Reversed-phase high-performance liquid chromatography coupled to ultraviolet and electrospray time-of-flight mass spectrometry on-line detection for the separation of eight tetracyclines in honey samples.

Eight antibiotics, chlortetracycline, demeclocycline, doxycycline, methacycline, minocycline, oxytetracycline, tetracycline and rolitetracycline, were separated and quantified in Spanish honey extracts of different floral origin using a commercial RP-C18 HPLC column and two different on-line detectors (diode array and electrospray time-of-flight mass spectrometry (ESI-TOF-MS) systems). Operating a linear gradient at a flow of 2 ml min(-1) the HPLC separation of the eight antibiotics was obtained within 10 min with good peak symmetry and an acceptable resolution (2.1) for the critical band pair rolitetracycline and oxytetracycline. Values of the numbers of theoretical plates (N) were comprised between 2328 and 19448 while the limits of detection in honey were within 0.02-1.03 microg kg(-1) in the case of UV detection and 0.05-0.76 microg kg(-1) for ESI-TOF-MS detection (operating in negative mode). A recovery study was carried out by preparing some quality control samples at four levels of concentration (10, 25, 50 and 100 microg kg(-1)) and percentages between 72% and 98% were attained.

Evaluation of amperometric detection for the liquid-chromatographic determination of tetracycline antibiotics and their common contaminants in pharmaceutical formulations.

A sensitive high-performance liquid chromatographic technique with amperometric detection has been developed for the determination of seven commercially used tetracyclines in bulk powders and pharmaceutical preparations. The technique is based on the oxidation of these compounds and their contaminants at the glassy carbon electrode. The extraction procedures are simple and the HPLC conditions separate the tetracyclines from their major degradation products. The chromatography was performed using a commercially available octadecylsilane column, with a mobile phase: KH2 PO4 (pH = 2.5; 0.05 M) - acetonitrile (84:16, v/v) and detection at 1.2 V. The technique permits the simultaneous determination of trace amounts of chlortetracycline, demeclocycline, doxycycline, methacycline, minocycline, oxytetracycline and tetracycline as well as the separation of their common impurities (epi, anhydro and epianhydro contaminants) with detection limits of 0.1-1.0 ng microl(-1) and recoveries of 99.1-100.4%. No interference was observed from the commonly present excipients in pharmaceutical formulations.

Isolation of doxycycline, 6-epidoxycycline and 2-acetyl-2- decarboxamidometacycline from commercial metacycline by preparative column liquid chromatography on silica gel.

Isolation of doxycycline, 6-epidoxycycline and 2-acetyl-2-decaboxamidometacycline from commercial metacycline was achieved by preparative column liquid chromatography on silica gel, previously impregnated with edetate (EDTA). Careful control of the pH of EDTA allowed fine tuning of the separation. The mobile phases were composed of dichloromethane, methanol and .1 mM EDTA at pH 9.0 or 6.0. Structures were confirmed with nuclear magnetic resonance spectroscopy. The presence of doxycycline and its 6-epimer in commercial metacycline has not previously been described. The presence of the 2-acetyl derivative was not surprising since analogous 2-acetyl derivatives have been identified in other tetracyclines.