Cryopreservation protocol induces reactive oxygen species formation and leads to a decrease in post-thawed sperm quality and fertilizing capacity. We investigated the effect of resveratrol and its optimal concentration on the quality of post-thawed boar semen and the level of malondialdehyde (MDA). Our investigations showed that the supplement of 50–100 µM resveratrol enhanced the quality of the post-thawed semen and decreased the MDA level. However, the maximum concentration of resveratrol (250 µM) tended to decrease sperm motility, sperm viability, intact acrosomes, and mitochondrial membrane potential. The protective or cytotoxic effect of resveratrol on the quality of frozen boar sperm depends on the concentration of resveratrol and extender composition as well as limiting antioxidant uptake capacity in sperm.p < 0.05). Furthermore, the 50 µM resveratrol group was significantly more protective than the control group in terms of intact acrosome, mitochondrial membrane potential, and level of MDA (p < 0.05). Nonetheless, the detrimental effect of resveratrol was found at a concentration of 250 µM. In conclusion, the addition of 50–100 µM resveratrol to a freezing extender is the optimal concentration for enhancing the quality of cryopreserved boar sperm.This study aimed to determine the effect of resveratrol and its optimal concentration on the quality of frozen-thawed (FT) boar sperm. Semen ejaculates were obtained from 13 Duroc boars aged between 1.5 and 3 years. The sperm sample was separated into 7 groups based on the concentrations of resveratrol in the freezing extender, which were 0 (control), 25, 50, 75, 100, 125, and 250 µM, respectively. The sperm was frozen using liquid nitrogen vapor and thawed at 50 °C for 12 s. After thawing, total motility, progressive motility, viability, intact acrosomes, mitochondrial membrane potential and level of MDA were assessed. The supplementation of 50–100 µM resveratrol improved the sperm motility and viability of FT sperm in comparison to the control group ( The cryopreservation procedure induces sperm damage in a number of ways under the impact of multiple factors, including significant fluctuations in temperatures, osmotic imbalances, and oxidative stress . Oxidaticysteine ,11, N-accysteine ,12 and gcysteine ,14. In rcysteine , resveracysteine , and curcysteine  also revResveratrol  is a natural polyphenolic compound found in grapes, mulberries, plums, peanuts, and other plant derived products . ResveraThirteen fertile Duroc boars (aged 1.5–3 years) utilized for routine artificial insemination in a commercial herd were used in this study. The boars were kept in an enclosed facility with an evaporative cooling system and separate enclosures. Each boar was fed a commercial diet containing 15–16% of protein twice daily, and water was provided ad libitum via water nipple.6 sperm/mL) was measured using Photometer SDM1 , subjective motility was analyzed at 400× using a light microscope, and sperm morphology was stained with William’s staining method and evaluated using a light microscope. Cryopreservation was only conducted on ejaculate that exhibited morphological normality and a motility rate of 80% [One ejaculate from each of the 13 individual boars was collected using the gloved-hand technique. During collection, sperm-rich fractions were collected after filtering through filter paper  . Within e of 80% ,31.v/v) ratio with the ModenaTM extender . The extended semen was carefully transferred into 50 mL centrifuge tubes, equilibrated at 15 °C for 120 min, and then centrifuged at 15 °C at 800× g for 10 min . After discarding the supernatant, the sperm pellet was re-suspended in a ratio of about 1-2:1 (v/v) with a lactose–egg yolk extender at a concentration of 1.5 × 109 spermatozoa/mL. The extender consisted of 80 mL of an 11% lactose solution and 20 mL of egg yolk. In this phase, the sperm sample was divided into 7 groups based on the resveratrol  concentration ). All of the sperm samples were chilled to 5 °C for 90 min. Each sample group was mixed with freezing extender, which is composed of 89.5% lactose–egg yolk extender with 9% (v/v) glycerol and 1.5% (v/v) Equex-STM® , to a final concentration of 1.0 × 109 sperm/mL [TM extender at 37 °C for 15 min [The conventional method of sperm freezing with liquid nitrogen was used to freeze all semen samples. After collection, the fresh semen was extended in a 1:1 . Briefly, 3 µL of a semen sample was pipetted into a pre-warmed disposable counting chamber  and maintained at 37 °C during analysis. At least 600 sperm cells were counted per analysis across five fields of each sample. The results of analysis were expressed as the percentage of total sperm motility and progressive motility as well as kinetic parameters such as curvilinear velocity , average pathway velocity , straight line velocity , the amplitude of the lateral head displacement , straightness , and linearity . Motile spermatozoa were defined when VCL ≥ 24 μm/s and ALH > 1 μm. The term “progressive motility” (PMOT) has been interpreted as the presence of sperm exhibiting a VCL ≥ 48 μm/s and VSL < 10 μm/s [Sperm motility was analyzed by computer-assisted sperm motility analysis (CASA)  and Ethidiumhomodimer-1 (EthD-1). Briefly, 10 µL of semen was combined with 2.7 µL of SYBR-14  and 10 µL of 1.17 µM EthD-1. The mixture was incubated at 37 °C for 15 min. Under a fluorescence microscope with a magnification of 400×, 200 sperms were evaluated. The sperm stained with SYBR-14/EhtD-1 were separated into viable sperm and non-viable sperm. The nuclei of living sperm with an undamaged plasma membrane turned green, while the nuclei of damaged membranes or dead sperm turned red . The resThe evaluation of acrosome integrity was conducted using a staining protocol of fluorescein isothiocyanate-labeled peanut agglutinin (FITC-PNA) and EthD-1 staining. A total of 10 µL of semen and 10 µL of EthD-1 were incubated at 37 °C for 15 min. Then, 5 µL of the mixture was applied to a glass slide and allowed to air dry. The slide was fixed with 95% ethanol for 30 s and then allowed to air dry. A total of 40 µL of FITC-PNA (100 µg/mL in PBS) was spread onto the slides and kept in a humid container at 4 °C for 30 min. The slides were then rinsed with cold PBS and air dried. At least 200 green-stained, live sperms were evaluated under a 1000× magnification fluorescence microscope and classified as intact acrosome or non-intact acrosome . The resl,l′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide (1.53 mM) . A total of 50 µL of diluted semen was mixed with 3 µL of a 2.4 mM propidium iodide (PI) and 3 µL of a 1.53 mM JC-1 solution in DMSO and incubated for 10 min at 37 °C in a dark container. Under a fluorescence microscope with a magnification of 400×, two hundred sperm were evaluated. Midpiece staining revealed that sperm with a high mitochondrial membrane potential were yellow-orange, while sperm with a low membrane potential were green  trichloroacetic acid was added to each tube, followed by 1 mL of 0.375% (w/v) thiobarbituric acid. The sample tubes were vortexed and boiled in a water bath for 10 min. Next, the samples were placed in an ice box to stop the reaction. Finally, the samples were centrifuged at 4 °C at 800× g for 10 min. The supernatant (2 mL) from each sample was transferred to a cuvette for analysis using a UV-visible spectrophotometer . Absorbance was read at 532 nm. The levels of MDA were calculated from the MDA standard curve and expressed as µM/mL [The MDA levels of semen samples were measured using the thiobarbituric acid reactive substance (TBARS) assay. Briefly, 250 µL of a semen sample was added with 250 µL of 0.2 mM ferrous sulphate and 250 µL of 1 mM ascorbic acid and incubated at 37 °C for 60 min. Thereafter, 1 mL of 15% , sperm viability, acrosome integrity, mitochondrial membrane potential, and level of MDA were analyzed using SPSS Statistics for Windows, version 26.0 . The data were tested for normality and then analyzed with a one-way ANOVA followed by Duncan’s multiple range test to determine significant differences in all parameters. A statistically significant difference was defined as p < 0.05). However, there was no significant difference in post-thawed sperm parameters among the 50, 75, and 100 µM resveratrol groups. For sperm kinetic parameters, the VCL, VSL, VAP, and ALH of the 50 µM resveratrol group were significantly higher when compared to the control group (p < 0.05). Nonetheless, the addition of resveratrol did not significantly affect the linearity and straightness of sperm movement. In this study, the highest total motility (49.2 ± 2.2%), sperm viability (52.2 ± 2.4%), and sperm with high mitochondrial membrane potential (60.7 ± 2.0%) were observed in the 50 µM resveratrol group. Nonetheless, the maximum concentration of resveratrol (250 µM) tended to decrease total sperm motility, sperm viability, intact acrosomes, and mitochondrial membrane potential. Furthermore, the 250 µM resveratrol group showed the lowest percentage of acrosome integrity.The descriptive statistics of the quality of fresh boar semen are presented in p < 0.05). The 50 µM resveratrol group had the lowest MDA level (1.08 ± 0.1 µM/mL). However, the level of MDA among treatment groups was not statistically significant.The effects of resveratrol on lipid peroxidation during cryopreservation are shown in It is noteworthy that sperm cryopreservation induced excessive ROS generation that led to lipid peroxidation of the sperm plasma membrane as well as DNA oxidation. These events contribute to the impairment of sperm structure and function, the reduction in sperm survival rate, and the decrease in fertilizing capability ,2. In thOur investigations are similar to those of other species, in that resveratrol was supplemented during the chilled storage and sperm cryopreservation processes. Bucak et al.  demonstrIt is well documented that sperm cells need energy to conserve their metabolism and function . It has In this study, it is proven that resveratrol, a natural polyphenolic antioxidant, is capable of inhibiting lipid peroxidation and improving sperm motility, viability, intact acrosomes, and mitochondrial membrane potential in frozen-thawed boar sperm. The addition of 50–100 µM resveratrol to a freezing extender enhanced sperm motility and the viability of cryopreserved boar sperm. However, a detrimental effect of resveratrol was found at a concentration of 250 µM.


Streptococcus agalactiae outbreak in tilapia has caused huge losses in the aquaculture industry worldwide. In Malaysia, several studies have reported the isolation of S. agalactiae, but no study has reported the isolation of S. agalactiae phages from tilapia or from the culture pond. Here, the isolation of the S. agalactiae phage from infected tilapia is reported and it is named as vB_Sags-UPM1. Transmission electron micrograph (TEM) revealed that this phage showed characteristics of a Siphoviridae and it was able to kill two local S. agalactiae isolates, which were S. agalactiae smyh01 and smyh02. Whole genome sequencing (WGS) of the phage DNA showed that it contained 42,999 base pairs with 36.80% GC content. Bioinformatics analysis predicted that this phage shared an identity with the S. agalactiae S73 chromosome as well as several other strains of S. agalactiae, presumably due to prophages carried by these hosts, and it encodes integrase, which suggests that it was a temperate phage. The endolysin of vB_Sags-UPM1 termed Lys60 showed killing activity on both S. agalactiae strains with varying efficacy. The discovery of the S. agalactiae temperate phage and its antimicrobial genes could open a new window for the development of antimicrobials to treat S. agalactiae infection.The  Streptococcus agalactiae, or Lancefield’s Groups B streptococcus (GBS), is a Gram-positive coccus that is beta-hemolytic and a facultative anaerobe. It is divided into ten molecular serotypes  based on the capsular polysaccharide (cps) gene and can be further classified based on clonal complex (CC), sequence type (ST), phage type, and virulence factor . Ex. Ex9 pfuProtein-coding gene prediction was performed using Glimmer3 v1.5  and Prodhttp://eggnog-mapper.embl.de/) (accessed on 5 October 2021) [The whole phage genome was searched for the closest relative using BLASTn against the default nucleotide collection (nr/nt) and virus database (taxid: 10239). Average nucleotide identity (ANI) of a phage was calculated using JSpecies with a cutoff of 95% to indicate similar taxonomic affiliation . The puter 2021)  and BLASer 2021) . Phylogeer 2021) , which wer 2021) . Geneiouer 2021)  and Cluser 2021)  alignmener 2021)  using de2O. The PCR was performed using Eppendorf® Mastercycler® Nexus Thermal Cycler  set to 98 °C for 2 min ; 35 cycles of 98 °C for 1 min, 57 °C for 30 s, and 72 °C for 1 min (amplification); and 72 °C for 5 min . PCR amplicons and plasmid pET were digested with NcoI and XhoI restriction enzymes  for 1 h at 37 °C before ligation using T4 Ligase  at 22 °C for 1 h. Recombinant DNA was introduced into E. coli using the traditional heat shock method. Positive clones were selected on LB with kanamycin plates and screened using colony PCR using forward (5′-TCCCGCGAAATTAATACGAC-3′) and reverse primers (5′-TATGCTAGTTATTGCTCAGC-3′) that recognized regions of pET-28a that flanked the multiple cloning site. For gene induction, overnight culture of recombinant E. coli was added into fresh LB medium and incubated with shaking at 37 °C. Upon reaching an OD600nm of 0.4, a final concentration of 0.5 mM IPTG was added into the culture and incubated at 18 °C for 24 h. Cells were harvested and washed using buffer  and sonicated for eight minutes with a resting period of 30 s. Soluble protein in the supernatant was applied into cOmplete His-Tag Purification Resin  to purify proteins that contained His tag. The eluted protein was concentrated in buffer using Pierce™ Protein Concentrator with a polyethersulfone membrane, 10 K molecular weight cutoff . The protein concentration was quantified using Pierce® BCA Protein Assay Kit  following the manufacturer’s protocol. The presence of Lys60 in crude as well as in purified fractions was detected using 12% SDS-PAGE with 10–180 kDa PageRuler™ Prestained Protein Ladder , followed by staining with Coomassie blue or Western Blot. After protein transfer from gel to Amersham Hybond polyvinylidene difluoride membrane , the membrane was stained with 1:1000 6× His-tag monoclonal antibody  followed by staining with 1:10,000 goat anti-mouse IgG secondary antibody, alkaline phosphatase (AP) . The substrate 5-Bromo-4chloro-3-indolyl-phospate (BCIP) was used, which reacted with nitro blue tetrazolium (NBT) for detection. The gel and Western Blot membrane was viewed using Amersham Imager 600 .A pair of forward primer containing NcoI site (5′-GCCCCATGGCAACTTATCAAGAATA-3′) and reverse primer containing XhoI site (5′-CCGCTCGAGCGGCATATCTGTTGCATCA-3′) was used to amplify the Lys60 gene following the PCR reaction—4 µL 5× Phusion HF Buffer , 1 µL 10 µM forward primer, 1 µL 10 µM reverse primer, 0.4 µL 10 mM dNTP mix, 0.2 µL Phusion™ Plus DNA Polymerase , and 1 µL 100 ng/µL DNA template and 12.4 µL HS. agalactiae at 600 nm absorbance. Briefly, the concentration of proteins was standardized to 645.6 µg/mL. Serial dilutions of 484.2 µg/mL, 322.8 µg/mL, 242.1 µg/mL, 161.4 µg/mL, and 80.7 µg/mL were prepared in buffer . Fifty mL of the early log phase of S. agalactiae culture (OD600nm = 0.2) was centrifuged, washed, and resuspended in 5 mL final volume of buffer. A volume of 178 µL of bacteria was mixed with 2 µL, 1M CaCl2  and dispensed into the 96-well plate in triplicate for each concentration of Lys60. A total of 20 µL of protein dilutions and empty buffer was added into bacterial suspensions, mixed, and incubated at 30 °C. At 5 min intervals, the absorbance at OD600nm was measured for a period of 2 h using iMark™ Microplate Reader . Additionally, the antimicrobial activity was determined by calculating the percentage of viable bacteria relative to control after treating with Lys60  or untreated for 0, 40, and 80 min. The CFU/mL of treated and untreated bacteria was determined and the percentage viability at each time point post-treatment was calculated based on the following formula.The lytic activity of Lys60 was determined using turbidity reduction assay of 600nm, and lysogeny efficiency between S. agalactiae smyh01 and smyh02, the data were analyzed using unpaired t-test where a p value of <0.05 was considered statistically significant. For the comparison of antimicrobial activity of Lys60 endolysin on S. agalactiae smyh01 and smyh02 between different time points post-infection, and between different enzyme concentration, the data were analyzed using two-way ANOVA with Tukey’s post hoc test, where a p value of <0.05 was considered statistically significant.The data were analyzed using GraphPad Prism 8. Firstly, the normality of datasets was determined using Shapiro–Wilk test, whereby most data were normally distributed (parametric data). For the comparison of burst size, absorbance at OD


There were no statistically significant differences in the concentration versus time curve of cabozantinib (AUCcabozantinib) between the control group and the RT2Gy×3 f’x and RT9Gy×3 f’x groups in the concurrent and the sequential regimens. However, compared to those in the control group, the Tmax, T1/2 and MRT decreased by 72.8% (p = 0.04), 49.0% (p = 0.04) and 48.5% (p = 0.04) with RT2Gy×3 f’x in the concurrent regimen, respectively. Additionally, the T1/2 and MRT decreased by 58.8% (p = 0.01) and 57.8% (p = 0.01) in the concurrent RT9Gy×3 f’x group when compared with the control group, respectively. The biodistribution of cabozantinib in the heart increased by 271.4% (p = 0.04) and 120.0% (p = 0.04) with RT2Gy×3 f’x in the concurrent and sequential regimens compared to the concurrent regimen, respectively. Additionally, the biodistribution of cabozantinib in the heart increased by 107.1% (p = 0.01) with the RT9Gy×3 f’x sequential regimen. Compared to the RT9Gy×3 f’x concurrent regimen, the RT9Gy×3 f’x sequential regimen increased the biodistribution of cabozantinib in the heart , liver , lung  and kidneys . No cabozantinib was detected in the brain in any of the groups. The AUC of cabozantinib is not modulated by irradiation and is not affected by treatment strategies. However, the biodistribution of cabozantinib in the heart is modulated by off-target irradiation and SBRT doses simultaneously. The impact of the biodistribution of cabozantinib with RT9Gy×3 f’x is more significant with the sequential regimen than with the concurrent regimen.The aim of this study was to evaluate the radiotherapy (RT)-pharmacokinetics (PK) effect of cabozantinib in concurrent or sequential regimens with external beam radiotherapy (EBRT) or stereotactic body radiation therapy (SBRT). Concurrent and sequential regimens involving RT and cabozantinib were designed. The RT–drug interactions of cabozantinib under RT were confirmed in a free-moving rat model. The drugs were separated on an Agilent ZORBAX SB-phenyl column with a mobile phase consisting of 10 mM potassium dihydrogen phosphate (KH Hepatocellular carcinoma (HCC) was the sixth most commonly diagnosed cancer and the third leading cause of cancer death worldwide in 2020 . The sysSorafenib inhibits HCC through the inhibition of vascular endothelial growth factor (VEGF) and partly through the inhibition of the RAS/RAF/MEK/ERK mitogen-activated protein kinase (MAPK) at the level of RAF ,6. In thP-glycoprotein (P-gp) could be affected by RT  × 100. The relative standard deviation (RSD) was calculated as follows: precision (% RSD) = [standard deviation (SD)/Cobs] × 100. The same data were used to determine both accuracy and precision. The intraday precision (% RSD) and accuracy (% bias) values were within ± 15%, which were considered to be in the acceptable experimental concentration range. Recovery was assessed by comparing the peak areas of the spiked samples post extraction with the standard solution at 0.05, 0.5 and 10 μg/mL.The intra-assay variabilities for cabozantinib were determined by quantitating six replicates at concentrations of 0.05, 0.1, 0.5, 1, 5 and 10 μg/mL using the HPLC method described above on the same day (intraday) and for six consecutive days (interday). The accuracy (% bias) was calculated as follows: accuracy (% bias) = [the nominal concentration . Adult male Sprague–Dawley rats (300 ± 20 g body weight) were provided by the Laboratory Animal Center at National Yang Ming Chiao Tung University . All animal experiments followed the guidelines and procedures for the care of laboratory animals at National Yang Ming Chiao Tung University.2Gy) and RT 9 Gy (RT9Gy) with three fractions with concurrent or sequential cabozantinib. The data were collected from six rats per group.A freely moving rat model was designed for the current study . Briefly0Gy (cabozatinib×3 d); (B) whole liver (2 Gy with 3 fractions) RT2Gy×3 f’x concurrent with cabozantinib (6 mg/kg) for 3 days; (C) whole liver RT2Gy×3 f’x sequential with cabozantinib (6 mg/kg) for 3 days; (D) SBRT (9 Gy with 3 fractions) RT9Gy×3 f’x concurrent with cabozantinib (6 mg/kg) for 3 days; and (E) RT9Gy×3 f’x sequential with cabozantinib (6 mg/kg) for 3 days  sham group, cabozantinib (6 mg/kg) only for 3 days (d) with RTr 3 days . Rats weBlood samples were collected via polyethylene tubing (PE-50) implanted into the jugular vein of each rat in a heparin-rinsed vial. An aliquot of 100–120 µL of blood was collected at time intervals of 0, 5, 15, 30, 45, 60, 90, 120, 150, 180, 210 and 240 min following drug administration. At each time point, 200 μL of blood was drawn into heparin-rinsed Eppendorf tubes and then centrifuged at 13,000 rpm for 10 min at 4 °C to obtain plasma. Plasma was stored at −20 °C until analysis. Each collected blood sample was transferred to a heparinized microcentrifuge tube and centrifuged at 13,000 rpm for 10 min. The resulting plasma (50 μL) was then mixed with 150 μL of internal standard solution (10 μg/mL). The denatured protein precipitate was separated by vortexing for 20 s and centrifuged at 13,000 rpm for 10 min at 4 °C.Six hours after cabozantinib administration for 3 days , blood samples were collected as mentioned above. The brain, liver, heart, spleen, lung and kidney were collected and weighed. These collected samples were stored at −20 °C until analysis.g for 10 min at 4 °C. The supernatant was collected, placed in brown Eppendorf tubes, and stored at −20 °C until analysis. Briefly, each organ sample (50 μL) was combined with 150 μL of IS solution (diethylstilbestrol) for protein precipitation. Finally, 20 μL of filtrate was subjected to HPLC analysis.The thawed organ samples were homogenized in 50% aqueous acetonitrile (the ratio of sample weight to volume was 1:5), and the homogenate was centrifuged at 13,000× 𝑡1/2), volume of distribution at steady state (Vss) and mean residence time (MRT), were calculated using the pharmacokinetics calculation software WinNonlin Standard Edition, Version 1.1  by a compartmental method. Relative bioavailability (RB %) = (AUCirradiated/AUCcontrol) × 100. The results are presented as the means ± standard deviations.Pharmacokinetic parameters, including the area under the concentration versus time curve (AUC), clearance (CL), elimination half-life  for Windows, version 20.0 . All data are expressed as the mean ± standard deviation (SD). One-way analysis of variance (ANOVA) was used for comparisons between groups, and statistically significant differences were defined as * 𝑝 < 0.05 or ** 9Gyx3f’x sequential regimen had a greater impact on the biodistribution of cabozantinib than the concurrent regimen. The current understanding of the systemic effects of cabozantinib with localized irradiation could be beneficial for exploring RT as a synergistic tool in HCC treatment strategies.Taken together, these data suggest that the PK of cabozantinib cannot be modulated by irradiation. However, the biodistribution of cabozantinib can be affected by RT, especially in the heart and kidney. The RT


We also investigate the underlying mechanism for this enhanced performance. Our results show that Ni/Ce0.8Gd0.2O2-δ at 600 °C can achieve nearly complete ammonia decomposition, resulting in a hydrogen production rate of 2008.9 mmol.g−1.h−1 with minimal decrease over 150 h. Density functional theory calculations reveal that the recombinative desorption of nitrogen is the rate-limiting step of ammonia decomposition over Ni. Our characterizations indicate that Ni/Ce0.8Gd0.2O2-δ exhibits a high concentration of oxygen vacancies, highly dispersed Ni on the surface, and abundant strong basic sites. These properties significantly enhance the associative desorption of N and strengthen the metal support interactions, resulting in high catalytic activity and stability. We anticipate that the mechanism could be applied to designing additional catalysts with high ammonia decomposition performance at relatively low temperatures.Ammonia decomposition is a promising method to produce high-purity hydrogen. However, this process typically requires precious metals  as catalysts to ensure high efficiency at relatively low temperatures. In this study, we propose using several Ni/Gd Hydrogen is considered an important carrier of energy for the future due to its clean and renewable characteristics ,2. Howev2O3 catalysts, using Co, Ni, and Fe as dopants to improve the catalytic activity and delved into the synergy between the dopants and Ni to disclose the mechanism for the enhanced performance. The results show that bimetallic CoNi/Al2O3 exhibits almost full ammonia decomposition at 700 °C with the stable trend in the temperature range of 650–750 °C. We can conclude from these results that the doping strategy can effectively enhance the performance of Ni. However, the catalytic performance is still unsatisfactory at temperatures <700 °C.Currently, the attention of numerous researchers has been directed toward transition-metal-based catalysts, such as Ni , Fe 13]13], and 2O4 catalysts with different support morphologies and studied the effect of the support morphologies on performance [2O4-LDH exhibits superior ammonia decomposition performance due to its high surface area and well-dispersed Ni. Additionally, the porous alumina matrix’s structural features were observed by Wang et al. [In addition to the doping strategy, the literature also suggests that the activity of Ni is contingent on the support, with the support properties exerting a significant influence on its ammonia decomposition performance ,21. Receformance . The fing et al.  to promoxCe1-xO2-δ as catalysts for ammonia decomposition by tuning the properties of GdxCe1-xO2-δ support. The results show both high ammonia conversion and stability for Ni/Gd0.2Ce0.8O2-δ at 600 °C. The improved performance was studied through a combination of DFT calculation and different physicochemical characterizations, revealing the fundamental mechanism behind it.In this study, we have synthesized several Ni/GdxCe1-xO2-δ supports and the active compound are in a crystalline phase. Importantly, based on the XRD patterns, there is no evidence of new compound formation or chemical reaction occurring between the support and the active compound. Scherer’s formula (Equation (1)) was used to evaluate crystallite sizes of the supports based on the (111) peak of GdxCe1-xO2-δ:0.1Ce0.9O2-δ, Gd0.2Ce0.8O2-δ, Gd0.3Ce0.7O2-δ, Gd0.4Ce0.6O2-δ are 5.15 Å, 5.17 Å, 5.20 Å, and 5.23 Å. We can attribute this discrepancy to the larger ionic radii of Gd3+ (1.19Å) compared with Ce3+ (1.01Å), as previously reported in the literature [The X-ray diffraction (XRD) patterns in terature .0.2Ce0.8O2-δ > Gd0.3Ce0.7O2-δ > Gd0.1Ce0.9O2-δ > Gd0.4Ce0.6O2-δ. This indicates that Gd0.2Ce0.8O2-δ shows the most oxygen vacancies among all the samples. Therefore, we can conclude that adjusting the molar ratio of Gd and Ce can increase the oxygen vacancy concentration, with Gd0.2Ce0.8O2-δ showing the highest concentration.The EPR profiles displayed in xCe1-xO2-δ with different Gd: Ce molar ratios. The results show the catalysts featured in the gathered spherical structures. In addition, the morphologies of the catalysts were found to be similar across all samples. This suggests that varying the molar ratio of Gd:Ce did not significantly impact the surface morphologies. Therefore, the difference in the catalytic activity was not influenced by the surface morphologies of the catalysts. 2/g. The actual Ni loading of all catalysts was estimated to be near 9.8%, which is only slightly lower than the nominal Ni amount. The Ni particle size of the catalysts was also not significantly influenced by the Gd: Ce molar ratio. However, comparing the Ni dispersion of the catalysts on the surface, we can find that Ni/Gd0.2Ce0.8O2-δ displays the highest Ni dispersion of 20.7%. In addition, the amount of Ni dispersion is positively related to that of the oxygen vacancy concentration. This observation is consistent with recent findings by Liu et al. that the oxygen vacancy on the surface of catalysts can promote the dispersion of active sites on the surface [Additional textural properties in terms of surface area, actual Ni loading, Ni dispersion, and Ni particle size are shown in  surface .2-TPR profiles of the catalysts were used to evaluate the metal–support interaction between the active sites (Ni) with the GdxCe1-xO2-δ support. As is shown in xCe1-xO2-δ support was usually reduced at 300–600 °C. It is difficult to differentiate the reduction peak of NiO with the significant metal–support interaction from that of GdxCe1-xO2-δ. However, the metal–support interaction can be evaluated by the number of reduction peaks at temperatures lower than 300 °C. After comparing the H2-profiles of the catalysts, it is evident that the reduction peaks of NiO/Gd0.2Ce0.8O2-δ are mostly located at temperatures above 300 °C, whereas the remaining catalysts exhibit partial reduction peaks at temperatures below 300 °C. In conclusion, NiO/Gd0.2Ce0.8O2-δ displays the most significant metal–support interaction compared with the other catalysts. Previous studies [0.2Ce0.8O2-δ results from the increased oxygen vacancy concentration, which is achieved through the manipulation of support properties.H studies ,29 demonxCe1-xO2-δ catalysts was evaluated in the temperature range of 300–700 °C with an interval of 50 °C. As we can see in xCe1-xO2-δ with different Gd: Ce ratios, we can observe that the support properties of the catalysts have a significant influence on their catalytic performance for ammonia decomposition. For Ni/Gd0.4Ce0.6O2-δ, the catalyst only started to react at a temperature of 400 °C. Its ammonia decomposition efficiency was only 80.8% at 600 °C. The catalyst’s ammonia decomposition efficiency only approached 100% when the temperature reached as high as 700 °C. For Ni/Gd0.2Ce0.8O2-δ, the decomposition reaction began at temperatures higher than 300 °C, and the reaction efficiency gradually increased to 29.9% at 450 °C, followed by a sharp increase to 100% at 600 °C and GHSV of 30,000 L.kg−1.h−1  for the catalysts were evaluated as 81.65 kJ/mol, 58.45 kJ/mol, 65.52 kJ/mol, and 83.02 kJ/mol for Ni/Gd0.1Ce0.9O2-δ, Ni/Gd0.2Ce0.8O2-δ, Ni/Gd0.3Ce0.7O2-δ, and Ni/Gd0.4Ce0.6O2-δ, respectively , which is higher than the majority of the catalysts reported in the literature, demonstrating its high potential for practical ammonia decomposition applications.To highlight the novelty of the catalysts in this study, we also compared the performance of Ni/Gdterature . The res2 was estimated to be 1.74 eV, which is much higher than that of the stepwise dehydrogenation processes. Therefore, the rate-limiting step of the ammonia decomposition reaction is the associative desorption of N.A DFT calculation was conducted to investigate the reaction route of ammonia decomposition over Ni (110), calculating the potential energy curves for each, thus uncovering the rate-limiting step during this process. The ammonia decomposition reaction was found to consist of two steps, as shown in xCe1-xO2-δ support on the abundance of the strong basic sites can be used to explain the enhanced performance of the Ni/Gd0.2Ce0.8O2-δ at medium temperatures.Previous studies ,33 have 2-TPD profiles of the Ni/ GdxCe1-xO2-δ profiles. The spectrum can be classified into two types according to the position of the CO2 desorption peaks. The peaks at temperatures <300 °C can be linked to the weak basic sites (Bronstone sites) and the peaks at high temperature (>450 °C) can be ascribed to the contribution of the strong basic sites (Lewis sites). Comparing the CO2-TPD profiles of the catalysts, we can find that all catalysts exhibit two CO2 desorption peaks at low and high temperatures. In addition, Ni/Gd0.2Ce0.8O2-δ had the highest area of the peak at a high temperature, followed by Ni/Gd0.3Ce0.7O2-δ, Ni/Gd0.1Ce0.9O2-δ, and Ni/Gd0.4Ce0.6O2-δ. This indicates that Gd0.2Ce0.8O2-δ support can significantly improve the abundance of the strong basic sites on the surface, benefiting the associative desorption of N on the surface, thus leading to enhanced catalytic performance toward the ammonia decomposition. 3-TPSR can yield crucial insights into catalyst surface reactions. Our DFT calculations have demonstrated that N recombinative desorption is the rate-limiting step, and the energy barriers can be estimated from N2 formation profiles obtained from NH3-TPSR 3·6H2O, Ce(NO3)3·6H2O, Gd(NO3)3·6H2O, and citric acid were dissolved in deionized water to form a solution with a metal ion/citric acid molar ratio of 1:1.1. After that, the solution was agitated and heated at 95 °C. The PEG-400 was employed as a gel promoter. The solution was heated at 95 °C overnight to remove all the water in the solution, thus forming the gel. After the evaporation of the water in the solution, the gel was dehydrated and annealed in air at 850 °C for 6 h to derive the final catalysts. The nominal mass ratio of metallic Ni was set as 10 wt%, while the nominal molar ratio of Ce:Gd varied between 9:1 and 6:4.Ni/Gd2/N2 for 1 h at 700 °C to reduce NiO before the characterization. The Scanning Electron Microscopy coupled with Energy Dispersive X-ray Spectroscopy (SEM-EDS), Quanta 200FEG with an Oxford EDS system was used for the morphological analysis of the catalysts. An ASAP 2010 instrument  was used to conduct the N2 physisorption characterization, which was used to determine the specific surface area of the catalysts. To quantify the actual content, an inductively coupled plasma optical emission spectrometry (ICP-OES) was carried out on an Optima 5300 DV instrument. The metal dispersion of Ni was assessed on an ASAP 2010C chemisorption analyzer (Micromeritics) using the static volumetric H2 chemisorption method. It was assumed that each nickel atom would adsorb a single hydrogen atom. XRD patterns were obtained using the PANalytical X’Pert-Pro powder X-ray diffractometer with CuKα radiation (0.1541 nm), with a fixed scanning step size of 0.0334° in the 10−80° range, and operated at 40 kV and 40 mA. All catalysts were pretreated in 250 mL/min 10% H2-TPR analysis using 10% H2/Ar mixture (25 mL·min−1) as the reducing agent. The procedure for conducting the H2-TPR reaction was as follows: First, 30 mg of catalysts was placed into the equipment. After that, the catalysts were pretreated at 140 °C under 30 mL/min He for 30 min to remove the possible moisture on the surface. Then, the samples were cooled down to room temperature to initiate the reduction reaction. The temperature was ramped at a rate of 10 °C·min−1 from room temperature to 800 °C.TPR Win v 1.50  was used for H2-temperature-programmed desorption (TPD) measurement was performed using the same instrument as for the H2-TPR. The characterization was conducted to evaluate the distribution of the basic sites on the surface of catalysts. Typically, the catalysts were pre-reduced in 10% H2/Ar at 600 °C for 4 h to remove all reducible oxygen, followed by cooling the catalysts to room temperature. After that, the catalysts were exposed to CO2 atmosphere at 60 °C for 1 h to adsorb CO2. After the termination of the adsorption reaction, the reactor was heated to 100 °C and flushed with He for 1 h to remove the physisorbed CO2. Finally, the reactor temperature was heated from 100 °C to 800 °C with a ramp of 10 °C/min. CO3-temperature-programmed surface reaction (TPSR) was measured using a homemade fixed-bed reactor. To begin the experiment, the reactor was first evacuated and heated to the desired temperature. Ammonia was then introduced into the reactor at a constant flow rate, and the temperature was gradually increased at 10 K/min. As the temperature increased, the reaction between the ammonia and the catalyst surface occurred, resulting in the production of various products. The effluent gas was continuously monitored by a Pfeiffer Vacuum mass spectrometer.The NHadsorbate and Esurface denote the total energies of the gaseous species and the clean slab.The Vienna Ab-initio Simulation Package (VASP) was used to conduct density functional theory (DFT) calculations using a five slab-layer Ni(110) and a p(3 × 3) unit cell. The calculations incorporated the Perdew–Burke–Ernzerhof (PBE) functional and the generalized gradient approximation (GGA) to calculate the exchange-correlation energy. The valence electron properties were modeled through the projector augmented-wave (PAW) approach, with an energy cutoff of 400 eV and a Monkhorst-Pack grid of (3 × 3 × 1) used for k-point sampling. To prevent interaction between adjacent slabs, a vacuum spacing of 15 Å in the z-direction was applied. The transition states of ammonia decomposition were determined using the dimmer method, with geometry optimization carried out using the force-based conjugated gradient method. Finally, the adsorption energy (Eads) was calculated as:2/N2. The reaction performance was evaluated using pure NH3 at 300–650 °C, at the gas hourly space velocity (GHSV) of 30,000 mL/(gcat.h). The residual NH3 in the effluent was absorbed in a 6 mol/L sulfuric acid solution. A mass flowmeter was used to monitor the flow rate of the outlet gas. The ammonia decomposition reaction was conducted in a homemade, lab-scale fixed bed reactor. A quartz tube  and a porous distributor were used to support the catalysts. An electric furnace was used to heat the reactor. The temperature of the reactor and the catalyst was controlled and monitored by two thermocouples to achieve the desired operating temperature. The details of the experimental setup used in this study are depicted in 3 conversion was calculated as follows: The NHSince ammonia is converted to N2 and H2 in the outlet gas, and a unit volume of ammonia is converted into two volumes of nitrogen and hydrogen mixture, the volume of ammonia converted is half the flow rate of the gas after the unconverted ammonia has been removed from the reactor outlet. That is, the converted ammonia flow rate was calculated as:Therefore, NH3 conversion can be calculated as follows:xCe1-xO2-δ for ammonia decomposition to obtain high-purity hydrogen. Ni/Gd0.2Ce0.8O2-δ shows almost full ammonia decomposition, leading to a high hydrogen production rate (2008.9 mmol.g−1.h−1) at 600 °C and 30,000 mL/(gcat.h). The performance persisted over 150 h long run and at variable temperatures of 550–600 °C. The mechanism investigations have manifested that Gd0.2Ce0.8O2-δ exhibits high oxygen vacancy concentration, highly dispersed Ni, and abundant strong active sites. These properties of Ni/Gd0.2Ce0.8O2-δ significantly improve the associative desorption of N and strengthen the metal–support interaction. Therefore, Ni/Gd0.2Ce0.8O2-δ shows satisfactory activity and stability at 600 °C.Tuning the support properties can enhance the activity and stability of Ni/ Gd


Emissions of volatile organic sulfur (S), selenium (Se),bromine(Br), and iodine (I) species from aquatic ecosystems represent animportant source of these elements into the atmosphere. Availablemethods to measure these species are either not sensitive enough ornot automated, which hinder a full understanding of species distributionand production mechanisms. Here, we present a sensitive and high-throughputmethod for the simultaneous and comprehensive quantification of S,Se, Br, and I volatile organic species in atmospheric and aqueoussamples using a preconcentration step onto sorbent tubes and subsequentanalysis by thermal desorption coupled to gas chromatography and inductivelycoupled plasma mass spectrometry (TD-GC-ICP-MS). Selected commerciallyavailable sorbent tubes, consisting of mixed porous polymer and graphitizedblack carbon, offered the highest trapping capacity and lowest lossof species when stored at −20 °C for 28 days after sampling.After optimization of the TD-GC-ICP-MS method, absolute detectionlimits were better than 3.8 pg, 9.1 fg, 313 fg, and 50 fg, respectively,for S, Se, Br, and I species. As a proof of concept, the concentrationsof target species were determined in aqueous and continuously collectedatmospheric samples during a cruise in the Baltic and North Seas.Moreover, unknown S, Br, and I volatile species were detected in bothaqueous and atmospheric samples demonstrating the full potential ofthe method. These emissions playa critical role in the global biogeochemical cycling of these elementsand have important environmental and health implications.8 For example, marine emissions of Br and I volatile species affectthe atmospheric oxidizing capacity through ozone depletion,9 whereas the oxidation products of DMS, the main volatile speciesin the sulfur cycle, contribute to the formation of cloud condensationnuclei and thus influence the Earth’s radiative budget.10Aquatic ecosystems, especiallymarine ones, are an important naturalsource of various volatile organic species to the atmosphere, notablythose containing sulfur (S), selenium (Se), and halogens such as bromine(Br) and iodine (I). Global marine emissions of S, Se, Br, and I areestimated at 17.6–34.4 Tg S y12 Volatile species of S and Se result frombiotic and abiotic degradation of metabolites,14 whereas those of I and Br arise from reactions between organic matterand reactive forms of I and Br16 or from phytoplanktonicand macroalgae activities.18 For S, Br, and I, respectively,the main marine volatile organic species are DMS, CHBr3, dibromomethane (CH2Br2), and iodomethane(CH3I).19 Average concentrationsof these species in the surface seawater are in the low picomolarto low nanomolar range, and their atmospheric mixing ratios are usuallyin the low part per trillion (pptv) range. Several minor species werealso reported for these elements in the low pM range, such as carbondisulfide (CS2), dibromochloromethane (CHBr2Cl), bromodichloromethane (CHBrCl2), iodo-ethane and iodo-propane(C2H5I and C3H7I), chloroiodomethane(CH2ICl), and diiodomethane (CH2I2).21 The concentrationof volatile Se species in surface (sea)waters are several orders ofmagnitude lower than the other elements, from the low fM to pM range,and have thus been scarcely analyzed.24 From these few measurementscarried out in estuarine or seawaters, dimethyl selenide (DMSe) wasthe main species detected, followed by dimethyl selenenyl sulfide(DMSeS) and dimethyl diselenide (DMDSe).Volatile organic species are producedthrough biological activityand/or photochemical processes occurring mainly in the upper watercolumn.3, CH2Br2, and CH3I, are usuallymeasured shipboard using purge and trap systems coupled to gas chromatography(GC) and specific detectors such as mass spectrometry (MS), flameionization detector (FID), or flame photometric detector (FDP).25 The advantage of using such online analyticalmethods is that they reduce the risk of potential losses and contaminationsthat might occur during sample storage and transport. However, thesetechniques present too high detection limits for the less concentratedvolatile species. For these latter, a prior preconcentration stepis required, which is usually achieved by cryogenic26 or sorbent trapping.27 Atmosphericsamples can also be collected in canisters, but the number of samplesis often limited due to the space requirement for their storage. Forthe analysis of volatile Se species in the environment, only cryogenictrapping, followed by gas chromatography and inductively coupled plasmamass spectrometry (GC-ICP-MS) analyses has been implemented to date.24 Although this method is sensitive, it remainsnonautomated and thus time consuming, and the necessity to bring liquidnitrogen in the field severely limits the number of samples that canbe collected and analyzed.22 Recently,promising results have been obtained for the preconcentration of volatileSe species onto multibed sorbent tubes and analysis by thermal desorption(TD) coupled to gas chromatography–mass spectrometry (GC-MS)28 as well as for the long-term storage of CHBr3 and CH2Br2 onto sorbent tubes29 or the quantification of DMS in coral incubationsusing a mix of porous material (Tenax TA) and molecular sieves (Sulficarb).30 However, the trapping of volatile organic speciesonto sorbents and their analysis has not been systematically investigatedso far for simultaneous volatile organic species present at environmentallyrelevant concentrations.The species presentin highest concentrations, i.e., DMS, CHBrIn this work, we present a new high-throughput,very sensitive,and multi-elemental method for the analysis of volatile species ofS, Se, Br, and I in atmospheric and aqueous samples using sorbenttrapping and analyses by an automated thermal desorption unit coupledto GC-ICP-MS (TD-GC-ICP-MS). We first fine-tuned the various desorptionsteps of the TD unit, GC program, and operating conditions of theICP-MS instrument to ensure optimal desorption, separation, and detectionof volatile species while keeping the analysis time within 15 minper sample. Then, we investigated several commercially available sorbenttubes for their retention and storage capacity as well as optimizedsampling and preconcentration for both dissolved gaseous species inaqueous samples using a purge and trap system and atmospheric samplesusing a commercially available automated autosampler. Our new methodallows for the simultaneous quantification of targeted species overa wide range of environmentally relevant concentrations, i.e., fromthe fM to nM range, as well as the detection of unknown volatile specieswith absolute detection limits (ADLs) ranging from the femtogram topicogram level depending on the element. Finally, we measured atmosphericand aqueous samples collected from the Baltic and North Seas, coveringvarious environmental conditions, to demonstrate the potential ofthis method.22.1124Xe (100 ppm) was purchased from Linde (Switzerland), whileargon, helium, hydrogen and nitrogen were purchased from PanGas AG(Switzerland). The volatile organic standards were purchased fromSigma-Aldrich (Switzerland) and VWR (Switzerland). Indications onboth gas and standards’ purity are given in Table S1. As no DMSeS standard is commercially available,it was quantified using the average of the calibration slopes of DMDSeand DMSe. Individual stock solutions (3000 mg·L–1) were prepared in amber glass vials by diluting the pure standardsin methanol  using gas-tight syringes . These stock solutions were sealed withPTFE-septum caps , covered with an aluminumfoil, and stored at −20 °C. As losses of volatile speciesare difficult to prevent and control, these solutions were regularlyreplaced, every 2 months at the latest. Fresh individual or mixeddiluted solutions were prepared daily in MeOH from the stock solutions.Then, 1 μL of these fresh solutions was loaded onto sorbenttubes via an injection loop  within a N2 flow of 50 mL·min–1 for three seconds to allow the transfer of the analytes from theMeOH solution to the gas phase and their sorption onto the sorbentmaterials while avoiding breakthrough.A mixture of He/2.2Table S2 presents the operating parameters ofthe analytical setup after the optimization presented in the secondsection of 124Xe and was found to be within 5% over an analysis periodof 20 h. Further details on the TD unit operation and optimizationare given in SI Section A.The analytical setup consists ofa thermal desorption unit coupled to a gas chromatography system fitted with an HP-5 column, hyphenated with an inductively coupledplasma mass spectrometer . Tables S3 and S4. The (re)conditioning programs of the varioussorbent tubes are given in Table S5. Calibrationcurves were obtained by analyzing five sorbent tubes, all loaded with1 μL of a working solution prepared as described above. Eachsorbent tube was analyzed with a different split ratio applied tothe focusing trap (varying from 3.8:1 to 75:1) to control the amountof volatile species entering the GC column . Further details on the calibration procedure and the determinationof detection limits are given in SI Section B.Six typesof commercially available multiple bed sorbent tubes ,odor/sulfur (SF), biomonitoring (BM), material emissions (ME), airtoxic (AT), and graphitized carbon (GR), all packed in inert stainless-steeltubes), were purchased from Markes International Limited. Their compositionsin terms of the sorbent material, target range, affinity to water,part number, and properties are summarized in 2.3n = 31) and aqueoussamples , collectedduring a 7-day cruise on the R/V Svea organized by the Swedish Meteorologicaland Hydrological Institute (SMHI) in the North and Baltic Seas inSeptember 2020. A detailed description of sampling conditions andcorrections applied to the data is available in SI Sections C and D along with a sampling map . Briefly, an in-house purge and trap system  with four parallel purging lineswas assembled for the sampling of dissolved volatile species. An automatedactive sampling device , including a sampling pump withvariable flow rate  was usedto sample atmospheric volatile species directly onto BM sorbent tubeswith a 4 h resolution (5 L of air at a flow rate of 21 mL·min–1).The sampling and analytical methods were assessed under environmentalconditions using atmospheric (33.13.1.1Table S6). GR and AT were the least performingsorbent tubes with average trapping capacities of 34 ± 26 and56 ± 35%, respectively. UN (94 ± 9%) and SF (93 ± 10%)overall performed better but were not adequate for low boiling point(BP) species, particularly DMS and DMSe. ME and BM overall presentedthe best trapping capacities with average recoveries of 94 ±4 and 99 ± 1%, respectively. However, BM showed better trappingcapacities for the volatile species with a low boiling point, notablyfor DMS (100 ± 4%), DMSe (100 ± 8), CS2 (100± 1%), and CH3I (100 ± 5%).The trappingcapacity of the six sorbent tubes was first determined at a low amount, the average trappingrecovery increased from 34 ± 26 to 44 ± 26% for GR and from56 ± 35 to 64 ± 36% for AT, indicating that some irreversibleadsorption or degradation occurred with lower amounts of volatilespecies. Considering this problem and their overall low trapping capacity,GR and AT sorbent tubes were not further considered. Unlike the othersorbent tubes tested here, AT and GR do not contain the porous polymerTenax TA (Table S3), suggesting its importancefor trapping the targeted species. ME and BM sorbent tubes, combininga graphitized carbon material with Tenax TA, presented the best trappingcapacity for the tested volatile species.In a second step, the trapping capacity was evaluatedfor 100 timeshigher amounts, i.e., 5 ng of volatile species. A comparison with0.05 ng is presented in 3.1.2Figure S4 for ME and SF sorbent tubes. In addition, Figures S5 and SI-6 show species recoveries at 14, 21, and 28 daysfor ME and BM sorbent tubes stored at 4 and 25 °C. Based on preliminaryresults showing a large broadening of the CS2 peak thatimpacted the quantification of DMS (not shown), UN sorbent tubes werediscarded. The recoveries after storage at – 20 °C for28 days were generally good with average values of 96 ± 20, 102± 7, and 100 ± 9%, respectively for SF, ME, and BM tubes.The lowest recoveries at −20 °C for all of the sorbenttubes were found for the species with low boiling points, e.g., CH3I (82 ± 21%), DMS (80 ± 22%), and DMSe (87 ±2%). Similar results were obtained for storage at – 80 °Cfor species with high boiling points; however, DMS (91 ± 11%),DMSe (94 ± 2%), and CH3I (98 ± 5%) showed betterrecoveries at −80 °C, indicating eventual losses occurringat higher storage temperatures.Species recoveries after storage at −80, −20, 4, and25 °C for 28 days are presented in 2Cl (91 ±5%), CHBr3 (91 ± 4%), and CH2I2 (89 ± 4%) at 25 °C. Overall, it can be concluded thatME, BM, and SF sorbent tubes have better trapping capacities at lowstorage temperatures with −20 °C being sufficient forstorage over one month. Based on the trapping and storage resultsas well as their hydrophobic character, BM sorbent tubes were selectedfor our application. SF could be an interesting alternative for thevolatile compounds mentioned above when low storage temperature at−20 °C is not available.Species recoveries after 14days of storage at 4 and 25 °C were on average 87 ± 26 and84 ± 24 and 81 ± 26 and 80 ± 23%, respectively, forBM and ME sorbent tubes, but they were much lower after 28 days, between10 and 37% depending on the species for both storage temperatures.These low recoveries demonstrate the inability of ME and BM sorbenttubes to retain the selected volatile species for a longer periodat storage temperatures of 4 and 25 °C. However, SF sorbent tubesshowed high recoveries after 28 days for CHBr3.23.2.1–1); the results are presented in Figure S7. Regarding the tube desorption time , 1 min was not long enough as many species, i.e., DMDS,DMDSe, CHBrCl2, CHBr2Cl, CHBr3, CH3I, CH2ICl, and CH2I2 accountedfor 3.3 ± 0.3 to 29 ± 1% of the total amount detected afterthe second tube desorption. No significant difference was observedfor tube desorption times of 2 and 4 min. For the desorption temperature,less than 5% of the initial amount loaded was detected after the seconddesorption regardless of the temperature applied . Therefore, 200 °C was chosen to preventexcessive TD maintenance resulting from too high desorption temperature.The efficiency of species desorption from the BM sorbent tubeswas evaluated for different times (1–4 min), temperatures (150–225°C), and gas flow rates  did not result in significantlydifferent recoveries of the volatile species. Only CH3Ishowed lower recoveries with increasing N2 flow rates.The loss of CH3I can be explained by a breakthrough effecton the focusing trap as CH3I is not well retained by theSF sorbent tube as demonstrated above. To prevent potential breakthroughthat might occur once the volatile species are transferred to thefocusing trap, we suggest using a low N2 flow rate. Basedon these results, we finally selected a tube desorption time of 2min at 200 °C and a N2 flow rate of 25 mL·min–1 (Different NmL·min–1 1 as thes3.2.2–1) wasrequired to avoid peak tailing and (ii) the residual peak areas ofeach species found after a second desorption of the focusing trapwere maximum 0.6% of the first peak areas regardless of the desorptiontime . To remove water vaporthat would potentially come from the sorption tubes and thus preservethe GC column, we also tested the effect of the initial temperatureof the focusing trap within the range from −30 to −5°C. The recoveries of the volatile species were found betterwhen the initial temperature of the focusing trap was higher , suggesting that lower temperatureshinder the adsorption of species to the sorbents present in the focusingtrap. The most important variations were observed when the initialtemperature of the focusing trap was set at −30 vs −20°C for DMS (85 ± 1 vs 92 ± 8%), CS2 (71± 15 vs 77 ± 7%), CHBrCl2 (93 ± 4 vs 90± 1%), and (96 ± 1 vs 76 ± 16%). At −30 °C,the average recovery for the tested volatile species was 86 ±15%, whereas better recoveries were observed at −10 and −5°C with 97 ± 10 and 98 ± 10%, respectively. Therefore,−5 °C was selected as the initial temperature for thefocusing trap, followed by a 3 min desorption at a heating rate of≥24 °C·min–1 (Preliminary tests showed that (i) the highest heating rate of thefocusing trap . However, significantbreakthrough was observed for CH3I, DMS, and CS2 . The breakthrough for CH3I varied from 57 ± 18 to 2 ± 1% and from 33 ±16 to 21 ± 21% for 5, 10, and 15 L, in the laboratory and inthe urban area, respectively. The decrease in breakthrough with increasingsampling volume demonstrates the low retention capacity of the BMsorbent for CH3I as soon as a higher sampling volume isapplied compared to the sorbent selection test (2 (<10 mL). A third BM sorbent tube connected in seriesat 15 L confirmed the presence of CH3I in the third tube(data not shown). The breakthrough increased from 2 ± 3 to 91± 2% and from 4 ± 2 to 31 ± 17% when the N2 volume increased from 5 to 15 L, respectively, for DMS and CS2. In the urban area, the breakthrough varied from 9 ±9 to 36 ± 24% and from 9 ± 11 to 37 ± 14%, with samplingvolumes from 5 to 15 L, for DMS and CS2, respectively.Similar results were obtained when sorbent tubes were flushed withurban air of varying humidity, suggesting that matrix effects arelimited. However, these conditions certainly do not represent allpossible sampling conditions, and future users would have to determinebreakthroughs for their specific applications. To both maximize thepreconcentration of species while minimizing DMS, CH3I,and CS2 losses from BM sorbent tubes, 5 L was selectedas the best compromise for a safe sampling of most of the target volatilespecies.For most of the species, the breakthrough remained negligible forall Nion test 3.1 where3.3.2Figure S9 as a function of the volumeof N2 (flow rate of 400 mL·min–1). It is important to note that the same results were obtained regardlessof the N2 flow rate applied  for the sameN2 volume demonstrating that the flow rate does not influencethe PT recovery and breakthrough. Moreover, the recovery was alsosimilar when artificial seawater (Milli-Q containing NaCl) or onlyMilli-Q was used . For specieswith a boiling point (BP) > 100 °C, i.e., CH2ICl,DMDS, CHBr2Cl, DMSeS, CHBr3, DMDSe, and CH2I2, recoveries increased with increasing N2 volume and reached between 57 ± 3% (CH2I2) and 100 ± 6% (CHBr3) at 25 L of N2, while the breakthrough remained negligible (<3.1%). For DMSand CS2, the recovery of the PT system was however maximalat 10 L N2 with 61 ± 2 and 35 ± 2%, respectively,while the breakthrough was limited to 11 ± 0.4 and 14 ±1%, respectively. The DMSe recovery also reached its maximum (28 ±2%) at 10 L, while the breakthrough was still very low (0.8 ±0.1%) pointing to a degradation of DMSe, likely by oxidation. Despitea rather low DMSe recovery in general, the low standard deviation(SD) values indicated a high reproducibility of DMSe trapping. A maximumrecovery of 14 ± 2% was observed for CH3I at 2 L ofN2. High breakthroughs were observed for CH3I regardless of the N2 volume (up to 68 ± 7% wasfound in the second tube). It was even detected in a third tube (datanot shown) demonstrating again the inability of the BM sorbent tubesto trap this species. When considering all of these results, 10 Lof N2 was selected as the best compromise between the recoveryfor most species and the breakthrough observed for DMS and CS2.Recoveries and breakthroughs of the PT system forthe various volatile species trapped onto BM sorbent tubes are presentedin 3.43.4.1–1 and 0.2–0.4 ppqv in aqueousand atmospheric samples, respectively, due to a very low Se instrumentbackground as a result of the efficient removal of interferences byH2 gas and a good ionization by the ICP. It should be notedthat potential interferences between CHBr3 on DMD80Se and generally from Br compounds on Se ones can be easily overcomeusing 78Se. MDLs for Br and I volatile species in aqueoussamples were in the low fmol·L–1 range (0.3–3.6fmol·L–1), while they ranged from 0.7 to 8.8ppqv in atmospheric samples. Both for atmospheric or aqueous samples,our newly developed analytical method achieves detection limits atleast 1 order of magnitude lower compared to the best methods availablethat are tailored for only a few species. Moreover, it should be notedthat the sampling, transport, and analyses based on sorbent tubesoffer certain advantages over canister sampling due to their easydeployment and implementation, low space requirement, straightforwardreconditioning, and reuse of sorption tubes (> 100 reconditioningsteps).Typical chromatogramsfor our target species after the full optimization of the TD-GC-ICP-MSare shown in 3.4.2Figure S12 (overlaid with standards),and the distribution of the measured concentrations is shown in 3I trappedon the sorbent tubes was always quantifiable, large uncertaintieson CH3I concentrations remained because of low PT recovery(2.1 ± 1.5%). For future work aiming at CH3I quantification,fine-tuning would be required to address this problem. Only in fewatmospheric samples, DMDSe concentrations were below DLs. The averageconcentrations of dissolved species found in the Baltic and NorthSeas were 1.8 ± 2.9 nM DMS, 7.4 ± 21 pM CHBr3, 3.4 ± 6.4 pM CH2Br2, and 0.94 ±0.81 pM DMSe. These concentrations are in good agreement, though onthe lower side (likely due to the low productivity of the waters atthe time of the cruise), with values previously reported in the literaturefor the main volatile species in aqueous samples: from 1 to 7 nM DMS,6 to 42 pM CHBr3, 1 to 9 pM CH2Br2, and 0.14 to 4.71 pM DMSe.37 The averages of the mixing ratio in atmospheric samples were 14.5± 13.9 DMS pptv, 5.3 ± 4.6 CHBr3 pptv, 0.35 ±0.13 CH2Br2 pptv, and 0.20 ± 0.09 DMSepptv, which are within the range described in the literature: 3.0–261pptv levels for DMS, 1–10 CHBr3 pptv, and 0.5–13.2CH2Br2 pptv.38 It is worth noting that some of the concentrationsmeasured with our method were below or close to the DLs of other techniques,i.e., for DMSe, DMDSe, and CH2ICl in atmospheric samplesas well as DMDSe, CHBrCl2, CHBr2Cl, CHBr3, and CH2ICl in aqueous samples  as confirmed with a standard. AlthoughMeSH has been difficult to measure due to its reactivity, it has beendetected in marine bacteria39 and in seawater.40 A peak containing both Br and I was detectedat 5.4 min corresponding to a theoretical BP (th. BP) of 131 ±1 °C that would match to CH2BrI (BP 139 °C),which was previously reported as a minor species.7 Regarding I, various peaks that did not match the standardswere detected in atmospheric and especially in aqueous samples. Thepeaks at 3.4 and 4.4 min were later confirmed with standards to beC2H5I and C3H7I, respectively,two species that have been previously detected in seawater.41 The large peak observed at 6.5 min (th. BP 172 ± 1 °C)could not be attributed to any species, and further work is thus neededto identify remaining unknowns as well as their importance in othermarine waters. However, these proof-of-concept analyses clearly showedthe potential of the method for the simultaneous quantification oftargeted volatile species and screening of unknown ones.Another advantageof using ICP-MS compared to other commonly used detectors lies inthe opportunity to detect unknown species. We defined unknown specieswhen no standard matched the peaks detected on the chromatograms.The major and minor unknown species detected in both atmospheric andwater samples are summarized in 4A novel and highly sensitivemethod for the determination of 13targeted S, Se, Br, and I volatile organic species in atmosphericand aqueous samples was successfully developed. Volatile species fromaqueous samples were first collected on commercially available sorbenttubes using a custom-made purge and trap system. Volatile speciesin air were collected using a commercially available autosampler thatcan be easily deployed in the field. The preconcentrated/trapped specieswere separated and quantified within 15 min using a high-throughput,automated thermal desorption unit coupled to GC-ICP-MS. Our methodpresents the lowest MDLs to date in the literature for all targetspecies in both compartments. Volatile species are stable on sorbenttubes stored at −20 °C for at least 28 days, allowingextended field campaigns. The potential for environmental sample analysis,including the detection of unknown species, was demonstrated by analyzingsamples collected from the North and Baltic Seas. This new multi-elementalmethod represents a great opportunity for a better characterizationof the environmental distribution of trace S, Se, Br, and I volatileorganic species and their cycling in aquatic ecosystems and in theatmosphere.


However, the underlying mechanisms are largely unclear. The functions of CaV1.2 channels are subtly modulated by splicing factor-mediated alternative splicing (AS), but whether and how CaV1.2 channels are alternatively spliced in diabetic heart remains unknown.L-type CaV1.2 channel functions were measured by whole-cell patch clamp, and intracellular Ca2+ concentration was monitored by using Fluo-4 AM.Diabetic rat models were established by using high-fat diet in combination with low dose streptozotocin. Cardiac function and morphology were assessed by echocardiography and HE staining, respectively. Isolated neonatal rat ventricular myocytes (NRVMs) were used as a cell-based model. Cardiac CaV1.2 channel with alternative exon 9* (CaV1.2E9*), but unchanged that with alternative exon 8/8a or exon 33. The splicing factor Rbfox2 expression is also increased in diabetic heart, presumably because of dominate-negative (DN) isoform. Unexpectedly, high glucose cannot induce the aberrant expressions of CaV1.2 exon 9* and Rbfox2. But glycated serum (GS), the mimic of advanced glycation end-products (AGEs), upregulates CaV1.2E9* channels proportion and downregulates Rbfox2 expression in NRVMs. By whole-cell patch clamp, we find GS application hyperpolarizes the current-voltage curve and window currents of cardiac CaV1.2 channels. Moreover, GS treatment raises K+-triggered intracellular Ca2+ concentration ([Ca2+]i), enlarges cell surface area of NRVMs and induces hypertrophic genes transcription. Consistently, siRNA-mediated knockdown of Rbfox2 in NRVMs upregulates CaV1.2E9* channel, shifts CaV1.2 window currents to hyperpolarization, increases [Ca2+]i and induces cardiomyocyte hypertrophy.We find that diabetic rats develop diastolic dysfunction and cardiac hypertrophy accompanied by an increased CaV1.2E9* channels and hyperpolarizes channel window currents. These make the channels open at greater negative potentials and lead to increased [Ca2+]i in cardiomyocytes, and finally induce cardiomyocyte hypertrophy in diabetes. Our work elucidates the underlying mechanisms for CaV1.2 channel regulation in diabetic heart, and targeting Rbfox2 to reset the aberrantly spliced CaV1.2 channel might be a promising therapeutic approach in diabetes-induced cardiac hypertrophy.AGEs, not glucose, dysregulates Rbfox2 which thereby increases CaThe online version contains supplementary material available at 10.1186/s12933-023-01894-5. The levels of AGEs in blood serum and the lysate of cardiac tissue were measured by using an AGEs ELISA kit . H9c2 rat cardiac myoblast cells were obtained from American Type Culture Collection .All animal works have been approved by and performed in accordance with the Institutional Animal Care and Use Committee of Nanjing Medical University and conformed to the Guide for the Care and Use of Laboratory Animals . Adult male Sprague-Dawley (SD) rats were fed with standard diet or high-fat diet (HFD) and tap water ad libitum at constant ambient temperature (25 °C) and humidity. Diabetic rat model was established by intraperitoneal injection of streptozotocin  for 5 consecutive days and fed with HFD. Control rats received the same volume of citrate buffer  and fed with standard diet. Blood glucose levels were measured using a glucometer , and the body weight of the rats were measured every 2 weeks. For tissue collection, the rats were euthanized by overdose CO2, 1 L/min). Left ventricle wall thickness and chamber dimensions were determined in the parasternal short-axis view (M-mode), from which measures of left ventricle ejection fraction (LVEF) and fractional shortening (FS) were derived. 2D images of the heart were obtained from the 4-chamber apical view to assess mitral blood inflow and tissue Doppler velocities. All measurements were averaged over 6 successive cardiac cycles and performed by an echocardiographer who was blinded throughout the whole experiment.Transthoracic echocardiographic measurements were performed as described in the previous study  with VevFor preparation of glycated serum (GS), dialyzed-fetal bovine serum (FBS) was incubated under sterile conditions with D-glucose (9 g/mL) at 37 °C for 3 weeks , 31. TheActb mRNA was used to normalize the expression of target genes. The 2−ΔΔCT method was used to analyze the data. Sequences of the primers are listed in Table The total RNA was extracted from cells or tissues by using Trizol reagent (Invitrogen), following the manufacturer’s instructions, and then 1 µg of total RNA was reverse-transcribed to cDNA using HiScript III QRT Super-Mix reverse transcriptase . RT-PCR was carried out using Rapid Taq Master Mix (Vazyme). 2% agarose gel was used to separate the PCR products. Quantitative real-time PCR (qPCR) was performed on Step-One Plus real-time PCR System . Reactions were performed in 96-well plates with 20 µL volume including 100 ng cDNA, 200 nmol/L of each primer and 10 µL AceQ qPCR SYBR Green Master Mix (Vazyme). NRVMs were extracted from SD rats of 1 to 2-days old by using Pierce Primary Cardiomyocyte Isolation Kit , which contains enzyme #1 (with papain) and enzyme #2 (with thermolysin). Neonatal hearts were placed into separate 1.5 mL centrifuge tubes with 300 µL HBSS. Cardiac tissues were then cut into small pieces with a curved scissors inside a sterile centrifuge tube. The minced tissue was then digested by adding 0.2 mL of enzyme #1 and 10 µL enzyme #2. Next, the tubes were incubated in a shaker under 37 °C for 30–35 min. The mixture was then transferred into another dish with DMEM medium (Gibco) containing 10% FBS (Gibco) and 1% penicillin/streptomycin (Sigma). NRVMs or H9c2 cells were transfected with siRNAs targeting Rbfox2 mRNA Table  with LipMembrane protein extraction was performed on ice. The heart tissue or cells were homogenized in the lysis buffer  added with protease inhibitor and dithiothreitol. Then vortex mixing was done for 30 s and place on the ice for 1 min, repeated for 5 cycles. The lysate was centrifuged at 12,000 rpm  for 10 min. The pellets were collected and homogenized in an extraction buffer solution. Then vortex mixed for 30 s, and place on the ice for 5 min, 5 cycles. The lysate was centrifuged at 12,000 rpm for 10 min again, and the supernatant was collected as the membrane protein.Total protein was extracted from cells or left ventricular tissues. The whole operation was carried out on ice. The collected cells or the ground tissues were lysed with RIPA lysis buffer, which added protease inhibitors. Lysates were centrifuged for 15 min at 12,000 rpm under 4℃. The supernatants were used for Western blotting after determining its protein concentration by Bradford assay . The protein was separated on 10% SDS-PAGE and transferred to PVDF membranes. After blocking, PVDF membranes were incubated with different primary antibodies overnight at 4℃ or 2 h at 25℃.V1.2 α1C , Na-K ATPase , RBM20 . The secondary antibodies used were HRP-conjugated goat anti-mouse  or anti-rabbit IgG , or rabbit anti-goat IgG . We used an imaging system  to visualize the blot with the enhanced chemiluminescence reagent .Primary antibodies against targeted proteins were used: β-actin , Rbfox2 , CaCSA of isolated NRVMs was determined by immunofluorescence staining. Briefly, the NRVMs were fixed in 4% paraformaldehyde for 15 min, permeabilized with 0.1% Triton X-100 in PBS for 5 min, then blocked with 0.3% BSA in PBS for 40 min at 25℃ and incubated with primary antibody α-actinin  overnight at 4℃. After washing with PBS 3 times for 5 min, the NRVMs were incubated with secondary antibody labeled with Alexa Fluor 488  for 1 h at 25℃. Then, DAPI was used to label the nuclei. The fluorescence signaling was captured by a confocal laser scanning microscope . Image J software was used to calculate surface area of the cells.2+]i was measured using Fluo-4 AM  as previously described i was calculated by the equation: Δ[Ca2+]i = ΔF/F0 = (F − F0)/F0, where F is the fluorescence intensity of NRVMs at any given time and F0 is the basal fluorescence intensity prior to an experimental manipulation./k), where Gmax is the maximum conductance, V is the testing potential, Erev is the reversal potential, V0.5 is half-activation potential, and k is slope rate. Steady-state activation (SSA) curve was derived from I–V protocol, and calculated by the equation of G = I / (V – Erev), where G is conductance, and the curve was fitted with the Boltzmann equation i elevation, our data showed that GS-induced hyperpolarized CaV1.2 channel kinetics at least partly raises K+-induced [Ca2+]i elevation in NRVMs.Since GS treatment hyperpolarized Cator Fig. A. ExtracVMs Fig. B. ThoughSERCA2a) , mediateNppa mRNA was slightly increased, and the levels of Nppb and Myh7 mRNA were obviously increased in GS-treated cells compared to NG-treated ones  CaV1.2 channels with alternative exon 9*, but not those with exon 8a or exon 33, are specifically increased in the heart from diabetic rats; (2) AGEs, not glucose, induces aberrant expression of Rbfox2, and upregulates CaV1.2E9* channels in cardiomyocyte; (3) AGEs/Rbfox2-mediated exon 9* insertion hyperpolarizes cardiac CaV1.2 window currents; and (4) AGEs application or knockdown of Rbfox2 increases [Ca2+]i, enlarges CSA and triggers the hypertrophic genes transcription in cardiomyocytes, which mediates cardiomyocyte hypertrophy (Fig. Camyocytes . Imbalan2+ handling, aberrant splicing of CaV1.2 channel is presented in different heart diseases, like myocardial infraction i, which leads to cardiac contraction [2+]i will initialize genes transcription in cardiomyocyte by excitation-transcription coupling i [2+]i in cardiomyocytes, which we thought is at least partly attributed to Rbfox2-mediated CaV1.2 AS. In the heart, increased Ca2+ influx through CaV1.2 channel raised fetal genes transcription, leading to cardiomyocyte hypertrophy i . This reertrophy , 38. HerV1.2 channel activity by LTCC blockers might have benefits in cardiac remodeling against cardiomyopathies in the murine model [V1.3 calcium channels which are also expressed in the heart. Second, CaV1.2 channels are widely expressed in different organs or systems [V1.2 channels on the other tissues including blood vessels and the brain. Therefore, targeting CaV1.2 splice isoform which specifically expressed in diseased heart might be a promising approach to manage cardiomyopathies [V1.2E9* channels produced enhanced channel functioning in diabetic hearts, we think by targeting CaV1.2 exon 9* or its regulatory factor Rbfox2 is possibly an optimal idea to manage DCM, which requires further investigations.Though the inhibition of enhanced Cane model , 48, thene model . We thinne model , 51, suc systems , therefoopathies . Here, iV1.2 channel is aberrantly spliced in diabetic heart with a specific manner, which enhances the channel function, and this effect was mediated by AGEs-induced Rbfox2 downregulation under diabetic hyperglycemia. Therefore, targeting Rbfox2 to reset the aberrantly spliced CaV1.2 channel might be a promising therapeutic approach in diabetes-induced cardiac hypertrophy.From the new dimension, we identified for the first time that CaBelow is the link to the electronic supplementary material.Supplementary Material 1


Electrochemical H2O2 generation has been studied in the past, but a lack of studies exists on miniaturized electrochemical platforms for the on-demand synthesis of H2O2 for antibacterial applications. In this article, a chemically modified cotton textile platform capable of in situ H2O2 production is demonstrated for E. coli deactivation. The cotton textile was modified by layer-by-layer coating with conductive carbon nanotubes/cellulose nanocrystals (CNT/CNC) and a polymer of polyaniline (PANI) decorated with anthraquinone (AQ), designated as the AQ@PANI@CNT/CNC@textile antibacterial patch. The AQ@PANI@CNT/CNC@textile antibacterial textile patch H2O2 production capabilities were evaluated using both electrochemical and colorimetric methods. The AQ@PANI@CNT/CNC@textile antibacterial patch electrochemically produced H2O2 concentrations up to 209 ± 25 µM over a 40 min period and displayed a log reduction of 3.32 for E. coli over a period of 2 h. The AQ@PANI@CNT/CNC@textile antibacterial patch offers promise for use as a self-disinfecting pathogen control platform.Hydrogen peroxide (H For example, some studies have examined the usage of H2O2 for the sterilization and reuse of personal protective equipment (PPE) to address global shortages and for environmental conservation motivations [2O2 cooling aerosol sprays were found to be highly effective in reducing the viral coronavirus load on used face masks [, the development of new methods for on-demand H2O2 production is an important area of research.Hydrogen peroxide -based process [2O2 as a byproduct, which is then distilled and concentrated [2O2 for integration into PPE [2O2 production have been explored. For instance, electrochemical technologies have utilized metal catalysts, such as platinum, to produce H2O2 [2 electrode capable of converting H2O into H2O2 at a rate of 0.29 µmol·L−1·cm−2·h−1, with a faradaic efficiency of 8% [2 into H2O2 [2O2 was generated at a rate of 1.80 mol·g−1·hr−1, with a faradaic efficiency of 95.8%, making their platform a promising new technology [The standard method of H process . In thisentrated ,11. Howento H2O2 . Utilizichnology .2O2 on the cathode when immersed in an electrochemical cell [− oxygen reduction reaction (ORR) transition, shown in Equations (1) and (2) [Both PANI and AQ are capable of generating Hcal cell ,13. When and (2) ,13., which becomes protonated to form H2O2  assembly of a cotton textile substrate integrated with layers of conductive carbon nanotube (CNT)/cellulose nanocrystal (CNC) composite and PANI decorated with AQ (AQ@PANI@CNT/CNC@textile). The AQ@PANI@CNT/CNC@textile platform has been successfully tested for electrochemical production of H2O2 for E. coli deactivation. The H2O2 production capabilities of the AQ@PANI@CNT/CNC@textile antibacterial patch were quantified using cyclic voltammetry (CV) and colorimetric methods. The H2O2-generating AQ@PANI@CNT/CNC@textile holds promise for potential integration into masks or other wearable devices for in situ disinfection.While effective, metal electrocatalysts are susceptible to anodic degradation and may require regeneration between uses . FurtherE. coli K-12 (Z-), tryptic soy agar, saline solution, and barium chloride were procured from Sigma Aldrich, Oakville, ON, Canada. Sulfuric acid, 30% H2O2, aniline, dipotassium phosphate, and monopotassium phosphate were purchased from Fisher Scientific, Denver, CO, USA. Carboxylic acid-functionalized multiwalled carbon nanotubes (CNT) (OD: 4–6 nm. 98%) were bought from TimesNano, China. Cellulose nanocrystals (CNC) were donated by Alberta Innovates, Canada. Stainless steel hypodermic needles  and textile bandages were bought from a local pharmacy in Edmonton, Alberta. Copper tape was purchased from 3M . All reagents were of analytical reagent grade. All aqueous solutions were prepared using 18.2 MΩ deionized (DI) Milli-Q water.Ammonium peroxydisulfate (APS), 3-(trimethoxysilyl)propyl methacrylate (TPM), methanol, silver nanoparticles, 2,6-diaminoanthraquinone, sodium hypochlorite, dimethylsulfoxide (DMSO), LB broth with agar (Lennox), potassium ferricyanide, sodium acetate trihydrate, 4-aminoantipyrine, phenol, acetic acid, horseradish peroxidase, 2O2 detection and the CNT/CNC/Ag (Ag/AgCl) reference needle were fabricated as described previously [2SO4 and 30% H2O2 in a 1:1 (v/v) ratio for 4 h, then washed with DI water and air dried. The oxidized needles were then silylated via immersion in a solution comprising 3-(trimethoxysilyl) propyl methacrylate (TPM)/DI water/methanol (2:1:8 v/v/v) for 4 hrs, then washed with DI water and air dried. To fabricate the CNT/CNC sensor electrode, the silylated needles were infused with a homogenous suspension comprising 1 mg·mL−1 CNT/4 mg·mL−1 CNC at a flow rate of 15 µL·min−1 at 80 °C. Homogenous CNT/CNC suspensions were first prepared by sonicating 1 mg·mL−1 CNT/4 mg·mL−1 CNC in DI water for 3 h [−1 CNT/4 mg·mL−1 CNC/5 mg·mL−1 Ag nanoparticle suspension (in DI water) at a flow rate of 15 µL·min−1. The needles were then immersed in 2 mL of sodium hypochlorite bleach overnight at room temperature and rinsed with DI water.The CNT/CNC sensor electrode for Heviously ,17. Brie−1 CNT/4 mg·mL−1 CNC suspension and dried on a hot plate at 50 °C. The PANI microparticles were produced by mixing 50 mL of 0.2 M aniline (in 1 M H2SO4) and 50 mL of 0.25 M APS initiator in an ice bath at 4 °C. The resulting PANI microparticle suspension was then centrifuged  and rinsed with DI water three times before being dried in a dehydrator at 70 °C. A 20 mg·mL−1 solution comprising dried PANI microparticles (in DI water) was sonicated for 3 h to create a homogenous suspension. A total of 8 mL of the 20 mg·mL−1 PANI suspension was drop cast onto the CNT/CNC@textile patch and air dried. Lastly, the PANI@CNT/CNC@textile patch was cut into 4 × 4 cm squares. To integrate AQ into the PANI@CNT/CNC@textile patches, 1.5 mL of 0.5 M 2,6-diaminoanthraquinone (in DMSO) was drop cast onto the patch surface and air dried to yield the AQ@PANI@CNT/CNC@textile antibacterial patches. The AQ was similarly drop cast onto the CNT/CNC@textile patches to yield AQ@CNT/CNC@textile patches. After air drying, the patches were gently rinsed in a beaker of DI water to remove any excess AQ. A 0.3 × 1.0 cm strip of copper tape was wrapped around one corner of each patch to serve as an electrical connection.To fabricate the antibacterial patch, an 8 × 8 cm cotton textile obtained from a local store was drop cast with 8 mL of a 1 mg·mLFourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), electrochemical impedance spectroscopy (EIS), voltammetric, and Raman spectroscopy techniques were used to characterize the AQ@PANI@CNT/CNC@textile antibacterial patch. FTIR spectra were recorded using a Bruker Tensor 27 FTIR instrument fitted with diamond attenuated total reflectance (ATR). SEM images were acquired from a Zeiss Sigma 300 VP field emission SEM. EIS was performed by immersing different patches in 25 mM potassium ferricyanide (in 0.1 M KCl) as a standard redox probe in a three-electrode electrochemical cell, with an in-house fabricated Ag/AgCl reference needle, and a platinum counter electrode. EIS data were acquired in the frequency range of 20.0 Hz–1 MHz at 6.00 mV of sinusoidal amplitude. CV was used to determine the electroactive surface area of the different patches. The CV was performed by immersing each patch in 25 mM potassium ferricyanide (in 0.1 M KCl) in a three-electrode electrochemical cell, using an Ag/AgCl reference needle and a platinum counter electrode. A Palmsens 4 potentiostat equipped with PSTrace software was used for all electrochemical measurements.2O2, antibacterial patches were immersed in 10 mL of 0.1 M phosphate buffer (pH 7.0) electrolyte in a three-electrode cell comprising a platinum counter electrode and an Ag/AgCl reference needle for CV application .To electrochemically generate H2O2 was electrochemically detected using CV  acquired from a CNT/CNC needle sensor running separately within the H2O2 generation cell. The H2O2 detection cell comprised a three-electrode system, with a platinum wire and Ag/AgCl reference needle acting as the counter and reference electrodes, respectively. Faradaic capacitance was determined from the CVs acquired using the CNT/CNC needle sensor and was calculated by averaging the cathodic current within the − 0.50 to 0 V voltage range and dividing it by the scan rate. The ∆capacitance was determined by subtracting the capacitance generated from the blank from that generated from the sample and dividing it by capacitance of the blank.The H2O2 generation. Briefly, the colorimetric reagent was prepared by combining 0.3681 g of sodium acetate trihydrate, 0.0102 g of 4-aminoantipyrine, 0.0047 g of phenol, 29.4 µL of glacial acetic acid, and D.I water to make 50 mL of solution. In a cuvette, 2 mL of the colorimetric reagent was mixed with 10 µL of 1.1 mg/mL horseradish peroxidase dissolved in D.I water, 500 µL of electrolyte, and brought to a volume of 3 mL using 490 µL of DI water [A colorimetric method was also used to detect HDI water . The absE. coli. E. coli K-12 (Z-) cells were streaked on tryptic soy agar (TSA) and incubated at 37 °C for 24 h. Single E. coli colonies were suspended in 0.85% (w/v) sterile saline, and the turbidity was adjusted to match a 0.5 McFarland standard, resulting in a bacterial suspension of approximately 108 cells·mL−1. The bacterial suspension was then diluted by combining 2.5 mL suspension with 2.5 mL of sterile saline. To incorporate E. coli into the antibacterial patch network for the deactivation test, 500 µL of dilute bacterial suspension was drop-casted onto the patch surface. CV  was subsequently performed via immersion in 10 mL of 0.1 M phosphate buffer under the same conditions used for electrochemical H2O2 generation. A total of 10 µL of phosphate buffer was drawn at various points in time , suspended in 990 µL of sterile water, and then serially diluted in a 1:9 ratio five times. The final diluted suspension was then cultured on LB agar and incubated at 37 °C for 16 h. Bacterial growth was visually evaluated by counting the number of colonies formed on the LB agar plate after the 16 h incubation period.Different antibacterial patch designs were tested for their ability to deactivate E. coli integration were examined using SEM. The CNT/CNC@textile patch, shown in E. coli cells, respectively. AQ is visualized as tiny crystals coated on the surface of the CNT/CNC fiber network, while E. coli cells appear as rounded cylinders  is desirable for high patch conductivity and effective signal transduction for H2O2 generation [ct values determined from the circuit fittings  as the standard redox probe. The Nyquist plots from EIS analysis and associated circuit fittings are shown in neration . The Rctfittings  for the −1) and invoking the Randles–Sevcik equation (The electroactive surface areas of the different patches were determined by running CV of the redox couple 25 mM potassium ferricyanide (in 0.1 M KCl) at different scan rates  an2O2 generated from various chemically modified patches. In the H2O2 generation cell, the patches acted as the working electrode for CV application in phosphate buffer. For the H2O2 detection system, a CNT/CNC sensor electrode was used as the working electrode for CV acquisition in the same electrochemical cell using a different potentiostat [2O2 are shown in Two three-electrode cells were utilized to produce and quantify the Hntiostat . Represe2O2, a calibration curve in the range of 0–400 µM H2O2 was used   . H2O2 prectively a. Additiectively a.2O2 production [2O2 was indicated by the absorbance peak at 505 nm, and this detection method had a calibration sensitivity of 4.53 × 10−4 µM−1 (2O2 generation from AQ@PANI@CNT/CNC@textile and PANI@CNT/CNC@textile patches are shown in 2O2 at a rate of 1.01 µM·min−1 (2O2 production occurred very slowly in the first 16 min before increasing linearly at a rate of 9.54 µM·min−1. The total concentration of H2O2 after 32 min of CV application was determined to be 36.8 and 143 µM for PANI@CNT/CNC@textile and AQ@PANI@CNT/CNC@textile patches, respectively (2O2 production (A colorimetric assay was used as an additional method to quantify Hoduction . The pre0−4 µM−1 , with a µM·min−1 b. For thectively b. Both qoduction . However2O2 is generated from O2 via a 2e− ORR assisted by the AQ@PANI@CNT/CNC@textile antibacterial patch [2O2 production for the AQ@PANI@CNT/CNC@textile antibacterial patches relative to the PANI@CNT/CNC@textile can be attributed to the enhanced catalytic activity imparted by the AQ layer [2O2 compared to other methods (2O2 generation.Mechanistically, Hal patch . AQ actsal patch . TherefoAQ layer . General methods . HoweverE. coli bacteria was tested. CV was performed on E. coli-integrated CNT/CNC@textile, PANI@CNT/CNC@textile, and AQ@PANI@CNT/CNC@textile patches immersed in phosphate buffer, and samples taken from the electrochemical cells at various times were grown on LB agar plates and counted (E. coli colonies formed from it at any point during CV application (E. coli-integrated AQ@PANI@CNT/CNC@textile antibacterial patch, and the log reduction was 3.32, whereas that of the PANI@CNT/CNC patch was less than 0.500 (E. coli bacteria over 2 h. This suggests that the AQ@PANI@CNT/CNC@textile patch may be useful for integration into face coverings or masks for enhanced microbe deactivation and sanitization [The ability of the AQ@PANI@CNT/CNC@textile antibacterial patch to deactivate  counted . The CNTlication . Antibaclication , calculaan 0.500 . As suchtization .2O2 in amounts up to 209 ± 25 µM. Although this concentration of H2O2 is lower than that reported in other studies conducted using PANI and AQ electrocatalysts [E. coli after 2 h of electrochemical H2O2 generation. These results are indicative of strong antibacterial activity, and the textile-based antibacterial patch has great potential to be integrated into PPE or face coverings for enhanced protection against pathogens. Overall, this study outlines a promising textile-based technology that may be further optimized for controlling various pathogens in food-safety- and health-related applications.The AQ@PANI@CNT/CNC@textile antibacterial patch described in this study produced Hatalysts , bacteri