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Biomedical subjects

M H Smit

Publications and source records attributed to M H Smit.

6 recordsLinked to original sources
PubMed1993

Toxin detection using a tyrosinase-coupled oxygen electrode.

An enzyme-based "electrochemical canary" is described for the detection of cyanide. The sensing system imitates cyanide's site of toxicity in the mitochondria. The terminal sequence of electron transfer in aerobic respiration is mimicked by mediator coupling of tyrosinase catalysis to an electro-chemical system. An enzyme-coupled oxygen electrode is created which is sensitive to selective poisoning. Biocatalytic reduction of oxygen is promoted by electrochemically supplying tyrosinase with electrons. Thus, ferrocyanide is generated at a cathode and mediates the enzymatic reduction of oxygen to water. An enzyme-dependent reductive current can be monitored which is inhibited by cyanide in a concentration-dependent manner. Oxygen depletion in the reaction layer can be minimized by addressing enzyme activity using a potential pulsing routine. Enzyme activity is electrochemically initiated and terminated and the sensor becomes capable of continuous monitoring. Cyanide poisoning of the biological component is reversible, and it can be reused after rinsing. The resulting sensor detects cyanide based on its biological activity rather than its physical or chemical properties.

Biosensing Techniques↗
PubMed1992

Reagentless enzyme electrode for the determination of manganese through biocatalytic enhancement.

An amperometric enzyme electrode is described for the detection and determination of manganese(II). The biosensor is based on the stimulation by manganese of the aerobic oxidation of substrates by horseradish peroxidase. A mediator, 1,2-naphthoquinone, is used as the substrate and is incorporated with the enzyme into a carbon-paste electrode. The resulting electrode acts as an enzyme-based oxygen sensor, which is sensitive to manganese. Electrochemical control of enzyme activity is achieved through substrate promotion of catalysis. Enzyme modulation by manganese can be switched on and off or adjusted through the appropriate selection of the applied potential. Currents are generated due to the bioelectrocatalytic reduction of oxygen in response to the introduction of manganese sulfate. A sustained current is achieved which is dependent on manganese concentration. Concentrations of 0.5 microM manganese or greater can be measured, and the sensor is reversible, as demonstrated by manganese removal. Biological selectivity for manganese provides a sensor which does not respond to other divalent cations tested, with the possible exception of cobalt. Reagentless, continuous sensing is achieved through substrate cycling.

Catalysis↗
PubMed1990

Cyanide detection using a substrate-regenerating, peroxidase-based biosensor.

An enzyme-based, dual working electrode system is described for the sensing of cyanide. Horseradish peroxidase (HRP) is incorporated as the sensing element. A continuous monitoring of oxidative activity by the enzyme results through the generation and regeneration of substrates at the electrode surfaces. Thus, HRP is oxidized by hydrogen peroxide generated from dissolved oxygen, at the primary electrode, and then reduced through the secondary electrode by mediated electron transfer using ferrocene as a carrier. Ferrocene regeneration at this electrode is proportional to the intrinsic activity of HRP. The dynamics of the system are investigated by using a rotating ring-disk electrode. The enzyme is immobilized to provide better control over its catalytic activity and to increase the lifetime of the biosensor. Cyanide inhibition of current can be modeled by reversible binding kinetics. Detection of cyanide is possible in submicromolar (ppb) concentrations, with a half maximal response at 2 microM. The response time for detection of introduced cyanide is within 1 s. The sensor can be operated between 5 and 40 degrees C, and cyanide inhibition is unaffected by pH changes between 5 and 8. The sensor is reproducible for cyanide determination and is stable for over 6 months.

Biosensing Techniques↗
PubMed1987

Terminal-group oxidation of retinol by mouse epidermis. Inhibition in vitro and in vivo.

Locally applied retinol is metabolized to retinoic acid in mouse epidermis in vivo. To characterize the oxidation system we investigated the ability of soluble extracts of hairless-mouse epidermis to convert retinol and retinal into retinoic acid. The extracts oxidized retinol to retinoic acid in two steps catalysed by two NAD+-dependent enzymes that were resolved on h.p.l.c. The first enzyme catalyses the reversible oxidation of retinol to retinal and is an alcohol dehydrogenase isoenzyme. The second enzyme oxidizes retinal to retinoic acid. Retinol oxidation by epidermal extracts was inhibited by the alcohol dehydrogenase inhibitor 4-methylpyrazole and by the polyene citral. The toxicity and relatively low potency at inhibiting the epidermal alcohol dehydrogenase isoenzyme curtailed the use of 4-methylpyrazole in vivo. However, citral significantly inhibited retinoic acid formation from retinol in the epidermis in vivo. The ability to inhibit the oxidation of retinol to retinoic acid in mouse epidermis provides a potential method to resolve the roles of retinol and retinoic acid in epithelial function.

Acyclic Monoterpenes↗
PubMed1987

The formation of all-trans-retinoic acid from all-trans-retinol in hairless mouse skin.

All-trans-retinoic acid formation from topically applied retinol has been demonstrated in the skin of skh/hr1 (hairless) mice. The all-trans-retinoic acid was identified on the basis of its chromatographic properties on HPLC at various pH values, its photoisomerization to reaction products identical to those formed from authentic all-trans-retinoic acid, and its co-chromatography with methyl retinoate after methylation with diazomethane. Topically applied retinol is about 2-fold less potent at inducing epidermal hyperplasia and 7-fold less potent at inhibiting the induction of epidermal ornithine decarboxylase by phorbol esters than all-trans-retinoic acid in this strain of mice. To elucidate the possible role all-trans-retinoic acid formation from retinol may have in these pharmacological activities, the epidermal and dermal all-trans-retinoic acid levels were compared in mice treated topically with retinol or [11-3H]-all-trans-retinoic acid. The levels of all-trans-retinoic acid found after retinol treatment were several orders of magnitude lower than those found after [11-3H]-all-trans-retinoic acid treatment, and they were insufficient to account for the difference in potencies between all-trans-retinoic acid and retinol. Retinol was eliminated from the epidermis at a rate similar to that of all-trans-retinoic acid after topical administration, but the initial tissue levels achieved were lower. These results suggest that the lower potencies of retinol may simply reflect lower tissue uptake.

Animals↗