Department of Pharmaceutical Analysis, China Pharmaceutical University, Key Laboratory of Drug Quality Control and Pharmacovigilance, Ministry of Education, Nanjing 210009, China.
Anal Biochem. 2012 Jan 15;420(2):171-6. doi: 10.1016/j.ab.2011.09.026. Epub 2011 Oct 3.
The measurement of membrane affinity is an important early screening step during drug discovery. However, classical methods for membrane affinity measurement are tedious and difficult to implement in high-throughput screening. This article describes a quantitative method for the measurement of membrane affinity by colorimetric assay based on polydiacetylene (PDA) sensors. Prepared lipid/PDA chromatic vesicles were used to model cell membranes. By measuring the colorimetric response of the chromatic vesicles when drug-membrane interactions occurred, membrane affinity constant K(b) could be calculated using a simple quantitative model. Under optimized preparation conditions, the calculated log(K(b)) values exhibited an in-batch relative standard deviation (RSD) of less than 4% and a between-batch RSD of less than 8% for all three reference compounds. The logarithm of K(b) of the six β-blockers exhibited excellent linear correlation with the logarithm of the liposome/water partition coefficient (K(m)) with R(2)=0.9793. For neutral compounds, the log(K(b)) of n-fatty alcohols correlated with the logarithm of the n-octanol/water partition coefficient (K(oct)) with a linear correlation coefficient R(2)=0.9833. This work provides a simple, convenient, and reproducible method for the rapid measurement of membrane affinity and presents important implications for high-throughput screening.
膜亲和力的测量是药物发现过程中一个重要的早期筛选步骤。然而，经典的膜亲和力测量方法繁琐且难以在高通量筛选中实施。本文描述了一种基于聚二乙炔（PDA）传感器的比色测定法来测量膜亲和力的定量方法。制备的脂质/PDA 彩色囊泡被用来模拟细胞膜。通过测量药物-膜相互作用时彩色囊泡的比色响应，可以使用简单的定量模型计算膜亲和力常数 K(b)。在优化的制备条件下，对于所有三种参考化合物，计算得到的 log(K(b))值在批内相对标准偏差（RSD）小于 4%，批间 RSD 小于 8%。六种β-受体阻滞剂的 K(b)的对数与脂质体/水分配系数（K(m)）的对数呈极好的线性相关，相关系数 R(2)=0.9793。对于中性化合物，n-脂肪醇的 K(b)的对数与正辛醇/水分配系数（K(oct)）的对数呈线性相关，相关系数 R(2)=0.9833。这项工作为快速测量膜亲和力提供了一种简单、方便和可重复的方法，并为高通量筛选提供了重要的启示。
