16,17α-epoxy progesterone application

16,17α-epoxyprogesterone is used as an intermediate in the manufacture of almost all steroid hormones, including dexamethasone, betamethasone, 6α-methyl prednisone, prednisone, and prednisone. Steroid hormone drugs are an important class of drugs second only to antibiotics. They have various functions such as anti-inflammatory, anti-immune, contraceptive, anti-cancer, and endocrine regulation. They are widely used in rheumatism, cardiovascular diseases, collagen diseases, and lymphatic leukemia. , Human organ transplantation, bacterial encephalitis, skin diseases, endocrine disorders, senile diseases, hormone-dependent tumors and other diseases.

Using digestrel acetate as a raw material, it is oxidized with hydrogen peroxide to form 3-hydroxy-5ene-16,17α-epoxy. Through Oppenauer oxidation, 3-hydroxy-5ene is converted into 3-keto-4ene structure. The 16,17α-epoxyprogesterone is finally obtained by steam distillation, extraction, concentration, and purification. The limiting factor of this technological level is that the effective reaction of Oppenauer oxidation is relatively low, resulting in many by-products in the reaction process, which affects the conversion rate and yield.

At present, with 100Kg of digestrel acetate, only 70～73Kg of 16,17α-epoxyprogesterone can be obtained. In addition, traditional chemical processes use a lot of organic solvents, the reaction conditions are severe, and steam distillation has to be used. The device causes high energy consumption and high emissions, and the Oppenauer oxidation reaction of 16,17α-epoxyprogesterone has become a bottleneck in the development of the entire steroid drug industry. Therefore, it provides a method that can increase the yield of 16,17α-epoxyprogesterone. And the method of mild response and green manufacturing has become an urgent problem to be solved.

Application[2][3]

16,17α-epoxyprogesterone is used as an intermediate in almost all steroid hormones including dexamethasone, 16-DPA, betamethasone, 6α-methyl prednisone, prednisone, prednisone, etc. Manufacturing. Examples of its applications are as follows:

1. Used for 11α hydroxylation of an important intermediate of steroid hormones.

Corticosteroids are a class of steroid compounds synthesized and secreted by the adrenal cortex. Its main function is to regulate water and salt metabolism and sugar metabolism in animals. The C11α hydroxylation of steroids is an indispensable step in the synthesis of various corticosteroids. The hydroxyl groups on steroids are very difficult for chemical synthesis. Except for the C17 position of steroids, it is easy to introduce a hydroxyl group, and other positions are difficult to introduce chemically.

Steroid C11α hydroxylation is a unique conversion reaction of microorganisms. This not only solves the problem of difficult drug synthesis and synthesis, but also increases the activity of corticosteroid compounds and makes them have a highly inhibitory effect of inflammation. A new way to find more effective new drugs has been created.

The preparation method includes the following steps: the first step is the propagation of the strain: the Aspergillus ochrae or the black rhizozyme strain is connected to the corresponding seed medium for cultivation; the second step is the emulsification of the substrate: the substrate 17α-hydroxy One of progesterone, 4-androstenedione or 16,17-α epoxy progesterone is emulsified under the action of surfactant; the third step, 16-DPA, fermentation culture: get from the first step After cultivating the Aspergillus oryzae spp. in the corresponding fermentation medium for a period of time, add sterilized 17α-hydroxyprogesterone, 4-androstenedione or 16,17-αepoxyprogesterone The emulsified liquid of one of them continues to be fermented and cultivated; the fourth step is to extract the finished product.

2. Synthesize mold oxide,

It includes the following steps:

(1) Preparation of mycelium: A. Add carbon source: glucose 60～85kg; nitrogen source: corn steep liquor 60～85kg; silkworm pupa powder 10～20kg; ammonium sulfate 5～10kg; defoamer: polyether 1000～2000ml once Add 1-4 tons of water to the first-level fermentation tank, adjust the PH value to 2-5, steam sterilize, and when the temperature is lowered to 26±4℃, connect 3-50 liters of spore suspension of Rhizopus nigricans to suspend The concentration of the solution is 100,000 to 3 million/ml, and air is introduced, stirred for 12-36h, and then transferred to the secondary fermentation tank;

B. Put 5-10 times the material according to the same material ratio as the first-level fermentation tank, sterilize, cool to 26±4℃, transfer all the liquid produced in step A, blow in air, stir for 18-36h, and put in 340kg Substrate Wohren's oxide, biological oxidation, sampling and microscopic examination, and chromatographic test at 25 ~ 30 ℃, after the substrate conversion ratio reaches 50-60%, raise the temperature to 30-60 ℃, discharge the bottom tank; C 、Press the mycelium liquid into the plate and frame mixer, filter out the fermentation liquid, air-dry the mycelium, and then crush the mycelium;

(2) Preparation of crude mold oxide: Put the pulverized mycelium into an extraction tank, add 5-10 times the weight of mycelium in acetone, extract 2-8 times, and concentrate the extract to crystallize to obtain crude mold oxide. ；

(3) Refining of mold oxide: Put the crude mold oxide into the refining tank, add a mixed solvent of toluene and chloroform of 10-15 times the weight of the crude mold oxide, and concentrate the whole solution. Repeat the refining 3-9 times and sample for testing. After the mold oxide content is greater than 97%, place the material in the bottom tank, drain the product, and dry the product in the oven, 16-DPA, sample and submit for inspection. The mold oxide content greater than 97% is mold oxide.

Preparation [1][4]

method 1:

1) Add sodium hydroxide and hydrogen peroxide in proportion to the epoxidation reaction. Acetic acid is comprehensively concentrated, and the toluene extract is azeotropically dehydrated.

2) Add aluminum isopropoxide and cyclohexanone in proportion to the Wöhren's oxidation reaction. Ethanol refined to obtain 16,17α-epoxyprogesterone.

Method 2: A microbial fermentation of 3-hydroxy-5ene-16,17α-epoxy compound to produce 16,17α-epoxy progesterone with mild reaction conditions, green and environmental protection, and can increase the yield of 16,17α-epoxyprogesterone Ketone method.

Biological fermentation of 3-hydroxy-5ene-16,17α-epoxy compound with Arthrobacter simplex Arthrobacter-simplex-AS1.94's own cholesterol oxidase to realize the biological production of 16,17α-epoxyprogesterone, instead of traditional The chemical process improves the conversion and yield of 16,17α-epoxyprogesterone. Among them, the substrate is 3-hydroxy-5-ene-16,17α-epoxy, Arthrobacter-simplex-AS1.94 is a catalytic strain, the yield of 16,17α-epoxyprogesterone after fermentation is greater than 90%, and the purity is Greater than 98.5%.

Main reference materials

[1] Zhang Anan; Li Jiuzhou; Bie Songtao. Biological method for preparing 16,17α-epoxyprogesterone CN201611263298.0, 16-DPA, application date 20161230

[2] Song Haolei; Zhang Yaoxun; Zhang Minran; Yang Ke. A method for preparing 11α hydroxylation of important intermediates of steroid hormones. CN201410174391.9, application date 20140428

[3] Liu Caitian; Zhao Youjie. New process for preparing mold oxide by mold fermentation biosynthesis. CN201110031709.4, application date 201110130

[4] Dioscorea chrysanthemum