Table 1.

A non-exhaustive list of measurements and standard calculations that may be performed on each sample during a fermentation characterization experiment

Goal	Measurement	Calculation	Description
Understand growth profiles	Biomass (OD, DCW, cytometry)	Growth rates	Calculation of maximum specific growth rates (μmax) during exponential phase, doubling time (td), timing of exponential growth within the fermentation, number of cell doublings within the fermentation (both main fermentation alone and including entire seed train)
Understand production rates and profiles	Product concentration, cell concentration	Production rates	Calculation of the rate of product formation (qp), both over the course of the fermentation and during specific intervals
Quantify residual substrate and substrate uptake rates and profiles	Substrate concentration, cell concentration	Substrate consumption rates	Calculation of the rate of substrate consumption (qs), both over the course of the fermentation and during specific intervals
Understand production efficiency	Product and substrate concentration	Production yields Carbon balances Yield coefficients for different substrates	Calculation of the efficiency of product formation (YSP = qp/qs), both over the course of the fermentation and during specific intervals Account for all the carbon substrate added to the fermentation in biomass, product, pathway intermediates and byproducts (including CO2) Calculation of the mass of cells produced per unit mass of substrate consumed—Yx/s during exponential growth
Identify and quantify pathway bottlenecks	Concentration of Pathway intermediates and byproducts	Byproduct formation	Calculation of the amounts of different byproducts/pathway intermediates formed during specific phases of the fermentation
Identify potentially limiting nutrients and substrates	Quantification of trace elements, Vitamins, AA, OA, Alcohols, N, P, S	Yield coefficients	Correlation of exhaustion of essential nutrients and decrease in growth rate or cessation of growth
Monitor cellular metabolism during fermentation	Offgas (CO2, O2 Ethanol)	Analysis of offgas and online measurements	Calculation of Oxygen Uptake Rates (OUR), Carbon Dioxide Evolution Rates (CER) and Respiratory Quotient (RQ) throughout the fermentation. Interpret OUR, CER, RQ, DO and pH trends in the context of identifying growth phases, metabolic shifts and loss of metabolic activity
Monitor the process and measure substrate uptake rates	Online measurements (pH, DO, feed rates, agitation, temperature)
Quantify viability loss during fermentation	Viability (cytometry, CFUs)	Viability	Quantification of viability loss during fermentation
Monitor cellular morphology and broth viscosity during fermentation	Microscopy Viscosity	Morphology Viscosity	Documentation of cellular morphology and viscosity during the fermentation

OD optical density, DCW dry cell weight, AA amino acids, OA organic acids, N nitrogen, P phosphate, S sulfate, DO dissolved oxygen, CFU colony forming units)