## 2 Results and Discussion
### Polyphenolic Compounds Analysis by HPLC Supported by FTIR and Confocal Microscopy
#### 2.1.2 FTIR Analysis  
To compare the differences and similarities of polyphenol peaks, which correspond to stretching vibrations of some functional groups, the FTIR spectrum of three investigated wines, ferulic acid, and rutin between the region of 4000 and 500 cm\({}^{-1}\) were combined in Figure 1. A FTIR spectral analysis of wine extracts reported similar absorption bands as obtained for the standards ferulic acid and rutin. Persimmon (3276 cm\({}^{-1}\)), pomergnate (3333 cm\({}^{-1}\)), and kiwifruit (3286 cm\({}^{-1}\)) wines (Figure 1A-C) showed one prominent characteristic peak, similar to the peaks observed in ferulic acid (3433 cm\({}^{-1}\)) and in rutin (3422 cm\({}^{-1}\)). Therefore, spectral bands in the range 3500-3300 cm\({}^{-1}\) can be attributed to the stretching vibrations of the O-H groups, characteristic of polyphenolic extracts. \begin{table}
\begin{tabular}{c c c c} \hline \hline
**Phenolic Acids** & **Pomergnate Wine** & **Kiwi Wine** & **Persimmon Wine** \\ \hline chlorogenic acid & 3.16 \(\pm\) 0.30 \({}^{\text{ab}}\) & 0.49 \(\pm\) 0.10 \({}^{\text{ac}}\) & 0.10 \(\pm\) 0.04 \({}^{\text{bc}}\) \\ gallic acid & 7.99 \(\pm\) 0.23 \({}^{\text{ab}}\) & 1.78 \(\pm\) 0.10 \({}^{\text{ac}}\) & 0.44 \(\pm\) 0.11 \({}^{\text{bc}}\) \\ caffeic acid & 1.09 \(\pm\) 0.25 \({}^{\text{ab}}\) & 0.10 \(\pm\) 0.01 \({}^{\text{ac}}\) & 0.06 \(\pm\) 0.00 \({}^{\text{bc}}\) \\ \(p\)-coumaric acid & 0.09 \(\pm\) 0.02 & \textless{LOD} & \textless{LOD} \\ protocatechuic acid & 0.22 \(\pm\) 0.03 \({}^{\text{ab}}\) & 0.76 \(\pm\) 0.10 \({}^{\text{ac}}\) & 0.05 \(\pm\) 0.02 \({}^{\text{bc}}\) \\ sum of phenolic acids & 12.5 & 3.13 & 0.65 \\ \hline \hline \end{tabular} The same letters in superscript indicate significant differences in the results within the row; levels of significance \(p<\) 0.05.  
\end{table} 
Table 1: Concentration of phenolic acids in freeze-dried fruit wines [mg/g]. n = 3. In this spectral range, polyphenolic extracts had vibration bands similar to acids, as in ferfulic acid (Figure 1D), however, a number of vibrational contributions of the -OH groups appeared [17]. Peaks (cm\({}^{-1}\)) at 2969, 2943, and 2930 appeared in ferulic acid, pomergnate, and kiwifruit wines. Rutin and persimmon wine showed similar peaks at 2936 cm\({}^{-1}\). The spectral bands located between 2969 and 2930 cm\({}^{-1}\) could be attributed to C-H stretching vibrations, being due to the stretching vibrations of the O-H groups [18]. Strong absorption bands in the region of 1720-900 cm\({}^{-1}\) corresponded to the fingerprint region (Figure 1A-C). According to [PERSON] [19], FTIR spectra in the fingerprint region between 1750 and 950 cm\({}^{-1}\) are characteristic of polyphenolic extracts in wines. A slight difference (1600-1000 cm\({}^{-1}\)) may be caused by the presence of protein and the gallic acid side chain in the investigated samples [18; 20]. One peak (cm\({}^{-1}\)) nearly at the same place (1721, 1723, and 1727) appeared only in the examined wines and was also found in other reports [19], suggesting the presence of the carbonyl C=O stretching band of protonated carboxylic acid, characteristic of the galloyl unit of hydrolyzable tannins. The bands (cm\({}^{-1}\)) observed a 1622, 1632, 1687, and 1650 for pomergnate and kiwifruit wines, and ferulic acid and rutin, respectively (Figure 1B-E), can be associated with the aromatic C=C stretching vibrations, present in the condensed tannins, as well as C=O stretching vibrations and the presence of unsaturated bonds in flavonoid structures. Peaks between 1465 and 1410, and 1403 and 1442 allocated the carboxylic acid (O-C-O) were observed in ferulic acid, pomergnate, and kiwifruit wines, and were not found in persimmon wine samples. The peak at 1444 cm\({}^{-1}\) exhibited the C=C-C of aromatic rings and was found only in kiwifruit  
Figure 1: FTIR spectra in the range 4000–500 cm\({}^{-1}\) of polyphenolic extracts: (**A**) persimmon, (**B**) kiwifruit, (**C**) pomergnate wines, (**D**) ferulic acid, and (**E**) rutin.