RESUMO
A size-exclusion chromatographic method using an aqueous solvent as eluent was developed for the determination of the molecular mass of procyanidins has been developed. Size-exclusion chromatography is a chromatographic technique used to determine molecular weight distribution of polymers. During the development work, PL aquagel-OH columns and several different eluent compositions were investigated. The best separation of procyanidins was obtained with an eluent consisting of water-methanol (50:50, v/v), NaNO3: 0.18 M, pH < pKa + 2. The results from this aqueous SEC method were compared with results obtained using SEC with organic solvents.
Assuntos
Cromatografia em Gel/métodos , Proantocianidinas/isolamento & purificação , Calibragem , Peso Molecular , Proantocianidinas/química , Vitis/químicaRESUMO
In this work, we wanted to devise a reliable method to characterize polymerized forms of tannins, their structural information and mass distribution. Size-exclusion chromatography (SEC) is a chromatographic technique used to determine molecular mass (weight) distributions of polymers. One important step in the data treatment is the modeling of the calibration curves. Polystyrenes (PS) are standards usually used because no commercial procyanidin (PC) standards are available. An off-line coupling of SEC and MALDI was carried out to measure differences between polystyrenes and procyanidins. Thus, a new calibration curve was established; from 1000 to 8000 Da, there is a good correlation between the MALDI and PS calibration curves, in this field the PS calibration is correct and enables true mass determination. For masses above 8000 Da, PS calibration overestimates the real molecular weight of PC, overestimation of 53%. And for masses below 1000 Da, PS calibration underestimates their real molecular weight (10-15%). This means that to truly characterize PC, calibration based on PC standards is required.
