High-throughput analysis of drug dissociation from serum proteins using affinity silica monoliths

A noncompetitive peak decay method was used with 1 mm× 4.6 mm id silica monoliths to measure the dissociation rate constants (k _d) for various drugs with human serum albumin (HSA) and α ₁-acid glycoprotein (AGP). Flow rates up to 9 mL/min were used in these experiments, resulting in analysis times of only 20-30 s. Using a silica monolith containing immobilized HSA, dissociation rate constants were measured for amitriptyline, carboplatin, cisplatin, chloramphenicol, nortriptyline, quinidine, and verapamil, giving values that ranged from 0.37 to 0.78 s ^-1. Similar work with an immobilized AGP silica monolith gave k _d values for amitriptyline, nortriptyline, and lidocaine of 0.39-0.73 s ^-1. These k _d values showed good agreement with values determined for drugs with similar structures and/or affinities for HSA or AGP. It was found that a k _d of up to roughly 0.80 s ^-1 could be measured by this approach. This information made it possible to obtain a better understanding of the advantages and possible limitations of the noncompetitive peak decay method and in the use of affinity silica monoliths for the high-throughput analysis of drug-protein dissociation.