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As part of the NERC-URGENT thematic programme, research was undertaken into the physicochemistry and bioreactivity of microscopic airborne particulate matter in south Wales. This paper reviews the collecting and characterisation methods used in the research; some of the results obtained are shown as examples. Four main collecting locations were chosen: Cardiff (urban); Port Talbot (urban/industrial); Park Slip West coal opencast pit (industrial/rural); the Black Mountains (rural/background). Collections initially used a 30-l/min Negretti PM10 filter collection system, however in the later stages of the project increased use was made of a 1100-l/min impaction system (nicknamed the super-sucker). This latter device was developed at Harvard University USA, however was adapted and optimised at Cardiff University. Methods for the extraction of PM10 off polycarbonate filters and polyurethane substrates were developed, with particular attention being paid to minimise physical or chemical changes during the extraction, and the extracts being in an appropriate state for bioreactivity assessment. Physicochemical characterisation of the PM10 included the empirical measurement of shape and size using electron microscopy and semi-automated image analysis. The determinations of the water-soluble and -insoluble chemical components were undertaken by ion chromatography and inductive coupled plasma-mass spectrometry. The bioreactivity of south Wales airborne particles is not covered by this review.

The water-soluble metal content of 1950s London smogs and modern particulate matter (PM) are associated with adverse health effects. This study aimed to elucidate the bioreactivity of these metals alone and in mixtures and to investigate the comparative bioreactivities of a surrogate mixture and a PM sample. These revealed similar bioreactivities. A bioreactivity hierarchy of these metals was established: Fe2+ > Cu2+ > Fe3+ > VO2+ > Zn2+ > As3+ = Pb2+ = Mn2+ = VO3-. Secondary components (i.e., chlorides, sulfates, nitrates) did not affect metal bioreactivity, whereas oxidation state was important. Synergism was observed between zinc and various metal ions (Cu2+, Fe3+, VO2+). In conclusion, low-valence transition metals are key to PM bioreactivity.

This study was undertaken primarily to identify genetic markers of oedema and inflammation. Mild pulmonary injury was induced following the instillation of the oedema-producing agent, bleomycin (0.5 units). Oedema was then confirmed by conventional toxicology (lavage protein levels, free cell counts and lung/body weight ratios) and histology 3 days post-bleomycin instillation. The expression profile of 1176 mRNA species was determined for bleomycin-exposed lung (Clontech Atlas macroarray, n=9). To obtain pertinent results from these data, it was necessary to develop a simple, effective method for bioinformatic analysis of altered gene expression. Data were log10 transformed followed by global normalisation. Differential gene expression was accepted if: (a) genes were statistically significant (P < or = 0.05) from a two-tailed t test; (b) genes were consistently outside a two standard deviation (SD) range from control levels. A combination of these techniques identified 31 mRNA transcripts (approximately 3%) which were significantly altered in bleomycin treated tissue. Of these genes, 26 were down-regulated whilst only five were up-regulated. Two distinct clusters were identified, with 17 genes classified as encoding hormone receptors, and nine as encoding ion channels. Both these clusters were consistently down-regulated. The magnitude of the changes in gene expression were quantified and confirmed by Q-PCR (n = 6), validating the macroarray data and the bioinformatic analysis employed. In conclusion, this study has developed a suitable macroarray analysis procedure and provides the basis for a better understanding of the gene expression changes occurring during the early phase of drug-induced pulmonary oedema. This work has been presented orally, in part at the British Association for Lung Research Summer Meeting, University of Brighton, 3-5 September, 2003 and in full at the British Toxicology Society Annual Congress, Heriot Watt University, Edinburgh, 21-24 April 2004.

Total suspended particulate (TSP) samples collected on glass fibre filters in London before (1955) and after (1958-1974) the Clean Air Act was examined for physicochemical characteristics and oxidative capacity. High-resolution microscopy identified most of the material as soot with smelter spheres, fly ash (FA), sodium chloride and calcium sulphate particles. Image analysis (IA) was used to show that most of the soot aggregates were less than 1 microm in size and contained chains of individual particles of 10-50 nm. Speed mapping of large agglomerates of the historic particles confirmed that the samples were enriched with soot probably derived from a sulphur-rich coal called nutty slack which was used extensively at this time. Inductively coupled plasma-mass spectrometry (ICP-MS) was used to examine elemental composition. Meaningful quantitation of certain elements (Mg, Al and Zn) proved impossible because they were in high quantities in the glass fibre filters. However, high quantities of Fe>Pb>Cu>Mn>V>As were detected which may explain in part the bioreactivity of the samples. Using a simple in vitro test of oxidative capacity (plasmid assay), one historic particulate sample (1958) showed three times the activity of a modern-day diesel exhaust particle (DEP) sample but ten times less activity than a modern-day urban ambient particle collection. Such studies are continuing to link particle physicochemical properties and bioreactivity with a wider range of the samples collected between 1955 and 74 and how such historic samples compare with present-day London ambient particles.

A high-volume cascade impact collector (1100 l/min air flow) was used to collect air samples in an industrial (Port Talbot) and an urban (Cardiff) site with the purpose of characterising both coarse (PM(10-2.5)) and fine (PM(2.5)) fractions comprising the total sample. PM(10-2.5) and PM(2.5) samples were collected by cascading air through two polyurethane foams on which particles impact and become deposited. Air sample collection rates are to some extent dependent on weather conditions, notably rainfall, humidity, and especially, wind direction, but samples show a very different and distinctive air particle composition between the two collection sites. Thus, although both Cardiff and Port Talbot are coastal sites and therefore have high contents in chlorides, Port Talbot is extremely rich in tiny Fe spherules (>30%, in both coarse and fine fractions) from a nearby steel plant. Mineralogical characterisation using SEM-EDX shows a clear fractionation between the particle composition in the PM fractions, with the coarse fraction being dominated by chlorides, sulphates (gypsum), and silicates, and the fine fraction having high proportions of ammonium sulphates and elemental and organic carbon compounds, most of the latter being linked to traffic pollution.

Atmospheric aerosol samples were collected during different prevailing wind directions from a site located close to a busy motorway, a major steelworks, and the town of Port Talbot (Wales, UK). A high-volume collector was used (1100 l/min), enabling relatively large amounts of particulate matter (PM(10-2.5) and PM(2.5)) samples to be obtained on a polyurethane foam [PUF, H(2)N-C(O)O-CH(2)CH(3)] substrate over periods of 2-7 days. Four samples were chosen to exemplify different particle mixtures: SE- and NE-derived samples for particles moving along and across the motorway, a NW-derived sample from the town, and a mixed SW/SE-derived sample containing a mixture of particles from both steelworks and motorway. The latter sample showed the highest average collection rate (0.9 mg/h, 13 microg/m(3)) and included a prominent pollution episode when rainy winds were blowing from the direction of the steelworks. Both NW and SE samples were collected under dry conditions and show the same collection rate (0.7 mg/h, 10 microg/m(3)), whereas the NE sample was collected during wetter weather and shows the lowest rate (0.3 mg/h, 5 microg/m(3)). Scanning electron microscopy (SEM) and energy-dispersive X-ray microanalysis system (EDX) analyses show all samples are dominated by elemental and organic carbon compounds (EOCC) and nitrates, with lesser amounts of sulphates, felsic silicates, chlorides and metals. ICP-MS analyses show the SW/SE sample to be richest in metals, especially Fe, Zn, Ni, and Mn, these being attributed to an origin from the steelworks. The SE sample, blown along the motorway corridor, shows enhanced levels of Pb, V, Ti, As, and Ce, these metals being interpreted as defining a traffic-related chemical fingerprint. The NW sample shows a very low metal content. DNA plasmid assay data on the samples show TM(50) values varying from 66 to 175 microg/ml for the adjusted whole sample and 89 to 203 microg/ml for the soluble fraction. The SW/SE-mixed metalliferous sample is the most bioreactive (both whole and soluble) and the soluble fraction of the metal-depleted NW sample is the least bioreactive. The metal content of the aerosol samples, especially soluble metals such as Zn, is suggested to be the primary component responsible for oxidative damage of the DNA, and therefore most implicated in any health effects arising from the inhalation of these particulate cocktails.

Objectives: A substantial amount of Montserrat volcanic ash, containing up to 24% of cristobalite (w/w), a fibrogenic crystalline silica, has been generated since the first documented eruption in 1995. The bioreactivity of the ash and its two major components: cristobalite and anorthite have been studied in vivo for a year following intratracheal instillation into rats., Methods: The rats (n=5) were instilled with a sterile vehicle solution (0.15 M NaCl) and/or three doses (1.0, 2.5 or 5.0 mg) of each of the dust, and were sacrificed at 13, 25 or 49 weeks post-instillation for quantitative biochemical and histopathological analyses in the lung and lymph nodes., Results: Cristobalite caused inflammation in the lung and granuloma in the hilar lymph nodes associated with significant size augmentation at 13 weeks post-instillation (P<0.05) and cristobalite (5.0 mg) induced fibrosis in the lung at 49 weeks post-exposure. However, the Montserrat volcanic ash caused inflammation in the lung at 49 weeks post-treatment without any fibrogenic response although the ash (5.0 mg) triggered significant lymph node enlargement without significant changes in the lung at 13 weeks post-treatment (P<0.05). Dose and time independent responses in the anorthite-exposed lung and lymph nodes suggest that a single instillation of 5.0 mg of poorly soluble mineral dust does not induce any change in the lung or lymph nodes., Conclusion: The ash produces inflammatory reactions in lymph nodes at 13 weeks post-instillation in rats. These effects are seen much earlier than any inflammatory reaction in the lung. The onset of the lung inflammation is delayed until 49 weeks post-exposure. Despite the high cristobalite content of the ash, there is no evidence of any lung fibrogenic responses.

A positive correlation has been established between increased levels of airborne particulate pollution and adverse health effects, the toxicological mechanisms of which are poorly understood. For toxicologists to unambiguously determine these mechanisms, truly representative samples of ambient PM10 are required. This presents problems, as PM10 collecting equipment commonly employed, such as the Tapered Element Oscillating Microbalance (TEOM), heat the inflow to exclude moisture or use fibrous filters, resulting in a PM10 sample that may have undergone significant chemical change on the filter surface or is contaminated by filter fibres. Other systems (i.e. Negretti and Partisol) can successfully collect PM10 without chemical alteration or filter contamination. Comparative collections from Port Talbot, S. Wales suggest that TEOMs and Negretti/Partisol systems collect different PM10's; the principle difference arising from the TEOM's heating chamber, which precipitates water-soluble ions and volatilises some organic components. This results in both the mass and composition of the PM10's being altered. Particle size distributions for Negretti and Partisol collections highlighted differences mainly attributed to different flow rates. The results of this work demonstrate that simple correlations between PM10 mass and adverse health effects are problematic. Furthermore, elucidation of the complex fractionation and chemical changes in different collectors is necessary.

This investigation focuses on the application of an in vitro assay in elucidating the role of lung lining fluid antioxidants in the protection against inhaled particles, and to investigate the source of bioreactivity in urban PM10 collections from South Wales. The Plasmid Assay is an in vitro method of assessing and comparing the oxidative bioreactivity of inhalable particles. This method has provided the basis of limited toxicological studies into various inhaled xenobiotics including asbestos, and more recently PM10. Carbon Black M120 and Diesel Exhaust Particles (DEP) were tested as PM10 surrogates, DEP displaying the greatest oxidative bioreactivity. Both urban PM2.5 (fine fraction) and PM2.5-10 (coarse fraction) (Cardiff, S. Wales, UK) caused significant damage, the coarse fraction displaying higher oxidative capacity. The soluble components were found to be responsible for most of the bioreactivity in both PM sizes. Low molecular components of fresh lung lavage were found to offer most antioxidant protection, and surrogate Epithelial Lining Fluid (sELF) showed significant amelioration of DNA damage by the coarse fraction but less effect against the fine. Overall, the coarse, soluble fraction of PM10 is a great source of oxidative bioreactivity, but natural pulmonary low molecular weight antioxidants can significantly ameliorate its effects.

Airborne particulate matter has been collected from within, and proximal to, an opencast coal mine in south Wales. This work forms the first part of a three year project to collect and characterise, then determine the possible toxicology of airborne particles in the south Wales region. High-resolution Field Emission SEM has shown that the coal mine dusts consist largely of an assemblage of mineral grains and vehicle exhaust particles. SEM-EDX has shown that the mineralogical make-up of the PM10 is complex, heterogeneous, and constantly changing. These findings are supported by analytical TEM-EPXMA. However, patterns can be determined relating the mineralogical composition of the airborne particles to collection locations and mining activities within the opencast. At our study opencast, Park Slip West, quartz, which has known health effects, never exceeded 30% of the total collection mass, and average levels were much less. Vehicle exhaust emissions was the largest source in terms of particle numbers. The mass of airborne particulate matter within the pit averaged approximately twice that of outside the pit: importantly however, this higher mass was due to relatively large, and non-respirable, mineral grains. This study demonstrates that the physicochemical and mineralogical characterisation of airborne particles from mining and quarrying is essential to quantify the respirable fraction, and to identify potentially hazardous components within the PM10.