Redox titration, equilibration

The pH dependence of the redox midpoint potential, En,(g ), of Qa is obtained by integrating over pH the proton uptake by DQa given in Fig. 2 (5). We find that Em(g > is -proportional to pH with a slope of -20 mV per pH. This is in striking contrast with a slope of -60 mV per pH found by redox titrations using dyes (4,10,14). The origin of this discrepancy is not understood at present. One expiration that has been invoked is that the redox titration may be affected by poor equilibration between Qa and the redox dyes. An alternate explanation is based on the way in which Qa was reduced in all direct proton uptake experiments, Qa was photoreduced, whereas in the redox titrations,... 

Electrochemical titrations of redox proteins with spectroscopic monitoring bear the advantage that the sample is not successively diluted with redox reagents, that titrations can be performed in cycles, and that the electrode potential (and thus the ox/red equilibrium of the sample) can be set at mV precision. This precision offers the possibility to analyze redox proteins with several cofactors that may be coupled and that may exhibit cooperativity. As an example, Fig. 2 shows the complete redox titration of cytochrome c oxidase, the terminal enzyme of the respiratory chain. Provided that fast equilibration (typically within seconds) at the electrode occurs, a full spectral data set can be obtained within some minutes. 

PHREEQE can calculate pH, redox potential, concentration of elements, molalities and activities of aqueous species, and mineral or gas mass transfer as a function of reaction progress. The program is capable of simulating reactions due to mixing, titrating, net irreversible reaction, temperature changes, and mineral- or gas- phase equilibration. 

A more accurate way to use potentiometric data is to prepare a Gran plot6-7 as we did for acid-base titrations in Section 11-5. The Gran plot uses data from well before the equivalence poinl (Ve) to locate Ve. Potentiometric data taken close to Ve are the least accurate because electrodes are slow to equilibrate with species in solution when one member of a redox couple is nearly used up. 

Xanthine is converted to uric acid at the molybdenum center of the enzyme, and the electrons are removed from the enzyme by oxidation of the flavin center. From early reductive titrations of xanthine oxidase with sodium dithionite, it was proposed that reducing equivalents were equilibrated among the four redox-active centers (Mo-co, two separate Fe2S2 centers, flavin) at a rate that was rapid relative to the overall catalytic rate of substrate turnover (243). Under such conditions, the flux of reducing equivalents through the enzyme should be influenced by the relative reduction potentials of the redox centers involved (244). Any effects of pH and temperature on the reduction potentials of individual redox components would affect the apparent rates of intramolecular transfer of the enzyme. 

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