Simultaneous quantification of lycopene, β-carotene, retinol and α -tocopherol in plasma after a simple extraction procedure: stability study and application to human volunteers

Abstract

Um método para quantificação simultânea de licopeno, β-caroteno, retinol e α-tocoferol por cromatografia líquida de alta eficiência (HPLC) com detecção no visível/fluorescente e eluição isocrática foi otimizado e validado. O método consiste de extração líquido-líquido rápida e simples e posterior quantificação do sobrenadante extraído por HPLC. Alíquotas de plasma foram estocadas a -20°C por três meses para estudo da estabilidade. Aplicação metodológica foi realizada em amostras fornecidas por pintores e indivíduos não expostos a tintas (n = 75). O ensaio foi linear para todas as vitaminas analisadas (r > 0,99). Precisões intradia e interdia apresentaram coeficiente de variação (CV) menor que 5%. Exatidões variaram de 0,29 a -5,80% e recuperações entre 92,73 e 101,97%. Amostras de plasma e sobrenadante extraído foram estáveis por até 60 dias a -20°C. Foi demonstrada uma diminuição significativa nas concentrações de licopeno, β-caroteno e retinol em indivíduos expostos quando comparados com os não-expostos (p < 0,05). O método é simples, reprodutível, preciso, exato e sensível, e pode ser utilizado na rotina de laboratórios clínicos.A method for the simultaneous quantification of lycopene, β-carotene, retinol and α-tocopherol by high-performance liquid chromatography (HPLC) with Vis/fluorescence detection with isocratic elution was optimized and validated. The method consists of a rapid and simple liquid-liquid extraction procedure and a posterior quantification of extracted supernatants by HPLC. Aliquots of plasma were stored at -20°C for three months for stability study. The methodology was applied to samples from painters and individuals not exposed to paints (n = 75). The assay was linear for all vitamins (r > 0.99). Intra-and inter-run precisions were obtained with coefficient of variation smaller than 5%. The accuracies ranged from 0.29 to -5.80% and recoveries between 92.73 and 101.97%. Plasma samples and extracted supernatants were stable for 60 days at -20°C. A significant decrease of lycopene, β-carotene and retinol concentrations in plasma from exposed individuals compared to non-exposed individuals (p < 0.05) was observed. The method is simple, reproducible, precise, accurate and sensitive, and can be routinely utilized in clinical laboratories.

Blood antioxidant nutrients in riparian villagers of the Brazilian Amazon: its associations with wet/dry seasons and modulation by sociodemographic determinants

“…It is known that the main sources of β-carotene are fruits and vegetables, and a previous study conducted in the same region revealed that the consumption of this type of food is much higher in the RS when compared to the DS 1 , which is consistent with the interseasonal pattern we found. Unlike those of plasma β-carotene, the levels of lycopene were higher in the DS, with mean levels found in both seasons lower than in other studies with other populations all over the world 34,35 , and even from studies with other Brazilians living in the southern region 17,25 . Plasma carotenoids levels found in this study are in good agreement with the lower limit found for North American populations 35 .…”

“…Interestingly, the population's RS plasma β-carotene levels are substantially higher than those currently found in other populations 13,34 , and even in other Brazilian areas 17,25 . During the dry season, plasma β-carotene levels drop considerably but still remain in the safe range (>0.30μM/L) recommended by WHO 24 .…”

“…Literature data of essential metals like copper (Cu), manganese (Mn), selenium (Se) and zinc (Zn) 3,14,16,[19][20][21][22][23] in blood, β-carotene, lycopene and vitamins (A and E) in plasma 17,24,25 are shown in Table 2, along with interseasonal comparisons of these parameters. Blood Cu, Se and Zn levels were significantly lower in the RS when compared to DS, whereas blood Mn levels did not differ significantly between seasons.…”

“…Thus, the information on food frequency consumed during the dry and rainy season in this study was based on a study of 2001 1 , which was performed in the community of Brasília Legal, nearby our study area. Another limitation is that we did not evaluate the nutrient contents of the main foods consumed in this region during the two study periods, while it is known that nutrient contents can be different among different places 17,32 . These limitations prevented us from identifying more specific exposure pathways to blood Cu, Mn, Se, Zn, carotenoids and vitamins in this population.…”

“…The quantification of plasma β-carotene, lycopene and vitamins (A and E) was in accordance with detailed procedures previously published 17 .…”