Department of Chemistry and Polymer Science

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Browsing Department of Chemistry and Polymer Science by Author "Alberts, P."

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    Applicatins of liquid chromatography-tandem mass spectrometry to wine analysis : targeted analysis and compound identification
    (Stellenbosch : Stellenbosch University, 2012-12) Alberts, P.; Stander, M. A.; De Villiers, A. J.; Stellenbosch University. Faculty of Science. Dept. of Chemistry and Polymer Science.
    ENGLISH ABSTRACT: The wine industry is an important sector of agriculture and wine analysis forms the basis of assessing compliance of its commodities with regulatory standards and research in this field. Liquid chromatography (LC) is extensively used for the determination of a wide range of nonvolatile wine components, but conventional detectors impose performance limitations on the technique that prevents its application to sophisticated analytical problems. In particular, conventional detectors for LC often lack the sensitivity and specificity for the determination of many wine compounds, especially trace level analytes, and furthermore, do not possess spectral capabilities for compound identification or structure elucidation. The hyphenation of mass spectrometry (MS) to LC has led to the introduction of a range of detectors that confers high levels of sensitivity and selectivity to the technique. In addition, a wide variety of MS architectures are available that are inherently suited for targeted analysis or structure elucidation studies. In this dissertation, the potential benefits of liquid chromatography – tandem quadrupole mass spectrometry (LC-MS/MS) to solve analytical problems relevant to the wine industry are explored. LC-MS/MS is a particularly versatile analytical technique because both mass analysers can be operated in full-spectrum mode or selected-ion monitoring, which, together with optional fragmentation, gives rise to four modes of operation that may be used for highly specific and sensitive targeted analysis or spectral investigations. In multiple reaction monitoring (MRM) mode, both analysers are set at single ion frequencies specific for the compound under investigation and one or more of its product fragments, respectively. MRM mode is ideally suited for trace level analysis in complex mixtures, even in cases where the target components are not resolved from interferences. In this study, MRM detection was used to solve challenges relevant to the wine industry for the selective quantitation of target analytes that could not be analysed by conventional LC methods. The application of this approach for the analysis of natamycin, ethyl carbamate (EC) and 3-alkyl-2- methoxypyrazines (MPs) in wine is demonstrated. Natamycin is an antimicrobial preservative that is not permitted in wine in the European Union. A rapid and sensitive method for the determination of natamycin was developed, and has been used since 2009 to regulate this vitally important sector of the South African wine export industry. EC is a natural carcinogen that occurs at trace level amounts in alcoholic products. It also has the potential to accumulate in wines and can occur in very high concentrations in some fruit brandies. The determination of EC is complicated by its physicochemical properties, and available analytical methods suffer from drawbacks such as the requirement for elaborate extraction procedures and high solvent consumption. A novel method for the determination of EC in wines, fortified wines and spirits is described and it was applied to perform an audit of the South African industry as well as to investigate factors responsible for its accumulation in alcoholic beverages. This work forms an integral part of the food safety mandate of the State and it ensures that export products comply with international norms for trade. MPs are ultra-trace-level aroma compounds that contribute to the varietal character of Sauvignon blanc wines. Their analytical determination is challenging due to their low levels of occurrence. The loading capacity of LC combined with the sensitivity and resolving power of MS was exploited to analyse concentrated extracts, in order to achieve very low limits of detection. The performance of the LC-MS/MS method enabled the quantitation of these compounds at their natural levels of occurrence, including the first quantitation and spectral confirmation of 3- ethyl-2-methoxypyrazine in wine. Extensive data pertaining to South African Sauvignon blanc wines are reported and statistical analysis is performed, reporting the correlation of variables such as vintage and origin as well as wine parameters such as malic acid with wine MPs. Furthermore, the application of LC-MS/MS for structural elucidation and screening of target classes of analytes was demonstrated for the analysis of red wine anthocyanins. The anthocyanidin-glycosides are responsible for the colour of red grapes and wine, contribute to the sensory properties of wine, and are also of interest due to their beneficial biological properties. Their determination is complicated by their large numbers and structural diversity, further exacerbated by diverse reactions during wine ageing as well as the lack of reference standards for most members of this class of compounds. Tandem MS in scan mode was used for the highly selective detection of glycosylated anthocyanins and derivatives, exploiting the predictable elimination of the sugar moiety in neutral loss mode. Concurrent survey scan experiments were used to unambiguously identify neutral loss detected compounds. The method therefore follows a simplified and structured approach for unambiguous peak identification based on elution order and mass spectral information to impart a high level of certainty in compound identification. In summary, the work presented in this dissertation demonstrates that LC-MS/MS is a versatile and powerful analytical approach for the analysis of diverse compounds of relevance to the wine industry. The sensitivity and specificity of MRM mode, and the selectivity and spectral capabilities of neutral loss and survey scan modes of MS/MS detection, is amply demonstrated by the applications presented in the dissertation.
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    Development of a novel LC-MS/MS method for the detection of adulteration of South African sauvignon blanc wines with 3-alkyl-2-methoxypyrazines
    (Stellenbosch : University of Stellenbosch, 2008-03) Alberts, P.; De Villiers, A. J.; Stander, M. A.; University of Stellenbosch. Faculty of Science. Dept. of Chemistry and Polymer Science.
    A method for the detection of adulteration of South African Sauvignon blanc wines, by enrichment with foreign sources of 3-alkyl-2-methoxypyrazenes, is described. The levels of three 3-alkyl-2-methoxypyrazenes (3-isobutyl-, 3- isopropyl- and 3-sec-butyl-2-methoxypyrazine) in South African Sauvignon blanc wines were measured with liquid chromatography-mass spectrometry. Sample preparation involved clean-up and pre-concentration by distillation followed by solvent extraction of the distillate with dichloromethane. Extracts were acidified and concentrated by evaporation and finally reconstituted to a fixed volume to affect quantitative pre-concentration of the samples. Sample extracts were separated with reversed phase liquid chromatography utilizing a phenyl-hexyl separation column. Residues were measured with liquid chromatography-mass spectrometry utilizing a tandem quadrupole mass spectrometric detector operated in multiple reaction monitoring mode for optimal trace level quantitation. Atmospheric pressure chemical ionization was utilized as electrospray ionization was found to suffer from quenching effects attributed to the sample matrix. Qualitative information was obtained from the relevant molecular ions as well as two secondary ion transitions (and one ion ratio) in each case. Recoveries obtained by the extraction procedure were better than 90% with coefficient of variance of better than 10% at concentrations from 1 to 100 ng/L. The limit of detection of the method was 0.03 ng/L and the limit of quantification 0.10 ng/L for the three analytes measured. The described LC-MS method is more sensitive for the determination of 3-alkyl-2-methoxypyrazines in wine than GC methods reported for the same purpose. From the experimental data, a set of parameters were established to discriminate adulterated South African Sauvignon blanc wines. It was demonstrated that the 3-isobutyl-2-methoxypyrazine concentration, despite showing considerable variance, was confined to a relatively narrow range spanning approximately two orders of magnitude (0.20 to 22 ng/L). A clear indication of possible maximum values for this compound in South African Sauvignon blanc wines was obtained from the analysis of a large number of samples (577), spanning most relevant wine producing regions and representing vintages 2003 to 2006. It was also demonstrated that South African Sauvignon blanc wines contain the major 3- alkyl-2-methoxypyrazenes in reasonably distinct relative amounts and that the said ratios of abundance may be used to elucidate authenticity. The expected effect of adulteration with green pepper extracts or some synthetic preparations on the 3-isobutyl-2-methoxypyrazine concentration as well as the relative abundances were also determined by characterizing the corresponding profiles in green peppers and some synthetic flavor preparations. Two adulterated samples in the dataset were identified by both outlined criteria. A limited number of wines of other cultivars were also analyzed. The results represent the most complete and accurate data on the 3-alkyl-2-methoxypyrazine content of South African Sauvignon blanc wines to date. A publication covering the work presented in this thesis is currently in preparation.
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    Quantitative survey of 3-alkyl-2-methoxypyrazines and first confirmation of 3-ethyl-2-methoxypyrazine in South African Sauvignon blanc wines
    (South African Society for Enology and Viticulture, 2013) Alberts, P.; Kidd, M.; Stander, M. A.; Nieuwoudt, Helene; Tredoux, A. G. J.; De Villiers, A. J.
    3-Alkyl-2-methoxypyrazines are important aroma components in Sauvignon blanc wine, contributing to the typical vegetative character associated with the cultivar. The analysis of methoxypyrazines is highly challenging and, as a result, limited quantitative data are available for South African Sauvignon blanc wines. In this study, liquid chromatography-tandem mass spectrometry was applied for the sensitive quantification of the three principal methoxypyrazines in 881 South African Sauvignon blanc wines. The total wine methoxypyrazine concentration exceeded the combined recognition threshold for the vegetative aroma in more than 60% of the samples, demonstrating the involvement of these compounds in the typical aroma of the cultivar. The analytical method was adapted to allow the accurate quantitation and spectral confirmation of 3-ethyl-2-methoxypyrazine in Sauvignon blanc wines for the first time. Concentrations ranged between 1.1 and 4.9 ng/L. Statistically significant relationships between wine methoxypyrazine concentrations and parameters related to the maturity of the grapes and to the climate, as well as geographical origin and vintage, were found.