Influence of in Vitro Assay Setup on the Apparent Cytotoxic Potency of Benzalkonium Chlorides

Groothuis, Floris A.; Timmer, Niels; Opsahl, Eystein; Nicol, Beate; Droge, Steven T.J.; Blaauboer, Bas J.; Kramer, Nynke I.

doi:10.1021/acs.chemrestox.8b00412

Cited by 20 publications

(20 citation statements)

References 54 publications

(122 reference statements)

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“…Similar calculations for T14, P16, S16, T16, T18, Q14, DDAC, and C 12‐16 ‐benzalkonium yield BCF baseline estimates that exceed empirical BCFs and BAFs by factors of 3.1, 70, 49, 42, 128, 33, 420, and 420, respectively. These lower‐than‐predicted empirical BCFs and BAFs may be due to problems with the measurements themselves, including issues associated with field sampling, such as the movements of sampled animals and overestimated bioavailability of aqueous chemical concentrations (Chen et al, 2014; Groothuis et al, 2019). Limitations on diffusive flux of permanently charged QACs across fish gills also may prolong the time to steady state or prevent systemic uptake entirely (Kierkegaard et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

Biotransformation Potential of Cationic Surfactants in Fish Assessed with Rainbow Trout Liver S9 Fractions

Droge

Armitage

Arnot

et al. 2021

Enviro Toxic and Chemistry

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Biotransformation may substantially reduce the extent to which organic environmental contaminants accumulate in fish. Presently, however, relatively little is known regarding the biotransformation of ionized chemicals, including cationic surfactants, in aquatic organisms. To address this deficiency, a rainbow trout liver S9 substrate depletion assay (RT‐S9) was used to measure in vitro intrinsic clearance rates (CLint; ml min–1 g liver–1) for 22 cationic surfactants that differ with respect to alkyl chain length and degree of methylation on the charged nitrogen atom. None of the quaternary N,N,N‐trimethylalkylammonium compounds exhibited significant clearance. Rapid clearance was observed for N,N‐dimethylalkylamines, and slower rates of clearance were measured for N‐methylalkylamine analogs. Clearance rates for primary alkylamines were generally close to or below detectable levels. For the N‐methylalkylamines and N,N‐dimethylalkylamines, the highest CLint values were measured for C10–C12 homologs; substantially lower clearance rates were observed for homologs containing shorter or longer carbon chains. Based on its cofactor dependency, biotransformation of C12–N,N‐dimethylamine appears to involve one or more cytochrome P450–dependent reaction pathways, and sulfonation. On a molar basis, N‐demethylation metabolites accounted for up to 25% of the N,N‐dimethylalkylamines removed during the 2‐h assay, and up to 55% of the removed N‐methylalkylamines. These N‐demethylation products possess greater metabolic stability in the RT‐S9 assay than the parent structures from which they derive and may contribute to the overall risk of ionizable alkylamines. The results of these studies provide a set of consistently determined CLint values that may be extrapolated to whole trout to inform in silico bioaccumulation assessments. Environ Toxicol Chem 2021;40:3123–3136. © 2021 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

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“…In addition, EC200 and CV75 were highly correlated with methacrylate esters (Figure 1). It has been reported that alkyl chain length can correlate with cytotoxicity of chemicals based on data with benzalkonium chloride with various alkyl chain length (Groothuis et al, 2019). From this information, the positive response of h‐CLAT in this study may be influenced by cytotoxicity derived from alkyl chain length of methacrylate esters.…”

Section: Discussionmentioning

confidence: 99%

Characterization of dermal sensitization potential for industrial or agricultural chemicals with EpiSensA

Mizumachi

LeBaron

Settivari

et al. 2020

J of Applied Toxicology

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The regulatory community is transitioning to the use of nonanimal methods for dermal sensitization assessments; however, some in vitro assays have limitations in their domain of applicability depending on the properties of chemicals being tested. This study explored the utility of epidermal sensitization assay (EpiSensA) to evaluate the sensitization potential of complex and/or “difficult to test” chemicals. Assay performance was evaluated by testing a set of 20 test chemicals including 10 methacrylate esters, 5 silicone‐based compounds, 3 crop protection formulations, and 2 surfactant mixtures; each had prior in vivo data plus some in silico and in vitro data. Using the weight of evidence (WoE) assessments by REACH Lead Registrants, 14 of these chemicals were sensitizers and, six were nonsensitizers based on in vivo studies (local lymph node assay [LLNA] and/or guinea pig studies). The EpiSensA correctly predicted 16/20 materials with three test materials as false positive and one silane as false negative. This silane, classified as weak sensitizer via LLNA, also gave a “false negative” result in the KeratinoSens™ assay. Overall, consistent with prior evaluations, the EpiSensA demonstrated an accuracy level of 80% relative to available in vivo WoE assessments. In addition, potency classification based on the concentration showing positive marker gene expression of EpiSensA was performed. The EpiSensA correctly predicted the potency for all seven sensitizing methacrylates classified as weak potency via LLNA (EC3 ≥ 10%). In summary, EpiSensA could identify dermal sensitization potential of these test substances and mixtures, and continues to show promise as an in vitro alternative method for dermal sensitization.

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“…Non-specific migration to plastics and binding to media constituents (for example, serum proteins, lipids) have been documented, and test chemicals may evaporate, degrade, or be metabolized. All these factors can result in an underestimation of potency [ 121 ]. This information can be obtained by measuring chemical distribution in different tissue compartments or by mathematical model predictions.…”

Section: Applications Of Ivive Approachesmentioning

confidence: 99%

IVIVE: Facilitating the Use of In Vitro Toxicity Data in Risk Assessment and Decision Making

Chang¹,

Tan

Allen³

et al. 2022

Toxics

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During the past few decades, the science of toxicology has been undergoing a transformation from observational to predictive science. New approach methodologies (NAMs), including in vitro assays, in silico models, read-across, and in vitro to in vivo extrapolation (IVIVE), are being developed to reduce, refine, or replace whole animal testing, encouraging the judicious use of time and resources. Some of these methods have advanced past the exploratory research stage and are beginning to gain acceptance for the risk assessment of chemicals. A review of the recent literature reveals a burst of IVIVE publications over the past decade. In this review, we propose operational definitions for IVIVE, present literature examples for several common toxicity endpoints, and highlight their implications in decision-making processes across various federal agencies, as well as international organizations, including those in the European Union (EU). The current challenges and future needs are also summarized for IVIVE. In addition to refining and reducing the number of animals in traditional toxicity testing protocols and being used for prioritizing chemical testing, the goal to use IVIVE to facilitate the replacement of animal models can be achieved through their continued evolution and development, including a strategic plan to qualify IVIVE methods for regulatory acceptance.

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“…ized solid-phase extraction (SPE) (Kramer et al, 2012;Neale et al, 2018). Alternatively, affinity chromatography has been applied (Groothuis et al, 2019). As an indirect approach, it is also possible to measure an activity that depends on the free concentration (e.g., receptor activation, enzyme inhibition or cell death induction) and then to extrapolate free concentrations from nominal concentrations in the same assay (Gülden et al, 2003(Gülden et al, , 2005).…”

Section: Comparison Of Concentrations and Extrapolation To Dosementioning

confidence: 99%

“…It is also a good occasion to remind the reader of the biokinetics equation The third term, K alb , refers to the distribution of compound between albumin and the aqueous phase, i.e., the albumin-water partition coefficient. There are several experimental methods to determine such equilibrium binding constants (Groothuis et al, 2019). The disadvantage is that an analytical method needs to be available for each drug that is considered.…”

Section: Compoundmentioning

confidence: 99%

Chemical concentrations in cell culture compartments (C5) – free concentrations

Kisitu

Hollert

Fisher

et al. 2020

ALTEX

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In biological systems (cell culture media, cells, body fluids), drugs/toxicants are usually not freely dissolved but partially bound to biomolecules; only a fraction of the chemical is free/unbound (fu). To predict pharmacological effects and toxicity, it is important that the fu of the drug is known. As the differences between free and nominal concentrations are determined by test system parameters (e.g., the protein and lipid content, and the type of surface material), comparison of nominal concentrations between two different new approach methods (NAM) may lead to faulty conclusions. The same problem exists when in vitro concentrations are compared to those in human subjects. Therefore, the respective fu of a chemical in a test system needs to be determined for in vitro-to-in vivo extrapolations (IVIVE). Besides direct measurements, prediction models can help to obtain fu. Here we describe a simplified approach to approximate fu and provide background information on the underlying assumptions. Comparative predictions and measurements of fu of various drugs are shown to exemplify the approach. Basic input data, like protein and lipid concentrations, are also provided. Beyond such test systems data, the only required chemical-specific inputs are the lipophilicity of the candidate drug and its ionization state, as determined by the dissociation constants of its acidic or basic groups. This overview is intended to be used by any lab scientist without specific toxicokinetics training to obtain an estimate of fu in a given cell culture medium.

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Influence of in Vitro Assay Setup on the Apparent Cytotoxic Potency of Benzalkonium Chlorides

Cited by 20 publications

References 54 publications

Biotransformation Potential of Cationic Surfactants in Fish Assessed with Rainbow Trout Liver S9 Fractions

Characterization of dermal sensitization potential for industrial or agricultural chemicals with EpiSensA

IVIVE: Facilitating the Use of In Vitro Toxicity Data in Risk Assessment and Decision Making

Chemical concentrations in cell culture compartments (C5) – free concentrations

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