Naphthalene Metabolites Research Articles

Cumulative body burdens of polycyclic aromatic hydrocarbons associated with estrogen bioactivation in pregnant women: Protein adducts as biomarkers of exposure

The objective of this research was to simultaneously analyze protein adducts of quinonoid metabolites of naphthalene and endogenous estrogen in serum albumin (Alb) derived from healthy pregnant women in Taiwan and to explore the correlations among them. The isomeric forms of cysteinyl adducts of naphthoquinones, including 1,2-naphthoquinone (1,2-NPQ) and 1,4-naphthoquinone (1,4-NPQ) as well as estrogen quinones, including estrogen-2,3-quinones (E2-2,3-Q) and estrogen-3,4-quinones (E2-3,4-Q), are characterized after adduct cleavage. Results showed that the median levels of cysteinyl adducts of 1,2-NPQ and 1,4-NPQ on serum albumin were 249-390 and 16.0–24.8 pmol g−1, respectively. Logged levels of 1,2-NPQ-Alb were correlated with logged levels of 1,4-NPQ-Alb (correlation coefficient r = 0.551, P < 0.001). Cysteinyl adducts of E2-2,3-Q-1-S-Alb, E2-2,3-Q-4-S-Alb, and E2-3,4-Q-2-S-Alb were detected in all subjects with median levels at 275-435, 162-288, and 197-254 pmol g−1, respectively. We also found a positive relationship between logged levels of E2-2,3-Q-4-S-Alb and those of E2-3,4-Q-2-S-Alb (r = 0.770, P < 0.001).We noticed that median levels of E2-2,3-Q-derived adducts (E2-2,3-Q-1-S-Alb plus E2-2,3-Q-4-S-Alb) in pregnant women were greater than those of E2-3,4-Q-2-S-Alb (∼2–3-fold). Taken together, this evidence lends further support to the theme that cumulative concentration of E2-3,4-Q is a significant predictor of the risk of breast cancer. Furthermore, we noticed that levels of 1,2-NPQ-Alb are positively associated with levels of E2-3,4-Q-2-S-Alb (r = 0.522, P < 0.001) and those of E2-2,3-Q-4-S-Alb (r = 0.484, P < 0.001). Overall, this evidence suggests that environmental exposure to polycyclic aromatic hydrocarbons may modulate estrogen homeostasis and enhance the production of reactive quinone species of endogenous estrogen in humans.

Urinary Polycyclic Aromatic Hydrocarbons and Childhood Obesity: NHANES (2001–2006)

Background: Polycyclic aromatic hydrocarbons (PAHs) are known carcinogens and suspected endocrine disruptors. Prenatal exposure to PAHs has been associated with obesity in early childhood.Objective: We examined the association of urinary PAH metabolites with adiposity outcomes [body mass index (BMI) z-score, waist circumference (WC), and rate of obesity] in children and adolescents.Methods: We performed whole-sample analyses of 3,189 individuals 6–19 years of age who participated in the 2001–2006 National Health and Nutrition Examination Survey. We performed multivariate linear and logistic regression to analyze the association of BMI z-score, WC, and obesity with concentrations of single urinary PAH compounds and the sum of PAHs. Furthermore, the analyses were stratified by developmental stage [i.e., children (6–11 years) and adolescents (12–19 years)].Results: BMI z-score, WC, and obesity were positively associated with the molecular mass sum of the PAHs and the total sum of naphthalene metabolites. Most associations increased monotonically with increasing quartiles of exposure among children 6–11 years of age, whereas dose–response trends were less consistent for adolescents (12–19 years of age). Neither total PAHs nor total naphthalene metabolites were associated with overweight in either age group, and there was little evidence of associations between the outcomes and individual PAHs.Conclusions: Total urinary PAH metabolites and naphthalene metabolites were associated with higher BMI, WC, and obesity in children 6–11 years of age, with positive but less consistent associations among adolescents.Citation: Scinicariello F, Buser MC. 2014. Urinary polycyclic aromatic hydrocarbons and childhood obesity: NHANES (2001–2006). Environ Health Perspect 122:299–303; http://dx.doi.org/10.1289/ehp.1307234

Identification of naphthalene metabolism by white rot fungus Pleurotus eryngii

The use of biomaterials or microorganisms in PAHs degradation had presented an eye-catching performance. Pleurotus eryngii is a white rot fungus, which is easily isolated from the decayed woods in the tropical rain forest, used to determine the capability to utilize naphthalene, a two-ring polycyclic aromatic hydrocarbon as source of carbon and energy. In the meantime, biotransformation of naphthalene to intermediates and other by-products during degradation was investigated in this study. Pleurotus eryngii had been incubated in liquid medium formulated with naphthalene for 14days. The presence of metabolites of naphthalene suggests that Pleurotus eryngii begin the ring cleavage by dioxygenation on C1 and C4 position to give 1,4-naphthaquinone. 1,4-Naphthaquinone was further degraded to benzoic acid, where the proposed terepthalic acid is absent in the cultured extract. Further degradation of benzoic acid by Pleurotus eryngii shows the existence of catechol as a result of the combination of decarboxylation and hydroxylation process. Unfortunately, phthalic acid was not detected in this study. Several enzymes, including manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase and 2,3-dioxygenase are enzymes responsible for naphthalene degradation. Reduction of naphthalene and the presence of metabolites in liquid medium showed the ability of Pleurotus eryngii to utilize naphthalene as carbon source instead of a limited glucose amount.

Characterization of Model Peptide Adducts with Reactive Metabolites of Naphthalene by Mass Spectrometry

Naphthalene is a volatile polycyclic aromatic hydrocarbon generated during combustion and is a ubiquitous chemical in the environment. Short term exposures of rodents to air concentrations less than the current OSHA standard yielded necrotic lesions in the airways and nasal epithelium of the mouse, and in the nasal epithelium of the rat. The cytotoxic effects of naphthalene have been correlated with the formation of covalent protein adducts after the generation of reactive metabolites, but there is little information about the specific sites of adduction or on the amino acid targets of these metabolites. To better understand the chemical species produced when naphthalene metabolites react with proteins and peptides, we studied the formation and structure of the resulting adducts from the incubation of model peptides with naphthalene epoxide, naphthalene diol epoxide, 1,2-naphthoquinone, and 1,4-naphthoquinone using high resolution mass spectrometry. Identification of the binding sites, relative rates of depletion of the unadducted peptide, and selectivity of binding to amino acid residues were determined. Adduction occurred on the cysteine, lysine, and histidine residues, and on the N-terminus. Monoadduct formation occurred in 39 of the 48 reactions. In reactions with the naphthoquinones, diadducts were observed, and in one case, a triadduct was detected. The results from this model peptide study will assist in data interpretation from ongoing work to detect peptide adducts in vivo as markers of biologic effect.

Urinary naphthol metabolites and chromosomal aberrations in 5-year-old children.

Exposure to naphthalene, an International Agency for Research on Cancer (IARC)-classified possible carcinogen and polycyclic aromatic hydrocarbon (PAH), is widespread, though resulting health effects are poorly understood. Metabolites of naphthalene, 1- and 2-naphthol, are measurable in urine and are biomarkers of personal exposure. Chromosomal aberrations, including translocations, are established markers of cancer risk and a biodosimeter of clastogenic exposures. Although prenatal (maternal) PAH exposure predicts chromosomal aberrations in cord blood, few studies have examined chromosomal aberrations in school-age children and none has examined their association with metabolites of specific PAHs. Using Whole Chromosome Paint Fluorescent in situ Hybridization, we documented chromosomal aberrations including translocations, in 113 five-year-old urban minority children and examined their association with concurrent concentrations of PAH metabolites measured in urine. We report that in lymphocytes, the occurrence and frequency of chromosomal aberrations including translocations are associated with levels of urinary 1- and 2-naphthol. When doubling the levels of urinary naphthols, gender-adjusted OR for chromosomal aberrations are 1.63 [95% confidence interval (CI), 1.21-2.19] and 1.44 (95% CI, 1.02-2.04) for 1- and 2-naphthol, respectively; and for translocations OR = 1.55 (95% CI, 1.11-2.17) and 1.92 (95% CI, 1.20-3.08) for 1- and 2-naphthol, respectively. Our results show that markers of exposure to naphthalene in children are associated with translocations in a dose-related manner, and that naphthalene may be a clastogen. Indoor exposure to elevated levels of naphthalene is prevalent in large regions of the world. This study is the first to present an association between a marker of naphthalene exposure and a precarcinogenic effect in humans.

Analysis of naphthalene adduct binding sites in model proteins by tandem mass spectrometry

The electrophilic metabolites of the polyaromatic hydrocarbon naphthalene have been shown to bind covalently to proteins and covalent adduct formation correlates with the cytotoxic effects of the chemical in the respiratory system. Although 1,2-naphthalene epoxide, naphthalene diol epoxide, 1,2-naphthoquinone, and 1,4-napthoquinone have been identified as reactive metabolites of interest, the role of each metabolite in total covalent protein adduction and subsequent cytotoxicity remains to be established. To better understand the target residues associated with the reaction of these metabolites with proteins, mass spectrometry was used to identify adducted residues following (1) incubation of metabolites with actin and protein disulfide isomerase (PDI), and (2) activation of naphthalene in microsomal incubations containing supplemental actin or PDI. All four reactive metabolites bound to Cys, Lys or His residues in actin and PDI. Cys17 of actin was the only residue adducted by all metabolites; there was substantial metabolite selectivity for the majority of adducted residues. Modifications of actin and PDI, following microsomal incubations containing 14C-naphthalene, were detected readily by 2D gel electrophoresis and phosphor imaging. However, target modifications on tryptic peptides from these isolated proteins could not be readily detected by MALDI/TOF/TOF and only three modified peptides were detected using high resolution–selective ion monitoring (HR–SIM). All the reactive metabolites investigated have the potential to modify several residues in a single protein, but even in tissues with very high rates of naphthalene activation, the extent of modification was too low to allow unambiguous identification of a significant number of modified residues in the isolated proteins.

Exposure to polycyclic aromatic hydrocarbons and serum inflammatory markers of cardiovascular disease

Polycyclic aromatic hydrocarbons (PAHs) are environmental and occupational carcinogens produced by the incomplete combustion of organic materials, such as coal and petroleum product combustion, tobacco smoking, and food cooking, that may be significant contributors to the burden of cardiovascular disease in human populations. The purpose of this study was to investigate associations between ten monohydroxy urinary metabolites of four PAHs and three serum biomarkers of cardiovascular disease (fibrinogen, homocysteine, and white blood cell count). Using data on 3219 participants aged 20 years and older from the National Health and Nutrition Examination Survey (NHANES) 2001–2004 dataset, the associations between PAH metabolites and serum inflammatory markers were analyzed using the Spearman correlations and multiple linear regression modeling. The PAH metabolites of naphthalene, fluorene, phenanthrene, and pyrene each showed both positive and negative correlations with homocysteine, fibrinogen, and white blood cell count (correlation coefficient range: −0.077–0.143) in nonsmoking participants. Using multiple linear regression models adjusted for age, gender, race/ethnicity, and body mass index, estimates of weighted geometric means of inflammatory marker levels were not significantly different between high and low levels (75th vs. 25th percentiles) for all PAH metabolites in nonsmoking subjects. The results of this study do not provide evidence for a relationship between PAH exposure (as measured by urinary levels of PAH metabolites) and serum biomarkers of cardiovascular disease after controlling for tobacco use.

Reprint of “Update of the reference and HBM values derived by the German Human Biomonitoring Commission”

In 2007, we reviewed the working principles and working procedures of the German Human Biomonitoring Commission together with the reference values and human biomonitoring (HBM) values derived up to that time. Since then, the Commission has decided to derive additionally HBM I values on the basis of tolerable daily intakes and has used and evaluated this new approach on the metabolites of (2-ethylhexyl) phthalate (DEHP) in urine. Furthermore, the Commission has derived a HBM I value for thallium in urine, has recinded the HBM values for lead in blood, and has updated the HBM values for cadmium in urine. Based on the representative data of the German Environmental Survey on Children from 2003 to 2006 (GerES IV), the Commission has updated the reference values for a large number of environmental pollutants in urine and blood of children in Germany. Since 2007, the Commission has derived new and updated reference values for PFOS and PFOA in human plasma, for thallium in urine, for aromatic amines in urine, for a comprehensive number of phthalate metabolites in urine, and for organochlorine pesticides in human breast milk. Furthermore, the Commission has evaluated background exposure levels for two naphthalene metabolites and acrylamide (using acrylamide-haemoglobin adduct) for the general population. This paper reports the new values, including those already published, in order to provide an updated overview.

Influence of pH and surface oxygen-containing groups on multiwalled carbon nanotubes on the transformation and adsorption of 1-naphthol

The pH-dependent behavior, including the transformation of 1-naphthol by oxidative polymerization to form precipitates in solution and the adsorption of 1-naphthol onto carbon nanotubes (CNTs), was examined. Neglecting the precipitate loss of 1-naphthol and possibly of similar chemicals may result in the overestimation of their adsorption and inadequate interpretation of the underlying adsorption mechanisms. Surface oxygen-containing groups on CNTs and the dissociated species of these groups can interact with the dissociated and neutral species of 1-naphthol in a way similar to polymerization, thus promoting the adsorption of 1-naphthol onto CNTs. Adsorption onto CNTs may reduce the polymeric precipitates of 1-naphthol in solution by possibly decreasing aqueous 1-naphthol concentrations. These observations and the underlying mechanisms are important for predicting the fate and risks of naphthalene and carbaryl in the environment because 1-naphthol is a primary metabolite of naphthalene and carbaryl. In addition, it is possible to enhance the removal of 1-naphthol and similar chemicals by controlling the pH and designing specific surface functional groups for CNTs.

Urinary biomarkers of occupational jet fuel exposure among air force personnel

There is a potential for widespread occupational exposure to jet fuel among military and civilian personnel. Urinary metabolites of naphthalene have been suggested for use as short-term biomarkers of exposure to jet fuel (jet propulsion fuel 8 (JP8)). In this study, urinary biomarkers of JP8 were evaluated among US Air Force personnel. Personnel (n=24) were divided a priori into high, moderate, and low exposure groups. Pre- and post-shift urine samples were collected from each worker over three workdays and analyzed for metabolites of naphthalene (1- and 2-naphthol). Questionnaires and breathing-zone naphthalene samples were collected from each worker during the same workdays. Linear mixed-effects models were used to evaluate the exposure data. Post-shift levels of 1- and 2-naphthol varied significantly by a priori exposure group (levels in high group>moderate group>low group), and breathing-zone naphthalene was a significant predictor of post-shift levels of 1- and 2-naphthol, indicating that for every unit increase in breathing-zone naphthalene, there was an increase in naphthol levels. These results indicate that post-shift levels of urinary 1- and 2-naphthol reflect JP8 exposure during the work-shift and may be useful surrogates of JP8 exposure. Among the high exposed workers, significant job-related predictors of post-shift levels of 1- and 2-naphthol included entering the fuel tank, repairing leaks, direct skin contact with JP8, and not wearing gloves during the work-shift. The job-related predictors of 1- and 2-naphthol emphasize the importance of reducing inhalation and dermal exposure through the use of personal protective equipment while working in an environment with JP8.

The Chemical Biology of Naphthoquinones and Its Environmental Implications

Quinones are a group of highly reactive organic chemical species that interact with biological systems to promote inflammatory, anti-inflammatory, and anticancer actions and to induce toxicities. This review describes the chemistry, biochemistry, and cellular effects of 1,2- and 1,4-naphthoquinones and their derivatives. The naphthoquinones are of particular interest because of their prevalence as natural products and as environmental chemicals, present in the atmosphere as products of fuel and tobacco combustion. 1,2- and 1,4-naphthoquinones are also toxic metabolites of naphthalene, the major polynuclear aromatic hydrocarbon present in ambient air. Quinones exert their actions through two reactions: as prooxidants, reducing oxygen to reactive oxygen species; and as electrophiles, forming covalent bonds with tissue nucleophiles. The targets for these reactions include regulatory proteins such as protein tyrosine phosphatases; Kelch-like ECH-associated protein 1, the regulatory protein for NF-E2-related factor 2; and the glycolysis enzyme glyceraldehyde-3-phosphate dehydrogenase. Through their actions on regulatory proteins, quinones affect various cell signaling pathways that promote and protect against inflammatory responses and cell damage. These actions vary with the specific quinone and its concentration. Effects of exposure to naphthoquinones as environmental chemicals can vary with the physical state, i.e., whether the quinone is particle bound or is in the vapor state. The exacerbation of pulmonary diseases by air pollutants can, in part, be attributed to quinone action.

Update of the reference and HBM values derived by the German Human Biomonitoring Commission

In 2007, we reviewed the working principles and working procedures of the German Human Biomonitoring Commission together with the reference values and human biomonitoring (HBM) values derived up to that time. Since then, the Commission has decided to derive additionally HBM I values on the basis of tolerable daily intakes and has used and evaluated this new approach on the metabolites of (2-ethylhexyl) phthalate (DEHP) in urine. Furthermore, the Commission has derived a HBM I value for thallium in urine, has recinded the HBM values for lead in blood, and has updated the HBM values for cadmium in urine. Based on the representative data of the German Environmental Survey on Children from 2003 to 2006 (GerES IV), the Commission has updated the reference values for a large number of environmental pollutants in urine and blood of children in Germany. Since 2007, the Commission has derived new and updated reference values for PFOS and PFOA in human plasma, for thallium in urine, for aromatic amines in urine, for a comprehensive number of phthalate metabolites in urine, and for organochlorine pesticides in human breast milk. Furthermore, the Commission has evaluated background exposure levels for two naphthalene metabolites and acrylamide (using acrylamide-haemoglobin adduct) for the general population. This paper reports the new values, including those already published, in order to provide an updated overview.

Impact of the Improved Patsari Biomass Stove on Urinary Polycyclic Aromatic Hydrocarbon Biomarkers and Carbon Monoxide Exposures in Rural Mexican Women

Background: Cooking with biomass fuels on open fires results in exposure to health-damaging pollutants such as carbon monoxide (CO), polycyclic aromatic hydrocarbons (PAHs), and particulate matter.Objective: We compared CO exposures and urinary PAH biomarkers pre- and postintervention with an improved biomass stove, the Patsari stove.Methods: In a subsample of 63 women participating in a randomized controlled trial in central Mexico, we measured personal CO exposure for 8 hr during the day using continuous monitors and passive samplers. In addition, first-morning urine samples obtained the next day were analyzed for monohydroxylated PAH metabolites by gas chromatography/isotope dilution/high-resolution mass spectrometry. Exposure data were collected during the use of an open fire (preintervention) and after installation of the improved stove (postintervention) for 47 women, enabling paired comparisons.Results: Median pre- and postintervention values were 4 and 1 ppm for continuous personal CO and 3 and 1 ppm for passive sampler CO, respectively. Postintervention measurements indicated an average reduction of 42% for hydroxylated metabolites of naphthalene, fluorene, phenanthrene, and pyrene on a whole-weight concentration basis (micrograms per liter of urine), and a 34% reduction on a creatinine-adjusted basis (micrograms per gram of creatinine). Pre- and postintervention geometric mean values for 1-hydroxypyrene were 3.2 and 2.0 μg/g creatinine, respectively.Conclusion: Use of the Patsari stove significantly reduced CO and PAH exposures in women. However, levels of many PAH biomarkers remained higher than those reported among smokers.

Tissue Distribution and Depuration Kinetics of Waterborne 14C-labeled Light PAHs in Mummichog (Fundulus heteroclitus)

Light polycyclic aromatic hydrocarbons (PAHs) of petrogenic origin are commonly found in estuaries and coastal areas. Though they are known to be toxic to fish, little is known about their uptake and tissue distribution. This paper reports on the results of a study on uptake, elimination, and tissue distribution of three waterborne 14C-labeled PAHs in the mummichog, Fundulus heteroclitus, using whole-body autoradiography. After a 24 h exposure to 1 μCi·L(-1) of 14C-naphthalene, 14C-1-naphthol, and 14C-phenanthrene, fish were transferred to clean water and tissue distribution examined after 0, 1, 3, 7, 14, and 21 days of depuration. All compounds were readily accumulated by fish and were also rapidly eliminated (t0.5 range=1.1 to 3.0 days). Most of the radioactivity in naphthalene- and phenanthrene-treated fish was found in gall bladder≫liver>intestinal lumen. In naphthol-exposed fish, an important labeling of some brain areas was observed. Brain of naphthalene-exposed fish was also labeled after 24 h depuration, indicating that exposure to naphthalene may result in metabolite accumulation in the brain. This is the first study showing that naphthalene, naphthol, and/or unidentified metabolite(s) can accumulate in brain tissues, which may impair normal brain function.

DNA adducts and strand breaks in workers exposed to vapours and aerosols of bitumen: associations between exposure and effect

To study the associations between exposure to vapours and aerosols of bitumen and genotoxic effects, a cross-sectional and cross-shift study was conducted in 320 exposed workers and 118 non-exposed construction workers. Ambient air measurements were carried out to assess external exposure to vapours and aerosols of bitumen. Hydroxylated metabolites of naphthalene, phenanthrene and pyrene were measured in urine, whereas (+)-anti-benzo[a]pyrene-7,8-diol-9,10-epoxide ((+)-anti-BPDE), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8oxodGuo) and DNA strand breaks were determined in blood. Significantly higher levels of 8-oxodGuo adducts and DNA strand breaks were found in both pre- and post-shift blood samples of exposed workers compared to those of the referents. No differences between exposed workers and referents were observed for (+)-anti-BPDE. Moreover, no positive associations between DNA damage and magnitude of airborne exposure to vapours and aerosols of bitumen could be observed in our study. Additionally, no relevant association between the urinary metabolites of PAH and the DNA damage in blood was observed. Overall, our results indicate increased oxidative DNA damage in workers exposed to vapours and aerosols of bitumen compared to non-exposed referents at the group level. However, increased DNA strand breaks in bitumen workers were still within the range of those found in non-exposed and healthy persons as reported earlier. Due to the lack of an association between oxidative DNA damage and exposure levels at the workplaces under study, the observed increase in genotoxic effects in bitumen workers cannot be attributed to vapours and aerosols of bitumen.

1,2-Dihydroxynaphthalene as biomarker for a naphthalene exposure in humans

The possibly carcinogenic properties of naphthalene are, regarding to its ubiquitary presence, of environmental-medical and occupational-medical importance. Seven isomeric dihydroxynaphthalenes (DHN) were examined for their suitability as biomarkers in human biomonitoring and to get insights in human naphthalene metabolism. We developed a GC-MS-method for the quantification of 1,2-, 1,4-, 1,5-, 1,6-, 1,7-, 2,6- and 2,7-DHN after solid phase extraction and derivatization with BSA/TMCS. The internal burden of DHN after exposure to naphthalene was determined by measuring urine collected from smokers and non-smokers among the general population and among occupationally exposed persons. The elaborated method can be regarded as specific and sensitive procedure to quantify the seven different DHN. In human urine, we detected 1,2-DHN as main metabolite in 54 of the 55 analysed samples. Median 1,2-DHN values (range) were 1012 μg/L (22-6477 μg/L) for workers and 8 μg/L (<LOD-62 μg/L) for controls. 1,4-, 1,7-, 2,6- and 2,7-DHN were quantified in 61-89% of the samples (range <LOD-113 μg/L). 1,5- and 1,6-DHN were not detected in human urine. In the urine samples of workers, median-concentrations of 1,2-DHN were about tenfold higher than those of the established biomarkers 1- and 2-naphthol, while in our control collective comparable concentration of 1,2-DHN and 1- and 2-naphthol were found. 1,2-DHN showed to be the most sensitive biomarker for an internal exposure to naphthalene. It is clearly superior to 1- and 2-naphthol which up till now have been used for biomonitoring purposes. Moreover 1,2-DHN is the precursor of the possibly ultimate carcinogenic metabolite of naphthalene, 1,2-naphthoquinone. This way the diagnostical reliability of 1,2-DHN predominates the other naphthalene metabolites. So out of analytical and diagnostical reasons 1,2-DHN seems to be the most promising biomarker for the estimation of environmentally and occupationally caused internal exposures to naphthalene.

Abstract B32: Levels of metabolites of naphthalene in urine predict translocations in peripheral blood lymphocytes in children

Abstract Chromosomal aberrations (CAs) and translocations (TRNS) (subtype of CAs) are a validated biomarker of increased cancer risk in adults. Prenatal exposure to semi-volatile air PAH predicts levels of stable CAs in neonatal (cord) blood. Urinary levels of 1 and 2 naphthol, metabolites of naphthalene, an IARC class2b possible carcinogen, appear elevated in the CCCEH birth cohort. We hypothesize that concurrent exposure to naphthalene is associated with development of CAs and TRNS in children. Whole blood from 113 five year old children (68 girls) participating in the CCCEH birth cohort composed of Dominican (DomAm) (N=66) and African-Americans (AfrAm) (N=47) was processed fresh for Whole Chromosome Paint (WCP)-Fluorescent In Situ Hybridization using WCP for chromosomes 1-6. 400 Cell Equivalents [CE]) were scored for each child. TRNS were scored in accordance with the PAINT convention and frequencies were expressed per 100 CE. Levels of PAH metabolites were measured using MSGC in spot urine samples (Sjodin laboratory, CDC) collected concurrently and adjusted for urine specific gravity (SG). We examined predictors of presence of CA's or TRNS. For predicting frequency of CA and TRNS, mean ratios were derived for doubling of naphthol levels using negative binomial models. Results: 35 children had CA's and 20 had TRNS. Geometric means among AfrAm were 3490.2 for 1naphthol, and 3005.1 for 2naphthol, and 3336.4 and 5256.6 for 1 &amp; 2naphthol in DomAm children. Levels of 1&amp; 2 napthol did not vary with presence of a smoker in the child's home. 2naphthol (but not 1napthol) was higher in girls (p= 0.08) and in DomAm (p=0.01). After adjusting for gender and ethnicity, presence of both CA's and TRNS was predicted by increasing levels of 1- and 2- naphthol: OR for presence of CA's with increasing 1naphthol: (1.23; 95%CI:1.01, 1.48); and with 2napthol (1.19; 0.90, 1.56), but the effect was significantly higher in DomAms for both 1naphthol (1.63;1.21, 2.19) &amp; 2naphthol (1.44; 1.02, 2.04). For TRNS, the OR was 1.55 (1.11,2.17) for 1naphthol, and 1.92 (1.20-3.08) for 2naphthol. Frequency of CA's was predicted by 1naph (MR1.38; 1.10, 1.72) in DomAm but not in AfrAm (RR0.90; 0.65, 1.25). The association between naphthols and frequency of TRNS did not differ by ethnicity. To examine possible dose-response, levels of 2naphthol were grouped into tertiles using the lowest tertile as the referent group. After adjusting for gender/ethnicity, the proportion with TRNS present increased with increasing 2naphthol (though trend p = 0.09). Children with 2napthol in the highest tertile were significantly more likely to have translocations (OR 4.29; 95% CI: 1.11-16.55) when compared with children in the lowest tertile. Similarly, children with increasing 2-naphthol levels had increasing frequency of TRNS though the overall trend was not significant: mean TRNS frequencies for children in the highest tertile of exposure had a ratio of 3.23; (95%CI: 0.85-12.50) compared with those in the lowest tertile. The effect of increasing naphthol on aberrations or translocations was not dominated by any one chromosome, in contrast to our findings in newborns and prenatal air-PAH. Together these findings support our earlier findings that PAH exposure is associated with formation of TRNS, and suggests a dose response pattern with exposure to naphthalene, and an ethnic difference in susceptibility to formation of CAs. Citation Information: Cancer Prev Res 2010;3(12 Suppl):B32.

A liquid chromatography/tandem mass spectrometry (LC-MS/MS) method for the determination of phenolic polycyclic aromatic hydrocarbons (OH-PAH) in urine of non-smokers and smokers

Polycyclic aromatic hydrocarbons (PAH) are products of the incomplete combustion of organic materials and, therefore, occur ubiquitously in the environment and also in tobacco smoke. Since some PAH have been classified as carcinogens, it is important to have access to suitable analytical methods for biomarkers of exposure to this class of compounds. Past experience has shown that measuring a profile of PAH metabolites is more informative than metabolites of a single PAH. Assessment of environmental and smoking-related exposure levels requires analytical methods with high sensitivity and specificity. In addition, these methods should be fast enough to allow high throughput. With these pre-conditions in mind, we developed and validated a high-performance liquid chromatographic method with tandem mass spectrometric detection (LC-MS/MS) for the determination of phenolic metabolites of naphthalene, fluorene, phenanthrene and pyrene in urine of smokers and non-smokers. Sample work-up comprised enzymatic hydrolysis of urinary conjugates and solid-phase extraction on C18 cartridges. The method showed good specificity, sensitivity, and accuracy for the intended purpose and was also sufficiently rapid with a sample throughput of about 350 per week. Application to urine samples of 100 smokers and 50 non-smokers showed significant differences between both groups for all measured PAH metabolites, and strong correlations with markers of daily smoke exposure in smoker urine. Urinary levels were in good agreement with previously reported data using different methodologies. In conclusion, the developed LC-MS/MS method is suitable for the quantification of phenolic PAH metabolites of naphthalene, fluorene, phenanthrene, and pyrene in smoker and non-smoker urine.

Does Switching to Reduced Ignition Propensity Cigarettes Alter Smoking Behavior or Exposure to Tobacco Smoke Constituents?

Since 2004, several jurisdictions have mandated that cigarettes show reduced ignition propensity (RIP) in laboratory testing. RIP cigarettes may limit fires caused by smoldering cigarettes, reducing fire-related deaths and injury. However, some evidence suggests that RIP cigarettes emit more carbon monoxide and polycyclic aromatic hydrocarbons, and smokers may alter their smoking patterns in response to RIP cigarettes. Both of these could increase smokers' exposures to harmful constituents in cigarettes. An 18-day switching study with a comparison group was conducted in Boston, MA (N = 77), and Buffalo, NY (N = 83), in 2006-2007. Current daily smokers completed 4 laboratory visits and two 48-hr field data collections. After a 4-day baseline, Boston participants switched to RIP cigarettes for 14 days, whereas Buffalo participants smoked RIP cigarettes throughout. Outcome measures included cigarettes smoked per day; smoking topography; salivary cotinine; breath CO; and hydroxylated metabolites of pyrene, naphthalene, phenanthrene, and fluorene. Because the groups differed demographically, analyses adjusted for race, age, and sex. We observed no significant changes in smoking topography or CO exposure among participants who switched to RIP cigarettes. Cigarette use decreased significantly in the switched group (37.7 cigarettes/48 hr vs. 32.6 cigarettes/48 hr, p = .031), while hydroxyphenanthrenes increased significantly (555 ng/g creatinine vs. 669 ng/g creatinine, p = .007). No other biomarkers were significantly affected. Small increases in exposure to phenanthrene among smokers who switched to RIP versions were observed, while other exposures and smoking topography were not significantly affected. Toxicological implications of these findings are unclear. These findings should be weighed against the potential public health benefits of adopting RIP design standards for cigarette products.

Environmental and biological monitoring of exposures to PAHs and ETS in the general population

The objective of this study was to analyse environmental tobacco smoke (ETS) and PAH metabolites in urine samples of non-occupationally exposed non-smoker adult subjects and to establish relationships between airborne exposures and urinary concentrations in order to (a) assess the suitability of the studied metabolites as biomarkers of PAH and ETS, (b) study the use of 3-ethenypyridine as ETS tracer and (c) link ETS scenarios with exposures to carcinogenic PAH and VOC. Urine samples from 100 subjects were collected and concentrations of monophenolic metabolites of naphthalene, fluorene, phenanthrene, and pyrene and the nicotine metabolites cotinine and trans-3′-hydroxycotinine were measured using liquid chromatography–tandem mass spectrometry (LC-MS/MS) to assess PAH and ETS exposures. Airborne exposures were measured using personal exposure samplers and analysed using GC–MS. These included 1,3-butadiene (BUT), 3-ethenylpyridine (3-EP) (a tobacco-specific tracer derived from nicotine pyrolysis) and PAHs. ETS was reported by the subjects in 30-min time–activity questionnaires and specific comments were collected in an ETS questionnaire each time ETS exposure occurred. The values of 3-EP (> 0.25 μg/m 3 for ETS) were used to confirm the ETS exposure status of the subject. Concentrations as geometric mean, GM, and standard deviation (GSD) of personal exposures were 0.16 (5.50) μg/m 3 for 3-EP, 0.22 (4.28) μg/m 3 for BUT and 0.09 (3.03) ng/m 3 for benzo(a)pyrene. Concentrations of urinary metabolites were 0.44 (1.70) ng/mL for 1-hydroxypyrene and 0.88 (5.28) ng/mL for cotinine. Concentrations of urinary metabolites of nicotine were lower than in most previous studies, suggesting very low exposures in the ETS-exposed group. Nonetheless, concentrations were higher in the ETS population for cotinine, trans-3′hydroxycotinine, 3-EP, BUT and most high molecular weight PAH, whilst 2-hydroxyphenanthrene, 3 + 4-hydroxyphenanthrene and 1-hydroxyphenanthrene were only higher in the high-ETS subpopulation. There were not many significant correlations between either personal exposures to PAH and their urinary metabolites, or of the latter with ETS markers. However, it was found that the urinary log cotinine concentration showed significant correlation with log concentrations of 3-EP ( R = 0.75), BUT ( R = 0.47), and high molecular weight PAHs (MW > 200), especially chrysene ( R = 0.55) at the p = 0.01 level. On the other hand, low correlation was observed between the PAH metabolite 2-naphthol and the parent PAH, gas-phase naphthalene. These results suggest that (1) ETS is a significant source of inhalation exposure to the carcinogen 1,3-butadiene and high molecular weight PAHs, many of which are carcinogenic, and (2) that for lower molecular weight PAHs such as naphthalene, exposure by routes other than inhalation predominate, since metabolite levels correlated poorly with personal exposure air sampling.