1

OFFICE OF CHEMICAL SAFETY

AND POLLUTION PREVENTION

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C. 20460

MEMORANDUM 

DATE: December 7, 2010

SUBJECT: Revised Hazard Assessment for Tier I Human Health Toxicity in Support of the 

Registration of AMV-1018 Technical, Containing 3-decen-2-one as its Active 

Ingredient (Amends EPA Memorandum from G. Casciano and C. Walsh through 

R. Jones to D. Benmhand dated June 16, 2010).

EPA File Symbol Number: 5481-LAI

Decision Number: 425383

DP Barcode: 373085

PC Code: 068403

CAS Number: 10519-33-2

MRID Numbers: 479426-09 thru -16, -18

FROM: Gina M. Casciano, M.S., Biologist /s/ 12/07/2010

Biochemical Pesticides Branch

Biopesticides & Pollution Prevention Division (7511P)

Colin G. Walsh, M.S., Biologist /s/ 12/07/2010

Biochemical Pesticides Branch

Biopesticides & Pollution Prevention Division (7511P)

THROUGH: Russell S. Jones, Ph.D., Senior Biologist /s/ 12/07/2010

Biochemical Pesticides Branch

Biopesticides & Pollution Prevention Division (7511P)

TO: Driss Benmhand, Regulatory Action Leader

Biochemical Pesticides Branch

Biopesticides & Pollution Prevention Division (7511P) 

ACTION REQUESTED

AMVAC Chemical Corporation requests registration of AMV-1018 Technical, a new technical 

grade active ingredient (TGAI) and manufacturing-use product (MP) containing 98.0% of the 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

2

new active ingredient (a.i.) 3-decen-2-one, a chemical occurring naturally in some foods and 

approved as a direct food additive by the U.S. Food and Drug Administration for use as a 

synthetic flavoring agent and adjuvant (21 CFR 172.515). This TGAI/MP product is for 

manufacturing-use only; specifically, for formulation into registered end-use products for use in 

or on agricultural commodities on growing plants. To fulfill Tier I Human Health Toxicity 

requirements for manufacturing-use products (40 CFR 158.2050), the applicant has submitted 

acute data, mutagenicity data, and data waiver requests for subchronic and developmental testing.

RECOMMENDATIONS AND CONCLUSIONS

The Tier I Human Health Toxicity submission is UNACCEPTABLE but upgradeable. To 

upgrade the submission to Acceptable, the registrant much resolve the deficiencies listed below: 

1. The Acute Toxicity Tier I submission is ACCEPTABLE. Studies submitted to fulfill acute 

toxicity data requirements (Table 1, below) indicate that AMV-1018 Technical is Toxicity 

Category IV for acute oral toxicity and acute dermal toxicity, Toxicity Category III for acute 

inhalation toxicity, and Toxicity Category II for primary eye irritation and primary dermal 

irritation. Additional data are not required. 

2. The residue study submitted with this application is UNACCEPTABLE. Testing for 

residues of 3-decen-2-one and its degradates did not begin until 16 days post-application. 

However, the draft product label for end-use product AMV-1018 EP (EPA File Symbol 

5481-LTR) indicates that potatoes may be shipped to consumers as early as 24 hours posttreatment (this appears under the Sprout Control Programs section on the label). Therefore, 

testing for residues beginning 24 hours after application is required. In addition, the applicant 

did not conduct the residue study using the maximum label application rate. Dietary concerns 

for 3-decen-2-one and its primary degradates, 2-decanol and 2-decanone, remain. 

3. The scientific rationale and cited data submitted to fulfill the 90-day oral toxicity data 

requirement are UNACCEPTABLE. Using the data presented by the applicant in their 

current residue study, the reviewer calculations indicate that the inclusion of a source of 3-

decen-2-one in the diet from pesticidal residues would potentially increase the average daily 

intake of this chemical 4,000 times based on residues found at 16 days post-application. In 

addition, the amount of active ingredient that can be applied to the potatoes, based on the 

maximum label application rates, exceeds the daily dose amounts tested in the 90-day oral 

studies cited for the three structurally similar compounds. Additional data and/or data waiver 

rationale must be submitted to determine if the observed residues, including residues at 24 

hours post-application, are of toxicological concern. 

4. The data waiver request submitted to fulfill the 90-day inhalation toxicity data requirement is 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

3

UNACCEPTABLE. The data waiver request is only valid if repeated occupational exposure 

via the inhalation route is not expected. This is the case when 3 -decen-2-one is used in 

warehouse air circulation systems (thermal fogging) and the storage facilities remains closed 

for 24 hours to personnel entry while the thermal fogging in the treated area is re-circulated 

through the potato pile. No instructions are provided on the AMV-1018EP (EPA File Symbol 

5481-LTR) label that indicates when the worker is allowed to reenter the treated area once 

the ventilation louvers are reopened to the outside after the 24 hour thermal fogging 

application. The occurrence of inhalation exposure is not clear of the other application 

method, spray drenching, for AMV-1018 90EC (EPA File Symbol 5481-LTN) and AMV1018 67.5EC (EPA File Symbol 5481-LAO). Appropriate precautions to mitigate inhalation 

exposure, including any applicable Restricted Entry Interval (REI) and Personal Protective 

Equipment (PPE), for all use patterns must be included on all product labels (see NOTES TO 

RAL 3 and 4, below).

5. The scientific rationale submitted to fulfill the developmental toxicity data requirement is 

UNACCEPTABLE. Significant dietary exposure is expected to occur and it is unclear as it 

whether or not residues will be of toxicological concern. Additional data and/or data waiver 

rationale for developmental toxicity must be submitted.

6. The mutagenicity data submitted to fulfill this data requirement are UNACCEPTABLE. 

Study results from the Mouse Lymphoma Assay are equivocal and no other test for 

mammalian gene mutation or chromosomal aberration was performed. The applicant must 

resolve this discrepancy.

NOTES TO RAL:

1. The Acute Toxicity data submission (see Conclusion 1, above) places AMV-1018 Technical

in Toxicity Category II.

2. The label for end-use product AMV-1018 EP (EPA File Symbol 5481-LTR) indicates that 

potatoes may be shipped to consumers as early as 24 hours post-treatment (this appears under 

the Sprout Control Programs section on the label). The labels for AMV-1018 90EC (EPA 

File Symbol 5481-LTN) and AMV-1018 67.5EC (EPA File Symbol 5481-LAO) do not 

include this information. The applicant must resolve this discrepancy. 

3. The labels for AMV-1018 90EC (EPA File Symbol 5481-LTN) and AMV-1018 67.5EC 

(EPA File Symbol 5481-LAO) mention the use pattern of thermal fogging (under the Use 

Precautions Sections) but do not include instructions (or worker protection language) for this 

type of application. The applicant must include complete application instructions and worker 

protection language for thermal fogging if this is an intended use pattern for these two 

products. In addition, the REI following thermal fogging should be included on all labels.
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

4

4. Since a 90-day inhalation study was not submitted, this use of AMV-1018 90EC (EPA File 

Symbol 5481-LTN) and AMV-1018 67.5EC (EPA File Symbol 5481-LAO) as spray 

solutions, as indicated on these product labels, must be accompanied by appropriate worker 

PPE to mitigate exposure (unless use is restricted to automated spray equipment only) and 

any applicable REI. Currently, there are no applicator PPE requirements for inhalation on the 

draft product labels. 

5. The amount, frequency, and timing of the proposed application of this biochemical were not

included in Tolerance Petition 9F7670. This information appears on the label for AMV-1018 

EP (EPA File Symbol 5481-LTR) only. The labels for AMV-1018 90EC (EPA File Symbol 

5481-LTN) and AMV-1018 67.5EC (EPA File Symbol 5481-LAO) do not include this 

information. The applicant must resolve this discrepancy.

SUMMARY OF DATA SUBMITED 

Toxicological Profile of 3-Decen-2-one and Hazard Assessment

The data presented in Table 1, below, are a summary of the toxicity data and information 

submitted to support the registration of the proposed manufacturing-use product AMV-1018 

Technical (EPA File Symbol No. 5481-LAI), with contains 98.0% 3-decen-2-one as its active 

ingredient. At this time, all data requirements have been adequately satisfied with the exception 

of 90-day oral toxicity, 90-day inhalation toxicity, developmental toxicity, and mutagenicity.

Table 1: Mammalian Toxicology Data Requirements for 3-decen2-one (40 CFR § 158.2050)

Study/OCSPP Guideline No. Results Toxicity 

Category/Description

MRID

Acute oral toxicity (rat)

(870.1100)

LD50 > 5,000 mg/kg body weight in female 

rats Toxicity Category IV 47942609

Acute dermal toxicity (rat)

(870.1200)

LD50 > 5,000 mg/kg body weight in male and 

female rats Toxicity Category IV 47942610

Acute inhalation toxicity (rat)

(870.1300)

LC50 0.52-2.04 mg/L in male rats and > 2.04 

mg/L in female rats Toxicity Category III 47942611

Primary eye irritation (rabbit)

(870.2400)

Test substance is moderately irritating 

to the eye. Toxicity Category II 47942612

Primary dermal irritation (rabbit)

(870.2500)

Test substance is severely irritating 

to the skin. Toxicity Category II 47942613

Dermal sensitization (guinea pig)

(870.2600) Test substance is not a dermal sensitizer. - 47942614

Hypersensitivity incidents

(885.3400) None reported. - -
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

5

Table 1: Mammalian Toxicology Data Requirements for 3-decen2-one (40 CFR § 158.2050)

Study/OCSPP Guideline No. Results Toxicity 

Category/Description

MRID

90-Day oral toxicity

(870.3100)

Waiver rationale submitted.

UNACCEPTABLE; 

Additional data and/or 

data waiver rationale 

is required (see 

discussion below).

47942617

90-Day dermal toxicity

(870.3250)

Not required; uses not to involve purposeful 

application to human skin. - 47942617

90-Day inhalation toxicity

(870.3465)

Waiver rationale submitted.

UNACCEPTABLE; 

Additional data and/or 

data waiver rationale 

is required (see 

discussion below).

47942617

Developmental toxicity

(870.3700)

Waiver rationale submitted.

UNACCEPTABLE; 

Additional data and/or 

data waiver rationale 

is required (see 

discussion below).

47942617

Mutagenicity

(870.5100, 5300 and 5375)

Test substance is negative in bacterial reverse

mutation assay. Test substance positive in 24-

hr exposure without metabolic activation in an 

in vitro mammalian gene mutation assay

(mouse lymphoma cell). 

UNACCEPTABLE;

Equivocal results are 

not adequate to fulfill 

data requirement (see 

discussion below).

47942615; 

47942616

Acute Toxicity

All Tier I Human Health Toxicity requirements for Acute Testing were satisfied with studies 

conducted according to OCSPP Harmonized Test Guidelines. All studies contained signed and 

dated Data Confidentiality, Good Laboratory Practice (GLP) Compliance, and Quality Assurance 

statements were provided. A Data Evaluation Record (DER) was written for each submitted 

study. DERs appear at the end of this memorandum. The results of the Acute Toxicity testing

(summarized in Table 1, above) place AMV-1018 Technical in Toxicity Category II. No 

hypersensitivity incidents or other adverse effects data was reported for the TGAI/MP.

CLASSIFCATION: These data are ACCEPTABLE; no additional data are required.

Subchronic Toxicity: 90-day Oral Toxicity

To satisfy the data requirement for 90-day oral toxicity, the applicant has submitted data waiver 

rationale stating that 3-decen-2-one: 1) occurs naturally in the diet; 2) is approved for use as a 

direct food additive by the U.S. FDA; 3) is not reported to be associated with subchronic toxicity; 

4) is metabolized into innocuous compounds; and 5) is structurally similar to compounds that did 

not cause adverse effects in rats exposed for 90-days. According to the results of the applicant 

performed residue study, the inclusion of a source of 3-decen-2-one in the diet from pesticidal 

use could potentially increase the current average daily intake of this chemical 4,000 times based 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

6

on residues found at 16 days post application. In addition, the amount of active ingredient that 

can be applied to the potatoes, based on the maximum label application rates, exceeds the daily 

dose amounts tested in the 90-day oral studies cited for the three structurally similar compounds.

Therefore, the Agency is requiring additional data and/or data waiver rationale to be submitted to 

determine if the residues observed, including residues at 24 hours post-application, are of 

toxicological concern. 

The applicant cites the occurrence of 3-decen-2-one in edible porcini mushrooms (Thomas, 

1973) and Iberian ham (Timon et al., 1998) (quantitative data not available) without report of 

incident or safety concern. Its use as a synthetic flavoring agent and adjuvant (21 CFR 172.515) 

indicates that it may be commonly consumed in other foods, however, specific information 

regarding this was not provided.

The Joint FAO/WHO Expert Committee on Food Additives (JEFCA) has characterized the 

metabolic pathways of 3-decen-2-one and determined that metabolism of the chemical results in 

the production of innocuous metabolites. Specifically, 3-decen-2-one is reduced to its 

corresponding alcohol, which is subsequently conjugated with glucuronic acid and excreted 

(JEFCA, 2003). In the environment, 3-decen-2-one degrades into the compounds 2-decanol and 

2-decanone. The Agency agrees with this characterization. 

Subchronic oral toxicity studies have been performed on three compounds structurally similar to 

3-decen-2-one, and reviewed by the Joint FAO/WHO Expert Committee on Food Additives

(JECFA, 2003). In these studies, the aliphatic secondary ketones, 2-octen-4-one and 1-octen-3-

one, and the secondary alcohol, 1-octen-3-ol, were used as test substances; 3-decen-2-one is an 

unsaturated, aliphatic, secondary ketone. All three 90-day studies revealed no clinical, 

hematological, or histopathological effects after 90-day exposure in rats to a single daily dose of 

6.7 mg/kg test substance (2-octen-4-one and 1-octen3-one), 12 mg/kg (1-octen3-ol, males), or 14 

mg/kg (1-octen-3-ol, females) (JECFA, 2003). These results are considered supplemental with 

regards to the 90-day oral toxicity data requirement for 3-decen-2-one because the amount of a.i. 

that can be applied to the potatoes, based on the maximum label application rates, exceed the 

single daily dose amounts given in the studies for these three structurally similar chemicals. In 

addition, the label indicates that potatoes can be shipped 24 hours post-application and there is 

no 24 hour residue data from the residue study. The earliest residue data in the residue study is 16 

days post-application.
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

7

O

3-Decen-2-one

Structures of compounds on which 90-day oral toxicity studies were performed:

1-Octen-3-ol 1-Octen-3-one 2-Octen-4-one

In summary, the information provided to fulfill the requirement for 90-day oral toxicity is 

inadequate. Residues of 3-decen-2-one and it’s degradates were detected up to 60 days post 

application on the potato internal tissue and potato peel tissue (see discussion below). 3-Decen-2-

one was observed in potato internal tissue up to 16 days post treatment (0.85 ng 3-decen-2-one 

per gram fresh weight potato) and in the peel tissue through the last testing interval of 60 days 

post treatment (403 ng/g). Residues of the primary degradates, 2-decanol and 2-decanone, were 

observed in the internal issue (2-decanol only) and the peel tissues (both chemicals) through 60 

days post treatment. According to these values, the inclusion of a source of 3-decen-2-one in the 

diet from pesticidal residues could potentially increase the current average daily intake of this 

chemical 4,000 times (see development toxicity discussion below) and exceed the daily dose 

amounts listed in the 90 day oral data cited for the three structurally similar chemicals based on 

maximum label application rates. Therefore, the Agency is requiring additional data and/or data 

waiver rationale for the 90-day oral toxicity data requirement be performed on the biochemical to 

determine if the residues observed, including residues at 24 hours post-application, are of 

toxicological concern.

No DERs were written for 90-day Oral Toxicity. Please refer to MRID 47942617 for further 

detail regarding waiver request rationale. 

CLASSIFCATION: The waiver request is UNACCEPTABLE; additional data and/or data 

waiver rationale are required. Please refer to #3 under the Recommendations and Conclusions 

section of this memorandum.

Subchronic Toxicity: 90-day Dermal Toxicity

A 90-day dermal toxicity study was not submitted and is it not required at this time for the use of 

3-decen-2-one in the proposed EPs. Prolonged exposure via the dermal route is not anticipated 

because the product is not purposefully applied to the skin or comparable prolonged human 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

8

exposure. The draft product labels for the three end-use products contain instructions for 

applicator Personal Protective Equipment (PPE) which includes: coveralls worn over long sleeve 

shirt and long pants, socks, chemical resistant footwear, chemical-resistant gloves, face shield or 

shielded safety glasses.

No DERs were written for 90-day Dermal Toxicity. Please refer to MRID 47942617 for further 

detail regarding waiver request rationale. 

CLASSIFCATION: These data are ACCEPTABLE; no additional data are required.

Subchronic Toxicity: 90-day Inhalation Toxicity

A 90-day inhalation toxicity study was not submitted. This study is not required for the use of 3-

decen-2-one in thermal fogging because label instructions limit the likelihood of significant 

levels of repeated inhalation exposure. Specifically, when 3-decen-2-one is applied to stored 

product via warehouse air circulation systems and the storage facilities remains closed for 24 

hours to personnel entry following application. A re-entry interval following thermal fogging 

should be included on the labels. However, if the end-use products containing 90% and 67.5% 3-

decen-2-one are used as spray solutions, as indicated on the product labels, the possibility of 

repeated exposure by workers exists, unless PPE or other precautions (such as the use of 

automated spray equipment only and no workers allowed in the facility during application) is 

added to these labels. Currently, there are no applicator PPE requirements for inhalation on the 

draft product labels. The applicant must clarify the discrepancies amongst the product labels. 

No DERs were written for 90-day Inhalation Toxicity. Please refer to MRID 47942617 for 

further detail regarding waiver request rationale. 

CLASSIFCATION: The waiver request is UNACCEPTABLE; additional data and/or data 

waiver rationale are required. Please refer to #4 under the Recommendations and Conclusions 

section of this memorandum.

Developmental Toxicity 

To satisfy developmental toxicity testing requirements, the applicant has submitted data waiver 

rationale stating that 3-decen-2-one: 1) occurs naturally in the diet, 2) is approved by the U.S. 

FDA as a direct food additive, 3) is metabolized into innocuous compounds, 4) has not been 

associated in the literature with developmental toxicity, and 5) is within the current intake levels, 

and well below the threshold levels, identified for flavoring agents in this compound’s structural 

class, as identified by JECFA (2003). Rationale statements 1-4, while true, are not adequate to 

completely fulfill the developmental toxicity date requirement (see discussion above). The 

Agency does not agree with statement 5, which is further discussed here. 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

9

In the JECFA (2003) report, these agents are divided into structural Classes I and II. 3-Decen-2-

one, an unsaturated, aliphatic ketone (see Figure 1, below), is placed in structural class II. The 

evaluation of these chemicals by structural class implies that they are toxicologically equivalent, 

although this is not specifically stated in the JECFA (2003) report. The threshold for human 

intake for chemicals in this class is 540 μg per person per day, as determined by JECFA (2003).

The calculated intake values, for all chemicals in this class combined, are 520 μg per person per 

day in Europe (only 20 μg less than the threshold value for this class of 540 μg) and 120 μg per 

person per day in the USA. The calculated intake of 3-decen-2-one, in particular, is 0.01 

μg/person/day in Europe and not determined for the United States (JECFA, 2003). 

O

Figure 1: Structure of 3-decen-2-one

The average daily intake (ADI) of 3-decen-2-one, resulting solely from its use in pesticide 

products as a potato sprout inhibitor, was calculated by the applicant based on the results of an 

applicant-performed residue study. In the study, potato peel and tissue for 3-decen-2-one residues 

were analyzed at 16, 23, 30, and 60 days post treatment. It is important to note that this is not 

agreement with the draft product labels for the associated end-use products containing 3-decen-2-

one as their active ingredients, which indicate that the potatoes can be treated and shipped to 

consumers 24 hours post-application (this appears under the Sprout Control Programs section on 

the label). Treatment in the residue study was at 0.75 mmol AMV-1018 (containing 98% 3-

decen-2-one as its active ingredient) per kg of fresh potato weight. This is equivalent to 0.116 

grams of product per kg of potatoes based on reviewer calculations. This is 4 times lower than 

the max label application rate (see Table 2, below) for thermal fogging. The study revealed 

residues of 3-decen-2-one in the potato internal tissue up to 16 days post application and in the 

potato peel up to 60 days post application. 

Table 2: Application Rates for End-use Products Containing 3-Decen-2-one as their Active Ingredient.

Product Percent A.I. Application 

Type Application Rate1

Conversion to 

grams product

per kg potatoes

AMV-1018 EP

(EPA File Symbol: 

5481-LTR)

98.0%

Thermal 

Fogger 

(undiluted)

No more than 17.5 fl oz 

product per ton of potatoes 

per storage season

0.482 grams product 

(98.0% pure)

per kg of potatoes

per storage season

AMV-1018 90EC

(EPA File Symbol: 

5481-LTN)

90.0% Spray

(diluted)

Mix at a rate of 1.0%-2.0% 

(v/v) in water and drench or 

spray at a rate of 0.25-0.5 

gallons per ton of potatoes2

0.0357 grams product 

(90% pure) per kg of 

potatoes3
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

10

AMV-1018 67.5EC

(EPA File Symbol: 

5481-LAO)

67.5% Spray

(diluted)

Mix at a rate of 1.3%-2.6% 

(v/v) in water and drench or 

spray at a rate of 0.25-0.5 

gallons per ton of potatoes2

0.0463 grams product 

(67.5% pure) per kg of 

potatoes4

1 per product label 2

Applicant must indicate max number of applications per storage season. 

3 Using 0.5 gallons of a 2.0% (v/v) solution per ton of potatoes.

4 Using 0.5 gallons of a 2.6% (v/v) solution per ton of potatoes.

Sixteen to 23 days post application, 3-decen-2-one made up 24-25% of measured residues. The 

other three-quarters of measured residue was almost entirely the degradate 2-decanol. The 

applicant states that, when evaluated by the criteria of the JECFA committee, this compound 

would be placed in “structural class I” of aliphatic acyclic secondary alcohol flavoring agents. 

The reviewers agree with this conclusion. The threshold for human intake for chemicals in this 

class is 1,800 μg per person per day, as determined by JECFA (2003). The actual intake values, 

for all chemicals in this class combined, was determined to be 58 μg per person per day in 

Europe and 16 μg per person per day in the USA. The actual intake of 3-decen-2-one is reported 

to be 0.01 μg/person/day in Europe and not determined for the United States (JECFA, 2003). 

A small amount of a second degradate, 2-decanone, was measured in the potato skin. The 

applicant states that this degradate is not considered to be of a concern because residues 

measured are very low (194.46 ng/g max in peel, 0 ng/g in tissue), and would be categorized by 

the JECFA method as a saturated aliphatic acyclic secondary ketone that is rapidly absorbed and 

eliminated from the blood (JECFA, 1999). 

When average potato intake for women of childbearing age is considered (U.S. EPA, 1997: age 

group 12-19 years, which is the group with highest potato intake of all adult groups and contains 

women of child-bearing age), whole potato (skin plus tissue) residue intake levels of 3-decene-2-

one, 2-decanol, and 2-decanone are applicant calculated as 41 μg/day, 221 μg/day, and 10 μg/day, 

respectively. This is well below the structural class threshold levels identified by JECFA (2003); 

3-decen-2-one represents structural class II, which has a threshold intake of 540 μg/day, and 2-

decanol represent structural class I, which has a threshold intake of 1,800 μg/day. However, a 

more appropriate comparison is to the current average daily intake of this compound. According 

to JECFA (2003), the average intake of 3-decen-2-one in Europe is 0.01 μg per person per day, 

or 0.0002 μg per kg body weight per day. These average daily intake (ADI) values indicate that 

consumption of residues of 3-decen-2-one at the levels detected in the applicant’s residue study, 

on average, are 4,000 times higher level than the amount currently consumed by the average 

European. Unfortunately, corresponding ADI values for the U.S. are not listed in the JECFA 

(2003) report, apparently due to lack of data. 

Therefore, the applicant concludes that the estimated exposure to 3-decen-2-one and it’s 

degradates from residues of proposed pesticide application are well within current acceptable 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

11

threshold intake levels (and no adverse effects have been reported in the literature) and that, 

therefore, it is not necessary to perform a guideline study for developmental toxicity. However,

based on reviewer calculations, the addition of 3-decen-2-one residues would be a very 

significant step towards the set threshold value of 540 μg/person/day for JECFA structural class

II (whose combined intake total is currently at 120 μg/person/day in the U.S.). Comparatively, a 

similar move in Europe would exceed the structural class threshold value, since the European 

daily intake value of structural class II is already 520 μg/person/day. 

Consequently, the Agency disagrees with the applicant’s conclusion that the estimated exposure 

to 3-decen-2-one and it’s degradates from residues of proposed pesticide application are well 

within current acceptable threshold intake levels. The Agency estimates that the inclusion of a 

source of 3-decen-2-one in the diet from pesticidal residues would potentially increase the 

average daily intake of this chemical 4,000 times (based on current ADIs in Europe). In addition, 

no residue data was taken at 24 hours post-application (the earliest was 16 days post-application), 

which would most-likely increase the estimated exposure amount of 3-decen-2-one. Therefore, 

the information provided to fulfill the requirement for developmental toxicity is inadequate. The 

Agency is requiring additional data and/or data waiver rationale to be submitted for this 

biochemical.

No DERs were written for Developmental Toxicity. Please refer to MRID 47942617 for 

complete waiver request rationale and MRID 47942618 for complete Residue Study. 

CLASSIFCATION: The waiver request is UNACCEPTABLE; additional data and/or data 

waiver rationale are required. Please refer to #5 under the Recommendations and Conclusions 

section of this memorandum.

Mutagenicity

To satisfy mutagenicity testing requirements, the applicant has submitted a bacterial reverse 

mutation assay (Ames Test) and a in vitro mammalian gene mutation (mouse lymphoma) assay. 

The test substance for each study was AMV-1018 Technical (EPA File Symbol No. 5481-LAI), 

containing 98.0% 3-decen-2-one. The test substance was found to be negative in the bacterial 

reverse mutation assay, and appears to be (somewhat equivocally) negative in the mouse 

lymphoma assay under 4 hour exposure test conditions, with and without metabolic activation, 

and mutagenic under 24 hour exposure conditions (without metabolic activation). There may be 

reproducible evidence of mutagenicity in this assay with and without S9 activation. In addition, 

colony sizing revealed an increase in small colonies in was observed all experiments. An increase 

in small colonies in addition to high mutant frequencies (as seen in “Experiment II” both with 

and without metabolic activation) indicates clastogenicity of the test substance. However, there is 

also evidence of marked cytotoxicity, and the osmolality for all experiments was outside the 

physiological range. The investigators conclude that, under the conditions of this assay, the test 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

12

substance is mutagenic; however, it is not clear that these results, given the increased osmotic 

pressure in particular and the marked cytotoxicity secondarily, would translate into an in vivo

system. The Agency is considering the mouse lymphoma assay to be equivocal. The applicant 

must resolve this discrepancy.

DERs were written for each submitted study and appear at the end of this memorandum.

CLASSIFCATION: The data are UNACCEPTABLE; additional data and/or data waiver 

rationale are required. Please refer to #6 under the Recommendations and Conclusions section of 

this memorandum.
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REFERENCES CITED

Thomas, A.F. (1973) An analysis of the flavor of dried mushroom, Boletus edulis. J. Agric. Food 

Chem. 21: 955-958.

Timon, M.L, Ventanas, J., Martin, L., Tejada, J.F., Garcia, C. (1998) Volatile compounds in 

supercritical carbon dioxide extracts of Iberian ham. J. Agric. Food Chem. 46: 5143-5150.

JECFA (1999) Joint FAO/WHO Expert Committee on Food Additives. WHO Food Additives 

Series 42. Saturated aliphatic acyclic secondary alcohols, ketones, and related saturated and 

unsaturated esters. International Programme on Chemical Safety.

JECFA (2003) Joint FAO/WHO Expert Committee on Food Additives. WHO Food Additives 

Series 50. Secondary alcohols, ketones, and related esters. International Programme on Chemical 

Safety. 
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DATA EVALUTATION RECORDS (DERs)

EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Acute Oral Toxicity - Rats (OCSPP 870.1100)

MRID NO: MRID 47942609

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98.57%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: Eurofins/Product Safety Laboratories

725 Cranbury Road

East Brunswick, NJ 08816

TITLE OF REPORT: Acute Oral Toxicity Up and Down Procedure in

Rats

AUTHOR: Jennifer Durando, B.S.

STUDY COMPLETED: July 24, 2009

STUDY NO: 27396
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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GOOD LABORATORY

 PRACTICE:

GLP Compliant

CONCLUSION: The oral LD50 is greater than 5,000 mg/kg body weight 

in female rats.

CLASSIFICATION: ACCEPTABLE -- TOXICITY CATEGORY IV

EXECUTIVE SUMMARY:

In an acute oral toxicity study (MRID 47942609), AMV-1018, containing 98.57% of the active 

ingredient 3-decen-2-one, was administered to four female rats according to the procedures for 

the performance of an acute oral toxicity limit test, as outlined in the U.S. EPA Health Effects 

Test Guidelines, OCSPP 870.1100. One animal died within two days of test substance 

administration. Prior to death, the animal was hypoactive and exhibited ano-genital staining, 

hunched posture, and soft feces. Gross necropsy revealed red intestines. Surviving animals 

exhibited similar symptoms, but recovered by Day 6, at which point they appeared active and 

healthy, gaining body weight over the entire 14-day observation period. No gross abnormalities 

were noted for these animals when euthanized and necropsied at the conclusion of the 14-day 

observation period. According to the results of this study, the acute oral LD50 of AMV-1018 is 

greater than 5,000 milligrams per kg body weight in female rats. This places AMV-1018 in 

TOXICITY CATEGORY IV. The study classification is ACCEPTABLE. 

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Colorless to pale yellow liquid

Lot No.: KB 147-36-1

Active Ingredient: 3-decen-2-one

Purity: 98.57% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

2. Vehicle and/or Positive Control: Test substance administered via gavage. No positive 

control (limit test). 
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3. Test Animals:

Number of Animals: 4

Species/Strain: Rat/Sprague-Dawley derived, albino.

Age/Body Weight: Young adult (9-10weeks)/170-186 grams at study initiation

Sex: Female, nulliparous and non-pregnant

Source: Received from Ace Animals, Inc., Boyertown, PA 

Housing: Singly, in suspended stainless steel cages with mesh floor

Diet: Purina Rodent Chow #5012

Water: filtered tap water supplied ad libitum by dispenser

4. Environmental Conditions:

Temperature: 19-23o

C

Humidity: 34-69%

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 7-15 days

B. STUDY DESIGN

1. In Life Dates: Start: April 22, 2009 End: May 15, 2009

2. Animal Selection: Healthy, naïve (not previously tested) female rats were selected for 

testing. 

3. Dose-selection Rationale: Dosing took place according to the limit test (OCSPP

870.1100). In total, four animals were dosed. Individual doses were calculated based on 

the initial body weight, taking into account the specific gravity of the test substance. 

4. Treatment Preparation, Administration, and Analysis: Prior to dosing, test animals 

were fasted overnight via the removal of feed from the cage. During fasting, rats were 

examined for health and weight (initial). Test substance was administered to the stomach 

using a stainless steel ball-tipped gavage needle attached to an appropriate syringe. 

Following administration, each animal was returned to its designated cage. Feed was 

replaced approximately 3-4 hours after dosing. 

C. METHODS
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1. Body Weight Measurements: Individual body weights of animals were collected three 

times: 1) initial weight before test substance administration, 2) on Day 7 following 

dosing, and 3) on Day 14 or after death. 

2. Cage-side Observations: Animals were observed for mortality, signs of gross toxicity, 

and behavioral changes. Observations took place during the first several hours post 

dosing and at least once daily thereafter for 14 days or until death occurred. Observations 

included gross evaluation of skin and fur, eyes and mucous membranes, respiratory, 

circulatory, autonomic and central nervous systems, somatomotor activity, and behavior 

pattern. 

3. Necropsy: At the end of the 14-day observation period, surviving rats were euthanized 

via CO2 inhalation. Gross necropsies were performed on these animals as well as the 

animal that did not survive the study. Examinations included tissues and organs of the 

thoracic and abdominal cavities. 

II. RESULTS

The first two animals dosed survived, however, the third animal dosed died within two days of 

test substance administration. Prior to death, the animal was hypoactive and exhibited ano-genital 

staining, hunched posture, and soft feces. Gross necropsy revealed red intestines. A fourth animal 

was dosed and survived. Therefore, the limit test outcome was O OX O (where O=survival and 

X=death). Since three or more animals survived, the LD50 for this substance is greater than the 

test dose (5,000 mg/kg). 

All surviving test animals exhibited ano-genital staining and/or were hypoactive, with hunched 

posture, piloerection, reduced fecal volume, soft feces, and facial stains following test substance 

administration. However, these animals recovered by Day 6 and appeared active and healthy, 

gaining body weight over the entire 14-day observation period. No gross abnormalities were 

noted for these animals when euthanized and necropsied at the conclusion of the 14-day 

observation period. 

III. DISCUSSION AND CONCLUSION

A. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, the acute oral LD50 of AMV1018 is greater than 5,000 milligrams per kg body weight in female rats. 
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B. REVIEWERS COMMENTS

The reviewer agrees with the investigators conclusion that, under the conditions of this study, the 

acute oral LD50 of AMV-1018 is greater than 5,000 milligrams per kg body weight in female 

rats. This places AMV-1018 in TOXICITY CATEGORY IV. There were no significant 

deviations from U.S. EPA Health Effects Test Guidelines, OCSPP 870.1100. The study is 

classified as acceptable/guideline (OCSPP 870.1100) and satisfies the requirements for an acute 

oral toxicity study in the rat. Signed and dated Data Confidentiality, Good Laboratory Practice 

(GLP) Compliance, and Quality Assurance statements were provided. 

C. STUDY DEFICIENCIES

No significant study deficiencies that would invalidate the study outcome were found. 
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EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Acute Dermal Toxicity - Rats (OCSPP 870.1200)

MRID NO: MRID 47942610

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98.57%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: Eurofins/Product Safety Laboratories

2394 U.S. Highway 130

Dayton, NJ 08810

TITLE OF REPORT: Acute Dermal Toxicity Study in Rats – Limit Test

AUTHOR: Jennifer Durando, B.S.

STUDY COMPLETED: July 24, 2009

STUDY NO: 27397

GOOD LABORATORY

 PRACTICE:

GLP Compliant
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CONCLUSION: The dermal LD50 for male and female rats was greater 

than 5,000 mg/kg body weight.

CLASSIFICATION: ACCEPTABLE -- TOXICITY CATEGORY IV

EXECUTIVE SUMMARY:

In an acute oral toxicity study (MRID 47942610), AMV-1018, containing 98.57% of the active 

ingredient 3-decen-2-one, was administered to ten rats (5 male and 5 female) according to the 

procedures for the performance of a limit test for acute dermal toxicity test, as outlined in the 

U.S. EPA Health Effects Test Guidelines, OCSPP 870.1200. One animal died within two days of 

test substance administration. Prior to death, the animal was hypoactive and exhibited prone 

posture. Gross necropsy revealed extremely red intestines. All surviving test animals exhibited 

dermal irritation at the dose site between Days 1 and Day 14. Seven of these animals were 

hypoactive, but all recovered by Day 2. All surviving animals gained body weight over the entire 

14-day observation period. No gross abnormalities were noted for the surviving animals when 

euthanized and necropsied at the conclusion of the 14-day observation period. According to the 

results of this study, the acute dermal LD50 of AMV-1018 is greater than 5,000 milligrams per kg 

body weight in male and female rats. This places AMV-1018 in TOXICITY CATEGORY IV. 

The study classification is ACCEPTABLE.

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Colorless to pale yellow liquid

Lot No.: KB 147-36-1

Active Ingredient: 3-decen-2-one

Purity: 98.57% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

2. Test Animals:

Number of Animals: 10 

Species/Strain: Rat/Sprague-Dawley derived, albino.

Age/Body Weight: Young adult (8-9 weeks)/ males 217-238 grams and 

females 182-208 at study initiation
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Sex: 5 Male and 5 Female - nulliparous and non-pregnant

Source: Received from Ace Animals, Inc., Boyertown, PA 

Housing: Singly, in suspended stainless steel cages with mesh floor

Diet: Purina Rodent Chow #5012

Water: filtered tap water supplied ad libitum by dispenser

3. Environmental Conditions:

Temperature: 20-23o

C

Humidity: 51-66%

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 7 days

B. STUDY DESIGN

1. In Life Dates: Start: April 28, 2009 End: May 12, 2009

2. Treatment Preparation and Animal Selection: On the day prior to test substance 

application, a group of animals was prepared by clipping hair of the dorsal area and the 

trunk. After clipping and prior to application, the animals were examined for health 

including any skin abnormalities. Initial body weights were recorded. From this group ten 

healthy, naïve (not previously tested) animals were selected for the test – five males and 

five females.

3. Dose-selection Rationale: Individual doses were calculated based on the initial body 

weight, taking into account the specific gravity of the test substance. 

4. Test Substance Administration and Analysis: The test substance was administered at 

5000 mg/kg of body weight, applied evenly over an area of approximately 2 inches x 3 

inches on the dorsal side of the animal where hair had been clipped. Then, the area was 

covered with a 2 in x 3 in gauze pad. The pad and entire trunk of the animal were 

wrapped with tape. After 24 hours of exposure to the test substance, the gauze pads were 

removed and the test sites were gently cleansed of any residual test substance. 

C. METHODS

1. Body Weight Measurements: Individual body weights of animals were collected three 

times: 1) initial weight before test substance administration, 2) on Day 7 following 

dosing, and 3) on Day 14 or after death. 
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2. Cage-side Observations: Animals were observed for mortality, signs of gross toxicity, 

and behavioral changes. Observations took place during the first several hours post 

application and at least once daily thereafter for 14 days or until death occurred. 

Observations included gross evaluation of skin and fur, eyes and mucous membranes, 

respiratory, circulatory, autonomic and central nervous systems, somatomotor activity, 

and behavior pattern. 

3. Necropsy: At the end of the 14-day observation period, surviving rats were euthanized 

via CO2 inhalation. Gross necropsies were performed on these animals as well as the one 

animal (female) that did not survive the study. Examinations included tissues and organs

of the thoracic and abdominal cavities. 

II. RESULTS

One female died within two days of test substance application. Prior to death this animal was 

hypoactive and exhibited prone posture. Gross necropsy of the deceased animal revealed 

extremely red intestines. 

All surviving test animals exhibited dermal irritation at the dose site between Days 1 and Day 14.

All surviving animals gained body weight over the entire 14-day observation period. Seven of 

these animals were hypoactive, but all recovered by Day 2. No gross abnormalities were noted 

for the surviving animals when euthanized and necropsied at the conclusion of the 14-day 

observation period.

III. DISCUSSION AND CONCLUSION

A. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, the acute dermal LD50 of 

AMV-1018 is greater than 5,000 milligrams per kg body weight in male and female rats.

B. REVIEWERS COMMENTS

The reviewer agrees with the investigators conclusion that, under the conditions of this study, the 

acute dermal LD50 of AMV-1018 is greater than 5,000 milligrams per kg body weight in male 

and female rats. This places AMV-1018 in TOXICITY CATEGORY IV. There were no 

significant deviations from U.S. EPA Health Effects Test Guidelines, OCSPP 870.1200. The 

study is classified as acceptable/guideline (OCSPP 870.1200) and satisfies the requirements for 

an acute dermal toxicity study in the rat. Signed and dated Data Confidentiality, Good Laboratory 

Practice (GLP) Compliance, and Quality Assurance statements were provided.
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C. STUDY DEFICIENCIES

No significant study deficiencies that would invalidate the study outcome were found. 
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EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Acute Inhalation Toxicity - Rats (OCSPP 870.1300)

MRID NO: MRID 47942611

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98.57%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: Eurofins/Product Safety Laboratories

2394 U.S. Highway 130

Dayton, NJ 08810

TITLE OF REPORT: Acute Inhalation Toxicity Study in Rats

AUTHOR: Jennifer Durando, B.S.

STUDY COMPLETED: July 24, 2009

STUDY NO: 27398
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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GOOD LABORATORY

 PRACTICE:

GLP Compliant

CONCLUSION: The acute inhalation LC50 is between 0.52 - 2.04 mg/L in 

male rats and is greater than 2.04 mg/L in female rats. 

CLASSIFICATION: ACCEPTABLE -- TOXICITY CATEGORY III

EXECUTIVE SUMMARY:

In an acute inhalation toxicity study (MRID 47942611), five male and five female rats were 

exposed to 2.04mg/L AMV-1018, containing 98.57% of the active ingredient 3-decen-2-one. 

Testing was in accordance with the procedures for the performance of an acute inhalation toxicity 

Limit test, as outlined in the U.S. EPA Health Effects Test Guidelines, OCSPP 870.1300. One

male died immediately following exposure to the test atmosphere, with no clinical signs 

observed. Two males died within two days of exposure to the test atmosphere. Surviving animals 

exhibited clinical signs, however, these animals recovered by Day 10 and appeared active and 

healthy for the remainder of the study. Due to the treatment related mortality observed, a second 

test was run with five naïve male rats and an exposure scenario of 0.52mg/L. One animal died 

within one day of exposure to the test atmosphere. According to these results, the acute 

inhalation LC50 of AMV-1018 is between 0.52 mg/L and 2.04 mg/L in male rats and is greater 

than 2.04 mg/L in female rats. This places AMV-1018 in TOXICITY CATEGORY III. The study 

classification is ACCEPTABLE.

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Colorless to pale yellow liquid

Lot No.: KB 147-36-1

Active Ingredient: 3-decen-2-one

Purity: 98.57% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

2. Delivery: Test substance was aerosolized as received from the Sponsor. 

3. Test Animals:
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Number of Animals: 15

Species/Strain: Rat/Sprague-Dawley derived, albino.

Age/Body Weight: Young adult (9-11 weeks)/males 267-346 grams and 

females 210-248 grams at study initiation

Sex: 10 males and 5 females (nulliparous and non-pregnant)

Source: Received from Ace Animals, Inc., Boyertown, PA 

Housing: Singly, in suspended stainless steel cages with mesh floor

Diet: Purina Rodent Chow #5012

Water: filtered tap water supplied ad libitum by dispenser

4. Apparatus and Equipment:

Exposure Apparatus: Nose-only inhalation chamber, internal volume 6.7 liters

Air Supply: Filtered air supplied by compressor and air tank (mixing)

Ambient Conditions: Humidity-Temperature Indicator/Monitor

Particle Size: Eight-stage Anderson Cascade Impactor

Chamber Concentration: Gravimetric samples collected using glass fiber filters 

attached to vacuum pump via filter holder and tubing

Treatment of Exhaust Air: Air is discharged from the chamber through a 0.5-inch tube 

attached to the base unit of the chamber. Treatment of 

exhausted air was not reported. 

5. Environmental Conditions:

Temperature: 19-23o

C

Humidity: 50-67%

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 13-20 days

B. STUDY DESIGN

1. In Life Dates: Start: April 20 and May 11, 2009 End: May 25, 2009

2. Pre-test Trials: Prior to study initiation, pre-test trials were conducted to establish 

procedures to achieve target chamber concentration (2.0 mg/L and 0.5 mg/L) and desired 

particle size distribution (less than or equal to 4 μm). Results revealed conditions 

favorable to achieve 2.04 mg/L chamber concentration at particle size distribution of 

2.8 μm, and 0.52 mg/L chamber concentration at particle size distribution 2.5 μm. 
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3. Test Conditions: Temperature and humidity within the chamber as well as the room 

were recorded every 15 minutes during the first hour of exposure and every 30 minutes 

thereafter. Chamber concentration measurements were taken via gravimetric 

measurements from the breathing zone of the animals, six times during each exposure. 

For each exposure level, the animals were exposed to the targeted chamber concentration 

for at least 4 hours. Nominal concentration of test substance was 13.72 mg/L for the 

2.0 mg/L study, and 6.73 mg/L for the 0.5 mg/L study. Actual (gravimetric) 

concentrations of test substance in the breathing zone were 2.04 mg/L and 0.52 mg/L, 

respectively.

4. Animal Selection, Assignment, and Dose-selection Rationale: On the day of exposure 

and the day prior to exposure, the rats were examined for health and were weighed. Five 

male and five female rats were selected for the 2.04 mg/L exposure. Based on the results 

of this exposure, five male rats only were selected for the 0.52 mg/L exposure. 

5. Treatment Preparation, Administration, and Analysis: For each exposure level, 

animals were exposed to the target chamber concentration for at least 4 hours (extended 

beyond 4 hours to allow the chamber to reach equilibrium). Chamber was then operated 

for an additional 15 minutes with clean air. Animals were removed, fur cleaned of excess 

test material, and returned to their cages. 

C. METHODS

1. Body Weight Measurements: Individual body weights of animals were collected three 

times: 1) initial weight before test substance administration, 2) on Day 7 following 

dosing, and 3) on Day 14 or after death. 

2. Cage-side Observations: Animals were observed for mortality, signs of gross toxicity, 

and behavioral changes. Observations took place during the first several hours post 

dosing and at least once daily thereafter for 14 days or until death occurred. Observations 

included gross evaluation of skin and fur, eyes and mucous membranes, respiratory, 

circulatory, autonomic and central nervous systems, somatomotor activity, and behavior 

pattern. 

3. Necropsy: At the end of the 14-day observation period, surviving rats were euthanized 

via CO2 inhalation. Gross necropsies were performed on these animals as well as the 

animal that did not survive the study. Examinations included tissues and organs of the 

thoracic and abdominal cavities. 

II. RESULTS
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At the 2.04 mg/L exposure level, one male died immediately following exposure to the test 

atmosphere, with no clinical signs observed prior to death. Two males died within two days of 

exposure to the test atmosphere, displaying signs of hypoactivity, abnormal respiration, nasal 

discharge, facial staining, hunched posture, and reduced fecal volume, and were cold to the 

touch. The surviving animals exhibited hypoactivity, hunched posture, abnormal respiration, 

reduced fecal volume, nasal and oral discharge and/or facial staining. However, these animals 

recovered by Day 10 and appeared active and healthy for the remainder of the study. Gross 

necropsy of the dead animals revealed edema, discoloration of the lungs and liver, gas distended 

yellow intestines, and/or rigor mortis. No gross abnormalities were noted for the surviving 

animals when euthanized and necropsied at the conclusion of the 14-day observation period.

At the 0.52 mg/L exposure level, one animal died within one day exposure to the test 

atmosphere. Prior to death, this animal was hypoactive and exhibited irregular respiration and 

moist rales. The surviving animals also exhibited hypoactivity, irregular respiration, and rales, as 

well as hunched posture, reduced fecal volume, and facial and/or ano-genital staining. However, 

these animals recovered by Day 7 and appeared active and healthy, gaining body weight over the 

entire 14-day observation period. Gross necropsy of the dead animal revealed extremely red 

lungs. No gross abnormalities were noted for the surviving animals when euthanized and 

necropsied at the conclusion of the 14-day observation period.

III. DISCUSSION AND CONCLUSION

A. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, the acute inhalation LC50 of 

AMV-1018 is between 0.52 mg/L and 2.04 mg/L in male rats and is greater than 2.04 mg/L in 

female rats. 

B. REVIEWERS COMMENTS

This study first used the Limit test approach for acute inhalation (five males and five females 

dosed at 2 mg/L). Due to the test substance related mortality seen as a result of this test (3 male 

deaths), further testing was required. The investigators’ then dosed five male rats only at 

0.52mg/L. One animal death was observed. Due to the results of these tests, the reviewer agrees 

with the investigators conclusion that, under the conditions of this study, the acute inhalation 

LC50 of AMV-1018 is between 0.52 mg/L and 2.04 mg/L in male rats and is greater than 2.04 

mg/L in female rats. This places AMV-1018 in TOXICITY CATEGORY III. There were no 

significant deviations from U.S. EPA Health Effects Test Guidelines, OCSPP 870.1300. The 

study is classified as acceptable/guideline (OCSPP 870.1300) and satisfies the requirements for 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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an acute inhalation toxicity study in the rat. Signed and dated Data Confidentiality, Good 

Laboratory Practice (GLP) Compliance, and Quality Assurance statements were provided. 

C. STUDY DEFICIENCIES

No significant study deficiencies that would invalidate the study outcome were found. 
3-decen-2-one DP Number: 373085

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EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Acute Eye Irritation (OCSPP 870.2400)

MRID NO: MRID 47942612

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98.57%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: Eurofins/Product Safety Laboratories

725 Cranbury Road

East Brunswick, NJ 08816

TITLE OF REPORT: Primary Eye Irritation Study in Rabbits

AUTHOR: Jennifer Durando, B.S.

STUDY COMPLETED: July 24, 2009

STUDY NO: 27399
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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GOOD LABORATORY

 PRACTICE:

GLP Compliant

CONCLUSION: Test substance is moderately irritating to the eye.

CLASSIFICATION: ACCEPTABLE -- TOXICITY CATEGORY II

EXECUTIVE SUMMARY:

In an acute eye irritation study (MRID 47942612), three female rabbits were exposed to 0.1 mL 

AMV-1018, which contains the active ingredient 3-decen-2-one at 98.57%. Testing was 

performed according to the procedures for the performance of a step-wise acute eye irritation test, 

as outlined in the U.S. EPA Health Effects Test Guidelines, OCSPP 870.2400. One hour after 

test substance instillation, all three treated eyes exhibited corneal opacity and conjunctivitis, and 

two treated eyes exhibited iritis. Severity decreased over time, with all animals free of ocular 

irritation by Day 10. All animals appeared active and healthy during the study, with no signs of 

toxicity aside from the eye irritation. The Maximum Mean Total Score (MMTS) for ocular 

irritation in this study is 24.3. According to these results, AMV-1018 is classified as moderately 

irritating to the eye. This places AMV-1018 in TOXICITY CATEGORY II. The study 

classification is ACCEPTABLE.

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Colorless to pale yellow liquid

Lot No.: KB 147-36-1

Active Ingredient: 3-decen-2-one

Purity: 98.57% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

2. Delivery and Control: The test substance was instilled into the right eye of the test 

animal. Substance was instilled as received by the Sponsor. The left eye remained 

untreated and served as a control. 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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32

3. Test Animals:

Number of Animals: 3

Species/Strain: Rabbit/New Zealand albino

Age/Body Weight: Young adult 

Sex: Female, nulliparous and non-pregnant

Source: Received from Robinson Services, Inc. Clemmons, NC 

Housing: Singly, in suspended stainless steel cages with mesh floor

Diet: Pelleted Purina Rabbit Chow #5326

Water: filtered tap water supplied ad libitum by dispenser

4. Environmental Conditions:

Temperature: 20-22o

C

Humidity: 36-70%

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 12 or 19 days

B. STUDY DESIGN

1. In Life Dates: Start: May 18, 2009 End: June 8, 2009

2. Animal Selection: Prior to test irritation, a group of female rabbits under went ocular 

examination involving a white light source and fluorescein dye. Three healthy, naïve 

animals (not previously tested) without pre-existing ocular irritation were selected for the 

test. 

3. Treatment Preparation, Administration, and Analysis: Study was conducted 

according to a step-wise procedure, observing the effects of installation on one animal 

before enrolling two more. Prior to all instillations, 2-3 drops of ocular anesthetic were 

placed into both eyes. Then, one-tenth of a millimeter of the test substance was instilled 

into the conjunctival sac of the right eye. The left eye remained untreated and served as a 

control. Ocular irritation was evaluated using a high-intensity white light and scored 

according to Draize, et al. (1944). 

C. METHODS

1. Body Weight Measurements: Not taken. 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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2. Cage-side Observations: Animals were observed for signs of gross toxicity, and 

behavioral changes at least once daily during the test period. Observations included gross 

evaluation of skin and fur, eyes and mucous membranes, respiratory, circulatory, 

autonomic and central nervous systems, somatomotor activity, and behavior pattern.

3. Necropsy: Not performed. 

II. RESULTS

One hour after test substance instillation, all three treated eyes exhibited corneal opacity and 

conjunctivitis, and two treated eyes exhibited iritis. Severity decreased over time, with all 

animals free of ocular irritation by Day 10. All animals appeared active and healthy during the 

study, with no signs of toxicity aside from the eye irritation. The Maximum Mean Total Score 

(MMTS) for ocular irritation in this study is 24.3.

III. DISCUSSION AND CONCLUSION

D. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, AMV-1018 is moderately 

irritating to the eye.

E. REVIEWERS COMMENTS

This study followed the step-wise procedure for an acute eye irritation study, as set forth in the 

U.S. EPA Health Effects Test Guidelines, OCSPP 870.2400. Three animals were dosed, with all 

three exhibiting eye irritation and no other signs of gross toxicity or abnormal behavior. Irritation 

cleared by Day 10. Therefore, the reviewer recommends that AMV-1018 be placed in 

TOXICITY CATEGORY II. There were no significant deviations from U.S. EPA Health Effects 

Test Guidelines, OCSPP 870.2400. The study is classified as acceptable/guideline (OCSPP

870.2400) and satisfies the requirements for acute eye irritation in the rabbit. Signed and dated 

Data Confidentiality, Good Laboratory Practice (GLP) Compliance, and Quality Assurance 

statements were provided. 

F. STUDY DEFICIENCIES

No significant study deficiencies that would invalidate the study outcome were found. 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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IV. REFERENCES

Draize, J.H., Woodward, G. and Calvery, H.O. (1944) Methods for the study of irritation and 

toxicity of substances applied topically to the skin and mucous membranes. J. Pharmacol. Exp. 

Ther. 82:377-390.
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

35

EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Acute Dermal Irritation (OCSPP 870.2500)

MRID NO: MRID 47942613

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98.57%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: Eurofins/Product Safety Laboratories

2725 Cranbury Road

East Brunswick, NJ 08816

TITLE OF REPORT: Primary Skin Irritation Study in Rabbits

AUTHOR: Jennifer Durando, B.S.

STUDY COMPLETED: July 24, 2009

STUDY NO: 27400
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

36

GOOD LABORATORY

 PRACTICE:

GLP Compliant

CONCLUSION: Test substance is severely irritating to the skin. 

CLASSIFICATION: ACCEPTABLE -- TOXICITY CATEGORY II

EXECUTIVE SUMMARY:

In an acute dermal irritation study (MRID 47942613), three male rabbits were exposed for four 

hours to 0.5 mL AMV-1018, which contains the active ingredient 3-decen-2-one at 98.57%. 

Testing was performed according to the procedures outlined in the U.S. EPA Health Effects Test 

Guidelines, OCSPP 870.2500. Animals exhibited severe irritation (severe erythema and 

moderate edema) through 72 hours post patch removal. Irritation was no longer observed by Day 

14. The Primary Dermal Irritation Index (PDII) calculated for this test substance was 5.5. 

According to the results of this study, AMV-1018 is classified as severely irritating to the skin 

and is placed in TOXICITY CATEGORY II. The study classification is ACCEPTABLE.

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Colorless to pale yellow liquid

Lot No.: KB 147-36-1

Active Ingredient: 3-decen-2-one

Purity: 98.57% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

2. Test Animals:

Number of Animals: 3

Species/Strain: Rabbit/New Zealand albino

Age/Body Weight: Young adult 

Sex: Male

Source: Received from Robinson Services, Inc. Clemmon, NC 

Housing: Singly, in suspended stainless steel cages with mesh floor

Diet: Pelleted Purina Rabbit Chow #5326
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

37

Water: filtered tap water supplied ad libitum by dispenser

3. Environmental Conditions:

Temperature: 19-22oC

Humidity: 40-66%

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 6 days

B. STUDY DESIGN

1. In Life Dates: Start: April 28, 2009 End: May 12, 2009

2. Treatment Preparation and Animal Selection: On the day prior to test substance 

application, a group of animals was prepared by clipping hair of the dorsal area and the 

trunk. After clipping and prior to application, the animals were examined for health 

including any skin abnormalities. From this group three healthy male animals, with no 

pre-existing skin irritation, were selected for the test.

3. Test Substance Administration and Analysis: Five-tenths of a milliliter of test

substance was administered to one 6 cm2 dose site (hair previously clipped). Then, the 

area was covered with a 1 inch x 1 inch gauze pad. The pad and entire trunk of the animal 

were wrapped with tape. Elizabethan collars were placed on animals to avoid 

ingestion/inhalation of the test substance. After 4 hours of exposure to the test substance, 

the gauze pads and collars were removed and the test sites were gently cleansed of any 

residual test substance. Skin irritation was scored according to Draize, et al. (1944). 

C. METHODS

1. Body Weight Measurements: Not taken. 

2. Cage-side Observations: Animals were observed for signs of gross toxicity, and 

behavioral changes at least once daily during the test period. Observations included gross 

evaluation of skin and fur, eyes and mucous membranes, respiratory, circulatory, 

autonomic and central nervous systems, somatomotor activity, and behavior pattern.

3. Necropsy: Not performed. 

II. RESULTS

One hour after patch removal, all three treated sites exhibited well-defined to severe erythema 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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and moderate edema. Severe irritation was observed through 72 hours post patch removal. 

Irritation (erythema and edema) was gone by Day 14. The Primary Dermal Irritation Index 

(PDII) calculated for this test substance was 5.5.

III. DISCUSSION AND CONCLUSION

A. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, AMV-1018 is classified as 

severely irritating to the skin.

B. REVIEWERS COMMENTS

This study followed the step-wise procedure for an acute dermal irritation study, as set forth in 

the U.S. EPA Health Effects Test Guidelines, OCSPP 870.2500. Animals exhibited severe 

irritation through 72 hours post patch removal. Therefore, the reviewer agrees with the 

investigators conclusion that, under the conditions of this study, AMV-1018 is classified as 

severely irritating to the skin. This places AMV-1018 in TOXICITY CATEGORY II. There were 

no significant deviations from U.S. EPA Health Effects Test Guidelines, OCSPP 870.2500. The 

study is classified as acceptable/guideline (OCSPP 870.2500) and satisfies the requirements for 

acute dermal irritation in the rabbit. Signed and dated Data Confidentiality, Good Laboratory 

Practice (GLP) Compliance, and Quality Assurance statements were provided. 

C. STUDY DEFICIENCIES

No significant study deficiencies that would invalidate the study outcome were found. 

IV. REFERENCES

Draize, J.H., Woodward, G. and Calvery, H.O. (1944) Methods for the study of irritation and 

toxicity of substances applied topically to the skin and mucous membranes. J. Pharmacol. Exp. 

Ther. 82:377-390.
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

39

EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Skin Sensitization (OCSPP 870.2600)

MRID NO: MRID 47942614

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98.57%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: Eurofins/Product Safety Laboratories

2394 U.S. Highway 130

Dayton, NJ 08810

TITLE OF REPORT: Dermal Sensitization in Guinea Pigs (Buehler Method)

AUTHOR: Jennifer Durando, B.S.

STUDY COMPLETED: July 24, 2009

STUDY NO: 27401
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

40

GOOD LABORATORY

 PRACTICE:

GLP Compliant

CONCLUSION: Test substance is not a dermal sensitizer.

CLASSIFICATION: ACCEPTABLE

EXECUTIVE SUMMARY:

In a dermal sensitization study (MRID 47942614), twenty healthy guinea pigs were exposed to 

topical applications of AMV-1018, which contains the active ingredient 3-decen-2-one at 

98.57%. Testing was performed according to the Buehler Method, as outlined in the U.S. EPA 

Health Effects Test Guidelines, OCSPP 870.2600. Twenty test animals were dosed three times 

each week for three weeks with either undiluted test substance or 75% w/w test substance in 

mineral oil. Twenty-seven days after the first dose was given, test animals and ten additional 

control animals were given a challenge dose of 1% w/w test substance in mineral oil to determine 

sensitization to the test material. No positive reactions were observed following challenge in 

either the test or the control animals. According to these results, AMV-1018 is classified as not a 

dermal sensitizer. The study classification is ACCEPTABLE.

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Colorless to pale yellow liquid

Lot No.: KB 147-36-1

Active Ingredient: 3-decen-2-one

Purity: 98.57% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

2. Delivery and Control: The test substance was topically applied (0.4 mL) either undiluted 

or at 75% w/w in mineral oil for an initial induction period. Test substance at 1% w/w in 

mineral oil was applied for the challenge application. A naïve control group of ten 

animals was treated at challenge only. 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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41

3. Test Animals:

Number of Animals: 38

Species/Strain: Guinea pigs/Hartley albino

Age/Body Weight: Young adult, 318-412 grams at experimental start

Sex: Female, nulliparous and non-pregnant 

Source: Received from Elm Hill Breeding Labs, Chelmsford, MA

Housing: Singly, in suspended stainless steel cages with mesh floor

Diet: Pelleted Purina Guinea Pig Chow #5025

Water: filtered tap water supplied ad libitum by dispenser

4. Environmental Conditions:

Temperature: 19-22o

C

Humidity: 42-74%

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 5-7 days

B. STUDY DESIGN

1. In Life Dates: Start: April 17, 2009 End: May 28, 2009

2. Animal Selection: On the day prior to test substance application, a group of animals was 

prepared by clipping hair of the dorsal area and the trunk. After clipping and prior to 

application, the animals were examined for health including any skin abnormalities. 

Initial body weights were recorded. From this group, eight healthy animals were selected 

for the preliminary irritation group, twenty for the test group, and ten for the naïve control 

group. Females only were chosen for the test and naïve control groups. 

3. Dose Selection: Dose was based on the results of a preliminary irritation test. Dose 

concentrations of 100%, 75%, 50%, 25%, 12%, 6%, 3%, and 1% (dilutions in mineral 

oil) were applied at 0.4 mL to an area of clipped fur using an occlusive chamber. The test 

area was then wrapped for 6 hours. The occlusive chamber was removed after 6 hours and

the test area was cleaned to remove any residual test substance. Test sites were examined 

and scored approximately 24 hours after application. The highest non-irritating 

concentration that produced a response in 4 guinea pigs was the 1% dilution, therefore, 

this concentration was chosen for the challenge phase. 

4. Treatment Preparation, Administration, and Analysis: Twenty test animals were 

dosed three times each week for three weeks with 0.4mL of either undiluted test 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

42

substance or 75% w/w test substance in mineral oil. Twenty-seven days after the first 

dose was given, test animals and ten additional control animals were given a challenge 

dose (1% w/w) to determine sensitization to the test material. Scores were given for 

incidence and for severity, based on the methods of Ritz and Buehler (1980). Sensitizer 

classification was based on the criterion of Robinson, et al. (1990). 

C. METHODS

1. Body Weight Measurements: Individual body weights (test group and control group) 

were recorded prior to experimental start and again on the day after challenge. 

2. Cage-side Observations: Not performed. 

3. Necropsy: Not performed. 

II. RESULTS

No positive reactions were observed following challenge in either the test or the control animals. 

(Very faint erythema was seen in some animals, but this is not considered a positive reaction). 

III. DISCUSSION AND CONCLUSION

A. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, AMV-1018 is not a dermal 

sensitizer. 

B. REVIEWERS COMMENTS

This study followed the Buehler method for skin sensitization determination, as set forth in the 

U.S. EPA Health Effects Test Guidelines, OCSPP 870.2600. No positive reactions were 

observed following challenge in either the test or the control animals. Therefore, the reviewer 

agrees with the investigators conclusion that AMV-1018 is not a contact sensitizer. There were 

no significant deviations from U.S. EPA Health Effects Test Guidelines, OCSPP 870.2600. The 

study is classified as acceptable/guideline (OCSPP 870.2600) and satisfies the requirements for 

skin sensitization in the guinea pig. Signed and dated Data Confidentiality, Good Laboratory 

Practice (GLP) Compliance, and Quality Assurance statements were provided. 

C. STUDY DEFICIENCIES
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

43

No significant study deficiencies that would invalidate the study outcome were found. 

IV. REFERENCES

Ritz, H. and Buehler, E. (1980) Planning, conduct, and interpretation of guinea pig sensitization 

patch tests. Current Concepts in Cutaneous Toxicity, V.A. Drill and P. Lazar (Eds.), Academic 

Press, New York, 1980, pg 25.

Robinson, M., Nusair, T., Fletcher, E., and Ritz, H. (1990) A review of the Buehler Guinea Pig 

Sensitization Test and its use in risk assessment process for human skin sensitization. 

Toxicology, 61: 91-107. 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

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EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: Bacterial Reverse Mutation Test (Salmonella 

typhimurium) OCSPP 870.5100

MRID NO: MRID 47942616

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: BSL BIOSERVICE Scientific Laboratories GmbH 

Behringstrabe 6/8

82152 Planegg, Germany

TITLE OF REPORT: Reverse Mutation Assay Using Bacteria

AUTHOR: Dr. Barbara Wallner

STUDY COMPLETED: July 7, 2009

STUDY NO: 091460
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

Human Health Toxicity

45

GOOD LABORATORY

 PRACTICE:

GLP Compliant

CONCLUSION: Test substance is non-mutagenic (negative) in this assay.

CLASSIFICATION: ACCEPTABLE 

EXECUTIVE SUMMARY:

In a reverse mutation assay (MRID 47942616), AMV-1018, containing 98% of the active 

ingredient 3-decen-2-one, was investigated for its potential to induce gene mutations via a plate 

incorporation test (Experiment I), and a pre-incubation test (Experiment II). Each experiment was 

conducted with five tester strains of Salmonella typhimurium (TA 98, TA 100, TA 1535, TA 

1537, and TA 102), six different test substance concentrations (0.0316, 0.100, 0.316, 1.0, 2.5 and 

5.0 μL/plate, and control scenario), and with and without metabolic activation. According to the 

results of this study, no biologically relevant increases in revertant colony numbers of any of the 

five tester strains were observed following treatment with AMV-1018 at any concentration level, 

neither in the presence or absence of metabolic activation, in either experiment. In the preincubation experiments, cytotoxicity was noted in all five tester strains at a dose concentration of 

5.0 μL/plate without metabolic activation and in tester strain TA 102 at a dose concentration of 

5.0 μL/plate with metabolic activation. The reference mutagens employed in the control 

scenarios induced a distinct increase in revertant colonies, indicating the validity of the 

experiments. Therefore, the test substance is considered to be non-mutagenic in this bacterial 

reverse mutation assay. The study classification is ACCEPTABLE. 

I. MATERIALS AND METHODS

A. MATERIALS

1. Test Substance: AMV-1018

Description: Pale yellow liquid

Lot No.: 3D2-2009/01

Active Ingredient: 3-decen-2-one

Purity: 98% w/w

Compound Stability: Expected to be stable for the duration of testing, outcome of 

stability test not available at time of study. 

CAS No of TGAI: 10519-33-2
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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46

2. Positive Controls: The reference mutagens sodium azide, 4-nitro-o-phenylene-diamine, 

and methyl methane sulfonate were tested in one or two of the test strains without 

metabolic activation. The reference mutagen 2-aminoanthracene was tested in all five test 

strains with metabolic activation. The OCSPP Test Guideline for this assay (OCSPP

870.5100) recommends that 2-Aminoanthracene not be used as the sole indicator of the 

efficacy of the S9 mix, and that a pro-mutagen requiring metabolic activation by 

microsomal enzymes also be used. The study authors did not follow this 

recommendation, however, the study is still considered to be acceptable. 

3. Test System: Five different strains of the bacterium Salmonella typhimurium. 

Strain: Characteristic:

TA 98 frame shift mutations

TA 100 base-pair substitutions

TA 1535 base-pair substitutions

TA 1537 frame shift mutations

TA 102 base-pair substitutions

All strains contain mutations in the histidine operon, thereby imposing a requirement for 

histidine in the growth medium. They also all contain the deep rough (rfa) mutation, 

which deletes the polysaccharide side chain of the lipopolysaccharides of the bacterial 

cell surface, thus increases the permeability of larger substances, and the uvrB mutation, 

which is a deletion of the uvrB gene coding for a protein of the DNA nucleotide excision 

repair system resulting in an increased sensitivity in detecting many mutagens. Tester 

strains TA 98, TA 100 and TA 102 contain the R-factor plasmid pkM101. These strains 

are reverted by a number of mutagens that are detected weakly or not at all in the parent 

strains. 

B. METHODS

1. Preparation of Bacteria: Samples of each tester strain were grown by culturing for 12h 

at 38.5 o

C in nutrient broth (8 g nutrient broth + 5 g NaCl) to the late exponential or early 

stationary phase of growth (appox. 109 cells/mL). For strains TA 98, TA 100, and TA 

102, a solution of 125 μL ampicillin (10 mg/mL) was added in order to retain the 

phenotypic characteristics of the strains. 

2. Preparation of Medium: Vogel-Donner Medium E agar plates with 2% glucose were 

either prepared by BSL BIOSERVICE GmbH or provided by a supplier. The investigators

indicate that quality controls were performed and sterilization was at 121o

C in an 

autoclave. Overlay agar containing L-histidine and biotin was prepared and sterilized at 

121oC in an autoclave.
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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3. Preparation of Metabolic Activation (S9): The S9 homogenate was prepared at the 

facility by inducing male Wistar rats with Phenobarbital (80 mg/kg bw) and βNaphthoflavone (100 mg/kg bw) for three consecutive days by oral route. The protein 

concentration in the S9 preparation was 33 mg/mL. The S9 mix was prepared according 

to Ames et al. (1973).

C. STUDY DESIGN

1. Exposure Concentrations: A pre-experiment was conducted for toxicity of the test 

substance at eight concentration levels. The following six were chosen for the main 

experiments: 0.0316, 0.100, 0.316, 1.0, 2.5 and 5.0 μL/plate.

2. Controls: Three plates per experiment were prepared for each of the following scenarios: 

solvent control, negative control, and reference mutagen solution (positive control). 

3. Procedures: For each strain and dose level, three plates were used. For each experiment, 

after solidification, the plates were inverted and incubated in the dark at 37o

C for at least 

24 hours. 

a. Plate Incorporation Experiment: The following materials were mixed in a test 

tube and poured over the surface of a minimal agar plate: 100μL test solution at 

one of six concentration levels for each of three controls, 500 μL of either S9 mix 

or S9 substitution buffer (for testing without metabolic activation), 100 μL of 

bacteria suspension, and 2000 μL of overlay agar. 

b. Pre-Incubation Experiment: 100 μL of the test substance preparation was 

preincubated with the tester strains (100 μL) and sterile buffer or the S9 (500 μL) 

for 60 minutes at 37o

C prior to adding the overlay agar (2000 μL) and pouring 

onto the surface of a minimal agar plate.

4. Data and Reporting: Data was collected using a ProtoCOL colony counter if conditions 

were favorable. If precipitation of the test substance precluded automatic counting, and 

for tester strains with low spontaneous mutation frequency, counting was done by hand. 

Evaluation of results for validity/mutagenicity was done in accordance with OCSPP Test 

Guideline 870.5100. Historical laboratory control data of the negative and the positive 

controls were included. 

II. RESULTS

The investigators reported that no dose-response related or biologically relevant increases in 

revertant colony numbers of any of the five tester strains were observed following treatment with 
3-decen-2-one DP Number: 373085

PC Code: 068403 EPA File Symbol No.: 5481-LAI

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AMV-1018 at any concentration level, neither in the presence or absence of metabolic activation, 

in either experiment. In the pre-incubation experiments, cytotoxicity was noted in all five tester 

strains at a dose concentration of 5.0 μL/plate without metabolic activation and in tester strain 

TA 102 at a dose concentration of 5.0 μL/plate with metabolic activation. The reference 

mutagens employed in the control scenarios induced a distinct increase in revertant colonies, 

indicating the validity of the experiments. Therefore, the test substance is considered to be nonmutagenic in this bacterial reverse mutation assay.

III. DISCUSSION AND CONCLUSION

A. INVESTIGATORS CONCLUSIONS

The investigators conclude that, under the conditions of this study, AMV-1018 is non-mutagenic.

B. REVIEWERS COMMENTS

The reviewer agrees with the investigators conclusion that, under the conditions of this study, 

AMV-1018 is non-mutagenic. There were no significant deviations from U.S. EPA Health 

Effects Test Guidelines, OCSPP 870.5100. The study is classified as acceptable/guideline 

(OCSPP 870.1500) and satisfies the requirements for a reverse mutation assay using bacteria. 

Signed and dated Data Confidentiality, Good Laboratory Practice (GLP) Compliance, and 

Quality Assurance statements were provided. 

C. STUDY DEFICIENCIES

No significant study deficiencies that would invalidate the study outcome were found. 

IV. REFERENCES

Ames, B.N., Durston, E.W., Yamasaki, E. and Lee, F.D. (1973) Carcinogens are mutagens: a

simple test system combining liver homogenates for activation and bacteria for detection. 

Proc. Natl. Acad. Sci. (USA) 70: 2281-2285. 
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EPA Primary Reviewer: 

Gina M. Casciano, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

EPA Secondary Reviewer: 

Colin G. Walsh, Biochemical Pesticides Branch Signature: ______________

Biopesticides and Pollution Prevention Division (7511P) Date: ______________

DATA EVALUATION RECORD

STUDY TYPE: In vitro Mammalian Cell Gene Mutation Test 

(OCSPP 870.5300)

MRID NO: MRID 47942615

DP BARCODE NO: 373086

DECISION NO: 425385

SUBMISSION NO: 864619

TEST SUBSTANCE: AMV-1018 

active ingredient: 3-decen-2-one

purity: 98%

PC CODE: 068403

SPONSOR: AMVAC CHEMICAL CORPORATION 

4100 East Washington Blvd.

Los Angeles, CA 90023-4406

TESTING FACILITY: BSL BIOSERVICE Scientific Laboratories GmbH 

Behringstrabe 6/8

82152 Planegg, Germany

TITLE OF REPORT: In vitro Mammalian Cell Gene Mutation Assay 

(Thymidine Kinase Locus/TK+/-) in Mouse Lymphoma 

L5178Y Cells with AMV-1018

AUTHOR: Michael Kraft

STUDY COMPLETED: September 1, 2009
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STUDY NO: 091462

GOOD LAB PRACTICE: GLP (Good Laboratory Practice) Compliant

CONCLUSION: Test substance is considered to be mutagenic under the 

conditions of the 24 assay without metabolic activation, 

presented here. Other results remain equivocal.

CLASSIFICATION: ACCEPTABLE

EXECUTIVE SUMMARY: In a mammalian cell gene mutation assay at the TK +/- locus

(MRID 47942615), L5178Y mouse lymphoma cells cultured in vitro were exposed to AMV1018 (3-decen-2-one; 98.57% a.i.) in dimethyl sulfoxide (DMSO) at the following 

concentrations: 0.05, 0.15, 0.22, 0.28, 0.31, 0.34, 0.37 mM (+ S9) for 4 hours; 0.005, 0.01, 0.02, 

0.05, 0.10, 0.12, 0.14, 0.16 mM (- S9) for 4 hours; 0.10, 0.15, 0.19, 0.23, 0.27, 0.31, 0.35, 0.38 

mM (+ S9) for 4 hours; and 0.0001, 0.0007, 0.004, 0.007, 0.014, 0.028, 0.034, 0.04 mM (- S9) 

for 24 hours. The S9 fraction (for metabolic activation) was derived from the livers of male 

Wistar rats induced with Phenobarbital (80 mg/kg bw) and β-Naphthoflavone (100 mg/kg bw). 

The solvent DMSO served as a negative control in the presence and absence of S9. 

Benzo(α)pyrene (BP) served as a positive control in the presence of S9. Ethylmethanesulfonate 

(EMS) and methylmethanesulfonate (MMS) served as positive controls in the absence of S9. 

Selection of test substance concentrations were based on a pre-experiment for cytotoxicity. No 

precipitation of the test substance was noted in the experiments. Growth inhibition was noted in 

all experiments (+/- S9), with marked cytotoxicity seen in several cases (one incident less than 

10%, several less than 20%). The pH values for the highest concentrations tested were within the 

physiological range. The osmolality for the solvent controls as well as for the highest test 

concentrations was found to be 465 mosmol/kg. Thus, the osmolality was above the 

physiological range. 

The reviewer agrees with the investigators conclusions that, under the conditions of this assay, 

the test substance AMV-1018 (3-decen-2-one) is mutagenic. However, it is not clear that the 

results, given the increase osmotic pressure in particular and the marked cytotoxicity secondarily, 

would translate into an in vivo system. In addition, equivocal results in the experiments with 

metabolic activation should be clarified with a follow-up experiment. 

The study is classified as acceptable/guideline and satisfies the guideline requirements (OCSPP

870.5300) for in vitro mutagenicity (mammalian forward gene mutation) data.

COMPLIANCE: Signed and dated Data Confidentiality, GLP Compliance, and Quality 

Assurance statements were provided.
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I. MATERIALS AND METHODS

A. MATERIALS

1. Test Material: AMV-1018

Description: Pale yellow liquid

Batch #: 3D2-2009/01

Purity: 98% a.i.

CAS # of TGAI: 10519-33-2

2. Control Materials

Negative control: The solvent dimethylsulfoxide (DMSO, 1%) served as the negative control.

Positive control: Non-activated: Ethylmethanesulfonate (EMS, 200 μg/mL and 500 μg/mL) 

Methylmethanesulfonate (MMS, 10 μg/mL)

Activated: Benzo[a]pyrene (B[a]P, 2.5-3.5 μg/mL)

3. Activation: The S9 fraction was derived from male Wistar rats (age, weight, and supplier 

not reported).

X Induced Aroclor 1254 X Rat X Liver

Non-induced X Phenobarbital Mouse Lung

None Hamster Other 

X Other (βNaphthoflavone)Other 

The S9 fraction was prepared at BSL BIOSERVICE GmbH (the testing facility). Quality control measures 

included a Salmonella typhimurium assay for biological activity and a sterility test. A stock of the 

supernatant containing microsomes was frozen in ampoules of 2 and 4.5 mL and stored at ≤ -75o

C. For 

preparation of the S9 mix, supernatant was thawed and mixed with cofactors to reach the following 

concentrations: 8 mM MgCl2, 33 mM KCl, 5 mM Glucose-6-phosphate, and 5 mM NADP, in a solution of

100 mM sodium-phosphate buffer, pH 7.4. 

4. Test Cells: Mammalian cells in culture

X Mouse lymphoma L5178Y cells V79 cells (Chinese hamster lung fibroblasts)

Chinese hamster ovary (CHO) cells list any others

Properly maintained? X Yes No

Periodically checked for Mycoplasma contamination? X Yes No

Periodically checked for karyotype stability? X Yes No

Periodically "cleansed" against high spontaneous background? X Yes No
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Media: Growth medium - RPMI 1640 medium, supplemented with 15% horse serum, 100 μg/mL 

penicillin/streptomycin, 1 mM sodium pyruvate, 2 mM L-glutamine, 25 mM HEPES, and 250 μg/mL 

amphotericin B; Treatment medium - growth medium with 3% horse serum for short-term exposure and 7.5% 

horse serum for long-term exposure; and Selection medium - growth medium supplemented with 5 μg/mL 

trifluorothymidine.

5. Locus Examined

X Thymidine kinase (TK) Hypoxanthine-guaninephosphoribosyl transferase (HGPRT)

Na+/K+

ATPase

Selection 

agent:

Bromodeoxyuridine (BrdU) 8-azaguanine (8-AG) Ouabain

Fluorodeoxyuridine (FdU) 6-thioguanine (6-TG)

X Trifluorothymidine (TFT; 5 μg/mL)

6. Test Compound Concentrations

Pre-test for Cytotoxicity, Activated: 0.005, 0.01, 0.05, 0.1, 0.2, 0.4 mM

(0.77, 1.54. 7.71, 15.43, 30.85, 61.70 μg/mL)*

Pre-test for Cytotoxicity, Non-Activated: 0.005, 0.01, 0.05, 0.1, 0.2 mM

(0.77, 1.54. 7.71, 15.43, 30.85 μg/mL)*

Mutagenicity Assay, Activated (4 hr): 0.05, 0.15, 0.22, 0.25, 0.28, 0.31, 0.34, 0.37 mM

(7.71, 23.14, 33.94, 38.56, 43.19, 47.82, 52.45, 57.07 μg/mL)*

Mutagenicity Assay, Non-Activated (4 hr) 0.005, 0.01, 0.02, 0.05, 0.10, 0.12, 0.14, 0.16 mM

(0.77, 1.54, 3.09, 7.71, 15.43, 18.51, 21.60, 24.68 μg/mL)*

Mutagenicity Assay, Activated (4 hr): 0.10, 0.15, 0.19, 0.23, 0.27, 0.31, 0.35, 0.38 mM

(15.43, 23.14, 29.31, 35.48, 41.65, 47.82, 53.99, 58.62 μg/mL)*

Mutagenicity Assay, Non-Activated (24 hr): 0.0001, 0.0007, 0.004, 0.007, 0.014, 0.028, 0.034, 0.04 mM

(0.015, 0.108, 0.617, 1.080, 2.160, 4.319, 5.245, 6.170 μg/mL)*

*molecular weight of the test substance is 154.25 g/mol

B. TEST PERFORMANCE

1. Cell Treatment

a. Cells were exposed to test compound, negative/solvent or positive controls for 4 hours 

(activated and non-activated) in Experiment I, and for 4 hours (activated) and 24 hours (nonactivated) in Experiment II.

b. After washing, cells were cultured during the expression period for 72 hours after the 4 

hour experiments and 48 hours after the 24 hour experiment at 37°C in 5% CO2/95% 

humidified air before cell selection.

c. After expression, a statistical number of 1.6 cells/well (192 wells/group) were cultured 

for 6 days without selective agent to determine cloning efficiency (viability), and 2000 
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cells/well (384 wells/group) were cultured for 11-14 days in selection medium to determine 

numbers of mutants.

2. Statistical Methods: 

Mutant frequencies were calculated from the natural log (ln) comparison of the dividend of 

negative cultures to total cultures seeded, of the selective medium over the non-selective 

medium (see below). No further statistical methods (i.e. comparison or regression) were 

performed. 

Mutation frequency = -ln [NC / TC (selective medium)] x 800

-ln [NC / TC (non-selective medium)]

NC: number of negative cultures

TC: total number of cultures seeded

3. Evaluation criteria

a. Assay validity 

According to the study authors, the assay was considered valid if the following criteria were 

met:

• The mutant frequencies in the negative or solvent control cultures were in the range of 

BSL BIOSERVICE historical control data (about 82 to 253 mutants per 106 cells).

• The positive controls (EMS, MMS, and B[a]P) induce significant increases (at least 2-

fold increase of mutant frequencies related to the comparable negative control values and 

higher than the historical range of negative controls) in the mutant frequencies.

• At least three out of four 96-well-plates from the TFT resistance-testing portion of the 

experiment are scorable.

• The absolute cloning efficiency ([number of positive cultures x 100]/ total number of 

seeded cultures) of the negative and/or solvent controls is >50%.

b. Positive result 

According to the study authors, the test article was considered mutagenic if:

• There was a clear and dose-related increase in the mutant frequency. 

• There was a biologically relevant response (at least a 2-fold increase of mutant 

frequencies related to the respective negative control values and higher than the historical 

range of negative controls) for at least one of the dose groups. 

• If combined with a positive effect in the mutant frequency, an increased occurrence of 
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small colonies (slow growth colonies) indicated by a low large/small colonies ratio (1.5 

times the ratio of clastogenic controls MMS and/or B[a]P) is an indication for potential 

clastogenic effects and/or chromosomal aberrations.

II. REPORTED RESULTS 

A. PRELIMINARY CYTOTOXICITY ASSAY: The pre-test for cytotoxicity was 

performed on solvent controls, negative controls, and the test material concentrations 

reported above (section I.6.). Observation of test material precipitation was not reported. 

Marked cytotoxicity (<10-20% relative survival) was seen at the highest test material 

concentration with and without metabolic activation (6.60% RSG at 0.4 mM with metabolic 

activation; 4.74% RSG at 0.2 mM without metabolic activation). When comparing the results 

of these experiments across test material concentration levels, higher cytotoxicity is observed 

in the experiments without metabolic activation than with metabolic activation. This data is 

reported in Tables 1-2 of the study report (page 24 of the study report) and are included as an 

Attachment to this DER. 

B. MUTAGENICITY ASSAY

The main experiment consisted of four parts: 

Experiment I, with metabolic activation (4 hr assay)

Experiment I, without metabolic activation (4 hr assay)

Experiment II, with metabolic activation (4 hr assay)

Experiment II, without metabolic activation (24 hr assay)

Test compound concentrations for each experiment are listed above (section I.6.). The pH 

values of the highest concentrations tested were within the physiological range. The 

osmolality for the solvent controls as well as for the highest concentrations were found to be 

465 mosmol/kg. Thus, the osmolality was above the physiological range. No precipitation of 

the test substance was noted in the experiments.

Dose-response related growth inhibition was observed in all four experimental trials. 

Mutant frequency (i.e. mutation factor, MF) was determined for all test substance

concentrations, and colony sizing was performed for the highest concentrations of the test 

item and for the negative and positive controls. EMS (500 and 200 μg/mL), MMS (10 

μg/mL) and B[a]P (3.5 μg/mL) were used as positive controls and showed distinct and 

biologically relevant effects in mutation frequency in all experiments.

In Experiment I with metabolic activation, all mutant values were within the range of 

historical control data of the test facility (93-253 mutants per 106 cells, Table 16, page 40 of 

the study report). The study authors report that no dose-response relationship could be 
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observed, however, the reviewer notes a dose-response related increase at the three highest 

test substance concentrations. The study authors report that the mutation frequencies found in 

the groups treated with the test item did not show a biologically relevant increase as 

compared to the solvent controls. This is based upon a mutation factor upper limit of 2.0 

(when compared to the solvent controls) for a negative outcome. The highest mutation factor 

observed was 1.91 and was seen in the highest dose group for this experiment (0.37mM, 

RTG of 12.22%). In addition, colony sizing (performed on the three highest dose groups) 

revealed an increased number of small colonies as compared to the negative and solvent 

controls. However, because no corresponding mutagenicity was found in this experiment (i.e. 

MF > 2.0), the investigators concluded that no clear statement could be made regarding these 

results, and therefore, to clarify these findings an independent repetition experiment was 

performed (see Experiment II with metabolic activation). The data for this experiment 

appears in Tables 3-5 (pages 25-26) of the study report, and are attached to this DER. 

In Experiment I without metabolic activation, all mutant values found were within or slightly 

above the range of historical control data of the test facility (82-238 mutants per 106 cells, 

Table 16, page 40 of the study report). A slight dose-response relationship could be observed. 

The highest mutant value of 265.61 mutants/106 cells at a concentration of 0.16 mM was 

regarded as not biologically relevant due to a mutagenicity factor (when compared to the 

solvent controls) of only 1.85. RTG for this dose level was 13.17%. Colony sizing was 

performed and all dose groups were considered as not clastogenic. The data for this 

experiment appears in Tables 6-8 (pages 27-28) of the study report, and are attached to this 

DER.

In Experiment II with metabolic activation, the mutant values at doses of 0.27, 0.31, and 0.38 

mM clearly exceeded the range of the historical control data (93-253 mutants per 106 cells, 

Table 16, page 40 of the study report), and a slight dose-response relationship was observed. 

In two dose groups (0.31 and 0.38 mM) the threshold value of 2.0 for the mutation factor was 

slightly exceeded. The mutation factor (compared to the solvent control values) at a 

concentration of 0.31 mM (with a RTG of 36.5 1%) was 2.22, and at a concentration of 0.38 

(with RTG of 9.2%) was 2.15. The investigators conclude that, since in no mutagenicity was 

evaluated up to a RTG of 12.22% in Experiment I with metabolic activation, the results for 

the experiments with metabolic activation are equivocal (i.e., inconclusive). For colony 

sizing, increases in the number of small colonies noted at doses of 0.31 mM and 0.38 mM 

lead the investigators to conclude a clastogenic effect of the test item because the threshold 

value for mutagenicity was exceeded in these same dose groups. The data for this experiment 

appears in Tables 9-11 (pages 29-30) of the study report, and are attached to this DER.

In Experiment II without metabolic activation, all mutant values found up to a dose of 0.014 

mM (2.160 μg/mL) were within the range of the historical control data (82-238 mutants per 

106 cells, Table 16, page 40 of the study report), however, doses of 0.028, 0.034, and 0.04 
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mM (4.319, 5.245, and 6.170 μg/mL) clearly exceeded the historical control range. In 

addition, in these dose groups the threshold MF value of 2.0 (when compared to the solvent 

control value) was exceeded. Mutation factors for the doses with mutants above historical 

control data were 2.84 at 0.028 mM with a RTG of 29.93%, 3.63 at 0.034 mM with a RTG of 

17.51%, and 4.13 at 0.04 mM with a RTG of 13.16%. These are considered by the study 

authors to be significant, indicating mutagenicity of the test compound in this 24-hour assay. 

Colony sizing revealed an increase of small colonies at the 0.034 mM dose, leading the 

investigators to conclude that this “is a hint at clastogenicity of the test item” because

corresponding mutagenicity was found in this dose group. The data for this experiment 

appears in Tables 13-15 (pages 32-33) of the study report, and are attached to this DER.

III. DISCUSSION AND CONCLUSIONS

A. INVESTIGATORS’ CONCLUSIONS: 

The investigators conclude that the test item AMV-1018 is mutagenic in the mouse 

lymphoma thymidine kinase locus (cell line L5178Y) without metabolic activation. Results 

for the experiments with metabolic activation are considered to be equivocal (i.e., 

inconclusive).

B. REVIEWER COMMENTS: 

OCSPP Test Guideline 870.5300 states that the criterion for determining a positive result in 

the in vitro Mammalian Cell Gene Mutation Test includes biological relevance and a 

concentration-related, reproducible increase in mutant frequency. Furthermore, the Guideline 

indicates that although most studies will give clearly positive or negative results, in rare cases 

the data set will preclude making a definite judgment about the activity of the test substance, 

with results remaining equivocal or questionable regardless of the number of times the 

experiment is repeated. In the case of AMV-1018 (3-decen-2-one) described above, the 

numbers of mutants per 106 cells exceeded the laboratory’s historical control data at 

particular dose levels in experiments with and without metabolic activation. In Experiment II 

without metabolic activation, these results exhibit a clear dose-response relationship. In 

addition, colony sizing revealed an increase in small colonies in all experiments performed. 

An increase in small colonies in addition to high mutant frequencies, as seen in Experiment II 

both with and without metabolic activation, indicates clastogenicity of the test substance. 

However, the Guideline also states that care should be taken to avoid conditions which would 

lead to results not reflecting intrinsic mutagenicity, which therefore would not be biologically 

relevant when translated to a living system. Such positive results which do not reflect 

intrinsic mutagenicity may arise from changes in pH, osmolality, or high levels of 
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cytotoxicity (OCSPP 870.5300; Scott, 1991; Moore, 2003). In the assay in question, the 

investigators report that pH values of the highest concentrations tested were within the 

physiological range, however, osmolality for the solvent controls as well as for the highest 

concentrations of test substance were both found to be 465 mosmol/kg. Thus, the osmolality 

was above the physiological range.

With regards to cytotoxicity, the Guideline states that if the maximum concentration of test 

substance investigated is based on cytotoxicity (rather than solubility) then it should result in 

approximately 10–20 percent, but not less than 10 percent, relative survival (i.e., relative total 

growth, RTG). This principle was generally adhered to in the assay, with only one dose level 

investigated (0.38 mM, Experiment II with metabolic activation) producing a RTG of less 

than 10%. However, of MF values exceeding 2.0 (five in total), three occurred at RTG levels 

less than 20%. This may be of significance when determining the validity of the assay. The 

Mouse Lymphoma Assay (MLA) MLA Workgroup of the International Workshop on 

Genotoxicity Tests (IWGT), assembled in 2002, considered the general issue of biological 

relevance in the assay and concluded that very small increases in MF can often be seen at 

high cytotoxicity levels, and that mutations induced under these circumstances would not 

normally occur in vivo and thus are not considered by the Workgroup to be biologically 

relevant (Moore, 2003). Cytotoxicity levels less than 20% and osmolality outside the 

physiological range are of concern to the reviewer. 

The OCSPP Guideline states that a positive result for the in vitro mammalian cell gene 

mutation test which indicates mutagenicity and is reproducible is most meaningful. There 

appears to be reproducible evidence of mutagenicity in this assay with and without S9 

activation. However, there is also evidence of marked cytotoxicity, and the osmolality for all 

experiments was outside the physiological range. The reviewer agrees with the investigators 

conclusions that, under the conditions of this assay, the test substance AMV-1018 (3-decen2-one) is mutagenic. However, it is not clear that these results, given the increase osmotic 

pressure in particular and the marked cytotoxicity secondarily, would translate into an in vivo

system. In addition, equivocal results from the experiments with metabolic activation should 

be clarified with a follow-up experiment. 

The study is classified as acceptable/guideline and satisfies the guideline requirements 

(OCSPP 870.5300, OECD 476) for in vitro mutagenicity (mammalian forward gene 

mutation) data.

C. STUDY DEFICIENCIES: None

D. REFERENCES CITED
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Moore, M.M. et al. (2003) Mouse Lymphoma Thymidine Kinase Gene Mutation Assay: 

International Workshop on Genotoxicity Tests Workgroup Report – Plymouth UK, 2002. 

Mutation Research 540: 127-140.

Scott, D. et al. (1991) Genotoxicity Under Extreme Culture Conditions. A report from 

ICPEMC Task Group 9. Mutation Research 257: 147–204.
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ATTACHMENT

Pre-test for Toxicity: 

Tables 1-2 from page 24 of MRID 47620516

Main Experiment I, with metabolic activation: 

Tables 3-5 from pages 25-26 of MRID 47620516

Main Experiment I, without metabolic activation: 

Tables 6-8 from pages 27-28 of MRID 47620516

Main Experiment II, with metabolic activation: 

Tables 9-11 from pages 29-30 of MRID 47620516

Main Experiment II, without metabolic activation: 

Tables 13-15 from pages 32-33 of MRID 47620516
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cc: G. Casciano, C. Walsh, R. S. Jones, D. Benmhend, BPPD Science Review File, IHAD/ARS

G. Casciano, Biologist, FT, PY-S: 12/07/2010