In fermentation, the energy conservation (ATP-synthesis) generally takes place by way of substrate-level phosphorylation and by way of decarboxylation of organic acids in certain cases.
Way # 1. Substrate-level Phosphorylation and Fermentation:
Substrate-level phosphorylation, is a mechanism by which high energy phosphate bonds from organic intermediates of the fermentation are transferred to ADP producing ATP. ATP synthesis via substrate-level phosphorylation can take place in various different ways; in all cases, the central point is the production of one or another high energy intermediate compound.
1. High energy intermediate compounds:
The high energy intermediate compounds are usually organic compounds containing a phosphate group or a coenzyme-A molecule, the hydrolysis of which is highly energy- releasing (exergonic). Many such high energy intermediate compounds are given in Table 26.1.
Since most of the intermediate compounds listed in Table 26.1 can couple directly to ATP synthesis, if an organism can form one or another of these compounds during fermentative metabolism, it can synthesize ATP.
Substrate-level phosphorylation is a more direct way of making ATP than via proton motive force (oxidative phosphorylation) but requires that the energy source couples directly to a high energy intermediate compound.
2. Pathways of formation of high energy intermediate compounds:
There are many pathways for the anaerobic degradation of various fermentable substances by microorganisms to high energy organic intermediate; some major ones are summarized in Fig. 26.2. Either one of these compounds or its related derivative is produced during each case leading to synthesis of ATP by substrate-level phosphorylation.
The maximum number of ATP synthesized per substrate-level phosphorylation during each pathway is two; many substrates synthesize even less.
Although the free energy released during a particular stage of substrate-level phosphorylation may be theoretically high, the net ATP synthesis does not cross the maximum limit of two, because:
(i) The microorganisms usually operate at considerably less than 100% efficiency, and
(ii) Some of the free energy is lost as heat.
For example, the anaerobic breakdown of glucose to ethanol and CO2 has a theoretical energy yield of -235 kJ/mole, sufficient to synthesize about 7 ATP molecules (each ATP molecule synthesis from ADP and Pi (inorganic phosphate) needs -31.8 kJ free energy), but only 2 ATP molecules are actually produced. 
Way # 2. Decarboxylations of Organic Acids:
There are certain fermentative pathways in which the catabolism of the substrate (breakdown of substrate releasing energy) is linked to ion pumps that establish a proton (H+) or sodium (Na+) gradient across the plasma membrane. The reason behind it is said to be the release of insufficient free energy to couple to the synthesis of ATP directly by substrate-level phosphorylation.
Succinate fermentation by Propionigenium modestum (with the help of Na+ gradient) and oxalate fermentation by Oxalobacter formigenes (with the help of H+ gradient) are the examples of such fermentations. In both cases the microorganisms couple the fermentation of dicarboxylic acids to membrane-bound energy-linked ion pumps.
The interesting and unique aspect of the fermentation metabolism of both P. modestum and O. formigenes is that ATP synthesis takes place without substrate-level phosphorylation or electron transport chain; however, chemiosmotic ATP synthesis still occurs as a result of Na+ or H+ pump linked to decarboxylation of organic acids. The mechanism of succinate fermentation by P. modestum is as follows, for convenience.
Succinate fermentation by Propionigenium modestum:
The overall reaction of succinate fermentation by Propionigenium modestum is the following:
Succinate2- + H2O → Propionate– + HCO–3
This overall reaction yields insufficient free energy (∆G° = -20.5 kJ/reaction) to couple to ATP synthesis directly by substrate-level phosphorylation, but nevertheless it serves as the sole energy-yielding reaction for growth of the microorganism.
This becomes possible because the decarboxylation of succinate by the bacterium couples to the extrusion of Na+ across the plasma membrane. A Na+ ATPase associated with the plasma membrane of P. modestum uses this sodium gradient to drive ATP synthesis (Fig. 26.3).
