# Influence of Altitude on Phytochemical Composition of Hemp Inflorescence: A Metabolomic Approach
## 2 Results and Discussion
### Quantification of Cannabinoids by HPLC-Q-Exactive-Orbitrap(r)-MS Analysis  
Based on the available commercial cannabinoids standards, quantification of inflorescence extracts was performed by applying our validated method [13; 24; 25], as explained in detail in the material and methods section. Quantitative data related to the analysis of the content of phytocannabinoids in the two infloresesc groups performed by means of the HPLC high-resolution mass spectrometry (HRMS, Q-Exactive-Orbitrap(r)-MS) are shown in Table 2. The quantification was performed for CBD (cannabidiol), \(\Delta^{9}\)-THC (delta-9- tetrahydroccannabinol), CBN (cannabinol), CBC (cannabichromene), CBG (cannabigerol), CBDV (cannabidivarin), \(\Delta^{9}\)-THCV (delta-9-tetrahydroocannabivarin) and the acid forms CBDA (cannabidiolic acid), \(\Delta^{9}\)-THCA (delta-9- tetrahydroccannabinolic acid), CBNA (cannabinolic acid), CBCA (cannabichromenic acid), CBGA (cannabigerolic acid), CBDVA (cannabidivarin acid) and \(\Delta^{9}\)-THCVA (delta-9-tetrahydrocannabivarin acid). In contrast to what was found for volatile terpenoids, the results obtained for cannabinoids were more uniform with respect to the cultivation site. It was therefore feasible to perform paired statistical evaluations combining the specimens from both locations. Since the Kompolti chemovar belongs to the fiber-type hemp, it is not surprising that CBDA was the most abundant phytocannabinoid with significantly higher concentration in the mountain than in the plains. Those values (both for M and P group) are higher than those recently reported by [32; 33]. The THC and THCA content were below legal limits, but the occurrence of traces of CBNA --THCA's non-enzymatic, oxidative product--should be taken into consideration when so-called \"total THC\" amount is concerned. Furthermore, it is noted that the metabolic pathway of CBGA (considered as the progenitor/precursor of all cannabinoids) was activated towards the production of CBCA, which occurred in considerably 3 times higher quantities in the samples cultivated at high altitude. The presence of similar quantities of CBD in the mountain and plains specimens indicated that geographical location did not significantly influence decarboxylation of CBDA. This process naturally occurs under the action of heat and light, but here it is more probable that it was caused by [PERSON]'s predisposition to have prolonged flowering in the late summer/beginning autumn season, when the average daily temperatures are moderately higher in both locations. Therefore, a genetically predisposed flowering season may have led to a partial conversion of the parent CBDA into its neutral counterpart, as was demonstrated for Futura and Finola 75 varieties [13]. \begin{table}
\begin{tabular}{c c c c c c} \hline \hline  & \multicolumn{2}{c}{**Mountain**} & \multicolumn{2}{c}{**Plains**} & \multicolumn{2}{c}{**Statistical**} \\ \hline  & **Mean** & **SD (\(\pm\))** & **Mean** & **SD (\(\pm\))** & **Significance** \\ \hline \multicolumn{6}{c}{Neutral forms} \\ \hline CBD & 5300 & 3500 & 6000 & 3800 & ns \\ \(\Delta^{9}\)-THC & \textless{}LOQ & / & \textless{}LOQ & / & / \\ CBN & \textless{}LOQ & / & \textless{}LOQ & / & / \\ CBC & 460 & 120 & 120 & 50 & 0.005 \\ CBG & 110 & 10 & 180 & 80 & \textless{}0.001 \\ CBDV & 250 & 400 & 450 & 40 & \textless{}0.001 \\ \(\Delta^{9}\)-THCV & \textless{}LOQ & / & \textless{}LOQ & / & / \\ \hline \multicolumn{6}{c}{Acid forms} \\ \hline CBDA & 99,600 & 24,800 & 68,220 & 15,000 & 0.01 \\ \(\Delta^{9}\)-THCA & 840 & 200 & 1010 & 400 & ns \\ CBNA & 40 & 4 & 50 & 10 & ns \\ CBCA & 1570 & 200 & 570 & 30 & 0.008 \\ CBGA & 7410 & 900 & 4510 & 400 & 0.015 \\ CBDVA & 310 & 70 & 240 & 20 & ns \\ \(\Delta^{9}\)- THCVA & \textless{}LOQ & / & \textless{}LOQ & / & / \\ \hline \hline \multicolumn{6}{c}{LOQ–limit of quantification 1 ug/g for all phytocannabinoids. ns: not significative} \\ \hline \hline \end{tabular}
\end{table}