RESUMO
Quantitation of progesterone (P(4)) in biological fluids is often performed by radioimmunoassay (RIA), whereas liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) has been used much less often. Due to its autoconfirmatory nature, LC-MS/MS greatly minimizes false positives and interference. Herein we report and compare with RIA an optimized LC-MS/MS method for rapid, efficient, and cost-effective quantitation of P(4) in plasma of cattle with no sample derivatization. The quantitation of plasma P(4) released from three nonbiodegradable, commercial, intravaginal P(4)-releasing devices (IPRD) over 192 h in six ovariectomized cows was compared in a pairwise study as a test case. Both techniques showed similar P(4) kinetics (P > 0.05) whereas results of P(4) quantitation by RIA were consistently higher compared with LC-MS/MS (P < 0.05) due to interference and matrix effects. The LC-MS/MS method was validated according to the recommended analytical standards and displayed P(4) limits of detection (LOD) and quantitation (LOQ) of 0.08 and a 0.25 ng/mL, respectively. The high selective LC-MS/MS method proposed herein for P(4) quantitation eliminates the risks associated with radioactive handling; it also requires no sample derivatization, which is a common requirement for LC-MS/MS quantitation of steroid hormones. Its application to multisteroid assays is also viable, and it is envisaged that it may provide a gold standard technique for hormone quantitation in animal reproductive science studies.
Assuntos
Cromatografia Líquida/veterinária , Progesterona/sangue , Espectrometria de Massas em Tandem/veterinária , Animais , Bovinos , Cromatografia Líquida/métodos , Sincronização do Estro/métodos , Feminino , Espectrometria de Massas em Tandem/métodosRESUMO
This paper describes the preparation and characterization of collagen films extracted from swim bladders of three species of tropical fishes: Arius parkeri (Gurijuba), Cynoscion acoupa (Pescada Amarela) and Cynoscion leiarchus (Pescada Branca). Collagen was extracted under acidic conditions (CH(3)COOH, 2.5 pH) and precipitated by the addition of NaCl up to 3.0 mol L(-1). The films were prepared in acrylic containers and dried in a vacuum atmosphere. The collagen films were characterized by hydroxyproline contents, thermal analysis, scanning electron microscopy and impedance spectroscopy. The determined values of 4-hydroxiproline and collagens in the films were: 105.23+/-4.48 and 873.2; 102.94+/-4.42 and 854.1; 100.65+/-4.80 and 835.8 mg g(-1) for A. parkeri, C. acoupa and C. leiarchus, respectively. Differential scanning calorimetry revealed high denaturation temperature peaks at temperatures ranging from 65.9 to 74.8 degrees C. The micrographs showed no fibrillar organization along the material, but spongy structure, with cavity diameters relatively uniform, at around 2 microm. The impedance spectroscopy presented a distributed relaxation process. A. parkeri's films showed piezoelectricity.