CH642083A5 - PROCESS FOR THE PREPARATION OF L-ALPHA-GLYCERYLPHOSPHORYLCHOLINE. - Google Patents
PROCESS FOR THE PREPARATION OF L-ALPHA-GLYCERYLPHOSPHORYLCHOLINE. Download PDFInfo
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- CH642083A5 CH642083A5 CH90080A CH90080A CH642083A5 CH 642083 A5 CH642083 A5 CH 642083A5 CH 90080 A CH90080 A CH 90080A CH 90080 A CH90080 A CH 90080A CH 642083 A5 CH642083 A5 CH 642083A5
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- Switzerland
- Prior art keywords
- glycerylphosphorylcholine
- lecithin
- preparation
- solution
- moles
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- 238000000034 method Methods 0.000 title claims description 13
- 238000002360 preparation method Methods 0.000 title claims description 7
- 239000008777 Glycerylphosphorylcholine Substances 0.000 title description 10
- SUHOQUVVVLNYQR-MRVPVSSYSA-N choline alfoscerate Chemical compound C[N+](C)(C)CCOP([O-])(=O)OC[C@H](O)CO SUHOQUVVVLNYQR-MRVPVSSYSA-N 0.000 title 1
- 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 claims description 18
- 229940067606 lecithin Drugs 0.000 claims description 17
- 235000010445 lecithin Nutrition 0.000 claims description 17
- 239000000787 lecithin Substances 0.000 claims description 17
- WQDUMFSSJAZKTM-UHFFFAOYSA-N Sodium methoxide Chemical compound [Na+].[O-]C WQDUMFSSJAZKTM-UHFFFAOYSA-N 0.000 claims description 12
- 238000006140 methanolysis reaction Methods 0.000 claims description 5
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 15
- 239000000243 solution Substances 0.000 description 10
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 9
- 229960004956 glycerylphosphorylcholine Drugs 0.000 description 9
- SUHOQUVVVLNYQR-MRVPVSSYSA-O glycerylphosphorylcholine Chemical compound C[N+](C)(C)CCO[P@](O)(=O)OC[C@H](O)CO SUHOQUVVVLNYQR-MRVPVSSYSA-O 0.000 description 8
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 7
- 230000007062 hydrolysis Effects 0.000 description 7
- 238000006460 hydrolysis reaction Methods 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 6
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 6
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 6
- YKYOUMDCQGMQQO-UHFFFAOYSA-L cadmium dichloride Chemical compound Cl[Cd]Cl YKYOUMDCQGMQQO-UHFFFAOYSA-L 0.000 description 6
- 238000002425 crystallisation Methods 0.000 description 6
- 230000008025 crystallization Effects 0.000 description 6
- 238000005227 gel permeation chromatography Methods 0.000 description 6
- 150000002632 lipids Chemical class 0.000 description 5
- 239000000203 mixture Substances 0.000 description 5
- 238000000746 purification Methods 0.000 description 5
- 239000011347 resin Substances 0.000 description 5
- 229920005989 resin Polymers 0.000 description 5
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 4
- -1 fatty acid esters Chemical class 0.000 description 4
- VDZOOKBUILJEDG-UHFFFAOYSA-M tetrabutylammonium hydroxide Chemical compound [OH-].CCCC[N+](CCCC)(CCCC)CCCC VDZOOKBUILJEDG-UHFFFAOYSA-M 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 229960001231 choline Drugs 0.000 description 3
- 235000014113 dietary fatty acids Nutrition 0.000 description 3
- 239000008344 egg yolk phospholipid Substances 0.000 description 3
- 239000000194 fatty acid Substances 0.000 description 3
- 229930195729 fatty acid Natural products 0.000 description 3
- 239000012467 final product Substances 0.000 description 3
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 102000004895 Lipoproteins Human genes 0.000 description 2
- 108090001030 Lipoproteins Proteins 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
- 230000003197 catalytic effect Effects 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- WORJEOGGNQDSOE-UHFFFAOYSA-N chloroform;methanol Chemical compound OC.ClC(Cl)Cl WORJEOGGNQDSOE-UHFFFAOYSA-N 0.000 description 2
- 238000004587 chromatography analysis Methods 0.000 description 2
- 238000003776 cleavage reaction Methods 0.000 description 2
- 238000001704 evaporation Methods 0.000 description 2
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 description 2
- 150000002500 ions Chemical class 0.000 description 2
- 210000004185 liver Anatomy 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 230000007017 scission Effects 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- PZNPLUBHRSSFHT-RRHRGVEJSA-N 1-hexadecanoyl-2-octadecanoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCCCC(=O)O[C@@H](COP([O-])(=O)OCC[N+](C)(C)C)COC(=O)CCCCCCCCCCCCCCC PZNPLUBHRSSFHT-RRHRGVEJSA-N 0.000 description 1
- 208000001378 Carbon Tetrachloride Poisoning Diseases 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 235000013400 Quercus lobata Nutrition 0.000 description 1
- 240000001749 Quercus lobata Species 0.000 description 1
- FKNQFGJONOIPTF-UHFFFAOYSA-N Sodium cation Chemical compound [Na+] FKNQFGJONOIPTF-UHFFFAOYSA-N 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 229910001860 alkaline earth metal hydroxide Inorganic materials 0.000 description 1
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000008346 aqueous phase Substances 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000004061 bleaching Methods 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 229910052793 cadmium Inorganic materials 0.000 description 1
- BDOSMKKIYDKNTQ-UHFFFAOYSA-N cadmium atom Chemical compound [Cd] BDOSMKKIYDKNTQ-UHFFFAOYSA-N 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
- OEYIOHPDSNJKLS-UHFFFAOYSA-N choline Chemical compound C[N+](C)(C)CCO OEYIOHPDSNJKLS-UHFFFAOYSA-N 0.000 description 1
- 238000013375 chromatographic separation Methods 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 235000013345 egg yolk Nutrition 0.000 description 1
- 210000002969 egg yolk Anatomy 0.000 description 1
- 235000013601 eggs Nutrition 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 239000003480 eluent Substances 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- 229910001385 heavy metal Inorganic materials 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 230000008595 infiltration Effects 0.000 description 1
- 238000001764 infiltration Methods 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- JILPJDVXYVTZDQ-UHFFFAOYSA-N lithium methoxide Chemical compound [Li+].[O-]C JILPJDVXYVTZDQ-UHFFFAOYSA-N 0.000 description 1
- 210000005228 liver tissue Anatomy 0.000 description 1
- 229960002523 mercuric chloride Drugs 0.000 description 1
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 description 1
- 239000000401 methanolic extract Substances 0.000 description 1
- NBTOZLQBSIZIKS-UHFFFAOYSA-N methoxide Chemical compound [O-]C NBTOZLQBSIZIKS-UHFFFAOYSA-N 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 125000001453 quaternary ammonium group Chemical group 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000000284 resting effect Effects 0.000 description 1
- 230000009291 secondary effect Effects 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- RMAQACBXLXPBSY-UHFFFAOYSA-N silicic acid Chemical compound O[Si](O)(O)O RMAQACBXLXPBSY-UHFFFAOYSA-N 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 235000012239 silicon dioxide Nutrition 0.000 description 1
- 229910001415 sodium ion Inorganic materials 0.000 description 1
- 239000011877 solvent mixture Substances 0.000 description 1
- 239000008347 soybean phospholipid Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 239000003643 water by type 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
Landscapes
- 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)
- Low-Molecular Organic Synthesis Reactions Using Catalysts (AREA)
Description
La presente invenzione ha per oggetto un procedimento per la preparazione industriale di L-a-glicerilfosforilcolina (I) allo stato di elevata purezza. The present invention relates to a process for the industrial preparation of L-a-glycerylphosphorylcholine (I) in the state of high purity.
È noto che la L-a-glicerilfosforilcolina (I) L-a-glycerylphosphorylcholine (I) is known
ch2oh « ch2oh «
ho-g-h ch2-0-p ho-g-h ch2-0-p
0 0
/ /
0 0
(I) (THE)
o-ch2-ch2-n(ch3)3 o-ch2-ch2-n (CH3) 3
somministrata per via intramuscolare o orale protegge in modo molto evidente il tessuto epatico dalla infiltrazione grassa indotta da dieta iperlipidica-ipoproteica o determinata da avvelenamento da tetracloruro di carbonio e pure protegge dalla constasi sperimentale. administered intramuscularly or orally, it protects the liver tissue from fat infiltration induced by hyperlipid-hypoprotein diet or determined by carbon tetrachloride poisoning and also protects from experimental findings.
È importante mettere in evidenza che la glicerilfosforilcolina entra nella costituzione delle lipoproteine. Ricerche di farmaco-cinetica dimostrano che la glicerilfosforilcolina si assorbe rapidamente dall'intestino o dal focolaio di inoculazione intramuscolare: dal circolo si distribuisce agli organi, specialmente al fegato, al rene ed al cervello. Particolarmente dal fegato è nuovamente reimmessa in circolo, incorporata nelle lipoproteine. Questo particolare dato spiega il motivo per il quale la glicerilfosforilcolina manifesta un'azione chiarificante il siero. It is important to highlight that glycerylphosphorylcholine enters the constitution of lipoproteins. Pharmacokinetic research shows that glycerylphosphorylcholine is rapidly absorbed from the intestine or the outbreak of intramuscular inoculation: from the circulation it is distributed to the organs, especially the liver, kidney and brain. Particularly from the liver it is reintroduced into the circulation, incorporated into the lipoproteins. This particular data explains the reason why glycerylphosphorylcholine exhibits a clarifying action on serum.
È altresì noto che (I) viene ottenuta per idrolisi della lecitina, in particolare della lecitina di uovo. It is also known that (I) is obtained by hydrolysis of lecithin, in particular egg lecithin.
La maggior difficoltà di questa operazione è costituita dalla purificazione; i prodotti secondari che accompagnano la L-a-glicerilfosforlicolina (I) ottenuta per idrolisi non soltanto ne diminuiscono il valore terapeutico, ma presentano spesso sfavorevoli effetti secondari. Non sono quindi mancati tentativi per ottenere (I) in forma particolarmente pura. The greatest difficulty of this operation is constituted by purification; the secondary products that accompany the L-a-glycerylphosphoricoline (I) obtained by hydrolysis not only decrease its therapeutic value, but often have unfavorable secondary effects. Therefore attempts to obtain (I) in a particularly pure form were not lacking.
Così, in USP 2 864 848 del 16 dicembre 1958 si descrive l'idrolisi in presenza di cloruro mercurico, allo scopo di precipitare i componenti secondari sotto forma di sali mercurici; ciò comporta la necessità di allontanare poi dalla glicerilfosforilcolina gli ioni mercurici (dei quali è superfluo sottolineare la tossicità), e ciò viene realizzato mediante un complicato trattamento con HiS e BaCC>3. Lo stesso brevetto, tuttavia, avverte che sovente tale trattamento non basta a eliminare tutti gli ioni mercurici, e a tal fine si rendono necessarie ulteriori operazioni. Per di più il prodotto finale dev'essere purificato attraverso la formazione di un complesso con cloruro di cadmio. L'operazione è quindi assai complicata, il prodotto finale non esente da metalli pesanti. Thus, in USP 2 864 848 of 16 December 1958 hydrolysis in the presence of mercuric chloride is described, in order to precipitate the secondary components in the form of mercuric salts; this entails the need to remove the mercuric ions (of which it is superfluous to underline the toxicity) from glycerylphosphorylcholine, and this is accomplished by means of a complicated treatment with HiS and BaCC> 3. The same patent, however, warns that often this treatment is not enough to eliminate all the mercuric ions, and further operations are necessary for this purpose. Furthermore, the final product must be purified through the formation of a complex with cadmium chloride. The operation is therefore very complicated, the final product not free of heavy metals.
Una tecnica analoga alla precedente è descritta in Biochemi-cal Préparation (Vol. 6, pag. 16-19); l'eliminazione finale del 5 cadmio è in questo caso effettuata mediante passaggio su una miscela di resine scambiatrici, che rende il procedimento ancora più complesso e deprime ulteriormente le rese. A technique similar to the previous one is described in Biochemi-cal Préparation (Vol. 6, pages 16-19); the final elimination of the 5 cadmium is in this case carried out by passing it on a mixture of exchange resins, which makes the process even more complex and further depresses the yields.
Brocherhoff [Journal Lipid Research 4, 96 (1963)] descrive la metanolisi di una soluzione molto diluita di lecitina (circa 21 io g/litro) con metossido di sodio in ragione di 3 moli di metossido per mole di fosfatide. Brocherhoff [Journal Lipid Research 4, 96 (1963)] describes the methanolysis of a very dilute solution of lecithin (about 21 g / liter) with sodium methoxide in the ratio of 3 moles of methoxide per mole of phosphatide.
In queste condizioni si verifica tuttavia, oltre alla desiderata scissione del legame acile-ossigeno, anche una parziale scissione del legame P-O, con formazione di sottoprodotti. Under these conditions, however, in addition to the desired cleavage of the acyl-oxygen bond, a partial cleavage of the P-O bond also occurs, with the formation of by-products.
15 Okui e collaboratori [Yakugaku Zasshi (12), 1206-9 (1964)] descrivono la preparazione di (I) da lecitina di uovo mediante idrolisi con idrossidi di metalli alcalinoterrosi. Il prodotto così ottenuto risulta notevolmente impuro di componenti secondari. 15 Okui and collaborators [Yakugaku Zasshi (12), 1206-9 (1964)] describe the preparation of (I) from egg lecithin by hydrolysis with alkaline earth metal hydroxides. The product thus obtained is remarkably impure of secondary components.
20 Brockerhoff e Yurkowski descrivono in una breve nota [Ca-nadian Journal of Biochemistry, 43, 1777 (1965)] la preparazione di (I) mediante idrolisi di lecitina con una base ammonica quaternaria (idrossido di tetrabutilammonio): le rese sono soddisfacenti, come ha posto in evidenza anche Chadha [Chem. 25 Phys. Lipids, 4, 104-108 (1970)]; tuttavia l'idrolisi compiuta con questa base, commercialmente costosa, non consente di ottenere glicerilfosforilcolina esente da idrossido di tetrabutilammonio, rendendo pertanto necessaria la cristallizzazione di (I). Inoltre questo procedimento male si presta per preparazioni su 30 larga scala, mancando di riproducibilità. 20 Brockerhoff and Yurkowski describe in a short note [Ca-nadian Journal of Biochemistry, 43, 1777 (1965)] the preparation of (I) by hydrolysis of lecithin with a quaternary ammonium base (tetrabutylammonium hydroxide): the yields are satisfactory, as Chadha [Chem. 25 Phys. Lipids, 4, 104-108 (1970)]; however the hydrolysis carried out with this commercially expensive base does not allow to obtain glycerylphosphorylcholine free from tetrabutylammonium hydroxide, therefore making the crystallization of (I) necessary. Furthermore, this procedure is not suitable for large scale preparations, lacking reproducibility.
Secondo Cuberò Robles e Roels [Chem. Phys. Lipids, 6, 31-38 (1971)], si può ottenere L-a-glicerilfosforilcolina mediante metanolisi di un estratto lipidico ottenuto da polvere di tuorlo d'uovo. Il metodo descritto da questi autori comprende l'e-35 strazione della polvere con cloroformio/metanolo, l'evaporazione della miscela di solventi, la dissoluzione del residuo in etere e il trattamento della soluzione eterea con metossido di litio. Quest'ultimo viene usato in ragione di 1 mole/mole di fosfatide. Ne risulta una miscela di GPC e di GPE che, previa 40 neutralizzazione con HCl metanolico, viene separata mediante cromatografia su silice. In pratica si è osservato che questa cromatografia fornisce GPC inquinata da acido silicico colloidale (come era prevedibile, dato che l'eluente è acqua), eliminabile soltanto mediante ripetute cristallizzazioni (notoriamente diffi-45 coltose) del prodotto finale. According to Cuberò Robles and Roels [Chem. Phys. Lipids, 6, 31-38 (1971)], L-a-glycerylphosphorylcholine can be obtained by methanolysis of a lipid extract obtained from egg yolk powder. The method described by these authors includes extracting the powder with chloroform / methanol, evaporating the solvent mixture, dissolving the residue in ether and treating the ether solution with lithium methoxide. The latter is used in the ratio of 1 mole / mole of phosphatide. The result is a mixture of GPC and GPE which, after 40 neutralization with methanolic HCl, is separated by silica chromatography. In practice it has been observed that this chromatography provides GPC polluted by colloidal silicic acid (as was foreseeable, since the eluent is water), which can only be eliminated by repeated crystallizations (notoriously diffi cult) of the final product.
Riassumendo, si può affermare che i processi sinora descritti per ottenere GPC di soddisfacente purezza comportano inconvenienti che ne rendono insostituibile l'attuazione economica, soprattutto su scala industriale. In summary, it can be said that the processes described so far to obtain GPCs of satisfactory purity entail drawbacks which make their economic implementation irreplaceable, especially on an industrial scale.
so Si è ora inaspettatamente trovato che si può ottenere GPC pura con alte rese e con un metodo relativamente semplice, attuabile senza difficoltà su scala industriale, se si effettua la metanolisi della lecitina purificata in soluzione molto concentrata e operando con quantità catalitiche di metossido di sodio. 55 II prodotto al quale si perviene ha un grado di purezza tale da renderne superflua sia la cristallizzazione (che, come si è detto è particolarmente difficoltosa) sia la formazione del complesso con CdCl2, sostanzialmente inattuabile su scala industriale. so It has now been unexpectedly found that pure GPC can be obtained with high yields and with a relatively simple method, which can be carried out without difficulty on an industrial scale, if the methanolysis of the purified lecithin is carried out in a very concentrated solution and operating with catalytic quantities of sodium methoxide . 55 The product to be obtained has a degree of purity such as to make it unnecessary both crystallization (which, as has been said, is particularly difficult) and the formation of the complex with CdCl2, substantially unworkable on an industrial scale.
Secondo l'invenzione, la metanolisi è condotta trattando per 60 60', a temperatura ambiente, una soluzione metanolica di lecitina purificata (39% in peso di lecitina) con quantità catalitiche di CH}ONa (da 0,01 a 0,1, di preferenza 0,05 moli/mole). Al termine della reazione gli esteri degli acidi grassi che si sono separati durante la reazione vengono allontanati e la soluzione 65 metanolitica residua è trattata con circa 8 equivalenti di resina cationica debole IR-C 50 (H +) per equivalente di metossido di sodio, allo scopo di eliminare lo ione sodio. According to the invention, methanolysis is carried out by treating for 60 60 ', at room temperature, a methanolic solution of purified lecithin (39% by weight of lecithin) with catalytic quantities of CH} ONa (from 0.01 to 0.1, preferably 0.05 moles / moles). At the end of the reaction the esters of the fatty acids which separated during the reaction are removed and the residual methanolytic solution 65 is treated with about 8 equivalents of weak cationic resin IR-C 50 (H +) per equivalent of sodium methoxide, at aim to eliminate the sodium ion.
L'eluato e il lavaggio della resina sono concentrati a piccolo The eluate and the resin wash are concentrated to small
3 3
642 083 642 083
volume mediante evaporazione a pressione ridotta (12 mm Hg) e, dopo diluizione con acqua, sono estratti con cloroformio per allontanare le ultime tracce degli esteri degli acidi grassi, le eventuali tracce di lecitina non deacilata o parzialmente deacila-ta e le tracce di sfingomieline cha a volte accompagnano la lecitina di partenza. Si evapora la fase acquosa a secco a pressione ridotta, quindi, se necessario, si decolora il prodotto con carbone SL 50. volume by evaporation at reduced pressure (12 mm Hg) and, after dilution with water, are extracted with chloroform to remove the last traces of the fatty acid esters, any traces of non-deacylated or partially deacylated lecithin and traces of sphingomieline cha sometimes accompany the starting lecithin. The aqueous dry phase is evaporated under reduced pressure, then, if necessary, the product is decoloured with SL 50 carbon.
Il prodotto ottenuto è esente da impurezze di idrolisi, da sali inorganici e da esteri degli acidi grassi e non mostra differenze significative rispetto a campioni ottenuti per formazione del complesso con CdCl2, successiva cristallizzazione, decomposizione del medesimo per passaggio su resine scambiatrici e cristallizzazione finale della L-a-glicerilfosforilcolina così ottenuta. The product obtained is free from hydrolysis impurities, inorganic salts and fatty acid esters and does not show significant differences compared to samples obtained by forming the complex with CdCl2, subsequent crystallization, decomposition of the same by passage on exchange resins and final crystallization of the La-glycerylphosphorylcholine thus obtained.
La L-a-glicerilfosforilcolina ottenuta col metodo dell'invenzione contiene generalmente circa il 15% di umidità. The L-a-glycerylphosphorylcholine obtained by the method of the invention generally contains about 15% humidity.
L'umidità può essere eliminata, se nesessario, mediante essiccamento su P2O5 sotto alto vuoto e successiva cristallizzazione da etanolo/dietiletere o da etanolo/acetato di etile; si ottiene così un solido incolore igroscopico a p.f. 130-131°C, di formula bruta CgHhoNOfiP. The humidity can be eliminated, if necessary, by drying on P2O5 under high vacuum and subsequent crystallization from ethanol / diethyl ether or from ethanol / ethyl acetate; in this way a colorless hygroscopic solid is obtained at m.p. 130-131 ° C, of brute formula CgHhoNOfiP.
Tale operazione è ovviamente inutile qualora si debba utilizzare la L-a-glicerilfosforilcolina in soluzione acquosa oppure in associazione ad altri principi attivi in specialità farmaceutiche iniettabili liofilizzate, come quasi sempre accade; è allora possibile utilizzare il prodotto amorfo tal quale senza ricorrere a successiva purificazione poiché la purezza del fosfatide deacila-to è più che soddisfacente sotto ogni punto di vista. This operation is obviously useless if it is necessary to use L-a-glycerylphosphorylcholine in aqueous solution or in combination with other active ingredients in freeze-dried injectable pharmaceutical specialties, as almost always happens; it is then possible to use the amorphous product as it is without resorting to subsequent purification since the purity of the deacylated phosphatide is more than satisfactory from every point of view.
Gli esempi che seguono valgono a illustrare il procedimento secondo l'invenzione, senza peraltro limitarla in modo alcuno. The following examples are worth illustrating the process according to the invention, without however limiting it in any way.
Esempio 1 Example 1
a) Purificazione della lecitina a) Purification of lecithin
Si impiega come materia prima la lecitina da uovo grezza (P alcalilabile: 2,7-2,85%; P della glicerilfosforilcolina: 2,24-2,52%). Raw lecithin from egg is used as raw material (alkalizable P: 2.7-2.85%; P of glycerylphosphorylcholine: 2.24-2.52%).
Una soluzione di 0,5 kg di lecitina grezza in cloruro di metilene 1 litro è versata, sotto agitazione a 5°C, in 6 litri di acetone. Dopo 60' si decantano le acque, quindi si ripete la purificazione sciogliendo il residuo in cloruro di metilene (1 litro) e versando nuovamente la soluzione in acetone (6 litri). Si filtra e si essicca il precipitato sotto vuoto (12 mm Hg) a temperatura ambiente (320 g). A solution of 0.5 kg of crude lecithin in methylene chloride 1 liter is poured, under stirring at 5 ° C, into 6 liters of acetone. After 60 'the waters are decanted, then the purification is repeated by dissolving the residue in methylene chloride (1 liter) and pouring again the solution in acetone (6 liters). The precipitate is filtered and dried under vacuum (12 mm Hg) at room temperature (320 g).
La soluzione di lecitina (320 g) in cloroformio-metanolo 1 : 1 (650 mi) è cromatografata su AI2O3 (1,4 kg) eluendo con cloroformio-metanolo 1 : 1 (3,5 litri). L'eluato, concentrato a secco sotto vuoto (12 mm Hg) alla temperatura di 40°, è ripreso con cloruro di metilene (350 mi), centrifugato e versato a 5°C, sotto agitazione in acetone (2,4 litri). The lecithin solution (320 g) in 1: 1 chloroform-methanol (650 ml) is chromatographed on AI2O3 (1.4 kg) eluting with 1: 1 chloroform-methanol (3.5 liters). The eluate, concentrated dry under vacuum (12 mm Hg) at a temperature of 40 °, is taken up in methylene chloride (350 ml), centrifuged and poured at 5 ° C, under stirring in acetone (2.4 liters).
Dopo riposo per 60' a 5° si filtra e si essicca il precipitato sotto vuoto (12 mm Hg) a temperatura ambiente ottenendo 210 After resting for 60 'at 5 °, the precipitate is filtered and dried under vacuum (12 mm Hg) at room temperature obtaining 210
g di lecitina pura. (P alcalilabile: 3,70-3,80%; P della glicerilfosforilcolina: 3,70-3,80%). g of pure lecithin. (Alkalizable P: 3.70-3.80%; P of glycerylphosphorylcholine: 3.70-3.80%).
b) Preparazione della L-a-glicerilfosforilcolina 5 La soluzione di lecitina purificata (500 g circa; 0,7 moli) in metanolo anidro (1 litro) contenente 0,039 moli di metilato di sodio è lasciata a riposo, a temperatura ambiente, per 60'. Si separa lo strato oleoso formatosi, quindi si percola la soluzione su resina IR C 50 H + . (80 g altezza dello strato = cm 50) 10 eluendo successivamente con metanolo (1 litro). Si concentra l'eluato sotto vuoto, (12 mm Hg) a 40° ad un volume di circa 1 litro, quindi si diluisce con acqua (55o mi) e si estrae con cloroformio (4 x 400 mi). b) Preparation of L-a-glycerylphosphorylcholine 5 The purified lecithin solution (about 500 g; 0.7 moles) in anhydrous methanol (1 liter) containing 0.039 moles of sodium methylate is left to rest, at room temperature, for 60 '. The oily layer formed is separated, then the solution is percolated on IR C 50 H + resin. (80 g layer height = 50 cm) 10 eluting successively with methanol (1 liter). The eluate is concentrated under vacuum, (12 mm Hg) at 40 ° at a volume of about 1 liter, then diluted with water (55o ml) and extracted with chloroform (4 x 400 ml).
Si concentra a secco la fase acquosa sotto vuoto (12 mm 15 Hg) a 40°, si scioglie il residuo in acqua (410 mi), si aggiungono 12 g di carbonio decolorante e si agita a temperatura ambiente per 15'. Si filtra su millipore e si elimina il solvente a pressione ridotta (12 mm Hg) a 40°C, ottenendo 165 g di L-a-glicerilfosforilcolina pura amorfa. The aqueous phase is concentrated to dryness under vacuum (12 mm 15 Hg) at 40 °, the residue is dissolved in water (410 ml), 12 g of bleaching carbon are added and the mixture is stirred at room temperature for 15 '. The mixture is filtered on millipore and the solvent is removed under reduced pressure (12 mm Hg) at 40 ° C, obtaining 165 g of pure amorphous L-a-glycerylphosphorylcholine.
20 H20 = 15,5% 20 H20 = 15,5%
P totale = 84,1% Total P = 84.1%
Fosforo dopo separazione cromatografica: 83,94% Phosphorus after chromatographic separation: 83.94%
Glicole vicinale: 84,59% Local glycol: 84.59%
Colina: 85%. Choline: 85%.
25 Non si notano prodotti di scissione del legame P-O, come si può verificare effettuando l'analisi del campione secondo Broc-kerhoff [J. Lipids Research^, 96 (1963)] 0 secondo Dawson [Biochem. J._75. 45 (I960)]. Un campione, essiccato su P2O5 a 0,05 mm Hg, è stato cristallizzato da etanolo 99% - etere etili-30 co, ottenendo un solido cristallino, bianco, igroscopico. P.f.: 130-131°C; aD = —2,84 (10% in H20). 25 No splitting products of the P-O bond are noted, as can be verified by carrying out the analysis of the sample according to Broc-kerhoff [J. Lipids Research ^, 96 (1963)] 0 according to Dawson [Biochem. J._75. 45 (I960)]. A sample, dried on P2O5 at 0.05 mm Hg, was crystallized from 99% ethanol - ethyl ether-30 co, obtaining a crystalline, white, hygroscopic solid. M.p .: 130-131 ° C; aD = —2.84 (10% in H20).
C = 37,50; H = 7,70; N = 5,38; P = 12,06; colina estere = 46,92; glicole vicinale: 99,8%. C = 37.50; H = 7.70; N = 5.38; P = 12.06; choline ester = 46.92; neighborhood glycol: 99.8%.
Calcolato: Calculated:
35 C = 37,35; H = 7,84; N = 5,45; P = 12,04; colina estere = 47,12. 35 C = 37.35; H = 7.84; N = 5.45; P = 12.04; choline ester = 47.12.
Esempio 2 Example 2
Si parte da lecitina da soia (P alcalilabile: 2%; P della GPC 40 minimo: 0,5%). It starts from soy lecithin (alkalizable P: 2%; minimum GPC 40 P: 0.5%).
Una sospensione di questa lecitina (600 g) in metanolo (600 mi) è agitata a temperatura ambiente per 60'. Si lascia a riposo a 5° per 60' quindi si decanta il solvente lavando il residuo con metanolo (300 mi). A suspension of this lecithin (600 g) in methanol (600 ml) is stirred at room temperature for 60 '. The mixture is left to rest at 5 ° for 60 'then the solvent is decanted by washing the residue with methanol (300 ml).
45 L'estratto metanolico è lasciato a riposo a 5°C per 18 h, quindi è filtrato e concentrato a secco a pressione ridotta (12 mm Hg) a 40°C (g 70) (P alcalilabile: 3,0-3,1%); P della glicerilfosforilcolina: 2,0-2,1%). 45 The methanolic extract is left to rest at 5 ° C for 18 h, then it is filtered and concentrated to dryness under reduced pressure (12 mm Hg) at 40 ° C (g 70) (alkalizable P: 3.0-3, 1%); P of glycerylphosphorylcholine: 2.0-2.1%).
La successiva purificazione del residuo fornisce 23 g di leci-50 tina pura. The subsequent purification of the residue provides 23 g of pure leci-50 thine.
Si procede come descritto nell'esempio 1 per la lecitina da uovo. Si ottiene GPC di eguale purezza, con rese sostanzialmente identiche. Proceed as described in Example 1 for egg lecithin. GPC of equal purity is obtained, with substantially identical yields.
v v
Claims (3)
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| IT25761/79A IT1123187B (en) | 1979-09-17 | 1979-09-17 | PROCESS FOR THE PREPARATION OF L-ALPHA-GLYCERYLPHOSPHORYLCHOLINE |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CH642083A5 true CH642083A5 (en) | 1984-03-30 |
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ID=11217655
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CH90080A CH642083A5 (en) | 1979-09-17 | 1980-02-05 | PROCESS FOR THE PREPARATION OF L-ALPHA-GLYCERYLPHOSPHORYLCHOLINE. |
Country Status (9)
| Country | Link |
|---|---|
| JP (1) | JPS5643290A (en) |
| AR (1) | AR225159A1 (en) |
| CH (1) | CH642083A5 (en) |
| DE (1) | DE3000246C2 (en) |
| ES (1) | ES8100307A1 (en) |
| FR (1) | FR2464961A1 (en) |
| GB (1) | GB2058792A (en) |
| IT (1) | IT1123187B (en) |
| MX (1) | MX6294E (en) |
Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AT383130B (en) * | 1984-05-15 | 1987-05-25 | Chemie Linz Ag | METHOD FOR THE PRODUCTION OF PHOSPHATIDYLCHOLINES AND PHOSPHATIDYLETHANOLAMINES SUBSTITUTED DIFFERENTLY AT C1 AND C2 OVER THE NEW COMPOUNDS 1-0-TRITYLGLYCEROPHOSPHOCHOLIN OR RELATED (1-0, N-DITYHOHOLINE) |
| IT1201477B (en) * | 1985-10-04 | 1989-02-02 | Istituto Chemioterapico | PROCEDURE FOR THE PREPARATION OF L-ALPHA-GLYCERYLPHOSPHORYLCHOLINE, L-ALPHA-GLYCERYLPHOSPHORYLETHANOLOMINE AND L-ALPHA-GLYCERYLPHOSPHORYLINOSITOL FROM RAW AND / OR DEOLEATE LECITHINS |
| JPH066744B2 (en) * | 1986-05-15 | 1994-01-26 | 住友金属工業株式会社 | Method for manufacturing high Ni-Fe alloy hot rolled steel sheet |
| JP2572757B2 (en) * | 1986-11-27 | 1997-01-16 | 川崎製鉄株式会社 | Billet processing method and billet processing apparatus in hot rolling equipment |
| FR2614621B1 (en) * | 1987-04-29 | 1989-08-11 | Ire Celltarg Sa | PROCESS FOR THE PURIFICATION OF PHOSPHATIDYLCHOLINES AND PRODUCTS OBTAINED |
| CN103193821A (en) * | 2013-03-29 | 2013-07-10 | 山东罗欣药业股份有限公司 | Synthesis method of L-alpha-glyceryl phosphoryl choline |
| CN109134532A (en) * | 2018-07-19 | 2019-01-04 | 芜湖福民生物药业股份有限公司 | The method that glycerolphosphocholine is prepared using powdered soybean phospholipid |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US2864848A (en) * | 1954-07-19 | 1958-12-16 | Ca Nat Research Council | Method of producing l-alpha-glycerylphosphorylcholine |
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1979
- 1979-09-17 IT IT25761/79A patent/IT1123187B/en active Protection Beyond IP Right Term
- 1979-12-13 GB GB7942951A patent/GB2058792A/en not_active Withdrawn
- 1979-12-19 AR AR279371A patent/AR225159A1/en active
- 1979-12-28 JP JP17395979A patent/JPS5643290A/en active Granted
-
1980
- 1980-01-04 ES ES487473A patent/ES8100307A1/en not_active Expired
- 1980-01-05 DE DE3000246A patent/DE3000246C2/en not_active Expired
- 1980-01-18 FR FR8001090A patent/FR2464961A1/en active Granted
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Also Published As
| Publication number | Publication date |
|---|---|
| IT1123187B (en) | 1986-04-30 |
| FR2464961A1 (en) | 1981-03-20 |
| ES487473A0 (en) | 1980-11-01 |
| JPS5643290A (en) | 1981-04-21 |
| JPS6341920B2 (en) | 1988-08-19 |
| IT7925761A0 (en) | 1979-09-17 |
| ES8100307A1 (en) | 1980-11-01 |
| FR2464961B1 (en) | 1985-05-17 |
| DE3000246C2 (en) | 1989-02-02 |
| GB2058792A (en) | 1981-04-15 |
| DE3000246A1 (en) | 1981-04-02 |
| MX6294E (en) | 1985-03-18 |
| AR225159A1 (en) | 1982-02-26 |
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