Direct electron transfer (DET) from bare spectrographic graphite (SPGE) or 3-mercaptopropionic acid-modified gold (MPA-gold) electrodes to Trachyderma tsunodae bilirubin oxidase (BOD) was studied under anaerobic and aerobic conditions by cyclic voltammetry and chronoamperometry. On cyclic voltammograms nonturnover Faradaic signals with midpoint potentials of about 700 mV and 400 mV were clearly observed corresponding to redox transformations of the T1 site and the T2/T3 cluster of the enzyme, respectively. The immobilized BOD was differently oriented on the two electrodes and its catalysis of O(2)-electroreduction was also massively different. On SPGE, where most of the enzyme was oriented with the T1 copper site proximal to the carbon with a quite slow ET process, well-pronounced DET-bioelectroreduction of O(2) was observed, starting already at >700 mV vs. NHE. In contrast, on MPA-gold most of the enzyme was oriented with its T2/T3 copper cluster proximal to the metal. Indeed, there was little DET-based catalysis of O(2)-electroreduction, even though the ET between the MPA-gold and the T2/T3 copper cluster of BOD was similar to that observed for the T1 site at SPGE. When BOD actively catalyzes the O(2)-electroreduction, the redox potential of its T1 site is 690 mV vs. NHE and that of one of its T2/T3 copper centers is 390 mV vs. NHE. The redox potential of the T2/T3 copper cluster of a resting form of BOD is suggested to be about 360 mV vs. NHE. These values, combined with the observed biocatalytic behavior, strongly suggest an uphill intra-molecular electron transfer from the T1 site to the T2/T3 cluster during the catalytic turnover of the enzyme.

The desorption of 2-mercaptoethanesulfonate (MES) spontaneously adsorbed on Au(111) has been studied by using both potential-step and voltammetric experiments. From the amount of gold oxide formed during the oxidation sweep in the fingerprint region it is shown that the adsorption process induces structural changes of the gold surface. It is also shown that together with the reductive desorption of MES ions a concomitant faradaic process occurs. The results suggest that this process is connected to the reduction of solvent on the structurally modified gold electrode. The reductive desorption process of MES undergone at more negative potentials is characterized by a single peak in the voltammetric response and the presence of a maximum in the chronoamperogram. It is shown that the logarithm of the maximum current, the time at which the maximum current appears, and the peak width at half height depend linearly on potential. An experimental protocol for the desorption/readsorption of MES based on a potential-step experiment followed by cyclic voltammetry is outlined as an appropriate tool to analyze simultaneously the desorption of adsorbed and readsorbed MES. A linear dependence between the two desorption peak potentials and the surface concentration of adsorbate was observed. Changes in the environment surrounding the adsorbed moities and in the potential of zero charge of the electrode are shown to be the factors ruling this dependence.

A new method is proposed for the determination of the potential of zero charge of gold electrodes modified with thiol monolayers. It makes use of the immersion technique, in combination with a vapor deposition protocol to build the thiol monolayers. As compared to previous methods, the present approach provides more accurate results, particularly in the case of long-chain alkanethiol monolayers, and it is applicable to any monolayer irrespective of its degree of hydrophilicity. Results are presented for a series of 12 alkanethiol monolayers and for 11-mercaptoundecanol and 11-mercaptoundecanoic acid monolayers. Good agreement is found between the variation of the potential of zero charge along the alkanethiol series with the corresponding change of the surface work function. The potential of zero charge of the 11-mercaptoundecanoic acid monolayer is shown to depend on the extent of dissociation of the acid, thus opening the possibility of applying this type of measurements to the study of surface ionization processes.

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