In order to improve the stability of printed biosensors, chemical bonding of mediators and enzymes to MgO-templated carbons (GMgOC) with a polymethacrylate glycidyl group has been investigated.1, 2) It has been reported that the binding of amino groups of mediators and enzymes to the glycidyl group significantly inhibits elution. On the other hand, the chemical bond between the glycidyl group and the mediator is not so strong depending on the solution pH. In this study, N-hydroxysuccinimide (NHS)-grafted porous carbons were prepared as a new immobilization method, and sensors using NHS-grafted porous carbons were fabricated and their response and stability were evaluated.NHSMA (N-Succinimidyl Methacrylate) was grafted onto the surface of MgO-templated carbons (MgOC) to enable the immobilization of amino group-containing mediators and enzymes by chemical bonding. This carbon is referred to as NHS-GMgOC in the following. Electrodes were prepared by screen-printing NHS-GMgOC ink as the working electrode, Ag/AgCl ink as the reference electrode, and carbon ink as the counter electrode on a polyimide substrate. The electrodes were modified by dropping the mediator thionine and the enzyme FAD-derived glucose dehydrogenase (FAD-GDH) on the working electrode. Concentration dependence and long-time stability were evaluated by chronoamperometry. A 0.1 mol dm-3 phosphate buffer (pH 7) containing 0.1-25 mmol dm-3 glucose was used as the measurement solution.Figure 1 shows the graft reaction mechanism. Electron irradiation of the MgOC surface in an argon atmosphere cleaves the C-H bonds on the carbon surface and generates carbon radicals. When NHSMA, a monomer with a carbon double bond, comes into contact with MgOC, the radicals on the MgOC surface react with the double bond of NHSMA, and radicals are also generated on the reacted NHSMA. This chain of reactions leads to the introduction of NHS esters on the carbon surface.Glucose biosensors were prepared by three different preparation methods, and their long-time stability was investigated (Fig. 2). NHS-GMgOC was prepared by washing carbon fixed with thionine by adding it to a thionine solution after preparation, and then inked (mixing method). The glucose sensor using the mixing method showed a current density of more than 1 mA cm-2 even after 60000 s. On the other hand, when the NHS-GMgOC was inked and the thionine solution was dropped after printing (casting method), almost no current density was observed after 60000 s. These results suggest that it is important for the immobilization of the mediator to allow the reaction with thionine to proceed sufficiently immediately after the preparation of NHS-GMgOC. When a chitosan membrane was added to the glucose sensor using the mixing method, the current value increased to 5 mA cm-2. This was attributed to the inhibition of FAD-GDH elution by the chitosan membrane.1) I. Shitanda et al., Bull. Chem. Soc. Jpn., 93 (2020) 32.2) R. Suzuki et al., Journal of Power Sources, 479 (2020) 228807.ACKNOWLEDGMENTSThis research was supported by AMED (Grant Number JP22be1004205). Figure 1
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