NAA Summary of Analytical Chemisrry and Residue Data DP#: D423409 

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY 

WASHINGTON, D.C. 20460 

MEMORANDUM 

Date: November 18, 2015 

OFFICE OF CHEMICAL SAFETY 

AND POLLUTION PREVENTION 

SUBJECT: 1-Naphthaleneacetic acid (NAA), its salts, ester, and acetamide. New Use in/on 

Pomegranate. Summary of Analytical Chemistry and Residue Data. 

PC Code: 05600 1, 056002, 056003, 056004, 

056007, & 056008 

Decision No.: f./9511 1 

Petition No.: 4E8310 

Risk Assessment Type: NA 

TXR No.: NA 

DP Barcodc: 423409 

Registration Nos.: 548 1-429 

Regula tory Action: Section 3 

Case No.: 0379 

CAS No.: 86-87-3 

M RID Nos.: 4945100 I 40 CFR: 180. 155 

FROM: Thurston G. Morton, Chemist 

Risk Assessment Branch 4 

Health Effects Division (7509P) 

THROUGH: Elissa Reaves, Ph.D., Chief 

Risk Assessment Branch 4 

Health Effects Division (7509P) 

And 

Danette 

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~ Science Advisory Council (ChemSAC) 

Yvonne Barnes, Chemist r. ~ ~~ 

RAB6/HED, 7509P, ChemSAC 

TO: Sidney Jackson/Barbara Madden 

Risk Integration, Minor Use & Emergency Response Branch 

Registration Division (7505P) 

Page 1 of 11 
NAA Summary of Analytical Chemistry and Residue Data DP#: 0423409 

Action Requested 

Under PP#4E83 l 0, The Interregional Research Project Number 4 (IR-4) has submitted a petition 

for the establishment of a permanent tolerance for residues of the plant growth regulator 1-

Naphthaleneacetic acid (NAA) for a Section 3 registration of TRE-HOLD Sprout Inhibitor A 11 2. 

The petitioner is proposing the establishment of the following permanent tolerance for residues 

of NAA: 

Pomegranate ............................................................. ........ ........ 0.05 ppm 

Discussio11 

1-Naphthaleneacetic acid (NAA), its salts, ester, and acetamide are plant growth regulators 

(PGR) which are collectively referred to as naphthalene acetates. The PGR activity of NAA is 

due to its structural similarity to the natural plant hormone indole acetic acid (lAA). They are 

currently registered for use on various orchard and fruit crops to prevent preharvest drop of fruits, 

thin fruit trees, and delay flower induction. They can also stimulate growth and delay leaf drop on 

ornamentals. 

New residue chemistry data on pomegranate were submitted to support the new use. 

Residues ofNAA were below the LOQ (<0.05 ppm) in/on pomegranate harvested 45 days 

following two spray applications of a I% ai solution of the 15.1 % EC formulation of NAA ethyl 

ester. 

Tolerances are established under 40CFR§ 180.155(a) for the residues of l-naphthaleneacetic acid, 

including its metabolites and degradates in or on the commodities. Compliance with the tolerance 

levels specified is to be detennined by measuring only 1-naphthaleneacetic acid and its 

conjugates, calculated as the Stoichiometric equivalent of 1-naphthaleneacetic acid, in or on the 

commodity. The established tolerances range from 0.05 ppm to 2.0 ppm. 

The current tolerance expression for NAA is in accord with our current guidance. 

Conclusion 

HEO has examined the residue chemistry database for NAA. With regards to NAA for 

PP#4E83 10, pending submission of analytical standards, there are no residue chemistry issues 

that would preclude granting a Section 3 registration for residues of NAA only in/on the 

following commodity: 

Pomegranate ............................................ ................................. 0.05 ppm 

A human health risk assessment is forthcoming. 

Page 2of 11 
NAA Summary of Analytical Chemistry and Residue Data DP#: D423409 

Background 

The chemical structure and nomenclature of I naphthaleneacetic acid (NAA), its salts, ester, and 

acetamide and the physicochemical properties are presented in Tables I and 2. 

Table l. Naphtha lene Acetates Nomenclature 

Chemical structure 0 0 

Nll 2 OH 

I 

~ ~ 

I 

~ ~ 

~ ~ ~ ~ 

Common name AA acctamidc AA 

Molecular Formula C12H11 0 C12l !io02 

Molecular Weight 185.23 186.20 

IUPAC name 2-( 1-naphthyl)acctamide 2-( 1-naphthyl)acetic acid 

CAS name 1-naphthalcneacctamide 1-naphthalcneacetic acid 

CAS# 86-86-2 86-87-3 

PC Code 05600 1 056002 

Chemical structure 0 0 

• - + 

0 K 0 NH4 

I 

~ ~ 

I 

~ ~ 

~ ~ ~ ~ 

Common name NAA potassium salt NAA ammonium salt 

Molecular Formula C12H1002K C12H13 0 2 

Molecular Weight 224.3 1 203.24 

IUPAC name potassium-2( 1-naphthyl)acetatc ammonium-2( 1-naphthyl)acetate 

CAS name 1-naphthalnene acetic acid. potassium salt 1-naphthaleneacctic acid, ammonium salt 

CAS# 151 65-79-4 25545-89-5 

PC Code 056003 056004 

Page 3of11 
NAA Summary of Analytical Chemistry and Residue Data DP#: D423409 

Table I. Naphtha lene Acetates Nomenclature 

Chemical structure 

0 Na 

Common namt: NAA sodium salt NAA ethyl ester 

Molecular formula 

Molecular Weight 208.2 214.26 

IUPAC name sodium-2( 1-naphthyl )acetate ethyl-2( 1-naphthyl)acetatc 

CAS name 1-Naphthalcncacctic acid. sodium salt 1-Naphthalencacetic acid. ethyl ester 

CAS # 61-31-4 2122-70-5 

PC Code 056007 056008 

Table 2. Physicochemical Properties of NAA 1 

Parameter Value Reference 

Active Ingred ient NAA acetamide 

Melting point/range 182-184 °c Farm Chemicals 1 landbook 

pl I of I% aqueous suspension 5.1 Product CSF 

Density or specific gravity 0.221 g/cm3 Product CSF 

Water solubility (20 C) not available 

Solvent solubility (20 C) not available 

Vapor pressure at 20 C not available 

Dissociation constant (pK.) not available 

Octanol/water partition coefficient (Kow) not available 

UV/vis absorption spectrum not available 

Active ingredient NAA 

Melting point/range 130 °c Farm Chemicals 1 landbook 

pl I of I% aqueous suspension 3.45 RD 0265117. 5115100. B. Kitchens 

Density or specific gravity 0.45 g/mL CB Nos. 3468 and 3469. 6/3/88. F. uhre 

Water solubility (20 C) 0.042 g/100 mL CB Nos. 3468 and 3469. 6/3/88. F. Suhre 

Solvent solubility (20 C) xylene 5.5 g/ I 00 mL CB Nos. 3468 and 3469. 6/3/88. F. Suhre 

CCI~ 1.06 g/100 mL 

freely soluble in acetone. ether. Parm Chcmicals I landbook 

and chloroform 

Vapor pressure at 20 C 0.3 mm Hg at 26 °c CB Nos. 3468 and 3469. 6/3/88. F. Suhrc 

Dissociation constant (pKa) 3. 16 x 10-' CO Nos. 3970 and 3971. 7/5/88. F. Suhrc 

Octanol/water partition coefficient (Kow) not applicable: polar compound 

UV/vis absorption spectrum not available 

Active ingredient NAA sodium salt 

Melting point/range >300 °c I CB Nos. 3468 and 3469. 6/3/88. F. Suhre 

Page 4 of 11 
NAA Summary of Analytical Chemistry and Residue Data DP#: D423409 

Table 2. Physicochemical Properties of NAA 1 

Parameter Value Reference 

pl I of 1% aqueous suspension 9. 1 Cl3 Nos. 3468 and 3469. 6/3/88. F. Suhre 

Density or specific gravity 0.46 g/mL CB Nos. 3468 and 3469, 6/3/88, F. Suhre 

Water solubility (20 °q 340 g/100 mL CB Nos. 3468 and 3469, 6/3/88, F. Suhre 

Solvent solubility (20 °C) insoluble in nonpolar solvents CB Nos. 3468 and 3469. 613188, F. Suhre 

Vapor pressure at 20 °c not available 

Dissociation constant (pKa) 3.16 x 10..s CB os. 3970 and 397 1. 715188, F. Suhre 

Octanol/water partition coemcient (Kow) not applicable: polar compound 

UV/vis absorption spectrum not available 

/\ctive ingredient NAA ethyl ester 

Boiling point/range > 150 °c Old un-reviewed Union Carbide data 

pl I of I% aqueous suspension not available 

Density or specific gravity I. I I at 20 °C Old un-reviewed Union Carbide data 

Water solubility (20 °C) insoluble Old un-reviewcd Union Carbide data 

. oh ent solubility (20 °C) soluble in xylene. toluene. Old un-reviewed Union Carbide data 

ethanol. acetone. and methyl 

ethyl ketone 

Vapor pressure at 20 °C not available 

Dissociation constant (pK.) not available 

Octanol/water partition cocnicient (Kow) not available 

UV/vis absorption spectrum not available 

1 No physicochemical properties information was available concerning the NAA potassium and ammonium salts. 

860.1 200 Directions for Use 

The petitioner has submitted copies of the proposed labels for Tre-Hold Sprout Inhibitor A-112. 

Tre-Hold Sprout Inhibitor A-112 is an emulsi fiable concentrate formulation containing 15.1 % 

NAA, ethyl ester (I lb/gal ae NAA). The proposed use directions are presented in Table 3 below. 

Table 3. Summa ry of Directions fo r Use of NAA 

Applic. Timing, Max. No. Max. 

Fonnulation Applic. Rate Seasonal PHI Type, and (lb ac/gal) Applic. per Applic. Rate (days) Equip. Season (lb ae/gal) 

Pomegranate 

TRE-Hold Sprout Inhibitor A- 0.078 lb ac/gal two per I 12, EPA Reg. No. 548 1-429; 10 oz. 45 days 

15.1 % NAA, ethyl ester ( I lb product/gallon year 0.156 

Directed spray implied to the lower ae/gal) ( !% w/v) 

tree trunks 

Use Directions and Limitations: Treat early before suckers attain 12" in length. Best results are 

attained when first treatment is made after the trees resume growth and suckers become visible. 

A minimum 60-day RTI is specified. The REI is 12 hours. Aerial application is prohibited. 

Conclusion. The proposed use directions are adequate to allow evaluation of the residue data 

relative to the proposed uses. 

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NAA Summary of Analytical Chemistry and Residue Data DP#: D423409 

860.1300 Nature of the Residue - Plants 

RED Residue Chemistry Chapter D299296, 10 MARCH 2004. G. Otakie 

Acceptable apple and olive metabolism studies have been previously submitted and reviewed. 

Based on the nature of residues identified from the apple and olive metabolism studies, HED has 

previously determined that the terminal residues of concern in plants are the parent compounds, 

NAA and its conjugates and that the qualitative nature of the residues in plants (i.e., orchard 

trees) is adequately understood. For purposes of the current petition the nature of the residue in 

plants is adequately understood. 

860.1300 Nature of the Residue - Livestock 

RED Residue Chemisrry Chapter D299296, 10 MARCH 2004, G. Otakie 

An acceptable goat metabolism study has been previously submitted and reviewed. Based on 

these data, HED has previously concluded that the qualitative nature of the residue in ruminants 

is adequately understood. The terminal residues of concern in ruminants are the parent 

compounds, NAA and its conjugates. 

The available ruminant metabolism data are adequate to support the proposed uses. There are no 

poultry feed items associated with currently registered or proposed food uses ofNAA; thus a 

poultry metabolism study is not required. 

860.1340 Residue Analytical Methods 

RED Residue Chemistry Chapter D299296, I 0 MARCH 2004, G. Otakie 

Enforcement and data collection methods: The residue analytical methods, currently listed in 

Volume II of the Pesticide Analytical Methods, are inadequate for the enforcement of tolerances 

for residues of 1-naphthaleneacetic ac id (Pesticide Reg. Sec. 180.1 55) and 1-

naphthaleneacetamide (Pesticide Reg. Sec. 180.309) in/on plant commodities because these 

methods use benzene, a hazardous or toxic reagent. 

An HPLC method using fluorescence detection (Method NAA-AM-001) for apples and peas and 

a similar method for olives and olive oil (Method NAA-AM-002) has been previously submitted 

for determination of NAA in plant commodities. These methods include extraction with water 

and incorporate a basic hydrolysis step to release bound residues. These methods do not use 

benzene or diazomethane whjch are being discouraged by the Agency for safety reasons. Theses 

methods have been subjected to successful independent laboratory validations. Acceptable 

recoveries were obtained from apples, olives and olive oil fortified with NAA at the method 

LOQ (0.0 l ppm) and at 1.0 ppm. This method is suitable for enforcement. 

Pomegranate samples were analyzed for residues of NAA using high performance liquid 

Page 6of11 
NAA Summary of Analytical Chemistry and Residue Data DP#: D423409 

chromatography with fluorescence detection (HPLC/UV) adapted from Morse Laboratories, Inc. 

analytical method NAA-AM-003. The method is a common moiety method that converts 

residues ofNAA acetamide and NAA ethyl ester to NAA. The limit of quantitation (LOQ; 

determined as the lowest level of method validation, LLMV) was 0.05 ppm. Acceptable method 

validation and concurrent recoveries [Concurrent recoveries were below the acceptable range of 

70% ; 61 % for NAA, and 60% for NAA-EE.] were reported for samples of pomegranate 

fortified with NAA and NAA ethyl ester at 0.05-1.0 ppm, thus validating the method. The 

fortification levels used in concurrent method recovery were adequate to bracket expected 

residue levels. Concurrent recoveries and residues in the treated samples were not corrected for 

residues in controls. 

860.1360 Multiresidue Methods 

0232463, 0294854, 30 OCT 2003, G. Otakie 

Multiresidue method (MRM) data have been submitted previously in support of the registration 

ofNAA, its salts, ester and acetamide. Naphthaleneacetamide was found to chromatograph 

satisfactori ly using Pesticide Analytical Manual (PAM) Vol. I, Section 302 DG 5. 

Naphthaleneacetamide was found to produce a 50% fu ll scale deflection (FSD) with a 50 

nanogram injection and the relative retention time to chlorpyrifos is approximately 0.8. Average 

recoveries for naphthaleneacetamide spiked into pears at approximately 0.1 ppm and 1.0 ppm 

were 97.8 and I 04%, respectively, for Section 302, method EI, without Flori sit cleanup. Apple 

spikes using this procedure could not be quantified due to interference in the apple matrix. 

Naphthaleneacetamide was found not to be recoverable through Florisil, so it would not be 

recoverable through Protocols E or F. Therefore, testing under PAM Vol. I Protocol E, for 

apples and pears, and Protocol F, for olives, was suspended prior to recovery experiments 

through the entire Sections 303 and 304 methods. FDA MRM's are not suitable as enforcement 

methods for NAA. 

860.1380 Storage Stability 

RED Residue Chemistry Chapter 0299296, 10 MARCH 2004, G. Orakie 

Adequate storage stability data have been submitted and reviewed for apples, pears, and olives. 

These data indicate that residues of 1-naphthaleneacetic acid are reasonably stable under frozen 

storage conditions in/on the above commodities for up to 12 months. In addition, storage 

stability data showed NAA and NAA ethyl ester stored in frozen avocado were stable for up to 

27 months and stored in frozen Rambuttan for up to 25 months. 

All pomegranate samples for the current submission were maintained frozen at the testing 

facilities, during shipping, and were stored frozen at the analytical laboratory until analysis. The 

maximum storage interval for samples between harvest and extraction was 23 1 days (7.6 

months). Samples were analyzed within 5 days of extraction. Concurrent recoveries were below 

the acceptable range, 6 1 % for NAA (see MRID# 4945100 I, p.154), and 60% for NAA-EE 

(p.149). Storage stability data generated concurrently with the field trial study demonstrate that 

residues of NAA and NAA ethyl ester, respectively, are stable in/on samples of pomegranate 

Page 7of 11 
NAA Summary of Analytical Chemistry and Residue Data OP#: 0423409 

stored frozen for up to 240 and 248 days (7.9-8.2 months); no 0-day data were provided. These 

data are acceptable to support the storage conditions and durations of sampl es from the submitted 

field trials; however, storage stability studies should always include a 0-day sampling interval to 

establish the residue levels present at the time samples are placed into storage [see OPPTS 

860.1380( d)(6)(i)]. 

860.1400 Water, Fish, and Irrigated Crops 

There are no proposed uses that are relevant to this guideline topic. 

860.1460 Food Handling 

There are no proposed uses that are relevant to this guideli ne topic. 

860.1480 Meat, Milk, Poultry, and Eggs 

RED Residue Chemistry Chapter 0 299296, I 0 MARCH 2004, G. Otakie 

Livestock 

There are no livestock feedstuffs associated with the proposed uses of naphthalene acetates. At 

this time, the Agency has determined that NAA is in Category 6(a)(3) (i.e., no reasonable 

expectations of finite residues of concern in meat and milk). 

Poultry 

There are no poultry feed items associated with the currently registered or proposed uses of 

naphthalene acetates; thus, a poultry feeding study is not required. Tolerances for the eggs and 

edible tissues of poultry need not be established. 

860.1500 Crop Field Trials 

4945100 t .der 

In pomegranate harvested 45 days following the last of two applications of a 15. l % EC 

formulation ofNAA ethyl ester directed to the base of the tree as a 1% w/v ai spray to run-off to 

pruned sprouts/suckers, residues of NAA were below the LOQ (<0.05 ppm) in/on all samples. 

Residue decline was not investigated. 

Page 8 of 11 
NAA Summary of Analytical Chemisrry and Residue Data DP#: 0 423409 

TABLE4. Residue Data from Pomegranate f ie ld Trials with NAA. 

Location: City. State: Zone Crop Matrix Total Rate Pl II (days) Residues2 (ppm) 

Year (Trial ID)1 Variety (% wlv ai [average] 

solution)' 

Lost ll ills. CA: 2011 10 Wonderful Fruit 2X 1% 45 <0.05. <0.05, <0.05. <0.05 

(CA63 and CA64) ro.051 

Gridley. CA: 2011 10 Wonderful Fruit 2 X 1% 45 <0.05, <0.05. <0.05. <0.05 

(CJ\65 and CA66) ro.051 

1 Two applications of a I% solution were made at each trial site. 2 Residues ofNAA ethyl ester were determined by a common moiety method that convens residues ofNAA acetamide and NAA 

eth) I ester to AA. The LOQ was 0.05 ppm. Values below the LOQ were not provided in the raw data. 

Table 5. Summary of Residues from Pomegranate f ield Trials with NAA ethyl ester. 

Commodity Total Rate1 Analyte Pl II , n· 

(% wlv ai (days) 

solution) 

Pomegranate 2 x 1% NAA~ 45 2 

1 Two appl ications of a I% solution were made at each trial site. 

~ n = no. of independent field trials. 3 Values based on total number of samples. 

Residue Levels (ppm) 

Min.

3 Max.3 Ll\FT.i HAFT-I Median4 

<0.05 <0.05 0.05 0.05 0.05 

Mean4 SD 

0.05 NII\ 

4 Values based on per-trial averages. LAFT= lowest average field trip. HAFT = highest average field trial. SD = standard 

deviation. For computation of the Ll\FT. I IAFT. median. mean. and standard deviation. values <LOQ arc assumed to be at the 

LOQ. NIA= Not applicable. 5 Residues ofNAA ethyl ester were determined by a common moiety method that converts residues of NA/\ acctamide and NAA 

ethyl ester to NAA. 

Conclusions 

The submitted pomegranate field trial data are adequate. The geographical representation of field 

trials are appropriate. In pomegranate harvested 45 days following the last of two applications of 

a 15.1 % EC formulation of NAA ethyl ester directed to the base of the tree as a I% w/v ai spray 

to run-off (0.078 lb ae/gal x 2 applications) to pruned sprouts/suckers, residues ofNAA were 

below the LOQ (<0.05 ppm) in/on all samples. Residue decline was not investigated. 

An acceptable method was used for residue quantitation, and adequate storage stability data were 

submitted to support sample storage durations and conditions for all analytes. 

860.1 520 Processed Food and Feed 

There are no processed commodities associated with the proposed uses in the current petition 

860.1650 Submittal of Analytical Reference Standards 

A supply of NAA analytical standard is not available in the National Pesticides Standards 

Repository. Analytical standards must be supplied. 

To date, the Repository has not received a supply ofNAA reference standard. As NAA is the 

key residue for enforcement, the petitioner is required to submit a 5-gram sample of reference 

Page 9of11 
NAA Summary of Analytical Chemistry and Residue Data DP#: 0423409 

standard and supporting documentation (see OCSPP Guideline 860.1650) to the repository at the 

address below, to the attention of either Theresa Cole or Thuy Nguyen. Note that the full 9-digit 

ZIP code is mandatory or the mail will be returned to the sender. 

USEPA 

National Pesticide Standards Repository/Analytical Chemistry Branch/OPP 

701 Mapes Road 

Fort George G. Meade, MD 20755-5350 

As a reminder to the petitioner, supplies of analytical standards must be replenished as requested 

by the repository as long as the tolerance is published in 40 CFR. 

860.1850/ 860.1900 Confined/Field Accumulation in Rotational Crops 

There are no rotational crop data requirements associated with the proposed uses in the current 

petition. 

860.1550 Proposed Tolerances 

Tolerances are established under 40CFR§ l 80. 155(a) for the combined residues of 1-

naphthaleneacetic acid and its conjugates calculated as 1-naphthaleneacetic acid from the 

application of 1-naphthaleneacetic acid, its ammonium, sodium, or potassium salts, ethyl ester, 

and acetamide. The established tolerances range from 0.05 ppm to 2.0 ppm. 

No Codex or Canadian maximum residue limits (MRLs) have been established for residues of 

(NAA), its salts, ester, and acetamide for the use in this petition. 

Table 6 Tolerance Summary for NAA 

Commodity I Established/Proposed I 

Tolerance (ppm) 

Recommended I CommenlS; Correct Co111111odity 

Tolerance (ppm) Definition 

Tolerances Proposed Under PP#4E8310 

Pomegranate I 0.05 I 0.05 I 

Page 10of 11 
NAA Summary of Analytical Chemistry and Residue Data DP#: 0423409 

Attachment l: International Residue Limits 

NAA (056002;) 

Summary of US and International Tolerances and Maximum Residue Limits 

Residue Definition: 

us Canada Mexico2 Codex 

40 CFR 180.155: None None 

Plant: tolerance levels specified is to be 

determined by measuring only 1-

naphthaleneacetic acid and its conjugates, 

calculated as the Stoichiometric equivalent of I -

naphthaleneacetic acid 

Livestock: NA 

Tolerance (ppm) IMrLi:imum Residue limit (mg/kg) 

Commodity' 

Mexico2 us Canada Codex 

Pomegranate 0.05 

Completed: M. Negussie/ 05/05/ 15 

1 Includes only commodities of interest for this action. Tolerance values should be the HED recommendations and 

not those proposed by the applicant. 

2 Mexico adopts US tolerances and/or Codex MRLs for its export purposes. 

F. DOCUMENT TRACKING 

RDI: T. Morton 11 /1 8/15; E. Reaves 11 /1 8/15 

Petition Number: 4E8310 

DP#: 423409 

PC Code: 129032 

Tcmpla1c Version Scplcmbcr 2005 

Page 11 of 11 
NAA [PC Code 056008]/lnterrcgional Research Project No. 4 [IR-4] 

B.7.6 Residues Resulting from Supervised Trials 

(Annex HA 6.3; Annex IIIA 8.3) 

B.7.6.1 Residues in Target Crops 

B.7.6.1. 1 Pomegranate 

Document ID: MRID No. 4945 100 I 

PMRA No. Not Available 

Report: Barney, W. (2011) NAA: Magnitude of the Residue on Pomegranate. 

Project Number: A5389. Unpublished study prepared by Interregional 

Research Project No. 4. 194 p. 

Guidelines: EPA OCSPP Hannonized Test Guideline 860.1 500 Crop Field Trials 

(August 1996) 

PMRA Regulatory Directive DIR98-02 - Residue Chemistry Guidelines, 

Section 9 - Crop Field Trials 

PMRA Regulatory Directive DIR20 I 0-05 - Revisions to the Residue 

Chemistry Crop Field Trial Requirements 

OECD Guideline 509 Crop Field Trial (September 2009) 

GLP Compliance: No deviations from regulatory requirements were reported which would 

have an impact on the validity of the study: 

Acceptability: The study is considered scientifically acceptable. The acceptability of this 

study for regulatory purposes is addressed in the forthcoming U.S. EPA 

Residue Chemistry Summary Document, DP# 423409. 

Evaluator: Thurston G. Morton, Chemist 

Registration Action Branch 4 

Health Effects Division (HED) (7509P) 

Note: This Data Evaluation Record (DER) was originally prepared under contract by Versar. Inc. (6850 Versar Center, 

Springfield. VJ\ 22 151 ; submitted 2/2/15). The DER has been reviewed by 1-1 ED and revised to reflect current Office of 

Pesticide Programs (OPP) policies. 

EXECUTIVE SUMMARY 

The Interregional Research Proj ect Number 4 (IR-4) has submitted field trial data for 1-

naphthaleneacetic acid (NAA) on pomegranate. Two field tri als were conducted in the United 

States during the 20 11 growing season in the North American Free Trade Agreement (NAFT A) 

Growing Zone 10 (CA). The above trial count reflects two pairs of trials conducted at the same 

locations that HED has detcnnined to be replicate trials. 

Each trial site consisted of one untreated plot and one treated plot. At each trial location, the 

treated plot received two applications of a 15. I% emulsi fiable concentrate (EC) fomrnlation of 

NAA ethyl ester (Tre-Hold Sprout Inhibitor A-112) directed to the base of the tree as a I% w/v 

ai spray to run-off to pruned sprouts/suckers. The reported % deviation from the target rate was 

S0.09. Applications were made at retreatment intervals of 60 days using ground equipment in 

EPA MRID #: 4945 1001 

Page I of7 
NAA [PC Code 056008)/lnterregional Research Project No. 4 [IR-41 

spray volumes of 13-23 gal/A. No adjuvants were used. Samples of pomegranates were 

harvested at a preharvest interval (PHI) of 45 days. 

All samples were maintained frozen at the testing facilities, during shipping, and were stored 

frozen at the analytical laboratory until analysis. The maximum storage interval for samples 

between harvest and extraction was 23 l days (7.6 months). Samples were analyzed within 5 

days of extraction. Storage stability data generated concurrently with the field trial study 

demonstrate that residues of NAA and NAA ethyl ester, respectively, are stable in/on samples of 

pomegranate stored frozen for up to 240 and 248 days (7.9-8.2 months); no 0-day data were 

provided. These data are acceptable to support the storage conditions and durations of samples 

from the submitted field trials; however, storage stability studies should always include a 0-day 

sampling interval to establish the residue leve ls present at the time samples are placed into 

storage [see OPPTS 860. I 380(d)(6)(i)]. 

Samples were analyzed for residues of NAA using high performance liquid chromatography with 

fluorescence detection (HPLC/UV) adapted from Morse Laboratories, Inc. analytical method 

NAA-AM-003. The method is a common moiety method that converts residues ofNAA 

acetamide and NAA ethyl ester to NAA. The limit of quantitation (LOQ; determined as the 

lowest level of method validation, LLMV) was 0.05 ppm. Acceptable method validation and 

concurrent recoveries were reported for samples of pomegranate fortified with NAA and NAA 

ethyl ester at 0.05-1.0 ppm, thus validating the method. The fortification levels used in 

concurrent method recovery were adequate to bracket expected residue levels. Concurrent 

recoveries and residues in the treated samples were not corrected for residues in controls. 

In pomegranate harvested 45 days following the last of two applications of a 15.1 % EC 

formulation of NAA ethyl ester directed to the base of the tree as a 1 % w/v ai spray to run-off to 

pruned sprouts/suckers, residues of NAA were below the LOQ (<0.05 ppm) in/on all samples. 

Residue decline was not investigated. 

I. MATERIALS AND METHODS 

A. MATERIALS 

Table B.7.6. 1.1 -1. Nomenclature for NAA. 

Common name NAA 

Identity 1-naphthalcneacctic acid 

CAS no. 86-87-3 

Company experimental name NIA 

Other synonyms (if applicable) NIA 

0 

OH 

I ~ ~ 

~ ~ 

EPA MRIO #: 4945 1001 

Page 2 of7 
NAA [PC Code 056008]/Interregional Research Project No. 4 [IR-4] 

Table B.7.6.1.1- 1. Nome nc la ture fo r NAA. 

Common name NJ\A ethyl ester 

Identity 1-Naplnhalcncacetic acid. ethyl ester 

CAS no. 2122-70-5 

Compa ny exper imental name NIJ\ 

Other synonyms (if a pplicable) NIA 

0 

O~CH 3 

I ~ ~ 

~ ~ 

Common name NJ\J\ acctamidc 

Identity 1-naphthaleneacetamidc 

CAS no. 86-86-2 

Compa ny experi mental name NIA 

Other synonyms (if applicable) NIA 

0 

NH2 

I 

~ ~ 

~ ~ 

B. Study Design 

1. Test Procedure 

Two field trials were conducted on pomegranate during the 201 1 growing season reflecting two 

applications of a 15.1 % EC formulation ofNAA ethyl ester (Tre-Hold Sprout Inhibitor A-11 2) 

directed to the base of the tree as a I% w/v ai spray to run-off to pruned sprouts/suckers. Field 

trial locations by NAFTA growing zone are summarized in Table B.7.6.1.1-2. 

The trials separated by <20 miles have been assessed for independence as deta iled in the table 

below. HED has determined that each pair of trials constitutes a single trial with replicate 

samples for purposes of 860.1500 data requirements. 

Independent Trial Determination1 

Trial Nos. Differences Decision 

Cl\63 & CJ\64 Same location and varietv Replicate 

*No other differences in criteria 

CA65 & CA66 Same location and varietv Replicate 

'No other differences in criteria 

1 J\11 assessments are based on the replicate trial guidance presented in draft memo 568_ Critcria for Independence of Trials 

04/23/2013 (EPA and PMRA). 

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NAA [PC Code 056008)/lnterregional Research Project No. 4 [IR-4] 

Table B.7.6.1.1-2. Trial Numbers a nd Geographical Locations. 

Crop No. NAfTA Growing Zone Total Trials 1 2 3 4 5 6 7 8 9 10 11 12 

Pomegranate Sub. 22 2 

Req. 1 3 or 2 1 As per Table I of 860.1500 for pomegranate. three trials with a minimum of 6 samples or two trials with a minimum of 8 

samples arc required. Geographic distribution is not specified for crops requiring 3 trials: ho\\ ever. per Table 6 of 860.1500. 

Zone I 0 accounts for 99% of U.S. pomegranate production. 2 I JED has concluded that two pairs of the four Zone 10 trials constitute single trials with replicate samples for purposes of 

860.1500 data requirements. 

Locations and detailed use patterns for the trials are provided in Table B.7.6.1.1-3. 

Table B.7.6.1.1-3. Study Use Pattern. 

Location: End-use Method of Application/ Timing of Volume Rate per Retrcatment Total Surfactant/ 

City, State: Product' Application (gal/A) Application Interval Ratc2 Adjuvant 

Year (% ai (days) (% 

(Trial ID) solution) Solution) 

Lost Hills. 15.1% I. Directed to base of tree: blooming/ 13 I - 2x 1% None 

CA: 20 11 EC fruiting 

(CA63) 2. Directed to base of tree: blooming 14 I 60 

/fruiting 

Lost I lills. 15.1% I. Directed to base of tree: blooming/ 16 I -- 2x 1% one 

CA: 2011 EC fruiting 

(CJ\64) 2. Directed to base of tree: blooming/ 15 I 60 

fruiting 

Gridley. 15.1% I. Directed to base of tree: flowering/ 21 I -- 2x 1% None 

CJ\:20 11 EC bloom 

(C/\65) 2. Directed to base of tree: fruiting 22 1 60 

Gridley. 15.1% I. Directed to base of tree: flowering 23 1 -- 2x 1% None 

CJ\: 2011 EC 2. Directed to base of tree: fruiting 22 I 60 

(CJ\66) 

1 15.1 % emulsiliablc concentrate ( ) EC fonnulation of NA/\ eth ) •I ester Tre-Hold S p rout Inhibitor A-11 2 2 Two applications of the I% solution were made at each trial site. 

Pomegranates were grown and maintained according to typical agricultural practices. Irrigation 

was used to supplement rainfall at all trial sites. No unusual weather conditions were reported to 

have adversely affected crop growth or yields during the study. 

Sample Handling and Preparation 

Single control and duplicate treated samples of pomegranate were harvested following 

application at a 45-day PHI. Samples were placed in frozen storage at the field sites within - 3.5 

hours of harvest and were shipped within 5-14 days of harvest frozen on dry ice via FedEx to the 

analytical laboratory, IR-4 Western Region Satellite Laboratory (Honolulu, HI). At the 

analytical laboratory, samples were homogenized in the presence of dry ice and were stored 

frozen (- -20 °C) until extraction for analysis. 

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NAA [PC Code 056008]/lntcrrcgional Research Project No. 4 [IR-4) 

2. Description of Analytical Procedures 

Samples were analyzed using an HPLC/UY method adapted from Morse Laboratories, Inc. 

Analytical Method NAA-AM-003 entitled ··Recommended Analytical Residue Method for 1-

Naphthal eneacetic Acid, 1-Naphthaleneacetamide, and/or 1-Naphthaleneacetic Acid Ethyl Ester, 

in Grape (red and white), Grapefruit, Tomato, and Green Bell Pepper Matrices". The method is 

a common moiety method that converts residues ofNAA acctamide and NAA ethyl ester to 

NAA. A complete description of the method including a list of minor modifications was 

included in the submission. 

Briefly, samples were extracted by blending with distilled water. Residues ofNAA ethyl ester 

were converted to NAA by base hydrolysis with 50% NaOH at reflux for 3 hours. The mixture 

was cooled and filtered, diluted with water, then partitioned with dichloromethane (DCM). The 

resulting aqueous phase was acidified to pH <2 with concentrated HCI and partitioned twice with 

DCM. The resulting organic phases were dried through sodium sulfate, reduced by rotary 

evaporation, and adjusted to volume with DCM, then applied to a silica gel solid phase 

extraction (SPE) column. Residues were eluted with DCM: I% acetic acid (99: I, v:v), and the 

eluate was subj ected to solvent exchange with acetonitrile:water (1: 1, v:v). The resulting extract 

was filtered for analysis by HPLC/UV. 

The LOQ (determined as the LLMV) was 0.05 ppm. We note that the petitioner also presented 

calculated values for the LOO and LOQ based on the recoveries obtained from samples fortified 

with NAA and NAA ethyl ester at the 0.05-ppm fortification level. The LOO for each matrix 

was calculated by multiplying the standard deviation of recovery measurements al the LLMV by 

the one-tailed t-statistic (confidence level not reported) for six replicates, and the LOQ was 

defined as 3x the LOO. The calculated LOQs were 0.025 1 and 0.0285 ppm, respectively, for 

NAA and NAA ethyl ester; corresponding calculated LOD values were 0.00835 and 0.00951 

ppm. 

II. RESULTS AND DISCUSSION 

Method performance was evaluated during method validation and by use of concurrent recovery 

samples. Pomegranate samples were fortified with NAA and NAA ethyl ester at 0.050-1.0 ppm 

for method validation and 0. 10 ppm for concurrent recoveries. Recoveries were within the 

acceptable range of 70-120%; therefore, the method was considered valid for the analysis of 

NAA inion pomegranate (Table B.7.6.1.1 -4). The fortification levels bracketed the measured 

residues. Concurrent recoveries were not co1Tected for apparent residues in controls. 

The detector response was linear (coefficient of determination, r2 =::0.9997) within the range of 5-

50 ng/mL. Representative chromatograms of control samples, fortified samples, and treated 

samples were provided. The control chromatograms generally had no peaks of interest above the 

chromatographic background. The fortified sample chromatograms contained onl y the analyte of 

interest, and peaks were symmetrical and wet I defined. Apparent residues in controls were 

below the LOQ (:S0.05 ppm). The reported residue values were not corrected for apparent 

residues in controls. 

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NAA [PC Code 056008)/lnterregional Research Project No. 4 [IR-41 

T a b le 8 .7.6.1.1-4. S umma r y of M ethod Valida tio n a nd Concu r rent Recoveries of NAA a nd NAA ethyl ester 

fro m Po megra na te. 

Matrix I Analyte I Fortification I Sample size I 

Level (ppm) (n) 

Method Validation 

I NA/\ I 0.050-1.0 I 12 I Pomegranate I NAA ethyl ester I 0.050-1.0 I 12 I 

Concurrent Recovery 

) I NAA I 0. 10 I I I 

I omegranate I N I 0. 10 I I AA I ethyl ester 

1 Recoveries were not corrected for apparent residues in controls. 2 Standard deviation is not calculated for sample sizes <3. 

Recoveries' I Mean ± Std. Dev. 2 

(%) (%) 

74-87 I 79 ± 4.6 

73-90 I 83 ± 5.6 

6 1 I 6 1 

60 I 60 

Samples were stored frozen for a maximum of 231 days (7.6 months) from harvest to extraction 

(Table B.7.6. I. l-5a). Samples were analyzed within 5 days of extraction. To support sample 

storage durations, a concurrent storage stability study was conducted with the field trial study; 

the results are presented in Table B.7.6. l.l-5b. The data demonstrate that residues ofNAA and 

NAA ethyl ester, respectively, are stabl e in/on samples of pomegranate stored frozen for up to 

240 and 248 days (7.9-8.2 months); no 0-day data were provided. These data are acceptable to 

support the storage conditions and durations of samples from the submitted field trials; however, 

storage stability studies should always include a 0-day sampling interval to establish the residue 

levels present at the time samples are placed into storage [see OCSPP 860.1380(d)(6)(i)). 

Table 8.7.6. 1.1-Sa. Summary of Sto rage Conditions. 

Matrix Analytc Storage Actual Storage Interval of Demonstrated Storage Stability 

(RAC or Extract) Temperature Duration1 

(OC) (days/months) 

Pomegranate Nl\A - -20 224-23 1 days Residues of NA/\ and NA/\ ethyl ester arc 

(7.4-7.6 months) stable inion samples of pomegranate stored 

AA ethyl ester frozen for 240 days (7.9 months) and 248 

days (8.2 months). respectively. 2 

1 Interval from harvest lo extraction. Samples were analy..:cd within 3-5 da) s of extrac11on. 2 Based on concurrent storage stability study. See Table B.7.6. 1. 1-5b 

T able B. 7.6. 1.1-Sb. Sta bility of NAA a nd NAA e thyl ester in Fr ozen Po megra na te. 

Analytc Spike Storage Fresh Fortification Stored Sample Mean Corrected % 

Level interval Recoveries (%) Recoveries (%) Recovery (%) recovery1 

(ppm) (days) 

NA/\ 0. 10 240 79 73. 73. 74 73 92.8 

NAA ethyl ester 0. 10 248 8 1 81.76.74 77 95. 1 

1 Corrected for recovery in freshly fortified samples. 

The results from the submitted field trials are presented in Tables B.7.6.1. 1-6 and B.7.6.1.1 -7. In 

pomegranate harvested 45 days following the last of two applications of a 15.1 % EC formulation 

ofNAA ethyl ester directed to the base of the tree as a 1% w/v ai spray to run-off to pruned 

sprouts/suckers, residues ofNAA were below the LOQ (<0.05 ppm) inion all samples. Residue 

decline was not investigated. 

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NAA [PC Code 056008]/lnterregional Research Project No. 4 [IR-4] 

TABLE 8 .7.6.1.1-6. Resid ue Data fro m Pom egranate Field Tria ls wit h NAA. 

Location: City. State: Zone Crop Matrix Total Rate Pl II (days) Rcsiducs2 (ppm) 

Year (Trial 10)1 Variety (% Solution) 1 laverage] 

Lost I lills. CA: 201 I 10 Wonderful Fruit 2X 1% 45 <0.05. <0.05. <0.05. <0.05 

(CA63 and CA6 ) [<0.05) 

Gridley. CA: 201 I 10 Wonderful Fru it 2X 1% 45 <0.05. <0.05. <0.05. <0.05 

(CA65 and CA66) f <0.05) 

1 Two applications of a I% solution were made at each trial site. 2 Residues ofNAA ethyl ester were determined by a common moiety method that converts residues ofNAA aeetamide and NAA 

eth)I ester to AA. TI1c LOQ was 0.05 ppm. Values below the LOQ were not provided in the raw data. 

Table 8 .7.6.1.1 -7. S ummary of Residues fro m Pomegranate Fie ld T r ials with NAA ethyl ester. 

Commodity Tota l Rate 1 Analyte PHI n2 

(%Solution) (days) 

Pomegranate 2X 1% AA5 45 2 

1 Two r1pplications of a I% solution were made at each trial site. 2 n = no. of lield trials. 1 Values based on total number of samples. 

Residue Levels (ppm) 

Min.

1 Max.3 LAFT'' HAf1'4 Median• Mean4 SD 

<0.05 <0.05 <0.05 <0.05 0.05 0.05 NIA 

4 Values based on per-trial averages. LAFT= lowest average licld trip. I IA FT = highest average lield trial. SD = standard 

deviation. For computation of the LAFr. HAFT. median. mean. and standard deviation. values <LOQ arc assumed to be at the 

LOQ. NIA= Not applicable. 5 Residues ofNAA ethyl ester were determined by a common moi.:ty method that converts residues ofNAA acetamide and NAA 

ethyl ester to NAA. 

III. CONCLUSIONS 

The pomegranate field trials are considered scientifically acceptable. In pomegranate harvested 

45 days following the last of two applications of a 15.1 % EC formulation of NAA ethyl ester 

directed to the base of the tree as a I% w/v ai spray to run-off to pruned sprouts/suckers, residues 

of NAA were below the LOQ (<0.05 ppm) in/on all samples. Residue decline was not 

investigated. 

An acceptable method was used for residue quantitation, and adequate storage stability data were 

submitted to support sample storage durations and conditions for all analytes. 

IV. REFERENCES 

None. 

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