CN103554206A - Method for separating hyocholic acid from pig's bile - Google Patents
Method for separating hyocholic acid from pig's bile Download PDFInfo
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- CN103554206A CN103554206A CN201310449525.9A CN201310449525A CN103554206A CN 103554206 A CN103554206 A CN 103554206A CN 201310449525 A CN201310449525 A CN 201310449525A CN 103554206 A CN103554206 A CN 103554206A
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Abstract
The invention discloses a method for separating hyocholic acid from pig's bile. The method includes the steps of esterification, extraction, recrystallization, hydrolysis, secondary recrystallization and the like. The method takes pig's bile as the extraction raw material, and after extraction of bilirubin from pig's bile, a hyodeoxycholic acid extracted filtrate is collected for separation, thus obtaining hyocholic acid. The pig's bile raw material is wide and easily available, thus reducing the preparation cost of hyocholic acid. The extraction process involved in the invention selects acetic anhydride as an esterification reagent, the hyocholic acid ester hydrolysis and recrystallization refining conditions are determined, and sodium hydroxide is adopted as a base catalyst. With the technological process, the total yield of hyocholic acid is up to over 5%.
Description
Technical field
The invention belongs to biological medicine technology field, be specifically related to a kind of from Fel Sus domestica the method for separated Iocholic acid.
Background technology
Bile is the product that human body or vertebrates liver cell secrete continuously, it is a kind of coloured alkaline liquid of viscosity with bitter taste, beyond its composition dewaters, mainly contain bile acide, cholate and bile pigment, also have in addition a small amount of cholesterol, lipid acid, Yelkin TTS, ionogen and protein etc.More common animal bile is generally bird gall, Fel Sus domestica, oxgall, Fel Ursi and Fel Serpentis etc. now.Bile acide is the main component of bile, account for 50%~70% of bile total amount, the physiological action of bile is mainly brought into play by bile acide exactly, when as medicine, that bile acide can be used for is analgesic, antispastic antibechic, regulate blood pressure, cardiac stimulant cholagogic, antibacterial anti-inflammatory and regulate immunity etc., and activity is strong, side effect is again little, cheap, is therefore subject to vast medication person's welcome.
In various animal biles, Fel Sus domestica is the most common and output is maximum, from ancient times, start to be just widely used in clinical, as by Fel Sus domestica treatment acute icteric infectious hepatitis, Whooping cough, acute enteritis and stomach ulcer etc., all can receive good therapeutic action.Main containing bile acids, bile pigment, Saliva Orthana, lipid and inorganics etc. in Fel Sus domestica, bile acide mainly comprises Hyodeoxycholic Acid, Iocholic acid, Chenodiol, 3 Alpha-hydroxies-6-oxygen-5 α-ursodeoxycholic acid and cholelith acid etc.Wherein Hyodeoxycholic Acid accounts for 40%, and Chenodiol accounts for 25%.
Iocholic acid chemistry 3 α by name, 6 α, 7 α-trihydroxy--5 β-ursodeoxycholic acid, molecular formula is C24H40O5, molecular weight 408.577, CAS 547-75-1, is white, needle-shaped crystals, is soluble in ethanol, and glacial acetic acid is water-soluble hardly, and fusing point is 188~189 ℃.
Iocholic acid has the effect that reduces blood cholesterol levels, can be used for treating hypertension and blood vessel scleratheroma; Gram-positive microorganism is had to good restraining effect, particularly better to the staphylococcic bacteriostatic action of white, in recent years report in addition that it has certain therapeutic action to human promyelocytic leukemia.It is also the main component of artificial Calculus Bovis and QINGKAILING, and can be used as the synthetic rare bile acide ursodesoxycholic acid of raw material.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind ofly take Fel Sus domestica and is the method for the separated Iocholic acid of raw material, and the method extraction process is simple, and without specific installation, raw material sources are extensively easy to get, and environmental pollution is little, and product purity is high.
For solving the problems of the technologies described above, the technical scheme that Bian of the present invention gets is as follows:
From Fel Sus domestica, a method for separated Iocholic acid, is characterized in that, comprises the steps:
(1) get and carried bilirubinic Fel Sus domestica tankage, through saponification, hydrolysis, decolouring and after being dried, obtain tan pig TOTAL BILE ACID crude product;
(2) get pig TOTAL BILE ACID crude product, after esterification, reagent obtains pig TOTAL BILE ACID ester powder with ethanol-extracted, recrystallization, then pig TOTAL BILE ACID ester is hydrolyzed under alkaline condition after, recrystallization obtains Hyodeoxycholic Acid white crystal for the second time;
(3) filtrate step (1) and (2) being obtained, washing lotion and recrystallization mother liquor merge, and decompression and solvent recovery obtains brown thick thing, adds esterifying reagent, carry out esterification and generate Iocholic acid ester under catalyst action;
(4) under alkaline condition, with alkane, extract Iocholic acid ester;
(5) Iocholic acid ester is carried out to recrystallization, the white powder that obtains Iocholic acid ester is dried to constant weight under infrared lamp;
(6) get the Iocholic acid ester that above-mentioned (5) make, after reaction that Iocholic acid ester is hydrolyzed under alkaline condition, carry out recrystallization, the white crystal obtaining is dried to constant weight under infrared lamp, can make Iocholic acid.
As improvement of the present invention, the esterifying reagent of above-mentioned steps (3) is diacetyl oxide or formic anhydride.
Further, the esterifying reagent of above-mentioned steps (3) is diacetyl oxide.
As improvement of the present invention, the catalyzer of above-mentioned steps (3) is sodium acetate or sodium formiate.
As improvement of the present invention, the alkali of above-mentioned steps (4) is NaOH.
As improvement of the present invention, the alkane of above-mentioned steps (4) is normal hexane.
As improvement of the present invention, the recrystallization reagent that above-mentioned steps (5) is used is benzene-sherwood oil system.
As improvement of the present invention, the recrystallization reagent that above-mentioned steps (6) is used is ethyl acetate.
Compared with prior art, beneficial effect of the present invention is:
The present invention is usingd Fel Sus domestica as extracting raw material, and Fel Sus domestica extracts after bilirubin, collects the filtrate of extracting after Hyodeoxycholic Acid and carries out separation and make Iocholic acid, and Fel Sus domestica raw material is extensively easy to get, and has reduced the preparation cost of Iocholic acid.In extraction process of the present invention, choosing diacetyl oxide is esterifying reagent, has determined the condition of the hydrolysis of Iocholic acid ester and recrystallizing and refining, determines and adopts sodium hydroxide as alkaline catalysts, and through this technical process, the total recovery of Iocholic acid reaches more than 5%.
Accompanying drawing explanation
Fig. 1 is process flow sheet of the present invention.
Embodiment
For ease of understanding the present invention, it is as follows that the present invention enumerates embodiment.Those skilled in the art should understand, described embodiment only, for helping to understand the present invention, should not be considered as concrete restriction of the present invention.
In the following example, the same meaning that all specialties of using and scientific words and those skilled in the art are familiar.
As shown in Figure 1, a kind of from Fel Sus domestica the method for separated Iocholic acid, comprise the steps:
(1) get and carried bilirubinic Fel Sus domestica tankage, through saponification, hydrolysis, decolouring and after being dried, obtain tan pig TOTAL BILE ACID crude product;
(2) get pig TOTAL BILE ACID crude product, after esterification, reagent obtains pig TOTAL BILE ACID ester powder with ethanol-extracted, recrystallization, then pig TOTAL BILE ACID ester is hydrolyzed under alkaline condition after, recrystallization obtains Hyodeoxycholic Acid white crystal for the second time;
(3) filtrate step (1) and (2) being obtained, washing lotion and recrystallization mother liquor merge, and decompression and solvent recovery obtains brown thick thing, adds esterifying reagent, carry out esterification and generate Iocholic acid ester under catalyst action;
(4) under alkaline condition, with alkane, extract Iocholic acid ester;
(5) Iocholic acid ester is carried out to recrystallization, the white powder that obtains Iocholic acid ester is dried to constant weight under infrared lamp;
(6) get the Iocholic acid ester that above-mentioned (5) make, after reaction that Iocholic acid ester is hydrolyzed under alkaline condition, carry out recrystallization, the white crystal obtaining is dried to constant weight under infrared lamp, can make Iocholic acid.
Embodiment 1
(1) get 250ml round-bottomed flask, add pig TOTAL BILE ACID crude product 10g, anhydrous methanol 50ml, 98% vitriol oil 0.7ml, stirring reaction 24h under normal temperature, complete through TLC detection reaction, without the residual rear stopped reaction of raw material, in system, add anhydrous Na HCO
3solid adjusts pH to neutral, elimination inorganic salt, filter cake methanol wash, merging filtrate and washing lotion, reclaim under reduced pressure methyl alcohol, obtain brown dope, add water 50ml to dissolve wherein remaining inorganic salt, then extract organism (50ml with ether gradation, 30ml, 20ml), merge organic phase, respectively with saturated NaHCO
3solution and saturated common salt water washing 1 time, anhydrous Na
2sO
4dry 2h, filters, and removes ether under reduced pressure, obtain brown thick thing 11.4g, in flask, add benzene 30ml, at 80 ℃ of oil bath pans, after dissolve complete, stop heating, after naturally cooling to room temperature, put into the about 16h of refrigerator, separate out solid, filter, filter cake is with benzene washing 3 times, obtain off-white color solid 4.9g, then carry out recrystallization with benzene, the white powder obtaining is dried to constant weight under infrared lamp, for 4.5g, i.e. pig TOTAL BILE ACID methyl esters;
(2) get 100ml round-bottomed flask, add successively Hyodeoxycholic Acid methyl esters (4.5g), NaOH solid (1.1g), ethanol 30ml, water 1ml, stirring and refluxing reaction 20h at 80 ℃, after completion of the reaction, decompression and solvent recovery, to dry, obtains faint yellow solid in TLC monitoring, add water 30ml, under vigorous stirring, add 10% dilute hydrochloric acid and adjust pH to 1~2, separate out a large amount of solids, continue to stir at normal temperatures 0.5h, filter, 10% dilute hydrochloric acid washing 3 times for filter cake, washes 3 times, obtains block white solid, drying under reduced pressure 8h at 50 ℃ is 4.5g.With re-crystallizing in ethyl acetate, after the white crystal obtaining is dried to constant weight under infrared lamp, be 4g, mp is 200~201 ℃, i.e. Hyodeoxycholic Acid.
(3) filtrate upper step being obtained, washing lotion and recrystallization mother liquor merge, and reclaim under reduced pressure benzene, to the greatest extent, obtains brown thick thing 6.7g.Add diacetyl oxide 54ml, sodium acetate 8g, back flow reaction 5h at 110 ℃, after TLC monitoring has been reacted, remove the diacetyl oxide of approximately 2/3 volume under reduced pressure, after system is down to room temperature naturally, add water 50ml to dissolve inorganic salt wherein, with ether gradation, extract organism (50ml, 30ml, 20ml), merge organic phase, first neutral to approaching by 1% NaOH solution washing organic phase, use again saturated NaHCO3 solution and saturated common salt water washing, anhydrous Na
2sO
4dry 2h, filters, and ether is removed in decompression, obtain brown thick thing 7.9g, in flask, add normal hexane 20ml, system is warming up to after 65 ℃ of stirrings are dissolved it completely and stops heating, naturally be down to room temperature, at room temperature stir 2h, filter, normal hexane washing 3 times for filter cake, after dry, obtain brown ceramic powder 1.7g, successively with methyl alcohol and benzene-sherwood oil system recrystallization, after being dried, obtain white powder 0.8g, mp184~186 ℃, i.e. Iocholic acid methyl esters triacetate.
(4) get 25ml round-bottomed flask, add successively Iocholic acid methyl esters triacetate (0.8g), NaOH solid (0.35g), ethanol 10ml, water 0.4ml, back flow reaction 20h at 80 ℃, TLC monitoring after completion of the reaction, decompression and solvent recovery, obtain pale solid, add water 10ml, under vigorous stirring, with 10% dilute hydrochloric acid, adjust PH to 1.5, separate out solid, continue under normal temperature and stir 0.5h, filter, filter cake respectively washs 3 times with 10% dilute hydrochloric acid and water successively, obtain white solid, at 50 ℃, after drying under reduced pressure, be 0.6g, with re-crystallizing in ethyl acetate, under infrared lamp, be dried to constant weight, obtain white powder 0.52g, mp188~189 ℃, be Iocholic acid, total recovery is 5.2%.
Embodiment 2
Compare with embodiment 1, (1) is identical with (2) step, and (3) and (4) step is as follows:
(3) filtrate upper step being obtained, washing lotion and recrystallization mother liquor merge, and reclaim under reduced pressure benzene, to the greatest extent, obtains brown thick thing 6.7g.Add formic anhydride 54ml, sodium formiate 8g, back flow reaction 5h at 110 ℃, after TLC monitoring has been reacted, remove the diacetyl oxide of approximately 2/3 volume under reduced pressure, after system is down to room temperature naturally, add water 50ml to dissolve inorganic salt wherein, with ether gradation, extract organism (50ml, 30ml, 20ml), merge organic phase, first neutral to approaching by 1% NaOH solution washing organic phase, use again saturated NaHCO3 solution and saturated common salt water washing, anhydrous Na
2sO
4dry 2h, filters, and ether is removed in decompression, obtain brown thick thing 7.9g, in flask, add normal hexane 20ml, system is warming up to after 65 ℃ of stirrings are dissolved it completely and stops heating, naturally be down to room temperature, at room temperature stir 2h, filter, normal hexane washing 3 times for filter cake, after dry, obtain brown ceramic powder 1.7g, successively with methyl alcohol and benzene-sherwood oil system recrystallization, after being dried, obtain white powder 0.8g, mp184~186 ℃, i.e. Iocholic acid methyl esters triacetate.
(4) get 25ml round-bottomed flask, add successively Iocholic acid methyl esters triacetate (0.8g), NaOH solid (0.35g), ethanol 10ml, water 0.4ml, back flow reaction 20h at 80 ℃, TLC monitoring after completion of the reaction, decompression and solvent recovery, obtain pale solid, add water 10ml, under vigorous stirring, with 10% dilute hydrochloric acid, adjust PH to 1.5, separate out solid, continue under normal temperature and stir 0.5h, filter, filter cake respectively washs 3 times with 10% dilute hydrochloric acid and water successively, obtain white solid, at 50 ℃, after drying under reduced pressure, be 0.6g, with re-crystallizing in ethyl acetate, under infrared lamp, be dried to constant weight, obtain white powder 0.50g, mp mp188~189 ℃, be Iocholic acid, total recovery is 5.0%.
Applicant's statement, person of ordinary skill in the field is on the basis of above-described embodiment, by the concrete content point value of above-described embodiment component, combined with the technical scheme of summary of the invention part, thereby the new numerical range producing, also be one of record scope of the present invention, the application, for making specification sheets simple and clear, is no longer enumerated these numerical ranges.
Claims (8)
1. a method for separated Iocholic acid from Fel Sus domestica, is characterized in that, comprises the steps:
(1) get and carried bilirubinic Fel Sus domestica tankage, through saponification, hydrolysis, decolouring and after being dried, obtain tan pig TOTAL BILE ACID crude product;
(2) get pig TOTAL BILE ACID crude product, after esterification, reagent obtains pig TOTAL BILE ACID ester powder with ethanol-extracted, recrystallization, then pig TOTAL BILE ACID ester is hydrolyzed under alkaline condition after, recrystallization obtains Hyodeoxycholic Acid white crystal for the second time;
(3) filtrate step (1) and (2) being obtained, washing lotion and recrystallization mother liquor merge, and decompression and solvent recovery obtains brown thick thing, adds esterifying reagent, carry out esterification and generate Iocholic acid ester under catalyst action;
(4) under alkaline condition, with alkane, extract Iocholic acid ester;
(5) Iocholic acid ester is carried out to recrystallization, the white powder that obtains Iocholic acid ester is dried to constant weight under infrared lamp;
(6) get the Iocholic acid ester that above-mentioned (5) make, after reaction that Iocholic acid ester is hydrolyzed under alkaline condition, carry out recrystallization, the white crystal obtaining is dried to constant weight under infrared lamp, can make Iocholic acid.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the esterifying reagent of described step (3) is diacetyl oxide or formic anhydride.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the esterifying reagent of described step (3) is diacetyl oxide.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the catalyzer of described step (3) is sodium acetate or sodium formiate.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the alkane of described step (4) is normal hexane or normal butane.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the alkali of described step (4) is NaOH.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the recrystallization reagent that described step (5) is used is benzene-sherwood oil system.
As claimed in claim 1 a kind of from Fel Sus domestica the method for separated Iocholic acid, it is characterized in that: the recrystallization reagent that described step (6) is used is ethyl acetate.
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Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN104041678A (en) * | 2014-05-28 | 2014-09-17 | 山东龙昌动物保健品有限公司 | Bile acid composition and preparation method thereof |
CN105646635A (en) * | 2016-03-15 | 2016-06-08 | 成都市新功生物科技有限公司 | Method for recycling cholic acid from chicken bile extraction industrial effluent |
CN107033059A (en) * | 2017-06-15 | 2017-08-11 | 马鞍山汇智生物技术有限公司 | A kind of new method that bilirubin and chenodeoxycholic acid are prepared by raw material of pig gall |
CN110003302A (en) * | 2019-04-30 | 2019-07-12 | 山东省药学科学院 | A kind of refining methd of 7- Ketolithocholsaeure |
CN111961105A (en) * | 2020-09-23 | 2020-11-20 | 安徽科宝生物工程有限公司 | Method for separating hyodeoxycholic acid from pig bile paste by extraction method |
-
2013
- 2013-09-27 CN CN201310449525.9A patent/CN103554206A/en active Pending
Non-Patent Citations (2)
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丁新腾等: "《从猪胆汁提取鹅去氧胆酸及猪胆酸》", 《医药工业》 * |
邓启华等: "《猪胆汁中三种主要胆汁酸的提取分离》", 《中国生化药物杂志》 * |
Cited By (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN104041678A (en) * | 2014-05-28 | 2014-09-17 | 山东龙昌动物保健品有限公司 | Bile acid composition and preparation method thereof |
CN104041678B (en) * | 2014-05-28 | 2016-05-11 | 山东龙昌动物保健品有限公司 | A kind of bile acid composition and preparation method thereof |
CN105646635A (en) * | 2016-03-15 | 2016-06-08 | 成都市新功生物科技有限公司 | Method for recycling cholic acid from chicken bile extraction industrial effluent |
CN107033059A (en) * | 2017-06-15 | 2017-08-11 | 马鞍山汇智生物技术有限公司 | A kind of new method that bilirubin and chenodeoxycholic acid are prepared by raw material of pig gall |
CN110003302A (en) * | 2019-04-30 | 2019-07-12 | 山东省药学科学院 | A kind of refining methd of 7- Ketolithocholsaeure |
CN110003302B (en) * | 2019-04-30 | 2021-05-11 | 山东省药学科学院 | Refining method of 7-ketolithocholic acid |
CN111961105A (en) * | 2020-09-23 | 2020-11-20 | 安徽科宝生物工程有限公司 | Method for separating hyodeoxycholic acid from pig bile paste by extraction method |
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