Hboa And Its Derivatives, Synthesis And Hepatoprotective Effect

Objective: To synthesize Ilicifoliosids alkaloid A and its derivatives, and to study their hepatoprotective effects and their related mechanisms. Methods: HBOA(TC-1) was obtained from 2-nitroresorcinol with reduction and subsequent condensation with urea by"one-pot-way", and acyl and benzylic derivatives(TC-2, TC-3, TC-4)were prepared through substitution reaction.HSC-T6 was incubated with different concentrations of TC-1, TC-2, TC-3, TC-4 (5, 1, 0.2, 0.04, 0.008mg/ml respectively) or without TC-1, TC-2, TC-3, TC-4 for 32 hours, respectively. The proliferation rate of the HSC-T6 was determined by MTT assay. The morphological changes of the HSC-T6 were observed with inverted fluorenscence microscopy.The acute toxicity was tested in mice by intragastric administration and median lethal dose (LD50) as well as 95% confidence interval was caculated by Bliss method.The acute hepatic injury models were established by carbon tetrachloride with intraperitoneal injection in mice. The activities of alanine transferase(ALT)in serum were investigated by chlorimetric method, pathological section of tissues were prepared and observed. Results: If 2-nitroresorcinol was the starting material, the yeild rates of TC-1, TC-2, TC-3, TC-4 were 60.08%, 68.22%, 64.62%, 74.91% respectively. The target compounds were identified by IR, 1H-NMR and 13C-NMR wave analysis, and the wave information was quite matched.MTT assay indicated that the proliferation rates of HSC-T6 among different dose groups were significantly reduced compared with the control group (all P < 0.05 or P < 0.01), except the TC-1, TC-2, TC-3 groups whose concentrations were 0.008mg/ml and the TC-4 group (P>0.05). TC-1, TC-2, TC-3 significantly inhibited the proliferation of HSC-T6 in a dose-dependent manner. The higher the concentrations, stronger the inhibitory effects were. After acridine orange staining, inverted fluorenscence microscopy showed the morphological changes of HSC-T6, including depletion in numbers, cells shrinkage, karyorrhexis and karyopycnosis, and the changes were more significant as the concentration increased.After intragastric administration, partial mice showed toxic reaction and death, the LD50 of TC-1, TC-2, TC-4 were 2.14,1.95,1.64g/kg. 95% confidence intervals were 1.89～2.33, 1.70～2.25, 1.41～1.85g/kg respectively. The maximum tolerated dose (MTD) of TC-3 was 2.34g/kg. The autopsy was carried out on the mice after their death and the pathology was not found in the primary organs of the mice.CCl4-induced acute liver injury model in mice was established successfully, compared with NC group, the serum ALT in model group increased a lot (P<0.01) and the pathological changes were obvious (P<0.01); compared with CCl4 group, the serum ALT and pathology in TC-1H, TC-2H, TC-3H , TC-1M, TC-2M, TC-3M groups(P<0.01) changed much more than TC-1L,TC-2L,TC-3L,TC-4H groups(P>0.05).Conclusion: The target compounds are synthesized successfully, inhibitory effects on cell proliferation of TC-1, TC-2, TC-3 really exist. TC-1, TC-2, TC-3, TC-4 have low toxicity with oral administration, and they are safe for clinical application. TC-1, TC-2, TC-3 have a good hepatoprotective effect for liver injury.