Anna Katharine Mansfield

A core–shell column approach to a comprehensive high-performance liquid chromatography phenolic analysis of Vitis vinifera L. and interspecific hybrid grape juices, wines, and other matrices following either solid phase extraction or direct injection

Four high-throughput reverse-phase chromatographic protocols utilizing two different core-shell column chemistries have been developed to analyze the phenolic profiles of complex matrices, specifically targeting juices and wines produced from interspecific hybrid grape cultivars. Following pre-fractionation via solid-phase extraction or direct injection, individual protocols were designed to resolve, identify and quantify specific chemical classes of compounds including non-anthocyanin monomeric phenolics, condensed tannins following acid hydrolysis, and anthocyanins. Detection levels ranging from 1.2 ppb to 27.5 ppb, analyte %RSDs ranging from 0.04 to 0.38, and linear ranges of quantitation approaching five orders of magnitude were achieved using conventional HPLC instrumentation. Using C(18) column chemistry, the non-anthocyanin monomeric protocol effectively separated a set of 16 relevant phenolic compounds comprised flavan-3-ols, hydroxycinnamic acids, and flavonols in under 14 min. The same column was used to develop a 15-min protocol for hydrolyzed condensed tannin analysis. Two anthocyanin protocols are presented, one utilizing the same C(18) column, best suited for anthocyanidin and monoglucoside analysis, the other utilizing a pentafluorophenyl chemistry optimized to effectively separate complex mixtures of coexisting mono- and diglucoside anthocyanins. These protocols and column chemistries have been used initially to explore a wide variety of complex phenolic matrices, including red and white juices and wines produced from Vitis vinifera and interspecific hybrid grape cultivars, juices, teas, and plant extracts. Each protocol displayed robust matrix responses as written, yet are flexible enough to be easily modified to suit specifically tailored analytical requirements.

A next-generation marker genotyping platform (AmpSeq) in heterozygous crops: a case study for marker-assisted selection in grapevine

Marker-assisted selection (MAS) is often employed in crop breeding programs to accelerate and enhance cultivar development, via selection during the juvenile phase and parental selection prior to crossing. Next-generation sequencing and its derivative technologies have been used for genome-wide molecular marker discovery. To bridge the gap between marker development and MAS implementation, this study developed a novel practical strategy with a semi-automated pipeline that incorporates trait-associated single nucleotide polymorphism marker discovery, low-cost genotyping through amplicon sequencing (AmpSeq) and decision making. The results document the development of a MAS package derived from genotyping-by-sequencing using three traits (flower sex, disease resistance and acylated anthocyanins) in grapevine breeding. The vast majority of sequence reads (⩾99%) were from the targeted regions. Across 380 individuals and up to 31 amplicons sequenced in each lane of MiSeq data, most amplicons (83 to 87%) had <10% missing data, and read depth had a median of 220–244×. Several strengths of the AmpSeq platform that make this approach of broad interest in diverse crop species include accuracy, flexibility, speed, high-throughput, low-cost and easily automated analysis.

Identification of Odor-Impact Compounds in Red Table Wines Produced from Frontenac Grapes

Frontenac (Vitis spp. MN 1047) is a recently introduced, cold-hardy red winegrape that is currently the most-planted cultivar in much of the Upper Midwest. Through descriptive analysis, a set of aroma attributes common to red Frontenac table wines has been described, but the volatile compounds responsible for the characteristic sensory notes of the product have not been investigated. In order to identify these odor active compounds, eight Frontenac table wines were evaluated using stir bar sorptive extraction (SBSE) combined with concurrent gas chromatography/olfactometry-mass spectrometry (GC/O-MS). Eight panelists evaluated GC/O effluent using qualitative detection frequency analysis. Twenty-four volatiles were identified in odor regions perceived by panelists, including five alcohols, 14 esters, one lactone, two acids, and two volatile phenols. Twenty-three of these were confirmed by linear retention index data in separate GC-MS analyses, and 23 were quantified in runs using a known concentration of internal standard. Similar analyses of wines produced from V. riparia clone #89, a parent of Frontenac, found 16 volatiles common to Frontenac wines. A brief study of Frontenac juice with two days of skin contact suggested that four volatiles found in the wine may originate in the fruit.

Impact of Processing Parameters on the Phenolic Profile of Wines Produced from Hybrid Red Grapes Maréchal Foch, Corot noir, and Marquette

Phenolic extraction in hybrid and interspecific wine grape cultivars is poorly understood, especially in terms of the impact of fermentation and enological conditions on condensed tannins and anthocyanins. Following fractionation via solid-phase extraction and high-performance liquid chromatography, phenolic profiles of must and wine from red hybrid grape cultivars Maréchal Foch, Corot noir, and Marquette were examined to assess the impact of enzyme and tannin addition, cold soak, and hot press during vinification. Across cultivars, hot press treatments resulted in the greatest extraction of condensed tannin, anthocyanin, and other monomeric phenolic compounds in musts, and treatments that increased skin contact time or cellular degradation during fermentation produced higher concentrations of tannins, anthocyanins, and flavonols. However, these increases were transient, evincing incomplete carryover into finished wines. Depending on initial must extraction, diglucoside forms of anthocyanins were either selectively extracted or selectively retained throughout fermentation when compared to their monoglucoside counterparts. Typical of hybrid grapes, tannin concentrations across cultivars were low, even under hot press conditions. For condensed tannins and anthocyanins, a cultivar-specific, stable-state concentration and phenolic profile emerged regardless of fermentation conditions. Due to the high levels of diglucoside anthocyanins and low levels of condensed tannins, it is expected that the color development and profile in these wines produced from hybrid grape cultivars will be dictated by the monomeric anthocyanins and their potential role in copigmentation processes involving other monomeric phenolic species, as opposed to the formation of polymeric color pigments.

Next Generation Mapping of Enological Traits in an F2 Interspecific Grapevine Hybrid Family.

In winegrapes (Vitis spp.), fruit quality traits such as berry color, total soluble solids content (SS), malic acid content (MA), and yeast assimilable nitrogen (YAN) affect fermentation or wine quality, and are important traits in selecting new hybrid winegrape cultivars. Given the high genetic diversity and heterozygosity of Vitis species and their tendency to exhibit inbreeding depression, linkage map construction and quantitative trait locus (QTL) mapping has relied on F1 families with the use of simple sequence repeat (SSR) and other markers. This study presents the construction of a genetic map by single nucleotide polymorphisms identified through genotyping-by-sequencing (GBS) technology in an F2 mapping family of 424 progeny derived from a cross between the wild species V. riparia Michx. and the interspecific hybrid winegrape cultivar, ‘Seyval’. The resulting map has 1449 markers spanning 2424 cM in genetic length across 19 linkage groups, covering 95% of the genome with an average distance between markers of 1.67 cM. Compared to an SSR map previously developed for this F2 family, these results represent an improved map covering a greater portion of the genome with higher marker density. The accuracy of the map was validated using the well-studied trait berry color. QTL affecting YAN, MA and SS related traits were detected. A joint MA and SS QTL spans a region with candidate genes involved in the malate metabolism pathway. We present an analytical pipeline for calling intercross GBS markers and a high-density linkage map for a large F2 family of the highly heterozygous Vitis genus. This study serves as a model for further genetic investigations of the molecular basis of additional unique characters of North American hybrid wine cultivars and to enhance the breeding process by marker-assisted selection. The GBS protocols for identifying intercross markers developed in this study can be adapted for other heterozygous species.

Quantifying the Contribution of Grape Hexoses to Wine Volatiles by High-Precision [U13C]-Glucose Tracer Studies

Many fermentation volatiles important to wine aroma potentially arise from yeast metabolism of hexose sugars, but assessing the relative importance of these pathways is challenging due to high endogenous hexose substrate concentrations. To overcome this problem, gas chromatography combustion isotope ratio mass spectrometry (GC-C-IRMS) was used to measure high-precision (13)C/(12)C isotope ratios of volatiles in wines produced from juices spiked with tracer levels (0.01-1 APE) of uniformly labeled [U-(13)C]-glucose. The contribution of hexose to individual volatiles was determined from the degree of (13)C enrichment. As expected, straight-chain fatty acids and their corresponding ethyl esters were derived almost exclusively from hexoses. Most fusel alcohols and their acetate esters were also majority hexose-derived, indicating the importance of anabolic pathways for their formation. Only two compounds were not derived primarily from hexoses (hexanol and isobutyric acid). This approach can be extended to other food systems or substrates for studying precursor-product relationships.

Accumulation and Prediction of Yeast Assimilable Nitrogen in New York Winegrape Cultivars

A three-year study was conducted to investigate the accumulation of yeast assimilable nitrogen (YAN) during the final weeks of ripening in seven winegrape cultivars grown in New York State and to assess the feasibility of predicting harvest YAN using linear regression models. Berry samples of Vitis vinifera cvs. Cabernet franc, Chardonnay, Merlot, Pinot noir, and Riesling and interspecific hybrid cvs. Noiret and Traminette were collected weekly from 49 vineyard sites across the Finger Lakes, Hudson Valley, Lake Erie, and Long Island growing regions from August through harvest. YAN concentrations in Cabernet franc, Riesling, and Traminette were generally low, averaging <100 mg/L annually, while Chardonnay and Pinot noir had average YAN concentrations >200 mg/L. During the ripening period, linear regression models predicted harvest YAN up to five weeks before harvest (R2 = 81.6%). A decrease in YAN during ripening was observed across cultivars, caused primarily by decreases in ammonia (AMM), as primary amino nitrogen levels remained stable. Population distributions were used to estimate appropriate prophylactic nitrogen additions for each cultivar, minimizing the risk of deficiency or excess; this was most difficult with Chardonnay, Noiret, and Pinot noir, which had the highest and most variable YAN concentrations and subsequently run the greatest risk of oversupplementation with prophylactic additions.

Reaction Kinetics of Monomeric Anthocyanin Conversion to Polymeric Pigments and Their Significance to Color in Interspecific Hybrid Wines

The color stability of red wines produced from interspecific hybrid grapes, which is partially dependent on anthocyanin diglucosides, is not well understood. In this study, the rate of decrease of monomeric anthocyanins as they polymerized to polymeric pigments due to the presence of excess catechin and acetaldehyde was measured in model wine using HPLC. Colorimetry was used to measure L*, a*, and b* values, hue angle, and change in color (ΔE). Concentrations of individual diglucosides decreased more slowly than monoglucosides. When monoglucosides and diglucosides were combined, the reaction rate of monoglucosides was slower than that of monoglucosides alone. Hue angles described transitions from red to red-orange, orange, or orange-yellow as anthocyanin-specific changes occurred. The evolution in color represents dynamic reactions between anthocyanins, catechin, and acetaldehyde. Consequently, wines containing high concentrations of diglucosides, such as those produced from interspecific hybrid grapes, will form less polymeric pigment than wines containing primarily monoglucosides.

Preharvest Prediction of Yeast Assimilable Nitrogen in Finger Lakes Riesling Using Linear and Multivariate Modeling

A three-year study was conducted to determine if regression models could be developed to predict yeast assimilable nitrogen (YAN) before harvest, using Riesling in the New York Finger Lakes region as a model. Berry samples were taken from 62 commercial Riesling vineyards around the Finger Lakes at veraison, two weeks before harvest, and harvest. Samples were measured for berry weight, Brix, pH, titratable acidity, ammonia, primary amino nitrogen, and yeast assimilable nitrogen (YAN). The average YAN concentration at harvest was 91.8 mg/L, and there were no significant differences in harvest YAN concentration among years. Linear regression models created using preharvest YAN concentrations (p < 0.05) had a cross-validated R2 (Q2) of 70%. Models using only preharvest ammonia had less predictive power (Q2 = 63%) but may allow winemakers more analytical flexibility than those requiring complete YAN measurements. Models created using multiple linear regression provided better predictive power (Q2 = 73.6%). Finally, a multivariate approach using partial least squares regression was used to create models with the highest predictive power (Q2 = 74.2%). The additional analysis required to obtain values for additional prediction variables may limit the practicality of multiple linear regression and partial least squares approaches. Because many winemakers are not able to perform the analyses required to calculate YAN during the busy time of harvest, the development of these regression models as predictive tools may allow winemakers to use preharvest analysis to calculate accurate supplemental nitrogen additions, allowing targeted supplementation and lowering the risk of excessive prophylactic additions.

Effects of Growth Temperature and Postharvest Cooling on Anthocyanin Profiles in Juvenile and Mature Brassica oleracea

The effects of growth temperatures on anthocyanin content and profile were tested on juvenile cabbage and kale plants. The effects of cold storage time were evaluated on both juvenile and mature plants. The anthocyanin content in juvenile plants ranged from 3.82 mg of cyanidin-3,5-diglucoside equivalent (Cy equiv)/g of dry matter (dm) at 25 °C to 10.00 mg of Cy equiv/g of dm at 16 °C, with up to 76% diacylated anthocyanins. Cold storage of juvenile plants decreased the total amount of anthocyanins but increased the diacylated anthocyanin content by 3-5%. In mature plants, cold storage reduced the total anthocyanin content from 22 to 12.23 mg/g after 5 weeks of storage in red cabbage, while the total anthocyanin content increased after 2 weeks of storage from 2.34 to 3.66 mg of Cy equiv/g of dm in kale without having any effect on acylation in either morphotype. The results obtained in this study will be useful for optimizing anthocyanin production.