Method development for the separation of carbohydrates predominant in foods by capillary electrophoresis

Orientadores: Helena Teixeira Godoy, Roy Edward BrunsTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia AlimentosResumo: Os carboidratos são compostos muito abundantes na natureza. Estão presentes em todas as etapas evolutivas, tanto em vegetais como animais. Quaisquer reações, de anabolismo ou catabolismo envolvem, de alguma forma, algum carboidrato. Na indústria alimentícia são responsáveis por diversos fenômenos que promovem efeitos químicos, físicos e sensoriais nos alimentos. A importância destes compostos acompanha etapas que vão desde a obtenção da matéria-prima, a formulação, o processamento, as características do produto final até o tratamento dos resíduos gerados. São compostos de ações biológicas e tecnológicas distintas, mas de características físico-químicas semelhantes e, por isso, de difícil separação. Diversos métodos já foram desenvolvidos para a análise desses compostos. No entanto, dentre a literatura pesquisada, nenhum método investigou a possibilidade de separação simultânea dos 13 carboidratos geralmente encontrados em alimentos: glicose, frutose, sacarose, lactose, galactose, lactulose, epilactose, arabinose, manose, maltose, xilose, ribose e maltotriose. Nesse estudo, foram avaliados os efeitos de três diferentes metodologias na separação desses compostos, envolvendo eletroforese capilar de zona, cromatografia eletrocinética micelar com tensoativo catiônico e tensoativo aniônico. Foram investigados diversos fatores que possam influenciar na separação, dentre eles, o tipo e a concentração do eletrólito, o pH, a concentração de surfactante, a voltagem, temperatura, adição de outros sais como tetraborato de sódio, fosfato de sódio, acetato de sódio e cloreto de sódio e a adição de solventes orgânicos como etanol e acetonitrila. Foram utilizadas técnicas estatísticas multivariadas para a otimização do pH, da concentração do eletrólito e do tensoativo. Através da função de Derringer e Suich foi possível realizar a avaliação simultânea de várias respostas e prever as condições analíticas das metodologias de separação. Utilizando a eletroforese capilar de zona foi possível obter a separação de 8 compostos, por cromatografia eletrocinética micelar contendo surfactante catiônico foi possível separar 9 compostos (sacarose, lactose, lactulose, glicose, arabinose, manose, frutose, xilose e ribose), enquanto que utilizando tensoativo aniônico foi obtida a separação dos 13 compostos. Os métodos estatísticos multivariados se apresentaram como uma valiosa ferramenta para o tratamento dos dados e a predição matemática das condições ótimas de separação. Além disso, permitiram a redução do número de experimentos aliada a obtenção de um grande número de informações a respeito dos sistemas estudadosAbstract: The carbohydrates are compounds very abundant in nature. They are present in all evolutionary steps, both in vegetables and animals. Any metabolism or synthesis reaction involves, in some way, a carbohydrate. In food industries, carbohydrates are compounds responsible for several phenomena that promote chemical, physical and sensorial effects. These compounds are important in raw material obtention, formulation, processing, final product characteristics and generated residue treatment. They are compounds having different biological and technological actions, but with similar physico-chemical characteristics and, for this reason, are hard of separate. Several methods have been developed for the analysis of these compounds. However, in the research literature, no method investigating the possibility of simultaneous separation of the 13 carbohydrates commonly found in foods: glucose, fructose, sucrose, lactose, galactose, lactulose, epilactose, arabinose, mannose, maltose, xylose, ribose and maltotriose was found. In this work, three different methodologies for the separation of these compounds were evaluated, involving capillary zone electrophoresis, micellar electrokinetic chromatography with cationic and anionic surfactants. Several factors that could influence the separation were assayed, electrolyte type and concentration, voltage, temperature, addition of sodium tetraborate, sodium phosphate, sodium acetate and sodium chloride salts and addition of organic solvents like ethanol and acetonitrile. Multivariate statistical techniques were used to optimize pH and electrolyte and surfactant concentrations. Employing the Derringer and Suich desirability function it was possible to make a simultaneous evaluation of several responses and predict the optimum analytical conditions of separation. Using the method of capillary zone electrophoresis it was possible to obtain the separation of 8 compounds, by micellar electrokinetic chromatography containing cationic surfactant. It was possible to separate 9 compounds (sucrose, lactose, lactulose, glucose, arabinose, mannose, fructose, xylose and ribose), when using an anionic surfactant. it was obtained the 13 compounds separation. The multivariate statistical methods were a valuable tool for data treatment and for mathematical prediction of the optimum separation conditions. Moreover, they allowed a reduction in number of experiments as well as the obtention of a great amount of information concerning the studied systemsDoutoradoDoutor em Ciência de Alimento

Protective effects of diet containing rutin against trichlorfon-induced muscle bioenergetics disruption and impairment on fatty acid profile of silver catfish Rhamdia quelen

Trichlorfon is an organophosphate insecticide that is widely used on fish farms to control parasitic infections. It has been detected in freshwater ecosystems as well as in fishery products. There is a growing body of evidence to suggest that certain feed additives may reduce or prevent pesticide-induced toxicity in fish. The aim of the present study was to determine whether acute exposure to trichlorfon would alter bioenergetic homeostasis and alter fatty acid profiles in muscles of silver catfish (Rhamdia quelen). We also sought to determine whether rutin prevents or reduces these effects. Cytosolic and mitochondrial creatine kinase (CK) and activities of complexes II-III and IV in muscle were significantly inhibited by exposure to 11 mg/L trichlorfon for 48 h compared to effects in the unexposed group. Total content of polyunsaturated fatty acids (omega-3 and omega-6) were significantly lower in muscle of silver catfish exposed to 11 mg/L trichlorfon for 48 h than in the unexposed group. Addition of 3 mg rutin/kg feed increased CK activity and prevented inhibition of complex IV activity, as well as preventing all alterations of muscle fatty acid profiles elicited by exposure to trichlorfon. No significant differences were observed between groups with respect to muscle adenylate kinase or pyruvate kinase activities, as well as total content of saturated and monounsaturated fatty acids. Our findings suggest that exposure (48 h) to 11 mg trichlorfon/L water inhibits cytosolic and mitochondrial CK activity in muscle. Trichlorfon also affects activities of complexes II-III and IV in respiratory chain, with important consequences for adenosine triphosphate production. The pesticide alters fatty acid profiles in the fish and endangers human consumers of the product. The most important finding of the present study is that inclusion of rutin improves bioenergetic homeostasis and muscle fatty acid profiles, suggesting that it reduces trichlorfon-induced muscle damage

Extração e separação cromatográfica em coluna aberta de pigmentos do cará-roxo (dioscorea trifida) com utilização de materiais alternativo

A presente pesquisa propôs realizar o estudo sobre a utilização materiais alternativos (como a argila montmorilonita) para a purificação de antocianinas oriundas do cará-roxo utilizando a separação cromatografia em coluna aberta

Yerba mate as an inexpensive source of analytical standards of chlorogenic acid isomers: an optimization study

Chlorogenic acid isomers have been increasingly studied because of their beneficial biological effects in humans. However, their commercial analytical standards are high cost, a fact that limits research. Yerba mate (Ilex paraguariensis) is a lowcost, natural plant matrix with marked contents of chlorogenic acids, but its potential as a source of analytical standards of chlogenic acids has never been investigated. Thus, this study aimed to optimize a method to extract, isolate, and purify analytical standards of six chlorogenic acids (3-caffeoylquinic, 4-caffeoylquinic, 5-caffeoylquinic, 3,4-dicaffeoylquinic, 3,5-dicaffeoylquinic, and 4,5-dicaffeoylquinic) from yerba mate (Ilex paraguariensis) using semi-preparative high-performance liquid chromatography. For this, sequential statistical multivariate approaches (central composite designs) were utilized. Using the optimized extraction conditions (5 mL of ethanol:water 26:74 v/v, stirring for 30 min at 60 °C), 1000 g of yerba mate gave a concentrated extract totalizing 21.57 g of chlorogenic acid isomers. We established optimized chromatographic conditions to obtain analytical standards of each compound individually, as well as to produce a mix containing all the compounds, with high yields and purities above 97%. Thus, the optimized conditions to obtain the standards have an excellent yield, employed a natural low-cost matrix, and used simple and automated processes with potential to produce in laboratory and industrial scale. These results show the potential of yerba mate as a novel source of standards of chlorogenic acids, and provide an effective method to produce them in laboratories worldwide, which may contribute to advance the research on these compounds.This work was supported by CAPES (Coordination for the Improvement of Higher Education Personnel), Project numbers 001 and 1267386.info:eu-repo/semantics/publishedVersio

Methylxanthines in 100 Brazilian herbs and infusions: determination and consumption

The contents of caffeine, theophylline and theobromine were studied in 100 plants commercialized in Brazil to investigate new sources of these compounds, and evaluate the consumption of these compounds when these plants are used as infusions (teas). Therefore, two extraction methods were performed, one used an ethanol solution and one used an aqueous infusion process. The extracts were analyzed by ultra-high-performance liquid chromatography. White, green, and black tea (Camellia sinensis) and yerba mate (Ilex paraguariensis) can be considered as potential sources of both caffeine and theobromine. Boldo (Peumus boldus) was a source of theobromine, and small-leaved lime (Tilia cordata), stonebreaker (Phylantus niruri), cow's foot (Bauhinia forficata) and calendula (Calendula officinalis) presented the highest contents of theophylline. This study highlighted herbs not yet investigated for these compounds. The results obtained may be useful in future studies to evaluate the use of these plants as sources of methylxanthines in foods or drugs312125133COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPES126738

Development of a method for simultaneous analysis of caffeine and taurine in energy drinks by micellar electrokinetic chromatography with diode-array detector

The objective of this study was to develop, optimize and validate a fast and reliable method for the simultaneous determination of caffeine and taurine contents by micellar electrokinetic chromatography with diode array detector, using direct and indirect detection concomitantly. Multivariate statistical techniques were used as a central composite design and the simultaneous optimization method of responses of Derringer and Suich were used for optimization. The method was applied in the analysis of 73 samples of energy drinks commercialized in Brazil. The optimized method employed a capillary tube with an extended bulb of 50 mu m i.d. x 72 cm total length, an electrolyte containing 16.20 mmol.L-1 of benzoic acid and 39.90 mmol.L-1 of SDS, a pH value of 7.26, + 30 kV voltage, direct detection of caffeine at 274 nm and indirect detection of taurine at 230 nm. Validation parameters have demonstrated the reliability and applicability of this method. It was found that more than 50% of the samples were out of the legal limits determined by the Brazilian government regarding the taurine content and 68 % contained caffeine below the value declared on the label. Therefore, the need for greater control concerning the composition of these drinks exists.39267368

Analysis of chlorogenic acids isomers and caffeic acid in 89 herbal infusions (tea)

Several plants used to prepare aqueous beverages (teas) are known for their health benefits due to the presence of bioactive compounds, such as isomers of 3-caffeoylquinic (3-CQ), 4-caffeoylquinic (4-CQ), 5-caffeoylquinic (5-CQ), 3,4-dicaffeoylquinic (3,4-DQ), 3,5-dicaffeoylquinic (3,5-DQ), 4,5-dicaffeoylquinic (4,5-DQ) and caffeic acid. This study analyses the presence of these compounds in infusions from 89 plants. We acquired these plants from public markets and analyzed them using ultra-performance liquid chromatography. We found the listed chlorogenic acids in 93% of infusions. The infusions containing highest concentrations were yerba mate (Ilex paraguariensis), with 52.6 mg in 100 mL of infusion, followed by assa-peixe (Boehmeria caudata), white tea (Camellia sinensis), winter’s bark (Drimys winteri), green tea (Camellia sinensis), and elderflower (Sambucus nigra), with 6.0 mg in 100 mL of infusion. Several plant infusions had not been previously studied. Compared to other foods and beverages, these infusions are considered potential sources of chlorogenic acids.737682CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPE