The effects of UV exclusion on the soluble phenolics of young Scots pine seedlings in the subarctic

The characteristics of UV-absorbing compounds, particularly soluble phenolics, were studied in needles of 63-day-old seed-grown Scots pine (Pinus sylvestris L.) seedlings of two provenances in a UV exclusion field experiment at Pallas-Ounastunturi National Park in Finnish Lapland (68 degrees N, 270 m a.s.l.). The experiment used the following plastic filters in exclosure treatments to manipulate the spectral balance of natural irradiance: (1) 'control' (a polyethene plastic filter); (2) 'UV-B exclusion' (a clear polyester filter); and (3) 'UV-B/UV-A exclusion' (a clear acryl plate). Polyethene transmitted 89% of the ambient levels of total UV (280-400 nm), polyester transmitted 75% of the total UV, but only 0.6% of the UV-B (280-315 nm) component, while acryl plate transmitted 0.2% of UV (280-360 nm). The research also included (4) 'Ambient' plants that were not subjected to any treatment exclosures. After the 58 day UV exclusion, significant (p<0.0001) differences due to treatments were determined for a kaempferol derivative, kaempferol 3-glucoside, and a quercetin derivative, the quantities of which ranged from 0.23 to 0.45, 0.42 to 1.34 and 0.39 to 0.75 micromol g FW(-1), respectively, depending on treatment and provenance. Overall, Scots pine seedlings grown at ambient UV radiation (PAS300, Caldwell's generalized Plant Action Spectrum (PAS) normalized at 300 nm, 72 mW m(-2)) or under a control had significantly (p<0.05) higher quantities of soluble phenolics than seedlings grown under UV-B or UV-B/UV-A exclusion treatments. There were no significant differences in the quantity of soluble phenolics between the two exclosure treatments or between the two Scots pine provenances. The sums of diacylated flavonol glucosides ranging from 3.75 to 4.55 micromol g FW(-1) depending on treatment and provenance, were already present at very low UV-levels under the UV-B/UV-A exclusion treatment. The present study indicated that soluble phenolics, particularly the diacylated flavonol glucosides, may provide an effective preformed protection for young Scots pine seedlings against UV-B and UV-A radiation.     

Genotoxic fungicide methyl thiophanate as an oxidative stressor inducing 8-oxo-7,8-dihydro-2′ -deoxyguanosine adducts in DNA and mutagenesis

Dimethyl 4,4′ -(O-phenylene)bis(3-thioallophanate), commonly known as methyl thiophanate (MT), is a systemic fungicide and suspected carcinogen to humans. In this study, the oxidative potential of this category-III acute toxicant has been ascertained based on its capacity of inducing reactive oxygen species (ROS) and promutagenic 8-oxo-7,8-dihydro-2′ -deoxyguanosine (8-oxodG) adducts in DNA. The discernible MT dose-dependent reduction in fluorescence intensity of a cationic dye rhodamine (Rh-123) in human lymphocytes and increased fluorescence intensity of 2′,7′-Dichlorodihydro fluorescein diacetate (DCFH-DA) treated cells signifies decreased mitochondrial membrane potential (Δ Ψ m) due to intracellular ROS generation. The ³²P-post-labeling assay demonstrated the MT-induced 8-oxodG adduct formation in calf thymus DNA. Thus, it is concluded that MT, as a potent oxidative stressor, produces ROS leading to mitochondrial dysfunction, oxidative DNA damage and mutagenesis.     

Modeling the photo-oxidation of dissolved organic matter by ultraviolet radiation in freshwater lakes: implications for mercury bioavailability

Uncertainties in projected ultraviolet (UV) radiation may lead to future increases in UV irradiation of freshwater lakes. Because dissolved organic carbon (DOC) is the main binding phase for mercury (Hg) in freshwater lakes, an increase in DOC photo-oxidation may affect Hg speciation and bioavailability. We quantified the effect of DOC concentration on the rate of abiotic DOC photo-oxidation for five lakes (DOC=3.27-12.3 mg L(-1)) in Kejimkujik National Park, Canada. Samples were irradiated with UV-A or UV-B radiation over a 72-h period. UV-B radiation was found to be 2.36 times more efficient at photo-oxidizing DOC than UV-A, with energy-normalized rates of dissolved inorganic carbon (DIC) production ranging from 3.8×10(-5) to 1.1×10(-4) mg L(-1)J(-1) for UV-A, and from 6.0×10(-5) to 3.1×10(-4) mg L(-1)J(-1) for UV-B. Energy normalized rates of DIC production were positively correlated with DOC concentrations. Diffuse integrated attenuation coefficients were quantified in situ (UV-A K(d)=0.056-0.180 J cm(-1); UV-B K(d)=0.015-0.165 J cm(-1)) and a quantitative depth-integrated model for yearly DIC photo-production in each lake was developed. The model predicts that, UV-A produces between 3.2 and 100 times more DIC (1521-2851 mg m(-2) year(-1)) than UV-B radiation (29.17-746.7 mg m(-2) year(-1)). Future increases in UV radiation may increase DIC production and increase Hg bioavailability in low DOC lakes to a greater extent than in high DOC lakes.     

Influence of solar UV radiation on the nitrogen metabolism in needles of Scots pine (Pinus sylvestris L.)

Needles of 20-year-old Scots pine (Pinus sylvestris L.) saplings were studied in an ultraviolet (UV) exclusion field experiment (from 2000 to 2002) in northern Finland (67 °N). The chambers held filters that excluded both UV-B and UV-A, excluded UV-B only, transmitted all UV (control), or lacked filters (ambient). UV-B/UV-A exclusion decreased nitrate reductase (NR) activity of 1-year-old needles of Scots pines compared to the controls. The proportion of free amino acids varied in the range 1.08-1.94% of total proteins, and was significantly higher in needles of saplings grown under UV-B/UV-A exclusion compared to the controls or UV-B exclusion. NR activity correlated with air temperature, indicating a “chamber effect”. The study showed that both UV irradiance and increasing temperature are significant modulators of nitrogen (N) metabolism in Scots pine needles.     

Damage to DNA caused by UV-B radiation in the desert cyanobacterium Scytonema javanicum and the effects of exogenous chemicals on the process

Radiation with UV-B increased the damage to DNA in Scytonema javanicum, a desert-dwelling soil microorganism, and the level of damage varied with the intensity of UV-B radiation and duration of exposure. Production of reactive oxygen species (ROS) also increased because of the radiation. Different exogenous chemicals (ascorbate acid, ASC; N-acetylcysteine, NAC; glyphosate, GPS; and 2-methyl-4-chlorophenoxyacetic acid, MCPA-Na) differed in their effect on the extent of DNA damage and ROS production: whereas NAC and ASC protected the DNA from damage and resulted in reduced ROS production, the herbicides (GPS and MCPA-Na) increased the extent of damage, lowered the rate of photosynthesis, and differed in their effect on ROS production. The chemicals probably have different mechanisms to exercise their effects: NAC and ASC probably function as antioxidant agents or as precursors of other antioxidant molecules that protect the DNA and photosynthetic apparatus directly from the ROS produced as a result of UV-B radiation, and GPS and MCPA-Na probably disrupt the normal metabolism in S. javanicum to induce the leaking of ROS into the photosynthetic electron transfer pathway following UV-B radiation, and thereby damage the DNA. Such mechanisms have serious implications for the use of environment-friendly herbicides, which, because they can destroy DNA, may prove harmful to soil microorganisms.     

Gaseous mercury emissions from unsterilized and sterilized soils: The effect of temperature and UV radiation

Mercury (Hg) emissions from the soils taken from two different sites (deciduous and coniferous forests) in the Adirondacks were measured in outdoor and laboratory experiments. Some of the soil samples were irradiated to eliminate biological activity. The result from the outdoor measurements with different soils suggests the Hg emission from the soils is partly limited by fallen leaves covering the soils which helps maintain relatively high soil moisture and limits the amount of heat and solar radiation reaching the soil surface. In laboratory experiments exposure to UV-A (365 nm) had no significant effect on the Hg emissions while the Hg emissions increased dramatically during exposure to UV-B (302 nm) light suggesting UV-B directly reduced soil-associated Hg. Overall these results indicate that for these soils biotic processes have a relatively constant and smaller influence on the Hg emission from the soil than the more variable abiotic processes.     

Long-term exposure to enhanced UV-B radiation has no significant effects on growth or secondary compounds of outdoor-grown Scots pine and Norway spruce seedlings

The effects of long-term enhanced UV-B radiation on growth and secondary compounds of two conifer species were studied in an outdoor experiment. Scots pine (Pinus sylvestris) seedlings were exposed for two growing seasons and Norway spruce (Picea abies) seedlings for three growing seasons to supplemental UV-B radiation, corresponding to a 30% increase in ambient UV-B radiation. The experiment also included appropriate controls for ambient and increased UV-A radiation. Enhanced UV-B did not affect the growth of the conifer seedlings. In addition, neither the concentrations of terpenes and phenolics in the needles nor the concentrations of terpenes in the wood were affected. However, in the UV-A control treatment the concentrations of diterpenes in the wood of Scots pine decreased significantly compared to the ambient control. Apparently, a small increase in UV-B radiation has no significant effects on the secondary compounds and growth of Scots pine and Norway spruce seedlings.     

Spectral attenuation of solar ultraviolet radiation in humic lakes in Central Finland

The attenuation of solar ultraviolet (UV) radiation in five lakes in Central Finland was evaluated through field measurements and/or by determining the optical properties of the lake water during summer 1999. Spectral UV irradiance in the air and at several depths underwater was measured in three lakes (Lake Palosj?rvi, Konnevesi, and Jyv?sj?rvi) with dissolved organic carbon (DOC) ranging from 4.9 to 8.7 mg l(-1) and chlorophyll a ranging from 1.6 to 16 g l(-1). According to the field measurements, 99% of the UV-B radiation was attenuated in approximately a half meter water column in the clearest lake. In the UV-A region at 380 nm, the corresponding attenuation occurred in the upper one meter. In a small humic lake (DOC 13.2-14.9 mg l(-1)) UV-B radiation was attenuated to 1% of the subsurface irradiance within the top 10 cm water column, whereas UV-A radiation (at 380 nm) penetrated more than twice as deeply (maximum 25 cm), as predicted from the absorption coefficients. These results suggest the importance of the dissolved fraction of lake water in governing the UV attenuation in lakes. This was seen from the significant relationship between the vertical attenuation coefficients (Kd) based on field measurements and the absorption coefficients (ad) derived from spectrophotometric laboratory scannings, as well as between Kd and DOC.     

Preliminary evidence of the role of hydrogen peroxide in the degradation of benzo[a]pyrene by a non-white rot fungus Fusarium solani

In order to study the enzymatic mechanisms involved in the successive steps of BaP degradation by a Deuteromycete fungus Fusarium solani, we developed an indirect approach by using inhibitors of enzymes. We used either specific inhibitors of peroxidases (i.e. salicylhydroxamic acid) and of cytochrome P-450 (i.e. piperonyl butoxyde) or inhibitors of both enzymes (i.e. potassium cyanide). Surprisingly, no expected decrease of BaP degradation was observed with most inhibitors tested. On the contrary, more BaP was degraded. Only butylated hydroxytoluene, which acts as a free radical scavenger, inhibited BaP degradation. The inhibition of these enzymes, which use H(2)O(2) as a cosubstrate, might have resulted in an increase of hydrogen peroxide availability in the fungal cultures. This enhancement could induce formation of reactive oxygen species (ROS) which might be the agents that initiate benzo[a]pyrene oxidation. This study proposed a hypothetic alternative metabolic pathway involved in PAH metabolism by Fusarium solani.     

The degradation of the cyanobacterial hepatotoxin nodularin (NOD) by UV radiation

This study investigates the decomposition of NOD by UV irradiation. Water solutions of pure NOD and NOD-containing Nodularia extract as well as Nodularia filaments collected on filters were exposed to UV-A, UV-B, and white fluorescent light (VIS) during 48 h experiments. In VIS, the toxin was fairly stable and only 3.8-4.6% of the original degraded. UV-B had the most pronounced effect on the NOD degradation rate. In the experiment, the overall loss of NOD was 0.27 and 0.77 micro g ml(-1)day(-1) for the solution of pure toxin and Nodularia extract and 0.28 micro g day(-1) for Nodularia filaments. Comparison of UV-B degradation rate in water and methanol extracts revealed higher stability of NOD in methanol. This might suggest that some hydrophobic components of Nodularia cell play a protective role against UV radiation. Additionally, chemical (LC-MS/MS) and biochemical (ELISA and PPIA) assays were employed to characterize the UV degradation products. LC-MS/MS analyses showed that in UV-B exposed sample, apart from NOD, there were three other compounds with molecular ion at m/z at 825.4. The fragmentation pattern of the ion was the same for all four compounds suggesting that they are geometrical isomers of NOD. The major degradation product, with a local absorption maximum at 242 nm, was active in both biochemical assays.     

Feasibility of ultraviolet-light-emitting diodes as an alternative light source for photocatalysis

The objective of this study was to determine whether ultraviolet-light-emitting diodes (UV-LEDs) could serve as an efficient photon source for heterogeneous photocatalytic oxidation (PCO). An LED module consisting of 12 high-power UV-A (lambda max = 365 nm) LEDs was designed to be interchangeable with a UV-A fluorescent black light blue (BLB) lamp for a bench scale annular reactor packed with silica-titania composite (STC) pellets. Lighting and thermal properties of the module were characterized to assess its uniformity and total irradiance. A forward current (I(F)) of 100 mA delivered an average irradiance of 4.0 mW cm(-2) at a distance of 8 mm, which is equivalent to the maximum output of the BLB, but the irradiance of the LED module was less uniform than that of the BLB. The LED and BLB reactors were tested for the oxidization of ethanol (50 ppm(v)) in a continuous-flow-through mode with 0.94 sec residence time. At the same average irradiance, the UV-A LED reactor resulted in a lower CO2 production rate (19.8 vs. 28.6 nmol L(-1) s(-1)), lower ethanol removal (80% vs. 91%), and lower mineralization efficiency (28% vs. 44%) than the UV-A BLB reactor. Ethanol mineralization was enhanced with the increase of the irradiance at the catalyst surface. This result suggests that reduced ethanol mineralization in the LED reactor relative to the BLB reactor at the same average irradiance could be attributed to the nonuniform irradiance over the photocatalyst, that is, a portion of the catalyst was exposed to less than the average irradiance. The potential of UV-A LEDs may be fully realized by optimizing the light distribution over the catalyst and utilizing their instantaneous "on" and "off" feature for periodic irradiation. Nevertheless, our results also showed that the current UV-A LED module had the same wall plug efficiency (WPE) of 13% as that of the UV-A BLB, demonstrating that UV-A LEDs are a viable photon source both in terms of WPE and PCO efficiency.     

Seasonal acclimation of the moss Polytrichum juniperinum Hedw. to natural and enhanced ultraviolet radiation

Short- and long-term changes in the methanol-extractable UV-absorbing compounds and biomass of the pioneer moss Polytrichum juniperinum in response to natural and enhanced UV radiation were studied. Under natural conditions, the compounds were found to fluctuate seasonally. In summer these compounds correlated negatively with irradiation. The concentration was low in July after a period of simultaneous heat, drought and high irradiation. Transient positive correlation between daily concentration and UV was seen in June. The concentration increased towards autumn and was relatively high under snow. Two enhanced UV experiments were performed. Seasonality in the compounds was again observed, with negative correlations with irradiation. During the first weeks, a transient inhibition of compound production was observed after the daily UV-B treatment. After six years of modulated UV-treatment in situ, photosynthesising biomass decreased under UV-B and increased under UV-A. A larger variation in the UV-absorbing compounds was observed under UV-B treatment.     

Assessment of oxidative stress due to exposure to pesticides in plasma and urine of traditional limited-resource farm workers: Formation of the DNA-adduct 8-hydroxy-2-deoxy-guanosine (8-OHdG)

The objective of this study was to assess the risk of genotoxicity caused due to oxidative stress using plasma and urinary levels of 8-hydroxy-2-deoxyguanosine (8-OHdG) in farm workers for six months during a growing season. Blood and urine samples were collected once a month for six months (June to November 2003) from farm workers (n = 15) and urban unexposed controls (n = 10). Plasma and urinary levels of 8-OHdG were evaluated by Enzyme Linked Immunosorbent Assay (ELISA) technique. There was no significant difference in the urinary levels of 8-OHdG between the farm workers and the control group, but there was an approximately four-fold increase in mean values of plasma 8-OHdG levels in the farm worker group (P ≤ 0.05).     

Photoinduced toxicity of retene to Daphnia magna under enhanced UV-B radiation

The effects of UV radiation on the acute toxicity of retene (7-isopropyl-1-methylphenanthrene) to Daphnia magna Straus were studied. Dehydroabietic acid (DHAA) from which retene is formed in the vicinity of pulp and paper industry was also studied. Pyrene, anthracene, and phenanthrene were used as model PAH compounds. The time taken for immobilization (ET50) was monitored under biologically effective UV-B dose rates of 240, 365, 565, and 650 mW m(-2) (UV-A and visible light also present). Median effective concentrations (EC50) were determined after a 15-min UV exposure (565 mW m(-2)) followed by 24 h in the dark. Retene (10-320 microg l(-1)) was not acutely toxic in the dark. The induction of phototoxicity was in agreement with the absorption properties of the compounds (absorption peak of retene at around 300 nm). Photoinduced toxicity followed an order pyrene > anthracene > retene. Phenanthrene and DHAA were not acutely phototoxic. Accumulation of the compound in Daphnia before UV exposure was essential. Some changes in the absorption spectra of the compounds were seen after a 5-h UV irradiation (565 mW m(-2)), but none of the irradiated compounds were acutely toxic without further UV exposure. Therefore, the enhanced acute toxicity was primarily due to internal photosensitization reactions rather than photomodification. The dissolved fraction of 25% pulp and paper mill effluent reduced phototoxicity by attenuating UV radiation. The phototoxicity of retene was a function of both the exposure concentration and the UV-B dose rate, but relatively high UV-B dose rates and concentrations were needed for the acute photoinduced toxicity.     

Assessment of oxidative stress due to exposure to pesticides in plasma and urine of traditional limited-resource farm workers: formation of the DNA-adduct 8-hydroxy-2-deoxy-guanosine (8-OHdG)

The objective of this study was to assess the risk of genotoxicity caused due to oxidative stress using plasma and urinary levels of 8-hydroxy-2-deoxyguanosine (8-OHdG) in farm workers for six months during a growing season. Blood and urine samples were collected once a month for six months (June to November 2003) from farm workers (n = 15) and urban unexposed controls (n = 10). Plasma and urinary levels of 8-OHdG were evaluated by Enzyme Linked Immunosorbent Assay (ELISA) technique. There was no significant difference in the urinary levels of 8-OHdG between the farm workers and the control group, but there was an approximately four-fold increase in mean values of plasma 8-OHdG levels in the farm worker group (P < or = 0.05).     

Oxidative DNA damage estimated by urinary 8-hydroxydeoxyguanosine: influence of taxi driving,smoking and areca chewing

Nitrogen oxides (NOx) and polycyclic aromatic hydrocarbons are common air pollutants generated from automobile exhaust and cigarette smoke. This study was to investigate urinary 8-hydroxydeoxyguanosine (8-OHdG) as an effective biomarker on DNA damage from traffic exhaust and/or smoking in exposed and non-exposed individuals. With subject consents, the levels of plasma NOx, urinary 1-hydroxypyrene (1-OHP) and 8-OHdG were determined for 95 male taxi drivers and 75 male community residents as the reference group. After adjusting for associate variables, there was a significant correlation between the levels of urinary 8-OHdG and 1-OHP but not NOx. The average level of urinary 8-OHdG was significantly higher in drivers than in community men (13.4+/-4.7 vs. 11.5+/-4.7 microg/g creatinine in mean+/-standard deviation). Compared with non-smoking community men, the multivariate logistic regression showed that the odds ratios (OR) of having elevated levels of urinary 8-OHdG (greater than the overall median value, 12.1 microg/g creatinine) were 6.6 (95% confidence interval (CI)=2.1-20.8) for smoking community men, 5.0 (95% CI=1.7-14.7) for non-smoking taxi drivers and 4.6 (95% CI=1.4-15.0) for smoking taxi drivers. Higher risk was also observed for areca quid chewers compared with non-chewers (OR=1.6; 95% CI=1.1-3.6). In conclusion, taxi driving and smoking may contribute independently to elevated DNA damage using urinary 8-OHdG levels as a sensitive biomarker. This effect is most potent on heavy smokers.     

Photochemical alteration of the molecular weight of dissolved organic matter

Molecular weight (MW) is a key control on the physical, chemical, and biological characteristics of dissolved organic mater (DOM). This study investigated the effect of photooxidation on the average MW of DOM by exposing DOM of diverse origins to simulated solar radiation at varying levels of dissolved oxygen and under different incident light wavelength regimes. During irradiation, high-molecular-weight fractions were destroyed and low-molecular-weight constituents were formed. The average MW decreased with irradiation time in all treatments in a manner that can be described by a quasi-exponential function, which suggests that solar radiation is incapable of completely mineralizing DOM even after prolonged exposure. Increasing the oxygen concentration accelerated the MW reduction while the removal of oxygen strongly suppressed this process. The fractional contributions from UV-B, UV-A, and visible radiations to full-spectrum photoinduced MW reduction varied considerably among the DOM samples examined, ranging from 19% to 60%, 17% to 36%, and 15% to 46%, respectively. The MW changes in time-series irradiations were inversely correlated with the ratio of the absorbance at 250 nm to that at 365 nm (i.e., the E₂/E₃ quotient). Photoinduced MW reduction was accompanied by a decrease of polydispersity, which is indicative of a reduced DOM heterogeneity.     

Impacts of Antioxidants on Hydroxyl Radical Production from Individual and Mixed Transition Metals in a Surrogate Lung Fluid

Inhalation of ambient particulate matter causes morbidity and mortality in humans. One hypothesized mechanism of toxicity is the particle-induced formation of reactive oxygen species (ROS) - including the highly damaging hydroxyl radical ((·)OH) - followed by inflammation and a variety of diseases. While past studies have found correlations between ROS formation and a variety of metals, there are no quantitative measurements of (·)OH formation from transition metals at concentrations relevant to 24-hour ambient particulate exposure. This research reports specific and quantitative measurements of (·)OH formation from 10 individual transition metals (and several mixtures) in a cell-free surrogate lung fluid (SLF) with four antioxidants: ascorbate, citrate, glutathione, and uric acid. We find that Fe and Cu can produce (·)OH under all antioxidant conditions as long as ascorbate is present and that mixtures of the two metals synergistically increase (·)OH production. Manganese and vanadium can also produce (·)OH under some conditions, but given that their ambient levels are typically very low, these metals are not likely to chemically produce significant levels of (·)OH in the lung fluid. Cobalt, chromium, nickel, zinc, lead, and cadmium do not produce (·)OH under any of our experimental conditions. The antioxidant composition of our SLF significantly affects (·)OH production from Fe and Cu: ascorbate is required for (·)OH formation, citrate increases (·)OH production from Fe, and both citrate and glutathione suppress (·)OH production from Cu. MINTEQ ligand speciation modeling indicates that citrate and glutathione affect (·)OH production by changing metal speciation, altering the reactivity of the metals. In the most realistic SLF (i.e., with all four antioxidants), Fe generates approximately six times more (·)OH than does the equivalent amount of Cu. Since levels of soluble Fe in PM are typically higher than those of Cu, our results suggest that Fe dominates the chemical generation of (·)OH from deposited particles in the lungs.     

Effects of ultraviolet-B radiation (UV-B) on growth and physiology of the dune grassland species Calamagrostis epigeios

Seedlings of Calamagrostis epigeios were exposed to four levels of UV-B radiation (280-320 nm), simulating up to 44% reduction of stratospheric ozone concentration during summertime in The Netherlands, to determine the response of this plant species to UV-B irradiation. After six weeks of UV-B treatment, total biomass of all UV-B treated plants was higher, compared to plants that had received no UV-B radiation. The increase of biomass did not appear to be the result of a stimulation of net photosynthesis. Also, transpiration rate and water use efficiency were not altered by UV-B at any exposure level. Pigment analysis of leaf extracts showed no effect of enhanced UV-B radiation on chlorophyll content and accumulation of UV absorbing pigments. UV-B irradiance, however, did reduce the transmittance of visible light (400-700 nm) of intact attached leaves, suggesting a change in anatomical characteristics of the leaves. Additionally, the importance of including an ambient UV-B treatment in indoor experiments is discussed.     

FIA/MS analysis of temporally ozonated benzo[a]pyrene and pyrene and their reaction products: inhibition of gap junction-mediated intercellular communication

In this study, the polycyclic aromatic hydrocarbons, benzo[a]pyrene (BaP) and pyrene, were subjected to temporal ozonation. The products from ozonation of 5 mg l(-1) BaP and 5 mg l(-1) pyrene, for varying time intervals (0, 1, 2, 3, 4, 5, 6, 8, 10, 20, and 30 min) were tested for their effects on gap junction-mediated intercellular communication (GJIC) in Clone 9 rat liver cells. Additionally, the ozonation products were also analyzed by flow injection analysis/mass spectrometry (FIA/MS) and the results were compared with the toxicity observed in the GJIC assay. Treatment of the Clone 9 cells with 5 mg l(-1) of ozonated BaP products resulted in a decrease in GJIC that was inversely proportional to the length of ozonation. The products from 1 min of ozonation resulted in a 92% decrease in the rate of GJIC, but with >5 min ozonation, the products did not suppress GJIC. In contrast, pyrene (0.5 mg l(-1)) required >10 min of ozonation to alleviate its effects on GJIC. FIA/MS, using atmospheric pressure chemical ionization (APCI), demonstrated products with higher molecular weights (MW) than their corresponding parent compounds, BaP (MW 252) and pyrene (MW 202). Ozonation of pyrene formed significantly fewer products than BaP. More importantly, pyrene ozonation products were constant from 1 to 10 min, while BaP ozonation products seemed to vary between time intervals. With the longer ozonation times (20 and 30 min), BaP and pyrene formed similar products (m/z peaks 157, 111, and 96). The suppression of GJIC by ozonated products seemed to correlate with oxidation of the aromatic ring framework. Further oxidation (longer ozonation times) to lower MW products correlated with restoration of normal GJIC.