PHYSIOLOGICAL AND BIOCHEMICAL TECHNICS 183 



One must know at a given temperature the gas volume of the flask and 

 manometer (to the liquid reference level in the closed arm) , the volume of 

 fluid in the flask, the gas being exchanged, the solubility of that gas, and 

 the density of the manometric fluid. 



The Warburg technic is commonly used to measure rates of O2 or H2 

 uptake, CO2 or H2 production, and respiratory quotients with a variety of 

 substrates. When properly used this method provides one of the most 

 versatile tools for the microbial physiologist. It is pertinent here to 

 point out that although the manometric apparatus itself allows only the 

 measurement of gas exchange, it is often desirable to subject the contents 

 of the vessel to chemical analysis at the conclusion of the manometric 

 experiment. After deproteinization and centrifugation, the supernatant 

 liquid may be analyzed for a variet}^ of common intermediates and end 

 products of metabolism, for many of which excellent micromethods are 

 available. It is quite possible with some bacteria and with certain sub- 

 strates to obtain an accurate dissimilation balance with the usual volume 

 of contents (about 3 ml) employed in the Warburg flask. 



Analytical Procedures 



The material in this section is usually presented under the heading of 

 "Fermentation Analysis." The less restricted title used above, however, 

 is considered more appropriate because fermentation represents only one 

 of the dissiiTiilatory mechanisms found in microorganisms. The term 

 ''fermentation," or intramolecular oxidation, is properly reserved for 

 those anaerobic processes in which the hydrogen acceptor originates from 

 the substrate. Other mechanisms include anaerobic oxidation or inter- 

 molecular oxidation w^herein carbonate, nitrate, sulfate, or an organic 

 compound acts as the hydrogen acceptor and respiration or aerobic oxida- 

 tion wherein oxygen serves as the hydrogen acceptor. Coupled reactions 

 between two substrates have also been described as, for example, the 

 Stickland reaction between pairs of amino acids (Stephenson, 1949). 

 These distinctions are important if a valid mechanistic interpretation of 

 the results is to be obtained and^ used to describe the probable inter- 

 mediary pathways of dissimilation. The majority of the studies pub- 

 lished to date deal with the products of carbohydrate, polyalcohol, and 

 organic acid dissimilation. The principles in all cases are the same. 



Measurement of dissimilatory products must include both their quali- 

 tative identification and quantitative determination. The quantity of 

 substrate dissimilated, as well as any substances reacting T\dth the sub- 

 strate during its breakdown (e.g., oxygen), must be accounted for 



