United States
Environmental Protection
1=1 m m Agency
EPA/690/R-09/047F
Final
9-15-2009
Provisional Peer-Reviewed Toxicity Values for
Picramic Acid
(CASRN 96-91-3)
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268

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COMMONLY USED ABBREVIATIONS
BMD
Benchmark Dose
IRIS
Integrated Risk Information System
IUR
inhalation unit risk
LOAEL
lowest-observed-adverse-effect level
LOAELadj
LOAEL adjusted to continuous exposure duration
LOAELhec
LOAEL adjusted for dosimetric differences across species to a human
NOAEL
no-ob served-adverse-effect level
NOAELadj
NOAEL adjusted to continuous exposure duration
NOAELhec
NOAEL adjusted for dosimetric differences across species to a human
NOEL
no-ob served-effect level
OSF
oral slope factor
p-IUR
provisional inhalation unit risk
p-OSF
provisional oral slope factor
p-RfC
provisional inhalation reference concentration
p-RfD
provisional oral reference dose
RfC
inhalation reference concentration
RfD
oral reference dose
UF
uncertainty factor
UFa
animal to human uncertainty factor
UFC
composite uncertainty factor
UFd
incomplete to complete database uncertainty factor
UFh
interhuman uncertainty factor
UFl
LOAEL to NOAEL uncertainty factor
UFS
subchronic to chronic uncertainty factor
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
PICRAMIC ACID (CASRN 96-91-3)
Background
On December 5, 2003, the U.S. Environmental Protection Agency's (U.S. EPA) Office of
Superfund Remediation and Technology Innovation (OSRTI) revised its hierarchy of human
health toxicity values for Superfund risk assessments, establishing the following three tiers as the
new hierarchy:
1)	U.S. EPA's Integrated Risk Information System (IRIS).
2)	Provisional Peer-Reviewed Toxicity Values (PPRTVs) used in U.S. EPA's Superfund
Program.
3)	Other (peer-reviewed) toxicity values, including
~	Minimal Risk Levels produced by the Agency for Toxic Substances and Disease
Registry (ATSDR),
~	California Environmental Protection Agency (CalEPA) values, and
~	EPA Health Effects Assessment Summary Table (HEAST) values.
A PPRTV is defined as a toxicity value derived for use in the Superfund Program when
such a value is not available in U.S. EPA's IRIS. PPRTVs are developed according to a
Standard Operating Procedure (SOP) and are derived after a review of the relevant scientific
literature using the same methods, sources of data, and Agency guidance for value derivation
generally used by the U.S. EPA IRIS Program. All provisional toxicity values receive internal
review by two U.S. EPA scientists and external peer review by three independently selected
scientific experts. PPRTVs differ from IRIS values in that PPRTVs do not receive the
multiprogram consensus review provided for IRIS values. This is because IRIS values are
generally intended to be used in all U.S. EPA programs, while PPRTVs are developed
specifically for the Superfund Program.
Because new information becomes available and scientific methods improve over time,
PPRTVs are reviewed on a 5-year basis and updated into the active database. Once an IRIS
value for a specific chemical becomes available for Agency review, the analogous PPRTV for
that same chemical is retired. It should also be noted that some PPRTV documents conclude that
a PPRTV cannot be derived based on inadequate data.
Disclaimers
Users of this document should first check to see if any IRIS values exist for the chemical
of concern before proceeding to use a PPRTV. If no IRIS value is available, staff in the regional
Superfund and Resource Conservation and Recovery Act (RCRA) program offices are advised to
carefully review the information provided in this document to ensure that the PPRTVs used are
appropriate for the types of exposures and circumstances at the Superfund site or RCRA facility
in question. PPRTVs are periodically updated; therefore, users should ensure that the values
contained in the PPRTV are current at the time of use.
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It is important to remember that a provisional value alone tells very little about the
adverse effects of a chemical or the quality of evidence on which the value is based. Therefore,
users are strongly encouraged to read the entire PPRTV document and understand the strengths
and limitations of the derived provisional values. PPRTVs are developed by the U.S. EPA
Office of Research and Development's National Center for Environmental Assessment,
Superfund Health Risk Technical Support Center for OSRTI. Other U.S. EPA programs or
external parties who may choose of their own initiative to use these PPRTVs are advised that
Superfund resources will not generally be used to respond to challenges of PPRTVs used in a
context outside of the Superfund Program.
Questions Regarding PPRTVs
Questions regarding the contents of the PPRTVs and their appropriate use (e.g., on
chemicals not covered, or whether chemicals have pending IRIS toxicity values) may be directed
to the U.S. EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
INTRODUCTION
Picramic acid, or 4,6-dinitro-2-aminophenol, is used as a chemical intermediate in the
manufacture of azo dyes, an explosive initiator, an oxidation base for dyeing furs, a reagent for
albumin, and an acid-base indicator (Hazardous Substances Data Bank [HSDB], 2008). Figure 1
shows its chemical structure.
HO	NhL
0
Figure 1. Structure of Picramic Acid
There is no assessment of picramic acid (2-amino-4,6-dinitrophenol) on IRIS
(U.S. EPA, 2008), in the Drinking Water Standards and Health Advisories list (U.S. EPA, 2006)
or in the Health Effects Assessment Summary Tables (HEAST; U.S. EPA, 1997). The Chemical
Assessments and Related Activities (CARA) list (U.S. EPA, 1991a, 1994) includes Health and
Environmental Effects Profiles (HEEPs) for aminophenols (U.S. EPA, 1985) and selected
dinitrophenols (U.S. EPA, 1984); however, neither of these include data on picramic acid.
Similarly, the Agency for Toxic Substances and Disease Registry (ATSDR, 1995) prepared a
toxicological profile of dinitrophenols, but this document did not include picramic acid.
Occupational exposure limits for picramic acid have not been derived by the American
Conference for Governmental Industrial Hygienists (ACGIH, 2008), the National Institute for
Occupational Safety and Health (NIOSH, 2008), or the Occupational Safety and Health
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Administration (OSHA, 2008). Assessments for picramic acid have not been performed by the
National Toxicology Program (NTP, 2005, 2008), the California Environmental Protection
Agency (CalEPA, 2008a,b), the International Agency for Research on Cancer (IARC, 2008), or
the World Health Organization (WHO, 2008).
Literature searches were conducted from 1960s through November 2008 for studies
relevant to the derivation of provisional toxicity values for picramic acid. Databases searched
include MEDLINE, TOXLINE (with NTIS), BIOSIS, TSCATS/TSCATS2, CCRIS, DART,
GENETOX, HSDB, RTECS, Chemical Abstracts, and Current Contents (last 6 months). A
Cosmetic Ingredient Review Expert Panel Report (CIREP, 1992) was also consulted for relevant
information.
REVIEW OF PERTINENT DATA
Human Studies
No relevant data regarding the toxicity of picramic acid to humans following chronic or
subchronic oral or inhalation exposure were located.
Animal Studies
No relevant data regarding the toxicity of picramic acid to animals following chronic or
subchronic oral or inhalation exposure were located.
Other Studies
Acute Toxicity
The acute oral LD50 for picramic acid in male HA/ICR mice was 378 mg/kg
(CIREP, 1992). Few details of the study are reported in the CIREP (1992) review article: only
that 36 adult male mice were tested using a wide range of doses, and that the LD50 was
determined by probit analysis.
Robbins (1944) reported that picramic acid in the diet of chicks at a concentration of 1%
resulted in lesions (lens opacities or cataracts) similar to those observed with 2,4-dinitrophenol at
a concentration of 0.25%. No further information was provided on the experiments conducted
with picramic acid. In the experiments with 2,4-dinitrophenol, the compound was mixed with
feed and given ad libitum to chicks (0.05-0.25% in feed) and ducks (0.25% in feed) for
durations ranging from 3 to 31 days. Each day, the fowl were examined for lens opacities and
their temperatures and body weights were recorded. Individual birds were sacrificed at intervals
from 5 hours to 30 days after commencement of exposure, and their lenses were examined
microscopically. Lens opacities were observed within hours of the initial administration of
2,4-dinitrophenol at a dietary concentration of 0.25%.
Other Routes
In a brief report published in French in 1933, Casier (1933) reported that sodium
picramate exposure caused hyperthermia, sometimes resulting in death, when administered
intravenously to dogs or via intravenous or intraperitoneal injection to pigeons. The author
indicated that the hyperthermic effects were similar to those occurring with dinitronaphthol,
dinitrophenol, or dinitrocresol, but were less severe and lasted longer.
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Pugh and Stone (1968) measured bile secretion, bromosulphophthalein excretion, and
body temperature in anaesthetized male and female dogs (strain and number not reported)
exposed via intravenous injection to doses of 5-53 mg/kg picramic acid. Picramic acid exposure
resulted in a moderate increase in bile flow (mean 97% increase in bile volume relative to
controls) and a slight decrease in bromosulphophthalein excretion (2% less than controls), but it
did not affect body temperature. By comparison, 2,4-dinitrophenol at a dose of 5 mg/kg caused a
larger increase in bile volume (153% of controls), an increase in bromosulphophthalein excretion
(16%) higher than controls), and an increase in body temperature (3.9°C).
The effects of dermal exposure to oxidative hair dyes containing picramic acid or sodium
picramate (at concentrations from 0.01 to 0.1%>) were assessed in subchronic, chronic, and
teratogenicity studies. The dye was mixed with an equal volume of 6%> hydrogen peroxide and
applied twice weekly at a dose of 1 mL/kg. No effects on hematology, blood chemistry,
urinalysis, or gross or microscopic findings were observed in New Zealand white rabbits (6/sex)
exposed dermally to a hair dye containing 0.1%> sodium picramate for 13 weeks; slight
thickening of the skin was noted at the application site (Burnett et al., 1976). Teratogenicity
testing of the same dye (with 0.1%> sodium picramate) applied to the shaven skin of 20 mated
female CD rats on gestation days 1, 4, 7, 10, 13, 16, and 19 (at 2 mL/kg of the dye solution)
revealed no effects on numbers of corpora lutea, implantation sites, live fetuses, resorption sites,
or the incidence of visceral or skeletal malformations (Burnett et al., 1976). The carcinogenicity
of an oxidative hair dye containing 0.01%> picramic acid was evaluated in a skin-painting study
using Swiss-Webster mice (Jacobs et al., 1984, as cited in CIREP, 1992). The dye was mixed
with an equal volume of hydrogen peroxide and applied to the skin once a week for 20 months.
Treatment caused no hematology or urinalysis changes, and did not increase the incidence of
tumors compared with controls (Jacobs et al., 1984, as cited in CIREP, 1992). Citing
unpublished studies, CIREP (1992) reported that an aqueous solution of 2.5%> picramic acid did
not cause skin irritation in three rabbits (strain not reported), but a solution of 2%> picramic acid
(1:10, 1:100, and 1:1000 dilutions in distilled water) caused mild skin sensitization in guinea pigs
(4/15 guinea pigs treated with the 1:10 dilution of 2%> picramic acid), and an aqueous solution of
2.5% sodium picramate caused "mild conjunctival inflammation" as ocular irritation in three
albino rabbits.
Toxicokinetics
No information on the pharmacokinetics of picramic acid in humans or animals was
located in the available literature either via oral or inhalation route. Wyman et al. (1992)
reported that the primary urinary metabolite of picric acid administered orally to rats was
picramic acid, which represented 18.5%> of the administered dose. Picramic acid was one of two
urinary metabolites (along with picric acid) observed in a study of rats exposed subcutaneously
to trinitrophenylmethylnitramine (Myers and Spinnato, 2007). The study did not quantify the
excretion of either picric acid or picramic acid. A similar metabolism of
trinitrophenylmethylnitramine (Tetryl) to picric acid and picramic acid was also observed in a
rabbit (strain not stated) oral study by Zambrano and Mandovano (1956). The exact
quantification of the excretion of picric acid and picramic acid was not provided, but the
picramic acid was present in the urine of "practically all the rabbits treated." The study authors
also stated that picramic acid can be excreted in a conjugated form (sulfoconjugates).
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Genotoxicity
With or without metabolic activation, picramic acid yielded positive results for reverse
mutation in Salmonella typhimurium strains TA97, TA98, TA100, TA1535, TA1537, and
TA1538 (Zeiger et al., 1988; Litton Bionetics, Inc., 1979a, as cited in CIREP, 1992;
Wyman et al., 1979; CIREP, 1992; Zieger et al. 1988) but gave equivocal results in
Saccharomyces cerevisiae strain D4 (CIREP, 1992). Picramic acid was not mutagenic at the TK
locus in cultured L5178Y mouse lymphoma cells at concentrations that were moderately toxic,
although there was some evidence of weak mutagenic activity with extremely toxic treatments
(2-4% relative growth). This compound did not increase the frequency of sister chromatid
exchanges (SCE) in L5178Y mouse lymphoma cells (CIREP, 1992). Picramic acid did not
induce chromosomal aberrations in the bone marrow of mice in a subchronic group given oral
doses up to 75.6 mg/kg (CIREP, 1992). In addition, picramic acid gave negative results in a
dominant lethal assay using CD-I mice (CIREP, 1992).
FEASIBILITY OF DERIVING PROVISIONAL SUBCHRONIC AND CHRONIC
ORAL RfD VALUES FOR PICRAMIC ACID
Due to a lack of data, no chronic or subchronic RfDs are developed. However, the
Appendix of this document contains a Screening Value (a screening RfD) based on an analog
treatment, which may be useful in certain instances. Please see the attached Appendix for
details.
FEASIBILITY OF DERIVING PROVISIONAL SUBCHRONIC AND CHRONIC
INHALATION RfC VALUES FOR PICRAMIC ACID
There are no toxicity data from which to derive a provisional inhalation RfC (p-RfC) for
picramic acid. Furthermore, there are no inhalation toxicity data for any of the structural analogs
identified for picramic acid, precluding derivation of a chronic screening RfC.
PROVISIONAL CARCINOGENICITY ASSESSMENT
FOR PICRAMIC ACID
Weight-of-Evidence Descriptor
Under the 2005 Guidelines for Carcinogen Assessment (U.S. EPA, 2005), there is
"Inadequate Information to Assess [the] Carcinogenic Potential' of picramic acid. There are no
data with which to assess the potential carcinogenicity of picramic acid in humans or animals.
Genotoxicity testing of picramic acid yielded positive results for mutagenicity in S. typhimurium,
equivocal results for mutagenicity in S. cerevisiae, negative results for mutagenicity and SCE
induction in mouse lymphoma cells, and negative results for chromosomal aberration induction
in mice.
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Quantitative Estimates of Carcinogenic Risk
The lack of data on the carcinogenicity of picramic acid precludes the derivation of
quantitative estimates of risk for either oral (p-OSF) or inhalation (p-IUR) exposure.
REFERENCES
ACGIH (American Conference of Governmental Industrial Hygienists). 2008. Threshold limit
values for chemical substances and physical agents and biological exposure indices. Cincinnati,
OH.
ATSDR (Agency for Toxic Substances and Disease Registry). 1995. Toxicological Profile for
Dinitrophenols. U.S. Department of Health and Human Services, Public Health Service. Online.
http://www.atsdr.cdc. gov/toxprofiles/tp.asp?id=729&tid=132.
Burnett, C., E.I. Goldenthal, S.B. Harris et al. 1976. Teratology and percutaneous toxicity
studies on hair dyes. J. Toxicol. Environ. Health. 1:1027-1040.
CalEPA (California Environmental Protection Agency). 2008a. Office of Environmental Health
Hazard Assessment. Search Chronic RELs. Online.
http://www.oehha.ca.gov/air/chronic rets/index.html
CalEPA (California Environmental Protection Agency). 2008b. Office of Environmental Health
Hazard Assessment. Search Toxicity Criteria Database. Online.
http://www.oehha.ca.gov/risk/Chem.icalDB/index.asp.
Casier, H. 1933. Hyperthermia-producing action of sodium picramate. Comptes Rendus des
Seances de la Societe de Biologie et de Ses Filiales. 114:554-555.
CIREP (Cosmetic Ingredient Review Expert Panel). 1992. Final report on the safety assessment
of Sodium Picramate. J. Am. Coll. Toxicol. 11(4):447-464.
Dodds, E.C. and J.D. Robertson. 1933. The clinical applications of dinitro-o-cresol. Lancet.
2:1137-1139.
Harvey, D.G., P.L. Bidstrup and J.A. Bonnell. 1951. Poisoning by dinitro-ortho-cresol. Some
observations on the effects of dinitro-ortho-cresol administration by mouth to human volunteers.
Br. Med. J. 2:13-16.
Horner, M.D. 1942. Dinitrophenol and its relation to formation of cataracts. Arch. Ophthal.
27:1097.
HSDB (Hazardous Substances Data Bank). 2008. 4,6-Dinitro-2-aminophenol. National Library
of Medicine. Online. http://toxnet.nlm.nih.gov/cgi-bin/sis/htmlgen7HSDB.
IARC (International Agency for Research on Cancer). 2008. Search IARC Monographs.
Online, http://monographs.iarc.fr/.
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Jacobs, M.M., C.M. Burnett, A.J. Penicnak et al. 1984. Evaluation of the toxicity and
carcinogenicity of hair dyes in Swiss mice. Drug Chem Toxicol. 7(6):573-586.
Mahlen, S. 1938. Zur kenntnis der katarakta bei dintroorthokresolbehandlung. Acta Ophthal.
16:563-572.
Myers, S.R. and J. A. Spinnato. 2007. Tissue distribution and elimination of N-methyl-
N-2,4,6-tetranitroaniline (tetryl) in rats. Arch. Toxicol. 81(12):841—848.
NIOSH (National Institute for Occupational Safety and Health). 2008. NIOSH Pocket Guide to
Chemical Hazards. Index by CASRN. Online, http://www.cdc.gov/niosh/npe/.
NTP (National Toxicology Program). 2005. 11th Report on Carcinogens. U.S. Department of
Health and Human Services, Public Health Service, National Institutes of Health, Research
Triangle Park, NC.
NTP (National Toxicology Program). 2008. Testing status of agents at NTP. Online.
http://ntp.niehs.nih.gov/ntp/roc/tocl 1 .htm.
OSHA (Occupational Safety and Health Administration). 2008. OSHA Standard 1915.1000 for
Air Contaminants. Part Z, Toxic and Hazardous Substances. Online.
https://www.osha.gov/pls/oshaweb/owadisp.show document?p table STANDARDS&p id=999
2.
Plotz, M. 1936. Dinitro-ortho-creosol: A metabolic stimulator and its toxic side-actions. N.Y.
State J. Med. 36:266-268.
Pugh, P.M. and S.L. Stone. 1968. The effect of 2,4-dinitrophenol and related compounds on
bile secretion. J. Physiol. 198(l):39-49.
Robbins, B.H. 1944. Dinitrophenol cataract: Production in an experimental animal. J.
Pharmacol. Exp. Ther. 80:264-271.
U.S. EPA. 1984. Health and Environmental Effects Profile (HEEP) for Dinitrophenols
(selected). Prepared by the Environmental Criteria and Assessment Office, Cincinnati, OH for
the Office of Solid Waste and Emergency Response, Washington, DC.
U.S. EPA. 1985. Health and Environmental Effects Profile (HEEP) for Aminophenols.
Prepared by the Environmental Criteria and Assessment Office, Cincinnati, OH for the Office of
Solid Waste and Emergency Response, Washington, DC.
U.S. EPA. 1991a. Chemical Assessments and Related Activities (CARA). Office of Health and
Environmental Assessment, Washington, DC. April.
U.S. EPA. 1991b. Integrated Risk Information System (IRIS). IRIS Summary of
2,4-Dinitrophenol (CASRN 51-28-5). Office of Research and Development, National Center for
Environmental Assessment, Washington, DC. Online.
http ://www. epa. gov/NCE A/iri s/ sub st/0152. htm.
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U.S. EPA. 1994. Chemical Assessments and Related Activities (CARA). Office of Health and
Environmental Assessment, Washington, DC. December.
U.S. EPA. 1997. Health Effects Assessment Summary Tables. FY-1997 Update. Prepared by
the Office of Research and Development, National Center for Environmental Assessment,
Cincinnati OH for the Office of Emergency and Remedial Response, Washington, DC. July.
EPA/540/R-97/036. NTIS PB97-921199.
U.S. EPA. 2002. Provisional Peer-reviewed Toxicity Values for 4,6-Dinitro-o-cresol
(CASRN 534-52-1): Derivation of a Chronic Oral RfD. Superfund Health Risk Technical
Support Center, U.S. Environmental Protection Agency, Cincinnati, OH.
U.S. EPA. 2004. Provisional Peer-reviewed Toxicity Values for Trinitrophenylmethylnitramine
(Tetryl) (CASRN 479-45-8). Superfund Health Risk Technical Support Center,
U.S. Environmental Protection Agency, Cincinnati, OH.
U.S. EPA. 2005. Guidelines for Carcinogen Risk Assessment. Risk Assessment Forum,
National Center for Environmental Assessment, Washington, DC. EPA/630/P-03/001F. Online.
http ://www. epa. gov/cancerauidelines/.
U.S. EPA. 2006. 2006 Edition of the Drinking Water Standards and Health Advisories. Office
of Water, Washington, DC. EPA/822/R-06/013. Washington, DC. Online.
http://water.epa.gov/drink/standards/hascience.cfm.
U.S. EPA. 2008. Integrated Risk Information System (IRIS). Online. Office of Research and
Development, National Center for Environmental Assessment, Washington, DC. Online.
http ://www. epa. gov/iris/.
WHO (World Health Organization). 2008. Online Catalogs for the Environmental Health
Criteria Series. Online.
http://www.who.int/ipcs/publications/ehc/ehc alphabetical/en/index.html.
Wyman, J.F., H.E. Guard, W.D. Won et al. 1979. Converstion of 2,4,6-Trinitrophenol to a
Mutgen by Pseudomonas aeruginosa. Appl Environ Microbiol. 37(2):222-226.
Wyman, J.F., M.P. Serve, D.W. Hobson et al. 1992. Acute toxicity, distribution, and
metabolism of 2,4,6-trinitrophenol (picric acid) in Fischer 344 rats. J. Toxicol. Environ. Health.
37(2):313-327.
Zambrano, A. and S. Mandovano. 1956. Urinary excretion of picric acid, picramic acid, and of
sulfoconjugation products in experimental tetryl poisoning. Folia Med. 39:162-171. (Italian)
Zeiger, E., B. Anderson, S. Haworth et al. 1988. Salmonella mutagenicity tests IV. Results
from the testing of 300 chemicals. Environ. Mol. Mutagen. 1 l(Suppl 12): 1-158.
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APPENDIX. DERIVATION OF A SCREENING VALUE
FOR PICRAMIC ACID
For reasons noted in the main PPRTV document, it is inappropriate to derive provisional
toxicity values for picramic acid. However, information is available for this chemical which,
although insufficient to support derivation of a provisional toxicity value, under current
guidelines, may be of limited use to risk assessors. In such cases, the Superfund Health Risk
Technical Support Center summarizes available information in an Appendix and develops a
"screening value." Appendices receive the same level of internal and external scientific peer
review as the PPRTV documents to ensure their appropriateness within the limitations detailed in
the document. Users of screening toxicity values in an appendix to a PPRTV assessment should
understand that there is considerably more uncertainty associated with the derivation of an
appendix screening toxicity value than for a value presented in the body of the assessment.
Questions or concerns about the appropriate use of screening values should be directed to the
Superfund Health Risk Technical Support Center.
Identification of Structural Analogs
Only compounds that have toxicity values available on IRIS, the current PPRTV status
table, or the HEAST (U.S. EPA, 1997) were considered as potential structural analogs to
picramic acid. The ChemlDplus Similarity Search function was used both as a tool to identify
candidate analogs and as a quantitative measure of structural similarity by which to select the
final analog to represent the subject chemical. The approach used to select potential analogs
consisted of two steps:
1)	Using available toxicokinetic information, upstream and/or downstream metabolites of
picramic acid in mammalian systems were identified and considered as potential analogs
if they had toxicity data from the pertinent sources (IRIS, PPRTV, HEAST).
2)	Using the ChemlDplus Similarity Search function, compounds structurally similar to
picramic acid were identified. Beginning with the highest degree of similarity,
compounds with structural similarity scores of at least 85% were identified, and IRIS, the
current PPRTV status table, and the HEAST were searched for the CAS registry numbers
and names of these compounds. The similarity score was later relaxed to at least 60% to
include any potential analogs.
Applying this approach, three potential analogs were identified for picramic acid. The
first candidate analog, trinitrophenylmethylnitramine (also known as tetryl or nitramine), was
identified because picramic acid is a metabolite of this compound. Picramic acid was one of two
urinary metabolites (along with picric acid) observed in a study in rats exposed subcutaneously
to trinitrophenylmethylnitramine (Myers and Spinnato, 2007). While picramic acid is also a
metabolite of picric acid (Wyman et al., 1992), picric acid does not have a toxicity value
available from IRIS, the PPRTV database, or the HEAST. The search of structural analogs by
the ChemlDplus similarity score identified trinitrophenylmethylnitramine (Tetryl),
4,6-dinitro-o-cresol (DNOC), and 2,4-dinitrophenol (2,4-DNP) as candidate analogs (see
Tables A-l and A-2).
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Table A-l. Physical-Chemical Properties of Picramic Acid and Candidate Analogs3

Picramic Acid
(4,6-Dinitro-
2-aminophenol)
T rinitrophenylmethylnitramine
(Tetryl or Nitramine)
4,6-Dinitro-o-cresol
(DNOC)
2,4 Dinitrophenol
(2,4-DNP)
Structure
HO NH,
N+— 0"
//
0
+-°"
^ N CH,
1 1 ,°~
A"
O — N+— O-
1 II
O" 0
OH O"
H3CV^LAo
0 0
II II
^ N+^
CASRN
96-91-3
479-45-8
534-52-1
51-28-5
Molecular formula
C6-H5-N3-05
C7-H5-N5-08
C7-H6-N2-05
ch4n2o5
Molecular weight
199.122
287.15
198.133
184.11
ChemID Plus Similarity Score (%)
100
62.68
88.59
85.47
Melting point (°C)
169
131.5
86.6
115.5
Boiling point (°C)
-
-
378
-
Vapor Pressure (mm Hg at 25°C)
4.16 x 10"7
5.66 x 10"8
1.06 x 10"4
3.9 x 10"4 at 20°C
Henry's Law Constant
(atm-m3/mole at 25 °C)
9.75 x 10"12
2.71 x 10"9
1.4 x 10"6
8.6 x 10"8 at 20°C
Water solubility (mg/L)
1400 at 22°C
74
198 at 20°C
2790 at 20°C
Log Kow
0.93
1.64
2.12
1.67
pKa
1
-
4.31
4.09 at 25°C
aData reported by ChemlDplus
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Table A-2. Comparison of Available Toxicity Data for Picramic Acid and Candidate Analogs3

Picramic Acid
(4,6-Dinitro-2-aminophenol)
T rinitrophenylmethylnitramine
(Tetryl or Nitr amine)
4,6-Dinitro-o-cresol
(DNOC)
2,4 Dinitrophenol
(2,4-DNP)
Structure
HO NH2
N+— 0"
//
0
+-°"
T r- ,*¦
0 — N+— 0"
1 l<
0" 0
OH 0"
V
"0 ^ 0
0 0
II II
^ wN+-v
CASRN
96-91-3
479-45-8
534-52-1
51-28-5
ChemID Plus Similarity
Score (%)
100
62.68
88.59
85.47
Human LDLOorLCLO
-
-
29 mg/kg (unreported route);
500 mg/kg (dermal)
36 mg/kg (oral)
Human TDL0 or TCL0
(Route, Effect)


7.5 mg/kg
(oral; sleepiness, headache)
1 mg/m3
(inlialation; CNS effects, cardiac and
gastrointestinal changes)

Oral LD50 in Mice (mg/kg)
378b
5000 (LDlq)
21
45
Intravenous LD50 in Dogs
(mg/kg)
150 (LDlq)
-
15 (LDlq)
15 (LDlq)
Subcutaneous LD50 in Rat
(mg/kg)
2100 (LDlq)
-
25.6
25
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Table A-2. Comparison of Available Toxicity Data for Picramic Acid and Candidate Analogs3

Picramic Acid
(4,6-Dinitro-2-aminophenol)
T rinitrophenylmethylnitramine
(Tetryl or Nitramine)
4,6-Dinitro-o-cresol
(DNOC)
2,4 Dinitrophenol
(2,4-DNP)
Chronic Oral RfD
(mg/kg-day)
Critical Effect
Source

4 x 10"3
Methemoglobin and serum
chemistry changes indicative of
hepatotoxicity in rats
U.S. EPA, 2004 (PPRTV)
1 x 10"4
Discolored conjunctivae, elevated
body temperature and basal
metabolic rate, perspiration, fatigue
in humans
U.S. EPA, 2002 (PPRTV)
1 x 10"4
Cataracts in humans
Derived in this Appendix with a
LOAEL of 1.2 mg/kg-day (Mahlen,
1938) and a composite UF of 10,000
2 x 10"3
Cataracts in humans
U.S. EPA, 1991b (IRIS)
aData reported by ChemlDplus except where noted
bCIREP (1992)
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Selection of Surrogate for Picramic Acid
The selection of a surrogate for picramic acid from among the three candidate analogs
took into consideration the physical-chemical properties of the compounds and available
information on their toxicity. The physical-chemical properties and available toxicity
information on picramic acid were compared with corresponding information on the three
candidate analogs in Tables A-l and A-2 below. The comparisons among the physical-chemical
properties of picramic acid and its candidate analogs suggest differences in properties that may
be important determinants of the compounds' behavior in biological systems. Specifically, water
solubility, Henry's Law constants, and logKow values differed widely among the analogs;
consequently, these data show that surrogate selection based on the physiochemical properties
alone may not be appropriate. The comparison of acute lethality information suggests that the
candidate analogs are either much less acutely lethal (Tetryl) or much more acutely lethal
(DNOC and 2,4-DNP). As Tetryl is less acutely lethal than picramic acid, and, therefore, could
potentially underestimate the toxicity of picramic acid in the long term, this compound was not
considered further as the surrogate for picramic acid. Furthermore, Tetryl was also excluded
based on lack of similar critical effect(s) to picramic acid, DNOC, and 2,4-DNP (see Table A-2).
There is limited direct information on potential target(s) or critical effect(s) of picramic acid
toxicity. Robbins (1944) reported that picramic acid administered in the diet of chicks resulted
in cataracts similar to those observed with 2,4-dinitrophenol. Therefore, it is likely that picramic
acid can cause cataract formation in humans as well as in chickens (chemical similarity between
picramic acid and 2,4-dinitrophenol is at least 85%; see Table A-l).
Comparison of DNOC and 2,4-DNP
A summary of the critical study is excerpted from the U.S. EPA (1991b) IRIS record for
2,4-DNP and reproduced here for the reader's convenience.
Over 100 anecdotal cases of cataracts resulting from therapeutic use of
2,4-dinitrophenol were reviewed. The length of time and amount of drug taken
varied among the population. It was estimated that over 1% of the population
administered 2,4-dinitrophenol developed cataracts. Data did not allow for
calculation of a NOEL; cataracts were observed in patients receiving as little as
2 mg/kg/day, the lower range of the recommended therapeutic dose.
Similarly, a summary of the critical studies is excerpted from the U.S. EPA (2002)
PPRTV document for DNOC and reproduced here for the reader's convenience.
The human studies, particularly Harvey et al. (1951), Dodds and Robertson
(1933), Plotz (1936) and Mahlen (1938), indicate that dosages of 0.35 to
~1 mg/kg-day constitute a LOAEL for humans ingesting DNOC for up to 1 year.
Critical effects include elevation of BMR and body temperature, excessive
perspiration, fatigue, discoloration of conjunctivae, and cataract formation. The
low end of the LOAEL range, 0.35 mg/kg-day (Plotz, 1936), is chosen as the basis
for the calculation.
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Even though a LOAEL of 0.35 mg/kg-day (Plotz, 1936) was originally chosen as the
most sensitive endpoint (not based on formation of cataracts in humans) and, therefore, as the
point of departure (POD) for DNOC in the U.S. EPA (2002) PPRTV, an alternate LOAEL of
1.2 mg/kg-day based on cataracts in humans (Mahlen, 1938) is specifically selected for the
derivation of a chronic screening RfD in this Appendix. Details of the Mahlen (1938) study is
summarized as the follows:
Mahlen (1938) reported on 73 patients in Sweden who had received DNOC at
weight-reduction dosages (~1 mg/kg-day) during a clinical study (DNOC was taken as a dieting
drug). Durations of treatment for these patients were not specified. After some time on
treatment, a nonjaundice yellowing of the skin and sclera were observed; however, reduction of
the dosage eliminated this symptom (reversible). Six of the patients were examined for cataracts
by other physicians and were found to be negative. The remaining 67 patients were examined by
the study author. Of these 67, 11 were eliminated from consideration because they had senile
cataracts (not treatment-related), had other conditions such as diabetes, or, in the case of one
patient, were not yet on DNOC treatment at the time of the examinations. Of the remaining
56 patients, 1 had bilateral cataracts attributable to DNOC. The affected patient was a 31-year
old woman had taken 83 mg/day (1.2 mg/kg-day based on her body weight of -69 kg at the time
of observation) of DNOC for 56 days and developed cataracts about 8 months later.
Mahlen (1938) also describes six other cases of cataracts associated with ingestion of
DNOC in Sweden. These cases were identified through a questionnaire sent by the Swedish
Board of Health to Swedish ophthalmologists. In all six cases, the patients were women (ages
ranged from 34-57 years), and their daily doses of DNOC ranged from 50-125 mg/day.
Although body weights were not specified, assuming body weights of 60 kg would result in
doses of 0.8-2.1 mg/kg-day. Durations of treatment ranged from 5-12 months, with occasional
additional treatment periods after 1-2 months without treatment. Symptoms and diagnosis of
cataracts occurred 2-10 months after the end of treatment. Based on the specific clinical case
with reported body weight (69 kg) and daily dose (83 mg/day), a LOAEL of 1.2 mg/kg-day is
identified for DNOC-induced cataract development.
As a result, this alternate endpoint and its associated LOAEL are deemed more
appropriate for DNOC in the limited context of serving as a potential surrogate for picramic acid.
The LOAEL of 1.2 mg/kg-day in a subchronic human study (Mahlen, 1938;
U.S. EPA, 2002) for DNOC and the LOAEL of 2 mg/kg-day in a subchronic human study
(Horner, 1942; U.S. EPA, 1991b) for 2,4-DNP both were based on formation of cataracts in
humans as the critical effects, suggesting that cataract formation may be a common critical effect
for the dinitrophenols (both DNOC and 2,4-DNP have a common core structure of
dinitrophenol). Therefore, cataract formation in humans is postulated to be the critical effect for
picramic acid because picramic acid belongs to this dinitrophenol chemical class.
For 2,4-DNP, U.S. EPA (1991b) derived a chronic RfD of 2 x 10"3 mg/kg-day based on
the LOAEL of 2 mg/kg-day as the POD and applied a composite UF of 1,000 (see U.S. EPA,
1991b for details). The composite UF includes a UFH of 10 for intraspecies variation, a UFS of
10 for extrapolation from subchronic-to-chronic duration, and a UFl of 10 for extrapolation from
LOAEL to NOAEL.
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Screening Chronic RfD
Although the U.S. EPA (2002) derived a p-RfD of 1 x 10"4 mg/kg-day for DNOC based
on the LOAEL of 0.35 mg/kg-day and a composite UF of 3,000 (see U.S. EPA, 2002 for details),
an alternate chronic p-RfD (as a potential surrogate for picramic acid), based on the LOAEL of
1.2 mg/kg-day1 for formation of cataracts in humans (Mahlen, 1938), is derived specifically for
this Appendix as follows:
Screening Chronic RfD = LOAEL UF
= 1.2 mg/kg-day ^ 10,000
= 0.0001 or 1 x 10"4 mg/kg-day
The composite UF of 10,000 for picramic acid (not for DNOC) is composed of the following:
•	UFr: A factor of 10 is applied for extrapolation to a potentially susceptible human
subpopulation because the data for evaluating susceptible human response are
insufficient.
•	UFl: A factor of 10 is applied for extrapolation from LOAEL to NOAEL because
the data for identifying a NOAEL are insufficient.
•	UFd: A factor of 10 is applied for database inadequacies—including lack of a
multigeneration reproduction study and a developmental study.
•	UFs: A factor of 10 is applied to derive the chronic RfD because of an adjustment
for exposure from sub chronic-to-chronic duration.
Notably, this composite UF is based on lack of toxicity information for picramic acid
(using default approach for each UF)—not for DNOC. This alternate p-RfD of
1 x 10"4 mg/kg-day is only applicable in this Appendix for comparison of common critical effect
(cataract formation) across a specific chemical class (dinitrophenols), and it should not be used
for DNOC by itself or elsewhere. In the future, if newer studies suggest that picramic acid could
cause similar responses (i.e., headache, excessive perspiration, and fatigue, elevated BMR, body
temperature, and greenish coloration of sclera) as shown for DNOC in Plotz (1936), then the
critical effect and POD of picramic acid, and the composite UF could be changed accordingly (as
stated in the U.S. EPA, 2002 document or the updated PPRTV for DNOC).
Both DNOC and 2,4-DNP are suitable candidates as the appropriate surrogate for
picramic acid because they have the same critical effect and target organ (cataracts; eyes) and
comparable acute lethality data. DNOC differs from picramic acid by only one functional group,
a methyl group instead of amino group, and 2,4-DNP lacks this additional functional group.
DNOC is, therefore, considered
1 A molecular weight (MW) adjustment is only necessary when a mechanism of action is elucidated and when
molecular targets have been identified. It is a common misconception that a MW adjustment must be applied
indiscriminately for the use of any surrogate approach when there is no mechanistic information (down to the
molecular level). This misconception stems from the practice of pharmaceuticals when one applies a
structure-activitiy relationship (SAR) to a series of congeners with a common and elucidated molecular mechanism
(e.g. tamoxifen vs. substituted tamoxifens binding to estrogen receptor alpha [ERa]). In general, a consideration of
the MW adjustment is not necessary if the MW of potential surrogate is less than a factor of 2 in comparison to the
MW of chemical of concern. If the MWs of potential surrogates are significantly higher (>2-fold) than the MW of
chemical of concern, then a common mechanism of action should be elucidated prior to the molecular-weight
adjustment.
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structurally more similar to picramic acid than 2,4-DNP. Based on the ChemlDplus similarity
score (88.59% for DNOC vs. 85.47% for 2,4-DNP; see Table A-2) and available RfDs (1 x 10"4
for DNOC vs. 2 x 10"3 for 2,4-DNP), DNOC was selected as the surrogate for picramic acid.
For picramic acid, the chronic p-RfD of 1 x 10"4 mg/kg-day, derived in this Appendix and
based on formation of cataracts in a subchronic human study (Mahlen, 1938), is recommended
only as a chronic screening RfD based on the surrogate analysis and structure-activity
relationship presented here. This alternate chronic p-RfD (a toxicity-based exposure level that is
specific to the target organ/effect of interest) uses the LOAEL of 1.2 mg/kg-day and employs a
composite UF of 10,000 (10 for intraspecies extrapolation, 10 for LOAEL-to-NOAEL
extrapolation, 10 for subchronic-to-chronic exposure duration extrapolation, and 10 for database
inadequacies) because data for evaluating developmental/reproductive toxicity for picramic acid
are inadequate.
Confidence in the critical study (low) and database (low) is the same as stated in the
PPRTV for DNOC (see U.S. EPA, 2002). Confidence in the overall surrogate approach is high
because the structural similarity is reasonably high (-89%) between picramic acid and DNOC
and because of the common target organ (eyes) and toxic effect (cataracts). However, due to the
high inherent uncertainty in the overall surrogate approach and low confidence in the critical
study and database, confidence in the screening chronic p-RfD is low.
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