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[摘要]
目的 研究染料木黄酮在Caco-2单细胞层模型上的吸收特征。方法 将染料木黄酮的细胞样品经高速离心后取上清液,以乙腈-醋酸铵水溶液为流动相,以睾丸酮为内标,采用超高效液相色谱(ultra performance liquid chromatography,UPLC)法检测。考察时间、pH值等因素对染料木黄酮吸收的影响,测定透过Caco-2单细胞层的染料木黄酮浓度并计算其表观渗透系数(Papp)。结果 Caco-2细胞对染料木黄酮的吸收在1 h内呈线性,药物摄取时间定为1 h;pH值对染料木黄酮的吸收没有显著影响。染料木黄酮的线性范围为0.625~40 μmol/L,日间、日内精密度均小于15%。Papp从基顶侧(apical,AP)到基底侧(basolateral,BL)为1.51×10?5 cm/s,从BL侧到AP侧为1.84×10?5 cm/s,其渗透系数Papp(BL-AP)/Papp(AP-BL)为1.22。结论 染料木黄酮在肠道的吸收主要以被动扩散为主,UPLC检测方法简单、灵敏度高,可用于研究染料木黄酮在Caco-2单细胞层模型上的吸收特征。
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[Abstract]
Objective To study the absorptive characteristics of genistein in Caco-2 cell monolayer model. Methods The genistein cell samples underwent high speed centrifugation, then the supernatant was collected and detected by UPLC method with acetonitrile and ammonium acetate water solution as mobile phase and testosterone as internal standard. The effects of time and pH value on genistein were investigated. The concentration of genistein across the Caco-2 cell monolayer model was determined and the apparent permeability coefficient (Papp) of genistein was calculated. Results The uptake of genistein in Caco-2 cells showed a linear range in 1 h and the uptake time was fixed at 1 h. pH value showed no significant influence on the absorption of genistein. The linear range of genistein was 0.625—40 μmol/L, with intra- and inter-day precision less than 15%. Papp (AP-BL) from apical side (AP) to basolateral side (BL) was 1.51 × 10?5 cm/s, whereas Papp (BL-AP) from BL to AP was 1.84 × 10?5 cm/s, and the efflux ratio of Papp (BL-AP)/ Papp (AP-BL) was 1.22. Conclusion Passive diffusion is the major process for the transport of genistsein across the intestine. The UPLC method is simple and sensitive, and it is applicable for the study on absorptive characteristics of genistein in Caco-2 cell monolayer model.
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