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D H Phillips

Publications and source records attributed to D H Phillips.

At least 127 records · Page 7Linked to original sources
PubMed1992

Detection of polycyclic aromatic hydrocarbon-DNA adducts in white blood cells from coke oven workers: correlation with job categories.

Polycyclic aromatic hydrocarbon (PAH)-DNA adducts were quantitatively determined by ultrasensitive radioimmunoassay (USERIA) and 32P postlabeling in 128 DNA samples from WBCs of 68 coke oven workers and a local control group of 13 workers. Forty-four samples had a detectable adduct level by USERIA, with a mean of 0.390 fmol adducts/micrograms DNA (12.9 adducts/10(8) nucleotides) in the exposed group compared to a mean of 0.316 fmol adducts/micrograms DNA (10.4 adducts/10(8) nucleotides) in the control group. The mean adduct level with 32P postlabeling was 0.05 fmol/micrograms DNA (1.67 adducts/10(8) nucleotides) for the exposed group and 0.046 fmol/microgram DNA (1.54 adducts/10(8) nucleotides for the control group. Based on job description the workers were divided in 4 groups: control, low-, medium-, and high-exposure group. Both methods produced a positive correlation coefficient between estimated exposure and PAH-DNA adduct levels. The significance levels determined with Kendall rank correlation were P = 0.0145 for USERIA and P = 0.0594 for 32P postlabeling. Adduct levels determined by 32P postlabeling showed a correlation with tobacco smoking in the control group. No significant correlation between PAH-DNA adduct levels measured by USERIA and 32P postlabeling was found. These results show that these methods recognize different parts of the complex exposures in a coke oven plant.

Coke↗
PubMed1992

HPLC separation of 32P-postlabelled DNA adducts formed from dibenz[a,h]anthracene in skin.

Mouse skin and human skin have been treated in vivo or in short-term organ culture with dibenz[a,h]anthracene (DB[a,h]A), the related 3,4- or 5,6-diols or the anti- or syn-3,4-diol 1,2-oxides. DNA hydrolysates have been 32P-postlabelled and the adducts present examined by HPLC using a phenyl-modified reverse phase column and, for comparison, by PEI-cellulose TLC and autoradiography. The adducts formed when the diol-epoxides were reacted with salmon sperm DNA were also examined. The results show that in mouse skin treated in vivo, the major adducts formed from DB[a,h]A and the 3,4-diol were the same and that two of them were more polar than those formed in skin or in DNA that had been treated with the related anti- or syn-diol epoxides. Human skin treated with DB[a,h]A in culture yielded an adduct profile that was qualitatively similar to the profiles obtained with mouse skin.

Animals↗
PubMed1992

Genotoxic potential of tamoxifen and analogues in female Fischer F344/n rats, DBA/2 and C57BL/6 mice and in human MCL-5 cells.

Chronic administration of tamoxifen to female rats causes hepatocellular carcinomas. We have investigated damage to liver DNA caused by the administration of tamoxifen to female Fischer F344/N rats or C57B1/6 or DBA/2 mice using 32P-postlabelling. Following the administration of tamoxifen for 7 days (45 mg/kg/day) and extraction of hepatic DNA, up to 7 radiolabelled adduct spots could be detected after PEI-cellulose chromatography of the 32P-labelled DNA digests. Tamoxifen caused a time-dependent increase in the level of adduct detected up to a value of at least 1 adduct/10(6) nucleotides after 7 days dosing. A dose response relationship was demonstrated over the range of 5-45 mg/kg/day (0.013-0.12 mmol/kg/day). On cessation of dosing there was a loss of adducts from the liver DNA. These adducts were not detected in DNA from vehicle-dosed controls or in DNA from kidney, lung, spleen, uterus or peripheral lymphocytes. Pyrrolidinotamoxifen caused a similar level of adduct formation as tamoxifen. In contrast, no significant adduct formation could be detected in liver DNA from rats given droloxifene or toremifene. Mice given tamoxifen (45 mg/kg/day for 4 days) showed levels of adducts in the liver which were 30-40% of those present in rats. Exposure of rat hepatocytes to tamoxifen in vitro, resulted in induction of unscheduled DNA synthesis, when preparations from rats which had been pretreated with tamoxifen in vivo were used. No such increase could be detected in hepatocytes from control rats, suggesting tamoxifen may induce enzymes responsible for its own activation. Tamoxifen induced a significant increase in micronucleus formation in a dose dependent manner in cultures of MCL-5 cells, a human cell line that expresses 5 different human cytochrome P450 isoenzymes, as well as epoxide hydrolase.

Animals↗
PubMed1992

Metabolic activation of the food mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) to DNA binding species in human mammary epithelial cells.

When incubated in suspension with the heterocyclic aromatic amine food mutagens 2-amino-3-methylimidazo [4,5-f]-quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), human mammary epithelial cell aggregates were found, by 32P-postlabelling analysis, to yield DNA that contained adducts. Analysis by HPLC of the 32P-labelled digests of mammary cell DNA indicated that in each case a major adduct peak corresponded to that produced in DNA in vitro by activated derivatives of the two compounds. The patterns of adducts obtained when DNA digests were separated by TLC on polyethyleneimine-cellulose plates were found to resemble those previously shown to be present in DNA of tissues of mice fed IQ or MeIQ. These results demonstrate the ability of human mammary epithelial cells to activate carcinogenic heterocyclic compounds known to be present in the human diet to DNA binding derivatives.

Biotransformation↗
PubMed1992

Detection and characterization by 32P-postlabelling of DNA adducts induced by a Fenton-type oxygen radical-generating system.

Reactive oxygen species can give rise to numerous modifications of DNA. We have investigated the formation of such modifications using the nuclease P1 digestion method of the 32P-postlabelling procedure for the detection of DNA damage. Analysis of DNA that had been treated with a Fenton-type system of copper (or iron) ions and H2O2 resulted in the detection of up to ten discrete 32P-labelled spots, displaying chromatographic characteristics similar to aromatic adducts, on PEI-cellulose TLC. Maximum total levels equivalent to 28 adducts/10(8) nucleotides were achieved after 15 min of treatment with Cu2+/H2O2. The formation of adducts was 1.5 times greater if single-stranded rather than double-stranded DNA was employed, suggesting an intrastrand effect. Experiments with 3'-deoxyribonucleotides demonstrated that the adducts detected did not represent base modifications such as 8-hydroxydeoxyguanosine or thymidine glycols. However, treatment of specific dinucleotides (dApdG and dApdA) was found to produce two major adducts that were chromatographically identical by TLC and HPLC to the two major adducts formed in DNA. It is proposed that these species with aromatic adduct-like characteristics are the result of the intrastrand linking of specific adjacent bases in DNA.

Animals↗
PubMed1992

Interaction of omeprazole with DNA in rat tissues.

There has been concern recently over the possible genotoxicity of omeprazole, a potent inhibitor of gastric acid secretion in humans. In order to investigate its possible DNA binding activity in vivo, 14C-labelled omeprazole was administered to male PVG rats, 8-10 weeks old, orally by gavage at a dose of 30 mg/kg. At various times after treatment, animals were killed and DNA isolated from the fundic and antral regions of the glandular stomach, and from the duodenum, ileum and colon. Scintillation counting of the DNA samples revealed a reproducible association of 14C that was not extractable with organic solvents and could not be removed from the DNA by column cartridge chromatography. DNA samples were digested enzymically to deoxyribonucleosides and analysed by reversed-phase high performance liquid chromatography (HPLC). With DNA from each tissue, most of the radioactivity eluted in a peak that was chromatographically distinct from normal nucleosides and from omeprazole itself. Chromatography following partial digestion of DNA gave rise to the same radioactivity elution profiles, whilst chromatography on hydroxyapatite resulted in retention of most of the radioactivity on the column using conditions under which DNA was eluted. These findings suggest either the formation in rat tissues of chemically-labile covalent omeprazole-DNA adducts or, more likely, a strong non-covalent interaction. The apparent DNA binding occurs rapidly in vivo and is short-lived, the maximum levels of radioactive incorporation (equivalent of up to 36 pmol omeprazole/mg DNA) occurring at 0.5 h in fundus, 0.5-1 h in antrum, 1 h in duodenum, 2 h in ileum and at 4-8 h in colon.

Animals↗
PubMed1991

Improved reversed-phase high-performance liquid chromatographic separation of 32P-labelled nucleoside 3',5'-bisphosphate adducts of polycyclic aromatic hydrocarbons.

32P-Postlabelling is a sensitive technique for the detection and analysis of carcinogen-DNA adducts. In this paper we describe the development of an improved high-performance liquid chromatography (HPLC) method for the separation of 32P-labelled 3',5'-bisphosphates of nucleosides modified by reactive derivatives of carcinogenic polycyclic aromatic hydrocarbons (PAH). Optimal resolution of the major 32P-postlabelled DNA adducts formed by the anti-diol-epoxides of ten PAH was achieved using a phenyl-modified silica gel column with a gradient of methanol in phosphate buffer at low pH and high ionic strength. Use of a radioactivity flow detector coupled to the HPLC apparatus allowed detection of subfemtomole quantities of labelled adducts.

Carcinogens↗
PubMed1991

DNA adducts in human and mouse skin maintained in short-term culture and treated with petrol and diesel engine lubricating oils.

Human and mouse skin samples maintained in short-term organ culture were treated topically with used engine oils from petrol- and diesel-powered vehicles. Mice were also treated topically in vivo for comparison. DNA was isolated and analysed by 32P-postlabelling and the labeled DNA digests were resolved on polyethyleneimine-cellulose tlc sheets. A large number of radioactive adduct spots were observed in DNA from skin treated with the used petrol-engine oil, indicating the formation of adducts by many components of the complex oil mixture. Total adduct levels were similar in mouse skin (both in vivo and in vitro) and in human skin, although qualitative differences in the adduct maps were apparent between the human and mouse skin DNA. Treatment with the used diesel engine oil produced adduct levels no greater than that of control samples in mouse skin (in vivo and in vitro), although significant levels were found in human skin DNA from one donor. The results correlate well with the carcinogenic activity of these oils in experimental animals, helping to substantiate the conclusion that petrol engine oils (but not diesel engine oils) may present a carcinogenic risk to man if appropriate measures to minimise skin contact are not observed.

Animals↗
PubMed1991

The in vitro metabolic activation of dibenz[a,h]anthracene, catalyzed by by rat liver microsomes and examined by 32P-postlabelling.

DNA has been incubated in vitro with dibenz[a,h]anthracene (DB[a,H]A) and the related 5,6-diol and 3,4-diol in the presence of 3-methylcholanthrene- or Aroclor 1254-induced rat liver microsomes. After incubation, the DNA was extracted and examined for the presence of aromatic adducts using the nuclease P1 modification of the 32P-postlabelling technique. The maps of PEI-cellulose plates and autoradiography showed that 92% of the radioactivity contained in DB[a,h]A-DNA adduct spots is derived from the related 3,4-diol and that about 50% of the adducts may be formed following the conversion of this diol to the bay-region anti- and syn-3,4-diol 1,2-oxides.

Animals↗
PubMed1991

Relationship between mutagenicity and DNA adduct formation in mammalian cells for fjord- and bay-region diol-epoxides of polycyclic aromatic hydrocarbons.

Chinese hamster V79 cells were treated with the anti- and syn-diastereomers of the bay- or fjord-region diol-epoxides of four polycyclic aromatic hydrocarbons, namely benzo[a]pyrene (BP), benzo[c]chrysene (BcC), benzo[g]chrysene (BgC) and benzo[c]phenanthrene (BcPh). The frequency of induction of 6-thioguanine-resistant mutations was determined, and the extent of formation of DNA adducts was measured by 32P-postlabelling. When expressed as mutation frequency per nanomoles compound per millilitre incubation medium, this group of chemicals expressed a 160-fold range in potency. In agreement with previous experimental studies, the anti-diol-epoxide of BcC was highly mutagenic, inducing in excess of 3 x 10(4) mutations/10(6) cells per nmol compound/ml. The mutagenic activities of the anti- and syn-diol-epoxides of BP were 10- and 100-fold lower, respectively. Both diol-epoxides of BgC, the syn-BcC and the anti-BcPh derivatives were also highly mutagenic, and only the syn-BcPh diol-epoxide was less mutagenic than the anti-diol-epoxide of BP. Determination of the levels of DNA adducts formed by the diol-epoxides indicated that the most mutagenic compounds were the most DNA reactive, although the fjord-region diol-epoxides gave rise to more complex patterns of adducts than those of the BP diol-epoxides. When the mutagenicity results were expressed as mutations per femtomoles total adducts formed, all compounds showed similar activities. Thus the potent mutagenicity of the fjord region diol-epoxides appears to be due to the high frequency with which they form DNA adducts in V79 cells, rather than to formation of adducts with greater mutagenic potential.

Animals↗
PubMed1991

Microsomal metabolism of dibenz[a,c]anthracene, dibenz[a,h]anthracene and dibenz[a,j]anthracene to bis-dihydrodiols and polyhydroxylated products.

Polar, ethyl acetate soluble metabolites formed in incubations of dibenz[a,c]anthracene (DB[a,c]A), dibenz[a,h]anthracene (DB[a,h]A) and the related DB[a,h]A 3,4-diol and dibenz[a,j]anthracene (DB[a,j]A) with 3-methylcholanthrene (3-MC)-induced rat liver microsomal preparations have been separated by HPLC and examined using fluorescence, UV and NMR spectroscopy. Metabolites with spectral properties consistant with their identification as the 3,4:8,9-bis-diol of DB[a,j]A and a 1,2,3,4,12,13-hexol derived from DB[a,c]A were found. DB[a,h]A was metabolized to three polar products identified as the 3,4:10,11-bis-diol and the related 1,2,3,4,8,9- and 1,2,3,4,10,11-hexols, which were also formed, together with the related 1,2,3,4-tetrol, from the DB[a,h]A 3,4-diol. The possible role of bis-diols in the metabolic activation of these three dibenzanthracenes is discussed.

Animals↗
PubMed1991

32P-postlabelling detection of aromatic DNA adducts in peripheral blood lymphocytes from aluminium production plant workers.

Aluminium production plant workers are exposed to a great number of airborne polycyclic aromatic hydrocarbons and epidemiological studies suggest that these workers are at increased risk of lung and bladder cancer. Blood samples from 46 workers at 2 primary aluminium plants and from 29 occupationally unexposed control individuals were analysed. DNA was isolated from the peripheral blood lymphocytes and aromatic DNA adducts were detected by 32P-postlabelling assay using the nuclease P1 digestion procedure for the enrichment of the adducts. The total levels of DNA adducts of exposed individuals varied from the detection limit of about 0.5 adducts/10(8) nucleotides up to 7.1 adducts/10(8) nucleotides and control adduct levels were up to 2.42 adducts/10(8) nucleotides. There was no significant difference between the mean adduct levels of the control group and of the individuals of one plant. However, the mean DNA adduct level obtained from workers of the second plant was significantly higher than that of the controls (p less than 0.001) and of the first plant (p less than 0.01), respectively. This difference can be attributed to differences in the design of technology and different levels of exposure at the 2 plants. The results of this study encourage further investigations of the use of peripheral white blood cells as marker cells and of 32P-postlabelling analysis for monitoring occupational exposure to mixtures of environmental carcinogenic pollutants.

Adult↗
PubMed1991

The metabolic activation of dibenz[a,h]anthracene in mouse skin examined by 32P-postlabelling: minor contribution of the 3,4-diol 1,2-oxides to DNA binding.

Dibenz[a,h]anthracene (DB[a,h]A) and the related 3,4-diol and anti- and syn-3,4-diol 1,2-oxides were applied to the shaved dorsal skin of groups of four C57Bl/CB1 mice. Twenty-four hours later the mice were killed, DNA isolated from the treated skin, hydrolysed and examined for the presence of aromatic adducts using the nuclease P1 modification of the 32P-postlabelling technique. Autoradiography of the maps obtained by chromatography on polyethyleneimine-cellulose plates showed that six DNA adduct spots that were derived from DB[a,h]A were also present in the DNA of skin treated with the DBA 3,4-diol and that, whilst four of these adduct spots were also seen in maps prepared from the DNA of skin treated with the anti-3,4-diol-1,2-oxide, they were not present in DNA from skin to which the syn-isomer had been applied. The identity of these adduct spots was confirmed by their coincidence when mixtures of different DNA hydrolysates were chromatographed together. Quantitatively, the highest levels of mouse skin modification were obtained with the diol-epoxides and the lowest with DB[a,h]A. The results suggest that most of the DNA adducts formed in DB[a,h]A-treated mouse skin arise through metabolism of the hydrocarbon to the related 3,4-diol and that some may be formed following the conversion of this diol to the bay-region anti-3,4-diol-1,2-oxide.

Animals↗
PubMed1991

Evaluation of DNA-binding activity of hydroxyanthraquinones occurring in Rubia tinctorum L.

The naturally-occurring anthraquinones (AQs), alizarin (1,2-dihydroxyanthraquinone) and lucidin (1,3-dihydroxy-2-hydroxymethylanthraquinone), were incubated with DNA in the presence of S9 mix. The isolated DNA was analysed by 32P-postlabelling for the presence of aromatic adducts. Only lucidin formed up to five different DNA adducts in the range from 0.995 to 3.05 adducts/10(8) nucleotides. Lucidin was also incubated with polynucleotides poly[d(A-T)] and polydG*polydC in the presence of S9 mix. Analysis of polydG*polydC revealed a similar adduct pattern to that obtained with lucidin-modified DNA. Alizarin, lucidin, a glycoside mixture containing alizarinprimeveroside and lucidinprimeveroside, and Rubia Teep (a herbal drug made from Rubia tinctorum containing lucidin) were incubated with primary rat hepatocytes for 24 h and the isolated DNA was analysed by 32P-postlabelling. Lucidin, the glycoside mixture and Rubia Teep gave rise to DNA adducts, but alizarin did not. Male Parkes mice were treated orally for 4 days with alizarin (10 mg/d), lucidin (2 mg/d), the glycoside mixture (20 mg/d) or Rubia Teep (1/2 tablet/d) and DNA was isolated from liver, kidney, duodenum and colon. Analysis by 32P-postlabelling revealed that lucidin, the glycoside mixture and Rubia Teep, but not alizarin, formed DNA adducts in all the tissues examined but that the adduct patterns were organ-specific.

Animals↗
PubMed1990

Influence of cigarette smoking on the levels of DNA adducts in human bronchial epithelium and white blood cells.

The presence of carcinogen-DNA adducts in human tissues is evidence of exposure to carcinogens and may be an indicator of cancer risk. DNA was isolated from non-tumorous bronchial tissue of 37 cigarette smokers, 8 former smokers and 8 non-smokers and analyzed for the presence of aromatic and/or hydrophobic DNA adducts in the 32P-post-labelling assay. Adducts were detected as bands of radioactive material when 5'-32P-labelled deoxyribonucleoside 3',5'-bisphosphates were chromatographed on polyethyleneimine-cellulose tlc plates, and the patterns indicated the formation of adducts by a large number of compounds. Adduct levels detected in DNA from non-smokers, former smokers and current smokers were 3.45 +/- 1.62, 3.93 +/- 1.92 and 5.53 +/- 2.13 adducts/10(8) nucleotides, respectively. The differences in adduct levels between smokers and former and non-smokers were statistically significant (p less than 0.01); and among the smokers, significant correlations were found between adduct levels and both daily cigarette consumption and total cigarette consumption (daily consumption X number of years smoked). DNA was also isolated from the peripheral-blood leukocytes of 31 heavy smokers (greater than 20 cigarettes/day) and 20 non-smokers and analyzed by 32P-post-labelling. Adduct levels in the smokers' samples were not significantly different from levels in the non-smokers' samples (2.53 +/- 1.31 and 2.12 +/- 1.44 adducts/10(8) nucleotides, respectively). Thus, evidence for carcinogen exposure was found in human bronchial epithelium, a target tissue for tobacco-induced tumour formation, but not in peripheral-blood cells, indicating possible limitations in the use of the latter as a surrogate, non-target tissue source of DNA for monitoring human exposure to inhaled carcinogens.

Adult↗
PubMed1990

Aromatic DNA adducts in white blood cells of coke workers.

White blood cell DNA adducts were measured in coke workers, local controls and countryside controls using the 32P-postlabelling technique. The method detected aromatic adducts including those formed by polycyclic aromatic hydrocarbons (PAHs). Coke workers are heavily exposed to PAHs particularly when working at the batteries. A difference in adduct levels was noted between the coke workers at the battery as compared to other jobs. The adduct levels in the non-battery were higher than those in the countryside controls.

Air Pollutants, Occupational↗