## 2 Results
### Comparison of the Chemical Composition of Green and White Asparagus in 2020  
To validate and follow-up on the results from the first year's analyses, in 2020, the same green and white varieties were harvested from the same field as in 2019. All varieties were harvested twice, with three weeks in between, to investigate potential genotype and harvest time-point differences. The sample preparation, the analyses, the examination and processing of the raw data were all carried out in the same way as in 2019.  
PCA was performed separately on the complete 2020 datasets, both for the GC-MS and the LC-MS data. The QCs again indicate good reproducibility of the analyses (average RSD based on compounds present in QCs < 20%) and the green and white asparagus samples are again separated along PC1 (Figure 4). Further statistical analyses showed the discriminatory metabolites for the two asparagus types of the 2020 season which corresponded to 64% and 75% of the detected VOCs and non-volatiles, respectively (Tables S3 and S6). In agreement with the 2019 data, aldehydes and alcohols were highly abundant in the green varieties while white spears were characterised with high levels of a few other aldehydes. The differences in the levels of all benzenoids and monoterpenoids between green and white asparagus were less distinct based on the differential PC1 values (Figure S2a). With regard to the non-volatiles, the metabolites separating the green and white spears comprised polyphenols and saponins, respectively (Figure S2b), confirming the findings of 2019. Additionally, PCA was performed on the combined 2019 and 2020 GC-MS data on the 61 identified volatiles detected in both seasons verifying the distinct differences in the volatile composition of green and white spears (Figure S3).  
Figure 3: PCA score plots based on (a) 84 volatiles and (b) 442 non-volatiles detected in the green and white asparagus spears harvested in 2019: the first two PCs are presented, and the explained variance is shown in parentheses on the axes. Colours indicate the asparagus type; symbol shapes indicate asparagus variety.  
The separation of the green varieties based on their volatile and non-volatile profiles along PC2 that was observed in 2019 data, was less evident in the 2020 data (Figure 4). Further analyses comparing the profiles of the three varieties were performed, separately for the green and white asparagus and only 6 volatiles and 0 non-volatiles were found at significantly different levels (adjusted \(p\) value \(\leq\) 0.05) between the three varieties in either the green or white type (Figures A1 and A2, Tables S2 and S5).  
Within each asparagus type ca. 25% of the total variation in the profiles of volatiles and non-volatiles was caused by the harvest time-point, which also appeared to be type dependent (Figures A1 and A2 in Appendix A). The compounds significantly changing in calendar week 23 compared to week 20 are 2-methylfuran, toluene, 2-methoxy-3-isopropylprazine, 2-methoxy-3-isobutylpyrazine and methanethiol for the white spears and chlorogenic acid, one asparagusic acid ester isomer, heptanal, 3-methylhiopropanol (methanol), nonanal and octanal for the green spears (Tables S2 and S5).