A P E R E S E A R C H C O U N C I L

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Statement on EPA Estrogenic Screening Results

for Nonylphenol and Nonylphenol Ethoxylates 

July 14, 2010 – EPA recently released results from the first round of the Agency’s 

toxicity testing on oil dispersant products, which showed that none of the eight 

dispersants tested, including those that contain nonylphenol ethoxylate (NPE) surfactants, 

displayed biologically significant endocrine disrupting activity. 1 2 

EPA used in vitro reporter gene assays, which are rapid lab-bench tests conducted in 

specially developed cell lines. The assay results determine, at a screening level, whether 

test compounds are capable of binding with hormone receptors and, if so, how their 

binding potency relates to natural human hormones, such as the estrogen, 17-estradiol.

In addition to screening the dispersant products, EPA also conducted limited testing 

directly on NPEs and two forms of nonylphenol (NP), branched and linear. Branched NP 

can be an intermediate formed during the biodegradation of commercial NPEs in the 

environment. Linear NP is laboratory chemical that is not used in commerce and is not a 

degradant found in the environment. 

In the EPA screening tests the estrogenic activity for branched NP ranged from 250 to 

80,000-fold less potent than 17-estradiol. Results previously reported in the published

literature found branched NP to be one thousand to one million-fold less potent than 17-

estradiol, depending on which assay was used. Short-chain NPE degradants (NPE1 and 

NPE2) have been found to be less estrogenic than NP, while longer chain NPE 

(NPEn≥4), which are most commonly found in commercial products, appear to have little 

to no estrogenic activity.

3 4 5 6 7 8 9 10 These results are from a variety of in vitro tests that 

measure biomarkers of exposure to estrogens (e.g., vitellogenin induction in fish liver 

cells) and in vivo tests, which measured estrogen mediated changes in living fish or 

rodents. EPA’s screening results appear to be fairly consistent with these previously 

reported results. 

Across all aquatic organisms, the weight-of-evidence shows that commercial NPEs (e.g., 

NPE9) are much less toxic than NP. 11,12 In 2006, EPA established Water Quality 

Criteria (WQC) for NP, the most toxic of the NPE environmental degradants. 13 WQC 

are surface water concentrations that, when met, will protect aquatic life. EPA develops 

WQC based on aquatic toxicity studies that measure adverse effects, such as growth, 

survival, and reproduction. 14 These types of effects can have broader impacts on species, 

such as population reductions. 
2

EPA’s WQC document for NP states: 

“Whole organism endpoints such as reproductive and growth effects are used to 

derive aquatic life ambient water quality criteria for nonylphenol. To the extent 

that such endpoints reflect the integration of molecular, biochemical and tissuelevel effects at the whole organism level, the nonylphenol criteria address the 

estrogenicity of nonylphenol.” 15

It is important to remember that the results of an in vitro assay are not directly relevant to 

exposures and effects in living organisms; bathing cells in a chemical solution is not the 

same as exposing living fish or other aquatic organisms in the lab or out in the 

environment. However, it is interesting to note that EPA’s in vitro estrogenic screening 

results for NP continue to support EPA’s chronic WQC for NP with effect concentrations 

that are 7 to 23 times greater than the chronic freshwater (6.6 μg/L) criteria and 28 to 88 

times greater than the chronic saltwater criteria (1.7 μg/L ).16

The US EPA WQC for NP were based on study results on ecological effects of this 

compound available through 2005. Studies published since that time continue to support 

the US EPA WQC.17 

EPA’s WQC for NP, in conjunction with the extensive body of data available for NPE, 

provide EPA with a more than adequate basis to assess the risk of these compounds in the 

aquatic environment. 

Contact Information: 

Barbara S. Losey

Deputy Director

info@aperc.org

(202) 419-1500

*****

The mission of the APE Research Council, which is composed of manufacturers, processors and raw 

material suppliers of alkylphenols (AP) and alkylphenol derivatives (e.g., alkylphenol ethoxylates (APE)), 

is to promote the safe use of AP and AP derivatives through research, product stewardship and outreach 

efforts, within the framework of responsible chemical management. For more information about AP and 

APE go to www.aperc.org. 

 

1

U.S. Environmental Protection Agency (US EPA). (Accessed 2010, June 30). Statement: EPA’s Toxicity Testing 

of Dispersants. http://www.epa.gov/bpspill/dispersants-testing.html

2U.S. Environmental Protection Agency (US EPA) Office of Research and Development (ORD). (2010, June 30). 

Analysis of Eight Oil Spill Dispersants Using In Vitro Tests for Endocrine and Other Biological Activity. 

http://www.epa.gov/bpspill/reports/EPADispersantInVitroReport30june2010FINALx.pdf
3

 

3

Routledge, E.J., & Sumpter, J.P. (1996). Estrogenic activity of surfactants and some of their degradation products 

assessed using a recombinant yeast screen. Environmental Toxicology and Chemistry, 15 (3), 241-248. 

4

Lee, P.C., & Lee, W. (1996). In vivo estrogenic action of nonylphenol in immature female rats. Bulletin of 

Environmental Contamination and Toxicology, 57, 341-348. 

5

Islinger, M., Pawlowski, S., Hollert, H., Völkl, A., & Braunbeck, T. (1999). Measurement of vitellogenin-mRNA 

expression in primary cultures of rainbow trout hepatocytes in a non-radioactive dot blot/RNAse protection-assay. 

The Science of the Total Environment, 233, 109-122. 

6

Jobling, S., & Sumpter J.P. (1993). Detergent components in sewage effluent are weakly oestrogenic to fish: An in 

vitro study using rainbow trout (Oncorhynchus mykiss) hepatocyte. Aquatic Toxicology, 27, 361-372. 

7 Dussault, E.B., Sherry, J.P., Lee, H.B., Burnison, B.K., Bennie, D.T., & Servos, M.R. (2005). In vivo estrogenicity 

of nonylphenol and its ethoxylates in the Canadian environment. Human and Ecological Risk Assessment, 11 (2), 

353–364

8 Balch, G., & Metcalfe, C. (2006). Developmental effects in Japanese medaka (Oryzias latipes) exposed to 

nonylphenol ethoxylates and their degradation products. Chemosphere, 62 (8), 1214-1223. 

9 Metcalfe, C.D., Metcalfe, T.L., Kiparissis, Y., Koenig, B.G., Khan, C., Hughes, R.J., Croley, T.R., March, R.E., & 

Potter, T. (2001). Estrogenic potency of chemicals detected in sewage treatment plant effluents as determined by in 

vivo assays with japanese medaka (Oryzias latipes). Environmental Toxicology and Chemistry, 20, 297–308. 

10 US Environmental Protection Agency (US EPA). (2006, February 23). Notice of availability of final aquatic life 

ambient water quality criteria for nonylphenol. Federal Register, 71 (36), 9337-9339. http://www.epa.gov/EPAWATER/2006/February/Day-23/w2558.htm.

11 Staples, C.A., Weeks, J., Hall, J.F., & Naylor, C.G. (1998). Evaluation of aquatic toxicity and bioaccumulation of 

C8- and C9-alkylphenol ethoxylates. Environmental Toxicology and Chemistry, 17, 2470-2480.

12 Staples, C., Mihaich, E., Carbone, J., Woodburn, K., & Klečka, G. (2004). A weight of evidence analysis of the 

chronic ecotoxicity of nonylphenol ethoxylates, nonylphenol ether carboxylates, and nonylphenol. Human and 

Ecological Risk Assessment, 10 (6), 999-1017.

13US EPA. (2006, February 23).

14 US EPA. (2006, February 23). 

15 US Environmental Protection Agency (US EPA). (2005). Aquatic life ambient water quality criteria -

nonylphenol. Report 822-R-05-005. US Environmental Protection Agency, Washington, DC, USA.

http://www.epa.gov/waterscience/criteria/nonylphenol/final-doc.pdf

16 In-vitro test results are expressed as AC50 or EC50, the concentration associated with the activity at the 50th 

percent point on the concentration response curve. Based on the concentration-response curves obtained by EPA 

using the Attagene, Inc. ER assay, the AC50 values for branched NP were 0.68 and 0.61 M for the ER trans and 

ER cis assays, respectively. Using a molecular weight of 220 g/mol for 4-br-NP, concentrations of 134 to 150 

g/L are calculated. Based on the results in EPA’s ER Agonist Assay T47D-KBluc, the log EC50 for branched NP

was reported as -6.657 M. Using a molecular weight of 220 g/mol for 4-br-NP, concentrations of 48.46 g/L are 

calculated.

17 Coady, K., Staples, C. Losey, B., and Klecka, G. (2009). A Hazard Assessment of Aggregate Exposure to 

Nonylphenol and Nonylphenol Mono- and Di-ethoxylates in the Aquatic Environment. Human and Ecological Risk 

Assessment. In press