Résumé
@#Objective - To optimize the extraction and quantification methods for the determination of S phenylmercapturic acid - Methods (SPMA) in urine with performance liquid chromatography mass spectrometry. The urine was hydrolyzed with 50.0% sulfuric acid. The hydrolysate was purified by solid phase extraction column. Purified samples were separated by C18 chromatographic column and detected by tandem mass spectrometry. The isotope labeled SPMA was used as the internal Results - standard. The internal standard curve was used for quantification. The linear range of SPMA was 0.50 50.00 μg/L with the correlation coefficient of 0.999 8. The detection limit and the lower limit of quantification were 0.05 and 0.17 μg/L, - - - - respectively. The recovery rate was 97.0% 102.0%. The within run and between run relative standard deviation were 0.6% 1.0% - and 1.7% 6.5%, respectively. The mass concentration of urinary SPMA in the occupational benzene exposure group was - vs P higher than the non occupational benzene exposure group by this method (median: 2.81 0.28 μg/g creatinine, <0.05). Conclusion Compared to the national standard method, this optimized method of solid phase extraction and internal standard for quantification eliminates the matrix effect. This method is accurate and precise, and is suitable for the determination of SPMA acid in urine.
Résumé
Objective To study the effect of Phenobarbital (PB) on experimental rats,observe the histological changes of immature brain and the expressions of apoptosis-related proteins Bcl-2 and Bax in neurons detected by immunohistochemistry,and to explore the influence and mechanism of PB on brain damage at therapeutic levels to immature brain maturation of rat in order to provide the theoretical and experimental base for clinic.Methods A total of 40 healthy 18-day-old Sprague-Dawley(SD) rats (male or female) were randomly assigned into 2 groups:normal saline (NS) treated as control group(20 cases),PB group(20 cases).Each group was further randomly divided into longterm(4 weeks) treated group and short-term(2 weeks) treated group(10 rats in each group).The rats in PB group received intragastricadministration with PB (30 mg/kg).The rats in control group were handled by injection of NS into stomach and abdomen according to 4 mL/kg.All performances were undertaken for twice every day.At the end of the therapeutic period,body and brain weight were measured when the rats were sacrificed.Histological studies on the tissues of frontal lobes and hippocampus were performed by Hematoxylin-Eosin(HE) staining and Nissl staining.Expressions of apoptosis-related proteins Bcl-2 and Bax in neurons were detected by immunohistochemistry.Results There were no significant differences in body and brain weights or histological studies index among control group as well as PB group before and after treatment for short term(P ＞0.05).Remarkable reduction of brain weight was only observed in immature rats exposed to PB compared to control group for long period,and significant neurodegeneration,neuronal necrosis were observed in immature rats exposed to PB compared to control group(all P ＜ 0.01).The expression of Bax protein in the frontal lobe increased significantly in immature rats receiving PB for long period comparing with control (P＜0.01).In contrast,expression of Bcl-2 protein did not change at therapeutic level.The ratio of Bax/Bcl-2 was obviously increased.Conclusions Chronic treatment with PB will result in significant neuronal apoptosis and necrosis and persistent cognitive impairment and brain damage to immature rates.Brain damage of PB at therapeutic level to immature brain may be irreversible.The significant expression of Bax protein in the frontal lobe and the high rate of Bax/Bcl-2 are probably the main reasons which cause brain weight decreasing and brain damage by PB in immature brain tissue.