these techniques. Some group members also told me that they use liquid chromatography-mass spectrometry (LC- MS), and it is a great help in analyzing their reaction products. Methods like atomic force microscopy, scanning electron microscopy, and surface FTIR are used in the group for characterization of the electrode surfaces they make. A week or so later: I have done my first electrochemical synthesis experiment. I did Electro-organic Reactions in undergraduate analytical lab, so it was not totally new. The reactor cell I used was like two connected beakers in a water jacket (Fig. 1). One beaker is and Redox Active Biomolecules: the working electrode compartment, and the other is the counter electrode compartment, separated by a salt bridge. A Student Diary* This is so that products from the two compartments do not combine to give by James F. Rusling and Albert J. Fry undesirable products. The reactor had a carbon cloth working electrode, which is a cool material, like a conducting It is early December, and I have been in chemistry grad school black fabric that you cut to the right for about 3 months. It is a big transition, and a bit tougher size with scissors. It has a large surface than I expected. The courses are not too hard, but the focus area to facilitate the catalytic reaction. The reactor has counter and reference here is very much on research. I have chosen the group in electrodes, and is hooked up to a which I will do my thesis research, and I am happy about that. device called a potentiostat to control This group does on organic and biological the voltage at the working electrode. The working electrode voltage is set to molecules. It was a hard choice to make because there is some produce enough energy to drive the cool research going in the labs of professors in this department. reaction, and you can also measure the But I feel that this group is right for me. It will expand on charge consumed by the reaction. By comparing the measured yield to the my previous undergraduate research experience in organic theoretical yield predicted from this and bioanalytical chemistry, and I will be exposed to modern charge using Faraday’s law, you can aspects of electrochemistry, materials science, and biochemistry. estimate the efficiency of the reaction. Careerwise, this experience could lead to opportunities in For the working electrode, the one that catalyzes the reaction, I learned how academia or in the pharmaceutical or specialty chemicals to make nanoparticle-polyion films.1 industries. Also, my new group is active in developing synthetic We were given the 40 nm diameter reactions as well as fundamental studies of redox manganese oxide nanoparticles from a chemistry. This gives me several options for a thesis research collaborating inorganic chemistry group. Then, we adsorbed a layer of polystyrene project. My professor says that I can start by taking some sulfonate onto the carbon cloth, washed training in both areas and we will decide together on a specifi c it, and then adsorbed a layer of a project sometime in the spring. I’ll be hooked up with an cationic polymer. The manganese oxide nanoparticles are negatively charged, advanced grad student or postdoc in each area, and they are so they were adsorbed on the cationic going to train me in the basic techniques. I have decided to polymer layer to make the electrode. record my experiences, and below is my diary. The reaction was to convert styrene to styrene oxide with this manganese oxide nanoparticle electrode.2 I determined Research in Organic This can involve direct attachment of the products with GC and then GC-MS, Electrochemical Synthesis monolayers of molecules to electrode and got my first experience using GC- surfaces, or deposition of a suitable MS. The first time I ran the reaction, I will have my first training session polymer or nanoparticles on an the yield was lousy. But I did it twice in organic electrochemistry soon. Our electrode. I have noticed instruments more and both times the yield of styrene group develops catalytic procedures for in the lab that are used in more oxide was good. My mentor said that we synthetic electrochemical reactions, and conventional organic synthesis, such could also use cyclic voltammetry to help also investigates their mechanisms. I as gas and liquid chromatographs, characterize the system, but we decided have learned that a major function of and UV-visible and Fourier transform to leave that for another time. the group is modification of electrode infrared (FTIR) spectrometers. That is surfaces to make them more selective good because I know how to use some of (continued on next page) for a particular synthetic reaction.

*Ed. Note: The authors of this piece are firmly established professionals; but they enjoyed playing students again to present the diverse and exciting area of organic and biological electrochemistry.

The Electrochemical Society Interface • Spring 2006 59 Electro-organic Reactions... plane pyrolytic graphite electrodes (continued from previous page) containing carboxylate groups were also found effective. So, surface chemistry plays an important role in modern Power Supply o protein electrochemistry. or Potentiostat Also, I found that amperometric H O 2 2 biosensors have been developed with MnO 2 Reference electrode polyions mediators to deliver electrons between carrbboonn e elelcetcrtordoede electrodes and . While this is Counter electrode not direct electron transfer, immobilized voltage applied enzymes in films can have high catalytic 2 activity, and the devices can be driven by gas the mediators. Electrochemical glucose sensors based on this principle with the glucose oxidase are used water routinely by diabetic patients to measure glucose in their blood. I saw one type advertised by the blues guitarist B. B. King on TV last week. Stir bar Working electrode Protein film voltammetry can be done with many types of films. FIG. 1. Typical reactor cell for benchtop electrochemical synthesis. Inset on the right illustrates can be coadsorbed with other chemicals the epoxidation of styrene on a magnesium oxide nanoparticle electrode. to give monolayers with reversible Figure 1 voltammetry on edge plane pyrolytic graphite electrodes. Self-assembled Br Br ��������� monolayers of organothiols on gold ���������� electrodes that adsorb or chemically N N ��� ��������� N bond with proteins can be used to give reversible electrochemistry. Insoluble surfactants and polyions of various types + 5,6 N2 can be used to make stable films. Cyclic voltammetry is a major

FIG. 2. Construction of an electrocatalytic electrode bearing triphenylamine groups. electrochemical tool for studying protein films. A typical setup is shown in Fig. 3. It is the usual cyclic voltammetry Bonding Electrocatalysts to scale by preparing a larger electrode setup using a potentiostat to control Electrode Surfaces for Synthesis using the same surface chemistry. the applied potential and measure the It is several weeks later, and I am I made the electrode in Fig. 2 on a 3 x current in an electrochemical cell. It doing a reaction with a different 3 in. piece of carbon cloth and used it in can be used for all sorts of electroactive catalytic electrode. In addition to what controlled potential electrolysis of benzyl materials; but in this case the protein I have already tried, there are other methyl ether. The cell was similar to the film is attached to the working electrode. ways to modify electrode surfaces. one in Fig. 1. I got a good yield of the I used this setup to obtain cyclic Some ways have the disadvantage expected product benzaldehyde, that I voltammograms for a peroxidase enzyme that the coating is removed readily, measured by gas chromatography. I think from the tuberculosis bacterium in a so that the electrode must be recoated I am getting the hang of this! lipid film. We got the enzyme from a on a regular basis. Two Frenchmen collaborator in Brooklyn. described a good method to beat this Voltammetric Studies of Protein Films These films are simple to problem.3 They found that reduction In preparation for the last phase make. I dissolved some lipid + of phenyldiazonium ion (C6H5N2 ) at a of my training, I was assigned to read (dimyristoylphosphatidyl choline) in graphite surface forms phenyl radicals, several review articles on protein film water and sonicated the dispersion which bind chemically to graphite voltammetry.5,6 I found that protein or to make vesicles. Then I added the electrodes, and that it is possible to enzyme films can be used for biosensors, protein in a neutral buffer, and spread carry out chemical reactions on the aryl bioreactors, and electrochemical a few microliters of the mixture on groups of the modified electrode. These studies of protein redox properties. a pyrolytic graphite disk electrode. ideas were used recently to produce an However, it is tricky, because adsorption This was dried overnight with a small electrocatalytic electrode by attaching of denatured proteins can make a bottle over the electrode to make the a triphenylamine electrocatalyst to a film that blocks electron transfer drying slow. Then the protein-film graphite fiber electrode (Figure 2).4 You on metal electrodes. About 25 years coated working electrode was placed reduce a diazonium ion containing the ago, researchers found that control of into the electrochemical cell, and cyclic triphenylamine nucleus (a well-known electrode surface chemistry was a key voltammetry was done. This involved a electrocatalyst) at a graphite electrode to good protein electron transfer. Early linear scan of voltage to negative values, and then brominate the electrode to success stories included the attachment followed immediately by another linear block undesired reactions at the para of monolayers of 4,4´-bipyridyl or related scan in the positive direction. The results positions. Voltammetry shows that the thiol derivatives to metal electrodes are shown in Fig. 4. This peroxidase is electrode catalyzes the oxidation of to promote direct electron exchange an iron enzyme, and the peaks are substrates such as silanes and benzyl between proteins and the electrodes. caused by reduction of the FeIII enzyme ethers. You can make electrodes for Before this, it was difficult to get direct to the FeII form on the forward scan, catalytic oxidations on a preparative protein voltammetry. A little later, edge followed by oxidation of the FeII enzyme

60 The Electrochemical Society Interface • Spring 2006 Equipment for protein film voltammetry

potentiostat Reduction insulator electrode III material Of Fe

reference Protein film N2 counter inlet working electrode

E-t waveform Cyclic E, V voltammetry Oxidation Electrochemical cell Of FeII time

FIG. 3. System used for cyclic voltammetry of protein films. FIG. 4. Background subtracted cyclic voltammogram at 0.1 V/s of Mycobacterium Tuberculosis catalase peroxidase in a thin film (~0.5 µm) of the lipid dimyristoylphosphatidyl choline on a pyrolytic graphite electrode in pH 6.1 buffer. to FeIII on the reverse scan. The results do fundamental studies, all the rates Editor, p. 170, Academic Press, San Diego tell me that the reaction is reversible, I mentioned above can be measured, (2001). because both peaks are nearly equal in and also all the spectroscopic methods 2. L. Espinal, S. L. Suib, and J, F. Rusling, J. Am. Chem. Soc., 126, 7676 (2004). height and at about the same voltage. We mentioned above can be used. For optical 3. J. Pinson and J.-M. Saveant, J. Am. Chem. know that these peaks are caused by the spectra, the films are usually made on Soc., 119, 201 (1997). enzyme because lots of characterization transparent indium-tin oxide electrodes, 4. B. T. Mayers and A. J. Fry, Organic Lett., 8, studies have been done on these protein- or you can do reflectance. So it seems 411 (2006). lipid films, such as visible absorbance that a lot of information about the redox 5. F. A. Armstrong, H. A. Heering, and J. Hirst, spectroscopy, circular dichroism, chemistry, redox states, and bioactivity Chem. Soc. Rev., 26, 169 (1997). reflectance infrared spectroscopy, of the enzyme can be obtained with 6. J. F. Rusling and Z. Zhang, in Biomolecular Films, J. F Rusling, Editor, p. 1, Marcel spectroelectrochemistry, electron spin these films. Dekker, New York (2003). resonance (ESR) spectroscopy, and other Outcome methods. All these experiments showed About the Authors results consistent with the native protein I have finally finished my initial JAMES F. RUSLING is a professor of chemistry and and not with the free iron heme group or training. I have learned a lot about pharmacology at the University of Connecticut. denatured protein or anything like that. organic and biological electrochemistry His research laboratory in on the Storrs campus. Another method used to make films in the past several months. I also found He is currently the chair of the ECS Organic and Biological Electrochemistry Division and in our lab is layer-by-layer deposition that there is an organization called ECS that holds great research meetings twice Editor for the Americas of Electrochemical of enzymes and polyions of alternate Communications. His research encompasses charges.6 This is the same method I used a year, and there is an Organic and biomolecule electrochemistry, biomedical sensors to make the manganese oxide films, so Biological Electrochemistry Division and arrays, bioreactors, and electrochemistry in it is versatile. Although it is a little more in that Society. Student membership is microemulsions. He may be reached at James. time-consuming than the lipid film cheap; I am definitely going to join! [email protected]. method, the films are usually stable and What did I learn about A. J. FRY is the E. B. Nye Professor of Chemistry in the Chemistry Department, Wesleyan good for biosensors. electrochemical research? In addition University, Middletown, Connecticut and Cyclic voltammetry can be used to to many new electrochemical and Secretary-Treasurer of the ECS Organic and get all sorts of information about the characterization techniques, I have been Biological Electrochemistry Division. His research protein film. The integral under the exposed to modern surface modification involves the anodic behavior of unsaturated silanes, application of quantum chemical peak gives the charge used for protein and film construction methods. Also I learned how electrochemistry is methods to problems in organic electrochemistry, electrolysis, which tells us how much electrocatalysis, and the electrochemical oxidation electroactive enzyme is present in the linked to many other areas like organic and reduction of unsaturated hydrocarbons. He film using Faraday’s law. A scan rate and inorganic chemistry as well as may be reached at [email protected]. study of the separation of the two peaks biology. It also utilizes peripheral as a function of the rate of voltage scan techniques such as chromatography, can be analyzed with theory to obtain all sorts of spectroscopy, and even electron transfer rate constants. Addition quantum mechanical computations and of hydrogen peroxide to the solution voltammetry simulations. Now I need caused the FeIII peak to increase, and the to decide on the specific area that I will FeII peak to disappear. This is because the specialize in for my thesis research, enzyme catalytically reduces hydrogen and dream up a project of my own. peroxide. The rate of this reaction can Everything was so interesting; it is going also be obtained by analysis of the to be a tough decision.  catalytic voltammetry peak. Also, this References type of catalytic current increase can be used in biosensors, in this case a sensor 1. Y. Lvov, in Handbook of Surfaces and Interfaces of Materials, Vol. 3. Nanostructured for hydrogen peroxide. If you want to Materials, Micelles, and Colloids, R.W. Nalwa,

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