Preparation of Test Material and Evaluation of a Proficiency Test for Official Food Control Laboratories on the Determination of Polycyclic Aromatic Hydrocarbons in Baby Food

A proficiency test on the determination of polycyclic aromatic hydrocarbons (PAHs) was organized by the German National Reference Laboratory for PAH in 2010. The test samples were produced by spiking cereal-based instant baby food with PAHs at concentration levels between 0.6 and 3.8 μg/kg; homogeneity and stability of the test material were verified before sample dispatch. Twenty-one official laboratories from Germany and Austria participated in the test and the evaluation of the test was done by applying methods of robust statistics. The individual performance was assessed with the help of z-scores. As to the quantitative results, the dispersion of data for the most important group of benzo(a)pyrene (BaP), benz(a)anthracene (BaA), benzo(b)fluoranthene (BbF), and chrysene (CHR) appeared to be acceptable, with a relative robust standard deviation ranging from 13.2% for BbF to 26.7% for BaA. In total, the performance of one laboratory had to be rated as unsatisfactory because of a result for BaP outside the limits of tolerance. The methods applied in the test may be considered to be comparable, as no significant effects in the distribution of data could be attributed to certain analytical procedures.     

Activation of Polycyclic Aromatic Compounds by cDNA-Expressed Phase I and Phase II Enzymes

A large number of human and other mammalian xenobiotic-metabolizing enzymes have been expressed in target cells of standard mutagenicity tests, such as Ames's Salmonella typhimurium strains and Chinese hamster V79 cells. These recombinant cells are useful for determining the ability of individual enzymes to activate (or inactivate) a given compound. In contrast to standard S9-mediated test systems, they also allow the detection of mutagenic metabolites that do not penetrate cell membranes--a situation often found with reactive phase II metabolites. We present mutagenicity data for benzo[ a ]pyrene and dibenzo[ a,l ]pyrene in V79-derived cells expressing human cytochrome P450 (CYP) 1A1, 1A2, and 1B1, and for 1-hydroxymethylpyrene, R - and S -1-( f -hydroxyethyl) pyrene, 4-hydroxycyclopenta[ def ]chrysene and N -hydroxy-2-acetylaminofluorene in V79-derived cells and/or Salmonella strains expressing the 11 human sulfotransferases (SULTs) identified. In some cases, allelic variants and orthologous enzymes from other mammalian species were also investigated. The data indicate that mutagenicity of many compounds is detected in the appropriate recombinant systems at extremely low substrate concentrations, that the activation of various promutagens is mediated with high specificity by only one or few enzyme forms, and that substantial differences may occur between alloenzymes from the same species and orthologous enzymes from different species. Such information could be important for understanding differences in susceptibility between tissues, species, individual genetic traits, and physiological states.     

Detection of Reactive Quinones in the Metabolism of Polycyclic Aromatic Hydrocarbons by the Formation of their Glutathione Conjugates

The biotransformation of polycyclic aromatic hydrocarbons to quinones by rat liver microsomes was investigated. The employment of an electrochemical detector allowed the specific detection of quinones separated by reverse phase HPLC with higher sensitivity as compared to UV detection. Microsomal incubations of benzo[a]pyrene (BP) resulted in the formation of 1,6-, 3,6- and 6,12-quinones, of naphthalene in the detection of naphthalene-1,4-quinone, whereas ortho-quinones could only be detected in trace amounts. Additional protein binding studies showed that only 9–22% of synthetic ortho-quinones could be recovered from microsomal incubations. In order to scavenge possible reactive quinone metabolites with glutathione (GSH) and to identify these metabolites, GSH-conjugates of naphthalene-1,2-quinone, naphthalene-1,4-quinone, chrysene-1,2-quinone, BP-7,8-quinone and BP-9,10-quinone were synthesized and spectroscopically characterized. After incubations of 1-naphthol or naphthalene with rat liver microsomes the GSH conjugate of naphthalene-1,2-quinone could be identified by cochromatography with the authentic reference compound.     

PHOTOCHEMICAL REACTION OF CHRYSENE IN ACETONITRILE/WATER

The photochemical reaction of chrysene under UVA light irradiation in acetonitrile/water yields 1,4- and 5,6-chrysenequinones and one lactone, H-benzo[d]naphtha[1,2-b]pyran-6-one. Solvent-dissolved oxygen is necessary for the photolysis. The presence of TiO₂, Al₂O₃, La₂O₃, KI, or I₂ decreased the photolysis rate. The free radical scavenger, Na₂S₂O₃, greatly suppressed the chrysene photolysis, indicating that a free radical process was involved. Also, a self-catalysis phenomenon for chrysene photolysis was observed and investigated.     

Syntheses and Identification of Benzo[C]Chrysene Metabolites

Like other PAHs, chrysenes are thought to exert their carcinogenicity via metabolic activation of proximally carcinogenic dihydrodiols to diol epoxides as ultimate carcinogens. Benzo[c] chrysene (B[c]C) is structurally intriguing among the PAH because it features both a bay region and a fjord region. Although B[c]C is carcinogenic and mutagenic, few data are available on its metabolic activation or the nature of its metabolites.

We have synthesized the B[c]C trans-1,2-, 7,8-, and 9,10-dihydrodiols from the appropriate methoxy-substituted bisnaphthyl olefins by photochemical cyclization. B[c]C was metabolized with S9 liver fraction from phenobarbital/β-naphthoflavone-treated rats. Dihydrodiols were formed on both terminal rings as well as in the K-region. 2-, 3-, and 10-HydroxyB[c]C were also identified as metabolites. In mutagenicity studies toward S. typhimurium TA100, 1,2-dihydrodiol was more mutagenic than B[c]C at doses above 1.25 μg/plate, whereas 9,10-dihydrodiol was toxic at doses above 1.25 μg/plate.     

Evaluation of Benzo[c]Chrysene Dihydrodiols in the Morphological Cell Transformation of Mouse Embryo Fibroblast C3H10T1/2CL8 Cells

The morphological cell transforming activities of three dihydrodiols of benzo[c]chrysene (B[c]C), trans?B[c]C-7,8-diol, trans?B[c]C-9,10-diol, and trans?B[c]C-1,2-diol were compared to those of B[c]C in order to study the possible routes of metabolic activation in transformable C3H10T1/2 mouse embryo fibroblasts. B[c]C-treated C3H10T1/2 cells exhibited a concentration-related increase in morphologically transformed foci over a concentration range of 0–3 μg/ml. At 3 μg/ml, B[c]C induced 1.23 Type II & III foci/dish, with 73% of the dishes exhibiting Type II or Type III foci, and a survival of 87%. trans?B[c]C-7,8-diol produced concentration-related responses over a range of 0–5 μg/ml. At 3 μg/ml, trans?B[c]C-7,8-diol produced 1.13 Type II & III foci/dish with 72% of the dishes exhibiting foci, and a survival of 76%. trans?B[c]C-9,10-diol was inactive as a morphological cell transforming agent over a concentration range of 0–3 μg/ml. trans?B[c]C-1,2-diol was also inactive as a morphological cell transforming agent over a concentration range of 0–3 μg/ml. These results suggest that a K-region dihydrodiol of B[c]C, trans?B[cC-7,8-diol, may play a role in the ability of B[c]C to morphologically transform C3H10T1/2 cells.     

内标校准液相色谱荧光法测定食用油中苯并（a）芘

采用内标苯并（b）屈校正,建立了正相固相净化-反相液相色谱-荧光分光光度法测定食用油中苯并（a）芘的方法。正相Plus Silica固相萃取柱作为净化柱,反相C18柱作为分离柱。以苯并（b）屈作为内标,测定了苯并（a）芘的校正因子。使用本方法,苯并（a）芘在液态食用油和固态食用油中的检出限分别为0.01和0.03μg/kg。苯并（a）芘在不同食用油回收率在83%～108%之间,方法的日内精密度和日间精密度分别小于7.7%和13.2%。     

A Coupled-Column HPLC Method for Routine Analysis of Various Chrysene and Benzo[a]Pyrene Metabolites in Urine

The carcinogenic potential of some PAHs leads to the necessity of expressive biological monitoring for people occupationally exposed to PAH. A highly automated, coupled-column, high performance liquid chromatography method for the simultaneous quantification of several chrysene and benzo[a]pyrene metabolites in urine of exposed subjects is presented. After enzymatic hydrolysis of the metabolites, the sample can be directly injected into the HPLC system. The instrumental set-up comprises an analytical column and a precolumn, connected via a 6-port switching valve. Clean-up and analyte preconcentration are automatically performed on the precolumn which is packed with copper-phthalocyanine modified silica. Afterwards, analytes are automatically transferred onto the analytical column where separation is carried out by elution with a methanol-water gradient. Detection of the analytes makes use of their natural fluorescence. Thus, clean-up and analytical separation require little time, making the method suitable for routine analysis in biomonitoring.     

An Abbreviated Synthesis of 7,12-Dimethylbenz[ a ]anthracene and Benzo[ c ]chrysene Metabolites Using the Suzuki Reaction

Efficient syntheses of important metabolites of 7,12-dimethylbenz[ a ]anthracene (DMBA) and benzo[ c ]chrysene (B[ c ]C), via Suzuki coupling reaction are described. This approach provides an excellent method for the preparation of 3-methoxy-DMBA 5 , 10-methoxy-B[ c ]C 14 and 2-methoxy B[ c ]C 20 , and hence for the corresponding dihydrodiols 6 , 15 , and 21 . Following a similar Suzuki reaction, DMBA-6(5 H )-one 8 was also synthesized in high yield.     

6，12-二溴屈的合成研究

含屈基团的OLED材料被认为是很有开发前景和实用价值的一类发光材料，6，12-二溴屈是合成其他屈衍生物重要的中间体。以磷酸三甲酯为溶剂、以溴素为溴化剂合成了6，12-二溴屈，对影响反应的重要因素进行了考察和讨论。反应的最佳条件为：反应温度80℃，溴素分批加入。可以得到75％的收率。