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采用高速逆流色谱法分离纯化石榴皮乙醇粗提物中的有效组分,在分析型高速逆流色谱上对分离参数进行了系统优化,并放大至制备型高速逆流色谱。从5种溶剂体系中,确定了正丁醇-甲醇-水(55:1:45,v/v)为最佳两相溶剂体系。在流动相流速2.0 m L/min,主机转速850 r/min,进样量150 mg条件下进行分离制备,获得3个峰(组分I、II、III)。通过微量热法测定了不同浓度的组分I、II抑制类志贺邻单胞菌Sh1生长代谢的热功率-时间曲线。利用软件Origin8.0,根据Logistic方程拟合出各浓度下细菌的生长速率常数k,并建立了生长速率常数与药物浓度的关系,进而计算出两组分的半数抑菌浓度分别为2.16、2.68mg/m L,以期为进一步研制石榴皮抗菌剂及临床应用提供科学理论依据。
The high performance countercurrent chromatography (HPLC) was used to separate and purify the ethanol extract from pomegranate peel. The separation parameters were systematically optimized by analytical high-speed countercurrent chromatography and amplified to preparative high-speed countercurrent chromatography. From the five solvent systems, n-butanol-methanol-water (55: 1: 45, v / v) was the best two-phase solvent system. The mobile phases were separated and prepared at a flow rate of 2.0 m L / min, a host rotation speed of 850 r / min and an injection volume of 150 mg. Three peaks (fractions I, II and III) were obtained. The thermodynamic power-time curves of different concentrations of components I and II for growth inhibition of Shigellobacterium shigellosis Sh1 were determined by microcalorimetry. Using software Origin8.0, according to Logistic equation, the growth rate constant k of bacteria at each concentration was fitted and the relationship between growth rate constant and drug concentration was established. Then the half inhibitory concentrations of the two components were calculated as 2.16 and 2.68 mg / m L, in order to provide a scientific basis for the further development of pomegranate bark antibacterial agents and clinical application.