Fiedler et al. cortisol was signiﬁcant (F = 4.90; df = 3, 347; p = 0.0024). The signiﬁcance of the main effect of stress was due to changes in cortisol levels from before the stressor (minute 90) to after the stressor (minute 125) (t = 4.00; p < 0.0001). When examining changes from minutes 110 to 125, on average, the cortisol levels decreased (from 0.178 to 0.154) for the no-stressor condition and increased slightly (from 0.140 to 0.147) for thestressor condition (t = 3.63; p = 0.0003) (Figure 4). Discussion The most striking result of our study was the lack of signiﬁcant subjective or objective health effects from exposure to mixtures of VOCs both with and without O3, despite signiﬁcant differences in the chemical composition of the air for the three conditions (Table 2). Numerous epidemiologic studies suggest that symptoms are significantly increased among workers in buildings with poor ventilation and mixtures of VOCs (Mendell 1993; Mendell et al. 2002; Sieber et al. 1996), many of which likely contain low levels of O3. Our present controlled acute exposure to similar chemical mixtures in young women, however, failed to support these epidemiologic ﬁndings. Relative to ambient air masked with a pulse of VOCs, neither the VOC nor the VOC + O3 exposures caused signiﬁcantly increased symptom reports, changes in cortisol, reduced neurobehavioral performance, or changes in lung function. Subjects were more likely to report some VOC physical symptoms in the VOC and VOC + O3 conditions relative to the MCA condition. However, these effects were of marginal signiﬁ- cance and became nonsigniﬁcant with appro- priate correction for multiple comparisons. Conversely, although stress did not exacerbate exposure effects, stress signiﬁcantly increased symptoms of anxiety. The effect of stress was further validated by the signiﬁcant difference in 10 E 1 S 8 ± ty 6 i r e v 4 e s an 2 e M0 15 60 Exposure begins 90 110 Stressor/math Minutes Figure 3. Mean anxiety symptom severity at each time point across exposures stress versus no stress. Bonferroni correction: p < 0.003. Time point compari- son for anxiety symptom sensitivity is as follows: change from minute 15 (baseline) to minute 110 (poststress), χ2 = 15.17 (p < 0.0001); change from minute 60 (after exposure onset) to minute 110 (post- stress), χ2 = 12.52 (p < 0.0004); change from minute 90 (prestress) to minute 110 (poststress), χ2 = 30.16 (p < 0.0001); change from minute 110 to minute 125, χ2 = 22.29 (p < 0.0001); change from minute 110 to minute 185 (poststress), χ2 = 11.64 (p < 0.0006). 1546 E0.5 S 1 )±0.4 L d /g0.3 µ ( l iso0.2 t r o c0.1 n a e M 0 cortisol for those who received the stressor relative to subjects who did not. There was a marginal (p < 0.08) increase in severity, but not incidence, of lower respiratory symptoms with the VOC and VOC + O3 exposure. However, there were no signiﬁcant changes in the lung function parameters attrib- utable either to the VOC or VOC + O3 expo- sure. This is consistent with work showing increased lower respiratory symptoms at 50 mg/m3 but not at 25 mg/m3 of VOCs alone, but no change in spirometry or increase in inﬂammatory mediators from induced spu- tum at either VOC concentration, again with- out O3 (Pappas et al. 2000). Although rodent bioassays had indicated that relatively higher concentrations of limonene–O3 oxidation products were irritating to the respiratory tract, at our exposure concentrations we did not see an effect on lung function and only a marginal effect on symptoms (Rohr et al. 2002; Wilkins et al. 2001). Indicators of nasal inﬂammation were also negative for our exposures (Laumbach et al., in press). In contrast to our present ﬁnd- ings, Kleno and Wolkoff (2004) reported increases in eye blink frequency among a small number of subjects with eye exposure only to limonene oxidation products and nitrate radi- cals relative to clean air. Although we did not measure eye blink frequency, symptoms of eye irritation were not signiﬁcantly greater in the VOC + O3 condition. Our present findings regarding neuro- behavioral performance are consistent with those of Otto et al. (1992), who reported no changes in neurobehavioral performance among subjects exposed to a similar mixture of 23 VOCs relative to clean air. However, the research group from the Technical University of Denmark reported several studies in which subjects were exposed to off-gassing from a 20-year-old carpet and showed reductions in productivity on tasks that simulate ofﬁce work (typing, calculations) (Wargocki et al. 1999, 2000a). Although the total VOCs were of similar concentrations between the exposure conditions with and without the carpet (~ 2.34 ppm), the composition of the chemical mixture differed substantially between the two conditions, and likely contained unidentiﬁed compounds associated with emissions from the No stress Stress Baseline Immediate poststress Prestress Poststress No stress Stress 125 185 Exposure ends carpet. In contrast to the negative ﬁndings in our present study, Wargocki et al. (1999, 2000a) also reported more symptoms (head- ache) and slower typing speed in response to the condition with the old carpet (polluted condition). Because the total VOC concentra- tion (2.35 ppm) was less than in our present study, symptom and performance differences could be ascribed to differences in the chemical composition, to unspecified biological com- ponents, or to differences in task demands. Neither acetone nor acetic acid was present in the VOC mixture used in our present study (only small amounts were produced from the O3 and VOC reactions), but such a specific chemical effect seems unlikely. Wargocki et al. (1999, 2000b) also suggested that simulated work tasks (typing, calculations) performed over a longer period of time (265 min) than in our present study were more sensitive to the effects of poor air quality than are standard neurobehavioral tasks. Thus, although the POL was given for approximately 20 min on two occasions during exposure, this did not require the sustained effort needed to perform continuous typing for 47 min on two occa- sions as in the Wargocki et al. (1999, 2000b) studies. However, the use of an old carpet as an exposure was ultimately not comparable with the present speciﬁc VOC mixture. Although some increased symptoms were observed in previous controlled exposures using similar “indoor air” mixtures (Hudnell et al. 1992; Prah et al. 1998), actually only a few out of many symptoms assessed in those studies were signiﬁcantly increased. A careful examina- tion of those symptoms exacerbated by expo- sure reveals some consistency with our present ﬁndings. Prah et al. (1998) reported that rela- tive to clean air, mixtures of VOCs increased ratings of nasal irritation, odor intensity, and air quality but not health (cough, sore throat) and cognitive symptoms (memory loss, dizziness). Similarly comparing responses of a VOC mix- ture with clean air, Hudnell et al. (1992) reported signiﬁcantly reduced air quality ratings and increased odor level, but they also reported increased symptoms of headache, eye irritation, drowsiness, and throat irritation. However, Hudnell et al. (1992) did not control for multi- ple statistical comparisons among the individual tests of 22 symptoms (p-value set at < 0.05). Furthermore, no previous indoor air study has “masked” the clean air condition to control for the effects of odor on symptoms. Several studies suggest that when subjects rate air contami- nated with various combinations of limonene, O3, and ofﬁce products (paper), they report dis- satisfaction with air quality (Knudsen et al. 2002; Tamas et al., in press). However, these studies did not measure health symptoms. To further clarify our findings, we con- ducted power calculations to estimate the size of the effect that would be necessary to detect 15 90 110 125 Minutes Figure 4. Cortisol over time for stress versus no stress regardless of exposure. VOLUME 113 | NUMBER 11 | November 2005 • Environmental Health Perspectives