The development of a simple HPLC method for detection of the sea lamprey pheromone 3-ketopetromyzonol sulfate (3kPZS) is reported. The ketone moiety of the bile alcohol reacts readily with 1-dimethylaminonaphthalene-5-sulfonyl (dansyl) hydrazine to form the hydrazone. This hydrazone has a strong UV absorbance at 333 nm and visible fluorescence at 518 nm. Solutions containing the dansyl derivatized 3kPZS are readily analyzed by reverse phase HPLC with UV detection. A simple water-methanol gradient elutes the derivatized 3kPZS in just over 10 minutes using a standard 150 mm column. Detector response has high linearity throughout the ppm range, and the detection limit by UV is below 100 ppb. This methodology is useful for high throughput analysis of large numbers of samples in the ppm range.