## 2 Analytical Methods for Phytocannabinoid Profiling
### Instrumental Analysis
#### 2.2.3 SFC Methods  
Despite being efficient, SFC is an analytical technique that has still not been fully exploited for the analysis, separation and quantification of cannabis plants and cannabis-based products, compared to GC and LC. The limited number of studies available report SFC as a fast (8-10 min runtime), cost-effective method with high specificity and separation power for phytocannabinoid profiling [195; 199; 200]. Prior to SFC analysis, derivatization and/or decarboxylation of phytocannabinoids is not required, thus reducing the risk of sample contamination (unlike GC). Additionally, SFC allows separation of the neutral from the acidic phytocannabinoids, simultaneously due to the properties of the supercritical fluids, offers shorter analysis time, better resolution and definitive identification in a single chromatogram of cannabis products when compared to both GC-MS and HPLC methods [199]. UHPSFC is shown to offer greater selectivity than UHPLC, but at the price of lower sensitivity, as a result of considerable variation of the refractive index of CO\({}_{2}\), resulting in greater baseline noise [188].  
Recently, the development of ultra-high performance SFC (UHPSFC) improved resolving power and efficiency, such that SFC has regained its popularity as an alternative phytocannabinoid profiling of cannabis plants and cannabis-based products. SFC combined with a 2 \(\upmu\)m particle size column offers rapid separation and when coupled to UV or MS detection, offers highly efficient analysis of the main phytocannabinoids, using inexpensive and environmentally friendly SC-CO\({}_{2}\) as solvent. On the other hand, UHPSFC is considered as highly orthogonal technique which provides different elution order and relative retention of the investigated components compared to UHPLC; therefore, in combination with MS, it could increase the discrimination power of phytocannabinoids in complex matrices.  
For the purpose of phytocannabinoid profiling, SFC using cyanopropyl silica packed column was employed [199], while the reported UHPLSFC methods used BEH 2-EP (2-ethylpyridine) column [195,200]. Evaluation of columns with different stationary phases, Torus 1-AA (1-aminoanthracene), Viridis BEH-2 EP (ethyl-pyridine) and Torus Diol (OH) revealed that the latter column achieved the highest number of (although not completely separated) peaks; thus, it is most appropriate for routine phytocannabinoid profiling [188]. SC-CO\({}_{2}\) is used with MeOH as co-solvent, with a constant (2%) [188] or gradually increasing concentration from 2% to 7% [199], often with the addition of 0.1% formic acid to improve peak shape of cannabinoid acids [188]. UHPSFC methods employ SC-CO\({}_{2}\) with isopropanol/MeCN (80:20, _v/v_) with 1% H\({}_{2}\)O and linear gradient [195,200]. SFC and UHPSFC are usually coupled either to PDA detectors [188] or to mass analyzers, such as APCI-QQQ [199] and ESI-Q mass analyser [195,200]. The reported linearity, sensitivity and specificity confirm the potential of SFC and UHPSFC to become the main profiling analytical platforms instead of GC- and LC-based methods in near future.