Alternative proton exchange membrane based on a bicomponent anionic nanocellulose system.

As integral parts of fuel cells, polymer electrolyte membranes (PEM) facilitate the conversion of hydrogen's chemical energy into electricity and water. Unfortunately, commercial PEMs are associated with high costs, limited durability, variable electrochemical performance and are based on perfluorinated polymers that persist in the environment. Nanocellulose-based PEMs have emerged as alternative options given their renewability, thermal and mechanical stability, low-cost, and hydrophilicity. These PEMs take advantage of the anionic nature of most nanocelluloses, as well as their facile modification with conductive functional groups, for instance, to endow ionic and electron conductivity. Herein, we incorporated for the first time two nanocellulose types, TEMPO-oxidized and sulfonated, to produce a fully bio-based PEM and studied their contribution separately and when mixed in a PEM matrix. Sulfonated nanocellulose-based PEMs are shown to perform similarly to commercial and bio-based membranes, demonstrating good thermal-oxidative stability (up to 190 °C), mechanical robustness (Young's modulus as high as 1.15 GPa and storage moduli >13 GPa), and high moisture-uptake capacity (ca. 6330 % after 48 h). The introduced nanocellulose membranes are shown as promising materials for proton-exchange material applications, as required in fuel cells.

Enhanced low-cost lipopeptide biosurfactant production by Bacillus velezensis from residual glycerin.

Biosurfactants (BSFs) are molecules produced by microorganisms from various carbon sources, with applications in bioremediation and petroleum recovery. However, the production cost limits large-scale applications. This study optimized BSFs production by Bacillus velezensis (strain MO13) using residual glycerin as a substrate. The spherical quadratic central composite design (CCD) model was used to standardize carbon source concentration (30 g/L), temperature (34 °C), pH (7.2), stirring (239 rpm), and aeration (0.775 vvm) in a 5-L bioreactor. Maximum BSFs production reached 1527.6 mg/L of surfactins and 176.88 mg/L of iturins, a threefold increase through optimization. Microbial development, substrate consumption, concentration of BSFs, and surface tension were also evaluated on the bioprocess dynamics. Mass spectrometry Q-TOF-MS identified five surfactin and two iturin isoforms produced by B. velezensis MO13. This study demonstrates significant progress on BSF production using industrial waste as a microbial substrate, surpassing reported concentrations in the literature.

Water-Repellent Spray for Textiles Using Plant Waste from Conifer Trees.

A superhydrophobic textile coating, applied by using a home drying machine, was developed as an aqueous dispersion of waxes that were extracted from recycled Christmas trees. Because the bulk extraction of waxes yielded a mixture of hydrophobic and hydrophilic compounds, a purification process was tested to determine if removing noncrystallizing wax components would enhance the performance of the coating. The performances of coatings created from the crude and enriched extracts were compared, and no significant difference in hydrophobicity was found. Moreover, although the enriched coating was slightly more breathable, there was not enough of an improvement to justify the additional purification steps, rendering the crude extract more industrially viable. Overall, Christmas tree waxes are readily sourced and are capable of producing superhydrophobic coatings without the need for a costly purification step.

Preparation of isolated guard cells, containing cell walls, from Vicia faba.

Stomatal movement, initiated by specialized epidermal cells known as guard cells (GCs), plays a pivotal role in plant gas exchange and water use efficiency. Despite protocols existing for isolating GCs through proplasting for carrying out biochemical, physiological, and molecular studies, protocals for isolating GCs with their cell walls still intact have been lacking in the literature. In this paper, we introduce a method for the isolation of complete GCs from Vicia faba and show their membrane to remain impermeable through propidium iodide staining. This methodology enables further in-depth analyses into the cell wall composition of GCs, facilitating our understanding of structure-function relationship governing reversible actuation within cells.

Development of a pH-responsive intelligent label using low molecular weight chitosan grafted with phenol red for food packaging applications.

In this study, we successfully developed a screen-printed pH-responsive intelligent label using low molecular weight chitosan grafted with phenol red (LCPR) as a colorant for screen printing ink. The LCPR was synthesized via a Mannich reaction, and its successful grafting was confirmed through FT-IR, UV-vis, and NMR spectroscopy. The LCPR exhibited lower crystallinity and thermal stability compared to low molecular weight chitosan (LC) and demonstrated zwitterionic behavior. To create intelligent labels, the LCPR-based ink was efficiently printed on cotton substrates with high resolution. The label exhibited remarkable sensitivity to buffer pH solutions and ammonia gas, leading to distinctive color changes from orange to red to purple. Additionally, the label showed excellent reversibility, storage stability, and leaching resistance to different food simulant solutions. The label was utilized to monitor shrimp freshness, successfully detecting a noticeable color shift upon spoilage. These findings highlight the significant potential of the LCPR-based label as an intelligent food packaging solution, offering pH-responsiveness and color stability for qualitative freshness detection of protein-rich food.

Proton Exchange Membranes from Sulfonated Lignin Nanocomposites for Redox Flow Battery Applications.

Redox flow batteries (RFBs) are increasingly being considered for a wide range of energy storage applications, and such devices rely on proton exchange membranes (PEMs) to function. PEMs are high-cost, petroleum-derived polymers that often possess limited durability, variable electrochemical performance, and are linked to discharge of perfluorinated compounds. Alternative PEMs that utilize biobased materials, including lignin and sulfonated lignin (SL), low-cost byproducts of the wood pulping process, have struggled to balance electrochemical performance with dimensional stability. Herein, SL nanoparticles are demonstrated for use as a nature-derived, ion-conducting PEM material. SL nanoparticles (NanoSLs) can be synthesized for increased surface area, uniformity, and miscibility compared with macrosized lignin, improving proton conductivity. After addition of polyvinyl alcohol (PVOH) as a structural backbone, membranes with the highest NanoSL concentration demonstrated an ion exchange capacity of 1.26 meq g-1, above that of the commercial PEM Nafion 112 (0.98 meq g-1), along with a conductivity of 80.4 mS cm-1 in situ, above that of many biocomposite PEMs, and a coulombic efficiency (CE), energy efficiency (EE) and voltage efficiency (VE) of 91%, 68% and 78%, respectively at 20 mA cm-2. These nanocomposite PEMs demonstrate the potential for valorization of forest biomass waste streams for high value clean energy applications.

Improvement in the Thermomechanical Properties and Adhesion of Wood Fibers to the Polyamide 6 Matrix by Sequential Ball Milling Technique.

The engineering thermoplastics industry has largely limited the use of natural fiber reinforcements due to their susceptibility to low-onset thermal degradation and water absorption. Therefore, in order to utilize these economically viable and environmentally friendly materials effectively through common composite fabrication methods such as hot pressing, safeguarding them from thermal degradation becomes essential. This work presents a viable industrially technique called sequential ball milling for processing unbleached softwood kraft pulp fibers (PF) with an engineering thermoplastics polyamide 6 (PA6) with high melting temperatures (>220 °C). An additional eco-friendly modification step that employs ball milling and cellulose nanocrystal (CNC) has been implemented in this study to enhance the mechanical properties of the composites. Special attention is given to fine-tuning key variables, such as milling duration and PF particle size, to produce optimal composites. Leveraging the ability of sequential ball milling to evenly distribute pulp fibers into PA6, a 160% increase in Young's modulus was achieved with the incorporation of 30 wt % PF. Importantly, the introduction of a 5 wt % CNC modifying agent elevated Young's modulus to 4.3 GPa, marking a 187% improvement over unmodified PA6. Diverse techniques, including rheological analyses, thermomechanical evaluations, morphological examinations, and assessments of moisture absorption, were utilized to validate the efficiency of the suggested processing approach and the modification phase.

Plant-based, aqueous, water-repellent sprays for coating textiles.

Novel superhydrophobic coatings, that are both biodegradable and biosourced, have the potential to revolutionize the water-repellent coating industry. Here, water-repellent coatings were prepared from commercially unavailable plant waxes, isolated using solvent extraction and characterized using DSC, GC-MS and DLS. In the first stage, a plant survey was conducted to identify an ideal plant source for the final spray, in which Whatman filter paper was submerged in a wax-solvent solution with recrystallization occurring upon air-drying. In the second stage, aqueous, PFC-free wax dispersions were prepared, coated onto textiles (cotton and polyester), and heat-treated with a home drying machine to allow for the spreading and recrystallization of the waxes. In both stages, SEM visualization verified the coating's morphology, and contact angle measurements showed them to be superhydrophobic. It was concluded that, using less coating material than commercial coatings, high-performing petroleum-free coatings could be made and applied onto textiles of various polarities.

Spatially Directed Pyrolysis via Thermally Morphing Surface Adducts.

Combustion is often difficult to spatially direct or tune associated kinetics-hence a run-away reaction. Coupling pyrolytic chemical transformation to mass transport and reaction rates (Damköhler number), however, we spatially directed ignition with concomitant switch from combustion to pyrolysis (low oxidant). A 'surface-then-core' order in ignition, with concomitant change in burning rate,is therefore established. Herein, alkysilanes grafted onto cellulose fibers are pyrolyzed into non-flammable SiO2 terminating surface ignition propagation, hence stalling flame propagating. Sustaining high temperatures, however, triggers ignition in the bulk of the fibers but under restricted gas flow (oxidant and/or waste) hence significantly low rate of ignition propagation and pyrolysis compared to open flame (Liñán's equation). This leads to inside-out thermal degradation and, with felicitous choice of conditions, formation of graphitic tubes. Given the temperature dependence, imbibing fibers with an exothermically oxidizing synthon (MnCl2 ) or a heat sink (KCl) abets or inhibits pyrolysis leading to tuneable wall thickness. We apply this approach to create magnetic, paramagnetic, or oxide containing carbon fibers. Given the surface sensitivity, we illustrate fabrication of nm- and µm-diameter tubes from appropriately sized fibers.

On the Roles of Cellulose Nanocrystals in Fiber Cement: Implications for Rheology, Hydration Kinetics, and Mechanical Properties.

Fiber cement reinforced with pulp fibers is one of the key drivers for the decarbonization of nonstructural building materials, where the inclusion of sustainable pulp fibers at high proportions (i.e., > 8 wt %) renders poor workability of fiber-cement slurry with a concomitant loss in mechanical strength. Petrochemical-derived superplasticizers, i.e., polycarboxylates (PCEs), are predominantly used in fiber cement (including cement mortars) because they dramatically improve (content <0.5 wt %) the slurry rheology but reduce the rate of hydration and weaken the strength of the cured composite. Thus, it is crucial to explore renewable and bio-based superplasticizers devoid of any negative traits (if possible) of the conventional PCEs. In this study, we examined wood-derived cellulose nanocrystals (CNCs) as a multifunctional additive in fiber cement (bleached pulp fiber content: 8 wt %). In fiber cement, variation of the content (0.02-4 wt %) of CNCs resulted in improvement in the shear thinning behavior of the fiber-cement slurry and thereafter increased the hydration kinetics at high CNC contents (2-4 wt %). Notably, the flexural strength of the composite also exhibited improvement upon the addition of CNCs; the maximum strength was observed at 4 wt % of CNCs. Overall, the beneficial roles of CNCs afforded >10 wt % (in-total) bio-based content in fiber cement without compromising the mechanical strength and curing time (compared to PCEs); hence, the findings of this study could unravel new avenues in interface engineering of cement composites leveraging the multifunctional features of biomaterials, thus enhancing sustainability.

Unraveling lignin degradation in fibre cement via multidimensional fluorometry.

Pulp fibre reinforced cement (fibre cement) has the potential to become a forerunner in mitigating the carbon dioxide (CO2) footprint of non-structural materials for residential and commercial structures. However, one of the significant bottlenecks in fibre cement is its poor chemical stability in the alkaline cement matrix. To date, probing the health of pulp fibre in cement is lengthy and laborious, requiring mechanical and chemical separations. In this study, we have demonstrated that it is possible to understand the chemical interactions at the fibre-cement interfaces by tracking lignin in a solid state without using any additional chemicals. For the first time, multidimensional fluorometry is employed for the rapid assessment of the structural change (degradation) of lignin in fibre cement as an indicator of pulp fibre health; providing an excellent platform for the germination of resilient fibre cement with high content of natural lignocellulosic fibre.

Direct milling: Efficient, facile, and green method for processing fibrillated cellulose/polymeric nanocomposites with boosted thermomechanical and rheological performance.

Bringing biobased nanomaterials into polymer manufacturing is essential to enhance polymers' properties and address the challenges posed by plastic waste. Using polymers such as polyamide 6 (PA6) in advanced industries, e.g., automotive sector, has been impeded as a direct consequence of their inability to meet the required mechanical properties. Herein, we utilize bio-based cellulose nanofibers (CNFs) to enhance the properties of PA6 by green processing, with no footprint on the environment. We address the issue of the dispersion of the nanofillers in polymeric matrices and demonstrate direct milling (cryo-milling and planetary ball milling) to facilitate a thorough integration of the components. Nanocomposites incorporating 1.0 wt% CNF, processed by pre-milling followed by compression molding, are shown to possess a storage modulus of 3.8 ± 0.2 GPa, Young's modulus of 2.9 ± 0.2 GPa, and ultimate tensile strength of 63 ± 3 MPa (all measured at room temperature). To show the superiority of direct milling in achieving these properties, other frequent approaches used to disperse CNF in polymers, such as solvent casting and hand mixing, are meticulously investigated and compared for the performance of their resulting specimens. The ball-milling method is demonstrated to provide PA6-CNF nanocomposites with excellent performance, better than solvent casting, with no associated environmental concerns.

Assessment of Dietary Intake in Three Cohorts of Advanced Age in Two Countries: Methodology Challenges.

OBJECTIVES: Dietary intake information is key to understanding nutrition-related outcomes. Intake changes with age and some older people are at increased risk of malnutrition. Application, difficulties, and advantages of the 24-hour multiple pass recall (24hr-MPR) dietary assessment method in three cohorts of advanced age in the United Kingdom (UK) and New Zealand (NZ) is described. PARTICIPANTS: The Newcastle 85+ study (UK) recruited a single year birth cohort of people aged 85 years during 2006-7. LiLACS NZ recruited a 10-year birth cohort of Maori (indigenous New Zealanders) aged 80-90 years and a single year birth cohort of non-Maori aged 85 years in 2010. MEASUREMENTS: Two 24hr-MPR were conducted on non-consecutive days by trained assessors. Pictorial resources and language were adapted for the New Zealand and Maori contexts. Detailed methods are described. RESULTS: In the Newcastle 85+ study, 805 (93%) participants consented to the 24-MPR, 95% of whom completed two 24hr-MPR; in LiLACS NZ, 218 (82%) consented and 203 (76%) Maori and 353 (90%) non-Maori completed two 24hr-MPR. Mean time to complete each 24hr-MPR was 22 minutes in the Newcastle 85+ study, and 45 minutes for Maori and 39 minutes for non-Maori in LiLACS NZ. Dietary assessment of participants residing in residential care and those requiring proxy respondents were successfully included in both studies. Most participants (83-94%) felt that data captured by the 24hr-MPR reflected their usual dietary intake. CONCLUSIONS: Dietary assessment using 24hr-MPR was successful in capturing detailed dietary data including information on portion size and time of eating for over 1300 octogenarians in the UK and New Zealand (Maori and non- Maori). The 24hr-MPR is an acceptable method of dietary assessment in this age group.

Northward Expansion of Amblyomma americanum (Acari: Ixodidae) into Southwestern Michigan.

Amblyomma americanum (Linnaeus) (Acari: Ixodidae) (lone star tick) is an aggressive, generalist parasite that vectors numerous important human and animal pathogens. In recent decades its geographic range has expanded northwards from endemic regions in the southeastern and southcentral United States. In 2019 five questing A. americanum ticks, comprising two life stages were detected at one site in southwestern Michigan, satisfying one CDC criterium for an established population for the first time in recent history in the state. To better characterize the extent of emerging A. americanum, we conducted active surveillance (i.e., drag sampling) in summer 2020 throughout Michigan's southern counties and detected one adult A. americanum from each of six widespread sites, including where they had been detected in 2019. A larger established population was identified at another site in Berrien County, which yielded 691 A. americanum comprising three life stages, and questing phenologies here were similar to that reported for other endemic regions. Statewide surveillance in 2021 revealed no A. americanum outside of Berrien County, but establishment criteria were met again at the two sites where established populations were first detected respectively in 2019 and 2020. These observations may represent the successful invasion of A. americanum into Michigan. Data from passive (1999-2020) and active surveillance (2004-2021) efforts, including a domestic animal sentinel program (2015-2018), are reported to provide context for this nascent invasion. Continued active surveillance is needed to help inform the public, medical professionals, and public health officials of the health risks associated with this vector.

Snail Based Carbonated-Hydroxyapatite Material as Adsorbents for Water Iron (II).

Carbonated hydroxyapatite (CHAp) adsorbent material was prepared from Achatina achatina snail shells and phosphate-containing solution using a wet chemical deposition method. The CHAp adsorbent material was investigated to adsorb aqua Fe(II) complex; [Fe(H2O)6]2+ from simulated iron contaminated water for potential iron remediation application. The CHAp was characterized before and after adsorption using infrared (IR) and Raman spectroscopy. The IR and the Raman data revealed that the carbonate functional groups of the CHAp adsorbent material through asymmetric orientation in water bonded strongly to the aqua Fe(II) complex adsorbate. The adsorption behaviour of the adsorbate onto the CHAp adsorbent correlated well to pseudo-second-order kinetics model, non-linear Langmuir and Freundlich model at room temperature of a concentration (20-100 mg L-1) and contact time of 180 min. The Langmuir model estimated the maximum adsorption capacity to be 45.87 mg g-1 whereas Freundlich model indicated an S-type isotherm curvature which supported the spectroscopy revelation.

Early Outpatient Treatment for Covid-19 with Convalescent Plasma.

BACKGROUND: Polyclonal convalescent plasma may be obtained from donors who have recovered from coronavirus disease 2019 (Covid-19). The efficacy of this plasma in preventing serious complications in outpatients with recent-onset Covid-19 is uncertain. METHODS: In this multicenter, double-blind, randomized, controlled trial, we evaluated the efficacy and safety of Covid-19 convalescent plasma, as compared with control plasma, in symptomatic adults (≥18 years of age) who had tested positive for severe acute respiratory syndrome coronavirus 2, regardless of their risk factors for disease progression or vaccination status. Participants were enrolled within 8 days after symptom onset and received a transfusion within 1 day after randomization. The primary outcome was Covid-19-related hospitalization within 28 days after transfusion. RESULTS: Participants were enrolled from June 3, 2020, through October 1, 2021. A total of 1225 participants underwent randomization, and 1181 received a transfusion. In the prespecified modified intention-to-treat analysis that included only participants who received a transfusion, the primary outcome occurred in 17 of 592 participants (2.9%) who received convalescent plasma and 37 of 589 participants (6.3%) who received control plasma (absolute risk reduction, 3.4 percentage points; 95% confidence interval, 1.0 to 5.8; P = 0.005), which corresponded to a relative risk reduction of 54%. Evidence of efficacy in vaccinated participants cannot be inferred from these data because 53 of the 54 participants with Covid-19 who were hospitalized were unvaccinated and 1 participant was partially vaccinated. A total of 16 grade 3 or 4 adverse events (7 in the convalescent-plasma group and 9 in the control-plasma group) occurred in participants who were not hospitalized. CONCLUSIONS: In participants with Covid-19, most of whom were unvaccinated, the administration of convalescent plasma within 9 days after the onset of symptoms reduced the risk of disease progression leading to hospitalization. (Funded by the Department of Defense and others; CSSC-004 ClinicalTrials.gov number, NCT04373460.).

Adjustable film properties of cellulose nanofiber and cellulose nanocrystal composites.

A novel nanocomposite comprised of cellulose nanocrystals (CNCs) and 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO) oxidized cellulose nanofibers (TOCNFs) was prepared through solution casting to evaluate potential improvements of the mechanical performance compared to individual reinforcements alone. Such materials can be implemented as mechanical reinforcements in polymer composites, especially when less weight is desired. Dissipative particle dynamics (DPD) simulations, in combination with polarized light microscopy and atomic force microscopy, were analyzed to evaluate the morphology of these combined cellulose nanomaterial (CNM) films. Our results indicate that TOCNFs provide enhanced translational mobility to CNCs which become incorporated near the crystalline domains of TOCNFs. This mobility enables CNCs to increase the rigidity of the network without sacrificing elongation and toughness. The combination of these materials provides improved ultimate tensile strength and elongation without sacrificing the Young's modulus. Therefore, a combination of these materials can be used to develop nanocomposites with enhanced mechanical properties.

The Influence of Pineapple Leaf Fiber Orientation and Volume Fraction on Methyl Methacrylate-Based Polymer Matrix for Prosthetic Socket Application.

This work reports on the use of low-cost pineapple leaf fiber (PALF) as an alternative reinforcing material to the established, commonly used material for prosthetic socket fabrication which is carbon-fiber-reinforced composite (CFRC) due to the high strength and stiffness of carbon fiber. However, the low range of loads exerted on a typical prosthetic socket (PS) in practice suggests that the use of CFRC may not be appropriate because of the high material stiffness which can be detrimental to socket-limb load transfer. Additionally, the high cost of carbon fiber avails opportunities to look for an alternative material as a reinforcement for composite PS development. PALF/Methyl Methacrylate-based (MMA) composites with 0°, 45° and 90° fiber orientations were made with 5-50 v/v fiber volume fractions. The PALF/MMA composites were subjected to a three-point flexural test to determine the effect of fiber volume fraction and fiber orientation on the flexural properties of the composite. The results showed that 40% v/v PALF/MMA composite with 0° fiber orientation recorded the highest flexural strength (50 MPa) and stiffness (1692 MPa). Considering the average load range exerted on PS, the flexural performance of the novel composite characterized in this work could be suitable for socket-limb load transfer for PS fabrication.

Understanding the Mechanical Reinforcement of Metal-Organic Framework-Polymer Composites: The Effect of Aspect Ratio.

The aspect ratio (AR) of filler particles is one of the most critical determinants for the mechanical properties of particle-reinforced polymer composites. However, it has been challenging to solely study the effect of particle AR due to the difficulties of controlling AR without altering the physical and chemical properties of the particle. Herein, we synthesized PCN-222, a zirconium-based porphyrinic metal-organic framework (MOF) with preferential longitudinal growth as a series of particles with ARs increasing from 3.4 to 54. The synthetic MOF conditions allowed for the chemical properties of the particles to remain constant over the series. The particles were employed as reinforcers for poly(methyl methacrylate) (PMMA). MOF-polymer composite films were fabricated using doctor-blading techniques, which facilitated particle dispersion and alignment in the PMMA matrix, as revealed by optical microscopy and wide-angle X-ray diffraction. Mechanical measurements showed that both elastic and dynamic moduli increased with particle AR and particle concentrations but started to decrease as particle loading increased beyond 0.5 wt % (1.12 vol %). The data obtained at low particle loadings were fitted well with the Halpin-Tsai model. In contrast, the percolation model and the Cox model were unable to adequately fit the data, indicating the mechanical reinforcement in our system mainly originated from efficient load transfer between particles and the matrix in the particle orienting direction. Finally, we showed that the thermal stability of composite films increased with the addition of MOF particles because of the high thermal degradation temperature and restricted polymer chain mobility.

Randomized Controlled Trial of Early Outpatient COVID-19 Treatment with High-Titer Convalescent Plasma.

BACKGROUND: The efficacy of polyclonal high titer convalescent plasma to prevent serious complications of COVID-19 in outpatients with recent onset of illness is uncertain. METHODS: This multicenter, double-blind randomized controlled trial compared the efficacy and safety of SARS-CoV-2 high titer convalescent plasma to placebo control plasma in symptomatic adults ≥18 years positive for SARS-CoV-2 regardless of risk factors for disease progression or vaccine status. Participants with symptom onset within 8 days were enrolled, then transfused within the subsequent day. The measured primary outcome was COVID-19-related hospitalization within 28 days of plasma transfusion. The enrollment period was June 3, 2020 to October 1, 2021. RESULTS: A total of 1225 participants were randomized and 1181 transfused. In the pre-specified modified intention-to-treat analysis that excluded those not transfused, the primary endpoint occurred in 37 of 589 (6.3%) who received placebo control plasma and in 17 of 592 (2.9%) participants who received convalescent plasma (relative risk, 0.46; one-sided 95% upper bound confidence interval 0.733; P=0.004) corresponding to a 54% risk reduction. Examination with a model adjusting for covariates related to the outcome did not change the conclusions. CONCLUSION: Early administration of high titer SARS-CoV-2 convalescent plasma reduced outpatient hospitalizations by more than 50%. High titer convalescent plasma is an effective early outpatient COVID-19 treatment with the advantages of low cost, wide availability, and rapid resilience to variant emergence from viral genetic drift in the face of a changing pandemic. Trial Registration: ClinicalTrials.gov number, NCT04373460.