Dynamic metabolic flux analysis using B-splines to study the effects of temperature shift on CHO cell metabolism

Abstract

Metabolic flux analysis (MFA) is widely used to estimate intracellular fluxes. Conventional MFA, however, is limited to continuous cultures and the mid-exponential growth phase of batch cultures. Dynamic MFA (DMFA) has emerged to characterize time-resolved metabolic fluxes for the entire culture period. Here, the linear DMFA approach was extended using B-spline fitting (B-DMFA) to estimate mass balanced fluxes. Smoother fits were achieved using reduced number of knots and parameters. Additionally, computation time was greatly reduced using a new heuristic algorithm for knot placement. B-DMFA revealed that Chinese hamster ovary cells shifted from 37 °C to 32 °C maintained a constant IgG volume-specific productivity, whereas the productivity for the controls peaked during mid-exponential growth phase and declined afterward. The observed 42% increase in product titer at 32 °C was explained by a prolonged cell growth with high cell viability, a larger cell volume and a more stable volume-specific productivity. Keywords: Dynamic, Metabolism, Flux analysis, CHO cells, Temperature shift, B-spline curve fitting