## 3 Results and Discussion
### Quantification of the Polyphenols with UPLC-Q-TOF-MS  
The phenolic profile of chestnut leaves was carried out by means of UPLC-Q-TOF-MS analyses. All of the compounds were identified by comparison of their retention times, elution orders, ESI-MS spectrometric data, and photodiode array PDA/UV-vis with authentic reference standards and/or the literature data. The chromatographic, spectroscopic and spectrometric data, as well as the quantitative amounts of individual compounds are shown in Table 2.  
In total, 18 phenolic compounds were putatively identified. The main class (six compounds) consisted of hydrolysate tannins. Hydrolysable tannins are polymers containing a center core of glucose esterified with gallic or ellagic acids (gallo- and ellagitannins, respectively). They exhibit anti-cancer, anti-angiogenic, antioxidant, anti-inflammatory and anti-ulcerative properties [34]. Compounds **2**, **4**, **5**, and **10** were characterized as ellagitannins, the isomers of galloyl-bis-HHDP-glucose. They all displayed a dominant precursor [M-H]- ion at _m/z_ 935 and ion at _m/z_ 467 corresponding to doubly charged molecular \begin{table}
\begin{tabular}{c c c} \hline \hline  & **TPC** & **TFC** \\  & **mg GAE/g** & **mg QE/g** \\ \hline _C. sativa_ leaf methanolic extract & 458.22 \(\pm\) 1.02 & 24.55 \(\pm\) 0.08 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Total phenolic (TPC) and flavonoid (TFC) content in _C. sativa_ leaf methanolic extract. ion [M-2H]\({}^{2-}\). Component **3** showed [M-H]- at _m/z_ 953 and the product ion at _m/z_ 785, which was received by the loss of 168 Da. It also produced a fragmentation ion at _m/z_ 301 coinciding to ellagic acid and at the same time being the most significant diagnostic ion of ellagitannins [35]. Therefore, this component was assigned to galloyl-chebuloyl-HHDP-glucose (chebulagic acid). In turn, Compound **9** was identified as galloyl glucose derivative--trigalloyl-glucose. It displayed parent ion [M-H]- at _m/z_ 635 with a typical for gallotannins fragmentation pathway with the loss of gallic acid moiety (170 Da). The second most abundant group of compounds were phenol glucose derivatives. Components **6** and **8** both exhibited pseudomolecular ions [M-H]- at _m/z_ 637 with fragmentation [M-H-170-162]- at _m/z_ 467 and 305 attributable to the loss of galloyl and glucose units, respectively. They were identified as chesnatin and isochsenatin. Peaks 13 and 16 both showed precursor ion [M-H]- at _m/z_ 937 that produced fragment ion [M-H-470]- at _m/z_ 467 due to the loss of the glucose, the 3,4,5-trihydroxybenzyl alcohol and the galloyl moieties; thus, they were tentatively characterized as chestanin and isochsenatin. Compound **11** showed [M-H]- at 469 and the daughter ion [M-H-300]- at _m/z_ 169, indicating the simultaneous loss of the 3,4,5-trihydroxybenzyl alcohol unit and the glucose molecule. Further, five constituents were identified as flavonoid glycosides, namely quercetin 3-\(O\)-glucuronide (**14**), quercetin 3-\(O\)-glucose (**15**), kaempferol 3-\(O\)-glucose (**17**) and isorhamethein 3-\(O\)-glucose (**18**). Their full mass spectra showed deprotonated molecules [M-H]- at _m/z_ 477, 463, 447 and 417, respectively. The fragmentation pathway displayed common diagnostic product ions at _m/z_ 301 (**14** and **15**), 285 (**17**) and 315 (**18**), confirming the presence of three different aglycones, corresponding to quercetin, kaempferol and isorhamethein, respectively, and, at the same time, the neutral loss of the sugar unit, which was represented by glucose (162 Da) and glucuronic moiety (176 Da). Additionally, Component 1 was found to be gallic acid, Compound **3** was methylgallate and Constituent 12 was ellagic acid pentoside. Gallic acid (**1**) fragmentation produced the [M-H]- ion at _m/z_ 169 and the MS/MS ion spectrum [M-H-44]- at _m/z_ 125, suggesting the loss of carbon dioxide. Methyl-gallate (**3**) was characterized by the diag