Table 1.

Insights gained from different metabolic modelling methods. The methods differ in modeled metabolic layers and scales, cellular resolutions, modelling assumptions, and results interpretability. The differences in method inputs are described in Figure 3. The here summarized assumptions are general ones and methods may have additional assumptions.

	Gained insight	Applicable to whole metabolism	Applicable to individual pathways or reactions	Metabolic layer	Assumptions	Results require post-processing summarization	Applied to individual cells
Pathway-level analysis	List of enriched pathways or data-defined pathways; pathway activity	Yes	Yes	Reactions, rarely metabolites	Often disregards reaction associations within pathways or assumes that pathways are independent; sometimes approximately accounts for mass balance	Usually not as pathway-level information is already available	Usually not
Constraint-based analysis	Whole-system metabolic predictions of reaction fluxes and sometimes metabolite concentrations	Yes, but often simplified to reduce the necessary compute resources	No	Reactions and metabolites	Metabolite steady-state and mass balance; may assume availability of all substrates	Yes; in exploratory analysis as results are on the level of reactions or metabolites and thus pathway-level analysis is often applied	Yes, but often pooled to reduce the necessary compute resources
Kinetic models	Metabolite and reaction fluxes predictions through time or under different parameter setting; model selection	Usually not	Yes	Reactions and metabolites	Mass balance and Michaelis–Menten kinetics; may assume constant availability of all substrates	No, due to small systems	Yes