Production of salicylate hydroxylase fromPseudomonas putidaUUC‐1 and its application in the construction of a biosensor

Abstract

The enzyme salicylate hydroxylase was produced from Pseudomonas putida UUC-1 in an 80 dm3 16 h batch fermentation process yielding, -9 K units per 60 dm3. This enzyme preparation has been studied for its application in the construction of biosensor systems, e.g. carbon paste electrodes, screen-printed carbon electrodes, and disposable carbon enzyme electrodes, which will be used for the rapid estimation of salicylate in blood samples. The linear range of the disposable carbon enzyme electrode in response to salicylate was achieved to 1.8 mmol dm-3.

A Polypyrrole-Bienzyme Electrode (Salicylate Hydroxylase-Polyphenol Oxidase) for the Interference-Free Determination of Salicylate

“…The K app M value (5 mM) for salicylate is in good accordance with that reported for the free enzyme (K M ¼ 1.9 -2.6 mM) [9,26,27]. This may illustrate the good permeability of the poly1 host matrix for the diffusion of salicylate and catechol.…”

“…This sensing scheme has already been employed by Dominguez et al [15]. However, the resulting bienzyme carbon paste electrode was less sensitive (51 mA M À1 cm À2 ) than the monoenzyme sensors based on the direct or mediated oxidation of catechol (0.112-6.6 A M À1 cm À2 ) [9,12,13]. This may be due to the coimmobilization of both enzymes into carbon paste that could reduce the accessibility to their active sites.…”

“…This enzyme (SH) catalyzes the hydroxylation of salicylate to catechol in the presence of dioxygen and reduced pyridine nucleotides (NAD(P)H). The majority of the SH-based biosensors involves an amperometric detection based on the direct or mediated oxidation of the enzymically generated catechol [7][8][9][10][11][12][13]. Owing to the bulkiness of reduced pyridine nucleotides, amperometric biosensors have been mainly elaborated by adsorption or covalent immobilization of SH for preserving a good access of substrate and coenzyme to the immobilized enzyme.…”

Microbial Enzymes with Special Characteristics for Biotechnological Applications

“…Pectinases have established their role in the fruit and juice industries [76]. Certain enzymes are specifically required in pharmaceutical industry for diagnostic kits and analytical assays [77,78,79,80]. …”

Electrochemical Biosensors for Salicylate and Its Derivatives

“…Zhou and co-workers [26] studied a feasible preparation of the enzyme salicylate hydroxylase from Pseudomonas putida UUC-1 and its application in salicylate biosensor preparation. They produced this enzyme through the culture of UUC-1 [60], using it in the construction of carbon paste electrodes from graphite with paraf®n oil.…”