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Effects of cerium oxide and ferrocene nanoparticles addition as fuel-borne catalysts on diesel engine particulate emissions: Environmental and health implications Zhi-Hui Zhang, and Rajasekhar Balasubramanian Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.7b00920 • Publication Date (Web): 27 Mar 2017 Downloaded from http://pubs.acs.org on March 29, 2017
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Environmental Science & Technology
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Effects of cerium oxide and ferrocene nanoparticles addition as fuel-
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borne catalysts on diesel engine particulate emissions: Environmental
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and health implications
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Zhi-Hui Zhang, Rajasekhar Balasubramanian*
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Department of Civil and Environmental Engineering, Faculty of Engineering, National University of Singapore, 1 Engineering Drive 2, E1A 02-19, Singapore 117576, Singapore
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* Corresponding address: Tel.: +65 65165135; fax: +65 67744202 E-mail address:
[email protected] (R. Balasubramanian).
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ABSTRACT
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This study systematically examined the potential impacts of doping CeO2 and Fe(C5H5)2
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nanoparticles as fuel-borne catalysts (FBCs) to ultralow sulfur diesel (ULSD) fuel on the
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physical, chemical and toxicological characteristics of diesel particulate matter (DPM). The
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FBCs-doped fuels are effective in promoting soot oxidation and reducing the DPM mass
29
emissions, but lead to a significant increase in the total particle counts due to the formation of
30
self-nucleated metallic nanoparticles. Compared to undoped ULSD, the FBCs-doped fuels result
31
in higher concentrations of particle-phase polycyclic aromatic hydrocarbons (PAHs) and n-
32
alkanes, higher fractions of organic carbon (OC) and water-soluble organic carbon (WSOC) in
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particles, show slight alterations in soot nanostructure, reduce soot ignition temperature and
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activation energy. Exposure of the human-type II cell alveolar epithelial cells (A549) to DPM
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derived from FBCs-doped fuels shows a decrease in cell viability and alterations in the global
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gene expression with a broad range of biochemical pathways. The overall variations in DPM
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characteristics are mainly caused by the catalytic combustion process, and are related to the type,
38
properties and contents of FBCs used in diesel fuel as well as the engine operating conditions.
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Environmental and health implications of the study are highlighted.
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Keywords: Diesel particulate matter; Fuel-borne catalysts; Cerium oxide; Ferrocene; Toxicity
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Environmental Science & Technology
INTRODUCTION
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High emissions of airborne particulate matter (PM) from diesel engines used in both on-
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road and off-road applications are of global concern as they have adverse impacts on urban air
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quality, human health, and also affect global climate change.1,
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technology and the application of post combustion-treatment systems in conjunction with diesel
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fuel reformulations are critically needed for abatement of diesel particulate matter (DPM)
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emissions.3 One of the fuel reformulation technologies under consideration is the addition of an
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ultra-low dose of fuel-borne catalysts (FBCs) in the form of organometallic nanoparticles to
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diesel fuels. This approach is explored for suppressing soot formation and/or promoting soot
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oxidation during the combustion process, and thus reducing DPM mass emissions.3-8
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additional consideration is that if FBCs are used in combination with diesel particulate filters
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(DPFs), the FBCs embedded in the emitted DPM could enhance soot oxidation in the DPFs and
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therefore promote DPFs regeneration.9 FBCs containing Ce, Fe and Pt are now commercially
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available, among which both Ce- and Fe-based FBCs have been increasingly used in Europe and
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elsewhere as diesel fuel additives to ensure fast and complete DPFs regeneration.7-11 These
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FBCs have also been used without DPFs in existing off-road diesel engines and commercial
58
vehicles to reduce soot and DPM emissions.6-8 However, these FBCs seem to be restricted from
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using in several countries. For example, in the U.S., current regulations restrict their use to on-
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road diesel engines. In Switzerland, FBCs can only be used in combination with DPFs, but their
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use without particle traps is generally prohibited. These restrictions have been placed because of
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concerns over the potential environmental and public health impacts of FBCs.
2
Improvements in engine
An
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DPM emissions vary significantly in physical characteristics and chemical composition
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between different engine types, engine operating conditions, and fuel formulations. Some recent
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studies have reported that the catalytic combustion process, caused by FBCs-doped fuels, not
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only reduce DPM mass concentrations efficiently, but also trend to alter the physical and
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chemical properties of DPM.7-8,
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composition of DPM into account are deemed to be more effective and efficient because the
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chemical characteristics of DPM rather than the DPM mass determine their potential influence on
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the environment and human health. In addition, the application of metal-based FBCs in diesel
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fuels inevitably causes the emissions of the metals themselves in DPM which, combined with
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changes in chemical composition and/or particle size distributions, might lead to changes in the
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overall toxicity of DPM.15-17 However, the potential adverse environmental effects and health
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risks resulting the use of Ce-based and Fe-based FBCs are poorly understood due the lack of
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systematic investigations.
12-14
Emission reduction strategies that take the chemical
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To address these knowledge gaps, we systematically investigated the changes in the
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physicochemical and toxicological characteristics of DPM emissions resulting from the diesel
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engine fuelled by CeO2- and Fe(C5H5)2- doped fuels by using a series of complementary
79
analytical techniques. The central goal of this work is to assess their potential environmental and
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human health implications. Specifically, we examined the influence of doping different
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concentrations of these additives to ultralow sulfur diesel (ULSD) fuel on particulate mass
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concentration as well as particle number concentration with their corresponding size distributions.
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We then analyzed the effects of these FBCs-doped fuels on the carbonaceous contents of the
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DPM including elemental carbon (EC), organic carbon (OC), and water-soluble organic carbon
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(WSOC). We selected PAHs and n-alkanes as appropriate chemical species of OC for further
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discussion as PAHs are suspected human carcinogens and n-alkanes represent an important class
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of the organic compounds originating from unburned fuel and/or lubricating oil. In addition, we
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studied the particle volatility and soot oxidation behavior to understand the effects of these
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FBCs-doped fuels on nanoparticle formation and soot oxidation reactivity. Determination of the
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influence of FBCs-doped fuels on the above-mentioned properties of DPM is essential for
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understanding how FBCs-doped fuels affect the DPM formation process and its chemical
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composition. The changes in the physico-chemical characteristics are likely to affect the toxicity
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of DPM. In order to examine this possibility, we exposed the particles derived from both FBCs-
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doped and undoped fuels to the human-type II cell alveolar epithelial cell line (A549) and
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assessed their toxicity by using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
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bromide) assay. The cDNA microarray technique was then used to assess the changes in the
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expression of gens A549 in an attempt to gain an insight into the complex mechanisms that
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underlie the health effects induced by DPM derived from FBCs-doped fuels in comparison with
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undoped diesel fuel. We selected an off-road diesel engine for this study as such engines are
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widely used and emit a substantial fraction of DPM on a global level due to the lack of effective
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environmental regulations.18 A long-term exposure to high concentration of freshly emitted DPM
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has been reported to pose a significant carcinogenic risk to occupational workers in various
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working microenvironments where off-road engines are frequently used.19
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EXPERIMENTAL SECTION
105 106
The essential details of the experimental work are explained briefly here, and more detailed information is given in Supporting Information (SI).
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Engine, Fuels and FBCs. The schematic of the experimental system is given in Figure
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S1. Experiments were carried out on a single-cylinder, naturally aspirated, four-stroke, direct-
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injection diesel engine (L70AE, Yanmar Corporation) connected to a 4.5 kW generator. The
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diesel engine has a capacity of 296 cm3 with bore and stroke of 78 mm and 62 mm, a fixed speed 5 ACS Paragon Plus Environment
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of 3000 rpm (revolutions per min). The more detailed specifications of the engine are provided in
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Table S1. Similar small off-road diesel engines have been used previously for investigating the
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influence of FBCs-doped fuels on engine exhaust emissions.4-8, 13, 14
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ULSD with less than 10-ppm (parts per million) by weight of sulfur was used as a base
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fuel, and the major properties of this fuel can be found elsewhere.20 The FBCs used in this study
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include cerium oxide (CeO2; Sigma-Aldrich, ≥ 99.9%) nanopowder with particle size of less than
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25 nm, and the analytical grade ferrocene (Fe(C5H5)2; Sigma-Aldrich, ≥ 98%) in the form of
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commercially available nanoparticles. CeO2 and Fe(C5H5)2 were added to ULSD in three
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different concentrations (25, 50, and 100 ppm Ce and Fe by weight, which are designated as
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Ce25, Ce50 and Ce100 for the CeO2-doped fuels, and Fe25, Fe50 and Fe100 for the Fe(C5H5)2-
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doped fuels, respectively.). Both CeO2 and Fe(C5H5)2 were added to the diesel fuel as per the
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established procedure, 8, 14, 21 which can be found in the SI.
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Particulate sampling and measurement. A two-stage Dekati mini-diluter (DI-2000,
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Dekati Ltd.) was used for diluting the engine exhausts for DPM sampling and online evaluation.
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The dilution ratio from each stage was determined, and all data presented in this article have
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been dilution-corrected to represent tailpipe conditions. DPM emissions from the first-stage
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diluter were collected onto 47 mm Teflon filters (Pall Life Sciences, Ann Arbor, MI, 2 µm pore
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size) and on 47 mm quartz fiber filters (Whatman, USA, 2.2 µm pore size) for the subsequent
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analyses, by concurrently using two Mini-Vol particulate samplers (Air metrics Ltd.; 5 L/min
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flow rate). The blank quartz fiber filters were pre-combusted in air (650 ℃ for 12 h) to remove
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any residual carbon contamination prior to being used for particles collection. Further details
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about determination of the dilution ratio and the handling of filters and DPM samples are
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provided in the SI. The number concentrations and size distributions of volatile and solid
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particles in the secondary dilution stage were measured by a Fast Mobility Particle Sizer (FMPS,
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Model 3091, TSI Incorporated, USA) for particles over the size range of 5.6 to 560 nm. In this
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setup, two diluters were used in series, with the first stage being heated to maintain a temperature
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of 190 ℃ to minimize thermophoretic deposition. During the solid particle number emission
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experiments, a thermodenuder (TD, Dekati Ltd) with a heated temperature of 265 ℃ was placed
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in-line between the secondary stage diluter and the FMPS for removing the volatile and semi-
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volatile compounds of particles. The TD diffusion losses were estimated using a widely used
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method,22 and the diffusion loss-corrected particle size distributions are presented in this article.
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Physical analyses. A thermogravimetric analyzer (Discovery TGA, TA instruments) was
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used to investigate the ignition temperature of soot followed by the method reported previously,
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22
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Arrhenius expression.23 Raman spectra of the DPM samples were recorded with a Renishaw
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microscope system (Renishaw, System 2000) using a 514 nm Ar ion laser as an excitation source.
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The spectra were then analyzed by using the Renishaw WIRE 2.0 software running under
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GRAMS/32 (Galactic, Levenberg–Marquardt nonlinear least-squares fitting algorithm) to obtain
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the D (Defect)/G (Graphite) band intensity ratios (ID/IG) for investigation of the graphite-like
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structure of soot particles. Further details of these analyses are provided in the SI.
and the activation energy for the soot was then estimated using a modified form of the
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Chemical analyses. A thermal/optical carbon aerosol analyzer (Sunset Labs, Forest
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Grove, Oregon, USA) was used to quantify EC and OC fractions of particles collected onto
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quartz filters according to the NIOSH 5040 reference method.24 Meanwhile, the particles
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collected on Teflon filters were extracted with organic-free Milli-Q water (>18 MΩ cm) using a
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water-bath sonicator (Elmasonic S 60H, ELMA, Germany) for 2 x 30 min. The extracts were
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then filtered through a 0.20 µm PTFE membrane filter (Satorius, Germany), acidified by 7 ACS Paragon Plus Environment
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ultrapure HCl (0.1 ml of 2M HCl), and purged for 2 min with ultrapure air. 100 µl of the
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acidified filtrate was injected into an organic carbon analyzer (TOC-VCSH, Shimadzu) for
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determination of WSOC.
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A
Unity/Ultra
thermal
desorption
system
coupled
with
Agilent
6890N
161
chromatography/5973N mass spectrometry (TD-GC/MS) was used for the determination of
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particle-phase PAHs and n-alkanes. The details of the method are provided by Ho and Yu.25 In
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this study, the 16 US priority PAHs were measured and then classified into three groups based
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on their molecular weights, namely low molecular weight (LMW, 2 and 3 rings) PAHs, medium
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molecular weight (MMW, 4 ring) PAHs and high molecular weight (HMW, 5 and 6 rings) PAHs,
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as suggested in our previous study.20 Therefore, the total-PAH data represents the sum of the 16
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individual PAHs. The overall toxicity is defined as the sum of Benzo[a]pyrene equivalent (total
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BaPeq), which was estimated based on the method of Nisbet and LaGoy.26 For particle-phase n-
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alkanes, concentrations of the homologous series from C14 to C26 were quantified. The
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concentrations of n-alkanes with molecular weight lower than C14 and higher than C26 were
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quite low and therefore are not reported in this article.
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Toxicological experiments. In this study, DPM emitted from pure ULSD and from 100
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ppm Ce- and 100 ppm Fe-doped fuels at medium engine load was chosen for a comparative
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evaluation of their toxicological characteristics. We have made this selection because particles
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emitted under these conditions contained the highest amount of Ce and Fe when the engine was
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operated with the most commonly used engine load. These particles were physically removed
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from the Teflon filters by sonication, and re-suspended in DMSO (Dimethyl sulfoxide,
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Bioreagents, Sigma-Aldrich) and subsequently in cell exposure medium using previously
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validated procedures,27 as summarized in the SI. The extraction efficiencies were evaluated, and
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the extracted particulate mass was efficiency-corrected.
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We exposed the particles to the human-type II cell alveolar epithelial cell line (A549).
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The cytotoxicity of these particles was measured by using the MTT assay and the resulting
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changes in the expression of global genes were carefully assessed by using the cDNA microarray
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technique. Method details are given in the SI. For the MTT assay, the cells were exposed to
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concentrations of particles in the cell culture wells that corresponded to 25, 50, 100, and 200
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µg/mL of particles, respectively, for 24 hours. Each sample was tested in six replicate wells, and
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the data on cell viability are reported as relative decrease compared to the control, considered as
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100% of viable cells. As for gene expression, the A549 cells were exposed to 50 µg/ml of
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particles for 6 hours. The gene expression level was considered significantly altered based on a
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1.5-fold cut-off for the fold-change. The shortlisting of genes obtained from microarray data was
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conducted using the Gene Ontology (GO) analysis. The identified gene list has undergone
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functional annotation clustering using the DAVID web resource (http://david.abcc.ncifcrf.gov/).
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The examined categories are Biological Processes and KEGG.
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QA/QC and statistical analysis. The analytical quality of the data obtained on the
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chemical composition of DPM was ascertained using the lower limit of detection (LOD),
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recovery, the linearity of calibration, and the relative standard deviation (RSD) of replicate
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analyses. The details of quality assurance and control (QA/QC) protocols for chemical analyses
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are provided in SI. The LODs for OC and EC were below 0.47 mg/m3. The RSD of duplicate
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analyses was lower than 5.8 % for TC (total carbon), and below 10% for OC and EC. The LOD
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for WSOC was below 0.6 µg/m3. After the correction for the blank contribution, the recoveries
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for WSOC were 82.9-94.8% by the TOC method, and the RSD of four replicate analyses was
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estimated to be within 6.5%. A five-point calibration for WSOC standard solution was
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performed, and the correlation coefficients (R2) obtained from the linear regression of the
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calibration curves was larger than 0.99. For PAHs and n-alkanes, the LODs were calculated to be
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in the range of 2.1–35.8 ng/m3 for PAHs and 2.9–29.2 ng/m3 for n-alkanes, respectively. The
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RSDs of four replicate analyses were 4.5 to 10.3% for the PAHs, and were 4.8 to 7.5% for n-
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alkanes, respectively. An eight-point calibration over a concentration range of 0.1– 5.0 ng for
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each PAH and n-alkane from the liquid standard mixtures (Sigma-Aldrich, Bellefonte, PA, USA)
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was established, and the correlation coefficients (R2) for linear regressions of the calibration
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curves was greater than 0.99. In this study, all analytical data were corrected for the average
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value of the blanks, and are reported as the mean values. The significance of the differences of
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all tested parameters between diesel fuel and FBCs-doped fuels, and between the two kinds of
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FBCs-doped fuels under the same FBC concentration was assessed by Student's t test at p < 0.05.
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The results of particle number size distributions are reported as the mean values, with the relative
215
standard deviations ranging from 2.5 to 3.2%, respectively. With the exception of particle-phase
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n-alkanes and particle number size distributions, all other reported values are presented as mean
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± SD (standard deviations).
218
RESULTS AND DISCUSSION
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Particulate mass and carbonaceous fractions. Figure 1(a) shows that both Ce- and Fe-
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doped fuels could effectively reduce the particulate mass concentrations, which is associated
221
with the variation of the EC and OC emissions, as evident from Figures 1(b) and (c), respectively.
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Specifically, the EC emissions generally decreased with an increase in both Ce and Fe
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concentrations in ULSD. The degree of the EC reduction at lower engine load was greater than
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that at higher engine load. In addition, the Fe100 was generally more effective in inhibiting EC 10 ACS Paragon Plus Environment
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emissions than the Ce100 at a higher engine load. Similar results in the soot or smoke emission
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reduction were reported for Ce-doped fuels, 4, 5 and for Fe-doped fuels.7, 8 Interestingly, we found
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that with the exception of Ce100, all other FBCs-doped fuels did not make significant changes in
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OC emissions. Thus, our finding suggests that the decrease in DPM emissions, caused by FBCs-
229
doped fuels, can be attributed mainly to the EC emission reduction. Moreover, the substantial
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reduction in EC emissions coupled with the relatively minor change in OC emissions for both
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Ce- and Fe-doped fuels contributes to the higher OC/EC ratios (Figure 1(d)) than that for
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undoped ULSD. Compared to Ce100, the higher EC emission reduction coupled with the lower
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OC emission reduction for Fe100 led to higher OC/EC ratios. These observations suggest that a
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high proportion of semi-volatile organics deposited on the surface of soot particles emitted from
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these FBCs-doped fuels, especially from Fe-doped fuels. Another interesting finding of our study
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is that the Ce50 and Ce100 led to a 3.7 to 8.1% higher particulate mass emission reduction than
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that of Fe50 and Fe100, indicating Ce-based FBC being more effective in inhibiting DPM
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emissions than that of Fe-based FBC under these conditions.
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Figure 1(e) shows that WSOC mass concentrations increased significantly for both Ce-
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and Fe-doped fuels at the high engine load, leading to the higher WSOC/OC (Figure 1(f)) ratios
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for these conditions. However, at low and medium engine loads, the higher WSOC/OC ratios
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only occurred for the addition of 100 ppm Ce to ULSD. In other cases, the WSOC/OC ratios for
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FBCs-doped fuels showed no significant difference compared to that for undoped ULSD. In
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addition, for each test condition, the WSOC/CO ratios for Ce-doped fuels showed slightly higher
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than that of Fe-doped fuel, but were not statistically significant. Our results indicate that the
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FBCs-doped fuels tend to increase the water soluble fraction of OC in these cases, suggesting an
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increased fraction of polar organic compounds in OC in these cases. These observations imply
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that the FBCs-doped fuels have potential to change in the particle toxicity due to the increased
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emission of both OC and WSOC fractions in DPM although they showed an overall decrease in
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total particle mass concentrations.
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A fundamental understanding of the formation of new particles and their growth in diesel
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engines may explain how FBCs-doped fuels affect the carbonaceous particulate emissions. The
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details can be found in the SI. Briefly, with the addition of FBCs to diesel fuel, the organic
254
component of ferrocene decomposes in the early stage of the combustion process before the
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onset of soot particle inception due to its lower melting point, 28 and is most dominantly present
256
in the form of amorphous Fe (III) oxide crystallizing to hematite α-Fe2O3.29 Then, the iron oxide
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nuclei and nanoparticles are formed for the subsequent deposition of the carbonaceous phase. As
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for CeO2, its nanoparticulate structure would remain unchanged due to its very high melting
259
point (2200–2400℃). During the combustion process, these metal oxide catalysts have a high
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potential to promote carbon oxidation due to the redox mechanisms, where metal oxide catalysts
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act as oxygen suppliers for the gasification of carbon and are then re-oxidized by external
262
oxygen.30-32 Therefore, the high external surface area of the catalyst is crucial for the oxidation of
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soot. In this study, we found that the doping of both CeO2 and Fe(C5H5)2 can lead to higher
264
counts of metallic nanoparticles as explained in following section. As the iron metal oxide
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particulates tend to affect the soot formation, they are likely to serve as condensation nuclei for
266
early than the soot inception and therefore enhance soot formation. Conversely, our observations
267
show that the higher concentrations of Fe in diesel fuel led to the lower EC emissions. We
268
therefore believe that the surface enrichment of the catalysts with the soot formed from FBCs-
269
doped fuels can facilitate easier oxygen transfer from the surrounding air to the metal oxide,
270
accelerating the redox process and consequently enhancing the oxidation behavior. With an
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increase in the engine load, the counts of metallic nanoparticles decreased as found in this study,
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which suggests a reduced amount of contact points between soot and catalysts, and therefore led
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to lower percentage reduction of soot emission. Moreover, when compared to CeO2, the
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decomposition of the same dose of Fe(C5H5)2 produced higher counts of smaller metal oxide
275
nuclei as evident from the higher counts of nanoparticle emissions (see the following section).
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These Fe-containing nanoparticles, combined with their higher mobility due to the lower melting
277
point, create more contact points for enhancing the soot/catalyst contact under diesel combustion
278
conditions, and therefore appear to be more effective than the cerium oxide catalyst as observed
279
in this study.
280
The addition of FBCs to diesel fuel could also considerably reduce the ignition 12, 13, 33
281
temperature (light-off temperature) of diesel soot as proposed by some previous studies,
282
which was further confirmed by our current study (see a later section).
283
temperature, the oxidation rate of diesel and soot in the last stage of engine combustion will
284
decrease leaving more unburned fuels left to condense on the surface of the soot. Therefore, the
285
reduction of ignition temperature may be one of the factors that led to higher OC emissions.
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Finally, the lower EC emissions may provide less surface area for OC adsorption, which may in
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turn lead to lower OC emission. The two factors compete with each other, and therefore lead to a
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minor variation of OC caused by FBCs-doped fuels.
With a drop in
289
Particle-phase PAHs and n-alkanes emissions. Compared to the undoped ULSD, the
290
total particle-phase PAHs concentrations generally increased with an increase in the Ce and Fe
291
contents in the fuels (Figure 2(a)). The percentage increase in both MMW-PAHs and HMW-
292
PAHs was higher than that of LMW-PAHs. Another important finding is that with the same dose
293
of FBCs in the fuel, the Fe-doped fuel led to higher total particle-phase PAHs emissions than that
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of the Ce-doped fuel. A similar effect of both additives on the total BaPeq was also observed in
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this study, suggesting that both Ce- and Fe-doped fuels tend to increase the toxicity of the DPM
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due to an increase in the particle-phase PAHs emissions. Our observation is consistent with a
297
previous report that the total particle-phase BaPeq was increased by CeO2-doped diesel fuels.8
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Figure 2(b) shows that for each fuel, the distribution of particle-phase n-alkanes represents a
299
similar bell-shaped curve with C21 as the most abundant. As in the case of FBCs, both Ce- and
300
Fe-doped fuels led to higher n-alkanes emissions compared to the undoped ULSD, with Fe-
301
doped fuels leading to higher emissions of n-alkanes than that of Ce-doped fuels. It is known that
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particle-phase PAHs and n-alkanes mainly originate from the unburned diesel fuel or/and
303
lubricating oil. Diesel fuels have a complex mixture of hydrocarbons with chain lengths mostly
304
between C12 and C26 while crankcase oils tend to have a longer chain of carbon atoms. Given
305
the nature of diesel fuel and lubricating oil combined with the distribution of particle-phase n-
306
alkanes observed in this study, it is reasonable to assume that most of the n-alkanes observed in
307
this study are a result of incomplete combustion of diesel fuel and not lubricating oil. The same
308
reason may also be responsible for higher PAHs emissions. This finding further confirms our
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previously proposed theory that a reduction in the engine ignition temperature due to FBCs can
310
cause an increase in unburned hydrocarbon emissions as discussed in the previous section.
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Compared to Ce-doped fuels, Fe-doped fuels resulted in a lower ignition temperature and
312
therefore higher particle-phase PAHs and n-alkanes emissions. Moreover, we found that the
313
relative increase in C16-C25, caused by FBCs-fuels, was much higher than that of both C14-C15
314
and C26. This can be explained on the rationale that the heavier n-alkanes such as C26 probably
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originated from the lubricating oil, and therefore the emissions of these species are expected to
316
show little variations. As for the shorter-chain alkanes such as C14-C16, on the one hand, they
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are more likely to undergo nearly complete combustion than the heavier ones. On the other hand,
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they are not easily attached to the soot particles due to their higher volatility than heavier n-
319
alkanes. As a result, they were only slightly affected by the addition of FBCs. The same reason
320
can also be used to explain the observation that the percentage increase in both MMW-PAHs and
321
HMW-PAHs was higher than that of LMW-PAHs.
322
Particle number concentrations and size distributions. As shown in Figures 3(a)-(f),
323
the variation of both volatile and solid particle number size distributions, caused by the FBCs-
324
doped fuels, seems to be associated with their proportion used in ULSD and the engine load. At
325
the low engine load (Figure 3(a)), both Ce- and Fe-doped fuels resulted in the formation of new
326
particles with the nucleation mode within the peak diameters of 10-20 nm. Progressive increases
327
in the concentrations of Ce and Fe in fuels resulted in the size distribution of these nucleation
328
mode particles being shifted upward and toward a larger size, leading to an increase in the
329
emission of total particle counts as shown in Table S2. Moreover, for each dose of FBCs in
330
ULSD, the peak of nucleation mode particles for the Fe-doped fuels was higher and shifted
331
toward a larger size than that of the peak associated with the Ce-doped fuels, resulting in the Fe-
332
doped fuels having higher total particle counts than those of Ce-doped fuels, as evident from
333
Table S2. At the medium engine load (Figure 3(b)), with the exception of 25 ppm Ce, all other
334
FBCs-doped fuels showed obvious peaks in the nucleation mode. However, at the high engine
335
load (Figure 3(c)), only the 100 ppm Fe-doped fuel showed a peak in the nucleation mode. In
336
addition, with an increase in the engine load, the peak in the distribution of nucleation mode
337
particles for FBCs-doped fuels became flatter, indicating that the counts of nucleation mode
338
particles decreased with an increase in the engine load. Table S2 reveals that for each engine
339
load, with an increase of Ce and Fe concentrations in ULSD, the concentrations of particles with
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340
diameter less than 20 nm increased significantly, while the concentrations of particles larger than
341
50 nm decreased generally. Table S2 also shows that the FBCs-doped fuels significantly
342
increased particles of less than 50 nm in some cases, including at the low engine load with all
343
tested additions of FBCs, with the exception of 25 ppm Ce at the medium engine load, and also
344
at the high engine load with 100 ppm Fe. Our findings indicate that the increase in the total
345
particle counts as well as the decrease in the geometric mean diameter (GMD) caused by FBCs-
346
doped fuels was a result of a significant increase in the counts of smaller particles, while the
347
reductions in large diameter particles were mainly responsible for the decrease in particulate
348
mass, as evident from Figure 1(a). Additionally, our observations also reveal that with the same
349
amount of FBCs doped in the diesel fuel, the increase in the counts of nanoparticles is higher at
350
the lower engine load, and also generally higher for the Fe-doped fuel than compared to the Ce-
351
doped fuel, suggesting the variations in particle counts are associated with the FBC types used
352
and the engine operating conditions. Another interesting finding from Figures 3(d)-(f) is that
353
after TD, the counts of nanoparticles emitted from both Ce- and Fe-doped fuels were
354
significantly reduced due to the suppression of the formation of new particles from volatile
355
species after the dilution and cooling processes, but did not disappear altogether, which were still
356
much higher than those from pure ULSD. Our findings, therefore, confirm that more and larger
357
self-nucleated metallic particles and their agglomerates are generated in the cylinder and emitted
358
to the engine exhaust with an increase in Ce and Fe concentrations in ULSD. The explanations
359
regarding the formation and the evolutionary processes of metallic nanoparticles, and their
360
influence of particle size distributions in response to changes in the proportion of FBCs used in
361
ULSD and the engine load are provided in the SI. The presence of nanoparticles containing Ce
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362
and Fe may have profound implications on human exposure and subsequent health effects, and
363
this topic is discussed in a later section.
364
Soot oxidation behavior and soot nanostructure. Compared to undoped ULSD, a
365
lower soot ignition temperature was obtained from all Ce- and Fe-doped fuels (Figure 4(a)). The
366
extent of the reduction in the ignition temperature seems to have increased with an increase in
367
the Ce concentrations in fuels. However, the reduction in the ignition temperature showed no
368
clear trends with an increase in Fe concentrations. For each engine load, the activation energy of
369
soot generally decreased with an increase in the concentrations of both Ce and Fe in the fuels.
370
Our results suggest that the use of Ce- and Fe-doped fuels offers co-benefits in that these fuels
371
not only produce lower engine output i.e. soot, but also make it easier to oxidize soot by after-
372
treatment systems. In addition, at the high engine load, the soot emitted from Fe100 showed a
373
lower activation energy than that from Ce100, indicating that the soot emitted from Fe100 can be
374
more easily oxidized than that from Ce100 under high engine load. We believe that the Ce and
375
Fe embedded in the diesel soot act as catalysts that reduce the activation energy needed to start
376
soot oxidation and lower the temperature at which soot is oxidized. It should be noted that both
377
Ce- and Fe-based FBCs could effectively promote soot oxidation during the engine combustion
378
process as discussed previously. The reasons responsible for lower soot (EC) emissions can also
379
be used to explain the results in this section. Apart from that, previous observations also
380
suggested that the change in the volatile organic fraction (VOF) in particles,23 the nanostructure
381
of soot
382
oxidation of soot, which might vary with the Ce and Fe concentrations in the fuels as well as the
383
engine operation conditions. In this study, the higher OC fraction in particles and the higher
384
counts of total particles, caused by FBCs-doped fuels, suggest that the soot from these FBCs-
34
as well as the spreading of the metal oxide on the soot surface 33 affects the catalytic
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doped fuels could have higher surface areas than that of undoped ULSD, and thus should also be
386
considered as one of factors that led to higher soot oxidation reactivity.
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The D (Defect)/G (Graphite) band intensity ratios (ID/IG) were used to further explore if
388
and how FBCs-doped fuels affect the oxidation of soot through the change in the graphite-like
389
structure of soot particles. With the decrease in this ratio, the graphite-like structure becomes
390
dominant, implying the soot is not easily oxidized.33 Figure 4(b) shows that the ID/IG ratios of the
391
soot particles emitted from 25 and 50 ppm Ce-doped fuels were lower than that from undoped
392
ULSD, indicating that the soot had more ordered graphite-like structures and lower amorphous
393
carbon concentrations. However, the soot emitted from all Fe-doped fuels and from the 100 ppm
394
Ce-doped fuel showed a marginal change in ID/IG ratios, indicating the nanostructure of the soot
395
from these fuels was not significantly changed. The variation in soot nanostructures from the
396
combustion of FBCs-doped fuels in diesel engines could be associated with the changes in the
397
combustion chemistry and in-cylinder combustion and pyrolysis conditions, such as temperature
398
and residence time, etc. The results from this study indicate that the factors that lead to the
399
changes in soot nanostructure are related to the type and amounts of FBCs doped in fuels. Our
400
results further reveal that the changed soot nanostructure, caused by FBCs-doped fuels, did not
401
significantly affect the soot oxidation based on our observations that the soot from Ce25 and
402
Ce50 had more ordered graphite-like structures but still had higher oxidation activity than that
403
from undoped ULSD.
404
Toxicological Studies. Exposure of the cells to a dose of 25, 50 and 100 µg/ml Ce100-
405
derived particles and a dose of 25 and 50 µg/ml Fe100-derived particles led to lower cell
406
viability than those to ULSD-derived particles (Figure 5). Meanwhile, with the exception of the
407
highest exposure does of 200 µg/ml, the cell viability for particles derived from Ce100 was 18 ACS Paragon Plus Environment
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slightly lower than that from Fe100, indicating that the Ce-based additive leads to the particles
409
having a higher cytotoxicity than that of the Fe-based fuel additive.
410
As shown in Table 1, a short time exposure of A549 cells to the ULSD-derived particles
411
significantly modulated 115 genes, with 70 of them being up-regulated and the 45 down-
412
regulated. While the Ce100 and Fe100-derived particles triggered the modulation of 148 genes
413
(102 up- and 46 down-regulated) and 106 genes (79 up- and 27 down-regulated), respectively.
414
The strikingly altered genes in this assay were widely involved in transcription, cell
415
proliferation/cell cycle/cell aging, metabolism, response to stimulus, cell adhesion, cell signal
416
transduction, response to organic substances/drugs and transport (Table S3). Specifically, we
417
observed that the particles emitted from Ce100 caused the down-regulation of genes of CTGF,
418
CDKN2D, OSGIN1, CYR61, DDIT3, UHMK1 and FOS while the particles emitted from Fe100
419
caused the down-regulation of genes of CTGF, OSGIN1, CYR61. Additionally, when compared
420
to undoped ULSD, the particles emitted from Fe100 significantly up-regulated gene DDIT3. The
421
regulation of these genes is involved in the function of cell proliferation inhibition and/or
422
apoptosis promotion.35
423
Interestingly, compared to the control, the particles emitted from both Ce100 and Fe100
424
up-regulated the cytochrome P450 family genes (CYP1A1 and CYP3A5), which can be altered
425
by PAHs through the aryl hydrocarbon receptor.36-39 Additionally, the increased emissions of
426
PAHs were also reported to be related to the modulation of the genes involved in signal
427
transduction, apoptosis, metabolic activation process and DNA damage.38 Apart from PAHs,
428
alkanes were shown to up-regulate several genes related to cell cycle.37 Therefore, it is possible
429
that the higher emissions of particle-phase PAHs and n-alkanes, caused by FBCs-doped fuels,
430
may contribute to the variation of gene expressions as observed in this study. Notably, when 19 ACS Paragon Plus Environment
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431
compared to the control, the particles emitted from Fe100 specifically induced an up-regulation
432
of genes related to the response to reactive oxygen species (ROS)/oxidative stress (ACOX2,
433
ALDH1A3, CYP1A1, DHRS9, HMOX2, FA2H and PPARGC1A). Moreover, among these up-
434
regulated genes, ACOX2 is reported as a shared genetic risk factor for preeclampsia and
435
cardiovascular diseases,
436
addition, the cytochrome P450 family genes together with ALDH1A3 are more potent in the
437
induction of many pro-carcinogenic genes (i.e. JUN, FOS).38 Here, we noticed a significant up-
438
regulation of P450 family genes, ALDH1A3 and their corresponding pro-carcinogenic gene FOS
439
was stimulated by the particles emitted from Fe100. These results suggest that Fe100 may
440
potentially increase the pathogenicity of the diesel exhaust.
40
and CYP1A1 is associated with the increased lung cancer risk. 39 In
441
Interestingly, when we compared to undoped ULSD, we found that the particles emitted
442
from Fe100 induced a significant up-regulation of DDIT3, EGR1, EGR2, FOS, which were
443
associated with multiple biological functions including transcription, cell proliferation, metabolic,
444
and response to ROS. The particles emitted from Ce100 only significantly up-regulated CFB and
445
SERPINA3, which were involved in the regulation of immune and inflammatory responses.41
446
These results are consistent with previous reports in the literature that the exposure of cells to
447
Fe2O3 nanoparticles through the inhalation route could affect biological targets through
448
generation of ROS by oxygenation of reduced Fe species, 16, 42 and a short time exposure of cells
449
to CeO2 nanoparticles may lead to an acute inflammatory response.17 Therefore, we believe that
450
emission of the metallic nanoparticles from FBC-doped diesel as found in this study may affect
451
the gene expression and cytotoxicity.
452
Apart from the genes with specific functions described above, our data also reveals a
453
broad range of biochemical pathways that are illustrated by genes regulating transcription, 20 ACS Paragon Plus Environment
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transport and cell signal transduction. This observation implies that apart from PAHs and metal
455
oxides, the changes in polar and nonpolar organic compounds in the OC fraction, the physical
456
properties of metallic nanoparticles, caused by FBCs-doped fuels, could also affect the biological
457
reactions occurring at the particles/cell interface and therefore may lead to synergistic or
458
antagonistic biological effects. Notably, the results from this preliminary study suggest that a
459
panel of cytochrome P450 family genes, and the genes that are related to oxidative stress and
460
inflammatory response proposed here deserve considerable attention when conducting further
461
studies on this topic.
462
Environmental and health implications. The study reveals that the use of FBCs-doped
463
fuels in diesel engines effectively reduces the DPM mass and EC emissions due to the enhanced
464
soot oxidation, caused by FBCs during engine combustion. The reduction in DPM mass tends to
465
improve urban air quality while that in EC is beneficial from the climate change mitigation
466
viewpoint as EC is a second major contributor to global warming after CO2. However, the FBCs-
467
doped fuels facilitate the formation of self-nucleated metallic nanoparticles, resulting in the
468
increased emissions of nanosize particles. In addition, the use of FBCs-doped fuels increases the
469
organic carbon (OC) fraction of particles and the water-soluble fraction of OC, suggesting an
470
increased fraction of both polar and non-polar organic compounds in particles, including the
471
specific toxic organic compounds such as PAHs. Another important observation is a decrease in
472
cell viability, as revealed by the in vitro toxicity assessment. Exposure of human epithelial cells
473
to DPM exhibited alterations in the global gene expression with a broad range of biochemical
474
pathways, which could be attributed to the changes in the physico-chemical characteristics of
475
DPM caused by the FBCs-doped fuels. Emission of ultrafine particles (diameter < 100 nm) with
476
high toxicity is of major health concern as such particles penetrate deeper in the lungs and can
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477
cross cellular membranes, extending their toxicological impacts on other organs.43 From the
478
findings of the study, it becomes clear that the DPM mitigation strategies involving the use of
479
chemical additives in diesel engines must take into account the different characteristics of DPM
480
holistically in order to provide both environmental and public health benefits.
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481
Limitations. The knowledge gained from this preliminary investigation with a small
482
diesel engine (gen-set) can be valuable for selecting appropriate fuel additives or designing after-
483
treatment devices and alternative combustion systems for reduction of DPM emission from
484
diesel engines. However, further studies will still be needed to investigate detailed new particle
485
formation mechanisms in large commercial diesel engines as well as in on-road engines and to
486
examine the influence of fuel properties and operating conditions on the detailed characteristics
487
of DPM while using toxic metal-containing chemical additives. Apart from PAHs and n-alkanes,
488
other toxic organic compounds such as nitrated- and oxygenated-PAHs, hopanes, steranes, and
489
quinones, should also be determined while using FBCs and their role in the toxicity of DPM
490
should be assessed. Expression profiles of genes induced by DPM in human epithelial cells
491
provided a fundamental understanding of possible biological mechanisms involved in pulmonary
492
diseases, caused by FBCs-doped fuels. The finding from the cDNA microarray technique needs
493
to be confirmed with additional analyses of gene and protein expressions in other conditions such
494
as varied exposure time and dose, and different human pulmonary cell lines using a
495
complementary technique such as quantitative real-time PCR (qPCR). Additionally, particles
496
emitted from different engine types and operating conditions will allow us to gain a better
497
understanding of how the changes in the physico-chemical characteristics of DPM, induced by
498
FBCs, trigger the multiple biological effects. It should be acknowledged that the protocols used
499
in the current toxicity study could significantly change the nature of particles emitted from the
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500
engine, which might not be representative of in vivo exposure. Therefore, in vivo inhalation
501
exposure studies using animal models over a long time of exposure are needed. Despite these
502
limitations, the findings from this study emphasize the importance of a balance that needs to be
503
maintained between the environmental regulation-driven strategies involving the use of FBCs to
504
reduce the emission of the DPM mass and the key environmental and public health benefits that
505
can be derived, leading to a win-win situation.
506
Supporting Information available: The schematic of the experimental system employed in this
507
study, specifications of the engine, the procedures of doping CeO2 and Fe(C5H5)2 to diesel fuel,
508
the methods of determination of the dilution ratio and the handling of filters and DPM samples,
509
the procedures of physical analyses and toxicological experiments, and the details of quality
510
assurance and control (QA/QC) protocols for chemical analyses. The discussion regarding the
511
DPM formation processes, the explanations regarding the formation and the evolutionary
512
processes of metallic nanoparticles and their influence of particle size distributions in response to
513
changes in the proportion of FBCs used in ULSD and the engine load.
514
ACKNOWLEDGMENTS
515
Z.-H. Zhang thanks the Singapore − Peking − Oxford Research Enterprise (COY-15-EWI-
516
RCFSA/N197-1) for providing scholarship in support of his doctoral study.
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(41) Kelz, M. B.; Dent, G. W.; Therianos, S.; Marciano, P. G.; Mclntosh, T. K.; Coleman, P. D.; Eberwine, J. H. Single-cell antisense RNA amplification and microarray analysis as a tool for studying neurological degeneration and restoration. Sci. Aging Knowl. Environ. 2002, 1, re1.
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(42) Acevedo-Morantes, C. Y.; Meléndez, E.; Singh, S. P.; Ramírez-Vick, J. E. Cytotoxicity and reactive oxygen species generated by ferrocenium and ferrocene on MCF7 and MCF10A cell lines. J. Cancer Sci. Ther. 2012, 4, 271-275.
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637 638 639 640
Environmental Science & Technology
(43) Oberdörster, G.; Maynard, A.; Donaldson, K.; Castranova, V.; Fitzpatrick, J.; Ausman, K.; Carter, J.; Karn, B.; Kreyling, W.; Lai, D.; et al. Principles for characterizing the potential human health effects from exposure to nanomaterials: Elements of a screening strategy. Part. Fibre Toxicol. 2005, 2, 8.
641 642 643 644 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661 662 663 664 665 666 667 668 669
27 ACS Paragon Plus Environment
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Page 28 of 35
Table 1 Functional classification of genes up- and down-regulated following exposure of A549 cells after 6 h to particles derived from both FBCs-doped and undoped ULSD fuels
Category Transcription Cell proliferation /growth/cycle/aging Metabolism Stimulus Cell adhesion Signal transduction Response to organic substance/inorganic substance/drug Response to reactive oxygen species/oxidative stress Regulation of inflammatory response Transport Total
ULSD vs Control
Ce100 vs Control
Fe100 vs Control Downregulated 0 3
Ce100 vs ULSD Upregulated 0 0
Fe100 vs ULSD Upregulated 5 2
Upregulated 8 10
Downregulated 13 0
Upregulated 0 15
Downregulated 11 10
Upregulated 3 13
13 8 4 9 10
15 11 2 0 11
15 10 0 5 15
5 8 2 0 0
17 8 0 19 5
6 4 2 0 5
0 0 0 0 0
4 0 0 0 0
0
6
0
0
7
0
0
2
3
0
5
0
3
0
2
0
9 70
2 45
9 102
0 46
5 79
2 27
0 17
0 7
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Environmental Science & Technology
Figure Captions Figure 1. Effect of FBCs-doped fuels on (a) the concentrations of diesel particulate matter (DPM) mass and carbonaceous species, (b) EC (elemental carbon), (c) OC (organic carbon), and (e) WSOC (water-soluble organic carbon)), and the ratios of (d) OC/EC and (f) WSOC/OC. Significantly different from the ULSD (p < 0.05).
b
a
Significantly different from between Ce-
doped fuel and Fe-doped fuel under the same doping dose (p < 0.05). Figure 2. Effect of FBCs-doped fuels on (a) the concentrations of particle-phase PAHs and their total BaPeq (benzo[a]pyrene equivalent), and (b) the concentration distributions of n-alkanes. LMW (low molecular weight, 2 and 3 rings)-PAHs, MMW (medium molecular weight, 4 rings)PAHs, HMW (high molecular weight, 5 and 6 rings)-PAHs. a Significantly different from the ULSD (p < 0.05). b Significantly different from between Ce-doped fuel and Fe-doped fuel under the same doping dose (p < 0.05). Figure 3. Effect of FBCs-doped fuels on (a-c) volatile (without TD) and (d-f) non-volatile (with TD) particle number size distributions. Figure 4. Effect of FBCs-doped fuels on (a) soot oxidation: ignition temperature and activation energy, and (b) the D (defect)/G (graphite) band intensity ratios (ID/IG). a Significantly different from the ULSD (p < 0.05).
b
Significantly different from between Ce-doped fuel and Fe-doped
fuel under the same doping dose (p < 0.05). Figure 5. Effect of particles derived from both ULSD and FBCs-doped fuels on cell viability. Significantly different from the ULSD (p < 0.05).
b
a
Significantly different from between Ce-
doped fuel and Fe-doped fuel under the same doping dose (p < 0.05).
29 ACS Paragon Plus Environment
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(b) 50
(a) 120 Particulate mass
EC
75% Load
75% Load
3
DPM mass concentration (mg/m )
Page 30 of 35
25% Load
50% Load a a
3
80
40 a,b a,b a,b a,b
EC (mg/m )
100
60
a a
a,b a,b a,b a,b
40 a a a,b a,b a,b a,b
50% Load a
30
a a
50% Load
75% Load
75% Load
a,b
a b
OC/EC
3
OC (mg/m )
50% Load
25% Load
a,b
30 a,b
b
30 a,b a,b a,b
20
a a
a,b a,b a,b a,b a a
10
10
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
(f) 0.30
WSOC 50% Load
a,b a,b a,b a,b a,b a,b
Fuel type
(e) 10 25% Load
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
0
Fuel type
75% Load
0.25
WSOC/OC
25% Load
a
4
WSOC/OC
6 a
a a
50% Load
a
0.20
3
WSOC (mg/m )
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
OC/EC
25% Load
40
8
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
(d) 50
OC
50
0
a a a,b a,b
Fuel type
40
20
75% Load
a,b a b
0.15 a,b a,b
0.10
a,b a,b
a a
2
a a
0.05
Fuel Type
Fuel type
Figure 1 30 ACS Paragon Plus Environment
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
0.00
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
0
a,b a,b
a a a a a,b a,b
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
0
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
60
a
20
Fuel type
(c) 70
a a
10
20 0
25% Load
Page 31 of 35
Environmental Science & Technology
1.5 50% Load
Total BaPeq HMW-PAHs MMW-PAHs LMW-PAHs
50
a,b a,b
a,b
40
a,b
a,b
a,b a,b a,b
a,b
a,b
0.6
a,b
a,b
20
0.9
a,b a,b
b
30
a,b
10
a,b
a,b
a
a
a,b
Fe100
Ce100
Fe50
Ce50
Fe25
Ce25
ULSD
0
1.2
a,b
0.3
0.0
Fuel type
50% Load
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
3
Particle-phase n-alkanes (µg/m )
(b) 500 400
300
200
100
0 14
16
18
20
22
Number of carbon atoms Figure 2 31 ACS Paragon Plus Environment
24
26
Total BaPeq
3
Particle-phase PAHs (µg/m )
(a) 60
Environmental Science & Technology
(a) 4 x 1014
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
14
2 x 10
50% Load, without TD
14
3
3
3 x 10
dN/dlog(Dp) (#/m )
14
dN/dlog(Dp) (#/m )
(b) 4 x 1014
25% Load, without TD
14
1 x 10
3 x 10
14
2 x 10
14
10
0
100
10
Diameter Dp (nm)
75% Load, without TD
14
2 X 10
25% Load, with TD
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
14
3
3 X 10
0
(f) 4 X 1014
50% Load, with TD
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
14
14
3
3
10
100 Diameter Dp (nm)
dN/dlog(Dp) (#/m )
10
dN/dlog(Dp) (#/m )
14
2 x 10
1 x 10
0
1 X 10
14
3 x 10
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
14
14
1 X 10
(e) 4 X 1014
100 Diameter Dp (nm)
(d) 4 x 1014
dN/dlog(Dp) (#/m )
3
dN/dlog(Dp) (#/m )
(c) 4 X 1014
2 X 10
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
1 x 10
0
3 X 10
Page 32 of 35
14
3 X 10
100 Diameter Dp (nm)
75% Load, with TD
14
14
2 X 10
14
1 X 10
0
0
10
100 Diameter Dp (nm)
10
Figure 3
32 ACS Paragon Plus Environment
100 Diameter Dp (nm)
ULSD Ce25 Ce50 Ce100 Fe25 Fe50 Fe100
Environmental Science & Technology
25% Load
640
250 a,b
a,b
480
a,b a,b a,ba,b a,b
a,b a,b a a
a a
a a
a,b
a,b
a,b
a,b
a,b
a,b a,b a a
a a
320
300
75% Load
50% Load
Ignition temperature Activation energy
o
Ignition temperature ( C)
(a) 800
a a a a
a a
a a a b a b
160
150 100 50
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
Fe100 Ce100 Fe50 Ce50 Fe25 Ce25 ULSD
0
200
0
Fuel type
(b) 2.0 ID/IG
50% Load b a,b
1.5
ID/IG
b a,b
1.0
0.5
Figure 4
33 ACS Paragon Plus Environment
Fe100
Fuel type
Ce100
Fe50
Ce50
Fe25
Ce25
ULSD
0.0
Activation energy (kJ/mol)
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Environmental Science & Technology
670 671 ULSD 672 Ce100673 Fe100674
100
90 Cell viability (% control)
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a,b a,b
80
a,b a,b
b a,b
70
60
50 25
50 100 Particulate dose (µg/ml)
Figure 5
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200
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Environmental Science & Technology
229x94mm (150 x 150 DPI)
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