Revealing spatial distribution and accessibility of cell wall polymers in bamboo through chemical imaging and mild chemical treatments.

Understanding the distribution and accessibility of polymers within plant cell walls is crucial for addressing biomass recalcitrance in lignocellulosic materials. In this work, Imaging Fourier Transform Infrared (FTIR) and Raman spectroscopy, coupled with targeted chemical treatments, were employed to investigate cell wall polymer distribution in two bamboo species at both tissue and cell wall levels. Tissue-level Imaging FTIR revealed significant disparities in the distribution and chemical activity of cell wall polymers between the fibrous sheath and fibrous strand. At the cell wall level, Imaging Raman spectroscopy delineated a distinct difference between the secondary wall and intercellular layer, with the latter containing higher levels of lignin, hydroxycinnamic acid (HCA), and xylan, and lower cellulose. Mild acidified sodium chlorite treatment led to partial removal of lignin, HCA, and xylan from the intercellular layer, albeit to a lesser extent than alkaline treatment, indicating susceptibility of these polymers to chemical treatment. In contrast, lignin in the secondary wall exhibited limited reactivity to acidified sodium chlorite but was slightly removed by alkaline treatment, suggesting stable chemical properties with slight alkaline intolerance. These findings provide valuable insights into the inherent design mechanism of plant cells and their efficient utilization.

Addition of sodium malonate alters the morphology and increases the critical flux during tangential flow filtration of precipitated immunoglobulins.

Recent studies have demonstrated that one can control the packing density, and in turn the filterability, of protein precipitates by changing the pH and buffer composition of the precipitating solution to increase the structure/order within the precipitate. The objective of this study was to examine the effect of sodium malonate, which is known to enhance protein crystallizability, on the morphology of immunoglobulin precipitates formed using a combination of ZnCl2 and polyethylene glycol. The addition of sodium malonate significantly stabilized the precipitate particles as shown by an increase in melting temperature, as determined by differential scanning calorimetry, and an increase in the enthalpy of interaction, as determined by isothermal titration calorimetry. The sodium malonate also increased the selectivity of the precipitation, significantly reducing the coprecipitation of DNA from a clarified cell culture fluid. The resulting precipitate had a greater packing density and improved filterability, enabling continuous tangential flow filtration with minimal membrane fouling relative to precipitates formed under otherwise identical conditions but in the absence of sodium malonate. These results provide important insights into strategies for controlling precipitate morphology to enhance the performance of precipitation-filtration processes for the purification of therapeutic proteins.

Solutes unmask differences in clustering versus phase separation of FET proteins.

Phase separation and percolation contribute to phase transitions of multivalent macromolecules. Contributions of percolation are evident through the viscoelasticity of condensates and through the formation of heterogeneous distributions of nano- and mesoscale pre-percolation clusters in sub-saturated solutions. Here, we show that clusters formed in sub-saturated solutions of FET (FUS-EWSR1-TAF15) proteins are affected differently by glutamate versus chloride. These differences on the nanoscale, gleaned using a suite of methods deployed across a wide range of protein concentrations, are prevalent and can be unmasked even though the driving forces for phase separation remain unchanged in glutamate versus chloride. Strikingly, differences in anion-mediated interactions that drive clustering saturate on the micron-scale. Beyond this length scale the system separates into coexisting phases. Overall, we find that sequence-encoded interactions, mediated by solution components, make synergistic and distinct contributions to the formation of pre-percolation clusters in sub-saturated solutions, and to the driving forces for phase separation.

High-valent metal-oxo species transformation and regulation by co-existing chloride: Reaction pathways and impacts on the generation of chlorinated by-products.

High-valent metal-oxo species (HMOS) have been extensively recognized in advanced oxidation processes (AOPs) owing to their high selectivity and high chemical utilization efficiency. However, the interactions between HMOS and halide ions in sewage wastewater are complicated, leading to ongoing debates on the intrinsic reactive species and impacts on remediation. Herein, we prepared three typical HMOS, including Fe(IV), Mn(V)-nitrilotriacetic acid complex (Mn(V)NTA) and Co(IV) through peroxymonosulfate (PMS) activation and comparatively studied their interactions with Cl- to reveal different reactive chlorine species (RCS) and the effects of HMOS types on RCS generation pathways. Our results show that the presence of Cl- alters the cleavage behavior of the peroxide OO bond in PMS and prohibits the generation of Fe(IV), spontaneously promoting SO4•- production and its subsequent transformation to secondary radicals like Cl• and Cl2•-. The generation and oxidation capacity of Mn(V)NTA was scarcely influenced by Cl-, while Cl- would substantially consume Co(IV) and promote HOCl generation through an oxygen-transfer reaction, evidenced by density functional theory (DFT) and deuterium oxide solvent exchange experiment. The two-electron-transfer standard redox potentials of Fe(IV), Mn(V)NTA and Co(IV) were calculated as 2.43, 2.55 and 2.85 V, respectively. Due to the different reactive species and pathways in the presence of Cl-, the amounts of chlorinated by-products followed the order of Co(II)/PMS > Fe(II)/PMS > Mn(II)NTA/PMS. Thus, this work renovates the knowledge of halide chemistry in HMOS-based systems and sheds light on the impact on the treatment of salinity-containing wastewater.

Freezing-enhanced degradation of azo dyes in the chloride-peroxymonosulfate system.

In this study, we investigated the freezing-induced acceleration of dye bleaching by chloride-activated peroxymonosulfate (PMS). It has been observed that the oxidation of chloride by PMS generates a free chlorine species, such as hypochlorous acid (HOCl), under mild acidic and circumneutral pH condition. This process is the major reason for the enhanced oxidation capacity for electron-rich organic compounds (e.g., phenol) in the chloride-PMS system. However, we demonstrated that the chloride-PMS system clearly reduced the total organic carbon concentration (TOC), whereas the HOCl system did not lead to decrease in TOC. Overall, the chemical reaction is negligible in an aqueous condition if the concentrations of reagents are low, and freezing the solution accelerates the degradation of dye pollutants remarkably. Most notably, the pseudo-first order kinetic rate constant for acid orange 7 (AO7) degradation is approximately 0.252 h-1 with 0.5 mM PMS, 1 mM NaCl, initial pH 3, and a freezing temperature of -20 °C. AO7 degradation is not observed when the solution is not frozen. According to a confocal Raman-microscope analysis and an experiment that used an extremely high dose of reactants, the freeze concentration effect is the main reason for the acceleration phenomenon. Because the freezing phenomenon is spontaneous at high latitudes and at mid-latitudes in winter, and the chloride is ubiquitous elsewhere, the frozen chloride-PMS system has potential as a method for energy-free and eco-friendly technology for the degradation of organic pollutants in cold environments.

The application of chitosan quaternary ammonium salt to replace polymeric aluminum ferric chloride for sewage sludge dewatering.

Inorganic coagulants such as poly aluminum ferric chloride (Al/Fe) are applied conventionally to sewage sludge dewatering and can be retained in the sludge cake, causing its conductivity to increase and generate secondary pollution. To reduce these disadvantages, there is a need to develop alternative, more sustainable chemicals as substitutes for conventional inorganic coagulants. In the present investigation, the application of a polymeric chitosan quaternary ammonium salt (CQAS) is explored as a complete, or partial, replacement for Al/Fe in the context of sludge dewatering processes. Laboratory experiments using digested sewage sludge showed that CQAS could effectively substitute for over 80 % of the Al/Fe inorganic coagulant in the sludge dewatering process. This substitution resulted in a reduction of sludge cake conductivity by more than 50 %. Simulation of sludge dewatering curves and imaging of the sludge surface indicated that the addition of CQAS led to an increase in nanosized pores, and a decrease in the specific resistance of the sludge filter cake as the dosage of Al/Fe decreased to around 30 %. The variations of fluorescence emission, quantum yield and carboxylic and amino groups, suggested that the chelating of Al/Fe decreased due to the bridging effects of CQAS. The CQAS had different flocculation bridging effects on various EPS fractions, which varied the amount of protein chelated with Al/Fe in each fraction. This study provides new information about the benefits of replacing conventional inorganic coagulants with natural organic polymers for sewage sludge dewatering, in terms of reduced sludge cake conductivity and greater dry solids content.

A novel approach for the facile synthesis of zinc oxide/carbon hybrid systems from corn distillers soluble: Surface modification and characterization for sustainable remediation.

Sustainable, efficient, and environmentally friendly ways to tailor the carbonaceous materials from bio sources with desired functionalities remain a challenge around the world. In this study, we represent a novel approach to synthesize carbon hybrid material based on Zinc Oxide/carbon (ZnO/C) hybrid systems by catalytic hydrothermal process via crosslinking reaction through nucleation and growth of ZnO particles at the functional groups of oxidized carbon material. This research explored the volarization of Condensed Corn Distillers Soluble (CDS) as a carbon precursor to synthesize biobased carbon spheres. Surface modification of the produced carbon spheres took place using zinc chloride (ZnCl2) during hydrothermal carbonization (HTC). Zinc chloride (ZnCl2) was used to function as a catalyst during HTC and functioned as a ZnO source to synthesize (ZnO/C) hybrid systems. Design Expert software v13 was used to design the hydrothermal carbonization (HTC) experiments and response surface methodology was used to find the optimized conditions for the preparation of carbon hybrid systems. The hydrothermal synthesis process introduced 3D stone like zinc oxide particles onto the carbon matrix. These particles were self-assembled onto the carbon framework to produce carbon hybrid systems with unique physical, chemical, structural and functional properties. Herein, the obtained carbon hybrid systems (ZnO/C) were investigated and discussed in detail. ZnO/C hybrid systems were analyzed for surface morphology using scanning electron microscopy (SEM) that presented a 3D spherical interconnected phase and XRD analyses were used for phase crystallinity that showed new crystalline phases such as hopeite and zincite after the ZnCl2 incorporation. Surface functional groups were also analyzed by FTIR and results confirmed the presence of hydrophilic groups such as -OH, CC, and COOH on the surface of ZnO/C hybrid carbon systems. This study provided the insightful guidance for tailoring novel design of multifunctional carbon material as an adsorbent/catalyst for various applications of sustainable remediation.

Suitability of inorganic coagulants for algae-laden water treatment: Trade-off between algae removal and cell viability, aggregate properties and coagulant residue.

Inorganic coagulants could effectively precipitate algae cells but might increase the potential risks of cell damage and coagulant residue. This study was conducted to critically investigate the suitability of polyaluminum (PAC), FeCl3 and TiCl4 for algae-laden water treatment in terms of the trade-off between algal substance removal, cell viability, and coagulant residue. The results showed that an appropriate increase in coagulant dosage contributed to better coagulation performance but severe cell damage and a higher risk of intracellular organic matter (IOM) release. TiCl4 was the most destructive, resulting in 60.85% of the algal cells presenting membrane damage after coagulation. Intense hydrolysis reaction of Ti salts was favorable for the formation of larger and more elongated, dendritic structured flocs than Al and Fe coagulants. TiCl4 exhibited the lowest residue level and remained in the effluents mainly in colloidal form. The study also identified charge neutralization, chemisorption, enmeshment, and complexation as the dominant mechanisms for algae water coagulation by metal coagulants. Overall, this study provides the trade-off analyses between maximizing algae substance removal and minimizing potential damage to cell integrity and is practically valuable to develop the most suitable and feasible technique for algae-laden water treatment.

Efficient extraction of lignin from moso bamboo by microwave-assisted ternary deep eutectic solvent pretreatment for enhanced enzymatic hydrolysis.

Applications of deep eutectic solvent (DES) systems to separate lignocellulosic components are of interest to develop environmentally friendly processes and achieve efficient utilization of biomass. To enhance the performance of a binary neutral DES (glycerol:guanidine hydrochloride), various Lewis acids (e.g., AlCl3·6H2O, FeCl3·6H2O, etc.) were introduced to synthesize a series of ternary DES systems; these were coupled with microwave heating and applied to moso bamboo. Among the ternary DES systems evaluated, the FeCl3-based DES effectively removed lignin (81.17%) and xylan (85.42%), significantly improving enzymatic digestibility of the residual glucan and xylan (90.15% and 99.51%, respectively). Furthermore, 50.74% of the lignin, with high purity and a well-preserved structure, was recovered. A recyclability experiment showed that the pretreatment performance of the FeCl3-based DES was still basically maintained after five cycles. Overall, the microwave-assisted ternary DES pretreatment approach proposed in this study appears to be a promising option for sustainable biorefinery operations.

Direct Synthesis of Glycosyl Chlorides from Thioglycosides.

Reported herein is a new method for the direct synthesis of glycosyl chlorides from thioglycosides using sulfuryl chloride at rt. A variety of thioglycosides and thioimidates could be used as substrates. Both acid- and base-sensitive protecting groups were found compatible with these reaction conditions. Preliminary investigation of the reaction mechanism indicates chlorination of the leaving group at the anomeric sulfur as the key step of the reaction.

Utility of Triethyloxonium Tetrafluoroborate for Chloride Removal during Sarcosine N-Carboxyanhydride Synthesis: Improving NCA Purity.

The clinical translation of polysarcosine (pSar) as polyethylene glycol (PEG) replacement in the development of novel nanomedicines creates a broad demand of polymeric material in high-quality making high-purity sarcosine N-carboxyanhydride (Sar-NCA) as monomer for its production inevitable. Within this report, we present the use of triethyloxonium tetrafluoroborate in Sar-NCA synthesis with focus on amino acid and chloride impurities to avoid the sublimation of Sar-NCAs. With a view towards upscaling into kilogram or ton scale, a new methodology of monomer purification is introduced by utilizing the Meerwein's Salt triethyloxonium tetrafluoroborate to remove chloride impurities by covalent binding and converting chloride ions into volatile products within a single step. The novel straightforward technique enables access to monomers with significantly reduced chloride content (<100 ppm) compared to Sar-NCA derived by synthesis or sublimation. The derived monomers enable the controlled-living polymerization in DMF and provide access to pSar polymers with Poisson-like molecular weight distribution within a high range of chain lengths (Xn 25-200). In conclusion, the reported method can be easily applied to Sar-NCA synthesis or purification of commercially available pSar-NCAs and eases access to well-defined hetero-telechelic pSar polymers.

A composite gel formed by konjac glucomannan together with Nano-CF obtained by FeCl3-citric acid hydrolysis as a potential fat substitute.

This study aims to fabricate composite gels using nano citrus fiber (Nano-CF) derived from the hydrolysis process of citric acid (CA) with FeCl3, with a simultaneous exploration of its potential as an substitute to fats. Investigation of varying FeCl3 concentrations (0.01 to 0.03 mmol/g of CA) revealed a significant enhancement in the water-holding and oil-retention capacity of the Nano-CF. The meticulous synthesis of the composite gels involved integrating nano citrus fibers with konjac glucomannan (KGM) through high-speed shearing, followed by a comprehensive evaluation of its microstructure and physicochemical attributes. Increasing the Nano-CF concentration within the gels led to a synergistic interaction with KGM, resulting in enhanced viscosity, improved thermal stability, and restricted water molecule mobility within the system. The gels initially displayed reduced firmness, resilience, and adhesive characteristics, followed by subsequent improvement. When the ratio of Nano-CF to KGM was 0.5:1, the composite gels exhibited texture parameters, viscosity, and viscoelastic stability comparable to whipped animal cream formulations. These findings provide a new idea for the application of Nano-CF/KGM composite gels in whipped cream.

Fluoride Ion Binding and Translocation in the CLCF Fluoride/Proton Antiporter: Molecular Insights from Combined Quantum-Mechanical/Molecular-Mechanical Modeling.

CLCF fluoride/proton antiporters move fluoride ions out of bacterial cells, leading to fluoride resistance in these bacteria. However, many details about their operating mechanisms remain unclear. Here, we report a combined quantum-mechanical/molecular-mechanical (QM/MM) study of a CLCF homologue from Enterococci casseliflavus (Eca), in accord with the previously proposed windmill mechanism. Our multiscale modeling sheds light on two critical steps in the transport cycle: (i) the external gating residue E118 pushing a fluoride in the external binding site into the extracellular vestibule and (ii) an incoming fluoride reconquering the external binding site by forcing out E118. Both steps feature competitions for the external binding site between the negatively charged carboxylate of E118 and the fluoride. Remarkably, the displaced E118 by fluoride accepts a proton from the nearby R117, initiating the next transport cycle. We also demonstrate the importance of accurate quantum descriptions of fluoride solvation. Our results provide clues to the mysterious E318 residue near the central binding site, suggesting that the transport activities are unlikely to be disrupted by the glutamate interacting with a well-solvated fluoride at the central binding site. This differs significantly from the structurally similar CLC chloride/proton antiporters, where a fluoride trapped deep in the hydrophobic pore causes the transporter to be locked down. A free-energy barrier of 10-15 kcal/mol was estimated via umbrella sampling for a fluoride ion traveling through the pore to repopulate the external binding site.

Polyamide Membranes with Tunable Surface Charge Induced by Dipole-Dipole Interaction for Selective Ion Separation.

Nanofiltration (NF) has the potential to achieve precise ion-ion separation at the subnanometer scale, which is necessary for resource recovery and a circular water economy. Fabricating NF membranes for selective ion separation is highly desirable but represents a substantial technical challenge. Dipole-dipole interaction is a mechanism of intermolecular attractions between polar molecules with a dipole moment due to uneven charge distribution, but such an interaction has not been leveraged to tune membrane structure and selectivity. Herein, we propose a novel strategy to achieve tunable surface charge of polyamide membrane by introducing polar solvent with a large dipole moment during interfacial polymerization, in which the dipole-dipole interaction with acyl chloride groups of trimesoyl chloride (TMC) can successfully intervene in the amidation reaction to alter the density of surface carboxyl groups in the polyamide selective layer. As a result, the prepared positively charged (PEI-TMC)-NH2 and negatively charged (PEI-TMC)-COOH composite membranes, which show similarly high water permeance, demonstrate highly selective separations of cations and anions in engineering applications, respectively. Our findings, for the first time, confirm that solvent-induced dipole-dipole interactions are able to alter the charge type and density of polyamide membranes and achieve tunable surface charge for selective and efficient ion separation.

Enhanced benzene vapor adsorption through microwave-assisted fabrication of activated carbon from peanut shells using ZnCl2 as an activating agent.

Herein, microwave-assisted activated carbon (MW-AC) was fabricated from peanut shells using a ZnCl2 activator and utilized for the first time to eliminate benzene vapor as a volatile organic compound (VOC). During the MW-AC production process, which involved two steps-microwave treatment and muffle furnace heating-we investigated the effects of various factors and achieved the highest iodine number of 1250 mg/g. This was achieved under optimal operating conditions, which included a 100% impregnation ratio, CO2 as the gas in the microwave environment, a microwave power set at 500 W, a microwave duration of 10 min, an activation temperature of 500 °C and an activation time of 45 min. The structural and morphological properties of the optimized MW-AC were assessed through SEM, FTIR, and BET analysis. The dynamic adsorption process of benzene on the optimized MW-AC adsorbent, which has a significant BET surface area of 1204.90 m2/g, was designed using the Box-Behnken approach within the response surface methodology. Under optimal experimental conditions, including a contact duration of 80 min, an inlet concentration of 18 ppm, and a temperature of 26 °C, the maximum adsorption capacity reached was 568.34 mg/g. The experimental data are better described by the pseudo-second-order kinetic model, while it is concluded that the equilibrium data are better described by the Langmuir isotherm model. MW-AC exhibited a reuse efficiency of 86.54% for benzene vapor after five consecutive recycling processes. The motivation of the study highlights the high adsorption capacity and superior reuse efficiency of MW-AC adsorbent with high BET surface area against benzene pollutant. According to our results, the developed MW-AC presents itself as a promising adsorbent candidate for the treatment of VOCs in various industrial applications.

Comparison of methods for activating sesame stalk lignin biochar for removing benzo[a]pyrene from sesame oil.

In this study, three activators and two activation methods were employed to activate sesame lignin-based biochar. The biochar samples were comprehensively characterized, their abilities to adsorb benzo[a]pyrene (BaP) from sesame oil were assessed, and the mechanism was analyzed. The results showed that the biochar obtained by one-step activation was more effective in removing BaP from sesame oil than the biochar produced by two-step activation. Among them, the biochar generated by one-step activation with ZnCl2 as the activator had the largest specific surface area (1068.8776 m3/g), and the richest mesoporous structure (0.7891 m3/g); it removed 90.53 % of BaP from sesame oil. BaP was mainly adsorbed by the mesopores of biochar. Mechanistically, pore-filling, π-π conjugations, hydrogen bonding, and n-π interactions were involved. The adsorption was spontaneous and heat-absorbing. In conclusion, the preparation of sesame lignin biochar using one-step activation with ZnCl2 as the activator was found to be the best for removing BaP from sesame oil. This biochar may be an economical adsorbent for the industrial removal of BaP from sesame oil.

Coupling and regulation mechanisms of the flavin-dependent halogenase PyrH observed by infrared difference spectroscopy.

Flavin-dependent halogenases are central enzymes in the production of halogenated secondary metabolites in various organisms and they constitute highly promising biocatalysts for regioselective halogenation. The mechanism of these monooxygenases includes formation of hypohalous acid from a reaction of fully reduced flavin with oxygen and halide. The hypohalous acid then diffuses via a tunnel to the substrate-binding site for halogenation of tryptophan and other substrates. Oxidized flavin needs to be reduced for regeneration of the enzyme, which can be performed in vitro by a photoreduction with blue light. Here, we employed this photoreduction to study characteristic structural changes associated with the transition from oxidized to fully reduced flavin in PyrH from Streptomyces rugosporus as a model for tryptophan-5-halogenases. The effect of the presence of bromide and chloride or the absence of any halides on the UV-vis spectrum of the enzyme demonstrated a halide-dependent structure of the flavin-binding pocket. Light-induced FTIR difference spectroscopy was applied and the signals assigned by selective isotope labeling of the protein moiety. The identified structural changes in α-helix and ß-sheet elements were strongly dependent on the presence of bromide, chloride, the substrate tryptophan, and the product 5-chloro-tryptophan, respectively. We identified a clear allosteric coupling in solution at ambient conditions between cofactor-binding site and substrate-binding site that is active in both directions, despite their separation by a tunnel. We suggest that this coupling constitutes a fine-tuned mechanism for the promotion of the enzymatic reaction of flavin-dependent halogenases in dependence of halide and substrate availability.

Organometallic Ru(II), Rh(III) and Re(I) complexes of sterane-based bidentate ligands: synthesis, solution speciation, interaction with biomolecules and anticancer activity.

In this study, we present the synthesis, characterization and in vitro cytotoxicity of six organometallic [Ru(II)(η6-p-cymene)(N,N)Cl]Cl, [Rh(III)(η5-C5Me5)(N,N)Cl]Cl and [Re(I)(CO)3(N,N)Cl] complexes, in which the (N,N) ligands are sterane-based 2,2'-bipyridine derivatives (4-Me-bpy-St-OH, 4-Ph-bpy-St-OH). The solution chemical behavior of the ligands and the complexes was explored by UV-visible spectrophotometry and 1H NMR spectroscopy. The ligands and their Re(I) complexes are neutral at pH = 7.40; this contributes to their highly lipophilic character (log D7.40 > +3). The Ru(II) and Rh(III) half-sandwich complexes are much more hydrophilic, and this property is greatly affected by the actual chloride ion content of the medium. The half-sandwich Ru and Rh complexes are highly stable in 30% (v/v) DMSO/water (<5% dissociation at pH = 7.40); this is further increased in water. The Rh(III)(η5-C5Me5) complexes were characterized by higher water/chloride exchange and pKa constants compared to their Ru(II)(η6-p-cymene) counterparts. The Re(I)(CO)3 complexes are also stable in solution over a wide pH range (2-12) without the release of the bidentate ligand; only the chlorido co-ligand can be replaced with OH- at higher pH values. A comprehensive discussion of the binding affinity of the half-sandwich Ru(II) and Rh(III) complexes toward human serum albumin and calf-thymus DNA is also provided. The Ru(II)(η6-p-cymene) complexes interact with human serum albumin via intermolecular forces, while for the Rh(III)(η5-C5Me5) complexes the coordinative binding mode is suggested as well. They are also able to interact with calf-thymus DNA, most likely via the coordination of the guanine nitrogen. The Ru(II)(η6-p-cymene) complexes were found to be the most promising among the tested compounds as they exhibited moderate-to-strong cytotoxic activity (IC50 = 3-11 µM) in LNCaP as well as in PC3 prostate cells in an androgen receptor-independent manner. They were also significantly cytotoxic in breast and colon adenocarcinoma cancer cell lines and showed good selectivity for cancer cells.

Interactive Effects of Copper-Silver Mixtures at the Intestinal Epithelium of Rainbow Trout: An In Vitro Approach.

While metals are present in mixture in the environment, metal toxicity studies are usually conducted on an individual metal basis. There is a paucity of data in the existing literature regarding specific metal-metal interactions and their effect on metal toxicity and bioavailability. We studied interactions of a silver (Ag)-copper (Cu) mixture at the intestinal epithelium using an intestinal cell line derived from rainbow trout (Oncorhynchus mykiss), the RTgutGC. Exposures were conducted in media containing different chloride concentrations (low chloride, 1 mM; high chloride, 146 mM), thus resulting in different metal speciation. Cytotoxicity was evaluated based on two endpoints, cell metabolic activity and cell membrane integrity. The Ag-Cu mixture toxicity was assessed using two designs: independent action and concentration addition. Metal mixture bioavailability was studied by exposing cells to 500 nM of Ag or Cu as a single metal or a mixture (i.e., 500 nM of Cu plus 500 nM of Ag). We found an antagonistic effect in the low-chloride medium and an additive/synergistic effect in the high-chloride medium. We found that Cu dominates over Ag toxicity and bioavailability, indicating a competitive inhibition when both metals are present as free metal ions in the exposure media, which supports our hypothesis. Our study also suggests different mechanisms of uptake of free metal ions and metal complexes. The study adds valuable information to our understanding of the role of metal speciation on metal mixture toxicity and bioavailability. Environ Toxicol Chem 2024;43:105-114. © 2023 SETAC.