Erythritol

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Endpoint:

two-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1996

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)

GLP compliance:

not specified

Limit test:

no

Justification for study design:

Not specified

Specific details on test material used for the study:

Batch no. not specified
Erythritol was obtained from Cerestar, Vilvoorde Belgium. The test material had a white, crystalline appearance and was reported to have a water content of <0.5% and a purity of 99.9% on a dry matter basis. HPLC analysis determined that the actual dietary concentrations varied between 95 and 110% of the nominal concentrations.

Species:

rat

Strain:

Wistar

Remarks:

Wistar-derived strain

Details on species / strain selection:

Cri:WI(WU)BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

Source : Charles River Wiga GmbH, Sulzfeld, Germany
On arrival, the rats were about 5 weeks old
Diets : provided as a meal mash in stainless steel cans. The food was covered by a perforated stainless steel plate to prevent spillage. Diet : CIVO cereal-based stock diet, food and water available ad libitum

Acclimatization :
Duration : 2 weeks
Rats housed in groups of 4
Male/female separated
Cages : Suspended stainless steel group cages with wire mesh floor and front under conventional conditions
Temperature : 19–24.7°C
Relative Humidity : 40 and 70%.
Photoperiod : 12h light/dark cycle
Diet : defatted soya bean meal (11.0%), fish meal (7.0%), meat and bone scraps (4.0%), whole ground wheat (36.0%) and maize (29.7%), brewer’s yeast (3.0%), alfalfa meal (3.0%), whey powder (2.0%), defatted bone meal (0.4%), salt and vitamin premixes (0.6%), and soya bean oil (3.0%). Analysis : contain crude protein (20.2%), crude fat (6.3%), crude fiber (3.3%), moisture (10.4%), calcium (0.87%), phosphorus (0.72%), magnesium (0.17%), as well as minerals, trace elements, and vitamins in adequate amounts.

Premating period and thereafter until termination :
Premating period : 10 weeks
Diet of the control group : 87.4% cereal-based stock diet, 10% wheat starch, 2.5% casein, and 0.1% d,l-methionine.
Diet of the treatment group : The three erythritol dose groups received the same diet, to which 2.5, 5, or 10% erythritol was added at the expense of wheat starch.
Initial body weight : 176.3 g for males (149.8 to 206.3 g) and 135.5 g for females (119.4 to 158.7 g)

Route of administration:

oral: feed

Vehicle:

other: erythritol administered in diet

Details on exposure:

Erythritol mixed into the diet: 87.4% cereal-based stock diet, 10% wheat starch (to which 2.5, 5, or 10% erythritol was added at the expense of wheat starch), 2.5% casein and 0.1% d,l-methionine.

Animais received the test article for 10 weeks prior to mating, during mating, gestation, and lactation. During the premating period, food consumption measured weekly for each cages by weighing the feeders. Individual food consumption of all mated females was also recorded weekly during pregnancy and lactation.

Actual intakes, first and second generation :

first generation :
actual intakes lactation during premating : male : 1.4, 2.8, 5.95 g/kg body weight/day
actual intakes during premating : female: 1.69, 3.42, 6.68 g/kg body weight/day
actual intakes during gestation : female : 1.56, 3.27, 6.44 mg/kg body weight/day
actual intakes during lactation : female : 3.79, 7.43, 15.81 mg/kg body weight/day

second generation :
actual intakes during premating : male : 1.61, 3.33, 7.03 mg/kg body weight/day
actual intakes during premating : female : 2, 3.4, 8.46 mg/kg body weight/day
actual intakes during gestation : female : 1.54, 3.18, 6.64 mg/kg body weight/day
actual intakes during lactation : female : 3.38, 7.48, 16.45 mg/kg body weight/day

From Day 0 onward, the rats received the test diets with 0 (control), 2.5, 5, and 10% erythritol. There was no adaptation period with stepwise increasing levels.

Details on mating procedure:

Three days before the start of the study, all rats were weighed and allotted to four groups of 24 rats/sex/group by computer randomization on basis of mean body weight.
At the end of the 10-week premating period, the animals were paired, according to numerical sequence, by caging one female with one male of the same dose group (F0-generation). The next morning the females were checked for evidence of mating. Females not showing a vaginal plug or sperm cells were returned to the male. This procedure was continued for 3 weeks. The day of detection of a vaginal plug of sperm cells was considered to be Day 0 of gestation. Upon evidence of copulation, males were returned to their original group cage. Positive females were housed individually in nesting cages for the birth and rearing of their pups (F1-generation).

Days 4 post-partum : litter adjusted to eight pups by eliminating extra pups by random selection to yields, as nearly as possible, four males and four females per litter.
Day 21 post-partum : pups weaned. After that, 24 male and 24 female pups of each dose group were selected at random and from as many litters as possible to rear the next generation. The remaining pups were discarded

Procedures followed to mate the F1-generation and to rear the F2-litters were the same as those described for the F0-generation.

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

The mean intake of erythritol was calculated per kilogram of body weight per day on the basis of the nominal dietary erythritol concentrations. Water consumption was not measured.

Duration of treatment / exposure:

10 weeks

Frequency of treatment:

At each meal, food and water available ad libitum

Details on study schedule:

Main steps of the study :
- Acclimatization : 2 weeks
- 3 days before the start of the study, all rats were weighed and allotted to four groups of 24 rats/sex/group kilo by computer randomization on basis of mean body weight.
- Premating : 10 weeks
- Mating after the premating. The next morning, if the females did not show a vaginal plug or sperm cells were returned to the male. Procedures continued for 3 weeks.
- Birth of the pups
- Days 4 post-partum : litter adjusted to eight pups by eliminating extra pups by random selection to yields, as nearly as possible, four males and four females per litter.
- Day 21 post-partum : pups weaned. After that, 24 male and 24 female pups of each dose group were selected at random and from as many litters as possible to rear the next generation. The remaining pups were discarded
- Sacrificed and gross examination of the F0 parent : at week 20
- The study was terminated at the end of weaning of the F2 pups.

Dose / conc.:

2.5 other: % at the expense of dietary starch

Dose / conc.:

5 other: % at the expense of dietary starch

Dose / conc.:

10 other: % at the expense of dietary starch

Dose / conc.:

0 other: % at the expense of dietary starch

No. of animals per sex per dose:

24 rats/sex/generation/dose group

Control animals:

yes, concurrent no treatment

yes, plain diet

Positive control:

Not specified

Parental animals: Observations and examinations:

Body weights during premating period : all prospective parent males and females were recorded weekly
Body weights after mating period : males were weighed weekly until sacrifice / Mated females were weighed on Days 0, 7, 14, and 21 of gestation and during lactation on Days 1, 7, 14, and 21 postpartum. / Females which did not have a positive smear or vaginal plug and females which did not deliver a litter were not weighed after the end of the premating and gestation period, respectively.

Clinical signs and abnormal behavior : all animals were checked daily

Oestrous cyclicity (parental animals):

Not specified

Sperm parameters (parental animals):

Not specified

Litter observations:

Clinical signs and abnormal behavior : all animals were checked daily
Litter size / number of male and female / number of pups with abnormalities : determined on days 1, 4 (before culling), 7, 14 and 21 postpartum
Body weights of entire litter : determined on days 1 4 (before culling), 7, 14 postpartum
Body weight of individual pups : on day 21, at weaning

Postmortem examinations (parental animals):

All parental animals of the F1-generation were sacrificed and necropsied
At termination, all surviving parental animals of the F0- and F1-generation were killed by decapitation under ether anesthesia and then examined for gross pathological changes.

Postmortem examinations (offspring):

Necropsy was performed on stillborn pups and pups dying during lactation.

Statistics:

clinicat findings were evaluated using Fisher's exact probability test; body weight and food consumption data were analysed using one way analysis of variance and Dunnett's multiple comparison tests; mating parameters were evaluated using Fisher's exact probability test; duration of gestation and litter size were evaluated using Kruskal-Wallis nonparametric analysis of variance followed by the Mann-Whitney U-test; pup body weights were analysed by analysis of variance followed by Dunnett's multiple comparison test; implantations were analysed using Kruskal-Wallis nonparametric analysis of variance followed by the Mann-Whitney U-test; pathologicai changes were analysed using Fisher's exact probability test. Ail tests were two-sided.

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

Transient diarrhea occurred in all F0-males and -females of the 10% erythritol group at the start of the study, but ceased in most animals with continued administration of erythritol.

Dermal irritation (if dermal study):

not specified

Mortality:

mortality observed, non-treatment-related

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weights and body weight gains were below those of controls in the F0-males and -females of the 10% erythritol group.
Males : the reduction of body weights was significant from Week 1 to 19
Female : this effect reached statistical significance in Week 4 only

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

10% erythritol : Food consumption of F0-males and -females was significantly below that of controls during the first week of treatment in the 10% erythritol group. Thereafter, it was significantly increased, in males through the end of the study in Week 19 and in females during Weeks 6–10 of the premating period and the first week of lactation
5% erythtritol : significant enhancing effect on food intake was seen only rarely (F0 females in Weeks 6, 9, and the first week of gestation)

Food efficiency:

not specified

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

not examined

Description (incidence and severity):

tap water were provided ad libitum throughout the study but not measured

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

Microscopic examination did not reveal any histopathological changes that could be related to the treatment. The abnormalities observed were common findings in rats of this strain and age. Moreover, the lesions were about equally distributed among the test groups and controls, or they occurred only in one or a few animals.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Reproductive function: oestrous cycle:

effects observed, non-treatment-related

Description (incidence and severity):

minor abnormalities were equally distributed among the different groups or occurred in one or a few rats only. Microscopic examination did not reveal any histopathological changes that could be related to the treatment. The abnormalities observed were common findings in rats of this strain and age. Moreover, the lesions were about equally distributed among the test groups and controls or they occurred only in one or a few animals.

Reproductive function: sperm measures:

effects observed, non-treatment-related

Description (incidence and severity):

minor abnormalities were equally distributed among the different groups or occurred in one or a few rats only. Microscopic examination did not reveal any histopathological changes that could be related to the treatment. The abnormalities observed were common findings in rats of this strain and age. Moreover, the lesions were about equally distributed among the test groups and controls or they occurred only in one or a few animals.

Reproductive performance:

no effects observed

Description (incidence and severity):

Mating was successful in all pairs of each treatment group in each generation except in F0-rats of the 10% erythritol group in which 22 of 24 females were mated. Precoital time was slightly shorter in the F0- than in the F1-generation but it was similar among the different treatment groups.
There were no significant differences between groups with regard to the number of pregnant females (18–22 per group in the F0-generation).

The female fertility index varied from 75 to 83%
The fecundity index from 75 to 92%
Both indices were unaffected by the erythritol treatment and were within normal limits

The gestation time did not differ between the groups.
The gestation index varied between 91 and 100%.

Postimplantation loss was moderate and similar among the different treatment groups. A few stillborn pups were observed but the live birth index was close to 100% in all groups.

Key result

Effect level:

> 7 530 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

reproductive performance

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Effect level:

> 7 670 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

reproductive performance

Remarks on result:

not determinable due to absence of adverse toxic effects

Clinical signs:

no effects observed

Description (incidence and severity):

Diarrhea was not observed in any rat of the F1-generation.

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

F1-males and -females of the 10% erythritol group exhibited reduced body weights, the difference from controls being statistically significant for males during Weeks 0–8 and 17–18 and for females during Weeks 0–4. However, the rates of body weight gain of males and females of this dose group were not different from those of the controls.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

10% erythtitol : Food consumption was consistently increased in F1-males and -females from the first measurement (about 2 weeks after weaning) to the end of the study (difference from controls statistically significant in all weeks except for males in Weeks 1 and 15)
5% erythtritol : significant enhancing effect on food intake was seen only rarely (F1-males in Weeks 8–10; and F1-females in Weeks 5–7 and 9, the last week of gestation, and the first week of lactation)

At the high-dose level, the erythritol intake amounts to about 6.5 g/kg body wt/day. However, in lactating females, it is more than twice as high.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Description (incidence and severity):

Minor abnormalities observed equally distributed among the different groups or occurred in one or a few rats only.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

Microscopic examination did not reveal any histopathological changes that could be related to the treatment. The abnormalities observed were common findings in rats of this strain and age. Moreover, the lesions were about equally distributed among the test groups and controls, or they occurred only in one or a few animals.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Description (incidence and severity):

There were no significant differences between groups with regard to the number of pregnant females (17–21 per group in the F1-generation)

The female fertility index varied from 71 to 88%
The fecundity index from 71 to 88%
Both indices were unaffected by the erythritol treatment and were within normal limits

The gestation time did not differ between the groups
The gestation index varied between 91 and 100%.

Postimplantation loss was moderate and similar among the different treatment groups. A few stillborn pups were observed but the live birth index was close to 100% in all groups

Key result

Effect level:

> 7 530 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

clinical signs

mortality

gross pathology

histopathology: non-neoplastic

reproductive performance

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Effect level:

> 7 670 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

clinical signs

mortality

gross pathology

histopathology: non-neoplastic

reproductive performance

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Effect level:

> 3 765 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

food consumption and compound intake

Remarks on result:

not determinable due to adverse toxic effects at highest dose / concentration tested

Key result

Effect level:

> 3 835 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

body weight and weight gain

food consumption and compound intake

Remarks on result:

not determinable due to adverse toxic effects at highest dose / concentration tested

Remarks on result:

not determinable due to absence of adverse toxic effects

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

pup mortality in the 10% erythritol group seemed to differ from that of the control group, in the F0-generation because it was higher than the control group. But the results were no significant.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

The group mean body weight of the F1-pups (derived from F0-parents) was significantly decreased in the 10% erythritol group on Day 21 of lactation.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

10% erythritol group : Food consumption was consistently increased in F1-males and -females of the 10% erythritol group from the first measurement (about 2 weeks after weaning) to the end of the study (difference from controls statistically significant in all weeks except for males in Weeks 1 and 15)
5% erythritol group : a significant enhancing effect on food intake was seen only rarely (F1-males in Weeks 8–10; and F1-females in Weeks 5–7 and 9, the last week of gestation, and the first week of lactation)

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

effects observed, non-treatment-related

Description (incidence and severity):

The sex ratio on Day 1 and Day 21 varied between 48 and 57% in the F0-generation (F1 pups)

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Visual examination of the pups revealed a few abnormalities such as small size (i.e., body weight of less than 75% of controls), subcutaneous hemorrhages at different sites, alopecia, and missing or dead tail tips. These findings were randomly distributed among the various groups or they occurred in one or a few pups only with the exception of smaller F1-pups in the 10% erythritol group which is in line with the lower pup body weights of this group

Histopathological findings:

not examined

Other effects:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

The mean number of live pups per litter was slightly higher in the 10% erythritol group of both generations.

Key result

Generation:

F1

Effect level:

>= 7 530 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

viability

mortality

gross pathology

other: sex-ratio

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Generation:

F1

Effect level:

> 7 670 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

viability

mortality

gross pathology

other: sex-ratio

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Generation:

F1

Effect level:

> 3 765 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

food consumption and compound intake

Remarks on result:

not determinable due to adverse toxic effects at highest dose / concentration tested

Key result

Generation:

F1

Effect level:

> 3 835 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

body weight and weight gain

food consumption and compound intake

Remarks on result:

not determinable due to adverse toxic effects at highest dose / concentration tested

Clinical signs:

not specified

Dermal irritation (if dermal study):

not examined

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

pup mortality in the 10% erythritol group seemed to differ from that of the control group, in the F1-generation (pup F2) because it was lower than the control group. But the results were no significant.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

In the F2 pups no difference was observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not specified

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

no effects observed

Description (incidence and severity):

The sex ratio on Day 1 and Day 21 varied between 50 and 57% in the F1-generation (F2 pups)

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

Other effects:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Key result

Generation:

F2

Effect level:

> 7 530 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

value from test report (7.6 mg/kg bw/d in the publication)

Sex:

male

Basis for effect level:

viability

mortality

food consumption and compound intake

gross pathology

other: sex-ratio

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Generation:

F2

Effect level:

>= 7 670 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

value from test report (7.6 mg/kg bw/d in the publication)

Sex:

female

Basis for effect level:

viability

mortality

food consumption and compound intake

gross pathology

other: sex-ratio

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Reproductive effects observed:

no

Conclusions:

The effects seen in parental animais consisted of body weight changes which were due to the reduced calorie intake due to erythritol added to the diet at high levels. This was concluded based on the lower body weights despite higher intake of food in the erythritol groups. In addition, no toxicological effects were apparent in the treated animais.
Despite the reduced body weights in high-dose animais, no effects on fetal development or pup body weights were observed, nor were any reproductive parameters affected. Some statistical differences were noted in pup mortality rates; however, the differences were contradictory, being increased in one generation but decreased in the next, or appeared only at lower dose.
As a result, the authors concluded that erythritol did not affect reproductive performance or fetal development in rats when administered in the diet at levels of up to 10% for 2 consecutive generations. Mean intakes of erythritol at the highest dose levels were reported to be 7.53 and 7.67 g/kg body weight/day for males and females, respectively.

Executive summary:

In an assay similar to OECD 416 (two generation reproductive toxicity study), the authors concluded that erythritol did not affect reproductive performance or fetal development in rats when administered in the diet at levels of up to 10% for 2 consecutive generations. Mean intakes of erythritol at the highest dose levels were reported to be 7.53 and 7.67 g/kg body weight/day for males and females, respectively.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Species:

rat