CN111116642A - Method for preparing natural L- α -glycerophosphatidylcholine - Google Patents
Method for preparing natural L- α -glycerophosphatidylcholine Download PDFInfo
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- CN111116642A CN111116642A CN202010096450.0A CN202010096450A CN111116642A CN 111116642 A CN111116642 A CN 111116642A CN 202010096450 A CN202010096450 A CN 202010096450A CN 111116642 A CN111116642 A CN 111116642A
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- glycerophosphatidylcholine
- phosphatidylcholine
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- 238000000034 method Methods 0.000 title claims description 23
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 53
- 239000011347 resin Substances 0.000 claims abstract description 33
- 229920005989 resin Polymers 0.000 claims abstract description 33
- 238000003756 stirring Methods 0.000 claims abstract description 31
- 238000006731 degradation reaction Methods 0.000 claims abstract description 30
- 238000001914 filtration Methods 0.000 claims abstract description 28
- 239000013078 crystal Substances 0.000 claims abstract description 26
- WTJKGGKOPKCXLL-RRHRGVEJSA-N phosphatidylcholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCC=CCCCCCCCC WTJKGGKOPKCXLL-RRHRGVEJSA-N 0.000 claims abstract description 22
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 claims abstract description 14
- 239000000203 mixture Substances 0.000 claims abstract description 14
- 150000001875 compounds Chemical class 0.000 claims abstract description 13
- 238000001816 cooling Methods 0.000 claims abstract description 11
- 239000008213 purified water Substances 0.000 claims abstract description 11
- 239000002798 polar solvent Substances 0.000 claims abstract description 9
- 230000003472 neutralizing effect Effects 0.000 claims abstract description 7
- 239000002253 acid Substances 0.000 claims abstract description 6
- 238000007670 refining Methods 0.000 claims description 29
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 24
- 239000012043 crude product Substances 0.000 claims description 23
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 18
- 239000002994 raw material Substances 0.000 claims description 15
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 claims description 14
- 239000003456 ion exchange resin Substances 0.000 claims description 12
- 229920003303 ion-exchange polymer Polymers 0.000 claims description 12
- WQDUMFSSJAZKTM-UHFFFAOYSA-N Sodium methoxide Chemical compound [Na+].[O-]C WQDUMFSSJAZKTM-UHFFFAOYSA-N 0.000 claims description 10
- 239000003795 chemical substances by application Substances 0.000 claims description 10
- 230000035484 reaction time Effects 0.000 claims description 7
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 claims description 6
- 239000000706 filtrate Substances 0.000 claims description 6
- QDRKDTQENPPHOJ-UHFFFAOYSA-N sodium ethoxide Chemical compound [Na+].CC[O-] QDRKDTQENPPHOJ-UHFFFAOYSA-N 0.000 claims description 4
- 150000007514 bases Chemical class 0.000 claims description 3
- 238000002425 crystallisation Methods 0.000 claims description 3
- 230000008025 crystallization Effects 0.000 claims description 3
- 238000006386 neutralization reaction Methods 0.000 claims description 3
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 claims description 3
- 238000004042 decolorization Methods 0.000 claims description 2
- 239000008349 purified phosphatidyl choline Substances 0.000 claims description 2
- 150000001805 chlorine compounds Chemical group 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 abstract description 3
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 15
- 239000007788 liquid Substances 0.000 description 11
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 description 9
- PEDCQBHIVMGVHV-UHFFFAOYSA-N glycerol Substances OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 9
- 239000000047 product Substances 0.000 description 7
- 239000012535 impurity Substances 0.000 description 5
- 150000003904 phospholipids Chemical class 0.000 description 5
- 239000008367 deionised water Substances 0.000 description 4
- 229910021641 deionized water Inorganic materials 0.000 description 4
- 238000000926 separation method Methods 0.000 description 4
- SUHOQUVVVLNYQR-MRVPVSSYSA-N choline alfoscerate Chemical compound C[N+](C)(C)CCOP([O-])(=O)OC[C@H](O)CO SUHOQUVVVLNYQR-MRVPVSSYSA-N 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- HZAXFHJVJLSVMW-UHFFFAOYSA-N 2-Aminoethan-1-ol Chemical compound NCCO HZAXFHJVJLSVMW-UHFFFAOYSA-N 0.000 description 2
- 235000010469 Glycine max Nutrition 0.000 description 2
- 244000068988 Glycine max Species 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000003729 cation exchange resin Substances 0.000 description 2
- 238000004587 chromatography analysis Methods 0.000 description 2
- 150000004665 fatty acids Chemical group 0.000 description 2
- -1 glycerol phosphatidylethanolamine Chemical class 0.000 description 2
- JZNWSCPGTDBMEW-UHFFFAOYSA-N glycerophosphatidylethanolamine Chemical compound NCCOP(O)(=O)OCC(O)CO JZNWSCPGTDBMEW-UHFFFAOYSA-N 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 239000000741 silica gel Substances 0.000 description 2
- 229910002027 silica gel Inorganic materials 0.000 description 2
- 238000010898 silica gel chromatography Methods 0.000 description 2
- IIZPXYDJLKNOIY-JXPKJXOSSA-N 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/CCCCC IIZPXYDJLKNOIY-JXPKJXOSSA-N 0.000 description 1
- XYZZKVRWGOWVGO-UHFFFAOYSA-N Glycerol-phosphate Chemical compound OP(O)(O)=O.OCC(O)CO XYZZKVRWGOWVGO-UHFFFAOYSA-N 0.000 description 1
- 238000005904 alkaline hydrolysis reaction Methods 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 239000003957 anion exchange resin Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000001925 catabolic effect Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- OEYIOHPDSNJKLS-UHFFFAOYSA-N choline Chemical compound C[N+](C)(C)CCO OEYIOHPDSNJKLS-UHFFFAOYSA-N 0.000 description 1
- 229960001231 choline Drugs 0.000 description 1
- 230000003930 cognitive ability Effects 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 235000014113 dietary fatty acids Nutrition 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000000194 fatty acid Substances 0.000 description 1
- 229930195729 fatty acid Natural products 0.000 description 1
- 239000004519 grease Substances 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 239000000787 lecithin Substances 0.000 description 1
- 229940067606 lecithin Drugs 0.000 description 1
- 235000010445 lecithin Nutrition 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 239000003495 polar organic solvent Substances 0.000 description 1
- 238000013341 scale-up Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 239000002351 wastewater Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic Table
- C07F9/02—Phosphorus compounds
- C07F9/06—Phosphorus compounds without P—C bonds
- C07F9/08—Esters of oxyacids of phosphorus
- C07F9/09—Esters of phosphoric acids
- C07F9/091—Esters of phosphoric acids with hydroxyalkyl compounds with further substituents on alkyl
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
Abstract
A process for preparing natural L- α -glycero-phosphatidylcholine includes such steps as dissolving high-purity phosphatidylcholine in polar solvent, adding alkaline compound, degradation reaction, neutralizing with acid, vacuum concentrating, layering to obtain the lower L- α -glycero-phosphatidylcholine, dissolving the L- α -glycero-phosphatidylcholine in alcohol while adding the mixture of resin and alumina, stirring, filtering, vacuum concentrating, cooling, crystallizing, filtering to obtain L- α -glycero-phosphatidylcholine crystal, dissolving the L- α -glycero-phosphatidylcholine crystal in purified water to obtain L- α -glycero-phosphatidylcholine solution, decoloring, filtering, vacuum concentrating, and further purifying to remove oily compound.
Description
Technical Field
The invention belongs to the technical field of natural medicine preparation, and particularly relates to a method for preparing natural L- α -glycerophosphatidylcholine.
Background
L- α -glycerophosphatidylcholine (L- α -GPC) is a product formed by hydrolysis of two fatty acid chains on a phospholipid molecule, and is a catabolic product of lecithin in a human body, plays an extremely important role in synthesis and metabolism of phospholipid, and has extremely important significance for improvement of human cognitive ability, maintenance of hormone level and the like.
At present, a method for preparing natural L- α -glycerophosphatidylcholine generally adopts soybean powdered phospholipid as an initial raw material, and comprises the steps of degrading the soybean powdered phospholipid by sodium methoxide, removing impurities such as glycerophosphatidylethanolamine, ethanolamine, glycerophosphatidylinositol, glycerophosphatidic acid and the like by a chromatography method, removing other impurities such as neutral grease, fatty acid, choline and the like by resin, decoloring by using activated carbon, and concentrating to obtain a product.
In the separation and purification methods adopted in chinese patents CN102516292A and CN102964377A, a GPC sample to be separated is first dissolved in a low carbon alcohol, and then a non-polar organic solvent is added to the low carbon alcohol solution to precipitate GPC.
The separation and purification method adopted in Chinese patents CN102875592A and CN103172659A is to pass GPC sample to be separated through cation exchange resin and anion exchange resin in turn, and the method has large waste water amount and low productivity.
The method adopted by the Chinese patent CN102093410A is that the alkaline hydrolysis liquid is firstly subjected to silica gel column chromatography separation, and then is subjected to cation exchange resin column chromatography separation, the method can obtain a high-content L- α -glycerol phosphatidylcholine product, but when the silica gel column chromatography is carried out, because an eluant can dissolve part of silica gel and enter GPC, the subsequent operation is difficult to remove silica gel impurities.
Disclosure of Invention
The invention aims to provide a method for preparing natural L- α -glycerophosphatidylcholine, which is beneficial to obviously improving the purity so as to provide convenience for subsequent treatment and simplify the operation process so as to meet the requirement of industrial scale-up production.
The task of the present invention is accomplished by a method for preparing natural L- α -glycerophosphatidylcholine, comprising the steps of:
A) preparing a L- α -glycerophosphatidylcholine crude product, putting high-purity phosphatidylcholine into a polar solvent to prepare a raw material solution, stirring and dissolving, adding an alkaline compound to carry out degradation reaction, controlling the weight ratio of the alkaline compound to the high-purity phosphatidylcholine, controlling the degradation reaction temperature and controlling the degradation reaction time, neutralizing with acid after the degradation reaction is finished, carrying out reduced pressure concentration, and layering to obtain a lower L- α -glycerophosphatidylcholine crude product;
B) preparing L- α -glycerophosphatidylcholine crystals, adding the crude L- α -glycerophosphatidylcholine obtained in the step A) into ethanol, stirring for dissolving, adding a mixture of resin and alumina, stirring for refining, filtering, concentrating the filtrate under reduced pressure, cooling for crystallization, and filtering to obtain L- α -glycerophosphatidylcholine crystals;
C) refining, namely putting the L- α -glycerophosphatidylcholine crystal obtained in the step B) into purified water to be dissolved into L- α -glycerophosphatidylcholine solution, adding a decolorizing agent for decolorization, filtering, and then concentrating under reduced pressure to obtain the high-purity natural L- α -glycerophosphatidylcholine.
In a specific embodiment of the present invention, the weight ratio of the basic compound to the highly purified phosphatidylcholine in the step A) is controlled to be 1: 20-200; in the raw material liquid, the mass concentration of the phosphatidylcholine is 100-500 g/L.
In another specific embodiment of the present invention, the polar solvent in step a) is ethanol, propanol or butanol; the alkaline compound is sodium ethoxide, sodium methoxide or sodium hydroxide; the degradation reaction temperature is controlled to be 0-40 ℃, and the degradation reaction time is controlled to be 3-22 h.
In still another embodiment of the present invention, the neutralization with acid in step A) refers to neutralization with acetic acid until the pH value is 6-7, and the mass concentration of the crude L- α -glycerophosphatidylcholine is 200 g/L.
In another embodiment of the present invention, the content of the highly pure phosphatidylcholine is greater than or equal to 96%.
In still another specific embodiment of the invention, the weight ratio of the L- α -glycerophosphatidylcholine crude product to the ethanol in the step B) is 1: 1-3, the mass ratio of the resin, the alumina mixture and the L- α -glycerophosphatidylcholine crude product is 0.01-0.3: 1, the stirring and refining temperature is 0-4 ℃, the stirring and refining time is 30-120min, the cooling and crystallizing temperature is-5 ℃, and the cooling and crystallizing time is 0.5-24 h.
In a more specific embodiment of the present invention, the resins are ion exchange resin 711 and D001 resin, and the volume ratio of the ion exchange resin to the D001 resin is 2.8-3.2: 1; the ethanol is absolute ethanol.
In still another embodiment of the present invention, the mass concentration of L- α -glycerophosphatidylcholine in the L- α -glycerophosphatidylcholine solution in step C) is 50-100 g/L.
In a still more specific embodiment of the invention, the mass ratio of the decoloring agent to the L- α -glycerophosphatidylcholine crystals in the step C) is 1: 100-200, and the filtering is performed by using a filter element of a 0.22 μm filter.
In yet another specific embodiment of the present invention, the decolorizing agent is activated carbon that removes chloride ions.
The technical scheme provided by the invention has the technical effects that as high-purity phosphatidylcholine is used as a raw material to carry out degradation reaction with an alkaline compound, impurities in phospholipid can be controlled from the source, the impurities such as glycerol phosphatidylethanolamine, glycerol phosphatidylinositol, glycerol phosphoric acid and the like generated by the reaction are reduced, and convenience is provided for subsequent treatment, as the mixture of resin and alumina is added into the crude product of L- α -glycerol phosphatidylcholine in the refining step B) to be stirred and refined, the oil compounds can be further purified and removed through crystallization, a foundation is laid for the purity of more than 99.5 percent and the molar yield of more than 75 percent, and as a filter element is adopted for filtering by a 0.22 micron filter without depending on chromatography, the whole preparation process is simple, and the requirement of industrial amplification production can be met.
The specific implementation mode is as follows:
example 1:
A) preparing a crude product of L- α -glycerophosphatidylcholine, putting high-purity phosphatidylcholine which is purchased from a commercial channel and has a content of more than 96%, namely a mass percentage content of more than 96% into ethanol serving as a polar solvent to prepare a raw material liquid, stirring and dissolving the raw material liquid, adding sodium ethoxide serving as an alkaline compound to carry out degradation reaction, wherein the weight ratio of the sodium ethoxide to the high-purity phosphatidylcholine is 1: 100, the degradation reaction temperature is 10 ℃, the degradation reaction time is 18h, neutralizing with acetic acid until the pH value is 6 after the degradation reaction is finished, and then carrying out reduced pressure concentration and layering to obtain a crude product of L- α -glycerophosphatidylcholine with a lower layer and a mass concentration of 100 g/L;
B) preparing L- α -glycerophosphatidylcholine crystals, adding the crude L- α -glycerophosphatidylcholine with the mass concentration of 100g/L obtained in the step A) into ethanol, stirring and dissolving, wherein the weight ratio of the crude glycerophosphatidylcholine to the ethanol is 1: 1, simultaneously adding a mixture of resin and alumina, stirring and refining, the stirring and refining temperature is 4 ℃, the stirring and refining time is 30min, filtering is carried out after refining is finished, the filtrate obtained by filtering is concentrated, cooling and crystallizing are carried out at the temperature of minus 5 ℃ for 24h, and filtering is carried out to obtain the L- α -glycerophosphatidylcholine crystals, wherein the mass ratio of the resin, the alumina mixture and the crude L- α -glycerophosphatidylcholine in the step is 0.08: 0.15: 1, the resin is an ion exchange resin 711 and a D001 resin, the volume ratio of the ion exchange resin 711 to the D001 resin is 3.2: 1, and the ethanol in the step is absolute ethanol;
C) and (3) refining, namely putting the L- α -glycerophosphatidylcholine crystal obtained in the step B) into purified water such as purified water, deionized water or deep desalted water to be dissolved into an L- α -glycerophosphatidylcholine solution, adding activated carbon for removing chloride ions as a decoloring agent into the L- α -glycerophosphatidylcholine solution, decoloring for 30min at 45 ℃, wherein the mass ratio of the activated carbon for removing the chloride ions to the L- α -glycerophosphatidylcholine crystal is 1: 150, filtering by using a filter element with the size of 0.22 mu m after decoloring is finished, and then concentrating under reduced pressure to obtain the L- α -glycerophosphatidylcholine with the purity of 99.65% and the molar yield of 75.81%.
Example 2:
A) preparing a L- α -glycerophosphatidylcholine crude product, putting high-purity phosphatidylcholine which is purchased from a commercial channel and has the content of more than 97 percent, namely the mass percentage content of more than 97 percent into butanol serving as a polar solvent to prepare a raw material liquid, stirring and dissolving the raw material liquid to obtain the phosphatidylcholine with the mass concentration of 100g/L, adding sodium methoxide serving as an alkaline compound to carry out degradation reaction, wherein the weight ratio of the sodium methoxide to the high-purity phosphatidylcholine is 1: 200, the temperature of the degradation reaction is 0 ℃, the time of the degradation reaction is 22h, neutralizing the raw material liquid with acetic acid until the pH value is 7 after the degradation reaction is finished, and then carrying out reduced pressure concentration and layering to obtain the L- α -glycerophosphatidylcholine crude product with the lower layer and the mass concentration of 200 g/L;
B) preparing L- α -glycerophosphatidyl choline crystals, adding the crude L- α -glycerophosphatidyl choline product with the mass concentration of 200g/L obtained in the step A) into ethanol, stirring and dissolving, wherein the weight ratio of the crude glycerophosphatidyl choline product to the ethanol is 3: 1, adding a mixture of resin and alumina, stirring and refining, the stirring and refining temperature is 0 ℃, the stirring and refining time is 120min, filtering after refining is finished, concentrating the filtrate obtained by filtering, cooling and crystallizing at 0 ℃ for 12h, and filtering to obtain the L- α -glycerophosphatidyl choline crystals, wherein the mass ratio of the resin to the mixture of the alumina and the crude L- α -glycerophosphatidyl choline product is 0.01: 0.3: 1, the resin is an ion exchange resin 711 and a D001 resin, the volume ratio of the ion exchange resin 711 to the D001 resin is 3: 1, and the ethanol in the step is absolute ethanol;
C) and (2) refining, namely putting the L- α -glycerophosphatidylcholine crystal obtained in the step B) into purified water such as purified water, deionized water or deep desalted water to be dissolved into an L- α -glycerophosphatidylcholine solution, adding activated carbon for removing chloride ions as a decoloring agent into the L- α -glycerophosphatidylcholine solution, decoloring for 30min at 45 ℃, wherein the mass ratio of the activated carbon for removing the chloride ions to the L- α -glycerophosphatidylcholine crystal is 1: 100, filtering by using a filter element with the size of 0.22 mu m after decoloring is finished, and then concentrating under reduced pressure to obtain the L- α -glycerophosphatidylcholine with the purity of 99.64% and the molar yield of 75.15%.
Example 3:
A) preparing a crude product of L- α -glycerophosphatidylcholine, putting high-purity phosphatidylcholine which is purchased from a commercial channel and has a content of more than 96.5%, namely a mass percentage content of more than 96.5% into propanol serving as a polar solvent to prepare a raw material liquid, stirring and dissolving the raw material liquid, adding sodium hydroxide serving as an alkaline compound to carry out degradation reaction, wherein the weight ratio of the sodium hydroxide to the high-purity phosphatidylcholine is 1: 20, the degradation reaction temperature is 40 ℃, the degradation reaction time is 3 hours, neutralizing with acetic acid after the degradation reaction is finished until the pH value is 6.3, carrying out reduced pressure concentration, and layering to obtain a crude product of L- α -glycerophosphatidylcholine with a lower layer and a mass concentration of 150 g/L;
B) preparing L- α -glycerophosphatidylcholine crystals, adding 150 g/L- α -glycerophosphatidylcholine crude product obtained in the step A) into ethanol, stirring and dissolving, wherein the weight ratio of the glycerol phosphatidylcholine crude product to the ethanol is 2.5: 1, simultaneously adding a mixture of resin and alumina, stirring and refining at the stirring and refining temperature of 1 ℃, stirring and refining for 90min, filtering after refining, concentrating the filtrate obtained by filtering, cooling and crystallizing at 2 ℃ for 5h, and filtering to obtain L- α -glycerophosphatidylcholine crystals, wherein the mass ratio of the resin, the alumina mixture and the L- α -glycerophosphatidylcholine crude product is 0.15: 0.08: 1, the resin is ion exchange resin 711 and D001 resin, the volume ratio of the ion exchange resin 711 to D001 resin is 2.8: 1, and the ethanol in the step is absolute ethanol;
C) and (2) refining, namely putting the L- α -glycerophosphatidylcholine crystal obtained in the step B) into purified water such as purified water, deionized water or deep desalted water to be dissolved into an L- α -glycerophosphatidylcholine solution, adding activated carbon for removing chloride ions as a decoloring agent into the L- α -glycerophosphatidylcholine solution, decoloring for 30min at 45 ℃, wherein the mass ratio of the activated carbon for removing the chloride ions to the L- α -glycerophosphatidylcholine crystal is 1: 200, filtering by using a filter element with the size of 0.22 mu m after decoloring is finished, and then concentrating under reduced pressure to obtain the L- α -glycerophosphatidylcholine with the purity of 99.58% and the molar yield of 76.42%.
Example 4:
A) preparing a crude product of L- α -glycerophosphatidylcholine, putting high-purity phosphatidylcholine which is purchased from a commercial channel and has a content of more than 97.3%, namely a mass percentage content of more than 97.3% into ethanol serving as a polar solvent to prepare a raw material liquid, stirring and dissolving the raw material liquid, adding sodium hydroxide serving as an alkaline compound to carry out degradation reaction, wherein the weight ratio of the sodium hydroxide to the high-purity phosphatidylcholine is 1: 150, the degradation reaction temperature is 25 ℃, the degradation reaction time is 8 hours, neutralizing with acetic acid after the degradation reaction is finished until the pH value is 6.7, carrying out reduced pressure concentration, and layering to obtain a lower L- α -glycerophosphatidylcholine crude product with a mass concentration of 180 g/L;
B) preparing L- α -glycerophosphatidylcholine crystals, adding the L- α -glycerophosphatidylcholine crude product with the mass concentration of 180g/L obtained in the step A) into ethanol, stirring and dissolving, wherein the weight ratio of the glycerophosphatidylcholine crude product to the ethanol is 2: 1, simultaneously adding a resin and alumina mixture, stirring and refining, the stirring and refining temperature is 2.5 ℃, the stirring and refining time is 60min, filtering is carried out after refining is finished, the filtrate obtained by filtering is concentrated, cooling and crystallizing are carried out at 5 ℃ for 0.5h, and filtering is carried out to obtain the L- α -glycerophosphatidylcholine crystals, wherein the mass ratio of the resin, the alumina mixture and the L- α -glycerophosphatidylcholine crude product in the step is 0.3: 0.01: 1, the resin is ion exchange resin 711 and D001 resin, the volume ratio of the ion exchange resin 711 to D001 resin is 3.1: 1, and the ethanol in the step is absolute ethanol;
C) and (3) refining, namely putting the L- α -glycerophosphatidylcholine crystals obtained in the step B) into purified water such as purified water, deionized water or deep desalted water to be dissolved into an L- α -glycerophosphatidylcholine solution, adding activated carbon for removing chloride ions as a decoloring agent into the L- α -glycerophosphatidylcholine solution, decoloring for 30min at 45 ℃, wherein the mass ratio of the activated carbon for removing the chloride ions to the L- α -glycerophosphatidylcholine crystals is 1: 180, filtering by using a filter element of 0.22 mu m after decoloring is finished, and then concentrating under reduced pressure to obtain the L- α -glycerophosphatidylcholine with the purity of 99.69% and the molar yield of 77.12%.
In conclusion, the technical scheme provided by the invention overcomes the defects in the prior art, successfully completes the invention task and truly realizes the technical effects of the applicant in the technical effect column.
Claims (10)
1. A method for preparing natural L- α -glycerophosphatidylcholine, comprising the steps of:
A) preparing a L- α -glycerophosphatidylcholine crude product, putting high-purity phosphatidylcholine into a polar solvent to prepare a raw material solution, stirring and dissolving, adding an alkaline compound to carry out degradation reaction, controlling the weight ratio of the alkaline compound to the high-purity phosphatidylcholine, controlling the degradation reaction temperature and controlling the degradation reaction time, neutralizing with acid after the degradation reaction is finished, carrying out reduced pressure concentration, and layering to obtain a lower L- α -glycerophosphatidylcholine crude product;
B) preparing L- α -glycerophosphatidylcholine crystals, adding the crude L- α -glycerophosphatidylcholine obtained in the step A) into ethanol, stirring for dissolving, adding a mixture of resin and alumina, stirring for refining, filtering, concentrating the filtrate under reduced pressure, cooling for crystallization, and filtering to obtain L- α -glycerophosphatidylcholine crystals;
C) refining, namely putting the L- α -glycerophosphatidylcholine crystal obtained in the step B) into purified water to be dissolved into L- α -glycerophosphatidylcholine solution, adding a decolorizing agent for decolorization, filtering, and then concentrating under reduced pressure to obtain the high-purity natural L- α -glycerophosphatidylcholine.
2. The method according to claim 1, wherein the weight ratio of the basic compound to the highly purified phosphatidylcholine in step A) is controlled to be 1: 20-200, and the concentration of the phosphatidylcholine in the raw material solution is 100-500 g/L.
3. The method of claim 1, wherein the polar solvent used in step A) is ethanol, propanol or butanol, the basic compound is sodium ethoxide, sodium methoxide or sodium hydroxide, the degradation reaction temperature is controlled to 0-40 ℃, and the degradation reaction time is controlled to 3-22 h.
4. The method for preparing natural L- α -glycerophosphatidylcholine according to claim 1, wherein the neutralization with acid in step A) is performed until the pH value is 6-7, and the crude L- α -glycerophosphatidylcholine has a mass concentration of 100-200 g/L.
5. The method for preparing natural L- α -glycerophosphatidylcholine according to claim 1 or 2, wherein the content of highly pure phosphatidylcholine is greater than or equal to 96%.
6. The method for preparing natural L- α -glycerophosphatidylcholine according to claim 1, wherein the weight ratio of L- α -glycerophosphatidylcholine crude product to ethanol in step B) is 1: 1-3, the mass ratio of the resin, the alumina mixture and the L- α -glycerophosphatidylcholine crude product is 0.01-0.3: 1, the temperature for stirring and refining is 0-4 ℃, the time for stirring and refining is 30-120min, the temperature for cooling and crystallizing is-5 to 5 ℃, and the time for cooling and crystallizing is 0.5-24 h.
7. The method for preparing natural L- α -glycerophosphatidylcholine according to claim 1 or 6, wherein the resin is ion exchange resin 711 and D001 resin, the volume ratio of the ion exchange resin to the D001 resin is 2.8-3.2: 1, and the ethanol is absolute ethanol.
8. The method for preparing natural L- α -glycerophosphatidylcholine according to claim 1, wherein the L- α -glycerophosphatidylcholine solution in step C) has a mass concentration of L- α -glycerophosphatidylcholine of 50-100 g/L.
9. The method for preparing natural L- α -glycerophosphatidylcholine according to claim 1, wherein the mass ratio of the decoloring agent to the L- α -glycerophosphatidylcholine crystals in step C) is 1: 100-200, and the filtering is performed by using a filter element as a 0.22 μm filter.
10. The method of claim 1 or 9, wherein the decolorizing agent is chloride-depleted activated carbon.
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