# An Optimized Terpene Profile for a New Medical Cannabis Oil
## 4 Discussion  
Here, we compared the concentrations of cannabinoids and terpenes of three MCOs obtained with three different extraction procedures in order to improve the therapeutic profile of the final product. In particular, quantitative analyses of terpenes were performed while several studies on MCOs did not investigate cannabis volatile compounds or performed qualitative or semi-quantitative approaches [31; 32]. For the development of the extraction protocol, this work mainly focused on temperatures and times of the decarboxylation and following steps. It is well known that the decarboxylation (pre-heating) of the starting plant material is necessary for the conversion from acid to neutral cannabinoid forms [34]. In fact, cannabinoid concentrations in the 3 MCOs (all methods including pre-heating of at least 100 \({}^{\circ}\)C) were found to be consistent with previous findings and in line with the quantities declared by the manufacturer (Bedrocan) in the plant material [30; 31; 32]. On the other hand, the application of high temperatures for a long time increased the risk of thermal volatile evaporation/degradation [35]. Thus, our working hypothesis was that an improved extraction method had to include a decarboxylation step resulting in high cannabinoid concentration and minimal loss of terpen. The innovative CERFIT method decarboxylated Cannabis flos at a moderate temperature (max 100 \({}^{\circ}\)C) in a microwave-assisted vacuum (to preserve the preparation from the external humidity) system: microwave was reported more effective than conventional extraction [35]. Moreover, we introduced a cyclical variation of the temperature in this preheating step in order to have rotation between heating (max 100 \({}^{\circ}\)C for 10 min) and cooling (100-25 \({}^{\circ}\)C for 25 min) and rest (25 \({}^{\circ}\)C for 55 min) reducing the thermal stress of the phyto-complex. In addition to SIFAP and CALVI procedures, a final step at \(-\)25 \({}^{\circ}\)C for 60 min was added to condense the evaporated components with no loss of volatile compounds. Moreover, it is important to highlight that our method permitted the full recovery of final MCO (i.e., starting from 5 g of Cannabis flos in 50 mL, we obtained 50 mL medical cannabis oil): this yield (100%) is given by a constant control of the filtration system applied before packaging. Finally, we could make a case for the good scalability of CERFIT method. In fact, we developed the present work starting from 5 g of Cannabis in 50 mL but we also conducted several experiments (data not shown) with major quantities (up to 30 g of Cannabis in 300 mL) maintaining the same Cannabis/matrix ratio (1:10) and the results overlapped both in terms of recovery and quali-quantitative cannabinoid and terpene profiles. A significant increasing trend resulted in the total terpene concentrations of the MCOs extracted with the three methods (SIFAP < CALVI < CERFIT). This finding could be also determined by the utilized extraction technique: ultrasound-assisted extraction was reported more effective than conventional procedures [36]. Moreover, in the CALVI method, ultrasounds were applied for 30 min at the water bath containing the flask with infloresences in MCT oil. CERFIT method used a probe sonicator inside the flask maintained in a refrigerated water bath. At the end of the extraction the water baths had a T of about 45 \({}^{\circ}\)C and 21 \({}^{\circ}\)C, respectively, and this difference could explain the major efficacy of CERFIT method since lower temperatures were reported to enhance the cavitation and consequently the yield of the extracted bioactive compounds [37]. Interestingly, total sesquiterpenes increased in MCO extracted with CERFIT method while \(\beta\)-caryophyllene concentration decreased. Thus, the extraction method appeared to influence the qualitative composition of the final oil product as well as the quantitative profile of volatile compounds. Remarkably, the final composition of MCO-2 and MCO-3 was further varied.