Piezoelectric Electrochemistry Studies On Several Biocatalysis And Chemocatalysis Systems

Bio/chemocatalyst modified electrodes have been widely used in bio/chemosensing, energy science, and electrosynthesis. Reactions using biocatalysts can proceed with higher efficiency and better selectivity under milder conditions than those using chemical catalysts. Bioelectocatalysis is virtually the discipline base of biosensors and biofuel cells (BFCs), and it is thus important to efficiently immobilize enzymes and other biocatalysts on electrodes for bioelectrocatalysis. In addition, the electrochemical techniques feature the three-in-one integration of synthesis, separation and analysis functions, it is thus of great significance to develop new characterization methods with the electrode as a research platform for bio/chemocatalysis systems. In this dissertation, recent advances in quartz crystal microbalance (QCM), enzymatic biosensors, enzymatic BFCs, and enzyme-catalyzed polymerization are briefly reviewed, and detailed studies on several bio/chemocatalysis systems are carried out using many instrumental analysis techniques including electrochemical quartz crystal microbalance (EQCM). The main contents are as follows.1. An electrochemical noise (ECN) device is utilized for the first time to study and characterize a glucose/air membraneless BFC and a monopolar glucose BFC. In the glucose/air membraneless BFC, ferrocene (Fc) and glucose oxidase (GOx) were immobilized on a multiwalled carbon nanotubes (MWCNTs)/Au electrode with a gelatin film at the anode; and laccase (Lac) and an electron mediator,2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonate) diammonium salt (ABTS), were immobilized on a MWCNTs/Au electrode with polypyrrole at the cathode. This BFC was performed in a stirred acetate buffer solution (pH 5.0) containing 40 mM glucose in air, with a maximum power density of 8μW cm-2, an open-circuit cell voltage of 0.29 V, and a short-circuit current density of 85μA cm-2, respectively. The cell current at the load of 100 kQ retained 78.9% of the initial value after continuous discharging for 15 h in a stirred acetate buffer solution (pH 5.0) containing 40 mM glucose in air. The performance decrease of the BFC resulted mainly from the leakage of the ABTS mediator immobilized at the cathode, as revealed by the two-channel quartz crystal microbalance technique. In addition, a monopolar glucose BFC was performed with the same anode as that in the glucose/air membraneless BFC in a stirred phosphate buffer solution (pH 7.0) containing 40 mM glucose, and a carbon cathode in Nafion-membrane-isolated acidic KMnO4, with a maximum power density of 115μW cm-2 an open-circuit cell voltage of 1.24 V, and a short-circuit current density of 202μA cm-2, respectively, which are superior to those of the glucose/air membraneless BFC. A modification of the anode with MWCNTs for the monopolar glucose BFC increased the maximum power density by a factor of 1.8. The ECN device is highly recommended as a convenient, real-time and sensitive technique for BFC studies.2. A simple chemical modification on the popular biocompatible biopolymer chitosan (CS) makes it feasible as an acid-resistant film matrix for biosensing and BFC applications in acidic media. To covalently immobilize Lac from Trametes versicolor that shows its maximum enzymatic activity in acidic solutions, CS was chemically modified with glutaraldehyde (GA) to form GA functionalized CS (GAfCS), which was then allowed to react with Lac to form a Lac-GAfCS composite that is robust in acidic solutions (two-step protocol), as confirmed by QCM and durability tests. The Lac-GAfCS-multiwalled carbon nanotubes (MWCNTs) modified glassy carbon electrode (GCE), Lac-GAfCS-MWCNTs/GCE, exhibited good catalytic activity towards O2 reduction in the presence of 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonate) diammonium salt (ABTS), and the pH-dependent enzymatic activity of the immobilized Lac towards O2 reduction was examined. A glucose/air biofuel cell was fabricated, with the Lac-GAfCS-MWCNTs/GCE and a glucose oxidase (GOx)-GAfCS-MWCNTs/GCE as the biocathode and the bioanode in Nafion-membrane-separated acetate buffer solutions (pH 5.0), respectively. The biofuel cell output a maximum power density of 9.6μW cm-2, an open-circuit cell voltage of 0.19 V, and a short-circuit current density of 114μA cm-2, respectively, as measured with an ECN device. Furthermore, the Lac-GAfCS-MWCNTs/GCE was applied to determine catechol in Britton-Robinson buffer solution (pH 3.0), with a linear range of 0.1-50μM and a limit of detection of 20 nM. In comparison with the direct use of GA for one-pot Lac-GA-CS or Lac-GA crosslinking to immobilize enzyme, the use of macromolecular GAfCS in the proposed two-step protocol was proven to be less harmful to enzymatic activity and thus more suitable for immobilizing Lac to construct the biofuel cell and biosensor.3. Pre-crosslinking enzyme molecules to CS with low-concentration crosslinker and then one-pot electrodeposition of the resultant complex can increase the enzyme load and sensitivities of thus-prepared biosensors (vs. conventional CS electrodeposition). GOx is chosen to examine the proposed protocol in detail. GOx was crosslinked to CS with low-concentration GA (0.08 wt%), and the electroreduction of added H2O2 increased the electrode-surface pH and triggered the electrodeposition of a GOx-GA-CS composite film. The GOx-GA-CS electrodeposition was monitored by an electrochemical quartz crystal microbalance and is theoretically discussed based on an electrogenerated base-to-acid titration model. The prepared first-generation enzyme electrode (CS-GA-GOx/Ptnano/Au) exhibits a current sensitivity as high as 102μA mM-1 cm-2 at 0.70 V vs SCE, being 13 times that of the CS-GOx/Ptnano/Au prepared similarly but without precrosslinking GOx to CS. UV-Vis spectrophotometric determination of the GOx remains in the supernatant liquids after pH-induced CS precipitation suggested a high enzyme load in the GOx-GA-CS film, and amperometric measurements suggested a negligible decrease in the enzymatic activity of GOx after its reaction with the low-concentration GA. Also, the proposed protocol works well for the precrosslinking manner of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide/N-hydroxysulfosuccinimide activation, the water-electroreduction-triggered CS electrodeposition, the second-generation biosensing mode, a 5.0-μm-radius Pt microelectrode, and immobilization of alkaline phosphatase for phenyl phosphate biosensing. Because electrodeposition has been so widely used to immobilize biomacromolecules, and it is always an important topic to increase the load and activity of the immobilized biomacromolecule, the proposed protocol of pretethering the target biomacromolecule to the electrodeposition precusor for immobilization of the biomacromolecule at high load/activity is recommended for wide applications.4. The facile preparation of polymer-enzyme-MWCNTs cast films accompanying in-situ Lac-catalyzed polymerization is described for electrochemical biosensing and biofuel cell applications. Lac-catalyzed polymerization of dopamine (DA) as a new substrate was examined in detail by UV-Vis spectroscopy, cyclic voltammetry (CV), QCM, and scanning electron microscopy. Casting the aqueous mixture of DA, Lac and MWCNTs on a GCE yielded a robust polydopamine (PDA)-Lac-MWCNTs/GCE that can sense hydroquinone (HQ) with 643-μA mM-1 cm-2 sensitivity and 20-nM detection limit (S/N=3). The DA substrate yielded the best biosensing performance, as compared with aniline, o-phenylenediamine or o-aminophenol as the substrate for similar Lac-catalyzed polymerization. Casting the aqueous mixture of DA, GOx, Lac and MWCNTs on a Pt electrode yielded a robust PDA-GOx-Lac-MWCNTs/Pt electrode that exhibits glucose-detection sensitivity of 68.6μA mM-1 cm-2. In addition, ABTS was also co-immobilized to yield a PDA-Lac-MWCNTs-ABTS/GCE that can effectively catalyze the reduction of O2, and it was successfully used as the biocathode of a membraneless glucose/02 BFC in pH 5.0 Britton-Robinson buffer. The proposed biomacromolecule-immobilization platform based on enzyme-catalyzed polymerization may be useful for preparing many other multifunctional polymeric bionanocomposites for wide applications.5. The electropolymerization of L-noradrenaline (NA) after Lac-catalyzed preoxidation to efficiently immobilize GOx at an Au electrode is described for sensitive amperometric biosensing of glucose. The Lac-catalyzed preoxidation of NA was studied by UV-vis spectrophotometry and electrochemical technique. The prepared glucose biosensor displayed a sensitivity of 38μA mM-1 cm-2 and a limit of detection of 0.4μM at 0.7 V vs. SCE, which are obviously better than those prepared via preoxidation-free conventional electropolymerization. The immobilized GOx retained high enzymatic specific activity, as quantified by EQCM and UV-vis spectrophotometry. The proposed Lac-catalyzed preoxidation strategy may have application potential in many fields, such as biosensing, biocatalysis, and BFCs.6. A two-channel EQCM is used to investigate the cyclic voltammetric behavior of two Prussian blue (PB) film-modified Au electrodes in a two-electrode configuration in aqueous solution. The redox peaks observed in the two-electrode cyclic voltammogram are assigned to the intrinsic redox transitions among the Everitt's salt, PB, and Prussian yellow for the film itself, the redox process of the Au substrate and the redox process of small-quantity ferri-/ferrocyanide impurities entrapped in the PB film, as also supported by UV-Vis spectroelectrochemical data. The electrocatalytic reduction of H2O2 was also examined in two-electrode system. The profile of the two-electrode solid-state cyclic voltammogram for the PB powder sandwiched between two gold-coated indium-tin oxide electrodes is similar to that for two PB-modified Au electrodes in aqueous solution, implying similar origins for the corresponding redox peaks. The two-channel EQCM method is expected to become a highly effective technique for the studies of the two-electrode electrochemical behaviors of many other species/materials.7. To investigate the catalytic mechanism of 8-quinolinolato manganese(III) complexes (Q3MnⅢ) epoxidation system, CV and QCM were employed to study catalytic Q3MnⅢ, Q3FeⅢ,5-nitryl-8-quinolinolato MnⅢCl and salen-MnⅢCl complexes. The results showed that the high catalytic efficiency for the epoxidation of olefins with Q3MnⅢcatalysts should be due to their special hexadentate binding structures that could be easily converted to the corresponding pentadentate with pendant hydroxyl groups via opening an axial Mn-O bond in the reaction media. The CV and QCM tests have given us much useful information on the liquid-phase catalysis mechanism, which are highly expected to find wider applications in catalysis science and technology.