## 2 Materials and Methods
### NMR Analyses  
The dried organic fraction of each sample was dissolved in 0.7 mL of a CDCl\({}_{3}\)/CD\({}_{3}\)OD mixture (2:1 \(v\)/\(v\)) and then placed into a 5 mm NMR tube. Finally, the NMR tube was flame sealed. Conversely, the dried hydroalcoholic phase of each sample was solubilized in 0.7 mL 400 mM phosphate buffer/D\({}_{2}\)O containing 1 mM solution of TSP as internal standard and then transferred into a 5 mm NMR tube. NMR spectra of all hydroalcoholic and organic extracts were recorded at 27 \({}^{\circ}\)C on a Bruker AVANCE 600 spectrometer operating at the proton frequency of 600.13 MHz and equipped with a Bruker multinuclear z-gradient 5 mm probe head. \({}^{1}\)H spectra were referenced to methyl group signals of TSP (\(\delta\) = 0.00 ppm) in D\({}_{2}\)O and to the residual CHD\({}_{2}\) signal of methanol (set to 3.31 ppm) in CD\({}_{3}\)OD/CDCl\({}_{3}\) mixture. \({}^{1}\)H spectra of hydroalcoholic extracts were acquired with 256 transients with a recycle delay of 5 s. The residual HDO signal was suppressed using a pre-saturation. The experiment was carried out by using 45\({}^{\circ}\) pulse of 6.5-7.5 us, 32 K data points. \({}^{1}\)H spectra of extracts in CD\({}_{3}\)OD/CDCl\({}_{3}\) were acquired with 256 transients, recycle delay of 5 s, and 90\({}^{\circ}\) pulse of 9-11 us, 32 K data points. The two-dimensional (2D) NMR experiments, such as \({}^{1}\)H-\({}^{1}\)H TOCSY, \({}^{1}\)H-\({}^{13}\)C HSQC, and \({}^{1}\)H-\({}^{13}\)C HMBC, were carried out under the same experimental conditions previously reported [24]. The integrals of 25 selected signals in hydroalcoholic extract (Table 1) were measured using the Bruker TOPSPIN software and normalized with respect to the resonance at 0.00 ppm, due to methyl group signal of TSP, normalized to 100. Results have been expressed in mg/100 g FW. The integrals of 6 selected signals in organic extract (Table 2) were also measured using the Bruker TOPSPIN software and normalized with respect to the resonance at 2.30 ppm, due to signal of total fatty acids, set to 100. The molar % values of fatty acids and sterols were calculated using the following equations:  
\[\%_{\rm\beta\text{-}SIT}=100\times 0.66I_{\rm\beta\text{-}SIT}/I_{\rm tot} \tag{1}\]  
\[\%_{\rm STIG}=100\times 0.66I_{\rm STIG}/I_{\rm tot} \tag{2}\]  
\[\%_{\rm TRI}=100\times 0.5I_{\rm TRI}/I_{\rm tot} \tag{3}\]  
\[\%_{\rm DI}=100\times I_{\rm DI}/I_{\rm tot} \tag{4}\]  
\[\%_{\rm MONO}=100\times(I_{\rm LNS}-2I_{\rm Di}-1.5I_{\rm TRI})/I_{\rm tot} \tag{5}\]  
\[\%_{\rm SAT}=100\times(I_{\rm FA}-I_{\rm DI}-0.5I_{\rm TRI}-I_{\rm MONO})/I_{ \rm tot} \tag{6}\]  
where \(\%_{\rm\beta\text{-}SIT}\), \(\%_{\rm STIG}\), \(\%_{\rm TRI}\), \(\%_{\rm OD}\), \(\%_{\rm MONO}\), \(\%_{\rm SAT}\) are molar % of \(\beta\)-sitosterol, stigmasterol, tri-unsaturated fatty acids, di-unsaturated fatty acids, mono-unsaturated fatty acids, and saturated fatty acids, respectively. I\({}_{\rm\beta\text{-}SIT}\), I\({}_{\rm STIG}\), I\({}_{\rm TRI}\), I\({}_{\rm DIS}\), I\({}_{\rm FA}\) are integrals (see Table 2), and I\({}_{\rm tot}\) is calculated according to the Equation (7)  
\[\rm I_{tot}=I_{\rm FA}+0.66I_{\rm\beta\text{-}SIT}+0.66I_{\rm STIG} \tag{7}\]  
\begin{table}
\begin{tabular}{c c c c} \hline
**ppm** & **Compounds** & **ppm** & **Compounds** \\ \hline
0.96 & Leu & 3.21 & Choline \\
0.99 & Val & 3.25 & \(\beta\)-Glucose \\
1.01 & Ile & 4.04 & Fructose \\
1.33 & Lact acid & 4.31 & Malic Acid \\
1.34 & Thr & 4.59 & \(\beta\)-Galactose \\
1.49 & Ala & 5.25 & \(\alpha\)-Glucose \\
2.30 & GABA & 6.91 & Tyr \\
2.35 & Glu & 7.34 & Phe \\
2.46 & Gln & 7.74 & Trp \\
2.55 & Citric Acid & 8.46 & Formic Acid \\
2.81 & Asp & 9.13 & Trigonelline \\
2.90 & Asn & & \\ \hline \end{tabular}
\end{table}
Table 1: Compounds and relative signals (ppm) selected for quantitative analysis in the hydroalcoholic extracts.