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Administrative data

Description of key information

The read across results indicate that the repeated dose toxicity of hydrogen peroxide – urea (1:1) relies on the toxicity of the breakdown product, hydrogen peroxide, as this is the case for acute toxicity. The lower NOAEL value for male mice (26 mg/kg bw/day) is taken for the calculation of a NOAEL for the target substance (71.8 mg hydrogen peroxide - urea (1:1)/kg bw/d) and derivation of DNEL values.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
ANALOGUE APPROACH JUSTIFICATION
Please refer to the attached read across justification in section 13.
Reason / purpose:
read-across source
Clinical signs:
not specified
Mortality:
mortality observed, non-treatment-related
Description (incidence):
ne male mouse died in the control group (the cause of death was undetermined), and one male mouse in the 3000 ppm group died on study day 43 (no histopathological findings).
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weights were significantly reduced only in male and female animals receiving 3000 ppm.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Dose-related reductions in both food and water consumption were observed in female animals receiving 300 ppm and greater, while among the males consistent reductions were observed at the top dose level.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
Dose-related reductions in both food and water consumption were observed in female animals receiving 300 ppm and greater, while among the males consistent reductions were observed at the top dose level
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Hydrogen peroxide related changes were observed only in the duodenum at terminal sacrifice in the 1000 and 3000 ppm groups of males and females, and in a single 300 ppm group male. Although the general architecture of the affected duodenum was normal, there was an increase in cross sectional diameter and a larger mucosal area with broader, more substantial villi when compared to those of control mice. The change was assessed as mucosal hyperplasia because of the increase in mucosal thickness and size of the villi.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Mucosal hyperplasia in the duodenum in animals at 1000 or 3000 ppm, and in one male at 300 ppm
Histopathological findings: neoplastic:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
100 ppm
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
food consumption and compound intake
water consumption and compound intake
gross pathology
histopathology: non-neoplastic
Key result
Dose descriptor:
NOAEL
Effect level:
26 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male
Basis for effect level:
body weight and weight gain
food consumption and compound intake
water consumption and compound intake
gross pathology
histopathology: non-neoplastic
Key result
Dose descriptor:
NOAEL
Effect level:
37 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
female
Basis for effect level:
body weight and weight gain
food consumption and compound intake
water consumption and compound intake
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
300 mg/L drinking water
System:
gastrointestinal tract
Organ:
duodenum
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
yes

Results are for the source substance hydrogen peroxide. Read across to the target substance can be made taking the molecular weights into account which results in a factor of 2.76. Resulting NOAEL values for the target substance:

male mice:      71.8 mg/kg bw /day

female mice:   102.1 mg/kg bw /day

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
71.8 mg/kg bw/day
Study duration:
subchronic
Species:
mouse
Quality of whole database:
Source substance data suffiicient for read across and assessment. NOAEL derivation based on data from H2O2 (worst case).
System:
gastrointestinal tract
Organ:
duodenum

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: dermal
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
ANALOGUE APPROACH JUSTIFICATION
Please refer to the attached read across justification in section 13.
Reason / purpose:
read-across source
Clinical signs:
no effects observed
Dermal irritation:
no effects observed
Description (incidence and severity):
No erythema, scab and other irritative response were noted at the application site in any of the experimental groups including the control group of the 4-week and the 25-week-study
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:
no effects observed
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Immunological findings:
no effects observed
Description (incidence and severity):
no change in the albumin /globulin ratio in either study
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Description (incidence and severity):
behaviour was unchanged in all treated groups
Histopathological findings: non-neoplastic:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
338.4 mg/kg bw/day
Based on:
test mat.
Remarks:
hydroegn peroxide- urea (1:1)
Sex:
male/female
Basis for effect level:
other: no adverse effects noted
Remarks on result:
other:
Remarks:
calculated for the target substance from the NOAEL for urea (source), taking into account molecular weights
Key result
Critical effects observed:
no

The NOAEL for the target substance (338.4 mg/kg bw and day) was calculated from the NOAEL value for urea (source; 216 mg/kg bw and day), taking into account molecular weights of 94 for the target substance and 60 for the source substance.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
338.4 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
Source substance data suffiicient for read across and assessment. NOAEL derivation based only on data from urea.

Repeated dose toxicity: dermal - local effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: dermal
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
ANALOGUE APPROACH JUSTIFICATION
Please refer to the attached read across justification in section 13.
Reason / purpose:
read-across source
Clinical signs:
no effects observed
Dermal irritation:
no effects observed
Description (incidence and severity):
No erythema, scab and other irritative response were noted at the application site in any of the experimental groups including the control group of the 4-week and the 25-week-study
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:
no effects observed
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Immunological findings:
no effects observed
Description (incidence and severity):
no change in the albumin /globulin ratio in either study
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Description (incidence and severity):
behaviour was unchanged in all treated groups
Histopathological findings: non-neoplastic:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
338.4 mg/kg bw/day
Based on:
test mat.
Remarks:
hydroegn peroxide- urea (1:1)
Sex:
male/female
Basis for effect level:
other: no adverse effects noted
Remarks on result:
other:
Remarks:
calculated for the target substance from the NOAEL for urea (source), taking into account molecular weights
Key result
Critical effects observed:
no

The NOAEL for the target substance (338.4 mg/kg bw and day) was calculated from the NOAEL value for urea (source; 216 mg/kg bw and day), taking into account molecular weights of 94 for the target substance and 60 for the source substance.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
16.92 mg/cm²
Study duration:
subchronic
Species:
rat
Quality of whole database:
Source substance data suffiicient for read across and assessment. NOAEL derivation based only on data from urea.

Mode of Action Analysis / Human Relevance Framework

The target substance dissolves in water and breaks down to hydrogen peroxide and urea. Toxicity is governed by hydrogen peroxide whereas oral or dermal toxicity of urea is negligible. As a metabolite of the intermediary metabolism, urea is produced and excreted mainly via urine in quantities of 25-30 g/day in humans (i.e. endogenous urea production is approx. 500 mg/kg bw/day). Excess exogenous urea is rapidly excreted even at large doses, as shown in ADME studies.

Hydrogen peroxide is also generated at in the intermediary metabolism and in the oxidative burst of immune cells. Concentrations are low and mainly enzymatically controlled (glutathione peroxidase, catalase). Hydrogen peroxide is an oxidant that may cause local irritation and corrosion at higher concentrations. External hydrogen peroxide is stable enough to be absorbed through the skin or mucous membranes and distributed with the blood stream to other sites. It is decomposed predominantly by glutathione peroxidase at low concentrations and by catalase at higher concentrations. The latter reaction leads to the formation of large amounts of oxygen (1 ml of 30% hydrogen peroxide may liberate 100 ml of oxygen [ECB, 2003; DFG, 2006]) which may cause embolism with ischaemia, local necrosis, and death. Accidental cases have been reported in the open literature.

Additional information

No repeated dose study could be located for hydrogen peroxide – urea (1:1) but since this substance readily dissolves in water and breaks down to the two components, hydrogen peroxide and urea, these can be used as source substance and data can be read across to the target substance, hydrogen peroxide – urea (1:1).

Hydrogen peroxide was subject of the EU priority substance program. The Risk Assessment Report provides detailed information on all aspects relevant for the risk assessment, including repeated dose toxicity. Of all studies addressed for this endpoint, the 90-day drinking water study using catalase-deficient male and female mice is considered to be the most reliable one.

Reportedly, the sub-chronic toxicity of hydrogen peroxide was examined in a drinking water study (FMC, 1997; dose levels: 0, 100, 300, 1000, and 3000 ppm) using catalase-deficient male and female mice (15/sex/group). At termination after 90 days, 10 animals/sex/group were sacrificed and examined (blood, body weights, gross pathology, histopathology. The latter included all gross lesions, tongue, oesophagus, stomach, duodenum, ileum, jejunum, caecum, colon, and rectum from all animals in all groups, and on all major organs including the sex organs in the high dose and control animals). The remaining 5 animals/sex/group continued on distilled water during a 6-week recovery period.

Effects during the treatment period (Days 0-90): There were no treatment-related deaths. Clear treatment-related, dose-dependent effects were noted among both females and males receiving 300, 1000 or 3000 ppm of H2O2. Body weights were significantly reduced only in male and female high-dose animals receiving 3000 ppm. Dose-related reductions in both food and water consumption were observed in female animals receiving 300 ppm and greater, while among the males consistent reductions were observed only at the top dose level. Among females 300 ppm (103 mg/kg/day) was a LOAEL based on significant reduction in water consumption. Histopathology revealed treatment-related effects only in the duodenum of males and females at 1000 and 300 ppm, and in one male at 300 ppm. Although the general architecture of the affected duodenum was normal, there was an increase in cross sectional diameter and a larger mucosal area with broader, more substantial villi when compared to those of control mice. The change was assessed as mucosal hyperplasia because of the increase in mucosal thickness and size of the villi. Mucosal hyperplasia was not found in 100 ppm group mice, neither among controls. No other histopathological changes were noted on the tongue, esophagus, stomach, duodenum, ileum, jejunum, caecum, colon, and rectum from all animals in all groups, and on all major organs including the sex organs in the high dose and control animals.

Recovery period: the most notable effect was increased water consumption observed in males that had received 3000 ppm, and among females that had received 300, 1000, or 3000 ppm. Mucosal hyperplasia was reversible, as no such lesion was seen in any dose group after the recovery period.

The NOAEL was 100 ppm (26 and 37 mg/kg bw/day) in this study for males and females, respectively, based on dose-related reductions in water and food consumption, and on the observation of duodenal mucosal hyperplasia (ECB, 2003).

 

Urea

The oral repeated dose toxicity of urea was examined in two 12 -month carcinogenicity screening assay. One study was performed with F344 rats and one study with C57BL/6 mice. The rodents (50/sex/group) were exposed to urea in the diet at concentrations of 4500, 9000 or 45000 ppm for 12 months. Five animals/sex/group were sacrificed at the end of the 365-day exposure period and a comprehensive list of tissues were investigated histopathologically; interim deaths were similarly investigated. All remaining animals were sacrificed after the 4-month recovery period and investigated histopathologically.

There were no signs of toxicity. Survival and bodyweights were unaffected by treatment. Gross and microscopic pathology did not reveal any treatment-related effects. The NOAEL for rats and mice was determined to be 45000 ppm.

The repeated dose dermal toxicity of urea was examined in male and female Wistar rats in a 4-week study (15 rats per dose and sex) and in a 25-week study (10 rats per dose and sex). An ointment containing urea at 0, 10, 20, and 40% was applied daily to the dorsal skin, dose levels used were 0, 56, 108, and 216 mg/kg bw and day. Examinations included observation of clinical signs and behaviour, food and water intake, body weight development, clinical chemistry, haematology, urinalysis, gross pathology, examination of organ weights, and histopathology at termination.

 

Several effects were seen in animals of the intermediate dose group, but there was no dose-related adverse effect or pathological change in body weights, food and water consumption, or effects on any organ, clinical chemistry, haematology or urinalysis parameters. Further, no local effects on the skin were seen at the application sites. Thus no toxicity was seen at either 4 or 25 weeks with urea doses up to and including 216 mg/kg bw and day which was the NOAEL in this study (Sato et al., 1977).

 

Read across

The studies above are considered to be valid and suitable for assessment. The results can be read across to hydrogen peroxide – urea (1:1), taking the molecular weights into account.

Source substance hydrogen peroxide:

Read across to the target substance can be made taking the molecular weights into account which results in a factor of 2.76.

Resulting NOAEL values for oral repeated dose toxicity for the target substance (source substance: hydrogen peroxide):

male mice:      subchronic NOAELmouse,oral  = 71.8 mg hydrogen peroxide – urea (1:1)/kg bw /day

female mice:   subchronic NOAELmouse,oral  = 102.1 mg hydrogen peroxide – urea (1:1)/kg bw /day

Resulting NOAEL values for dermal repeated dose toxicity for the target substance (source substance: urea):

The subchronic NOAELrat, dermal for urea was 216 mg/kg bw and day. Taking into account molecular weights of 94 for the target substance and 60 for the source substance, this results in a subchronic NOAELrat, dermal of 338.4 mg hydrogen peroxide – urea (1:1)/kg bw /day

Conclusions

The read across results indicate that the repeat dose toxicity of hydrogen peroxide – urea (1:1) relies on the toxicity of the breakdown product, hydrogen peroxide, as this is the case for acute toxicity. The lower value for male mice (26 mg/kg bw/day) is taken for the calculation of a NOAEL for the target substance (71.8 mg hydrogen peroxide - urea (1:1)) and derivation of DNEL values.

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on these data the substance is not considered to be classified for repeated dose toxicity under Regulation (EC) No 1272/2008, as amended for the tenth time in Regulation (EC) No 2017/776.