Study On The Lipase-catalyzed Resolution Of Tetrahydro-1H-benzoimidazole-5-carboxylate And Its Applications

With the continuous development of modern medicine, people are increasingly concerned about drug safety. Chiral drugs in clinical medicine has been widely used, its pharmacological effects in vivo are achieved through the strict match between chiral macromolecules and the molecular recognition. That is to say, there exist specific interactions between drugs and receptor. Only one enantiomer of chiral drugs is suitable for the combination with the active site or receptor. Typically, one enantiomer combined with the active site or receptor selectively and work against the disease. The other enantiomer are ofen noneffective, some even have side effect. Therefore, a great deal of emphasis was devoted to the development and application of chiral drugs.High level enantioselectivity is one of the important catalytic properties of lipase, the establishment and development of the non-aqueous enzymology increased the importance of lipase in organic synthesis.The application of lipase in non-aqueous system facilitated the preparation of optically pure compounds and play a vital role in the synthesis of drugs, pesticide and flavors.In this paper, lipase was chose as the resolution catalyst, and the optically pure (R)-4, 5, 6, 7-tetrahydro-benzimidazole-5-carboxylic acid was obtained through the stereoselective hydrolysis of the ester. Reaction conditions were optimized in ordet to achieve satisfying stereoselectivity. We also explained the mechanism of the enantioselective hydrolysis under molecular level and studied application of tetrahydro-1H-benzimidazole -5-carboxylic acid. 1. The synthesis of methyl 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylate. In this paper, the reaction conditions of cyclization, esterification, and reduction were investigated in the synthesis of methyl 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylate.Reaction time of cyclization was 6 hours, the more suitable ratio of 3,4-Diaminobenzoic acid and formic acid was 1:40. The suitable ph value in after-treatment was 4.The esterification was performed in two steps. First, the acyl chloride was prepared in 4 hours. Then, the reaction of acyl chloride and methanol was performed in 3h. The more suitable ratio of benzimidazole-5-carboxylic acid, thionyl chloride and methanol was 1:7:40. Addition of cosolvent increased the yield of the product.Methyl 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylate was obtained by the hydrogenation of methyl 4,5,6,7-benzimidazole-5-carboxylate sulfate. Reaction conditions such as ratio of reactants , hydrogen pressure, reaction temperature and reaction time were examined. The optimal reaction conditions: 20 mol% Pd-C, 230oC. 6 Mpa, 21h.2. Analysis of the 4,5,6,7-tetrahydro benzimidazole -5-carboxylic acid ester. In this paper, the optical purity of the 4,5,6,7-tetrahydro benzimidazole-5-carboxylic acid ester was determined by chiral HPLC analysis with CHIRALPAK AS-H, CHIRALPAK AD-H and CHIRALPAK OD-H column. Hexane, isopropanol and ethanol were used as mobile phase. CHIRALPAK OD-H column was found to be the more suitable column, the optimal mobile phase was hexanone: ethanol = 97:3. Flow rate is 1.0 ml/min and the wave lenth is 254 nm.3. Study on the lipase catalysed resolution of methyl 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylate. The synthesis of (R)-4,5,6,7-tetrahydro-benzimidazole-5-carboxylic acid via the lipased catalysed resolution of methyl 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylate under mild reaction condition was investigated. Different lipases were screened in the stereoselective hydrolyzation of the model compound. PSL exhibited excellent catalytic activity, high enantioselectivity was obtained.Then, we examined the influences of reaction conditon on the resolution reaction. Experimental data showed that the catalytic activitity of lipase was influenced dramatically by reaction temperature. High enantioselectivity was obtained under lower reaction temperature. Increasing of the reaction temperature resulted in the decrease of enantioselectivity. This may be caused by the denaturalization of lipase under higher reaction temperature. The more suitable Ph value of reaction medium (buffer) was 8. The catalytic activitity and stereoselectivity of PSL was also influenced by the length of alkane chain of esters. From the kinetic curves of enzyme catalysed resolution we can came to the conclusion that PSL showed no interfacial activation and substrate inhibition and the resolution was completely controlled by the dynamics.The method based on the resolution of methyl 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylate was applied to the stereoselective hydrolysis of other alkyl esters, good conversions and enantioselectivitied were obtained. This indicated that the new method established by us was universal for the chiral resolution of 4,5,6,7-tetrahydro-1H-benzoimidazole-5-carboxylates.4. The application of (R)-4,5,6,7-tetrahydro-benzimidazole-5-carboxylic acid. Chiral intermediate (R)-4, 5, 6, 7-tetrahydro-benzimidazole-5-carboxylic acid was prepared through the lipase catalysed kinetic resolution of Methyl 4, 5, 6, 7-tetrahydro-benzimidazole-5-carboxylate. Then, we afforded opitically pure ramosetron hydrochloride after two steps of reaction,the yield is 24.6%.