Polycyclic aromatic hydrocarbon skin permeation efficiency in vitro is lower through human than pigskin and decreases with lipophilicity.

Polycyclic aromatic hydrocarbons (PAH) are persistent environmental pollutants, which occasionally appear as contaminants in consumer products. Upon dermal contact, transfer of PAH into the stratum corneum (s.c.) and migration through the skin may occur, resulting in this class of highly toxic compounds to become bioavailable. In this study, dermal penetration through human and porcine skin of 24 PAH, comprising broad molar mass (M: 152-302 g/mol) and octanol-water partition coefficient (logP: 3.9-7.3) ranges, was evaluated via Franz diffusion cell in vitro assays. More lipophilic and potentially more toxic PAH had decreased permeation rates through the rather lipophilic s.c. into the more hydrophilic viable (epi-)dermis. Furthermore, human skin was less permeable than pigskin, a commonly used surrogate in skin penetration studies. In particular, the s.c. of human skin retains a greater share of PAH, an effect that is more pronounced for smaller PAH. Additionally, we compared the skin permeation kinetics of different PAH in pigskin. While small PAH (M < 230 g/mol, logP < 6) permeate the skin quickly and are detected in the receptor fluid after 2 h, large PAH (M > 252 g/mol, logP ≥ 6) do not fully permeate the skin up to 48 h. This indicates that highly lipophilic PAH do not become bioavailable as readily as their smaller congeners when transferred to the skin surface. Our data suggest that pigskin could be used as a surrogate for worst case scenario estimates of dermal PAH permeation through human skin.

Levels of nicotine and tobacco-specific nitrosamines in oral nicotine pouches.

BACKGROUND: Nicotine pouches without tobacco are new products that deliver nicotine into the body via the oral mucosa. There is a lack of independent research on the chemical composition and product characteristics of these products, contributing to uncertainties regarding product regulation. This study sought to address knowledge gaps by assessing levels of nicotine and screening for tobacco-specific nitrosamines (TSNAs) in a sample of these products. METHODS: Nicotine pouches (n=44) and nicotine-free pouches (n=2) from 20 different manufacturers were analysed regarding their contents of nicotine and TSNAs by gas chromatography with flame ionisation and liquid chromatography-tandem mass spectrometry, respectively. Product labelling and pH values of aqueous extracts were determined. RESULTS: Nicotine contents of products ranged from 1.79 to 47.5 mg/pouch; median product weight, pH, and proportion of free-base nicotine were 0.643 g, 8.8, and 86%, respectively. A clear labelling of the nicotine content was missing on 29 products and nicotine strength descriptions were ambiguous. TSNAs were detected in 26 products, with a maximum of 13 ng N-nitrosonornicotine/pouch. CONCLUSION: Although nicotine pouches may potentially be a reduced risk alternative for cigarette smokers or users of some other oral tobacco products, nicotine contents of some pouches were alarmingly high. Presence of carcinogenic TSNAs in the nicotine pouches is of serious concern. Better manufacturing processes and quality control standards should be implemented. Labels of nicotine strength on most products are misleading. A strict regulation regarding nicotine contents and its labelling would be advisable.

Mono-n-hexyl phthalate: exposure estimation and assessment of health risks based on levels found in human urine samples.

Mono-n-hexyl phthalate (MnHexP) is a primary metabolite of di-n-hexyl phthalate (DnHexP) and other mixed side-chain phthalates that was recently detected in urine samples from adults and children in Germany. DnHexP is classified as toxic for reproduction category 1B in Annex VI of Regulation (EC) 1272/2008 and listed in Annex XIV of the European chemical legislation REACH; thereby, its use requires an authorisation. Health-based guidance values for DnHexP are lacking and a full-scale risk assessment has not been carried out under REACH. The detection of MnHexP in urine samples raises questions about the sources of exposure and concerns of consumer safety. Here, we propose the calculation of a provisional oral tolerable daily intake value (TDI) of 63 µg/kg body weight/day for DnHexP and compare it to intake levels corresponding to levels of MnHexP found in urine. The resulting mean intake levels correspond to less than 0.2% of the TDI, and maximum levels to less than 5%. The TDI was derived by means of an approximate probabilistic analysis using the credible interval from benchmark dose modelling of published ex vivo data on reduced foetal testosterone production in rats. Thus, for the dose associated to a 20% reduction in testosterone production, a lower and upper credible interval of 14.9 and 30.0 mg/kg bw/day, respectively, was used. This is considered a conservative approach, since apical developmental endpoints (e.g. changed anogenital distance) were only observed at higher doses. In addition, we modelled various scenarios of the exposure to the precursor substance DnHexP from different consumer products, taking measured contamination levels into account, and estimated systemic exposure doses. Of the modelled scenarios including the application of sunscreen (as a lotion or pump spray), the use of lip balm, and the wearing of plastic sandals, and considering conservative assumptions, the use of DnHexP-contaminated sunscreen was highlighted as a major contributing factor. A hypothetical calculation using conservative assumptions for the latter resulted in a margin of safety in relation to the lower credible interval of 3267 and 1007 for adults and young children, respectively. Most importantly, it was found that only a fraction of the TDI is reached in all studied exposure scenarios. Thus, with regard to the reported DnHexP exposure, a health risk can be considered very unlikely.

Lessons learned in a decade: Medical-toxicological view of tattooing.

Tattooing has been part of the human culture for thousands of years, yet only in the past decades has it entered the mainstream of the society. With the rise in popularity, tattoos also gained attention among researchers, with the aim to better understand the health risks posed by their application. 'A medical-toxicological view of tattooing'-a work published in The Lancet almost a decade ago, resulted from the international collaboration of various experts in the field. Since then, much understanding has been achieved regarding adverse effects, treatment of complications, as well as their regulation for improving public health. Yet major knowledge gaps remain. This review article results from the Second International Conference on Tattoo Safety hosted by the German Federal Institute for Risk Assessment (BfR) and provides a glimpse from the medical-toxicological perspective, regulatory strategies and advances in the analysis of tattoo inks.

Less efficient skin penetration of the metal allergen Pd2+ compared to Ni2+ and Co2+ from patch test preparations.

BACKGROUND: Contrary to Ni2+- and Co2+-induced allergic contact dermatitis (ACD), reactions against Pd2+ are rare. However, Pd2+ activates a larger T cell fraction in vitro, suggesting an inefficient skin penetration. OBJECTIVES: This study compares Ni2+, Co2+ and Pd2+ skin penetration from commonly used diagnostic patch test preparations (PTPs) and aqueous metal salt solutions. METHODS: Using Franz diffusion cell assays, we applied the metals in PTPs (5% NiSO4, 1% CoCl2, 2% PdCl2 and 3% Na2PdCl4) and in solution to pigskin for 48 h, thereby mirroring the time frame of a patch test. The different compartments were analysed individually by inductively coupled plasma mass spectrometry. RESULTS: Metal ions were mainly retained in the upper stratum corneum layers. After application of PTPs, concentrations in the viable skin were lower for Pd2+ (1 and 7 µM) compared to Ni2+ and Co2+ (54 and 17 µM). CONCLUSIONS: Ni2+ and Co2+ penetrated the skin more efficiently than Pd2+ and thus may sensitize and elicit ACD more easily. This was observed for ions applied in petrolatum and aqueous solutions. We hypothesize that the differently charged metal complexes are responsible for the varying skin penetration behaviours.

Physiologically based modelling of dermal absorption and kinetics of consumer-relevant chemicals: A case study with exposure to bisphenol A from thermal paper.

Bisphenol A (BPA) is one of the best studied industrial chemicals in terms of exposure, toxicity, and toxicokinetics. This renders it an ideal candidate to exploit the recent advancements in physiologically based pharmacokinetic (PBPK) modelling to support risk assessment of BPA specifically, and of other consumer-relevant hazardous chemicals in general. Using the exposure from thermal paper as a case scenario, this study employed the multi-phase multi-layer mechanistic dermal absorption (MPML MechDermA) model available in the Simcyp® Simulator to simulate the dermal toxicokinetics of BPA at local and systemic levels. Sensitivity analysis helped to identify physicochemical and physiological factors influencing the systemic exposure to BPA. The iterative modelling process was as follows: (i) development of compound files for BPA and its conjugates, (ii) setting-up of a PBPK model for intravenous administration, (iii) extension for oral administration, and (iv) extension for exposure via skin (i.e., hand) contact. A toxicokinetic study involving hand contact to BPA-containing paper was used for model refinement. Cumulative urinary excretion of total BPA had to be employed for dose reconstruction. PBPK model performance was verified using the observed serum BPA concentrations. The predicted distribution across the skin compartments revealed a depot of BPA in the stratum corneum (SC). These findings shed light on the role of the SC to act as temporary reservoir for lipophilic chemicals prior to systemic absorption, which inter alia is relevant for the interpretation of human biomonitoring data and for establishing the relationship between external and internal measures of exposure.

Unique and common TCR repertoire features of Ni2+ -, Co2+ -, and Pd2+ -specific human CD154 + CD4+ T cells.

BACKGROUND: Apart from Ni2+ , Co2+ , and Pd2+ ions commonly trigger T cell-mediated allergic contact dermatitis. However, in vitro frequencies of metal-specific T cells and the mechanisms of antigen recognition remain unclear. METHODS: Here, we utilized a CD154 upregulation assay to quantify Ni2+ -, Co2+ -, and Pd2+ -specific CD4+ T cells in peripheral blood mononuclear cells (PBMC). Involved αß T cell receptor (TCR) repertoires were analyzed by high-throughput sequencing. RESULTS: Peripheral blood mononuclear cells incubation with NiSO4 , CoCl2 , and PdCl2 increased frequencies of CD154 + CD4+ memory T cells that peaked at ~400 µM. Activation was TCR-mediated as shown by the metal-specific restimulation of T cell clones. Most abundant were Pd2+ -specific T cells (mean 3.5%, n = 19), followed by Co2+ - and Ni2+ -specific cells (0.6%, n = 18 and 0.3%, n = 20) in both allergic and non-allergic individuals. A strong overrepresentation of the gene segment TRAV9-2 was unique for Ni2+ -specific TCR (28% of TCR) while Co2+ and Pd2+ -specific TCR favorably expressed TRAV2 (8%) and the TRBV4 gene segment family (21%), respectively. As a second, independent mechanism of metal ion recognition, all analyzed metal-specific TCR showed a common overrepresentation of a histidine in the complementarity determining region 3 (CDR3; 15% of α-chains, 34% of ß-chains). The positions of the CDR3 histidine among metal-specific TCR mirrored those in random repertoires and were conserved among cross-reactive clonotypes. CONCLUSIONS: Induced CD154 expression allows a fast and comprehensive detection of Ni2+ -, Co2+ -, and Pd2+ -specific CD4+ T cells. Distinct TCR repertoire features underlie the frequent activation and cross-reactivity of human metal-specific T cells.

Oral nicotine pouches with an aftertaste? Part 2: in vitro toxicity in human gingival fibroblasts.

Nicotine pouches contain fewer characteristic toxicants than conventional tobacco products. However, the associated risks in terms of toxicity and addiction potential are still unclear. Therefore, endpoints of toxicity and contents of flavoring substances were investigated in this study. The in vitro toxicity of five different nicotine pouches and the reference snus CRP1.1 were studied in human gingival fibroblasts (HGF-1). Cells were exposed to product extracts (nicotine contents: 0.03-1.34 mg/mL) and sampled at different time points. Cytotoxicity, total cellular reactive oxygen species (ROS) levels, and changes in the expression levels of inflammatory and oxidative stress genes were assessed. Flavor compounds used in the nicotine pouches were identified by GC-MS. Cytotoxicity was observed in two nicotine pouches. Gene expression of interleukin 6 (IL6) and heme oxygenase 1 (HMOX1) was upregulated by one and three pouches, respectively. ROS production was either increased or decreased, by one pouch each. CRP1.1 caused an upregulation of IL6 and elevated ROS production. Toxicity was not directly dependent on nicotine concentration and osmolarity. A total of 56 flavorings were detected in the five nicotine pouches. Seven flavorings were classified according to the harmonized hazard classification system as laid down in the European Classification, Labelling and Packaging regulation. Nine flavorings are known cytotoxins. Cytotoxicity, inflammation, and oxidative stress responses indicate that adverse effects such as local lesions in the buccal mucosa may occur after chronic product use. In conclusion, flavorings used in nicotine pouches likely contribute to the toxicity of nicotine pouches.

Oral nicotine pouches with an aftertaste? Part 1: screening and initial toxicological assessment of flavorings and other ingredients.

Nicotine pouches are oral products that deliver nicotine without containing tobacco. Previous studies mainly focused on the determination of known tobacco toxicants, while yet no untargeted analysis has been published on unknown constituents, possibly contributing to toxicity. Furthermore, additives might enhance product attractiveness. We therefore performed an aroma screening with 48 different nicotine-containing and two nicotine-free pouches using gas chromatography coupled to mass spectrometry, following acidic and basic liquid-liquid extraction. For toxicological assessment of identified substances, European and international classifications for chemical and food safety were consulted. Further, ingredients listed on product packages were counted and grouped by function. Most abundant ingredients comprised sweeteners, aroma substances, humectants, fillers, and acidity regulators. 186 substances were identified. For some substances, acceptable daily intake limits set by European Food Safety Agency (EFSA) and Joint FAO/WHO Expert Committee on Food Additives are likely exceeded by moderate pouch consumption. Eight hazardous substances are classified according to the European CLP regulation. Thirteen substances were not authorized as food flavorings by EFSA, among them impurities such as myosmine and ledol. Three substances were classified by International Agency for Research on Cancer as possibly carcinogenic to humans. The two nicotine-free pouches contain pharmacologically active ingredients such as ashwagandha extract and caffeine. The presence of potentially harmful substances may point to the need for regulation of additives in nicotine-containing and nicotine-free pouches that could be based on provisions for food additives. For sure, additives may not pretend positive health effects in case the product is used.

Artificial intelligence and machine learning disciplines with the potential to improve the nanotoxicology and nanomedicine fields: a comprehensive review.

The use of nanomaterials in medicine depends largely on nanotoxicological evaluation in order to ensure safe application on living organisms. Artificial intelligence (AI) and machine learning (MI) can be used to analyze and interpret large amounts of data in the field of toxicology, such as data from toxicological databases and high-content image-based screening data. Physiologically based pharmacokinetic (PBPK) models and nano-quantitative structure-activity relationship (QSAR) models can be used to predict the behavior and toxic effects of nanomaterials, respectively. PBPK and Nano-QSAR are prominent ML tool for harmful event analysis that is used to understand the mechanisms by which chemical compounds can cause toxic effects, while toxicogenomics is the study of the genetic basis of toxic responses in living organisms. Despite the potential of these methods, there are still many challenges and uncertainties that need to be addressed in the field. In this review, we provide an overview of artificial intelligence (AI) and machine learning (ML) techniques in nanomedicine and nanotoxicology to better understand the potential toxic effects of these materials at the nanoscale.