Skin sensitisation prediction using read-across, an illustrative next generation risk assessment (NGRA) case study for vanillin.

Skin sensitisation is a key adverse human health effect to be addressed in the safety assessment of cosmetic ingredients. Regulatory demands and scientific progress have led to the development of a Next Generation Risk Assessment (NGRA) framework, relying on the use of New Approach Methodologies (NAM) Defined Approaches (DA) and read-across instead of generating animal data. This case study illustrates the application of read-across for the prediction of the skin sensitisation potential of vanillin at the hypothetical use concentration of 0.5% in a shower gel and face cream. A three-step process was applied to select the most suitable analogues based on their protein reactivity, structural characteristics, physicochemical properties, skin metabolism profile and availability of skin sensitisation data. The applied read-across approach predicted a weak skin sensitiser potential for vanillin corresponding with a Local Lymph Node Assay EC3 value of 10%. Based on this EC3 value a point of departure of 2500 µg/cm2 was derived, resulting in an acceptable exposure level (AEL) of 25 µg/cm2. Because the consumer exposure levels (CEL) for the face cream (13.5 µg/cm2) and shower gel (0.05 µg/cm2) scenarios were lower than the AEL, the NGRA concluded both uses as safe.

In vivo toxicity assessment of eugenol and vanillin-functionalised silica particles using Caenorhabditis elegans.

The toxicological properties of different silica particles functionalised with essential oil components (EOCs) were herein assessed using the in vivo model C. elegans. In particular, the effects of the acute and long-term exposure to three silica particle types (SAS, MCM-41 micro, MCM-41 nano), either bare or functionalised with eugenol or vanillin, were evaluated on different biological parameters of nematodes. Acute exposure to the different particles did not reduce nematodes survival, brood growth or locomotion, but reproduction was impaired by all the materials, except for vanillin-functionalised MCM-41 nano. Moreover, long-term exposure to particles led to strongly inhibited nematodes growth and reproduction. The eugenol-functionalised particles exhibited higher functionalisation yields and had the strongest effects during acute and long-term exposures. Overall, the vanillin-functionalised particles displayed milder acute toxic effects on reproduction than pristine materials, but severer toxicological responses for the 96-hour exposure assays. Our findings suggest that the EOC type anchored to silica surfaces and functionalisation yield are crucial for determining the toxicological effects of particles on C. elegans. The results obtained with this alternative in vivo model can help to anticipate potential toxic responses to these new materials for human health and the environment.

Effects of essential oil components exposure on biological parameters of Caenorhabditis elegans.

The extensive use of essential oil components in an increasing number of applications can substantially enhance exposure to these compounds, which leads to potential health and environmental hazards. This work aimed to evaluate the toxicity of four widely used essential oil components (carvacrol, eugenol, thymol, vanillin) using the in vivo model Caenorhabditis elegans. For this purpose, the LC50 value of acute exposure to these components was first established; then the effect of sublethal concentrations on nematodes' locomotion behaviour, reproduction, heat and oxidative stress resistance and chemotaxis was evaluated. The results showed that all the components had a concentration-dependent effect on nematode survival at moderate to high concentrations. Carvacrol and thymol were the two most toxic compounds, while vanillin had the mildest toxicological effect. Reproduction resulted in a more sensitive endpoint than lethality to evaluate toxicity. Only pre-exposure to carvacrol and eugenol at the highest tested sublethal concentrations conferred worms oxidative stress resistance. However, at these and lower concentrations, both components induced reproductive toxicity. Our results evidence that these compounds can be toxic at lower doses than those required for their biological action. These findings highlight the need for a specific toxicological assessment of every EOC application.

Sex-related deposition and metabolism of vanisulfane, a novel vanillin-derived pesticide, in rats and its hepatotoxic and gonadal effects.

A series of vanillin derivatives have recently been synthesized as effective candidate antiviral agents, with vanisulfane exhibiting pronounced curative and protective activities against cucumber mosaic virus and potato virus Y. However, research on some new pesticides usually ignores their various metabolites and sex-related toxicity. Assisted by 14C labeling, a trial was conducted to investigate the tissue distribution, excretion, and metabolism of vanisulfane in male and female rats for the first time. The results showed that 83.30-87.51% of applied 14C activity was excreted in urine and feces within 24 h of oral administration, and 14C was most abundant in the liver and kidney in both sexes. Interestingly, sex differences were observed in the experiment, with lower body clearance in males than in females 24 h after treatment and preferences for biliary and renal excretion of the pesticide in male and female rats, respectively. A high degradation rate was found for vanisulfane in the plasma; thus, the metabolites of vanisulfane were investigated using liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) combined with 14C labeling. One glucuronic acid conjugate and two oxidation metabolites were detected, supporting the monitoring of vanisulfane in vivo. Additionally, rats exposed to vanisulfane exhibited hepatic steatosis in both sexes, along with mild gonadal effects in males. This research offers an effective method for conducting environmental behavioral research and provides new insights for evaluating the potential risks of novel pesticides in mammals from a sex perspective.

A benzaldehyde-indole fused chromophore-based fluorescent probe for double-response to cyanide and hypochlorite in living cells.

With the rapid development of various industries, cyanide (CN-) and hypochlorite (ClO-) have a tremendously adverse effect on the health of humans and animals. In this study, a fluorescent probe HHTB based on a benzaldehyde-indole fused chromophore was designed to detect cyanide and hypochlorite simultaneously. The synthesized probe was found to have strong anti-interference ability. In addition, the designed probe could respond rapidly to ClO- in just 80 s, while the color changed visibly from red to colorless. Moreover, the response time to CN- was longer (about 160 s), with the apparent color change from red to light red. The ratiometric and colorimetric absorbance variation of HHTB was due to the nucleophilic attack of CN- on the indole CN functional group and the strong oxidization of ClO- which destroyed the CC bonds and the conjugation systems. Furthermore, the probe HHTB responding to ClO- and CN- presented high sensitivity, as the calculated detection limits were 1.18 nM and 1.40 nM, respectively. The probe was also found to have low biological toxicity and was used in living cells successfully. Therefore, it has good application prospect in the field of cell imaging and biomedicine. The binding mechanism of HHTB-CN and the reaction mechanism of HHTB and ClO- were further elucidated by a series of experiments.

Dihydropyriculol produced by Pyricularia oryzae inhibits the growth of Streptomyces griseus.

Dihydropyriculol is a major secondary metabolite of Pyricularia oryzae. However, the biological activity of dihydropyriculol has not been reported. Here, we showed that dihydropyriculol has inhibitory activity against Streptomyces griseus. Localization analysis of dihydropyriculol revealed that dihydropyriculol could reach to S. griseus under confrontation culture. These results suggest that dihydropyriculol can be used as a chemical weapon against S. griseus.

RIFM fragrance ingredient safety assessment, p-tolualdehyde, CAS Registry Number 104-87-0.

The existing information supports the use of this material as described in this safety assessment. p-Tolualdehyde was evaluated for genotoxicity, repeated dose toxicity, developmental and reproductive toxicity, local respiratory toxicity, phototoxicity, skin sensitization potential, and environmental safety. Data from read-across analog benzaldehyde (CAS # 100-52-7) show that p-tolualdehyde is not expected to be genotoxic. Data from read-across analog cuminaldehyde (CAS # 122-03-2) provided p-tolualdehyde a No Expected Sensitization Induction Level (NESIL) of 1100 µg/cm2 for the skin sensitization endpoint. The repeated dose toxicity, developmental and reproductive toxicity, and local respiratory toxicity endpoints were completed using the threshold of toxicological concern (TTC) for a Cramer Class I material, and the exposure to p-tolualdehyde is below the TTC (0.03 mg/kg/day, 0.03 mg/kg/day, and 1.4 mg/day, respectively). The phototoxicity/photoallergenicity endpoints were evaluated based on data from read-across analog 4-ethylbenzaldehyde (CAS # 4748-78-1); p-tolualdehyde is not expected to be phototoxic/photoallergenic. The environmental endpoints were evaluated; p-tolualdehyde was found not to be persistent, bioaccumulative, and toxic (PBT) as per the International Fragrance Association (IFRA) Environmental Standards, and its risk quotients, based on its current volume of use in Europe and North America (i.e., Predicted Environmental Concentration/Predicted No Effect Concentration [PEC/PNEC]), are <1.

Are atranols the only skin sensitizers in oakmoss? A systematic investigation using non-animal methods.

Oakmoss and treemoss absolutes are the major natural extracts of concern as potential sources of skin sensitizers in cosmetics and personal care products (PCP). Two single constituents, atranol and chloroatranol, have been identified as primary culprits in both lichens, and industrial self-regulation has been proposed to limit their contents to less than 100 ppm. Nonetheless, evidence points to the presence of additional candidate skin sensitizers in these multicomponent extracts. These observations, along with a lack of data from non-animal alternative methods and the chemical variability of commercial absolutes, prompted further investigation of oakmoss absolute along with altranol-like compounds in these extracts. The major chemical constituents of a commercial sample were identified by two independent analytical techniques, GC-MS and HPLC-DAD-MS. The crude oakmoss extract and pure compounds were assayed with two in chemico methods (HTS-DCYA and DPRA) to gauge their chemical reactivity. Activation of inflammatory responses in vitro was also investigated by KeratinoSens™ and human cell line activation tests (h-CLAT). Based on weight of evidence, orcinol, ethyl orsellinate, and usnic acid were classified as candidate sensitizers, along with both atranols and oakmoss extract.

Comparative cytotoxic study of silica materials functionalised with essential oil components in HepG2 cells.

This work evaluated the cytotoxic effect of different EOCs-functionalised silica particle types. The in vitro toxicity of eugenol and vanillin-immobilised SAS, MCM-41 microparticles and MCM-41 nanoparticles was evaluated on HepG2 cells, and compared to free EOCs and pristine materials. The results revealed that free essential oil components and bare silica had a mild cytotoxic effect on HepG2 cells. However, the comparative study showed that free eugenol and vanillin had a milder cytotoxic effect than the equivalent concentrations of immobilised components on the different silica particles, while differences in cell viability between the bare and functionalised particles relied on the type of analysed material. The most cytotoxic materials were eugenol and vanillin-functionalised MCM-41 micro with IC50 values of 0.19 and 0.17 mg/mL, respectively, at 48 h exposure. Differences in cytotoxicity between functionalised particles may be attributed to the density of the functional components on their surface as a result of the functionalisation reaction performance for different materials. The study of the physico-chemical properties of particles demonstrated that cationic nature and increased hydrophobicity could be responsible for promoting cell-particle interactions for the eugenol and vanillin functionalised silica particles, enhancing their cytotoxic behaviour.

2-Hydroxy-4-methoxybenzaldehyde from Hemidesmus indicus is antagonistic to Staphylococcus epidermidis biofilm formation.

Staphylococcus epidermidis (SE) is an opportunistic nosocomial pathogen that accounts for recalcitrant device-related infections worldwide. Owing to the growing interest in plants and their secondary metabolites targeting bacterial adhesion, this study was intended to uncover the anti-biofilm potential of Hemidesmus indicus and its major constituent 2-hydroxy-4-methoxybenzaldehyde (HMB) against SE. The minimum biofilm inhibitory concentration (MBIC) of H. indicus root extract and HMB were found to be 500 and 250 µg ml-1, respectively. The results of time-dependent biofilm inhibition and mature biofilm disruption assays confirmed that HMB targets initial cell adhesion. Furthermore, interference by HMB in the expression of adhesin genes (icaA, aap and bhp) and biofilm components was associated with an increased susceptibility of SE to oxidative stress and antibiotics. To conclude, this study reports for the first time HMB as a potential drug against SE biofilms.

Cuminaldehyde potentiates the antimicrobial actions of ciprofloxacin against Staphylococcus aureus and Escherichia coli.

Escherichia coli and Staphylococcus aureus are important agents of urinary tract infections that can often evolve to severe infections. The rise of antibiotic-resistant strains has driven the search for novel therapies to replace the use or act as adjuvants of antibiotics. In this context, plant-derived compounds have been widely investigated. Cuminaldehyde is suggested as the major antimicrobial compound of the cumin seed essential oil. However, this effect is not fully understood. Herein, we investigated the in silico and in vitro activities of cuminaldehyde, as well as its ability to potentiate ciprofloxacin effects against S. aureus and E. coli. In silico analyses were performed by using different computational tools. The PASS online and SwissADME programmes were used for the prediction of biological activities and oral bioavailability of cuminaldehyde. For analysis of the possible toxic effects and the theoretical pharmacokinetic parameters of the compound, the Osiris, SwissADME and PROTOX programmes were used. Estimations of cuminaldehyde gastrointestinal absorption, blood brain barrier permeability and skin permeation by using SwissADME; and drug likeness and score by using Osiris, were also evaluated The in vitro antimicrobial effects of cuminaldehyde were determined by using microdilution, biofilm formation and time-kill assays. In silico analysis indicated that cuminaldehyde may act as an antimicrobial and as a membrane permeability enhancer. It was suggested to be highly absorbable by the gastrointestinal tract and likely to cross the blood brain barrier. Also, irritative and harmful effects were predicted for cuminaldehyde if swallowed at its LD50. Good oral bioavailability and drug score were also found for this compound. Cuminaldehyde presented antimicrobial and anti-biofilm effects against S. aureus and E. coli.. When co-incubated with ciprofloxacin, it enhanced the antibiotic antimicrobial and anti-biofilm actions. We suggest that cuminaldehyde may be useful as an adjuvant therapy to ciprofloxacin in S. aureus and E. coli-induced infections.

Antinociceptive and antineuropathic effects of cuminaldehyde, the major constituent of Cuminum cyminum seeds: Possible mechanisms of action.

ETHNOPHARMACOLOGICAL RELEVANCE: In Iranian traditional medicine, Cuminum cyminum is a unique medicinal herb for pain relief. Cuminaldehyde has been distinguished as the major constituent of C. cyminum seeds; even though, the analgesic effect of cuminaldehyde has not yet been examined. AIM OF THE STUDY: The nobility of this study was to assess cuminaldehyde effect on nociceptive and neuropathic pains; furthermore, evaluation of its possible mechanisms of action. MATERIALS AND METHODS: Hot plate, formalin, and acetic acid-induced writhing tests were used to evaluate nociception in mice. Naloxone (opioid receptors antagonist), L-arginine (nitric oxide (NO) precursor), L-NAME (NO synthase inhibitor), sodium nitroprusside (NO donor), methylene blue (guanylyl cyclase inhibitor), sildenafil (phosphodiesterase inhibitor), and glibenclamide (KATP channel blocker) were used to determine the implication of opioid receptors and L-arginine/NO/cGMP/KATP channel pathway. Allodynia and hyperalgesia were investigated in the CCI (chronic constriction injury) model of neuropathic pain in rats. The ELISA method was used to measure the inflammatory cytokines in serum samples of rats. The entire chemicals were intraperitoneally injected. RESULTS: Cuminaldehyde (100 and 200 mg/kg) significantly decreased the latency to nociceptive response in the hot plate test. The outcome of cuminaldehyde was completely antagonized by naloxone (2 mg/kg). Formalin- and acetic acid-induced nociception was significantly inhibited by cuminaldehyde (12.5-50 mg/kg). The antinociceptive effect of cuminaldehyde was reversed in writhing test by L-arginine (200 mg/kg), sodium nitroprusside (0.25 mg/kg), and sildenafil (0.5 mg/kg); however, L-NAME (30 mg/kg) and methylene blue (20 mg/kg) enhanced the effect of cuminaldehyde. Glibenclamide (10 mg/kg) did not alter the antinociceptive effects of cuminaldehyde. In the CCI-induced neuropathy, cuminaldehyde (25-100 mg/kg) significantly alleviated allodynia and hyperalgesia and decreased the serum levels of TNF-α and IL-1ß. CONCLUSION: It was attained magnificently that cuminaldehyde exerts antinociceptive and antineuropathic effects through the involvement of opioid receptors, L-arginine/NO/cGMP pathway, and anti-inflammatory function.

Bioactivity of some Apiaceae essential oils and their constituents against Sitophilus zeamais (Coleoptera: Curculionidae).

Sitophilus zeamais is a key pest of stored grains. Its control is made, usually, using synthetic insecticides, despite their negative impacts. Botanical insecticides with fumigant/repellent properties may offer an alternative solution. This work describes the effects of Anethum graveolens, Petroselinum crispum, Foeniculum vulgare and Cuminum cyminum essential oils (EOs) and (S)-carvone, cuminaldehyde, estragole and (+)-fenchone towards adults of S. zeamais. Acute toxicity was assessed by fumigation and topical application. Repellence was evaluated by an area preference bioassay and two-choice test, using maize grains. LC50 determined by fumigation ranged from 51.8 to 535.8 mg L-1 air, with (S)-carvone being the most active. LD50 values for topical applications varied from 23 to 128 µg per adult for (S)-carvone > cuminaldehyde > A. graveolens > C. cyminum > P. crispum. All EOs/standard compounds reduced significantly the percentage of insects attracted to maize grains (65-80%) in the two-choice repellence test, whereas in the area preference bioassay RD50 varied from 1.4 to 45.2 µg cm-2, with cuminaldehyde, (S)-carvone and estragole being strongly repellents. Petroselinum crispum EO and cuminaldehyde affected the nutritional parameters relative growth rate, efficiency conversion index of ingested food and antifeeding effect, displaying antinutritional effects toward S. zeamais. In addition, P. crispum and C. cyminum EOs, as well as cuminaldehyde, showed the highest acetylcholinesterase inhibitory activity in vitro (IC50 = 185, 235 and 214.5 µg mL-1, respectively). EOs/standard compounds exhibited acute toxicity, and some treatments showed antinutritional effects towards S. zeamais. Therefore, the tested plant products might be good candidates to be considered to prevent damages caused by this pest.

Quantification of selected aroma compounds in e-cigarette products and toxicity evaluation in HUVEC/Tert2 cells.

During recent years, the number of consumers using so-called e-cigarettes, which are electrical devices to aerosolize a liquid consisting of propylene glycol, glycerol, optional nicotine and flavoring chemicals, has been increasing. Aromas vary from common flavors such as mint to more unusual flavors such as buttermilk or pepperoni pizza. Consumers today can buy e-concentrates that consist of propylene glycol and aroma to blend their own desired flavor at home. Little is known about the composition and concentration of various aroma molecules in the different e-liquids and e-concentrates. In addition, the process of EU-wide regulation is still ongoing. The aim of this research study was to identify and quantify possible undesirable aroma compounds in e-liquids and e-concentrates. Flavoring chemicals such as estragole, benzaldehyde and cinnamaldehyde were quantified. The measurements were carried out on a GC-MS system. The results show the presence of highly concentrated flavoring compounds and limonene oxide in lemon-flavored e-concentrates. In the final step, samples and single-aroma standards were tested for their toxicity to HUVEC/Tert2 cells, where some single-flavoring chemicals such as cinnamic aldehyde revealed significant toxic effects.

Comparison of the metabolism of 10 cosmetics-relevant chemicals in EpiSkin™ S9 subcellular fractions and in vitro human skin explants.

An understanding of the bioavailability of topically applied cosmetics ingredients is key to predicting their local skin and systemic toxicity and making a safety assessment. We investigated whether short-term incubations with S9 from the reconstructed epidermal skin model, EpiSkin™, would give an indication of the rate of chemical metabolism and produce similar metabolites to those formed in incubations with human skin explants. Both have advantages: EpiSkin™ S9 is a higher-throughput assay, while the human skin explant model represents a longer incubation duration (24 hours) model integrating cutaneous distribution with metabolite formation. Here, we compared the metabolism of 10 chemicals (caffeine, vanillin, cinnamyl alcohol, propylparaben, 4-amino-3-nitrophenol, resorcinol, 4-chloroaniline, 2-amino-3-methyl-3H-imidazo[4,5-F]quinoline and 2-acetyl aminofluorene) in both models. Both models were shown to have functional Phase 1 and 2 enzymes, including cytochrome P450 activities. There was a good concordance between the models with respect to the level of metabolism (stable vs. slowly vs. extensively metabolized chemicals) and major early metabolites produced for eight chemicals. Discordant results for two chemicals were attributed to a lack of the appropriate cofactor (NADP+ ) in S9 incubations (cinnamyl alcohol) and protein binding influencing chemical uptake in skin explants (4-chloroaniline). These data support the use of EpiSkin™ S9 as a screening assay to provide an initial indication of the metabolic stability of a chemical applied topically. If required, chemicals that are not metabolized by EpiSkin™ S9 can be tested in longer-term incubations with in vitro human explant skin to determine whether it is slowly metabolized or not metabolized at all.