## 3 Results and Discussion
### Volatile Compounds
#### 3.1.3 Horseradish  
A total of 75 compounds were identified or tentatively identified in the headspace of horseradish roots, 38 of which are newly reported (Table 2). As with wasabi, the peaks with the highest relative abundances were dominated by GHPs. Compounds with the highest relative abundances were allyl ITC (**5/6**, 7.4%/39.3%), phenethyl ITC (**24**, 32.5%), thiuraneacetonitrile (**77**, 3.6%), 4-pentenyl ITC (**14**, 2.3%), (Z)-3-hexen-1-ol (**31**, 1.9%), and _sec_-butyl ITC (**9**, 1.8%; Figure 2). As stated in Section 3.1.2, allyl ITC was identified as two distinct peaks (**5** and **6**). As in wasabi, **6** was of the greatest abundance and odour intensity, producing a very strong, pungent, garlic, mustard, and horseradish-like aroma (Table 3), whereas **5** produced a medium intensity, pungent horseradish smell. Thus far, the presence of two peaks has not been addressed or explained satisfactorily within the literature, with only one previous paper reporting the same phenomenon of separate and distinct allyl ITC peaks [71]. _Sec_-butyl ITC (**9**) produced a medium-weak intensity aroma that was vegetative and radish-like (Table 3). The compound was also present in wasabi at a medium intensity. The compound has been previously reported in horseradish as having green, chemical, and mustard like aromas [21], and it is known to activate the human Transient Receptor Potential Ankyrin 1 (TRPA1). This receptor is known to act in response to environmental irritants, and several ITCs identified in this study are known to activate it to varying degrees (isopropyl ITC, **3**; isobutyl ITC, **10**; allyl ITC, **5/6**; 3-butenyl ITC, **11**; 4-pentenyl ITC, **14**; benzyl ITC, **22**; phenylethyl1 ITC, **24**; [72]). Phenylethyl ITC (**24**) is known to be a key constituent of horseradish aroma, and our data are in agreement with previous reports [73]. Assessors described the compound as radish and gooseberry-like, with a sweet note. It had a high odour intensity and contributed significantly to the odour profile of roots. Likewise, 4-pentenyl ITC (**14**) was observed to have the same odour attributes as previous reports [1] and those found for wasabi in this study, but at a lower intensity. By contrast, pentyl ITC (**15**) was present at much lower relative intensities to other GHPs (0.2%, Table 2) but produced a strong, green, rotten, and cabbage-like aroma. Thiiraneacetonitrile (**77**) has been previously reported in horseradish [22] and is an epithonitrile hydrolysis product of singorin. To our knowledge, no previous studies have described the odour of this compound. We found it to have a sweaty, gas-like aroma of medium intensity (Table 3). 2-Isopropyl-3-methoxypyrazine (**102**, 0.1%) and 2-_sec_-butyl-3-methoxypyrazine (**106**, 0.4%) have been previously described and characterised in horseradish roots [25] as having green and pepper-like aromas. Our data agree with previous reports but found the compounds to be of very high aroma intensity, despite relatively low abundances within the headspace (Table 3). Assessors described the compounds as rotten, earthy, potato-like, and vegetative. We report several compounds previously unidentified in horseradish including GHPs, isocyanates, alcohols, aldehydes, and a ketone and ester. As in wasabi root, cyclopropane ITC (7, 0.1%) produced an intense aroma containing horseradish, garlic, onion, and sulphur notes (Table 3). Therefore, it is likely to be a significant contributor to root odour and the volatile profile, despite its very low abundance, which may be a reason why it has not been previously detected and/or reported. As discussed in Section 3.1.2, it is unknown if the presence of isocyanates is linked with GSLs and their hydrolysis products. Allyl isocyanate (**99**, <0.1%) and phenethyl isocyanate (**107**, 0.4%) were both observed for the first time in horseradish roots. Given that high abundances of allyl ITCs (**5/6**) and phenethyl ITC (**24**) were observed, it seems likely that isocyanates may be derived from them and/or directly from parent GSLs.