Oroboros 1989

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Oroboros 1989

Mitochondria and Cell Research - the Oroboros Ecosystem

  • Project supported by the Fonds zur Förderung der wissenschaftlichen Forschung, Austria, FWF Project P7162-BIO: Thermodynamics and respiratory control in aerobic and anoxic mitochondria.
The prototype of the Oxygraph was developed and applied in this project (1989-1992).
Dr. Josef Martin Bergant (FWF) initiated the contact between the research group of Dr. E. Gnaiger (principal investigator; Dept. Zoophysiology, Univ. Innsbruck) and industry (Fa. Paar, Graz).
This lead finally to the foundation of Oroboros Instruments as a spin-off from basic research supported by the FWF.

FWF Project Summary

E. Gnaiger, Univ. Innsbruck, Austria
SUMMARY(FWF Project P7162-BIO)
Despite the advances in elucidating the chemosmotic and enzymatic mechanisms of oxidative phosphorylation, the much debated theory on the dynamics of mitochondrial respiration remains controversial. Questions on the interface of thermodynamic and kinetic control mechanisms are unresolved. In the present project experiments will be designed to test the hypothesis that respiratory flux in coupled isolated mitochondria is under thermodynamic control under a much wider range of conditions than previously conceived. A new nonequilibrium thermodynamic concept, developed within the frame of the project, will have important implications on our general view of the dynamics of chemical reactions, providing a basis for the separation of thermodynamic and kinetic effects.
As empirical tests, studies of isolated mitochondria are proposed using respirometric, biochemical and direct calorimetric methods under aerobic, hypoxic and anoxic conditions. Electron microscopic analyses will serve as a control of the quality of isolation of mitochondria from various mammalian tissues and from euryoxic invertebrates. Correlations between mitochondrial ultrastructure and respiratory flux will be investigated under incubation conditions exceeding the thermodynamic control range, to test the quantitative importance of mitochondrial structure‑function relations in the control of flux.
The conceptual and methodological developments in the present project will improve our understanding of respiratory control in isolated mitochondria and tissues under physiological and pathological conditions. The proposed concept and the microcalorimetric method have elicited an interest in collaboration on an international and national basis in a wide range of relevant research, spanning from regulatory biology, cardiology, issues of pathological long-term hypoxia, to ecologically important aspects of microbial energetics.
An improved theory on the thermodynamics of irreversible chemical processes is required because conflicting relations are observed between mitochondrial oxygen flux and phosphorylation potential. Depending on the incubation conditions, linear or entirely irregular flux/force relations are observed within identical limits of departure from equilibrium (Jacobus 1985). The outline of a new theory on the dynamics of chemical reactions is an integral part of the project proposal. The theory will be developed on the basis of the fundamental equations of diffusion (Einstein 1905). The form of Einstein's diffusion equation, J = -u c F, where u and F are mobility and force (gradient of the chemical potential) respectively, does not generally predict linear relations between flow and force. This is due to the fact that concentration, c, is variable. Einstein's diffusion equation contrasts with the form of the thermodynamic linear phenomenological laws, J = -L F. The conductivity, L, for diffusion is the product of mobility and concentration. In a linear concentration gradient the conductivity is a nonlinear function of the force, which is considered prohibitive for the use of Onsager linear thermodynamics. Therefore, we are forced to drop the nonequilibrium thermodynamic paradigm of flux/force linearity in the near-equilibrium region.
Next, a form of Einstein's diffusion equation for discontinuous systems is required, which reveals an important analogy between the concentration difference in Fick's First Law of diffusion and osmotic pressure. The concentration (activity) difference times RT is the diffusion pressure, linearly related to diffusive flow by the mobility. Generalized fluxes are linearly related to generalized pressures far beyond the near-equilibrium range, in contrast to the restricted range of approximate flux/force linearity. For chemical reactions, the "reaction pressure" is defined in analogy to osmotic pressure, as the product of the chemical driving force and a concentration term, the generalized "free activity".
A successful application is presented of the concept on "chemical reaction pressure" by providing the first explanation for the flux‑force linearity observed in a kinetically complex gas reaction (data from Prigogine et al. 1948). More importantly, linear as well as irregular flux/force relations on liver and heart mitochondria (Jacobus 1985; and original data provided by Dr.W.E. Jacobus) are fully consistent with the new flux/pressure concept. In all test cases analyzed so far, linear flux/pressure relations were obtained. The concept offers an explanation of extended flux/force linearities, defines the limits of the linear flux/force range, and rationalizes irregular flux/force relations in terms of the linear relationship between flux and pressure of chemical reactions. This provides a significantly improved method, for understanding the integrated operation of thermodynamic and kinetic mechanisms in the control of metabolic flux.