# The Effect of Type and Duration of Digestive Enzyme Treatment on Coffee Bean Composition
## 3 Results and Discussion  
### pH Values during Enzymatic Modification  
At the beginning and after a specified modification time, the pH values of samples and controls were measured (Table 1). The values measured immediately after preparation of the mixtures (0 h) were comparable for beans of different species and ranged between 4.13-4.92 and 6.95-7.01 for samples with the addition of 0.01 M HCl and phosphate buffer, respectively. In the case of the former ones, an increase in pH values over time was observed, with generally higher values observed with longer times. In the case of samples prepared using phosphate buffer, the opposite trend was noted--a slight (0.2-0.5) decrease in pH values after modification was observed. In controls, a slightly decreasing trend was noticeable with prolonged duration of the process.  
The applied HCl solution did not lower the pH of the coffee bean mixtures to a value close to 2, where pepsin exhibits maximum activity. However, literature data indicate that  
\begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**Modification**} & \multicolumn{2}{c}{**Pepsin**} & \multicolumn{2}{c}{**Trypsin and \(\alpha\)-Amylase**} \\ \cline{2-5}  & **Control** & **Modified** & **Control** & **Modified** \\ \hline \hline \multicolumn{5}{c}{Arabica beans} \\ \hline
0 & 4.92 \(\pm\) 0.17 \({}^{\text{a}}\) & 4.35 \(\pm\) 0.25 \({}^{\text{a}}\) & 6.95 \(\pm\) 0.01 \({}^{\text{a}}\) & 6.97 \(\pm\) 0.04 \({}^{\text{a}}\) \\
3 & 5.64 \(\pm\) 0.04 \({}^{\text{b}}\) & 5.00 \(\pm\) 0.01 \({}^{\text{b}}\) & 6.76 \(\pm\) 0.01 \({}^{\text{b}}\) & 6.73 \(\pm\) 0.03 \({}^{\text{b}}\) \\
6 & 5.66 \(\pm\) 0.02 \({}^{\text{b}}\) & 5.54 \(\pm\) 0.09 \({}^{\text{b}}\) & 6.71 \(\pm\) 0.02 \({}^{\text{b}}\) & 6.81 \(\pm\) 0.02 \({}^{\text{b}}\) \\
12 & 5.78 \(\pm\) 0.03 \({}^{\text{b}}\) & 5.61 \(\pm\) 0.02 \({}^{\text{b}}\) & 6.57 \(\pm\) 0.02 \({}^{\text{b}}\) & 6.64 \(\pm\) 0.02 \({}^{\text{c}}\) \\ \hline \multicolumn{5}{c}{Robusta beans} \\ \hline
0 & 4.66 \(\pm\) 0.11 \({}^{\text{a}}\) & 4.29 \(\pm\) 0.13 \({}^{\text{a}}\) & 6.99 \(\pm\) 0.05 \({}^{\text{a}}\) & 6.98 \(\pm\) 0.06 \({}^{\text{a}}\) \\
3 & 5.59 \(\pm\) 0.08 \({}^{\text{b}}\) & 5.2 \(\pm\) 0.03 \({}^{\text{b}}\) & 6.89 \(\pm\) 0.06 \({}^{\text{b}}\) & 6.75 \(\pm\) 0.02 \({}^{\text{c}}\) \\
6 & 5.64 \(\pm\) 0.09 \({}^{\text{b}}\) & 5.28 \(\pm\) 0.1 \({}^{\text{b}}\) & 6.83 \(\pm\) 0.08 \({}^{\text{b}}\) & 6.88 \(\pm\) 0.04 \({}^{\text{b}}\) \\
12 & 5.64 \(\pm\) 0.04 \({}^{\text{b}}\) & 5.54 \(\pm\) 0.02 \({}^{\text{b}}\) & 6.74 \(\pm\) 0.02 \({}^{\text{c}}\) & 6.72 \(\pm\) 0.02 \({}^{\text{c}}\) \\ \hline \hline \end{tabular} All results expressed as mean value \(\pm\) standard deviation. In columns, different letters indicate statistically significant differences between results.  
\end{table}
Table 1: pH values of samples during enzymatic modification.  
pepsin shows activity at pH values below 6.5, which is also confirmed by the data from the manufacturer of the enzyme used [20]. Overall, conditions created by usage of applied solutions (HCl and phosphate buffer) were suitable for observing the activity of selected digestive enzymes.  
Literature data show that the pH value of green coffee bean extracts can be in the range of 4-5; these values vary depending on the geographical origin and processing method [21; 22]. In this work, observed changes in pH values could be explained by the extraction of coffee compounds. During the sterilization process, high temperature might have led to an increase in beans' volume and loosening of their structure, significantly affecting the extraction rate. Additionally, the applied digestive enzymes could have caused the release of additional buffering compounds (including proteins and peptides) from the coffee matrix, which could have influenced the pH of the environment.