Determination of 8-chloroadenosine 3',5'-monophosphate in dog plasma by capillary electrophoresis.

Abstract

A method for the quantitative determination of 8-chloroadenosine 3',5'-monophosphate (8-Cl-cAMP) in dog plasma by capillary zone electrophoresis (CE) has been developed and validated. Samples of plasma (with 2'-O-monobutyryladenosine-3',5'-monophosphate as internal standard) were deproteinized with two volumes of acetonitrile. The supernatant was evaporated and reconstituted in water. A BioFocus 2000 system (Bio-Rad, Hercules, CA, USA) was used. The UV detector was set at 261 nm. The samples were loaded into uncoated fused silica capillary (40 cm x 50 microm) by pressure injection. A running electrolyte contained 30 mM SDS, 100 mM Tris, pH 7.55, with 20% of methanol added. The typical analytical conditions were: voltage, 18 kV; injection, 12 psi x s; capillary and carousel temperature were 20 degrees C. The linear relationship was observed between 0.063--2.00 microM using the time-corrected peak area ratio of 8-Cl-cAMP to internal standard with correlation coefficient greater than 0.99. The intra-day and inter-day coefficients of variation (CV's) were less than 12%. The developed method was used for the analysis of plasma samples from beagle dogs (n=12) to examine the toxicity of the anticancer drug, 8-Cl-cAMP, following two, 5-day cycles of continuous intravenous infusion at various doses of 8-Cl-cAMP as the sodium salt.