key: cord-0269546-w9tfr9lr
authors: Shen, Hongchen; Zhou, Zhe; Wang, Haihuan; Zhang, Mengyang; Han, Minghao; Shen, Yun; Shuai, Danmeng
title: Photosensitized Electrospun Nanofibrous Filters for Capturing and Killing Airborne Coronaviruses under Visible Light Irradiation
date: 2021-07-30
journal: bioRxiv
DOI: 10.1101/2021.07.29.454404
sha: 5645ca7dfca846bc97c9cdb77722808c516e84ce
doc_id: 269546
cord_uid: w9tfr9lr

To address the challenge of the airborne transmission of SARS-CoV-2, photosensitized electrospun nanofibrous membranes were fabricated to effectively capture and inactivate coronavirus aerosols. With an ultrafine fiber diameter (∼ 200 nm) and a small pore size (∼ 1.5 µm), the optimized membranes caught 99.2% of the aerosols of the murine hepatitis virus A59 (MHV-A59), a coronavirus surrogate for SARS-CoV-2. In addition, rose bengal was used as the photosensitizer for the membranes because of its excellent reactivity in generating virucidal singlet oxygen, and the membranes rapidly inactivated 98.9% of MHV-A59 in virus-laden droplets only after 15 min irradiation of simulated reading light. Singlet oxygen damaged the virus genome and impaired virus binding to host cells, which elucidated the mechanism of disinfection at a molecular level. Membrane robustness was also evaluated, and no efficiency reduction for filtering MHV-A59 aerosols was observed after the membranes being exposed to both indoor light and sunlight for days. Nevertheless, sunlight exposure photobleached the membranes, reduced singlet oxygen production, and compromised the performance of disinfecting MHV-A59 in droplets. In contrast, the membranes after simulated indoor light exposure maintained their excellent disinfection performance. In summary, photosensitized electrospun nanofibrous membranes have been developed to capture and kill airborne environmental pathogens under ambient conditions, and they hold promise for broad applications as personal protective equipment and indoor air filters. Synopsis Photosensitized electrospun nanofibrous filters with excellent capture-and-kill performance against coronaviruses were designed and implemented to prevent the airborne transmission of COVID-19. Table of Contents

Airborne transmission of SARS-CoV-2 has been recognized as an important route for spreading The electrospun membranes have a reduced pore size (tens of nanometers to several micrometers), 64 an increased specific surface area, and enhanced porosity to enable efficient aerosol filtration and 65 a low pressure drop in filtration. 7 Furthermore, the surface and volume charges within the 66 electrospun membranes improve aerosol filtration through electrostatic attraction. 10, 11 Particularly, 67 our previous study has underscored that the electrospun membranes caught up to 99.9% of 68 coronavirus aerosols whose size was within or close to that of the most penetrating particles in 69 mechanical air filtration. 12 However, like most masks and air filters currently used on the market, 70 electrospun membranes only physically capture viral aerosols but they do not inactivate the viruses, 71 which could lead to secondary contamination and potential infection (e.g., via the contact of the 72 contaminated masks/air filters or viruses dislodged from the masks/filters). 13

The goal of our study is to (i) develop highly efficient and robust photosensitized electrospun 75 nanofibrous membranes that can both physically capture and chemically disinfect coronavirus 76 aerosols, and (ii) elucidate the mechanism of photooxidation and inactivation of coronaviruses at 77 5 a molecular level. We selected dye photosensitizers as antimicrobial additives for electrospinning, 78 because they produce highly reactive singlet oxygen ( 1 O2) for effective and rapid virus disinfection, 79 they can be excited under the irradiation of visible light that is readily available in indoor 80 environments, they are low cost, and some of them have been approved by the U.S. Food and Drug 81 Administration for biomedical applications and thus pose little to no health risks to humans (e.g., 82 rose bengal (RB)). Our membranes enable a self-cleaning feature for disinfecting coronaviruses We first optimized the composition for preparing the photosensitized electrospun nanofibrous 89 membranes to yield the best NaCl aerosol filtration efficiency, the lowest pressure drop in filtration, 90 and the highest reactivity for producing 1 O2 under visible light irradiation. Next, we tested the 91 filtration efficiency and inactivation kinetics of coronavirus aerosols to best understand 92 performance of the optimum membranes for controlling the airborne transmission of COVID-19. 93 We then conducted a thorough mechanistic investigation on how the membranes inactivated electrospinning of 10-20 wt% of PVDF, the solution feeding rate, electric field, and 110 electrospinning duration was maintained at 0.6 mL h -1 , 1 kV cm -1 , and 20 min, respectively. For 111 electrospinning 8 wt% of PVDF, the solution feeding rate, electric field, and electrospinning 112 duration was kept at 0.4 mL h -1 , 1 kV cm -1 , and 30 min, respectively, to reduce bead formation.

The dyes including 0.3 wt% of RB, 0.015 wt% of methylene blue hydrate (MB), 0.3 wt% of crystal 114 violet (CV), 0.015 wt% of (-)-riboflavin (RF), and 0.003 wt% of toluidine blue O (TBO) were 115 used. Mass percentage of PVDF and the dyes was calculated with respect to the total mass of the 116 electrospinning dope solution, and the maximum dye concentration was selected based on dye 117 solubility in the solution. The two-layer photosensitized electrospun membranes were denoted as 118 PVDFx-dye name (e.g., PVDF15-RB). Membranes that were electrospun from 15 wt% of PVDF 119 without dyes onto a layer of PP fabrics (VWR ® Basic Protection Face Mask) were also fabricated 120 for comparison (i.e., PVDF15). Three-layer photosensitized electrospun membranes (here referred to as sandwiched membranes), 123 which consisted of an additional PP fabric top layer (Amazon, skin friendly non-woven fabrics) 124 on the top of the two-layer photosensitized electrospun membranes, were particularly assembled 125 for aerosol filtration and pressure drop tests. The sandwiched structure protected the electrospun 126 layer from destruction during and after filtration (e.g., removal from the filter holder). Based on 127 our previous study, the PP fabrics had negligible impact on aerosol removal and pressure drop, 128 because of their very large pore size compared with the aerosol size (~120 µm versus < 2 µm) and with a lamp-to-membrane distance of 15 cm) and simulated indoor light (3% 7W white LED with 143 a lamp-to-membrane distance of 33 cm), and the steady-state 1 O2 concentration ([ 1 O2]ss) was 144 calculated by dividing the measured first-order decay rate constant of FFA by the second-order 145 8 reaction rate constant between 1 O2 and FFA (1.2 × 10 8 M -1 s -1 ). Simulated reading light had a 146 higher photon flux and optical power density than those of simulated indoor light, and they were 147 recorded in Figure S1 and Table S1 (AvaSpec-2048 Fiber Optic Spectrometer). FFA was 148 monitored by high performance liquid chromatography (Shimadzu LC-20AT Prominence). for RT-qPCR quantification, as described in Section 2.4. Photooxidation damaged the viral genes 223 and prevented its RT-qPCR quantification, and only the intact genes were able to be determined. The filtration efficiency of NaCl aerosols was further examined for those PVDFx-RB membranes.

Our previous study has demonstrated the filtration efficiency of NaCl aerosols was a conservative (Figures 2a and 2b) . In contrast, in the (Figure 2 and 2b) . That is said, 98.9% of coronaviruses were inactivated after only 15 min (Figure S4) , and thus much more 398 rapid coronavirus inactivation was observed under light exposure with a stronger intensity. We 399 also found that the ORF5 gene damage rate of MHV-A59 in the droplets was lower than that of 400 aerosolized MHV-A59 captured on the membranes after exposure to simulated reading light (kgene 401 = 0.0195 ± 0.0030 versus 0.0461 ± 0.0044 min -1 , p < 0.05, Figure 2 and 2b) . The slower ORF5 402 gene damage could be attributed to the shorter lifetime and diffusion length of 1 O2 in water 403 compared with that in the air (2 μs in distilled water versus 2.80 s in the air), due to energy 404 dissipation resulted from the collision between 1 O2 and water molecules. 35, 36 The dissolved oxygen 405 concentration in water was also lower than the oxygen concentration in the air (< 10 mg L -1 in the 406 water versus 275 mg L -1 in the air at 25 ℃), which might limit 1 O2 production in the aqueous 407 phase. 37 Since 1 O2 was the key ROS in photosensitization and it did not bias damaging the viral 408 genome and inactivating the viruses, the liquid setup is considered as a conservative system to 409 evaluate the inactivation of MHV-A59 aerosols captured on the photosensitized electrospun 410 membrane. Therefore, it is reasonable to speculate that the inactivation of infectious or viable 411 MHV-A59 in the aerosols captured on the membrane will be much faster than that in the droplets, 412 though the infectivity of viral aerosols was not quantifiable in our study. 

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