Updated project metadata. Under well-defined laboratory conditions, we grew R. pomeroyi DSS-3 and A. macleodii MIT1002 in batch cultures on a monosaccharide (glucose) and organic acid (acetate), provided either individually or in combination, and all at the same carbon equivalent. This batch culturing approach mimicked bacterial DOC assimilation in short-lived substrate ‘hot spots’, such as those formed by high phytoplankton extracellular release at peak photon availability. Measurements were made of bacterial metabolite uptake, respiration, and biomass accumulation through a growth cycle. Insights into bacterial core metabolism came from gene and protein expression measured at intervals during growth. Curated genome-scale models (flux balance analysis; FBA) were used to explore the metabolic foundation of CO2 production for insights into determinants of BGE and bCUE.