Development of a HPLC method for the determination of losartan urinary metabolic ratio to be used for the determination of CYP2C9 hydroxylation phenotypes
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Pedro Dorado
and
CEIBA.FP Consortium of the Ibero-American Network of Pharmacogenomics RIBEF
Abstract
Background: Losartan is metabolized to losartan carboxylic acid (E-3174) by the polymorphic cytochrome CYP2C9. The aim of the study was to develop a high-performance liquid chromatographic (HPLC) method with fluorescence detection for simultaneously measuring losartan and its metabolite E-3174 in urine to evaluate the losartan urinary metabolic ratio (MR: losartan/E-3174) for CYP2C9 phenotyping in humans.
Methods: The compounds were separated in a reversed-phase chromatographic column and detected by fluorescence at a wavelength of 250 nm for excitation and of 370 nm for emission.
Results: No analytical interferences with endogenous compounds were found, and the extraction recoveries were over 88%. Limits of quantification of 2 ng mL–1 for losartan and 5 ng mL–1 for E-3174 were achieved, as well as good reproducibility with coefficients of variation of <9% in all cases. Analyses with the present HPLC method show significant differences (p<0.05) in losartan MRs between the four CYP2C9 genotype groups in 13 Spanish healthy volunteers.
Conclusions: The method developed is simple and affordable, as well as sensitive and reliable to calculate the MR. Therefore, it appears to be useful for CYP2C9 phenotyping using losartan as a drug test in populations, such as Hispanics with different allele combinations.
This study was partially supported by the Institute of Health Carlos III-FIS and the European Union (FEDER) Grants PI10/02010, PI10/02758, and CP06/00030 (PD); Gobierno de Extremadura, Consejería de Empleo, Empresa e Innovación and the European Union (Fondo Social Europeo) Grant PD10199 (MEGN); and Consejería de Sanidad y Dependencia PRIS10043; and Gobierno de Extremadura AEXCID Cooperación Extremeña (11IA002) to the SIFF-RIBEF network (Red Iberoamericana de Farmacogenética y Farmacogenómica; www.ribef.com).
The following members listed by country in alphabetical order (*Group Coordinator), constitute the “CEIBA.FP (Consorcio Europeo Ibero-Americano de Farmacogenética de Poblaciones)/RIBEF (Red Iberoamericana de Farmacogenética)”; “CEIBA.FP (European Ibero American Consortium for Population Pharmacogenetics)/RIBEF (Ibero Latino American Network of Pharmacogenetics)”:
Group 01: Authors: Graciela E. Moya*, Verónica Ferreiro. Institutions: UCA Pontificia Universidad Católica Argentina and Fundación GENOS, Buenos Aires, Argentina.
Group 02: Author: Eduardo Tarazona-Santos*. Institution: UFMG Universidade Federal de Minas Gerais, Belo Horizonte, Brasil.
Group 03: Authors: Jenny Fiedler* (1), Luisa Herrera (1), Romina Rojas-Ponce (2). Institutions: UCH Universidad de Chile, Santiago (1) and UCON Universidad de Concepción, Concepción, Chile (2).
Group 04: Authors: Alba P. Sarmiento*, Angélica Borbón. Institution: PUJ Pontificia Universidad Javeriana, Bogotá, Colombia.
Group 05: Authors: Ramiro Barrantes*, Gerardo Jiménez-Arce, Carolina Céspedes. Institution: UCR Universidad de Costa Rica, San José, Costa Rica.
Group 06: Authors: Idania Rodeiro* (1), Mayra Álvarez (2), Bárbaro Pérez (2), Luis R. Calzadilla (3), René Delgado (4), Diadelis Remirez (5). Institutions: CEBIMAR (1); Facultad de Medicina Calixto García (2); Centro Comunitario de Salud Mental La Habana Vieja (3); Centro para las Investigaciones y Desarrollo de los Medicamentos, CIDEM (4); Centro para el Control Estatal de la Calidad de los Medicamentos, Equipos y Dispositivos Médicos, CEMED (5), La Habana, Cuba.
Group 07: Authors: Enrique Terán* (1,2), Natalia Heras (2), Leonardo J. Beltrán (2), Francisco Hernández (3). Institutions: USFQ Universidad San Francisco de Quito (1), UCE Universidad Central de Ecuador, Quito (2) and UG Universidad Estatal de Guayaquil (3), Ecuador.
Group 08: Authors: Rocío Ortiz-López*, Augusto Rojas-Martínez, Lourdes Garza-Ocañas, Yadira X. Pérez-Páramo. Institution: UANL Universidad Autónoma de Nuevo León, Monterrey, Nuevo León, México.
Group 09: Authors: Marisol López-López* (1), Alberto Ortega-Vázquez (1), Elisa Alonso-Vilatela (2), Nancy Monroy-Jaramillo (2), Teresa Corona Vázquez (2), Petra Yescas-Gómez (2), Adriana Ochoa-Morales (2). Institutions: UAM Universidad Autónoma Metropolitana (1) and Instituto Nacional de Neurología y Neurocirugía Manuel Velasco Suárez (2), México DF, México.
Group 10: Authors: Martha G. Sosa-Macías*, Carlos Galaviz-Hernández, Ismael Lares-Aseff; Blanca Lazalde. Institution: IPN Instituto Politécnico Nacional-CIIDIR, Durango, México
Group 11: Author: Ronald Ramirez-Roa*. Institution: UNAN Universidad Nacional Autónoma de Nicaragua, León, Nicaragua.
Group 12: Authors: Manuela Grazina*. Institution: UC.PT University of Coimbra, Portugal.
Group 13: Authors: Adrián LLerena*, Pedro Dorado, Eva M. Peñas-Lledó, Jesús Cobaleda, M. Eugenia G. Naranjo, Fernando de Andrés. Institutions: UEX University of Extremadura, Badajoz, Spain.
Group 14: Authors: Francisco E. Estévez-Carrizo*, Nicolás González-Vacarezza. Institution: UM Universidad de Montevideo, Uruguay.
Conflict of interest statement
Authors’ conflict of interest disclosure: The authors stated that there are no conflicts of interest regarding the publication of this article.
Research funding: None declared.
Employment or leadership: None declared.
Honorarium: None declared.
References
1 Timmermans PB, Wong PC, Chiu AT, Herblin WF, Benfield P, Carini DJ, et al. Angiotensin II receptors and angiotensin II receptor antagonists. Pharmacol Rev 1993;45:205–51.Search in Google Scholar
2 Stearns RA, Chakravarty PK, Chen R, Chiu SH. Biotransformation of losartan to its active carboxylic acid metabolite in human liver microsomes: role of cytochrome P4502C and 3A subfamily members. Drug Metab Dispos 1995;23:207–15.Search in Google Scholar
3 Yun CH, Lee HS, Lee H, Rho JK, Jeong HG, Guengerich FP. Oxidation of the angiotensin II receptor antagonist losartan (DuP 753) in human liver microsomes: role of cytochrome P4503A(4) in formation of the active metabolite EXP3174. Drug Metab Dispos 1995;23:285–9.Search in Google Scholar
4 Yasar Ü, Tybring G, Hidestrand M, Oscarson M, Ingelman-Sundberg M, Dahl ML, et al. The role of CYP2C9 polymorphism in losartan oxidation. Drug Metab Dispos 2001;29:1051–6.Search in Google Scholar
5 Miners JO, Birkett DJ. Cytochrome P4502C9: an enzyme of major importance in human drug metabolism. Br J Clin Pharmacol 1998;45:525–38.10.1046/j.1365-2125.1998.00721.xSearch in Google Scholar
6 Yasar U, Forslund-Bergengren C, Tybring G, Dorado P, Llerena A, Sjöqvist F, et al. Pharmacokinetics of losartan and its metabolite E-3174 in relation to the CYP2C9 genotype. Clin Pharmacol Ther 2002;71:89–98.10.1067/mcp.2002.121216Search in Google Scholar
7 Dorado P, Berecz R, Norberto MJ, Yasar U, Dahl ML, LLerena A. CYP2C9 genotypes and diclofenac metabolism in Spanish healthy volunteers. Eur J Clin Pharmacol 2003;59:221–5.10.1007/s00228-003-0588-0Search in Google Scholar
8 LLerena A, Dorado P, Berecz R, González AP, Peñas-LLedó EM. Effect of CYP2D6 and CYP2C9 genotypes on fluoxetine and norfluoxetine plasma concentrations during steady-state conditions. Eur J Clin Pharmacol 2004;59:869–73.10.1007/s00228-003-0707-ySearch in Google Scholar
9 Sullivan-Klose TH, Ghanayem BI, Bell DA, Zhang ZY, Kaminsky LS, Shenfield GM, et al. The role of the CYP2C9-Leu359 allelic variant in the tolbutamide polymorphism. Pharmacogenetics 1996;6:341–9.10.1097/00008571-199608000-00007Search in Google Scholar
10 van der Weide J, Steijns LS, van Weelden MJ, de Haan K. The effect of genetic polymorphism of cytochrome P450 CYP2C9 on phenytoin dose requirement. Pharmacogenetics 2001;11:287–91.10.1097/00008571-200106000-00002Search in Google Scholar
11 Odani A, Hashimoto Y, Otsuki Y, Uwai Y, Hattori H, Furusho K, et al. Genetic polymorphism of the CYP2C subfamily and its effect on the pharmacokinetics of phenytoin in Japanese patients with epilepsy. Clin Pharmacol Ther 1997;62:287–92.10.1016/S0009-9236(97)90031-XSearch in Google Scholar
12 Aithal GP, Day CP, Kesteven PJ, Daly AK. Association of polymorphisms in the cytochrome P450 CYP2C9 with warfarin dose requirement and risk of bleeding complications. Lancet 1999;353:717–9.10.1016/S0140-6736(98)04474-2Search in Google Scholar
13 The Human Cytochrome P450 (CYP) Allele Nomenclature Committee. Available at: www.CYPalleles.ki.se. Accessed March 19, 2012.Search in Google Scholar
14 Romkes M, Faletto MB, Blaisdell JA, Raucy JL, Goldstein JA. Cloning and expression of complementary DNAs for multiple members of the human cytochrome P450IIC subfamily [published erratum appears in Biochemistry 1993;32:1390]. Biochemistry 1991;30:3247–55.10.1021/bi00227a012Search in Google Scholar PubMed
15 Rettie AE, Wienkers LC, Gonzalez FJ, Trager WF, Korzekwa KR. Impaired (S)-warfarin metabolism catalysed by the R144C allelic variant of CYP2C9. Pharmacogenetics 1994;4:39–42.10.1097/00008571-199402000-00005Search in Google Scholar PubMed
16 Rettie AE, Haining RL, Bajpai M, Levy RH. A common genetic basis for idiosyncratic toxicity of warfarin and phenytoin. Epilepsy Res 1999;35:253–5.10.1016/S0920-1211(99)00017-0Search in Google Scholar
17 Ninomiya H, Mamiya K, Matsuo S, Ieiri I, Higuchi S, Tashiro N. Genetic polymorphism of the CYP2C subfamily and excessive serum phenytoin concentration with central nervous system intoxication. Ther Drug Monit 2000;22:230–2.10.1097/00007691-200004000-00016Search in Google Scholar
18 del Rosario Brunetto M, Contreras Y, Clavijo S, Torres D, Delgado Y, Ovalles F, et al. Determination of losartan, telmisartan, and valsartan by direct injection of human urine into a column-switching liquid chromatographic system with fluorescence detection. J Pharm Biomed Anal 2009;50:194–9.10.1016/j.jpba.2009.04.015Search in Google Scholar
19 Zhang M, Wei F, Zhang YF, Nie J, Feng YQ. Novel polymer monolith microextraction using a polymethacrylic acid-ethylene glycol dimethacrylate) monolith and its application to simultaneous analysis of several angiotensin II receptor antagonists in human urine by capillary zone electrophoresis. J Chrom A 2006;1102:294–301.10.1016/j.chroma.2005.10.057Search in Google Scholar
20 Yeung PKF, Jamieson, A, Smith GJ, Fice D, Pollak PT. Determination of plasma concentrations of losartan in patients by HPLC using solid phase extraction and UV detection. Int J Pharm 2000;204:17–22.10.1016/S0378-5173(00)00453-1Search in Google Scholar
21 Polinko M, Riffel K, Song H, Lo MW. Simultaneous determination of losartan and EXP3174 in human plasma and urine utilizing liquid chromatography/tandem mass spectrometry. J Pharm Biomed Anal 2003;33:73–84.10.1016/S0731-7085(03)00348-0Search in Google Scholar
22 Shah HJ, Kundlik ML, Patel NK, Subbaiah G, Patel DM, Suhagia BN, et al. Rapid determination of losartan and losartan acid in human plasma by multiplexed LC-MS/MS. J Sep Sci 2009;32:3388–94.10.1002/jssc.200900287Search in Google Scholar
23 Ritter MA, Furtek CI, Lo M-W. An improved method for the simultaneous determination of losartan and its major metabolite, EXP3174, in human plasma and urine by high performance liquid chromatography with fluorescence detection. J Pharm Biomed Anal 1997;15:1021–9.10.1016/S0731-7085(96)01948-6Search in Google Scholar
24 Iwasa T, Takano T, Hara K, Kamei T. Method for the simultaneous determination of losartan and its major metabolite, EXP-3174, in human plasma by liquid chromatography-electrospray ionization tandem mass spectrometry. J Chromatogr B Biomed Sci Appl 1999;734:325–30.10.1016/S0378-4347(99)00358-8Search in Google Scholar
25 Llerena A, Dorado P, Ramírez R, González I, Alvarez M, Peñas-Lledó EM, Pérez B, Calzadilla LR. CYP2D6 genotype and debrisoquine hydroxylation phenotype in Cubans and Nicaraguans. Pharmacogenomics J 2012;12:176–83.10.1038/tpj.2010.85Search in Google Scholar PubMed
26 Dorado P, Heras N, Machín E, Hernández F, Teran E, Llerena A. CYP2D6 genotype and dextromethorphan hydroxylation phenotype in an Ecuadorian population. Eur J Clin Pharmacol 2012;68:637–4410.1007/s00228-011-1147-8Search in Google Scholar PubMed
27 Sosa-Macías M, Dorado P, Alanis-Bañuelos RE, Llerena A, Lares-Asseff I. Influence of CYP2D6 deletion, multiplication, -1584C–>G, 31G–>A and 2988G–>a gene polymorphisms on dextromethorphan metabolism among Mexican tepehuanos and mestizos. Pharmacology 2010;86:30–6.10.1159/000314334Search in Google Scholar PubMed
28 Dorado P, Sosa-Macias MG, Peñas-Lledó EM, Alanis-Bañuelos RE, Wong ML, Licinio J, et al. CYP2C9 allele frequency differences between populations of Mexican-Mestizo, Mexican-Tepehuano, and Spaniards. Pharmacogenomics J 2011;11:108–12.10.1038/tpj.2010.29Search in Google Scholar PubMed
©2012 by Walter de Gruyter Berlin Boston
Articles in the same Issue
- Masthead
- Masthead
- Editorial
- Maintain the goals of Drug Metabolism and Drug Interactions
- Reviews
- Cytochrome P450 2B6: function, genetics, and clinical relevance
- In silico pharmacology for a multidisciplinary drug discovery process
- Original Articles
- In vitro effects of three antidepressant drugs on plasma paraoxonase activity
- Development of a HPLC method for the determination of losartan urinary metabolic ratio to be used for the determination of CYP2C9 hydroxylation phenotypes
- Interaction of methocarbamol and yeast sucrase induces enzyme inhibition
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Articles in the same Issue
- Masthead
- Masthead
- Editorial
- Maintain the goals of Drug Metabolism and Drug Interactions
- Reviews
- Cytochrome P450 2B6: function, genetics, and clinical relevance
- In silico pharmacology for a multidisciplinary drug discovery process
- Original Articles
- In vitro effects of three antidepressant drugs on plasma paraoxonase activity
- Development of a HPLC method for the determination of losartan urinary metabolic ratio to be used for the determination of CYP2C9 hydroxylation phenotypes
- Interaction of methocarbamol and yeast sucrase induces enzyme inhibition
- Evaluation of the effect of statin use on the acenocoumarol and phenprocoumon maintenance dose
- Exploring structurally conservative blocks as universal templates for modeling eukaryotic cytochrome P450s
