1206
Anal. Chem. 1982, 54, 1206-1208
1
--87 - +70
I
-+I03
+310
I
,
,
,
,
,
,
I ~
450
500
550
EOOnm
Figure 3. Spectra of a 16 WM solution of cytochrome c in a solution with 1 mM V(EDTA) and 100 mM sodium phosphate buffer pH 7.0. The
potentials at which each of the spectra were measured are given at the right. The spectra were measured separately and collected in one figure. The spectra are not corrected for the mediator background changes. +0.25 V vs. NHE for cytochrome c at pH 7.0. This is in the range found for the enzyme under similar conditions (ref 10 and citations therein) using other mediators. The n value obtained from the slope was 1. The poise potential measured by the Au test electrode was assumed to represent the equilibrium voltage of the solution phase and, hence, that of the electroactive enzyme species. This was substantiated in the case of cytochrome c; the Eo' measured in the presence of millimolar vanadium mediator is in the range found with other mediators and methods (IO). The potentiometric indicator electrode can be any conducting material such as platinum, gold, or glassy carbon. In this study the same material as composes the working electrode was used. An alternative method of measuring the solution potential was also investigated. One can switch out the potentiostat and connect the reference and working electrodes to the voltmeter to measure the poise potential. This arrangement would simplify construction by eliminating one reference electrode, the separate potentiometric indicator electrode, and their connectors. Similar (within a few millivolts) poise values were obtained when this switching was done manually. In use, following the initial f i g of the pneumatic mixing tube, no gas bubbles are introduced into the optical path as long as the tube from the pneumatic mixer is beneath the surface of the solution. Also, there is little or no effect on the noise in the spectrum from the pulse stirring unless part of the beam passes over the top of the sample solution in the cell and, thus, encounters a varying sample volume. A further point of concern is worth mentioning. In a spectroelectrochemical cell with a long path length, the sample is significantly lower in concentration than in a more typical, short (
