Separation of negatively charged isomeric quinones in acidic solution by capillary electrophoresis with reductive electrochemical detection

Abstract

A capillary electrophoresis method with reductive electrochemical detection was developed for the separation of the novel enzyme cofactor pyrroloquinoline quinone (PQQ) and three isomeric analogues. Tuning the reduction potential of the o-quinone moiety to a value more positive than the reduction potential for oxygen was accomplished by adjusting the capillary buffer to pH 2, thus eliminating the need for deoxygenation. To counter the suppression of the electroosmotic flow (EOF) at pH 2, a negative separation voltage of –22.5 V was applied to a 25 µm id capillary resulting in migration of the anionic isomers toward the electrochemical detector. Fast and efficient separation was achieved in 0.15 mM phosphate buffer at pH 2. A mass detection limit for PQQ of 2 fmol was obtained with end-column detection. This approach may find utility for the separation and sensitive detection of a wide range of reducible quinones.