Durability Analysis of Cold Spray Repairs: Phase I-Effect of Surface Grit Blasting.

This paper presents the results of an extensive investigation into the durability of cold spray repairs to corrosion damage in AA7075-T7351 aluminium alloy specimens where, prior to powder deposition, the surface preparation involved grit blasting. In this context, it is shown that the growth of small naturally occurring cracks in cold spray repairs to simulated corrosion damage can be accurately computed using the Hartman-Schijve crack growth equation in a fashion that is consistent with the requirements delineated in USAF Structures Bulletin EZ-SB-19-01, MIL-STD-1530D, and the US Joint Services Structural Guidelines JSSG2006. The relatively large variation in the da/dN versus ΔK curves associated with low values of da/dN highlights the fact that, before any durability assessment of a cold spray repair to an operational airframe is attempted, it is first necessary to perform a sufficient number of tests so that the worst-case small crack growth curve needed to perform the mandated airworthiness certification analysis can be determined.

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.

Computing the Fatigue Life of Cold Spray Repairs to Simulated Corrosion Damage.

This paper summarises the findings of an investigation into the durability of cold spray repairs, also known as supersonic particle deposition or SPD repairs, to simulated corrosion damage in AA7075-T7351 aluminium alloy specimens. A feature of this paper is that it is the first to show how to perform the mandatory durability analysis of repaired corroded structures, where the corroded material is first removed by machining and then repaired using cold spray, in a fashion consistent with the requirements delineated in USAF Structures Bulletin EZ-19-01, MIL-STD-1530D, and the US Joint Services Structural Guidelines JSSG2006.

A simplified index to quantify the irritation/corrosion potential of chemicals - Part I: Skin.

Skin irritation is a key human health endpoint assessed by in vitro and in vivo methods. The OECD TG 404 guideline (in vivo) is based on erythema and oedema translated semi-quantitatively into Draize scores, providing hazard statements for substance classification following EUCLP/UNGHS criteria. Draize scores require quantitation from subjective in vivo observations, to obtain a scoring index, the Primary Irritation Index (PII). However, it is not recognised under REACH due to translating difficulties, notably the cut-off limit for classification and non-inclusion of corrosive effects. The aim of this study was to determine if classification can be driven by just one of the observed effects, erythema only, to create a Simplified Irritation Index (SIISKIN). This simplifies the scoring calculation and reduces subjectivity. A quantitative approach with cut-off limits is thus proposed for classification. Substances can be classified as non-irritant, potentially irritant, irritant, or corrosive. The Simplifed Irritation Index (SIISKIN) is based on validated studies, representing multiple chemical groups. A significant correlation between SIISKIN and the harmonised classification was observed, and a proportionate relationship between the SIISKIN and the corresponding PII. The index proved to be useful in the development of an in silico model.

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.

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.

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.

Differences in classification for skin corrosion/irritation in EU and Ukraine: Case study of alternative (in vitro and in silico) methods application for classification of pesticide active ingredient imazamox.

In Ukraine Globally Harmonized System of classification of chemicals has not been implemented yet. In this article we analyze differences between GHS/CLP classification systems and Hygienic Classification of Pesticides by the Degree of Hazard currently in force in Ukraine in respect of approach and criteria for classification of effects on skin. As a case study, we conducted in silico modelling of herbicide imazamox using skin irritation/corrosion modules of ToxTree. The prediction of ToxTree was "Not Corrosive to skin". Then skin irritation and skin corrosion in vitro tests (OECD TGs 439, 431) were conducted. Classification of this substance based on in vitro and in vivo results according to GHS/CLP was the same, while it was not possible based on in vitro results to assign certain hazard class of Ukrainian classification due to difference in its and GHS/CLP criteria. However, ongoing process of harmonization of Ukrainian legislation with EU will give opportunity not only use alternative methods, but also adopt most recent advances and incorporate data from non-animal methods directly into classification criteria.

A way forward of alternative methods in China: Implementation of skin corrosivity potential using in vitro reconstructed human epidermis.

The purpose of present study was to investigate the applicability of reconstructed human epidermis model to identify skin corrosive UN GHS Categories 1A, 1B/1C and non-corrosive chemicals in China. By using a commercialized reconstructed human epidermis model, China EpiSkin™ which had been proven to be applicable as a stand-alone test method to predict skin irritation in previous study, the predictive capacity of corrosion was assessed with 76 chemicals that included 30 reference chemicals recommended by OECD TG 431 in this study. The latter reference chemicals were tested in three runs, the within-laboratory reproducibility reached 100%, the accuracy was 90% for distinguishing corrosive and non-corrosive chemicals and 80% for sub-categorization (Cat. 1A vs Cat. 1B/1C vs non corrosive). Additional 46 chemicals were also tested, and the overall accuracy for sub-categorization of all 76 tested chemicals was 80.3% with 91.7% sensitivity for Category 1A, 82.1% sensitivity for category 1B/1C and 75% specificity which met all required predictive capacity by OECD. The present study results show that China EpiSkin™ model can be applied to predict sub-categorization 1A and 1B/1C of corrosive chemicals. The availability of skin corrosion in vitro test method provides the applicability of in vitro non-animal testing method for chemicals widely used in various industries, and will further support the implementation and promotion of alternative methods in China.

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.

Verification and application of the rat skin transcutaneous electrical resistance test ( TER) as an alternative method to replace the animal skin corrosion test / 中国实验动物学报

Objective In order to verify an alternative method for the skin corrosion test by using transcutaneous electrical resistance ( TER) test, and to optimize the implementation criteria in OECD TG 430 procedure. Methods According to the OECD TG 430 procedure, Wistar rat skin was used to test the TER values of 16 reference chemicals, and selected the most optimal standard via different implementation criteria. The program B was chosen to make inter-laboratory comparison between 5 laboratories by testing 11 chemicals, which were identified as the optimal standard. Results After the TER test, the result of corrosion test of 16 chemicals were accordant with the reference data ( Kappa value＝0. 64). The program B was the most optimal implementation criteria, and the specificity was 66. 7% and sensitivity was 100%. There were no significant differences between the corrosion estimations of 5 laboratories, and the concordance rate of the 5 laboratories was 72. 7%. Conclusions Transcutaneous electrical resistance (TER) test is an feasible and efficient tool for skin corrosion testing, and may become a good interim test to replace the in vivo test with this ex vivo test in cosmetics chemical safety assessment, thus, to reduce the animal usage in our country.

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).

Skin Corrosion and Irritation Test of Nanoparticles Using Reconstructed Three-Dimensional Human Skin Model, EpiDermTM.

Effects of nanoparticles (NPs) on skin corrosion and irritation using three-dimensional human skin models were investigated based on the test guidelines of Organization for Economic Co-operation and Development (OECD TG431 and TG439). EpiDermTM skin was incubated with NPs including those harboring iron (FeNPs), aluminum oxide (AlNPs), titanium oxide (TNPs), and silver (AgNPs) for a defined time according to the test guidelines. Cell viabilities of EpiDermTM skins were measured by the 3-(4, 5-dimethylthi-azol-2-yl)-2.5-diphenyltetrazolium bromide based method. FeNPs, AlNPs, TNPs, and AgNPs were non-corrosive because the viability was more than 50% after 3 min exposure and more than 15% after 60 min exposure, which are the non-corrosive criteria. All NPs were also non-irritants, based on viability exceeding 50% after 60 min exposure and 42 hr post-incubation. Release of interleukin 1-alpha and histopathological analysis supported the cell viability results. These findings suggest that FeNPs, AlNPs, TNPs, and AgNPs are 'non-corrosive' and 'non-irritant' to human skin by a globally harmonized classification system.

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.

An integrated testing strategy for in vitro skin corrosion and irritation assessment using SkinEthic™ Reconstructed Human Epidermis.

The SkinEthic™ Reconstructed Human Epidermis (RHE) method has been formally adopted for the regulatory assessment of skin irritation (OECD TG 439) and corrosion (OECD TG 431). Recently, the OECD adopted an Integrated Approach on Testing and Assessment (IATA) for skin corrosion and skin irritation (OECD GD 203), which provides guidance on the integration of existing and new information in a modular approach for classification and labelling. The present study aimed to evaluate the use of the SkinEthic™ RHE model within the proposed OECD IATA. Data on 86 substances were integrated in a bottom-up and top-down testing strategy to assess their capacity for EU CLP and UN GHS classifications. For EU CLP, strategies showed an accuracy of 84.8% to discriminate non-classified from classified substances, 94.4% to discriminate corrosive from non-corrosive substances, and 68.5% to discriminate the four (sub)-categories. For UN GHS, strategies showed an accuracy of 89.5% to discriminate non-classified from classified substances, 93.4% to discriminate corrosive from non-corrosive substances, and 74.2% to discriminate four GHS (sub)-categories (excluding Category 3). In conclusion, the integration of SkinEthic™ RHE irritation and corrosion data in a bottom-up and top-down testing strategy allows the classification of substances according to EU CLP and UN GHS.

Use of HPLC/UPLC-spectrophotometry for detection of formazan in in vitro Reconstructed human Tissue (RhT)-based test methods employing the MTT-reduction assay to expand their applicability to strongly coloured test chemicals.

A number of in vitro test methods using Reconstructed human Tissues (RhT) are regulatory accepted for evaluation of skin corrosion/irritation. In such methods, test chemical corrosion/irritation potential is determined by measuring tissue viability using the photometric MTT-reduction assay. A known limitation of this assay is possible interference of strongly coloured test chemicals with measurement of formazan by absorbance (OD). To address this, Cosmetics Europe evaluated use of HPLC/UPLC-spectrophotometry as an alternative formazan measurement system. Using the approach recommended by the FDA guidance for validation of bio-analytical methods, three independent laboratories established and qualified their HPLC/UPLC-spectrophotometry systems to reproducibly measure formazan from tissue extracts. Up to 26 chemicals were then tested in RhT test systems for eye/skin irritation and skin corrosion. Results support that: (1) HPLC/UPLC-spectrophotometry formazan measurement is highly reproducible; (2) formazan measurement by HPLC/UPLC-spectrophotometry and OD gave almost identical tissue viabilities for test chemicals not exhibiting colour interference nor direct MTT reduction; (3) independent of the test system used, HPLC/UPLC-spectrophotometry can measure formazan for strongly coloured test chemicals when this is not possible by absorbance only. It is therefore recommended that HPLC/UPLC-spectrophotometry to measure formazan be included in the procedures of in vitro RhT-based test methods, irrespective of the test system used and the toxicity endpoint evaluated to extend the applicability of these test methods to strongly coloured chemicals.

The usefulness of the validated SkinEthic™ RHE test method to identify skin corrosive UN GHS subcategories.

The SkinEthic™ Reconstructed Human Epidermis (RHE) test method has been adopted within the context of OECD TG 431 for distinguishing corrosive and non-corrosive chemicals. The EU CLP classification system requires subcategorising of corrosive chemicals into the three UN GHS subcategories 1A, 1B and 1C. Since the SkinEthic™ RHE method was originally validated to discriminate corrosives from non-corrosives, the present study was undertaken to investigate its usefulness to discriminate skin corrosive UN GHS subcategories. In total 84 substances were tested in three independent runs and two prediction models (PM) were assessed, representing a pre-defined validated prediction model (PM-A) and an alternative one defined post-hoc (PM-B). The results obtained with both PM were reproducible, as shown by the ⩾92.9% concordance of classification between runs for discriminating corrosives versus non-corrosives, and the ⩾85% concordance for discriminating the GHS subcategories versus non-corrosives. Moreover results confirmed a high sensitivity of the SkinEthic™ RHE method to predict corrosives (94.9%) and good specificity (⩾73.7%) independent of the PM applied. Regarding the identification of UN GHS corrosive subcategories, PM-A resulted in 86.1% correct classifications of the GHS subcategory 1A. When using the PM-B, the identification of GHS subcategory 1B-and-1C substances improved, with 63.4% correct sub-categorisation. If considering the 30 reference chemicals as recommended in the recently revised OECD TG 431 (2013), PM-A and PM-B achieved 78.9% and 83.3% accuracy respectively for the identification of GHS subcategories and non-corrosives. They correctly predicted 90% of GHS subcategory 1A and 80% of GHS non-corrosive substances independent of the PM used. In conclusion, the SkinEthic™ RHE test method is highly reproducible and sensitive for discriminating corrosive from non-corrosive substances. Furthermore it allows reliable identification of skin corrosive GHS subcategory 1B-and-1C substances using the PM-A and PM-B, and of GHS subcategories 1A using the PM-B. Due to its high sensitivity, the test method provides high safety standards for skin corrosion testing.

Sub-categorisation of skin corrosive chemicals by the EpiSkin™ reconstructed human epidermis skin corrosion test method according to UN GHS: revision of OECD Test Guideline 431.

The EpiSkin™ skin corrosion test method was formally validated and adopted within the context of OECD TG 431 for identifying corrosive and non-corrosive chemicals. The EU Classification, Labelling and Packaging Regulation (EU CLP) system requires the sub-categorisation of corrosive chemicals into the three UN GHS optional subcategories 1A, 1B and 1C. The present study was undertaken to investigate the usefulness of the validated EpiSkin™ test method to identify skin corrosive UN GHS Categories 1A, 1B and 1C using the original and validated prediction model and adapted controls for direct MTT reduction. In total, 85 chemicals selected by the OECD expert group on skin corrosion were tested in three independent runs. The results obtained were highly reproducible both within (>80%) and between (>78%) laboratories when compared with historical data. Moreover the results obtained showed that the EpiSkin™ test method is highly sensitive (99%) and specific (80%) in discriminating corrosive from non-corrosive chemicals and allows reliable and relevant identification of the different skin corrosive UN GHS subcategories, with high accuracies being obtained for both UN GHS Categories 1A (83%) and 1B/1C (76%) chemicals. The overall accuracy of the test method to subcategorise corrosive chemicals into three or two UN GHS subcategories ranged from 75% to 79%. Considering those results, the revised OECD Test Guideline 431 permit the use of EpiSkin™ for subcategorising corrosive chemicals into at least two classes (Category 1A and Category 1B/1C).