Determination of isatin, an endogenous monoamine oxidase inhibitor, in urine and tissues of rats by HPLC.

1. We have previously identified isatin as one of the endogenous monoamine oxidase (MAO) inhibitors in the urine and the brain of stroke-prone spontaneously hypertensive rats (SHRSP), using gas chromatography-mass spectrometry (GC-MS). 2. In this study, we attempted to develop a convenient assay to determine isatin using high performance liquid chromatography with an ultraviolet detector (HPLC-UV). The standard curve for authentic isatin was linear at a range from 2 to 20 nmol per ml. The coefficient of variance was within 3% for both intra-assay and inter-assay. The sensitivity was 20 pmol per 10 microl of urine sample. 3. Isatin concentration correlated significantly and positively with endogenous MAO activity (tribulin-like activity) in both urine (r=0.924, P<0.001) and kidney extracts (r=0.862, P<0.01). There was a significant difference in urinary isatin between Wistar Kyoto rats (WKY) and SHRSP. Oral administration of isatin increased urinary isatin concentration and systolic blood pressure in WKY. 4. Determination of isatin using HPLC-UV may be useful for elucidating role of isatin in various conditions of stress and disease.