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Herein is delineated a first systematic framework for the definition of structure‐antioxidant property relationships in the dihydroxynaphthalene (DHN) series. The results obtained by a combined experimental and theoretical approach revealed that 1,8‐DHN is the best performing antioxidant platform, with its unique hydrogen‐bonded peri‐hydroxylation pattern contributing to a fast H atom transfer process. Moreover, the comparative analysis of the antioxidant properties of DHNs carried out by performing DPPH and FRAP assays and laser flash photolysis experiments, revealed the higher antioxidant power associated with an α‐substitution pattern (i. e. in 1,8‐ and 1,6‐DHN) with respect to DHNs exhibiting a β‐substitution pattern (i. e. in 2,6‐ and 2,7‐DHN). DFT calculations and isolation and characterization of the main oligomer intermediates formed during the oxidative polymerization of DHNs supported this evidence by providing unprecedented insight into the generation and fate of the intermediate naphthoxyl radicals, which emerged as the main factor governing the antioxidant activity of DHNs. [ABSTRACT FROM AUTHOR]

Formation and structural modification of oxygenated polycyclic aromatic hydrocarbons (oxyPAHs) by UV irradiation on minerals have recently been proposed as a possible channel of PAH transformation in astrochemical and prebiotic scenarios of possible relevance for the origin of life. Herein, it is demonstrated that high‐energy proton‐beam irradiation in the presence of various meteorites, including stony iron, achondrite, and chondrite types, promotes the conversion of two representative oxyPAH compounds, 1‐naphthol and 1,8‐dihydroxynaphthalene, to complex mixtures of oxygenated and oligomeric derivatives. The main identified products include polyhydroxy derivatives, isomeric dimers encompassing benzofuran and benzopyran scaffolds, and, notably, a range of quinones and perylene derivatives. Addition of urea, a prebiotically relevant chemical precursor, expanded the range of identified species to include, among others, quinone diimines. Proton‐beam irradiation of oxyPAH modulated by nitrogen‐containing compounds such as urea is proposed as a possible contributory mechanism for the formation and processing of insoluble organic matter in meteorites and in prebiotic processes. [ABSTRACT FROM AUTHOR]

Developing effective strategies to increase the chemical stability and to fine‐tune the physico‐chemical properties of melanin biopolymers by rational control of π‐electron conjugation is an important goal in materials science for biomedical and technological applications. Herein we report that poly‐1,8‐dihydroxynaphthalene (pDHN), a non‐nitrogenous, catechol‐free fungal melanin mimic, displays a high degree of structural integrity (from MALDI‐MS and CP/MAS 13C NMR analysis), a strong radical scavenging capacity (DPPH and FRAP assays), and an unusually intense EPR signal (g=2.0030). Morphological and spectral characterization of pDHN, along with deassembly experiments in ionic liquids, indicated amorphous aggregates of small globular structures with an estimated stacking distance of 3.9 Å and broadband absorption throughout the visible range. These results indicate that DHN‐based melanins exhibit a high structural integrity and enhanced antioxidant and free‐radical properties of potentially greater biomedical and technological relevance than for typical indole‐based eumelanins. [ABSTRACT FROM AUTHOR]

Despite the growing scientific and technological relevance of polydopamine (PDA), a eumelanin-like adhesive material widely used for surface functionalization and coating, knowledge of its structural and physicochemical properties, including in particular the origin of paramagnetic behavior, is still far from being complete. Herein, we disclose the unique ability of ionic liquids (ILs) to disassemble PDA, either as a suspension or as a thin film, up to the nanoscale, and to establish specific interactions with the free radical centers exposed by deaggregation of potential investigative value. Immersion of PDA-coated glasses into four different ILs ([C1C1im][(CH3O)HPO2], [C1C1im][(CH3O)CH3PO2], [C1C1im][(CH3O)2PO2], [N1888][C18:1]) at room temperature caused the fast and virtually complete removal of the coating as determined by UV-visible spectroscopy and scanning electron microscopy (SEM). Transmission electron microscopy (TEM) analysis of the colored supernatants from PDA suspensions in ILs revealed the presence of nanostructures not exceeding 50 nm in diameter. Electron paramagnetic resonance (EPR) analysis indicated profound IL-dependent modifications in signal intensity, line-width, and g-factor values of PDA. These differences were interpreted in terms of a partial conversion of C-centered radicals into O-centered semiquinone-type components following destacking and interaction with the anion component in ILs. The discovery of ILs as a powerful tool to disassemble PDA under mild conditions provides a new entry both to detailed investigations of this biopolymer on the nanoscale and to mild removal of coatings from functionalized surfaces, greatly expanding the scope of PDA-based surface functionalization strategies. [ABSTRACT FROM AUTHOR]

The oxidative polymerization of 5,6-dihydroxyindoles and related hydroxyindoles at pH<3 is diverted from the usual eumelanin-forming pathway to produce mixtures of symmetric and asymmetric triazatruxenes (TATs), which could be separated and characterized for their opto-electronic properties with the aid of TD-DFT calculations. Data showed that the asymmetric isomers exhibit higher fluorescence quantum efficiencies, lower HOMO-LUMO gaps, better film homogeneity, and a more definite aggregation behavior than the symmetric counterparts, suggesting promising applications in organic electronics. The enhanced luminance exhibited by the OLED devices fabricated with blends of the synthesized TATs in poly-9-vinylcarbazole confirmed the potential of the asymmetric skeleton as new versatile platform for light-emitting materials. [ABSTRACT FROM AUTHOR]

5,6-Dihydroxyindole ( 1) and its N-methyl derivative ( 2), key eumelanin building blocks, were inserted into zeolite L by sublimation at 175 °C for 5 days. At a 10 mg/300 mg indole/zeolite ratio, the resulting hybrids displayed a stable deep red coloration. CP/MAS 13C NMR and UV/Vis spectroscopy of the red species suggested the generation and accommodation of quinonoid biindole derivative(s) within the void space of the acidic zeolite channels. Removal of the zeolite matrix by treatment with HF gave a stable species that could be separated by HPLC and characterized by mass spectrometry as an oxygenated biindole derivative (or a mixture of isomers), suggesting addition of water to the original dimer and subsequent re-oxidation. The characterization was corroborated by optimized molecular geometries and simulated UV spectra with density functional calculations. Loading 1 or 2 into the larger pores of SBA-15 type mesoporous silica resulted in black eumelanin-type polymers, confirming channel size dependence over the polymerization process. [ABSTRACT FROM AUTHOR]

The 'Haptenation theory' concerns the multicausal pathogenesis of vitiligo ending ultimately in the (partial) disappearance of melanocytes from the skin and/or hairs. The melanocyte specificity is attributed to the tyrosinase-catalysed production of haptogenic ortho-quinones that covalently bind to tyrosinase or other melanosomal proteins to generate neo-antigens. These latter, in turn, trigger an immunological cascade resulting in a melanocyte-specific delayed-type hypersensitivity reaction that eliminates melanocytes and produces the characteristic depigmentation. This causal chain of events is critically discussed with special reference to factors modifying the process and the possible influence of various biochemical changes, such as raised levels of catecholamines and epidermal hydrogen peroxide, which have been reported to be associated with the onset of vitiligo. This all adds up to the typical vitiligo reaction pattern or syndrome, which demands a treatment strategy involving most of the already known therapies. Similar pathogenetic mechanisms might be engaged in the enhancement of cellular immunity (vaccination) against melanoma. [ABSTRACT FROM AUTHOR]

Abstract: We present the application of a novel isotope dilution method, named Alternate Isotope-Coded Derivatization Assay (AIDA), to the quantitative analysis of hydrazone derivatives of malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) in exhaled breath condensate (EBC) samples using liquid chromatography–tandem mass spectrometry. AIDA is based on the alternate derivatization of the analyte(s) with reagents that are available in two pure isotopic forms, respectively “light” and “heavy”, by using light-derivatized standards for the quantification of the heavy-derivatized analytes, and vice versa. To this purpose, 2,4-dinitro-3,5,6-trideuterophenylhydrazine (d 3 -DNPH) has been synthesized and used as “heavy” reagent in combination with commercial “light” DNPH. Using the AIDA method, any unknown concentration of the analyte in the matrix can be calculated without the need of a calibration curve. An external calibration method has been also investigated for comparative purpose. The stability of DNPH and d 3 -DNPH derivatives was verified by excluding any exchange of hydrazones with each other. In the range of concentrations of biological interest, e.g., 2–40nM for MDA and 0.5–10nM for 4-HNE, the derivatization reactions of MDA and 4-HNE with DNPH and d 3 -DNPH showed overlapping kinetics and comparable yields. The MS response of both DNPH and d 3 -DNPH hydrazones was similar. The precision of AIDA, calculated as %RSD, was within 3.2–8% for MDA and 4.5–11% for 4-HNE. Accuracy was tested by analyzing a spiked EBC pool sample and acceptable results (accuracy within 98–108% for MDA and 93–114% for 4-HNE) were obtained by AIDA after subtraction of the blank, which was not negligible. The results of quantitative analysis of MDA and 4-HNE in EBC samples obtained by AIDA assay with four analyses per sample were in good agreement with those obtained by external calibration method on the same samples. [Copyright &y& Elsevier]

We developed a new method for the simultaneous quantitative determination of 8-oxo-7,8-hydro-2′-deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydroguanine (8-oxoGua), 8-oxo-7,8-dihydroguanosine (8-oxoGuo), and the corresponding non-oxidized forms, 2′-deoxyguanosine (dGuo), guanine (Gua) and guanosine (Guo), in human urine samples by liquid chromatography–tandem mass spectrometry. Differences in the ionization of analytes in different urine batches with variable matrix effects were effectively compensated for by internal standardization with stable isotope-labelled analytes. The method was sensitive enough to allow the determination of background levels of these biomarkers and was applied to characterize the inter- and intraindividual variability of biomarkers in the diurnal profile of concentrations in 24 healthy volunteers. When normalized for creatinine, none of the biomarkers was affected by sampling time, thus ruling out any circadian rhythm for nucleic acid oxidation in urine. [ABSTRACT FROM AUTHOR]

Abstract: This study investigated nucleic acid oxidation associated with exposure to benzene at low levels in 239 workers recruited among traffic policemen, taxi drivers and gasoline pump attendants of the city of Parma (Italy). Biomarkers of exposure, namely urinary t,t-muconic acid (t,t-MA) and S-phenylmercapturic acid (S-PMA), urinary cotinine, and urinary biomarkers of nucleic acid oxidation, namely 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo), 8-oxo-7,8-dihydroguanosine (8-oxoGuo) and 8-oxo-7,8-dihydroguanine (8-oxoGua) were determined by liquid chromatography–tandem mass spectrometry. Relevant polymorphisms of NAD(P)H:quinone oxidoreductase (NQO1), glutathione S-transferases M1-1 (GSTM1), T1-1 (GSTT1), and A1 (GSTA1) were characterized by polymerase chain reaction-based methods in a subgroup of subjects. Biomarkers of nucleic acid oxidation were correlated with each other (r ≥0.32, p <0.0001) and with exposure biomarkers (r ≥0.28, p <0.0001). Multiple linear regression models including age, sex and smoking habits as independent variables demonstrated that benzene exposure is associated with oxidation damage to nucleic acid, particularly to RNA (p <0.0001) and is modulated by the NQO1 polymorphism. The study confirmed a significant modulating effect of GSTM1 (p =0.010), GSTT1 (p =0.023) and GSTA1 (p =0.048) polymorphisms on S-PMA excretion, with a significant interaction between GSTM1 and both GSTT1 and GSTA1 (p =0.006 and p =0.037, respectively). [Copyright &y& Elsevier]

To evaluate a combined biomonitoring approach based on both cobalt and tungsten determination in workers of the hard metal alloy sector. We enrolled 55 workers from a factory producing cutting tools for carpentry. Combined workroom air and biological monitoring of both cobalt and tungsten relied on inductively coupled plasma mass spectrometry determinations. Metals were determined on plasma, blood and urine samples. Urine samples from 34 unexposed subjects were also analyzed. Tungsten was determined in every collected sample. Workers showed significantly higher urinary tungsten levels than controls (pre-shift values of 4.12 vs. 0.06 μg/l on average; P < 0.0005). Both airborne and biological levels of tungsten prevailed among workers involved in wet-grinding activities. The element was excreted at higher urinary levels than cobalt and showed lower circulating (blood, plasma) concentrations. Exposure–dose relationships were apparent for tungsten biomarkers. Obtained results may contribute to the development of biomarkers of exposure to tungsten. The association of such biomarkers to traditional determinations of cobalt in blood and/or urine may substantially improve the exposure assessment of workers employed in cemented carbide industries. [ABSTRACT FROM AUTHOR]

Abstract: This study investigated nucleic acid oxidation associated with styrene exposure, mRNA expression levels of hOGG1 gene and the role of the genetic polymorphism Ser326Cys of human 8-oxoguanine DNA N-glycosylase 1 (hOGG1) in 60 styrene-exposed workers and 50 unexposed clerks. Biomarkers of exposure (styrene in blood, mandelic and phenylglyoxylic acids and 4-vinylphenol in urine) and urinary biomarkers of nucleic acid oxidation, namely 8-oxo-7,8-dihydro-2′-deoxyguanosine (U-8-oxodGuo), 8-oxo-7,8-dihydroguanosine (U-8-oxoGuo) and 8-oxo-7,8-dihydroguanine (U-8-oxoGua) were determined by liquid chromatography–tandem mass spectrometry. The levels of 8-oxodGuo adduct and 2′-deoxyguanosine (dGuo) were measured by HPLC in DNA from white blood cells (WBC). Genomic DNA and RNA from blood samples were used to characterize the Ser326Cys polymorphism and the mRNA expression levels of the hOGG1 gene, respectively, by PCR-based methods. Exposed workers showed lower values of 8-oxodGuo/105 dGuo ratio in WBC-DNA but higher concentrations of U-8-oxoGuo compared to controls (p =0.002 and p =0.008, respectively, t-test for independent samples). In the whole group, all urinary biomarkers of nucleic acid oxidation correlated with both the sum of mandelic and phenylglyoxylic acids (rho >0.33, p <0.0001) and 4-vinylphenol (rho >0.29, p <0.001), whereas 8-oxodGuo/105 dGuo in WBC showed a negative correlation with exposure parameters (rho <−0.24, p <0.02). Subjects bearing the hOGG1 Ser/Ser genotype showed lower values of 8-oxodGuo/105 dGuo in WBC than those with at least one variant Cys allele (0.34±0.16 vs 0.45±0.21, p =0.008). In the subgroup of hOGG1 Ser/Ser subjects, laminators showed lower levels of WBC 8-oxodGuo/105 dGuo ratio and significantly higher concentrations of U-8-oxoGua than controls (p =0.07 and p =0.01, respectively, t-test for independent samples). Interestingly, workers showed higher levels of hOGG1 expression compared to controls (p <0.0005). Styrene exposure seems to be associated with oxidation damage to nucleic acids, particularly to RNA and with an induction of the BER system. [Copyright &y& Elsevier]

A unified convenient strategy for the synthesis of 5,6-dihydroxyindole-derived 2,7'-, 2,2'-, and 2,3'-biindolyls is reported, which is based on proper manipulation of key o-ethynylaniline precursors. By the same methodology 5,6- diacetoxy-7-iodoindole can also be obtained in good yield. [ABSTRACT FROM AUTHOR]

The article discusses the chemistry and metabolic fate of nitrated lipids which have remained badly elucidated despite of the mounting interest in nitrolinoleic acids and related polynsaturated fatty acids as a new class of bioactive signaling lipids. Accordingly, nitrated derivatives of unsaturated fatty acids were discovered as significant products of lipid modification under physiological conditions colligated with oxidative stress and lifted productions of nitric oxide (NO).

Abstract: A comparative evaluation of urinary biomarkers was carried out to characterize benzene exposure in a group of 100 traffic policemen of the city of Parma (Italy). All subjects were monitored once, in two consecutive days characterized by similar climatic conditions but preceded by two windy days. Benzene ambient concentration measured by municipal air monitoring stations was 1μg/m3 (Day 1) and 2μg/m3 (Day 2). Personal exposure to ambient concentrations of benzene, toluene, ethylbenzene and xylene (BTEX) was assessed by using Radiello® passive-diffusive samplers in a subgroup of 24 workers. Benzene metabolites, t,t-muconic acid (t,t-MA) and S-phenylmercapturic acid (S-PMA) were determined by isotopic dilution liquid chromatography–tandem mass spectrometry on spot urine samples collected at the end of the shift. Urinary benzene (U-B) was determined by solid-phase microextraction gas chromatography–mass spectrometry. Airborne benzene concentration expressed as median [and interquartile range] was 6.07 [0.28–9.53]μg/m3, as assessed by personal sampling. Urinary concentrations of biomarkers in the whole group were 41.8 [34.1–89.8]μg/g creatinine for t,t-MA, 0.67 [0.23–1.32]μg/g creatinine for S-PMA, and 0.16 [0.13–0.26]μg/l for U-B. Smokers eliminated significantly higher concentrations of unchanged BTEX and benzene metabolites than non-smokers (p <0.05). When traffic policemen were distinguished into indoor (n =31) and outdoor workers, no significant differences were observed for either airborne benzene or urinary biomarkers. Significantly lower concentrations of S-PMA and U-B were determined in samples collected at Day 1 as compared to Day 2 (p <0.0001 and p =0.003, respectively) suggesting that these biomarkers are enough sensitive and specific to detect changes in airborne benzene concentration even at few μg/m3. [Copyright &y& Elsevier]

Abstract: Nitration of unsaturated fatty acids is a (patho)physiologically important pathway of lipid modification induced by nitric oxide-derived species. We report herein on the unexpected chain rearrangement undergone by (13S,9Z,11E)-13-hydroxyoctadeca-9,11-dienoic acid (1), a linoleic acid metabolite, when exposed to nitrating agents of biological relevance. At pH 7.4 and at room temperature, reaction of 1 with peroxynitrite (ONOO−) as well as Fe2+-EDTA/H2O2/NO2 − and horseradish peroxidase/H2O2/NO2 − led to the formation of two nitration products, which could be isolated as the methyl esters and were identified as diastereoisomeric methyl (12S)-10,11-epoxy-12-hydroxy-9-nitromethylheptadecanoates by extensive 1H, 13C, 15N NMR and MS analysis. [Copyright &y& Elsevier]

A procedure for analysis of melanin-pigmented tissues based on alkaline hydrogen peroxide degradation coupled with high-performance liquid chromatography (HPLC) ultraviolet determination of pyrrole-2,3,5-tricarboxylic acid (PTCA) for eumelanin and 6-(2-amino-2-carboxyethyl)-2-carboxy-4-hydroxybenzothiazole (BTCA) and 1,3-thiazole-2,4,5-tricarboxylic acid for pheomelanin was recently developed. Despite advantages related to the degradation conditions and sample handling, a decrease of the reproducibility and resolution was observed after several chromatographic runs. We report herein an improved chromatographic methodology for simultaneous determination of PTCA and BTCA as representative markers of eumelanin and pheomelanin, respectively, based on the use of an octadecylsilane column with polar end-capping with 1% formic acid (pH 2.8)/methanol as the eluant. The method requires conventional HPLC equipments and gives very good peak shapes and resolution, without need of ion pair reagents or high salt concentrations in the mobile phase. The intra-assay precision of the analytical runs was satisfactory with CV values ≤4.0% (n = 5) for the two markers which did not exceed 8% after 50 consecutive injections on the column over 1 week. The peak area ratios at 254 and 280 nm ( A280/ A254: PTCA = 1.1, BTCA = 0.6) proved a valuable parameter for reliable identification of the structural markers even in the most complex degradation mixtures. The method can be applied to various eumelanin and pheomelanin pigmented tissues, including mammalian hair, skin and irides, and is amenable to be employed in population screening studies. [ABSTRACT FROM AUTHOR]

Abstract: Biological monitoring (BM) and biomarkers are widely applied in occupational toxicology. BM is mainly aimed at (i) defining the existence of an occupational exposure; (ii) quantifying the level of internal dose; (iii) verifying that exposure limits (BEI®, BAT, BLV) are respected. As compared to ambient monitoring, BM is more expensive and complex. Several biomarkers are available for the same chemical and the meaning of the marker may depend on the sampling time. Therefore, practical issues, including cost and selection of an adequate sampling strategy, should be dealt with when planning a BM program for specific purposes. In addition, several biological and analytical sources of variability may influence biomarker levels, thus making the interpretation of BM data a difficult task. However, we should recognize that the main aim of BM is not to reduce, but to explain biological variance. The decreasing trend in occupational exposure levels highlighted the specificity problems of traditional biomarkers of exposure and prompted the research to the development of new biomarkers, e.g. unchanged volatile compounds in urine, minor metabolites, DNA and protein adducts. Depending on the scope and context (research or routine) different requirements of biomarkers can be envisaged in terms of validation and acceptable variability. [Copyright &y& Elsevier]

Abstract: An integrated approach based on ambient and biological monitoring, the latter including both biomarkers of exposure and susceptibility, was applied to characterize benzene exposure in a group of 37 taxi drivers of the city of Parma (Italy). Airborne benzene concentrations were assessed by 24h personal sampling and work-shift sampling inside the taxicab using passive samplers (Radiello®). Benzene metabolites, trans,trans-muconic acid (t,t-MA) and S-phenylmercapturic acid (S-PMA), and urinary cotinine as biomarker of smoking habits were measured by isotopic dilution liquid chromatography tandem mass spectrometry in both pre-shift (PS) and end-of-shift (EOS) samples. Urinary benzene (U-B) levels were determined by solid-phase microextraction gas chromatography–mass spectrometry in EOS samples. Relevant polymorphisms of microsomal epoxide hydrolase, NAD(P)H:quinone oxidoreductase, glutathione S-transferases M1-1 (GSTM1), T1-1, and A1 were characterized by PCR-based methods. Mean airborne benzene concentration was 5.85±1.65μg/m3, as assessed by 24h personal sampling integrating for work-shift, indoor or general environment activities. Significantly, higher benzene concentrations were detected in the taxicab during the work-shift (7.71±1.95μg/m3, p <0.005). Smokers eliminated significantly higher concentrations of U-B and S-PMA than non-smokers in EOS samples [geometric mean (geometric S.D.): 2.58 (4.23) versus 0.44 (1.79)μg/l for U-B; 3.79 (1.50) versus 2.14 (1.87)μg/gcreat. for S-PMA, p <0.002]. Within smokers, S-PMA concentrations significantly increased at the end of the work-shift compared to pre-shift values (p <0.05). t,t-MA showed a similar behaviour, although differences were not significant. In the narrow range examined, no correlation was observed between air benzene concentration and urinary biomarkers. All benzene biomarkers but EOS t,t-MA were correlated with U-cotinine (p <0.05). GSTM1 polymorphism significantly modulated S-PMA excretion, as subjects bearing the GSTM1pos genotype [3.61 (1.15)μg/gcreat.] excreted significantly higher S-PMA concentrations than GSTM1null subjects [2.19 (1.18)μg/gcreat., p <0.05]. [Copyright &y& Elsevier]

Abstract: In this paper, we provide a systematic analysis of glyoxal (1) formation from a range of monosaccharides and related compounds, to determine their potential role as sources of this α-oxoaldehyde in vivo. Substrates were reacted with the Fenton reagent (Fe2+/EDTA/H2O2) and the mixtures were analyzed by HPLC using the 6-hydroxy-2,4,5-triaminopyrimidine fluorimetric assay. The rank order of hexoses and their derivatives as glyoxal sources was found to be fructose>glucose=mannose=galactose>glucose-6-phosphate>mannitol. Within the pentose group, arabinose and ribose gave the higher yields of 1 followed by deoxyribose and its adenine N-glycosides and ribulose. Among the tested substrates, three-carbon compounds, that is, trioses and glycerol, but not glyceraldehyde-3-phosphate, were by far the most effective sources of 1. The effects of H2O2 and Fe2+/EDTA concentrations as well as of other metal ions were also investigated. [Copyright &y& Elsevier]

Abstract: An efficient one-pot procedure for the preparation of 10β,17β-dihydroxyestra-1,4-dien-3-one (p-quinol, 1, 75%) is reported, involving oxidation of 17β-estradiol with potassium permanganate. Similar treatment of 17β-estradiol with sodium chlorite led to 10β-chloro-17β-hydroxyestra-1,4-dien-3-one (2) in 44% yield along with smaller amounts 4-chloro-10β,17β-dihydroxyestra-1,4-dien-3-one (3), 2,10β-dichloro-17β-hydroxyestra-1,4-dien-3-one (4), and 4,10β-dichloro-17β-hydroxyestra-1,4-dien-3-one (5). [Copyright &y& Elsevier]

Abstract: Recent advances on biomarker research are discussed, primarily relying on experience gained with technologies based on liquid chromatography–tandem mass spectrometry (LC–MS–MS). Determination of urinary metabolites of industrial chemicals (n-hexane, benzene, toluene, and styrene) in samples from occupationally exposed workers and controls was performed by LC–MS–MS with either electrospray (ESI) or atmospheric pressure chemical ionization (APCI), as appropriate. Both phase I and II metabolites (glucuronides, sulfates, and mercapturic acids) can be detected with little or no sample manipulation, thus allowing the identification of a number of artifacts and “new” metabolites. However, experimental evidence indicates the need for properly addressing the matrix effect, which is always associated with the analysis of biological samples. Both efficient sample preparation and the use of isotopically labeled internal standards seem to be necessary to develop validated quantitative methods. [Copyright &y& Elsevier]

Abstract: The reactions of 5-S-cysteinyldopa (1), l-α-methyldopa (2) and dl-m-tyrosine (3) with d-glucose were investigated at 90°C in phosphate buffer at pH ranging from 5.0 to 9.0. Whereas 1 gave mainly the double Maillard condensation product N,N′-bis(1″-deoxy-d-fructos-1″-yl)-5-S-cysteinyldopa, as an inseparable mixture of β-d-fructopyranosyl and α,β-d-fructofuranosyl derivatives, 2 and 3 gave both Maillard and Pictet–Spengler products, although to different extents and with different regio- and stereochemistry. A peculiar pattern of reactivity was displayed by 2 which gave, besides the Maillard product and the expected 6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline C-1 diastereoisomeric pairs, the unprecedented 7,8-dihydroxy-1,2,3,4-tetrahydroisoquinoline derivative via the ortho cyclization pathway. Pictet–Spengler cyclization of 2 and 3 proceeded with Felkin–Anh-type asymmetric induction, favouring the 1R isomer throughout the pH range 5.0–9.0. These results, which highlight the first example of carbohydrate-derived 7,8-dihydroxytetrahydroisoquinoline, provide new insights into the factors governing competition between Maillard and Pictet–Spengler condensation pathways. [Copyright &y& Elsevier]

Abstract: Oxidation of 4-nitro-17β-estradiol (1) with the peroxidase/H2O2 system gave the symmetric C2-linked dimer (3) through phenoxy radical coupling. Similar oxidation of 2-nitro-17β-estradiol (2), in which the nitro group is coplanar with the aromatic ring, yielded 9α- and 9β-hydroxy-2-nitro-17β-estradiol (4a,b), (17β)-2-nitroestra-1(10),2,4,9(11)-tetraene-3,17-diol (5), and (12α,17β)-2-nitroestra-1(10),2,4,9(11)-tetraene-3,12,17-triol (6). With higher concentrations of H2O2, the novel secoestra-1(10),2,4-trien-9-one derivative 7 was obtained from 2. Theoretical calculations suggested that the peculiar behavior of 2 may be due to the generation of a relatively stable radical intermediate at C9, which would then be converted to the reactive quinone methide 8. The chemistry described in this paper appears to be an intriguing example of control of the site of substitution over evolution of phenoxy radicals, and opens new vistas toward selective oxyfunctionalization of the estrane skeleton. [Copyright &y& Elsevier]

Abstract: A method for the determination of volatile chlorinated hydrocarbons, namely dichloromethane (DCM), trichloroethylene (TCE), and perchloroethylene (PCE), in urine samples was developed using headspace solid phase microextraction (HS-SPME) gas chromatography–mass spectrometry (GC–MS). HS-SPME was performed using a 75μm Carboxen-polydimethylsiloxane fiber. Factors, which affect the HS-SPME process, such as adsorption and desorption times, stirring, salting-out effect, and temperature of sampling have been evaluated and optimized. The highest extraction efficiency was obtained when sampling was performed at room temperature (22°C), from samples saturated with salt and under agitation. Linearity of the HS-SPME-GC–MS method was established over four orders of magnitude and the limit of detection was 0.005μg/l for all the compounds. Precision, calculated as %R.S.D. at three different concentration levels, was within 1–8% for all intra- and inter-day determinations. The method was applied to the quantitative determination of TCE and PCE in human urine samples from exposed (TCE, n =5; median, 9.32μg/l and PCE, n =39; median, 0.58μg/l) and non-exposed individuals (n =120; median concentrations, 0.64, 0.22 and 0.11μg/l for DCM, TCE and PCE, respectively. In addition, two cases of acute accidental exposure to DCM are reported, and the elimination kinetics in blood and urine was followed up. The calculated half-lives of urinary and blood DCM were, respectively, 7.5 and 8.1h for one subject and 3.8 and 4.3h for the other. [Copyright &y& Elsevier]

Abstract: Oxidation of 5×10-3M noradrenaline in aqueous phosphate buffer, pH 7.4, with K3Fe(CN)6, NaIO4 or Fe2+/EDTA/H2O2 followed by extraction with ethyl acetate and acetylation with Ac2O/Pyr led to a main reaction product which was isolated and identified as 4-[bis-(1H-5,6-diacetoxyindol-2-yl)methyl]-1,2-diacetoxybenzene, an unprecedented [bis-(indol-2-yl)methyl]-benzene derivative unsubstituted on the 3-position of the indole rings. This product was also obtained in 40% yield by reaction of 5,6-dihydroxyindole with 3,4-dihydroxybenzaldehyde. Other components of the oxidation mixture were 1-acetyl-3,5,6-triacetoxyindole, derived from noradrenolutin, and 5,6-diacetoxyindole, originating from cyclisation/dehydration of the o-quinone of noradrenaline, along with some 3,4-diacetoxybenzaldehyde. Inspection of the aqueous phase revealed the presence of 3,4-dihydroxymandelic acid and 3,4-dihydroxybenzaldehyde, derived from oxidative breakdown of the 2-amino-1-hydroxyethyl chain via a p-quinomethane intermediate. These results disclose new aspects of the oxidative chemistry of noradrenaline beyond the aminochrome stage and provide a route to novel [bis-(indol-2-yl)methyl]-benzene derivatives of potential pharmacological interest. [Copyright &y& Elsevier]

Abstract: Biological monitoring and biomarkers are used in occupational toxicology for a more accurate risk assessment of occupationally exposed people. Appropriate and validated biomarkers of internal dose, like urinary metabolites, besides to be positively correlated with external exposure, have a predictive value to the risk of adverse effects. The application of liquid chromatography–mass spectrometry (LC–MS) in occupational and environmental toxicology, although relatively recent, has been demonstrated valid in the determination of traditional biomarkers of exposure, as well as in metabolism studies aimed at investigating minor metabolic routes and new more specific biomarkers. This review presents selected applications of LC–MS to the study of the metabolism of industrial chemicals, like n-hexane, benzene and other aromatic hydrocarbons, styrene and other monomers employed in plastic industry, as well as to other chemicals used in working environments, like pesticides used by farmers, and antineoplastic agents prepared by hospital personnel. Analytical and pre-analytical factors, which affect quantitative determination of urinary metabolites, i.e. sample preparation, matrix effect, ion suppression, use of internal standards, and calibration, are emphasized. [Copyright &y& Elsevier]

Objective. To apply mass spectrometry-based techniques for the determination of background levels of metabolites of widespread organic solvents, such as styrene, in unexposed subjects from the general population. Methods. The study population consisted of 129 healthy Italian subjects, not occupationally exposed to styrene (67 men; 37% smokers). Urinary levels of mandelic acid (MA) and phenylglyoxylic acid (PGA) were determined by liquid chromatography–tandem mass spectrometry (LC-MS/MS). Results. Using LC-MS/MS, we were able to detect the peaks of MA and PGA in 100% of samples. The distribution of styrene metabolites was log-normal, the geometric means of MA and PGA concentrations being 0.443 mg/g creatinine [geometric standard deviation (GSD) 2.34] and 0.107 mg/g creatinine (GSD 3.49), respectively. The reference intervals estimated for MA and PGA were 0.084–2.339 and 0.009–1.238 mg/g creatinine, respectively. Conclusion. The application of highly sensitive and selective mass spectrometry-based analytical techniques could be useful in the definition of reference values for metabolites of organic pollutants and for better assessment of low-level occupational exposure to organic solvents. [ABSTRACT FROM AUTHOR]

Nitration of 17β-estradiol by H2O2 and nitrite in the presence of various peroxidases, viz. horseradish peroxidase, lactoperoxidase, and peroxidase-containing homogenates from bovine uteri, was systematically investigated to assess on a chemical basis its potential relevance to the mechanisms of impairment of estrogen functions under oxidative/nitrosative stress conditions. In the presence of excess nitrite 17β-estradiol reacted smoothly to give 2-nitroestradiol (1), 4-nitroestradiol (2), and 2,4-dinitroestradiol (3). With 10–300 μM estradiol, formation yields of 1–3 were 12–55%, but dropped to 1% or less at lower estrogen concentration, for example, 1 μM, or in plasma as the reaction medium. Time course analysis showed that 2 is the prevalent nitration product under conditions of slow generation of nitrating species, suggesting some regioselectivity for estradiol nitration at C-4, whereas 1 prevails with bolus addition of reagents, due to faster degradation of 2. Competition experiments carried out with 15NO2- showed that 2 is about twice more susceptible to nitration than 1 as determined by 15N NMR analysis of the resulting 3. The biological effects of 1 and 2 were preliminarily tested on in vitro bovine embryo cultures. When 1 and 2 were substituted to the standard 17β-estradiol in the oocyte maturation, a significant decrease in both cleavage and blastocyst efficiency was observed in the case of 1 but not 2. Overall, these results suggest that estradiol nitration is a potential pathway of hormonal dysfunction and toxicity but would require elevated estrogen levels of questionable physiological relevance. [Copyright &y& Elsevier]

A new and simple method for the determination of fat-soluble vitamins (retinol, α-tocopherol, and β-carotene) in human serum was developed and validated by using liquid chromatography–tandem mass spectrometry with atmospheric pressure chemical ionization (LC-APCI-MS-MS). Different solvent mixtures were tested to obtain deproteinization and extraction of the analytes from the matrix. As a result, a volume of 240 μL of a 1:1 (v/v) ethanol/ethyl acetate mixture added to 60 μL of serum was found to be suitable for both protein precipitation and antioxidants solubilization, giving the best recovery for all three analytes. Deproteinized samples (20 μL) were injected after dilution, without the need for concentration or evaporation to dryness and reconstruction of the sample. Vitamins were separated on a C-8 column using a 95:5 (v/v) methanol/dichloromethane mixture and ionized in the positive-ion mode; detection was performed in the selected-reaction monitoring mode. Linearity of the LC-APCI-MS-MS method was established over 5 orders of magnitude for retinol and α-tocopherol, whereas in the case of β-carotene it was limited to 4 orders. Lower limits of quantitation were 1.7, 2.3, and 4.1 nM for retinol, α-tocopherol, and β-carotene, respectively. Serum concentrations of retinol, α-tocopherol, and α+β-carotene determined in a group of healthy volunteers were 2.48, 38.07, and 0.50 μM, respectively, in samples collected in winter (n=122) and 2.69, 45.88, and 0.90 μM during summer (n=66). [ABSTRACT FROM AUTHOR]

Discusses the involvement of several redox-related mechanisms in the toxicities of xenobiotics that triggers enhanced sensitivity in Fanconi anemia cells. Association of chromium toxicity with oxidative stress; Formation of DNA or protein-DNA cross-links; Fanconi anemia cell deficiency in withstanding oxidative stress; Relationship of the Fanconi anemia phenotype with a prooxidant state.

Styrene is a chemical widely used in the plastic industry. The main pathway of styrene metabolism in humans occurs via the oxidation to styrene-7,8-oxide (7,8-SO). The aim of this study was the investigation of a minor metabolic route, involving the oxidation of the arene moiety of styrene, by means of the characterization of the conjugated urinary metabolites of 4-vinylphenol (4-VP). 4-vinylphenol-glucuronide (4-VP-G) and -sulfate (4-VP-S), were measured by liquid chromatography electrospray tandem mass spectrometry (LC–ESI–MS/MS) from 174 workers belonging to three cohorts recruited in European countries and from 26 volunteers exposed to 50 mg/m3 (11.8 ppm) of styrene for 8 h. The 4-VP conjugates represented about 0.5–1% of the total excretion of styrene metabolites. Both 4-VP-G and 4-VP-S are eliminated with a monophasic kinetic, the glucuronide being excreted faster (half-time, 2.2 ± 0.2 h) than the sulfate (half-time 9.7 ± 1.7 h). The urinary 4-VP was found to be significantly correlated both with airborne styrene (r = 0.607, p < 0.001) and the sum of MA and PGA (r = 0.903, p < 0.001 in “end-of-shift” samples). Apart from 7,8-SO, 4-VP is the only styrene metabolite not shared with ethylbenzene and therefore thought to be a highly specific marker of styrene exposure. However, a measurable background excretion of 4-VP was also found in all urine samples from controls not occupationally exposed to styrene. This background appears to be highly correlated to smoking (p < 0.001) and possibly also to the dietary intake of styrene or 4-VP. Consequently, the use of 4-VP as a biomarker of styrene exposure is recommended for exposures exceeding 1 ppm. [Copyright &y& Elsevier]

Ni2+, a toxic, carcinogenic and allergenic agent, affected both the kinetic and chemical courses of the Fe2+-induced oxidation of arachidonic acid (AA) in 0.05 M phosphate buffer (pH 7.4) and at 37 °C. At 10 μM concentration, Ni2+ decreased the rate of oxidation of peroxide-free AA (200 μM) promoted by 50 μM Fe2+, as determined by measurement of thiobarbituric acid reactive species (TBARS) and 1H NMR analysis. However, in the presence of low levels of peroxides (e.g. 2%), Ni2+ exerted a significant stimulatory effect on Fe2+-induced AA oxidation and TBARS formation. 1H NMR analysis showed that Ni2+ (10 μM) enhanced formation of genotoxic alkenals including 4-hydroxy-2-nonenal (4-HNE, GC/MS evidence) by Fe2+-promoted degradation of both AA and 15-hydroperoxy-5,8,11,13-eicosatetraenoic acid (15-HPETE) methyl esters. The observed stimulatory effects of Ni2+ on peroxide breakdown and cytotoxic aldehyde formation provide an attractive explanation to the enhanced sensitization capacity of nickel in inflammatory states compared to normal states. [Copyright &y& Elsevier]

In the presence of excess H2O2, oxidation of dopamine was diverted from the usual pigment-forming pathway to afford 6-hydroxydopamine and then a colorless reaction mixture comprising a polar non-extractable product. The latter was obtained in 20% yield by oxidation of 6-hydroxydopamine and was tentatively formulated as the novel 5-(2-aminoethyl)-2-hydroxy-5-(3-hydroxy-2-oxotetrahydro-1aH-oxireno[2,3]cyclopenta[1,2-b]pyrrol-3a(4H)-yl)cyclohex-2-ene-1,4-dione by extensive spectral analysis and conversion to a tetraacetyl derivative. Mechanistic experiments suggested that formation of the product proceeds via 6-hydroxydopamine by H2O2-dependent epoxidation and cyclization steps followed by dimerization and ring contraction with decarboxylation. [Copyright &y& Elsevier]

Two workers were accidentally exposed to unusually high styrene concentrations (>1000 ppm) for about 30 min. In addition to the main styrene metabolites, mandelic acid (MA) and phenylglyoxylic acid (PGA), other minor metabolites, including specific mercapturic acids, (R,R)- and (S,R)-N-acetyl-S-(1-phenyl-2-hydroxyethyl)-l-cysteine [(R,R)-M1 and (S,R)-M1] and (R,R)- and (S,R)-N-acetyl-S-(2-phenyl-2-hydroxyethyl)-l-cysteine [(R,R)-M2 and (S,R)-M2], 4-vinylphenol-glucuronide and -sulfate, and phenylglycine, were determined by Liquid Chromatography Electrospray Tandem Mass Spectrometry (LC-ESI-MS/MS) in urine samples collected 12, 24, 36, 48, 75 and 99 h after the episode. The genotypes of microsomal epoxide hydrolase, glutathione-S-transferases M1-1 (GSTM1), T1-1 (GSTT1) and P1-1 (GSTP1) were characterized by PCR-based methods. The two subjects showed similar peak levels of MA and PGA, as well as 4-vinylphenol conjugates, whereas mercapturic acids were five times higher in the subject bearing the GSTM1pos than in the GSTM1null subject. Also, relative proportions of diasteroisomers of mercapturic acids were influenced by the GSTM1 polymorphism. [Copyright &y& Elsevier]

Investigates the Maillard reaction of L-3,4-Dihydroxyphenylalanine (L-DOPA) and dopamine with D-glucose under physiologically relevant conditions to specifically elucidate the nature of competing pathways. Reaction of L-DOPA with D-glucose; Reactions of Dopamine and L-tyrosine with D-glucose; Effects of transition metal cations on the reaction of L-DOPA with D-glucose.

Describes a novel octahydropyridobenzothiazepine metabolite in human urine. Details on a biomimetic formation from the melanogen 5-S-cysteinyldopa and formaldehyde; Relevance of the Pictet-Spengler condensation in forming the compound; Critical role of sulfur substituents in subverting regiochemistry of Schiff bases cyclization.

Four melanin polymers prepared from different precursors (5,6‐dihydroxyindole, 5,6‐dihydroxyindole 2‐carboxylic acid, 1,8‐dihydroxynaphthalene and dopamine) have been adopted as photocatalysts in the reductive dehalogenation of a series of model α‐halogen carbonyl derivatives. The best performing melanin was that obtained from 5,6‐dihydroxyindole 2‐carboxylic acid, which allowed to dehalogenate efficiently both bromomalonate esters and phenacyl bromide; on the other hand, chloromalonate esters were almost unreactive under the same reaction conditions. Electrochemical and control experiments enabled to elucidate the reaction mechanism, demonstrating the key role of electrons as charge carriers in the observed photocatalytic process. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of citronella oil obtained from the leaves of Cymbopogon nardus (L.) Rendle, when used as a sensory additive for all animal species. The EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) concluded that citronella oil from C. nardus is of low concern for long‐living and reproductive animals at the use levels in complete feed of 3.5 mg/kg for laying hens and rabbits, 6 mg/kg for sows and dairy cows, 9.5 mg/kg for sheep/goats and horses, 2.0 mg/kg for cats and 10 mg/kg for dogs. For short‐living animals (species for fattening), the additive was considered of no concern at concentrations of 18 mg/kg in chickens for fattening, 24 mg/kg in turkeys for fattening, 20 mg/kg for piglets, pigs for fattening, veal calves (milk replacer), cattle for fattening, sheep/goats for meat production, horses for meat production and rabbits for meat production, and 30 mg/kg for salmonids. The conclusions were extrapolated to physiologically related minor species. For any other species, the additive is considered of low concern at 2.0 mg/kg complete feed. The use of citronella oil in animal feed is expected to be of no concern for the consumers and for the environment. The essential oil under assessment should be considered as irritant to skin and eyes and as a dermal sensitiser. When handling the essential oil, exposure of unprotected users to methyleugenol may occur. Therefore, to reduce the risk, the exposure of the users should be minimised. Since the leaves of C. nardus and its preparations were recognised to flavour food and its function in feed would be essentially the same as that in food, no further demonstration of efficacy was considered necessary. [ABSTRACT FROM AUTHOR]

Studies the oxidation of 5,6 dihydroxyindole in acidic aqueous media. Characterization of the products formed by acid promoted oxidation of 1a; Experimental procedures utilized; Formation of diindolocarbazoles; Description of the oxidation of indolic compounds in acidic media.

This technical report implements the EFSA MIXTOX guidance document for animal health risk assessment of combined exposure to multiple chemicals in essential oilsusing whole mixture and component‐based approaches (WMA and CBA).The harmonised step wise approaches from the MIXTOX guidance are applied to the problem formulation, exposure assessment, hazard assessment and risk characterisation using conservative scenarios, available data and tools. Options for the refinement of the CBA are proposed for the exposure assessment and hazard assessment (toxicity and toxicokinetic data and models) steps as well as for the application of risk characterisation methods. For exposure assessment, exposure metrics are derived for the whole mixture or for each component of the assessment groupusing the maximum proposed use levels of the essential oil in feedcombined with the feed consumption patterns corrected for body weight for each farm animal species. For hazard assessment, combined chronic toxicity on a species‐specific basis requires data collection of reference points from chronic tolerance studies in each farm species or (sub)‐chronic toxicity studies on test species. For the WMA, the chronic toxicity of the whole mixture is used and divided by uncertainty factors.For the component‐based approach, the chronic toxicity for each substance in each assessment group is collected. For risk characterisation using WMA and CBA, a margin of exposure approach and a combined margin of exposure approach are applied respectively. Options for refining the approachesfor the component‐based approach are proposedincluding the use ofspecific toxic effectsfor the grouping of chemicals as well as the use of toxicokinetic data and generic physiologically‐based kinetic models for farm animal species. [ABSTRACT FROM AUTHOR]

Astrochemistry and astrobiology, the fascinating disciplines that strive to unravel the origin of life, have opened unprecedented and unpredicted vistas into exotic compounds as well as extreme or complex reaction conditions of potential relevance for a broad variety of applications. Representative, and so far little explored sources of inspiration include complex organic systems, such as polycyclic aromatic hydrocarbons (PAHs) and their derivatives; hydrogen cyanide (HCN) and formamide (HCONH2) oligomers and polymers, like aminomalononitrile (AMN)-derived species; and exotic processes, such as solid-state photoreactions on mineral surfaces, phosphorylation by minerals, cold ice irradiation and proton bombardment, and thermal transformations in fumaroles. In addition, meteorites and minerals like forsterite, which dominate dust chemistry in the interstellar medium, may open new avenues for the discovery of innovative catalytic processes and unconventional methodologies. The aim of this review was to offer concise and inspiring, rather than comprehensive, examples of astrochemistry-related materials and systems that may be of relevance in areas such as surface functionalization, nanostructures, and hybrid material design, and for innovative technological solutions. The potential of computational methods to predict new properties from spectroscopic data and to assess plausible reaction pathways on both kinetic and thermodynamic grounds has also been highlighted. [ABSTRACT FROM AUTHOR]

The catalytic application of chiral phosphoric acids (CPAs) from 2004 to date represents a true milestone for asymmetric organocatalysis. However, not only the free acids can be conveniently employed in numerous different asymmetric synthetic methodologies, thus being strictly compliant to the concept of “organocatalysis”, but also their metal salts. This review focuses on asymmetric reactions in which the catalyst is a chiral alkali or alkaline‐earth metal phosphate. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP Panel) was asked to deliver a scientific opinion on the safety for the users of the feed additive consisting of ferric tyrosine chelate (TYFER™) when used as a zootechnical additive for chickens, turkeys and minor poultry species for fattening or reared for laying/breeding. The European Commission request follows a previous opinion of the FEEDAP Panel. In that opinion, the Panel identified several risks for the users of the additive; it was listed that it posed a risk to users by inhalation, should be considered as an irritant to skin, eyes and mucous membranes, and also that, due to its nickel content, should be considered as a dermal and respiratory sensitiser. In the current application, the applicant proposed a maximum content of nickel (50 mg/kg). No changes in the manufacturing process have been reported by the applicant. In the absence of new data, the FEEDAP Panel reiterates its previous conclusion that the additive should be as an irritant to skin, eyes and mucous membranes and as a dermal and respiratory sensitiser. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of cajuput oil obtained from fresh leaves of Melaleuca cajuputi Maton & Sm. ex R. Powell and Melaleuca leucadendra (L.) L., when used as a sensory additive for all animal species. The FEEDAP Panel concluded that cajuput oil is safe up to the maximum proposed use levels in complete feed of 30 mg/kg for sows and dogs, 50 mg/kg for horses and ornamental fish, 40 mg/kg for salmon and 5 mg/kg for cats. For the other species, the calculated safe concentrations were 18 mg/kg for chickens for fattening, 26 mg/kg for laying hens, 23 mg/kg for turkeys for fattening, 37 mg/kg for pigs for fattening, 31 mg/kg for piglets, 78 mg/kg for veal calves (milk replacer), 69 mg/kg for cattle for fattening and sheep/goats, 45 mg/kg for dairy cows and 28 mg/kg for rabbits. These conclusions were extrapolated to other physiologically related species. For any other species, the additive is safe at 5 mg/kg complete feed. The use of cajuput oil in water for drinking was considered safe provided that the total daily intake does not exceed the daily amount considered safe when consumed via feed. No concerns for consumers and the environment were identified following the use of the additive up to the highest safe use level in feed. The essential oil under assessment should be considered as an irritant to skin and eyes, and as a dermal and respiratory sensitiser. Since cajuput oil was recognised to flavour food and their function in feed would be essentially the same as that in food, no further demonstration of efficacy was considered necessary. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of Patent Blue V as a sensory feed additive for non‐food‐producing animals. The additive is already authorised for use with non‐food‐producing animals. The applicant has not provided evidence that the additive currently on the market complies with the existing conditions of authorisation. The FEEDAP Panel cannot conclude whether the additive remains safe for the target species due to the non‐compliance with the specifications and the lack of adequate data on the potential aneugenicity of the additive. In the absence of data, the FEEDAP Panel cannot conclude on the potential of the additive to be a dermal and eye irritant nor a dermal and respiratory sensitiser. Since the potential genotoxicity of the additive was not ruled out, the exposure to the additive of the unprotected users should be minimised. The Panel retains that the previously made conclusion on the efficacy remains valid. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the assessment of the application for renewal of authorisation of sodium bisulphate (SBS) as a feed additive for all terrestrial animal species (category: technological additive; functional group: preservative), and for all terrestrial animal species other than cats, mink, pets and other non‐food‐producing animals (category: technological additive; functional group: acidity regulator). EFSA has also been asked to assess the new use of the product as an acidity regulator and flavouring compound in all pets and other non‐food‐producing animals except aquatic animals. The applicant provided evidence that the additive currently in the market complies with the existing conditions of authorisation. There is no evidence that would lead the FEEDAP Panel to reconsider its previous conclusions. Thus, the Panel concluded that the additive remains safe for all terrestrial animal species, consumer and the environment under the authorised conditions of use. The FEEDAP Panel considers that the proposed new use would not introduce risks not already considered in the previous assessment and therefore the same conclusions on all terrestrial animal species, consumers of products from animals fed the additive and the environment would apply. Regarding user safety, the additive is irritant to the skin, eyes and the respiratory tract, and should be considered a skin and respiratory sensitiser. There is no need to assess the efficacy of the additive in the context of the renewal of the authorisation. The Panel considers that the additive has the potential to be efficacious as an acidity regulator and sensory additive (flavouring compound) in feed for pet and non‐food‐producing animals (except aquatic animals). [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety of microcrystalline cellulose and carboxymethyl cellulose as technological feed additives for all animal species. In its previous opinions on the safety and efficacy of the products, the FEEDAP Panel could not conclude on proper identification and characterisation as required for a feed additive. The occurrence of potential toxic impurities could also not be assessed. Based on the new data provided, the feed additives microcrystalline cellulose and carboxymethyl cellulose were properly identified and characterised and were shown to meet the specifications set for their use as food additives. Therefore, the conclusions of the safety reached in the previous opinions for microcrystalline cellulose and carboxymethyl cellulose meeting the food additive specifications apply to the microcrystalline cellulose and carboxymethyl cellulose under assessment as feed additives. The additives are considered safe for all animal species, the consumer and the environment. In the absence of data, the FEEDAP Panel is not in the position to conclude on the safety for the user. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety of hydroxypropyl methyl cellulose and methyl cellulose as technological feed additives for all animal species. In its previous opinions on the safety and efficacy of the products, the FEEDAP Panel could not conclude on proper identification and characterisation as required for a feed additive. The occurrence of potential toxic impurities could also not be assessed. Based on the new data provided, the feed additives hydroxypropyl methyl cellulose and methyl cellulose were properly identified and characterised and were shown to meet the specifications set for the food additives. Therefore, the conclusions of the safety assessments reached in the previous opinions for hydroxypropyl methyl cellulose and methyl cellulose meeting the food additive specifications, apply to the hydroxypropyl methyl cellulose and methyl cellulose under assessment as feed additives. The additives are considered safe for all animal species, the consumer and the environment. In the absence of data, the FEEDAP Panel is not in the position to conclude on the safety for the user. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety of ethyl cellulose as a technological feed additive for all animal species. In its previous opinions on the safety and efficacy of the product, the FEEDAP Panel could not conclude on proper identification and characterisation as required for a feed additive. The occurrence of potential toxic impurities could also not be assessed. Based on the new data provided, the feed additive ethyl cellulose was properly identified and characterised and was shown to meet the specifications set for the food additive. Therefore, the conclusions of the safety assessment reached in the previous opinions for ethyl cellulose meeting the food additive specifications, apply to the ethyl cellulose under assessment as a feed additive. The feed additive is considered safe for all animal species, the consumer and the environment. In the absence of data, the FEEDAP Panel is not in the position to conclude on the safety for the user. [ABSTRACT FROM AUTHOR]