Escrutinio optimizado de amplio espectro cualitativo y alternativo de aplicación forense por HPLC/ DAD para drogas y medicamentos
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Fecha
2011-10
Autores
Barrantes Oconitrillo, Elisa M.
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Editor
Universidad Nacional (Costa Rica)
Resumen
Con el desarrollo de este trabajo, se implementó un método de escrutinio de amplio espectro de drogas y medicamentos, utilizando cromatografía líquida de alta resolución (HPLC), con detector de arreglo de diodos (DAD), el cual fue aplicado a extractos obtenidos con las técnicas líquido — líquido (LLE) y fase sólida (SPE).
La columna utilizada fue una Discovery RP-Amida C16, de 25 cm de longitud, 3 mm de diámetro interno y un tamaño de partícula de 5 pm. El método desarrollado se basó en el uso de metanol y amortiguador de fosfato (25 mM) a un pH de 3,0; con gradiente lineal con intervalos isocráticos en la composición de la fase móvil y gradientes de flujo, utilizando como modificador de par iónico el octilsulfonato de sodio (2 mM) a 35 oc.
A partir de la metodología desarrollada, con disoluciones de materiales de referencia, se generaron tablas de índices de retención con respecto al estándar interno utilizado (acetanilida 25 pg / mL). Se creó una biblioteca con los 79 espectros UV / Vis de las diferentes drogas y medicamentos obtenidos en cada cromatograma, para ayudar en la identificación futura en el análisis de muestras.
En cuanto a la optimización cromatográfica y el análisis de robustez de la metodología, se investigó el efecto de cuatro factores: el uso de dos modificadores de par iónico, el uso de un aditivo amínico como modificador de tensión superficial, el pH del amortiguador utilizado en la fase móvil y la temperatura del horno. Se utilizó un diseño factorial completo a dos niveles y cuatro factores, con tres réplicas cada uno. Los efectos del pH y el uso del modificador amínico resultaron altamente significativos, a un 95 % de confianza.
La mayoría de sustancias estudiadas por cromatografía líquida, se analizaron por cromatografía de gases (GC) con detector de masas (MS) de impacto electrónico (IE), esto con el fin de analizar la capacidad de detección de ambas técnicas.
Con el desarrollo de ambas metodologías se dotó al Departamento de Ciencias Forenses de herramientas analíticas para el escrutinio de drogas y medicamentos. Debido a la gran variedad de polaridades presentes en los analitos, incluso de una misma familia o grupo farmacológico, ambas técnicas representan un complemento analítico. Todo lo anterior en pro de la mejora en el sistema judicial costarricense.
With the development of this work, a broad-spectrum drug and medication screening method was implemented, using high-performance liquid chromatography (HPLC), with a diode array detector (DAD), which was applied to extracts obtained with the liquid — liquid (LLE) and solid phase (SPE) techniques. The column used was a Discovery RP-Amida C16, 25 cm long, 3 mm internal diameter and a particle size of 5 pm. The developed method was based on the use of methanol and phosphate buffer (25 mM) at a pH of 3.0; with linear gradient with isocratic intervals in the composition of the mobile phase and flow gradients, using sodium octylsulfonate (2 mM) as ion-pair modifier at 35 oC. Based on the methodology developed, with solutions of reference materials, retention index tables were generated with respect to the internal standard used (acetanilide 25 pg/mL). A library with the 79 UV/Vis spectra of the different drugs and medicines obtained in each chromatogram was created, to help in future identification in sample analysis. Regarding the chromatographic optimization and the robustness analysis of the methodology, the effect of four factors was investigated: the use of two ion pair modifiers, the use of an amine additive as a surface tension modifier, the pH of the buffer used in mobile phase and oven temperature. A complete factorial design was used at two levels and four factors, with three replicates each. The effects of pH and the use of the amine modifier were highly significant, at 95% confidence. Most of the substances studied by liquid chromatography were analyzed by gas chromatography (GC) with an electronic impact (IE) mass detector (MS), in order to analyze the detection capacity of both techniques. With the development of both methodologies, the Department of Forensic Sciences was provided with analytical tools for the scrutiny of drugs and medicines. Due to the great variety of polarities present in the analytes, even from the same family or pharmacological group, both techniques represent an analytical complement. All of the above in favor of improving the Costa Rican judicial system.
With the development of this work, a broad-spectrum drug and medication screening method was implemented, using high-performance liquid chromatography (HPLC), with a diode array detector (DAD), which was applied to extracts obtained with the liquid — liquid (LLE) and solid phase (SPE) techniques. The column used was a Discovery RP-Amida C16, 25 cm long, 3 mm internal diameter and a particle size of 5 pm. The developed method was based on the use of methanol and phosphate buffer (25 mM) at a pH of 3.0; with linear gradient with isocratic intervals in the composition of the mobile phase and flow gradients, using sodium octylsulfonate (2 mM) as ion-pair modifier at 35 oC. Based on the methodology developed, with solutions of reference materials, retention index tables were generated with respect to the internal standard used (acetanilide 25 pg/mL). A library with the 79 UV/Vis spectra of the different drugs and medicines obtained in each chromatogram was created, to help in future identification in sample analysis. Regarding the chromatographic optimization and the robustness analysis of the methodology, the effect of four factors was investigated: the use of two ion pair modifiers, the use of an amine additive as a surface tension modifier, the pH of the buffer used in mobile phase and oven temperature. A complete factorial design was used at two levels and four factors, with three replicates each. The effects of pH and the use of the amine modifier were highly significant, at 95% confidence. Most of the substances studied by liquid chromatography were analyzed by gas chromatography (GC) with an electronic impact (IE) mass detector (MS), in order to analyze the detection capacity of both techniques. With the development of both methodologies, the Department of Forensic Sciences was provided with analytical tools for the scrutiny of drugs and medicines. Due to the great variety of polarities present in the analytes, even from the same family or pharmacological group, both techniques represent an analytical complement. All of the above in favor of improving the Costa Rican judicial system.
Descripción
Barrantes Oconitrillo, E. M.(2011). Escrutinio optimizado de amplio espectro cualitativo y alternativo de aplicación forense por HPLC/ DAD para drogas y medicamentos. [Tesis de Licenciatura]. Universidad Nacional, Heredia, C.R.
Palabras clave
DROGAS, DRUGS, PRODUCTOS FARMACEUTICOS, PHARMACEUTICAL PRODUCTS, CROMATOGRAFIA, ANALISIS QUIMICO, MEDICINA FORENSE