**3. Results and Discussion**

As reported in the literature, the major constituents of cuticular waxes extracted by SFE processing plus high-pressure fractionation of the vegetable matter were paraffinic compounds and, among them, heptacosane, nonacosane, hentriacontane and tritriacontane showed the larger percentages [32–38]. Moreover, odd-carbon-atom hydrocarbons were largely more represented than the homologous, even-carbon atoms. This is an interesting characteristic from an applicative point of view; indeed, differently from paraffins coming from fossil fractions, odd-carbon-atom hydrocarbons are largely more compatible with the human skin and can be applied in cosmetics and health care products [9,39–42].

A photograph of the cuticular waxes extracted during SFE processing of cannabis inflorescence is reported in Figure 1. In all cases, the precipitated material looks like a white powder, sometimes with a light smell, similar to that of the starting vegetable species.

**Figure 1.** Macroscopic and qualitative example of cuticular waxes extracted by SFE processing of cannabis inflorescence.

Eleven GC-MS traces of the produced cuticular waxes are summarized in Figure 2, for overall comparison purposes. These traces can give a qualitative perspective of the compounds present in the various plants tested and their relative abundance.

**Figure 2.** Comparison of GC−MS traces of cuticular waxes extracted by SC−CO2 plus fractional separation from different vegetable matter, studied in this work.

Extensive identification of the cuticular waxes extracted and analyzed in this work is reported in Table 1. In particular, Figure 2 and Table 1 confirm that the most abundant compounds present in the cuticular waxes extracted by SC-CO2 are C27, C29 and C31 for all the vegetable species studied. These results are in agreement with the previous literature related to the same vegetable matter [32–39]. However, data in the literature are referred only to straight paraffins. Analysis performed in this work demonstrates, instead, the presence of some high-molecular-weight paraffinic alcohols (namely, C24, C26, C28 and C30). The largest percentages of these compounds are found in marjoram (16.22%), tobacco (5.94%) and lavender (5.48%). Additionally, aldehydes and traces of a paraffinic acid, namely octacosanoic acid, are detected in jasmine and tobacco. More specifically, C28 and C30 aldehydes are the most widespread compounds and the largest percentage of C28 aldehyde is found in tobacco (6.57%) and marjoram (6.40%).

The prevalence of long-chain alkanes is confirmed, and they range from C23 to C33, with a prevalence of odd paraffins, as C27, C29 and C31, that largely confirm as the major components, though their relative proportions vary from one vegetable species to another [43,44]. Some small quantities of paraffinic alcohols, aldehydes and fatty acids are also identified, as expected. They all show the same carbon atoms' skeleton of the identified paraffins, with the further presence of a functional group: i.e., alcoholic, aldehydic or acid group.

*Separations* **2022**, *9*, 80



#### **4. Conclusions**

In the present work, a detailed study on the composition of cuticular waxes extracted and fractionated by SFE is reported. GC-MS analysis confirmed that the separation from the other extractable materials was accurate, and these products can be considered a sort of fingerprint of the specific vegetable matter. C27, C29 and C31 were the most abundant compounds found in the investigated vegetable materials, in line with the previous findings reported in the literature. Moreover, the specific selectivity of SC-CO2 extraction towards non-polar or slightly polar compounds makes these cuticular waxes suitable for higher added-value applications, such as in the medical and pharmaceutical field, in which purity and biocompatibility are key features that justify the selection of a more complex extraction process with respect to the traditional ones.

**Author Contributions:** Conceptualization, E.R.; methodology, M.S.; validation, L.B. and E.R.; formal analysis, M.S.; investigation, L.B. and M.S.; resources, E.R.; data curation, L.B. and M.S.; writing—original draft preparation, L.B. and E.R.; writing—review and editing, L.B.; supervision, E.R. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Conflicts of Interest:** The authors declare no conflict of interest.
