## 2 Analytical Methods for Phytocannabinoid Profiling
### Sample Preparation Techniques
\begin{table}
\begin{tabular}{l c c c} \hline \hline \multicolumn{1}{c}{**Sample Preparation Technique**} & \multicolumn{1}{c}{**Advantages**} & \multicolumn{1}{c}{**Disadvantages**} \\ \hline HS-SPME & - & programmable automated operation; & - & specific to GC-based methods only; \\ HS-SPME & - & improved chromatographic & - & applicable mostly for simple \\  & - & peak shape; & & matrices (herbal material) \\ \hline  & - & “green” extraction method; & & \\  & - & ensures stability of thermolabile & & \\  & & and light-sensitive & - & rarely used \\ SFE & & phytocannabinoids; & - & high price \\  & - & high extraction yields; & & \\  & - & ability to separate & & \\  & phytocannabinoids from terpenes & & & \\ \hline FUSE, UAE & - & low solvent and energy & - & applicable mostly for simple \\  & - & consumption & & matrices (herbal material) \\ \hline SPE & - & most suitable for food matrices & - & laborious and time-consuming \\  & - & “green” extraction technique & - & high price \\ \hline MHD & - & simultaneous extraction of terpenes & - & more commonly used for extraction \\  & & and phytocannabinoids & & of essential oils \\  & - & simultaneous decarboxylation & - & high price \\ \hline  & - & analyte extraction and & & \\  & & preconcentration in a single, & & \\ CPE & & solvent-free step & - & low extraction efficiency for \\  & - & avoidance of analyte loss during & & phytocannabinoids \\  & & solvent evaporation & - & time consuming \\  & - & low price & & \\ \hline CPC & - & allows for large-scale extraction of & - & high solvent consumption and \\  & & phytocannabinoids with high & & waste generation \\  & & efficiency & - & high price \\ \hline \hline \end{tabular}
\end{table} 
Table 2: _Cont._  
\begin{table}
\begin{tabular}{l c} \hline \hline \multicolumn{1}{c}{**Solvent/Solvent Mixture**} & \multicolumn{1}{c}{**References**} \\ \hline MeCN & [89; 90] \\ \hline MeCN + 1\% acetic acid & [65] \\ \hline MeCN saturated with n-hexane & [91] \\ \hline MeOH & [38; 84; 50; 54; 56; 68; 78; 79; 85; 89; 92; 93; 94; 95; 96; 97; 98; 99; 100; 101; 102; 103; 10  
Absolute EtOH is the most preferred organic solvent for maceration and LLE due to its great affinity for phytocannabinoid structure [54; 115; 146] that leads to high extraction efficiencies. EtOH is, however, known to co-extract significant amount of pigments and ballast from cannabis plant material, much more than CHCl\({}_{3}\), enhancing matrix interferences [44]. To avoid this, \(n\)-hexane is used [87]. For highly aqueous cannabis-based products, such as coffee beverages, 100% MeCN is preferred LLE solvent [90]. CHCl\({}_{3}\) is preferred for extraction of dried glandular plant material (leaves), cannabis resin and reefers (mixtures of tobacco with powdered resin or herbal cannabis), yielding 99% extraction efficiency, much higher than light petroleum [44]. Hexane was found to be the worst performer in terms of recovery of all phytocannabinoids, while EtOH was shown to possess the most appropriate polarity for cannabinoid compounds [40]. Prior to TLC/HPTLC analysis, maceration and/or LLE is the extraction technique of choice. Phytocannabinoids are extracted from _cannabis_ plant material using MeOH, according to Ph.Eur., Ph.Helv. and DAB [34; 35; 36] and published studies [147], dichloromethane, according to AHP [33], solvent of choice (petroleum ether, MeOH, \(n\)-hexane, toluene, CHCl\({}_{3}\) and solvent mixtures, e.g., MeOH/CHCl\({}_{3}\) (9:1, _v/v_)), according to UNODC [5]. Prior to maceration/LLE, lactose-containing cannabis products (cream cheese, butter, coffee beverages with milk) require lactase pre-treatment/lactose hydrolysis in order to avoid matrix interferences [90]. Maceration and LLE of semisolid fatty/oily matrices of cannabis-based products, such as butter, margarine, chocolate bars, nonpolar topical ointments and balms requires warming of the samples on a hot plate after addition of the extraction solvent in order to melt the matrix.