Masters Degrees (Food Science)

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Browsing Masters Degrees (Food Science) by Author "Beelders, Theresa"

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    HPLC method development for the characterisation of the flavonoid and phenolic acid composition of rooibos (Aspalathus linearis) infusions
    (Stellenbosch : Stellenbosch University, 2011-12) Beelders, Theresa; De Villiers, A. J.; Joubert, E.; De Beer, D.; Sigge, G. O.; Stellenbosch University. Faculty of AgriSciences. Dept. of Food Science.
    ENGLISH ABSTRACT: Rooibos tea, produced from the endemic South African fynbos plant Aspalathus linearis, has various healthpromoting benefits which can largely be attributed to its phenolic composition. In this study, the reversedphase liquid chromatographic (RP-LC) separation of the principal phenolic constituents of aqueous rooibos infusions was optimised on conventional high-performance liquid chromatography (HPLC) instrumentation. The phenolic constituents comprised dihydrochalcones (aspalathin and nothofagin), flavones (orientin, isorientin, vitexin, isovitexin, luteolin, luteolin-7-O-glucoside and chrysoeriol), flavonols (quercetin, isoquercitrin, hyperoside and rutin), a hydroxycinnamic acid (ferulic acid) and a phenylpropanoid (phenylpyruvic acid glucoside, PPAG). A systematic approach towards method development was adopted: resolution was optimised by simultaneous optimisation of gradient conditions and temperature on a sub-2 6m phase to exploit the benefits of this phase for fast routine analyses. The developed HPLC method, utilising photodiode-array (PDA) detection, yielded complete separation of the 15 target analytes on the 1.8 µm C18 column, thermostatted to 37°C, within 37 min (total analysis time: 50 min). The method was successfully validated and demonstrated its suitability for the fast, quantitative analysis of aqueous infusions of unfermented and fermented rooibos. Mass spectrometric (MS) and tandem MS detection confirmed peak purity and permitted the tentative identification of 13 additional phenolic compounds, including a flavonol Odiglycoside (quercetin-3-O-robinobioside), a luteolin-6-C-pentoside-8-C-hexoside and a novel C-8-hexosyl derivative of aspalathin reported here for the first time. The HPLC-PDA method was subsequently applied to a large number of fermented rooibos samples representative of different production seasons (2009, 2010 and 2011) and quality grades (grades A, B, C and D) to capture as much potential variation in the phenolic composition as possible. Production season had no clear effect on the levels of the individual phenolic compounds in ‘cup-of-tea’ rooibos infusions, whilst high quality tea (grades A and B) was associated with higher levels of phenolic compounds and soluble solids than low quality tea (grades C and D). Steam-pasteurisation of the plant material, required to obtain a product of high microbiological quality, induced significant reductions in the mean values of most of the phenolic compounds in rooibos infusions. The major phenolic constituents of steam-pasteurised, fermented rooibos were isoorientin and orientin, whilst quercetin-3-O-robinobioside, PPAG and aspalathin were also present in high concentrations. Representative content values of the major phenolic compounds present in a typical ‘cup-of-tea’ rooibos infusion were thus obtained and the generated data are suitable for inclusion in food composition databases. The application of comprehensive two-dimensional liquid chromatography (LCxLC) was investigated as an alternative approach for the detailed investigation of rooibos phenolics. The combination of hydrophilic interaction chromatography (HILIC) in the first dimension and RP-LC in the second dimension offered different separation selectivities and hence a high degree of orthogonality. HILICxRP-LC provided a significant improvement in resolution, as is evident from practical peak capacities in excess of 2000 and 800 for the off-line and on-line methods, respectively. Further optimisation, particularly of the first dimension separation, is however required to improve the LCxLC separation of complex rooibos phenolic fractions.