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Background and Aims: Winemakers from a commercial winery observed sensory differences in Cabernet Sauvignon wines made from three pruning treatments in a single vineyard, particularly in mouthfeel characteristics. This study examined the relationships between wine composition and wine sensory characteristics, then related these to berry weight and composition and wine quality scores. Methods and Results: Cabernet Sauvignon from three pruning treatments � Machine, Cane and Spur � was harvested at commercial harvest date, and replicate wines were made from each for three vintages. The composition of the wines from all three pruning systems was generally similar. Differences in individual descriptive attributes did not separate the wines from the three treatments, or across vintages, despite differences in overall quality scores. Principal component analysis (PCA) could separate the wines by pruning and by vintage using wine composition or sensory parameters. Higher concentrations of anthocyanins, tannins and phenolics in berries did not always result in higher concentrations in wines. Conclusions: In this study, higher wine tannin or wine phenolic concentrations did not result in higher wine astringency, and wine colour measures and phenolic composition were not good indicators of individual wine sensory properties or wine quality. Wine composition was not necessarily directly influenced by berry composition. Significance of the Study: Few studies focus on the berry to wine to sensory continuum, particularly over more than one vintage or in a commercial context. This study highlighted how complex the relationships among berries, wine sensory properties and wine quality can be, particularly within a single vineyard.

Background and Aims: Commercial winemakers observed differences in wine sensory properties among wines produced from vines of three different pruning treatments in a single vineyard. To clarify the relationships between berries and wine quality, this study examined berry size and berry composition and the quality rating for wines produced from the pruning treatments. Methods and Results: The study was conducted over three seasons. Berries from Machine-, Cane- and Spur-pruned vines were sampled at commercial harvest for analysis of berry size and berry phenolic composition. Wines made from each treatment were assessed for quality by a panel of winemakers. Machine berries were lighter and had higher concentrations of anthocyanins, tannins and total phenolics than Cane or Spur. Machine wines had the lowest quality scores. Comparing vintages, berries from 2004 were lighter, but did not always differ in phenolic composition to other vintages, and wines from 2004 had lower quality scores than the other vintages. Vintage effects were generally stronger than pruning effects. Conclusions: In this study, high berry anthocyanins, total phenolics and tannin concentration measures were not good indicators of wine quality scores. Changes in vineyard treatments and, in particular, vintage influences, produce incremental, but potentially important, changes to berry size and composition and to wine quality. Significance of the Study: This study is unique in its focus on the commercial reality of producing wines to a specific style and quality, while bringing scientific rigour to investigating the relationships between berries and wine quality in this vineyard over a number of vintages.

An obscure sesquiterpene, rotundone, has been identified as a hitherto unrecognized important aroma impact compound with a strong spicy, peppercorn aroma. Excellent correlations were observed between the concentration of rotundone and the mean 'black pepper' aroma intensity rated by sensory panels for both grape and wine samples, indicating that rotundone is a major contributor to peppery characters in Shiraz grapes and wine (and to a lesser extent in wine of other varieties). Approximately 80% of a sensory panel were very sensitive to the aroma of rotundone (aroma detection threshold levels of 16 ng/L in red wine and 8 ng/L in water). Above these concentrations, these panelists described the spiked samples as more 'peppery' and 'spicy'. However, approximately 20% of panelists could not detect this compound at the highest concentration tested (4000 ng/L), even in water. Thus, the sensory experiences of two consumers enjoying the same glass of Shiraz wine might be very different. Rotundone was found in much higher amounts in other common herbs and spices, especially black and white peppercorns, where it was present at approximately 10000 times the level found in very 'peppery' wine. Rotundone is the first compound found in black or white peppercorns that has a distinctive peppery aroma. Rotundone has an odor activity value in pepper on the order of 50000-250000 and is, on this criterion, by far the most powerful aroma compound yet found in that most important spice.

In a polar fraction of an extract obtained from quince fruit juice capillary gas chromatography and capillary gas chromatography-mass spectrometry revealed the occurrence of 4-hydroxy-7,8-dihydro-β-ionone. Biomimetic studies with another 4-oxygenated ionone derivative from quince fruit, were carried out. Heat treatment of this diol gave a number of volatile C 13 norisoprenoids. After preparative HPLC separation 1 H and 13 C NMR analyses revealed the structure of the main isomer. This component was also detected as trace constituent in quince fruit juice

Mousy off-flavor is an insidious and economically disastrous microbiologically derived spoilage characteristic of wine and other fermented beverages. Tainted wines are rendered unpalatable and there is currently no satisfactory procedure for removal of the off-flavor. Here we report the confirmation of that both d- and l-lysine can act as a precursor for the formation of mousy off-flavor N-heterocycles. Further, through the use of stable isotope feeding experiments, we could establish that a pentylamine group from lysine is incorporated into the piperideine moiety of two off-flavor N-heterocycles. A biochemical pathway for the formation of mousy off-flavor compounds is proposed.

'Black pepper' aroma and flavor is important to some Australian Shiraz red wine styles but the aroma compounds involved have yet to be identified, and no objective analytical method to assess 'pepper' grape aromas is available to date. Samples of potentially 'spicy'/'peppery' grapes were obtained from vineyards in South Australia and Victoria over two vintages. The important sensory attributes of the grapes, including the aroma descriptor 'pepper', were rated by a sensory panel. The sensory study revealed a strong correlation between the intensity of 'pepper' aroma and the intensity of 'pepper' flavor perceived on the palate. The grape homogenates were analyzed by static headspace GC-MS using a cool inlet system. Vectors obtained by analysis of over 13 000 individual mass spectra per grape sample were then subjected to multivariate analyses. Both principal component analysis and partial least-squares regression were used to develop multivariate models based on mass spectra and aroma descriptors to explain the intensity of the rating of the 'pepper' character. Corresponding differences in mass spectra and aroma were observed among vineyards and from the same vineyards in different years. Additional optimization of the methodology enabled selection of a single region of the GC-MS chromatogram that allowed prediction of 'pepper' aroma intensity with a correlation coefficient0.98 and led to the identification of alpha-ylangene, a tricyclic sesquiterpene. To assess the potential of alpha-ylangene as a marker for this sensory characteristic, a method for alpha-ylangene analysis of grapes and wine using HS-SPME-GC-MS was developed. Although not a significant aroma compound by itself, alpha-ylangene was a satisfactory marker for the 'pepper' aroma in grapes and wine, and its concentration showed similar discrimination between 'peppery' vineyards and vintages as that obtained using the multivariate models. Despite its presence in grapes, we could not detect alpha-ylangene in wine.

The modern wine industry needs tools for process control and quality assessment in order to better manage fermentation or bottling processes. During wine fermentation it is important to measure both substrate and product concentrations (e.g. sugars, phenolic compounds), however, the analysis of these compounds by traditional means requires sample preparation and in some cases several steps of purification are needed. The combination of visible/near-infrared (Vis/NIR) spectroscopy and chemometrics potentially provides an ideal solution to accurately and rapidly monitor physical or chemical changes in wine during processing without the need for chemical analysis. The aim of this study was to assess the possibility of combining spectral and multivariate techniques, such as principal component analysis (PCA), discriminant partial least squares (DPLS), or linear discriminant analysis (LDA), to monitor time-related changes that occur during red wine fermentation. Samples (n = 652) were collected at various times from several pilot scale fermentations with grapes from either Cabernet Sauvignon or Shiraz varieties, over three vintages (2001-2003) and scanned using a monochromator instrument (Foss-NIRSystems 6500, Silver Spring, MD) in transmission mode (400-2,500 nm). PCA was used to demonstrate consistent progressive spectral changes that occur through the time course of the fermentation. LDA using PCA scores showed that regardless of variety or vintage, samples belonging to a particular time point in fermentation could be correctly classified. This study demonstrates the potential of Vis/NIR spectroscopy combined with chemometrics, as a tool for the rapid monitoring of red wine fermentation.

From the glycosidic extract of pinuela fruit; 1-O-β- d -glucopyranosyl anthranilate was isolated by adsorption chromatography on XAD-2, followed by rotation locular counter-current chromatography, multilayer coil counter-current chromatography and HPLC. Identification was carried out by NMR ( 1 H, 13 C, 1 H 1 H and 13 C 1 H COSY) and HPLC-MS/MS under electrospray ionization.

Motivated by the identification of numerous novel tetrahydro-beta-carboline-carboxylic acids in food samples, we studied the reactions of tetrahydro-beta-carbolines in the presence of nitrosating agents. The anticipated formation of nitroso derivatives from unsubstituted tetrahydro-beta-carbolines, and from tetrahydro-beta-carboline-3-carboxylic acids was indicated by HPLC-MS/MS analysis and validated by the characteristic product ion spectra of the respective nitroso compounds. In addition, oxidative decarboxylation resulted in formation of the corresponding dihydro-beta-carbolines, and in the generation of the beta-carbolines harman or norharman. Subsequently, we studied the reactivity of tetrahydro-beta-carboline-1-carboxylic acids derived from the Pictet-Spengler condensation of indole amines with alpha-oxo acids. Again, in the presence of nitrosating agents the rapid disappearance of the starting material was obvious, but no nitroso derivatives could be observed. Instead, further HPLC-MS/MS studies demonstrated that dihydro-beta-carbolines were the major products of tetrahydro-beta-carboline-1-carboxylic acids. Finally, we demonstrated that freshly isolated nitroso-precursors spontaneously decomposed to yield harman alkaloids. In conclusion, we revealed that nitroso-tetrahydro-beta-carbolines can represent intermediates involved in the generation of beta-carbolines, and we established a novel pathway for the formation of harman alkaloids from nutritional tetrahydro-beta-carbolines.

Human Vgamma9delta2 T lymphocytes are suggested to play an important role in the immune response to various microbial pathogens. In contrast to alphabeta T cells, gammadelta T lymphocytes recognize small, non-protein, phosphate-bearing antigens (phosphoantigens) in a major histocompatibility complex-independent manner. Four different phosphoantigens termed TUBag1 to TUBag4 with a common 3-formyl-1-butyl-pyrophosphate moiety and isopentenyl-pyrophosphate have been isolated and identified from mycobacteria. However, natural occurring gammadelta T cell ligands from other bacterial species were not characterized so far. Here, we describe the structural identification of the two compounds responsible for the gammadelta T cell-stimulating capacity of Escherichia coli as similar to the mycobacterial phosphoantigens 3-formyl-1-butyl-pyrophosphate and its M(r) 275 homologue TUBag2. In addition, E. coli phosphoantigens exert bioactivities on gammadelta T cells with similar potencies to the mycobacterial phosphoantigens at 5-15 nm concentration. Furthermore, our results clearly prove that the deoxyxylulose 5-phophate pathway (also referred to as Rohmer metabolic route of isoprenoid biosynthesis) is essential for the biosynthesis of the phosphoantigens in E. coli. Because this pathway is absent from human cells, it proves an ideal target for focusing efficiently the antimicrobial selectivity of human gammadelta T lymphocytes.

The formation of various carbohydrate-derived beta-carbolines was observed when model reactions of tryptophan with glucose were studied by means of HPLC with diode array detection, as well as by means of HPLC-MS. Isolation of these compounds and subsequent characterization by tandem mass spectrometry and NMR spectroscopy led to the identification of diastereomeric 1-(1,3,4,5-tetrahydroxypent-1-yl)-9H-pyrido[3,4-b]indoles (1a/b), 1-(1,4,5-trihydroxypent-1-yl)-9H-pyrido[3,4-b]indoles (2a/b), and E/Z isomers of 1-(1,5-dihydroxypent-3-en-1-yl)-9H-pyrido[3,4-b]indole (3a/b). HPLC-MS was used to prove the presence of these novel beta-carboline alkaloids in various food samples. In addition, quantitative determination of beta-carbolines 1a, 1b, and 2a/b in numerous products was achieved by means of HPLC with fluorometric detection. Concentrations ranged from 12 to 1922 microg/L for 1a and 1b and from 3 to 644 microg/L for 2a/b. The highest concentrations of all carbohydrate-derived beta-carbolines under study were found in ketchup, soy sauce, and fish sauce.

After intake of food or herbal medicinal products containing quercetin glycosides, the systemic availability of the genuine glycoside, as well as the systemic occurrence of the aglycone or conjugates of this polyphenol has been a matter of dispute. Consequently, we designed this study to develop a reliable method for determination of quercetin and its metabolites. Following consumption of fried onions five different glucuronides of quercetin could be identified in human plasma samples by means of HPLC-UV-MS/MS. Selective determination of the target compounds was achieved by simultaneous UV (254 nm) and MS/MS detection with selected reaction monitoring experiments using positive mode electrospray ionisation. In contrast, neither the free flavonol nor the genuine glycoside could be detected in plasma. Identification of the quercetin glucuronides detected in vivo was confirmed by comparison with authentic reference compounds synthesised enzymatically using glucuronyl transferase from rabbit liver.

In extracts prepared from various fruits as well as in fruit juices a single tryptophan glycoconjugate was detected by HPLC-MS analysis. Product ion spectra demonstrated the N-glycosidic linkage of a hexose moiety to the indole nitrogen. For structure elucidation, the novel tryptophan glycoside was isolated from pear juice and identified as N1-(β-d-glucopyranosyl-4C1)-l-tryptophan by 1H, HH−COSY and 13C NMR spectroscopy. Finally, we disclosed the biosynthetic origin of the novel tryptophan metabolite by demonstrating the enzymatic glycosylation of deuterium-labeled tryptophan, which was applied to pear fruit. Keywords: Tryptophan-N-glucoside; electrospray ionization; HPLC-MS/MS; biosynthesis

Tetrahydro-s-carboline (THC)-carboxylic acids are readily formed by Pictet-Spengler condensation of tryptophan or tryptamine with various carbonyl compounds. In plants, mammals, and by chemical oxidation studies, it has been demonstrated that THC-carboxylic acids represent precursors of s-carboline (e.g. harman) alkaloids (Herbert and Mann, 1980; Rommelspacher and Susilo, 1985; Bobbitt and Willis, 1980). With respect to Parkinson’s disease, generation of potent neurotoxins from s-carbolines by enzymatic N-methylation has been reported (Collins et al., 1992).

Isopentenyl diphosphate (IPP), an important precursor of isoprenoid biosynthesis in prokaryotic and eukaryotic organisms, has been shown to activate Vgamma9/Vdelta2 T cells, the major subset of human gammadelta T cells. The biosynthesis of IPP has been first described as the acetate/mevalonate pathway. Recently, 1-deoxy-D-xylulose 5-phosphate (DOXP) and 2-C-methyl-D-erythritol 4-phosphate have been shown to be key metabolites in the DOXP pathway also leading to the formation of IPP in some eubacteria such as Escherichia coli. Here we report that the low molecular mass fraction of extracts from bacteria using the DOXP pathway induces Vgamma9/Vdelta2 T cell activation, while analogous preparations from bacteria using the classical mevalonate pathway fail to do so. Addition of 1-deoxy-D-xylulose potentiates the ability of E. coli extracts to activate Vgamma9/Vdelta2 T cells. As the amounts of IPP present in the bacterial preparations are not sufficient to induce significant Vgamma9/Vdelta2 T cell activation, our data suggest that compounds other than IPP associated with the DOXP pathway are responsible for Vgamma9/Vdelta2 T cell activation.

The synthesis of different cycloSal-phosphotriesters of the acyclic nucleoside analogues acyclovir (ACV), penciclovir (PCV) and T-penciclovir (T-PCV) as potential new lipophilic, membrane-soluble pronucleotides is described. The introduction of the cycloSal moiety was achieved by using reactive cyclic chlorophosphane reagents. In addition to the cycloSal-PCV monophosphate (MP) phosphotri-esters, a second derivative bearing an acetyl group at the second primary alcohol function was prepared. In hydrolysis studies the cycloSal-ACVMPs showed the expected range of hydrolytic stability dependent on the substituent in the masking group (8–17 h). In contrast, the cycloSal-PCVMP derivatives exhibited a 11- to 15-fold increase in hydrolytic lability as compared to the corresponding cycloSal-ACVMP derivatives. We demonstrated that the free primary alcohol group is responsible for this rate acceleration because cycloSal-OAc-PCVMP, in which the hydroxyl group was blocked by acetylation, did not show the aforementioned acceleration. Unexpectedly, the hydrolysis product was not PCVMP but according to NMR and mass spectrometry it was cycloPCVMP (cPCVMP). The title compounds were evaluated in vitro for their ability to inhibit herpes simplex virus type 1 (HSV-1) and thymidine kinase-negative (TK–) HSV-1 replication in Vero cells. The cycloSal-ACVMP compounds exhibited high antiviral activity in HSV-1-infected cells. More importantly, one derivative retained all activity from the wild-type virus strain in HSV-1/TK–-infected Vero cells. The PCV derivatives were markedly less active. The reason for the failure of the cycloSal-PCVMPs seems to be due to the formation of cPCVMP instead of the desired PCVMP.

Peroxidase active preparations from three Ancistrocladus species and from Triphyophyllum peltatum have been partially purified. They catalyze the oxidative dimerization of korupensamines A and B to michellamines A and C, respectively, as well as the mixed coupling to michellamines A, B, and C. All of these enzymes consist of single polypeptides of approximately 65 kDa with peroxidase activity as monomers. They were characterized as soluble proteins predominantly localized in the leaf phloem of all species examined. Comparative studies with various naphthol precursors revealed an unexpected substrate specificity. Only korupensamines were dimerized by the enzymes, while other monomers, even if closely related, were not accepted as substrates. The coupling reactions described here represent the first direct synthesis of michellamines from korupensamines without previous protection of these precursors.

A rapid and sensitive method is described for the detection of 1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid, 1-methyl-1,2,3,4- tetrahydro-beta-carboline-3-carboxylic acid and 1-methyl-1,2,3,4-tetrahydro-beta-carboline by electrospray ionization tandem mass spectrometry coupled to liquid chromatography. In combination with selected reaction monitoring (SRM) detection limits of 3 ng ml-1 (ca. 75 fmol on column) were established by the use of model solutions. Due to the excellent selectivity and sensitivity of SRM no sample preparation step was required prior to analysis of food samples. In addition, any artifactual formation of tetrahydro-beta-carbolines could be excluded. Application of the method revealed that all food samples analyzed contained both tetrahydro-beta-carboline-carboxylic acids at ng ml-1 to microgram ml-1 concentrations, whereas 1-methyl-1,2,3,4-tetrahydro- beta-carboline was identified in most samples as a minor constituent.

2,5-Dimethyl-4-hydroxy-3[2H]-furanone 6′-O- malonyl-β- d -glucopyranoside was isolated from a glycosidic extract of strawberry fruit (Fragaria x ananassa, cv. Senga Sengana) by means of countercurrent chromatography and reverse-phase HPLC. Identification was achieved by comparison of chromatographic and 1H, 13C and 2D-NMR, as well as mass spectral data with those of the synthesized reference compound. In ripe strawberry fruit, a 1:1 ratio of the malonylated glucoside to its deacylated glucoconjugate was determined by on-line liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry.

A sensitive and selective method to determine diazepam, N-desmethyldiazepam, nitrazepam, flunitrazepam and medazepam in human serum and urine was established employing solid-phase extraction and high-performance liquid chromatography-electrospray tandem mass spectrometry (HPLC-ESI-MS-MS) with the selected reaction monitoring (SRM) mode. Using diazepam-d 5 and N-desmethyldiazepam-d 5 as internal standards, recoveries between 90.4 and 109.7% were determined. The routine quantification limit was set at 2 ng/ml for serum and urine at a signal-to-noise ration of 10:1.

A hydrolysis study of 3-methyl-cycloSal-PCVMP 2 is described. Surprisingly, phosphotriester 5 released in this study not the expected PCVMP, but cycloPCVMP.

A new method was developed for the analysis of heterocyclic aromatic amines (HAA) using HPLC-ESI-MS/MS with selected reaction monitoring (SRM). Detection limits between 0.1 and 50 ng/mL were achieved.

Long-chain alkadien(trien)ylphenols, i.e., cardanols, cardols, and anacardic acids, as well as their acetylated and methylated derivatives were used in enzymatic dioxygenation catalyzed by soybean lipoxygenase-1 (LOX-1). Kinetic studies revealed the influence of functional groups, chain length, number of double bonds, and the distance of the double bonds from the terminal end. Free functional groups were found to be a structural prerequisite for high affinity and turnover number; exceptional kinetic parameters were obtained for ω-6 compounds possessing the same chain length as linoleic acid. The regioselectivity and enantioselectivity of LOX-1 catalysis were investigated with 3-[(Z,Z)-8‘,11‘-pentadecadienyl]phenol as the representative substrate, yielding 3-[12‘-(S)-hydroperoxy-(Z,E)-8‘,10‘-pentadecadienyl]phenol as the main product. Our extensive chromatographic and spectroscopic studies comprised HPLC−MS/MS for differentiation of the regioisomers, 1H NMR spectroscopy for determination of the double-bond...