Polymeric maleimido-quaternary ammonium (QA) compounds have been shown to function as molecular tape measures when covalently tethered to external cysteine residues of a Shaker K(+) channel (Blaustein R.O., P.A. Cole, C. Williams, and C. Miller. 2000. Nat. Struct. Biol. 7:309-311). For sufficiently long compounds, the cysteine-maleimide tethering reaction creates a high concentration, at the channel's pore, of a TEA-like moiety that irreversibly blocks current. This paper investigates a striking feature of the maleimide-cysteine tethering kinetics. Strong blockers-those that induce substantial levels (>80%) of irreversible inhibition of current-react with channel cysteines much more rapidly than weak blockers and, when delivered to channels with four cysteine targets, react with multiexponential kinetics. This behavior is shown to arise from the ability of a strong blocker to concentrate its maleimide end near a channel's cysteine target by exploiting the reversible pore-blocking affinity of its QA headgroup.

Quaternary alkyl ammonium compounds (QAACs) are produced in large quantities for use as surfactants and disinfectants and also found in soils, sediments, and surface waters, where they are potentially involved in the selection of antibiotic resistance genes. Micelle formation influences fate and effects of QAACs. The critical micelle concentration (CMC) of six homologs of benzylalkylammonium chlorides (BAC) was determined in deionized water, 0.01 M CaCl2 solution, and aqueous soil extracts, using both spectrofluorometric and tensiometric methods. Additionally, eight organic model compounds were employed at concentrations of 15 mg C L-1 as background solutes in order to test the effect of dissolved organic carbon (DOC) on CMCs. Results found CMCs decreased with an increasing length of the alkyl chain from 188 mM for BAC-C8 to 0.1 mM for BAC-C18. Both methods yielded similar results for measurements in water and CaCl2 solution; however, the spectrofluorescence method did not work for soil extracts due to fluorescence quenching phenomena. In soil extracts, CMCs of BAC-C12 were reduced below 3.7 mM, while the CMC reduction in soil extracts was less pronounced for BAC-C16. Besides ionic strength, molecular structures of BACs and dissolved organic compounds also affected the CMC. The number of carboxyl groups and small molecular weights of the DOC model compounds reduced the CMCs of BAC-C12 and BAC-C16 at pH 6. This study highlights that CMCs can be surpassed in soil solution, pore waters of sediments, or other natural waters even at (small) concentrations of QAACs typically found in the environment.

The aim of this study is to determine the effect of the addition of benzalkonium chloride and cetylpyridinium chloride in three commercial root canal sealers.

During the pandemic of COVID-19, the amounts of quaternary ammonium compounds (QACs) used to inactivate the virus in public facilities, hospitals and households increased, which raised concerns about the evolution and transmission of antimicrobial resistance (AMR). Although QACs may play an important role in the propagation of antibiotic resistance gene (ARGs), the potential contribution and mechanism remains unclear. Here, the results showed that benzyl dodecyl dimethyl ammonium chloride (DDBAC) and didecyl dimethyl ammonium chloride (DDAC) significantly promoted plasmid RP4-mediated ARGs transfer within and across genera at environmental relevant concentrations (0.0004-0.4 mg/L). Low concentrations of QACs did not contribute to the permeability of the cell plasma membrane, but significantly increased the permeability of the cell outer membrane due to the decrease in content of lipopolysaccharides. QACs altered the composition and content of extracellular polymeric substances (EPS) and were positively correlated with the conjugation frequency. Furthermore, transcriptional expression levels of genes encode for mating pairing formation (trbB), DNA replication and translocation (trfA), and global regulators (korA, korB, trbA) are regulated by QACs. And we demonstrate for the first time that QACs decreased the concentration of extracellular AI-2 signals, which was verified to be involved in regulating conjugative transfer genes (trbB, trfA). Collectively, our findings underscore the risk of increased disinfectant concentrations of QACs on the ARGs transfer and provide new mechanisms of plasmid conjugation.

Quaternary ammonium compounds (QACs) are inexpensive and readily available disinfectants, and have been widely used, especially since the COVID-19 outbreak. The toxicity of QACs to humans has raised increasing concerns in recent years. Here, a new type of QACs was synthesized by replacing the alkyl chain with zinc phthalocyanine (ZnPc), which consists of a large aromatic ring and is hydrophobic in nature, similar to the alkyl chain of QACs. Three ZnPc-containing disinfectants were synthesized and fully characterized. These compounds showed 15-16 fold higher antimicrobial effect against Gram-negative bacteria than the well-known QACs with half-maximal inhibitory (IC50) values of 1.43 μM, 2.70 μM, and 1.31 μM, respectively. With the assistance of 680 nm light, compounds 4 and 6 had much higher bactericidal toxicities at nanomolar concentrations. Compound 6 had a bactericidal efficacy of close to 6 logs (99.9999% kill rate) at 1 μM to Gram-positive bacteria, including MRSA, under light illumination. Besides, these compounds were safe for mammalian cells. In a mouse model, compound 6 was effective in healing wound infection. Importantly, compound 6 was easily degraded at working concentrations under sunlight illumination, and is environmentally friendly. Thus, compound 6 is a novel and promising disinfectant.

With the pandemic of COVID-19, the application of quaternary ammonium compounds (QACs), which can be used in SARS-CoV-2 disinfection products, has increased substantially. QACs cumulated in sewer system are ultimately deposited and enriched in sludge. QACs in the environment can adversely affect human health and the environment. In this study, a liquid chromatography-mass spectrometry method was established for the simultaneous determination of 25 QACs in sludge samples. Ultrasonic extraction and filtration of the samples was performed using a 50 mM hydrochloric acid-methanol solution. The samples were separated by liquid chromatography and detected in multiple reaction monitoring mode. The matrix effects of the sludge on the 25 QACs ranged from - 25.5% to 7.2%. All substances showed good linearity in the range of 0.5-100 ng/mL, with all determination coefficients (R2) greater than 0.999. The method detection limits (MDLs) were 9.0 ng/g for alkyltrimethylammonium chloride (ATMAC), 3.0 ng/g for benzylalkyldimethylammonium chloride (BAC), and 3.0 ng/g for dialkyldimethylammonium chloride (DADMAC). The spiked recovery rates were in the range of 74-107%, while the relative standard deviations were in the range of 0.8-20.6%. Considering its sensitivity, accuracy, and easy operation, the proposed method in this study was used to determine 22 sludge samples collected from a comprehensive wastewater treatment plant. The results showed that the concentrations of ΣATMACs, ΣBACs, and ΣDADMACs were 19.684, 3.199, and 8.344 μg/g, respectively. The main components included ATMAC-C16, ATMAC-C18, ATMAC-C20, ATMAC-C22, BAC-C12, and DADMAC-C18:C18, with concentrations exceeding 1.0 μg/g. The concentration relationships of different components in the congeners showed that some components were of similar origin.

Standard cleaning processes may not remove all the soiling typically found in food industry, such as carbohydrates, fats, or proteins. Contaminants have a high impact in disinfection as their presence may reduce the activity of disinfectants. The influence of alginic acid, bovine serum albumin, yeast extract, and humic acids was assessed on the antimicrobial activities of benzalkonium chloride and cetyltrimethyl ammonium bromide against Bacillus cereus vegetative cells and Pseudomonas fluorescens. The bacteria (single and consortium) were exposed to surfactants (single and combined) in the absence and presence of potential disinfection interfering substances. The antimicrobial effects of the surfactants were assessed based on the bacterial respiratory activity measured by oxygen uptake rate due to glucose oxidation. The tested surfactants were efficient against both bacteria (single and consortium) with minimum bactericidal concentrations ranging from 3 to 35 mg·L(-1). The strongest effect was caused by humic acids that severely quenched antimicrobial action, increasing the minimum bactericidal concentration of the surfactants on P. fluorescens and the consortium. The inclusion of the other interfering substances resulted in mild interferences in the antibacterial activity. This study clearly demonstrates that humic acids should be considered as an antimicrobial interfering substance in the development of disinfection strategies.

The usage of quaternary ammonium compounds (QACs) as disinfectants has increased dramatically since the outbreak of COVID-19 pandemic, leading to potentially accelerated emergence of antibiotic resistance. Long-term exposure to subinhibitory level QACs can lead to multidrug resistance, but the contribution of mutagenesis to resistance evolution is obscure. In this study, we subcultured E. coli K-12 under subinhibitory (0.25 × and 0.5 × Minimum Inhibitory Concentration, MIC) or inhibitory (1 × and 2 × MIC) concentrations of benzalkonium chloride (BAC, mono-chained) or didecyldimethylammonium chloride (DDAC, twin-chained) for 60 days. The sensitivity of QAC-adapted cells to five typical antibiotics decreased significantly, and in particular, the MIC of rifampicin increased by 85 times. E. coli adapted faster to BAC but developed 20-167% higher antibiotic resistance with 56% more mutations under DDAC exposure. The broader mutations induced by QACs, including negative regulators (acrR, marR, soxR, and crp), outer membrane proteins and transporters (mipA and sbmA), and RNA polymerase (rpoB and rpoC), potentially contributed to the high multi-drug resistance. After QACs stresses were removed, the phenotypic resistance induced by subinhibitory concentrations of QACs was reversible, whereas that induced by inhibitory concentrations of QACs was irreversible. The different patterns and molecular mechanism of antibiotic resistance induced by BAC and DDAC is informative to estimating the risks of broader QACs present at varied concentrations in the environment.

Foodborne diseases are of major concern as they have a significant impact on public health, both socially and economically. The occurrence of cross-contamination of food in household kitchens is a serious threat and the adoption of safe food practices is of paramount importance. This work aimed to study the effectiveness and durability of a commercial quaternary ammonium compound-based surface coating which, according to the manufacturer, retains its antimicrobial activity for 30 days, and is suitable for all types of hard surfaces for the prevention and/or control of cross-contamination. For that, its antimicrobial efficacy, killing contact time and durability on three different surfaces-polyvinyl chloride, glass, and stainless-steel-against three pathogens-Escherichia coli ATCC 25922, Acinetobacter baumannii ESB260 and Listeria monocytogenes Scott A-were tested according to the current antimicrobial treated surfaces efficacy test (ISO22196:2011). The results showed that the antimicrobial coating was effective against all pathogens with a reduction of >5.0 log CFU/cm2 in less than one minute for the three surfaces, but its durability was less than one week on all surfaces cleaned in the usual manner. Additionally, trace amounts (≤0.2 mg/kg) of the antimicrobial coating, which may migrate into food when contacting the surface, did not show cytotoxicity to human colorectal adenocarcinoma cells. The suggested antimicrobial coating has the potential to significantly reduce surface contamination, ensure surface disinfection and reduce the likelihood of cross-contamination in domestic kitchens, although it is less durable than suggested. The use of this technology in household settings is an attractive complement to the existing cleaning protocols and solutions that are already in place.

Quaternary ammonium compounds (QACs) have been extensively used in the community, healthcare facilities, and food chain, in concentrations between 20 and 30,000 mg/L. Enterococcus faecalis and Enterococcus faecium are ubiquitous in these settings and are recognized as nosocomial pathogens worldwide, but QACs' activity against strains from diverse epidemiological and genomic backgrounds remained largely unexplored. We evaluated the role of Enterococcus isolates from different sources, years, and clonal lineages as hosts of QACs tolerance genes and their susceptibility to QACs in optimal, single-stress and cross-stress growth conditions. Only 1% of the Enterococcus isolates included in this study and 0.5% of publicly available Enterococcus genomes carried qacA/B, qacC, qacG, qacJ, qacZ, qrg, bcrABC or oqxAB genes, shared with >60 species of Bacillota, Pseudomonadota, Actinomycetota, or Spirochaetota. These genes were generally found within close proximity of antibiotics and/or metals resistance genes. The minimum inhibitory concentrations (MIC) and minimum bactericidal concentrations (MBC) of benzalkonium chloride (BC) and didecyldimethylammonium chloride ranged between 0.5 and 4 mg/L (microdilution: 37°C/20 h/pH = 7/aerobiosis) for 210 E. faecalis and E. faecium isolates (two isolates carrying qacZ). Modified growth conditions (e.g., 22°C/pH = 5) increased MICBC/MBCBC (maximum of eightfold and MBCBC = 16 mg/L) and changed bacterial growth kinetics under BC toward later stationary phases in both species, including in isolates without QACs tolerance genes. In conclusion, Enterococcus are susceptible to in-use QACs concentrations and rarely carry QACs tolerance genes. However, their potential gene exchange with different microbiota, the decreased susceptibility to QACs under specific environmental conditions, and the presence of subinhibitory QACs concentrations in various settings may contribute to the selection of particular strains and, thus, require a One Health strategy to maintain QACs effectiveness. IMPORTANCE Despite the increasing use of quaternary ammonium compounds (QACs), the susceptibility of pathogens to these antimicrobials remains largely unknown. Enterococcus faecium and Enterococcus faecalis are susceptible to in-use QACs concentrations and are not main hosts of QACs tolerance genes but participate in gene transfer pathways with diverse bacterial taxa exposed to these biocides. Moreover, QACs tolerance genes often share the same genetic contexts with antibiotics and/or metals resistance genes, raising concerns about potential co-selection events. E. faecium and E. faecalis showed increased tolerance to benzalkonium chloride under specific environmental conditions (22°C, pH = 5), suggesting that strains might be selected in settings where they occur along with subinhibitory QACs concentrations. Transcriptomic studies investigating the cellular mechanisms of Enterococcus adaptation to QACs tolerance, along with longitudinal metadata analysis of tolerant populations dynamics under the influence of diverse environmental factors, are essential and should be prioritized within a One Health strategy.

Quaternary ammonium compounds (QACs) are among the most effective antimicrobial agents that have been used for more than a century. However, due to the growing trend of bacterial resistance and high toxicity of QACs, research in this field remains a pressing matter. Recent studies of the structure-activity relationship suggest that the introduction of the amide functional group into QAC structures results in soft variants that retain their antimicrobial properties while opening the possibility of fine-tuned activity regulation. Here, we report the synthesis and structure-function study of three structurally distinct series of naturally derived soft QACs. The obtained 3-amidoquinuclidine QACs showed a broad range of antibacterial activities related to the hydrophobic-hydrophilic balance of the QAC structures. All three series yielded candidates with minimal inhibitory concentrations (MIC) in the single-digit μM range. Time-resolved growth analysis revealed subtle differences in the antibacterial activity of the selected candidates. The versatile MIC values were recorded in different nutrient media, suggesting that the media composition may have a dramatic impact on the antibacterial potential. The new QACs were found to have excellent potential to suppress bacterial biofilm formation while exhibiting low ability to induce bacterial resistance. In addition, the selected candidates were found to be less toxic than commercially available QACs and proved to be potential substrates for protease degradation. These data suggest that 3-amidoquinuclidine QACs could be considered as novel antimicrobial agents that pose a low threat to ecosystems and human health.

Quaternary ammonium compounds (QUATs) are commonly found in cleaning products, disinfectants, hand sanitizers, and personal care products. They have been used for >50 years and are considered safe when used according to directions. Recent papers report reduced fertility and neural tube defects in rodents after low-level exposures. To determine if QUATs interfere with mammalian reproduction and development, we conducted a methodical assessment of all available data.

Quaternary ammonium compounds (QACs) are amphiphilic molecules displaying a broad-spectrum of antibacterial activity. QACs are commonly used antiseptics in industrial, home and hospital settings. Given the emergence of the QAC-resistant bacteria, there is an urgent need to design new QACs with good antimicrobial activity, able to escape the host resistance mechanism. Therefore, a series of QACs derived from quinuclidine-3-ol and an alkyl chain of variable length (QOH-C3 to -C14), was designed and synthesized. The antimicrobial potential of the new monoquaternary QACs was surveyed against seventeen strains of emerging food spoilage and pathogenic microorganisms, including clinical multidrug-resistant ESKAPE isolates. The QOH-C14 proved to have the strongest antimicrobial activity. It was highly active against all pathogens tested, particularly against the Gram-positive bacteria with minimal inhibitory concentrations (MICs) ranging from 0.06 to 3.9 μg/mL, and fungi exerting the MIC90 between 0.12 and 3.9 μg/mL. The potency of QOH-C14, confirmed that alkyl chains are an important part of the structure with their lengths playing a critical role in bioactivity of these compounds. The atomic force microscopy images show the disruption of a cell membrane upon the treatment with QOH-C14. These results were additionally confirmed by flow cytometry and fluorescence microscopy. A relatively low toxicity toward healthy human cells underline that QOH-C14 has a potential as new QAC antimicrobial candidate.

A diverse series of 43 novel "soft antimicrobials" based on quaternary ammonium pyridoxine derivatives which include six-membered acetals and ketals of pyridoxine bound via cleavable linker moieties (amide, ester) with a fragment of fatty carboxylic acid was designed. Nine compounds exhibited in vitro promising antibacterial activity against Gram-positive and Gram-negative bacterial strains with MIC values comparable with reference antiseptics miramistin, benzalkonium chloride and chlorohexidine. On various clinical isolates, the lead compounds 6i and 12a exhibited antibacterial activity comparable with that of benzalkonium chloride while higher than that of miramistin. Moreover, 6i and 12a were able to kill bacteria embedded into the matrix of mono- and dual species biofilms. The treatment of bacterial cells by either 6i and 12a lead to fast depolarization of the membrane suggesting that the membrane is an apparent molecular target of compounds. 6i and 12a were non mutagenic neither in SOS-chromotest nor in Ames test and non-toxic in vivo at acute oral (LD50 > 2000 mg/kg) and cutaneous administration (LD50 > 2500 mg/kg) on mice. Taken together, our data allow suggesting described active compounds as promising starting point for the new antibacterial agents development.

Quaternary ammonium compounds (QACs) are commonly used in a variety of consumer, pharmaceutical, and medical products. In this study, bioaccumulation potentials of 18 QACs with alkyl chain lengths of C8-C18 were determined in the in vitro-in vivo extrapolation (IVIVE) model using the results of human hepatic metabolism and serum protein binding experiments. The slowest in vivo clearance rates were estimated for C12-QACs, suggesting that these compounds may preferentially build up in blood. The bioaccumulation of QACs was further confirmed by the analysis of human blood (sera) samples (n = 222). Fifteen out of the 18 targeted QACs were detected in blood with the ΣQAC concentrations reaching up to 68.6 ng/mL. The blood samples were collected during two distinct time periods: before the outbreak of the COVID-19 pandemic (2019; n = 111) and during the pandemic (2020, n = 111). The ΣQAC concentrations were significantly higher in samples collected during the pandemic (median 6.04 ng/mL) than in those collected before (median 3.41 ng/mL). This is the first comprehensive study on the bioaccumulation and biomonitoring of the three major QAC groups and our results provide valuable information for future epidemiological, toxicological, and risk assessment studies targeting these chemicals.

Quaternary ammonium compound based disinfectants are commonly used in pig and poultry husbandry to maintain farm hygiene. However, studies have shown that subinhibitory concentrations of these disinfectants may increase antibiotic resistance. Investigation of antibiotic susceptibility is usually assessed via the microbroth dilution method, although this conventional culture-based technique only provides information on the bacteriostatic activity of an antimicrobial agent. Therefore, experiments were performed to investigate the effect of prior benzalkonium chloride (BKC) exposure on the viability of subsequent ciprofloxacin (CIP) treated Escherichia coli.

The spore-forming bacterium Bacillus anthracis causes anthrax, an often-fatal infection in animals. Therefore, a rapid and reliable strategy to decontaminate areas, humans, and livestock from B. anthracis is very critical.

Equine piroplasmosis has become a global problem of the equine husbandry sector. Haemoprotozoans evolved very quickly and developed resistance against most of the current available drugs. Phospholipid membrane synthesis by choline kinase enzyme is vital for propagation of intra-erythrocytic protozoa parasites. This pathway was targeted in the present study. Quaternary ammonium salts (QAS) and their analogues act against choline and hamper the biosynthesis process for phosphatidylcholine. We analysed anti-T. equi activity of three QAS - decamethonium bromide (DMB), decyl trimethyl ammonium bromide (DTAB) and dodecyl trimethyl ammonium bromide (DDTAB). Theileria equi parasites in vitro treated with different concentrations of DMB, DDTAB and DTAB. Drug treated T. equi failed to multiply further in the viability test. The IC50 value of DMB, DDTAB and DTAB for growth inhibition of T. equi was 14.0 μM, 469.51 nM and 558.40 nM, respectively. DMB, DDTAB and DTAB treated T. equi parasites were observed to be devoid of internal structures, showing pyknotic and degenerative appearances. Various concentration of DMB, DDTAB and DTAB were analysed for their cytotoxicity and haemolytic activity on horse's PBMCs and RBCs. DMB was less than 10% cytotoxic to PBMCs, while DDTAB and DTAB were 40%-50% cytotoxic at 1000 μM concentrations. The respective CC50 values were 7202.96 μM, 1026.26 μM and 1263.95 μM. DMB and DTAB showed least haemolytic activity (<3%); whereas DDTAB was more haemolytic to RBCs at highest concentration of 2000 μM. The respective CC50 values of these drugs were 224495.3 μM, and 39101.35 μM; 713.54 μM. Specific selective index for DMB, DDTAB and DTAB values with respect to host's PBMC and RBC cells, were 514.50, 2185.81, 2263.52 and 16035.38, 1519.75, 70023.91, respectively. These data indicated its non-toxicity to host's cells and selective potential of anti-T. equi in vitro activity.

Alkyltrimethylammonium compounds (ATMACs), dialkyldimethylammonium compounds (DADMACs) and benzylalkyldimethylethylammonium compounds (BACs) are quaternary alkylammonium compounds (QAAC), which are released into the environment in large quantities after their use in cleaning agents and disinfectants. Despite their potential role as selective agents promoting resistance against QAACs as well as antibiotics, there is a lack of data for QAACs in soil due to the lack of sensitive analytical methods. Therefore, we present a robust and fast method for the extraction and quantification of concentrations of these compounds in soil and sewage sludge. The method is based on ultrasonic extraction (USE) with a mixture of acetonitrile and HCl followed by a solid phase extraction (SPE) cleaning step and a subsequent quantification of concentrations with high performance liquid chromatography with mass spectrometry (HPLC-MS/MS) in multi mass reaction mode (MRM). The proposed method is suitable for the quantification of ATMACs (chain length C-8 to C-16), BACs (C-8 to C-18) and DADMACs (C-8 to C-16). The achieved limits of quantification (LOQ) range from 0.1 μg kg-1 to 2.1 μg kg-1. The recovery rates of spiked soil samples for non-deuterated homologues were between 47% and 57%. The analysis of sewage sludge samples and soil samples revealed that BAC-C12 was the most abundant QAAC with concentrations up to 38600 μg kg-1 in sewage sludge and up to 81 μg kg-1 in a Mexican soil that was irrigated with wastewater. Overall, the presented methods open perspectives for effectively studying fate and effects of QAACs in soils.

Antimicrobial disinfectants are used as primary treatment options against pathogens on surfaces in healthcare facilities to help prevent healthcare associated infections (HAIs). On many surfaces, pathogenic microorganisms exist as biofilms and form an extracellular matrix that protects them from the antimicrobial effects of disinfectants. Disinfectants are used as all-purpose antimicrobials though very few specifically make biofilm efficacy claims. The objective of this study was to evaluate the efficacy of eight registered disinfectants (six registered by the Environmental Protection Agency and two products registered in by the European Chemical Agency) with general bactericidal claims, but currently no biofilm efficacy claims, against Staphylococcus aureus ATTC-6538 and Pseudomonas aeruginosa ATCC-15442 biofilms. We hypothesized that hydrogen peroxide and sodium hypochlorite disinfectant products would be more effective than quaternary ammonium chlorides.