Article pubs.acs.org/est
ATR-IR Study of Ozone Initiated Heterogeneous Oxidation of Squalene in an Indoor Environment Dong Fu,†,# Chunbo Leng,†,‡,# Judas Kelley,† Guang Zeng,†,‡ Yunhong Zhang,‡ and Yong Liu*,† †
Department of Chemistry, University of Colorado Denver, Denver, Colorado 80217, United States The Institute of Chemical Physics, Key Laboratory of Cluster Science, School of Chemistry, Beijing Institute of Technology, Beijing 100081, China
‡
ABSTRACT: There has been a surge of interest in interfacial ozone chemistry for its application in indoor air quality and public health. Squalene, one of the most abundant ozone reactive constituents in an indoor environment, has received increasing attention lately, and a number of studies have been devoted to its heterogeneous interaction with ozone in actual and simulated settings. At present, there is still a large discrepancy in the measurement of the reactive uptake coefficient of ozone onto a squalene surface, and knowledge about this system remains incomplete. In this work, we investigated the ozone initiated heterogeneous oxidation of squalene using attenuated total reflection infrared spectroscopy (ATR-IR). We measured pseudo-first-order rate constants and uptake coefficients based on time dependent absorbance changes in CC (1668 cm−1) and CO (1730 cm−1) vibration bands. The uptake coefficients are (1.7 ± 0.2) × 10−4 from the CC band and (5.1 ± 0.7) × 10−4 from the CO band. The latter is likely an upper limit of reaction probability for ozone uptake onto squalene. Studies of temperature (5−32 °C) and relative humidity (0 and 80% RH) dependence revealed that indoor temperatures and RHs did not affect reaction kinetics. The insignificant RH effect is probably due to the weak interaction between water and squalene molecules. We quantitatively characterized the hydrophilicity and redox activity of squalene before and after exposure to ozone for the first time, and observed considerable enhancements in both hydrophilicity and redox activity during reaction. This may imply that ozone initiated heterogeneous oxidation could pose a higher public health risk in an indoor environment, and it may help explain some of the adverse health effects associated with elevated indoor pollutants.
1. INTRODUCTION
Squalene, a major component of lipid found in human skin, has been reported as one of the most abundant ozone reactive constituents in indoor environments.16 Over the past five years, heterogeneous reaction of ozone with squalene has been examined in both simulated and actual indoor settings,2,4,5,7,9,11,16 and people have started to recognize the pivotal role that squalene plays in indoor ozone chemistry as the most important ozone scavenger.16 As revealed in these studies,2,4,5,7,9,11,16 squalene can readily react with ozone, and the reaction products include mono- and bifunctional group compounds such as aldehydes, ketones, and carboxylic acids. As of today, most studies have centered on reaction product identification, and only two kinetics studies have been conducted by Wells et al.7 and Petrick and Dubowski.11 Unfortunately, reaction probabilities of ozone uptake onto squalene in these two studies differ by a factor of 45. Moreover, at present there is only one study of the effects of environmental conditions such as temperature and relative humidity on this important chemistry.11 It is evident that there is still a large discrepancy in the measurement of the reactive
Atmosphere is a complex, dynamic gaseous system, and air pollution exists in both outdoor and indoor environments. People in industrialized countries, on average, spend 80−90% of their time indoors where concentrations of some pollutants are often much greater than those outdoors.1 Lately, research in ozone interaction with myriads of organic compounds at indoor interfaces has garnered increasing attention,2−19 for many pervasive household chemicals such as detergents, disinfectants, and air-refreshers, once applied, they can be adhered to various indoor surfaces for an extended period of time and interact with ozone. The indoor interfacial chemistry is markedly important because compared to outdoor settings, the available surface area relative to the building volume is extremely large, and their heterogeneous reaction probabilities can be greatly enhanced by several orders of magnitude relative to the counterparts in gas phase.20 Also, products of the chemistry are often highly complex and include carcinogens, irritants, free radicals, secondary organic aerosols, and other oxidation products, all of which can contribute to the poor air quality in homes.20 Epidemiological evidence has demonstrated that exposures to elevated indoor pollutants can cause hypersensitive pneumonitis, asthma, legionellosis, and sick building syndrome (SBS).21,22 © 2013 American Chemical Society
Received: Revised: Accepted: Published: 10611
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O3/H2O/air, in which air was the carrier gas. Two mass controllers installed upstream were used to control flow rates of the gas mixture. An ozone generator (Pen-Ray) was used to generate ozone by flowing dry air at 200 mL/min through a UV light source, and then the ozone flow was diluted by a second flow of dry air of 800 mL/min. The flow system has an air change rate of 200 min−1. The ozone concentration measured downstream by an ozone monitor (UV-100, Ozone solutions) was 250 ± 15 ppb. In a similar ATR-IR study, Petrick and Dubowski11 investigated the heterogeneous reaction of ozone and squalene over a wide range of ozone concentrations (40 ppb to ∼20 ppm) and observed no secondary oxidation reactions when the ozone level was ≤250 ppb. Humidification in the reactor was achieved by switching the dry airflow to a path through a dew point generator (V-gen, Instruquest), and RH (80 ± 1%) was controlled by adjusting the dew point temperature. The setup allowed O3 and H2O concentrations to be varied independently. A circulation bath was used to achieve reaction temperature control by circulating a heat transfer fluid through stainless steel cooling blocks which were covered by silicone for insulation. Two thermocouples were placed inside the inlet and outlet of the reactor. Averaged readings from two thermocouples were used to determine the reaction temperatures. Since it usually took several minutes to establish an ozone flow with a stable concentration, an additional ozone bypass line was installed to minimize uncertainty resulting from the changes in ozone concentration. Ozone flow was kept on throughout the experiments and was switched back and forth between bypass line and reaction line through the flow reactor. Before each reaction, the reacting gas flow was switched to the bypass line, and a background spectrum of the clean ZnSe crystal was taken. Squalene thin film was prepared by dispensing a 60 μL solution of 4 mM squalene (99% purity, Aldrich) in carbon tetrachloride (99% purity, Aldrich) directly onto the ATR crystal using a micropipet. The amount of solution was sufficient to completely cover the ATR crystal surface. The flow reactor was then sealed, and additional N2 flow was directed through the reactor to evaporate the solvent. Removal of the solvent was confirmed by the disappearance of the C−Cl band in the IR spectra. The thickness of the thin film was estimated to be ∼0.25 μm, much less than the penetration depth of the evanescent wave so that the bulk of the film could be fully probed by the IR beam. Infrared spectral collections were started once the ozone flow was switched to the reaction line. In all experiments, spectroscopic measurements were carried out simultaneously with the exposure, and spectra were acquired automatically using the macro function built in the OMNIC program over 32 scans at a resolution of 4 cm−1. For each reaction condition, experiments were repeated at least 3 times. The redox activity of samples was measured by DTT assay, and details about the method have been discussed elsewhere.33 In brief, the squalene samples were extracted by sonicating in 10 mL of acetonitrile for 30 min. Afterward, 50 μL aliquots of the extracts were taken into 1 mL of potassium phosphate buffer at pH 7.4 with 200 μM DTT (in excess). The solution was incubated at 37 °C for different amounts of time (10−60 min). At the designated times, 1 mL of 10% trichloroacetic acid was added to the mixture, followed by 4 mL of 0.4 M Tris-HCl at pH 8.9 containing 20 mM EDTA. 100 μL of 10 mM DTNB was added to react with the remaining DTT to generate 5mercapto-2-nitrobenzoic acid, and its concentration was determined by absorption at 412 nm from a UV−vis
uptake coefficient of ozone onto a squalene surface, and our knowledge about this key indoor reaction remains limited and incomplete. Redox activity of chemicals can induce a sequence of redox reactions in living organisms, which result in formation of reactive oxygen species (ROS). ROS in cells are typically generated through electron transfer from biological reducing agents such as NADH and NADPH to dissolved oxygen to produce O2−• and H2O2 with the catalytic assistance of electron transfer enzymes and redox active chemical species such as redox active metals and organics.23−25 Oxidative stress, which is often associated with an increase in ROS level, reflects an imbalance in oxidants and antioxidants in cells and has been regarded as a major toxicological mechanism for causing damage to cell structures.26 Squalene’s interaction with ozone has been known to generate products, which may be respiratory and skin irritants.16 However, to the best of our knowledge, no data is available about how the interfacial ozone oxidation quantitatively alters the redox activity of squalene. Additionally, recent studies on chemical aging of atmospheric organics have demonstrated an enhancement in the hydrophilicity of reaction products.27−29 Studies on penetration enhancement for skin care products and transdermal drugs have shown that adequate hydrophilicity and lipophilicity would facilitate penetration through the stratum corneum of skin and cell membrane.30,31 Similarly, hydrophilicity enhancement has been discussed in a recent review article as a simple and effective approach to improve cell/tissue compatibility with biomedical materials.32 Since squalene is lipophilic in nature; one may speculate that its interaction with ozone could result in a change in hydrophilicity, which would affect condensed phase reaction products’ ability to penetrate through the skin and cell membranes, in turn, facilitating the ROS formation in the cells and imposing an increased public health risk. Motivated by the large discrepancy in reaction probability of squalene/ozone and the lack of knowledge of change in redox activity and hydrophilicity of squalene upon exposure to ozone, we investigated the ozone initiated heterogeneous oxidation of squalene using an attenuated total reflection infrared spectroscopy (ATR-IR). The main objectives of this work are to 1) reduce uncertainty in the squalene/ozone reaction kinetics and provide recommended data and 2) characterize properties of squalene during interaction with ozone to offer more insights into the ozone interfacial chemistry and its potential health impact. In this work, we derived pseudo-first-order rate constants and uptake coefficients from absorbance changes in both CC and CO bands as a function of time at different temperatures (5−35 °C) and RHs (0 and 80%) related to an indoor environment. We also quantitatively determined hydrophilicity and redox activity of squalene before and after exposure to ozone for the first time using ATR-IR spectroscopy and dithiothreitol (DTT) assay.
2. EXPERIMENTAL SECTION Experimental setup is similar to the one used in our previous study.29 The heterogeneous reactions of squalene with ozone under different conditions were monitored in real time by an ATR-IR spectrometer (Nicolet 6700) equipped with a liquidnitrogen cooled MCT detector. A multireflection sampler equipped with a ZnSe ATR crystal (refractive index: 2.4) was placed into a customized flow-through stainless steel chamber (volume: ∼5 cm3). The penetration depth is estimated to be 1.1 μm at 1700 cm−1. Reacting gaseous species were made of 10612
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−C(CH3) wagging vibration shows peaks at 1381 cm−1. When the squalene thin film is exposed to ozone, the initial heterogeneous oxidation is expected to proceed via the ozonolysis mechanism.11 Figure 2 shows the spectral evolution of the squalene thin film during exposure to ozone over time at room temperature and under dry condition. As shown in the Figure 2, in the high wavenumber region bands associated with alkyl stretches decreased with increasing ozone exposure time. In the middle wavenumber region, in addition to the decrease in absorption at 1668 cm−1, once the reaction started, a new peak located at about 1730 cm−1 began to appear and increase accordingly as the reaction proceeded. The band was reportedly attributable to CO stretching vibration of ketones, aldehydes, and carboxylic acids.34 Along with the increase in absorption at 1730 cm−1, in the fingerprint region, additional bands including C−H deformation vibration near 1379 cm−1 and several C−O stretch absorption in the range of 1300−1000 cm−1 increased with increasing exposure times. All these spectral observations have demonstrated that squalene readily reacted with ozone, which was consistent with previous studies.2,4,5,7,9,11,16 Due to the complex spectral overlap in the high wavenumber and fingerprint regions, it was difficult to separate those peaks quantitatively. As a result, only the spectral changes in the C O band (1730 cm−1) and the CC bands (1668 cm−1) were utilized in this work to derive kinetics of the heterogeneous oxidative reaction of squalene. Although ozone molecules have absorption in mid-IR region,34 we did not observe any noticeable absorption from ozone molecules in our ATR-IR setup when we had ozone flow through the reactor. Thus, the contribution of ozone absorption to the infrared spectra can be neglected. 3.2. Rate Constants and Uptake Coefficients. The method for determining the reaction rate constants and reactive uptake coefficients was similar to the ones used in previous ozone oxidation studies, and details have been discussed elsewhere.11,29,35−37 Based on Beer’s law, absorbance of the CC band at ∼1668 cm−1 is linearly proportional to the concentration of squalene. Likewise, values of the CO absorption band at ∼1730 cm−1 are linearly proportional to the concentration of reaction products. In the present study, the ozone:squalene molar ratio was kept >10 throughout the reaction by careful selection of ozone concentration, flow rate, reaction time, and squalene amount to ensure the pseudo-firstorder condition was met. As a result, the absorbance values of both CC and CO bands are expected to follow an
spectrometer. The rate of DTT consumption was proportional to the concentration of the catalytically active redox-active species in the samples. Hydrophilicity of samples was determined by its interaction with water using the identical ATR-IR setup for heterogeneous reaction study. During the water uptake study, the ozone flow was turned off. Moist air was generated by the dew point generator, and RH was controlled by adjusting the dew point temperature.
3. RESULTS AND DISCUSSION 3.1. IR Spectra of Ozone-Processed Squalene Film. Squalene is a triterpene with 6 unconjugated double bonds. Figure 1 shows an infrared spectrum of squalene thin film at
Figure 1. Infrared spectrum of fresh squalene thin film at room temperature.
room temperature before exposure to ozone. Generally, main absorption features arise from −CH3 and −CH2 groups. As shown in the figure, there are peaks located at 3050, 2966, 2918, and 2853 cm−1, which can be attributed to CH stretch, −CH3 asymmetric stretch, −CH2 asymmetric stretch, and −CH2 symmetric stretch, respectively. The peak near 1668 cm−1 is assigned to the CC stretching vibration. The −CH2 bending vibration is observed at 1448 cm−1, while the
Figure 2. Infrared spectra of squalene thin film at different ozone exposure times during the reaction. Conditions: [O3] ∼ 250 ppb, room temperature, and RH ∼ 0%. The Y-axis stands for absorbance. 10613
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Figure 3. Temporal changes in the infrared spectra shown focusing on CO and CC spectral bands: (a) CO for ∼3 h; (b) CC for ∼3 h; (c) CO for ∼10 min; and (d) CC for ∼10 min. The absorbance data (solid circles) was exponentially fit (red solid lines) to derive the kapp.
kapp(CC)‑overall and kapp(CO)‑overall is relatively larger than what we have observed in the case of linoleic acid/ozone reaction.29 Such discrepancy likely originated from the highly unsaturated nature of squalene and complexity of the reaction products. The reaction between ozone and unsaturated hydrocarbon is expected to proceed via addition of ozone to the double bond to form an unstable primary ozonide, followed by a Criegee intermediate and aldehyde/ketone formation. The Criegee intermediate can further decompose or continue to react with other fragments, yielding bifunctional group products.11 There are six double bonds in a squalene molecule, and ozone molecules are expected to have no preference toward any of them based on electron withdrawing groups, stability of cations, or hindrance.11,38 Therefore, the gain rate of CO group may be faster than the loss rate of CC groups. In addition, as shown in Figures 3a and 3b, the oxidation took place very rapidly initially and started to level off after about 2 h. If the squalene is exposed to ozone for an extended period of time, as pointed out by Petrick and Dubowski,11 it is possible that many of the reaction products get involved in a secondary reaction with ozone or with the formed OH and alkyl radicals, forming secondary oxidation products such as acetaldehyde, glyoxal, and pyruvic acid. In our experiments, the reactions typically lasted about 3 h, and as a result, there may be some secondary reactions occurring toward the end of the reaction. The slight redshift of carbonyl group from 1730 to 1724 cm−1, as the reaction progressed, may be indicative of some secondary reactions. Given the potential complication from the secondary chemistry, we also analyzed the kinetics data only from the beginning of the reaction (approximately first 10 min) and the kapp(CC)‑initial and kapp(CO)‑initial are (2.8 ± 0.3) × 10−4 s−1 and (8.0 ± 0.8) × 10−4 s−1, respectively (Figures 4c and 4d). The kapp(CC)‑initial is slightly larger than kapp(CC)‑overall; this is probably because at the beginning of the reaction ozone was
exponential pattern with reaction time. Figures 3a and 4b present the time dependent absorbance profiles of CC (1668
Figure 4. The temperature dependence of kapp(CC)‑initial and γ(CC)‑.
cm−1) and CO (1730 cm−1) bands as the squalene thin film was exposed to ozone at ∼250 ppb at room temperature and under dry conditions as a function of time. The circles are absorbance data, and the solid lines are from exponential curvefitting. As seen in Figures 3a and 3b, the exponential function fits the time dependent absorbance profiles of both vibration bands markedly well, which confirms that the reaction in this study is under pseudo-first-order conditions. The apparent overall pseudo-first-order rate constants kapp(CC)‑overall and kapp(CO)‑overall acquired based on curve-fitting of the time dependent absorbance profiles of CC (1668 cm−1) and C O (1730 cm−1) bands are (2.5 ± 0.3) × 10−4 s−1 and (6.3 ± 0.6) × 10−4 s−1, respectively. Note that the difference in 10614
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to be insensitive to temperature variations, and no significant effect of temperature outside experimental margin of error was observed. This result is consistent with Petrick and Dubowski11 who measured uptake coefficients at 24 °C, 37 °C, and 58 °C and found no temperature dependence. According to Petrick and Dubowski’s study,11 the ozone and squalene heterogeneous reaction followed a Langmuir−Hinshelwood mechanism. Therefore, it likely involves two processes: ozone molecules are first adsorbed on the surface of the squalene thin film and quickly reach equilibrium with the gas phase; then the adsorbed ozone molecules react with the squalene on the surface at a relatively slower rate. In the present study, we are unable to determine whether squalene at subsurface layers participated in the reaction. Recent experiments have revealed that the overall reaction is more likely to be dominated by heterogeneous surface reaction over homogeneous bulk liquid reaction;40,41 therefore, the observed lack of temperature dependence may be due to low activation energy of surface-bound ozone reaction with squalene. Similar weak temperature dependence was also observed in other studies of ozone reaction with unsaturated organic liquid surfaces.42,43 Nevertheless, recent studies of ozone/oleic acid and ozone/linoleic acid reactions have revealed much stronger temperature dependent reaction kinetics when organic species are present in a liquid and solid mixture.29,36 Clearly, more studies on the effects of ambient temperature on ozone initiated heterogeneous reaction are needed for future work. The RH effect on kinetics was also studied at 0% and 80% with ozone concentration of ∼250 ppb and a temperature of 298 K. The kapp(CC)‑initial and the γ(CC)‑initial values obtained at 80% are (3.0 ± 0.4) × 10−4 s−1 and (1.8 ± 0.2) × 10−4, respectively. Our results show that differences in both the kapp and the γ obtained at RH = 0% and 80% are statistically insignificant. Similar RH effects were also reported by Dubowski and co-workers in their studies of heterogeneous oxidation of cypermethrin and squalene with ozone.11,37 However, our recent study of linoleic acid oxidation by ozone revealed water vapor can promote the reaction kinetics.29 Such disagreement may be explained by differing physiochemical properties of the organic species. Linoleic acid is a carboxylic acid, and it can absorb water much more readily than squalene both before and after ozone exposure.29 According to experimental study and molecular dynamic simulations, surface bound water can affect uptake of molecular species at surface layer.44 Since O3 molecules present an increased solubility in water (109 mL/L at 25 °C),45 nearly 13 times more soluble than O2, under high RH conditions, when O3 molecules collide with the linoleic acid film, some of them might be captured and dissolved into the surface water layers. This would augment residence time of surface adsorbed ozone and, in turn, enhance the heterogeneous reaction. On the other hand, although squalene thin film illustrates enhanced hydrophilicity after oxidation likely due to the formation of carbonyl containing moiety reaction products (see section 3.4), its ability to uptake water is still much weaker compared to linoleic acid. As a result, the reaction rate of squalene and ozone is not greatly affected by a higher humidity condition. 3.4. Hydrophilicity of Products. Chemical aging of organic surfaces may enhance their interaction with water.27−29 To understand how the ozone initiated oxidation affects hydrophilicity of squalene, three sets of water uptake experiments were carried out on unreacted and reacted squalene thin films using the identical ATR-IR setup for
more likely to interact exclusively with squalene. On the other hand, as the reaction progressed, the probability of ozone interaction with unsaturated moieties in reaction products became increasingly higher. As such, the kapp(CC)‑initial should be a better measure of reaction kinetics than kapp(CC)‑overall. Note the kapp(CO)‑initial is also slightly larger than the kapp(CO)‑overall, indicating that carbonyl group formation was probably more efficient at the early stage of the heterogeneous reaction. With the apparent first-order rate constants kapp, the uptake coefficient, γ, could be estimated by the following equation39 ⎛P c ⎞S d[Squalene] = −γ ⎜ O3 ̅ ⎟ A ⎝ 4RT ⎠ V dt
(1)
where d[Squalene]/dt = −kapp[Squalene], and [Squalene] [molecules cm−3] is the initial concentration of squalene (∼1.25 × 1021 molecules cm−3); c ̅ [cm s−1] is the mean speed of ozone molecules in gas phase; and SA/V [cm−1] is the surface area-to-volume ratio of the squalene thin film (∼3.91 × 104 cm−1). In the present study, the surface area of the squalene thin film was estimated to be 4.5 cm2, which is the available geometric surface area of the ZnSe crystal. Using the average kapp(CC)‑initial and kapp(CO)‑initial values, we obtained the γ(CC)‑initial and γ(CO)‑initial to be (1.7 ± 0.2) × 10−4 and (5.1 ± 0.7) × 10−4, respectively. Before our study, there were only two kinetics studies of ozone uptake onto squalene surfaces. Wells et al.7 first reported an ozone reactive uptake coefficient of (4.5 ± 1.4) × 10−4 by squalene thin film on a glass plate surface using the field and laboratory emission cell automation and control system. Petrick and Dubowski11 later employed an ATR-IR flow system and obtained a γ value of 1.0 × 10−5. Despite great similarities in the reaction system between Petrick and Dubowski’s11 and our studies, results obtained in this work are actually an order of magnitude higher than theirs. At this time, it is unclear what causes such a disagreement. It should be noted that experimental condition and data analysis method in both studies are fairly different. In this work, we selected amount of squalene, film thickness, ozone concentration, flow rate, and reaction time carefully to ensure sufficient infrared absorption, minimal secondary chemistry complication, and experiments in conformity with pseudo-first-order condition. In addition, to better measure reaction kinetics, we utilized time dependent absorbance data from CC and CO bands for both initial and overall stages. We also attempted to reanalyze Petrick and Dubowski’s data but were unsuccessful because the reaction time was not clearly conveyed in their paper.11 On the other hand, our γ values are much closer to Wells et al.,7 especially the γ(CO)‑initial. Albeit an excellent agreement between our γ(CO)‑initial and theirs,7 we believe the γ(CC)‑initial of (1.7 ± 0.17) × 10−4 should be more accurate and reliable considering the inherent complexity in the absorbance measurement from the CO band. The γ(CO)‑initial of (5.1 ± 0.7) × 10−4 is likely an upper limit for the reaction probability of ozone uptake by squalene. It should be pointed out that Wells et al. also stated that the (4.5 ± 1.4) × 10−4 observed in their study was among the highest measured values for ozone reaction with any indoor surface species.7 3.3. Temperature and RH Effects. The temperature dependent experiments were carried out under the condition of [O3] = ∼250 ppb, RH = ∼0%, and temperatures varying from 278 to 305 K. Figure 4 shows kapp and the corresponding γ versus temperature. As seen, both the kapp and the γ were found 10615
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squalene was exposed to ozone, even for 30 min, hydrophilicity of chemically transformed thin films was evidently enhanced. There was a considerable amount of water uptake even under lower RH conditions probably because of the formation of more oxygenated compounds with higher water uptake capacity. The amount of water adsorbed on the oxidized thin film at RH = 80% was more than double the squalene thin film without prior exposure to ozone. With an increase in exposure time from 30 min to 3 h, the hydrophilicity of the thin film was further improved, and the amount of water adsorbed on the oxidized thin film at RH = 80% was tripled. In spite of the appreciable enhancement in hydrophilicity of oxidized squalene thin film, when compared with linoleic acid,29 squalene still presented much weaker hydrophilicity. Squalene thin films before and after exposure ozone picked up only about 1/2 and 1/3 the amount of water at RH = 80% as compared to linoleic acid, indicating that the linoleic acid/ozone reaction yielded more hydrophilic products than those from the squalene/ozone reaction. This may help explain why no RH effect on reaction kinetics was observed. 3.5. Redox Activity of Products. DTT assay is a measure of redox activity of chemical samples related to their ability to induce a stress protein in cells. Rate of DTT loss is proportional to the concentration of the catalytically active redox-active species in the samples, and it can be acquired from the slope of time dependent absorbance of 5-mercapto-2-nitrobenzoic acid (Figure 6). As seen in Figure 6 (left), there is an excellent linearity of absorbance of 5-mercapto-2-nitrobenzoic acid as a function of incubation time in the analysis of squalene samples. From the curve-fitting, the DTT loss rate can be obtained. Figure 6 (right) presents consumption rates for different squalene samples after exposure to ozone at 250 ppb for various times (0, 30, 60, 90, 120, 150, and 180 min). Results illustrated that the DTT loss rates increased rapidly over 1 order of magnitude even after 30 min of exposure, indicating a significant enhancement in redox activity. As the reaction progressed, the continuous ozone exposure continued to increase the redox activity of reacted squalene in an almost linear fashion. After 90 min of exposure, the enhancement seemingly slowed down, and the DTT loss rate started to level off after 120 min. This indicated that the reaction may have approached completion after 120 min. The time dependent
heterogeneous reaction kinetics study. The only difference between the two studies was whether ozone flow was on or off. Using the same setup in both studies allowed us to stop the heterogeneous reaction at any stage and quickly switch to water uptake investigation. For the ozone treated squalene samples, reactions were conducted under two different conditions: T = 298 K, RH = 0%, 0.5 h and T = 298 K, RH = 0%, 3 h with [O3] ∼ 250 ppb. The method for the hydrophilicity experiments is similar to our previously reported ones.29,46,47 Absorbance of the OH stretching band in the 2750−3660 cm−1 region was employed to represent the adsorbed water content. Since other groups, such as alkyl stretching bands, also contribute to the absorbance intensity in the region of 2750−3660 cm−1, the spectrum recorded at RH = 0% for each film was subtracted from those recorded at higher RHs. Figure 5 shows the water
Figure 5. Changes in water absorbance of three different squalene thin films with RH.
absorbance values of the three different squalene thin films at different RHs. Before exposure to ozone, unlike linoleic acid, squalene is highly hydrophobic. Little water uptake was observed until RH became higher (70−80%). Once the
Figure 6. Absorbance of 5-mercapto-2-nitrobenzoic acid as a function of incubation time (left). DTT loss rate of squalene films exposed to ozone for different times (right). 10616
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support from National Natural Science Foundation of China (20933001). The authors are also very grateful to anonymous reviewers for their valuable comments and help.
DTT loss behavior observed here was consistent with the evolution of the infrared spectra (Figure 2), i.e, few spectral changes were observed after 120 min. Recently, McWhinney et al.48 observed a similar monotonic increase in the redox activity of ozone oxidized particles from a two-stroke engine. Our calculated DTT consumption rates are in reasonable agreement with theirs (∼5 × 10−7 M min−1) under similar exposure conditions (i.e, [O3] × exposure time). According to the literature,2,4,5,7,9,11,16 the ozone/squalene reaction yields a variety of oxygenated products including mono- and bifunctional compounds that contain carbonyl, carboxyl, and αhydroxyl ketone groups. These compounds probably have similar redox activity as the reaction products in McWhinney’s study.48 In the present study, ozone concentrations slightly higher than typical indoor environments (5−100 ppb) were used. The heterogeneous reaction of squalene/ozone follows a Langmuir−Hinshelwood mechanism,11 and the observed pseudo-first-order rate constant is nearly linearly dependent on ozone concentration up to 5 ppm. As a result, our exposure condition can also be represented by ∼750 ppb•h, which is approximately equivalent to one day’s exposure in a typical indoor environment. Squalene is prevalent in an indoor environment. Recent studies16,49,50 have indicated that products from the ozonolysis of squalene, such as dicarbonyl 4-oxopentanal and 3isopropenyl-6-oxo-heptanal, can induce irritation and allergic response upon exposure. Our results have shown that squalene can increase its redox activity by nearly 20-fold upon ozone exposure at ∼750 ppb•h. An enhanced redox activity in cells can damage cellular components such as proteins, lipids, and DNA. In addition, reactive oxidative species can also cause disruptions in normal cellular signaling mechanisms.36 For these reasons, oxidative stress, which is associated with an increased redox activity, has been attributed to development of a number of human diseases.51 In addition to the reported induced irritation and allergic response in previous studies,49,50 squalene ozonolysis products may cause other health problems related to oxidative stress. Furthermore, our study also reveals that heterogeneous oxidation can increase hydrophilicity of the condensed phase products. There are three major pathways of human exposure from pollutants: inhalation; dermal contact; and hand-to-mouth transfer. Since recent literature has demonstrated hydrophilicity can improve skin and cell membrane penetration of moisturizer and transdermal drugs,30,31 and cell and tissue compatibility of biomedical materials,32 it is conceivable that enhancements in hydrophilicity and redox activity from squalene upon ozone exposure could pose a higher health risk for human beings in an indoor environment. This may help explain some of the adverse health effects associated with elevated indoor pollutants.
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AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. Author Contributions #
Fu and Leng contributed equally to this work.
Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This work is supported by Research Corporation for Science Advancement (Grant # 20192). Y.H.Z. acknowledges the 10617
dx.doi.org/10.1021/es4019018 | Environ. Sci. Technol. 2013, 47, 10611−10618
Environmental Science & Technology
Article
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