## 3 Results and Discussion
### Yield and Synthesis Efficiency of BC

Table 1: The influence of culture conditions on analysed enzymes stability. It may be a consequence of the specific properties of \(\beta\)-Gal and the ability to conduct transgalactosylation reaction, which can reduce the amount of available glucose in the reaction medium during the cultivation period of _K. xylinus_[38]. The lowest yield of BC production was found in the cultures of _K. xylinus_ grown in a medium containing starch. The cultures cultivated with the addition of 5.0, 10, and 20 U/mL glucoamylase units obtained approximately 3.06, 3.23, and 3.11 g/L of BC, respectively. Cultures containing a higher amount of added enzyme, 40 U/mL, resulted in slightly lower amounts of CB, 2.41 g/L. In the control culture without enzyme, insignificant amounts of CB not exceeding 0.32 g/L were obtained (Figure 3c). Moreover, the macrostructure of obtained pellicles was highly disordered. The efficiency of BC synthesis in the cultures conducted in HS medium containing starch and glucoamylase was at a level not exceeding 31% for added enzyme amounts of 5.0, 10, and 20 U/mL. Significant lower efficiency was also observed for 40 U/mL, reaching only 24% (Figure 3f). The reason for this difference could be the faster decrease in the pH of the medium as a result of the faster release of glucose from starch, further oxidised to gluconic acid by _K. xylinus_ cells. [33]. The high amount of GLA units allowed a short time to saturate the culture medium with glucose to the concentration, which caused its excessive oxidation to gluconic acid, which was manifested by the highest acidification of the medium and the limitation of the BC synthesis. Attenuated Total Reflectance in Fourier Transformation InfraRed Spectroscopy (ATR-FTIR) and X-ray Diffraction (XRD) Analysis of Bacterial Cellulose  
The ATR-FTIR and XRD were used for assessment of the influence of enzymes-assisted cultivation on the molecular BC properties. The ATR-FTIR spectra of BC obtained from _K. xylinus_ cultures maintained in HS medium with various substrates and glycolhydrolases are shown in Figure 4. In the range 800 cm\({}^{-1}\) to 1800 cm\({}^{-1}\), all spectra contained a typical for BC set of bands with intense signals from adsorbed water at 1641 cm\({}^{-1}\), \(\delta\)CH2 (symmetric) of C-6 at 1427 cm\({}^{-1}\), \(\delta\)COH in the plane from C-2 and C-3 at 1134 cm\({}^{-1}\), \(\delta\)OC of \(\beta\)-glycosidic linkage at 1160 cm\({}^{-1}\), \(\delta\)CO at C-6 at 1030 cm\({}^{-1}\), and \(\delta\)COC of \(\beta\)-glycosidic bonds of amorphous BC phase at 897 cm\({}^{-1}\). In the second part of spectra at range from 2600 cm\({}^{-1}\) to 3600 cm\({}^{-1}\) were seen the bands specific for \(\delta\)OH for hydrogen and covalent bonds at 3334 cm\({}^{-1}\) and \(\delta\)CH at 2895 cm\({}^{-1}\)[24]. The ATR-FTIR spectra analysis showed no presence of significant changes in bands intensivity that could point at the modification of the BC molecular structure by the action of the analysed enzymes and used as carbon sources, oligo- and polysaccharides. The impact on the BC synthesis process at the cellular level or alteration of BC microfibrils structure by constituents added to the cultivation media can be demonstrated by alteration of BC crystallinity and porosity [39]. Nevertheless, the IR crystallinity index's values (LOI, TCI) did not differ considerably between the BC samples obtained from cultivation media with different enzymes and substrates (Table 2). The XRD analysis was consistent with ATR-FTIR and showed that the addition of enzymes into cultivation media has no influence on BC microfibrils' molecular structure (Figure 4b). The crystallite sizes of BC from control cultures and with the addition of enzymes ranged from 5.4 nm to 5.9 nm, consistent with earlier reported values [36]. Moreover, no significant differences in the crystallinity (CrI) of BC microfibrils were detected, which indicates the neutrality of the used glycolhydrolases for the BC synthesis process.