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Σε αυτήν τη διατριβή, πραγματοποιήθηκε ανάπτυξη μίας μεθοδολογίας SPE-GC/MS και χημειομετρική ανάπτυξη μίας μεθοδολογίας SPME-GC/MS για τον προσδιορισμό 21 ελεύθερων και γλυκοζιτικά δεσμευμένων πρωτογενών αρωματικών συστατικών (τερπενικών ενώσεων, C6 αλκοολών, C13 νορισοπρενοειδών και βενζολικών παραγώγων) σε λευκούς ξηρούς οίνους.Αρχικά, πραγματοποιήθηκε συγκριτική μελέτη μεταξύ επτά εμπορικά διαθέσιμων SPE μικροστηλών για το διαχωρισμό και την απομόνωση των ελεύθερων και των πρόδρομων αρωματικών ενώσεων με SPE. Την καλύτερη επίδοση, με κριτήριo την ανάκτηση σήματος των ελεύθερων και μετά από ενζυμική υδρόλυση το αναλυτικό σήμα των δεσμευμένων συστατικών, είχε η μικροστήλη Isolute ENV+.Για την ανάπτυξη της μεθοδολογίας SPΜE-GC/MS, αρχικά πραγματοποιήθηκε συγκριτική μελέτη μεταξύ πέντε εμπορικά διαθέσιμων ινών SPΜE, με δειγματοληψία από την υγρή και την αέρια φάση του δείγματος. Επιλέχθηκε μικροεκχύλιση από την υγρή φάση με ίνα PDMS/DVB καθώς με αυτό το συνδυασμό παρατηρήθηκε ικανοποιητική ανάκτηση και η βέλτιστη αναπαραγωγιμότητα. Με εφαρμογή μερικού πειραματικού σχεδιασμού (Plackett-Burman), εξετάσθηκε η επίδραση στη μικροεκχύλιση επτά παραγόντων και επιλέχθηκαν εκείνοι με τη μεγαλύτερη στατιστική σημαντικότητα. Στη συνέχεια, για τη βελτιστοποίηση των πέντε σημαντικότερων παραγόντων εφαρμόσθηκε τροποποιημένη μέθοδος Simplex.Οι δύο μέθοδοι επικυρώθηκαν για τον ποσοτικό προσδιορισμό των 21 ελεύθερων αρωματικών συστατικών με αξιολόγηση ως προς την ειδικότητα, τη γραμμικότητα, τα όρια ανίχνευσης και ποσοτικοποίησης, την ανάκτηση, την αναπαραγωγιμότητα, την επαναληψιμότητα και την αβεβαιότητά. Στη συνέχεια, εφαρμόσθηκαν επιτυχώς, για την ανάλυση 20 λευκών μονοποικιλιακών οίνων από ελληνικές και διεθνείς ποικιλίες αμπέλου Vitis vinifera.

The concentration of yeast assimilable nitrogen (YAN) content in grape musts of selected representative vineyards of Nemea wine region was determined for 3 consecutive years. It was also of interest to investigate the effect of climatic conditions and vineyard altitude on YAN concentration. Amino acids were analyzed by High-Pressure Liquid Chromatography (HPLC), and 18 were identified and quantified. The majority of grape samples presented a deficiency in yeast assimilable nitrogen required for successful wine fermentation according to the literature. None of the samples had ammonia nitrogen content more than 2 mg N/L. The predominant amino acids quantified were arginine followed by proline, glutamine, and glutamic acid. Based on the ratio of arginine and proline, Agiorgitiko is probably weak or net arginine accumulator. A general tendency of increasing amino acid content with temperature was reported, while the opposite was evident for rainfall. Climatic conditions and cultivar are more likely the most important parameters affecting amino acid concentration in grapes.

β-D-glucans are proposed to have many health benefits. It is therefore important to have methods which can distinguish these from other carbohydrates present in natural products, as well as giving glucan content and structural information. Correlations between features in the CP/MAS spectra of β-D-glucans and enzyme assay determined β-D-glucan content were generally found to be poor. The β-D-glucan in dry and hydrated forms of the mushroom Ganoderma lucidum was investigated in detail by spectral peak fitting to the anomeric carbon C1 region in CP/MAS NMR spectra. Hydrated samples gave spectra with enhanced resolution and suggested that a clear distinction between β-D-glucans and other carbohydrates could be possible in the anomeric carbon C1 region. Chemical shift values for a range of carbohydrate polymers, which can be found alongside β-D-glucans, as well as the values for various linkages are given. Contamination by other carbohydrates and buffer salts is discussed.

&beta, Glucan, isolated from the mushroom Pleurotus ostreatus, at a concentration of 0.4%, was used in the manufacture of reduced-fat white-brined cheese from sheep milk. Control reduced-fat cheese was also produced from the same milk without the addition of &beta, glucan. The resultant cheeses were examined for their physicochemical characteristics, color and textural properties, and level of proteolysis and lipolysis. Furthermore, cheeses were evaluated organoleptically. In general, there were no statistical differences in the physicochemical characteristics and proteolysis levels found between both cheeses. The addition of &beta, glucan improved textural properties, and the cheeses received favorable grades for all the organoleptic characteristics. There were no flavor defects (such as a bitter taste) described by the panellists in this study. Generally, the addition of &beta, glucan did not significantly affect total free fatty acid content, however, at 180 days of ripening and storage, cheeses with the addition of &beta, glucan had a higher (p &lt, 0.05) content than cheeses without &beta, glucan. The major fatty acids were acetic acid and capric acid.

A critical comparison was made of seven solid-phase extraction (SPE) sorbents for the fractionation and isolation of 21 important free and glycosidically-bound varietal volatile aroma compounds. The sample was subjected to SPE and the free aromatics were eluted with dichloromethane followed by elution of the glucoconjugates with methanol; after fractionation, the free fraction was analyzed directly by GC-MS while the sugar-bound fraction was enzymatically hydrolyzed to liberate the free compounds before analysis by GC-MS. The extraction efficiency for the free compounds was evaluated based on the analytes’ signal recovery and for the glycosidically-bound compounds in terms of the relative peak areas. The best results for both the free and bound fractions were obtained with the Isolute ENV+ resin. Following selection of the most efficient SPE material, a GC-MS method was validated (in terms of selectivity, linearity, limits of detection (LODs) and limits of quantification (LOQs), recovery, repeatability, within-laboratory reproducibility and uncertainty) for the quantitative determination of the free primary volatiles in white wines. Validation results are presented at 4 fortification levels (10, 50, 200 and 500 µL−1). Regarding linearity, the correlation coefficient of the matrix-matched calibration plots was ≥0.99 for all the compounds. The LOQs were in the range 0.6–17.5 µg L−1. Recoveries ranged from 61% to 120% while the% relative standard deviation of the within-laboratory reproducibility was in the range 1.3% to 17.7%. Finally, the% expanded uncertainty ranged from 3.1% to 40.3%. The method has been successfully applied to the analysis of 20 white wine samples.

In this work, the influence of cluster thinning on the amino acids concentration of musts and wines was studied. Three vineyards were selected from Peza district of Crete where the white vine variety Vilana is cultivated. One part of each vineyard, containing 100 vines was divided in two sections. The first was used as reference while in the second section, cluster thinning to 50%, just before veraison, was applied. The experimentation was repeated for two continuous years. In 1996, the reference sections yielded 35.5 – 46.7 t/h while the other sections, where cluster thinning took place, yielded 16.5 – 23.0 t/h. The next year, the yields were 43.2 – 48.4 t/h and 12.6 – 14.0 t/h respectively. In all musts and wines, 18 amino acids were determined with the use of HPLC and proline was determined colorimetrically. The volatile components of the wines were also measured because, as it is known, the amino acids contribute to the formation of some higher alcohols, ethyl esters and acetates. From this work the following conclusions were made:The musts and the wines from Vilana variety are poor in amino acids and their levels vary from one year to another. This variation is related to the climatic conditions of the period of maturation but the profile of these acids seems invariable.The musts of Vilana variety deriving from vineyards with cluster thinning, which lead to low yield, contain higher levels of amino acids. This is important, given that a must rich in amino acids, positively influences not only the fermentation process evolution but also the enrichment of the wines in volatile components. Particularly for the Vilana variety, which is poor in amino acids and without any typical aroma, the high yields have a negative effect on their wine quality.

In this work, a solid-phase microextraction/gas chromatography–mass spectrometry (SPME/GC–MS) methodology was developed for the determination of 21 free and glycosidically-bound volatile varietal aroma compounds in wines. Initially, a comparison was made of 5 commercially available SPME fibers for the isolation/preconcentration of the target compounds in the headspace (HS) and direct immersion (DI) modes. The statistical significance of the microextraction variables was evaluated using a 2-level Plackett–Burman experimental design; the most significant variables were further optimized using a modified Simplex procedure. Using the selected conditions, a GC–MS method was fully validated for the quantitative determination of the 21 free primary aroma compounds. The hydrophilic bound precursors were isolated by solid-phase extraction (SPE), enzymatically hydrolyzed to liberate them as the free compounds and further detected by SPME/GC–MS. The method has been successfully applied to the analysis of 20 Greek white wine samples.

Aims: To study the population dynamics of indigenous malolactic bacteria in a Greek winery and to examine their potential to produce detrimental levels of biogenic amines (BA) under winemaking conditions. Methods and Results: Although the wines studied were of different vintage, grape variety and enological characteristics, molecular typing of malolactic bacteria revealed only a low number of strains within the single-species populations of Oenococcus oeni that developed during spontaneous fermentations. Strain MF1, originating primarily from the vineyards surrounding the winery invariably predominated in almost all samples. HPLC analysis showed a slight increase in the BA, putrescine, tyramine and phenylethylamine after malolactic conversion, while histamine, methylamine and ethylamine remained unaffected. No correlation could be established between the BA profiles and the bacterial compositions or the amino acid concentrations in wine samples studied. Conclusions: A certain regional bacterial flora is established in the winery that prevails in spontaneous malolactic fermentations (MLF) irrespective of the wine characteristics. In all cases, the BA content of the wines after malolactic conversion was within enologically acceptable levels. Significance and Impact of the Study: This is the first report on the malolactic bacteria occurring naturally in spontaneous MLF in Greek red wines and a preliminary assessment of their impact on wine safety in relation to BA.

Aims: To assess ochratoxin A (OTA) presence in Greek wines, according to the production area and the technology, as OTA risk is higher in wines from southern Europe. The effect of fining agents was examined in order to investigate OTA reduction.Methods and results: Dry and sweet wines issued from research programs and trade wines were analyzed, according to the official HPLC method. Greek dry wines do not exceed the European limit of 2 ng/mL. The amount of OTA (in ng/mL) varies between 0 - 1,18 for dry white wines, 0 -1,00 for dry red wines. Among sweet wines, only fortified wines without alcoholic fermentation range between 0,50 -7,64, while other sweet wines range between 0,05 -1,73. Dry yeast reduced OTA by 25 - 60 % according to dosage and duration (1-10 weeks) while the effect of bentonite was 33 - 40 % reduction for a treatment of 3-6 days.Conclusion: The picture for Greek wine production is positive, even in case of a future lowering of the European limit. OTA risk seems higher in the islands, particularly for some dessert wines. Results from wine treatment with fining agents support the theory of OTA adsorption onto suspended solids and the beneficial effect of yeast presence.Significance and impact of study: Assessment of OTA danger for Greek wines and information concerning the effect of fining agents contribute to the production of wines free from contaminants and support wine economy.