## 3 Discussion
For comparison to both non-destructive methods, a random 5 berry sample was collected, crushed, and analyzed under controlled laboratory conditions. Statistical analysis using PCA indicated that sampling by PVF chambers and modified glass vials detected similar VOC emission profiles across all 4 cultivars. There was 1 outlier from the glass vial method, indicating a higher than average concentration of styrene in La Crescent grapes. This data could provide evidence of styrene as a product of 2-phenylethanol synthesis from yeast cells [44] (p. 309) present during sampling. It should be noted that 2-phenylethanol variable is positively correlated with principal component 2, orthogonal to styrene. It is expected to have more VOCs detected at higher relative concentrations in crushedberry analysis because of the release of juices and volatiles bound within the berry skin and pulp. Research is warranted to compare headspace SPME analyses of crushed berries with conventional analytical methods such as liquid-liquid extraction [45]. Berry VOCs were sampled within a 3-month growing period each year for 2 years. VOC profile in 2012, sampled by PVF chambers and crushed berries were similar in profile. Data points from the PCA fall close to the origin throughout the year in 2012, not shown. This indicated that VOCs collected via PVF chambers did not show noticeable changes from veraison to harvest. VOC profile in 2013, sampled by modified glass vials and crushed berries were similar in profile (points near the origin) until the end of August, with the exception of the outlier on 14 August 2013, Figure 3. In La Crescent and St. Croix cultivars grown in Iowa, there is a movement towards higher than average VOC emission on 24 August 2016, Figure 4. In Iowa, VOC development was still trending above average at harvest on 3 August 2016 for St. Croix and on 29 September 2016 for La Crescent, not shown. VOCs emitted from Frontenac, Marquette, and St Croix cultivars grown in South Dakota started to develop and deviate from average later than Iowa on 29 August 2016, Figure 5. VOCs emitted from La Crescent grown in South Dakota start to trend above average on 3 September 2016, Figure 6. VOC emissions returned to average levels between 29 August and 5 September (harvest) in South Dakota Frontenac berries, Figure 7. The same decreasing trend was observed in South Dakota Marquette between 5 September and 8 September 2016, not shown. Similar to Iowa, the increased VOCs emitted from South Dakota St. Croix and La Crescent do not decline by harvest, not shown. Differences in microclimate of Iowa and South Dakota plots did not affect VOC emissions from 4 cold-hardy grape cultivars. Little difference in VOC emissions is expected from Marquette and Frontenac because of a shared pedigree. Greater changes in VOC emissions was observed between destructive crushed berry analysis and non-destructive in-vivo analysis methods, but not within the non-destructive methods. In Iowa and South Dakota plots, VOCs emitted from St. Croix and La Crescent cultivars continued to change from veraison through harvest. VOCs emitted in-vivo from Frontenac and Marquette cultivars in South Dakota started to decline 8 days and 3 days before harvest, respectively. More research is warranted in order to make recommendations to viticulturists regarding ideal harvest time for maximum aromas in the cold-hardy grapes. Linking correlations between viticultural practices can enhance the quality of wines for new cold-climate cultivars. Several improvements to the proposed in-vivo sampling are warranted. Addition of internal standard (IS) [46], for example a small vial with a membrane for controlled emission of IS during sampling (e.g., inside a PVF bag) would to ensure that sampling temperature and SPME fiber variables are controlled. This information would help to normalize sampling variables in field conditions and potentially help with data quality. Secondly, IS addition would enable quantification of volatiles.