Phosphonium Modi�ed Nanocellulose Membranes with High Permeate Flux and Antibacterial Property for Oily Wastewater Separation

Nanocellulose membranes could e�ciently separate oily wastewater because of their superhydrophilic and underwater superoleophobic property and nanoporous structure. However, the practical application and storage of nanocellulose membranes were limited by their low water permeation �ux and easy corrosion by bacteria, respectively. Herein, nanocellulose membranes with high permeate �ux and antibacterial property were fabricated by grafting tetrakis(hydroxymethyl) phosphonium chloride (THPC) onto the surface of tunicate cellulose nano�bers (TCNFs) via esteri�cation reaction. The introduction of THPC groups with tetrahedral structure on the surface of TCNFs signi�cantly improved the pore size and interlayer space of nanocellulose membranes, resulting in the increasing of water permeation �ux. These THPC@TCNF membranes were superhydrophilic and underwater superoleophobic, which could effectively separate various oil/water nanoemulsion. Moreover, THPC@TCNF membranes possessed excellent durability, mechanical stability, and cycling performance. Due to the presence of positively charged phosphonium groups, THPC@TCNF membranes exhibited excellent antibacterial property against B. subtilis that is a typical Gram-positive bacterium presenting in oily wastewater. This works provided a facile method to endow nanocellulose membrane with high permeate �ux and antibacterial property.


Introduction
Oily wastewater produced from petrochemical industry and human daily life have attracted worldwide environmental concerns, therefore, high-performance materials for oil/water separation are imminently required (Gupta et  effectively separate oil/water nanoemulsions, which was attributed to the nano-scale pore size and super wettability of the membrane (Cheng et al. 2017). However, the nanoporous structure of membranes produced by dense assembly of nanocellulose always reduced their permeate ux, which severely limited their separation e ciency (Kwon et al. 2021). According to the Hagen-Poiseuille Eq. (1): Herein, we tried to fabricate antibacterial nanocellulose membranes with high permeate ux by grafting tetrakis(hydroxymethyl) phosphonium chloride (THPC) onto tunicate cellulose nano bers (TCNFs). THPC with ame retardancy and antibacterial property is an inexpensive by-product of phosphine tail gas (Olkiewicz et al. 2015). Owing to its tetrahedral structure and broad-spectrum antibacterial characteristics (Palermo et al. 2019;Peng et al. 2020), THPC was employed as pore-forming and antibacterial agent to endow membranes with enlarged pore size and antibacterial property. After esteri cation reaction, the average pore size and thickness of THPC@TCNF membranes increased to 62.2 ± 2.9 nm and 21.1 µm, respectively. The THPC@TCNF membrane could separate oil/water nanoemulsion with water permeate ux of 1858 L m − 2 h − 1 bar − 1 and oil rejection of 99%. Furthermore, the resultant membrane exhibited good mechanical stability, cycle performance, and antibacterial property. diameter: 50 mm) as a support was bought from Tianjin Jinteng Co., Ltd (Tianjin, China). 2,2,6,6tetramethyl-1-piperidinyloxy (TEMPO) was purchased from Sigma-Aldrich (America).

Fabrication of tunicate cellulose nano bers (TCNFs)
Tunicate cellulose was isolated from the mantle of tunicate by NaOH boiling and H 2 O 2 bleaching according to our previous method (Huang et al. 2019). Afterwards, tunicate cellulose (1 g) was dispersed in deionized water (99 g), followed by the addition of NaBr (0.1 g) and TEMPO (0.016 g). After TEMPO was completely dissolved, NaClO (6.2 g, 12%) was used to start the TEMPO oxidation, and NaOH (1 M) was applied to maintain the reaction medium pH of 10 ~ 10.5 under 400 rpm stirring for 12 h. Finally, the TEMPO-oxidized tunicate cellulose suspension was centrifuged and dialyzed to neutrality, and the TCNFs suspension was obtained by mechanical stirring in an ordinary soymilk machine.

Fabrication of THPC@TCNF membranes
TCNF suspension (12 mL, 0.06 wt%) was freely deposited on the Nylon 66 supported membrane by vacuum-assisted ltration to obtain a TCNF layer (5.73 g m − 2 ). Subsequently, the Nylon 66 membrane coated with TCNF layer was dried, and nally stripped from the Nylon 66 support to obtain selfsupporting TCNF membrane. Subsequently, a certain amount of THPC (2 ~ 32 wt%) with DMAP (5 wt%) were dissolved in DMSO. The TCNF membranes were then soaked with above solution at room temperature for 24 h. Finally, THPC@TCNF membranes were fabricated by washing with deionized water and drying.

Characterization
The morphologies of TCNFs and THPC@TCNFs were visualized with transmission electron microscopy (TEM, JEM-2100, Japan). The morphology of membranes was observed with scanning electron microscope (SEM, Zeiss SIGMA, Germany). The chemical structures of THPC@TCNF membranes were analyzed with Fourier transform infrared spectrometer (FTIR, NICOLET 5700, Thermo Fisher Scienti c, USA). The constituent elements of the membranes before and after modi cation were analyzed by X-ray photoelectron spectrometer (XPS, ESCALAB250Xi, Thermo Fisher Scienti c, USA). The distribution of P and Cl element was observed with energy disperse spectroscopy (EDS, Oxford UltimMax 40, Oxford, UK).
The wettability of membranes was characterized by contact angle measurements (DSA100, Krüss, Germany). The oil concentration of the ltrates was measured with infrared spectrometer oil content analyzer (FYHW-2000B, Fangyuan, China).

Oil/water nanoemulsion separation
Four types of emulsions containing oil droplets of 50 ~ 500 nm were prepared by dispersing petroleum ether, chloroform, pump oil, and dimethicone into water with the help of surfactants, respectively. Details on the preparation of various nanoemulsions were provided in the Supporting Information. In a typical oil/water nanoemulsion separation process, the effective lter membrane area was 1.54 cm 2 and various nanoemulsions were fed into the separation unit at a constant pressure of 0.5 bar. According to the volume of ltrate collected in 5 min, the water ux J (L m − 2 h − 1 bar − 1 ) was calculated by substituting into Eq. (2): , t (h) and ΔP (bar) was water permeate volume, effective ltration membrane area, permeation time, and transmembrane pressure, respectively. Besides, the oil rejection R (%) was obtained according to Eq. (3): where C f (mg mL − 1 ) and C 0 (mg mL − 1 ) were the oil concentrations in the ltrate collected after ltration and in the un ltered oil/water emulsion, respectively. The infrared spectrometer oil content analyzer was used to measure the value of C f .
For the cycle performance of THPC@TCNF membranes in the oil/water separation process, the membranes were soaked in pure water for a few seconds after each cycle and dried for the next cycle.

Antibacterial property
The antibacterial properties of various membranes were appraised using Bacillus subtilis (B. subtilis), which is a typical Gram-positive bacterium presenting in oily wastewater. In the blend culture test, a complete membrane was soaked in the diluted B. subtilis solution (10 7 CFU mL − 1 , 2 mL). After cultured at 37°C for 1 h, bacterial solution (100 µL) was spread on the Luria-Bertani (LB) culture medium. After culturing at 37°C for 24 h, the number of colonies was counted. For the bacterial ltration test, the diluted bacterial liquid (10 2 CFU mL − 1 , 10 mL) was deposited on the membranes by vacuum-assisted ltration. The membranes were kept under wet condition for 1 h, and then the morphology of bacterial was observed by SEM.

Fabrication and morphology of THPC@TCNF membranes
The fabrication process of THPC@TCNF membrane was displayed in Fig. 1. Firstly, The TEMPO oxidized TCNFs suspension was deposited on nylon 66 supported membrane by vacuum assisted ltration. After drying, the supported membrane was removed to obtain self-supporting TCNF membrane ( Figure S1a).
After TEMPO oxidation, a lot of carboxyl groups appeared on the surface of TCNFs. By immersing TCNF membrane into THPC solution, THPC was grafted onto the surface of TCNFs in the membrane via esteri cation reaction. Finally, THPC@TCNF membrane was obtained after washing with water and drying ( Figure S1b). Bene ting from the swelling effect of DMSO, the tetrahedral structure of THPC, and the electrostatic repulsion of positively charged phosphonium groups, the pore size and thickness of THPC@TCNF membrane would be signi cantly improved in comparison with TCNF membrane. Therefore, THPC@TCNF membrane was expected to have enhanced permeate ux and antibacterial property.
TCNFs prepared by TEMPO-oxidized tunicate cellulose had ber-like morphology, whose average length, average width, and aspect ratio were 10.0 ± 0.3 µm, 6.5 ± 0.2 nm, and ~ 1540, respectively ( Figure S2). After vacuum-assisted ltration, TCNF membrane exhibited nanoporous structure with an average pore size of 50.4 ± 2.5 nm (Fig. 2a), which could generate capillary force and easily form a hydration layer during oil/water separation process (Peng et al. 2022). After modi cation, THPC@TCNF membrane displayed a relatively loose surface with a larger pore size (62.2 ± 2.9 nm), as shown in Fig. 2b. According to Hagen-Poiseuille equation, the enlarged pore size of membranes would improve their permeate ux during oily wastewater puri cation. Besides, the thickness of TCNF membrane also augmented from 11.8 µm to 21.1 µm after reacting with THPC, whilelarger pore could be clearly discovered in the cross-section of THPC@TCNF membrane (Fig. 2c). THPC modi ed TCNFs that isolated from membrane had ber-like morphology, and some black dots could be observed on the surface of TCNFs ( Figure S4). These results indicated that THPC was successfully grafted onto TCNFs via esteri cation reaction and the morphology of TCNFs was not changed.

Structure of THPC@TCNF membranes
To comprehend the chemical structure of THPC@TCNF membrane, FTIR spectra of TCNF membrane and THPC@TCNF membrane were contrasted. As displayed in Fig. 3a, a new absorption band appeared at 1725 cm − 1 in the spectrum of THPC@TCNF membrane, which was ascribed to ν C=O in the ester bond formed through esteri cation of -COOH and -OH (Kang et al. 2017). The absorption band located at 3426 cm − 1 corresponding to -OH stretching vibration became stronger in the spectrum of THPC@TCNF membrane due to the introduction of THPC. Moreover, a new characteristic peak at 133.5 eV could be discovered in the XPS spectrum of THPC@TCNF membrane (Fig. 3b), which could be assigned to phosphor element of THPC. In the high-resolution P 2p spectra, the characteristic peak could only be found in the spectrum of THPC@TCNF membrane ( Figure S5), revealing the presence of phosphonium groups in the membrane. In the C 1s spectrum of THPC (Fig. 3c), different types of carbon atoms mainly included alphatic carbon (C-H, 284.68 eV), hydroxyls (C-O, 286.30 eV), and carbon-phosphorus covalent bond (C-P, 286.80 eV), where related peaks shifted to 284.91 eV, 286.55eV, and 288.15eV, respectively, in the spectrum of THPC@TCNF membrane (Fig. 3d), indicating that the binding of THPC and TCNFs resulted in a higher binding energy of various types of carbon atoms. Furthermore, a new XPS peak at 288.15 eV in C 1s spectrum of THPC@TCNF membrane, which was assigned to carboxyl groups (O-C = O) due to presence of TCNFs.
In addition, the distribution of phosphonium groups in the membrane was also investigated by EDS mapping of P and Cl elements. As shown in Fig. 3e and Figure S6, green and red dots were uniformly distributed in the images, revealing that phosphonium groups were uniformly distributed in the membrane. To determine degree of substitution of phosphonium groups, the carboxyl content of THPC@TCNF membranes prepared with different dosage of THPC was measured by conductance titration. As displayed in the Fig. 3f, the carboxyl group content of TCNF membrane was 1.25 mmol g − 1 , because TCNFs were prepared by TEMPO-oxidization strategy and carboxyl groups generated on the surface of TCNFs. When the dosage of THPC increased from 0-16%, carboxyl content gradually decreased from 1.25 mmol g − 1 to 0.28 mmol g − 1 . As the dosage of THPC further increased to 32%, the carboxyl content changed slightly because of the steric hindrance effect of phosphonium groups.

Wettability and separation performance of THPC@TCNF membranes
The wettability of the membranes is strong depended on their chemical composition and surface topography, which is crucial to the separation e ciency and fouling resistance during oily wastewater puri cation (Li et al. 2021a; Zhang et al. 2017). As shown in Fig. 4a, a water droplet could quickly spread out on the surface of THPC@TCNF membrane in air, indicating the superhydrophilicity of membrane. Four kinds of oils, including petroleum ether, pump oil, dimethicone, and chloroform, were employed to study the wettability of THPC@TCNF membrane. The underwater oil contact angles were 158.7°, 164.6°, 160.4°, and 158.1°, for petroleum ether, pump oil, dimethicone, and chloroform, respectively (Fig. 4b).
These results indicated that various oil droplets with different density and viscosity could be repelled by the THPC@TCNF membranes, revealing their underwater superoleophobic properties. The wettability of THPC@TCNF membranes was consistent with that of TCNF membranes ( Figure S7), which con rmed that the introduction of THPC into TCNF membranes did not change their wettability.
As exhibited in Fig. 4c, the underwater anti-oil adhesion property of the THPC@TCNF membrane was evaluated with dynamic approach-compress-detach test. Under water, after contacting the surface of the THPC@TCNF membrane, the chloroform droplets were slowly compressed downward to form an ellipse. When gradually lifted, the chloroform droplets could quickly overcome the adhesion force and detach from the THPC@TCNF membrane surface without any deformation. The results illustrated that the THPC@TCNF membrane was expected to have low oil adhesion and excellent antifouling performance during oil/water separation.
To ascertain the optimal dosage of THPC, the water permeate ux of different THPC@TCNF membranes was shown in Figure S8. As THPC dosage increased from 0 to 16 wt%, the water ux of membranes increased from 1208 ± 35 L m − 2 h − 1 bar − 1 to 1931 ± 64 L m − 2 h − 1 bar − 1 , which was attributed to the enlarged pore size of the membranes. When the dosage of THPC increased to 32 wt%, the water permeate ux of membrane was 1928 ± 63 L m − 2 h − 1 bar − 1 . This result was agreed with the result of carboxyl content, suggesting that the water permeate ux of membrane was dependence of carboxyl content in the membrane. Thus, THPC@TCNF membrane that was prepared by using 16% THPC was selected for the puri cation of oily wastewater. Four kinds of oil/water nanoemulsions with average particle size of 200 nm were prepared, which showed bright pathway under the irradiation of a laser pen because of Tyndall effect (Fig. 5a-d).
To investigate the durability of THPC@TCNF membrane, the separation of petroleum ether/water nanoemulsion for 60 min was displayed in Fig. 5g. During the continuous separation of petroleum ether/water nanoemulsion, the water permeate ux of THPC@TCNF membrane was stable, which remained at 1735 ± 58 L m − 2 h − 1 bar − 1 after 60 min separation. Until 180 min, the ux was still higher than half of the initial ux and remained at 1121 ± 35 L m − 2 h − 1 bar − 1 ( Figure S9a). In addition, as shown in Fig. 5h, the cycle performance of THPC@TCNF membrane was also measured. Both water permeate ux and oil rejection of THPC@TCNF membrane hardly decreased after 10 cycles. Even after 20 cycles, the water ux slightly decreased to 1523 L m − 2 h − 1 bar − 1 , while the oil rejection was still as high as 99.02% ( Figure S9b), demonstrating excellent durability of THPC@TCNF membrane.

Antibacterial activity and mechanical stability
The presence of bacteria in oily wastewater could decrease the separation e ciency of membranes during long-term use (Bethke et al. 2018), so antibacterial properties of membranes are critical for the practical application of membranes. As shown in Fig. 6a, a slight increase in the colony forming units on the TCNF membrane could be observed in comparison with the control group, because cellulose is a kind of polysaccharide. Impressively, no colony of B. subtilis was observed on THPC@TCNF membrane, because all B. subtilis were effectively killed after contacting with THPC@TCNF membrane for 1 h. Compared to control and TCNF membrane, the antibacterial e ciency of THPC@TCNF membrane calculated according to the colony forming units before and after contact almost 100% (Fig. 6b). After contacting with membranes for 1 h, the morphology of B. subtilis cultured on TCNF membrane was B. subtilis, whereas the surface of B. subtilis cultured on THPC@TCNF membrane became loose and fractured, as shown in Fig. 6c and Figure S10, because quaternary phosphonium salts are capable of destroying a wide range of microorganisms by destabilizing bacterial cell walls ). The antibacterial property of HPC@TCNF membrane would be bene tting to prevent the corrosion of bacteria.
Furthermore, both bending test and abrasion test were employed for evaluating the mechanical stability of the THPC@TCNF membranes. As presented in Fig. 6d, the pump oil contact angle still maintained at 163° after 200 bending cycles. The surface morphology of THPC@TCNF membrane hardly changed before and after bending test (Fig. 6e). Additionally, abrasion test of THPC@TCNF membrane was also carried out by rubbing THPC@TCNF membrane on a 2000 mesh sandpaper (Fig. 6f). The THPC@TCNF membrane was adhered to a glass slide with a weight of 100 g, and the membrane was subsequently dragged on the sandpaper for 10 cm in the horizontal and vertical directions, which was a wear cycle. After 30 cycles, some ne moving traces appeared on the surface of the THPC@TCNF membrane, the complete structure of the membrane was not destroyed (Fig. 6g). Simultaneously, the underwater OCA remained at an angle of 159° (Fig. 6f), showing underwater superoleophobicity of the membrane. These results revealed that the THPC@TCNF membranes have good bending and wear resistance properties. Mechanical stability is a necessary condition to ensure long-term high-e ciency oil/water separation, which was bene cial to the practical application of THPC@TCNF membranes in the eld of oil/water separation.

Conclusions
We successfully fabricated THPC@TCNF membranes through grafting tetrahedral-structured THPC onto the surface of TCNFs via esteri cation reaction. THPC effectively increased the surface average pore size and interlayer spacing of TCNF membranes, resulting in the enhancement of water permeate ux of membrane. On the other hand, the positively charged phosphonium groups endowed the TCNF membrane with good antibacterial properties, which could prevent the membrane from the corrosion of bacteria. These THPC@TCNF membranes exhibited good superhydrophilicity, underwater superoleophobicity, and nanoporous structure, which enabled them to separate various oil/water nanoemulsions. Moreover, THPC@TCNF membranes possessed excellent durability, mechanical stability, and cycle performance. This work provided a simple strategy for fabricating high water permeate ux and antibacterial membranes, which had great potential in oily wastewater treatment.

Declarations Notes
Page 10/17 The authors declare no competing nancial interest.    Contact angle measurement of THPC@TCNF membranes with respect to wettability. Photograph of a water drop in air (a), underwater various types of oil drops on THPC@TCNF membranes (b). Dynamic approach-compress-detach oil-adhesion test photograph for THPC@TCNF membrane (c).

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