Double enzyme‐catalyzed microreactors using capillary electrophoresis

Abstract

This work evaluates the concept of a double enzyme-catalyzed microreactor using capillary electrophoresis (CE). Migrating in a capillary under electrophoresis conditions, plugs of substrate and two enzymes are injected separately in buffer and allowed to react. Extent of reaction and product ratios were subsequently determined by CE. This concept is demonstrated using two model systems: the conversion of adenosine triphosphate (ATP) to adenosine diphosphate (ADP) and adenosine monophosphate (AMP) by hexokinase (HK, EC 2.7.1.1) and apyrase (APY, EC 3.6.1.5), respectively, in the conversion of glucose to glucose-6-phosphate and inorganic phosphate, respectively, and the conversion of nicotinamide adenine dinucleotide, reduced form (NADH), to nicotinamide adenine dinucleotide (NAD) and back to NADH by lactate dehydrogenase (LDH, EC 1.1.1.27) and glucose-6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49), respectively, in the conversion of pyruvate to lactate and glucose-6-phosphate (glc-6-P) to 6-phosphogluconate, respectively. These procedures illustrate the use of the capillary as a double microreactor and the ease of quantitation of reaction products under conditions of electrophoresis.

Response surface examination of the relationship between experimental conditions and product distribution in electrophoretically mediated microanalysis

“…In EMMA, differential electrophoretic mobility is utilized to merge distinct zones of analyte and analytical reagent(s) under the influence of an electric field [15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32]. The reaction is then allowed to proceed within the region of reagent overlap either in the presence or absence of an applied potential, and the resultant product is transported to the detector under the influence of an electric field (Fig.…”

Electrophoretically mediated reaction of glycosidases at a nanoliter scale

“…There are basically two ways of mixing two components in a capillary under electrophoretic conditions, the continuous format [7][8][9][10][11][12][13] and the plug-plug format [12,[14][15][16][17][18]. In contrast to the use of a capillary filled with one of the reactants in the continuous format, the plug-plug format is based on a plug-plug interaction.…”

Electrophoretically mediated microanalysis

“…1A) [6,7,[12][13][14][15][16][17][18][19][20][21]. This type of electrophoretically mixing of the reactants was used in the pioneering work of Bao and Regnier [6].…”