Internal Standard Capillary Electrophoresis as a High-Throughput Method for pK_aDetermination in Drug Discovery and Development

Abstract

A novel high-throughput method for determining acidity constants (pKa) by capillary electrophoresis (CE) is developed. The method, based on the use of an internal standard (IS-CE), is implemented as a routine method for accurate experimental pKa determination of drugs undergoing physicochemical measurements in drug discovery laboratories. Just two electropherograms at 2 different pH values are needed to calculate an acidity constant. Several ISs can be used in the same buffer and run to enhance precision. With 3 ISs, for example, the pKa of the test compound (TC) can be obtained in triplicate in less than 3 min of electrophoresis. It has been demonstrated that the IS-CE method eliminates some systematic errors, maintaining, or even increasing the precision of the results compared with other methods. Furthermore, pH buffer instability during electrophoretic runs is not a problem in the IS-CE method. It is also proved that after 16 h of electroseparation using the same buffer vial, pH may change by around one unit; but the pKa calculated by the IS-CE method remains constant. Thus, IS-CE is a powerful high-throughput method for pKa determination in drug discovery and development.

Assessing Physicochemical Properties of Drug Molecules via Microsolvation Measurements with Differential Mobility Spectrometry

“…Unfortunately, existing methodologies lack the resolution to discriminate between closely related structural analogues or motifs. 15 , 16 Moreover, while progress has been made in streamlining these assays, 17 − 19 they still require considerable resources and experience suboptimal reproducibility. 20 , 21 …”

Determining molecular properties with differential mobility spectrometry and machine learning

“…However, it is often challenging to discriminate between closely related structural analogues using the current state-of-the-art techniques 20 , 21 . Moreover, while exceptional progress has been made in refining these methods 22 – 24 , they still require considerable resources and often experience issues with reproducibility 25 , 26 . As a result, efforts to develop complementary techniques for assaying molecular properties are ongoing.…”

Determination of acidity constants, ionic mobilities, and hydrodynamic radii of carborane‐based inhibitors of carbonic anhydrases by capillary electrophoresis

“…CE is a powerful separation method suitable for the physicochemical characterization of ionogenic compounds [20] including determination of their acidity constant, p K a [21, 22]. CE is widely used for the determination of the p K a s of acidic, basic, and amphoteric compounds in aqueous [23–33], hydro‐organic [34], and organic solvents [35, 36]. Recently, a new computer program AnglerFish was introduced for direct determination of the thermodynamic p K a s and limiting ionic mobilities from the raw CE experimental data [37].…”

“…The analytes need not be quite pure if their admixtures can be separated during the CE run, several analytes can be studied simultaneously, the method is relatively simple, can be automated and is cost effective and environment friendly. The disadvantage of time‐consuming CE analyses at low pH because of the suppressed EOF can be solved by accelerating the experiments using pressure‐assisted CE [26, 39], or by using internal standards with similar known p K a values [24, 25], or by dynamic coating of capillary wall [40]. Acid–base and other physicochemical properties of various sulfonamides were studied by CE [40–43].…”