## 3 Results and Discussion
### Multivariate Lipid Signature
Metabolite data collected in the LC-MS positive ((a), n = 2 components, R\({}^{2}\)Y = 0.99, Q\({}^{2}\)Y = 0.64) and negative ((b), n = 2 components, R\({}^{2}\)Y = 0.61, Q\({}^{2}\)Y = 0.20) ionization modes. The analysis of complex lipids in the positive mode (Table 2) allowed us to annotate, among discriminant compounds, cholesteryl esters, sphingomyelins and their metabolites ceramides phosphate, phosphatidylcholines, phosphatidylethanolamineamines, diacylglycerols, and monoacylglycerols. Several triacylglycers and sterols were found to be discriminant, although they were differently expressed (upregulated or downregulated). \begin{table}
\begin{tabular}{c c c} \hline \hline
**Chemical Class** & **Metabolite** & **Regulation b ** \\ \hline \hline \multirow{4}{*}{Sterols} & Caffeine & Up \\  & Theobromine/Theophylline c  & Up \\  & 6-Amino5-(formyl-N-methylamino)-1,3-dimethyl uracil (1,3-7-DAU) & Up \\  & & demethylylacet1 (1,3-7-DAU) & Up \\  & & Cer(d14:1/28:0) & Up \\  & & GlcCer(d18:1/24:0) & Down \\  & & GlcCer(d15:1/18:0) & Down \\  & & Acetoacetic acid & Down \\  & & Propionic Acid & Up \\  & & Palmitic Acid & Down \\  & & Oleic Acid & Down \\  & & Myristic Acid & Down \\  & & 2-Hydroxyadipic acid & Down \\  & & Stearic Acid & Down \\  & & & \\ Lysophosphatidylethanolamineamines & Lysyler (18:1/0:0) & Up \\  & & ST 22:4:0\({}_{4}\) & Down \\  & & ST 29:4:0\({}_{4}\) & Down \\  & & ST 25:3\({}_{0}\) & Up \\  & & ST 23:4:0\({}_{2}\) & Down \\  & & ST 23:5\({}_{0}\) & Down \\  & & ST 27:4:0\({}_{4}\) & Down \\  & & ST 27:2\({}_{0}\) & Up \\  & & ST 23:5\({}_{0}\) & Up \\  & & ST 23:4\({}_{0}\) & Down \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Main discriminant metabolites a in the milk of goats fed SCG vs. control, detected in the negative ionization mode. \begin{table}
\begin{tabular}{c c c} \hline \hline
**Chemical Class** & **Metabolite** & **Regulation b ** \\ \hline \hline \multirow{8}{*}{Caffeine and caffeine metabolites} & Caffeine & Up \\  & Theobromine/Theophylline c  & Up \\  & 6-Amino5-(formyl-N-methylamino)-1,3-dimethylthiazol (1,3-7-DAU) & Up \\  & & dimethyl uracil (1,3-7-DAU) & Up \\  & Cer(d18:0/2:4:1) & Up \\  & Cer(d14:1/28:0) & Up \\  & Glc(d18:1/24:0) & Down \\  & & GlcCer(d15:1/18:0) & Down \\  & & Acetoacetic acid & Down \\  & & Propionic Acid & Up \\  & & Palmitic Acid & Down \\  & & Oleic Acid & Down \\  & & Myristic Acid & Down \\  & & 2-Hydroxyadipic acid & Down \\  & & Stearic Acid & Down \\  & & Stearic Acid & Down \\ Lysophosphatidylethanolamineamines & Lysyler (18:1/0:0) & Up \\  & & ST 22:4:0\({}_{4}\) & Down \\  & & ST 29:4:0\({}_{4}\) & Down \\  & & ST 25:3\({}_{0}\) & Up \\  & & ST 23:4:0\({}_{2}\) & Down \\ Sterols & ST 23:5\({}_{0}\) & Down \\  & & ST 27:4:0\({}_{4}\) & Down \\  & & ST 27:2\({}_{0}\) & Up \\  & & ST 23:5\({}_{0}\) & Up \\  & & ST 23:4\({}_{0}\) & Down \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Main discriminant metabolites a in the milk of goats fed SCG vs. control, detected in the negative ionization mode. Cholesteryl esters (CE) were found upregulated in milk samples of goats on the SCG diet. This result agrees with a recent study on human plasma [28] that evidenced an increase in CE in individuals consuming coffee. CE plays a crucial role in lipid metabolism as a transport form for essential fatty acids. Interestingly, the following CE of high-polyunsaturated fatty acids, i.e., CE(20:4), CE(22:4), and CE(22:5), were found upregulated; these fatty acids are present in the milk of ruminants in very low concentrations (\(<\)1% of fatty acids) [6]. In milk, the glycerophospholipids (PL) are represented by four main classes: phosphatidylethanolamine (PE, 26.4-72.3% of total PL), phosphatidylcholine (PC, 8.0-45.5%), phosphatidylserine (PS, 2.0-16.1%), and phosphatidylinositol (PI, 1.4-14.1%) [29]. Supplementing the diet with SCG increased the milk contents of PC and PE, indicating a significant effect on phospholipid synthesis in the mammary gland.