## 3 Results and Discussion
### \(\beta\)-Galactosidase Recovery and Concentration Using Ultrafiltration
The evaluation of all process factors--including the flow and its variations throughout the process, the enzymatic recovery and purification values, as well as the concentration and retention parameters for the protein and enzymatic activity--is necessary to define the conditions for carrying out ultrafiltration [1; 21]. Based on the previous results, the effect of the operating pressure was assessed using the 50 kDa membrane at pH 7.5, which presented higher purification factor and enzyme recovery values and a slight decrease in the permeate flux during \(\beta\)-galactosidase ultrafiltration. Figure 3 presents these results. After comparing the initial flux with the final flux in the process at different pressures (Figure 3), decreases of approximately 3.5%, 38.4%, and 47.6% were observed, respectively, indicating that the increase in pressure causes a significant decrease in permeate flux during the process. In the runs operated at low pressures, the concentration was gradual, leading to a slight drop in the permeate flux, a consequent reduction in the operating time, and an increase in the process yield (734.1 U/h) when compared to those at higher pressures (484.0 and 453.7 U/h for pressures of 2.0 and 2.5 kgf/cm\({}^{2}\), respectively). Additionally, a comparison between pressures of 2.0 and 2.5 kgf/cm\({}^{2}\) also showed the effect of higher pressures on the membrane concentration process, where an inversion of permeation efficiency was observed over time, especially at 25 min. Despite initially presenting a higher flux, the pressure of 2.5 kgf/cm\({}^{2}\) resulted in the faster concentration of the proteins (see the description of the effects below), altering the permeation capacity of the membrane,which made the process less efficient over time when compared to the process using a pressure of 2.0 kgf/cm\({}^{2}\), for example. After 35 min, no difference was observed between the pressures tested, and from this point onwards the processes seem equivalent. However, it is important to consider the advantages that reduce the operating costs and make the process viable, such as the constant filtration rate and low flow variation, the permeation of smaller particles present in the material, the membrane deformation, as well as consumption of gas used in the membrane permeation process [12; 52; 53]. The reduced recovery and purification factors of the enzyme in the systems operating with higher pressures may be associated with greater flux reductions caused by the membrane deformation (related to the compaction that occurs depending on the time of use, the constant conditions of high pressure, and the material used, which influences the permeability and increases the filtration resistance), concentration polarization, and fouling due to the applied pressure, leading to the inefficiency of the process; however, the occurrence of these phenomena is inevitable, can harm the performance of the membrane, and influence the process viability [54; 55]. The decrease in resistance with increasing pressure is generally not observed in permeation experiments. In general, the opposite contradictory trend is observed; that is, the higher pressure results in the increased formation of the second layer on the membrane, thereby reducing the permeability [56; 57]. Although working at reduced pressures with lower flux may seem inevitable, the results indicate that under less polarized conditions, the fouling is lower and the permeate flux is less affected during the process. Sodium and potassium chloride were added to the enzymatic extract of \(\beta\)-galactosidase, as these salts can change the ionic strength of the solution [58], act as protective agents of the enzyme [59], and modify the permeability during the separation of some compounds [58; 60; 61]. However, the addition of NaCl and KCl in the \(\beta\)-galactosidase extract revealed no positive effect compared to the control test (without added salt) (Table 2) and reduced the activity, purity, concentration, and enzyme recovery.