Inhibition of Escherichia coli Cell-Free β-Galactosidase Activity by Binary Mixtures of Water-miscible Solvents

Abstract

The inhibitory effects of ethanol-dimethylsulfoxide (DMSO) and ethanol-N,N-dimethylformamide (DMF) binary mixtures on the activity of cell-free β-galactosidase from Escherichia coli were assessed. The concentration-response relationships of the individual solvent and the mixtures were fitted to a Gormpertz model to estimate the median inhibitory concentrations (EC50) and No-Observable-Effect-Concentration (NOEC) thresholds. The EC50 values estimated are 29.246 ± 2.986% (ethanol), 28.112 ± 0.471% (DMSO), and 18.244 ± 0.674% (DMF). The NOEC values are 11.265 ± 1.121% (ethanol), 6.4047 ± 0.564% (DMSO), and 1.897 ± 0.427% (DMF). The order of inhibitory effects is DMF > DMSO > ethanol. The mixture NOEC values ranged from 10.081% (6:4 ethanol-DMSO mixture) to 13.619% (9:1 ethanol-DMSO mixture). In the case of ethanol-DMF mixtures, NOEC values ranged from 6.843% in 6:4 ethanol-DMF mixture to 10.599% in 9:1 ethanol-DMF mixture. All solvent mixtures with higher EC50 and NOEC values are less inhibitory than the individual DMSO and DMF. The inhibitory effects of the ethanol-DMSO and ethanol-DMF binary mixtures were predicted using the concentration addition (CA) model, toxic index (TI) and model deviation ratio (MDR). The TI for the ethanol-DMSO mixtures ranged from 1.076 ± 0.046 to 1.142 ± 0.032, while that of ethanol-DMF ranged from 1.027 ± 0.015 to 1.136 ± 0.024. These values are marginally higher than 1; thus, the combined effect is considered additive. The study provides fundamental information on the sub-inhibitory concentrations of ethanol, DMSO, and DMF, or their mixtures, for use in the permeabilization of E. coli cells for a β-galactosidase activity assay.