INVESTIGATION ON TOXICITIES OF ALTERNATIVES FOR CHLOROFLUOROCARBONS AND ITS RISK ASSESSMENT USING MOLECULAR AND BIOLOGICAL MARKERS.

取1-溴丙烷染毒大鼠的尿液。将100μL尿加入400μL饱和氯化钠溶液中，用500μL乙酸乙酯提取3次。收集的乙酸乙酯层用干燥的亚硫酸钠脱水，然后用离心式蒸发器干燥。干燥样品用40μ的乙酸乙酯L溶液溶解，用四叔丁基二硫代谢物衍生化。结果表明，代谢产物N-乙酰-S-丙基半胱氨酸具有剂量依赖性，可作为内暴露标志物。用丙酮-草酸混合液从血样中提取珠蛋白。在氮气中酸解球蛋白，用LC-MS/MS鉴定S-丙基半胱氨酸。将组织匀浆，得到可溶性组分。用夹心式酶免疫分析法检测神经元特异性蛋白γ-烯醇化酶和胶质细胞特异性蛋白β-S100。谷胱甘肽(GSH)和谷胱甘肽(GSSG)测定采用酶循环法。结果表明，大脑中的伽马烯醇化酶特异性降低，1-溴丙烷对中枢神经系统有较大的不良影响。α-enolase免疫组织化学染色显示α-enoalse对睾丸支持细胞有明显的特异性定位。酶免疫定量显示睾丸α-烯醇化酶呈剂量依赖性增加，提示1-溴丙烷对支持细胞的作用。

Urine was collected from the rats exposed to 1-bromopropane. One hundred μL of urine was added 400μL of saturated sodium chloride solution, and then extracted with 500μL of ethyl acetate for three times. The collected ethyl acetate layer was dehydrated with dried sodium sulfoxide, and dried with centrifuging evaporator. The dried sample was resolved with 40μL of ethylacetate and derivatized with TBDMS. The result showed the metabolite N-acetyl-S-propylcysteine was dose-dependent and available as an internal exposure marker.Blood was collected from the rats exposed to 1-bromopropane. Globin was extracted from blood samples using acetone-oxalic acid mixture. Globin was hydrolyzed with acid under nitrogen, and S-propylcysteine was identified using LC-MS/MS.The cerebrum, cerebellum, brainstem and spinal cord were taken from the rats exposed to 1-bromopropane under deep anesthesia. The tissue was homogenized and the soluble fraction was obtained. Neuro-specific protein γ-enolase and glia-specific protein β-S100 was quantified with a sandwich type enzyme immunoassay. Glutatione (GSH and GSSG) was also measured by enzymatic cycling method. The result showed that gamma-enolase was decreased specifically in the cerebrum and 1-bromopropane has a great adverse effect on the central nervous system.The testes were taken from the rats exposed to 1-bromopropane under deep anesthesia. Immunostaining with α-enolase showed clearly specific localization of α-enoalse to Sertoli cells in the testis. The quantification with enzyme immunoassay showed dose-dependent increase in alpha-enolase in the testis, suggesting the effects of 1-bromopropane on Sertoli cells.