Epigenetic toxicity of hydroxylated biphenyls and hydroxylated polychlorinated biphenyls on normal rat liver epithelial cells

2-Biphenylol, 3-biphenylol, 2,2'-biphenyldiol, 3,3'-biphenyldiol, 3-chloro-2-biphenylol, and 4,4'-dichloro-3-biphenylol were evaluated using the scrape-loading/dye transfer (SL/ DT) technique to determine in vitro gap junctional intercellular communication (GJIC) in a normal rat liver epithelial cell line as a measure of the epigenetic toxicity. Cytotoxicity was determined using the neutral red uptake assay. A dose range of 0-300 microM was examined. Only 3,3'-biphenyldiol and 4,4'-dichloro-3-biphenylol induced cytotoxicity within the tested dose ranges. Noncytotoxic doses were selected for evaluation of epigenetic toxicity. 4,4'-Dichloro-3-biphenylol was most inhibitory to GJIC at the lowest dose. The cytotoxicity and GJIC inhibitory effects observed for 4,4'-dichloro-3-biphenylol might be, although not exclusively, a consequence of the lipophilic nature of this chemical. 3-Chloro-2-biphenylol was least inhibitory to GJIC. 3-Chloro-2-biphenylol was less inhibitory to GJIC than 2-biphenylol because of the presence of the chlorine functional group, which appears to attenuate the toxic effect of the ortho-hydroxyl group. Although cells were capable of complete recovery of GJIC after removal of each of the chemicals, only with 2,2'-biphenyldiol and 4,4,'-dichloro-3-biphenylol did the cells demonstrate partial recovery without removal of the chemical. The more noncoplanar conformation of 2,2'-biphenyldiol and 2-biphenylol might explain their more inhibitory behavior in comparison to 3,3'-biphenyldiol and 3-biphenylol, respectively.     

Role of temperature and enzyme induction in the biotransformation of polychlorinated biphenyls and bioformation of hydroxylated polychlorinated biphenyls by rainbow trout (Oncorhynchus mykiss)

Hydroxylated PCBs (OH-PCBs) are metabolites of polychlorinated biphenyls (PCBs) that have recently been found in the plasma of Great Lakes fish. Studies have shown that the ability of laboratory-held rainbow trout (Oncorhynchus mykiss) to bioform OH-PCBs from dietary mixtures of PCB congeners is complex and may be attributed to factors such as temperature and/or enzyme induction. Past studies have also suggested that CYP1A- and 2B-like enzymes are the likely mechanism for forming OH-PCBs, but this has not been directly studied in a controlled setting. To address these issues, we exposed rainbow trout (-80 g) to dietary concentrations of a mixture of three Aroclors (1248, 1254, and 1260), at three water temperatures (8, 12, and 16 oC), as well as additional PCBs known to induce CYP1A- and CYP2B-like isoforms in mammals. PCB half-lives in trout were inversely related to water temperature, but biotransformation of PCBs was positively related to water temperature. Thirty-one OH-PCBs were observed in trout plasma after 30 days of dietary exposure to the Aroclor mixtures, although approximately 40% of the sigmaOH-PCBs concentrations were OH-PCB for which no standards were available. Concentration of OH-PCBs in the trout plasma increased with increasing temperature and with the addition of CYP2B-like inducing congeners but not with the addition of CYP1A-inducing congeners to food. The results of this study provide the first in vivo evidence that rainbow trout are responsive to CYP2B-like induction by PCBs and that this enzyme system can influence PCB concentrations and OH-PCB formation in fish.     

Factors affecting phase I stereoselective biotransformation of chiral polychlorinated biphenyls by rat cytochrome P-450 2B1 isozyme

In vitro incubations of rat cytochrome P-450 (CYP) 2B1 isozyme with three chiral polychlorinated biphenyl (PCB) congeners (PCBs 45, 95, and 132) were performed to investigate factors affecting phase I stereoselective biotransformation. Rat CYP2B1 preferentially biotransformed the second-eluting atropisomers of PCBs 45 and 95 at low substrate concentration ranges (≤15 μM). Biotransformation competition by different congeners was also observed, with increasing competition at higher chlorination. Competition decreased the biotransformation rates of each congener stereoselectively, affecting atropisomeric composition. No atropisomeric enrichment was observed for PCB 132 upon incubation of the racemate. However, under the same conditions, significant differences in biotransformation kinetics were observed in individual atropisomer incubations, indicating that (+)-PCB 132 and (-)-PCB 132 were competitively biotransformed. Homology modeling and docking studies suggested that each atropisomer had different interactions with rat CYP2B1 and could dock with the isozyme at different locations. This is one possible explanation for stereoselective biotransformation and competition of chiral PCBs at the molecular level. Our results suggest that the lack of predictive capability for stereoselectivity of PCBs and other chiral pollutants in biota may be due to competitive and/or inhibitory activities of different substrates, including individual enantiomers of the same compound.     

Polychlorinated biphenyls and hydroxylated polychlorinated biphenyls in plasma of bottlenose dolphins (Tursiops truncatus) from the Western Atlantic and the Gulf of Mexico

Polychlorinated biphenyls (PCBs) and hydroxylated metabolic products (OH-PCBs) were measured in plasma collected from live-captured and released bottlenose dolphins (Tursiops truncatus) from five different locations in the Western Atlantic and the Gulf of Mexico in 2003 and 2004. In 2004, the sum (sigma) of concentration of PCBs in plasma of dolphins sampled off Charleston, SC [geometric mean: 223 ng/g of wet weight (w.w.)] was significantly higher (p<0.05) than concentrations detected in animals from the Indian River Lagoon, FL (sigmaPCBs: 122 ng/g w.w.) and the Sarasota Bay, FL (sigmaPCBs: 111 ng/g w.w.). The PCB homolog profiles were similar among locations. Concentrations of OH-PCBs were significantly higher (p<0.05) in plasma of dolphins from Charleston, SC (sigmaOH-PCBs for 2003: 126 ng/g w.w.; 2004: 138 ng/g w.w.) than animals from Florida (sigmaOH-PCBs ranged from 6 to 47 ng/g w.w.) and Bermuda (8.3 ng/g w.w.); however, concentrations in the Charleston samples did not differ from animals captured in Delaware Bay, NJ (57 ng/g w.w.). The sigmaOH-PCBs constituted 2-68% of the total PCB concentrations in plasma. Dichloro- to nonachloro-OH-PCBs were quantified using high-resolution gas chromatography mass spectrometry, but only around 20% of OH-PCBs could be identified by comparison to authentic standards. Results from this study show that OH-PCB are important environmental contaminants in dolphins and suggest that PCBs, decades after their ban, may still constitute a threat to wildlife.     

Effect of ionol on cytochrome P-450 induction in the liver of rats by polychlorinated biphenyls

A single oral administration of a mixture of polychlorinated diphenyls (50 and mg/kg) leads to a dose-dependent increase in the cytochrome P-450 content in rat liver. The synthetic antioxidant ionol administered singly and repeatedly (30 mg/kg) does not exert any substantial influence on the hemoprotein content in the liver. Preliminary administration of ionol potentiates the action of the mixture of polychlorinated diphenyls on cytochrome P-450 induction in the liver of experimental animals.     

Detection of hydroxylated polychlorinated biphenyls (OH-PCBs) in the abiotic environment: surface water and precipitation from Ontario, Canada

Hydroxylated PCBs (OH-PCBs) are well-known metabolites of PCBs in organisms, but there has been no direct study of their presence in the abiotic environment. In this study, OH-PCBs were determined in samples of rain, snow, and surface waters from sites in Ontario, Canada. OH-PCBs were quantified by gas chromatography with high-resolution mass spectrometry (GC-HRMS)in order to provide complete characterization of all OH-PCB homolog groups. OH-PCBs and PCBs were detected in all the samples analyzed, although half of the sigmaOH-PCBs could not be identified even with 71 individual congener standards. Total concentrations of OH-PCBs (sigmaOH-PCBs) in water ranged from 0.87 to 130 pg/L and from 230 to 990 pg/g in particulate organic matter. Total fluxes of those compounds in snow and rain were from < 1 to 100 pg/m2 and from < 1 to 44 pg/ m2/day, respectively. Higher sigmaOH-PCB fluxes in rain were found in southern Ontario than in a remote north-central Ontario site possibly reflecting greater sources of precursor PCBs near urban areas. Relatively higher sigmaOH-PCB concentrations were found in surface waters from sites near sewage treatment plant (STP) outfalls in the cities of Toronto (130 pg/L) and Hamilton (35 pg/L) than in offshore samples from Lake Ontario (1.6 pg/L). The results indicate that STPs are one of the sources of OH-PCBs for lake waters in this region. Similar homolog and congener profiles in rain and offshore surface water samples suggest that atmospheric deposition is the predominant source at offshore sites. This is the first report to detect the OH-PCBs in the abiotic environment.     

Morphologic changes and cytochrome P-450 induction in the liver of the rat after exposure to polychlorinated biphenyls in various diets

In is shown that administration of sovol, a mixture of polychlorinated diphenyls, in a dose of 500 mg/kg (about 1:10 LD50) induces pronounced changes in the rat liver structure, in particular, fat accumulation and diminution of RNA lump number in hepatocyte cytoplasma, alteration of nuclei, decrease in the number of hepatocytes and binuclear cells in the visual field. Structural disorders in the liver are observed during a long period (up to 5 months) and, to a certain extent, depend on the composition of the lipid component of the ration. Certain differences are recorded in the character of the morphological changes in the liver after single and repeated injections of the agent. No distinct relationship is recorded between the manifestation of hepatocyte fat infiltration and other morphological changes in the liver, and the index of microsomal cytochrome P-450 induction during sovol action.     

The effect of a lipid food component on enzymatic activity of rat liver endoplasmic reticulum upon the action of polychlorinated biphenyls

The authors have studied the character of changes in the content of cytochrome P450 and b5, in the oxidation rate of amidopyrine, dimethyl-aniline and aniline, in the NADPH- and ascorbate-dependent lipid peroxidation systems, as well as in glucose-6-phosphatase and acetylesterase activities in the liver microsomes of the rats on semisynthetic diets, including 50% (according to calorific value) of butter or sunflower oil, or receiving fat-free diet (0.5% of sunflower oil) in different terms (4 and 70 days) after a single intragastric administration of a mixture of polychlorinated diphenyls, chlorinated biphenyl (500 mg/kg). It is shown that the degree and character of the microsomal enzymes studied, as well as the changes in the liver structure under the action of chlorinated biphenyl depend, to a certain extent, on the quality and quantity of fat in the diet.     

Gas chromatographic analysis in the detection of anaerobes

By means of gaschromatography a rapid (ca 30 min) diagnosis of anaerobic infections is possible. The principle of a simple technique regarding the equipment and the laboratory-chemical process is presented.     

食品包装材料中丙烯酰胺在大鼠肝微粒和S9中的体外代谢研究

目的研究食品包装材料中丙烯酰胺(acrylamide)的体外代谢情况,并确证其代谢产物。方法采用肝微粒体及肝S9体外温孵法,优化代谢条件,对丙烯酰胺进行了体外代谢研究,并用液相色谱-串联质谱法(liquid chromatography-tandem mass spectrometry,LC-MS/MS)检测并确证丙烯酰胺的代谢物。结果通过检测结果发现,丙烯酰胺分别在肝S9和肝微粒体作用下发生显著的代谢反应,其代谢产物之一为环氧丙酰胺。结论丙烯酰胺能通过代谢转化成毒性更强的环氧丙酰胺,因此,监控食品、饮水和周围环境中的丙烯酰胺的含量对于维护人们的健康具有重要的意义。     

Enantiomeric fraction of selected chiral polychlorinated biphenyls in cow, goat, and ewe milk and dairy products by heart-cut multidimensional gas chromatography: first results

The concentration of the most relevant polychlorinated biphenyl (PCB) congeners has been determined in milk and dairy products (i.e., cheese and yogurt, from cow, goat, and ewe). In addition, the enantiomeric composition of 11 atropisomeric PCB has been determined for the first time in this type of product, using heart-cut multidimensional gas chromatography and 2 enantioselective columns for the unambiguous determination of both enantiomers. Results showed a deviation from the racemic composition for PCB 135, 136, 176, 171, and 183 in the samples analyzed. Whereas PCB 135, 136, and 176 showed an enrichment of the first eluted enantiomer, congeners 171 and 183 showed an enrichment of the first or second eluted enantiomer depending on the dairy product and species. The remaining congeners analyzed (PCB 84, 91, 95, 132, 149, and 174) did not present a clear enrichment of any of the enantiomers. Therefore, differences in the enantiomeric composition of some of the target atropisomeric PCB have been found among the 3 species and among the milk samples and the related dairy products analyzed. Enantioselective species-dependent processes, as well as enantioselective processes carried on by microorganisms during the fermentation and ripening in the latter could be possible explanations for the differences observed. However, changes in the enantiomeric composition are still not well understood, and further investigation in this direction is recommended.     

Measurement of polychlorinated biphenyls in fish tissue by gas chromatography with electron capture detection and enzyme-linked immunosorbent assay

Polychlorinated biphenyls in fish tissue were analyzed by enzyme-linked immunosorbent assay (ELISA) and gas chromatography with electron capture detection (GC/ECD) methods. Fish samples were collected in 2000 and 2001 during an Indiana fish survey. For samples with concentrations ranging from 0.05 to 0.5 ppm (n = 40) and 0.5 to 5.0 ppm (n = 12), the ELISA results were not significantly different from GC/ECD (P < 0.05). This research has demonstrated the effectiveness of analysis of fish samples by ELISA in the development of fish consumption advisories that accurately articulate health risks from the consumption of contaminated fish.     

酸消化-气相色谱法测定鱼肉中指示性 多氯联苯研究

目的建立酸消化-气相色谱法测定鱼肉中指示性多氯联苯的方法。方法用硫酸溶液消化样品,以正己烷+二氯甲烷(50:50,v:v)为提取溶液萃取、浓硫酸净化后用毛细管气相色谱法测定。结果采用该方法对鳕鱼中多氯联苯标准物质GBW(E)100131进行检测,检测结果与参考物定值吻合,回收率为93.7%~104.4%,相对标准偏差为2.75%~4.65%,检出限0.22~0.44μg/kg。结论该方法具有简单、快速、测定灵敏度高和精确度高的特点,能满足鱼肉中指示性多氯联苯分析的要求,值得推广应用。     

Effect of a phosphatidylcholine-enriched diet on the structural and functional parameters of rat liver microsomes

The authors studied the influence of the diet enriched with phosphatidylcholine on the structural-functional parameters of microsomes: lipid peroxidation activity, the levels of cytochromes b5 and P-450, microviscosity of the lipid phase of the membrane. In experiments in vivo it was found that phosphatidylcholine per se, not combined with antioxidants, did not show antioxidant properties. Biological effects of lecithin could be explained by its role as a catalyst of the monooxygenase system in the liver, based on the correlation (r = +0.8) between the high level of cytochrome b5 and the degree of eximerization of the fluorescent probe pyrene in the microsomes.     

加速溶剂萃取-气相色谱法检测鱼粉中7种指示性多氯联苯方法研究

目的 建立采用加速溶剂萃取-气相色谱检测鱼粉中7种指示性多氯联苯残留的方法。方法 采用加速溶剂萃取法, 以正己烷-二氯甲烷(1:1, V:V)为提取剂对鱼粉样品进行提取, 提取温度为120 ℃, 压力为10 MPa, 静态提取时间为8 min, 循环提取2次。用浓硫酸进行净化后, 用气相色谱法进行分析测定。结果 多氯联苯在5~500 ng/mL浓度范围内线性关系良好, 相关系数为0.9986~0.9999, 方法定量限为3.0~5.0 μg/kg, 3个添加水平的平均回收率为85.5%~104%, 相对标准偏差为2.61%~8.26%。结论 本方法精密度好, 准确度高, 操作简单、快速, 适用于鱼粉中多氯联苯的定量检测。     

Development of a screening analytical method for the determination of non-dioxin-like polychlorinated biphenyls in chicken eggs by gaschromatography and electron capture detection

ABSTRACT

A sensitive and reproducible screening analytical method is here proposed for the determination of six non dioxin-like polychlorinated biphenyls (NDL-PCBs, congener 28, 52, 101, 138, 153, 180) in chicken eggs based on accelerated solvent extraction (ASE) procedure for the fat extraction and determination, a solid phase extraction (SPE) sample clean-up process, and a gas chromatography – electron capture detection (GC-ECD) analysis. The optimized chromatographic separation, in less than 25 min, returned good responses for the six NDL-PCBs in the range of 2.5–60.0 µg L^?1, with correlation coefficients always higher than 0.9995. Instrumental limits of detection were between 0.08–0.35 µg L^?1, corresponding to 0.05 and 0.23 ng g^?1 fat in the matrix, while method detection limits, calculated on spiked egg samples, ranged from 1.6 to 3.5 ng g^?1 fat. The method has been extensively validated in terms of selectivity, sensitivity, recovery, precision, ruggedness and measurement uncertainty, following the European Directives.     

气相色谱内标法测定鱼肉中多氯联苯不确定度评定

目的建立一种气相色谱内标标准曲线法测定鱼肉中7种指示性多氯联苯不确定度的分析方法。方法依据JJF 1059.1-2012《测量不确定度评定与表示》,通过建立测量结果数学模型,系统分析实验过程的不确定度影响因素,对各不确定度来源进行了量化与合成,最终得到气相色谱内标标准曲线法测定鱼肉中多氯联苯不确定度的关键影响因素。结果标准曲线拟合、标准溶液配制、样品重复性测定以及样品中内标物的加入是鱼肉中多氯联苯测量不确定度的关键影响因素。结论本研究为水产品中多氯联苯不确定度分析提供了较为详尽的参考依据,更加有利于多氯联苯测量结果的科学评价和广泛应用。