Metodologias para determinação de ácidos graxos em alimentos por técnicas cromatográficas, eletroforéticas e espectroscópicas

Abstract

The addition of hydrogenated vegetable oils to food has been restricted worldwide due to the harmful effects that industrial trans fatty acids (TFA) can cause to human health. The development of new analytical methods is needed to attend legal requirements, and fast methodologies for TFA determination in foods were the focus of this thesis. Initially, a method for the determination of total TFA of octadecenoic acid in foods was optimized through Capillary Zone Electrophoresis with Direct Absorption Detection within the Ultraviolet Region. Optimum instrumental parameters were studied by using a background electrolyte previously described in the literature to direct detection, through an univariate evaluation of capillary length and a 32 factorial design having voltage and temperature as factors. Sample preparation was then studied, and comprised only alkaline hydrolysis of triacylglycerols. The validation of the developed method comprised the evaluation of linearity, specificity, precision, accuracy, robustness, and limits of detection and quantification. Relative standard deviation ≤ 5,72 % were obtained for instrumental precision when considering peak area, peak height and migration time, in a quadruplicate injection of standards. The calculated detection and quantification limits were 0,17 and 0,54 mmol L-1, respectively. In addition, no significant evidence was found within the 95% confidence interval when the methods by Capillary Electrophoresis and Gas Chromatography were compared. Among all the methods available in the literature for the determination of total TFA from octadecenoic acid using Capillary Electrophoresis, this is the method that provides the fastest result so far, with an analysis time less than 4 min. A new method for the separation of major TFA positional isomers of octadecenoic acid in food from natural and industrial origin (vaccenic and elaidic acids) was then developed using Capillary Zone Electrophoresis Modified with Cyclodextrins and Direct Absorption Detection within the Ultraviolet Region. The electrolyte was optimized univariately for this novel separation, and instrumental parameters, such as voltage and temperature, were optimized by a 22 factorial design. Sample preparation was evaluated for spreadable cheese, and only the alkaline hydrolysis of triacylglycerols was effective for the preparation of this matrix. The running time was less than 25 min and the estimated detection limit for elaidic and vaccenic acids was 0,02 mmol L-1. In addition, the mechanism of this separation was studied through molecular modeling. This approach presents a potential application in the discrimination of major isomeric TFA from natural and industrial origin, which is already required in legislations of several countries. Finally, other methodologies for the determination of fatty acids in food matrices were evaluated through chromatographic (Gas Chromatography with Flame Ionization Detector and Mass Spectrometry Detector, as well as Thin-layer Chromatography) and spectroscopic techniques (Infrared, Raman and Nuclear Magnetic Resonance) associated with chemometric tools, complementary to Capillary Electrophoresis, and were briefly discussed at the end of this thesis.