Dibenzyl ether

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

Benzyl alcohol, benzoic acid and its sodium and potassium salt can be considered as a single category regarding human health, as they are all rapidly metabolised and excreted via a common pathway within 24hrs (OECD SIDS for Benzoates). In a 4-generation study with benzoic acid no effects on reproduction were seen (NOAEL 750 mg/kg). No compound related effects on reproductive organs (gross and histopathology examination) could be found in the (sub) chronic studies in rats and mice with benzyl acetate, benzyl alcohol, benzaldehyde, sodium benzoate and supports a non-reprotoxic potential of these compounds. In addition, data from reprotoxicity studies on benzyl acetate (NOAEL >2000 mg/kg bw/d; rats and mice) and benzaldehyde (tested only up to 5 mg/kg bw; rats) support the non-reprotoxicity of benzyl alcohol and benzoic acid and its salts (OECD SIDS for Benzoates).

Link to relevant study records

Reference

Endpoint:

multi-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Maximum reliability for read across-studies Critical study in OECD risk assessment, 2004: Meets generally accepted scientific standards, well documented and acceptable for assessment

Principles of method if other than guideline:

4-generation diet-study. Cited from OECD risk assessment: "A robust protocol, according to standards at that time, was used. Taking into account the reputation of the investigators a high quality has to be assumed."

GLP compliance:

no

Limit test:

no

Species:

rat

Strain:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

initial body weight:40-50 g

Route of administration:

oral: feed

Details on exposure:

Oral feed (first 8 weeks paired feed technique; afterwards ad libitum)

Duration of treatment / exposure:

Exposure period: generation 1 and 2: lifelong; generation 3: 16 weeks; generation 4: until breeding

Frequency of treatment:

Continuously in diet

Details on study schedule:

Lifelong (4-generations)

Remarks:

Doses / Concentrations:
approx. 375 and 750 mg/kg bw (0.5 and 1 % in diet)
Basis:

No. of animals per sex per dose:

20

Control animals:

yes

Details on study design:

The mean compound consumption was calculated according to Lehman, A.J., Assoc. Food Drug Off. Q. Bull. 18, 66 (1954).

Dose descriptor:

NOAEL

Effect level:

>= 750 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No indications for parental toxicity at highest dose of 750 mg/kg; reference: OECD risk assessment, 2004

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

>= 750 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No indications for fetal toxicity at highest dose of 750 mg/kg; reference: OECD risk assessment, 2004

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

>= 750 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: No indications for fetal toxicity at highest dose of 750 mg/kg; reference: OECD risk assessment, 2004

Reproductive effects observed:

not specified

"In all 4 generations no influence on growth (weight, weight gain and food efficiency (measured by protein efficiency) and organ weights was found. In all 4 generations, no effects on fertility ("Fortpflanzung") and lactation ("Aufzucht der Jungen") was found. The animals of the 3rd generation were sacrificed and examined histopathologically after 16 weeks (after lactation of the pups). No remarkable histopathological findings were found.

In the paper no information is given on the organs investigated, however the robustness of the total study, the reputation of the investigators, as well as the reputation of the Professor who did the histopathologic investigation, a high quality has to be assumed.

From other parameters it can be assumed that as a minimum the brains, heart, liver, kidney, testis and were examined.

Feeding of 0.5 % led to prolongation of survival compared to controls. In addition a so called "Alters-Paarung" after 48 weeks gave no influence on start of menopause."

Executive summary:

Cited from OECD 2004: "In a 4-generation study 20 rats/sex/group were dosed continuously by diet with 375 or 750 mg/kg/day benzoic acid. In all 4 generations, no effects on fertility (“Fortpflanzung”) and lactation (“Aufzucht der Jungen”) were found. In addition a so-called “Alters-Paarung” after 48 weeks gave no influence on start of menopauze. NOAEL (Parental) > 750 mg/kg/day, NOAEL (F1 Offspring) > 750 mg/kg/day, NOAEL (F2 Offspring) > 750 mg/kg/day."

(Kieckebusch & Lang, 1960, OECD 2004)

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

750 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

Scientifically acceptable and sufficiend documented for evaluation.

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Dibenzyl ether (CAS 103-50-4)_Read-across Justification Reproductive toxicity

No studies on reproduction are available for dibenzyl ether (CAS 103-50-4).

For substances manufactured or imported in quantities of 10 tones or more, a screening test for reproductive/developmental toxicity (OECD 421 or 422) is required according REACH Annex VIII (REGULATION (EC) No 1907/2006).

This study does not need to be conducted if a pre-natal developmental toxicity study or a EOGRTS (Extended One-Generation Reproductive Toxicity Study ) is available (COMMISSION REGULATION (EU) 2015/282 of 20 February 2015 amending Annexes VIII, IX and X to Regulation (EC) No 1907/2006 ), Annex VIII, 8.7.1 column 2).

According to REACH Annex XI the standard testing regime set out in Annexes VII to X can be adapted if testing does not appear scientifically necessary.

This is the case for substances of structural similarity where a read-across is justified. The similarity may be based on a common functional group (Annex XI, 1.5 [1]) or common precursors and/or the likelihood of common breakdown products via physical and biological processes, which result in structural similar chemicals (Annex XI, 1.5 [2]).

The following information it taken into account to waive the screening test for reproductive/ developmental toxicity (OECD 421 or 422).

• A 4-generation study with benzoic acid a predicted metabolite of dibenzyl ether
• A screening test for reproductive/developmental toxicity (OECD 422) with 4-hydroxybenzoic acid a predicted metabolite of dibenzyl ether.
• A Prenatal Developmental Toxicity Study (OECD 414) with ditolyl ether a chemically related compound
• Detailed information on reproductive organ toxicity in repeated dose toxicity studies

1) Waiving according to Annex XI, 1.5 [2] based on read-across with the 4-generation study of benzoic acid and the reproductive/developmental screening test of 4-hydroxybenzoic acid

Dibenzyl ether is predicted to metabolize into benzylalkohol, benzaldehyde, benzoic acid and hydroxylated compounds e.g. 4-hydroxybenzoic acid. Taking into account all available data on reproductive toxicity/fertility and on repeated dose toxicity (examinations of reproductive organs) for dibenzyl ether, the benzoates group and 4-hydroxybenzoic acid there is sufficient and reliable data, including a reliable 4-generation reproduction study with benzoic acid and an OECD combined repeat dose and reproductive/developmental toxicity study with 4-hydroxybenzoic acid to conclude that the predicted metabolites are not toxic to fertility.

2) Waiving based on read-across with ditolyl ether (Annex XI 1.5 [1]) and use of a pre-natal developmental toxicity study (Annex VIII, 8.7.1 column 2).

Dibenzyl ether is chemically similar to ditolyl ether based on structural similarity and a common functional ether group. A read-across between dibenzyl ether and ditolyl ether is justified based on chemical and toxicological similarities. Although no screening test for reproductive/developmental toxicity is available for both compounds a comprehensive developmental toxicity study according to OECD Guideline 414 is available for ditolyl ether. Since no screening test for reproductive/developmental toxicity (OECD 421 or 422) is required according REACH Annex VIII when a pre-natal developmental toxicity study is available, the developmental toxicity data of ditolyl ether are very relevant to waive the screening test for reproductive/developmental toxicity.

3) Waiving based on Weight-of-evidence with available reproductive data on dibenzyl ether (Annex XI, 1.2)

In addition, there are no indications of reproductive toxicity in comprehensive repeated dose toxicity studies with dibenzyl ether investigating the reproductive organ weights and histopathology.

Overall conclusion:

The screening test for reproductive/ developmental toxicity (OECD 421 or 422) for dibenzyl ether is waived based on the above mentioned scientific rational.

Detailed justification:

No indication of reproductive toxicity in comprehensive repeated dose toxicity studies with dibenzyl ether (CAS 103-50-4):

Dibenzyl ether has a low acute toxicity for oral and dermal toxicity. The LD50 are > 2000 mg/kg bw.

In a sub-chronic oral feed study of 91 days, No clinical signs of toxicity were observed during the course of the study; there were no deaths. There was a decrease in erythrocyte count, haemoglobin and haematocrit in mid- and high-dose females at the 6-wk interval, which appeared to be resolving at the 12-wk interval, with the exception of the haematocrit in the mid- and high-dose females and erythrocyte counts in the mid-dose females. As some of the decreases appeared to resolve during the intervening 6 wk, and since the erythrocyte count and haematocrit were significantly elevated in males and the values were within normal limits for the laboratory for this strain, these and other effects in the haemograms were considered to be transient and not toxicologically significant.

There was a statistically significant increase in absolute and relative liver weight in high-dose females at study termination. Other increases occurred sporadically throughout the groups of females in one or another of the paired organs and were not considered to be biologically significant.

No macroscopic or microscopic abnormalities were seen that could be attributed to treatment with the test substance. The NOAEL was 620 mg/kg bw/day.

There were no observations on organ weight or histopathology on testes (with adnexa) and ovaries. Overall there is no indication of reproductive toxicity in this sub-chronic toxicity study with dibenzyl ether.

There is no indication of reproductive toxicity in predicted metabolites:

The metabolism of dibenzyl ether can be predicted by the QSAR toolbox Version 3.2 (See chapter Toxicokinetics and separate expert statement; Schlecker, 2014). The autoxidation and hydrolysis simulator within the toolbox indicate benzyl alcohol (CAS No 100-51-6), benzyl aldehyde (CAS 100-52-7) and benzoic acid (CAS 65-85-0) as potential metabolites.

Benzylalkohol, benzaldehyde and benzoic acid are considered as a category of substances with similar toxicological and metabolic properties by different expert committees (e.g. OECD HPV program 2004, IPCS 2005, SCF 2002).

The OECD SIDS for ”Benzoates” UNEP publication (2004) concludes the following:

Benzyl alcohol, benzoic acid and its sodium and potassium salt can be considered as a single category regarding human health, as they are all rapidly metabolised and excreted via a common pathway within 24hrs. Systemic toxic effects of similar nature (e.g. liver, kidney) were observed.

For benzoic acid repeated dose oral toxicity studies give a NOAEL of 800 mg/kg/day. For the salts values > 1000 mg/kg/day are obtained. At higher doses increased mortality, reduced weight gain, liver and kidney effects were observed. For benzyl alcohol the long-term studies indicate a NOAEL > 400 mg/kg bw/d for rats and > 200 mg/kg bw/d for mice. At higher doses effects on bodyweights, lesions in the brains, thymus, skeletal muscle and kidney were observed. It should be taken into account that administration in these studies was by gavage route, at which saturation of metabolic pathways is likely to occur. It can be concluded that benzoic acid and its salts exhibit very low repeated dose toxicity. Benzyl alcohol exhibits low repeated dose toxicity.

OECD risk assessment (2004) certifies that “In addition no compound related effects on reproductive organs (gross and histopathology examination) could be found in the (sub) chronic studies in rats and mice with benzyl acetate, benzyl alcohol, benzaldehyde, sodium benzoate…” This is further evidenced by the fact that most of the sub-chronic dose studies available were conducted including investigations on sperm morphology and vaginal cytology (Morrissey et al 1988).

Of most value is an early four-generation study with benzoic acid (Kieckebusch & Lang, 1960; relevant study in OECD risk assessment on benzoates group, 2004). Cited from OECD 2004: "In a 4-generation study 20 rats/sex/group were dosed continuously by diet with 375 or 750 mg/kg/day benzoic acid.” The OECD certifies the four-gen. study that “A robust protocol, according to standards at that time, was used. Taking into account the reputation of the investigators a high quality has to be assumed." The study revealed no effects on fertility and lactation in all 4 generations. Additionally the so-called “Alters-Paarung” after 48 weeks gave no influence on start of menopause. According to OECD 2004 the quantitative results were NOAEL (Parental) > 750 mg/kg/day, NOAEL (F1 Offspring) > 750 mg/kg/day, NOAEL (F2 Offspring) > 750 mg/kg/day.

Another study on Fertility is available for benzaldehyd. Cited from OECD 2004: "A single study was conducted to examine the potential reproductive toxicity of benzaldehyde, and the report was available as a translation from Romanian. A group of 10 rats of breeding age were given 2 mg benzaldehyde in oil (type not specified) by gavage every other day for 32 weeks, equivalent to about 5 mg/kg bw per day. Ten controls were used. Two pregnancies in each rat, one at 75 days and one at 180 days, were studied. The end-points examined included the number of pregnant females, number of offspring born, pup body weight at days 7 and 21 post partum, and pup viability.

At the end of treatment, the body weights of control and treated rats were similar: 265 g and 260 g, respectively. It was reported that fewer females in the group given benzaldehyde than in the control group became pregnant; however, no data or statistical analyses were presented. The authors concluded that treatment did not significantly modify any of the parameters studied. No further details were available. The NOAEL was about 5 mg/kg bw per day" (Sporn et al. 1967, cited from OECD 2004).

Taking into account all available data on reproductive toxicity/fertility and on repeated dose toxicity (examinations of reproductive organs) for the benzoates group there is sufficient and reliable data, including a reliable 4-generation reproduction study on benzoic acid, to conclude that this group of compounds is not toxic to fertility.

The liver metabolism of dibenzyl ether can be predicted by the QSAR toolbox Version 3.2 (See chapter Toxicokinetics and separate expert statement; Schlecker, 2014). The predicted metabolites have hydroxylgroups in meta or para position in one or both aromatic rings or two hydroxyl-groups in meta/para-position to the ether bound in one aromatic ring. There are fertility data available for one hydroxylated compound, 4-hydroxybenzoic acid (CAS No 99-96-7) and the OECD SIDS; UNEP publication (1999), summarizes the toxicity and fertility as follows:

Oral LD50 of 4-hydroxy benzoic acid for rats is more than 2,000 mg/kg. This chemical is considered to be slightly irritating to skin and moderate to eyes, and a mild skin sensitizer. In an OECD combined repeat dose and reproductive/developmental toxicity study in rats at 40, 200 and 1,000 mg/kg/day, this chemical induced rale and rhinorrhea, indicative of imitation to respiratory tract irritation, and small fluctuation of blood chemistry with no changes of histopathological findings and organ weights. These changes of blood chemistry are considered not to be adverse.

Therefore, no sign of toxic effects in repeated dose toxicity testing were detected at the highest dose of 1,000 mg/kg/day. Reproductive toxicity was not observed up to the highest test dose of 1000 mg/kg/day, suggesting no reason for concern.

Conclusion on fertility assessment based on read across to predicted metabolites:

Taking into account all available data on reproductive toxicity/fertility and on repeated dose toxicity (examinations of reproductive organs) for dibenzyl ether, the benzoates group and 4-hydroxybenzoic acid there is sufficient and reliable data, including a reliable 4-generation reproduction study with benzoic acid and an OECD combined repeat dose and reproductive/developmental toxicity study with 4-hydroxybenzoic acid to conclude that the predicted metabolites are not toxic to fertility.

Additional information on ditolyl ether (CAS 28299-41-4), a chemically related compound, supports that dibenzyl ether is not toxic to reproduction:

Dibenzyl ether is chemically quiet similar to ditolyl ether based on structural similarity and a common functional group. This assumption is supported by similar toxicological results of toxicological studies in mammalians (see attached data matrix).

Therefore a read-across with ditolyl ether is justified and data of reproduction studies can be conveyed from ditolyl ether to dibenzyl ether.

In a Prenatal Developmental Toxicity Study (according to OECD Guideline 414) 25 inseminated Wistar rats per group were orally administered daily doses of 0, 100, 300, and 1000 mg kg bw on days 6-15 of pregnancy. Dams were examined regarding body weight, appearance and behaviour. On day 20 of gestation dams were killed and the foetuses delivered by caesarean section were examined for morphological changes. In all dose groups, no indications of teratogenesis were observed. At 1000 mg/kg bw/d, there were signs of maternal toxicity. Signs of embryotoxicity were reduced number and decreased weights of the fetuses at the high dose level.

Doses up to and including 300 mg/kg bw/g were tolerated without any signs of maternal toxicity and without toxic signs on embryonal and foetal development. Result: NOEL = 300 mg/kg bw/day for maternal and embryonal/foetal toxicity (Renhof, 1986).

In conclusion, a read-across between dibenzyl ether and ditolyl ether is justified based on chemical and toxicological similarities.

Although no screening test for reproductive/ developmental toxicity (OECD 421 or 422) is available for both compounds a comprehensive developmental toxicity study according to according to OECD Guideline 414 is available for ditolyl ether. Since no screening test for reproductive/developmental toxicity (OECD 421 or 422) is required according REACH Annex VIII when a pre-natal developmental toxicity study is available, the developmental toxicity data of ditolyl ether are very relevant to waive the screening test for reproductive/developmental toxicity.

Data Matrix, Analogue Approach:

	Target Chemical	Source chemical
CAS #	103-50-4	28299-41-4
Chemical Name	dibenzyl ether	ditolyl ether
Physico-Chemical Data
Physical state at 20°C and 101.3 kPa	Liquid at 20°C (IUCLID 5.6.)	Liquid at 20°C
Appearance	colourless liquid (IUCLID 5.6.)	organic liquid at 20 °C and 101.3 kPa
Melting Point	4 °C at 1013 hPa (IUCLID 5.6.)	-
Boiling Point	298 °C at 1013 hPa (IUCLID 5.6.)	283 °C at 1013 hPa
Density	1.043 at 20°C (IUCLID 5.6.)	1.034 at 20°C
Vapour Pressure	0.00137 hPa at 25 °C (IUCLID 5.6.)	0.0012 hPa at 20 °C
Water Solubility	42 mg/L at 20 °C (IUCLID 5.6.)	2.82 mg/L at 20 °C
Pow	3.31 (IUCLID 5.6.)	4.3 at 25°C
pH value 20°C	-	-
Mammalian Toxicity
Dermal irritation/corrosion	weight of evidence: not irritating (Mihail 1977, Kynoch 1978)	not irritating (Maertins 1989)
Eye irritation	not irritating (Mihail 1977)	not irritating (Maertins 1989)
Dermal sensitization	weak sensitizer in guinea pig (Sharp 1978)(modified Draize technique)	ambiguous (Diesing , Flucke 1991)
Mutagenicity (bacteria)	Ames test: negative (Solokowski 2015)	Ames test: negative(Herbold 1982)
Mutagenicityin vitro cytogenetic assays in mammalian cells	MNT: negative(Bohnenberger 2015)	In vivo study available
Mutagenicityin vitro gene mutation assays in mammalian cells	HPRT Test: negative(Wollny 2015)	In vitro UDS: negative(Lehn 1990)
Mutagenicity in vivo assays	In vivo micronucleus test: negative (Wild 1983)
Acute toxicity (oral)	LD50rat: 3860 mg/kg bw (Kynoch et al. 1978)LD50 rat: ca. 4818 mg/kg bw (Loeser 1976)	LD50rat: ca. 3622 mg/kg bw (Loeser 1986) LD50mouse: 200 — 2000 mg/kg bw (Bomhard, Groening 1990)LD50 guinea pig: LD50: 200 — 2000 mg/kg bw (Bomhard , Groening 1990)LD50 hamster: > 2000 mg/kg bw (male)Weight of evidence: Xn, R22 for ditolylether is proposed by the author
Acute toxicity (inhalation)	No data	LC50 (7 h): > 521 mg/m³ air (Pauluhn 1984)
Acute toxicity (dermal)	LD50 rabbit > 5000 mg/kg bw(Kynoch et al. 1978)	LD50: > 2587 mg/kg bw (Kroetlinger 1984)
Repeated dose toxicity subacute	No data	Dose-range finding study (no NOAEL defined) (Kroetlinger 1984)
Repeated dose toxicity subchronic	Subchronic feeding study in rats, NOAEL : 620 mg/kg/day, (increased rel. and absolute liver weights), (Burdock 1992)	Subchronic feeding study in rats:NOAEL: 132 mg/kg bw/dayLOAEL: 425 mg/kg bw/d: decreased body weights, increased liver weights(Kroetlinger, Schilde 1988 (Ownership: Lanxess)
Screening test reproduction- /developmental toxicity	No data	No data
Developmental toxicity	No data	OECD TG 414 rat:NOAEL (maternal toxicity): 300 mg/kg bw/day LOAEL: 1000 mg/kg bw/day (body weight gain reduced during the whole gestation; treatment-related deaths)NOAEL (embryo toxicity): 300 mg/kg bw/dayLOAEL (embryotoxicity): 1000 mg/kg bw/day (reduced number and decreased weights of the fetuses(Renhof 1986)(Ownership: Lanxess)
Multigenerational study	No data	No data
Toxicokinetic	QSAR, Schlecker, 2015and Information/Assumptions regarding Toxicokinetics and metabolism (expert judgment)	Oral and ip. Application:biological half-life of about 1.5 hours, more than 90% of the intragastrically administered dose was excreted in urine and faeces within 3 days; > 80% dose in the urine(Law 1983)
Carcinogenicity study	No data	No data
	# Data from IUCLID5.6. data set Dibenzyl ether	# Data from ECHA disseminated dataset of Ditolyl ether (2015-02-11)

References:

OECD (Organisation for Economic Co-operation and Development), 2001, SIDS Initial Assessment Report for 13th SIAM (Bern, 7th – 9th November 2001) on Benzoates: Benzoic acid, Sodium benzoate, Potassium benzoate, Benzyl alcohol https://hpvchemicals.oecd.org/ui/handler.axd?id=aa89d225-a2a7-4ed5-b8d6-c06b5e30b45b

OECD (Organisation for Economic Co-operation and Development), 1999, SIDS Initial Assessment Report for for 9th SIAM (France, June 29-July 1, 1999) on 4-Hydroxybenzoic acid

https://hpvchemicals.oecd.org/ui/handler.axd?id=CD93235E-9715-4766-A0BC-4B219347AEF2

IPCS 2005 (International Programme on Chemical Safety) CICAD 26 (Concise International Chemical Assessment Document 26) (corrigendum 2005/ 2000) on benzoic acid and sodium benzoate, published under Joint Sponsorship of United Nations Environment Programme, International Labour Organization, and World Health Organization, Wissenschaftliche Verlagsgesellschaft mbH, D-70009 Stuttgart 10

SCF 2002. SCIENTIFIC OPINION

Flavouring Group Evaluation 54, Revision 1 (FGE.54Rev1) (2009):

Consideration of benzyl derivatives evaluated by JECFA (57th meeting) structurally related to benzyl alcohols, benzaldehydes, a related acetal, benzoic acids and related esters evaluated by EFSA in FGE.20Rev1 (2009) 1 Scientific Opinion of the Panel on Food Contact Materials, Enzymes, Flavourings and Processing Aids (CEF). The EFSA Journal (2009) 1025, 1-73

Mihail F, Plastikator BA (Dibenzylether) - Untersuchungen zur Haut- und Schleimhaut-vertraeglichkeit, Bayer AG - Institut fuer Toxikologie, Wuppertal-Elberfeld, Germany, Company study no. T 2050575 (1977)

Kynoch SR and Ligget MP, Irritant effects of oxyde de benzyle on rabbit skin, Ineos, Rport no. 8913/3D/78 (1978)

Sharp DW, The sensitization potential of some perfume ingredients tested using a modified Draize procedure, Toxicology 9, 261-271 (1978)

Sokolowski A, Dibenzyl ether, CAS 103-50-4: Salmonella typhimurium and Echerichia coli reverse mutation assay, Harlan Cytotest Cell Research GmbH (Harlan CCR), In den Leppsteinswiesen 19, 64380 Rossdorf/Germany, Report no. 1648301 (2015)

Bohnenberger S, Dibenzyl ether, CAS 103-50-4: Micronucleus Test in Human Lymphocytes In vitro, Harlan Cytotest Cell Research GmbH (Harlan CCR), In den Leppsteinswiesen 19 64380 Rossdorf, Germany, Report no. 1648303

Wollny HE, Dibenzylether: Gene mutation assay in Chinese hamster V79 Cells In vito (V79 / HPRT), Harlan Cytotest Cell Research GmbH (Harlan CCR), In den Leppsteinswiesen 19, 64380 Rossdorf, Germany, Report no. 1648302 (2015)

Wild DK, Gocke M, and Eckhardt K, Study of artificial flavoring substances for mutagenicity in the Salmonella/Microsome, BASC and micronucleus tests, Food Chem. Toxicol., 21, 707-719 (1983)

Kynoch SR, Lloyd GK and Andrews CD, Acute oral toxicity to rats of oxyde de benzyle, Report no. 9154/D5/78 (1978)

Loeser E, Akute orale Toxizitaet - Plastikator BA, short report, Bayer AG - Institur fuer Toxikologie - Wuppertal-Elberfeld, Germany, Company no. T 2049108 81976)

Kynoch SR and Lloyd GK, Acute dermal toxicity to rats of oxyde de benzyle, Huntingdon Research Centre, Report no. 9153/D6/78 (1978)

Burdock GA and Ford RA, Safety evaluation of dibenzyl ether, Food Chem Toxicol 30, 7, 559-566 (1992)

Maertins T, Ditolylether - study for skin and eye irritation/corrosion in rabbits according to OECD guideline no 404 and 405, Bayer AG, Report no. 18606 (1989)

Diesing L, Flucke W, Ditolylether - Untersuchungen auf hautsensibilisierende Wirkung bei Meerschweinchen (Maximierungstest nach Magnusson und Kligman sowie modifizierter Epikutantest nach Buehler), Bayer AG, Report no. 19986 A (1991)

Herbold B, Traenkmittel VP OC 4900 - Salmonella/Mikrosomen-Test zur Untersuchung auf punktmutagene Wirkung, Bayer AG , Institut für Toxikologie, 42096 Wuppertal , Germany, Report no. 11141 (1982)

Lehn H, Ditolylether - Mutagenicity test on unscheduled DNA synthesis in rat liver primary cell cultures in vitro, Bayer AG, Report no. 19446 (1990)

Herbold B, Baylectrol 4900 - Mikronucleus-Test an der Maus zur Pruefung auf mutagene Wirkung, Bayer AG, Report no. 12945 (1984)

Loeser E, Diphyl DT - Untersuchung zur akuten oralen Toxizitaet an maennlichen Wister-Ratten, Bayer AG, Report no. 15250 (1986)

Bomhard E, Groening P, Ditolylether - Vergleichende Untersuchungen zur akuten oralen Toxizitaet an maennlichen B6C3F-Maeusen, maennlichen syrischen Goldhamstern und Meerschweinchen, Bayer AG, Report no. 19746 (1990)

Pauluhn J, Baylectrol 4900 - Untersuchungen zur akuten inhalativen Toxizitaet (Inhalations-Risiko-Test), Bayer AG, Report no. 12793 (1984)

Kroetlinger F, Investigations of the acute dermal toxicity in male and female Wistar-rats, Bayer AG (1984)

Kroetlinger F, Baylectrol 4900 Orientierende toxikologische Untersuchungen zur Dosisfindung fuer einen subchronischen Versuch (Fuetterungsversuch ueber 3 Wochen), Bayer AG, company no. T3017429 (1984)

Kroetlinger F, Schilde B, Baylectrol - Subchronische toxikologische Untersuchungen an Ratten (Fuetterungsversuch ueber 3 Monate), Bayer AG, Report no. 16353 (1988)

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Law FCP, Song YY, Chakrabarti S, Disposition and metabolism of diphenylether in rats, Xenobiotica 13, 627-633 (1983)

Short description of key information:
Benzyl alcohol, benzoic acid and its sodium and potassium salt can be considered as a single category regarding human health, as they are all rapidly metabolised and excreted via a common pathway within 24hrs (OECD SIDS for Benzoates). In a 4-generation study with benzoic acid no effects on reproduction were seen (NOAEL 750 mg/kg). No compound related effects on reproductive organs (gross and histopatology examination) could be found in the (sub) chronic studies in rats and mice with benzyl acetate, benzyl alcohol, benzaldehyde, sodium benzoate and supports a non-reprotoxic potential of these compounds. In addition, data from reprotoxicity studies on benzyl acetate (NOAEL >2000 mg/kg bw/d; rats and mice) and benzaldehyde (tested only up to 5 mg/kg bw; rats) support the non-reprotoxicity of benzyl alcohol and benzoic acid and its salts (OECD SIDS for Benzoates).

Cited from OECD 2004: "In a 4-generation study 20 rats/sex/group were dosed continuously by diet with 375 or 750 mg/kg/day benzoic acid. In all 4 generations, no effects on fertility and lactation were found. NOAEL (Parental) > 750 mg/kg/day, NOAEL (F1 Offspring) > 750 mg/kg/day, NOAEL (F2 Offspring) > 750 mg/kg/day."

Justification for selection of Effect on fertility via oral route:
Benzoic acid is expected as a main metabolite of dibenzyl ether (see section 7.1 toxicokinetics, metabolism). Therefore data on reproduction studies can be conveyed from benzoic acid to dibenzyl ether by read-across.

Effects on developmental toxicity

Description of key information

Cited from OECD SIDS for Benzoates: "A study using pregnant Wistar rats, dosed with 700, 1400, 2800, 5600 mg/kg sodium benzoate in the diet during the entire gestation showed no statistical difference in organ and bone abnormalities of fetuses between experimental groups and controls; growth of treated offsprings was similar to controls in rats dosed with 1400 mg/kg/day; reduced food intake and decreased body weight of the pregnant rats especially in the 5600 mg/kg group; 100% perinatal death rate; organ abnormalities of fetuses involved eye, brain and kidneys, in addition abnormalities of the skeletal system were found in rats dosed with >2800 mg/kg/day. The authors concluded that the effects on the dams and fetuses at the 2800 and 5600 levels were due to reduced maternal feed intake in these groups, leading to malnutrition, NOAEL Maternal toxicity: 1400 mg/kg bw, NOAEL Teratogenicity: 1400 mg/kg bw. "

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Maximum reliability for read across-studies Critical study in OECD risk assessment, 2004.

Principles of method if other than guideline:

Study with pregnant Wistar rats, dosed with 700, 1400, 2800, 5600 mg/kg sodium benzoate in the diet during the entire gestation; examinations included body weight, food consumption, number of life and dead fetuses, visceral and skeletal investigations of fetuses.

GLP compliance:

not specified

Species:

rat

Strain:

Wistar

Route of administration:

oral: feed

Duration of treatment / exposure:

during entire gestation period

Frequency of treatment:

continuously in diet

Control animals:

yes

Details on study design:

Study with pregnant Wistar rats, dosed with 700, 1400, 2800, 5600 mg/kg sodium benzoate in the diet during the entire gestation; examinations included body weight, food consumption, number of life and dead fetuses, visceral and skeletal investigations of fetuses.

Dose descriptor:

NOAEL

Effect level:

1 400 mg/kg bw/day (nominal)

Basis for effect level:

other: maternal toxicity

Dose descriptor:

NOAEL

Effect level:

1 400 mg/kg bw/day (nominal)

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

No statistical difference in organ and bone abnormalities of fetuses between experimental groups and controls; growth of treated offsprings was similar to controls in rats dosed with 1400 mg/kg/day; reduced food intake and decreased body weight of the pregnant rats especially in the 5600 mg/kg group; 100% perinatal death rate; organ abnormalities of fetuses involved eye, brain and kidneys, in addition abnormalities of the skeletal system were found in rats dosed with >2800 mg/kg/day. The authors concluded that the effects on the dams and fetuses at the 2800 and 5600 levels were due to reduced maternal feed intake in these groups, leading to malnutrition.

Executive summary:

Cited from OECD 2004: "A study using pregnant Wistar rats, dosed with 700, 1400, 2800, 5600 mg/kg sodium benzoate in the diet during the entire gestation showed no statistical difference in organ and bone abnormalities of fetuses between experimental groups and controls; growth of treated offsprings was similar to controls in rats dosed with 1400 mg/kg/day; reduced food intake and decreased body weight of the pregnant rats especially in the 5600 mg/kg group; 100% perinatal death rate; organ abnormalities of fetuses involved eye, brain and kidneys, in addition abnormalities of the skeletal system were found in rats dosed with >2800 mg/kg/day. The authors concluded that the effects on the dams and fetuses at the 2800 and 5600 levels were due to reduced maternal feed intake in these groups, leading to malnutrition, NOAEL Maternal toxicity: 1400 mg/kg bw, NOAEL Teratogenicity: 1400 mg/kg bw."

(Onodera et al. 1978, OECD 2004)

Reference 2

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
[see 'Attached justification']

Reason / purpose for cross-reference:

read-across source

Duration of treatment / exposure:

during entire gestation period

Frequency of treatment:

continuously in diet

Dose descriptor:

NOAEL

Effect level:

1 400 mg/kg bw/day (nominal)

Basis for effect level:

other: maternal toxicity

Dose descriptor:

NOAEL

Effect level:

1 400 mg/kg bw/day (nominal)

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

No statistical difference in organ and bone abnormalities of fetuses between experimental groups and controls; growth of treated offsprings was similar to controls in rats dosed with 1400 mg/kg/day; reduced food intake and decreased body weight of the pregnant rats especially in the 5600 mg/kg group; 100% perinatal death rate; organ abnormalities of fetuses involved eye, brain and kidneys, in addition abnormalities of the skeletal system were found in rats dosed with >2800 mg/kg/day. The authors concluded that the effects on the dams and fetuses at the 2800 and 5600 levels were due to reduced maternal feed intake in these groups, leading to malnutrition.

Executive summary:

Cited from OECD 2004: "A study using pregnant Wistar rats, dosed with 700, 1400, 2800, 5600 mg/kg sodium benzoate in the diet during the entire gestation showed no statistical difference in organ and bone abnormalities of fetuses between experimental groups and controls; growth of treated offsprings was similar to controls in rats dosed with 1400 mg/kg/day; reduced food intake and decreased body weight of the pregnant rats especially in the 5600 mg/kg group; 100% perinatal death rate; organ abnormalities of fetuses involved eye, brain and kidneys, in addition abnormalities of the skeletal system were found in rats dosed with >2800 mg/kg/day. The authors concluded that the effects on the dams and fetuses at the 2800 and 5600 levels were due to reduced maternal feed intake in these groups, leading to malnutrition, NOAEL Maternal toxicity: 1400 mg/kg bw, NOAEL Teratogenicity: 1400 mg/kg bw."

(Onodera et al. 1978, OECD 2004)

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 400 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Scientifically acceptable and sufficient documented.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Benzyl alcohol, benzoic acid and its sodium and potassium salt can be considered as a single category regarding human health, as they are all rapidly metabolised and excreted via a common pathway within 24hrs (OECD SIDS for Benzoates).

In rats for sodium benzoate dosed via food during the entire gestation developmental effects occurred only in the presence of marked maternal toxicity (reduced food intake and decreased body weight) (NOAEL = 1400 mg/kg bw). For hamster (NOEL: 300 mg/kg bw), rabbit (NOEL: 250 mg/kg bw) and mice (CD-1 mice, NOEL: 175 mg/kg bw) no higher doses (all by gavage) were tested and no maternal toxicity was observed. For benzyl alcohol: NOAEL= 550 mg/kg bw (gavage; CD-1 mice). LOAEL = 750 mg/kg bw (gavage mice). In this study maternal toxicity was observed e. g. increased mortality, reduced body weight and clinical toxicology.

(OECD SIDS for Benzoates).

Additional informationon a chemically related compound supports the non-reprotoxicity of dibenzyl ether regarding developmental toxicity/teratogenicity:

Dibenzyl ether is quiet similar to ditolyl ether (CAS n° 28299-41-4) based on structural similarity and a common functional group. This assumption is supported by similar toxicological results of toxicological studies in mammalians (see attached data matrix).

Therefore a read-across with ditolylether is justified and data of reproduction studies can be conveyed from ditolyl ether to dibenzyl ether.

In a Prenatal Developmental Toxicity Study (according to OECD Guideline 414) 25 inseminated Wistar rats per group were orally administered daily doses of 0, 100, 300, and 1000 mg kg bw on days 6-15 of pregnancy. Dams were examined regarding body weight, appearance and behaviour. On day 20 of gestation dams were killed and the foetuses delivered by caesarean section were examined for morphological changes. In all dose groups, no indications of teratogenesis was observed. At 1000 mg/kg bw/d, there were signs of maternal toxicity. Signs of embryotoxicity were reduced number and decreased weights of the fetuses at the high dose level.

Doses up to and including 300 mg/kg bw/g were tolerated without any signs of maternal toxicity and without toxic signs on embryonal and foetal development. Result: NOEL = 300 mg/kg bw/day for maternal and embryonal/foetal toxicity (Renhof (Bayer AG), 1986).

Justification for selection of Effect on developmental toxicity: via oral route:
Benzoic acid is expected as a main metabolite of dibenzyl ether (see section 7.1 toxicokinetics, metabolism). Therefore data on reproduction studies can be conveyed from benzoic acid and sodium benzoate to dibenzyl ether by read-across. Studies were conducted in rats, rabits, mice and hamsters. For rats 2 stuides are available; in one study a NOAEL of > 175 mg/kg bw (highest applied dose) was found, whereas in the other study no indications for teratogenicity at a dose of 1400 mg/kg bw was determined, which was the highest dose without maternal toxicity (reference: OECD risk assessment, 2004).

Toxicity to reproduction: other studies

Additional information

No data.

Justification for classification or non-classification

The proposed metabolites of dibenzyl ether (benzoic acid and 4-hydroxybenzoic acid) are not toxic to reproduction.

Additionally ditolyl ether as a structural analogue or surrogate for dibenzyl ether is tolerated without any signs on embryonal and foetal development in doses without signs of maternal toxicity.

According to CLP classification criteria (Regulation (EC) No 1272/2008) a classification of dibenzyl ether for reprodution toxicity is not justified.