182 MANUAL OF MICROBIOLOGICAL METHODS 



sure type commonly referred to as the Barcroft respirometer and the 

 constant-volume type or Warburg instrument. Inasmuch as the War- 

 burg type is by far the most widely employed in this country, further 

 comments will be concerned with this apparatus. 



The fundamental principle involved is that quantitative changes in the 

 amount of a gas can be measured by determining pressure changes as 

 long as the temperature and volume of the gas are kept constant. The 

 apparatus consists of a flask or vessel having one or more side arms 

 equipped with stoppers and commonly containing a small center well 

 sealed to the bottom. The flask is attached to a manometer having one 

 closed and one open arm and usually containing Krebs' solution (Krebs, 

 1951) or Brodie's solution (density 1.03) rather than mercury (density 

 13.6), thus increasing the sensitivity of the pressure changes. The vessel 

 is placed in a water bath at constant temperature, and the system shaken 

 in order to increase the rapidity of gas exchange between the liquid and 

 gaseous phases. The level of the liquid in the closed arm of the manom- 

 eter is brought to a previously calibrated point (usually 150 or 250 mm), 

 and the level in the open arm recorded. In this manner the system is 

 always read at constant volume, and the recorded values represent pres- 

 sure changes. Inasmuch as the manometers are influenced by barometric 

 changes in pressure by virtue of having one end open to the atmosphere, 

 a thermobarometer (flask containing water attached to a manometer) is 

 included in all experiments. The pressure changes (positive or negative) 

 recorded for the thermobarometer are used to correct all readings of the 

 manometers attached to an experimental A^essel. 



The prime purpose of the side arm of the flask is to allow separation of 

 the components of the system under investigation so that one may con- 

 trol the initial reaction time. For example, a bacterial suspension in a 

 suitable buffer may be placed in the main compartment of the flask and 

 the substrate in the side arm. After temperature equilibration, the sub- 

 strate is added by removing the manometer from the water bath, closing 

 the open end of the manometer with one's finger, and tipping the system 

 at an angle sufficient to allow the material in the side arm to pour into 

 the main compartment. The manometer is then returned to the water 

 bath, and readings taken at suitable intervals, depending primarily on 

 the speed of the particular reaction under study. The stopper of the 

 side arm may be a venting plug, thus allowing introduction of gases of 

 known composition other than air. The center well is used primarily to 

 hold alkali to absorb CO2 as, for example, in the measurement of O2 

 uptake or H2 output. 



In actual operation certain information is required in order to make 

 possible the quantitative calculation of the gas evolved or taken up. 



