Discussion 123 



kinase-lactic dehydrogenase system, ATP by using the phosphoglycerol 

 kinase-glyceraldehyde phosphate dehydrogenase system. Our results 

 are given in Table I. For each experiment the concentration of ATP 



Table I (Hess) 



ATP/ADP RATIO IN ISOLATED LIVER MITOCHONDRIA (STATE 4) 



Concentration 

 [iMlml. 



Expt. No. 



1 

 2 

 3 

 4 

 5 



Conditions: phosphate buffer, pH 7-4 (0-01 M) 3 -hydroxy butyrate 10 (jlM, Mg^+ 50 (XM, 

 mitochondria: 8-8 mg. biuret value; nil. 



and ADP, the sum of both and the ratio is given. Experiment 1 shows 

 the results giving the endogenous concentration without addition of 

 external ADP. Here the steady state concentration of ADP is very low 

 and well within the range we would expect from the titration data for 

 the ADP affinity (K^) for the respiratory chain below 56 [jlm which have 

 been given by Prof. Chance. In experiments 2-5, ADP was added from 

 outside to the mitochondria, which were deproteinized and analysed 

 after state 4 was reached according to the oxygen kinetics recorded 

 with a platinum electrode. Here, again, the steady state concentration 

 of ADP is very low and within the expected range, but not as low as in 

 experiment 1. This small increase might be due to methodological 

 factors. The time required for deproteinization and analytical treat- 

 ment of the samples is enough to allow some ATP to be broken down to 

 ADP and to spoil the analysis. However, the values for the ATP/ADP 

 ratio illustrate quite well the high efficiency of the mitochondria in 

 keeping the theoretically expected steady state value of ADP. 



In order to demonstrate a pathological, abnormal efficiency of mito- 

 chondria, it is useful to influence the ATP/ADP ratio by means of tri- 

 iodothyronine or thyroxine, the uncoupling action of which was shown 

 in 1951 (Martins, C, and Hess, B. (1951). Arch. Biochem., 33, 486). The 

 influence of tri-iodothyronine on the ratios in mitochondria is shown in 

 Table II. Tri-iodothyronine was added to the mitochondria immediately 

 prior to the addition of ADP. The percentage decrease in the ratio 

 (right-hand column) demonstrates the inefficiency of the mitochondria in 

 maintaining the normal ratio as a function of the concentration of tri- 

 iodothyronine. 



With regard to the phenomenon of respiratory control as applied to 

 intact cells, I shall give some data on the glucose-oxygen stoichiometry 

 in ascites tumour cells. The addition of glucose to these cells causes an in- 

 crease and, after 30-60 seconds, a strong blockage of oxygen uptake as 



