The reactions between the halogenofluorescein-mercury compounds such as mercurochrome and penicillins such as penicillin G (PCG), ampicillin (ABPC), sulbenicillin (SBPC) and amoxicillin (AMPC) were investigated. Detection tests and fluorometric determinations of these penicilins are proposed using the coloring system of mercurochrome and penicillin reactions. The detection limit of penicillins, as ABPC, was 0.8 μg/ml, and the calibration curves were rectilinear up to about 5 μg/ml (about 1.5 X 10-5 M) of PCG, ABPC, SBPC and AMPC at 535 nm. The relative standard deviation was 2.7% for 1.4 μg/ml of ABPC (n=5).