1                    NAC/Interim (NO2)/Proposed (N2O4): December 2008
 2
 3
 4
 5
 6
 7
 8
 9
10   ACUTE EXPOSURE GUIDELINE LEVELS (AEGLs)

11

12                         FOR
13

14                NITROGEN DIOXIDE

15               (CAS Reg. No. 10102-44-0)

16

17               NITROGEN TETROXIDE

is               (CAS Reg. No. 10544-72-6)
19
20
21
22
23
24
25
26                INTERIM/PROPOSED
27
28
29
30

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1                                       PREFACE
 2
 3          Under the authority of the Federal Advisory Committee Act (FACA) P. L. 92-463
 4   of 1972, the National Advisory Committee for Acute Exposure Guideline Levels for
 5   Hazardous Substances (NAC/AEGL Committee) has been established to identify, review
 6   and interpret relevant toxicologic and other scientific data and develop AEGLs for high
 7   priority, acutely toxic chemicals.
 8
 9          AEGLs represent threshold exposure limits for the general public and are
10   applicable to emergency exposure periods ranging from 10 minutes to 8 hours. Three
11   levels— AEGL-1, AEGL-2 and AEGL-3— are developed for each of five exposure
12   periods (10 and 30 minutes, 1 hour, 4 hours, and 8 hours) and are distinguished by
13   varying degrees of severity of toxic effects. The three AEGLs are defined as follows:
14
15          AEGL-1 is the airborne concentration (expressed as parts per million or
16   milligrams per cubic meter [ppm or mg/m3]) of a substance above which it is predicted
17   that the general population, including susceptible individuals, could experience notable
18   discomfort, irritation, or certain asymptomatic, non-sensory effects. However, the effects
19   are not disabling and are transient  and reversible upon cessation of exposure.
20
21          AEGL-2 is the airborne concentration (expressed as ppm or mg/m3) of a
22   substance above which it is predicted that the general population, including susceptible
23   individuals, could experience irreversible or other serious, long-lasting adverse health
24   effects or an impaired ability to escape.
25
26          AEGL-3 is the airborne concentration (expressed as ppm or mg/m3) of a
27   substance above which it is predicted that the general population, including susceptible
28   individuals, could experience life-threatening health effects or death.
29
30          Airborne concentrations below the AEGL-1 represent exposure levels that can
31   produce mild and progressively increasing  but transient and nondisabling odor, taste, and
32   sensory irritation, or certain asymptomatic, non-sensory effects. With increasing airborne
33   concentrations above each AEGL, there is a progressive increase in the likelihood of
34   occurrence and the severity of effects described for each corresponding AEGL. Although
35   the AEGL values represent threshold levels for the general public, including susceptible
36   subpopulations, such as infants, children, the elderly, persons with asthma, and those with
37   other illnesses, it is recognized that individuals, subject to unique or idiosyncratic
38   responses, could experience the effects described at concentrations below the
39   corresponding AEGL.

-------
      NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1                                  TABLE OF CONTENTS
 2
 3    LIST OF TABLES	5
 4    LIST OF FIGURES	5
 5    SUMMARY	6
 6    1. INTRODUCTION	10
 7    2. HUMAN TOXICITY DATA	12
 8      2.1. Acute Lethality	12
 9      2.2. Nonlethal Toxicity	13
10        2.2.1. Case Reports	13
11        2.2.2. Epidemiologic Studies	15
12        2.2.3. Experimental Studies	17
13      2.3. Developmental/Reproductive Toxicity	21
14      2.4. Genotoxicity	22
15      2.5. Carcinogenicity	22
16      2.6. Summary	22
17    3. ANIMAL TOXICITY DATA	22
18      3.1. Acute Lethality	22
19        3.1.1. Dogs	23
20        3.1.2. Rabbits	23
21        3.1.3. Guinea Pigs	24
22        3.1.4. Rats	25
23        3.1.5. Mice	26
24      3.2. Nonlethal Toxicity	27
25        3.2.1. Monkeys	27
26        3.2.2. Dogs	27
27        3.2.3. Rabbits	28
28        3.2.4. Sheep	29
29        3.2.5. Guinea Pigs	29
30        3.2.6. Hamsters	30
31        3.2.7. Ferrets	30
32        3.2.8. Rats	31
33        3.2.9. Mice	34
34      3.3. Developmental/Reproductive Toxicity	35
35      3.4. Genotoxicity	35
36      3.5. Subchronic and Chronic Toxicity/Carcinogenicity	35
37      3.6. Summary	36
38    4. SPECIAL CONSIDERATIONS	36
39      4.1. Metabolism and Disposition	36
40      4.2. Mechanism of Toxicity	37
41      4.3. Oxides of Nitrogen	38
42      4.4. Other Relevant Information	39
43        4.4.1. Species Variability	39
44        4.4.2. Susceptible Populations	39
45        4.4.3. Concentration-Response Relationship	40
46        4.4.4. Susceptibility to infection	41
47    5. DATA ANALYSIS FOR AEGL-1	42
48      5.1. Summary of Human Data Relevant to AEGL-1	42
49      5.2. Summary of Animal Data Relevant to AEGL-1	43
50      5.3. Derivation of AEGL-1	43
51    6. DATA ANALYSIS FOR AEGL-2	43
52      6.1. Summary of Human Data Relevant to AEGL-2	44
53      6.2. Summary of Animal Data Relevant to AEGL-2	44
54      6.3. Derivation of AEGL-2	45
55    7. DATA ANALYSIS FOR AEGL-3	45

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1      7.1. Summary of Human Data Relevant to AEGL-3	45
 2      7.2. Summary of Animal Data Relevant to AEGL-3	46
 3      7.3. Derivation of AEGL-3	46
 4    8. SUMMARY OF AEGLS	47
 5      8.1. AEGL Values and Toxicity Endpoints	47
 6      8.2. Comparison with Other Standards and Criteria	48
 7    9. REFERENCES	51
 8    APPENDIX A: Derivation of AEGL Values	64
 9    APPENDIX B: Derivation Summary for AEGL Values for Nitrogen Oxides	68
10    APPENDIX C: Time Scaling Category Plot for Nitrogen Oxides	72
11
12

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1                                    LIST OF TABLES
 2
 3   SI:  Summary of AEGL Values for Nitrogen Dioxide	9
 4   S 2:  Summary of AEGL Values for Nitrogen Tetroxide	9
 5
 6   TABLE LPHYSICOCHEMICAL DATA FOR NITROGEN DIOXIDE	 11
 7   TABLE 2: PHYSICOCHEMICAL DATA FOR NITROGEN TETROXIDE	12
 8   TABLE 3: Effects of acute exposure to high NO2 concentrations	13
 9   TABLE 4: Summary of NO2 Mortality for Five Species	27
10   TABLES: AEGL-IValues for Nitrogen Dioxide and Nitrogen Tetroxide	43
11   TABLE 6: AEGL-2 Values for Nitrogen Dioxide and Nitrogen Tetroxide	45
12   TABLE 7: AEGL-3 Values for Nitrogen Dioxide and Nitrogen Tetroxide	47
13   TABLES: Summary of AEGL Values for Nitrogen Dioxide (mg/m3 [ppm])	47
14   TABLE 9: Summary of AEGL Values for Nitrogen Tetroxide (mg/m3 [ppm])	48
15   TABLE 10: Extant Standards and Guidelines for Nitrogen Dioxide	49
16
17
18                                   LIST OF FIGURES
19
20   Figure 1: Category plot of AEGL values and effects of nitrogen dioxide on humans and animals	73
21

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1                                         SUMMARY
 2
 3          Nitrogen oxide compounds occur from both natural and anthropogenic sources.
 4   Nitrogen dioxide (NO2) is the most ubiquitous of the oxides of nitrogen and has the
 5   greatest impact on human health. Nitrogen tetroxide (TSPzO^ is a component of rocket
 6   fuels. NC>2 exists as an equilibrium mixture of NC>2 and N2O4 but the dimer is not
 7   important at ambient concentrations (U.S. EPA 1993). The two compounds are phase-
 8   related forms with N2O4 favored in the liquid phase and NO2 favored in the gaseous
 9   phase. As a result when ^64 is released it vaporizes and dissociates into NO2, making
10   it nearly impossible to generate a significant concentration of N2©4 at atmospheric
11   pressure and ambient temperatures, without generating a vastly higher concentration of
12   NO2. Very few inhalation toxicity data are available on N2O4. Thus, the AEGL values
13   were developed based on data for NC>2, the predominant form, and values are considered
14   applicable to all nitrogen oxides.  Values for N2O4 in units of ppm have been calculated
15   on a molar basis as presented below.
16
17          NC>2 is an irritant to the mucous membranes and may cause coughing and dyspnea
18   during exposure. After less severe exposure, symptoms may persist for several hours
19   before subsiding (NIOSH 1976).  With more severe exposure, pulmonary edema ensues
20   with signs of chest pain, cough, dyspnea, cyanosis, and moist rales heard on auscultation
21   (NIOSH 1976, Douglas et al. 1989).  Death from NO2 inhalation is caused by
22   bronchospasm and pulmonary edema in association with hypoxemia and respiratory
23   acidosis, metabolic acidosis, shift of the oxyhemoglobin dissociation curve to the left,
24   and arterial hypotension (Douglas et  al. 1989). A characteristic of NO2 intoxication after
25   the acute phase is a period of apparent recovery followed by late-onset bronchiolar injury
26   that manifests as bronchiolitis fibrosa obliterans (NIOSH 1976, NRC 1977, Hamilton
27   1983, Douglas et al. 1989).  In addition, experiments with laboratory animals indicate
28   that exposure to NO2 increases susceptibility to infection (Henry et al. 1969, U.S. EPA
29   1993) due, in part, to alterations in host pulmonary defense mechanisms (Gardner et al.
30   1969).
31
32          For AEGL-1 a concentration  of 0.5 ppm was adopted for all time points.
33   Although the response of asthmatics  to NO2 is variable,  asthmatics were identified as a
34   potentially susceptible population. The evidence indicates that some asthmatics exposed
35   to 0.3-0.5 ppm NO2 may respond with either subjective  symptoms or slight changes in
36   pulmonary function of no clinical significance. In contrast, some asthmatics did not
37   respond to NO2 at concentrations of 0.5-4 ppm.  Because of the weight of evidence, the
38   study by Kerr et al. (1978, 1979) was considered the most appropriate for derivation of
39   AEGL-1 values. They reported that 7/13 asthmatics experienced slight burning of the
40   eyes, slight headache, and chest tightness or labored breathing with exercise during
41   exposure to  0.5 ppm for 2 hours; at this concentration the odor of NO2 was perceptible
42   but the subjects became unaware of it after about 15 minutes.  No changes in any
43   pulmonary function tests were found immediately  following the chamber exposure (Kerr
44   et al. 1978, 1979).  Therefore, 0.50 ppm was considered a no-adverse-effect level for the
45   asthmatic population.  Since asthmatics are potentially the most susceptible population,
46   no uncertainty factor was applied.  Time scaling was not done because adaptation to mild

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   sensory irritation occurs. In addition, animal responses to NC>2 exposure have
 2   demonstrated a much greater dependence upon concentration than upon time; therefore,
 3   extending the 2-hour concentration to 8 hours should not exacerbate the human response.
 4
 5          Supporting studies for AEGL-1 report findings similar to the key studies.
 6   Significant group mean reductions in FEVi (-17.3% with NC>2 vs -10.0% with air) and
 7   specific airway conductance (-13.5% with NC>2 vs -8.5% with air) occurred in asthmatics
 8   after exercise during exposure to 0.3 ppm for 4 hours and 1/6 individuals experienced
 9   chest tightness and wheezing (Bauer et al. 1985). The onset of effects was delayed when
10   exposures were by oral-nasal inhalation as compared to oral inhalation and may have
11   resulted from scrubbing within the upper airway. In a similar study, asthmatics exposed
12   to 0.3 ppm for 30 minutes at rest followed by  10 minutes of exercise had significantly
13   greater reductions in FEVi (10% vs 4% with air) and partial expiratory flow rates at 60%
14   of total lung capacity, but no symptoms were reported (Bauer et al. 1986). In a
15   preliminary study with 13 asthmatics exposed to 0.3 ppm for 110 minutes, slight cough
16   and dry mouth and throat and significantly greater reduction in FEVi occurred after
17   exercise (11% vs 7%); however, in a larger study, no changes in pulmonary function
18   were measured and no symptoms were reported following exposure of 21 asthmatics to
19   concentrations up to 0.6 ppm for 75 minutes (Roger et al. 1990).
20
21          Human data were also used as the basis for AEGL-2. Three healthy male
22   volunteers experienced definite discomfort from exposure to 30 ppm for 2 hours
23   (Henschler et al. 1960).  Three individuals exposed to 30 ppm for 2 hours perceived an
24   intense odor upon entering the chamber, but the odor perception quickly diminished and
25   was completely absent after 25-40 minutes. One individual experienced a slight tickling
26   of the nose and throat mucous membranes after 30 minutes, the two others after 40
27   minutes. From 70 minutes on, all subjects experienced a burning sensation and an
28   increasingly severe cough for the next 10-20 minutes, but coughing decreased from 100
29   minutes on. However, the burning sensation continued and moved into the lower
30   sections of the airways and was finally felt deep in the chest. At this time, marked
31   sputum  secretion and dyspnea were noted. Toward the end of the  exposure,  the subjects
32   reported the exposure conditions to be bothersome and barely tolerable. A sensation of
33   pressure and increased sputum secretion continued for several hours after cessation of
34   exposure (Henschler et al. 1960). The point of departure is considered a threshold for
35   AEGL-2 since the effects noted by the subjects would not impair the ability to escape and
36   the effects were reversible after cessation of exposure.
37
38          AEGL-3 values were based on animal  data and supported by a human case report.
39   Exposure of monkeys to 50 ppm for 2 hours was used to derive the AEGL-3 values.
40   Monkeys (n = 2-6/group) were exposed to 10-50 ppm NO2 for 2 hours with respiratory
41   function monitored during exposure (Henry et al. 1969).  NO2 exposure alone resulted in
42   a markedly increased respiratory rate and decreased tidal volume during exposures to 50
43   and 35 ppm, but only slight effects at 15 and 10 ppm.  Mild histopathological changes in
44   the lungs were noted after exposure to 10 and  15 ppm, however, marked changes in lung
45   structure were observed after exposure to 35 and 50 ppm. The alveoli were expanded
46   with septal wall thinning, bronchi were inflamed with proliferation or erosion of the

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   surface epithelium, and lymphocyte infiltration was seen with edema.  In addition to the
 2   effects on the lungs, interstitial fibrosis (35 ppm) and edema (50 ppm) of cardiac tissue,
 3   glomerular tuft swelling in the kidney (35  and 50 ppm), lymphocyte infiltration in the
 4   kidney and liver (50 ppm), and congestion and centrilobular necrosis in the liver (50
 5   ppm) were observed.
 6
 7          The AEGL-3 values are supported  by human data for a welder.  Pulmonary
 8   edema, confirmed on X-ray and requiring  medical intervention, resulted from exposure to
 9   approximately 90 ppm for up to 40 minutes (Norwood et al. 1966). If this exposure
10   scenario is  used for derivation of AEGL-3  values with an uncertainty factor of 3 the
11   values are nearly identical to those derived using the data in the monkey. In addition, the
12   AEGL-3  values are below the concentrations at which lethality first occurred in five
13   animal species: 75 ppm for 4 hours in the dog and 1 hour in the rabbit, 50 ppm for 1 hour
14   in the guinea pig, and 50 ppm for 24 hours in the rat and mouse (Hine et al.  1970).
15
16          For AEGL-2 and AEGL-3, the 10- and 30-minute, and 1-, 4-, and 8-hour AEGL
17   endpoints were calculated from Cn x t = k  using n = 3.5 (ten Berge et al. 1986).  The
18   value of n was calculated by ten Berge et al. using the data of Hine et al. (1970) in five
19   species of laboratory animal. A total uncertainty factor of 3 was applied which includes a
20   3 for intraspecies variability and a 1 for interspecies variability. Use of a greater
21   intraspecies uncertainty factor was not considered necessary because the mechanism of
22   action is not expected to differ greatly among individuals. Because human data were
23   used as the point of departure for AEGL-2, the endpoint in the monkey study is below the
24   definition of AEGL-3, human data support the AEGL-3 point of departure and derived
25   values, the  mechanism of action does not vary between  species with the target at the
26   alveoli, and due to the similarities of the respiratory tract between humans and monkeys,
27   additional interspecies uncertainty factors  are not considered necessary.
28
29          The values for the three AEGL classifications for the five time periods are listed
30   in Table S  1 for NO2 and Table S 2 for N2O4.

-------
      NITROGEN OXIDES
                                         NAC/Interim (NO2)/Proposed (N2O4): 12/2008
S 1: Summary of AEGL Values for Nitrogen Dioxide
Classification
AEGL-lb
(Non-
disabling)
AEGL-2
(Disabling)
AEGL-3
(Lethal)
10-Minute
0.94
mg/m3
(0.50 ppm)
38 mg/m3
(20 ppm)
64 mg/m3
(34 ppm)
30-Minute
0.94
mg/m3
(0.50 ppm)
28 mg/m3
(15 ppm)
47 mg/m3
(25 ppm)
1-Hour
0.94
mg/m3
(0.50 ppm)
23 mg/m3
(12 ppm)
38 mg/m3
(20 ppm)
4-Hour
0.94
mg/m3
(0.50 ppm)
15 mg/m3
(8.2 ppm)
26 mg/m3
(14 ppm)
8-Hour
0.94
mg/m3
(0.50 ppm)
13 mg/m3
(6.7 ppm)
21 mg/m3
(11 ppm)
Endpoint" (Reference)
slight burning of the eyes,
slight headache, chest
tightness or labored
breathing with exercise in
7/13 asthmatics (Kerr et
al. 1978, 1979)
burning sensation in nose
and chest, cough,
dyspnea, sputum
production in normal
volunteers (Henschler et
al. 1960)
marked irritation;
histopath in lungs; fibrosis
and edema of cardiac
tissue; necrosis in liver;
no deaths in monkeys
(Henry etal. 1969)
      aSome effects may be delayed.
      bThe sweet odor of NO2 may be perceptible to most individuals at this concentration; however, adaptation
      occurs rapidly.
S 2: Summary of AEGL Values for Nitrogen Tetroxide
Classification
AEGL-lb
(Non-
disabling)
AEGL-2
(Disabling)
AEGL-3
(Lethal)
10-Minute
0.94
mg/m3
(0.25 ppm)
38 mg/m3
(10 ppm)
64 mg/m3
(17 ppm)
30-Minute
0.94
mg/m3
(0.25 ppm)
28 mg/m3
(7.6 ppm)
47 mg/m3
(13 ppm)
1-Hour
0.94
mg/m3
(0.25 ppm)
23 mg/m3
(6.2 ppm)
38 mg/m3
(10 ppm)
4-Hour
0.94
mg/m3
(0.25 ppm)
15 mg/m3
(4.1 ppm)
26 mg/m3
(7.0 ppm)
8-Hour
0.94
mg/m3
(0.25 ppm)
13 mg/m3
(3.5 ppm)
21 mg/m3
(5.7 ppm)
Endpoint" (Reference)
slight burning of the eyes,
slight headache, chest
tightness or labored
breathing with exercise in
7/13 asthmatics (Kerr et
al. 1978, 1979)
burning sensation in nose
and chest, cough,
dyspnea, sputum
production in normal
volunteers (Henschler et
al. 1960)
marked irritation;
histopath in lungs; fibrosis
and edema of cardiac
tissue; necrosis in liver;
no deaths in monkeys
(Henry etal. 1969)
 9
10
11
aSome effects may be delayed.
bThe sweet odor of NO2 may be perceptible to most individuals at this concentration; however, adaptation
occurs rapidly.

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   1. INTRODUCTION
 2
 3          Nitrogen dioxide (NO2) is the most ubiquitous of the oxides of nitrogen and has
 4   the greatest impact on human health. NO2, which exists as an equilibrium mixture of
 5   NO2 and ^64 (nitrogen tetroxide), is a reddish-brown gas with a sweet odor, is heavier
 6   than air, and reacts with water (U.S. EPA 1993, Mohsenin 1994).  NC>2 is shipped under
 7   pressure and the equilibrium between NO2 and N2O4 is altered with change in pressure
 8   with N2O4 becoming predominant at very high pressures. NO2 is a free radical with
 9   sufficient stability to exist in relatively high concentrations in ambient air (Mohsenin
10   1994).
11
12          The major source of atmospheric NC>2 is from the combustion of fossil fuels for
13   heating, household appliances, power generation, and in motor vehicles. Consequently,
14   the chemical is a major contributor to smog and a concern for indoor air quality.
15   Ambient levels in urban air pollution episodes in the US have been measured between 0.1
16   and 0.8 ppm as a maximum hourly average with short-term peaks  as high as 1.27 ppm.
17   Indoor NO2 concentrations may reach a maximum 1 hour level of 0.25 to 1.0 ppm with
18   peak levels as high as 2-4 ppm where gas appliances or kerosene heaters are used
19   (Mohsenin 1994).
20
21          N2O4 is a commonly used as a rocket propellant (Yue et al. 2004). Limited
22   toxicity data on N2O4 show effects similar to those of NC>2.
23
24          No toxicity data or information on the uses or sources of nitrogen trioxide (N2O3)
25   were found.  Information on the chemical interactions of N2Os with the other oxides of
26   nitrogen was not available.  Therefore, N2Os will not be considered further here.
27
28          NC>2 is an irritant to the mucous membranes and may cause coughing and dyspnea
29   during exposure. After less severe exposure, symptoms may persist for several hours
30   before subsiding (NIOSH 1976).  With more severe exposure,  pulmonary edema ensues
31   with chest pain, cough, dyspnea, cyanosis, and moist rales heard on auscultation  (NIOSH
32   1976, Douglas et al. 1989).  Death from NO2 inhalation is caused by bronchospasm and
33   pulmonary edema in association with hypoxemia and respiratory acidosis, metabolic
34   acidosis, shift of the oxyhemoglobin dissociation curve to the left, and arterial
35   hypotension (Douglas et al. 1989). A characteristic of NO2 intoxication after the acute
36   phase is a period of apparent recovery followed by late-onset bronchiolar injury that
37   manifests as bronchiolitis fibrosa  obliterans (NIOSH 1976, NRC 1977, Hamilton 1983,
38   Douglas etal. 1989).
39
40          Selected physicochemical properties of nitrogen dioxide and of nitrogen tetroxide
41   are listed in Tables 1 and 2, respectively.
42
43
                                             10

-------
NITROGEN OXIDES
NAC/Interim (NO2)/Proposed (N2O4): 12/2008
TABLE 1: PHYSIC OCHEMICAL DATA FOR NITROGEN DIOXIDE
Parameter
Common name
Synonyms
CAS registry no.
Chemical formula
Molecular weight
Physical state
Vapor pressure
Vapor density (air =1)
Melting/boiling point
Flamability
Solubility in water
Conversion factors in air
Reactivity
Value
nitrogen dioxide

10102-44-0
NO2
46.01
reddish-brown gas
720 torr at 20°C; 800 mm Hg at
25°C
1.58
-9.3°C/21.15°C
does not burn
0.037 niL/mL at 35°C
1 ppm = 1.88 mg/m3
1 mg/m3 = 0.53 ppm
decomposes in water forming nitric
oxide and nitric acid
Reference



Budavari et al. 1996
Budavari et al. 1996
Budavari et al. 1996
ACGIH 1991, U.S. EPA 1990
Budavari et al. 1996
Budavari et al. 1996
Budavari et al. 1996
Mohsenin 1994
U.S. EPA 1993
Budavari et al. 1996
                                    11

-------
     NITROGEN OXIDES
                                   NAC/Interim (NO2)/Proposed (N2O4): 12/2008
TABLE 2: PHYSICOCHEMICAL DATA FOR NITROGEN TETROXIDE
Parameter
Common name
Synonyms
CAS registry no.
Chemical formula
Molecular weight
Physical state
Vapor pressure
Vapor density (air = 1)
Melting/boiling point
Flamability
Solubility in water
Conversion factors in air
Reactivity
Value
dinitrogen dioxide

10544-72-6
N2O4
92.01
colored liquid
760mmHgat21°C
1.45at20°C
-9.3°C/21.5°C
No data
No data
1 ppm= 3.70 mg/m3
1 mg/m3 = 0.27 ppm
Reacts violently with organic
compounds; reacts with water
Reference



Lide 1988
Lide 1988
Lide 1988
Lide 1988
Lide 1988
Kushneva and Gorshkova 1999,
Lide 1988


calculated
Kushneva and Gorshkova 1999,
Lide 1988
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
14
15
16
17
2. HUMAN TOXICITY DATA
2.1. Acute Lethality

       Book (1982) used allometric scaling based on minute volume and LCso values for
NC>2 for 5 animal species to calculate a human 1-hour LCso of 174 ppm. Concentrations
of >200 ppm are reported to induce immediate symptoms of bronchospasm and
pulmonary edema and may cause syncope, unconsciousness, and quick death (Douglas et
al. 1989).

       Clinical responses to "acute" inhalation of high concentrations of NC>2 based on
occupational exposures, are given in Table 3 (NRC 1977).  Durations of exposures were
not specified except for the  statement that workers in a nitric acid manufacturing plant in
Italy were exposed to average concentrations of 30-35 ppm for an unspecified number of
years with no adverse signs or symptoms.
                                           12

-------
     NITROGEN OXIDES
                                     NAC/Interim (NO2)/Proposed (N2O4): 12/2008
TABLE 3: Effects of acute exposure to high NO2 concentrations
Concentration (ppm)
0.4
15-25
25-75
150-300+
Effect
approximate odor threshold
respiratory and nasal irritation
reversible pneumonia and bronchiolitis
fatal bronchiolitis and bronchopneumonia
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
From NRC 1977.
2.2. Nonlethal Toxicity
2.2.1. Case Reports

       Probably the most well known occupational manifestation of NO2 toxicity is that
of silo filler's disease. In a silo, the gas that accumulates above the silage is depleted of
oxygen, is rich in carbon dioxide, and contains a mixture of nitrogen oxides, mainly
NC>2 which can reach concentrations of 200 to 4000 ppm within 2 days (Lowry and
Schuman 1956, Douglas et al. 1989).  The term silo filler's disease was first used by
Lowry and Schuman in 1956 in an article in which they described the clinical
progression of the disease: inhalation of irritant gas from a silo; immediate cough and
dyspnea with a sensation of choking; 2-3 week period after exposure of apparent
remission; second phase of illness accompanied by fever with progressively more severe
dyspnea, cyanosis, and cough; inspiratory and expiratory rales; discrete nodular
densities on the lung; and neutrophilic leukocytosis (Lowry and Schuman 1956).
Douglas et al. (1989) reported on 17 patients examined at the Mayo  clinic between 1955
and 1987 after exposure to silo gas. Eye irritation was described during exposure, acute
lung injury occurred in 11 individuals, and 16 had either persistent or delayed symptoms
of dyspnea, cough, chest pain, and rapid breathing.  One patient died and at autopsy
diffuse alveolar damage with hyaline membranes and hemorrhagic pulmonary edema
and acute edema of the airways were observed. Bronchiolitis fibrosa obliterans
developed in one patient many years later; however, prophylactic administration of
corticosteroids may have prevented chronic obstructive pulmonary disease (COPD) in
the other patients. Similar case reports and  outcomes of silo filler's  disease and
industrial exposure were described in earlier literature (Grayson 1956, Lowry and
Schuman 1956, Milne 1969).

       A welder developed shortness of breath and chest discomfort during the use of an
acetylene torch for metal-cutting in a poorly ventilated water main; the worker had spent
approximately 30 minutes welding in the confined space before being forced to vacate.
Several hours later, the worker became so short of breath that he couldn't sleep that night.
Chest X-ray 18 hours after exposure revealed pulmonary edema and a pulmonary
function test showed 42% of the predicted value for FVC. The individual was admitted
to the hospital and treated with antibiotics and oxygen. The patient fully recovered by 21
days after exposure. Simulation of the accident produced an NO2 concentration of 90
ppm within 40 minutes and total oxides of nitrogen in excess of 300 ppm (Norwood et al.
                                             13

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   1966). It can be assumed that the individual was exposed to at least 90 ppm during the
 2   welding operation and that the outcome could have been more severe, or even fatal,
 3   without medical intervention.
 4
 5          An outbreak of NO2-induced respiratory illness was reported among players and
 6   spectators at two high school hockey games (Hedberg et al. 1989).  Patients presented
 7   with acute onset of cough, hemoptysis, and/or dyspnea during or within 48 hours of
 8   attending the hockey game. No changes in lung function were measured 10 days and 2
 9   months after exposure. NO2 concentrations were not measured in the arena during the
10   outbreak, but the source was traced to a malfunctioning motor on the ice resurfacer.
11   Other cases of respiratory illness in hockey players, referees, and spectators have been
12   associated with elevated nitrogen dioxide levels in the arena due to malfunctioning
13   resurfacers or ventilation systems combined with elevated carbon monoxide levels (Smith
14   et al. 1992, Soparkar et al. 1993, Morgan 1995, Karl son-Stiber et al. 1996). Attempts to
15   measure NO2 concentrations in the arenas or reconstruct the situations were described by
16   the authors as not indicative of the actual exposure scenario which resulted in adverse
17   effects.
18
19          Morley and Silk (1970) described a number of cases in which welders involved in
20   ship repair and shipbuilding were exposed to nitrous fumes.  Symptoms included
21   dyspnea, cough, headache, tightness/pain in chest, nausea, and cyanosis. Most patients
22   recovered after treatment with oxygen and antibiotics; however, one man died 43 days
23   later from viral pneumonia. Two individuals admitted to the hospital with cyanosis,
24   dyspnea, and pulmonary edema, were exposed to a concentration of NO2 measured at 30
25   ppm during a 40-minute welding operation. However, the authors noted that 7 other
26   individuals present at the time were unaffected.
27
28          A railroad tank car ruptured at a chemical plant releasing a cloud of NO2 in a
29   small community (Bauer et al. 1998).  In the first 30 hours after the release, the most
30   common symptoms reported in emergency room visits were headache, burning eyes, and
31   sore throat. Most air samples collected 3-7 hours after the release showed concentrations
32   of 0 ppm with one sample showing 1.4 ppm. No attempt was made to correlate
33   symptoms with estimated exposure.
34
35          Acute toxic reactions were described in four fireman who were exposed to NO2
36   which originated from a leak in a chemical plant (Tse and Bockman 1970).
37   Concentrations were not reported and exposure durations were defined as "barely a few
38   minutes" to "about ten minutes."  Initial responses, which cleared within several days,
39   included headache, a dry hacking cough, pulmonary edema, sinusitis, and/or upper
40   respiratory tract irritation.  Four to six weeks after exposure, three of the patients
41   developed fever, chest tightness, shortness of breath, and a productive cough; these
42   subsided and the patients remained asymptomatic. The  fourth patient developed
43   chronic pulmonary insufficiency, consisting of dyspnea on exertion, despite normal
44   chest X-ray.
45
46          A large number of patients were treated for respiratory complaints following
                                             14

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   release of a cloud of ^64 from a railroad tank car.  The most common symptoms were
 2   headache, burning eyes, and sore throat; an abnormal lung exam and an abnormal chest
 3   x-ray were also reported for some individuals, but these findings were not further defined
 4   (Bauer et al. 1996).  No pulmonary edema or deaths were attributed to the accident.
 5   However, six individuals were later diagnosed with reactive airways dysfunction
 6   syndrome (RADS) three months after exposure (Conrad et al. 1998). Concentrations of
 7   various oxides of nitrogen in the cloud were not reported.
 8
 9          Four cases of exposure to unknown concentrations of nitrous fumes were reported
10   for individuals involved in either the use of an oxyacetylene burner during a leak at a
11   chemical plant, or in shotfiring (Jones et al. 1973).  Three patients presented with
12   pulmonary edema, one of which progressed to bronchiolitis obliterans; the fourth patient
13   presented with clinical features of bronchi olitis  obliterans. All recovered completely
14   following corticosteroid treatment.
15
16   2.2.2. Epidemiologic Studies
17
18          Several  epidemiological studies associating ambient NC>2 exposure with an
19   increase in the prevalence of respiratory illness  have been inconclusive. Increased odds
20   ratios (1.2-1.7) were found for bronchitis, chronic cough, and chest illness but not for
21   wheeze and asthma in children from six U.S. cities with annual average NC>2 levels
22   0.0065-0.0226 ppm (Dockery et al. 1989).  No association was found between long-term
23   differences in NC>2 levels (change of 0.0106 ppm/6-week average) and mean annual rates
24   of respiratory episodes in children from urban and rural regions in Switzerland,  however
25   the duration of symptoms was increased (Braun-Fahrlaender et al. 1992).  An increase in
26   the cases of croup in children was associated with total  suspended particulate matter and
27   NC>2 (Schwartz et al. 1991)  and decreased lung function in children was linked to sulfur
28   dioxide in combination with NO2 (Mostardi et al. 1981). Symptoms of chronic
29   obstructive pulmonary disease have been linked to exposure to total oxidants (>0.1 ppm),
30   NO2, and/or sulfates, but not to NO2 alone (Detels et al. 1981, Euler et al.  1988).
31   Combined effects of NC>2, SC>2, particulate matter, H^S, and other pollutants were
32   considered as contributing factors to a positive association between the occurrence of
33   upper respiratory infections and living in polluted areas of Finland for children <2 years
34   and 6 years old (Jaakkola et al. 1991).
35
36          In a more recent study, children from 12 communities in California were assessed
37   for respiratory disease prevalence and pulmonary function (Peters et al. 1999a,b).
38   Wheeze prevalence was correlated with levels of both acid and NC>2 in boys, whereas
39   regression analysis showed  that NC>2 was significantly associated with lower FVC, FEVi,
40   and maximal midexpiratory flow in girls. When these data were further analyzed by
41   month (Millstein et  al. 2004), wheezing during the spring and summer months was not
42   associated with either nitric acid or NC>2. However among asthmatics, the monthly
43   prevalence of asthma medication use was associated with monthly levels of ozone, nitric
44   acid, and acetic acid (Millstein et al. 2004).  Similar results were reported for eight areas
45   of Switzerland in which a 10 |ig/m3 average increase in NC>2  exposure was associated
46   with decreases in FVC (Schindler et al. 1998).
                                             15

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2          Several recent studies have attempted to describe the correlation between NO2
 3   levels and mortality or respiratory symptoms by pooling large datasets from multiple
 4   cities or countries. One of these studies used information collected from up to 12 cities in
 5   Canada.  These authors found that an approximate 20 ppb increase in NO2 was positively
 6   associated with a 2.25% increase in mortality (Burnett et al. 2004), intrauterine growth
 7   retardation (odds ratio 1.14-1.16) (Liu et al. 2007), a 17.72% increase in SIDS incidence
 8   (Dales et al. 2004), increased numbers of hospitalizations due to cardiac disease (Cakmak
 9   et al. 2006), and greater asthma hospitalizations in children 6-12 years of age (Lin et al.
10   2003). However, many of the positive findings in Canada were also positively correlated
11   with other pollutants such as particulate matter, ozone, and sulfur dioxide.  Similarly, a
12   significant association of NO2 with cardiovascular and respiratory mortality was found in
13   30 European cities (Samoli et al. 2006) and in nine French cities (Le Tertre et al. 2002),
14   but evidence of confounding effects of black smoke, sulfur dioxide, and/or ozone were
15   also found in both studies.
16
17          Asthma and allergy prevalence in conjunction with NO2 levels have also been
18   assessed in multiple city or country studies. Positive correlations were found for asthma
19   attacks, tightness in the chest, wheeze, and/or allergic rhinitis in children from eight
20   Japanese communities (Shima et al. 2002) and in  13 areas of Italy with the most
21   pronounced effects in the warmer Mediterranean areas (de Marco et al. 2002). An
22   increased incidence of morning symptoms was associated with a 6-day average increase
23   in NO2 (odds ratio 1.48) in asthmatic children from eight US cities (Mortimer et al.
24   2002). In a cross-sectional study of five countries, long-term NO2 concentrations were
25   correlated with sensitivity to inhaled allergens, but not to prevalence of bronchitis or
26   asthma (Pattenden, et al. 2006). No association was found between NO2 concentrations
27   and asthma, allergic rhinitis, and atopic dermatitis in children from six French cities
28   (Penard-Morand et al. 2005).
29
30          Epidemiological studies associating indoor NO2 have also been inconclusive.
31   One study found no evidence of any short-term association between prevalence of
32   respiratory symptoms in infants and median indoor and outdoor NO2 levels of 6.8 and
33   12.6 ppb, respectively (Farrow et al. 1997).  Similarly, no associations were found with
34   indoor NO2 and wheeze and asthma in children from seven Japanese communities (Shima
35   and Adachi 2000). Other studies found a significant increase in the occurrence of sore
36   throat, colds, and absences from school among children exposed to hourly peak levels of
37   >80 ppb NO2 from unvented gas heating in the classrooms (Pilotto et al.  1997), increased
38   respiratory illness in children from homes using gas cooking where NO2 concentrations
39   in the children's bedroom ranged from 4-169 ppb (Florey et al. 1979),  and slight
40   decreases in forced vital capacity and peak expiratory flow among adult asthmatics
41   exposed to >0.3 ppm while cooking on a gas range (Goldstein et al. 1988). Similarly,
42   Neas et al.  (1991) found that a 15 ppb increase in household annual NO2 mean was
43   associated with an increased cumulative incidence of attacks of shortness of breath with
44   wheeze, chronic wheeze, chronic cough, chronic phlegm, or bronchitis in children.
45
46          As part of a review of the National Ambient Air Quality Standards (NAAQS) for
                                             16

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   NC>2, U.S. EPA (1995) conducted a meta-analysis of studies which examined the
 2   respiratory effects on children living in homes with gas stoves.  Conclusions drawn from
 3   that analysis were that children ages 5-12 years old had an increased risk of about 20%
 4   for developing respiratory symptoms and disease with each increase of 0.015 ppm in
 5   estimated 2-week average NC>2 exposure (mean weekly concentrations in bedrooms
 6   0.008-0.065 ppm) and that no evidence for increased risk was found for infants <2 years
 7   old. Several limitations of this meta-analysis have been noted, including the following:
 8   uncertainty between monitored vs. actual exposure concentration; peak and average
 9   exposures could not be distinguished by the method used; and confounding effects of
10   other gas combustion by-products.  In context of the NAAQS review it was noted that
11   indoor exposures do not mimic outdoor exposures (U.S. EPA 1995).
12
13          Several occupations result in exposure to NC>2 concentrations higher than ambient
14   levels. In diesel bus garage workers, NC>2 concentrations of >0.3 ppm, along with
15   respirable particulates, were associated with work-related symptoms of cough, itching,
16   burning or watering eyes, difficult breathing, chest tightness, and wheeze; but, there were
17   no reductions in pulmonary function (Gamble et al. 1987). In contrast, no relationship
18   was found between respiratory symptoms or decline in FEVi among British coalminers
19   and exposure to peak NC>2 concentrations up to 14 ppm; controls were matched for age,
20   dust exposure, smoking habit, coal rank, and type of work (Robertson et al. 1984).  No
21   differences in pulmonary  function were noted among shipyard welders exposed to
22   average concentrations of 0.04 ppm oxides of nitrogen (Peters et al. 1973).  Slight
23   increases in prevalence of bronchitis (17.2% vs  12.6%) and colds (37.5% vs 30.7%) were
24   noted in traffic officers exposed to automobile exhaust containing mean concentrations of
25   0.045-0.06 ppm NO2 (Speizer and Ferris 1973).
26
27          In conclusion, indoor air quality may be more significant than outdoor air quality
28   to the prevalence of respiratory illness due to NC>2. A review of epidemiology studies
29   which assessed ambient quality (U.S. EPA 1993) yielded insufficient evidence to reach
30   any conclusion about the  long- or short-term health effects of NC>2.  Further, review of
31   epidemiology studies that assessed indoor air quality in homes with gas stoves, found
32   that meta analysis yielded insufficient evidence that NC>2 had an effect on infants 2 years
33   and younger while several considerations limited the interpretation of the positive
34   results for children aged 5-12 years.
35
36   2.2.3. Experimental Studies
37
38   Healthy Subjects
39
40          The odor threshold for NC>2 in air has been reported as 0.4 ppm for recognition
41   and 4.0 ppm for less than 100% identification (NIOSH 1976). In an experimental
42   study, the odor of NC>2 was  perceived by 3/9 volunteers exposed to 0.12 ppm and by
43   8/13 subjects at 0.22 ppm. At concentrations of <4 ppm, the volunteers perceived the
44   odor for 1-10 minutes, but the duration of perception was not directly related to
45   concentration. The olfactory response to NC>2 returned 1-1.5 minutes after cessation of
46   exposure (Henschler et al. 1960).  There appears to be a difference between perception
                                             17

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   and recognition level concentrations and the volunteers perceiving the odor at the
 2   lowest concentrations were described as "olfactorily sensitive."
 3
 4          Exposures of healthy individuals to <2 ppm NC>2 have shown no effects on
 5   pulmonary function or symptoms.  In several studies, healthy men and/or women were
 6   exposed to 0.6 ppm NC>2 for 1-3 hours with intermittent or continuous exercise.  No
 7   significant effects were observed in any study on pulmonary function, cardiovascular
 8   function, metabolism, or symptoms of exposure (Folinsbee et al. 1978, Adams et al.
 9   1987, Frampton et al. 1991, Hazucha et al. 1994). No changes in pulmonary function
10   occurred following exposure to 1.5 ppm for 3 hours or to a baseline of 0.05 ppm with
11   intermittent peaks of 2 ppm, however, continuous exposure to 1.5 ppm for 3 hours
12   resulted in a slight but significantly greater fall in FEVi and FVC in response to
13   carbachol (Frampton et al. 1991).  Pulmonary function was not affected in competitive
14   athletes exposed to 0.18 and 0.30 ppm for 30 minutes during heavy exercise (Kim et al.
15   1991) or in healthy adults exposed to 0.3 ppm for 4 hours with intermittent exercise
16   (Smeglin et al. 1985).
17
18          Studies at higher concentrations of NC>2 indicate an apparent threshold before
19   pulmonary function is affected.  No changes in pulmonary function, airway reactivity, or
20   indications of irritation were measured in healthy adults exposed to 1 ppm for 2 hours, 2
21   ppm for 3 hours (Hackney et al. 1978), 2 ppm for 4 hours (Devlin et al. 1992), 3 ppm for
22   2 hours (Goings et al. 1989) or to 2.3 ppm for 5 hours (Rasmussen et al. 1992).  Normal
23   subjects exposed to 2 ppm for 1  hour developed an increase in airway  reactivity to
24   methacholine challenge without changes in lung volume or pulmonary function
25   (Mohsenin 1988). No statistically  significant effects on airway resistance, symptoms,
26   heart rate, skin conductance, or self-reported emotional state were found in healthy
27   volunteers exposed to 4 ppm NO2 for 1 hour and 15 minutes with intermittent light and
28   heavy exercise (Linn and Hackney 1983).  However, a significant decrease in mean (n =
29   11) alveolar O2 partial pressure by  8 mm Hg and a significant increase in mean (n = 11)
30   airway resistance from 1.51 to 2.41 cm H2O/(L/s) occurred in healthy volunteers exposed
31   to 5 ppm for 2 hours with 6/11 individuals responding (von Nieding et al. 1979).
32   Similarly, a 10-minute exposure to 4-5 ppm resulted in increased expiratory and
33   inspiratory flow resistance in five healthy males; the effect was greatest 30 minutes after
34   exposure (Abe 1967).
35
36          Henschler et al. (1960) performed several experiments on healthy, male
37   volunteers. They reported that a 2-hour exposure to 20 ppm did not cause any irritation
38   when preceded by several exposures to lower concentrations during the preceding days;
39   however, exposure to 30 ppm for 2 hours caused definite discomfort.  Three individuals
40   exposed to 30 ppm for 2 hours perceived an intense odor upon entering the chamber, the
41   odor quickly diminished and was completely absent after 25-40 minutes.  One individual
42   experienced a slight tickling of the nose and throat mucous membranes after 30 minutes,
43   the two others after 40 minutes.  From 70 minutes on, all  subjects experienced a burning
44   sensation and an increasingly severe cough for the next 10-20 minutes, but coughing
45   decreased from 100 minutes on. However, the burning sensation continued and moved
46   into the lower sections of the airways and was finally felt deep in the chest. At this time,
                                             18

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   marked sputum secretion and dyspnea were noted. Toward the end of the exposure, the
 2   subjects reported the exposure conditions to be bothersome and barely tolerable. A
 3   sensation of pressure and increased sputum secretion continued for several hours after
 4   cessation of exposure (Henschler et al. 1960).
 5
 6          In a similar experiment (Henschler and Liitke 1963) groups of 4 or 8 healthy,
 7   male volunteers were exposed to 10 ppm for 6 hours or to 20 ppm for 2 hours. All
 8   subjects upon entering the chamber noted the odor which diminished rapidly. At 20 ppm
 9   minor scratchiness of the throat was felt after about 50 minutes and 3/8 experienced slight
10   headaches toward the end of the exposure period. Methemoglobin levels remained
11   within the normal range in all subjects after exposure.
12
13          Biochemical changes in bronchoalveolar lavage fluid (BALF) and blood have also
14   been studied following exposure of healthy adults to NC>2. Exposures to 2 ppm for 4
15   (Devlin et al.  1992) or 6 hours (Frampton et al. 1992) caused an influx of
16   polymorphonuclear leukocytes in BALF, 2.3 ppm for 5 hours resulted in a decrease in
17   serum glutathione peroxidase activity (Rasmusen et al. 1992), 1  and 2 ppm for 3 hours
18   caused a decrease in RBC membrane acetylcholinesterase activity, 2 ppm for 3 hours
19   resulted in an increase in peroxidized RBC lipids and glucose-6-phosphate
20   dehydrogenase activity (Posin et al. 1978), and exposure to 3 or 4 ppm for 3 hours
21   resulted in a decrease in a-1-protease inhibitor activity but not in enzyme concentration in
22   BALF (Mohsenin and Gee 1987).  Following exposure to 2 ppm for 4 hours, neutrophilic
23   inflammation was detected in bronchial washings but no changes in inflammatory cells
24   were observed in endobronchial biopsy samples (Blomberg et al. 1997).  Mucociliary
25   activity was completely stopped in healthy individuals 45 minutes after a 20-minute
26   exposure to 1.5 and 3.5 ppm (Helleday et al. 1995).
27
28   Asthmatic Subjects
29
30          Studies on the effects of NC>2  on  pulmonary function in asthmatics are
31   inconclusive and conflicting. No consistent changes in pulmonary function or reported
32   symptoms have been found in exercising asthmatic adults and adolescents exposed to
33   0.12 or  0.18 ppm for 40 minutes (Koenig et al. 1987), 0.12 ppm for 1 hour at rest (Koenig
34   et al. 1985), 0.2 ppm for 2 hours with intermittent exercise (Kleinman et al. 1983), 0.3
35   ppm for 30 minutes (Rubinstein et  al. 1990), 1 hour (Vagaggini et al. 1996), or 4 hours
36   with exercise (Morrow and Utell 1989),  0.5 ppm for  1 hour at rest (Mohsenin 1987), up
37   to 0.6 ppm for 75 minutes with intermittent exercise (Roger et al. 1990), and up to 1 ppm
38   for 4 hours (Sackner et al. 1981). No statistically significant differences between control
39   and NC>2 exposure were found for airway resistance,  symptoms, heart rate, skin
40   conductance,  or self-reported emotional  state for asthmatics exposed to 4 ppm for 75
41   minutes with intermittent exercise (Linn and Hackney 1984).
42
43          Kerr et al. (1978, 1979) studied the effects of NC>2 on pulmonary function and
44   reported other symptoms which were not included in many other studies.  The subjects
45   were asked to keep note of symptoms they experienced during exposure to 0.5 ppm for 2
46   hours, specifically cough, sputum,  irritation of mucus membranes, and chest discomfort.
                                            19

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   At this concentration the odor of NO2 was perceptible but the subjects became unaware
 2   of it after about 15 minutes. Seven of 13 asthmatics reported symptoms with exposure,
 3   compared with only one of 10 normal subjects and one of 7 subjects with chronic
 4   bronchitis.  In the group of asthmatics, two had slight burning of the eyes, one had a
 5   slight headache, three reported chest tightness, and one had labored breathing with
 6   exercise, compared with slight nasal  discharge in the normal and chronic bronchitis
 7   individuals. No changes in any pulmonary function tests were found immediately
 8   following the exposure.
 9
10          Significant group mean reductions in FEVi (-17.3% with NO2 vs -10.0% with air)
11   and specific airway conductance (-13.5% with NO2 vs -8.5% with air) occurred in
12   asthmatics  after exercise during  exposure to 0.3 ppm for 4 hours and 1/6 individuals
13   experienced chest tightness and wheezing (Bauer et al. 1985). The onset of effects was
14   delayed when exposures were by oral-nasal inhalation compared to oral inhalation; the
15   delay may have resulted from scrubbing within the upper airway.  In a similar study, 15
16   asthmatics  exposed to 0.3 ppm for 20 minutes at rest followed by 10 minutes of exercise
17   had significantly greater reductions in FEVi (-10% vs -4% with air) and partial expiratory
18   flow rates at 60% of total lung capacity, but no symptoms were reported (Bauer et al.
19   1986). In a preliminary study with 13 asthmatics exposed to 0.3 ppm for 110 minutes,
20   slight cough and dry mouth and  throat and significantly greater reduction (-11% vs -7%)
21   in FEVi occurred after exercise,  however, in a larger study, no changes in pulmonary
22   function were measured and no symptoms were reported following exposure of 21
23   asthmatics  to concentrations up to  0.6 ppm for 75 minutes (Roger et al. 1990). The mean
24   drop in FEVi for asthmatics during a 3-hour exposure to 1 ppm NO2 (-2.5%) with
25   intermittent exercise was significantly greater than the drop during air (-1.3%) exposure;
26   in BALF, levels of 6-keto-prostaglandinia were decreased and levels of thromboxane B2
27   and prostaglandin D2 were increased after NO2 exposure (Torres et  al. 1995).
28
29          Studies on the effects of NO2 on airway hyperreactivity in asthmatics have also
30   been inconclusive. Methacholine responsiveness in  asthmatics was not increased
31   following exposure to 0.25 ppm  for 20 minutes at rest plus 10 minutes of exercise
32   (Torres and Magnussen 1991)  or by exposure to 0.1 ppm for 1 hour at rest (Hazucha et
33   al. 1983). Exposure to 0.1 ppm for 1 hour caused an increase of specific airway
34   resistance in 3/20 asthmatics (the remaining 17 individuals had little or no response to
35   NO2 exposure) and enhanced the bronchoconstrictor effect of carbachol in 13/20
36   asthmatics, but the remaining  7 subjects were unaffected. When the study was repeated
37   in 4 individuals (two responders and two non-responders) at 0.2 ppm, the results were
38   variable; the two non-responders were still unaffected, while one responder had an
39   equal response and the other had a greater response to carbachol challenge compared
40   with their responses to 0.1  ppm (Orehek et al. 1976). Slight but significant potentiation
41   of airway reactivity in asthmatics occurred from exposure to 0.5 ppm NO2 for 1 hour
42   followed by methacholine challenge  (Mohsenin 1987), 0.3 ppm for 40 minutes
43   followed by isocapnic cold air hyperventilation (Bauer et al. 1986), 0.2 ppm for 2 hours
44   followed by methacholine challenge  (Kleinman et al. 1983), and 0.25 ppm for 30
45   minutes followed by isocapnic hyperventilation (Torres and Magnussen 1990).  A
46   significantly greater fall in FEVi from challenge with house dust mite antigen was
                                            20

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   reported for asthmatics compared to controls (-7.76% vs -2.85%) following exposure to
 2   0.4 ppm for 1 hour (Tunnicliffe et al. 1994), but no significant changes were found in a
 3   similar study using a 6-hour exposure (Devalia et al. 1994). Exposure of asthmatics to
 4   0.4 ppm for 3 hours significantly decreased the amount of inhaled allergen required to
 5   decrease FEVi by 20% but no changes in airway responsiveness occurred following
 6   exposure to 0.2 ppm for 6 hours; these results suggest a concentration threshold rather
 7   than a duration effect (Jenkins et al. 1999).
 8
 9          Folinsbee (1992) conducted a meta-analysis of twenty studies which measured
10   airway responsiveness in asthmatics following NO2 exposure. Eight different agents
11   were used in these studies to induce non-specific airway responsiveness and the analysis
12   was limited to exposures in the range of 0.2 to 0.3 ppm.  The fraction of asthmatic
13   subjects with an increase in airway responsiveness was significant (p < 0.01) following
14   exposures at rest, but not with exercise. When only those studies which used a
15   cholinergic agonist were analyzed, similar results were found in that a  greater proportion
16   of subjects showed an increased response in resting exposures than in exercising
17   exposures.
18
19   Subjects with Chronic Lung Disease
20
21          The results of studies on the effects of NO2 on pulmonary function in patients
22   with chronic lung disease or bronchitis are  also conflicting. No significant differences in
23   pulmonary function or symptom reporting were observed in patients with chronic
24   respiratory illness exposed to 0.3 ppm for 4 hours at rest (Hackney et al.  1992), in
25   patients with chronic obstructive pulmonary disease (COPD) exposed to NO2 levels up to
26   2 ppm for 1 hour with intermittent exercise (Linn et al. 1985), and in patients with
27   chronic bronchitis exposed to 0.5 ppm for 2 hours with exercise (Kerr et al. 1978, 1979).
28   In contrast to these reports, forced expiratory volume of COPD patients significantly
29   decreased from 18.8 L after air to 13.6 L after exposure to 0.3 ppm for 1  hour (Vagaggini
30   et al. 1996).  A significant reduction in forced vital capacity that progressed during
31   exercise (-1.2 to -8.2%) occurred in elderly COPD patients exposed to 0.3 ppm for 4
32   hours while no effects were seen in an age and gender matched healthy control group
33   (Morrow and Utell 1989, Morrow et al. 1992).
34
35          The effects of NO2 on respiratory gas exchange were investigated in patients with
36   chronic bronchitis. Inhalation of 4 and 5 ppm for 15-60 minutes significantly decreased
37   the CO diffusing capacity and arterial pO2 with no progressive changes noted over time.
38   Exposure to 5 ppm over 15 minutes resulted in  an average decrease in  CO diffusion
39   capacity of 3.8 mL/min/torr and a decrease in arterial pO2 from an average of 76.5 to 71.3
40   torr. A slight, but statistically significant, increase in airway resistance (approximately
41   20-30% above the initial value) was measured at concentrations of 1.6-5 ppm for 5
42   minutes; no effects occurred at or below 1.5 ppm (von Nieding et  al. 1973, von Nieding
43   and Wagner  1979).
44
45   2.3. Developmental/Reproductive Toxicity
46
                                             21

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1          No information was found regarding the developmental or reproductive toxicity
 2   of NC>2 in humans.
 3
 4   2.4. Genotoxicity
 5
 6       No information was found regarding the genotoxicity toxicity of NC>2 in humans.
 7
 8   2.5. Carcinogenicity
 9
10          No information was found regarding the carcinogenicity of NC>2 in humans.
11
12   2.6. Summary
13
14          In humans,  exposure to >15 ppm NC>2 in healthy individuals causes immediate
15   irritation with pulmonary edema followed by a latent period of apparent recovery. A
16   second phase of symptoms can occur after several hours or several days with the
17   development of fever with progressively more severe dyspnea, cyanosis, and cough, and
18   inspiratory and expiratory rales.  An  estimation of the concentration causing death in
19   humans is approximately >150 ppm, but no duration of exposure was given. Most case
20   reports do not contain concentrations or durations of exposure, however, welders exposed
21   to 30 and 90 ppm for 40 minutes experienced varying degrees of dyspnea, cough,
22   headache, chest tightness, nausea, and cyanosis with hospitalization required for
23   pulmonary edema,  confirmed by X-ray, at the higher concentration (Norwood et al. 1966,
24   Morley and Silk 1970).  Similar symptoms and findings of respiratory complaints were
25   reported following release of a cloud of ^64 from a railroad tank car (Bauer et al. 1996).
26
27          Epidemiological studies on the long-term effects of elevated NC>2 are
28   conflicting.  It is likely that increases in respiratory illnesses are due to NC>2 in
29   combination with other pollutants and that short-term peak concentrations are more
30   detrimental than chronic, low-level exposures. Evidence suggests that children ages 5-
31   12 have a greater risk for developing respiratory disease from long-term exposure to
32   higher concentrations, but that infants do not.
33
34          Experimental studies with both healthy and asthmatic individuals are
35   inconclusive. Negative results have been obtained in many studies with exposures up
36   to 4 ppm for 1  hour, however, other studies report positive effects on pulmonary
37   function at lower concentrations.  It should be noted that in the studies which found
38   statistically significant differences with NC>2 exposure, the changes were within 10% of
39   the measured value after air only exposure and of questionable biological significance
40   even for asthmatics. However, the available evidence also suggests  that asthmatics
41   may experience an increase in airway responsiveness from concentrations of 0.2-0.3
42   ppm.
43
44   3. ANIMAL TOXICITY DATA
45   3.1. Acute Lethality
46
                                            22

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1          Acute lethality data from NO2 exposures were located for several species. One
 2   group of investigators (Hine et al. 1970) studied the effects of varying concentration and
 3   duration of exposure in five different species of laboratory animal; these results are
 4   described separately by species below and summarized in Table 4 at the end of this
 5   section. In this study, deaths generally occurred within 2-8 hours after exposure and the
 6   majority within 24 hours. Additional data from rabbit, rat, and mouse studies were
 7   available and are in good agreement with the results of the Hine et al. study. LCso values
 8   for N2O4 were listed for four species, but duration of exposure was not given; effects
 9   were similar to those described following NO2 exposure.
10
11   3.1.1. Dogs
12
13          Greenbaum et al. (1967) exposed mongrel dogs (n = I/exposure) to 0.1% (1000
14   ppm) NO2 for 136 minutes, 0.5% (5000 ppm) for 5-45 minutes, or to 2% (20,000 ppm)
15   for 15 minutes. All dogs that were exposed to either 0.5% for 35-45 minutes or to 2% for
16   15 minutes died. Respiration became shallow and gasping with death due to pulmonary
17   edema.  Fluid was visible in the tracheobronchial tree at necropsy. Cyanosis due to
18   methemoglobin formation (78%) was noted in one animal given 2% for 15 minutes. At
19   concentrations of 0.5% and 2%, arterial pO2 and systemic arterial pressure were reduced.
20   The authors stated that pulmonary edema was caused by the action of nitrogen dioxide on
21   the alveolar lining fluid,  forming nitric and nitrous acids which, in turn, cause denaturing
22   of proteins, rupture of lysosomes, and the development of chemical pneumonitis.
23
24          Hine et al. (1970) studied the effects of varying concentration and duration of
25   NO2 exposure on mongrel dogs. Animals (n = 1-4) were exposed to 5-250 ppm NO2 for
26   periods of 30 minutes to 24 hours.  At concentrations of >40 ppm, signs of toxicity
27   included lacrimation, reddening of the conjunctivae,  and increased respiration which
28   became labored and difficult as the concentration increased. Mortalities were first
29   observed at 75 ppm for 4 hours (Table 4).  Terminally, respiration became gasping and
30   spasmodic and lung edema was observed at necropsy. Histological findings in the lungs
31   of decedents included bronchiolitis, desquamated bronchial epithelium, infiltration by
32   polymorphonuclear cells, and edema.
33
34          Kushneva and Gorshkova (1999) list an LC50 of 260 mg/m3 (70 ppm) for N2O4,
35   with the cause of death pulmonary edema; duration of exposure was not given and no
36   experimental details were included.
37
38   3.1.2. Rabbits
39
40          The 15-minute LCso for the rabbit (strain not specified; n = 5) was 315 ppm.
41   Clinical signs of toxicity included severe respiratory distress,  eye irritation, 10-15% body
42   weight suppression for two days, and death; time to death varied from 30 minutes to 3
43   days.  Gross pathology revealed darkened areas on the surface of the lungs.
44   Histopathology of the lungs of the survivors 7 and 21 days after exposure showed focal
45   accumulation of intraalveolar macrophages, some proliferation of the alveolar lining
46   epithelium, and varying amounts of inflammatory cells (Carson et al. 1962).
                                            23

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1
 2          Hine et al. (1970) studied the effects of varying concentration and duration of
 3   NO2 exposure on rabbits (strain not specified). Animals (n = 2-8) were exposed to 5-200
 4   ppm NO2 for 30 minutes up to 24 hours. At concentrations of >40 ppm, signs of toxicity
 5   included lacrimation, reddening of the conjunctivae, and increased respiration which
 6   became labored and difficult as the intoxication increased. One mortality was observed
 7   at 75 ppm for 60 minutes but none occurred at 2 hours making the death after the 1 hour
 8   exposure questionable (Table 4).  Terminally, respiration became gasping and spasmodic
 9   and lung edema was observed at necropsy. Histological findings in the lungs of
10   decedents included bronchiolitis, desquamated bronchial epithelium, infiltration by
11   polymorphonuclear cells, and edema.
12
13          In a similar study, rabbits (strain not specified; n = 3) were exposed to 125, 175,
14   250, 400, 600, or 800 ppm NO2 for 10 minutes (Meulenbelt et al. 1994). Two of three
15   animals given 800 ppm died 7-21  hours after exposure. Lung weights were significantly
16   higher and lung homogenates contained greater amounts of protein and higher levels of
17   lactate dehydrogenase (LDH), glutathione peroxidase, and glucose-6-phosphate
18   dehydrogenase activity in animals exposed to >250 ppm. Bronchoalveolar lavage fluid
19   from animals exposed to > 175 ppm contained greater amounts of protein and albumin,
20   and higher levels of LDH and angiotensin converting enzyme activity than unexposed
21   controls and all treated groups had increased numbers of neutrophilic leucocytes. Dose-
22   related increases in severity of centriacinar catarrhal pneumonitis, macrophage influx,
23   and neutrophilic leucocytes were observed on histopathological examination of the lungs.
24   Edema occurred at >250 ppm, subpleural hemorrhaging at >400 ppm, and desquamation
25   of the bronchiolar epithelium was seen at >600 ppm.
26
27          Kushneva and Gorshkova  (1999) list an LC50 of 320 mg/m3 (86 ppm) for N2O4,
28   with the cause of death pulmonary edema; duration of exposure was not given and no
29   experimental details were included.
30
31   3.1.3. Guinea Pigs
32
33          Hine et al. (1970) also studied the effects of varying concentration and duration of
34   exposure in the guinea pig (strain  not specified). Animals (n = 2-6) were exposed to 5-
35   200 ppm NO2 for 30 minutes up to 8 hours. At concentrations of >40 ppm, signs of
36   toxicity included lacrimation, reddening of the conjunctivae, and increased respiration
37   which became labored and  difficult as the intoxication increased. Mortalities were first
38   observed at 50 ppm for 1 hour (Table 4). Terminally, respiration became gasping and
39   spasmodic and lung edema was observed at necropsy. Histological findings in the lungs
40   of decedents included bronchiolitis, desquamated bronchial epithelium, infiltration by
41   polymorphonuclear cells, and edema.
42
43          To determine the sensitivity of adult and  neonate animals to NO2 inhalation,
44   Duncan-Hartley guinea pigs aged 5, 10, 21, 45, 55, and 60 days were exposed
45   continuously for 3 days to 2 or 10 ppm (Azoulay-Dupuis et al. 1983). A total of 17-27
46   animals were studied in each age group and litter exposures prior to weaning included the
                                            24

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   dam.  At 10 ppm, clinical signs of toxicity in adults over 45 days old included moving
 2   with difficulty, reduced food and water consumption, and hyperventilation. Body weight
 3   gain was decreased until 21 days and body weight was reduced after 45 days in all
 4   exposed animals. These effects were most pronounced in the dams. Mortality in the
 5   high-exposure group increased with age with 4% of 5-day olds, up to 60% of 55-day
 6   olds, and 67% of dams dying. Further, most of the older animals died after the first 24
 7   hours of exposure whereas the younger animals died later in the 3-day period.  At 2 ppm,
 8   lung histopathology was normal until animals were 45 days of age when thickening of the
 9   alveolar walls, infiltration by PMNs, and alveolar edema were observed; in dams
10   bronchioles were devoid of cilia in some areas. At 10 ppm, guinea pigs of all ages were
11   affected by these changes which were more pronounced in older animals.
12
13   3.1.4. Rats
14
15          Five-, 15-, 30-, and 60-minute LCso values for the male rat (100-120 g; strain
16   not specified; n = 10) are 416, 201, 162, and 115 ppm, respectively. Clinical signs of
17   toxicity included severe respiratory distress, eye irritation, 10-15% body weight
18   suppression, and death; time to death varied from 30 minutes to 3  days. Gross
19   pathology revealed darkened areas on the surface of the lungs, and, in some instances,
20   purulent nodules involving the entire lungs of some of the survivors (Carson et al.
21   1962).
22
23          An older  study (Gray et al. 1954) reported LCso values for male rats (200-300 g;
24   strain not specified;  n = 10) of 1445 ppm for 2 minutes,  833 ppm for 5 minutes, 420
25   ppm for 15 minutes, 174 ppm for 30 minutes, 168 ppm for 60 minutes, and 88 ppm for
26   240 minutes.  Deaths were attributed to pulmonary edema.  The differences in LCso
27   values between this  study and Carson et al. (1962) may be due to the differences in size
28   and age of the rats used in each.
29
30          Meulenbelt et al. (1992a,b) investigated the effects of both concentration and
31   duration of exposure in Wistar rats. The effect of concentration was studied by exposing
32   6-9 rats/group to 25, 75, 125, 175 or 200 ppm NC>2 for 10 minutes. No signs of toxicity
33   were observed at 25 ppm.  Stertorous respiration was heard in animals exposed to 175
34   and 200 ppm. Rats exposed to >75 ppm had significantly  increased lung weight, and
35   subpleural hemorrhages and pale discol orations of the lung were observed grossly.
36   Histologically, the lungs from these animals showed atypical pneumonia, edema, focal
37   desquamation of the terminal bronchiolar epithelium, increased numbers of macrophages
38   and neutrophilic  leucocytes, and interstitial thickening of the centriacinar septa (175 and
39   200 ppm only) with the severity increasing at the higher concentrations.  One rat died 14-
40   20 hours after exposure in both the 175 ppm and 200 ppm groups. Biochemical changes
41   in bronchoalveolar lavage fluid included concentration-dependent increases in protein
42   and albumin concentrations, angiotensin converting enzyme activity,  p-glucuronidase
43   activity, and neutrophilic leukocytes.
44
45          Duration  of exposure was investigated by exposing 6 rats/group to either 175 ppm
46   for 10, 20, or 30  minutes or to 400 ppm for 5, 10, or 20 minutes (Meulenbelt et al.
                                            25

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   1992a,b).  Stertorous respiration was heard in animals for all exposure times at both
 2   concentrations and lung weight was significantly higher than that of the controls at all
 3   exposure durations.  At 175 ppm, 5/6 rats died in the 20 and 30 minute groups and at 400
 4   ppm, 6/6 rats died in the 10 and 20 minute groups. Necropsy revealed foamy,
 5   seroanguinous fluid  in the trachea,  subpleural bleeding, and pale discoloration.
 6   Histological alterations were similar to those described above.  Methemoglobin levels,
 7   measured after exposure to 175 ppm for 10 minutes, were not elevated, but plasma nitrate
 8   levels were significantly higher than controls.
 9
10          Hine et al. (1970) also studied the effects of varying concentration and duration of
11   exposure in Long-Evans rats. Animals (n= 4-31) were exposed to 5-250 ppm NO2 for
12   durations of 30 minutes up to 24 hours. At concentrations of >40 ppm, signs of toxicity
13   included lacrimation, reddening of the conjunctivae, and increased respiration which
14   became labored and difficult as the intoxication increased. Mortalities were first
15   observed at 50 ppm for 24 hours (Table 4).  Terminally, respiration became gasping and
16   spasmodic and lung  edema was observed  at necropsy. Histological findings in the lungs
17   of decedents included bronchiolitis, desquamated bronchial epithelium, infiltration by
18   polymorphonuclear cells, and edema.
19
20          Kushneva and Gorshkova (1999) list an LC50 of 105 mg/m3 (28 ppm) for N2O4,
21   with the cause of death pulmonary  edema; duration of exposure was not given and no
22   experimental details were included.
23
24   3.1.5. Mice
25
26          BALB/c mice (n = 5-7) were exposed to 5, 20, or 40 ppm NO2 for 12 hours
27   (Hidekazu and Fujio 1981). Body weight was markedly decreased  1 and 2 days after
28   exposure to 20 and 40  ppm and 3/38 (7.8%) animals exposed to 40 ppm died within 2
29   days of the exposure.
30
31          Hine et al. (1970) studied the effects of varying concentration and duration of
32   exposure in Swiss-Webster mice.  Animals (n= 5-14) were exposed to 5-250 ppm NO2
33   for durations of 30 minutes up to 24 hours.  At concentrations of >40 ppm, signs of
34   toxicity included lacrimation, reddening of the conjunctivae, and increased respiration
35   which became labored and difficult as the intoxication increased. Mortalities were first
36   observed at 50 ppm for 24 hours (Table 4).  Terminally, respiration became gasping and
37   spasmodic and lung  edema was observed  at necropsy. Histological findings in the lungs
38   of decedents included bronchiolitis, desquamated bronchial epithelium, infiltration by
39   polymorphonuclear cells, and edema.
40
41          Kushneva and Gorshkova (1999) list an LC50 of 190 mg/m3 (51 ppm) for N2O4,
42   with the cause of death pulmonary  edema; duration of exposure was not given and no
43   experimental details were included.
                                            26

-------
     NITROGEN OXIDES
NAC/Interim (NO2)/Proposed (N2O4): 12/2008
TABLE 4: Summary of NO2 Mortality for Five Species
Cone.
(ppm)
50
75
100
150
200
Time (hr)
1
6
24
1
2
4
8
0.5
2
4
8
0.5
1
2
4
0.08
0.17
0.33
0.50
Rat
0/17
0/12
3/10
3/31
1/12
7/12
12/12
0/5
8/8
29/29
2/10
10/13
10/12
4/4
6/12
8/12
5/5
4/4
Mouse
0/5
0/5
5/10
1/6
2/6
5/6
6/6
2/10
13/14
10/10
10/10
-
4/6
6/6
6/6
Guinea Pig
1/6
4/6
1/4
3/4
2/4
4/4
1/2
3/4
3/4
3/3
2/2
Rabbit
0/4
0/4
1/8
0/6
2/8
6/8
1/3
2/4
3/4
1/6
3/4
0/2
1/2
2/4
Dog
0/1
0/2
0/2
0/2
1/3
1/4
0/2
1/3
2/2
2/3
2/2
2
3
4
5
6
7
Data from Hine et al. 1970, p.
majority within 24 hours.

3
3

.2.
.2.

Nonlethal Toxicity
1. Monkeys
                            206; deaths generally occurred within 2-8 hours after exposure and the
 9          Squirrel monkeys (n = 2-6/group) were exposed to 10-50 ppm NC>2 for 2 hours
10   with respiratory function monitored during exposure (Henry et al. 1969).  Exposure to 35
11   or 50 ppm resulted in a markedly increased respiratory rate and decreased tidal volume
12   during exposure which returned to normal by the seventh day post exposure. Only slight
13   effects on respiratory function were noted at 15 and 10 ppm. Mild histopathological
14   changes in the lungs were noted after exposure to 10 and 15 ppm, however, marked
15   changes in lung structure were observed after exposure to 35 and 50 ppm. At 35 ppm,
16   areas of the lung were collapsed with basophilic alveolar septa, in other areas the alveoli
17   were expanded with septal wall thinning, and the bronchi were moderately inflamed with
18   some proliferation of the surface epithelium. At 50 ppm, extreme vesicular dilatation of
19   alveoli or total collapse was observed, lymphocyte  infiltration  was seen with extensive
20   edema, and surface erosion of the epithelium of the bronchi was observed. In addition to
21   the effects on the lungs, interstitial fibrosis (35 ppm) and edema (50 ppm) of cardiac
22   tissue, glomerular tuft swelling in the kidney (35 and 50 ppm), lymphocyte infiltration in
23   the kidney and liver (50 ppm), and congestion and centrilobular necrosis in the liver (50
24   ppm) were observed. Although no animals died following the single exposure to 50 ppm,
25   one animal died following a second exposure two months after the first exposure
26   suggesting that some of the lesions were irreversible.
27
28   3.2.2. Dogs
                                             27

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2          Carson et al. (1962) conducted a series of experiments on dogs (strain not
 3   specified; n = 2) at target concentrations of NO2 approximately 50% and 25% of the
 4   LC50 for the rat (see Sec. 3.1.5).  The actual analyzed concentrations varied slightly, but
 5   were within 10% of target. Dogs exposed to 164 ppm for 5 minutes, 85 ppm for 15
 6   minutes, or 53 ppm for 60 minutes (approximately 50% of the rat LCso) had some
 7   respiratory distress during exposure, a mild cough, and eye irritation all of which cleared
 8   within two days after exposure. Dogs exposed to 125 ppm for 5 minutes, 52 ppm for 15
 9   minutes, or 39 ppm for 60 minutes (approximately 25% of the rat LCso) showed only
10   mild sensory effects.  No gross or microscopic lesions were noted in any dog.
11
12          Greenbaum et al. (1967) exposed mongrel dogs (n = 1) to 0.1% (1000 ppm) NO2
13   for 136 minutes or to 0.5% (5000 ppm) for 5-45 minutes. A concentration of 0.1% did
14   not cause death and the one dog exposed to this concentration remained in good
15   condition throughout the exposure. Exposures to 0.5% for 15 and 22 minutes were not
16   lethal but resulted in respiratory distress which gave rise to anxiety for about 2 hours
17   then resolved without therapy. Histopathologic examination of the lungs was not
18   performed.
19
20          No treatment-related changes in behavior or clinical signs were observed in
21   mongrel dogs (n = 1) exposed to 10-40 ppm NO2 for 6 hours (Henschler and Liitke
22   1963).
23
24          Mongrel dogs (number of animals not stated) exposed to 20 ppm NO2 for up to 24
25   hours showed minimal signs of irritation and changes in behavior. Microscopic lesions
26   were described as questionable evidence of lung congestion and interstitial inflammation
27   for up to 48 hours postexposure (Hine et al. 1970).
28
29          Pulmonary ultrastructural changes were examined in beagle dogs (n = 1) exposed
30   to 3-16 ppm NO2 for 1 hour (Dowell et al. 1971). Intraalveolar edema occurred in most
31   dogs exposed to >7 ppm and was associated with impaired surfactant activity and lung
32   compliance. Ultrastructural alterations included wide-spread bleb formation,  loss of
33   pinocytic vesicles, and mitochondrial swelling of endothelial cells.  Exposure to 3 ppm
34   resulted in bleb formation in the  alveolar endothelium (observed by EM) without
35   biochemical or physiological  changes.
36
37   3.2.3. Rabbits
38
39          Rabbits (strain not specified; number of animals not given) exposed to 20 ppm
40   NO2 for up to 24 hours showed minimal signs of irritation and changes in behavior.
41   Microscopic lesions were described as questionable evidence of lung congestion and
42   interstitial inflammation for up to 48 hours postexposure (Hine et al. 1970).
43
44          Rabbits exposed to 10 ppm NO2 for 2 hours showed accelerated alveolar
45   particle clearance (Vollmuth et al. 1986) and altered pulmonary arachidonic acid
46   metabolism (Schlesinger et al. 1990).  Continuous exposure of rabbits to 3.6 ppm
                                            28

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   NC>2 for 6 days did not cause morphological changes in the lungs (Hugod 1979).
 2
 3          Rabbits (strain, sex, number not given) exposed continuously for up to 20
 4   hours to 7, 14, or 28 ppm NO2 had an increase in polymorphonuclear leukocytes in
 5   the lavage fluid throughout the exposure (Gardner et al.  1977).
 6
 7   3.2.4. Sheep
 8
 9          Lung mechanics, hemodynamics, and blood chemistry were assessed in crossbred
10   sheep (n = 5-6) exposed by nose-only or lung-only (to mimic mouth breathing) to 500
11   ppm NC>2 for 15 minutes; in another group exposed by lung-only, bronchoalveolar lavage
12   fluid content was examined after a 20 minute exposure to 500 ppm (Januszkiewicz and
13   Mayorga 1994). No changes in hemodynamics or blood chemistry occurred in  either
14   group. Mean inspired minute ventilation was  significantly increased, resulting in
15   increased breathing rate and decreased mean tidal volume, in the lung-only exposure
16   group, but not the nose-only group. Both nose-only and lung-only exposure groups had
17   significantly increased lung resistance and decreased dynamic lung compliance.
18   Histopathologic examination of tissue from the lung-only exposed group revealed
19   exudative fluid distributed in a patchy lobular pattern with mild neutrophil infiltration;
20   little evidence of exudation was seen in the nose-only exposed group. Epithelial cell
21   number and total protein in bronchoalveolar lavage fluid were significantly increased in
22   the NC>2 exposed animals while macrophage number was decreased.
23
24          Airway reactivity to aerosolized carbachol  was evaluated in crossbred sheep (n =
25   4-10) exposed to 7.5 or 15 ppm NC>2 for 2 hours (Abraham et al. 1980). Group means for
26   pulmonary resistance, bronchial reactivity to carbachol,  and static lung compliance were
27   similar to those from controls at both concentrations.  However, following exposure to
28   7.5 ppm NC>2,  5 of 9 animals showed an increase in pulmonary resistance after carbachol
29   exposure of 57% above baseline and following exposure to 15 ppm NC>2, 9 of 10 animals
30   responded with either bronchoconstriction or hyperreactivity. In a concurrent
31   experiment, sheep were exposed to 15 ppm NC>2 for 4 hours (Abraham et al. 1980).
32   Mean pulmonary resistance was significantly increased from the preexposure value, but
33   there were no changes in pulmonary hemodynamics or clinical signs of distress.
34
35   3.2.5. Guinea Pigs
36
37          Guinea pigs (strain not specified; number of animals not given) exposed to 20
38   ppm NC>2 for up to 24 hours showed minimal signs of irritation and changes in behavior.
39   Microscopic lesions were described as questionable evidence of lung congestion and
40   interstitial inflammation for up to 48 hours postexposure (Hine et al. 1970).  Guinea pigs
41   exposed to 9 and 13 ppm for 2 hours or to 5.2 and  6.5 ppm for 4 hours had significantly
42   increased respiratory rate and decreased tidal volume with complete recovery after
43   cessation of exposure (Murphy et al. 1964).
44
45          Hartley guinea pigs (n = 5-16) maintained on an ascorbic acid-deficient diet had
46   increased lung lavage fluid protein following exposure to 4.8 ppm NC>2 for 3 hours and
                                            29

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   increased wet lung weight, increased nonprotein sulfhydryl and ascorbic acid content of
 2   the lungs, and decreased a-tocopherol content of the lungs following exposure to 4.5 ppm
 3   for 16 hours.  These changes were not seen in animals maintained on normal guinea pig
 4   diets (Hatch et al. 1986).  Similarly, vitamin C-deficient male Hartley guinea pigs (n = 3-
 5   12) exposed to 1, 3,  or 5 ppm for 72 hours had significantly increased protein and lipid
 6   content in lavage fluid (Selegrade et al. 1981). No effects were seen at 0.4 ppm. At 5
 7   ppm,  50% of the animals died and histopathology revealed multifocal interstitial
 8   pneumonia. When the exposure to 5 ppm was shortened to 3 hours, lavage protein was
 9   increased with the peak effect 15 hours post-exposure.
10
11          Guinea pigs  (strain not specified; n = 12-18) were exposed to 20, 40, or 70 ppm
12   NO2 for 30 minutes  followed by a 30-minute exposure to aerosolized albumin; this
13   regimen was repeated 5-7 times at intervals of several days (Matsumura  1970). During
14   the first exposure to 70 ppm, labored breathing, though not severe, was observed in
15   "some" animals, but was not seen with subsequent exposures.  Immediately after the fifth
16   exposure to antigen, one-half of the animals in the 70 ppm group showed enhanced
17   airway sensitization (anaphylactic attacks). No effects were seen at 20 or 40 ppm.
18
19          Changes in airway responsiveness to histamine were investigated in Hartley
20   guinea pigs (number of animals not given) exposed to 7-146 ppm NO2 for 1 hour
21   (Silbaugh et al. 1981). Pulmonary function measurements and histamine challenge tests
22   were performed 2 hours before and at about 10 minutes, 2 and 19 hours after NO2
23   exposure. At 10 minutes after exposure, increased sensitivity to histamine occurred at
24   concentrations >40 ppm but returned to baseline thereafter. Concentration-related
25   significantly increased breathing frequency and decreased tidal volume were measured
26   at 10 minutes (exact concentrations not specified) and remained correlated with
27   concentration at 2 and 19 hours.
28
29   3.2.6. Hamsters
30
31          Syrian golden hamsters (n = 5) were administered 28 ppm NO2 for 6, 24, or 48
32   hours and histopathological changes in the lungs were examined by light and electron
33   microscopy (Case et al. 1982, Gordon et al. 1983).  The bronchiolar epithelium showed
34   ciliary loss and surface membrane damage, loss of ciliated cells, and epithelial flattening
35   at 24 and 48 hours and epithelial hyperplasia, nonciliated cell hypertrophy, and loss of
36   tight junctions between type I pneumocytes at 48 hours.
37
38   3.2.7. Ferrets
39
40          Weanling domestic ferrets (n = 4-6), 6 weeks of age, were exposed to 5, 10, 15,
41   or 20  ppm NO2 for 4 hours (Rasmussen 1992). A transient inflammatory response was
42   evident as a significantly increased number of neutrophils in the lavage fluid up to 48-
43   hours postexposure at all concentrations. Morphometrically, dose-related decreased
44   alveolar size and thickened alveolar walls indicative of exposure were observed in the
45   lungs.
46
                                            30

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   3.2.8. Rats
 2
 3          Only one study was found in which rats were exposed to N2O4 and pulmonary
 4   lesions were similar to those described following NO2 exposure. Male Wistar rats
 5   exposed to 43 ppm N2O4 for 15 minutes had increased lung weight, lung edema, and
 6   hemorrhaging (Yue et al. 2004). The chamber atmosphere was generated by injecting
 7   liquid N2O4 and heating to evaporate it. Thus it is likely that much of the dimer was
 8   converted to NO2.
 9
10          Pulmonary injury from NO2 as indicated by increases in lung weight was assessed
11   in male Fischer 344 rats (n = 6-12) following exposure to 10, 25, or 50 ppm for 5,  15, or
12   30 minutes or to  100 ppm for 5 or 15 minutes (Stavert and Lehnert 1990).  No significant
13   changes in lung weight occurred in rats exposed to 10 ppm for 30 minutes or to 25-50
14   ppm for up to 15 minutes. Significant increases in lung wet weight and right cranial lobe
15   dry weight were found following exposure to 50 ppm for 30 minutes or to 100 ppm for 5
16   and 15 minutes.  However, histological evidence of lung injury was seen in animals
17   exposed to 25 ppm for 30 minutes, 50 ppm for >5 minutes, and 100 ppm for 5 and 15
18   minutes.  Findings included accumulation of fibrin, increased numbers of
19   polymorphonuclear leukocytes and macrophages, extravasated erythrocytes, and type II
20   pneumocyte hyperplasia, the severity of which increased with concentration and duration
21   of exposure.
22
23          In a more expanded study, Lehnert et al. (1994) determined that exposure
24   concentration was more important than exposure duration in the severity of lung injury.
25   Male Fischer 344 rats (n = 8-12) were  exposed to 25, 50, 75, 100,  150, 200, or 250 ppm
26   NO2 for durations ranging from 5-30 minutes. Lung wet weight was significantly
27   increased following exposure to > 150 ppm for 5 minutes, 100 ppm for 15 minutes, or 75
28   ppm for 30 minutes and further increases were observed as exposure duration increased.
29   The pulmonary edematous response to a given concentration was not proportional to
30   duration, however,  increasing concentrations produced proportional increases in lung wet
31   weight when similar exposure durations were compared. Histologically, fibrin and type II
32   cell hyperplasia were observed following 5-minute exposures to >50 ppm the severity
33   increased proportionally to concentration. As further confirmation of concentration-
34   dependent lung injury, rats were exposed to 1-minute bursts of 500-2000 ppm. The
35   severity of the resulting pulmonary edema (as measured by  lung wet weight) was directly
36   proportional to exposure concentration. The authors concluded that brief exposures to
37   high concentrations of NO2 are more injurious than longer duration exposures to lower
38   concentrations. Dietary taurine (an antioxidant) was not protective against the increase in
39   lung wet weight and exercise potentiated the severity of the pulmonary edema.
40
41          The concentration-dependent response of the lung to NO2 was confirmed in
42   another study in which Sprague-Dawley rats (n = 5-6) were exposed to 3.6-14.4 ppm for
43   durations of 6-24 hours/day for 3 days (Gelzleichter et al. 1992).  Increases in protein
44   content and cell types in lavage fluid demonstrated that the magnitude of lung injury was
45   a function of exposure concentration.
46

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1          Carson et al. (1962) conducted a series of experiments at NO2 concentrations
 2   approximating 50%, 25%, and 15% of the rat LC50 levels. At the 50% level, rats (strain
 3   not specified; n = 30) exposed to 190 ppm for 5 minutes, 90 ppm for 15 minutes, or 72
 4   ppm for 60 minutes showed signs of severe respiratory distress and eye irritation lasting
 5   about two days; lung-to-body weight ratios were significantly increased during the first
 6   48 hours after exposure.  Pathological examination showed darkened areas of the lungs,
 7   pulmonary edema,  and an increased incidence of chronic murine pneumonia.  Rats
 8   exposed to 104 ppm for 5 minutes, 65 ppm for 15 minutes, or 28 ppm for 60 minutes
 9   (about 25% of the LCsos) showed some respiratory distress or mild signs of nasal
10   irritation during exposure but lung-to-body weight ratios were increased only at the 104
11   and 65 ppm levels.  No gross lesions were observed, but pulmonary edema was seen
12   microscopically. No adverse clinical signs of toxicity or pathological changes were
13   seen in rats exposed at 15% of the LCso (74 and 33 ppm for 5 and 15 minutes,
14   respectively).
15
16          Histological changes were examined in the lungs of male rats (strain and number
17   of animals not given) continuously exposed to 17 ppm NC>2 (Stephens et al. 1972).
18   After 2 hours of exposure, there was some precapillary and postcapillary engorgement
19   in the alveoli.  Loss of cilia and occasional alveolar type I cell swelling were detectable
20   by 4 hours, the terminal bronchiolar epithelium had become uniform by 16 hours,
21   maximal macrophage numbers were reached by 24 hours, cellular hypertrophy had
22   begun by 48 hours, and mitotic figures became more prevalent in the epithelium of the
23   terminal bronchiole between 16 and 48 hours. Type I alveolar cells appeared to be the
24   most sensitive to NC>2 insult.
25
26          Results similar to those described above were obtained in a morphological study
27   of the Wistar rat lung (number of animals not given) following exposure to 20 ppm NC>2
28   for 20 hours (Hayashi et al. 1987). Cytoplasmic blebbing occurred in a small number of
29   type I cells immediately  after exposure.  Swelling and hyperplasia of type II cells and
30   pinocytotic vesicles of endothelial cells in capillaries followed by interstitial edema in the
31   alveolar walls were observed between days 5-15 postexposure.  Twenty days after
32   exposure the lesions lessened and the lungs appeared normal after 35 days. Other studies
33   have confirmed alveolar and interstitial edema, bronchiolitis, bronchiolar epithelial cell
34   hyperplasia, loss of cilia, necrosis of type I cells, and/or type II cell hyperplasia 1-3  days
35   after exposure to 26 ppm NO2 for 24 hours (Schnizlein et al. 1980, Hillam et al. 1983) or
36   to 20 ppm for 24 hours (Rombout et al. 1986).
37
38          Long-Evans rats  (number of animals not given) exposed to 20 ppm NO2 for up
39   to 24 hours showed minimal signs of irritation and changes in behavior.  Microscopic
40   lesions were described as questionable evidence of lung congestion and interstitial
41   inflammation for up to 48 hours postexposure (Hine et al. 1970).
42
43          The effects  of NO2 on the lung were compared between  Sprague-Dawley neonatal
44   and adult rats (number of animals not given). Animals,  1-40 days old, were continuously
45   exposed to 14 ppm for 24, 48, or 72 hours. Prior to weaning (20 days old), exposure
46   resulted in only minor injury and loss of cilia from epithelial cells lining the terminal
                                            32

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   airways.  Subsequent to weaning, there was a progressive increase in lung injury with
 2   maximum response reached at about 35 days of age (Stephens et al.  1978).
 3
 4          In a similar study to determine the sensitivity of adult and neonate animals to NO2
 5   inhalation, Wistar rats (number of animals not given) aged 5-60 days were exposed
 6   continuously for 3 days to 2 or 10 ppm (Azoulay-Dupuis et al. 1983). Litter exposures
 7   prior to weaning included the dam.  No clinical signs of toxicity or deaths were observed
 8   in animals of any age except for body weight loss in dams of the 10 ppm-group. At 2
 9   ppm, lung histopathology was normal in all animals. At 10 ppm in animals 45 days old
10   and older, fibrinous deposits were observed in the alveoli and the tracheal and
11   bronchiolar epithelia were occasionally devoid of cilia.
12
13          Alterations in lavage fluid have been assessed in male Long Evans rats (n = 6)
14   following exposure to 10, 20, 30, or 40 ppm for 4 hours. Cell-free lavage fluid contained
15   elevated lactate dehydrogenase (LDH), malate dehydrogenase (MDH), isocitrate
16   dehydrogenase (IDH), glucose-6-phosphate dehydrogenase (GDH), acid phosphatase
17   (AP), and aryl sulfatase (AS) activity levels after exposure to >30 ppm.  Total protein and
18   sialic acid were increased after exposure to >20 ppm. Protein and sialic acid
19   concentrations and AP activity level were similar to those in plasma indicating
20   transudation into the airways (Guth and Mavis  1985). The increases  in LDH,  MDH, and
21   GDH activity were significantly attenuated in animals maintained on diets providing
22   1000 mg/kg of a-tocopherol, suggesting that lipid peroxidation is involved in NO2
23   induced lung injury (Guth and Mavis 1986). Antioxidants in the lung were depleted, lipid
24   peroxidation products were elevated, and total cell count in BALF and alveolar
25   macrophage count were decreased while epithelial cell count was increased following
26   exposure of male Sprague-Dawley rats (n = 5) to 200 ppm for 15 minutes (Elsayed et al.
27   2002). Another study found changes in fatty acid composition of alveolar lavage
28   phospholipids following exposure of Wistar rats (n = 6) to 20 ppm NO2 for 12 hours
29   (Kobayashi et al. 1984).  Increases in lavageable protein, polymorphonuclear
30   lymphocytes, and alveolar macrophages were also observed following exposure of male
31   Fischer 344 rats (n not given) to 100 ppm for 15 minutes (Lehnert et al.  1994).
32
33          Changes in minute ventilation, VE, were measured in male Fischer 344 rats (n =
34   12) following exposure to 100, 300, or  1000 ppm NO2 for 1-20 minutes (Lehnert et al.
35   1994). In general, reductions in VE were greater with the higher concentrations. For
36   example, reductions of about 7% and 15% were measured during 15- and 20-minute
37   exposure to  100 ppm, while reduction of about 20% and 28% were measured during 1-
38   and 2-minute exposures to 1000 ppm.  Similarly, male Sprague-Dawley rats (n  = 5)
39   exposed to 200 ppm for 15 minutes showed a decrease in minute ventilation that was due
40   to a decline in tidal volume but not in frequency of breathing (Elsayed et al. 2002).
41
42          Male Porton rats (n = 4) were exposed to an atmosphere of oxides of nitrogen
43   that was produced by mixing nitrogen dioxide and nitric oxide (Brown et al. 1983).
44   Exposures were to 518 ppm for 5 minutes or 1435  ppm for 1 minute. No clinical signs
45   of toxicity were observed during exposure to either concentration but "stertorous
46   respirations" appeared within 24 hours. Histologically, initial lung damage showed
                                            33

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   thickening and blebbing of the alveolar epithelium followed by a latent period of about 6
 2   hours after which development of edema of the interstitium and alveolar septum was
 3   observed. The early changes were attributed to a direct oxidant effect.
 4
 5          Changes in lung immunity after NC>2 exposure have been described as increased
 6   specific IgE, IgA, and IgG liters following exposure to 87 ppm for 1 hour (Siegel et al.
 7   1997) or 5 ppm for 3  hours (Gilmour 1995), increased number of IgG anti-sheep red
 8   blood cell antibody-forming cells in the lung-associated lymph nodes (Schnizlein et al.
 9   1980), and cell proliferation in the spleen and thoracic lymph nodes (Hillam et al. 1983)
10   following exposure to 26 ppm for 24 hours.
11
12   3.2.9. Mice
13
14          Swiss-Webster mice (number of animals not given) exposed to 20 ppm NC>2 for
15   up to 24 hours showed minimal signs of irritation and changes in behavior.
16   Histologically, there was questionable evidence of lung congestion and interstitial
17   inflammation for up to 48 hours postexposure (Hine et al. 1970). The voluntary
18   running activity of mice on an activity-wheel was 80% and 17% of preexposure levels
19   during 6-hour exposures to 7.7 and 20.9 ppm, respectively (Murphy et al. 1964).
20
21          Female CD-I  mice (n = 29-60) were examined for phenobarbital-induced
22   sleeping time after a 3 hour, whole body exposure to 0.125-5.0 ppm NC>2 (Miller et al.
23   1980). Sleeping time was significantly increased in animals exposed to >0.25 ppm
24   compared with air exposed controls.  The authors stated that no effects in males were
25   observed until after three days of exposure (data not included).  In contrast, the effect in
26   females decreased after the third day of exposure suggesting some tolerance may have
27   developed.
28
29          The alveolar septum from two female NMRI mice was examined
30   microscopically thirty-six hours after exposure to 35 ppm for 6 hours (Dillmann et al.
31   1967). Morphometric measurements found that the arithmetic mean thickness of the
32   alveoli was approximately 1.5x that of unexposed controls. No changes in the numbers
33   or types of cells present were observed and no interstitial edema was  seen with
34   ultrastructure examination by electron microscopy.
35
36          Male CD-I mice (n = 5-9) were exposed to  50-140 ppm NC>2 for 1 hour and
37   biochemical and histological responses assessed immediately and 48-hours after exposure
38   (Siegel et al. 1989). Immediately after exposure to  140 ppm, cell death was visible in the
39   terminal bronchioles  and there were significant increases in protease inhibitor activity,
40   pulmonary protein, and lung wet weight.  Two days after exposure to 140 ppm, the
41   histological damage was exacerbated with complete obliteration of the alveolar structure,
42   progressive edema and congestion of the lungs, hypertrophy and hyperplasia of the
43   epithelial cells, and increased numbers of intraalveolar macrophages and neutrophils.
44   Also two days after exposure, there were dose-related increases in p-glucuronidase,
45   lactate dehydrogenase, and choline kinase activity as well as increased protease inhibitor
46   activity, pulmonary protein, and lung wet weight.
                                            34

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2          To examine the effects of NC>2 on gaseous exchange in the lung, JCL:ICR mice (n
 3   =6) were exposed to 5,  10, or 20 ppm for 24 hours (Suzuki et al. 1982).  Significantly
 4   increased lung wet weight and lung water content occurred at 10 and 20 ppm. In animals
 5   exposed to 5 ppm, the gaseous exchange and metabolic rate of 62 and CO2 were
 6   accelerated while in animals exposed to 10 and 20 ppm, gaseous exchange in the lung
 7   was inhibited.
 8
 9          Continuous exposure of C56B1/6 mice (n = 60) to 20 ppm NC>2 for four days
10   resulted in significantly decreased food consumption and body weight, but no deaths
11   (Bouley etal. 1986).
12
13   3.3. Developmental/Reproductive Toxicity
14
15          The postnatal effects of prenatal exposure to NC>2 were investigated (Tabacova et
16   al. 1985). Pregnant Wistar rats (n = 20) were exposed to 0.265, 0.053, 0.53, or 5.3 ppm
17   NC>2 for 6 hours/day throughout pregnancy. Maternal effects were not reported or
18   discussed. Pup viability and body weight of the 5.3 ppm-group were significantly (p <
19   0.05) less than those of controls on lactation day 21. Exposure to >0.53 ppm resulted in
20   developmental delays and exposure to >0.053  ppm caused disturbances in neuromotor
21   development. Also at the two highest concentrations, hexobarbital sleeping time was
22   increased in the offspring and correlated with altered biochemical parameters in the
23   liver.
24
25   3.4. Genotoxicity
26
27          Three-week old male Sprague-Dawley rats were exposed by inhalation to 8, 15,
28   21, or 27 ppm NC>2 for 3 hours, maintained overnight before sacrifice, and lung cells
29   isolated.  At concentrations of >15 ppm,  there was a concentration-related increase in
30   mutation to ouabain resistance in lung cells. Concentration-dependent increases in
31   chromosome aberrations were observed at 8 and 27 ppm NC>2, the only concentrations
32   analyzed for aberrations (Isomura et al. 1984).
33
34   3.5. Subchronic and Chronic Toxicity/Carcinogenicity
35
36          Respiratory effects in humans from long-term environmental exposure to NC>2
37   have been discussed with the epidemiology studies in Section 2.2.2.
38
39          The effect of NC>2 on promotion of lung tumorigenesis induced by 7V-bis(2-
40   hydroxypropyl) nitrosamine (BHPN) was investigated in male Wistar rats (Ichinose et al.
41   1991). Animals were given a single intraperitoneal injection of 0.5 g BHPN/kg body
42   weight at 6 weeks of age and exposed to 0.04, 0.4, or 4.0 ppm NC>2 for 17 months. The
43   incidence of pulmonary tumors in rats exposed to BHPN plus 4 ppm NC>2 was 12.5%
44   (n.s.) with adenomas found in 4/40 rats (10%) and adenocarcinomas found in 1/40 rats
45   (2.5%). One adenoma was found in the control group (2.5%) and one in the 0.04 ppm
46   group, but none in the 0.4 ppm group. In addition, marked bronchiolar mucosal
                                            35

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   hyperplasia was found in 17/40 rats (42.5%, p < 0.001) in the BHPN plus 4.0 ppm NO2
 2   group.
 3
 4   3.6. Summary
 5
 6          Five- to 60-minute LCso values for NO2 in the rat ranged from 416 to 115 ppm,
 7   respectively in one study (Carson et al. 1962) and from 833 to 168 ppm in another study
 8   (Gray et al. 1954).  The 15-minute LC50 for rabbits was 315 ppm (Carson et al. 1962). In
 9   a study using varying concentration and duration of exposure, the first mortalities were
10   observed in dogs at 75 ppm for 4 hours,  in rabbits at 75 ppm for 1 hour, in guinea pigs at
11   50 ppm for 1 hour, and in rats and mice at 50 ppm for 24 hours (Hine et al. 1970).
12   Histological alterations of the lungs following death included bronchiolitis, desquamated
13   bronchial epithelium, infiltration by polymorphonuclear cells, and edema. Enhanced
14   susceptibility to infection was shown in monkeys following exposure to 50 ppm for 2
15   hours (Henry et al. 1969) and in mice following exposure to 2 or 3.5 ppm for 3 hours
16   (Ehrlich 1978).
17
18          Pulmonary  edema and histological alterations induced by exposure to NO2 have
19   been characterized in dogs, sheep, guinea pigs, hamsters, rats, and mice.  Numerous
20   studies in rats have confirmed alveolar and interstitial edema, bronchiolitis, bronchiolar
21   epithelial cell hyperplasia, loss of cilia, necrosis of type I cells, and/or type II cell
22   hyperplasia 1-3 days after exposure to 26 ppm NO2 for 24 hours (Schnizlein et al. 1980;
23   Hillam et al. 1983) or to 20 ppm for 20 (Hayashi et al. 1987) or 24 hours (Rombout et
24   al. 1986).
25
26          Neonates appear less  sensitive to NO2 than adult animals with progressive
27   increases in lung injury and deaths seen in older rats and guinea pigs (Stephens et al.
28   1978, Azoulay-Dupuis et al.  1983).
29
30          Only one study was found in which rats were exposed to N2O4 and pulmonary
31   lesions were similar to those  described following NO2 exposure. No studies with
32   exposure to N2Os were found.
33
34   4. SPECIAL CONSIDERATIONS
35   4.1. Metabolism and Disposition
36
37          Total respiratory tract absorption of NO2 by humans exposed to 0.29-7.2 ppm for
38   <30 minutes during quiet respiration and during exercise has been measured at 81-90%
39   and 91-92%, respectively in healthy adults and 72% and 87%, respectively in asthmatics
40   (U.S. EPA 1993).  In monkeys exposed to 0.30-0.91 ppm NO2 for <10 minutes, 50-60%
41   of the inspired gas  has been shown to be retained during quiet respiration with
42   distribution throughout the lungs (Goldstein et al. 1977).  While the isolated rat lung,
43   ventilated with 5 ppm for 90  minutes, retained 36% of the NO2 (Postlethwait and Mustafa
44   1981), the majority of labeled NO2 (exposure parameters not specified) was retained by
45   the upper respiratory tract of the rat (Russell et al. 1991).
46
                                            36

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1          Pulmonary absorption of NO2 has been studied using in vivo and in vitro models.
 2   Uptake appears to be governed by the reaction between inhaled NO2 and constituents of
 3   the pulmonary surface lining layer to form nitrite (Postlethwait and Bidani 1990, 1994,
 4   Saul and Archer 1983). NO2 uptake is saturable with absorption proportional to inspired
 5   dose (Postlethwait and Bidani 1994) and has been shown to increase as temperature
 6   increases to a maximum of 10.6 jig NO2/min in an isolated lung model (Postlethwait and
 7   Bidani 1990).  The predominant reaction in the lungs involves hydrogen abstraction by
 8   readily oxidizable tissue components such as proteins and lipids to form nitrous acid and
 9   the nitrite radical (Postlethwait and Bidani 1994) and reaction with water to form nitrous
10   and nitric acids (Goldstein et al. 1977).
11
12          Distribution of inhaled NO2 or its metabolites is via the blood stream
13   (Goldstein et al. 1977). Nitrite formed in the lungs is oxidized to nitrate by
14   interactions with RBCs after diffusion into the vascular space (Postlethwait and
15   Mustafa  1981). Exposure of mice to 40 ppmNO2 produced slight (0.2%)
16   nitrosylhemoglobin but no methemoglobin (Oda et al. 1980) and an increase in both
17   nitrite and nitrate that reached equilibrium in 10 and 30 minutes, respectively (Oda et
18   al.  1981). After cessation of exposure, nitrite had a half-life of several minutes and
19   nitrate had a half-life of about 1 hour (Oda et al. 1981). Urinary excretion of nitrate
20   has been shown to be linearly related to the NO2 concentration administered via
21   inhalation (Saul and Archer 1983).
22
23   4.2. Mechanism of Toxicity
24
25          NO2 is an irritant to the mucous membranes and may cause coughing and dyspnea
26   during exposure.  After less severe exposure, symptoms may persist for several hours
27   before subsiding (NIOSH 1976). With more severe exposure, pulmonary edema ensues
28   with signs of chest pain, cough, dyspnea, cyanosis, and moist rales heard on auscultation
29   (NIOSH 1976, Douglas et al. 1989). Death from NO2 inhalation is caused by
30   bronchospasm and pulmonary edema in association with hypoxemia and respiratory
31   acidosis, metabolic acidosis, shift of the oxyhemoglobin  dissociation curve to the left,
32   and arterial hypotension (Douglas et al. 1989). A characteristic of NO2 intoxication after
33   the acute phase is a period of apparent recovery followed by late-onset bronchiolar injury
34   that manifests as bronchiolitis fibrosa obliterans (NIOSH 1976, NRC 1977, Hamilton
35   1983, Douglas et al. 1989).
36
37          Toxicity from acute exposure can be described in one of three categories: 1)
38   immediate death after very heavy exposure, 2) delayed symptoms with development of
39   edema within 48 hours, and 3) apparent recovery from immediate effects but later chronic
40   chest disease of varying severity (NRC 1977, Hamilton 1983).  Morphological and
41   biochemical changes in the lungs during these phases were studied in mice exposed to
42   140 ppm for 1  hour (Siegel et al. 1989). Immediately after exposure, cell death was
43   noted in  areas adjacent to the distal terminal bronchioles, and protease inhibitor activity,
44   lung protein content, and lung wet weight were significantly elevated. Two days after
45   exposure, the histological damage was exacerbated with  complete obliteration of the
46   alvoelar  structure, progressive edema and congestion of the lungs, hypertrophy and
                                             37

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   hyperplasia of the epithelial cells, increased numbers of intraalveolar macrophages and
 2   neutrophils. Also two days after exposure, there were dose-related increases in P-
 3   glucuronidase, lactate dehydrogenase, and choline kinase activity as well as increased
 4   protease inhibitor activity, pulmonary protein, and lung wet weight.  Pulmonary injury is
 5   characterized by loss of ciliated cells, disruption of tight capillary junctions, degeneration
 6   of type I cells, and proliferation of type II cells (Siegel et al. 1989, Elsayed 1994).
 7
 8          The predominant reaction in the lungs involves hydrogen abstraction by readily
 9   oxidizable tissue components such as proteins and lipids to form nitrous acid and the
10   nitrite radical (Postlethwait and Bidani 1994 U.S. EPA 1995) and reaction with water to
11   form nitrous and nitric acids (Greenbaum et al. 1967, Goldstein et al. 1977). This
12   reaction can lead to one mechanism by which NO2 causes pulmonary injury, lipid
13   peroxidation. NO2 is a free radical  that can attack unsaturated fatty acids in the cell
14   membrane forming carbon and oxygen centered radicals in a chain reaction (Ainslie
15   1993, Elsayed 1994, U.S. EPA 1995). This hypothesis is supported by studies on the
16   effects of antioxidants on NO2 exposure in humans and animals. Four-week
17   supplementation with vitamins C and E before exposure to 4 ppm for 3 hours resulted in
18   a marked  decrease in the amount of conjugated dienes and attenuated the decrease in
19   elastase activity inhibitory capacity in the alveolar lining fluid of healthy, human
20   volunteers (Mohsenin 1991).  Guinea pigs maintained on an ascorbic acid-deficient diet
21   had increased lung lavage fluid protein following exposure to 4.8 ppm NO2 for 3 hours
22   and increased wet lung weight, increased nonprotein sulfhydryl and ascorbic acid content
23   of the lungs, and decreased a-tocopherol content of the lungs following exposure to 4.5
24   ppm for 16 hours.  These changes were not seen in animals maintained on normal guinea
25   pig diets (Hatch et al. 1986). Rats exposed to 30 and 40 ppm for 4 hours had elevations
26   of lactate  dehydrogenase (LDH), malate dehydrogenase (MDH), and glucose-6-
27   phosphate dehydrogenase (GDH) activity in lavage fluid which were significantly
28   attenuated in animals maintained on diets providing 1000 mg/kg of a-tocopherol (Guth
29   and Mavis 1986). Another study found changes in fatty acid composition of alveolar
30   lavage phospholipids following exposure of rats to 10 ppm NO2 for 12 hours (Kobayashi
31   etal. 1984).
32
33   4.3. Oxides of Nitrogen
34
35          NO2 exists as an equilibrium mixture  of NO2 and N2C>4 but the dimer is not
36   important at ambient concentrations (U.S. EPA 1993).  The two compounds are phase-
37   related forms with N2C>4 favored in the liquid phase and NO2 favored in the gaseous
38   phase. An equilibrium distribution is reached which favors the lowest energy state in
39   the phase.  As a result when N2C>4 is released  it vaporizes and dissociates into NO2,
40   making it nearly impossible to generate a significant concentration of N2C>4 at
41   atmospheric pressure and ambient temperature, without generating a vastly higher
42   concentration of NO2. A rate for this reaction was not found, and because of this effect,
43   almost no inhalation toxicity data are available on N2C>4.  No information was found on
44   the interactions of N2Os.
45
46          Another oxide of nitrogen, nitric oxide (NO), is unstable in air and undergoes
                                             38

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   spontaneous oxidation to NC>2.  If the exposure concentration of NO is not high enough
 2   to be lethal due to methemoglobin formation, the victim can recover completely.  On
 3   the other hand, concentrations of NO2 that are not rapidly lethal may cause persistent
 4   effects and in some cases cause death from pulmonary edema after a delay of several
 5   days (NIOSH 1976).  In photochemical smog, NO2 absorbs sunlight of wavelengths
 6   between 290-430 nm and decomposes to NO and O (U.S. EPA 1993). If NO is of
 7   concern, reference should be made to the AEGL technical support document for nitric
 8   oxide.
 9
10   4.4. Other Relevant Information
11   4.4.1. Species Variability
12
13          Several studies indicate that there is a size-dependent species sensitivity to NO2
14   with larger animals apparently less sensitive than smaller animals such as rodents. Dogs
15   showed only mild signs of irritation at concentrations that caused pulmonary edema in
16   rats (Carson et al.  1962). Dogs  also survived exposures to 1000 ppm for 136 minutes and
17   5000 ppm for up to 22 minutes (Greenbaum et al. 1967) and sheep survived exposure to
18   500 ppm for 15-20 minutes (Januszkiewicz and Mayorga 1994). In contrast, 15-minute
19   and 1-hour LCso values in the rat ranged from 201-420 ppm and 115-168 ppm,
20   respectively (Gray et al. 1954, Carson et al. 1962). Based on the data  available, humans
21   are not more sensitive than larger laboratory animals. For example, irritation was
22   reported for humans exposed to 30 ppm for 2 hours (Henschler et al. 1960), in dogs
23   exposed to 20 ppm for 24 hours (Hine et al. 1970), and in monkeys  exposed to 35 ppm
24   for 2 hours (Henry et al. 1969).
25
26          Elsayed et al. (2002) examined species variability through dosimetry; the
27   calculated total inspired dose from  experimental measurements in rats and sheep was
28   compared to the theoretical dose of an average human.  Whether normalized for body
29   weight, lung volume, or alveolar surface area, the total effective dose was rats » sheep >
30   humans. Taking physiologic and anatomical factors into consideration, rats had a much
31   higher effective dose than the larger animals.  The authors concluded that NO2 toxicity is
32   associated with inhaled-dose distribution per unit lung volume or lung surface rather than
33   per unit body mass (Elsayed et al. 2002).
34
35   4.4.2. Susceptible Populations
36
37          For chronic, low-level exposures, U.S. EPA (1995) has identified two
38   populations as potentially at risk from NO2 exposure: children ages  5-12 and persons
39   with pre-existing respiratory disease.  Conclusions drawn from epidemiology studies
40   were that children ages 5-12 years old had an increased risk of about 20% for
41   developing respiratory symptoms and disease with each increase of 0.015 ppm in
42   estimated 2-week  average NO2 exposure (mean weekly concentrations in bedrooms
43   0.008-0.065 ppm) and that no evidence for increased risk was found for infants <2 years
44   old. This conclusion is supported somewhat by animal data in which adult animals were
45   more sensitive than neonates to the effects of NO2  (Azoulay-Dupuis et al. 1983,
46   Stephens et al. 1978). Reduced ventilatory reserves may prevent individuals with
                                            39

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   respiratory disease from resuming normal activity following exposure to NO2 (U.S. EPA
 2   1995). However, it is not certain whether these populations are also at particular risk
 3   from acute exposure scenarios.
 4
 5          Taken together, the data summarized in section 2.2.3 indicate that some
 6   asthmatics exposed to 0.3-0.5 ppm NO2 may respond with either subjective symptoms or
 7   slight changes in pulmonary function of no clinical significance. At approximately these
 8   same concentrations of NO2, subsequent exposure of asthmatics to an agent that causes
 9   non-specific airway responsiveness resulted in slight hyperreactivity, but the response is
10   not more severe than to NO2 alone (e.g. while some asthmatics respond to a bronchial
11   challenge and to NO2, the response to the challenge is not additively increased from prior
12   exposure to NO2). In contrast, some asthmatics did not respond to NO2 with changes in
13   pulmonary function or symptoms at concentrations of 0.5-4 ppm. The responses of
14   healthy individuals to NO2 exposures are also variable, with some, but not all, having
15   slight changes in pulmonary function following exposure to 5 ppm. All reported
16   responses in both asthmatic and healthy subjects at the concentrations discussed were
17   slight and of questionable biological or clinical significance.
18
19          Conclusions regarding differences in susceptibility between healthy and asthmatic
20   individuals are difficult to draw from the available data because of the high variability in
21   responses among  both groups.  There is only one study which has measured the responses
22   of both healthy and asthmatic individuals with the same study protocol (Linn and
23   Hackney 1983). Dose-response patterns are not discernable at these low concentrations
24   and clear thresholds are not apparent. Some individuals reported clinical symptoms in
25   the absence of changes in pulmonary function, while other individuals had measurable
26   changes in pulmonary function tests but no symptoms. One proposed explanation for the
27   variability in the responses of asthmatics to inhaled NO2 is the existence of a subgroup of
28   "responders."  From one laboratory, several asthmatics were identified as equally
29   responsive to 0.3  ppm in more than one study (Bauer et al. 1985, 1986).  However, the
30   investigators could find no common identifiers for these "responders" such as degree of
31   baseline obstruction or their inherent airway reactivity to carbachol or cold air (Utell
32   1989). Although  some individuals had a measurable response at lower concentrations,
33   the magnitude of the reported changes was not biologically or clinically significant in
34   either asthmatics or healthy individuals.
35
36   4.4.3. Concentration-Response Relationship
37
38          As discussed below for AEGL-2 and -3 levels, extrapolations were made to each
39   of the time points using Cn x t = k where n = 3.5 (ten Berge et al. 1986).  The value of n
40   was calculated by ten Berge et al. based on the data of Hine et al. (1970). The large value
41   of n indicates that concentration is more important than duration for the effects of
42   exposure to NO2.  Support for this supposition also comes from Gardner et al. (1979)
43   who showed that  short-term exposure to high concentrations resulted in greater effects (as
44   measured by mortality in an infectivity model in mice) than exposure to lower
45   concentrations administered over a longer duration.
46
                                             40

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   4.4.4. Susceptibility to infection
 2
 3          To determine the effects of NC>2 on resistance to infection, squirrel monkeys were
 4   challenged with Klebsiellapneumoniae within 24 hour after exposure. No deaths
 5   occurred from exposure to NC>2 alone, however, 3/3 monkeys died within 72 hours after
 6   50 ppm NC>2 exposure for 2 hours followed by K. pneumoniae challenge; massive
 7   infection was present in the lungs and other organs. Exposure of monkeys to 10 ppm for
 8   2 hours followed by K. pneumoniae challenge 3-5 days later did not result in death of the
 9   animals, but bacteria were still present in lung tissue at necropsy up to 46 days after
10   challenge indicating reduced clearence (Henry et al.  1969).
11
12          Numerous studies have reported enhanced susceptibility of mice to infectious
13   agents following exposure to NC>2. Most of these studies have been reviewed by U.S.
14   EPA (1993) and only a few are described here. Gardner et al. (1977) demonstrated that
15   the concentration-time relationship was linear for 20% mortality using an infectivity
16   model in mice challenged with Streptococcuspyogenes; NC>2 exposures ranged from
17   0.5-28 ppm for 6 minutes to 12 months. Similarly, mortality was increased in mice
18   challenged with S. pyogenes in response to short-term exposure to a high concentration
19   of NC>2 compared to a lower concentration administered over a longer duration when
20   the concentration x time product was held constant.  A single 3-hour exposure to 2.0 or
21   3.5 ppm NC>2 enhanced the susceptibility of three strains of mice to streptococcal
22   pneumonia and influenza infection as seen by excess mortality and reduced survival
23   time (Ehrlich 1978). Mice exercised in a motorized wheel during exposure to 3 ppm
24   for 3 hours followed by challenge with S. pyogenes had significantly increased
25   mortality compared to non-exercised animals (Tiling et al.  1980). Pulmonary bacterial
26   defenses against Staphylococcus aureus were suppressed following exposure of Swiss
27   mice to concentrations of >4 ppm for 4 hours (Jakab 1987).  Significantly decreased
28   pulmonary bactericidal activity was shown in Swiss mice infected with S. aureus then
29   exposed to 7, 9.2, or 14.8 ppm NC>2 for 4 hours, or exposed to 2.3  or 6.6 ppm for 17
30   hours prior to infection.  Histologically the lungs of mice exposed to >9.2 ppm for 4
31   hours showed vascular hyperemia while those from mice exposed to >2.3 ppm for 17
32   hours had minor vascular hyperemia and interstitial edema (Goldstein et al. 1973).
33   Enhanced susceptibility to infection was observed in CD-I mice exposed to 5 ppm for 6
34   hours/day on two consecutive days prior to inoculation with murine cytomegalovirus,
35   followed by exposure to 5 ppm NC>2 for 6 hours/day for four consecutive days; there
36   was no histological evidence of lung injury  (Rose et al. 1989). Continuous exposure of
37   mice to 20 ppm NC>2 for 4 days resulted in impairment of acquired resistance
38   (decreased EDso) of C57B1/6 mice immunized prior to Klebsiella pneumoniae
39   challenge (Bouley etal. 1986).
40
41          Alterations in host defense mechanisms have been demonstrated in rabbits. Male
42   and female New Zealand rabbits exposed for 3 hours to varying concentrations of NC>2
43   had an increase in polymorphonuclear leukocytes obtained by pulmonary lavage at >8
44   ppm with the peak infiltration 6-9 hours after the end of exposure (Gardner et al. 1969).
45   In other experiments, these authors demonstrated that the response persisted up to 72
46   hours post exposure and that phagocytic activity was inhibited.
                                            41

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2          Mice have also been used extensively as a model for immune function
 3   alterations following NC>2 exposure.  Decreases in splenic and thymic weights,
 4   cellularity, plaque-forming cell (PFC) responses, and hemagglutinins (HA), along with
 5   decreased body weight, were observed in C56B1/6 mice exposed to 20 ppm NC>2 for 48
 6   hours (Azoulay-Dupuis et al. 1985).  Significant suppression of primary antibody
 7   responses (HA and PFC) were also seen following exposure of BALB/c mice to 20 or
 8   40 ppm for 12 hours (Hidekazu and Fujio 1981).  Phytohemagglutinin and bacterial
 9   lipopolysaccharide responses were depressed in mice exposed continuously to 0.5 ppm
10   or to 0.1 ppm with daily 3-h peaks (5 days per week) of either 0.25 ppm, O.Sppm, or 1.0
11   ppm (Maigetter et al. 1978).  Other effects of NC>2 on cellular and humoral immunity
12   have been reviewed by U.S. EPA (1993) but are not relevant to  derivation of AEGL
13   values.
14
15   5. DATA ANALYSIS FOR AEGL-1
16
17          AEGL-1 is the airborne concentration (expressed as parts per million or
18   milligrams per cubic meter [ppm or mg/m3]) of a substance above which it is predicted
19   that the general population, including susceptible individuals, could experience  notable
20   discomfort, irritation, or certain asymptomatic, non-sensory effects. The effects are not
21   disabling and are transient and reversible upon cessation of exposure.
22
23   5.1. Summary of Human Data Relevant to AEGL-1
24
25          The evidence indicates that some asthmatics exposed to  0.3-0.5 ppm NC>2 may
26   respond with either subjective symptoms or slight changes in pulmonary function of no
27   clinical significance. At approximately these same  concentrations of NC>2 some
28   asthmatics may show slight hyperreactivity to a bronchial challenge, but the response is
29   no more severe than the response to NC>2 alone (e.g. while some asthmatics respond to a
30   bronchial challenge and to NC>2, the response to the challenge is not additively increased
31   from prior exposure to NC>2). In contrast, some asthmatics did not respond to NC>2 at
32   concentrations of 0.5-4 ppm. The responses of healthy individuals to NC>2 exposures are
33   also variable, with some, but not all, responding to 5 ppm.
34
35          Kerr et al. (1978, 1979) reported that 7/13 asthmatics experienced slight burning
36   of the eyes, slight headache, chest tightness, or labored breathing with exercise  during
37   exposure to 0.5 ppm for 2 hours; at this concentration the odor of NC>2 was perceptible
38   but the subjects became unaware of it after about 15 minutes.  No changes in  any
39   pulmonary function tests were found immediately following the chamber exposure (Kerr
40   et al. 1978, 1979).  Significant group mean reductions in FEVi (-17.3% vs -10.0%) and
41   specific airway conductance (-13.5% vs -8.5%) occurred in asthmatics after exercise
42   during exposure to 0.3 ppm for 4 hours and 1/6 individuals experienced chest tightness
43   and wheezing (Bauer et al. 1985). The onset of effects was delayed when exposures were
44   by oral-nasal inhalation compared with oral inhalation. This delay may result from
45   scrubbing within the upper airway. In a similar study, asthmatics  exposed to  0.3 ppm for
46   30 minutes at rest followed by 10 minutes of exercise had significantly greater reductions
                                            42

-------
     NITROGEN OXIDES
                                    NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
in FEVi (10% vs 4% with air) and partial expiratory flow rates at 60% of total lung
capacity, but no symptoms were reported (Bauer et al. 1986). In a preliminary study with
13 asthmatics exposed to 0.3 ppm for 110 minutes, slight cough and dry mouth and throat
and significantly greater reduction (11% vs 7%) in FEVi occurred after exercise,
however, in a larger study, no changes in pulmonary function were measured and no
symptoms were reported following exposure of 21 asthmatics to concentrations up to 0.6
ppm for 75 minutes (Roger et al. 1990).  The mean drop in FEVi for asthmatics during a
3-hour exposure to 1 ppm NO2 (2.5%) with intermittent exercise was significantly greater
than the drop  during air (1.3%) exposure with intermittent exercise; in BALF, levels of 6-
keto-prostaglandinia were decreased and levels of thromboxane B2 and prostaglandin D2
were increased after NO2 exposure (Torres et al. 1995).

5.2. Summary of Animal Data Relevant to AEGL-1

       Animal data relevant to derivation of AEGL-1 are limited.  Slight irritation was
noted in squirrel monkeys exposed to 10 and 15 ppm for 2 hours (Henry  et al. 1969)
and mild sensory effects occurred in dogs exposed to 125 ppm for 5 minutes, 52 ppm
for 15 minutes, or 39 ppm for 60 minutes (Carson et al. 1962).

5.3. Derivation of AEGL-1

       The study by Kerr et al. (1978, 1979) was considered the most appropriate to use
as the basis for AEGL-1 values. Exposure of asthmatics to 0.5 ppm NO2 for 2 hours
resulted in clinical signs but no changes  in pulmonary function.  Since asthmatics are
potentially the most susceptible population, no uncertainty factor was applied. Therefore,
a concentration of 0.94 mg/m3 (0.50 ppm NO2 or 0.25 ppm N2C>4) was adopted for all
time points (Table 5) because adaptation to mild sensory irritation occurs. In addition,
animal responses to NO2 exposure have demonstrated a much greater dependence upon
concentration than upon time; therefore,  extending the 2-hour concentration to 8 hours
should not exacerbate the human response.
TABLE 5: AEGL-1 Values for Nitrogen Dioxide and Nitrogen Tetroxide
Chemical
N02
N204
10-minute
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
30-minute
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
1-hour
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
4-hour
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
8 -hour
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
32
33
34
35
36
37
38
39
6. DATA ANALYSIS FOR AEGL-2

       AEGL-2 is the airborne concentration (expressed as ppm or mg/m3) of a
substance above which it is predicted that the general population, including susceptible
individuals, could experience irreversible or other serious, long-lasting adverse health
effects or an impaired ability to escape.
                                            43

-------
     NITROGEN OXIDES                NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2   6.1. Summary of Human Data Relevant to AEGL-2
 3
 4          Human data relevant to AEGL-2 are limited but consistent. Henschler et al.
 5   (1960) performed several experiments on healthy, male volunteers and found that
 6   exposure to 30 ppm for 2 hours caused definite discomfort. Three individuals exposed to
 7   30 ppm for 2 hours perceived an intense odor upon entering the chamber, the odor
 8   quickly diminished and was completely absent after 25-40 minutes.  One individual
 9   experienced a slight tickling of the nose and throat mucous membranes after 30 minutes,
10   the two others after 40 minutes. From 70 minutes on, all subjects experienced a burning
11   sensation and an increasingly severe cough for the next 10-20 minutes, but coughing
12   decreased from 100 minutes on.  However, the burning sensation continued and moved
13   into the lower sections of the airways and was finally felt deep in the chest. At this time,
14   marked sputum secretion and dyspnea were noted. Toward the end of the exposure, the
15   subjects condition was described as bothersome and barely tolerable. A sensation of
16   pressure and increased sputum secretion continued for several hours after cessation of
17   exposure (Henschler et al. 1960).  In a similar experiment (Henschler and Liitke 1963)
18   groups of 4 or 8 healthy male volunteers were exposed to 10 ppm for 6 hours or to 20
19   ppm for 2 hours. All subjects, upon entering the chamber, noted the odor which
20   diminished rapidly. At 20 ppm minor scratchiness of the throat was felt after about 50
21   minutes and 3/8 experienced slight headaches towards the end of the exposure period.
22
23   6.2. Summary of Animal Data Relevant to AEGL-2
24
25          Several animal studies are relevant to AEGL-2 derivation. Hine et al. (1970)
26   noted lacrimation, reddening of the conjunctivae, and increased respiration in 5  species
27   exposed to >40 ppm for varying durations. Lethality did not occur until concentrations
28   and durations reached 75  ppm for 4 hours in the dog and 1 hour in the rabbit, 50 ppm for
29   1 hour in the guinea pig, and 50 ppm for 24 hours in the rat and mouse.  At 20 ppm for 24
30   hours, all species showed minimal signs of irritation and changes in behavior with
31   histopathological lesions described as questionable evidence of lung congestion and
32   interstitial inflammation.
33
34          Exposure of monkeys to 35 ppm for two hours resulted in irritation as measured
35   by changes in lung function and microscopic lesions in the lung (Henry et al. 1969).
36   The histological lesions in the lung were characterized by Siegel et al. (1989) following
37   exposure of mice to 140 ppm for 1 hour.  Carson  et al. (1962) conducted a series of
38   experiments in dogs and rats. Mild irritation and  some respiratory effects, but no gross
39   or microscopic lesions, were noted in dogs exposed to 53 or 39 ppm for 1 hour while
40   rats exposed to 72 ppm for 1 hour showed signs of severe respiratory distress and eye
41   irritation as well as gross lesions in the lung and evidence of infection.
42
43          Developmental delays and disturbances in neuromotor development were reported
44   for rat pups following maternal exposure (Tabacova et al. 1985). However, these effects
45   were reported to have occurred at levels near ambient concentrations and are well below
46   those of most other studies in both humans and animals.
                                            44

-------
     NITROGEN OXIDES
                                     NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
14
15
16
17
18
19
6.3. Derivation of AEGL-2

       Based on both human and animal data, it appears that a concentration of >30 ppm
NC>2 is required before marked irritation, discomfort, and respiratory effects occur.
Therefore, a concentration of 30 ppm for a 2-hour exposure of humans (Henschler et al.
1960) was used to derive AEGL-2 values.  The point of departure is considered a
threshold for AEGL-2 since the effects noted by the subjects would not impair the ability
to escape and the effects were reversible after cessation of exposure. Values scaled for
the derivation of the 10- and 30-minute and 1-, 4-, and 8-hour AEGL-2 endpoints were
calculated from Cn x t = k using n = 3.5 (ten Berge  et al. 1986).  The value of n was
calculated by ten Berge et al. from the data of all species together from Hine et al. (1970).
An intraspecies uncertainty factor of 3 was applied  to account for sensitive
subpopulations because the mechanism of action of a direct acting irritant is not expected
to differ greatly among individuals.  With additional uncertainty factors the values would
be inconsistent with some of the experimental data  for asthmatics, such as the no-
adverse-effect concentration of 4 ppm in the study by Linn and Hackney (1984). AEGL-
2 values are presented in Table 6.
TABLE 6: AEGL-2 Values for Nitrogen Dioxide and Nitrogen Tetroxide
Chemical
N02
N204
10-minute
38 mg/m3
(20 ppm)
38 mg/m3
(10 ppm)
30-minute
28 mg/m3
(15 ppm)
28 mg/m3
(7.6 ppm)
1-hour
23 mg/m3
(12 ppm)
23 mg/m3
(6.2 ppm)
4-hour
15 mg/m3
(8.2 ppm)
15 mg/m
(4.1 ppm)
8 -hour
13 mg/m3
(6.7 ppm)
13 mg/m
(3. 5 ppm)
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
       These levels are not expected to cause severe effects as coal miners were exposed
to peakNO2 concentrations of 14 ppm without adverse consequences (Robertson et al.
1984) and it can be assumed that the peak levels were not sustained longer than a few
minutes.  Similar AEGL-2 values are derived using the exposure of 140 ppm for 1 hour
in the mouse (Siegel et al. 1989) and an uncertainty factor of 10 or the exposure of 35
ppm for 2 hours in the monkey (Henry et al.  1969) and an uncertainty factor of 3. If the
animal data from either Hine et al. (1970) or Carson et al. (1962) are used for the basis  of
derivation, the AEGL-2 values are even more conservative  than those derived with the
use of human data.

7. DATA ANALYSIS FOR AEGL-3

       AEGL-3 is the airborne concentration (expressed as ppm or mg/m3) of a
substance above which it is predicted that the general population, including susceptible
individuals, could experience life-threatening health effects or death.

7.1. Summary of Human Data Relevant to AEGL-3

       A welder was hospitalized with pulmonary edema after exposure to
                                            45

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1   approximately 90 ppm for 30-40 minutes (Norwood et al. 1966). It is possible that
 2   without medical intervention, the exposure could have been fatal.
 3
 4          Concentrations of NO2 above 150 ppm are probably fatal to humans due to
 5   bronchospasm and pulmonary edema (NRC 1977, Douglas et al. 1989). A human 1-hour
 6   LCso of 174 ppm was estimated from data in 5 animal species (Book 1982), however,
 7   these are not considered valid experimental data on which to base AEGL-3. No other
 8   human data were relevant to derivation of AEGL-3.
 9
10   7.2. Summary of Animal Data Relevant to AEGL-3
11
12          Squirrel monkeys (n = 2-6/group) were exposed to 10-50 ppm NO2 for 2 hours
13   with respiratory function monitored during exposure (Henry et al. 1969). NO2 exposure
14   alone resulted in a markedly increased respiratory rate and decreased tidal volume
15   during exposures to 50 or 35 ppm, but caused only slight effects at 15 and 10 ppm.  Mild
16   histopathological changes in the lungs were noted after exposure to 10 and 15 ppm,
17   however, marked changes in lung structure were observed after exposure to 35 and 50
18   ppm.  At 35 ppm, areas of the lung were collapsed with basophilic alveolar septa, in
19   other areas the alveoli were expanded with septal wall thinning, and the bronchi were
20   moderately inflamed with some proliferation of the surface epithelium.  At 50 ppm,
21   extreme vesicular dilatation or total collapse of alveoli was observed,  lymphocyte
22   infiltration was seen with extensive edema, and surface erosion of the epithelium of the
23   bronchi was observed. In addition to the effects on the lungs, interstitial fibrosis (35
24   ppm) and edema (50 ppm)  of cardiac tissue, glomerular tuft swelling in the kidney (35
25   and 50 ppm), lymphocyte infiltration in the kidney and liver (50 ppm), and congestion
26   and centrilobular necrosis in the liver (50 ppm) were observed.
27
28          Rats exposed to 72  ppm for 60 minutes (approximately 50% of the LD50) showed
29   signs of severe respiratory  distress and eye irritation lasting about 2 days; lung-to-body
30   weight ratios were significantly increased during the first 48 hours after exposure (Carson
31   etal. 1962).
32
33          Lethality in 5 animal species first occurred at exposure concentrations and
34   durations of 75 ppm for 4 hours in the dog and 1 hour in the rabbit, 50 ppm for 1 hour
35   in the guinea pig, and 50 ppm for 24 hours in the rat and mouse (Hine et al. 1970).  In
36   general, the larger animals, including humans, are less susceptible to toxicity from NO2
37   inhalation than are the rodents.
38
39   7.3. Derivation of AEGL-3
40
41          The data from the monkey are considered the best available for derivation of
42   AEGL-3 values. Signs of marked irritation and severe lung histopathology were observed
43   from exposure to 50 ppm for 2 hours.  This exposure scenario was extrapolated to the 10-
44   and 30-minute and 1-, 4-, and 8-hour time points using the equation Cn x t = k where n =
45   3.5 (ten Berge  et al. 1986). The value of n was calculated by ten Berge et al. from the
46   data of all species together from Hine et al. (1970).  A total uncertainty factor of 3 was
                                            46

-------
     NITROGEN OXIDES
                                    NAC/Interim (NO2)/Proposed (N2O4): 12/2008
     applied which includes a 3 for intraspecies variability and a 1 for interspecies variability.
     Use of a greater intraspecies uncertainty factor was not considered necessary because the
     mechanism of action is not expected to differ greatly among individuals. Because the
     endpoint in the monkey study is below the definition of AEGL-3, human data support the
     point of departure and derived values, and due to the similarities of the respiratory tract
     between humans and monkeys, additional interspecies uncertainty factors are not
     considered necessary.  The mechanism of action of NO2 does not vary between species
     with the target at the alveoli.  AEGL-3 values for NO2 and N2O4 are listed in Table 7.
TABLE 7: AEGL-3 Values for Nitrogen Dioxide and Nitrogen Tetroxide
Chemical
NO2
N2O4
10-minute
64 mg/m
(34 ppm)
64 mg/m3
(17 ppm)
30-minute
47 mg/m3
(25 ppm)
47 mg/m3
(13 ppm)
1-hour
38 mg/m3
(20 ppm)
38 mg/m3
(10 ppm)
4-hour
26 mg/m3
(14 ppm)
26 mg/m3
(7.0 ppm)
8 -hour
21 mg/m3
(1 1 ppm)
21 mg/m3
(5. 7 ppm)
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
       The AEGL-3 values are supported by human data from the welder.  Pulmonary
edema, confirmed on X-ray, resulted from exposure to approximately 90 ppm for up to
40 minutes (Norwood et al. 1966). If this exposure scenario is used for derivation of
AEGL-3 values with an uncertainty factor of 3 the 10- and 30-minute and 1-, 4-, and 8-
hour values are 45, 33, 27, 18, and 15 ppm, respectively.  Similar results are obtained
using the exposure of rats to 72 ppm for 1 hour (Carson et al. 1962) and an  uncertainty
factor of 3. In addition, the AEGL-3 values are below the concentrations at which
lethality first occurred in five animal species (Hine et  al. 1970).
8. SUMMARY OF AEGLS
8.1. AEGL Values and Toxicity Endpoints

       The derived AEGL values for various levels of effect and durations of exposure
are summarized in Tables 8 and 9.  Values were derived based on data from human and
animal exposures to NO2 and are considered applicable to the other oxides of nitrogen.
Values for N2O4 in units of ppm have been calculated on a molar basis as presented
below.
TABLE 8: Summary of AEGL Values for Nitrogen Dioxide (mg/m3 [ppm])
AEGL Level
AEGL-1
AEGL-2
AEGL-3
10-minute
0.94 (0.50)
38 (20)
64 (34)
30-minute
0.94 (0.50)
28(15)
47 (25)
1-hour
0.94 (0.50)
23 (12)
38 (20)
4-hour
0.94 (0.50)
15 (8.2)
26 (14)
8 -hour
0.94 (0.50)
13 (6.7)
21(11)
30
31
                                            47

-------
     NITROGEN OXIDES
                                   NAC/Interim (NO2)/Proposed (N2O4): 12/2008
TABLE 9: Summary of AEGL Values for Nitrogen Tetroxide (mg/m3 [ppm])
AEGL Level
AEGL-1
AEGL-2
AEGL-3
10-minute
0.94 (0.25)
38(10)
64 (17)
30-minute
0.94 (0.25)
28 (7.6)
47(13)
1-hour
0.94 (0.25)
23 (6.2)
38(10)
4-hour
0.94 (0.25)
15(4.1)
26 (7.0)
8 -hour
0.94 (0.25)
13 (3.5)
21 (5.7)
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
8.2. Comparison with Other Standards and Criteria

       Standards and guidance levels for workplace and community exposures to NC>2
are listed in Table 10. No standards or guidance for exposure to N2O4 were found.  The
ACGffl recommends a TLV of 3 ppm for workers (ACGffl 2003) while the OSHA PEL
is a ceiling of 5 ppm (OSHA 1999). The NIOSH IDLH is 20 ppm (NIOSH 1996) which
is exactly between the 30-minute AEGL-2 and AEGL-3 values. The IDLH is reported as
based on acute inhalation data in humans, but no primary references were listed in the
documentation; NIOSH notes that the IDLH may be a conservative value due to the lack
of relevant acute toxicity data for workers exposed to concentrations above 20 ppm.
ERPGs (AIHA 2003), based on human and animal data, are similar to the 1-hour AEGL
values. The NRC's 1-hour EEGL is 1 ppm (NRC 1985) for workplace conditions. The
occupational exposure limits from ACGIH, Germany, The Netherlands, and Sweden are
2-5 ppm.

       In addition to the standards listed in Table 10, air quality standards have also
been developed for NO2. The National Ambient Air Quality Standard is 0.053 ppm
(U.S. EPA 1997) with Significant Harm Levels of 2 ppm  for a 1-hour average and 0.5
ppm for a 24-hour average (U.S.  EPA 1987a). The Level of Concern is 5 ppm (U.S.
EPA 1987b). The state of California has adopted 0.25 ppm as the standard for a 1-hour
exposure to protect sensitive individuals (Cal. EPA 1992).
                                          48

-------
      NITROGEN OXIDES
NAC/Interim (NO2)/Proposed (N2O4): 12/2008
TABLE 10: Extant Standards and Guidelines for Nitrogen Dioxide
Guideline
AEGL-1
AEGL-2
AEGL-3
ERPG-1 (AfflA)3
ERPG-2 (AfflA)
ERPG-3 (AfflA)
EEGL (NRC)b
IDLH (NIOSH)C
REL-STEL (NIOSH)d
PEL-STEL (OSHA)e
PEL-TWA (OSHA)f
TLV-TWA
(ACGIH)g
TLV-STEL
(ACGIH)h
MAK (Germany)1
MAK Peak Exposure
(Germany)"
MAC (The
Netherlands)1"
OEL-LLV (Sweden)1
OEL-CLV (Sweden)"1
Exposure Duration
10 minute
0.50 ppm
20ppm
34 ppm





30 minute
0.50 ppm
15 ppm
25 ppm




20 ppm
1 ppm
1 ppm




5 ppm


5 ppm




5 ppm
1 hour
0.50 ppm
12 ppm
20 ppm
1 ppm
15 ppm
30 ppm
1 ppm











4 hour
0.50 ppm
8.2 ppm
14 ppm



0.25 ppm











8 hour
0.50 ppm
6.7 ppm
11 ppm



0.12 ppm



5 ppm (C)
3 ppm

5 ppm

2.0 ppm
2 ppm

 1    a ERPG (Emergency Response Planning Guidelines, American Industrial Hygiene Association)
 2    (AIHA 2003)
 3        The ERPG-1 is the maximum airborne concentration below which it is believed nearly all individuals
 4        could be exposed for up to one hour without experiencing other than mild, transient adverse health
 5        effects or without perceiving a clearly defined objectionable odor.
 6        The ERPG-2 is the maximum airborne concentration below which it is believed nearly all individuals
 7        could be exposed for up to one hour without experiencing or developing irreversible or other serious
 8        health effects or symptoms that could impair an individual's ability to take protective action.
 9        The ERPG-3 is the maximum airborne concentration below which it is believed nearly all individuals
10        could be exposed for up to one hour without experiencing or developing life-threatening health effects.
11
12    bEEGL (Emergency Exposure Guidance Levels) National Research Council (NRC 1985) The EEGL is
13            the concentration of contaminants that can cause discomfort or other evidence of irritation or
14            intoxication in or around the workplace, but avoids death, other severe acute effects and long-term
15            or chronic injury.
16
17    CIDLH (Immediately Dangerous to Life and Health, National Institute of Occupational Safety and
18            Health) (NIOSH 1996) represents the maximum concentration from which one could escape
19            within 30 minutes without any escape-impairing symptoms, or any irreversible health effects.
20            The IDLH for nitrogen dioxide is based on acute inhalation toxicity data in humans.
                                                    49

-------
      NITROGEN OXIDES                 NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1
 2    dNIOSH REL-STEL (Recommended Exposure Limits - Short Term Exposure Limit) (NIOSH
 3            2003) is defined analogous to the ACGIH TLV-STEL.
 4
 5    eOSHA PEL-STEL (Permissible Exposure Limits - Short Term Exposure Limit) (NIOSH
 6            2003) is defined analogous to the ACGIH-TLV-STEL.
 7
 8    fOSHA PEL-TWA (Occupational Health and Safety Administration, Permissible Exposure Limits -
 9    Ceiling)
10            (OSHA 1999) is defined analogous to the ACGIH-TLV-TWA, but is for exposures of no
11            more than 10 hours/day, 40 hours/week. (C) denotes a ceiling.
12
13    gACGIH TLV-TWA (American Conference of Governmental Industrial Hygienists, Threshold
14            Limit Value - Time Weighted Average) (ACGIH 2003) is the time-weighted average
15            concentration for a normal 8-hour workday and a 40-hour workweek, to which nearly all workers
16            may be repeatedly exposed, day after day, without adverse effect.
17
18    hACGIH TLV-STEL (Threshold Limit Value - Short Term Exposure Limit) (ACGIH 2003) is
19            defined as a ISminute TWA exposure which should not be exceeded at any time during the
20            workday even if the 8-hour TWA is within the TLV-TWA.  Exposures above the TLV-TWA up
21            to the STEL should not be longer than 15 minutes and should not occur more than 4 times per
22            day. There should be at least 60 minutes between successive exposures in this range.
23
24    'MAK (Maximale Argeitsplatzkonzentration [Maximum Workplace Concentration]) (Deutsche
25            Forschungsgemeinschaft [German Research Association] 2002) is defined analogous to the
26            ACGIH-TLV-TWA.
27
28    JMAK Spitzenbegrenzung (Peak Limit [Category 1,1]) (German Research Association 2002) constitutes
29            the average concentration to which workers can be exposed for a period up to 15 minutes with no
30            more than 1 excursion per work shift and a minimum of 1 hour between excursions.
31
32    kMAC (Maximaal Aanvaaarde Concentratie [Maximal Accepted Concentration]) (SDU
33            Uitgevers [under the auspices of the Ministry of Social Affairs and Employment], The
34            Hague, The Netherlands 2000) is defined analogous to the ACGIH-TLV-TWA.
35
36    'OEL-LLV (Occupational Exposure Limits - Level Limit Value) (Swedish National Board of
37            Occupational Safety and Health 1996) is an occupational exposure limit value for exposure during
38            one working day.
39
40    mOEL-CLV (Occupational Exposure Limits - Ceiling Limit Value) (Swedish National Board of
41            Occupational Safety and Health 1996) is an occupational exposure limit value for exposure
42            during a reference period of fifteen minutes.
43
44    8.3. Data Adequacy and Research Needs
45
46           Data on the effects of NC>2 on asthmatics and individuals with respiratory disease
47    were inconsistent  and inconclusive. Additional studies that correlate severity of disease
48    with individual responses would be helpful.
49
                                                 50

-------
      NITROGEN OXIDES                 NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1    9. REFERENCES
 2
 3    Abe, M.  1967.  Effects of mixed NO2-SO2 gas on human pulmonary functions. Bull. Tokyo Med. Dent.
 4            Univ. 14:415-433.
 5
 6    Abraham, W.M., Welker, M., Oliver, W., Jr., Mingle, M., Januszkiewicz, A.J., Wanner, A., and Sackner,
 7            M.  1980. Cardiopulmonary effects of short-term nitrogen dioxide exposure in conscious sheep.
 8            Environ. Res. 22:61-72.
 9
10    ACGIH.  1991.  American Conference of Governmental Industrial Hygienists, Inc.  Nitrogen Dioxide. In:
11            Documentation of the Threshold Limit Values and Biological Exposure Indicies, 6th ed., ACGIH,
12            Cincinnati, OH, pp. 11081110.
13
14    ACGIH.  2003.  American Conference of Governmental Industrial Hygienists, Inc.  TLVs and BEIs
15            Based on the Documentation of the Threshold Limit Values for Chemical Substances and
16            Physical Agents and Biological Exposure Indicies. ACGIH, Cincinnati, OH, pp. 44.
17
18    AIHA (American Industrial Hygiene Association). 2003.  The AIHA 2003 Emergency Response Planning
19            Guidelines and Workplace Environmental Exposure Level Handbook.  Amer. Ind. Hyg. Assoc.,
20            Fairfax, Virginia.
21
22    Adams, W.C., Brookes, K.A., and Schelegle, E.S. 1987. Effects of NO2 alone and in combination with O3
23            on young men and women. J. Appl. Physiol. 62:1698-1704.
24
25    Ainslie, G. 1993. Inhalational injuries produced by smoke and nitrogen dioxide. Respir. Med. 87:169-174.
26
27    Azoulay-Dupuis, E., Torres, M., Soler, P., and Moreau, J. 1983. Pulmonary NO2 toxicity in neonate
28            and adult guinea pigs and rats. Environ. Res. 30:322-339.
29
30    Azoulay-Dupuis, E., Levacher, M., Muffat-Joly, M., Pocidalo,  J.J.  1985. Humoral immunodepression
31            following acute NO2 exposure in normal and adrenalectomized mice.  J.  Toxicol. Environ.
32            Health 15:149-162.
33
34    Bauer, M.A., Utell, M.J., Morrow, P.E., Speers, D.M., and Gibb, F.R.  1985. Route of inhalation
35            influences airway responses to 0.30 ppm nitrogen dioxide in asthmatic subjects. Am. Rev.
36            Respir. Dis. 131:A171.
37
38    Bauer, M.A., Utell, M.J.,  Morrow, P.E., Speers, D.M., and Gibb, F.R.   1986. Inhalation of 0.30 ppm
39            nitrogen dioxide potentiates exercise-induced bronchospasm in asthmatics. Am.  Rev. Respir.
40            Dis.  134:1203-1208.
41
42    Bauer, U., Berg, D., Kohn, M., and Meriwhether, R.  1996. Community exposure to nitrogen tetroxide.
43            Am. J. Epidemiol. 143:844.
44
45    Bauer, U., Berg, D., Kohn,  M.A., Meriwether, R.A., and Nickle, R.A.   1998. Acute  effects of
46            nitrogen dioxide after accidental release. Public Health Reports 113:62-70.
47
48    Blomberg, A., Krishna, M.T., Bocchino, V., Biscione, G.L., Shute, J.K., Kelly, F.J., Frew, A.J., Holgate,
49            S.T., and Sandstom,  T.  1997. The inflammatory effects of 2 ppm NO2 on the airways of healthy
50            subjects. Am. J. Respir. Crit. Care Med. 156:418-424.
51
52    Book,  S.A.  1982. Scaling toxicity from laboratory animals to people: an example with nitrogen
53            dioxide. J. Toxicol. Environ. Health 9:719-725.
54
                                                    51

-------
      NITROGEN OXIDES                   NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1    Bouley, G., Azoulay-Dupuis, E., and Gaudebout, C. 1986. Impaired acquired resistance of mice
 2            to Klebsiella pneumoniae infection induced by acute NOa exposure. Environ. Res.
 3            41:497-504.
 4
 5    Braun-Fahrlaender, Ch., Ackermann-Liebrich,  U.,  Schwartz,  I, Gnehm, H.P., Rutishauser, M.,  and
 6            Wanner, H.-U.  1992. Air pollution and respiratory symptoms in preschool children. Am. Rev.
 7            Respir. Dis. 145:42-47.
 8
 9    Brown, R.F.R., Clifford, W.E., Marrs, T.C., and Cox, R.A.  1983. The histopathology of rat lung
10            following short term exposures to mixed oxides of nitrogen (NOX). Br. J. Exp.  Path. 64:579-
11            593.
12
13    Budavari, S., O'Neil, M.J., Smith, A., Heckelman, P.E., and Kinneary, J.F. (Eds.)  1996.  The Merck
14            Index, 11th ed. Rahway, NJ:Merck and Co., Inc., p. 1135.
15
16    Burnett, R.T., Stieb, D., Brook, J.R., Cakmak, S., Dales, R., Raizenne, M., Vincent, R., Dann, T.
17            2004.   Associations between short-term changes  in  nitrogen dioxide  and mortality in
18            Canadian cities.  Arch. Environ. Health 59:228-236.
19
20    Cakmak, S., Dales, R.E., and Judek, S.  2006. Do gender, education, and income modify the effect
21            of air pollution gases on cardiac disease? J. Occup. Environ. Med. 48:89-94.
22
23    Cal. EPA.  1992.  California Environmental Protection Agency.  Review of the One-Hour Ambient Air
24            Quality Standard for Nitrogen Dioxide. State of California, Air Resources Board.
25
26    Carson, T.R., Rosenholtz, M.S., Wilinski, F.T., and Weeks, M.H.  1962. The responses of animals
27            inhaling  nitrogen dioxide for single,  short-term exposures. Am. Ind. Hyg. Assoc. J. 23:457-
28            462.
29
30    Case, B.W.,  Gordon, R.E., and Kleinerman, J.  1982.  Acute bronchiolar injury following nitrogen
31            dioxide exposure: a freeze fracture study. Environ. Res. 29:399-413.
32
33    CFR.  1997.  Code of Federal  Regulations. National primary  and secondary  ambient air quality
34            standards for nitrogen dioxide. 40 CFR (§50.11) p. 8.
35
36    Conrad, E., Lo, W., deBiosblanc, B.P., and Shellito, J.E.  1998.  Reactive airways dysfunction syndrome
37            after exposure to dinitrogen tetroxide. South. Med. J. 91:338-341.
38
39    Dales, R., Burnett, R.T., Smith-Doiron, M., Stieb, D.M., and Brook, J.R.  2004.  Air pollution and sudden
40            infant death syndrome. Pediatrics 113:628-631.
41
42    de Marco, R., Pili, A., Ferrari, M., According  S., Giammanco, G., Bugiani,  M., Villani, S., Ponzio, M.,
43            Bono, R., Carrozzi, L., Cavallini,  R.,  Cazzoletti,  L., Dallari, R., Ginesu, F., Lauriola, P.,
44            Mandrioli, P., Perfetti, L., Pignato, S., Pirina, P., and Struzzo, P. 2002.  The impact of climate and
45            traffic-related NO2 on the prevalence of asthma and allergic rhinitis in Italy. Clin. Exp. Allergy
46            32:1405-1412.
47
48    Detels, R.,  Sayre, J.W., Coulson, A.H., Rokaw, S.N., Massey, F.J., Jr., Tashkin, D.P., and Wu, M.-M.
49            1981. The UCLA population studies of chronic obstructive respiratory disease. IV. Respiratory
50            effect of long-term exposure to photochemical oxidants, nitrogen dioxide, and sulfates on current
51            and never smokers.  Am. Rev. Respir. Dis. 124:673-680.
52
53    Devalia, J.L., Rusznak, C., Herdman, M.J., Trigg, C.J., Tarraf, H., and Davies, RJ.  1994. Effect of
54            nitrogen dioxide and sulphur dioxide on airway response of mild asthmatic patients to allergen
55            inhalation. Lancet 344:1668-1671.
56
                                                     52

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1    Devlin, R., Horstman, D., Becker, S., Gerrity, T., Madden, M, and Koren, H.  1992. Inflammatory
 2            response in humans exposed to 2.0 ppm NC>2. Am. Rev. Respir. Dis. 145:A456.
 3
 4    Dillmann, G., Henschler, D., and Thoenes, W. 1967.  [Effects of nitrogen dioxide on lung alveoli in
 5            the mouse. Morphometric-electron microscopic studies.] Archiv fur Toxikologie 23:55-65.
 6            (translated from German).
 7
 8    Dockery, D.W., Speizer, F.E., Stram, D.O., Ware, J.H., Spengler, J.D, and Ferris, E.G., Jr.  1989.
 9            Effects of inhalable particles on respiratory health of children.  Am. Rev. Respir. Dis.
10            139:587-594.
11
12    Douglas, W.W., Hepper, N.G.G., and Colby, T.V. 1989. Silo-filler's disease. Mayo Clin. Proc. 64:291-
13            304.
14
15    Dowell, A.R., Kilburn, K.H., and Pratt, P.C.  1971. Short-term exposure to nitrogen dioxide: Effects
16            on pulmonary ultrastructure, compliance, and the surfactant system.  Arch. Intern. Med.
17            128:74-80.
18
19    Ehrlich, R. 1978. Interactions of various pollutants on causation of pulmonary disease.  Report: ISS
20            EPA/600/1-78-057, Order No. PPB-288363,  48 pp.
21
22    Elsayed, N.M.  1994. Toxicity of nitrogen dioxide: an introduction. Toxicology 89:161-174.
23
24    Elsayed, N.M., Gorbunov, N.V., Mayorga, M.S., Kagan, V.E., and Januszkiewicz, A.J.  2002.
25            Significant pulmonary response to a brief high-level, nose-only nitrogen dioxide exposure: an
26            interspecies dosimetry perspective. Toxicol. Appl. Pharmacol.  184:1-10.
27
28    Euler, G.L., Abbey, D.E., Hodgkin, J.E., and Magie, A.R.  1988. Chronic obstructive pulmonary disease
29            symptom effects of long-term cumulative exposure to ambient levels of total oxidants and
30            nitrogen dioxide in California Seventh-day Adventist residents. Arch. Environ. Health 43:279-
31            285.
32
33    Farrow, A., Greenwood, R., Preece, S., Golding, J. 1997. Nitrogen dioxide, the oxides of nitrogen, and
34            infants' health symptoms.  Arch. Environ. Health 52:189-194.
35
36    Florey,  C.du V., Melia, R.J.W., Chinn, S., Goldstein, B.D., Brooks, A.G.F., John, H.H., Craighead, I.E.,
37            and Webster, X. 1979. The relationship between respiratory illness in primary school children
38            and the use of gas for cooking. Ill - Nitrogen dioxide, respiratory illness, and lung infection. Int.
39            J. Epidemiol. 8:347-353.
40
41    Folinsbee, L.J.  1992. Does nitrogen dioxide exposure  increase airways responsiveness? Toxicol. Indus.
42            Health 8:273-283.
43
44    Folinsbee, L.J., Horvath, S.M., Bedi, J.F., and Delehunt, J.C. 1978. Effect of 0.62 ppm NO2 on
45            cardiopulmonary function in young male nonsmokers.  Environ. Res. 15:199-205.
46
47    Frampton, M.W., Morrow, P.E., Cox, C., Gibb, R., Speers, D.M., and Utell, M. 1991. Effects of nitrogen
48            dioxide exposure on pulmonary function and airway reactivity in normal humans. Am. Rev.
49            Respir. Dis. 143:522-527.
50
51    Frampton, M.W., Voter, K.Z., Morrow, P.E., Roberts, N.J., Jr., Gavras,  J.B., and Utell, M.J. 1992.
52            Effects of NO2exposure on human host defense.  Am. Rev. Respir. Dis. 145:A455.
53
54    Gamble, J., Jones, W., and Minshall, S.  1987. Epidemiological-environmental study of diesel bus garage
55            workers: acute effects of NO2 and respirable  paniculate on the respiratory system. Environ. Res.
56            42:201-214.
                                                     53

-------
      NITROGEN OXIDES                   NAC/Interim (NO2)/Proposed (N2O4):  12/2008


 1
 2    Gardner, D.E., Holzman, R.S., and Coffin, D.L. 1969. Effects of nitrogen dioxide on pulmonary cell
 3            population.  J. Bacteriol. 98:1041-1043.
 4
 5    Gardner, D.E., Coffin, D.L., Pinigin, M.A., and Sidorenko, G.I. 1977. Role of time as a factor in the
 6            toxicity of chemical compounds in intermittent and continuous exposures. Part I. Effects of
 7            continuous exposure. J. Toxicol. Environ. Health 3:811-820.
 8
 9    Gardner, D.E., Miller, F.J., Blommer, E.J., and Coffin, D.L. 1979. Influence of exposure mode on the
10            toxicity of NO2. Environ. Health Perspect. 30:23-29.
11
12    Gelzleichter, T.R., Witschi, H., and Last, J.A. 1992. Concentration-response relationships of rat lungs to
13            exposure to oxidant air pollutants:  a critical test of Haber's law for ozone and nitrogen dioxide.
14            Toxicol. Appl. Pharmacol. 112:73-80.
15
16    German Research Association (Deutsche Forschungsgemeinschaft). 2002. List of MAK and BAK Values,
17            2002. Commission for the Investigation of Health Hazards of Chemical Compounds in the Work
18            Area, Report No. 36. Weinheim, Federal Republic of Germany: Wiley VCH.
19
20    Gilmour, M.I. 1995. Interaction of air pollutants and pulmonary allergic responses in experimental
21            animals. Toxicology 105:335-342.
22
23    Goings, S.A.J., Kulle, T.J., Bascom, R., Sauder, L.R., Green, D.J., Hebel, J.R., Clements, M.L. 1989.
24            Effect of nitrogen dioxide exposure on susceptibility to influenza A virus infection in healthy
25            adults. Am. Rev. Respir. Dis. 139:1075-1081.
26
27    Goldstein, E., Eagle, M.C., and Hoeprich, P.O. 1973. Effect of nitrogen dioxide on pulmonary
28            bacterial defense mechanisms.  Arch. Environ. Health 26:202-204.
29
30    Goldstein, E., Peek, N.F., Parks, N.J., Hines, H.H.,  Steffey, E.P., and Tarkington, B.  1977. Fate and
31            distribution of inhaled nitrogen dioxide in  Rhesus monkeys. Am. Rev. Resp. Dis. 115:403-
32            412.
33
34    Goldstein, I.F., Lieber, K., Andrews, L.R., Kazembe, F., Foutrakis, G., Huang, P., and Hayes, C.  1988.
35            Acute respiratory effects of short-term exposures to nitrogen dioxide. Arch. Environ. Health
36            43:138-142.
37
38    Gordon, R.E., Case, B.W., and Kleinerman, J.  1983. Acute NO2 effects on penetration and transport
39            of horseradish peroxidase in hamster respiratory epithelium. Am. Rev. Respir. Dis.
40             128:528-533.
41
42    Gray, E.Le B., Patton, P.M., Goldberg, S.B., and Kaplan, E. 1954. Toxicity of the oxides of nitrogen.
43            II. Acute inhalation toxicity of nitrogen dioxide, red fuming nitric acid, and white fuming
44            nitric acid. Arch. Ind. Hyg. Occup. Med. 10:418-422.
45
46    Grayson, R.R.  1956. Silage gas poisoning:  nitrogen dioxide pneumonia, a new disease in agricultural
47            workers.  Ann. Int. Med. 45:393-408.
48
49    Greenbaum, R., Bay, J., Hargreaves, M.D.,  Kain, M.L., Kelman, G.R., Nunn, J.F., Prys-Roberts, C.,
50            and Siebold, K. 1967. Effects of higher oxides of nitrogen on the anaesthetized dog. Brit. J.
51            Anaesth. 39:393-404.
52
53    Guth, D. J. and Mavis, R.D. 1985. Biochemical assessment of acute nitrogen dioxide toxicity in rat lung.
54            Toxicol. Appl. Pharmacol. 81:128-138.
55
56    Guth, D.J. and Mavis, R.D. 1986. The effect of lung -tocopherol content on the acute toxicity of
                                                     54

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1            nitrogen dioxide. Toxicol. Appl. Pharmacol. 84:304-314.
 2
 3    Hackney, J.D., Thiede, F.C., Linn, W.S., Pedersen, E.E., Spier, C.E., Law, D.C., and Fischer, D.A. 1978.
 4            Experimental studies on human health effects of air pollutants. IV. Short-term physiological and
 5            clinical effects of nitrogen dioxide exposure. Arch. Environ. Health 33:176-181.
 6
 7    Hackney, J.D., Linn, W.S., Avol, E.L., Shamoo, D.A., Anderson, K.R., Solomon, J.C., Little, D.E.,
 8            and Peng, R.-C. 1992. Exposures of older adults with chronic respiratory illness to nitrogen
 9            dioxide.  A combined laboratory and field study.  Am. Rev. Respir. Dis. 146:1480-1486.
10
11    Hamilton, A. 1983. Nitrogen compounds. In: Hamilton and Hardy's Industrial Toxicology, 4th ed.
12            Boston: John Wright-PSG, Inc. pp.184-186.
13
14    Hatch, G.E., Slade, R., Selgrade, M.K., and Stead, A.G. 1986. Nitrogen dioxide exposure and lung
15            antioxidants in ascorbic acid-deficient guinea pigs.  Toxic. Appl. Pharmacol. 82:351-359.
16
17    Hayashi, Y., Kohno, T., and Ohwada, H.  1987. Morphological effects of nitrogen dioxide on the rat
18            lung. Environ. Health Persp. 73:135-145.
19
20    Hazucha, M.J.,  Ginsberg, J.F., McDonnell, W.F., Haak, E.D., Jr., Pimmel, R.L., Salaam, S.A., House,
21            D.E., andBromberg,  P. A.  1983. Effects of 0.1 ppm nitrogen dioxide on airways of normal
22            and asthmatic subjects. J. Appl. Physiol.: Respirat.  Environ. Exercise Physiol.  54:730-739.
23
24    Hazucha, M.J.,  Folinsbee, L.J., Seal, E., and Bromberg, P.A.  1994. Lung function response of
25            healthy women after sequential exposures to NO2 and O3. Am. J. Respir. Crit. Care Med.
26             150:642-647.
27
28    Hedberg, K., Hedberg, C.W., Iber, C, White, K.E., Osterholm, M.T., Jones, D.B.W., Flink, J.R., and
29            MacDonald, K.L.  1989. An outbreak of nitrogen dioxide-induced respiratory illness among ice
30            hockey players. J. Am. Med. Assoc. 262:3014-3017.
31
32    Helleday, R., Huberman, D., Blomberg, A., Stjernberg, N., and Sandstrom, T.  1995. Nitrogen dioxide
33            exposure impairs the  frequency of the mucociliary activity in healthy subjects.  Eur. Respir. J.
34            8:1664-1668.
35
36    Henry, M.C., Ehrlich, R., and Blair, W.H. 1969. Effect of nitrogen dioxide on resistance of
37            squirrel monkeys to Klebsiellapneumoniae infection. Arch. Environ. Health 18:580-587.
38
39    Henschler, D. and Liitke, W. 1963.  [Methemoglobin formation due to inhalation of low concentrations
40            of nitroso gases.]  Internal. Arch. Gewerbepath. 20:362-370. (translated from German).
41
42    Henschler, D., Stier, A., Beck, H., and Neuman, W. 1960.  [Odor threshold of a few important irritant
43            gasses (sulfur dioxide, ozone, nitrogen dioxide) and observations in humans exposed to low
44            concentrations.] Archiv fur Gewerbepathologie und Gewerbehygiene 17:547-570.
45            (translated from German).
46
47    Hidekazu, F. and Fujio, S. 1981. Effects of acute exposure to nitrogen dioxide on primary antibody
48            response.  Arch. Environ. Health 36:114-119.
49
50    Hillam, R.P., Bice, D.E., Hahn, F.F., and Schnizlein, C.T.  1983. Effects of acute nitrogen dioxide
51            exposure on cellular immunity after lung immunization. Environ. Res. 31:201-211.
52
53    Hine, C.H., Meyers, F.H., and Wright, R.W. 1970. Pulmonary changes in animals exposed to nitrogen
54            dioxide, effects of acute exposures. Toxicol. Appl. Pharmacol. 16:201-213.
55
56    Hugod, C.  1979. Effect of exposure to 43 ppm nitric oxide and 3.6 ppm nitrogen dioxide on rabbit
                                                     55

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1            lung. A light and electron microscopic study.  Int. Arch. Occup. Environ. Health 42:159-167.
 2
 3    Ichinose, T., Fujii, K., and Masaru, S. 1991. Experimental studies on tumor promotion by nitrogen
 4            dioxide. Toxicology 67:211-225.
 5
 6    Illing, J.W., Miller, F.J., and Gardner, D.E.  1980. Decreased resistance to infection in exercised mice
 7            exposed to NO2 and O3. J. Toxicol. Environ. Health 6:843-851.
 8
 9    Isomura, K., Chikahira, M, Teranishi, K., and Hamada, K. 1984. Induction of mutations and chromosome
10            aberrations in lung cells following in vivo exposure  of rats to nitrogen oxides. Mut. Res. 136:119-
11             125.
12
13    Jaakkola, U.K., Paunio, M., Virtanen, M., and Heinonen, O.P.  1991. Low-level air pollution and
14            upper respiratory infections in children. Am. J. Public Health 81:1060-1063.
15
16    Jakab, G.J. 1987. Modulation of pulmonary  defense mechanisms by acute exposures to nitrogen dioxide.
17            Environ. Res. 42:215-228.
18
19    Januszkiewicz, AJ. and Mayorga, M.A.  1994. Nitrogen dioxide-induced acute lung injury in sheep.
20            Toxicology 89:279-300.
21
22    Jenkins, H.S., Devalia, J.L., Mister, R.L., Bevan, A.M., Rusznak, C., and Davies, RJ.  1999. The effect
23            of exposure to ozone and nitrogen dioxide on the airway response of atopic asthmatics to
24            inhaled allergen. Dose- and time-dependent effects. Am. J. Respir. Crit. Care Med. 160:33-39.
25
26    Jones, G.R., Proudfoot, A.T.,  and Hall, J.I. 1973. Pulmonary effects of acute exposure  to nitrous fumes.
27            Thorax 28:61-65.
28
29    Jorres, R. and Magnussen, H.  1990. Airways response of asthmatics after a 30 min exposure, at resting
30            ventilation, to 0.25 ppmNO2 or 0.5 ppm SO2. Eur. Respir. J. 3:132-137.
31
32    Jorres, R. and Magnussen, H. 1991. Effect of 0.25 ppm nitrogen dioxide on the airway response to
33            methacholine in asymptomatic asthmatic patients. Lung 169:77-85.
34
35    Jorres, R., Nowak, D., Grimminger, F., Seeger, W., Oldigs, M., and Magnussen,  H. 1995. The effect of
36             1 ppm nitrogen dioxide on bronchoalveolar lavage cells and inflammatory mediators in normal
37            and asthmatic subjects. Eur. Respir. J. 8:416-424.
38
39    Karlson-Stiber, C., Hojer, J.,  Sjoholm, A., Bluhm, G., Salmonson, H.  1996. Nitrogen  dioxide
40            pneumonitis in ice hockey players.  J. Intern. Med. 239:451-456.
41
42    Kerr, H.D., Kulle, T. J., Mcllhany, M.L., and Swidersky, P. 1978. Effects of nitrogen dioxide on
43            pulmonary function  in human subjects. An environmental chamber study. Report:  ISS
44            EPA/600/1-78/025; Order no. PB-281 186, 20  pp.
45
46    Kerr, H.D., Kulle, T.J., Mcllhany, M.L., and Swidersky, P. 1979. Effects of nitrogen dioxide on
47            pulmonary function  in human subjects: An environmental chamber study. Environ. Research
48             19:392-404.
49
50    Kim, S.U., Koenig, J.Q., Pierson, W.E., and Hanley, Q.S. 1991. Acute pulmonary effects of nitrogen
51            dioxide exposure during exercise in competitive athletes. Chest 99:815-819.
52
53    Kleinman, M.T., Bailey, R.M., Linn, W.S., Anderson, K.R., Whynot, J.D., Shamoo, D.A., and Hackney,
54            J.D.  1983. Effects of 0.2 ppm nitrogen dioxide on pulmonary function and response to
55            bronchoprovocation in asthmatics.  J. Toxicol. Environ. Health 12:815-826.
56
                                                    56

-------
      NITROGEN OXIDES                   NAC/Interim (NO2)/Proposed (N2O4):  12/2008


 1    Kobayashi, T., Noguchi, T., Kikuno, M., and Kubota, K. 1984. Effect of acute nitrogen dioxide
 2             exposure on the composition of fatty acid associated with phospholipids in alveolar
 3             lavage. Chemosphere 13:101-105.
 4
 5    Koenig, J.Q., Covert, D.S., Morgan, M.S., Horike, M., Horike, N., Marshall, S.G., and Pierson,
 6             W.E.  1985. Acute effects of 0.12 ppm ozone or 0.12 ppm nitrogen dioxide on pulmonary
 7             function in healthy and asthmatic adolescents. Am. Rev. Respir. Dis.  132:648-651.
 8
 9    Koenig, J.Q., Covert, D.S., Marshall, S.G., van Belle, G., and Pierson, W.E. 1987. The effects of ozone
10             and nitrogen dioxide on pulmonary function in healthy and in asthmatic adolescents.  Am. Rev.
11             Respir. Dis. 136:1152-1157.
12
13    Kushneva, V.S. and Gorshkova, R.B. (Eds.)  1999.  [Nitrogen tetroxide.  In: Reference Book on
14            Toxicology and Hygienic Standards (MAC) of Potentially Hazardous Chemical Substances.]
15            IzdAT:Moscow.  272pp. (translated from Russian)
16
17    Lehnert, B.E., Archuleta, D.C., Ellis, T., Session, W.S., Lehnert, N.M., Gurley, L.R., and Stavert,
18             D.M.  1994. Lung injury following exposure of rats to relatively high mass concentrations of
19             nitrogen dioxide.  Toxicology 89:239-277.
20
21    Le Tertre, A., Quenel, P., Eilstein, D., Medina, S., Prouvost, H., Pascal, L., Boumghar, A.,  Saviuc, P.,
22             Zeghnoun, A., Filleul, L., Declercq, C., Cassadou, S., and Le Coaster, C. 2002.  Short-term
23             effects of air pollution on mortality in nine French cities: a quantitative summary.  Arch.
24             Environ, Health 57:311-319.
25
26    Lide, D.R. (Ed.) 1988. CRC Handbook of Chemistry and Physics, 69th Edition.  Boca Raton:CRC Press,
27            Inc.
28
29    Lin,  M., Chen, Y., Burnett, R.T., Villeneuve, P.J., and Krewski, D. 2003. Effect of short-term exposure to
30            gaseous pollution on asthma hospitalization in children: a bi-directional case-crossover analysis.
31            J. Epidemiol. Community Health 57:50-55.
32
33    Linn, W.S. and Hackney, J.D. 1983. Short-term human respiratory effects of nitrogen dioxide:
34             determination of quantitative dose-response profiles. Phase 1. Exposure of healthy volunteers
35             to 4 ppm NO2. NTIS order no. PB84-132299. 30 pp.
36
37    Linn, W.S. and Hackney, J.D.  1984. Short-term human respiratory effects of nitrogen dioxide:
38             determination of quantitative dose-response profiles. Phase 2. Exposure of asthmatic volunteers
39             to 4 ppm NO2. NTIS order no. PB85-104388. 31 pp.
40
41    Linn, W.S., Shamoo, D.A., Spier, C.E., Valencia, L.M., Anzar, U.T., Venet, T.G., Avol,  E.L.,  and
42             Hackney, J.D. 1985. Controlled exposure  of volunteers with chronic  obstructive pulmonary
43             disease to nitrogen dioxide. Arch. Environ. Health 40:313-317.
44
45    Liu, S., Krewski, D., Shi, Y., Chen, Y., and Burnett, R.T. 2007.  Association between maternal exposure
46             to ambient air pollutants during pregnancy and fetal growth restriction.  J. Expo. Sci,  Environ.
47             Epidemiol. 17:426-432.
48
49    Lowry, T. and Schuman, L.M. 1956. "Silo-filler's disease" -  a syndrome caused by nitrogen dioxide.
50             J. Am. Med. Assoc. 162:153-160.
51
52    Maigetter, R.Z., Fenters, J.D., Findlay, J.C., Ehrlich, R., and Gardner, D.E. 1978. Effects of
53             exposure to nitrogen dioxide on T and B cells in mouse spleens. Toxicol. Letters 2:157-161.
54
55    Matsumura, Y.  1970. The effects of ozone, nitrogen dioxide, and sulfur  dioxide on the experimentally
56             induced allergic respiratory disorder in guinea pigs. 1. The effect on sensitization with albumin
                                                     57

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1            through the airway.  Am. Rev. Resp. Dis. 102:430-437.
 2
 3    Meulenbelt, I, Dormans, J.A.M.A., Marra, M., Rombout, P.J.A., and Sangster, B. 1992a. Rat model to
 4            investigate the treatment of acute nitrogen dioxide intoxication. Human Exp. Toxicol. 11:179-
 5            187.
 6
 7    Meulenbelt, I, van Bree, L., Dormans, J.A.M.A., Boink, A.B.T.J., and Sangster, B.  1992b. Biochemical
 8            and histological alterations in rats after acute nitrogen dioxide intoxication.  Human Exp. Toxicol.
 9            11:189-200.
10
11    Meulenbelt, J., van Bree, L., Dormans, J. A.M. A., Boink, A.B.T.J., and Sangster, B.  1994.
12            Development of a rabbit model to investigate the effects of acute nitrogen dioxide
13            intoxication.  Human Exp. Toxicol.  13:749-759.
14
15    Miller, F.J., Graham, J.A., Illing, J.W., and D.E. Gardner. 1980. Extrapulmonary effects of NO2 as
16            reflected by pentobarbital-induced sleeping time in mice.  Toxicol. Lett.6:267-274.
17
18    Millstein, J., Gilliland, F., Berhane, K., Gauderman, W.J., McConnell, R., Avol, E., Rappaport, E.B.,
19            and Peters, J.M. 2004.  Effects of ambient air pollutants on asthma medication use and
20            wheezing among fourth-grade school children from 12 southern California communities
21            enrolled in the children's health study. Arch. Environ. Health 59:505-514.
22
23    Milne, J.E.H. 1969. Nitrogen dioxide inhalation and bronchiolitis obliterans. A review of the literature
24            and report of a case. J. Occup. Med. 11:538-547.
25
26    Mohsenin, V. 1987. Airway responses to nitrogen dioxide in asthmatic subjects. J. Toxicol. Environ.
27            Health 22:371-380.
28
29    Mohsenin, V., 1988. Airway responses to 2.0 ppm nitrogen dioxide in normal subjects.  Arch. Environ.
30            Health 43:242-246.
31
32    Mohsenin, V. 1991. Lipid peroxidation and antielastase activity in the lung under oxidant stress: role
33            of antioxidant defenses. J. Appl. Physiol. 70:1456-1462.
34
35    Mohsenin, V. 1994. Human exposure  to oxides of nitrogen at ambient and supra-ambient
36            concentrations. Toxicology 89:301-312.
37
38    Mohsenin, V. and Gee, J.B.L. 1987. Acute effect of nitrogen dioxide exposure on the functional
3 9            activity of alpha-1 -protease inhibitor in bronchoalveolar lavage fluid of normal subjects.
40            Am. Rev. Respir. Dis. 136:646-650.
41
42    Morgan, W.K.C.  1995.'Zamboni disease'Pulmonary edema in an ice hockey player.  Arch. Intern.
43            Med. 155:2479-2480.
44
45    Morley, R. and Silk, S.J. 1970. The industrial hazard from nitrous fumes. Ann. Occup. Hyg. 13:101-107.
46
47    Morrow, P.E. and Utell, M. J.  1989. Responses of susceptible subpopulations to nitrogen dioxide.
48            Cambridge, MA: Research Rept. Health Effects Inst, report no. 23.
49
50    Morrow, P.E., Utell, M.J., Bauer, M.A., Smeglin, A.M., Frampton, M.W., Cox, C., Speers, D.M., and
51            Gibb, F.R.  1992. Pulmonary performance of elderly normal subjects and subjects with chronic
52            obstructive pulmonary disease exposed to 0.3 ppm nitrogen dioxide.  Am. Rev. Respir. Dis.
53            145:291-300.
54
55    Mortimer, K.M., Neas, L.M., Dockery, D. W., Redline, S., and Tager, IB.  2002.  The effect of air
56            pollution on inner-city children with asthma. Eur. Respir. J. 19:699-705.
                                                     58

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1
 2    Mostardi, R.A., Woebkenberg, N.R., Ely, D., Atwood, G., Conlon, M., Jarrett, M., and Dahlin, M.  1981.
 3            Project summary: Air pollution and health effects in children residing in Akron, Ohio.  Report:
 4            ISS EPA-600/1-81-004; Order no. PB 81-152498.
 5
 6    Murphy, S.D., Ulrich, C.E., Frankowitz, S.H., and Xintaras, C.  1964.  Altered function in
 7            animals inhaling low concentrations of ozone and nitrogen dioxide. Am. Ind. Hyg.
 8            Assoc. J. 25:246-253.
 9
10    Neas, L.M., Dockery, D.W., Ware, J.H., Spengler, J.D., Speizer, F.E.,  and Ferris, E.G.  1991.
11            Association of indoor nitrogen dioxide with respiratory symptoms and pulmonary function in
12            children. Am. J. Epidemiol. 134:204-219.
13
14    NIOSH. 1976. National Institute for Occupational Safety and Health.  NIOSH criteria for a
15            recommended standard.... occupational exposure to oxides of nitrogen (nitrogen dioxide and
16            nitric oxide). U.S. Department of Health, Education, and Welfare, Washington, D.C., HEW
17            publication No. (NIOSH) 76-149, 195pp.
18
19    NIOSH. 1996. National Institute for Occupational Safety and Health.  Nitrogen Dioxide.
20            Documentation for Immediately Dangerous to Life or Health Concentrations (IDLHs).
21            NIOSH, Cincinnati,  OH, last updated 8/16/96.
22            
23
24    NIOSH. 2003. National Institute for Occupational Safety and Health.  NIOSH Pocket Guide to
25            Chemical Hazards. NIOSH, Cincinnati, OH, online version. Accessed January, 2004.
26
27    Norwood, W.D., Wisehart, D.E., Earl, C.A, Adley, F.E., and Anderson, D.E.  1966. Nitrogen dioxide
28            poisoning due to metal-cutting with oxyacetylene torch.  J. Occup. Med. 8:301-306.
29
30    NRC. 1977. National Research Council. Medical and Biologic Effects of Environmental Pollutants.
31            Nitrogen Oxides. NRC, National Academy of Sciences, Washington, DC. 333pp.
32
33    NRC. 1985. National Research Council. Emergency and Continuous Exposure Guidance Levels for
34            Selected Airborne Contaminants.  Volume 4. pp. 83-95. National Academy Press, Washington,
35            DC.
36
37    Oda, H., Nogami, H., Nakajima, T.  1980. Reaction of hemoglobin with nitric oxide and nitrogen
38            dioxide in mice. J. Toxicol. Environ. Health 6:673-678.
39
40    Oda, H., Tsubone, H., Suzuki, A., Ichinose, T., and Kubota, K.  1981. Alterations of nitrite and nitrate
41            concentrations in the blood of mice exposed to nitrogen dioxide.  Environ. Res. 25:294-301.
42
43    Orehek, J., Massari,  J.P., Gayrard, P., Grimaud, C., and Charpin, J. 1976. Effect of short-term,
44            low-level nitrogen dioxide exposure on bronchial sensitivity of asthmatic patients.  J. Clin.
45            Invest. 57:301-307.
46
47    OSHA. 1999. Occupational Safety and Health Administration.  Table Z-l. Limits for Air
48            Contaminants. 29 CFR (§1910.1000), p. 14.
49
50    Pattenden, S., Hoek,  G., Braun-Fahrlander, C., Forastiere, F., Kosheleva, A., Neuberger, M., and Fletcher,
51            T. 2006. NO2 and children's respiratory symptoms in the PATY study. Occup. Environ. Med.
52            63:828-835.
53
54    Penard-Morand, C.,  Charpin, D., Raherison, C., Kopferschmitt, C., Caillaud, D., Lavaud, F., and
55            Annesi-Maesano, I.  2005. Long-term exposure to background air pollution related to
56            respiratory  and allergic health in schoolchildren.  Clin. Exp. Allergy 35:1279-1287.
                                                    59

-------
      NITROGEN OXIDES                   NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1
 2    Peters, J.M., Murphy, R.L.H., Ferris, E.G., Burgess, W.A., Ranadive, M.V., and Perdergrass, H.P.
 3             1973. Pulmonary function in shipyard welders an epidemiologic study. Arch. Environ.
 4             Health 26:28-31.
 5
 6    Peters, J.M., Avol, E., Navidi, W., London, S.J., Gauderman, W.J., Lurmann, F., Linn, W.S., Margolis, H.,
 7             Rappaport, E., Gong, H., Jr., and Thomas, D.C.  1999a. A study of twelve southern California
 8             communities with differing levels and types of air pollution. I. Prevalence of respiratory
 9             morbidity. Am. J. Resp. Crit. Care Med. 159:760-767.
10
11    Peters, J.M., Avol, E., Navidi, W., London, S.I, Gauderman, W.J., Lurmann, F., Linn, W.S., Margolis, H.,
12             Rappaport, E., Gong, H., Jr., and Thomas, D.C.  1999b. A study of twelve southern California
13             communities with differing levels and types of air pollution. II. Effects on pulmonary function.
14             Am. J. Resp. Crit. Care Med. 159:768-775.
15
16    Pilotto, L.S., Douglas, R.M., Attewell, R.G., and Wilson, S.R. 1997. Respiratory effects associated with
17             indoor nitrogen dioxide exposure in children. Int. J. Epidemiol. 26:788-796.
18
19    Posin, C., Clark, K., Jones, M.P., Patterson, J.V., Buckley, R.D., and Hackney, J.D. 1978. Nitrogen
20             dioxide inhalation and human blood biochemistry. Arch. Environ. Health 33:318-324.
21
22    Postlethwait, E.M. and Bidani, A. 1990. Reactive uptake governs the pulmonary air space removal of
23             inhaled nitrogen dioxide. J. Appl. Physiol. 68:594-603.
24
25    Postlethwait, E.M. and Bidani, A. 1994. Mechanisms of pulmonary NO2 absorption. Toxicology 89:217-
26             237.
27
28    Postlethwait, E.M. and Mustafa, M.G. 1981. Fate of inhaled nitrogen dioxide in isolated perfused rat
29             lung. J. Toxicol. Environ. Health 7:861-872.
30
31    Rasmussen, R.E.  1992. Effects of acute NC>2 exposure in the weanling ferret lung. Inhal. Toxicol. 4:373-
32             382.
33
34    Rasmussen, T.R., Kjaergaard, S.K., Tarp, U., and Pedersen, O.F.  1992. Delayed effects of NO2
35            exposure on alveolar permeability and glutathione peroxidase in healthy humans. Am. Rev.
36            Respir. Dis.  146:654-659.
37
38    Robertson, A., Dodgson, J., Collings, P., and Seaton, A. 1984. Exposure to oxides of nitrogen: respiratory
39             symptoms and lung function in British coalminers.  Br. J. Ind. Med. 41:214-219.
40
41    Roger, L.J., Horstman, D.H., McDonnell, W., Kehrl, H., Ives, P.J., Seal, E., Chapman, R., and Massaro,
42             E. 1990. Pulmonary function, airway  responsiveness, and respiratory symptoms in asthmatics
43             following exercise inNO2. Toxicol. Ind. Health 6:155-171.
44
45    Rombout, P. J.A., Dormans, J.A.M.A., Marra, M., and van Esch, J.  1986. Influence of exposure
46             regimen on nitrogen dioxide-induced morphological changes in the rat lung. Environ. Res.
47             41:466-480.
48
49    Rose, R.M., Pinkston, P., and Skornik, W.A. 1989. Altered susceptibility to viral respiratory infection
50             during short-term exposure to nitrogen dioxide. Res. Rep. Health Eff. Inst. 24:1-24.
51
52    Rubinstein, I., Bigby, E.G., Reiss, T.F., and Boushey, H.A., Jr. 1990. Short-term exposure to 0.3 ppm
53             nitrogen dioxide does not potentiate airway responsiveness to sulfur dioxide in asthmatic
54             subjects.  Am. Rev.  Respir. Dis. 141:381-385.
55
56    Russell, M.L., Need, J.L., Mercer, R.R., Miller, F.J., and Crapo, J.D. 1991. Distribution of inhaled
                                                     60

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1             [15O]-NO2 in the upper and lower respiratory tracts of rats. Am. Rev. Respir. Dis.
 2             134:A704.
 3
 4     Sackner, M.A., Birch, S., Friden, A., and Marchetti, B. 1981. Effects of breathing low levels of nitrogen
 5             dioxide for four hours on pulmonary function of asthmatic adults.  Am. Rev. Respir. Dis. 123:151.
 6
 7     Samoli, E., Aga, E., Touloumi, G., Nisiotis, K., Forsberg, B., Lefranc, A., Pekkanen, J. Wojtyniak, B.,
 8             Schindler, C., Niciu, E., Brunstein, R., Dodic Fikfak, M., Schwartz, J., Katsouyanni, K.  2006.
 9             Short-term effects of nitrogen dioxide on mortality: an analysis within the APHEA project. Eur.
10             Respir. 127:1129-1138.
11
12     Saul, R.L. and Archer, M.C.  1983. Nitrate formation in rats exposed to nitrogen dioxide. Toxicol.
13             Appl. Pharmacol. 67:284-291.
14
15     Schindler, C., Ackermann-Liebrich, U., Leuenberger, P., Monn, C., Rapp, R., Bolognini, G., Bongard, I-
16             P., Brandli, O., Domenighetti, G., Karrer, W., Keller, R., Medici, T.G., Perruchoud, A.P.,
17             Schoni, M.H.,  Tschopp, J.-M, Villiger, B., Zellweger, J.-P., and the SAPALDIA Team.  1998.
18             Associations between lung function and estimated average exposure to NO2 in eight areas of
19             Switzerland. Epidemiology 9:405-411.
20
21     Schlesinger, R.B., Driscoll, K.E.,  Gunnison, A.F., and Zelikoff, J.T.  1990. Pulmonary arachidonic
22             acid metabolism following acute exposures to ozone and nitrogen dioxide. J. Toxicol.
23             Environ. Health 31:275-290.
24
25     Schnizlein, C.T., Bice, D.E.,  Rebar, A.H., Wolff, R.K., and Beethe, R.L.  1980. Effect of lung damage by
26             acute exposure to nitrogen dioxide on lung immunity in the rat.  Environ. Res. 23:362-370.
27
28     Schwartz, J.,  Spix,  C., Wichmann, H.E., and Malin, E.  1991. Air pollution and acute  respiratory illness
29             in five German communities. Environ. Res. 56:1-14.
30
31     SDU Uitgevers (Ministry of Social Affairs and Employment). 2000. Nationale MAC
32             (Maximum Allowable Concentration) List, 2000. The Hague, The Netherlands.
33
34    Selegrade, M.K., Mole,  M.L., Miller, F.J., Hatch, G.E., Gardner, D.E., an Hu., P.C. 1981. Effect of NO2
35            inhalation and vitamin C deficiency on protein and lipid accumulation in the  lung. Environ. Res.
36            26:422-437.
37
38     Shima, M. and Adachi,  M. 2000. Effect of outdoor and indoor nitrogen dioxide on respiratory
39             symptoms in schoolchildren. Int. J. Epidemiol. 29:862-870.
40
41     Shima, M., Nitta, Y., Ando, M., and Adachi, M. 2002. Effects of air pollution on the prevalence and
42             incidence  of asthma in children. Arch. Environ. Health 57:529-535.
43
44     Siegel, P.D., Bozelka, B.E., Reynolds, C., and George, W.J. 1989. Phase-dependent response of the lung
45             to NO2 irritant insult. J. Environ. Path. Toxicol. Oncolog. 9:303-315.
46
47     Siegel, P.O., Al-Humadi, N.H., Nelson, E.R., Lewis, D.M., and Hubbs, A.F. 1997. Adjuvant effect of
48             respiratory irritation on pulmonary allergic sensitization: time and site dependency.  Toxicol.
49             Appl. Pharmacol. 144:356-362.
50
51     Silbaugh, S.A., Mauderly, J.L., and Macken, C.A.  1981. Effects of sulfuric acid and nitrogen
52             dioxide on airway responsiveness of the guinea pig. J. Toxicol. Environ. Health 8:31-45.
53
54     Smeglin, A.M., Utell, M.J., Bauer, M.A., Speers, D.M., Gibb, F.R., and Morrow, P.E. 1985. Low-level
55             nitrogen dioxide exposure does not alter lung function in exercising healthy subjects. Am. Rev.
56             Respir. Dis. 131:A171.
                                                     61

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1
 2    Smith, W., Anderson, T., and Anderson, H.A.  1992. Nitrogen dioxide and carbon monoxide intoxication
 3            in an indoor ice arena - Wisconsin, 1992.  MMWR Weekly 41:3 83 -3 85.
 4            
 5
 6    Soparkar, G., Mayers, I., Edouard, L., and Hoeppner, V.H. 1993. Toxic effects from nitrogen dioxide
 7            in ice-skating arenas. Can. Med. Assoc. J.  148:1181-1182.
 8
 9    Speizer, F.E. and Ferris, E.G., Jr. 1973. Exposure to automobile exhaust. I. Prevalence of respiratory
10            symptoms and disease. Arch. Environ. Health 26:313-318.
11
12    Stavert, D.M. and Lehnert, B.E. 1990. Nitric oxide  and nitrogen dioxide as inducers of acute
13            pulmonary injury when inhaled at relatively high concentrations for brief periods.  Inhal.
14            Toxicol. 2:53-67.
15
16    Stephens, R.J., Freeman, G., and Evans, M.J.  1972. Early response of lungs to low levels of nitrogen
17            dioxide - light and electron microscopy. Arch. Environ. Health  24:160-179.
18
19    Stephens, R.J., Sloan,  M.F., Groth, D.G., Negi,  D.S., and Lunan, K.D.   1978.  Cytologic response of
20            postnatal rat lungs to O3 or NO2 exposure. Am. J. Pathol. 93:183-200.
21
22    Suzuki, A.K., Tsubone, H., and Kubota, K.  1982. Changes of gaseous exchange in the  lung of mice
23            acutely exposed to nitrogen dioxide. Toxicol. Letters 10:327-355.
24
25    Swedish National  Board  of Occupational Safety and Health.   1996.  Occupational  Exposure Limit
26            Values, Adopted 28th August 1996. p. 58.
27
28    Tabacova, S., Nikiforov, B., and Balabaeva, L. 1985. Postnatal effects of maternal exposure to
29            nitrogen dioxide. Neurobehav. Toxicol. Teratol. 7:785-789.
30
31    ten Berge, W.F., Zwart,  A., and Appelman, L.M.   1986. Concentration-time  mortality response
32            relationship of irritant and systemically acting vapours and gases.  J. Hazard. Mat.  13:301-
33            309.
34
35    Tse, R.L. and Bockman,  A.A.  1970. Nitrogen dioxide toxicity: report of four cases in firemen.
36            J.A.M.A. 212:1341-1344.
37
38    Tunnicliffe,  W.S.,  Burge, P.S., and Ayres, J.G.  1994. Effect of domestic concentrations of nitrogen
39            dioxide on airway responses to inhaled allergen in asthmatic patients. Lancet 344:1733-1736.
40
41    U.S. EPA. 1987a. U.S. Environmental Protection Agency. §51.151 Significant Harm Levels, p. 639.
42            Subpart H Prevention of Air Pollution Emergency Episodes.  40 CFR.
43
44    U.S. EPA. 1987b. U.S. Environmental Protection Agency. Technical Guidance for Hazards Analysis.
45            Emergency Planning for Extremely Hazardous Substances. U.S. EPA, FEMA,  and U.S. DOT,
46            Washington, DC.
47
48    U.S. EPA. 1990. U.S. Environmental Protection Agency. Health and Environmental Effects Document for
49            Nitrogen Dioxide. Office of Health and Environmental Assessment, U.S. EPA,  Cincinnati, OH.
50            133 pp.
51
52    U.S. EPA. 1993. U.S. Environmental Protection Agency. Air Quality Criteria for Oxides of Nitrogen, Vol.
53            I-III. Office of Research and Development, U.S. EPA, Research Triangle Park, NC.
54
55    U.S. EPA. 1995. U.S. Environmental Protection Agency. Review of the National Ambient Air Quality
56            Standards for Nitrogen Dioxide. Assessment of Scientific and Technical Information.  OAQPS
                                                    62

-------
      NITROGEN OXIDES                  NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1            Staff Paper. Office of Air Quality, U.S. EPA, Research Triangle Park, NC. 95 pp.
 2
 3    U.S. EPA. 1997. U.S. Environmental Protection Agency. §50.11 National Primary and Secondary Ambient
 4            Air Quality Standards for Nitrogen Dioxide, p. 8. 40 CFR.
 5
 6    Utell, M.J. 1989. Asthma and nitrogen dioxide: A review of the evidence. ASTM STP 1024, M.J. Utell
 7            and R. Frank, Eds., American Society for Testing and Materials, Philadelphia, pp. 218-223.
 8
 9    Vagaggini, B.,  Paggiaro, P.L., Giannini, D., Di Franco, A.,  Cianchette, S., Carnevali, S., Taccola, M,
10            Bacci,  E., Bancalari, L., Dente, F.L., and Giuntini, C.  1996. Effect of short-term NO2 exposure on
11            induced sputum in normal, asthmatic and COPD subjects. Eur. Respir. J. 9:1852-1857.
12
13    Vollmuth, T.A., Driscoll, K.E.,  and  Schlesinger, R.B.   1986.  Changes  in early alveolar particle
14            clearance due  to  single  and repeated nitrogen dioxide  exposures  in the rabbit. J.  Toxicol.
15            Environ. Health 19:255-266.
16
17    von Nieding, G., Krekeler, H., and Fuchs,  R.  1973. Studies of the  acute  effects of NO2 on  lung
18            function: influence on diffusion, perfusion and ventilation in the lungs. Int. Arch. Arbeitsmed.
19            31:61-72.
20
21    von Nieding, G. and Wagner, H.M.  1979. Effects of NO2 on chronic bronchitics. Environ. Health Perspect.
22            29:137-142.
23
24    von Nieding, G., Wagner, H.M., Krekeler, H., Lollgen, H., Fries, W., and Beuthan, A.  1979. Controlled
25            studies of human exposure to single and combined action of NO2, O3, and SO2. Int. Arch. Occup.
26            Environ. Health 43:195-210.
27
28    Yue, M.-x., Peng, R.-y., Wang, Z.-g., Wang, D.-w., Yang, Z.-h., and Yang, H.m.  2004.  Characteristics of
29            acute and chronic intoxication induced by rocket propellant nitrogen tetroxide.  Space Med.
30            Medical Eng. 17:117-120. (Chinese with English abstract; partially translated).
31
                                                     63

-------
   NITROGEN OXIDES            NAC/Interim (NO2)/Proposed (N2O4): 12/2008
i                APPENDIX A: Derivation of AEGL Values
2
                                   64

-------
     NITROGEN OXIDES
                                    NAC/Interim (NO2)/Proposed (N2O4): 12/2008
 1
 2
 3
 4
 5
 6
 7
 8
 9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
Key Study:

Toxicity endpoint:


Time scaling:

Uncertainty factors:

Modifying factor:

Calculations:
                    Derivation of AEGL-1 for Nitrogen Oxides
Kerretal., 1978, 1979

slight burning of the eyes, slight headache, chest tightness or
labored breathing with exercise in 7/13 asthmatics exposed to
0.5 ppm for 2 hours

Not applied

None

None

None; 0.50 ppm value applied across AEGL-1 exposure
durations
Appropriate chemical-specific data were not available for derivation of AEGL-1 values
for N2O4. NC>2 exists as an equilibrium mixture of NC>2 and ^64 but the dimer is not
important at ambient concentrations (U.S. EPA, 1993). When ^CMs released it
vaporizes and dissociates into NC>2.  Thus, the AEGL values were developed based on
data for NO2 and are considered applicable to all nitrogen oxides. Values for N2O4 in
units of ppm have been calculated on a molar basis as presented in the text.
                                            65

-------
     NITROGEN OXIDES
                                    NAC/Interim (NO2)/Proposed (N2O4): 12/2008
                                   Derivation of AEGL-2
 9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
     Key Studies:
                    Henschler et al., 1960
     Toxicity endpoints:   burning sensation in nose and chest, cough, dyspnea, sputum
                         production in normal volunteers exposed to 30 ppm for 2 hours
 8   Time scaling:
                    C3'5 x t = k; the value of n was calculated by ten Berge et al. (1986)
                    from the data of Hine et al. (1970).
Uncertainty factors:   3 for intraspecies variability

Modifying factor:    None

Appropriate chemical-specific data were not available for derivation of AEGL-2 values
for N2O4. NO2 exists as an equilibrium mixture of NO2 and N2O4 but the dimer is not
important at ambient concentrations (U.S. EPA, 1993). When N2C>4is released it
vaporizes and dissociates into NO2.  Thus, the AEGL values were developed based on
data for NO2 and are considered applicable to all nitrogen oxides. Values for N2O4 in
units of ppm have been calculated on a molar basis as presented in the text.
Calculations:
C3'5 x t = k
                    (30 ppm/sy-3 x 2 hours = k
                    6324.56 ppm-hours = k
10-minute AEGL-2:  C = (6324.56 ppm-hours/0.167 hours)
                    C = 20 ppm

30-minute AEGL-2:  C = (6324.56 ppm-hours/0.5 hour)173'5
                    C = 15 ppm
                                                      1/3.5
1-hour AEGL-2:
4-hour AEGL-2:
8-hour AEGL-2:
C = (6324.56 ppm-hours/1 hour)173'5
C = 12 ppm
C = (6324.56 ppm-hours/4 hours)
C = 8.2 ppm

C = (6324.56 ppm-hours/8 hours)
C = 6.7 ppm
                                                  1/3.5
                                                  1/3.5
                                            66

-------
     NITROGEN OXIDES               NAC/Interim (NO2)/Proposed (N2O4): 12/2008


 1                                 Derivation of AEGL-3
 2
 3   Key Studies:         Henry etal., 1969
 4
 5   Toxicity endpoint:    signs of marked irritation, but no deaths in monkeys exposed to 50
 6                       ppm for 2 hours
 7
 8   Time scaling:        C3'5 x t = k; the value of n was calculated by ten Berge et al. (1986)
 9                          from the data of Hine et al. (1970).
10
11   Uncertainty factors:  3 for intraspecies variability;  1 for interspecies variability
12
13   Modifying factor:    None
14
15   Appropriate chemical-specific data were not available for derivation of AEGL-3 values
16   for N2O4.  NO2 exists as an equilibrium mixture of NO2 and N2O4 but the dimer is not
17   important at ambient concentrations (U.S. EPA, 1993). As a result when N2O4 is released
18   it vaporizes and dissociates into NO2.  Thus, the AEGL values were developed based on
19   data for NO2 and are considered applicable to all nitrogen oxides. Values  for N2O4 in
20   units of ppm have been calculated on a molar basis as presented in the text.
21
22   Calculations:         C3'5 x t = k
23                       (50 ppm/3)3'5 x 2 hours = k
24                       37,801 ppm-hours = k
25
26   10-minute AEGL-3:  C = (37,801 ppm-hours/0.1667 hours)173'5
27                       C = 34 ppm
28
29   30-minute AEGL-3:  C = (37,801 ppm-hours/0.5 hours)173'5
30                       C = 25 ppm
31
32   1-hour AEGL-3:     C = (37,801 ppm-hours/1 hours)173'5
33                       C = 20 ppm
34
35   4-hour AEGL-3:     C = (37,801 ppm-hours/4 hours)173'5
36                       C =  14 ppm
37
38   8-hour AEGL-3:     C = (37,801 ppm-hours/8 hours)173'5
39                       C =  11 ppm
40
                                            67

-------
    NITROGEN OXIDES             NAC/Interim (NO2)/Proposed (N2O4): 12/2008

i     APPENDIX B: Derivation Summary for AEGL Values for Nitrogen
2                                 Oxides
                                    68

-------
NITROGEN OXIDES
                 NAC/Interim (NO2)/Proposed (N2O4): 12/2008
            AEGL-1 VALUES for Nitrogen Dioxide and Nitrogen Tetroxide
Chemical
NO2
N204
10 minute
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
30 minute
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
1 hour
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
4 hour
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
8 hour
0.94 mg/m3
(0.50 ppm)
0.94 mg/m3
(0.25 ppm)
 Key Reference:
Keir, H.D., Kulle, T.J., Mcllhany, M.L., and Swidersky, P.  1978.
Effects of nitrogen dioxide on pulmonary function in human subjects. An
environmental chamber study.  Report: ISS EPA/600/1-78/025; Order
no. PB-281 186,20pp.

Kerr, H.D., Kulle, T.J., Mcllhany, M.L., and Swidersky, P.  1979.
Effects of nitrogen dioxide on pulmonary function in human subjects:
An environmental chamber study. Environ. Research 19:392-404.
 Test Species/Strain/Number:  Human subjects; sex not given; 13 asthmatics with exercise
 Exposure Route/Concentrations/Durations:
    Inhalation:  0.5 ppm NO2 for 2 hours
 Effects:      slight burning of the eyes, slight headache, chest tightness, or labored
              breathing in 7/13 subjects
 Endpoint/Concentration/Rationale:
      Mild symptoms of discomfort in asthmatics.
 Uncertainty Factors/Rationale:
    Total uncertainty factor: none
      Interspecies: NA; human data used
      Intraspecies: 1 - asthmatics were used as the test population
 Modifying Factor: none
 Animal to Human Dosimetric Adjustment: not applicable
 Time Scaling: Extrapolation to time points was not conducted because adaptation to mild
              sensory irritation occurs. In addition, animal responses to NO2 exposure have
              demonstrated a much greater dependence upon concentration than upon time;
              therefore, extending the 2-hour concentration to 8 hours should not exacerbate
              the human response.
 Data Quality and Support for the AEGL Values: AEGL-1 values are considered conservative
 and should be protective of the toxic effects of NO2 outside those expected as defined under
 AEGL-1.
                                       69

-------
NITROGEN OXIDES
           NAC/Interim (NO2)/Proposed (N2O4): 12/2008
            AEGL-2 VALUES for Nitrogen Dioxide and Nitrogen Tetroxide
Chemical
NO2
N204
10 minute
38 mg/m3
(20 ppm)
38 mg/m3
(10 ppm)
30 minute
28 mg/m3
(15 ppm)
28 mg/m3
(7.6 ppm)
1 hour
23 mg/m3
(12 ppm)
23 mg/m3
(6.2 ppm)
4 hour
15 mg/m
(8.2 ppm)
15 mg/m
(4.1 ppm)
8 hour
13 mg/m
(6.7 ppm)
13 mg/m
(3.5 ppm)
 Key Reference:
Henschler, D., Stier, A., Beck, H., and Neuman, W. 1960. Odor
threshold of a few important irritant gasses (sulfur dioxide, ozone,
nitrogen dioxide) and observations in humans exposed to low
concentrations.  Archiv fur Gewerbepathologie und
Gewerbehygiene 17:547-570.
 Test Species/Strain/Number: human, healthy male, 10-14
 Exposure Route/Concentrations/Durations:  0.5-30 ppm for up to 2 hours
 Effects:
    0.5 ppm: metallic taste
    1.5 ppm: dryness of the throat
    4 ppm: sensation of constriction
    25 ppm: prickling of the nose
    30 ppm: burning sensation in nose and chest, cough, dyspnea, sputum production
 Endpoint/C oncentrati on/Rati onal e:
         Humans exposed to 30 ppm for 2 hours experienced
         pronounced irritation.  The point of departure is
         considered a threshold for AEGL-2 since the effects
         noted by the subjects would not impair the ability to
         escape and the effects were reversible after cessation of
         exposure.
 Uncertainty Factors/Rationale:
    Total uncertainty factor: 3
      Interspecies:        NA, human data used
      Intraspecies:        3 - mechanism of action of a direct acting irritant is not expected
                         to differ greatly among individuals
 Modifying Factor: not applicable
 Animal to Human Dosimetric Adjustment: not applicable
 Time Scaling: Cn x t = k where n = 3.5 (ten Berge et al., 1986)
 Data Quality and Support for the AEGL Values: AEGL-2 values should be protective of the
 toxic effects of NO2 outside those expected as defined under AEGL-2.  The values are
 supported by occupational monitoring data.
                                       70

-------
NITROGEN OXIDES
           NAC/Interim (NO2)/Proposed (N2O4): 12/2008
Chemical
NO2
N204
10 minute
64 mg/m
(34 ppm)
64 mg/m3
(17 ppm)
30 minute
47 mg/m3
(25 ppm)
47 mg/m3
(13 ppm)
1 hour
38 mg/m3
(20 ppm)
38 mg/m3
(10 ppm)
4 hour
26 mg/m3
(14 ppm)
26 mg/m3
(7.0 ppm)
8 hour
21 mg/m3
(11 ppm)
21 mg/m3
(5.7 ppm)
            AEGL-3 VALUES for Nitrogen Dioxide and Nitrogen Tetroxide
 Key Reference:
Henry, M.C., Ehrlich, R., and Blair, W.H.  1969.  Effect of
nitrogen dioxide on resistance of squirrel monkeys to Klebsiella
pneumonias infection. Arch. Environ. Health 18:580-587.
 Test Species/Strain/Number: monkeys, 2-6/group
 Exposure Route/Concentrations/Durations: Inhalation, 10, 15, 35, 50 ppm for 2 hours
 Effects:
      50 ppm: marked increase in respiratory rate and decrease in tidal volume, microscopic
                lesions in lung (determinate for AEGL-3)
      35 ppm: increase in respiratory rate and decrease in tidal volume, microscopic lesions in
                lung
      10 and 15 ppm: slight changes in lung function
 Endpoint/Concentration/Rationale:  50 ppm resulted in marked effects on lung function but
                                  no deaths
 Uncertainty Factors/Rationale:
    Total uncertainty factor: 3
      Interspecies:
      Intraspecies:
 1 - the endpoint in the monkey study is below the definition of
 AEGL-3, human data support AEGL-3 point of departure and
 derived values, the mechanism of action does not vary between
 species with the target at the alveoli, and due to the similarities of
 the respiratory tract between humans and monkeys
 3 - mechanism of action of a direct acting irritant is not expected
 to differ greatly among  individuals.
 Modifying Factor: not applicable
 Animal to Human Dosimetric Adjustment: not applicable
 Time Scaling: Cn x t = k where n = 3.5 (ten Berge et al., 1986)
 Data Quality and Support for the AEGL Values: The study is of high quality and the AEGL-3
 values are supported by human data.
                                       71

-------
   NITROGEN OXIDES            NAC/Interim (NO2)/Proposed (N2O4): 12/2008

i   APPENDIX C: Time Scaling Category Plot for Nitrogen Oxides
2
                                   72

-------
NITROGEN OXIDES
NAC/Interim (NO2)/Proposed (N2O4): 12/2008
              1000.0
                               Chemical Toxicity - TSD All Data
                                     Nitrogen Dioxide
                        60    120    180   240   300   360    420    480
                                        Minutes
Figure 1: Category plot of AEGL values and effects of nitrogen dioxide on humans and animals
Source


NAC/AEGL-
NAC/AEGL-
NAC/AEGL-
NAC/AEGL-
NAC/AEGL-

NAC/AEGL-2
NAC/AEGL-2
NAC/AEGL-2
NAC/AEGL-2
NAC/AEGL-2

NAC/AEGL-3
NAC/AEGL-3
NAC/AEGL-3
NAC/AEGL-3
NAC/AEGL-3

Norwood et al., 1966
Morley and Silk, 1970
Species




















human
human
Sex




















m

#
Exposures






















ppm


0.5
0.5
0.5
0.5
0.5

20
15
12
8.2
6.7

34
25
20
14
11

90.0000
30.0000
Minutes


10
30
60
240
480

10
30
60
240
480

10
30
60
240
480

40
40
Category


AEGL
AEGL
AEGL
AEGL
AEGL

AEGL
AEGL
AEGL
AEGL
AEGL

AEGL
AEGL
AEGL
AEGL
AEGL

2
1
                                           73

-------
NITROGEN OXIDES
NAC/Interim (NO2)/Proposed (N2O4): 12/2008
Henschleretal., 1960
multiple studies
Frampton et al, 1991
Linn and Hackney, 1983, 1984
von Nieding et al., 1979
Kleinman et al., 1983
Sackner et al., 1981
Kerretal., 1978
Roger etal., 1990
Roger etal., 1990

Mine et al., 1970
Mine et al., 1970
Mine et al., 1970
Mine etal., 1970

Henry etal., 1969
Mine et al., 1970
Carson et al., 1962
Carson et al., 1962
Carson et al., 1962
Mine et al., 1970

Henschler and Lutge, 1963
Bauer etal., 1985

Mine etal., 1970
Carson etal., 1962
Carson etal., 1962
Meulenbeltetal., 1992
Hidekazu and Fujio, 1981
Henschler and Lutke, 1963
Mine etal., 1970
Mine etal., 1970
human
human
human
human
human
human
human
human
human
human

dog
rat
mouse
rabbit

monkey
dog
rat
rat
rat
rat

human
human

guinea pig
rabbit
rat
rat
mouse
dog
guinea pig
mouse




































































30.0000
0.6000
1.5000
4.0000
5.0000
0.2000
1.0000
0.5
0.3000
0.6

75
100
100
75

50
20
190
90
72
20

20
0.3

50
315
115
200
40
40
20
20
120
180.0
180
75
120.0
120.0
240.0
120
110
75

240
240
240
60

120
1440
5
15
60
1440

120
240

60
15
60
10
720
360
1440
1440
1
0
0
0
1
0
0
1
1
0

PL
3
3
PL

2
1
2
2
2
1

1
1

PL
PL
PL
2
PL
0
1
1
For Category 0 = No effect, 1 = Discomfort, 2 = Disabling, PL = Partially Lethal, 3 = Lethal
                                         74

-------