OFFICE OF PREVENTION, PESTICIDES, AND TOXIC SUBSTANCES UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTO N, D.C. 20460

MEMORANDUM TXR No. 0051358 Date: 21-November-2002 Subject: Alpha Olefin Sulfonate Sodium: (PC Code 000905). Section 3 Registration for

Application of EH-2001 for the Control of Richardson’s Ground Squirrels and Wyoming Ground Squirrel. Human Health Risk Assessment.DP Barcodes: D275422, D275424, D282331, and D2865 16. Submission: S598325, S598325, S608203, a nd S608203. Case Number: 070054. From: PV Shah, Ph.D., Toxico logist Guruva Reddy

, D.V.M.; Ph.D., Toxic

ologist Mark Dow, Ph.D., B

iologist Registration Ac

tion Branch 1; He

alth Effects Divi

sion (RAB1/HED; 7509C) Through: G. Jeffrey H

erndon, Branc

h Senior Scientist RAB1/HED (7509C) To: Meredith Law s/Geraldine McCa

nn; PM Team 4 Registration Div

ision (7505C) And Kathryn Boyle Lower Toxicity Pesticide Chemical Focus Group Registration Div

ision (7505C)

ACTION REQUESTED: The Health Effe cts Division (HED)

of the Office of

Pesticide Progra

ms (OPP) is charged with estimating the risk to human health from exposure to pesticides. The Registration Division (RD) of OPP has requested that HED evaluate toxicology and worker e xposureas part of a joint review process between PMRA (Canada) and US EPA for the end-use product EH2001 (product con

taining ye llow mustard seed [PC C ode 014901] and sulf

onic acids, C14-16

-olefin sulfonate [PC Code 000905]) for the control of Richardson’s Ground Squirrels and Wyoming Ground Squirrel in rangeland, ornamental plantings, orchards, golf courses, parks, nurse ries, and non-crop rights-of-way.2

BACKGROUND: The Registration Action Branch 1 (RAB1) of the HED reviewed the data provide d in the MRID 45551102, 45551103, 45307616, and 45551101. In addition, RAB1 reviewed the available toxicological studies on alpha olefin sulfate (C14-16) from the open literature. A decision document on alpha olefin sulfate (C14-16) summarizing all available toxicity studies, has been submitted to Kathryn Boyle (L ower Toxicity Pesticide Chemical Focus Group) by Dr . Guruva Reddy of RAB1. This document has been reviewed by RAB1 toxicologists and Karen Whitby , Branch Chief of RAB1. This document is also included in this memo in its entirety. USE PATTERN - EH-2001 The product is dilut

ed in a ratio of 4 L

concentrate to 9 1 L water. I t is delivered via s

ome modification of a hand-held spray wand with an aspirating nozzle to facilitate foaming action. The nozzle is inserte

d as far as possibl

e into a burrow an

d the burrow is fill

ed with foam. The dilute is applied at 20-55 psi and it takes about 5 - 60 seconds (average 15 seconds) to treat one burrow. Approximately 3.4 liters of foam are administered per burrow or about 0.9 g al. The number of burrows treated per day is highly variable depending upon population density but is has been estimated that 215 gallons of dilute could be used in an 8 hour work day. Upon administration of the foam, a wire mesh basket is placed over the burrow opening. If no activity is observed after about one minute, the bur

row entrance is

to be filled with ea

rth and tamped fir

mly. Use of th

is product is limited to areas in Canada of southern Alberta, Saskatchewan and the extreme Southwest of Manitoba and in the U.S., of I

daho, Montana, No

rth Dakota, Wy

oming, South Dakot

a, Colorado, Minne

sota and Nevada. CONCLUSION: Based on available toxicity data and anticipated use pattern, RAB1 of the HED concluded that a

quantitative risk a

ssessment is not requiredfor the end-use product EH-2001 for the control of Richardson’s ground squirrel and Wyoming ground squirre l. This conclusion is based on the follow ing:

• Based on the proposed use pattern and other information provided to HED for review, only short term exposure via inhalation or dermal routes of exposure are anticipated. Exposure by the oral route is not anticipated. HED expects that the material will be applied by some type of lowpressure hand-

wand apparatus a

nd that the volume of

material per use

/application will be

relatively small.

• No significant toxicity was observed in the 28-day de rmal toxicity study in the rabbit, 91-day dermal toxicity study in the rat, 2-y ear chronic/carcinogenicity study in the rat, or the 92-week chronic/carci

nogenicity

study in mice. No systemic toxicity via de

rmal route was obs

erved in all three species studied.

• The active ingredient is in Toxicity Category IV via inhalation. Therefore, no significant inhalation toxicity is anticipated from the proposed use.

• Liver hypertrophy

seen in the one-week feeding study in rats is due to adaptive cha nges and not considered as a

toxicologically

significant ef

fect. This effe

ct was not observe

d in the subchronic and carcinogenicity feeding studies in rats. No sig nificant systemic toxicity was observed in the subchronic feeding studies in rats at doses up to 1,000 mg/kg/day or in the c arcinogenicity feeding stu dy in rat s at dos es u p to 195 to 2 59 m g/kg /da y.3

• There was no evidence of carcinogenicity in mice or rats either by dermal or oral routes of exposure.

• Mutagenicity studies indicate no conclusive evidence of mutage nic effects of alpha olefin.

• No evidence of q

ualitative or qua

ntitative suscepti

bility was obs

erved in the deve

lopmental toxicity studies in mice, ra

ts and rabbits.

• Developmental toxicity was observed at or above maternally toxic doses (300 mg/kg/day or above) in mice, rats and rabbits. The effects of concern (i.e. maternal deaths) seen in these studies a re probably due to bolus administration (i.e. gavage) and/or surfa ctant property of the chemical. The maternal toxicity

is the most likely

cause of fetal d

eaths and skeleta

l variations seen

in these studies. This is fur

ther supported by

the fact that no sig

nificant sy

stemic toxicity

was observed in feeding (oral) subchronic and chronic studies in rats. The maternal and fetal dea ths would not be expected to occur via the dermal route, which is the anticipated route of exposure because no systemic toxicity were observed in subchronic and chr onic dermal toxicity studies. In addition, considering poor dermal absorption of alpha olefin (reported value of 0.6%), it would require almost 30,000 mg/kg/day oral equivalent doses to produce developmental eff ects and these types of exposure concentrations are not likely to occur in the work environment.

• An available published peer reviewed dermal absorption study indicates poor de rmal absorption (0.6%) of alpha olefin through the skin of rats. Although, there are several re porting/deficiencies in th is st udy, th e stud y adequa tely in dicat es ver y poor d ermal absor ptio n of al pha ol efin i n rats . In this study, dermal absorption was measured following dermal application of radiolabelled a lpha olefin to the intact skin and also through the damaged skin in which the stratum corneumwas removed prior to application of the test material. It is generally believed that the primary barrier property of the skin resides in the stratum corneum , the upper most layer of the skin. Approximately

0.6% of the applie

d dose was recov

ered following

24 hour exposure dur

ation via intact skin. Approximately 50.4% of the applied dose was recovered following 30 hour exposure duration via damaged skin. In these sets of experiments, several guideline repor ting/deficiencies were noted. Despite all of the guideline deficiencies, the study w as well conducted and does provide valuable

information on der

mal absorption in ra

ts that could be use

d for regulator

y risk assessment. I

f one looks at the r

esults of intact an

d damaged skin (

without barrier p

roperty), it showed that only 0.6% of the applied dose was recovered throug h intact skin compared to 50.4% of the applied dose through the damaged skin. One might expect 100% dermal absorption through the damaged skin (barrier less skin), however, in this study only half of the applied dose was recovered in various tissue examined. Based on this estimates, one could extrapolate approximately 1.2% dermal absorption via intact skin. Therefore, this study does provide a useful calculate d dermal absorption value of 1.2% dermal absorption of alpha olefin through the skin.

• The poor dermal a

bsorption of alpha

olefin is also suppo

rted by the r

esults of dermal to

xicity studies in mice, rats and rabbits which indicate no systemic toxicity at doses up to 100 mg/kg /day (HDT) in the subc

hronic toxicity

studies and 38.9 mg

/kg/day (H

DT) in a dermal c

arcinogenicit

y study.

• The end-use-pr

oduct is a severe

eye irrita nt, however, the p

roposed labeling

is explicit regarding possible ocular injury and directs handle rs to use the appropriate Personal Protective Equ ipm ent to a voi d in

jur y.

• Based upon the me

thod of delivery

, the point of applic

ation, low applica

tion pressures, r

elatively small volumes of material that might be used per day and upon the label directed Persona l4

Protective Equipme

nt, HED expects tha

t dermal, inhalati

on and ocular expos

ure to be neglig

ible if applied as directed by the proposed labeling. I n addition, based on the specific sites of application and the fact that the burrows are sealed with soil after the application is complete, HED believes the post-application exposure to handlers or non-handlers is also negligible.

• Finally, only 6.91% of the alpha olefin is prese nt in the product. Based on the low concentration of alpha olefin in the product available for exposure, the low dermal absorption and the lack of demonstrated systemic toxicity by the dermal route and oral feeding studies, RAB1 of the HED has determined t

hat no quantitative r

isk assessment is r

equired.5 

### Lower Toxicity

Pesticide Chem

ical Focus Group Decision Docum

ent

" -Olefin Sulfonate Sodium EH-2001 (Alpha-Olefin Sulfonate Sodium and White Mustard Seed) HAZARD CHARACTE RIZATION Sodium " -Olefin Sulfonates are long-chain sulfonic acids which are used in the Cosmetic I ndustry as surfactants or cleansing agents. International J. of Toxicology , 17 (Suppl.5) 39 - 65, 1998, published the findings of Cosmetic Ingr edient Review Expe

rt Panel on sodium " -olefin sulfonates. The review compiled experimental data concerning the safety information of Sodium C14 - 16 Olefin Sulfonate, Sodium C12 - 14 Olefin

Sulfonate, Sodium C14

- 18 Olefin Sulfona

te, and Sodium C16 - 18

Olefin Sulfonate. Sodium olefin sulfonates are primarily the mixtures of sodium hydroxy alkane sulfonates and sodium alkane

sulfonates as show

n in figures belo

w. Since the reg

istrant submitted the

acute studies with Sodium C14 - 16 Olefin Sulfonates, this review of the toxicity studies is primarily on C14 -16 sulfonates only, with exceptions being the developmental and mutagenicity studies which used other sulfona

tes. Sodium C14 - 16 " -Olefin Sulfonate has low acute toxicity (Tox. Cat. III ) via oral, and dermal routes of exposure. It is mild skin irritant (Tox. Cat. III) and not a skin sensitizer. It is a severe eye irritant (Tox . Cat I) causi ng corn eal op acity l astin g 7 days p ost ap plica tion . The i nhala tion LC 50is > 229 mg/L for powdered aerosol flakes (90% a.i.), which indicate low toxicity via inhalation route. No subchronic, repeat dermal, developmental or mutagenicity da ta are submitted as required under Tier 1 data requirements for Non-food use chemical. However, the review by the Cosmetic Ingredient Review Expert Pane

l published in the I

nternational J. of T

oxicology, 1 7 (Suppl.5) 39 - 65, 1 998, did provide the necessary toxicity information to fulfill Tier 1 Data r equirements. In subchronic feeding studies in rats there was no systemic toxicity observed at doses up to 1000 mg/kg /day (Limit Dose). Systemic toxici

ty was not obs

erved in 90-day

dermal study

in rabbit but result

ed in mild to moderat

e skin irritation at 100 mg/kg/day. In chronic toxicity /carcinogenicity study in ra ts decreased body weight in males and females was observed at 195 and 259 mg/kg/day , respectively (LOAEL

) and the NOAEL was 96 and 132 mg/kg/day , respectively. Twice dermal application of 0.2 mL (30% a.i) for 92 weeks in rats did not elicit any systemic toxicity up to 38.9 mg/kg/day (NOAEL). The AOS was not carcinogenic in both the feeding and dermal carcinogenic ity studies. The AOS (C14 - 16, C14 - 18) were not mutagenic in in vitro mut agen ici ty as says in 4 sep arat e tr ial s. In a ho st- med iat ed as say, the AOS (C14 - 16) caused positive response in salmonella typhimurium TA 1530. This positive response was probably due to incomplete ether extraction and higher pH (11.1). When the pH of the extract was adjusted with sulfuric acid to 8.5, the salmonella typhimurium TA 1530 test was negative. Developmental toxicity using C14 - 18 AOS in mice, rats and rabbits, did not cause developmental toxicity below maternal toxicity. In Mice materna l toxicity NOAEL was 2 mg/kg/day based on reduced body weights and litter loss observed at 300 mg/kg/da y (LOAEL). T

he Developmental6

NOAEL was 300 mg/kg/day ba sed on decreased pup weight, major malformations (cleft palate) and minor skeletal anomalies. In the rat there was no maternal or developmental toxicity was observed up to 600 mg/kg/day (NOAEL; HDT). Ra bbit appears to be sensitive species for developmental toxicity. Mat erna l to xic ity w as s imi lar to m ice i.e ., t he N OAE Ls (2 mg/ kg/d ay) an d LOA ELs ( 300 mg/ kg/d ay) are same as mice. The developmental NOAEL is 2 mg/kg/day and LOAEL is 300 mg/kg/day based on increased incidence of minor skeletal anomalies and extra ribs. The AOS was readily absorbed from the GI tract, and rapidly excreted in the ur ine (72%) and feces (22%) within 24 hours. About 4.3% was excreted in the bile within 12 hours after oral administration. Blood levels peaked in 3 hours then declined. Less than 0.03% was retained in the tissues after 24 hours. Upon intravenous administration, half of the administered dose was excreted within 1 hour and 90% was excreted 0 - 6 hours. The data suggest metabolites may be more polar than the parent and the intact AOS could bind with proteins while metabolites can not. Dermal absorption study indicate about 0.6% was absorbed from the intact skin. The End-Use formulation EH-2001 has low acute toxicity (Tox. Cat. IV) via oral, der mal and inhalation routes

of exposure. I

t is not a skin irritan

t (Tox. Cat. IV). It is a skin sen sitizer. The skin sensitization potential is probably due to impurities such as unsaturated and chloro-1,3-sultones. The formulation is a severe eye irritant (Tox. Cat I) ca using corneal opacity lasting 7 day s post application. File Symbol: 72919-R Tolerance Reassessments ?: Yes [40 CFR 180.1001( c); sodium " -olefin sulfonate (sodium C 14- C 16 )(olefin sulfonate) ] Chemical Name: Alpha-Olefin Sulf onate Sodium (AOS) Chemical Category/Group: Alpha-Olefin Sulfonate Sodium - Surfactant CAS. Reg. No.: 68439-57–6 PC Code (active): 000905 PC Code (inert): 900631 and 900905 1. Structure: 

### R-CH-CH 2 

### -CH 2 

### -SO 2 

### -Na sodium 3-hydr

oxyalkane su

lfonate 

### R-CH=CH-CH 2 

### (OH)-SO 2 

### -Na sodium 2,3-alkane

sulfonate7

Sodium alkane sulfonates (AOS) are primarily a mixture of sodium hydroxy alkane sulfonate and sodium alkane sulf

onate. The sodium C 14 -C 16 AOS is also identified as tetradecenesulfonate or sodium tetradecensulfonate. 2. Alpha-Olefin Sulfonate Sodiu m Several types of studies were submitted to the Agency : (1) complete studies for review and evaluation, (2) articles from open-leterature, (3) an assessment by a Cosmetic Ingredient Review (CIR) expert panel, and (4) an assessment by the Soap and D etergent Association. The Agency does not have access to the studies that support the assessments. A toxicity profile of the information available for C 14 -C 16olefin sulfonate, sodium salt is below. Robust summaries of the submitted studies and artic

les from open-lite

rature and toxicity

profile are liste

d below: Oral Toxicity Species/Strain: Rat/Albino (HSD:SD) Conclusion: LD 50

Males = 2161 mg/kg Females = 1895 mg/kg Combined = 2000 mg/kg Tox Cat. III. Method: Groups of 5 young immature rats/sex/group were given a single oral dose of 1000, 2000 or 3000 mg/kg (purity unknown). Animals were observed for clinical signs of toxicit y for 2 weeks pos t-dosing. GLP: Yes Test Substance: Witconate AOK (Alpha-Olefin Sulfonate Sodium; Sodium c14-16), Lot # 3379,

purity - unknown Results: No deaths in 1000 mg/kg group. Two males and 3 females in 2000 mg/kg group a nd all animals in the 3000 mg/kg group died on day 1. Clinical signs of toxicity was seen at all dose levels which included pilorection, salivation, polyuria, diarrhea, respirator y chirp and gurgle , ptosis, decreased act ivity, nasal d ischarge, gas ping, aggressiveness, sensitivity to touch, exophthalmos and bloating were seen up to 10 days post dosing. Body weig hts of surviving animals were not effected by treatment except one male at

the 2000 mg/kg a

nd one females at

the 1000 mg/kg g

roups lost weight. Necropsy of the surviving animals was unremar kable. Animals died on the test exhibited discolored gastrointestinal contents and filled with gas, staining of muzzle, and matting and staining of genital hair and ventral hair.8

Reference: Kuhn, J. (1996) Acute Oral Toxicity of Witconate AOK in Rats. Stillmeadow, Inc.,

Sugar Land, T exas. Labora tory St udy No. : 2830 -96. J uly 1, 1 996. M RID 45307612. Unpublished. Rel iab ili ty: (2) valid with restrictions. Purity was not provided. Acute Dermal Toxicity Species/Strain: Albino Rabbit/New Z ealand White Conclusion: LD 50 > 2020 mg/kg

Severe dermal irritant; included erythema, edema, shallow latera l fissuring and coriaceousness. Tox Cat. III. Method: Groups of 5 male and 5 female rabbits were applied a single dose of 2020 mg/kg of

Witconate AOK in 0.535 mL deionized water/g of substance, covered with surgical gauze, secure

d with non-irritati

ng adhesive tap

e and maintained i

n place orthoped

ic stockinette. After 24 hours, the bandages were removed, the site was gently wiped with clean cloth and dermal irritation was scored at approximately 60 minutes, and on Days 4, 7, 10 and 14. GLP: Yes Test Substance: Witconate AOK (Alpha-Olefin Sulfonate Sodium; Sodium C14-16), Lot # 3379,

Purity - unknown Results: There was no mort ality during

the study. Th

ere were no cli

nical signs of toxic

ity were observed, except for a ruptured abscess in one animal which was considered unrelated to treatment. One male lost < 3% of body weight during the study. Severe skin irritation was observed which included erythema, edema, shallow lateral fissuring and co

riaceousness. Reference: Kuhn, J. (1996) Acute Dermal Toxicity of Witconate AOK in Rabbits. Stillmeadow,

Inc., S ugar Lan d, Tex as. Lab orato ry Stud y No.: 2 829-9 6. Ju ne 7, 1 996. M RID 45307613. Unpublished. Rel iab ili ty: (2) valid with restrictions. Purity was not provided. Acute Inhalation Toxicity Species/Strain: Rat/unknown9

Conclusion: LC 50> 229 mg/L for C14 - 16 flakes (90% a.i.) 1 hour exposure

LC 50> 221 mg/L for C14 - 16 sprayed dried f ormulation (17% a.i.) 1 hour exposure. Tox. Cat. Not determined Method: No experimental details were provided. Groups of 10 rats were exposed for 1 hour

to a powdered aerosol of either C14-16 AOS flake (90% active) or a spray -dried formulation containing 17% C14-16 AOS at concentrations of 229 mg/L and 221 mg/L, respectively. Five ra ts were sacrificed and examined grossly after the exposure, but did not specify the time of sacrifice. The remaining rats wer e necropsied 14 days after exposure. GLP: No. Test Substance: C14-16 AOS (Alpha- Olefin Sulfonate Sodium) Results: There were no deaths. The rats appeared normal except for increased pr eening behavior. Mild petechial hemorrhages in lungs were seen in 2 rats exposed to flakes and in one rat exposed to spray-dried formulation. Necropsy of the rats killed after 14 days were unremarkable. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: Low. Guideline method was not used. Primary Eye Irritation Species/Strain: Albino Rabbits/New Zealand White Conclusions: Corneal opacity was present 7 day s post application. Tox Cat. I. Method: 0.1 ml undiluted sulframin AOS (Lot # U3541, purity - unknown) was instilled in the

one conjunctival sac of 3 male and 3 females albino rabbits and observed at 24, 48, and 72 hours and at 4 and 7 days after treatment for ey e irritation. GLP: No Test Substance: Sulframin AOS (Lot # U3541, purity - unknown)10

Results: Corneal opacity was present in 4/6 rabbits 7 day s post treatment. Conjunctival irritation was present in 5/6 rabbits 7 days post treatment. The maximum irritation score was 26.8 a

t 48 hours and 15.5 a

t 7 days. Reference: Sabol, E. (1980) Ac ute Eye I rritation Study

of Sulframin AOS in Ra

bbits. Stillmeadow, Inc., Houston, TX 77036. Laboratory Study No.: 1930-80. November 12, 1980. MRID 45307614. Unpublished. Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and some experimental

details such as purity were not listed. Primary Dermal Irritation Species/Strain: Albino Rabbits/New Zealand White Conclusions: Sulframin AOS is moderately irritating to rabbit skin. Erythema a nd edema were observed at both th

e intact and abra

ded skin sites post 72

hours. Tox Cat. III. Method: The backs of 3 mal e and 3 female al

bino rabbits were

shaved 24 hours pr

ior to treatment. Two sites were identified on the backs, one intact and the other abraded. The backs were

abraded using a

hypodermic

needle. Each te

st site was applied

with 0.5 mL of undiluted sulframin AOS, covered with gauze pad, secured with adhesive tape. Twenty

-four hours afte

r treatment, dre

ssing was remove

d, and the animals were observed f

or erythem

a, edema, and es

char formation a

t 24, and 72 hours. GLP: No Test Substance: Sulframin AOS (Lot # U3541, purity - unknown) Results: Sulframin AOS is moderately irritating to rabbit skin. Erythema a nd edema were observed at both the intact and abraded skin sites post 72 hours. The mean score for erythema a

nd edema were 2.0 and 1.33, respe ctively. The

primary irr

itation score is 3.33. There was no signs of ulcerati on, necrosi s, or irrit ation (oth er than fiss uring) present 72 hours a

fter treatment. Reference: Sabol, E. (1980) Ac ute Dermal I

rritation Study

of Sulframin AOS in Ra

bbits. Stillmeadow, Inc., Houston, TX 77036. Laboratory Study No.: 1931-80. November 19, 1980. MRID 45307615. Unpublished. Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and some experimental

details such as purity were not listed.1 Calculated based on the assumption that an 250 g adult rat would consume 20 g/da y. 11 Dermal Sensitization Species/Strain: Guinea Pig/Hartley Conclusions: Sulframin AOS ( (Alpha-Olefin Sulfonate Sodium; Sodium C14-16) is not a dermal

sensitizer Method: Delayed Contact Hyperse nsitivity method by Buehler was employ ed to evaluate the sensitiz ation pote ntial of Su lframin AOS . Ten male an d ten female s guinea pigs were induced with applications of 25%w/v sulframin formulation is distilled water for 6 hours, once

a week for a tota

l of 3 applications. Following 2 wee ks after last induction exposure, the guinea pigs were challenged with 10% w/v sulframin AOS in distilled water.

Ten (5/sex; naive c

ontrol) guinea p

igs were also c

hallenged with 10% formulation. All the treated guinea pigs and naive controls were rechalleng ed with 5% formulation. GLP: No Test Substance: Sulframin AOS (EXP. 5113-161A), purity - unknown Results: No animal showed positive signs of sensitization during the first, second, or third

challenges at e

ither the 24 or 48 ho

ur observations. Reference: Kreuzmann, J. (1988) Skin Sensitization Study of C-14 to 16 AOS in Gu inea Pigs. Hill Top Biolabs, Inc., Miamiville, OH 45146. Laboratory Study No.: 88-3168-21. July 18, 1988. MRID 45357703. Unpublished. Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and some experimental

details such as purity were not listed. Subchronic Oral Toxicity a) Range-Finding Study Species/Strain: Rat/unknown Conclusion: NOAEL = 0.625% (125 mg/kg/day 1 ) LOAEL = 1.25% (250 mg/kg/day ), based on increased liver to body weight ratio.12

Method: Groups of rats were fed diets containing 0.625, 1.25, or 1.5% AOS (70% C14: 30%

C16; 125, 250 or 300 mg/kg/day, respectively ) for 7 days. No other details were provided. GLP: NO Test Substance: AOS (Al pha -Ol efi n Su lfo nat e So diu m; 7 0% C 14: 30% C16 ), u nkn own pur ity. Results: No systemic t oxicity was not

ed up to 125 mg/kg

/day. A slig

ht increase in live

r to body weig

ht ratio was noted i

n males. At 300 mg/

kg/day, a significant dec

reased in body weig

hts was noted for ma

les on day 2 a

nd day 7 in fe

males. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and some experimental

details such as purity was not listed. b) 90-day Feeding Stud y Species/Strain: Rat/unknown Conclusion: NOAEL $ 1000 mg/kg/day(HDT; Limit Dose)

LOAEL was not established Method: Groups of rats (n umber not specifi

ed) were fed di

ets containing A

OS (Alpha-Olefin Sulfonate Sodium 89.7%) at doses of 40, 200 or 1,000 mg/kg/day for 90 day s. No other details were provided. GLP: No Test Substance: AOS (Alpha-Olefin Sulfonate Sodium 89.7%). Results: No changes in hematology, clinical chemistry , feed consumption, gross or histopathologica

l lesions were not n

oted at the highe

st dose tested. A sli

ght increase in the liver to body weight ratio was noted. In the abse nce of histopathological and liver enzyme changes, the increased ratio was c onsidered as adaptive change. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423.13

Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and some experimental

details la cking. c) 91-day Feeding st

udy Species/Strain: Rat/unknown Conclusion: NOAEL $ 500 mg/kg/day(HDT)

LOAEL was not established Method: Groups of rats (number not specified) were fed diets containing C14-16 AOS

(Alpha-Olefin Sulfonate Sodium34% a.i.) at doses of 50, 150 or 500 mg/kg/day for 91 days. No other details were provided. GLP: No Test Substance: C14-16 AOS (Alpha-Olefin Sulfonate 34% a.i.) Results: No treatment-related toxicological or histopathological changes were noted at the

highest dose tested. Minor variations in hematology parameters wer e noted. No details were provided. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and some experimental

details la cking. Repeat Dermal Toxicity a) 28-Day Dermal Toxicity Species/Strain: Rabbits/unknown Conclusion: NOAEL $ 2mg/animal i.e., 0.8 mg/kg (base

d on 2.5 kg avera

ge weight of a

rabbit) for 1%aqueous solution of C14-16 AOS or a 1% AOS formulation LOAEL Was not established. Method: Rabbits were dermally treated with 0.2 ml (2 mg assuming 90% purity ) of a 1% aqueous solution of C14-16 AOS or a 1% AOS formulation for 28 days. No other details were provided.14

GLP: No Test Substance: C14-16 AOS or its formulation Results: Not irritating to rabbit skin. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: (2) valid with restrictions. Study not performed to the guidelines method was not

used. b) 91-day Dermal Toxicity Species/Strain: Rat/unknown Conclusion: NOAEL < 100 mg/kg/day (HDT) LOAEL = 100 mg/kg/day, based on mild to mo

derate skin irrita

tion. Method: Six rabbits were dermally treated with 2 ml/kg/day of a 5% aqueous solution of AOS (34% active) for 91 days. No other details were provided. GLP: No Test Substance: AOS (Alpha-Olefin Sulfonate Sodium 34%). Results: Mild to moderate skin irritation. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: (2) valid with restrictions. Study not performed to guidelines and experimental

details were not provided. Chronic Oral Toxicity- Rat Species/Strain: Rat/Sprague-Dawley Conclusion: NOAEL = 2500 ppm (125 mg/kg/day).

LOAEL= 5000 ppm (250 mg/kg/day), based on decreased body gains in males and females.15

Method: Alpha-Olefin Sulfonate (97.7% a.i., 60.4 Alkenyl sulphonate:39.6% hy droxyalkane sulphonate w/w) was administered in diet to groups of 50 male and 50 females rats at dose levels of 0, 1000, 2500, or 5000 ppm (0, 39, 96 or 195 mg/kg/day for males and 0, 57, 132, and 259 mg/kg/day for females) for 104 wee ks. Food consumption and body weights were recorded weekly . Urinalysis was conducted from 10 male and 10 female rats from the control and the high-dose group at 12, 25, 51, 77 and 103 weeks. Hematology and clinical chemistries were done 10/se x from the control and the high-dose groups at 13, 25, 52, 78 and 103 weeks. At the end of the treatment period all surviving animals were sacrificed, gross and histopathology of the organs performed as recommended by the guidelines. Appropr iate statistical analysis was performed to assess tumor incidence. GLP: No Test Substance: Alpha-Olefin Sulfonate (97.7% pure) Results: There was no treated-related effects on the clinical signs, food consumption,

hematology and clinical chemistries, organ weig hts and gross and histopathology. There was a slight depression in food consumption in females in 5000 ppm group, but the animals recovered during the second year of treatment. Body weight gains were significantly less in males (16%) and females (28%) during 14 - 26 weeks. Pancreatic islet tumors in 1000 ppm male rats (4), adrenal tumors in 2500 ppm males (8), thyroi

d tumors in 2500 ppm fe

males. These inci

dences consider

ed not related to treatment since t

here was no dose

response. Mammar

y tumors wer

e observed in females in all treated groups. The tumor evidence did not support that AOS was tumorogenic. Reference : Hunter, B., Be nson, H. (1976) L

ong-Term Toxicity

of Surfactant Alp

ha Olefin Sulphonate (AOS) in the Rat. Huntingdon Research Center, Huntingdon, UK. Toxicology 5:359-370. Rel iab ili ty: (2) valid with restrictions. Although the study was not performed as per the

guidelines, but the findings are peer reviewed. Therefore, the re sults are considered valid and could be used for regulatory purposes. Carcinogenicity - Dermal a) Rat Species/Strain: Rat/Long-Evan Conclusion: No evidence of carcinogenicity by application of 38.9 mg/kg C14-16 (38.9%) AOS twice weekly

for 2 year

s.16

Method: 50 rats/sex were tr eated for 2 y

ears as follows: a) deionized water b) hydrolyzed composite sample of C14-16 AOS and C16-18 AOS (30.0% ac tive) c) partially hydrol yzed sample of AOS (30.9% ) identical to (ii) but containin g a residual level of sultone d) commercial C14-16 AOS (38.9% active) Each preparation was applied as a 10% active (v/v) aqueous solution twice weekly to the clipped backs at a dose level of 1 mL/kg for 2 y ears. No other experimental details were provided. GLP: No Test Substance: Commercial C14-16 AOS (38.9% active) Results: Mean body w eights, food con

sumption, hematolog

y, urinaly sis, mortality

, and gross and histopathology were comparable to controls. No increase d incidence of tumors were in treated animals were observed. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: (4) not assignable. b) Mice Species/Strain: Mice/Swiss Webster Conclusion: No signific ant toxic ity or histop athology Method: 40/sex/group were treated three times a week for 92 weeks 0.02 mL of te st material applied shaved interscapular surface. The study is pertinent only for alpha olefins of C14-16. The six treatment groups are:

C 20% C14-18 AOS

C 25% C14-18 AOS

C 20% C14-16 AOS

C 25% C14-16 AOS

C 6.7% C16-1,4-sultone

C 8.3% C16-1,4-sultone GLP: No17

Test Substance: C14-16 AOS Results: No significant toxicity or histopathology attributable to treatment wa s found. Reference: Little, D. (1993) Environmental and human safety of major surf actants. Volume I. Anionic surfactants. Part 4. Alpha Olefin Sulfonates. National Technical Service (NTIS) report No. PB94-102423. Rel iab ili ty: (4) not assignable. Developmental Toxicity Species/Strain: Mice/strain unknown

Rats/strain unknown Rabbits/strain unknown Conclusions: Mice

Maternal Toxicity NOAEL = 2 mg/kg/day Maternal Toxicity LOAEL = 300 mg /kg/day, based reduced body weights, and litter loss; deaths at 600 mg/kg/day Developmental Toxicity NOAEL = 300 mg/kg/day Developmental To

xicity LOA

EL = 600 mg/kg

/day, base d on decreased p

up weights and major malformations (cleft palate) and minor skeletal anomalies Rats Mater nal To xici ty NOAE L $ 600 mg/kg/day (HDT) Maternal Toxicity LOAEL - not established Devel opmen tal To xici ty NOAE L $ 600 mg/kg/day (HDT) Developmental Toxicity LOAEL - not established Rabbits Maternal Toxicity NOAEL= 2 mg/kg/day Maternal Toxicity

LOAEL = 300 mg/kg/day , based reduced

body weig

hts, mortality and litter loss; all rabbits died at 600 mg/kg/day Developmental Toxicity NOAEL= 2 mg/kg/day Developmental Toxicity LOAEL= 300 mg /kg/day, based on increased incidence of minor skeletal anomalies and extra ribs. Method: Alpha Olefin Sulfonate was administered by gavage during gestation days 6 - 15 of pregnancy in rats and mice and 6 - 18 day s of pregnancy in rabbits at dose levels of 0, 0.2, 2, 300 and 600 mg/kg/day. Twenty rats and mice/group and 13 rabbits/group were used in the study. No further experimental details were provided, but looking at the data presented, maternal, litter and major formations, minor anomalies and18skeletal variations were evaluated which should be adequate to establish the endpoints for materna

l and developmenta

l toxicities. GLP: No Test Substance: Alpha Olefin Sulfo nate ( 60.4% alke

nyl sulfonat

e:39.6% hyd

roxy-alkane

sulphonate w/w) Results: Rats

Systemic toxicity was not observed at any dose level tested. Litter parameters (litter size, embryonic deaths, litter weig ht, mean pup weight), fetal abnormalities, ma

lformation and va

riations were una

ffected at all dose levels. Mice At 0.2 and 2 mg/kg/day, body w eights were reported slightly reduced a nd there was no clinical signs of toxicity were exhibited. At 300 mg/kg pilo-erection, reduced activity and weight loss was observed. At 600 mg/kg, there w as mortality (6/20), clinical signs and reduced body weights were r eported. At 300 mg/kg no dams died, but 5 dams their entire litter. The higher incidence litter loss resulted in the mean high embryonic loss and low mean litter size in the 300 and 600 mg/kg/day group. Excluding the litter loss the litter size and embryonic loss is comparable to controls. Mean pup weights at all doses were significantly lower tha n the controls, however, the decrease o

f 36% at the high d

ose at the matern

ally toxic dose

s appear treatm

entrelated. At other doses there was a decrease of 10 - 15% pup body weights, however, this decrease was due above laboratory concur rent pup body weights (1.23 vs 1.04). Similarly, the mean litter weights (8.95) at 600 mg/kg/day group considered treatment-related. Cleft palate was observed in 4 pups (from 3 litters) in 600 mg/kg/day and in 2 pups (from litter) at 300 mg/kg/day . The incidence was 1.3, 0, 0, 1.4 and 5.4% in the c

ontrol, 0.2, 2, 300 a

nd 600 mg/kg/day

groups. Rabbit All rabbits in 600 mg/kg/day group died and showed anorexia, diarrhe a and weight loss before death. At 300 mg/kg/day 1/13 rabbits died and survivors showed anorexia and weight loss. Autopsy revealed gastrointestinal chang es. There were no treatment-rela

ted systemic

toxicity/litter

effects at 0.2 a

nd 2 mg/kg/day

. At a maternal toxic dose of 300 mg/kg/day litter parameters were not a ffected, however, litter and fetal weights decreased 24 and 20%, respectively , compared to the controls. The authors suggested that the fetal effects were probably due to maternal toxicity. No major malformations and minor visceral anomalies were affected, but incidences minor skeletal anomalies (23 vs 6.7%) and proportion of pups extra ribs (86.9 vs 59%) significantly increased, compared to the controls.19

Reference: Palmer, A., Readshaw, M., Neuff, A. (1975) Assessment of the teratogenic potential

of Surfactants. Huntingdon Research Center, Huntingdon, UK. Toxicology (3): 107 - 113. Rel iab ili ty: (2) valid with restrictions. Although the study was not performed as per the

guidelines, but the findings are peer reviewed. Therefore, the re sults are considered valid and could be used for regulatory purposes. Gene Mutation Bacterial mutagenicity Testing a AOS Concentrat

ion (max) Assay (if i ndicated) Strain Tested Result

In vitro 10,000 ppm Reversio n plate Salmonella typhi murium TA 1535, 1536, 1537, 1538 Escherichia coli B/r WP try and try her Negative ( strains 1536, and 1538 smaller c olony siz e than His

+ strains but were not reve

rtants) 100 μg/plate Ames Salmonella typhi murium TA 98, 100, 1535, 1537, 1538 Negative
100 μg Salmonella typhi murium TA 98, 100 Negative 2 μg/plate Salmonella typhi murium TA 1535, 1536, 153 7, 1538 (7 preperations) Salmonella typhi murium TA 1535 (4 AOS compounds) Negative

In vivo 283 mg/kg AOS C14-16 (2 8.4% activ e) Host-mediat ed (rat) Salmonella typhi murium TA 1530, 1534 (+) in TA 1530 (-) in TA 1 534 Positive r esponse in T

A1530 could be due to: a) incomplete ether extraction b) higher pH 11 .3; when adju sted to pH 8.5 with H 2 SO 4 yielded negative respons e. AOS C14-16 (2 8.4% activ e) i.m. 0.1 mL/1% solution Host-mediated (mouse) Salmonella typhi murium TA 1535 Negative

a Copied from registrants re sponse to Agency de

ficiency letter 7/19/2001. Toxico logy Data. MRID 45551102. Metabolism Species/Strain: Rat/Wistar Conclusions: The metabolic fate of 14 C-AOS (a mixture of sodium alkenyl sulfonate and 14 Csodium-3-hy

droxy alkane

sulfonate) was r

eadily abso

rbed from GI

tract, and rapid

ly20 excreted in urine (72%) and feces (22%) within 24 hours, while 4.3% was excreted in the bile within 12 h

ours. By int

ravenous dosing

50% of the dose wa

s excreted in the urine within 1 hour and 90% in 6 hours. Results suggest no accumulation of AOS occurs and it is rapidly absorbed, metabolized and excreted. Method: Male Wistar rats were dosed with a single oral and intravenous dose of 100 mg/kg

(50 μCi) and 10 mg/kg (5 μCi) 14 C-AOS (a 55:45 mixtur e of sodiu m-3-hy droxy alkane sulfonate and sodium alkenyl sulfonate which was labeled 14 C). The mass balance, metabolic profile, and blood kinetics were evaluated. GLP: No Test Substance: 14 C-A OS ( a 55 :45 mix tur e of s odi um- 3-h ydro xy al kan e su lfo nat e and sod ium alk enyl

sulfonate which was labeled 14 C). Grea ter tha n 98 % rad io c hem ica l pu rit y. Results: The COS was readi ly absorbed

(about 80%) from

GI tract, an

d rapidly exc

reted in the urine (72%) and feces (22%) within 24 hours, while 4.3% was excreted in the bile within 12 hours after oral administration. Blood levels of radioactivity peaked 3 hours after dosin

g and then decli

ned. Tissue resid

ues in the cecum w

ere 0.8% but it was less than 0.03% in other tissues 24 after oral administration. Upon intravenous administration, half of the administered dose was excreted within 1 hour and 90% was excreted 0 - 6 hours in the urine. No intact labeled COS was found in the urine after oral or intravenous administration. The data suggest metabolites may be more polar than the parent and the intact COS could bind with proteins while the metabolites cannot. Reference: Inoue, S., O’Grodnic, J., Tomizawa, S. (1982) Metabolism of alpha-Olefin

sulfonate (AOS) in Rats. Biological Science Laboratory , Lion Corporation, Odawara, Kanagawa 256, Japan. Fundamental and Applied Toxicology 2: 130 - 138. May/June, 1982. Rel iab ili ty: (2) valid with restrictions. Although the study was not performed as per the

guidelines, but the findings are peer reviewed. Therefore, the re sults are considered valid and could be used for regulatory purposes. Dermal Absorpt

ion: Species/Strain: Rats/Wistar Conclusion: Dermal absorption 0.6%. Method: Groups of 3 male rats were applied with 0.5 ml of a 0.2% solution of 14 C-AOS

(specific activity 6.55 μCi/mg) to the dorsal skin as follows: (a) the intact skin dried21 naturally after application, (b) the intact skin wiped off 0.5 hour after a pplication, (c) the intact skin wipe

d off 1 1⁄2 hour afte

r application, (d

) the intact skin wi

th plastic cup contain ing 14 C-AOS solution and (e) the damaged skin without the stratum corneumdried naturally after application. Bile and urine were collected at 1, 3, 6, 9, 12, and 24 hours in all groups and in addition in groups an 30 hour samples were collected. The animals in groups (a) to (d) were sacrificed after 24 hour s and group (e) were sacrificed after 30 hours. The samples after appropria te preparation radioactivity was analyzed using liquid scintillation counting. GLP: No Test Substance: 14 C-AOS ( (a 55:45 mixture of 3-hydroxytetradecane- 1- 14 C sulfonate and $

-

tetradecane-1- 14 C sulfonate). Results: Under the experimental conditions (a), (b), and (c) the percutaneous absorption of 14 C-AOS was almost completed by 1.5 hour after application. The excretion of radioactivity in urine and bile was highest during 3 - 6 hours and g radually decreased and continued even after 70 - 90 hours after application. Under the experimental conditions (d) a small amount of radioactivity was continuously absorbe d from the skin. Under conditions (e) about 50% of the administered dose was excreted in the urine and bile, and the recoveries were 36.26% in the urine and 1.83% in the bile and 12.28% in the organs after 30 hours after application. Based on the results the calculated dermal absorption factor for intact skin is 0.6% Reference: Minegishi, K., Osawa, M., Yamaha, T. (1977) Percutaneous Absorption of " -Olefin

Sulfonate (AOS) in

Rats. National I

nstitute of Hy

gienic Science

s, Tokyo. Che

m. Pharm. Bull. 25: 821-825. Rel iab ili ty: (2) valid with restrictions. Although the study was not performed as per the

guidelines, but the findings are peer reviewed. Therefore, the re sults are considered valid and could be used for regulatory purposes. 4. EH-2001 End-Use Product Oral Toxicity Species/Strain: Albino Rat/Sprague-Dawley Conclusion: LD 50 > 5050 mg/kg in males and females

Tox. Cat. IV. Method: Five and 5 female rats were administered by g avage single dose of 3.35 mL/kg (density - 1.509 g/mL; 5050 mg/kg) EH- 2001 (Batch # 20000209) and were22 observed for clinical signs of toxicity at least three times a day on the day of dosing and once daily

for 14 days

. GLP: Yes Test Substance: EH-2001 (batch # 20000209), purity - unknown Results: The re wa s no mor tal ity d uri ng th e st udy. Cli nic al s igns inc lud ed d ecre ased act ivi ty,

diarrhea, polyurea, sensitivity to touch and withdra wn testes in males on the day of dosing. Body weights were not effec ted. No unremarkable gross pathology was observed. Reference: Kuhn, J. (2000) Acute Oral Toxicity in Rats. Stillmeadow, Inc., Sugar L and, Texas 77478. Laboratory Study No.: 5627-00. March 28, 2000. MRID 45307603. Unpublished. Rel iab ili ty: 1. Guideline method was used. Acute Dermal Toxicity Species/Strain: Albino Rabbits/New Zealand White Conclusions: LD 50> 2020 mg/kg in males and females

Tox. Cat. IV Method: Groups of 5 male and 5 female rabbits were applied a single dose of 1.34 mL/kg

(density - 1.5090 g/mL; 2020 mg/kg) of EH -2001, covered with surgical gauze, secured with non-

irritating adhe

sive tape and main

tained in place w

ith orthopedic stockinette. After 24 hours, the bandages were removed, the site was gently washed with water and wiped clean with cloth and dermal irritation was scored at approximately 60 minutes, and on Days 4, 7, 11 and 14. GLP: Yes Test Substance: EH-2001 (batch # 20000209), purity - unknown Results: There were no deaths during the study. Soft feces wa s observed in two males on day of dosing. Body weights were not effec ted. One male lost < 3% of body weight during the study. Slight to moderate irritation was observed during the first 4 day s of treatment, subsided by Day 11, except one male a nd 2 females still had slight irritation. By Day 14, desquamation (2 male s and one female) and alopecia were present (one male). Necropsy was unremarka ble.23

Reference: Kuhn, J. (2000) Acute Dermal Toxicity Study in Rabbits. Stillmeadow, Inc., Suga r Land, T exas 7 7478. Labora tory St udy No. : 5628 -00. M arch 3 0, 200 0. MR ID 45307604. Unpublished. Rel iab ili ty: 1. Guideline method was used. Acute Inhalation Species/Strain: Rat/Sprague-Dawley Conclusion: LC 50> 2.37 mg/L

Tox. Cat. IV Method: In a 500 L nose-only stainless stee l chamber, 5 males and 5 females rats were exposed to EH-2001 (batch # 20000209) aerosol concentrations (gravimetric) of 2.37 mg/L for 4 hours. Concentration of test substance (from the breathing zone) was determined gravimetrically twice per hour a nd nominally at the end of the exposure. Particle size was determined twice during the exposure, using Cascade impactor, at a rate of 7.3 L/min for a duration of 10 seconds. The MMAD and particle size distr

ibutions were ca

lculated. Obser

vations for clinic

al signs of toxicity were made frequently on the day of exposure and at least daily thereafter for 14 day s. Body weig

hts were determi

ned just prior to the

inhalation exposure

and on Days

7 and 14. GLP: Yes Test Substance: EH-2001 (batch # 20000209), purity - unknown Results: There was no mortality during the study . Decreased activity and pilorection were observed in all males and females for 6 hours; and were asymptomatic by Day 7. Body weights were unaffected. Ne cropsy was unremarkable. The exposure concentration in the chamber was 2.37 mg/L (gravimetric), the mean MMAD was 2.8 μm and the geometric standard deviation was 1.5. Reference: Kuhn, J. (2000) Acute Inhalation Toxicity Study in Rats. Stillmeadow, I nc., Sugar Land, T exas 7 7478. Labora tory St udy No. : 5629 -00. A pril 1 3, 200 0. MR ID 45307605. Unpublished. Rel iab ili ty: 1. Guideline method was used. Primary Eye Irritation Species/Strain: Albino Rabbits/New Zealand White24

Conclusion: Corneal opacity did not resolve by Day 21. Conjunctival redness and chemosis was

present on Day 14 and redness on day 17. Tox. Cat. I Method: 0.1 ml undiluted EH-2001 (Lot # 20000209, purity - unknown) was placed into the

conjunctival sac of right eye of 1 male and 5 females a lbino rabbits and observed at 1, 2 4, 4 8, a nd 7 2 ho urs and at 4 , 7, 10, 14, 17 a nd 2 1 da ys af ter tre atm ent for eye irritation. All eye were washed after with deionized water af ter 24 observation. Left eye acted as untreated con

trol. GLP: Yes Test Substance: EH-2001 (batch # 20000209), purity - unknown Results: The maximum average eye irritation score of 37.0 vs 80 (maximum score), obtained

at 24 hours after

treatment. Corne

al opacity w

as present in 6/6 a

t 24 hours, and 3/6 at 17 days and 1/6 at 21 day evaluation. I ritis was present 1/6 eyes at 48 and was cleared by 72 hours. Conjunctival redness was seen in 5/6 rabbits at 72 hours and 1/6 at day 17

and was clear by

day 21. Chemo

sis was present 6/6

at 24 hours and 1/6 at 14 days and was clear by 17 days. Therefore, EH-2001 is considered a moderate eye irritant

and is Tox. Cat. I. Reference: Kuhn, J. (2000) Acute Eye Irritation Study in Rabbits. Stillmeadow, Inc., Sugar Land, T exas 7 7478. Labora tory St udy No. : 5630 -00. A pril 5 , 2000 . MRID 45307606. Unpublished. Rel iab ili ty: 1. Guideline method was used. Acute Dermal Irritation Species/Strain: Albino Rabbits/New Zealand White Conclusion: The Primary Irritation I ndex (PII) = 0.6 Tox. Cat. IV Method: Three males and 3 females rabbits

were tested at e

ach site (right

) with 0.5 mL undiluted EH-2001. After application of the test material, the treatment site was covered with semipermeable dressing and maintained in contact with skin. At the end of 4 hours, the

dressing was re

moved, washed wit

h tap water, cle

aned dry wi

th cloth and observed for erythema and edema formation at 1, 24, 48 a nd 72 hours and on Day 7 after removal of the bandages. The skin reactions were evaluated as per Draize (1994).25

GLP: Yes Test Substance: EH-2001 (batch # 20000209), purity - unknown Results: Erythema was observed in five rabbits 1 hour post removal of bandag es and cleared of erythema by Day

7. The Primary Irritation I ndex score was 3.5 and the mean score was 0.6. Th

e Primary I

rritation score w

as 1.75. There wa

s no edema observed. Therefore, EH-2001 was considered slightly irritating and has a Tox. Cat. IV. Reference: Kuhn, J. (2000) Acute Dermal Irritation Study in Rabbits. Stillmeadow, Inc., Sug ar Land, T exas 7 7478. Labora tory St udy No. : 5631 -00. M arch 1 6, 200 0. MR ID 45307607. Unpublished. Rel iab ili ty: 1. Guideline method was used. Dermal Sensitization Species/Strain: Guinea Pig/Hartley-Albino Conclusion: Induced dermal sensitization . Also produced faint to moderate irritation. Method: Modified Buehler method was employed. Male and female g uinea pigs were assigned into 2 groups, designated as Groups I (5/sex) and I I (10/sex). Group I remained untreated and acted as naive control group. Group I I animals were induce with 0.5 mL EH-2001 once a week for 3 weeks. The animals were re sted for 2 weeks, then all animals (the treated animals along with naive controls) were challenged with 0.4 mL of undiluted test substance. Observation of skin reactions were made about

24 hours and 48 aft

er each treatm

ent. Observation

s for skin reactions were made about 48 hours after challenge. DNCB (1-c hloro-2,4- dinitrobenze) was

used as positive co

ntrol. GLP: Yes Test Substance: EH-2001 (batch # 20000209), purity - unknown Results: The test substance produced no irrit

ation in the naive c

ontrols animals af

ter a single treatment at cha

llenge. The tes

t substance produ

ced a very

faint to moderate irritation in the a

nimals of test gro

up. The averag

e skin reaction sc

ore was 0.6 vs 0 for the naive control. The positive control DNCB produced a mean score of 0.4.26

Reference: Kuhn, J. (2000) Skin Sensitization Study in Guinea Pigs. Stillmeadow, Inc., Sugar

Land, T exas 7 7478. Labora tory St udy No. : 5632 -00. M ay 3, 20 00. MR ID 45307608. Unpublished. Rel iab ili ty: 1. Guideline method was used. 3. Toxicity Profile

Guideline #/ Test Substance Study Ty pe - Dose Levels Results Alpha-Olefin Sulfonate Sodiu m 81-1 C14 -1 6 AOS Acute Oral - Rat (1996) MRID 45307612 Doses (mg/k

g): 0, 1000, 2000, or 3000 for males and females Acceptable/

guideline LD 50males = 2161 mg/kg

LD 50females = 1895 mg/kg Combined LD 50= 2000 mg/kg Tox. Cat. III

81-2 C14 -1 6 AOS Acute D ermal To xicity - Rabbits ( 1996) MRID 45307613 Doses (mg/k

g): 2020 single dermal dose Acceptable/

guideline Dermal LD 50males and females > 2020 mg/kg

Tox. Cat. III

81-3 C14 -1 6 AOS Powdered aerosol flakes (90% a.i.) Sprayed dri

ed formula

tion (17% a.i.) Acute Inha lation To xicity - Rat (1993) MRID - N/A Doses (mg/L) : 229 (powdered aerosol fl

akes C14 - 16, 90% a.i .) or 221 mg/L sprayed dried formulat ion (17% a.i . C14 - 16) for 1 hour Unacceptable/

guideline howe ver it is valu able informati

on for label

ing purposes. No deaths. Mild petechial hemorrhages in lungs of 2/5 and 1/5 rats exposed to flakes an d spray-dried formulations, respectively. LC 50> 229 mg/L for powdered aerosol f lakes LC 50> 221 mg/L for sprayed dr ied formul

ation27

81-4 C14 -1 6 AOS Primary Eye Irritation - Rabbit (1980) MRID 45307614 Doses: 0.1 mL Acceptable/

guideline Cornea l opacity was p resent in 4/6 ra bbits 7 days posttreatment. Co njunctival irritation was presen t in 5/6 rabb its 7 days post-tr

eatment. The maximum i rritati on score was 26.8 at 48 hours and 15.5 at 7 d ays post-treatm ent. Tox. Ca t. I 81-5 C14 -1 6 AOS Primary Derma l Irri tation - Rabbit ( 1980) MRID 45307615 Dose: 0. 5 mL Acceptable/

guideline Moderate skin irri tant. Eryt

hema and edema was observed at both the intact and abraded skin sites post 72 hours application . The mea n erythema a nd edem a scores we re 2.0 and 1.33, respectively. There were no signs of ulcerati on, necrosis, or irr

itati on present

72 hours post

applicat ion Tox. Cat. III 81-6 C14 -1 6 AOS Dermal Sensitization - Guinea Pig ( 1988) MRID 45357703 Doses: C14-16 a

t 25% for induction and 10 and 5% for challenge Acceptable/

guideline C14-16 A OS was not a dermal sensiti zer .

NA C14 -16 AOS (70% C14: 30% C16) One Week Feeding St udy - Rat (1993) MRID N/A Doses: 0, 0. 625%, 1.25% and 1.5% (0, 125,

250 and 300 mg/kg/day, respectively) fo r 7 days Acceptable/

nonguideline NOAEL = 125 mg/kg/day LOAEL = 250 mg/kg/day based on in

creased l iver to body weight ratios

82-1a AOS (8 9.7% a .i.) 90-day Fe eding Stud y - Rats (1993) MRID N/A Doses: 0, 40,

200, or 1,000 mg/kg/day Unacceptable/

guideline No changes in hematology , clinical chemistr

y, feed consumption, gross or histopat hological lesions were noted at any dose le vel. There was slight increas e in the liver to body weight ratios was noted in the absence of any histopatho logical or en zymatic finding s. NOAEL $ 1000 mg/kg/day (Limit Dose) LOAEL was not est

ablished. 82-1a C14-16 (34% a .i.) 91-day Feedi ng Study Rats (1993) MRID N/A Doses: 0, 50,

150, or 500 mg/kg/day Unacceptable/

guideline NOAEL > 500 mg/kg/day

LOAEL was not establish ed28

82-2b C14-16 AOS 28-day D ermal To xicity - Rabbit (1993) MRID N/A Doses: 1% aqueous

C14- 16 AOS or formula

tion containi ng 1% AOS for 28 days. NOAEL > 2 mg/animal i.e., 0.8 mg/kg (based on 2.5 kg average weight of

the rabbi

t) for 1% aqueous solut ion of C14-16 AOS o r formulation containing 1 % AO S for 28 d ays produc ed no effec t on the intact rab bit skin.

82-3b AOS (3 4% a.i) 90-day D ermal To xicity - Rabbit (1993) MRID N/A Doses: 2 mL/kg/day o f a 5% (10 0 mg/kg/da y) aqueous solut

ion of AOS (34% a.i.) for 90 d ays. Unacceptable/

guideline NOAEL < 100 mg/k

g/d ay ( HDT) LOAE L = 100 mg/kg/day b ased on m ild to mod erate skin irritation.

83-5 C14-16 AOS (9 7.9% a .i., alkenyl sulphonat

e 60.4: 39.6% hydroxyalkane sulphonate w/w

) Chronic

toxicity/carcino genicity - rat (1976) MRID N/A Doses: 0, 1000,

2500, or 5000 ppm (0, 39, 96,

or 195 mg/kg/day for mal es and 0, 57, 132,

or 259 mg/kg/day for females) for 104 weeks Acceptable/

guideline NOAEL = 2500 ppm

LOAEL = 5 000 ppm, based on decreased body weight gains in males and females. No evidence of carcinoge nicity

.

83-2 Essentiall

y hydrolyzed C14-1 6 and C 16-1 8 AOS (30% a.i.)
Part ial ly h ydrolyzed AOS (30.9% a.i.) + contains residual levels of sul

tone Commer cia l C14- 16 AOS (38 .9%) Carcinog enicity Derm al - Rat (1993) MRID N/A Doses: 0.02 mL twice a week for 92 weeks Unacceptable/

guideline No treatm ent-related system ic toxicity was ob served.

NOAEL > 38.9 mg/kg/twice a week. No evidence of carcinoge nicity

.

83-2 20% C14- 18 AOS 25% C14- 18 AOS 20% C14- 16 AOS 25% C14- 16 AOS 6.7% C16-1,4-sultone 8.3% C16-1,4-sultone Carcinog enicity Derm al - Mice (1993) MRID N/A Doses: 0.02 mL twice a week for 92 weeks Unacceptable/

guideline No significan t toxicity/histopatho logy attributab le to

treatme nt was f ound.29

83-3a C14-18 AOS (60.4 % alkenyl sulfonate:39 .6%hyd roxyalkane sulphonat

e w/w) Develo pmental T oxicity -

Mice (1975) MRID N/A Doses: 0, 0.2, 2, 300 and 600 mg/kg/da

y on gestat ion days 6 - 15 Acceptable/

nonguideline Maternal Toxi city NOAEL = 2 mg/kg/day Maternal Toxi

city LOAEL = 300 mg/kg/da y, based on reduced body we

ights, and li

tter l oss; deat hs at 600 mg/kg/day. Developmen

tal Toxi city NOAEL = 300 mg/kg/day Developmen

tal Toxi city LOAEL = 600 mg/kg/day, based on decre ased pup weights, and m ajor malfo rmations (cle ft palate) and minor skeletal ano malies.

83-3a C14-18 AOS (60.4 % alkenyl sulfonate:39 .6%hyd roxyalkane sulphonat

e w/w) Develo pmental T oxicity -

Rats (1975) MRID N/A Doses: 0, 0.2, 2, 300 and 600 mg/kg/da

y on gestat ion days 6 - 15 Acceptable/

nonguideline Maternal Toxi city NOAEL $ 600 mg/k g/d ay ( HDT) Maternal Toxi

city LOAEL not established Developmen

tal Toxi city NOAEL $ 600 mg/k g/d ay ( HDT) Developmental Toxicity LOAEL not established

83-3b C14-18 AOS (60.4 % alkenyl sulfonate:39 .6%hyd roxyalkane sulphonat

e w/w) Develo pmental T oxicity -

Rabbits ( 1975) MRID N/A Doses: 0, 0.2, 2, 300 and 600 mg/kg/da

y on gestat ion days 6 - 18 Acceptable/

nonguideline Maternal Toxi city NOAEL = 2 mg/kg/day Maternal Toxi

city LOAEL = 300 mg/kg/da y, based on reduced body weig hts, mortality and litter loss; all rabbits died at 60 0 mg/kg/da y. Developmen

tal Toxi city NOAEL = 2 mg/kg/day Developmen

tal Toxi city LOAEL = 300 mg/kg/day, based on increased incidence of minor skeletal anomalies and extra ribs.

84-4 AOS products (21 -

38% a.i.) In in vitro mutagenicity MRID 45551102 Doses: 2, 100 a

nd 100 μg/plate; 10 000 pp m in reversion plat

e assay Negative in 4 separate tests

84-4 C14-16 AOS (2 8.4% a .i.) In in vivo mutagenicity Host-mediated - r

at MRID 45551102 Doses: 283 mg/

kg (28.4% a.i. C14- 16) (+ ) in Salmonell a typhimur

ium TA 1530 (-) in Salmonell

a typhimur

ium TA 1534 Positiv e response

in TA1530 could be du e to: a) incompl

ete eth er extra ction b) higher p H 11.3 ; when adju sted to pH 8.5 with sulfuric acid yielded neg

ative response

.30

85-1 C14-18 AOS (55% mixture of sodium-3- hydroxy alkane sulfonate:45 sodium alkenyl su

lfonate) Metabolism - Rat (1982) MRID N/A Single oral and intrav enous dose of

100 mg/kg (50 μCi

) and 10 mg/kg (5 μCi

) a 55:45 mixture of sodium-3- hydroxy alka ne sulfonate and sodiu m alkenyl sulfon ate whic h was C14 labeled. Acceptable/

nonguideline The AO S was read ily absorbed from GI tra ct, and rapid ly excreted i

n the uri ne (72%) and feces (22%) wit hin 24 hours; 4.3% was excret ed in bile wi

thin 12 hours af

ter oral administrat

ion. Blood levels

peaked in 3 hours th

en declined. Less

than 0.03% was retai

ned in the ti

ssues aft er 24 hours. Upon intravenous administration, half of t he administered dose was excreted wit hin 1 hour and 90% was excreted 0 - 6 hours.

The data sug

gest met abolites may be more polar than the parent and the intact AOS could bind with proteins w hile metabo lites can not.

85-3 AOS (55% 3- hydroxyt etradecan

e-1- 14 C sul fon ate :45 % $

- tetradecane-1- 14 C sulfonate) Dermal Absorption - Rat (1977) MRID N/A Doses: 0.5 mL of

a 0.2% solution of 14 C-AOS (specif ic acti vity 6. 55 μCi/mg) w as applied to the dorsal skin and various scenarios were attempt ed. Acceptable/

nonguideline Der mal absorption -

0.6%.

EH-2001 End-Use Product 81-1 Acute Oral Toxicity - Rat (2000) MRID 45307603 Groups of 5 mal

e and 5 females rats were given a single oral dose

of 3.35 mL/kg (dens

ity - 1.509 g/mL; 5050 mg/

kg) of EH-2001 formulation. Acceptable/

guideline Oral LD 50> 5050 m g/kg in males and females.

Tox. Cat. IV31

81-2 Acute D ermal To xicity - Rabbit (2000) MRID 45307604 Doses: G roups of 5 male and 5 female rabbits were applied with a single dose of 2020 mg/kg to 10% clipped body surface area for 24 hours. Acceptable/

guideline Dermal LD 50> 2020 mg/kg (Limit Dose) in males and

females. Tox. Cat. IV

81-3 Acute Inhalat ion - Rat (2000) MRID 45307605 Doses: G roups of 5 male and 5 female rats were exposed nose only to aerosol con

centrat ion of 2.37 mg/L EH-2001 for 4 hours. Acceptable/

guideline LC 50> 2.37 mg/L for m ales and females.

Tox. Cat. IV

81-4 Primary Eye Irritation - Rabbit (2000) MRID 45307606 Dose: 0.1 mL of

EH-2001 was instilled int

o the conjunctiv al sac of 1 ma le and 5 fem ale rabbits; eye were washed 24 hours after instil

lation. Acceptable/

guideline Corneal opacity was not resolved by Day 21.Conjunctival redness and chem osis were present on D ay 14 and redne ss on Day 17. Tox. Ca t. I

81-5 Primary Derma l Irri tation - Rabbit (

2000) MRID 45307607 Dose: Groups of (3 males and 3 fem ales) rabb its were derm ally exposed to 0.5 mL of EH-2001 for 4 hours. Acceptable/

guideline EH-2001 did not induce irritation and erythema at the 1 hour

post application. Irritat ion was resolved by 72 hours. The mean prim ary irritation inde x (PII) was 0 .6 i.e., slightly irritating. Tox. Cat. IV32

81-6 Dermal Sensitization - Guinea Pig (

2000) MRID 45307608 Dose: In modified Buehler method guinea pigs were exposed to

EH2001. Acceptable/

guideline There w as a 100% incidence o f reaction to ch allenge with

EH-2001. EH-2001 is a dermal sensitizer.