Comparison of the Tetrazolium Salt Assay for Succinate Dehydrogenase with the Cytosensor Microphysiometer in the Assessment of Compound Toxicities

Abstract

The cytosensor microphysiometer (a biosensing instrument for detecting cellular metabolism) was compared to the established tetrazolium salt assay as a chemosensitivity test. Two coumarin compounds, 7-hydroxycoumarin and esculetin, were examined to determine their effect on the cellular metabolism of A431 cells over a 24-h exposure period. In the tetrazolium salt assay, 7-hydroxycoumarin caused suppression of the succinate dehydrogenase activity at concentrations greater than 10 μg/ml. Esculetin exerted a more serious effect on succinate dehydrogenase, with decreases in activity observed at greater than 1 μg/ml. The observed effect was dose-dependent for both compounds examined. The metabolic activities of cells exposed to 100 μg/ml of drug were 90.37 ± 2.8 and 71.62 ± 2.96 (n = 3), of control values, for 7-hydroxycoumarin and esculetin, respectively. Using the cytosensor microphysiometer to assess metabolic activities, a similar pattern of inhibition was observed, with esculetin more detrimental to cellular metabolism than 7-hydroxycoumarin. The effect was dose- and time-dependent for both compounds. 7-Hydroxycoumarin (100 μg/ml) caused the cellular metabolic rate to drop to 44.21 ± 5.34% (n = 4) of the control metabolic rate, while 100 μg/ml esculetin caused the metabolic rate to fall to 21.5 ± 4.54% (n = 4) of the control rate. The cytosensor method proved to be superior to the tetrazolium salt assay for a number of reasons, which are discussed in this paper.