/

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON D.C., 20460

OFFICE OF

CHEMICAL SAfETY AND

POLLIJTK)N PREVENTION

FROM:

THROUGH:

pc : 079701

DP Barcode: D374490

MEMORANDUM

SUBJECf: Registration Review - Problem Formulation for d-Limonene

(CAS Reg. No. 5989-27-5)

Jim Goodyear, Ph.D., Biologist f)~ j ;l. CJ J "

Silvia Term es, Ph.D., Chemist 50 f...-'-'y............

Daoa Spatz, Braoch Chief ~-::: ""- L 6/, \JIIU

E

Enviro

nvironm

nmental

ental Ri

Fa

sk

te and

Branch

Effec

3

t

~~" s Division

TO: Russell Wasem, Chemical Review Manager

Risk Management and Implementation Branch 1

Pesticide Re-evaluation Division

The Environmental Fate and Effects Division (EFED) has completed the preliminary prob lem

formulation (attached) for the ecological risk, environmental fate, endangered species, and

drinking water assessments to be conducted as part ofthe Registration Review of d-limonene.

The problem Cannulation draws information from both open literature and from studies

submitted by the technical registrants in response to data requirements. This document is

intended to provide an overview ofwhat is currently known regarding the environmental fate and

ecological effec ts associated with d-Iimonene and outlines uncertainties regarding attributes of

the parent compound and its transformation products. It describes the prelimi nary ecological risk

hypothesis and the processes that will be used during the completion of drinking water and

eco logical risk assessments in support ofregistration review.
1

REGISTRATION REVIEW

PROBLEM FORMULATION FOR: 

Limonene (d-Limonene) 

 PC Code 079701; CAS Reg No.5989-27-5 

 

 

PREPARED BY: 

Jim Goodyear, Ph.D., Biologist 

Silvia Termes, Ph.D., Chemist 

APPROVED BY: 

Dana Spatz, Branch Chief 

Environmental Risk Branch 3 

Environmental Fate and Effects Division 

Office of Pesticide Programs
2

The purpose of this problem formulation is to provide the foundation for the ecological risk 

assessment being conducted for d-Limonene (“limonene,” unless otherwise specified). As such, 

it articulates the purpose and objectives of the risk assessment, evaluates the nature of the 

problem, and provides a plan for analyzing the data and characterizing the risk (EPA, 1998). This 

problem formulation is intended to identify data gaps, uncertainties, and potential assumptions 

used to address those uncertainties. 

Nature of Regulatory Action 

As part of the implementation of the new Registration Review program pursuant to Section 3(g) 

of the Federal Insecticide, Fungicide Act (FIFRA), the Agency is beginning the re-evaluation of 

limonene as a pesticide active ingredient to determine whether limonene continues to meet 

FIFRA standard for registration. This problem formulation for the environmental fate and 

ecological risk assessment chapter in support of the registration review of d- limonene 

(limonene) is intended for the initial docket opening phase of the review process. 

Stressor Source and Distribution 

Nature of the Chemical Stressor

Limonene (d-limonene) is a naturally occurring chemical obtained from the rind of citrus fruits. 

Limonene is a hydrocarbon classified as a cyclic terpene. Specifically, it is a monoterpene. 

Limonene has a chiral (stereogenic) carbon. Thus, limonene occurs as an enantiomeric pair (i.e., 

as two non-superimposable mirror-image chemical structures and therefore, each enantiomer is 

optically active1

. The limonene enantiomer under review is the d-enantiomer (i.e., the 

dextrorotatory enantiomer). This dextrorotatory enantiomer has the R-absolute configuration. 

Unless otherwise specified, the term “limonene” is used interchangeably with “d-limonene.” 

The mode of action of limonene is believed to be inhibition of acetylcholine esterase and 

phosphatase activity2

. As an herbicide, d- limonene is a non-systemic, contact herbicide that 

strips the waxy rind or cuticle from weeds, causing death of the plant3. 

 

1 The chirality of limonene arises from an asymmetric carbon atom (i.e., carbon is the chiral/stereogenic center). 

The odors of l-limonene and d-limonene are different; l-limonene smells piney and turpentine like and d-limonene 

has a pleasing orange scent. This is associated with enantiomer-specific interactions with chiral olfactory receptors. 

2 Kostyukovsky M, Rafaeli A., Gileadi C., Demchenko N. and Shaaya ,E. 2002. Activation of octopaminergic 

receptors by essential oil constituents isolated from aromatic plants: possible mode of action against insect pests. 

Pest Management Science, Vol. 58 (11), pp 1101-1006 

3 Stephen O. Duke, Joanne G. Romagni, Franck E. Dayan 2000. Natural products as sources for new mechanisms of 

herbicidal action. Crop Protection 19 (2000) 583-589 
3

Overview of Pesticide Usage

Limonene is used as an acaricide, herbicide, insecticide, and repellent/feeding depressant. 

Its use patterns are food/feed and non-food/ non-feed. A summary of use sites and groups for 

food/feed patterns are shown in Table 1 and in Table 2 for non-food/non-feed. 

Limonene is not applied aerially. It can be applied using sprayers (ready-to-use containers, 

hooded sprayers, pump sprayers, and pressurized containers); aerosol containers (cans); 

impregnated strips; injection equipment; or with a sponge. There is a wide range of limonene 

concentrations and application rates in formulated products. 

Table 1. Food/Feed Use Sites and Groups 

Use Sites Groups 

Terrestrial Citrus fruits, cole crops, flavoring/spice crops, 

fruiting vegetables, grapes, leafy vegetables, legume 

vegetables, pome fruits, root and tuber vegetables, 

small fruits, tree nuts 

Indoor Household/domestic dwellings 

Food handling areas 

Commercial egg packing plants 

Food processing plant premises (non-food contact) 

Food Stores/markets/supermarket premises 

Food/grocery/marketing storage/distribution 

facilities 

Meat processing plant premises (non-food contact) 

Greenhouses Not specified 

 

Table 2. Non-food/Non-feed Use Sites and Groups 

Use Sites Groups 

Terrestrial Citrus; Christmas trees; Forest trees; Grapes; Non 

agricultural outdoor buildings; Nonagricultural 

rights-of-ways/fencerows,hedgerows; Nonagricultural cultivated arrears/soil; Nursery stock; 

Ornamental/shade trees/herbaceous plants/lawn and 

turf; nonflowering plants/ woody shrubs and vines; 

Pome fruits; Small fruits; Stone fruits Tree nuts 

Indoor Automobiles, taxis, limousines, recreational 

vehicles, tires 

Commercial/institutional/industrial 

premises/equipment 

Commercial storage/warehouse premises 

Dogs/canine (adults/puppies) 

Hospitals/medical institutions premises 

(human/veterinary) 

Household/domestic dwellings/premises/handling 

areas 

Human body/clothing while being worn 

Pet living/sleeping quarters 

Refuse/solid waste containers (garbage cans); Wide 

area/general indoor treatment; Wood protection 

treatment to buildings/products 
4

Outdoor Cats (adults/kittens commercial transportation 

facilities 

Commercial/institutional/industrial 

premises/equipment 

Household/domestic dwellings/premises 

Manure 

Outdoor buildings and structures; Paths/patios; 

Private roads/sidewalks 

Ships and boats; Recreational areas; Refuse/solid 

waste sites; Sewage systems; Ships and boats; Wide 

area/general indoor treatment (public health use; 

Wood protection treatment to buildings/products) 

Greenhouse Not specified 

The highest application rate (34.6 lb ai/A) corresponds to use as a terrestrial burn-down 

herbicide. The herbicide is applied to terrestrial food/feed crops as well as non-food/non-feed 

crops and outdoor premises. The herbicide can be applied by broadcast or as a spot treatment. 

When the application rate of 34.6 lb ai/A is used, the herbicide is broadcasted over the target 

vegetation. 

Integration of Available Information 

Regulatory History of Limonene

 

Limonene (d-limonene) has been registered in the United States as an insecticide since 1958. In 

1971, limonene was registered as an antimicrobial and in 1983 as a dog and cat repellent. In 

1988, the Agency considered d-limonene as an inert when used in antimicrobial products 

(40CFR 180.001 (c) (2)). Limonene was exempted for the requirement of a tolerance in 1994 

when it is used as a solvent or fragrance in pesticide formulations. Limonene is currently 

registered as an acaricide, herbicide, insecticide, and repellent/feeding depressant. 

In September 1994, the Agency issued a “Re-registration Eligibility Decision” (RED; EPA-738-

F-94-030) for d-Limonene. It was concluded that pesticide products containing limonene would 

not pose unreasonable risks or adverse effects to humans and the environment. Therefore, the 

1994 RED concluded that all products containing limonene were eligible for re-registration. 

Because the 1994 RED was issued prior to the 1996 enactment of the Food Quality Protection 

Act (FQPA), no drinking water assessment was performed at that time. However, the TRED 

concluded that given its rapid volatilization from water, it is unlikely that d-limonene will occur 

in drinking water sources resulting from any of the registered uses. 

Limonene is also regulated by the U.S. Food and Drug Administration (FDA) as a food additive 

or flavoring and as a fragrance additive. The FDA recognizes d-limonene as a Generally 

Recognized as Safe (GRAS) chemical (21 CFR 182.60).

Other non-pesticide uses of limonene include: (a) solvents (substitute for chlorinated 

hydrocarbons, chlorofluorocarbons, and other solvents); (b) degreasing of metals; (c) cleaning 

agent in the printing industry and household cleaning products; (d) solvent for paint. 
5

Previous Ecological Risk Assessments

The ecological risk assessment in the Re-registration Eligibility Decision (RED) completed in 

September 1994 (EPA 738-R-94-034) is based on a limited number of environmental fate and 

ecological toxicity studies. Exposure concentrations were estimated using simple screening level 

exposure models. The risk assessment concluded that minimal risk is anticipated for birds, 

mammals, and aquatic animals for the pesticidal uses of limonene. EFED believed at that time 

that applications of limonene did not pose any undue risks to endangered species. 

Exposure Characteristics 

Chemical identification and physical-chemical properties of limonene are shown in Table 3. 

Whenever possible, experimental physical-chemical properties are reported. 

Table 3. Identity and Physical-Chemical Properties of the Stressor 

Type of Information Information 

Common name d-Limonene 

Chemical name IUPAC: (R)-4-isopropenyl-1-

methylcyclohexene 

or p-mentha-1,8-diene 

CAS (4R)-1-methyl-4-(1-

methylethenyl)cyclohexene 

Other Names: D-Limonene; Cyclohexene, 1-methyl-4-

(1-methylethenyl)-, (R)-; p-Mentha-1,8-diene, (R)-(+)-;

(+)-(R)-Limonene; (+)-(4R)-Limonene; (+)-p-Mentha1,8-diene; (R)-(+)-Limonene; Carvene; D-(+)-Limonene; 

Dipentene; Limonene; Limonene, (+)-; (R)-1-methyl-4-

(1-methylethenyl)cyclohexene; Dextro-limonene; (γ)-

Carvene; (R)-4-Isopropenyl-1-methyl-1-cyclohexene; pMentha-1,8-diene; R(γ)-Limonene; 4-Isopropenyl-1-

methyl-1-cyclohexene; (R)-limonene. 

For other names consult EPA’s Substance Registry 

Service 

Pesticide type Acaricide, herbicide, insecticide, repellent/feeding 

depressant 

Chemical class Terpene (monoterpene) 

CAS Reg. No. d- limonene: 5989-27-5

Racemate: 7705-14-8 

 l- Limonene: 5989-54-8 

US. EPA Chemical Code Number 079701 

Empirical Formula C10H16

Smiles String C(=CCC(C(=C)C)C1)(C1)C

Chemical structure 
6

Molecular mass (g/mol) 136.23 

Physical State Liquid (colorless) 

Specific gravity 0.841 g/cm3

Melting point,  ̊C -74.35 

Boiling point,  ̊C 175.5-176 

Vapor pressure 20  ̊C

2 mmHg; 2 torr; 2.66 x 103

 Pa (Product chemistry data 

(465111-03) 

25  ̊C

1.4 mmHg; 1.4 torr; 1.92 x 103

 Pa (Estimated at 25  ̊C 

(EPISuite) 

Henry’s Law Constant 3.19 x 10-1 atm-m3/mole; 3.85 x 104 Pa-m3/mole 

Solubility in water (mg/L) Insoluble (465111-03) 

13.8 mg/L (Massaldi and Kin,1973) 

Massaldi and Kin as cited by Smith, T.G. and LeMaguer, M. (1980) 

Solubilities of terpenic essential oil components in aqueous solutions. 

J. Chem. Eng. Data, 1980, 25 (2), pp 150–152. 

Log Kow (n-octanol/water partition coefficient) 4.8 (Experimental) 

Log Ko-air (octanol-air partition coefficient) 4.26 (estimated; EPISuite) 

pKa/pKb Not applicable 

Optical Rotation (Technical Product) > + 98 (465111-03) 

Environmental Fate

Persistence Limonene does not hydrolyze. Its cyclohexene ring and ethylene groups are 

resistant to hydrolysis. Therefore, hydrolysis is not expected in soils or aquatic 

systems. 

Under normal environmental conditions, limonene is not likely to be metabolized 

by soil microorganisms. Limonene has been reported to metabolize in some pure 

microbial cultures.4

In the atmosphere, volatilized limonene undergoes rapid bimolecular gas-phase 

reactions (i.e., the reactants are gases) with ozone, nitrate radicals, and 

photochemically generated hydroxyl radicals. 

Transport Limonene binds strongly to soil (Koc > 1000 L/kg), therefore it is expected to be 

relatively immobile in soils. However, the vapor pressure (1.4 mmHg at 25  ̊C) 

and Henry’s Law Constant 3.19 x 10-1 atm-m3

/mole indicate that limonene readily 

volatilizes from soil and water. However, this dissipation process competes with 

binding. 

 

4 Amaral, J.A., Keins, A., Richards, S.R., and Knowles, R. 1998. Effect of Selected Monoterpenes om Methane 

Oxidation, Denitrification, and Aerobic Metabolism by Bacteria in Pure Culture. Applied and Environmental 

Microbiology, Vol 64(2), pp 520-525. 
7

Transformation The products formed by gas-phase reactions depend on the type of reactant 

(i.e., ozone, hydroxyl and nitrate free radicals). The products formed by the nitrate radical are 

simpler than those formed by the hydroxyl radical. 

Dissipation Pathways Volatilization from soil, other surfaces, and water are the major 

dissipation pathways for limonene. 

Bioaccumulation The Log Kow of 4.83 indicates potential bioaccumulation. 

Detections in Environmental Media (Environmental Levels)

Limonene residues have been detected in rural, urban, suburban, and forest areas in the U.S. and 

in other countries (CICADS document, 1998; Footnote 3). It has been detected in air, surface 

water, ground water, seawater, wastewater, and ice as well as in soils and sediments. Limonene 

has been detected in solid waste composting facilities. Levels of limonene are varied, but the 

limited data from 1998 do not allow correlation with usage or geographical region. However, it 

appears that there is a temporal variability in the number of detections. 

EFED has data from 270 samples collected in ambient surface of water. Only one had a 

concentration of limonene above 0.5 μg/L (which is believed to be the limit of detection). That 

sample was 1.2 μg/L. 

Ecological Effects 

Consistent with the process described in the Overview Document (EPA, 2004), this risk 

assessment will use a surrogate species approach in its evaluation of d-limonene. Toxicological 

data generated from surrogate test species, which are intended to be representative of broad 

taxonomic groups, are used to extrapolate to potential effects on a variety of species (receptors) 

included under these taxonomic groupings. 

Acute and chronic toxicity data from studies submitted by pesticide registrants along with the 

available open literature are used to evaluate the potential direct effects of d-limonene to the 

aquatic and terrestrial receptors identified in this section. This includes toxicity data on the 

technical grade active ingredient, degradates, and when available, formulated products (e.g. “SixPack” studies). 

The open literature studies are identified through EPA’s ECOTOX database 

(http://cfpub.epa.gov/ecotox/), which employs a literature search engine for locating chemical 

toxicity data for aquatic life, terrestrial plants, and wildlife. The evaluation of both sources of 

data can also provide insight into the direct and indirect effects of d-limonene on biotic 

communities from loss of species that are sensitive to the chemical and from changes in structure 

and functional characteristics of the affected communities. 

No ecological incidents were found in the Agency’s Ecological Incident Information System 

EIIS database. 
8

Table 4 summarizes the available data used to evaluate the ecological toxicity and risk of 

limonene in the 1994 assessment. 

Table 4. Summary of data used to evaluate ecological effects of limonene. 

Measures of effect Species Toxicity Value Toxicity 

Classification 

Toxicity 

Category 

(Acute) 

MRID 

(Year) Notes 

Single dose avian 

toxicity Bobwhite1 >2,000 mg/kg Core Practically Nontoxic 

109340 

(1982) 

Mallard 2 >5,000 mg/kg Core Practically 

Nontoxic 

109341 

Dietary avian (1982) 

toxicity Bobwhite1 >5,000 mg/kg Core Practically 

Nontoxic 

109341 

(1982) 

96-hr Freshwater 

fish, cold water Rainbow Trout 3 205 ppm Supplemental Practically 

Nontoxic 

109343 

(1982) 

96-hr Freshwater 

fish, warm water Fathead minnow 4 595 ppm Supplemental Practically 

Nontoxic 

109344 

(1982) 

Freshwater 

invertebrates Daphnia magna 17-20 ppm Supplemental Slightly 

Toxic 

109345 

(1982) 

Formulated product 

(dog repellent) 

Avian Dietary 

toxicity Bobwhite1 >5,600 mg/kg Core Practically 

Nontoxic 

146988 

(1984) 

96-hr Freshwater 

fish, cold water Rainbow Trout 3 80 ppm Core 

Slightly 

Toxic 146085 

(1984) 

Freshwater 

invertebrates Daphnia magna 39 ppm Core Slightly 

Toxic 

146085 

(1984) 

Mammalian 

Acute oral Laboratory rat 5 >5,000 mg/kg Core Practically 

Nontoxic 

EFED 

Science 

Chapter 

(1994) 

Technical Grade 

1 Colinus virginianus, 2

 Anas platyrhynchos, 3 Oncorhynchus mykiss, 4 Pimephales promelas, 

5 Rattus norvegicus 

Avian and Aquatic Effects 

The toxicity effects for avian species are “Practically Nontoxic” for both the technical and 

formulated products. Formulated limonene is “Practically Nontoxic” to aquatic vertebrates and 

“Slightly Toxic” to freshwater invertebrates. Technical limonene is “Slightly Toxic” to aquatic 

species. 
9

Mammalian Acute Effects 

The acute oral LD50 value for the Norway rat is greater than 5000 mg/kg (5 g/kg), which 

classifies it as practically nontoxic. Granular limonene is registered as a mammalian repellent 

and is categorized as “Practically Nontoxic.” The “granules” are diatomaceous earth; therefore, 

their size is very small (typically 10 to 200 μm). It is not anticipated that “granules” would be 

consumed in sufficient quantities to be a risk to mammals. 

Ecosystems Potentially at Risk 

In general, the ecosystems at risk are those in close proximity to the use areas. These include 

any water bodies and terrestrial ecosystems adjacent to use areas that may receive runoff. Risks 

are considered for aquatic animals (fish, invertebrates, amphibians) and plants, assumed to occur 

in small, static ponds and marine water bodies in areas adjacent to the use site and terrestrial 

animals (birds, reptiles, terrestrial amphibians, mammals and insects) and plants, assumed to 

exclusively occur in the treated area. 

Organisms of potential concern broadly include fish, aquatic invertebrates and plants, birds, 

amphibians, reptiles, mammals and terrestrial invertebrates and plants. Based on the known 

effects of limonene, the organisms of concern could be terrestrial and aquatic invertebrates and 

plants. 

The assessment endpoints are intended to reflect population sustainability and community 

structure within ecosystems and hence relate back to ecosystems at risk. If risks are expected for 

given species/taxa based on the screening-level assessment, then risks might be expected to 

translate to higher levels of biological organization. 

Assessment Endpoints 

Assessment endpoints are defined as “explicit expressions of the actual environmental value that 

is to be protected.” Defining an assessment endpoint involves two steps, 1) identifying the 

valued attributes of the environment that are considered to be at risk, and 2) operationally 

defining the assessment endpoint in terms of an ecological entity (i.e., a community of fish and 

aquatic invertebrates) and its attributes (i.e., survival and reproduction). Therefore, selection of 

the assessment endpoints is based on valued entities (i.e., ecological receptors), the ecosystems 

potentially at risk, the migration pathways of pesticides, and the routes by which ecological 

receptors are exposed to pesticide-related contamination. The selection of clearly defined 

assessment endpoints is important because they provide direction and boundaries in the risk 

assessment for addressing risk management issues of concern. Changes to assessment endpoints 

are typically estimated from the available toxicity studies, which are used as the measures of 

effects to characterize potential ecological risks associated with exposure to a pesticide, such as 

limonene. 

The most sensitive toxicity endpoints are used from surrogate test species to estimate treatmentrelated direct effects on acute mortality and chronic reproductive, growth and survival 

assessment endpoints. Toxicity tests are intended to determine effects of pesticide exposure on 
10

birds, mammals, fish, terrestrial and aquatic invertebrates, and plants. These tests include shortterm acute, sub-acute, and reproduction studies and are typically arranged in a hierarchical or 

tiered system that progresses from basic laboratory tests to applied field studies. The toxicity 

studies are used to evaluate the potential of a pesticide to cause adverse effects, to determine 

whether further testing is required, and to determine the need for precautionary label statements 

to minimize the potential adverse effects to non-target animals and plants. 

Conceptual Model

For a pesticide to pose an ecological risk, it must reach ecological receptors in biologically 

significant concentrations. An exposure pathway is the means by which a pesticide moves in the 

environment from a source to an ecological receptor. For an ecological pathway to be complete, 

it must have a source, a release mechanism, an environmental transport medium, a point of 

exposure for ecological receptors, and a feasible route of exposure. 

Risk Hypothesis

Based on an examination of the physical/chemical properties of d-limonene, the fate and 

disposition in the environment, and method of application, the following ecological risk 

hypothesis is being employed for this baseline risk assessment: Limonene will affect the growth, 

reproduction, and survival of non-target species when it is used as per label directions. The 

major transport pathway for d-limonene is volatilization. 
11

Conceptual Diagram 

Analysis Plan Options 

In Registration Review, pesticide ecological risk assessments will follow the Agency’s 

Guidelines for Ecological Risk Assessment, will be consistent with the paper titled “Overview of 

the Ecological Risk Assessment Process in the Office of Pesticide Programs, U.S. Environmental 

Protection Agency” (“Overview Document”) (January 2004), and will be done in accordance 

with Section 7 of the Endangered Species Act. 

The ecological risk assessment for limonene will be performed only for uses of limonene as a 

pesticide. Non-pesticide uses will not be assessed, but published data can be included in the 

overall environmental exposure of limonene. Exposure will be evaluated using modeling and 

monitoring data. In the ecological risk reassessment for limonene, the Agency will include data 

previously used in the 1994 RED, as well as open literature identified using EPA’s ECOTOX 

database. Data on terrestrial and aquatic plants are needed and the results will be used in the risk 

assessment for plants. 

Limonene is volatile. The use of atmospheric transport models may be necessary because offtarget deposition of limonene on vegetation and aquatic ecosystems could potentially affect nonStressor Application of Limonene

Direct Ground Deposition

Volatilization/

Wind Suspension Leaching

Non-Target Plants:

•Plant growth abnormalities

•Lethality

•Developmental/reproductive

effects

Source:

Transport Pathway

Terrestrial Animals:

•Lethality

•Reproductive/developmental

effects

Aquatic Animals and Plants:

•Effects to survival, growth, and 

reproduction

•Reductions in cell density/ 

biomass in aquatic plants

Altered Terrestrial Community Assemblages:

Reduced food supply and edge habitat availability

Altered Aquatic Community Assemblages:

Effects to riparian habitat availability

Effects to species abundance/diversity

Changes in physical/chemical parameters

Source:

Exposure Media

Exposure Point

Receptors

Direct Effects

Indirect Effects

Terrestrial Food Residues

(foliage, fruit, insects)

Direct Contact

& Ingestion

Terrestrial

Vertebrates and 

Invertebrates

Direct Contact/

Root Uptake

Terrestrial

Upland

Plants

Surface Water

Sediment Groundwater

Gill/Integument

Uptake

Aquatic

Invert &

Vertebrates

Upland Soil and 

Foliage

Riparian/Wetland

Foliage/Soil

Spray

Drift Runoff/

Erosion

Uptake/

Adsorption

Riparian/

Wetland

Plants

Aquatic

Plants

Plant Population

Reduced Population

growth
12

target plants. Volatilization rates will be estimated using volatilization models recommended by 

EFED and will be integrated in the models used to estimate concentrations of limonene in 

surface water. If necessary, published gas-phase kinetics data will be used to estimate the 

degradation of gaseous limonene in the atmosphere (i.e., reactions with photochemically 

generated hydroxyl radicals, ozone, and nitrogen oxide radicals). A source for these data is the 

National Institute of Science and Technology (NIST) Kinetics Database. 

The Health Effects Division (HED) concluded in 1994 that it is unlikely that d-limonene will 

occur in drinking water sources (surface; ground) resulting from any of the registered and 

proposed uses as an active ingredient or when used as an inert ingredient. Residues in drinking 

water were thus not included in the dietary risk assessments. Given the current use pattern, 

EFED plans to conduct a drinking water assessment in registration review. 

Exposure concentrations in aquatic ecosystems will consider the use of the Tier I GENEEC2 

and/or Tier II PRZM-EXAMS models using actual or estimated environmental fate/physicalchemical data. 

Endangered Species

Consistent with the Agency’s responsibility under the Endangered Species Act (ESA), EPA will 

evaluate risks to federally-listed threatened and/or endangered (listed) species from registered 

uses of limonene. This assessment will be conducted in accordance with the Overview 

Document, provisions of the ESA, and the Services’ Endangered Species Consultation 

Handbook . 

The assessment of effects associated with registrations of limonene is based on an action area. 

The action area is considered the area directly or indirectly affected by the federal action, as 

indicated by the exceedance of Agency LOCs used to evaluate direct or indirect effects. The 

Agency’s approach to defining the action area under the provisions of the Overview Document 

considers the results of the risk assessment process to establish boundaries for that action area 

with the understanding that exposures below the Agency’s defined LOCs constitute a no-effect 

threshold. For the purposes of this assessment, attention will be focused on the footprint of the 

action (i.e., the area where limonene application occurs), plus all areas where offsite transport 

(i.e., spray drift and runoff) may result in potential exposure that exceeds the Agency’s LOCs. 

Specific measures of ecological effect that define the action area for listed species include any 

direct and indirect effects and/or potential modification of its critical habitat, including reduction 

in survival, growth, and reproduction as well as the full suite of sublethal effects available in the 

effects literature. Therefore, the action area extends to a point where environmental exposures 

are below any measured lethal or sublethal effect threshold for any biological entity at the whole 

organism, organ, tissue, and cellular level of organization. In situations where it is not possible 

to determine the threshold for an observed effect, the action area is not spatially limited and is 

assumed to be the entire United States. 
13

Clean Water Act 

Limonene is not identified as a cause of impairment for any water bodies listed as impaired 

under section 303(d) of the Clean Water Act, based on information provided at 

http://iaspub.epa.gov/tmdl_waters10/attains_nation_cy.cause_detail_303d?p_cause_group_id=88

5. 

In addition, no Total Maximum Daily Loads (TMDL) have been developed for limonene, based 

on information provided at 

http://iaspub.epa.gov/tmdl_waters10/attains_nation.tmdl_pollutant_detail?p_pollutant_group_id

=885&p_pollutant_group_name=PESTICIDES. More information on impaired water bodies and 

TMDLs can be found at http://www.epa.gov/owow/tmdl/. 

EPA invites submission of water quality data for this pesticide. To the extent possible, data 

should conform to the quality standards in Appendix A of the OPP Standard Operating 

Procedure: Inclusion of Impaired Water Body and Other Water Quality Data in OPP’s 

Registration Review Risk Assessment and Management Process (see: 

http://www.epa.gov/oppfead1/cb/ppdc/2006/november06/session1-sop.pdf), in order to ensure 

they can be used quantitatively or qualitatively in pesticide risk assessments. 

Endocrine Disruptor Screening Program 

As required under FFDCA section 408(p), EPA has developed the Endocrine Disruptor 

Screening Program (EDSP) to determine whether certain substances (including pesticide active 

and other ingredients) may have an effect in humans or wildlife similar to an effect produced by 

a “naturally occurring estrogen, or other such endocrine effects as the Administrator may 

designate.” The EDSP employs a two-tiered approach to making the statutorily required 

determinations. Tier 1 consists of a battery of 11 screening assays to identify the potential of a 

chemical substance to interact with the estrogen, androgen, or thyroid (E, A, or T) hormonal 

systems. Chemicals that go through Tier 1 screening and are found to have the potential to 

interact with E, A, or T hormonal systems will proceed to the next stage of the EDSP where EPA 

will determine which, if any, of the Tier 2 tests are necessary based on the available data. Tier 2 

testing is designed to identify any adverse endocrine related effects caused by the substance, and 

establish a dose-response relationship between the dose and the E, A, or T effect. 

Between October 2009 and February 2010, EPA issued test orders/data call-ins for the first 

group of 67 chemicals, which contains 58 pesticide active ingredients and 9 inert ingredients. 

This list of chemicals was selected based on the potential for human exposure through pathways 

such as food and water, residential activity, and certain post-application agricultural scenarios. 

This list should not be construed as a list of known or likely endocrine disruptors. 

Limonene is not among the group of 58 pesticide active ingredients on the initial list to be 

screened under the EDSP. Under FFDCA sec. 408(p) the Agency must screen all pesticide 

chemicals. Accordingly, EPA anticipates issuing future EDSP test orders/data call-ins for all 

pesticide active ingredients. 
14

For further information on the status of the EDSP, the policies and procedures, the list of 67 

chemicals, the test guidelines and the Tier 1 screening battery, please visit our website: 

http://www.epa.gov/endo/. 

Preliminary Identification of Data Gaps 

Environmental Fate 

No guideline environmental fate studies have been submitted. Submission of guideline studies 

was waived in 1992. For the reassessment, up-to-date open literature and databases will be used 

to generate an integrated environmental fate assessment (persistence, transformation, transport, 

and dissipation routes) of limonene. EFED believes that the behavior of limonene in the 

environment is well documented and that guideline studies are not necessary. 

Ecological Effects 

The 1994 assessment did not include a terrestrial or aquatic plant assessment because limonene 

was not registered as an herbicide at that time. Terrestrial and aquatic plant toxicity data are not 

available. The data are needed to evaluate the risk of limonene to non-target plants.
15

Guideline Number: 850.4100 and 850.4150

Test Substance: Limonene TEP

Title: Seedling emergence and vegetative vigor studies (Tier 2) 

Test Species: Terrestrial plant species as specified by 850.4100 and 850.4150

Rationale for Requiring the Data 

There are no toxicity data available for terrestrial monocots and dicots. A risk assessment for exposure to 

monocots and dicots is needed based on limonene’s use pattern. The data will be utilized to determine 

whether the Agency’s current measures are effective for protecting terrestrial plants. 

Risk to terrestrial plants has not been quantified in previous assessments, and no terrestrial plant seedling 

emergence or vegetative vigor toxicity data are currently available for limonene. Based on the updated data 

requirements in 40 CFR part 158 (October 2007), these data are required to support all terrestrial use 

patterns, including those relevant to limonene. These phytotoxicity studies should be conducted using the 

maximum label application rate of limonene typical end-use product (TEP). Without these studies, the 

Agency would have to presume risk to listed and non-listed terrestrial plants and to organisms that depend 

on terrestrial plants for sustenance, shelter, or reproduction. Potential risks, however, would not be able to 

be quantified. 

Practical Utility of the Data 

How will the data be used? 

Tier II seedling emergence and vegetative vigor toxicity data will be used to quantitatively estimate 

potential risks to terrestrial plants associated with the use of limonene. These data will be used to define an 

action area for endangered species and explore risk mitigation strategies. The data will reduce uncertainties 

associated with the risk assessment for terrestrial plants and will improve the Agency’s understanding of 

the potential effects of limonene on terrestrial plants. In the absence of data, risk to terrestrial plants will be 

presumed. 

How could the data change the Agency’s decision or impact the Agency’s future decision-making? 

The value of these seedling emergence and vegetative vigor studies would be in quantifying and potentially 

refining risks to non-target terrestrial plants. Defining an action area for endangered species and exploring 

risk mitigation strategies cannot be evaluated without these data. Without terrestrial plant toxicity data for 

limonene, the Agency cannot determine if the current application rates of limonene may result in direct 

adverse effects to terrestrial plants and/or indirect effects (e.g., via loss of habitat/cover, riparian vegetation 

effects, etc.). If future endangered species risk assessments are performed without these data, the Agency 

will assume that limonene “may affect” terrestrial plants directly and listed species from other taxa 

indirectly, and use of limonene may need to be restricted in areas where listed species could be exposed. 

The lack of these data will limit the flexibility the Agency and registrants have in coming into compliance 

with the Endangered Species Act and could result in use restrictions for limonene that are unnecessarily 

severe. 
16

Guideline Number: 850.4400 and 850.5400 

Test Substance: Limonene (TEP)

Title: Aquatic vascular and non-vascular plant toxicity tests; Tier II

Test Species: Aquatic plant species as specified by 850.4400 and 850.5400 

Rationale for Requiring the Data

Risk to aquatic plants has not been considered in previous assessments, and no aquatic vascular or nonvascular plant toxicity data are currently available for limonene. Based on the updated data requirements in 

40 CFR part 158 (October 2007), these data are required to support all terrestrial use patterns, including those 

relevant to limonene. (Tier II phytotoxicity tests would be required if a 50 percent or greater detrimental 

effect is observed in the Tier I test). Without this study, the Agency would have to presume risk to listed and 

non-listed aquatic vascular plants, but would not be able to quantify the risk. 

Practical Utility of the Data

 How will the data be used? 

Aquatic plant toxicity data will be used to quantitatively estimate the risk of limonene to aquatic plants and to 

define an action area for endangered species and explore risk mitigation strategies. These data will greatly 

reduce uncertainties associated with the risk assessment for aquatic plants. In the absence of aquatic plant 

data for limomene, quantitative risk estimation will not be possible, and risk will be presumed. Determination 

of the distance downstream from a treated field to which risk may extend and application rate reductions 

resulting in no LOC exceedances cannot be quantified without these data. 

How could the data impact the Agency’s future decision-making?

Because aquatic plants form the basis of the food chain for aquatic habitats and significantly contribute to 

overall water quality, a solid understanding of the potential risks to aquatic plants is essential for sound 

environmental risk management decision-making. Aquatic plant toxicity data will allow the Agency to 

quantitatively characterize the potential risks of limonene to aquatic ecosystems. If future endangered 

species risk assessments are performed without these data, the Agency would have to assume that limonene 

"may affect" listed aquatic plants directly and other endangered species indirectly via reduction in the prey 

base and/or loss of habitat or cover. The lack of these data will limit the flexibility the Agency and 

registrants have in complying with the Endangered Species Act and could result in unnecessary use 

restrictions.