What are the assumptions made during the calculation of net gain of ATP?

Calculating the net gain of ATP for each glucose molecule oxidised is doable, but it could only be a theoretical exercise in practise. These calculations can only be made based on the following assumptions:

i.There is a sequential, orderly pathway is in function, with one substrateforming the next and glycolysis, TCA cycle, and ETS pathway occurringone after the other.

ii.The NADH produced during glycolysis is transferred to the mitochondriaand undergoes oxidative phosphorylation. None of the intermediates inthe pathway is used to make another compound.

iii.Only glucose is respired. No other alternative substrates enter thepathway at any of the intermediate stages.

These kinds of assumptions, however, are not valid in a living system. All pathways occur simultaneously and do not occur one after the other. Substrates enter the pathways and are withdrawn from them as needed. ATP is used as and when it is required. Multiple factors influence enzymatic rates. As a result, aerobic respiration of one molecule of glucose can result in a net gain of 36 ATP molecules.