## 3 Results and Discussion
### Birefringence of Cassava Starches in HPP-treated Cassava Flours
#### 3.3.1 Short-Range Order of Cassava Starches  
Short-range order is defined as the double-helical order and is related to variations in the ratio of the amounts of ordered to an unordered (amorphous) fraction within the starch granules [42]. The FTIR spectrum band at 1300-800 cm\({}^{-1}\) is sensitive to starch granule's C-O-H bending [43]. Most previous works have identified bands at 1047, 1022, and 995 cm\({}^{-1}\) to be sensitive to changes in the short-range order, representing the amount of ordered or crystalline starch, the amount of amorphous starch, and the influence of water content on the intramolecular hydrogen bonding, respectively [23]. The absorbance ratio of 1047/1022 cm\({}^{-1}\) was used to quantify the ordered starch to amorphous starch [21], with a larger value being more crystalline [22], whereas 1022/995 cm\({}^{-1}\) measures the molecular order of starch double helices, with an increased organization at a lower ratio [21; 24]. No interactions were detected between the processing variables for 1047/1022 cm\({}^{-1}\), but a significant pressure effect was found (Table 1). Table 3 shows a notable decrease in the ratio, especially at 600-30 min. The FTIR spectra shown in Figure 3 for 875-1200 cm\({}^{-1}\) also \begin{table}
\begin{tabular}{c c c c c c c} \hline
**Source of Variation** & **Abs** & **Abs** & **\%RC** & **\%SDS** & **\%SDS** & **\%RS** \\  & **1047/1022** & **1022/995** & & & & & \\ \hline Pressure & 0.002 & \textless{}0.001 & \textless{}0.001 & \textless{}0.001 & \textless{}0.001 & \textless{}0.001 \\ FC & 0.061 & 0.601 & \textless{}0.001 & 0.011 & 0.010 & \textless{}0.001 \\ Time & 0.053 & 0.307 & 0.014 & 0.062 & 0.010 & 0.005 \\ Pressure \(*\) FC & 0.191 & 0.939 & \textless{}0.001 & 0.147 & 0.020 & 0.151 \\ Pressure \(*\) HT & 0.237 & 0.045 & 0.008 & 0.002 & 0.456 & 0.003 \\ FC \(*\) HT & 0.139 & 0.450 & 0.929 & 0.368 & 0.601 & 0.351 \\ Pressure \(*\) FC \(*\) HT & 0.418 & 0.909 & 0.027 & 0.091 & 0.462 & 0.494 \\ \hline \end{tabular}
\end{table} 
Table 1: Summary of probability values after three-way ANOVA analysis of absorbance ratios, %RC, and starch digestion fractions as affected by pressure, flour concentration (FC), and holding time (HT). \begin{table}
\begin{tabular}{c c c c c c} \hline
**Source of Variation** & _To_ & _Tp_ & _Tc_ & _Trange_ & \(\Delta H\) & _DG_ \\ \hline Pressure & \textless{}0.001 & \textless{}0.001 & \textless{}0.001 & 0.039 & \textless{}0.001 & \textless{}0.001 \\ FC & 0.018 & 0.323 & 0.797 & 0.216 & \textless{}0.001 & \textless{}0.001 \\ Time & \textless{}0.001 & 0.110 & 0.196 & \textless{}0.001 & \textless{}0.001 & \textless{}0.001 \\ Pressure \(*\) FC & \textless{}0.001 & 0.015 & 0.590 & 0.194 & \textless{}0.001 & \textless{}0.001 \\ Pressure \(*\) HT & \textless{}0.001 & 0.024 & 0.960 & \textless{}0.001 & \textless{}0.001 & \textless{}0.001 \\ FC \(*\) HT & 0.104 & 0.874 & 0.466 & 0.111 & 0.502 & 0.377 \\ Pressure \(*\) FC \(*\) HT & 0.039 & 0.902 & 0.697 & 0.191 & \textless{}0.001 & \textless{}0.001 \\ \hline \end{tabular}
\end{table} 
Table 2: Summary of probability values after three-way ANOVA analysis of thermal properties of cassava flour as affected by pressure, flour concentration (FC), and holding time (HT). displayed a decreased absorbance at pressure \(\geq\)500 MPa, similar to HPP-treated quinoa flour [11]. A notable decreasing band for 1047 cm\({}^{-1}\) and an increasing 1022 cm\({}^{-1}\) band were also observed, especially for 600-10% treatments. These imply a reduction in the short-range order and an increase in the amorphous starch fraction during gelatinization [23; 42]. On the other hand, the absorbance ratio for 1022/995 cm\({}^{-1}\) had a significant two-way interaction between pressure and time, with both having a significant simple main effect (Table 1). [PERSON], [PERSON], and [PERSON] [43] also found this ratio to be a valuable tool to monitor loss of structure, but a thorough assessment of spectra suggested greater complexity.