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Título
HPLC‐QTOF method for quantifying 11‐ketoetiocholanolone, a cortisol metabolite, in ruminants' feces: Optimization and validation
Autor
Facultad/Centro
Área de conocimiento
Título de la revista
Ecology and Evolution
Número de la revista
18
Cita Bibliográfica
Molina-García, L., Pérez, J. M., Sarasa, M., Ureña-Gutiérrez, B., Espinosa, J., & Azorit, C. (2018). HPLC-QTOF method for quantifying 11-ketoetiocholanolone, a cortisol metabolite, in ruminants” feces: Optimization and validation. Ecology and Evolution, 8(18), 9218-9228. https://doi.org/10.1002/ECE3.4285
Editorial
Wiley Open Access
Fecha
2018
ISSN
2045-7758
Resumen
[EN]Studies of animal ecology can benefit from a quantified understanding of eco-physiological
processes and, in particular, of the physiological responses in free-ranging
animals to potential stressors. The determination of fecal cortisol metabolites
as a noninvasive method for monitoring stress has proved to be a powerful tool.
High-performance
liquid chromatography coupled with tandem mass spectrometry
(HPLC-MS/
MS) has emerged as the most accurate method for avoiding problems
related to the nonspecificity of immunoassays. In this study, we optimize and validate
a reliable method using HPLC-MS/
MS for quantifying 11-ketoetiocholanolone
(11-k),
a representative fecal cortisol metabolite in ruminants. An appropriate extraction
and purification procedure was developed taking into account the complex nature of
feces. The final extract obtained was then analyzed with HPLC-MS/
MS using a
quadrupole-time-
of-
fly
(QTOF) tandem mass spectrometer with an electrospray ionization
interface operating in positive mode, which allowed an unequivocal determination
of the metabolite due to its accurate mass capabilities. After rigorous
optimization of both sample extraction and the HPLC-QTOF
parameters, making use
of feces from free-ranging
Iberian ibex, ideal conditions were established. Matrix-matched
standards were used to calibrate the method. The limit of detection and
quantification was 13-and
40-ng/
g, respectively. The validation of the method was
performed with recoveries in the range of 85–110%, a figure much higher than the
60% obtained with the previous extraction methods used in our laboratory, and with
relative standard deviations (RSDs) no higher than 15% for the complete analytical
procedure, including extraction and analysis. The time required for the fecal 11-k
analysis was greatly reduced in comparison with the previous work carried out in our
laboratory. This is the first time that QTOF mass detection coupled with HPLC has
been validated for 11-k
quantification in feces from free-ranging
ruminants such as
Iberian ibex. Given the high selectivity and sensitivity attained, our method could
become a useful tool for noninvasive stress quantification in ruminants.
Materia
Palabras clave
Peer review
SI
ID proyecto
- info:eu-repo/grantAgreement/Junta de Andalucía/Otras actuaciones de fomento de la I+D+I/RNM-118/ES/Biologia De Las Especies Cinegeticas Y Plagas
- info:eu-repo/grantAgreement/Junta de Andalucía/Otras actuaciones de fomento de la I+D+I/ RNM-175/ES/ Biodiversidad Y Desarrollo Sostenible
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