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Browsing by Subject "UPLC"

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  • Approaches to folyl polyglutamate analysis 
    Yang, Yingying (2013)
    The literature review presented the effects of the polyglutamate chain on the biological and nutritional properties of folates and the main methods used for folate assays, with a special emphasis on the approaches to studying intact polyglutamates. A brief introduction regarding safety aspects of folate fortification was also given. The aim of this study was to develop a UPLC-FLR/PDA method for simultaneous determination of polyglutamyl folate vitamers. Chromatographic conditions were optimised for the resolution of polyglutamyl 5-methyltetrahydrofolates and major naturally-occurring monoglutamates. Method validation was conducted for both the UPLC method and affinity chromatography. Applicability of the validated method was evaluated on lupin flour, faba bean flour, and dry yeast, which were subjected to preparatory treatments with and without deconjugation. In addition, the effects of the sequential modification of preparatory treatments on the folate content and composition were investigated by using both the UPLC method and Lactobacillus rhamnosus assay. A desirable separation of target polyglutamates and monoglutamates was successfully achieved on the BEH C18 UPLC column within 11 minutes. The optimised UPLC method showed satisfactory selectivity, linearity, and sensitivity for the determination of methylated polyglutamates in the femtomole range and monoglutamates in the picogram range. Affinity chromatography showed satisfactory recoveries for polyglutamyl 5-methyltetrahydrofolates, but not for 5-formyl polyglutamates. In all three selected foods, 5-methyltetrahydrofolate was the dominant folate vitamer. Meanwhile, the analysis of undeconjugated samples showed that in the intact methylated folate pools, pentaglutamate predominated in legume flours and heptaglutamate in dry yeast. In addition, different sequences of enzyme and purification pretreatments were found to significantly affect both the total measurable folates and the folate profiles. Our standard preparatory procedures comprising simultaneous treatments with amylase and conjugase, then protease and affinity purification resulted in the greatest yield of total folates, but UPLC analysis indicated incomplete deconjugation. However, a modification in which deconjugation was conducted as the last step enhanced hydrolysis efficiency.
  • Physical and chemical characterization of two quinoa side streams and separation of valuable components from them 
    Penttilä, Anna (2022)
    Quinoa is a South American crop plant and the abrasive pearling of its seeds produces abrasion waste as a side stream. The aim of this study was to examine the chemical and physical differences between two different side streams and extract some valuable components from two abrasive side streams. The side streams were obtained with two different pearling methods and are referred to in this study as fine and coarse side stream. Four different extraction media; water, 0.4 M sodium chloride, 70% (v/v) ethanol and pH 9.0 sodium hydroxide, were used solubilize protein and carbohydrates from the side streams. Electrophoresis (SDS-PAGE) was used to detect the protein composition of the side streams and ultrafiltration (UF) of aqueous extract together with UPLC of UF permeate were used to separate and detect mono- and disaccharides from the side streams. The coarse side stream had bigger particle size and higher density and it contained more protein and fat than the fine side stream which instead had higher ash and fibre content than the coarse side stream. Mainly albumins and globulins as well as some glutelins were detected from water, saline, and alkaline extracts of both side streams. The coarse side stream’s ultrafiltration permeate contained sucrose, fructose, and glucose - sucrose being most abundant and glucose least. The fine side stream’s permeate contained glucose and some fructose, but no sucrose. This study found quinoa pearling side streams to be a potential source for carbohydrates and protein, but the differences in the compositions of the side streams significantly affect their potential for wider use. Further studies are needed to investigate the protein yields and mono and disaccharide content of extracts with other extraction media. This study also raises further research topics, such as the effect of pre-treatment methods on the mono and disaccharide yields.

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