Simultaneous determination of multibenzodiazepines by HPLC/UV: Investigation of liquid-liquid and solid-phase extractions in human plasma


Autoria(s): BORGES, Keyller Bastos; FREIRE, Ellen Figueiredo; MARTINS, Isarita; SIQUEIRA, Maria Elisa Pereira Bastos de
Contribuinte(s)

UNIVERSIDADE DE SÃO PAULO

Data(s)

19/10/2012

19/10/2012

2009

Resumo

A method for simultaneous determination of seven benzodiazepines (BZPs) (flunitrazepam, clonazepam, oxazepam, lorazepam, chlordiazepoxide, nordiazepam and diazepam using N-desalkylflurazepam as internal standard) in human plasma using liquid-liquid and solid-phase extractions followed by high-performance liquid chromatography (HPLC) is described. The analytes were separated employing a LC-18 DB column (250 mm x 4.6 mm, 5 mu m) at 35 degrees C under isocratic conditions using 5 mM KH(2)PO(4) buffer solution pH 6.0: methanol: diethyl ether (55:40:5, v/v/v) as mobile phase at a flow rate of 0.8 mL min(-1). UV detection was carried out at 245 nm. Employing LLE, the best conditions were achieved with double extraction of 0.5 mL, plasma using ethyl acetate and Na(2)HPO(4) pH 9.5 for pH adjusting. Employing SPE, the best conditions were achieved with 0.5 mL plasma plus 3 mL 0.1 M borate buffer pH 9.5, which were then passed through a C18 cartridge previously conditioned, washed for 3 times with these solvents: 3 mL 0.1 M borate buffer pH 9.5,4 mL Milli-Q water and 1 mL acetonitrile 5%, finally the BZPs elution was carried with diethyl ether: n-hexane: methanol (50:30:20). In both methods the solvent was evaporated at 40 degrees C under nitrogen flow. The validation parameters obtained in LLE were linearity range of 50-1200 ng mL(-1) plasma (r >= 0.9927), limits of quantification of 50 ng mL(-1) plasma, within-day and between-day CV% and E% for precision and accuracy lower than 15%, and recovery above 65% for all BZPs. In SPE, the parameter obtained were linearity range of 30-1200 ng mL(-1) plasma (r >= 0.9900), limits of quantification of 30 ng mL(-1) plasma, within-day and between-day CV% and E% for precision and accuracy lower than 15% and recovery above 55% for all BZPs. These extracting procedures followed by HPLC analysis showed their suitable applicability in order to examine one or more BZPs in human plasma. Moreover, it could be suggested that these procedures might be employed in various analytical applications, in special for toxicological/forensic analysis. (c) 2008 Elsevier B.V. All rights reserved.

Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)

BIOTA/FAPESP (Rede BIOprospecTA)

Financiadora de Estudos e Projetos (FINEP)

Conselho Nacional cle Desenvolvimento Cientifico e Tecnologico (CNPq)

Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES)

Identificador

TALANTA, v.78, n.1, p.233-241, 2009

0039-9140

http://producao.usp.br/handle/BDPI/20041

10.1016/j.talanta.2008.11.003

http://dx.doi.org/10.1016/j.talanta.2008.11.003

Idioma(s)

eng

Publicador

ELSEVIER SCIENCE BV

Relação

Talanta

Direitos

restrictedAccess

Copyright ELSEVIER SCIENCE BV

Palavras-Chave #Benzodiazepines #Flunitrazepam #Clonazepam #Oxazepam #Lorazepam #Chlordiazepoxide #Nordiazepam #Diazepam #N-Desalkylflurazepam #Human plasma #HPLC #LLE #SPE #CHROMATOGRAPHY-MASS SPECTROMETRY #NITROGEN-PHOSPHORUS DETECTION #COLUMN-SWITCHING TECHNIQUE #GAS-CHROMATOGRAPHY #WHOLE-BLOOD #HUMAN URINE #HUMAN-SERUM #POTENTIOMETRIC DETERMINATION #CAPILLARY-ELECTROPHORESIS #QUANTITATIVE-ANALYSIS #Chemistry, Analytical
Tipo

article

original article

publishedVersion