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Master's Dissertation
DOI
10.11606/D.59.2007.tde-17082007-114009
Document
Author
Full name
Bruno José Gonçalves da Silva
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2007
Supervisor
Committee
Nassur, Maria Eugenia Queiroz (President)
Augusto, Fabio
Lopes, Norberto Peporine
Title in Portuguese
Avaliação técnica SPME/LC na análise de antidepressivos em amostra de plasma para fins de monitorização terapêutica
Keywords in Portuguese
Antidepressivos
HPLC
in-tube SPME
monitorização terapêutica
SPME
Abstract in Portuguese
As recentes técnicas miniaturizadas de preparo de amostra, microextração em fase sólida (SPME) e in tube SPME, apresentam uma série de vantagens em relação aos métodos clássicos de extração (extração líquido-líquido e extração em fase sólida), tais como: não requer instrumentação analítica sofisticada, utilização de pequenas quantidades de solventes orgânicos, rápido processo operacional, permite automação das análises, a reutilização das fases extratoras, e integra em um único sistema a extração, concentração e introdução da amostra no cromatográfico. Esta dissertação tem como objetivo a padronização, validação e comparação dos métodos SPME/LC-UV com dessorção off line e in tube SPME/LC-UV, para a análise dos antidepressivos da nova geração (mirtazapina, citalopram, paroxetina, duloxetina, fluoxetina e sertralina) em amostras de plasma para fins de monitorização terapêutica. As variáveis: fase extratora, pH da matriz, tempo e temperatura de extração e de dessorção e força iônica apresentaram grande influência na eficiência do processo SPME. O método SPME/LC-UV padronizado, apresentou limite de quantificação (LQ) de 25 a 50 ng mL-1, ampla faixa de linearidade (LQ ? 500 ng mL-1, r2 > 0,9970) e precisão inter ensaios com coeficientes de variação menor que 15% para todos os analitos. Apesar das baixas taxas de recuperação obtidas, de 8,1% (citalopram) a 17,1% (mirtazapina), o método SPME/LC-UV apresentou seletividade e sensibilidade analítica adequada. As variáveis: pH da matriz, fluxo e número de ciclos aspirar/dispensar e volume de amostra apresentaram grande influência na eficiência do processo in tube SPME. A etapa de precipitação de proteínas do plasma, anterior ao processo de extração, foi necessária para a eliminação dos compostos endógenos. O método in tube SPME/LC-UV padronizado apresentou seletividade adequada, precisão inter ensaios com coeficiente de variação menor que 10%, LQ de 20 a 50 ng mL-1, linearidade na faixa de concentração do LQ a 500 ng mL-1, com r2 > 0,9983 para todos os analitos e recuperação absoluta de 5,32% (mirtazapina) a 43,5% (sertralina). A técnica in tube SPME, quando comparada à SPME, permitiu a automação das análises, menor exposição do analista às amostras biológicas e solventes orgânicos, menor tempo de análise e menor volume de amostra de plasma. A eficácia dos métodos, SPME/LC-UV e in tube SPME/LC-UV, foi comprovada através das análises de amostras de plasma de pacientes em terapia com os antidepressivos, para fins de monitorização terapêutica.
Title in English
Evaluation of SPME/LC technique in the antidepressants analysis in plasma sample for ends of therapeutic monitoring
Keywords in English
Antidepressants
HPLC
in-tube SPME
SPME
therapeutic drug monitoring
Abstract in English
The recent miniaturized sample techniques preparation, solid phase microextraction (SPME) and in tube SPME, present several advantages when compared with classic extraction methods (liquid-liquid extraction and solid phase extraction), such as: it does not require sophisticated analytical instrumentation, use small organic solvent amounts, fast operational process, automation of the analyses, reuse extraction phases, and incorporates, into a single procedure, sample extraction, concentration and sample introduction. The aim of this work is development, validation and comparison of methods SPME/LC-UV with off line desorption and in tube SPME/LC-UV, for analyses of antidepressants of the new generation (mirtazapine, citalopram, paroxetine, duloxetine, fluoxetine and sertraline) in plasma samples for therapeutic drug monitoring. Variables: extraction phase, matrix pH, time and temperature of extraction and desorption and ionic strength showed great influence in SPME process efficiency. The method SPME/LC-UV presented limit of quantification (LOQ) variety from 25 to 50 ng mL-1, wide range the of linearity (LOQ 500 ng mL-1, r2 > 0.9970) and interassays precision with coefficient of variation lower than 15% for all analytes. Although the low recovery, from 8.1% (citalopram) to 17.1% (mirtazapine), the method SPME/LC-UV presented adequate selectivity and analytical sensitivity. Variables: matrix pH, flow and number of aspirate/dispense cycles and sample volume showed great influence in the in tube SPME process efficiency. The protein precipitation of the plasma steps, previous to the extraction process, was necessary for the endogenous compounds elimination. The method in tube SPME/LC showed adequate selectivity, interassays precision with coefficient of variation lower than 10%, LOQ variety from 20 to 50 ng mL-1, linearity in range concentration from LOQ to 500 ng mL-1, with r2 > 0.9983 for all analytes and recovery from 5.32% (mirtazapine) to 43.5% (sertraline). The technique in tube SPME, compared with the SPME, permitted the automation of the analyses, minor exposition of the analyst to the biological samples and organic solvent, shorter analyses time and minor plasma sample volume. The effectiveness methods, SPME/LC-UV and in tube SPME/LC-UV, was proven through the analyses of plasma samples of patients in therapy with antidepressants, for therapeutic drug monitoring.
 
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Dissertacao-PDF.pdf (917.47 Kbytes)
Publishing Date
2007-09-10
 
WARNING: The material described below relates to works resulting from this thesis or dissertation. The contents of these works are the author's responsibility.
  • SILVA, Bruno J G. Simultaneous determination of nontricyclic antidepressants in human plasma by Solid-Phase Microextra. Journal of Analytical Toxicolgy, 2007, vol. 31, n. 6, p. 313-320.
  • SILVA, Bruno José Gonçalves, LANçAS, Fernando Mauro, and QUEIROZ, Maria Eugênia Costa. In-tube solid-phase microextraction coupled to liquid chromatography (in-tube SPME/LC) analysis of nontricyclic antidepressants in human plasma [doi:10.1016/j.jchromb.2007.12.006]. Journal of Chromatography B [online], 2008, vol. 862, n. -1, p. 181-188.
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