Classification of skin sensitizing substances: a comparison between approaches used by the DFG-MAK Commission and the European Union legislation.

A systematic classification of substances (or mixtures of substances) with regard to various toxicological endpoints is a prerequisite for the implementation of occupational safety strategies. As its principal task the "Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area" of the "Deutsche Forschungsgemeinschaft" (DFG-MAK Commission) derives and recommends maximum workplace concentrations and biological tolerance values (MAK and BAT values) based exclusively on scientific arguments. Several endpoints are evaluated separately in detail, e.g. carcinogenicity, risks during pregnancy, germ cell mutagenicity or contribution to systemic toxicity after cutaneous absorption. Skin- and airway sensitization is also considered; the present paper focuses on these two endpoints.

Skin sensitization testing in potency and risk assessment.

The purpose of this article is to review, and make recommendations for, the use of relevant skin sensitization test methods, for the purposes of determination of relative potency and the threshold dose necessary for the induction of skin sensitization, and for risk assessment. In addressing the first area, the utility of three guinea pig tests (the guinea pig maximization test, the occluded patch test, and the open epicutaneous test) of the local lymph node assay (LLNA) and of human volunteer testing for the assessment of relative potency and identification of thresholds for sensitization were considered. The following conclusions were drawn. (1) Although attempts have been made to modify the guinea pig maximization test for the purposes of deriving dose-response relationships, this method is usually unsuitable for determination of relative sensitizing potency. (2) Guinea pig methods that do not require the use of adjuvant and which employ a relevant route of exposure (the occluded patch test and the open epicutaneous test) are more appropriate for the assessment of relative skin-sensitizing potency. (3) The LLNA is suitable for the determination of relative skin sensitizing potency, and the adaptation of this method for derivation of comparative criteria such as EC3 values (the estimated concentration of test chemical required to induce a stimulation index of 3 in the LLNA) provides an effective and quantitative basis for such measurements. (4) For all the methods identified above, potency is assessed relative to other chemical allergens of known skin sensitizing potential. The estimation of likely threshold concentrations is dependent upon the availability of suitable benchmark chemicals of known potency for human sensitization. (5) Human testing (and specifically, the Human Repeat Insult Patch Test) can provide information of value in confirming the absence of skin sensitizing activity of formulations and products under specific conditions of use and exposure. Based on the above, the following recommendations are made. (1) If results are already available from suitable guinea pig tests, then judicious interpretation of the data may provide information of value in assessing relative skin sensitizing potency. This option should be explored before other analyses are conducted. (2) The LLNA is the recommended method for new assessments of relative potency, and/or for the investigation of the influence of vehicle or formulation on skin sensitizing potency. (3) Whenever available, human skin sensitization data should be incorporated into an assessment of relative potency. With respect to risk assessment, the conclusion drawn is that all the available data on skin-sensitizing activity in animals and man should be integrated into the risk-assessment process. Appropriate interpretation of existing data from suitable guinea pig studies can provide valuable information with respect to potency, as the first step in the development of a risk assessment. However, for de novo investigations, the LLNA is the method favored for providing quantitative estimations of skin-sensitizing potency that are best suited to the risk assessment process. Finally, human testing is of value in the risk assessment process, but is performed only for the purposes of confirming product safety.

Three-generation reproductive toxicity study of dietary bisphenol A in CD Sprague-Dawley rats.

Bisphenol A (BPA) was evaluated at concentrations of 0, 0.015, 0.3, 4.5, 75, 750, and 7500 ppm ( approximately 0.001, 0.02, 0.3, 5, 50, and 500 mg/kg/day of BPA) administered in the diet ad libitum to 30 CD((R)) Sprague-Dawley rats/sex/dose for 3 offspring generations, 1 litter/generation, through F3 adults. Adult systemic toxicity at 750 and 7500 ppm in all generations included: reduced body weights and body weight gains, reduced absolute and increased relative weanling and adult organ weights (liver, kidneys, adrenals, spleen, pituitary, and brain), and female slight/mild renal and hepatic pathology at 7500 ppm. Reproductive organ histopathology and function were unaffected. Ovarian weights as well as total pups and live pups/litter on postnatal day (PND) 0 were decreased at 7500 ppm, which exceeded the adult maximum tolerated dose (MTD). Mating, fertility, gestational indices; ovarian primordial follicle counts; estrous cyclicity; precoital interval; gestational length; offspring sex ratios; postnatal survival; nipple/areolae retention in preweanling males; epididymal sperm number, motility, morphology; daily sperm production (DSP), and efficiency of DSP were all unaffected. At 7500 ppm, vaginal patency (VP) and preputial separation (PPS) were delayed in F1, F2, and F3 offspring, associated with reduced body weights. Anogenital distance (AGD) on PND 0 was unaffected for F2 and F3 males and F3 females (F2 female AGD was increased at some doses, not at 7500 ppm, and was considered not biologically or toxicologically relevant). Adult systemic no observed adverse effect level (NOAEL) = 75 ppm (5 mg/kg/day); reproductive and postnatal NOAELs = 750 ppm (50 mg/kg/day). There were no treatment-related effects in the low-dose region (0.001-5 mg/kg/day) on any parameters and no evidence of nonmonotonic dose-response curves across generations for either sex. BPA should not be considered a selective reproductive toxicant, based on the results of this study.

Chemical substances and contact allergy--244 substances ranked according to allergenic potency.

From 1985 to 2001 a group consisting of thirty experts including dermatologists from universities, representatives from the chemical industry and from regulatory authorities elaborated and consequently decided on the potency ranking of chemicals with contact allergenic properties. These chemicals were defined either as synthetic chemicals or as chemicals identified as ingredients in natural products. On 244 substances clinical and experimental data on humans and results of animal tests as documented in the scientific literature were carefully collected and evaluated. This careful evaluation and assessment of these chemicals clearly demonstrate that ranking of substances according to their allergenic potency is possible and justified. It was decided to rank the most potent contact allergens in Category A of substances having significant allergenic properties. Substances with a solid-based indication of a contact allergenic potential and substances with the capacity of cross-reactions were listed in Category B and substances with insignificant or questionable allergenic effects were listed in Category C. An assessment of these compiled data is published here. Three Appendices give a comprehensive overview of the 98 substances listed in Category A, the 77 substances listed in Category B and the 69 substances listed in Category C.

Skin sensitisation testing--new perspectives and recommendations.

Various methodological aspects of skin sensitisation testing have been explored, particularly in the context of animal welfare considerations and reliability and sensitivity of test methods. Recommendations are made for the conduct of current and proposed OECD skin sensitisation tests with respect to appropriate test configurations for the purposes of hazard identification and labelling, and the requirement for positive controls. Specifically, the following aspects of guinea pig sensitisation test methods have been addressed: (1) the number of test and control animals required; (2) the option of using joint positive controls between independent laboratories; (3) the choice of positive control chemicals; (4) the optimal conduct and interpretation of rechallenge; and (5) the requirement for pretreatment with sodium lauryl sulfate. In addition, the use of the murine local lymph node assay (LLNA) has been considered. A number of conclusions have been drawn and recommendations made as follows: In many instances, particularly with the conduct of the guinea pig maximisation test, it is acceptable to halve the number of test and control animals used. An optional scheme for the conduct of joint positive control studies within a co-ordinated group of laboratories is appropriate. Only one positive control chemical (alpha-hexyl cinnamic aldehyde) is necessary for the routine assessment of assay sensitivity. The proper conduct and interpretation of rechallenge can provide valuable information and confirmation of results in guinea pig sensitisation tests. Sodium lauryl sulfate should no longer be used as a pretreatment in the guinea pig maximisation test. The LLNA is a viable and complete alternative to traditional guinea pig test methods for the purposes of skin sensitisation hazard identification. These recommendations provide the opportunity for both animal welfare benefits and improved hazard identification.

When should a substance be designated as sensitizing for the skin ('Sh') or for the airways ('Sa')?

In the List of MAK and BAT Values compounds are designated with 'Sa' ('sensitizing for the airways') or 'Sh' ('sensitizing for the skin') if, according to scientific evidence, they are allergens. Mainly based on suggestions by a WHO working group and based on our own experience, extended criteria have been elaborated by the working group 'skin and allergy' of the Commission of the Deutsche Forschungsgemeinschaft for the Investigation of Health Hazards of Chemical Compounds in the Work Area, which are presented in this article. They serve as guidelines for deciding which substances have to be labelled 'Sa' and 'Sh', respectively, for the prevention of sensitization and subsequent allergic diseases in workers. Although in some special cases their strict application may not be deemed necessary or possible, the proposed new criteria should be used to make the procedure of classification of substances: 1) more rational, 2) more consistent, 3) more comprehensible, and 4) more transparent. This paper informs readers working scientifically or administratively in this field and invites a critical discussion of the issue.

Joint positive control testing in guinea pig skin sensitization tests. A harmonized approach.

A scheme for the performance of positive control studies within a coordinated group of laboratories was proposed (joint positive control testing). The procedure has been described, as well as the first results of the validation phase of this joint positive control testing project. Adoption of this proposal within the participating six laboratories would lead to a reduction in the number of guinea pigs required for reliability and sensitivity checks from current approximate 12 studies per year down to 2 studies, i.e., 150-300 fewer animals per year. Another benefit would be the use of a harmonized, and therefore more comparable, method to perform guinea pig tests and interpret the data. In the validation phase of joint reading of the positive control studies, the congruency of reading could clearly be demonstrated. From the experience gained up to now, it was possible to draw the conclusion that a coordinated interlaboratory approach for positive control testing was fully acceptable and an improvement with regard to animal welfare.