Document ID: EPA-HQ-OPP-2018-0150-0002
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2018-05-18T04:00Z

EPA REGISTRATION DIVISION COMPANY NOTICE OF FILING FOR PESTICIDE PETITIONS PUBLISHED IN THE FEDERAL REGISTER  

EPA Registration Division contact: [P.V. Shah; 703-308-1846]

INSTRUCTIONS:  Please utilize this outline in preparing the pesticide petition.  In cases where the outline element does not apply, please insert "NA-Remove" and maintain the outline. Please do not change the margins, font, or format in your pesticide petition. Simply replace the instructions that appear in green, i.e., "[insert company name]," with the information specific to your action.

TEMPLATE:

[Bayer CropScience Biologics GmbH]

IN-11085

	EPA has received a pesticide petition (IN-11085) from [Bayer CropScience Biologics GmbH], [c/o SciReg, Inc., 12733 Director's Loop, Woodbridge, VA 22192] requesting, pursuant to section 408(d) of the Federal Food, Drug, and Cosmetic Act (FFDCA), 21 U.S.C. 346a(d), to amend 40 CFR part 180

(Options (pick one)
   
To establish an exemption from the requirement of a tolerance for [titanium dioxide (CAS Reg. No. 13463-67-7) under 40 CFR 180.920] in or on the raw agricultural commodity.  EPA has determined that the petition contains data or information regarding the elements set forth in section 408 (d)(2) of  FDDCA; however, EPA has not fully evaluated the sufficiency of the submitted data at this time or whether the data supports granting of the petition. Additional data may be needed before EPA rules on the petition. 

A. Residue Chemistry

	1. Plant metabolism. [NA-remove]

	2. Analytical method. [NA-remove]

	3. Magnitude of residues. [NA-remove]

B. Toxicological Profile

	1. Acute toxicity.  [Titanium dioxide is classified as Toxicity Category IV for acute oral and inhalation toxicity and primary dermal irritation, and Toxicity Category III for primary eye irritation. Titanium dioxide is not a dermal sensitizer. No studies on the acute dermal toxicity of titanium dioxide were located; however, no toxicity via the dermal route of exposure is expected because, as an insoluble, solid material, titanium dioxide is not adsorbed via the skin. 

Acute oral toxicity

In an acute oral toxicity study, a single dose of ultrafine particle type C (uf-C) titanium dioxide (80:20 ratio of anatase:rutile; particle size 0.140 um) suspended in water was administered by oral gavage to one female Crl:CD(SD) rat each at a dose of 175, 550, or 1750 mg/kg and to three females at a dose of 5,000 mg/kg. The rats were observed for mortality, clinical signs of toxicity, and body weight effects for 14 days. A complete necropsy was performed on each rat at the time of sacrifice. There were no mortalities. The only clinical sign observed was grey feces. There were no effects on body weights nor were there any gross findings at necropsy. The oral LD50 of uf-C titanium dioxide was determined to be greater than 5,000 mg/kg.  

In another acute oral toxicity study, ultrafine titanium dioxide particles (0.155 um; > 99% pure) suspended in 0.5% hydroxypropyl methylcellulose K4M were administered by oral gavage to nine male and eight female CD-1(ICR) mice at a dose of 5,000 mg/kg. Mice were sacrificed two weeks later. All mice survived the 14 day period. Body weights and liver, spleen, and kidney weights were unaffected. None of the serum parameters evaluating liver toxicity, nephrotoxicity, and cardiac toxicity were affected by treatment. The livers of male and female mice examined microscopically had hydroscopic degeneration around the central vein and spotty hepatocytic necrosis. Upon microscopic examination, the heart, lung, testes, ovary, and spleen had no findings. The LD50 was determined to be greater than 5,000 mg/kg.

In a limit test conducted according to OECD Guideline 401, the acute oral LD50 was greater than 2,000 mg/kg bw in both sexes of rats.

Acute Inhalation Toxicity

In an acute inhalation study, groups of Crl:CD(SD)IGS BR male rats were intratracheally instilled with multiple formulations of titanium dioxide at doses of 0, 2, or 10 mg/kg. The particles were suspended in saline. Twenty-four hours, one week, and 1 and 3 months after instillation, eight rats/time point were sacrificed. The lungs of four rats/time point were assessed using bronchoalveolar biomarkers and the lungs of the other four rats were examined for histopathology/cell proliferation. The numbers of cells recovered by bronchoalveolar lavage from the lungs of the titanium dioxide exposed groups were not significantly different from the controls at any time point after exposure. At 2 mg/kg, H-11,567 and H-11,569 produced a short-term pulmonary inflammatory response as shown by an increase in the numbers and percentage of bronchoaveolar lavage neutrophils at the 24 hour time point. The same inflammatory response was also observed at 24 hours in all groups exposed to 10 mg/kg. Exposures to 10 mg/kg H-l1,567 and H-l1,569 produced a more sustained lung inflammatory response which persisted to the one month time point for H-11,569 and to three months for H-11,567. Examination of the bronchoaveolar lavage fluid from the H-11,565 and H-11,566 groups treated at 10 mg/kg revealed an increase in lactate dehydrogenase values at the 24 hour time point that were not observed at the later time points. Exposures to H-11,569 produced significant increases in lactate dehydrogenase values at the one week time point. The H-11,567 formulation produced a sustained increase in this parameter through the one month time point. Transient significant increase in bronchoaveloar fluid microprotein values were observed in the lungs of 10 mg/kg H-11,566 and H-11,568 at the 24 hour time point, but did not persist. H-11,569 produced a significant increase in microprotein at one week. H-11,567 caused sustained increased in lung permeability endpoints through one week post exposure. No increase in alkaline phosphatase values or in BrdU lung cell proliferation rates in parenchymal cells was observed.

Microscopic examination of the lungs of the rats instilled with the titanium dioxide formulations revealed no significant morphological differences between the formulations. The lung tissues had particle-laden macrophages at bronchoalveolar junctions following instillation. There were no other findings. The NOAEL was determined to be 10 mg/kg.

In another acute inhalation toxicity study, six male ChR-CD rats were nose-only exposed to 4.99 and 6.82 mg/L titanium dioxide for four hours. Rats were observed and weighed daily for 14 days. No mortality was observed. Immediately following exposure, the rats exposed to 6.82 mg/L titanium dioxide displayed irregular respiration, gasping, lethargy, red eye discharge, and nasal discharge. No remarkable clinical signs were noted at the 4.99 mg/L dose level. Both test groups showed slight weight loss between days 1 to 2 with subsequent normal weight gain from days 3 to 14. The LC50 was determined to be greater than 6.82 mg/L.

The acute inhalation LC50 values in male rats exposed to titanium dioxide powders were greater than 3.43 mg/L (particle size < 3.5 um was 56%, MMAD 3.2 um) and greater than 5.09 mg/L (particle size < 3.5 um was 20%, MMAD 7.0 um). No mortality, body weight changes, or clinical signs were observed. Gross pathology revealed mottled lungs in 2/5 males and 3/5 females exposed to titanium dioxide (particle size < 3.5 um was 56%), as well as pale lungs in 3/8 males and 1/5 females exposed to titanium dioxide (particle size < 3.5 um was 20%).

Primary Eye Irritation

In an acute eye irritation study, uf-C titanium dioxide (80:20 ratio of anatase:rutile; particle size ~ 0.140 um) was evaluated for eye irritation in New Zealand white rabbits according to OCSPP 870.2400. Approximately 57 mg of uf-C titanium dioxide was instilled into one eye of each animal. The eye remained unwashed. One animal was treated initially. No severe irritation occurred, so an additional two rabbits were treated. The eyes were evaluated at 1, 24, 48, and 72 hours following instillation of the test material. The test material caused conjuctival redness (score 1 or 2) for all three rabbits. There was no corneal involvement as determined by fluorescent staining. The treated eyes were normal at 24 or 48 hours after instillation.  

An acute eye irritation/corrosion test was performed according to OECD Guideline 405. Conjunctival redness (score of 1 or 2) was observed at the 1 and 24 hour examinations. The treated eyes of three rabbits were normal by 24 or 48 hours after instillation of the test substance. Based on these results, titanium dioxide was not irritating to the eye of rabbits.

In two other eye irritation studies performed according to OECD Guideline 405, the results also showed that the test substance was not irritating to the eye of rabbits.

Primary Dermal Irritation 

In an acute dermal irritation study, uf-C titanium dioxide (80:20 ratio of anatase:rutile; particle size ~ 0.140 um) was evaluated for dermal irritation in new Zealand white rabbits according to OECD Guideline 404. A single dose of 0.5 g uf-C titanium dioxide was applied to the shaved back after being moistened with 0.25 mL of distilled water. The application area was covered with a 2-ply square gauze pad and held in place with non-irritating tape and covered with porous tape. After a four hour exposure period, the test substance was removed with warm water and soap and the sites were evaluated by the Draize scoring system at 1, 24, 48, and 72 hours after removal. No skin irritation was observed during the study; therefore, uf-C titanium dioxide was considered non-irritating to the skin.  

In another skin irritation assay performed in three rabbits/sex according to OECD Guideline 404, slight erythema occurred in 2/6 animals at 1 hour, 3/6 animals at 24 hours, and 1/6 animals both at 48 and 72 hours. In addition, mild erythema occurred in 1/6 animals at 1 hour. No edema was observed. Titanium dioxide was considered to be a non-irritant in this study.
 
Dermal Sensitization

In a skin sensitization study, uf-C titanium dioxide (80:20 ratio of anatase:rutile; particle size ~ 0.140 um) was evaluated for dermal sensitization in the local lymph node assay in mice according to OECD Guideline 429. Five groups of five female CBA/JHsd mice were dosed for three consecutive days with 0% (vehicle), 5%, 25%, 50%, or 100% titanium dioxide. A positive control (hexylcinnamaldehyde) group was also tested. On day five, 3H-thymidine was injected into the tail vein of each mouse, and the mice were sacrificed five hours later. 

Cell proliferation in the draining auricular lymph nodes of the mice was evaluated. 
A stimulation index (SI) was derived for each group. A response was considered positive 
if the SI was >= 3.0. SIs of less than 3.0 were obtained for the titanium dioxide treated groups. The positive control produced the expected positive response. Therefore, uf-C titanium dioxide was not considered a dermal sensitizer.  

A Buehler test was performed with titanium dioxide using guinea pigs in accordance with OECD Guideline 406. Sensitization reactions were not observed in any of the 20 animals treated with titanium dioxide at both 24 and 48 hours after the challenge application. No clinical signs of toxicity were observed.]

	2. Genotoxicty. [Results of mutagenicity testing with titanium dioxide were mixed; however, the majority of the results were negative. In addition, EPA concluded that, overall, the studies reviewed in support of the tolerance exemption under 40 CFR 180.1195 demonstrate that titanium dioxide is not mutagenic.  

Bacterial Reverse Mutation Test

A bacterial reverse mutation test was conducted with titanium dioxide according to OECD Guideline 471. Titanium dioxide was tested with the standard plate incorporation method in S. typhimurium strains TA98, TA100, TA1535, and TA1537 and E. coli strain WP2uvrA, both in the presence and absence of metabolic activation. Dosage levels tested 
were 100, 200, 500, 1,000, 2,500, and 5,000 ug/plate. Titanium dioxide did not induce any significant, reproducible increases in the observed number of revertant colonies in any of the tester strains, either in the presence or absence of metabolic activation.  

In a bacterial reverse mutation test, uf-C titanium dioxide (80:20 ratio of anatase:rutile; particle size ~ 0.140 μm) was tested using the plate incorporation method. Titanium dioxide was tested in Salmonella tester strains TA98, TA100, TA1535, and TA1537 and E. coli strain WP2uvrA, both in the presence and absence of metabolic activation (Arochlor induced rat liver). The vehicle was sterile water. Dosage levels tested were 100, 333, 1,000, and 5,000 μg/plate. The appropriate positive controls were used. uf-C titanium dioxide did not produce a positive response in any of the test strains, either in the presence or absence of metabolic activation.

In Vitro Mammalian Chromosome Aberration Test

In an in vitro mammalian chromosome aberration test, uf-C titanium dioxide (80:20 ratio of anatase:rutile; particle size ~ 0.140 μm) was tested for chromosomal aberrations in vitro in Chinese hamster ovary cells in the presence and absence of metabolic activation (Aroclor-induced rat liver S9). The vehicle was Milli-Q water. Cytogenetic evaluations of structural aberrations and numerical aberrations were conducted in 200 cells in metaphase at 750, 1,250, and 2,500 μg/mL for the four hour non-activated test; at 62.5, 125, and 250 μg/mL for the four hour activated test; and at 25, 50, and 100 μg/mL for the 20 hour non-activated test. The appropriate positive controls were included. uf-C TiO2 did not produce structural or numerical aberrations under the conditions of this study.

Mammalian Erythrocyte Micronucleus Test

A mammalian erythrocyte micronucleus test was conducted with titanium dioxide according to OECD Guideline 474. Male and female Crl:CD1(ICR) mice were administered a single dose of the test material by oral gavage at dose levels of 500, 1,000, and 2,000 mg/kg. Cyclophosphamide was used as the positive control. Animals were sacrificed at 24 and 48 hours after dose administration and the bone marrow collected. Bone marrow was processed and erythrocytes were collected. At least 2,000 
polychromatic erythrocytes (PCE) per animal were scored for the presence of micronuclei. The proportion of PCEs among 1,000 total erythrocytes was determined for each animal. No statistically significant increases in micronucleated PCE frequency were observed in any of the treated mice at either time point.  
  
In an in vitro micronucleus assay, titanium dioxide was tested for its potential to induce micronuclei formation in human whole blood cultures. Blood cultures were treated with 0, 1, 2, 3, 5, 7.5, or 10 uM titanium dioxide. The formation of micronuclei were examined in the blood lymphocytes. Micronuclei were increased at titanium dioxide concentrations of 5 μM and above. When tested in the presence of either boric acid or borax, the titanium dioxide response was significantly reduced.

In another in vitro micronucleus assay, various forms of titanium dioxide in human bronchial epithelial BEAS 2B cells were tested using the cytokinesis-block micronucleus test. Rutile (nanosize) titanium dioxide stated to be 99.5% pure but found to contain < 5% silicon dioxide as a coating (particle size 10 x 40 nm), rutile (fine) titanium dioxide oxide (99.9% pure; particle size < 5 jam) and anatase titanium dioxide nanopowder (99.7% pure; particle size < 25 nm) were tested. Actual particle size was determined by transmission electron microscopy. Concentrations tested were 1 to 100 μg/cm[2], which were selected based on a preliminary cytotoxicity test. Only nanosized anatase induced micronuclei in BEAS 28B cells after 72 hours, but not at 24 and 48 hours. There was no dose response and there was only a doubling at the two doses that were positive for inducing micronuclei.

In Vitro Sister Chromatid Exchange

In an in vitro sister chromatid exchange assay, titanium dioxide was tested for its potential to induce sister-chromatid exchanges (SCE) in human whole blood cultures. Blood cultures were treated with 0, 1, 2, 3, 5, 7.5, or 10 uM titanium dioxide. The formation of SCE was examined in the blood lymphocytes. SCE were increased in a dose-related manner at titanium dioxide concentrations of 2 μM and above. When tested in the presence of either boric acid or borax, the titanium dioxide response was significantly reduced.

In Vitro Comet Assay

In a comet assay, various forms of titanium dioxide in human bronchial epithelial BEAS 2B cells were tested. Rutile (nanosize) titanium dioxide stated to be 99.5% pure but found to contain < 5% silicon dioxide as a coating (particle size 10 x 40 nm), rutile (fine) titanium dioxide oxide (99.9% pure; particle size < 5 nm), and anatase titanium dioxide nanopowder (99.7% pure; particle size < 25 nm) were tested. Actual particle size was determined by transmission electron microscopy. Concentrations tested were 1 to 100 μg/cm[2], which were selected based on a preliminary cytotoxicity test. The comet assay was conducted twice. The titanium dioxide forms were dispersed as agglomerates with an average size ranging from 3.8 pm (fine rutile) to 5.5 μm (nanosized anatase) as determined by transmission electron microscopy. The dispersions also contained nanosized particles. All three forms caused DNA damage in the comet assay. Nanosized anatase and fine rutile showed similar induction of DNA damage. Nanosized rutile was less effective. However, none of the titanium dioxide forms produced a potent response.

In another comet assay, nano and microsized particles of titanium dioxide were used on human alveolar type-II-like epithelial cells (A594 cells). The cells were exposed to 40 and 80 μg/ml of each particle size for four hours. To measure DNA damage as single strand breaks and alkali labile sites, the alkaline version of the comet assay was used. To investigate oxidative DNA lesions mainly oxidized purines, the FormamidoPyrimidine DNA Glycosylase enzyme was added. Thirty-five cells in duplicate were analyzed. DNA damage was measured as percent DNA in the tail. Particle size was determined to be an average of 63 nm for the nanosized titanium dioxide particles and 1 μm for the microsized titanium dioxide particles. A significant increase in DNA damage was observed for both nano and microsized titanium dioxide particles at 80 μg/mL. At 40 μg/mL, both particle sizes caused an increase in DNA damage; however, the increase was only significant with the microsized titanium dioxide particles.

DNA Damage Formation of 8-oxoguanine in Rats
 
In a DNA damage formation of 8-oxoguanine study, female Wistar rats (five per group) were exposed by instillation to a single dose of 0.15, 0.3, 0.6, and 1.2 mg of titanium dioxide P25 (untreated, hydrophilic surface) or titanium dioxide T805 (silanized, hydrophobic surface) particles, suspended in 0.2 mL of physiological saline containing 0.25% lecithin. The controls were instilled with the vehicle or with a single dose of 0.6 mg quartz DQ12 as a positive control. The rats were euthanized 90 days after instillation. Eight micron frozen sections of the left lobe were used for immunohistochemical detection of 8-oxoguanine (8-oxoGua) by a polyclonal antibody in the DNA of individual lung cells. The fluorescence intensity of 500 individual cells per animal was determined. The genotoxic effects in the lung of two types of commercially available titanium dioxide at low doses relevant to the working environment were examined. Although, the companies supplying the titanium dioxide particles claimed their particle size was 20 nm, electron microscopy demonstrated that the particles were aggregated and agglomerated. Sonication was not successful in breaking up the particles to a size of 20 nm. In the quartz-exposed animals, ninety days after exposure, a significant increase in the amount of 8-oxoGua in the DNA of lung cells was detected. In contrast, the amount of 8-oxoGua, as a marker of DNA damage, was at the level of the control in animals exposed to titanium dioxide P25 or titanium dioxide T805.]

	3. Reproductive and developmental toxicity. [Titanium dioxide has low reproductive/developmental toxicity. In addition, EPA concluded that based on the weight of evidence from subchronic and chronic studies in rodents that were reviewed to support the tolerance exemption under 40 CFR 180.1195, titanium dioxide does not present a reproductive hazard.

A prenatal developmental toxicity study with titanium dioxide was conducted according to OECD Guideline 414. CRL rats were exposed to the test substance by oral gavage at dose levels of 0, 100, 300, and 1,000 mg/kg once daily, seven days per week from gestation day 5 through gestation day 19. Clinical observations were made daily. Body weight and food consumption were measured on gestation days 5, 8, 11, 14, 17, and 20. Animals were sacrificed on gestation day 20. Females were examined macroscopically for any structural abnormality or pathological changes. The ovaries and uterus were examined. Fetal evaluations included examination for external, soft tissue, skeletal, and head abnormalities. There were no treatment-related effects on clinical observations, body weights, or food consumption. There were no treatment-related effects in the females or fetuses or significant developmental effects observed at any dose. Therefore, the NOAEL for maternal and developmental toxicity was 1,000 mg/kg. 

In a non-guideline male reproductive parameter evaluation study, male albino rats (six per group) were placed on diets containing 0, 1, and 2% titanium dioxide for a period of 65 days. Body weights were recorded at study initiation and at study termination. Clinical signs of toxicity were monitored. Serum levels of testosterone, nitrite, superoxide dismutase activity, glutathione reductase activity, and malondialdehyde levels were evaluated. The cauda epididymis was minced in saline and examined for sperm motility, sperm cell concentration, sperm abnormalities, and live percent spermatozoa. The testis, seminal vesicles, and liver were fixed for histopathological examination.

Depression, anorexia, and white feces were noted in the titanium dioxide treated groups (incidence not provided). Body weights were statistically significantly decreased in both treated groups compared to the control: 14.6% decrease at the low dose and 14% at the high dose. Sperm motility was depressed at both dosage levels but the decrease was statistically significant at the high dose only. Sperm cell concentration, sperm viability, and serum testosterone levels were statistically decreased and the number of sperm abnormalities were statistically increased at both titanium dioxide concentrations tested. Malondialdehyde levels were unaffected by treatment; however, glutathione reductase and superoxide dismutase activities, and nitrite levels were statistically increased at both dosage levels compared to control levels. 

Macroscopically, the seminal vesicles of the treated rats showed hypertrophy at the low dose and atrophy at the high dose. Microscopically, the lining of the seminal vesicles of all rats fed 1% titanium dioxide had hyperplastic changes with little or absence of secretion "beside edema of trabiculae" while at 2% titanium dioxide, there was a reduction in number and size of the epithelial cell lining of the tubuloaveolar glands. At the low dose, the seminiferous tubules revealed mild spermatogenesis with congested interstitial blood vessels with endotheliosis. Thickened tunica albuginea with degenerated spermatogonial cell layers and spermatocytes with the absence of spermatogenesis were also observed, particularly at the high dose. Microscopic examination of the liver revealed hepatocytic degenerative changes which varied from vacuolar and hydropic degeneration to cell death at the low dose and increased in severity to mainly steatosis of the hepatic cells as the high dose. A NOAEL was not observed in this study. However, a LOAEL was observed at 1% titanium dioxide in the diet which is equivalent to approximately 500 mg/kg/day.

Another reproductive toxicity study in male mice conducted with nano-sized titanium dioxide, resulted in a LOAEL of 500 mg/kg/day and a NOAEL of 200 mg/kg/day based on decreases in the liver, heart and kidneys weights, and a significant increase in serum biochemical parameters such as ALT, ALT/AST, and BUN (P < 0.05). The 500 mg/kg/day group also showed significantly reduced sperm density and motility, increased sperm abnormality, and germ cell apoptosis, but no obvious pathological changes in the liver, kidney, spleen, testis, and epididymis.

Titanium dioxide was investigated in a reproductive and developmental toxicity screening test in rats according to OECD Guideline 421. Titanium dioxide was administered by oral gavage to 10 animals per sex at 0 or 1,000 mg/kg bw/day (limit test) to male rats from two weeks prior to mating, during the mating period, and approximately two weeks post mating, and to female rats from two weeks prior to mating, during the mating period, gestation period, and three days after lactation. During the observation period, there were no dose-related effects on clinical signs, body weights, food consumption, mating, gestation, delivery, organ weights, necropsy, and histopathology in parents. No dose-related changes in clinical signs, body weights, viability index, external malformations, and sex ratios were noted in pups. This study found no indication of any reproductive toxicity in parent animals or developmental toxicity in pups. Therefore, the NOAEL for reproductive and developmental toxicity was 1,000 mg/kg bw/day.]

	4. Subchronic toxicity. [Many repeat-dose oral, dermal, and inhalation studies were conducted in various species (rats, mice, dogs, and hamsters) at very high doses that indicate low toxicity via the oral and dermal routes of exposure for titanium dioxide. No endpoint of concern via the oral route of exposure was identified by EPA from the studies reviewed to support the tolerance exemptions under 40 CFR 180.1195. In a 28-day lung instillation study, one formulation of titanium dioxide had a slight fibrogenic effect and the other formulations caused lung changes consistent with that of a nuisance dust. 

Repeat-dose Oral Toxicity 

In a subchronic oral toxicity study, two groups of 10 male and 10 female rats were administered 0 and 10% (approximately 100,000 ppm) N.F. grade titanium dioxide in their diet for 30 to 34 days. No treatment-related effects were observed in this study.

In a National Cancer Institute (NCI) 90-day oral toxicity study in rats and mice, 10 males and 10 females of each species were administered titanium dioxide at doses of 6,250, 12,500, 25,000, 50,000, and 100,000 ppm. Ten males and ten females of each species also received basal diets. In both species, there were no deaths and dosed animals had mean body weight gains that were comparable to those of the controls. No gross or microscopic pathology was found that could be related to the administration of titanium dioxide in either species at doses up to 100,000 ppm (equal to 5,000 mg/kg/day in rats).

In another 90-day repeat-dose oral toxicity study, albino rats were fed diets containing 0, 1.0, 3.16, and 10% titanium dioxide coated-mica (TCM). No compound related effects on behavior, growth, food consumption, or mortality were observed. Organs and tissues showed neither consistent gross changes nor histopathological changes that could be attributed to dietary administration of TCM.

In a fourth 90-day repeat-dose oral toxicity study, three groups of two dogs were orally administered 0.05, 0.1, and 0.15 g of titanium dioxide. Every five days, the dose was increased by the same amounts. One dog of each group was kept for one month, the other for two months. No toxic effects were seen in the dogs administered titanium dioxide.

In a fifth 90-day repeat repeat-dose toxicity study, pigment grade titanium dioxide was administered by oral gavage to ten male and ten female Crl:CD(SD) rats at doses of 0, 100, 300, and 1,000 mg/kg according to OECD Guideline 408. Animals were observed daily for clinical observations. Body weight and food consumption were measured weekly. Blood and urine were collected at sacrifice for evaluation of hematology and clinical chemistry and urinalysis parameters, respectively. A neurobehavioral examination was conducted during week 13. After 92 or 93 days of dosing, animals were sacrificed and examined macroscopically; tissues were taken for microscopic examination. No mortality was observed. No treatment-related effects were noted in clinical signs, body weights, food consumption, hematology, clinical chemistry, or urinalysis parameters, and macroscopic or microscopic evaluations. The NOAEL was determined to be 1,000 mg/kg.  

In a final 90-day repeat-dose oral toxicity study, pigment grade titanium dioxide was administered by oral gavage to ten male and ten female Crl:CD(SD) rats at doses of 67, 258, and 962 mg/kg according to OECD Guideline 408. Animals were observed daily for clinical observations. Body weight and food consumption were measured weekly. Blood and urine were collected at sacrifice for evaluation of hematology and clinical chemistry and urinalysis parameters, respectively. A neurobehavioral examination was conducted during week 13. After 92 or 93 days of dosing, animals were sacrificed and examined macroscopically; tissues were taken for microscopic examination. No mortality was 
observed. No treatment-related effects were noted in clinical signs, body weights, food consumption, hematology, clinical chemistry, or urinalysis parameters, and macroscopic or microscopic evaluations. The NOAEL was determined to be 962 mg/kg.  

Repeat-dose Dermal Toxicity 

A seven day repeat-dose dermal toxicity study was conducted with titanium dioxide in Balb-c mice. 15% of the titanium dioxide was mixed with Vaseline and lanolin (2:1) to produce an ointment. 0.2 g of this ointment was applied to the ventral skin of the mice once daily for seven days. Animals were sacrificed after seven days and a gross necropsy was conducted. Tissues, including liver, kidney, heart, and lung were collected and examined microscopically. There was no mortality during the study and no treatment-related clinical observations were noted. No treatment-related observations were noted in the tissues examined microscopically.  

A 14-day repeat-dose dermal toxicity study was conducted with titanium dioxide in Wistar rats. Rats were administered the test material topically at dose levels of 14, 28, 42, and 56 mg/kg once daily for 14 days. Animals were sacrificed after 14 days and skin was collected and examined microscopically. No treatment-related observations were noted, including in the skin evaluations.  

A 56-day repeat-dose dermal toxicity study was conducted with titanium dioxide in Hairless Wistar Yagi rats. 4 mg/cm[2] of the test material was applied topically to the dorsal skin of the rats once a day for 56 days. After 2, 4, and 8 weeks, skin samples were taken from the treated skin and evaluated microscopically. Animals were sacrificed after 56 days and a gross necropsy was conducted. Tissues, including lung, liver, spleen, and kidney, were collected and examined microscopically. No treatment-related observations were noted, including in the skin evaluations.  

Repeat-dose Inhalation Toxicity

In a 28-day inhalation toxicity study, twenty-five male Chr:CD rats/group were exposed to one of six formulations (85 to 99%) of titanium dioxide for six hours per day, five days a week for a total of 20 exposures. A control group of 15 male rats was exposed to air only. Ten rats per group were subjected to a clinical laboratory evaluation (hematology, clinical chemistry, and urinalysis) following the 20th exposure and five rats per group were subjected to a clinical laboratory evaluation after a 14 day recovery period. Urine aminolevulinic acid concentrations and blood aminolevulinic acid dehydrase activity were also determined. Histopathology was conducted on the tissues of five rats per group on the last day of exposure and at 14 days, and 3, 6, and 12 months after exposure. Weights of the heart, liver, lungs, spleen, testis, and thymus were recorded. The exposure concentrations were determined gravimetrically. The particle size (mass medium diameter) ranged from 1.3 to 1.8 μm. 

Body weight was reduced in the H-11,567 titanium dioxide formulation group but was similar to the control group's body weight after the recovery period. No significant effects were observed with the other titanium dioxide formulations. The only notable clinical finding was lung noises in two rats (days 8 and 11) in the H-11,567 treated group; however, this finding was absent by study day 15. Four mortalities occurred during the study: two in the control group, one in the H-11,565 treated group, and one in the 
H-11,566 treated group. 

Neutrophils were significantly elevated in the H-11,564 (high dose only), H-11,566, 
H-11,567, H-11,568, and H-11,569 treated groups. These values remained elevated after the 14 day recovery period. 

The mean absolute and relative lung weights of rats exposed to the six titanium dioxide formulations, except for the low dose, H-11,565, group, were statistically elevated when compared to the control group. The lung weights remained elevated after the 4 week recovery period through the 12 month recovery period, except for the H-11,564 treated rats at both concentrations tested. The mean lung weight was still elevated at the high dose when compared to the control group.

At the end of the four-week exposure period, the inhaled dust particles of all test groups
were heavily concentrated with the bronchioles, alveolar ducts, and alveoli. The dust
particles were mostly phagocytized by the intra-alveolar macrophages. In addition, dust cell accumulation and alveolar cell hyperplasia were observed in all rats exposed to 
H-11,564 at both test concentrations. A marked reduction in dust cells and alveolar cell proliferation was observed at the end of the 14 day and three month recovery periods; however, dust cells were still observed at the six and twelve month recovery periods.

Rats exposed to H-11,567 and H-11,569 exhibited hyperplasia of the type II pneumocytes
and disintegration of dust cells at the three month recovery period. The hyperplasia
persisted to the 12 month recovery period. Light collagen deposition which represents a
fibrogenic response was observed in the H-11,567 treated group. H-11,564, H-11,565, 
H-11,566, H-11,568, and H-11,569 groups exhibited slight alveolar cell hyperplasia at the end of the four week exposure.

It was concluded that except for H-11,567, which exhibited a slight fibrogenic
response, the titanium dioxide formulations caused lung changes consistent with a nuisance dust.

In another repeat-dose inhalation toxicity study, female rats were exposed for six hours/day, five days/week for four weeks by nose-only inhalation to 0, 0.1, 1.0, or 10 mg/m[3] of titanium dioxide (MMAD: 1.3 um, GSD: 2.6, respirable fraction was not available) and the lung burdens were determined at one week after the end of the exposure. The lungs were evaluated by analysis of bronchoalveolar lavage fluid (BALF) at 1, 8, and 24 weeks after the end of the exposure and by histopathology at 24 weeks. With lung burdens up to 420 ug/g lung, titanium dioxide elicited no changes in BALF parameters at any time after exposure, nor were any histopahological findings observed. Therefore, the NOAEC was considered to be 10 mg/m[3].

In another repeat-dose inhalation toxicity study (whole body), 65 female rats were exposed to 0, 10, 50, or 250 mg/m[3] of titanium dioxide (rutile) for six hours/day, five days/week for 13 weeks with recovery groups held for an additional 4, 13, 26, or 52 weeks post-exposure (MMAD: 1.44 um, Geometric SD (GSD): 1.71, respirable fraction was not available). No deaths occurred during the exposure period. Lung and lung-associated lymph node burdens of titanium dioxide increased in a concentration-dependent manner. Pulmonary overload was achieved in rats at 50 and 250 mg/m[3]. Inflammation was seen at 50 and 250 mg/m[3] by the evidence of increased numbers of macrophages and neutrophils and incidences of soluble inflammation markers. Inflammatory responses remained elevated throughout the entire post-exposure recovery period at 250 mg/m[3]. Pulmonary lesions with progressive epithelial and fibroproliferative changes were observed at 250 mg/m[3]. These epithelial changes were also manifested in rats as evidenced by an increase in alveolar cell labeling at 250 mg/m[3] in cell proliferation studies. Based on these results, the NOAEC was determined to be 10 mg/m[3].

A fourth repeat-dose inhalation toxicity study was conducted to examine burdens of titanium dioxide (rutile) in the lung and lung associated lymph nodes and selected lung responses in mice and hamster (73 females per concentration). Animals were exposed to 
0, 10, 50, or 250 mg/m[3] pigmentary titanium dioxide for six hours/day, five days/week for 13 weeks with recovery groups held for an additional 4, 13, 26, or 52 weeks (46 weeks for hamster) post-exposure (MMAD: 1.39 um in mice, 1.36 um in hamster). Pulmonary parameters including inflammation, cytotoxicity, lung cell proliferation, and histopathologic alterations were assessed. Pigmentary titanium dioxide burdens in the lung and lymph nodes increased in a concentration-dependent manner. Inflammation was noted at 50 and 250 mg/m[3], as evidenced by increases in macrophage and neutrophil numbers and in soluble indices of inflammation in BALF. Based on the results, the NOAEC for mice and hamsters was 10 mg/m[3].

In a final repeat-dose inhalation toxicity study, male rats were exposed to aerosols of titanium dioxide (rutile) at concentrations of 0, 25, or 50 mg/m[3]. Rats were exposed by whole-body inhalation seven hours/day, five days/week (25 mg/m[3] exposure for 209 days, and 50 mg/m[3] for 118 days). The MMAD (GSD) for titanium dioxide was 2.1 μm (2.2). There were six time points and generally 12 animals per concentration were used. The lung burdens at the final exposure points were 24 and 17 mg/g for the high and low-dose groups, respectively. The mean lymph node burdens and number of polymorphonuclear cells (PMN) were elevated with increasing exposure. The higher levels of inflammation occurred concurrently with the higher lymph node burdens, after 69 and 139 days in the 50 and 25 mg/m[3] groups, respectively. The predicted averages for percent PMN were, for the high and low dose groups, respectively, 28% and 16%. The mean numbers of alveolar macrophages obtained did not change significantly compared to control animals. Titanium dioxide showed no significant fibrogenic activity. Therefore, the LOAEC for titanium dioxide was considered to be 25 mg/m³ based on the increased mean number of neutrophils with exposure-related lymph-node burdens.]

	5. Chronic toxicity. Titanium dioxide was evaluated for carcinogenicity by oral administration in mice and rats, by inhalation in rats and mice, by intratracheal administration in hamsters, rats, and mice, by subcutaneous injection in rats, and by intraperitoneal administration in mice and rats. Titanium dioxide did not induce tumors by the oral, intraperitoneal, or subcutaneous routes of exposure in rats or mice. In one inhalation study, the incidence of benign and malignant lung tumors was increased in female rats. In another inhalation study, the incidences of lung adenomas were increased in high-dose male and female rats. Cystic keratinizing lesions that were diagnosed as squamous-cell carcinomas but re-evaluated as nonneoplastic pulmonary keratinizing cysts were also observed in the high-dose female rats. Two inhalation studies in rats and one in mice were negative. From these data, the International Agency for Research on Cancer (IARC) concluded that there is sufficient evidence in experimental animals for the carcinogenicity of titanium dioxide; however, there is inadequate evidence in humans for the carcinogenicity of titanium dioxide. The overall conclusion was that titanium dioxide is possibly carcinogenic to humans (Group 2B). However, the IARC also stated that titanium dioxide is a poorly soluble low toxicity (PSLT) particle, which can elicit overloading of lung clearance, chronic inflammation, and lung tumors in rats following prolonged exposure at sufficiently high concentrations of particles. The tumors present in the lungs may have been a localized fibrogenic effect caused by overloading the lungs with ultrafine titanium dioxide particles. In contrast to the results in rats, inhalation effects were not observed in mice and hamsters and may be a rat-specific threshold phenomenon, dependent upon lung overloading at high exposure concentrations and possibly of little relevance to humans. Epidemiological data suggest that there is no carcinogenic effect associated with workplace exposure to titanium dioxide dust.

It is important to note that rats are known to be an extremely sensitive species for developing tumors in the lungs when overloaded with poorly soluble, low toxicity dust particles. Rat lungs process particles very differently compared to larger mammals, such as dogs, primates, or humans. This sensitivity in the lungs has not been observed in other rodent species, such as mice or hamsters; therefore, using the rat model to determine carcinogenicity of titanium dioxide in humans can be misleading, as extrapolation of species-specific data to humans is erroneous.

In addition, the titanium dioxide used in pesticide formulations is considered pigmentary grade, not ultrafine or nanosize. Consequently, the tumors observed from inhalation exposure to ultrafine particles of titanium dioxide are not relevant for assessing exposure to the type of titanium dioxide used in pesticide formulations. This position is consistent with the National Institute of Occupational Health and Safety's (NIOSH) recent assessment that ultrafine, but not fine titanium dioxide, would be considered a "potential occupational carcinogen." The NIOSH Current Intelligence Bulletin "Occupational Exposure to Titanium Dioxide" concludes that "[t]he lung tumors observed in rats after exposure to 250 mg/m[3] [titanium dioxide] were the basis for the original NIOSH designation of TiO2 [titanium dioxide] as a "potential occupational carcinogen." However, because this dose is considered to be significantly higher than currently accepted inhalation toxicology practice, NIOSH concluded that the response at such a high dose should not be used in making its hazard identification." NIOSH concluded that the data is insufficient to classify fine titanium dioxide as a potential occupational carcinogen.

Chronic Oral Toxicity/Carcinogenicity 

In a chronic oral toxicity study, two guinea-pigs, two rabbits, two cats and one dog were fed technical grade titanium dioxide for 390 days. The dog received 9 g/day, rabbits and cats, 3 g/day, and guinea pigs 0.6 g/day. No signs of systemic toxicity were observed in these animals.

In a bioassay of titanium dioxide performed by the NCI, 1979 In U.S. EPA, 2012, groups of 50 rats of each sex and 50 mice of each sex were administered titanium dioxide in the diet at one of two doses, either 25,000 or 50,000 ppm (approximately 1,250 mg/kg/day and 2,500 mg/kg/day, respectively) for 103 weeks and then observed for one additional week. A control group consisted of 50 untreated rats of each sex and 50 untreated mice of each sex.

Administration of the titanium dioxide had no appreciable effect on the mean body weights of rats or mice of either sex. Besides the presence of white feces, no other clinical signs were observed to be related to the administration of titanium dioxide. In the female rats, C-cell adenomas or carcinomas of the thyroid occurred at incidences that were dose related but were not high enough to be of significance. Therefore, these tumors were not considered to be related to the administration of titanium dioxide.

In the male and female mice, no tumors occurred in dosed groups at incidences that were significantly higher than those for corresponding control groups. It was concluded that under the conditions of the bioassay, titanium dioxide was not carcinogenic by the oral route for Fischer 344 rats or B6C3F1 mice.  

Chronic Inhalation Toxicity/Carcinogenicity 

In an inhalation study following OECD Guideline 453, titanium dioxide (rutile) was administered via inhalation (dry aerosol, whole body) to 50 rats/sex/concentration at 0 or 5 mg/m[3] for six hours/day, five days/week for 24 months. The MMAD (GSD) was 1.1 um (1.6) and the respirable fraction was 78%, equivalent to 3.87 +- 0.28 mg/m[3]. A 5% incidence of lung fibrosis was seen in the titanium dioxide-exposed groups. After exposure, minor changes were observed in the cytologic pattern of the BALF. Lymphoid
hyperplasia of the lung-associated lymph nodes was observed in the titanium dioxide-exposed group. Therefore, the LOAEC was considered to be 5 mg/m[3] in rats.

Groups of 50 male and 50 female Sprague-Dawley rats, eight weeks of age, were exposed by inhalation to 0 or 15.95 mg/m[3] titanium dioxide (99.9%; < 0.5 μm; purity unspecified) for six hours per day, five days per week for 12 weeks. The rats were sacrificed at
140 weeks. Average survival was 116 and 113 weeks for control and treated males, and
114 and 120 weeks for control and treated females, respectively. At the end of the study,
39 and 44 control and treated males and 45 and 45 control and treated females, respectively, were still alive. No significant differences in body weights or incidence of
tumors were observed (lung and other respiratory tract tumors were benign; other
neoplasms seen in the lung were metastases from tumors of other sites) between treated
and control groups.

Groups of 100 male and 100 female CD rats, five weeks of age, were exposed by inhalation to 0, 10, 50, or 250 mg/m[3] titanium dioxide (rutile; 99% pure; MMAD, 1.5 to 1.7 μm; ~ 84% of dust particles < 13 μm) for six hours per day, five days per week for two years, at which time all surviving rats were sacrificed. No differences in mortality, body weights, or clinical signs were observed. The incidence of lung tumors was increased in both male and female high-dose rats [adenomas: 2/79, 1/71, 1/75, and 12/77 (P < 0.001) control, low-, mid- and high-dose males, respectively; 0/77, 0/75, 0/74, and 13/74 (P < 0.001) females, respectively; squamous-cell carcinomas: 0/79, 0/71, 0/75, and 1/77 males and 0/77, 1/75, 0/74, and 13/74 (P < 0.001) females, respectively]. One anaplastic carcinoma occurred in a low-dose male (Lee et al., 1985a,b, 1986 In IARC, 2010). Difficulty was experienced in distinguishing between keratinizing squamous metaplasia and squamous-cell carcinomas. The 15 squamous-cell carcinomas reported were reevaluated by Warheit and Frame, who described 11 of the squamous-cell carcinomas as non-neoplastic pulmonary keratinizing cysts.

Groups of 50 male and 50 female SPF Fischer 344 rats, eight weeks of age, were exposed to titanium dioxide (5.0 +- 0.7 mg/m[3]; 99.5% rutile; MMAD, 1.1 μm) for six hour per day, five days per week or air only (control) for 24 months then maintained in clean air for a further 1.5 months. No treatment-related effects on lifespan or causes of death were observed. No differences in tumor development were seen between the groups (one adenoma and one adenocarcinoma in treated animals and two adenomas and one adenocarcinoma in controls).

A group of 100 female Wistar rats, seven weeks of age, were exposed by inhalation to
titanium dioxide (P25; MMAD, 0.80 μm) for 18 hours per day, five days per week for up to 24 months (7.2 mg/m3 for the first four months, then 14.8 mg/m3 for four months, and 
9.4 mg/m3 for 16 months) and then maintained in clean air for a further six months. 
A control group of 220 animals was maintained in clean air. After 30 months, 32/100 treated rats had lung tumors (20 benign squamous-cell tumors, three squamous cell carcinomas, four adenomas, and 13 adenocarcinomas) in contrast to only 1/217 controls (one adenocarcinoma). Lung tumor incidence was 19/100 when benign squamous-cell tumors were not included.

Intratracheal Carcinogenicity 

Groups of 24 and 22 female A/J mice, 20 weeks of age, received a single intratracheal
instillation of a suspension of 0.5 mg titanium dioxide (> 99.9% pure; size unspecified) in
saline or saline alone (control), respectively, and were maintained until 105 weeks of age.
No differences in the incidence of lung tumors (17/24 versus 19/22 controls) or tumor
multiplicity (2.24 +- 1.35 versus 1.42 +- 0.77) were noted.

Groups of 24 or 48 female SPF Wistar (HsdCpb:WU) rats, 8 to 9 weeks of age, received weekly intratracheal instillations under carbon dioxide anesthesia of one of three types of titanium dioxide. The first type was P25: hydrophilic, majority anatase; mean particle size, ~ 0.025 um; density, 3.8 g/mL; specific surface area, 52 m[2]/g. The second type was P805 (AL 90 003-2): hydrophobic; mean particle size, 0.021 um; density, 3.8 g/mL; specific surface area, 32.5 m[2]/g. The third type was AL 23 203-3: hydrophilic, anatase; mean particle size, ~ 0.2 um; density, 3.9 g/mL; specific surface area, 9.9 m[2]/g. The dusts were suspended by ultrasonification in 0.4 mL 0.9% phosphate buffered sodium chloride solution, and Tween 80 was added (1.0%) as a detergent to improve the homogeneity of the dosed suspensions. A control group was maintained untreated. The following table summarizes the experimental groups and the doses instilled. 

Dose levels and tumor results of the intratracheal installation study in mice

Type of 
titanium dioxide
Dose instilled
Lungs with benign tumors (%)[a]
Lungs with malignant tumors (%)[a]
Lungs with total tumors (%)[a]
Lungs with metastases of other tumors (%)
Control
0
0
0
0
0
P25, hydrophilic
5 x 3 mg
5 x 6 mg
10 x 6 mg
21.4
17.4
23.9
31.0
50.0
45.7
52.4
67.4
69.6
14.3
15.2
15.2
P805, AL90, hydrophobic
15 x 0.5 mg
30 x 0.5 mg
0.0
6.7
0.0
0.0
0.0
6.7
9.1
6.7
AL23, anatase, hydrophilic
10 x 6 mg
20 x 6 mg
15.9
38.6
13.6
25.0
29.5
63.6
11.4
2.3
a Primary lung tumor types diagnosed; benign: adenoma, epithelioma; malignant: adenocarcinoma, squamous-cell carcinoma; lungs with one or more malignant tumors may additionally have had benign tumors.

Rats were inspected for clinical signs of morbidity and mortality twice per weekday and once a day on weekends. The experiment was terminated at 30 months unless rats were killed when moribund or diagnosed with a growing subcutaneous tumor. Because of acute toxicity, the number of animals exposed to the hydrophobic titanium dioxide was reduced. After death of the animals and before necropsy of the thoracic and abdominal cavity, lungs were insufflated in situ with formalin via the trachea. In particular, the surface of the lung was inspected and lesions were recorded. Lungs were embedded in paraffin and sections were stained with haematoxylin and eosin. All suspected tumor tissues that were taken from other sites were also examined for histopathological lesions, especially for tumors that might be primary tumors with lung metastases. The above table summarizes the incidences of lung tumors for each group. Statistically significant increases in benign and/or malignant lung tumors were observed with both types of hydrophilic titanium dioxide.

Groups of 24 male and 24 female Syrian golden hamsters, 6 to 7 weeks of age, received
intratracheal instillations of 0 (control) or 3 mg titanium dioxide (particle size: 97% 
< 5 um; 51% < 0.5 μm) in 0.2 mL saline once a week for 15 weeks. The animals were observed until spontaneous death. All control and treated hamsters died by weeks 110 to 120 and 70 to 80, respectively, after the beginning of the experiment. The respiratory tract and other organs with gross lesions were examined histopathologically. No respiratory tract tumors were found in the treated groups compared with two tracheal papillomas that were found in untreated controls.

Subcutaneous Carcinogenicity 

Groups of 20 male and 20 female Sprague-Dawley rats, 13 weeks of age, received a single subcutaneous injection into the flank of 1 mL saline (control) or 30 mg of one of three preparations of titanium dioxide (> 99% pure, coated with antimony trioxide; > 95% pure, coated with aluminium oxide; or > 85% pure, coated with both compounds) in 1 mL saline. All rats were observed until spontaneous death, which occurred as late as 136, 126, 146, and 133 weeks in the control and three titanium dioxide-treated groups, respectively. No tumors were observed at the site of the injection in any group.

Intraperitoneal Carcinogenicity 

Groups of 30 or 32 male Marsh-Buffalo mice, 5 to 6 months of age, received a single
intraperitoneal injection of 0 (control) or 25 mg titanium dioxide (purity, > 98%; manually ground) in 0.25 mL saline. All survivors (10 control and 13 treated mice)
were sacrificed 18 months after treatment. No difference in the incidence of local or distant tumors was observed between treated and control animals.

As part of a large study on various dusts, three groups of female Wistar rats received intraperitoneal injections of titanium dioxide in 2 mL 0.9% saline solution. The first group received a total dose of 90 mg/animal in five weekly injections; the second group received a single injection of 5 mg/animal; and the third group received three weekly injections of 2, 4, and 4 mg/animal. One concurrent group of Wistar rats (controls), five weeks of age, received a single injection of saline alone. Average lifespans were 120, 102, 130, and 120 weeks, respectively. No intra-abdominal tumor was reported in 47 and 32 rats that were examined in the second and third groups; six of 113 rats (5.3%) examined in the first group had sarcomas, mesotheliomas, or carcinomas of the abdominal cavity (numbers unspecified). Two of 32 controls (6.3%) had abdominal tumors (tumor type not specified). In a similar experiment with female Sprague Dawley rats that received single intraperitoneal injections of 5 mg/animal titanium dioxide, 2/52 rats (3.8%) developed abdominal tumors (tumor type not specified) (average lifespan, 99 weeks).

Groups of female Fischer 344/Jslc rats (n=330; number of rats per group unspecified),
five weeks of age, received intraperitoneal injections of one of several man-made mineral
fibers, including titanium oxide (rutile) whiskers [fiber length, ~ 2.5 um; fiber diameter,
~0.125 um (estimated from a figure)]. The fibers were given in doses of 5, 10 or 20 mg
with 1 mg of dust suspended in 1 mL saline before injection. The greatest volume
administered in a week was 5 mL. The fiber concentration of titanium oxide whiskers was 639x10[3]/μg. Two years after administration, peritoneal mesotheliomas were induced by silicon carbide whiskers (fiber concentration, 414x10[3]/μg; cumulative incidence, 70 to 100%) and potassium titanate whiskers (fibre concentration, 594x10[3]/μg; cumulative
incidence, 20 to 77%), but not by titanium dioxide whiskers.]

	6. Animal metabolism. [NA-remove]

	7. Metabolite toxicology. [NA-remove]

	8. Endocrine disruption. [NA-remove]

C. Aggregate Exposure

	1. Dietary exposure. [NA-remove]

	i. Food. [Titanium dioxide has low acute, subchronic, and reproductive/ developmental toxicity and is not mutagenic or carcinogenic (via the exposure routes most likely to occur through contact/consumption of food and feed crops treated with pesticide formulations containing titanium dioxide; oral and dermal routes of exposure).

There are multiple tolerance exemptions for titanium dioxide. Titanium dioxide is exempt from the requirement of a tolerance under 1) 40 CFR 180.920 when used as a pigment/ coloring agent in plastic bags used to wrap growing banana (preharvest) and as a colorant on seeds for planting; 2) 40 CFR 180.930 when used as a pigment/colorant in pesticide formulations for animal tags; 3) 40 CFR 180.1195 when used as UV protectant in microencapsulated formulations of the insecticide lambda cyhalothrin at no more than 3.0% by weight of the formulation; 4) 40 CFR 180.1195 as a UV-stabilizer at no more than 5% in pesticide formulations containing the active ingredient napropamide; and 5) 
40 CFR 180.1195 when used as a colorant in pesticide formulations intended for varroa mite control around bee hives at a maximum concentration of 0.1% by weight of the pesticide formulation.

Titanium dioxide is also approved by FDA for use as a colorant in food (21 CFR 73.575); in drugs (21 CFR 73.1575); and in cosmetics (21 CFR 73.2575 and 73.3126).

While it is difficult to develop a precise estimate of total human exposure to titanium dioxide, its low toxicity at relatively high doses indicate that expected exposures are likely to be of low toxicological concern. No endpoint of concern via the oral or dermal routes of exposure were identified by EPA from the studies reviewed to support the tolerance exemptions under 40 CFR 180.1195. In addition, EPA did not conduct a quantitative assessment for dietary, non-dietary, or drinking water exposures. Therefore, it can be concluded that dietary exposure to titanium dioxide should not present concern to human health.]

	ii. Drinking water. [Titanium dioxide has low acute, subchronic, and reproductive/ developmental toxicity and is not mutagenic or carcinogenic (via the exposure routes most likely to occur through contact/consumption of food and feed crops treated with pesticide formulations containing titanium dioxide; oral and dermal routes of exposure).

There are multiple tolerance exemptions for titanium dioxide. Titanium dioxide is exempt from the requirement of a tolerance under 1) 40 CFR 180.920 when used as a pigment/ coloring agent in plastic bags used to wrap growing banana (preharvest) and as a colorant on seeds for planting; 2) 40 CFR 180.930 when used as a pigment/colorant in pesticide formulations for animal tags; 3) 40 CFR 180.1195 when used as UV protectant in microencapsulated formulations of the insecticide lambda cyhalothrin at no more than 3.0% by weight of the formulation; 4) 40 CFR 180.1195 as a UV-stabilizer at no more than 5% in pesticide formulations containing the active ingredient napropamide; and 5) 
40 CFR 180.1195 when used as a colorant in pesticide formulations intended for varroa mite control around bee hives at a maximum concentration of 0.1% by weight of the pesticide formulation.

Titanium dioxide is also approved by FDA for use as a colorant in food (21 CFR 73.575); in drugs (21 CFR 73.1575); and in cosmetics (21 CFR 73.2575 and 73.3126).

While it is difficult to develop a precise estimate of total human exposure to titanium dioxide, its low toxicity at relatively high doses indicate that expected exposures are likely to be of low toxicological concern. No endpoint of concern via the oral or dermal routes of exposure were identified by EPA from the studies reviewed to support the tolerance exemptions under 40 CFR 180.1195. In addition, EPA did not conduct a quantitative assessment for dietary, non-dietary, or drinking water exposures. Therefore, it can be concluded that drinking water exposure to titanium dioxide should not present concern to human health.]

	2. Non-dietary exposure. [Titanium dioxide has low acute, subchronic, and reproductive/ developmental toxicity and is not mutagenic or carcinogenic (via the exposure routes most likely to occur through contact/consumption of food and feed crops treated with pesticide formulations containing titanium dioxide; oral and dermal routes of exposure).

There are multiple tolerance exemptions for titanium dioxide. Titanium dioxide is exempt from the requirement of a tolerance under 1) 40 CFR 180.920 when used as a pigment/ coloring agent in plastic bags used to wrap growing banana (preharvest) and as a colorant on seeds for planting; 2) 40 CFR 180.930 when used as a pigment/colorant in pesticide formulations for animal tags; 3) 40 CFR 180.1195 when used as UV protectant in microencapsulated formulations of the insecticide lambda cyhalothrin at no more than 3.0% by weight of the formulation; 4) 40 CFR 180.1195 as a UV-stabilizer at no more than 5% in pesticide formulations containing the active ingredient napropamide; and 5) 
40 CFR 180.1195 when used as a colorant in pesticide formulations intended for varroa mite control around bee hives at a maximum concentration of 0.1% by weight of the pesticide formulation.

Titanium dioxide is also approved by FDA for use as a colorant in food (21 CFR 73.575); in drugs (21 CFR 73.1575); and in cosmetics (21 CFR 73.2575 and 73.3126).

While it is difficult to develop a precise estimate of total human exposure to titanium dioxide, its low toxicity at relatively high doses indicate that expected exposures are likely to be of low toxicological concern. No endpoint of concern via the oral or dermal routes of exposure were identified by EPA from the studies reviewed to support the tolerance exemptions under 40 CFR 180.1195. In addition, EPA did not conduct a quantitative assessment for dietary, non-dietary, or drinking water exposures. Therefore, it can be concluded that non-dietary exposure to titanium dioxide should not present concern to human health.]

D. Cumulative Effects [Based on EPA' previous reviews of titanium dioxide for an exemption of a tolerance under 40 CFR 180.1195, EPA did not make a common mechanism of toxicity finding for titanium dioxide and any other substances and determined that titanium dioxide does not appear to produce a toxic metabolite produced by other substances.]

E. Safety Determination

	1. U.S. population. [Titanium dioxide has low acute, subchronic, and reproductive/developmental toxicity and is not mutagenic or carcinogenic (via the exposure routes most likely to occur through contact/consumption of food and feed crops treated with pesticide formulations containing titanium dioxide; oral and dermal routes of exposure).

When titanium dioxide is used on raw agricultural commodities in accordance with good agricultural practice, it is expected to meet EPA's reasonable certainty of no harm requirement. Based on the low toxicity profile of titanium dioxide, there is no reason to believe that adverse effects on the U.S. population will result from the use of this inert ingredient in pesticide formulations. 

Also, during EPA's most recent review of data to support the exemption from a requirement of a tolerance for titanium dioxide under 40 CFR 180.1195, the Agency concluded that due to titanium dioxide's low potential hazard and the lack of a hazard endpoint, it was determined that a quantitative risk assessment using safety factors applied to a point of departure protective of an identified hazard endpoint is not appropriate for titanium dioxide.]

	2. Infants and children. [Titanium dioxide has low acute, subchronic, and reproductive/developmental toxicity and is not mutagenic or carcinogenic (via the exposure routes most likely to occur through contact/consumption of food and feed crops treated with pesticide formulations containing titanium dioxide; oral and dermal routes of exposure).

When titanium dioxide is used on raw agricultural commodities in accordance with good agricultural practice, it is expected to meet EPA's reasonable certainty of no harm requirement. Based on the low toxicity profile of titanium dioxide, there is no reason to believe that adverse effects on the U.S. population will result from the use of this inert ingredient in pesticide formulations. In addition, there is no reason to believe that infants and children will be disproportionately at risk to the use of titanium dioxide in pesticide formulations.

Also, during EPA's most recent review of data to support the exemption from a requirement of a tolerance for titanium dioxide under 40 CFR 180.1195, the Agency concluded that due to titanium dioxide's low potential hazard and the lack of a hazard endpoint, it was determined that a quantitative risk assessment using safety factors applied to a point of departure protective of an identified hazard endpoint is not appropriate for titanium dioxide. For the same reasons that a quantitative risk assessment based on a safety factor approach is not appropriate for titanium dioxide, a 10-fold FQPA safety factor is not needed to protect the safety of infants and children.]

F. International Tolerances

	[International tolerances, if any, are not known for titanium dioxide.]