| Purpose:This paper focused on chemical components and estab-lished the determination method of index component in QNM. Besides, we studied on anticerebral ischemia pharmacodynamics and pharmac-okinetics of PG, for the purpose of researching the mechanisms and material basis, which could provide research foundation for the development and utilization of local TCM resources.Methods and results:Established in vitro HPLC method, we select acetonitrile-0.1%formic acid with gradient elution and investigate several kinds of mobile phase, columns and wavelengths to detect40compounds from QNM for the first time. Besides, the main monomer compound,(+) pinoresinol-β-D-glucopyranoside (PG) has been researched methodo-logy of content determination and the content of PG in QNM has accounted for55.55mg/10ml average.Study on the mechanism of the protective effects of QNM against cerebral damage induced by suture method of focal ischemia reperfusion in rats. The model of focal brain ischemia reperfusion injury was esblished by middle cerebral artery occlusion (MCAO). The results showed that the rats have obvious behavioral problems and brain tissue necrosis after operation. Histopathology results of cerebral ischemia experiments indicated that QNM2.7ml/kg,5.4ml/kg,10.8ml/kg could reduce tissue lesions and neurons apoptosis of ischemic area compared with the model group, nimodipine as a positive control. The results of serum cytokines showed that QNM each dose group could significantly increased serum CAT, NO, SOD levels and reduced MDA level compared with positive control and model group (P<0.05, P<0.01). Compared with model group, QBM10.8ml/kg,5.4ml/kg could significantly reduce brain tissue ICAM-1level (P<0.05, P<0.01); QNM10.8ml/kg could significantly increased BDNF level (P<0.01); QNM10.8ml/kg,5.4ml/kg could significantly reduce IL-6level (P<0.05, P<0.01). TTC staining results showed that QNM each dose group could reduce the weight percentage of cerebral infarction in ischemic area, but compared with model group, the difference was not statistically significant. HE staining and immunohistochemistry results in cerebral ischemia repe-rfusion part showed that QNM2.7ml/kg,5.4ml/kg,10.8ml/kg group could significantly reduce bax expressions compared with model group (p<0.05, P<0.01), which might be related to inhibiting brain tissue lesions and the process of the neuronal apoptosis in ischemic area. Each dose group of QNM could significantly increase serum CAT level compared with the model group and nimodipine group (p<0.05, P<0.01); QNM5.4ml/kg could increase serum SOD (P<0.05, compared with the model group);QNM5.4ml/kg and10.8ml/kg could significantly lower serum MDA levels (P<0.05, compared with the model group). Compared with the model group, QNM10.8ml/kg could significantly increase BDNF, NGF levels and reduce ICAM-1, IL-6levels (P<0.05, P<0.01) and QNM5.4ml/kg could significantly increase NGF level compared with model group (P<0.01); TTC staining showed that all QNM groups could significantly reduce the weight infarct ratio of cerebral ischemic area compared with model group (P<0.05, P<0.01).PG pharmacokinetic research by tail intravenous administration in rats. Established in vitro HPLC method, selected acetonitrile-0.1%formic acid (25:75, v/v) for isocratic elution, the method is specific, and has no matrix interference. We selected10time points to collect rat plasma samples from orbital venous plexus. The acquisition time last for2hours. The analyte can be enriched in the collected plasma samples after protein precipitation and other managements. During the research we selected acetonitrile, methanol and ethyl acetate for protein precipitation; the final choice was acetonitrile which works best. Experimental data after model fitting with3P97, the results showed that pharmacokinetics of turpentine glycoside presented to be a double-compartment model after iv, the main pharmacokinetic parameters see in the following. A7.000μg/ml, α0.238min-l, B3.2351ng/ml, β0.01Omin-1, V(c)1.793(mg/kg)/(μg/ml), CL(s)0.051mg/kg/min/(μg/ml), tl/2β72.872min, K210.083min-1\K100.037min-1, K120.112min-1, AUC361.647(μg/ml)*min, T1/2α3.130min. The results showed that dynamic PG in vivo presented a rapid distribution and slow elimination in many tissues, which might play important roles in the clinical efficacy and possible pathway.Conclutions:In this paper, we have established HPLC methods for chemical composition analysis and quantified the index component named PG in QNM. The pharmacodynamic results of intervention MCAO induced cerebral ischemia and cerebral ischemia reperfusion model in rats show that QNM has a protective effect for ischemic injury. The mechanisms may be closely related to the inflammatory cytokines inhibition, free radicals elimination, neurotrophic factor level up-regulation, infarct volume reduction and neuronal apoptosis inhibition of QNM. In vito pharmacokinetic results of PG in rats showed that QNM may play an efficacy material basis role in protection of cerebral ischemia injury in brain tissue. |
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