The pH in reversed micelles as imposed by the dihydrogen/proton redox couple and indicated by viologens and cytochrome c3 using hydrogenase as redox catalyst.

ABSTRACT

The pH values in reversed micelles were measured, making use of the hydrogenase enzyme as redox catalyst short-circuiting the viologen oxidized/semiquinone redox states. The hydrogenases from Desulfovibrio vulgaris (Hildenborough) and from Megasphaera elsdenii were applied. The observed pH values in reversed micelles were not dependent on the type of hydrogenase. Two cationic [cetyltrimethylammonium bromide and dodecylammonium propionate (DAP)] and two anionic sodiumdodecyl sulphate, sodium di(ethylhexyl)sulfosuccinate types of reversed micelles were used in combination with viologens having distinguishable valencies. It was observed that, in the cationic-reversed micelles, the dissociation constant for the semiquinone dimer had about the same value as compared to bulk water, while this value was significantly higher in the anionic-reversed micelles. Furthermore, the dissociation constant was independent of the concentration of viologen semiquinone in the reversed micelle, indicating that exchange kinetics are faster than the dimerisation process. With the exception of DAP, a linear relation exists, pH = a.pHrm + b, between the pH of the bulk water and the pH as measured in the reversed micelle (pHrm). In all these cases the value of a is smaller than unity, the value of b ranges between 1.6-2.7. For DAP the pHrm is always around 7. In DAP-reversed micelles, the counter-ion propionate probably serves as an internal buffer. Using cytochrome c3 as pH indicator in combination with N,N'-di(3-aminopropyl)-4,4'-bipyridinium)4+ to take care of electron transfer, in cetyltrimethylammonium-bromide-reversed micelles the pHrm is about the same as indicated by the viologen; in SDS-reversed micelles the pHrm is always lower than that indicated by N,N'-di(3-aminopropyl)4,4'-pyridinium4+. In contrast to cytochrome c3 from D. vulgaris, which in reversed micelles cannot become reduced directly by its D. vulgaris hydrogenase, the hydrogenase of M. elsdenii is able to reduce its ferredoxin directly.