**UNITED STATES ENVIRONMENTAL PROTECTION AGENCY WASHINGTON, D.C. 20460** 

OFFICE OF CHEMICAL S AFETY AND POLLUTION PREVENTION September 11, 2015 **MEMORANDUM SUBJECT:**  PC Code: 900177 ; Dimethyl Sulfoxide ; Human Health Risk Assessment

and Ecological Effects Assessment to Support Proposed Exemption from the Requirement of a Tolerance When Used as Inert Ingred ients in Pesticide Formulations .

**All PC Code:** 900177 **DP Barcode:** N/A **Decision No.: N/A**  **Regulatory Action:** Inert Tolerance Exemption; 40 CFR 180.9 2 0 . **Petition No: IN**  **- 10**  **713 All CAS**  **Reg. No**  **: 67**  **- 6**  **8 -**  **5 Inert Tracking No:**  **N/A FROM:**  Lisa Austin , Ph.D., Toxicologist

Chemistry , Inert s and Toxicology Assessment Branch ( CI T AB) Registration Division (7505P) **THROUGH:** Kerry

L eifer, Team Leader Chemistry , Inerts and Toxicology Assessment Branch (CITAB)2 Registration Division (7505P) **TO:** PV Shah , Ph.D., Branch Chief

Chemistry , Inerts and Toxicolog y Assessment Branch (CITAB) Registration Division (7505P) **EXECUTIVE SUMMARY** EPA received a petition ( PP # IN - 10713 ) from ISK BioSciences ( 7470 Auburn Rd., Suite A, Concorde, OH 44077 ) , requesting that 40 CFR 180.920 be amended b y modifying the exemption from the requirement of a tolerance for residues of dimethyl sulfoxide ( here in referred to DMSO, CAS Reg . No. 67 - 6 8 - 5 ) in c ycl aniliprol e formulations for pre - harvest applications including post - emergence use. DMSO will be used as a diluent in c ycl aniliprole formulations (60.02% (w/w) with upper and lower limits of 61.82% and 58.22%, respectively) under 40 CFR p art 180.9 2 0 . DMSO has an ex isting tolerance exemption for use as a pesticidal inert under 40 CFR 180.920 as a solvent or co - solvent for formulations used before crop emerges from soil or prior to formation of edible parts of food plants (EPA, 2006). EPA published the notice of filing (NOF) for this petition in the Federal Register on March 4, 2015 ( 80 FR 11611 ). No comments were received in response to this notice. The following documents are available on DMSO: the IU CLID summary (2003) and the OECD SIDS document (2000). The Agency reassessed DMSO in 2006. As no new data w ere submitted the agency

relied primaril y on and reproduced data from the se documents and previous assessment. Below is a summary of the critical studies that comprise the toxicity profile of DMSO. For more details on all available toxicity studies please refer to the aforementioned documents included in MRID 49388701. DMSO has low acute toxicity via the oral and dermal routes in rats and mice and the inhalation route in rats . The acute oral LD50 > 7,92 0 mg/kg in rats and mice. The acute dermal LD50 > 40,000 mg/kg in rats and mice. The acute inhalation L C50 > 1600 mg/m 3 ( equivalent to 277 mg/kg) in rats. It is a dermal , ey e and gastric irritant in rats and rabbits . Itis a sensitizer in guinea pigs.3

Overall sy stemic toxicity with regard to DMSO is low . The targe t organ of toxicity is the ey e . C hanges in the lens of the ey es such as refractile changes in the lens and lens composition is seen in various animals at doses above the limit dose (1000 mg/kg/day). Subchronic and c hronic toxicity studies were availab le in the rat, rabbit and dog. In subchronic studies in dogs and rabbits, ey e effects were observed at > 2,500 mg/kg/day following up to 24 weeks of DMSO treatment. In chronic oral toxicity studies in the dog, and rabbit , toxicity

was manifested as decreased body - weight gain and /or change s in the refractive index of the nuclear region in the ey e at doses > 1 mL/kg/day (1100 mg/kg/day). Reproduction/developmental toxicity studies were available for review. In the rat, n either maternal nor fetal effects were observed up to 1000 mg/kg/day , the limit dose. Maternal ( decreased bod yw eight gain and food consumption ) and fetal ( decreased fetal body weight and delay ed rib ossification ) effects were observed at 5000 mg/kg/day (5 times the limit dose) . Reproduction was also not effected up to 1000 mg/kg/day . In rabbits, maternal toxicity was manifested as r educed food consumption and body weight gain at 1000 mg/kg/day . The no observable adverse effect level (NOAEL) was 300 mg/kg/day . Developmental toxicity was not observed up to 1000 mg/kg/day . Classical carcinogenicity studies with DMSO were not availabl e. However, cancer initiation/promotion studies are available and were used to ascertain the cancer potential of DMSO. In a study

in rats, DMSO (50 ppm , equivalent to 5 mg/kg/day) in the drinking water did not a ffect the latency

or frequency

of tumors induced b y 7,12 - dimethylbenz[a] anthracene (DMBA) , a known carcinogen. In an initiation - promotion study in mice, DMSO (0.1 mL ) didn’t induce skin tumors following a single dermal application of DMBA. Also, tumors were not observed in rats when DMSO (0.02 mL ) was applied dermally . There are mixed results in mutagenicity studies available with DMSO , but the weight of evidence showed no genotoxic risk from exposure to DMSO. Dermal toxicity

studies we re available in the rabbit and dog . In subchronic and chronic toxicity studies with rabbits and dogs, lenticular changes were observed at doses 1.1 g/kg/day (1100 mg/kg/day). In toxicity

studies with rats via the inhalation route of exposure, systemic toxicity was not observed at doses up to 0.964 mg/ L ( equivalent to 250 mg/kg/day). The lowest observed adverse concentration (LOAEC) was 2.783 mg/1 ( equivalent to 722 mg/kg/day) based on respiratory tract ir ritation.4

Neurotoxicity

studies were not available for review, however, evidence of potential neurotoxicity

was not observed in the submitted studies. In addition, functional observation battery parameters were not affected following inhalation exposur e in rats for 13 weeks. An immunotoxicity

study with DMSO was available for review. DMSO administered via the inhalation route of exposure caused an increase in the number of antibody secreting cells in male rats at concentrations > 0.310 mg/ L (equivale nt to 80 mg/kg/day) . However, this effect is not considered adverse. In mice, 5% DMSO ( equivalent to 27,000 - 52,000 mg/kg/day) administered in the drinking water for 6 weeks caused did not cause immunotoxicity . In the rat and monkey , DMSO administered via the oral and/or dermal route was rapidly absorbed, metabolized and excreted. Excretion was primarily via urine, feces was a minor route in the rat only . The major metabolite w as dimethyl sulfone . D imethyl sulfide , another metabolite, is eliminated through the breath . There was no bioaccumulation. EPA has determined that reliable data show the safety of infants and children would be adequatel y protected if the FQPA SF were reduced to 1X. Available toxicity

data on DMSO demonstrated no adverse effects at doses less than 1 , 1 00 mg/kg/day . Therefore, since there are no endpoints of concern for DMSO a qualitative dietary exposure risk assessment is appropriate. DMSO is used as an inert ingredient in pesticide products that could re sult in resident ial exposure. However, based on the lack of toxicity , a quantitative exposure assessment from “residential exposures” was not performed. The Agency

believes that establishing the tolerance exemption to include 40 CFR § 180.920 in a cyclaniliprole formulations will not significantly /measurabl y increase occupational exposure. The change in use pattern would be expanded to include post - emergence use in pesticide products on all pre - harvest food commodities. Based on the regulatory history of DMSO and no new toxicological information to indicate otherwise, it is not necessary to quantitatively

assess occupational exposure.5

DMSO has low toxicity

to aquatic organisms. Also, it is not expected to be toxic to mammals. Iti s not expected to persist in the environment and does not bioaccumulate. It i s biodegradable. Potential areas of environmental justice concerns, to the extent possible, were considered in this human health risk assessment, in accordance with U.S. Executive Order 12898, "Federal Actions to Address Environmental Justice in Minority Populations and Lo w - Income Populations," http://www.eh.doe.gov/oepa/guidance/justice/eo12898.pdf . Taking into consideration all available information on DMSO ( CAS Reg. No. 67 - 68 - 5) , EPA concludes that there is a reasonable certainty that no harm will result to the general population or to infants and children from aggregate exposure to DMSO . Therefore, the establishment of an exemption from tolerance under 40 CFR 180.9 2 0 for residues of DMSO f or use as an inert ingredient (diluent) at a level not to exceed 62% in pesticide formulations used before crop emerges from soil or prior to formation of edible parts of food plants; for pesticide formulations used after crop emerges but before harvest, provided that the potential for increased residues of the formulation’s active ingredient(s) in or on food commodities has been assessed, can be considered safe under section 408 of the FFDCA. **2 .**  **BACKGROUND** On May 14 , 201 4 IS K BioSciences (7470 Auburn Rd., Suite A, Concorde, OH 44077) , submitted a petition ( IN - 107 13 ) requesting that 40 CFR 180.920 be amended by modifying the exemption from the requirement of a tolerance for residues of DMSO ( CAS Reg. No. 67 - 68 - 5) in c yc laniliprole formulat ions for pre - harvest applications including post - emergence use. DMSO will be used as a diluent in c yc laniliprole formulations (60.02% (w/w) with upper and lower limits of 61.82% and 58.22%, respectively) under 40 CFR part 180.920. EPA published the notic e of filing (NOF) for this petition in the Federal Register on March 4, 2015 (80 FR 11611). No comments were received in response to this notice. Currently , DMSO has an existing tolerance exemption for use as a pesticidal inert under 40 CFR 180.920 as a solvent or co - solvent for formulations used before crop emerges from soil or prior to formation of edible parts of food plants (EPA, 2006).6

This document provides an assessment of the risk to human health and the environment for DMSO when used as an ine rt ingredient in pestici de formulations applied pre - harvest to agricultural commodities. **3 .**  **PHYSICAL AND CHEMICAL PROPERTIES** DMSO was previousl y reassessed in 2006 b y the agency. The following phy sical and chemical properties were reproduced from the reassessment document. **Table 1**  **. Physical and Chemical Properties of**  **DMSO Parameter**  **Value Source** 

Structure

EPA Inert Reassessment : One Exemption from the Requirement of a Tolerance for Dimethyl s ulfoxide (CAS Reg. No. 67 - 98 - 5) (2006) (MRID 49388701) Chemical Name Dimethyl Sulfoxide CAS # 67 - 68 - 5 Molecular Weight (g/mol) 78.13 Common Names Delton, Demasorb, Dimexide, Domoso, Methyl sulfoxide, Methyl sulfinylmethane Phy sical State Practically colorless liquid or crystals Melting Point (°C) 18.5 Boiling Point (°C) 189 Vapor Pressure ( mm Hg @ 25

°C ) 0.61 Partition Coefficient ( Log Kow) - 1.35 Water Solubility (mg/L at 25  C) 1.0x10 6 ;Miscible Henry's La w Constant ( atm m 3 /mol at 25  C) 1.51 x 10 - 97

**4 .**  **Hazard A**  **ssessment** The Agency reassessed DMSO in 2006. The following toxicity data are reproduced from that document (MRID 49388701) . Toxicity data are also reproduced from the IUC LID summary and OECD SI DS (MRID 49388701). **Acute Toxicity Studies Table**  **2 .**  **Acute Toxicity Profile of DMSO** 

**Study Type Results**  Acute Oral

– Rat LD50 = 14.5 - 28.3 g/kg (14,000 - 28,300 mg/kg) Acute Oral

– Mouse LD50 = 16.5 - 21. 4 g/kg (16,500 - 21,400 mg/kg) Acute Oral

– Rat LD50 >20 mL/kg Acute Oral

– Mouse LD50 =20 mL/kg Acute Oral

– Rat LD50 >15 mg/kg (50% DMSO) Acute Oral

– Mouse LD50 >14 mg/kg (50% DMSO) Acute Oral

– Mouse LD50 = 21400 mg/kg Acute Oral

– Mouse LD50 = 7920 mg/kg Acute Dermal

– Rat 40 - 50 g/kg (40,000 - 50,000 mg/kg) Acute Dermal

– Mouse 40 g/kg (40,000 mg/kg)8 **Study Type Results**  Acute Inhalation - Rat Mild pulmonary irritation (edema) 1600 mg/m3 ( equivalent to 277 mg/kg) (4 hr) 2900 mg/m3 ( equivalent to 3008 mg/kg) (24 hr) 2000 mg/m3 ( equivalent to 3458 mg/kg) (40 hr) Eye Irritation - Rabbit Slight conjunctivitis at 24 - hour observation period; cleared by 48 hours Eye Irritation - Rabbit Undiluted; no effects Eye Irritation - Rabbit Slight erythema Sensitization - Guinea pig Not a sensitizer **Acute Oral Toxicity** DMSO administered orally is of low acute toxicity

to rats and mice. After massive single doses, experimental animals exhibited rapid breathing, restlessness, and coma, leading to hy pothermia and death within a few hours (Gosselin, e tal., 1984 as cited in MRID 49388701 ). Administration of pure DMSO (100%) resulted in serious and rapid modification of the red blood cells, along with "certain coagulation defects" (Caujolle, et al., 1967 as cited in MRID 49388701 ). Lethal oral doses caused ataxia, myasthenia, decreased motor activity , and brad ypnea s hortly

after administration (Wi lson, et al., 1965 as cited in MRID 49388701 ). Non lethal oral doses produced only decreased motor activity , although polydipsia and poly uria were noted in rats following doses of 20 g/kg ( equivalent to 20,000 mg/kg) . Hy peremia and inflammation in the ey es of Sp rague - Dawley rats w ere observed following single oral doses of > 13 g/kg ( equivalent to 1 3,000 mg/kg) DMSO ( MRID 49388701 ) . **Acute Dermal Toxicity** DMSO is of low acute toxicity by the dermal route. Studies have demonstrated that DMSO is a skin irritant when used in sufficientl y high or multiple doses, but it is not a serious ey e irritant. (Fishman, 1969 as cited in MRID 49388701).9

**Acute Inhalation Toxicity**  In a study conducted by

Fishman et. al., Sprague - Dawley rats were exposed to DMSO at concentrations of approximately2900 mg/m 3 (equivalent to 3008 mg/kg) for 24 hours, 2000 mg/m 3 (equivalent to 3458 mg/kg) for 40 hours, or 1600 mg/m 3 (equivalent to 277 mg/kg) for 4 hours. Others were exposed to 200 mg/m 3 7 hours a day , 5 day s a week for a total of 30 exposures. Mild pulmonary irritation was observed at >1600 mg/m 3 . The LC50 > 1600 mg/m 3 (Fishman, 1969 as cited in MRID 49388701).

**Sensitization** DMSO did not induce skin sensitization in guinea pigs using the Buehler test (Marzulli and Maguire, 1982 ), the Magnusson and Kligman maximization method (Marzulli and Maguire, 1982; Nakamura et al., 1994 ) and the Draize test, the Cyclophophamide/CFA assay and the Split adjuvant test (Marzulli and Maguire, 1982) (As cited in MRID 49388701) . **Repeated Dose Studies** Subchronic Toxicity - Oral Dog: Young adult beagle dogs (6/sex/group) received daily oral doses of 0, 2.5, 5, 10, 20, or 40 g/kg/day (0, 2,500; 5,000; 10,000; 20,000; 40,000 mg/kg/day) DMSO, 5 day s/week for up to 23 weeks (Rubin and Barnett, 1967). Dose levels of 20 and 40 g/kg/day were not tolerated well and were reduced. After 9 weeks of administration, changes in the lens of the ey e were observed in dogs receiving 5 g/kg/day , and by the 18th week, all dogs at the lowest dose level were affected similarly . After 23 weeks, treatment was stopped and the dogs were observed for 31 weeks. Ey e changes persisted after withdrawal of DMSO b ut became less pronounced. The lowest observable adverse effect level (LOAEL) was 2.5 g/kg/day (2,500 mg/kg/day). The NOAEL10

was not established. Although, a NOAEL was not established it was noted that effects were seen above the limit dose, improved after treatment cessation and ey e opacity was not observed. Rabbit: New Zealand rabbits were treated orall y (drinking water) with 0.5 g/kg/day or 10 g/kg/day (500 or 10,000 mg/kg/day) of DMSO for 24 weeks (Wood and Wirth, 1969). The 10 g/kg rabbits exh ibited progressive changes in the optical lenses when examined with the biomicroscope at 2, 6, 12, and 24 weeks. The effect was characterized by refractile changes, and there were alterations in the composition of lens proteins. The no adverse effect lev el (NOAEL) was observed in the rabbits dosed at 0.5 g/kg/day (500 mg/kg/day) . The LOAEL was 10 g/kg/day (10,000 mg/kg/day) based on progressive changes in the optical lenses.

Chronic Toxicity Rat: In a chronic oral tox icity

study , 50 Sprague - Dawley rats/sex/group were administered DMSO [a 50% aqueous solution containing DMSO doses of 0, 1, 3, or 9 mL/kg/day (1100, 3300, or 9900 mg/kg/day)] via gavage 5 day s/week for 18 months. Clinical signs, body weight, and food consumption were monitored, and periodic assessments of hematological, clinical chemistry and urinary parameters, ophthalmoscopic examination, and gross and histopathological examinations were performed. There was a dose - related depression of weight gain in both sexes, with the excepti on of males receiving 1 mL/kg (in males, ca. - 20% at the top dose and ca. - 10% at the mid dose and in females, ca. - 20% at the top dose and ca - 10% at the mid and low doses). There was no accompan yi ng reduction of food intake. A slight reduction of h emog lobin and packed - cell volume (PCV) in male rats receiving 9 mL/kg ( 9900 mg/kg/day) was observed. Examination of the ey e revealed no changes in the retina or vitreous hum o r. No peripheral (equatorial) opacities were seen and there was no difference in incidence of polar opacities between the test and control animals. Prominent nuclear annuli were seen in a small number of11

animals towards the end of the study , but there was no dose - relationship to suggest an y treatment - related effects. The only relevant finding was some degree of change in the refractive index of the nuclear region in 3 rats receiving 9 mL/kg ( 9900 mg/kg/day) . The LOAEL = 9 mL/kg ( 9900 mg/kg/day) based on decreased weight gain in both sexes and the change in the refractive index of the nuclear region in the ey e. The NOAEL= 3 mL/kg/day (3300 mg/kg/day) (Noel et al, 1975 as cited in MRID 49388701). Dog: In a chronic oral tox icity

study , groups of 5 pur e - bred Pembrokeshire Corgi dogs/sex/group were administered DMSO [a 50% aqueous solution containing DMSO doses of 0, 1, 3, or 9 mL/kg/day

(1100, 3300, or 9900 mg/kg/day)] via gavage 5 day s/week for 18 months or 24 months . Clinical signs, body weight, and food consumption were monitored, and periodic assessments of hematological, clinical chemistry and urinary

parameters, ophthalmoscopic examination, and gross and histopathological examinations were performed. N o treatment - related effects were observed on body weight. At the mid - and high - dose levels, persistent diuresis but no renal damage was observed. In creased PCV and hemoglobin levels were observed at the high dose, although the ery throcy tes had normal hemoglobin concentrations and were of normal si ze. Lenticular changes (alterations in the refractive index of the central portion of the lens, lens opacit y, opalescence in the central region of the lens, and/or changes in vitreous humor) and biochemical changes in the lens (an increase in insoluble pr otein and reductions in soluble protein, glutathione, and water) were observed. Ocular changes were evident before 10 weeks at the high dose, with obvious progression with continued dosing. The sequence of changes occurred at the 3 mL/kg/day dose level, although the onset of changes was delay ed. At the low dose, nuclear refractive changes were observed after 6 months, but none of these dogs had opalescence. There were no marked clinical signs and onl y one accidental death occurred. Occasional isolated bouts of vomiting were seen at 9 mL/kg/day and transitory

"head shaking" was temporaril y observed during weeks 11 and 12 at this and the 3 mL/kg/day level. No adverse effects on body weight and food intake were recorded. Electrocardiogram ( ECG) records w ere normal throughout except for a transient, minimal slowing of the heart rate in recordings made after 4 weeks. Terminal radiology of excised bone showed no evidence of12

osteoporosis. Laboratory investigations confirmed the persistence of diuresis in do gs receiving 3 mL/kg and above but no renal damage resulted. Persistently

increased PCV, h emoglobin levels, and total red cell count were observed at 9 mL/kg. The red cells had normal h emoglobin concentrations (MCHC) and were of normal size (MCV). Bone marrow examination prior to termination revealed no evidence of toxic changes. It seemed possible that the constant diuresis had resulted in a balance with a slightly higher degree of h emoconcentration than is normally found. Ocular effects were observed after 5 - 10 weeks dosing in the dogs receiving 9 mL/kg including central (nuclear) lenticular changes with alteration of the refractive index (myopia) and by the fifth month, transitory equatorial opacities, central (nuclear) op alescence, and changes in the vitreous humor. Similar effects were observed in dogs receiving 3 and 1 mL/kg but they occurred more slowl y, respectively after 16 and 31 weeks of treatment. No abnormalities, other than those in the ey e, were detected duri ng macroscopic or microscopic examination of organs . Dogs withdrawn from the study at 18 weeks showed partial (high - dose) to complete (mid dose) recovery

by two y ears. No other histopathological abnormalities were observed. The LOAEL is 1100 mg/kg/day , based on ophthalmologic changes, and thus, no NOAEL could be determined for this stud y (Noel et al, 1975 as cited in MRID 49388701). Although, a NOAEL was not established it was noted that the ey e effects observed at 1100 mg/kg/day were less severe , occurred after 16 weeks of treatment , were reversible and ey e opacity was not observed . Reproduction/Developmental Toxicity Screening Test Rat: In a study performed according to the OECD Guideline # 421 and GLP, four groups of 12 male and 12 female Sprague - Dawley rats received DMSO (purity 99.977%), daily , by

oral (gavage) administration, before mating and through mating and, for the females, through13

gestation until day 21 post - partum, at dose - levels of 0, 100, 300 or 1,000 mg/kg/d ay (dosing volume: 5 mL/kg in purified water). Clinical signs and mortality were checked daily . Bod y weight and food consumption were recorded weekl y until mating and then at designated intervals throughout gestation and lactation. The animals were paired for mating and the dams were allowed to litter and rear their progen y until day 22 postpartum. The total litter sizes and numbers of pups of each sex were recorded after birth, pup clinical signs were recorded dail y and pup body weights were recorded on day s 1 and 4 post - partum and then weekl y until day 21 post - partum. The males were sacrificed after completion of the mating period. The bod y weight and selected organs weights were recorded and a macroscopic postmortem examination of the principal thoracic and abdominal organs was performed, with particular attention paid to the reproductive organs. A microscopic examination was performed on selected organs for all males in the control and high - dose group. The dams were sacrificed on day

22 postpartum (or on day 25 post - coitum for females which did not deliver or 24 day s after the end of the pairing period for unmated females) and a macroscopic examination of the principal thoracic and abdominal organs was performed, with particular attention paid to the reproductive org ans. In females that were apparently non - pregnant, the presence of implantation scars on the uterus was checked using ammonium sulfide staining technique. A microscopic examination was performed on selected organs of females in the control and high - dose group. The litters were culled on day 4 postpartum to four males and four females whenever possible and non - selected pups were sacrificed and retained. Selected pups sacrificed on day

22 post - partum and pups prematurely sacrificed or found dead were carefully examined for gross external abnormalities and a macroscopic post - mortem examination was performed. There were no unscheduled deaths and no treatment - related clinical signs during the study in males or females. There were no effects of treatment with DMSO on male body weight gains or food consumption. All DMSO treated female groups had slightly , but not statistically significant, lower body

weight gains than the controls during the pre - pairing period ( - 11, - 26 and

- 22% at 100, 300 and 1,000 mg/kg/day , respectively). However, on day 15, the body

weight differences between the treated females and the controls were less than 3% (control 268g, treated 264, 262 and 261 g at 100, 300 and 1,000 mg/kg/d ay , respectivel y). No effect was observed on the body weight gains during gestation or lactation periods, as well as on food consumption. No DMSO related effects were noted on mating and fertility parameters. There were no effects on estrus stages, on the mean number of day s taken to mate and no effect on the mean duration of gestation. There were no effects of treatment with the test item on pup mortality and survival, mean pup bod y weight, body weight gain or sex ratio. Pup necropsy finding was lim ited to a slightly increased incidence of dilated renal pelvis in the group treated at 1,000 mg/kg/d ay . However, this effect was not statistically significant, in th e range of the historical values and there was no dose - relationship between the group s treated at 100 or 300 mg/kg/d ay .14

Marginally

higher absolute (+10%) and relative (+8%) liver weights were noted in the FO males given 1,000 mg/kg/d ay . There were no treatment - related macroscopic findings. The treatment with DMSO at the highest dose - level di d not affect the incidences of microscopic findings in testes, epididy mides and ovaries. The NOAEL for parental toxicity was c onsidered to be 1000 mg/kg/d ay , the highest dose tested . The NOAEL for reproductive performance (mating and fertility) was considered to be 1,000 mg/kg/d ay and the NOAEL for toxic effects on the progen y was 1,000 mg/kg/d ay (Arkema, 2007 as cited in MRID 49388701 ). Rabbit: In a study to evaluate the potential toxic eff ects of DMSO on embry onic and f etal development, DMSO was administered daily by oral gavage at 100, 300 or 1,000 mg/kg/d ay to pregnant New Zealand white rabbits during the period of organogenesis (day 7 to day 28 post coitum). This study

was conducted according OECD guidelines #414 and GLP. No signs of maternal toxici ty were observed at 100 mg/kg/d ay . At 300 mg/kg/d ay , the signs of maternotoxicity

were confined to a slightly lower food consumption compared to the control group ( - 18%) during the first two day s of treatment but was not thereafter statisti cally different from the control group. At 1,000 mg/kg/d ay , the signs of maternotoxicity compared to the control group were: a slightl y lower food consumption during the first part of the study (days 7 to 15: - 17%) with a more marked effect during the fir st two day s of treatment (day s 7 to 9: - 32%) and slightly

lower body

weight gain during the first part of the study (day s 7 to 15: - 37%) with a more marked effect during the first two day s of treatment, where a slight body weight loss was recorded. There was no abortion at any dose - level. There was no effect, at an y dose - level, on the number of corpora lutea, implantation sites, post - implantation loss, number of live fetuses, fetal body weight and sex - ratio. The embryofetal development was not affected and there were no teratogenic effects at an y dose - level. The NOAEL for maternal toxicity was 300 mg/kg/d ay and the LOAEL was 1000 mg/kg/day based on reduced food consumption and body weight gain . T he NOAEL for embryofetal15

development was 1,000 mg/kg/d ay , the highest dose tested (Atofma, 2002b as cited in MRID 49388701). Carcinogencit y

Initiation/Promotion Studies : Rat: The effects of DMSO on the tumorigenic activity of 7, 12 - dimethylbenz[a] anthracene (DMBA) was investigated in rats. Three groups of 50 male Sprague - Dawley rats were given 20 mg DMBA b y gavage. In the two first groups, DMBA treatment was followed by 50 ppm DMSO in their drinking water 3 day s before or 3 day s after DMBA administration, respectivel y. The third group received no DMSO and served as untreated controls. The animals were treated or observed for up to 18 months, DMSO had no effect on the latency or frequency

of tumors induced b y DMBA. Rats receiving DMSO weighed more and had fewer tumors than did the controls at the end of the 18 - month study period. This was suggestive that DMSO decreased the total number of tumors, although the difference between treated and control rats did not reach statistical significance (OECD SIDS, MRID 49388701) . Two groups of 40 Sprague - Dawley rats were treated dermall y, 3 times a week, for at least 26 weeks, with 0.02 mL of a 1% solution of 9, 10 - dimethylbenzanthracene (DMBA) dissolved either in DMSO or acetone. A third group (40 animals) received onl y DMSO (0.0 2 mL ). The animals were observed weekl y and the number of tumors counted. Spe cimens were taken from all tumo rs for histological examination at the end of the experiment. In the rats treated with DMBA dissolved in acetone, the time of appearance of the fi rst tumor was shorter (26 weeks) compared to DMBA in DMSO (34 weeks) and the number of papillomas was greater (141 versus 32). No skin tumor was observed in the rats tre ated with DMSO alone (OECD SIDS, MRID 49388701). Mouse:16

In 20 IC R/Ha Swiss mice, dermal application of 0.1 mL DMSO, 3 times weekly over a period of 400 day s, after a primary treatment with DMBA (applied once only , 20ug in 0.1 mL acetone), induced no skin tumors (OECD SIDS, MRID 49388701). Dermal Toxicity Rab bits : Groups of male and female New Zealand white rabbits received daily applications of 50% or 90% aqueous solution of DMSO to normal and abraded skin at volumes equivalent to 0, 1.5, 2,7, 4.5 or 8.1 mL undiluted DMSO per kg (0, 1,650; 2,970 ; 4,950 and 8,910 mg/kg/d ay , respectivel y). Treatments continued for 6 months; animals were kept under observation for an additional 12 weeks after treatment was terminated. Each rabbit was observed daily , weighed weekl y and its water consumption was recorded durin g week 26. Ophthalmoscopic examination was performed on all animals before dosing commenced and then after 5, 8, 14, 20, 22, 28 and 33 weeks. Hematological inve stigations comprising PCV, h emoglobin, total and differential white cell count, and erythrocy t e sedimentation rate , were performed initially and at 4, 12, 26 and 32 weeks. At termination, each animal was subjected to a post mortem examination, with subsequent organ weight anal ys is and histopathology . Rabbits received dermal applications of DMSO to normal and abraded skin for a period of 23 weeks, when ocular changes were observed. Treatment was withheld from animals showing ocular changes; the remaining animals continued to receive DMSO applications for the scheduled 26 weeks (6 months). Macro scopic and hematological examinations, organ weight anal ys is, and histopathology did not reveal an y adverse effects. There were no changes in dermal morphology except for random occurrences of inflammatory reaction. After 23 weeks of treatment, adverse o cular (lenticular) effects were observed at all dose levels (1, 5, 3 and 8 rabbits with a normal skin and 3, 7, 4 and 11 rabbits with an abraded skin, respectivel y at 1.5, 2.7, 4.5, and 8.1 mL/kg). These were restricted to the lens, and consisted of nucle ar refractive changes. There was no effect on the peripheral lens, vitreous humor, or retina. The systemic NOAEL was not established. The sy stemic LOAEL was 1.5 mL/kg (1650 mg/kg/day) based on lenticular changes (OECD SIDS, in MRID 49388701) . In another stud y, m edical grade DMSO was applied daily to the shaved backs of groups of 10 rabbits for 30 day s at the dose of 0, 1 and 5 g/kg (1000 and 5000 mg/kg/day) . Blood was drawn by cardiac puncture from each rabbit 1 and 7 day s prior to treatment, and on treatment day s 1, 7,17

and 30. Serum chemistry , serum enzy me levels and liver alcohol deh ydrogenase levels were evaluated. Ey e lenses of each rabbit were examined with a biomicroscope before and during treatment. At the end of the study , all rabbits w ere autopsied and examined for gross pathology . There was no mortality . At both 1 and 5 g/kg/day (1000 and 5000 mg/kg/day) of DMSO administration resulted in elevated values for serum lactic deh ydrogenase . Transient elevations of the serum hemoglobin values were observed at 1 g/kg, as well as serum hemoglobin, total bilirubin and glucose at 5 g/kg. There were no gross lesions in DMSO or control animals examined at the end of the study . None of the animals treated with 1 g/kg/day display ed lenticular changes. All of the rabbits treated with 5 g/kg/day display ed microscopic ocular changes, which were first detected after 1 0 - 15 days treatment . The LOAEL was 5 g/kg/day (5000 mg/kg/day) based on ocular changes. The NOAEL was 1 g/kg/day (1000 mg/kg/day) (OECD SIDS, MRID 49388701) .

Inhalation Tox i city Rat: In 2 range - finding studies, three groups of 5 males and female Crl:CD®BR rats were exposed, by inhalation (snout only), to an atm osphere of droplets and/or vapo r generated from DMSO for 6 hours each day , over a period of 14 or 28 day s. The rats were killed on the day following the last exposure. The achieved study mean anal yz ed respirable concentrations of DMSO in air were 0, 0.520 (vapo r), 1.390 (> 64% of droplets < 7 um) and 5.36 0 mg/1 (> 64% of droplets < 7 um) for the 14 - day study and 0, 0.132 (vapo r), 0.507 (vapo r) and 1.886 (75% of droplets < 7um) mg/1 for the 28 - day study . In both studies, clinical signs both during exposures and at rest were monitored and recorded. Bod y weight wa s recorded twice weekl y and food and water consumption w ere recorded dail y. In the 28 - day study , body

rectal temperature was recorded on one occasion during the week prior the start of exposure and immediately after the 6 - hour exposure on week 1 and 4 for the control and high dose animals. Inthe 14 - day study , laboratory

investigations comprising anal ys is of h ematological and blood chemistry parameters were conducted at the end of the treatment period. Inboth studies, following sacrifice of the animals rats were subjected to detailed macroscopic examinations followed by preservation of tissues. Adrenals, kidneys, liver, lungs, spleen and testes were weighed and, the lary nx, lungs, nasal passages, trachea, and any gross lesions were subject to histopatho logical examination with in addition, ey es, kidney s and liver, in the 14 - day study .18

There were no unscheduled deaths during these studies. There were no clinical signs attributable to the administration of DMSO. There were no treatment - related effects on body temperature. In the 14 - day study , the mean body

weight gain of h igh and i ntermediate dose rats (both sexes) and l ow dose female rats was lower than control values. The mean cumulative food consumption of h igh and i ntermediate dose rats (both sexes) and l ow dose female rats was lower than the concurrent c ontrols. The mean cumulative water consumption of h igh dose female rats was higher than that of controls. None of these effects on the body weight gain and the foo d and water consumption were observed in the 28 - day study . In the 14 - day

study, there were no findings considered to be of toxicological importance in h ematology and blood biochemistry parameters investigated prior to sacrifice. There were no treatment - r elated macroscopic findings at necrops y and mean organ weights of all test groups were similar to control values in both studies. Aggregations of macrophages in the lungs, epithelial hy perplasia in the lary

nx and eosinophilic inclusions in the olfactory e pithelium of the nose were detected at the h igh dose level in the 14 - day

study but not in the 28 - day

study . Based on the 28 - day

study , the NOAEC for s ys temic toxicity and local effects on the respiratory tract is estimated to be >1.886 mg/1 (489 mg/kg/day) (OECD SIDS, MRID 49388701). In a study compliant with the OECD guideline #413 and GLP, three groups of rats (each of 10 males and 10 females) of the Crl:CD®BR strain were exposed to DMSO, 6 hours a day , 7 day s a week, for 13 weeks using a snout - onl y exposure s ys tem. A fourth group, acting as control, was exposed to air only . An additional 10 male and 10 female rats concurrently exposed at the c ontrol and h igh dose levels were retained following the final exposure for a further 4 weeks to assess the reversibility of any adverse findings. The stud y was designed to compl y with OECD, US EPA OPPTS guidelines and GLP. The mean anal y z ed chamber concentrations of DMSO were 0.310 mg/1, 0.964 mg/1 and 2.783 mg/1 ( equivalent to 80, 2 50, and 722 mg/kg/day) for groups 2 ( l ow dose), 3 ( i ntermediate dose) and 4 ( h igh dose) respectively . Throughout the stud y, the animals were checked twice a day for mortalit y and clinical signs. Bod y weight and food consumption were recorded weekl y. Water consumption was recorded daily . The ey es of all main and recovery group rats were examined once before the start of dosing using a Keeler indirect ophthalmoscope. Similarly , all m ain and r ecovery group rats in groups 1 and 4 were examined during w e ek 13. Functional observations were conducted on 10 male and 10 female rats from each group, pre - treatment, on week 12 and at the end of the recovery period. Vaginal smears were prepared daily from all female rats during weeks 8 and 9 of the study

for as sessment of the oestrus cycle. Blood and urine samples for h ematology , biochemistry

and urinal ysis were collected from all main group rats during week 13 of dosing. All main group animals were killed following 13 weeks of dosing and subject to a detail19

m acroscopic examination. Sperm samples from rats from all groups were assessed for motility , sperm morphology

and number. Histopathological examinations were performed on all scheduled tissues. A high dose group male was sacrificed on humane grounds in week 13 due to the condition of the upper incisors and subsequent weight loss. Treatment - related clinical signs consisted of red staining around the nose of rats exposed to 0.964 and 2.783 mg/1. Rats exposed to 2.783 mg/1 gained less weight over the expo sure period compared with the controls. The trend was reversed during the recovery phase. However, the differences were small ( - 17% in males and - 16% in females compared to controls) and in the absence of any other indications of sy stemic toxicity

probab ly reflected a loss of appetite caused by the mildly

irritant nature of DMSO. Differences between control and treated groups for food and water consumption were minimal and of no toxicological importance. Ophthalmic examination, functional observation b attery , oestrus cycle, sperm anal ys is, h ematology , biochemistry , urinaly sis, macroscopic pathology and organ weights were unremarkable. Treatment - related microscopic changes were found in the nasal passages and phary nx of high dose males and females after treatment for 90 day s, but not in rats from the low and intermediate dose groups. The pathology in the nasal passages comprised of lesions in the inferior ventral medial meatus and an increased degree of eosinophilic inclusions in the olfactory epithelium. Inthe phary nx, prominent goblet cells were present in the majorit y of high dose rats. In rats killed after the recovery period, changes were still evident in the nasal passages of high dose male and females and also in the phary nx of the females. Ac cording to these results, the no adverse effects concentrations (NOAEC) could b e established at 0.964 mg/1 for res piratory tract irritation and 2, 783 mg/1 ( equivalent to 722 mg/kg/day) for sy stemic toxicity (OECD SIDS, MRID 49388701). Neurotoxicity Neurotoxicity

studies were not available for review, however, evidence of potential neurotoxicity

was not observed in the submitted studies. In addition, functional observation battery parameters were not affected following inhalation exposure in rats for 13 weeks (OECD SIDS, MRID 49388701) . Mutagenicity In vitro:20

DMSO was negative for mutagenicity in the reverse mutation assay s in strains TA97, TA98, TA100, TA102, TA 1535, TA1537, and TA1538 at concentrations up to 300, 000 u g/ plate (WHO, 2000). DMSO was mutagenic in Salmonella typhimurium strains TA1537 and TA2637 at concentrations of 0.1 - 0.4 mL/plate and in E. coli WP2uvrA at concentrations of 0.2 - 0.4 mL/plate, with and without metabolic activation (Hakura, et a/., 1993); however, the concentrations were relatively high with some being cy totoxic. Additionally , DMSO was mutagenic in the umu test with Salmonella typhimurium strains TA1535/pSK1002 carrying the umu C - lacZ fusion gene (Nakamura, et al ., 1990). The umu gene expression was detected only at high - dose levels (5% - 15%). The level of p - galactosidase activity , which reflects umu expression was increased in a concentration - related pattern (121 - 313 units compared to 90 units in control). There was a dose - related increase in gene conversions at ce rtain loci of log phase cells of the D4 strain of Saccharomyces cerevisiae following exposure to dose levels up to 1.4 M for 4 hours at 37°C. The gene conversions were attributed to the metabolic conversion of DMSO to a geneticall y active compound by cy tochrome P - 450 mixed function oxidation reactions. In a study performed with a method comparable to the OECD Guide - line no. 471, DMSO was tested in two separate laboratories in the gene mutations assay in bacteria. Concentrations of DMSO (100, 333, 10 00, 3333, and 10,000 ug) were added to overnight culture s of Salmonella ty phimurium strains TA97, TA98, TA100, TA1535, TA1537, and S - 9 mix or buffer were incubated without shaking for 20 minutes. The top agar was added and the contents of the tubes were m ixed and poured onto the surfaces of petri dishes. His+ (histidine dependent) colonies arising on plates were machine - counted after two day s incubation. Initial testing was without metabolic activation, with 10% rat liver S - 9, or with 10% hamster liver S - 9. After a negative result was obtained, DMSO was retested without S - 9 and with 30% S - 9 from rat and hamster. Positive controls were sodium azide (TA 1535 and TA100), 9 - aminoacridine or ICR - 191 (TA 97 and TA1537) and 4 - nitro - o - phen yl enediamine (TA98) wi thout S9 and 2 - aminoanthracene (all strains) with S9: The positive control chemicals induced a significant increase of the revertant frequency in all tester strains, either with or without metabolic activation. DMSO was negative, in the presence and absen ce of metabolic activation, in all tester strains (OECD SIDS , MRID 49388701). In addition DMSO was not mutagenic in the Salmonella typhimurium mutagenicity assay using the standard plate incorporation protocol, both in the presence and absence of metabol ic activation (OECD SIDS , MRID 49388701). DMSO did not induce dominant lethality in male rats injected intraperitoneally at a dose level of 1 mL/kg/day

for 10 weeks (Sheu and Green, 1979 as cited in MRID 49388701 ), or in21

Swiss mice injected with doses up to 10 g/kg twice at intervals of a bout 20 hours (Aravindakshan, et al ., 1975 as cited in MRID 49388701 ). In vivo: In a study performed with a method comparable to the OECD Guide - line no. 473, DMSO was tested with Chinese hamster ovary (CHO) cells in culture at concentrations of 0, 499, 1,500 and 4,990 u g/mL , in the presence and absence of liver microsomal fraction (S9 ) from Aroclor 1254 - induced male Sprague - Dawley rats, to determine its in vitro cy togenetic potential. Medium control was used with each assay with or without S9. Mitomycin C was used in the experiments without metabolic activation, and cyclophosphamide was used in the experiments with activation as positive controls. The positive control chemicals induced a significant increase of the frequency of the chromosomal aberrations. DMSO did not induce cell toxicity or cell cycle delay , and did not induce an increase in the incidence of chromosomal aberrations (OECD SIDS, MRID 49388701). In an in vivo micronucleus assay performed according to the OECD guideline no. 474 and GLP, groups of 6 male and 6 female Han Wistar rats received 5 dail y i.p. injections at the dose levels of 0, 200, 1,000 and 5,000 mg DMSO/kg bw/d ay . The animals were sacrificed 24 h ours after the last dosing and bone marrow collected and evaluated for the presence of micronucleated polychromatic ery

throcy tes. The negative and positive controls in the study were purified water and cyclophosphamide, respectivel y. There was no increas e in the incidence of micronuclei in the polychromatic ery

throcy tes of the bone marrow of male and female Han Wistar treated with DMSO as compared to concurrent controls (OECD SIDS, MRID 49388701). Immunotoxicity As part of a 13 - week inhalation toxicity study , four satellite groups of Sprague - Dawley rats (5/sex/group) were exposed snout - only , 6 h ours /day , 7 d ay s /week, for 28 day s to mean anal yz ed chamber concentrations of 0, 0.310, 0.964 and 2.783 mg /1 ( equivalent to 0, 80, 250 and 722 mg/kg/day) DMSO . To act as a positive control, a further group of satellite animals were injected i.p. with cyclophosphamide at a dose of 50 mg/kg 2 day s prior to termination. The study was designed to compl y with OECD/US EPA OPPTS guidelines and GLP regulations. Sheep Red Blood Cell (SRBC) - specific antibod y secreting cells (ASCs) were enumerated using a modification of the Jerne Plaque Forming Cell (PFC) assay .22

Inhalation exposure to DMSO led to an apparent increase in the number of antibody secreting cells in male rats, compared with controls. Changes in the total number of antibody secreting cells in the spleen (PFC/spleen) and also changes in the relative numbers of antibody secreting cells, associated with changes in other spleen cell populations (PFC/106 cells) were measured. Since DMSO increased both PFC/106 cells and PFC/spleen in male rats, the effect is likely to be a genuine enhancement, not simply a broad spectrum increase in spleen cell numbers. Itis worth noting that there was a high degree of i nter - animal variation in the PFC response. This is almost certainl y linked to the genetic background of the Sprague - Dawley rats, which are an outbred population. The adaptive immune response is known to have strong genetic linkage and work from other sources has shown that while outbred species are popular models in toxicology they have an inherent variability in immune responses. Based in the high variability observed in male rats, the lack of effects in female rats and the lack of effect on the other e nd - points relevant for immunotoxicity

(spleen and thy

mus weights, ly

mphocy te count) reported in the 13 - week stud y, these data are of doubtful toxicological significance . The NOAEL was 2.783 mg /1 ( equivalent to 722 mg/kg/day), the highest dose tested (OEC D SIDS, MRID 49388701) .

Metabolism Rat:

35 S - DMSO (0.55 g/kg ( equivalent to 550 mg/kg) , approximately 0.5 uCi) was administered to male Sprague - Dawley

rats orall y or dermally, the animals were killed at various times and the tissues were assay ed for total radioactivity . After an oral dose the concentration of radioactivity in plasma was maximal ( equivalent to 600 u g equivalent DMSO/mL ) between 0.5 and 4 h ours, then declined to equivalent to 70 u g eq. DMSO/mL at 24 h ours, 0.5 h ours after administrati on. Kidney , spleen, lung, heart and testes appeared to have somewhat higher levels ( equivalent to 60 to 70% of the plasma concentration) than liver, fat, small intestine, brain, skeletal muscle and red cells ( equivalent to 50%). Concentrations in the tes tes, brain, skeletal muscle and heart increased (60 to 85% of the plasma concentration) at 4h ours , but remained virtuall y constant in other tissues. Levels had declined to minimal values in plasma and all tissues after 24 h ours . The ratio of DMSO 2 to DMSO in rats 4 h ours after oral administration of 35 S - DMSO was found to be virtually constant in liver, testes, kidney , spleen, small intestine, heart and plasma, averaging about 6.5% (range of 4.1 - 10.6% for23

tissues of 2 rats). Thus, the major part, at least, of radioactivity present in tissues seems to be represented b y DMSO. After dermal administration, tissue concentrations of radioactivity were also appreciable after 0.5 h ( equivalent to 300 ug eq. DMSO/mL ), but were somewhat lower than after an oral dose. The concentration of radioactivity in plasma was maximal ( equivalent to 600 ug eq. DMSO/mL ) 2 h ours after the administration, and then declined to equivalent to 70 ug eq. DMSO/mL at 24 h ours . At 0.5 h ours , levels in the spleen, liver and lungs were higher ( equivalent to 70% of the plasma concentration) than the other tissues (40 to 60%). Concentrations in the liver, testes, kidney , spleen, brain, lungs, skeletal muscle, heart, plasma and red cells increased after 4 h ours to values comparable t o those after an oral dose. Levels in the fat and small intestine remained virtually constant. All tissue concentrations had declined to minimal values after 24 h ours (OECD SIDS, MRID 49388701). Monkey : The pharmacokinetics of DMSO in rhesus monkeys w as studied during and after a 14 - day oral administration of a dose l evel of 3 g/kg as a 50% solution in water. DMSO and DMSO 2 were measured in serum, urine and feces b y gas - liquid chromatograph y. The absorption of DMSO was rapid. After the first adminis tration, an average peak serum concentration of 2.3 mg/mL was observed after about 4 hour s , which declined relatively rapidl y to about 0.95 mg/mL after 24 hour s . DMSO half - life was 16 hour and its elimination rate constant was about 4% per hour. DMSO 2 became detectable in serum after about 2 h ours , rose slowly and reached about 0.18 mg/mL at 24 hour s . With continued daily

oral administration, serum DMSO rose slightly from 0.95 to 1.1 mg/mL on day 2. DMSO and DMSO 2 reached a stead y s tate concentratio n of about 0.9 mg/mL and 0.34 mg/mL after 4 day s, respectivel y. After the last DMSO dose, serum DMSO declined rapidly and was not detected after 72 h ours . The mean DMSO 2 serum concentration declined slowly over the next 96 h ours and trace amounts were det ected at 120 h ours . DMSO 2 half - life was calculated to be about 38 h ours and its elimination rate constant about 2% per h our . About 60% of an orally

administered dose was excreted in the urine as DMSO and about 16% as DMSO 2 . Neither DMSO nor DMSO 2 could be detected in feces. The pulmonary elimination of DMSO was not quantified in this study but was detectable as a sweet smell in the exhaled air (OECD SIDS, MRID 49388701).24

**Table 3 . Toxicology Profile for**  **DMSO Study Type**  **Doses Results**  2 3 - we ek - Oral Toxicity Study - Dog - diet 0, 2.5, 5,10, 20, or 40 g/kg/day (0, 2,500; 5,000; 10,000; 20,000; 40,000 mg/kg/day) NOAEL was not established. LOAEL: 2,500 mg/kg/day based on changes in the lens of the eye . 24 - we ek - Oral Toxicity Study - Rabbit - drinking water 0, 0.5 or 10 g/kg/day (0, 500 or 10,000 mg/kg/day) NOAEL = 0.5 g/kg/day ( 500 mg/kg/day) . LOAEL = 10 g/kg/day ( 1 0, 000 mg/kg/day) based on changes in the optical lenses . 18 - Month - Oral Toxicity Study - Rat - Gavage - (5 days/week) 0,1, 3, or 9 mL/kg/day (1100, 3300, or 9900 mg/kg/day) NOAEL = 3 mL/kg/day (3300 mg/kg/day) . LOAEL = 9 mL/kg/day ( 99 00 mg/kg/day) based on decreased body weight gain and the change in the refractive index of the nuclear region in the eye . 18 and 24 - Months - Oral Toxicity Study - Dog Gavage - (5 day s/ week) 0,1, 3, or 9 mL/kg/day (1100, 3300, or 9900 mg/kg/day) NOAEL was not established. LOAEL = 1 mL/kg/day ( 1100 mg/kg/day) based on ophthalmologic changes. Reproduction/Development al Toxicity Screening - Rat Gavage 0, 100, 300 and 1000 mg/kg/day Maternal NOAEL = 1000 mg/kg/day. Maternal LOAEL was not established. Reproduction NOAEL = 1000 mg/kg/day. Reproduction LOAEL was not established. Developmental NOAEL = 1000 mg/kg/day. Developmental LOAEL was not established. Developmental Toxicity Rabbit - Gavage 0, 100, 300 and 1000 mg/kg/day) Maternal NOAEL = 300 mg/kg/day. Maternal LOAEL = 1000 mg/kg/day based on reduced body weight and food consumption. Developmental NOAEL = 1000 mg/kg/day. Developmental LOAEL was not established. Cancer Initiation/Promotion - Rat 20 mg DMBA - gavage 50 ppm DMSO - drinking water 3 days before or 3 days after for 18 months No increase d tumorigenesis.25

**Study Type Doses**  **Results** Cancer Initiation/Promotion - Rats Derm al - 3 times/week - 34 weeks DMSO (0.02 mL ) Tumors were not observed.

Cancer Initiation/Promotion - Mice Derm al - 3 times/week - 400 days DMBA (applied once only, 20ug in 0.1 mL acetone) DMSO - 0.1 mL (3 times/week - 400 days) No increased tumorigenesis.

6 Month - Derm al Toxicity Study - Rabbit 0,1.5, 2.7, 4.5, or 8.1 mL /kg/day 0, 1,650; 2,970; 4,950 or 8,910 mg/kg/day) S ys temic NOAEL was not established . Sy stemic LOAEL = 1.5 mL/kg (1650 mg/kg/day) based on lenticular changes . 30 days - Derm al Toxicity Study - Rabbit 0, 1 and 5 g/kg (1000 and 5000 mg/kg/day) Sy stemic NOAEL = 1 g/kg (1000 mg/kg/day). Sy stemic LOAEL = 5 g/kg (5000 mg/kg/day) based on ocular changes. 14 Days Inhalation Toxicity - Rat - (range finding study) 0, 0.52, 1.39 and 5.36 mg/1 ( equivalent to 135, 360, and 1390 mg/kg/day) Sy stemic NOAEC was not established. Sy stemic LOAEC = 0.52 mg/1 ( equivalent to 135 mg/kg/day) based on decreased mean body weight gain. 14/28 Days Inhalation Toxicity - Rat - (range finding study) 0, 0.132, 0.507 and 1.886 mg/1 ( equivalent to 34 , 131 , and 489 mg/kg/day). Sy stemic NOAEC = 1.886 mg/1 ( equivalent to 489 mg/kg/day) . Sy stemic LOAEC was not established. 90 Days Inhalation Toxicity - Rat 0, 0.3 , 10, 0.964 and 2.783 mg/1 ( equivalent to 80, 250, and 722 mg/kg/day) Sy stemic NOAEC = 2.783 mg/1 ( equivalent to 722 mg/kg/day) . Sy stemic LOAEC wa s not established. Ames Test 100 - 300 mg/plate Negative.

Dominant lethal assay in Rodents - intraperitoneal injection. 1 mL/kg/day for 10 weeks(rats) 10 g/kg twice at intervals of about 20 hours (mice) Not clastogenic26

**Study Type Doses**  **Results** Sister chromatid exchange assay in CHO cells up to 4, 999 u g/mL +/ - S9 Not clastogenic . Micronucleus assay in rat bone marrow 0,200, 1,000 and 5,000 mg/kg/day Not clastogenic Plaque - Forming Cell Assay

- Mice - Inhalation 0, 0.310, 0.964 and 2.783 mg/1 ( equivalent to 0, 80, 250 and 722 mg/kg/day) NOAEL 2.783 mg /1 ( equivalent to 722 mg/kg/day). LOAEL wa s not established. Metabolism - Rat - Oral and Derm al 0.55 g/kg ( equivalent to 550 mg/kg) DMSO is rapidly absorbed, metabolized and excreted. Excretion was primarily via urine, feces was a minor route. The major metabolite was dimethyl sulfone. Dimethyl sulfide, another metabolite, is eliminated through the breath. It did not bioaccumulate. 14 - Day Metabolism Rhesus Monkey 3 g/kg/day ( equivalent to 300 0 mg/kg/day) DMSO is rapidly absorbed, metabolized and excreted. Excretion was primarily via feces . The major metabolite wa s dimethyl sulfone. Dimethyl sulfide, another metabolite, is eliminated through the breath. It did not bioaccumulate. **5 .**  **Toxicity Endpoint Selection** The available toxicity

studies indicate that DMSO has low toxicity . These data demonstrated effects at 1 1 00 mg/kg/day (above the limit dose) and no toxicity at doses up to 5 00 mg/kg/day . Although, the lowest NOAELin the dat abase is 300 mg/kg/day in a developmental study in rabbits , it was not selected because of the dose spread , as effects were no t observed in rabbits treated with 500 mg/kg/day in the 24 week oral toxicity study . Therefore, since no endpoint of concern was identified for DMSO , a qualitative risk assessment is appropriate. **6 .**  **Special Considerations for Infants and Childre n**  Section 408 of the FFDCA provides that EPA shall apply an additional margin of safet y for infants and children in the case of threshold effects to account for prenatal and post natal toxicity27

and the completeness of the database on toxicity and exposure un less EPA determines that a different margin of safety will be safe for infants and children. EPA concludes that the FQPA safet y factor could be reduced to 1X for DMSO for the following reasons: i. The database is considered complete for FQPA assessment b ased on the toxicity information on DMSO . The following acceptable studies are available: Subchronic (rat , do g , rabbit, monkey) Chronic (rat, dog, monkey) Reproduction (rat) Developmental toxicity

study (rat , rabbit ) ii. F etal susceptibility was not observed in an y of the reproduction/devel opmental toxicity studies with rats or rabbits administered DMSO . iii. Neurotoxicity

studies were not available for review. However, signs of potential neurotoxicity

were not observed in an y of the submitted studies with DMSO . EPA concluded that neither a neurotoxicity nor a developmental neurotoxicity study is required at this time. iv. An immunotoxicity study was available for review. In a Jerne Plaque Forming Cell (PFC) assay , the total number of antibody secreting cells in the spleen and the relative numbers of antibod y secreting cells were increased. However, this effect is not considered adverse. Therefore, EPA concluded that the immunotoxicity study is not required at this time. Taking into consideration the available information, EPA concludes the additional 10X FQPA safety

factor can be reduced to 1X.

**7 .**  **Exposure Assessment** 28

In evaluating dietary

exposure to DMSO , EPA considered exposure under the petitioned for exemptions from the requirement of a tolerance. EPA assessed dietary exposures from DMSO in food as follows: **Dietary Exposure** Dietary exposure can occur from eating foods containing residues of DMSO . Because no hazard endpoint of concern was identified for the acute and chronic dietary assessment (food and drinking water), a quantitative dietary exposure risk assessment was not conducted. **Residential Exposure (non -**  **dietary exposure)** The term “residential exposure” is used in this document to refer to non - occupational, non dietary exposure (e.g., textiles (clothing and diapers), carpets, swimming pools, and hard surface disinfection on walls, floors, tables). DMSO is used as an inert ingredient in pesticide products that could result in short - and intermediate - term residential exposure. However, based on the lack of toxicity , a quantitative exposure assessment from residential exposures was not performed. **8 .**  **Aggregate Risk** Based on the lack of an y endpoints of concern, EPA concludes that there is a reasonable certaint y that no harm will result to the general population or to infants and children from aggregate exposure DMSO residues. **9 .**  **Occupational Exposure/Risk Pathway** The Agency

believes that establishing the tolerance exemption to include 40 CFR § 180.9 2 0 in a proprietary

product will not significantly /measurabl y increase occupational exposure. The change in use pattern would be expanded to include post - emergence use in pestici de products on all pre - harvest food commodities. Based on the regulatory history of DMSO and no new toxicological information to indicate otherwise, it is not necessary to quantitatively assess occupational exposure.29

**10 .**  **Cumulative Exposure**  **:** Section 408(b)(2)(D)(v) of FFDCA requires that, when considering whether to establish, modify , or revoke a tolerance, the EPA consider "available information" concerning the cumulative effects of a particular pesticide's residues and "other substances that have a common mechanism of toxicity ." Unlike other pesticides for which EPA has followed a cumulative risk approach based on a common mechanism of toxicity , EPA has not found DMSO to share a common mechanism of toxicity with an y other substances and DMSO does not appear to produce a toxic metabolite produced b y other substances. An endpoint of concern was not identified for DMSO in various dietary and non - dietary

exposure scenarios. Additionally , the risk to aggregate exposure of DMSO was below the Agen cy ’s concern. For information regarding EPA's efforts to determine which chemicals have a common mechanism of toxicity and to evaluate the cumulative effects of such chemicals, see the policy statements released by EPA's Office of Pesticide Programs conce rning common mechanism determinations and procedures for cumulating effects from substances found to have a common mechanism on EPA's website at http://www.epa.gov/pesticides/cumulative/. **11 .**  **Enviro n**  **mental Fate** Detailed information regarding environmental fate can be found in the OECD SIDS Dimethyl Sulfoxide d ocument found in MRID 49388701. Below is a summary of environmental fate reproduced from that document. DMSO is mainly

found in water and soil. In the atmosphere DMSO concentrations are low and associated with aqueous phase. Stability and behavio r are strongl y related to the ph ys ico chemical characteristics of the compartment considered but DMSO can be considered as mobile in soil and relatively stable in water. **Ecotoxicity** Detailed information regarding ecotoxicity can be found in the OECD SIDS Dimethyl Sulfoxide document found in MRID 49388701. Below is a summary of ecotoxicity

reproduced from that document.30

Acute toxicity

studies, carried out for some of them according to guidelines similar to OECD guidelines, reveal 48 - hour EC50's ranging from 24,600 to 58,200 mg/Lfor daphnid ( Daphnia magna ) and 96 - hour L C50's ranging from 32,300 to 43,000 mg/L for fish according to the species considere d (eg. Ictalurus punctatus, Lepomis cyanellus ). Modeling calculation for algae indicates 96 - hour EC50 value of about 400 mg/L. On this basis DMSO can be considered non toxic for aquatic compartment. **12 .**  **Enviro n**  **mental Justice** Potential areas of environmental justice concerns, to the extent possible, were considered in this human health risk assessment, in accordance with U.S. Executive Order 12898, "Federal Actions to Address Environmental Justice in Minority Populations and Lo w - Income Populations," http://www.eh.doe.gov/oepa/guidance/justice/eo12898.pdf ). As a part of every

pesticide risk assessment, OPP considers a large variet y of consumer subgroups acco rding to well - established procedures. In line with OPP policy , HED estimates risks to population subgroups from pesticide exposures that are based on patterns of that subgroup’s food and water consumption, and activities in and around the home that involv e pesticide use in a residential setting. Extensive data on food consumption patterns are compiled by the USDA under the Continuing Survey of Food Intake by

Individuals (CSFII) and are used in pesticide risk assessments for all registered food uses of a p esticide. These data are anal yz ed and categorized by

subgroups based on age, season of the y ear, ethnic group, and region of the country . Additionally , OPP is able to assess dietary exposure to smaller, specialized subgroups and exposure assessments are performed when conditions or circumstances warrant. Whenever appropriate, non - dietary

exposures based on home use of pesticide products and associated risks for adult applicators and for toddlers, youths, and adults entering or playing on treated areas po st - application are evaluated. Further considerations are currentl y in development as OPP has committed resources and expertise to the development of specialized software and models that consider exposure to b ys tanders and farm workers as well as lifest yl e and traditional dietary patterns among specific subgroups. **1 3**  **. Human Studies**  This assessment relies in part on data from studies in which adult human subjects were intentionally

exposed to a pesticide. These studies, listed below, have received the appropriate ethical review for use in risk assessment.31

The PHED Task Force, 1998. The Pesticide Handler Exposure Database (PHED), Version 1.1. Task Force members: Health Canada, U.S. Environmental Protection Agency , the California Department of Pesticid e regulation, and the American Crop Protection Association; released August 1998. **1 4**  **. Risk Characterization** EPA received a petition (PP#I N - 10713) from ISK BioSciences (7470 Auburn Rd., Suite A, Concorde, OH 44077), requesting an exemption from the requir ement of a tolerance for dimethyl sulfoxide (here in referred to DMSO, CAS Reg. No. 67 - 68 - 5 ) in c ycl aniliprole formulations for pre - harvest applications including post - emergence use. DMSO will be used as a diluent in c yc laniliprole formulations (60.02% (w/w) with upper and lower limits of 61.82% and 58.22%, respectivel y) under 40 CFR part 180.920. DMSO has an existing tolerance exemption for use as a pesticidal inert under 40 CFR 180.920 as a solvent or co - solvent for formulations used before crop emerges from soil or prior to formation of edible parts of food plants (EPA, 2006). DMSO has low acute toxicity via the oral and dermal in rats and mice and inhalation route in rats. The acute oral LD50 > 7,920 mg/kg in rats and mice. The acute dermal LD50 > 40,000 mg/kg in rats and mice. The acute inhalation L C50 > 1600 mg/m 3 (~277 mg/kg) in rats. It is a dermal, ey e and gastric irritant in rats and rabbits. It is a sensitizer in guinea pigs. Overall s ys temic toxicity with regard oral and dermal exposure to DMSO is low also. The target organ of toxicity is the ey e. Changes in the ey es, such as refractile changes in the lens and lens composition are seen in various animals at doses above the limit dose (1000 mg/kg/day). Sy ste mic t oxicity

is not observed following exposure to DMSO up to 1000 mg/kg/day , the limit dose, in the subchronic, chronic or reproduction/developmental toxicity studies via oral, dermal or inhalation exposures in rats, dogs and rabbits . DMSO is not expected to be carcinogenic based on the lack of mutagenicity and the lack of tumor formation in cancer initiation/promotion studies. It is not neurotoxic nor immunotoxic. Toxicity via the inhalation route of exposure is limited to portal of entry effects at 2.783 mg/1 (equivalent to 722 mg/kg/day). In the rat and monkey , DMSO administered via the oral and/or dermal route is rapidly absorbed, metabolized and excreted. Excretion is primarily via urine, feces was a minor route in32

the rat only . The major metabolite was dimethyl sulfone. Dimethyl sulfide, another metabolite, is eliminated through the breath. There is no bioaccumulation. The FQPA SF was reduced to 1X. Available toxicity data on DMSO demonstrated no adverse effects at doses up to 1,100 mg/kg/day . Therefore, since there are no endpoints of concern for DMSO a qualitative dietary , non - dietary and occupational exposure risk assessment s were appropriate. DMSO has low toxicity

to aquatic organisms. Also, it is not expected to be toxic to mammals. Itis not expected to persist in the environment and does not bioaccumulate. It i s biodegradable. Potential areas of environmental justice concerns, to the ex tent possible, were considered in this human health risk assessment, in accordance with U.S. Executive Order 12898, "Federal Actions to Address Environmental Justice in Minority Populations and Low - Income Populations," http://www.eh.doe.gov/oepa/guidance/justice/eo12898.pdf . Taking into consideration all available information on DMSO (CAS Reg. No. 67 - 68 - 5), EPA concludes that there is a reasonable certainty that no harm will result to the general population or to infants and children from aggregate exposure to DMSO. Therefore, the establishment of an exemption from tolerance under 40 CFR 180.920 for residues of DMSO for use as an inert ingredient (diluent) at a level not to exceed 6 2% in pesticide formulations used before crop emerges from soil or prior to formation of edible parts of food plants; for pesticide formulations used after crop emerges but before harvest, provided that the potential for increased residues of the formulati on’s active ingredient(s) in or on food commodities has been assessed, can be considered safe under section 408 of the FFDCA.33

**REFERENCES** MRID 49388701 Freeman, E. (20 14 ) Compilation of References for the Petition for Dimethyl Sulfoxide (DMSO) (CAS RN 67 - 68 - 5) as a Pesticide Inert Ingredient in Non - Food Use Formulations and For Exemptions from the Requirement of a Tolerance in Accordance with 40 CFR §180.920 for Post - Emergence Pre - Harvest use in the Cyclaniliprole Formulations .