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 1     those animals exposed to the mixture.  Edema of pulmonary connective tissue was more
 2     pronounced and alveolar Type 2 cells became swollen. Although this study was not
 3     quantitative, qualitative  observations led the authors to conclude that the lesions were not
 4     due to O3 alone but that NO2 played some, albiet undefined, contributory role.
 5          The effect of exposure duration on interaction was also noted in studies of Schlesinger
 6     et al. (1990, 1991). Rabbits exposed to 0.3 ppm O3 + 3 ppm NO2 for 2 h showed
 7     synergistic increases in certain BAL eicosanoids obtained immediately after exposure,
 8     whereas animals exposed to the same mixture for 2 h/day for 14 days showed no interaction
 9     for the same parameters.
10          A number of studies examined other biochemical responses to O3/NO2 mixtures (e.g.,
11     sulfhydryl metabolism and the activity of certain enzymes).  Some of these discussed in the
12     previous  O3 criteria document were found to involve synergism (e.g., Mustafa et al., 1984).
13     More recent studies of lung biochemistry also suggest that O3 and NO2 interact
14     synergistically. Ichinose and Sagai (1989) exposed rats and guinea pigs to 0.4 ppm
15     O3, 0,4 ppm NO2, or a mixture of the two pollutants continuously for 2 weeks.  No change
16     in lung peroxide production was observed in rats, but the mixture synergistically increased
17     peroxide levels in guinea pigs. The guinea pig showed no change in lung antioxidant content
18     following any exposure, whereas the mixture synergistically increased antioxidant levels hi
19     rat lung.   The conclusion of a significant interaction was based upon relative changes from
20     air controls following exposure to the mixture compared to changes following exposure to
21     each pollutant alone using the t-test, with synergism defined as a change greater than the sum
22     of the responses to individual pollutants; no specific test for interaction was performed.
23          Ichinose and Sagai (1989) also noted that levels of antioxidant enzymes in rat or guinea
24     pig lungs were variously affected by exposure to the above mixture. For example, GST was
25     decreased in both species exposed only to O3, but the mixture produced a reduction of this
26     enzyme in guinea pigs and  no change in rats. Thus, the occurrence of interaction was
27     dependent on endpoint as well as species.  This latter finding likely reflected interspecies
28     differences in biochemical defenses  against oxidant pollutants, given the results of a study by
29     Sagai et  al. (1987) with four animal species. This  study suggested that observed species
30     differences in lipid peroxide formation following exposure were  related to the relative content
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 1     of antioxidants and the specific composition of phospholipids and their fatty acids. The
 2     guinea pig was the most sensitive animal, and the hamster was the most resistant.
 3          The effects of exposures to O3/NO2 mixtures on lung lipid peroxides and antioxidant
 4     activity have been examined in a number of other studies (Ichinose et al., 1988; Lee et al.,
 5     1990; Sagai and Ichinose,  1991) and the results generally confirm those noted above, namely
 6     that such mixtures tend to  produce synergistic interaction.   However, there is also some
 7     evidence for antagonism.  Takahashi and Miura (1989) examined effects on the pulmonary
 8     xenobiotic system of rats exposed 1 or 2 mo to a mixture of 0.2 ppm O3  + 4 ppm NO2, as
 9     well as to each pollutant alone.  Ozone induced an increase in lung cytochrome P-450
10     content, but the activity of these enzymes was reduced by the addition of NO2  to the
11     exposure atmosphere; that is, the mixture resulted in levels intermediate between those found
12     with O3 or NO2 when given alone.  However, the reduction in enzyme activity induced by
13     NO2 was restricted  to those systems that had been increased by exposure to O3 alone. The
14     authors suggested that antagonism was due to the production of undefined secondary reaction
15     products in the exposure atmosphere.  A similar explanation was proposed to explain
16     observed synergism of lung antioxidant activity in another study (Lee et al., 1990).  Thus,
17     the response to any secondary product likely depends upon the endpoint examined, assuming
18     that the same reaction products  were formed in these two  studies.
19          The role of exposure parameters in producing an interaction between simultaneously
20     inhaled O3 and NO2 was examined by Gelzleichter et al. (1992a). Rats were exposed to
21     various combinations of O3 and NO2 for various durations (6, 8,  12, and 24 h),  such that the
22     C x T products were identical  for each of four exposure  sets.  As indicated in Table 6-24,
23     as the exposure duration increased, the exposure concentration of each component of the
24     mixture decreased.  Lavaged protein levels and recovered cells were the  endpoints.  For each
25     exposure combination, the additive response was predicted from results of exposure to each
26     pollutant alone, and then synergism was indicated when there was deviation from additivity.
27     Responses to exposure to  either O3 or NO2 alone for 6, 8, or 12 h showed that the product
28     of C x T was a constant  for the observed biological effects.  However,  less severe changes
29      occurred when delivery was at  the lowest dose rate (i.e., when the lowest concentration of
30      each pollutant was  delivered over the 24-h exposure duration). Exposure at higher  dose rates
31      (i.e., 6 to 12 h) increased the magnitude of the response.  Thus, the degree of response to

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 1     each pollutant alone was not a constant function of C x T throughout the entire range of
 2     dose rates, but was concentration driven, and was not identical at the highest and lowest
 3     rates.  Responses following exposure to the mixture  did not follow C x T even over the
 4     range of dose rates in which it was constant following exposure to the pollutants individually.
 5     Thus, interaction, in this case synergism, appeared to be concentration dependent, in that the
 6     response was disproportionately greater at the higher concentrations (higher dose rates) of the
 7     constituent pollutants in the mixture.  The response following exposure to the mixtures
 8     appeared to be a function of peak concentration, rather than of cumulative dose.  More
 9     recently, Rajini et al. (1993) noted that analysis of all kinetics following similar exposure to
10     mixtures also did not follow a C x T relationship.
11          All of the studies described above involved simultaneous exposure to O3 and NO2.
12     However, ambient exposure to these pollutants has temporal patterns, and exposure to one
13     agent may then alter the response to another subsequently inhaled agent.  The realism of
14     these studies is somewhat dependent upon their relationship to actual temporal patterns of
15     pollutants in ambient air (i.e., whether one  material  is the precursor of the other, as is the
16     case for O3 and NO^.  As described in the previous O3 criteria document (U.S.
17     Environmental Protection Agency, 1986), Fukase et al.  (1978) exposed mice for 7 days to
18     3 to 15 ppm NO2 for 3 h/day, followed by  1 ppm O3 for 3 h/day, and noted an additive
19     effect on the level of lung GSH.
20          Yokoyama et al.  (1980) exposed rats to 5 ppm NGj or 1 ppm O3 for 3 h/day, or to
21     NO2 for 3 h followed by O3 for 3 h/day, for various total durations up to 30 days,  and
22     assessed lung mechanics in postmortem lungs,  lung histology, and enzyme activity in
23     subcellular fractions  of lung tissue.  The activity of phospholipase A2 in the mitochondria!
24     fraction was increased in those animals exposed to O3 only  or to O3 after NO2, and the
25     response in the latter was significantly greater than that in the former.  A decrease in activity
26     of lysolecithin acyltransferase in the supernatent fraction was found only in those animals
27     exposed to both NO2 and O3.  Pulmonary mechanics showed a change in pulmonary
28     resistance (as a function of elastic recoil pressure) in the O3-only and NO2/O3-exposed
29     animals.  Histologically, the lungs of the animals exposed to both NC^ and O3 appeared
30     similar to those exposed to O3 alone.  However, a slight degree of epithelial necrosis in
31     medium bronchi not  found with either NO2 or O3 alone was seen in the animals  exposed to

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 1     both pollutants.  In addition, damage at the bronchoalveolar junction appeared to be
 2     somewhat more marked in animals exposed to both gases than in those exposed to O3 alone.
 3     This study suggested that sequential  exposures produced responses that were, in most cases,
 4     not greatly different from those due  to O3 alone.
 5          Aside from sequential exposures, simulation of ambient exposure scenarios  involving
 6     NO2 and O3 has been performed by  examining the effects of a continuous baseline exposure
 7     to one concentration of both pollutants, with  superimposed short-term peaks to a higher level
 8     of one or both.  The endpoint generally examined in this regard has been bacterial resistance.
 9     Studies reported in the previous criteria document (e.g., Ehrlich et al., 1979; Ehrlich, 1983)
10     in which mice were exposed to O3 under various scenarios of baseline concentrations of NO2
11     upon which were superimposed daily "peak" exposures to NO2, or a combination of NO2
12     and O3, suggested that exposure  with peaks can enhance response to pollutant mixtures,  and
13     that the sequence of peak exposures  was important in producing reduced resistance to
14     infection that was different from that due to exposure to the baseline concentration only.
15          As a comparison, toxicologic interactions for infectivity involving simultaneous
16     exposure to NO2 and O3 discussed in the previous O3 criteria document were found generally
17     to be additive following acute exposures, with each pollutant contributing to the observed
18     response when its concentration reached the threshold at which it would have affected
19     bacterial resistance when administered alone (Goldstein et al., 1974).  If the exposure level
20     of either NO2 or O3 was below this threshold, then the response was  due solely  to the
21     constituent inhaled at the more toxic concentration (Ehrlich et al., 1977).
22          More recently, Graham et al.  (1987) examined resistance to respiratory infection
23     (as measured by bacterial-induced mortality) in mice continuously exposed (for 15 days,
24     24 h/day) to baseline levels of an NO2/O3 mixture with two  daily, 1-h peaks of the mixture
25     at very high, high, intermediate, and low exposure concentrations (see Table 6-25 for
26     concentrations).  Animals were also exposed to the same baseline levels of either NO2 or
27      O3 onto which were superimposed two daily, 1-h peaks of the same single gas in
28     concentrations as above.  At the low concentration, only NC^ increased mortality.  At the
29      intermediate exposure level, the mixture was synergistic, whereas NO2 alone increased
30      mortality and O3 had no effect.  At the high exposure level, the combined exposure was
31      again synergistic, even though exposure to each gas separately also increased mortality.

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 1     A similar effect was seen at the very high level, although the combined exposure just missed
 2     statistical significance for synergism. These results are consistent with those of the earlier
 3     studies reported in the previous O3 criteria document and support the conclusion that
 4     response depends upon the specific exposure pattern. The results of Graham et al. (1987)
 5     are also consistent with those from the earlier studies with simultaneous exposures, in that
 6     the mixture did not produce any interaction when exposure to either gas alone was
 7     ineffective.
 8          The relationship between exposure and response is very complex, and seems to depend
 9     upon exposure duration, the ratio of O3 and NO2 concentrations, and other factors that may
10     include the production of secondary reaction products within the exposure atmosphere.  This
11     complexity was highlighted by the  study of Gelzleichter et al. (1992b), who examined effects
12     of combined or sequential exposures of rats to mixtures of Qj and NO2 at various
13     concentrations ranging from 0.2 to 0.8 ppm O3 and 3.6 to 14.4 ppm NO2.  Sequential
14     exposures consisted of 6 h of O3 at night, followed by 6 h of NO2 during the  day, or vice
15     versa;  concurrent exposures were for 6 h/day for 3 days. Various endpoints were examined,
16     and it was  noted that  sequential and concurrent exposures did not result in the same response.
17     Thus, lavage protein levels were increased additively with sequential exposure (in any
18     pollutant order), but were found to be greater than additive  with concurrent exposure.
19     An increase in the number of lavaged epithelial cells was additive for the 03 night/NO2 day
20     sequence, antagonistic for the NO2 night/O3 day sequence, and additive for concurrent
21     exposure.  An increase in the number of lavaged PMNs was additive for both sequential
22     conditions and was synergistic for  concurrent exposure. It was concluded that production of
23     synergism depended upon the concentration of each pollutant within the mixture, and
24     additivity would result for any endpoint when the  concentration of each component of the
25     mixture  fell below a certain threshold level. However, these threshold concentrations were
26     endpoint specific,  with some being more sensitive than others;  it was speculated that the least
27     sensitive assays were  based upon changes that were reversible,  whereas the most sensitive
28     ones were  irreversible.  The authors also noted that the extent of chemical reaction within the
29     O3/NO2 mixture atmosphere was related to the extent  of lexicological interaction, suggesting
30     that interaction was due to production of some secondary reaction product, which in this case
31     was suggested to be nitrogen pentoxide.  This particular chemical had also been suggested in

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 1     earlier studies to be responsible for interactions following exposure to NO2/O3 atmospheres
 2     (e.g., Diggle and Gage,  1955).
 3          The concentrations at which synergism occurred in the study of Gelzleichter et al.
 4     (1992b) discussed above (namely,  ^0.4 ppm O3 and ^7.2 ppm NO2) were higher than
 5     those that are generally found in ambient air. However, the threshold concentration for
 6     interaction was dependent upon exposure dose rate, with higher rates leading to lower
 7     threshold concentrations for synergism.  This suggests that a threshold may occur at ambient
 8     concentrations with longer term exposures.
 9          Although interaction is clearly modulated by environmental exposure factors, such as
10     concentration, duration of exposure, or specific exposure regime, host factors may also play
11     a role.  Mautz et al. (1988) examined the effect of exercise on rats exposed to mixtures of
12     O3 and NO2. Exercise modified the toxic interactions of combined pollutants, resulting in
13     synergistic interaction occurring at lower exposure concentrations of the constituent pollutants
14     than with exposure at rest.  Thus, a similar magnitude of response, namely an increase in the
15     extent of focal lesions in lung parenchyma,  was noted 2 days following a 4-h exposure to
16     0.6 ppm O3 + 2.5 ppm NO2 at rest, or with a shorter (3-h) exposure to lower
17     concentrations, 0.35 ppm O3  + 0.6 ppm  NO2, with exercise.  In both cases,  the response
18     was different from that due to either pollutant given alone. Furthermore, a greater response
19     was noted with a 3-h exposure to  0.6 ppm O3 + 2.5 ppm NO2 with exercise than to the
20     same mixture for the same exposure duration at rest. Thus, exercise also increased the
21     response at similar concentrations compared to rest. The effect of exercise was ascribed to
22     an increase in delivered dose or dose rate, due to increased VE. The ability of exercise to
23     enhance response to a pollutant mixture was also noted by Bhalla et al. (1987).
24           The study  of Mautz et al.  (1988) above also provided further evidence suggesting that
25      chemical reactions within the exposure atmosphere may play some role in toxicologic
26      interaction. In this case, nitric acid (HNO3) vapor was noted at concentrations ranging from
 27      0.02 to 0.73 ppm, depending upon the concentrations of the primary constituents.
 28      As discussed below, acids have been found to interact with O3. This study also found
 29      interaction to occur at a concentration of one of the components, namely NO2, that had no
 30     effect when administered alone.  Although this appears to contrast with the conclusions of
 31      Graham et al. (1987) above, the endpoints in these two studies were quite different.

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 1           Another aspect of pollutant interaction involves the ability of O3/NO2 mixtures to affect
 2      the course of other lung lesions (e.g., malignant tumor colonization).  Richters (1988)
 3      exposed mice to a mixture of 0.15 ppm O3 + 0.35 ppm NO2 for 7 h/day, 5 days/week for
 4      12 weeks, following which they were injected (iv) with viable melanoma cells.  The mice
 5      were sacrificed 3 weeks later and the lungs were examined for melanoma colonies.  Although
 6      exposure to O3 alone produced no change in the percentage of animals with colonies or in
 7      the average number of colonies per lung (compared to air control), exposure to the mixture
 8      produced an increase in the latter, suggesting to the authors that it faciliated cancer cell
 9      colonization.  However, the exact role played by O3 in the mixture is not clear because a
10     previous study had indicated that NO2 alone facilitates blood-borne cancer cell spread  to the
11      lungs (Richters and Kuraitis,  1981). Furthermore, the experimental model used is not
12      generally accepted as representing metastatic mechanisms.
13           Ichinose and Sagai (1992) also examined the ability of an O3/NO2 mixture to promote
14     primary lung tumor development. Rats were  injected (intraperitoneally) with
15      AMns(2-hydroxypropyl) nitrosamine (BHPN) and then were exposed for 13 mo to a mixture
16     of 0.05 ppm O3 + 0.4 ppm NO2, O3 alone, or to clean air (chamber control).  Although the
17     NO2 exposure was continuous, the O3 exposure was intermittent, with the concentration
18     altered between 0 and 0.1 ppm following a sine curve from 9 to 19 h of each day (resulting
19     in a daily mean concentration of 0.05 ppm).   One other group of rats served as a room
20     control, maintained in a clean room for 24 mo following injection of BHPN. After an 11-mo
21     recovery period, all animals were autopsied.  Compared to clean air-exposed animals, lung
22     tumor incidence was increased in mice exposed to the mixture; O3 alone did not increase the
23     tumor incidence.  However, the authors noted that tumor incidence in the room control group
24     was not different from that in the group exposed to the mixture, and suggested that the clean
25     air (chamber control) group should be used as a control in interpreting the data from the
26     pollutant-exposed animals.  The  enhanced incidence in mixture-exposed animals was
27     suggested to be due to synergistic increases in lipid peroxidation, which was noted in  other
28     studies (see Table 6-25). A complication in interpreting this  study is that a previous study
29     (Sagai and Ichinose, 1991) had suggested tumor development in animals exposed to an
30     O3 and NO2 mixture without BHPN, although this latter study involved a longer exposure
31     duration and somewhat higher pollutant concentrations.

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 1      6.4.2.2   Acidic Compounds as Copollutants
 2           Binary mixtures containing acids comprise another type of commonly examined
 3      exposure atmosphere.  Most of these involved acidic sulfate aerosols as the copollutant.
                                                                           3
 4      Peak (1-h) ambient levels of sulfuric acid (H2SO4) are estimated at 75 /tg/m  , with longer
 5      (12-h) averages about one-third of this (Spengler et al., 1989).
 6           Earlier studies that employed simultaneous single, repeated, or continuous exposures of
 7      various animal species to mixtures of acid sulfates and O3 found responses for several
 8      endpoints, including tracheobronchial mucociliary clearance, alveolar clearance, pulmonary
 9      mechanics, and lung morphology,  to be due solely to the O3 (U.S. Environmental Protection
10      Agency,  1986; Cavender et al., 1977; Moore and Schwartz, 1981; Phalen et al., 1980; Juhos
11      et al., 1978). However, synergism was noted for bacterial infectivity in mice (Grose et al.,
12     1982), for response to antigen in mice (Osebold et al., 1980), and for effects on lung protein
13     content and the rate of collagen synthesis in rats (Last et al., 1983, 1984a; Last and Cross,
14     1978). More recent studies performed since publication of the previous O3  criteria document
15     support the earlier finding of synergism between O3 and acid sulfates on lung biochemistry,
16     while providing possible explanations for underlying mechanisms.
17          Last et al. (1986) exposed rats for 7 days to O3 alone (at 0.96 ppm) and to mixtures of
18     O3 with one of three aerosols, namely sodium chloride, sodium sulfate, or ammonium sulfate
19     [(NH4)2SO4] (all at 5 mg/m3); only the (NH4)2SO4 was acidic. Lung protein content,
20     proline content, collagen synthesis rate, fibroblast numbers hi parechymal lesions, and the
21     volume of these lesions were examined following exposure. Mixtures of O3 with sodium
22     chloride or sodium sulfate produced changes that did not differ from those found with
23     O3 alone.  On the other hand, mixtures of (NIL^SC^ with O3 resulted in increases in all of
24     the measured parameters, and the increases were greater in magnitude than those due to
 25      O3 alone;  synergism was concluded. These results suggested  that acidity was necessary for
 26     synergism of the aerosols with O3.  This was further supported by demonstrating that
 27     significant interaction of O3 with  H2SO4, which is much more acidic than (NH4)2SO4,
 28     occurred at lower concentrations than was noted for mixtures  of O3 and (NH4)2SO4 (Warren
 29     and Last,  1987); interaction was suggested with H2SO4 concentrations as low as 40 jig/m
 30     (with 0.2 ppm O3) for some lung biochemical endpoints.  The studies above did not use any
 31     specific statistical test for interaction, and conclusions of interaction were based upon

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 1     findings of significant differences from responses following exposure to 63 alone and the
 2     absence of detectable responses following exposure to H2SO4 aerosols at the same and higher
 3     concentrations.
 4           Warren et al. (1986) found synergistic interaction with the above endpoints following
                                                  ^
 5     7 days of exposure to 0.2 ppm O3 plus 5 mg/m  (NH4)2SO4. However, exposure for only
 6     3 days produced responses that were not different from those noted with O3 alone.  This
 7     seems to indicate that the duration of exposure is a factor affecting the occurrence of any
 8     interaction.  However, exposure duration may also affect the type of interaction. In a study
 9     by Schlesinger et al. (1992a) in which rabbits were exposed to a mixture of 0.1 ppm O3 plus
                q
10     125 /tg/m  H2SO4 for 2 h/day, 5 days/week, a synergistic increase in bronchial epithelial
11     secretory cell number was noted after 4 mo of exposure, whereas antagonism was noted
12     following 8 mo of continued exposure.
13          The mechanism underlying interaction between acid sulfates and O3 is not known.  Last
14     et al. (1986) noted that similar sites of deposition for O3 and acid aerosols favored
15     synergism. A synergistic response of biochemical indices in rat lung with exposure to
16     1,000 /ig/m3 H2SO4 with 0.6 ppm  O3 was found when the acid droplet diameter was
17     0.5 /*m, whereas no increase compared to the O3-only response was  noted when the droplet
18     diameter was 0.02 /*m.  Apparently, the larger particles that deposited to a greater extent
19     within the bronchioalveolar junction,  the major target site for O3, were most interactive.
20          Observed synergism between O3 and acid sulfates  in rats was also suggested to be due
21     to a shift in the local microenvironmental pH of the lung following deposition of acid,
22     enhancing effects of O3 by producing a change in the reactivity or residence time of
23     reactants, such as free radicals, involved in O3-induced  tissue injury  (Last et al., 1984a).
24     If this was the only explanation, then the effects of O^ should be consistently enhanced by
25     the presence of acid in an exposure atmosphere.  However, in the study of Schlesinger et al.
26     (1992b), in which rabbits were exposed for 3 h to combinations of O3 at 0.1, 0.3, and
27     0.6 ppm with H2SO4  (0.3 /*m) at 50, 75, and 125 jDtg/m3, antagonism was noted when
28     evaluating stimulated production of superoxide anion by AMs harvested by lavage
29     immediately after exposure to 0.1 or 0.3 ppm O3 in combination with 75 or 125 jttg/m3
30     H2SO4, and also for AM phagocytic activity at all of the O3/acid combinations. Mixtures of
31     O3 (0.6 ppm) and another acid, namely HNO3 vapor (1,000 /ig/m3), (Nadziejko et al.,  1992)

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 1     also produced antagonism for certain aspects of the function of AMs harvested from acutely
 2     exposed rats. Although the deposition sites of both acid and O3 should be comparable in
 3     these two studies, perhaps the particular cellular endpoints examined are subject to this type
 4     of interaction.
 5          Last (1989) observed an apparent all-or-none response in rats exposed to the acid
 6     sulfate/O3 mixtures.  That is, there was no concentration-response relationship between the
 7     concentration of acid in the mixture and the extent of change in various endpoints, compared
 8     to effects observed with O3 alone.  In the study of Schlesinger et al. (1992b), a similar
 9     phenomenon was noted, but in this case, the concentration of O3 in the mixture did not
10     always influence the response compared to that seen with acid alone.  Thus, exposure-
11     concentration-response relationships noted with individual pollutants may not necessarily
12     reflect responses following exposure to their mixtures. This is consistent with the results of
13     Gelzleichter et al. (1992a) for mixtures of O3 and NO2.
14           The above studies involved simultaneous exposures to O3 and acidic pollutants, but
15     some studies involving sequential exposures to O3 and acid  sulfate aerosols were described in
16     the previous O3  criteria document.  For example, Gardner et al. (1977) found an additive
17     increase in infectivity when mice were exposed to 0.1 ppm  O3 for 3 h prior to a
18     2-h exposure to  900 jwg/m3 H2SO4,  whereas no difference from air control was noted when
19     the acid was administered prior to O3. Grose et al. (1980)  noted a reduction in ciliary
20     activity in isolated trachea! sections obtained from hamsters exposed to 0.1 ppm O3 for
21     3 h followed by exposure to  1,090 jwg/m3 H2SO4 for 2 h that was less in magnitude than that
22     found with exposure to acid alone (O3 alone had no effect).
23            More recently, Silbaugh and Mauderly (1986) examined the ability of O3 to increase
24     susceptibility to a subsequent exposure to H2SO4 in terms of producing airway constriction.
25     Guinea pigs were exposed to 0.8 ppm O3 for 2 h followed by H2SO4 (12 mg/m  for 1 h).
26     An increased volume of trapped gas in the lungs (the metric of constriction) was  seen with
27     both O3 alone and with the mixture, but the response to the latter did not differ from that due
28     to the former, and acid alone had no effect.  Thus, preexposure to  O3 in this case did not
29      affect response to a subsequent exposure to acid.
 30           Chen et al. (1991) examined the reverse exposure scenario, namely whether exposure to
 31      H2SO4 affected subsequent response to O3.  Guinea pigs were exposed to H2SO4 or H2SO4

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 1     coated on ultrafine zinc oxide particles.  A 1-h exposure to 0. 15 ppm O3 following a
 2     1-h exposure to acid (300 /*g/m , 0.09 fim) did not alter the response seen with acid alone,
 3     namely a decline in DLCO-  However, when single (1-h) or multiple (3-h/day, 7-day)
 4     exposures to acid-coated zinc oxide (24 or 84 jug/m equivalent H2SO4) were followed by a
 5     1-h exposure to 0.15 ppm O3, the effect on DLco appeared to be greater than additive,
 6     although no specific statistical test for interaction was performed.  This study suggested that
 7     prior exposure to acid increased the susceptibility of the guinea pig to subsequent exposure to
 8     O3, but it also showed that the manner in which the acid was delivered affected whether or
 9     not any interaction occurred.  It is likely that the number concentration of particles was
10     greater in the zinc oxide-H2SO4 aerosol than in the H2SO4 aerosol, and the interaction may
1 1     reflect this greater particle number.
12
13     6.4.2.3   Other Copollutants
14          Although the bulk of the database for binary mixtures of O3 involves NO2 or acids, a
15     few studies examined responses  to combinations of O^ with other pollutants.
16          Reuzel  et al. (1990) exposed rats to mixtures of O3 (0.2, 0.4, 0.6 ppm) +
17     formaldehyde (HCHO) (0.3 to 3.0 ppm).  Although exposure to the mixtures did not alter the
18     nature or extent of histological lesions, namely cilia loss and epithelial hyperplasia, compared
19     to exposure to each pollutant alone, a site-specific synergistic increase in turnover of nasal
20     epithelial cells was found with all concentrations of HCHO together with 0.4 ppm Oj.
21     A lack of such response  with 0.8 ppm O3 was ascribed to an O3 -induced alteration in
22     breathing pattern, which reduced the delivered dose. It was,  however, noted that interaction
23     occured  only when one constituent of the mixture was administered at cytotoxic
24     concentrations, an exposure scenario that rarely occurs in ambient air.  In any case, the
25     authors concluded that because cell proliferation likely plays a role in carcinogenesis, and
26     that if mixtures potentiate cell proliferation, then exposure to pollutant mixtures may increase
27     cancer risk.
28          Mautz et al.  (1988) exposed rats for 3 h, at both rest and exercise, to a mixture of
29     0.6 ppm O3  + 10 ppm HCHO.   A synergistic increase in nasal epithelial cell turnover
30     followed exposure with exercise, whereas exposure  at rest resulted in no difference from that
31     seen with HCHO alone.  Likewise, exposure to the mixture with  exercise resulted in an

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 1     increase in the number of focal lesions in lung parenchyma compared to either O3 or HCHO
 2     alone, but exposure at rest resulted in a lower incidence of lesions than seen with O3 alone.
 3     This latter observation was ascribed to an effect of the HCHO upon breathing pattern,
 4     producing a change in inhaled dose of O3 that did not occur with exercise.
 5          Nishikawa et al. (1992) examined the effect of sequential exposure to cigarette smoke
 6     and O3 in altering airway responsivity to inhaled brochoconstrictor challenge and trachea!
 7     vascular permeability in guinea pigs.  Animals were exposed to 1 ppm Oj for 0.5 h followed
 8     by 5 puffs of cigarette smoke, or to 1 ppm O3 for  1.5 h followed by  10 puffs.  Exposure to
 9     O3 and five puffs increased responsivity and vascular permeability immediately after
10     exposure, whereas no effect on either endpoint was noted with either pollutant given alone.
11     Exposure to O3 and 10 puffs also increased responsivity and permeability, but to the same
12     extent as did the lower concentration  mixture or exposure  to O3 alone, whereas exposure to
13     10 puffs of smoke only increased responsivity. Thus, sequential exposure to O3 and cigarette
14     smoke enhanced the magnitude of response compared to either pollutant alone, but the
15     duration of response was not altered.
16          The potential role of O3 in enhancing fibrotic lung disease by interaction with silica was
17     examined by Shiotsuka et al. (1986),  who  exposed rats with developing silica-induced
18     fibrosis to O3 at 0.8 ppm for 6 h/day, 5 day/week for 37  exposure days.  Silica had been
19     instilled (2,  12, or 50 mg) on Day 1 of the study and exposure to O3 began on Days 3 or
20     4 postinstillation.  There was found to be no interaction between silica and O3 in
21     development of fibrosis, as assessed biochemically (lung content of hydroxyproline) or
22     histopathologically.  Although an increase was found in the ratio of hydroxyproline to total
 23     protein in the group exposed to the mixture and instilled with the highest amount of silica,
 24      this was not considered by the authors to be biologically significant.
 25
 26     6.4.3    Complex (Multicomponent) Mixtures Containing Ozone
 27          Ambient pollution in most areas is a complex mix of more than two chemicals, and a
 28     number of studies have examined the effects of exposure  to multicomponent atmospheres
 29     containing O3.  Some of these attempted to simulate photochemical reaction products
 30     occurring under actual atmospheric conditions.  However, the results of these studies are
 31     often difficult to interpret due to chemical interactions between the components and the

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 1      resultant production of variable amounts of numerous secondary reaction products, and a lack
 2      of precise control over the ultimate composition of the exposure environment. In addition,
 3      the role of O3 in the observed biological responses is often obscure.
 4           One type of experimental multicomponent atmosphere that has been examined is
 5      ultraviolet-irradiated and nonirradiated automobile exhaust mixtures.  Irradiation leads to the
 6      formation of photochemical reaction products that are biologically more active than those in
 7      nonirradiated mixtures.  Such mixtures are characterized by total oxidant concentrations
 8      (expressed as O3) in the range of 0.2 to 1.0 ppm. Although the effects described following
 9      exposure were not necessarily uniquely characteristic of O3 and although O3 could have been
10     responsible for some or even most of them, the biological effects have been difficult to
11      associate with any one particular component in most cases. Effects of exhaust mixtures on
12      different species have been discussed in the previous O3 criteria document (U.S.
13      Environmental Protection Agency, 1986).  Pulmonary function changes were demonstrated in
14     guinea pigs after short-term exposures to irradiated exhaust and in dogs after long-term
15      exposure to both irradiated and nonirradiated exhaust mixtures.
16          Additional studies of complex mixtures have been performed since publication of the
17     previous O3 criteria document.  Kleinman et al.  (1985) exposed rats (Sprague-Dawley, male,
18     7 weeks, nose-only) for 4 h to atmospheres, designed to represent photochemical pollution,
19     consisting of 0.6 ppm (1,180 ng/m3) O3 + 2.5 ppm (4,700 jttg/m3) NO2 + 5 ppm
20     (13,100 /ig/m3) sulfur dioxide (SO2) + particles.  The paniculate phase consisted of
21     1,000 /tg/m3 of either H2SO4 or (NH4)2SO4, laced with iron sulfate [Fe^SO^] and
22     manganese sulfate. The metallic salts act as catalysts for the conversion of sulfur IV into
23     sulfur VI, and the incorporation of gases into the aerosol droplets.  The respiratory region
24     was examined for morphological effects.  A confounding factor in these studies was the
25     production of HNO3 vapor in atmospheres that contained O3  and NO2, a phenomenon
26     discussed previously,  and nitrate in those that contained O3 and (NH4)2SO4, but not NO2.
27     Nevertheless, a significant enhancement of tissue damage was noted with exposure to
28     atmospheres  containing H2SO4, or secondarily produced HNO3, compared to those
29     containing (NH4)2SO4, a less acidic compound.   In addition,  there was some suggestion that
30     the stronger acidic atmospheres resulted in a greater area of the parenchyma becoming
31     involved in lesions, which  were characterized by a thickening of alveolar walls, cellular

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 1     infiltration in the interstitium, and an increase in free cells within alveolar spaces.
 2     An increased rate of nasal epithelial cell turnover was noted following exposure to
 3     atmospheres containing paniculate acids compared with exposure to either O3 alone or to a
 4     binary mixture of O3 + NO2.  Furthermore, exercise seemed to potentiate the nasal and
 5     parenchymal responses to the complex mixtures containing strong acids (Kleinman et al. ,
 6     1989), a finding similar to that with binary mixtures of O3 and NO2,  or O3 and HCHO,
 7     previously discussed.
 8          Bhalla et al. (1987) examined the effects of a seven-component atmosphere, similar to
 9     that above, on epithelial permeability of rat lungs (Sprague-Dawley, male, 47 to 52 days
10     old). The animals were exposed for 2 h (chambers, relative humidity [RH]  = 85%) to the
11     following:  O3 (0.6 ppm) + NO2 (2.5 ppm) + SO2 (5 ppm)  + ferric oxide
12     (Fe2O3) (241  /ig/m3) + (NH4)2SO4 (308 to 364 jig/m3) 4- Fe^SO^ (411 to 571 /*g/m3) +
13     MnSO4 (7 to 9 /ig/m3).  The response to this mixture was compared to that following
14     exposure to O3 (0.6 ppm) + NO2 (2.5 ppm); O3 alone (0.6 or 0.8 ppm); or NO2 alone (6 or
15     12 ppm).  As above, the complex mixture was found to result in production of HNC^, in this
16     case at measured concentrations of 1,179 to 2,558 /ig/m3 (0.46 to 1.02 ppm); the O3 + NO2
17     atmosphere also resulted in some HNO3 vapor formation. Epithelial permeability was  found
18     to increase immediately following exposure to  O3, O3 + NO2, or to the complex mixture.
19     Although the magnitude of this change was similar following exposure to O3 alone or in
20     combination with other pollutants, there was increased persistence of effect after exposure to
21     either the binary or complex mixture.
22           Prasad et al.  (1988) used a similar multicomponent atmosphere and examined effects on
23     AM surface receptors.  Rats (Sprague-Dawley, male, 200 g) were exposed for 4 h/day, for
24     7 or 21 days to a mixture of O3 (0.3 ppm) + NO2 (1.2 ppm) + SO2 (2.5 ppm) +
25      (NH4)2S04 (270 /xg/m3)  + Fe^SO^  (220 ptg/m3) + MnSO4 (4 ^g/m3) +
26      Fe2O3 (150 jug/m3), or to O3 alone. Both the mixture and O3 alone resulted in a decrease  in
27      Fc receptor activity beginning immediately after the last exposure.  Exposure to the complex
28      atmosphere for 7 days resulted in a response similar to that seen with C^ alone, but
29      continued exposure to this mixture for up to 21 days resulted in an even greater reduction in
 30     receptor function compared to O3 alone.  However, as with most studies of complex
 31      mixtures, although the response to the mixture was  different from that found with O3, the

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 1     role of other constituents was not clear. Phagocytic function of AMs was also examined
 2     following exposure to the mixture, but there were no O^-only controls for comparison.
 3          Mautz et al. (1985a) examined the effects of a complex mixture upon pulmonary
 4     mechanics in exercising dogs.  Exposures  (nose-only) were for 200 min to a mixture of
 5     O3 (0.45 to 0.7 ppm) + SO2 (4.8 to 5.2 ppm) + H2SO4 (800 to 1,200 /*g/m3, 0.2 /mi) +
 6     catalytic salts of Fez  (SO4)3 and MnSO4.  A greater increase in resistance and decrease in
 7     compliance was found with the complex atmosphere than with 63 alone,  but the effect was
 8     ascribed to the presence of the H2SO4.  Although synergism was implied, it could not be
 9     definitively concluded because the mixture was not tested without Oj.
10          Mautz et al. (1991) further examined the ability of components of acidic fogs to alter
11     the response to O3.  Rats (Sprague-Dawley, male, 7  weeks, n = 12/group) were exposed for
12     4 h (nose-only; temperature  =  22 to 23 °C, RH = 82 to 83%) to 0.4 ppm 03 or to a
13     mixture of 0.4 ppm O3 + 670 /tg/m3 HNO3 vapor + 610 /ig/m3 H2SO4 particles (0.32 /*m).
14     Exposure to either O3 or the mixture resulted in comparable changes, namely development of
15     a rapid, shallow breathing pattern, a decrease in fatty acid composition of pulmonary
16     surfactant,  and focal  parenchymal lesions with thickened alveolar septa and cellular
17     infiltration.  The lack of any modulation of the O3-induced effects by acids prompted the
18     authors to raise the question of the  sensitivity of rats to inhaled acids.  Although responses to
19     any pollutant are somewhat species dependent, there is  some evidence that rats are not the
20     most sensitive species to acidic aerosols (U.S. Environmental Protection  Agency, 1989).
21     As discussed previously, the extent of interaction within any one species of animal is
22     endpoint dependent, and it is likely that the sensitivity of various endpoints is species
23     dependent. Thus, rats do show biochemical changes (e.g., in collagen metabolism) with
24     exposure to fairly low levels of acidic aerosols in combination with 03 (see Table 6-10),
25     although these involved longer duration exposures.  In any case,  the underlying reasons for
26     the lack of interaction in the complex mixture study above remain unclear.
27          Kleinman et al. (1989) exposed  rats  (Sprague-Dawley, male, 7 weeks, nose-only) to a
28     mixture of O3 (0.8 ppm)  + SO2 (5 ppm)  + H2SO4 or  (NH^SC^  (1,000 /ig/m3) at high RH
29     (85%), and noted a delay in early clearance of inert particles from  the lungs, compared to
30     air-exposed controls. However, it is difficult to relate any effects to the 03 because
31     responses to O3 alone were not examined.

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 1           The ideal complex mixture is one that actually exists in the ambient environment.
 2      Saldiva et al.  (1992) exposed rats (Wistar, male, 2 mo) for 6 mo to actual atmospheres of
 3      Sao Paulo, Brazil, with controls maintained for the same period of time in a clean, rural
 4      environment.  The mean pollution levels over the exposure period were as follows:
 5      0.011 ppm O3,  1.25 ppm CO, 29.05 /xg/m3 SO2, and 35.18 /xg/m3 particles. The animals
 6      exposed to the urban air showed evidence of bronchial secretory cell hyperplasia, ciliary
 7      structural changes, increased viscosity of mucus, and impaired mucociliary clearance.
 8      Although chronic exposure to air pollution may result in pulmonary dysfunction, the specific
 9      components producing the response could not be determined.
10          Inhalation exposures to air pollutants are, of course, the ideal way to assess interaction,
11      but in vitro exposures may provide indications of potential interactions.  Shiraishi and
12     Bandow (1985) exposed Chinese hamster V79 cells for 2 h to photochemical reaction
13     products produced from the reaction of propylene and NO2 in a smog chamber. The
14     resultant exposure atmospheres consisted of various proportions of propylene (0.07 to
15     0.16 ppm), N02 (0.22 to 0.28 ppm), O3 (0.09 to 0.38), PAN (0.04 to 0.41 ppm),  and
16     HCHO (0.23 to 1.50 ppm).  Exposures to NO2 and O3 alone were also performed.  All of
17     the complex mixtures resulted in an increased frequency of sister chromatid exchange and
18     growth inhibition. The effects of the mixture were greater than those due to either O3 or
19     NO2 alone for  sister chromatid exhange, but growth inhibition was similar to that induced
20      solely by O3.  The authors concluded that the observed effects were not due to any single
 21      compound within the mixture, but rather to various compounds producing multiple effects.
 22
 23      6.4.4   Summary
 24           It is difficult to summarize the role that O3 plays in response to exposure to binary
 25      mixtures, and  it is even harder to determine its role in response to multicomponent
 26     atmospheres.  One of the problems in understanding interactions is that although the specific
 27     mechanisms of action of the individual pollutants within a mixture may be known, the exact
 28     bases for toxic interactions have not been clearly elucidated.  There  are, however, certain
 29     generic mechanisms that may underlie pollutant interactions.  One is physical, involving
 30     adsorption of one pollutant onto another and subsequent transport to more or less sensitive
 31     sites, or sites  where one of the components  of the mixture normally would not deposit in

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 1     concentrated amounts.  This, however, probably does not play a major role in O3-related
 2     interactions.  A second mechanism involves production of secondary product(s) that may be
 3     more lexicologically active than the primary materials. This has been demonstrated or
 4     suggested in a number of studies as a basis for interaction between O3 and NO2.  A third
 5     mechanism involves biological or chemical alterations at target sites that affect response to
 6     the O3 or the copollutant.  This has been suggested to underly interactions with mixtures of
 7     O3 and acid sulfates.  A related mechanism is an O3- or copollutant-induced physiological
 8     change, such as alteration  in ventilation pattern, resulting in changes in the penetration or
 9     deposition of one pollutant when another is present.  This has been implicated in enhanced
10     responses to various O3-containing mixtures with exercise.
11          Evaluation of interactions between O3 and other copollutants is a complex procedure.
12     Responses are dependent upon a number of host and environmental  factors, such that
13     different studies using the  same copollutants may show different types or magnitudes of
14     interactions.  The occurrence and nature of any interaction is dependent upon the endpoint
15     being examined, and is also highly related to the specific conditions of each study, such as
16     animal species, health status, exposure method, dose, exposure sequence, and the
17     physiocochemical characteristics of the copollutants. Because of this, it is difficult to
18     compare studies, even those examining similar endpoints,  that were performed under
19     different exposure conditions.  Thus, any description of interactions is really valid only for
20     the specific conditions of the study in question and cannot be generalized to all conditions of
21     exposure to a particular chemical mixture.  Furthermore, it is generally not possible to
22     extrapolate the effect of pollutant mixtures from studies on the effects of each component
23     when given separately.  In any case, what can be concluded from the database is that
24     interactions of O3-containing mixtures are generally synergistic, but antagonism has been
25     noted in a few studies, depending upon the various factors noted above, and O3 may produce
26     more significant biological responses as a component of a mixture than when inhaled alone.
27     Furthermore, although most studies have shown that interaction occurs only at higher than
28     ambient concentrations with acute exposure, some have demonstrated interaction at more
29     environmentally relevant levels (e.g., 0.05 to 0.1 ppm O3 with NO^ with repeated
30     exposures.
31

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 1     6.5   SUMMARY AND CONCLUSIONS
 2     6.5.1    Introduction
 3          In the past 30 years, thousands of research studies on the effects of Qj in laboratory
 4     animals have been reported in the literature.  This body of evidence presents a clear picture
 5     of the types of alterations O3 can cause on respiratory tract host defense mechanisms,
 6     biochemistry, structure, and lung function.  Less is known about carcinogenic potential and
 7     effects on organs distant from the lungs.  These types of effects are observed in many animal
 8     species from mice to nonhuman primates, lending credence to the qualitative extrapolation of
 9     these effects to humans.   The major issue is what levels, durations, and patterns of exposure
10     are capable of causing these effects in humans.  Extrapolation is discussed in Chapter 8.
11     Suffice it to say here that the animal lexicological studies both assist in interpreting
12     observations made in O3-exposed humans and extend our knowledge of potential human
13     hazards that can never be studied adequately in humans.
14          This summary and conclusion section deals exclusively with the effects of O3, alone
15     and  in mixture.  Other photochemical oxidants either have been evaluated elsewhere (NO2
16     and  formaldehyde; U.S. Environmental Protection Agency, 1993; Grindstaff et al., 1991) or
17     in an earlier O3 criteria document (U.S. Environmental Protection Agency, 1986).  This
18     section is first organized  by respiratory tract effects, systemic effects,  and effects of
19     mixtures. Between this chapter and the animal lexicological  chapter in the 1986 document
20     (U.S. Environmental Protection Agency,  1986), over 1,000 references are cited. Although
21     all of them contribute to  choosing and understanding the key  issues to be summarized here,
22     there must obviously be a highly selective choice which references to  include here.
23     Generally, the papers discussed here were selected either because they represent lowest
24     effective concentration for an endpoint or they significantly influenced a particular
25     conclusion.
26
27     6.5.2   Molecular Mechanisms of Effects
28           Molecular mechanisms (the manner in which chemical reactions of O3  are translated
29     into biological effects) are alluded to in different sections  of  this document.  Studies which
30     link O3  chemistry with O3 effect measurements would greatly strengthen the theoretical basis
31     for  understanding the biological effects of O3. They would also allow examination of the

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 1     similarity between animals and humans, thus strengthening interspecies extrapolations.
 2     Ozone has been shown to react directly with a variety of biomolecules that are present in
 3     both animals and humans.  Most of the attention has been centered on polyunsaturated fatty
 4     acids and carbon-carbon double bonds, although reactions with sulfhydryl, amino, and some
 5     electron-rich compounds may be equally  important.  Free radicals may be involved, and
 6     antioxidant defenses appear to lessen the effect of these reactions.  A "molecular target" for
 7     03—the biomolecules most affected by reaction with O3 or most crucial in mediating the
 8     observed  responses, has not been identified for any of the endpoints studied.  In fact, the
 9     target may be different  for different endpoints.
10           An important concept in evaluating molecular targets was recently elucidated by Pryor
11     (1992) who suggests (based on reaction and diffusion rate data) that the 03 molecule does not
12     penetrate through cell membranes or even the surfactant layer of the lung.  Instead, a
13     "reaction cascade" forms intermediates (organic or oxygen free radicals, lipid
14     hydroperoxides, aldehydes, hydrogen peroxide, etc.), which penetrate into the cells, causing
15     the biological effects observed  (Pryor et al., 1991).  Confirmation of such O3-induced free
16     radical  autoxidation of lipids has been sought in vivo, but the indirect nature  of the
17     measurement methods produced equivocal results.  More direct evidence has  been obtained
18     by Kennedy  et al. (1992) who used electron spin-trapping methods to  measure a
19     concentration-related increase in radical adducts of the lipid fraction of lungs  from
20     O3-exposed rats. Increased radical signals were detected after a 2-h exposure to >0.5 ppm
21     O3,  but because the rats' respiration was stimulated by CO2, the effective dose would be
22     greater than  it appears.   Oxidized (oxygenated) biomolecules that result from reaction with
23     O3 may also mediate the effects of O3.  Studies by Hatch et al.  (1994) show  that crude
24     fractions of the lung lining layer become labeled with oxygen-18 after exposure to oxygen-
25     18-labeled O3. The label is concentrated in the airway  lining layers, and the amount of
26     oxygen-18 incorporation in this layer appears to be correlated with effects of
27     O3 (permeability and inflammation) in both rats and humans.  These findings are  consistent
28     with the hypothesis that O3 reacts with the lining of the lung, that the same types of
29     interactions occur in both animals and humans, and that these reactions lead to similar
30     effects.
31

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 1     6.5.3    Respiratory Tract Effects
 2     6.5.3.1  Effects on Host Defenses
 3          Several systems defend the respiratory tract of the host against infectious and neoplastic
 4     disease as well as nonviable inhaled particles.  All these systems can be affected by O3.  The
 5     mucociliary clearance system moves particles deposited on the mucous layer (either through
 6     deposition from the air stream or entry of cells/cellular debris from the alveoli) upwards and
 7     out of  the lower respiratory tract.  The nasal passages also have an effective clearance
 8     system.  Concentrations as low as 0.15 ppm O3 (8 h/day, 6 days)  caused structural changes
 9     in the  nasal respiratory epithelium (e.g., ciliated cell necrosis, shortened cilia) of monkeys
10     (Harkema et al., 1987).  Ciliated cells are also lost or damaged in the conducting airways of
11     the lower respiratory tract after short exposures (e.g., 0.96 ppm, 8 h,  monkeys; Hyde et al.,
12     1992).  Mucous chemistry also is changed (McBride et al., 1991). Sufficient morphologic
13     damage would be expected to have functional consequences. Acute exposures (0.6 ppm, 2 h)
14     slow mucociliary particle clearance in rabbits, but repeated exposures  (up to  14 days) caused
15     no effects.  Alveolar clearance is slower and involves clearance of particles through
16     interstitial pathways to the lymphatic system or movement of particle-laden AMs up to the
17     bottom of the mucociliary  escalator.   Effects on alveolar clearance are concentration-
18     dependent. A single 2-h exposure of rabbits to 0.1 ppm accelerated clearance up to 14 days
19     postexposure, exposure to 0.6 ppm caused no effect, and a higher concentration  (1.2 ppm)
20      slowed alveolar clearance (Driscoll et al., 1986).  Alveolar clearance of asbestos particles
21      was slowed by a 6-week exposure to  an urban pattern of O3 (Pinkerton et al., 1989).
22           Alveolar macrophages are the first line of defense against microbes and nonviable
23      particles that reach the pulmonary region of the lung.  They phagocytize particles, kill
 24     microbes, and interact with lymphocytes in the development of an immune response.  Thus,
 25     their proper functioning is critical. Alveolar macrophages from several species of animals
 26     exposed acutely to O3 can exhibit decreased  phagocytosis; decreased lysosomal enzyme
 27     activities and superoxide anion radical production, both of which function in killing bacteria;
 28     alterations in membrane morphology; chromosomal damage; decreased cytotoxicity to tumor
 29     cells; increased release  of PGE2 and  PGF2a; and alterations in the number of AMs.
 30     Phagocytic changes are the most investigated. Exposure of rabbits to level as low as
 31     0.1 ppm (2 h/day) decreased non-specific phagocytosis (of latex microspheres) after 2 or 6,

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 1     but not 13, days of exposure (Driscoll et al.,  1987).  Recovery from the single 2-h exposure
 2     to 0.1 ppm was complete by 7 days postexposure.  This pattern of response was confirmed in
 3     mice  for Fc-receptor mediated phagocytosis (Gilmour et al., 1991; Canning et al., 1991).
 4           The humoral- and cell-mediated immune system of the lung is also affected by O3.
 5     Generally, T-cell dependent immunity is more susceptible than B-cell dependent immunity,
 6     but most immune functions examined have exhibited effects.  However, because relatively
 7     few studies have been conducted, it is not currently possible to adequately interpret the
 8     impacts of O3-induced alterations on the immune system (e.g., decreases in mitogenic
 9     responses of T cells, alterations in T:B cell ratios in the MLN).  Only a few studies have
10     attempted to correlate immunological changes and infectious disease outcome.  Van Loveren
11     et al. (1988) infected rats with Listeria, exposed them to 03 for 1 week (0.26 to 1.02 ppm)
12     and measured several endpoints.  Ozone increased Listeria-induced mortality and severity of
13     lesions in the lung and liver. They interpreted these  findings as due to O3-induced impaired
14     clearance of the bacteria caused by decreased AM function and decreased cellular immunity
15     (e.g., decreased delayed-type hypersensitity and decreased T/B cell ratios in MLN).
16           A reasonably large body of evidence indicates that the impact of 03 on one or several
17     host defense mechanisms leads to the inability of animals  to fight bacterial  infection and
18     alters the course of viral infection.  Antibacterial models are more commonly used. Mice
19     exposed for 3 h to 0.4 ppm  O3 have decreased intrapulmonary killing of S.  zooepidemicus
20     (Gilmour et al., 1993a; Gilmour and Selgrade, 1994).  Similar results  have been obtained  for
21     S. aureus at a slightly higher concentration (Goldstein et al., 1971b).   Correlations have been
22     made between O3 exposure and decreases in AM phagocytosis, decreases in bactericidal
23     activity, growth of bacteria in the lungs, presence of bacteria in the blood,  and mortality in
24     mice  (Coffin and Gardner,  1972b; Gilmour and Selgrade, 1994). The lowest O3 exposure
25     causing increased streptococcal-induced mortality is 0.08 ppm for 3 h in mice (Coffin et al.,
26     1967;  Coffin and Gardner,  1972b; Miller et al., 1978). However, prolonged intermittent
27     exposure to 0.1 ppm for 15  weeks only slightly increased the mortality (Aranyi et al., 1983),
28     and continuous exposure for 15 days to 0.1 ppm with two daily 1-h peaks (5 days/week) to
29     either 0.3 or 0.5 ppm did not enhance mortality in the same model  system  (Graham et al.,
30     1987).  Prolonged exposure (1 to 2 weeks) also did not affect bactericidal activity to
31     S. aureus (Gilmour et al., 1991).

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 1          Generally, short-term exposure to O3 does not affect viral titers in the lungs of mice
 2     infected with influenza virus, but reduced numbers of lung tissue T and B cells reduced
 3     antibody titers to the virus, mortality, lung pathology, and increased lung wet weight do
 4     occur (Selgrade et al.,  1988; Jakab and Hmieleski, 1988).  Ozone also enhances
 5     postinfluenzal alveolitis and structural changes that begin at 30 days postinfection (Jakab and
 6     Bassett, 1990). The complexity of the interaction of viral infection and O3 exposure  is
 7     further illustrated by Selgrade et al. (1988) who found that the effects of O3 on influenza
 8     virus infection was dependent on the temporal relationship of O3 exposure and day of
 9     infectious challenge. Also, interferon, which can be induced by viral infection, mitigates the
10     O3-induced lung lessons in mice, raising the possibility that certain stages of viral infection
11     may have interactions with the lung that are  different from other stages (Dziedzic and White,
12     1987b).
13
14     6.5.3.2   Effects on Inflammation and Permeability
15          The barrier function of the respiratory  tract is disrupted by O3, allowing cellular and
16     fluid components from the blood to enter the lung and allowing certain types of  substances in
17     the lung to enter the blood.  Markers of inflammation generally included increased proteins
18     and PMNs in BAL.  Concurrent with these events, but not necessarily interdependently,
19     AMs liberate more arachidonic acid, which results in the production of biologically active
20     LTs and PGs.  Similar responses are observed in mice, rats, rabbits,  guinea pigs, hamsters,
21     and nonhuman primates.  After acute exposure, the lowest effective concentration that
22     increases BAL protein and number of PMNs is 0.12 ppm (mice, 24 h of exposure, BAL
23     immediately after exposure) (Kleeberger et al.,  1993a).  However, typically the  increase in
24     BAL protein is maximal roughly 16 to 24 h postexposure. In rats exposed to 0.8 ppm for
25      6-h and examined by lavage and morphometry at several times postexposure, the increase in
26      nasal PMNs occurs  sooner and wanes about the time that these cells are increasing in number
27      in the lungs (Hotchkiss et al., 1989a,b).  It  should be recognized that BAL can measure the
28      protein and cells accessible by lavage,  including the resident material (i.e., may include
 29      protein from O3-induced cellular destruction) and the material entering from the
 30      tissue/circulation.  Thus,  interstitial inflammation, which has been observed in several
 31      species microscopically, is not detectable by BAL.

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 1          Several C x T studies have been conducted in mice using BAL protein as an endpoint.
 2     In two studies, there was combination of various C's (0.1 to 2.04 ppm) and T's (1 to 12 h),
 3     resulting in a number of different C x T products (Rombout et al.,  1989; Highfill et al.,
 4     1992). Both of these studies showed that the influence of T increased as C increased.  That
 5     is, there was no simple relationship of Ca X T* = constant product. However, at the lowest
 6     C x T products, there was a more equivalent influence of C and T.  Gelzleichter et al.
 7     (1992b) used a single C  x T product composed of a variety of Cs and Ts for up  to 3 days of
 8     exposure.  The 24 h/day exposure group had less response than the  other groups  which
 9     responded equivalently.  Effects of longer term exposure on permeability and inflammation
10     are more complex to interpret (also see subsequent discussion on lung structure).
11     Histological examination of rat lungs exposed to 0.5 ppm O3 for 5 days showed more
12     inflammatory cells in the alveoli after 5  days (2.25 h/day) of exposure compared  to 1 day of
13     exposure (Tepper et al., 1989).  In contrast, the increase in BAL PMNs that occurred after
14     Day 1 of exposure of rats had resolved by Day 4 (7 h/day) (Donaldson et al., 1993).
15          Some studies suggest that although protein and PMN increases are observed
16     concurrently, this may be more of a function of the experimental design than the  biological
17     sequence of events.  For example, in rats depleted of PMNs with anti-PMN serum, Oj did
18     not increase BAL PMNs, but BAL protein was still increased (Pino et al., 1992b).  Also
19     Young and Bhalla (1992) observed an increase in trachea! protein earlier than increased
20     tracheal PMNs.  They interpreted this and other related results to suggest that the recruited
21     PMNs may serve to sustain an increase in permeability.
22
23     6.5.3.3  Effects on  Structure, Function, and Biochemistry
24          Theoretically, and in some cases empirically, lung structure, function, and biochemistry
25     are linked.  Correlations are not exact because of differences in available measurement
26     methods (e.g., most lung function tests used do not sensitively measure the function of the
27     smallest airways, where the "classical" O3 lesion is observed) and some independence of
28     effects (e.g., a transient change in breathing frequency would not be morphologically
29     detectable).  Also, most biochemical measurements are made of whole lung, rather than focal
30     areas of damage, and only some enzyme activities measured would be expected to be
31     correlated to structure and/or function (e.g., collagen metabolism, antioxidant metabolism).

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 1           After acute exposure to O3, the most commonly observed effect in several species is
 2      tachypnea (increased fB and decreased VT) with little (if any) change in VE; the lowest
 3      exposure causing tachypnea was 0.2 ppm for 3 h in rats (Mautz and Bufalino, 1989).  Other
 4      effects reported after acute exposure to  < 1 ppm include increased RL and decreased Cdyn,
 5      TLC, VC, FRC, RV, FVC, DLCO, and N2  slope (e.g., Fouke et al., 1991; Mautz et al.,
 6      1985b; Miller et al., 1988).  However,  these changes are not observed in all studies,
 7      probably due to differences in animal species, measure method, and exposure protocols.
 8      With rare exception, concentrations well in excess of 1 ppm O3 are required to increase
 9      airway reactivity.
10           Two C x T studies of pulmonary function using acute exposure periods have been
11      performed.  Costa et al. (1989) found that FVC, DLcO, and N2 slope decreased with
12      increasing C x T products in rats and that the influence of T is greater at higher Cs.
13      In guinea pigs, Nishikawa et al.  (1990) observed that airway responsiveness to methacholine
14     increased at higher  C x T products (e.g., at 90 ppm • min, but not at 30 ppm  • min). The
15     authors concluded that T was an important factor in the 63 response.
16          When rats were exposed for 5 days (2.25 h/day, with CO2 to stimulate ventilation
17     equivalent to light exercise in humans)  to 0.35, 0.5, and 1 ppm 03, the change in shape of
 18     the flow-volume curve and tachypnea peaked occurred on Days 1 and 2, but by Day 5, there
 19     was no difference from control (except at 1 ppm) (Tepper et al., 1989).  This attenuation is
20     similar to that observed in humans. However, in other, similar groups of animals,
21     histological changes in the lung progressed and BAL protein remained elevated.  Other
22     similarities between laboratory animals and humans in their pulmonary function responses to
 23      short-term O3 exposure are explored in Chapter 8.
 24           Ozone causes similar types of alterations in lung morphology in all laboratory animal
 25      species studied.  The most affected cells are the ciliated epithelial cells of the airways and
 26     Type 1 cells in the gas exchange region. Within the nasal cavity, anterior portions of the
 27     respiratory and transitional epithelium  are affected. Cilia are lost or damaged; some ciliated
 28     cells become necrotic, are lost,  and are replaced with nonciliated cells.  Mucous secreting
 29     cells are affected.
 30           The centriacinar region (CAR, the junction of the conducting airways and the gas
 31     exchange region) is a primary target, possibly because it receives the greatest dose of

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 1     O3 delivered to the lower respiratory tract (see Chapter 8) and has Type 1 epithelial cells
 2     covering a large surface area.  Even though there are significant interspecies differences in
 3     the structure of the CAR (e.g., primates, including humans, have respiratory bronchioles,
 4     which are rudimentary or absent in laboratory animals such as rats or mice), it is the target
 5     in all species studied.  Exposure to O3 causes loss of cilia or necrosis of the ciliated cells,
 6     leaving a bare basement membrane that is replaced by nonciliated bronchiolar cells, which
 7     may become hyperplastic after longer exposures.  Mucous secreting  cells can be affected, but
 8     not as significantly as ciliated cells. Type 1 cells are also damaged and can be sloughed
 9     from the surface; Type 2 cells, which are thicker, replace them. Sometimes Type 2 cells
10     differentiate into Type 1 cells.  This epithelial remodeling is accompanied by an
11     inflammatory response in the CAR, primarily consisting of an increase  in numbers  PMNs in
12     the earlier stages and increase in numbers of AMs in later stages; interstitial edema occurs.
13     With increased duration of exposure, alveolar septa in the CAR thicken due to increased
14     matrix, basement membrane, collagen, fibroblasts, and a thickened alveolar epithelium.
15           These patterns of change have different relationships to duration of exposure,  as
16     illustrated by Dungworth (1989) (see Figure 6-3; Section 6.2.4.5).  Inflammatory changes
17     peak after a few days of O3 exposure; are still observable, but to a much lesser degree, in
18     tissue during months of exposure;  and begin to return to control values after exposure  ceases.
19     In contrast, epithelial hyperplasia rapidly increases during about the  first week of exposure,
20     plateaus as exposure continues, and begins to decrease slowly when  exposure stops.
21     Interstitial fibrosis requires months of exposure to be observed microscopically and increases
22     slowly, but when exposure ceases, it can still persist or continue to increase.  Numerous
23     studies using several different species and experimental approaches support  these findings.
24     Only a few of the studies (primarily those using more sensitive morphometric measurements)
25     are selected here to illustrate key points and show correlations with pulmonary function and
26     lung biochemistry.  Only rat and nonhuman primate studies are discussed because most
27     investigations were conducted on them.  At equivalent exposures, nonhuman primates appear
28     to be more responsive than rats (Section 6.2.4).
29           Generally short-term exposures to concentrations ^0.2 ppm O3 do not cause  changes in
30     the nasal cavities of rats or nonhuman primates detectable by LM, except for inflammation
31     and an occasional delayed postexposure finding of mild hyperplasia.  For example,  Hotchkiss

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 1     et al.  (1989a) reported inflammation in the nasal epithelium of rats up to 66 h after a 6-h
 2     exposure to levels as low as 0.12 ppm O3; there was no necrosis, loss of cilia, or hyperplasia
 3     even at 1.5 ppm.  After 3 days (22 h/day) of exposure, >0.4 ppm (but not 0.2 ppm) caused
 4     loss of cilia and hyperplasia and metaplasia of the nasal epithelium of rats (Reuzel et al.,
 5     1990). Nonhuman primates appeared to be more responsive. Harkema et al. (1987)
 6     observed that exposure  to 0.15 or 0.3 ppm for 6 or 90 days (8 h/day) caused necrosis of
 7     ciliated cells, shortened cilia, and increased mucous  granule cells in the respiratory
 8     epithelium; alterations in cell numbers were also found in the transitional epithelium.
 9          Within the CAR, a number of alterations occur.  In rats and monkeys ciliated and
10     Type  1 cells become necrotic and are sloughed from the epithelium as soon as  the first 2 to
11     4 h of an exposure to about 0.5 ppm (Stephens et al.,  1974a,b).  Repair, as shown by
12     increased DNA synthesis by nonciliated bronchiolar  and Type 2  cells, begins by about 18 to
13     24 h of exposure (Evans et al., 1976; Stephens et al.,  1974a; Castleman et al., 1980),
14     although cell damage continues (Castleman et al.,  1980).  The lesion is fully developed by
15     about 3 days of continuous exposure,  after which the rate or repair exceeds the rate of
16     damage.  The increase in antioxidant enzyme activities (e.g., succinate oxidase, G6PD, and
17     6PGD) parallels the increase in Type  2 cells, which  are rich in these enzymes; the  increase
18     in the Type 2 cell population is probably responsible for these biochemical changes (Bassett
19     et al., 1988a; U.S. Environmental Protection Agency,  1986).
20           Lesions in the CAR are one of the hallmarks of O3 toxicity, having been well
21     established.  The study  by Chang et al. (1992) provides examples of some of the patterns of
22     cellular alterations.  Chang et al. (1992) exposed rats to an urban pattern of O3 (0.06 ppm
23     background, 7 days/week on which were superimposed 9-h peaks [5 days/week] slowly rising
24     to 0.25 ppm) for 78 weeks and made periodic examinations of the CAR TB and proximal
25     alveoli by TEM morphometry during and after exposure.   Type  1 cells had a larger volume
26     at Week 13  and increased numbers at Weeks 13 and 78; there was no such changes at
27     17 weeks after exposure ceased.  Type 2 cell volume per area of basement membrane
28     increased immediately after Week 78 and was still increased 17  weeks after exposure ceased.
29     Interstitial cells and matrix were increased after Weeks 1,  13, and 78, but returned to control
30     by 17 weeks after exposure ceased.  However, epithelial and endothelial basement membrane
31     were thickened and accompanied by increased collagen fibers at the later examination times

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 1     and 17 weeks after the 78-week exposure ended.  In TBs, surface areas of ciliated and
 2     nonciliated cells decreased during exposure. Pulmonary function studies conducted in
 3     identically exposed groups of rats were consistent with the morphometric findings (Tepper
 4     et al., 1991).  Generally expiratory resistance was increased (suggesting central airway
 5     narrowing), but it was only significantly different from control at 78 weeks.  Tidal volume
 6     was reduced at all evaluation times.  Overall, breathing frequency was reduced, but no single
 7     evaluation time was significant. Monkeys exposed to a higher concentration of
 8     O3 (0.64 ppm, 1 year) also showed increased resistance and decreased flows, which were
 9     interpreted as central and peripheral airway narrowing; during a 3-mo postexposure period,
10     decreases in static lung compliance persisted (Wegner, 1982).
11          Several studies have demonstrated distal airway remodeling.  This bronchiolization of
12     the alveoli  is so named because bronchial epithelium replaces the Type 1 and 2 cells typical
13     of alveolar ducts, resulting in the appearance of respiratory bronchioles in rats  and an
14     increase volume  fraction and volume of respiratory bronchioles in monkeys.  This has been
15     observed at exposures as low as 0.5 ppm O3 (50 days) in rats (Moore and Schwartz, 1981)
16     and as low as 0.25 ppm (8 h/day, 18 mo) in monkeys (Tyler et al., 1988). Inflammation
17     occurs concurrently, perhaps indicating an  influence on remodeling. In monkeys,  such
18     bronchiolization  can persist 6 mo after a 1-year (8 h/day) exposure to 0.64 ppm ends (Tyler
19     etal.,  199 Ib).
20          Exposure regimens can have unexpected impacts on experimental outcomes.   Several
21     investigations of combinations  of O3 "episodes" or O3 "seasons" with clean-air periods have
22     been examined.  In the first of these, Last  et al. (1984b) compared air control rats to
23     2 groups of rats  exposed to 0.96 ppm.  One group received a 90-day (8 h/day) exposure
24     ("daily") the other had intermittent units of 5 days of O3 (8 h/day) and 9 days of air, such
25     that there were 35 O3 exposure days over the 90-day period ("episodic").  Both groups  had
26     equivalent  increases in lung  collagen.  Using a similar exposure regimen, Barr et al. (1990)
27     found equivalent CAR remodeling and volumes of CAR lesions in both groups. In contrast,
28     respiratory bronchiole thickness increased in the daily group only  and the CAR interstitium
29     increased in thickness only in the episodic  group.  Monkeys were studied more extensively
30     after a daily (8 h/day) exposure to 0.25 ppm for 18 mo and a "seasonal" exposure only
31     during the  odd months of the 18-mo period (Tyler et al., 1988). Most morphometric

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 1     measurements were similar between the two groups (e.g., both had respiratory bronchiolitis).
 2     However, only the daily group had an increased number of AMs in the lumen and
 3     interstitium.  Only the seasonal group had increased lung collagen  content; increased chest
 4     wall compliance, suggesting delayed lung maturation; and increased inspiratory capacity.
 5     This body of work indicates that under these types of exposure circumstances, the simple
 6     product of C x T does not predict the outcome.  Indeed half the O3 (on a C  XT basis)
 7     caused equivalent or more effects than a "full" O3 exposure.
 8          The complexity of understanding C  X T relationships is  further illustrated by Chang
 9     et al. (1991).  She and her colleagues compared two different  exposure regimens (one a
10     square wave and the other an urban pattern O3) on the basis of C  x T products.  There was
11     a linear relationship between C  x T products and the increase in Type 1 cell volume in the
12     CAR; a similar observation on Type 2 cell volumes was less robust. There was not such
13     relationship for other morphometric endpoints in the same animals.  Cell proliferation in the
14     nasal epithelium does not increase linearly with increasing C x T, but does increase linearly
15     with increasing C (Henderson et al., 1993).
16          Long-term exposure also thickens CAR alveolar septa, due to an increase in
17     inflammatory cells, fibroblasts, and amorphous extracellular matrix (Fujinaka et al., 1985;
18     Barry et al., 1985; Zitnik et al., 1978).  There is some morphological evidence of mild
19     fibrosis (i.e., local increase in collagen) in CAR interalveolar septa (Last et al., 1979;
20     Boorman et al., 1986;  Chang et al.,  1992; Pickrell et al., 1987b; Freeman et al., 1974;
21     Moore and  Schwartz, 1981). Biochemical evidence supports these findings, even though
22     biochemical approaches would be expected to be less sensitive because the whole lung (rather
23     than focal lesions) is examined.  Last et al. (1979) directly demonstrated the  correlation by
24     observing increased  collagen histologically and biochemically  (collagen synthesis rate) in rats
25     similarly exposed to 0.5 to 2.0 ppm for 7 to 21 days.  The increase became greater with
26     increasing concentration and duration of exposure. Similar correlations were observed at a
27     higher concentration by Pickrell et al. (1987b).  The increased collagen content can persist
28     after exposure ceases (Chang et al.,  1992; Hussain et al., 1976a,b; Last et al., 1984b), but
29      some, not all, studies suggest that higher concentrations (>0.5 ppm) may be required for
30      such persistence (Last and Greenberg,  1980;  Pickrell et al., 1987b). Collagen cross-links
31      were studied in monkeys  exposed to 0.61 ppm O3 for 1 year  (8 h/day) (Reiser et al., 1987).

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 1     Earlier examination of these same monkeys revealed that collagen content was increased
 2     (Last et al., 1984b).  When specific collagen crosslinks were measured, the increase in
 3     "abnormal" cross-links observed immediately after exposure remained in the lungs at 6 mo
 4     postexposure.
 5          As discussed above, O3 can cause fibrotic changes.  However, there is no evidence that
 6     O3 causes emphysema, using the currently accepted morphological definition of emphysema.
 7          These morphologic/morphometric and biochemical findings of fibrotic changes are
 8     supported by some pulmonary function studies.  For example, rats exposed for up to
 9     78 weeks using the same urban exposure protocol as Chang et al. (1992) exhibited reduced
10     lung volume and hastened N2 washout patterns, consistent with a "stiffer" lung (i.e.,
11     restrictive lung disease) (Costa et al., 1994).
12
13     6.5.3.4   Genotoxicity and Carcinogenicity of Ozone
14          At this point in time, a significant amount of research has been conducted to determine
15     whether O3 is  genotoxic or carcinogenic.  Many of the earlier experiments have flaws in
16     experimental design or have used O3 concentrations far above levels that could occur in the
17     ambient air. In evaluating the data, a number of conclusions can be made.  In vitro exposure
18     of naked plasmid DNA to very high O3 concentrations results in single and double-strand
19     breaks in the DNA, as confirmed by gel electrophoresis and electron microscopy studies
20     (Hamelin, 1985). Testing of O3 in various mutagenesis assays has led to marginal or small
21     results in a number of assays and negative results in other assay systems.  Ozone is not
22     mutagenic in Salmonella strains TA98, TA100, TA104, and TA1535 and causes, at most,
23     weak effects in strains TA102 that are not strictly concentration-dependent (Dillon  et al.,
24     1992; Victorin and Stahlberg, 1988a,b).  Extremely high concentrations of 03 (50 ppm)
25     caused  mutation to streptomycin resistance in E. coli and caused various types of mutations
26     in the yeast, S. cerevisiae, but it was a weak mutagen compared to known strong mutagens
27     in the yeast system (L' Herault and Chung, 1984; Dubeau and Chung, 1982).  Ozone was
28     not mutagenic in the AT.  tabacam or Tradescantia mutation assay systems (Gichner et al.,
29     1992).  Hence, overall, the data on the mutagenicity of O3 are mixed:  negative in six
30     assays,  marginally positive in one system, and weakly positive in two systems.  The present
31     data indicate that O3 is,  at most, a weak mutagen, but further data are needed in mammalian

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 1     cell systems to draw definitive conclusions regarding this point. There are some data
 2     indicating that O3 may cause chromosome breakage in cultured cells, but in vivo animal
 3     studies are conflicting (Zelac et al., 1971a,b; Tice et al., 1978).  A human study with an
 4     appropriate experimental design was negative (McKenzie et al., 1977; McKenzie, 1982).
 5          Regarding carcinogenicity,  O3 has been shown to induce morphological transformation
 6     in cultured  C3H/10T1/2 mouse embryo cells and in SHE cells, and to cause a synergistic
 7     transformation in cells treated also with gamma radiation (Borek et al., 1986, 1989b).
 8     However, the results could be due to interactions of O3  with the culture medium that
 9     generate chemical species different from those produced in vivo.  Whole animal
10     carcinogenesis assays performed in  strain A mice have demonstrated marginal increases in
11     tumor yield that were not statistically significant or concentration-dependent (Hassett et al.,
12     1986; Last  et al., 1987).  The NTP study demonstrated that O3 was not a tumor promoter or
13     a co-carcinogen when NNK-treated male  F344/N rats were exposed for 2 years to 0.5  ppm
14     O3 (National Toxicology Program,  1993). This NTP study showed no evidence of
15     carcinogenic activity in male or female F344/N rats, equivocal evidence of carcinogenic
16     activity in male B6C3Fi mice, and some  evidence of carcinogenic activity in female B6C3FJ
17     mice at a high concentration (1.0 ppm).  Hence,  O3 may be a weak pulmonary carcinogen
18     only in female B6C3FJ mice at toxic concentrations.
19          At present, O3 is shown to be nonmutagenic in some assay systems; at most, weakly
20     mutagenic in a few  assay systems; and clastogenic in vitro but not in vivo.  Ozone can
21     transform cells in vitro. Ozone does not cause concentration-dependent tumor induction that
22     is statistically significant in hamsters, Wister male rats, F344/N male or female rats, male or
23     female A/J mice, or Swiss-Webster male mice. There  are ambiguous data for pulmonary
24     carcinogenesis in male B6C3Fj mice and weak carcinogenesis data in female B6C3Fj  mice
25     from chronic exposure to 1.0 ppm O3. Therefore, O3 has been shown to be a carcinogen
26     only in female E6C3Fl mice in one experiment.  Ozone needs to be studied further  in well-
27     designed experiments to determine definitively whether it can significantly induce tumors
28     reproducibly in B6C3Fj mice and other species of rats  and mice in a concentration-dependent
29     manner.  Because the rat and  mice inhalation carcinogenesis experiments do not agree and
30     mechanistic data to assist in interpretation are either not useful or not available, these
31     pulmonary carcinogenesis results in female B6C3Fi mice cannot be extrapolated to humans.

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 1     Hence, there is no justification to consider O3 a human carcinogen at this time.  Further,
 2     since it required a chronic exposure to 1 ppm O3 to induce pulmonary tumors in mice, it is
 3     possible that pulmonary toxicity that does not occur at lower levels contributed to the tumor
 4     development.
 5
 6     6.5.3.5   Risk Factors
 7          Factors that increase the delivered dose of O3> decrease biochemical defense
 8     mechanisms, or increase cellular sensitivity can increase the impact of a given O3 exposure.
 9     The most commonly studied factors include exercise, age, and nutrition.
10          As  discussed in Chapter 8, exercise increases the dose of O3 delivered to the
11     respiratory tract and alters its distribution. As would be expected, exercise during exposure
12     enhances the effect of O3.  This has been demonstrated by Mautz et al. (1985b) who showed
13     that exercising rats had more extensive lung lesions than those at rest.  Similarly, Tepper
14     et al. (1990, 1994) found that rats were more responsive to O3 when coexposed to CO2 to
15     increase  ventilation, simulating exercise.
16          A number of studies have been conducted to compare the effects of O3  on various ages
17     of mice and rats, from 1 day old to older adults.  Interpretation of these studies is difficult
18     because prior to weaning, the huddling behavior of the neonates with their dams as well as
19     the bedding material (present in some  studies) may have affected the concentration of O3 in
20     the breathing zone and hence the subsequent  delivered dose.   Generally, in short-term
21     exposure biochemical  studies of antioxidant metabolism, there was a decrease or no  change
22     in enzyme activity in neonates.  As age increased after weaning, the typical increase in
23     antioxidant metabolism became greater with age (Elsayed et  al., 1982; Tyson et al.,  1982;
24     Lunan et al., 1977; Mustafa et al., 1985). Stephens et al (1978) found that morphological
25     effects did not occur in animals exposed prior to weaning at 21 days of age.   This may
26     explain the results of Barry et al. (1985, 1988) who found no morphometric  differences in
27     the CAR and TB in rats that started a  42-day exposure at ages of 1 day and 42 days.
28     In identically exposed rats, however, Raub et al. (1983) found more, though admittedly
29     subtle, pulmonary function changes in the youngest group of animals.  Yokoyama et al.
30     (1984) did not detect any age-related differences in lung function of rats at 4-, 7-, and
31     10-weeks of age. Although O3-induced increases in BAL protein and PMNs do not show an

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 1      age-dependence, BAL prostaglandins increased sooner and more leukocytes were dead in
 2      younger (13-days old) rats, compared to adults (e.g., 16- weeks old) (Gunnison et al., 1990,
 3      1992a). Age (5- versus 9-weeks old) did not influence the O3-induced decrease in lung
 4      bactericidal activity (Gilmour et al., 1993a).
 5           The literature on O3-exposed pregnant animals is extremely sparse.  Exposure of rats
 6      (1 ppm 6 h) on day 17 of pregnancy or days 3, 13, and 20 of lactation caused a greater
 7      increase in lung permeability and inflammation than that observed in nonpregnant rats
 8      (Gunnison etal., 1992b).
 9           Numerous reports document that animals made vitamin E  deficient are more susceptible
10      to the biochemically detected effects of O3 (e.g.,  lipid changes, antioxidant metabolism
11      changes) (U.S. Environmental Protection Agency, 1986; Pryor, 1991).  Generally, the
12     research shows that although vitamin E deficiency enhances susceptibility to lung biochemical
13     changes, there is not a proportionate relationship between vitamin E supplementation (above
14     normal levels) and protection from O3. Also, vitamin E deficiency did not alter the impact
15     of O3 on lung structure (Chow et al.,  1981).  Vitamin C deficiency also has an influence.
16     Guinea pigs deficient in vitamin C  had a greater  (compared to vitamin C-normal animals)
17     when exposed acutely to 0.5, but not  1.0 ppm O3 (Slade et al., 1989).
18
 19      6.5.4   Systemic Effects
 20           Theoretical studies (Pryor, 1992) indicate that the O3 molecule does not penetrate to the
 21      blood, yet there are numerous reports of systemic effects (i.e., effects on lymphocytes,
 22      erythrocytes, serum,  CNS, parathyroid gland, circulatory system,  and liver).  Possibly one or
 23      several of the reaction products of O3 (see Section 6.2.1) penetrates the lung tissue,  or
 24      perhaps some systemic responses are secondary to pulmonary effects.  Although a variety of
 25      clinical chemistry changes occur after O3  exposure, they cannot be interpreted and will not
 26     be discussed here (see U.S. Environmental Protection Agency, 1986 and Section 6.3).
 27     Effects on systemic immunity are  discussed earlier (6.5.3.1).
 28
 29     6.5.4.1   Central Nervous System and Behavioral Effects
 30           Acute exposure to O3 caused transient changes in behavior.  The lowest exposure
 31      causing effects was 0.12 ppm for  6 h in rats;  wheel-running activity decreased  (Tepper et al.,

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 1     1985; Tepper and Weiss, 1986).  Because exercising animals were exposed in these studies
 2     (i.e., they received a higher dose of O3), it is not surprising that higher 03 concentrations
 3     (0.5 ppm, 6 h) are required to affect sedentary behavior (e.g., operant behaviors such as
 4     lever pressing for food reinforcement) (Weiss et al.,  1981). Mice show aversive responses
 5     to O3 (60s, 0.5 ppm) by terminating O3 exposure (Tepper et al., 1985).  The lowest
 6     exposures causing effects are impacted by the type of reward.  For example, 03 had less
 7     effect on behaviors to avoid electric shock (Ichikawa et al., 1988) than on behaviors to obtain
 8     food or access exercise (Tepper et al.,  1982, 1985; Weiss et al., 1981).
 9
10     6.5.4.2   Cardiovascular Effects
11          In rats, O3  can cause bradyarrhythmia at exposures as low as 0.1 ppm for 3 days;
12     bradycardia, at exposures as low as 0.2 ppm for 2 days; and decreased mean arterial blood
13     pressure, at exposures as low  as 0.5 ppm for 6 h (Arito et al., 1990, 1992;  Uchiyama and
14     Yokoyama,  1989; Watkinson et al., 1993; Yokoyama et al., 1989;  Uchiyama et al., 1986).
15     There is an  interaction between some of these responses and thermoregulation in the rat.  For
16     example, when heart rate decreased, the core temperature of the exposed rats also decreased;
17     and when exposures were conducted at higher ambient temperature, there was no change in
18     core temperature or heart rate (Watkinson et al.,  1993).  Such interactions add to the
19     complexity of extrapolating this type of response  to humans, and therefore,  without more
20     information, qualitative extrapolation would be highly speculative.
21
22     6.5.4.3   Reproductive and Developmental Effects
23          No reports of "classical" (e.g., 2-generation studies) reproductive assays with O3 were
24     found.  Kavlock et al. (1979,  1980) performed several developmental toxicity experiments in
25     rats.  Pregnant rats exposed intermittently (8 h/day) from 0.44 to 1.97 ppm (ppm during
26     early, mid-, or late- gestation or  the entire period of organgensis (days 6 to 15) had no
27     significant teratogenic effects.  Continuous exposure during mid gestation increased the
28     resorption of embryos.  Postnatal growth and behavioral development were  also investigated.
29     There was no effect on neonatal  mortality (up to  1.5 ppm).  Pups from dams  exposed
30     continuously to 1 ppm during mid- or late gestation weighed less 6 days after birth.  Pups
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 1      from pregnant rats exposed continuously to 1 ppm during late gestation had delays in
 2      behavioral development (e.g., righting, eye opening).
 3
 4      6.5.4.4  Other Systemic Effects
 5           A number of investigations have shown the effects of O3 on the pituitary-thyroid-
 6      adrenal axis,  as evidenced by changes in circulating hormones and morphological changes in
 7      the thyroid and parathyroid glands.  (U.S. Environmental Protection Agency, 1986).
 8      No newer studies could be found.
 9           Several  approaches have been used to study the effects of O3 on the liver: increase in
10      sleeping time following the injection of drugs (e.g., pentobarbital) metabolized by the liver,
11      drug pharmacokinetics, and changes in liver enzymes.  The lowest exposure causing
12     increased sleeping time from pentobarbital was 0.1 ppm for at least 15 or  16 days (3 h/day)
13     in female mice (Graham et al.,  1981).  In three species of animals, only females were
14     affected (Graham et al., 1981).   Pentobarbital pharmacokinetics was marginally (p = 0.06)
15     slowed in mice exposed to 1  ppm O3 for 3 h  (Graham et al., 1985); theophylline clearance
16     was slowed in older rabbits exposed to 0.3 ppm O3 for 5 days  (3.75 h/day) (Canada and
17     Calabrese, 1985). Ozone has caused both increases, decreases, and no changes in liver
18     xenobiotic metabolism, depending upon the exposure and enzyme being measured (U.S.
19     Environmental Protection Agency, 1986).
20
21     6.5.5   Effects of Mixtures
22           Humans in the real world are exposed to complex mixtures of gases and particles.
 23      Sufficient evidence exists to  know that the health outcome is dependent on the mixture, but
 24      the relative role (or even  the exact identity) of the "major" components is not known.
 25      Because of this, it is  crucial to evaluate the health effects of O^ in light of epidemiological,
 26     human clinical, and animal  lexicological studies.  For the purposes of this document, an
 27     interaction is considered to  occur when the response to the mixture is statistically
 28     significantly higher (synergism) or lower (antagonism) than the sum of the individual
 29     pollutants.  Most animal lexicological studies of interactions have been conducted with binary
 30     mixtures (predominantly NO2 and H2SO4).  The rarer reports on complex mixtures are
 31     interesting, but less helpful because often they did not include a group exposed only to 03,

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 1     and therefore, knowledge of the role of O3 is confounded. Thus, only the binary mixture
 2     studies will be summarized here.  This research has demonstrated that exposure to O3 in
 3     combination with another chemical can result in antagonism, additivity, or synergism,
 4     depending on the animal species, exposure regimen, and endpoint studied.  Interpretation is
 5     further complicated by the fact that most studies used exposure regimens unlike the real
 6     world in terms of ratios of pollutant concentrations, "natural" sequencing of exposure
 7     patterns, and other factors. For example, when O3 and NO2 exposures were sequential
 8     (in any order), there was an additive increase in BAL protein, as compared to a synergistic
 9     increase when the exposures were concurrent (Gelzleichter et al., 1992a).
10          A range of interactions has been shown with O3 and NO2 combinations.  For example,
11     a 2-week exposure to an O3-NO2 mixture (0.4 ppm of both) synergistically increased
12     antioxidants in the lungs of rats,  but not guinea pigs; peroxide levels were synergistically
13     increased in guinea pigs, but not rats; GST activity was decreased in guinea pigs and
14     unchanged in rats (Ichinose and Sagai, 1989). Most of the interaction studies using lung
15     biochemical endpoints display synergism.  A rare exception was the antagonism to the
16     increase in lung cytochrome P-450 content caused by 0.2 ppm O3 (1 to 2 mo) when the rats
17     were coexposed to 4 ppm NO2 (Takahashi and Miura,  1989). Combinations of various acute
18     exposure durations and O3 and NO2 concentrations did not follow a C x T relationship for
19     increased lung permeability, but were synergistic at higher C x T products (Gelzleichter
20     et al., 1992b). For pulmonary host defenses against bacterial infection, the interaction is
21     dependent on the exposure pattern.  Graham et al. (1987) showed that a 15-day exposure of
22     mice to mixtures of O3 and NO2, each having a baseline  level with two daily 1-h peaks of
23     the pollutant, resulted hi synergism when exposure to either gas alone caused an increase in
24     bacterial-induced mortality.
25          Both synergistic and antagonistic interactions have been found with combinations of
26     O3  and acidic sulfates.  Warren et al. (1986) reported that with 3 days of exposure to
                              'j
27     0.2 ppm O3 plus 5  mg/m (NH4)2SO4,  O3 alone was responsible for increasing BAL protein,
28     collagen synthesis rate, and other parameters; but, by 7 days of exposure, synergism
29     occurred.  When rabbits were exposed for 4 mo (2 h/day, 5 days/week) to 0.1 ppm O3 plus
                •5
30     125 /*g/m  H2SO4, there was a synergistic increase in epithelial secretory cell number,
31     whereas 8 mo of exposure resulted in antagonism  (Schlesinger et al., 1992a).  Antagonism

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1     was also observed for effects or certain AM functions after acute exposures to O3-H2SO4
2     mixtures (Schlesinger et al., 1992b).  Sequential exposures to O3 and H2SO4 have also been
3     examined.  Exposure to O3 did not influence the subsequent effects of H2SO4 on
4     bronchoconstriction in guinea pigs (Silbaugh and Mauderly, 1986).  Gardner et al. (1977)
5     found an additive increase in bacterial infectivity when mice were exposed acutely to
6     0.1 ppm O3 before, but not after, H2SO4.
7          In summary, the animal lexicological studies clearly demonstrate the major complexities
8     and potential importance of interactions, but do not provide a scientific basis for predicting
9     the results of interactions under untested ambient exposure scenarios.
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 i          7.  HUMAN HEALTH EFFECTS OF OZONE AND
 2               RELATED PHOTOCHEMICAL OXIDANTS
 3
 4
 5      7.1   INTRODUCTION
 6          In the previous chapter, results of ozone (O3) studies in laboratory animals were
 7      presented in order to understand the wide range of potential effects that might occur in
 8      exposed human populations and to expand our understanding of the mechanisms of
 9      O3 toxicity and the basic exposure-response relationships for O3.  The concept of
10      quantitatively extrapolating results from laboratory animals to humans is further explored in
11      Chapter 8.  Whenever possible, however, risk assessment and risk management of pollutants
12      should be based on direct evidence of their health effects in human populations. Information
13      on human health responses to O3 can be obtained through controlled human exposure studies
14     on volunteer subjects or through field and epidemiological studies of populations that are
15      exposed to ambient air containing O3. Controlled human studies typically use fixed
16     concentrations of O3 under carefully regulated environmental conditions, whereas realistic
17      O3 exposure conditions occur in field and epidemiology studies, but are more variable.  The
18      primary purpose of all these studies, however, is to obtain exposure-response data for O3.
19     This chapter will summarize the results of controlled human, field, and epidemiologic studies
20     on the health effects of exposure to O3 that have been published in the peer-reviewed
21      literature.  Further evaluation of the most important key information from this chapter, as it
22     relates to the rest of the document, will be provided in the remaining chapters.
23         Most of the scientific information selected for review and comment in this chapter
24     comes from the literature published since completion of the previous O3 criteria document
25     (U.S. Environmental Protection Agency,  1986).  Some of these newer studies were briefly
26     reviewed in the supplement to that document (U.S. Environmental Protection Agency, 1992),
27     but more intense evaluation of these studies is included here. In order to give a broader
28     overview of the known human health  effects of O3, the older literature is summarized and
29     specific studies whose data were judged to be significant because of their usefulness in
       December 1993                          7-1       DRAFT-DO NOT QUOTE OR CITE

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 1     deriving the current National Ambient Air Quality Standards (NAAQS) are briefly discussed.
 2     The reader should, however, be referred to the more extensive discussion of these "key"
 3     studies in the previous document.  Other, older studies are also briefly discussed in this
 4     chapter if they are (1) open to reinterpretation because of newer data, or (2) potentially
 5     useful in deriving revised NAAQS for O3.  To further aid in the development of this chapter,
 6     summary tables of the relevant O3 literature are included for each of the major subsections.
 7     In summarizing the human health  effects literature, changes from control are described if
 8     they were  statistically significant at a probability (p) value less than 0.05.  A specific p value
 9     is provided, however, if it aids understanding of the data, particularly trends toward
10     significance, or if major effects need to be emphasized.  Where appropriate, critique of a
11     statistical procedure also is mentioned.
12
13
14     7.2    CONTROLLED HUMAN EXPOSURE STUDIES
15     7.2.1   Pulmonary Function Effects of One- to Three-Hour Ozone
16             Exposures
17     7.2.1.1  Healthy Subjects
18     Introduction
19          The pulmonary  responses observed in healthy human subjects exposed to ambient
20     O3 concentrations consist of a decreased inspiratory capacity, a mild bronchoconstriction, a
21     rapid shallow breathing pattern during exercise, and  subjective symptoms of cough and pain
22     on deep inspiration.  In addition,  O3 has been shown to result hi airway hyperresponsiveness
23     as demonstrated  by an increased physiological response to a nonspecific stimulus.  The
24     decrease in inspiratory capacity results in a decrease in forced vital capacity (FVC) and total
25     lung capacity (TLC)  and in combination with bronchoconstriction contributes to a decrease  in
26     the forced expiratory volume in one second (EEVj).  However, it is important to stress that
27     in many of the studies reporting the effects of O3 concentrations critical to the standard-
28      setting process (i.e.,  <0.30 ppm),  the observed decrements in FEV! to a large extent reflect
 29      decrements in FVC of a similar magnitude (i.e., a decreased inspiratory capacity) and, to a
 30      lesser extent, increases in central and/or peripheral airway resistance (RaW).
        December 1993                           7-2       DRAFT-DO NOT QUOTE OR CITE

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 1          The majority of controlled human studies have been concerned with the effects of
 2     various O3 concentrations in healthy subjects performing continuous exercise (CE) or
 3     intermittent exercise (IE) for variable periods of time.  Controlled human exposure studies of
 4     this type have provided the strongest and most quantifiable concentration-response data on
 5     the health effects of O3. As a result of these studies, a large body of data regarding the
 6     interaction of O3 concentration,  minute ventilation, and duration of exposure is available.
 7     The most salient observations  from these studies are  (1) O3 concentration is more important
 8     than either ventilation or exposure duration in determining pulmonary responses;  and
 9     (2) normal healthy subjects exposed to O3 concentrations  <0.12 ppm (the level of the
10     current NAAQS) develop significant reversible, transient decrements in pulmonary function if
11     ventilation and/or duration of  exposure are increased sufficiently.  There is typically a large
12     intersubject varability in physiologic and symptomatic responses to O3;  however, with most
13     individuals these responses tend to be reproducible.  The relationship among response
14     variables such as spirometry,  resistance measurements,  symptoms, and  nonspecific bronchial
15     responsiveness is yet to be fully determined, but the generally weak associations suggest that
16     several response mechanisms  may be operant.   In addition, a growing  number of studies  are
17     beginning to provide insight into the relationship between regional dosimetry (see Chapter 8),
18     mechanisms of pulmonary responses elicited by acute O3 exposure, and tissue level events
19     within the airways.  This type of information promises to provide further insight into the
20     health effects relevance of O3-induced pulmonary  responses with regard to determining which
21     individuals are at greatest risk from ambient O3 exposure.
22          In this section, the effects  of acute single 1-  to 3-h O3 exposures on pulmonary
23     function in healthy subjects are  examined by reviewing studies that investigate (1) the
24     O3 dose-response relationship; (2) intersubject variability, individual sensitivity, and the
25     association between responses; and (3) mechanisms of pulmonary function responses  and the
26     relationship between tissue-level events and functional responses.  Recent single O3 exposure
27     studies of greater than 3 h duration are reviewed in Section 7.2.2. These single-exposure,
28     long-duration studies are beginning to provide important insights  into the O3 concentration X
29     ventilation X time interaction and the minimum O3 concentration required to elicit a
30     significant pulmonary response.  Important key studies of less than 3 h duration that have
31     contributed to the dose-response database and other studies that have contributed  to a better

       December 1993                            7.3       DRAFT-DO NOT QUOTE OR CITE

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 1      understanding of O3-induced pulmonary responses in healthy individuals are summarized in
 2      Table 7-1.  Table 7-1 summarizes several studies reviewed in the previous air quality criteria
 3      document (U.S. Environmental Protection Agency, 1986) and its supplement (U.S.
 4      Environmental Protection Agency, 1992), as well as  studies published since completion of
 5      these earlier documents.  Not reviewed in this section are studies that examine changes in
 6      airway responsiveness induced by O3 inhalation  (see  Section 7.2.3).
 7
 8      The Ozone Concentration-Response Relationship
 9           At-Rest Exposures. No new studies examining the acute effects of a single exposure to
10      O3 concentrations below 1 ppm in resting humans have been published since the 1986  U.S.
11      Environmental Protection Agency (EPA) criteria document (U.S. Environmental Protection
12      Agency, 1986).  Seven studies (Young et al.,  1964;  Bates et al., 1972; Silverman et al.,
13     1976; Folinsbee et al., 1978;  Horvath et al., 1979; Kagawa  and Tsuru, 1979; Konig et al.,
14     1980) examining 2-h at-rest exposures were discussed in the 1986 EPA criteria  document
15     (U.S. Environmental Protection Agency,  1986)  involving 91 healthy subjects (74 males,
16     17 females) exposed to  O3  concentrations ranging from 0.10 to 1.00 ppm.  The lowest
17     concentration at which significant reductions in  FVC and ¥EVl were reported was 0.50 ppm
18     (Folinsbee et al., 1978; Horvath et al., 1979).  Reports of increases in airway resistance are
19     inconsistent in resting human subjects exposed to O3 concentrations below 1.00 ppm.
20
21           Exposure  with Exercise.  Bates et al. (1972) and Hazucha et al. (1973) were the first
 22      investigators to  examine the effect of increasing ventilation via exercise during  O3 inhalation
 23      on pulmonary function  responses. The intermittent  exercise protocol used consisted of the
 24      subjects alternating rest and light exercise on a  cycle ergometer at a rate sufficient to  double
 25      resting minute ventilation for 15 min during a period of 2 h.
 26          Hazucha et al. (1973) observed significant decreases in forced expiratory  endpoints at
 27     0.37 ppm O3 (p < 0.05) and 0.75 ppm O3 (p  < 0.001), with subjects exposed to 0.75 ppm
 28     O3 having the greatest  decrements.  After exposures, all subjects complained to varying
 29     degrees of substernal soreness, chest tightness,  and  cough.  The important findings from
         December 1993                           7-4       DRAFT-DO NOT QUOTE OR CITE

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TABLE 7-1. CONTROLLED HUMAN EXPOSURE TO OZONE3
r:
n>
1
5
u>
•^J
U)
O
§
1
I
3
g
Ozone
Concentration
ppm
Hg/m
Exposure
Duration and Exposure
Activity Conditions
Number
and
Gender of
Subjects
Subjects
Characteristics6
Observed Effect(s)
References
Healthy Adult Subjects at Rest
0.00
0.25
0.50
0.75
0.00
0.37
0.50
0.75
0.00
0.50
Healthy
0.00
0.08
0.10
0.12
0.14
0.16
0.00
0.10
0.15
0.20
0.25
0.00
0.12
0.18
0.24
0.00
0.12
0.18
0.24
0.30
0.40

0
490
980
1,470
0
726
980
1,470
0
980
2h NA
2h NA
2h NA
8M
5 F
20 M
8F
40 M

Young, healthy
adults

FVC decreased with 0.50- and 0.75-ppm 03 exposure compared
with FA; 4% nonsignificant decrease in mean VO2inax following
0.75 ppm 03 compared with FA exposure.
Decrease in FEVj, ^2S%VC' an(^ ^50%VC w*"1 0-75 ppm
03 exposure compared to FA.
Decrease in forced expiratory volume and flow.
Horvath et al. (1979)
Silverman et al. (1976)
Folinsbee et al. (1978)
Exercising Adult Subjects
0
157
196
235
274
314
0
196
294
392
490
0
235
353
470
0
235
353
470
588
784

2hIE Tdb = 32 ;C
(4xl5minat RH = 38%
VE =
68 L/min)
2 h IE Tdb = 22 °C
(4 x 14min RH = 50%
treadmill at
mean Vg =
70.2 L/min)
1 h competitive Tdb = 23-26 °C
simulation RH = 45-60%
exposures at
mean VE =
87 L/min
2.5 h IE Tdb = 22 CC
(4xl5min RH = 40*
treadmill
exercise
[VE =
65 L/min])

24 M
20 M
10 M
20 M
22M
20 M
21 M
20 M
29 M

Young, healthy
adults
Young, healthy
NS
10 highly
trained
competitive
cyclists
Young, healthy
adults

No significant changes in pulmonary function measurements.
FVC, FEVj, FEF25.755J, SGaw, 1C, and TLC all decreased with
(1) increasing 03 concentration, and (2) increasing time of
exposure; threshold for response was above 0. 10 ppm but below
0.15 ppm 03.
Decrease in FVC and FEV1-0 for 0.18- and 0.24-ppm
O3 exposure compared with FA exposure; decrease in exercise
time for subjects unable to complete the competitive simulation at
0.18 and 0.24 ppm 03, respectively.
Significant decrease in FVC, FEVj , and FEF25_75% at 0.12 ppm
63; decrease in VT and increase in f and SR,W at 0.24 ppm 03.

Linn et al. (1986)
Kulle et al. (1985)
Schelegle and Adams (1986)
McDonnell et al. (1983)

-------
TABLE 7-1 (cont'd). CONTROLLED HUMAN EXPOSURE TO OZONE3
0
1
cr
n
—
i














-j
ON




O
C


§'

M
O
H
0

Q
M
O
O
Ozone
Concentration

ppm
Healthy
0.12
0.18
0.24
0.30
0.40







0.12
0.18
0.24
0.30
0.40


0.00
0.12
0.18
0.24
0.30
0.40






0.00
0.12
0.20


fig/m
Exposure
Duration and Exposure
Activity Conditions
Number
and
Gender of
Subjects

Subjects
Characteristics6 Observed Effect(s)


References
Exercising Adult Subjects (cont'd)
235
353
470
588
784







235
353
470
588
784


0
235
353
470
588
784






0
235
392

2 x 2.5 h IE Tdb = 22 °C
(4 x 15 min RH = 40%
treadmill
exercise
[ VE =
35 L/min/m
BSA]).
Exposure
separated by
48 ± 30 days
and 301 ±
77 days
2 x 2.5 h IE Tdb = 22 °C
(4 X 15 min RH = 40%
treadmill
exercise
[ ^P ~
35 L/min/m2
BSA])
2.5 h IE Tdb = 22 °C
(4 X 15 min RH = 40%
treadmill
exercise
[ VE =
25 L/min/m2
BSA])





1 h CE (mean Tdb = 31 °C
VE = 89
L/min)

8M
8M
5M
5M
6M







290 M






17 WM/15 BM/15 WF/15
BF
15 WM/15 BM/15 WF/16
BF
15 WM/17BM/17 WF/15
BF
16 WM/15 BM/17 WF/16
BF
15 WM/15 BM/15 WF/15
BF
15 WM/15 BM/15 WF/15
BF
15 M
2F


Young, healthy Pulmonary function variables SRaw and Vg were not significantly
adults different in repeat exposures, indicating that the response to
0.18 ppm 03 or higher is reproducible.









Young, healthy 03 concentration and age predicted FEVj decrements; it was
adults concluded that age is a significant predictor of response (older
subjects being less responsive to 03).




Young, healthy Decreases in FEVj for all levels of 03 as compared with FA;
whites and increase in SRaw with 0.18 ppm O3 and greater compared with FA;
blacks black men and women had larger FEV] decrements than white men,
and black men had larger FEVj decrements than white women.








Highly trained Decrease in Vgmax, VO2inax, VTmax, work load, ride time,
competitive FVC, and FEVj with 0.20 ppm Oj exposure, but not significant
cyclists with 0.12 ppm 03 exposure, as compared to FA exposure.

McDonnell et al.
(1985b)










McDonnell et al. (1993)






Seal et al. (1993b)











Gong et al. (1986)

-------
TABLE 7-1 (cont'd). CONTROLLED HUMAN EXPOSURE TO OZONE3
o
I
Ozone
Concentration Exnogure
ppm
Duration and
Hg/m Activity
Exposure
Conditions
Number
and
Gender of
Subjects
Subjects
Characteristics
Observed Effect(s)
References
^O Healthy Exercising Adult Subjects (cont'd)
i
o
6
o
1
i
0.00
0.16
0.24
0.32
0.00
0.20
0.00
0.20
0.30
0.00
0.20
0.35
0.00
0.21
0.00
0.21
0.00
0.25
0
314
470
627
0
392
0
392
588
0
392
686
0
412
0
412
0
490
1 h CE (mean
57 L/min)
4hffi
(4 x 50 min
cycle
ergometry or
treadmill
running [ VE
= 40 L/min])
30-80 min
CE cycle
ergometry
(VE = 33or
66 L/min)
1 h CE or
competitive
simulation
(mean VE =
77.5 L/min)
1 h CE (75%
VCynax)
1 h CE cycle
ergometry
(mean VE =
80 L/min)
IhCE
(mean VE =
63 L/min)
Tdb =
RH =
Tdb =
RH =
Tdb =
•c
RH =
Tdb =
"C
RH =
Tdb =
°C
RH =
Tdb =
RH =
Tdb =
RH =
32 °C
42-46%
20 °C
50%
20-24
40-60%
23-26
45-60%
19-21
60-70%
22.5 °C
58.8%
20 °C
70%
42 M
8F
11 M
3F(FA
exposure);
9M
exposure)
8M
10 M
6M
1 F
14 M
1 F
19 M
7F
Competitive
bicyclists
Adult, healthy
NS
Aerobically fit
Well-trained
distance
runners
Well-trained
cyclists
Highly fit
endurance
cyclists
Active
nonathletes
Small decrements in FEVj at 0.16 ppm with larger decrements at
0.24 ppm 03.
Decrease in FVC, FEVj, VT, and SRaw and increase in f with
Oj exposure compared with FA; total cell count and LDH increased in
isolated left main bronchus lavage and inflammatory cell influx occurred
with 03 exposure compared to FA exposure.
03 effective dose was significantly related to pulmonary function
decrements and exercise ventilatory pattern changes; multiple regression
analysis showed that 03 concentration accounted for the majority of the
pulmonary function variance.
Decrease in FVC, FEVj, and FEF25_75% with 0.20 and 0.35 ppm 03
exposure compared with FA; Vj decreased and f increased with
continuous 50-min 03 exposures; three subjects unable to complete
continuous and competitive protocols at 0.35 ppm 03.
Decrease in FVC, FEVj, FEF25_75%, and MW with 0.21 ppm 03
compared with FA exposure.
No significant differences in the effects of albuterol on metabolic data,
pulmonary function, airway reactivity, and exercise performance vs.
placebo; decrease in VEmax during 03 conditions.
FVC, FEVlt and MW all decreased with 0.25 ppm 03 exposure
compared with FA.
Avol et al. (1984)
Aris et af. (1993a)
Adams et al. (1981)
Adams and Schelegle
(1983)
Folinsbee et al. (1984)
Gong et al. (1988)
Folinsbee et al. (1986)

-------
TABLE 7-1 (cont'd). CONTROLLED HUMAN EXPOSURE TO OZONE3
CT Ozone
5. Concentration
1
VO
VO
U)
-J
00
d
6
o
1
$
ppm
Healthy
0.00
0.25
0.00
0.30
0.00
0.35
0.00
0.37
0.50
0.75
0.00
0.40
0.00
0.40
Hg/m
Exposure
Duration and Exposure
Activity Conditions
Number
and
Gender of Subjects
Subjects Characteristics
Observed Effect(s)
References
Exercising Adult Subjects (cont'd)
0
490
0
588
0
686
0
726
980
1,470
0
784
0
784
1 h CE NA
cycle
ergometer
(VE= 2
30 L/min/m
BSA)
1 h CE cycle NA
ergometry
(mean Vg =
60L/min)
1 h CE cycle Tdb = 21-25
ergometry °C
(mean VE = RH = 45-60%
60L/min)
2 h IE cycle
ergometry
(VE = 2.5x
rest)
2 h IE Tdb = 22 °C
treadmill RH = 40%
exercise
( VE = 50 to
75 L/min)
1 h CE NA
treadmill
exercise;
( VE =
20 L/min/m2
BSA)
5 M Young, healthy
2F NS
5 M Normal
14 M Moderately fit,
young, healthy
adults
20 M Young, healthy
8 F adults
8 M Young, healthy
NS
20 M Young, healthy
NS
12 A % decrease in FEVj . Significant elevation of sub stance P and
8-epi-PGFj in segmental airway washing, but not bronchoalveolar
lavage fluid.
Decrease in FVC and FEVj, and increase in SR,,W 1 h post-Oj
exposure; increase in percent neutrophils at 1, 6, and 24 h post-Oj
exposure compared with FA in first aliquot "bronchial" sample.
Neutrophils peaked at 6 h post-Oj in "bronchial" sample. Percent
neutrophils elevated at 6 and 24 h post-Oj in pooled aliquots.
Significant decreases in FVC and FEVj with 03 exposure compared to
FA exposure; FVC and FEVj decreases with 03 exposure were
significantly attenuated with indomethacin compared to no drug and
placebo; SRgW increases were not effected by indomethacin.
Decrease in FVC with 0.50 ppm and FEVj with 0.50 and 0.75 ppm
03 compared to FA; decrease in ^25% VC ""^ "-37 and 0.75 and
VjQ^vc with 0.37, 0.50, and 0.75 ppm Oj exposure compared to
FA.
Decreases in FVC, FEVj, VT, and TLC, and increases in SR,,W and f
with 03 exposure compared with FA. Atropine pretreatment abolished
O3-induced increase in SRaw and attenuated FEVj and Ff-F25-7S%
response.
VT fell by 25 % , and O3 uptake efficiency in the lower respiratory
tract fell by 9% during 03 exposure.
Hazbunetal. (1993)
Schelegleetal. (1991)
Schelegleet al. (1987)
Silverman et al. (1976)
Beckett et al. (1985)
Gerrity et al. (1993a)

-------
TABLE 7-1 (cont'd). CONTROLLED HUMAN EXPOSURE TO OZONE8
g Ozone
g. Concentration11 Exoosure
Cl>
t£ PPm
Duration and
fig/m Activity
Exposure
Conditions
Number
and
Gender of Subjects
Subjects Characteristics0 Observed Effect(s)
References
£O Healthy Exercising Adult Subjects (cont'd)
0.00
0.40
0 2hIE
784 (4 x 15 min
heavy treadmill
exercise
NA
1 1 M Young, healthy No correlation between pulmonary function and inflammatory endpoints
NS measured in BAL fluid obtained 18 h after exposure; increase in percentage of
PMNs, total protein, albumin, IgG, and neutrophil elastase; decrease in
percentage of macrophages with Oj exposure compared to FA exposure.
Koren et al. (1989a)
35 L/min/m
BSA])
0.00 0
0.40 784
O 0.00 0
vb 0.40 784
0
j> 0.00 0
2 0.40 784
i
5 0.00 0
^ 0.40 784
.- 0.60 1,176
1
a
g
2 h IE Tdb = 22 °C
(4 x 15 min RH = 40%
heavy treadmill
exercise
35 L/min/m
BSA])
2 h IE Tdb = 22 °C
(4 X 15 min RH = 50%
bicycle
ergometry
[VE =
30 L/min/m
BSA])
1 h CE Tdb = 22 °C
(treadmill RH = 40%
exercise; Vc =
20 L/min/m
BSA)
2hIE Tdb = 71.5 "C
(4 x 15 min RH = 55%
cycle ergometry
at 100 W for
males and 83 W
for females)

10 M Young, healthy PMN, PGE2, and IL-6 were higher in BAL fluid obtained 1 h post-Oj exposure
NS than 18 h; fibronectin and urokinase-type plasminogen activator were higher 18
h post-Oj exposure than 1 h.
13 M NS Indomethacin pretreatment and Oj exposure resulted in a significantly smaller
decrease in FVC and FEVj than 03 exposure alone; airway hyperresponsiveness
was not significantly affected by indomethacin pretreatment.
22 M Young, healthy Significant decreases in FVC, FEV,, FEVj/FVC, and FEF25_75%. The half-
NS width of an expired aerosol bolus was significantly increased, suggesting an
ozone-induced change in small airway function.
7 M Healthy Increase in airway responsiveness to methacholine challenge, in mean percentage
3 F NS of neutrophils, and in PGF2a, TXB2, and PGE2 concentrations measured in
BALF 3 h after 0.40- and 0.60-ppm 03 exposure compared with FA exposure.

Koren et al. (1991)
Ying et al. (1990)
Keefe et al. (1991)
Seltzer et al. (1986)

-------
                               TABLE 7-1 (cont'd).  CONTROLLED HUMAN EXPOSURE TO OZONE3
o>
u>
Ozone
Concentration1" Exoosure
Duration and Exposure
ppm fig/m Activity Conditions
Number
and
Gender of Subjects
Subjects Characteristics0 Observed Effect(s)
References
Healthy Exercising Adult Subjects (cont'd)
0.00 0 2 h IE Tdb = 21 °C
0.50 980 (4 x 15 min RH = 40%
treadmill
exercise;
40L/min)
0.00 0 2 h IE NA
0.75 1,470 (4 x 15 min
light [50 W]
cycle
ergometry)
1 8 M Healthy, young Decrease in VC, VT, and maximal transpulmonary pressure, and increase
adults in SR^ and f with O3 exposure compared to FA exposure; lidocaine
inhalation partially reversed the decrease in VC.
13 M 4 light S, Decrease in FVC, FEV^ ERV, 1C, and FEF50% after 1 h exposure to
9 NS 0.75 ppm 03; decrease in VO2max, VTmax, VEmax, maximal
workload, and heart rate following 0.75-ppm 03 exposure compared with
FA.
Hazucha et al. (1989)
Folinsbeeet al. (1977b)
    aSee glossary of terms and symbols for abbreviations and acronyms.
     Grouped by rest and exercise; within groups listed from lowest to highest 03 concentration.
     Age range in years or as mean + SEM.

-------
 1     these early studies were that the exercise-induced increase in minute ventilation accentuated
 2     the observed pulmonary response at any given O3 concentration and lowered the minimum
 3     O3 concentration at which significant pulmonary responses were observed.  Subsequently, the
 4     interaction between O3 concentration and minute ventilation was examined by using similar
 5     intermittent exercise protocols in which both O3 concentration and level of minute ventilation
 6     were varied.
 7           Silverman et al.  (1976)  and  Folinsbee et al. (1975) exposed a group of 20 males and
 8     8 females to 0.37, 0.50, or 0.75 ppm for 2 h while resting or exercising intermittently.  The
 9     IE protocol used alternated 15 min of rest with 15 min of exercise, sufficient to increase the
10     minute ventilation (VE) by a factor of 2.5 over the value at rest.  The submaximal exercise
11     responses of the subjects were tested postexposure using a three-stage cycle  ergometer test,
12     with loads adjusted to 45, 60, and 75% of maximum oxygen uptake (VO2max) (Folinsbee
13     et al., 1975). Pulmonary function responses were related to the total inhaled dose or the
14     "effective dose" of O3 calculated as the product of concentration, exposure time, and
15     ventilation.  Neither submaximal exercise  oxygen uptake (VO2) nor VE were significantly
16     affected by any level of O3 exposure. However, a significant increase in respiratory
17     frequency (f), and a significant decrease in tidal volume  (VT) at the 75% VO2max workload,
18     was observed.  The relationship between the effective dose of O3 and the mean percent
19     change in selected measures of lung function was analyzed using linear regression.  Forced
20     vital capacity, maximum expiratory flow at 25 and 50%  of FVC (Vmax25% and Vmax50%.
21     respectively), and  FEVj were found to have a significant linear correlation with the effective
22     dose.  The description of the relationship between O3 pulmonary function decrements and
23     effective dose was apparently improved by the use of a second-order polynomial model  in
24     which effective dose was used as  the independent variable.
25           Though the investigations of Silverman et al.  (1976) and others (Bates et al., 1972;
26     Hackney et al., 1975; Hazucha et al., 1973) clearly demonstrate {he potentiating effects  of
27     exercise on O3 responses, the level of exercise used in these studj|$5 was  low, requiring
28     increases in VE of only 2 to 2.5 times resting, a level of exercisp lower than walking at
29     5.5 km/h (DeLucJa and Adams, 1977).  In order to address this concern, DeLucia and
30     Adams (1977) exposed six healthy nonsmoking male  subjects on 12 separate occasions to
31     FA, 0.15, and  0.30 ppm O3 for 1 h, while at rest,  and while exercising continuously at

       December 1993                          7_n      DRAFT-DO NOT QUOTE OR  CITE

-------
 1     workloads that required 25, 45, and 65% of the subjects' VO2max. They observed a
 2     significant time-dependent increase in f during the 65%  VO2max, 0.30-ppm O3 exposure and
 3     a significant decrease in FEVj and forced expiratory flow at 25 to 75 % of FVC (FEF25_75%)
 4     immediately following this same exposure.
 5          These initial studies, which clearly demonstrated the potentiating effects of exercise on
 6     human responses to acute O3 exposure, provided the impetus for a series of studies (Adams
 7     et al.,  1981; Folinsbee et al., 1978; McDonnell et al.,  1983; Kulle et al., 1985; Linn et al.,
 8     1986) designed to more accurately define O3 dose-response relationships.   These
 9     investigations utilized both IE (Folinsbee et al., 1978; McDonnell et al., 1983) and CE
10     (Adams et al., 1981) of varying intensity.  Folinsbee et al.  (1978) exposed four groups of
11     10 subjects each to FA,  0.10, 0.30, and 0.50 ppm O3 for 2 h.  One group was exposed
12     while at rest and the other three groups were exposed while performing IE at levels requiring
13     a ventilation of 30, 50, or 70 L/min.  These combinations of ventilation and O3 concentration
14     resulted in a range of total inhaled dose of 0.00 to 4.41 mg O3.  Adams et al.  (1981)
15     exposed eight trained male subjects to FA, 0.20, 0.30, and 0.40 ppm O3  while exercising
16     continuously at two different workloads (35 and 62% of VO2max) for durations ranging
17     from 30 to 80 min.  Each subject completed all 18 protocols with at least 3 days between
18     each.  The findings from these two studies confirmed that significant pulmonary responses
19     occurred at 0.30 ppm when subjects exercised  at moderately heavy workloads.  It was further
20     demonstrated, by multiple regression analysis,  that the O3 effective dose was a better
21     predictor of response than O3 concentration, VE, or duration of exposure, alone.  Multiple
22     regression analysis also revealed that the majority of variance for pulmonary function
23     responses was accounted for by O3 concentration, followed by VE. In the Adams et al.
24      (1981) study, in which both workload and duration of exposure were varied, duration of
25      exposure  was observed  to be the poorest predictor of response for all parameters analyzed.
26      However, the minor impact of changes in exposure  duration could have been an artifact of
27      the limited combinations of ventilation and durations of exposure used by these investigators.
28           McDonnell et al. (1983) conducted  a study with  the primary purpose of discerning the
 29      lowest concentration of O3 at which group mean decrements in pulmonary function occur  in
 30     heavily exercising healthy men.  In order to determine a concentration-response relationship,
 31      six groups of subjects (n = 20 to 29) were  exposed to either an FA control or one of five

        December 1993                           7-12       DRAFT-DO NOT QUOTE OR CITE

-------
 1     03 concentrations (0.12, 0.18, 0.24, 0.30, or 0.40 ppm) at a VE of 67 L/min and exposure
 2     duration of 2.5 h (15-min rest, 15-min exercise.  These investigators observed small
 3     significant changes in FVC, FEVb FEF25_75%, and cough at 0.12 ppm O3 and concentration-
 4     dependent responses in all variables measured  (FVC, FEV^ FEF25_75%, specific airway
 5     resistance [SR^], f, VT, and subjective symptoms) at O3 concentrations > 0.24 ppm.
 6          Kulle et al. (1985) also conducted a similar study on healthy, nonsmoking men
 7     performing intermittent exercise at a VE of 70 L/min for an exposure duration of 2 h,
 8     (16-min rest, 14-min exercise).  Twenty subjects were exposed to an FA control or one of
 9     four O3 concentrations  (0.10, 0.15, 0.20, or 0.25 ppm). These investigators observed a
10     significant concentration x time interaction at  0.15 ppm O3 for FVC, FEVls FEF25_75%, and
11     in all variables measured (FVC, FEVj, FEF25_75%, SR^, f, VT, and subjective symptoms)
12     at O3 concentrations greater than 0.15  ppm.
13          Linn et al. (1986) exposed 24 healthy, well-conditioned male subjects (18 to 33  years of
14     age) for 2 h to 0.00, 0.08, 0.10, 0.12, 0.14, or 0.16 ppm O3, using  an intermittent exercise
15     protocol (15-min rest, 15-min exercise; VE =  68 L/min) combined with an ambient heat
16     stress (32 °C and 38%  relative humidity).  They observed no statistically significant changes
17     in forced expiratory endpoints and symptoms after exposure to  O3 concentrations from
18     0.08 to 0.14 ppm.  These authors observed a small (-2.3%) but significant (p  < 0.05)
19     reduction in FEV, following the 2-h 0.16-ppm O3 exposure that was not associated with
20     symptoms of respiratory discomfort.
21          More recently, Seal et al. (1993b) examined whether gender or race differences in
22     responsiveness to O3 exist.  The authors exposed 372 white and black males and females
23     (n > 90 in each gender-race group) once for 2.33 h to 0.0, 0.12, 0.18, 0.24, 0.30, or
24     0.40 ppm O3 using an intermittent exercise protocol (15-min rest, 15-min exercise;
        *                9
25     VE = 25 L/min/m  BSA). Statistical  analysis (nonparametric two-factor analysis of
26     variance) of the percent changes from baseline for FEVj, SRaw, and  cough responses
27     demonstrated no significant differences in responsiveness to  O3  between the race-gender
28     groups studied.  Changes in FEVl5 SR^, and  cough were first noted at 0.12, 0.18, and
29     0.18 ppm O3,  respectively, for the group as a whole. It is difficult to compare  the results
30     from this study with others that have examined the 0$ concentration-response relationship in
31     healthy adult males because the authors did not present a separate analysis of male responses.

       December 1993                          7_13       DRAFT-DO  NOT QUOTE OR CITE

-------
 1     For further evaluation of the influence of gender and race on O3 responsiveness,  see
 2     Section 7.2.1.3.
 3          The observation of significant decrements in pulmonary function in heavily  exercising
 4     healthy subjects at O3 concentrations of 0.20 ppm and lower has been confirmed  by
 5     numerous investigators (Adams and Schelegle, 1983; Avol et al., 1984; Folinsbee et al.,
 6     1984; Gong et al., 1986) who utilized 1-h continuous heavy exercise exposure protocols.
 7     Adams and Schelegle (1983) and Folinsbee et al. (1984) observed significant decrements in
 8     FVC and FEVj in well-trained subjects exposed to 0.20 ppm O3 while exercising with a
 9     VE of approximately 80 L/min. Avol et al. (1984) observed small but significant decrements
10     in FVC and FEVt in a group  of 50 competitive cyclists (42 males, 8 females) exposed to
11     0.16 ppm O3  while exercising with a VE of 57 L/min in combination with added heat stress
12     (32 °C). Similarly, Gong et al. (1986) observed modest but significant decrements in FVC
13     and FEVj in  a group of 17 top-caliber endurance cyclists exposed to 0.12 ppm O3 while
14     exercising at  approximately 70% of their VO2max (mean VE = 89 L/min) with an added
15     heat stress  (32 °C). In addition to the above studies that used continuous exercise, Schelegle
16     and Adams (1986) observed significant reductions in FVC and FEVj  and increased
17     symptoms of respiratory discomfort following exposure to 0.18 ppm O3, but not 0.12 ppm
18     O3, in a group of competitive endurance athletes exposed while  performing a competitive
19     simulation  consisting of a 30-min warm-up followed by a 30-min competitive bout (mean
20     VE over entire protocol = 87 L/min).
21           The results of the studies reviewed above demonstrate that the minimum
22     O3 concentrations between 0.12 to 0.18 ppm are required to elicit significant pulmonary
23     function decrements and increased symptoms in healthy young adult subjects exposed while
24     performing moderate to severe IE and CE of from 1 to 3 h duration.
25           Retrospective analysis by Hazucha (1987) confirmed the previously reported (Adams
26     et al., 1981;  Folinsbee et al., 1978) dominant role that O3 concentration plays in determining
27     O3-induced responses.  Hazucha (1987) analyzed data from studies that utilized intermittent
28     exercise protocols of 2 h in duration.  While controlling for ventilation, he found that the
29      data best fit a model that was a quadratic function of O3 concentration.  Based on his
 30      analysis, Hazucha (1987) also concluded that he could not define an O3 concentration below
 31      which no pulmonary function response would be elicited.

        December 1993                           7-14      DRAFT-DO NOT QUOTE OR CITE

-------
 1     Intersubject Variability, Individual Sensitivity, and the Association Between Responses
 2          Bates et al. (1972) noted that variation in sensitivity and response was evident for
 3     various symptoms and pulmonary functions they assessed following O3 exposure.  This
 4     observation of large intersubject variability in response to O3 has also been reported by
 5     numerous other investigators (Adams et al., 1981; Folinsbee et al., 1978; McDonnell et al.,
 6     1983; Kulle et al.,  1985) and is illustrated by data from Kulle et al. (1985) plotted in
 7     Figure 7-1. The description of the factors that contribute to intersubject variability is
 8     important for the understanding of individual responses, mechanisms of response, and health
 9     risks associated with acute exposures.  The effect of this large intersubject variability on the
10     ability to predict individual responsiveness to O3 was recently demonstrated by McDonnell
11     et al. (1993).  These investigators analyzed the data of 290 white male subjects (18 to
12     32 years of age) who inhaled either 0.00, 0.12, 0.18, 0.24, 0.30, or 0.40 ppm O3 for
13     2  h while performing an IE protocol (VE = 35 L/min/m2 BSA) to identify personal
14     characteristics (i.e., age, height, baseline pulmonary functions, presence of allergies, and
15     past smoking history) that might predict individual differences in FEVj response.  Of the
16     personal characteristics  studied,  only age contributed significantly to intersubject
17     responsiveness (younger subjects being more responsive), accounting for 4%  of the observed
18     variance.   Interestingly, O3  concentration accounted for only 31 % of the variance, clearly
19     demonstrating the importance of as yet undefined intrinsic individual characteristics that
20     determine  responsiveness  to O3.
21          McDonnell et al. (1985b) examined the reproducibility of individual responses to
22     O3 exposure in healthy  human subjects exposed twice, with from 21 to 385 days separating
23     exposures  (mean = 88 days). This investigation was conducted in order to determine
24     whether the observed intersubject variability is primarily due to real differences in
25     O3 responsiveness among subjects, or  whether it can be accounted for by other sources of
26     variability. The authors examined  the reproducibility of FVC, FEV], FEF25.75%,  SR^,
27     cough, shortness of breath (SB), pain on deep inspiration (PDI), VT, and f responses induced
28     by O3 exposure to concentrations ranging from 0.12 to 0.40 ppm.  Reproducibility was
29     assessed using the intraclass correlation coefficient (R), which directly measures the
30     reproducibility of responses by incorporating into a single measure all the information
31     contained in the correlation coefficient, slope, and intercept obtained in linear

       December 1993                           7.15       DRAFT-DO NOT QUOTE OR CITE

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                      22-
                      20-
                      18
                      16 -|
                      J4
                      12-1
                    k*H
                    C
                    'c 10
                    o
                    1  M
                    *
                    DC  6-
                        4 _

                        2-
                        0-
                       -2-
                                    0.10       0.15        0.20
                                             Ozone (ppm)
                      0.25
      Figure 7-1.  Individual dose response curves for five separate subjects exposed to 0.10,
                  0.15, 0.20, and 0.25 ppm O3 for 2 h with moderate intermittent exercise.
                  Illustrates the wide variability in responsiveness to O3 from individual to
                  individual.
      Source:  Kulle et al. (1985).
1     regression analysis.  Similar to the more routinely used correlation coefficient, R is equal to
2     one when two identical measurements occur in the same subject; and the "worst" possible
3     coefficient is equal to l/(n — 1), which approaches zero for a large n.  The ranking of most
4     to least reproducible for the responses studied was FVC (R = 0.92), FEV^ (R  = 0.91),
5     FEF25.75%  (R = 0.83), cough (R = 0.77), SB (R = 0.60), SRaw (R = 0.54),  PDI
6     (R = 0.37), f (R = -0.20), and VT (R = -0.03).  The value of R was significantly
7     different from zero for FVC, FEV,, FEF25_75%, cough, SB,  and SR^.  McDonnell et al.
8     (1985b) concluded that the reported large intersubject variability in magnitude of response
       December 1993
7-16
DRAFT-DO NOT QUOTE OR CITE

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 1     was due to large differences in the intrinsic responsiveness of individual subjects to
 2     O3 exposure. However, the factors that contribute to this large intersubject variability
 3     remain undefined.
 4          The examination of intersubject variability is complicated by a poor association between
 5     the various O3 responses.  In their study investigating O3 dose-response relationships,
 6     McDonnell et al. (1983) observed very low correlation between changes in SRaw and FVC
 7     (r = -0.16) for 135 subjects exposed to O3 concentrations ranging from 0.12 to 0.40 ppm
 8     for 2.5 h.
 9
10     Mechanisms of Acute Pulmonary Responses
11          The pulmonary responses observed during and following acute exposure to O3 at
12     concentrations between 0.10 and 0.50 ppm in normal healthy human subjects include
13     decreases in TLC, 1C, FVC, FEVl5 FEF25_75%,  and VT; and increases in SR^, f, and
14     airway responsiveness. Ozone exposure has also been shown to result in the symptoms of
15     cough, PDI, SB, throat irritation, and wheezing. When viewed as a whole, changes in these
16     specific parameters can be categorized into four general  responses, including alterations in
17     lung volumes, airway caliber, bronchomotor responsiveness, and symptoms.  The absence of
18     consistent associations between the various responses from individual to individual suggests
19     that the functional responses  observed are the result of multiple interactions within the
20     respiratory tract.  These interactions may be the  result of O3 action on the biochemical,
21     anatomical, and physiological systems of the respiratory tract.  In turn, these factors
22     determine O3 dose distribution and the resulting  cellular and reflex responses.
23          Bates et al. (1972) observed that the most significant decrement in pulmonary  function
24     was the reduction in the transpulmonary pressure at maximal inspiratory volume without a
25     concomitant decrease in static compliance.  This would suggest an inhibition of maximal
26     inspiratory effort after O3 exposure and result in reductions in 1C.  These authors speculated
27     that this inhibition is an early result of stimulation of rapidly adapting pulmonary stretch
28     receptors, or "irritant receptors", located hi the major bronchi. Since  1972, when this initial
29     hypothesis was published, numerous studies have examined the underlying mechanisms
30     leading to the functional responses observed hi human subjects.  These mechanistic  studies
31     have used both animal models and human subjects.  This discussion of mechanisms  will

       December 1993                          747      DRAFT-DO NOT QUOTE OR CITE

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 1     focus on studies that used human subjects, but also will cover those animal studies that have
 2     direct relevance to O3-induced functional responses.
 3          The acute inhalation of ambient concentrations of O3 by healthy human subjects has
 4     been shown to result in a dose-dependent increase in airway resistance  (Folinsbee et al.,
 5     1978; McDonnell et al., 1983; Kulle et al., 1985; Seal et al., 1993b).  This O3-induced
 6     increase in airway resistance has been shown to be poorly correlated with changes in forced
 7     expiratory endpoints (McDonnell et al.,  1983).  Ozone-induced increases in airway resistance
 8     have a rapid onset (Beckett et al., 1985) compared with the gradual development of
 9     decrements in forced expiratory endpoints (Kulle et al., 1985).  Ozone-induced increases in
10     airway resistance also appear to be greater in atopic subjects as a group (Kreit et  al., 1989;
11     McDonnell et al., 1987), although this does not appear to be the case for O3-induced
12     decrements in FVC and symptoms.  Taken together, these observations suggest that different
13     pathways lead to  O3-induced decrements in 1C and to  O3-induced increments in airway
14     resistance.
15           Increases in airway resistance induced by O3 have been shown to be blocked by
16     atropine sulfate pretreatment in human subjects (Beckett et al., 1985; Adams, 1986).  This
17     inhibition suggests that the release of acetylcholine from parasympathetic postganglionic
18     fibers that innervate airway smooth muscle plays a role in this response.  However, the
19     observation that acute O3 inhalation also results in a hyperresponsiveness to methacholine, a
20     cholinergic agent (Holtzman et al.,  1979), suggests the possibility that acute O3 exposure can
21     also increase the sensitivity of airway smooth muscle  to acetylcholine independent of a reflex
22     mechanism involving cholinergic postganglionic nerves. The role that an increase in airway
23     smooth muscle sensitivity to the endogenous release of acetylcholine might play in
24     O3-induced increases in airway resistance has not been studied.
25           Analyses by Colucci (1983) have suggested that the increase in airway resistance is not
 26      as large as would be expected when O3 exposure is combined with moderate to heavy
 27      exercise.  However, the observation that circulating epinephrine levels increase as a function
 28      of the relative workload in exercising human subjects (Galbo, 1983; Warren and Dalton,
 29      1983) suggests that stimulation of airway smooth muscle beta-adrenoreceptors may counteract
 30     airway smooth muscle contraction  induced by O3 exposure.  The observations by Beckett
         December 1993                           7-18      DRAFT-DO NOT QUOTE OR CITE

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 1     et al. (1985) that the beta-agonists abolish O3-induced bronchoconstriction is consistent with
 2     this possibility.
 3          Another question to be addressed with regard to O3-induced increases in airway
 4     resistance is where along the airway (central versus peripheral airways) is the increase in
 5     resistance produced?  Studies of acutely and subchronically exposed animals have
 6     demonstrated tissue damage in the central acinar region (Castleman et al. , 1977; Mellick
 7     et al.,  1977), as well as increases in peripheral resistance and reactivity (Gertner et al.,
 8     1983a,b,c; Beckett et al., 1988).  The observations by Kreit et al.  (1989) and Keefe et al.
 9     (1991) of small significant decreases in the ratio of FEVj/FVC in normal subjects exposed to
10     0.40 ppm O3 suggest that O3 exerts an effect on the small peripheral airways of humans.
11     Keefe et al. (1991) examined this possibility using an inhaled aerosol bolus dispersion
12     technique in 22 healthy,  nonsmoking male subjects exposed to 0.40 ppm O3 for  1 h using a
13     CE protocol ( VE = 20 L/min/m  BSA).   The bolus dispersion technique is not dependent
14     upon vital capacity maneuvers and compares the profile of a bolus of small (0.5 to 1.0
15     aerosol particles injected into the inspired airstream (at a fixed lung volume) with the profile
16     of the bolus during expiration.  Dispersion of the bolus during expiration can be affected by
17     increases in turbulence within the airway, the development of asymmetries in ventilation due
18     to unequal regional time constants within the lung, and an increase in aerosol deposition in
19     the small airways.  Keefe et al. (1991) observed that O3 exposure in their subjects resulted in
20     a significant increase in dispersion of an aerosol bolus (without an increased aerosol
21     deposition) that was not correlated with changes in SRaw.  These findings suggest that
22     exposure to 0.40 ppm O3 under the conditions of this experiment results in changes in small
23     airway function that are not detectable by more conventional  techniques.
24           Ozone-induced alterations in ventilatory pattern have been observed in exercising dogs
25     (Lee et al., 1979) and humans (Adams et al., 1981; Folinsbee et al., 1978; McDonnell et al.,
26     1983; Kulle et al., 1985).  In exercising humans, O3 exposure has been shown to result in a
27     decrease in VT and an increase in f in the absence of any change in VE.  A rapid shallow
28     breathing pattern is consistent with the maintenance of an appropriate ventilation with a
29     reduced tidal volume.  Reduction of tidal volume is probably related to the reduction of
30     inspiratory capacity and is anecdotally related to reduction in breathing discomfort caused by
31     pain on deep inspiration.

       December 1993                           749       DRAFT-DO NOT QUOTE OR CITE

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 1          Lee et al. (1979), who produced a reversible vagotomy by cooling the vagus nerves to
 2     0 °C, abolished the rapid shallow breathing induced by O3 inhalation in conscious dogs.
 3     More recently, Schelegle et al. (1993) have shown in anesthetized dogs exposed to O3 that
 4     cooling the cervical vagus nerves to 7 °C did not abolish the observed O3-induced rapid
 5     shallow breathing pattern and bronchoconstriction, but cooling the vagus nerves to 0 °C did
 6     abolish both the rapid shallow breathing and the bronchoconstriction.  These findings suggest
 7     that O3  stimulates nonmyelinated C fiber afferents arising from the lung, whose conduction is
 8     not totally blocked at 7 °C, but is totally blocked at 0 °C.  This conclusion is consistent with
 9     the findings of Coleridge et al. (1993) that bronchial C fibers are the only receptors that are
10     stimulated directly during O3 inhalation in anesthetized dogs.  If similar bronchial C fibers
11     were stimulated or sensitized in humans exposed to  O3, this could explain the O3-induced
12     rapid shallow breathing observed during exercise, as well as the subjective symptoms
13     associated with taking a deep inspiration.
14          Hazucha et al.  (1989) exposed 11 healthy normal volunteers to FA and 0.50 ppm
15     O3 for 2 h while performing moderate intermittent exercise.  Ozone exposure induced a
16     significant decrement in FVC, which was associated with a marked fall in inspiratory
17     capacity without an increase in residual volume.  Spraying of the upper airway with lidocaine
18     aerosol  by these subjects was immediately followed  by restitution of FVC toward control
19     values.   Hazucha et al. (1989) concluded that O3 inhalation stimulates lidocaine-sensitive
20     tracheal and laryngeal airway receptors, which leads to an involuntary inhibition of full
21     inspiration, a reduction in FVC, and a concomitant  decrease in maximal expiratory  flow rates
22     in humans.
23          The airway  afferents  blocked by lidocaine in the Hazucha et al. (1989) investigation
24     remain  undefined. However, it  seems likely that the lung afferents involved are the same
25     ones that result in O3-induced rapid shallow breathing in dogs (i.e., bronchial C fibers).
26     When stimulated  with exogenous chemicals in animal experiments, bronchial C fibers induce
27     a reflex apnea (Coleridge and Coleridge, 1986).  In dogs,  this reflex apnea involves the
28     inhibition of inspiratory neurons, expiratory neurons,  and y- and a-motoneurons  in the
29      intercostal nerves (Koepchen et al., 1977; Schmidt and Wellhoner, 1970).  Such  a reflex
 30      response in humans  would explain the reflex inhibition of maximal inspiration consequent to
 31      acute O3 exposure.

        December 1993                           7-20       DRAFT-DO NOT QUOTE OR CITE

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 1          Data consistent with an O3-induced stimulation of bronchial C fibers in human subjects
 2     has recently been published by Hazbun et al. (1993). These investigators observed a
 3     significant increase in substance P, the neurotransmitter released from the afferent endings of
 4     bronchial C fiber during excitation, in segmental airway  washings of seven (2  female/5 male)
 5     healthy, nonsmoking subjects after a 1 h CE (VE = 30 L/min/m BSA) exposure to
 6     0.25 ppm O3.  Substance P was not elevated in bronchoalveolar lavage (BAL) fluid after air
 7     exposure. In addition, the segmental airway substance p levels were significantly correlated
 8     (r  = 0.89; p <  0.05) with an elevated airway concentration of 8-epi-prostaglandin F2a, a
 9     marker of oxidative free radical reactions. These results are consistent with (1) an increased
10     release of substance P secondary to an increased discharge of bronchial C fibers induce by
11     O3 inhalation, and/or (2) an O3-induced inhibition of neutral endopeptidase, the enzyme that
12     degrades substance P, within the airways.
13          Lung C fibers have been shown to be stimulated by prostaglandin £2 and other lung
14     autacoids (Coleridge et al., 1978, 1976). Interestingly, Schelegle et al. (1987),
15     Eschenbacher et al. (1989), and Ying et al. (1990)  have  shown that pretreatment with the
16     cyclooxygenase inhibitor indomethacin reduces—and in some subjects totally abolishes—
17     O3-induced pulmonary function decrements in human subjects.  Schelegle et al. (1987)
18     examined whether O3-induced pulmonary function decrements could be inhibited by the
19     prostaglandin synthetase inhibitor indomethacin in healthy human subjects. Fourteen college-
20     age males completed six 1-h exposure protocols consisting of no drug, placebo, and
21     indomethacin (Indocin SR, 75 mg every 12 h for 5 days) pretreatments, with filtered air and
22     O3 (0.35 ppm) exposure within each pretreatment.  Pretreatments were delivered weekly in
23     random order in a double-blind fashion.  Exposures consisted of  1 h exercise on a cycle
24     ergometer with work loads set to elicit a mean VE  of 60 L/min.  Statistical analysis revealed
25     significant (p < 0.05) effects for FVC and FEVj across pretreatment, with no drug versus
26     indomethacin and placebo versus indomethacin comparisons being significant.  These
27     findings  suggest that cyclooxygenase products of arachidonic acid, which are reduced by
28     indomethacin inhibition of cyclooxygenase, play a role in the development of pulmonary
29     function  decrements.  These and similar findings by Eschenbacher et al. (1989) and Ying
30     et al. (1990) suggest that the release of some cyclooxygenase product consequent to
31     O3 inhalation plays a role in O3-induced pulmonary function decrements.  This idea is

       December 1993                          7_2i       DRAFT-DO NOT QUOTE OR CITE

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 1     supported by the findings of Koren et al. (1991), who obtained a positive correlation between
 2     O3-induced pulmonary function decrements and the level of prostaglandin £2 in BAL fluid
 3     collected within 1  h after the end of exposure in human subjects who varied greatly in
 4     O3 responsiveness.
 5          The release of cyclooxygenase products of arachidonic acid from injured airway
 6     epithelium can thus be viewed as a link in a cascade of events, which begins with the initial
 7     reaction of O3  with the tissues and ends with the observed pulmonary function responses.
 8     The apparent components of this chain of events include factors that influence (1) O3 delivery
 9     to the tissue (i.e.,  the inhaled concentration, breathing pattern, and airway geometry);
10     (2) O3 reactions with components in airway surface liquid and/or epithelial cell membranes;
11     (3) local tissue responses, including injury and inflammation; and (4) stimulation of neural
12     afferents (bronchial C fibers) and the resulting reflex responses. More studies need to be
13     conducted to determine how each event in this cascade contributes to the pulmonary
14     responses induced by acute O3 inhalation.
15          The influence that individual responsiveness has on this cascade of events has not been
16     determined; however, recent data suggest that individual O3 responsiveness may feed back
17     and influence the distribution of O3 dose within the lung. Gerrity et al. (1993a) tested the
18     hypothesis that O3-induced rapid shallow breathing helps to limit the dose of O3 reaching the
19     lower respiratory tract.  They found that the degree of O3-induced rapid shallow breathing
20     (25% decrease in  VT) was significantly correlated (p <  0.004) with a decrease in 03 uptake
21     efficiency of the lower respiratory tract measured using a fast responding O3 analyzer.   This
22     observation may explain the recent data of Schelegle et al.  (1991) and Aris et  al. (1993a) that
23     suggest individual responsiveness to O3 as  measured by FEVl decrements may be negatively
24     correlated with the number of neutrophils present in bronchoalveolar lavage samples. These
25     limited findings indicate that more studies are needed that examine the interrelationship
26     between the responsiveness to O3, the distribution of dose within the airway, and resulting
27     airway inflammation.
28
29
        December 1993                            7-22      DRAFT-DO NOT QUOTE OR CITE

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 1     7.2.1.2  Subjects with Preexisting Disease
 2     Introduction
 3          Ten studies (Konig et al., 1980; Linn et al., 1982a; Koenig et al., 1985; Linn et al.,
 4     1983a; Linn et al., 1978; Solic et al., 1982; Kehrl et al., 1985; Superko et al., 1984;
 5     Silverman, 1979; Kulle et al., 1984) examining the pulmonary responses to acute
 6     O3 exposures of less than 3 h in patients with preexisting disease were discussed in the 1986
 7     EPA criteria document (U.S. Environmental  Protection Agency, 1986).  This section
 8     examines the effects of O3 exposure on pulmonary function in subjects with preexisting
 9     disease by reviewing O3 exposure studies that utilized subjects with (1) chronic obstructive
10     pulmonary disease (COPD), (2) asthma, (3) allergic rhinitis, and (4) ischemic heart disease.
11     Due to their important health implications, all of the available studies  are reviewed and
12     summarized in Table 7-2.
13
14     Subjects with Chronic Obstructive Pulmonary Disease
15          In five of the studies cited above, the C^-induced pulmonary  function responses of
16     patients with mild to moderate COPD were examined (Konig et al., 1980; Linn et al., 1982a;
17     Linn et al., 1983a; Solic et al., 1982; Kehrl  et al., 1985).  No significant changes in
18     pulmonary function or symptoms were  reported  in any of the studies of the effects of O3 in
19     patients with COPD.  Four of these studies (Linn et al.,  1982a; Linn  et al.,  1983a; Solic
20     et al.,  1982; Kehrl et al., 1985) examined the effects of O3 concentrations between 0.10 and
21     0.30 ppm O3 in 66 mild to moderate COPD  patients using mild IE exposure protocols of 1  to
22     2 h duration.  The total exercise time in all three of these studies was 30 min with intensity
23     being  variable (exercise VE approximately 14 to 28 L/min).  Linn et  al.  (1982a) observed a
24     small but significant reduction in arterial oxygen saturation in 25 mild to moderate COPD
25     patients at the end of the 0.12 ppm O3  exposure for 1 h (absolute  mean difference = 1.3%,
26     p < 0.05). Similarly, Solic et al.  (1982) observed a small reduction  in arterial oxygen
27     saturation in 13 mild to moderate COPD patients at the end of a 0.20-ppm O3 exposure for
28     2 h (absolute mean difference = 0.48%, p < 0.008).  In contrast, Kehrl  et al.  (1985) did
29     not find a significant effect on arterial oxygen saturation  in 13 mild to moderate COPD
30     patients after exposure to 0.30 ppm O3 using the same IE exposure protocol used by Solic
31     et al. (1982).  Similarly, Linn et al. (1983a)  found no significant effect on arterial oxygen

       December 1993                           7-23       DRAFT-DO NOT QUOTE OR CITE

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TABLE 7-2.  OZONE EXPOSURE IN SUBJECTS WITH PREEXISTING DISEASE8
2 Ozone
§• Concentration ExpOsure
1
g
i
o
d
ppm
Duration and Exposure
ftg/m Activity Condition
Subjects with
0.00
0.12
0.00
0.18
0.25
0.00
0.20
0.00
0.30
0.00
0.41
0
236
0
353
490
0
392
0
588
0
804
Chronic Obstructive Pulmonary Disease
1 h IE Tdb = 25 "C
(2 x 15 min RH = 50%
light bicycle
ergometry)
1 h IE Tdb = 25 °C
(2 x 15 min RH = 50%
light bicycle
ergometry)
2 h IE Tdb = 22 °C
(4 x 7.5 min RH = 40%
light
treadmill
running)
2 h IE Tdb = 22 °C
(4 x 7.5 min RH = 40%
light
treadmill
running)
3 h daily Tdb = 22 °C
(1 X 15 min RH = 50%
light bicycle
ergometry
during each
exposure) for
5 days
Number
and
Gender of Subjects
Subjects Characteristics

18 M 8 smokers,
7 F 14 ex-smokers,
3 nonsmokers;
FEV^FVC =
32-66%
15 M 15 smokers,
13 F 11 ex-smokers,
2 nonsmokers;
FEVj/FVC =
36-75%
13 M 8 smokers,
4 ex-smokers,
1 nonsmoker;
productive
cough;
FEVj/FVC =
46-70%
13 M 9 smokers,
4 nonsmokers;
FEV^FVC =
37-65%
17 M All smokers;
3 F productive
cough;
FEVj/FVC =
56-82% and/or
FEV3/FVC =
75-93%
Observed Effect(s)

No significant changes in pulmonary function measurements;
small significant decrease in arterial O^ saturation.
No significant changes in pulmonary function measurements;
no significant change in arterial O^ saturation.
No significant changes in pulmonary function measurements;
small significant decrease in arterial 0% saturation.
No significant changes in pulmonary function measurements or
arterial 02 saturation.
Decrease in FVC and FEVj with 0.41 ppm 03 compared with
FA exposure.
References

Linn et al. (1982a)
Linn et al. (1983a)
Solic et al. (1982)
Kehrl et al. (1985)
Kulle et al. (1984)
Subjects with Heart Disease
0.00
0.20
0.30
0
392
588
40 min CE NA
treadmill
walking
6 M Coronary heart
disease with
angina pectoris
threshold
No significant changes in pulmonary function measurements,
exercise ventilatory pattern, oxygen uptake, or cardiovascular
parameters.
Superko et al. (1984)

-------
TABLE 7-2 (cont'd). OZONE EXPOSURE IN SUBJECTS WITH PREEXISTING DISEASE3
o
§
sr
8
t— *
VO
vO
U)










Ozone
h
Concentration"

ppm

Aig/m

Exposure
Duration and
Activity
Number
and
Exposure Gender of Subjects
Conditions Subjects Characteristics



Observed Effect(s)



References
Subjects with Allergic Rhinitis
0.00
0.18

0.00
0.50




0
353

0
980




2hffi
(4 X 15 min)

4h rest





NA 26 M History of
allergic rhinitis

Tdb = 20-24 °C 6 M History of
RH = 40-48% 6 F seasonal allergic
rhinitis; acute
response to nasal
challenge with
antigen
Increased respiratory symptoms, SRj,,,, and reactivity to histamine
with Oj exposure and decreased FVC, FEVj, and FEF25_75%
with Oj exposure compared to FA.
Increase in upper and lower respiratory symptom scores, cell
influx, epithelial cells with O3 exposure compared to FA; no
effect on acute allergic response to nasal antigen challenge
between 03 and FA exposure.


McDonnell et al.
(1987)

Bascom et al. (1990)





Adult Subjects with Asthma




-jj
N)
<-^i


O
§>
M
I
O
o
25
0
H
O
c
o
m

i
0.00
0.10
0.25



0.00
0.12



0.00
0.12











0
196
490



0
236



0
236











2 h light IE
(2 x 15 min
on treadmill,
vE =
10-20 L/min)

1 h rest




0.75 h m
vE =
30 L/min
(15 min rest,
15 min
exercise,
15 min rest)
followed by
15 min
exercise
inhaling
0.10 ppm
SO2
Tdb = 21 ° C; 7 M Stable mild
RH = 40% asthmatics with
FEVj > 70%
and
methacholine
responsiveness
NA 7 M, 8 F Never smoked,
asthmatics mild stable
asthmatics with
exercise-induced
asthma
Tdb = 22° C; 8 M, 5 F Asthmatics
RH = 75 % asthmatics, ages classified on
12 to 18 years basis of positive
clinical history
and
methacholine
challenge.
Asymptomatic at
time of study.




No significant differences in FEVj observed for Oj-filtered air
exposures or postexposure exercise challenge.




Exposure to 0.12 ppm Oj did not effect pulmonary function.
Preexposure to 0.12 ppm 03 at rest did not effect the magnitude
or time course of exercise-induced bronchoconstriction.


Filtered air followed by SOj and 03 alone did not cause
significant changes in pulmonary function. Ozone followed by
SOj resulted in significant decrease in FEVj (8%) and V^^Q^
(15%) and a significant increase in RT (19%).









Weymer et al. (1993)





Femandeset al.
(1993)



Koenig et al. (1990)

-------
TABLE 7-2 (cont'd). OZONE EXPOSURE IN SUBJECTS WITH PREEXISTING DISEASE8
«
8
3
H
I—*
v§
LtJ














^J
1
M



M
%^
J>
H
bL
O
2
^-«
o

O
d
g
a
IAJ
/~\
8
Ozone
Concentration Exposure
Duration and Exposure
ppm fig/m Activity Conditions
Adult Subjects with Asthma (cont'd)
0.00 0 1.5 h IE, Tdb = 22°C
0.12 236 VE = RH = 65%
0.24 472 25 L/min









0.00 0 6.5h/dayIE NA
0.12 236 (6 x 50 min)
(2 days of
exposure),
VE =
31 L/min
(asthmatic),
VE =
34 L/min
(healthy)
0.00 0 1 h rest NA
0.12 236
0.00 0 2 h IE Tdb = 31 °C
0.20 392 (4 x 15 min RH = 35%
at 2x rest
VE cycle
ergometry)

0.00 0 2 h rest NA
0.25 490



Number
and
Gender of
Subjects

4M, 4F
(nonasthmatics);
5M.5F
(asthmatics)








8M.7F
(nonasthmatics);
13 M, 17 F
(asthmatics)






4M
3F
20 M
2F




5M
12 F





Subjects
Characteristics

Physician-
diagnosed
asthma
confirmed with
methacholine
challenge test.
All nonsmokers
and
asymptomatic at
time of study.
Nine were
atopic.
Asthmatics
classified on
basis of positive
clinical history,
previous
physician
diagnosis, and
low PDjO' Mild
to severe
asthmatics.
Mild, stable
asthma
Physician
diagnosed
asthma;
6 smokers,
9 exsmokers,
7 nonsmokers
Nonsmoking
asthmatics
selected from a
clinical practice




Observed Effect(s)

No significant changes in pulmonary and nasal function
measurements in either asthmatics or nonasthmatics. Significant
increase in nasal lavage white cell count and epithelial cell
following 03 exposure in asthmatics only.








Significant increase in bronchial reactivity to methacholine in both
asthmatics and nonasthmatics. FEVj decreased 9.6% in
asthmatics and 1.7% in nonasthmatics, with difference not being
significant.






Increase in bronchial responsiveness to allergen; no change in
baseline airway function.
No significant changes in pulmonary function measurements;
significant blood biochemical changes.




No significant changes in pulmonary function measurements.







References

McBride et al. (1993)











Linn et al. (1993)









Molfino et al. (1991)

Linn et al. (1978)





Silverman (1979)

-------
                TABLE 7-2 (cont'd). OZONE EXPOSURE IN SUBJECTS WITH PREEXISTING DISEASE3
0
1
cr
Ozone
Concentration1 Exoosure
ppm
Duration and Exposure
ftg/m Activity Conditions
Number
and
Gender of
Subjects
Subjects
Characteristics0 Observed Effect(s)
References
£O Adult Subjects with Asthma (cont'd)

0.00
0.40
0 2 h ffi Tdb = 22 °C
784 (4 x 15 min RH = 50%
cycle
ergometry)
4M, 5F
(normals);
4M, 5 F
(asthmatics)
Asthmatics as Decrease in FVC and 1C with 03 in asthmatics; increase in airway
diagnosed by a responsiveness to methacholine in asthmatics with 03 and FA;
physician; asthmatic subjects had significantly greater decreases in FEVj and
history of chest f^25-75% w*m ®3 exposure than did normal subjects.
tightness and
wheezing
Kreit et al. (1989)
Eschenbacher et al.
(1989)
Adolescent Subjects with Asthma
to
0
d
o
1
0.00
0.12
0.00
0.12
0.00
0.12
0.18
0 1 h rest Tdb = 22 °C
235 RH ^ 75%
0 1 h IE Tdb = 22 °C
235 (2 x 15 min RH a 75%
treadmill
walking at
mean Vg =
32.5 L/min)
0 40 min IE NA
235 (1 X 10 min
353 treadmill
walking at
mean VE =
32.5 1/min)
4M, 6F
(normals);
4M, 6F
(asthmatics)
5M, 8F
(normals);
9M, 3 F
(asthmatics)
4M, 9F
(normals);
8M, 8F
(asthmatics)
Asthmatics had a Decrease in FRC with 03 exposure in asthmatics; no consistent
history of atopic significant changes in pulmonary functional parameters in either
extrinsic asthma group or between groups.
and exercise-
induced
bronchospasm
Asthmatics Decrease in maximal flow at 50% of FVC in asthmatics with
selected from a 03 exposure compared to FA; no significant changes with
clinical practice combined O3-NO2 exposure.
and had
exercise-
induced
bronchospasm
Asthmatics had Decrease in FEVj and increase in Rj in normals and asthmatics
allergic asthma, with 0.12 and 0.18 ppm 03 exposure compared to FA; no
positive consistent differences between normals and asthmatics.
responses to
methacholine,
and exercise-
induced
bronchospasm
Koenig et al. (1985)
Koenig et al. (1988)
Koenig et al. (1987)
 See glossary of terms and symbols for abbreviations and acronyms.
 Grouped by rest and exercise; within groups listed from lowest to highest 03 concentration.
°Age range in years or as mean ± SEM.

-------
 1      saturation in 28 mild to moderate COPD patients exposed to 0.18 and 0.25 ppm O3 for 1 h.
 2      The combined observations of these studies indicate that persons with COPD are not
 3      responsive to O3 concentrations of 0.30 ppm and lower in combination with mild exercise.
 4      However, this conclusion should be viewed within the context of the low total inhaled dose
 5      of O3 involved in the above studies, in that studies in healthy subjects using similar total
 6      inhaled doses have also not shown significant pulmonary function effects.  Interpretation of
 7      these studies is also complicated by the wide range of the pulmonary function impairment of
 8      the patients studied (FEV^FVC from 0.30 to 0.70), their variable smoking history, and the
 9      fact that these patients are older (» 60 years of age). The inconsistency of the observed
10     small decreases in arterial oxygen saturation makes the interpretation of the clinical
11      significance of this data difficult and uncertain.
12          Despite similar limitations, Kulle et al. (1984) observed small (<4%) significant
13     (p <  0.05) decreases in FVC and forced expiratory volume in 3 s (FEV3) in  20 smokers
14     (age range 31 ot 51 years) diagnosed with mild chronic bronchitis exposed to 0.40 ppm
15     O3 for 3 h using an IE protocol (one 15-min exercise period beginning 1 h prior to end of
16     exposure, VE approximately 29 to 38 L/min).  In addition, Kulle et al. (1984) observed that
17     repeated daily exposure over a 5-day period led to an adaptation of these forced expiratory
18     endpoints, and that this adaptive response did not  last longer than 4 days.  The pulmonary
19     responses induced by O3 exposure in this study were associated with only  mild symptoms.
20
21     Subjects with Asthma
22           Three studies examining the pulmonary responses to acute O3 exposures in adult (Linn
23      et al., 1978; Silverman,  1979) and adolescent (Koenig et al., 1985) asthmatics were
 24      discussed in the 1986 EPA criteria document (U.S.  Environmental Protection Agency, 1986).
 25      Significant decrements in group mean pulmonary  function  were not observed for adult
 26     asthmatics exposed for 2 h at rest (Silverman,  1979) or with light IE (Linn et al., 1978) to
 27     O3 concentrations of 0.25 ppm or less. However, it should be noted that although group
 28     mean pulmonary function responses were not significantly affected in these studies, there
 29     were responsive asthmatic subjects who had obvious decrements in pulmonary function.
 30          Koenig and coworkers (Koenig et al., 1985, 1987, 1988) conducted  a series of studies
 31     examining the pulmonary responses of adolescent asthmatics and nonasthmatics (11 to

        December  1993                          7-28      DRAFT-DO NOT QUOTE OR CITE

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 1     19 years of age) exposed to low levels of O3.  Koenig et al. (1985) found no significant
 2     changes in pulmonary function or symptoms in 10 adolescent normal and asthmatic subjects
 3     (four male, six female)  who inhaled 0.12 ppm O3 for 1 h at rest.  The asthmatic subjects in
 4     this study were characterized as having histories  of atopic (Type I, immunoglobulin E PgE]
 5     mediated) asthma and exercise-induced bronchospasm.  Subsequently, in two separate studies
 6     of similar groups of adolescent asthmatics and nonasthmatics, Koenig and coworkers  (Koenig
 7     et al., 1987, 1988) observed no significant changes in pulmonary function or symptoms
 8     following exposure to 0.12 and 0.18 ppm O3 with intermittent moderate exercise up to 1 h,
 9     although a small significant decrease in flow at 50% of FVC was observed in the adolescent
10     asthmatics exposed to 0.12 ppm O3.
11          Kreit et al. (1989) and Eschenbacher et al.  (1989) have demonstrated that exposure to
12     0.40 ppm 03 with heavy IE (exercise VE = 30 L/min/m2 BSA) for 2 h elicits a significant
13     decrease in FVC, FEVj, FEVyFVC, and FEF25_75% in both normal and asthmatic subjects.
14     In these studies, O3 exposure elicited significantly greater decrements in FEV19 FEVYFVC,
15     and FEF25_75% in asthmatic subjects.  In contrast, Kreit et al. (1989) and Eschenbacher et al.
16     (1989) found no significant difference between asthmatic and normal subjects in FVC and
17     subjective symptoms. In addition, the effect of O3 exposure on bronchial responsiveness as
18     measured by the concentration of methacholine needed to increase SRaw 100% (PC^SR^)
19     was  also studied.  The  asthmatic subjects had a significant decrease in PCi00SRaw following
20     filtered air and O3 exposure.  In comparison, the normal subjects  had a significant decrease
21     in PCjQgSR^ following O3 exposure, with the percent decrease in mean PC10oSRaw after
22     O3 exposure being similar in normal and asthmatic subjects, although the asthmatic patients'
23     baseline PC100SRaw was significantly lower than that of the normal subjects. The findings
24     from this study indicate that if the total inhaled dose is increased sufficiently by either
25     increasing VE during exposure or O3 concentration, mild to moderate asthmatics  will respond
26     with a greater obstructive response than will normal subjects.
27          Linn et al. (1993) have reported responses  of healthy (n = 15) and asthmatic (n = 30)
28     subjects to 0.12 ppm O3 and 100 /*g/m3  of respirable sulfuric acid (H2SO4) aerosol,  alone
29     and  in combination using the EPA prolonged-exposure protocol (see Section 7.2.2).  They
30     observed a significant O3-induced reduction in FEVl that was statistically significant and an
31     increase in airway responsiveness to methacholine for all subjects combined. The asthmatic

       December 1993                           7-29      DRAFT-DO NOT QUOTE OR CITE

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 1     subjects demonstrated a statistically significant decrease in FEVj as a function of exposure
 2     duration regardless of pollutant exposure.  In addition, Linn et al. (1993) observed a greater
 3     reduction in FEV] following O3 alone in the asthmatics as compared to the nonasthmatics
 4     (-8.6% versus -1.7%), although this difference was not statistically significant.  Despite
 5     the lack of a significant difference between asthmatics' and nonasthmatics'  group mean FEV]
 6     responses  with O3 exposure, the responses observed in the asthmatics may be considered
 7     more important because their average was already significantly depressed by the underlying
 8     illness.
 9           The findings from the above studies comparing the pulmonary function responses
10     following O3 exposure in asthmatic and nonasthmatic subjects  suggest that asthmatics are at
11     least  as sensitive, if not more sensitive to the acute effects of O3 inhalation.  The  underlying
12     mechanism that would explain a possible increased responsiveness of asthmatic  subjects to
13     O3 is undefined.  One possible mechanism could be that asthmatic subjects have an
14     exaggerated airway inflammatory response to acute O3 exposure.  A recent study  conducted
15     by McBride et al. (1993) would support this hypothesis.  McBride et al. (1993) exposed
16     10 asymptomatic asthmatic subjects with a history of allergic rhinitis and eight nonallergic
17     healthy subjects to filtered air and 0.12 and 0.24 ppm O3 for 90 min using a light IE
18     protocol (VE = approximately 25 L/min). Pulmonary function tests, posterior
19     rhinomanometry, and nasal lavage were performed before exposure and 10 min, 6 h,  and
20     24 h after exposure.  No significant  changes in pulmonary or nasal function were found in
21     either the allergic asthmatic or nonallergic nonasthmatic subjects. The allergic  asthmatic
22      subjects had a significant increase in the number of white blood cells in nasal lavage fluid
23      10 min and 24 h following the 0.24-ppm Oj exposure.  In addition, a significant  increase in
24      epithelial cells was present 10  min after exposure to 0.24 ppm O3 in the asthmatic subjects.
25      No significant cellular changes were observed in the nonasthmatic subjects.  These data
26      indicate that the upper airways of asthmatic individuals are more sensitive to the acute
 27      inflammatory effects of O3 than those of nonallergic nonasthmatic subjects.
 28           The above studies compared the effects of O3 inhalation on pulmonary function in
 29     asthmatic and normal subjects, but do not address the effect of preexposure to ambient
 30     concentrations of O3 on the responsiveness of asthmatic subjects to other  respiratory
 31      challenges, including other irritant gases, allergens,  and exercise.  Koenig et al. (1990)

        December 1993                           7-30       DRAFT-DO NOT QUOTE OR CITE

-------
 1     reported an increase in the bronchial response to an SO2 challenge in a group of
 2     13 asymptomatic adolescent asthmatic subjects following inhalation of 0.12 ppm O3 for
 3     45 min using a light to moderate intermittent exercise protocol (VE = approximately
 4     30 L/min).
 5          In a recent study, Molfino et al. (1991) investigated whether resting exposure to
 6     0.12 ppm O3 for 1 h potentiates the airway response to inhaled allergen in seven mild
 7     asthmatic  patients with seasonal symptoms of asthma and positive skin tests for ragweed or
 8     grass.  This study was conducted during 4 separate weeks during the winter when ambient
 9     allergen levels were low.  In each  week, there were 3 consecutive study days.  On Days 1
10     and 2, subjects underwent methacholine challenges, whereas on Day 2, the subjects received
11     one of four combined challenges in a single blind design:  air breathing followed by
12     inhalation of allergen  diluent, O3 exposure followed by inhalation of allergen diluent, air
13     breathing followed by inhalation of allergen, and O3 exposure followed by inhalation of
14     allergen.  Molfino et al. (1991) observed no significant differences in baseline FEVj after
15     O3 exposure, but did observe a significant reduction in the provocative concentration of
16     allergen required to reduce FEVj 15 %.  This study was limited by its small subject number
17     and the results  were confounded by possible ordering effects with the "O3 exposure followed
18     by allergen protocol"  being  the last protocol for all but one subject. Despite these
19     limitations, the findings suggest that O3 concentrations as low as 0.12 ppm may increase the
20     bronchial responsiveness to  allergen in atopic subjects and clearly indicates the need for more
21     studies that address this potentially significant health problem.
22          In order to examine whether preexposure to O3 results in exacerbation of
23     exercise-induced asthma, two studies were recently conducted (Fernandes et al.,  1993;
24     Weymer et al.,  1993). Fernandes et al. (1993) preexposed 15 stable mild asthmatics who
25     had never smoked with exercise-induced asthma to 0.12 ppm O3 for 1 h at rest  followed by
26     6 min exercise challenge test and found no significant effect on either the magnitude or time
27     course of exercise-induced bronchoconstriction.  Similarly, Weymer et al. (1993) observed
28     that preexposure to either 0.10 or  0.25 ppm O3 for 60 min while performing light IE did not
29     enhance or produce exercise-induced asthma in seven otherwise healthy adult subjects with
30     stable  mild asthma. Although the  results of these studies would suggest that preexposure to
31     O3 neither enhances or produces exercise-induced asthma in asthmatic subjects,  the small

       December 1993                           7_31      DRAFT-DO NOT QUOTE OR CITE

-------
 1     sample size and the relatively low total inhaled doses used in the above studies limit coir
 2     ability to draw  any conclusions.
 3
 4     Subjects with Allergic Rhinitis
 5          McDonnell et al. (1987) exposed 26 adults (18 to 30 years of age) with allergic rhinitis
 6     to clean air and 0.18 ppm O3 for 2 h using an IE protocol (VE = 64 L/min at 15-min
 7     intervals).  The study subjects  with allergic rhinitis did not have a history of asthma-like
 8     symptoms. Following O3 exposure, the subjects with allergic rhinitis exhibited significant
 9     (p < 0.01) increases  in respiratory symptoms, airway reactivity to histamine, and SRaw and
10     significant decreases in FVC, FEVj,  and FEF25_75% when compared to clean air exposure.
11     When compared to normal subjects without allergic rhinitis similarly exposed to 0.18 ppm
12     O3, the subjects with allergic rhinitis were no more responsive to O3 based on symptoms,
13     forced expiratory parameters, or airway reactivity to histamine aerosols,  although subjects
14     with allergic rhinitis did have a small but significantly greater increase in SR^.  The data on
15     subjects with allergic rhinitis and asthmatic subjects suggest that both of  these groups  have a
16     greater rise in airway resistance to O3 with a relative order of airway responsiveness to
17     O3 being normal < allergic < asthmatic.
18           Bascom et al. (1990) conducted a study to characterize the upper respiratory response
19     to acute  O3 inhalation, nasal challenge with antigen, and the combination of the two.
20     Bascom et al.  (1990) exposed  12 resting asymptomatic subjects with a history of allergic
21     rhinitis in a randomized, crossover design on each of 2 days, separated by 2 weeks, to clean
22     air or 0.5 ppm O3 for 4 h.  Following exposure, subjects underwent nasal challenge with
23     four doses of antigen (1, 10, 100, and 1,000 PNU ragweed or grass).  Upper and lower
24     airway symptoms were rated and nasal lavage was performed before and after clean air and
25     0.5 ppm O3 exposure, and following each antigen challenge.  Exposure to O3 caused
26      significant increases in upper and lower airway symptoms, a mixed inflammatory cell influx
27      with a sevenfold increase in nasal lavage neutrophils, a 20-fold increase  in eosinophils and a
28      10-fold increase in mononuclear cells as well as an apparent sloughing of epithelial cells.
29      There was a significant increase in nasal lavage albumin concentration following
30      O3 exposure.   When expressed as a  change from the postexposure values, there was no
31      significant difference between O3 and clean air exposure in antigen-induced upper and lower

        December 1993                           7-32      DRAFT-DO NOT QUOTE OR CITE

-------
 1     airway symptoms, cells, albumin and mediators (histamine and TAME-esterase activity).
 2     These results suggest that acute exposure to O3 does not alter the acute response to nasal
 3     challenge with antigen.
 4
 5     Subjects with Ischemic Heart Disease
 6           One study has been conducted examining the cardiopulmonary effects of acute
 7     O3 inhalation in patients with ischemic heart disease.  Superko et al. (1984) exposed six
 8     middle-aged males with angina-symptom-limited exercise tolerance for 40 min to filtered air
 9     and to 0.20 and 0.30 O3 while exercising continuously according to a protocol simulating
10     their  angina-symptom-limited exercise training prescription (mean VE = 35 L/min).
11     No significant pulmonary function impairment or evidence of cardiovascular strain induced
12     by O3 inhalation was observed. The low workloads were dictated by the patient's angina-
13     symptom-limited exercise tolerance and these low workloads acted to "protect" them from
14     O3-induced effects by limiting the total inhaled dose.
15
16     7.2.1.3   Influence of Gender, Age, Ethnic, and Environmental Factors
17     Gender Differences
18           As was noted in the previous O3 criteria document (U.S. Environmental Protection
19     Agency, 1986), the pulmonary function responses to O3 of only  a small number of female
20     subjects  have been evaluated under controlled laboratory conditions.  Although the database
21     on females has expanded (see Table 7-3), there are still fewer data than for males.  Most
22     studies involving mixed groups of male and female subjects include too few female subjects
23     to allow for meaningful comparisons between the responses of the sexes, or fail to consider
24     the question at all.  There are, however, a few studies including only female subjects.
25     Several studies cited in the 1986 O3 criteria document suggested that females might be more
26     responsive to O3 than males (Horvath et al., 1979;  Gliner et al., 1983; Gibbons and Adams,
27     1984; Lauritzen and Adams, 1985).  De Lucia et al. (1983), on the other hand, did not find
28     significant differences in the responses of young men and  young women to 63 exposure.
29           Messineo and Adams (1990) hypothesized that differences previously observed between
30     the responses of males and females exposed to O3 were related to differences in lung size
31     between the sexes.  They addressed this  issue by selecting two groups of 14 women each.
        !
       December 1993                           7.33       DRAFT-DO NOT QUOTE OR CITE

-------
TABLE 7-3. GENDER DIFFERENCES IN PULMONARY FUNCTION RESPONSES TO OZONE3
1
O"
rt>
vO
VO
W












-j
OJ
•£>•


o
x^
55
H
O
t— ^
Z
o
H
1
O
90
Ozone
Concentration
Exposure Duration
ppm
0.00
0.12
0.18
0.24
0.30
0.40

0.18




0.30

0.20
0.30




0.30








/ig/m and Activity
0
235
353
470
588
784

353




588

392
588




588








2.33 h
, O
VE = 25 L/min/ni
BSA
(one
exposure/subject)


1 h (mouthpiece)
CE
VE » 47 L/min




1 h (mouthpiece)
IE (20 min exercise)
VE * 28 L/min for
men
VE » 23 L/min for
women
1 h (mouthpiece)
CE
VE « 70 L/min for
men
VE « 50 L/min for
women



Exposure
Conditions'"
Mean T = 22 °C
MeanRH = 4%
treadmill




T = 21-25 °C
RH = 45-60%
cycle




T « 22 °C
RH > 75%
treadmill



T = 21-25 °C
RH = 45-60%
cycle






Number and
Gender of
Subjects
30-33 F and
30-33 M in
each
concentration
group; total of
372 individuals
participated
14 F




14 F

9M

10 F



20 M

20 F







Subject
Characteristics
Healthy NS, 18-35 years,
blacks and whites





Mean FVC =
5.11 ± 0.53 L,
NS, 20-24 years

Mean FVC =
3.74 ± 0.30 L,
NS, 19-23 years
55-74 years, NS

56-74 years, NS



18-30 years, NS

19-25 years, NS








Observed Effect(s)
Decrements in FEVj,
increases in SRaw and cough,
correlated with 03 dose.
There were no significant
differences between the
responses of males and
females.
Significant dose/response
effect on FVC and FEVj;
lung size had no effect on
percentage decrements in
FVCorFEV].


No changes in spirometry in
men or women. Women had
significant 13% increase in
Rj following exposure,
which was sustained at
20 min postexposure.
Significant decrements in
FVC, FEV1; and FEF25_75%
following 03 exposure.
No significant differences
between men and women for
spirometry or SRaw.





Reference
Seal et al. (1993b)






Messineo and Adams
(1990)





Reisenauer et al. (1988)





Adams et al. (1987)

-------
         TABLE 7-3 (cont'd).  GENDER DIFFERENCES IN PULMONARY FUNCTION RESPONSES TO OZONE3
o
1
1-J
>J
-n
Ozone
Concentration
Exposure Duration
ppm fj,g/m and Activity
0.45 882 2 h
IE
VE « 27.9 L/min
for men
VE « 25.4 L/min
for women
0.45 882 2 h
IE
Mean VE =
28.5 L/min for men
Mean VE =
26. 1 L/min for
women
0.48 941 2 h
IE
VE « 25 L/min
Exposure
Conditions0
T = 24 °C
RH = 58%
cycle
MeanT = 23.1
MeanRH =
46.1%
cycle/treadmill
T = 21 °C
WBGT
cycle
Number and
Gender of
Subjects
8M
8F
°C 10 M
6F
10 F
Subject
Characteristics
Healthy NS,
51-69 years
Healthy NS,
56-76 years
Healthy NS,
60-89 years
Healthy NS,
64-71 years
Healthy NS,
19-36 years
Observed Effect(s)
Range of responses in FEVj :
0 to -12% (mean = -5.6%).
No significant difference in
responses of men and women.
Tendency for women to have
greater effects.
Mean decrement in FEVj =
5.7%. Decrements in FVC and
FEVj were the only pulmonary
functions significantly altered by
03 exposure. No significant
differences between responses
of men and women.
Mean decrement in
FEVj = 22.4%.
Significant decrements in all
Reference
Drechsler-Parks et al.
(1987a,b)
Bedi et al. (1989)
Horvath et al. (1986)
                                                                                spirometry measurements.
                                                                                Results not significantly
                                                                                different from a similar study
                                                                                on males (Drechsler-Parks
                                                                                etal., 1984).
 See glossary of terms and symbols for abbreviations and acronyms.
 Listed from lowest to highest O3 concentration.
^WBGT = 0.7 Twetbulb + 0.3 Tdry bulb or globe.
 Age range in years or as mean ± SEM.

-------
 1      One group had a mean FVC of 5.11 L, and the other group had a mean FVC of 3.74 L.  All
 2      subjects were 19 to 24 years of age and were healthy nonsmokers who had not lived in a
 3      high air pollution area for at least 6 mo. The subjects completed three 1-h CE (VE =
 4      47 L/min) exposures: (1) FA, (2) 0.18 ppm O3, and (3) 0.30 ppm O3.  The mouthpiece
 5      exposures were presented in random order, at least 4 days apart, and all were performed
 6      when the subject was in the follicular phase of her menstrual cycle.  Two subjects in the
 7      small lung group and one in the large lung group were unable to complete the 0.30 ppm
 8      O3 exposure. Both groups had similar O3-induced percentage decrements in all measures of
 9      lung function, regardless of lung size, leading to the conclusion that lung  size, per se, is not
10     systematically related to percentage decrements in FEV1 consequent to O3 exposure.
11           Horvath et al. (1986) exposed 10 healthy, young, nonsmoking females, 19 to 36 years
12     of age (mean age 23.6 years) to 0.48 ppm O3 or FA for 2 h in an environmental chamber
13     while they exercised intermittently at a target ventilation of 25 L/min. The subjects engaged
14     in three 20-min cycle ergometer exercise periods alternated with four 15-min rest periods.
15     The exposures were a minimum of 1 week apart.  The responses  of these subjects were
16     compared with those of a group of 10 young males who had earlier completed the same
17     protocol (Drechsler-Parks et al., 1984). There were no statistically significant differences in
18     the responses based on gender.  The female subjects had decrements of 18.8, 22.4, and
19     30.8% in FVC, FEVl5 and FEF25.75%, respectively, compared to 19.8, 25.0, and 31.9% for
20     the male subjects.  On an individual basis, 4 of the  10 males and 3  of the 10 females had
21     decrements  of 30% or more in FEV]^ following the  exposure to 0.48 ppm O3.  One male
22     subject did not respond to the O3 exposure.  It was  noted, however, that  the female subjects
23     inhaled an absolute dose  of O3 about 22% less than the male subjects due to a slightly lower
24     exercise VE and the inherently lower resting VE of females compared to males. However,
25     if O3 dose was related to BSA or to FVC, the females inhaled slightly higher relative doses
 26     of O3 than the males.
 27           Adams et al. (1987) compared the responses of 20 young men (18 to 30 years of age)
 28      and 20 young women (19 to 35 years of age) to exposure to 0.30 ppm 05 via mouthpiece.
 29     All subjects were healthy nonsmokers with clinically normal pulmonary function.  None had
 30     a history of significant allergies, and none had resided  in a high  air pollution area for  at least
 31      3 mo.  The subjects completed 1-h CE exposures (mean VE » 70 L/min for males and

        December  1993                         7-36       DRAFT-DO NOT QUOTE OR CITE

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 1     50 L/min for females) to FA and 0.30 ppm O3.  The exposures were given in random order
 2     and were separated by a minimum of 5 days.  Ozone exposure induced significant
 3     decrements in FVC, FEVj, and FEF25_75% compared to FA exposure.  Three females and
 4     four males were unable to complete the O3 exposure.  Females experienced mean decrements
 5     of 14.2, 20.3, and 24.5% in FVC, FEVl5 and FEF25.75%, respectively, compared to mean
 6     decrements of 15.8% in FVC, 23.8% in FEV1? and 35.7% in FEF25.75% for males. There
 7     were no statistically significant differences between the spirometry or SRaw responses related
 8     to gender.  Because the female subjects inhaled a substantially smaller absolute dose of
 9     O3 due to the considerably lower exercise VE, yet had similar decrements in pulmonary
10     function compared to men, the authors concluded that females are more responsive to
11     O3 than males.  In this  study,  the female subjects inhaled a lower relative dose of
12     O3 compared to males,  if expressed on the basis of BSA, but a similar relative dose if
13     expressed on the basis of FVC.
14          Seal et al.  (1993b) reported on 372 healthy black and white men and women between
15     18 and 35 years of age  who  each were assigned to complete one 2.33-h exposure to FA,
16     0.12, 0.18, 0.24, 0.30,  or 0.40 ppm O3.  Subjects exercised intermittently on a motor-driven
                                        •                     0
17     treadmill at a work load inducing a  VE of about 23 L/min/m  BSA for women and about
18     24.5 L/min/m2 BSA for men.  Although female subjects inhaled about 22%  less total dose of
19     O3 than males in each exposure-concentration group, there were no significant differences in
20     the changes in FEVj 0,  SRaw,  or cough ratings between males and females among either
21     blacks or whites. Women also inhaled a lower absolute dose of O3 than men.
22          Drechsler-Parks et al. (1987a,b) compared the responses of eight men and eight women
23     between 51 and 76  years of age to FA and 0.45 ppm O3.  The subjects were all healthy
24     nonsmokers who were long-term residents of a relatively low pollution area. The subjects
25     participated in 2-h intermittent exercise (20 min rest/20 min exercise at  VE =  25 L/min)
26     environmental chamber  exposures that were presented in random order and were separated
27     by at least 1 week.   Except for FEV3, there were no statistically significant differences
28     between the responses of the men and women subjects, although women had slightly larger
29     mean decrements in FVC and  FEVj 0 than men.  Individual decrements in FVC and FEVj 0
30     ranged from 0 to about  12% for both male and female subjects.  Based on FEVj, two
31     females and three males had no response to the O3 exposure.  Male subjects inhaled a

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 1      somewhat larger absolute effective dose of O3 due to a higher exercise and resting VE.
 2      If VE was normalized to BSA, females inhaled a larger dose of O3 than males.  If VE was
 3      normalized to FVC, the relative inhaled doses of O3 were similar.
 4           Reisenauer et al.  (1988) reported on the pulmonary function responses of 9 men and
 5      10 women between 55 and 74 years of age who were exposed to 0.0, 0.20, and 0.30 ppm
 6      O3.  The three exposures were presented in random order and at the same time of day for
 7      each subject.  The subjects were exposed via mouthpiece for 1 h, during which seven men
 8      exercised for  10 min and rested for 50 min, and the other two men and all of the women
 9      alternated two 20-min  rest periods and two 10-min exercise periods.  Ventilation rates were
10      about 28 L/min for men and 23 L/min for women, although if VE is  normalized to BSA, the
11      relative VE for males and females was similar. All data were pooled, regardless of the total
12     exercise tune. There were no significant changes in any parameter of pulmonary function in
13     the males.  Females had no significant changes in any spirometric parameter, but did have a
14     small (13%) increase in total respiratory resistance (RT) following the 0.30 ppm
15     O3 exposure, which remained at this level 20 min postexposure.
16          Bedi et  al. (1989) reported on the responses of 10 men and 6 women (60 to 89 years of
17     age) exposed for 2 h to FA or 0.45 ppm O3 for 3 consecutive days.  Only the first O3 day
18     results will be discussed in this section.  The issue of repeated exposures is addressed in
19     Section 7.2.1.4.  Exposures were conducted at the same time of day, on consecutive days,
20     with the FA exposure always conducted first. The subjects alternated 20-min exercise
21     periods (mean VE =  28.5 L/min for men and 26.1 L/min for women) and 20-min rest
22     periods throughout the 2-h chamber exposures.  If VE is normalized to BSA, women inhaled
 23      slightly higher relative doses of O3; but if normalized to FVC, women inhaled a slightly
 24      lower  relative dose of O3 than men.  There were no  statistically significant group mean
 25      differences between the responses of men and women subjects.  The mean decrements in
 26     FVC and FEVj following the O3 exposure for the 16 subjects were 2.8 and  5.7%,
 27     respectively.  In an exploratory analysis, the subjects were divided into two  groups based on
 28     whether their decrement in FEV^^ following the first O3 exposure compared to the FA
 29     exposure was >5% or <5%.  There were eight subjects in each group, with the sensitive
 30     group consisting of two females and six males.  The mean post-O3 exposure decrement in
 31     FEVt was 320 mL for the sensitive group,  versus 21 mL for the nonresponsive group.

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 1      Similar patterns of response were evident in FVC and FEV3 0.  There were no significant
 2      changes in any flow parameter, maximum voluntary ventilation (MVV), expiratory reserve
 3      volume, or functional residual capacity.
 4           The question as to whether there is a difference hi responsiveness to O3 between men
 5      and women remains unresolved. Different conclusions can be arrived at depending on
 6      whether VE is normalized to body or lung size when the inhaled doses of O3 are calculated.
 7      The sensitive  subgroup studied by Bedi et al. (1989) included six males and two females,
 8      suggesting that older males may be more responsive to O3 than females. However,
 9      Reisenauer et al. (1988) found a significant increase in RT only in women.  Horvath et al.
10     (1986), Adams et al. (1987), and Drechsler-Parks et al. (1987a,b) suggested that because
11      their female subjects had similar pulmonary function responses to their male subjects, even
12     though the females inhaled less O3,  females were more responsive than males.  Messineo and
13      Adams (1990) suggested that some factor other than absolute lung size accounted for
14     observed differences between males and females. Their two groups of females with widely
15      different lung sizes experienced similar decrements  in pulmonary function following
16     equivalent exposures.  Although the currently available literature suggests that females may
17     be somewhat  more responsive to O3 than males, the question is not settled. Further,
18     comparative studies have included only small subject groups, except for Seal et al. (1993b),
19     and often only group mean data are presented, with little information about individual
20     responses.  Small subject groups with wide differences among the inherent responsiveness of
21      the individuals in the groups can have disproportionate effects on group mean data and on
22     conclusions reached.
23
24     Hormonal Influences
25          Seal et al. (1993a) compared the pulmonary function responses of 48 white and
26     55 black women (18 to 35 years of age) whose menstrual phase was known at the time of a
27     single 2.3-h exposure to 0.18, 0.24, 0.30, or 0.40 ppm O3.  Subjects performed intermittent
28     treadmill exercise (VE = 20 L/min/m2 BSA) during the first 2 h of exposure.  There were
29     no significant effects for SR^ or cough that could be related to menstrual cycle. There was
30     a race x menstrual phase interaction for FEVj.  However, when the groups of black and
31     white women were analyzed separately, there was no significant main effect for menstrual

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 1     cycle phase.  The significance of the observed interaction between race and menstrual cycle
 2     phase is unknown.
 3          Gerbase et al.  (1993) compared the pulmonary function responses of six healthy,
 4     nonsmoking women to a 130-min exposure to 0.35 ppm O3 4 to 8 days after the onset of
 5     menses and 4 to 8 days after ovulation.  Subjects performed intermittent exercise at a
 6     workload that induced a VE of 10 x FVC.  Ovulation was confirmed by a blood
 7     progesterone test.  Spirometry was performed pre- and  25-min post-O3 exposure. Although
 8     resting VE was the same during both exposures, exercise load had to be reduced 30% during
 9     the luteal phase in order to match the ventilatory response to exercise during the follicular
10     phase.  There were  no significant effects related to phase of the menstrual cycle.  The
11     authors concluded that menstrual phase does not need to be considered in experimental
12     design.  One problem with the study is that the postexposure measurements were made
13     25 min after the conclusion of the exposure.  Typically, pulmonary function decrements
14     begin to reverse once exposure ends; thus, any pulmonary function changes that did occur
15     could be expected to be reduced  at 25-min post-O3 exposure compared to immediately after
16     exposure.
17          Acute O3 exposure has been shown to induce short-term airway inflammation (see
18     Section 7.2.4) induced by prostaglandins, among other inflammatory substances.  It has also
19     been demonstrated that progesterone inhibits prostaglandin production in the uterine
20     endometrium, which fluctuates as the progesterone concentration varies throughout the
21     menstrual cycle.  Fox et al. (1993) investigated the hypothesis that O3 exposure during the
22     follicular phase, when progesterone  concentration is lowest, might result in greater
23     pulmonary function responses due to reduced anti-inflammatory influences of progesterone.
24     Nine nonsmoking women completed 1-h mouthpiece exposures to FA and 0.30 ppm O3 while
25     exercising continuously (VE about 50 L/min) during both the follicular and mid-luteal phases
26     of two to four ovulatory menstrual cycles. There were no differences in any pulmonary
27     function responses to FA related to  menstrual phase, nor was there a difference in the mean
28     FVC decrements  following the follicular or mid-luteal phase O3 exposures. The O3-induced
29     decrements hi FEVj and FEF25_75% were significantly larger during the follicular phase than
 30     during the mid-luteal phase.  The authors speculated that the difference between the FEVj
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 1     and FEF25_75% responses to the two O3 exposures could be due to differences in circulating
 2     progesterone, and the effect of progesterone on prostaglandin activity.
 3          Available data (see Table 7-4) do not permit a conclusion regarding the influence of the
 4     menstrual cycle on responses to O3 exposure.  Two of the three studies available, Fox et al.
 5     (1993) and Gerbase et al. (1993), were performed with small groups of subjects and resulted
 6     in opposite conclusions.  Seal et al. (1993a) compared race (black versus white) and
 7     menstrual phase, obtaining a significant interaction between race and phase, but post-hoc
 8     analysis failed to establish a basis for the interaction, leaving the implications of the study
 9     unclear.
10
11     Age Differences
12          It has been hypothesized that age may be a factor in responsiveness to O3. Although
13     children make up a large proportion of the population, few controlled laboratory studies of
14     the pulmonary function effects of any air pollutant have been reported on subjects under
15     age 18. Field and epidemiological studies (see Section 7.4) attempting to relate ambient air
16     pollutant exposure to pulmonary function  in children have suggested that children may be
17     more responsive to ambient air pollution than young adults.
18          The previous O3 criteria document (U.S. Environmental Protection Agency, 1986)
19     included only one laboratory exposure study in which children were the subjects. McDonnell
20     et al. (1985a) evaluated the pulmonary function responses of 23 boys between 8 and 11 years
21     of age to 0.00 and 0.12 ppm O3 in random order.   The boys alternated  15-min rest and
                        .                o
22     exercise periods (VE = 35 L/min/m  BSA) for the first 120 min of the 150-min exposure.
23     Forced expiratory spirometry and respiratory  symptoms were measured  before exposure and
24     at 125 min of exposure, whereas airway resistance was  measured before exposure began and
25     after 145 min of exposure.  The group mean decrement in FEVj following the O3 exposure
26     was 3.4%, compared to 4.3%  for a group of  young adult males who had earlier completed
27     the same protocol (McDonnell et al., 1983).  It should be noted that the absolute VE for the
28     children (39.4 L/min) and adults (65.0 L/min) was similar if normalized for BSA (about
29     35 L/min/m  BSA). Assuming that adjusting ventilation for differences in BSA is an
30     appropriate normalizing technique, these children appeared to experience similar O3-induced
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D
TABLE 7-4.  HORMONAL INFLUENCES ON PULMONARY FUNCTION RESPONSES TO OZONE3
3 Ozone
g- Concentration5 ETOMim
H^
^O
\O ppm
0.12
0.24
0.30
0.40
0.30
-j
^
0.35
d
Duration and Exposure Number and Subjects
Atg/m Activity Conditions Gender of Subjects Characteristics0
235 2.3 h IE NA 48 WF Healthy NS,
470 VE = 55 BF 18-35 years
588 20 L/min/m2 BSA
784
588 1 h CE NA 9 F Healthy NS,
VE - 50 L/min ^£Ll
cycles,
20-34 years
686 130 min NA 6F Healthy NS,
regular
menstrual cycles
(age not stated)
Observed Effect(s) References
Significant menstrual cycle phase X race Seal et al. (1993a)
interaction for FEVj. No significant
menstrual cycle phase effect when blacks
and whites were analyzed separately.
No significant menstrual phase effects
for SRaw or cough score.
FEV! decreased 13. 1 % during the mid- Fox et al. (1993)
luteal phase and 18.1% during the
follicular phase. Decrement in
FEF25_75% was significantly larger
during the follicular phase than the mid-
luteal phase. Changes in FVC were
similar in both phases.
Changes in FVC, FEVj, FEF25.75%, Gerbase et al.
Vmax50%> *&& ^msx25% were similar (1993)
during both the follicular and luteal
phases.
    See glossary of terms and symbols for abbreviations and acronyms.
    Listed from lowest to highest O3 concentration.
   cAge range in years or as mean ± SEM.

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 1     pulmonary effects as adults.  The children reported no symptoms, although the adults
 2     reported a small but statistically significant increase in cough following O3 exposure.
 3          It continues to be the case that controlled laboratory studies of the effects of exposure
 4     to air pollutants are rarely performed with children as subjects (see Table 7-5).  Avol et al.
 5     (1987) have reported on the pulmonary function responses of 33 healthy boys and 33  healthy
 6     girls having a mean age of 9.4 years.  The children completed exposures to purified air and
 7     outdoor ambient air that was drawn into an environmental chamber.  Ambient temperature
 8     averaged about 33  °C.  Exposures  were 1 h in duration, were  separated by a minimum of
 9     2 weeks, and were conducted from June through September, beginning in the early afternoon
10     when ambient air pollutant concentrations generally peak.  The subjects performed
11     continuous exercise throughout the  hour of exposure.  Boys and girls exercised at similar
12     VE, 22 to 23 L/min.  It should be  noted that the ambient exposure included the full range of
13     air pollutants present in the outdoor air mix on the days of the exposures, except for small
14     fractions of O3  and paniculate lost  in the inlet duct.  Concentrations of O3, nitrogen dioxide
15     (NO2), total suspended particulate (TSP), paniculate nitrate, paniculate sulfate, paniculate
16     sodium, and particulate ammonium were measured throughout the exposures. The
17     O3 concentration during the ambient air exposures averaged 0.113 ± 0.033 ppm, whereas it
18     averaged 0.003 +  0.002 ppm during the purified air exposures. The children consistently
19     had a decline in pulmonary function with time, following both FA and ambient air
20     exposures.  Typical mean decrements in FVC and FEV1 were 50 mL or less.  A correlation
21     analysis between inhaled O3 dose and the mean net pre- to postexposure volume change
22     (in liters) in FEVl5 including  all valid pulmonary tests, indicated an r of -0.27 (p < 0.05).
23     The investigators have also published similar studies on adolescents and adults (Avol et al.,
24     1984, 1985a).  The responses of the adolescents and adults to both exposures were not
25     substantially different from those of the children whose results are reported here.  The
26     authors further noted that the  children seemed to have difficulty performing consistent,
27     reproducible pulmonary function tests, a factor which could have impacted on these results.
            Several studies comparing the pulmonary function responses of healthy and asthmatic
       adolescents to O3 exposure have appeared in the literature. The responses of the asthmatics
       are presented in Section 7.2.1.2; only data on normal adolescent subjects will be discussed in
       this section.

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TABLE 7-5. AGE DIFFERENCES IN PULMONARY FUNCTION RESPONSES TO OZONE8
8
I
Co
VO
u>
-J
£
0
§
2,
S
1
i
Ozone
Concentration Exposure Duration
3
ppm jug/m and Activity
0.113
+
other
ambient
pollutants
0.12
0.12
0.18
0.18
0.24
0.30
0.40
0.20
0.30

221
235
235
353
353
470
588
784
392
588

In
CE
VE « 22 L/min
1 h (mouthpiece)
IE
VE = 4-5 X resting
40 min (mouthpiece)
IE
10 min exercise at
VE = 32.6 L/min;
40 min (mouthpiece)
IE
10 min exercise at
VE = 41.3 L/min
2.3 h
IE
VE = 20 L/min/m2
BSA
1 h (mouthpiece)
IE (20 min)
VE « 28 L/min for
men
VE « 23 L/min for
women

Exposure
Conditions
T = 32.7 °C
RH « 43%
cycle
T = 22 °C
RH = 75%
treadmill
NA
treadmill
NA
T » 22 °C
RH S: 75%
treadmill

Number and
Gender of Subject
Subjects Characteristics6
33 M
33 F
5 M
7 F
3 M
7 F
4M
6F
48 WF
55 BF
9M
10 F

NS for both groups,
mean age = 9.4 years
Healthy NS,
12-17 years
Healthy NS,
14-19 years
Healthy NS,
18-35 years
Black and white
Healthy NS,
55-74 years

Observed Effect(s)
No differences in responses
of boys and girls. Similar
decrements (<5% on
average) following both
purified air and ambient air
(03 at 0.11 ppm) exposures.
No significant changes in any
pulmonary function in
healthy subjects.
No significant change in
FEVj; increased Rf with
exposure to 0.18 ppm O3.
Some subjects responded to
5-10 mg/mL methacholine
after 0. 18-ppm 03 exposure,
whereas none responded to
25 mg/mL methacholine at
baseline bronchochallenge.
Older women had smaller
changes in FEV1 than
younger women. No age-
related differences in SRaw
or cough score.
No change in any spirometry
measure. Women had 13 %
increase in RT after
0.30-ppm exposure.

Reference
Avol et al. (1987)
Koenig et al. (1988)
Koenig et al. (1987)
Seal et al. (1993b)
Reisenauer et al. (1988)

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         TABLE 7-5 (cont'd).  AGE DIFFERENCES IN PULMONARY FUNCTION RESPONSES TO OZONE3
o
CD
cr
UJ
d
6
o
Ozone
Concentration Exposure Duration
O
ppm jig/m and Activity
0.45 882 2 h
IE
VE ~ 26 L/min
0.45 882 2 h
IE
Mean VE =
28.5 L/min for men
Mean VE =
26.1 L/min for
women
0.45 882 2 h
IE
VE « 26 L/min
0.45 882 1 h
CE
VE « 26 L/min
2h
IE
VE « 26 L/min
Exposure
Conditions
T « 23 °C
RH = 53%
cycle
T = 23 °C
RH = 46%
cycle/treadmill
T « 24 °C
RH = 63%
cycle
T « 23 °C
RH = 58%
cycle/treadmill
Number and
Gender of
Subjects
8M
8F
10 M
6F
8M
8F
7M
5F
Subject
Characteristics0
Healthy NS
51-76 years
Healthy NS
60-89 years
Healthy NS
51-69 years
Healthy NS
56-76 years
Healthy NS
60-79 years
(all hi 60s
except one
79-year-old)
Observed Effect(s) Reference
Mean decrement hi Drechsler-Parks et al.
FEV! = 5.6 ± 13%; range of (1987a,b)
decrements = 0-12%.
Mean decrement in Bedi et al. (1989)
FEVj = 5.7%; eight subjects had a 5%
or greater difference between their
response to 03 and FA, and the other
eight had less than a 5% difference
between their responses to FA and
0.45 ppm 03.
13 subjects had decrements in FEVj Bedi et al. (1988)
on three separate exposures to 0.45 ppm
within 5% of their mean response to the
three exposures. The other
three subjects were not reproducible.
Symptom reports did not correlate well
with pulmonary function changes.
Comparison of 1-h CE protocol and 2 h Drechsler-Parks et al.
IE protocol indicated no difference (1990)
between the changes hi pulmonary
function following the two protocols.
See glossary of terms and symbols for abbreviations and acronyms.
Listed from lowest to highest O3 concentration.
Age range hi years or as mean ± SEM.

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 1          Koenig et al. (1987) reported on 20 adolescents, 14 to 19 years of age, who were
 2     exposed for 40 min to air, 0.12 ppm O3, or 0.18 ppm O3 via a mouthpiece system.  Ten
 3     subjects were exposed to each O3 concentration, but not all subjects were exposed to both
 4     concentrations. None of the healthy subjects had a history of asthma or allergies, and all had
 5     pulmonary function within the predicted range, based on age, sex, and height.  There was a
 6     5- to 7-min break in exposure for pulmonary function test performance following 30 min of
 7     resting exposure,  followed by a 10-min exercise period (VE = 32.6 ± 6.4 L/min for the
 8     0.12-ppm O3 exposure and 41.3 ± 9.3 L/min for the 0.18-ppm O3 exposure).  Changes in
 9     FEV1 were not significant following any exposure.  After exposure to 0.18 ppm O3, RT was
10     increased 15%.
11          Koenig et al. (1988)  have also reported on the pulmonary function responses of another
12     group of 12 healthy adolescents (12 to 17 years of age) to 1-h exposures to air and 0.12 ppm
13     03.  The subjects were exposed by mouthpiece to air and 0.12 ppm O3 while alternating
14     15 min periods of exercise (VE =  32.8 ± 6.0 L/min) with 15 min periods of rest (VE =
15     8.8 ± 1.2 L/min).  Tests of pulmonary function included forced expiratory spirometry and
16     RT.  Healthy subjects had no significant alterations in any parameter of pulmonary function
17     consequent to either air  or 0.12-ppm O3 exposure.
18          Although few data are available on the responses of healthy adolescents exposed to 03,
19     the limited existing data do not identify adolescents as being either more or less responsive
20     than young adults.
21          At the time the 1986 O3 criteria document was released, no studies  specifically
22     evaluating the pulmonary function responses of older adults had been reported.  Several
23     studies (Folinsbee et al., 1985; Adams et al., 1981) that included a few middle-aged
24     individuals among the subjects were suggestive that there might be a decrease in
25     O3 responsiveness with  advancing age. Several reports have since appeared, collectively
26     suggesting that healthy older adults (i.e.,  over 50 years of age) are generally minimally
27     responsive to O3, although some individuals remain responsive to O3.
28           Drechsler-Parks et al. (1987a) reported on eight men and eight women between 51 and
29     76 years of age who were exposed for 2 h to FA or 0.45 ppm O3.  The subjects were
 30      healthy nonsmokers with normal baseline pulmonary function.  The chamber exposures
 31      involved alternating 20-min rest and exercise periods (VE averaged 27.9 ± 0.29 L/min for

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 1     men and 25.4 ± 0.80 L/min for women). Exposures were presented in random order at
 2     least 1 week apart.  The only significant difference related to sex was in FEV3, in which
 3     women had larger decrements than men.  There were no significant decrements in any other
 4     parameter of pulmonary function related to O3 exposure, except when the data of all
 5     16 subjects  were pooled, resulting in significant mean decrements of 5.3 ± 1.3% in FVC,
 6     and 5.6 ± 1.3% in FEVj.  The range of individual decrements in FEVj was from 0 to 12%.
 7     Two women and three men had no response following the O3 exposure. The subjects
 8     reported more symptoms following the O3 exposure than the FA exposure. Seven subjects
 9     reported cough, nine reported sore throat, and six reported chest tightness. The authors
10     compared their results on older adults with a group of young adults who had completed the
11     same protocol. Both the older men and women inhaled slightly higher doses of O3 than the
12     young men  and women (10.23 x  104 L for older men versus 10.12  x 10"4 L for young
13     men, and 8.48 x 10"4 L for older women versus 7.94 X 10"4 L for young women).
14     However, older men had a mean decrement in FEVj of 4.2% versus 23.7% for young men,
15     whereas older women had a mean decrement in FEVj  of 7.0% versus 14.7% for young
16     women.  The decrements of the older subjects were also compared to published values for
17     young adults, with the older subjects studied consistently showing smaller changes in
18     pulmonary functions than young adults. These comparisons indicated that these older adults
19     were less responsive to O3 exposure than typical young adults, in terms of pulmonary
20     function changes and symptom reports.
21          Reisenauer et  al.  (1988) reported on the pulmonary function responses of 19 healthy
22     adults between 55 and 74 years of age.  All were nonsmokers with baseline pulmonary
23     function within the  predicted normal range.  None had a history of asthma, atopy, or
24     cardiovascular disease, and none responded to a baseline methacholine bronchochallenge
25     (up to 25 mg/mL).  Subjects were exposed by mouthpiece to 0.0, 0.20, or 0.30 ppm O3 for
26     1 h.  Seven men rested for 50 min and exercised for 10 min, whereas the other two men and
27     all women alternated two 20-min rest periods with two 10-min exercise periods; VE was
28     approximately 28 L/min for men and 23 L/min for women. The three  exposures were
29     presented in random order at the same time of day for each individual,  but the  separation
30     between exposures  is not stated.  The only significant change in pulmonary function was a
31     13% increase in RT with exposure to 0.30 ppm O3 in women only, which was  sustained for

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 1      at least 20 min postexposure.  The authors concluded, based on the increase in RT in the
 2      older women, that older adults were at increased risk for pulmonary function changes with
 3      near ambient O3 exposure.  However, RT is a highly variable parameter, and no other
 4      changes were significant.  Given the large number of variables tested, this isolated result is
 5      possibly related to the large number of statistical tests performed.  In contrast, the results of
 6      Koenig et al. (1987), from 10 healthy adolescents exposed to 0.18 ppm  O3 using a similar
 7      protocol, reported a mean decrement in FEVj 0 of 2%, and an increase  of 10.5% in RT at
 8      2 to 3 min postexposure.  At 7 to 8 min postexposure, the increase in RT was 15.3%.
 9      Comparison of these results to those of Reisenauer et al. (1988) at 0.20 ppm O3 supports the
10      contention that younger individuals are more responsive to O3 than older individuals, in that
11      no changes in spirometry were noted in the older adults exposed to 0.30 ppm C^, although
12     older women showed increased RT with 0.30 ppm O3 exposure.  Adolescents had alterations
13     in both spirometry and RT with exposure to 0.18 ppm O3.
14          Bedi et al. (1988) reported that older men and women 51 to 76 years of age who
15     completed three exposures to  0.45 ppm O3 did not respond equivalently to each of three
16     exposures.  The subjects were healthy nonsmokers with baseline pulmonary function within
17     predicted normal limits. The subjects alternated 20-min exercise (VE was approximately
18     26 L/min) and  20-min rest periods throughout the 2-h chamber  exposures. There was a
19     minimum of 1  week between exposures, although separations ranged from 1 to 4 weeks
20     between exposures  1 and 2, and between 1 and 7 weeks between exposures 2 and 3.
21     Analysis of variance indicated no difference between the group  mean responses to the three
22     exposures.  The data were then subjected to a correlation analysis, which led to the
23     conclusion that the responses within an individual subject were  not reproducible.  McDonnell
 24      et al. (1985b), on the other hand, found good reproducibility of pulmonary function
 25      responses after exposure to various concentrations of O3 between 0.12  and 0.40 ppm in
 26      young adult males between 18 and 30 years of age.
 27           Seal et al. (1993a) compared the pulmonary function responses of 48 white and
 28      55 black women (18 to 35 years) who each completed a 2.3-h exposure to 0.18, 0.24, 0.30,
 29     or 0.40 ppm O3.  The subjects participated in only one exposure each while intermittently
 30     exercising (VE = 20 L/min/m2 BSA) during the first 2 h of the exposure.  Older subjects
 31      within the age range tested had smaller decrements in I!EVl than younger subjects.

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 1          One simple method to estimate the O3 exposure dose is to calculate the product of
 2     O3 concentration (parts per million), VE (liters per minute), and exposure duration
 3     (minutes).  Research on young adults (Folinsbee et al., 1978; Adams et al., 1981) has
 4     demonstrated that the order of relative importance of the three factors is O3 concentration,
 5     VE, and exposure duration.  Drechsler-Parks et al. (1990) investigated the relative role of
 6     the three components of effective dose in 12 healthy, nonsmoking adults between 61 and
 7     79 years of age.  The subjects were exposed to both FA and 0.45 ppm, once while they
 8     performed a 1-h continuous exercise protocol and once while they performed a 2-h
 9     intermittent exercise protocol in which they alternated 20-min exercise periods and 20-min
10     rest periods.  Mean VE ranged from 25.2 to 27.3 L/min among the four exposures.
11     Exposures were separated by at least 1  week.  Regardless of protocol, O3 exposure induced
12     significant decrements in FEV0 5, FEVj, FEV3, and peak flow rate compared to FA
13     exposure.  There  were significant decrements in FEF25-75%, FEF50%, FEF25%, and MVV
14     following all four exposures. The only significant difference between the responses to the
15     1- and 2-h O3 protocols was in FEV0 5. The total number of symptoms reported was 10 for
16     the 1-h FA exposure, 6 for the 1-h O3  exposure, and 12 for both the 2-h FA and 2-h
17     O3 exposures.  It appears that resting ventilation during the 2 h protocol had a lesser effect
18     compared to exercise ventilation. This supports earlier reports that the O3 concentration is
19     the most significant factor among the three factors that contribute to effective dose (Adams
20     et al.,  1981; Folinsbee et al., 1978; Hackney et al., 1975).
21          Available data, though on a limited number of subjects, consistently indicate that
22     responsiveness to O3 is decreased in persons over 50 years of age compared to young adults.
23     Although there are few data available on adults in their thirties and forties, the statistical
24     modeling study of McDonnell et al. (1993) on  subjects from 18 to 32 years of age suggests
25     that responsiveness  to O3 is already diminishing by age 30,  and that the most responsive
26     individuals are likely to be less  than 25 years of age. The results of Bedi et al. (1988)
27     suggest that older adults may be less reproducible in their responses to O3 than young adult
28     males  (McDonnell et al., 1985b); however, this finding is based on only 16 subjects and
29     should be confirmed before being considered conclusive.
30
31

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 1     Ethnic and Racial Factors
 2          Young white males have been the most frequently studied population in published
 3     reports on pulmonary function responses to O3.  There is concern, however, that responses
 4     to O3 may  be influenced by ethnic differences based on the observation that blacks have
 5     smaller lungs than whites for a given standing or sitting height (Rossiter and Weill, 1974;
 6     McDonnell and Seal, 1991). Thus, an equivalent inhaled volume of O3 could result in a
 7     larger O3 dose per unit of lung tissue in blacks compared to whites, potentially inducing
 8     greater effects in blacks than whites exposed to O3 under the same conditions.  Seal et al.
 9     (1993b) evaluated the pulmonary function responses  of 372 individuals, black, white, male,
10     and female (n  > 90 per group), between 18 and 35  years of age who were exposed to 0.00,
11     0.12, 0.18, 0.24, 0.30,  or 0.40 ppm O3. Each subject was randomly assigned to an
12     exposure group and participated in only one experimental session.  The protocol involved
13     environmental chamber  exposure for 2.33 h to the assigned condition.  During the first
14     2 h of exposure, the subjects alternated 15-min rest periods and 15-min exercise periods
                          2
15     (VE = 25  L/min/m BSA).  Spirometric and plethysmographic measurements were made at
16     5 and 20 min following the final exercise period.  The initial nonparametric analysis of the
17     percentage changes in FEVj indicated that FEVj responses increased with increasing
18     O3 concentration, and a group effect occurred that was independent of O3 concentration.
19     There was an O3 effect, but no group effect or group  x O3 interaction for SR^, indicating
20     an increase in SR^ with increasing O3 concentration.  Both group and O3 effects were
21     significant for cough, but  the interaction was not significant. A post hoc analysis, using a
22     different statistical method on the absolute changes in FEVl5 indicated that the black males
23     experienced significant decrements in  FEVj  following exposure to 0.12 ppm O3, whereas
24     black women and white men and women did not have significant decrements in PEVl at
25     O3 concentrations below 0.18 ppm. These results are not  easily explained because there was
26     no gender difference among whites and no racial difference among women. Furthermore,
27     the black men had significantly greater decrements in FEV^ at only some of the
28      O3 concentrations studied (see Figure 7-2),  Although the results can be considered
29      suggestive of an ethnic difference, more subjects must be studied before the issue of ethnic
30      difference in O3 responsiveness can be considered settled.  It should be noted that although
31      this study  included a large number of subjects, each subject participated in only one

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                        0.00        0.12        0.18        0.24
                                         Ozone Concentrator)  (ppm)
                                                             0.30
0.40
       Figure 7-2. Mean percent change (± SFJVQ in post minus pre values of FEV! for each
                   gender-race group.  Open bars =  white women; cross-hatched bars =
                   black women; hatched bars = white men; solid bars = black men.
       Source: Seal et al. (1993).
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
experiment. Thus, the range of individual responsiveness could have been different between
groups.

Environmental Factors
     A number of environmental factors,  such as ambient temperature and humidity, season
of the year, route of inhalation, and smoking history have been hypothesized to potentially
impact on responses to O3 exposure in additive or synergistic ways.  None of these
potentially interacting agents has been adequately addressed  in the extant O3 literature.
Although O3 concentrations in Los Angeles, for example, generally are highest on hot, dry
days, most research on responses to O3 exposure has been conducted under temperature and
humidity conditions  not substantially different from those typical of indoor environments.
The few studies that included temperature  and humidity as experimental factors have
       December 1993
                                               7-51
                                                   DRAFT-DO NOT QUOTE OR CITE

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 1     produced equivocal results (see p.  10-44 in the 1986 O3 criteria document).  No new reports
 2     of temperature or humidity effects have appeared since the 1986 O3 criteria document.
 3          It has been suggested that the effects induced by  oral inhalation of O3 may be greater
 4     than those induced when subjects breathe nasally or oronasally because the scrubbing
 5     function of the nasal passages and upper airways is bypassed.  This issue has potential
 6     importance in comparing results from different laboratories because some groups utilize
 7     environmental chambers with the subjects breathing freely, whereas others use obligatory
 8     mouthpiece exposure systems.  [For additional information on this subject, see Chapter 8.]
 9          Hynes et al. (1988) had 17 nonsmoking, nonallergic individuals (18 to 31  years of age)
10     exercise continuously for 30 min (VE about 30 L/min) while exposed to 0.40 ppm O3.  The
11     subjects completed the protocol once while breathing orally and once while breathing nasally.
12     There was no baseline FA exposure.  No significant difference in any pulmonary function
13     was evident when the oral and nasal exposure responses were compared.
14          Adams et al. (1989) compared the pulmonary function responses to oral and oronasal
15     inhalation of O3.  Six healthy young males (23 to 30 years of age) completed five exposures
16     to 0.40 ppm O3.  There was no FA baseline exposure. Exposures lasted 30 or 75 min,
17     during which the subjects exercised continuously at 75 or 30 L/min (effective dose for all
18     exposures was about 900 ppm • L).  The exposures were presented in random order with at
19     least 4 days between exposures.  Subjects breathed on one occasion through a mouthpiece
20     exposure system while  wearing a noseclip and through a facemask system configured for
21     either oral breathing  only, or oronasal breathing,  on the other four occasions. There were no
22     significant differences in the pulmonary function changes related to the  two facemask
23     configurations.  Group mean decrements in FEVj ranged between 0.67 and 0.92 L (about
24      15 to 17%) among the four exposures. The only significant differences in the FVC and
25     FEVj decrements between the mouthpiece system and the 30 min, 75 L/min facemask
26     configured for oral breathing only was that the decrements with the mouthpiece system were
27     larger.  With the facemask, FEV! decreased 0.70 ± 0.32 L compared to 0.92  ± 0.44 L
28     with the mouthpiece. The subjects did not report any differences in symptomatology among
29     the five exposures.   A potential confounding factor in the facemask studies compared to the
30      mouthpiece exposure is whether or not O3 degrades inside the mask either on the mask
31      surface or by contact with the facial skin, thereby altering the O3 concentration of the inhaled

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 1     air, at least with short-term exposures.  The authors concluded that the lack of significant
 2     difference in FEVj responses between the four facemask exposures supports the contention
 3     that breathing route during moderate and heavy continuous exercise does not alter O3-induced
 4     pulmonary function and subjective symptoms effects in humans.
 5          Earlier studies (see p. 10-38 in the 1986 O3 criteria document) have suggested that
 6     cigarette smokers are less responsive to O3 than nonsmokers.  Frampton et al. (1993)
 7     compared  the pulmonary function responses of 18 current smokers (^3 pack-years smoking
 8     history) and 18  never smokers to 0.22 ppm O3.  All subjects had normal spirometry and no
 9     evidence of exercise-induced bronchospasm.  Exposures were 4 h in duration, during which
10     subjects exercised for 20 min out of each 30 min of exposure (VE = 25 L/min/m2 BSA).
11     Spirometry and  specific airway conductance were measured after 2 h of exposure and at the
12     conclusion of the 4 h exposure period.  Smokers had smaller responses  in all measures  of
13     pulmonary function tested compared to nonsmokers.
14          The  question as to whether reactivity to O3 returns with cessation  of cigarette smoking
15     has been addressed by Emmons and Foster (1991).  Thirty-four individuals with no history of
16     asthma or obvious respiratory  disease who enrolled in a smoking cessation program were
17     randomly  assigned  to an O3 group (n =  18) or an FA group (n = 16).   The  subjects ranged
18     from 24 to 58 years of age, and had a group mean smoking history of 33.9 ± 13 pack-years.
19     Most of the subjects had baseline pulmonary functions somewhat below predicted values,
20     based on age, height, and sex.  Subjects completed 2 h exposures to 0.42 ppm O3 or FA, as
21     assigned, prior  to beginning the smoking cessation program.  The  subjects rested during the
22     exposures, except for 5 min of exercise at 150 kg • m/min (no VE  given) at the beginning of
23     the last half-hour of exposure.  Nine subjects in the O3 group and six in the FA  group
24     completed the 6-mo smoking cessation program  and repeated their assigned exposures at the
25     end of the program.  Prior to beginning the smoking cessation program, both the FA and
26     O3 groups had pre- to postexposure changes  in FVC, FEVj, and FEF25_75% within the
27     variability of repeated tests.  The O3 group had  a significant mean change in  FEF25.75% of
28     -22.5%,  comparing postexposure to preexposure, whereas the FA group had a
29     nonsignificant -12% change.  Changes in FVC and FEVj were not significant in either
30     group.  It should be noted that smoking cessation led to a group mean improvement in
31     baseline FEF25_75% of 22.9%.  The post-O3 exposure values for FEF25_75% were similar

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 1      following the initial and the post-smoking cessation exposures.  Thus, the difference in the
 2      FEF25_75% decrement with O3 exposure post-smoking cessation was largely due to the
 3      improvement that ensued from 6 mo of abstinence from smoking.
 4           The results of Emmons and Foster (1991) and Frampton et al. (1993) suggest that
 5      active smoking blunts responsiveness to O3 and that cessation of smoking for 6 mo leads to
 6      improved baseline pulmonary function and possibly the reemergence of O3 responsiveness.
 7
 8      7.2.1.4  Repeated Exposures to Ozone
 9           Repeated daily exposure to O3 hi the laboratory setting leads to attenuated changes in
10     spirometry and symptom responses that were initially termed "adaptation" (Hackney et al.,
11      1977a). A series of repeated exposure studies, performed in various laboratories, was
12     reviewed in the previous criteria document (U.S. Environmental Protection Agency, 1986).
13     The spirometric responses to repeated O3 exposure typically showed that the response was
14     increased on the second exposure day to concentrations in the range of 0.40 to 0.50 ppm
15     O3 in exposures accompanied by moderate exercise (see Table 7-6). Thus, the response was
16     enhanced on the second consecutive day. Mechanisms for enhanced responses had not been
17     established, although it was hypothesized that persistence of O3-induced damage for greater
18     than 24 h may have contributed to the larger Day 2 response.  An enhanced Day 2 response
 19     was less obvious or absent in exposures that were repeated at lower concentrations or that
20     caused relatively small group mean O3-induced decrements in spirometry.  Two reports (Bedi
 21      et al., 1985; Folinsbee et al.,  1986) indicated that enhanced spirometric responsiveness was
 22      present within 12 h and lasted for at least 24 and possibly 48 h, but was clearly absent after
 23      72 h.  After 3 to 5 days of consecutive daily exposures  to O3, responses were markedly
 24      diminished or absent.  One study (Horvath et al., 1981) suggested that the rapidity of this
 25      decline in  response was related to the magnitude of the subjects' initial responses to O3  or
 26     their "sensitivity".  Finally, the persistence of the attenuation of spirometric and symptom
 27     responses has been studied (Horvath et al.,  1981; Linn et al., 1982b; Kulle et al., 1982).
 28     These studies indicate that the attenuation of response is relatively short-lived, being  partially
 29     reversed within 3 to 7 days and typically abolished within  1 to 2 weeks.  Repeated exposures
 30     separated by 1  week (for up to 6 weeks) do not apparently cause any lessening of the
 31     spirometric response (Linn et al., 1982b).

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               TABLE 7-6.  CHANGES IN FORCED EXPIRATORY LUNG VOLUME AFTER REPEATED
                                             DAILY EXPOSURE TO OZONE3
O" Ozone
*"* Concentration
£
M^*
U>













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 1           Folinsbee et al. (1993a) (also see Devlin et al., 1993b) exposed a group of healthy
 2      males (preliminary n = 8 or 9) to 0.40 ppm O3 for 2 h/day on 5 consecutive days.  Subjects
 3      performed heavy intermittent exercise ( VE = 60 to 70 L/min, 15 min rest/15 min exercise).
 4      Decrements in FEVj averaged 21.8, 28.6, 24.6, 9.1, and 5.5% on the 5 exposure days.
 5      Baseline preexposure FEY^ decreased from the first day's preexposure measurement and was
 6      depressed by an average of about 7% on the third day.  This study illustrates that with high
 7      concentration and heavy exercise exposures, spirometry  and symptom responses are  not
 8      completely recovered within 24 h.
 9           Besides the absence of pulmonary function responses after several days of O3 exposure,
10     symptoms of cough and chest discomfort usually associated with O3 exposure are generally
11      absent (Folinsbee et al., 1993a,b; Linn et al.,  1982; Foxcroft and Adams, 1986;  Folinsbee
12     et al., 1980).  In addition, airway responsiveness to methacholine is increased with an initial
13     O3 exposure (Holtzman et al., 1979; Folinsbee et al., 1988), may be further increased with
14     subsequent exposures (Folinsbee et al., 1993b), and shows  a tendency for the increased
15     response to diminish with repeated exposure (Kulle et al., 1982; Dimeo et al.,  1981).
16     A number of possible explanations for the initially enhanced and then lessened response may
17     be related to changes that are occurring in pulmonary epithelia as a consequence of
18     O3 exposure.  Inflammatory responses (Koren et al., 1989a), epithelial damage, and changes
19     in permeability (Kehrl et al., 1987) could be invoked to explain at least a portion of these
20     responses.  By blocking spirometric and symptom responses with indomethacin pretreatment,
21     Schonfeld et al.  (1989) demonstrated that these responses were not enhanced by repeated
22     exposure.  However, the mechanisms of these responses with regard to repeated exposures in
23     humans remains to be elucidated.
24           Recent studies of repeated O3 exposures have addressed some other features of the
25     responses (see Table 7-7).  A series of reports from the Rancho Los Amigos group  have
26     examined changes in response to O3 as a result of the season of the year in the
27     Los Angeles/South Coast Air Basin. The purpose of this study (Linn et al., 1988; also
 28     Hackney et al.,  1989; Avol et al., 1988) was to determine whether responsive subjects
 29      (n =  12), identified during an initial screening following a period of low ambient O3
 30      exposure,  would remain responsive after regular ambient exposure during the  "smog season".
 31      Responses of so-called "nonresponsive" subjects (n =  13)  were also examined across  the

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         TABLE 7-7. PULMONARY FUNCTION EFFECTS WITH REPEATED EXPOSURES TO OZONE
                                                                              ,a
Ut
Ozone Concentration
	 Exposure Exposure
ppm /ig/m Duration and Activity Conditions
6~l2 235 6.6 h 18~°C
50-min exercise/ 1 0-min 40 % RH
rest, 30-min lunch 5 consecutive
VE = 38.8 L/min daily exposures







018 353 2h JT^C
IE (heavy) 35% RH
VE = 60-70 L/min (Screen exposures
(35 L/min/m BSA) in spring 1986;
second exposures
in summer/fall
1986 and winter
1987 and spring
1987 for
responders and
nonresponders
only)
0.20/0.20 392/392 1 h 21 to 25 °C
0.35/0.20 686/392 CE at 60 L/min 40-60% RH
0.35/0.35 686/686 (three 2-day pairs
of exposures)






Number and Subject
Gender of Subjects Characteristics
17 M Healthy NS










59 adult Responders:
Los Angeles Age =
residents 19-40 years
12 responsive 6 atopic
13 nonresponsive 2 asthmatic
4 normal

Nonresponders:
Age =
18-39 years
13 normal

15 M Healthy
aerobically
trained NS
FVC = 4.24 -
6.98 L





Observed Effect(s) Reference
FEVj responses were maximal on first day Folinsbee et al. (1993b)
of exposure (- 13 %), less on second day (also see Table 7-9)
(—9%), absent thereafter. Symptom
responses only the first 2 days.
Methacholine airway responsiveness was at
least doubled on all exposure days, but was
highest on the second day of ozone. Airway
responsiveness was still higher than air
control after 5 days of ozone exposure.
Trend to lessened response, but it was not
achieved after 5 days.
Responders had ApEVj = 12.4% after Linn et al. (1988)
initial screening, nonresponders had no (also see Hackney et al., 1989)
change. Responders had nonsignificant
response in late summer or early winter, but
were responsive again in early spring (spring
1986, -385 mL; Autumn 1986, -17 mL;
winter 1987, +16mL; spring 1987,
—347 mL). Nonresponders did not change
with season. Suggests that responders
responses may vary with ambient exposure,
but nonresponders generally remain
nonresponsive.
Consecutive days of exposure to 0.20 ppm Brookes et al. (1989)
produced similar responses on each day
(-5.02, -7.80); 0.35/0.20 ppm pair caused
increased response to 0.20 ppm on second
day (-8.74); 0.35/0.35 ppm caused much
increased response on day 2 (—15.9,
-24.6). Symptom responses were worse on
the second exposure to 0.35 ppm, but not
with second exposure to 0.20 ppm.
-------
d
8
n>
TABLE 7-7 (cont'd). PULMONARY FUNCTION EFFECTS WITH REPEATED
EXPOSURES TO OZONE3
Sg Ozone Concentration
S PPm ^
6 mo
Healthy NS
61 years for M
and 65 years for
F
FVC = 4.97 L
for M and
3.11 LforF
Healthy NS
60-89 years
(median age =
65 years; mean
FVC = 3.99 L;
mean FEVj =
3. OIL;
FEVj/FVC range
= 61-85%)

Observed Effect(s) Reference
Largest FEVj decrease on second of 4 days Foxcroft and Adams (1986)
Oj exposure (—40% mean 1). Trend for
adaptation not complete in 4 days.
VO2tnax decreased with single acute 03
exposure (—6%), but was not significant
after 4 days of 03 exposure (-4%).
Performance time was less after acute 03
(211 s) exposure than after FA (253 s).
No differences between responses to Schonfeld et al . (1989)
exposures separated by 72 or 120 h.
Enhanced FEVj response at 24 h (- 16.1 %
versus —30.4%). Possible enhanced
response at 48 h (—14.4% versus
—20.6%). Similar trends observed for
respiratory pattern and SRaw.
Spirometric changes were not reproducible Bedi et al. (1988)
from time to time after ozone exposure
(r < 0.50). Repeat exposures to air
yielded consistent responses.
Overall increase in symptoms, but no single Bedi et al. (1989)
symptom increased significantly. FVC
decreased 111 mL and 104 mL on Days 1
and 2, respectively. FEVj fell by 171 and
164 mL and FEV3 fell by 185 and
172 mL. No significant changes on Days 3
and 4 or with FA. FEVj changes were
-5.8%, -5.6%, -1.9%, and -1.7% on
the four 03 days.

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                     TABLE 7-7 (cont'd).  PULMONARY FUNCTION EFFECTS WITH REPEATED
                                             EXPOSURES TO OZONEa

§, Ozone Concentration

I^Q ppm Mg/m Duration and Activity
U> 0.45 882 2h
(+ 0.30 IE (20 min rest, 20 min
PAN) exercise)
VE = 27 L/min







Exposure
Conditions
22 °C
60% RH
Five days
consecutive
exposure to PAN
+ 0,




Number and
Gender of Subject



Subjects Characteristics Observed Effect(s) Reference
3 M Healthy NS
5 F Mean age =
24 years






FEVj decreased =19% with 03 alone, Drechsler-Parks et al. (1987b)
* 15% on Day 1 of 03 + PAN, =5% on (also see Table 7-13)
Day 5 of Oj + PAN, =7% 3 days after
5 days of 03 + PAN, =15% after 5 days
of 03 + PAN. Similar to O3 adaptation
studies, 03 responses peaked after 2 days,
were depressed 3 days later, and responses
returned 7 days later. PAN probably had
no effect on adaptation to 03.
    See glossary of terms and symbols for abbreviations and acronyms.
    Listed from lowest to highest 03 concentration.
    Age range in years or as mean + SEM.
71
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 1     year.  The subjects were exposed to 0.18 ppm O3 on four occasions, spring, fall, winter, and
 2     the following spring.  Only 17 subjects (8 responders) participated hi the final spring
 3     exposures.  The marked difference in FEVj response seen initially (-12.4% versus +1%)
 4     was no longer present after the summer smog season (fall test) or 3 to 5 mo later (whiter
 5     test).  However, when the reduced subset of subjects was exposed during the following
 6     spring, the responsive subjects again had significantly larger changes in FEVj.  Seasonal
 7     changes in FEVj response to O3 in the responsive and nonresponsive subjects are shown
 8     below.
 9
10                        AFEV^ Spring     AFEVj Fall     AFEX^ Winter    AFEVj Spring
       	(mL)	(mL)	(mL)	(mL)
11      Responders            -385            -17             +16             -347
12      Nonresponders         +28             +90             +34             +81
13     =^^^======^^=====^^
14
15          These results suggest a seasonal variability hi response that may be attributed to
16     increased ambient O3 exposure during the summer months.  It must be noted that the
17     "responders"  included subjects  who had a history of complaints from ambient air pollution.
18     Furthermore, this group included a significant proportion of allergic individuals whose
19     seasonal allergies could have contributed to their varying responses. Historically, however,
20     studies with the subjects drawn from the population of Los Angeles residents have reported
21     reduced responses to O3 exposure in the laboratory compared to nonresidents (Hackney
22     etal., 1976,  19775).
23          Brookes et al.  (1989) recently reexamined a  hypothesis previously tested 5y Gliner
24     et al. (1983), namely that repeated exposure to one concentration can alter response to
25     subsequent exposure to a different O3 concentration.  Gliner et al.  (1983) had previously
26     shown that the response to 0.40 ppm O3 was not influenced by previously being exposed to
27     0.20 ppm O3 for 2 h on 3 consecutive days. Brookes et al. (1989) tested whether exposure
28     to 0.20 or 0.35 ppm O3 would change subsequent response to 0.20 or 0.35 ppm O3.  They
29     found increased  responses  to 0.20 ppm for both preexposures  (AFEVj =  -5.02%, -7.80%,
30     and -8.74% foF 0.20 ppm acutely,  0.20 ppm after 0.20 ppm, and 0.20 ppm after 0.35 ppm,
31     respectively), but this trend was significant only for the higher concentration.  Although not

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 1      statistically significant, the response increase seen on the second exposure day at 0.20 ppm is
 2      similar to that seem by Gliner et al. (1983).  These observations suggest that although
 3      preexposure to low concentrations of O3 may not influence response to higher concentrations,
 4      preexposure to a high concentration of O3 may significantly increase response to a lower
 5      concentration on the following day.
 6           Schonfeld et al. (1989) confirmed previous observations of Bedi et al. (1985) and
 7      Folinsbee et al. (1986) that the period of enhanced responsiveness to Oj following an initial
 8      exposure persists for about 24 to 48 h,  but is absent by 72 h after the initial exposure.  In a
 9      series of paired exposures to 0.35 ppm  with continuous heavy exercise separated by intervals
10     of 1, 2, 3,  or 4 days, they found that the responses to the second exposure were clearly
11      increased at 24 h (AFEVj  = —16.1% and  —30.4% for the first and second exposures,
12      respectively) and possibly also at 48 h (^FEVl = -14.4% and -20.6%).  Similar trends
13      were observed for other physiological variables such as SRaw and respiratory pattern during
14     exercise. With a 3- or 4-day interval between exposures, the responses to the two exposures
15      were similar.
16          Foxcroft and Adams (1986) demonstrated that decrements in exercise performance seen
17     after a 1-h exposure to 0.35 ppm (continuous heavy exercise) were less after 4 consecutive
18     days of O3 exposure than they were after a single acute exposure.  Maximal aerobic power
19     and performance time on a progressive bicycle exercise test were reduced 6%  and
20     42  s respectively, from FA control, after a single 0.35 ppm exposure.  After 4 consecutive
21      days of 1-h exposures, the maximal aerobic power was reduced only 4% and the
22     performance time by only 14 s; these differences from FA control were not statistically
23     significant.  Despite the change in exercise performance, Foxcroft and Adams (1986) did not
24     show the attenuation of FEVj response seen in many previous studies (Folinsbee et  al., 1980;
25     Linn et al., 1982b). However, these investigators selected known O3-sensitive subjects
26     whose FEY^ decrements exceeded 30% on the first 3  days of exposure.  The large
27     magnitude of these responses,  the trend for the responses to decrease on the third and fourth
28     day, the decreased  symptom responses,  and the observations of Horvath et al.  (1981) that
29     O3-sensitive subjects adapt more slowly, suggest that attenuation of response would have
30     occurred if the exposure series had been continued for another 1 or 2 days.  These
31     observations support the contention advanced by Horvath et al. (1981) that the progression of

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 1     attenuation of response is a function of "O3 sensitivity." Furthermore, these results suggest
 2     that exercise responses after O3 exposure may be limited, either voluntarily  or involuntarily,
 3     more by subjective symptoms than by alterations in gas exchange consequent to changes in
 4     ventilatory function.
 5          Bedi et al. (1989) examined the responses of elderly subjects (median age,  65 years) to
 6     four exposures to 0.45 ppm O3 for 2 h with mild intermittent exercise.  The first three
 7     exposures were on consecutive days, with the fourth  exposure following the third by 3 days.
 8     Changes in FEVj on the first two exposure days averaged —5.8% and  —5.6%,  about half
 9     the response expected in a group of healthy young males (-12.7%; Folinsbee et al., 1978).
10     There were no significant changes in FEY^ on the third (-1.9%) and fourth (-1.7%)
11     exposure days.  Symptom responses were negligible, although there was an overall increase
12     in symptoms  on the first day of O3 exposure compared to air exposure.  Despite the high
13     concentration of the exposure, there was no enhancement of the spirometry  response on the
14     second day of exposure.  Although similar observations have been made in  previous studies
15     producing small changes in spirometry (Folinsbee et  al., 1980, 1993a) with repeated
16     exposures, the responses of older subjects are not sufficiently understood to explain these
17     responses.  Bedi et al. (1988) had previously reported that responses to O3  in the older
18     subjects tended to be less reproducible, although this observation alone could not explain
19     these responses.
20          Drechsler-Parks et al. (1987b) examined response to repeated exposures to 0.45 ppm
21     O3 plus 0.30 ppm peroxyacetyl nitrate (PAN). Exposures to O3 and O3 plus PAN yielded
22     similar changes in spirometry  (AFEVj =  —19% and —15%, respectively).  Thus, PAN did
23     not increase responses to O3.  Repeated exposure to  the PAN plus O3 mixture resulted in
24     similar changes as those that would be seen with Oj  exposure alone. Responses in FEVj
25     exceeded -30%  on the second exposure and fell to less than -5% after the fifth day.  The
26     attenuation of response persisted 3 days after the repeated exposures, but was absent after
27     7 days.  These observations suggest that PAN does not influence the attenuation response to
28      repeated O3 exposure. If the PAN responses are  considered negligible, this study confirms
29      the observation that the attenuation of O3 responses with chamber exposures lasts no longer
 30      than 1 week.
        December 1993                           7-62      DRAFT-DO NOT QUOTE OR CITE
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 1          Repeated prolonged exposure to low concentrations of O3 has recently been examined
 2     (Horvath et al., 1991; Folinsbee et al., 1993b; Linn et al., 1991, 1992).  Horvath et al.
 3     (1991) exposed subjects for 2 consecutive days to 0.08 ppm using the 6.6-h prolonged-
 4     exposure protocol (see Section 7.2.2).  They observed small pre- to postexposure changes in
 5     FEVj (—2.5%) on the first exposure, but no change on the second day.  Linn et al. (1993)
 6     observed a 1.7% decrease in FEVj after a 6.5-h exposure to 0.12 ppm in healthy subjects.
 7     A second consecutive exposure yielded even smaller responses.  With exposure to a mixture
 8     of O3 plus 100 jitg/m3 of sulfuric acid aerosol, there was a 4.2% decrease in FEVl on the
 9     first exposure day. In a group of asthmatics exposed under similar conditions, the FEVj
10     response on the first day was -8.6% (O3) and -11.6% (O3 plus acid).  After adjustment for
11     the  exercise effect (-4.6%), the responses (-4% and -7%) were still greater than those of
12     nonasthmatics.  Responses were slightly reduced on the second day of exposure.
13          Folinsbee et al. (1993b) exposed 17 subjects to 0.12 ppm O3 for 6.6 h on 5 consecutive
14     days.   Spirometry responses were typified by changes in FEV1 that reached  -13% on the
15     first day and -9% on the second day of exposure.  No significant differences in spirometry
16     responses between FA and subsequent O3 exposures were observed.  Symptom responses
17     were also greatest on the first exposure day and were largely absent  from the third day on.
18     Methacholine responsiveness was tested using a single dose of methacholine and then
19     comparing changes (as a ratio) in airway resistance between methacholine and saline
20     aerosols.  The responses to FEVj and methacholine testing are shown below.
21          Methacholine responsiveness was increased (over the clean air  response) throughout the
22     5 days of O3 exposure, although it reached a peak on the second day and there was a trend
23     for responsiveness to decrease after 5 days in some subjects.  These results suggest that
24     repeated exposure to low levels of O3, despite the  attenuation of symptoms and pulmonary
25     function changes, is not without hazard.  It is likely that some epithelial damage persists that
26     contributes to the enhanced response to methacholine throughout the exposure series.
27     However, it must be noted that, in this study,  subjects were initially selected based on their
28     FEVj  response to 0.16 ppm O3 for 4 h.   This may in part explain the greater FEVt
29     responses seen in this study, but there was no correlation between individual FEVj
30     decrements and changes in methacholine responsiveness.  Furthermore, the Horvath et al.
31     (1991) subjects were exposed to only 0.08 ppm and they  were somewhat older than the

       December 1993                           7-63      DRAFT-DO NOT QUOTE OR CITE
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 1      Folinsbee et al. (1993b) subjects; the Linn et al. (1991) subjects, on the other hand, had
 2      lower ventilation during exercise and were residents of Los Angeles accustomed to exposure
 3      to these levels of O3 (see Chapter 4 for typical O3 concentrations).
 4
5
6
7
8
9
10

A%FEV,
SRaw Ratio
Based on
Day
-12.
3.
1
79
67
studies cited
Day 2
-8.73
4.55
here and in
Day 3
-2.54
3.99
the previous
Day
-0.
3.
4
6
24
criteria document
Day
+0.
3.
5
Clean
2 +1.
74 2.
(U.S.
Air
1
22
Environmental
11     Protection Agency, 1986), several conclusions can be drawn about repeated  1- to 2-h
12     O3 exposures.  Repeated exposures to O3 can cause an enhanced (i.e., greater) response on
13     the second day of exposure. This enhancement of response appears to be dependent on the
14     interval between the exposures (24 h causes the greatest increase) and is absent with intervals
15      >3  days.  An enhanced response also appears to depend to some extent upon the magnitude
16     of the initial response.  Small responses to the first O3 exposure are less likely to result in an
17     enhanced response on the second day of O3 exposure. Repeated daily exposure also results
18     in attenuation of spirometric responses, typically after 3 to 5 days of exposure.   This
19     attenuated response persists for less than 1 or possibly 2 weeks.  In temporal conjunction
20      with the spirometry changes, symptoms induced by O3, such as cough and chest discomfort,
21      are  also attenuated with repeated exposure.  Ozone-induced changes in airway responsiveness
22      attenuate somewhat more slowly than spirometric and symptom responses.  Attenuation of
23      the  changes in airway responsiveness may also persist longer than changes in spirometry,
24      although this has only been studied on a limited basis.  In longer duration-lower
25      concentration studies that do not cause an enhanced second-day response, the attenuation of
26      response to O3 appears to proceed more rapidly.
27
28     7.2.1.5   Effects on Exercise Performance
 29     Introduction
 30           An early epidemiological study examining race performances in high  school cross-
 31      country runners (Wayne et al.,  1967) suggested that exercise performance is depressed by
 32     inhalation of ambient oxidant air pollutants.  Wayne et al.  (1967) suggested that the
        December 1993                           7-64      DRAFT-DO NOT QUOTE OR CITE
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 1     detrimental effects of oxidant air pollutants on race performance may have been related to
 2     increased airway resistance or to the associated discomfort in breathing, thus limiting
 3     runners' motivation to perform at high levels.  The effects of acute O3 inhalation on exercise
 4     performance have been evaluated in numerous controlled human studies.  These studies can
 5     be divided into two categories, those that examine the effects of acute O3 inhalation on
 6     maximal oxygen uptake and those that examine the effects of acute O3 inhalation on the
 7     ability to complete strenuous continuous exercise protocols up to 1 h in duration. Five
 8     studies (Folinsbee et al., 1977b; Horvath et al., 1979; Folinsbee et al.,  1984; Adams and
 9     Schelegle, 1983; Savin and Adams, 1979) examining the affects of acute O3 exposures on
10     exercise performance were discussed in the 1986 EPA criteria document (U.S.
11     Environmental Protection Agency, 1986). This section  summarizes the studies reviewed in
12     that document and reviews newer studies published since that examine the effect of acute
13     O3 inhalation on (1) maximal oxygen uptake and (2) endurance performance.  Studies are
14     also summarized in Table 7-8.
15
16     Effect on Maximal Oxygen Uptake
17          Three studies (Folinsbee et al., 1977b; Horvath et al.,  1979; Savin and Adams, 1979)
18     examining the effects of acute O3 exposures on VO2max  were discussed in the  1986 EPA
19     criteria document (U.S. Environmental Protection Agency, 1986). Of these studies, only one
20     showed a reduction in VO2max.  Folinsbee et al. (1977b) observed that VO2max was
21     significantly (p  < 0.01) decreased 10.5% following a 2-h exposure to 0.75 ppm O3 with
22     light IE. Reductions in VO2max were accompanied by a 9.5% decrease in maximum
23     attained workload (p  < 0.01), a  16% decrease in maximum ventilation (p < 0.01), and a
24     6% decrease in maximum heart rate (p < 0.05).  The 16%  decrease in maximum ventilation
25     was associated with a 21 % decrease in VT.  In addition, the O3 exposure resulted in a
26     22.3% decrease in FEVj  and subjective symptoms of cough and chest discomfort.
27     In contrast, Horvath et al. (1979) did not observe a change in VO2max or other maximum
28     cardiopulmonary endpoints in male and female subjects  exposed at rest to 0.75 ppm O3 for
29     2 h, although FVC was significantly decreased 10% (p  < 0.05).  Similarly, Savin and
30     Adams (1979) observed no effect on maximum attained workload or VO2max in nine
31     subjects exposed to 0.30 ppm O3 while performing a progressively incremented exercise test

       December 1993                           7.55      DRAFT-DO NOT QUOTE  OR CITE
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TABLE 7-8. OZONE EFFECTS ON EXERCISE PERFORMANCE3
g Ozone
g. Concentration
cs
>— ppm
S 0.00
0.06-
0.07
0.12-
0.13
0.00
0.12
0.18
0.24
0.00
0.12
0.20
ON 0.00
°^ 0.20
0.35
O
S 0.00
jri °-2i
i
y o.oo
U 0.25
§ 0.00
H 0-25
o °-50
C °'75
Exposure
Duration and Exposure
fig/m Activity Conditions
0
120-
140
245-
260
0
235
353
470
0
235
392
0
392
686
0
412
0
490
0
490
980
1,470
CE ( VE = Tdb = 23-24.5 °C;
30-120 L/min) RH = 50-53%
16-28 min
progressive
maximum
exercise protocol
1 h competitive Tdb = 23-26 °C
simulation RH = 45-60%
exposures at
mean VE =
87 L/min
1 h CE Tdb = 31 °C
VE = 89 L/min
1 h CE or Tdb = 23-26 °C
competitive RH = 45-60%
simulation at
mean VE = 77.5
L/min
1 h CE at 75% Tdb = 19-21 °C
VO2max RH = 60-70%
1 h CE Tdb = 20 °C
VE = 63 L/min RH = 70%
2 h rest NA
Number
and
Gender of Subjects
Subjects Characteristics
12 M Athletic
12 F
10 M Highly trained
competitive
cyclists
15 M Highly trained
2 F competitive
cyclists
10 M Well-trained
distance runners
6 M Well-trained
1 F cyclists
19 M Active
7 F nonathletes
8M
5F
Observed Effect(s) References
Reduced maximum performance time and increased Linder et al. (1988)
respiratory symptoms during 03 exposure.
Decrease in exercise time of 7.7 min and 10.1 min Schelegle and Adams (1986)
for subjects unable to complete the competitive
simulation at 0.18 and 0.24 ppm 03, respectively;
decrease in FVC and FEVj for 0.18- and 0.24-ppm
03 exposure compared with FA exposure.
Decrease in VEmax, VO2max, VTmax, workload, Gong et al. (1986)
ride time, FVC, and FEVj with 0.20 ppm 03
exposure, but not significant with 0.12-ppm 63
exposure, as compared to FA exposure.
VT decreased and f increased with continuous 50- Adams and Schelegle (1983)
min 03 exposures; decrease in FVC, FEVj, and
FEFy.75% from FA to 0.20 ppm and FA to 0.35
ppm O3 exposure in all conditions; three subjects
unable to complete continuous and competitive
protocol at 0.35 ppm 03.
Decrease in FVC, FEVj, FEF25.75sj, and MW Folinsbee et al. (1984)
with 0.21 ppm Oj compared with FA exposure.
FVC, FEVj, and MW all decreased with 0.25 ppm Folinsbee et al. (1986)
Oj exposure compared with FA.
FVC decreased with 0.50- and 0.75-ppm 03 Horvath et al. (1979)
exposure compared with FA; 4% nonsignificant
decrease in mean VO2max following 0.75 ppm 03
compared with FA exposure.
-------
                           TABLE 7-8 (cont'd).  OZONE EFFECTS ON EXERCISE PERFORMANCE8
g Ozone
1" Concentration1" Exoosure
JJj
f£ ppm
S °-00
0.35


0.00
0.75



Duration and Exposure
jUg/m Activity Conditions
0 50 min CE NA
686 VE = 60 L/min


0 2 h IE NA
1,470 (4 X 15 min light
[50 W] bicycle
ergometry)

Number
and
Gender of Subjects
Subjects Characteristics
8 M Trained
nonathletes


13 M 4 light S,
9NS





Observed Effect(s)
VT decreased, f increased with 50-min O3
exposures; decrease in FVC, FEVj, FEF25-75%>
performance time, VO2max, VEmax, and HR,,^
from FA to 0.35 ppm Oj exposure.
Decrease in FVC, FEVj, ERV, 1C, and FEF50%
after 1-h 0.75 ppm 03 exposure; decrease in
VO2inax, Vijinax, Vgmax, maximal workload
and heart rate following 0.75 ppm 03 exposure
compared with FA.


References
Foxcroft and Adams (1986)



Folinsbee et al. (1977b)




     See glossary of terms and symbols for abbreviations and acronyms.
     Listed from lowest to highest 03 concentration.
<1
-0
-------
 1     to volitional fatigue lasting 30 min. In addition, Savin and Adams (1979) observed no
 2     significant effect on pulmonary function, performance time, maximum heart rate,  or
 3     anaerobic threshold,  although maximum ventilation was significantly reduced 7%.
 4          More recent findings of Foxcroft and Adams (1986) and Gong et al. (1986)  support the
 5     earlier observations of Folinsbee et al. (1977b).  Foxcroft and Adams (1986) observed
 6     significant (p  < 0.05) reductions in performance time (16.7%), VO2max (6.0%), maximum
 7     ventilation (15.0%),  and maximum heart rate (5.6%) in eight aerobically trained males
 8     during a rapidly incremented VO2max test following 50 min exposure to 0.35 ppm O3 with
 9     CE (exercise  VE =  60 L/min).  Similarly, Gong et al. (1986)  found significant (p < 0.05)
10     reductions in performance tune (29.7%), VO2max (16.4%), maximum ventilation (18.5%)
11     and maximum workload (7.8%) in 17 top-caliber endurance cyclists during a rapidly
12     incremented VO2max test following 1-h exposure to 0.20 ppm O3 with very heavy
13     CE (VE = 90 L/min) and the addition of ambient heat stress (31 °C).  In both studies
14     (Foxcroft and Adams, 1986; Gong et al., 1986),  the reductions in maximal exercise
15     endpoints were accompanied by significant decrements in pulmonary function and marked
16     subjective symptoms of respiratory discomfort.  More recently, Linder et al. (1988) observed
17     small decrements in performance time during a progessive maximal exercise test at
18     O3  concentrations as low as 0.06 ppm. These small effects were associated with increased
19     respiratory symptoms and small, inconsistent changes in FEVj.  Hence, it appears that
20     maximal oxygen uptake is reduced if it is preceded by an O3 exposure entailing a sufficient
21     total inhaled dose of O3 to result in significant pulmonary function decrements and/or
22     subjective symptoms of respiratory  discomfort.
23
24     Effect on Endurance Exercise Performance
25           Two  studies (Adams and Schelegle, 1983; Folinsbee et al., 1984) that addressed the
26     effects of acute O3 exposures on the ability of highly trained subjects to complete strenuous
27     continuous exercise protocols were discussed in the 1986 EPA criteria document (U.S.
28     Environmental Protection Agency,  1986).
29           Adams and Schelegle (1983) exposed 10 well-trained distance runners to filtered air,
30     0.20 ppm O3, and 0.35 ppm O3 while the runners exercised on a bicycle ergometer at
31     workloads simulating either a 1-h steady-state "training"  bout or a 30-min warm-up followed

        December 1993                          7-68      DRAFT-DO NOT QUOTE OR CITE
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 1     immediately by a 30-min "competitive bout".  The exercise levels in the steady-state training
 2     bout were of sufficient magnitude (68% of their VO2max) to increase mean VE to 80 L/min.
 3     The  VE averaged over the entire competitive simulation was also 80 L/min, whereas the
 4     mean VE during the 30-min competitive bout was 105 L/min. Subjective symptoms
 5     increased as a function of O3 concentration for both training and competitive protocols.
 6     In the competitive protocol, four runners exposed to 0.20 ppm O3 and nine exposed to
 7     0.35 ppm O3 indicated that they could not have performed maximally.  Three subjects were
 8     unable to complete  both the training and competitive protocols at 0.35 ppm O3, and a fourth
 9     failed to  complete only the competitive ride.
10          Folinsbee et al.  (1984) exposed six well-trained  men and one well-trained woman to
11     0.21 ppm O3 while they exercised continuously on a bicycle ergometer for 1 h at 75% of
12     their VO2max (VE = 81 L/min).  Following O3 exposure, FVC and FEVl were
13     significantly reduced and the subjects reported  symptoms of laryngeal and/or trachea!
14     irritation and chest  soreness and tightness upon taking a deep breath.  Anecdotal reports
15     obtained  from  the cyclists suggested that their performance would have been limited if they
16     experienced similar symptoms during competition.
17          Avol et al.  (1984) exposed 50 well-conditioned cyclists to 0.0, 0.08,  0.16, 0.24, and
18     0.32 ppm O3 for 1  h in ambient heat (32 °C) while exercising continuously
19     (VE = 57 L/min).  Reductions in FEVj and symptoms increased in a concentration-
20     dependent manner,  being initially detected at 0.16 ppm O3.  Three and 16 cyclists could not
21     complete the 1-h exposure to 0.16 and 0.24 ppm O3,  respectively, without a reduction in
22     workload.  Similarly, in their study of the effects of O3 exposure on  VO2max, Gong et al.
23     (1986) reported that 6 of 17 highly trained endurance cyclists were not able to complete  1-h
24     exposure to 0.20 ppm O3 with very heavy CE  (VE = 90 L/min) and the addition of ambient
25     heat stress (31 °C).
26          In a study designed to determine the effects of the inhalation of low ambient
27     O3 concentrations on simulated competitive endurance performance, Schelegle and Adams
28     (1986) exposed 10  highly trained endurance athletes to 0.12, 0.18, and 0.24 ppm O3 while
29     performing a  1 h "competitive" protocol. The competitive protocol used in this study was
30     similar to that used by Adams and Schelegle (1983) except that the workload during the final
31     30 min competitive bout was more intense, being selected based on the maximum workload

       December 1993                          7.59      DRAFT-DO NOT QUOTE OR CITE
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 1     (approximately 86% of their VO2max, mean VE =  120 L/min) each subject could maintain
 2     for 30 min while breathing FA.  All subjects completed the FA exposure, whereas one, five,
 3     and seven subjects could not complete the 0.12-, 0.18-, and 0.24-ppm Oj exposures,
 4     respectively.  Following 0.18- and 0.24-ppm O3 exposures, FVC  and FEVj were
 5     significantly reduced (p < 0.05) and subjective symptoms  were significantly elevated
 6     (p <  0.05).  No significant effect of O3 was found for metabolic or ventilatory pattern
 7     responses.  Similarly, Folinsbee et al. (1986) found that highly trained runners experienced a
 8     reduced run time on a treadmill (speed and grade set at approximately 80% of their subjects
 9     VO2max) when exposed to 0.18 ppm O3 compared with filtered air. These subjects did
10     have significantly elevated symptoms of respiratory discomfort and significantly decreased
11     FVC and FEVl5 whereas arterial oxygen saturation at the end of the run was not affected by
12     O3 exposure.
13          Determining the mechanisms leading to the observed decrements in maximal oxygen
14     uptake and the inability to complete strenuous exercise protocols is problematic.  As stated
15     by Astrand and Rodahl (1977) "the capacity for prolonged rhythmic muscular exercise is
16     limited by an interrelated composite of cardiorespiratory,  metabolic, environmental, and
17     psychological factors."  Many investigators cited above have concluded that the observed
18     reductions in exercise performance appeared to be due to symptoms limiting the ability of
19     their  subjects to perform.  However, in every case,  this is a conclusion achieved by
20     exclusion and not by the demonstration of a causal relationship.  Other  factors could also
21     contribute to O3-induced decrements in exercise performance.  One possibility is that
22      stimulation of neural receptors in the airways may result in an inhibition of alpha-motor
23      nerve activity to respiratory muscles during inspiration (Koepchen et al., 1977; Schmidt and
24      Wellhoner, 1970), resulting in the observed decrease in VT and at the same time increasing
25      the subject's sensation of respiratory effort.  This mechanism would not be directly related to
26      symptoms of discomfort,  but because of the common role of airway neural afferents,  may be
27      difficult to discern from the  affects of symptoms of respiratory discomfort.  Indeed, a reflex
 28      inhibition of the ability to inspire would be consistent with the reduced VTs following
 29      O3 exposure in subjects performing  maximal exercise and would be consistent with the
 30     development of a physiologically induced ventilatory limitation to maximal oxygen uptake.
 31

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 1     7.2.2   Pulmonary Function Effects of Prolonged (Multihour) Ozone
 2             Exposures
 3          Since 1988, a series of studies have described the responses of subjects exposed to
 4     relatively low O3 concentrations from 0.08 to 0.16 ppm for durations of 4 to 8 h (see
 5     Table 7-9). These studies have demonstrated statistically significant changes in spirometry,
 6     airway resistance, symptoms, and airway responsiveness during and after such exposures.
 7          The only related study to be cited in the previous criteria document (U.S.
 8     Environmental Protection Agency, 1986) was that of Kerr et al. (1975), who exposed
 9     subjects for 6 h to 0.50 ppm, but with only two brief 15-min periods of moderate exercise
10     during the exposure.  Small changes in spirometry were observed.  Because of the minimal
11     exercise level and the high O3 concentration, these results cannot be compared to the more
12     recent  studies.
13          The first prolonged exposure study involving low concentrations and a substantial
14     amount of moderate exercise  was reported by Folinsbee et al. (1988). The basic protocol
15     used by these investigators has been used in a number of subsequent investigations and is
16     worth describing in some detail. The exposures lasted 6 h and 35 min (i.e., 6.6 h). Except
17     for a 35-min lunch break (during which 03 exposure continued at rest) after 3 h, the subjects
18     exercised at a moderate level (with a ventilation of about 40 L/min) for 50 min of each hour.
19     Pulmonary function tests were conducted during the 10-min rest period and at the beginning
20     and end of exposure. The exposure was intended to simulate a day of heavy outdoor work
21     or play. For convenience, this protocol is referred to as the EPA prolonged-exposure
22     protocol.
23          In this study (Folinsbee et al., 1988), a group of 10 subjects was exposed to clean air
24     and 0.12 ppm O3 for 6.6 h.  Forced vital capacity and FEVl decreased in a roughly linear
25     fashion throughout the exposure and had fallen by 8.3 and 13%, respectively, by the end of
26     the exposure.  Symptoms of cough and chest discomfort were increased and airway
27     responsiveness to methacholine was approximately doubled after O3 exposure.  There was
28     a wide range  of response and three subjects had FEVj  decrements of 25% or greater,
29     whereas the three least sensitive subjects had less than 5% change in FEVj.
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2

I

§
                 TABLE 7-9.  PULMONARY FUNCTION EFFECTS AFTER PROLONGED EXPOSURES TO OZONE3
Ozone Concentration
ppm
0.08
0.10
0.12




Hg/m Duration and Activity Conditions
157 6.6 h 18 °C
196 IE (6 x 50 min) 40% RH
235 VE « 39 L/min




Number and
Subjects Characteristics Observed Effect(s)
22 M Healthy NS FVC and FEVj decreased throughout the
exposure; FEVj decrease at end exposure
was 7.0, 7.0, and 12.3%, respectively.
FEVj change > 15% occurred in 3, 5, and
9 subjects at 0.08, 0.10, and 0.12 ppm,
respectively. Methacholine responsiveness
increased by 56, 89, and 121 %, respectively.

Reference
Horstman et al. (1990)






                              See Horstman et al. (1990)
                              and Folinsbee et al. (1988)
A lognormal model was fitted to FEVj data. Larsen et al. (1991)
Model parameters indicate Oj concentration
had greater effect than  VE or duration
(estimated exponent for [Oj] = 4/3).
                              See Folinsbee et al. (1993b)
See Folinsbee et al. (1993b). Slight
increases in clearance of ^T"c-DTPA
occurred after 03 exposure (t = 10%).
Suggests possible increase in permeability.
Kehrl et al. (1991)

0.08
0.10
0.08
0.12
(a) 0.12
(b) Varied from 0.0
to 0.24 (increased
by 0.06 ppm/h then
decreased by
0.06 ppm/h)
See McDonnell et al. (1991a)
157 6.6 h
196 IE (6 x 50 min)
VE = 40 L/min
157 6.6 h
IE (6 x 50 min)
VE = 35-38 L/min
(1 day of air, 2 days of Oj)
235 6.6 h
IE (6 x 50 min)
VE = 42.6 L/min
235 8h
IE (8 x 30 min)
VB = 40 L/min

18 °C
40% RH
25 °C
48% RH
18 °C
40% RH
(1 exp. to clean
air; 1 exp to 03)
22 °C
40% RH
<3 ftg/m TSP

38 M Healthy NS
Mean age =
25 years
5 F Healthy NS
6 M Age = 30-45 years
10 M Healthy NS
23 M Healthy NS
No change in ySmTc-DTPA clearance. Kehrl et al. (1989b)
Possible masking of response by exercise.
FEVj, decreased 8.4% at 0.08 ppm and McDonnell et al. (1991a)
11.4% at 0.10 ppm. Symptoms of cough,
PDI, and SB increased with Oj exposure.
FVC decreased 2.1%, FEVj decreased 2.2% Horvath et al. (1991)
on first day of Oj exposure; no change on
second Oj day.
FEVj decreased by 13 % after 6.6 h. FVC Folinsbee et al. (1988)
dropped 8.3 % . Cough and PDI increased
with Oj exposure. Airway responsiveness to
methacholine doubled after 03 exposure
(t 100%).
(a) FEVj decreased 5% by 6 h and remained Hazucha et al. (1992)
at this level through 8 h.
(b) FEVj change mirrored 03 concentration
change with a lag time of ~ 2 h. Max
decrease of 10.2% after 6 h. FEVj change
was reduced in last 2 h of exposure.
-------
O TABLE 7-9 (cont'd). PULMONARY FUNCTION EFFECTS AFTER PROLONGED EXPOSURES TO OZONE4
3 Ozone Concentration
£5 PPm lig/m Duration and Activity
^ 0.12 235 6.5h/day
'O IE (6 x 50 min)
(2 days of exposure)
VE = 3 1 L/min (asthmatic)
VE = 34 L/min (healthy)
0.12 235 6.6 h
IE (6 x 50 min)
VE = 38.8 L/min
0.16 314 4h
IE (4 x 50 min)
VE = 38.9 L/min
Exposure
Conditions
21 °C
50% RH
18 °C
40% RH
(5 consecutive
days of exposure
to 03, 1 day
exposure to C A) .
18 °C
40% RH (one
exposure to 63,
no control
exposure).
Number and
Gender of Subject
Subjects Characteristics Observed Effect(s)
15
(8M,
7F)
30
(13 M,
17 F)
17 M
15 M
Healthy NS Bronchial reactivity to methacholine increased with
Age 22-41 03 exposure in healthy subjects. FEVj decreased 2%
(pre- to postexposure) in healthy subjects and 7.8% in
Asthmatic NS asthmatics. Responses were generally less on the
Age 18-50 second day. Two healthy and four asthmatics had
FEVj decreases >10%.
Healthy NS FEVj decreased by 12.8, 8.7, 2.5, 0.6, and +0.2 on
Days 1-5 of 03 exposure. Methacholine airway
responsiveness increased by > 100% on all exposure
days. Symptoms increased on the first O3 day, but
were absent on the last three exposure days.
Healthy NS FVC decreased 9.5% and FEVt decreased 16.6% .
FEVj/FVC ratio decreased from 0.79 to 0.73.
Reference
Linn et al. (1993)
Folinsbeeetal. (1993b)
Folinsbee et al. (1993b)
-J a
U See glossary of terms and symbols for abbreviations and acronyms.
Age range in years or as mean + SEM.
-------
 1          In order to extend these initial observations, Horstman et al. (1990) used the same
 2     protocol to expose a group (n = 22) of subjects to clean air and three different
 3     O3 concentrations (0.08, 0.10,  and 0.12 ppm).  Similar responses were shown at 0.12 ppm
 4     as in the previous study, with the exception that the symptom responses were less in the new
 5     group of subjects.  A similar pattern of response in spirometry, airway resistance, and
 6     airway responsiveness, but of smaller magnitude, was seen at the two lower concentrations.
 7     The mean FEVi responses during the four exposures are shown in Figure 7-3.  The
 8     responses  were dependent on concentration and exposure duration (ventilation was not
 9     varied) and averaged 7, 8, and 13% at the three O3 concentrations.  Larsen et al. (1991)
10     have used these data (Horstman et al., 1990) to develop a "dose-response"  relationship for
11     percent change in FEV^ as a function of O3 concentration and exposure duration.  The
12     lognormal multiple linear regression model suggested that FEVj responses  were
13     approximately linear with duration of exposure but that O3 concentration plays a more
14     important role.  The exponent of approximately 4/3 suggests that doubling O3 concentration
15     would be  similar to increasing  exposure duration  by about 2'/2 times.
16          A series  of additional studies were conducted at 0.08 and 0.10 ppm.  The primary aim
17     of these studies was to study changes in cells and inflammatory mediators from BAL (see
18     Section 7.2.4).  McDonnell  et  al.  (1991a) reported an  8.4% decrease in FEVl at 0.08 ppm
19     and an 11.4% decrease at 0.10 ppm. These responses were slightly  larger than seen in the
20     previous Horstman et al.  (1990) study.  The duration—FEVj response data were fit to a
21     three parameter logistic model, which significantly improved the amount of variance
22     explained by the model compared to a linear model. This is consistent with exploratory
23     analyses in the Folinsbee et  al. (1988) report. The reasonably good  fit to the logistic model
24     suggests that the O3-pulmonary function response relationship may have a  sigmoid shape.
25     The main importance of this observation is that it suggests there is a response plateau.  That
26     is, there is some level of response (i.e., plateau)  to a given O3 concentration and exercise
27     ventilation level (i.e., dose rate) beyond which the FEV^ response tends not to increase with
28     increasing duration of exposure.
29           In the fourth study in this series (Folinsbee et al., 1993b),  17 subjects were exposed to
 30     0.12 ppm for 6.6 h on 5 consecutive days. Subjects who were not responsive to O3 were not
 31      selected to participate in this study.  Responses in FEV] on the first of these exposures

        December 1993                           7.74      DRAFT-DO NOT QUOTE OR CITE
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             FEV,   During Air and  Ozone Exposure
          4,500

          4,400

          4,300

      ^ 4,200

      :>" 4,100
      UJ
      LJ.
          4,000

          3,900

          3,800
                               Air
                               0.08 ppm

                               O.IOppm

                               0.12 ppm
                        12345
                            Exposure Duration (h)
     Figure 7-3. The forced expiratory volume in one second (FEV}) is shown hi relation to
                exposure duration (hours) at four different O3 concentrations:
                (1) — <0.01 ppm, (2) • = 0.08 ppm, (3) • = 0.10 ppm, and (4) A =
                0.12 ppm. A 35-min resting exposure period was interposed between the
                end of third and the beginning of the fourth hour. There were six 50-min
                exercise periods (minute ventilation « 39 L/min) during the exposure; these
                measurements were made 5 min after the end of each exercise. The total
                exposure duration was 6.6 h. The standard error of the mean (not shown)
                for these FEV} averages ranged from 120 to 160 mL.

     Source: Horstman et al. (1990).
1     averaged -12.8%.  Again, symptom responses were modest with a significant increase in a

2     lower respiratory symptom score on the first exposure day. A significant increase in airway
3     responsiveness to methacholine was also shown. The response to the repeated exposures is
4     discussed in Section 7.2.1.4 on repeated exposures to 03. In addition, 15 subjects were
      December 1993
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 1     exposed to 0.16 ppm for 4 h using the same hourly exposure protocol as described above.
 2     In these subjects, FVC decreased -9.5% and FEVj declined -16.6%.
 3          Folinsbee et al. (1991) took the FEVl response data from all four studies conducted at
 4     the EPA Health Effects Research Laboratory, using the same prolonged exposure protocol,
 5     and examined the distribution of responses among the subjects at the three concentrations.
 6     This response distribution is illustrated graphically in Figure 7-4.  This figure illustrates that
 7     FEVj decrements as large  as 30 to 50% have been observed with prolonged exposure to
 8     O3 concentrations equal to or less than 0.12 ppm.  This response distribution also allows one
 9     to determine the number or percentage of subjects with responses in excess of a certain level.
10     The proportion of subjects with an FEVj decrease in excess of 10% is shown. With air
11     exposure, no one exceeded this response level; however, 46% of the subjects exposed to
12     0.12 ppm using this exposure protocol had greater than a 10%  drop hi FEVt after 6.6 h.
13     This response  distribution also  illustrates the wide range of response to O3 under these
14     exposure conditions and reinforces the observation by others (McDonnell et al., 1983;
15     Horvath et al., 1981) of a  substantial  range of individual response to O3.
16          Kehrl et  al. (1989b) reported that clearance of radiolabeled diethylene triamine
17     pentacetic acid ( "Tc-DTPA)  was not altered by the 0.08-ppm exposure (see McDonnell
18     et al.,  1991a for exposure details).  Subsequent studies  (Kehrl et al., 1991) suggested that a
19     small acceleration of clearance may have occurred with exposure to 0.12 ppm for 6.6 h.
20     Increased clearance of labeled  99mTc-DTPA from the lung is thought to indicate increased
21     permeability of the  respiratory epithelium (Kehrl et al.,  1987).  These preliminary data
22     (Kehrl et al.,  1991) have not been published.  Other summaries of the four studies from the
23     EPA laboratory, however, have been published (McDonnell et al., 1991b; Horstman et al.,
24      1988a,b; Horstman et al., 1991).
25          Horvath et al. (1991) examined the responses of healthy middle aged (ages 30 to 45)
26      men and women to 0.08 ppm O3 for  6.6 h using the EPA prolonged-exposure protocol.
27      When compared with the clean air exposure, FEVj decreased about 5 % with O3 exposure.
28      However, the variability of this response among this small heterogeneous group of subjects
29      was enough to preclude statistical significance of this observation.  No significant changes
 30      were observed with a second exposure on the next day.  On the first day of Oj exposure,
        December 1993                           7-76      DRAFT-DO NOT QUOTE OR CITE
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                     Distribution of Percent Change in FEVj
                      = 87
                     AIR
             i  10 15 20 2S 30 38 «
N-60
 0.08
                              0510152025903543
                                      Percent of Subjects
> 10% Drop
inFB< °%
26%
31%
46%
       Figure 7-4. The distribution of response for 87 subjects exposed to clean air and at least
                  one of 0.08, 0.10, or 0.12 ppm is shown here.  The O3 exposures lasted
                  6.6 h,  during which time the subjects exercised for 50 min of each hour
                  with a 35-min rest period at the end of the third hour.  The abscissa
                  indicates the number of subjects.  The bar labels on the ordinate indicate
                  decreases in FEVj, expressed as percent change from baseline. For
                  example, the bar labeled 10 indicates the number of subjects with a
                  decrease in FEV, of >5% but ^10%; the bar labeled -5 indicates
                  improvement in FEVj of >0% but <5%.  The rectangle across the bottom
                  of the  graph indicates the percentage of subjects at each O3 concentration
                  with a decrease of FEVj hi excess of 10%.

       Source: Folinsbee et al. (1991).
 1     7 of the 11 subjects reported chest tightness.  The authors point out that the range of
 2     variability in response in their study was similar to that reported by Folinsbee et al.  (1988)

 3     and Horstman et al. (1990), although fewer of their subjects experienced large negative

 4     changes in FEV!.  One possible explanation for the differences between the findings of

 5     Horvath et al. (1991) and Horstman et al. (1990) may be that the subjects in Horvath's study

 6     were significantly older, which may result in reduced responses to O3, as Drechsler-Parks

 7     et al. (1987a) have shown (see Section 7.2.1.3). The ventilation during exercise (37 to

 8     39 L/min) was similar to that reported by Horstman et al. (1990).  An additional FEVi

 9     measurement was made in this study at the end of the lunch period (i.e., after 40 min of

10     rest).  At this time, the small decrements in FEVj seen after the third exercise were reversed
       December 1993
                                              7-77
                 DRAFT-DO NOT QUOTE OR CITE
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 1     and the FEV^ was similar to the response in FA at the same time point.  Although
 2     spirometry was not measured at this time in the other prolonged-exposure studies (Folinsbee
 3     et al.,  1988; Horstman et al., 1990), it was noted that the decline in FEV^ was attenuated
 4     between the third and fourth postexercise measurement. These observations suggest that
 5     subject's lung function may indeed have  improved during the lunch rest period.
 6          Linn et al. (1991, 1992) have reported responses of healthy and asthmatic subjects to
 7     0.12 ppm O3 using the EPA prolonged-exposure protocol.  [These results are presently
 8     available as a pair of abstracts and a preliminary project report.] They observed a small
 9     (2.0%) decrease in FEV^ which was statistically significant, and an increase in airway
10     responsiveness to methacholine.  The functional responses in asthmatics were greater than
11     those of the healthy subjects, a decrease  in FEV^ of 7.9%.  They observed smaller responses
12     on a second consecutive day of exposure, as did Horvath et al. (1991) and Folinsbee et al.
13     (1993b).  The ventilation averaged about 34 and 31 L/min in the healthy and asthmatic
14     subjects, respectively.  The FEVj responses observed in this study, although statistically
15     significant, are much lower than those observed by EPA investigators (Folinsbee et al.,
16     1988;  Horstman et al., 1990; Folinsbee et al., 1993b).  The smaller responses may be due to
17     previous ambient exposures, lower ventilations, or a larger proportion of O3-insensitive
18     subjects in Los Angeles.  Only 1 of 15 healthy subjects experienced an FEVj decrement in
19     excess of 10%, whereas 9 of 30 asthmatics had an FEVj decrement in excess of 10%.
20     Asthmatic responses ranged from +12% to —35%.
21          To further explore the factors that  determine responses to O3, Hazucha et al.  (1992)
22     designed a protocol to examine the effect of a varying, rather than a constant,
23     O3 concentration. In this study, subjects were exposed to both a constant level of 0.12 ppm
24     O3 for 8 h and to an O3 level that ramped linearly from 0 to 0.24 ppm for the first 4 h and
25     then ramped linearly from 0.24 to 0 over the second  4 of the 8 h exposure (triangular
26     concentration profile). Subjects performed moderate exercise for the first 30 min of each
27      hour.  The overall exposure dose for these two exposures, calculated as the O3 concentration
28       x  exposure duration X ventilation, was almost identical (difference  <1%). With exposure
29      to  the constant 0.12 ppm O3, the FEVj  declined approximately 5% by the fifth hour of
 30     exposure and remained at that level for  the remainder of the exposure.  These responses are
 31      illustrated in Figure 7-5.  This observation clearly indicates a response plateau, suggested  in

        December 1993                           7-78      DRAFT-DO NOT QUOTE OR CITE
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       8T
        cc
               Steady vs. Variable Ozone Concentration
                                  FEV, Responses
4.5


4.4


4.3


4.2


4.1
            3.9
                            246
                            Exposure Duration (h)
                                            8
     Figure 7-5. The forced expiratory volume in one second (FEVj) is shown in relation to
                exposure duration (hours) under three conditions: (1) • clean air exposure,
                (2) • a constant concentration of 0.12 ppm O3, and (3) A a variable
                O3 concentration increasing from 0.0 to 0.24 ppm over the first 4 h and
                then decreasing from 0.24 to 0.0 ppm over the second 4 h. Duration of
                exposure (hours) is shown on the abscissa.  Mean FEVj (liters) is shown on
                the ordinate. Subjects exercised (minute ventilation * 40 L/min) for
                30 min during each hour; FEVX was measured at the end of the intervening
                rest period. Total exposure duration was 8 h. Standard error of the mean
                for these FEVt averages (not shown)  ranged from 120 to 150 mL.

      Source: Hazucha et al. (1992).
1     other studies (Horstman et al., 1990), with an exposure regimen that produces relatively

2     small changes in lung function.
3         With the triangular O3 concentration profile, the ¥EVV decreased almost twice as much

4     after 6 h of exposure. The initial response over the first 3 h was minimal and then there was

5     a substantial decrease in FEVj, corresponding to the higher average O3 concentration, that
      December 1993
                                7.79      DRAFT-DO NOT QUOTE OR CITE
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 1     reached a nadir after 6 h.  Despite continued exposure to a lower O3 concentration
 2     (<0.12 ppm), the FEVj began to improve and was reduced by only 5.9%  at the end of the
 3     8-h exposure.  (However, note that the average O3 concentration in the eighth hour was
 4     0.03 ppm).  This study illustrates two important points for those planning to model responses
 5     to O3 exposure.  First, the model should include a response plateau. It is intuitively obvious
 6     that there must be a limit to the acute decrease that can occur in FEV^  However, from this
 7     study, it is clear that the response plateau must be dependent upon the O3 concentration
 8     because much larger decreases in FEVl occur with exposure to higher concentrations than
 9     0.12 ppm.  Secondly, the response to O3 exposure is dependent on the dose rate (some
10     function of O3 concentration and ventilation) as well as the cumulative dose (some function
11     of dose rate and exposure duration), at least when O3  concentration is varied.  This study
12     also affirms the observation (Folinsbee et al.,  1978; Adams et al., 1981; Hazucha, 1987;
13     Larsen et al.,  1991) that O3 concentration is a more important factor in determining
14     O3 responses than either exposure duration or the volume of air breathed during the
15     exposure.
16
17     7.2.3   Increased Airway Responsiveness
18          Increases in airway responsiveness are an important consequence of exposure to 0^.
19     Increased airway responsiveness indicates that the airways are predisposed to
20     bronchoconstriction induced by a variety of stimuli (e.g., specific allergens, sulfur dioxide,
21     cold air, etc.). Airway responses are usually  measured  by having the individual forcefully
22     exhale into a spirometer designed to measure expiratory flow rates (e.g., FEVj) or, less
23     commonly, by measuring airway resistance in a body plethysmograph.  In order to determine
24     the level of airway  responsiveness, airway function is measured before and immediately after
25     the inhalation of small amounts of an aerosolized bronchoconstrictor drug  (e.g., methacholine
26     or histamine).  The dose of the bronchoconstrictor drug is increased in a step-wise fashion
27     until a predetermined degree of airway response (e.g., a 20% drop in FEVj or a 100%
28      increase in airway resistance) has occurred.  The dose of the bronchoconstrictor drug that
29      produced the aforementioned response is often referred  to as the PD20 (i.e., the provocative
30      dose that produced a 20% drop in FEVj) or the PD10o  (i.e., the provocative dose that
31      produced a 100% increase in airway resistance).

        December 1993                           7-80      DRAFT-DO NOT QUOTE OR CITE
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 1           A high level of bronchial responsiveness is a hallmark of asthma.  However, varying
 2      degrees of increased airway responsiveness may occur in other lung disease (e.g., chronic
 3      bronchitis or viral respiratory infections) or in healthy asymptomatic individuals.  The range
 4      of nonspecific bronchial responsiveness, as expressed by the PD20 for example, is at least
 5      1,000-fold from the most sensitive asthmatics to the least sensitive healthy subjects (see
 6      Figure 7-6).  The average PD20 for healthy subjects is 10 to 100 times that of mild to
 7      moderate asthmatics (Chatham et al., 1982; Cockcroft et al., 1977). Atopic or allergic
 8      individuals without asthma typically have a lower PD2o than healthy individuals (Townley
 9      et al.,  1975; Cockcroft et al., 1977), being intermediate in responsiveness between healthy
10     subjects and mild asthmatics. Increasing severity of asthma, as indicated by increased
11      symptoms or medication usage, is associated with a decreased PD20.  Mild asthmatics may
12      have a PD20 that is 10 times higher than that of moderate/severe asthmatics (Cockcroft et al.,
13      1977). A low PD20 in nonasthmatics is also associated with increased symptoms and a
14     reduced baseline FEV1 (Kennedy et al., 1990).  The  average changes in airway
15      responsiveness induced by O3 range from 150 to 500%. This means that a healthy subject
16     with a PD20 of 20 units would decrease to a PD20 between 13 and 4 units.  Therefore, with a
17     pronounced O3-induced change in airway responsiveness, a healthy subject could move from
18     the normal range into the upper half of the mild asthmatic range of airway responsiveness.
19          Results of studies reporting changes in airway responsiveness following 03  exposure
20     are summarized in Table 7-10.  Increased airway responsiveness associated with  O3 exposure
21      was first reported by Golden et al. (1978), who studied histamine-responsiveness in eight
22     healthy men after exposure to 0.6 ppm O3 for 2 h at rest and found that the histamine-
23     induced ARaw for the group was 300% greater 5 min after O3 exposure than at baseline.
24     Two of their subjects, however, had an increased response to histamine 1 week or greater
25     after exposure, raising the possibility that high O3 levels can result in more persistent
26     increases  in airway responsiveness.   Later, Holtzman et al. (1979) found in 16 nonasthmatic
27     subjects that a 10-breath methacholine or histamine challenge increased SR^ almost twice as
28     much after O3 as after air exposure, but this effect resolved after 24 h. Atopic subjects
29     showed similar increases in responsiveness to histamine after O3  exposure.  The authors
30     concluded that the increased nonspecific bronchial responsiveness after O3 exposure was not
        December 1993                           7-81      DRAFT-DO NOT QUOTE OR CITE
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                           Arbitrary Logarithmic Scale
       0.01             0.10             1.00             10.0            100
        *— severe asthma	+
                           •*— moderate asthma—>
                                        *	mild asthma	»
                                                      allergy only—+
                                                          +	healthy
      Figure 7-6.  Airway function can be measured before and immediately after the
                  inhalation of an aerosolized bronchoconstrictor drug like methacholine.
                  The provocative dose that produces a 20% drop in FEVj (PD^) has been
                  used to express the range of nonspecific bronchial responsiveness.
 1      related to atopy. Konig et al. (1980) found increased responsiveness to inhaled acetylcholine
 2      after a 1-h exposure to  627 and 1,960 jwg/m3 (0.32 and 1.0 ppm, respectively).  Folinsbee
 3      and Hazucha (1989) found increased airway responsiveness in 18 females 1 and 18 h after a
 4      70-min exposure to 0.35 ppm O3 when compared to air.  Taken together, these studies
 5      suggest that O3-induced increases in airway responsiveness usually resolve 18 to 24 h after
 6      exposure, but may persist in some  individuals for longer periods.
 7          Dimeo et al.  (1981) were the first to investigate "adaptation" to the increases in airway
 8      responsiveness following O3 exposure. Over 3 days of a 2 h/day exposure to 0.40 ppm O3,
 9      they found progressive  attenuation  of the increases in airway responsiveness such that after
10      the third day of O3 exposure histamine airway responsiveness was no longer different from
11      the sham exposure levels.  Kulle et al. (1982) extended these findings by exposing two
12      groups of healthy volunteers (n =  48) to 0.40 ppm O3 for 3 h/day for 5 days in a row in an
13      experiment  designed to study the process of adaptation to O3 and found that there was a
14     significantly enhanced response to  methacholine after the first 3 days of exposure, but this
15      response  slowly normalized by the end of the fifth day.  Thus, the attenuation of C^-induced
16     increases in airway responsiveness followed the same tune course as attenuation of other
17     pulmonary function changes.
       December 1993                          7-82      DRAFT-DO NOT QUOTE OR CITE
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TABLE 7-10. INCREASED AIRWAY RESPONSIVENESS FOLLOWING
                 OZONE EXPOSURES3
^f Ozone
*"* • Concentration
"— '
vo












i
oo



o

K
i
5
2
1
O
g
§
M
g
n

ppm
0.00
0.08
0.10
0.12

0.00
0.10
0.32
1.00
0.00
0.12
0.20

0.00
0.12




0. 12 ppm O
0. 12 ppm O
air- 100 ppb

air-antigen
/j

0
157
196
235

0
196
627
1,960
0
235
392

0
235




3-100ppbSO2
3-0.12ppm O3
SO2


Number and
Exposure Exposure Gender of
Duration and Activity Conditions Subjects
6.6 h 18 °C 22 M
IE at =39 L/min 40% RH



2 h NA 14



1 h at VE = 89 L/min 31 °C 15 M
followed by 3-4 min at 35% RH 2 F
» 150 L/min

6.6 h with IE at NA 10 M
«25 L/min/m2 BSA




45 min in first 75% RH 8 M
atmosphere and 15 min in 22 °C 5 F
second.
IE
1 h at rest NA 4 M, 3 F
0. 12 ppm O3-antigen

0.00
0.35


0
686


70 min with IE at NA 18 F
40 L/min

auDjeci
Characteristics
Healthy NS,
18-32 years
old


NS



Elite
cyclists


NS
18-33 years
old



Asthmatic
12-18 years
old

Asthmatic
21-64 years
old
19-28 years
old


Observed Effect(s)
33, 47, and 55% decreases in cumulative
dose of methacholine required to produce
a 100% increase in SRaw after exposure to
O3 at 0.08, 0.10, and 0.12 ppm,
respectively.
Increased airway responsiveness to
methacholine immediately after exposure
at the two highest concentrations of O3.

Greater than 20% increase in histamine
responsiveness in one subject at 0. 12 ppm
O3 and in 9 subjects at 0.20 ppm C^.

Approximate doubling of mean
methacholine responsiveness after
exposure. On an individual basis, no
relationship between O3-induced changes
in airway responsiveness and FEVj or
FVC.
Greater declines in FEVj and Vmax50%
and greater increase hi respiratory
resistance after O3-SO2 then after O3-O3
or air-SO2.
Increased bronchoconstrictor response to
inhaled ragweed or grass after O3
exposure compared to air.
PDjoo decreased from 59 CIU after air
exposure to 41 CIU and 45 CIU, 1 and
18 h after 63 exposure, respectively.

Reference
Horstman
et al. (1990)



Konig et al.
(1980)


Gong et al.
(1986)


Folinsbee
et al. (1988)




Koenig et al.
Q990)


Molfino
et al. (1991)

Folinsbee
and Hazucha
H989)
-------
                   TABLE 7-10 (cont'd). INCREASED AIRWAY RESPONSIVENESS FOLLOWING
                                             OZONE EXPOSURES3
Ozone
Concentration

ppm
0.20
0.40
0.40

0.00
0.40
0.00
0.40




0.00
0.60


0.00
0.60





fig/m Duration and Activity
392
784
784

0
784
0
784




0
1,176


0
1,176





2 h with IE at 2 X resting
2 h with IE at 2 X resting
2 h/day for 3 days

3 h/day for 5 days in a row

2 h with IE at
VE = 53-55 L/min




2 h at rest



2 h with IE at 2 X resting






Number and
Exposure Gender of Subject
Conditions Subjects
22 °C 12 M
55% RH 7 F


13 M
11F
22 °C 8 M
50% RH 10 F




NA 5M
3 F


22 °C 11 M
55% RH 5 F






Characteristics Observed Effect(s)
NS
21-32 years
old

NS

9 asthmatics
(5 F, 4 M)
9 healthy
(5 F, 4 M)
18-34 years
old
NS
22-30 years
old

9 atopic
7 nontopic
NS




110% increase in ASR^ to a 10-breath histamine
(1.6%) aerosol challenge after exposure to O3 at
0.40 ppm, but no change at 0.20 ppm. Progressive
adaptation of this effect over 3-day exposure.
Enhanced response to methacholine after first 3 days,
but this response normalized by Day 5.
Decreased PC100SR from 33 mg/mL to 8.5 mg/mL in
healthy subjects afterwO3. PC100Sn fell from
0.52 mg/mL to 0.19 mg/mL in asffimatic subjects after
exposure to O3 and from 0.48 mg/mL to 0.27 mg/mL
after exposure to air.

300% increase hi histamine-induced ARaw 5 min after
O3 exposure; 84% increase and 50% increase 24 h and
1 week after exposure (p > 0.05). Two subjects had an
increased response to histamine 1 week after exposure.
Ten-breath methacholine or histamine challenge
increased SRaw > 150% in 16 nonasthmatics after O3.
On average, the atopic subjects had greater responses
than the nontopic subjects. The increased
responsiveness resolved after 24 h. Atropine
premedication blocked the O3-induced increase hi airway
responsiveness .

Reference
Dimeo
et al.
(1981)

Kulle et al.
(1982)
Kreit et al.
(1989)




Golden
et al.
(1978)

Holtzman
et al.
(1979)




See glossary of terms and symbols for abbreviations and acronyms.
Listed from lowest to highest O3 concentration.
Age range hi years or as mean ± SEM.
-------
 1           Gong et al. (1986) demonstrated increased airway responsiveness to histamine at
 2      0.20 ppm O3 in 17 vigorously exercising elite cyclists who were exposed for 1 h.  Folinsbee
 3      et al. (1988) found an approximate doubling of the mean methacholine responsiveness in a
 4      group of healthy volunteers exposed for 6.6 h to 0.12 ppm O3.  However,  on an individual
 5      basis, they found no relationship between O3-induced changes in airway responsiveness  and
 6      those in FVC and FEV^ suggesting that these two processes occurred by different
 7      mechanisms.  Horstman et al. (1990) corroborated and extended Folinsbee's observations by
 8      demonstrating significant decreases in the PD100 in 22 healthy subjects  immediately after a
 9      6.6-h exposure to concentrations of 03 as low as 0.08 ppm.  Because methacholine
10     challenges were not conducted at later time points  in any of these studies, the duration of the
11      increased airway responsiveness after ambient-level O3 exposure could  not be determined.
12          Kreit et al. (1989) were first to investigate the change in airway responsiveness that
13     occurs after O3 exposure in individuals with asthma.  They exposed nine mild asthmatics
14     (baseline PC100SRaW < 1.5 mg/mL) for 2 h to 0.40 ppm  O3 with IE and found that the
15     baseline  PC^SR^ declined from 0.52 to 0.19 mg/mL after O3 as compared to 0.48 to
16     0.27 mg/mL after air.
17           No doubt exists that O3, even at ambient concentrations, produces acute increases in
18     airway responsiveness.  Whether O3 exposure causes protacted  increases in airway
19     responsiveness in healthy individuals or even induces or predisposes  to asthma is a more
20     difficult  question to answer (see Section 7.4.2).  However, the  increases in airway
21     responsiveness following O3 exposure, even if short in duration, may have important clinical
22     implications.  Several studies have been conducted specifically  to determine the significance
23     of acute  increases in airway responsiveness after O3 exposure.  These studies, designed to
24     test the hypothesis that an O3 exposure heightens the response to  a subsequent
25     bronchoconstrictor challenge, have exposed asthmatics to  O3 or air and, then, to a known
26     bronchoconstrictor agent to compare the pulmonary function changes after O3 to those after
27     air. Koenig et al. (1990) demonstrated that a 45-min exposure to 0.12 ppm O3 followed by a
28      15-min exposure to 120 ppb sulfur dioxide (SO2) caused greater changes in FEV^
29     respiratory resistance, and  Vmax50% in 14  adolescent asthmatics than an air-SO2 exposure
30     combination.
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 1          Molfmo et al. (1991) examined the effects of a 1-h resting exposure to 0.12 ppm O3 on
 2     the response to a ragweed or grass allergen inhalation challenge.  Asthmatic subjects were
 3     exposed twice to air and twice to ozone, once per week over a period of 4 weeks.  Two
 4     allergen challenges were performed, once after air and once after ozone exposure.  The other
 5     air and ozone exposures were followed by a placebo challenge.  A ragweed allergen extract
 6     was used for six of the  seven subjects. The order of experiments  was not randomized; six of
 7     the seven subjects were exposed to the ozone/allergen condition last and five of the seven
 8     were exposed to the air/placebo condition first.  Allergen responsiveness was expressed as
 9     the allergen concentration needed to cause a 15%  reduction in FEVj  or PC15.  The PC15 was
10     lower after the O3 exposure than after the air exposure (p = 0.04).  These observations
11     suggest that allergen-specific airway responsiveness is increased after O3 exposure.  Although
12     it is expected that specific bronchial reactivity will be increased by O3 exposure based on the
13     marked increases in nonspecific  bronchial responsiveness induced  by O3 exposure, such a
14     response would not have been anticipated under these mild exposure  conditions where lung
15     function or symptomatic responses have not been  observed.  The experimental design flaw in
16     this study makes it difficult to assess the validity of conclusions based on the statistical
17     analysis.  These results are provocative but should be considered preliminary until this
18     experiment can be repeated.
19           Contrary to these  studies, Bascom et al. (1990) found no increase in responses to
20     antigen nasal challenge in a group of allergic subjects exposed to 0.5 ppm O3 for 4 h when
21     compared to the air-antigen challenge (see also Section 7.2.4.6).  Because more than half of
22     the population in the United States lives in regions that experience ambient C^ concentrations
23      at the levels used in these studies, O3 may be a clinically important cofactor in the response
24     to airborne bronchoconstrictor substances in individuals with asthma. It is plausible that this
25      phenomenon could contribute to increased asthma exacerbations and, even, increased hospital
26      admissions for asthma exacerbations (see Section 7.4.1). Whether the increased airway
27      responsiveness  following O3 exposure produces an accentuated bronchoconstrictor response
28      to inhaled allergens or  SO2 in healthy individuals or those with other lung diseases is
29      unknown.
 30           Several studies have been undertaken to determine the mechanism of O3-induced
 31      increases in airway responsiveness.  Early experiments in dogs (Lee et al., 1977) and

        December  1993                          7-86      DRAFT-DO NOT QUOTE OR CITE
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 1     humans (Golden et al., 1978) suggested an important role for vagal reflexes because vagal
 2     nerve cooling and atropine inhibited the increase in histamine-induced bronchoconstriction
 3     caused by O3.  Subsequent studies, however, revealed that O3 exposure increased
 4     bronchomotor responses to cholinergic stimuli (e.g., acetylcholine and methacholine) in dogs
 5     (Holtzman et al., 1983) and humans (Seltzer et al., 1986) and that bilateral vagotomy did not
 6     inhibit O3-induced hyperresponsiveness to subcutaneous histamine in guinea pigs (Gordon
 7     et al., 1984).  These data provide strong evidence that O3-induced increased airway
 8     responsiveness  is mediated, at least in part, by cholinergic receptors on airway smooth
 9     muscle cells. Interestingly, Gordon et al. also noted that isometric tension in guinea pig
10     tracheal smooth muscle and lung parenchymal strips in response to histamine and carbachol
11     was not affected by exposure to Oj, suggesting that O3 affected the hi vivo milieu
12     surrounding the smooth muscle rather than producing direct effects on the smooth muscle
13     itself.
14           Osebold et al. (1980), having shown that ovalbumin-sensitized mice were much more
15     sensitive to inhaled albumin after O3 exposure (compared to air), hypothesized that the
16     increased epithelial permeability caused by O3 may allow greater penetration of
17     bronchoconstrictor substances, including methacholine and histamine, and that this would
18     lead to increased airway  responsiveness.  However, Roum and Murlas (1984) suggested that
19     the increased epithelial permeability after O3 could not totally explain this phenomenon
20     because parenteral cholinergic challenge after O3 more reproducibly caused bronchospasm
21     than did inhalation challenge  with methacholine. The increased responsiveness to parenteral
22     compared to inhaled cholinergic challenge may, however, have been due to increased
23     bronchial blood flow after O3 exposure.  Therefore, their findings do not exclude increased
24     epithelial permeability as the cause of increased airway responsiveness after 63 exposure.
25           Holtzman et al. (1983)  first pointed out that O3-induced acute inflammation may  be
26     important in the induction of the increased airway responsiveness.  In mongrel dogs exposed
27     to O3, they found bronchial wall neutrophil infiltration in those animals that developed
28     increased airway responsiveness to acetylcholine, but not in animals that failed to develop
29     increased airway responsiveness.  O'Byrne et al. (1984) later demonstrated that hydroxyurea
30     simultaneously decreased peripheral blood leukocyte counts, decreased neutrophil influx into
31     bronchial tissue, and increased airway responsiveness in dogs exposed to O3.  Both

       December 1993                           7.37      DRAFT-DO NOT QUOTE OR CITE
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 1     O3-induced increased airway responsiveness and bronchial tissue neutrophil influx returned
 2     6 weeks after treatment was discontinued when peripheral leukocyte counts had normalized.
 3     Seltzer et al. (1986) found a larger percentage of neutrophils (30.8 versus 8.0%) in BAL
 4     fluid after O3 exposure in their subjects that had a greater than threefold decrease in the
 5     provocative concentration that caused a > 8 L x cm H2O/L/s increase in SRaw for
 6     methacholine compared to those that had less than a twofold decrease.  These data suggest a
 7     possible association between inflammation and increased airway responsiveness after
 8     O3 exposure. However, McDonnell et al. (1990a,b) found no significant relationship
 9     between changes in airway responsiveness or FEVj  and inflammatory response as judged by
10     changes in BAL neutrophil levels. The concentration of PGE2 was associated with changes
11     in both FEVj and changes in airway responsiveness. Folinsbee et al. (1993) found no
12     association between either symptoms or FEVt responses and changes in airway
13     responsiveness in subjects exposed to 0.12 ppm O3 for 6.6 h.  Furthermore, FEVi responses
14     were attenuated after three exposures, whereas changes in airway responsiveness persisted
15     for a longer period (see also Section 7.2.1.4).  Schelegle et al. (1991)  examined the time
16     course of O3-induced airway neutrophilia and found no correlation between FEVj changes
17     and individual increases in neutrophil levels.   If anything, the least sensitive FEVX
18     responders tended to have fewer neutrophils in their airways.
19          However, Evans et al. (1988) found that the increased airway responsiveness in rats
20     caused by  O3 was not associated with an increase in tracheal mucosal neutrophils or vascular
21     permeability.  In the first of several studies with a prostaglandin inhibitor, indomethacin did
22     not attenuate the increase in airway responsiveness in humans exposed to 0.4 ppm O3 for
23     2 h (Ying et al.,  1990), but did ameliorate the effect of O3 on spirometric endpoints.
24     Kleeberger and Hudak (1992) observed a marked reduction in PMN influx in O3-exposed
25      mice given indomethacin without any change in O3-induced increases in permeability, as
26      indicated by BAL protein. However, Hazucha et al. (1993) found no effect of ibuprofen on
27      PMN levels or protein in the BAL fluid of O3-exposed humans (also see Section 7.2.4.5).
28      Seltzer et al. (1986) and Koren et al.  (1989a,b) found that O3  increases a large number of
 29      BAL inflammatory mediators (including prostaglandin E^ [PGEj], prostaglandin F2 [PGF2a],
 30     and thromboxane B2 [TXB2]), one or more of which may play a role  in the increase in
        December 1993                           7-88      DRAFT-DO NOT QUOTE OR CITE
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 1     airway responsiveness after O3 exposure.  To date, only PGE2 has been shown to be
 2     associated with the increased airway responsiveness (McDonnell et al., 1990a,b).
 3          The role of reactive oxygen metabolites or neuropeptide mediators in the increase in
 4     airway responsiveness after O3 has not been investigated.  Furthermore, there has been no
 5     direct assessment of alterations in nerve afferents, changes in neurotransmitter
 6     concentrations, changes in smooth muscle postsynaptic receptors,  or modulation of nerve
 7     signal transmission by inflammatory mediators as these pertain to the increase in airway
 8     responsiveness after O3.  In conclusion, although the mechanism of O3-induced increases in
 9     airway responsiveness is not completely understood, it appears to be a consequence of
10     cellular or biochemical changes in the airway (see Section 7.2.4 and Tables 7-11 and 7-12).
11     Because these alterations are part of a complex process, it comes  as no surprise that the
12     mechanistic studies on O3-induced increases in airway responsiveness  have not pinpointed an
13     isolated derangement.
14
15     7.2.4  Inflammation and Host Defense
16     7.2.4.1   Introduction
17          In general, inflammation can be considered as the host response  to injury, and the
18     induction of inflammation can be accepted as evidence that injury has  occurred.
19     Inflammation induced by exposure of humans to O3 can have several outcomes:
20     (1) inflammation induced by a single exposure (or several exposures over the course of a
21     summer) can resolve entirely; (2)  continued acute inflammation can evolve into a chronic
22     inflammatory state; (3) continued inflammation can alter the structure  or function of other
23     pulmonary tissue, leading to diseases such as fibrosis or emphysema;  (4) inflammation can
24     alter the body's host defense response to inhaled microorganisms, particularly in potentially
25     vulnerable populations such as the very young and old; and (5) inflammation can alter the
26     lung's response to other agents such as allergens or toxins. It is also possible that the profile
27     of response can be altered in persons with preexisting pulmonary disease (e.g., asthma or
28     COPD) or smokers.
29          The previous O3 criteria document (U.S. Environmental Protection Agency, 1986)
30     contained no studies in which inflammation was measured in humans exposed to 03.  Since
31     then, the use of fiberoptic bronchoscopy has made possible the  sampling of cells and  fluids

       December 1993                           7.39       DRAFT-DO NOT QUOTE OR CITE
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TABLE 7-11. INFLAMMATORY EFFECTS OF CONTROLLED HUMAN
 EXPOSURE TO OZONE (BRONCHOALVEOLAR LAVAGE STUDIES)3
Sr

<,
O
O
I
3
8
R
Ozone Concentration
ppm
0.00
0.08
0.10
0.00
0.20
0.00
0.30
0.00
0.40
0.00
0.40
0.00
0.40
0.00
0.40
3
0
157
196
0
392
0
588
0
784
0
784
0
784
0
784

- Exposure Activity Level
Duration ( Vg)
6.6 h IE (40 L/min)
six 50-min
exercise periods
+ 10 min rest;
35 min lunch
4 h IE (50 min at
40 L/min, 10 min
rest)
1 h (mouth- CE (60 L/min)
piece)
2 h IE (70 L/min) at
15-min intervals
2 h IE (70 L/min) at
15-min intervals
2 h IE (70 L/min) at
15-min intervals
2 h IE (70 L/min) at
15-min intervals
Number and
Gender of
Subjects0
18 M
(18-35 years)
15 M
13 F
(21-39 years)
5M
11 M
11 M
11 M
10 M
(18-35 years)
Observed Effect(s)
BAL fluid 18 h after exposure to 0.1 ppm O3 had
significant increases in PMNs, protein, PG^, fibronectin,
IL-6, lactate dehydrogenase, and a-1 antitrypsin compared
with the same subjects exposed to filtered air. Similar but
smaller increases in all mediators after exposure to
0.08 ppm 03 except for protein and fibronectin.
Decreased phagocytosis of yeast by alveolar macrophages
was noted at both concentrations.
Bronchial lavage, bronchial biopsies, and BAL done 18 h
after exposure. BAL shows changes similar to other
studies. Airway lavage shows increased cells, LDH,
IL-8. Biopsies show increased number of PMNs.
Significantly elevated PMNs in the BAL fluid 1,6, and
24 h after exposure, with peak increases at 6 h.
BAL fluid 18 h after exposure had significant increases hi
PMNs, protein, albumin, IgG, PGE2, plasminogen
activator, elastase, Complement C3a, and fibronectin.
Macrophages removed 1 8 h after exposure had changes in
the rate of synthesis of 123 different proteins as assayed
by computerized densitometry of two-dimensional gel
protein profiles.
BAL fluid 18 h after exposure contained increased levels
of the coagulation factors Tissue Factor and Factor VH.
Macrophages in the BAL fluid had elevated Tissue Factor
mRNA.
BAL fluid 1 h after exposure to 0.4 ppm O3 had
significant increases hi PMNs, protein, PGE2, TXB2,
IL-6, LDH, a-1 antitrypsin, and Tissue Factor compared
with the same subjects exposed to FA. Decreased
phagocytosis of yeast by alveolar macrophages.
Reference
Devlin et al.
(1990, 1991)
Koren et al.
(1991)
Aris et al.
(1993a)
Schelegle et al.
(1991)
Koren et al.
(1989a,b)
Devlin and
Koren (1990)
McGee et al.
(1990)
Koren et al.
(1991)
Devlin et al.
(1993)
-------
                 TABLE 7-11 (cont'd). INFLAMMATORY EFFECTS OF CONTROLLED HUMAN
                             TO OZONE (BRONCHOALVEOLAR LAVAGE STUDIES)8
n> Ozone Concentration
,
VO
u>












-------
TABLE 7-12. INFLAMMATORY AND HOST DEFENSE EFFECTS OF CONTROLLED
3
y
H-*
*?
Ui

-jj
vb
K>
o
6
o
1
-------
            TABLE 7-12 (cont'd). INFLAMMATORY AND HOST DEFENSE EFFECTS OF CONTROLLED
                        HUMAN EXPOSURE TO OZONE (CLEARANCE STUDIES)3
if
VO
O
I
O
1
0
1
o
Ozone Concentration11 Number and
ppm
0.00
0.40
0.00
0.50
0.00
0.25
0.50
0.00
0.25
0.50
1.00
0.00
0.25
0.50
1.00
0.00
0.30
1.00
bxposure Activity Level uenaer or
fig/m Duration ( Vg) Subjects0
0
784
0
784
0
490
980
0
490
980
1,960
0
490
980
1,960
0
588
1,960
See glossary of terms
Listed from lowest to
cAge range in years or
2 h IE (70 L/min) at 8 M
15 min intervals (20-30
years)
2.25 h IE (70 L/min) at 16 M
15 min intervals (20-30
years)
6 h Human nasal
epithelial cells
1 h Airway epithelial
cell line
1 h Airway epithelial
cell line and
alveolar
macrophages
1 h Alveolar
macrophages
and symbols for abbreviations and acronyms.
highest O3 concentration.
as mean ± SEM.
Observed Effect(s)
Subjects inhaled "^Tc-DTPA 75 min after exposure.
Significantly increased clearance of ""Tc-DTPA from the
lung hi O3-exposed subjects. Subjects had expected
changes in FVC and SRaw.
Similar design and results as earlier study (Kehrl et al. ,
1987). For the combined studies the average rate of
clearance was 60% faster in O3-exposed subjects.
Increased in ICAM-1, IL-6, IL-1, and TNF expression at
0.5 ppm. No increase hi IL-8 expression. No increases
at 0.25 ppm.
Dose-dependent increased secretion of PGE2, TXB2,
PGF2Q,, LTB4, and LTD4. More secretion basolaterally
than apically.
Increased secretion of IL-6, IL-8, and fibronectin by
epithelial cells, even at lowest O3 concentration. No
O3-induced secretion of these compounds by macrophages.
Dose-dependent increases hi PGE2 production, and
decreases hi phagocytosis of sheep erythrocytes. No
O3-induced secretion of IL-1, TNF, or IL-6.

Reference
Kehrl et al.
(1987)
Kehrl et al.
(1989a)
Beck et al.
(1993)
McKinnon
et al. (1993)
Devlin et al.
(1993a)
Becker et
al. (1991)

n
-------
 1     lining the respiratory tract of humans for many markers (Reynolds, 1987).  Bronchoalveolar
 2     lavage primarily samples the alveolar region of the lung; however, the use of small volume
 3     lavages (Rennard et al., 1990) or balloon catheters also allows sampling of the airways.
 4     Nasal lavage (NL) allows sampling of cells and fluid removed from the nasal passages.
 5          In the past 6 years, several studies have analyzed BAL and NL cells and fluid from
 6     humans exposed to O3 for markers of inflammation and lung damage (see Tables 7-11 and
 7     7-12).  The presence of neutrophils (polymorphonuclear leukocytes [PMNs]) in the lung has
 8     long been accepted as a hallmark of inflammation and has been taken as the major indicator
 9     that O3 causes inflammation in the lungs of humans.  Soluble mediators of inflammation (or
10     its resolution) such as cytokines and arachidonic acid  metabolites have also been measured in
11     the BAL fluid of humans exposed to O3. Cytokines that have been reported most often are
12     interleukin (IL)-6 and IL-8, although IL-1  and tumor  necrosis factor (TNF) have also been
13     studied.  Soluble metabolites of arachidonic acid involved in inflammation and host defense
14     such (e.g., PGE2 and PGF2a, thromboxane, and leukotrienes [LTs] such as LTB4) have also
15     been reported hi the BAL fluid of humans exposed to O3. In addition to their role in
16     inflammation, many of these compounds have bronchoconstrictive properties, and may be
17     involved in increased airway hyperreactivity observed following O3 exposure.
18          Under normal circumstances, the epithelia lining the large and small airways develop
19     tight junctions and restrict the penetration of exogenous particles and macromolecules from
20     the airway lumen into the interstitium and blood, as well as restricting the flow of plasma
21     components into the airway lumen.  However, several studies (see Table 7-12) report that
22     O3 disrupts the integrity of the epithelial cell barrier in human airways, as measured by
23     increased passage of radiolabeled compounds out of the airways,  as well as passage of
24     markers of plasma influx such as albumin, immunoglobulin,  and total BAL fluid protein into
25     the airways.  In addition, markers of epithelial cell damage such as lactate dehydrogenase
26     (LDH) have also been measured hi the BAL fluid of  humans exposed to O3.
27          Inflammatory cells of the lung such as macrophages, monocytes, and PMNs also
28     constitute an important component of the pulmonary host defense system.  In their
29     unstimulated  state, they present no danger to surrounding pulmonary cells and tissues, but
30     upon activation, they are capable of generating free radicals and enzymes with microbicidal
        December 1993                          7.94      DRAFT-DO NOT QUOTE OR CITE
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 1     activities, but also with the potential of damaging nearby cells.  Animal studies have assessed
 2     the effect of O3 on host defense system function by measuring the lethality to an inhaled
 3     bacteria (see Chapter 6, Section 6.2.3). Although these experiments are not directly
 4     comparable to human studies, the ability of macrophages removed from O3-exposed
 5     individuals to phagocytose and kill microorganisms in culture has been assessed.
 6          Other soluble factors that have been  studied include those involved with fibrin
 7     deposition and degradation (Tissue Factor, Factor Vn, and plasminogen activator), potential
 8     markers of fibrogenesis (fibronectin, platelet derived growth factor), and components of the
 9     complement cascade (C3a).
10
11     7.2.4.2   Inflammation Assessed by Bronchoalveolar Lavage
12          Seltzer et al. (1986) were the first to demonstrate that exposure of humans to
13     O3 resulted in inflammation in the lung.  In this study, 10 volunteers were exposed to
14     0.4 ppm or 0.6 ppm O3 for 2 h while undergoing exercise, and BAL was performed
15     3 h later.  Bronchoalveolar lavage fluid from subjects exposed to O3 contained 7.8-fold more
16     neutrophils compared with BAL fluid from the same subjects exposed to filtered air.
17     Additionally, BAL fluid from O3-exposed subjects contained increased levels of PGE2,
18     pGF2Q;, and TXB2 compared to fluid from air-exposed subjects.  A report by Koren et al.
19     (1989a,b) also described inflammatory changes in the lungs of humans exposed to Oj.
20     Eleven subjects were exposed to 0.4 ppm  O3 for 2 h while undergoing IE at 70 L/min in a
21     study designed to simulate adults working outdoors or children actively playing.
22     Bronchoalveolar lavage was performed 18 h after O3 exposure.  Subjects exposed to O3 had
23     an eightfold increase in neutrophils in the BAL fluid, confirming the observations of Seltzer
24     et al.  In addition, Koren et al. reported a twofold increase in BAL fluid protein, albumin,
25     and immunoglobulin G (IgG) levels, suggestive of increased epithelial cell permeability as a
26     result of O3 exposure.  There was also a 12-fold increase in IL-6 levels in the BAL fluid.
27     Interleukin-1  and TNF were not present in detectable levels in the BAL fluid of any subject.
28     There  was, however, a twofold increase in the proinflammatory eicosanoid PGE2, as well as
29     a twofold increase in the complement component C3a. This study also  provided evidence for
30     stimulation of fibrogenic processes in the  lung by demonstrating significant increases in two
31     components of the coagulation pathway, Tissue Factor and Factor VH (McGee et al., 1990),

       December 1993                           7.95       DRAFT-DO NOT QUOTE OR CITE
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 1     as well as urokinase plasminogen activator and fibronectin (Koren et al., 1989a).  Taken
 2     together, these two studies demonstrate that exposure of humans to moderate levels of
 3     O3 results in an inflammatory reaction in the lung, as evidenced by substantial increases in
 4     neutrophils and proinflammatory compounds. Furthermore, these studies demonstrate that
 5     both cells and mediators capable of damaging pulmonary tissue are increased after
 6     O3 exposure, as are compounds that play a role in fibrotic and fibrinolytic processes.
 7          Although animal studies have shown that the airway is a major site of O3-induced
 8     inflammation, few human studies have confirmed this finding because BAL primarily
 9     samples cells and fluid in the terminal bronchioles and alveoli. However, isolated lavage of
10     the mainstream bronchus using balloon catheters or the more traditional BAL using  small
11     volumes of saline have the ability to preferentially measure O3-induced changes in the large
12     airways. In one study, isolated airway lavage was performed on 14 subjects 18 h after
13     exposure to 0.2 ppm O3 while undergoing moderate exercise (Aris et al., 1993a). Increases
14     in total  lavagable cells, LDH, and IL-8 were reported.  In contrast with Schelegle et al.
15     (1991),  there was no increase in PMNs found in the bronchial fluid; however, bronchial
16     biopsies showed increased numbers of PMNs in airway tissue.
17          The data suggestive of O3-induced changes in epithelial cell permeability described by
18     Koren et al. (1989a), Devlin et al.  (1991), and Koren et al. (1991) support earlier work in
19     which epithelial cell permeability as measured by increased clearance of  "'Tc-DTPA from
20     the lung of humans exposed to O3 was demonstrated (Kehrl et al., 1987).  In that study,
21     eight healthy subjects who inhaled   '"Tc-DTPA just prior to exposure to air or 0.4 ppm
22     O3 for 2 h while undergoing heavy exercise (65 L/min) had  increased clearance of the
23     compound.  Kehrl et al. (1989a) reported similar observations on an additional 16 subjects.
24     For the combined group of 24 subjects exposed for 2 h to 0.4 ppm O3, the average clearance
25     rate was 60% faster than that observed after air exposure, strongly suggesting increased
26     permeability from the airway lumen and alveolar space to the blood and interstitial  spaces.
27     The average O3-induced decrement in FVC in these subjects was —10%.  These changes in
28     permeability are most likely associated with acute inflammation and could  potentially allow
29      better access of inhaled antigens and other substances to the submucosa.
 30           Studies in which human alveolar macrophages and airway epithelial cells were exposed
 31      to O3 in vitro suggest that most of the components found in increased levels in the  BAL fluid

        December 1993                          7-96      DRAFT-DO NOT QUOTE OR CITE
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 1     of O3-exposed humans are produced by epithelial cells.  Macrophages exposed to 0.1, 0.3,
 2     and 1.0 ppm O3 for 1 h showed small increases in PGE2 in cells exposed to the latter two
 3     concentrations, but no change in superoxide anion or cytokine production (Becker et al.,
 4     1991).  In contrast, airway epithelial cells exposed to 0.1, 0.25, 0.5, and 1.0 ppm O3 for
 5     1 h showed large dose-dependent increases in PGE^, TXB2, LTB4, LTC4, and LTD4
 6     (McKinnon et al., 1993).  These cells also showed increases in IL-6, IL-8, and fibronectin at
 7     O3 concentrations as  low as 0.1 ppm (Devlin et al., 1993a). Interestingly, macrophages
 8     removed 18 h later from subjects exposed to 0.4 ppm O3 for 2 h while undergoing
 9     intermittent heavy exercise (Koren et al.,  1989a) showed changes in the rate of synthesis of
10     123 different proteins as measured by quantitative computerized densitometry of two-
11     dimensional gel protein profiles.  However, macrophages exposed to O3  in vitro only showed
12     changes in the rate of synthesis of six proteins, suggesting that most of the changes seen in
13     the in vivo exposed macrophages were due to actions resulting from mediators released by
14     other cells following  O3 exposure, which then altered macrophage function.
15          Numerous controlled human studies  have shown that humans exposed to O3 for
16     5 consecutive days experience decrements in pulmonary function on the first and second
17     days, but the decrements diminish with each succeeding day so that by the fifth day, O3 no
18     longer induces decrements in lung function (see Section 7.2.1).  This phenomenon has been
19     termed "adaptation".  These studies  did not address the question of whether repeated
20     exposure to O3 also induced "adaptation"  to inflammation or lung damage, although two
21     animal studies suggest that although some markers of inflammation may  be diminished,
22     underlying damage to lung epithelial cells continues (Tepper et al., 1989; VanBree et al.,
23     1993).  In a recent study (Devlin et  al., 1993b), humans were exposed to 0.4 ppm O3 for
24     5 consecutive days (2 h/day while undergoing IE) and then were exposed to Oj a single time
25     either  10 or 20 days laters. The results show that numerous indicators of inflammation (e.g.,
26     PMN influx,  IL-6, IL-8, PGI^, BAL protein, fibronectin, macrophage phagocytosis) show
27     "adaptation" (i.e., there is a complete disappearance of response and values are different
28     from those observed  in the same individual after 5 days of exposure to FA).  These
29     attenuated responses  show a gradual recovery.  Ten days later, some of these markers have
30     regained full  susceptibility, but others have not regained  susceptibility even after 20 days.
       December 1993                           7.97      DRAFT-DO NOT QUOTE OR CITE
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 1     In agreement with animal studies, some markers (LDH, elastase) never show adaptation,
 2     indicating that tissue damage may continue to occur during repeated exposure.
 3
 4     7.2.4.3   Inflammation Induced by Ambient Levels of Ozone
 5          Devlin et al. (1991) reported an inflammatory response in humans exposed to levels of
 6     O3 at or below 0.12 ppm.  In this  study,  10 volunteers were exposed to FA, 0.10 ppm O3,
 7     and 0.08 O3 ppm for 6.6 h while undergoing moderate exercise (40 L/min), and underwent
 8     BAL 18 h later.  An additional eight subjects were exposed to filtered air and 0.08 ppm O3.
 9     Increased levels of neutrophils and IL-6 were found in BAL fluid of subjects exposed to
10     0.10 and 0.08 ppm O3.  There were also increases in most of the other compounds reported
11     by Koren et al. (1989a,b), including fibronectin and PGE^.  In this study, alveolar
12     macrophage phagocytic capability  was also monitored, and it was reported that macrophages
13     removed from humans exposed to both O3 concentrations had decreased ability to
14     phagocytose Candida albicans opsonized with complement.  Comparison of the magnitude of
15     inflammatory changes observed in this study and by Koren et al.  (1989a,b), when normalized
16     for differences in concentration, duration of exposure, and ventilation,  suggest that lung
17     inflammation from O3 may occur as a consequence of exposure to ambient levels while
18     exercising.  Although the mean changes in IL-6, PGE2, and neutrophils reported by Devlin
19     et al. (1991) were  low, there was  a considerable range of response among the individuals
20     participating in the study.  Thus, although some of the study population showed little or no
21     response to O3,  others had increases in IL-6 or PMNs that were as large or larger as those
22     reported by Koren et al. (1989a,b) when subjects were exposed to 0.4 ppm O3.
23     Interestingly, those individuals who had the largest increases in inflammatory mediators in
 24     this study did not necessarily have the largest decrements in pulmonary function,  suggesting
 25      separate mechanisms underlying these two responses to O3.  These data suggest that,
 26      although the population as a whole may have a small inflammatory response to low levels of
 27      O3, there may be  a significant subpopulation that is very sensitive to these low levels of O3.
 28      Furthermore,  even a small inflammatory response (if it recurs) in the population as a whole
 29      should not be discounted.
 30
 31

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 1     7.2.4.4   Time Course of Inflammatory Response
 2          The time course of the inflammatory response to O3 in humans has not been fully
 3     explored.  Studies in which BAL was performed 1 h (Devlin et al., 1990;  Koren et al.,
 4     1991) or 3 h (Seltzer et al.,  1986) after exposure to 0.4 ppm O3 demonstrate that the
 5     inflammatory response is quickly initiated, and the data reported by Koren et al. (1989a,b)
 6     indicate that even 18 h after exposure, inflammatory mediators such as IL-6 and PMNs are
 7     still substantially elevated.  However, a comparison of these studies shows there are
 8     differences in the magnitude of response of some indicators depending on  when BAL is
 9     performed after O3 exposure.  Ozone-induced increases in PMNs, IL-6, and PGE^ are
10     greater 1 h after O3 exposure, whereas BAL levels of fibronectin and plasminogen activator
11     are greater 18 h after exposure.  Still other compounds (protein, Tissue Factor) are equally
12     elevated both 1 and 18 h after O3 exposure.  Schelegle et al. (1991) exposed five subjects to
13     filtered air or 0.3 ppm O3 for 1 h with a ventilation of 60 L/min.  Each subject was exposed
14     to O3 on three separate occasions and BAL was performed either 1, 6, or 24 h after
15     exposure.  In addition, the BAL was separated into two fractions: the first 60 mL wash was
16     designated the "proximal airways" fraction (PA), and the remaining three  60 mL washes
17     were pooled and designated the "distal airways and alveolar surface" fraction (DAAS). The
18     percent of PMNs in the PA sample was  statistically elevated at 1, 6, and 24 h after
19     O3 exposure, with a peak response at 6 h. The percent of PMNs in the DAAS sample was
20     elevated at only the 6 and 24 h time points, with equivalent elevations at each time.
21     However, more extensive studies  will be needed to better define the kinetics of appearance
22     and disappearance of the various inflammatory mediators observed in the  lungs of humans
23     exposed to O3.
24
25     7.2.4.5  Effect of Anti-Inflammatory Agents on Ozone-Induced Inflammation
26          Previous studies (Schelegle et al., 1987; Eschenbacher et al., 1989)  have shown that
27     indomethacin, an anti-inflammatory agent that inhibits cyclooxygenase products of
28     arachidonic acid metabolism, is capable of blunting the well-documented decrements in
29     pulmonary function observed in humans exposed to O3. In a recent study, 10 healthy male
30     volunteers were given 800 mg ibuprofen, another anti-inflammatory agent that blocks
31     cyclooxygenase metabolism, or a placebo 90 min prior to a 2-h exposure to 0.4 ppm O3.

       December 1993                          7.99      DRAFT-DO NOT QUOTE OR CITE
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 1     An additional 200 mg was administered following the first hour of exposure.
 2     Bronchoalveolar lavage was performed 1 h after the exposure.  As expected, subjects given
 3     ibuprofen had blunted decrements in lung function following O3 exposure compared to the
 4     same subjects given a placebo (Hazucha et al.,  1993).  Bronchoalveolar lavage fluid from
 5     subjects given ibuprofen also had reduced levels of the cyclooxygenase product PGEz as well
 6     as IL-6, but no decreases were observed in PMNs, fibronectin, permeability, LDH, or
 7     macrophage phagocytic function (Hazucha et al., 1993).  These data suggest that although
 8     anti-inflammatory agents may blunt O3-induced decrements in FEVj and increases in PGI^,
 9     most inflammatory mediators are elevated in the BAL of these subjects.  Additional studies
10     will  be needed to determine the role, if any,  played by PGE2 and IL-6 in the pulmonary
11     function decrements and increases in PMNs, fibronectin, and other components  in the BAL
12     fluid of humans after O3 exposure.
13
14     7.2.4.6  Use of Nasal Lavage to Assess Ozone-Induced Inflammation in the Upper
15              Respiratory Tract
16          Bronchoalveolar lavage has proven to be a powerful research tool to analyze changes in
17     the lung following exposure of humans to xenobiotics.  However, because BAL is expensive,
18     somewhat invasive, and requires specialized personnel and facilities, it is usually done only
19     with small numbers of subjects and in selected medical centers.  Therefore, there is
20     increasing interest in the use of NL as a tool in assessing C^-induced inflammation in the
21     upper respiratory tract, which is the primary portal for inspired air, and therefore the first
22     region of the respiratory tract to come in contact with airborne xenobiotics.  Nasal lavage is
23     simple and rapid to perform, is noninvasive, and allows collection of multiple sequential
24     samples from the same person.  Graham et al. (1988)  reported increased levels of PMNs in
25     the  NL fluid of 21 humans exposed to 0.5 ppm O3 at rest for  4 h on 2 consecutive days,
26     with NL performed immediately before and immediately after each exposure as well as
27     22 h after the second exposure.  Nasal lavage fluid contained elevated PMN levels at all
28     postexposure times tested, with peak values occurring immediately prior to the  second day of
29      exposure.  There were no changes in PMN  levels at any time in 20 subjects exposed to clean
 30      air  for 2 consecutive days. Bascom et al. (1990) exposed 12 subjects with allergic rhinitis to
 31      filtered air or 0.5 ppm O3 at rest for 4 h, followed immediately by NL.  They  reported  a
 32      7-fold increase in PMNs, a 20-fold increase in eosinophils, and a 10-fold increase in
        December 1993                           7-100      DRAFT-DO NOT QUOTE OR CITE
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 1      mononuclear cells following 03 exposure, as well as a 2.5-fold increase in albumin.  Graham
 2      and Koren compared inflammatory mediators present in both the NL and BAL fluids of
 3      humans exposed to O3.  The same 11 subjects  who were exposed to 0.4 ppm O3 for 2 h with
 4      BAL performed 18 h later, as described earlier (Koren et al., 1989a,b), also underwent NL
 5      immediately before, immediately after, and 18 h after each exposure (Graham and Koren,
 6      1990).  There were significant increases in PMNs in the NL fluid taken both immediately
 7      after exposure as well as the next day. Increases in NL and BAL PMNs were similar
 8      (6.6- and 8-fold, respectively), demonstrating a qualitative correlation between changes in the
 9      lower airways as assessed by BAL and the upper respiratory tract as assessed by BAL.
10     Furthermore, all individuals who had increased PMNs in BAL fluid also had increased
11      PMNs in NL fluid, although the NL PMN increase could not quantitatively predict the BAL
12     PMN increase.  Albumin, a marker of epithelial cell permeability, was increased  18 h later,
13     but not immediately after exposure.  There were no changes in PGE^, plasminogen activator,
14     or LTC4/D4/E4 (Graham and Koren,  1990). However, tryptase,  a constituent of mast cells
15      and contained in the same granules as histamine, was found in elevated levels immediately
16     after O3 exposure, but not 18 h later  (Koren et al., 1990). These studies suggest that NL
17     may serve as a  sensitive and reliable tool to detect inflammation in the upper airways of
18     humans exposed to xenobiotics.
19
20     7.2.4.7  Changes in Host Defense Capability Following Ozone Exposure
21           Concern about the effect of O3 on human host defense capability derives from
22     numerous animal studies demonstrating that exposure to as little as 0.08 ppm O3 increases
23     mortality in rodents subsequently challenged with aerosolized or instilled bacteria (Gardner
24     et al., 1982; Van Loveren et al., 1988, see Chapter 6, Section 6.2.3).  Because of the
25     difficulty in repeating these types of studies with humans, there are little data available on the
26     direct effects of O3 on human host defense systems.  A study of experimental rhinovirus
27     infection in susceptible volunteers failed  to show any effect of 5 consecutive days of
28     O3 exposure on the clinical evolution or host response to a viral challenge (Henderson et al.,
29     1988).  In this  study, 24 young males were inoculated with type  39 rhinovirus
30     (1,000 TCID-50) administered as nose drops.  Half were then exposed to 0.3 ppm O3
31     (6 h per day) for 5 consecutive days while undergoing intermittent light exercise, and half

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 1     were exposed to clean air under the same regimen.  There was no difference in rhinovirus
 2     tilers in nasal secretions between the O3-exposed and control groups, nor were there any
 3     differences in levels of interferon gamma or PMNs in NL fluid and blood lymphocyte
 4     proliferative response to rhinovirus antigen. However, recent findings that rhinovirus can
 5     attach to the intracellular adhesion molecule (ICAM) receptor  on respiratory tract epithelial
 6     cells (Greve et al., 1989) and that O3 can up-regulate the ICAM receptor on nasal epithelial
 7     cells (Beck et al., 1994) suggest that more studies are needed to explore more fully the
 8     potential interaction between O3 exposure and viral infectivity.
 9          In a single study, human macrophage host defense  capacity was measured in vitro in
10     macrophages removed from subjects exposed to low levels of O3 (0.08 and 0.10 ppm) for
11     6.6 h while undergoing moderate exercise.  Macrophages from O3-exposed subjects had
12     significant decrements in complement receptor (but not Fc receptor) mediated phagocytosis of
13     Candida albicans compared with macrophages removed  from  air-exposed individuals (Devlin
14     et al.,  1991).  These data show that acute in vivo exposure of humans to 63 results in
15     impairment of alveolar macrophage host defense capability, potentially resulting in decreased
16     ability to phagocytose and kill inhaled microorganisms in vivo.  Human alveolar
17     macrophages have also been exposed to O3 in vitro to investigate whether changes in
18     macrophage host defense functions are due to a direct effect of O3  on macrophages or
19     secondary effects resulting from lung injury and inflammation.  Becker et al. (1991) exposed
20     macrophages to 0.1 to 1.0 ppm O3 for 1 h and showed a dose-dependent decrease in
21     phagocytosis of antibody-coated sheep  erythrocytes; a small increase in PG!^;  and
22     production of significantly lower levels of IL-1, TNF, and IL-6 upon stimulation with
23     lipopolysaccharide when compared with air-exposed cells (Becker et al., 1991).  Although
24     there are several studies in which animals have been exposed to bacteria or virus in
25     conjunction with O3 exposure, which provide little evidence to suggest that O3 impairs the
26      immune system's ability to fight viral infections, there is insufficient human data to know
 27      whether O3 exposure affects viral  infectivity.  However, there is potential cause for concern
 28      that O3 may render humans and animals more susceptible to a subsequent bacterial challenge.
 29           There are two studies that have investigated the effect of O3 exposure on mucociliary
 30      clearance of inhaled particles, with conflicting results.   In one study (Foster et al., 1987),
 31      seven male volunteers inhaled radiolabeled ferric oxide  (99mTc-Fe2O3) particles and were

        December 1993                          7-102      DRAFT-DO NOT QUOTE OR CITE
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 1     then exposed to FA, 0.2 ppm O3, and 0.4 ppm O3 for 2 h while undergoing light IE.  They
 2     observed a concentration-dependent increase in rate of particle clearance 2 h after exposure,
 3     although increased clearance was primarily confined to the peripheral airways in subjects
 4     exposed to 0.2 ppm O3.  In the second study (Gerrity et al., 1993), 15 male or female
 5     subjects were exposed to clean air or 0.4 ppm O3 for 1 h while undergoing CE (40 L/min).
 6     Two hours after exposure, subjects inhaled 99mTc-Fe2O3 particles and clearance was
 7     measured with a gamma camera for the next 3 h and then the next morning.  There was no
 8     difference in the clearance rate of particles in air and O3-exposed subjects.  The discrepancy
 9     between these studies may be explained by differences in exposure protocol, time of particle
10     inhalation, or time of clearance measurement, or the presence of cough immediately
11     following O3 exposure, which may have accelerated clearance in the first study.  Further
12     studies will be needed to better define the relationship between O3 exposure and mucociliary
13     clearance.
14
15     7.2.5   Extrapulmonary Effects of Ozone
16          It is still believed that O3 immediately reacts on contact with respiratory system tissue
17     and is not absorbed or transported to extrapulmonary sites to any significant degree (see
18     Chapter 8).  A  number of laboratory animal studies presented hi Chapter 6, however, suggest
19     that reaction products formed by the interaction of O3 with respiratory system fluids or
20     tissues may produce effects measured outside the respiratory tract—either in the blood, as
21     changes in circulating blood lymphocytes, erythrocytes, and serum, or as changes in the
22     structure or function of other organs, such as the parathyroid gland, the heart, the liver, and
23     the central nervous system (see Section 6.3).  Very little is known, however, about the
24     mechanisms by which O3 could cause these extrapulmonary effects.
25          The results from human exposure studies discussed in the previous criteria document
26     (U.S. Environmental Protection Agency, 1986) were not able to demonstrate any consistent
27     extrapulmonary effects (see Section 10.6 of the 1986  document).  Early studies on peripheral
28     blood lymphocytes collected from human volunteers did not find any significant genotoxic or
29     functional changes at O3 exposures of 0.4 to 0.6 ppm for up to 4 h/day.  Limited data on
30     human subjects available at the tune the 1986 criteria document was published also indicated
31     that 0.5 ppm O3 exposure for over 2 h caused transient changes in blood erythrocytes and

       December 1993                          7-103     DRAFT-DO NOT QUOTE OR CITE
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 1      sera (e.g., erythrocyte fragility and enzyme activities), but the physiological significance of
 2      these studies remains questionable.  The conclusions drawn from these early studies raises
 3      doubt that cellular damage or altered function is occurring to circulating cells  at
 4      O3 exposures under 0.5 ppm.
 5           New studies on the potential extrapulrnonary effects of in vivo O3 exposure of human
 6      subjects published since the publication of the previous criteria document (U.S.
 7      Environmental Protection Agency,  1986) have not been very definitive.  Johnson et al.
 8      (1986) exposed 11 male nonsmokers to 0.5 ppm O3 for 4 h on 2 consecutive  days. When
 9      compared to air controls, O3 exposure did not result in any significant change in the activity
10     of blood plasma a-1-proteinase inhibitor.  Schelegle et al. (1989) exposed 20  O3-sensitive,
11      healthy young men to 0.20 and 0.35 ppm with heavy exercise (VE = 50 L/min).  Plasma
12     concentrations of PGF2a were elevated after 40 and 80 min of exposure to the higher
13     O3 level (0.35 ppm).  It is likely, however, that the elevation of this ecosanoid in the  blood
14     was due to either increased production or decreased metabolism of PGF2a in  the lung.
15          The demonstration in the previous section (Section 7.2.4) of an array of inflammatory
16     mediators and immune modulators  released at the airway surface provides a possible
17     mechanism for effects to occur elsewhere in the body.  Additional in vivo studies are  needed,
18     therefore, in order to determine if there are significant extrapulmonary effects of
19     O3 exposure in humans and at what levels of exposure they might occur.
20
21     7.2.6   Ozone Mixed with  Other Pollutants
22           Although it is well-known that polluted air contains a large number of chemical  species,
23     the most common approach to evaluating air pollution effects under laboratory conditions  has
24     been assessment of responses consequent to exposure to single pollutants. This has been the
25     case for a variety of reasons, not the least of which is the  problem inherent in adequately
26     controlling the concentrations of multiple pollutants simultaneously. Further, atmospheric
 27      chemistry is very complicated, and it  is difficult to adequately assess the exposure mixture as
 28      the number of constituent pollutants increases.  Observed effects may be related to unknown
 29      reaction products, the monitored pollutants being only surrogates.  Other problems inherent
 30      in mixture studies involve considerations such as whether pollutants are presented
 31      simultaneously, or in sequential or overlapping patterns.  Ideally, the selected pattern should

        December 1993                           7-104     DRAFT-DO NOT QUOTE OR CITE
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 1     at least approximate one that occurs in the ambient environment. In spite of these
 2     difficulties, information from mixture studies is important from the standpoint of attempts to
 3     better understand responses of humans to the complex mixture of ambient air.
 4          The previous O3 criteria document (U.S Environmental Protection Agency, 1986)
 5     evaluated the limited data base of information available on mixtures of O3 with one or more
 6     pollutants, concluding that pulmonary function changes were no more than additive, and in
 7     most cases, attributable to O3 alone.  Several new studies have since appeared in which
 8     human subjects were exposed to mixtures of two or more pollutants, or to individual
 9     pollutants sequentially (Table 7-13), extending the database for controlled studies.
10     Epidemiological studies have also investigated mixtures of pollutants and have found
11     evidence suggestive of synergistic effects (see Section 7-4).
12
13     7.2.6.1  Ozone and Sulfur-Containing Pollutants
14          Horvath et al. (1987) compared the pulmonary function responses of male subjects
15     (19 to 29 years of age) with normal baseline pulmonary function to four experimental
16     conditions:  (1) FA,  (2) 0.25 ppm O3, (3) 1,200 to 1,600 /*g/m3 H2SO4 aerosol, and
                                             2
17     (4) 0.25 ppm O3 + 1,200 to 1,600 /xg/m H2SO4 aerosol.  Exposures were completed in
18     random sequence, a minimum of 1 week apart, and were conducted at 35 °C and 83%
19     relative humidity.  Subjects alternated 20-min rest and exercise (VE = 30 to 32 L/min)
20     periods throughout the 2-h exposures.  The results indicated that neither O3 alone nor
21     O3 mixed with H2SO4 aerosol had  significant effects on any pulmonary function, metabolic,
22     or ventilatory parameter.
23           Koenig et al. (1990) evaluated sequential O3 (0.12 ppm)  and  SO2 (0.10 ppm) exposures
24     in 13 allergic, asthmatic adolescents (12 to 18 years of age). Three subjects used no regular
25     medications,  the other 10 used one or more of beta-adrenergic agents, theophylline, and
26     antihistamines.   All subjects had a  PC20 for methacholine of 10 mg/mL or less.  Subjects
27     took their morning medication on experiment days if needed, but at least 4 h elapsed between
28     any medication use and the start of the experiment.  The subjects participated in three
29     exposures (22 °C and 75% relative humidity), which were presented in random order and at
30     least  1 week apart. The three exposures were (1) air - SO2,  (2) O3 -  O3,  and
31     (3) O3 - SO2.  The mouthpiece exposures were 1 h in duration, during which the subjects

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                         TABLE 7-13. OZONE MIXED WITH OTHER POLLUTANTS8
-jj
t—*
o


Concentration
ppm
Exposure Number and
Duration and Exposure Gender of
3 c
/tg/m Pollutant Activity Conditions Subjects
Subject
Characteristics

Observed Effect(s)

Reference
Peroxyacetyl Nitrate
0.485
0.27



0.45
0.30

0.45
0.60
0.13


951 O3
1,337 PAN



882 O3
1,485 PAN

882 O3
1,128 N02
644 PAN


Nitrogen-Containing
0.12
0.30


0.20







235 O3
564 N02


392 O3
500 HNO3
H2O





2 h T = 21 °C 10 F
IE WBGT
VE « 25 L/min


2 h T « 22 °C 3 M
IE RH « 60% 5 F
VE « 27 L/min
2 h T = 24 °C 16 M
IE RH = 55-58% 16 F
VE = 25 L/min


Pollutants
1 h (mouthpiece) T = 22 °C 5 M
IE RH = 75% 7 F
VE = 4-5 times
resting
5 h T = 20 °C 6 M
IE (50 min/h RH = 5% 4 F
exercise)
VE « 40 L/min
2 h HNO3 or
H2O fog or air,
followed by 1-h
break, followed
Healthy NS
19-36 years



Healthy NS
Mean age = 24 years

Healthy NS
16 subjects,
19-26 years;
16 subjects,
51-76 years

Healthy NS
12-17 years


Healthy NS
Minimum of 10%
decrement in FEVj
after 3 h exposure to
0.20 ppm O3 with
50 min exercise/h


Exposure to the mixture of
PAN + O3 induced decrements
in FVC and FEVt averaging 10%
greater than observed following
exposure to O3 alone.
No differences between
responses to exposure to O3
alone and Oj + PAN.
No differences between
responses to O3 alone,
QI + NO2, 03 + PAN, or
03 + NO2 + PAN.


No significant changes hi any
pulmonary function with O3
alone or O3 -1- NO2.

Exposure to HNO3 or H2O fog
followed by O3 induced smaller
pulmonary function decrements
than air followed by O3 .




Horvath et al.
(1986)



Drechsler-Parks
et al. (1987b)

Drechsler-Parks
et al. (1989)




Koenig et al.
(1988)


Aris et al. (1991)







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December 1993
                        7-107
                                   DRAFT-DO NOT QUOTE OR CITE
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TABLE 7-13 (cont'd). OZONE MIXED WITH OTHER POLLUTANTS8
I
1
§
u>
•^J
H"k
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00
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6
o
1
1
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Exposure
Concentrationb Duration and Exposure
ppm £ig/m Pollutant Activity Conditions0
Sulfur-Containing Pollutants (cont'd)
0.12 235 O3 6.5 h T = 21 °C
100 H2SO4 2 consecutive RH = 50%
days
50 min exercise/h
VE = 29 L/min
0.12 235 O3 1.5 h with IE for T = 22 °C
0.30 564 NO2 two consecutive RH = 65%
70 H2SO4 days; VE »
0.050 HNO3 23.2 L/min
0.25 490 O3 2 h T = 35 °C
1,200- H2S04 IE RH = 83%
1,600 aerosol VE = 30-
32 L/min
Number and
Gender of Subject
Subjects Characteristics

Nonasthmatic 22-41 years
8M NS
7F
Asthmatic 1 8-50 years
13 M NS
17 F
Asthmatic 12-19 years
adolescents NS
22 completed
study
15 M
7F
9 M Healthy NS
19-29 years
Observed Effect(s) Reference

Exposure to O3 or O3 -t- H2SO4 Linn et al.
induced significant decrements hi (1993)
forced expiratory function.
Differences between O3 and O3 +
H2SO4 were at best marginally
significant. O3 is the more important
pollutant for inducing respiratory
effects. A few asthmatic and
nonasthmatic subjects were more
responsive to O3 -1- H2SO4 than to
O3 alone.
No significant pulmonary function Koenig et al.
changes following any exposure (1993)
compared to response to clean air.
Six additional subjects started the
study, but dropped out due to
uncomfortable symptoms.
No significant effects of exposure to Horvath et al.
O3 alone or combined with H2SO4 (1987)
aerosol.
aSee glossary of terms and symbols for abbreviations and acronyms.
Grouped by pollutant mixture.
°WBGT = 0.7 Twet bulb + 0.3 Tdiy bulb or globe.
Age range in years or as mean ± SEM.
-------
 1     breathed one test gas for 45 min, followed by a second gas for the final 15 min.  Subjects
 2     exercised at a VE of about 30 L/min during the second and fourth 15-min segments of the
 3     exposure.  Pulmonary functions were measured 2 to 3 and 7 to 8 min postexposure.
 4     Changes in ¥EVl and RT were significantly greater following the O3 - SO2 exposure than
 5     following the other two exposures.  Although the subject group was small, the results
 6     indicate that O3 exposure may potentiate responses to SC^ exposure in asthmatic adolescents.
 7     It should be noted that the SO2 concentration (0.100 ppm) used in this study  is a subthreshold
 8     level.
 9          Linn et al. (1993) evaluated the pulmonary function and symptom responses of
10     15 atopic or normal subjects and 30 asthmatic subjects exposed to FA,  0.12 ppm O3,
11     100 /xg/m  respirable H2SO4 aerosol, and a mixture of the two pollutants. The chamber
12     exposures were 6.5 h in duration, during which the subjects walked on a treadmill (VE about
13     29 L/min) for 50 min of each hour.  There was a 30 min lunch period  following the third
14     hour.  Pulmonary function and symptom responses were  measured preexposure and during
15     the hourly 10 min breaks, and a methacholine bronchochallenge  test was performed following
16     each exposure.  Relative to  responses to the FA exposure, H2SO4 aerosol exposure alone
17     induced no significant alteration in pulmonary function, symptoms, or bronchial reactivity to
18     methacholine.  Exposure to  O3 alone, or mixed with H2SO4 aerosol, induced significant
19     decrements in forced expiratory function and increased bronchial reactivity.  Both effects
20     were greater on the first of  two consecutive days of exposure.  Group mean  lung function
21     and methacholine reactivity  changes were somewhat larger following O3 + H2SO4 aerosol
22     compared to exposure to O3 alone, but the differences were at best marginally significant,
23     and usually nonsignificant, depending on the function tested.  However, there were a  few
24     individual subjects who showed significantly larger pulmonary function decrements following
25     the exposure to O3 + H2SO4 than following exposure to O3 alone.  The authors concluded
26     that O3 is more important than H2SO4 aerosol in inducing pulmonary dysfunction in normal,
27     atopic  and asthmatic adults.  There does, however,  appear to be  a more sensitive
28     subpopulation that does respond to O3 + H2SO4 aerosol  more strongly  than the average
29     adult.
30          Kagawa (1986) exposed Japanese men to three mixtures (1) O3  (0.30 ppm) + NO2
31     (0.30 ppm) + H2S04 (200 /ig/m3), (2) O3 (0.15 ppm) + NO2 (0.15 ppm) + H2SO4

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 1     (200 /ig/m3), or (3) O3 (0.15 ppm) + N02 (0.15 ppm) + SO2 (0.15 ppm)+ H2SO4
 2     (200 j«g/m ).  Exposures were 2 h in duration, and subjects exercised for a total of 20 min
 3     during exposure 1 or 60 min during exposures 2 and 3.  Some of the subjects were smokers.
 4     Reported symptoms were attributed to O3 exposure, whereas small decrements in airway
 5     conductance (^10%) were observed following exposures to mixtures 1 and 2.  Although the
 6     magnitude of the FEVj decrement is not stated, a possible decrease was observed after
 7     exposure 3. The responses observed with these mixed exposure conditions were no different
 8     than responses reported for exposures to similar concentrations of O3, indicating no enhanced
 9     response due to the presence of the other pollutants in the mixtures.
10
11     7.2.6.2   Ozone and Nitrogen-Containing Pollutants
12          Adams et al. (1987) reported on the responses of 20 males and 20 females (18 to
13     30 years of age), all healthy nonsmokers, exposed to (1) FA, (2) 0.30 ppm  O3, (3) 0.60 ppm
14     NO2, and (4)  0.30 ppm  O3 + 0.60 ppm NO2.  Subjects were exposed via mouthpiece for
15     1 h, during which they exercised continuously at VE of about 70 L/min for males and
16     50 L/min for females.  The exposures were presented in random order, a minimum of 5 days
17     apart. There  were no differences in any pulmonary function between the O3 and
18     NO2 +  O3 exposures, except for SR^, which was lower following NO2  + O3 than
19     following O3 alone for both men and women.
20          Koenig et al. (1988) exposed  14 male and 10 female adolescents to FA, 0.30 ppm NO^
21     0.12 ppm O3, and 0.30 ppm NO2 + 0.12 ppm O3. Twelve of the subjects were healthy
22     normals, and  the other 12 were allergic asthmatics. The asthmatics, except for one who took
23     no regular medications,  used one or more of beta-adrenergic agents, theophylline, and
24     antihistamines.  Asthmatic  subjects took their morning medications if needed, but refrained
25     from medication use for at least 4 h prior to the exposures. The mouthpiece exposures were
26     1 h in duration, during which the subjects exercised hi 15-min periods (mean VE = 32.8 ±
27     6.0 L/min), alternated with 15-min rest periods. Neither normal nor asthmatic subjects
28     evidenced changes in any measure  of pulmonary function following O3 or NO2 +
29     O3 exposure.
30           Aris et al. (1991) examined pulmonary function responses  to a 3-h exposure to
31     0.20 ppm O3 following a 2-h exposure  to nitric acid (HNO3) or water (H2O) fog. This is a

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 1     common pattern of pollutant exposure in coastal California areas.  Subjects were 10 healthy
 2     adults 21 to 31 years of age.  They were prescreened for a decrement of 10% or greater in
 3     FEVj following 3 h of exposure to 0.20 ppm 63, during which they exercised for 50 min of
 4     each hour (VE = 40 L/min).  Decrements in FEVt following the screening O3 exposure
 5     ranged from 15 to 49%.  The three exposures (FA +  O3, H2O fog + O3, and HNO3
 6     fog + O3) were presented in random order, and were separated by a minimum of 2 weeks.
 7     The authors hypothesized that exposure to acidic fog, followed by  O3 exposure, would
 8     induce greater decrements in  FVC and FEVt than H2O fog or air followed by O3 exposure.
 9     In fact,  both HNO3 and H2O fog exposure seemed to ameliorate the effect of subsequent
10     O3 exposure on FEVj and FVC, although only the difference between the FEVj responses to
11     FA + O3 and H2O fog + O3 was significant.  The results of comparisons between baseline
12     methacholine response and that following the exposures indicated that the O3 sensitive
13     subjects were more sensitive to methacholine than O3 nonsensitive subjects, who participated
14     only in the sensitivity screening O3 exposure, suggesting that O3 responsiveness may be
15     related to nonspecific airway  responsiveness.
16          Aris et al. (1993b)  further examined pulmonary responses to combined 0^ and HNO3
17     exposures. Ten healthy, nonsmoking adults, 19 to 41  years of age, were exposed to filtered
18     air, 500 /xg/m of HNO3 gas plus 0.20 ppm O3, or to 0.20 ppm O3 alone.  The exposure
19     protocol was  4 h in duration, with 50 min IE at 40 L/min alternating with 10-min rest
20     periods  each hour. Pulmonary function was measured during each rest period, whereas
21     BAL, proximal airway lavage, and bronchial biopsies were performed 18 h after completion
22     of each  exposure. Mean FEVj and FVC decreased, and mean SR^ and respiratory
23     symptom scores increased across both the HNO3  + O3 and O3 exposures.  The results
24     indicated, however, that HNO3 combined with  O3 did not exacerbate the pulmonary function
25     decrements or respiratory symptoms caused by  O3 alone. Similarly, there were no
26     statistically significant differences in the cellular or biochemical constituents in either the
27     BAL or proximal airway lavage fluids or in the bronchial biopsy specimens between the
28     HNO3 +  O3  and the O3 exposures.  The authors  concluded that HNO3 does not potentiate
29     the inflammatory response produced by O3 in healthy individuals.
30          The objective of a study by Koenig et al.  (1993) was to investigate possible interactions
31     between oxidants (0.12 ppm O3 + 0.30 ppm NO^ and H2SO4 aerosol (70 /*g/m3) or HNO3

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 1     aerosol (0.05 ppm).  Twenty-two adolescent allergic asthmatics who also had exercise-
 2     induced bronchospasm and a positive response to a standardized methacholine
 3     bronchochallenge test completed all exposures.  Subjects inhaled FA, O3 + NO2, O3  + NO2
 4     + H2SO4, or O3 + NO2 + HNO3 through a mouthpiece for 90 min on 2 consecutive days.
 5     Each pair of exposures was separated by at least 1 week. Subjects exercised (VE about
 6     10 times FVC) and rested in alternating 15-min periods. Pulmonary function (FVC, FEV1?
 7     Vmax50%, Vmax75%, and RT) were measured before and after each exposure and on the day
 8     following the second consecutive exposure, at which time only pulmonary  function was
 9     evaluated and a methacholine bronchochallenge was performed.  Six additional subjects
10     began the study, but dropped out before completion due to uncomfortable symptoms
11     associated with the exposures.  None dropped out following an FA exposure.  There were no
12     statistically significant changes in any measured parameter of pulmonary function following
13     the three pollutant-containing exposures, compared to following the FA exposure, contrary to
14     expectations.  (See also Section 7.4 for related epidemiological studies.)
15
16     7.2.6.3  Ozone, Peroxyacetyl Nitrate, and More Complex Mixtures
17          Horvath et al. (1986) exposed 10 healthy young women (19 to 36 years of age) to
18     (1) FA, (2) 0.48 ppm O3, (3) 0.27 ppm PAN, and (4) 0.48 ppm O3 + 0.27 ppm PAN. The
19     chamber exposures were 2 h in duration,  during which subjects alternated 20-min exercise
20     periods (VE  = 25 L/min) and 15-min rest periods. Exposures were completed in random
21     order, and were at least 1 week apart.  Exposure to PAN alone did not  induce any significant
22     changes in pulmonary function. Both O3 and PAN + O3 exposure induced significant
23     decrements in FVC, FEVj, and FEF25.75%; however, the decrements following the
24     PAN + O3 exposure were significantly larger (average of about 10%),  suggesting interaction
25     between PAN and O3. It should be noted that typical peak ambient PAN concentrations are
26     about 0.050 ppm. Symptom reports indicated that O3 + PAN exposure induced greater
27     subjective stress than exposure to O3 alone.
28          Drechsler-Parks et al. (1987b) exposed eight healthy young adults (mean age 24 years)
29     to a mixture of 0.30 ppm PAN + 0.45 ppm O3 on 5 consecutive days to evaluate possible
30     desensitization. Subjects were reexposed to the PAN + O3 mixture on the third and seventh
31     days following the last consecutive day of exposure to evaluate the  persistence of the

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 1     desensitization response.  Desensitization occurred with the same pattern and time sequence
 2     as has been reported for O3 alone.  The largest group mean decrements occurred following
 3     the second exposure,  and the subjects became progressively less responsive with subsequent
 4     exposures. Two subjects failed to desensitize fully with 5 consecutive days of exposure.
 5     Puhnonary function changes after the follow-up exposures indicated that the desensitization
 6     response is relatively short lived, in that the desensitization began to abate between 3 and
 7     7 days following the fifth consecutive day of exposure.  These results are consistent with
 8     those of similar studies using exposure to O3 only (Horvath et al., 1981; Kulle et al., 1982),
 9     suggesting that PAN had no additional effect on the desensitization response to O3.
10     A greater number of symptoms was reported following all PAN + Oj exposures than
11     following exposure to O3 alone.
12          Drechsler-Parks et al.  (1989) studied 16 older men and women (51 to 76 years of age)
13     and 16 young men and women (19 to 26 years of age) who each completed 2-h chamber
14     exposures to FA, 0.45 ppm O3,  and mixtures of 0.45 ppm O3 with 0.60 ppm NO2 and/or
15     0.13 ppm PAN.  Subjects alternated 20-min exercise (VE about 25 L/min) and rest periods.
16     Exposure to O3 alone and in all  combinations induced significant decrements in FVC, FEVj,
17     and FEF25_75% in the young group.  In the older group, these same three variables were
18     significantly decreased only with NO2 + O3 exposure.  Exposure of the older subjects to
19     PAN + O3 induced significant decrements only in FVC and FEVj. The PAN + NO2 +
20     O3 exposure induced a significant decrement only in FVC in the older subjects.  Both young
21     and older subjects reported  more symptoms following the mixture exposures than following
22     exposure to O3 alone.  These puhnonary function results following the exposure to
23     O3 + PAN are in contrast to those reported by Drechsler-Parks et al.  (1984) and Horvath
24     et al.  (1986) on young adults exposed to 0.45 ppm  O3  -I- 0.30 ppm PAN. The results of
25     both earlier studies suggested an interaction between O3 and PAN, in that puhnonary
26     function decrements following the mixture exposure were approximately 10% larger than
27     following exposure to O3 alone,  whereas there were no significant puhnonary function effects
28     with exposure  to PAN alone.  A likely explanation for this discrepancy is that the PAN
29     concentration used by Drechsler-Parks et al. (1989) was slightly less than half that used by
30     Drechsler-Parks et al.  (1984) and Horvath et al. (1986). Thus, if the additional effect of
31     PAN is linear, an additional effect of PAN + O3 would be expected to be less than 5%,

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 1     which would probably not be detected because it is within the variability of the pulmonary
 2     function measurements.  In any case, ambient PAN concentrations are considerably less than
 3     even 0.13 ppm. This indicates that even if PAN and O3 do interact in some way in their
 4     effects on pulmonary function, at typical ambient concentrations of Oj and PAN, effects can
 5     be attributed to O3 alone.
 6
 7     7.2.6.4   Summary
 8          Information on interactive effects between O3 and other pollutants remains sparse at this
 9     time.  However, several comments can be made when the material discussed above is
10     summarized collectively.  It is clear that O3 is responsible for the largest share of observed
11     effects when subjects are exposed to the mixtures of O3 and other pollutants that have been
12     studied to date. There is no evidence that simultaneous exposure of healthy individuals to
13     ambient concentrations of O3 plus NO2,  PAN, H2SO4, HNO3, or SO2 results in significant
14     interaction.  However, Aris et al. (1991) have reported that HNOg and H2O fog exposure
15     ameliorates the pulmonary function effects of a subsequent O3 exposure. Koenig et al.
16     (1990) found that preexposure to O3 induced significant pulmonary function decrements in
17     allergic asthmatic adolescents following a sequential SO2 exposure.  Both the O3 and SO2
18     concentrations  were at subthreshold levels.
19          Both studies that have reported potentially significant effects have involved sequential
20     exposure  protocols, in contrast to the simultaneous exposure protocols, which have not
21     generally shown effects beyond those that would be expected at the O3 concentration used.
22     It may be that certain preexposures predispose an individual to responses following a
23     subsequent exposure; however, this question  remains far from being resolved. Further, these
24     results are related only to spirometry and plethysmography and may not be applicable to
25     other possible endpoints.
26
27
28     7.3   SYMPTOMS AND PULMONARY FUNCTION IN CONTROLLED
29           STUDIES OF  AMBIENT AIR  EXPOSURES
30          Controlled O3 exposure studies under a variety of different experimental conditions
31     have generated a large amount of informative exposure-effects data.  However, complete

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 1     laboratory simulation of the pollutant mix present in ambient air is impossible on practical
 2     grounds.  Thus, the exposure effects of one or several artificially generated pollutants (i.e.,
 3     a simple mixture) on symptoms and lung function may not be comparable to those in ambient
 4     air where complex  mixtures of pollutants likely exist.  This section reviews two types of
 5     studies that utilize a mobile laboratory  or a hypobaric chamber to  investigate the acute effects
 6     of 63  during exposures to ambient air or altitude, respectively.  These studies can be
 7     designed to determine the independent  effects of O3 as well as possible interactions among
 8     many pollutants and other conditions present in typical ambient air.
 9
10     7.3.1   Mobile Laboratory Studies
11          Quantitatively useful information  on the effects of acute exposure to photochemical
12     oxidants on symptoms  and pulmonary function originated from field studies using a mobile
13     laboratory, as presented in the previous criteria document (U.S. Environmental Protection
14     Agency, 1986). These studies offer the advantage of studying the effects of ambient air on a
15     local subject  population by combining the experimental methods of both epidemiology and
16     controlled-exposure studies. Field studies using mobile exposure chambers can expose
17     subjects to ambient air, FA without pollutants, or FA containing artificially generated
18     concentrations of O3 that are comparable to those measured in the ambient environment.
19     The exposure air can also be conditioned to a desired temperature and humidity.  As a result,
20     measured health responses in ambient air can be compared to those found in more artificial
21     or controlled conditions. The mobile laboratory shares many of the same limitations of
22     stationary exposure laboratories (e.g., limited number of both subjects and artificially
23     generated pollutants for testing).  Ambient air studies in the mobile laboratory are dependent
24     on ambient conditions, which can be unpredictable, uncontrollable, and not completely
25     characterizable. Logistical problems (space, power, and locations with local interfering
26     outdoor conditions) limit access to many ambient pollution sites of interest.
27          As summarized in Table 7-14, investigators at the Rancho Los Amigos Medical Center
28     in California used a mobile laboratory  and demonstrated that respiratory effects in
29     Los Angeles  residents are related to O3 concentration and level of exercise (Linn et al.,
30     1980,  1983b; Avol  et al., 1983,  1984,  1985a,b,c, 1987). Such effects include pulmonary
31     function decrements at O3 concentrations of 0.144 ppm in exercising healthy adolescents

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                            TABLE 7-14.  ACUTE EFFECTS OF OZONE IN AMBIENT AIR IN
                                    FIELD STUDDZS WITH A MOBILE LABORATORY8
   Mean uzone
  Concentration
 ppm      pg/m
 0.113      22T"
 ± .033     ± 65
                      Amoient
Temperature    Exposure
    (°C)       Duration
             	Th—
   Activity
 Level ( Vg)     Number of Subjects
CE (22 L/min)  66 hei
         children
(8-11 years)
       Observed Effects)	Reference
 No significant changes in      Avol et al. (1987)
 forced expiratory
 function and respiratory
 symptoms after
 exposure to 0.113 ppm O3 in
 ambient air.
33 ± 1
 0.144      282        32  ± 1        TE     CE (32 L/min)  59 healthy adolescents
 ± .04S    ±84                                          (12-15 years)
"Olll300       32 ± 2        Tl     CE (53 L/min)  50 healthy adults
 ± .025     ± 49                                           (competitive bicyclists)
 Small significant decreases in   Avol et al. (1985a,b)
 FVC(-2.1%),FEV075
 (-4.0%), FEV! (-4.2%), and
 PEFR (-4.4%) relative to
 control with ao recovery during
 a 1 -h postexposure rest; no
 significant increases in
 symptoms.
 0.156      306       33 ± 4         Th     CE (38 L/min)  48 healthy adults
 ± .055     ± 107                                          50 asthmatic adults
Mild increases in lower
respiratory symptom
scores and significant decreases
in FEV, (-5.3%)andFVC;
mean changes in ambient air
were not statistically different
from those in purified air
containing 0.16 ppm 03.
                                                                                   Avol et al. (1984, 1985c)
No significant changes for total  Linn et al. (1983b)
symptom score or forced       Avol et al. (1983)
expiratory performance in
normals or asthmatics; however,
FEVj remained low or
decreased further (-3%) 3 h
after ambient air exposure in
asthmatics.
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TABLE 7-14 (cont'd). ACUTE EFFECTS OF OZONE IN AMBIENT AIR IN
        FIELD STUDIES WITH A MOBILE LABORATORY8
S3
f
K-»
^o
vo
LO
-J
i»i
*— *
•J
Mean Ozone
Concentration
ppm ng/m
0.165 323
± .059 ± 115
0.174 341
± .068 ± 133
Ambient
Temperature0
(°C)
33 ± 3
33 ± 2
Exposure
Duration
1 h
2h
Activity
Level ( Y£) Number of Subjects
CE (42 L/rnin) 60
(7
IE (2 X resting) 34
at 15-min 30
intervals
"healthy" adults
were asthmatic)
"healthy" adults
asthmatic adults
Observed Effect(s)
Small significant decreases in FEV i
(-3.3%) and FVC with no recovery
during a 1-h postexposure rest; TLC
decreased and AN2 increased slightly.
Increased symptom scores and small
significant decreases in FEVj
(-2.4%), FVC, PEFR, and
TLC in both asthmatic and healthy
subjects; however, 25/34 healthy
subjects were allergic and "atypically"
reactive to polluted ambient air.
Reference
Linn et al. (1983b)
Avol et al. (1983)
Linn et al. (1980, 1983b)
See glossary of terms and symbols for abbreviations and acronyms.
Ranked by lowest level of 63 in ambient air, presented as the mean ± SD.
cMean ± SD.
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 1     (Avol et al., 1985a,b) and increased respiratory symptoms and pulmonary function
 2     decrements at 0.153 ppm in heavily exercising athletes (Avol et al., 1984,  1985c) and at
 3     0.174 ppm in lightly exercising normal and asthmatic subjects (Linn et al., 1980,  1983b).
 4     The observed effects were typically mild, and generally no substantial differences were seen
 5     between asthmatic and nonasthmatic subjects.  Postexposure pulmonary function decrements
 6     appeared to last several hours longer in the asthmatics, but no significance test was reported
 7     for this difference (Avol et al., 1983;  Linn et al., 1983b). The medication status of the
 8     asthmatic subjects during the studies was not reported,  although medications were
 9     temporarily withheld prior to exposures.  The subjects' clinical severity was likely mild,
10     based upon their baseline lung function and exercise capability.  Many of the normal subjects
11     with a history of allergy appeared to be more responsive to O3 than "nonallergic" normal
12     subjects (Linn et al., 1980, 1983b), although a standardized evaluation of atopic status was
13     not performed.  Direct comparative studies of exercising athletes (Avol et al., 1984, 1985c)
14     with chamber exposures to oxidant-polluted ambient air (mean O3 concentration of 0.15 ppm)
15     and purified air containing a controlled concentration of generated O3 at 0.16 ppm showed no
16     significant differences in lung function and symptoms, suggesting that coexisting ambient
17     pollutants had minimal contribution to the measured responses under the typical summer
18     ambient conditions in Southern California.  Effects of copollutants in other regions of the
19     country remain to be investigated with the mobile laboratory. These field studies emphasize
20     the importance of adequate characterization of subjects and the ambient air, exercise levels,
21     duration of exposure, and individual variations hi sensitivity in interpreting observed
22     exposure effects.  Although these factors need to be investigated over a wider range of
23     experimental conditions, the results from these field studies are, so far, consistent with those
24     from controlled human  exposure studies.  Short-term respiratory effects of summer ambient
25     oxidant pollution in Southern California are predominantly, if not entirely, caused by ambient
26     O3 in typical healthy or asthmatic residents, according to mobile laboratory studies (Avol
27     etal., 1984, 1985c).
28
29     7.3.2   High-Altitude Studies
30           Symptoms and pulmonary function resulting from exposure to O3 in commercial aircraft
31     flying at high altitudes and in altitude-simulation studies were reviewed in the previous

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 1      criteria document (U.S. Environmental Protection Agency,  1986).  Much attention has
 2      focused on the health effects in flight crew and, specifically, flight attendants because of their
 3      physical activities at altitude and exposure patterns to peak  levels of cabin 03.  The most
 4      quantitatively useful information was based on a series of hypobaric studies of normal
 5      nonsmoking subjects who were exposed to 1,829 m (6,000 ft) and O3 at concentrations of
 6      0.2 and 0.3 ppm for 3 or 4 h (Lategola et al., 1980a,b).  Increased symptoms and pulmonary
 7      function decrements occurred at 0.30 ppm but not at 0.2 ppm under light exercise conditions.
 8      However, the exposure conditions did not reflect higher (peak) O3 concentrations reported to
 9      occur in certain aircraft at high altitudes or the higher cabin altitudes attained by new-
10      generation commercial aircraft.
11           No reports have subsjequently appeared in the literature that specifically study the health
12      effects of aircraft cabin O3.  However, O3 levels were reported to be very low (average
13      concentration 0.01  to 0.02 ppm) during 92 randomly selected smoking and nonsmoking
14     flights in 1989 (Nagda et al., 1991).  None of the flights exceeded the time-weighted average
15      standard  of 0.10 ppm (during any 3-h interval) promulgated by the U.S. Federal Aviation
16     Administration, perhaps related to the use of O3-scrubbing catalytic filters (Melton, 1990).
17     However, in-flight O3 exposure is possible because catalytic filters are not necessarily in
18      continuous use during flight.
19
20
21      7.4   FIELD AND EPIDEMIOLOGY STUDIES
22     7.4.1   Acute Effects of Ozone Exposure
23     7.4.1.1  Introduction
24          Field and epidemiology studies addressing the acute effects of O3 on lung function
25     decrements and increased morbidity and mortality in human populations involve those
26     combinations of environmental conditions, copollutant levels, and activity levels present
27     under real-world conditions of O3 exposure.  This real-world relevance is an advantage over
28     animal or human chamber studies.  Thus, results of such studies are essential components of
29     an understanding of overall effects of O3. However, the conditions under which
30     epidemiologic studies are carried out cannot be controlled in the same way that they can in
31     experimental studies.  Parameters that may be difficult or impossible to estimate or control

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 1     outside the laboratory include actual O3 exposures, levels of temperature, relative humidity,
 2     allergens, correlated pollutants other than O3, and breathing rates and activity patterns of
 3     subjects.  Variations in these factors can be important sources of variability in data and
 4     results, and may, under certain conditions, lead to biases (e.g., confounding) in results.
 5     These and other issues of importance in the interpretation of epidemiology study results are
 6     discussed in each relevant section below.
 7           The above noted weaknesses of epidemiologic studies of O3 health effects were
 8     highlighted in the previous O3 criteria document (U.S. Environmental Protection Agency,
 9     1986), which reached the conclusion that, because of such factors, epidemiologic studies on
10     the acute effects of O3 on lung function do not provide information that is quantitatively
11     useful in the standard-setting process.  Since  publication of 1986 O3 criteria document,
12     however, results have become available from a substantial number of well-conducted,
13     individual-level field studies and aggregate-level time-series studies.  In the following
14     section, these studies will be collectively evaluated to determine to what extent epidemiologic
15     studies may now play a quantitative role in the evaluation of the NAAQS for O3.
16
17     7.4.1.2   Individual-Level Studies
18           The studies discussed in this section fall into three main categories: summer camp
19     studies, exercise studies, and daily life studies. Summer camp  studies involve collection of
20     sequential (usually daily) data on lung function, respiratory symptoms, and environmental
21     conditions over the course of 1 or 2-week camps.  Exercise studies are unique in that lung
22     function and respiratory symptoms are measured before and after each of a series of discrete
23     exercise events in the presence of ambient air pollution.  Daily life studies measure lung
24     function, respiratory symptoms, and/or exacerbation of existing respiratory diseases, along
25     with environmental variables at regular intervals in the course of normal daily activities of a
26     population.  These  include studies in healthy adults and school-aged children as well as
27     studies in individuals with preexisting disease (e.g., asthma). Medication use may also be
28     monitored in asthmatics.
29           The important differences among the three study types relate mainly to issues of
30     exposure assessment.  Because subjects are mainly out-of-doors or in well-ventilated cabins,
31     exposure estimation errors are usually minimized in camp and exercise studies.  In contrast,

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 1     larger exposure estimation errors may occur in daily life studies.  Camp and daily life studies
 2     enable assessment of effects of cumulative O3  exposures, whereas exercise studies limit
 3     attention to rather brief exposures.  Exercise studies offer the potential of assessing individual
 4      VE values and O3 doses during the relevant exposure period.
 5           Although the study designs differ in some ways, the central design feature of all of
 6     these study types is the collection of repeated measurements on individuals.  This feature is
 7     exploited in data analysis  by having each subject serve as his or her own control. For
 8     continuous outcomes such as lung function, subject-specific linear regressions are usually
 9     performed with lung function (or change in lung function) as the outcome variable and 63  or
10     other environmental factors  as the explanatory variable(s).  The regression slope is a measure
11     of individual lung function response to O3. The mean slope across individuals is often used
12     as a measure of the average population response.  A more statistically valid approach
13     involves computing the mean slope with weighting proportional to the inverse variances of
14     the individual slopes.   An alternative approach has been to use analysis of covariance
15     methods to fit a population-pooled slope and separate, subject-specific y-intercepts.  To date,
16     no studies have used nonlinear (e.g., quadratic) models, which in chamber studies have been
17     shown to better describe the functional relationship between low-level O3 exposures  and lung
18     function decrements (Hazucha, 1987).
19
20     Issues in the Interpretation of Individual Level Studies
21           The most basic question affecting the interpretation of acute O3 epidemiology studies is
22     whether, and to what extent, the associations observed between O3 and decreased pulmonary
23     function are causally related to O3 and not merely due to confounding by some other
24     factor(s) (e.g., temperature, allergens, time trends in spirometry, or other pollutants).
25     By definition, a confounder is an unmeasured, or unaccounted-for, variable that both has an
26     effect on lung function and is also correlated with O3 concentrations.  Variables that satisfy
27     only one of these latter two conditions are not confounders.  For example, a variable that
28     affects lung function,  but is independent of O3, would add variation to lung function
29     measurements, but would not confound an O3/lung function analysis.  A variable that
30     correlates with O3, but does not directly affect lung function (in the range of measurements),
31     would not confound an analysis of O3 effects.  Other variables might modify the effect of

       December 1993                          7-121      DRAFT-DO NOT QUOTE  OR CITE
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 1     O3 on lung function, thereby increasing or decreasing the O3 effect under the conditions of
 2     study. Epidemiologists refer to this latter phenomenon as  "effect modification".  The
 3     presence of effect modification does not bias the results of a study, and also can provide
 4     insights into the range of effect magnitudes (e.g., slope of lung function development on
 5     O3 level) that occur under varying environmental conditions.
 6           Ambient air temperature often exhibits a moderate to high correlation over time with
 7     O3 in acute epidemiology studies due, in part, to the dependence of O3 formation rate on
 8     light intensity.  Among the studies reviewed in this section, correlations ranging from
 9     -0.06 to 0.90  (mean = 0.51) have been reported.  Correlations between O3 and relative
10     humidity, when reported, have been in the range  -0.4 to  -0.6.  Several human chamber
11     studies have examined the possible effects of temperature and relative humidity on lung
12     function independent of O3, with somewhat mixed results (Stacy et al., 1982; Folinsbee
13     et al., 1985;  Eschenbacher et al., 1992).  Two studies have reported  increases in FEVl at
14     high temperature (30 and 37 °C) and 60% relative humidity (Stacy et al., 1982;
15     Eschenbacher et al., 1992), whereas another reported no effect on FEVj at 35 °C, but a drop
16     at 40 °C (Folinsbee et al.,  1985).  Referring to results of acute Oj epidemiology studies,
17     Eschenbacher and colleagues (1992) concluded based on their results  that "the associations
18     found between  ambient O3 and daily changes in ventilatory function cannot be attributed to
19     the heat and  humidity stress often associated with high O3 concentrations."  Temperatures
20     observed in the epidemiology studies reviewed in the present section  have mainly been below
21     30 °C,  with  occasional peaks as high as 35 °C.  It should be noted that subjects studied
22     epidemiologically will usually have had an opportunity to acclimate to ambient temperatures
23     prior to, or soon after, the start of the  study. In any event, given the laboratory findings, a
24     significant confounding role for temperature in these studies is unlikely. The possibility
25     exists, however, that  temperature may  semetimes act as an effect modifier.   Folinsbee et  al.
26     (1977a) reported that high  temperature potentiated O3 effects on lung function, perhaps by
27     increasing VE  for a given workload. Effect modification in the opposite direction is also
28     possible due to curtailment of activity on very hot days.
29           Exposure to specific allergens can influence lung  function in individuals who have
30     diseases characterized by IgE-mediated antigen-antibody interactions  (i.e., atopy) and may
31     also affect individuals who have an atopic tendency (e.g., as assessed by positive prick skin

       December 1993                         7-122     DRAFT-DO NOT QUOTE OR CITE
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 1      test or serum levels of total IgE) without diagnosed clinical disease.  Raizenne and colleagues
 2      (1989) detected positive reactions to one or more allergens by skin prick in 49% of 96 young
 3      nonasthmatic females enrolled in a summer camp study.  Few data are available on the
 4      correlation between O3 and allergen levels during acute epidemiology studies.  However,
 5      because both variables are to some extent influenced by weather patterns, some correlation
 6      seems likely. Thus, a possible confounding role of airborne allergens in such studies cannot
 7      be ruled out. Because of the specific nature of individual antigen sensitization,  and
 8      uncertainty regarding the  full set of relevant allergens in a given setting, attempts to measure
 9      and statistically control for allergen levels on a group level in epidemiology studies may not
10      be very effective.
11           The potential effects of time trends in spirometry due to training effects are also of
12      concern.  There have been several recent studies that have looked at time trends in serial
13      lung function measurements (mainly FEVj and peak expiratory flow rate [PEFR])
14      independent  of air pollution effects (Raizenne et al., 1989; Avol et al., 1990; Hoek and
15      Brunekreef,  1992). In each case, average FEV, measurements have been observed to decline
16      steadily over the first few  measurements and then to stabilize or recover slightly to a flat
17      pattern.  Average FVC measurements follow a similar pattern. In contrast, PEFR often has
18      been observed to increase  steadily over successive measurements.  Similar patterns have been
19      observed in studies with intervals between lung function measurements ranging from half a
20      day to 1 week.  The consistency of these observations across studies suggests that they
21      represent real phenomena that should be recognized in designing and analyzing  studies
22      involving repeated lung function measurements.  However, time trends will result in
23      confounding of O3  effects  only if, by chance, the trend correlates with temporal variations in
24      O3 concentrations.  Such chance correlations could be either positive or negative, and, if
25      present, would have a larger impact  (i.e., produce an undesirable degree of confounding) on
26      studies in which all subjects begin the study simultaneously and have few follow-up
27      measurements.  Studies that focus on daily changes in lung function may be less impacted by
28      this phenomenon.
29           It is also important to consider the roles of other pollutants as possible confounders or
30      effect modifiers.  In the studies to be reviewed in this section, levels of measured
31      copollutants (e.g., SO2, NO2, sulfate and acid aerosols) were present at levels well below

        December  1993                          7_123      DRAFT-DO NOT QUOTE OR CITE
-------
 1      those that have produced lung function decrements in healthy subjects following short-term
 2      exposures in chamber studies (see Section 7.2.6).  In contrast, an extensive and growing
 3      database is available from chamber studies documenting the independent acute effects of
 4      ambient-level O3 on lung function (see Section 7.2.1).  Although direct lung function effects
 5      of other pollutants at typical ambient concentrations  seems unlikely, it has been suggested
 6      that the effects of O3 may be potentiated by coexposure or previous exposure to other
 7      pollutants, most notably acid aerosols (Spektor et al., 1988b).  Some data from animal
 8      studies suggest interactive effects of O3 and acid exposures for certain pulmonary outcomes
 9      (see Chapter 6).  However, to date, analyses directed towards this phenomenon in field
10      studies of human lung function (via analysis of the relationship between acid aerosol levels
11      and residuals  from  regressions of lung function on C^)  have proven negative (Spektor et al.,
12      1988a,b).  That is, after controlling for the influence of O3, no significant association
13      between acid  aerosol peaks  and lung function decrements  has been  observed.  Acid aerosol
14      episodes,  which often occur coincident with high O3 levels in the summer in the northeastern
15      United States, may extend for several hours or days. The possible potentiating effects of
16      prolonged acid peaks on O3 effects require further study under controlled conditions before
17      this issue can be further resolved.
18           In epidemiologic studies, activity levels are difficult to control and to measure, although
19      this varies with study type (see below).  Chamber studies have shown clearly that lung
20      O3 doses  and associated functional effects increase as a function of physical activity level
21      (Hazucha, 1987).  Epidemiologic study designs have often been chosen that result in
22      relatively high subject activity levels (exercise studies and camp studies), but generally the
23      studies have been carried out without quantitative  information on VE distributions across
24      subjects and across time.
25           Variations in  activity levels will introduce variability in the relationship between
26      personal exposure and personal dose.  If this variability occurs primarily between subjects, it
27      will result in  differing O3 doses to people exposed to the  same O3 level and yield differences
28      in response that may be misinterpreted as O3 sensitivity variations.  If variability in activity
29      levels occurs  over time for  a given subject, it will add error to the functional relationship
30      linking lung function and O3 exposure.  In either case,  the influence of activity level
31      variability is to add dose estimation error (or misclassification).  Estimates of VE based on

        December 1993                           7-124     DRAFT-DO NOT QUOTE OR CITE
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 1     heart rate measurements can be derived using subject-specific calibrations under
 2     representative ranges of exercise levels and types (Samet et al., 1993; Raizenne and
 3     Spengler, 1989).  However, the utility of such VE estimates for reducing dose-
 4     misclassification errors in acute O3 epidemiology studies has not yet been demonstrated
 5     (Kinney, 1986; Spektor et al., 1988b; Raizenne and Spengler, 1989). This is partly due to
 6     the logistical difficulties associated with collecting accurate data, and may also be due to the
 7     fact that, for a given subject, VE variations across days are usually small in comparison to
 8     O3 concentration variations.  The same issues arise in the context of O3 exposure
 9     misclassification in "daily life"  studies (see below), where outdoor O3 concentrations are
10     used to estimate exposures of subjects who spend substantial amounts of time indoors during
11     the period over which lung function measurements take place.
12
13     Camp Studies of Lung Function in Children
14           Summer camp studies provide the most extensive and reliable information on the acute
15     pulmonary effects  of O3 under natural conditions.  Camp studies involve the collection of
16     sequential (usually daily) data on lung function on each of a large number of children, along
17     with concurrent measurements of O3 exposures  and other environmental factors over the
18     course of a single-week or multiweek summer camp. Data analyses usually consist of
19     estimating the linear association between lung function and environmental variables on an
20     individual basis (allowing each  subject to serve  as his/her own control) and then testing the
21     mean population association for statistical significance.  As noted,  summer camps offer the
22     significant advantage that subject exposures are  especially well estimated based upon on-site,
23     outdoor O3 monitoring.  In addition, these studies assess the pulmonary effects of natural
24     diurnal patterns of O3 exposures,  which often involve broad daytime peaks.
25           Since the last O3  criteria document (U.S. Environmental Protection Agency,  1986),
26     eight camp studies have been reported. Design characteristics and results are summarized in
27     Table 7-15.  Six of these studies have focused just on normal (i.e., nonasthmatic) children,
28     one focused on asthmatics exclusively (Thurston et al.,  1993c), and one looked at both
29     normal and asthmatic children (Raizenne et al.,  1987).  Although methods and results varied
30     somewhat across studies, this group of studies collectively provides substantial evidence for
       December 1993                          7-125      DRAFT-DO NOT QUOTE OR CITE
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                         TABLE 7-15.  ACUTE EFFECTS OF PHOTOCHEMICAL OXTOANT POLLUTION:
                                                       LUNG FUNCTION IN  CAMP STUDIES8
       Pollutants/Environmental Variables
              Study Description
                                                                                                       Results and Comments
                                                     Reference
Hourly O3 ranged from = 10 to 110 ppb.  SOj,
NOX, 03, S04~, H2S04, pH, PM)0, PM2.5, RH,
T, barometric pressure, wind speed, and direction
were also measured.
Effects of pollutants and other environmental
variables on symptoms and lung function were
examined in children attending a summer camp at
Lake Couchiching, about 100 km norm of
Toronto, ON. Study was conducted 6/30-7/8/83;
n = 52, 23 nonasthmatic (11 males, 12 females)
and 29 asthmatics (16 males, 13 females), avg.
age =  12.1 years.  Symptom questionnaire and
function tests given twice daily to each child
between 7:30-9:30a.m. and 4:30-6:3* p.m.
Children's activity levels net estimated.
Strongest association between lung function
and environmental variables was in
nonasthmatics, with FVC decrements
significantly correlated (p < 0.01) with
lagged-avg. 804, PM2.5, and T.  Unlagged
PEFR significantly correlated with 1 h Oj.
Also, significant association of T with all
lung function indices in nonasthmatics, but
not in asthmatics. Coefficient of variation
stable across morning and evening tests.
Raizenne et al. (1987)
1-h Oj ranged from < It to 143 ppb; max 1-h Oj
> 10» ppb on 14 days of total study (6 weeks).
For other pollutants and variables measured, see
Raizenne et al. (1987) because same protocol used
here as in *at study.
(a) Effects of pollutants and other environmental
variables on lung function were examined in girls
attending one of three 2-week Girl Guide camp
sessions on north shore of Lake Erie.  Cohort (n
= 194) screened by MC and skin prick testa for
!• common respiratory allergens; 5 asthmatics
withdrawn from the study (n = 99). Lung
fiinction tests administered twice daily. Children's
activity levels net estimated.
(») Subset of 12 girls (7 MC + , 5 MC-) studied
pre- and postexercise on 1 low-pollution (control)
day and 1 peak pollution dav (episode, 1 h Oj >
139 ppb, S04~ >  80pg/m ).
(a) Associations between aerometric data
and lung function measurements were not
reported by pollutant in this reference.
Aggregate analysis for full study not
reported. Lung function changes reported
for 5 episode days only. FEVj decrements
statistically significant on 2 episode days for
methacholine nonrespensive subjects.

(b) Group mean FVC increased postexercise
in the n = 12 subset by 46 mL. 71 mL in
MC- and 17 mL in MC+. Pollution effect
not statistically significant
Raizenne et al. (1917; 1989)
Continuous 1-h Oj, S(>2, N(>2, and acid aerosols
(as r^SOjj); 1-h 03 range = 40-143 ppb; max
12-h acid particle concentration = 28 fig/m in
one episode; FP SO4   100 /zg/m  for peak hour.
Time-activity model used to evaluate likely
cumulative (6 h) Oj and H2SO4 exposures/doses
experienced by children in above Lake Erie Girl
Guide camp study, summer 1986.  See Raizenne
et al. (1987, 1989) for protocol and related
information.  Dosimetry model was developed for
relating heart rate (from a 12-min, graded cycle
ergometer test) to ventilation and then to Oj and
H2SO4 dose. Also, five randomly selected
children wore portable heart-rate monitors,
providing data for use in the dosimetric model.
Application of the dosimetry model used to
estimate individual 6-h cumulative doses for
03 and H2SO4 exposures on one control
and one episode day indicated negative
trend in lung function (PEFR) as cumulative
dose increased for both 03 and HjSC^,
although slopes for each did not differ
significantly from zero (p > 0.10).
Raizenne and Spengler (1989)
-------
1
if
Co
                         TABLE 7-15 (cont'd).   ACUTE EFFECTS OF PHOTOCHEMICAL OXIDANT POLLUTION:
                                                            LUNG FUNCTION IN CAMP STUDIES3
              Pollutants/Environmental Variables
             Study Description
                                                                                                         Results and Comments
                                                                                                                                                    Reference
•^J
I—*
to
       Max 1 h 03 ranged from 40 to »100 ppb, with
       max 1 h > 80 ppb on 9 of 27 days of Oj
       recorded  O3, SO4 H2SO4, PM15, PM2.5, T,
       humidity, and wind speed and direction measured.
       Levels not reported for SC>2, pH, NC^, and
Effects of pollutants and other environmental
variables on respiratory functions in 91 children
(53 boys, 38 girls; ages 8-15) attending 2 to
4 weeks of summer camp at Fairview Lake, NJ.
Subsets were n = 37 for all 4 weeks, n = 34 for
first 2 weeks only, n = 20 for last 2 weeks only.
Symptom questionnaire; FVC, FEVj, MMEF by
spirometry; and PEFR by mini-Wright flow
meter were measured once/test day (most of days
in camp) sometime between 11:00 a.m. and
6:30 p.m.  All children had validated spirometric
data for  >7 days of their 2- or 4-week camp
stay. Activity levels of the children were not
estimated.  Respiratory health status determined
by parental questionnaire only. Children slept in
screened-in shelters but otherwise were exposed
to ambient air 24 h/day.  Average regression
slopes for respiratory function vs. max. 1-h Oj
concentration reported for the full cohort, for
boys and girls separately, and for subsets in
attendance for all 4 weeks and for respective
2-week sessions.  Regressions also repeated for
data below  80 and 60 ppb 1-h 03, and for data
with THI < 78 °F.
Average regression slopes (±SE) were
-1.03 ± 0.24 and -1.42 + 0.17 mL/ppb for
FVC and FEVj, respectively; and -6.78 ±
0.73 and -2.48 ± 0.26 mL/s/ppb for PEFR
and MMEF, respectively.  Most slopes of
regression significant at p < 0.05
(differences from zero).  Not clear if slopes
for data subsets significantly different from
each other (e.g., function vs. 03 < 60 ppb
and function vs. 03 < 80 ppb).  No formal
analysis performed for possible concentration
threshold.
                                                                                                                                         Spektor et al. (1988a)
-------
                          TABLE 7-15 (cont'd).  ACUTE EFFECTS OF PHOTOCHEMICAL OXTOANT  POLLUTION:
                                                             LUNG FUNCTION IN CAMP STUDIES3
              Pollutants/Environmental Variables
             Study Description
                                                                                                            Results and Comments
                                                      Reference
NJ
09
       Maximal I h 03 concentrations ranged from
       approximately 40 to 150 ppb over the course of
       the study.  12-h average aerosol acidity
       measurements ranged from near 0 to 18.6 fig/m
       (H2SC>4 equivalent).  T and RH measured but
       levels not reported.  THI reached a maximum of
       81 °F. All environmental measurements were
       made on site.
Effects of 03 and other environmental variables
on lung function studied in a group of
46 children (13 girls, 33 boys; ages 8-14) at a
4-week, 1988 summer camp in southwestern
New Jersey (Fair-view Lake). Same location
used in previous camp study by same
investigators.  Subjects had no history of lung
diseases or atopy. Two lung function
measurement periods each day (a.m. and p.m.)
along with collection of respiratory symptom
data. Datt collected during or after periods of
rain were excluded from analysis. Results for
FVC, FEVt, FEVi/FVC, FEF25.75%, and
PEFR reported. Linear day-of-study  trends were
examined for lung function. Subject-specific
linear regressions were performed relating lung
function in a.m., p.m., and p.m.-a.m.
differences to 03 averaged over various periods.
Average slopes across subjects were tested for
significant differences from zero.  Regressions
were repeated after excluding days with 03 at or
above 120 ppb. Regression residuals  were  tested
for correlation with THI and H   concentrations.
No significant linear day-of-study effect seen
for any of the lung function variables tested,
but the linear model may not have been
optimal for testing this effect. In a subset of
35 subjects with at least 2 consecutive days of
lung function measurements, mean regression
slopes of a.m. lung function variables on
previous-day mean or 1-h maximum 03 were
all significantly negative (e.g., mean  slope of
FEVi on 1-h maximum 03 was -0.50
±0.12 mL/ppb).  These results suggest a
possible carry-over effect from previous-day
03 exposures.  In the full set of 46 subjects,
regressions of p.m. lung function on previous
hour 03, maximum 1-h 03 for same day, or
average 03 for day were significantly
negative in most cases (e.g., mean slope of
FEVj on previous hour 03 was -1.60
±0.30 mL/ppb).  All regressions of the
p.m.-a.m. lung function differences on
intervening 03 concentrations were
significantly negative (e.g., mean slope of
FEVj on mean 03 between a.m. and p.m.
measurements was -0.63 {+ 0.09] mL/ppb).
No correlation seen between regression
residuals and THI or H   concentrations,
indicating there was no remaining effect of
these variables with lung function after
accounting for O3. However, no models
were fit that included 03 and these variables
simultaneously, nor were interaction effects
tested for.  The strong and consistent
associations between lung function
decrements and 03 concentrations in this
study contrast  with results reported from
studies in Canada  and California at similar
levels of 03.
Spektoret al. (1991);
Spektor and Lippmann (1991)
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 I
 U>
                          TABLE 7-15 (cont'd).  ACUTE EFFECTS OF PHOTOCHEMICAL OXTOANT POLLUTION:
                                                              LUNG FUNCTION  IN CAMP STUDIES3
       Pollutants/Environmental Variables
                                                                   Study Description
                                                                                                     Results and Comments
                                                                                                                                                 Reference
-J
to
1 h O3 preceding lung function measurements
ranged from 25 to 245 ppb.  Pollutants measured
on site included O3; NO2 (range: 0 to 40 ppb),
SO2 (range.  1 to 8 ppb); and fine (mean =
23.9 ng/m ), coarse (mean = 36.6 fig/m ), and
total (mean = 59 jug/m ) PMjg mass.  T averaged
21.5 °C  (range:  13.5 to 25.5 °C) and RH
averaged 43.3%.
Effects of O3 and other environmental factors on
lung function examined in 43 children
(24 female, 19 male; ages 7-13) attending one of
3 sequential weeks (three subjects stayed an
additional week) of summer camp in the
San Bernardino Mountains of California.  Camp
was at 5,710 ft above sea level. Lung function
measured by spirometry up to three times daily
on each subject; analytical measures included
FVC, FEV^ and PEFR.  No report of
respiratory data derived from questionnaires.
Subject activity levels prior to lung function
testing were not characterized.  Campers slept in
well-ventilated cabins.  Subjects came mostly
from homes in the Los Angeles Basin, and were
thus likely to have been exposed to high
O3 levels prior to camp. Simple linear
regression models were fit on an individual basis
(subject-specific slopes) and by pooling across
individuals (common population slope) to
determine the linear relations between the three
lung function variables and various O3 metrics
(1-h average preceding hour of spirometry, 1-h
average 2 h previous to hour of spirometry, or
6-h average preceding spirometry).  The
common slope model was repeated separately for
morning,  noon, and evening lung function
measurements, and separately for data with 1-h
O3 levels above and below 120 ppb. Multiple
regression models were fit that  included O3
along with T, RH, and coarse and fine
paniculate matter mass.
The population-pooled regression slopes
(+SE) of FVC and FEVj on previous hour
O3 were -0.40 (+  0.10) and -0.38
(± 0.09) mL/ppb,  respectively (p < 0.0001
in both cases); for  PEFR, the regression
slope was -0.13 (+ 0.36) mL/s/ppb (not
significant).  Similar, though slightly more
negative, slopes were obtained using 2-h and
6-h average O3. Interpretation of differences
across the three O3 metrics is substantially
hampered by the high correlations among
them (r > 0.90).  When T, RH, and coarse
and fine paniculate matter mass were
included with O3 in multiple regression
models, the O3 slopes increased in  absolute
magnitude to -0.68 (+ 0.16) and -0.76 (±
0.15) mL/ppb for FVC and FEVj,
respectively, and to -1.91  (± 0.63) for
PEFR. Technical problems with the
temperature sensor in the first week of the
study did not appear to influence these
results. Data were split on the basis of
whether or not the  maximum 1-h O3
concentration in the 6 h preceding spirometry
was above 120 ppb. Regression slopes
relating lung function and previous  1-h, 3-h,
and 6-h average O3 were more negative, and
only statistically significant, in the high
concentration stratum.  This result is
consistent with the  nonlinear (e.g., quadratic)
relationships between lung function and 03
exposure observed in chamber studies.
Because levels of pollutants other than
O3 were quite low  (NOj and SO^, and/or
were uncorrelated with 03 levels (paniculate
matter), the regression results reported from
this well-conducted study are likely to
represent real influences of 03 on lung
function.
                                                                                                                                             Higgins et al. (1990);
                                                                                                                                             Gross etal. (1991)
-------
                   TABLE  7-15 (cont'd).  ACUTE  EFFECTS OF PHOTOCHEMICAL  OXTOANT POLLUTION:
                                                       LUNG FUNCTION  IN CAMP STUDIES8
       Pollutants/Environmental Variables
              Study Description
                                                                                                      Results and Comments
                                                                                                    Reference
Daily maximum Oj concentrations ranged from
approximately 60 to  160 ppb (derived visually
from figure presented in paper).  Other pollutants
measured on site included SOj, NO2, CO, total
hydrocarbons, and size-segregated paniculate
matter mass.  Aside from 03, all gaseous pollutant
levels reported to be very low (data not
presented). 24-h TSP concentrations ranged from
H to 54 ng/m . Airborne allergen data collected.
T ranged from 1« t» 15 °C at night and from
25 to 35 °C during day. RH ranged from 39 to
45% at night and from 5 t» 29% during day.
Effects of 03 and other environmental variables
on lung iunction examined in 293 children
(139 girls,  154 boys; ages 8-17) attending one of
six 1-week camp sessions at a summer camp
located in the mountains near Idyllwild, CA,
190 km southeast of Los Angeles (altitude:
1,570 m).  Lung function measured twice daily
on each camper (a.m.:  0730 to 0930; p.m.:
16OB to 1930). Analyses presented for FVC,
FEVj, PEFR, and fEe^_i^%.  Symptom
questionnaires completed prior to each test.
Used repeated measures analysis of variance
model to test for day-of-study and a.m./p.m.
effects on lung function independent of pollution
concentrations. Linear regressions of morning,
afternoon, and p.m.-a.m. difference of lung
function on 03 were performed with
simultaneous control of day and a.m./p.m.
effects. Upper and lower quartiles of
distribution of individual FEV|/Oj regression
slopes were examined with respect to subject
characteristics. Changes in FEVj over several
days analyzed  in relation to intervening
integrated 03 concentrations.
Significant day-of-study effect observed for
FVC and FEVj characterized by steady drop
over first few days of measurement, followed
by partial reversal later in week.  For PEFR,
p.m. measurements were significantly higher
than a.m. measurements.  Controlling for day
and a.m./p.m. effects, the authors reported
that no consistent 03 effects on lung function
were observed. The a.m. lung function
measurements had a significant positive
correlation with 03 averaged over the
previous 1,  8, or  24 h.  The p.m.
measurements reported  to have no correlation
with 03.  p.m.-a.m.  lung function differences
were negatively correlated with previous 8-h
average 03  concentrations, but not with
previous 1-h 03 concentrations. No
quantitative results reported for the above
lung function/03 findings.  There were no
discernable  differences between subjects in
the upper and lower quartiles of the
distribution  of individual regression slopes of
a.m.-p.m. FEVj difference on previous 1 h
03. Regressions  of change in FEVj over
several days (four separate intervals ranging
from approximately 8 h to approximately
80 h) with integrated 03 concentrations
yielded negative slopes  ranging from -0.41 to
-1.46 mL/ppb, one of which was statistically
significant.  The time-trends in FVC and
FEVj measurements observed in this  study
are qualitatively consistent with those seen in
some other summer camp studies. The lack
of consistent negative slopes relating lung
function with 03 concentrations contrasts
with other, eastern U.S., summer camp
studies at similar 03 levels.
Avol et al. (1990);
Avol et al. (1991)
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I
VO
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U>
                    TABLE 7-15  (cont'd).   ACUTE EFFECTS OF PHOTOCHEMICAL OXIDANT POLLUTION:
                                                       LUNG FUNCTION  IN CAMP STUDIES3
        Pollutants/Environmental Variables
                                                            Study Description
                                                                                                           Results and Comments
                                                      Reference
 O3 data collected at a site 8 mi from camps
 Daily 1-h O3 maxima ranged from approximately
 40 ppb to approximately 200 ppb.  12-h aerosol
 acidity concentrations ranged between 14 and
 360 neq/m .  T and RH data obtained from a
 nearby site.
                                                    Report of data collected during two simultaneous
                                                    summer camps located 2 mi apart in central New
                                                    Jersey in 1988. 34 subjects were studied,
                                                    including 20 camp counselors (ages 14-35) and
                                                    14 campers (ages  9-13).  Study spanned 19 days.
                                                    Spirometry and respiratory symptom data
                                                    collected each afternoon.  Analysis of FVC,
                                                    FEV],  and PEFR in relation to Oj and T using
                                                    linear regression within camps and subject types
                                                    (i.e., counselors vs. campers).
Regressions of lung function on 1-h and 8-h
average O3 within several subject subsets
yielded inconsistent results, with some mean
slopes apparently significantly positive, and
one negative mean  slope, highlighted by
authors, of borderline significance
(p < 0.10).
                                                                                                                                           Berry et al. (1991)
 Daily 1-h maximum O3 concentrations ranged
 from 70 to  160 ppb in 1991 and from 10 to
 63 ppb  in 1992  On-site measurements also made
 for acid aerosols  (approximately 20 to
 110 nmoles>/m in 1991 and 15 to 55 nmoles/m  in
 1992) and temperature (between 21 and 32 °C
 over 2 years)
                                                    Effects of O3 and acid aerosols on peak flow,
                                                    respiratory symptoms, and medication usage in
                                                    asthmatic children evaluated at two 1-week
                                                    summer camps (June of 1991  and 1992) in the
                                                    Connecticut River Valley. 52 and 55 subjects
                                                    were studied in 1991 and 1992, respectively,
                                                    ranging in age from 7-13. Peak flow measured
                                                    twice daily (approximately 9:00 a.m. and
                                                    5:00 p.m.).  Combining data from the two
                                                    studies, individual regressions of daily change in
                                                    FEVj on O3 or H  concentrations were
                                                    performed.
In subjects without asthma exacerbations
during the camps, statistically significant,
negative mean slopes were found relating
APEFR and O3 or H  concentrations.  The
correlation between these two pollutants was
not reported. The mean slopes were
-2.3 (± 0.7) mL/s/ppb for O3, and
-1.2 (± 0.6) mL/s/nmol/m for H+. In the
case of O3, a scatter-plot with APEFR
demonstrated an apparently linear trend.
In contrast, the H  regression results
appeared to be driven entirely by one data
point.
                                                                                                                                           Thurstonet al. (1993c)
See glossary of terms and symbols for abbreviations and acronyms.
Cited in U.S. Environmental Protection Agency (1992).
-------
 1     associations between ambient O3 exposures, together with other pollutants, and acute
 2     decrements in lung function. Interpretation of these associations as causal is supported by
 3     evidence of biological plausibility.  For example, the well-documented direct effects of O3 on
 4     lung function in human chamber studies; the evidence, also from chamber studies, indicating
 5     a lack of direct effects of other collinear environmental factors (e.g., temperature and acid
 6     aerosols) at the levels these factors occur in the camp studies reported to date; exposure-
 7     response relationships; and consistency across studies all provide strong support.  Camp
 8     studies involving asthmatic children have generally yielded lung function/O3 associations that
 9     are similar in absolute magnitude to those observed in  nonasthmatics (Raizenne et al., 1987;
10     Thurston et al., 1993c); however,  the health significance of a given drop in FEVj  may be
11     greater for those with preexisting,  compromised respiratory function.
12           Although similar study designs have been employed in most of the camp studies
13     summarized in Table 7-15, differences in analytical methods have made quantitative
14     comparisons between studies difficult to  interpret. In particular, it has not been clear to  what
15     extent differences in results across studies may be due  to differences in study characteristics
16     (e.g.,  O3 effect potentiation by other pollutants and activity levels) as opposed to differences
17     in data analysis methods.
18           Recently, Kinney  et al. (1993) reported a reanalysis of data from six of the camp
19     studies summarized in Table 7-15  using uniform analytical methods. For each  study,
20     afternoon lung function data (FEV,) were regressed on concurrent 1-h O3 concentrations
21     using  an analysis of covariance model that included subject-specific intercepts and a single,
22     pooled slope on O3.  The study-specific  slopes computed with this model ranged from
23      -0.29 to  -1.34 mL/ppb across the six studies (Table 7-16).  All but one of these slopes
24     were statistically significant (p <  0.02).  The inverse  standard-error weighted mean slope for
25     the six studies  was -0.64 mL/ppb (p = 0.008).  Slopes from the two Fairview Lake, NJ,
26     studies (about -1.3 mL/ppb) were consistently greater in absolute magnitude than the slopes
27     from the other four studies (which ranged from about  -0.3 to -0.5 mL/ppb).  Overall,
28     however, these results indicate a quantitative consistency among studies that is  not as readily
29     apparent in the absence of the combined analysis.
30           It is  not clear why the New Jersey studies have yielded larger slopes  than the other
31      studies.  Possible explanations include greater subject  activity levels (resulting in higher

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            TABLE 7-16.  SLOPES FROM REGRESSIONS OF FORCED EXPIRATORY
               VOLUME IN ONE SECOND ON OZONE FOR SIX CAMP STUDIES8
Study Name
Fairview Lake, 1984
Fairview Lake, 1988
Lake Couchiching
Lake Erie
San Bernardino Mountains
Pine Springs Ranch
All studies
Slope ± SE (mL/ppb)
-1.34 ± 0.13
-1.29 ± 0.28
-0.44 ± 0.51
-0.29 ± 0.10
-0.47 ± 0.13
-0.32 ± 0.13
-0.64 ± 0.18
p-Value
<0.0001
0.0001
0.39
0.003
0.0006
0.013
0.008
Reference
Spektor et al. (1988a)
Spektor et al. (1991)
Spektor and Lippmann (1991)
Raizenne et al. (1987)
Raizenne et al. (1987, 1989)
Higgins et al. (1990)
Gross et al. (1991a)
Avol et al. (1990, 1991)

       b
lFor each study, data were analyzed in one model that fit subject-specific intercepts, but one, pooled,
O3 slope.  See glossary of terms and symbols for abbreviations and acronyms.
'Slope is the weighted mean of six study-specific slopes. The SE is the weighted SE of mean slope.
       Source:  Kinney et al. (1993).


 1      O3 doses at a given exposure level), potentiation of the O3 effect by other pollutants (such as
 2      acid aerosols), the relative absence of O3 tolerance in the New Jersey studies, or confounding
 3      by airborne allergens.  There are no firm data on activity levels across the six studies.  Thus,
 4      whereas this factor surely contributes to the random  variability within and between studies, it
 5      is not known whether activity levels were substantially and systematically higher in the
 6      New Jersey studies.  Potentiation of the O3 effects on lung function in asthmatics by acid
 7      aerosols has been demonstrated in a recent chamber  study in which O3 exposure was
 8      administered  1 day following a 3-h exposure to  100  jwg/m3 H2SO4 (Frampton et al., 1992).
 9      Although the relevance of these data to the nonasthmatic subjects who experienced much
10      lower acid levels at northeastern summer-camps is not clear, they do demonstrate that
11      potentiation can  occur between these pollutants.  However, this factor cannot, alone, explain
12      the observed  differences across camp results,  because a camp study in southern Ontario
13      (Raizenne et al., 1989), which yielded relatively low FEVl  slopes on O3, experienced sulfate
14     aerosol levels that were comparable to those seen in New Jersey.  Similarly, whereas
15      tolerance due to prior exposures to high O3 levels has  been suggested as an  explanation for
16     the smaller slopes seen in the California studies, it cannot explain the smaller slopes in
17     southern Ontario.  Data have not been reported  on comparative levels of airborne allergens

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 1     during the various camp studies. None of the subjects in the two New Jersey studies
 2     reported a history of asthma or atopy, minimizing the likelihood of confounding by airborne
 3     allergens. However, given the lack of allergen data, and the potential for substantial
 4     numbers of "silent hyper-responders" (Raizenne et al., 1989), this possibility cannot be
 5     completely discounted. Thus,  no one factor seems adequate to explain the differences in
 6     results across studies. Quite possibly these differences reflect the combined influence of
 7     several of the factors discussed above.  Indeed, given the many possible sources of camp-to-
 8     camp variability, it is surprising that results are as consistent as they are across six studies by
 9     three investigative groups.
10           Several investigators have reported regression results both for 1-h average O3 and for
11     longer averaging times (e.g., 6 to 8 h) (Higgins et al., 1990; Avol et al., 1990, 1991;
12     Spektor et al., 1988a; Spektor et al., 1991).  In general, similar results have been obtained
13     regardless of the averaging time.  Attempts to draw conclusions  regarding the relative
14     importance of short-term  peaks and longer term  averages from such analyses have been
15     hampered by the high degree of correlation between 1-h and multihour averages.   Until
16     better analytical methods  are found for dealing with this problem, comparative results will
17     remain difficult to interpret.
18
19     Lung Function in Exercising  Subjects
20           This subsection discusses studies involving lung function measurement immediately
21     before and after a series of discrete outdoor exercise activities in the presence of air
22     pollution. This design is similar in principle to the ambient chamber studies conducted in the
23     early 1980's (see Section 7.3), in which subjects exercised under a specified protocol in a
24     chamber ventilated with ambient air.  Here, however, there  is typically less control imposed
25     over exercise duration and intensity, and less assessment of achieved VE.  Compensating  to
26     some extent for this diminished control is the relative ease of collecting numerous repeated
27     measurements at varying ambient O3 levels on the same subjects, improving the precision of
28     dose-response estimation.  In contrast to camp studies, duration of relevant 03 exposure is
29     assumed to be known—as defined by the length  of each exercise event.
30           Results from three exercise studies (Selwyn et al., 1985; Spektor et al., 1988b; Hoek
31     and Brunekreef, 1992; Hoek et al., 1993a) are summarized in Table 7-17.  One of the

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                                TABLE 7-17.  ACUTE EFFECTS OF PHOTOCHEMICAL OXIDANT POLLUTION:
                                                       LUNG FUNCTION IN EXERCISING SUBJECTS3
               Pollutants/Environmental Variables
                                                                   Study Description
                                                       Results and Comments
                                                                                                   Reference
vj

OJ
        1-h 03 concentration ranged from 21-124 ppb,
        max. THI =78 °C: max acidic aerosol (as
        H2SO4) = 9 /tg/m _during study. SO2, NOX,
        PM,5> PM2.5, S04~, N03, NH4+, T, and RH
        measured but not reported.
Effects of Og on respiratory function and
symptoms examined in 30 nonsmoking adults
(2 of 10 females non-Caucasian) exercising
almost daily outdoors (Tuxedo, NY) for 15 to
55 min (average ca. 30 min) from July to early
August, 1985. Pre- and postexercise lung
function measured and questionnaire answered
postexercise. Pulse rate, calibrated to VE
indoors, taken postexercise. Exercise regimen
self-selected but constant for each subject over
the course of study. Dosimetry estimated and
linear regressions done for pulmonary function
changes vs. (1) mean O3 concentration during
exercise, and (2) inhaled Oj dose.  Persistence
of effects tested by linear regressions of before-
exercise lung function on previous-day O3 during
exercise. Subjects screened only  by
questionnaire; two with previous history of
asthma  but asymptomatic.
Significant (p < 0.01) decrements in FVC,
FEV^ PEFR, FEF25_75%, and FEV,/FVC
associated with O3.  For example, the mean
slope of AFEVj on O3 across all subjects
was -1.35 mL/ppb  (+  0.35).  No
persistence of effects seen. No symptoms
reported by subjects. Mean decrements
showed unexpected inverse relationship with
calculated Vg levels, as indicated by
regressing pulmonary function changes and
postexercise function against inhaled O3
during exercise.  VE ranges given, but not
group or subset means.  Subjects not
screened for atopy.  Exercise done in Sterling
Forest, wooded research park, on paved
roads or trails.
Spektoret al. (1988b)
        15-min peak O3 measured during runs averaged
        47 ppb (range:  4 to 135 ppb).  Ambient T
        averaged 29.4 °C (range: 18.0 to 37.8  °C).
        RH averaged 62.6% (range: 37.0 to 88.0%).
        Levels of other pollutants were low, median values
        were SO2, 3 ppb; NO2, 6 ppb; FP, 10 jig/m3.
        Median of subject-specific correlations of O3 and
        RH correlated was -0.42.
Effects of O3 on lung function change during
running outdoors was examined in
24 conditioned, recreational runners (6 women,
18 men, ages 29-47) at a track 30 mi southeast
of Houston, TX, from May to October, 1981.
All runs were 3 mi in length, and each subject
performed at a near-constant heart rate for the
duration of the study. An average of 28 runs
completed by each subject during the study.
Spirometry carried out before and after each run,
with analysis of FVC, FEVj, FEF2j_75^, and
FEFfj 2.j 2L- Chang6 in each lung function
variable was regressed, for each subject, on
15-min maximum O3 measured during the run.
The mean slope across subjects was tested for
significance.  The regression was repeated with
T and RH in the model.
Mean slope of FEVj on O3 alone was -
0.4 mL/ppb (p = 0.03).  In regressions that
included T and RH, the 05 slope dropped to
-0.07 (not significant). Although T reached
high levels during study, a substantial direct
effect of T or RH on lung function, relative
to that of O3, seems unlikely. A possible
potentiating role  of high T and RH on VE,
and corresponding O3 dose, cannot be ruled
out.  Lung function effect observed in simple
O3 model seem likely to be a valid reflection
of O3 effects under varying environmental
conditions.
Selwyn et al. (1985)
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                   TABLE 7-17 (cont'd).  ACUTE EFFECTS OF PHOTOCHEMICAL OXIDANT POLLUTION:
                                               LUNG FUNCTION  IN EXERCISING SUBJECTS3
        Pollutants/ Environmental Variables
             Study Description
          Remits and Comments
                                                                                                                                            Reference
 1-h maximum Oj concentrations during study
 ranged from 50 to 240 fig/m  (25 to 120 ppb).
 The highest 4-h average PM2.5 level was
 70 fig/m , the highest 4-h average sulfate
 concentration was 21 /ig/m , the highest 24-h
 average NO2 concentration was 51 /ig/m .  T data
 were collected but levels were not reported.
The relationship between lung function change
and 03 exposures during outdoor exercise
examined in a population of 83 children
(43 girls, 40 boys; ages not given) in
Wageningen, The Netherlands. Study covered
the period from late May to mid-July, 1989.
Lung function assessed using hand-held peak
flow meters before and after various outdoor
sports training exercises listing approximately
1 h.  Change in PEFR regressed on 03, 03 X
exercise duration, and T for each subject and
distribution of slopes examined. Postexercise
PEFR analyzed in relation to same and previous
day 1-h O3 maximum, and T. Analyses repeated
in subsets of subjects with varying levels of
correlation between 03 and T during their series
of exercise events.
For 55 children with at least four sets of
before and after exercise peak flow
measurements, the mean slope of the PEFR
change on Oj during exercise was 0.035
(± 0.030) mL/s/jtg/m . For 65 subjects with
at least four postexercise measurements, the
mean slope of PEFR on previous hour Oj
was 0.080 (± 0.023), which is statistically
significant, but in the nonplausible direction.
Adjustment for T resulted in negative mean
slopes, but these are difficult to interpret
because of the high statistical  correlation
between same-day O3 and T (r = 0.86).
Exercise events were of low intensity as
compared with chamber studies and with the
Tuxedo runners study (Spektor et al., 1988b).
Significant exposures may have occurred
prior to the exercise period. H   levels were
low (<5 ng/m ) as measured simultaneously
at three other nonurban sites in The
Netherlands. The possibility of a physical
effect of temperature on mini-Wright peak
flow meter measurements was noted by
authors.
Hoekand Brunekreef (1992)
Hoeket al. (1993a)
See glossary of terms and symbols for abbreviations and acronyms.
-------
 1      studies (Selwyn et al., 1985) was discussed in the previous O3 criteria document (U.S.
 2      Environmental Protection Agency, 1986), but is reviewed again here because of its apparent
 3      consistency with the more recent runners study of Spektor et al. (1988b).
 4           Certain design variations across studies are worth noting.  In the Houston runners
 5      study, each of 24 recreational  runners performed spirometry before and after a series of
 6      approximately 28 runs on a track from late spring to early fall (Selwyn et al., 1985).  Each
 7      run was  3 mi long and each subject attempted to maintain a similar heart rate across all runs.
 8      Minute ventilation was not  assessed. In the adult runner/walker study carried out in Tuxedo,
 9      NY, subjects were allowed to  choose their own exercise level and duration, but again were
10      encouraged to maintain a steady heart rate for the duration of the study (Spektor et al.,
11      1988b).  Minute ventilation of each subject while running was estimated by measurement of
12      VE during a treadmill test that achieved a heart rate typical of that subject's experience while
13      running.
14           In contrast to these two studies, the study in Wageningen, The Netherlands, involved
15      lower and more variable exercise levels, without any specific attempt to control exercise
16      intensity (Hoek and Brunekreef, 1992;  Hoek et al., 1993a).  Here, subjects engaged in sports
17      training and skills development activities that were characterized by the investigators as low
18      to moderate in intensity.  Lung function change after exercise was assessed using peak flow
19      meters.
20           Although the designs varied somewhat, O3 exposure levels were similar in the three
21      studies:  in  Houston,  15-min peaks while running varied from 4 to 135 ppb; in Tuxedo,  1-h
22      O3 levels ranged from 21 to 124 ppb; in Wageningen,  1-h maxima on study days ranged
23      from 30  to  120 ppb.
24           The two studies involving fairly intense exercise yielded significant mean slopes of
25      AFEVj (i.e., FEVj after exercise minus FEVj before exercise) regressed on O3 levels
26      measured during exercise, whereas the Wageningen study did not. The mean slope observed
27      in the Tuxedo study across  all  subjects  was -1.35 mL/ppb (±  0.35), but was reduced to
28      -0.55 (± 0.45) in the group of 10 runners who achieved the highest  VE values during
29      exercise.  The mean slope reported from the Houston study was similar to the latter number:
30      -0.4 mL/ppb (± 0.16).  The  large effect level observed in the Tuxedo study led Spektor
31      et al. (1988b) to speculate that 65 effects may have been potentiated by other pollutants such

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 1      as acid aerosols; however, this phenomenon was not demonstrated analytically from the
 2      available acid monitoring data.  In the Houston study, the O3 effect became small and
 3      nonsignificant when temperature and relative humidity were added to the model.  In spite of
 4      this observation, and given the available knowledge base on the independent effects of
 5      O3 and temperature on lung function, it seems reasonable to interpret both of these well-
 6      conducted studies as demonstrating acute effects of O3 on lung function with moderate to
 7      heavy exercise.   The negative findings of the Wageningen study are  difficult to interpret, but
 8      may be related to the low exercise intensities achieved,  exposures (and, perhaps,  associated
 9      O3 tolerance) that occurred prior to the exercise period  under study,  or some subtle physical
10      effect of ambient temperature on the peak flow measurements.
11
12      Lung Function  in Daily Lafe Studies
13           This set of studies is characterized by the assessment of lung function, respiratory
14      symptoms, and environmental factor associations in the  course  of people's daily lives.  This
15      section discusses only the lung function data from these studies. For logistical reasons,
16      studies of this kind have usually involved either spirometry conducted at regular intervals
17      (every 1 to 3 weeks) in schools (Kinney et al., 1989; Castillejos et al.,  1992; Hoek et al.,
18      1993b), or self-administered peak flow measurements in subjects of various ages  over various
19      periods (Vedal et al.,  1987; Krzyzanowski et al., 1989). Although daily life studies have the
20      worthwhile goal of characterizing air pollution effects on respiratory health in the real world,
21      they may suffer from significant exposure assessment uncertainties owing to the use of
22      outdoor O3 monitoring, the incomplete and variable penetration of O3 indoors,  and the
23      preponderance of time spent indoors by study subjects.  This problem is probably less severe
24      for the studies involving schoolchildren, who often spend substantial time outdoors after
25      school, when O3 levels may be elevated.  Indeed, three of the  school-based studies have
26      found statistically significant associations between lung function and  previous-day O3 levels
27      (Castillejos et al., 1992; Kinney et al., 1989; Hoek et al., 1993b).  Another difficulty in
28      interpreting the results of these studies is the possible role of seasonal factors (e.g., pollens,
29      epidemics of respiratory infection, changes in activity patterns) as potential confounders of
30      the  analyses.
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 1           In addition to these general limitations inherent in the study design,  several of the
 2      studies summarized in Table 7-18 have other problems that limit their utility for assessing
 3      O3 effects on lung function.  The study of Vedal and colleagues (1987), though well
 4      conducted, took place from September through May, a period when O3 levels are generally
 5      low and other potential respiratory insults may dominate.  The statistical significance of
 6      results from  a study carried out in Tucson, AZ, is difficult to interpret because of the
 7      multiple statistical tests performed (Krzyzanowski et al., 1989).
 8           The remaining studies, though subject to the general  criticisms noted previously,
 9      provide suggestive evidence that ambient O3 may play a role in short-term lung function
10      declines among children engaged in their normal daily routines (Kinney et al.,  1989;
11      Castillejos et al., 1992; Hoek et al.,  1993b).  The Mexico City study of Castillejos and
12      colleagues (1992) is especially  noteworthy because of the novel observation of FEVj and
13      FEF25_75% decrements that were strongly related to O^ levels averaged  over 24 to
14      168 h previous to spirometry, but not to previous-hour O3 levels.  The  strength of these
15      associations (measured by the ratio of the regression slope to its standard  error) increased
16      steadily as averaging time increased.  As noted by the authors, these results may reflect an
17      inflammatory response in the airways rather than the well-known acute physiological
18      response.  Ozone levels observed throughout this 6-mo study were high by U.S. standards;
19      1-h average O3 concentrations in the hour preceding lung function measurements ranged from
20      14 to 287 ppb, with a mean of 99 ppb.
21
22      Panel Studies of Symptom Prevalence
23           Many field and epidemiological studies reviewed both in the last criteria document
24      (U.S.  Environmental Protection Agency,  1986) and in the  previous section of this document
25      reported results that indicated associations between ambient oxidant exposures and various
26      measures  of respiratory effects (e.g., irritative respiratory  symptoms and acute pulmonary
27      function decrements)  in children and adults.  The aggregation of individual studies provides
28      reasonably good evidence for an association between  ambient photochemical oxidants and
29      acute respiratory effects and a database that is generally coherent,  consistent, and biologically
30      plausible.  In addition, other studies of irritative symptoms in children and adults were also
31      reported in the 1986 document.  For example, Hammer et al.  (1974) reported qualitative

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                         TABLE 7-18.  ACUTE EFFECTS OF PHOTOCHEMICAL OXTOANT POLLUTION:
                         DAILY LIFE STUDDZS OF LUNG FUNCTION AND RESPDXATORY SYMPTOMS3
PolIutants/EnvironmentHl Variables
                                                            Study Description
                                                                                                          Results and Comments
                                                                                                                                                      Reference
Max 03 (1-h) concentrations ranged from 3 to
63 ppb.  Other ambient pollutants measured_were
N02, TSP, IP, RSP. FP, SC>2, and FP SQf.
                                        Lung function measured by spirometry for
                                        154 children ages 10-12 years (90 males,
                                        64 females) in Kingston and Harriman, TN
                                        Spirometry done between 10 a.m. and 1 p.m. on
                                        up to 6 days at least 1 week apart during February
                                        to April, 1981. Child-specific linear regression
                                        models of FVC, FEV0 75, MMEF, and V75% fit
                                        on 1-h O3 max. and 24-h FP and FP SO£".
                                        Means ± SD of distributions of estimated child-
                                        specific slopes computed and tested for
                                        significance by <-test.
Significantly negative mean slopes on 03 for all lung
function variables. For example, mean slope of
FEV0 75 on O3 was -0.99 mL/ppb (±0.36).
Among regressions on FP and FP 804 , only one
statistically significant mean slope (i.e., positive mean
slope of MMEF on FP). Results insensitive to outlier
audits and inconclusive for sensitivity variation.
Association between fitted slopes and individual
characteristics not significant. Low O3 levels raise
plausibility questions.
Kinney (1986)
Kinney et al. (1989)b
1-h average O3 concentrations in hour preceding
spirometry ranged from 14 to 287 ppb, with mean
of 99 ppb. No other pollutants measured.
T ranged from 3  9 to 27.8 °C.  RH ranged from
18.9 to 92.3%
                                        Effects of O3 on lung function examined during
                                        regular school hours in a group of 148 children
                                        (65 girls, 83 boys; ages 7-9) from three  schools in
                                        Mexico City.  Spirometry and symptom data
                                        (cough/phlegm) collected between 0800 and
                                        1400 houri every 2 weeks over the period January
                                        through June,  1988. To account for lung growth
                                        over the study period, residuals from lung
                                        function prediction equations were used in
                                        analyses. Analyses limited to 143 subjects with at
                                        least seven valid measurements.  Schools were not
                                        air conditioned, and windows were usually open.
                                        Schools and subject residences all within 5 km of
                                        O3 monitoring site. Associations between O3 and
                                        lung function (FVC, FEVj, and fEF2S-75lt)
                                        examined by computing the weighted mean of
                                        subject-specific regression slopes relating these
                                        variables.  Various O3 averaging times (from  1 h
                                        to 168 h) were tested.  After analyzing population
                                        as a whole, regressions were repeated in subject
                                        subsets defined by sex, report of chronic
                                        symptoms, and maternal smoking. Overall
                                        regressions repeated with T and RH  in model  with
Only FVC had a statistically significant negative mean
slope in relation to previous hour O3 concentration
(-0.059 + 0.23 mL/ppb). This slope is
approximately 1 order of magnitude lower than those
observed in some camp studies. Both FEVj and
FEF25_75% had significant negative associations with
O3 averaged over the previous 24, 48, and 168 h.
For example, the mean slope of FEVj on 48-h
average O3 was -0.592 + 0.109 mL/ppb. The
authors speculated that the FVC result reflects the
acute, reversible effects of O3 on one's ability to take
a deep breath, whereas the FEVj  and FEF2S-75%
observations may  reflect inflammatory effects of more
prolonged O3 exposures. It should be noted that both
FVC and  FEVj had significant negative slopes on 1-h
maximum O3 measured in the previous 24 h.
Adjustment for T  and RH diminished somewhat the
associations between lung function and O3.
Associations between lung function decrements and
O3 exposure often appeared larger in children with
chronic respiratory symptoms than in those without,
and in children of mothers who were current smokers;
however,  these results were not statistically
confirmed.
Castillejos et al. (1992)
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o
8
I
\o
VO
U)
                   TABLE 7-18 (cont'd).  ACUTE  EFFECTS OF PHOTOCHEMICAL  OXTOANT POLLUTION:
                        DAILY LIFE STUDIES OF LUNG FUNCTION AND RESPIRATORY  SYMPTOMS3
             Pollutants/Environmental Variables
                                                                 Study Description
                                                                                                    Results and Comments,
                                                                                                                                               Reference
Daily maximum 1-h Oj concentrations on days
prior to lung function testing ranged from 7 to
206 fjig/m  (3.5 to 103 pph). Levels of other
pollutants measured (SO2, NO2, PMjrj, and
aerosol H  ) were reported to he low during study.
Ambient T (range:  5 to 31  °C) and some pollen
data were also collected
Associations between morning lung function and
previous day Og examined during school in
533 children (ages 7-11) from seven schools in
three towns in the Netherlands. Towns were
selected without local pollution sources and with
low levels of pollutants other than Oj.  Study
spanned the period from March through July,
with lung function measurements collected every
2-3 weeks. An overall time trend was fit to the
lung function data to account for lung growth
Data on FVC, FEV^  PEFR, and FEF25_755{
analyzed in relation to previous-day 1-h
maximum O-j concentrations using subject-
specific linear regressions followed by analysis
of mean slopes.  Intersubject variations in
responsiveness to 03 were tested via an F-test.
The influence of chronic respiratory symptoms
and other subject characteristics (e.g., age, sex)
on O3 responsiveness was examined. Models
that included other pollutants were also
considered.
Negative, usually statistically significant
mean slopes seen for lung function regressed
on previous-day  1-h maximum 03 for the
seven individual schools. Over all
533 subjects, mean regression slopes for FVC
and FEVj were -0.20 ± 0.05 and -0.21 ±
0.04 mL//ig/m ,  respectively; and for PEFR
and FEF25_7j% were -0.72 + 0.22 and
-0 45 + 0.12 mL/s/jig/m , respectively.
These coefficients may be doubled to convert
to slopes in terms of ppb. The authors report
that adding SO2,  NO2, or PM10 did not
materially change the Oj slopes. There was
evidence for inter-subject variation in Oj
responsiveness, but this variation was not
statistically related to available subject
characteristics data. T data not included in
models, perhaps due to high correlation with
Oj. The lung function/O3 relationships noted
above are qualitatively similar to those
reported in the 1988 Fairview Lake camp
study and the Mexico City school children's
study
Hoek and Brunekreef (1992)
Hoek et al. (1993b)
-------
I
«
U)
                          TABLE 7-18 (cont'd).   ACUTE EFFECTS OF PHOTOCHEMICAL OXIDANT  POLLUTION:
                                DAILY LIFE STUDIES OF LUNG FUNCTION AND RESPIRATORY SYMPTOMS3
              Pollutants/Environmental Variables
              Study Description
                                                                                                             Results and Comments
                                                       Reference
-J

NJ
       1-h maximum O3 concentrations on PEFR
       measurement days ranged from 20 to 103 ppb.
       No other pollutants assessed, but ambient T data
       included.
Relationship between daily peak flow
measurements and ambient O3 concentrations in a
population sample of 732 subjects (both adults
and children) over 2-week periods during normal
daily activities in Tucson, AZ.  Peak flow
assessed using hand-held peak flow meters up to
four times per day.  PEFR measurements on
initial 2 days for each subject dropped to avoid
possible learning effects, leaving a series of up to
12 measurement days per subject. Population
pooled regression slopes computed for PEFR on
1-h and 8-h avenge Oj for children (ages <  15)
and adults (ages > 15), controlling for residual
auto-correlations.  Outcome measures included
PEFR diurnal variability and afternoon PEFR
levels.  Besides O3, potential explanatory
variables included T, average time outdoors, acute
respiratory infections, asthma, and environmental
tobacco smoke exposure.
Significant positive associations observed
between 03 concentrations and PEFR diurnal
variability; the effect magnitude was greatest
in asthmatic subjects.  In children only, noon
PEFR was suppressed on days with higher O3
levels.  The uncertain relationship between
central site O3 levels and personal exposures
in this southwestern community was not
addressed. Although the statistical models
employed were appropriate and well chosen, it
appears that a substantial amount of
exploratory data analysis was performed prior
to selection of results to present in paper,
leading to uncertainties regarding the statistical
validity of the hypothesis tests presented.
Krzyzanowski et al. (1989)
       Means and range of max. daily 1-h values:
       O3 mean = 32.4 /ig/m .range = 0-129 ng/m ;
       SO2 mean  =  51.2 ^g/m .  range = 18-176 jug/m ;
       NC>2 mean = 40.5 fig/m , range =  12-79 fig/m ;
       CoH mean = 0.38 CoH units, range =
       0.1-1.3 CoH units; T mean = 1.3 °C, range =
       -22° to +22 °C.
Follow-up study (September, 1980-April, 1981)
of pollutant-respiratory symptom relationships in
subsets of children from 1979 Chestnut Ridge
cross-sectional study of > 4,000 elementary
school children. Subsamples selected from six
schools in study area with consistently higher
levels of air pollution during previous 4 years.
Subsamples (three) stratified by reported
symptoms. One or more of following measures
taken for 144 children: diaries, symptom
questionnaire, spirometry. Telephone follow-up
each 2 weeks on diaries, spirometry done at
school, pollutants (including O3) measured at one
monitor (data from 17 monitors for SO2 generally
reflected in data at single monitor).  Diary panel
study covered 8 mo; successive PEFR spirometry
studies of 9 weeks each done in respective groups
of the three Subsamples.
Relationships of maximum hourly SO2>
O3, and CoH and minimum.  T for each 24-h
period to daily upper and lower respiratory
illness, wheeze, and PEFR were evaluated
using multiple regression models adjusted for
illness occurrence or levels of PEFR on
preceding day.  No air pollutant was strongly
associated with respiratory illness or with
PEFR.  Authors concluded that this study can
best be interpreted as showing no acute effects
of studied pollutants on respiratory symptoms
or PEFR in children at levels lower than the
current NAAQS, but also noted that
conclusion must be tempered by relatively low
levels of pollutants encountered and possibility
of exposure misclassiftcation.
Vedal et al. (1987)
      aSee glossary of terms and symbols for abbreviations and acronyms.
       Cited in U.S  Environmental Protection Agency (1992).
-------
 1      associations between ambient oxidant levels and symptoms such as eye and throat irritation,
 2      chest discomfort, cough, and headache at total oxidant levels greater than 0.15 ppm in young
 3      adults (nursing students).  Wayne et al. (1967) reported a high correlation (R2 = 0.89)
 4      between ambient total oxidant levels (1 h prior to competition) and impaired exercise
 5      performance (running time) in high school students during cross-country  track meets in
 6      Los Angeles, CA.  Symptoms were not measured, but Wayne speculated that chest
 7      discomfort from oxidant inhalation impaired exercise performance.  Although results such as
 8      these are consistent with evidence from controlled human exposure studies, precise
 9      characterization of ambient pollutants and environmental conditions and rigorous statistical
10      analyses were lacking in the studies.  Thus, the primarily qualitative data from these and
11      other studies were not satisfactory to provide quantitative conclusions about the relationship
12      of ambient O3 concentrations and acute respiratory illness.
13           Schwartz (1992), Schwartz and Zeger (1990), and Schwartz et al. (1988) reanalyzed the
14      original diary data of student nurses reported earlier by Hammer et al. (1974) (Table 7-19).
15      The nurses were told that the diaries were part of a prospective study of  viral infections.
16      Logistic regression  models including time series analyses were used to control for
17      autocorrelation effects that are frequently present in time series data.  The reanalysis for
18      daily prevalence rates of symptoms (Schwartz et al., 1988) confirmed that ambient oxidants
19      were significantly associated with cough and eye discomfort.  However,  earlier reported
20      associations between oxidants and headache or chest discomfort were not confirmed. Cough
21      was the one symptom that showed an apparent threshold near 0.20 ppm total oxidants, which
22      approximates the threshold value reported by Hammer et al.  (1974). Further reanalysis  of
23      the diary data (Hammer et al., 1974) by Schwartz (1992) and by Schwartz  and Zeger (1990)
24      for the effects of air pollutants on the risk of new episodes of respiratory and other
25      symptoms and on their durations revealed interesting findings.  The mean plus or minus
26      standard deviation (SD) level of oxidants was 0.102 ± 0.074 ppm.  In logistic regression
27      models, an increase in oxidant concentration by one SD (0.074 ppm) was associated with a
28      17%  increased risk of chest discomfort and a 20% increased risk of eye irritation.  These
29      associations were highly  significant (p <  0.001).  In addition, photochemical oxidants were
30      significantly (p < 0.0001) associated with the duration of episodes of cough, phlegm, and
31      sore throat.

        December 1993                          7-143      DRAFT-DO NOT QUOTE OR  CITE
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                     TABLE 7-19.  ACUTE EFFECTS OF PHOTOCHEMICAL OXIDANT POLLUTION:
                                                     SYMPTOM PREVALENCE3
Pollutants and Environmental Variables
         Study Description
       Results and Comments
                                                                                                                     Reference
Total oxidant, CO, SO2, NO, and NO2
measured; total oxidant concentrations
reached episodic levels (maximum
1-h/day < 0.4-0.5 ppm); mean daily
temperature was 71.8 °C.
Reanalysis of daily diary study of
student nurses working and living at
schools in Los Angeles (see U.S.
Environmental Protection Agency,
1986, for details of Hammer et al.,
1974). This series of papers
reexamines the nurses' data using
logistic regression models and time-
series methods to account for serial
correlation (autocorrelation) of
symptoms on successive days.  The
effects of total oxidants on daily
prevalence rates of symptoms, risks
of developing new symptoms
(episodes), and duration of episodes
were analyzed.
 Associations found between total
 oxidants and prevalence of cough and
 eye irritation, confirming part of
 findings of original study.
 Association with cough only at
 oxidant concentrations above
 approximately 0.20 ppm.  Previously
 reported associations between
 oxidants and chest discomfort and
 headache (Hammer et al., 1974) not
 confirmed. Oxidants associated with
 increased risk (incidence) of chest
 discomfort and eye irritation and
 duration of episodes of cough,
 phlegm, and sore throat.  Duration of
 symptoms showed dose-response
 relationships even below current
 NAAQS for O$.  Findings suggest
 different effects of oxidants on
 symptom characteristics.  Lack of
 daily paniculate measurements, small
number of subjects, and
heterogeneous individual responses
 restrict quantitative interpretation of
results.  Lung function was not
measured.
Schwartz (1992)
Schwartz and Zeger (1990)
Schwartz et al. (1988)
-------
                 TABLE 7-19 (cont'd).  ACUTE EFFECTS OF PHOTOCHEMICAL OXTOANT POLLUTION:
                                                     SYMPTOM PREVALENCE3
Pollutants and Environmental Variables
        Study Description
          Results and Comments
          Reference
Daily O3, NO2, SO2, and CH and every
sixth-day sulfates measured at one site
(Azusa).  1-h daily maximum O3
0.1 ppm, 7-h average O3 0.07 ppm;
sulfates 8.43 /tg/m ; maximum
temperature was 22.4 °C.
Reanalysis of daily diaries
completed during 181-day survey
period (September 1978 to March
1979) by 756 children and
572 adults (Krupnick et al., 1990)
and 321  nonsmoking adults (Ostro
et al., 1993) living in Glendora,
Covina,  or Azusa, CA  (see Flesh
et al., 1982 for details).  Presence
or absence of 19 (upper and lower)
respiratory and 2 nonrespiratory
symptoms recorded daily. Presence
or absence of symptoms analyzed in
a pooled cross-sectional time-series
model.   Nonpollution factors,
including sex, gas  stove use, day of
study, and a chronic disease
indicator were included in final
regression models  used to measure
effects of ambient  air pollution.
Logistic regression analyses for
entire sample to determine effect of
each pollutant on health endpoints.
Lagged effects of each  pollutant and
effects hi individuals (n = 74)
without air conditioners and those
with preexisting respiratory infection
were analyzed.
Logistic regression model indicated
significant associations between incidence
of lower respiratory symptoms and healthy
nonsmoking adults (but not among
smokers, children, or patients with chronic
respiratory disease); 1-h daily maximum O3
levels OR = 1.22, 95% CI of 1.11-1.34,
for a 0.1 ppm change; 7-h average O3 level
(OR =  1.32, 95% CI of 1.14-1.52); and
ambient sulfates (OR = 1.30, 95% CI of
1.09-1.54, for a 10 /tg/m3 change).  CH
was significantly related to daily symptoms
in children. Gas stove in the home was
associated with lower respiratory tract
symptoms (OR =  1.23, 95% CI of
1.03-1.47), as were the effects of O^ in
subgroups without residential air
conditioner (OR = 1.24) and with
preexisting respiratory infection
(OR =1.24).  All the above increased risks
were statistically significant (p < 0.05).
Interpretation of results limited by selection
of sample; undersampling of young adults;
aggregation of symptoms of all severity
levels into one measure; possible reporting
bias; and absence  of indoor exposure,
aeroallergen, and  lung function data.
Krupnick et al. (1990)
Ostro et al. (1993)
See glossary of terms and symbols for abbreviations and acronyms.
-------
 1          Krupnick et al. (1990) and Ostro et al. (1993) reanalyzed daily health data from over
 2     5,000 children and adults living in the Los Angeles area during a 6-mo period (September
 3     1978 to March 1979) (Table 7-19). The original study was reported by Flesh et al. (1982)  of
 4     the U.S. Environmental Protection Agency. The presence or absence  of daily respiratory
 5     symptoms associated with daily exposure to ambient O3 and other air pollutants was analyzed
 6     in a pooled cross-sectional time-series model.  Krupnick et al. (1990) reported statistically
 7     significant effects of O3 levels on daily reported respiratory symptoms in healthy nonsmoking
 8     adults, but not among smokers, children, and patients with chronic respiratory disease.
 9     Ostro et al.  (1993) evaluated the daily reports of 321 nonsmoking adults and, using a logistic
10     regression model,  found a statistically significant association between the incidence of lower
11     respiratory tract symptoms and 1-h daily maximum and 7-h average O3 levels (22 and 32%
12     increased risk, respectively, with 0.10 ppm increase in O3) and ambient sulfates (30%
13     increased risk with 10-^g/m3 change). The lower  respiratory tract effects of O3 were greater
14     in the subgroups with gas stoves, without residential air conditioners, and with preexisting
15     respiratory infection.  Interpretation of the results is limited by the selection of the sample
16     for analysis;  undersampling of young adults; aggregation of symptoms of all  severity levels
17     into one measure;  possible reporting bias; and absence of indoor exposure, outdoor
18     aeroallergen, and lung function data.
19          The results from the above panel studies suggest a modest but biologically plausible
20     relationship between short-term exposure to ambient oxidants/O3 and respiratory symptoms.
21     The interpretation of these recent reanalyses is limited  by several factors.  Heterogeneous
22     individual responses occur and analyses of grouped data may possibly miss susceptible
23     subgroups.  The lack of specific measurements of  O3 and/or other pollutants (especially
24     particulates) and of personal exposure or risk variables weaken the assessment of confounders
25     and effect modifiers.  In addition, the overall data analysis pertains to small and very
26     selected samples that have uncertain representativeness to the general population.
27
28     Aggravation of Existing Respiratory Diseases
29           Prior epidemiological data on the effects of ambient O3 levels in subjects with existing
30      respiratory disease have been  difficult to interpret  due  to methodological limitations (U.S.
31      Environmental Protection Agency, 1986).  Exacerbation  of asthma and other health endpoints

        December 1993                           7-146      DRAFT-DO NOT QUOTE  OR CITE
-------
 1      has been subsequently evaluated, and more recent studies have observed possible increases in
 2      symptom aggravation or changes in lung function of asthmatic subjects in relation to
 3      increased O3 or total oxidant levels,  as well as interactions between O3 concentrations and
 4      temperature. However, no consistent pattern of findings for aggravation of symptoms or
 5      lung function changes has been reported for patients with other types of chronic lung disease.
 6      Some of the major issues in interpreting results from studies of respiratory exacerbations
 7      have been inadequate sample size and characterization of the study subjects, lack of
 8      information on the possible effects of medications, the absence of records for all days on
 9      which symptoms could have occurred, inadequate interpretation of the clinical significance of
10      measured changes, the role of confounders and effect modifiers (e.g., temperature, humidity,
11      particulates,  and aeroallergens), and  personal or group characterization of indoor-outdoor
12      exposures.  For example, Whittemore and Korn (1980) and Holguin et al.  (1985) found small
13      increases in the probability of asthma attacks associated  with previous attacks, decreased
14      temperature, and incremental increases in oxidant and O3 concentrations. Lebowitz et al.
15      (1982, 1983, 1985) and Lebowitz (1984) reported effects in asthmatics,  such as decreased
16      PEFR and increased respiratory symptoms, that were related to the interaction of O3  and
17      temperature. None of these studies adequately assessed  possible effect modification by other
18      pollutants, particularly inhalable particulates, which may have independent effects.
19          Epidemiological studies published since the 1986 criteria document (U.S.
20      Environmental Protection Agency, 1986) have attempted to control for many methodological
21      issues  (e.g.,  with [1] better estimates of exposure to pollutants [as well as O3] and
22      environmental variables that can confound or modify responses, [2] serial measurements  of
23      pulmonary function for determining correlations with pollutants and other environmental
24      variables, [3] better biomedical characterization of cohorts, and [4] more robust analytical
25      approaches that control for autocorrelation of environmental variables and health responses).
26      Recent studies have generally provided further evidence  that supports a relationship between
27      ambient O3/oxidant concentrations and respiratory morbidity in asthmatic subjects
28      (Table 7-20).
29          Gong (1987) studied the relationship between air quality and respiratory status of
30      83 asthmatic subjects living in a high-oxidant area of Los Angeles County.  The study
31      covered February to December 1983, but data analyses were limited to a 230-day period

        December  1993                          7-147      DRAFT-DO NOT QUOTE OR CITE
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                             TABLE 7-20.  AGGRAVATION OF EXISTING RESPIRATORY DISEASES BY
                                                PHOTOCHEMICAL OXIDANT POLLUTION8
     Pollutants and Environmental Variables
                                                  Study Description
                                                  Results and Comments
                                                   Reference
oo
Air pollutant measurements for April to
November 1983 used in statistical analyses.
Daily maximum of NOX, SO2, CO, THC
less than California standards or NAAQS.
SO4= >25 fig/m on 4 days;
TSP> 100 /ig/m3 on 78% of days with
data. Daily maximum 1-h average 03
concentrations (from continuous
monitoring) = 0.01-0.11 ppm on
103 days, 0.12-0.19  ppm on 65 days,
0.2-O.34 ppm on 60  days, and
0.35-0.38 ppm on 3  days.  Outdoor
aeroallergens sampled with  Roto-Rod:
spores, pollens,  grasses, molds,
miscellaneous debris; all generally low
except for group of common molds (rusts,
smuts, mushroom) present in
thousands/cm on sampler.  Mean (±SD)
daily temperature at  1 p.m. during
200 days:  26 ± 11  °C, range 13-41 °C;
128 days with £ 24  °C.
Effects of pollutants and other
environmental variables on respiratory
symptoms and PEFR evaluated in 11-mo
population study of asthmatics living in
high-O3 area (Glendora) of Los Angeles
County, CA.  Detailed questionnaires
given at outset on medical/occupational
histories and personal factors, including
general activity patterns; psychological
testing (Asthma Symptom Checklist,
State-Trait Anxiety  Inventory, etc.) also
given, once during good air period and
once during smoggy period.  Lung
function (spirometry) and bronchodilator
responses measured at outset in all
subjects.  Daily diaries (checked
2 X/week), mini-Wright peak flow
meters, and Nebulizer Chronolog
attached to metered-dose inhaler used to
record symptoms, day and night PEFR,
and medication use, respectively.
Multiple regression  analyses  for overall
group; then subsets  (two groups of
"responders") analyzed separately  and
compared with rest  of cohort.
Eight of 91 subjects completing study (of
109 recruited) showed no variability in asthma
status during the 230-day study.  Respiratory
status of final study population (n = 83 with
generally mild or stable asthma) as a whole not
related,  either clinically or statistically, to
maximum 1-h average O3 on Days t, t-1, t-2,
t-3 for any respiratory variable even when
adjusting for medication use, symptoms,  and
PEFR on Day t-1.  Subset analyses showed
association of O3 with symptoms and with day and
night  PEFR hi subjects in top quartile for
respiratory measures, but association did not
follow a consistent  relationship with ambient
O3 concentrations.   VE levels during outdoor time
not estimated.  Outcomes not related to time
outdoors vs.  indoors or to outdoor time on "clean"
versus "smoggy" days.  Subsets ("responders")
differed from rest of cohort  mainly hi scores of
Asthma  Symptom Checklist  for factors
representing fatigue, hyperventilation, and rapid
breathing, but there was no difference in
responders between clean and and smoggy
periods.  Aeroallergens from maple, oak, beech,
and elm trees showed significant (and clinically
relevant) relationships to respiratory variables.
Exposure assessment limited by outdoors-only
monitoring and lack of time-activity data.
                                                                                                                               Gong (1987)
                                                                                                                               Gong et al. (1985)
-------
g
U)
                    TABLE 7-20 (cont'd). AGGRAVATION OF EXISTING RESPIRATORY DISEASES BY
                                           PHOTOCHEMICAL OXTOANT POLLUTION8
     Pollutants and Environmental Variables
                                                 Study Description
                                                  Results and Comments
                                                   Reference
Hourly outdoor O3, CO, NO2, and TSP
measured from three stations.  Hourly
maxima used for O3 and CO; daily CO
and NO2 derived as weighted measures for
each cluster sampling site and daily values
were used.  Sample of homes monitored
inside and outside for particulates and
gases and evaluated for housing
characteristics (e.g.,  gas stove  usage).
Meteorological variables measured daily.
Temperature data not reported.
Effects of outdoor and indoor air
pollutants and aeroallergens evaluated in
a 2-year study of 22 subjects with
asthma, 33 with airway obstructive
disease, 30 atopies, and 14 normals
living in a dry arid environment
(Tucson,  Arizona). Subjects part of a
community population sample of
117 families  (see U.S. Environmental
Protection Agency, 1986, for details of
Lebowitzetal., 1982, 1983, 1985;
Lebowitz, 1984) and had well-
characterized symptoms, medication use,
lung function, methacholine response (in
a subsample), and immunological status.
Daily diaries (acute symptoms,
medication use, and doctors' visits) and
daily PEFR (2x/day) performed for
3 mo, 2-4 X /study period.  Duration
spent outdoors recorded.  Spectral time
series analyses used to evaluate each
respiratory response variable for
periodic tendencies and covariance
(dependent and independent) functions as
processes in time in the different
groups.
Asthmatics had greatest number of respiratory
complaints, which were related to the presence of
gas stoves, active smoking, humidity, and
temperature.  Ozone was associated with peak
flow (late  spring and with temperature), wheeze
(Day t-3 with humidity),  and productive cough
(Day t-2).  O3 (Day t-3) was related to productive
cough during the summer in allergic subjects.
Outdoor gases and meteorological variables
significantly related to symptoms and PEFR both
independently and  as effect modifiers.
No significant O3 effect in patients with
obstructive disease or in normals. Small number
of subjects and study days and lack of indoor NO2
and PM10  measurements, measured pollutant
values, and effect estimates limit quantitative
interpretation of study.
                                                                                                                              Lebowitz et al.
                                                                                                                              (1987)
-------
                            TABLE 7-20 (cont'd).  AGGRAVATION OF EXISTING RESPIRATORY
                                  DISEASES BY PHOTOCHEMICAL  OXTOANT POLLUTION3
Pollutants and Environmental Variables
          Study Description
             Results and Comments
                                                                                                                                  Reference
-J

o
Outdoor O3 levels measured hourly by
three stations and maximum 1-h and 8-h
average values were used to represent
O3 levels for all subjects on a given day.
For each day of the study, the mean of the
maximum 8-h O3 average for the
4 preceding days was calculated to be an
index of cumulative exposure.  PMjQwas
measured daily at one station. Mean
± SD of maximum 1-h O3  concentrations
was 0.055 ± 0.014 ppm (range:  0.015-
0.092 ppm), moving average maximum 8-h
O3 levels were 0.046 + 0.013 ppm (0.09-
0.082 ppm). Maximum daily outdoor
temperature was 87 °F (30  °C) per person-
day, maximum PM10 was 187 j«g/m
(mean 42 /ig/m ).
Temporal effect of ambient O$
concentration on PEFR during 30-mo
study period in 287 children (13% with
physician-diagnosed asthmatics) and
523 nonsmoking adults (9% asthmatics)
in the Tucson community population
sample.  Mini-Wright peak flow meters
used four or fewer times/day but for
only 2-week periods and only one meter
was assigned/household.  Children's
tests were supervised by adult and initial
2 days of observation were eliminated
from analysis.  Symptoms from daily
diaries were also used in analysis.
Random-effects longitudinal model was
used for analyses to account for
autocorrelation of PEFR values.
Multifactorial ANCOVA was used to
analyze day-to-day changes in daily
average PEFR and symptom prevalence
rates (the dependent variables) in
relation to 8-h O3 values on the same
day and previous days (lags of 0 and 1).
Analyzed PEFR data limited to at least
12 measurements for at least 6 days in 78 % of
children and 74% of adults.  Noon PEFR in
nonasthmatic and asthmatic children was lower
with higher 1-h maximum O3 levels:
-11.9 L/min/0.1 ppm €5 (p < 0.05) and
-31.0 L/min/0.1 ppm 0$ (p < 0.1), respectively.
Effect of 8-h O3 mean  on evening PEFR only seen
in asthmatic  children, possibly reflecting a
cumulative Oj  response during course of day.
Among adults, evening PEFR was decreased in
asthmatics who spent more time outdoors on days
with higher Oj concentrations (time X
concentration effect). The ANCOVA model
showed significant interactive effects of Oj  X
temperature  x  PMjQ on daily average PEFR.
Daily rates of allergic-irritant symptoms increased
with the maximum 8-h O3 average (> 0.056 ppm)
on the previous day and increased more with
interactions of  O3  X temperature X PM10.
Missing PEFR data,  possible overestimation of
outdoor O3 exposure, large variability of
responses in  asthmatics, medication use on days
with high O3 levels,  relatively low O3 levels, and
uncertain effects of indoor and outdoor allergens
and respiratory infections limit interpretation.
                                                                                                                              Lebowitz et al.
                                                                                                                              (1991)
                                                                                                                              Krzyzanowski et al.
                                                                                                                              (1992)
-------
p-t   	
>—   Hourly O3, twice daily (9:00 a.m. and
^O   9:00 p.m.) acidic aerosols (sulfates, SO4,
     and H  ), and pollen counts were measured
     on site.  Hourly T, RH, and O3 measured
     from nearby monitors.  In 1991, pollution
     levels increased daily until Day 5, when
     maximum 1-h O3 reached 0.154 ppm and
     daytime H  and sulfate levels were
     245 nm/m  and 26.7 /xg/m , respectively.
     In  1992, air quality was better (e.g., the
     highest daily 1 -h maximum O3 was
     0.063 ppm). Temperature data not
     reported.
                            TABLE 7-20 (cont'd).  AGGRAVATION OF EXISTING RESPIRATORY
                                  DISEASES BY PHOTOCHEMICAL OXTOANT POLLUTION3
Pollutants and Environmental Variables
Study Description
                                                                                             Results and Comments
Reference
                                       Effects of ambient summertime haze air
                                       pollution on asthmatic children
                                       (ages 7-13) attending 1-week asthma
                                       camp in Connecticut River Valley were
                                       evaluated during June 1991  (n = 50)
                                       and 1992 (n = 55). PEFR and
                                       symptoms (2x/day) and number of
                                       as-needed (p.r.n.) inhaled bronchodilator
                                       treatments given by on-site physician
                                       during each study day were recorded.
                                       Correlations between health outcomes
                                       and air pollutants were performed.
                           In 1991, daily total number of p.r.n. treatments    Thurston et al.
                           highly correlated (r > 0.80) with maximum O3,   (1992a,  1993c)
                           804, daytime H , and maximum temperature, but
                           only SO4 (r = 0.97) and H+ (r = 0.985) were
                           significant (p < 0.05) and remained so after
                           temperature was included in the analysis.  Daily
                           pollen counts were not associated with as-needed
                           treatments.  Afternoon chest symptoms (cough,
                           phlegm, and wheeze) and changes in morning-
                           afternoon PEFR values (excluding children given
                           medication) were significantly correlated
                           (p < 0.05) with O3 and H+, respectively.
                           Scheduled medications did not apparently provide
                           a protective effect (X2 = 3.25, p = 0.067),
                           although the failure to achieve statistical
                           significance is not unexpected given the small
                           sample size.  In 1992, change in PEFR
                           (magnitude not reported), chest symptoms, and the
                           fewer daily exacerbations (maximum 27 vs. 37 in
                           1991) were not significantly correlated with
                           pollution, pollen, or temperature.  Only sore
                           throat, runny nose, and eye irritation were
                           correlated with pollen counts. Although the data
                           are only hi preliminary form, the 1991 results
                           appear consistent with an effect of summertime
                           haze air pollution on PEFR, chest symptoms, and
                           asthma exacerbations. The 1992 results are
                           consistent with less health effects owing to cleaner
                           ambient conditions. Small number of subjects and
                           study days and lack of results for other pollutants
                           limit the interpretation of the studies.
See glossary of terms and symbols for abbreviations and acronyms.
-------
 1     (April 15 through November 30) because of staggered entry of subjects into the study and the
 2     high frequency of missing or incomplete data encountered in the earlier part of the study
 3     period.  Regression and correlation analyses between 03 and average symptom scores,
 4     asthma medication index (AMI), and day and night PEFR across subjects showed weak,
 5     nonsignificant relationships.  These daily outcome variables were compared across days with
 6     maximum 1-h-average O3 in three ranges:  <0.12 ppm, 0.12 to 0.19 ppm, and >0.20 ppm;
 7     "no statistical or clinical significance was detected."  Individual exposures and activity
 8     patterns  were not estimated in these two analyses. Multiple regression analyses also
 9     indicated the lack of a significant overall relationship between O3 (and their independent
10     variables) and respiratory status, despite the use of lagged variables and the inclusion of
11     other pollutants,  meteorological variables, aeroallergens, and AMI.  Total suspended
12     particulates directly affected PEFR, but the relationship was not consistent in the analysis.
13     Aeroallergens showed significantly negative relationships to respiratory variables, but only
14     the effect of certain molds was considered  clinically relevant.  Temperature and humidity
15     showed no significant effect on the respiratory variables on this study.
16           Although there was no significant overall effect of O3 on respiratory variables in the
17     83 asthmatic subjects, multiple regression analysis of subjects whose O3  coefficients on
18     various days were in the top quartile for dependent variables (respiratory measures) showed
19     significant and consistent effects of O3 on Day t and the previous day  (Day t-1). Multiple
20     regression testing of subsets for associations of  symptom score or day or night PEFR on the
21     same day's O3 and the previous day's value of the same responses showed highly significant
22     O3 coefficients for all three respiratory measures.
23           The clinical significance of responses in symptom scores and day and night peak flow
24     was evaluated for all subjects by individual regression analyses.  No subject had evidence of
25     significant worsening of symptoms attributable to O3 during the study. Adult subjects with
26     high scores in fatigue, hyperventilation, dyspnea, congestion, and rapid breathing in the
27      Asthma Symptom Checklist had more negative  slope coefficients for O3 than subjects with
28      low-to-moderate scores on  the checklist. "Re&ponders" (statistically identified by multiple
29      regression analysis) scored consistently higher in the factors representing fatigue,
30      hyperventilation, and rapid breathing.  The higher scores of these responders,  however,
31      "were apparently not associated with differences in  ambient O3 concentrations since the test

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 1     scores were similar during relatively low (first test) and high (second test) O3 days.  The
 2     significance of the psychological results is unclear at this time."
 3          Lebowitz et al. (1987) performed time series analysis to evaluate daily respiratory
 4     responses to outdoor and indoor air pollutant and aeroallergen exposures in potentially
 5     sensitive adults living in a dry climate (Tucson, Arizona).  Daily symptoms and PEFR were
 6     recorded in well-characterized groups  of asthmatics, allergic subjects, patients with chronic
 7     airways obstruction, and asymptomatic healthy controls (total sample size of 204) over
 8     2 years.  Daily diaries included acute  symptoms, medication use, and doctors' visits.
 9     A sample of homes was evaluated for  environmental characteristics and was monitored
10     indoors and outdoors for gases and particulates,  in addition to stationary outdoor monitors.
11     Asthmatics showed the most respiratory responses.  Outdoor O3 levels were significantly
12     (p < 0.05) related to wheeze, productive cough, and peak flow (late spring) in the asthmatic
13     group.  Statistical interactions between O3 and smoking, presence of a gas stove, maximum
14     temperature, and minimum humidity (R  = 0.49) were found.  The other groups did not
15     demonstrate an O3 effect, except for the atopic group, which had increased summertime
16     productive cough related to O3 levels.  Thus,  these results  indicate an O3 effect on asthmatics
17     and that statistical interactions between O3 and other environmental factors are significantly
18     related to symptoms and peak flow. On the other hand, the results are largely descriptive
19     and qualitative without adequate effect estimators.
20           A subsequent analysis of the same community population sample in Tucson (Lebowitz
21     et al., 1991; Krzyzanowski et al., 1992) evaluated the temporal relationship between PEFR
22     and ambient O3  in 287 children and 523 nonsmoking adults.  During part of the study
23     period, ambient particulates (paniculate matter of mass median aerodynamic diameter 10 jum
24     or less [PM10]) were collected daily at one monitoring station.  A random-effects longitudinal
25     model and multifactorial analysis of covariance were  used for analyses.  During the study
26     period, the maximum ambient O3 concentrations  were relatively low (i.e., the 1-h maximum
27     never exceeded 0.092 ppm).  In children, noon peak  flows were decreased on days when
28     there was a high O3 concentration.  Children with physician-confirmed asthma experienced
29     the greatest decrease in noon peak flow. Evening peak flow was also significantly related to
30     O3 in children, especially asthmatic children,  suggesting a cumulative O3 response during the
31     course of the day.  Among adults, evening peak flows were decreased in asthmatics who

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 1     spent more time outdoors on days when O3 levels were high.  After adjustment for
 2     covariates, significant statistical interactions of 8-h O3 levels with participate matter (PM10)
 3     and temperature on daily PEFR were found.  There was a significant increase in allergic-
 4     irritant symptom rates related to prolonged exposure to O3 (maximum 8-h average on the
 5     previous day and the interactions of O3, temperature, and humidity).  The study had some
 6     methodologic problems (e.g., missing daily PEFR data in many subjects, lack of information
 7     about specific hours spent outdoors, medication usage, and relatively low O3 levels during
 8     the study period). Nonetheless, the data analyses, control of confounders, and overall
 9     exposure assessment strengthen the conclusions of the study (i.e., that the respiratory
10     response to O3  is acute, occurs more often in asthmatics, and increases as temperature and
11     PMjo increase).
12          The  respiratory effects of ambient O3 and other coexisting pollutants were evaluated
13     during a 1-week asthma camp located in the Connecticut River Valley during June of 1991
14     and  1992  (Thurston et al., 1992a, 1993c).  Each child (age 7 to 13 years) participated in the
15     same daily activities all week. Peak flow and symptoms were recorded twice a day, as well
16     as the number of as-needed (p.r.n.) treatments of inhaled bronchodilator administered by an
17     on-site physician during each day (each representing an exacerbation of asthma).  Hourly
18     measurements of O3 and twice daily samples of acidic aerosols (sulfates, SO4, and hydrogen
19     ions, H+) were collected.   The preliminary results (manuscript in preparation) indicate a
20     strong association between the ambient air pollution mix and the occurrence of asthmatic
21     exacerbations in children.  During 1991, pollution levels progressively increased until Day 5,
22     when the  1-h maximum O3 concentration reached 0.154 ppm, and the daytime (9:00 a.m. to
23     9:00 p.m.) H+ and SO4 concentrations were 254 nm/m3 and 26.7 jig/™3, respectively.  The
24     correlations of the daily total number of p.r.n.  treatments required with daily  maximum O3,
25     daytime SO4 and H+, and maximum temperature were all high (r > 0.8), but only SO4
26     (r = 0.97) and H+ (r = 0.98) were significant (p <  0.05) given the small number of days
27     involved.   Afternoon  symptoms (cough, phlegm,  and wheeze) and morning-afternoon change
28     in PEFR  (without medication) were significantly correlated (p < 0.05) with 03 and H  ,
29     respectively.  During 1992, the local air quality was better (e.g., the daily 1-h maximum
30     O3 concentration was only 0.063 ppm).  There were fewer asthmatic exacerbations
31     (maximum of 27 versus 37 in 1991), and they  were not significantly correlated with

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 1      pollution, pollen, or temperature.  Pollutants were not significantly correlated with symptoms
 2      or PEFR.  Overall, the 1991 data indicate a coherence in the associations of summertime
 3      haze air pollution with peak flow, chest symptoms, and asthma exacerbations hi children.
 4      The lack of correlation during 1992 was likely due to the improved air quality and indirectly
 5      supports the results of the previous year.  An adequate interpretation of these preliminary
 6      results is limited by the small number of subjects and study days and the lack of results for
 7      other pollutants.  These camp studies remain to be reported in more detail.
 8            The above epidemiological studies have generally supported a direct association
 9      between ambient O3/oxidant concentrations and acute respiratory morbidity in asthmatics.
10      The recent studies have strengthened their conclusions by improvements or new approaches
11      in the estimations of O3 exposure, confounders,  and  effect modifiers, characterization of the
12      subjects and serial measurements of their responses;  and analytical approaches.  Thus, the
13      aggregate results can  be viewed as biologically and temporally plausible, consistent, and
14     coherent to some extent.  However, the studies share certain  deficiencies such as small
15      numbers of subjects (which may reduce statistical power) and the lack of significant data
16     about individual responses and  their distribution. The independent effect of ambient 63, as
17     estimated by statistical models in epidemiological studies, is difficult, at best, to clearly
18     differentiate from those of copollutants because O3 (or another pollutant such as H+)  may
19     only  be acting as an indicator of the toxic potency of the ambient mixture of pollutants.
20     This, in combination  with  measurement error and uncontrolled associations with other
21     factors, complicates analytical findings about the relationships among components of an
22     ambient mixture and  may not accurately disentangle the effects of O3 in a biologically
23     appropriate fashion.
24
25     7.4.1.3   Aggregate  Population Time Series Studies
26           Aggregate population, or "ecological", time series studies are epidemiological
27     investigations in which the associations between air pollution and human health outcomes are
28     evaluated over time in the population as a whole (e.g., with respect to deaths per day in a
29     given city) and  for which outcomes and exposures are not matched for the individuals within
30     the population.  Indeed, aggregate population time series studies of extreme air pollution
31     episodes have provided some of the clearest evidence of the adverse effects of air pollution

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 1      on humans.  For example, during the historic December 1952 London Fog episode, in which
 2      extremely high sulfur oxide and paniculate matter air pollution levels were experienced,  total
 3      mortality in Greater London rose from roughly 300 to 900 deaths/day, and acute respiratory
 4      hospital admissions rose from 175 to 460/day (Ministry of Health of Great Britain, 1954).
 5      At more routine levels of air pollution, any effects of air pollution are necessarily less
 6      obvious, and, as  shall be discussed below, methodological issues exist as to the proper
 7      analysis and interpretation of such aggregate population time series data.
 8           The previous criteria document (U.S. Environmental Protection Agency, 1986)
 9      discussed several methodological issues with regard to the epidemiological studies of O3  and
10      photochemical oxidants available at that time. Limitations identified included interferences
11      by, or interactions with, other pollutants and meteorological factors in the ambient
12      environment; lack of comprehensive exposure issue assessments, such as individual activity
13      patterns and evaluation of pollutant monitor appropriateness; difficulty in identifying the
14      responsible oxidant species; and inadequate characterization of the study population.
15      However, most of these criticisms are not relevant to time series studies.  For example,
16      because the same population is being followed from day-to-day, the study population acts as
17      its own "control", obviating the need for a detailed population characterization.  Also, the
18      use of central site monitoring data can be appropriate in these studies for two reasons:
19      (1) although O3 concentrations can vary  spatially within an airshed, they are usually highly
20      correlated across sites over time, so that correlational time series studies are not as dependent
21      on detailed exposure assessments as are, for example, cross-sectional studies; and (2) if the
22      ultimate use of these studies is to be included as criteria for ambient standards, the attainment
23      of which  is evaluated at central monitoring stations, then these are the data most relevant for
24      analysis.  Of the concerns raised by the previous criteria document regarding epidemiological
25      studies, then, the most important to these studies is the potential for other correlated
26      environmental factors to confound the unique identification of O3 as a critical causal factor in
27      any health effects found via time series analyses of aggregate population data.
28           One aspect of evaluating tune series epidemiologic studies of the health effects of air
29      pollution that was not directly  raised by the previous criteria document, but which can be
30      crucial to their proper interpretation, is the statistical question as to how each study has
31      addressed the potentially confounding influences of long-wave (e.g., seasonal) variations in

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 1      the health outcome data.  The seasonality of morbidity and mortality was explicitly
 2      mentioned in Hippocrates' treatise on "Airs, Waters, and Places" and has itself been much
 3      studied over the years (Hechter and Goldsmith, 1961). In respiratory diseases such as
 4      asthma, this seasonality of admissions is very common, due in part to the multifactorial
 5      nature of these diseases.  For example, spring and fall rises in pollen and winter influenza
 6      epidemics superimpose long-wave cycles on the day-to-day variations in respiratory hospital
 7      admission rates.  Such long-wave cycles need to be addressed as part of any time series
 8      analysis for two reasons:  (1) they result in strong autocorrelations, which violate the
 9      underlying assumptions of most statistical approaches used to analyze such data;  and (2) their
10     inclusion can lead to misleading conclusions, in that the long-wave relationships  would likely
11      obscure the acute (i.e., short-wave) effects being evaluated. The need to address seasonal
12      cycles in respiratory disease time series data in order to avoid spurious long-wave dominated
13      correlations has been long recognized (e.g., Ipsen et al., 1969), but has all too often been
14     ignored or inadequately addressed in the published literature. Lipfert (1993), in his literature
15      review of hospitalization and air pollution,  also noted the need for such studies to take into
16     account both weekly and seasonal temporal patterns in the data.  Thus, an important criterion
17     for the evaluation of aggregate population time series studies of the acute morbidity and
18     mortality effects of O3 is whether or not the authors have appropriately addressed long-wave
19     periodicities in the data as part of their analysis.
20           There are a variety of statistical approaches available to address long-wave confounding
21     in time series analyses, each having advantages and disadvantages.  The primary goal in
22     invoking such procedures is to eliminate the long-wave autocorrelation "noise" in the data
23     without inadvertently removing any O3-related health effects  "signal" at the same time.
24     In particular, steps that address autocorrelation in the model but also remove or  explain
25     short-wave variance in the health outcome variable of interest (e.g., by applying prefilters to
26     the series that affect periodicities down to a few days, or by analyzing the residuals from
27     prior regressions of the outcome variable on "control" variables that are correlated with 03,
28     such as temperature) carry with them the risk of also removing short-wave associations of
29     interest before the actual analysis has begun.  Similarly,  the inclusion of autoregressive terms
30     to remove all autocorrelation has the advantage of being a very conservative approach, but
31     may also alter any short-wave O3-related health effects signals present in the process.  Less

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 1     severe methods of autocorrelation removal (e.g., the selection of more limited study time
 2     periods during that the series is quasi-stationary, applying longer wave prefilters that do not
 3     affect cycles less than a week in periodicity, calculating deviations from annual cycle sine-
 4     cosine fits of the data series, or using longer term averages of control variables, such as
 5     temperature, in simultaneous regressions) may not remove autocorrelation as completely as
 6     the previously mentioned methods, but have the advantage that they are less likely to affect
 7     the short-wave data variations of interest.  Furthermore, although there are standard
 8     regression diagnostics available to determine whether autocorrelation remains a significant
 9     problem (e.g., the Durbin-Watson statistic), no such check exists to determine whether the
10     more severe autocorrelation removal methods have also inadvertently removed an O3-health
11     effects association of interest. Thus, although steps must be taken in time series analyses to
12     address the  potentially large biases resulting from long-wave (e.g., seasonal)
13     autocorrelations,  care must be taken not to also remove the signal of interest when dealing
14     with the autocorrelation problem.
15
16     Emergency Room Visits and Hospital Admissions
17          Many investigators have evaluated the associations between hospital emergency room
18     visits or hospital admissions and air pollution. Hospital admissions  are far more common (as
19     counts per day) than, for example,  mortality, thereby providing greater statistical reliability
20     and avoiding the distributional complications that may be presented  by low counts.  Also,
21     admission to the  hospital is a well defined endpoint, having the desirable feature that every
22     patient must have been seen by a physician and deemed sick enough to require
23     hospitalization.  Emergency room (ER) visits provide even larger daily counts, but are not as
24     well defined an endpoint because the patient usually decides whether or not to attend the  ER.
25     Furthermore, in a well designed study in Quebec, Canada,  hospital admission diagnosis at
26     discharge was found to be very  reliable, with the study confirming the classification of
27     respiratory  admissions in general 92% of the time,  and asthma admissions 95% of the time
28      (Delfino et al., 1993).  Daily series of hospital admissions thus represent an especially useful
29      research resource for the investigation of the human health consequences of O3 exposure.
30           Hospital admission and emergency room visit studies that have considered
31      O3 associations  are  summarized in  Table 7-21.  In  the previous criteria documents (U.S.

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                 TABLE 7-21.  HOSPITAL ADMISSIONS/VISITS IN RELATION TO PHOTOCHEMICAL
                                      OXIDANT POLLUTION:  TIME SERIES STUDIES8
Concentration(s)
ppm
0.11 -0.28 avg
max 1 h during
low and high
periods,
respectively
(Not reported)

Pollutant Study Description
Oxidant Comparison of admissions to Los Angeles
County Hospital for respiratory and cardiac
conditions during smog and smog-free
periods from August to November, 1954.
Oxidant Respiratory and cardiovascular admissions to
Los Angeles County Hospital for residents
living within 8 mi of downtown Los Angeles
between August and December, 1954.
Results and Comments
No consistent relationship between admissions and high
smog periods; however, statistical analyses were not
reported. Clear seasonal trends in admissions
(increasing from summer to winter) not addressed.
Inconclusive results; partial correlation coefficients
between total oxidants and admissions were variable.
Method of patient selection was not given. Other
pollutants were not considered. Seasonal trend not
Reference
California Dept. of
Public Health
(1955b, 1956b,
1957b)
Brant and Hill
(1964b)
Brant (1965)
                                                                   addressed.
(Not reported)    Oxidant
Admissions of Blue Cross patients to Los
Angeles hospitals with > 100 beds between
March and October, 1961; daily average
concentrations of oxidant, 03, CO, SO2,
NO2,  NO, and PM by Los Angeles air
pollution control districts.
                                                                                                                             1967b)
Correlation coefficients between admissions for         Sterling et aL
allergies, eye inflammation, and acute upper and lower  (1966
respiratory infections and all pollutants were statistically
significant; correlations between cardiovascular and
other respiratory diseases were significant for oxidant,
Og, and SO2; significant positive correlations were
noted with length of hospital stay for SO2, NO2, and
NOX.  Correlations were not significant for T and RH
or for pollutants with other disease categories.  Reported
seasonal variations in admissions and pollution not
addressed.
(Not reported)    Oxidant
Admissions for all adults and children with
acute respiratory illness in four Hamilton,
Ontario hospitals during the 12 mo from
July 1, 1970, to June 30, 1971; city-average
pollution monitoring for OX(KI), SC^, PM,
CoH,  CO, NOX, HC, T, wind direction and
velocity, RH, and pollen.
Correlation found between admissions and an air
pollution index for SO2 and CoH; negative correlation
between T and admissions; and nonsignificant negative
correlations found with concentrations of OX(K1).
However, clear, long-wave trends (e.g., seasonality) not
addressed.
Levy et al. (1977°)
-------
            TABLE 7-21 (cont'd). HOSPITAL ADMISSIONS/VISITS IN RELATION TO PHOTOCHEMICAL
	^	OXIDANT POLLUTION;  TIME SERIES STUDIES8	
Concentration(s)
      ppm
                  Pollutant
             Study Description
               Results and Comments
     Reference
(Not reported)        O3     Emergency room visits for cardiac and
                            respiratory disease in two major hospitals in
                            the city of Chicago during April 1977 to
                            April  1978; 1-h concentrations of O3, SO2,
                            NO2,  NO, and CO from an EPA site close to
                            the hospital, 24-h concentrations of TSP,
                            SO2, and NO2 from the Chicago Air
                            Sampling Network.
                                         No significant association between admissions for any   Namekata et al.
                                         disease groups and O3, CO, or TSP; SO2 and NO      (1979C)
                                         accounted for part of the variation of ER visits for
                                         respiratory and cardiovascular admissions.  Questionable
                                         analysis, including lack of control for confounding,
                                         possible seasonality in admissions (time series not
                                         shown), and model overspecification (e.g., use of wind
                                         speed).
0.07 and 0.39        O3      Emergency room visits and hospital
avg max 1 h                 admissions for children with asthma
during low and               symptoms during periods of high and low air
high periods,                 pollution in Los Angeles from August 1979 to
respectively                  January 1980; daily maximum hourly
                            concentrations of 03, SO2> NO, N02, HC,
                            and CoH; weekly maximum hourly
                            concentrations of SO4~ and TSP; biweekly
                            allergens and daily meteorological variables
                            from regional monitoring stations.
                                         Asthma positively correlated with CoH, HC, NC^, and  Richards et al.
                                         allergens on same day and negatively correlated with O3 (1981°)
                                         and SO2; asthma positively correlated with NO2 on
                                         Days 2 and 3 after exposure; correlations were stronger
                                         on Day 2 for most variables; nonsignificant correlation
                                         for SO4~ and TSP.  Monthly admissions and pollution
                                         data  indicate strong seasonality, which is not accounted
                                         for.  This questionable analysis results in (likely
                                         spurious) positive correlations with CoH, HC, and NO2
                                         and negative correlations with O3.
0.03 and 0.11    Oxidant
avg max 1 h for
low and high
areas,
respectively
Daily hospital emergency room admissions in
four Southern California communities during
1974-1975. Maximum hourly average
concentrations of oxidant, NO2, NO, CO,
SO2, CoH; 24-h average concentrations of
PM and SO4~; and daily meteorological
conditions from monitoring sites 8 km from
the hospitals.
Admissions significantly associated with oxidant and
temperature in all locations.  Long-term trends and
day-of-week effects appropriately controlled, but not
seasonality. Path analysis guided regression used to
discriminate among the intercorrelated pollutant and
meteorological factors, indicating O3 to be most
important only at the highest O3 site.  However,  lack of
catchment area population figures and inadequate
seasonality adjustments prevent quantitative use of
results.
Goldsmith et al.
(1983C)
-------
 2!
 i—>
 u>
OS
               TABLE 7-21 (cont'd).  HOSPITAL ADMISSIONS/VISITS IN RELATION TO PHOTOCHEMICAL
                                   OXTOANT POLLUTION: TIME SERIES STUDIES3
Concentration(s)
ppm
0.03-0.12
avg of max
1 -h/day for
15 stations
0.025-0.075
3 -mo avg of
monthly means
from all city
sites
0.001-0.085
mean 1-h max
O3 (avg of 1 1
sites)
0-0. 13 avg of
max 1 h/day for
two stations
Pollutant Study Description
O3 Admissions to 79 acute-care hospitals in
Southern Ontario for the months of January,
February, July, and August in 1974, 1976-
1983. Hourly average concentrations of
particulate (CoH), O3, SO2, NO2, and daily
temperature from 15 air sampling stations
within the region.
O3 Analysis of quarterly hospital admission rates
for childhood asthma in Hong Kong during
1983-1987 (n = 19). Quarterly means of
SO2, NO2, NO, O3, TSP, and RSP
considered.
O3 Analysis of emergency room visits, by cause,
to acute care hospitals in the Vancouver, BC,
area July 1984-October 1986. SO2, NO2,
O3, SO4, and temperature considered.
O3 Emergency room admissions for COPD in
Barcelona, Spain during 1985-1986. 24-h
avg SO2 and BS city wide averages. 1-h max
SO2, CO, NO2, and O3 obtained from two
stations.
Results and Comments
Excess respiratory admissions most strongly associated
(p<.001) with O3, sulfate, and T during
July and August with 24- and 48-h lag. No such
associations exist for nonrespiratory (control)
diseases. Seasonality minimized by selection of
narrow study period, and day-of-week effects controlled.
SO4~ and O3 highly intercorrelated (r = 0.65), making
effect discrimination difficult. A lack of independent
regression coefficients prevents quantitative application of
results.
Concludes that asthma is negatively correlated with SO2,
but not with O3. Questionable analysis, including use of
quarterly means and lack of seasonality controls.
Summer (May-October) total emergency (but not
respiratory) visits significantly correlated with temperature
and O3. Day-of-week effects addressed. Seasonality
reduced by study period selection, but opposing within
season cycles in asthma visits and O3 not addressed, which
may have weakened reported O3 -respiratory visit
relationship. Also, O3 levels much lower than in previously
studied Southern Ontario.
A weak but statistically significant association found between
COPD admissions and levels of SO2, BS, and CO, after
accounting for seasonality and autocorrelation and during
eliminated from the analysis based on its Initial overall
Reference
Bates and Sizto
(1983, 1987,
1989)
Bates (1985)
Tseng and Li
(1990)
Bates et al.
(1990)
Sunyer et al.
(1991)
                                                          negative correlation with admissions (prior to long-wave
                                                          controls).  Thus, no conclusions regarding Q$ can be made
                                                          from this work.
-------
              TABLE 7-21 (cont'd).  HOSPITAL ADMISSIONS/VISITS IN RELATION TO PHOTOCHEMICAL
	OXIDANT POLLUTION;  TIME SERIES STUDIES3	
Concentration(s)
      ppm
                  Pollutant
                    Study Description
                Results and Comments
     Reference
0-0.04 daily
mean
       Hospital admissions for asthma in Helsinki,
       Finland during 1987 to 1989; 24 h average
       SO2, NO2, TSP, and 03 city wide averages.
After accounting for daily minimum temperature, NO2   Ponka (1991)
and O3 were significantly correlated on the same day as
admissions, while O3 was most significant on the prior
day (p  = 0.006). However, long wave peaks (e.g., in
April for asthma) were not addressed and
autocorrelation was not assessed.
0.06-0.13 mean
of 1000 to
1500 hours O3
(0.12 1-hmax
was  exceeded on
42 of 226 total
study days.
whereas 0.08
was  exceeded on
102  days)
O3      Emergency room visits for asthma,
        bronchitis, and finger wounds
        (a nonrespiratory control) at nine hospitals in
        central New Jersey were analyzed for the
        period May-August 1988 and 1989. Daily
        values of 03 and SO2 obtained from nearest
        of five monitoring sites. Barometric
        pressure, T, RH, and visibility (as an index
        of sulfate) obtained from a Newark net
        station.
Bivariate correlations indicated asthma visits to be
strongly negatively correlated with T and weakly
negatively correlated with O3, suggesting a seasonality
influence, despite limitation to the O3 season.
However, simultaneous regression of asthma visits on
all environmental variables yielded significant (positive)
03 and (negative) T coefficients only, suggesting that T
acted as a long-wave control variable, revealing the
short-wave O3 relationship with asthma.   Day-of-week
effects on visits found unimportant. No environmental
associations seen with bronchitis or control cases (finger
cuts).
Cody et al. (1992)
0.00-0.05 avg of
daily means
from 22 stations
O3     Admissions to 79 acute-care hospitals in
       Southern Ontario for the months of January,
       February, July, and August in 1979-1985.
       Hourly average O3, SO2, NO2, T, RH, wind,
       speed, barometric pressure, and daily average
       TSPandSO4=.
An elaborate reanalysis of the Bates and Sizto (1989)
data set augmented to 1985.  Long-wave influences
controlled using time period subsets and AR modeling.
Despite possible overspecification of models (e.g., use
of wind speed) and AR filtering of the short wave,
results confirm Bates and Sizto's overall conclusions
regarding significant O3 associations. Response to air
pollution estimated to be 19-24% of summer respiratory
admissions, although the exact contribution by O3 to the
total was not estimated.
Lipfert and
Hammerstrom
(1992)
0.01-0.05 3-mo
avg of daily
means from all
city sites
O3     Age-specific quarterly asthmatic hospital
       discharge rates in Hong Kong during
       1983-1989 examined in relation to quarterly
       mean levels of TSP, RSP, NOj, NOX, and
       03 (n = 27).
Concludes that asthma morbidity is correlated with
paniculate matter, but not O3. Questionable analysis,
including use of quarterly means and lack of seasonality
controls.
Tseng et al. (1992)
-------
8
I
I
vo
U)
             TABLE 7-21 (cont'd).  HOSPITAL ADMISSIONS/VISITS IN RELATION  TO PHOTOCHEMICAL
	OXIDANT POLLUTION;  TIME SERIES STUDIES3	
Concentration(s)
           ppm
                      Pollutant
                                         Study Description
                                                                Results and Comments
                                                        Reference
0.03-0.21
1-h daily max at
central site in
each area
O3      Daily emergency admissions to acute care
        hospitals for asthma, total respiratory, and
        control disease categories in the New York
        City, Albany, and Buffalo, NY, metropolitan
        areas during June-August 1988 and 1989;
        daily 1-h maximum 03 and temperature and
        daily average sulfate and acid aerosols (H  )
        considered.
Significant positive associations found for O3, SO4  ,
and H   with asthma and total respiratory admissions,
but not for control categories.  Long-wave and
day-of-week effects removed, and T effects controlled.
Strongest O3 associations in higher pollution year (1988)
and in more urban population centers (Buffalo and New
York, NY).
Thurston et al.
(1992b)
     0.01-0.16
     1-h daily max at
     central site
                     O3     Daily admissions to 22 acute care hospitals in
                            Toronto, Ontario, for asthma, total
                            respiratory, and control disease categories
                            during July and August 1986, 1987, and
                            1988; daily 1-h maximum O3, SO2, NO^, T,
                            and daytime (9:00 a.m.-5:00 p.m.) SO4~~ and
                            H   considered.
                                                 Significant positive correlations found for O3, H  ,
                                                 SO4 = , PM10, and TSP with asthma and for total
                                                 respiratory admissions, but not for SO2 or NO2, and not
                                                 with control admissions.  Long-wave and day-of-week
                                                 effects removed. Multivariate regressions and
                                                 sensitivity analyses suggested that O3 was the pollutant
                                                 of primary importance, but H  may potentiate O3
                                                 effects.  Except for H  , all PM metrics considered
                                                 became nonsignificant  when entered into regressions
                                                 simultaneously with O3.  Ozone significant even after
                                                 dropping days  >0.12 ppm.
                                                    Thurston et al.
                                                    (1993)
    0.01-0.15            O3      Daily emergency respiratory admissions to
     1-h daily max                168 acute care hospitals in Ontario, Canada,
                                 during May-August 1983-1988 were related
                                 to daily levels of O3 and SO4 at the nearest
                                 of 22 and 9 monitoring sites, respectively.
                                 Admissions broken into 0-1, 2-34, 35-64,
                                 and 65 4- age groups, and by geographical
                                 subregion.
                                                                      Ozone and SO4   positively and significantly associated
                                                                      with admissions for asthma and COPD in all age
                                                                      groups.  Associations consistent across regions.
                                                                      Seasonal and day of week effects addressed prior to
                                                                      analysis.  Analyses also controlled for individual
                                                                      hospital influences.  No pollutant associations found for
                                                                      nonrespiratory control admissions.  Simultaneous
                                                                      regressions suggest O3  to be more important than
                                                                      SO4=.
                                                                                                     Burnett et al.
                                                                                                     (1993)
-------
             TABLE 7-21 (cont'd).  HOSPITAL ADMISSIONS/VISITS IN RELATION TO PHOTOCHEMICAL
	OXIDANT POLLUTION;  TIME SERIES STUDIES3	
Concentrations)
      ppm
                  Pollutant
                    Study Description
                                                                                  Results and Comments
                                                      Reference
0.02-0.16
1-h avg daily
max 0.01-0. 12
8-h avg daily
max
O
                           Daily numbers of emergency asthma visits by
                           patients 1-16 years old to an inner city
                           hospital in Atlanta, GA, during June-August
                           1990 were related to daily levels of O3, SO2,
                                 , pollen, and T.
Hospital visits were found to be significantly higher on   White et al. (1993)
days when the previous day's 1-h max O3 exceeded
0.11. No relationship was found below 0.11, or with
8-h avg daily maximum O3.  Day-of-week effects were
accounted for.  Seasonality effect reduced by study
period selection, but probable long-wave seasonal cycles
superimposed on the day-to-day fluctuations were not
directly addressed, which likely weakened the reported
O3-admissions associations.
0.02-0.09           O3     Daily respiratory admissions for patients
1-h avg daily                 >65 years of age in Detroit, MI during 1986
max (10th to                to 1989 were related to daily levels of O3,
90th percentile)             PM10, and temperature.
                                               Pneumonia and COPD respiratory admissions were
                                               found to be significantly associated with both PM10 and
                                               O3, even after eliminating non-compliance days.
                                               Monthly dummy variables were employed to account for
                                               seasonal variations, but day of week effects were not
                                               addressed.  Asthma admissions were not associated with
                                               pollution, but this was attributed to the very low counts
                                               in this category for the elderly.
                                                                                                                     Schwartz (1993)
aSee glossary of terms and symbols for abbreviations and acronyms.
 Reviewed in U.S. Environmental Protection Agency (1978).
°Reviewed in U.S. Environmental Protection Agency (1986).
-------
 1      Environmental Protection Agency,  1978,  1986), such studies were found to give inconsistent
 2      results for reasons that were not apparent. A common weakness of many of those studies,
 3      however, was a failure to control adequately for seasonal differences in hospital usage and
 4      O3 concentration. Therefore, each of the updated critiques in this table now includes an
 5      evaluation as to how the data were (or were not) controlled for long-wave influences (e.g.,
 6      seasonally). With this factor taken into account, the older studies' varying results are now
 7      more understandable.  In a number of these studies, documented long-wave periodicities in
 8      the data had been ignored, resulting in either nonsignificant associations (i.e., California
 9      Department of Public Health, 1955, 1956, 1957;  Brant and Hill, 1964; Brant, 1965; Levy
10      et al., 1977; and Namekata et al., 1979) or even  significant negative correlations between
11      O3 and hospital visits/admissions (Richards et al., 1981) as a result of the generally higher
12      respiratory admission rates in the colder months,  when O3 levels are at their lowest.  Two
13      studies that did not control for seasonably still reported significant positive correlations
14      between hospital admissions  and oxidants (Sterling et al., 1966,  1967; Goldsmith et al.,
15      1983), though Sterling et al.  excluded the winter months from the analysis and Goldsmith
16      et al. did detrend the data.  Also, unlike any of the previously cited studies, both of these
17      analyses controlled for day of week effects on hospital admission rates (e.g., due to
18      consistently lower admissions on weekends),  an important factor in hospital admissions
19      variations that must also be addressed (see Sterling et al., 1966). Moreover, the one
20      previously reviewed study that adequately controlled for both long-wave and day-of-week
21      influences (Bates and Sizto,  1983, 1987, 1989) reported  very significant associations
22      (p < 0.001) between O3 levels and summertime  (July and August) respiratory hospital
23      admissions.  However, other intercorrelated environmental variables (e.g., acidic sulfates)
24      may also have  been cofactors in this association.  Overall, a review of these older studies
25      suggests that, if the  data are  appropriately analyzed, a significant association may be found
26      between elevated ambient O3 concentrations and acute increases  in daily respiratory hospital
27      admissions.
28           Since the last criteria document  (U.S. Environmental Protection Agency, 1986),
29      a number of new ER visit and hospital admissions studies have been completed, a few of
30      which share some of the same statistical flaws found in many of the older studies.  For
31      example, Tseng and Li (1990) and Tseng et al. (1992) failed  to control for the seasonally of

        December 1993                          7-165      DRAFT-DO NOT QUOTE OR CITE
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 1     the admissions and of the pollutants in their statistical analyses of quarterly hospital
 2     admissions in Hong Kong, causing them to report no associations with O3, but a significant
 3     (and likely spurious)  negative correlation of age-specific asthma admissions with quarterly
 4     mean SO2 in the first of these papers, and a significant (and also likely spurious) positive
 5     association with TSP in the second paper, but no association with O3.  Sunyer et al. (1991)
 6     failed to consider seasonality in their initial evaluation  of an O3 relationship with COPD
 7     hospital admissions in Barcelona, Spain; causing them  to eliminate O3  from consideration in
 8     the study and any evaluation of possible health effects.  Also,  Bates et al. (1990), using a
 9     largely descriptive approach, characterized the seasonal periodicities of Vancouver, BC,
10     respiratory ER visits.  Their subanalysis of the warm season (May through October) included
11     a dominant fall asthma peak, which would obscure any summertime O3 associations and,
12     therefore, little can be inferred from this data analysis  about the existence or nonexistence of
13     an acute relationship  between O3 and Vancouver hospital visits for respiratory causes. Ponka
14     (1991) showed significant O3 associations with asthma hospital admissions in Helsinki over a
15     3-year period. The model also included temperature, but did  not directly address noted long-
16     wave variations in both admissions and pollution.  Thus, whether a study has  adequately
17     addressed statistical confounding by the prominent long-wave  cycles in respiratory hospital
18     admissions series, which are clearly dominated by other causes (e.g., the spring pollen, fall
19     respiratory infection, and winter influenza seasons), continues to be a crucial criterion in
20     evaluating the usefulness of a study's results.
21          Fortunately, there are also a number of new studies that have addressed both long-wave
22     and day-of-week influences in  their analyses.  Cody et al. (1992) did not directly control for
23     seasonality, but they  did narrow their analysis of central New Jersey hospital ER visits  to the
24     high O3 season (May through August).  Even so,  their initial  correlational analysis yielded
25     negative associations between hospital visits and both temperature and O3, which suggests
26     that within season long-wave effects existed (e.g., generally higher asthma visits in May,  at
27     the end of the pollen season, when O3 and temperature are lower on average  than in July or
28     August). However, the authors did conduct subsequent regressions of respiratory visits on
29     both temperature and O3 simultaneously, yielding a significant positive coefficient for O3 and
30      a negative coefficient for temperature, which  suggests that the inclusion of temperature may
31      have indirectly accounted for the long wave cycle, allowing the positive short wave O3-visit

        December 1993                          7-166      DRAFT-DO NOT QUOTE OR CITE
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 1      relationship to be seen.  Day of week influences were considered, but found to be
 2      unimportant for these emergency room visit data. No such pollution-hospital visit
 3      relationship was found for finger cut (i.e., control disease) visits.
 4           Thurston et al. (1992b) analyzed unscheduled (emergency) admissions to acute care
 5      hospitals in three New York State metropolitan areas during the summers of 1988 and 1989.
 6      Environmental variables considered included daily 1-h maximum O3 and 24-h average SO4
 7      and acid aerosol (H+) concentrations, as well as daily maximum temperature recorded at
 8      central sites in each community. Long-wave periodicities in the data were reduced by
 9      selecting a June through August study period.  However, because of remaining within-season
10      long-wave cycles in the data series (i.e.,  day-to-day fluctuations superimposed on an annual
11      cycle in admissions),  they were prefiltered using sine and cosine  waves with annual
12      periodicities. Day of week  effects were also controlled via regression.  These adjustments
13      resulted in nonsignificant autocorrelations in the data series and also improved the pollution
14      correlations with admissions.  For example, in New York City, the  same day O3-asthma
15      correlation rose from a  nonsignificant r = 0.04 in the raw data to a significant
16      r = 0.24 after prefiltering.  This shows the importance of addressing long-wave cycles in
17      such data, even  when these  data come from within a single season.  In contrast, correlations
18      between the pollution data and hospital admissions for nonrespiratory control diseases were
19      nonsignificant both before and after prefiltering. The strongest O3-respiratory admissions
20      associations were found during the high pollution 1988 summer and in the most urbanized
21      communities considered (i.e.,  Buffalo and New York City).  After controlling for
22      temperature effects via simultaneous regression,  the summer haze pollutants (i.e., 804",
23      H  , and O3) remained significantly related to total respiratory and asthma admissions.
24      However, these pollutants' high intercorrelation prevented the clear discrimination of a single
25      pollutant as the causal agent.  Depending on the index pollutant,  the admission category, and
26      the city considered, it was found that summer haze pollutants accounted for roughly 5 to
27      20% of June through  August total respiratory and asthma admissions, on average, and that
28      these admissions increased approximately 30% above average on the highest pollution days.
29      Based on  these  models'  regression coefficients,  daily respiratory admissions were estimated
30      to rise 1.4 (± 0.5)  and  3.1  (± 1.6) admissions per hundred ppb  O3 per million persons in
31      New York City and Buffalo, NY, respectively.

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 1           Lipfert and Hammerstrom (1992) reanalyzed the Bates and Sizto (1989) hospital
 2      admissions data set for 79 acute-care hospitals in  southern Ontario, incorporating more
 3      elaborate statistical methods and extending the data set through  1985.  Long-wave influences
 4      were once again reduced by using the short study periods previously employed by Bates and
 5      Sizto (e.g.,  July and August only for summer), as well as by employing prewhitening  and
 6      autoregressive procedures to the data.  Day of week effects were also controlled.  In
 7      addition, the models were much more extensively specified, including a variety of new
 8      meteorological variables which may have caused some confounding with the pollutant
 9      variables (e.g., wind speed correlated at r =  -0.55 with NO2). Despite the application of
10      prefiltering  (which may have inadvertently removed some of the short-wave signal of
11      interest) and possible model overspecification (e.g., the inclusion of wind speed), summer
12      haze pollutants (i.e., O3, SO4, and SO2) were still found to have significant effects on
13      hospital admissions in  southern Ontario.  In contrast, pollution associations with hospital
14      admissions for accidental causes became nonsignificant in these models.  Although air
15      pollution concentrations were generally within U.S. air quality standards, the pollutant mean
16      effect accounted for 19 to 24% of all summer respiratory admissions, though the
17      "responsible" pollutant(s) could not be selected by the authors with certainty.
18           Burnett et al. (1993) also employed the Ontario acute care hospital database to analyze
19      the effects of air pollution on hospital admissions, but their analysis considered all of Ontario
20      and analyzed the data from each individual hospital, rather than aggregating the counts by
21      region. Slow moving  temporal cycles, including seasonal and yearly effects, were removed
22      (via an 19-day moving average equivalent high pass filter) and day-of-week effects were
23      controlled prior to the  analysis.  Poisson  regression techniques were employed because of the
24      low daily admission counts at individual hospitals. Ozone displayed a positive association
25      with respiratory admissions in 91 % of the 168 hospitals, and 5% of summertime (May
26      through August) respiratory admissions were attributed to O3. Positive associations were
27      found in all age groups (0 to 1, 2 to 34, 35 to 64, and 65+).  A parallel analysis of
28      nonrespiratory admissions showed no such associations, which indicates the association
29      specificity.  Ozone was found to be a stronger predictor of admissions than SO4, which
30      accounted for an additional 1 % of summertime respiratory admissions.  Temperature had no
31      effect on the pollution-respiratory admission relationship.  Based upon the reported relative

        December 1993                          7-168     DRAFT-DO NOT QUOTE OR CITE
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 1      risks, this analysis indicates that O3 is associated with 1.3 (± 0.2) respiratory admissions/day
 2      per hundred ppb O3 per million persons in the province of Ontario.
 3          Thurston et al. (1993) focused their analysis of respiratory hospital admissions in the
 4      Toronto metropolitan area during the  summers (July through August) of 1986 to 1988, when
 5      they directly monitored for strong paniculate acidity (H+) pollution on a daily basis at
 6      several sites in that city.  Long-wave  cycles, and their associated autocorrelations, were
 7      removed by first applying an annual periodicity sine-cosine fit to the data (as well as day-of-
 8      week dummy variables) and analyzing the resulting residuals.  Strong and significant positive
 9      associations with asthma and respiratory admissions were found for both O3 and H  , and
10      somewhat weaker significant associations with SO4~,  PM2 5, PM10, and TSP, as measured
11      at a central site in downtown Toronto. No such associations were found for SO2 or NO2,
12      nor for any pollutant with nonrespiratory control admissions. Temperature was only weakly
13      correlated with respiratory admissions and became nonsignificant when entered in regressions
14      with air pollution indices.  Simultaneous regressions and sensitivity analyses indicated that
15      O3 was the summertime haze constituent of greatest importance to respiratory and asthma
16      admissions, though elevated H   was  suggested as a possible potentiator of this effect.
17      During multipollutant, simultaneous regressions on admissions,  O3 was consistently the most
18      significant.  Of the particle metrics, only H+ remained statistically significant when entered
19      into the admissions regressions simultaneously with O3.  Sensitivity analyses also showed that
20      dropping all days above the current U.S. O3 standard  of 0.12 ppm (2 of a total 117 days) did
21      not significantly change the O3 coefficients.  The simultaneous  O3, H+, and temperature
22      model indicated that 21 ± 8%  of all respiratory admissions during the three summers were
23      associated with O3 air pollution, on average, and that  admissions rose an estimated
24      37 + 15%  above otherwise expected  on the highest O3 day (0.159 ppm).  Regression results
25      for the three summers also indicated that daily respiratory admissions increased an estimated
26      2.1 ±  0.8 admissions per day per hundred ppb O3 per million people in the city of Toronto.
27      Moreover, despite differing health care systems, the Toronto regression results for the
28      summer of  1988 were remarkably  consistent with previously reported results for that same
29      summer in Buffalo, NY, (see Table 7-22).
30          White et al. (1993) abstracted daily emergency room visit records from June through
31      August,  1990, at a large inner city hospital in Atlanta, GA.  Daily counts of visits for asthma

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TABLE 7-22. COMPARISON OF REGRESSIONS OF DAILY SUMMERTIME RESPIRATORY
    ADMISSIONS ON POLLUTION AND TEMPERATURE IN TORONTO, ONTARIO,
                BUFFALO, NEW YORK, FOR THE SUMMER OF 1988a
5?
1— *
^D
^D








^j
i
h- »
-J
0


	
1
g
u
1
3
O

City and Year
Toronto
1988 summer

Toronto
1988 summer

Buffalo
1988 summer

Buffalo
1988 summer

Respiratory Admissions
Category
Total Respiratory
(mean = 14. I/day)

Total Asthma
(mean = 9.5/day)

Total Respiratory (mean
= 25.0/day)

Total Asthma
(mean = 7. I/day)

aSee glossary of terms and symbols for abbreviations
Temperature, Pollutant
Model Specification
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
T(LG2),
S04~(LGl)b
H+(LG1)
O3 (LG1)
S04=(LG1)
H+(LGO)
03 (LG1)
S04=(LGO)
H+(LGO)
03 (LG2)
S04=(LG1)
H+(LG1)
03 (LG3)
Pollutant
Regression
Coefficient
(adm//tg/m 710
persons
0.07
0.18
0.011
0.04
0.13
0.007
0.11
0.35
0.015
0.03
0.09
0.006
±
±
±
±
±
±
±
±
±
±
±
±
± SE)
0.03C
0.09d
A
0.005°
0.02d
0.07d
A
0.004C
0.04C
0.12C
J
0.008
0.02d
0.05d
0.002C
Pollutant Mean
Effect (% ± SE)
13.3 ± 5.3
7.7 ± 3.9
26.4 ± 11.8
13.0 ± 6.8
8.1 ± 4.5
25.3 ± 14.9
8.0 ± 2.7
6.4 ± 2.2
18.4 ± 9.9
7.0 ± 3.9
5.6 ± 3.3
23.9 ± 10.1
Max/Mean Pollutant
Relative Risk
(
1.41
1.50
1.34
1.40
1.53
1.32
1.22
1.47
1.25
1.29
1.43
1.25
±
±
±
±
±
±
±
±
±
±
±
±
±
SE)
0.16
0.25
0.15
0.21
0.29
0.19
0.12
0.16
0.09
0.12
0.26
0.14
and acronyms.
bLG = lag between exposure and admission, in days.
cp < 0.01 (one-way
p < 0.05 (one-way

Source: Thurston et

test).
test).

al. (1993).














































-------
 1      or reactive airway disease by patients 1 to 16 years of age were related to daily levels of 03,
 2      SO2, PM10, pollen, and temperature.  Seasonality was likely reduced by the study period
 3      selection, though no effort was made to address possible within-season long-wave cycles in
 4      the data.  Day-of-week and temperature effects were controlled as part of a Poisson model
 5      employed so as to address the small admission numbers at a single hospital.  This model
 6      yielded a 1.42 admissions rate ratio (p = 0.057) for the number of asthma visits following
 7      days with O3 levels equal to or exceeding a 1-h maximum of 0.11 ppm, which is consistent
 8      with the relative risk values reported by Thurston et al. (1992b, 1993).  No admissions
 9      relationship with  O3 was seen below 0.11 ppm, or with 8-h average O3.
10           Schwartz (1993) analyzed O3 and PM10 air pollution relationships with daily hospital
11      admissions of persons 65 years or older in the Detroit, MI metropolitan statistical area during
12      1986 to 1989. Daily counts for pneumonia (mean = 15.7/day), asthma (mean = 0.75/day),
13      and all other chronic obstructive pulmonary diseases (mean = 5.8/day) were regressed on the
14      pollution variables and various seasonal, trend and temperature dummy variables, using
15      Poisson modeling.  However, day-of-week effects were not addressed.  Ozone was analyzed
16      with respect to both its daily  24 h average and 1 h maximum.  Autoregressive analyses and
17      residuals plots indicated no remaining autocorrelation in the model. Both O3 and PM10 were
18      significant in simultaneous pollutant models for pneumonia and COPD, but not asthma
19      (which was ascribed to the low daily counts for this  category).  These simultaneous
20      coefficients were reportedly similar to those from the single pollutant models, though the
21      correlations of the coefficients were not provided. Dropping all days exceeding the 1 h
22      maximum O3 standard did not change the size of the O3 coefficients, which remained
23      significant (p<0.01). Based  on the regression coefficients and data presented, it can be
24      estimated that the mean  effect for O3 (11.6%) was double that for PM10 (5.7%) in the
25      pneumonia model, but comparable for COPD (12.2% for O3 vs. 10.2% for PM10).  On an
26      absolute scale, these results imply that O3 was associated with 1.9 (±0.2) respiratory
27      admissions by the elderly/day per hundred ppb (as a 1 h max) per million persons in the
28      Detroit metropolitan area. This estimate does not include admissions by persons less than
29      65 years of age (which would likely included higher asthma admissions, for example), so that
30      the total respiratory admissions associated with O3 in the entire population would likely be
31      higher than estimated from this work.

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 1     Daily Mortality
 2          Past studies of the possible association of O3 (oxidants) with human mortality
 3     summarized in prior O3 criteria documents (U.S. Environmental Protection Agency, 1978,
 4     1986) were sometimes suggestive of an association, but were each flawed in some way.
 5     These studies are included in Table 7-23, with annotation as to the document in which they
 6     were reported. Most  of these studies considered daily mortality in Los Angeles, CA, during
 7     the 1950s and 1960s.   Unlike most  historical hospital admissions studies, many of these
 8     studies did recognize and attempt to control for seasonality in the data series.  Notable
 9     exceptions are the California Department of Public Health studies (1955, 1956, and 1957),
10     which were further weakened by their qualitative treatment of the air pollution data. The
11     Mills (1957a,b) analyses also employed a questionable exposure assessment method (the
12     Standard Research Institute smog index), which diminishes its usefulness. Massey et al.
13     (1961) reported no significant correlations between community differences in mortality and
14     differences in oxidant levels over time, but compared two communities having very different
15     populations (e.g.,  age distributions), a likely confounder in such cross-sectional comparisons.
16     Mills (1960), while reporting mortality-oxidant associations and effects (370 respiratory and
17     cardiovascular deaths/year), did not control for potential  temperature influences on mortality
18     below 96 °F daily maximum. Hechter and Goldsmith (1961) reanalyzed the Mills (1960)
19     data using a simple sine wave seasonality correction and obtained significant oxidant
20     correlations until they  applied an autocorrelation adjustment, which reportedly caused the
21     pollutant-mortality correlations to drop to nonsignificance (results not presented).  Biersteker
22     and Evendijk  (1976) conducted a t-test of difference analysis of two summers of time series
23     data from Rotterdam during 1974 to 1975. Although significant mortality differences could
24     be seen during 1975 heat-pollution  episodes (0.05 < O3  < 0.125 ppm), no significant
25     mortality increase could be seen during the cleaner and cooler summer's episodes
26     (0.05 < O3  < 0.08 ppm).  Statistical time series methods to address probable confounding
27     by temperature effects were needed. Overall, the various exposure assessment and statistical
28     analyses weaknesses in the studies reported in previous O3 criteria documents have prevented
29     the drawing of definitive conclusions in those past documents as to whether or not there  is a
30     significant association between O3 and human mortality.
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3
                     TABLE 7-23. DAILY MORTALITY ASSOCIATED WITH EXPOSURE
                            TO PHOTOCHEMICAL OXIDANT POLLUTION3
•jj
^—'
-J
Concentration(s)
ppm
< 1 .0 peak
(undefined)
<0.38 max
1-h (?)/day
(Not reported)
0.10-0.42
(undefined)
for 1 48 days
of 1949
(Not reported)
0.02-0.37
average of 1-h
daily max from
all Los Angeles
sites

Pollutant Study Description
Oxidant Relationship between daily concentrations of
photochemical oxidants and daily mortality
among residents of Los Angeles County aged
65 years and over the periods August through
November, 1954, and July through
November, 1955.
Oxidant Data extended to include the period from
1956 through the end of 1959.
Oxidant Relationship between daily maximum oxidant
concentrations and daily cardiac and
03 respiratory mortaility in Los Angeles for the
periods 1947 to 1949; August 1953 through
December 1954; and January through
September, 1955.
Oxidant Comparison of daily mortality in two
Los Angeles County areas similar in T, but
with different levels of daily maximum and
mean oxidant levels (KI); SO2 and CO
concentrations were also measured.
Oxidant Daily respiratory and cardiac death counts for
Los Angeles County during 1956 to 1958
related to daily maximum oxidant
concentrations. All days above 96 °F daily
maximum T eliminated from analysis. Each
day's average of daily oxidant maxima was
related to that day's deviation from monthly
mean mortality.
Results and Comments
Heat had a significant effect on mortality; no consistent
association between mortality and high oxidant
concentrations in the absence of high temperature.
However, seasonal trends were not addressed, and
pollution data treatment was qualitative.

Positive relationship between daily maximum oxidant
concentrations and mean daily death rates on high-smog
versus low-smog days. Questionable exposure analysis,
including use of the "SRI smog index".
No significant correlations between differences hi
mortality and differences in pollutant levels. However,
the populations differed in socioeconomic and age
distribution characteristics.
A stratification of the mortality deviations versus oxidant
concentration revealed increasing mortality with
increasing oxidant concentration, even in the cooler
months. The use of deviations addresses data
seasonal! ty. It is estimated that over 300 deaths/year in
Los Angeles are associated with oxidants. However,
the lack of T controls below 96 °F is a major weakness.
Reference
California
Department of
Public Health
(1955b, 1956b,
1957b)
Tucker (1962)
Mills (1957ab,bb)
Massey et al.
(1961)
Mills (1960)
-------
                         TABLE 7-23  (cont'd).  DAILY MORTALITY ASSOCIATED WITH EXPOSURE
	TO PHOTOCHEMICAL OXIDANT POLLUTION8	
Concentrations)
      ppm
                  Pollutant
                       Study Description
                Results and Comments
    Reference
0.05-0.21
  monthly
  avg
Oxidant   Reanalysis of the relationship between daily
          maximum oxidant concentrations (KI) and
          daily mortality from cardiac and respiratory
          diseases in Los Angeles for the years 1956
          through 1958.
Used deviations from sine wave fit to reduce seasonal! ty   Hechter and
of pollution and mortality, but fit of monthly variations    Goldsmith (1961)
was inadequate.  Significant correlations found between
pollutants and mortality for cardiorespiratory diseases, but
(poorly documented) autocorrelation adjustments by
authors reportedly reduced these associations to
nonsignificance.
0.003-0.128
  max 1-h/day
  O3      Relationship between daily mortality and daily Mortality significantly higher during relatively high
          1 -h maximum concentrations of 03 in
          Rotterdam, The Netherlands during the
          months of July and August, 1974 and 1975.
pollution (0.05  < O3  < 0.125) and heat episodes in
1975.  However, no significant mortality difference due
to moderate O3 episodes (0.05 < O3 < 0.08) in 1974, in
the absence of high T.  Such simplistic analyses of time
series data makes interpretation difficult.
Biersteker and
Evendijk (1976)c
0.02-0.29            03      Total, respiratory, and cardiovascular         Frequency domain analysis indicated a significant short   Shumway et al.
six site mean                 mortality in Los Angeles County during 1970  wave O3-mortality association, but this was not          (1988)
of daily 1-h                  to 1979 related to O3, CO, SO2, NO2, HC,   investigated further. The filtered (i.e., long-wave) data
max                         particulate matter, daily max T, and RH.      analysis indicated O3 to be a nonsignificant contributor to
                             Low-pass filter used to eliminate short-wave   seasonal variations in mortality.
                             associations so that only seasonal associations
                             could be studied.
0.02-0.29            O3      Shumway et al. (1988) 1970 to 1979
six site mean                 Los Angeles mortality dataset reanalyzed
of daily 1-h                  using a high-pass filter to allow investigation
max                         of short-wave (acute) associations with
                             environmental variables, after removing
                             seasonably effects.  Environmental variables
                             considered included temperature, relative
                             humidity, extinction coefficient, carbonaceous
                             particulate matter, SO2, NO2, CO, and O3.
                                                    Filtered environmental and mortality data analyses
                                                    demonstrated significant associations between short-term
                                                    variations in total mortality and pollution, controlling for
                                                    temperature.  Day-of-week effects found not to affect the
                                                    relationships. Of the pollutants considered, O3 had the
                                                    strongest association with total mortality. Similar results
                                                    found for cardiovascular deaths, but not for respiratory
                                                    deaths (for which only T was  significant),
                                                      Kinney and
                                                      Ozkaynak (1991)
-------
cr
VO
                  TABLE 7-23 (cont'd). DAILY MORTALITY ASSOCIATED WITH EXPOSURE
                            TO PHOTOCHEMICAL OXTOANT POLLUTION3
Concentration(s)
ppm
0.000-0.064
24-h avg
^ TN, no
exceedances of
0.12 ppm 1-h
max; in MO,
5 exceedances
with max =
0.15 ppm)
0.00-0.21
daily 1 -h max
See glossary of
Reviewed in U.
Pollutant Study Description
Oj Associations between total daily mortality and
air pollution were investigated in St. Louis,
MO, and Kingston-Harriman, TN, during
September 1985 through August 1986.
Environmental variables considered include
T, RH, PM]0, PM2 5, sulfate, aerosol
acidity, SO2, NO2, and O3.
O3 Associations between total and cause-specific
daily mortality and air pollution were
investigated for New York City during April
through September, 1971 to 1976.
Environmental variables considered included
T, RH, CoH, particulate matter, and O3.
terms and symbols for acronyms and abbreviations.
S. Environmental Protection Agency (1978).
Results and Comments
Statistically significant daily mortality associations were
found with PMjQ, but not with 03. Autocorrelation
removed via season indicators, multiple T/climate
variables, and AR modeling. The nonsignificant O3
coefficient may have been contributed to by the more
conservative autocorrelation removal measures taken,
lower O3 concentrations, and shorter study period,
relative to other recent mortality studies.
Seasonality controlled using high pass filter, as in the
prior Los Angeles analysis. Controlling for T, RH, and
CoH, the previous day's O3 (lag 1) was significant
(p < 0.01) in the total mortality regression. The
elasticity of the resulting C^ regression coefficient is
consistent with the Los Angeles elasticity value.

Reference
Dockery et al.
(1992)
Kinney and
Ozkaynak (1992)

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 1           Although there have been relatively few O3 mortality studies conducted and published
 2     since the last criteria document (U.S. Environmental Protection Agency, 1986), the statistical
 3     methods and pollution data employed in these studies have improved, as compared with the
 4     older studies discussed above.  Shumway et al. (1988) focused on long-wave variations in
 5     mortality, finding that O3 was a nonsignificant contributor to seasonal variations in
 6     Los Angeles mortality during 1970 to 1979.  As might have been expected, temperature was
 7     found to be the principal environmental factor influencing seasonal mortality fluctuations.
 8     This paper's exploratory frequency domain analysis did indicate a significant short-wave
 9     (i.e., cycles on the order of a few days in period) association between O3 and mortality, but
10     this result was not pursued in the subsequent regression analyses.
11           Kinney and Ozkaynak (1991) reanalyzed the 1970 to 1979  Los Angeles County
12     mortality and  environmental data set for short-wave pollution-mortality associations using
13     seasonal and day-of-week controls. After prefiltering the environmental and mortality time
14     series using a high-pass filter, significant associations were demonstrated between air
15     pollution and  short-wave (acute) variations in total mortality, even after controlling for
16     temperature influences. Day-of-week effects were also accounted for, but were found not to
17     affect pollutant-mortality associations. In the regression  models considered, the  1-day lag of
18     O3 concentration gave the strongest pollutant associations with total mortality. This
19     O3 coefficient was  statistically separable from the other significant pollutants in the analysis
20     (CO, NO2,  and paniculate matter), although these other  three pollutants were too
21     intercorrelated to separate from each  other.  Expressed as an elasticity,  the O3 regression
22     coefficient (0.03  ± 0.01 deaths/ppb)  indicated that a 1 % increase in O3 concentration was
23     associated with a 0.015% increase in total mortality.  This result would imply an O3 mean
24     effect of on the order  of 1.5% of total mortality throughout the  year (i.e.,  830 total
25     deaths/year).  Regression results  for cardiovascular deaths (average = 87/day) were
26     qualitatively similar to those for total mortality (average = 152/day), but only temperature
27     was significant for respiratory deaths (average = 8/day), probably due to low count number
28     effects for this category (i.e., Poisson models may have  been  required). Overall, although
29     the Shumway et  al. (1988) analysis of these  1970 to 1979 Los Angeles  data indicates that
30     disease factors and other pollutants dominate the overall seasonal cycles in mortality in Los
31     Angeles, the Kinney and Ozkaynak (1991) short-wave analysis documents that O3 explained a

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 1     small but statistically significant portion of day-to-day variations in total mortality in that
 2     city.
                                                          «
 3          Dockery et al. (1992) conducted an analysis of total daily human mortality in St. Louis,
 4     MO, and Kingston-Harriman, TN, during a  1-year period from September 1985 through
 5     August 1986 aimed primarily at assessing the effects of paniculate matter on mortality. One
 6     of the strengths of this study is the fact that multiple air pollutants were measured and
 7     considered. Thus, as part of the analysis, O3 and other gaseous pollutants were also
 8     considered and found to have nonsignificant associations with mortality in these cities. The
 9     statistical analysis addressed autocorrelation in the mortality data through the use of multiple
10     climate indices (i.e., daily mean temperature, hot day, cold day, humid day, hot  and humid
11     day, season, and interactive terms) and through the  incorporation of autoregressive modeling.
12     This approach is more conservative than that employed by Kinney and Ozkaynak (1991) and
13     others, and the lack of a significant O3 coefficient in this analysis  may be due in  part to the
14     statistical modeling, which may or may not have affected an O3 mortality relationship in the
15     data in the process of conservatively addressing autocorrelation and controlling for
16     temperature (which is usually correlated with O3 over time).  Also, the lack of any
17     03 associations with total mortality may  in part be due to the relatively low O3 levels found
18     in these particular communities (especially in Kingston-Harriman,  where no O3 exceedances
19     occurred) during the study year (maximum 24-h mean O3 <0.065 ppm).  Overall, this study
20     did not show an association between O3 and mortality, but this may be a product of the
21     particular methodological and exposure characteristics of this study vis-a-vis the identification
22     of O3  health effects.
23          Kinney and Ozkaynak (1992) analyzed the associations between daily air pollution
24     concentrations and total and cause-specific mortality in New York City, NY, during April
25     through September, 1971 to 1976, in a manner similar to their prior Los Angeles analysis
26     (Kinney and Ozkaynak, 1991).  Environmental variables considered included the daily
27     concentrations of SO2, paniculate matter (coefficient of haze [CoH]), O3, temperature,
28     relative humidity, and visibility. Long-wave cycles were removed prior to the analysis using
29     a high-pass filter.  Prior to regression analyses, cross correlations  between mortality and each
30     environmental variable were used to determine the optimal lag for each variable.  A multiple
31     simultaneous regression yielded significant coefficients for 1-day lagged O3  (p = 0.004), and

       December  1993                          7-177      DRAFT-DO NOT QUOTE OR CITE
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 1     same day CoH (p = 0.003), temperature (p = 0.0001), and relative humidity (p = 0.0002).
 2     Expressed as an elasticity, the O3 coefficient (0.058 ± 0.03 deaths/ppb) indicated that
 3     a 1 % increase above the mean O3 concentration was associated with a 0.019% increase in
 4     mortality, which  is very close to the value found by these authors for Los Angeles, CA.
 5
 6     7.4.1.4   Summary and Conclusions
 7           Recent epidemiology studies addressing the acute effects of ambient O3 have yielded
 8     significant associations with a wide range of health outcomes, including lung function
 9     decrements, aggravation of preexisting respiratory disease, and increases in daily hospital
10     admissions and mortality.  Individual-level camp and exercise studies clearly indicate that
11     lung function can decrease in an exposure-related manner in response to O3 exposures
12     occurring in ambient air. The combined results of these studies provide useful, quantitative
13     information on the pulmonary effects of ambient O3 exposures.  Results from daily life
14     studies, although more difficult  to interpret quantitatively due to exposure assessment
15     uncertainties, are qualitatively consistent with camp and exercise studies.  There is limited
16     evidence from  several studies suggesting that ambient O3-induced lung function decrements
17     may persist for up to 24 h.  Results from lung function epidemiology  studies are generally
18     consistent with those of human  chamber studies. An O3-related worsening of symptoms in
19     selected groups of healthy individuals and adverse changes in symptoms, lung function, and
20     medication  use in asthmatics have been observed qualitatively and,  to  a lesser extent,
21     quantitatively.  The relationship is consistent, temporally plausible, and moderately coherent.
22     Subsequent research should add more quantitative characterization of individual exposure and
23     speciation of copollutants and their clinical effects, as well as larger numbers of subjects.
24           Emergency room visit and hospital admission studies considered in this document
25     collectively indicate that, when  the major confounders to such analyses are  addressed (e.g.,
26      seasonally  and day of week effects), consistent associations are seen between acute
27     occurrences of respiratory morbidity and O3 exposure.  The evidence is especially strong for
28      hospital admissions, as the association has been seen by numerous  researchers at a variety of
29      localities using a wide  range of appropriate statistical approaches.  Although the absolute size
30      of the effect varied somewhat across localities and statistical approaches, these analyses
31      suggest that, in the summertime (when many other respiratory illness  causes have abated),

        December  1993                           7-178      DRAFT-DO NOT QUOTE OR CITE
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 1      O3 air pollution is associated with a substantial portion (on the order of 10 to 20%) of all
 2      respiratory hospital visits and admissions. Moreover, certain of these analyses also indicate
 3      that, on the highest O3 days, this pollutant's estimated contribution can increase to the point
 4      where it is associated with nearly half of all respiratory hospital admissions.  Moreover,
 5      significant associations are also seen between O3 and hospital visits/admissions below the
 6      U.S.  standard of 0.12 ppm 1-h daily maximum O3.
 7           As was also the case for the O3-hospital admissions time series studies, many of the
 8      older O3-mortality studies had methodological or statistical weaknesses that prevented clear
 9      conclusions.  However, since the previous Criteria Documents, two out of the three relevant
10      new studies (Kinney and Ozkaynak, 1991, 1992) indicated consistent and statistically
11      significant effects by O3  on short-term  (acute) human mortality.  These studies considered
12      two very different locales (Los Angeles, CA, and New York City, NY), though the statistical
13      methods employed were  similar.  The one relevant new study that did not show any
14      O3 association (Dockery  et al., 1992) employed more conservative climate and
15      autocorrelation control methods and was conducted over a much shorter time period than the
16      other two studies. Also, the two studies  that showed O3-mortality associations considered
17      urban areas experiencing 1-h maximum O3 concentrations above 0.15 ppm, whereas the
18      other study areas (eastern Tennessee and St. Louis, MO)  did not.  Thus, although the
19      analysis of daily series of human mortality and air pollution  has yielded small but statistically
20      significant associations with O3, the sensitivity of these associations to statistical modeling
21      methods and to  O3 concentration level  needs further investigation.
22
23      7.4.2   Chronic Effects of Ozone  Exposure
24      7.4.2.1   Introduction
25          At the time of the publication of the previous EPA air quality criteria document (U.S.
26      Environmental Protection Agency, 1986), little useful data were available on the chronic
27      effects of O3 exposure.  Table 11-10 of that document summarized the limited number of
28      studies available at that time and concluded "...it is unlikely  that any of these studies can be
29      used to develop  quantitative exposure-response relationships for ambient oxidant exposures.
30      Further study of well-defined populations over long periods of time  is required before any
31      relationship between photochemical oxidants and the progression of chronic diseases can  be

        December 1993                           7_179      DRAFT-DO NOT QUOTE OR CITE
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 1     conclusively demonstrated from population studies."  The document noted that existing
 2     studies failed to demonstrate any consistent relationship between chronic oxidant exposure
 3     and changes in pulmonary function, chronic symptoms, chromosomal abnormalities, or
 4     chronic disease mortality.
 5          The largest study that had been performed at the time of the 1986 criteria document
 6     was that  of Detels et al.  at the University of California at Los Angeles (UCLA)  (Detels
 7     et al., 1979, 1981; Rokaw et al., 1980).  This study employed a population based sample of
 8     households in  selected communities in the Los Angeles Basin.  A standardized interview was
 9     administered, and individuals underwent  various tests of lung function.  Air pollution data
10     were derived from a network of monitoring stations maintained by the South  Coast Air
11     Quality Management District (California  Air Resources Board). The  usefulness  of the
12     findings of this study was considered to be limited due to a number of factors:  (1) testing in
13     the several study communities occurred at different times over a 4-year period, (2) little data
14     were presented on self-selection (completion rates between 70 to 79%) and migration in and
15     out of the study communities, (3) reproducibility of the pulmonary function measurements
16     could not be demonstrated consistently, (4) mixed ethnicity of the study population,
17     (5) inadequate data on individual exposure and the failure to adjust exposure  estimates for
18     migration in and out of the study areas, and (6) the methods employed for the comparisons
19     of health effects.
20          The 1986 criteria document also summarized the first of the Adventist Health Smog
21     (AHSMOG) studies (Hodgkin et al., 1984) on the occurrence of COPD in relation to chronic
22     air pollution exposures.  However, the data from this first publication were felt to be of
23     limited value because only symptom data were reported and the exposure assessment was
24     insufficient.
25          The 1992 supplement to the 1986 criteria document (U.S. Environmental Protection
26     Agency,  1992) focused most of its attention on more acute health effects attributed to
27     O3 exposure and provided no new data on chronic effects in humans.  The document did
28     summarize important laboratory animal data derived from multiday exposure studies (c.f.,
29     Table 3-5; U.S. Environmental Protection Agency, 1992) that provide a pathologic
30     framework for the occurrence  of chronic health effects in humans.  Data from the primate
31     studies are particularly useful (Tyler et al., 1988; Reiser et al., 1987; Rao et al., 1985a,b;

       December 1993                           7-180      DRAFT-DO NOT QUOTE OR CITE
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 1      Moffatt et al., 1987; Hyde et al.,  1989; Harkema et al., 1987a,b).  Respiratory bronchiolar
 2      changes (bronchiolitis, cell hypertrophy, and shift to more cuboidal cells), alterations of
 3      collagen cross linking and inflammatory cells in the interstitium and airway lumen have been
 4      observed with prolonged exposure of monkeys to levels as low as 0.25 ppm for up to 18 mo.
 5      Of importance for the pathogenesis of chronic effects is the observation by Tyler et al.
 6      (1988) that intermittent exposures were associated with continued injury during nonexposure
 7      periods.  See Chapter 6, Section 6.2.4 for a more detailed discussion of the laboratory
 8      animal data.
 9
10      7.4.2.2   Recent Epidemiological Studies of Effects of Chronic Exposure
11           By the very nature of the problem of the establishment of a link between chronic
12      exposure to O3 and the occurrence of chronic health effects, epidemiological studies remain
13      the principal, if not the only, studies from which data can be sought. As has  been noted in
14      the 1986 document, principal problems for such studies relate to (1) the specification of
15      individual exposures over the relevant periods of life of the study subjects; (2) the coincident
16      effects of other oxidant species (e.g., NOX and derivative acid  species) and other air
17      pollutants (acid aerosols, paniculate species); (3) seasonal effects that relate to pollutant and
18      meteorologic factors that effect specific pulmonary  function measurements relevant during the
19      course of longitudinal studies or over studies that utilize multiple cross-sectional samples; and
20      (4) control for effects of factors such as occupational exposures, cigarette smoking, etc.
21      In addition, past epidemiologic studies have not had access to any human histologic
22      specimens in relation to the exposure groups under study nor have specific mechanisms been
23      investigated to explain any of the  symptom or functional outcomes observed.
24
25      Histologic and Immunologic Effects
26           Sherwin has presented some provocative preliminary, histologic data that he uses to
27      offer a hypothesis on the importance of pathologic changes in the centriacinar region (CAR)
28      of the lung in relation to chronic pulmonary effects of oxidant air pollution (Sherwin, 1991;
29      Sherwin and Richters,  1991). Only the publication that presents the primary data is reviewed
30      (Sherwin, 1991) because there is some redundancy  in the two available publications.
31      Sherwin and Richters obtained lungs from 107 subjects, 15 to 25 years of age, who died of a

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 1     sudden death without evidence of overt disease, lived in Los Angeles County, had no autopsy
 2     evidence or history of drug use, and had no lung trauma.  Abnormalities of the CAR were
 3     evaluated by a pathologist who was "blinded" to basic demographic data. Centriacinar
 4     region disease was defined as the extension of a respiratory bronchiolitis into the proximal
 5     acinar structures (i.e., chronic inflammatory cells and histiocytes into alveolar ducts, sacs
 6     and alveoli immediately adjacent to a respiratory bronchiole).  The odds ratio for severe
 7     CAR disease in subjects who lived in metropolitan Los Angeles versus those who lived in
 8     other cities in Los Angeles County was 4.0 (95% confidence limit [CL], 1.4 to 11.3; this
 9     author's calculation based on data in Sherwin Tables 2 and 3).
10           Unfortunately, no exposure data (or lifetime residence data)  were available for the
11     subjects in the Sherwin study, nor were smoking histories, cotinine results, or occupational
12     histories available.  The smoking history data is of critical importance because a respiratory
13     bronchiolitis has been shown to be an early lesion found in the pulmonary airways of young
14     smokers (Niewoehner et al., 1974).  Additional problems for this study were the fact that
15     most subjects  were of low socioeconomic status and only 10 of the subjects were female.
16     Furthermore,  the study is limited by a lack of quantitative morphometry on the lung
17     specimens and by the lack of a control group from an ambient environment with low oxidant
18     pollution.  Therefore, although Sherwin and Richter's data are of considerable interest,
19     particularly in relation to the primate exposure data cited previously,  they currently are not
20     of value in the determination of appropriate human exposure levels for O3, nor do they even
21     establish the fact that the oxidant environment found in metropolitan Los Angeles, indeed, is
22     responsible for the observed lesions.
23           Zwick et al. (1991) carried out a study of allergic sensitization and cellular immune
24     responses in children from four schools in two Austrian cities.  Subjects were median
25     11 years of age.  Two years of meteorologic data and continuously measured levels of SO2,
26     NO2, and O3  were available for both cities. Monitors were within 2 km of the study
27     schools, except for one O3 monitor that was 13 km from a school in the "high" 03 area.
28      "Allergic diseases" (rhinitis, conjunctivitis, and asthma), response to  prick test antigens, total
29     IgE concentration,  number of subjects with IgE > 100 kU/L, and total IgG concentration did
30     not differ between the subjects in the two cities.  Adjustment for  sex, age, active and passive
31     smoking, and types of cooking and home heating did not alter the results.  Children from the

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  1      "high O3" environment had small,  but statistically significant, decreases in the absolute and
  2      relative numbers of OKT4+ (helper/inducer) T-cells and OKNK+ (natural killer) cells and
  3      increases in OKT8+ (suppressor) T-cells. Adjustment for active and passive smoking and
  4      recent respiratory illness did not alter the results.  The frequency of subjects with a
  5      measurable PD2o to histamine also was increased in the high O3 area.  No relationship
  6      between the T-cell findings and PD20 or any of the other immunologic markers are provided.
  7           The Zwick et al. (1991) results are limited by lack of any exposure data and by lack of
  8      detail for the O3 and other ambient air pollution data.  Except for data on the average
  9      percentage time above specific levels of O3,  there are  no useful data that can be applied to
 10      the observations  reported.  Moreover, the differences observed in the various T-cell subsets
 11      were relatively small and of questionable  biological significance.  There are no analyses that
 12      relate the T-cell findings to the clinical and functional  data (see Table 7-24) that are reported.
 13      Finally, although the communities were said  to be similar on all meteorologic and other
 14      ambient pollution data, inspection of the author's Table 1 (Zwick et al., 1991) indicates that
 15      the mean (averaging time not given) NO2 levels in the "low" O3 community were fourfold
 16      greater than those in the "high" O3 community (42 ng/m3 versus  11 jwg/m3).   No data on
 17      acid species or particulates are provided, although both study cities were free of heavy
 18      industry and heavy traffic.
 19           Calderon-Garciduenas et al. (1992) have studied chronic exposure to the ambient air of
 20      southwestern Metropolitan Mexico  City in relation to histologic abnormalities of the nasal
 21      mucosa.  The exposed group consisted of subjects who spent at least 8 h per day while
 22      working at a Naval hospital in southwestern Metropolitan Mexico City. Ninety-two percent
 23      of the group lived in same area as the hospital and all  had lived in southwestern Metropolitan
 24      Mexico City for  >2 mo (n = 47).  Controls consisted of (1) subjects who lived in Veracruz
 25      and who had not left this area over a period of at least 5 years  before the onset of the study
 26      (n = 12); and (2) new arrivals (<30 days residence in southwestern Metropolitan Mexico
 27      City) at the Naval hospital who came from low-O3, "non-polluted" ports (n = 17).   Nasal
28      biopsies were obtained for all subjects in May through  June, 1990, as were histories on
29      residence, smoking, occupation, allergies, etc.  All three groups were matched for age, sex,
30      and occupation.  There were no difference in familial allergy history or personal smoking
31      (specific data not given in paper).  There was a progressive increase in both nasal symptoms

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     TABLE 7-24.  PATHOLOGIC AND IMMUNOLOGIC CHANGES ASSOCIATED WITH CHRONIC OZONE EXPOSURE8
Concentrations(s) Pollutants and
ppm
iiii v 11 uiiiiiciiuti
/ig/m Variables
Study Description
Results and Comments Reference
     Not Provided
            Not provided,
            Los Angeles County,
            not further specified
                     Autopsy study of lungs from sudden
                     death victims 15 to 25 years whose
                     residence was Los Angeles County;
                     examination of lungs for inflammatory
                     changes in CAR of lungs.
                                    Most severe CAR disease in residents of  Sherwin and
                                    metropolitan Los Angeles County versus  Richters (1991)
                                    other county areas; data limited by lack
                                    of smoking history, personal exposure
                                    and occupational data; interesting
                                    hypothesis—not useful for standards.
•^J
00
     0.095-0.188
     time metric
     not given
186-368
                     Study of allergic sensitization and
                     cellular immune responses in children
                     median age 11 years in two Austrian
                     cities, 1989.
                                    Small increases in OKT4 +
                                    (helper/inducer) and OKT8 +
                                    (suppressor) T-cells and small decrease
                                    in natural killer cells in "high" ozone
                                    community; t number of subjects with
                                    measureable PD2Q histamine in "high"
                                    ozone area; no relationship between
                                    T-cell findings and any clinical
                                    immunologic measure, lung function, or
                                    PD2o; meaning of results unclear.
                                     Zwick et al.
                                     (1991)
     0.150-0.275
     monthly
     average
Approx.
294-539
03
Study of nasal histology in persons
living in southwestern Mexico City and
Veracruz; subjects matched on age,
sex, occupation; similar allergy and
smoking histories.
Increased occurrence of nasal dysplasia
in southwestern Mexico City residents,
especially those with more than 5 years
residence; no data on other air
pollutants; data not relevant for U.S.
conditions because Mexico City residents
are exposed to  O3 levels between 0.100
and 0.400 ppm for several hours each
day all year long, with relatively few
days below 0.100 ppm.
Calderon-
Garciduenas et al.
(1992)
     aSee glossary of terms and symbols for acronyms and abbreviations.
-------
  1      and nasal histologic abnormalities in relation to presumed O3 exposure (Veracruz <  new
  2      arrivals  < chronic residents of southwestern Metropolitan Mexico City).  The principal
  3      histologic lesion was basal cell hyperplasia, with squamous cell metaplasia and mucosal
  4      atrophy occurring less frequently.  Only 11 % of those with greater than 60 days residence in
  5      Southwestern Metropolitan Mexico City showed normal mucosa.
  6           Unfortunately, no ambient air data were presented for SO2 or particles, which are said
  7      to be low relative to other parts of the city.  In addition, because the monthly average
  8      maximal O3  concentrations are (and have been since late 1986) well above the current U.S.
  9      1-h standard of 120 ppb, the Calderon-Garciduenas et al.  (1992) data are of limited value to
 10      understanding low ambient O3 exposures.  (This conclusion probably applies even if one
 11      considers the different concentrations represented by a given parts-per-billion value at
 12      different altitudes.) Subjects in southwestern Metropolitan Mexico City are  subjected to
 13      O3 levels of between  100 and 400 ppb for several hours per day in the winter and spring.
 14      Despite the lack of data on other air pollutants and specific exposure data for individual
 15      subjects, this study does provide useful evidence to suggest upper respiratory damage as a
 16      consequence of prolonged exposure to ambient air containing high levels of O3.
 17
 18      Pulmonary Function,  Respiratory Symptoms, and Chronic Respiratory Disease
 19           The Adventist Health Smog Study.  Since the publication of the 1986 criteria document
 20      and its 1992 supplement (U.S. Environmental Protection Agency, 1986, 1992), a number of
 21      studies have  been published that attempt to define chronic respiratory system health effects in
 22      relationship to ambient O3 concentrations (Hodgkin et al.,  1984; Euler et al., 1987, 1988;
 23      Abbey et al., 1991a,b) (see Table 7-25).  Among these, the series of publications from the
 24      AHSMOG study (Hodgkin et al., 1984;  Euler et al.,  1987, 1988; Abbey et al., 1991a,b) are,
 25      perhaps, the  most important and the most useful  for issues that are related to O3 standards.
 26      Therefore, these studies will be discussed first and as a set.
 27          The basic population for these studies represents California-resident, Seventh-Day
 28      Adventists aged >25  years of age who had lived 11 years or longer (as of August 1976) in
29      either a high-oxidant polluted area (South Coast Air Basin [Los Angeles and vicinity], and a
 30      portion of the nearby Southeast Desert Air Basin) and two low-pollution areas (San Francisco
31      and San Diego).  This  sample was supplemented by an additional group of subjects who met

        December  1993                          7_185      DRAFT-DO NOT QUOTE OR CITE
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I
u>
          TABLE 7-25.  EFFECTS OF CHRONIC OZONE EXPOSURE ON PULMONARY FUNCTION, RESPIRATORY
                                        SYMPTOMS, AND CHRONIC RESPIRATORY DISEASE3
           Concentrations(s)
          ppm
                                      Pollutants and
                                      Environmental
                                        Variables
           Study Description
                                                                                                    Results and Comments
                                      Reference
     0.033 median
     average annual
     hourly value
                         65
Study of relationship of air pollution to
levels of FVC, FEV1; and PEFR based on
1976-80 supplement to NHANES and data
from EPA SAROAD monitoring system;
subjects 6-24 years of age; exposure
values based on hourly O3 values for
previous 365 days; data for TSP, NO2,
and SO2;  and data for important
demographic, smoking, and health
covariates.
Nonlinear relationship between annual
average O3 and function measurements
with threshold at approximately
0.040 ppm;  findings limited by inability
to control for multiple pollutants effects,
relatively crude assignment of exposure;
data consistent with effect on forced
flow at O3 levels at or below 0.12 ppm.
Schwartz
(1989)
~J
oo
     0.034-0.050
     90th percentile
     annual mean
     1 h daily max.
                        67-98
1983-1984 cross-sectional study of 2nd-6th
grade students in Ontario and Manitoba,
Canada; data on SO2, NO2, nitrates, and
sulfates; respiratory health, demographic,
smoking, and home cooking fuel data; and
spirometry.
Ontario town had more O3 days
> 0.080 ppm; small decrements (<2%)
in FVC and FEVj were found in the
Ontario town compared to the Manitoba
town; any O3 possible effects were
completely confounded with SO4 effects.
Stern et al.
(1989)
     0.024-0.031
     Annual mean
     1 h daily max.
                        47-61
1985-1986 cross-sectional study of 3,945
7- to 11-year-old children from five rural
towns in Ontario and five towns in
Saskatchewan, Canada; data on SO2,
sulfates, NO3, NO2, and PM10;
respiratory health multiple covariates;
spirometry including flow at mid lung
volumes.
Ontario towns had higher levels of O3
and SO4 in summer months and for
90  and 99  percentiles of distributions;
90  percentile mean 1-h maxima were
80 ppb versus 47  ppb for Oa and
11.5 /tg/m versus 3.1 /ig/m  for SO4;
magnitude of FEV, and FVC effects
was similar to Stern et al. (1989); no
effect for raid volume flows, except for
subjects with asthma; coincidence of
increased O3 and  SO4 preclude definite
statements concerning O3 effects.
Stern et al.
(1993)
-------
n>

I
Ui
 TABLE 7-25 (cont'd). EFFECTS OF CHRONIC OZONE EXPOSURE ON PULMONARY FUNCTION, RESPIRATORY
                                   SYMPTOMS, AND CHRONIC RESPIRATORY DISEASE3
    Concentrations(s)
         ppm
               /ig/m
Pollutants and
Environmental
  Variables
Study Description
                                                                                               Results and Comments
    Reference
     0.008-0.118
     average
     hourly
     concen-
     trations
     1974-1979
               16-231
                Study of chronic respiratory symptoms in
                adults with use of 1979 National Health
                Interview Survey data and 1974-1979 EPA
                SAROAD data; data on respiratory health,
                demography, and smoking; and data for
                TSP.
                            Data for only 29% of those eligible
                            could be used; average hourly
                            03 concentration over period 1973-1979
                            associated with report of sinusitis and
                            hayfever after control for covariates and
                            TSP; no association with asthma or
                            emphysema; large amount of data
                            reduction, lack of adequate exposure
                            assignment, lack of occupational
                            exposure histories, and lack of adequate
                            data on other pollutant make results very
                            difficult to interpret.
Portney and
Mullahy (1990)
oo
-J
0.015-0.052    29-102
average
HMV
                Cross-sectional study of children ages
                6-15 years in a community in Austrian alps
                divided into three zones based on SC^, NC>2,
                and O^; respiratory health, demographic, and
                spirometry data.
                            Only difference in respiratory history
                            was increased adjusted prevalence of
                            asthma in zone with highest O3 (6.4%;
                            0.052 ppm HMV) versus the zones with
                            lower 63 concentrations (4.8%;
                            0.015 ppm HMV; 2.7%, 0.026 ppm
                            HMV); no meaningful differences in
                            spirometry indices; data limited by use
                            of single monitoring site for 1,200 km
                            area; effects of SO2 and NC>2 on  asthma
                            prevalence not well studied.
                                                                                                                           Schmitzberger
                                                                                                                           et al. (1993)
-------
TABLE 7-25 (cont'd).
oo
00
                             EFFECTS OF CHRONIC OZONE EXPOSURE ON PULMONARY FUNCTION, RESPIRATORY
                                     SYMPTOMS, AND CHRONIC RESPIRATORY DISEASE3
Concentrations(s)
3
ppm ^g/m
0.10-0.20 196-392
3-mo mean
daily peak
hourly
values for
Lancaster
and
Glendora,
respectively
0.04-0.07 78-137
mean peak
daily peak
hourly
values 1972-
1981; Long
Beach and
Lancester,
respectively
(Not reported)
Pollutants and
Environmental
Variables Study Description
Oxidants 5 year follow-up of Lancaster and
Glendora, CA, cohorts; from UCLA
population study of CORD restricted to
nonsmoking, non-Hispanic whites,
7-59 years.
Oxidants 5-6 year follow-up of Lancaster and Long
Beach, CA, cohorts from UCLA CORD
study; Long Beach with higher NO2, SO4,
and TSP then Lancaster.
Oxidant Prevalence of respiratory symptoms in
nonsmoking Seventh Day Adventists
Results and Comments
No difference in respiratory symptoms over
follow-up for either community; across all age
groups, slope of Phase III of N2— washout
deteriorated more rapidly in Glendora; in
subjects ^ 14 years of age, more rapid
decrease in spirometric indices in Glendora;
interpretation hampered by large losses to
follow-up, inability to disentangle multiple
pollutant effects.
All reported excess functional decline for
Long Beach likely due to bias in decline
estimates between locations; data not useful
with regard to possible O3 effects.
Slightly increased prevalence of respiratory
symptoms in high pollution area; after
Reference
Detels et al.
(1987)
Detels et al.
(1991)
Hodgkin
etal. (1984)
                                             residing for at least 11 years in high (South
                                             Coast) and low (San Francisco, San Diego)
                                             photochemical air pollution areas of
                                             California; ARB regional air basin
                                             monitoring data for oxidants, NO2, SO2,
                                             CO, TSP, and SO4 from 1973 to 1976.
                                                                           adjusting for covariables, 15% greater risk for
                                                                           COPD due to air pollution (not specific to
                                                                           oxidants); past smokers had greater risk than
                                                                           never smokers; when past smokers were
                                                                           excluded, risk factors were similar. In
                                                                           addition, insufficient exposure assessment and
                                                                           confounding by environmental conditions limit
                                                                           the quantitative use of this study.
-------
 1-t
VO
u>
                TABLE 7-25 (cont'd). EFFECTS OF CHRONIC OZONE EXPOSURE ON PULMONARY, FUNCTION
      	RESPIRATORY SYMPTOMS, AND CHRONIC RESPIRATORY DISEASE3	
      Concentrations(s)
      ppm
               /ig/m
                         Pollutants and
                         Environmental
                           Variables
           Study Description
             Results and Comments
   Reference
     Not reported
                       Oxidant
Cross-sectional analysis of above
populations; uses hours above various
"threshold" values for oxidants, TSP, SO2
based upon California, EPA, and World
Health Organization max. levels; period
covered 1966-1976; data available for
important covariates (sex, occupation,
environmental tobacco smoke, race, age,
education, past smoking).
OX{10) most significantly associated with COPD
after adjustment for covariates; number of hours
above higher thresholds less significant; when TSP,
SO2> and OX(10) entered in same regression,
TSP(200) only pollutant associated with COPD; high
correlation between OX(10), TSP(200), and SO2
(hours more than 4 pphm). Improved exposure
assessment over previous paper; however, no clear
statement possible about effects of oxidants due to
colinearity with TSP and SO2.
Euler et al.
(1988)
-jj
t—*
00
     Not reported
                       03
Same as Abbey et al, (199la) but analysis
applied to COPD severity and a "multi-
pollutant" analysis performed; also
evaluated effect of using data for different
time periods of ambient air monitoring.
Cumulative incidence of COPD symptoms when each  Abbey et al.
                                                                              pollutant entered separately similar to above study;
                                                                              joint effects of OZ(10) and TSP(200) and mean
                                                                              concentrations of each pollutant evaluated only for
                                                                              cumulative asthma incidence; TSP(200) entered
                                                                              logistic regression in preference to OZ(10) but mean
                                                                              03 concentration entered in preference to mean TSP;
                                                                              change in asthma severity associated with mean
                                                                              O3 concentration (1977-87) and with exceedance
                                                                              frequency for OZ(10), OZ(12), and TSP(200)
                                                                              considered separately; findings for asthma severity
                                                                              similar to cumulative incidence when TSP and
                                                                              03 evaluated together; in no analysis did TSP and
                                                                              O3 both remain jointly significant, nor were there any
                                                                              interactions; data unable unequivocally to disentangle
                                                                              effects of individual pollutants.
                                                                                                                               (1993)
     See glossary of terms and symbols for acronyms and abbreviations.
-------
 1     the 11-year residence requirement but who came from low-exposure rural areas in California,
 2     The total, baseline sample (March 1977) comprised 8,572 individuals, of whom
 3     7,267 enrolled.  From this group, 109 current smokers and 492 subjects who had lived
 4     outside of the designated areas for a portion of the previous  11 years were excluded.
 5     Detailed respiratory illness and occupational histories were obtained.  In these studies,
 6     "COPD" refers to "definite chronic bronchitis", "definite emphysema", and "definite asthma"
 7     as defined by the study questionnaire. Measures of pulmonary function are not included.
 8          Air monitoring data were obtained from the California Air Resources Board (ARE)
 9     monitoring system. Ninety-nine percent of the subjects  (excluding the rural supplement)
10     lived at a distance from the nearest ARE monitoring site that was considered to provide
11     relatively reliable concentration estimates for the outdoor, ambient environment at their
12     residence.  Concentrations at the monitors  were interpolated to the centroid of each
                                                                                 2
13     residential zip code from the three nearest  monitoring sites with the use of a 1/R
14     interpolation.  Subsequent development of  exposure indices took account of the improvements
15     in ARE data after 1973.  Data were available for total oxidants, O3, TSP, SO2, NO2,  carbon
16     monoxide, and SO4 (excluding 1973  to 1975).
17          The initial report from this study was summarized in Table 11-10 of the 1986 Criteria
18     document (U.S. Environmental Protection  Agency, 1986). Based upon a multiple logistic
19     regression that adjusted for smoking, occupation, race, sex,  age, and education, it was
20     estimated that residence in the South Coast Air Basin conferred a 15 % increase in risk for
21     prevalent COPD.  No estimates of exposure  were provided and the data were considered to
22     be of limited utility.
23          In their 1988 publication, Euler et al. provided exposure estimates based upon the
24     cumulative number of hours, over 11 years prior to the baseline,  that individuals lived in
25     environments at various oxidant thresholds, beginning at 10 pphm (OX[10]) (196 /ig/m ) and
26     the total dosage to which they would be exposed.  The estimates in this report did not correct
27     for time spent indoors.  When the OX(10) was the only pollutant considered, each 750 h/year
28     increment in exposure was associated with a 20% increase in risk for COPD in a multiple
29     logistic regression analysis that adjusted for effects of occupation, passive exposure to
30     tobacco smoke, personal smoking, sex, age, race, and education (baseline data only).
31     Moreover, the data were compatible with  a threshold effect at 10 pphm. However, when

        December 1993                          7-190      DRAFT-DO NOT QUOTE OR CITE
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 1     hours above a TSP concentration of 200 /xg/m3 (TSP[200]) and SO2 concentration of 4 pphm
 2     were included in the logistic regression model, only TSP(200) was associated with the
 3     occurrence of COPD.  No significant interactions were found between the various pollutant
 4     thresholds. The authors noted that their failure to control for time spent indoors may have
 5     led to an underestimation of the oxidant effect. Moreover, the fact that 74%  of the variance
 6     of OX(10) was explained by the other pollutants certainly reduced the power of this  study to
 7     detect an independent effect of oxidants on the occurrence of COPD.  The authors also noted
 8     the limitations imposed by the cross-sectional nature of the data that were used in this
 9     analysis.  Thus, on the basis of this study, no clear statement could be made about the
10     chronic respiratory system effects of oxidant exposure.
11          A major improvement in the methods for assessment of exposure was presented in
12     Abbey et al (1991a).  Previous exposure estimates were refined by the computation of
13     "excess concentrations" (concentration minus cutoff, summed over all relevant time periods
14     and corrected for missing data).  Exposures also were corrected for time spent at work and
15     time away from residence,  with estimates provided for the environments where work
16     occurred and for geographic areas away from  residence.  The quality of the interpolations
17     (in terms of distance of monitor from residence zip codes) also was evaluated and
18     incorporated  into the estimates. Adjustments were made for the time spent indoors by
19     individuals.  New indices were developed that were based upon O3, rather than on total
20     oxidants.  The investigators demonstrated correlation coefficients of 0.98 between monthly
21     mean total oxidants  and O3 at concentrations < 12 pphm.  (It should be noted that a more
22     appropriate comparison would have been between the  mean and the differences of the two
23     measurements.)
24          The  above estimates were applied to data that included 6 years of follow-up of the
25     study population (Abbey et al., 1991b).  This analysis focused on incident occurrence of
26     obstructive airways disease (AOD—same definition as for COPD above).  Incident symptoms
27     of AOD were significantly associated with hours above several TSP thresholds, but not with
28     hours above any O3 threshold. There was a suggestion of an association between hours
29     above 10 pphm O3 (OZ[10]) and the 6-year cumulative incidence of asthma (relative risk
30     [RR] for 500 h/year above OZ[10] =  1.40 [95%  CL =  0.90 to 2.34]) and definite bronchitis
31     (RR 1.20  [95% CL  = 0.97 to 1.53]).  Approximately 43% of the study population

       December  1993                           7-191      DRAFT-DO NOT QUOTE OR CITE
-------
 1     experienced at least 500 h in excess of the OZ(10) standard.  Cumulative incidence estimates
 2     were adjusted with the use of Cox proportional hazard models for the same variables noted in
 3     the original publication of Hodgkin et al.  (1984) as well as the presence of possible
 4     symptoms in 1977 and childhood respiratory illness history.  None of the analyses included
 5     both O3 and TSP thresholds.  No data are provided on the details of the subjects available for
 6     the prospective analysis and their representativeness versus the entire base population.
 7     Therefore, assuming no bias due to selective loss to follow-up, these data are consistent with
 8     a small O3 effect and are limited by the same considerations of colinearity and subsequent
 9     reduction of power noted above.
10           Another analysis by Abbey et al. (1993) evaluated changes  in respiratory  symptom
11     severity with the TSP and O3 thresholds noted above.  In this analysis, logistic regression,
12     rather than Cox proportional hazard modeling, was used to assess cumulative incidence of
13     components of the COPD/AOD complex; and multiple, linear regression was used to
14     evaluate changes in symptom  severity.  When O3 was considered by itself, there was a trend
15     toward an increased risk of asthma for a 1,000 h average annual increment in the OZ(10)
16     standard  (RR = 2.07, 95% CL = 0.98 to 4.89). In this analysis, there was a suggestion
17     that recent ambient O3 concentrations were more related to cumulative incidence than past
18     concentrations.   Change in asthma severity score was significantly associated with the  1977
19     to 1987 average annual exceedance frequency for O3 thresholds of 10 and 12 pphm.
20     No significant effects were found for COPD or bronchitis alone.  In contrast to the above
21     study of  cumulative incidence, the investigators carried out an analysis in which TSP(200)
22     and OZ(10) were allowed to compete for  entry into a model to evaluate asthma cumulative
23     incidence and changes in severity.  In the cumulative incidence model that employed
24     exceedance frequencies (number of hours above  threshold), TSP(200) entered before OZ(10);
25     when average annual mean concentrations were used, O3 entered before TSP.  From this, the
26     authors concluded that both TSP and O3 were relevant to asthma cumulative incidence.
27     In no case did both pollutants remain significant simultaneously hi the same regression.
28     No interactions were observed between TSP and O3 for either metric. A similar result was
29     observed for change in asthma severity.  As in previous analyses, there was  a high
30     correlation between TSP(200) and OZ(10) exceedance  frequencies (0.72) and their respective
31     average annual mean concentrations (0.74).

       December 1993                          7-192      DRAFT-DO NOT QUOTE  OR CITE
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 1          The AHSMOG study represents the most extensive effort to date to provide realistic
 2     exposure estimates within the constraints of a large, population-based study.  Moreover, the
 3     exposure estimates for oxidants/O3 have been tied to current standards and have taken into
 4     account many of the sources of inaccuracy and imprecision in the assignment of exposures to
 5     individuals (short of detailed personal monitoring).  As such, they do represent a
 6     considerable improvement over all other studies to date.  Nonetheless, these  data do not
 7     provide unequivocal evidence of an effect of O3 on the outcomes that were studied.  This
 8     largely is due to the difficulty of partitioning effects between O3 and particulates.  However,
 9     it fairly can be concluded that, if there is an effect of exposure to O3 on the  occurrence of
10     the COPD/AOD symptom/disease complex as defined in these studies, such an effect is
11     occurring at exposure levels at or  below 0.12 ppm.
12
13          Other Studies. Subsequent to the publication of the 1986 criteria document, two
14     additional publications have emerged from the UCLA study (Detels et al.,  1987, 1991).  The
15     data presented are derived from the same population bases that were used in  previous
16     publications,  and they are therefore subject to the same limitations that were  cited hi the
17     introduction to this section.
18          In 1987, Detels et al. reported a 5-year follow-up study of white, non-Hispanic subjects
19     from the Lancaster and Glendora study areas.  The 12- and 3-mo mean peak hourly total
20     oxidant values from 1972 to 1982 for Lancaster and Glendora were 7 and  10 pphm and
21     11 and 20 pphm, respectively. Only 47 and 58% of subjects, respectively, were retested
22     with both the questionnaire and measures of lung function.  Effects of air pollution on the
23     days of testing were evaluated by  comparing lung function test results hi a subgroup of
24     individuals who were tested three  times at  3- to 4-mo intervals.  No effect was observed,  but
25     the power to  find differences was  low.  Over the follow-up period,  there were no changes in
26     reported respiratory symptoms for either community.  In adults (^19 years of age) who
27     never smoked, all spirometric and nitrogen-washout results showed more rapid deterioration
28     in Glendora.  Differences were significant  only for mid-expiratory flows and for slope of
29     Phase ffl from the nitrogen-washout curve. The effects were greater in females, in whom
30     changes in  FEV1  also were significant.  In subjects less than 19 years of age, only changes in
       December 1993                           7.193      DRAFT-DO NOT QUOTE OR CITE
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 1     slope of Phase in were significant, although FVC in Glendora females was lower than that
 2     observed in Lancaster.
 3          The results of this study remain limited by the lack of adequate exposure data and the
 4     failure to control for the possible effects of other ambient pollutant differences between the
 5     communities.  Problems with loss-to-follow-up represent a significant issue,  especially for the
 6     pulmonary function measurements, given that approximately 50% of the original subjects
 7     were not available for repeated testing.  Baseline comparability is also of concern because
 8     subjects who were retested in Lancaster had a better slope of Phase HI than  those not
 9     retested. Because this measure most consistently differed between the two study
10     communities, the possibility of selection bias is very real.  Overall, these results do not
11     strengthen the usefulness of this study for the attribution of an effect of oxidant exposure on
12     respiratory health.
13          The 1991 report from the UCLA group compared Lancaster  with Long Beach,  the latter
14     area with relatively high levels of SO2, sulfates, nitrogen oxides, and hydrocarbons as well
15     as increased total oxidant levels (mean daily hourly peak values, 1972 to 1982, 30 pphm
16     versus 110 pphm, respectively) (Detels et al., 1991).  As above, the analysis was restricted
17     to non-Hispanic whites who never smoked cigarettes and with 5 years of follow-up.  Only
18     47% of the Lancaster cohort and 44% of the Long Beach cohort had  pulmonary function
19     retested on  two occasions.  Over the age range 25 through 59 years, changes in slope of
20     Phase m of the nitrogen-washout curve and most spirometric indices  were significantly
21     worse in Long Beach, compared to  Lancaster. In subjects under 25 years, there were
22     significant differences in slope of Phase ffl, especially in subjects 7 to 10 years of age.
23           All of the limitations identified for the 1989 report apply to this report as well.
24     Moreover, comparison between the two communities of the interlaboratory differences
25     (mobile laboratory versus UCLA reference laboratory; 3%  sample) indicated that average
26     annual decrements in FEVj were exaggerated by  -13 mL/year (standard error ±7 mL/year)
27     in Long Beach versus -2 mL/year (±7 mL/year) in Lancaster. Application of this
28     difference to the data in their Table 6A would suggest that the "significant"  difference in
29     FEVj for both males and females may be largely, if not entirely,  due to bias.  Thus, all of
30     the  functional difference reported in this study are suspect on this basis alone.  This, of
        December 1993                           7-194      DRAFT-DO NOT QUOTE OR CITE
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  1      course, ignores any additional biases that may have been due to the large losses to follow-up
  2      in both communities.
  3           A number of additional studies have addressed data relevant to the chronic effects of
  4      O3 on respiratory health (see Table  7-24).
  5           Schwartz (1989) evaluated the effect of air pollution on children and young adults ages
  6      6 to 24 years with the use of data derived from NHANES (National Health and Nutrition
  7      Examination Survey, February 1976 to 1980).  All individuals in each census tract were
  8      assigned average pollutant values derived from monitors located  within 10 miles of the
  9      centroid of the census tract.  Average hourly values for the 365 days preceding spirometry
10      were used, and an annual average was created for O3 (EPA Storage and Retrieval of
11      Aerometric  Data [SAROAD] database).  For O3 (chemiluminescence and ultraviolet  [UV]
12      spectroscopy), six of the seven hourly readings between 11:00 a.m. and 5:00 p.m. were
13      required to include a day's data.  Only 1,005 of the 3,922 (25.6%) of the subjects lived close
14      enough to a monitor to have O3 exposures assigned to them.  Data for TSP, NO2, and SO2
15      were assigned to 47.1, 13.6,  and 21.2%, respectively. Analyses were restricted to
16      consideration of single pollutants because the author reported that there was insufficient
17      overlap between the locations where data were available for all or any combination of
18      pollutants.  Data for a variety of relevant personal and demographic covariates were
19      available. Statistical analyses appropriate to the correlation structure of the data (induced by
20      the sampling design of NHANES) were utilized.  There was an nonlinear relationship
21      between the annual hourly average O3 concentration and FVC, FEVl5 and PEFR.
22      A threshold of effect around 0.04 ppm was  observed, above which there appeared to be a
23      linear decline in FVC (only data shown graphically).  The effect persisted after control for
24      sex, race, age, family income, educational level, chronic respiratory symptoms, and  smoking
25      history.  Results were little affected  by region or use of a 2-year averaging time. Ozone
26      levels above the threshold were significantly associated with an FVC <70%, a result not
27      seen for TSP but observed for NO2.
28           The major limitation of the Schwartz (1989) analysis is the  inability  to distinguish
29      between the effects of O3,  TSP, and NO2 and the choice of only a single metric for
30      O3  (hourly average). Support for the former concern can be seen in the similarity of the
31      effects of NO2 and O3 in the logistic regression analyses, which suggests that the results

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 1     could reflect the joint effect of a number of species in a complex oxidant environment.  The
 2     operating assumption that near-term (1 to 2 years) exposure reasonably reflects a "lifetime"
 3     of exposure is highly suspect in the mobile U.S. population.  In fact, restriction of the
 4     analysis to subjects who still resided in the state in which they were born led to slight
 5     reductions of O3 effect, especially for FEV, and PEFR. Despite these limitations, the data
 6     do suggest that, for children and young adults, if there is a chronic O3 effect (or, more
 7     accurately for these data, a subacute effect) on lung function, it could occur at levels  at or
 8     below  120 ppb.  However, the particular pattern of exposure (peaks, season, etc.) that may
 9     be relevant cannot be discerned from these data.
10          In 1989, Stern et al.  reported a cross-sectional study,  conducted in 1983 to 1984,  of the
11     relationship between respiratory health effects of second through sixth grade children  in
12     two Canadian communities (one in southern Ontario and one in southern Manitoba).  The
13     Ontario region was characterized by low levels of gaseous pollutants (SC^ and NO2) and
14     moderately elevated levels of paniculate sulfate, fine particles, and O3.  Frequent episodes of
15     elevated sulfate and O3 concentrations occurred in the summer and early fall.  The Manitoba
16     community was not subject to the same pattern of transported air pollutants.  Gases and
17     O3 (measured by chemiluminescence) were sampled continuously, and TSP, sulfates,  and
18     total nitrates were sampled every sixth day.  Fixed monitoring stations were established at
19     the center of each community, and monitoring was carried  out from October 1983 to  April
20     1984.  Ozone measurements in Ontario were derived from  sites between 35 to 45 km from
21     the study area. Average annual maximum O3 concentrations were similar in the two
22     communities (0.136 ppm and 0.130 ppm; Ontario and Manitoba, respectively), but the
23     frequency of elevated O3 events (>0.080 ppm, Canadian standard for 1-h max) was more
24     frequent in Ontario (30 days) than  in Manitoba (3 days) in  1983.  Ninety-two percent of
25     subjects (n  = 1,317) provided data from detailed questionnaires, but only 70% (1,010)
26     provided spirometric data  (tested in fall and winter months).  There were no meaningful
27     differences in the prevalence of all of the respiratory health outcomes studied after
28     adjustment for parental smoking, gas cooking, sex, length of residence, parental education,
29     and past respiratory illness history. Ontario children had a 2% lower FVC (adjusted  for age,
30     sex, height, and parental smoking) and a  1.7% lower FEV);  both differences were
31     statistically significant. The differences were somewhat greater when children with

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  1      underlying respiratory illness or symptoms were excluded from the analyses.  These data are
  2      very difficult to interpret in relation to O3 due to the marked colinearity between the O3 and
  3      sulfate levels in Ontario.  Moreover, the differences observed in lung function are very small
  4      (an average of 50 mL and 40 mL for FVC and FEV1? respectively) and of questionable
  5      importance without further follow-up data on the subjects.  Such follow-up data would need
  6      to attempt to identify whether the small decrements observed are "across the board" with
  7      respect to the overall population or the result of decrements in a susceptible subset  of the
  8      population, particularly a set of children at the lower end of the pulmonary function
  9      distribution.  In these later children, small decrements might be associated with adverse
 10      respiratory effects as a result of their already lowered (absolute or relative) levels of lung
 11      function.
 12           Stern and colleagues  (Stern et al.,  1993) extended the 1983 to  1984 Stern study to
 13      10 rural Canadian communities.  Five towns in southwestern Ontario and five towns in
 14      Saskatchewan were selected and studied between September 1985 and March 1986. Children
 15      7 to 11 years of age were  studied (n =  3,945) with techniques similar to the previous study.
 16      In 1986, SO2, NO2, and O3 were continuously monitored through a 10-site network; one site
 17      in each town. Particulates were sampled every 3 days for 24 h in Saskatchewan and every
 18      6 days in Ontario.  Annual mean 1-h maximum O3 concentrations were slightly higher in
 19      Ontario, but  the 90th  and 99th percentile values were much greater (90th:  80 ppb versus
20      47 ppb, 99th:  115 ppb versus 57 ppb).  This was particularly true for the months of June to
21      August.  The levels of PM10 and nitrate did not vary between the areas and were well within
22      the Canadian Ambient Air Quality Objectives. Annual mean SO4 levels were 3 to 4 tunes
23      greater in Ontario communities (6.6 /mi/m3 versus  1.9 /*g/m3).
24           The adjusted (age, sex, parental education,  gas cooking, parental  smoking) prevalence
25      of respiratory symptoms did not differ between the 10 communities. Adjusted (height,
26      weight, plus above adjustment factors) FVC and  FEV, averaged 1.7%  and 1.3%  less,
27      respectively,  in the five Ontario towns.  No differences were observed  for PEFR,
28      FEF25_75%, or Vmax50%. The results did not change when the analysis was restricted to life-
29      long residents or to children without respiratory symptoms.  Although not statistically
30      significant, Ontario children with doctor-diagnosed current asthma had FEF25_75% and
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 1      Vmax50% levels that were 6.6% and 6.5%, respectively, lower than similar children in
 2      Saskatchewan.  Overall,  the prevalence of asthma was 4% for the entire sample.
 3           These results are consistent, in terms of the magnitude of the FEVj and FVC effects,
 4      with those of the previous Stern study (Stern et al., 1989).  In addition,  these data provide
 5      suggestive evidence of enhanced effects for children with current asthma. The two major
 6      limitations of the study are recognized by the authors:  (1) the effects observed cannot be
 7      attributed to O3 or to SO4 (or acid) aerosols and could be due to either part of the pollutant
 8      mixture or attributed to the combination of the component, and (2) the differences in the
 9      mean values that  reported do not take into account the variability in the pulmonary function
10      distribution and the variability of responses across the distribution (see above).  It would
11      have been helpful, in this latter regard, had the investigators provided data on the distribution
12      of the lung  function measurements in the asthmatic subject, in whom there appeared to be a
13      larger effect than in the nonasthmatics.
14           Portney and Mullahy (1990) used the 1979 U.S. National Health Interview Survey and
15      EPA  SAROAD data to explore the relationship between O3 and TSP and chronic respiratory
16      disease.  Average hourly O3 concentrations from 1974 to 1979 were used;  data from 1974 to
17      1979  and data from 1979 alone were evaluated.  Individuals were matched to the nearest
18      centroid of the census tract in which they lived in 1979.  Individuals were excluded if  they
19      lived  >20 miles  from the nearest monitor. Only 29.3% of the 4,500 adults surveyed  who
20      participated in the smoking and respiratory disease supplemental interview and  for whom
21      residential data were available could be included.  Seven different model specifications
22      (probit analysis) evaluated cumulative (5-year) and 1-year effects of O3 on  various
23      respiratory diseases.  Hourly average O3  concentrations, but not TSP concentrations, over
24      1974  to 1979 were significantly associated with the report of sinusitis and hay fever after
25      control for smoking, sex, income,  race, education, temperature, and stability of residence.
26      In contrast, neither O3 nor TSP were associated with reported asthma and emphysema.
27      An enormous amount of data reduction, the lack of individual exposure data, lack of
28      specification of the age and sex distribution of the study population, lack of data on
29      occupational exposures, the use of a single O3 metric, and the limit formulation of the
30      paniculate data all severely limited the usefulness of these data.
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 1          Kilburn et al. (1992) studied the effect of "air pollution" on expiratory flows and vital
 2     capacity in Mexican-American children in Los Angeles. In 1984, 556 second and fifth
 3     graders were studied, and 251  of these were studied again in 1987.  The analytical strategy,
 4     the losses to follow-up, and the lack of reasonable exposure data make the data from this
 5     study virtually uninterpretable.
 6          A study by Castillejos et al.  (1992) evaluated the effects of acute exposures to ambient
 7     O3 concentrations on pulmonary function and respiratory symptoms.  One-hundred and forty-
 8     eight 9-year-old children  in the southwest part of Mexico City were studied between January
 9     and June 1988. Weekly spirometric measurements were made over 10 weeks.  Ambient air
10     data were obtained from the monitoring system maintained by Mexican government and
11     included  hourly values for temperature, relative humidity, and O3 concentration.  Ozone
12     concentration exceeded 120 ppb on 74% of days and "frequently" exceeded 240 ppb.
13     No data are presented for SO2 or particulates, which are said to be low relative to other parts
14     of the  city.  All subjects had to live within 5 km of a monitoring station.  The study
15     demonstrated that levels of FEVj and FEF25_75% were associated with mean hourly O3 levels
16     in the preceding 24, 48, and 168 h.  The authors interpreted their data as consistent with an
17     subacute effect of O3 on measures derived  from spirometry that may be due to an
18     "inflammatory process".  However,  this interpretation seems at odds with the statement in
19     the paper that the initial FEVj measurements for the group did not differ from those
20     observed in a comparable age group in the Harvard Six Cities Study who were not exposed
21     to O3 concentrations as high as those reported in this study.  If the overall level of
22     pulmonary function of this group does not  differ from those children who live in ambient
23     environments with far lower O3 concentrations, the data would suggest that the subacute
24     effects observed are not translated into persistent abnormalities,  at least as can be observed
25     with spirometry.
26          In a brief letter to the Lancet, Austrian investigators (Schmitzberger et al., 1992)
27     described a cross-sectional study of the effects of O3 on the respiratory health of
28     1,156 children, ages 6 to 15 years.  Pulmonary function in two different areas with differing
29     "annual" O3 concentrations (actual metric on which "annual" based not given) were
30     compared (52 ppb versus 26 ppb).  No differences were observed for FVC.   All flow
31     measures (FEVj, FEF50, and FEF75) were significantly lower in the children in the

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 1     "high"-O3 area.  These data are of limited value for a variety of reasons, the most important
 2     being lack of individual exposure data, lack of data on other pollutants, the uninterpretable
 3     specification of "annual" O3 concentration, lack of data on chronic respiratory illness
 4     (especially asthma) and the lack of data on smoking  for the teenage members of the subject
 5     group.
 6          Schmitzberger et al. followed up their preliminary data (Schmitzberger et al., 1992)
 7     with a full report of their study that included additional subjects (Schmitzberger et al.,  1993)
 8     An area in the Austrian Tyrolian alps was studied.  Three zones were identified based upon
 9     ambient air conditions:  (1) Zone 1 was characterized by annual mean SO2 (UV fluorescence)
10     of 20 /xg/m3, monthly mean NO2 (Palmes tubes) of  17 ppb, and annual mean
11     O3 (chemiluminescence) of 15 ppb (max. half-hour mean  =102 ppb);  (2) Zone 2 was
12     characterized by values of 14 /*g/m3, 13 ppb, and 26 ppb (112  ppb), respectively; and
13     (3) Zone 3 was characterized by 12 /xg/m3, 8 ppb, and 52 ppb  (146 ppb), respectively,
14     Children ages 6 to 15 years who lived in the study areas for ^3 years were enrolled.
15     Respiratory health questionnaire  data and forced expiratory flows were obtained.  Full data
16     were available from 81%  of the  enrolled  subjects.  Adjusted (age, sex, environmental
17     tobacco smoke, socioeconomic status, and home heating) levels of FVC and forced flows did
18     not follow the gradient in O3 concentrations.  Although Zone 3 differed significantly from
19     Zone 2 on several measures, there were no meaningful differences with Zone 1.  Adjusted
20     asthma prevalence was highest in Zone 3 (6.4% versus 4.8 and 2.7% for Zones 1 and  2,
21     respectively).  There were no differences for other respiratory  symptoms.  Although the
22     authors conclude that "residence in the area of elevated O3 increases the risk...of low small
23     airway-related lung function", careful inspection of the data does not support this conclusion.
24     This conclusion is based upon the supposed increased frequency of FEVj of less  than 70% in
25     Zone 3 relative to the other zones, although the specific data are not provided. Moreover,
26     the mean levels for all functional measurements are lowest in Zone 1,  the zone with the
27     lowest O3 concentrations and the highest SO2 and NO2 concentrations.  This study is
28     handicapped by the lack of any information that could be used  to access  individual exposures.
29     Moreover, only a single monitoring station was employed that  was placed at the  center of
30      Zone  1, which itself was at the center of the study area (1,200 km2).  No information is
31     provided as to how the concentrations of the various pollutants were estimated for Zones 2

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 1      and 3.  Therefore, these data virtually are of no quantitative value for standard-setting
 2      purposes.
 3
 4      Other Chronic Disease Morbidity and Mortality
 5           Only the Adventist Health Study has provided any data on possible O3-related health
 6      effects other than those related to the respiratory system or malignant disease of the
 7      respiratory system (Abbey et al., 1991b; Mills et al., 1991) (see Table 7-26).  The
 8      population studied and the assignment of exposures has been presented previously (Hodgkin
 9      et al., 1984; Euler et al., 1988; Abbey et al., 1991a).
10           In their initial study based on 6 years of follow-up, Mills et al. (1991) found that for
11      500 h in excess of the O3 10-pphm threshold, there was a relative risk of 2.24 for respiratory
12      cancer incidence after adjustment for a number of factors listed previously.  When the TSP
13      200 yug/m  and O3 10 pphm thresholds were allowed to compete for entry into a Cox
14      proportional hazards model for respiratory cancer  incidence, the O3 threshold entered in
15      preference to TSP. Ozone exposure was not associated with excess respiratory cancer
16      mortality or incidence of nonrespiratory cancer over the 6-year follow-up period.
17           A second chronic  disease study  from the Adventist population extended the above
18      observations to include myocardial infarction and all-cause mortality (Abbey et al.,  1993).
19      Incident chronic respiratory disease also was included in this analysis.  Ambient levels of
20      O3 were not associated  with incidence of myocardial infarction at any of the threshold indices
21      that were tested. Neither the mean concentration of O3 nor any of the thresholds were
22      associated with incidence of chronic respiratory  diseases, as previously defined.  However,
23      there was a trend toward an association between 6-year cumulative incidence of asthma and
24      500-h exceedance of the OZ(10) threshold (RR  =  1.40, 95% CL =  0.99 to 2.34).
25
26      7.4.2.3   Conclusions
27           The body of data that has accumulated since  publication of the previous air quality
28      criteria document for O3 (U.S. Environmental Protection Agency,  1986) provides largely
29      suggestive evidence for health effects of chronic O3 exposure.   Most of the studies suffer
30      from one or another of the following limitations:  (1) simplistic assignment of exposure in
31      terms of choice of O3 metrics,  assignment of exposure, or adequate adjustment for relevant

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 I
 CT
 CD
 VO
 VO
                       TABLE 7-26.  EFFECTS OF CHRONIC OZONE EXPOSURE ON THE INCIDENCE OF
                                           CARDIOVASCULAR AND MALIGNANT DISEASES
Concentrations(s)
 ppm
/ig/m
Pollutants and
Environmental
  Variables
Study Description
                                                                               Results and Comments
                                                                                               Reference
S
     Not reported
                 Hodgkin et al.      Hodgkin et al. (1984) and Abbey et al.
                 (1984)            (199la); analysis based upon exceedance
                                   frequencies 1973-1977 and cancer
                                   cumulative incidence 1977-1982.
                                                             Exceedance of OZ(10) threshold
                                                             borderline associated with respiratory
                                                             cancer; no  association with mean
                                                             concentration; multipollutant analysis with
                                                             TSP(200) and OZ(10), only QZ(10)
                                                             entered the logistic regression for
                                                             respiratory  malignancy; TSP(200) was
                                                             significant for females for all malignancy;
                                                             no association between 03 and of any
                                                             measure of cancer mortality; overall
                                                             results suggestive of O3 effect on
                                                             respiratory  cancer morbidity  at level of
                                                             exposure within range experienced by
                                                             large percentage of study population.
                                                                                        Mills et al. (1991)
     Not reported
                 See above
                         See above
                                                   No association between any O3 threshold
                                                   and all cause mortality or incidence of
                                                   myocardial infarction.
                                                                                                           Abbey et al. (1991b)
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  1     covariates;  and (2) lack of ability to isolate effects related to O3 from those of other
  2     pollutants, especially the paniculate fraction. The Adventist Health Study has made
  3     substantive progress in the problem of the assignment of individual exposures (Abbey et al.,
  4     1991a). Unfortunately, the results from this study cannot disentangle the effects of chronic
  5     O3 exposure from those due to chronic exposure to the paniculate fraction of ambient
  6     pollution.  The study also lacks sufficient power to evaluate the possibility of interactions
  7     between O3 and paniculate pollution in relation to health effects. Thus,  the overall data are
  8     not conclusive, but current evidence is suggestive of possible health effects from chronic
  9     exposure to O3, and research efforts should  continue to look for responses in populations
 10     exposed to levels of O3 at or below 0.12 ppm.
 11
 12
 13     7.5   SUMMARY AND CONCLUSIONS
 14     7.5.1   Controlled Human Studies  of Ozone Exposure
 15     7.5.1.1   Effects on Pulmonary Function
 16     Healthy Subjects
 17          Controlled human O3 exposure studies  have provided the strongest and most
 18     quantifiable exposure-response data on the health effects of O3.  This chapter reviews the
 19     results of studies involving healthy, young adult subjects exposed to O3 concentrations
 20     ranging from 0.08 to 0.75 ppm O3 while at rest or during CE or IE of varying intensity for
 21      periods of up to 8 h.  In many  of these studies, small  sample size and suboptimal
 22     experimental design limit the ability to generalize from a small sample to the larger
 23      population.  Of particular concern in considering studies with small sample sizes is the risk
 24      of making a beta (Type H)  error, the incorrect conclusion that no difference exists between
25      treatments when comparisons are not significantly different.  The likelihood of making  a
 26      Type H error greatly limits the ability to determine the minimum O3 concentration that
27      results in a significant pulmonary response in the larger population.  As a result, the
28      conclusions  drawn from many of the studies  cited in this chapter may underestimate the
29      presence of responses at low O3 concentrations in healthy, young adults.
30          Results from  studies of at-rest  exposures to  O3 for 2 h in healthy adult subjects have
31      demonstrated decrements  in forced expiratory volumes and flows occurring at and above

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 1     0.50 ppm O3 (Folinsbee et al., 1978; Horvath et al., 1979).  Airway resistance is not clearly
 2     affected during at-rest exposure to these O3 concentrations.
 3          With moderate IE for 2 h, eliciting a VE of 30 to 50 L/min, decrements in forced
 4     expiratory volumes and flows,  secondary to decreases in 1C, have been observed in healthy
 5     adult subjects at and above 0.30 ppm O3 (Folinsbee et al., 1978;  Seal et al., 1993b).  With
 6     IE (VE  ^ 65 L/min), pulmonary symptoms and decrements in forced expiratory volumes
 7     and flows are present following 2 h exposures to 0.12 ppm  O3 (McDonnell et al., 1983).
 8     Symptoms are present and decrements in forced expiratory volumes and flows occur at
 9     0.16 to 0.24 ppm O3 following 1 h of continuous heavy exercise  (VE = 55 to 90 L/min)
10     (Adams and Schelegle, 1983; Folinsbee et al., 1984; Avol et al.,  1984; Gong et al., 1986)
11     and following 2 h intermittent heavy exercise (VE « 65 to  68 L/min) (McDonnell et al.,
12     1983; Kulle et al., 1985; Linn  et al., 1986).  With longer exposures of 4 to 8 h duration,
13     responses have been observed at lower O3 concentrations and lower ventilation rates.  In the
14     range of concentrations between 0.08 and 0.16 ppm, a number of studies using moderate IE
15     and durations between 4 and 8  h have shown  significant responses under the following
16     conditions:  0.16 ppm for 4 h of IE at  VE «  40 L/min (Folinsbee et al., 1993),  0.08 to
17     0.12 ppm for 6.6 h of IE at VE « 35 to 40 L/min (Folinsbee et  al., 1988; Horstman et al.,
18     1990), and 0.12 ppm for 8 h of IE at VE « 40 L/min (Hazucha  et al., 1992). Symptom and
19     spirometry responses were increased with increased duration of exposure,  increased
20     O3 concentration, and increased total ventilation.  Airway resistance is only modestly
21     affected with moderate or even heavy exercise combined with O3 exposure to concentrations
22     as high as 0.50 ppm O3 (Folinsbee et al., 1978; McDonnell et al., 1983; Seal et al., 1993).
23     Increased fB and decreased VT, while maintaining VE, occur with exposure to 0.20 to
24     0.24 ppm O3 when combined with heavy exercise for 1 to 2.5 h  (McDonnell et al., 1983;
25     Adams and Schelegle, 1983).  Differences in response to a  given O3 concentration among
26     individuals have been shown to be reproducible (Gliner et al., 1983; McDonnell et al.,
27     1985b), indicating some individuals are consistently more responsive to O3 than others.
28          Group mean decrements in pulmonary function can be roughly estimated when
29     expressed as a nonlinear function of effective (i.e., exposure) dose of O3, the simple product
30     of O3 concentration,  mean ventilation, and exposure duration (Silverman et al., 1976;
31     Folinsbee et al., 1978; Adams et al., 1981).  The O3 concentration ([O3]) appears to make a

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 1     greater impact on the pulmonary function response than does VE or exposure duration
 2     (Folinsbee et al., 1978; Adams et al., 1981) and indeed, Larsen et al. (1991) suggest an
 3     exponent of approximately 4/3 for [O3]. Another way of expressing this relationship is that
 4     doubling [O3] under any given exposure scenario will have a greater impact on spirometry
 5     responses than doubling either VE or exposure duration. However, at any given
 6     O3 concentration, the major external determinants of response are VE and exposure duration.
 7     Because of the broad range of intersubject variability, and the inability to identify
 8     characteristics that influence this variability (other than age), efforts to estimate or model
 9     individual responses have so far been fruitless (McDonnell et al.,  1993).  Nevertheless,
10     prediction of group mean FEVj responses  using the variables of [O3], VE,  and exposure
11     duration can be successful (Adams et al., 1981; Folinsbee et al., 1978, 1988; Hazucha,
12     1987; Hazucha et al., 1992; Larsen  et al., 1991; McDonnell et al.,  1993).
13          In acute O3 exposure studies of 3 h or less in duration, the responses  observed during
14     and following acute exposure to O3  at concentrations between 0.12 and 0.50 ppm in normal
15     healthy human subjects include decreases in TLC, 1C, FVC, FEVl5 FFJF25_75%, and VT and
16     increases in SR^, fB, and airway responsiveness. Ozone exposure has also been shown to
17     result in the symptoms of cough, PDI,  SB, throat irritation, and wheezing.   Similar responses
18     are seen with 4- to 8-h exposures in the O3 concentration range between 0.08 and 0.16 ppm.
19          When viewed collectively, these physiological and symptom responses may be separated
20     into four general categories, including (1)  symptoms, (2) changes in lung volume or
21     spirometry,  (3) changes in airway resistance, and (4) changes  in airway responsiveness.
22     These  categories are based on the absence of correlation between  spirometry responses and
23     change in airway resistance or airway responsiveness.  The attenuation by atropine of Raw,
24     but not spirometry, responses supports  the notion of independent mechanisms. The
25     attenuation by indomethacin or ibuprofen of spirometry responses but not changes in  airway
26     resistance or airway responsiveness  also supports this categorization. A bronchodilator,
27     albuterol, given to healthy subjects prior to O3 exposure did not prevent changes in
28     spirometry,  symptoms, or airway responsiveness.  Symptoms ratings represent reflex
29     responses (e.g., cough) or a perceptual evaluation of consciously appreciated afferent
30     information (e.g., chest tightness, pain on  deep inspiration)  and it is therefore somewhat
31     difficult to separate these responses  from the more objective physiological responses.

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 1     However, cough and pain on deep inspiration are related temporally to spirometry and
 2     breathing pattern responses (i.e., volume-related changes).  In repeated exposure studies,
 3     changes in spirometry and breathing pattern become attenuated with the same time course as
 4     the changes in symptom responses.
 5          Recent multihour O3 exposure studies indicate that spirometry and symptom responses
 6     to concentrations as low as 0.08 ppm  occur in healthy subjects with exposures lasting 6 to
 7     8 h. Prolonged exposures (8 h) at lower Oj concentrations (0.12 ppm) also indicate that
 8     there is a plateau of response to O3 (Hazucha et al., 1992).  Although suggested in previous
 9     studies (Gliner et al., 1983),  such a plateau is difficult to verify with the typical duration of
10     less than 2 h and the large responses seen with higher concentrations. The level of the
11     response plateau (i.e., the spirometry  decrement at which the response no longer changes)
12     must be dependant upon the dose rate of exposure (i.e., the product of [03] and  VE) because
13     the magnitude of response at a higher dose rate may greatly exceed the response plateau seen
14     at a lower  dose rate.  Prolonged exposure studies also suggest that O3-induced spirometry
15     responses depend upon the immediate exposure history. With relatively low dose rates (e.g.,
16     Hazucha et al., 1992), responses to exposure that occurred 2 to 4 h previously may influence
17     the current response.  The cumulative effect of exposures has not been studied at higher dose
18     rates, but greater persistence of effects may be expected based on the longer recovery period
19     at higher doses rates.
20          Recovery from O3 exposure has not been systematically investigated in a large group of
21     subjects, but available information indicates that an initial phase  of recovery proceeds
22     relatively rapidly and some 40 to 65 % of the acute response appears to be recovered within
23     about 2 h (Folinsbee and  Hazucha, 1989). However, there is some indication that the
24     spirometric responses, at  least to higher O3 concentrations, are not fully recovered within
25     24 h (Folinsbee and Horvath, 1986; Folinsbee et al., 1993a). Collectively, these
26     observations suggest that  there is a rapid recovery of O3 induced spirometry and symptom
27     responses, which may occur during resting exposure to O3 (Folinsbee et al.,  1977b) or as
28     O3 concentration is reduced during exposure (Hazucha et al., 1992), and a slower phase,
29      which in some cases may take at least 24 h to complete. Repeated exposure studies at higher
30      concentrations typically show that the response to O3 is enhanced on the second of several
31      days of exposure.  This enhanced response suggests a residual effect of the previous

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 1     exposure, about 22 h earlier, even though the preexposure spirometry may be the same as on
 2     the previous day.  The absence of the enhanced response with repeated exposure at lower
 3     O3 concentrations may be the result of a more complete recovery and/or less damage to
 4     pulmonary tissues.
 5           Studies of repeated daily exposure to O3 have shown that O3-induced changes in
 6     spirometry, symptoms, airway resistance, airway responsiveness, and airway inflammation
 7     are attenuated with repetitive exposure.  At higher dose rates, symptom and spirometry
 8     responses may be enhanced on the second exposure.  Attenuation of response within 3 to
 9     5 days is a consistent finding in repeated exposure studies, regardless of O3 exposure dose
10     rate,  although attenuation of response occurs after fewer exposures at the lower dose rates.
11     The attenuation of response appears to occur more rapidly in less responsive individuals
12     (Horvath et al., 1981) or in responsive subjects exposed to lower O3 dose rates  (Folinsbee
13     et al., 1979, 1993b). Loss of attenuation is relatively rapid with responsiveness being
14     partially restored within 4 to 7 days (Kulle et al.,  1982; Linn et al., 1982) and normal
15     responsiveness within 1 to 2 weeks after a series of 4 or 5 daily O3 exposures.  The
16     attenuation of airway responsiveness may occur somewhat more slowly than for symptom
17     and spirometry responses.  Airway inflammation also appears to  attenuate, but less
18     completely than the  spirometry responses and with a  more gradual  recovery (Devlin et al.,
19     1993b; Folinsbee et al.,  1993b).  Some markers of inflammation (e.g., LDH and elastase)
20     have  not demonstrated attenuation.
21           The mechanisms leading to the observed pulmonary responses induced by O3 are
22     beginning to be better understood. The available descriptive data suggest several possible
23     mechanisms, some leading to alterations in lung volumes, symptoms, and exercise breathing
24     pattern, and others leading to increases  in central and peripheral  airway resistance.  These
25     mechanisms appear to involve (1) O3 reactions with the airway lining fluid and/or epithelial
26     cell membranes; (2) local tissue responses, including  injury and inflammation; and
27     (3) stimulation of neural  afferents (bronchial C fibers) and the resulting reflex responses and
28     symptoms.  More studies need to be conducted to determine how each event in this cascade
29     contributes to the pulmonary responses induced by acute ozone inhalation in human subjects.
30
31

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 1     Subjects with Preexisting Disease
 2           Of the subpopulations studied, those with preexisting limitations in pulmonary function
 3     and exercise capacity are of primary concern in evaluating the health effects of 03. Studies
 4     on the acute effects of O3 inhalation in subjects with various preexisting diseases were
 5     reviewed.  Inherent in these studies are several limitations that, at present, hamper our ability
 6     to make a definitive conclusions regarding the relative O3 responsiveness of subjects studied.
 7     Furthermore,  it is necessary to clarify whether their responses are representative of the larger
 8     population with preexisting disease.  These limitations include subject selection, the absence
 9     of standardized methods of characterization, and the limited range of exposure doses utilized.
10           These limitations are evident in studies on subjects with COPD,  chronic bronchitis, and
11     ischemic heart disease.  For patients with COPD performing light to moderate IE, no
12     decrements in pulmonary function were observed for  1- and 2-h exposures to
13     O3 concentrations of 0.30 ppm and less (Linn et al., 1982a, 1983a; Solic et al., 1982; Kehrl
14     et al., 1985) and only small decreases in forced expiratory volume were observed for
15     3  h exposures of chronic bronchitics to 0.41 ppm O3 (Kulle et al.,  1984).  Small decreases in
16     arterial blood oxygen saturation have also been observed in some of these studies,  but the
17     interpretation of these results and their clinical significance is uncertain.
18           Similar limitations also apply to the early studies examining O3 effects in adult and
19     adolescent asthmatics.  Decrements in pulmonary function were not observed for adult
20     asthmatics exposed for  2 h at rest (Silverman, 1979) or with intermittent light exercise (Linn
21     et al., 1978) to O3 concentrations of 0.25 and less. Similarly,  no significant changes  in
22     pulmonary function or symptoms were found in adolescent asthmatics exposed for 1 h at rest
23     to 0.12 ppm O3 (Koenig et al.,  1985) and in adolescent asthmatics and nonasthmatics
24     exposed to 0.12 and 0.18 ppm O3 with intermittent moderate exercise up to 1 h (Koenig
25     et al., 1987, 1988), although a small decrease in forced expiratory flow at 50% of FVC was
26     observed in asthmatics  after exposure to 0.12 ppm O3.  More recent observations by Kreit
27     et al. (1989), Eschenbacher et al.  (1989), and Linn et al. (1993) suggest that mild to
28     moderate asthmatics are at least as sensitive to the acute effects of O3 inhalation as healthy
29     subjects when they are exposed to O3 under conditions that elicit a significant response in
30     healthy subjects.  Kreit et  al. (1989) and Eschenbacher et al. (1989) exposed adult asthmatic
31     and nonasthmatic subjects  to 0.40 ppm  O3 with intermittent moderate exercise for 2 h and

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 1     observed a greater response in airway resistance,  FEVj, and FEF25_75% in the asthmatic
 2     subjects, although changes in FVC and  symptoms were similar in both groups.  Ozone
 3     exposure also resulted in a marked increase in airway responsiveness to methacholine in both
 4     the asthmatic and nonasthmatic subjects. These responses take on greater importance when it
 5     is considered that the observed O3-induced pulmonary effects were superimposed upon
 6     preexisting impairment of pulmonary function and airway responsiveness.  In addition, the
 7     observations of Koenig et al.  (1990) and Molfino et al. (1991) suggest the possibility that
 8     acute exposure to O3 at doses that do not produce measurable pulmonary function decrements
 9     may increase the responsiveness of asthmatics to inhaled  SO2 or antigens.
10
11     7.5.1.2   Effects on Exercise Performance
12          Endurance exercise performance and  VO2max may  be limited by acute exposure to
13     O3 (Adams and Schelegle, 1983; Schelegle and Adams, 1986; Gong et al., 1986; Foxcroft
14     and Adams, 1986; Folinsbee et al., 1977b; Under et al.,  1988).  Gong et al. (1986) and
15     Schelegle and Adams (1986) found that significant reductions in maximal endurance exercise
16     performance may occur in well conditioned athletes while performing continuous exercise
17     (VE > 80 L/min) for 1 h at O3 concentrations >0.18 ppm.  Whereas, data from Linder
18     et al. (1988) suggest that small decrements in maximal exercise performance may occur at
19     O3 concentrations less than 0.18 ppm.   The mechanisms that lead to these responses and the
20     minimum O3 concentration at which these effects occur have not yet been clearly defined.
21     Reports from studies of exposure to O3  during high-intensity exercise indicate that breathing
22     discomfort associated with maximal ventilation may be an important factor in limiting
23     exercise performance.  However, these  studies do not exclude the possibility that some as yet
24     undefined physiological mechanism may limit exercise performance.
25
26     7.5.1.3   Effects on Airway Responsiveness
27          Ozone exposure causes an increase in nonspecific airway responsiveness as indicated by
28     a reduction in the concentration of methacholine or histamine required to produce a given
29     reduction in FEVj or increase in SRaw.   Increased airway responsiveness is an important
30     consequence of exposure to O3 because  its presence means that the airways are predisposed
31     to narrowing  upon inhalation of a variety of stimuli  (e.g., specific allergens, SO2, cold air).

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 1     Markedly increased airway responsiveness is a classical feature of asthma and may also be
 2     present with other respiratory diseases (e.g., chronic bronchitis and acute viral infections)
 3     and even in a sizeable percentage of the healthy asymptomatic population. Many studies
 4     have demonstrated O3-induced increases in nonspecific airway responsiveness in healthy
 5     subjects after  1 to 2 h exposure to concentrations  in the range of 0.2 to 0.6 ppm (Golden
 6     et al.,  1976; Holtzman et al., 1979; Konig et al.,  1980; Dimeo et al., 1981; Gong et al.,
 7     1986; Folinsbee et al., 1989) and after 6.6 h exposure to concentrations in the range of
 8     0.08 to 0.12 ppm (Folinsbee et al., 1988; Horstman et al.,  1990).  Ozone-induced increases
 9     in airway responsiveness tend to resolve within 24 h after exposure but may persist in
10     selected individuals for longer periods (Golden et  al., 1978).
11          Ozone exposure of asthmatic subjects,  who characteristically have increased airway
12     responsiveness at baseline,  can cause further increases in responsiveness (Rreit et al., 1989).
13     The difference in baseline airway responsiveness between healthy and mild asthmatic subjects
14     may be as  much as 100-fold, whereas the changes in airway responsiveness induced by
15     O3 are  typically two-  to fourfold.  Similar relative changes  in airway responsiveness are seen
16     in asthmatics exposed to O3 despite their markedly different baseline airway responsiveness.
17     One study  (Molfino et al.,  1991) has been published suggesting an increase in specific (i.e.,
18     allergen-induced) airway reactivity.  This response was observed after a 1-h resting exposure
19     of atopic asthmatics to 0.12 ppm O3. One of the important aspects of this observation of
20     increased airway responsiveness after O3 exposure is that this represents a plausible link
21     between ambient O3 exposure and increased hospital admissions for asthma. However,
22     experimental design flaws preclude the use of this study in  the determination of a LOEL.
23          Changes in airway responsiveness after O3 exposure appear to be resolved more slowly
24     than changes in FEVj or respiratory symptoms.  Furthermore, in studies of repeated
25     exposure to O3, changes in airway responsiveness tend to be  somewhat less  susceptible to
26     attenuation with consecutive  exposures than changes in FEVj (Dimeo et al., 1981; Kulle
27     et al., 1982a; Folinsbee  et  al.,  1993). The question of whether chronic O3 exposure can
28     induce  a persistent increase (or decrease) in airways responsiveness has not been adequately
29     studied.
30           Increases in airway responsiveness  do not appear to be strongly associated with
31     decrements in lung function or increases in  symptoms.  A number of observations support

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  1      this conclusion. First, asthmatics who have widely different baseline airway responsiveness
  2      show similar FEVj changes after O3 exposure to those seen in healthy subjects (Kreit et al.,
  3      1989).  The changes in airway responsiveness are not correlated with changes in FEVl5
  4      although a weak association has been seen with increased airway responsiveness and PGE2
  5      levels in BAL fluid (McDonnell et al., 1990).
  6           The mechanism of O3-induced  increases in airway responsiveness is only partially
  7      understood, but it appears to be a consequence of cellular and biochemical changes in airway
  8      tissue.  Airway inflammation may be temporally associated with the presence of increased
  9      airway  responsiveness (Holtzman et al., 1983; O'Byrne et al.,  1984; Seltzer et al., 1986),
10      but at least one study in rats demonstrated that increased airway responsiveness could be
11      induced by O3  exposure in the absence of an influx of neutrophils (i.e., airway inflammation)
12      into the airway mucosa (Evans et al., 1988).  Interestingly, indomethacin treatment prior to
13      exposure in one human study blocked the effect of O3 on FEVj and FVC but not on airway
14      responsiveness  (Ying et al., 1990).  Because cyclooxygenase inhibitors have not been
15      effective at blocking the O3-induced  influx of neutrophils into BAL fluid in other studies
16      (Hazucha,  1991; Kleeberger and Hudak, 1992), this result provides further evidence that
17      inflammation is not required for the induction of increased airway responsiveness.
18
19      7.5.1.4   Inflammation and Host Defense Effects
20           A number of studies clearly show that a single acute exposure  (1 to 4 h) of humans to
21      moderate concentrations of O3 (0.2 to 0.6 ppm) while exercising at moderate to heavy levels
22      results in a number of cellular and biochemical changes in the lung, as assessed by
23      measurement of bronchoalveolar lavage constituents (Seltzer et al., 1986; Kehrl et al., 1987,
24      1989; Koren et al., 1989a,b, 1991; Schelegle et al., 1991; McGee et al.,  1990; Aris et al.,
25      1993; Devlin et al.,  1993b).  These exposures result in an inflammatory response
26      characterized by increased numbers of PMNs, increased permeability of the epithelial cells
27      lining the respiratory tract, cell damage, and production of proinflammatory cytokines and
28      prostaglandins.   This response can be detected as early as 1 h after exposure (Koren et al.,
29      1991; Schelegle et al., 1991) and persists for at least  18 h (Koren et al., 1989; Aris et al.,
30      1994).  The response profile of these mediators is not  adequately defined, although it is  clear
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 1     that the time course of response varies for different mediators and cells (Schelegle et al.,
 2     1991, Koren et al., 1989, 1991).
 3          A single study (Devlin et al., 1991) provides evidence that many of these changes also
 4     occur in humans exposed to near-ambient levels of O3 (0.08 to 0.10 ppm) with moderate
 5     exercise for 6.6 h.  Decrements in the ability of alveolar macrophages to phagocytose
 6     microorganisms were also reported in this study.
 7          Ozone also causes inflammatory changes in the nose, as  indicated by increased levels of
 8     PMNs and albumin, a marker for increased epithelial cell permeability. Increases in tryptase
 9     levels immediately after O3 exposure, suggested the release of mast cell products.
10          There appears to be no strong correlation between any of the measured cellular and
11     biochemical changes and changes in lung function measurements, suggesting that different
12     mechanisms may be responsible for these processes.   Alternatively, the absence of a
13     correlation may reflect either the temporal misalignment of these measurements,  the fact that
14     changes detected in the lavage fluid do not quantitatively reflect events occurring in tissues
15     where functional or symptomatic events originate, or that lavage fluid may not be collected
16     from the same lung region primarily  implicated in pulmonary  function responses. The idea
17     of different mechanisms is supported by a study in which ibuprofen, a cyclooxygenase
18     inhibitor,  blunted the O3-induced decrements in lung function  without altering the O3-induced
19     increase in PMNs  or epithelial cell permeability, although ibuprofen did change the
20     concentration  of a number of mediators, some of which may be related to changes in
21     function (Hazucha et al., 1993).
22          In vitro  studies suggest that epithelial cells are the primary target of O3 in the lung and
23     that O3 induces them to produce many of the mediators found in the BAL fluid of humans
24     exposed to O3. Although O3 does not induce alveolar macrophages to produce these
25     compounds in large quantities, it does directly  impair their ability to phagocytose and kill
26     microorganisms.
27
28     7.5.1.5  Factors Modifying Responsiveness  to Ozone
29           Many variables that at least have potential for influencing response to O3 remain
30      inadequately addressed in the available clinical data.  Factors such as smoking status, age,
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 1     gender, race or ethnic group, season, and mode of breathing during exposure have been
 2     evaluated for their potential influence on responses to O3 exposure.
 3           Information derived from O3 exposure of smokers is limited.  Some degree of
 4     attenuation appears to occur in active smokers (Frampton et al.,  1993), which may be
 5     reversed following smoking cessation (Emmons and Foster, 1991), but available results
 6     should be interpreted with caution.  The possibility of age-related differences in response to
 7     O3 has been explored to some extent since the publication of the previous O3 criteria
 8     document (U.S.  Environmental Protection Agency, 1986).  Young adults historically have
 9     provided the subject population for air pollutant exposure studies. Pulmonary function
10     responsiveness appears to decrease with age, although symptom rates remain similar to
11     young adults (Drechsler-Parks et al., 1987b,  1989, 1990; Bedi et al., 1988; Reisenauer
12     et al., 1988; McDonnell et al., 1993).  The limited information available on the responses of
13     children and adolescents to O3 (McDonnell et al.,  1985a; Avol et al.,  1985a, 1987; Koenig
14     et al., 1987, 1988) does not indicate that children and adolescents are either more or less
15     responsive than young adults.  Of the studies that have investigated  gender differences in
16     responsiveness to O3, some (Lauritzen and Adams, 1985; Horvath et al., 1986; Adams
17     et al., 1987; Drechsler-Parks et al., 1987a,b; Messineo and Adams, 1990) have suggested
18     that women are more responsive to O3 than men.  However, the absence of consistent
19     findings with respect to gender differences indicates that it cannot be concluded that  men and
20     women respond differently to O3.  Comparison of responses across  gender, racial, ethnic,
21     and age groups is complicated by the determination of equivalent exposures. For example,
22     women and children  have smaller lungs than adult men. Thus with  a given exposure
23     concentration, duration,  and ventilation, humans with smaller lungs  will presumably receive a
24     large relative intrapulmonary exposure.  Some attempts have been made to normalize
25     responses according to BSA or lung capacity (e.g., FVC).  The only study in which this
26     factor has been systematically investigated (Messineo and Adams, 1990) found no influence
27     of lung size on the spirometry responses under identical exposure ([Qj], VE, and T)
28     conditions.  Three studies (Fox et al.,  1993; Gerbase et al., 1993; Seal et al., 1993a) have
29     compared pulmonary function responses of women during different phases of the menstrual
30     cycle, but the  results are conflicting. The responses of black and white young adults to
31     various concentrations of O3 have been compared in one study (Seal et al.,  1993b).  The data

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 1      suggested that black males experienced significant decrements in pulmonary function at a
 2      lower concentration of O3 than white males, but there were no differences among the
 3      responses of white males and black and white females.  Thus, the question of ethnic or racial
 4      differences in responsiveness to O3 is inadequately answered, and the available results should
 5      be interpreted with caution. No new studies are available on the effects of heat stress (i.e.,
 6      increased temperature or relative humidity) on O3 responses.  One study (Linn et al., 1988)
 7      suggests that sensitivity to 03 may be related to seasonal variations in ambient
 8      O3 concentrations.  This finding needs to be confirmed if seasonal variations in
 9      responsiveness are to be considered both in design of future studies and in the risk
10      assessment process.  Two studies (Hynes et al., 1988; Adams et al., 1989) have reported that
11      differences in the inhalation route (e.g., oral versus nasal or oronasal) appears to be of
12      negligible importance in the responses of exercising adults to O3 exposure.  Studies of
13      O3 uptake in the upper airway  (Gerrity et al., 1988) confirm  the negligible differences
14      between oral and nasal inhalation (also see Chapter 8).  None of these potential influences on
15      O3 responsiveness (age, gender, race, hormonal fluctuations, smoking, seasonal variations in
16      responsiveness,  and ambient environmental factors) has  been  thoroughly investigated.
17      However, the observation that  healthy older adults appear to  be less responsive  to
18      O3 exposure than  young adults has been  confirmed to the point that it can be considered in
19      risk assessment.  Nevertheless, this does not  fully address the question of age differences
20      because children and adolescents remain inadequately studied.
21
22      7.5.1.6   Extrapulmonary Effects of Ozone
23           It is still believed that O3 immediately reacts on contact with respiratory system tissue
24      and is not absorbed or transported to extrapulmonary sites to any significant degree.
25      A number of laboratory animal studies reported in the previous chapter (Chapter 6) and early
26      studies on human  subjects reported in this chapter suggest that reaction products formed by
27      the interaction of O3 with respiratory system fluids or tissues may produce effects measured
28      outside the respiratory tract—either in the blood,  as changes  in  circulating blood
29      lymphocytes, erythrocytes, or serum, or as changes in the structure or function of other
30      organs,  such as the parathyroid, the heart, the liver, and the  central nervous system.
31      No extrapulmonary effects have been reported to date in other organ systems of O3-exposed

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 1     human subjects, except for limited data indicating that acute (1- to 2-h) exposures with
 2     exercise at concentrations ^0.35 ppm O3 caused transient changes in blood cells and plasma.
 3      The interpretation of all these effects in regards to potential human health risk at ambient
 4     levels of exposure (<0.35 ppm O3) is not clear.  However, the demonstration in this chapter
 5     of an array of inflammatory mediators and immune modulators released at the airway surface
 6     in response to O3 provides a possible mechanism for effects to occur outside of the lung.
 7     Additional studies are needed, therefore, in order to determine if there are any significant
 8     extrapulmonary effects of O3 exposure and at what levels of exposure they might occur.
 9
10     7.5.1.7   Effects of Ozone Mixed with Other Pollutants
11          No significant enhancement of respiratory effects (i.e., more than additive) has been
12     consistently demonstrated for mixtures of O3 with SO2, NO2,  H2SO4, HNO3, or paniculate
13     aerosols, or with multiple combinations of these pollutants.  There is general agreement
14     among studies of simultaneous exposure of healthy adults and  asthmatic adolescents to
15     mixtures of O3 and NO2, SC>2, H2SO4, or HNO3 that pulmonary function responses are not
16     significantly different from those following exposure to O3 alone when compared to studies
17     conducted at the same O3 concentration.  Exposure to high PAN concentrations (i.e.,
18     0.30 ppm) combined with O3 has been reported to induce greater pulmonary function
19     responses than  exposure  to O3 alone (Horvath et al.,  1986), but when the PAN concentration
20     is reduced to the ambient range,  any additional effect of PAN in the mixture appears to be
21     negligible (Drechsler-Parks et al., 1989).
22          In addition to simultaneous exposures to pollutant mixtures, studies of the responses to
23     O3 exposure either preceded or followed by another pollutant have been performed.  To the
24     extent that these exposure sequences mimic real ambient conditions, the results could be
25     useful in the risk assessment process.  Koenig et al. (1990) demonstrated that exposure of
26     allergic (and probably asthmatic) adolescents to O3 and then to SO2 resulted in significant
27     pulmonary function decrements not seen with an O3-O3 sequence or filtered air-SO2
28     sequence.  These results  can also be interpreted in light of the fact that O3 increases
29     nonspecific bronchial  responsiveness and that the increased SO2 responses may simply reflect
30     this increased responsiveness.  Such responses  would  be unlikely in nonatopic healthy
31     adolescents. Other studies (Aris et al., 1991; Hazucha et al.,  1994) have assessed the

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 1     responses to O3 after previous exposure to another pollutant. Aris et al.  (1991) found that
 2     preexposure to water or HNO3 fog appeared to attenuate responses to O3 whereas Hazucha
 3     et al. (1994) observed an increased airway responsiveness after O3 exposure preceded by
 4     NO2 exposure relative to O3 exposure alone.  These findings are intriguing, but must be
 5     replicated before they can be useful for quantitative risk assessment.  Much remains unknown
 6     with reference to responses to  air pollutant mixtures.  Only a limited number of pollutant
 7     combinations and exposure protocols have been investigated, subject groups are small and
 8     may not be representative of the general population.  Few studies have included more than
 9     two pollutants, and most combinations have been evaluated in single studies.  Furthermore,
10     only rarely are endpoints other than pulmonary function and plethysmography measured.
11     Additional studies are, therefore, required to evaluate the relationships between O3 and the
12     complex mix of pollutants found in the ambient environment.
13
14     7.5.2   Field and Epidemiology Studies of Ozone Exposure
15          Individual-level camp and exercise studies provide useful, quantitative information on
16     the exposure-response relationships linking human lung function declines with O3 exposure
17     occurring in ambient air.  Their utility derives largely from  the reliability with which
18     individual exposures can be estimated using outdoor measurements in studies of these kind.
19     Although it usually has not been possible to isolate O3 exposures from other copollutants
20     (e.g., acid aerosols) and environmental factors (e.g., temperature) in the  design of such
21     studies, the available body of evidence now strongly supports a dominant role of O3 in the
22     observed lung function decrements.
23          The most extensive epidemiologic database on pulmonary function responses to ambient
24     O3 comes from camp studies.  Six recent key studies from three separate research groups
25     provide a combined database on  individual exposure-response relationships  for 616 children
26     ranging in age from 7 to 17 years, each with at least 6 sequential measurements of FEVj and
27     previous-hour O3 exposures while attending summer camps  (Avol et at.,  1990; Higgins
28     et al., 1990; Raizenne et al., 1987; Raizenne et al.,  1989; Spektor et al., 1988a; Spektor
29     et al., 1991).  When analyzed together using consistent methods, these data yielded an
30     average relationship between FEVj and previous-hour O3 concentration of  -0.64 mL/ppb
31     (Kinney et al., 1993).  The highest 1-h O3 levels measured in 5 of the 6 studies ranged from

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 1      100 to 160 ppb, with one study reporting concentrations as high as 245 ppb.  Minimum
 2      O3 values ranged from 10 to 60 ppb.  Although the regression results noted above were
 3      based on 1-h O3 levels, exposure in camp studies usually extended for multiple hours.
 4      Because of the high level of correlation between single- and multiple-hour averages in the
 5      studies, these results may therefore, represent, to some extent, the influence of multihour
 6      exposures.  In addition to the camp study results, two key studies involving lung function
 7      measurements before and after well-defined exercise events in adults have yielded exposure-
 8      response slopes of -0.40 and -1.35  mL/ppb (Spektor et al., 1988b; Selwyn et al., 1985).
 9      Ozone concentrations during exercise  events of approximately 0.5 h duration ranged from
10      4 to 135 ppb in these studies. Consistent with chamber studies, there is no clear evidence
11      from individual-level studies for a response threshold for the average population effects of
12      O3 on pulmonary function decline.  However, as  with chamber studies, there is evidence that
13      responsivity varies across individuals.  Thus, pulmonary function decline as a function of
14      ambient O3  exposure for an individual may be either greater than  or less than the mean
15      responses noted above.
16          Recent results of daily-life studies also support a consistent relationship between
17      ambient O3/oxidant exposure and acute respiratory morbidity  in the population.  Respiratory
18      symptoms (or exacerbation of asthma) and decrements in PEFR occur with increasing
19      ambient O3,  especially in asthmatic children (Lebowitz et al., 1991; Krzyzanowski et al.,
20      1992; Thurston et al., 1992a, 1993b).  Concurrent temperature, particulates, H+,
21      aeroallergens, and asthma severity or  medication  status may also contribute as independent or
22      modifying factors.  The aggregate results show greater responses in asthmatic individuals
23      than in nonasthmatics (Lebowitz et al.,  1991; Krzyzanowski et al., 1992), indicating that
24      asthmatics constitute a sensitive group in epidemiologic studies of oxidant air pollution.
25          Recent aggregate population time series studies of O3-related health effects provide
26      relevant evidence of acute responses, even below  a  1-h maximum  of 0.12 ppm.  Emergency
27      room visits, hospital admissions, and mortality have all been examined as possible outcomes
28      of exposure to O3.  In the case of ER visits, the evidence is limited and somewhat mixed
29      (e.g., Bates et al., 1990; Cody et al.,  1992; White et al., 1993), probably due to the fact that
30      ER visits are not a very well defined endpoint because they are dependent upon the patient to
31      decide whether or not to attend the ER. Mortality is, however, a well-defined endpoint

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 1     (though cause of death is not nearly so certain), and two new, well-designed studies indicate
 2     a significant association between O3 and total morality in both Los Angeles, CA, and
 3     New York City, NY, even after controlling for the potentially confounding effects of
 4     temperature and paniculate matter (Kinney and Ozkaynak, 1991, 1992).  These studies
 5     suggest an effect on the order of 0.4 to 0.8 total deaths/day per 100 ppb O3 per million
 6     persons.  Both cities experienced peak 1-h maximum O3 concentrations above 0.2 ppm
 7     during their respective study periods.  At  lower concentrations, over a shorter time span, and
 8     using more conservative statistical methods, a third recent study (Dockery et al., 1992) did
 9     not detect a significant O3 association with mortality.  The strongest and most consistent
10     evidence of O3 effects, both above and  below 0.12 ppm O3, however, is provided by the
11     multiple studies that have been conducted  over the last decade on summertime daily hospital
12     admission for respiratory causes in various locales in eastern  North America (Bates and
13     Sizto,  1983, 1987,  1989; Thurston et al.,  1992b,  1993a,b; Lipfert and Hammerstrom, 1992;
14     Burnett et al.,  1993). These studies have consistently shown  that O3 air pollution is
15     associated with an increased incidence of  admissions,  accounting for roughly one to three
16     excess respiratory hospital admissions per hundred ppb O3 per million persons.   This
17     association has been shown to remain even after statistically controlling for the  possible
18     confounding effects of temperature and copollutants (e.g., H   , SO4, and PM10), as well as
19     when considering only concentrations below 0.12 ppm O3. Furthermore, these results imply
20     that O3 air pollution can account for a substantial portion of summertime hospital admissions
21     for respiratory causes on the most polluted days.   Overall, the aggregate population time
22     series  studies considered in this chapter provide strong evidence that ambient exposures to
23     O3 can cause significant exacerbations of preexisting respiratory disease in the  general public
24     at concentrations below 0.12 ppm O3, and suggest that  higher O3 levels may also cause a
25     much  smaller but statistically significant increase in total daily mortality.
26           Studies of chronic health effects that may relate to long-term exposure to  ambient
27     pollutants still have not provided unequivocal evidence  in support of respiratory or other
28      health effects that result directly from chronic O3  exposure.  However, the aggregate
29      evidence to date suggests that chronic O3 exposure could  be  responsible for health effects in
30      exposed populations, and research efforts should continue to  look for effects at O3 levels
31      <£0.12 ppm.

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 1          The most useful set of data has been provided by the AHSMOG studies (Hodgkin
 2     et al.,  1984; Euler et al., 1987, 1988; Abbey et al.,  1991a,b, 1993).  These studies have
 3     provided the most refined measures of chronic exposure to date (including adjustment for
 4     quality of the monitoring data as determined by  distance of monitoring sites from subject
 5     residences, topography, time spent indoors, and time spent at work).  The most consistent
 6     effects that can be attributable, in part, to O3 relate to an  increase in 10-year cumulative
 7     incidence of asthma (KR =  2.07 for each 1,000 h above 10 pphm) and an increase in asthma
 8     severity.  Unfortunately, for the entire set of studies, the colinearity between O3 and TSP
 9     reduces the confidence that effects can be attributed to O3 alone, O3 in combination with the
10     paniculate fraction of ambient pollution, or the combination of the two. Some support for an
11     effect  on persons with asthma also can be derived from a  recent Canadian study (Stern et al.,
12     1993)  that demonstrated non-statistically significant 6.6 and 6.5% reductions in FEF25-75%
13     and Vmax50% for people living in Ontario relative to those in Saskatchewan.  Again,
14     however,  the effects of O3 are impossible to disentangle from the other contributors such  as
15     the acid summer haze that characterizes the United States  east of the Mississippi River.
16          Most of the other studies that purport to demonstrate effects that relate to O3 exposure
17     are sufficiently limited in one or another of their design or analysis features to minimize their
18     usefulness for the setting of standards.  However, most of the studies  are compatible with
19     regard to  low-level effects on the upper and lower respiratory tract. In this latter regard,  the
20     overall body of studies of health effects that relate to chronic exposure to ambient 0)3 are
21     quite consistent with observations summarized in the previous section.
22          A major issue that virtually has been ignored in epidemiologic studies of chronic
23     O3-related health effects relates to the effect of measurement error (both in terms of ozone
24     exposure and important covariates) on the estimates of the magnitude of the effects that have
25     been attributed to O3.  Unfortunately,  given current uncertainties about the relevant metrics
26     by which  chronic O3 exposure should  be evaluated and the lack of an  accepted methodology
27     for the assignment of "lifetime" exposures and the highly  variable measurement-error
28     structure that is likely to be  associated with the important  covariates (e.g., cigarette smoking,
29     competing exposures in the work environment, lifetime activity patterns), it is difficult to
30     provide estimates either for the direction or magnitude of  any biases that might exist for
31     current estimates of effects on lung function of specific respiratory disease or symptom

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1     outcomes. This issue should be addressed specifically in future studies, with specific
2     attention given as to how data derived from personnel monitoring of O3 exposure can be
3     utilized for the assignment of more long-term exposures.
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27
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 i      8.  EXTRAPOLATION OF ANIMAL TOXICOLOGICAL
 2                                DATA TO HUMANS
 3
 4
 5      8.1   INTRODUCTION
 6           A full evaluation of the health effects of ozone (03) requires an integrated interpretation
 7      of human clinical, epidemiological, and animal lexicological studies. Each of these three
 8      research approaches has inherent strengths and limitations.  Animal toxicological data are
 9      valuable because they provide concentration- and duration-response information on a fuller
10      array of effects and exposures than can be studied in humans.  However, historically, use of
11      animal toxicological data have been limited because of difficulties in quantitative
12      extrapolation to humans. Recent advances in the state-of-the-art of extrapolation have
13      reduced several uncertainties, as will be discussed in this chapter.
14           Qualitative animal-to-human extrapolation is generally accepted because O3 causes
15      similar types of effects in several animal species, from mouse to  nonhuman primate
16      (Chapter 6). Also when similar endpoints (e.g., inflammation and pulmonary function) have
17     been examined in O3-exposed animals and humans, similar effects are observed. However,
18      quantitative extrapolation is the goal and is more controversial. That is, if a certain exposure
19      causes a specific effect in animals, what exposure is likely to cause that same effect in
20     humans? Such an extrapolation requires an integration of dosimetry and species sensitivity.
21      Dosimetry is defined as the dose delivered to a site in the respiratory tract.  As can be seen
22     in Section 8.2, substantial information is available on dosimetry in several species, including
23     humans.  Dosimetric studies that are referenced in the earlier O3  criteria document (U.S.
24     Environmental Protection Agency, 1986) are only briefly summarized here; newer research is
25     the focus.  Species sensitivity, discussed in Section 8.3, refers to the sensitivity of a specific
26     species to the delivered dose. For example, even if the same dose of O3 were delivered to a
27     specific respiratory tract site in rats and humans, differences in species sensitivity to that
28     dose are likely because of variations in defense mechanisms  and perhaps other factors.
29     Section 8.3 also provides a more holistic approach to extrapolation by quantitatively
30     comparing exposure-response data obtained in animals and humans.  The final Section 8.4 is


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 1     intended to draw the forgoing information together, reaching conclusions about the potential
 2     for human health effects based on animal studies.
 3          Although this chapter focuses on animal-to-human extrapolation, dosimetric studies can
 4     also be used to elucidate interpretations of the human studies described in Chapter 7.  For
 5     example, knowledge of dosimetry in humans as related to age and exercise can enhance
 6     understanding of human susceptibility factors.
 7
 8
 9     8.2  OZONE DOSIMETRY
10     8.2.1   Introduction
11          Dosimetry refers to measuring or estimating the quantity or rate of a chemical absorbed
12     by target sites within the respiratory tract. The compound most directly responsible for toxic
13     effects may be the inhaled gas, 03, or its chemical reaction products. Complete
14     identification of the actual toxic agents and their integration into dosimetry is a complex issue
15     which has not been resolved. Thus, most dosimetry investigations are concerned with the
16     dose of the primary inhaled chemical.  In this context, a further confounding aspect can be
17     the units of dose (e.g., mass retained per breath, mass retained per breath per body weight,
18     mass retained per breath per respiratory  tract surface area). That is, when comparing dose
19     between species, what is the relevant measure of dose? This question has not been
20     answered; units are often dictated by the type of experiment and/or by a choice made by the
21     investigators.
22          Experimental and theoretical (dosimetry modeling) studies are used to obtain
23     information on dose. Experiments have been carried out to obtain direct measurements of
24     absorbed O3 in the total respiratory tract (RT), in the upper respiratory tract (URT; region
25     proximal to the tracheal entrance), and in the lower respiratory tract (LRT; region distal to
26     tracheal entrance);  however, experimentally obtaining dosimetry data in smaller regions or
27     locations, such as specific airways or in  the centriacinar region (junction of conducting
28     airways and gas exchange region) where lesions due to O3 occur (see Section 6.2.4), is
29     extremely difficult.  Nevertheless, experimentation is important for determining dose, making
30     dose comparisons between  subpopulations and between different species, assessing


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 1     hypotheses and concepts, and in validating mathematical models that can be used to predict
 2     dose at specific respiratory tract sites and under more general conditions.
 3          Theoretical studies are based on the use of mathematical models developed for the
 4     purposes of simulating the uptake and distribution of absorbed gases in the tissues and fluids
 5     of the respiratory tract. Because the factors affecting the transport and absorption of gases
 6     are general to all mammals, a model that uses appropriate species and/or disease-specific
 7     anatomical and ventilatory parameters can be used to describe absorption in the species and
 8     in different-sized, aged, or diseased members of the same species.  Importantly, models may
 9     also be used to make interspecies and intraspecies dose comparisons, to compare and
10     reconcile data from different experiments, to predict dose in conditions not possible or
11     feasible experimentally, and to better understand the processes involved in toxic effects.
12
13     8.2.2    Summary of 1986 Review of Experimental and Theoretical
14              Dosimetry
15          Presented is a summary of the more relevant experimental and theoretical dosimetry
16     studies contained in the previous O3 criteria document (U.S. Environmental Protection
17     Agency, 1986).  The reader is referred to this document for completeness.
18          Experiments on the nasopharyngeal removal of O3 in laboratory animals suggested that:
19     (1) the fraction of O3 uptake depends inversely on flow rate (Yokoyoma and Frank, 1972),
20     (2) uptake was greater for nose than mouth breathing  (Yokoyoma and Frank, 1972), and
21     (3) tracheal and chamber concentrations were linearly related (Yokoyoma and Frank, 1972;
22     Miller et al., 1979).  Only one investigation measured uptake by the LRT, finding 80 to 87%
23     uptake by the LRT of dogs (Yokoyoma and Frank,  1972).  At the time, there were no
24     reported results for human URT or LRT uptake.  With the exception of two relatively crude
25     studies by Clamann and Bancroft (1959) and Hallett (1965), there were no data on O3 uptake
26     in humans at the time of the earlier Criteria Document (U.S. Environmental Protection
27     Agency, 1986).
28          Several mathematical dosimetry models had been developed to simulate the processes
29     involved in O3 uptake and to predict O3 uptake by various regions and sites within the
30     respiratory tract. The model of Aharonson et al. (1974) had been used to analyze
31     nasopharyngeal uptake data.  Applied to O3 data, the  model indicated that the average mass

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  1      transfer coefficient of this region and the mass retained increased with increasing air flow,
  2      but the percent uptake decreased.
  3           Models had been developed to simulate LRT uptake (Miller et al., 1978, 1985).  The
  4      models were very similar in their treatment of O3 in the airways and airspaces and hi their
  5      use of morphometric data to define the dimensions of the air compartments and liquid lining.
  6      Both of the 1978 and the 1985 models of Miller and co-workers took into account reactions
  7      of O3 with constituents of the liquid lining. However, the models of Miller et al.  differed in
  8      their treatment of chemical reactions in the liquid lining, and the later model included
  9      transport and chemical reactions within tissue and blood,  whereas the first Miller et al. model
10      did not (an instantaneous reaction at the liquid-tissue interface was assumed, so that the
11      O3 concentration here was defined as zero).  In both models, tissue dose was  defined as the
12      O3 flux to the liquid-tissue interface.  The models of Miller et al. predicted O3 tissue dose to
13      be relatively low in the trachea, to increase to a maximum in or near the centriacinar region,
14      and then to decrease distally. This was characteristic for both the animal and the human
15      simulations (Miller et al., 1978, 1985).
16           Prior to 1986, there were no experimental results that were useful in judging the
17      validity of the modeling efforts. However, a comparison of the results of Miller and
18      co-workers with morphological data (that show the centriacinar region to be most effected by
19      O3; see Section 6.2.4) indicated qualitative agreement between the site of predicted maximum
20      tissue dose and the site of observed maximum morphological damage in  the pulmonary
21      region.
22
23      8.2.3   Experimental Ozone Dosimetry Data
24      8.2.3.1   Introduction
25           Models of Oj uptake in the respiratory tract have reached a scale of sophistication that
26      provide some highly specific predictions regarding the location and magnitude of O3 dose.
27      However, before these models can be exploited to their fullest degree in extrapolating dose
28      within and between species, validation of the models with experimental data is essential.
29      This section will review the experimental data base upon which the modeling of
30      O3 dosimetry is both based and  validated.  This will help facilitate discussion of the models
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 1     themselves in subsequent sections. Table 8-1 provides a summary of all post-1986
 2     experimental O3 dosimetry studies.
 3
 4     8.2.3.2  Animal (In Vivo) Ozone Dosimetry Studies
 5          The model predictions of Overton et al. (1987) based on the original model of Miller
 6     et al. (1985) provided specific predictions about the regional and total uptake efficiencies of
 7     O3 in laboratory rats.  It was therefore necessary to test these predictions with actual data.
 8     The  first data on total RT uptake of O3 in rats were obtained by Wiester et al. (1987).
 9     Ozone uptake was measured in 30 awake unanesthetized Sprague-Dawley rats receiving a
10     nose-only exposure.  A rat was situated within a plethysmograph that could continuously
11     monitor the animal's breathing pattern. Air with O3 flowed by  rats' noses at 1,200 mL/min
12     for 1 h at a concentration of 0.3, 0.6, or  1.0 ppm O3. Determination of RT 03 uptake was
13     determined by mass balance. The uptake was the difference in mass in the upstream air and
14     the air downstream of the rat. Total RT O3 uptake efficiency of approximately 40% was
15     measured and was independent of O3 concentration.  During data acquisition, the animals had
16     an average tidal volume (VT) of about 2.8 mL, an average breathing frequency (fB) of about
17     150  breaths per minute (bpm), and an average minute ventilation (Vg) of about 400 mL/min.
18     This study was followed by another  (Wiester et al., 1988), in which total RT uptake  was
19     measured in three strains of rats and in the guinea pig.  Specifically, Fischer 344, Sprague-
20     Dawley,  Long Evans rats, and Hartley guinea pigs were all exposed for 1 h to 0.3 ppm O3;
21     F-344 rats also received a 0.6 ppm exposure. Uptake was measured as in the previous
22     experiment (Wiester et al., 1987). Total  RT uptake of O3  was  species-independent and
23     averaged 47%. This was higher than in the previous study because a different calculation
24     method for fractional uptake was used. Wiester et al. (1987) corrected all flows used in
25     uptake calculations for body temperature and relative humidity.  In their later work,
26     however, they found that this correction was not warranted, resulting in slightly higher
27     computed O3 uptake efficiency (Wiester et al., 1988).
28          All rats in the Wiester et al. (1988)  study had fBs between 112 to  132 bpm, VTs
29     between 2.4 to 2.8 mL, and VEs between 299 and 364 mL/min. The guinea pigs had a
30     VT of 2.4 mL (not different from rats), an fB of 77 bpm, and a VE of 188 mL/min.  As in
31     the first experiment, exposure concentration did  not affect uptake.

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                                      TABLE 8-1 (cont'd).  EXPERIMENTAL STUDIES ON OZONE DOSIMETRY8
                                   Species
             In Vivo/In Vitro        (Strain)
     Uptake
                     Breathing Patterns
                          Results
Reference
             In vitro
                              Pig
                              Sheep
Trachea
                  Vjjjj = 50 mL/s to      Unidirectional 03 uptake efficiencies of trachea decreased with
                  200 mL/s              increasing flow from 0.5 to 0.15 for the sheep and 0.12 for the pig.
                                        Mass transfer coefficients were generally independent of flow.
                                                                                                    Ben-Jebria et al. (1991)
             In vitro
                              N/A
                                                N/A
                                                                  N/A
                                                                                        Ozonolysis studies on various unsaturated fatty acids, rat
                                                                                        erythrocyte ghost membranes and rat BAL. Dominant processes are
                                                                                        the production of aldehydes and peroxides due to reactions between
                                                                                        03 and olefms.
                                                                                                    Pryoretal. (1991)
             In vivo
                              Human
20-200 mL depth    VT = 500 mL
into respiratory
tract
                                                                  Vtas = 250 mL/s
                                        Uptake efficiencies by measuring recovery of 03 boluses delivered
                                        at 20 mL increments into lung to depth of 200 mL.  At deepest
                                        depth, only 6% of 03 could be recovered. Ozone uptake by
                                        conducting airways larger than predicted by Miller et al. (1985).
                                                                                                                                                    Hu et al. (1992b)
00
             In vivo
                              Human
                                                20-200 mL depth
                                                into respiratory
                                                tract
                  VT = 500 mL
                  Vj^ = 150, 250, 500,
                  750, 1,000 mL/s
                                        Same technique as Hu et al. (1992b), but investigating flow effects.
                                        Increasing flow caused marked shift of delivered Oj toward the
                                        periphery of the conducting airways (i.e. , the greater the inspiratory
                                        flow the greater the amount of 03 delivered to the lung periphery),
                                        where it is available for absorption.  Mass transfer coefficients in
                                        upper airways independent of flow, but in conducting airways they
                                        increase proportional to flow. Lung liquid lining mass transfer
                                        coefficient computed to be 1 .4 cm/s in the URT, falling to
                                        0.17 cm/s in the respiratory airways. Reaction rate constant
                                        between 03 and the lung liquid lining was computed as 7.3 X
                                        10  s"  in the URT, falling to 8.2 X 10  s"  in the distal conducting
                                        airways.
                                                            Hu et al. (1993)
            In vivo
                              Human
                                                20-200 mL depth
                                                into respiratory
                                                tract
                  VT = 500 mL
                  Vj^ = 250 mL/s
                                        Comparison of 03 bolus uptake between oral and nasal routes.
                                        Nose was found to be 30% more efficient at removing 03 from air
                                        stream than the mouth.
                                                            Ben-Jebria et al. (1993)
            In vitro
                              Rat
                              (Sprague-Dawley)
Lung
                  VT  = 2.71 mL
                  fB = 50 - 103 bpm
                  FRC = 4 and 8 mL
Perfused and non-perfused rat lungs ventilated with 1 ppm 03.
Uptake efficiency of lungs  dropped from 95% at 50 bpm to about
50% at 103 bpm. No change in uptake efficiency when lungs
inflated from FRC = 4 mL to 8 mL.
Postlethwait et al. (1993)
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  1           These data are in disagreement with the model predictions of Overton et al. (1987) who
  2      made predictions of O3 uptake in two different rat anatomical lung models (Kliment, 1973
  3      versus Yeh et al., 1979).  They conducted simulations in both anatomical models, varying
  4      fB and VT at fixed VE; they also  varied functional residual capacity (FRC) as a fraction of
  5      total lung capacity (TLC).  The Kliment anatomical lung model gave consistently high
  6      predictions for uptake when compared with actual data.  The predictions using the Yeh et al.
  7      (1979) model came closer. Total RT uptakes (not including the head) for the Yeh et al.
  8      (1979) model were predicted to range between 46 and 60% at fB = 154 bpm and
  9      VT  = 1.25 mL and between 70 and 80% for fB = 81 bpm and VT = 2.4 L.  However,
10      when the fact that these predictions do not include the head of the animal is considered, it is
11      evident that the model predictions overestimate the total RT uptake in rats. The question is
12      whether the measurements are accurate or whether there is a problem with the model
13      formulation. The data of Postlethwait et al.  (1993) (see below) suggested that the data of
14      Wiester et al. (1987, 1988) may be reasonable.  Their data in the excised rat lung suggested
15      a clear inverse dependence of lung uptake on fB. At a VT of 2.51 mL,  the O3 uptake
16      efficiency of the excised lung fell from nearly unity at fB = 50 bpm to almost 50%  at
17      fB = 103 bpm.  For extrapolation purposes, a key question here is what fB should be
18      considered as the normal resting fB of a rat.   Although Wiester et al. (1987, 1988) allowed
19      their rats to acclimate to the plethysmograph by monitoring fB and only  began uptake
20      measurements after fB had plateaued at a minimum, it is still uncertain whether frequencies
21      of 120 to 150 bpm are reasonable. In a summary of studies of the pulmonary function of
22      rats  in response to O3, Tepper et  al. (1993) found typical breathing frequencies of 100 bpm.
23      Although the models appear to overestimate the O3 uptake efficiency of  the rat respiratory
24      tract, the discrepancy is not large, and the near agreement indicates that the O3 dosimetry
25      models have predictive capability.
26           In addition to data on the total RT O3 uptake efficiency, in vivo data on regional
27      O3 dosimetry in animal models have begun to emerge.  Hatch and Aissa (1987) and Aissa
28      and  Hatch (1988) first described a method to measure O3 uptake in animals by exposing them
29      nose-only,  while in a plethysmograph, to O3 enriched with 18O, a stable isotope of oxygen.
30      After exposure, bronchoalveolar lavage fluid (BAL) and respiratory tract tissue were assayed
                  1 R
31      for excess   O using isotope ratio mass spectrometry.  One problem with this technique is

        December  1993                           g_9       DRAFT-DO NOT QUOTE  OR CITE
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 1     that all of the absorbed 18O cannot be accounted for. The technique used by Hatch and
                 _                               * O     | Q
 2     Aissa (1987) involves the detection of excess   O in  O3 reaction products after tissue
 3     pyrolysis. Thus, 18O3 that is degraded to H218O or 18O2 is lost and cannot be detected in
 4     dry tissue.  Eight male F-344 rats (four previously exposed chronically to O3 for 1 year:
                                                                                  1 C
 5     0.06 ppm baseline with 1 h spikes of 0.25 ppm) were exposed for 2 h to 1 ppm of  O-
 6     enriched O3 (Hatch et al., 1989). During exposure, the rats breathed at 150 bpm, a VT of
 7     2.05 mL, and a VE of 290 mL/min.  After exposure, the lung, trachea, and head were
 8     separately analyzed for   O.  Overall, the animals took up 54.3% of inspired O3.  Although
 9     this value of O3 uptake efficiency is higher than that found by Wiester et al. (1987, 1988),
10     considering the fact that the coefficient of variation for O3 uptake efficiency measurements is
11     around 20% in all studies, the result  of Hatch and Aissa (1987) is consistent with the data of
12     Wiester et al. (1987,  1988). Of the O3 taken up by the animals, 49.6% was taken up by the
13     head, 6.7% by the larynx/trachea, and 43.6% by the lungs. By assuming equal uptake
14     efficiencies by compartments on inspiration and expiration, inspiratory uptake of O3 by these
15     regions  was computed. It was determined that the rat nasopharynx  had an inspiratory
16     efficiency of 17.4% and that the larynx/trachea removed 2.7% of the remaining O3.
17          This technique has recently been extended to humans.  Hatch et al. (1993) showed that
18     when human subjects were exposed to 0.4 ppm 18O3 while intermittently exercising at VE =
19     60 L/min for 2 h, the amount of recovered 18O in lavagable cells indicated that the humans
20     received 3-4 times the O3 dose than rats exposed to 0.4 ppm for 2 h at rest. This ratio was
21     derived from the concentration of 18O in the BAL constituents of both rats and humans.
22     Consequently,  to compare absorbed 18O3 doses between rats and humans using BAL requires
                                                                                  18
23     the assumption that the amount of lavagable cell membrane available to react with  O3 is
24     comparable between the two species.  The difference between rats and humans could be
25     accounted for by the fact that the humans were exercising whereas the rats  were not.
                                                   1 R
26     However, as was noted above, not all absorbed  O3 can be accounted for.
27
28     8.2.3.3   Animal (In Vitro) Ozone Dosimetry Studies
29          The use of whole, intact animals to study O3  uptake is needed to ascertain the actual
30     amounts of O3 absorbed. However,  it is also important to understand some of the more
31     fundamental processes governing O3 uptake, such as the biochemistry of Oyiiquid and

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 1      O3/tissue interactions to determine chemical reaction rates essential to O3 dosimetry models.
 2      Furthermore, the use of intact animals does not allow more precise detenninations of the role
 3      of physiological parameters on O3 uptake.  For this reason, there have been some limited
 4      attempts at utilizing animal tissue explants and whole lungs to study O3 uptake.
 5           Ben-Jebria et al. (1991) studied O3 uptake by the trachea of sheep and pigs to
 6      investigate mass transfer coefficients.  Ozone boluses of 1 ppm were passed through excised
 7      tracheae. Tracheae were obtained from a slaughter house 0.5 to 2  h after slaughter, and
 8      although they were kept coated with physiologic saline, they were not maintained at body
 9      temperature, possibly resulting in underestimation of in vivo uptake.  The lengths and
10      diameters of the pig  tracheae were not too different from human tracheal dimensions.  The
11      flow dependence of mass uptake and the mass transfer coefficient K were determined for
12      flows between 50 mL/s and 200 mL/s. Uptake efficiencies in the pig decreased with
13      increasing flow from about 0.5 to 0.12 and in the sheep decreased from about 0.5 to 0.15.
14      Mass transfer coefficients were generally independent of flow (K = 0.5 cm/s in pigs  and
15      0.35 cm/s in sheep), indicating the lack of dependence of uptake on gas-phase diffusion
16      processes. This contrasts with the conclusion of Aharonson et al. (1974) for the nasopharynx
17      of dogs  where they observed that the slight inverse dependence of uptake on flow observed
18      by Yokoyama and Frank (1972) leads one to conclude that the mass transfer coefficient for
19      the nasopharynx of the dog should increase with flow, suggesting a role of the boundary
20      layer in  limiting diffusion of O3 to the wall of the nasopharynx.  The different geometries of
21      a trachea and a nasopharynx may account for the differing observations.
22           A significant feature of the Ben-Jebria et al. (1991) study was the use of a rapidly
23      responding O3 analyzer.  In order to conduct their O3 uptake studies, Ben-Jebria and  Ultman
24      (1989) and Ben-Jebria et al. (1990) developed a rapidly responding O3 analyzer.  The
25      analyzer relies on reaction of O3 with alkenes such as ethylene, propylene, cyclohexane, etc.
26      Ten alkenes were tested.   Ninety percent step-response  times of 130 to  540 ms were achieved
27      with varying degrees  of linear response with O3 concentration.  The authors concluded that
28      the best  alkene was 2-methyl-2-butene, with optimum 10 to 90% responses of 110 ms and
29      minimum detectable limits of O3 of 18 ppb. Interference with CO2, however, was found,
30      requiring measurements of CO2 to correct the analyzer  response.
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 1          Postlethwait et al. (1993) used an isolated rat lung preparation to investigate the effects
 2     of vascular perfusion, inspired dose, temperature, and distal lung surface area on
 3     O3 absorption by the LRT. Vascular perfusion had little or no effect on uptake efficiency of
 4     O3.  When the lung was exposed to 1 ppm O3 and ventilated with a VT of 2.71 mL, FRC of
 5     4 mL,  and fB of 50 bpm, uptake efficiency was 95%.  As fB increased with fixed VT, uptake
 6     efficiency began to drop, reaching nearly 50% at fB of 103 bpm.  When the lung temperature
 7     was dropped from 37 to 25 °C, uptake efficiency dropped from 95 to 85% at 50 bpm.  This
 8     drop was exploited to investigate other factors (such as flow, volume, and lung surface area)
 9     governing uptake because it moved RT uptake further below 100%.  The observation of a
10     dependence of uptake on temperature indicates that uptake efficiency is chemical reaction-
11     dependent, thus possibly coupling uptake to reaction product formation.
12          Another interesting result from this study was the lack of dependence of uptake on
13     FRC.  When FRC was doubled from 4 to 8 mL at 25 °C, fractional O3  uptake was
14     unchanged.  This latter result suggests that O3 uptake is virtually complete by the time
15     O3 reaches the alveolar spaces of the lung.  Otherwise it would have been expected that the
16     uptake efficiency would have risen  with increased FRC.
17          To further investigate the reactions of 03 with the lung, Pryor et al. (1991) performed
18     ozonolysis studies of various unsaturated fatty acids (UFAs), rat erythrocyte ghost
19     membranes, and rat BAL. These studies demonstrated significant production of hydrogen
20     peroxide and aldehydes and that production of hydrogen peroxide was primarily due to
21     reactions between O3 and olefins.  The authors concluded that the reaction of O3 with UFAs
22     hi the lung fluid lining and cell membranes produce hydrogen peroxide and aldehydes that
23     may be important mediators in the  toxicity of O3.  The quantitative results of these studies
24     led Pryor (1992) to hypothesize about the degree to which O3 reacts with the liquid lining of
25     the lung and with lung tissue.  A simple model calculation was performed using the Einstein-
26     Smoluchowski equation to estimate the half-life of O3 in bilayers and cell membranes.  Pryor
27     (1992) concluded that a substantial fraction of O3 reacts in the bilayer and that only  in
28     regions of the lung where the lung lining fluid layer is less than 0.1 jum will O3 penetrate to
29     tissue and only then  will O3 react in cell membranes before penetrating further.  The overall
30     conclusion is that the toxic effects  of O3 may be mediated not just by O3 directly but by
31     reactive intermediates such as aldehydes and hydrogen peroxide.  This would bring into

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 1      question what the relevant dose of O3 is. Is it the total dose, the dose to the liquid lining,
 2      the tissue dose, or the dose of reactive intermediates delivered to tissue?
 3
 4      8.2.3.4   Human Ozone Dosimetry Studies
 5           Significant progress has been made since the previous criteria document (U.S.
 6      Environmental Protection Agency, 1986) in the area of human Oj dosimetry.  Studies have
 7      been conducted defining total and regional RT uptake; the dependence of uptake on
 8      physiological parameters; and the role of uptake in modulating response.
 9           Gerrity et al. (1988) reported on measurements of O3 uptake by the extrathoracic
10      airways (airways proximal to the posterior oropharynx) and intrathoracic airways (airways
11      distal to the posterior oropharynx) in 18 healthy, young male volunteers.  Ozone uptake was
12      measured by placing a small polyethylene catheter through the nose and positioning the distal
13      tip in the posterior oropharynx.  Breath-by-breath samples of O3 were collected, and the peak
14      plateau concentrations were compared with chamber concentrations. The effects on uptake of
15      O3 concentration (0.1, 0.2, 0.4 ppm), fB (12 and 24 bpm at fixed VT), and mode of
16      breathing (oral, nasal, and oronasal) were tested. The O3 analyzer had a moderately rapid
17      response with a 90%  response time of 700 ms.  Inspiratory VT ranged between 754 mL and
18      848 mL; mean inspiratory flow at 12 bpm was 350 mL/s; at 24 bpm it was 634 mL/s. The
19      authors measured extrathoracic uptake efficiency of O3  on inspiration to be approximately
20      40%, with intrathoracic uptake efficiency (inspiration plus expiration) being approximately
21      92%.  They essentially found no effect of O3 concentration on uptake (intrathoracic uptake
22      was significantly higher at 0.4 ppm, but the difference was very small). They did find that
23      both intrathoracic and extrathoracic uptake decreased with increasing fB (at fixed VT), falling
24      by about 7 % for extrathoracic uptake and about 3 % for intrathoracic uptake when
25      fB increased from  12  bpm to 24 bpm.  The finite response time of the analyzer may have
26      affected the results at the 24 bpm fB by overestimating extrathoracic uptake and
27      underestimating intrathoracic uptake.  However, because uptake was defined relative to
28      plateaus of concentration, the response time of the analyzer was adequate to reach a plateau
29      in the 1.2 s inspiratory time at 24 bpm. It is important to note here that when utilizing the
30      data from this study to compare with other studies and models, the uptake efficiencies
31      measured here are comparable to steady-state unidirectional measurements of uptake.

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 1     Another feature is that Gerrity et al. (1988) consistently measured a small, non-zero plateau
 2     of O3 on expiration.  This plateau is not consistent with the suggestion from models of
 3     O3 uptake and the data of Postlethwait et al. (1993) (nor with later work of Gerrity et al.
 4     [1993b] as presented below) that no O3 should be washed out from lung volumes beyond the
 5     conducting airways.  This observation of Gerrity et al. (1988) may have been an artifact of
 6     how O3 was measured by sampling from the posterior oropharynx.  There may have been
 7     entrainment of O3 in the pharyngeal airspaces that was washed out after expiration of dead
 8     space air. Regardless, the concentration of O3 exhaled from  the alveolar phase of washout
 9     was very low.
10          One of the most startling results from the work of Gerrity et al. (1988) was the finding
11     that there was only a small,  but statistically significant, difference between uptake by the
12     nose and the mouth.  The mouth had  about a 10% (in relative terms) greater uptake
13     efficiency than the nose.  The combined oronasal passage had an uptake efficiency greater
14     than the nose by another  8%.  This suggests that persons who nasally breathe are at no less
15     risk than persons who oro-nasafly breathe.  Adams et al. (1989) investigated this possibility
16     by comparing functional responses in subjects acutely exposed to O3 while either orally or
17     oro-nasally breathing.  Healthy subjects were exposed on five separate days  to 0.4 ppm O3.
18     In the first four exposures,  subjects were exposed  by  facemask (with or without noseclip) for
19     30 min at an exercise level of 75 L/min or for 75  min at exercise level of 30 L/min.  The
20     fifth exposure was for 30 min at 75 L/min, with exposure through a mouthpiece.  There
21     were no differences in pulmonary functional response (forced expiratory volume at 1 s
22     [FEVJ or forced vital capacity [FVC]  or forced expiratory flow [FEF25_75%]) with facemask
23     exposure among all experimental groups (i.e., no noseclip, VE, or time effect). Pulmonary
24     function response was, however, greater with a mouthpiece.  Adams et al. (1989)  speculated
25     that the greater response  with the mouthpiece was due to C^  scrubbing by the facemask or by
26     facial hair.  It may also have been due to different oral configurations imposed by a
27     mouthpiece.  Hynes et al. (1988) also investigated whether functional responses were affected
28     by the mode of breathing.  Healthy subjects were  exposed to 0.4 ppm O3 for 30 min in an
29     exposure chamber.  On two different occasions, each subject breathed either through the  nose
30     or the mouth exclusively. There was no difference in pulmonary function response between
31     these two routes of exposure. Taken together, the studies of Adams et al. (1989) and Hynes

       December 1993                           8-14        DRAFT-DO NOT QUOTE OR CITE
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 1     et al. (1988) are consistent with the observations of Gerrity et al.  (1988) on the equal
 2     efficiency of all routes of breathing for extrathoracic O3 scrubbing.
 3          This study was followed by another study (Gerrity and McDonnell,  1989; Gerrity
 4     et al.,  1989, 1993a) in which the relationship between O3 uptake and functional response was
 5     investigated.  Healthy subjects  were exposed to 0.4 ppm O3 for 1 h while continuously
 6     exercising at 40 L/min.  Ozone uptake was measured at the beginning and at the end of
 7     exposure while the subjects were still exercising,  using the technique of Gerrity et al. (1988).
 8     In contrast to the work of Gerrity et al. (1988), uptake was computed in this study by
 9     integrating concentration times flow instead of using peak plateau measurements.  Also, in
10     this work, the 90%  response time of the analyzer was 1.2 s (compared with 0.7 s in the
11     previous work). The authors found that about 40% of the inspired O3 was taken up by the
12     URT (i.e., the same as the extrathoracic airways described in Gerrity et al., 1988) during
13     inspiration and that  this  did not change  during exposure. Total RT uptake efficiency was
14     approximately 80%, and it did not change during exposure. However, LRT (i.e., the
15     intrathoracic airways described in Gerrity et al., 1988) uptake efficiency fell during exposure
16     from 68 to 62% and was correlated with the O3-induced fall in VT (VT fell from 1,650 to
17     1,239 mL; fB increased  from 25.2 to 34.8 bpm; inspiratory flow fell from 1,506 to
18     1,357 mL/s; VE slightly increased from 40.8 to 40.9 L/min), suggesting that the
19     VT reduction may have  a protective effect on dose delivered to the periphery of the lung.
20     It is not likely that the finite analyzer response time affected uptake measurements.  Evidence
21     of this is the lack of dependence of URT uptake on changes in VT or f.  The low values for
22     uptake in the LRT may  have been due to artifact from the relatively slow response time of
23     the analyzer which was  approximately equal to inspiratory and expiratory times.  As a check
24     on their results, the authors compared their data with the previous work of Gerrity et al.
25     (1988) by computing uptake by the original technique using peak plateau concentrations.
26     When that was done,  the URT uptake efficiencies were  17%  and 22% at  the beginning and
27     end of exercise, respectively; the LRT efficiencies were 96% and 92% at the beginning and
28     end of exercise. The URT change computed this way was not significant, but the LRT
29     efficiency drop was.  When looked at in this manner, the data from this experiment  are
30     consistent with those from the previous experiment.
       December 1993                           8-15       DRAFT-DO NOT QUOTE OR CITE
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 1          Because there is a need to compare human O3 uptake data with rat O3 uptake data, it is
 2     essential that there be confidence in the reliability of the different approaches.  To help
 3     establish the comparability of techniques, Wiester et al. (1993) measured total RT uptake in
 4     humans using a similar, though obviously scaled-up, system to that used for rats (Wiester
 5     et al., 1987, 1988).  Healthy subjects breathed 0.3 ppm O3 while seated in an exposure
 6     chamber.  Their faces were placed in a sealed facemask.  The facemask was attached to a
 7     large tube through which chamber air was circulated with a pump at a rate of «40 L/min.
 8     Upstream  and downstream O3 concentrations were measured continuously, as was ventilation
 9     with an induction  plethysmograph.  Subjects breathed at rest either through their nose or
10     their mouth: average fB = 16 bpm, VT = 598 to 642 mL, VE = 9.2 to 9.8 L/min.  While
11     nose breathing,  73 %  of inspired O3 was taken up  by the total RT;  and while mouth
12     breathing, 76%  of inspired O3  was taken up, which was  significantly higher than what was
13     found with nose breathing.  This difference is probably not, however, biologically
14     significant.  Significant negative correlations between fB and uptake in both mouth and nose
15     breathers were found. Similar correlations were found with VE.  No correlations were found
16     between uptake and VT or any other measure of breathing pattern or pulmonary function.
17          The  observations in Wiester et al. (1993) of a slight increase of total RT uptake
18     efficiency with oral breathing, and the inverse correlation of total RT uptake efficiency with
19     fB are consistent with those of Gerrity et al. (1988). Furthermore, the data on total RT
20     uptake are consistent overall with that of Gerrity  et al. (1988,  1993a).  The data from Gerrity
21     et al. (1988) reporting total RT uptake efficiencies of about 95 % were based on minimum
22     plateau  measurements, thus reflecting uptake during steady-state flow conditions, as opposed
23     to the cyclical conditions of actual breathing.  The data of Genity et al. (1993a), on the other
24     hand (in which total RT efficiencies of 80% were reported), were obtained by integrating the
25     product of concentration and flow, thus more accurately reflecting the acutal mass uptake of
26     O3 during cyclical breathing.  As was reported by Gerrity et al. (1993a), when they
27     computed uptake using the methodology from Gerrity et al. (1988), they found that the
28     total RT uptake measurements  were comparable.   Thus total RT mass uptake efficiencies at
29     rest of 80% are not unreasonable.
30          Hu (1991), Hu  et al. (1992b,  1993), and  Ultman et al. (1993) took a different approach
31     to measuring respiratory tract uptake of O3. They exploited the development of a rapid

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  1      responding O3 analyzer (Ben-Jebria and Ultman, 1989 and Ben-Jebria et al., 1990) to
  2      measure the recovery of small boluses of O3 delivered to different volumetric depths of the
  3      RT.  Ozone uptake was measured in a set of four experiments.  In the baseline experiments,
  4      absorption of O3 boluses was measured in healthy male subjects at rest. The O3 boluses
  5      were 10 mL in size,  with a peak concentration of 3 ppm.  The O3 analyzer characteristics
  6      were:  sample flow of 400 mL/min, 2-methyl-2-butene as reactive alkene, 10 to 90% step-
  7      response time of 110 ms, and lower detectable detection limit of 18 ppb. In the baseline
  8      experiments, the VT  was 500 mL and the inspiratory and expiratory flow rates were
  9      250 mL/s. In a complete set of measurements, bolus recovery was examined for
10      penetrations of 10 to 200 mL in 10 mL increments.  In the second set of experiments, the
11      effects of flow were measured by measuring bolus recovery as a function of penetration
12      depth for flows of 150, 250, 500, 750, and 1,000 mL/s at  a fixed  VT of 500 mL.  In a third
13      set of experiments, bolus recovery was measured as a function of penetration depth at a flow
14      of 250 mL/s and VT  of 500 mL, comparing the bolus delivered to a rubber mouthpiece or  to
15      a nasal cannula and thereby examining potential uptake differences between the two
16      pathways.  In the fourth set of experiments, the effects of Oj concentration on uptake were
17      determined by delivering boluses with peak concentrations  of 0.5,  1.0, 2.0 and 4.0 ppm.
18      The latter experiments were conducted because acute studies in isolated dog airways showed
19      that absorption efficiency was inversely related to inhaled concentration between 0.1 to
20      20 ppm (Vaughan et  al., 1969; Yokoyama and Frank, 1972).  However, later experiments  in
21      guinea pigs, rabbits (Miller et al., 1979), and humans (Gerrity et al., 1988) showed a lack  of
22      concentration dependence.  The dependence or lack of dependence of uptake efficiency on
23      O3  concentration provides information on the order of reactions of O3 with lung fluid lining
24      and tissue. Under steady-state conditions, concentration  independence of uptake efficiency
25      suggests  that first-order processes are playing a role.  However, because O3  absorption is
26      coupled to both interfacial transfer (gas-phase to solute O3) and subsequent reaction, at face
27      value one cannot conclude that saturated absorption rates are solely due to saturation of the
28      reaction components.
29          In all four experiments, Hu and colleagues computed  the first three moments of the
30      inspired and expired bolus distributions with respect to volume.  The zeroth moment of a
31      bolus is the O3 mass  contained in the bolus. Thus the zeroth moments of the inspired  and

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 1     expired boluses were used to compute O3 uptake efficiency (or absorbed fraction) A,
 2     breakthrough volume VB (the mean volume of the exhaled bolus), and bolus dispersion a2.
 3     The first moment of a bolus is its mean volumetric position.  The first moment on inspiration
 4     gives the penetration volume (Vp), and the first moment on expiration gives the breakthrough
 5     volume (VB).  In the absence of any O3 uptake,  a  longitudinally mixed bolus should have
 6     Vg=Vp.  The second moment of a bolus is its variance.  The difference in variance between
 7     the expired and inspired bolus (a ) is a measure of gas mixing, or dispersion, in  the lung.
 8          In the baseline experiments,  the breathing pattern was a resting pattern with a VT of
 9     500 mL and an average inspiratory flow of 250 mL/s.  These experiments were performed
10     on 9 male subjects and showed that almost all O3 was absorbed beyond a penetration depth
11     of 180 mL.  Only about 6% of inhaled 63 was recovered at the 180 mL penetration depth,
12     and beyond that depth, it was very difficult to obtain an accurate measurement of recovery.
13     The investigators also found that VB was greater than V  at penetration depths less than
14     100 mL, after which VB leveled out at a constant value.  Dispersion was insensitive to
15     penetration depth. An important finding of the baseline experiment was that at quiet resting
16     ventilation, about 50% of the O3 mass in a bolus inhaled through the upper airways  is taken
17     up by the upper airways.  To compare these data with results of Gerrity et al.  (1988), it is
18     necessary to assume that inspiratory and expiratory uptake efficiencies are equal.  Then the
19     unidirectional uptake efficiency of the upper airways to a depth comparable to that at which
20     Gerrity et al. (1988) positioned their sampling catheter is about 30%, which is approximately
21     25% less  than the 40% results of Gerrity et al. (1988).  This difference might be due to the
22     presence of a mouthpiece in the experiments of Hu and colleagues which could reduce the
23     uptake efficiency of the oral pathway. The functional response data of Adams et al.  (1989)
24     suggest that this might be the case.
25           The flow experiments showed that there was a general right shifting of the  A-Vp curves
26     with increasing flow (i.e., increasing flow causes a deeper penetration of O3 into the lung
27     with lower fractional uptake by the conducting airways).  Eventually, all of the O3 is still
28     absorbed. Breakthrough volume showed a similar pattern at all flows (i.e., greater than
29     penetration volume at small Vp but flattening  out at larger Vp).  As flow increased,  the level
30     of the plateau increased. Dispersion,  though constant as a function of Vp at all flows,
31     increased linearly with increasing flow.

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 1          The studies of Hu and colleagues investigating the role of exposure route in modulating
 2     O3 uptake efficiencies reported that the nose absorbed approximately 30 % more than the
 3     mouth.  This result is at variance with the findings of Gerrity et al.  (1988) indicating that
 4     there was only a slightly higher uptake by the oral pathway when compared to the nasal
 5     pathway.  Gerrity et al. (1988) studied uptake by the two pathways without the use of a
 6     mouthpiece or any other delivery system. Subjects were free to breathe naturally.  It is
 7     possible that the use of mouthpieces and  nasal canulas in the studies of Hu and colleagues
 8     caused artifacts, resulting in their findings for nasal and oral uptake efficiencies.  The study
 9     of Adams et al. (1989) showing enhanced pulmonary function response to O3  during a
10     mouthpiece exposure compared to facemask/oral exposure supports this. Finally, the
11     concentration-dependence studies showed that uptake efficiency was not affected by the
12     concentration of inspired O3 between 0.3 to 4 ppm, implying that O3 uptake is governed by
13     linear processes.
14          One of the very unique features of  the approach to measuring uptake efficiency taken
15     by Hu and colleagues is that the O3  bolus recovery data can be used to derive local mass
16     transfer coefficients for the conducting airways.  Regional mass transfer coefficients derived
17     experimentally in this way can then  be used as input into mathematical model simulations,
18     thereby potentially leading to more accurate models of O3 dose to the respiratory tract.
19          Hu and colleagues define the parameter Ka (s"1) as one that is suitable to characterize
20     local O3  absorption.  It is the product of the overall mass transfer coefficient K (cm/s) that
21     reflects the combined contribution of diffusion and chemical reaction to uptake, and the local
22     surface/volume ratio a (cm" ).  From the A-Vp curves, these investigators derived values for
23     Ka.  Thus, the mass transfer coefficients that were experimentally derived depended upon
24     assumptions about airway anatomy and morphology. As a result of the various experiments,
25     these investigators found a number of important properties for Ka:
26
27            •  The proximal subcompartment of the nose has a Ka that is 70% larger than
28              Ka for the proximal mouth compartment (see, however, the comment made
29              above regarding the nose/mouth differences).
30
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 1            •  Ka's in the upper airway compartment were between 1.20 and 2.24 s"1 and
 2               relatively insensitive to flow, indicating that diffusion resistance of
 3               O3 through the gas boundary layer is much less than through the mucus
 4               film. These data also are consistent with the pig and sheep tracheae
 5               experiments of Ben-Jebria et al. (1991).
 6
 7            •  In the proximal and distal conducting airway subcompartments, (Ka)"1 was
 8               linearly related to (flow)"1, suggesting that the gas phase absorption rate
 9               constant is directly proportional to flow.  In lower airways, therefore,
10               diffusion resistance of the gas boundary layer is important. Hu and
11               colleagues concluded that the gas boundary layer contributes 80 to 90% of
12               the overall diffusion resistance in the central airway compartment.
13
14            •  The mass transfer coefficient in the lung liquid lining (kj) is estimated to
15               fall from 1.4 cm/s in the URT to 0.17 cm/s in the respiratory airways.
16
17            •  The chemical reaction rate between O3 and the lung liquid lining was
18               estimated to be 7.3 x 106, 2.3 x  106, and 8.2 x 105  s'1 in the upper
19               airways, proximal conducting airways, and distal conducting airways
20               respectively.
21
22            •  The overall mass uptake coefficients determined in the work of Hu (1991)
23               are significantly higher than those used in the model of Miller et al. (1985).
24               If the mass transfer coefficients in the model are adjusted upward, the total
25               uptake efficiency would be higher than measurements have shown,
26               requiring a downward adjustment of pulmonary region mass transfer
27               coefficients.
28
29           Gerrity et al. (1993b) took a somewhat more conventional approach in an attempt to
30      acquire regional information on O3 absorption in the human respiratory tract.  Healthy
31      subjects underwent transnasal bronchoscopy while in an exposure chamber in which 0.4 ppm

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 1     O3 was present. Subjects were asked to breathe at 12 bpm. Inspired and expired air was
 2     sampled through a teflon catheter that had been passed through the biopsy channel of the
 3     bronchoscope and positioned approximately in the center of the airway lumen.  The air was
 4     drawn into a rapid response O3 analyzer (Ben-Jebria and Ultman, 1989 and Ben-Jebria et al.,
 5     1990) with a 90% response time of 250 mL/s while using ethylene as the reactive alkene.
 6     Average VT was 810 mL, and average inspiratory flow was 320 mL/s.  Air was sampled for
 7     5 breaths from  above the vocal cords, at the entrance to the trachea,  above the main
 8     bifurcation carina,  and midway through bronchus intermedius. Flow was measured
 9     simultaneously  by a pneumotach attached to a simple cylindrical mouthpiece.  Before and
10     after each measurement, a set of samples from the mouth was collected for reference.
11     Uptake was defined as the fraction of O3 mass lost across any anatomical segment; mass was
12     determined by integrating the product of O3 concentration and flow.  By way of comparison
13     with the other human O3 uptake studies, Gerrity et al. (1993a) found that total RT uptake of
14     O3 measured in this fashion was 91 %.  This is higher than the resting data of Wiester et al.
15     (1993); however, the average VT in this study of 810 mL compared with the 600 mL VT
16     reported by Wiester et al. (1993) may account for this difference.  When Gerrity et al.
17     (1993b) compute the unidirectional uptake efficiencies between the mouth and the various
18     sampling sites,  they find that 17.6%, 27.0%, 35.5% and 32.5% of O3 passing into the mouth
19     is taken up by structures up to the vocal cords, upper trachea, main bifurcation carina, and
20     bronchus intermedius, respectively.  They also computed the unidirectional uptake
21     efficiencies across individual airway segments which were: 17.6% between the mouth and
22     just above the vocal cords, 12.8% from above the vocal cords to the upper trachea, 11.5%
23     from the upper trachea to the main bifurcation carina, and essentially zero between the carina
24     and bronchus intermedius. The uptake between the mouth and just above the vocal cords is
25     considerably lower than what was measured earlier by Gerrity et al. (1988), even considering
26     the fact that in the earlier study peak plateaus were used.  As has been noted earlier, it is
27     possible that the mouthpiece played a role in reducing the uptake efficiency of the mouth.
28     The uptake efficiency of O3 across the trachea is in line with  the data from sheep and porcine
29     tracheae at the higher flow rates (Ben-Jebria et al., 1991).  The present data are also
30     consistent with  the bolus uptake data of Hu (1991) and colleagues, which were also acquired
31     with a mouthpiece.  When the  O3 bolus data are used to compute unidirectional uptake

       December 1993                          8-21       DRAFT-DO NOT QUOTE OR CITE
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 1     efficiencies (assuming that the segmental efficiencies are the same on inspiration and
 2     expiration), the Hu et al. (1992b) data yield uptake efficiencies of 21, 36, 44, and 46%
 3     between the mouth and the vocal cords, upper trachea, main bifurcation carina, and bronchus
 4     intermedius, respectively.  The O3 bolus data are, therefore, in good accord with the data of
 5     Gerrity et al. (1993b). The measured uptake efficiencies across airway segments clearly
 6     appear to be higher than those predicted by the model of Miller et al. (1985). If higher
 7     uptake coefficients in the conducting airways were used in the model of Miller et al.  (1985),
 8     the model would overestimate total RT O3 uptake.  To adjust for this, pulmonary uptake
 9     coefficients would have to be reduced.  Unfortunately , the data of Hu and  associates cannot
10     provide information beyond the conducting airways.
11          Gerrity et al. (1993b) also measured  O3 washout volumes (i.e., the expired volumes
12     required to cause a specified drop in O3 concentration). These type of data provide insight
13     into the location of major sites of O3 uptake.  At the mouth, the 90% washout volume was
14     142 mL; at the upper trachea, the 90% washout volume was 62 mL.  By the time the entire
15     anatomical dead space of the  lungs was washed out, the Oj concentration had fallen to zero
16     (Gerrity et al.,  1993b).  It is unclear whether the absence of recovered O3 after washout of
17     the conducting airways was due to O3 not penetrating beyond the conducting airways or all
18     of the  O3 penetrating beyond the conducting airways being absorbed.  The  latter possibility is
19     more likely based on the observations of Hu (1991).
20          In assessing the work of Gerrity et al. (1993b), it is significant to note that these
21     investigators measured expired plateaus of O3 concentration that were zero. This contrasts
22     with the earlier work of Gerrity et al. (1988, 1993a) in which a non-zero expiratory plateau
23     was observed.  The non-zero expiratory plateau may have been due to a number of factors
24     that are unclear.  Since ethylene was the reacting alkene  in all cases, it is unlikely that
25     interference with other gases such as CO2 was responsible.  Another possibility is that O3  in
26     the early expiratory phase became entrained in the posterior oropharynx and persisted for the
27     duration of expiration.
28
29     8.2.3.5   Intel-comparison of Ozone Dosimetry Studies
30          The previous sections emphasized the methods and results of individual experimental
31      studies on O3 dosimetry.  This section will focus on comparisons of the in vivo studies with

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  1     each other and will draw upon these comparisons to arrive at conclusions regarding the utility
  2     of these data for extrapolation purposes.  The discussion will be divided into three sections,
  3     focusing on  total RT uptake efficiency, unidirectional URT uptake efficiency, and LRT
  4     uptake efficiency.
  5          There  are two categories that distinguish various data sets among each other.  The first
  6     category is the mouth/nose category  listed in the Tables 8-2 to 8-4.  Studies indicated by
  7     M or N were performed with unencumbered breathing by either oral (M) or nasal (N)
  8     breathing. Unencumbered in the present context means the absence of a mouthpiece or
  9     canula.  Data listed as M/N are pooled from data encompassing oral and nasal breathing.
10     Data shown  as mouthpiece or nasal canula are data acquired using these devices to deliver
11     the O3 to the animal or human subject, and/or to measure flow.
12          The second category is the method used to compute O3 uptake efficiency.  There are
13     essentially two methods. One method, refered to as the steady-state method, relies  on
14     measuring the loss of O3 from a steady air flow moving across  an anatomical structure.
15     An example is the data of Yokoyama and Frank (1972) in which a constant flow of ozonated
16     air through the URT of a dog was maintained by a trachea! canula attached to a pump.
17     Uptake efficiency was measured by changes in equilibrium O3 concentration.  Another
18     example is the study of Gerrity et al. (1988) which used the steady-state method by
19     measuring the peak inspiratory and minimum expiratory O3 concentrations through a catheter
20     in the posterior pharynx. These measurements were compared  to the ambient chamber
21     concentration to obtain uptake efficiencies of the URT and LRT.  The second method is
22     refered to as the non-steady state method. This method uses the integration of the product of
23     flow and O3 concentration to compute O3 masses that are, in turn, used to compute uptake.
24     The studies of Wiester et al.  (1988) in rodents,  and Wiester et al. (1993) in humans are an
25     example of this technique, as is the study of Gerrity et al. (1993a).
26
27     Total Respiratory Tract Uptake Efficiency
28          Table 8-2 provides a summary of in vivo data in all animal species of RT O3 uptake
29     efficiency (F^.  The data reported for the studies of Gerrity et al. (1988,  1993a) have been
30     adjusted from the published values to account for the fact the Ft cited in those papers did not
31     include uptake in the URT on expiration.  To make the adjustment, the URT uptake

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                   TABLE 8-2.  TOTAL RESPIRATORY TRACT UPTAKE DATA8
Species
Human
Human
Human
Human
Human
Human
Human
Human
Human
Human
Human
Human
Human
Rat (F-344)
Rat (SD)
Rat (LE)
Rat (F-344)
Guinea Kg
Mouth/Note
M
N
M/N
M/N
M
M
M
M
Mouthpiece
M
N
Mouthpiece
Mouthpiece
N
N
N
N
N
Method
Steady
Steady
Steady
Steady
Non-Steady
Non-Steady
Steady
Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
Non-Steady
VT(mL)
832
754
832
778
1,650
1,239
1,650
1,239
825
631
642
500
1,000
2.8
2.4
2.7
2.6
2.4
Inspiratory
Flow (mL/s)
509
456
350
634
1,360
1,360
1,350
1,360
330
539
514
250
250
12.2
9.6
12.3
11.3
7.5
fB (bpm)b
18
18
12
24
25
35
25
35
12
16
16
15
7.5
118
123
132
113
77
Ft
0.97
0.96
0.97
0.96
0.88
0.87
0.97
0.95
0.91
0.76
0.73
0.86
0.93
0.44
0.46
0.48
0.54
0.53
Reference
Gefrity et al. (1988)°
Gerrity et al. (1988)
Gerrity et al. (1988)
Gerrity et al. (1988)
Gerrity et al. (1993a)b
Gerrity et al. (1993a)
Gerrity et al. (1993a)
Gerrity et al. (1993a)
Gerrity et al. (1993b)
Wiester et al. (1993)
Wiester et al. (1993)
Hu et al. (1992b)
Hu et al. (1992b)
Wiester et al. (1988)
Wiester et al. (1988)
Wiester et al. (1988)
Hatch et al. (1989)
Wiester et al. (1988)
       °M=mouth exposure by natural breathing; N=nasal exposure by natural breathing; M/N=pooled data from mouth and nasal exposure;
        Mouthpiece=exposureby mouthpiece; Nasal Canula=exposure by nasal canula; Steady=uptake computed during constant unidirectional
        flow; Non-Steady=uptake computed by integration during cyclic breathing; Ft=total RT uptake.
        fB is either the measured frequency or is computed based on reported flows and volumes.
       'Total RT uptake reported by Gerrity et al. (1988) and Gerrity et al. (1993a) did not include the contribution from URT uptake efficiency
        during expiration.  The data as listed here include an expiratory URT contribution assuming it equals inspiratory URT uptake efficiency.
1
2
3
4
5
6
7
efficiency upon expiration was assumed to equal to the inspiratory uptake efficiency.  The
Ft data listed for the study of Hu et al. (1992b) were derived from their bolus recovery data
by integrating the data over the desired VT.  Because Hu et al. (1992b) could not recover
boluses from a depth greater than 220 mL, it was assumed that any bolus delivered to a
depth greater than 220 mL was absorbed completely.  The derivation of Ft from the bolus
data was done for VTs of 500 mL and  1,000 mL.
       December 1993
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DRAFT-DO NOT QUOTE OR CITE
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                TABLE 8-3. UNIDIRECTIONAL UPPER RESPIRATORY TRACT
                                   UPTAKE EFFICIENCY DATA8
Species
Human
Human
Human
Human
Human
Human
Human
Human
Human
Human
Human
Dog (Beagle)
Dog (Beagle)
Dog (Beagle)
Dog (Beagle)
Rat (F-344)
Guinea Pig
Rabbit
Mouth/Nose
M
N
M/N
M/N
M
M
M
M
Mouthpiece
Mouthpiece
Mouthpiece
Nasal Canula
Nasal Canula
Mouthpiece
Mouthpiece
N
N
N
Method
Steady
Steady
Steady
Steady
Non-Steady
Non-Steady
Steady
Steady
Non-Steady
Non-Steady
Non-Steady
Steady
Steady
Steady
Steady
Non-Steady
Steady
Steady
Inspiratory
Flow (mL/s)
509
456
350
634
1,360
1,360
1,360
1,360
337
250
250
83.3
667
83.3
667
11.3
2.7
16.7
fB (bpm)
18
18
12
24
25
35
25
35
12
15
15
N/AC
M/A
N/A
N/A
113
N/A
N/A
FU«
0.40
0.43
0.41
0.38
0.37
0.41
0.16
0.22
0.18
0.30
0.47
0.72
0.37
0.34
0.12
0.17
0.62
0.41
Reference
Gerrity et al. (1988)
Gerrity et al. (1988)
Gerrity et al. (1988)
Gerrity et al. (1988)
Gerrity et al. (1993a)
Gerrity et al. (1993a)
Gerrity et al. (1993a)
Gerrity et al. (1993a)
Gerrity et al. (1993b)
Ultman et al. (1993)
Ultman et al. (1993)
Yokoyama and Frank (1972)
Yokoyama and Frank (1972)
Yokoyama and Frank (1972)
Yokoyama and Frank (1972)
Hatch et al. (1989)
Miller et al. (1979)
Miller et al. (1979)
       aM=mouth exposure by natural breathing; N = nasal exposure by natural breathing; M/N=pooled data from mouth and nasal exposure;
       Mouthpiece=exposure by mouthpiece; Nasal Canula=exposure by nasal canula; Steady=uptake computed during constant unidirectional
       flow; Non-Steady=uptake computed by integration during cyclic breathing; Ft=total RT uptake.
       fg is either the measured breathing frequency or is computed from flows and VT.
       N/A=not applicable.
1           To assess the consistency of the data, it is useful to examine it as a function of flow.
2      Figure 8-1 shows Ft as a function of inspiratory flow for all human studies.  The Ft from the
3      bolus recovery data of Hu et al.  (1993) are shown here for VTs of 500 mL,  1,000 mL, and
4      1,500 mL.  The  constitency of the data suggests that with respect to Ft there is good
5      agreement among the various experimental methods for humans. The data clearly show that
6      Ft decreases with increasing flow and increases with increasing  VT, all of  which is
7      qualitatively consistent with model predictions.
      December 1993
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           TABLE 8-4. LOWER RESPIRATORY TRACT UPTAKE EFFICIENCY DATA8
Species
Human
Human
Human
Human
Human
Human
Human
Human
Mouth/Nose
M/N
M/N
M
M
M
M
Mouthpiece
Mouthpiece
Method
Steady
Steady
Non-Steady
Non-Steady
Steady
Steady
Non-Steady
Non-Steady
VT (mL/8)
832
778
1,650
1,239
1,650
1,239
844
500
Inspiratory
Flow (mUs)
350
634
1,360
1,360
1,360
1,360
337
250
fB (bpm)b
12
24
25
35
25
35
12
15
Fjft Reference
0
0
0
0
0
0.
.93
.89
.68
.62
.96
.92
0.95
0.
.78
Oerrity et al.
Gerrity et al.
Oerrity et al.
Oerrity et al.
Gerrity et al.
Oerrity et al.
Gerrity et al.
(1988)
(1988)
(1993a)
(1993a)
(1993a)
(1993a)
(1993b)
Hu et al. (1992b)
         Human        Mouthpiece   Non-Steady     1,000

         Dog (Beagle)    N/AC        Non-Steady      168
         Dog (Beagle)    N/A
         Rat (F-344)     N
Non-Steady
Non-Steady
             168
2.6
250

112

168

 11.3
                     7.5     0.89   Hu et al. (1992b)

                    20      0.87   Yokoyama and Frank
                                  (1972)
                    30      0.83   Yokoyama and Frank
                                  (1972)
113
0.33   Hatch et al. (1989)
       *M=mouth exposure by natural breathing; N=nasal exposure by natural breaming; M/N=pooled data from mouth and nasal exposure;
        Mouthpiece=exposure by mouthpiece; Nasal CanuU=exposure by nasal canula; Steady=uptake computed during constant unidirectional
        flow; Non-Steady=uptake computed by integration during cyclic breathing; Ft=total RT uptake.
        fjj is either the measured breathing frequency or is computed from flows and Vj.
       cN/A=snot applicable.
 1           One observation is quite prominent. The rat data of Wiester et al. (1988) and Hatch
 2      et al. (1989) (not shown in Figure 8-1) are considerably lower than the human data.  Even if
 3      it is assumed that the rats were breathing up to three times resting ventilation (equivalent to
 4      an  inspiratory flow in humans of approximately 1,000 mL/s), the rat data would still be
 5      significantly lower than what was measured in humans.  The consistency of the human data
 6      of Wiester et al. (1993) (in which the same methodology was used to measure Ft in humans
 7      as was used for rats) with the other human data strongly suggests that the low Ft in rats is
 8      not a function of the methodology employed.  Overall, the evidence reasonably points to the
 9      conclusion that Ft in a rat is smaller than in a human.
10
        December  1993
                    8-26
                DRAFT-DO NOT QUOTE OR CITE
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                                  Vj-SOOml
                                           Hu9tal.,(1993)
      ,-1500ml      VT-g26ml
             Q«rrity«tal, (19936)
                                 V,-1239nl     V,-1850nl     VI-631ml
                               Q«rrily«(l.,(1993a) Q«rrtty»tal.,(1993) WNMtwet«l,(19fl3)
                                    »DO
                                1
                              0.9
                              0.8
                           UJ
                              0.7
(0
Q.
D
0>
0 0.6
N
0
0.5




	


i 1,1,1,1,1,1,
                                 0   200   400   600  800 1,000  1,200 1,400 1.600
                                            Inspiratory Flow (mL/s)
        Figure 8-1. Total respiratory tract uptake as a function of inspiratory flow in humans.
 1      Unidirectional Upper Respiratory Tract Uptake
 2           Table 8-3 summarizes the data on the unidirectional O3 uptake efficiency of the URT
 3      (FUrt)-  *n general, Furt describes the uptake efficiency of anatomical structures proximal to
 4      the larynx. A possible exception to this are the data of Hu et al. (1992b).  In that study, and
 5      other bolus studies, it was assumed that the URT is the volume of the respiratory tract
 6      50 mL distal to the lips.  This may or may not include the larynx.
 7           The wide variety of URT uptake data in different species and under different flow
 8      conditions allows one to  make some intra- and interspecies comparisons.  To make
 9      comparisons among different species, however, requires assumptions about the scaling of
10      breathing patterns among species.  Gerrity (1989) examined nasopharyngeal uptake data
11      available in different species at that time by examining the data as a function of the ratio of
       December 1993
8-27
DRAFT-DO NOT QUOTE OR CITE
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 1     predicted resting flow to measured flow (scaled flow).  The predicted flows were obtained
 2     using the allometric equations of Guyton (1947).  Ozone uptake efficiency of the URT was
 3     then described as a single function of scaled flow and species weight.  When newer data
 4     become available, this approach can be extended to URT data acquired by both mouth and
 5     nasal breathing.
 6           Figure 8-2 shows l-Furt for nasal breathing plotted as a function of the ratio between
 7     predicted and measured flow. As with the previous work of Gerrity (1989), all of the data,
 8     except the Beagle dog data, are  roughly consistent with each other.  The body weight
 9     dependence of the Gerrity  (1989) analysis is illustrated by the two dashed lines representing
10     the predicted range of flow dependences between rats (lower  line) and humans  (upper line).
11     All species, except the Beagle dogs, fall within these boundaries.
12           Figure 8-3 shows the data  for l-Furt by the oral route plotted hi the  same manner.  All
13     of the data are for humans except for the two Beagle dog data points.  Several observations
14     are worth noting. First, the data for the Beagle dog appear to be consistent with the human
15     data. The analysis of Gerrity (1989) predicts that the human data would generally lie above
16     the dog data, though not to a large degree.  This  suggests that the oral passage of the dog
17     may have similar O3 scrubbing properties as the human oral passage.  Second, within the
18     human data, use of a mouthpiece appears to reduce the uptake efficiency of the oral
19     passageway.  The closed diamonds are the data from the study of Gerrity et al. (1993a)
20     which involved unencumbered breathing. These data are generally lower than  the data of
21     Hu et al. (1993)  and the data of Gerrity et al.  (1993b).  The fact that the Hu et al. (1993)
22     and Gerrity et al. (1993b) data are consistent with each other supports this speculation. This
23     observation may account for the result of Ultman et al. (1993) that the uptake efficiency of
24     the URT is greater by the nasal pathway than by  the oral pathway, which is counter to the
25     observations of Gerrity et al. (1988).
26
27     Lower Respiratory Tract Uptake
28           Table 8-4 summarizes the data on the uptake efficiency of the LRT  tract (Flrt).  In the
29     current discussion, F,rt is the uptake efficiency of the LRT relative to the concentration of
30     O3 entering the LRT.  The human data of Gerrity et al. (1988,  1993a, 1993b)  and the rat
31     data of Hatch et al. (1989) include the larynx in the LRT.  The Beagle dog data of

       December 1993                           8-28      DRAFT-DO NOT  QUOTE OR CITE
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                                Natural      Natural      Canute      Canute
                               Steady State  Non-Steady State Steady State  Non-Steady State
                            0.7
                            0.5
                         LU

                         i
                          
-------
                                 Natural      Natural      Canute      Canute
                               Steady State  Non-Steady State Steady State  Non-Steady State
                                  •         »         D         0
                         UJ
                            0.7
                            0.5
                            0.3
                            0.2
                            0.1
  D
Bragl*
                                        I
                                                            Boagto
                               0.1     0.2  Q.3    0.5      1      23
                                   Predicted, Resting Flow/Measured Row
      Figure 8-3.  Unidirectional uptake efficiency in the upper respiratory tract by the oral
                  pathway.  The ratio of predicted resting flow to measured flow scales the
                  flow to allow for interspecies comparison.  The closed squares and
                  diamonds at a scaled flow of 0.25 are from Gerrity et al. (1993a); the closed
                  square at  a scaled flow of 0.65 is from Gerrity et al. (1988); the open
                  diamond at a scaled flow of 0.97 is from Gerrity et al. (1993b); the
                  remaining open diamonds are from Hu et al. (1993).
1     observation is that the single data point of Gerrity et al. (1993b) is approximately 20% higher
2     than the data  of Hu et al. (1993) at a comparable flow and VT.  Second, the data of Gerrity
3     et al. (1993a) are markedly lower than what the data of Hu et al. (1993) suggest for
4     comparable flows and VTs.  In assessing this discrepancy, it is important to keep in mind
5     that the URT is defined in the work of Hu et al. (1993) as a fixed volume of 50 mL distal to
6     the lips.  This is an inaccurate definition that could influence greatly the estimation of LRT
7     uptake efficiency.   The coherence of all of the data on Ft and Furt by the oral pathway
      December 1993
             8-30
DRAFT-DO NOT QUOTE OR CITE
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                              1 ,-
                          -£0.9
                          Ul
                             0.8
                          q> ._
                          § °'7
                             0.6
                                    \ft-500n4
                                    VHESml
                                               \ft-1000mt
                                              Hu •(•!., (1883)
                                                         Vt-1500ml
                                               VM239mi     wiesoml
                                             G«rt*y«tal.,(1983a) Qwr«y«tal.,(1M3)
                                                 *          D
                                                                  Q
                                    J	I	L.
                                                  I  I	I	I	1	1	1_
                                0   200  400  600  800 1,000  1,2001,4001,600
                                         Inspiratory  Flow (mL/s)
       Figure 8-4. Uptake efficiency of the lower respiratory tract as a function of inspiratory
                   flow in humans.
 1     suggests that F^ should also be consistent among studies. If it is assumed that the data
 2     represented by the Gerrity et al. (1993a, 1993b) studies more accurately reflect human LRT
 3     uptake efficiency, then the flow dependence for F^ would be considerably steeper than
 4     suggested by the data of Hu et al. (1993).
 5          Finally, the F^ data of Yokoyama and Frank (1972) in Beagle dogs were acquired with
 6     flows that were about 100% and 150%  of resting flow rates and VTs approximately 150% of
 7     normal (Guyton, 1947).  When flow is  scaled in the dogs, the F^s of 0.87 and 0.83 at the
 8     two flow rates are consistent with the human data of Gerrity et al. (1993b) but higher than
 9     the Hu et al. (1993) data.
10
       December 1993
8-31
DRAFT-DO NOT QUOTE OR CITE
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 1     8.2.4   Dosimetry Modeling
 2     8.2.4.1  Background
 3          Table 8-5 presents a summary of theoretical studies of the uptake of O3 by the
 4     respiratory tracts (or regions) of humans and laboratory animals that have become available
 5     since the 1986 review.  Although there are ten investigations listed, there are only five
 6     distinct dosimetry models (with respect to groups of co-workers and independent model
 7     formulation):  The models of (1) Overton et al. (1987) and Miller et al. (1985), (2) Hanna
 8     et al. (1989), (3) Grotberg et al. (1990)—although they consider two reaction schemes,
 9     (4) Mercer et al. (1991), and (5) Hu et al. (1992a).  In some cases, several references have
10     been grouped into one investigation. This is because the multiple studies came from the
11     same co-workers or laboratory, but added to or were complementary to the original or
12     common dosimetry modeling theme.
13          Major factors affecting the local uptake of reactive gases in the respiratory tract are
14     respiratory tract morphology and anatomy; the route of breathing (nose or mouth); the depth
15     and rate of breathing (VT and fB); the physicochemical properties of the gases; the processes
16     of gas transport;  and the physicochemical properties of the liquid lining of the respiratory
17     tract, respiratory tract tissue, and capillary blood. A detailed discussion of these factors can
18     be found in Overton (1984), Ultman (1988), and Overton and Miller (1988),
19          Because all of the dosimetry models listed in Table 8-5 were developed to simulate
20     uptake in the LRT or the total RT, they have some aspects in common. This includes the
21     formulation of O3 transport  and wall loss in the air compartments of the RT, the use of
22     species-dependent morphometric models or data to define air and liquid lining compartment
23     dimensions,  and  a description of the transport and loss of O3 in the liquid lining and tissue.
24          In all the dosimetry models that have become available since 1986, except for Ultman
25     and Anjilvel (1990) and Grotberg et al. (1990) which are  discussed later, O3 transport and
26     loss processes in air compartments are approximated in terms of a one-dimensional, time
27     dependent, partial  differential equation of continuity.  This type of equation accounts  for
28     axial convection, axial dispersion or diffusion, and the loss of O3 by absorption at the gas-
29     liquid interface.  The use of this approximation is very common in modeling the transport in
30     the LRT of gases such as oxygen, nitrogen, helium, carbon dioxide (COj), etc. (e.g.,
31     Scherer et al., 1972; Paiva, 1973; Chang and Fartii, 1973; Yu, 1975; Pack et al., 1977) and

       December 1993                           8-32      DRAFT-DO NOT QUOTE OR CITE
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                                         TABLE 8-5.   THEORETICAL OZONE DOSIMETRY INVESTIGATIONS8
              Species and Region
             Modeled/ Anatomical
                   Model
 Liquid Lining and Tissue
 Transport and Chemical
       Reactions
                                                            Dosimetry Model /Subject of Investigation
                                                                                      Results, Predictions
                                                                                                                          Reference
             Guinea pig
             LRT/Kliment
             (1973), Schreider
             and Hutchens
             (1980); rat LRT/
             Kliment (1973), Yeh
             etal. (1979)
Miller et al. (1985)        Enhanced Miller et al. (1985). Investigates the
                        effect on predictions of anatomical models,
                        FRC, ventilation, and TB liquid lining rate
                        constant. Simulates 03 uptake in anatomical
                        models of rat lobes.
                                           With respect to different anatomical models for the same and
                                           different species: qualitative similarity in the shape of net
                                           and tissue dose versus airway number curves, but significant
                                           differences in regional fractional uptakes. Maximum tissue
                                           dose in vicinity of PAR.  PAR dose decreases with
                                           increasing time of flight to this region. Maximum tissue
                                           dose in the vicinity of the first PUL region segment of
                                           anatomical models.
                                                       Overton et al.
                                                       (1987)
OO
             Human, rat, rabbit,
             and guinea pig
             LRT/Not specified
Miller et al. (1985) and
Overton et al. (1987)
Miller et al. (1985) and Overton et al. (1987).
Compares 03 dose profiles of human, rat,
guinea pig, and rabbit.  Uses model and
experimental data to estimate 03 dose-response
curves for decrements in FEVj  (humans) and
for BAL proteins in rat, guinea pig, and rabbit.
Compares LRT uptake predictions to the human
experimental data of Gerrity et al. (1988).
Similarity among species in the shape of the airway segment
curve:  tissue dose increases distally in the TB region,
reaches a maximum in the in the first PUL region segment
for human, rat, and guinea pig and in the last TB region
segment of the rabbit, and then decreases distally in the PUL
region.  Predictions of uptake distal to the oropharynx are in
agreement with Gerrity et al. (1988).
Miller et al. (1988)
             Rat total RT/URT:
             Schreider and Raabe
             (1981); TB: Uses
             rat cast data (Raabe
             etal., 1976) to
             define TB region
             paths; PUL: Yeh
             et al. (1979) for
             model of the acinus
TB and PUL region mass
transfer coefficients based
on Overton et al. (1987)
Enhanced Overton et al. (1987). Illustrates
dose distribution along the longest and shortest
(as defined by time of flight) paths from the
trachea to the most distal alveoli.
Three-fold difference in PAR doses of the shortest and
longest paths from trachea to PAR. Dose distributions along
the longest or shortest path were qualitatively similar to
Overton et al. (1987) with maximum tissue dose in the first
PUL region generation.
Overton et al.
(1989)
             Human LRT
             (newborn to
             adult)/TB:  based on
             Yeh and Schum
             (1980); PUL:  based
             on Hansen and
             Ampaya (1975)
Miller et al. (1985)        Enhanced Overton et al. (1987). From various
                        sources, develops age-dependent LRT
                        anatomical models.  For quiet and maximal
                        exercise breathing, applies the dosimetry model
                        of Miller et al. (1985) to several ages from birth
                        to adult, illustrating the LRT distribution of
                        absorbed 03.
                                           For quiet breathing, the LRT distribution of dose, the
                                           percent uptake, and the PAR dose are not very sensitive to
                                           age; but are more sensitive during exercise. Regardless of
                                           age and breathing state, the largest 03 dose occurs in the
                                           PAR.  No uptake in the URT.
                                                       Overton and Graham
                                                       (1989); Miller and
                                                       Overton (1989)
-------
                                   TABLE 8-5  (cont'd).  THEORETICAL OZONE DOSBMETRY INVESTIGATIONS1
              Species and Region
              Modeled/ Anatomical
                    Model
                      Liquid Lining and Tissue
                      Transport and Chemical
                            Reactions
   Dosimetry Model /Subject of Investigation
                  Results, Predictions
                                                                                                 Reference
             Human total RT/URT:
             Hanna and Scherer
             (1986); LRT:  Weibel
             (1963)
                     Time-dependent molecular
                     diffusion and first order
                     reactions in liquid lining
                     and interstitium; transfer
                     through epithelium
                     modeled as a permeability
                     process—no reactions in
                     this layer. URT and LRT
                     liquid lining rate
                     constants:  50 and 1 times
                     that of Miller et at.
                     (1985), respectively
Model development.  Lung dimensions scaled to
those of a young male and a young female.
Contrasts LRT airphase concentrations during
exercise and rest.  Compares male and female
airphase 03 concentrations and male and female
subepithelial concentrations.
URT uptake may be greater in cold than in warmer air.
For the ventilatory parameters used:  (1) Subepithelial
Oj concentrations are a maximum in the terminal
bronchioles or the first respiratory bronchioles, and
(2) These concentrations are greater in the female than
male for most of the RT.
Hanna et al. (1989)
oo
u>
*>•
Human, distal
segment of a lobe/is
based on Horsfield
etal. (1971)
                                  Similar to Miller et al.
                                  (1985)
Monte Carlo simulation; transport processes
defined in terms of probabilities based on
physical and chemical principles.  The effect of
lung asymmetries on the distribution of uptake
in the pulmonary region.
Tissue dose in the PAR along the shortest path is
approximately 50% larger than that along the longest path.
Ultman and Anjilvel
(1990)
             Human LRT/Weibel
             (1963)
                     Investigates two
                     formulations:
                     Miller et al.  (1978) and
                     Miller et al.  (1985)
Model development; assumes quasi-steady
conditions.  Air phase concentration and tissue
dose profiles for the two reaction schemes, for
various ventilatory parameters and various
liquid lining transport and chemical parameters.
Compares predictions of first order reaction
scheme to results of Miller et al. (1985).
This model does not conserve mass.  Predications should     Orotberg et al.
only be considered qualitatively. Maximum tissue dose in    (1990);
respiratory bronchioles for both chemical reaction schemes.   [Grotberg (1990)]
             Rat LRT/based on
             serial reconstruction
             of a set of
             intrapulmonary
             airways and their
             ventilatory units
             combined with a
             single path from the
             larynx to the
             reconstructed set
             based on Yeh  et al.
             (1979)
                     TB and PUL region mass
                     transfer coefficients based
                     Overton etal. (1987)
Model development.  Illustrates the influence of
ventilatory unit size and proximal anatomic dead
space and on the uptake and distribution of
inhaled 03 in ventilatory units.

[Illustrates the influence of ventilatory unit
volume on the distribution of inhaled 03 within
ventilatory units.]
Ventilatory unit uptake is significantly influenced by both
proximal airway deadspace and ventilatory unit volume.
Flux of 03 to air-liquid interface in the proximal portions
of larger ventilatory units are significantly greater man in
smaller units.
Mercer et al. (1991)

[Mercer and Crapo
(1993)]
-------
                                   TABLE 8-5 (cont'd).  THEORETICAL OZONE DOSIMETRY  INVESTIGATIONS'
              Species and Region
              Modeled/Anatomical
                    Model
                       Liquid Lining and Tissue
                       Transport and Chemical
                             Reactions
                                                             Dosimetry Model /Subject of Investigation
                                                             Results, Predictions
                                                                                                 Reference
             Rat ventilatory unit/
             Mercer et al. (1991)
                      Mercer et al. (1991)
Mercer et al. (1991). Along a path distally
from a bronchiolar-alveolar duct junction ,
compares experimentally determined changes in
ventilatory unit wall thickness due to an O3
exposure to dosimetry model predictions of flux
to the air-liquid interface.
As a function of distance from the broncho-alveolar duct     Pinkerton et al.
junction, experimentally determined relative changes in      (1992);
ventilatory unit wall thickness due to Oj exposure are very   Miller and Conolly
similar to predicted relative fluxes to the air-liquid          (1993)
interface.
             Human total RT/URT:
             Fredberg et al.
             (1980);
             LRT:  Weibel (1963)
                      Pseudo steady-state
                      diffusion and first order
                      reactions combined with
                      biochemical data of Miller
                      et al. (1985)
Model development.  Illustrates LRT
distribution of (1) airphase concentration at
various times during the breathing cycle and
(2) Oj flux (dose) to liquid lining and to tissue.
Flux of 03 to air-liquid interface decreases distally; flux to
tissue increases along the conducting airways, reaches a
maximum in the terminal bronchioles then decreases
rapidly in the gas exchange region.
Hu et al. (1992a)
oo
 URT = upper respiratory tract; total RT = total respiratory tract; LRT = lower respiratory tract; PAR = proximal alveolar region; generally, for modeling purposes, the first pulmonary
region
 generation or model segment; PUL = pulmonary region; FRC = functional residual capacity; TB = tracheobronchial region; FEVj  = forced expiratory volume at 1 s.
 Refers to  the theoretical or mathematical formulation aspects of gas transport and reactions without the specification of morphometric and physiological parameter values.
-------
 1     has been assumed to be applicable to O3.  Ultman and Anjilvel (1990) use a Monte Carlo
 2     method to simulate O3 uptake. Based on the physical and chemical principles of mass
 3     transport in the respiratory tract, probabilities are assigned to the fate of a molecule in a way
 4     so as to account for convection, dispersion, and loss to the liquid lining.
 5           Dosimetry models published since 1986 can be grouped according to how transport and
 6     chemical reactions are modeled in respiratory tract fluids and tissues:  those based on the
 7     formulation of (1) Miller (1977) and Miller et al. (1978) who used an  instantaneous reaction
 8     scheme and (2) Miller et al. (1985)  who used a quasi-steady first order reaction scheme.
 9     These two approaches are discussed in the earlier criteria document  (U.S. Environmental
10     Protection Agency, 1986). In addition to the use of similar formulations for liquid and tissue
11     transport/reactions, all of the post 1986 studies use essentially the same biochemical data of
12     either Miller et al. (1985) for humans or Overton et al. (1987) for laboratory animals.  The
13     implication is that most of the studies are expected to predict qualitatively similar results.
14           There are minor variations on the second chemical reaction formulation.  Hanna et al.
15     (1989) use a time-dependent diffusion reaction equation, instead of time-independent (quasi-
16     steady) equation as do Miller et al.  (1985). Based on the rate constants used by Hanna et al.
17     (1989) and on discussions in  Miller et al. (1985) and in Grotberg et al. (1990), one can infer
18     that the modeled transport processes in the liquids and tissues are essentially quasi-steady,
19     which is equivalent to the second formulation.  Another variation uses mass transfer
20     coefficients determined by the second formulation and the biochemical assumptions of Miller
21     et al. (1985) or Overton et al. (1987). The liquid and tissue transport/reaction formulation
22     for specific investigations is indicated in column 3  of Table 8-5.
23           In addition to the assumptions and the formulation of equations that describe the
24     transport and loss of O3 in the respiratory tracts of humans or laboratory animals, it is
25     important to evaluate whether simulation results reflect accurate solutions to the mathematical
26     dosimetry model formulation. Of the five distinct model formulations listed in Table 8-5,
27     Overton et al. (1987), Mercer et al. (1991), and Hu et al. (1992a) discuss most or all of the
28     relevant issues of stability, solution convergence, and mass conservation.  In addition, using
29      steady unidirectional flow in a straight tube as a test case, they report successfully simulating
30     analytical solutions to their equations of transport and uptake.  Neither Hanna et al. (1989),
31      Ultman and Anjilvel (1990), nor Grotberg et al. (1990) address the  issue of accuracy.  There

        December 1993                           8-36       DRAFT-DO NOT QUOTE OR CITE
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 1     is no reasonable way to judge whether or not the solutions of Hanna et al.  (1989) or of
 2     Ultman and Anjilvel (1990) accurately represent solutions to their dosimetry model
 3     assumptions and formulations; however, with the exception of Grotberg et  al. (1990), there
 4     are no reasons to assume that solutions of these models are not accurate.
 5           Because the Grotberg et al. (1990) model formulation is different than the others, an
 6     explanation is needed.  Based on the smallness of relevant parameters, Grotberg et al. (1990)
 7     assume quasi-steady conditions for O3 concentration and air velocity in the air compartments
 8     and obtain approximate analytical solutions to the tune-independent three-dimensional
 9     equation of continuity for a model airway, and apply the results to the morphometric model
10     of Weibel (1963).  One advantage of analytical solutions is that they account naturally for
11     parameters (such as dispersion and gas phase mass transfer coefficients) or local processes
12     (e.g., possibility of high uptake at airway entrances) that must be known and estimated for,
13     or incorporated into, the one-dimensional approach.  Grotberg et al. (1990) carry out
14     simulations using anatomical and physiological conditions based on Miller et al. (1985) and
15     compare their results to the results of the latter.  Although their predictions are qualitatively
16     similar to Miller et al. (1985), they predict significantly larger pulmonary tissue doses (up to
17     as much as 10 times) than do Miller et al. (1985).  A comparison of the pulmonary region
18     doses predicted by Grotberg et al. (1990) to those of Miller et al. (1985) indicates that the
19     Grotberg et al. (1990) model does not conserve mass:   it predicts that the pulmonary region
20     absorbs over 3.4 times the amount of O3 inhaled. The overprotection may be an artifact of
21     the quasi-steady approximation, because effects due to differences in the time of flights from
22     the trachea to different LRT locations are not taken into account. In any case, the
23     quantitative predictions reported by Grotberg et al.  (1990) are questionable.
24           Chemical data more recent than used for the dosimetry models in Table 8-5 show that
25     compounds other than unsaturated fatty acids (the only  compound with which O3 is assumed
26     to react in the models using the second chemical  reaction formulation) are  as reactive or
27     more reactive with O3 (Pryor, 1992).  Using these data, estimates of O3 diffusion
28     coefficients in the liquid lining and lipid bilayers, layer thicknesses, and data on the
29     concentrations of biocompounds in these layers, Pryor (1992) estimates that (1) most of any
30     O3 that penetrates into a cell bilayer reacts within the layer, very little if any penetrates to
31     the cell interior, and (2) O3 will  not penetrate the liquid lining  where it is greater than

       December 1993                           8-37      DRAFT-DO NOT QUOTE OR CITE
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 1     0.1/*m thick.  Several relevant comments are given by Pryor (1992):  (1) the calculations
 2     are considered a crude first approximation; (2) the possibility that a small fraction of 63 may
 3     penetrate the bilayer and reach the cell interior can not be excluded; and (3) surfactant layers
 4     can be very thin, and some cells may not be protected very well or at all.
 5           If the conclusions of Pryor (1992) are essentially correct, they have implications for
 6     past and future dosimetry modeling studies because: (1) past investigations have
 7     underestimated the reactivity of O3 with biocompounds; (2) with respect to cellular damage,
 8     products of O3 reactions in the liquid lining may be the main toxic compound; (3) increasing
 9     the value of rate constants would have no effect on predictions of dosimetry models using the
10     instantaneous reaction scheme, because  rate constants in this approach are assumed to be
11     infinite,  however, increasing the concentration of reacting biocompounds would  increase
12     uptake; (4) use of only unsaturated fatty acid data (with a first order reaction scheme) results
13     in an underestimate of the reactivity of O3 in the liquid lining and an overestimate of the
14     O3 tissue dose, but a possible underestimation of the toxic dose that is due to reaction
15     products; and (5) with higher O3 reaction rates, the first order chemical reaction formulation
16     would result in larger predicted uptakes.
17
18     8.2.4.2   Dosimetry Model Predictions
19     Similarity of Model Predictions
20           A survey of dosimetry modeling results shows that in some areas of investigation there
21     is a  qualitative similarity in predictions by models of different groups of investigators for
22     different species or sub-populations.
23           (1)    Distribution ofLRTdose (dose profiles or dose versus generation).  As shown in
24                 Figure 8-5, beginning at the trachea, net dose (O3 flux to air-liquid  interface)
25                 slowly decreases distally in the tracheobronchial  region and rapidly  decreases
26                 distally in the pulmonary region.  Tissue  dose (O3 flux to liquid-tissue interface)
27                 is very low in the trachea, increases to a  maximum in the terminal bronchioles or
28                 first generation in the pulmonary region,  and decreases rapidly distally from this
29                 location, (e.g., Miller et al., 1978, 1985, 1988; Overton et al.,  1987, 1989;
30                 Overton and Graham, 1989; Grotberg et al., 1990; Hu et al., 1992a).
        December 1993                            8-38      DRAFT-DO NOT QUOTE OR CITE
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                 10"
              cr>
                  10~8-
                  101
                    10
                  Zone
                 Order
              Generation
           A  Generation









1 ^
O
O
O
0



Human -
Rat
Guinea Pig
Rabbit
(No

•.
2 3
2 3
2 4
2 4

•~.u 	
VT (mL)
	 • 	 8OO.O
-• — -•- 1 .88
	 2.63
1 3.20
absorption In tha


4
4 s e
e a 10
e 8 10


f (bpm)
1S.O
ee.o
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38.8
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12 14 181
i . 1.1 .1
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-


Rabbit
Guinea Pig
Rat
Human
                  10-
                                       <&-'•'
                               •^,.:-*
                            •"   Human 	^~
                                  Rat	--
                             Guinea Pig ——-
                                Rabbit
                VT (mL)  f (bpm)
                8OO.O   1S.O
                   1.88  86.O
                   2.63  8O.8
                  13.2O
                                   (No absorption In the URT)
                  Order
              Generation
            B Generation
O
O
O
O
2
2
2
2
3
3
4
4
4
e
e
s
a
8
 e
10
10
 B
 7
12
12
 8
14
14
 e 7
 0 10 11
1B 18 17
1« 17 18
12
18
SO
13
21
22
 8  Rabbtt
14 Guinea Pig
23 Rat
23 Human
      Figure 8-5. Net dose (A) and tissue dose (B) versus sequential segments along
                  anatomical model airway paths for human, rat, guinea pig, and rabbit.
                  In general, each segment represents a group of airways or ducts, with
                  common features as defined by the designers of the anatomical model
                  (human and rat: generation; guinea pig:  order;  rabbit:  zone).  For a
                  given species the plotted dots represent a predicted dose that corresponds to
                  a given segment.  The dots have been joined by lines for ease of
                  interpreting  the plots; these lines do not represent predicted values except
                  where they intercept the dots. TB =  tracheobronchial region.
                  P = pulmonary region.

      Source: Overtoil and Miller (1988).
1          If O3 were the only toxic agent and all the tissues of the LRT were equally sensitive to
2     the same dose, the models predict that the greatest morphological damage would occur in the
3     vicinity of the junction of the conducting airways and the pulmonary region and decrease
4     rapidly (distally) from this area, which is consistent with observations in laboratory animals
      December 1993
                  8-39
                         DRAFT-DO NOT QUOTE OR CITE
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 1      (see Section 6.2.4).  On the other hand, using the best estimates of morphometric and
 2      physiologically based biochemical parameters of Miller et al. (1978,  1985) and Overton et al.
 3      (1987), the models predict extremely (relatively) low tissue doses in the trachea and large
 4      bronchi; this suggests very little or no tissue damage should occur there, which is contrary to
 5      observations (see Section 6.2.4). However, this is moot, if as suggested by Pryor (1992),
 6      the toxic substances are primarily reaction products of and not O3 itself.  In this case, the
 7      O3 net local dose, not the local O3 tissue dose, may be a better estimator of local toxic tissue
 8      dose, since the rate of production of products would be related to the rate of O3 uptake.
 9           (2)    Effect of exercise or increased ventilation.  The effect of exercise is to slightly
10                 increase the tracheobronchial region dose and significantly increase the
11                 pulmonary region total dose (mass of O3) and the centriacinar region dose (mass
12                 per unit surface area),  (e.g., Miller et  al., 1979, 1985; Overton et al., 1987,
13                 1989; Overton and Graham, 1989; Hanna et al.,  1989; Grotberg et al., 1990.)
14
15           (3)    Effect of respiratory tract inhomogeneity.   Models have predicted that the further
16                 the proximal alveolar region is from the trachea, the less the O3 tissue dose
17                 (mass of O3 absorbed per unit surface area) to the proximal alveolar region.
18                 (For modeling purposes, the proximal alveolar region has been defined as the
19                 first pulmonary generation or the first pulmonary region  model segment along a
20                 path; this region is a part of the centriacinar region.) Overton et al.  (1989)
21                 predicted a threefold greater proximal alveolar region dose for the shortest path
22                 relative to the longest path in rats.  Ultman and Anjilvel  (1990)  simulated
23                 O3 distribution in a small segment (< 1 %) of the distal airways  of an asymmetric
24                 anatomic model of the human lung.  They found that the O3 tissue dose
25                 (mass/cm2) in the proximal alveolar region along the shortest path was
26                 approximately 50% greater than that along the longest path.  Mercer et al.
27                 (1991) found that path distance and ventilatory unit size affect dose: predicted
28                 dose in the proximal segments (essentially, the proximal  alveolar region) of the
29                 larger ventilatory  units  (with the smallest proximal dead  space) are significantly
30                 larger than the average proximal segment doses.  Further, for the small sample
31                 of ventilatory units modeled (43), Mercer  et al. (1991) predicted a range of

        December 1993                           8-40       DRAFT-DO NOT QUOTE OR CITE
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 1                proximal segment doses of greater than a factor of 6.  Because the proximal
 2                alveolar regions of Ultman and Anjilvel (1990) and of Mercer et al.  (1991)
 3                belonged to a "local cluster" and there are many clusters with varying distances
 4                from the trachea, a greater variability than 50% and a factor of 6, respectively,
 5                in proximal alveolar region doses is to be expected. Mercer and Crapo (1993)
 6                illustrateed the effect of ventilatory unit volume alone on the distribution of dose,
 7                predicting that a 2.3 times  bigger unit receives 1.9 times the dose (mass per
 8                surface area) of the smaller unit at the entrance of the unit.
 9          The variability of predicted proximal alveolar region and, by inference, centriacinar
10     region doses  suggests the magnitude of lexicological effects for different centriacinar regions
11     are different.  This prediction is consistent with the observations of Schwartz et al. (1976)
12     and Boorman et al. (1980) of variation of damage among different centriacinar regions of the
13     same rat. It  is reasonable to assume that variable damage at equivalent but different
14     morphological locations also occur in humans.
15
16     Specific Topics
17          Effect of Assumptions About Anatomical Dimensions.  For rats and guinea pigs,
18     Overton et al. (1987) used two morphometrically based anatomical models  (rat anatomical
19     models:  Kliment, 1973, and Yeh et al., 1979; guinea pig anatomical models: Kliment,
20     1973, and Schreider and Hutchens, 1980) to investigate the influence of anatomical model
21     formulation on predicted uptake.  Results with all four anatomical models in combination
22     with different ventilatory parameters showed a qualitative similarity in the shapes of the dose
23     profiles, but  the two anatomical  models for the same species resulted in considerable
24     differences in predicted  percent total and pulmonary region uptakes.
25
26          Respiratory Tract  Uptake in  Human Adults and Children.  Overton and Graham
27     (1989) used several sources of data on  age-dependent LRT dimensions and structure to
28     construct theoretical LRT anatomical models for humans from birth to adult. The
29     O3 dosimetry model of Miller et al. (1985) was used to estimate the regional and local
30     uptake of O3.  For the percent uptake (84 to 88%) during quiet breathing, the LRT
31     distribution of absorbed O3 and the centriacinar O3 tissue dose are not very sensitive to age.

       December 1993                           8-41       DRAFT-DO NOT QUOTE OR CITE
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 1     For heavy exercise or work, regional percent uptakes are more dependent on age than during
 2     quiet breathing, and total uptakes range from 87 to 93%.  Generally, the total quantity of
 3     O3 absorbed per minute increases with age.  For all conditions simulated, the largest
 4     O3 tissue dose is predicted to occur in the centriacinar region.  Miller and Overton (1989)
 5     present similar results. Because uptake by the URT was not simulated and because this
 6     region can be assumed to have an important effect on LRT uptake, a comparison of
 7     predictions of LRT uptakes in children and adults should be viewed with caution.  On the
 8     other hand,  URT uptake probably has little effect on the shape of the dose curves.
 9
10     Dosimetry Modeling Results Compared to Dosimetry Data
11           Based  on the experimental conditions discussed in  Gerrity et al. (1988) and using the
12     model and parameters of Miller et al. (1985), Miller et al. (1988) simulated the uptake of
13     O3 distal to  the oropharynx of human subjects.  For the target fBs of 12  and 24 bpm and VTs
14     ranging from 0.4 to over 1.6 L,  the simulation results were  in good agreement with the
15     breath-by-breath experimental data.  The average experimental LRT uptake efficiency was
16      «0.91 as compared to the 0.89 prediction given by Miller et al. (1985)  for the region distal
17     to the oropharynx. It should be  kept in mind, however, that values for uptake efficiency
18     from the Gerrity et al. (1988) study were derived from the raw data using a steady-state
19     method, whereas the models of Miller et al.  (1985) and Miller et al. (1988) utilize cyclic
20     flow, thus making the predictions more appropriate for comparison with uptake data from
21     non-steady-state methods.  From an analysis  hi Gerrity et al. (1993a), it appears that total RT
22     uptake computed by either steady-state or non-steady state methods differ by only about 10%
23     in relative terms.
24           A further refinement of the models of Miller et al. (1985) and Overton et al. (1987) has
25     been  made in which an URT has been added, thus allowing  comparisons with total RT
26     uptake efficiency  data. The refined model utilizes assumptions shown in Table 8-6.
27           Based  on Table 8-6, the mass transfer coefficients for the nasopharyngeal and
28     tracheobronchial regions of the rat  were computed using two sets of data. Firstly,
29     calculations were based on the total uptake and the 18O3 nasopharyngeal uptake data of Hatch
30     et al. (1989), which were the averages for 8  rats.  The second set of data was that of the
31     8 rats.  Both approaches gave essentially the same set of parameters.  The first method

       December 1993                           8-42       DRAFT-DO NOT QUOTE OR CITE
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 1     reproduced the two uptakes exactly; for the second, the simulation results deviated from the
 2     experimental values by an average of «23%.  Given the assumptions and results, predictions
 3     of total uptake are expected to be realistic, but predictions of respiratory tract dose
 4     distribution should be considered speculative.
 5          Using the human assumptions (Table 8-6), the bolus-response experiment of Hu et al.
 6     (1992b) was simulated.  Results for the experimental bolus fractional uptakes were good,
 7     over-predicting by «5%; the results for the remaining two experimental variables deviated
 8     from the experimental values by »30%. As a consequence of the assumptions and results,
 9     simulations with flow rates of «250 mL/sec are expected  to overpredict by no more than
10     « 5 %, and predictions of the distribution of dose within the respiratory are speculative.
11          Given the assumptions of Table 8-6, Table 8-7 shows a comparison between
12     experimental total RT uptake efficiency data and model predictions for humans.  Use of the
13     rat assumptions in conjunction with the model will be discussed later (Section 8.4.3).
14          The model predictions show good agreement  with the total RT uptake efficiency data of
15     Gerrity et al.  (1988), Gerrity et al. (1993a), and Hu et al.  (1992b).  In all cases, the
16     predictions are within 10% of the measured  values. The agreement with the data of Hu
17     et al. (1992b) is  even better.
18          The model prediction for the data of Wiester et al. (1993) is less accurate.  Comparison
19     of the Wiester et al. (1993) data with the Hu et al. (1993) data (Figure 8-1) show, however,
20     that they are in good agreement with each other.  It would thus appear the VT dependence of
21     the model does not necessarily reflect the real world.  However, the general agreement
22     between  the model predictions  and data are quite good.
23          Although the models are capable of making reasonable predictions of total RT uptake
24     efficiency, their  accuracy on the regional level remains uncertain.  The O3 bolus data (Hu
25     et al., 1992b, 1993; Ultman et al., 1993); and the  airway  uptake efficiency data (Gerrity
26     et al., 1993b) bring  into question the model predictions about uptake in the conducting
27     airways.  These  latter data sets suggest that the models of Miller et al.  (1985, 1988) may
28     underestimate the O3 uptake coefficients in the conducting airways.  Ultman et al. (1993)
29     show in their analysis of the bolus data that  the reactivity of O3 with the lung liquid lining
30     decreases with increasing depth into the lung.  This could  imply that more O3 is taken up in
31     central airways than had previously been thought.  The predictions presented in Table 8-7

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    TABLE 8-6.  ASSUMPTION FOR APPLICATION OF DOSIMETRY MODEL
             TO BREATHING FREQUENCY RESPONSES TO OZONE
Species
 Mode of
Breathing
  Respiratory Tract Morphology
      Mass Transfer Coefficients
Human   Oral
Rat
Nasal
 LRT structure from Weibel (1963)
 Volume of oral cavity through
 larynx and surface to volume
 ratio (S/V) from Hu et al.
 (1992a,b)
 Dead space volume (Vj) and FRC
 from Hart et al. (1963); TB
 region volume at PRC equals V^
 minus oropharyngeal volume
 Proximal  alveolar region defined as
 first respiratory bronchiole
 Pulmonary region expands;
 tracheobronchial (TB)
 region does not expand

• Nasopharyngeal (NP) dimensions
 from  Schreider and Raabe (1981)
 TB region from Yeh et al. (1979)
 Volumes  and surface areas of LRT
 isotropically scaled to FRC
• Pulmonary region from Mercer
 et al. (1991)
• TB and pulmonary regions
 uniformly expand during
 breathing
- Proximal alveolar region is first
 generation of pulmonary region
                                           Mass transfer coefficients for each
                                           oropharyngeal segment and each TB
                                           generation defined as Ka/(S/V) where
                                           S/V for the TB region is from Weibel
                                           (1963) dimensions reduced to FRC.
                                           Ka from Hu et al. (1992b)
                                           Pulmonary mass transfer coefficient is
                                           0.10 cm/a (Miller et al. [1985])
NP and TB mass transfer coefficients
estimated using data of Hatch et al.
 (1989)
Mass transfer coefficient of pulmonary
region = 0.137 cm/s; inferred from
Pinkerton et al. (1992)
     TABLE 8-7. COMPARISON OF TOTAL RESPIRATORY TRACT UPTAKE
                        DATA WITH MODEL PREDICTIONS
VT(mL)
832
832
500
1,000
1,650
1,239
631
fB (bpm)
12
24
15
7.5
1,360
1,360
16
Measured Ft
0.97
0.96
0.86
0.93
0.88
0.87
0.76
Predicted Ft
0.96
0.93
0.89
0.94
0.95
0.93
0.94
Data Source
Gerrity et al. (1988)
Gerrity et al. (1988)
Hu et al. (1992b)
Hu et al. (1992b)
Gerrity et al. (1993a)
Gerrity et al. (1993a)
Wiester et al. (1993)
December 1993
                                8-44
                                DRAFT-DO NOT QUOTE OR CITE
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  1     based on the assumptions of Table 8-6, however, represent a revision of the Miller et al.
  2     (1985) model in that the mass transfer coefficients are derived from the actual human data of
  3     Hu et al. (1992b).
  4          Because the utility of dosimetry models is in their ability to facilitate interspecies
  5     extrapolation, it is important to compare predictions with animal data as well as human data.
  6     Using the Overton et al. (1987) model formulation and parameters, Overton et al. (1989)
  7     developed a formula that can be used to calculate respiratory tract uptakes in rats, given their
  8     VTs, and fBs.  For comparison purposes, the data of Wiester et al. (1987, 1988) and Hatch
  9     et al. (1989) can be used.  The average uptake efficiency for the rat from these data is 0.45.
10     Based on the VTs and fBs of these animals, an average LRT uptake of 0.61 is computed
11     using Overton et al.  (1989).  If Overton et al. (1989)  had included the effects of URT
12     uptake, their model would have predicted  more than 0.61.
13
14
15     8.3  SPECIES SENSITIVITY: LUNG  FUNCTION AND
16           INFLAMMATORY ENDPOINTS EXEMPLIFYING AN APPROACH
17     8.3.1   Introduction
18          Quantitative extrapolation of animal-based O3 toxicity data to the human circumstance
19     requires a paradigm that includes both an estimate of target tissue dose (dosimetry) and an
20     algorithm which relates the responsiveness of the test  species to that of the human (species
21     sensitivity).  This paradigm can be depicted as an extrapolation parallelogram (Figure 8-6)
22     which conceptualizes a relationship between chronic animal study data and long-term human
23     health effects based upon understanding of acute effects in both  species (Graham and Hatch,
24     1984).  Though recent  studies have begun to elucidate the underlying  mechanisms
25     determining response, the bulk of the present O3 toxicity data base in animals and humans
26     remains largely descriptive.  Hence,  only a simplified application of this paradigm is feasible
27     at this time.  The following section will attempt to harmonize a  selective literature on acute
28     human and animal responses to O3 exposure  (already reviewed in detail in Chapters 6 and 7)
29     with what is known about the dosimetry of O3 in an effort to discern relative species
30     sensitivity.  To construct an argument that is plausible for this test application, focus is on
31     endpoints for which there are sufficient data in both humans and test animal species and for

       December 1993                          g-45      DRAFT-DO NOT QUOTE OR CITE
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                            Acute Effect
                                Human
                                                                      Chronic Effect
                                                                          Human
        Acute Effect
           Animal
                                                   Chronic Effect
                                                       Animal
         —	  Experimental correlation
         	   Extrapolation
       Figure 8-6.  Parallelogram paradigm for utilizing animal data for human health
                   predictions.  Acute homologous endpoints serve as the basis for
                   extrapolating chronic effects hi humans from animal data.
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
which exposure scenarios are similar. The endpoints compared include measures of
pulmonary function and markers of lung inflammation, most notably bronchoalveolar (BAL)
protein and cells.  When possible, other influencing parameters,  such as ventilation
augmentation and antioxidants within the lung will also be discussed.  The body of in vitro
cell studies has not been included because  of the difficulties in interpretation associated with
dosimetry and culturing systems.  The reader is referred to Section 7.2.5 and the recent
review by Koren et al. (1993). The goal here is to develop a hypothetical model for the
assessment of species specific sensitivity with acute O3 exposure that can serve as a
framework to better predict human responses, especially with regard to chronic effects. The
complex issue of whether controlled human clinical studies accurately reflect population-
based responses also will not be considered in this discussion.
       December 1993
                                         8-46
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  1      8.3.1.1   Dosimetry
  2           This topic has been discussed in detail in Section 8.2.  Recent studies by Hatch et al.
  3      (1993) utilizing the nonradioactive isotope of oxygen, 18O, to label O3, have shown that
  4      exposure of exercising humans (60 L/min) and resting rats to 0.4 ppm for 2 h resulted in
                       18
  5      3 or 4 times the   O dose (as adduct) to the BAL constituents of humans as compared to
  6      those of F344 male rats. This 3- to 4-fold difference appeared to be due to the exercise-
  7      stimulated hyperventilation of the humans when  compared to the rat and compared favorably
  8      with indices of effect (i.e., BAL cells and protein at 24 h).  Only when the rats were
  9      exposed to 2 ppm for 2 h did their 18O3 labeling of BAL constituents approximate that of the
 10      human.  Thus, on the basis of this study of cellular and protein influx due to O3 injury, the
 11      rat and human appear to have similar sensitivity  to O3 when exercise is considered.
                                    1 8
 12      Additional related studies with   O3 indicate that deposition in the respiratory tract is a
 13      cumulative function of ventilation over the initial period of exposure which would lend
 14      support to these findings (Santrock et al., 1989).  Attempts to compare animal data obtained
 15      without exercise to human study data with exercise would thus underestimate the dose to the
 16      lung and presumably the resultant risk of effect.  Studies of O3 effects (but without
 17      assessment of dose) in exercising rodents have confirmed this conclusion (Mautz et al.,
 18      1985).
 19           Exercise is only one factor that can alter dose and effect.  Studies in laboratory animals
 20      which incorporate other factors,  such as time of day  or diurnal rhythms (Van Bree et al.,
 21      1992), animal strain (Pino et al., 1991;  Costa et al.,  1993a), or nutrition (Section 6.2.5),
 22      also show substantial modification of response to O3, and thus emphasize the need for careful
 23      consideration of exogenous factors when attempting to compare or extrapolate study findings.
 24      It is likely that a similar range of factor-dependent variability exists within human test
 25      subjects.
 26
 27      8.3.2   Homology of Response
28          The concept of species sensitivity actually consists of two integrated components.   The
29      first, homology of response, indicates whether the outcome seen in the animal test species
30      represents the  same biological response in the human.  In many cases, a measurement of the
31      same endpoint in both species can be presumed to reflect the same toxic phenomenon and/or

        December 1993                           3.47      DRAFT-DO NOT QUOTE OR CITE
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 1     mechanism (i.e., the pulmonary irritant-induced tachypnea) (see below).  On the other hand,
 2     there may be endpoints which, while homologous, are not expressed similarly.  One such
 3     example would be the burning discomfort of sensory irritation in the human and the pause on
 4     tidal expiration seen in rodents. The second component of species sensitivity relates the
 5     dose-response curve for given homologous responses. Alterations in permeability of the air-
 6     blood barrier of the lung appear to reflect true species differences in sensitivity to pulmonary
 7     irritants such as O3 (Hatch et al., 1986).  Ideally, these elements of species sensitivity should
 8     flow directly into extrapolation formulae developed to integrate animal and human research
 9     data.
10
11     8.3.2.1  Lung Function Endpoints as Homologous Indicators
12          Lung function studies of small mammals have provided basic physiological information
13     important to the understanding of both normal and diseased lungs (Snider and Sherter, 1977;
14     Harkema et al., 1982; Raub  et al., 1982; Mauderly,  1984; Costa, 1985).  Animal lung
15     function tests, adapted from  those used clinically, have proven useful in describing the nature
16     and severity of lung injury as well as distinguishing toxicant-induced effects in the central or
17     peripheral airways from those that exist in the parenchyma.  In practice, the interpretation of
18     functional changes detected in animals derives from knowledge and experience in human
19     pulmonary medicine.  Supporting this view in theory is the allometric data base in normal
20     mammals in which the lung  function variables associated with ventilation and aerobic
21     metabolism scale systematically to body mass over nearly seven orders of magnitude (Stahl,
22     1967; Leith,  1976). The lung function studies of O3 toxicity hi animals and humans,
23     considered in the present discussion, are described in detail in Sections 6.2.5 and 7.2 of the
24     current document or in the previous O3 criteria document (U.S. Environmental Protection
25     Agency, 1986).
26
27     8.3.2.2   Inflammatory and Antioxidant Endpoints as Homologous Indicators
28           Inflammation of pulmonary airways and airspaces is best described as a cascade of
29     events networking infiltrating leukocytes, plasma proteins, and cell-derived mediators which
30     function presumably to defend or repair (but may further damage) the injured lung (see
31     Sections 6.2.2 and 7.2.5; Koren et al., 1993).  Key  markers of the basic inflammatory

       December 1993                          8-48       DRAFT-DO NOT QUOTE OR CITE
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 1     process include plasma-derived proteins (including, albumin, globulins, plasmin, etc.) and a
 2     primary inflammatory cell, the polymorphonuclear leukocyte (PMN).  Of these two markers,
 3     plasma-derived proteins in the acute phase are generally thought to represent a "leak" from
 4     the vasculature to the airspace lumen. Hence, under controlled temporal conditions, plasma
 5     protein residing in the airspace and accessible by BAL can be used as a proportional marker
 6     of effect, which in turn should be related to dose. The presence of PMNs in the airspaces is
 7     a bit more complex because of the signals involved in recruiting these cells into the lung
 8     lumen after injury and  the cascade of events apparently involved in their poesis from the
 9     vasculature to the lung lumen. For the purposes of species comparison, plasma-derived
10     protein (nonspecific) and the proportion of PMNs among total cells  as sampled by BAL will
11     be emphasized as primary indices of damage and inflammation within the lung.
12          Antioxidant substances in lung tissue (Slade et al., 1985) and BAL fluid and cells
13     (Slade et al., 1993) have been identified and quantified for humans and several laboratory
14     animal species. The species profiles of these antioxidants in the lung tissue and their
15     respective BAL cells and fluid can differ appreciably (Table 8-8), but collectively they
16     appear to play a significant role in defense of the lung against both endogenous and
17     exogenous oxidant challenge.  Ascorbate and vitamin E in particular appear to have major
18     functions in protecting  the lung from O3 challenge (Section 6.2.6; Slade et  al., 1989;
19     Crissman et al., 1993;  Koren et al.,  1989b; Elsayed et al., 1988) and when their levels are
20     manipulated in vivo, either can influence the degree of toxic outcome.  Hence, the
21     measurement of basal and O3 response levels of these antioxidants in BAL  cells and fluids is
22     useful in assessing the  qualitative and quantitative responses among  humans and laboratory
23     test species.
24
25     8.3.3   Studies of Lung Function
26     8.3.3.1   Confounding Influences in Lung Function Studies
27          Ideally, a system  for measuring pulmonary function  in small animals would
28     approximate that used in humans with cooperative, unrestrained  subjects. However, in
29     animal studies, this is usually not possible. Fortunately, certain measures (e.g., static lung
30     volumes, diffusion capacity) appear to be minimally influenced by sensory reflex or muscular
31     activity in  spite of unnatural stresses  or blunting of responses caused by anesthetic or

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             TABLE 8-8. PULMONARY ANTIOXIDANT SUBSTANCES IN VARIOUS
                       LABORATORY ANIMAL SPECIES AND HUMANS8
Mouse
Ascorbate
tissue 41 ±4
BAL cells0 —
BAL fluid0 -
Glutathione
tissueb 62 ±3
BAL cells0 -
BAL fluidc —
Tocopherol
tissue 1.0 ±0.1
BAL cellsd -
BAL fluidd -
Uric Acid
tissue —
BAL cells0 -
BAL fluid0 -
Hamster Rat
26±2 34±2
— 50.3 ±5.4
— 199.4±9.1
61 ±2 50±2
— 14.8±2.7
- 12.1 ±5.0
1.0±0.1 2.1±0.1
- 577.7 ±83.1
- 0.6 ±0.2
— 0.35 ±0.05
- <0.01
- 4.3 ±0.6
Guinea Pig Rabbit
39±1 27 ±3
17.9±1.4 —
28.8±2.2 —
83 ±3 83 ±3
14.6±2.4 —
li.2±1.9 —
2.0±0.2 1.4±0.2
454.5 ±58.2 —
1.4±0.5 —
4-.14±0.24 -
0.8 ±0.1 —
2.7 ±0.4 -
Human
22 ±7
3.5±0.1
21.4±2.8
7±1
2.9±0.5
20.4±3.8
0.8±0.1
95.1 ±23.4
47.2±3.8
_
0.07 ±0.03
15.9±2.5
       aData (mean ± SE) extracted and summarized from Slade et al. (1985, 1993).
       Data expressed as mg/100 g wet tissue.
       Data expresses as nmol/mg protein.
       Data expressed as nmol/mg lipid phosphorus.
 1     physical immobilization. On the other hand, some measurements, typically those involved in
 2     the assessment of ventilatory mechanics, can be profoundly influenced by these and other
 3     factors, such as ambient and toxicant-altered body core temperature, thus confounding
 4     cross-species comparisons.  Because a major emphasis of this section is the comparison of
 5     lung function data of animals and humans, it is important that the reader realize potentially
 6     confounding influences borne by studies of lung function in rodents when compared to
 7     analogous measurements in humans.  These  are discussed briefly below.
 8
 9     Anesthesia
10          Anesthesia alters pulmonary function measurements in both humans (Rehder et al.,
11     1975) and laboratory animals (Skornick and  Brain, 1990; Lamm et al.,  1982; Rich et al.,
       December 1993                          8-50      DRAFT-DO NOT QUOTE OR CITE
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 1      1979).  In general, ventilation is reduced and changes in ventilatory patterns occur (Pavlin
 2      and Hornbein,  1986; Belville et al., I960; Hunter et al., 1968; Siafakas et al.,  1983).
 3      In humans, anesthesia can decrease compliance and functional residual capacity (FRC), and it
 4      can also increase airway resistance (Rehder et al., 1974, 1975).  In small laboratory
 5      mammals, an analogous decrease in FRC occurs, though apparently via a different
 6      physiological mechanism (Lamm et al.,  1982).  Additional anesthesia-related effects include a
 7      blockade of irritant reflexes (Weissberg  et al., 1976) and alteration of ventilatory patterns in
 8      response to carbon dioxide (CO2) (Martin-Body and Sinclair, 1985).  Hence, while not
 9      invalidating experimental results, choice of anesthetic agent may affect the measured
10      response and may confound cross-species comparison.
11
12      Restraint
13           To enable collection of small animal pulmonary function data without the use of
14      anesthesia, some type of physical immobilization is usually required.  Restraint may range
15      from minimally restrictive, allowing turning and some locomotion, to extremely confining,
16      as occurs when animals are inserted into nose-only exposure tubes. Although restraint
17      reduces movement artifacts and permits  attachment of delicate probes or sensors,
18      immobilization can also produce undesirable physiological disturbances such as changes in
19      body core temperature (Nagasaka et al., 1979), hypermetabolism (Nagasaka et al., 1980),
20      increased expiratory CO2 (Jaeger and Gearhart, 1982), changes in ventilation and ventilatory
21      pattern (Lai et  al., 1978; Mauderly, 1986) and gastric lesions (Toraason et al.,  1980).  Such
22      stress-related responses are poorly  understood and their influence on toxicologic responses
23      may well pass unnoticed unless specifically examined.
24
25      Temperature
26           Although not widely appreciated, toxicant-induced changes in thermoregulatory function
27      can modify the results of toxicological studies (Gordon et al., 1988; Gordon, 1991). Recent
28      studies indicate that exposure to 0.37, 0.5  and 1.0 ppm O3 also can decrease core
29      temperature (Tco), heart rate, and blood pressure over 2 or more hours in unrestrained,
30      unanesthetized  rodents maintained at normal  room temperature (Uchiyama et al., 1986;
31      Watkinson et al., 1992).  On the other hand,  when rats were restrained in a head-out body

        December 1993                           8-51       DRAFT-DO NOT QUOTE OR CITE
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 1     plethysmograph and exposed to the same concentration of O3 (1.0 ppm), as in the Uchiyama
 2     et al. (1986) and Watkinson et al. (1992) studies, no change in blood pressure was observed
 3     (Tepper et al., 1990).  The discordance between these findings may be the result of restraint
 4     stress which has been shown to increase Tco (Nagasaka et al., 1979) and in this circumstance
 5     could have blunted the decrease in Tco associated with O3 exposure.
 6          While O3-induced changes in heart rate and Tco may be unique to rodents, this
 7     phenomenon has not been well studied in humans. It is possible that because of their larger
 8     thermal mass and different thermoregulatory mechanisms, humans do not exhibit similar
 9     changes in these parameters upon exposure. For example, rectal temperature increased by
10     the same amount in both air and  0.4 ppm O3 groups of humans during a 2-h exposure at
11     35 °C (Bedi et al., 1982).  The effects on Tco may have been confounded because the
12     subjects performed moderate exercise during alternate 15 min periods during exposure.
13     On the other hand, women intermittently exercising in moderate (24 °C) and hot (35  °C)
14     ambient conditions showed no change in Tco related to O3 exposure, but did show less of an
15     increase in heart rate (2.7%) than did air-exposed (8.1 %) subjects at 35 °C (Gibbons and
16     Adams, 1984).  It should be noted, however, that several studies have  shown potentiation of
17     human lung  function responses associated with increased ambient temperature and
18     O3 exposure (Folinsbee et al., 1977; Gibbons and Adams, 1984).  The full importance of
19     temperature  in relating rodent and human responsiveness to O3 remains to be understood.
20
21     Exercise and Ventilation
22          Exercise has long been employed in human studies to enhance the effects of air
23     pollutants, especially O3 (Folinsbee and Raven,  1984). Exercise appears to exacerbate
24     functional effects by increasing the inhaled dose (Hatch et al., 1993) and possibly by shifting
25     the deposition of the pollutant to more sensitive  pulmonary sites (Gerrity and Wiester, 1987).
26     While exercise can be used in laboratory animals to enhance deposition of O3, no direct
27     methods for measuring ventilation or breathing mechanics are available for small animals
28     during exercise. Alternatively in an attempt to mimic the increase in ventilation produced by
29     exercise in humans, studies employing restrained animals have used CO2 as a ventilatory
30     stimulant.  Carbon dioxide (8-10%) maximally increases minute ventilation (VE) 3 to 5 times
31     in rodent species; CO2 in excess of 10% will result in a reduction in ventilation (Wong and

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 1     Alarie, 1982; Tepper et al., 1988). This increase in VE is equivalent to light (2 to 3 x
 2     resting VE) or moderate exercise (4 to 6 x VE) in humans (U.S. Environmental Protection
 3     Agency,  1986).  In many O3 studies in humans,  both heavy  (7 to 8  x VE) or very heavy
 4     (>9 x VE) exercise has been used.  Similar increases in ventilation cannot be attained in
 5     small animals using the CO2 challenge technique, thus posing a limitation for animal studies
 6     attempting to make direct comparisons with human studies.
 7          The application of this CO2-challenge methodology in O3-exposed rats (0.25 to 1.0 ppm
 8     for 2.25  h with 15 min alternating hyperventilation) clearly demonstrates enhanced
 9     pulmonary irritation as reflected in breathing pattern changes during exposure (Tepper et al.,
10     1988, 1990). The breathing pattern alterations typical of O3 exposure appeared to be larger
11     than would be predicted based solely  on increased dose, suggesting that CO2 challenge
12     during O3 exposure may have enhanced deposition at critical lung sites (Tepper et al., 1989).
13     This augmented response was clearly reflected in the large increases  in protein observed in
14     the BAL (Costa et al.,  1988b).  In postmortem studies, rats  exercised during exposure have
15     been found to have exacerbated lung pathology, thus appearing to confirm this hypothesis
16     (Mautz et al., 1985) and suggesting that exercise  may, in fact, enhance toxicity
17     disproportionate to the apparent dose of toxicant.
18
19     8.3.3.2   Acute Exposure Data
20           Two corners of the parallelogram paradigm can be readily constructed from data
21     gathered in empirical studies of acute O3 exposure in humans and laboratory animals.  These
22     studies have the bulk of the data with the highest frequency  of common  endpoints which can
23     be compared.  Hence,  the following discussion will focus on several categories of
24     homologous lung function and BAL study-data which have been obtained from humans and
25     animals exposed similarly to O3.  The human studies were drawn from the large existing data
26     base on lung function and represent typical  responses.  The  corresponding BAL data are
27     more limited and are used to the extent possible.   In contrast, the animal studies  selected for
28     comparison  are highly  selective and represent  a rather small data base involving similar
29     exposure scenarios and homologous endpoints. This approach, of necessity, excludes  the
30     large majority of animal studies, not because they do not contain important toxicologic data
        December 1993                           8-53       DRAFT-DO NOT QUOTE OR CITE
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 1     on O3, but rather, they are disparate in their exposure parameters or biologic endpoints
 2     which can be readily tied to those available in humans.
 3
 4     Tidal Breathing
 5          In humans, O3 produces pulmonary irritation, a response associated with cough and
 6     substernal soreness (Chapter 7).  While these symptoms are difficult to assess in animals,
 7     exposure to sufficient concentrations of O3 produces reflex alterations in tidal breathing that
 8     can be objectively measured.  Most notably, the response is an increase in  fB that is usually
 9     accompanied  by a decrease in VT (tachypnea), whereas VE may not be altered.  Although
10     the magnitude of the tachypneic response is  variable depending on the species and exposure
11     conditions, this endpoint is quite sensitive and consistent across many species (e.g., guinea
12     pigs, cats, dogs, rats, monkeys, and humans).
13          To examine the cross-species response to O3, data from human and animal studies
14     reporting immediate postexposure alterations in fB were evaluated.  Most human studies
15     employed an  exercise regimen during O3 exposure to increase dose. On the other hand, few
16     animal studies have used exercise, relying rather on high exposure concentrations or
17     CO2-induced hyperventilation. Three representative human studies were selected because
18     they used a large range of concentrations and ventilation rates.  Selected data from  these
19     three human  studies are compared to the available animal data in Figure 8-7 and discussed
20     further below.
21          Folinsbee et al. (1975) examined the upper limits of the O3 concentration-response
22     curve in human subjects exposed either at rest or while performing light/moderate
23     intermittent exercise (29 L/min;  »5x resting VE). Measurements  of fB were obtained during
24     exercise following a 2-h exposure to 0.37, 0.5, or 0.75 ppm O3.  Concentration-dependent
25     increases in fB were observed in both the resting and exercise group, although the magnitude
26     of the change in the exercise group was greater.  The role of exercise in altering the
27     O3-induced changes in fB was examined further by DeLucia and Adams (1977) who exposed
28     humans to 0.15 or 0.3 ppm for 1 h while the subjects exercised at one of four ventilation
29     levels (1 x, 3x, 4x and 6x resting VE).  The magnitude of fB response increased with
30     concentration and exercise level, but was only significant in the 6x exercise group at
31     0.15 and 0.3 ppm.  Lastly, the lower limits of the concentration-response were explored by

       December 1993                          8-54      DRAFT-DO NOT QUOTE OR CITE
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               160
               150
            <§  140
e 130
o>

I120
            •B  110

            |
               100
            LL
                90
                80
            0.2     0.4     0.6     0.8      1

                          Ozone (ppm)
                                                              1.2
                                                                 Human, Rest, 2
                                                               Folinsbeeetal. (1975)
                                                                 Human, Light, 2
                                                               Folinsbeeetal. (1975)

                                                                	H	

                                                                Human, Heavy, 1
                                                            DeLucia and Adams (1977)
                                                                         Human, Very Heavy, 2.5
                                                                         McDonnell etal. (1983)
                                                                           Guinea Pig, Rest, 2
                                                                           Amdur etal. (1978)
                                                               Guinea Pig, Rest, 2
                                                               Murphy etal. (1964)
                                                                	-A-	

                                                                Rat, Light, 2.25
                                                               Tapper etal. (1990)


                                                                  Rat, Rest, 3
                                                              Mautz and Bufalino (1989)
           1.4
      Figure 8-7.  Comparison of changes in frequency of breathing after O3 exposure in
                   humans and animals.  Data are expressed as percent of the control
                   response.  Right hand legend indicates species, exercise level, exposure
                   duration in hours, and on the second line, the reference.  Human data are
                   plotted with solid lines and open symbols, while animal data are plotted
                   with broken lines (differentiated by species) and closed symbols.
1     McDonnell et al. (1983) where subjects performing very heavy exercise (65 L/min; »10 x

2     resting VE), were exposed to 0.12, 0.18,  0.24, 0.30 or 0.40 ppm O3 for 2.5 h.  Significant

3     changes in fB were observed at all exposure levels.

4          Although several animal  studies have evaluated tidal breathing changes during and after

5     O3 exposure, only four studies have examined multiple concentrations such that comparisons

6     to human data can be made.  Unanesthetized, restrained guinea pigs were exposed for 2 h to

7     0.34, 0.68,  1.08, or 1.34 ppm O3 via nose cones, while tidal breathing was measured using

8     a constant volume plethysmograph (Murphy et al., 1964).  A similar experimental
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 1     preparation was used by Amdur et al. (1978) to evaluate the respiratory response of guinea
 2     pigs to 0.2, 0.4 and 0.8 ppm O3.  In both of these experiments, a monotonic increase in
 3     fB was observed; however, the animals studied by Murphy et al.  (1964) were uniformly more
 4     sensitive to O3 than those of Amdur et al. (1978).  Mautz and Bufalino (1989) measured
 5     breathing patterns in awake restrained rats exposed for 3-h to 0.2, 0.4, 0.6, and 0.8 ppm O3.
 6     Concentration-related increases in fB were observed up to 0.6 ppm, but the responses to
 7     0.6 and 0.8 were the same. In another study, awake rats were exposed to 0.12, 0.25, 0.5,
 8     and 1.0 ppm O3 for 2.25 h in head-out pressure plethysmographs where CO2-stimulated
 9     breathing was incorporated to augment ventilation (Tepper et al., 1990).  With the added
10     CO2,  rats appeared to be of similar responsiveness to O3 as were guinea pigs. In general, as
11     depicted in Figure 8-7, restrained guinea pigs and rats appeared to be as responsive as the
12     lightly exercising humans, and clearly more responsive than the humans exposed at rest.
13     Only  with strenuous exercise does the response of humans appear to exceed that of rodents.
14          In addition to similar concentration-related effects in humans and animals, the time-
15     related effects of O3 exposure appear to be similar.  To demonstrate this homology,
16     Mauderly  (1984) compared the time course of response to O3 in  humans and guinea pigs
17     exposed under somewhat similar conditions.  Humans were exposed to 0.75 ppm O3 for
18     2 h while engaging  in nonstrenuous, intermittent exercise at 15 min intervals (Folinsbee
19     et al., 1975). In another study, respiratory parameters were measured at 30 min intervals
20     during and for 4 h postexposure  (Bates and Hazucha, 1973). Similarly, unanesthetized,
21     restrained guinea pigs were exposed to 0.68 ppm O3 for 2 h as part of a concentration-
22     response study (described above), with respiratory function assessed at 15 min intervals
23     during and for 3.5 h postexposure (Murphy et al.,  1964).  In both guinea pigs and humans,
24     fB increased and VT decreased; both parameters  then returned toward control values during
25     the postexposure period.  The percent change from  control in fB and VT was nearly the
26     same, throughout the exposure and postexposure periods, indicating that a similar
27     concentration of O3 («0.7 ppm) produced similar temporal alterations in ventilation.  Again,
28     the guinea pigs would appear to  be slightly more responsive than humans since they were
29     exposed to a lower concentration (0.68 ppm) at rest, while the humans were exposed to
30     0.75  ppm with light intermittent exercise.
31

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 1     Mechanics
 2          Breathing mechanics have been examined in several animal and human-O3 exposure
 3     studies, but there is little similarity between the data bases in the concentrations or specific
 4     techniques used.  Bates et al. (1972) examined breathing mechanics in resting and in lightly
 5     exercised (2 x resting VE) humans exposed for 2 h to 0.75 ppm O3.  Although no
 6     concentration-response data were obtained, increased lung resistance (R^ and decreased
 7     dynamic compliance (Cdyn) were found for both resting (+22 and  -12%) and exercising
 8     (+67 and -51%) subjects exposed to O3.  In a similar study, Hazucha et al. (1989)  exposed
 9     men for 2 h to 0.5 ppm O3 using intermittent moderate (40 L/min) exercise, and  found a
10     significant 12.5% increase in airway resistance (RaW), although no concomitant change in
11     Cdya was detected.  McDonnell et al. (1983), using a broad range of O^ concentrations
12     (0.12, 0.18, 0.24, 0.3 and 0.4 ppm; 2 h) and very heavy exercise (65  L/min), reported
13     concentration-dependent increases in Raw.  In humans at rest or performing light exercise,
14     however, a 2-h exposure at near-ambient O3 concentrations would be expected to induce only
15     modest increases in R^ and no changes in Cdyn (Hazucha, 1987).
16          While relatively high O3 concentrations (> 1.0 ppm) showed effects on RL and Cdyn in
17     animals (Murphy et al., 1964), only three  studies  in animals have evaluated these parameters
18     at lower, more relevant O3 concentrations. Watanabe et al. (1973) studied anesthetized,
19     paralyzed,  and mechanically ventilated cats exposed via a steel tracheal tube to  either 0.25,
20     0.5 or 1.0 ppm O3 for between 2 and 6.5  h.  Measurements of breathing mechanics were
21     recorded every 30 min. With increasing O3 concentration and exposure duration, RL
22     increased and, to a lesser extent, Cdyn decreased.  Bronchoconstriction at 0.25 ppm O3 was
23     reversed following atropine (a parasympathetic receptor blocker), but only partially  reversed
24     at the two higher concentrations, suggesting the involvement of more than
25     bronchoconstriction in the increase in RL at these  levels.  Unfortunately, relating  the
26     concentrations used in this study to other animal or human studies  is difficult because
27     exposure through a tracheal tube would eliminate  scrubbing of O^  by the nasal and
28     oropharynx and would likely exaggerate the pulmonary O3 dose (Gerrity et al., 1988).
29          Other studies have attempted to examine breathing mechanics in unanesthetized animals
30     with natural nasal breathing and avoidance of potential anesthesia-related blunting of reflex
31     responses.   Murphy et al. (1964) exposed  unanesthetized guinea pigs to several

       December  1993                           8-57       DRAFT-DO NOT QUOTE OR CITE
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 1     concentrations of O3 for 2 h and measured ventilation, as previously discussed, and RL.
 2     At concentrations less than 1 ppm, O3 had no effect, but RL increased 48% and 147%  at
 3     1.08 and 1.34 ppm, respectively. Using a similar test system, Amdur et al. (1978) observed
 4     no significant alteration of RL in unanesthetized guinea pigs during a 2-h exposure to 0.2,
 5     0.4,  or 0.8 ppm O3. However, Cdyn decreased significantly at 0.4 and 0.8 ppm O3.
 6     In analogous studies in unanesthetized rats, Tepper et al. (1990) observed no significant
 7     changes in RL or Cdyn after a 2.25 h exposure to 0.12, 0.25, 0.5, or 1.0 ppm O3, in spite of
 8     intermittent 15 min periods of exercise-like hyperventilation induced by CO^.
 9          Although increased resistance is demonstrable in guinea pigs, cats, dogs and humans, a
10     comparison of percent change in resistance from control measurements after an acute («2 h)
11     O3 exposure (Figure 8-8) suggests that humans are more likely to bronchoconstrict due to
12     O3 exposure than rodents. Neither of the guinea pig studies (Murphy et al.,  1964; Amdur
13     et al., 1978) nor the rat study (Tepper et al., 1990) showed a significant increase in RL at
14     less than 1 ppm. However, closer examination of the human data reveals that the McDonnell
15     et al. (1983) study employed very heavy exercise, and most of the studies included in the
16     Hazucha (1987) model used moderate to heavy exercise.  Thus, again the inhaled dose  would
17     likely be greater than in spontaneously breathing animals.  A more comparable study in
18     humans that employed only light exercise reported that 0.25, 0.37 and 0.5 ppm for
19     2 h resulted in minimal, nonsignificant 118%, 124%, and  104% increase in R^ (Hackney
20     et al., 1975).  Likewise, the Bates et al.  (1972) data  obtained in subjects at rest and with
21     light exercise (0.75  ppm O3) also argue against an unusually high O3 responsiveness in
22     humans relative to test animals for this endpoint when exercise-related dose is considered.
23          In general, similar findings have been observed using the measurement of Cdyn;
24     however, the response decrements were more variable and of smaller magnitude. Given the
25     distal deposition of  O3, as indicated  by morphological studies (Section 6.2.4), it is surprising
26     that  so little attention has been given to this parameter.  Available data suggest that these
27     changes in Cdyn are of little biological significance for ambient exposures.
28
29     Airway Responsiveness
30          The ability of O3 to increase airway responsiveness to nonspecific bronchoconstricting
31     stimuli in both humans and other mammalian species has been known for at least a decade

       December 1993                           8-58       DRAFT-DO NOT QUOTE OR CITE
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               200
               180
            o 160
            E
            o
            O
               140
            to
            '(5
               120
               100
                80
                60
                        0.2       0.4       0.6       0.8

                                   Ozone (ppm)
                          Human, Very Heavy, 2.5
                          McDonnell at a). (1983)

                             Human, Rest, 2
                            Bates at al. (1972)

                                 A

                             Human, Light, 2
                            Bates etal. (1972)
                                 #•


                           Human, Combined, 2
                             Hazucha (1987)

                            Human, Light, 2
                          Hackney etal. (1975)

                            	H	

                              Cat, Rest, 2
                          Watanabe etal. (1973)
                                                                            Rat, Light, 2.25
                                                                           Tepperetal. (1990)
                                                                           Guinea Pig, Rest, 2
                                                                           Amduretal. (1978)
      Figure 8-8.  Comparison of changes in resistance after O3 exposure in humans and
                   animals. Data are expressed as percent of the control response. Right
                   hand legend indicates species, exercise level,  exposure duration in hours,
                   and on the second line, the reference. Human data are plotted with solid
                   lines and open symbols.  The line labelled, Hazucha (1987), is a model of
                   predicted response.  Animal data are plotted with dashed lines
                   (differentiated by species) and closed symbols.
1     (Sections 6.2.5 and 7.2.4).  However, airway responsiveness is perhaps the least understood

2     response to O3, particularly in the context of species comparisons.  Humans clearly exhibit

3     increases in airway responsiveness at environmental 03 exposure levels (Gong et al.,  1986;

4     McDonnell et al.,  1987; Folinsbee et al., 1988; Horstman et al., 1990), while analogous

5     responses in animals at O3 concentrations below 1 ppm are  controversial. Most studies of

6     airway responsiveness in laboratory animals focus on the development of asthma-like models

7     to elucidate generic mechanisms of airway responsiveness and utilize concentrations as high
      December 1993
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 1     as 3 ppm for brief periods of time to injure the airways.  Hence, anything more than a
 2     qualitative comparison between animal species and humans is tenuous and thus will not be
 3     further discussed in this section.  Details of the methodologies of the laboratory animal and
 4     human bronchoreactivity studies can be obtained elsewhere in the reviews of pulmonary
 5     function found in Sections 6.2.6 and 7.2.3, respectively.
 6
 7     Elasticity and Diffusion
 8          The integrity of the pulmonary air-blood barrier is essential for efficient exchange of
 9     oxygen (O^ and CO2.   This fragile epithelial interface with matrixed interstitial connective
10     tissues and capillaries possesses inherent elastic properties and presents a finite resistance to
11     O2 diffusion to the blood. Although the elastic and diffusionary properties of the blood-air
12     barrier are not implicitly linked to one another functionally, both properties can be readily
13     quantified and compared between humans and laboratory animals (Costa, 1985).  When
14     combined, assessment of these functional properties is often sufficient to evaluate pathologic
15     or toxic events  in the distal reaches of the  lung.  For this reason and the  fact that O3 deposits
16     in the deep lung, the effects of O3 on these parameters will be discussed together.
17          Inhaled O3 is known to penetrate to the depths of the lung and preferentially deposit in
18     the smallest airways and its proximal acini (Section 8.2).  Somewhat surprisingly, relatively
19     few studies in humans have sought to characterize potential functional impairments at the air-
20     blood interface. The reasons for this are likely two-fold: First, in the early studies of the
21     health effects of O3 on humans, static compliance and diffusing capacity of CO (DLcO) were
22     affected at only very high concentrations, well above what would be considered
23     environmentally relevant.  Second, from a practical perspective, these measurements proved
24     to be considerably more tedious to perform than the forced expiratory measurement, which
25     sensitively detects  O3-induced alterations (discussed below).  Nevertheless,  there are
26     sufficient data in humans exposed acutely to O3 to allow a reasonable comparison of these
27     endpoints with their more abundant animal homologs.
28          The earliest studies leave little doubt that O3 is edemagenic at high concentrations in
29     virtually all mammalian species. In the past, occupational exposures of 2 to 3 ppm C^ were
30     not uncommon, and a 9 ppm peak exposure has been reported  (Kleinfeld et al., 1957;
31     Challen et al.,  1958).  The resultant worker symptoms and signs, including chest radiograms,

       December 1993                           8-60      DRAFT-DO NOT QUOTE OR CITE
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  1      were consistent with the manifestations of edema  reported in experimental animals
  2      (i.e., increased lung weight and stainable edema in the airspaces) (Stokinger, 1965).  Lung
  3      function, however, was not typically measured in  these work-related exposures.  In a later
  4      study of arc welders exposed to 0.2 to 0.3 ppm O3, little, if any, convincing evidence of
  5      functional impairment, in terms of altered lung volumes, maximal expiratory flow rates, and
  6      DLco was obtained (Young et al., 1963). To further explore the possible effects of  O3,
  7      Young and coworkers (1964) subjected 11 human volunteers at rest to controlled atmospheres
  8      of 0.6 to 0.8 ppm O3 via mouthpiece for 2 h.  Small reductions in vital capacity (VC),
  9      forced expiratory volume in 0.75 s, dynamic and static  lung compliance, and intrapulmonary
10      gas distribution were observed, but only the 25 %  fall in Dl^o  proved to be statistically
11      significant.  Similar, but considerably more variable effects on  lung function were reported
12      by Hallett (1965) in 10  subjects exposed to 1 to 4 ppm for 30 min. Nonetheless, of
13      10 exposed subjects, 7 showed  at least a 20% drop in DLCO.  Like Young and coworkers,
14      Hallett interpreted these changes to indicate lung edema, in agreement with the hypothesis
15      that the deep lung irritant O3 was having its effect at the alveolar level.  Interestingly,
16      additional work from the same laboratory of the Young study (Bates et al., 1972) found that
17      resting subjects receiving nasal  exposure to 0.75 ppm O3 for 2  h resulted in a nonsignificant
18      3% reduction in DLCO. However, in a limited test group, the co-imposition of light
19      exercise, which doubled ventilation, enhanced this response («12%). It appears that the
20      nasal (Bates et al.,  1972) versus mouthpiece (Young et al., 1964) routes of exposure  were
21      instrumental in the differential response, because it is likely that the mouthpiece diminished
22      what scrubbing occurs when exposure is via the unencumbered  mouth in human test subjects
23      as reported by Gerrity et al. (1988).  Since these early studies,  there have been no additional
24      controlled human acute  studies that have examined alterations in DI^o at O3 concentrations
25      below 0.6 ppm.
26          Analogous animal  studies  of acute O3 exposure indicate that the general pattern of
27      functional impairment is similar to that reported in human studies. Anesthetized and
28      ventilated cats showed a general decline in VC,  static lung compliance, or DI^o with
29      exposures up to 6.5 h of 0.26 to 1.0 ppm O3 (Watanabe et al.,  1973). The responses of the
30      20 animals were variable, and these declines, which did not achieve overall statistical
31      significance, were thought to  be largely secondary to the substantial (36 to 200%) increases

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 1     in R^. In a more complex study design, rats were exposed for 2 or 7 h to 0.5 or 0.8 ppm
 2     O3 with intermittent 8% CO2 to hyperventilate (~2 to 3 X  resting VE) the animals as an
 3     exercise analogue to human exposures (Costa et al., 1988a).  The DLcO values were reduced
 4     by about 10% at both 0.5 ppm time-points and by about 12% with a 2-h exposure to
 5     0.8 ppm.  Exposure to 0.8 ppm for 7 h, however, greatly exacerbated the alveolar lesion
 6     with a resultant 40%  reduction in DL^.  Static compliance, unaffected by the other
 7     exposure conditions,  was affected only at this latter exposure duration.  These O3-induced
 8     effects, particularly the reductions in DLCO, appeared to correlate with the degree of lung
 9     edema in affected animals as had been surmised for the acutely exposed humans.  With the
10     multitude of more recent studies of O3 at ambient levels, alterations in static lung compliance
11     or DLCO are rarely reported in either humans or animals.
12
13     Forced Expiration
14          Reductions in FVC and FEVj have become the hallmarks of acute lung  dysfunction in
15     humans after O3 exposure (Chapter 7).  These measures are sensitive to O3 levels as low as
16     0.12 ppm for as little as 2 h when intermittent heavy exercise is included during  the exposure
17     (McDonnell et al., 1983) and show cumulative dysfunction resulting from 6.6 h of lower
18     levels of this oxidant (0.08 and 0.10 ppm) when nearly continuous, moderate  exercise is
19     employed (Horstman et al., 1990).  Reductions in the FEV1 and FVC induced by O3 are
20     believed to be partly  the result of pain-mediated interruption of maximal inspiration (Hazucha
21     et al.,  1989). Exactly what level of tissue injury or inflammation correlates with these
22     functional deficits is unclear and is an active area of research (Section 7.2.4).
23          Most studies of O3 in experimental animals make little effort to mimic human study
24     designs, thereby impeding the extrapolation of their results to humans.  Recently, however,
25     rat studies involving periods of intermittent CO2-induced hyperventilation to enhance
26     dosimetry, have attempted to capitalize on the qualitative similarity of the rat  and human
27     maximum expiratory flow volume (MEFV) curves as a potentially sensitive endpoint of
28     toxicity (Costa et al., 1988a; Tepper et al., 1989).  In the rat, FVC does indeed  decrease
29     with O3 exposure, though the magnitude of response is apparently less  than that observed in
30     humans.  As in the human, the reduced FVC in the animal  appears not to be  the result of a
31     change in residual volume (RV). Total lung capacity (TLC) may be reduced  slightly, but

       December 1993                          8-62      DRAFT-DO NOT QUOTE OR CITE
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 1     lung compliance does not change.  However, it is premature to assume a common
 2     mechanism for the FVC  reductions in the rat and human.  Unlike the human, pain on
 3     inspiration in the animal  model is likely not an issue since the animal is anesthetized during
 4     the procedure and is brought to TLC by a defined airway pressure (»30 cm H2O).  Since
 5     general anesthesia is known to diminish sensory afferent stimuli, an analogous O3-induced
 6     fall in rat FVC  should expectedly have been blunted, if not totally eliminated.  To what
 7     extent anesthesia mitigates the rat response or that there are inherent species differences in
 8     dosimetry or sensitivity is not clear from these studies. Nevertheless, comparison of model-
 9     predicted FVC changes in humans (Hazucha, 1987) with analogous rat data (Costa et al.,
10     1988a) would suggest that this response in the anesthetized rat is about half that of the human
11     (Figure 8-9)
12
13     Studies of Inflammation and Antioxidant Content of Bronchoalveolar Lavage Fluid
14          Both humans and animals exhibit a PMN inflammatory response with associated
15     changes in lung permeability after acute exposure to O3.  Recent studies indicate that humans
16     exposed to concentrations as low as 0.08 ppm for 6.6 h with moderate exercise (40 L/min)
17     exhibit a fourfold increase in the percentage PMNs when BAL is obtained 18-h postexposure
18     (Devlin et al., 1993).  To date, animal studies at comparable exposure levels are rare,
19     (Hotchkiss et al.,  1989) and exercise enhancement of exposure dose has yet to be
20     incorporated. As noted above,  the issue of dosimetry is critical if extrapolation at such levels
21     is to be attempted. Nevertheless, in the one acute rat study at 0.12 ppm for 6 h, an increase
22     in nasal lavage derived PMNs was noted 18 h postexposure with no similar change in PMN
23     number in the BAL (Hotchkiss et al., 1989).  In contrast, in the same study when higher
24     concentrations of O3 (0.8 and 1.5 ppm) were used, BAL PMNs were elevated, but no
25     changes were observed in the nose washings.  Such a "competetive" nasal-pulmonary
26     response  has yet to be directly studied in humans.  Nevertheless, the data support the general
27     hypothesis that there is comparability between inflammatory responsiveness of rats and
28     humans.  More  direct comparison of laboratory animal with human inflammatory responses
29     can be drawn from studies at higher concentrations when the nasal/lung competetive response
30     in the rat is skewed to  the lung and, like in the lung function comparison,  analogous
       December 1993                          3.53      DRAFT-DO NOT QUOTE OR CITE
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                                                              ••-•A
                                                                        Human, R»tt, 2 h
                                                                        Hazucha(1987)
                                                                      Huron, V«y Heavy, 2 h
                                                                        Hazucha (19
                                                                        Rat,Ught,4h
                                                                       Costa etaL(1988a)
                                                                         Rat,Ught,7h
                                                                       Costa etaL(1988a)
                                                                         Rat,Reit,8h
                                                                       OMM*tal.(1988a)
                                  0.4
 0.6     0.8
Ozone (ppm)
     1.2
       Figure 8-9.  Comparison of changes in forced vital capacity after O3 exposure in
                   humans and animals.  Data are expressed as percent of the control
                   response.  Right hand legend indicates species, exercise level, exposure
                   duration, and on the second line, the reference. Human data are plotted
                   with solid lines from the equation of Hazucha (1987), with the shaded area
                   representing the predicted range of decrements expected between light
                   exercise (top line) and very heavy exercise (bottom line). Rat data are
                   plotted with dashed lines and closed symbols.
 1      exposure conditions can be more directly compared. These studies are tabulated in

 2      Table 8-9 and discussed in more detail below.

 3           Four representative human studies of lung inflammatory responses after acute

 4      O3 exposure can be compared with existing acute animal data from studies of analogous

 5      design.  Seltzer et al. (1986) exposed moderately exercising (83 to 100 watts) subjects to

 6      0.4 or 0.6 ppm O3 for 2 h with BAL obtained 3 h postexposure.  The BAL (combined

 7      0.4 and 0.6 ppm groups) contained an average 7.8-fold more PMNs than after filtered air.

 8      Protein levels in the BAL were not assayed in this study.  In separate studies, humans were

 9      exposed to 0.4 ppm  for 2 h with heavy exercise (60 L/min) with BAL samples collected at

10      1 and 18 h postexposure (Devlin et al., 1993a; Koren et al., 1989b).  These two studies
       December 1993
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  1      showed that the inflammatory response is quickly initiated postexposure.  The PMN and
  2      protein content were elevated at both times (1 h:  PMN  18.2X; protein 1.2 x) with the
  3      18 h timepoint (PMN 8x; protein 2.2x) being higher for protein, but somewhat less for
  4      PMNs. Schelegle et al. (1991) followed the time course of the the inflammatory response
  5      (1-, 6-, and 24-h postexposure) after an exposure of 0.3 ppm for 1 h with exercise
  6      (60 L/min). The PMN content of the combined airway  and alveolar BAL was elevated at
  7      6 h (3X) and 24 h (2.5 x) only, with the 6-h point representing the peak. Protein  content of
  8      the BAL did not change significantly at any point.
  9          The many studies of O3-induced inflammation in laboratory animals are reviewed in
10      Section 6.2.3. For the purpose of species comparison, only selected studies (Table 8-9) will
11      be considered.  The spectrum of exposure conditions used in the various animal studies
12      makes difficult the direct comparison among laboratory test species.  However, one study by
13      Hatch  and coworkers (1986) specifically addressed this question by exposing mice, guinea
14      pigs, rats, rabbits, and hamsters under identical conditions (0.2, 0.5, 1.0, and 2.0 ppm for
15      4 h), followed at 18 h postexposure with BAL and assay for protein.  Guinea pigs  were  most
16      responsive, responding at 0.2 ppm, whereas mice, hamsters and rats began responding at
17      1.0 ppm and rabbits responded only to 2.0 ppm.  Only one study involving BAL assessment
18      of PMNs and protein in exposed monkeys has been published (Hyde et al., 1992).
19      At 0.96 ppm (8 h),  the monkeys had a significant inflammatory response, but it is difficult to
20      assess  monkey  responsiveness relative to the human  for this endoint. Assuming a linear
21      C x T relationship, the monkey data appear on the order of the guinea pig response.
22      However, none of these species showed BAL protein increases approximating those reported
23      in human studies.
24          In recent studies (0.4 ppm for 2 h,  with BAL 16 to 18 h postexposure) (Slade et al.,
25      1989;  Crissman et al., 1993), guinea pigs  made vitamin C-deficient exhibited enhanced
26      responsiveness to O3, making them comparable to that of the exercised humans of Koren
27      et al. (1989b) (Figure 8-10).  Similarly, when rats were  exposed to 0.5 ppm O3 for 2 h with
28      intermittent CO2-induced hyperventilation (Tepper et  al., 1993) to mimic mild-moderate
29      exercise (three- to fivefold VE), the BAL protein, as  well as PMN responses  at 18-h
30      postexposure compared favorably with those data of Koren and coworkers (1989a,b)
31      (Figure 8-11).

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     TABLE 8-9. POLYMORPHONUCLEAR LEUKOCYTE AND PROTEIN
           PERMEABILITY RESPONSE TO OZONE BY SPECIES

Exposure
Parameters
0.4-0.6
2h

0.4
2h

0.4
2h

0.3
Ih

0.96
8h



0.2
0.5
1.0
2.0
4h










0.5
1.0
4h



0.5
0.8
2or7h




Exposure
Conditions
Exercise
(15 min/alt;
83-100 W)
Exercise
(15 min/alt;
60 L/min)
Exercise
(15 min/alt;
60 L/min)
Exercise
(15 min/alt;
60 L/min)
Daytime;
Rest



Daytime;
Rest













Daytime;
Rest;
Vit. C
(AH2+/-)


Daytime;
(15 min/alt
for 2 h;
45 min/alt
for 7 h);
3-5 X VE


Species (Strain)
Human


Human


Human


Human


Rhesus monkey




(a) Mouse
(Swiss Albino)

(b) Guinea Pig
(Harley)

(c) Rat
(Sprague-Dawley)

(d) Hamster
(Golden Syrian)

(e) Rabbit
(NZW)
M
Guinea Pig
(Hartley)
M



Rat
(F344)
M



PMN Protein
Fold8 Folda
BAL Time Increase Increase
3 h 7.8 Not done


18 h 8.0 2.2


1 h 18.2 1.2


1,6, 24 h 3.0 at 6 h No change
2.5 at 24 h

1,12,24 27atlh 3 at 1 h
72, 168 h 19 at 12 h 3 at 12 h
24 at 24 h 8 at 24 h*
6 at 72 h 3 at 72 h
3 at 168 h 1 at 168 h
16 h Not done (a) 1.8 at 1.0 ppm
3.2 at 2.0 ppm

(b) 1.4 at 0.2 ppm
2.0 at 0.5 ppm
4.1 at 1.0 ppm
4.5 at 2.0 ppm

(c) 2.1 at 1.0 ppm
3.6 at 2.0 ppm

(d) 1.5 at 1.0 ppm
2.6 at 2.0 ppm

(e) 2.7 at 2.0 ppm
16 h Not done For 0.5 ppm;
1.1 for AH2+,
2.1 for AH2".
For 1.0 ppm;
2.4 for AH2+,
2.7 for AH2"
1 h Not done For 0.5 ppm;
1.2at6h,
2.1 at7h.
For 0.8 ppm;
1.5 at 2 h,
3.3 at 7 h


Reference
Seltzer et al.
(1986)

Koren et al.
(1989a,b)

Devlin et al.
(1990)

Schelegle
et al. (1991)

Hyde et al.
(1992)



Hatch et al.
(1986)













Slade et al.
(1989)




Costa et al.
(1988a)




"Fold represents (03 response/air response).

December 1993
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                          Basal Levels of Ascorbic Acid
              3500

              3000

            12500
            en

            :§ 2000

            | 1500

            «: 1000

               500
                     Humans
          Normal
              Guinea Pigs
           AH2-def
                    Effect of Ozone Inhalation (0.4 ppm, 2 hr)
                                  on BAL Protein
                                                             B
                                                       p- 0.004
                                           p - 0.025
                     Humans
Rats
Normal      AH2-def
    Guinea Pigs
Figure 8-10. Composite of data from Slade et al. (1989), Koren et al. (1989b), and
            Crissman et al. (1993) comparing basal BAL ascorbate levels (A) to
            O3-induced changes in BAL protein (B). Ozone-exposures (0.4 ppm; 2 h;
            16 to 18 h BAL) of humans (exposed with exercise), rats (exposed resting),
            and guinea pigs (exposed resting) with (AH2-def) and without (normal)
            ascorbic acid  deficiency.
December 1993
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                                                1        1.5
                                            Ozone (ppm)
       Figure 8-11.  Composite of data comparing BAL PMNs 16 to 18 h after O3 exposure
                    (0.4 ppm; 2 h) of humans (with exercise) (Korea et al., 1989b) to those of
                    rats exposed to O3 (0.5 to 2.0 ppm) at rest or hyperventilated with CO2
                    (Tepper et al., 1993).
 1          Within a given laboratory animal species, responses among strains can also differ
 2     appreciably, as recently demonstrated in rats by Pino et al. (1991) and Costa et al. (1993a).
 3     These studies indicated that Wistar rats exhibit greater inflammatory responses (protein and
 4     PMN) to O3 than Sprague-Dawley and Fischer-344 rats after an 8-h exposure to 0.5, 1.0,
 5     and 1.5 ppm with BAL sampled at 2 or 24 h later.  Similarly, mice strains (C3H/HeJ and
 6     B6C57/6) (Kleeberger et al., 1990) and Sprague-Dawley rat substrains (Costa et al., 1985)
 7     have been shown to possess specific genetic susceptibility to high levels of O3 (2 ppm).
 8     In the case  of the mice, the responsive strain is 7 times (at the 6-h postexposure peak) as
 9     susceptible  as the resistant strain for the PMN response to 2 ppm; the protein response is
10     twice (at the 24-h postexposure) that of the resistant strain.  In the Sprague-Dawley substrain,
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 1     protein extravasation into the alveolar lumen immediately postexposure is 40% higher in the
 2     responsive strain than the resistant (no other time points were examined).
 3           Humans have an order of magnitude less ascorbate in BAL fluid as compared to the rat,
 4     but they have nearly twice  the glutathione, 4 times the uric acid, and 80 times the vitamin E
 5     (as normalized to lipid P -  surfactant) (Slade et al., 1993) (Table 8-8). However,  on a BAL
 6     cell protein basis, the ratios clearly favor the rat for all of these antioxidants,  with the
 7     exception of uric acid which is generally not high in rats because of species differences in
 8     protein and prime nucleotide catabolism (urea being the major N-by-product with rat).   The
 9     guinea pig most closely resembles the human for ascorbate.  Since these antioxidants are
10     thought to function in the defense against oxidant challenge, it would appear critical to
11     appreciate their presence and function when attempting to interpret data for extrapolation.
12     Unfortunately, the data base in both animals and, particularly, humans with regard to these
13     antioxidants is quite limited.  Supplementation and deprivation studies with vitamins C  and E
14     have  shown that these antioxidants have some role  in protecting against the effects of O3 in
15     both animals (Elsayed et al., 1988; Slade et al.,  1989; Crissman et al., 1993)  and  in humans
16     (Chatham et al.,  1987).   Of the animal models, ascorbate-deprived guinea pigs appear to
17     have  BAL ascorbate levels  most like humans, with a protein permeability response (without
18     exercise in the animal) very similar to the human exposed to the same concentration
19     (0.4 ppm for 2 h) with exercise.  However, Crissman et al. (1993) also reported that
20     reduced-ascobate levels in BAL (18 h postexposure) increase in  the human,  while BAL
21     reduced-ascorbate levels in the rat decrease.  Whether this relates to the distinctly different
22     basal levels of this vitamin and is associated  with the disparate protein responsiveness
23     (ignoring exercise) is unclear because deficiency in animals (guinea pigs) appears more
24     critical to the responsiveness at low ambient-like concentrations  than at higher concentrations
25     (1.0 ppm).  While it would appear that vitamin C is involved in the interplay  between O3 and
26     the exposed subject (human and animal), there is not full coherence of the data. For
27     example, Hatch et al. (1986) showed that the rabbit was the least responsive to 03 in terms
28     of BAL protein, but this  species has among the lowest tissue levels of vitamin C (Slade
29     et al., 1985).  However,  rabbits apparently have  a low propensity to form lipid peroxides
30     (Arakawa et al.,  1986), an  expected product  of lung lipid-O3 interaction (Pryor, 1992).
       December 1993                           g_69       DRAFT-DO NOT QUOTE OR CITE
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 1     Thus, to interpret interspecies sensitivity only in terms of basal antioxidant levels, however
 2     templing this may be, would be premature and overly simplistic at this time.
 3
 4     8.3.3.3   Prolonged Exposure Studies
 5          The previous sections of this text have utilized lung function responses to acute
 6     O3 exposure in an attempt to elucidate the relative sensitivity among laboratory test species
 7     and humans and thereby complete two corners of the parallelogram paradigm.  Because acute
 8     responses represent only part of the extrapolation paradigm, temporal-based exposure
 9     responsiveness will  also be considered, despite the relative paucity of comparative data
10     between laboratory  animals and humans in that context.  Again, the criteria for selection of
11     studies was similarity in exposure scenario in an effort to best extact species-specific
12     response differences.  The discussion  will focus on relative adaptability of acute functional
13     changes and associated BAL-derived findings after repeated exposures and the coherence of
14     the findings from prolonged human  and animal exposure studies and epidemiological results.
15
16     Lung Function Studies
17          Reversal ("attenuation") of pulmonary function decrements using a scenario of repeated
18     exposure to O3 has  been reported for  both humans and laboratory animals.   At least 9 studies
19     between 1977 and  1984 have documented that, for repeated exposures between 0.2  and
20     0.5 ppm O3 (2 h/day, up to 5 days), spirometric changes were most severe on the first or
21     second day of exposure, waned over the next 3 days of exposure and by the fifth day, had
22     returned to control pre-exposure levels (Section 7.2.1.4). In the only animal study  using a
23     similar exposure protocol and analogous experimental design, Tepper et al.  (1989) showed
24     that rats initially displayed a tachyneic response to O3 that attenuated after  5 consecutive days
25     of exposure, a pattern quite similar  to that of humans.  Exposures were for 2.25 h and
26     included challenge with CO2 during alternate 15 min periods to augment ventilation (2 to
27     3 X resting VE - equivalent to light exercise in humans).  As in the human studies, the
28     functional changes were largest on Day 1 or 2, depending on the parameter and the
29     O3 concentration (0.35, 0.5 and 1.0 ppm).  Attenuation of the shape constant of the flow-
30     volume curve of the rats was also observed over this period.  Thus, under analogous
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 1     conditions of exposure, both the humans and the rats exhibited similar initial functional
 2     responses to O3 with full and kinetically similar reversal of effects.
 3          More difficult is the direct comparison between human and animal lung function
 4     responses to prolonged (several weeks) O3 exposure, largely because of the limited
 5     availability of controlled human study data.  In the only study of its kind, Bennett (1962)
 6     exposed 12 human subjects at rest to 0.2 or 0.5 ppm for 3 h/day, 6 days/week for
 7     12 consecutive weeks.   Although no effects were discernable early in the exposure, there
 8     appeared to be a small, but significant O3-induced reductions in FEVj (and small,
 9     nonsignificant reductions in FVC), particularly in the last weeks of the study.  This reduction
10     in FEVj would more likely reflect obstructive changes within the lung at these points late in
11     time rather than the pain-mediated reductions that are seen with acute O3, which adapt away
12     after a few days of exposure.  The lack of concentration-related decrements in FEVl  and
13     FVC is somewhat unsettling, but regardless, after 9 postexposure weeks in clean air,  all
14     measured effects had reversed.
15           Unfortunately,  there are no directly parallel animal studies to compare to this limited
16     data base.  But, if the 2-fold sensitivity difference between the rat to human FVC response
17     (see Figure 8-12) is assumed, a number of animal studies may be considered comparable for
18     the purposes of this discussion.  On the one hand, rats exposed to 0.2 or 0.8 ppm O3 for
19     6 h/day, 5  days/week for 12  weeks were reported to exhibit some degree of small airway
20     obstruction based on the MEFV curves, but little, if any reduction in FVC or Dl^o was
21     observed (Costa et al.,  1983).  Others have reported analogous marginal increases in  rat TLC
22     or its component volumes (Bartlett et  al., 1974; Costa et al., 1983; Raub et al., 1983) or in
23     regional airway resistances (Yokoyama et al.,  1984)  after intermittent or continuous
24     exposures to >0.25  ppm O3 for 4 to  12 weeks,  which would not be unexpected with distal
25     airway  or lung damage.  Actual pathology in the distal lung tends to be focal and difficult to
26     correlate precisely with the marginal functional impairment.
27          The limited functional data available in monkeys generally agree with the pattern of
28     distal lung pathophysiology reported in rats.  When exposed to 0.5 ppm for 90 days
29     (8 h/day), monkeys exhibited a slight  increase in lung distensibility (Eustis et al., 1981).
30     Likewise, monkeys exposed to 0.25 ppm (8 h/day for 18 months) exhibited increased  chest
-------
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  1      conditions of exposure, both the humans and the rats exhibited similar initial functional
  2      responses to O3 with full and kinetically similar reversal of effects.
  3           More difficult is  the direct comparison between human and animal lung function
  4      responses to prolonged (several weeks) O3 exposure, largely because of the limited
  5      availability of controlled human study data. In the only study of its kind, Bennett (1962)
  6      exposed 12  human subjects at rest to 0.2 or 0.5 ppm for 3 h/day, 6 days/week for
  7      12 consecutive weeks.  Although no effects were discernable early in the exposure, there
  8      appeared to be a small, but significant O3-induced reductions in FEV1 (and small,
  9      nonsignificant reductions in FVC), particularly in the last weeks of the study.  This reduction
10      in FEVi would more likely reflect obstructive changes within the lung at these points late in
11      time rather than the pain-mediated reductions that are seen with acute O3, which adapt away
12      after a few days of exposure.  The lack of concentration-related decrements in FEVl and
13      FVC is somewhat unsettling, but regardless, after 9 postexposure weeks in clean air, all
14      measured effects had reversed.
15           Unfortunately, there are no directly parallel animal studies to compare to this limited
16      data base.  But, if the  2-fold sensitivity difference between the rat to human FVC response
17      (see Figure  8-12) is assumed, a number of animal studies may be considered comparable for
18      the purposes of this discussion.  On the one hand, rats exposed to 0.2 or 0.8 ppm  O3 for
19      6 h/day, 5 days/week for  12 weeks were reported to exhibit some degree of small airway
20      obstruction based on the MEFV curves, but little, if any reduction in FVC or DLgQ was
21      observed (Costa et al., 1983).   Others have reported analogous marginal increases in rat TLC
22      or its component volumes (Bartlett et al., 1974; Costa et al.,  1983; Raub et al., 1983) or in
23      regional airway resistances (Yokoyama et al., 1984) after intermittent or continuous
24      exposures to >0.25 ppm O3 for 4 to  12 weeks, which would  not be unexpected with distal
25      airway or lung damage. Actual pathology in the distal lung tends to be focal and difficult to
26      correlate precisely with the marginal functional impairment.
27           The limited functional data available in monkeys generally agree with the pattern of
28      distal lung pathophysiology reported in rats.  When exposed to 0.5 ppm for 90 days
29      (8 h/day), monkeys exhibited a slight increase in lung distensibility (Eustis et al.,  1981).
30      Likewise, monkeys exposed  to 0.25 ppm (8 h/day for 18 months) exhibited increased chest
       December 1993                           g_71       DRAFT-DO NOT QUOTE OR CITE
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  1     Morphometric analyses of the end-airways and distal lung regions of O3 exposed monkeys
  2     consistently show altered cell profiles and interstitial restructuring, even when functional
  3     changes are marginal which, like in the rat, likely reflects the large functional reserve of the
  4     integrated lung.  Thus, while these collective data from subchronic animal studies suggest a
  5     reasonably homologous distal lung response to O3, many of these linkages in functional
  6     outcomes remain uncertain in terms of what to anticipate in the human response.
  7          Clearly, the question of potential lung impairment resulting from a near-lifetime
  8     exposure to O3 ranks among the most pressing concerns of this toxicant.  The animal data,
  9     while demonstrating that chronic O3 can induce changes in the structure and function of the
 10     lung, have yet to provide definitive evidence of potential disease in disability in humans
 11     exposed to O3 over prolonged periods of their lives.  The  existing epidemiologic studies
 12     (Chapter 7;  Section 7.4.2), too, merely provide suggestive evidence that persistent or
 13     progressive deterioration in lung function may be associated with long-term oxidant pollutant
 14     exposure (Detels et al., 1981, 1987). Most recently, Detels and coworkers (1991) reported
 15     decrements in FEV] and nitrogen washout across all age groups in areas where oxidant
 16     pollution is high.  Similarly,  analysis of the pulmonary function data from the National
 17     Health and Nutrition Examination Survey (NHANES H) showed loss of lung function when
 18     annual averages of ambient O3  exceeded 0.04 ppm (Schwartz, 1989).  This pattern of
 19     impairment is consistent qualitatively with the chronic animal studies (Costa et al., 1993b).
 20
 21      Studies of Inflammation and Antioxidant Content of the Bronchoalveolar Lavage Fluid
 22          The virtual absence of human  BAL study  data after repeated or prolonged exposures to
 23      O3 hinders the comparison of non acute human and animal inflammatory responses.
 24     However, the recent study of Devlin et al. (1993b) suggests that the PMN and protein
 25      responses to repeated daily exposures to 0.4 ppm  (2 h with intermittent exercise for
 26      5 consecutive days) attenuate, much as do the functional responses.  Hence, for these BAL
 27      parameters (exceptions - e.g., lactate dehydrogenase activity - a marker of cell injury), there
 28      is indeed an apparent reversal phenomenon for several of the components of acute
 29      inflammation when exposures are continued over the 5-day exposure period.  Rat studies
30      largely appear to show similar attenuation to O3, but this response seems to be influenced by
31      exposure patterns or conditions  (Bassett et al.,  1988; Tepper et al., 1989;  Van Bree et al.,

        December 1993                           8.73       DRAFT-DO NOT QUOTE OR CITE
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 1     1989).  The study with the most similar design to the human protocol (Tepper et al., 1989)
 2     showed some reduction in BAL protein with repeated exposure involving intermittent
 3     hyperventilation with CO2, but over the 5-day period,  the protein levels remained
 4     significantly elevated^ cells were not evaluated in this study.  Interestingly, vitamin C and
 5     glutathione levels in the BAL fluid increased over the 5-day course of exposure, a response
 6     consistent with an upregulated antioxidant role in the adaptative mechanism.  Van Bree and
 7     coworkers (1992), on the other hand, reported that 5 consecutive days (12 h/day) of 0.4 ppm
 8     O3 resulted in complete reversal of both BAL albumin and PMN measures to the control
 9     values.  It should be noted,  however, that other biomarkers and mediators within the BAL
10     were not fully recovered which might suggest a slower reversed time frame or continued
11     O3-induced pathogenesis, a conclusion of the Tepper et al. (1989) study.
12           In guinea pigs made deficient in vitamin C and exposed to O3 (0.2, 0.4, or 0.8 ppm)
13     continuously for 7 days, attenuation of the functional and inflammatory endpoints appeared
14     nearly complete in  spite of the deficiency  (Kodavanti et al., 1993). Other antioxidants, not
15     altered basally, were upregulated more by the O3 challenge; the small residual reservoirs of
16     ascorbate, which persisted in the nearly 98% deficiency state of the animals, were apparently
17     mobilized to the site of injury allowing repair to proceed.  Likewise, chronically exposed rats
18     have elevated BAL ascorbate indicative of the oxidant burden and the ongoing repair (Grose
19     et al., 1988). Prolonged exposures up to  18 mo appear to sustain a low-grade interseptal
20     inflammation and evidence of lung matrix remodeling in both rats and monkeys, suggesting
21     that humans would behave similarly.  However,  such data are not presently available from
22     humans.
23
24     8.4   QUANTITATIVE EXTRAPOLATION
25     8.4.1   Introduction
26          Advances in dosimetry since the previous O3 criteria document (U.S. Environmental
27     Protection Agency,  1986) fall into five major areas: (1) greater sophistication of model
28     applications (e.g., Overton et al., 1989; Mercer  et al., 1991), (2) appearance of experimental
29     uptake data that can be compared to model predictions (Wiester et al.,  1987,  1988, 1993;
30     Hatch et al.,  1989, Gerrity et al.,  1988, 1993a,b), (3) experiments specifically designed to
31     estimate model parameters (Hu et al.,  1992b), (4) a better understanding of the role of O3 in

       December 1993                           8-74      DRAFT-DO NOT QUOTE OR CITE
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  1      the liquid linings and tissues of the respiratory tract (Pryor, 1992), and (5) better
  2      understanding of anatomical aspects (Mercer et al.,  1991).  The role of these advances in
  3      interspecies dosimetric extrapolation follows.
  4           With the information available for rats, reasonably reliable predictions of the flux of
  5      O3 to the air-liquid lining interface of lexicologically important regions, such as the
  6      centriacinar region, is possible.  There are two main investigations that make this feasible:
  7      (1) Hatch et al. (1989), who estimated the percent uptake and the fraction of the retained
  8      O3 that is in the URT, trachea, and lung of rats, and (2) Pinkerton et al.  (1992) (with
  9      elaboration by Miller et al., 1993) who illustrate the basic correctness of modeling
 10      assumptions for ventilatory units. (The judgement of basic correctness  is based on the
 11      assumption that the dose causing the response is proportional to the flux of O3  to the air-
 12      liquid lining interface.)  Using this information, regional mass  transfer coefficients could be
 13      estimated which would allow the prediction of local respiratory tract O3 doses in rats exposed
 14      under general conditions.
 15           The results of the investigation of Hu et al. (1992b) can be  used to estimate URT and
 16      tracheobronchial region model parameters, but may  not be sensitive enough to determine
 17      pulmonary region parameters.  If uptake is not confined to the  URT and tracheobronchial
 18      regions, then their mass transfer coefficients alone would not be sufficient to account for total
 19      RT uptake (e.g., as measured by Wiester et al., 1993 or Gerrity et al.,  1988), and the
 20      difference in predicted  (without pulmonary region uptake) and  experimental uptakes could be
 21      used to estimate the pulmonary region mass transfer coefficient.  Unfortunately, no dosimetry
 22      data for the human pulmonary region are available.
 23           Despite limitations, the O3 dosimetry data that have been  obtained over the past several
 24      years, coupled with the advances in modeling,  suggest that there has been continual
 25      convergence between the model predictions and experimental observations.  Given the many
 26      areas of consistency between models and experiments, it is valuable to begin to employ these
 27      models to provide the dosimetric basis for animal-to-human extrapolation.  One of the
 28      greatest sources of uncertainty in such an application of dosimetry models is the lack of full
29      understanding of the appropriate O3 target site (e.g., upper/lower airways, airways,
30      pulmonary region) from which a particular response is initiated (especially for functional
31      endpoints). However, reasonable assumptions can be made to narrow the target site.

        December 1993                           8.75       DRAFT-DO NOT QUOTE OR CITE
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 1          The first application of dosimetry models given in Section 8.4.2 is an examination of
 2     delivered dose versus response within a given species.  This is followed in Section 8.4.3 by
 3     some interspecies comparisons of delivered dose versus response.
 4
 5     8.4.2    Intraspecies Delivered Dose Response
 6          Assuming that changes in FEVj in humans exposed to O3 are mainly the result of
 7     O3 pulmonary tissue dose, Miller et al. (1988) constructed a dose-response curve. They
 8     plotted decrements in FEV| versus predicted cumulative pulmonary region O3 tissue dose
 9     scaled to body mass  (Figure 8-13).  The concentration-response data are from McDonnell
10     et al. (1983), in which 135  healthy subjects  were exposed to 0.0, 0.12, 0.18, 0.24, 0.3, and
11     0.4 ppm O3. Exposure was for 2.5 h  with intermittent heavy exercise.  Miller et al. (1988)
12     used the average weight and height of the subjects to estimate FRC that was used in the
13     model to simulate O3 dose.   The exercise breathing parameter data were used along with an
14     estimate of resting breathing parameters.  Figure 8-13 is similar in shape to the
15     concentration-response curve of McDonnell  et al. (1983).  Differences between these two
16     curves,  however,  are accounted for by the translation between exposure concentration and
17     O3 dose.
18          Another example of the use of computed delivered dose to interpret intraspecies
19     response is provided by the work of Pinkerton et al. (1992) and Miller and Conolly (1993).
20     Pinkerton et al. (1992) examined the relation between actual tissue response of rats
21     chronically exposed to O3 and a prediction of O3 dose  as a function of distance from the
22     bronchiole-alveolar duct junction to ventilatory units.  Using these data, Miller and Conolly
23     (1993) plotted the predicted O3 dose and the observed change in wall thickness due to the
24     exposure versus distance from the bronchiole-alveolar duct junction (Figure 8-14). Even
25     though considerable variability in the thickness change can be inferred from the data, the two
26     curves,  scaled to their values at the junction, show a remarkable similarity and suggest a
27     basic correctness in regards to the ventilatory unit model parameters.
28
29     8.4.3    Interspecies Delivered Dose Response
30          The illustrations presented in this section are based on dosimetric estimates  for humans
31     and rats based on the existing or currently modified theoretical  models  (Miller et al., 1985;

       December 1993                           8-76       DRAFT-DO NOT QUOTE OR CITE
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         1.0
         0.9-
         0.8-
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         0.6-
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 Pulmonary Tissue Dose (ng Qj /g body weight)
 1
20
 1
22
                              24
      Figure 8-13.  Changes in FEVt versus pulmonary tissue dose.  Plotted are decrements in
                   FEVj (liters) for human subjects against predicted pulmonary tissue dose
                   normalized to body weight.  In order, from left to right, the dose values
                   correspond to 0, 0.12, 0.18, 0.24, 0.3, and 0.4 ppm O3 exposure
                   concentrations. The continuous curve was an "eye fit".
      Source:  Miller et al. (1988)
1     Miller et al., 1988; Overton et al., 1987).  One functional and one inflammatory endpoint
2     will be provided drawing from the fB and BAL protein data described in Section 8.3.
3     Because the diversity of exposure scenarios across species is so great, the window of
4     exposure parameters has been narrowed to minimize exposure-based differences in relating
5     species responsiveness.
6          Since the theoretical  models developed by Miller et al. (1985, 1988) and Overton et al.
7     (1987) estimate the dose distribution to the respiratory tract on a per breath basis, a
      December 1993
                      8-77
DRAFT-DO NOT QUOTE OR CITE
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                           Dose
                    "T
                                                I	1	T
                   100    200   300   400    500    600   700   800
                    Distance from Bronchiolar-Alveolar Duct Junction, urn
      Figure 8-14. Relationship between change hi alveolar wall thickness and predicted
                   O3 dose as a function of distance from the bronchiole-alveolar duct
                   junction (BADJ).  Rats were exposed to 0.98 ppm for 8 h/day for 90 days.
      Source: Miller and Conolly (1993).
1     minimum quantitative description of tidal breathing (VT and fB) is needed to utilize the
2     theoretical models of O3 deposition.  The need for detailed breathing parameters, therefore,
3     severely restricts the application of the model to studies providing such data.  Unfortunately,
4     breath-by-breath parameters over the the course of an exposure are not normally measured or
5     reported in most publications. Two studies, each  in humans and rats, providing adequate
6     detail over the course of the exposure allowed the  model computations to proceed for this
7     illustration (Figure 8-15).  The human studies examined were DeLucia and Adams (1977),
8     who exposed the subjects to 0.15 and 0,3 ppm O3 for 1 h with continuous exercise (65%
9     Vo max) periods and Beckett et al. (1985), who exposed the subjects to 0.4 ppm O3 with
      December 1993
                                      8-78
DRAFT-DO NOT QUOTE OR CITE
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  1      alternating 15 min exercise (70 L/min).  The rat studies evaluated were Tepper et al. (1989),
  2      who exposed the rats to 0.35,  0.5, and 1.0 ppm O3 for 2.25 h with alternating 15-min
  3      periods of CO2-induced hyperventilation (2 to 3 X resting VE) and Mautz and Bufalino
  4      (1989), who exposed the rats to 0.8 ppm O3 for 3 h (at rest).  The response parameter was
  5      the ratio of O3-altered fB to control fB (fO3 / fcnt). Dose rate was the average dose to the
  6      proximal alveolar region (PAR) computed  from time 0 to the time of the maximum fo3 / fcnt.
  7      The PAR was chosen as the target based on the perception that O3 acts on deep lung stretch
  8      receptors directly or indirectly (via tissue fluid pressure—edema) as the stimulus for
  9      tachypnea.  A number of assumptions were necessary to  implement the deposition model for
10      comparison of the human and rat in this context. These  are noted in Table 8-6.  Though
11      admittedly not validated, Figure 8-15 would suggest that the response of the rats not only
12      exceeded that of the human, but was initiated  at a lower dose rate to the targeted lung
13      region.  This conclusion is in general agreement with that emanating from Figure 8-7, but
14      the dose-to-target approach appears to accentuate the apparent difference in sensitivity for
15      this endpoint in favor of the rat.
16          Using an analysis similar to that applied  above for fB, Miller et al. (1988) related
17      pulmonary tissue dose (normalized to body weight) to BAL protein from rats, guinea pigs,
18      and rabbits (Hatch et al., 1986) for O3 concentrations of 0 to 2.0 ppm for 4 h. These values
19      have been  supplemented in Figure 8-16 to  include the results of the human  study of Koren
20      et al. (1989).  As can be seen  in the illustration, there appears to be a log-normal relationship
21      between BAL protein (/-eg/ml) and dose to the  pulmonary region, the purported site of plasma
22      leakage to  the airspace lumen.  This relationship would support the contention that there is a
23      mechanistic consistency  in response across species which may exhibit a quantitative
24      sensitivity factor for use in further  quantitative interspecies extrapolation. This sensitivity
25      factor is evident from the clustering together of data from different animal species.
26
27
28      8.5   SUMMARY AND CONCLUSIONS
29      8.5.1    Ozone Dosimetry
30          There have been significant advances in O3 dosimetry since publication of the previous
31      O3 criteria document (U.S. Environmental  Protection Agency, 1986) that better enable

        December  1993                           g-79      DRAFT-DO NOT QUOTE OR CITE
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     • DeLucia and Adams (1977)
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     D Tepperetal., (1989)
     + Mautz and Bufalino (1989)
               0.0       0.2       0.4        0.6        0.8       1.0       1.2
                            Predicted average PAR dose rate (ng Qj/crrrVmin)
                                                                            1.4
      Figure 8-15.  The maximum ratio of O3-altered breathing frequency to control breathing
                   frequency (fO3 / fcnt) at various O3 concentrations versus predicted
                   average dose rate to the proximal alveolar region (PAR first generation
                   distal to terminal bronchioles).  The average is for the tune from the
                   initiation of exposure to the time of occurrence of the maximum ratio.
                   Beckett et al. (1985) and Mautz and Bufalino (1989) provided sufficient
                   ventilatory data for only one O3 concentration.
1     quantitative extrapolation with marked reductions in uncertainty. Prior to 1986, there were

2     limited data on O3 uptake in laboratory animals (Yokoyama and Frank, 1972; Miller et al.,

3     1979), essentially no reliable data in humans (Clamann and Bancroft, 1959; Hallett, 1965),

4     only one realistic model of Oj dose (Miller et al., 1978,  1985), and no data on O3 reaction

5     kinetics in lung lining fluids.  At the present time, data gaps in all of these areas have begun

6     to fill in.  Experiments  and  models describing the uptake efficiency and delivered dose of
      December 1993
                                      8-80
DRAFT-DO NOT QUOTE OR CITE
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                             20      40      60      80      100     120     140    160
                               Pulmonary Tissue Dose (ng Qj /g body weight)
      Figure 8-16.  Protein in the BAL for several laboratory animal species and humans as
                    related to the estimated pulmonary dose (normalized per g body weight).
      Source: Modified from Miller et al. (1988).
1      O3 in the RT of animals and humans are beginning to present a clearer picture than has
2     previously existed.
3          The total RT uptake efficiency of rats at rest is approximately 0.50 (Wiester et al.,
4     1987,  1988; Hatch et al., 1989). Data in excised rat lungs support these in vivo findings,
5     and further indicate that O3 uptake efficiency is chemical reaction-dependent (Postlethwait
6     et al.,  1993).  Within the total RT of the rat, 0.50 of the O3 taken up by the total RT is
7     removed in the head, 0.07 in the larynx/trachea, and 0.43 in the lungs (Hatch et al., 1989).
8     The regional uptake efficiency data in the rat have been useful in estimating O3 mass transfer
9     coefficients for the rat.
      December 1993
                                      8-81
DRAFT-DO NOT QUOTE OR CITE
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 1           Ozone dosimetry models require input of regional mass transfer coefficients. Limited
 2      studies have been conducted to quantitate the mass transfer coefficients of lung tissue directly
 3      using excised animal tissue. In pig and sheep tracheae, mass transfer coefficients were
 4      determined for unidirectional flow conditions and were found to be independent of flow,
 5      suggesting a lack of dependence of O3 uptake on gas-diffusion processes (Ben-Jebria et al.,
 6      1991) .  These findings contrast with Aharonson et al. (1974) who found that the mass
 7      transfer coefficient in dog nasopharynx increased as a function of increasing flow.
 8           In humans at rest, the total RT uptake efficiency is between 0.80 and 0.95  (Gerrity
 9      et al., 1988; Hu et al.,  1992; Wiester et al, 1993).  At VTs around 500 mL, total RT uptake
10      efficiency falls from about 0.9 to 0.75 as flow increases from 250 to 1,000 mL/s. As VT
11      increases, uptake efficiency increases and flow dependence lessens, suggesting that at high
12      VT, uptake may be gas  diffusion limited. At a VT around 1500 mL, total RT uptake falls
13      from 0.96 at a  flow of 250 mL/s to 0.92 at a flow of 1,000 mL/s.  The studies of Gerrity
14      et al., (1988) and Wiester et al., (1993) indicate that the mode of breathing (oral  versus nasal
15      versus oro-nasal) has little effect on URT or on total RT uptake efficiency. This  observation
16      is supported by experiments comparing pulmonary function response as a function of mode
17      of breathing (Adams et  al., 1989; Hynes et al.,  1988).  Uftman et al.  (1993), however, found
18      that URT uptake efficiency was lower when mouthpiece breathing was compared  with nasal
19      breathing.  One possible explanation of the discrepancy among the studies is that  a
20      mouthpiece may decrease URT uptake efficiency in comparison  with unencumbered
21      breathing.  The enhanced physiologic response to O3 shown by Adams et al. (1989) with
22      mouthpiece breathing  supports this concept.
23           To obtain data on  regional O3 uptake efficiency in humans, Gerrity et al. (1993b)
24      measured O3 concentrations at various anatomical sites (from the mouth to bronchus
25      intermedius) in spontaneously breathing  humans.  They found that the unidirectional uptake
26      efficiency of the trachea was similar to that of the sheep and pig trachea (Ben-Jebria et al.,
27      1991), suggesting a similar mass transfer coefficient behavior in the human trachea.  Gerrity
28      et al. (1993b) also found that the uptake efficiencies between the mouth and various
29      anatomical sites in the total RT agreed well with the O3 bolus data of Hu et al. (1992).  Both
30      the Hu et al. (1992) and Gerrity et al. (1993b) data indicate that the mass transfer
31      coefficients of  the large conducting airways are larger than had been previously thought.

        December 1993                          8-82      DRAFT-DO NOT QUOTE OR CITE
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 1           When all of the animal and human in vivo O3 uptake efficiency data are compared,
 2      there is a good degree of consistency across data sets.  This agreement raises the level of
 3      confidence with which these data sets can be used to  support dosimetric model formulations
 4           In the area of mathematical model formulation,  there have been several models
 5      developed since 1986. They can be grouped according to how transport and chemical
 6      reactions  are modeled:  instantaneous reactions or quasi-steady first order reactions.  The
 7      models (Overton et al., 1987;  Miller et al.,  1988; Overton et al., 1989; Hanna et al., 1989;
 8      Grotberg  et al., 1990) predict  that  net O3 dose to lung lining fluid plus tissue gradually
 9      decreases distally from the trachea toward the end of the tracheobronchial region, and then
10      rapidly decreases in the pulmonary region.  When the dose of O3 to lung tissue is computed
11      theoretically, it is low in the trachea, increases to a maximum in the terminal bronchioles of
12      the first generation  of the pulmonary region, and then decreases rapidly distally into the
13      pulmonary region.  The models also provide insight into the role that increased ventilation
14      plays in enhancing O3-induced responses.  The increased VT and flow, associated with
15      exercise in humans or CO2-stimulated ventilation increases in rats, shifts O3 dose further into
16      the periphery of the lung, causing  a disproportionate increase in distal lung dose. This
17      prediction is supported by the  data of Postlethwait et al. (1993) in  excised rat lungs, and Hu
18      et al. (1992) and Gerrity et al. (1993b) in human lungs.
19           Ozone dosimetry models have also enabled examination of regional dosimetry among
20      parallel and serial anatomical structures.  When an asymmetric lung morphology is used in
21      dosimetric models,  the variation of O3  dose among anatomically equivalent ventilatory units
22      as a function of path length from the trachea varies as much as six-fold (Overton et al.,
23      1989; Mercer et al., 1991; Mercer and Crapo, 1993).  This could have significant
24      implications for regional or localized damage to lung tissue.  Whereas the average lung dose
25      might be at a level that would be considered insignificant, local regions of the lung may
26      receive significantly higher than  average doses and therefore be at greater risk for chronic
27      effects.
28           Theoretical models have  also been applied to make predictions about delivered doses
29      from exposure scenarios  that are not  necessarily achievable experimentally.  Overton and
30      Graham (1989) and Miller and Overton (1989) have scaled the  human lung dimensions to
31      account for age variations.   They predicted that LRT  uptake efficiency is not sensitive to age

        December 1993                           g_83       DRAFT-DO NOT QUOTE OR CITE
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 1     at resting ventilation, but is age-dependent when exercise conditions are invoked.  The total
 2     quantity of O3 absorbed/minute is predicted to increase with age during heavy work or
 3     exercise.
 4
 5     8.5.2   Species Sensitivity
 6          Examining functional parameters measured analogously in humans and various animal
 7     species discloses remarkable similarity in acute O3-induced effects and dysfunctions.  The
 8     tachypneic response to this oxidant is clearly concentration-dependent in both humans and
 9     animals and shows parallel exacerbation when hyperventilation (e.g., exercise or CO2) is
10     superimposed.  Indeed, rodents appear to be slightly more responsive than humans in this
11     regard.  What is not known is whether this is evidence of pulmonary irritant sensitivity,
12     perhaps as a prelude to toxicity, or whether tachypnea is a defensive posture taken by the
13     respiratory system to minimize distal lung O3 deposition.  Airway or lung resistance in
14     humans is not appreciably affected by acute exposure to O3 except under conditions of heavy
15     exercise; animals appear to need high level exposures or special preparations which bypass
16     nasal scrubbing.  Dynamic lung compliance, on the other hand, tends to decrease across
17     species.  However, the evidence in both animals and humans is not as strong as one might
18     expect,  given the distal lung deposition of this poorly soluble oxidant.
19           Ozone-induced spirometric changes, the hallmark of response in humans, also occur in
20     exposed rats, though the relative responsiveness of these alterations in the rodent appears to
21     be about half that of the human. It is unclear, however, the degree to which anesthesia  (rat)
22     and the comparability of hyperventilation induced  by CO2 (rat) or exercise (human) may
23     influence this difference in responsiveness.  Collectively, the acute functional response of
24     laboratory animals to O3 appears quite homologous to that of the human. Likewise, the
25     studies of BAL constituents indicate that the influx of inflammatory cells and protein from
26     the serum are influenced by species, but perhaps to a less extent than by ventilation and
27     antioxidant status, since adjustment for these factors can modulate responses to approximate
28     animal responses to those  of humans. Unfortunately, these influential factors are rarely
29     measured and even less often, controlled.
30           When humans are exposed repeatedly for several consecutive days, lung function
31     decrements  subside, and normal spirometric parameters are regained.  This phenomenon of

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  1      functional attenuation has also been demonstrated in rats, not only in terms of spirometry,
  2      but also in terms of the classic tachypneic ventilatory response.  Full or partial attenuation of
  3      the BAL parameters appears to also to occur in both rats and humans, but exposure scenario
  4      appears to play a role.  This leads to the question of potential lung impairment resulting from
  5      a near-lifetime exposure to O3.  Existing epidemiologic studies provide only suggestive
  6      evidence that persistent or progressive deterioration in lung function is associated with long-
  7      term oxidant pollutant exposure.  These long-term effects are thought to be expressed in the
  8      form of maximum airflow or spirometric abnormalities,  but the foundation for this
  9      conclusion remains weak and hypothetical. Animal study data,  while suggesting that Oj has
10      effects on lung function at near-ambient levels, present a variable picture of response which
                                   /
11      may or may not relate to technical conditions of exposure or some other yet undiscovered
12      variable of response.  Thus, a cogent  interpretation of the animal findings as definitive
13      evidence of chronic deterioration of lung function would be difficult at this time.  However,
14      the subtle functional defects apparent after 12 to 18  mo of exposure and the detailed
15      morphometric assessments of the O3-induced lesions do appear consistent with the modicum
16      of studies focusing on long-term effects in human populations.  Based on the apparent
17      homology of these responses between  humans and laboratory animals, animal studies provide
18      a means to more directly assess such chronic health concerns.
19
20      8.5.3   Quantitative Extrapolation
21           The agreement between theoretical models of O3 uptake and  experimental
22      determinations of O3 uptake efficiency now provide a basis upon which responses may be
23      examined as a function of delivered O3 dose instead of exposure concentration of O3.
24      By examining responses as a function  of delivered dose,  the goal of quantitative extrapolation
25      between species  can be approached.
26           The use of delivered dose to investigate responses has been examined in two contexts:
27      intraspecies  comparisons and interspecies comparisons.  With respect to intraspecies
28      comparisons, Miller et al. (1988) assumed that the relevant dose mediating the human
29      pulmonary function response was the pulmonary  tissue dose. They then utilized the
30      breathing patterns, exposure concentrations, and pulmonary function responses from the
31      human  studies of McDonnell et al. (1983) to predict the dose-response.  They found that

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 1     there was general agreement between the shapes of the concentration-response curves and the
 2     dose-response curves and that differences could be accounted for by the translation between
 3     exposure concentration and O3 dose. In another example dealing with intraspecies
 4     comparisons, Miller and Conolly (1993) compared the distribution of predicted O3 tissue
 5     dose to a ventilatory unit in a rat as a function of distance from the bronchoalveolar duct
 6     junction with the distribution of alveolar wall thickening as a function of the same distance
 7     measure.  Miller and Conolly (1993) found remarkable consistency between the predicted
 8     dose distribution and the response distribution, (i.e., as predicted delivered dose declined,
 9     response declined).
10          In an attempt to make an interspecies comparison of dose and response, existing or
11     currently modified models of Miller et al. (1985), Miller et al. (1988), and Overton et al.
12     (1987) were used to predict doses among species for two difference types of responses.
13     In the first case, the tachypneic response  to O3 as a function of dose was analyzed.  The
14     maximum ratio of O3-altered fB to control fB was plotted as a function of the average
15     centriacinar dose over the period from the beginning of exposure to the point of maximum
16     fB ratio. Rat and human data were used  for this comparison.  It was found that at
17     comparable O3 doses, the responses of rats  greatly exceeded that of humans and were
18     initiated at lower doses.  By examining the dose-response instead of the concentration-
19     response,  the difference in tachypneic response between rats and  humans is magnified.
20     In another example, an analysis similar to Miller et al. (1988) was performed to examine
21     recovered BAL protein as a function of O3 dose to the pulmonary region.  The species
22     considered were the rat,  guinea pig, rabbit, and human. In all cases, the BAL protein
23     response followed a log-linear relationship,  suggesting a consistency of response across
24     species.  Yet the data from different species tended to cluster together, suggesting species-
25     specific sensitivity factors.
26
27     8.5.4   Conclusions
28           There is an emerging consistency among a variety of O3 dosimetry data sets and
29     between the experimental data and theoretical predictions of O3 dose.  The convergence of
30     experimental data with theoretical predictions lends a degree of confidence to the use of
31     theoretical models to predict total and regional O3 dose.  Data have been acquired in animals

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  1      and humans that provide confirmation of total RT uptake efficiency predictions and provide
  2      regional mass transfer coefficients that enhance the predictive capacity of theoretical models.
  3      It is clear that the total RT uptake efficiency of rat is considerably less than that of humans
  4      on a per breath basis.  This difference is still one that is not totally accounted for in
  5      theoretical models, but further refinements may narrow this discrepancy.  Both the
  6      theoretical models, and experimental data indicate that regional dose distribution and the
  7      dependence of this distribution on breathing pattern can account for the role exercise plays in
  8      enhancing acute O3-induced effects.
  9           Comparison of O3-induced responses among animal species show that the acute
10      functional and inflammatory responses of laboratory animals to O3 is quite homologous to
11      human responses, although the detailed regional dose-response relationship appears to be
12      species dependent.  At equivalent delivered doses to the centriacinar region, rat tachypneic
13      responses are considerably larger than humans.  Inflammatory responses (influx of cells and
14      protein) may be less species dependent.  On an exposure concentration basis, the relative
15      spirometric response of rats is less  than humans by about one-half.  A detailed dose-response
16      comparison has not been done, but differences in regional delivered dose could account for
17      the difference.
18           Laboratory animals and  humans demonstrate similar attenuation of functional and
19      inflammatory responses with repeated exposure to O3, raising the question  of potential lung
20      impairment in humans resulting from near life-time exposure.
21           There are  still uncertainties that remain to be resolved before precise  quantitative
22      extrapolation can be used to accurately predict human chronic O3 response. Now that the
23      technology for accurate regional dose determinations in animals and humans is available,
24      more fully integrated studies examining dose-response are required.
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                              APPENDIX A
                 GLOSSARY OF TERMS AND SYMBOLS
99mTc-DTPA
AD
AHSMOG
AM
AMI
ANOCOVA
AOD
AR
ARE
ATP
BAL
BALF
BHNP
BF
bkg
BM
Brdu
BS
BSA
BW
CA
CAR
CC10
Cdyn
CE
CI
CI
CIU
CL
Radiolabeled diethylene triamine pentaacetic acid
Radiolabeled ferric oxide
Alveolar duct
Adventist Health Smog studies
Alveolar macrophage
Asthma medication index
Analysis of covariance
Obstructive airways disease
Autoregressive
California Air Resources Board
Adenosine triphosphate
Bronchoalveolar lavage
Bronchoalveolar lavage fluid
JV-bis (2-hydroxypropyl) nitrosamine
Black female
Background
Black male
Bromodeoxyuridine
Black smoke
Body surface area
Body weight
Clean air
Centriacinar region

Dynamic compliance
Constant exercise
Cardiac index
Confidence interval
Cumulative methacholine inhalation units
Confidence limit
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CO
CO2
CoH
ConA
COPD
CORD
CXT
Cu
DAAS
DR
DTPA
EEG
EPA
ER
ERV
ESC
f
F
F344
FA
FEF
FEF
    25-75%
FEV3
FEV0.75
FP
FRC
FVC
GDT
G6PD
GR
Carbon monoxide
Carbon dioxide
Coefficient of haze
Concanavalin A
Chronic obstructive pulmonary disease
Chronic obstructive respiratory disease
Concentration times duration of exposure
Copper
Distal airways and alveolar surface fraction
Diffusing capacity for CO
Disulfide reductase
Diethylenetriaminapentaacetate
Electroencephalogram
U.S. Environmental Protection Agency
Emergency room
Expiratory reserve volume
Epithelial secretory cell
Respiratory frequency
Female
Fischer 344
Filtered air
Breathing frequency (also fR)
Forced expiratory flow
Forced expiratory flow between 25 and 75 % of vital capacity
Ferric oxide
Iron sulfate
Forced expiratory volume in 1 s
Forced expiratory volume in 3 s
Forced expiratory volume in 0.75 s
Fine particle
Functional residual capacity
Forced vital  capacity
Glutathione-disulfide transhydrogenase
Glucose-6-phosphate
Glutathione reductase
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GSH
GSHPx
GST
H+
HC
HCHO
HMV
HNO3
H2O
HR
HRmax
H2SO4
1C
ICAM
ICD
IE
Ig
IL
ip
IP
IU
iv
LDH
LM
LT
M
MAP
MC
MLN
MMEF
Mn
MnSO4
MVV
N2
Glulathione
Glutathione peroxidase
Glutathione-S-transferase
Hydrogen ion
Hydrocarbon
Formaldehyde
Half-hour mean value
Nitric acid
Water
Heart rate
Maximum heart rate
Sulfuric acid
Inspiratory capacity
Intracellular adhesion molecule
Nicotinamide adenine diphsophate-specific isocitrate dehydrogenase
Intermittent exercise
Immunoglobulin (IgA, IgE, IgG, IgM )
Interleukin (IL-1, IL-6, IL-8)
Intraperitoneal
Inhalable particles
International Units
Intravenous
Lactate dehydrogenase
Light microscopy
Leukotriene (LTB4, LTC4, LTD4, LTE4)
Male
Mean arterial blood pressure
Methacholine challenge
Mediastinal lymph modes
Maximum mid-expiratory  flow
Manganese
Manganese sulfate
Maximum voluntary ventilation
Nitrogen
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AN2

NA
NAAQS
NADH
NADPH
NADPH-CR
NEP
NH4+
NHANES
(NH4)2S04
Nffl
NK
NL
NO
NO2
N03
NOX
NPSH
NS
NZW
02
03
OR
OX (10)
OX
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 PDI
 PE
 PEFR
 PG
 6PGD
 PHA
 PM2.5
PM
    10
PM15

PMN
p.r.n.
PUFA
R
Raw
RB
RH
RL
RR
RSP
RT
RV
S
SAROAD

SB
SD
S-D
SE
SEM
SEM
Provocative dose that produces a 100% decrease in forced expiratory
  volume in 1 s
Pain on deep inspiration
Postexposure
Peak expiratory flow rate
Prostaglandin (PGD2, PGE^ PGE2 PGFla, PGF2a)
6-Phosphogluconate
Phytochemagglutinin
Particulate matter of mass median aerodynamic diameter 2.5
  or less
Particulate matter of mass median aerodynamic diameter 10
  or less
Particulate matter of matter of mass median aerodynamic diameter
  15/um or less
Polymorphonuclear leukocyte
As needed (pro re nata)
Polyunsaturated fatty acid
Intraclass  correlation coefficeint
Airway resistance
Respiratory bronchiole
Relative humidity
Lung resistance
Relative risk
Respirable suspended particulates
Total respiratory resistance
Residual volume
Smoker
Storage and Retrieval of Aerometric Data, U.S.  EPA
  centralized data base; superseded by AIRS (q.v.)
Shortness  of breath
Standard deviation
Sprague-Dawley
Standard error
Standard error of the mean
Scanning electron microscopy
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SHAPE
SMG
S02
S04
S04=
SOD
SRBC
T
TB
TEARS
Tdb
THC
Tffl
TLC
TNF
TSP
TSP (200)

TX
UCLA
UV
VC
vs
VT
VTmax
V
  25%VC
VEmax
Specific airway conductance
Simulation of Human Activity and Pollutant Exposure
Small-mucous-granule
Sulfur dioxide
Sulfate
Sulfate ion
Superoxide dismutase
Specific airway resistance
Sheep red blood cell
Temperature
Terminal bronchiole
Thiobarbituric acid reactive substances
Dry bulb temperature
Total hydrocarbon content
Temperature-humidity index
Total lung capacity
Tumor necrosis factor
Total suspended particulates
Number of hours above a total suspended paniculate concentration of
 200 Atg/m3
Thromboxane (A2, B2)
University of California at Los Angeles
Ultraviolet
Vital capacity
Volume per surface area
Tidal volume (also TV)
Maximum tidal volume
Volume fraction
Lung volume at 25 % of the vital capacity
Lung volume at 50 % of the vital capacity
Alveolar ventilation
Minute ventilation; expired volume per minute
Maximum minute ventilation
Maximum expiratory flow at 25 %  of the vital capacity
Maximum expiratory flow at 50%  of the vital capacity
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VO2               Oxygen uptake by the body
VO2max            Maximal oxygen uptake (maximal aerobic capacity)
W                 Watt
WBGT             Wet bulb globe temperature
WF                White female
WM               White male
Zn                 Zinc
ZnO               Zinc oxide
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