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A sensitive and selective HPLC/ESI-MS/MS assay for the simultaneous quantification of 16-dehydropregnenolone and its major metabolites in rabbit plasma.
J Chromatogr B Analyt Technol Biomed Life Sci. 2006 Jan 02; 830(1):54-63.JC

Abstract

A sensitive, selective and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric assay was developed and validated for the simultaneous quantification of 16-dehydropregnenolone (DHP) and its five metabolites 4,16-pregnadien-3, 20-dione (M(1)), 5-pregnene-3beta-ol-20-one (M(2)), 5-pregnene-3beta, 20-diol (M(3)), 5-pregnene-3beta-ol-16, 17-epoxi-20-one (M(4)) and 5,16-pregnadien-3beta, 11-diol-20-one (M(5)) in rabbit plasma using dexamethasone as internal standard (IS). The analytes were chromatographed on Spheri-5 RP-18 column (5 microm, 100 mm x 4.6 mm i.d.) coupled with guard column using acetonitrile:ammonium acetate buffer (90:10, v/v) as mobile phase at a flow rate of 0.65 ml/min. The quantitation of the analytes was carried out using API 4000 LC-MS-MS system in the multiple reaction monitoring (MRM) mode. The method was validated in terms of linearity, specificity, sensitivity, recovery, accuracy, precision (intra- and inter-assay variation), freeze-thaw, long-term, auto injector and dry residue stability. Linearity in plasma was observed over a concentration range of 1.56-400 ng/ml with a limit of detection (LOD) of 0.78 ng/ml for all analytes except M(3) and M(5) where linearity was over the 3.13-400 ng/ml with LOD of 1.56 ng/ml. The absolute recoveries from plasma were consistent and reproducible over the linearity range for all analytes. The intra- and inter-day accuracy and precision method were within the acceptable limits and the analytes were stable after three freeze-thaw cycles and their dry residues were stable at -60 degrees C for 15 days. The method was successfully applied to determine concentrations of DHP and its putative metabolites in plasma during a pilot pharmacokinetic study in rabbits.

Authors+Show Affiliations

Pharmacokinetics and Metabolism Division, Central Drug Research Institute, Chattar Manzil Palace, Lucknow, India.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Validation Study

Language

eng

PubMed ID

16290073

Citation

Suryawanshi, Satyendra, et al. "A Sensitive and Selective HPLC/ESI-MS/MS Assay for the Simultaneous Quantification of 16-dehydropregnenolone and Its Major Metabolites in Rabbit Plasma." Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences, vol. 830, no. 1, 2006, pp. 54-63.
Suryawanshi S, Singh SK, Gupta RC. A sensitive and selective HPLC/ESI-MS/MS assay for the simultaneous quantification of 16-dehydropregnenolone and its major metabolites in rabbit plasma. J Chromatogr B Analyt Technol Biomed Life Sci. 2006;830(1):54-63.
Suryawanshi, S., Singh, S. K., & Gupta, R. C. (2006). A sensitive and selective HPLC/ESI-MS/MS assay for the simultaneous quantification of 16-dehydropregnenolone and its major metabolites in rabbit plasma. Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences, 830(1), 54-63.
Suryawanshi S, Singh SK, Gupta RC. A Sensitive and Selective HPLC/ESI-MS/MS Assay for the Simultaneous Quantification of 16-dehydropregnenolone and Its Major Metabolites in Rabbit Plasma. J Chromatogr B Analyt Technol Biomed Life Sci. 2006 Jan 2;830(1):54-63. PubMed PMID: 16290073.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A sensitive and selective HPLC/ESI-MS/MS assay for the simultaneous quantification of 16-dehydropregnenolone and its major metabolites in rabbit plasma. AU - Suryawanshi,Satyendra, AU - Singh,S K, AU - Gupta,R C, Y1 - 2005/11/09/ PY - 2005/04/20/received PY - 2005/09/13/revised PY - 2005/10/14/accepted PY - 2005/11/18/pubmed PY - 2006/3/11/medline PY - 2005/11/18/entrez SP - 54 EP - 63 JF - Journal of chromatography. B, Analytical technologies in the biomedical and life sciences JO - J Chromatogr B Analyt Technol Biomed Life Sci VL - 830 IS - 1 N2 - A sensitive, selective and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric assay was developed and validated for the simultaneous quantification of 16-dehydropregnenolone (DHP) and its five metabolites 4,16-pregnadien-3, 20-dione (M(1)), 5-pregnene-3beta-ol-20-one (M(2)), 5-pregnene-3beta, 20-diol (M(3)), 5-pregnene-3beta-ol-16, 17-epoxi-20-one (M(4)) and 5,16-pregnadien-3beta, 11-diol-20-one (M(5)) in rabbit plasma using dexamethasone as internal standard (IS). The analytes were chromatographed on Spheri-5 RP-18 column (5 microm, 100 mm x 4.6 mm i.d.) coupled with guard column using acetonitrile:ammonium acetate buffer (90:10, v/v) as mobile phase at a flow rate of 0.65 ml/min. The quantitation of the analytes was carried out using API 4000 LC-MS-MS system in the multiple reaction monitoring (MRM) mode. The method was validated in terms of linearity, specificity, sensitivity, recovery, accuracy, precision (intra- and inter-assay variation), freeze-thaw, long-term, auto injector and dry residue stability. Linearity in plasma was observed over a concentration range of 1.56-400 ng/ml with a limit of detection (LOD) of 0.78 ng/ml for all analytes except M(3) and M(5) where linearity was over the 3.13-400 ng/ml with LOD of 1.56 ng/ml. The absolute recoveries from plasma were consistent and reproducible over the linearity range for all analytes. The intra- and inter-day accuracy and precision method were within the acceptable limits and the analytes were stable after three freeze-thaw cycles and their dry residues were stable at -60 degrees C for 15 days. The method was successfully applied to determine concentrations of DHP and its putative metabolites in plasma during a pilot pharmacokinetic study in rabbits. SN - 1570-0232 UR - https://www.unboundmedicine.com/medline/citation/16290073/A_sensitive_and_selective_HPLC/ESI_MS/MS_assay_for_the_simultaneous_quantification_of_16_dehydropregnenolone_and_its_major_metabolites_in_rabbit_plasma_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1570-0232(05)00728-2 DB - PRIME DP - Unbound Medicine ER -