UNITED STATES ENVIRONMENTAL PROTECTION AGENCY 

WASHINGTON D.C., 20460 

OFFICE OF CHEMICAL SAFETY AND POLLUTION PREVENTION 

PC Code: 002201 

DP Barcode: 427826 

Date: September 8, 2015 

MEMORANDUM 

Subject: 

From: 

Through: 

To: 

Registration Review: Problem Formulation for the Environmental Fate, Ecological Risk, 

Endangered Species, and Drinking Water Assessm nts f r S badilla Alkaloids 

_/ q/,;/')..01) Freeborn Jewett, M.S., Biologisv-:::J 

He Zhong, Ph.D., Biologist 9 I 'if I,. o/ ..r 

Environmental Risk Branch I 

Environmental Fate and Effects Division (7507P) 

o~ q/s/1) 

Greg Orrick, Risk Assessment Process Leader ~ ~ \ g \ \ .j 

Sujatha Sankula; Ph.D., Branch Chief h~A~-1'--.......J ~ Environmental Risk Branch I ' -=- ,,. ' 

Environmental Fate and Effects Division (7507P) 

Roy Johnson, Chemical Review Manager 

Jill Bloom, Team Leader 

Linda Arrington, Branch Chief 

Risk Management and Implementation Branch V 

Pesticide Re-Evaluation Division (7508P) 

1 
2

1. Executive Summary:

The Environmental Fate and Effects Division (EFED) has completed the problem formulation for the 

environmental fate, ecological risk, endangered species, and drinking water exposure assessments to be 

conducted as part of the Registration Review of the insecticide sabadilla alkaloids (CAS Number 8051-02-

3). Sabadilla alkaloids refers to a mixture of several alkaloids, veratridine, cevadine, sabadilline, 

sabadinine, and sabadine, which are derived from the crushed seeds of the sabadilla lily (Schoenocaulon 

officinale). The sabadilla alkaloid mixture is composed primarily of cevadine and veratridine, with 

smaller amounts of other alkaloids such as sabadine, sabadinine, and sabadilline (Tomlin, 2003). One 

product containing sabadilla alkaloids is currently registered for use under the name Veratran D (0.2% 

ai) (EPA reg. #1021-2600), for the control of thrips in avocados, mangos and citrus. Veratran D is allowed 

for use in certified organic operations under the USDA National Organic Program. It is formulated as a 

wettable powder that is mixed with water and applied by aerial or ground application. The label for 

Veratran D requires clarification, as the current language does not specify a maximum annual 

application rate, a maximum number of applications/year, or a minimum retreatment interval. 

Functioning as the first stage of the risk assessment process for Registration Review, this problem 

formulation provides an overview of what is currently known about the environmental fate and 

ecological effects associated with sabadilla alkaloids. It also describes the preliminary ecological risk 

hypothesis and analysis plan for evaluating and characterizing drinking water exposure and risk to nontarget species and the environment in support of the Registration Review of sabadilla alkaloids. This 

document also recommends studies to address uncertainties related to the environmental fate and 

ecological effects of sabadilla alkaloids.

There are no recent actions on sabadilla alkaloids. Sabadilla alkaloids was last assessed in 2004 in the 

Registration Eligibility Decision (RED; EPA, 2004a). This assessment was conducted with limited data and 

relied primarily on predicted toxicity and fate values to characterize risk. These predicted toxicity values 

should not be used in future assessments.

Recommended Studies:

The list of recommended studies reflects a tiered approach. The first tier of recommended studies is 

needed to confirm the assumption from the RED that no risk exceedances exist (i.e. RQ<LOC) for

terrestrial and aquatic taxa. If risk concerns are identified, a second tier of data is recommended. In the 

absence of fate data, EFED will assume stability of the sabadilla alkaloid mixture. 

Table 1: A Summary of Recommended Data for Sabadilla Alkaloids

Taxa Test Substance

(For Tier I and 

Tier II) 

Tier I Tier II

Eco Data

Aquatic 

Invertebrates

TGAI1

 850.1010 Freshwater 

Invertebrate Acute Toxicity 

 850.1035 Estuarine/Marine 

Invertebrate Acute Toxicity (Mysid 

shrimp)

 

1

Sabadilla Alkaloids TGAI refers to a mixture of sabadilla alkaloids that would be representative of the active 

ingredient found in a typical commercially available formulation of Veratran D. Sabadilla alkaloids is a powder 

derived from dried seeds of plants in the lily family, and it has been reported that there is significant variation (>3 
3

Taxa Test Substance

(For Tier I and 

Tier II) 

Tier I Tier II

 850.1300 Freshwater 

Invertebrate Life Cycle 

 850.1025 Estuarine/Marine 

Invertebrate Acute Toxicity (Eastern 

Oyster)

 850.1350 Estuarine/Marine 

Invertebrate Life Cycle

Benthic 

Invertebrates

TGAI --  850.1735 Whole sediment2

: 

Invertebrates Subchronic: Freshwater 

amphipod, Hyalella azteca, and midge, 

Chironomus dilutus

 850.1740 Whole sediment2: 

Invertebrates Subchronic: 

Estuarine/marine amphipod

Leptocheirus plumulosus

 Non-Guideline Chronic Sediment 

Toxicity Test (if risks are identified in 

subchronic tests)

Fish TGAI  850.1075 Freshwater Fish 

Acute Toxicity (coldwater 

species)

 850.1400 Freshwater Fish 

Early-Life Stage (coldwater 

species)

 850.1075 Freshwater Fish Acute 

Toxicity (warm water species)

 850.1075 Estuarine/Marine Fish Acute 

Toxicity

 850.1400 Freshwater Fish Early-Life 

Stage (warm water species)

 850.1400 Estuarine/Marine Fish EarlyLife Stage

Aquatic Plants TGAI  850.4400 Aquatic Vascular 

Plant Toxicity Test using 

Lemna spp.

 850.4500 Algal Toxicity 

Pseudokirchneriella 

subcapitata

 850.4500 Marine diatom- Skeletonema 

costatum

 850.4500 Freshwater diatom-Navicula 

pelliculosa

 850.4550 Blue Green Algae/ 

Cyanobacterium-Anabaena flos-aquae

Birds TGAI  850.2100 Avian Acute Oral 

Toxicity (Bobwhite Quail)

 850.2300 Avian 

Reproduction (Bobwhite 

Quail)

 850.2100 Avian Acute Oral Toxicity 

(Mallard Duck)

 850.2100 Avian Acute Oral Toxicity 

(Passerine species)

 850.2200 Avian Dietary Toxicity 

(Bobwhite Quail and Mallard Duck)

 850.2300 Avian Reproduction (Mallard 

Duck)

 850.2200 Avian Dietary Toxicity

 

fold) in the concentrations of veratridine and cevadine (Hare and Morse, 1997) in the finished product. As a result, 

all studies conducted with sabadilla alkaloid TGAI should be carried out with the same relative proportions of 

veratridine, cevadine, sabadinine, and sabadine in the mixture. Analytical confirmation of the concentration of 

each component (veratridine, cevadine, sabadinine, and sabadine) must be provided in the study report for each 

test. Furthermore, the registrant needs to submit confirmatory analysis that demonstrates that the sabadilla 

alkaloids TGAI used for testing is representative of the composition of sabadilla alkaloids contained within a 

Veratran D, the typical end use product (TEP). 

2 Needed if KOC values are analytically confirmed to be consistent with those predicted by EPISuite. 
4

Taxa Test Substance

(For Tier I and 

Tier II) 

Tier I Tier II

Terrestrial 

Invertebrates

TGAI  Non-Guideline Tier I 

Honeybee Adult Acute Oral 

Toxicity

 Non Guideline Tier IHoneybee Adult Chronic 

Oral Toxicity

 Non Guideline Tier IHoneybee Larval Acute and 

Oral Exposure

 Non-Guideline Nectar and Pollen 

Residue Study for Insect Pollinators 

 Non-Guideline Semi-Field Testing for 

Honeybees 

 850.3040 Field Testing for Pollinators 

Terrestrial 

Plants

TEP3

 850.4100 Seedling 

Emergence and Seedling 

Growth 

 850.4150 Terrestrial Plants: Vegetative 

Vigor

Fate Data

Test Substance Tier I Data Requirement Tier II Data Requirement

Veratridine4

 835.2120- Hydrolysis

 835.2240- Aqueous 

Photolysis 

 835.2410- Soil Photolysis

 835.4100-Aerobic Soil 

Metabolism 

 835.4300- Aerobic Aquatic 

Metabolism

 835.4400- Anaerobic 

Aquatic Metabolism 

 835.1230/835.1240 

Leaching Adsorption/ 

Desorption 

 850.1730- Accumulation in 

Fish 

 850.6100- ECM/ILV (soil and 

water) 

--

Cevadine4--  835.4300- Aerobic Aquatic Metabolism

 835.4100- Aerobic Soil Metabolism

 835.2120- Hydrolysis

TEP3

 835.6100- Terrestrial Field 

Dissipation 

--

 

3 Veratran D

4 Two main insecticidal components of sabadilla alkaloid mixture
5

1. Introduction

The purpose of this problem formulation is to provide an understanding of what is known about the 

environmental fate and ecological effects of sabadilla alkaloids. This document will provide a plan for 

analyzing data relevant to sabadilla alkaloids and for conducting ecological risk, endangered species, and 

drinking water exposure assessments for its registered uses. This problem formulation is intended to 

identify data gaps, uncertainties, and potential assumptions used to address those uncertainties relative 

to characterizing the ecological risk associated with the registered uses of sabadilla alkaloids.

The term “sabadilla alkaloids” refers to a class of insecticidal compounds that consists of a mixture of 

chemicals rather than a single chemical moiety. Sabadilla alkaloids (PC Code: 002201) is derived from 

the crushed seeds of the sabadilla lily (Schoenocaulon officinale). The Registration Eligibility Decision 

(USEPA, 2004a) evaluated sabadilla alkaloids as a potential low-risk pesticide due to a lack of toxicity, 

and a risk assessment was performed on the alkaloids cevadine, veratridine, sabadine, and sabadilline. 

The insecticidal action of this chemical class is primarily the result of two of the alkaloids, veratridine 

(CAS: 71-62-5) and cevadine (CAS: 62-59-9) (Allen et al., 1945; Hare and Morse, 1997; Ikawa et al., 1945). 

Sabadilla alkaloids is currently registered for use under the name of Veratran D (0.2% ai) (EPA reg. 

#1021-2600) for the control of thrips in avocados, mangos and citrus. Veratran D allowed for use in 

certified organic operations under the USDA National Organic Program. It is formulated as a wettable 

powder that is mixed with water and applied by aerial or ground application. 

Limited data are available to characterize risk to terrestrial and aquatic taxa. Open literature studies 

provide qualitative endpoints on the toxicity of sabadilla alkaloids to freshwater fish, mammals, and 

terrestrial invertebrates. Uncertainty related to the available open-literature data, and a lack of 

registrant-submitted data related to the environmental fate and toxicity effects of sabadilla alkaloids,

means that it is difficult to characterize the risk picture of sabadilla alkaloids to non-target species. In the 

absence of data, future risk assessments may be conducted based on conservative assumptions. In 

addition, there are label deficiencies that require clarification. The current label for the only registered 

product, Veratran-D, does not specify a maximum annual application rate, a maximum number of 

annual applications, or a minimum retreatment interval.

Use Characterization

a. Mode of Action

The mode of action of sabadilla alkaloids is a sodium channel effect (delayed channel closure and an 

increase in the probability of channel opening) resulting in nerve excitation (repetitive firing), prolonged 

impulse duration and depolarization of nerve membrane potential, resulting in death. These alkaloids 

bind at the neurotoxin receptor 2 associated with voltage gates sodium channels to cause Na+ channels 

to stay open during a sustained membrane depolarization (Catterall, 1975). The consequential Na+ 

influx, either by itself or by causing a maintained depolarization, leads to many secondary effects such as 

increasing pump activity, Ca2+ influx, and in turn, exocytosis.

b. Overview of Pesticide Use and Usage

The most recent available usage summary of sabadilla alkaloids is available for California in 2012 

(Pesticide Action Network, 2015), and is presented in Table 2. In 2012, 66.2 lbs ai were applied over 

3,928 acres.
6

The Biological and Economic Analysis Division (BEAD) provided a Screening Level Usage Analysis (SLUA) 

(USEPA, 2014a) to provide estimates of the actual usage of sabadilla alkaloids on agricultural crops. The 

screening level usage profile (SLUA) produced by BEAD and located in the docket (USEPA, 2014a) and 

summarized in Table 3, lists the use patterns for the current uses of acequinocyl. Primary uses in terms 

of average weight of applied product of sabadilla alkaloids are avocados, grapefruit, lemons, oranges 

and tangerines. These data represent annual data from 2004-2012.

Sabadilla alkaloids is currently registered under a single formulated product , Veratran D, which is 

composed of 0.2% sabadilla alkaloids and 99.8% other ingredients and is registered for use on avocados, 

mangoes and citrus. The single maximum application rate for Veratran-D is 15 lbs formulated product 

per acre (0.03 lbs ai/A) for aerial application and 20 lbs formulated product per acre (0.04 lbs ai/A) for 

ground application to avocados, citrus and mangos. The current label directs applicators to, “reapply as 

necessary, usually at a 10-14 day interval.” Therefore, multiple applications will be assessed.

Table 2: Estimates of the Actual Usage of Sabadilla Alkaloids on Agricultural Crops in California in 2012 

(Pesticide Action Network, 2015).

Crop Gross Pounds ai Application rate (lb ai /A) Acres Treated

Tangerines 53 0.02 3,305

Avocado 7 0.03 237

Oranges 2.93 0.02 157

Outdoor Container Nursery 1.33 0.02 39.3

Grapefruit 0.82 0.02 80.0

Tangelos 0.60 0.02 30.0

Citrus 0.48 0.02 80.0

Statewide Total 66.2 0.02 3,928

Table 3: Estimates of the Actual Usage of Acequinocyl on Agricultural Crops (USEPA, 2014a).

Crop

Average Percent Crop Treated

Lbs. A.I. Average Maximum

Avocados <500 <2.5 5

Grapefruit <500 <1 <2.5

Lemons <500 <1 <2.5

Oranges <500 <1 <2.5

Tangerines <500 <2.5 <2.5

2. Conclusions from Previous Actions

a. Ecological Risk Assessment

Sabadilla alkaloids was last assessed in 2004 for the Registration Eligibility Decision (RED; EPA, 2004a). 

The previous assessment focused on the alkaloids veratridine, cevadine, sabadine, and sabadinine. 

Sabadilline was mentioned in the RED as a component of the alkaloid mixture, but no confirmation of 
7

the existence of this alkaloid could be located in the literature, so it will not be discussed as part of this

document. 

Due to the lack of registrant-submitted data on the fate and effects characteristics, the RED relied on 

EPISuite,5 a collection of physical/chemical property and environmental fate/effects estimation 

programs. ECOSAR, part of the suite of programs, was developed by the EPA’s Office of Pollution 

Prevention Toxics and was utilized to estimate fate characteristics and aquatic toxicity of sabadilla 

alkaloids. ECOSAR employs quantitative structure activity relationships (QSARs) to predict aquatic 

toxicity of veratridine, cevadine, sabadine, and sabadinine based on their structural similarity to 

chemicals for which toxicity has previously been measured. Reliability of the predictive capacity of 

ECOSAR modeling is determined based on how closely the modeling program can estimate the toxicity 

of the parent compound. 

A single open-literature study measured the acute toxicity of the formulated product, Red Devil Dust 

(0.8% ai; currently non-registered formulation) on bluegill sunfish (MRID 25601008). The toxicity 

endpoint for bluegill sunfish calculated in this study indicated that Red Devil Dust is four orders of 

magnitude more toxic than the ECOSAR-predicted toxicity values for veratridine, cevadine, sabadine, 

and sabadinine. This indicates that there is too much uncertainty to use ECOSAR-predicted toxicity 

values to qualitatively assess risk to aquatic taxa at this time. The RED (USEPA, 2004a) used ECOSAR to 

provide a rough characterization of risk to aquatic taxa and this approach should not be used in future 

assessments. 

The RED (USEPA, 2004a) characterized risk to terrestrial organisms based on limited studies conducted 

with rats and honeybees. The RED concluded that there may be risks from exposure of total sabadilla 

alkaloids to listed and non-listed mammals. This conclusion was based on an LD50 of 44 mg ai/kg-bw, 

estimated using a probit analysis of data from two acute rat toxicity studies. Since the RED, another 

study with rats has been submitted that reported an endpoint of >10mg ai/kg bw. The RED also 

concluded that sabadilla alkaloids are practically non-toxic on a contact basis to honeybees based on an 

LD50= 12.3 μg formulated product/Bee. The formulation was Veratrine (CAS: 8051-02-3; Purity not 

reported), which is no longer registered. 

b. Drinking Water Exposure Assessments

The most recent drinking water assessment (DWA) was conducted in support of the RED (USEPA, 

2004a). Calculation of the EDWCs was carried out with aerobic soil half-lives that were estimated using 

the Biodegradation Probability Program (BIOWIN) contained in EPIWIN v.3.11 (2000). This program 

estimates the probability of the rapid aerobic biodegradation of an organic chemical in a mixed 

population of microorganisms. 

Tier I Estimated Drinking Water Concentrations (EDWCs) were calculated by incorporating predicted 

degradation values in FIRST v. 1.0 (surface water) and SCIGROW v. 2.3 (ground water). The peak surface 

water concentrations ranged between 1.6 ppb and 1.8 ppb. The average annual surface water 

concentrations ranged between 0.05 ppb and 0.07 ppb, depending on the alkaloid assessed. Ground 

water concentrations were estimated to be 0.0007 ppb. Monitoring data were not available for use in 

this assessment.

 

5 http://www.epa.gov/opptintr/exposure/pubs/episuite.htm
8

Due to the uncertainty related to the use of estimated fate data in the BIOWIN without any empiricallyderived data to compare for accuracy, the EDWC values predicted in that assessment should be 

interpreted with caution and EFED recommends against their use in subsequent risk characterization. 

c. Clean Water Act Programs

Sabadilla alkaloids is not identified as a cause of impairment for any bodies listed as impaired under 

section 303(d) of the Clean Water Act (as of 11/2014).6 A Total Maximum Daily Load (TMDL) has not 

been developed.7 The Agency 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 Process8, in order to ensure they can be used quantitatively or 

qualitatively in pesticide risk assessments.

3. Environmental Fate and Transport

The estimated physical and chemical properties of sabadilla alkaloids: veratridine, cevadine, sabadinine, 

and sabadine are summarized in Table 4. These steroidal alkaloids are characterized by a close structural 

relationship with sterols. They contain a perhydro-1,2-cyclopentanophenanthrene nucleus and are 

often associated with glycosidic complexes in plant tissues, where they hydrolyze to free the alkaloid. 

Veratridine and cevadine are characterized by the presence of an oxygen bridge between carbons 4 and 

9 and a shared nitrogen on the fifth and six rings and terminal esters on ring one. As shown in Table 4,

the compounds are of low water solubility and have high pKa (>9 for veratridine; McKinney et al. 1986). 

No submitted data are available on the physical and chemical properties or the environmental fate of 

sabadilla alkaloids. From the open literature data, sabadilla alkaloids are reported to be very unstable in 

the presence of light and decompose rapidly by photodegradation (Metcalf 1995; Tomlin 2003); 

however, no rates were provided. This qualitative information suggests that photolysis on the surface of 

plants, water, and soil is a likely degradation process. A field trial with Veratran-D formulation showed 

that levels of cevadine and veratridine on leaves declined to 60% from the initial deposit within 20 h of 

application and to undetectable levels within 7 d (Hare and Morse, 1997). Pelletier and Jacobs (1953) 

reported that a gentle saponification of cevadine yields a new alkanolamine, veracevine, which contains 

one double bond and eight hydroxyl groups.

The volatilization of sabadilla alkaloids from soil and water surfaces is not considered an important

environmental pathway based on the estimated vapor pressures and Henry's Law constants. 

The dissociation constants (pKa) of sabadilla alkaloids are above 9 suggesting these compounds tend to 

exist in the neutral, protonated form in water and moist soils, and not as the free base. Based upon the 

estimated Koc values, these alkaloids may have very low mobility in soils. Consequently, leaching to 

groundwater is not expected to be an important environmental fate process. However, log Kow 

estimates are low, indicating that these alkaloids may not sorb substantially to suspended solids and 

sediment in aquatic systems. The environmental fate of sabadilla alkaloids is uncertain in the absence of 

guideline-compliant environmental fate studies. Physical and chemical information is not available for 

sabadilline (USEPA 2004a RED). In the absence of fate data, future assessments will assume stability. 

 

6http://iaspub.epa.gov/tmdl_waters10/attains_nation_cy.cause_detail_303d?p_cause_group_id=885

7http://iaspub.epa.gov/tmdl_waters10/attains_nation.tmdl_pollutant_detail?p_pollutant_group_id=885&p_pollut

ant_group_name=PESTICIDES

8http://www.epa.gov/oppfead1/cb/ppdc/2006/november06/session1-sop.pdf
9

Table 4. Physical and Chemical Properties of Sabadilla Alkaloids 

Parameter Value Reference

Chemical Veratridine USEPA, 2004a

Chemical Name (3beta,4alpha,16beta)-4,12,14,16,17,20-Hexahydroxy-4,9-

epoxycevan-3-yl 3,4-dimethoxybenzoate

EPI Suite v 4.1

CAS Number 71-62-5

Structure

SMILES C[C@H]1CC[C@H]2[C@@]([C@]3([C@H](C[C@]4([C@@H

]5CC[C@H]6[C@]7([C@]5(C[C@]4([C@@H]3CN2C1)O)O[

C@@]6([C@H](CC7)OC(=O)C8=CC(=C(C=C8)OC)OC)O)C)O)

O)O)(C)O

Chem Spider9

Molecular formula C36H51NO11

EPI Suite v 4.1

Molecular weight 673.8

Vapor Pressure (25o C) 1.51 ×10-24 mm Hg

Henry’s Law constant 4.65 ×10-25 atm m3 mol-1

Log KOW 0.678

KOC 3.2x 105

Water Solubility mg/L) 141

Dissociation constant 

(pKa)

9.54 RED

Chemical Cevadine USEPA, 2004a

Chemical Name (3beta,4alpha,9beta,16beta)-4,12,14,16,17,20-

Hexahydroxy-4,9-epoxycevan-3-yl (2Z)-2-methyl-2-

butenoate

EPI Suite v 4.1

CAS Number 62-59-9

Structure

 

9 CSID:391289, http://www.chemspider.com/Chemical-Structure.391289.html (accessed 15:06, Jun 4, 2015)
10

Table 4. Physical and Chemical Properties of Sabadilla Alkaloids 

Parameter Value Reference

SMILES C/C=C(/C)\C(=O)O[C@H]1CC[C@]2([C@H]3[C@@]1(O[C

@@]24C[C@]5([C@@H]6CN7C[C@H](CC[C@H]7[C@@]([

C@]6([C@H](C[C@]5([C@@H]4CC3)O)O)O)(C)O)C)O)O)C

Chem Spider

Molecular formula C32H49NO9

EPI Suite v 4.1

Molecular weight 591.7

Vapor Pressure (25o C) 5.68 ×10-20 mm Hg

Henry’s Law constant 8.69 ×10-22 atm m3 mol-1

Log KOW 0.89

KOC 2.0 x 105

Water Solubility 

(mg/L)

320.4

Dissociation constant 

(pKa)

9.2 RED

Chemical Sabadinine USEPA, 2004a

Chemical Name (3alpha,4alpha,16beta)-4,9-Epoxycevane3,4,12,14,16,17,20-heptol

EPI Suite v 4.1

CAS Number 124-98-1

Structure

SMILES C[C@H]1CC[C@H]2[C@@]([C@]3([C@H](C[C@]4([C@@H

]5CC[C@H]6[C@]7([C@]5(C[C@]4([C@@H]3CN2C1)O)O[

C@@]6([C@@H](CC7)O)O)C)O)O)O)(C)O

Chem Spider

Molecular formula 509.65

EPI Suite v 4.1

Molecular weight C27H43NO8

Vapor Pressure (25o C) 3.0 ×10-19 mm Hg

Henry’s Law constant 1.0 ×10-20 atm m3 mol-1

Log KOW -1.20

KOC 1.3 x105

Water Solubility mg/L) 6.727 x 104

Dissociation constant 

(pKa)

9.2 RED

Chemical Sabadine USEPA, 2004a

Chemical Name (3beta,4alpha,16beta)-4,12,14,16,17,20-

Hexahydroxycevan-3-yl acetate EPI Suite v 4.1

CAS Number 124-80-1
11

Table 4. Physical and Chemical Properties of Sabadilla Alkaloids 

Parameter Value Reference

Structure

SMILES C[C@H]1CC[C@H]2[C@@]([C@]3([C@H](C[C@]4([C@@H

]5CC[C@@H]6[C@@H]([C@H](CC[C@@]6([C@H]5C[C@]

4([C@@H]3CN2C1)O)C)OC(=O)C)O)O)O)O)(C)O

Chem Spider

Molecular formula 537.7

EPI Suite v 4.1

Molecular weight C29H47NO8

Vapor Pressure (25o C) 1.26 ×10-19 mm Hg

Henry’s Law constant 1.37 ×10-18 atm m3 mol-1

Log KOW 0.32

KOC 1.6x105

Water Solubility mg/L) 2232

Dissociation constant 

(pKa)

9.2 RED

4. Receptors

Consistent with the process described in the Overview Document (USEPA, 2004b), a risk assessment for 

sabadilla alkaloids will rely on a surrogate species approach. Toxicological data generated from 

surrogate test species, which are intended to be representative of broad taxonomic groups, will be used 

to extrapolate the potential effects on a variety of species (receptors) included under these taxonomic 

groupings. Table 5 provides a summary of the available endpoints that could be used to characterize the 

potential effects of sabadilla alkaloid exposure. 

No registrant-submitted data are available for toxicity of sabadilla alkaloids to aquatic species. Open 

literature suggested a formulated product containing sabadilla alkaloids, Red Devil Dust® (0.8% ai), is 

very highly toxic to fish, however Red Devil Dust is no longer registered for use. It is unclear how this 

toxicity is related to the toxicity of the individual components of the mixture, as the mixture was not 

characterized. The RED estimated an LD50 44 mg ai/kg-bw for acute exposure to rats based the probit 

analysis of two studies provided by the California Environmental Protection Agency in 2001. The validity 

of these data could not be confirmed, as the original studies and reviews of these studies could not be 

located. A registrant-submitted study conducted with rats using Veratran-D did not record any toxicity 

effects up to the maximum dosage of 10 mg formulation/kg-bw. The authors did not correct for % a.i, so 

the LD50 for the sabadilla alkaloids contained in Veratran-D, assuming a 0.2% a.i. as written on the 

current label, would be >10 mg ai/kg-bw. As a result of this non-definitive endpoint, the acute endpoint 

for rats is presumed to be 44 mg ai/kg-bw. Sabadilla alkaloids is practically nontoxic to adult honeybees 

on an acute contact basis.
12

Table 5: Available Toxicity Endpoints for Sabadilla Alkaloids

Taxa Study Type % ai Species Toxicity Value & 

Category

MRID & 

Status

Endpoints 

Affected

Freshwater 

fish 

Acute 

850.1075

TEP

0.8

Bluegill Sunfish

Lepomis 

macrochirus

LC50=89 μg ai1/L 

NOAEC <49 μg ai/L 

Very Highly Toxic

25601008

Supplementalquantitative

Characterized 

toxicity of TEP 

(Red Devil 

Dust 

formulation 

that is no 

longer 

registered)

Mortality

Mammals 

Acute oral 

870.1100

TEP 

(0.2%)2

Rat 

Rattus norvegicus

LD50 >5000 mg 

Veratran-D/kg-bw or 

LD50 >10 mg ai/kg-bw

46372101

SupplementalQualitativeSabadilla 

alkaloids 

present in 

mixture could 

not be 

characterized

None

Terrestrial 

invertebrates

Adult Acute 

Contact

850.3020

NR Honeybee Apis 

mellifera

48-hr LD50= 12.33 μg 

formulation/bee

Practically Nontoxic

00001999

SupplementalQualitative, 

Characterized 

toxicity of TEP 

(Veratrine, a

formulation 

that is no 

longer 

registered. 

Purity was not 

reported)

None

2Assumed based on current Veratran-D label. 

NR=Not reported

The RED (USEPA, 2004a) characterized toxicity values using the Ecological Structure Activity Relationship 

(ECOSAR) program within the EPI Suite of environmental fate/effects programs. EPI Suite is a screeninglevel tool and should not be used if acceptable measured values are available. Because of the lack of 

empirically-derived data assessing the toxicity of the various alkaloids to aquatic taxa, there is no basis 

for assessing the reliability of the predicted values for the other aquatic taxa until more data are 

submitted. 
13

a. Degradate toxicity

The degradate veracevine is formed from the hydrolysis of veratridine (Pelletier and Jacobs, 1953). Hare

and Morse (1997) compared the toxicity of the insecticidal components of Veratran-D, veratridine and 

cevadine, with that of the degradate, veracevine. The authors reported that veracevine was 500-3,000 

times less toxic than cevadine and veratridine to the adult female citrus thrip, Scirtothrips citri. Figure 1 

compares the toxicity of veratridine, cevadine and veracevine to the target pest species, thrips. EFED will 

conduct a formal review of this study to ensure that the methods and information reported are suitable 

for use in risk assessment. 

Figure 1: Comparison of the Toxicity of Veratridine, Cervadine and degradate Veracevine to 

Citrus Thrips (Hare and Morse, 1997)

b. Ecological Incidents

The Ecological Incident Information System or EIIS maintained by the Environmental Fate and Effects 

Division, and the Avian Incident Monitoring System (AIMS) maintained by the American Bird 

Conservancy, were searched on June 1, 2015 and no ecological incidents were recorded resulting from 

the use of sabadilla alkaloids.

In addition to the incidents recorded in EIIS and AIMS, additional incidents are reported to the Agency in 

aggregated form. Pesticide registrants report certain types of incidents to the Agency as aggregate 

counts of incidents occurring per product per quarter. Ecological incidents reported in aggregate reports 

include those categorized as ‘minor fish and wildlife’ (W-B), ‘minor plant’ (P-B), and ‘other non-target’ 

(ONT) incidents. ‘Other non-target’ incidents include reports of adverse effects to insects and other 

terrestrial invertebrates. There have been no reported incident summaries for sabadilla alkaloids as of 

June 1, 2015.
14

5. Exposure Pathways of Concern 

EFED's environmental exposure models estimate the potential exposure of plants and animals to 

pesticide residues in aquatic and terrestrial environments based on registered labels including maximum 

sabadilla alkaloid application rates and intervals. Aquatic models can also be utilized to estimate 

pesticide residues in drinking water for use in the dietary exposure models. A more detailed description 

of aquatic and terrestrial models can be found at the following website: 

http://www.epa.gov/pesticides/science/models_db.htm. 

The Screening Imbibition Program (SIP v.1.0) is used during the problem formulation phase of

Registration Review to provide an upper-bound estimate of exposure of birds and mammals to 

pesticides through drinking water alone. SIP does not aggregate the drinking water exposure route with 

other exposure routes (i.e., dietary, inhalation, dermal).

The Screening Tool for Inhalation Risk (STIR v.1.0) is used during the problem formulation phase of 

Registration Review to assess the potential for exposure to birds and mammals through inhalation. The 

exposure pathways that are assessed by this tool include both droplet inhalation and vapor phase 

inhalation. STIR is intended to determine if exposure is likely and not whether or not the potential for 

risk exists. If STIR predicts that exposure is likely, additional inhalation data may be necessary to 

adequately assess risk due to the inhalation exposure pathway. 

Due to a lack of toxicological endpoints and information on the chemical properties of sabadilla 

alkaloids, SIP and STIR could not be used to predict the potential for exposure to non-target species 

through inhalation or drinking water.

6. Analysis Plan

a. Stressors of Concern

The stressors of ecological concern for terrestrial/aquatic plants and animals are the primary 

components of the commercially available formulation of sabadilla alkaloids: veratridine, cevadine, 

sabadine, and sabadinine. Sufficient data are not available to exclude any alkaloids in the sabadilla 

alkaloid mixture as potential stressors of concern.

i. Ecological Risk Assessment

Open literature indicates that the primary insecticidal actors of the sabadilla alkaloid mixture are 

veratridine and cevadine (Pelletier and Caventou, 1819; Allen et al. 1945; Ikawa at al., 1945; Walton, 

1947; Bergmann et al., 1958; Hare and Morse, 1997; Humeres and Morse, 2006). Hare and Morse (1997) 

show that the toxicity of each varies among species. 

In the absence of the requested studies, EFED may not be able to assess risk to certain taxa or may need 

to rely on data with tremendous uncertainty. It may be possible to use toxicity endpoints from chemicals 

with structural similarities or modes of action, but this course of action could overestimate risk to 

certain taxa. For example, sabadilla alkaloid’s effect on sodium nerve channels is a mode of action 

shared with the pyrethrins and pyrethroids. 

Upon receiving the Tier I aquatic and terrestrial toxicity studies, toxicity endpoints for sabadilla alkaloids 

will be approximated by deriving a toxicological endpoint for each of the four constituents on a 
15

proportional basis, assuming that the toxicity of the mixture is conservatively explained by the amount 

of each constituent separately. In this manner, the observed toxicity of the mixture could be adjusted

based on the amount of each of the four compounds. These compound-specific estimates of toxicity can 

be compared with the relative expected amounts of each constituent in environmental media and could 

account for differences in formulation composition as well as exposure refinements resulting from 

submission of fate data. This approach also accounts for any possible synergistic effects that would not 

be reflected if alkaloid components were tested individually. In the absence of fate data, stability of the 

alkaloids will be assumed for modeling purposes. 

ii. Drinking Water Assessment

The last drinking water assessment (DWA) was conducted in 2004 as part of the RED (DP 304789). A new 

DWA will be conducted as part of the registration review, which will include the alkaloids veratridine 

and cevadine as well as any other alkaloids or degradates of concern as identified using guidelinecompliant environmental fate and toxicity data. 

b. Measures of Exposure

EFED will use standard available exposure models to evaluate potential exposures to aquatic and 

terrestrial organisms as described at http://www.epa.gov/pesticides/science/models_db.htm. 

i. Available Monitoring Data

EFED will consider available monitoring data from federal and state agencies or other sources. The 

Agency welcomes submission of monitoring data.

c. Measures of Effect

Ecotoxicity data presented in Table 5 of this problem formulation may be used to calculate risk 

quotients. Any additional information submitted by the registrant or found in the open literature prior 

to conducting the risk assessment will also be considered. The open literature studies will be identified 

using EPA’s ECOTOXicology database (ECOTOX10), which employs a literature search engine for locating 

chemical toxicity data for aquatic life, terrestrial plants, and wildlife. 

7. Endangered Species Assessment

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

risks to federally listed threatened and endangered (listed) species from registered uses of pesticides in 

accordance with the Joint Interim Approaches developed to implement the recommendations of the 

April 2013 National Academy of Sciences (NAS) report, Assessing Risks to Endangered and Threatened 

Species from Pesticides. The NAS report outlines recommendations on specific scientific and technical 

issues related to the development of pesticide risk assessments that EPA and the Services must conduct 

in connection with their obligations under the ESA and FIFRA. In November 2013, EPA, the U.S. Fish and 

Wildlife Service, National Marine Fisheries (the Services), and USDA released a white paper containing a 

summary of their joint Interim Approaches for assessing risks to listed species from pesticides. These 

Interim Approaches were developed jointly by the agencies in response to the NAS recommendations, 

and reflect a common approach to risk assessment shared by the agencies as a way of addressing 

scientific differences between the EPA and the Services. Details of the joint Interim Approaches are 

contained in the November 1, 2013 white paper, Interim Approaches for National-Level Pesticide 

 

10 http://cfpub.epa.gov/ecotox/
16

Endangered Species Act Assessments Based on the Recommendations of the National Academy of 

Sciences April 2013 Report.

Given that the agencies are continuing to develop and work toward implementation of the Interim 

Approaches to assess the potential risks of pesticides to listed species and their designated critical 

habitat, this ecological problem formulation supporting the Preliminary Work Plan for sabadilla alkaloids

does not describe the specific ESA analysis, including effects determinations for specific listed species or 

designated critical habitat, to be conducted during registration review. While the agencies continue to 

develop a common method for ESA analysis, the planned risk assessment for the registration review of 

sabadilla alkaloids will describe the level of ESA analysis completed for this particular registration review 

case. This assessment will allow EPA to focus its future evaluations on the types of species where the 

potential for effects exists, once the scientific methods being developed by the agencies have been fully 

vetted. Once the agencies have fully developed and implemented the scientific methods necessary to 

complete risk assessments for listed species and their designated critical habitats, these methods will be 

applied to subsequent analyses of sabadilla alkaloids as part of completing this registration review.

8. Endocrine Disruptors Screening Program 

Sabadilla alkaloids was not included in either the first or second group of chemicals issued an order to 

conduct Tier I EDSP testing, for more information on the EDSP program, visit 

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

9. Preliminary Identification of Data Gaps

Table 6 presents a list of the fate and effects data gaps, as well as a list of studies that EFED is 

recommending for submission. Aquatic and terrestrial toxicity studies will characterize the toxicity of the

active ingredient found in Veratran-D. Sabadilla alkaloids is derived from dried seeds of plants in the lily 

family and it has been reported that there is significant variation (>3 fold) in the concentrations of 

veratridine and cevadine depending on the year that the product was produced (Hare and Morse, 1997). 

As a result, toxicity studies on aquatic and terrestrial taxa must submit a report of the % composition of 

veratrine, cevadine, sabadine, and sabadinine contained in the test mixture, and a consistent test 

mixture composition must be maintained across all tests. Terrestrial plant seedling emergence and 

growth studies should be conducted with Veratran-D TEP. 

The recommendations include representative species from each taxonomic group that includes fish, 

aquatic invertebrates, terrestrial plants, aquatic plants, and birds. The requested data reflect a tiered 

approach to characterizing risk of sabadilla alkaloids to terrestrial and aquatic taxa. The first tier of 

requested studies will be used to confirm that no risk exceedances exist (i.e. RQ>LOC) to representative 

terrestrial and aquatic taxa. If risk concerns for a particular taxa are identified, then a second tier of data

for that taxa is recommended. 

In the absence of fate data, future assessments will assume stability for sabadilla alkaloids. The tier I set 

of fate studies will characterize the fate properties of the largest molecule, veratridine, which is 

suspected to be the final product of the biosynthesis pathway in the sabadilla lily seeds. If risk concerns 

are identified for any taxa, Tier II fate data will characterize the environmental fate characteristics of the 

second primary insecticidal component of sabadilla alkaloids, cevadine.
17

Table 6. Environmental Fate and Effects Data Gaps and Recommended Studies for Sabadilla Alkaloids

Data Requirement Data 

Gap?

Tier I Tier II Test 

Substance

Comments

850.1010

Freshwater Invertebrate 

Acute Toxicity 

Yes Yes No TGAI11--

850.1035

Estuarine/Marine 

Invertebrate

Yes No Yes TGAI --

850.1025 

Oyster Toxicity 

Yes No Yes TGAI --

850.1300

Freshwater Invertebrate Life 

Cycle (Chronic) 

Yes Yes No TGAI --

850.1350

Estuarine/Marine 

Invertebrate Life Cycle

Yes No Yes TGAI --

850.1735 Subchronic 

Freshwater Sediment 

Toxicity Test

Yes No Yes TGAI Sediment invertebrate toxicity 

data are required under 40 CFR 

Part 158 if any of the following 

exist:

A. The soil partition coefficient 

(Kd) is ≥ 50

B. The log Kow is ≥ 3.

C. The Koc is ≥ 1,000.

The Koc for Veratridine is 

32,000, 20,000 for cevadine, 

13000 for sabadinine, and 

16000 for sabadine, indicating 

the potential for risk to benthic 

invertebrates. Because of the 

lack of half-life values, it is 

uncertain whether a subchronic 

or chronic toxicity would better 

characterize risk to benthic 

organisms. As a result, a tiered 

testing strategy (as outlined in 

USEPA, 2014b) may be 

850.1740 Subchronic 

Estuarine/marine Sediment 

Toxicity Test

Yes No Yes TGAI

Non Guideline Chronic 

Sediment Toxicity Test 

Yes12 No Yes TGAI 

 

11 Sabadilla Alkaloids (TGAI) refers to a mixture of sabadilla alkaloids that would be representative of the active 

ingredient in a typical commercially available formulation of Veratran-D. As sabadilla alkaloids is a seed powder 

derived from dried seeds of plants in the lily family, it has been reported that there is significant variation (>3 fold) 

in the concentrations of veratridine and cevadine (Hare and Morse, 1997). As a result, studies conducted with 

sabadilla alkaloid TGAI should be carried out with a formulation of sabadilla alkaloids where the concentrations of 

veratridine, cevadine, sabadinine, and sabadine in the mixture are consistent for each study and an analytical 

confirmation is provided in the study report. Registrant needs to submit confirmatory analysis of the test 

substance to confirm that sabadilla alkaloids TGAI is representative of the composition of a typical product.
18

Data Requirement Data 

Gap?

Tier I Tier II Test 

Substance

Comments

considered for implementing 

the necessary Data Call-In 

requirements. In this approach, 

sediment toxicity testing would 

be conducted using the 

subchronic (10-d) sediment 

toxicity test guidelines (OSCPP 

850.1735 and 850.1740). 

Pending a comparison of 

estimated environmental 

concentrations (EEC) in 

sediment and pore water with 

toxicity results from the 10-d 

subchronic sediment toxicity 

studies, one or more chronic 

sediment toxicity studies would 

be recommended. 

850.1075

Freshwater Fish Acute 

Toxicity 

Yes Yes Yes TGAI Tier I- Coldwater Species

Tier II-Warmwater Species

850.1075

Estuarine/Marine Fish Acute 

Toxicity 

Yes No Yes TGAI --

850.1400 

Freshwater Fish Early-Life 

Stage 

Yes Yes Yes TGAI Tier I- Coldwater Species

Tier II-Warmwater Species

850.1400

Estuarine/Marine Fish Early 

Life Stage

Yes No Yes TGAI --

850.4400

Aquatic Vascular Plant 

Toxicity Test using Lemna 

spp.

Yes Yes No TGAI --

850.4500

Algal Toxicity

Pseudokirchneriella 

subcapitata

Yes Yes No TGAI --

850.4500

Marine diatom Skeletonema

costatum

Yes No Yes TGAI --

850.4500

Freshwater diatom Navicula 

pelliculosa

Yes No Yes TGAI --

850.4550

Blue Green Algae/ 

Cyanobacterium

Yes No Yes TGAI --
19

Data Requirement Data 

Gap?

Tier I Tier II Test 

Substance

Comments

Anabaena flos-aquae

850.2100

Avian Acute Oral Toxicity 

(Upland game, passerine or 

waterfowl species)

Yes Yes Yes TGAI Tier I- Bobwhite Quail

Tier II- Mallard Duck and a 

passerine species

850.2200 Avian Dietary 

Toxicity 

Yes No Yes TGAI Upland game and waterfowl 

species

850.2300

Avian Reproduction 

Yes Yes Yes TGAI Tier I- Bobwhite Quail

Tier II- Mallard Duck

850.3020

Tier I- Honeybee Acute Adult 

Contact Toxicity

No No No NA --

Non-Guideline (OECD 213)-

Tier I Honeybee Acute Adult 

Oral Toxicity

Yes Yes No TGAI --

Tier I- Honeybee Adult 

Chronic Oral Toxicity

Yes Yes No TGAI --

Tier I- Honeybee Larval 

Acute and Oral Exposure

Yes Yes No TGAI --

850.3030-Tier II- Honeybee 

Toxicity of Residues on

Foliage

No No No NA The acute contact 48-hr 

LD50 was 12.33 μg ai/bee. 

This study is only triggered if 

contact toxicity is less than 

11 μg ai/bee.

Non Guideline Tier II-Nectar 

and Pollen Residue Study for 

Insect Pollinators

Yes No Yes TEP --

Non Guideline (OECD 75)-

Tier II Semi-Field Testing for 

Honeybees

Yes No Yes TEP --

850.3040 - Field Testing for 

Pollinators

Yes No Yes TEP --

850.4100- Seedling 

Emergence and Seedling 

Growth

Yes Yes No TEP

850.4150- Vegetative Vigor Yes No Yes TEP

Fate Data

835.2120- Hydrolysis Yes Yes Yes Tier I –

Veratridine

Tier IICevadine

835.2240-Aqueous 

Photolysis 

Yes Yes No Veratridine

835.2410- Soil Photolysis Yes Yes No Veratridine
20

Data Requirement Data 

Gap?

Tier I Tier II Test 

Substance

Comments

835.4100-Aerobic Soil 

Metabolism 

Yes Yes Yes Tier I –

Veratridine

Tier IICevadine

--

835.4300- Aerobic Aquatic 

Metabolism

Yes Yes Yes Tier I –

Veratridine

Tier IICevadine

--

835.4400-Anaerobic Aquatic 

Metabolism 

Yes Yes No Veratridine --

835.1230/835.1240 Leaching 

Adsorption/ Desorption 

Yes Yes No Veratridine --

835.6100- Terrestrial Field 

Dissipation 

Yes Yes No TEP --

850.1730- Accumulation in 

Fish 

Yes Yes No Veratridine --

850.6100- ECM/ILV (soil and 

water) 

Yes Yes No Veratridine --
21

Justifications for Non-Guideline Data Requirements: 

Study Title: Tier I- Pollinator Chronic Oral Toxicity, Adult

Rationale for Requiring the Data

Terrestrial invertebrates are likely to be impacted if exposed to pesticides in various use settings. Pesticide 

residues may be transferred to pollen and/or nectar of treated plants and subsequently brought back to the 

hive. Therefore, potential chronic effects to adult honeybees and other pollinators from oral exposure to 

some pesticides could exist. Currently available toxicity studies do not address possible lethal and sublethal 

effects of chronic oral exposure on adult terrestrial invertebrates and will assist in determining whether the 

sensitivity of adult bees differs from that of earlier life stages. Because of the potential for pollen and nectar 

to be contaminated with pesticide residues, and subsequently brought back to the hive, it is important to 

determine the chronic oral toxicity of this compound to adult honeybees and other pollinators.

The Office of Pesticide Programs has made available a guidance regarding ecological testing for 

invertebrates with the honeybee. The guidance discusses Tier I laboratory-based chronic oral toxicity 

studies of individual adult honeybees as a critical component of the screening-level risk assessment process 

for examining potential risks from specific routes of exposure. The guidance can be found at: 

http://www2.epa.gov/pollinator-protection/pollinator-risk-assessment-guidance . Study design elements 

for the chronic 10-day oral toxicity test with honeybees are similar to the OECD Test Guideline 213 acute 

oral toxicity test http://www.oecd-ilibrary.org/environment/test-no-213-honeybees-acute-oral-toxicitytest_9789264070165-en .

Practical Utility of the Data

How will the data be used?

The Tier I chronic oral toxicity data on adult bees serve as a foundation for the screening-level assessment 

of potential risk to non-target organisms including federally listed threatened or endangered species and 

non-listed terrestrial invertebrate insects, including pollinators, from chronic oral exposures to pesticides. 

The data will be used to reduce uncertainties associated with the risk assessment for terrestrial 

invertebrates and will improve EPA’s understanding of the potential direct and indirect lethal and sublethal 

effects on a broad range of terrestrial species, particularly insect pollinators and to determine whether 

adult toxicity differs substantially from other life stages evaluated in other Tier I tests. If chronic oral effects 

data for adults are not available, risks to terrestrial insects from chronic oral exposure will be assumed.

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

The data will inform the determination required under FIFRA or the ESA as to whether continued 

registration of a pesticide is likely to result in unreasonable adverse effects to non-target species or is likely 

to adversely affect listed threatened or endangered species and/or their designated critical habitat. 

Without these data, EPA may need to presume risk which will limit the flexibility of pesticide products to 

comply with FIFRA and the ESA, and could result in use restrictions. 

Study Title: Tier I- Pollinator Acute Oral Toxicity, Larvae

Rationale for Requiring the Data

Terrestrial invertebrates are likely to be impacted if exposed to pesticides in various use settings. Pesticide 

residues may be transferred to pollen and/or nectar of treated plants and subsequently brought back to the 

hive where larvae and pupae may be exposed. Therefore, potential adverse effects to developing bees and 

other insect pollinators could result from exposure to pesticide residues. Available toxicity studies do not 

address possible effects on brood (larvae and pupae) survival/development. Because of the potential for 

pollen and nectar to be contaminated with pesticide residues, and subsequently brought back to the hive, it 

is important to determine the acute oral toxicity of this compound to bee brood. 
22

The Office of Pesticide Programs has made available a guidance regarding ecological testing for terrestrial 

invertebrates using the honeybee as a surrogate test species. The guidance discusses Tier I laboratorybased acute toxicity studies of individual honeybee larvae as a critical component of the screening-level risk 

assessment process for examining potential risks from specific routes of exposure. The guidance can be 

found at: http://www2.epa.gov/pollinator-protection/pollinator-risk-assessment-guidance . Additional 

guidance on larval honeybee toxicity test design can be found in OECD Test Guideline 237 

http://www.oecd-ilibrary.org/environment/test-no-237-honey-bee-apis-mellifera-larval-toxicity-test-singleexposure_9789264203723-en .

Practical Utility of the Data

How will the data be used?

The Tier I acute oral toxicity data on honeybee larvae serve as a foundation for the screening-level 

assessment of potential risk to non-target organisms such as federally listed threatened or endangered and 

non-listed terrestrial invertebrate insects, including pollinators, and/or modify their designated critical 

habitat from acute oral exposures to pesticides. The data will be used to reduce uncertainties associated 

with the risk assessment for terrestrial invertebrates and will improve EPA’s understanding of the potential 

effects on terrestrial species and whether there is a differential sensitivity of larval bees relative to adult 

bees. If acute oral effects data for larvae are not available, risks to terrestrial insects from acute oral 

exposure will be assumed

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

The data will inform the determination required under FIFRA or the ESA as to whether continued 

registration of a pesticide is likely to result in unreasonable adverse effects to non-target species or is likely 

to adversely affect listed threatened or endangered species and/or modify their designated critical habitat. 

Without these data, EPA may need to presume risk which will limit the flexibility of pesticide products to 

comply with FIFRA and the ESA, and could result in use restrictions. 

Study Title: Tier I- Pollinator Chronic Oral Toxicity, Larvae

Rationale for Requiring the Data

Terrestrial invertebrates are likely to be impacted if exposed to pesticides in various use settings. Pesticide 

residues may be transferred to pollen and/or nectar of treated plants and subsequently brought back to the 

hive where larvae and pupae may be exposed. Therefore, potential effects to developing bees could result 

from chronic oral exposure to pesticide residues. Available toxicity studies do not address possible chronic 

effects on brood (larvae and pupae) survival. Because of the potential for pollen and nectar to be 

contaminated with pesticide residues, and subsequently brought back to the hive, it is important to 

determine chronic larval/pupal toxicity and whether adult emergence is adversely affected. This study will 

provide information on whether honeybee larvae differ in sensitivity from adult bees following chronic 

exposure. 

The Office of Pesticide Programs has made available a guidance regarding ecological testing for 

invertebrates with the honeybee. The guidance discusses Tier 1 laboratory-based chronic oral toxicity 

studies of individual honeybee larvae as a critical component of the screening-level risk assessment process 

for examining potential risks from specific routes of exposure. The guidance can be found at: 

http://www2.epa.gov/pollinator-protection/pollinator-risk-assessment-guidance.

Additional information on larval honeybee toxicity repeat exposure test design can be found in the oECD 

draft guidance 
23

http://www.oecd.org/env/ehs/testing/Draft_GD_honeybees_rep_exp_for_2nd_CR_25_November_2013.p

df .

Practical Utility of the Data

How will the data be used?

The Tier I chronic oral toxicity data on bee larvae serve as a foundation for the screening-level assessment 

of potential risk to non-target organisms including federally listed threatened or endangered and non-listed 

terrestrial invertebrate insects, including pollinators, from chronic oral exposures to pesticides. The data 

will be used to reduce uncertainties associated with the risk assessment for terrestrial invertebrates and 

will improve EPA’s understanding of the potential direct and indirect lethal and sublethal effects on a broad 

range of terrestrial species, particularly insect pollinators. These data will also assist in determining 

whether early life stages of the bee differ in their sensitivity to pesticides relative to adults. If chronic oral 

effects data for larvae are not available, risks to terrestrial insects from chronic oral exposure will be 

assumed.

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

The data will inform the determination required under FIFRA or the ESA as to whether continued 

registration of a pesticide is likely to result in unreasonable adverse effects to non-target species or is likely 

to adversely affect listed threatened or endangered species and/or modify their designated critical habitat. 

Without these data, EPA may need to presume risk which will limit the flexibility of pesticide products to 

comply with FIFRA and the ESA, and could result in use restrictions. 

Study Title: Tier II Semi-Field Toxicity Testing (tunnel/enclosure studies)

Rationale for Requiring the Data

Tier II studies are conditional on the outcome of the screening-level assessment where acute and/or chronic 

risk levels of concern have been exceeded for terrestrial invertebrates. Terrestrial invertebrates are likely to 

be impacted if exposed to pesticides in various use settings. Pesticide residues may be transferred to pollen 

and/or nectar of treated plants and subsequently brought back to the hive and may adversely affect 

developing brood (egg, larvae, and pupae) and adult bees. Screening-level (Tier I) studies of individual bees 

do not address possible effects and/or exposure to pesticide residues at the colony-level. Because of the 

potential for pollen and nectar to be contaminated with pesticide residues, and subsequently brought back 

to the hive, it is important to determine whether bee colonies may be negatively affected under relatively 

controlled exposure conditions of a semi-field study. In addition to providing effects data, these studies can 

provide data on exposure as pesticide residues in pollen/nectar of treated plants.

The Office of Pesticide Programs has made available a guidance regarding ecological testing for 

invertebrates with the honeybee. The guidance describes the tiered testing process and can be found at: 

http://www2.epa.gov/pollinator-protection/pollinator-risk-assessment-guidance . Additional information 

on honeybee colony studies under semi-field conditions can be found in the OECD Guidance 75 

http://www.oecd.org/officialdocuments/publicdisplaydocumentpdf/?cote=env/jm/mono%28 . 

Practical Utility of the Data

How will the data be used? 

Tier II colony-level data will be used to assess potential risk to non-target organisms including listed and 

non-listed terrestrial social invertebrate species and to determine whether effects observed in the 

screening-level (Tier I) laboratory-based studies of individual bees are evident in colony-level studies under 

semi-field conditions. The Tier II semi-field test of whole colonies is a relatively controlled study, i.e., bees 

are confined to a specific area, that is designed to represent potential field-level exposure and account for 

hive dynamics, which are not achievable from other pollinator studies. This study will be used to determine 
24

whether adverse effects to insect pollinators at the whole colony level, may result for the use of pesticides 

and will help to refine risk estimates derived in the screening-level risk assessment for beneficial terrestrial 

invertebrates. Measured residues in pollen/nectar can also be used to refine risk estimates derived from 

model-based or default values in the screening-level assessment.

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

The data will inform the determination required under FIFRA or the ESA as to whether continued 

registration of a pesticide is likely to result in unreasonable adverse effects to non-target species or is likely 

to adversely affect federally listed threatened or endangered species or their designated critical habitat. 

Without these data, EPA may need to presume risk which will limit the flexibility of pesticide products to 

comply with FIFRA and the ESA, and could result in significant use restrictions.

Study Title: Tier II Semi-Field Toxicity Testing (feeding studies)

Rationale for Requiring the Data

Pesticide residues may be transferred to pollen and/or nectar of treated plants and subsequently brought 

back to the hive, and may adversely affect developing brood (egg, larvae, and pupae) and adult bees. Tier II 

feeding studies are conditional on the outcome of the screening-level assessment where acute and/or 

chronic risk levels of concern have been exceeded for terrestrial invertebrates based on Tier I studies of 

individual bees. Feeding studies utilize free foraging bee colonies that are “dosed” with specific quantities of 

test material and represent a means of ensuring exposure to the test material through spiked pollen and/or 

sugar solutions fed to the colony while still allowing the bees to forage freely. Since bee colonies are not 

confined to enclosures, colonies can be exposed for longer duration periods without subjecting the bees to 

stress that typically results from Tier II tunnel studies. Available toxicity studies of individual bees (Tier 1) 

conducted to support screening-level assessments do not address possible effects and/or exposure to 

pesticide residues at the colony-level. Because of the potential for pollen and nectar to be contaminated 

with pesticide residues, and subsequently brought back to the hive, it is important to determine whether 

bee colonies may be negatively affected where bees are free foraging and have the option to 

collect/consume alternative forage items beyond the spiked food. Since multiple dose levels can be more 

readily tested, feeding studies can help to define dose-response relationships at the whole colony level.

The Office of Pesticide Programs has made available a guidance regarding ecological testing for invertebrates 

with the honeybee. The guidance describes the tiered testing process and can be found at: 

http://www2.epa.gov/pollinator-protection/pollinator-risk-assessment-guidance . Additional information on 

honeybee colony feeding studies can be found in the EPPO Guidance 170 at: 

http://www.nationalbeeunit.com/index.cfm?pageId=187 .

Practical Utility of the Data

How will the data be used?

Tier II colony feeding data will be used to assess potential risk to non-target organisms including listed and 

non-listed terrestrial social invertebrate species. The colony feeding study is designed to represent 

potential field-level exposure and account for hive dynamics using longer duration exposure periods than 

are possible in Tier II tunnel studies. The study will be used to determine whether potential adverse effects 

to insect pollinators at the whole colony level when bees are able to forage naturally beyond the spiked 

food. Results from the feeding study will help to refine the screening-level risk assessment for beneficial 

terrestrial invertebrates that were based on Tier I studies on individual bees. Since feeding studies can help 

to define a dose-response relationship at the colony level, the studies can provide a means of determining 

exposure thresholds below which the likelihood of adverse effects on colonies may be low.
25

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

Tier II colony-level data will be used to refine screening-level risk estimates derived using Tier I laboratorybased data on individual bees. The Tier II data will inform the determination required under FIFRA or the 

ESA as to whether continued registration of a pesticide is likely to result in unreasonable adverse effects to 

non-target species or is likely to adversely affect federally listed threatened or endangered species or their 

designated critical habitat. Without these data, EPA may need to presume risk which will limit the flexibility 

of pesticide products to comply with FIFRA and the ESA, and could result in significant use restrictions.

Study Title: Residues in Pollen and Nectar

Rationale for Requiring the Data

Terrestrial invertebrates are likely to be impacted if exposed to pesticide residues in various use settings. 

Pesticide residues may be transferred to pollen and/or nectar of treated plants and subsequently brought 

back to hive where all life stages may be exposed. For some pesticides, the quantification of pollinatorrelevant residues in treated flowering plants is needed, since pollinators will be exposed to residues from 

either current or prior season applications (due to the potential for residues to accumulate in plants and 

trees). Residues in edible/transportable-to-hive parts of treated trees and plants, including (where 

appropriate), but not limited to, guttation water, sap/resins, whole plant tissue (e.g., leaves, stems), as well 

as blooming, pollen-shedding, and nectar producing parts (i.e., flowers and, if present, extra-floral nectaries) 

of plants may inform the potential for risk. Studies should be designed to provide residue data for crops 

and application methods of concern.

The Office of Pesticide Programs has made available a guidance regarding ecological testing for 

invertebrates with the honeybee. The guidance can be found at: http://www2.epa.gov/pollinatorprotection/pollinator-risk-assessment-guidance.

Practical Utility of the Data

How will the data be used?

Measured residue data will be used to refine conservative estimates of pesticide exposure and reduce 

uncertainties associated with the Tier I exposure assessment by providing direct measurements of pesticide 

concentrations resulting from actual use settings. Measured residues may provide a more realistic 

understanding of exposure through contact or ingestion with which to calculate risk quotients for individual 

bees as well as to characterize exposure to the colony. If measured residue data are not available, risk 

estimates for terrestrial insects will be based on model-generated or default values used to support the 

screening-level assessment.

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

The data will inform the determination required under FIFRA or the ESA as to whether continued 

registration of a pesticide is likely to result in unreasonable adverse effects to non-target species or is likely 

to adversely affect federally listed threatened or endangered species or their designated critical habitat. 

Without these data, EPA will have to rely on conservative estimates of exposure which will limit the 

flexibility of pesticide products to comply with FIFRA and the ESA, and could result in use restrictions. 
26

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