National Toxicology Program

National Toxicology Program

Abstract for IMM87033 - para-nitrotoluene (CASRN 99-99-0)

The following abstract presents results of a study conducted by a contract laboratory for the National Toxicology Program (NTP). The findings have not been peer reviewed and were not evaluated in accordance with the levels of evidence criteria established by the NTP on March 2009 (see The findings and conclusions for this study should not be construed to represent the views of the NTP or the US Government.

The Immunotoxicity of Para-nitrotoluene
(CAS No. 99-99-0) in Female B6C3F1 Mice

NTP Report Number IMM87033


The mono-nitrotoluenes are synthesized by the nitration of toluene. Para-nitrotoluene (p-nitrotoluene) is a colorless solid at ambient temperature. The mono-nitrotoluenes are slightly soluble in aqueous solution but are readily soluble in organic solvents. The nitrotoluenes have been detected in air, water and soil.1, 2 Meta- and para-nitrotoluene have been identified by the National Toxicology Program (NTP) for detail study. P-nitrotoluene was the first mono-nitrotoluene to be investigated for its potential adverse actions on the immune system.

The objective of the studies reported here was to determine the potential immunotoxicity of p-nitrotoluene in mice exposed by the oral route. The studies were divided into three areas: the more standard toxicology studies provided the base against which to compare the immunology studies; the immunology studies provided data on specific cellular targets of the xenobiotic; and the host resistance studies were used for a global assessment of the ability of the exposed animal to resist pathogenic challenge.

Table ES-1 shows a summary of the standard toxicology studies. Mice exposed to p-nitrotoluene by gastric gavage gained body weight over the 2 week experimental period in a manner similar to the control groups. Of the selected organs weighed, the spleen was the only organ to show an increase in weight. The liver of mice exposed to 400 and 600 mg/kg showed mild to moderate swelling of the hepatocytes adjacent to the central veins. The hepatocyte swelling appeared to be reversible and there was no evidence of necrosis. The selected hematological and serum chemistries were unaffected by p-nitrotoluene exposure. Bone marrow cellularity and the number of CFU/M and CFU/GM were unaffected by p-nitrotoluene exposure.

Table ES-2 summarizes the immunology studies of mice exposed to p-nitrotoluene. P-nitrotoluene suppressed the IgM response to sRBC and the DHR response to KLH. There was a slight (13%) decrease in the percentage of T lymphocytes in the spleen. All of the other immune parameters were unaffected by exposure to p-nitrotoluene. These parameters included macrophage numbers and function, natural killer cell activity, response to mitogens and allogeneic cells, and serum complement and interferon levels.

Table ES-3 summarizes the host resistance studies in mice exposed to p-nitrotoluene. Exposure of mice to p-nitrotoluene decreased resistance to Listeria monocytogenes. Resistance to Streptococcus pneumoniae and Plasmodium yoelii were unaffected by exposure as was the resistance to the two tumor models, the PYB6 tumor and the B16F10 melanoma.

The three phases of these studies indicate that the liver and, to an equal or greater extent, the immune system are a target for toxicity of p-nitrotoluene. The decreased host resistance to Listeria monocytogenes can be attributed to the decrease in T lymphocytes and, as seen in the number and activity as expressed, to the decreased delayed hypersensitivity response to Keyhole limpet hemocyanin. The decrease in IgM response did not correlate with a host resistance model, most likely because no effect was seen on the IgG response.

Table ES-1
Parameter Results Maximum Effect DoseComments
Periodic Weights Increase in
weight gain
(weight gain)
600 mg/kg
Gross Pathology None
Histopathology Moderate to mild
swelling of hepatocytes
 400 mg/kg
Organ Weights
        Liver Increased 7% 600 mg/kg
        Spleen Increased 14% No significance
at high dose
Based on
trend analysis
        Lungs No effect
        Thymus No effect
        Kidney No effect
        Brain No effect
        RBC's No effect
        Retics Increased 29% No significance
at high dose
Based on
trend analysis
        Leukocytes No effect
        Leukocyte Diff
          Lymphocytes No effect
          Neutrophils No effect
Serum Chemistries
        SGPT No effect
        Albumin Increased 25% No significance
at high dose
Based on
trend analysis
        BUN No effect
        Glucose No effect
        Total Protein Increased 17% No significance
at high dose
Based on
trend analysis
Bone Marrow
        Cellularity No effect
        DNA Synthesis No effect
        Stem Cells
          CFU-GM No effect
          CFU-M No effect

Table ES-2
Parameter Results Maximum Effect Dose
T Cells Decreased 13% Yes
B Cells No effect
IgM AFC to SRBC Decreased 54% Yes
IgG AFC to SRBC No effect
DHR to KLH Decreased 31% Yes
Mitogens- Con A No effect
        - PHA No effect
        - LPS No effect
        - Medium No effect
MLR No effect
NK Cell Activity No effect
Serum Complement
      CH50 No effect
      C3 No effect
Macrophage (RES) No effect
Macrophage (Peritoneal)
      Cell # No effect
        Lymphocytes No effect
        Poly's No effect
        Monocytes No effect
      Phagocytosis Covaspheres No effect
      Phagocytosis CRBC No effect
      Adherence of CRBC No effect
        5'ND No effect
        LAP No effect
        PDE No effect
        ß Glucuronidase No effect

Table ES-3
MODEL Results Maximum Effect Dose Comments
Listeria monocytogenes Decreased resistance 41 % increase in
600 mg/kg
Streptococcus pneumoniae No effect
PYB6 No effect
B16F10 No effect
Plasmodium yoelii No effect

1 Pellizzari, E.D. (1978): Quantitation of Chlorinated Hydrocarbons in Previously Collected Air Samples. EPA 450/3-78-412.
2 Howard, P.H., Santodonato, J., Saxena, J., Malling, J., and Greninger, D. (1976): Investigation of Selected Potential Environmental Contaminants: Nitroaromatics. U.S. EPA, RTP, NC, p.1407 EPA 560/2-76-010.

Report Date: November 1987

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