Effect of phospholipids,Triton X-100 and biological membranes on redox systems involving tetrazolium salt reduction. Its implications for the assay of enzymatic activities

The influence of phospholipids and Triton X-100 on the time course of chemical and enzyme-mediated reductions of a commonly used tetrazolium salt, MTT, was studied. MTT reduction was followed by the absorbance changes at 570 nm. With ascorbate as reducing agent, a 3-fold increase in the initial rates of the absorbance changes and a 24 % increase in the final absorbance values were observed in the presence of Triton X-100 micelles or phospholipid vesicles. The enzyme-mediated reduction of MTT with NADH generated by the NAD-dependent lactate dehydrogenase was also enhanced in the presence of Triton X-100, phospholipids or erythrocyte membranes. No enhancement was observed following the enzymatic generation of NADH at 340 nm in the absence of MTT. The above findings were interpreted as arising from: a) solubilization or reduced MTT in the detergent micelles or phospholipid vesicles which favors the redox reaction occurring in the aqueous fase, and b) changes in the spectral properties of reduced MTT in aqueous and lipid-like media.