*2.5. Profile of Volatile Compounds*

The ozonation process affected the chemical composition of the sea buckthorn berries. The group of compounds that are particularly susceptible to decomposition under the influence of this process are volatile compounds (Table 3). In sea buckthorn fruits, these compounds belong to main two groups: organic acid esters (compounds with a fruity smell) and terpene derivatives. The first group of compounds are aliphatic derivatives that are resistant to the oxidation process. HS-SPME analysis showed that the percentages of these substances vary depending on the ozonation conditions used, but their mutual proportions are similar. The differences in the percentage composition are due to the loss of other compounds that were most likely susceptible to the ozone decomposition process. The greatest differences were observed for carvacrol presence. Carvacrol is a

phenol derivative that was identified only in the control sample (non-ozonated fruit). This is an interesting observation because the aromatic compounds show a moderate tendency to ozonolysis [29]. The process carried out with the use of gaseous ozone mainly affects the surface of the ozonated raw material. The secondary metabolites are often found on the surface of the fruit in tissues capable of storing hydrophobic substances. They are often components of the cuticle, which is a barrier rich in hydrophobic compounds. Terpenes are hydrophobic substances that are often found in the cuticle. One of the components of the volatile fraction of sea buckthorn berries is carvacrol. Carvacrol is a component with a low boiling point and a relatively high vapor pressure, which makes the compound susceptible to the surface action of ozone. The HS-SPME technique used makes it possible to identify volatile compounds present in the headland phase. The lack of carvacrol in this phase in the case of ozonated fruit (Figure 3) indicates that it has been removed from the top layers of sea buckthorn berries, but this does not exclude its presence in deeper structures. It should be noted, however, that the lack of this substance in the headspace will likely affect the subjective smell of ozonated fruits. A cursory sensory analysis did not reveal the absence of foreign odors. Ozonolysis can lead to the formation of carbonyl derivatives, which often exhibit strong aromatizing properties. These compounds can arise as oxidation products of unsaturated fatty acids in the deeper layers of plant tissues [30,31]. However, they arise from long-chain unsaturated hydrocarbon derivatives. In the case of sea buckthorn berries, the aromatic hydrocarbon derivative (carvacrol) was degraded, and presumably degraded into highly volatile compounds (glyoxal), the presence of which was not detected. No significant losses of the esters, which are mainly responsible for the fruity fragrance, were observed during the ozonation process. As shown by Slynko et al. [32], these esters are the main ingredients in the scent of sea buckthorn berries and represent 71.45% of the ingredients of Hippophae scents. In the case of ozonated fruit, the proportion of these esters ranges from 97% (100 ppm, 30 min) to 60.98% for non-ozonated fruit.

**Figure 3.** Chromatogram SPME-GC for the volatile fraction of *Hippophae rhamnoides* L. fruit.




## **3. Materials and Methods**

## *3.1. Plant Material*

The research material was acquired from a local producer in a neighborhood of Rzeszów (southern Poland) after harvest. The plant material was analyzed at the Department of Food Chemistry and Toxicology University of Rzeszów.
