2009 Progress Report: The Chemical Properties of PM and their Toxicological Implications

EPA Grant Number: R832413C003
Subproject: this is subproject number 003 , established and managed by the Center Director under grant R832413
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).

Center: Southern California Particle Center
Center Director: Froines, John R.
Title: The Chemical Properties of PM and their Toxicological Implications
Investigators: Cho, Arthur K. , Froines, John R. , Fukuto, Jon , Harkema, Jack , Kumagai, Yoshito
Institution: University of California - Los Angeles , Michigan State University , University of Tsukuba
EPA Project Officer: Chung, Serena
Project Period: October 1, 2005 through September 30, 2010 (Extended to September 30, 2012)
Project Period Covered by this Report: August 1, 2008 through July 31,2009
RFA: Particulate Matter Research Centers (2004) RFA Text |  Recipients Lists
Research Category: Health Effects , Air

Objective:

The objective of the research in Project 3 is to characterize the chemical properties of air pollutants that relate to their effects on cells and on intact organisms.  By relating these chemical properties to biological effects, simple chemical assays can be used to predict the potential hazards of air pollutants. 

Progress Summary:

Assays
Three assays have been developed in the course of the particle center activities to quantitatively assess the capacity of a given sample to carry out chemical reactions relevant to mechanisms of toxicity. 
a.   The DTT based redox assay mimics the redox cycling of redox active chemical species that occurs intracellular utilizing cellular electron donors such as ascorbate and NADPH.  It determines redox activity due both to some metals and redox active organic species.
b.   A metal based redox assay involving the Fenton reaction was developed using the conversion of salicylate to dihydroxy benzoate isomers (DHBA) by hydroxyl, the major product of the reaction. 
c.   An assay based on the ability of electrophiles to inactivate the thiol enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by covalent bond formation with its key thiol function was developed to determine electrophile content of air pollutant samples. 
 
These assays, together with some cellular response analyses, have been used to characterize pollutants from two sources, a diesel engine and ambient air in Riverside, CA.  The objective in this period was to characterize the chemical properties of the samples in terms of the reactions above and relate the chemistry with cell responses.  In the case of the induction of the stress protein, hemeoxygenase-1 (HO-1), the results are semi quantitative and permitted a comparison or correlation analysis of the cellular and the chemical data.  In general, the DTT and GAPDH activities correlated with the HO-1 induction capacity, consistent with the role of oxidative stress in the responses.
  
Results
a. DEP studies
The assays of the DEP studies are summarized in Table 1.
Table 1.
 
 
 
 
 
 
 
 

Correlations were observed between assay data for redox, electrophile and the induction of cell stress, indicated by the protein, hemeoxygenase-1. These activities also correlated with quinones and PAHs.  We anticipated that quinones would be a major contributing species as they are both redox active and contain electrophiles, and the results are consistent with these expectations.  PAHs correlate with quinones because of the physical chemical properties, but would have to be bioactivated before they will be toxicologically significant. 
 
In further support for the role of quinones, the organic extract of one sample was subjected to the reductive acetylation procedure used to inactivate quinones in the quinone assay.  The modified organic extract was devoid of redox activity, lost about 90% of its electrophile content and about 80% of its HO-1 induction capacity (Shinyashiki, et al., 2009b).  These results support the notion that quinone or quinone-like chemical species are responsible for the chemical reactivities and cellular responses observed. 
 
A partial separation of the redox and electrophilic activities was observed by assaying aqueous suspensions of the particles before and after filtration.  Most ( 90%) of the total DTT based redox activity was associated with the particle fraction whereas 75% of the electrophilic activity was found in the filtrate, i.e., water soluble.  The metal based DHBA activity was also insoluble in water, with 100% of the activity retained by particles.  The insoluble nature of the redox activity in aqueous suspensions raised the possible involvement of humic like substances (HULIS) on the particles.  HULIS are polymeric organic substances containing a variety of functional groups such as carboxyl and alcohol which can chelate metals;removal of HULIS from particles requires strong alkali (Havers, et al., 1998).  The water soluble electrophiles likely reflect low molecular weight carbonyl compounds such as those identified in ambient air by Cahil and his associates (Seaman, et al., 2006; Spada, et al., 2008).
 
A more recent study examined the effect of heating on DEP activities.  The study was based on the report by Biswas et al. (Biswas, et al., 2009), who showed that a thermal denuder, run at 150oC, removed virtually all of the redox activity of diesel exhaust from a diesel engine controlled by a dynamometer.  Consistent with those findings, the GAPDH assay showed that all of the electrophiles in the DEPs from were removed by heating at 100oC .  Thus, a large portion of the redox and electrophiles from diesel exhaust appears to be volatile at 100 to 150oC. 
 
b. Ambient air samples
A large scale collection study is in progress, examining ambient air from Riverside, USC and Westwood.  These sites reflect a receptor site, a source site from urban traffic and a predominately small vehicle trafficked freeway, respectively.  The particles are collected on filters and the vapors trapped in resins placed below the filters.  The Riverside samples have been examined using the assays above and assays for effects on cell signaling pathways.  The study found redox activity to be concentrated in the particle phase, reflecting in part, the presence of metals, whereas the electrophiles, which are organic, appear to be mostly volatile. 
 
Although the cellular actions of the particle phase have not yet been evaluated, the vapor phase components were found to induce HO-1, a marker for inflammatory responses, at concentrations equivalent to about 1 m3.  This volume is highly relevant to exposure since human inhalation rates are about 20 m3 per day.  The contributions of the particle phase to stress induction would be in addition to this but would be redox based actions.

Future Activities:

Using the quantitative assays developed to characterize air samples for their capacity to catalyze toxicologically relevant reactions, we have found that most of the redox activity in aqueous suspensions is associated with particles, either diesel exhaust or ambient PM2.5.  The vapor phase of ambient air samples was found to exhibit most of the electrophilic activity, and preliminary experiments with DEPs indicate that their electrophiles are also volatile. 
 
The Riverside air study demonstrated the importance of vapor phase constituents in the overall actions of air pollutants.  Both protective and inflammatory pathways were activated and the chemical assays indicated that organic based redox and electrophilic activities were concentrated in this phase.  When the analyses of the other air samples are completed, assessment of the different sources of pollutants and the resulting distribution between particle and vapor phases will be determined. 
 
Although both redox and electrophilic activity have been associated with the increase in adaptation and inflammatory responses, the roles of redox or reactive oxygen generation per se and that of electrophiles has not been identified.  Future studies will address this ambiguity in two basic approaches.  In one, we will separate redox and electrophilic activities using observed physical chemical differences and use the separated, or near separated fractions in cell experiments.  In the second approach, we will perform dose and time dependent cell studies to evaluate the contributions of prooxidants and electrophiles in activating the two major cellular responses, inflammation and adaptation.
 
Our future studies will include other approaches to the separation of the redox and electrophilic components of air samples.  These will include filtration of particle suspensions, the use of sephadex-thiol preparations to trap electrophiles in an aqueous milieau and selective extractions of particles and resin traps of air samples.  The objective of these studies is twofold, one to characterize the physical properties of components exhibiting redox and electrophilic activity and the second is to use the partially separated fractions of the two activities in cellular studies to determine their effects

References:

Biswas S, Verma V, Hu S, Herner J, Ayala A, Schauer JJ, Cassee FR, Cho AK and Sioutas C. Redox activity of semi-volatile and non volatile particulate matter (PM) from heavy-duty vehicles retrofitted with emission control technologies. Environmental Science & Technology 2009;43:3905-3912.
Havers N, Burba P, Lambert J and Klockow D. Spectroscopic characterization of humic-like substances in airborne particulate matter. Journal of Atmospheric Chemistry 1998;29:45-54.
Seaman VY, Charles MJ and Cahill TM. A sensitive method for the quantification of acrolein and other volatile carbonyls in ambient air. Analytical Chemistry 2006;78:2405-2412.
Spada N, Fujii E and Cahill TM. Diurnal cycles of acrolein and other small aldehydes in regions impacted by vehicle emissions. Environmental Science of Technology 2008;42:7084-7090.


Journal Articles on this Report : 7 Displayed | Download in RIS Format

Other subproject views: All 47 publications 27 publications in selected types All 27 journal articles
Other center views: All 241 publications 157 publications in selected types All 157 journal articles
Type Citation Sub Project Document Sources
Journal Article Ayres JG, Borm P, Cassee FR, Castranova V, Donaldson K, Ghio A, Harrison RM, Hider R, Kelly F, Kooter IM, Marano F, Maynard RL, Mudway I, Nel A, Sioutas C, Smith S, Baeza-Squiban A, Cho A, Duggan S, Froines J. Evaluating the toxicity of airborne particulate matter and nanoparticles by measuring oxidative stress potential—a workshop report and consensus statement. Inhalation Toxicology 2008;20(1):75-99. R832413 (2008)
R832413 (2009)
R832413 (Final)
R832413C001 (2007)
R832413C001 (2008)
R832413C001 (Final)
R832413C002 (2008)
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R832413C003 (2009)
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  • Journal Article Biswas S, Verma V, Schauer JJ, Cassee FR, Cho AK, Sioutas C. Oxidative potential of semi-volatile and non volatile particulate matter (PM) from heavy-duty vehicles retrofitted with emission control technologies. Environmental Science & Technology 2009;43(10):3905-3912. R832413 (2009)
    R832413 (Final)
    R832413C001 (2009)
    R832413C001 (Final)
    R832413C003 (2009)
    R832413C003 (2010)
    R832413C003 (Final)
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  • Journal Article DiStefano E, Eiguren-Fernandez A, Delfino RJ, Sioutas C, Froines JR, Cho AK. Determination of metal-based hydroxyl radical generating capacity of ambient and diesel exhaust particles. Inhalation Toxicology 2009;21(9):731-738. R832413 (2009)
    R832413 (Final)
    R832413C001 (2009)
    R832413C001 (Final)
    R832413C003 (2009)
    R832413C003 (2010)
    R832413C004 (2010)
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  • Journal Article Eiguren-Fernandez A, Miguel AH, Lu R, Purvis K, Grant B, Mayo P, Di Stefano E, Cho AK, Froines J. Atmospheric formation of 9,10-phenanthraquinone in the Los Angeles air basin. Atmospheric Environment 2008;42(10):2312-2319. R832413 (2007)
    R832413 (2008)
    R832413 (Final)
    R832413C003 (2007)
    R832413C003 (2008)
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  • Journal Article Eiguren-Fernandez A, Miguel AH, Di Stefano E, Schmitz DA, Cho AK, Thurairatnam S, Avol EL, Froines JR. Atmospheric distribution of gas-and particle-phase quinones in Southern California. Aerosol Science and Technology 2008;42(10):854-861. R832413 (2008)
    R832413 (Final)
    R832413C003 (2009)
    R832413C003 (2010)
    R832413C003 (Final)
    R827352 (Final)
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  • Journal Article Shinyashiki M, Eiguren-Fernandez A, Schmitz DA, Di Stefano E, Li N, Linak WP, Cho S-H, Froines JR, Cho AK. Electrophilic and redox properties of diesel exhaust particles. Environmental Research 2009;109(3):239-244. R832413 (2008)
    R832413 (Final)
    R832413C003 (2009)
    R832413C003 (2010)
    R832413C003 (Final)
    R832413C004 (2010)
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  • Journal Article Taguchi K, Shimada M, Fujii S, Sumi D, Pan X, Yamano S, Nishiyama T, Hiratsuka A, Yamamoto M, Cho AK, Froines JR, Kumagai Y. Redox cycling of 9,10-phenanthraquinone to cause oxidative stress is terminated through its monoglucuronide conjugation in human pulmonary epithelial A549 cells. Free Radical Biology and Medicine 2008;44(8):1645-1655. R832413 (2007)
    R832413 (2008)
    R832413C003 (2007)
    R832413C003 (2008)
    R832413C003 (2009)
    R832413C003 (2010)
    R827352 (Final)
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  • Supplemental Keywords:

    Bioavailability, metabolism, chemicals, oxidation-reduction, biochemistry, ambient air, particulate matter, oxidative stress, RFA, Health, Scientific Discipline, Air, particulate matter, Health Risk Assessment, Risk Assessments, Biochemistry, Ecology and Ecosystems, atmospheric particulate matter, particulates, chemical assys, particle matrix, human health effects, PM 2.5, chemical characteristics, toxicology, airway disease, airborne particulate matter, cardiovascular vulnerability, air pollution, human exposure, vascular dysfunction, cardiovascular disease, human health risk

    Progress and Final Reports:

    Original Abstract
  • 2006 Progress Report
  • 2007 Progress Report
  • 2008 Progress Report
  • 2010 Progress Report
  • 2011
  • Final Report

  • Main Center Abstract and Reports:

    R832413    Southern California Particle Center

    Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
    R832413C001 Contribution of Primary and Secondary PM Sources to Exposure & Evaluation of Their Relative Toxicity
    R832413C002 Project 2: The Role of Oxidative Stress in PM-induced Adverse Health Effects
    R832413C003 The Chemical Properties of PM and their Toxicological Implications
    R832413C004 Oxidative Stress Responses to PM Exposure in Elderly Individuals With Coronary Heart Disease
    R832413C005 Ultrafine Particles on and Near Freeways