双酚A和对-壬基酚雌激素受体的竞争性结合

Estrogen receptor competitive binding assay of bisphenol A and para-nonylphenol

目的 用雌激素受体竞争性结合试验检测双酚A(BPA)、对 -壬基酚 (p -NP)的雌激素样活性。 方法 将SD大鼠进行人工卵巢去除术 ,饥饿受体 14d后提取子宫胞浆雌激素受体 (ER) ,受体蛋白定量 ,作雌二醇 (E2 )非标记配体饱和分析 ,以确定E2 的IC50 。随后进行BPA和 p -NP的竞争性取代实验 ,观察BPA ,p -NP是否能与3 H -E2竞争性结合大鼠子宫胞浆ER。结果 E2 的IC50 是 1× 10 -9mol/L ,BPA、p -NP的IC50 分别为 2 0 8× 10 -5mol/L和2 6 6R10 -5mol/L ,BPA、p -NP的相对结合亲和力系数 (RBA)分别为 0 0 0 4 8和 0 0 0 38,BPA与ER的结合能力稍高于 p -NP。 结论 BPA和 p

Objective The estrogen-like activity of bisphenol A and p-nonylphenol was examined with the estrogen receptor competitive binding assay.Methods Adult healthy female SD rat were ovarietomized and sacrificed 2 weeks later.Cytosol estrogen receptor were abstracted from the uteri.Aliquots of 150?μg cytosol protein were added with 1?μCi/mol 3H-E 2 50?μl and increasing concentrations of unlabeled E 2,BPA and p-NP,the total volume of reponse was 200?μl.The mixtures were incubated at 4?℃ 16-18 hours.DCC methods removed free labeled-ligand.At last,the radioactivity was measured.Results The percent inhibitory values (IC 50 ) of E 2 was 1×10 -9 mol/L.BPA and p-NP can inhibit of 3H-E 2 binding to rat uterine cycotol estrogen receptor with an IC 50 of 2.08×10 -5 mol/L and 2.66×10 -5 mol/L,respectively.The relative binding affinity(RBA) of BPA and p-NP were 0.004?8 and 0.003?8,respectively.The binding capacity of BPA to ER was bigger than that of p-NP.Conclusion Binding to ER maybe was the mechanism that BPA and p-NP mimicked estrogen-like activity.