Sanja Kežić

Dermal absorption of cis-1,3- dichloropropene vapour: human experimental exposure

1. The relevance of skin absorption of cis-1,3-dichloropropene (cis-1,3-DCP) vapour as a route of entry compared to inhalatory uptake has been assessed in human volunteers under controlled exposure conditions. 2. Five adults (four males and one female) were dermally exposed on the forearm and hand during 45 min to 86 mg/m3 cis-1,3-DCP. 3. Dermal uptake was assessed by determination of the main cis-1,3-DCP metabolite in urine: the mercapturic acid conjugate of cis-1,3-DCP (cis-1,3-DCP-MA). 4. When whole-body dermal exposure to vapour is compared to inhalatory exposure, the uptake through the skin is estimated to be about 2-5% of the inhalatory absorption.

Determination of 2,5-hexanedione, a metabolite of n-hexane, in urine : evaluation and application of three analytical methods

Three methods for the determination of 2,5-hexanedione (2,5-HD) in urine were compared in order to assess their applicability for toxicokinetic studies and biological monitoring of occupational exposure to n-hexane. Two of them were based on derivatization, followed by gas chromatography and electron-capture detection. Of these two, one is a modification of the other, already published, method. The third one involves direct extraction of 2,5-HD followed by gas chromatography and flame-ionization detection. To determine 2,5-HD in urine of workers occupationally exposed to n-hexane, the most straightforward method, direct extraction of 2,5-HD from urine, has been proven to be the most suitable. However, in case of very low concentrations of 2,5-HD in urine, or analysis of small samples of blood, e.g. in kinetic studies, it is necessary to use a more sensitive procedure. The sensitivity of the methods based on the derivatization of 2,5-HD followed by electron-capture detection, was, as expected, much higher in terms of analytical reliability. By using these methods, however, precautions are necessary to avoid a matrix effect.

Metabolic capacity and interindividual variation in toxicokinetics of styrene in volunteers

The aim of the present study was to assess the interindividual variation in styrene toxicokinetics and to correlate this variation with the individual metabolic capacity for cytochromeP450 (CYP), CYP2E1, CYP1A2 and CYP2D6. Twenty male volunteers were exposed on separate occasions to 104±3 and 360 ± 20 mg/m3 of styrene for 1 h while performing 50 W physical exercise on a bicycle ergometer. Styrene concentrations in blood and mandelic (MA) and phenylglyoxylic acid (PGA) in urine were measured. The metabolic capacity was assessed by phenotyping with chlorzoxazone (CYP2El), caffeine (CYP1A2), dextromethorphan (CYP2D6) and antipyrine (CYP450). In addition, for the main styrene-metabolising enzyme, CYP2E1, genotyping for the genetic polymorphisms of the gene was performed. The average pulmonary retention of styrene was 62 ± 7% at both exposure concentrations, and the 24-h excretion of MA and PGA accounted for 58% of the dose at both concentrations. The interindividual variation in styrene kinetics ranged from 19% for the terminal half-life (t1/2,β) of styrene to 41% for the cumulative excretion of MA and PGA. However, no correlation between the apparent blood clearance of styrene (CLapp), t1/2,β of styrene or excretion of MA and PGA on one hand, and the individual metabolic capacity on the other hand was found. Although other explanations cannot be excluded, this lack of correlation might be due to the high apparent blood clearance (1.4 1/ min) of styrene, indicating that styrene metabolism is liver–blood-flow-dependent.

Determination of methyl ethyl ketone and its metabolites in urine using capillary gas chromatography

A method is described for the determination of the concentration of methyl ethyl ketone and its metabolites: 2-butanol, 3-hydroxy-2-butanone and the meso- and d,l-isomers of 2,3-butanediol in urine. The analytes were isolated from urine by solid-phase extraction and analysed by capillary gas chromatography. The recovery rates were 50-70% for the 2,3-butanediol isomers and 88-96% for the other analytes. The precision of the method ranged from 5 to 12% (S.D.%). The detection limit was 1.0 and 1.4 mg/l for meso- and d,l-2,3-butanediol, respectively, and ranged from 0.1 to 0.15 mg/l for the other analytes.

Gas chromatography-electron capture determination of styrene-7,8-oxide enantiomers.

The enantiomers of styrene-7,8-oxide (phenyloxirane, SO) were determined using a method based on base catalysed hydrolysis with sodium methoxide. The oxirane ring opening resulted in formation, without racemisation, of the enantiomeric pairs of the two regional isomers, 2-methoxy-1-phenylethanol and 2-methoxy-2-phenylethanol. The structure of these regional isomers was confirmed by gas chromatography-mass spectrometry (GC-MS) and proton nuclear magnetic resonance (1H-NMR). To improve sensitivity of determination, the formed methoxy alcohols were subsequently derivatised with pentafluoropropionic anhydride enabling electron capture detection. This derivatization proceeded also without racemisation and the formed pentafluoropropionyl derivatives were separated on two serially coupled columns, a non-chiral AT 1705 and a chiral CP Chirasil-Dex-CB. As internal standard 2S,3S-(-)-2-methyl-3-phenyloxirane was used. The limit of quantitation of the method was 0.2 microM. The repeatability of the method was assessed at two concentration levels (2.5 and 25 microM) and ranged from 6 to 9% for both enantiomers. The method was applied to the determination of the rate and enantioselectivity of the cytochrome P-450 dependent oxidation of styrene to SO enantiomers in human liver microsomes.

Stereochemical metabolism of styrene in volunteers

Objectives: To study the stereochemistry of styrene metabolism in volunteers, and its interindividual variability. Methods: Twenty healthy male volunteers (aged 18–37 years) were exposed to 360 mg/m3 styrene for 1 h while they performed 50 W physical exercise. Venous blood was drawn during and for up to 2 h after exposure. Urine was collected at time-intervals up to 24 h after exposure. The following parameters were determined: styrene, free and conjugated styrene glycol (SG) in blood, and conjugated SG, mandelic acid (MA) and phenylglyoxylic acid (PGA) in urine. Results: Average pulmonary retention of styrene was 62%. Excretion of the acidic metabolites MA and PGA accounted for 58% of the pulmonary uptake. The average maximum concentration (Cmax) and area under the curve (AUC) of free (R)-SG in blood were 1.3 and 1.7 times higher than those of (S)-SG respectively; the half-life of (R)-SG was longer (82 vs 62 min, P < 0.005). Cmax and AUC of the conjugated SG enantiomers in blood did not differ, but again half-life for (R)-SG was longer (72 vs 64 min, P < 0.05). Cumulative excretion and renal clearance of conjugated (S)-SG in urine were three and four times higher, respectively, than that of (R)-SG. Cumulative excretion of (S)-MA was 1.6 times higher than (R)-MA. Interindividual differences in the kinetic parameters of the metabolites were two- to threefold. Conclusions: The enantiomeric excess found was different for each metabolite under study, implying different enantioselectivity and/or enantiospecificity of the enzymes and carrier-proteins involved in the biotransformation and excretion. The use of these metabolites as biological indicators for prediction of the enantiomeric excess of the toxic metabolite styrene-7,8-oxide (SO) is therefore not justified. Interindividual differences in the stereochemical metabolism of styrene are moderate.

Associations of TNFα -308G>A, TNFα -238G>A, IL-1α -889C>T and IL-10 -1082G>A Genetic Polymorphisms with Atopic Diseases: Asthma, Rhinitis and Dermatitis.

Background: Polymorphisms of cytokine genes are an interesting focus for association studies involving atopic diseases due to their role in immune cell communications during inflammation. The aim of this study was to investigate associations of TNFα -308G>A, TNFα -238G>A, IL-1α -889C>T and IL-10 -1082G>A polymorphisms with atopic diseases with adjustment for confounding lifestyle and environmental factors. Methods: This study was performed on 356 Croatian students. The diagnosis of atopic asthma, atopic rhinitis and atopic dermatitis was based on symptoms reported by the modified International Study of Asthma and Allergies in Childhood questionnaire and a positive skin prick test (SPT) to at least one common inhalatory allergen. Genetic polymorphisms were genotyped using the polymerase chain reaction-based technique. The influence of personal (gender, body mass index, parental history of atopic disease), lifestyle (cigarette smoking, pet ownership) and environmental (urban/rural residency, residency in continental/Mediterranean region) factors reported in the questionnaire was investigated by univariate and multivariate analysis. Results: Compared to the control subjects, univariate analysis showed a significant negative association of the TNFα -308G>A polymorphism with atopic asthma, atopic dermatitis, asthma and skin symptoms and positive SPT. These observations were confirmed in a multivariate model only for atopic dermatitis and skin symptoms (atopic dermatitis: OR = 0.27; 95% CI 0.07-1.00; p = 0.050; skin symptoms: OR = 0.29; 95% CI 0.10-0.83; p = 0.021). Conclusions: The results indicate a protective role of TNFα -308G>A genetic polymorphisms regarding atopic dermatitis and skin symptoms even after controlling for personal, lifestyle and environmental factors. Further studies are needed to elucidate the molecular patterns of this association in atopic dermatitis and other chronic inflammatory skin disorders.

Gas chromatographic determination of 2,3-butanediol isomers in urine.

Recent studies have revealed that 2,3-butanediol is involved in the metabolic pathway of ethanol, acetone and methyl ethyl ketone. In humans, 2,3-butanediol is one of the main known metabolites of methyl ethyl ketone. However, isolation of such a polar compound from urine followed by gas chromatography (GC) is generally known to be troublesome and to lack good recovery and precision. The method described here effectively deals with both problems in a single-step procedure.

Contents Vol. 227, 2013

G. Argenziano, Reggio Emilia M. Augustin, Hamburg W.H. Boehncke, Geneve L. Borradori, Bern R.P. Braun, Zürich R. Cerio, London L. Cerroni, Graz O. Chosidow, Créteil A. Dupuy, Rennes J.W. Fluhr, Berlin L. French, Zürich F. Furukawa, Wakayama M. Gilliet, Lausanne A.K. Gupta, London, Ont. R. Happle, Freiburg R.J. Hay, London P. Itin, Basel G. Kaya, Geneva R.G.B. Langley, Halifax, N.S. J.M. Mascaro, Jr., Barcelona K. Matsunaga, Nagoya Y. Miyachi, Kyoto L. Naldi, Bergamo F. Nestle, London C. Paul, Toulouse T. Shiohara, Tokyo Th. Simonart, Brussels H.P. Soyer, Brisbane L. Thomas, Lyon R.M. Trueb, Zürich T.J. Yoon, Jinju Ch.C. Zouboulis, Dessau International Advisory Board