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目的:建立肝水解肽体外生物学活性测定方法。方法:将不同浓度肝水解肽作用于正常人肝L02细胞,WST-8比色法测定L02细胞增殖情况。同时对实验条件,包括稀释液、细胞接种密度、药物作用时间等进行探讨,建立肝水解肽体外活性测定方法,并用该方法对2个厂家生产的2批产品进行活性检测。结果:获得了比较稳定可靠的实验参数:稀释液为RPMI-1640培养液,细胞接种密度为4.0×104个.mL-1,药物作用时间为48~72 h,肝水解肽浓度为0.5,0.25,0.12,0.06,0.03,0.015 mg.mL-1。由这些实验参数建立了肝水解肽活性测定方法。用该方法检测的2批产品活性均合格,重复实验3次,每次实验结果均一致,RSD均低于10%。结论:该方法可用于肝水解肽体外生物学活性测定。
Objective: To establish a method for the determination of biological activity of hepatic hydrolytic peptide in vitro. Methods: Different concentrations of hepatic hydrolyzing peptide were applied to normal human L02 cells. The proliferation of L02 cells was determined by WST-8 colorimetric assay. At the same time, the experimental conditions, including dilution, cell seeding density, drug action time and so on were discussed to establish a method for the determination of in vitro activity of hepatic hydrolytic peptide. Two batches of products produced by two manufacturers were tested by this method. Results: The stable and reliable experimental parameters were obtained: the dilution was RPMI-1640 medium, the cell inoculation density was 4.0 × 104 cells.mL-1, the drug action time was 48-72 h, the concentration of hepatic hydrolyzate peptide was 0.5, 0.25 , 0.12, 0.06, 0.03, 0.015 mg.mL-1. From these experimental parameters, a method for the determination of hepatic hydrolytic peptide activity was established. The activity of the two batches of products tested by the method was qualified, and the experiment was repeated three times. The results of each experiment were consistent and the RSDs were less than 10%. Conclusion: This method can be used to determine the biological activity of hepatic hydrolytic peptide in vitro.