Heat production and chemical changes were measured in untreated and dinitrofluorobenzene (DNFB)-treated muscles during isometric tetani. Levels of total creatine (Ct), free creatine, ATP, ADP, AMP, inorganic phosphate, glucose-1-phosphate, glucose-6-phosphate, fructose-6-phosphate, fructose-1,6-diphosphate, pyruvate, phosphoenolpyruvate, and lactate were measured. Changes in inosinic acid (IMP) were also measured. 2. DNFB effectively inhibited the creatine kinase reaction (Lohmann reaction). 3. Our major finding is that even after effective treatment with DNFB the observed heat plus work after 2 sec and 5 sec of stimulation is significantly greater than the enthalpy change produced by the measured chemical changes. This confirms that an unidentified exothermic process occurs during muscle contraction; this conclusion was reached previously from studies of untreated muscle. 4. The unexplained heat plus work is unlikely to be derived from glycolytic reactions since under anaerobic conditions no formation of lactate, pyruvate, phosphoenolpyruvate or fructose-1,6-diphosphate could be detected in either untreated or DNFB-treated muscles even 34 sec after a series of three 5 sec isometric tetani. 5. In the first 2 sec of stimulation the unexplained heat plus work is less in DNFB-treated muscles than in untreated muscles. However from 2 to 5 sec of stimulation the unexplained heat plus work is the same in treated and untreated muscles.
