Formation of reactive intermediates and metabolites: effects of macrolide antibiotics on cytochrome P-450.
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A method for the analysis of several macrolide and ionophore antibiotics, as well as tiamulin, from soil was developed using pressurized liquid extraction (PLE), reversed-phase liquid chromatography and atmospheric pressure chemical ionisation (APCI) tandem mass spectrometry (LC-APCI+-MS-MS). The analytes were extracted from soil by PLE in 30 min and the extracts were cleaned up by solid-phase extraction (SPE) on a diol SPE cartridge. Liquid chromatographic (LC) separation of the antibiotics was achieved in 35 min. Recovery experiments were performed using spiked soil and concentrations varying from 1 to 2000 microg/kg. By using a macrolide internal standard the recovery rates for the macrolides erythromycin and roxithromycin ranged from 43 to 94% (RSD 20-23%), for the ionophore salinomycin the recovery rate was 76% (RSD 29%), while the pleuromutilin tiamulin was completely recovered. The limits of detection ranged from 0.2 to 1.6 microg/kg. In soil samples a maximum concentration of 0.7 microg/kg tiamulin was found.
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Of four strains of Mycoplasma pneumoniae, highly resistant to erythromycin and related antibiotics, three were homogeneously resistant, but the fourth showed heterogeneous resistance, with only 1% of the cells manifesting this property. Stable, homogeneous resistance was generated in this strain in the presence of erythromycin, whereas the heterogeneous resistance was lost spontaneously on passage in the absence of antibiotics, or on treatment with acridine orange. The mechanism of induction of stable resistance appears to be different from that seen in Staphylococcus aureus.
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