Dynamic modelling of tetrazolium-based microbial toxicity assay-a parametric proxy of traditional dose-response relationship.

Microbial toxicity of test substances in tetrazolium assay is often quantified while referring to their IC50 values. However, the implication of such an estimate is very limited and can differ across studies depending on prevailing test conditions. In this work, a factorial design-based end-point microbial toxicity assay was performed, which suggests a significant interaction (P= 0.041) between inoculum and tetrazolium dose on formazan production. Subsequently, a dynamic model framework was utilized to capture the nonlinearities in biomass, substrate, formazan profiles and to project the toxicant inhibition parameter as a robust alternative to IC50 value. Microbial growth, glucose uptake and formazan production in the presence or absence of toxicant (Cu2+) from designed batch experiments were used for sequential estimation of model parameters, and their confidence intervals. A logistic growth model with multiplicative inhibition terms for formazan content and toxicant concentration fits the experimental data reasonably well (R2>0.96). Dynamic relative sensitivity analysis revealed that both microbial growth and formazan production profiles were sensitive to toxicant inhibition parameter. The modelling framework not only provides a better insight into the underlying toxic effect but also offers a stable toxicity index for the test substances that can be extended to design a versatile, robust in vitro assay system.