Gavin L. Sacks

Effects of cluster light exposure on 3-isobutyl-2-methoxypyrazine accumulation and degradation patterns in red wine grapes ( Vitis vinifera L. Cv. Cabernet Franc).

The effects of light exposure on 3-isobutyl-2-methoxypyrazine (IBMP) accumulation and degradation in Vitis vinifera L. cv. Cabernet Franc berries were assessed by comparison of shaded and exposed clusters within the same vine throughout a growing season. Twenty-seven vines were shoot-thinned to create regions of high and low cluster-light exposure within each vine. Samples were collected at 10 time points starting from 5 to 130 days postbloom. The experimental design allowed for intravine comparison of IBMP levels between treatments at each time. Vine-to-vine variability of IBMP and the correlation of IBMP to malic acid were also evaluated. Cluster exposure reduced accumulation of IBMP at all preveraison time points by 21-44%, but did not increase postveraison degradation. Significant vine-to-vine variability in IBMP content was observed, with the highest level of IBMP in shaded berries in the most vigorous block of vines. Although IBMP concentration by weight decreased significantly due to dilution just prior to color change (veraison), no significant IBMP degradation per berry occurred until after color change (day 70 postbloom). By contrast, malic acid degradation began prior to color change, and malic acid concentrations were not affected by cluster exposure preveraison, but were affected postveraison. A survey of 13 sites in New York state (Seneca Lake) showed that IBMP concentrations at 2 weeks preveraison were highly correlated (R(2) = 0.936, p < 0.0001) to levels at harvest, whereas classic grape maturity indices at harvest were uncorrelated with IBMP at harvest. In summary, light exposure conditions critically influence IBMP accumulation but not IBMP degradation.

Comprehensive Two-Dimensional Gas Chromatography Combustion Isotope Ratio Mass Spectrometry

We report the first coupling of comprehensive two-dimensional gas chromatography (GC x GC) to online combustion isotope ratio mass spectrometry (C-IRMS). A GC x GC system, equipped with a longitudinally modulated cryogenic system (LMCS), was interfaced to an optimized low dead volume combustion interface to preserve <300 ms full width at half-maximum (fwhm) fast GC peaks generated on the second GC column (GC2). The IRMS detector amplifiers were modified by configuration of resistors and capacitors to enable fast response, and a home-built system acquired data at 25 Hz. Software was home-written to handle isotopic time shifts of less than one bin (40 ms) and to integrate peak slices to recover isotope ratios from cryogenically sliced peaks. The performance of the GC x GCC-IRMS system was evaluated by isotopic analysis of urinary steroid standards. Steroids were separated by a nonpolar GC1 column (30 m x 0.25 mm, 5% phenyl), modulated into multiple 4- or 8-s cryogenic slices by the LMCS, and then separated on a polar GC2 column (1 or 2 m x 0.1 mm, 50% phenyl). GC2 peak widths from a 1-m column averaged 276 ms fwhm. Steroid standard sliced peaks were successfully reconstructed to yield delta(13)C VPDB values with average precisions of SD(delta(13)C) = 0.30 per thousand and average accuracies within 0.34 per thousand, at 8 ng on column. Two steroids, coeluting in GC1, were baseline separated in GC2 and resulted in delta(13)C VPDB values with average precisions of SD(delta(13)C) = 0.86 per thousand and average accuracies within 0.26 per thousand, at 3 ng on column. Results from this prototype system demonstrate that the enhanced peak capacity and signal available in GC x GC is compatible with high-precision carbon isotope analysis.

Rapid measurement of 3-alkyl-2-methoxypyrazine content of winegrapes to predict levels in resultant wines.

We describe an optimized protocol for analysis of the herbaceous smelling 3-alkyl-2-methoxypyrazines (MPs) in whole berries that predicts MPs in resultant red wines. Berries are homogenized by bead-milling with a deuterated standard prior to headspace solid phase microextraction (HS-SPME) and quantification by two-dimensional gas chromatography time-of-flight-mass-spectrometry (GCxGC-TOF-MS). In the case of 3-isopropyl-2-methoxypyrazine (IPMP), GCxGC-TOF-MS successfully resolved interferences that coeluted with the analyte in the first dimension. HS-SPME parameters (pH, queue time, incubation time, extraction time, extraction temperature) were optimized by a statistical experimental design. Good method accuracy was observed (consistent ratio of unlabeled analyte to labeled standard) at 10 min extraction times when 80 degrees C extraction temperatures were employed, although increasing sensitivity was observed for longer extraction times (up to 140 min). Standard addition of 3-isobutyl-2-methoxypyrazine (IBMP) and IPMP into preveraison and harvest ripe berry matrices showed good linearity (r(2) >0.99 in all cases), with limits of detection ranging from 0.6 to 1.8 pg/g. The protocol was validated by comparing IBMP in 16 lots of Cabernet Franc berries (range = undetectable to 18.4 pg/g) to the resulting wines (range = undetectable to 14.5 pg/g). Berry and wine MP content were strongly correlated, (r(2) = 0.97, p < 0.0001). Following correction for CO(2) loss, the observed concentration of IBMP in wines was 67 +/- 13% of the IBMP concentration observed in berries.

Timing of Cluster Light Environment Manipulation during Grape Development Affects C13 Norisoprenoid and Carotenoid Concentrations in Riesling

Sunlight exposure of winegrape clusters is frequently reported to increase C(13)-norisoprenoids in resulting wines, but the timing and mechanism of this influence is not well understood. Fruit zone leaf removal was applied to Vitis vinifera cv. Riesling at three timings: 2, 33 and 68 days past berry set (PBS), and compared to an untreated control. Free and total 1,1,6-trimethyl-1,2-dihydronaphthalene (TDN), vitispirane and beta-damascenone were measured in juice and wines, and carotenoid profiles were determined in grapes at midseason and maturity. Significantly higher total TDN was observed in grapes from the 33-day PBS treatment compared to the control and other treatments (195 microg/L vs 54-87 microg/L). Total vitispirane in juice was also significantly increased in the 33-day PBS treatment, while total beta-damascenone was reduced in the 68-day PBS treatment compared to the control. Existing HPLC protocols were modified to allow for quantification of zeaxanthin in V. vinifera berries, and zeaxanthin was determined to be significantly higher in the 33-day PBS treatment than the control or other treatments (p < 0.05). Total TDN in juice correlated with free TDN in wine, with 11.0% +/- 2.5% of total juice TDN converted to free TDN in wine. In contrast, total vitispirane increased significantly during fermentation, and was not correlated with vitispirane in juice. In summary, leaf removal at 33 days PBS significantly increased zeaxanthin in Riesling grapes midseason, total TDN and vitispirane in the juice of mature Riesling grapes, and free and total TDN in finished wine, while earlier or later leaf removal had no effect.

Comparison of odor-active compounds in grapes and wines from vitis vinifera and non-foxy American grape species.

Native American grape (Vitis) species have many desirable properties for winegrape breeding, but hybrids of these non-vinifera wild grapes with Vitis vinifera often have undesirable aromas. Other than the foxy-smelling compounds in Vitis labrusca and Vitis rotundifolia , the aromas inherent to American Vitis species are not well characterized. In this paper, the key odorants in wine produced from the American grape species Vitis riparia and Vitis cinerea were characterized in comparison to wine produced from European winegrapes (V. vinifera). Volatile compounds were extracted by solid-phase microextraction (SPME) and identified by gas chromatography-olfactometry/mass spectrometry (GC-O/MS). On the basis of flavor dilution values, most grape-derived compounds with fruity and floral aromas were at similar potency, but non-vinifera wines had higher concentrations of odorants with vegetative and earthy aromas: eugenol, cis-3-hexenol, 1,8-cineole, 3-isobutyl-2-methoxypyrazine (IBMP), and 3-isopropyl-2-methoxypyrazine (IPMP). Elevated concentrations of these compounds in non-vinifera wines were confirmed by quantitative GC-MS. Concentrations of IBMP and IPMP were well above sensory threshold in both non-vinifera wines. In a follow-up study, IBMP and IPMP were surveyed in 31 accessions of V. riparia, V. rupestris, and V. cinerea. Some accessions had concentrations of >350 pg/g IBMP or >30 pg/g IPMP, well above concentrations reported in previous studies of harvest-ripe vinifera grapes. Methyl anthranilate and 2-aminoacetophenone, key odorants responsible for the foxiness of V. labrusca grapes, were undetectable in both the V. riparia and V. cinerea wines (<10 μg/L).

Impact of Shoot and Cluster Thinning on Yield, Fruit Composition, and Wine Quality of Corot noir

Cluster thinning (CL), shoot thinning (ST), and a combination of the two practices (ST+CL) were applied to vigorous Corot noir (hybrid sp.) winegrapes grown at a commercial vineyard in the Finger Lakes region of New York State. Yield was reduced by CL (by up to 4.8 kg/vine) but not ST in 2008, and by ST (by up to 2.0 kg/vine) and not CL in 2009; however, high pruning weights (up to 4.5 kg/vine in 2008) and low crop-load ratios (ranging from 2.3 to 7.1) indicated that vines in the study were undercropped regardless of treatment. CL increased soluble solids by up to 2.5 Brix in 2008 and by up to 0.8 Brix in the ST+CL treatment in 2009, while ST increased soluble solids in 2009 by 0.8 Brix. The treatments had variable impacts on wine anthocyanin, berry skin tannin, berry seed tannin, and wine tannin depending on year. Wine tannin (42 to 64 mg/L) and tannin extractability (5 to 6%) were both very low compared to values typically observed in red wines produced from V. vinifera. Using a two-alternative forced choice test, panelists reported ST+CL wines were fruitier than the control and ST wines and that ST wines were less fruity than the control in both years. An economic analysis indicated that in order for growers/wineries to maintain their economic welfare, bottle prices would have to increase by

Impact of Shoot Thinning and Harvest Date on Yield Components, Fruit Composition, and Wine Quality of Marechal Foch

0.02 to

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

0.41 depending on the practice and year to compensate for additional labor costs and lost yield in implementing these crop-load management practices.

Calibration and Data Processing in Gas Chromatography Combustion Isotope Ratio Mass Spectrometry

Marechal Foch grapevines were subjected to shoot thinning (~15 shoots per meter of row and no shoot thinning) in combination with two harvest dates (early harvest and late harvest) in a factorialized treatment arrangement for two years (2007 and 2008). With shoot thinning, yields were reduced by 3.1 to 7.2 kg per vine and clusters were reduced by up to 59 clusters per vine, while berry weight increased by 0.03 to 0.09 g. Shoot thinning reduced crop load by 4.3 to 7.8 kg yield per kg pruning weight, and increased soluble solids in 2008 by 0.7 to 1.2 Brix. Shoot thinning increased berry anthocyanins by 1.25 to 2.24 mg/g fresh skin weight malvidin-3-glucoside, but no corresponding increase was observed in wine anthocyanins. Delaying harvest resulted in increases of soluble solids (0.5 to 2.3 Brix) and berry anthocyanins (0.32 to 1.48 mg/g) and significantly higher anthocyanins in finished wines. Both late harvest and shoot-thinning treatments resulted in decreased six-carbon alcohols (3 to 33%) in finished wines. The total concentration of tannin in Foch fruit was comparable to that of some vinifera (0.75 to 1.05 mg/berry catechin equivalents). However, the extractability of tannins during winemaking was very low compared to most vinifera (2 to 4%), in part likely due to the low skin tannin concentration. Using a two-alternative forced choice test, panelists reported that later harvest 2008 wines were more “fruity” than their early harvest counterparts for both treatments and that shoot thinning did not affect fruitiness.

Protein-Precipitable Tannin in Wines from Vitis vinifera and Interspecific Hybrid Grapes (Vitis ssp.): Differences in Concentration, Extractability, and Cell Wall Binding

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.