## Data Evaluation Record on tbe Acute Toxicity ofTebufenozide to Aqu.atic Vascular Plants, 

## Lemnagibba

_ _ _____ _ _ _ _ _ _____________ _::EPA MRJD Number 485330-03 Data Requirement:

Test material: Tebufenozide Common name Chemical name : IUPAC EPA DP Barcode OECD Data Point EPA MRID EPA Guideline 0392144 OECD Guideline 221 485330-03 850.4400 Purity: 98.6% CAS name : 3 ,5-dimethylbenzoic acid l-(1,1-dimetbylethyl)-2-(4-ethylberzoyl)hydrazide CAS No.: 112410-23 -8 Synonyms Primary Reviewer: Joan Gaidos Signature: Senior Scientist, Cambridge Environmental, Inc
. Date: 3/ 15/12 Primary EPA Reviewer: Andrew Sayer

**~ ~~** 



# 4 EP A/OPP /EFEDIERB4

- I - Date: 5/10112 Secondary EPA Reviewer(s): Thomas Steeger,



### Ph **.~ ~~Date:**  5/1 011 2 EPA/OPP/EFEDIERB4 y

**-~~** EPA PC Code 129026 Date Evaluation Completed: 4/27/ 12 CITATION: Porch, J. 2011. Tebufenozide: A 7-Day Static-Renewal Toxicity Test with Du ckweed 

### (LemnagibbaG3). Unpublished study performed by Wildlife International, Ltd. , Easton, Maryland. Laboratory project no.: 437A122A
. Study sponsored by Nippon Soda Co ., Ltd., Chiyoda-ku, Tokyo , Japan. Study completed July 7, 20II . EXECUTIVE SUMMARY: In a 7-day acute toxicity study, the freshwa ter floating aquatic vascular plants duckweed 

### Le ,nna gibba were exposed to tebufenozide technical grade active ingredient (98.6% ai) at nominal concentrations ofO

**(r eg**  **ative** control), 0 (solvent [DMF] contro l ) , 0.031, 0.063, 0
. 13 , 0.25, 0.50 and 1.0 mg ai/L under static
.renewal conditiors. Measured concentra tions were <LOQ (<0
. 0200, negative and solvent [DMF] controls), 0.031, 0.064, 0.13, 0.26 , OA5 , and 0.94 mg ai/L
. All comparisons were made relative to the negative control. Frond density in the negative control averaged 273 fronds per rep yie lding inhibitions of

-2 , I , I0, 2, 0 and 9% in the mean-measured 0.031, 0.064 , 0.13 , 0.26 , 0.45, and 0.94 mg ai/L treatment groups as compared to the control. The NOAEC and EC50 values were 0.94 and >0.94 mg ai/L, respectively (based on mean-me as 1 tred concentrations). Frond biomass (dry weight) averaged 32.4 mg in the negative control , yielding inhibitions o f -4 , -4, -2, -11, -1 , and 0% in the mean-meas ured 0.031, 0.064 , 0.13, 0.26, 0.45 , and 0 .94 mg ai/L treatment groups, rcsp t :ctively. The NOAEC and EC5 0 values were 0
.94 and >0.94 mg ailL, respectively (based on mean-measured concentrations)
. Growth rate (based on frond density) averaged 0.446 days" 1 in the negative control, yielding inh ibitions of

### -1, 0, 3, 

### I , 0 , and 3% in the mean-measured 0.031, 0.064, 0.13, 0.26 , 0.45, and 0
. 94 mg ai/L treatment groups, respectively. The NOAEC and EC 50 values were 0
.94 and >0 .94 mg ailL, respectively (based on mean-me asttred concentrations). After 

### 7 days ofexposure, the number ofchlorotic fronds observed in the 

### 0.031, 0.064 , 0.1 3, 0.26, 0.45 and 

### 0.94 mg/ L treatment groups was I, I , 0, 0, 2 and 0 representing less than I% ofthe total number o ffr ords in each group. Necrotic Page I of 15Data Evaluation Re c ord on the Acute Toxicity of Tebufe . nozide to Aquatic Vascular Plants, Lemnagibba EPA M RID N umber 485330 - 03 fronds in the 0.031, 0.064 , 0. 1 3, 0.26 , 0.45 and 0 . 94 mg/L also accounted for l ess than 1% of the total frond number in

each group . Plants in the ne g ative and so l vent control groups and in all of the treatment groups appeared norma l throughout the t est with n o overt signs of toxicity observed. This toxicity s t udy is**clas ific** **d** as scient i fically sound and is cons i stent with guideline recommendations for a T ier II aquatic p l ant toxicity study using t h e aquatic vascular plant Lemna gibba. Results Synopsis Test Organ i sm: Lem n a gibba

T est Type: Sta t ic r en e wal Frond density ECo s: NA 95% C. L: N/ A EC 50 : >0 . 94 mg ai ,. L 95% C .l. : N / A NOAEC: 0.94 mg ai / L L OAEC: >0 . 94 m g a i / L Biomass EC 05 : NA 95% C. L : N/ A EC 50 : >0 . 94 mg ai i L 95% C . I.: NIA NOAEC: 0.94 mg ai t L L OAEC: >0.94 mg a i/L Growth rate (based anj r oiUI density) EC 05 : NA 95%**.:** ';:-J / A ECso: >0 . 94 mg ai · 'L 95% C.l.: N / A NOAEC: 0.94 mg ai l L L OAEC: > 0 . 94 mg a i/L Endpoint(s) Effec t ed : Cou l d not be determined due to a lack of toxicity. M ost sensitive endpo i nt: C ou l d not be determine d due to a lac k of t oxicity. I. MATERIALS AND M ETHODS GUIDELINE FOLLOWED: This study was co n ducted acco r ding to a protocol based on procedures outlined in the U.S. Environmenta l Pr o tect i on Agency Ser i es 850 - Ecolog i cal Effects T est Gu i delines (draft), OP P TS N u mber 850.4400 (1996). D eviations from OPPTS 850.4400 were no t ed : COMPLIANCE :

l. The pH at test i n i tia t ion was already higher than recommended at 8.0-8.1. By tes t t ermination t he pH was 9.2-9.3; O P PTS guide li nes suggest that pH remain in the range of7 . 5 ± 0.1. 2. T h e physico-chemical properties of the test material were not reported. These deviations did not impact the accep t abi l ity of the study . Signed and dated GLP , Data Confiden t ia l ity, and Qua l ity Assurance statemen t s P age 2 of 15

## Data Evaluation Record on the Acute Toxicity of Tebufenozide to Aqt.atic Vascular Plants, 

## Lemnagibba

**--- ---**  **-- --**  **--- --**  **---- --**  **-- -=E**  **P~A** MRIO Number 4853 30-03

A. MATERIALS : 

### 1. Test material Description: Lot No./Batch No . : Purity: Stability of compound under test conditions:

Storage conditions of test chemicals: were provided. This study was conducted in 

### : ompliance with the Good Laboratory Practice Standards as published by the U.S. Environmental Protection Agency , 40 CFR Part 160 (1989) ; OECO Principles of GLP (ENV/MC /CHEM (98) 17); and Japan MAFF (I I NohSan, Notification No. 6283 , Agri cultural Production Bur ea

u, 1999), wilh the following exception: Periodic analyses of well water an d soil fo r pote:ttial contaminants were not performed in accordance with GLP standards, tut were performed using a certified laboratory and standar d U.S. EPA analy-ical methods.
Tebufenozide (TGAI) Solid 6326308 98.6%

Method validations using fortified matrix samples yielded recoveries of97. 1± 

### 6.3%. Analytical determinations performed at time 0 yielded recoveries ranging from 98.3 to 106% of nominal. At 3 (oltl so luti on prior to renewal) and 5 (new solution at renewal) days, recoveries were 97.6% to 109% of nominal, respectively. At 7 

### days (old sol ution at test termination) , recoveries were 96
.2% to II

### 0% ofnominal. Based on these data, the test material was stable under the conditions. (OECD recommends water solubility , stabilit y in wat er and light, pKa , Pow , and vaporpressure oftest compound) 

### The test material was stored at ambient condit10m
. in the dark. Table 1 

### lists the chemical physical properties oftebufenozide. T bl 1 Ph
. h . 1

a e IYSICOC 

### em 1 ca propert' 

### fT b

**~** 'd 

### 1 es 0 

### e u enoz• e. Parameter Values Comments Water solubility 0.83 ppm OP Barcode 0226011 Vapor pressure 2.25 X I 

### 0 ' 8 

### mm Hg OP Barcode 0226011 UV absorption Not reported


# -- pK, Not reported 

## - 4.25 Page 3 of 15Data E v a luation**Re, ord**  on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, Lemnagibba EPA MRID Number 485330-03 Parameter Values C om ments logKow DP Barcode 0226011 2 . Test organism: Name: Du ck we ed, Lemna gibba G3 Strain, if provided: No t reported Source: In- h ous e cultures w ere origina ll y ob tain e d from t h e U .S. Department of Agriculture and maintained i n culture medium at W il dlife In t ernational, Ltd., Eas ton, Maryla nd Age of inoc u l u m : The inoculum was taken from a stock culture that had been transferred to fresh medium 7 days before testing Method of**ul tivation** **:** Plants we r e cultured in 20X Algal Assay Proc edure (AA P) culture medium B . STUDY DESIG N : I. Experimenta l C o nditions a. Ran ge-finding study : Non e reported. b. Definit iv e Study T bl 2 E a e : xper.menta I P · ara m eters Pa rameter Details Remarks ----- --------------- Criteria - Acclimation period: 7 da y s C ultu r ing media and co n di tion s : Duckweed were acclimated to the (same as test or no t ) culture medium; th e cu l turing conditions we r e the same as test Health: (any m ortality ob served) Not reported Test system Fronds were transferred to newly S tati c / static renewal Static-renewal prepared test solutions on days 3 and 5. Renewal r ate fo r stat i c r t · mewal N / A ----- ----- ----- ----- EPA expects the test concentrations to be renewed every 3 to 4 days (one renewal for the 7 day test , 3-4 renewals for the 14 day test). Test vesse l s w e r e placed in a Incub at i o n facility t emperature-contr?lled environmental c hamb e r. Page 4 of 15

## Data Evaluation Record on the Acute Toxicity of Tebufenozide to Aqu atic Vascular Plants, Lemnagibba

Param eter

Duration ofthe test

Test vessel Material: (glass/stainless ste el) Size: Fill volume:
Details ofgrowth medium name pH at test initia tion: pH at test termination: Chelator used: Carbon source : 7 days

Glass 250mL 100 

### rnL

8 .0-8
. I 9.2

-9.3 Yes NaHC0 3 

### Details

If non-standard nutrient medium N/A was used, detailed composition provided (Yes/No)
Dilution water source /type: pH: water pretreatment (if any) : Total Organic Carbon: particulate matter: metals: pesticides: ch lorine: Purified well water Adjusted to 7.5 

### ± 0
. I if necessary Water was sterilized Not reported Not reported Not reported Not reported Not reported
Page 

### 5 of 15 EPA MRJD Number 485330-03
R emarks Crit e ria

EPA r equ.res a duration of 14 days
. Seven day studies will be accept ed for review by tl, e Ag ency .



### EP A re commends the following c ulture me dia: ModifiedHoagland ' s E+ or 2 0X-AAP. 

### C helating agents (e.g. EDTA) are recommend ed in the nutrient medium for optimum cell growt h. l ower concentrations of chelating agents (down to one-third of the normal concentration recommended for AAP medium) may be used ir. the nutrient medium used for t est SOJ'ution preparation 

## if it is suspected that the chelator will interact w ' th the test material
. ASTM reference, £1415-9/and D 3978-80 ( reapproved 19 87)
.

EPA rt?COt1mends a pH of

- 5
. 0
. A solution p !J of 7.5 is acceptable 

### iftype 20X-AAP n ltrient media is used
.Data Evaluation Re c ord on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, Lemnagibba EPA MRJD Number 485330-03 -

Parameter Details Remarks -------------------- Criteria Indicate how the test ma t erial i s A primary stock so l ution was Concentration of DMF in solvent

added to the medium (a d ded prepared by m i x i ng a calculated control was 0.1 mL DMF /L. directly or used stock so ution ) amount (no t reported) of tes t material into solvent dimethylformamide (DMF) and bringing i t to volume with 20X AAP medium in a 500 mL vessel. Aeration or agitation Neither Sediment used (for root 2 d ag u atic N I A vascular plants)

Origin: Textural classification**(~ 'Osand,**  silt, and clay) : Organic carbon(%): Geographic locat i on:

Numb e r of reQiicates Control : 3 Solvent control: 3 Treatments : 3 Number of p l ants / rep l ic R te 12 plants per replicate - ---- - -------------- EPA requires 5 plants. Num b er of fronds/p l ant 4 fronds per plant --- - --- ------------- EPA requires 3 fronds per plant. T est concentrations Nominal: 0 (negative contro l ), 0 (solvent ---------- - - ------- - control), 0.031, 0.063, 0.13 , 0.25, EPA requires at least 5 tes t 0 . 50, and 1 . 0 mg ai!L concentrations with a dose range of2X or Mean-Measured : 3X progression. < LOQ ( < 0.0200 , nega t ive and solvent [DMF] con t ro l ), 0 . 031, 0.064, 0. 1 3 ,

0.26, 0 . 45 , and 0.94 mg a i / L Solven t (type, percentag e, i f used) Dimethy lf ormamide (DMF), 0.1 mL DMF/L Page 6 of 15

### Data Evaluation Record on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, 

## Lemnagibba EPA MRJD Number 485330-03 

### Parameter Details Rema rks



# --- -- - ---- ----- -- -- - Criteria


# - Method and interval of analytical Samples taken from day 0, day 3 (old) verification and day 5 (spent) so lutions and day 7 spent solutions were analyzed via HPLC with UV detection (wavelength of220 run). Test conditions Temperature: 24.0-26.7°C Photoperiod: Continuous Light intensity an d quality: 6220-6790 lux Warm-white fluore scent bulbs Reference chemical (ifused} name: N/A concentrations: Other parameters , if any None



### 2. Observations : T bl 3 Ob

a 

### e : 

### servahon parameters 

# I Parameters 

# I Details 

# I Renarks/Criteria 

# I

Parameter s measured (e.g.,:

- frond density nu

mber of fronds , plant dry weight -b iomass (dry weight) or other toxicity symptoms) -growth rate (frond density)

- growth rate

- frond condition Measurement technique for frond Frond density was determined via number and other end points direct counting. Frond condit ion was vi sually determined. Biomass was measured by drying the plants at 60 ° C for 

### ca. 48 hours prior to weighing. Growth rate s were determined using equations that took into account frond number or biomass at the beginning and end ofthe test. Observation intervals Frond density and condition were determined at days 0 , 3 , 5 , and 7. Page 7 of 15Data Evaluation**Re ord**  on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, Lemnagibba EPA MRID Number 485330-03 I Parameters = I Details I Remarks/Criteria I Biomass was determined at study initiation and termination. Other observations, if a n y None Indicate whether there ""'as a n Yes; frond density was 273 fronds exponential growth in th e control per repl i cate in the control at 7 days . Were raw data included ? Yes II . RESULTS and DISC U SS I ON : A. INHIBITORY E F FECTS: According to the stud y authors, frond density in the negative control averaged 273 fronds per rep y i elding inhibitions of - 2, I, I 0, 2, 0 and 9% i n t h e mean-measured 0.031, 0.064, 0.13, 0.26, 0.45 , and 0.94 mg ai / L trea t ment groups as compared to the con tr o l ( T able 4) . Percent inhibition was based on pooled controls. The study author-reported

NOAEC and ECso va ' ues were 0 . 94 and >0.94 mg ai / L, respectively (based on mean - measured concentrations). Frond biomass (dry wei gh t) averaged 32.4 mg in the nega t i ve control, yielding inhibitions of -4, -4, -2 , -II, -I , and 0% i n the mean-mea s ured 0. 031, 0.064, 0.13, 0.26, 0.45, and 0 . 94 mg ai/L treatment groups, respectively (Table 4a). The study auth o r- r e p orted NOAEC and EC 50 values were 0.94 and >0.94 mg ai / L , respectively (based on mean-measured con etm tra t ions). Growth rate (based o n fr o n d density) averaged 0.446 days·• in the negative control , yielding inhibitions of - 1, 0, 3, I, 0, and 3% in the mea : 1- m easured 0 . 031, 0.064, 0 . 13, 0.26 , 0.45 , and 0.94 mg a ilL treatment groups, respectively (Table 4a). The stud)- author-reported NOAEC and EC 50 values were 0.94 and > 0.94 mg a ilL, respective l y (based on mean-measured conc e ntra t ions) . After 7 days of expo 5- ure , t he number of ch l orotic fronds observed in the 0.031, 0.064, 0.13, 0 . 26, 0.45 and 0.94 mg!L treatment groups was 1, I, 0, 0, 2 and 0 representing less than I %of the total number of fronds in each group. Necrotic fronds i n th e 0 . 031 , 0.064, 0.13, 0.26, 0.45 and 0 . 94 mg!L also accounted for Jess than I% of the total frond number in eac h group . P lants in the negative and solvent control groups and in all of the treatment groups appeared normal throughout the test with no overt signs of toxicity observed. Page 8 of 15

## Data Evaluation Record on the Acute Toxicity of Tebufenozide to AqtJ
. atic Vascular Plants, Lemna gibba EPA MRID Number 485330-03 T bl 4 Effi 

### f b fi 

### f d

a e 

### : ect o te u enozt e on ron number o fd k d 

### L ibb 

## * uc wee . , emna 

### g• a Treatment Mean-measured (and Frond number /replica t e (mean) Initial frond nominal) concentration number /test 7 days m

g ai/L solution 3 days 5 days fro nd m· mber % inhibition• 

### Negative control 12 35 95 27 3 N/A Solvent control 12 34 90 260 N/A 0.031 (0.031) 12 39 104 27 9 -2 0.064 (0. 063) 12 37 93 270 l 

### 0.13 (0.13) 12 36 93 247 10 0.26 (0.25) 12 35 90 269 2 0.45 (0
. 50) 12 34 92 

### 27t 0 0.94 ( 1.0) 12 34 88 24£ 9



### .. . 

## * Percent mhibitlon based on pooled controls a 

### Percent inhibition based on comparison to negative control. Table 4a: Effect of tebufenozide on g rowth rate and dry weight of duckweed, L e mna gibba

## * Growth rate Growth rate Dry weight Treatment Mean-measured Initial frond based on frond D ·y weight % % (and nominal) concentration mg number/test density Inhibition • (m g, mean) Inhibition • ailL solution (days · 1
, mean) 

### Negative control 12 0.446 N/A 32
.4 NIA 

### Solvent control 12 0.439 N/A 32.2 N/A 0.031 (0
. 031) 12 0.449 -1 33.9 -4 

### 0.064 (0.063) 12 0.445 0 33.6 -4 0 .13 (0.13) 12 0 .431 3 33.1 -2 0.26 (0.25) 12 0.444 1 35.9 -11 0.45 (0.50) 12 0.446 0 32.8 -1 

### 0.94 ( 1.0) 12 0.432 3 32.4 0



### . . . 

## * Per cent mhibition based on pooled controls a 

### Percent inhibition based on comparison to negative control.

Page 9 of 15Data Evaluation**Re ord**  on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, Lemnagibba EPA MRID Number 485330-03 Table 5: Statistical endpoi n t values (study author-reported) Statistical Endp o int Frond Density Biomass Growth rate (based on frond density) NOAEC (m g ai / L) 0 . 94 0 . 94 0.94 EC o s (95% CI; mg ai/L) N / A N/A N / A LOAEC (mg ai/L) >0.94 >0 . 94 >0.94 EC 50 (95% CI; mg ai/L) >0.94 >0 .94 >0 .94 B. REPORTED ST AT ISTICS: The study authors an a lyzed frond density, biomass, and growth rate (b ased on frond density and biomass) using mean-measured conc e ntrat i ons and SAS® (SAS Institute, Cary , NC; Versio n 8.2). T he data were tested for nonnality and homog .:: ne i ty of variance (p $ 0.05) using Shapiro- Wilks ' and Levene's tests, respectively. Th e data were then analyzed u 5- ing A N OV A and Dunnett's t-tes t. Since there was less than 50% inhibition in endpoin t s in any treatment group, day 7 EC , values were not calcu l ated. I nhib i tions were determined based on the negative controls . Page 10 of 1 5

## Data Evaluation Record on the Acute Toxicity ofTebufenozide to Aqu:atic Vascular Plants, 

## Lemnagibba EPA MRID Number 485330-03 

### C. VERIFICATION OF STATISTICAL RESULTS: Statistical Method: The reviewer analyzed frond density , biomass, and growth rate data using both initial-measured and mean-measured concentrations for analysis
. The replicate data were frrst tested for normality using the Shapiro Wilks ' tests and for homogeneity of variances using Levene ' s test in Tox stat 3.5. All endpoint data met the assumptions ofANOVA, and were thus analyzed using Dunnett' s test to determi ne t he NOAEC. There were no statistically significant inhibitions (a=0
. 05) compa red to the negative control for any < mdpoint. Analysis of frond density at 7 days yielded no significant differences (a=0.05)
. The EC 50 

### values f or each of the endpoints were empirically determined to be greater than the highest concentration tested
. The reviewer' s results , based on the statistical verification presented in Appendix I , are presented in the Executive Summary and Conclusions sections of this data evaluation record (DER)
. Based on Mean-measured Concentrations: 

### Frond density ECos : NA 95 % C. l.: N/A ECso : >0.94 mg ai/L 95 % C. 

### I.: N /A NOAEC : 0.94 mg ai/L LOAEC: > 0.94 mg ai/L Probit Slope : Could not be determined 

### Biomass ECo 5 : 

### NA 95% C.I.: N /A ECso: >0.94 mg ai/L 95% C. I.: N/A NOAEC: 0.94 mg ai/L LOAEC: >0.94 mg ai/L Probit Slope : Could not be determined 

### Growth rate (based on frond density) EC0 5 : 

### NA 95% C.l.: N/A EC 5

### o: >0
.94 mg ai /L 95 % C.I.: N/A NOAEC: 0.94 mg ai/ L LOAEC: > 0
.94 mg ai/L Probit Slope: Cou ld not be determined Endpoint(s) Effected: Could not be determined due to a lack oftoxicity
. Most sensitive endpoint: Could not be determined due to a lack oftoxicity. 

### D. STUDY DEFICIENCIES: None. 

### E. REVIEWER'S COMMENTS : The reviewer' s and the study authors' results were in agreement. The most sensitive endpoint could not be determined due to lack oftoxicity
. The reviewer' s results (and the study author ' s EC sJ value for growth rate) are presented in the Executive Summary and Conclusions sections ofthis data evalu a tion 'ecord (DER). The in- life phase ofthe study was conducted from May 20 to May 27, 20 II. 

### F. CONCL USIONS: Page II 

### of 15Data Evaluation Re e or d on th e Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, Lemnagihha EPA MRID Number 485330-03 This study is scientif : cally sound and classified as ACCEPT ABLE. The most sensitive endpoint could not be determined due to a l a ck of toxicity. The 7-day NOAEC and EC 50 values were 0.96 and >0 .96 mg ai / L, based on mean-m easured conc e ntra t ions and the absence of statistically s ignific ant effects at the highest concentration tested ..

Based on Mean-mea s ured Concentrations: F rond density EC 0 5 : NA 95% C. L: N/ A EC 50 : >0.9 4 mg aiiL 9 5% C.I.: N/A NOAEC: 0.94 mg ai 1 L LOA EC: >0.94 mg a i/L Probit Slope: Could n ot be determined Biomass ECo 5 : NA 95% C. l.: N/ A EC 5 0 : >0 .94 mg ai 1 L 95% C.l.: N/A NOAEC: 0.94 mg ai ' L LOAEC : >0 .94 mg a i / L

Probit Slope: Could n ot be determined Growtlr rate (based o n f r ond density) EC 05 : NA 95% C. I. : N / A EC 50 : > 0.94 mg a i / L 95% C.l.: N I A NOA EC: 0.94 mg ai / L LOAEC: >0.94 mg a i / L Probit Slope: Could n ot be determined Endpoin t( s) Effec ted : C o u l d not be determined due to a l ack of tox icity . Most sensitive endpoint: C ould not be determined due to a lac k of toxicity. III. REFERENCES: Organization for Eco n omi c Cooperation and Development. 2006. Guidelines for Testing of Chemicals , Guideline 221: Lemna sp . , Gro w th I n hibition Test. Adopted 23 March 2006. U.S . E nvi ronmental P ro t ection Agency. 1996. Series 850 - Ecological Effects Tes t Guidelines (draft), OPPTS Numbe r 850.4400: A quatic Plant Toxicity Test using Lemna spp., Tiers I and II. Washington , D . C . ASTM S tandard Guicle E 1415-91 E. 1991. Standard Guide for Conducting Acute Toxicity Tests with Lemna gibba GJ. American Socie t y for Testing and Materials , Philadelphia, PA. Microsoft Corporation, Microsoft Exce l2000 . Copyright 1985 - 1999. West, Inc. and David D. G u lley, 1996. TOXSTAT Version 3.5. Western Ecosystems Techno logy, Inc. Cheyenne, Wyoming. Page 12 of 15

### Data Evaluation Record on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, 

## Lemnagibba EPA MRJD Number 485330-03 

### APPENDIX I. OUTPUT OF REVIEWER'S STATISTICAL VERIFICATION: T itle: 3303 frond no . 7 day F ile: 3303 fd Transform: Shap i ro - Wi l k' s Tes t for Normality D 8587.3333 

### w = 0.9363 Critical 

## w w 0 .884 0 (alpha 0 .916 0 (alpha 0.01 0 .05 N N 24) 24 ) NO TR\NSFORMATION

Data PASS norma lity t es t (alpha = 0.01). Continue analys i s. Title : 3303 frond no . 7 day File: 3303f d Transform : N O TRJ;NSFORMATION Levene' s Test for Homogeneity of Var iance

SOURCE Between With i n (Error ) To ta l
Critical F ANOVA Table OF 

### ss

7 535. 3333 16 61 24 .6667 23 6660.0000
4
. 025 9 (alpha = 0
. 01 , df 2.657 2 (a l pha = 0 .05, df 7, 16) 7, 16) MS F 76.47 6 2 0.1 99 8 382.7917 (p-value = 0.9809)

Since F < Critical F FAIL TO REJECT Ho: All equal (al pha 0.01)
Title: 3303 frond no. 7 day File: 3303f d Transform : NO TRANSFORMATION ANOVA Table



# - -- -- -- - - --- - ---- ---------- - ---- - --- --- ------- -- - --- --- - -------·-- --- ---- - --- SOURCE OF 

### ss MS F

**------- ---**  **-- -----**  **--- ------**  **--- ----** 
**-- --**  **-- --**  **-----**
**-- ----**  **-- ---**  **--·~- ----**  **---- ---**  Between 7 3071.6250 4 38.8036 0.8176 W

ith in (Error ) 16 8587. 3333 536.7083



# ----- ---- - --- - --- ---- -- ---- - --- - ---------- - ----- - --- ------ --- -· ·--- - -- ------- Tot a l 23 116 58. 9583



# ----- ---- ----- --- --- --- - ---- --- - --- -- ----- ----- ---------- -- -- -· ·----- -- -- - --- (p

- va l ue 

### = 0
. 5864) 

### Page 13 

### of 1 5Data Evaluation Re • ; ord on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, Lemnagibba EPA MRID Number 485330-03 Critica l F 4 . 0259 (alpha = 0 .0 1, df = 7,16) 2 . 6572 (alpha = 0.05, df = 7, 1 6) Since F < Cr i t ical F FAIL TO REJEC T Ho: All equal (alpha Tit l e : 3303 fr on1 no . 7 day

0 . 05)

Fi l e : 3303f d Transform : NO TRANSFORMATION Dunnett's T e st TABLE 1 OF 2 H o : Control<Treatment TRANSFOR MED ME AN CALCULATED IN SIG GROUP IDENTIFICATION MEAN ORIGINAL UNI TS T STAT 0 . 05 -------------------- ----------- ------------------ 1 Neg. Control 273 . 3333 273 . 3333 2 solven:: control 259 . 6667 259 . 6667 0.7225 3 0 . 031 2 79 . 0000 279.0000 -0.29 96 4 0 . 064 2 70 . 3333 270.3333 0.1586 5 0 .1 3 24 7 . 0000 247.0000 1 . 3921 6 0 . 26 268.666 7 268 . 6667 0.2467 7 0 . 45 2 73 . 6667 273.6667 -0 .0176 8 0 . 94 248.0 000 248.0000 1 .3393 Dunnett critical value = 2 . 5 6 00 (1 T a il ed, alpha = 0.05, df = 7,16) Tit l e: 3303 frond no . 7 day File: 33 0 3fd Transform : NO T RANSFORMA TION Dunnett's Test. TABLE 2 OF 2 Ho:Contro l <Treatment -------------------·--- ------------ --- - --- ----- - -- -- - -- -- - --------- ------ --- NUM OF MIN SIG DIFF G ROU P IDENTIFICATIO N REPS (IN ORIG . UNIT S) ----------- --------- ------- ------- - - -- -- -- - 1 Neg. Control 3 2 so l ven:: contro l 3 48 .4 243 3 0.03 1 3 48 .4 243 4 0.064 3 48 . 4243 5 0. 13 3 48.4243 6 0 . 26 3 48.4243 7 0 . 45 3 48.4243 8 0 . 94 3 48 . 42 4 3 TEBUFENO.TXT : 3303 fro n d no. 7 day (V i a Nuthatch) W i lliams Test [One
- Sided Test for Decrease, alpha = 0 .0 50000 l Dose 0 0.03 1 0 . 06 4 0 . 13 0 .2 6 0 . 45

Isoton e Mea ns 271 271 27::> 263 263 263

T - bar P - value Significance - 0.258

- 0.2374 0.2098 0.2098 0 . 2098

N .S. N .S. N.S . N.S. N.S. Pa g e 14 o f 15

% OF DIFFE R E NCE CONTROL FROM CONTROL -- - -- -- - ---- --- --- - 1 7 . 7 1 3.6667 1 7 . 7 -5. 6667 1 7 . 7 3 . 0000 17 . 7 26 . 3333 17. 7 4 . 6667

17 .7 - 0 . 3333 17. 7 25 . 3333

### Data Evaluation Record on the Acute Toxicity ofTebufenozide to Aquatic Vascular Plants, 

## Lemnagibba EPA MRID Number 485330-03 0.94 248 1. 146 N.S.

"* " = Signifi cant; "N
. S."=No t Significan t .



### Page 15 of 15