## 3 Results  
### FTIR Spectra of Citin and Degree of Acetylation  
In a previous study [9], we analyzed the different steps of the extraction process (i.e., demineralization, deproteinization and decolorization) from cicada sloughs collected in 2017. The FTIR characterization showed that the raw material and the demineralized sample presented a single peak at approximately 1660 cm\({}^{-1}\), while after the deproteinization two individual peaks appeared at 1620 cm\({}^{-1}\) and 1660 cm\({}^{-1}\). This phenomenon was attributed to the removal of proteins since the peak at 1660 cm\({}^{-1}\) might be assigned to the amide I vibrational modes [12]. Furthermore, the presence of two individual peaks also suggested that the extracted chitin was the \(\alpha\)-polymorph, while the presence of a single peak is attributed to the \(\beta\)-polymorph [13]. In order to validate these observations, we collected the FTIR spectra of the raw material and of the intermediary products of the extraction of chitin from cicada sloughs harvested in 2019 and 2020--see Figure 11. We found that the FTIR spectra for the extractions performed on cicada sloughs harvested \begin{table}
\begin{tabular}{c c} \hline \hline
**Abbreviations** & **Corresponding Sample** \\ \hline CH-17 & chitin extracted from cicada sloughs harvested in 2017 \\ \hline CH-19 & Chitin extracted from cicada sloughs harvested in 2019 \\ \hline CH-20 & Chitin extracted from cicada sloughs harvested in 2020 \\ \hline CS-80 & Commercial chitosan with a _DD_ of \(\approx\)80 \% \\ \hline CS-90 & Commercial chitosan with a _DD_ of \(\approx\)90 \% \\ \hline CS-95 & Commercial chitosan with a _DD_ of \(\approx\)95 \% \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Abbreviations of the samples used through the text. in 2019 and 2020 are very similar to each other. In both the extraction processes, the raw material and the demineralized product present only one peak at 1660 cm\({}^{-1}\) while two peaks appear at 1620 cm\({}^{-1}\) and 1660 cm\({}^{-1}\) for the deproteinized and decolorized products, confirming that proteins were efficiently removed, and that chitin is in its \(\alpha\)-polymorph. According to [PERSON] et al. [3]\(\alpha\)-chitin has two types of hydrogen bonding: intra-sheet and inter-sheet. The absorption band at 1660 cm\({}^{-1}\) correspond to hydrogen bonding between the carbonyl groups (C=O) of amide I and amide II (NH) while the absorption band at 1620 cm\({}^{-1}\) correspond to the hydrogen bonding between the CH\({}_{2}\)OH side chain and the carbonyl group (C=O). Since the amide I of proteins absorbs at wavelengths comprised between 1600 and 1700 cm\({}^{-1}\), the presence of proteins and the interactions of the latter with chitin, lead to the appearance of a single peak, at wavelengths comprised between 1620 and 1660 cm\({}^{-1}\), in the samples before deproteinization (i.e., raw material and demineralized sample). Here, in order to evaluate the quality of the final product (i.e., chitin extracted from cicada sloughs harvested in three different years), we compare the FTIR spectra (shown in Figure 1) of the samples CH-17, CH-19, and CH-20 among them and with a commercial chitin. The three extracted chitin present similar spectra and, in particular, very similar to the spectrum of the commercial chitin. The FTIR spectra were used for the measurements of the degree of acetylation (\(DA\)). In Table 2 are presented the average \(DA\) of CH-17, CH-19 and CH-20. The results show that chitin extracted from cicada sloughs collected in three different years had a different degree of acetylation. In particular, CH-17 and CH-19 were more acetylated than CH-20, suggesting that chitin extracted from cicada sloughs harvested in 2020 was different from the others. To better understand if the difference in the \(DA\) is due to the extraction process or to an intrinsic difference within the source, we compared the FTIR spectra of the raw materials (cicada sloughs) harvested in 2017, 2019 and 2020.