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Fetzara Lake, considered one of the most important wetlands in northeastern Algeria, was designated a Ramsar site in 2002. The waters in its watershed are affected by salinity, which influences their suitability for irrigation. To identify the factors influencing the quality of these surface waters, geochemical and statistical analyses were carried out on the basis of the results of chemical analyses of 51 samples collected, during two monitoring campaigns, from all the tributaries in the watershed. The findings show the dominance of three hydrochemical facies over the two campaigns: Na-Cl facies (55.17% and 22.73%) characterizes the waters water from Fetzara Lake outlet (drainage channel and wadi Meboudja), in relation to the influx of saliferous elements due to water evaporation in the lake. Ca-Mg-Cl (27.59% and 40.91%) and Ca-Mg-HCO3 (13.79%. and 13.79%) facies characterize the waters of the remaining tributaries, reflecting the dissolution of carbonate formations and the alteration of the Edough metamorphic basement. Multivariate statistical analysis, using principal component analysis (PCA), shows three water types: highly mineralized (EC > 3000 µS/cm), moderately mineralized (1000 < EC < 3000 µS/cm), and weakly mineralized (EC < 1000 µS/cm). Evaporation and silicate weathering are the main mechanisms controlling water mineralization according to the different bivariate plots. Furthermore, cation exchange indices (CAI-I and CAI-II) reveal that these reactions involve the adsorption of Na+ and K+ onto clay minerals, as well as the simultaneous release of Ca2+ and Mg2+ ions. Finally, the various quality indices (SAR, %Na, RSC and KR) revealed that the water in 36% of tributaries is unsuitable for irrigation. These findings will provide important information on surface water quality in the study area, particularly for irrigation purposes, and will contribute to the thoughtful and sustainable management of this resource.

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Human urine, a highly saline solution rich in plant-available nutrients, leaves behind significant organic matter after nutrient recovery, necessitating additional treatment for environmental protection. While nutrient recovery from human urine is well-documented in the literature, research on the safe handling of the residual liquid phase is notably lacking. This study investigates nutrient recovery from source-separated human urine using clinoptilolite for the ion exchange/adsorption process and evaluates the safe management of the residual liquid through anaerobic granular sludge and a second-stage of sorption. The results indicated that the ion exchange/adsorption process, using an ammonium loading of 15 mg NH4+/g clinoptilolite, removed the majority of nutrients, achieving 82% ammonium removal and 100% phosphorus removal, along with 30% removal of organic matter. The residual liquid phase from the nutrient removal stage was treated separately with anaerobic digestion and a second-stage of sorption for further processing. Results showed that anaerobic processing removed 68%-84% of organic matter, with no additional nitrogen removal observed as expected, and produced 0.20-0.46 L CH4/L urine. The second-stage of sorption removed 59%-62% of organic matter and nearly all nitrogen. Both processes effectively removed organic matter, with sorption also eliminating nitrogen and anaerobic processing potentially generating biogas, making them recommended for improving the quality of the residual liquid phase before final disposal.

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Large synthetic oligonucleotides such as guide ribonucleic acid (gRNA), a critical reagent in clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 genome editing, have complex higher order structures (HOS) inherent in their design. In this study, we first developed a generic anion exchange chromatography (AEX) method for the comprehensive analysis of a 100mer single guide ribonucleic acid (sgRNA) impurity profiling. AEX demonstrated superior resolution compared to other common chromatographic methods employed for sgRNA analysis, such as Ion-Pairing Reversed Phase Liquid Chromatography (IP-RPLC) and Hydrophilic Interaction Chromatography (HILIC). Moreover, we discovered AEX's potential in probing the HOS of RNAs by adjusting the temperature and using organic additives. Our study also highlighted that sgRNA possesses a unique HOS distinctly different from other therapeutic nucleic acids, such as antisense oligonucleotides and messenger RNAs.

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Alluvial fans and deltas are two environments with different hydrochemical conditions. Their junction zones, as mixing environments, are variably influenced by different processes, leading to variable environmental conditions. The purpose of this study is to investigate groundwater quality in the junction zone of these environments in the northern part of the Jazmourian depression (known as the Rudbar plain) in southeastern Iran to determine the dominant processes, assess arsenic and fluoride health risks, and evaluate irrigation water quality. A total of 33 samples from deep drilled wells were taken, and the concentrations of major ions and elements were determined. Additionally, statistical and hydrochemical analyses were undertaken. The dominant processes in the delta are evaporation and ion exchange, while the dominant process in the fan environment is silicate hydrolysis. Among the samples, 26.7% were mainly affected by the delta, and 73.3% were mainly affected by fan conditions. Although the majority of groundwater samples were suitable for irrigation based on quality standards, a significant portion exceeded the acceptable level for Na%. Non-carcinogenic health risk assessments indicated that arsenic hazard risks exceeded thresholds in 63.3% of cases for children and 36% for adults. Carcinogenic health risks associated with arsenic and fluoride exceeded acceptable levels in 4 and 2 stations, respectively. Elevated As concentrations contribute to a greater average health risk in parts of fans environment.

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Membrane chromatography devices are a viable alternative to packed-bed resins and enable highly productive purification cascades for monoclonal antibodies and Fc-fusion proteins. In this study, ion exchange and protein A membrane chromatography performances were assessed and compared with their resin counterparts. Protein A dynamic binding capacities were higher than 50 g/L for two of the tested membranes and with a residence time of 0.2 min. For polishing, it was observed that aggregate clearance was generally less performant with membrane separation when compared to resins with similar ligands. However, the comparable yield and increased productivity of membranes could be enough to consider their implementation. In addition, lifetime studies demonstrated that the performance of membranes remained robust over cycles. One hundred cycles were reached for most of the tested membranes with no impact on the process performance nor product quality. Finally, purification cascades were fully operated with membranes, from capture to polishing, reaching good levels of host cells proteins (less than 50 ppm) and aggregates (equal to or less than 1%). The outcome of this study demonstrated that resin chromatography could be fully replaced by membranes for monoclonal antibody and Fc-fusion protein purification processes.

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The electrochemical reduction of CO2 (ERCO2) has emerged as one of the most promising methods for achieving both renewable energy storage and CO2 recovery. However, achieving both high selectivity and stability of catalysts remains a significant challenge. To address this challenge, this study investigated the selective synthesis of formate via ERCO2 at the interface of In2O3 and Bi2O3 in the InBiO6 composite material. Moreover, InBiO6 was synthesized using indium-based metal-organic frameworks as precursor, which underwent continuous processing through ion exchange and thermal reduction. The results revealed that the formate Faradaic efficiency (FEformate) of InBiO6 reached nearly 100 % at -0.86 V vs. reversible hydrogen electrode (RHE) and remained above 90 % after continuous 317-h electrolysis, which exceeded those of previously reported indium-based catalysts. Additionally, the InBiO6 composite material exhibited an FEformate exceeding 80 % across a wide potential range of 500 mV from -0.76 to -1.26 V vs. RHE. Density-functional theory analysis confirmed that the heterogeneous interface of InBiO6 played a role in achieving optimal free energies for *OCHO on its surface. Furthermore, the addition of Bi to the InBiO6 matrix facilitated electron transfer and altered the electronic structure of In2O3, thereby enhancing the adsorption, decomposition, and formate production of *OCHO.

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Advancing cathode materials is crucial for the broader application of aqueous zinc-ion batteries (ZIBs) in energy storage systems. This study presents amorphous H/VO4 (HVO), a novel cathode material engineered by substituting H+ for Mg2+ in Mg2VO4 (MgVO), designed to enhance performance of ZIBs. Initial exploration of MgVO through ab initio molecular dynamics (AIMD) simulations and density functional theory (DFT) calculations revealed a favorable Mg2+ and Zn2+ exchange mechanism. This mechanism notably reduces electrostatic interactions and facilitates ion diffusion within the host lattice. Building upon these findings, in this work, theoretical calculations analysis indicated that amorphous HVO offers a higher diffusion coefficient for Zn2+ ions and fewer electrostatic interactions compared to its crystalline MgVO precursor. Subsequent empirical validation is achieved by synthesizing amorphous HVO using a rapid ion-exchange process, effectively replacing Mg2+ with H+ ions. The synthesized amorphous HVO demonstrated 100% capacity retention after 18000 cycles at a current density of 2 A g-1 and exhibited exceptional rate performance. These findings underscore the significant potential of HVO cathodes to enhance the durability and efficiency of aqueous ZIBs, positioning them as promising candidates for future energy storage technologies.

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BACKGROUND: The integration of positron emission tomography (PET) and magnetic resonance imaging (MRI) holds promise for advancing diagnostic imaging capabilities. The METRICS project aims to develop cyclotron-driven production of 52Mn for PET/MRI imaging. RESULTS: Using the 52Cr(p,n)52Mn reaction, we designed chromium metal targets via Spark Plasma Sintering and developed a separation procedure for isolating 52Mn. Labeling tests were conducted with traditional chelators (i.e. S-2-(4-Isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane tetraacetic acid) and the 1.4-dioxa-8-azaspiro[4.5]decane-8- carbodithioate ligand to produce radioactive complexes suitable for PET/MRI applications. Our methodology yielded high-quality 52Mn suitable for PET radiopharmaceuticals and PET/MRI imaging. Preliminary studies on phantom imaging using microPET and clinical MRI demonstrated the efficacy of our approach. CONCLUSIONS: The developed technology offers a promising avenue for producing 52Mn and enhancing PET/MRI imaging capabilities. Further in vivo investigations are warranted to evaluate the potential advantages of this hybrid imaging technique.

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This study examines the critical interaction between seasonal precipitation variability and forest maturity in determining ion deposition patterns in rehabilitated forest ecosystems. This research was conducted in rehabilitated forest sites in Bintulu, Sarawak, Malaysia that had ecologically similar plant distribution, species, and age in each planting area. This facilitated the standardization of rainfall deposition in the different study plots which streamlined the study of these specific facets of ecosystem dynamics. The goal is to understand how seasonal changes and the age of the forest influence the chemical composition of the flux that relates to the movement and deposition of nutrients through the forest ecosystem. This flux is a key factor in the health of the forest ecosystem and nutrient cycling. Using ion exchange resin (IER) samplers, we accurately measured and compared the deposition of different ions (Ca2+, Na+, Fe2+, Cu2+, NO3 -, NH4 + and SO4 2-) across different seasons and forest ages. The deposition of Ca2+ and NH4+ was significantly lower in the low-precipitation season than in the high-precipitation season in all forest stands, regardless of the year they were established (1996, 1999, 2002, 2005, and 2009). In contrast, ions such as Na+, Fe2+, Cu2+, NO3 - and SO4 2- showed no clear seasonal fluctuations. In addition, the study shows that through-fall in forest stands from 2002, 2005 and 2009 had higher concentrations of Ca2+ in both seasons than in 1996 and 1999. Interestingly, forest stands from 2009 and 2002 had elevated levels of Na+ and SO42- in seasons with low precipitation, while stands from 1996 had higher levels in seasons with high precipitation. Our results emphasize the crucial role of precipitation amount and canopy age in determining ion deposition in forest ecosystems. By demonstrating the significant influence of precipitation seasonality and forest maturity on the chemical composition of throughfall, this study contributes to a deeper understanding of nutrient dynamics in developing forest landscapes and provides valuable insights for ecological restoration measures.

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Nylon 54 is a novel, biodegradable polyamide with excellent thermal resistance and water absorption properties. It can be polymerized using bio-based cadaverine and succinic acid as monomers. Traditional separation methods isolate individual monomers from the fermentation broth through acidification or alkalization, resulting in significant amounts of waste salts; however, synchronous separation of dibasic acids and diamines has not been reported. This study investigated an integrated process for the separation and extraction of nylon 54 salts from a co-fermentation broth without acidification or alkalization. We meticulously optimized the operational parameters of the integrated process to achieve maximum separation efficiency. Following microfiltration, ultrafiltration, and decolorization, the bacterial eliminating rate was ≥99.83%, and the protein concentration was ≤40 mg/L. The absorbance of the decolorized solution was ≤0.021 at 430 nm, and the recovery rate of nylon 54 salt reached 97%. Then, the pretreated solution was passed through sequential chromatographic columns, which effectively removed organic acid by-products (such as acetic acid and lactic acid), SO4 2-, and NH4 + from the fermentation broth, resulting in a cadaverine yield of 98.01% and a succinic acid yield of 89.35%. Finally, by concentrating and crystallizing the eluent, the simulated fermentation broth yielded nylon 54 salt with a purity of 99.16% and a recovery rate of 58%, and the real fermentation broth yielded nylon 54 salt with a purity of 98.10% and a recovery rate of 56.21%. This integrated process offers a sustainable and environmentally friendly pathway for the complete biosynthesis of nylon 54 salt and has the potential to be extended to the preparation of other nylon salts.

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Bacterial extracellular vesicles (BEVs) have extraordinary biotechnological potential, but traditional purification methods lack desirable scalability and commonly co-isolate protein impurities, limiting clinical translation. Anion exchange chromatography (AEC) separates molecules based on differences in net charge and is widely used for industrial biomanufacturing of protein therapeutics. Recently, AEC has recently been applied for purification of EVs from both mammalian and bacterial sources. Since most bacteria produce BEVs with a negative surface membrane change, AEC can potentially be widely used for BEV purification. Here, we describe a method utilizing high-performance AEC (HPAEC) in tandem with size-based tangential flow filtration for improved BEV purification. We have previously found this method can reduce co-isolated protein impurities and potentiate anti-inflammatory bioactivity of probiotic BEVs. Thus, this method holds promise as a scalable alternative for improved BEV purification.

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Extracellular vesicles (EVs) are relatively recently discovered biological nanoparticles that mediate intercellular communication. The development of new methods for the isolation and characterization of EVs is crucial to support further studies on these small and structurally heterogenous vesicles. New scalable production methods are also needed to meet the needs of future therapeutic applications. A reliable inline detection method for the EV manufacturing process is needed to ensure reproducibility and to identify any possible variations in real time. Here, we demonstrate the use of an inline Raman detector in conjunction with anion exchange chromatography for the isolation of EVs from human platelets. Anion-exchange chromatography can be easily coupled with multiple inline detectors and provides an alternative to size-based methods for separating EVs from similar-sized impurities, such as lipoprotein particles. Raman spectroscopy enabled us to identify functional groups in EV samples and trace EVs and impurities in different stages of the process. Our results show a notable separation of impurities from the EVs during anion-exchange chromatography and demonstrate the power of inline Raman spectroscopy. Compared to conventional EV analysis methods, the inline Raman approach does not require hands-on work and can provide detailed, real-time information about the sample and the purification process.

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This paper employs a high-throughput parallel batch (microtiter plate) adsorption screen with sequential salt step increases to rapidly generate protein elution profiles for multiple resins at different pHs using a protein library. The chromatographic set used in this work includes single mode, multimodal anion-exchange (MMA), and multimodal cation-exchange (MMC) resins. The protein library consists of proteins with isoelectric points ranging from 5.1 to 11.4 with varying hydrophobicities as determined by their retention on hydrophobic interaction chromatography. The batch sequential experiments are carried out using one protein at a time with a wide set of resins at multiple pH conditions, thus enabling simple microtiter plate detection. A mathematical formulation is then used to determine the first moment of the distributions from each chromatogram (sequential step elution) generated in the parallel batch experiments. Batch data first moments (expressed in salt concentration) are then compared to results obtained from column linear salt gradient elution, and the techniques are shown to be consistent. In addition, first moment data are used to calculate one-resin separability scores, which are a measure of a resin's ability, at a specified pH, to separate the entire set of proteins in the library from one another. Again, the results from the batch and column experiments are shown to be comparable. The first moment data sets were then employed to calculate the two-resin separability scores, which are a measure of the ability of two resins to synergistically separate the entire set of proteins in the library. Importantly, these results based on the two-resin separability performances derived from the batch and column experiments were again shown to be consistent. This approach for rapidly screening large numbers of chromatographic resins and mobile phase conditions for their elution behavior may prove useful for enabling the rapid discovery of new chromatographic ligands and resins.

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The management of produced water (PW) generated during oil and gas operations requires effective treatment and comprehensive chemical and toxicological assessment to reduce the environmental risks associated with reuse or discharge. This study evaluated a treatment train that included a low-temperature thermal distillation pilot system followed by granular activated carbon (GAC) and zeolite post-treatment for processing hypersaline Permian Basin PW. Our study provides a unique and comprehensive assessment of the treatment efficiency considering a targeted chemical scheme together with whole effluent toxicity (WET) tests across four trophic levels regarding aquatic critical receptors of concern (ROC): Raphidocelis subcapitata, Vibrio fischeri, Ceriodaphnia dubia, and Danio rerio. The distillate from the thermal distillation process met various numeric discharge standards for salinity and major ions. However, it did not meet toxicity requirements established by the United States National Pollutant Discharge Elimination System program. Subsequent post-treatment using GAC and zeolite reduced the concentration of potential stressors, including volatile organics, NH3, Cd, Cr, Zn, and Mn in the final effluent to below detection limits. This resulted in a consistent toxicity reduction across all WET tests, with no observable adverse effects for R. subcapitata, C. dubia, and D. rerio (no observed effect concentration >100%), and V. fischeri effects reduced to 19%. This study realizes the feasibility of treating PW to non-toxic levels and meeting reuse and discharge requirements. It underscores the importance of implementing integrated treatment trains to remove the contaminants of concern and provides a systematic decision framework to predict and monitor environmental risks associated with PW reuse.

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A design procedure for the separation of charge variants of a monoclonal antibody (mAb) was developed, which was based on the coupling of cation-exchange chromatography (CEX) and anion-exchange chromatography (AEX) under high loading conditions. The design of the coupled process was supported by a dynamic model. The model was calibrated on the basis of band profiles of variants determined experimentally for the mAb materials of different variant compositions. The numerical simulations were used to select the coupling configuration and the loading conditions that allowed for efficient separation of the mAb materials into three products enriched with each individual variant: the acidic (av), main (mv) and basic (bv) one. In the CEX section, a two-step pH gradient was used to split the loaded mass of mAb into a weakly bound fraction enriched with av and mv, and a strongly bound fraction containing the bv-rich product. The weakly bound fraction was further processed in the AEX section, where the mv-rich product was eluted in flowthrough, while the av-rich product was collected by a step change in pH. The choice of flow distribution and the number of columns in the CEX and AEX sections depended on the variant composition of the mAb material. For the selected configurations, the optimized mAb loading density in the CEX columns ranged from 10 to 26 mg mL-1, while in the AEX columns it was as high as 300 or 600 mg mL-1, depending on the variant composition of the mAb material. By proper selection of the loading condition, a trade-off between yield and purity of the products could be reached.

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Research on the recovery of rare earth elements from wastewater has attracted increasing attention. Compared with other methods, biosorption is a simple, efficient, and environmentally friendly method for rare earth wastewater treatment, which has greater prospects for development. The objective of this study was to investigate the biosorption behavior and mechanism of Yarrowia lipolytica for five rare earth ions (La3⁺, Nd3⁺, Er3⁺, Y3⁺, and Sm3⁺) with a particular focus on biosorption behavior, biosorption kinetics, and biosorption isotherm. It was demonstrated that the biosorption capacity of Y. lipolytica at optimal conditions was 76.80 mg/g. It was discovered that the biosorption process complied with the pseudo-second-order kinetic model and the Langmuir biosorption isotherm, indicating that Y. lipolytica employed a monolayer chemical biosorption process to biosorb rare earth ions. Characterization analysis demonstrated that the primary functional groups involved in rare earth ion biosorption were amino, carboxyl, and hydroxyl groups. The cooperative biosorption of rare earth ions by Y. lipolytica was facilitated by means of surface complexation, ion exchange, and electrostatic interactions. These findings suggest that Y. lipolytica has the potential to be an effective biosorbent for the removal of rare earth elements from wastewater.

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The large-sized perovskite single-crystal sheet (SCS) serves as the ideal research platform for perovskite photodetectors due to its outstanding carrier photophysics, pronounced geometric aspect ratio, and ultrahigh material utilization rate. However, its performance in flexible device applications is relatively lackluster due to the rigid and brittle nature of the three-dimensional cubic lattices. In this work, the indium tin oxide (ITO)-based multimillimeter-sized MAPbBr3 SCS is transformed into MAPbI3 SCS via ion exchange strategy. Significantly, we proposed and implemented a polymer-controlled transfer strategy─utilizing the dichloromethane (DCM) solution of poly(methyl methacrylate) (PMMA)─to nondestructively transfer the whole perovskite SCS off the ITO substrates and subsequently adhere it onto a flexible polyethylene terephthalate (PET) substrate of interdigital electrode, thereby fabricating a lateral-structured photodetector with a PMMA-SCS-Au-PET multilayer configuration. The tight self-encapsulation between the top PMMA membrane and the bottom PET substrate imparts excellent waterproof stability and concurrently excellent mechanical flexibility to these devices; additionally, the MAPbI3 device exhibits comprehensively superior performance to the MAPbBr3 one. This work represents a proactive attempt and exploration of the high-performance advancement of large-sized SCS photodetectors, undoubtedly introducing novel momentum and solutions to this domain.

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Improved measurement and analysis technologies are needed for investigating nanoparticle generation characteristics in sewage treatment plants. Single-particle inductively coupled plasma-mass spectrometry (spICP-MS) can be used to analyze metal nanoparticle characteristics. However, during spICP-MS analysis of environmental samples, high concentrations of ionic materials obscure the signals of particulate materials by increasing background signals. This can increase the threshold value for separating background and particle signals and increase the background-equivalent diameter (BED). In this study, particle size distributions in influent and effluent collected from sewage treatment plants were investigated using an improved spICP-MS method combining spICP-MS with ion-exchange resin (IER) column pretreatment. The ion removal effect of the IER column was first examined using a synthetic mixture of Ag nanoparticles (AgNPs) and ions. The method was then applied to wastewater from six different sewage treatment plants using an optimal IER packing of 5 g. The ion removal efficiency for samples containing a proper mixture of AgNPs and Ag ions was 99.98%, and the BED significantly decreased from 73.0 ± 1.0 to 6.1 ± 0.3 nm. Particle size distributions measured in the treatment plant influent and effluent ranged from 28.5 nm (Co) to 220.3 nm (Mg) and from 26.8 nm (Co) to 291.8 nm (Mg), respectively. spICP-MS/IER enabled the detection of smaller particles by removing ions from the sample and significantly decreasing the size detection limit. The results of this study offer a reference for developing predictive models for removing metal nanoparticles during sewage/wastewater treatment.

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Ion exchange chromatography-tandem mass spectrometry (IEC-MS/MS) has recently become the preferred method for detecting ionic substances in tea. In this study, an IEC-MS/MS method was developed for the rapid determination of chlorate and perchlorate residues in tea samples. The optimal sample extraction process, pretreatment column, and chromatographic and mass spectrometric conditions were systematically investigated. In the optimal process, the tea samples were ultrasonically extracted with methanol-water (13∶7, v/v), and a PRiME HLB SPE column was used to purify the sample extract. An AceChrom Hybri-A IEC column (150 mm×2.1 mm, 5.0 µm) was used for separation, and 100 mmol/L ammonium acetate-acetonitrile (40∶60, v/v) was used as the mobile phase for isocratic elution. The flow rate was 0.3 mL/min, the column temperature was 40 ℃, and the injection volume was 5.0 µL. The mass spectrometric data were collected in negative electrospray ionization mode combined with multiple reaction monitoring (MRM) mode to achieve the rapid and accurate separation and qualitative analysis of the desired chemical components. Quantification was performed using the internal standard (IS) method. The measurement results showed a good linear relationship when the mass concentrations of chlorate and perchlorate were between 2.00-200 and 1.00-100 µg/L, respectively, with correlation coefficients (r2) greater than 0.9990. The average recoveries of chlorate and perchlorate at three spiked levels of low, medium, and high ranged from 88.54% to 97.25% with relative standard deviations (RSDs, n=7) of 3.2%-5.2%. The limits of detection for chlorate and perchlorate were 12.0 and 8.0 µg/kg, respectively, while the limits of quantification were 40.0 and 26.6 µg/kg, respectively. The results of tests conducted to assess the linearity, specificity, accuracy, precision, and applicability of the method to the analysis of chlorate and perchlorate in 15 tea samples collected from a local market demonstrated its validity for the routine analysis of tea samples. The proposed method is simple, rapid, sensitive, and accurate, and can meet requirements for the rapid screening and quantitative analysis of residual trace chlorate and perchlorate in large quantities of tea samples.

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Charge variants are one of the most important quality attributes for protein therapeutics, including antibody drug conjugates (ADCs). ADCs are conjugation products between monoclonal antibodies (mAbs) and highly potent payloads. After attaching a payload, the charge profile of a mAb can be modified due to the change in net charge or surface charge. In this study, we present a unique challenge of charge assay development that arises from a desirable engineering of ADCs that incorporates the hydrolysis-prone succinimide-thioether conjugation chemistry. This engineered hydrolysis at conjugation sites is usually not complete during conjugation process and continuously progressing during mild stress. This hydrolysis also creates a carboxylic functional group, which manifests as acidic peaks in the ADC charge profiles. As a result, ion exchange chromatograms become sensitive measurements of this hydrolysis, which often masks the charge profile change due to other important post-translational modifications. In this study, two approaches were explored to address this unique challenge: to remove the hydrolysis heterogeneity by incubating ADCs under high pH conditions to drive complete hydrolysis; and to analyze charge variants at the subunit level after IdeS digestion. Acceptable charge profiles and quantitative integration results were successfully obtained by both approaches.