Pure vitamin D3 product and preparation process thereof

Abstract

The invention provides a vitamin D3 pure product and a preparation process thereof. The provided preparation process comprises the following steps: photochemical reaction, column chromatography separation and thermal isomerization reaction. According to the method, ultraviolet light sources with different wavelengths of a photochemical reactor are utilized to promote the raw material 7-dehydrocholesterol to be subjected to photochemical ring-opening reaction and photocatalytic isomerization reaction, and after photochemical isomerization, chromatographic purification is performed, and then thermal isomerization reaction is performed to obtain the vitamin D3. The preparation process provided by the invention can be used for obtaining a pure vitamin D3 product which has needle-like crystals, high yield, high crystal refining yield and high purity and conforms to pharmacopoeia regulations, and is a novel process which is high in raw material conversion rate, few in by-products and more efficient and environment-friendly in production.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical chemistry, and in particular to a pure vitamin D3 product and a preparation process thereof. Background Art

[0002] Vitamin D3 is a sterol derivative, also known as cholecalciferol, which is commonly used for the prevention and treatment of vitamin D deficiency. It is a fat-soluble vitamin that can be dissolved in organic matter and is an indispensable vitamin for the growth, development, reproduction and maintenance of health of humans and animals. Its main physiological role in the human body and animals is to promote the absorption of calcium and phosphorus, ensure the supply of calcium and phosphorus in the body and the calcification of bones, and maintain the balance of blood calcium and blood phosphorus.

[0003] Vitamin D3 in humans and animals is mainly obtained through the photochemical reaction of 7-dehydrocholesterol in the skin under the irradiation of ultraviolet rays in sunlight. In real life, the vitamin D3 we take mainly comes from two sources: one is obtained from the liver of deep-sea fish, such as cod liver; the other is obtained by chemically opening the ring of 7-dehydrocholesterol through light.

[0004] CN114230500A discloses a method for preparing vitamin D3, which includes a primary photochemical reaction, a secondary photochemical reaction and a thermal isomerization reaction. In the primary photochemical reaction, ultraviolet light with a wavelength of less than 290 nm is absorbed by a specific filter liquid, and in the secondary photochemical reaction, a specific filter liquid is added to absorb ultraviolet light with a wavelength of less than 330 nm, thereby increasing the content of vitamin D3 obtained after the thermal isomerization reaction.

[0005] CN114369047A discloses a method for crystallizing vitamin D3. The method provided by the invention comprises the following steps: a) dissolving resinous vitamin D3 oil in an organic solvent, then adding activated carbon for adsorption treatment, and then filtering to obtain a filtrate; b) cooling the filtrate, adding vitamin D3 seed crystals, and then performing gradient cooling crystallization, and then performing solid-liquid separation and drying to obtain needle-shaped vitamin D3 crystals; the gradient cooling crystallization comprises: firstly cooling the temperature to 5-15°C under stirring conditions and maintaining stirring for crystallization until needle-shaped crystals precipitate, then stopping stirring; and then continuing to cool to -20-0°C and standing for crystallization.

[0006] The prior art vitamin D3 preparation process usually involves first dissolving the raw material 7-dehydrocholesterol in a solvent for a light-induced ring-opening reaction in a photoreactor, then flowing the reaction solution into a second light-induced reactor for a transposition reaction and then a heating isomerization reaction, and then performing chromatography separation or no chromatography separation after obtaining vitamin D3 resin oil. Due to the problem of wide illumination spectrum and impurities not being separated in time, the general vitamin D3 yield is low and there is a lot of waste, which seriously affects the overall benefits of the enterprise. Summary of the invention

[0007] In view of the above-mentioned defects of the prior art, the present invention provides a preparation process of pure vitamin D3. The preparation process provided by the present invention can be used to prepare pure vitamin D3 that meets the requirements of the pharmacopoeia and has needle-shaped crystals, high yield, high crystal refining yield and high purity.

[0008] The present invention provides a preparation process of pure vitamin D3, which comprises the following steps, calculated by weight:

[0009] (1) adding 9 to 12 parts by weight of 7-dehydrocholesterol to 300 to 400 parts by weight of a solvent, heating the mixture to 38 to 43° C. to dissolve the mixture, and then adding 0.5 to 1.5 parts by weight of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; opening a photochemical reactor, introducing a first ultraviolet light filtering circulating liquid, and then introducing the 7-dehydrocholesterol solution into the photochemical reactor, irradiating the solution with an ultraviolet light source at a flow rate of 8 to 10 L / min, a reaction temperature of 25 to 40° C. and nitrogen to perform a first photochemical reaction, and then introducing a second ultraviolet light filtering circulating liquid, and then introducing the first ultraviolet light filtering circulating liquid into the photochemical reactor. The chemical reaction liquid is introduced into a photochemical reactor, and a second photochemical reaction is carried out under the conditions of an introduction flow rate of 8 to 10 L / min, a reaction temperature of 25 to 40° C. and nitrogen by irradiation with an ultraviolet light source to obtain a second photochemical reaction liquid, and the conversion rate is controlled at 50% to 65%; the second photochemical reaction liquid is collected, and 80 to 90% of the solvent is recovered by reduced pressure distillation at 20 to 30° C., and then 7-dehydrocholesterol raw materials are recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10° C. to -5° C. for 10 to 12 hours, and then filtered under reduced pressure, and the solvent is recovered from the filtrate to obtain a mixture of pre-vitamin and vitamin D3, that is, a vitamin mixture;

[0010] (2) mixing and dissolving the vitamin mixture obtained in the above step (1) with an adsorption solvent in a weight ratio of 1:5-10 to obtain a mixed solution; adding the mixed solution to a chromatography column, eluting and separating with an eluting solvent, collecting in sections, tracking with a high performance liquid chromatography analysis method until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component;

[0011] (3) The high-content pre-vitamin component obtained in the above step (2) is heated at a temperature of 40°C to 65°C and a vacuum degree of -0.070 to 0.095 MPa for 3 to 8 hours to perform a thermal isomerization reaction until the pre-vitamin is converted into vitamin D3, and then concentrated by reduced pressure distillation to obtain vitamin D3 oil; the vitamin D3 oil and the crystallization solvent are mixed and dissolved in a weight ratio of 1:5-10 to obtain a dissolving solution; the dissolving solution is frozen and crystallized at -8 to 5°C for 8 to 20 hours, and then filtered and dried to obtain pure vitamin D3.

[0012] Preferably, the solvent in step (1) is at least one selected from a mixed solvent of C3-C3 alcohol, C3-C3 alcohol and C5-C7 alkane.

[0013] Further preferably, the solvent is selected from at least one of methanol, ethanol, a mixed solvent of methanol and n-hexane, a mixed solvent of ethanol and n-hexane, and a mixed solvent of ethanol and n-heptane.

[0014] Further preferably, among the solvents, the mixed solvent of methanol and n-hexane is a mixture of methanol and n-hexane in a volume ratio of 2.5:(1 to 1.2); the mixed solvent of ethanol and n-hexane is a mixture of ethanol and n-hexane in a volume ratio of 2.5:(1 to 1.2); and the mixed solvent of ethanol and n-heptane is a mixture of ethanol and n-heptane in a volume ratio of 2.5:(1 to 1.2).

[0015] Preferably, the first ultraviolet light filtering circulating liquid in step (1) is a self-prepared copper sulfate aqueous solution with a concentration of 1.5 to 7 wt %; the second ultraviolet light filtering circulating liquid is a self-prepared copper sulfate aqueous solution with a concentration of 4.5 to 10 wt %.

[0016] Preferably, in the first photochemical reaction in step (1), the 7-dehydrocholesterol solution undergoes a photochemical reaction under the irradiation of an ultraviolet light source with a wavelength range of 280 to 300 nm; and in the second photochemical reaction, the first photochemical reaction solution undergoes a photochemical reaction under the irradiation of an ultraviolet light source with a wavelength range of 310 to 350 nm.

[0017] Further explained, the reaction process of obtaining the pure vitamin D3 from the raw material 7-dehydrocholesterol through photochemical reaction and thermal isomerization reaction includes the following two steps:

[0018] A. Photo-ring-opening reaction of raw material 7-dehydrocholesterol: 7-dehydrocholesterol is dissolved in a solvent to obtain a raw material solution; the raw material solution is pre-adjusted to a reaction temperature, and then irradiated with an ultraviolet light source with a wavelength range of 280 to 300 nm under nitrogen protection to obtain a solution containing a ring-opening reaction intermediate, i.e., the first photochemical reaction solution;

[0019] B. Photocatalytic isomerization purification reaction: The first photochemical reaction liquid is irradiated with an ultraviolet light source in the wavelength range of 310 to 350 nm under nitrogen protection to perform photocatalytic isomerization, thereby obtaining a solution mainly containing pre-vitamin D3; the solution mainly containing pre-vitamin D3 is subjected to chromatographic separation, and then heated to perform hydrogen shift isomerization, thereby obtaining pure vitamin D3.

[0020] The present invention utilizes ultraviolet light sources of different wavelengths in a photochemical reactor to promote photochemical ring-opening reaction and photocatalytic isomerization reaction of the raw material 7-dehydrocholesterol, and after photochemical isomerization, chromatography purification is performed and then thermal isomerization reaction is performed to obtain a pure vitamin D3 product that meets the requirements of the pharmacopoeia and has needle-shaped crystals, a relatively high yield, a high crystal refining yield and high purity.

[0021] Preferably, the adsorption solvent in step (2) is selected from at least one of C6 to C12 alkanes.

[0022] Further preferably, the adsorption solvent is selected from at least one of n-hexane and n-heptane.

[0023] Preferably, the stationary phase of the chromatography column in step (2) is selected from silica gel and activated alumina.

[0024] Preferably, the elution solvent in step (2) is selected from at least one of C6 to C9 alkanes.

[0025] Further preferably, the elution solvent is selected from at least one of n-hexane and n-heptane.

[0026] Preferably, the crystallization solvent in step (3) is selected from at least one of C2 to C9 ester solvents.

[0027] Further preferably, the crystallization solvent is methyl acetate.

[0028] In the photochemical reaction of step (1) of the present invention, the photochemical product previtamin generated in the photochemical reaction will absorb light again to generate a secondary photochemical reaction to generate byproducts leucosterol and tachysterol. If the conversion rate of 7-dehydrocholesterol is too high, the byproducts generated by the secondary photochemical reaction will increase greatly, which not only reduces the yield of vitamin P3, but also increases the difficulty of separation and purification. Therefore, the present invention controls the conversion rate of 7-dehydrocholesterol to 50% to 65%, so as to obtain a higher photochemical product vitamin mixture and improve selectivity, and at the same time, the concentration of byproducts can be controlled at a lower level.

[0029] The vitamin mixture obtained in step (1) of the present invention contains a high content of previtamin, vitamin D3, tachysterol and unreacted 7-dehydrocholesterol. Since the properties of previtamin and vitamin D3, tachysterol and 7-dehydrocholesterol are very different, column chromatography is used for separation in step (2) of the present invention to achieve a good separation effect. In the column chromatography step, the adsorbent is very important for column chromatography separation. The present invention preferably uses silica gel and alumina as the stationary phase of column chromatography, and also optimizes the adsorption solvent and elution solvent in the column chromatography solvent system, which is conducive to improving the separation efficiency.

[0030] Preferably, the step (3) can also be: the high-content pre-vitamin component obtained in the above step (2) is heated for 3 to 8 hours at a temperature of 40°C to 65°C and a vacuum degree of -0.070 to 0.095 MPa to perform a thermal isomerization reaction until the pre-vitamin is converted into vitamin D3, and then concentrated by reduced pressure distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with n-hexane in a weight ratio of 1:20 to 40 to obtain a vitamin D3 solution, and then the vitamin D3 solution is mixed and stirred with an adsorbent in a weight ratio of 1:0.1 to 0.3 for 15 to 25 minutes; and then the adsorbent is removed by a handheld magnet. The adsorbent is separated from the solution and washed with n-hexane, and the separated adsorbent is mixed with acetonitrile in a weight ratio of 1:50 to 100, and ultrasonically treated for 5 to 10 minutes to desorb vitamin D3; the adsorbent is then separated from the solution with a magnet and washed with acetonitrile; the acetonitrile solution containing the desorbed vitamin D3 is filtered through a polytetrafluoroethylene membrane with a pore size of 0.45 μm, the filtrate is collected, and concentrated under reduced pressure to obtain vitamin D3 oil; the vitamin D3 oil is mixed and dissolved with a crystallization solvent in a weight ratio of 1:5 to 10 to obtain a dissolved solution; the dissolved solution is frozen and crystallized at -8 to 5°C for 8 to 20 hours, and then filtered and dried to obtain pure vitamin D3.

[0031] Preferably, the adsorbent is a magnetic adsorbent, and its preparation method comprises the following steps, measured by weight:

[0032] 2 to 3 parts by weight of FeSO 4 7H 2O is added to 130 to 180 parts by weight of a graphene oxide aqueous suspension having a concentration of 1.5 to 3 mg / mL, and then 0.06 to 0.08 parts by weight of lycopodium spore pollen microcapsules and 10 to 15 parts by weight of anhydrous ethanol are added, mixed and stirred to obtain a mixture; the mixture is ultrasonically treated for 30 to 50 minutes, and then a 20 to 30 wt% ammonia aqueous solution is used to adjust the pH to 10 to 12, and then heated at 85 to 95° C. for 7 to 10 hours to obtain a magnetic hydrogel; the magnetic hydrogel is separated from the solution using a handheld magnet, and washed with distilled water, and then the hydrogel is placed at 18 to 22° C. until the gel is solidified, and then freeze-dried for 45 to 50 hours to obtain a magnetic adsorbent;

[0033] Or, 2 to 3 parts by weight of FeSO 4 7H 2 O is added to 130 to 180 parts by weight of a graphene oxide aqueous suspension with a concentration of 1.5 to 3 mg / mL, mixed and stirred to obtain a mixture; the mixture is ultrasonically treated for 10 to 20 minutes, and then a 20 to 30 wt% ammonia aqueous solution is used to adjust the pH to 10 to 12, and then heated at 85 to 95° C. for 4 to 8 hours to obtain a magnetic hydrogel; the magnetic hydrogel is separated from the solution by using a handheld magnet, and washed with distilled water, and then the hydrogel is placed at 18 to 22° C. until the gel is solidified, and then freeze-dried for 22 to 26 hours to obtain a magnetic adsorbent.

[0034] Further preferably, the preparation method of the lycopodium spore pollen microcapsules comprises the following steps: mixing lycopodium spore powder and acetone in a weight ratio of 1:5 to 10, filtering after refluxing and stirring at 45 to 60°C for 1.5 to 3 hours, washing the filtrate with ethanol and water until the filtrate is clear, and then drying to obtain dried lycopodium spores; mixing the dried lycopodium spores with a 0.1 to 0.3 g / mL phosphoric acid aqueous solution in a weight ratio of 1:10 to 20, filtering after refluxing and stirring at 70 to 80°C for 7 to 10 hours, washing with water and then drying to obtain lycopodium spore pollen microcapsules.

[0035] The present invention comprises the following steps: 4 7H 2O, graphene oxide and lycopodium spore pollen microcapsules are mixed, and after ultrasonic treatment, they are heated to obtain magnetic hydrogel, and the magnetic hydrogel is freeze-dried after solidification to prepare a magnetic adsorbent. The inventors found that the introduction of a magnetic adsorbent in the preparation process of pure vitamin D3 for adsorption operation can significantly improve the yield, crystal refining yield and purity of pure vitamin D3. The reason for the analysis may be that the magnetic adsorbent has a higher specific surface area and a porous structure, which enables it to provide a large number of adsorption sites, and has high selectivity for vitamin D3, while the adsorption capacity for impurities is weak. This high adsorption capacity and selective adsorption help to improve the recovery rate of vitamin D3, and then improve the refining yield and purity of the crystal. Among them, the introduction of lycopodium spore pollen microcapsules may provide additional adsorption sites or improve the physicochemical properties of the magnetic adsorbent, thereby improving the efficiency of adsorption and desorption of the adsorbent. In addition, magnetic adsorbents can be quickly separated using an external magnetic field. This property allows the adsorbent to be quickly removed from the solution after the adsorption process is completed, reducing the loss of vitamin D3 in traditional separation steps such as filtration and centrifugation, thereby increasing the yield.

[0036] The present invention also provides a pure vitamin D3 product, which is prepared by the above-mentioned preparation process.

[0037] Beneficial effects of the present invention:

[0038] 1. Compared with the prior art, the advantage of the present invention is that after the photochemical reaction, the present invention does not obtain vitamin D3 resin oil through post-treatment and thermal isomerization process and then conduct subsequent separation, but fully utilizes the thermal isomerization equilibrium of vitamin D3 and pre-vitamin and the large difference in properties between pre-vitamin and other isomers. First, a high content of pre-vitamin is obtained by column chromatography, and then high-purity pure vitamin D3 crystals that meet the requirements of the pharmacopoeia are obtained by thermal isomerization and crystallization. The reaction and separation efficiency are improved, which is higher than the yield of the prior art of first obtaining vitamin D3 resin oil and then conducting subsequent separation process. It has the advantages of simple process, low cost and no environmental pollution, and can fully achieve the purpose of green chemical industry.

[0039] 2. In order to prevent the open-ring compound generated in the photochemical reaction from generating leucosterol intermediates and tachysterol intermediates under illumination conditions, thereby affecting the generation of pre-vitamin D3, due to the high-pressure mercury lamp used in the photochemical reaction having a wide spectrum and containing wavelengths that are unfavorable to the reaction of the present invention, the present invention configures ultraviolet light filtering circulating fluids of different concentrations to absorb ultraviolet light below 280nm and above 310nm, and ultraviolet light below 300nm and above 360nm, respectively, so as to reduce the light that is unfavorable for maintaining the reaction balance of the compound pre-vitamin D3 in the reaction system, so as to adjust the reaction balance of the photochemical reaction and shift the reaction balance toward the direction of generating the compound pre-vitamin D3, thereby maximizing the proportion of pre-vitamin D3 in the photochemical reaction.

[0040] Furthermore, since the properties of the previtamin after the photochemical reaction are very different from those of vitamin D3, tachysterol, and 7-dehydrocholesterol, the present invention uses column chromatography for separation in the preparation process step (2), thereby achieving a good separation effect and greatly improving the yield and quality of vitamin D3.

[0041] 3. Compared with the prior art, the preparation process of pure vitamin D3 of the present invention has the following advantages: ① By selecting filter liquids of different concentrations for ultraviolet light treatment and then irradiating the mixture, the selectivity of the pre-vitamin and vitamin D3 is increased, which is significantly better than the result of general ultraviolet light photochemical reaction; ② The present invention preferably uses silica gel and alumina as the stationary phases of column chromatography, respectively, and performs column chromatography separation on the photochemical reaction products and the isomerized products, and at the same time optimizes the column chromatography separation solvent system to improve the complete separation of each component; ③ The obtained vitamin mixture is first subjected to column chromatography to obtain a high content of pre-vitamin, and then high-purity pure vitamin D3 crystals that meet the requirements of the pharmacopoeia are obtained by thermal isomerization and crystal dissolution chromatography; ④ Introducing a magnetic adsorbent for adsorption operation during the preparation process improves the yield of pure vitamin D3, the crystal refining yield and the purity. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Attached Figure 1 The reaction process of obtaining vitamin D3 by photochemical reaction and thermal isomerization reaction of 7-dehydrocholesterol in Example 1 of the present invention;

[0043] Attached Figure 2 This is a liquid chromatogram of the vitamin D3 standard and sample for the vitamin D3 content detection in Example 6 of the present invention. DETAILED DESCRIPTION

[0044] Parameters for specific chemical substances used, sources.

[0045] Graphene oxide, brand: Zhenzhun, item number: MS-21005, diameter: 3μm, thickness: 1nm, purity: 99%;

[0046] Lycopodium spore powder, particle size: 20μm, content: 99%.

[0047] Example 1

[0048] A preparation process for pure vitamin D3, comprising the following steps:

[0049] (1) Add 10 kg of 7-dehydrocholesterol (95 wt%) to a solvent formed by mixing 250 L of methanol and 110 L of normal hexane, heat to 40° C., mix and dissolve, then add 0.8 kg of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; turn on a photochemical water cooler, nitrogen, a circulating water pump, an ultraviolet lamp, and a 5 wt % copper sulfate aqueous solution, turn on a magnetic pump, and pass the 7-dehydrocholesterol solution into a photochemical reactor, and irradiate with an ultraviolet light source with a wavelength of 290 nm under a flow rate of 8.5 L / min, a reaction temperature of 35° C., and nitrogen conditions to perform a first photochemical reaction to obtain a first photochemical reaction solution; then pass a 9 wt % copper sulfate aqueous solution into the reactor, Then, the first photochemical reaction liquid is introduced into a photochemical reactor, and a second photochemical reaction is carried out by irradiation with an ultraviolet light source with a wavelength of 330 nm under the conditions of an introduction flow rate of 8.5 L / min, a reaction temperature of 35° C. and nitrogen, to obtain a second photochemical reaction liquid, and the detection conversion rate of the second photochemical reaction liquid is 58%; then, the second photochemical reaction liquid is collected, and 80% of the solvent (n-hexane and methanol) is recovered by reduced pressure distillation at 30° C., and then the 7-dehydrocholesterol raw material is recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10° C. for 10 hours, and then filtered under reduced pressure to obtain a solution containing a large amount of pre-vitamins, and the filtrate is completely recovered to obtain a vitamin mixture;

[0050] (2) mixing and dissolving the obtained vitamin mixture with n-hexane in a weight ratio of 1:8 to obtain a mixed solution; adding the mixed solution to a silica gel chromatography column containing 100 kg, eluting and separating with n-hexane, collecting in sections, tracking with high performance liquid chromatography until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component;

[0051] (3) The obtained high-content pre-vitamin component is heated at a temperature of 55°C and -0.095Mpa for 6 hours to perform a thermal isomerization reaction to convert the pre-vitamin into vitamin D3, which is then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a dissolving solution; the dissolving solution is frozen and crystallized at -10°C for 12 hours, and then filtered and vacuum dried to obtain a pure vitamin D3 product.

[0052] Example 2

[0053] A preparation process for pure vitamin D3, comprising the following steps:

[0054] (1) Add 10 kg of 7-dehydrocholesterol (95 wt%) to a solvent formed by mixing 250 L of ethanol and 110 L of n-heptane, heat to 40° C., mix and dissolve, then add 0.8 kg of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; turn on a photochemical water cooler, nitrogen, a circulating water pump, an ultraviolet lamp, and a 6 wt % copper sulfate aqueous solution, turn on a magnetic pump, and pass the 7-dehydrocholesterol solution into a photochemical reactor, and irradiate with an ultraviolet light source with a wavelength of 290 nm under a flow rate of 8.5 L / min, a reaction temperature of 35° C., and nitrogen conditions to perform a first photochemical reaction to obtain a first photochemical reaction solution; then pass a 9 wt % copper sulfate aqueous solution into the reactor, Then, the first photochemical reaction liquid is introduced into a photochemical reactor, and a second photochemical reaction is carried out by irradiation with an ultraviolet light source with a wavelength of 330 nm under the conditions of an introduction flow rate of 8.5 L / min, a reaction temperature of 35° C. and nitrogen, to obtain a second photochemical reaction liquid, and the detection conversion rate of the second photochemical reaction liquid is 56%; then, the second photochemical reaction liquid is collected, and 80% of the solvent (n-heptane and ethanol) is recovered by reduced pressure distillation at 35° C., and then the 7-dehydrocholesterol raw material is recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10° C. for 10 hours, and then filtered under reduced pressure to obtain a solution containing a large amount of pre-vitamins, and the filtrate is completely recovered to obtain a vitamin mixture;

[0055] (2) mixing and dissolving the obtained vitamin mixture with n-hexane in a weight ratio of 1:8 to obtain a mixed solution; adding the mixed solution to a silica gel chromatography column containing 100 kg, eluting and separating with n-hexane, collecting in sections, tracking with high performance liquid chromatography until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component;

[0056] (3) The obtained high-content pre-vitamin component is heated at a temperature of 55°C and -0.095Mpa for 6 hours to perform a thermal isomerization reaction to convert the pre-vitamin into vitamin D3, which is then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a dissolving solution; the dissolving solution is frozen and crystallized at -10°C for 10 hours, and then filtered and vacuum dried to obtain a pure vitamin D3 product.

[0057] Example 3

[0058] A preparation process for pure vitamin D3, comprising the following steps:

[0059] (1) adding 10 kg of 7-dehydrocholesterol (95 wt%) to a solvent formed by mixing 250 L of ethanol and 110 L of normal hexane, heating to 40° C. to mix and dissolve, and then adding 0.8 kg of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; turning on a photochemical water cooler, nitrogen, a circulating water pump, an ultraviolet lamp, and a 2.5 wt% copper sulfate aqueous solution, turning on a magnetic pump, and passing the 7-dehydrocholesterol solution into a photochemical reactor, and irradiating the 7-dehydrocholesterol solution with an ultraviolet light source having a wavelength of 290 nm at a flow rate of 10.0 L / min, a reaction temperature of 35° C., and nitrogen to perform a first photochemical reaction, and then passing a 5.5 wt% copper sulfate aqueous solution into the reactor. liquid, then the first photochemical reaction liquid is introduced into a photochemical reactor, and a second photochemical reaction is carried out under the conditions of an introduction flow rate of 10.0L / min, a reaction temperature of 35°C and nitrogen by irradiation with an ultraviolet light source with a wavelength of 330nm to obtain a second photochemical reaction liquid, and the detection conversion rate of the second photochemical reaction liquid is 58%; then the second photochemical reaction liquid is collected, and 80% of the solvent (n-hexane and ethanol) is recovered by vacuum distillation at 30°C, and then the 7-dehydrocholesterol raw material is recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10°C for 10h, and then filtered under reduced pressure to obtain a solution containing a large amount of pre-vitamins, and the filtrate is completely recovered to obtain a vitamin mixture;

[0060] (2) mixing and dissolving the obtained vitamin mixture with n-hexane in a weight ratio of 1:8 to obtain a mixed solution; adding the mixed solution to a silica gel chromatography column containing 100 kg, eluting and separating with n-hexane, collecting in sections, tracking with high performance liquid chromatography until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component;

[0061] (3) The obtained high-content pre-vitamin component is heated at a temperature of 55°C and -0.095 MPa for 6 hours to perform a thermal isomerization reaction to convert the pre-vitamin into vitamin D3, which is then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a dissolving solution; the dissolving solution is placed at -10°C for freezing and crystallization for 8 hours, and then filtered and vacuum dried to obtain a pure vitamin D3 product.

[0062] Example 4

[0063] A preparation process for pure vitamin D3, comprising the following steps:

[0064] (1) Add 10 kg of 7-dehydrocholesterol (95 wt%) to a solvent formed by mixing 250 L of methanol and 110 L of normal hexane, heat to 40° C., mix and dissolve, then add 0.8 kg of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; turn on a photochemical water cooler, nitrogen, a circulating water pump, an ultraviolet lamp, and a 5 wt % copper sulfate aqueous solution, turn on a magnetic pump, and pass the 7-dehydrocholesterol solution into a photochemical reactor, and irradiate with an ultraviolet light source with a wavelength of 290 nm under a flow rate of 8.5 L / min, a reaction temperature of 35° C., and nitrogen conditions to perform a first photochemical reaction to obtain a first photochemical reaction solution; then pass a 9 wt % copper sulfate aqueous solution into the reactor, Then, the first photochemical reaction liquid is introduced into a photochemical reactor, and a second photochemical reaction is carried out by irradiation with an ultraviolet light source with a wavelength of 330 nm under the conditions of an introduction flow rate of 8.5 L / min, a reaction temperature of 35° C. and nitrogen, to obtain a second photochemical reaction liquid, and the detection conversion rate of the second photochemical reaction liquid is 58%; then, the second photochemical reaction liquid is collected, and 80% of the solvent (n-hexane and methanol) is recovered by reduced pressure distillation at 30° C., and then the 7-dehydrocholesterol raw material is recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10° C. for 10 hours, and then filtered under reduced pressure to obtain a solution containing a large amount of pre-vitamins, and the filtrate is completely recovered to obtain a vitamin mixture;

[0065] (2) mixing and dissolving the obtained vitamin mixture with n-hexane in a weight ratio of 1:8 to obtain a mixed solution; adding the mixed solution to a silica gel chromatography column containing 100 kg, eluting and separating with n-hexane, collecting in sections, tracking with high performance liquid chromatography until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component;

[0066] (3) The obtained high-content pre-vitamin component was heated at a temperature of 55° C. and -0.095 MPa for 6 hours to perform a thermal isomerization reaction, and the pre-vitamin was converted into vitamin D3, and then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil was mixed and dissolved with n-hexane in a weight ratio of 1:30 to obtain a vitamin D3 solution, and then the vitamin D3 solution was mixed with a magnetic adsorbent in a weight ratio of 1:0.13, and stirred for adsorption for 20 minutes; then the adsorbent was separated from the solution by a handheld magnet, and washed with n-hexane, The separated adsorbent was mixed with acetonitrile in a weight ratio of 1:80, and ultrasonically treated for 8 minutes to desorb vitamin D3; the adsorbent was then separated from the solution by a magnet and washed with acetonitrile; the acetonitrile solution containing the desorbed vitamin D3 was filtered through a polytetrafluoroethylene membrane with a pore size of 0.45 μm, the filtrate was collected, and it was concentrated under reduced pressure to obtain vitamin D3 oil; the vitamin D3 oil was mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a solution; the solution was frozen and crystallized at -10°C for 12 hours, then filtered and vacuum dried to obtain pure vitamin D3.

[0067] The preparation method of the magnetic adsorbent comprises the following steps:

[0068] 2.6 parts by weight of FeSO 4 7H 2 O is added to 160 parts by weight of a graphene oxide aqueous suspension with a concentration of 2.2 mg / mL, and then 0.075 parts by weight of lycopodium spore pollen microcapsules and 13 parts by weight of anhydrous ethanol are added, mixed and stirred to obtain a mixture; the mixture is ultrasonically treated for 45 minutes, and then the pH is adjusted to 11 using a 25wt% ammonia solution, and then heated at 92°C for 9 hours to obtain a magnetic hydrogel; the magnetic hydrogel is separated from the solution using a handheld magnet, and washed with distilled water, and then the hydrogel is placed at 20°C until the gel solidifies, and then freeze-dried for 47 hours to obtain a magnetic adsorbent.

[0069] The preparation method of lycopodium spore pollen microcapsules comprises the following steps: mixing lycopodium spore powder and acetone at a weight ratio of 1:8, filtering after refluxing and stirring at 55°C for 2 hours, washing the filtrate with ethanol and water until the filtrate is clear, and then drying to obtain dried lycopodium spores; mixing the dried lycopodium spores with a 0.2 g / mL phosphoric acid aqueous solution at a weight ratio of 1:15, filtering after refluxing and stirring at 75°C for 8.5 hours, washing with water and then drying to obtain lycopodium spore pollen microcapsules.

[0070] Example 5

[0071] A preparation process of pure vitamin D3, which is mainly different from that of Example 4 in that the preparation method of the magnetic adsorbent in step (3) comprises the following steps:

[0072] 2.6 parts by weight of FeSO 4 7H 2 O is added to 160 parts by weight of a graphene oxide aqueous suspension with a concentration of 2.2 mg / mL to obtain a mixture; the mixture is ultrasonically treated for 15 minutes, and then the pH is adjusted to 11 using a 25wt% ammonia solution, and then heated at 92°C for 6 hours to obtain a magnetic hydrogel; the magnetic hydrogel is separated from the solution using a handheld magnet and washed with distilled water, and then the hydrogel is placed at 20°C until the gel solidifies, and then freeze-dried for 24 hours to obtain a magnetic adsorbent.

[0073] Example 6

[0074] A method for detecting vitamin D3 content, the steps of which are as follows:

[0075] (1) Preparation of standard solution

[0076] In a dark environment, mix and dissolve the vitamin D3 standard (purity ≥ 99%, brand: Yingxin Laboratory, item number: TX20875) with anhydrous ethanol to prepare a standard stock solution with a theoretical concentration of 1 mg / mL of vitamin D3; take 10 mL of the above standard stock solution and dilute it with methanol to prepare a standard solution with a concentration of 3 μg / mL;

[0077] (2) Sample processing

[0078] ① Saponification: Weigh 12.00 g of the pure vitamin D3 prepared in Example 1 of the present invention as a sample to be tested, add 1.8 g of ascorbic acid, 1.2 g of pyrogallic acid, 0.12 g of 2,6-di-tert-butyl-p-cresol, 120 mL of anhydrous ethanol and 120 mL of a 40 wt % potassium hydroxide aqueous solution in sequence, then heat and stir at 68° C. for 30 min for saponification, and cool to 28° C. to obtain a saponified solution;

[0079] ② Extraction: transfer the saponified solution to a 1000mL separatory funnel, add 120mL petroleum ether for extraction, extract 3 times, combine the extracts obtained, remove trace water through anhydrous sodium sulfate, and collect in a round-bottom flask; place the round-bottom flask in a rotary evaporator at 45°C until nearly dry, take it out and blow dry it with a nitrogen blower to obtain a sample;

[0080] ③Concentrate the volume: Wash the sample in the round-bottom flask obtained in step ② with n-hexane in small amounts several times and transfer it to a 20 mL constant volume flask to obtain a constant volume sample solution for standby use;

[0081] ④ Purification: sequentially elute the activated silica gel column with 5 mL of dichloromethane and 5 mL of n-hexane, transfer 1.0 mL of the sample solution after constant volume to the silica gel column, then sequentially elute with 5 mL of n-hexane and 5 mL of a mixed solution containing 49.6% (v, v) cyclohexane, 49.6% (v, v) n-hexane, and 0.8% (v, v) isopropanol to remove impurities, then add 5 mL of a mixed solution of ethyl acetate: n-hexane (40:60) to elute and collect vitamin D3 from the silica gel column, then blow dry with a nitrogen blower, add 1.0 mL of methanol to dissolve, filter through a 0.22 μm filter membrane, and wait for HPLC determination;

[0082] (3) High performance liquid chromatography analysis

[0083] The filtrate obtained in step ④ was injected into a high performance liquid chromatograph for analysis.

[0084] The chromatographic conditions during the high performance liquid chromatography analysis in step (3) are as follows:

[0085] Chromatographic column: HPLC SB C18 column (0.45 μm, 5 mm × 150 mm);

[0086] Mobile phase: 95% methanol and 5% water;

[0087] Flow rate: 0.3 mL / min;

[0088] Column temperature: 40°C;

[0089] UV detector (DAD) detection wavelength: 264nm;

[0090] The injection volume was 5 μL.

[0091] Test results: See the attached liquid chromatograms of the vitamin D3 standard and sample. Figure 2 .

[0092] The vitamin D3 content detection method provided by the present invention is an efficient and effective quantitative method. By adopting a solid phase extraction-high performance liquid chromatography HPLC method to detect the vitamin D3 content, the problems existing in the traditional high performance liquid chromatography HPLC method, such as long pretreatment time, easy destruction of effective ingredients, high technical requirements for operators, and large number of laboratory instruments, are improved. After saponification, extraction, and constant volume, vitamin D3 is directly purified by a solid phase extraction column and sampled into a reverse phase HPLC-DAD liquid phase system, which can achieve rapid peak output in a short time, greatly shortening the experimental time and improving the efficiency of the experiment.

[0093] Comparative Example 1

[0094] A preparation process of pure vitamin D3, which is different from Example 4 mainly in that activated carbon is added in step (3) instead of magnetic adsorbent for adsorption operation, that is, step (3) is:

[0095] The obtained high-content pre-vitamin component was heated at a temperature of 55°C and -0.095Mpa for 6 hours to carry out thermal isomerization reaction, and the pre-vitamin was converted into vitamin D3, and then concentrated by vacuum distillation to obtain vitamin D3 conversion oil; the vitamin D3 conversion oil was mixed and dissolved with n-hexane in a weight ratio of 1:30 to obtain a vitamin D3 solution, and then the vitamin D3 solution was mixed with activated carbon in a weight ratio of 1:0.13, stirred and adsorbed for 20 minutes; then filtered through a polytetrafluoroethylene membrane with a pore size of 0.45μm, the filtrate was collected, and concentrated under reduced pressure to obtain vitamin D3 oil; the vitamin D3 oil was mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a dissolving solution; the dissolving solution was placed at -10°C for freezing and crystallization for 12 hours, then filtered, and vacuum dried to obtain pure vitamin D3.

[0096] Comparative Example 2

[0097] A preparation process of pure vitamin D3, which is mainly different from Example 4 in that the vitamin mixture obtained in the actual operation is subjected to a heating isomerization reaction to obtain the vitamin D3 conversion oil and then subjected to column chromatography separation, that is, the preparation process includes the following steps:

[0098] (1) The operation of step (1) is the same as that of Example 4 to prepare a vitamin mixture;

[0099] (2) The obtained vitamin mixture is heated at a temperature of 55° C. and -0.095 MPa for 6 hours to perform a thermal isomerization reaction, and the pre-vitamin is converted into vitamin D3, and then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the obtained vitamin D3 conversion oil is mixed and dissolved with n-hexane in a weight ratio of 1:8 to obtain a mixed solution; the mixed solution is added to a silica gel chromatography column containing 100 kg, and eluted and separated with n-hexane, and the fractions are collected and tracked by high performance liquid chromatography until the vitamin mixture is completely washed out, and the eluate is collected; the solvent is removed from the eluate by vacuum reduction to obtain a high-content vitamin D3 oil;

[0100] (3) Mix and dissolve high-content vitamin D3 oil and n-hexane in a weight ratio of 1:30 to obtain a vitamin D3 solution, then mix the vitamin D3 solution with a magnetic adsorbent in a weight ratio of 1:0.13 and stir to adsorb for 20 minutes; then use a handheld magnet to separate the adsorbent from the solution and wash it with n-hexane, mix the separated adsorbent with acetonitrile in a weight ratio of 1:80, and ultrasonically treat for 8 minutes to desorb the vitamin D3; then separate the adsorbent from the solution with a magnet and wash it with acetonitrile; filter the acetonitrile solution containing the desorbed vitamin D3 through a polytetrafluoroethylene membrane with a pore size of 0.45 μm, collect the filtrate, and concentrate under reduced pressure to obtain vitamin D3 oil; mix and dissolve the vitamin D3 oil and methyl acetate in a weight ratio of 1:8 to obtain a solution; place the solution at -10°C for freezing and crystallization for 12 hours, then filter and vacuum dry to obtain pure vitamin D3.

[0101] The preparation method of the magnetic adsorbent is consistent with that of Example 4.

[0102] Comparative Example 3

[0103] A preparation process of pure vitamin D3, which is different from that of Example 1 mainly in that no ultraviolet light filtering circulating fluid is added in step (1), that is, the preparation process comprises the following steps:

[0104] (1) adding 10 kg of 7-dehydrocholesterol (95 wt%) to a solvent formed by mixing 250 L of methanol and 110 L of normal hexane, heating to 40° C. to mix and dissolve, and then adding 0.8 kg of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; turning on a photochemical water cooler, nitrogen, a circulating water pump, and an ultraviolet lamp, turning on a magnetic pump, and passing the 7-dehydrocholesterol solution into a photochemical reactor, and irradiating the solution with an ultraviolet light source having a wavelength of 290 nm under a flow rate of 8.5 L / min, a reaction temperature of 35° C., and nitrogen conditions to perform a first photochemical reaction, thereby obtaining a first photochemical reaction solution; and then passing the first photochemical reaction solution into a photochemical reactor. In a device, a second photochemical reaction is carried out under irradiation with an ultraviolet light source with a wavelength of 330 nm under the conditions of an inlet flow rate of 8.5 L / min, a reaction temperature of 35° C. and nitrogen, to obtain a second photochemical reaction liquid, and the detection conversion rate of the second photochemical reaction liquid is 58%; then the second photochemical reaction liquid is collected, and 80% of the solvent (n-hexane and methanol) is recovered by reduced pressure distillation at 30° C., and then the 7-dehydrocholesterol raw material is recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10° C. for 10 hours, and then filtered under reduced pressure to obtain a solution containing a large amount of pre-vitamins, and the filtrate is completely recovered to obtain a vitamin mixture;

[0105] (2) mixing and dissolving the obtained vitamin mixture with n-hexane in a weight ratio of 1:8 to obtain a mixed solution; adding the mixed solution to a silica gel chromatography column containing 100 kg, eluting and separating with n-hexane, collecting in sections, tracking with high performance liquid chromatography until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component;

[0106] (3) The obtained high-content pre-vitamin component is heated at a temperature of 55°C and -0.095Mpa for 6 hours to perform a thermal isomerization reaction to convert the pre-vitamin into vitamin D3, which is then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a dissolving solution; the dissolving solution is frozen and crystallized at -10°C for 12 hours, and then filtered and vacuum dried to obtain a pure vitamin D3 product.

[0107] Comparative Example 4

[0108] A preparation process for pure vitamin D3, which is different from Example 4 mainly in that the column chromatography operation in step (2) is omitted, that is, the preparation process comprises the following steps:

[0109] (1) The operation of step (1) is the same as that of Example 4 to prepare a vitamin mixture;

[0110] (2) The obtained vitamin mixture was heated at a temperature of 55°C and -0.095 MPa for 6 hours to perform a thermal isomerization reaction, and the pre-vitamin was converted into vitamin D3, and then concentrated by vacuum distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil was mixed and dissolved with n-hexane in a weight ratio of 1:30 to obtain a vitamin D3 solution, and then the vitamin D3 solution was mixed with a magnetic adsorbent in a weight ratio of 1:0.13 and stirred for adsorption for 20 minutes; then the adsorbent was separated from the solution by a handheld magnet, and washed with n-hexane, and The separated adsorbent is mixed with acetonitrile in a weight ratio of 1:80, and ultrasonically treated for 8 minutes to desorb vitamin D3; the adsorbent is then separated from the solution by a magnet and washed with acetonitrile; the acetonitrile solution containing the desorbed vitamin D3 is filtered through a polytetrafluoroethylene membrane with a pore size of 0.45 μm, the filtrate is collected, and concentrated under reduced pressure to obtain vitamin D3 oil; the vitamin D3 oil is mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a solution; the solution is frozen and crystallized at -10°C for 12 hours, then filtered, and vacuum dried to obtain pure vitamin D3.

[0111] The preparation method of the magnetic adsorbent is consistent with that of Example 4.

[0112] Comparative Example 5

[0113] A preparation process of pure vitamin D3, which is different from Example 1 mainly in that the column chromatography operation in step (2) is omitted, that is, the preparation process comprises the following steps:

[0114] (1) The operation of step (1) is the same as that of Example 1 to prepare a vitamin mixture;

[0115] (2) The obtained vitamin mixture is heated at 55°C and -0.095 MPa for 6 hours to perform thermal isomerization reaction, and the pre-vitamin is converted into vitamin D3, which is then concentrated by vacuum distillation to obtain vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with methyl acetate in a weight ratio of 1:8 to obtain a solution; the solution is frozen and crystallized at -10°C for 12 hours, and then filtered and vacuum dried to obtain pure vitamin D3.

[0116] Test Example 1

[0117] The vitamin mixture content prepared in step (1) of Examples 1 to 3 and Comparative Example 3 was tested and analyzed respectively. Since the steps (1) of Examples 4 to 5, Comparative Examples 1 to 2 and Comparative Examples 4 to 5 were the same as those of Example 1, the test was not repeated. The specific test results are shown in Table 1 below:

[0118] Table 1 Test results of the content of vitamin mixture in Examples 1 to 3 and Comparative Example 3

[0119]

[0120] As can be seen from the test results in Table 1, it is found that in Examples 1 to 3 and Comparative Example 3, the contents of previtamin and vitamin D3 in Examples 1 to 3 using ultraviolet light filtering circulating fluid are higher than those in Comparative Example 3 without using ultraviolet light filtering circulating fluid, and the content of tachysterol is lower than that in Comparative Example 3, and the content of raw material 7-dehydrocholesterol is less. The reason for the analysis may be that the ultraviolet light filtering circulating fluid may play a role in selectively filtering ultraviolet light of a specific wavelength, by absorbing or filtering out wavelengths that are not conducive to the generation of previtamin D3, while retaining wavelengths that are conducive to the conversion of 7-dehydrocholesterol into previtamin D3, thereby improving the generation efficiency of previtamin D3, and reducing the occurrence of side reactions, thereby reducing the content of tachysterol and other unwanted products. In addition, the ultraviolet light filtering circulating fluid may also help to improve the conversion rate of the light reaction, so that more 7-dehydrocholesterol participates in the reaction, reducing the amount of unreacted 7-dehydrocholesterol, which is reflected in the fact that the content of 7-dehydrocholesterol retained in Examples 1 to 3 is less.

[0121] By comparing Examples 1 to 3, it was found that the different concentrations of the ultraviolet light-filtered circulating fluids in Examples 1 to 3 resulted in different tachysterol contents in the test results, with Example 1 having the lowest content. The reason for this may be that the use of a specific concentration of ultraviolet light-filtered circulating fluid may increase the selectivity of the photoreaction, allowing more 7-dehydrocholesterol to be converted into previtamin D3 rather than into tachysterol or other byproducts.

[0122] Test Example 2

[0123] Finished product testing and analysis

[0124] The weights of the pure vitamin D3 products of Examples 1 to 5 and Comparative Examples 1 to 5 dried to constant weight were weighed and recorded respectively; the product morphology of the pure vitamin D3 products prepared in Examples 1 to 5 and Comparative Examples 1 to 5 was observed and the crystal state was recorded, and the purity, optical rotation, and absorption coefficient of the pure vitamin D3 products prepared in Examples 1 to 5 and Comparative Examples 1 to 5 were detected, analyzed, and recorded according to the standards of the "Chinese Pharmacopoeia 2020 Edition". The test results are shown in Table 2 below;

[0125] Crystal purification yield test

[0126] In the process of chemical synthesis or purification, the refining yield is an important parameter to measure the efficiency of the purification process, reflecting the utilization rate of raw materials in the purification process. The higher the refining yield, the less raw materials are lost in the purification process, and the higher the purification efficiency. In addition, medicinal vitamin D3 crystals have strict requirements on purity and crystal morphology. A high refining yield means that purer and better-shaped crystals can be obtained, meeting the standards of pharmaceutical grade.

[0127] Vitamin D3 conversion oil was prepared according to the preparation process of the pure vitamin D3 provided in Examples 1 to 5 and Comparative Examples 1 to 5 of the present invention, and 1 kg of the vitamin D3 conversion oil prepared in Examples 1 to 5 and Comparative Examples 1 to 5 was taken respectively, and the pure vitamin D3 was prepared according to the subsequent operations of the preparation process of the pure vitamin D3 in Examples 1 to 5 and Comparative Examples 1 to 5 respectively, and the final pure vitamin D3 was recorded, and the crystal refining yield (%) of Examples 1 to 5 and Comparative Examples 1 to 5 was calculated according to the final mass of the pure vitamin D3. The crystal refining yield (%) is as follows:

[0128] Crystal purification yield (%) = (mass of needle-shaped vitamin D3 crystals obtained / mass of vitamin D3 conversion oil input) × 100%

[0129] The specific test results are shown in Table 2 below.

[0130] Table 2

[0131]

[0132]

[0133]

[0134] According to the data in Table 2, the pure vitamin D3 prepared in Examples 1 to 5 of the present invention has an optical rotation in the range of +105 to +112 degrees, an absorption coefficient in the range of 465 to 495, and a crystal refining yield of more than 85%, a purity of more than 99%, and the obtained crystals are all pure vitamin D3 products that meet the requirements of the Pharmacopoeia.

[0135] It can be seen from the test results in Table 2 that by comparing Examples 1 to 5 and Comparative Examples 1 to 5, it can be found that the pure products of Examples 1 to 5 and Comparative Examples 1 to 2 are needle-shaped crystals, while Comparative Examples 3 to 4 are a mixture of needles and powders, and the product morphology is different; the yield of the pure products of Examples 1 to 5 and Comparative Example 1 is higher than that of Comparative Examples 2 to 5; the purity is more than 99%, which is higher than that of Comparative Examples 2 to 5; the optical rotation and absorption coefficient of Examples 1 to 5 and Comparative Examples 1 to 5 are both within the specified range; the crystal refining yield of Examples 1 to 5 and Comparative Examples 1 to 5 is more than 85%, which is higher than that of Comparative Examples 2 to 5; the above analysis results show that the pure vitamin D3 prepared by the preparation process of Examples 1 to 5 and Comparative Example 1 has good yield, crystal refining yield and purity, the product morphology is needle-shaped crystals, and Example 4 shows the highest yield, crystal refining yield and purity.

[0136] Compared with Example 1, Example 2 and Example 3 show lower yield, crystal refining yield and purity than Example 1. The reason may be that Example 2 and Example 3 use different solvents and solvent ratios, which may affect the solubility of 7-dehydrocholesterol and the efficiency of the photochemical reaction. The polarity and boiling point of the solvents are different, which may affect the reaction rate and the separation efficiency of the product, resulting in lower yield, crystal refining yield and purity than Example 1. In addition, the concentration and flow rate of the ultraviolet light filtering circulating fluid in Example 2 and Example 3 during the photochemical reaction are different from those in Example 1, which may affect the conversion rate and selectivity of the photochemical reaction. The freezing crystallization time of Example 2 and Example 3 is different from that of Example 1, and the length of the freezing crystallization time may affect the growth and formation of crystals, thereby affecting the purity and yield of the product.

[0137] Compared with Example 1, the product morphology of Comparative Example 3 is a mixture of needles and powders, and the yield, crystal refining yield and purity are significantly lower than those of Example 1. Analysis shows that the reason may be that the preparation process of Comparative Example 3 does not introduce ultraviolet light filtering circulating liquid for ultraviolet light treatment, which may reduce the efficiency and selectivity of the photochemical reaction, resulting in the yield, crystal refining yield and purity being significantly lower than those of Example 1, and affecting the growth and formation of its product crystals.

[0138] Compared with Example 1, the product morphology of Comparative Example 5 is a mixture of needles and powders, the yield is reduced to 1.43 kg, the crystal refining yield is reduced to 72.3%, and the purity is reduced to 95.21%. The reason may be that the column chromatography separation operation is missing in the preparation process of Comparative Example 5, which may increase the impurity content in the product, thereby reducing the yield, crystal refining yield and purity, and affecting the growth and formation of its product crystals. Column chromatography separation operation helps to improve the purity of vitamin D3, but may reduce the yield.

[0139] Compared with Example 4, the step (3) of the preparation process of Example 1 does not introduce a magnetic adsorbent for adsorption operation, and its yield, crystal refining yield and purity are significantly lower than those of Example 4, indicating that the magnetic adsorbent can effectively remove impurities and improve the purity of the product, thereby improving the yield and quality. The reason for this may be that the magnetic adsorbent has a high specific surface area and a porous structure, which enables it to provide a large number of adsorption sites and has high selectivity for vitamin D3, while its adsorption capacity for impurities is weak. This high adsorption capacity and selective adsorption help to improve the recovery rate of vitamin D3, thereby improving the refining yield and purity of the crystals. In addition, the magnetic adsorbent can be quickly separated using an external magnetic field. This feature allows the adsorbent to be quickly removed from the solution after the adsorption process is completed, reducing the loss of vitamin D3 in traditional separation steps such as filtration and centrifugation, thereby improving the yield.

[0140] In the preparation process of the magnetic adsorbent of Example 5, lycopodiella spore pollen microcapsules were not introduced, and its yield, crystal purification yield and purity were lower than those of Example 4, which may indicate that the introduction of lycopodiella spore pollen microcapsules can enhance the selectivity and adsorption efficiency of the magnetic adsorbent.

[0141] Comparative Example 1 uses activated carbon instead of magnetic adsorbent for adsorption operation, showing a yield, crystal refining yield and purity significantly lower than those of Example 4, which indicates that magnetic adsorbents may have higher selectivity and adsorption efficiency than traditional activated carbon.

[0142] In the preparation process of Comparative Example 2, the vitamin mixture is subjected to a heating isomerization reaction to obtain the vitamin D3 conversion oil, which is then separated by column chromatography. Compared with Example 4, the yield, crystal refining yield and purity are greatly reduced. This may be because the vitamin D3 resin oil obtained by the thermal isomerization reaction is then subsequently separated, which may result in impurities not being separated in time, resulting in a decrease in the vitamin D3 yield, crystal refining yield and purity.

[0143] The preparation process of Comparative Example 4 lacks column chromatography separation operation, and its product morphology is a mixture of needles and powders, the yield is reduced to 1.53 kg, the crystal refining yield is reduced to 78.4%, and the purity is reduced to 96.25%. This shows that column chromatography separation operation can help to obtain a high content of pre-vitamin, and then obtain high-purity pure vitamin D3 crystals that meet the requirements of the pharmacopoeia through thermal isomerization and crystallization, which is beneficial to improve the reaction and separation efficiency, thereby increasing the reaction yield and the purity of vitamin D3, and is an indispensable operating step in the vitamin D3 preparation process.

Claims

1. A process for preparing pure vitamin D3, characterized in that: The method comprises the following steps, in parts by weight: (1) adding 9 to 12 parts by weight of 7-dehydrocholesterol to 300 to 400 parts by weight of a solvent, heating the mixture to 38 to 43° C. to dissolve the mixture, and then adding 0.5 to 1.5 parts by weight of butylated hydroxyanisole to obtain a 7-dehydrocholesterol solution; opening a photochemical reactor, introducing a first ultraviolet light filtering circulating liquid, and then introducing the 7-dehydrocholesterol solution into the photochemical reactor, irradiating the solution with an ultraviolet light source at a flow rate of 8 to 10 L / min, a reaction temperature of 25 to 40° C. and nitrogen to perform a first photochemical reaction, and then introducing a second ultraviolet light filtering circulating liquid, and then introducing the first ultraviolet light filtering circulating liquid into the photochemical reactor. The chemical reaction liquid is introduced into a photochemical reactor, and a second photochemical reaction is carried out under the conditions of an introduction flow rate of 8 to 10 L / min, a reaction temperature of 25 to 40° C. and nitrogen by irradiation with an ultraviolet light source to obtain a second photochemical reaction liquid, and the conversion rate is controlled at 50% to 65%; the second photochemical reaction liquid is collected, and 80 to 90% of the solvent is recovered by reduced pressure distillation at 20 to 30° C., and then 7-dehydrocholesterol raw materials are recovered by filter pressing, and the filtrate is collected, and the filtrate is stirred and frozen and crystallized at -10° C. to -5° C. for 10 to 12 hours, and then filtered under reduced pressure, and the solvent is recovered from the filtrate to obtain a mixture of pre-vitamin and vitamin D3, that is, a vitamin mixture; (2) mixing and dissolving the vitamin mixture obtained in the above step (1) with an adsorption solvent in a weight ratio of 1:5-10 to obtain a mixed solution; adding the mixed solution to a chromatography column, eluting and separating with an eluting solvent, collecting in sections, tracking with a high performance liquid chromatography analysis method until the vitamin mixture is completely eluted, and collecting the eluate; removing the solvent from the eluate under reduced pressure to obtain a high-content pre-vitamin component; (3) The high-content pre-vitamin component obtained in the above step (2) is heated at a temperature of 40°C to 65°C and a vacuum degree of -0.070 to 0.095 MPa for 3 to 8 hours to perform a thermal isomerization reaction until the pre-vitamin is converted into vitamin D3, and then concentrated by reduced pressure distillation to obtain a vitamin D3 conversion oil; the vitamin D3 conversion oil is mixed and dissolved with a crystallization solvent in a weight ratio of 1:5-10 to obtain a dissolving solution; the dissolving solution is frozen and crystallized at -8 to 5°C for 8 to 20 hours, and then filtered and dried to obtain a pure vitamin D3 product.

2. The process for preparing pure vitamin D3 according to claim 1, characterized in that: The solvent is selected from at least one of C3-C3 alcohols, mixed solvents of C3-C3 alcohols and C5-C7 alkanes; preferably, the solvent is at least one of methanol, ethanol, a mixed solvent of methanol and n-hexane, a mixed solvent of ethanol and n-hexane, and a mixed solvent of ethanol and n-heptane.

3. The process for preparing pure vitamin D3 according to claim 2, characterized in that: Among the solvents, the mixed solvent of methanol and n-hexane is a mixture of methanol and n-hexane in a volume ratio of 2.5:(1-1.2); the mixed solvent of ethanol and n-hexane is a mixture of ethanol and n-hexane in a volume ratio of 2.5:(1-1.2); and the mixed solvent of ethanol and n-heptane is a mixture of ethanol and n-heptane in a volume ratio of 2.5:(1-1.2).

4. The process for preparing pure vitamin D3 according to claim 1, characterized in that: The first ultraviolet light filtering circulating liquid in step (1) is a self-prepared copper sulfate aqueous solution with a concentration of 1.5-7wt%; the second ultraviolet light filtering circulating liquid is a self-prepared copper sulfate aqueous solution with a concentration of 4.5-10wt%.

5. The process for preparing pure vitamin D3 according to claim 1, characterized in that: In the first photochemical reaction in step (1), the 7-dehydrocholesterol solution undergoes a photochemical reaction under the irradiation of an ultraviolet light source with a wavelength range of 280 to 300 nm; in the second photochemical reaction, the first photochemical reaction solution undergoes a photochemical reaction under the irradiation of an ultraviolet light source with a wavelength range of 310 to 350 nm.

6. The process for preparing pure vitamin D3 according to claim 1, characterized in that: The adsorption solvent in step (2) is selected from at least one of C6-C12 alkanes; the stationary phase of the chromatography column is selected from one of silica gel and activated alumina; and the elution solvent is selected from at least one of C6-C9 alkanes.

7. The process for preparing pure vitamin D3 according to claim 1, characterized in that: The crystallization solvent in step (3) is selected from at least one of C2 to C9 ester solvents.

8. The process for preparing pure vitamin D3 according to claim 1, characterized in that: The step (3) may also be: The high-content pre-vitamin component obtained in the above step (2) is heated at a temperature of 40° C. to 65° C. and a vacuum degree of -0.070 to 0.095 MPa for 3 to 8 hours for thermal isomerization reaction until the pre-vitamin is converted into vitamin D3, and then concentrated by reduced pressure distillation to obtain vitamin D3 conversion oil; The vitamin D3 conversion oil and n-hexane are mixed and dissolved in a weight ratio of 1:20 to 40 to obtain a vitamin D3 solution, and then the vitamin D3 solution and the adsorbent are mixed and stirred in a weight ratio of 1:0.1 to 0.3 for adsorption for 15 to 25 minutes; then the adsorbent is separated from the solution by a handheld magnet and washed with n-hexane, and the separated adsorbent is mixed with acetonitrile in a weight ratio of 1:50 to 100, and ultrasonically treated for 5 to 10 minutes to desorb the vitamin D3; then the adsorbent is separated from the solution by a magnet and washed with acetonitrile; the acetonitrile solution containing the desorbed vitamin D3 is filtered through a polytetrafluoroethylene membrane with a pore size of 0.45 μm, the filtrate is collected, and concentrated under reduced pressure to obtain vitamin D3 oil; the vitamin D3 oil and the crystallization solvent are mixed and dissolved in a weight ratio of 1:5 to 10 to obtain a dissolving solution; the dissolving solution is frozen and crystallized at -8 to 5°C for 8 to 20 hours, and then filtered and dried to obtain pure vitamin D3.

9. The process for preparing pure vitamin D3 according to claim 8, characterized in that: The adsorbent is a magnetic adsorbent, and its preparation method comprises the following steps, calculated by weight: 2 to 3 parts by weight of FeSO4·7H2O are added to 130 to 180 parts by weight of a graphene oxide aqueous suspension having a concentration of 1.5 to 3 mg / mL, and then 0.06 to 0.08 parts by weight of lycopodium spore pollen microcapsules and 10 to 15 parts by weight of anhydrous ethanol are added, mixed and stirred to obtain a mixture; the mixture is ultrasonically treated for 30 to 50 minutes, and then a 20 to 30 wt% ammonia aqueous solution is used to adjust the pH to 10 to 12, and then heated at 85 to 95° C. for 7 to 10 hours to obtain a magnetic hydrogel; the magnetic hydrogel is separated from the solution using a handheld magnet, and washed with distilled water, and then the hydrogel is placed at 18 to 22° C. until the gel solidifies, and then freeze-dried for 45 to 50 hours to obtain a magnetic adsorbent; Alternatively, 2 to 3 parts by weight of FeSO4·7H2O are added to 130 to 180 parts by weight of a graphene oxide aqueous suspension having a concentration of 1.5 to 3 mg / mL, and the mixture is mixed and stirred to obtain a mixture; the mixture is ultrasonically treated for 10 to 20 minutes, and then a 20 to 30 wt% ammonia aqueous solution is used to adjust the pH to 10 to 12, and then heated at 85 to 95° C. for 4 to 8 hours to obtain a magnetic hydrogel; the magnetic hydrogel is separated from the solution using a handheld magnet, and washed with distilled water, and then the hydrogel is placed at 18 to 22° C. until the gel is solidified, and then freeze-dried for 22 to 26 hours to obtain a magnetic adsorbent. The preparation method of lycopodium spore pollen microcapsules comprises the following steps: mixing lycopodium spore powder and acetone at a weight ratio of 1:5-10, filtering after refluxing and stirring at 45-60°C for 1.5-3h, washing the filtrate with ethanol and water until the filtrate is clear, and then drying to obtain dried lycopodium spores; mixing the dried lycopodium spores with a 0.1-0.3g / mL phosphoric acid aqueous solution at a weight ratio of 1:10-20, refluxing and stirring at 70-80°C for 7-10h, filtering, washing with water and then drying to obtain lycopodium spore pollen microcapsules.

10. A pure vitamin D3 product, characterized in that: Prepared by the process described in any one of claims 1 to 9.

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