## 3 Results and Discussion
### Performance Testing of Synthesized Membranes
It is well known that almost all bacteria, Gram-positive and Gram-negative alike, carry a net negative charge [55]. Gram-positive bacteria possess negatively charged teichoic acids (due to the abundance of phosphate groups) which are intimately associated with the peptidoglycan cell wall or, in some cases, connected to the inferior cell surface membrane [15]. The overall negative charge conferred by Gram-negative bacterial species can be attributed to their negatively charged outer covering composed mainly of phospholipids and lipopolysaccharidesaccharides [56]. It is reasonable to assume that neat PES membranes, being inherently hydrophobic, turn water molecules away from their surface due to the absence of hydrogen bonds at the boundary layer [57]. The vacant surface allows for the settlement of bacteria and other fouling agents. Conversely, hydrophilic membrane surfaces such as PES/GO surfaces attract water molecules, rapidly forming a hydration layer which prevents the subsequent colonization of bacteria at its surface [58]. The addition of AG to PES membranes might also contribute to the enhancement of the membrane's antibacterial properties [46]. The exact mechanisms behind AG's antibacterial properties are as of yet unknown. Whilst some studies suggest the presence of antimicrobial enzymes such as  
Figure 11: Rejection and fouling data for PES/GO membranes. (**a**) BSA rejection values, (**b**) FRR (flux recovery ratio) % for different membranes and (**c**) the different types of fouling from the BSA solution filtration tests. oxidases, peroxididases and pectinases [59], other studies have hypothesized that the high salt content, particularly the divalent cations found in AG, was responsible for the antibacterial action [60]. Next, the antibiofouling properties of the fabricated membranes alongside the two commercial membranes were tested. As described in the methods section, the membranes were subjected to _E. coli_ suspension being pipetted on the active surface of the membranes and left incubated overnight at 37 \({}^{\circ}\)C. Figure 13a displays the DW flux before incubation (first 20 min) and the DW flux for two filtration cycles of 20 min after the 24 h incubation with bacterial suspension. While all membranes exhibit a drop-in flux after bacterial incubation, M8 samples showed the highest flux recovery as seen in Figure 13b. The normalized flux for M8 not only outperformed the neat PES membrane, however also the commercial Nadir and TriSep membranes (2.5 fold and 3-fold increase, respectively). When comparing the flux performance during filtration of _E. coli_ bacterial suspensions over extended time periods, M8 membrane showed only a small decline in flux over the entire duration of the filtration test (see Figure 14a). Conversely, both M1 and the commercial Tri  
Figure 12: FESEM images of PES/GO membrane surfaces after incubation with _E. coli_ suspension (top panel) and incubation with _B. subtilis_ suspension (bottom panel). Figure 13: Change in flux over time before and after overnight incubation (denoted as the red arrow) of different PES/GO membranes with _E. coli_ suspension (**a**) and normalized flux data for each membrane (**b**). a decline in flux, especially in the latter half of the filtration period. The difference is highlighted in the normalized flux graph that is seen in Figure 14b where M8 samples had a normalized flux that is more than double that of M1 and with a 31% increase compared to its commercial counterpart. In a parallel experiment with the same set up where the bacterial suspension was replaced with real locally obtained TSE, similar results were obtained where M8 samples fared better than the other two membranes (see Figure S4). Interestingly, the normalized flux for M8 was over two folds greater than the commercial membrane's normalized flux (see Figure S4). The antibiofouling properties of the PES membranes were further tested alongside the UP 150 (Nadir) membrane after incubation in _E. coli_ suspension at different incubation times (see Figure 15).