Cross-Linked LDH Crystals for Lactate Synthesis Coupled to Electroenzymatic Regeneration of NADH

Abstract

Lactate dehydrogenase (LDH) was crystallized from concentrated ammonium sulfate solution and cross-linked with glutaraldehyde to afford long-lived enzymatically active cross-linked crystals (LDH-CLC). The crystals were employed in an electrolytic cell for lactate production from pyruvate, in which the cathode consisted of a carbon electrode containing a coating of lipoamide dehydrogenase (LiDH) immobilized with methyl viologen under a Nafion membrane. This cell was more effective than a similar cell containing LDH in soluble form. An even greater improvement in performance was achieved by chemically binding a viologen derivative to the LiDH and using an electrode based on this modified enzyme in a cell containing LDH-CLC. The activity of the LDH-CLC is much less sensitive to pH than that of the soluble enzyme.

Electrochemical reduction of NAD+ on a polycrystalline gold electrode

“…Electrochemical systems using chemically modified [13][14][15][16][17][18] and enzyme-mediated electrodes [19][20][21][22][23][24] have been proposed, and the yield in the active 1,4-NADH produced ranged from ca. 75% on a cholesterolmodified gold-amalgam electrode [13] to ca.…”

Direct regeneration of NADH on a ruthenium modified glassy carbon electrode

“…However, in these two papers [9,10] the authors did not report the yield of the active NADH. Fry et al [11][12][13] have published a series of papers on the reduction of NAD + to NADH on glassy carbon and vitreous carbon electrodes modified by a layer of immobilized methyl viologen and lipoamide dehydrogenase. They showed that the enzyme-catalyzed reduction of NAD + on such modified electrodes results in the production of enzymatically active NADH, but the amount of active NADH produced was not explicitly reported.…”

A kinetic study of NAD+ reduction on a ruthenium modified glassy carbon electrode

“…However, they did not report the yield of active NADH. Fry et al [7][8][9] have published a series of papers on the reduction of NAD þ to NADH on glassy carbon and vitreous carbon electrodes modified by a layer of immobilized methyl viologen and lipoamide dehydrogenase. They showed that the enzyme catalyzed reduction of NAD þ on such modified electrodes results in the production of enzymatically active NADH, but the amount of active NADH produced was not explicitly reported.…”