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Key value for chemical safety assessment

Effects on fertility

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Endpoint:
three-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions.
Qualifier:
according to
Guideline:
other: No guideline specified, but conforms to the standard 3 generation 2 litters per generation MGS normally used at that time.
Deviations:
not specified
GLP compliance:
no
Remarks:
Study pre-dates GLP
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, USA.
- Weight at study initiation: (P) Males: 130 - 135 g; Females: 110 - 149 g
- Diet (e.g. ad libitum): Ad libitum
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
This was a three generation multigeneration study with two matings (two litters) per generation. The F1a, F2a and F3a litters were sacrificed at weaning, and the F1b and F2b litters raised and used for breeding, and the F3b killed at weaning.
Details on mating procedure:
- M/F ratio per cage: 1 M and 2 F per cage
- Length of cohabitation: 21 days on each occassion
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
No data
Duration of treatment / exposure:
Groups of 8 males and 16 females were used for all generations. 14 weeks before mating.
From beginning of the study until sacrifice of parents P0, and from weaning till sacrifice for the parents of the F1 and F2-generations. The high dose group P animals were sterile so only controls, low and mid dose groups were taken to the F2 and F3 generations.
Frequency of treatment:
No data
Details on study schedule:
Offspring were culled to 8 per litter at 24 h after delivery.
Remarks:
Doses / Concentrations:
0, 1030, 3080 or 10300 ppm borax (0, 117, 350 and 1,170 ppm boron) in the diet, equivalent to 0, 50 (5.9), 155 (17.5) and 518 (58.5) mg borax (mg B)/kg bw/day respectively
Basis:
nominal in diet
No. of animals per sex per dose:
8 males, 16 females per group
Control animals:
yes, plain diet
Positive control:
No data
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: weekly


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly


BODY WEIGHT: Yes
- Time schedule for examinations: Weekly


FOOD CONSUMPTION AND COMPOUND INTAKE: Yes, weekly
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data


WATER CONSUMPTION AND COMPOUND INTAKE : No
Oestrous cyclicity (parental animals):
No data
Sperm parameters (parental animals):
No data, except in the high dose group in which histology of the testes was performed.
Litter observations:
Number and sex of pups, still births, livebirths, presence of gross abnormalities, weight gain, physical or behavioural abnormalities.
Postmortem examinations (parental animals):
Organ weights: Uterus, ovaries, testes, brain, liver, kidneys, spleen, thyroid and adrenals
Histopathology: All parental animals were necropsied and a range of tissues preserved in formalin but not examined histologically except for testes, ovaries and uterus of the high dose group only.
Postmortem examinations (offspring):
Organ weights: Uterus, ovaries, testes, brain, liver, kidneys, spleen, thyroid and adrenals
Histopathology: 5 of each sex from all groups of the F3b litters were necropsied and a range of tissues fixed in formalin but not examined histologically.
Statistics:
No data
Reproductive indices:
No data
Offspring viability indices:
No data
Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
not specified
Histopathological findings: non-neoplastic:
not specified
Other effects:
no effects observed
Reproductive function: oestrous cycle:
not specified
Reproductive function: sperm measures:
effects observed, treatment-related
Reproductive performance:
effects observed, treatment-related
Parent males:
Rats exposed to the high dose of 518 mg/kg borax (corresponding to a level of 58.5 mg B/kg bw) had reduced bodyweights though food intake was not affected and they were sterile. Microscopic examination of the atrophied testes of all males in this group showed no viable sperm. There were no adverse effects on reproduction reported at exposures of 5.9 and 17.5 mg B/kg bw. The authors reported no adverse effects on fertility, lactation, litter size, progeny weight or appearance in rats exposed to either 5.9 or 17.5 mg B/kg bw. Also, no gross abnormalities were observed in the organs from these dose groups.

Parent females:
The high dose groups of the P0 generation had reduced bodyweight without any effect on food intake. Evidence of decreased ovulation in about half of the ovaries examined from the females exposed to 58.5 mg B/kg bw and only two of 16 females produced a litter (one of which was cannibalised within 24 h) when mated with control male animals. There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.

The high dose group (58.5 mg B/kg bw) males and females showed clinical signs of toxicity with rough fur, scaly tails, respiratory distress and inflamed eyelids.
Dose descriptor:
LOAEL
Effect level:
518 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on sterility in males and females.
Remarks on result:
other: Equivalent to 1170 boron in the diet
Dose descriptor:
NOAEL
Effect level:
155 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: based on sterility in males and females.
Remarks on result:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
LOAEL
Effect level:
58.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: based on sterility in males and females.
Remarks on result:
other: Equivalent to 1170 ppm boron in the diet.
Dose descriptor:
NOAEL
Effect level:
17.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: based on sterility in males and females.
Remarks on result:
other: Equivalent to 350 ppm boron in the diet.
Clinical signs:
not specified
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
not specified
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
not specified
Histopathological findings:
not specified
F1 and F2 males and females: There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
155 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
Remarks on result:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
17.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
Remarks on result:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
155 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
17.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: No adverse effects in mid and low dose groups in any generation.
Reproductive effects observed:
not specified

Parameter

 

Control

Low dose

Medium dose

High dose

 

Generation

m

f

m

f

m

f

m

f

 

 

Mortality

Incidence

P

0

0

0

0

0

0

1

0

 

 

F1

0

0

0

0

1

0

0

0

 

 

F2

0

0

0

0

0

0

0

0

 

 

Food consumption

% of control

Not affected

 

 

Body weight gain

% of control

 

-

-

-

-

-

-

¯

¯

 

 

Clinical Observations

Incidence

 

-

-

-

-

-

-

+

+

 

 

Organ weights

% of control

Only effect noted was increase in absolute and relative thyroid wt. in low and mid dose groups (not thought to biologically significant)

Pathology

Histopathologic examination

Incidence

Evidence of testis atrophy in high dose males of P0 generation.

Evidence in ovary of reduced ovulation in high dose females.

Reproductive Performance

 

P0 to F1a

F1b to F2b

F2b to F3b

 

cont

low

mid

high

cont

low

mid

cont

low

mid

 

Mating index: (No. pregnant/No. mated)

%

63

69

75

0

80

75

94

69

88

100

 

Fertility index: No. litters born/No. Pregnant

%

100

100

100

-

100

100

100

91

100

100

 

Birth index

Live birth index: No.pups alive/No. born

%

98

98

100

 

99

92

99

100

100

100

 

Litter size

Mean

12

12

13

 

12

15

12

12

12

12

 

Pup weight

at 24h (g)

Mean

7.0

6.8

7.1

 

6.4

6.2

7.0

6.0

7.0

8.0

 

Sex ratio

Male/female

6/6

6/6

6/7

 

6/6

8/7

5/7

6/6

7/5

6/6

 

Lactation index:

Pup wt. at weaning

 

55

55

51

 

56

58

53

48

52

52

 

Conclusions:
Rats exposed to the high dose of 518 mg/kg bw of borax (corresponding to a level of 58.5 mg B/kg bw) were sterile. Microscopic examination of the atrophied testes of all males in this group showed no viable sperm. The authors also reported evidence of decreased ovulation in the majority of ovaries examined from the females exposed to 58.5 mg B/kg bw and no litters were obtained from these high dose females when mated with control male animals. There were no adverse effects on reproduction reported at exposures of 50 and 155 mg/kg bw borax ( 5.9 and 17.5 mg B/kg bw). The authors reported no adverse effects on fertility, lactation, litter size, progeny weight or appearance in rats exposed to either 5.9 or 17.5 mg B/kg bw. Also, no gross abnormalities were observed in the organs examined from either parents or weanlings from these dose groups. Based on these study data, the authors concluded that exposure of rats at levels up to 17.5 mg B/kg bw in the diet in a 3 generation reproduction study was without adverse effect. Data presented with boric acid data to support the data obtained with boric acid.
Endpoint:
three-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Comparable to guideline study with acceptable restrictions. This study is conducted on an analogue substance. Read-across is justified on the following basis: In aqueous solutions at physiological and acidic pH, low concentrations of simple inorganic borates such as boric acid, disodium tetraborate decahydrate, disodium tetraborate pentahydrate, boric oxide and disodium octaborate tetrahydrate will predominantly exist as undissociated boric acid. At about pH 10 the metaborate anion (B(OH)4-) becomes the main species in solution (WHO, 1998). This leads to the conclusion that the main species in the plasma of mammals and in the environment is un-dissociated boric acid. Since other borates dissociate to form boric acid in aqueous solutions, they too can be considered to exist as un-dissociated boric acid under the same conditions. For comparative purposes, exposures to borates are often expressed in terms of boron (B) equivalents based on the fraction of boron in the source substance on a molecular weight basis. Some studies express dose in terms of B, whereas other studies express the dose in units of boric acid. Since the systemic effects and some of the local effects can be traced back to boric acid, results from one substance can be transferred to also evaluate the another substance on the basis of boron equivalents. Therefore data obtained from studies with these borates can be read across in the human health assessment for each individual substance. Conversion factors are given in the table below. Conversion factor for equivalent dose of B Boric acid H3BO3 0.175 Boric Oxide B2O3 0.311 Disodium tetraborate anhydrous Na2B4O7 0.215 Disodium tetraborate pentahydrate Na2B4O7•5H2O 0.148 Disodium tetraborate decahydrate Na2B4O7•10H2O 0.113 Disodium octaborate tetrahydrate Na2B8O13•4H2O 0.210 Sodium metaborate (anhydrous) NaBO2 0.1643 Sodium metaborate (dihydrate) NaBO2•2H2O 0.1062 Sodium metaborate (tetrahydrate) NaBO2•4H2O 0.0784 Sodium pentaborate (anhydrous) NaB5O8 0.2636 Sodium pentaborate (pentahydrate) NaB5O8∙5H2O 0.1832 References: WHO. Guidelines for drinking-water quality, Addendum to Volume 1, 1998.
Qualifier:
according to
Guideline:
other: No guideline specified, but conforms to the standard 3 generation 2 litters per generation multi-generation studies normally used at that time.
Deviations:
not applicable
GLP compliance:
no
Remarks:
Study pre-dates GLP
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Caesarean-derived from Charles River
Weight at study initiation: (P) Males: 121 - 150 g; Females: 110 - 147 g
- Diet: Ad libitum
- Housing: Prior to initiation of the first breeding phase, the animals were maintained in individual cages and fed their respective diets for 14 weeks until they reached maturity.
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
Rats were exposed from beginning of the study until sacrifice of parents P0 , and from weaning till sacrifice for the parents of the F1 and F2-generations.
The high dose group P animals were sterile so only controls, low and mid dose groups were taken to the F2 and F3 generations.

DIET PREPARATION
- Mixing appropriate amounts with (Type of food): The test material was incorporated into the basal diet on a weight/weight basis and thoroughly mixed in a twin-shell blender to provide the desired dietary levels.
Details on mating procedure:
- M/F ratio per cage: 1:2
- Length of cohabitation: 21 days on each occasion
- Any other deviations from standard protocol: This is a three generation multigeneration study with two matings (two litters) per generation. The F1a, F2a and F3a litters were sacrificed at weaning, and the F1b and F2b litters raised and used for breeding, and the F3b killed at weaning.

24 h after birth, the litters were reduced to a maximum of eight pups to be nursed. The F1A litters were discarded when they reached 21 days of age. The parents in the control and two lower test groups were remated to produce their second (F1B) litters. At the time of weaning 16 females and 8 males from the control and two test groups were selected at random and designated as the second parental generation (P2) for continuation of the reproduction study. All excess weanlings were discarded.
The experimental design for the high level test group (0.67 %) was altered due to failure of the P1 parents to produce litters. In order to determine whether the female reproductive system was affected, the P1 females in the high level group were mated with males of the same strain and approximately the same age, which had received only the control diet. The males remained in the breeding cage for 8 h each day. To prevent the males from feeding on teh test diet, no food was available to the animals during the daily mating period.
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
No data
Duration of treatment / exposure:
Groups of 8 males and 16 females were used for all generations and were exposed from beginning of the study until sacrifice of parents P0, and from weaning till sacrifice of the F1- and F2-generations.
The high dose group P animals were sterile so only controls, low and mid dose groups were taken to the F2 and F3 generations.
Frequency of treatment:
Daily
Details on study schedule:
This is a three generation multigeneration study with two matings (two litters) per generation. The F1a, F2a and F3a litters were sacrificed at weaning, and the F1b and F2b litters raised and used for breeding, and the F3b killed at weaning.
From beginning of the study until sacrifice of parents P0, and from weaning till sacrifice for the parents of the F1 and F2-generations.
The high dose group P animals were sterile so only controls, low and mid dose groups were taken to the F2 and F3 generations.
Remarks:
Doses / Concentrations:
0, 670, 2000 or 6700 ppm boric acid (0, 117, 350 and 1,170 ppm boron) in the diet, equivalent to 0, 34 (5.9), 100 (17.5) and 336 (58.5) mg boric acid (mg B)/kg bw/day.
Basis:

No. of animals per sex per dose:
8 males and 16 females per group
Control animals:
yes, plain diet
Details on study design:
- Rationale for animal assignment: By stratified randomisation
Positive control:
No data
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Weekly

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly


BODY WEIGHT: Yes
- Time schedule for examinations: Weekly


FOOD CONSUMPTION AND COMPOUND INTAKE: Yes, weekly
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

Oestrous cyclicity (parental animals):
No data
Sperm parameters (parental animals):
Sperm parameters were not done in the high dose group in which histology of the testes were performed.
Litter observations:
Number and sex of pups, stillbirths, live births, presence of gross abnormalities, weight gain, physical or behavioural abnormalities; culled to 8 per litter at 42 h after delivery.
Records were maintained on the number of conceptions, number and size of litters, deaths and weights of the pups at 24 h and at weaning. The pups were observed for gross signs of abnormalities.
Postmortem examinations (parental animals):
After completion of the second cycle (F1B) of the first breeding phase, all P1 animals in the control and two lower test groups were sacrificed (34th week of study). The males in the high level group were sacrificed after completion of the 27th week and the females after completion of the 46th week of the study. Gross necropsies were performed and representative tissues from each rat were preserved in 10 % formalin. Weights were obtained for brain, thyroid, liver, spleen, kidneys, adrenals and testes in all groups; and ovaries and uterus in the high level group. Organ/body weight ratios were obtained. Individual blood samples and pooled samples of brain, liver and kidney (all groups) and testis, ovary and uterus (high level only) were frozen for possible future analysis. The ovaries and uteri preserved from the high level females were examined microscopically.
After completion of the second breeding phase, all P2 animals were sacrificed and after completion of the third breeding phase, all P3 animals were savrificed. Necropsies were performed and the animals were observed for gross signs of pathology. The following tissues from eight males and eight females in the P2 and P3 control and test groups were preserved in 10 % formalin: Brain, thyroid, lung, heart, liver, kidney, adrenal, stomach, pancreas, small intestine, large intestine and gonad. Necropsies were also performed on 5 male and 5 female F3B weanlings from the control and two lower level test groups and representative tissues preserved in 10 % formalin.

Postmortem examinations (offspring):
No data
Statistics:
Terminal body weights, organ weights and organ/body weight ratios for the P1 animals were examined by the analysis of variance, of F-test, at the 5 % probability level. Before completing each F test, the variances were tested for heterogeneity by the method of Bartlett. If the variances were homogeous, the F-test could be applied in the normal fashion, and if a significant F value was obtained those groups significantly different from control could be determined by the method of Scheffe.
In those instances of heterogeneous variances, the samples were examined for extreme values by Sachs' test for rejection of measurements. If no legitimate unbiased adjustment to the variance could be made by rejection of "outliers", comparison test to control was effected by the Fisher-Behrens modified t-test. Breeding indices were analysed by the chi-square test of significance.
Reproductive indices:
No data
Offspring viability indices:
No data
Clinical signs:
not specified
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
not specified
Histopathological findings: non-neoplastic:
not specified
Other effects:
not specified
Reproductive function: oestrous cycle:
not specified
Reproductive function: sperm measures:
effects observed, treatment-related
Reproductive performance:
effects observed, treatment-related
Parent males:
Rats of the P0 generation exposed to the high dose of 336 mg/kg bw boric acid (corresponding to a level of 58.5 mg B/kg bw) had reduced bodyweights though food intake was not affected and they were sterile. Microscopic examination of the atrophied testes of all males in this group showed no viable sperm. There were no adverse effects on reproduction reported at exposures of 5.9 and 17.5 mg B/kg bw. The authors reported no adverse effects on fertility, lactation, litter size, progeny weight or appearance in rats exposed to either 5.9 or 17.5 mg B/kg bw. Also, no gross abnormalities were observed in the organs from these dose groups.

Parent females:
The high dose groups of the P0 generation had reduced bodyweight without any effect on food intake. Evidence of decreased ovulation in about half of the ovaries examined from the females exposed to 58.5 mg B/kg bw and only one of 16 females produced a litter when mated with control male animals. There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
Dose descriptor:
LOAEL
Effect level:
336 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Equivalent to 1170 ppm in the diet. Based on sterility.
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
LOAEL
Effect level:
58.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: Based on sterility. Testicular atrophy, reduced fertility (no offspring from high dose females mated with untreated males).
Dose descriptor:
NOAEL
Effect level:
17.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: Based on sterility. Testicular atrophy, reduced fertility (no offspring from high dose females mated with untreated males).
Clinical signs:
effects observed, treatment-related
Mortality / viability:
not specified
Body weight and weight changes:
not specified
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
not specified
Histopathological findings:
not examined
F1 males:
There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
F1 females:
There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.

F2 males:
There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
F2 females:
There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.

The high dose group (58.5 mgB/kg bw) males and females showed clinical signs of toxicity with rough fur, scaly tails, respiratory distress and inflamed eyelids.
The high dose group P animals were sterile so only controls, low and mid dose groups were taken to the F2 and F3 generations.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
100 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
Remarks on result:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
17.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: There were no adverse effects on reproduction and no gross abnormalities were observed in the organs at exposures of 5.9 and 17.5 mg B/kg bw.
Remarks on result:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
100 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Equivalent to 350 ppm boron in the diet.
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
17.5 mg/kg bw/day
Based on:
element
Sex:
male/female
Basis for effect level:
other: No adverse effects in mid and low dose groups in any generation.
Reproductive effects observed:
not specified

Table for reproductive toxicity:

Parameter

 

control

low dose

medium dose

High dose

 

Generation

m

f

m

f

m

f

m

f

 

 

Mortality

incidence

P

0

1

0

0

0

0

0

0

 

 

 

 

F1

0

1

0

0

0

0

0

0

 

 

 

 

F2

0

0

0

0

0

0

0

0

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Food consumption

% of control

not affected

 

 

 

 

 

 

 

 

 

 

Body weight gain

% of control

 

-

-

-

-

-

-

¯

¯

 

 

Clinical Observations

specify effects

Incidence

 

-

-

-

-

-

-

+

  +

 

 

Organ weights

% of control

only effect noted was increase in absolute wt. of thyroid in low dose group and relative thyroid wt. in low and mid dose groups (not thought to biologically significant)

Pathology

 

 

 

 

 

 

 

 

 

 

 

 

Histopathologic examination

specify effects

Incidence

Evidence of testis atrophy in high dose males of P0 generation.

Evidence in ovary of reduced ovulation in high dose females.

Reproductive Performance

 

P0 to F1a

F1b to F2b

F2b to F3b

 

cont

low

mid

high

cont

low

mid

cont

low

mid

 

Mating index: (No. pregnant/No. mated)

%

62

88

81

0

80

94

94

69

94

94

 

Fertility index: No. litters born/No. Pregnant

%

100

100

100

-

100

100

100

91

100

100

 

Number of implantation sites

Mean

 

 

 

 

 

 

 

 

 

 

 

Duration of pregnancy

Mean

 

 

 

 

 

 

 

 

 

 

 

Birth index

 

 

 

 

 

 

 

 

 

 

 

 

Live birth index: No.pups alive/No. born

%

98

96

97

 

99

99

98

100

99

99

 

Gestation index

 

 

 

 

 

 

 

 

 

 

 

 

Litter size

Mean

12

11

11

 

12

13

12

12

13

11

 

Litter weight

Mean

 

 

 

 

 

 

 

 

 

 

 

Pup weight at 24h (g)

Mean

7.0

7.2

6.7

 

6.4

6.5

6.7

6.0

7.0

7.0

 

Sex ratio

Male/female

6/6

6/5

5/6

 

6/6

7/6

6/6

6/6

7/6

6/5

 

Survival index

 

 

 

 

 

 

 

 

 

 

 

 

Viability index

 

 

 

 

 

 

 

 

 

 

 

 

Lactation index: Pup wt. at weaning

 

55

50

52

 

56

53

51

48

51

55

 

Conclusions:
Rats exposed to the high dose of 336 mg/kg bw boric acid (corresponding to a level of 58.5 mg B/kg bw) were sterile. Microscopic examination of the atrophied testes of all males in this group showed no viable sperm. The authors also reported evidence of decreased ovulation in about half of the ovaries examined from the females exposed to 58.5 mg B/kg bw and only 1/16 matings produced a litter from these high dose females when mated with control male animals. There were no adverse effects on reproduction reported at exposures of 34 and 100 mg/kg bw boric acid (5.9 and 17.5 mg B/kg bw). The authors reported no adverse effects on fertility, lactation, litter size, progeny weight or appearance in rats exposed to either 5.9 or 17.5 mg B/kg bw. Also, no gross abnormalities were observed in the organs examined from either parents or weanlings from these dose groups. Based on these study data, the authors concluded that exposure of rats at levels up to 17.5 mg B/kg bw in the diet in a 3 generation reproduction study was without adverse effect.
Read-across is justified on the basis detailed in the rationale for reliability above. This study is therefore considered to be of sufficient adequacy and reliability to be used as a supporting study and no further testing is justified.
Effect on fertility: via oral route
Dose descriptor:
NOAEL
127.8 mg/kg bw/day
Additional information

Assessment entity approach

"Brazing fluxes" are mixtures of boron-containing constituents (potassium(fluoro)borates), which undergo chemical exchanges (anion exchange) and condensation reactions (e.g. formation of oligoborates, polyborates) upon mixing and further manufacturing. This results in a complex mixture of potassium borates, which cannot be fully chemically characterised for substance identity. Thus, according to the definition under REACH, such brazing fluxes must be described as a UVCB substance.

 

Data specifically on the UVCB substance to be registered are not available. An assessment entity approach is followed based on the transformation products of this UVCB uppon dissolution in aqueous media. The substance is highly soluble and forms complex boron, potassium and fluoride constituents. The quantitatively predominanttransformation product of this UVCB is represented by boric acid, which is assumed to be the determinant of human health effects because of its classification and its toxicity. For this reason, the assessment is based on information for “borates” (including potassium borate, boric acid and other borate substances).

 

Based on the information provided below, it may safely be assumed that under physiological conditions the chemical speciation of most of the unknown potassium boron compounds corresponds to boric acid. Thus, from a chemical point of view, there is no reason to assume that brazing fluxes would behave differently than boric acid and/or borates under physiological conditions.

 

The basis of this assessment entity approach is further justified by the following reasoning:

In aqueous solutions at physiological and acidic pH, low concentrations of simple inorganic borates such as boric acid B(OH)3, potassium pentaborate (K2B10O16*8H2O), potassium tetraborate (K2B4O7*4H2O), disodium tetraborate decahydrate (Na2B4O7.10H2O; borax), disodium tetraborate pentahydrate (Na2B4O7*5H2O; borax pentahydrate), boric oxide (B2O3) and disodium octaborate tetrahydrate (Na2B8O13*4H2O) will predominantly exist as undissociated boric acid. Above pH 9 the metaborate anion (B(OH)4-) becomes the main species in solution (WHO, 1998). This leads to the conclusion that the main species in the plasma of mammals and in the environment is undissociated boric acid. Since other borates dissociate to form boric acid in aqueous solutions, they too can be considered to exist as undissociated boric acid under the same conditions.

For comparative purposes, exposures to borates are often expressed in terms of boron (B) equivalents based on the fraction of boron in the source substance on a molecular weight basis. Some studies express dose in terms of B, whereas other studies express the dose in units of boric acid. Since the systemic effects and some of the local effects can be traced back to boric acid, results from one substance can be transferred to also evaluate the another substance on the basis of boron equivalents. Therefore data obtained from studies with these borates can be read across in the human health assessment for each individual substance. Conversion factors are given in the table below.

 

Substance

Formula

Conversion factor for equivalent dose of B (multiply by)

Boric acid

H3BO3

0.1748

Boric Oxide

B2O3

0.311

Disodium tetraborate anhydrous

Na2B4O7

0.2149

Disodium tetraborate pentahydrate

Na2B4O7•5H2O

0.1484

Disodium tetraborate decahydrate

Na2B4O7•10H2O

0.1134

Disodium octaborate tetrahydrate

Na2B8O13·4H2

0.2096

Sodium metaborate (anhydrous)

NaBO2

0.1643

Sodium metaborate (dihydrate)

NaBO2·2H2O

0.1062

Sodium metaborate (tetrahydrate)

NaBO2·4H2O

0.0784

Sodium pentaborate (anhydrous)

NaB5O8

0.2636

Sodium pentaborate (pentahydrate)

NaB5O8∙5H2O

0.1832

 Dipotassium tetraborate (anhydrous)

 

 K2B4O7

 

 0.185

 

 Dipotassium tetraborate (tetrahydrate)

 

 K2B4O7.4H2O

 

 0.1415

 

 Potassium pentaborate (anhydrous)

 

 B5KO8

 

 0.244

 

 Potassium pentaborate (tetrahydrate)

 

 B5KO8.4H2O

 

 0.1843

 

 

Reference: WHO. Guidelines for drinking-water quality, Addendum to Volume 1, 1998

 

  

Discussion (effects on fertility, data for borates):

Effects on male fertility of borates have been investigated in detail. A dose related effect on the testis was observed in rats, mice and deer mice, with confirmation from limited studies in dogs. Effects in rats start with reversible inhibition of spermiation after 14 days (at 39 mg B/kg bw/day) and 28 days (at 26 mg B/kg bw/day). At doses equal to and above 26 mg B/kg bw/day testicular atrophy, degeneration of seminiferous tubules and reduced sperm counts were observed. Male fertility was further investigated in two serial mating studies of treated male rats with untreated female rats. Infertility of treated males correlated well with germinal aplasia. Similar effects on male fertility were described in deer mice (Peromyscus maniculatus) after treatment with boric acid. Fertility studies in rats (two three-generation study with for boric acid and disodium tetraborate decahydrate) and mice (a continuous breeding study with boric acid) further support effects on testes as the underlying cause for reduced male fertility.

Diminished sperm production may be due to testicular effects on germ cell, Sertoli cell, or Leydig cell function or act via an alteration of the pituitary-hypothalamic axis. There is an indication that LH and FSH are elevated under boric acid treatment (Lee et al., 1978) and that serum testosterone may be decreased in CD-1 mice and F344 rats (Grizzle et al., 1989; reviewed in Fail et al., 1991; Treinen & Chapin, 1991). The decrease in prostate weight at 111.3mg B/kg bw/day observed by Fail et al. (1991) might be caused by reduced testosterone levels.

A NOAEL of 17.5 mg B/kg bw/day for effects on female fertility was derived in the Transitional Annex XV dossier (TD 2008) based on Weir (1966c-d) and Fail et al. 1991. However, the TD failed to adequately distinguish between effects on female fertility and effects on development. Fertility is generally defined in males as the ability to produce sperm which are capable of producing fertilisation of an ovum leading to conception.  In females, it is defined as the ability to produce and release ova which can be fertilised leading to conception.  To test fertility in animals males and females are pretreated to cover the period of development of the sperm and eggs, then mate and treat until the time of implantation, around Day 6 following mating, and then stop treatment in the females.   To test for effects on development pregnant females are treated from Day 6 till the end of pregnancy. Neither the Weir and Fisher multigeneration study nor the Fail RACB studies were performed with this division of treatments. They both treated animals continuously before and during pregnancy and also after delivery.

In a three generation study in rats groups of 8 males and 16 females were treated with boric acid or disodium tetraborate decahydrate equivalent to 0, 5.9, 17.5 and 58.8 mg B/kg bw/day (Weir 1966c, d). An attempt was made to study the fertility of the P1 females at the top dose level by mating them with untreated males but only one litter of 16 pairs was produced. This highest dose level was clearly clinically toxic to the females after 2-3 weeks of dosing, with rough fur, scaly tails, inflamed eyelids and staining of the fur on the face and abdomen. The mating procedure to test the fertility of the females was not a satisfactory one. To avoid treatment of the males used for pairing, food was withdrawn from the cages of the females for 8 hours per day during the pairing process, and this is known to be very stressful to laboratory rats. There was no evidence on whether mating actually occurred for any of the rats, and no vaginal examinations for the presence of sperm were carried out. The females of the top dose P1 generation were sacrificed after 45 weeks of treatment and histopathological examination of the ovaries and uterus carried out. In the ovaries the presence of corpora lutea was regarded as a major indication of cyclic function, and these were found in 7 of 15 females, with reduced or absent function in the remaining 8 animals. The changes in the ovaries were not clearly different from those of controls.  No treatment related changes were found in the uterus. No changes were found that could account for the reduced litter production, and no conclusions could be drawn about fertility in the top dose females.  Comparable results were found in the Weir and Fisher multigeneration study on borax, with clear testicular atrophy at the top dose levels in males, and no clear explanation of the reduced number of litters in the top dose females, using the same unsatisfactory mating technique.  The authors of the study concluded that testis atrophy was clearly produced in males at the top dose level, but that the evidence of the decreased ovulation in females did not account for the reduced number of litters in the cross mating study in females.  Thus the Weir and Fisher studies produced clear evidence of adverse effects on male fertility, but did not produce clear evidence for an adverse effect on female fertility.

In a continuous breeding study of boric acid in Swiss mice (NTP, 1990; Fail et al., 1991), the three administered doses were 1000 ppm (26,6 mg B/kg bw/day), 4500 ppm (111,3 mg B/kg bw/day) and 9000 ppm (220,9 mg B/kg bw/day). A dose-related effect on the testis (testicular atrophy and effects on sperm motility, morphology and concentration) was noted; fertility was partially reduced at 111 mg B/kg bw/day, and absent at 221 mg B/kg bw/day.

For cross over mating only the mid dose group (111.3 mg B/kg bw/day) could be mated with control animals, since the high dose produced no litter. Indices of fertility for mid dose males with control females, control males with mid dose females and control males with control females were 5 %, 65 % and 74 %, respectively. The according indices of mating (incidence of copulatory plugs) were 30 %, 70 % and 79 %. This indicates that the primary effect was seen in males, however, slight effects were also noted in females. Live pup weight (adjusted for litter size) was significantly reduced compared to control litters, the average dam weight was significantly lower on postnatal day 0 compared to control dams and the average gestational period of the mid dose females was 1 day longer than in control females. The latter finding has also been observed in the developmental toxicity study by Price et al. (1996, see section 5.9.2).

In task 4 of this continuous breeding study control animals and low-dose F1 animals were mated because in the 9000 ppm groups no litters and in the 4500 ppm group only 3 litters were produced. While mating, fertility and reproductive competence were un-altered compared to control, the adjusted pup-weight (F2) was slightly but significantly decreased. F1 females had significantly increased kidney/adrenal and uterus weights and the oestrus cycle was significantly shorter compared to control females. A crossover mating study of controls and 4500 ppm groups confirmed the males as the affected sex. Necropsy at 27 weeks confirmed reduced testes weight, seminiferous tubule degeneration, decreased sperm count and motility and increase in abnormal sperm. In females at 27 weeks, 4500 ppm boric acid was toxic with decreased liver, kidney and adrenal weights, but no effect on oestrous cycles, mating, number of litters and number of pups. In F1 males a reduction in sperm concentration was observed, but no other sperm parameters were influenced.

While in this study the NOAEL for females of the F0-generation is 1000 ppm this is a LOAEL for males of the F0-generation (motility of epididymal sperms was significantly reduced: 78% ± 3 in controls vs. 69% ± 5 at 1000 ppm). For the F1-generation 1000ppm can be identified as a LOAEL, based on the 25% reduction of sperm concentration in males at this dose. Further, though normal in number, the F2-pups had reduced adjusted bodyweights at 1000 ppm, which is therefore also a LOAEL for F2-generation.

The authors concluded that the male is the most sensitive sex and that the testis is the primary target organ for boron. The NOAEL for testicular pathology in the present mouse study is probably 1000 ppm (26mg B/kg bodyweight). While males are more sensitive to boron induced toxicity, data also suggest an effect of boron on the female reproductive system. A reduced number of pups per litter and number of pups born alive at high dose levels are in agreement with earlier reports and could result from an effect of boron to alter implantation or to disrupt cell division in the embryo. This is supported by results of developmental toxicity studies in rats and mice in which higher dose levels can reduce the number of implants. Although F1 females had significantly increased kidney/adrenal and uterus weights and the oestrus cycle was significantly shorter compared to control female, similar effects were not observed in the 4500 ppm dose group, therefore the NOAEL in females was the dose level in diet of 4500 ppm, 846 mg/kg bw of boric acid or equivalent to 148 mg B/kg bodyweight.

In conclusion, the effects described in the Fail study on fertility show that 4500 ppm (111.3mgB/kg bw) is a NOAEL for the females, and that other small effects in females are the result of developmental toxicity for which a NOAEL of <1000ppm (26.6mg B/kg bw) may be valid.

No further studies on the effects of boron on female fertility were reported by the National Toxicology Program team who published several other studies on the mechanism of action of boron on male fertility and on spermatogenesis. No effects on steroidogenic function were found in Leydig cells, and no clear mechanism of action to cause testis atrophy was identified by Ku and Chapin (1994).

Although boron has been shown to adversely affect male reproduction in laboratory animals, male reproductive effects attributable to boron have not been demonstrated in studies of highly exposed workers.For further information on epidemiologic studies with workers exposed to high concentrations of boron, please refer to chapter 7.10.2 of this dossier and the respective endpoint summary.

In lack of data specifically for braze fluxes, and because borates are considered the transformation product driving the human health effects for brazing fluxes (see 'discussion`), it is considered justified to use the DNELS based on “boron” content from borate substances after stoichiometric recalculation for B content of the brazing fluxes.


Short description of key information:
A multigeneration study in the rat (Weir, 1966) gave a NOAEL for fertility in males of 17.5 mg B/kg/day.
No multigeneration studies with dipotassium tetraborate were available however since all the borates will exist as undissociated boric acid under physiological and environmental conditions, the toxicology of all these simple borates is similar on an equivalent boric acid basis or boron basis. Therefore the data for boric acid and disodium tetraborate decahydrate can be read across to the other borates for toxicological effects.
The following oral data were obtained (NOAEL):
Dipotassium tetraborate (anhydrous): 94.6 mg/kg bw/day
Dipotassium tetraborate (tetrahydrate): 123.7 mg/kg bw/day

Transferred to the brazing fluxes with ~13.7 wt-% boron: 127.8 mg/kg bw/day

Effects on developmental toxicity

Description of key information
No studies with brazing fluxes or dipotassium tetraborate were available however since all the borates will exist as undissociated boric acid under physiological and environmental conditions, the toxicology of all these simple borates is similar on an equivalent boric acid basis or boron basis. Therefore the data for boric acid and disodium tetraborate decahydrate can be read across to the other borates for toxicological effects.
A benchmark dose of 10.3 mg B/kg bw/day for developmental toxicity developed by Allen et al. (1996) was based on the studies of Heindel et al. (1992), Price, Marr & Myers (1994) and Price et al. (1996).
Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
19/11/1993 to 26/01/1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
GLP guideline study. This study is conducted on an analogue substance. Read-across is justified on the following basis: In aqueous solutions at physiological and acidic pH, low concentrations of simple inorganic borates such as boric acid, disodium tetraborate decahydrate, disodium tetraborate pentahydrate, boric oxide and disodium octaborate tetrahydrate will predominantly exist as undissociated boric acid. At about pH 10 the metaborate anion (B(OH)4-) becomes the main species in solution (WHO, 1998). This leads to the conclusion that the main species in the plasma of mammals and in the environment is un-dissociated boric acid. Since other borates dissociate to form boric acid in aqueous solutions, they too can be considered to exist as un-dissociated boric acid under the same conditions. For comparative purposes, exposures to borates are often expressed in terms of boron (B) equivalents based on the fraction of boron in the source substance on a molecular weight basis. Some studies express dose in terms of B, whereas other studies express the dose in units of boric acid. Since the systemic effects and some of the local effects can be traced back to boric acid, results from one substance can be transferred to also evaluate the another substance on the basis of boron equivalents. Therefore data obtained from studies with these borates can be read across in the human health assessment for each individual substance. Conversion factors are given in the table below. Conversion factor for equivalent dose of B Boric acid H3BO3 0.175 Boric Oxide B2O3 0.311 Disodium tetraborate anhydrous Na2B4O7 0.215 Disodium tetraborate pentahydrate Na2B4O7•5H2O 0.148 Disodium tetraborate decahydrate Na2B4O7•10H2O 0.113 Disodium octaborate tetrahydrate Na2B8O13•4H2O 0.210 Sodium metaborate (anhydrous) NaBO2 0.1643 Sodium metaborate (dihydrate) NaBO2•2H2O 0.1062 Sodium metaborate (tetrahydrate) NaBO2•4H2O 0.0784 Sodium pentaborate (anhydrous) NaB5O8 0.2636 Sodium pentaborate (pentahydrate) NaB5O8∙5H2O 0.1832 References: WHO. Guidelines for drinking-water quality, Addendum to Volume 1, 1998.
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
not specified
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl: CD VAF/Plus (Sprague Dawley)
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Inc., Raleigh, USA.
- Age at study initiation: 9 weeks of age
- Weight at study initiation: 219 - 296 g
- Housing: Individually in solid-bottom polycarbonate cages with stainless steel wire lids

IN-LIFE DATES: From: 19/11/1993 To: 26/01/1994
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food): Purina Certified Rodent Chow. Within each of the two study replicates, each concentration of boric acid in feed was formulated independently in a quantity sufficient for use across the period of administration for both study phases within that replicate, except that two batch formulations were prepared at 0.025 % in Replicate 1.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis of dosed feed by ICP indicated that the added concentrations of boric acid in feed were within 90.9 - 109.1 % of their nominal values. Boron levels in control rodent chow were less than 0.00281 % expressed as boric acid (i.e. less than 28.1 μg/g boric acid).
Mixes of the ground rodent chow were homogenous with the mean boric acid for 250 μg/g found at 96.3 % of nominal and a relative standard deviation of 2.7 %. For the 2000 μg/g dose level, the mean boric acid level found was 93.8 % of nominal with a relative standard deviation of 1.2 %.
The chow dosed with 250 μg/g boric acid was stable at room temperature over a period of 7 days with a recovery of 96.8, 100.4 and 100.7 % if nominal boric acid on days 0, 3 and 7 respectively. Similarly, after 7 days at room temperature, 20000 μg/g recoveries of 98.3, 96.3 and 93.7 % of nominal were found on days 0, 3, and 7 respectively. The same feed formulations were stable for 65 days when refrigerated.
Details on mating procedure:
Time-mated. Males and females were cohabited (1:1) overnight and mating was assessed by examination for sperm in the vaginal lavage of each cohabited female or the presence of a copulatory plug. The study was performed in two replicates with two study phases embedded in each replicate. There were 6 consecutive breeding dates in each replicate and the replicates were separated by 15 days between the last breeding date for replicate 1 and the first breeding date of replicate 2.
Duration of treatment / exposure:
Days 0 - 20 post mating (phase I)
Days 0 - 20 post mating then on normal diet until termination on day 21 postpartum (phase II)
Groups of 28-32 females were used for both phase I and phase II. In phase I the dams were killed on Day 20 for detailed fetal examination. In phase II the dams were allowed to deliver and the pups reared to weaning and then killed for full visceral and skeletal examination as for phase I.
Frequency of treatment:
Daily
Duration of test:
Days 0 - 20 post mating
No. of animals per sex per dose:
Phase I (developmental toxicity phase): 28 - 32 per group
Phase II (postnatal phase): 28 - 32 per group
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale:
The dose levels were selected to meet the principle objectives of the study. Specifically the proposed study was designed to determine whether the reduction of foetal body weight at 0.001 % boric acid was repeatable, whether the NOAEL for foetal weight reduction was could be determined and whether the induction of foetal seketal anomalies at 0.2 % boric acid was repeatable, whether the NOAEL for skeletal effects could be determined and whether the incidence of skeletal anomalies changed during postnatal life in the control and boric acid-exposed groups.
The period of exposure for all dietary concentrations of boric acid was GD 0 to 2, the same as for a previous study and would therefore allow determination of whether the effects were repeatable and establish and NOAEL under the same conditions.

- Rationale for animal assignment:
On GD 0, timed-mated females were assigned to dose groups and study phase by stratified randomisation so that the body weights did not differ among groups within either study phase.
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data


WATER CONSUMPTION AND COMPOUND INTAKE : Yes
- Time schedule for examinations: Over 3-day periods


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: Uterus
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No data
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
other - foetuses alive and dead. Empty uteri were stained with ammonium sulphide to look for implantation sites.
Fetal examinations:
- External examinations: No data
- Soft tissue examinations: No data
- Skeletal examinations: No data
- Head examinations: No data
Statistics:
Statistical procedures for selected experimental endpoints were based on SAS software. Except for nominal scale measures, data were reported as mean ± SEM. The litter was the experimental unit for all measures of developmental toxicity. An alpha level of 0.05 was applied to all group-wise, pair-wise or trend tests, except for Bartlett's Test or as otherwise noted below. Statistical results were reported by providing p values for ANOVA as p < 0.05 or p < 0.01 for pairwise tests; and as p < 0.05, < 0.01 or < 0.001 for trend tests in order to provide information equivalent to that obtained from the computer-generated output for these analyses.
Analyses for effects of boric acid were conducted within each study phase as follows. Except for nominal scale measurements, data was analysed by Bartlett's test for homogeneity of variance with an alpha level of 0.001. ANOVA was used to determine the significance of dose effects, replicate effects and dose x replicate interactions. When a significant (p < 0.05) main effect of dose occurred, Dunnett's test (one-tailed) and William's test were used to compare each boric acid-treated group to the vehicle control group for that measure, except that a two-tailed Dunnett's test was used for maternal organ and body weight parameters, maternal food and water consumption, foetal or pup body weight and percent males per litter. In three cases when the main effect for dose approached significance ) p = 0.0505 for maternal water intake of GD 18 to 19 in Phase I, and p = 0.0571 for absolute or relative maternal food intake on GD 0 to 3 in Phase II, the Dunnett's and Williams' tests were also applied. An arcsine-square root transformation was performed on all litter-derived percentage data prior to analysis by ANOVA. A test for linear trend was used to determine the significance of the dose-response relationship for parametric analyses.
Indices:
No data
Historical control data:
No data
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
No clinical effects or effects on bodyweight. There was an increase in relative kidney weight in the highest dose group in phase I of the study but not in Phase II. The authors concluded that there was little evidence of maternal toxicity at any of the doses tested (Phase I or 11).
Dose descriptor:
LOAEL
Effect level:
143 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
76 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
76 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEL
Effect level:
55 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Dose descriptor:
LOAEL
Effect level:
25 mg/kg bw/day
Based on:
element
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
13.3 mg/kg bw/day
Based on:
element
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
13.3 mg/kg bw/day
Based on:
element
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEL
Effect level:
9.6 mg/kg bw/day
Based on:
element
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Developmental effects were found in foetuses from animals exposed to 76 mg/kg bw boric acid (13.3 mg B/kg bw) and above (Phase I) which are mainly associated with foetotoxic activity. Specifically, a reduction in the mean foetal bodyweight per litter (6 % compared to controls) was observed in Phase I foetuses at 13.3 mg B/kg bw. At this dose, skeletal changes which included an increased incidence of short rib XIII (considered a malformation by authors of this study but a variation by most workers) and an increased incidence of wavy rib (considered a variation) were also observed. At the high dose for Phase I animals of 143 mg/kg bw boric acid (25 mg B/kg bw), the bodyweight reductions and skeletal changes were more pronounced. The reduction in incidence in extra rib on Lumbar I (a variation) which was noted in the previous rat study was not statistically significant here due to the low incidence in control animals (3.2 % in controls in this study compared to 14 % in the study from Heindel et al, 1992). There was no evidence of any increase in external or visceral malformations in any treatment group. The authors concluded that the NOAELs for the prenatal and postnatal study phases (Phase I and Phase II) were 9.6 and 12.9 mg B/kg low/d, respectively.
The animals from the Phase II group which were killed on postnatal day 21 showed no reduction in pup bodyweight in any group at any time point compared to controls, which indicates full recovery in the offspring already by postnatal Day 0 from treatment-related bodyweight effects. The rib variations observed in the foetuses (wavy rib) from Phase I were not observed in any dose group in Phase II. Only at the highest dose in Phase II (25.3 mg B/kg bw) was an increased incidence of short rib XIII observed.
Dose descriptor:
NOAEL
Effect level:
9.6 mg/kg bw/day
Based on:
element
Sex:
not specified
Basis for effect level:
fetal/pup body weight changes
skeletal malformations
Dose descriptor:
LOAEL
Effect level:
13.3 mg/kg bw/day
Based on:
element
Sex:
not specified
Basis for effect level:
fetal/pup body weight changes
skeletal malformations
Abnormalities:
not specified
Developmental effects observed:
not specified

Maternal effects Phase I and II:

Parameter

Control

0.025 dose

0.05 dose

0.075 dose

0.10 dose

0.20 dose

Number of dams examined

60

60

60

60

60

60

Clinical findings during application of test substance

none significant in any group

 

 

 

 

 

Mortality of dams (%)

0

0

0

0

0

0

Abortions

0

0

0

0

0

0

Body weight gain

no sig differences

 

 

 

 

 

Food consumption

no sig differences

 

 

 

 

 

Water consumption

if test substance is applied with drinking water

no sig differences

 

 

 

 

 

Pregnancies

pregnancy rate or %

56/60

56/60

55/60

58/60

55/60

55/60

Necropsy findings in dams dead before end of test

none

 

 

 

 

 

Litter response (Caesarian section data) Phase I:

 

Control

0.025

dose

0.05

dose

0.075

dose

0.10

dose

0.20

dose

Number of pregnancies

27

29

27

29

30

27

Corpora lutea           No. per dam

                                  ±SEM 

19.4

0.7

19.3

0.3

19.4

0.7

17.9

0.6

19.0

0.6

19.0

0.7

Implantations           No. per dam

                                   ± SEM

16.4

0.7

16.5

0.6

16.6

0.7

15.8

0.7

16.4

0.7

16.0

0.7

Resorptions               % per litter

                                   ±SEM

9.5

3.6

3.3

1.0

2.6

0.8

3.9

0.8

6.7

3.4

4.8

1.1

Total number of live foetuses

417

461

437

437

471

411

pre-implantation loss            %

                                               ±SEM

(%)           

13.8

4.1

13.5

2.7

13.2

3.5

12.8

3.7

11.5

4.0

14.1

4.1

post-implantation loss          %

                                               ±SEM

(%)

9.5

3.6

3.3

1.0

2.6

0.8

4.7

1.0

6.7

3.4

4.8

1.1

total number of litters

26

29

27

29

29

27

fetuses / litter

 

 

 

 

 

 

live fetuses / litter                 number

                                               ±SEM

ratio

16.0

0.4

15.9

0.6

16.2

0.7

15.1

0.7

16.2

0.6

15.2

0.7

Litters with one or more late foetal deaths

0

0

0

7

0

0

foetus weight (mean)              male

                                               female

                                               average

3.71

3.52

3.61

3.64

3.47

3.56

3.62

3.45

3.54

3.60

3.38

3.50

3.48

3.27

3.38

3.23

3.04

3.16

placenta weight

(mean) [g]

not recorded

 

 

 

 

 

crown-rump length (mean)[mm]

not recorded

 

 

 

 

 

Foetal sex ratio

[ratio m/f]

not reported

 

 

 

 

 

Examination of the foetuses:

Parameter

Control

0.025

dose

0.05

dose

0.075

dose

0.10

dose

0.20

dose

External malformations       mean

% offspring per litter           ±SEM

0.4

0.3

0.0

0.0

0.4

0.4

0.4

0.3

0.0

0.0

3.7

3.7

External variations               mean

% offspring per litter            ±SEM

0.2

0.2

0.0

0.0

0.0

0.0

0.5

0.3

0.0

0.0

0.0

0.0

Skeletal malformations         mean

% offspring per litter            ±SEM

2.0

0.7

0.9

0.6

1.6

0.6

2.5

0.7

3.5

1.2

4.3

1.5

Skeletal variations                mean

% offspring per litter            ±SEM

10.0

2.0

3.4

1.0

6.5

1.8

5.3

1.4

7.4

2.1

12.1

3.0

Visceral malformations         mean

% offspring per litter            ±SEM

39.9

7.0

40.6

6.3

44.3

6.5

42.9

6.6

45.4

6.9

46.4

6.5

Visceral variations               mean

% offspring per litter            ±SEM

1.4

0.8

2.1

1.2

2.9

1.5

0.0

0.0

1.3

0.7

0.5

0.5

Conclusions:
The teratogenicity of the test substance was assessed according to OECD guideline 414. There was no evidence of developmental toxicity in offspring of rats fed boric acid in diet throughout gestation up to 0.075 % (55 mg/kg bw boric acid). At 0.100 % boric acid (76 mg/kg bw boric acid) there was reduced fetal bodyweight, short and wavy ribs, and these effects disappeared during the postnatal period. Similar but more marked effects were observed at the highest dose of 0.200 % (143 mg/kg bw boric acid) and apart from the short 13th rib, they also disappeared during the postnatal period.
Read-across is justified on the basis detailed in the rationale for reliability above. This study is therefore considered to be of sufficient adequacy and reliability to be used as a supporting study and no further testing is justified.
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
No data
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Comparable to a guideline study. This study is conducted on an analogue substance. Read-across is justified on the following basis: In aqueous solutions at physiological and acidic pH, low concentrations of simple inorganic borates such as boric acid, disodium tetraborate decahydrate, disodium tetraborate pentahydrate, boric oxide and disodium octaborate tetrahydrate will predominantly exist as undissociated boric acid. At about pH 10 the metaborate anion (B(OH)4-) becomes the main species in solution (WHO, 1998). This leads to the conclusion that the main species in the plasma of mammals and in the environment is un-dissociated boric acid. Since other borates dissociate to form boric acid in aqueous solutions, they too can be considered to exist as un-dissociated boric acid under the same conditions. For comparative purposes, exposures to borates are often expressed in terms of boron (B) equivalents based on the fraction of boron in the source substance on a molecular weight basis. Some studies express dose in terms of B, whereas other studies express the dose in units of boric acid. Since the systemic effects and some of the local effects can be traced back to boric acid, results from one substance can be transferred to also evaluate the another substance on the basis of boron equivalents. Therefore data obtained from studies with these borates can be read across in the human health assessment for each individual substance. Conversion factors are given in the table below. Conversion factor for equivalent dose of B Boric acid H3BO3 0.175 Boric Oxide B2O3 0.311 Disodium tetraborate anhydrous Na2B4O7 0.215 Disodium tetraborate pentahydrate Na2B4O7•5H2O 0.148 Disodium tetraborate decahydrate Na2B4O7•10H2O 0.113 Disodium octaborate tetrahydrate Na2B8O13•4H2O 0.210 Sodium metaborate (anhydrous) NaBO2 0.1643 Sodium metaborate (dihydrate) NaBO2•2H2O 0.1062 Sodium metaborate (tetrahydrate) NaBO2•4H2O 0.0784 Sodium pentaborate (anhydrous) NaB5O8 0.2636 Sodium pentaborate (pentahydrate) NaB5O8∙5H2O 0.1832 References: WHO. Guidelines for drinking-water quality, Addendum to Volume 1, 1998.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
not specified
GLP compliance:
yes
Limit test:
no
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Hazleton Research Products Inc., Denver, PA, USA
- Age at study initiation: 5 months of age
- Weight at study initiation: 2690-4380 g
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
VEHICLE
- Concentration in vehicle: 55 mg/mL boric acid
- Amount of vehicle: 5 mg/mL boric acid
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
No data
Details on mating procedure:
- Impregnation procedure: Artificial insemination; designated Day 0
Duration of treatment / exposure:
Groups of 30 rabbits were used treated on Day 6 - 19 post-mating
Frequency of treatment:
No data
Duration of test:
Terminated on Day 30 of gestation
No. of animals per sex per dose:
30 females/group
Control animals:
yes
Details on study design:
No data
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 30
- Organs examined: Uterus
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: No data
- Number of late resorptions: No data
- Other:
Uteri were stained with ammonium sulphide as no implantations were visible.
Litter size, number of dead foetuses and foetal weight were assessed.
Fetal examinations:
- External examinations: Yes, including assessment for cleft palate
- Soft tissue examinations: Yes by dissection (Staples) and sex determined
- Skeletal examinations: Yes, all foetuses were skinned and cleaned and stained with alcian blue/alizarin red S
- Head examinations: Yes
Statistics:
No data
Indices:
No data
Historical control data:
No data
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Pregnant does exhibited no overt symptoms attributable to boric acid toxicity except in the high dose group. A decreased food intake (30 % reduction vs. controls during exposure period) and decreased maternal bodyweight were observed. Vaginal bleeding was noted at 43.5 mg B/kg bw between gestational days 19 - 30. All high-dose animals with vaginal bleeding had no live foetuses at sacrifice. At mid dose, increased body weight gain not clearly adverse. The authors considered 43.5 mg B/kg bw as the LOAEL for pregnant does and 21.8 mg B/kg bw as the NOAEL for maternal toxicity.
Dose descriptor:
LOAEL
Effect level:
250 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
125 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
250 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEL
Effect level:
125 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Dose descriptor:
LOAEL
Effect level:
43.5 mg/kg bw/day
Based on:
element
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
21.8 mg/kg bw/day
Based on:
element
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
43.5 mg/kg bw/day
Based on:
element
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEL
Effect level:
21.8 mg/kg bw/day
Based on:
element
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
At the highest dose in this study of 250 mg/kg bw boric acid (43.5 mg B/kg bw/day), 90 % of implants/litter were resorbed compared to 6 % for controls, and 73 % had complete litter loss (0 % in controls). In the mid and low dose groups, no difference in percentage resorptions per litter was seen, compared to controls.
Average foetal bodyweight per litter was 92 % of the controls at the high dose (43.5-mg B/kg bw) but even at this exposure, it did not reach statistical significance possibly due to the low number of pups surviving (14 fetuses from 6 litters).
An increased incidence of malformed live foetuses/litter was observed at 43.5 mg B/kg bw, primarily due to cardiovascular defects (72 % for major defects of heart and/or great vessel in the high-dose group vs. 3 % in controls). In the mid and low dose groups, there was no increase in malformations per litter or total malformations. There were no variations between any groups concerning the incidence of skeletal malformations.
The only skeletal variations of interest was a dose related reduction in the incidence of extra ribs on Lumbar I which the authors did not consider to be toxicologically important.
Since no definitive developmental effects were observed in animals exposed to either 62.5 or 125 mg/kg bw boric acid (10.9 or 21.8 mg B/kg bw/day), the authors concluded that 125 mg/kg boric acid per day (21.8 mg B/kg bw/day) was the NOAEL for developmental toxicity.
Dose descriptor:
NOAEL
Effect level:
21.8 mg/kg bw/day
Based on:
element
Sex:
not specified
Basis for effect level:
changes in litter size and weights
other: major defects of heart and/or great vessel
Dose descriptor:
LOAEL
Effect level:
43.5 mg/kg bw/day
Based on:
element
Sex:
not specified
Basis for effect level:
changes in litter size and weights
other: major defects of heart and/or great vessel
Abnormalities:
not specified
Developmental effects observed:
not specified

Maternal effects

Parameter

Study

control

data

Low

dose

Medium

dose

High

dose

Number of dams examined

30

30

30

30

Clinical findings during application of test substance

 

 

 

reduced food and bodyweight

Mortality of dams (%)

0

1

1

0

Abortions

0

0

0

3

Body weight gain               day 6-19

                                             day 0-30

93

357

132

493

97

543

-137*

226

Food consumption g/kg/day  day 0-6

                                                 day 6-19

                                                 day 19-25

48.1

38.8

36.9

48.0

40.0

37.0

48.9

38.7

40.0

46.4

26.6*

44.9

Pregnancies  % pregnant at sacrifice

75

89

87

96

Necropsy findings in dams dead before end of test

 

gavage error lungs

stomach damage

 

* P < 0.05.

 

Litter response (Caesarean section data)

Parameter

Control

data

Low

dose

Medium

dose

High

dose

Corpora lutea                        number

                                                 ±SEM

mean No. per dam

12.2

0.7

10.7

0.5

11.5

0.4

10.0*

0.7

Implantations sites per litter     number

                                                     ±SEM

total/number of dams

9.5

0.8

8.4

0.6

8.3

0.5

8.6

0.7

Resorptions % per litter               %

total/number of dams          ±SEM

6.3

2.4

5.9

1.9

7.7

2.1

89.9

5.0

total number of foetuses

159

175

153

14

total number of litters

18

23

20

6

live foetuses / litter                 number

state ratio                               ±SEM

8.8

0.8

7.6

0.6

7.7

0.5

2.3*

0.8

dead foetuses / litter        %

state ratio

0

2.8

0.4

0

foetus weight (mean)                   weight (g)

                                                        ±SEM

44.8

1.5

46.5

1.4

45.7

1.2

41.1

2.7

Foetal sex ratio    % male per litter

                                   ±SEM

50

5

51

4

55

4

69

10

* P<0.05


 

Examination of the foetuses

Parameter

Control

data

Low

dose

Medium

dose

High

dose

External malformations*

 % foetuses per litter  ±SEM

0.8

0.8

1.4

1.0

1.0

1.0

11.1*

8.2

External malformations    No. of foetuses

1

2

1

2

Skeletal malformations*

% foetuses per litter ±SEM

19.9

5.4

19.9

4.0

24.3

6.4

38.9

20.0

Skeletal malformations     No. of foetuses

30

39

44

4

Visceral malformations*

% foetuses per litter  ±SEM

7.3

1.9

5.9

2.0

7.4

2.0

80.6*

16.3

Visceral malformations      No. of foetuses

13

11

12

11

% foetuses with cardiovascular       % malformations                               ±SEM

2.7

1.6

3.1

1.5

4.2

1.3

72.2*

16.5

* P<0.05

Conclusions:
The highest dose was very toxic to dams and 90 % of implants were resorbed at the highest dose level, and 72 % of surviving foetuses had cardiac or great vessel malformations or increase in resorptions were reported in the mid and low dose groups.
Read-across is justified on the basis detailed in the rationale for reliability above. This study is therefore considered to be of sufficient adequacy and reliability to be used as a supporting study and no further testing is justified.
Effect on developmental toxicity: via oral route
Dose descriptor:
BMDL05
70.1 mg/kg bw/day
Additional information

Assessment entity approach

"Brazing fluxes" are mixtures of boron-containing constituents (potassium(fluoro)borates), which undergo chemical exchanges (anion exchange) and condensation reactions (e.g. formation of oligoborates, polyborates) upon mixing and further manufacturing. This results in a complex mixture of potassium borates, which cannot be fully chemically characterised for substance identity. Thus, according to the definition under REACH, such brazing fluxes must be described as a UVCB substance.

 

Data specifically on the UVCB substance to be registered are not available. An assessment entity approach is followed based on the transformation products of this UVCB uppon dissolution in aqueous media. The substance is highly soluble and forms complex boron, potassium and fluoride constituents. The quantitatively predominant transformation product of this UVCB is represented by boric acid, which is assumed to be the determinant of human health effects because of its classification and its toxicity. For this reason, the assessment is based on information for “borates” (including potassium borate, boric acid and other borate substances).

 

Based on the information provided below, it may safely be assumed that under physiological conditions the chemical speciation of most of the unknown potassium boron compounds corresponds to boric acid. Thus, from a chemical point of view, there is no reason to assume that brazing fluxes would behave differently than boric acid and/or borates under physiological conditions.

 

The basis of this assessment entity approach is further justified by the following reasoning:

In aqueous solutions at physiological and acidic pH, low concentrations of simple inorganic borates such as boric acid B(OH)3, potassium pentaborate (K2B10O16*8H2O), potassium tetraborate (K2B4O7*4H2O), disodium tetraborate decahydrate (Na2B4O7.10H2O; borax), disodium tetraborate pentahydrate (Na2B4O7*5H2O; borax pentahydrate), boric oxide (B2O3) and disodium octaborate tetrahydrate (Na2B8O13*4H2O) will predominantly exist as undissociated boric acid. Above pH 9 the metaborate anion (B(OH)4-) becomes the main species in solution (WHO, 1998). This leads to the conclusion that the main species in the plasma of mammals and in the environment is undissociated boric acid. Since other borates dissociate to form boric acid in aqueous solutions, they too can be considered to exist as undissociated boric acid under the same conditions.

For comparative purposes, exposures to borates are often expressed in terms of boron (B) equivalents based on the fraction of boron in the source substance on a molecular weight basis. Some studies express dose in terms of B, whereas other studies express the dose in units of boric acid. Since the systemic effects and some of the local effects can be traced back to boric acid, results from one substance can be transferred to also evaluate the another substance on the basis of boron equivalents. Therefore data obtained from studies with these borates can be read across in the human health assessment for each individual substance. Conversion factors are given in the table below.

 

Substance

Formula

Conversion factor for equivalent dose of B (multiply by)

Boric acid

H3BO3

0.1748

Boric Oxide

B2O3

0.311

Disodium tetraborate anhydrous

Na2B4O7

0.2149

Disodium tetraborate pentahydrate

Na2B4O7•5H2O

0.1484

Disodium tetraborate decahydrate

Na2B4O7•10H2O

0.1134

Disodium octaborate tetrahydrate

Na2B8O13·4H2

0.2096

Sodium metaborate (anhydrous)

NaBO2

0.1643

Sodium metaborate (dihydrate)

NaBO2·2H2O

0.1062

Sodium metaborate (tetrahydrate)

NaBO2·4H2O

0.0784

Sodium pentaborate (anhydrous)

NaB5O8

0.2636

Sodium pentaborate (pentahydrate)

NaB5O8∙5H2O

0.1832

 Dipotassium tetraborate (anhydrous)

 

 K2B4O7

 

 0.185

 

 Dipotassium tetraborate (tetrahydrate)

 

 K2B4O7.4H2O

 

 0.1415

 

 Potassium pentaborate (anhydrous)

 

 B5KO8

 

 0.244

 

 Potassium pentaborate (tetrahydrate)

 

 B5KO8.4H2O

 

 0.1843

 

 

Reference: WHO. Guidelines for drinking-water quality, Addendum to Volume 1, 1998

Discussion (effects on developmental toxicity, data for borates):

Developmental effects have been observed in three species, rats, mice and rabbits. The most sensitive species being the rat with a NOAEL of 9.6 mg B/kg bw/day. This is based on a reduction in mean foetal body weight/litter, increase in wavy ribs and an increased incidence in short rib XIII at 13.3 mg B/kg bw/day. The reduction in foetal body weight and skeletal malformations had reversed, with the exception of short rib XIII, by 21 days post natal. At maternally toxic doses, visceral malformations observed included enlarged lateral ventricles and cardiovascular effects.

The NOAEL for this endpoint is 9.6 mg B/kg bw/day corresponding to 55 mg boric acid/kg bw/day; 85 mg disodium tetraborate decahydrate/kg, 65 mg disodium tetraborate pentahydrate/kg and 44.7 mg disodium tetraborate anhydrous/kg.

The critical effect is considered to be decreased fetal body weight in rats, for which the NOAEL was 9.6 mg/kg body weight per day. A benchmark dose developed by Allen et al. (1996) was based on the studies of Heindel et al. (1992), Price, Marr & Myers (1994) and Price et al. (1996). The benchmark dose is defined as the 95% lower bound on the dose corresponding to a 5% decrease in the mean fetal weight (BMDL05). The BMDL05of 10.3 mg/kg body weight per day as boron is close to the Price et al. (1996) NOAEL of 9.6 mg/kg body weight per day.

There is no evidence of developmental effects in humans attributable to boron in studies of populations with high exposures to boron (Tuccar et al 1998; Col et al. 2000; Chang et al. 2006).

BDML05:

Potassium pentaborate anhydrous: 55.7 mg/kg bw/day

Potassium pentaborate tetrahydrate: 72.8 mg/kg bw/day

Brazing fluxes (13.7 wt-% boron): 70.1 mg/kg bw/day (stoichiometric recalculation)

Justification for classification or non-classification

Boric acid and disodium tetraborate are classified under the 1stATP to CLP as Repr. 1B; H360FD.

However, text of the 30th ATP as published in the EU Official Journal, 15 September 2008 stated that “The classification and labelling of the substances listed in this Directive should be reviewed if new scientific knowledge becomes available. In this respect, considering recent preliminary, partial and not peer-reviewed information submitted by industry, special attention should be paid to further results of epidemiological studies on the Borates concerned by this Directive including the ongoing study conducted in…”

While boron has been shown to adversely affect male reproduction in laboratory animals, there was no clear evidence of male reproductive effects attributable to boron in studies of highly exposed workers (Whorton et al. 1994; Sayli 1998, 2001; Robbins et al. 2010; Scialli et al. 2010). Not only are these the most exposed workers, but the Chinese worker study is themost sensitive study that has been carried out as semen analysis was performed, a very sensitive detection system for testicular damage. There is no evidence of developmental effects in humans attributable to boron in studies of populations with high exposures to boron (Tuccar et al 1998; Col et al. 2000; Chang et al. 2006).

Comparison of Blood, Semen and Testes Boron Levels in Human and Rat

A comparison of blood, semen and target organ boron levels in studies of laboratory animals and human studies shows that boron industry worker exposures are lower than untreated control rats. Background boron levels in standard rat chow are high (10-20 ppm), as a result control rats in toxicity studies receive 45 times more boron than background exposure in humans. Blood boron levels in female control rats is about 0.23 µg B/g (Price et al. 1997), approximately equal to the blood levels in boron industry workers in China, Turkey and U.S. of 0.25, 0.22 and 0.26 µg B/g, respectively (Scialli et al. 2010; Culver et al. 1994; Duydu et al. 2011). Plasma and seminal vesicle fluid (the major component of semen) boron levels in untreated male control rats were 1.94 and 2.05 µg B/g, respectively, while boron levels in testes in rats dosed at the rat fertility LOAEL (26 mg B/kg) was 5.6 µg B/g (Ku et al. 1991,1993). Values in male control rats were higher than corresponding boron levels in the highest exposed Chinese boron industry workers with blood boron levels of 1.56 µg B/g and 1.84 µg B/g in semen (Scialli et al. 2010). Blood and semen boron levels in highly exposed Turkish boron workers were also lower than control rats with levels of 0.22 and 1.88 µg B/g, respectively (Duydu et al. 2011). Boron levels in testes of rats dosed at the rat fertility LOAEL was over 3x the blood boron levels in highest exposure group of Chinese boron industry workers. The blood level at the lowest animal LOAEL (13 mg B/kg) was 1.53 µg B/g, about 6 times greater than typical boron industry workers (Price et al. 1997). No adverse effects on sperm were seen in Turkish boron industry workers or in the most highly exposed subgroup of Chinese boron industry workers drinking boron contaminated water (mean blood level 1.52 µg B/g, the human NOAEL). Only under extreme conditions do human levels reach those of the animal LOAEL: the subgroup of Chinese boron workers who also drank contaminated water. Since no boron accumulation occurs in soft tissues (testes) over plasma levels biological monitoring in humans provide direct comparison to test animal target organ boron levels. Workers in boron mining and processing industries represent the maximum possible human exposure however their blood and semen boron levels are less than levels in untreated control rats. This provides an explanation why studies of highly exposed boron industry workers have shown no adverse effects and demonstrates that maximal possible exposures in humans are insufficient to cause reproductive toxicity effects. Graphs comparing the rodent and human exposure, blood, semen and tissue boron levels are presented in Appendix C.

A weight of evidence approach was used in evaluating numerous independent studies on the determination of the hazard of boric acid to humans. Information that was considered together included results of in vitro tests, animal data, occupational exposure data, epidemiological studies and mechanistic data.

Extensive evaluations of sperm parameters in highly exposed workers in Turkey and China have demonstrated no effects on male fertility. No evidence of developmental effects in humans attributable to boron (B) has been observed in studies of populations with high exposures to boron. Although the epidemiological studies have methodological deficiencies, collectively these studies consistently show an absence of effects in highly exposed populations.

Workers in boron mining and processing industries represent the maximum possible human exposure. However a comparison of blood, semen and target organ boron levels in studies of laboratory animals and human studies shows that boron industry worker exposures are lower than untreated control rats.

Mechanistic data provide possible explanations for the absence of developmental and reproductive effects in humans exposed to high levels of boron. Recent studies provide evidence that boric acid may act by similar mechanisms in causing developmental effects in mice as sodium salycilate (the natural deacetylated form of aspirin and a rodent teratogen) including effects on Hox gene expression and inhibition of embryonic histone deacetylases. Although aspirin is known to cause developmental effects in laboratory animals, controlled human studies have not demonstrated developmental effects in humans. Similar mechanisms of action of boric acid and aspirin, and the absence of developmental effects in humans ingesting aspirin suggest that boric acid related developmental effects in humans are unlikely. 

Additionally, zinc levels in soft tissue in humans is over 2 times greater than in comparative tissues in rats (King et al. 2000; Yamaguchi et al. 1996), which explain in part the absence of fertility and developmental effects in humans. Zinc has been shown to protect against testicular toxicity of cobalt and cadmium (Anderson et al. 1993), and the developmental effects of cadmium (Fernandez et al. 2003). There is evidence that zinc interacts with boric acid in the body reducing the toxicity of boric acid. The interaction of zinc and boric acid is evident by the low acute toxicity of zinc borate (absorbed as boric acid and zinc) with a LD50 value greater than 10,000 mg/kg-body weight in rats (Daniels 1969) compared to disodium tetraborate pentahydrate (similar % boron composition as zinc borate) with a LD50 value of 3300 mg/kg-body weight. Furthermore, no toxic effects were observed in the testes of males (a target organ of boric acid) administered 1000 mg zinc borate/kg/day in a 28-day repeated dose oral gavage toxicity study, equivalent dose of boron of 50 mg B/kg bodyweight (Wragg et al. 1996). The LOAEL for testicular effects is 26 mg B/kg body weight. 

Based on the total weight of evidence, the data show that it is improbable that boric acid or potassium pentaborate will cause reproductive or developmental effects in humans. Therefore, based on a total weight of evidence, Category 2 H361d: suspected human reproductive toxicant, suspected of damaging the unborn child is considered the appropriate classification.

Because borates are considered the transformation product driving the human health effects for brazing fluxes (see 'discussion`),

this weight of evidence approach is also applicable for brazing fluxes.