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WO2019218445A1 - Procédé de synthèse d'acide acétohydroxamique inhibiteur d'uréase - Google Patents

Procédé de synthèse d'acide acétohydroxamique inhibiteur d'uréase Download PDF

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Publication number
WO2019218445A1
WO2019218445A1 PCT/CN2018/095475 CN2018095475W WO2019218445A1 WO 2019218445 A1 WO2019218445 A1 WO 2019218445A1 CN 2018095475 W CN2018095475 W CN 2018095475W WO 2019218445 A1 WO2019218445 A1 WO 2019218445A1
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WO
WIPO (PCT)
Prior art keywords
reaction
ethyl acetate
hydroxylamine hydrochloride
acetohydroxamic acid
amount
Prior art date
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Ceased
Application number
PCT/CN2018/095475
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English (en)
Chinese (zh)
Inventor
王加启
赵圣国
王芃芃
郑楠
张养东
李松励
李慧颖
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Institute of Animal Science of CAAS
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Institute of Animal Science of CAAS
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Filing date
Publication date
Application filed by Institute of Animal Science of CAAS filed Critical Institute of Animal Science of CAAS
Publication of WO2019218445A1 publication Critical patent/WO2019218445A1/fr
Anticipated expiration legal-status Critical
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/105—Aliphatic or alicyclic compounds
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K50/00—Feeding-stuffs specially adapted for particular animals
    • A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C259/00—Compounds containing carboxyl groups, an oxygen atom of a carboxyl group being replaced by a nitrogen atom, this nitrogen atom being further bound to an oxygen atom and not being part of nitro or nitroso groups
    • C07C259/04—Compounds containing carboxyl groups, an oxygen atom of a carboxyl group being replaced by a nitrogen atom, this nitrogen atom being further bound to an oxygen atom and not being part of nitro or nitroso groups without replacement of the other oxygen atom of the carboxyl group, e.g. hydroxamic acids
    • C07C259/06—Compounds containing carboxyl groups, an oxygen atom of a carboxyl group being replaced by a nitrogen atom, this nitrogen atom being further bound to an oxygen atom and not being part of nitro or nitroso groups without replacement of the other oxygen atom of the carboxyl group, e.g. hydroxamic acids having carbon atoms of hydroxamic groups bound to hydrogen atoms or to acyclic carbon atoms

Definitions

  • the invention relates to the field of preparation of animal husbandry additives, in particular to a method for synthesizing a urease inhibitor acetohydroxamic acid.
  • Microorganisms in the rumen can secrete urease, which decomposes urea in the diet into ammonia.
  • the ammonia can be used by microorganisms to synthesize microbial proteins to provide nitrogen to the body.
  • Inhibition of urease activity, slowing down the decomposition rate of urea in the rumen of ruminants, can improve the urea utilization rate and microbial protein synthesis efficiency of ruminants.
  • Acetoxyhydroxamic acid is a urease inhibitor.
  • the hydroxylamine structure (-NHOH) in the molecule of the hydroxamic acid oxidizes the sulfhydryl group (-SH) of the adjacent metal nickel (Ni) in the urea structure to a disulfide bond (- SS-), forming a urease inhibitor-urease binary complex, thereby inhibiting urease activity.
  • the inhibition process is reversible, reducing the hydrolysis rate of urea by urease, and ensuring the slow release of ammonia.
  • Scientific and rational addition of urease inhibitor can effectively slow down the decomposition rate of dietary urea in the rumen of ruminants, regulate the synthesis of rumen microbial proteins, improve the utilization of urea and nitrogen by ruminants, and finally improve the performance of ruminants.
  • the present invention provides a method for synthesizing acetohydroxamic acid (AHA), which comprises the following synthesis step: hydroxylamine hydrochloride and ethyl acetate are reacted in an alkaline solvent system, after completion of the reaction Extracted with an organic solvent,
  • AHA acetohydroxamic acid
  • the alkaline solvent is preferably a mixed solvent of sodium hydroxide, methanol and water.
  • the molar ratio of the sodium hydroxide to the hydroxylamine hydrochloride is (1-3):1, preferably 2:1.
  • the molar ratio of hydroxylamine hydrochloride to ethyl acetate is 1: (1.2 - 1.5), preferably 1: 1.2.
  • the temperature of the reaction is 35 to 45 ° C, for example, 40 ° C.
  • the organic solvent is preferably ethyl acetate.
  • the temperature of the extraction is preferably from 70 to 80 °C.
  • the sodium hydroxide is selected to be added in portions, and for example, sodium hydroxide may be added in portions before the reaction and in the middle of the reaction.
  • the synthesis method further comprises the step of crystallizing the product using a small amount of acetone after extraction and concentration using ethyl acetate.
  • the present invention also provides an acetohydroxamic acid obtained by the above-described production method as a rumen microbial urease inhibitor, specifically, for use as a ruminant feed additive.
  • the inventors of the present application found that the yield and purity of the product of the acetohydroxamic acid prepared by using ethyl acetate and hydroxylamine hydrochloride in a basic solvent are high, for example, the molar yield can be as high as 80% or more, and the purity is 98% or more. Moreover, the raw materials used in the method are cheap and easy to obtain, the reaction conditions are mild, and the operation steps are simple.
  • the synthesis method of the present invention can realize industrial mass production of acetohydroxamic acid and still ensure high yield and high purity.
  • the solvent used in the production process can be recycled and the preparation method is green economy.
  • the synthesis method of the present invention can obtain high purity and good product properties by simple extraction of the extractant ethyl acetate without complicated purification steps, and the obtained product can be directly used as a ruminant feed additive.
  • Figure 1 is a nuclear magnetic spectrum of acetohydroxamic acid.
  • Figure 2 is a graph showing the inhibition of rumen bacterial urease activity by acetohydroxamic acid.
  • the reaction system is based on a final product yield of 1000 g: in this example, the amount of hydroxylamine hydrochloride is 1300 g (18.7 mol), the amount of ethyl acetate is 1978 g (86.3 mmol), the amount of methanol is 2.6 L, and the amount of water is 2.6 L.
  • the progress of the reaction was monitored by UV lamp; the purity of the compound was detected by liquid chromatography.
  • the purity of the product was 98.0%.
  • the structure of the product was confirmed by nuclear magnetic resonance spectroscopy and carbon spectroscopy. As shown in Figure 1, the product obtained was shown to be acetohydroxamic acid.
  • the reaction system the amount of hydroxylamine hydrochloride is 5.0g (71.9mmol), the amount of ethyl acetate is 7.0g (79.1mmol), the amount of sodium hydroxide is 5.75g (143.75mmol), the amount of methanol is 10ml, and the amount of water is 10ml;
  • the ratio of each component: hydroxylamine hydrochloride: ethyl acetate: sodium hydroxide: methanol: water 1.0 eq: 1.1 eq: 2.0 eq: 2v: 2v (where eq is a molar ratio; v is a volume ratio).
  • the amount of hydroxylamine hydrochloride is 5.0g (71.9mmol)
  • the amount of ethyl acetate is 7.0g (79.1mmol)
  • the amount of methanol is 10ml
  • the amount of water is 10ml
  • the amount of sodium methoxide is 7.76g (143.75mmol) or hydrogen.
  • the reaction system the amount of hydroxylamine hydrochloride is 5.0 g (71.9 mmol), the amount of ethyl acetate is 7.0 g (79.1 mmol), the amount of methanol is 10 ml, the amount of water is 10 ml, and the amount of sodium hydroxide is 5.75 g (143.75 mmol);
  • the ratio of each component: hydroxylamine hydrochloride: ethyl acetate: water: methanol: NaOH 1.0 eq: 1.1 eq: 2v: 2v: 2.0 eq (where eq is a molar ratio; v is a volume ratio).
  • Table 4 shows the effect of different reaction temperatures on AHA yield.
  • the reaction system the amount of hydroxylamine hydrochloride was 5.0 g (71.9 mmol), and the amounts of ethyl acetate were 7.0 g (79.1 mmol), 7.6 g (86.3 mmol) and 8.2 g (93.5 mmol), respectively, and the amount of methanol was 10 ml.
  • reaction conditions of the present invention facilitate the progress of the reaction, and a product having high yield and high purity can be obtained.
  • the inhibitory effect of the product acetohydroxamic acid on the rumen urease activity of ruminants is expressed by the IC50 value, which is the concentration of acetohydroxamic acid which inhibits 50% urease activity under certain reaction conditions.
  • fresh rumen fluid was taken, filtered through four layers of sterile gauze, and the filtrate was subjected to differential centrifugation (500 ⁇ g centrifugation for 20 min and 12000 ⁇ g centrifugation for 15 min) at 4 ° C to obtain liquid phase rumen microorganisms.
  • the feed residue was repeatedly washed twice with 50 mmol/l HEPES buffer (4-hydroxyethylpiperazineethanesulfonic acid) (pH 7.5) to obtain a solid phase rumen microorganism.
  • HEPES buffer (4-hydroxyethylpiperazineethanesulfonic acid)
  • the solid and liquid microorganisms were mixed and added to a buffer of PBS (phosphate buffered saline) (pH 7.5) under low temperature conditions (40% strength, three times, 30 seconds each time), and the supernatant was used as a crude enzyme extract.
  • AHA inhibits the mapping of urease activity curves.
  • urea 50 mmol
  • water bathed at 37 ° C for 30 min followed by the addition of 1.5 ml of sodium nitrophenolate and 1.5 ml of NaClO-NaOH.
  • the above reaction system was subjected to a water bath at 37 ° C for 30 min, and then the light absorption value was measured at 625 nm.
  • the protein content in the crude enzyme extract was determined by Bradford Kit (Beijing Kangrun Chengye Biotechnology Co., Ltd., Beijing, China).
  • Urease activity is defined as the nanomolar amount of ammonia (nmol/min.mg) produced per minute per milligram of crude enzyme protein.
  • Inhibitory activity 1 - urease activity of AHA added / urease activity without AHA added ⁇ 100%.
  • the experimental results are shown in Fig. 2. It can be seen from the results of Fig. 2 that acetohydroxamic acid has an inhibitory effect on urease in the crude enzyme extract of rumen fluid of dairy cows, and the IC50 value of acetohydroxamic acid is 5.9 under the current reaction conditions. mM, similar to 5.34 mM reported by Jones (1968).

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Polymers & Plastics (AREA)
  • Organic Chemistry (AREA)
  • Animal Husbandry (AREA)
  • Zoology (AREA)
  • Engineering & Computer Science (AREA)
  • Food Science & Technology (AREA)
  • Birds (AREA)
  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)

Abstract

L'invention concerne un procédé de synthèse d'un acide acétohydroxamique inhibiteur d'uréase. Le procédé consiste à : faire réagir du chlorhydrate d'hydroxylamine avec de l'acétate d'éthyle dans un système de solvant alcalin, et procéder à l'extraction du résultat, après l'achèvement de la réaction, avec un solvant organique, CH3COOC2H5+NH2OH·HCl→CH3CONHOH. Le procédé de synthèse permet d'obtenir la production en masse à l'échelle industrielle d'acide acétohydroxamique, et peut toujours garantir un haut rendement et une pureté élevée de celui-ci, le solvant utilisé dans le procédé de production pouvant être recyclé. Le procédé de préparation est écologique et économique.
PCT/CN2018/095475 2018-05-15 2018-07-12 Procédé de synthèse d'acide acétohydroxamique inhibiteur d'uréase Ceased WO2019218445A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN201810463175.4A CN110483336A (zh) 2018-05-15 2018-05-15 一种脲酶抑制剂乙酰氧肟酸的合成方法
CN201810463175.4 2018-05-15

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WO2019218445A1 true WO2019218445A1 (fr) 2019-11-21

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116606222A (zh) * 2023-05-29 2023-08-18 铜仁贵江科技有限责任公司 一种乙酰氧肟酸合成工艺及其应用
CN118206469A (zh) * 2024-05-21 2024-06-18 峰成医药科技(天津)有限公司 一种3-羟基-4-（三氟甲基）苯甲腈的合成方法

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1384097A (zh) * 2002-05-24 2002-12-11 陕西富士达农业科技有限公司 乙酰氧肟酸的制备工艺
CN102976968A (zh) * 2012-12-01 2013-03-20 张家港市大伟助剂有限公司 一种甲氧胺盐酸盐的制备方法
CN104529815A (zh) * 2014-12-04 2015-04-22 宁波欧迅化学新材料技术有限公司 合成2,4-二硝基苯氧胺的方法
CN106800580A (zh) * 2017-01-12 2017-06-06 中国药科大学 甾醇类衍生物及其制备方法和应用

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1384097A (zh) * 2002-05-24 2002-12-11 陕西富士达农业科技有限公司 乙酰氧肟酸的制备工艺
CN102976968A (zh) * 2012-12-01 2013-03-20 张家港市大伟助剂有限公司 一种甲氧胺盐酸盐的制备方法
CN104529815A (zh) * 2014-12-04 2015-04-22 宁波欧迅化学新材料技术有限公司 合成2,4-二硝基苯氧胺的方法
CN106800580A (zh) * 2017-01-12 2017-06-06 中国药科大学 甾醇类衍生物及其制备方法和应用

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
LIU, QIANG ET AL.: "Synthesis of O- Benzylhydroxylamine Hydrochloride from Polyvinyl Alcohol Alcoholysis Liquor", CHINESE JOURNAL OF SYNTHETIC CHEMISTRY, vol. 21, no. 5, 31 December 2013 (2013-12-31), pages 611 - 614, XP055653857 *
SU , LANHUI ET AL.: "Study on Synthetic Technics and Conditions of Acetohy-Droxamic Acid", CHINA FEED, no. 14, 31 December 2005 (2005-12-31), pages 15 - 16, 24 *
SU , LANHUI ET AL.: "Synthesis of Acetohydroxamic Acid", FINE AND SPECIALTY CHEMICALS, vol. 14, no. 3/4, 21 February 2006 (2006-02-21), pages 18 - 19, 22 *
SUN, GUANGQIANG ET AL.: "Synthesis of Cyclohexenone Herbicide-Clethodim", MODERN AGROCHEMICALS, vol. 10, no. 1, 28 February 2011 (2011-02-28), pages 24 - 26, XP055653851 *

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