Ester reaction products of 1,4-Benzenedicarboxylic acid with C11-14 iso-alcohols, C13-rich

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 weeks, with additional testing for recovery

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented study report equivalent or similar to OECD guideline 408: GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

GLP compliance:

yes

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

416-740-6

EC Name:

-

Molecular formula:

Can vary from C30H53O4 (di-C11 rxn product) to C36H65O4 (di-C14 rxn product)

IUPAC Name:

Ester reaction products of 1,4-Benzenedicarboxylic acid with C11-14 iso-alcohols, C13-rich

Details on test material:

Assumed 100% purity (Lot number 97455). Light yellow liquid with mild odor, stored at room temperature

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

120 total (60 males, 60 females) obtained from Charles River Laboratories (Kingston, New York 12484), identified as Albino Rats (Outbred) VAF/Plus® and Crl:CD® (SD)IGS BR. 100 total (50 males, 50 females) were used in the study.

Females were nulliparous and non-pregnant.

Approximately 4 weeks at receipt, approximately 6 weeks at initiation of dosing. Weights at initiation of dosing were mean 205 grams (range 182-233 grams) for males and 160 grams (range 119-175 grams). Individual weights of animals placed on test were within ±20% of the mean weight for each sex.

Animals were acclimated for 14 days. All animals were examined during the acclimation period to confirm suitability for study.

More animals than required for the study were purchased and acclimated. Animals considered unsuitable for the study on the basis of pretest physical examinations were eliminated prior to random selection for group assignment. Disposition of all animals not utilized in the study is maintained in the study file.

Animals considered suitable for study were distributed into 2 groups of 15 animals per sex and 2 groups of 10 animals per sex by a computerized random sort program so that body weight means for each group were comparable. Each rat was identified with a metal ear tag bearing its assigned animal number. The assigned animal number plus the study number comprised the unique animal number for each animal. If the tag was lost, it was replaced. In addition, each cage was provided with a cage card which was color-coded for dose level identification and contained study number and animal number information. Animals were monitored by the technical staff for any conditions requiring possible veterinary care and treated as necessary.

Animals were doubly housed in elevated, stainless steel, wire mesh cages during the first week of the acclimation period and individually housed thereafter. Certified Rodent Diet, No. 5002; (Meal) (PMI Nutrition International, St. Louis, Missouri) was available without restriction. Fresh food was presented weekly. Analysis of each feed lot used during this study was performed by the manufacturer. Results are maintained on file at the Testing Facility. Water was available without restriction via an automated watering system (Elizabethtown Water Company, Westfield, New Jersey). Water analyses are conducted by Elizabethtown Water Company, Westfield, New Jersey (Raritan-East Millstone Plant) to ensure that water meets standards specified under the EPA Federal Safe Drinking Water Act Regulations (40 CFR Part 141). In addition, water samples are collected biannually from representative rooms in the Testing Facility; chemical and microbiological water analyses are conducted on these samples by a subcontract laboratory. Results of all water analyses are maintained on file at the Testing Facility. There were no known contaminants in the feed or water which were expected to interfere with the results of this study.

A twelve hour light/dark cycle controlled via an automatic timer was provided. Temperature was monitored and recorded twice daily and maintained within the specified range to the maximum extent possible. Excursions outside the specified range were not considered to have affected the integrity of the study.
Desired: 18 to 26°C
Actual: 17 to 28°C
Relative humidity was monitored and recorded once daily and maintained within the specified range to the maximum extent possible. Excursions outside the specified range were not considered to have affected the integrity of the study.
Desired: 30 to 70%
Actual: 32 to 78%

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 1% methylcellulose (Sigma Chemical Company, Lots 105H0489 and 126H1424)

Details on oral exposure:

Animals were dosed by oral gavage once per day with a volume of 5 mL/kg/day. Doses were:
Group I - 0 mg/kg/day
Group II - 50 mg/kg/day
Group III - 500 mg/kg/day
Group IV - 1000 mg/kg/day

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The appropriate amount of test article was weighed out and placed in the beaker into which vehicle was added until the proper volume was reached. Mixture was homogenized for approximately 3 minutes and then stirred with a stir bar/plate for at least 30 minutes or until mixed well.

Doses were prepared once weekly and refrigerated when not in use.

Analyses to determine homogeneity, stability, and concentration of the test and/or control articles with carriers under the conditions of this study were performed by the Testing Facility. Prior to initiation of the study, batches of low-concentration and high- concentration dose solutions were prepared. Three samples each from the top, middle and bottom portion of each mixture were taken for homogeneity analysis. Duplicate samples of the low- and high-concentration dose solutions were assayed for stability 4, 7, and 14 days after preparation (samples for homogeneity assays, evaluated on the day of preparation, were used to establish concentration at time of preparation). All dose levels were assayed weekly for the first four weeks and monthly thereafter (1 sample per concentration was taken and two subsamples were analyzed).

Duration of treatment / exposure:

Ninety days

Frequency of treatment:

Once per day

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 other: mg/kg/day (actual ingested)

Remarks:

Doses / Concentrations:
0
Basis:
actual ingested

Doses / concentrations 2

Dose / conc.:

50 other: mg/kg/day (actual ingested)

Remarks:

Doses / Concentrations:
50 mg/kg/day
Basis:
actual ingested

Doses / concentrations 3

Dose / conc.:

500 other: mg/kg/day (actual ingested)

Remarks:

Doses / Concentrations:
500 mg/kg/day
Basis:
actual ingested

Doses / concentrations 4

Dose / conc.:

1 000 other: mg/kg/day (actual ingested)

Remarks:

Doses / Concentrations:
1000 mg/kg/day
Basis:
actual ingested

No. of animals per sex per dose:

Primary test group was 10 per sex per dose (80 total)
Recovery was tested with 5 per sex per dose from control and high dose group (20 total)

Control animals:

yes, concurrent vehicle

Details on study design:

This study was conducted for Mobil Business Resources Corporation. It was designed to assess the test substance for potential toxicity when administered orally, via gastric intubation, to Sprague-Dawley CD® rats at dose levels of 50, 500 and 1000 mg/kg/day for a period of at least 90 days and to assess reversiblity of any effects during a four-week recovery period. Data from this study may serve as a basis for classification and labeling of the test article.

This study was designed to meet or exceed the requirements of OECD (Organization for Economic Co-Operation and Development): Guidelines for Testing of Chemicals No. 408: “Subchronic Oral Toxicity - Rodent: 90-day Study” adopted 12 May 1981. In addition, the study was designed in accordance with the Draft Document (March 1997) (May 1996), OECD Guidelines for the Testing of Chemicals, Proposal for Updating Guideline 408, Repeated Dose 90-Day Oral Toxicity Study in Rodents.

This study was conducted in compliance with OECD Principles of Good Laboratory Practices ENV/MC/CI-IEM/(98)17 and EEC Good Laboratory Practices (90/18/EEC).

Positive control:

None

Examinations

Observations and examinations performed and frequency:

---MACROSCOPIC EXAMINATIONS---
Complete macroscopic postmortem examinations were performed on all animals killed at a scheduled sacrifice interval immediately after death. The macroscopic postmortem examination included examination of the external surface and all orifices; the external surfaces of the brain and spinal cord; the organs and tissues of the cranial, thoracic, abdominal and pelvic cavities and neck; and the remainder of the carcass for the presence of macroscopic morphologic abnormalities. Animals were fasted prior to scheduled sacrifices.
Animals which were sacrificed for humane reasons were similarly examined as soon as it was practically possible.

---ORGAN WEIGHTS---
Organs [adrenal glands, aorta, bone and bone marrow (sternum/femur), bone (femur with joint), brain (medulla/pons, cerebrum and cerebellum, optic chiasma), esophagus, eyes, Harderian gland, head, heart, kidneys, large intestine (cecum, colon, rectum), larynx, liver, lungs (with mainstem bronchi), lymph nodes (cervical, mesenteric and submandibular), mammary gland, nerve (sciatic), ovaries, pancreas, Peyer’s patches, pharynx, pituitary gland, prostate gland, salivary glands (submandibular), seminal vesicles, skeletal muscle (Biceps femoris), skin, small intestine (duodenum, ileum,jejunum), spinal cord (cervical, thoracic, lumbar), spleen, stomach, testes with epididymides, thymus, thyroid/parathyroid glands, tongue, trachea, urinary bladder, uterus (body/horns) with cervix, vagina, and tissues with macroscopic findings including tissue masses] were weighed for all animals at the scheduled sacrifice intervals. Prior to weighing, the organs were carefully dissected and properly trimmed to remove adipose and other contiguous tissues in a uniform manner. Organs were weighed as soon as possible after dissection in order to avoid drying. Paired organs were weighed together.

---CLINICAL CHEMISTRY---
-Method of Blood Collection-
Blood for clinical chemistry studies was collected into tubes with no anticoagulant, allowed to clot, and centrifuged to obtain serum.

-Collection Intervals-
Termination: Week 14
Recovery: Week 18

-Number of Animals Bled/lnterval-
Termination: 80 animals (10/sex/group)
Recovery: 20 animals (5/sex/control and high-dose groups)

-Analysis of Blood Samples-
Blood samples were analyzed using a Hitachi 717 Automatic Analyzer (Boehringer Mannheim Corporation) as follows:
Aspartate aminotransferase (Kinetic - Modified IFCC Technique)
Alanine aminotransferase (Kinetic - Modified IFCC Technique)
Alkaline phosphatase (AMP Buffer - Modified Bessey-Lowry-Brock Technique)
Blood urea nitrogen (Modified Urease Technique)
Creatinine (Jaffe Reaction - Kinetic - Alkaline Picrate)
Glucose (Hexokinase Method)
Cholesterol - enzymatic (Cholesterol esterase-cholesterol oxidase Method)
Triglycerides (GPO Triglyceride-lipase Method)
Total protein (Biuret Technique)
Albumin (Bromocresol Green Method)
Globulin (calculated value; total protein - albumin)
Albumin/globulin ratio (calculated value; albumin ÷ globulin)
Total bilirubin (Modified Jendrassik and Grof Method)
Sodium (Ion Selective Electrode)
Potassium (Ion Selective Electrode)
Chloride (Ion Selective Electrode)
Calcium (Cresolphthalein Complexane Method)
Inorganic phosphorus (Phosphomolybdate-UV Method)
Gamma-glutamyl transferase (Kinetic – Szasz, G.)

---HEMATOLOGY---
-Method of Blood Collection-
Blood for hematology studies was collected into tubes containing EDTA anticoagulant. Blood for coagulation studies was collected into tubes containing sodium citrate anticoagulant.

-Collection Intervals-
Termination: Week 14
Recovery: Week 18

-Number of Animals Bled/Interval-
Termination: 80 animals (10/sex/group)
Recovery: 20 animals (5/sex/control and high-dose groups)

-Analysis of Blood Samples-
Blood samples were analyzed using a Teclinicon H-1 Hematology System (Bayer Corporation) as follows:
Hemoglobin concentration
Hematocrit
Erythrocyte count
Platelet count
Mean corpuscular volume
Mean corpuscular hemoglobin
Mean corpuscular hemoglobin concentration
Total leukocyte counts
Differential leukocyte count (When questionable values were obtained, manual differential leukocyte counts and absolute value calculations were performed for verification)
Absolute lymphocytes (calculated value; total WBC x % lymphocyte value ÷ 100)
Absolute segmented neutrophils (calculated value; total WBC x % segmented neutrophil ÷ 100)

Blood samples were analyzed using a Photo-optical clot detection system, COAG-A-MATE® X2 (Organon Teknika Corp) as follows:
Prothrombin time
Activated partial thromboplastin time

Blood samples were also examined for erythrocyte morphology (method not disclosed)

Sacrifice and pathology:

---NECROPSY INFORMATION---
-Method of Euthanasia-
Exsanguination following carbon dioxide inhalation.

-Necropsy Intervals-
Termination: Week 14
Recovery; Week 18

-Number of Animals-
Termination: 80 animals (10/sex/group)
Recovery: 19 animals (up to 5/sex/control and high-dose groups)

-Necropsy Order-
A necropsy schedule was established to ensure that approximately equal numbers of males and females were examined on each day of necropsy and that examination of animals of both sexes were performed at similar times of the day throughout the necropsy period. In addition, animals were sacrificed in the following order: one rat each was selected from the control, high-, mid-, and low-dose groups and sacrificed in that order. This sequence was repeated until all the rats were sacrificed.

---HISTOPATHOLOGY---
-TISSUES PRESERVED AND EXAMINED HISTOPATHOLOGICALLY-
The tissues listed above were obtained at the scheduled sacrifice intervals and preserved for all animals. In addition, slides of the indicated tissues were prepared and examined microscopically for the 10 animals per sex from the control and high-dose groups sacrificed at the terminal sacrifice interval. In addition, indicated tissues were examined microscopically for Animal No. 1584 which was sacrificed for humane reasons during the recovery period. Any abnormalities not noted during macroscopic examinations which were seen during histology processing were recorded.

-Preservatives-
All tissues - 10% neutral buffered formalin.
Eyes were placed in glutaraldehyde/paraformaldehyde initially and then retained in 10% formalin. Lungs and urinary bladder were infused with formalin prior to their immersion into a larger volume of the same fixative.

-Processing-
After fixation, the tissues and organs from all animals were routinely processed, embedded in paraffin, cut at a microtome setting of 4- 5 microns, mounted on glass slides, stained with hematoxylin and eosin and examined by light microscopy at Huntingdon Life Sciences, Eye Research Center. The bones were decalcified in formic acid.

Other examinations:

---NEUROBEHAVIORAL STUDIES---
Testing was staggered over four sessions with approximately equal numbers of animals per sex per dose group in each session. Noise level in the functional observational battery room was maintained within a level of 55 to 65 decibels by a White Noise Generator, Model MDF 5001 (MDF Products, Inc. Danbury, Connecticut). Temperature, humidity, noise level and illumination of each room were measured and recorded to ensure that variations in environmental conditions were minimal during all evaluations. The functional observational battery was performed for all animals before evaluation of motor activity.

--MOTOR ACTIVITY--
-Method-
Using a modified version of Schulze’s procedures (Schulze, 1990), the locomotor activity of all animals was monitored using an automated Photobeam Activity System (San Diego Instruments, Inc., San Diego, California). Sessions were 60 minutes in length; each session was divided into twelve 5-minute intervals. The time of testing was balanced across treatment groups.

-Frequency-
Pretest: Week 0
Termination: Week 13
Recovery: Week 17

-Temperature-
Temperature was monitored and recorded twice daily, just prior to the first evaluation and just after the last evaluation.
Desired: 18 to 26°C
Actual: 22 to 24 °C

-Relative Humidity-
Relative humidity was monitored and recorded twice daily, just prior to the first evaluation and just after the last evaluation.
Desired: 30 to 70%
Actual: 34 to 78%

-Noise Level-
Noise level was monitored using a Digital Sound Level Meter, Model 840029 (Sper Scientific, Ltd., Scottsdale, Arizona) and recorded twice daily, just prior to the first evaluation and just after the last evaluation.
Desired: 55 to 65 dB
Actual: 57 to 65 dB

-Illumination-
Illumination was monitored using a Photometer I (Quantum Instruments, Inc., Garden City, New York) and recorded twice daily, just prior to the first evaluation and just after the last evaluation.
Desired: <80 footcandles
Actual: <33 footcandles

--FUNCTIONAL OBSERVATIONAL BATTERY
-Method-
A functional observational battery (Moser, 1989) was performed on all animals. Evaluations were performed “blind”, i.e., the observer did not know the identity of the animal’s dose group. Time of testing was counter-balanced across treatment groups.
The following evaluations were performed as part of the functional observational battery:

Home Cage Evaluations: posture, vocalization and palpebral closure.

Handling Evaluations: reactivity to general stimuli (handling); assessment of signs of autonomic function: lacrimation, salivation, altered fur appearance, or red/crusty deposits around eyes.

Open Field Evaluations: arousal level and gait; count of urination and defecation; convulsions, tremors, abnormal movements or behaviors, excessive or repetitive actions; piloerection and exophthalmos.

Reflex Assessments: response to visual (approach response) and auditory (finger snap) stimuli; response to a tail pinch; pupillary function.

Grip Strength (Meyer et al., 1979): Grip strength was measured using a Grip Strength Meter (Columbus Instruments International Corporation, Columbus, Ohio).

Landing Foot Splay: A small dot of paint was applied to each animal’s hindpaws. Each animal was then dropped on to a flat surface from a height of two feet. The distance between the marks left by the hindpaws was measured in centimeters.

Hindlimb Extensor Strength: Animals were held in a vertical position facing the observer with a firm grasp around the thorax. The observer placed one finger against the bottom of each hindpaw and pressed towards the animal. Muscular resistance and pressure exerted by the animals were scored.

Air Righting Ability: Animals were held upside down and dropped from a height of two feet into a container of bedding. The landing position of the animal was recorded.

Body Weight: Animals were removed from their cages and weighed using a Mettler Balance, Model PE4000 (Mettler Instrument Corporation, Hightstown, New Jersey).

Statistics:

See "Any other information on materials and methods incl. tables" below

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not examined

Description (incidence and severity):

no neoplasias noted

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Remarks on result:

not determinable due to absence of adverse toxic effects

2

Dose descriptor:

NOEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Remarks on result:

not determinable due to absence of adverse toxic effects

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The no observed adverse effect level for oral gavage of 1,4-Benzenedicarboxylic Acid, Di-C11-14-Isoalkyl Ester, C13-Rich is 1000 mg/kg/day. This finding does not warrant the classification of 1,4-Benzenedicarboxylic Acid, Di-C11-14-Isoalkyl Ester, C13-Rich as a specific target organ toxicant under the new Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP) or under the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.

Executive summary:

1,4-Benzenedicarboxylic Acid, Di-C11-14-Isoalkyl Ester, C13-Rich was administered by oral gavage to 50 male and 50 female Sprague Dawley rats at 0, 50, 500, or 1000 mg/kg for 13 weeks. A subset of animals (5 per sex per dose) from the 0 and 1000 mg/kg/day were monitored for an additional 4 weeks as a recovery group. No adverse effects were noted for physical observations, body weights, food consumption, motor activity levels, functional observational batteries, organ weights or macroscopic and microscopic pathology. At the end of the study, the no observed adverse effect level was determined to be greater than 1000 mg/kg. This finding does not warrant the classification of 1,4-Benzenedicarboxylic Acid, Di-C11-14-Isoalkyl Ester, C13-Rich under the new Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP) or under the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.