Rapid measurement of the monoamine content in small volumes of rat plasma

A method for the simultaneous measurement of serotonin, catecholamines, and their metabolites, 5-hydroxy-indoleacetic acid, homovanillic acid, and 3, 4-dihydroxyphenylacetic acid, by ultrafiltration and microbore high-performance liquid chromatography with dual electrochemical detection in small plasma volumes was established. Unlike the traditional assays which require at least 1-2 ml of plasma for each measurement, the present method uses only a 20-microliters sample volume. Since blood loss is minimal, repeated blood sampling from a single animal as a rat becomes practicable. The present microassay provides low detection limits (signal-to-noise ratio = 3) for all analytes (0.2-0.5 pg per 5-microliters injection or 50-120 pg/ml plasma). Isocratic separation of these analytes on a microbore column is achieved within 15 min. This rapid and sensitive method can be used as a routine research tool in various physiological or pharmacokinetic studies especially in small animals.