Registration Dossier

Administrative data

Description of key information

Upon repeated exposure of rats to DEGDBE toxic effects on hematology, liver, kidney and speen were observed. The red blood cell is the presumed target system,  the mode of action being the systemic exposure to 2-butoxyacetic acid. Humans are expected to be less susceptible. 

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
28 day treatment+14 day recovery
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Recently performed Guideline study
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
Test System:

Species/strain: Wistar rats, Crl: WI(Han) (Full Barrier)
Source: Charles River, 97633 Sulzfeld, Germany
Sex: male and female; the female animals were non-pregnant and nulliparous.
Age at the first
administration: males: 7-8 weeks old, females: 7-8 weeks old.
Body weight at the
beginning of the study: males: 151 - 182 g, (mean: 164.3 g, ± 20% = 131.54-197.31 g), females: 128 -150 g, (mean: 137.66 g, ± 20% = 110.13-164.19 g).

The animals were derived from a controlled full-barrier maintained breeding system (SPF). According to Art. 9.2, No. 7 of the German Act on Animal Welfare the animals were bred for experimental purposes.

Housing and Feeding Conditions:
- Full barrier in an air-conditioned room
- Temperature: 22 ± 3°C
- Relative humidity: 55 ± 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: 10 x / hour
- Free access to Altromin 1324 maintenance diet for rats and mice (lot no. 1530)
- Free access to tap water, sulphur acidified to a pH of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals)
- The animals were kept individually in IVC cages, type III H, polysulphone cages on Altromin saw fibre bedding (lot no. 110811)
- Certificates of food, water and bedding are filed at BSL BIOSERVICE
- Adequate acclimatisation period (at least 5 days)
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
The animals of the main groups were treated with the test item or vehicle on 7 days per week for a period of 28 days. The test item formulation or vehicle was administered at a single dose to the animals by oral gavage at an application volume of 4 mL/kg bw.
The animals of the recovery groups (control and high dose) were dosed along with the main groups for 28 days. After 28 days of administration the animals of the recovery groups were observed for a period of 14 days (recovery period).
For each animal the individual dosing volume was calculated on the basis of the body weight most recently measured.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The % nominal of formulation samples meant for the nominal concentration verification taken in study week 1, study week 2, study week 3 and study week 4 was in the range of 88 -104 % in LD group, 99-117% in MD group, 87-101% in HMD group and 97-121% in HD group.
The % nominal of formulation samples meant for the stability verification was 70-84% in LD group and 77-105% in HD group.
The % nominal of formulation samples meant for the homogeneity verification taken in study week 1 was 94-108% in LD group and 118-123% in HD group.
Duration of treatment / exposure:
28 days
Frequency of treatment:
7 days per week for a period of 28 days
Remarks:
Doses / Concentrations:
25, 100, 250 and 750 mg/kg body weight/ day
Basis:
other:
No. of animals per sex per dose:
50 animals (25 males and 25 females) were included in the main study (5 male and 5 female animals per group). The main study included one control (C) and four dose groups (LD, MD, HMD, HD).
In addition, 20 animals (5 male and 5 female animals per group) were included in the control and high dose groups and were observed for a period of 14 days following the last administration.
Control animals:
yes, concurrent vehicle
Details on study design:
Preparation of the Test Item Formulations:

The required amount of test item for each dose concentration was weighed separately into a tared plastic vial on a suitable precision balance and the vehicle (corn oil) was added to give the appropriate final concentration of the test item. Afterwards, the test sample was thoroughly mixed. Homogeneity of the test item in the vehicle was ensured by using a vortex machine. The vehicle was selected as suggested by the sponsor and on the basis of the test item’s characteristics.

Dose Formulation Analysis:

Each dosing concentration were analysed for nominal concentration and homogeneity of the test item in the vehicle at various intervals. Stability analysis was performed on samples collected from the top and bottom dose levels.

Samples for the nominal concentration verification were taken in study week 1, 2, 3 and 4.
Samples for homogeneity were taken from the top, middle and bottom of the high dose and low dose preparation in study week 1.

Samples for stability analysis were taken from top and bottom dose level in study week 1 at 0 hr and 6 hrs from high dose and low dose preparation.

All concentration samples were stored frozen (approximately -20°C) until the transport. The samples were transported to the test site packed in dry ice.
The dose formulation analysis was performed at GLP-certified contract laboratory IBACON GmbH, Arheilger Weg 17, D-64380 Rossdorf, Germany.

Preparation of the Animals:

Prior to the start of the treatment period a detailed clinical observation outside the home cage was made. Before the first administration all animals used for the study were weighed and assigned to the experimental groups with achieving a most homogenous variation in body weight throughout the groups of males and females.

Administration of Doses:

The animals of the main groups were treated with the test item or vehicle on 7 days per week for a period of 28 days. The test item formulation or vehicle was administered at a single dose to the animals by oral gavage at an application volume of 4 mL/kg bw.
The animals of the recovery groups (control and high dose) were dosed along with the main groups for 28 days. After 28 days of administration the animals of the recovery groups were observed for a period of 14 days (recovery period).
For each animal the individual dosing volume was calculated on the basis of the body weight most recently measured.

Body Weight, Food Consumption:

The body weight was recorded once before the assignment to the experimental groups, on the first day of administration and weekly during the treatment and recovery period. To achieve a more accurate individual dosing volume, two additional body weight measurements were performed during each week of the treatment period.
Food consumption was measured weekly during the treatment period the recovery period.

Clinical Observation:

All animals were observed for clinical signs during the entire treatment period of 28 days. The recovery animals were observed for an additional period of 14 days following the last administration.
General clinical observations were made at least once a day, preferably at the same time each day and considering the peak period of anticipated effects after dosing. The health condition of the animals was recorded. Twice daily all animals were observed for morbidity and mortality except for weekends and public holidays when observations were made daily once.
Detailed cageside observations included spontaneous activity, lethargy, recumbent position, convulsions, tremors, apnoea, asphyxia, vocalisation, diarrhoea, changes in skin and fur, eyes and mucous membranes (salivation, discharge), piloerection and pupil size.
Ophthalmological examination, using an ophthalmoscope were made on all animals before the first administration and in the last week of the treatment period as well at the end of the recovery period in the recovery animals.

Functional Observation

Once before the first exposure, as well as once in the fourth week of exposure multiple detailed behavioural observations were made outside the home cage using a functional observational battery of tests . These tests were conducted in all animals.

Haematology:

One day after the last administration, blood was sampled from all surviving animals of the main study for a haematological evaluation. Blood from the recovery animals was sampled at the end of the recovery period. After overnight fasting of the animals, blood from the abdominal aorta was collected in EDTA-coated tubes prior to or as part of the sacrifice of the animals.

Blood Coagulation:

One day after the last administration, blood was sampled from all surviving animals of the main study for an evaluation of the coagulation parameters. Blood from the recovery animals was sampled at the end of the recovery period. After overnight fasting of the animals, blood from the abdominal aorta was collected in citrate tubes prior to or as part of the sacrifice of the animals.

Clinical Biochemistry:

One day after the last administration, blood was sampled from all surviving animals of the main study for an evaluation of the clinical biochemistry. Blood from the recovery animals was sampled at the end of the recovery period. After overnight fasting of the animals, blood from the abdominal aorta was collected in serum separator tubes just prior to or as part of the sacrifice of the animals.

Urine sample collection and urinalysis:

Urine samples were collected from animals as mentioned in below table (Table 5), kept overnight in metabolic cages. The samples were collected on day 9, day 29, day 31 and day 43. The urine samples were stored at -80oC until the metabolite analysis was performed.
This phase of the study was not performed under GLP.

Urinalysis was performed with the portion of samples collected from above mentioned animals on day 9, day 29, day 31 and day 43. Additionally, urine colour/appearance was recorded.

Pathology:

Gross necropsy
All animals from the main and recovery groups were subjected to a detailed gross necropsy on day 29 for main group animals and day 43 for recovery group animals which included careful examination of the external surface of the body, all orifices and the cranial, thoracic and abdominal cavities and their contents.
The wet weight of the organs of all sacrificed animals was recorded as soon as possible. Paired organs were weighed separately. Organ weights of animals found dead or euthanised for animal welfare reasons were not recorded.

The tissues from all animals were preserved in 10% neutral buffered formalin except eyes, testes and epididymides that were fixed in Modified Davidson’s fixative. All animals found moribund and/or intercurrently euthanised were also subjected to a gross necropsy and the organs preserved for the histopathological examination.

Histopathology:

The afore-listed organs of all animals were examined histologically after preparation of paraffin sections and haematoxylin-eosin staining.

All organs and tissues were evaluated in all animals of the control and high dose group sacrificed at the end of the treatment period. In addition, liver, kidney, spleen, thymus and eye were evaluated in all intermediate group animals sacrificed at the end of the treatment period and in all recovery group animals.
All gross lesions identified in any animal were examined.
Histological processing of tissues to microscope slides was performed at the GLP-certified contract laboratory Propath UK Ltd. (test site for tissue processing), Willow Court, Netherwood Road, Hereford HR2 6JU, England. Histopathological evaluation was performed at the GLP-certified contract laboratory KALEIDIS – Consultancy in Histopathology (test site for histopathology), 6 rue du Gers, 68300 Saint-Louis, France. Blocking, embedding, cutting, H&E staining and scientific slide evaluation were performed according to the corresponding SOP’s of the test sites.



Statistics:
A statistical assessment of the results of the body weight, food consumption, parameters of haematology, blood coagulation and clinical biochemistry and absolute and relative organ weights were performed for each gender by comparing values of dosed with control animals of the main groups using a one-way ANOVA and a post-hoc Dunnett Test. High dose and control values of the recovery groups were compared using a Student’s t-Test. These statistics were performed with GraphPad Prism 5.01 software (p<0.05 was considered as statistically significant).
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
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:
effects observed, treatment-related
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Details on results:
Animal Survival:

Two female animals from the HD group and 1 female animal from HD recovery group died prematurely during the 28-day treatment period and no mortalities were recorded at the other dose levels. Predominant histological findings in the decedents were seen in the liver, kidney and lymphoid organs. These premature deaths were considered to be most likely to be related to toxicity.
All surviving main group animals were sacrificed on day 29 and recovery group animals were sacrificed on day 43.

Clinical Observations:

There were clinical signs recorded in treatment group male and female animals. In males the predominant clinical signs recorded were alopecia, salivation, piloerection and moving the bedding. These finding were more pronounced in HD group males. These clinical findings in male HD group were considered to be related to treatment without adversity. In females, the predominant clinical signs were abonormal breathing, recumbency, piloerection, salivation, discoloured urine, bradykinesia, ataxia and hypotonia. There were also some sporadic clinical signs noted in few animals during the study. Considering the mortality in HD group, these signs were considered to be adverse.

Functional Observation Battery:

No relevant effects were observed in any of the parameters of the functional observation battery before and at the end of the treatment period. There were no biologically relevant differences in body temperature between the groups. There were no ophthalmoscopic findings in any of the animals of this study.
However there were minor changes in 3 females of HD group. The changes were seen for equilibrium reflex (3/3 animals), positional passivity (2/3 animals), and startle/ auditory response (1/ 3 animals). These finding were not noted after the cessation of dose application. Hence, these findings were considered not likely to be an adverse effect.

Body Weight Development:

In males, there was no statistically significant difference observed for mean body weight in treatment and recovery group animals during the study period when compared to corresponding control group. There was statistical significant decrease observed for body weight gain in HD group. This decrease was assumed to be due to lower body weight gain in 2/10 animals (animal no. 22 and 23) in HD group. This change was considered to be not likely to be adverse. There was no statistical difference observed for animals between control and HD groups at the end of the recovery period.
In females, there was no statistically significant difference observed for mean body weight and weight gain in treatment and recovery group animals during the treatment or recovery period when compared to corresponding control group. However, there was substantial decrease in body weight gain noted in LD and HMD group in week 4. This decrease was due to considerable decrease in body weight gain of isolated animals (animal no 34, 41 and 43). There was no dose response pattern observed and hence was not considered to have toxicological relevance.
There was also substantial decrease noted for weight gain in HD recovery group in week 5. During last week of recovery there was slight decrease in mean daily weight gain in HD group without attaining the statistical significance. This difference in body weight gain was considered not likely to be an adverse effect.

Food Consumption:

In males and females, no statistical significant changes were recorded in treatment or recovery group animals when compared to the corresponding control group. However, there was slight decrease noted in males in HD group (week 1), HMD group (week 2), MD, HMD and HD groups (week 4). In females, slight decrease was noted in HD group in week 1. The changes in food intake corroborated only to body weight change only at HD group, which was related to treatment. The changes recorded at lower dose levels (LD, MD and HMD) were of no toxicological relevance.

Haematology and Blood Coagulation:

In males, there was statistically significant decrease recorded for mean RBC, Hb and Hct in HD group, statistically significant increase in mean MCV and RE values in HD group when compared to corresponding control group. The differential count indicated statistically significant increase in Neu and Mono values in LD group, statistically significant decrease in Lymph value in LD group, statistically significant decrease in Eos values in LD, MD, HMD and HD groups wen compared to the corresponding control group. The changes in RBC, Hb, Hct, MCV, Neu, Lymph, Mono and RE did not show dose response pattern. Considering the minimal or mild multifocal extramedullary hemopoiesis in the spleen observed in 4/5 males of HD group, the changes noted in HD group in association with histopathological changes in spleen were considered to be due to treatment. The changes for mean Eos showed a clear dose response, but the WBC values of treatment groups were comparable to control group. Considering the tendency towards more atrophy/regression of thymus in the males of HD group and changes only being restricted to HD group, the changes at lower dose levels (LD, MD and HMD) were considered of no toxicological relevance.
The measurement of blood coagulation indicated statistically significant increase in mean aPTT values in HMD and HD groups. Considering the values being within the range of historical control data, the finding was considered not likely to be an adverse effect.
During the recovery period there was statistically significant increase in mean Hct value in HD group. In the absence of histopatholohical changes in spleen in recovery group animals, the finding was not likely to be related to treatment.

In females, there was statistically significant increase in mean MCV and MCH in HD group, statistically significant decrease in mean MCHC value in HD group and statistically significant increase in mean RE values in HMD and HD groups. The above changes showed dose relationship from HMD to HD group. Considering the dose dependency and a tendency towards higher spleen weights, minimal or mild multifocal extramedullary hemopoiesis in the spleen observed in 2/3 females of HD group the changes in HD group was considered to be due to treatment.
There were no statistically significant changes recorded for blood coagulation and differential count parameters.
During the recovery period there was statistically significant increase in mean HB, MCH values in HD group, statistically significant increase in mean Luc value in HD group, statistically significant decrease in mean RE values in HD group. There was also significant decrease in mean aPTT value in HD group when compared to the control. In the absence of histopathological changes in recovery group females that could support the above findings, these findings were considered not likely to be adverse.

Clinical Biochemistry:

In males, there were no statistically significant changes recorded for the measured clinical biochemistry parameters in treated main group animals. However, there was considerable decrease in mean AP values in HMD group, substantial increase in mean Crea value in HMD and HD group, substantial increase in mean TBIL values in HMD and HD groups, substantial decrease in mean TBA value in all treatment groups when compared to corresponding control group. All above changes did not show dose response relationship. These changes were considered not likely to be an effect of treatment. In recovery males, there was statistically significant decrease noted for mean Gluc in HD group. The value being within the historical control range, the change was not considered to be of toxicological relevance.
In females, there was statistically significant decrease in mean ASAT, TP and Urea values in HD main group when compared to control. These changes were not likely to be related to treatment. In recovery females, there was statistically significant decrease noted for mean ASAT value and increased noted for mean Chol value in HD group. Due to histopathological kidney changes in recovery animals, the increase Chol was related to treatment.

Urinalysis:

Urinalysis was performed with the portion of samples collected from animals of control and treatment group animals on day 9, day 29, day 31 and day 43.
Males: There were high level of ketone recorded in most of HD group animals (day 29), high level of BIL in 2/5 HDR animals when compared to concurrent control animals (recovery day 3), high level of BIL and protein in 2/5 HDR animals when compared to concurrent control animals (recovery day 15),
In females, there were high level of BIL in 3/5 animals (against 1/5 control animals) in HDR group (recovery day 3). There were high level of protein in 2/5 animals in HDR group (recovery day 15).
These changes seen in males and females correlated with histological changes in kidney. Hence, the effect was considered to be of toxicological relevance.

Pathology:

There were few macroscopic findings recorded at necropsy. The macroscopic findings are listed as follows,
Males- C, LD, HMD, HD and CR group: no findings; MD group: small testes (1/5 animals), small epididimides (1/5 animals); HDR group: red discoloured mandibular lymphnodes (1/5 animals), yellow discoloured (cyst like) left axillary lymph node (1/5 animals), multiple areas of yellow tissue on ilieum (1/5 animals), grey discoloured lung (1/5 animals), enlarged thymus (1/5 animals).

Females- C: fluid distention of uterus (with oviduct and cervix) (1/5 animals), LD, HMD and HD: no findings; MD: fluid distention of uterus (with oviduct and cervix) (1/5 animals); CR: red discoloured axillary lymph node (1/5 animals), red discoloured thymus (1/5 animals), fluid distention of uterus (with oviduct and cervix) (1/5 animals); HDR: yellowish fluid content in stomach (1/5 animals).

The macroscopic findings noted at terminal or recovery sacrifice were considered not likely to be related to treatments.

Organ Weight:

In males, there was statistically significant decrease noted for absolute heart and thymus weight in HD group. There was statistically significant decrease in relative brain weight of thymus in HD group. There was statistically significant increase in relative body weight of spleen in HD group. The changes in thymus and spleen weights were confirmed by histopathologhical changes. Therefore, the changes were related to toxicity.

In females, there was statistically significant increase noted for absolute liver and spleen weight in HD group. There was statistically significant increase in absolute spleen weight in HD recovery group. There was statistically significant increase in relative brain weight of liver in HD group. There was statistically significant increase in relative brain weight of spleen in HD recovery group. There was statistically significant increase in relative body weight of liver in HMD and HD groups and relative body weight of spleen in HD groups. There was statistically significant increase in relative body weight of spleen HD recovery group. Considering the hepatocellular hypertrophy of liver and minimal or mild multifocal extramedullary hemopoiesis of spleen in HD group, the changes in HD group were considered to be due to toxicity.

Histopathology:

Three females in HD group were sacrificed moribund during the early treatment period. Predominant histological findings in the decedents were seen in the liver, kidney and lymphoid organs and their death was considered likely to be adverse.

In the liver, minimal centrilobular or panlobular hepatocellular hypertrophy was seen in the majority of animals of HD group, as well as in one single male of HMD group. The kidney showed minimal golden-brown pigment in the corticotubular epithelium in a proportion of animals of HD group and one single female of HMD group. Minimal or mild multifocal extramedullary hemopoiesis in the spleen was observed in the majority of animals of HD group. In the thymus, there was a tendency towards more atrophy/regression in the males of HD group, confirming lower thymus weights recorded.

After the recovery period, the kidney change was still observed, the other findings had resolved completely.

Dose Formulation Analytics

The % nominal of formulation samples meant for the nominal concentration verification taken in study week 1, study week 2, study week 3 and study week 4 was in the range of 88 -104 % in LD group, 99-117% in MD group, 87-101% in HMD group and 97-121% in HD group.
The % nominal of formulation samples meant for the stability verification was 70-84% in LD group and 77-105% in HD group.
The % nominal of formulation samples meant for the homogeneity verification taken in study week 1 was 94-108% in LD group and 118-123% in HD group.





Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'
Critical effects observed:
not specified
Conclusions:
DEGDBE was investigated for its repeated dose toxicity according to the Guideline OECD 407. The NOAEL of 100 mg/kg bw was obtained.
Executive summary:

DEGDBE was investigated for its repeated dose toxicity according to the OECD Guideline 407- Five rats per dose and per sex were treated with DEGDBE per gavage at doses of 0, 25, 100, 250 and 750 mg/kg bw. Additional five rats per dose and per sex were treated with 0 and 750 mg/kg bw and were allowed to recover for 14 days.

At 750 mg/kg bw three out of ten females did not survive the treatment. Alteration in liver, kidney and lymphoid organs were observed in decedents. Clinical signs indicative of intoxication occurred at 750 mg/kg bw and were more pronounced for females.

No effect was observed upon functional observation batteries and ophthalmoscopic observation and no significant effect was observed on body weight, body weight gain and food consumption.

The alterations in hematology at 750 mg/kg bw and partially also at 250 mg/kg bw were indicative of hemolysis and inflammatory status. No significant effect was found for clinical biochemistry. At doses of 750 and 250 mg/kg bw the organ weight changes were observed for spleen, liver and thymus together with the histopathology findings, extramedullary hemopoiesis in the spleen and hepatocellular hypertrophy in the liver.

Based on the findings above, the dose level of 100 mg/kg bw was considered to be the NOAEL.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
100 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The data on DEGDBE is limited to one 28-day oral toxicity study. The data on source chemicals comprise studies on diverse species and up to 2 year chronic toxicity studies. The quality of whole database is considered to be sufficiently robust for a reliable hazard assessment on DEGDBE.

Additional information

The category approach is used to asses the repeated dose toxicity of DEGDBE. Following aspects will be discussed:

a)     Category definition and justification

b)   Repeated dose toxicity of DEGDBE based on category building

a) Category approach justification:

The repeated dose toxicity of DEGDBE is to be assessed by read-across consideration. Three structurally related glycol ethers are identified as suitable surrogates. Together with DEGDBE, a category building is proposed to increase the robustness of the read-across consideration.

a1)Category members and chemical structures: the similarity in their structure is given by presence of butoxylated ethylene glycol at terminal position (Butyl-O-CH2-OH-).

Ethylene glycol butyl ether (EGBE)*

CAS 111-76-2

Butyl-O-CH2-OH

Diethylene glycol butyl ether (DEGBE)*

CAS 112-34-5

Butyl-O-CH2-O-CH2-OH

Diethylene glycol dibutyl ether (DEGDBE)**

CAS 112-73-2

Butyl-O- CH2-O-CH2-O-Butyl

Polyethylene glycol dibutyl ether (PolyEGDBE)***

CAS 31885-97-9

Butyl-O- CH2-O-CH2-O-CH2-O-Butyl

* EGBE and DEGBE are extensively investigated substances and reviews on their toxicity profiles are available in public domain (i.e. EU Risk Assessment Report, 2-Butoxyethanol (EGBE), CAS 111 -76 -2, 2008; Opinion on Diethylene Glycol Monobutyl Ether (DEGBE), SCCP/1043/06, 2006). **target chemical. ***Clariant own data, details provides in corresponding endpoint study record.

a2) The proposed grouping is justified by the common mode of action, namely systemic exposure to 2-butoxyacetic acid (2-BAA) and/or butoxyethoxyacetic acid (BEAA):

- EGBE: 2-BAA is the major urinary metabolite (summarized in EU risk assessment, 2008)

- DEGBE: BEAA is the major urinary metabolite (Deisinger et al. 1989)

- DEGDBE: in 28-day study (Clariant own data) the urinary 2-BAA determination was incorporated; 750 mg/kg bw external dose level corresponded to 1400 mg/L 2-BAA in urine.

- PolyEGDBE: no experimental data is available; BEAA and/or 2-BAA as metabolite can be reasonably assumed due to the observed RBC reduction and indication of compensatory increased hematopoietic activity.

a3) The proposed grouping is justified by the comparable toxicity profiles, which reflects the toxicity action of 2-BAA and/or BEAA. Both metabolites are known to induce hemolysis (Udden 2002; Udden 2005).

- EGBE: hemolytic action demonstrated in acute and repeated dose toxicity studies (summarized in EU risk assessment, 2008)

- DEGBE: i.e. in 2 and 13 week oral toxicity studies (Johnson et al. 2005)

- DEGDBE: in 28-day study (Clariant own data) RBC reduction and hematuria was evident.

- PolyEGDBE: in dose-range finding study for OECD 422 (Clariant own data), RBC reduction was evident together with compensatory increased hematopoietic activity.

 

b) Repeated dose toxicity of DEGDBE based on category building

 

Data availability on DEGDBE is limited to one 28 -day oral toxicity study. The toxic effects found at 750 mg/kg bw comprised alteration in hematology and effects on liver, spleen, and kidney. Some of these effects were present in rats treated with 250 mg/kg bw, and thus considered to be possibly adverse. The NOAEL of 100 mg/kg bw was identified.

In order to derive the chronic toxicity of DEGDBE, following aspects will be discussed.

b1) comparative findings in studies with DEGDBE and in NTP studies with EGBE (NTP, 2000)

b2) comparative findings in studies with DEGDBE and 13 -week oral toxicity study with DEGBE (Johnson et al., 2005)

b3) systemic burden of 2 -butoxyacetic acid in rats treated for 8 days, 28 days and after recovery of 14 days

b4) assessment of DEGDBE based on the EU assessment on EGBE and DEGBE

Relevant data are provided as endpoint study records.

 

b1) comparative findings in studies with DEGDBE and in NTP chronic toxicity studies with EGBE (NTP, 2000)

Rats were exposed to EGBE by inhalation up to concentrations of 500 ppm for 14 week or up to concentrations of 125 ppm for 104 weeks. The study identified hemolysis as primary toxic effect. Further affected organs were liver, spleen and thymus. Comparable findings were obtained in the 28-day repeated dose toxicity with DEGDBE. Especially the hematology alteration patterns were identical: reduced RBC, increased or decreased Hgb, increased Hct, increased MCV, increased MCH and decreased MCHC.

 

b2) comparative findings in studies with DEGDBE and 13 -week oral toxicity study with DEGBE (Johnson et al., 2005)

Rats were exposure to DEGBE for 13 weeks via drinking water. The effects found in this study are consistent to those found in the 28 -day study with DEGDBE, namely hematology alteration, oragan weight changes for liver and kidney. The major metabolite of DEGBE is butoxyethoxyacetic acid and its potency towards hemolysis is reported to be far lower (Udden, 2005) than that of 2 -butoxyacetic acid, explaning the enhanced toxicity potency of DEGDBE when compared to DEGBE.

 

b3) systemic burden of 2 -butoxyacetic acid in rats treated for 8 days, 28 days and after recovery of 14 days

In the 28 -day study the 24 -hours urine samples were collected after 8 day, 28 days and after reovery of 14 days (the endpoint study record provided under 7.1. in IUCLID) and these samples were analyzed for 2-butoxyacetic acid concentratiions. Comparable values were obtained in the samples of 8 and 28-day treatment and very low levels in the samples of recovery animals. These values are demonstrating that the systemic exposure is not increasing after 8 days exposure and that the bioaccumulating effect cannot be expected. Based hereupon, it can be derived that the toxicity found in the 28 -day studies is likely to reflect the chronic toxicity as well.

 

b4) assessment of DEGDBE based on the EU assessment on EGBE and DEGBE

According to the official EU documents (EU Risk Assessment 2 -Butoxyethanol Cas 111 -76 -2, ECB, 2008; Opinion on Diethylene Glycol Monobutyl Ether (DEGBE) SCCP/1043/06, 2006) EGBE or DEGDBE are agents causing hemolysis in rats and mice. Humans are likely to be more resistent, so that toxic effects in such a degree found in the animals studies cannot be expected. Based on the category building it is justified to apply the same consideration also for DEGDBE. The chronic toxicity of DEGDBE for humans is expected to be low.

Based on the provided data in b1)-b4), together with the justification for category building, the chronic toxicity of DEGDBE can be reasonably derived. No further animal testing is recommended.

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Well performed Guideline study on registration substance

Justification for classification or non-classification

DEGDBE induced hemolytic effects in rats upon repeated exposure, the mode of action being the systemic exposure to 2 -butoxyacetic acid. Based on the existing knowledge that humans are less susceptible to 2 -butoxyacetic acid, no significant hazard can be derived for the endpoint chronic toxicity or specific target organ toxicity for humans.

No classification is warranted.