A System for LogD Screening of 96-Well Plates Using a Water-Plug Aspiration/Injection Method Combined with High-Performance Liquid Chromatography−Mass Spectrometry

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Abstract

A system for screening of the octanol/water distribution coefficient (LogD) using automatic sampling of 96-well plates was developed. The high-speed assay for LogD (HSLogD) screening uses a water-plug aspiration/injection method combined with high-performance liquid chromatography-mass spectrometry (HPLC-MS). The method is useful for LogD analysis of highly hydrophobic compounds, where the concentration of the compound in the octanol phase is much higher than that in the water phase. In the case of LogD analyses, the conventional shake-flask method has been widely used, but it is difficult to increase the throughput of the shake-flask method because the lower water phase is carefully separated by manual separation without contamination of the upper octanol phase. We attempted to develop an automatic sampling method instead of manual separation to increase the throughput of the measurement. In initial attempts at automatic sampling, contamination of the octanol phase occurred when sampling of the water phase was made. This was because the octanol phase entered the sampling needle as it passed through to the lower water phase. This contamination was prevented by taking up a few microliters of water into the needle as a plug before sampling of the water phase (the water-plug aspiration/injection method). LogD values of some common drugs measured using the HSLogD agreed with reported LogD values (0 < LogD < 5).

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