Study On The Synthesis Process Of Optically Pure(6S)-5-methyltetrahydrofolate And Its Amino Acid Ester Salt Derivatives

Tetrahydrofolate derivative(6S)-5-methyltetrahydrofolate,is a new type of folate drugs,can be directly transformed and used by human body.Meanwhile,compared with other drugs,(6S)-5-methyltetrahydrofolate has the characteristics of conspicuous therapeutic effects,complete functions and tiny secondary effect.But currently,there are no relevant domestic manufacturers of(6S)-5-methyltetrahydrofolate in China,the supply of products is all dependent on imports,at the same time,Folic acid,the raw material of tetramethylfolic acid,has a large output in China.Therefore,the study of preparing(6S)-5-methyltetrahydrofolate with folic acid as the raw material has important practical significance and broad market prospects.Although(6S)-5-methyltetrahydrofolate has many advantages,it is unstable,and it needs to be converted into the corresponding salt to increase its stability.At present,the common(6S)-5-methyltetrahydrofolate drugs mainly exist in the form of calcium salt or glucosamine salt,but excessive intake of calcium ions or glucose will bring some potential danger to the body.Amino acids,as the constituent compounds of organisms,have the best compatibility with organisms.Therefore,there are few side effects of using amino acids which are necessary and cannot be synthesized by human body as the ligand of(6S)-5-methyltetrahydrofolate.Consequently,the selection of amino acids as ligands of(6S)-5-methyltetrahydrofolate is an important research direction for the future upgrading of such products.This paper studies the preparation of(6S)-5-methyltetrahydrofolate and its amino acid ester salt with folic acid as the raw material.The details are as follows:1.Synthesis of(6S)-tetrahydrofolate: folic acid was reduced by sodium borohydride to obtain tetrahydrofolate with a yield of 97.0%.p-Toluenesulfonic acid was used as the resolving agent.Using the difference in solubility of tetrahydrofolate p-toluenesulfonate of S configuration and R configuration in water,optically pure(6S)-tetrahydrofolate p-toluenesulfonate was obtained.Then under alkaline conditions,(6S)-tetrahydrofolate was obtained.The optimal reaction conditions were determined as follows: the ratio of tetrahydrofolate and p-toluenesulfonic acid was1:1.3,water was used as the solvent,and the reaction continued at 45°C for 7h.After two times of recrystallization,the light yellow(6S)-tetrahydrofolate was finally obtained.The yield was 45.3%.2.Synthesis of(6S)-5-methyltetrahydrofolate:(6S)-tetrahydrofolate underwent formylation reaction with formaldehyde under alkaline conditions and was reduced by sodium borohydride to obtain(6S)-5-methyltetrahydrofolate.By optimizing the reaction route and comparing it with the reaction route of N-octyl-D-glucosamine as the resolving agent to prepare optically pure(6S)-5-methyltetrahydrofolate,it was determined that the reaction route for preparing(6S)-5-methyltetrahydrofolate with folic acid as the raw material and p-toluenesulfonic acid as the resolving agent was the optimal route,and the optimal reaction conditions were: water was used as solvent,the ratio of(6S)-tetrahydrofolate,formaldehyde andboron sodium hydride was 1:9:3,and formaldehyde was added dropwise at 0-5°C and the reaction was maintained for25 min.After sodium borohydride was added,the reaction was maintained at 60°C for1 h.Adjust the p H of the reaction system with glacial acetic acid could obtain(6S)-5-methyltetrahydrofolate.The yield was 91.5% and the purity was 97.6%.3.Recovery and utilization of(6R)-tetrahydrofolate p-toluenesulfonate.In this article,the p H of the by-product(6R)-tetrahydrofolate-p-toluenesulfonate solution was adjusted to 3-4 and filtered with suction to obtain free(6R)-tetrahydrofolate,In the environment of 0°C and p H=7,the oxidation reaction was carried out with oxygen as the oxidant to obtain the product then folic acid was recovered with the yield of 88.5%.4.Synthesis of amino acid ethyl ester.The eight amino acids which were necessary for humans were used to prepare thirteen corresponding amino acid methyl esters,ethyl esters,and n-butyl esters by neutralization and esterification reactions,and their yields were between 47.2% and 81.5%.5.Synthesis of(6S)-5-methyltetrahydrofolate amino acid ester.5g of(6S)-5-Methyltetrahydrofolate was dispersed in water and reacted with free amino acid ester in the mixed solvent of water,acetonitrile and ether,to obtain the thirteen corresponding(6S)-5-methyltetrahydrofolate amino acid ester with the yield of70.6%-96% and the purity of 97.2%-98.2%.HPLC method was established in this paper to monitor the reaction process and the optical purity of the products.The products and intermediates were characterized by NMR,HRMS and FT-IR.