## 3 Discussion  
The effect of vineyard practices on grape and wine aroma merit study. The claims to a regions' wines by sensory attributes need to be scientifically correlated to bolster the local economies. Otherwise consumers are inundated with marketing claims. This study attempted to compare microclimates of Iowa and South Dakota during 3 months of the growing season, over 2 years. The Iowa plot is in USDA plant hardness zone 5a [39]. In comparison, the South Dakota plot is in USDA plant hardness zone 4b [39]. These metrics were obtained from the National Oceanic and Atmospheric Administration, formerly the National Climatic Data Center [40]. Preliminary analyses using ANOVA were also completed (shown in 'ANOVA in-vivo' spreadsheet, Supplementary Materials). Type 1 sum of squares analysis indicated statistical significance of method, cultivar, date and time, method and cultivar interaction, and method and site and cultivar interactions (shown in Table 11). Post-hoc Tukey HSD test is (Table 12) shows differences within a method, cultivar, date and time, method and cultivar interaction, and method and site and cultivar interactions. PCA analyses were used to determine key volatile compounds emitted. Although significant differences were noted using ANOVA and Tukey HSD for total VOCs emitted, the more detailed analysis with PCA (focused on individual VOCs) accounted for less than 70% variance. This was the case for all but one case (Figure 5) of Frontenac grown in South Dakota in 2013. Statistical analysis using PCA indicated that Frontenac and Marquette were most similar in total VOC emission profile (i.e., clustered around the origin). St. Croix cultivars had a higher positive correlation with the first principal component. Seventeen VOCs with correlation \(\geq\)0.300 are listed in a section on statistical analysis. Any differences in the soil and microclimate of these two sites affected overall VOCs emitted from La Crescent and St. Croix cultivars during this research. It is cautioned that these differences could also be affected by the genetics of the cold-hardy hybrids. Frontenac and Marquette share similar parentage [41]. The advantages of the modified glass vials over PVF film chambers are its compact design for field sampling, reusability, reduced background contamination from glass, and isolation of VOCs emitted from a single berry. Vacuum-assisted headspace SPME sampling has been used in carefully controlled laboratory settings, to successfully achieve shorter sampling times at lower sampling temperature with good sensitivity and precision to extract polychlorinated biphenyls (PCB) from water [42]. This novel sampling device was the logical next step to isolate VOCs emitted from grape berries during development. This sampling technique is comparable to a viticulturalist'smelling' a grape, and detecting only the volatile compounds emitted through the grape skin. These VOCs are recognized as \"free\" aroma compounds not bound to a sugar moiety within the berry [43]. This could allow for monitoring of VOCs to measure berry ripeness by instrumental methods. Grapes get softer as they develop, and some cultivars are prone to slip skin (i.e., the grape skin slips easily from the fruit pulp). A disadvantage in using modified glass vials for grape sampling in this research is that increased vacuum was needed as the grapes developed and softened, and sometimes broke the grape skin, more often in the St. Croix cultivar. Another confounding element could be the presence and interference of volatile compounds on the grape skin but not produced by the grape (i.e., pesticide residues, naturally occurring yeasts and molds). Non-destructive, sampling of VOCs emitted in-vivo from cold-hardy grapes was conducted using 2 methods. PVF film sampling chambers with custom SPME sampling port was used to monitor whole cluster VOC emissions. Modified glass vials supported SPME sampling of individual berries.