Validación del método para la cuantificación de alcohol en sangre por cromatografía gaseosa con detector de ionización de llama y espacio de cabeza y correlación con el método enzimático / Validation of the method for the quantification of blood alcohol by gaseous chromatography with flame ionization detector and head and correlation space with the enzymatic method

El presente estudio describe la estandarización del método para la cuantificación de alcohol en sangre por Cromatografía Gaseosa con Detector de Ionización de llama y espacio de cabeza. La separación del alcohol de una matriz compleja como la sangre fue llevada a cabo de manera exitosa utilizando una columna ELITE-1, la fase móvil o gas de arrastre es una mezcla de hidrógeno y aire, inyector split y detector de ionización de llama. El método fue validado con los siguientes parámetros: especificidad, linealidad, precisión, exactitud, límite de detección y límite de cuantificación. También se realizó la prueba de aptitud del sistema. El método fue específico para el alcohol, la respuesta fue lineal en el rango de 0,125 ­ 1,0 mg/mL de concentración del analito. El valor del coeficiente de variación o desviación estándar relativa (C.V. o DSR) para la precisión fue óptimo. La recuperación media fue de 99,06%. Y el límite de detección y cuantificación resultaron óptimos para la cuantificación de alcohol en sangre en el rango definido. Finalmente el método cromatográfico fue correlacionado con el método analítico (previamente validado) actualmente utilizado en la División de Dosage Etílico del Instituto de Investigaciones Técnicas de la Universidad Policial Mariscal Antonio José de Sucre, mediante el análisis estadístico de las respuestas analíticas de ambos métodos se obtuvo una muy buena correlación por lo que se concluye que el método cromatográfico puede ser utilizado para los fines consiguientes.

The present study describes the standardization of the method for the quantification of alcohol in blood by Gas Chromatography with Flame Ionization Detector and headspace. The separation of the alcohol from a complex matrix such as blood was successfully carried out using an ELITE-1 column, the mobile phase or entrainment gas is a mixture of hydrogen and air, split injector and flame ionization detector. The method was validated with the following parameters: specificity, linearity, precision, accuracy, limit of detection and limit of quantification. The system fitness test was also performed. The method was alcohol specific, the response was linear in the range of 0.125 - 1.0 mg / mL analyte concentration. The value of the coefficient of variation or relative standard deviation (C.V. or DSR) for precision was optimal. The mean recovery was 99.06%. And the limit of detection and quantification were optimal for the quantification of blood alcohol in the defined range. Finally the chromatographic method was correlated with the enzymatic method (previously validated) currently used in the Ethical Dosage Division of the Institute of Technical Investigations of the Mariscal Antonio José de Sucre Police University, through the statistical analysis of the analytical responses of both methods a very good correlation was obtained by which it is concluded that the chromatographic method can be used for the consequent purposes.

Determinaçäo de colesterol em ovos: comparaçäo entre um método enzimático e um método por cromatografia líquida de alta eficiência / The determination of cholesteral in eggs: a comparison of an enzymatic method with that of high performance liquid chromatography

Os métodos enzimáticos apesar de serem baratos e simples são pouco utilizados na análise de colesterol em alimentos, uma vez que a enzima reage com qualquer esterol presente na amostra. Considerando que ovos possuem apenas colesterol, o objetivo deste trabalho foi otimizar um método por cromatografia líquida de alta eficiência (CLAE). Em ambos os métodos foi realizado saponificação direta das amostras e extração da matéria insaponificável com hexano. No método enzimático foi utilizado um kit (LABORLAB SA) e a absorbância foi determinada a 499 nm, 90 minutos após a reação. Na CLAE utilizou-se coluna C18, 100x4,6 mm x 4um (Chromolith, Merck), fase móvel acetonitrila:isopropanol (85:15) e fluxo de 2mL/min. Os métodos foram validados através de testes de recuperação, material de referência certificado de ovo em pó (SRM 1846,NIST) e receptibilidade. Os valores médios de colesterol nos ovos foram 1363 +- 47 e 1364 +- 40mg/100g de gema para o método enzimático e por CLAE, respectivamente, não havendo diferenças significativas (p>0,01) entre os resultados obtidos pelos dois métodos. Ambos os métodos podem ser utilizados com confiabilidade para determinação de colesterol em gema de ovos, sendo que o método enzimático possui vantagem de ser bem menos oneroso que o por CLAE. (AU)

Enzymatic methods are simple and of low-cost, but are nevertheless little used in thedetermination of cholesterol in foods as the enzyme reacts with any sterol, present in the sample. As eggsonly have cholesterol, the objective of this study was to optimize an enzymatic method for the determinationof cholesterol in eggs and compare it to a high performance liquid chromatography (HPLC) method. Forboth methods, the samples were saponified directly and the unsaponifiable matter extracted with hexane.For the enzymatic method, a test kit (LABORLAB S.A) was used and the absorbance read at 499 nm, 90minutes after the reaction. For HPLC, a C18, 100 x 4.6 mm x 4 µm (Chromolith, Merck) column was used,the solvent being acetonitrile:2-propanol (85:15 v/v) with a flow rate of 2 mL/min. The methods werevalidated according to recovery test, the use of egg standard reference material (SRM 1846, NIST), andrepeatability. The mean result for the enzymatic method was 1363 ± 47 and for HPLC 1364 ± 40 mg/100g of yolk. There were no significant differences (p>0.01) found between the results of the two methods.Both methods are reliable for use in the determination of cholesterol in yolk, the enzymatic method showingthe additional advantage of being cheaper than HPLC. (AU)