Spectro-electrochemistry

Electrochemistry should be included in pre-school curriculum!

Electrochemistry is an old discipline that is finding new and exciting applications in biological and chemical studies. Electrochemistry deals with reduction/oxidation reactions that take place on the surface of an electrode in the presence of electric potential.

Comparing to many typical chemical applications, expansion of bio-analytical and bio-engineering areas of electrochemistry is relatively recent. Nevertheless, electrochemistry is a promising, and in many respects unique approach in studies on redox-active proteins, enzymes, and their models. Its practical benefits come from simplicity of controlling and delivering electrical potential. In many cases electrochemical reactions can be reversed on demand making it ideal for studying reversible systems. Such flexibility makes electrochemistry a ideal companion for integration with other analytical techniques, such as spectroscopy.

From the perspective of biological studies there are two key applications for electrochemistry. First, electrochemistry can be used as a sensitive standalone analytical technique primarily focusing on the kinetics and thermodynamics of redox reactions and coupled steps. Second, electrochemistry provides a fine tool for precise non-invasive manipulation of biological samples and in conjunction with other analytical techniques. Presence of distinct color changes in most redox-active samples makes spectro-electrochemistry - a combination of spectroscopy and electrochemistry - an ideal combination that allows to resolve individual components even in complex systems.

X1 transmission thin layer electrochemical cell

In out studies on metalloenzymes and related chemical models we use so-called thin-layer variant of electrochemistry, where redox reactions take place only in a thin layer of sample covering electrode. This approach has multiple advantages for biological studies. Effective lack of diffusion in thin sample gives better resolution in analysis of multi-component systems. Sample volume as small as 0.05 ml or less preserves precious analyte. Very thin sample layer (0.01 mm) over transparent electrodes allows to use transmission electrochemistry in combination infra-red spectroscopy. At larger layer thickness (0.03-0.1 mm) thin layer electrochemistry can be combined with UV/Vis absorption spectroscopy.

750 grit gold mesh used as a transparent electrode in comparison with a 1/16" disk.

Recently we developed a new thin-layer transmission electrochemical cell. The 3-rd (X2) and 4-th (X1) generation cells were designed using Computer-Aided Design software and built from scratch at the MSU. The X2 cell provides over 90o clear field of view primarily intended for simultaneous spectral measurements, such as UV/Vis and IR transmission or combination of either method with photolysis, or reflective measurements. The X1 cell has a narrow field of view for a single-beam transmission measurements but uses smaller sample volume and gives much faster electrochemical response due to smaller electrode area. Both cell versions incorporate miniature Ag/AgCl reference electrode and allow accurate voltammetric measurements and can use most solid working electrode materials.

Optically transparent electrodes are key components of transmission spectro-electrochemical cells that define both electrochemical kinetics and optical properties of the cell. We use two types of transparent - traditional fine gold mesh electrodes and thin-film boron-doped diamond electrodes on solid substrates developed by Dr. Swain's group. Diamond film electrodes offer remarkable stability and good visible to IR transmission. They are also robust and resistive to fouling. Gold mesh electrodes are relatively fragile and require surface modification to avoid spontaneous unfolding of proteins non bare metal. However, metal mesh is completely achromatic and has unsurpassed transmission in the far UV spectral region.

6-step time-resolved cyclic potential step of thin layer cytochrome c.

Taken together, our recent efforts in developments in spectro-electrochemical methodology significantly improved sensitivity and stability of electrochemical and spectroscopic components and dramatically improved reliability of method. These improvements now allow us to study fine subtle spectral details over a broader spectral range. Example shown here illustrates stable, reversible spectral changes upon redox cycling of ferri/ferro cytochrome c at wavelengths as short as 210 nm. This region is of a particular importance for our research interests as characteristic absorption changes are expected in this region in most amino acid radicals and yet essentially nothing is known about far UV properties of such common redox cofactors as hemes, for example.We are working to expand such studies to other model proteins and enzymes. In parallel efforts we are working on methods of immobilization of biological samples on surfaces such as electrodes, particularly on increasing density of immobilized molecules to the level suitable for spectrolectrochemical studies.