Open AccessTurnover Rates and Intracellular Pool Size Distribution of Citrate Cycle Intermediates in Normal, Diabetic and Fat‐Fed Rats Estimated by Computer Analysis from Specific Activity Decay Data of 14 C‐Labeled Citrate Cycle Acids
Author(s) -
Sauer Frank,
Erfle James D.,
Binns Michael R.
Publication year - 1970
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1970.tb01173.x
Subject(s) - flux (metallurgy) , chemistry , citric acid cycle , steady state (chemistry) , citric acid , in vivo , metabolite , isotope , metabolism , biochemistry , endocrinology , biology , physics , microbiology and biotechnology , organic chemistry , quantum mechanics
Specific radioactivity curves of citrate, malate, glutamate and aspartate were measured in vivo in liver from normal, fat‐fed and diabetic rats with [1‐ 14 C]acetate as precursor at time intervals of 0.1 to 20 min. A mathematical model of 14 pools and 40 interconnecting flux rates been constructed and by computer analysis on the Univac 1108 flux rates and intra‐ and extra‐mitochondrial metabolite pool distribution calculated from the measured specific radioactivity curves. Flux rates and pool distributions were varied by using an automatic minimization routine to minimize the difference between observed and theoretical values until an acceptable fit of computer generated specific radioactivity time curves to experimentally observed specific radioactivities was obtained. Steady state kinetics were assumed, i.e. the model was fixed with no change in pool sizes over the measured time interval. The main conclusions were:1 A state of non‐equilibration of isotope exists for the acetyl‐CoA and acetylcarnitine, citrate, malate, aspartate, glutamate and α‐ketoglutarate pools distributed between cytoplasmic and mitochondrial compartments. 2 Cytoplasmic pools of citrate, acetyl‐CoA and acetylcarnitine account for the slow rate of radioactivity decay observed for these metabolites in vivo . 3 Key flux rates were acetate entry into the citrate cycle of 0.36, 0.28 and 1.05 μmoles × min −1 × g liver −1 in normal, fat‐fed and diabetic rats. Citrate turnover rates were 1.21, 0.37 and 1.94 μmoles × min −1 × g liver −1 for normal, fat‐fed and diabetic rats. The acetate pool was 0.23 μmoles/g liver in normal, 0.44 μmoles/g liver in fat‐fed and 0.38 μmoles/g liver in diabetic rats. Metabolite pool distribution was greater in the extra‐ than intramitochondrial space: the shift was most marked in the diabetic rat where the intra‐ to extramitochondrial malate distribution was in the ratio of 1:21.5.