The Characteristics and the Cytotoxic Effects of Particulate Matter in the Ambient Air of the Chiang Mai Lamphun Basin in Northern Thailand

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Narongpan Chunram1, Usanaee Vinitketkumnuen2, Richard L. Deining3, and Richard M. Kamens4

'Department of Applied Science, Faculty of Science and Technology, Chiang Mai Rajabhat University, Chiang Mai 50300, Thailand department of Biochemistry, Faculty of Medicine, Chiang Mai University,

Chiang Mai 50200, Thailand department of Chemistry and Biochemistry, California State University at Fullerton, Fullerton, CA 92834 department of Environmental Science and Engineering, University of North Carolina, Chapel Hill, NC 27599

There are growing concerns about the deteriorating air quality in the Chiang Mai-Lamphun Basin in Northern Thailand resulting from accelerating development and construction, increasing industrial emissions, expanding transportation, extensive open burning and the continuing drought. Recent studies of the levels of fine particulate matter (PM2 5 and PMjo) in the Basin have highlighted their seasonal variations, correlations with other pollutants, and toxicological properties. These are reviewed and implications for future research are summarized.

© 2009 American Chemical Society


The effects of airborne pollutants on human health have been widely recognized and extensively examined. Results of numerous studies around the world have documented the increases in respiratory distress, pulmonary disease and cancer, leading to increased hospital admissions and higher mortality associated with a variety of pollutants (1-10). Of special concern is particulate matter (PM) of less than 10 microns (PMi0), and, especially, of the fine particles of less than 2.5 microns (PM2 s) (//). Air pollution problems in urban areas are especially significant because of the large numbers of people that can be affected by the numerous and varied sources of pollutants in these densely settled areas. In addition, rural areas surrounding cities often have significant impact on urban centers because of the transport of materials under varying climatological, atmospheric and geophysical conditions.

This review highlights some of the important data and conclusions of recent investigations (12-15) that focus on the rapidly growing Chiang Mai-Lamphun basin in Northern Thailand (Figure 1). The work reviews the data on the levels and distribution of <10 \im and <2.5 [im particulate matter, PM10 and PM2 5, respectively, along with the mutagenicity and cytotoxicity of samples collected on filters in the Chiang Mai-Lamphun Basin, providing information about the extent of the problem and the potential health effects.

The adjacent provinces of Chiang Mai (population 1.5 million) and Lamphun (0.45 million) are situated in the northern part of Thailand in the mountainous and forest-covered region near Myanmar (Burma) and Laos. The abundant rainfall and fertile hills and valleys in the region support agricultural crops such as rice, sugar cane, citrus, coffee and vegetables. It is one of the most attractive year-round tourists destinations in Southeast Asia. The Chiang Mai provincial capital city (metropolitan population 0.7 million) and nearby provincial capital city of Lamphun incorporate urban, suburban and industrial areas and are located in a basin between mountain ranges. This fast-growing region that is experiencing many environmental stresses.

The three distinct climatic seasons in Northern Thailand have significant effects on air quality. The rainy season extends from 1 June to 15 October, the winter, or dry, season is from 16 October to 15 March, and the summer season is from 16 March to 31 May. The basin geography creates the conditions for frequent thermal inversions at certan times of the year, especially in the cooler winter months, trapping pollutants and limiting air flow and mixing. The year-round heavy vehicular traffic (not subject to inspection and much of it relying on diesel fuel) and increasing industrial development make noticeable contributions to the atmosphere by lowering the air quality. Cycles of construction of roads, superhighways and buildings make periodic and significant contributions to dust and particulate levels.

Figure 1. Map of Thailand highlighting the Chiang Mai-Lamphun Basin.

It is common agricultural practice in the months of January through April to burn dry crops to reduce the vegetation and prepare the fields for planting. Also, frequent forest fires in the dry (winter) season are often set by villagers in order to clear dense underbrush in forested areas of the surrounding mountains. Contributions from fires in neighboring Myanmar, and even from as far-away as Indonesia, add to the particulate load in the atmosphere. Although air quality is generally best during the rainy season, there may still be periods of locally high levels of pollutants. As a result of all of these conditions, there are frequent increases in hospital admissions and significant health threats from the high levels of pollutants many days of the year.

A recent study (16) found that lung cancer rates in Chiang Mai, Thailand, were found to be the second highest in the world. The high incidence of cancer, especially of lung cancer, in Chiang Mai relative to other places in Thailand was reported in 1993 (17) and considerable effort has been devoted to examining the contributions from components in polluted air. In 2002, a study (12) was published on the levels and mutagenecity of airborne PMio and PM25, in urban areas of Chiang Mai using data collected during the period March 1998 to October 1999.

One of the greatest challenges in determining the specific agents that cause the detrimental effects of air pollution on organisms is the complexity of the samples. It is clear that some of the most potent toxins are the polycyclic aromatic hydrocarbons, PAH's, but a number of other organic and inorganic compounds are also capable of inducing cellular changes. Initial screening studies usually are usually based on genotoxic or mutagenic properties of a mixture of materials extracted from filter samples by applying extracts to specific cells or cell lines (18-28). Studies focusing on the levels and impact of particulate matter and other pollutants in the Chiang Mai - Lamphun Basin are summarized below, along with their key findings.


Particulate samples were collected for 24-hour periods using mini-volume air samplers (AIRmetrics MiniVol portable samplers, USA) on 47 mm fiber-film filters (Pallflex, USA) with <2.5 ¡xm and <10 nm cutoffs for PM25 and PM,0 measurements, respectively, operating at 2.5 L/min.. Sampling sites in Chiang Mai included a residential apartment and a research laboratory, both near heavy traffic areas a rural area with limited direct automotive traffic exposure, a commercial market on a busy street, a site in an area known to have a high incidence of lung cancer. In Lamphun, they included both industrialized and urban areas. For daily measurements, filters were conditioned in an electronic dessicator at 25°C and relative humidity of 50% before and after sample collection and weighed on a micrometric balance (Sartorius AG, Germany) to the nearest 0.001 fig. Meterological data (wind direction, wind speed and humidity levels) were collected with standard equipment mounted near selected sites, and levels of S02, N02 and 03 used for correlations were obtained from local and provincial governmental pollution monitoring agencies (13).

For toxicity studies following mass measurements (14), filters collected over one month's time were were cut into small pieces, pooled, and sonicated for 15 minutes in 200 mL of dichloromethane. After addition of anhydrous sodium sulfate, the extract was filtered through Whatman No. 41 filter paper. Sonication was repeated two more times and the samples were dried at 35°C on a rotary evaporator. For mutagenicity studies on the residue, the Ames test utilized Salmonella typhimurium strains TA98 and TA100, with proper controls, as described previously (12). Revertant colonies per plate were counted and the toxic effects were examined under a stereomicroscope.

For cell viability and DNA fragmentation studies (75), the residue was quantitatively resolved in culture media and filtered through a Millipore membrane to ensure sterility and stored in the dark at 4°C in sealed vials. Human alveolar type II-derived cell line A549 and alveolar macrophages MH-S were grown on F-12KDMEM(1:1) and RPMI-1640 media, respectively, supplemented with 10% fetal bovine serium (FBS), 100 U/mL penicillin, and 10 mg/mL streptomycin in a humidified atmosphere at 37°C and 5% C02. The 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide (MTT) assay was used for cell viability measurements, as described earlier (12). DNA fragementation was examined by agarose gell electrophoresis with ethidium bromide staining, following lysis in buffers, incubation with RNase A and then Proteinase K, precipitation with NaCl, centrifugation, washing, and dissolution in TE buffer.

Results and Discussion

The daily levels of PM25 and PM10 were measured at several sites in the Basin covering the period of June 2004 through May 2006. The results from the residential sampling site are illustrated in Figure 2 for the period of June 2004 to May 2005, a typical annual pattern for all sites.

Average seasonal variations at two sampling sites are summarized in Table I for the period June 2004 to May 2006. As noted in the table, the end of the winter (dry) season and the beginning of the summer season show extremely high levels of particulate matter. The winter season is characterized by cold weather and frequent thermal inversions with limited dispersion from low wind speeds, and increased biogenic emissions from local burning. During those times, average PM2 5 mass concentrations at various sites ranged from 29.9 (rural area) to 44.5 |ig/m3 (commercial market) at a miniumum, and the maximum 24hour levels ranged from 115.1 fig/m3 (research laboratory) to 257.5 ng/m3 (high lung cancer area). The new 2006 USEPA 24-hour standard for PM2 5 of 35 |ig/m3 was exceeded 30-50% of the days during the study periods.

A number of studies have focused on meteorological contributions to particulate matter and correlations with other constituents (29-31). Correlations of PM2 5 with PM10 with other atmospheric constituents, S02 and N02, in the Chiang Mai-Lamphun Basin are positive and high, with significant variation for different wind directions and speeds. With respect to ozone, 03, the unusually high level in summer is consistent with the suggestion (32) that atmospheric chemistry, rather than transport, may be responsible for these high summer concentrations. High concentrations of particulate matter occur with moderate southerly winds (6-12 m/s blowing from the industrial areas and the international airport, moderate temperature (20-25 °C), and low humidity (near 50%) during winter.

Time (Days)

Figure 2. Daily levels of PM25 and PM10 (/¿g/m3) at a residential site during the period of June 2004 (1/6/04) through May 2005 (31/5/05).

(Modifiedfrom Reference 13 .)

Time (Days)

Figure 2. Daily levels of PM25 and PM10 (/¿g/m3) at a residential site during the period of June 2004 (1/6/04) through May 2005 (31/5/05).

(Modifiedfrom Reference 13 .)

The 2002 study (12) demonstrated mutagenicity to Salmonella typhimurium strain TA100 without activation for extracts of airborne particulate matter collected in winter in Chiang Mai. Sites included a fifth-floor balcony of a multidisciplinary building Chiang Mai University, the ground level outdoor patio of a residential home near the same high-traffic area, and a downtown market area. Monthly average PM2.s levels ranged from 17 to 138 ng/m3 and PMio levels ranged from 27 to 173 fig/m3, and daily levels in February and March reaching as high as 208 ng/m3. Mutagenicity increased in the presence of metabiolic activation (S9 mix) and appeared to track particle concentrations. Indirect mutagenicity in airborne particulate matter extracts were detectable during the winter months, October to March. Direct-acting mutagenicity was detected at one site, and mutagenic activity was higher in the presence of enzyme activation.

The more recent study (14) utilizing Salmonella typhimurium provides additional information about the toxicity of particulate matter in the 24-hour filter samples collected in urban areas of Chiang Mai from June 2004 to May 2006. With both indoor and outdoor sampling, the types of mutations induced by the complex environmental samples were found to be the same types as those

Table I. Seasonal averages of PM25 and PMi0 for two sites in the Chiang Mai-Lamphun Basin from June 2004 to May 2006




















Std Dev
















Std Dev
















Std Dev



Note: All numeric values are in jig/m3 Source: Modified from Reference 13

Note: All numeric values are in jig/m3 Source: Modified from Reference 13

in humans exposed to the same environmental mutagens. Indirect-acting mutagenicities were detected in both sites in all samples. The seasonal trends were higher levels in the winter months and lower to undetectable levels in the summer and rainy seasons. It has been noted that organic extractable matter from air particles and different combustion sources are carcinogenic in animals and mutagenic in short-term bioassay tests (23). Also, vehicle emissions have been shown account for most of the mutagenic activity associated with air particles in urban areas (24). Additional contributions from cooking and tobacco smoke are likely. Finally, during high ozone (03) and N02 episodes, OH radicals can facilitate formation of nitro-PAHs which have higher mutagenicity than PAHs (25).

Another study (15) using human cell line A549 (alveolar type II-derived cell line) and MH-S alveolar macrophages, involved screening for cytotoxicity and apoptosis (cell death) induction in cultured lung cells and macrophages, using 24-hour filter samples for the period from July 2004 to May 2005. MTT (3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide) assays in human lung cells and alveolar macrophages were carried out, along with DNA fragmentation to detect apoptosis induction. The PM25 and PMi0 components were cytotoxic to lung cells and alveolar macrophages, and, at one site in Lamphun province, apoptosis of alveolar macrophages was induced. High cytotoxicity (about 40%) was observed for 72-hr exporsure to PM2.5 and PMi0 extracts, with larger effects on MH-S than on A549 lung cells. Apoptosis of alveolar macrophages was observed at one site in Lamphun province for samples collected in May, near the end of the summer season. These results are important because they imply that exposure to ambient particulate matter may be related to short-term and as well as long-term respiratory and health problems.


The studies in the Chiang Mai-Lamphun basin described above suggest that there is a growing need for regular and continuous monitoring of PM2.5 and PM10 levels. Particulate matter formation is not limited to only urban areas, as can be clearly seen by high levels of smoke and haze in much of the region due to open burning and dry conditions. Sampling sites outside of the central city have noticeably high levels of particulates, as well. If pollution regulations are to be established and enforced, and if the health of the people is to be improved, monitoring to obtain solid ongoing scientific information must continue and results must be made available to government officials and local planners.

Additional toxicological research is needed to more closely examine detrimental effects of the PM25 and PM10 on human health. More detailed analytical measurements involving identification and quantitation of particular chemical species need to be developed in order to identify specific compounds in the local particulate matter that are responsible for the adverse health effects and facilitate the fingerprinting of possible sources. Finally, improved regulation and monitoring, greater public awareness and education, and expanded involvement of public health professionals and local leaders will contribute greatly to solving this ongoing problem and lead to improved health.


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