| Objective:This experiment need to establish the method to detect the concentration of kaempferol in serum and tissue samples in rats first of all. Effects of kaempferol on the pharmacokinetics and Tissue accumulation were using RP-HPLC at different time.Methods:Selection of Wister rats, discuss metabolic regulation of kaempferol in rats. The HPLC with an analytical column Diamonsil C18(250mm×4.6mm5um) was used. Methanol-water65:35(v/v) was used as mobile phase at the flow rate of1.0mL·min-1. The column temperature was40℃and detection wavelength was at370nm. Treatment of rats biological specimens and spectrum of color behaviors:Suction rats vacant blood plasma100μl, add500μl acetonitrile precisely. Take the samples of hearts, livers, spleens, lungs and kidney of rats, weight respectively and precisely, add isotonic Na chloride and acetonitrile. An aliquot (20μl) was injected into the HPLC system after filter with Millipore filter. Results:1. The founded linearity of kaempferol was in the range of0.03to5.03μg·mL-1(R2=0.999). The experiment of precision:Suction the rats vacant blood plasma and the vacant homogenate, added0.05μg·mL-1,2.50μg·mL-1,5.00μg·mL-1kaempferol standard solutions100respectively, made the kaempferol concentration in plasma or in tissue homogenate be0.05μg·mL-1,2.5μg·mL-1,5.0μg·mL-1manipulation is similar to treatment of rats vacant blood plasma. Treated and determined for5times per day. and continued to extract and determine for3days, calculated the interassay precisions and day-to-day variabilities. The relative standard deviation (RSD) of inter-day and intra-day were all less than9.87%. The experiment of stability:manipulation is similar to treatment of rats vacant blood plasma, determine the relevant peak areas at0,2,4,6,8,10,12.24hour respectively. The relative standard deviation (RSD) of inter-day was less than8.53%. The experiment of recovery:Suction the rats vacant blood plasma and the vacant homogenate, added kaempferol standard solutions, manipulation is similar to treatment of rats vacant blood plasma. The average recovery rate of kaempferol was100.12%, with RSD of1.84%.2. Pharmacokinetics experiment design:The concentration-time data and pharmacokinetics parameters conformed to a2-compartment model. The pharmacokinetics after Vena caudalis intravenous injection such as a、β、T1/2a、T1/2β、K10、K21、K12、AUC、CL、V、V2、Cmax were 0.7357min-1、0.02883min-1、0.9421min、24.0446min、0.0465min-1、0.4561min-1、0.2620min-1、402.1261ug/mL-min、0.0230mL-min、0.4955mL、0.3846mL.18.7892μg/mL.3、The distribution of kaempferol was rapid and wide. At one minute after injection, all tissues had rather high concentration. The lung and liver were higher than the other tissues. The concentration of kaempferol eliminated conspicuously within60min, but in the liver and lung are still obvious content. Conclusion:1. In this research, we developed a HPLC method to determine the concentration of kaempferol in biological samples in rats. The analysis method offers high sensitivity, good repeatability, and specificity, which is suitable for the study of pharmacokinetics and distribution regularity of kaempferol in rats.2. Pharmacokinetic parameters, drug half-life is24.0446min, the clearance rate is0.0230mL·min, peak concentration of18.7892μg·mL-1. Kaempferol in rats in vivo pharmacokinetic process conform to basic characteristics of two compartment model.3. The distribution of kaempferol was wide and rapid in rats, there are no significant difference in the distribution between male and female rats. Heart, liver, spleen, lung, kidney and other major organizations are detected in kaempferol, but the elimination rate fast, the concentration of kaempferol eliminated conspicuously within60min. But drug still have in liver and lung tissue, to determine the initial kaempferol tissue metabolism in liver, lung tissue. |
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