroxybutanone using wheat B starch.
  • the wheat B starch produced during the production of gluten is used for the fermentation production of 3-hydroxybutanone after sugar production. Adopting the above process method can not only effectively reduce the production cost of 3-hydroxybutanone, but also effectively increase the added value of wheat B starch, so it has good practical application value.
  • the first aspect of the present invention provides the application of wheat B starch in the fermentation production of 3-hydroxybutanone.
  • the second aspect of the present invention provides a process method for producing 3-hydroxybutanone using wheat B starch.
  • the process method includes: preparing sugar from wheat B starch to obtain a starch sugar solution, and using the starch sugar solution as a raw material to prepare fermentation In the medium, the 3-hydroxybutanone producing strain is inoculated into the fermentation medium for fermentation to produce 3-hydroxybutanone.
  • the wheat B starch is mixed with water to obtain a starch syrup, the concentration of the starch syrup is controlled to be 10-15 Bé, and then the "dual enzyme method" sugar production process is used to prepare the starch sugar liquid.
  • the initial concentration of glucose is controlled to be 120-160 g/L;
  • the fermentation medium also includes an inorganic nitrogen source; furthermore, the inorganic nitrogen source may be urea.
  • the inorganic nitrogen source may be urea.
  • the third aspect of the present invention provides 3-hydroxybutanone prepared by the above-mentioned process for producing 3-hydroxybutanone using wheat B starch.
  • 3-Hydroxybutanone azeotropes with water can be collected by vacuum distillation or multi-effect concentration.
  • Starch B is brought into pentosan and other unfermented components to be enriched in the bottom distillation. Compared with glucose raw materials, it does not increase the extraction cost of 3-hydroxybutanone.
  • the bottom distillate can be further disposed of as feed.
  • the application of wheat B starch in the fermentation production of 3-hydroxybutanone is provided.
  • the present invention reports for the first time that the starch sugar solution of wheat B starch is used as the medium component for the fermentation and production of 3-hydroxybutanone by the production of strains. Tests have verified that only a small amount of inorganic nitrogen source needs to be added to the medium to achieve the 3-hydroxybutanone is produced by fermentation, and the yield and conversion rate of 3-hydroxybutanone are relatively high.
  • a process method for producing 3-hydroxybutanone using wheat B starch includes: preparing sugar from wheat B starch to obtain a starch sugar solution, using the starch sugar solution as The raw materials are prepared to obtain a fermentation medium, and 3-hydroxybutanone production strains are inoculated into the fermentation medium for fermentation to produce 3-hydroxybutanone.
  • the wheat B starch can be selected from B starch syrup separated during the processing of gluten; or can be selected from B starch products prepared by direct drying of B starch;
  • the wheat B starch may be:
  • commodity B starch (dry powder) produced by wheat starch manufacturers.
  • the wheat B starch is mixed with water to obtain a starch syrup, the concentration of the starch syrup is controlled to be 10-15 Bé, and then the "dual enzyme method" sugar production process is used to prepare the starch sugar liquid.
  • the fermentation medium further includes an inorganic nitrogen source; furthermore, the inorganic nitrogen source may be urea.
  • the fermentation medium includes the above-mentioned starch sugar solution and an inorganic nitrogen source, and the inorganic nitrogen source is urea;
  • the initial concentration of glucose is controlled to be 120-160 g/L, and the concentration of urea is 1-2 g/L.
  • the 3-hydroxybutanone producing strain is preferably Bacillus, more preferably Bacillus subtilis; in the embodiment of the present invention, the 3-hydroxybutanone producing strain is It is Bacillus subtilis BS168D.
  • the process method further includes the collection and purification of 3-hydroxybutanone; specifically, the specific method for collection and purification is: collecting the fermentation broth of 3-hydroxybutanone for fractionation treatment, and The obtained 3-hydroxybutanone fraction is separated by salt extraction and distillation to obtain 3-hydroxybutanone crystals; the specific purification method can also refer to CN 201310426868.3.
  • the specific method of fractionation treatment includes: collecting the 3-hydroxybutanone fermentation broth by vacuum distillation or multi-effect concentration method, and the distillation temperature is controlled to 55-75 °C; the collection of fractions accounts for 75-85% of the total volume of the fermentation broth and stop fraction collection.
  • 3-hydroxybutanone azeotropes with water the 3-hydroxybutanone in the fermentation broth can be collected by vacuum distillation or multi-effect concentration, and the B starch raw material is brought into pentosan and other unfermented components to enrich In bottom distillation, the extraction cost of 3-hydroxybutanone is not increased compared with the raw material of glucose.
  • 3-hydroxybutanone prepared by the above-mentioned process for producing 3-hydroxybutanone using wheat B starch is provided.
  • Bacillus subtilis BS168D (2,3-butanediol dehydrogenase gene (bdhA) deletion Bacillus subtilis 168 engineering strain) (Junjiao Zhang, Xiangying Zhao, Jiaxiang Zhang, Chen Zhao, Jianjun Liu, Yanjun Tian&Liping Yang( 2017) Effect of deletion of 2,3-butanediol dehydrogenase gene (bdhA) on acetoin production of Bacillus subtilis, Preparative Biochemistry and Biotechnology, 47: 8,761-767).
  • the above-mentioned strains can be obtained by the public from Shandong Food Fermentation Industry Research and Design Institute.
  • B Starch Obtained Take the fresh wheat starch slurry produced during the production of gluten and centrifuge it with a centrifuge at 3000 speed for 2 seconds, remove the supernatant liquid, collect the intermediate fluid slurry part, and use it for the preparation of starch sugar solution.
  • Preparation of sugar solution mix the separated and washed starch with water and adjust the concentration of starch slurry to 10-15 Bé. Add appropriate amount of amylase to make sugar by double-enzyme method based on the total amount of starch. After the saccharification is completed, centrifugation is performed, and the clear liquid is collected for the fermentation of 3-hydroxybutanone.
  • Seed culture medium glucose 50g/L, yeast extract 10g/L, corn steep liquor 10g/L, sodium chloride 5g/L, drinking water preparation, pH 7.0.
  • Shake flask fermentation 500ml triangular flask with 50ml liquid volume, 5% inoculation volume, and culturing for 3-4 days under optimal culture conditions.
  • Fermentation tank fermentation The fermentation adopts a laboratory scale (5-50L) fermentation tank, and the appropriate temperature and relative dissolved oxygen concentration are controlled according to the needs during the fermentation process. Cultivate until the residual glucose drops below 0.1g/L to end the fermentation.
  • Example 1 The effect of culture medium on the fermentation of 3-hydroxybutanone
  • Example 2 50L fermentation with B starch syrup as raw material
  • Fresh wheat B starch syrup is used to prepare starch sugar using conventional double-enzyme sugar production technology. After saccharification, the solid residue is removed by centrifugation. The residue can be washed with equal volume of water, centrifuged, and centrifuged to separate the clear liquid, adjust the appropriate glucose concentration, and add it to the fermentation Inside the tank, sterilize. In addition, 2g/L urea was added (sterilized separately, added before inoculation). Inoculate according to the inoculation amount of 5%, the initial sugar concentration after inoculation is 138g/L, and the appropriate temperature and relative dissolved oxygen concentration are controlled during the fermentation process. After culturing for 58 hours, the glucose concentration was determined to drop below 0.1g/L, and the fermentation was terminated. A total of 35L of fermentation broth was collected, and the content of 3-hydroxybutanone was determined to be 53.2g/L.

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Abstract

一种利用小麦B淀粉生产3-羟基丁酮的工艺方法，属于农产品深加工技术领域。具体工艺方法包括：小麦B淀粉制糖得淀粉糖液，以所述淀粉糖液为原料配制获得发酵培养基，将3-羟基丁酮生产菌种接种至发酵培养基中进行发酵生产得3-羟基丁酮。采用上述工艺方法，既能有效降低3-羟基丁酮的生产成本，同时还有效增加了小麦B淀粉的附加值，因此具有良好的实际应用价值。

Description

一种利用小麦B淀粉生产3-羟基丁酮的工艺方法 技术领域
本发明属于农产品深加工技术领域，具体涉及一种利用小麦B淀粉生产3-羟基丁酮的工艺方法。
背景技术
公开该背景技术部分的信息仅仅旨在增加对本发明的总体背景的理解，而不必然被视为承认或以任何形式暗示该信息构成已经成为本领域一般技术人员所公知的现有技术。
3-羟基丁酮(acetoin)又名乙偶姻、乙酰甲基甲醇，是一种应用广泛的食用香料和重要C4平台化合物，在食品、医药、化工、烟草等领域具有广泛应用。目前，市场上的3-羟基丁酮产品主要是化学合成法生产的，价格约为10-15万元/吨。化学法生产原料来源受限、副产物较多，产品也不符合天然、绿色、安全的消费理念，限制了其在食品行业的应用发展。另外，化学法生产的3-羟基丁酮纯度偏低，限制了其作为平台化合物在医药、化工等行业的应用推广。3-羟基丁酮为多种微生物代谢产物，可以葡萄糖为原料经微生物转化生产。利用生物转化生产的3-羟基丁酮具有原料来源丰富、产物纯度高、产品安全性高等优点，产品市场价格为化学法产品的2倍(25-30万元/吨)，产品附加值较高。发明人开展生物技术生产3-羟基丁酮多年，筛选的枯草芽孢杆菌3-羟基丁酮产率高，具有较好的工业应用潜力(赵祥颖等，一株产高纯度3-羟基丁酮的枯草芽孢杆菌，中国专利，ZL2007100134025，CN101016530B，2007；赵祥颖等，一株产3-羟基丁酮的芽胞杆菌及其应用，中国专利ZL201310289934.7，2013；范宜晓等，产3-羟基丁酮菌株的筛选及产物分析，食品发酵工业，2012,38(11):42-46.)。目前报道的生物转化生产3-羟基丁酮技术，基本都是以葡萄糖为原料，另外，发酵培养基中还需要添加酵母浸出物和玉米浆等有机氮源和硫铵、脲等无机氮源，以及少量的其 他无机盐，原料成本约占总生产成本的大约60-70％。
谷朊粉生产是小麦主要的深加工方式之一，小麦中蛋白含量大约12-13％，淀粉含量65-70％，因此，在生产谷朊粉的过程中会联产大量的小麦淀粉。小麦淀粉根据颗粒大小的不同分为A淀粉和B淀粉，A淀粉颗粒较大，通过离心分离相对容易，可以进一步精制作为商品淀粉出售。B淀粉是颗粒较小的淀粉，生产中和小麦淀粉浆中戊聚糖、残余蛋白等非淀粉成分一起组成三相离心机分离中间相组分，该组分干物中淀粉成分大约占80％，粘度大，持水性强，增值处置困难。发明人发现，B淀粉目前生产中主要用于酒精发酵或是干燥后作为饲料出售，产品附加值比较低。
发明内容
针对目前谷朊粉生产过程中产生的小麦B淀粉低值化利用的问题，本发明提供一种利用小麦B淀粉生产3-羟基丁酮的工艺方法。具体的，将谷朊粉生产过程中产生的小麦B淀粉制糖后用于3-羟基丁酮的发酵生产。采用上述工艺方法，既能有效降低3-羟基丁酮的生产成本，同时还有效增加了小麦B淀粉的附加值，因此具有良好的实际应用之价值。
为了实现上述技术目的，本发明的技术方案如下：
本发明的第一个方面，提供小麦B淀粉在发酵生产3-羟基丁酮中的应用。
本发明的第二个方面，提供利用小麦B淀粉生产3-羟基丁酮的工艺方法，所述工艺方法包括：小麦B淀粉制糖得淀粉糖液，以所述淀粉糖液为原料配制获得发酵培养基，将3-羟基丁酮生产菌种接种至发酵培养基中进行发酵生产得3-羟基丁酮。
进一步的，对小麦B淀粉制糖方法没有限制，如常用“双酶法”制糖工艺即可；
更进一步的，将所述小麦B淀粉加水调浆得淀粉浆，控制淀粉浆浓度 为10～15Bé，然后采用“双酶法”制糖工艺制备淀粉糖液。
进一步的，所述发酵培养基中，控制葡萄糖起始浓度为120～160g/L；
进一步的，所述发酵培养基还包括无机氮源；更进一步的，所述无机氮源可以是脲。本发明研究发现，以B淀粉糖浆为原料进行3-羟基丁酮发酵不需要添加酵母浸出物，并且3-羟基丁酮产率和转化率都明显高于葡萄糖。
本发明的第三个方面，提供上述利用小麦B淀粉生产3-羟基丁酮的工艺方法制备得到的3-羟基丁酮。
以上一个或多个技术方案的有益技术效果：
1)小麦谷朊粉生产过程中分离的B淀粉部分，干物约占40％，干物中淀粉约占80％左右，其他还含有蛋白质、脂肪、粗纤维、灰分等。研究发现以此为原料进行3-羟基丁酮发酵，只需要添加少量的无机氮源，不需要添加酵母浸出物等价格昂贵的有机氮源。同时B淀粉成本远低于葡萄糖，因此，利用小麦B淀粉可大幅降低3-羟基丁酮生产的原料成本；
2)3-羟基丁酮与水共沸，可以通过减压蒸馏或多效浓缩收集发酵液中的3-羟基丁酮，B淀粉原料带入戊聚糖等其他未发酵成分富集到底馏中，与葡萄糖原料相比并不增加3-羟基丁酮的提取成本。同时底馏物可以进一步作为饲料处置。
3)将小麦淀粉分离获得B淀粉用于生产3-羟基丁酮，产品附加值高，实现了小麦B淀粉的高附加值利用，因此具有良好的实际应用之价值。
具体实施方式
应该指出，以下详细说明都是例示性的，旨在对本发明提供进一步的说明。除非另有指明，本文使用的所有技术和科学术语具有与本发明所属技术领域的普通技术人员通常理解的相同含义。
需要注意的是，这里所使用的术语仅是为了描述具体实施方式，而非意图限制根据本发明的示例性实施方式。如在这里所使用的，除非上下文另外 明确指出，否则单数形式也意图包括复数形式，此外，还应当理解的是，当在本说明书中使用术语“包含”和/或“包括”时，其指明存在特征、步骤、操作、器件、组件和/或它们的组合。应理解，本发明的保护范围不局限于下述特定的具体实施方案；还应当理解，本发明实施例中使用的术语是为了描述特定的具体实施方案，而不是为了限制本发明的保护范围。
本发明的一个具体实施方式中，提供小麦B淀粉在发酵生产3-羟基丁酮中的应用。本发明首次报道了将小麦B淀粉的淀粉糖液作为生产菌种发酵生产3-羟基丁酮的培养基成分，经试验验证，只需要向培养基中再添加少量的无机氮源即可实现对3-羟基丁酮的发酵生产，且3-羟基丁酮产率和转化率均较高。
本发明的又一具体实施方式中，提供一种利用小麦B淀粉生产3-羟基丁酮的工艺方法，所述工艺方法包括：小麦B淀粉制糖得淀粉糖液，以所述淀粉糖液为原料配制获得发酵培养基，将3-羟基丁酮生产菌种接种至发酵培养基中进行发酵生产得3-羟基丁酮。
本发明的又一具体实施方式中，所述小麦B淀粉可选自谷朊粉加工过程中分离的B淀粉浆；也可以选自B淀粉直接干燥制备得到的B淀粉产品；
本发明的又一具体实施方式中，所述小麦B淀粉可以是：
取谷朊粉生产过程中产生的新鲜小麦淀粉浆用离心机离心，转速3000rpm离心时间2秒，移去上层清液，收集中间流动性粉浆部分；
或，三相卧螺离心机谷朊粉生产工艺中分离的中间相，分离谷朊粉后得到的B淀粉浆；
或，小麦淀粉生产厂家生产的商品B淀粉(干粉)。
本发明的又一具体实施方式中，对小麦B淀粉制糖方法没有限制，如常用“双酶法”制糖工艺即可；
本发明的又一具体实施方式中，将所述小麦B淀粉加水调浆得淀粉浆， 控制淀粉浆浓度为10～15Bé，然后采用“双酶法”制糖工艺制备淀粉糖液。
本发明的又一具体实施方式中，所述发酵培养基还包括无机氮源；更进一步的，所述无机氮源可以是脲。本发明研究发现，以B淀粉糖浆为原料进行3-羟基丁酮发酵不需要添加酵母浸出物，并且3-羟基丁酮产率和转化率都明显高于以葡萄糖为原料所进行的发酵。
本发明的又一具体实施方式中，所述发酵培养基包括上述淀粉糖液和无机氮源，所述无机氮源为脲；
本发明的又一具体实施方式中，所述发酵培养基中，控制葡萄糖起始浓度为120～160g/L，脲浓度为1～2g/L。
本发明的又一具体实施方式中，所述3-羟基丁酮生产菌种优选为芽孢杆菌，进一步优选为枯草芽孢杆菌；在本发明的实施例中，所述3-羟基丁酮生产菌种为枯草芽孢杆菌BS168D。
本发明的又一具体实施方式中，所述工艺方法还包括对3-羟基丁酮的收集纯化；具体的，所述收集纯化具体方法为：收集3-羟基丁酮发酵液进行分馏处理，将获得的3-羟基丁酮馏分加盐萃取精馏分离提取即获得3-羟基丁酮结晶；具体纯化方法亦可参考CN 201310426868.3进行。
本发明的又一具体实施方式中，所述分馏处理具体方法包括：将3-羟基丁酮发酵液采用减压蒸馏或多效浓缩的方式进行馏分的收集，所述蒸馏温度控制为55-75℃；收集馏分占发酵液总体积的75-85％停止馏分收集。在本发明中，3-羟基丁酮与水共沸，可以通过减压蒸馏或多效浓缩收集发酵液中的3-羟基丁酮，B淀粉原料带入戊聚糖等其他未发酵成分富集到底馏中，与葡萄糖原料相比并不增加3-羟基丁酮的提取成本。
本发明的又一具体实施方式中，提供上述利用小麦B淀粉生产3-羟基丁酮的工艺方法制备得到的3-羟基丁酮。
以下通过实施例对本发明做进一步解释说明，但不构成对本发明的限制。应理解这些实施例仅用于说明本发明而不用于限制本发明的范围。
实验材料
实验菌种：枯草芽孢杆菌BS168D(2,3-丁二醇脱氢酶基因(bdhA)缺失Bacillus subtilis 168工程菌株)(Junjiao Zhang,Xiangying Zhao,Jiaxiang Zhang,Chen Zhao,Jianjun Liu,Yanjun Tian&Liping Yang(2017)Effect of deletion of 2,3-butanediol dehydrogenase gene(bdhA)on acetoin production of Bacillus subtilis,Preparative Biochemistry and Biotechnology,47:8,761-767)。上述菌种公众可以从山东省食品发酵工业研究设计院获得。
B淀粉获得：取谷朊粉生产过程中产生的新鲜小麦淀粉浆用离心机离心，转速3000离心时间2秒，移去上层清液，收集中间流动性粉浆部分，用于淀粉糖液制备。
糖液的制备：将分离洗涤后的淀粉加水调浆，调整淀粉浆浓度10～15Bé，根据淀粉总量计算加入适量淀粉酶进行双酶法制糖(酶制剂加量按照生产厂商推荐量添加)，糖化结束后，离心分离，收集清液部分用于3-羟基丁酮发酵。
种子培养基：葡萄糖50g/L，酵母浸出物10g/L，玉米浆10g/L，氯化钠5g/L，饮用水配制，pH 7.0。
3-羟基丁酮发酵
取淀粉糖液，调节至适当葡萄糖浓度(120-160g/L)，配制3-羟基丁酮发酵培养基。对照实验采用口服葡萄糖配制。培养基经灭菌后接入菌种进 行发酵。
摇瓶发酵：500ml三角瓶装液量50ml，接种量5％，在最佳培养条件下培养3-4天。
发酵罐发酵：发酵采用实验室规模(5-50L)发酵罐，发酵过程中根据需求控制适当的温度和相对溶解氧浓度。培养至残留葡萄糖降至0.1g/L以下结束发酵。
实施例1培养基对3-羟基丁酮发酵的影响
实验用葡萄糖为对照进行3-羟基丁酮发酵实验，考察氮源添加对小麦B淀粉糖浆3-羟基丁酮发酵的影响，结果见下表：
Figure PCTCN2020133657-appb-000001
实验结果显示，以B淀粉糖浆为原料进行3-羟基丁酮发酵不需要添加酵母浸出物和玉米浆，并且3-羟基丁酮产率和转化率都明显高于葡萄糖。
实施例2以B淀粉糖浆为原料50L发酵
采用新鲜小麦B淀粉浆采用常规双酶制糖工艺制备淀粉糖，糖化结束后离心分离去固体残渣，残渣可以等体积水洗涤，离心分离，合并离心分离清液，调整适当的葡萄糖浓度，加入发酵罐内，灭菌。另添加2g/L脲(单独杀菌，接种前加入)。按照5％的接种量接种，接种后初始糖浓度为138g/L，发酵过程中控制适当温度和相对溶解氧浓度。培养58小时，测定葡萄糖浓度降至0.1g/L以下，结束发酵。共收集发酵液35L，测定3-羟基丁酮含量为53.2g/L。
取上述发酵液10L在70～80℃进行减压蒸馏，收集馏分8.3L，测定馏分中3-羟基丁酮的含量55.6g/L，3-羟基丁酮回收率86.7％。
应注意的是，以上实例仅用于说明本发明的技术方案而非对其进行限制。尽管参照所给出的实例对本发明进行了详细说明，但是本领域的普通技术人员可根据需要对本发明的技术方案进行修改或者等同替换，而不脱离本发明技术方案的精神和范围。

Claims (10)

  1. 小麦B淀粉在发酵生产3-羟基丁酮中的应用。
  2. 一种利用小麦B淀粉生产3-羟基丁酮的工艺方法，其特征在于，所述工艺方法包括：小麦B淀粉制糖得淀粉糖液，以所述淀粉糖液为原料配制获得发酵培养基，将3-羟基丁酮生产菌种接种至发酵培养基中进行发酵生产得3-羟基丁酮。
  3. 如权利要求2所述的工艺方法，其特征在于，所述小麦B淀粉选自谷朊粉加工过程中分离的B淀粉浆或B淀粉直接干燥制备得到的B淀粉产品。
  4. 如权利要求2所述的工艺方法，其特征在于，将所述小麦B淀粉加水调浆得淀粉浆，控制淀粉浆浓度为10～15Bé，然后采用“双酶法”制糖工艺制备淀粉糖液。
  5. 如权利要求2所述的工艺方法，其特征在于，所述发酵培养基还包括无机氮源；优选的，所述无机氮源是脲。
  6. 如权利要求2所述的工艺方法，其特征在于，所述发酵培养基包括淀粉糖液和无机氮源；
    优选的，所述无机氮源为脲；
    优选的，所述发酵培养基中，控制葡萄糖起始浓度为120～160g/L，脲浓度为1～2g/L。
  7. 如权利要求2所述的工艺方法，其特征在于，所述3-羟基丁酮生产菌种为芽孢杆菌；优选为枯草芽孢杆菌；进一步优选的，所述3-羟基丁酮生产菌种为枯草芽孢杆菌BS168D。
  8. 如权利要求2所述的工艺方法，其特征在于，所述工艺方法还包括对3-羟基丁酮的收集纯化；
    优选的，所述收集纯化具体方法为：收集3-羟基丁酮发酵液进行分馏处理，将获得的3-羟基丁酮馏分加盐萃取精馏分离提取即获得3-羟基丁酮结 晶。
  9. 如权利要求8所述的工艺方法，其特征在于，所述分馏处理具体方法包括：将3-羟基丁酮发酵液采用减压蒸馏或多效浓缩的方式进行馏分的收集，所述蒸馏温度控制为55-75℃；收集馏分占发酵液总体积的75-85％停止馏分收集。
  10. 权利要求2-9任一项所述利用小麦B淀粉生产3-羟基丁酮的工艺方法制备得到的3-羟基丁酮。
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Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111607622B (zh) * 2020-05-15 2022-02-22 山东省食品发酵工业研究设计院 一种利用小麦b淀粉生产3-羟基丁酮的工艺方法
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Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101161818A (zh) * 2007-09-29 2008-04-16 上海天之冠可再生能源有限公司 一种部分替代玉米发酵生产丙酮丁醇的方法
CN100455658C (zh) * 2007-01-29 2009-01-28 山东省食品发酵工业研究设计院 一株产高纯度3-羟基丁酮的枯草芽孢杆菌
CN102352396A (zh) * 2011-09-01 2012-02-15 河南永昌飞天淀粉糖有限公司 一种小麦淀粉生产功能淀粉糖的方法
WO2013023713A1 (en) * 2011-05-18 2013-02-21 Scale Biofuel, ApS Solar-assisted volatile fermentation products production processes
CN103333842A (zh) * 2013-07-10 2013-10-02 山东省食品发酵工业研究设计院 一株产3-羟基丁酮的芽孢杆菌及其应用
CN103483167A (zh) * 2013-08-27 2014-01-01 山东鲁北药业有限公司 3-羟基丁酮的一种纯化方法
CN103524315A (zh) * 2013-09-18 2014-01-22 山东省食品发酵工业研究设计院 一种加盐萃取蒸馏分离发酵液中3-羟基丁酮的方法
CN110305913A (zh) * 2019-06-28 2019-10-08 山东省食品发酵工业研究设计院 一种糖质原料生产四甲基吡嗪的方法
CN111607622A (zh) * 2020-05-15 2020-09-01 山东省食品发酵工业研究设计院 一种利用小麦b淀粉生产3-羟基丁酮的工艺方法

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101049133B (zh) * 2007-04-26 2011-07-06 华南理工大学 小麦b淀粉多酶法制取发酵用含氮糖浆的方法
CN101294143B (zh) * 2008-06-20 2010-11-03 南京工业大学 一种产3-羟基丁酮的菌株及其应用
CN102899364A (zh) * 2012-09-14 2013-01-30 日照鲁信金禾生化有限公司 一种小麦b淀粉发酵生产柠檬酸的方法
CN104328099A (zh) * 2014-11-13 2015-02-04 河南天冠纤维乙醇有限公司 一种重组毕赤酵母生产β-葡聚糖酶的方法
CN105624219A (zh) * 2016-03-07 2016-06-01 江苏国信协联能源有限公司 一种以小麦淀粉乳发酵生产柠檬酸的方法
CN105820988B (zh) * 2016-06-08 2019-06-11 中国石油大学(华东) 一株芽孢杆菌及其在高温发酵生产乙偶姻和2,3-丁二醇中的应用
CN107129959B (zh) * 2017-06-28 2020-12-18 广西科学院 生产(r)-乙偶姻基因工程菌株的构建方法及其应用

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN100455658C (zh) * 2007-01-29 2009-01-28 山东省食品发酵工业研究设计院 一株产高纯度3-羟基丁酮的枯草芽孢杆菌
CN101161818A (zh) * 2007-09-29 2008-04-16 上海天之冠可再生能源有限公司 一种部分替代玉米发酵生产丙酮丁醇的方法
WO2013023713A1 (en) * 2011-05-18 2013-02-21 Scale Biofuel, ApS Solar-assisted volatile fermentation products production processes
CN102352396A (zh) * 2011-09-01 2012-02-15 河南永昌飞天淀粉糖有限公司 一种小麦淀粉生产功能淀粉糖的方法
CN103333842A (zh) * 2013-07-10 2013-10-02 山东省食品发酵工业研究设计院 一株产3-羟基丁酮的芽孢杆菌及其应用
CN103483167A (zh) * 2013-08-27 2014-01-01 山东鲁北药业有限公司 3-羟基丁酮的一种纯化方法
CN103524315A (zh) * 2013-09-18 2014-01-22 山东省食品发酵工业研究设计院 一种加盐萃取蒸馏分离发酵液中3-羟基丁酮的方法
CN110305913A (zh) * 2019-06-28 2019-10-08 山东省食品发酵工业研究设计院 一种糖质原料生产四甲基吡嗪的方法
CN111607622A (zh) * 2020-05-15 2020-09-01 山东省食品发酵工业研究设计院 一种利用小麦b淀粉生产3-羟基丁酮的工艺方法

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
XU HUI, JIA SHI-RU, LIU JIAN-JUN: "Present Research Situation of Acetoin Production by Biological Method", SHANDONG FOOD FERMENT, vol. 36, no. 11, 1 January 2010 (2010-01-01), pages 137 - 144, XP055866942, DOI: 10.13995/j.cnki.11-1802/ts.2010.11.005 *
ZHANG JUNJIAO, ZHAO XIANGYING, ZHANG JIAXIANG, ZHAO CHEN, LIU JIANJUN, TIAN YANJUN, YANG LIPING: "Effect of deletion of 2,3-butanediol dehydrogenase gene (bdhA) on acetoin production of Bacillus subtilis", PREPARATIVE BIOCHEMISTRY AND BIOTECHNOLOGY, DEKKER, NEW YORK, NY, US, vol. 47, no. 8, 14 September 2017 (2017-09-14), US , pages 761 - 767, XP055859477, ISSN: 1082-6068, DOI: 10.1080/10826068.2017.1320293 *

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