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Abstract(s)
Na presente dissertação são descritas duas metodologias analíticas para a identificação e quantificação de compostos fenólicos, nomeadamente a vanilina e o 3,4-dihidroxibenzaldeído, em amostras de concentrado obtido através de nanofiltração do efluente produzido na etapa inicial do processamento da cortiça destinada à remoção de contaminantes, designada como ―cozimento‖.
Utilizando a cromatografia líquida de alta eficiência (HPLC) foi realizada a optimização analítica e separação cromatográfica. Utilizou-se a metodologia de análise de HPLC pelo método de eluição por gradiente com coluna de fase reversa C18 5μm (250 x 4mm) à temperatura de 25°C, com um fluxo de eluição de 1mL min-1 e e a detecção foi feita por um detector por arranjo de diodos.
A separação cromatográfica utilizou como fases móveis as misturas A: ácido fosfórico 0,01M e B: água – metanol – ácido fosfórico 0,01M (70:28:2 (v/v)), usando o seguinte programa de gradiente: gradiente linear de 0% a 40% de B em 3 minutos, gradiente linear de 40% a 60% de B em 22 minutos, 60% de B isocrático em 18 minutos, gradiente linear de 60% a 80% de B em 12 minutos e por fim 80% de B isocrático em 5 minutos.
A dissertação apresenta a segunda parte dedicada à optimização da metodologia analítica, utilizando a extracção em fase sólida (SPE) como método de preparação de amostra, com objectivo de extrair os compostos fenólicos presentes nas amostras em questão.
Os métodos analíticos foram validados no que diz respeito à sensibilidade, curva analítica e linearidade, limite de detecção (LD) e de quantificação (LQ), precisão, exatidão (recuperação) e robustez, podendo ser aplicados à análise da vanilina e 3,4-dihidroxibenzaldeído nas amostras em questão.
As curvas analíticas apresentaram valores de R2 de 0,9996 para a vanilina e 0,9997 para o 3,4-dihidroxibenzaldeído.
A metodologia para determinação de compostos fenólicos mostrou-se seletiva e linear para uma faixa de trabalho de 1 a 6 ppm para a vanilina e de 1 a 5 para o 3,4-dihidroxibenzaldeído e apresentou limites de detecção e quantificação para a vanilina de 0,14 e 0,43 ppm e para o 3,4-dihidroxibenzaldeído de 0,11 e 0,32 ppm, respectivamente.
O método mostrou-se adequado à análise dos compostos fenólicos e todos os parâmetros de validação encontram-se dentro dos limites sugeridos para validação de métodos cromatográficos.
In the present study are described two analytical methods for identification and quantification of phenolic compounds, such as vanillin and 3,4-dihydroxybenzaldehyde in concentrated samples obtained through nanofiltration effluent produced in the initial stage of the processing of cork intended for removing contaminants, referred to as "cooking". Using high performance liquid chromatography (HPLC) it was performed an analytical optimization and chromatographic separation. It was used a methodology for analysis by HPLC gradient elution method with reverse phase column C18 column 5μm (250 x 4mm) at 25 ° C, with a elution flow of 1mL min-1 and detection was performed by a diode array detector. The chromatographic separation used mixtures as mobile phases A: 0.01 M phosphoric acid and B: water - methanol - 0.01 M phosphoric acid (70:28:2 (v/v)), using the following gradient program: linear gradient from 0% to 40% B in 3 min, linear gradient from 40% to 60% B in 22 min, 60% isocratic B in 18 minutes, linear gradient of 60% to 80% B in 12 minutes and finally isocratic 80% B in 5 minutes. This thesis presents the second part dedicated to the optimization of analytical method, using solid phase extraction (SPE) as a method of sample preparation in order to extract the phenolic compounds present in the samples in question. The analytical methods were validated with regard to sensitivity, linearity, and analytic curve, detection limit (LD) and quantification (LOQ), precision, accuracy (recovery) and robustness can be applied to the analysis of vanillin and 3,4-dihydroxybenzaldehyde in the samples in question. The analytical curves present values of R 2 of 0.9996 for vanillin and 0.9997 for 3,4- dihydroxybenzaldehyde. The methodology for the determination of phenolic compounds shown to be selective and linear for a working range of 1 to 6 ppm for vanillin and 1 to 5 for the 3,4- dihydroxybenzaldehyde and showed limits of detection and quantification of 0.14 to 0.43 ppm for vanillin and 0.11 to 0.32 ppm for 3,4-dihidroxibenzaldeído respectively. The method was suitable for the analysis of phenolic compounds and all of the validation parameters are within the limits suggested for validation of chromatographic methods.
In the present study are described two analytical methods for identification and quantification of phenolic compounds, such as vanillin and 3,4-dihydroxybenzaldehyde in concentrated samples obtained through nanofiltration effluent produced in the initial stage of the processing of cork intended for removing contaminants, referred to as "cooking". Using high performance liquid chromatography (HPLC) it was performed an analytical optimization and chromatographic separation. It was used a methodology for analysis by HPLC gradient elution method with reverse phase column C18 column 5μm (250 x 4mm) at 25 ° C, with a elution flow of 1mL min-1 and detection was performed by a diode array detector. The chromatographic separation used mixtures as mobile phases A: 0.01 M phosphoric acid and B: water - methanol - 0.01 M phosphoric acid (70:28:2 (v/v)), using the following gradient program: linear gradient from 0% to 40% B in 3 min, linear gradient from 40% to 60% B in 22 min, 60% isocratic B in 18 minutes, linear gradient of 60% to 80% B in 12 minutes and finally isocratic 80% B in 5 minutes. This thesis presents the second part dedicated to the optimization of analytical method, using solid phase extraction (SPE) as a method of sample preparation in order to extract the phenolic compounds present in the samples in question. The analytical methods were validated with regard to sensitivity, linearity, and analytic curve, detection limit (LD) and quantification (LOQ), precision, accuracy (recovery) and robustness can be applied to the analysis of vanillin and 3,4-dihydroxybenzaldehyde in the samples in question. The analytical curves present values of R 2 of 0.9996 for vanillin and 0.9997 for 3,4- dihydroxybenzaldehyde. The methodology for the determination of phenolic compounds shown to be selective and linear for a working range of 1 to 6 ppm for vanillin and 1 to 5 for the 3,4- dihydroxybenzaldehyde and showed limits of detection and quantification of 0.14 to 0.43 ppm for vanillin and 0.11 to 0.32 ppm for 3,4-dihidroxibenzaldeído respectively. The method was suitable for the analysis of phenolic compounds and all of the validation parameters are within the limits suggested for validation of chromatographic methods.
Description
Keywords
Cromatografia líquida de alta eficiência Compostos fenólicos por HPLC Vanilina Cromatografia