Skin Corrosion Test Research Articles

Evaluation of a New In Chemico Skin Corrosion Test

The purpose of this study is to evaluate a novel macromolecular test method for the identification of dermal corrosives. The simple in chemico test procedure involves allowing the material to be tested to interact with a skin biomarker for corrosivity and then adding a detection reagent. The corrosivity of the test substance is predicted based on the measured macromolecular damage, which results in reduced optical density of the detection reagent as compared with controls. This study aims to determine if such an extremely simple, cell-free test method can accurately identify dermal corrosives. To determine predictivity and repeatability, we tested 60 chemicals (30 in vivo dermal corrosives and 30 in vivo dermal noncorrosives; all tested in triplicate) representative of a broad range of chemical classes, functional groups, mixtures, and levels of toxicity. Validation results indicate the GHS multicategory and packing group assignment accuracy is on par with that of the Reconstructed Human Epidermis test method and the Membrane Barrier test method and for the global identification of corrosives, the method has a considerably higher accuracy (98 % vs. ∼80 %).

P21 Advancing dermatological care: a comprehensive evaluation of the steroid-free GL-1 formulation

Abstract Introduction and aims The prolonged use of topical steroids introduces notable dermatological risks, including the onset of permanent stretch marks, bruising, discoloration and vascular abnormalities. This study presents an innovative strategy by formulating GL-1, a steroid-free treatment amalgamating seaweed bioactives, sesame oil and naturally occurring salts and minerals. The objective of this study was to address the limitations of conventional topical steroids and introduce pioneering solutions to dermatological care. Methods To assess the safety and efficacy of GL-1 (product 1), a comprehensive battery of in vitro tests was employed. The skin corrosion test (OECD 431) established the noncorrosive nature of GL-1, validating its safety on reconstructed human epidermis. The skin irritation test (OECD 439) demonstrated the nonirritating attributes of GL-1, aligning with the United Nations Globally Harmonized System of Classification and Labelling guidelines. Additionally, the bovine corneal opacity and permeability test (OECD 437) affirmed the noninducing nature of GL-1 in relation to eye irritation or serious eye damage. Results Preliminary outcomes from dermal clinical trials indicate promising Results for GL-1. In vitro experiments on HaCaT and human dermal fibroblasts, adult (HDFa) cell lines highlighted the noncytotoxicity of GL-1, contrasting with the ‘Cortopin’, which exhibited cytotoxicity. GL-1 exhibited a proliferative effect in HDFa cells, signifying its potential for regenerative applications. Furthermore, both GL-1 and ‘Cortopin’ displayed antioxidant activity in HaCaT cells, underscoring the additional merits of our formulation. Conclusions Our steroid-free formulation, GL-1, demonstrates exceptional safety and efficacy through a meticulous series of in vitro assessments and initial clinical trials. These findings position GL-1 as a promising candidate for diverse dermal applications, offering a secure alternative to traditional topical steroids. The potential impact of GL-1 transcends its therapeutic efficacy, emphasizing its robust safety profile validated through multiple in vitro evaluations.

Evaluating the QileX-RhE skin corrosion test for chemical subcategorization in accordance with OECD TG 431

Skin corrosion testing is integral to evaluating the potential harm posed by chemicals, impacting regulatory decisions on safety, transportation, and labeling. Traditional animal testing methods are giving way to in vitro alternatives, such as reconstructed human epidermis (RhE) models, aligning with evolving ethical standards. This study evaluates the QileX-RhE test system's performance for chemical subcategorization within the OECD TG 431 framework. Results demonstrate its ability to differentiate subcategories, accurately predicting 83% of UN GHS Category 1A and 73% of UN GHS Category 1B/1C chemicals with 100% sensitivity in corrosive prediction. Additionally, this study provides a comprehensive assessment of the test method's performance by employing nuanced parameters such as positive predictive value (PPV), negative predictive value (NPV), post-test odds and likelihood rations, offering valuable insights into the applicability and effectiveness of the QileX-RhE test method.

Earthworm half-pipe assay: A new alternative in vivo skin corrosion test using invertebrates

As a result of the efforts to introduce the principle of the 3Rs (replacement, reduction, and refinement) into animal testing, alternative in vitro skin corrosion test methods have been developed and standardized globally. However, alternative in vitro skin corrosion test methods have some limitations in terms of the use of humanely killed rats or commercial models and kits. The present study focused on the applicability of invertebrates as alternative in vivo skin models. Even though earthworm skin comprises the same biomolecules as human skin, the possibility of using earthworm skin as an alternative for skin testing remains unexplored. In this study, we developed a half-pipe tool for earthworm skin corrosion testing and optimized the test protocol. Subsequently, the applicability of the earthworm half-pipe assay for corrosion testing with six chemicals, including inorganic acids, organic acids, and alkalis, was investigated using stereomicroscopy and electron microscopy. It was observed that the specific concentrations for earthworm skin corrosion were lower than those for animal or in vitro tests. Therefore, the sensitivity of the earthworm half-pipe assay indicates that it could be useful as a screening tool before conducting in vivo animal tests or in vitro skin tests. This new method can contribute to research on alternative skin corrosion tests by reducing ethical issues, time, and cost while achieving effective results.

Ex Vivo Human Skin: An Alternative Test System for Skin Irritation and Corrosion Assays.

Native human skin has been reported in the literature as being an important experimental model for studying skin biology. Studies performed by our group have shown that ex vivo skin, from elective plastic surgery, maintains the biological characteristics of native skin under specific culture conditions. As such, it might be a feasible model for the safety and efficacy testing of topical substances. While Brazil is at the forefront of global regulation implementation, Brazilian researchers are not always able to transfer certain widely used protocols to their laboratories, particularly protocols that involve the use of reconstructed tissues with limited viability, such as those for skin corrosion (OECD TG 431) and irritation testing (OECD TG 439). In this study, we investigated the applicability of the ex vivo skin model to the evaluation of irritation and corrosion potential of a number of proficiency substances described in TG 431 and TG 439. The skin fragments were standardised in size and diameter, and placed into cell culture inserts. The experimental protocol was conducted according to TG 431 and TG 439. The results obtained show that ex vivo skin could represent a promising tool for the evaluation of irritation and corrosion potential of substances (subject to inclusion and exclusion criteria), as recommended by OECD guidelines. While this is a proof-of-concept study, the use of ex vivo skin should be considered for such testing.

Accounting for Precision Uncertainty of Toxicity Testing: Methods to Define Borderline Ranges and Implications for Hazard Assessment of Chemicals

For hazard classifications of chemicals, continuous data from animal‐ or nonanimal testing methods are often dichotomized into binary positive/negative outcomes by defining classification thresholds (CT). Experimental data are, however, subject to biological and technical variability. Each test method's precision is limited resulting in uncertainty of the positive/negative outcome if the experimental result is close to the CT. Borderline ranges (BR) around the CT were suggested, which represent ranges in which the study result is ambiguous, that is, positive or negative results are equally likely. The BR reflects a method's precision uncertainty. This article explores and compares different approaches to quantify the BR. Besides using the pooled standard deviation, we determine the BR by means of the median absolute deviation (MAD), with a sequential combination of both methods, and by using nonparametric bootstrapping. Furthermore, we quantify the BR for different hazardous effects, including nonanimal tests for skin corrosion, eye irritation, skin irritation, and skin sensitization as well as for an animal test on skin sensitization (the local lymph node assay, LLNA). Additionally, for one method (direct peptide reactivity assay) the BR was determined experimentally and compared to calculated BRs. Our results demonstrate that (i) the precision of the methods is determining the size of their BRs, (ii) there is no “perfect” method to derive a BR, alas, (iii) a consensus on BR is needed to account for the limited precision of testing methods.

Real world-like simulations show efficient predictive power of in vitro skin corrosion tests used as stand-alone and in combination and how can toxicologists take advantage of them

Historically, performance of in vitro toxicology test methods has been evaluated and considered for regulatory purposes based on sensitivity & specificity values derived from validation studies. Other indicators are however useful to evaluate in vitro tests such as positive & negative predictive values (PPV & NPV), likelihood ratios (LRs) or odds ratio (OR). These indicators, which are routinely used in diagnostic tests, if adapted and adequately applied to in vitro tests would help determine their realistic predictive power in real world-like situation and help risk assessors know how they can rely on in vitro tests used for their safety assessments. This paper performs a series of simulations considering the actual distribution in ECHA C&L inventory of skin corrosive chemicals to calculate several of these indicators of performance (PPV, NPV, LRs, OR). It shows applied examples of predictive power on EpiSkin™ and SkinEthic™ RHE two validated in vitro skin corrosive tests, explains how to build testing strategies based on these examples, compares so called ‘bottom-up’ and ‘top-down’ approaches, and demonstrates the number of tests required, and how risk assessors can practically take advantage of this.

Evaluation of toxicity profiles of rare earth elements salts (lanthanides)

The presented study aims to extend the knowledge of toxicological profile of rare earth elements salts (REEs). The basal toxicity test performed comprised assessment of cytotoxicity (3T3 Balb/c Neutral Red Uptake Test) that allows for calculation of LD50 (rats) on the basis of the concentration which leads to a 50% reduction in cell growth (IC50). Environmental toxicity was addressed by the Tubifex tubifex (T.t.) express test. The in vitro skin irritation (OECD TG 439) and skin corrosion tests (OECD TG 431) utilizing the 3D in vitro reconstructed human epidermal model EpiDerm (MatTek IVSL, SK) were used for assessment of skin irritation and corrosion potential hazard of REEs. Mutagenic effects were determined using the bacterial reverse mutation assay (Ames Test) on 5 Salmonella typhimurium strains with and without metabolic activation (OECD TG 471). Endocrine disruption was evaluated by means of a yeast-based assay YES/YAS (Xenometrix, CH). Skin sensitization was assessed using the LuSens assay, based on a genetically modified human keratinocyte cell line (OECD TG 442D). The tested REEs have no potential of mutagenicity or skin sensitization, exhibit very weak endocrine disruption potential and only exceptional local irritation/corrosion effects for thulium (III) chloride anhydrous, but have acute and chronic toxic effects on the aquatic environment.

A ready-to-use integrated in vitro skin corrosion and irritation testing strategy using EpiSkin™ model in China

The need of in vitro alternative methods has been increasing in toxicology research as well as in cosmetic industry in China recently. Following the establishment of China EpiSkin™ skin corrosion and irritation testing methods, both as stand-alone in vitro tests according to Organization for Economic Co-operation and Development (OECD) TG 431 and TG 439, the present study aims to evaluate the use of these two methods within the Integrated Approach on Testing and Assessment (IATA). The IATA, adopted by OECD as Guidance Document 203, provides guidance on the integration of existing and new data in a modular approach for classification and labelling of chemicals according to Globally Harmonized System of classification and labeling of chemicals (GHS) issued by the United Nations (UN). By applying bottom-up and top-down integrated testing strategies to a set of 60 chemicals representing various chemicals classes (organic acid/base/neutral, inorganic acid/base/salt, and surfactant) and physical states (liquid and solid), the results demonstrated that both strategies reached a high overall accuracy of 83.3% to distinguish non-classified, Category 2, Category 1B/1C and Category 1A according to UN GHS, identically. In conclusion, the integration of China EpiSkin™ skin corrosion and irritation testing data into either bottom-up or top-down strategy allows accurate assessment of potential skin hazard of chemicals. It brings a future extension of application of alternative methods and implementation of alternative testing strategies in China.

Human-Derived In Vitro Models Used for Skin Toxicity Testing Under REACh.

In regulatory toxicology, in vivo studies are still prevailing, and human-derived in vitro models are mostly used in testing for local toxicity to the skin and the eyes. A single in vitro model may be limited to address one or few molecular or cellular events leading to adverse outcomes. Hence, in many instances their regulatory use involves the combination of several in vitro models to assess the hazard potential of test substance. A so-called defined approach combines different testing methods and a 'data interpretation procedure' to obtain a comprehensive overall assessment which is used for the regulatory hazard classification of the test substance.Validation is a prerequisite of regulatory acceptance of new testing methods: This chapter provides an overview of the method development from an experimental method to a test guideline via application of GIVIMP (good in vitro method practice), standardization, validation to the regulatory adoption as an OECD test guidelines. Quandaries associated with the validation towards reference data from in vivo animal studies with limited accuracy and limited human relevance are discussed, as well as uncertainty and limitations arising from restricted applicability and technical and biological variance of the in vitro methods.This chapter provides an overview of human-derived in vitro models currently adopted as OECD test guidelines: From the first skin corrosion tests utilizing reconstructed human epidermis models (RhE), to models to test for skin irritation, phototoxicity, eye irritation, and skin sensitization. The latter is using a battery of different methods and defined approaches which are still under discussion for their regulatory adoption. They will be a vanguard of future applications of human-derived models in regulatory toxicology. RhEs for testing of genotoxicity and of dermal penetration and absorption, have been developed, underwent validation studies and may soon be adopted for regulatory use; these are included in this chapter.

Regarding the references for reference chemicals of alternative methods

The selection of reference and proficiency chemicals is an important basis for method validation and proficiency evaluations. Reference chemicals are a set of test substances used by a method developer to evaluate the reliability and relevance of a new method, in comparison to reference data (usually to a validated reference method). Proficiency chemicals, as defined in OECD Guidance Document on Good In Vitro Method Practices, are defined post validation as a subset of the reference chemicals or other chemicals with sufficient supporting data that are used by naïve laboratories to demonstrate technical competence with a validated test method. Proficiency chemicals should cover different physical states, several chemical classes within the applicability domain of the method and yield the full range of responses (in the validated reference method and in vivo), they shall be commercially available (at non-prohibitive costs) and have high quality reference data. If reference and subsequent proficiency chemicals are chosen without sufficient evidence for their inclusion, both test method evaluation and demonstration of technical proficiency can be hampered. In this report we present cases in which the selection of reference chemicals led to problems in the reproduction of the reference results and demonstration of technical proficiency: The variability of results was not always taken into account in selection of several reference substances of the LLNA (OECD TG 429). Based on the available reference data one proficiency chemical for the Corrositex skin corrosion test (OECD TG 435) should be replaced. Likewise, the expected in vitro result for one of the proficiency chemicals for the BCOP (OECD TG 437) was difficult to reproduce in several labs. Furthermore, it was not possible to obtain one of the proficiency chemicals for the Steroidogenesis Assay (OECD TG 456) at non-prohibitive costs at a reasonable purity. Based on these, we recommend changes of current proficiency chemicals lists with established OECD Test Guidelines and provide recommendations for developing future sets of reference chemicals.

Skin corrosion test: a comparison between reconstructed human epidermis and full thickness skin models

Currently, there is a strong global trend towards the development of in vitro models to replace the use of animals in safety evaluation tests. Reconstructed Human Epidermis (RHE) models have been employed as an alternative method to animal testing of skin corrosion and irritation potential of chemical compounds. However, the consequences of an absence of the dermal compartment in these models should be considered since the cross-talk between fibroblasts and keratinocytes is fundamental for promoting proper epidermal stratification, homeostasis, inflammatory response and wound healing. In this study, we compare in-house developed models of Reconstructed Human Epidermis (i.e. USP-RHE) and full thickness skin (i.e. USP-FTS) regarding their response when submitted to skin corrosion assays, based on Guideline 431 (OECD). The results show that both models correctly classified the four substances tested (2-phenylethyl bromide, benzylacetone, lactic acid, octanoic acid) as corrosive or non-corrosive. Furthermore, we have demonstrated higher cell viability of the USP-FTS model compared to the USP-RHE model, a sign of its improved barrier function, following the exposure to the substances test on the corrosion assay. This emphasizes the importance of employing in vitro models that are more physiologically relevant and that better mimic the in vivo situation for the toxicological screening of substances.

Limited applicability of in vitro skin corrosion and irritation tests for agrochemical formulations

Limited applicability of in vitro skin corrosion and irritation tests for agrochemical formulations

Regulatory accepted but out of domain: In vitro skin irritation tests for agrochemical formulations

Several in vitro methods have gained regulatory acceptance for the prediction of skin irritation and corrosion. However, the test guidelines for the majority of in vitro methods do not state whether they are applicable to agrochemical formulations. Hence, we would like to share the results from our routine skin corrosion and irritation testing of agrochemical formulations in which both in vitro (according to OECD TG 431 and OECD TG 439) and in vivo (according to OECD TG 404) tests were conducted as regulatory requirements. The in vitro skin irritation test did not correlate well with the CLP classification by in vivo results (44% sensitivity, 60% specificity, and 54% accuracy, based on 65 data pairs). This indicates a lack of applicability of the current protocol of the in vitro skin irritation test for agrochemical formulations. The data set did not contain formulations which were skin corrosive in vivo and hence its applicability could not be assessed. The correlation of in vitro skin corrosion testing to formulations which were not corrosive in vivo was, however, high (95% specificity based on 81 data pairs).

Use of ECVAM validated EpiDerm skin corrosion test (EpiDerm SCT) for sub-categorization according to the UN GHS

Use of ECVAM validated EpiDerm skin corrosion test (EpiDerm SCT) for sub-categorization according to the UN GHS

Evaluation of fibrin-based dermal-epidermal organotypic cultures for in vitro skin corrosion and irritation testing of chemicals according to OECD TG 431 and 439

Reconstructed human epidermis (RhE) models have been used for in vitro testing of the potential harmful effects of exposure to chemical compounds on health. In the past, skin irritation and corrosion were evaluated in animal models; however, in recent years, due to the bioethics implications of the method and, to minimize the use of experimental animals, alternative procedures have been proposed. The Organisation for Economic Co-operation and Development (OECD) in its test guidelines (TG) 431 and 439 indicates the requirements for validating new methods for the evaluation of skin corrosion and irritation, respectively. Here, we present an in-house human dermal-epidermal model, useful for the performance of these tests. Using the methods described in this work, it was possible to obtain human fibrin-based dermal-epidermal organotypic skin cultures (ORGs) displaying similar histological characteristics to native skin and expressing specific differentiation epithelial proteins. The end points to classify a substance as irritant or corrosive were cell viability evaluated by MTT assay, and cytokine release measured by BD CBA for human inflammatory cytokines. According to the MTT test, the ORGs correctly classified irritating and corrosive substances. Moreover, the cytokine release assay was difficult to interpret in the context of testing chemical hazard classification. Further experiments are needed to validate this new model for the evaluation of surfactants because the fibrin matrix was affected in the presence of these substances.

Standardization of in vitro skin corrosion test using reconstructed human epidermis tissues (RhE Model)

The corrosion inhibitory effect of two 3,5-bis(disubstituted)-4-amino-1,2,4-triazole derivatives, namely 3,5-bis(4-tolyl)-4-amino-1,2,4-triazole (K1) and 3,5-bis(3,4-dimethoxyphenyl)-4-amino-1,2,4-triazole (K2), for mild steel in 2 M H3PO4 has been studied using electrochemical impedance spectroscopy (EIS), Tafel polarization curves and weight loss measurements. It was found that the inhibition efficiency of the two investigated inhibitors increases with increase in concentration of inhibitors and decreases with increase in temperature. K1 and K2 show corrosion inhibition efficiency of 80.00 and 86.35%, respectively at 10− 3 M and 308 K. Impedance experimental data revealed a frequency distribution of the capacitance, simulated as constant phase element. Potentiostatic polarization study showed that K1 and K2 are mixed-type inhibitors in 2 M H3PO4. The results obtained from electrochemical and weight loss studies were in reasonable agreement. The adsorption of K1 and K2 on steel surface obeys Langmuir's adsorption isotherm. The correlation between inhibition efficiency and molecular structure of the inhibitors is investigated by determination of chemical indexes, which were performed using density functional theory (DFT) at (B3LYP/6-31G) (d,p) level. Monte Carlo simulations were further performed to simulate the adsorption of K1 and K2 on Fe (110) surface in the presence of water and the results show that K2 is more effective as corrosion inhibitor for mild steel in 2 M H3PO4 medium than K1.

Application of MatTekIn VitroReconstructed Human Skin Models for Safety, Efficacy Screening, and Basic Preclinical Research

Abstract During the past three decades, great progress has been achieved in the development of in vitro reconstructed human epidermal (RhE) tissue models. This technology is now available via various manufacturers of commercially available tissue models, and has provided opportunities for researchers worldwide to reduce or even completely avoid in vivo experimentation for screening and prediction of cutaneous toxic effects of substances and mixtures. A major step forward in recognition of RhE models as alternative tools in toxicology was their full regulatory acceptance for in vitro skin corrosion testing in 2004 as OECD Test Guideline 431. International multicenter validation studies performed during 2004–2008 under the auspices of European Centre for the Validation of Alternative Methods (ECVAM) provided further acceptance of RhE models for in vitro skin irritation testing (OECD TG 439). The use of RhE as alternatives to animal experimentation is also promoted in the new EU regulatory framework for the ...

Two novel prediction models improve predictions of skin corrosive sub-categories by test methods of OECD Test Guideline No. 431

Alternative test methods often use prediction models (PMs) for converting endpoint measurements into predictions. Two PMs are used for the skin corrosion tests (SCTs) of the OECD Test Guideline No. 431 (TG 431). One is specific to EpiSkin™ test method, whereas EpiDerm™, SkinEthic™ RHE and epiCS® share a common PM. These methods are based on reconstructed human epidermis models wherein cell viability values are measured. Their PMs allow translating those values into sub-categories of corrosive chemicals, Category 1A (Cat1A) and a combination of Categories 1B/1C (Cat1BC), and identifying non-corrosive (NC) chemicals. EpiSkin™'s PM already results in sufficiently accurate predictions. The common PM of the three others accurately identifies all corrosive chemicals but, for sub-categorization, an important fraction of Cat1BC chemicals (40–50%) is over-predicted as Cat1A. This paper presents a post-hoc analysis of validation data on a set of n=80 chemicals. It investigates: why this common PM causes these over-predictions and how two novel PMs that we developed (PMvar1 and PMvar2) improve the predictive capacity of these methods. PMvar1 is based on a two-step approach; PMvar2 is based on a single composite indicator of cell viability. Both showed a greater capacity to predict Cat1BC, while Cat1A correct predictions remaining at least at the same level of EpiSkin™. We suggest revising TG 431, to include the novel PMs in view of improving the predictive capacity of its SCTs.

Analysis of the validated epiderm skin corrosion test (EpiDerm-SCT) and prediction model for sub-categorization according to the UN-GHS and EU-CLP

Analysis of the validated epiderm skin corrosion test (EpiDerm-SCT) and prediction model for sub-categorization according to the UN-GHS and EU-CLP