## 2 Advancements in Ultrafiltration for Protein and Peptide Separation
### Electroultrafiltration (EUF) and Electrodialysis with UF Membrane (EDUF)
Removal of ions is governed by  
\[c_{f}-c_{d}=I/(z\;F\;Q) \tag{8}\]  
The removal rate or concentration difference between diluate or product (\(c_{d}\)) and feed (\(c_{f}\)) is determined by applied current, feed flowrate (\(Q\)), ionic valence \(z\), and Faraday constant \(F\). Only ionic substances are transported in through ion-exchange membranes in ED, while solvent is almost unaffected by the electrical field. CEM is permeable for cations while excluding anions. In contrast, AEM allows the permeation of anions but rejects cations. The introduction of UF into the ED module allows one to separate ionic components that have different molecular weights. EDUF can separate salt and protein into different compartments [76], and may be employed within either separation or elution modes as depicted in Figure 2.  
Separation mode can be used to attain high productivity, although with the cost of not-too-selective isolation. Since multiple feeds are flowed through, this will divide the protein based on charge, each on concentrated and diluted side. In Figure 2a, negatively charged A is the primary target and positively charged B is the secondary target, making CEM embrace the concentrated (A-rich) stream and AEM embrace the diluted (B-rich) stream. The elution mode aims to achieve higher purification, but that requires the sacrifice of its productivity. So, as only one feed is passed through, the majority of targeted compounds would be collected in either the diluted or concentrated stream. In Figure 2b, owing to the targeted and negatively charged A, CEM interface will directly be accessible toward the concentrated (rich A) stream while AEM faces lower concentration of both A and B (compared to separation mode). In an experiment to isolate \(\alpha\)-lactalbumin and bovine hemoglobin, at initial concentration of 0.1 g L\({}^{-1}\), 99% of \(\alpha\)-lactalbumin purity was obtained at concentrate side by elution mode. Meanwhile, superior yield by separation mode exceeds the elution mode by 20% [78]. Promisingly, EDUF can enhance the bioactivity of isolate like portrayed in fractionating herring mild hydrolysate that attained 214%-increase in antioxidant activity [98], and 125 ug mL\({}^{-1}\) of 0.3-0.5 kDa cationic peptides exhibit in vitro noncompetitive inhibition toward ACE and dipeptidyl peptidase IV (DPP-IV) after single-step 4-h treatment [79]. Figure 2: Electrodialysis with UF membrane (EDUF) in (**a**) separation and (**b**) elution mode. CEM: cation-exchange membrane; AEM: anion-exchange membrane; UFM: UF membrane. In order to increase the purity and selectivity, in particular, permeate flux to compete that of UF with the same membrane and length of the process, configuration of EDUF can be arranged as disclosed in another review [32]. Hitherto, there are six stand-out configurations relevant in EDUF practices, which are described in Figure 3a-f, of which configuration (a) is the most popular. Furthermore, integration of EDUF with other UF process and physical pretreatment of feeds are sought to grow the acceptance of EDUF in industries. First, when treating soy protein isolate, operation of EDUF at pH 3.0 and 6.0 amplified the antioxidant capacity owing to higher concentration polar peptides with lysine moieties (0.4-0.5 kDa) from anionic compartment compared to the NF when tested in vitro against reactive oxygen species by human neuroblastoma cell line. Of isolates generated from NF and EDUF at pH 9.0, only the capability of degrading H\({}_{2}\)O\({}_{2}\) was observed. Thus, sequence integration of EDUF-NF would be advantageous to refine the UF permeate [99]. Second, pretreatment of defatted flaxseed protein isolate with high hydrostatic pressure (HHP) at 400 MPa HHP and 21 \({}^{\circ}\)C for 20 min, and further processing with EDUF resulted in isolate (recovered in KCl) rich in arginine, which lowers systolic blood pressure [100]. Arginine itself cannot be separated using conventional electrodialysis since it adulterated the CEM in basic EDUF [101].