ABSTRACT
In order to obtain an active and stable oxidation reactor for daily use in biochemical laboratory we decided to immobilize galactose oxidase orientedly through a carbohydrate chain to the magnetic carriers. We used hydrazide derivatives of non-magnetic and magnetic bead cellulose and of magnetic and non-magnetic poly(HEMA-co-EDMA) microspheres. Activation of the enzyme molecules was done by sodium periodate in the presence of supplements (fucose, CuSO4, catalase). Orientedly immobilized galactose oxidase presents high storage stability and lower susceptibility to inappropriate microenvironmental conditions. Reactor reactivated by three pulses of D-galactose retained practically 100% of its native activity after 6 months. The positive properties of both magnetic carriers were entirely confirmed.
Subject(s)
Cellulose/chemistry , Enzymes, Immobilized/chemistry , Galactose Oxidase/chemistry , Methacrylates/chemistry , Animals , Immunoglobulin G/chemistry , Magnetics , Microspheres , Nuclear Magnetic Resonance, Biomolecular , Periodic Acid/chemistry , Polyhydroxyethyl Methacrylate/chemistry , SwineABSTRACT
Hydrogen sulphide is produced in human large intestine by anaerobic fermentation and may play a pathogenic role. An analytical method for determination of sulphide in whole blood using an extractive alkylation technique was optimised and validated for this purpose. The sample was mixed with organic phase containing pentafluorobenzyl bromide as an alkylating agent. The benzalkonium chloride was used as a phase-transfer catalyst. The quantitative determination was performed using GC-MS technique in selected ion monitoring mode. The blood levels of sulphide of healthy controls were measured (35-80 microM/l). The method is versatile, reproducible (RSD=2.7%) and suitable for research of anaerobic fermentation in vivo.
