Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

One study is available. This study was performed prior to the introduction of GLP.

Link to relevant study records

Referenceopen allclose all

Endpoint:
three-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
no guideline followed
Principles of method if other than guideline:
A reproduction study spanning 3 generations of rats reared and maintained on diets containing 0.0% and 0.5% HMP (Hexametaphosphate). Per generation two different litters were produced; one was sacrificed and the other was bred to produce the next generation. Matings were carried out between 16 females and 8 males in each group when the rats were 100 days old. Observations were limited to: the numbers of pups born, pup mortality up to 21 days, organ weights and histological changes of tissues and organs.
GLP compliance:
no
Remarks:
study predates GLP
Limit test:
no
Species:
rat
Strain:
other: Rochester (Ex-Wistar 1923)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: The original groups of rats were mated at 100 days
- Weight at study initiation: Bodyweight was approximately 70 grams for males and females at the start of the 100 day exposure.
- Fasting period before study:
- Housing: cages
- Diet (e.g. ad libitum): ad libitum,
- Water (e.g. ad libitum): ad libitum

ENVIRONMENTAL CONDITIONS
no data

IN-LIFE DATES: From: *** To: ***
Route of administration:
oral: feed
Type of inhalation exposure (if applicable):
other: not applicable
Vehicle:
unchanged (no vehicle)
Details on exposure:
DIET PREPARATION

Diet mixtures were prepared using a basal ration of Purina Fox Chow Meal into which the appropriate amounts of sodium hexametaphosphate were mixed by a mechanical mixer. At weekly intervals. Diets were stored during the week in galvanized iron pails with covers.
Details on mating procedure:
- M/F ratio per cage: 1male: 2 female
- Length of cohabitation: 7 days
- Proof of pregnancy: no data
- Unsuccessful pairing replacement of first male by another male with proven fertility?: no.
- Further matings after two unsuccessful attempts: no data
- After successful mating each pregnant female was caged (how): no data
- other: males were rotated so that at each mating a different male would be placed in the cage with the female.

Details on mating schedules are tabulated below. - Table 1.
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
not applicable
Duration of treatment / exposure:
Exposure took place for 100 days prior to mating, 21 days of gestation and during lactation for the P1, F1b, F2b. The F3b were exposed for their first 21days after which time they were sacrificed. Litters were culled to ten pups with approximately 5/sex/group on postnatal day 5.
For the F1, F2, F3-generations, offspring were weaned and exposed during an interim period of 10 days and then during a mating period of 7 days. Dams were exposed continually during gestation and lactation. Offspring of the second litter were again mated for 7 days after a 10 day interim period following weaning. The first litters (F1a, F2a, F3a) were sacrificed at 30 days of age. The F1c and F3b litters were sacrificed after 21 days.
Frequency of treatment:
daily in feed
Details on study schedule:
- F1 parental animals were mated at 100 days of age in order to produce the sacrificial group F2a, and at 151 days F1 animals were mated to produce the next parental generation for F2b.
- Per female the litters were culled to ten pups with approximately 5/sex/group. There are no details on the selection of parental animals.
- Age at mating of the mated animals in the study: 100 days to produce the sacrifical litter and at 151 days to produce the next parental generation.

More details on protocol are tabulated below. - Table 1.
Remarks:
Doses / Concentrations:
0.0% and 0.5%
Basis:
nominal in diet
No. of animals per sex per dose:
16 females and 8 males were used to produce the subsequent generation. P1 , F1b , F2b , F3b.
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: Two doses were considered; 0.05% and 0.5%. After 100 days on these diets the 0.5% group exhibited no depression of body weight and was selected for use in this study.
- Other: The first litters born per generation were sacrificed, the F1a, F2a, F3a. The second litters born in a generation, the F1b, F2b, were mated to produce the next generation. The F1c was an additional litter produced and treated the same as the F1a.
Positive control:
no data
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: No

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were measured weekly during the 100 day pre-mating exposure for each generation.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Not measured

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Not measured
Oestrous cyclicity (parental animals):
not evaluated
Sperm parameters (parental animals):
not evaluated
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 5 postpartum: yes
- excess pups were sacrificed to leave 10 per litter.

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring:
[number of pups, pups/litter, pup mortality up to 21 days of age, weight gain per litter]

GROSS EXAMINATION OF DEAD PUPS: No
Postmortem examinations (parental animals):
SACRIFICE
- The first litters prodced per generation, the F1a, the F2a and the F3a were sacrificed at 30 days of birth.
- There is no data on the fate of the parents after they were mated to produce each generation.

GROSS NECROPSY
no data

HISTOPATHOLOGY
No histopathological examinations were performed for parental rats.

ORGAN WEIGHTS
No organ weight analyses were performed for parental rats.
Postmortem examinations (offspring):
SACRIFICE
- The non-parental litters, F1a , F1c, F2a , F3a , and F3b were sacrificed after 21 days.
- Observations amongst these litters were limited to litter weights on the 21st day. In the F3b histopathological and organ weight examinations were performed.

GROSS NECROPSY
F3b examined only.

HISTOPATHOLOGY / ORGAN WEIGTHS
The ten males and ten females from final litter, the F3b litter, were evaluated for body weight, histopathology (liver, kidneys, gonads, lungs, brain, stomach, heart, spleen, adrenal, stomach, large and small intestines, bladder, lymph nodes, gut, marrow, pancreas) and organ weights (liver, kidneys, testes, lungs, brain, stomach, heart, spleen).
Statistics:
no data
Reproductive indices:
no data
Offspring viability indices:
no data
Clinical signs:
not examined
Body weight and weight changes:
not examined
Food consumption and compound intake (if feeding study):
not examined
Organ weight findings including organ / body weight ratios:
not examined
Histopathological findings: non-neoplastic:
not examined
Other effects:
not examined
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)

In Table 2. a summary is presented of the reproductive performance.
Number of females mated: In all cases 15 or 16 females were mated after a 7-day cohabitation with one male (1 male caged with 2 females).
Number of pregnancies: Most of the pregnancies were successful. For the control rats there were 10 to 15 pregnancies per litter and for the rats maintained on the diet containing 0.5% HMP there were 10 to 13 in each mating.
Number of rats born: Comparable between the control and 0.5% group, the average number of pups ranged from 8-10 in both the controls and 0.5% group.
Key result
Dose descriptor:
NOAEL
Effect level:
0.5 other: %
Based on:
test mat.
Sex:
male/female
Key result
Dose descriptor:
NOAEL
Effect level:
ca. 250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: dose level calculated
Key result
Critical effects observed:
no
Clinical signs:
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
not examined
Histopathological findings:
no effects observed
VIABILITY (OFFSPRING)
The number of pups surviving at 21 days was 65-122 in the control groups and 69-111 in the 0.5% HMP group.
The average weight of the pups at 21 days was 41.8 - 48.7 g in the control groups and 40.9 - 45.4g in the 0.5% HMP group.

BODYWEIGHT (OFFSPRING)
After 21 days the newly born rats surviving to 21 days were weighed per litter, differences between the control and HMP-fed rats were not considered to be significant. See Table 2.

F1a - The average bodyweight of rats on the HMP diet was less than that of the control group.
F1b - The average bodyweight of rats on the HMP diet was less than that of the control group.
F1c - The average bodyweight of rats on the HMP diet was greater than that of the control group - based on this the authors considered the decreased average litter weights of the F1a and F1b to be mere coincidental variations.

F2a - The average bodyweight of rats on the HMP diet was less than that of the control group.
F2b - There was a small difference, less than a gram, in the average weight of the control and HMP-fed rats. There was a peculiar delay of the HMP-fed rats within the first week on the diet, but growth was rapidly recovered and surpassed that of the control group.

F3a - The average weight of the HMP-fed and control rats was almost identical at 42.5 and 42.4 gram respectively
F3b - The average weight of the MHP-fed rats was greater than that of the controls.

ORGAN WEIGHTS (OFFSPRING)
When the F3b rats were at weaning age they were sacrificed. The author described the organ weights and organ weight body weight ratios as, being within normal ranges. See Table 3

HISTOPATHOLOGY (OFFSPRING)
No histological changes were found that were attributed to the presence of HMP in the diet. No tumours were present and only normal tissues were found.
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
0.5 other: % w/w
Based on:
test mat.
Sex:
male/female
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: dose level calculated
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
0.5 other: % w/w in diet
Based on:
test mat.
Sex:
male/female
Key result
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: dose level calculated
Key result
Key result
Reproductive effects observed:
no

Table 2. Reproductive performance

 Generation & Litter

F1a

F1b

F1c

F2a

F2b

F3a

F3b

 Controls

Females

16

15

15

16

15

16

16

Pregnancies

14

10

10

11

10

15

11

 

 

 

 

 

 

 

 

Total pups

107

95

77

94

93

145

115

Avg. pups/litter

7.6

9.5

9.6

8.5

9.3

9.7

10.4

 

 

 

 

 

 

 

 

Mortality (days 0-5)

0

1

3

0

10

8

10

Mortality (days 6-21)

0

1

5

3

4

9

6

 

 

 

 

 

 

 

 

Total rats surviving to day 21

107

84

65

87

77

122

90

Avg. weight of surviving rats

48.7

45.2

42.0

44.9

41.8

42.4

42.9

0.5% Hexametaphosphate group

Females

16

15

15

16

16

16

16

Pregnancies

11

10

13

13

10

12

10

 

 

 

 

 

 

 

 

Total pups

83

100

126

118

89

119

97

Avg. pups/litter

8.3

10.0

9.7

9.1

8.9

9.9

9.7

 

 

 

 

 

 

 

 

Mortality (days 0-5)

6

15

3

11

2

1

3

Mortality (days 6-21)

5

1

0

9

11

3

0

 

 

 

 

 

 

 

 

Total rats surviving to day 21

69

77

111

95

73

108

87

Avg. weight of surviving rats

43.1

43.5

44.7

44.3

40.9

42.5

45.4

Table 3 - Organ Weights

Group

No. of Rats

Body Wt.

Liver

Kidneys

Testes

Lungs

Brain

Stomach

Heart

Spleen

Control

Males

10

111

4.84

1.28

1.20

0.80

1.59

1.04

0.55

0.59

0.5%HMP-fed males

10

102

4.74

1.16

1.04

0.82

1.49

1.07

0.56

0.37

Control Females

10

96

4.03

1.06

n/a

0.69

1.43

1.02

0.53

0.64

0.5%HMP-fed females

10

90

3.71

1.03

n/a

0.67

1.54

1.09

0.47

0.36

Conclusions:
Three generations of rats reared and maintained on diets containing 0.5% HMP showed no adverse effects to the test material - Control and test data were comparable across the range of observations made. Observations were limited to reproductive performance, pup mortality, organ weights and tissue and organ histopathology. Based on this evidence sodium metaphosphate is not considered to be classified as a reproductive toxicant according to Regulation (EC) 1272/2008 (EU CLP).

Although this study is limited and deficient by modern standards, it is still possible to make some valid scientific conclusions from the data. Adequate data are reported to show no effects on fertility or reproductive performance or on offspring growth and development over three generations with two litters per generation. Thus, the study should be considered acceptable for the dose level tested, 0.5% test material in the diet.
Endpoint:
three-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
Well documented study with the following deviations: Only one dose level was included in the study. Test material was not analyzed in diet. No impurity profile of test material was provided. Source of animals was not specified. The group size was smaller than the 20 females and 10 males required in OECD guideline 416. There was no recorded acclimation period for the P generation. Only one dose tested. Daily observation of clinical signs of parents and observations of abnormal behaviour of offspring are not reported. Body Weights: parental body weights were not recorded during gestation and lactation. Food consumption was not determined. Estrus cycle was not evaluated. Sperm parameters were not included. Offspring sex and sex ratios were not determined. Litter body weight was reported; however, individual pup body weights were not recorded. There is no report of gross pathology and necropsy observations. Organ weight data for the uterus, ovaries, epididymides, prostate, seminal vesicles, pituitary gland, adrenals and thyroid gland were not included. Histopathology was conducted only on the F3b offspring and did not include the following organs: uterus, cervix, vagina, epididymides, seminal vesicles, prostate, coagulating gland, pituitary gland. Note that the known target organ (kidney) was evaluated. Statistics are not reported. This study is considered to fulfil the data requirements as a supporting study for REACH.
Qualifier:
no guideline followed
Principles of method if other than guideline:
Three generations of rats were reared on diets containing 0 and 0.5% sodium tripolyphosphate. Two litters were examined in each generation. Matingwas carried out between 16 females and 8 males of each group when the rats were 100 days old.
GLP compliance:
no
Remarks:
Study predates GLP
Limit test:
no
Species:
rat
Strain:
other: Rochester Strain (Ex-Wistar 1923)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Weight at study initiation: 60 g
- Diet: e.g. ad libitum
- Water :e.g. ad libitum
Route of administration:
oral: feed
Details on exposure:
Exposure took place for 100 days prior to mating, 21 days of gestation and during lactation. For the F1, F2, F3-generations, offspring were weaned and exposed during an interim period of 10 days and then during a mating period of 7 days. Dams were exposed continually during gestation and lactation. Offspring of the second litter were again mated for 7 days after a 10 day interim period following weaning. The first litters (F1a, F2a, F3a) were sacrificed at 30 days of age. The second litters (F1b, F2b, F3b) were used to produce the succeeding generation.
Details on mating procedure:
Each male was caged with two females for 7 days until conception occurred.
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
Exposure took place for 100 days prior to mating, 21 days of gestation and during lactation. For the F1, F2, F3-generations, offspring were weaned and exposed during an interim period of 10 days and then during a mating period of 7 days. Dams were exposed continually during gestation and lactation. Offspring of the second litter were again mated for 7 days after a 10 day interim period following weaning. The first litters (F1a, F2a, F3a) were sacrificed at 30 days of age. The second litters (F1b, F2b, F3b) were used to produce the succeeding generation.
Remarks:
Doses / Concentrations:
0.5% in diet
Basis:
nominal in diet
No. of animals per sex per dose:
P1, F1, F2, F3: 16 females and 8 males
Control animals:
yes, concurrent vehicle
Sperm parameters (parental animals):
Not evaluated
Litter observations:
number of pups, stillbirths, live births, weight gain per litter
Not Evaluated: sex of pups, presence of gross anomalies
physical or behavioural abnormalities, , no individual pup body weights
Postmortem examinations (offspring):
F3b generation groups of male (8/group) and female (16/group) rats were fed control diet or diet with 0.5% test material for 100 days (P1) generation. Body weights were measured weekly during the 100 day pre-mating exposure for each generation. Each male was caged with two females for 7 days until conception occurred. Pregnancy rates were determined. The first litter born of the parental animals was designated as the F1a litter. Upon delivery, offspring were counted and average litter weights determined. On day 5 pups were culled to 10 per litter. Body weights per litter were determined on day 21. Pup survival was determined on days 1-5 and days 6-21. The F1a litter was sacrificed at 30 days postnatally. Parental animals were mated again 10 days after the sacrifice of the F1a litter. Mating took place for 7 days. A second litter (F1b) was born after approximately 21 days of gestation. Upon delivery, offspring were counted and the average litter weights determined. On day 5 pups were culled to 10 per litter. Body weights per litter were determined on day 21. Pup survival was determined on days 1-5 and days 6-21. Following weaning of the F1b litter, 8 males and 16 females for the control group and the treated group were fed their respective diets for 100 days. Again two litters (F2a and F2b) were produced. Parental animals were mated for 7 days following 10 days after the sacrifice of the F2a litter (pups were 30 days old). Following birth of the F2b litter the same offspring and parental parameters were measured. Eight males and 16 females of the F2b litter were designated for continual exposure for another 100 days. Upon completion of the exposure of the F2b litter, animals were mated again to produce the F3a and F3b litters in the same manner described above. The ten males and ten females from final litter, the F3b litter, were evaluated for body weight, histopathology (liver, kidneys, gonads, lungs, brain, stomach, heart, spleen, adrenal, stomach, large and small intestines, bladder, nodes, marrow, pancreas) and organ weights (liver, kidneys, testes, lungs, brain, stomach, heart, spleen). Individual animal data were collected and recorded throughout the study.
Organ weights:
liver, kidneys, gonads, lungs, brain, stomach, heart, spleen on F2b offspring.
Not evaluated: Uterus, ovaries, epididymides (total and cauda), prostate, seminal vesicles, pituitary, thyroid, adrenal glands
Histopathology:
Kidneys, heart, brain, spleen, lungs, ovaries, testes, adrenal, bladder, lymph nodes, gut, bone marrow, pancreas on F3b animals.
Not Evaluated: Vagina, uterus with cervix, epididymis, seminal vesicle, prostate coagulating gland
Key result
Dose descriptor:
NOAEL
Effect level:
0.5 other: % w/w
Based on:
test mat.
Sex:
male/female
Key result
Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: dose level calculated
Key result
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
0.5 other: % w/w
Based on:
test mat.
Sex:
male/female
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: calculated
Key result
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
0.5 other: % w/w in diet
Based on:
test mat.
Sex:
male/female
Key result
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: calculated
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no

This study included only one dose level of the test material, 0.5% in the diet. 

NOAEL calculated in accorance with Appendix F, guidelines for the preparation of toxicological working papers for the joint FAO/WHO expert committee on food additives, December 2000.

There were occasional fluctuations in the fertility index.  The lower fertility index in the F2a, F3a and F3b litters is considered spurious and not treatment-related.  The fertility indices of the treated group were greater than the control group for some litters (F1a, F2b); the same as the control group for the F1b litter, and less than the control group for other litters (F2a, F3a, F3b).  These variations are not considered to show a treatment-related effect on fertility.  The duration of pregnancy, live birth index, litter size, litter weights, survival index, viability index and lactation index were all comparable between control and treated groups.  Organ weights were similar between control and treated groups.  There were no treatment-related histopathological findings.

Parameter

Control

High dose

Generation

M

F

M

F

Mortality

Incidence

P

0

0

0

0

F1

0

0

0

0

F2

0

0

0

0

F3

0

0

0

0

Food consumption

% of control

NA

NA

NA

NA

Body weight gain

% of control

100

100

100

100

Clinical Observations

specify effects

Incidence

NA

NA

NA

NA

Organ weights (F3b)

% of control

95 -103

95 -103

95 -103

95 -103

Pathology

Histopathologic examination: F3b

Immature testes

Renal cyst

Parasites in intestine

Focal pulmonary hemorrhage

Parasites in renal pelvis

Incidence

10

-

-

-

-

-

-

1

1

-

10

1

1

1

-

-

-

-

-

2

Reproductive Performance

Mating index: P

-

NA

NA

F1

-

NA

-

NA

F2

-

NA

-

NA

F3

-

NA

-

NA

Fertility index: P, la

-

87.5

-

93.8

F1b

-

75

-

75

F2a

-

82.3

-

68.8

F2b

-

76.5

-

81.3

F3a

81.3

-

68.8

F3b

80

-

62.5

Number of implantation sites

Mean

-

NA

-

NA

Duration of pregnancy: P, 1a

Mean days

24.7

24.06

F1b

25.1

-

25.25

F2a

23.36

-

24.91

F2b

25

-

24.4

F3a

25.8

-

25.1

F3b

25.2

-

25

Birth index

Live birth index: P, 1a

-

93.8

-

100

F1b

93.8

-

100

F2a

100

-

100

F2b

100

-

93.8

F3a

93.8

-

100

F3b

100

87.5

Gestation index

Litter size: P, 1a

Mean

-

8.9

-

9.2

F1b

11

-

13

F2a

9.2

-

9.5

F2b

10.7

-

10.2

F3a

10.4

-

10.2

F3b

8.4

-

10.5

Litter weight: P, 1a

Mean, day 21, g

-

307

-

283

F1b

348

-

392

F2a

279

-

341

F2b

379

-

325

F3a

270

349

F3b

319

-

388

Pup weight, 1a

Mean: day 21, g

36.8

-

37.8

F1b

42.2

-

40.5

F2a

42.9

-

42.1

F2b

40.3

-

41.5

F3a

41.4

-

43.7

F3b

45.6

-

41.9

Sex ratio

Male/female

NA

NA

Survival index: P, 1a

Day 5

91.4

-

95.6

F1b

91.6

-

97.4

F2a

93.3

-

88.6

F2b

96.7

-

93.4

F3a

82.4

-

85.2

F3b

74.8

-

98.8

Viability index: P, 1a

Day 21

-

86.2

-

81.1

F1b

75

-

74.3

F2a

74.2

-

84.8

F2b

85

-

77

F3a

72.8

-

72

F3b

76.2

-

88

Lactation index: P, 1a

Day 21

86.2

-

81.1

F1b

75

-

74.3

F2a

74.2

-

84.8

F2b

85

-

77

F3a

72.8

-

72

F3b

76.2

-

88

Sperm characterization

NA

NA

Number

% of control

NA

NA

Deformations

% of control

NA

NA

Conclusions:
The study is relevant to the hazard evaluation of STPP, but limited in data on the endpoints tested because only one dose was included. In addition, histopathological evaluation of all major reproductive organs, except testes and ovaries, was not conducted. Therefore, this study is considered to be Klimisch code 2: reliable with restrictions. Although the restrictions are significant, the investigation is well documented and scientifically acceptable. Reliable conclusions can be drawn from this study based on the data reported with consideration of the limitations. It can be concluded that STPP did not induce adverse effects on fertility and reproduction in rats when fed for three generations at a level of 0.5% in the diet. This no-effect level provides a basis for some risk assessment even though not all parameters currently included in the OECD 416 guideline were evaluated. Key Restrictions: Only one dose level was tested and important data on the reproductive organs were not collected; no evaluation of some sex organs, no sperm evaluations, individual pup data not evaluated, Specific deficiencies may be addressed by reviewing other studies on STPP and on similar inorganic phosphates. Together, the data on several phosphates indicate no major concerns regarding reproductive or fertility.
Endpoint:
three-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
Well documented study with the following deviations: Only one dose level was included in the study. Test material was not analyzed in diet. No impurity profile of test material was provided. Source of animals was not specified. The group size was smaller than the 20 females and 10 males required in OECD guideline 416. There was no recorded acclimation period for the P generation. Only one dose tested. Daily observation of clinical signs of parents and observations of abnormal behaviour of offspring are not reported. Body Weights: parental body weights were not recorded during gestation and lactation. Food consumption was not determined. Estrus cycle was not evaluated. Sperm parameters were not included. Offspring sex and sex ratios were not determined. Litter body weight was reported; however, individual pup body weights were not recorded. There is no report of gross pathology and necropsy observations. Organ weight data for the uterus, ovaries, epididymides, prostate, seminal vesicles, pituitary gland, adrenals and thyroid gland were not included. Histopathology was conducted only on the F3b offspring and did not include the following organs: uterus, cervix, vagina, epididymides, seminal vesicles, prostate, coagulating gland, pituitary gland. Note that the known target organ (kidney) was evaluated. Statistics are not reported. Study has been reviewed by an expert assessor (see expert report attached) and determined to be suitable for use for the purposes of REACH registration.
Qualifier:
no guideline followed
Principles of method if other than guideline:
Groups of male (8/group) and female (16/group) rats were fed control diet or diet treated with 0.05% test material for 100 days (P1) generation. Body weights were measured weekly during the 100 day pre-mating exposure for each generation. Each male was caged with two females for 7 days until conception occurred. Pregnancy rates were determined. The first litter born of the parental animals was designated as the F1a litter. Upon delivery, offspring were counted and average litter weights determined. Pup survival was determined on days 1-5 and days 6-21. On day 5, pups were culled to 10 per litter. Body weights per litter were determined on day 21. The F1a litter was sacrificed at 30 days postnatally. Parental animals were mated again 10 days after the sacrifice of the F1a litter (ten-day interim period). Mating took place for 7 days. A second litter (F1b) was born after approximately 21 days of gestation. Upon delivery, offspring were counted and the average litter weights determined. Pup survival was determined on days 1-5 and days 6-21. On day 5 pups were culled to 10 per litter. Body weights per litter were determined on day 21. Following weaning of the F1b litter, 8 males and 16 females from the F1b litter from the control group and the treated group were fed their respective diets for 100 days. Again two litters (F2a and F2b) were produced in the same sequence described for the F1 litters. Parental animals were mated for 7 days after the ten-day interim period after the sacrifice of the F2a litter (pups were 30 days old). Following birth of the F2b litter the same offspring and parental parameters were measured. Eight males and 16 females of the F2b litter were designated for continual exposure for another 100 days. Upon completion of the exposure of the F2b litter, animals were mated again to produce the F3a and F3b litters in the same manner described above. The ten males and ten females from the final litter, the F3b litter, were evaluated for body weight, histopathology (liver, kidneys, gonads, lungs, brain, stomach, heart, spleen, adrenal, stomach, large and small intestines, bladder, bone marrow, muscle, pancreas, lymph nodes, gut) and organ weights (liver, kidneys, testes, lungs, brain, stomach, heart, spleen). Individual animal data were collected and recorded throughout the study.
GLP compliance:
no
Remarks:
study predates GLP
Limit test:
yes
Species:
rat
Strain:
other: Rochester Strain (Ex-Wistar 1923)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: no data
- Age at study initiation: (P) x wks; (F1) x wks
- Weight at study initiation: Body weight was approximately 60 grams for males and 59 grams for females at the start of the 100 day exposure.
- Fasting period before study: no data
- Housing: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: not applicable


Route of administration:
oral: feed
Type of inhalation exposure (if applicable):
other: not applicable
Vehicle:
unchanged (no vehicle)
Details on exposure:
No data
Details on mating procedure:
- Length of cohabitation: 7 days
- Any other deviations from standard protocol: Litters were culled to ten pups on day 5 of lactation.
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
no data
Duration of treatment / exposure:
Exposure took place for 100 days prior to mating, 21 days of gestation and during lactation. Litters were culled to ten pups with approximately 5/sex/group on postnatal day 5. For the F1, F2, F3-generations, offspring were weaned and exposed during an interim period of 10 days and then during a mating period of 7 days. Dams were exposed continually during gestation and lactation. Offspring of the second litter were again mated for 7 days after a 10 day interim period following weaning. The first litters (F1a, F2a, F3a) were sacrificed at 30 days of age. The second litters (F1b, F2b) were used to produce the succeeding generation. The F3b pups were used for the histopathology evaluation and organ weight determinations.
Frequency of treatment:
daily, ad libitum
Details on study schedule:
Beginning with the original groups of rats, 16 female and 8 male rats were removed at the age of 100 days and mated. The matings were carried out in such a way that male rats were rotated through cages each housing two female rats.

For the F1, F2, F3-generations, offspring were weaned and exposed during an interim period of 10 days and then during a mating period of 7 days.
Offspring of the second litter were again mated for 7 days after a 10 day interim period following weaning.

The first litters (F1a, F2a, F3a) were sacrificed at 30 days of age. The second litters (F1b, F2b) were used to produce the succeeding generation. The F3b pups were used for the histopathology evaluation and organ weight determinations.

Litters were culled to ten pups with approximately 5/sex/group on postnatal day 5.
Dose / conc.:
0.05 other: % w/w nominal in diet
No. of animals per sex per dose:
P1, F1, F2, F3: 16 females and 8 males
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale:
The reproduction study began with 16 female and 8 male rats from the control diet group and from the group given the diet containing 1% of TMP in the 2-year study. When it was discovered that the male rats given a 1% Trimetaphosphate diet were not maintaining normal growth, it was obvious that this level was unsuitable one for reproduction study. Consequently additinoal diet groups were started at 0.1% and at 0.05% to be sure that no detectable effect could be laid to the TMP in the diet. The continued stress of repeated production of generations and litters of rats could then be tested. In 1957 it was decided to conduct a reproduction study on rats maintained on the diet containing 0.05% trimetaphosphate.

- Rationale for animal assignment (if not random): no data

- Other: no data
Positive control:
no data
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: no data
- The animals were monitored for general well-being.

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly during the 100 day pre-mating period, but not evaluated during gestation and lactation.
Oestrous cyclicity (parental animals):
Not evaluated.
Sperm parameters (parental animals):
The following were not evaluated: testis weight, epididymis weight, daily sperm production, sperm count in testes, sperm count in epididymides, enumeration of cauda epididymal sperm reserve, sperm motility, sperm morphology.
Litter observations:
number of pups, stillbirths, live births, weight gain per litter

The following was not evaluated: sex of pups, presence of gross anomalies, physical or behavioural abnormalities. No individual pup body weights.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals.
- Maternal animals: All surviving animals.

GROSS NECROPSY
- Gross necropsy consisted of [external and internal examinations including the cervical, thoracic, and abdominal viscera.]

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table [#] were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
ORGAN WEIGHTS; F3b:
liver, kidneys, testes, lungs, brain, stomach, heart, spleen on 10/sex/group from the F3b offspring at 21 days of age.
The following was not evaluated: Uterus, ovaries, epididymides (total and cauda), prostate, seminal vesicles, pituitary, thyroid, adrenal glands

HISTOPATHOLOGY; F3b:
Kidneys, heart, brain, spleen, lungs, liver, ovaries, testes, adrenal, urinary bladder, stomach, large and small intestines, muscle, bone marrow, muscle, pancreas, gut and lymph noted on F3b animals.
The following was not evaluated: Vagina, uterus, epididymis, seminal vesicle, prostate
coagulating gland.
Not evaluated
Key result
Dose descriptor:
NOAEL
Effect level:
0.05 other: % w/w
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects noted
Key result
Dose descriptor:
NOAEL
Effect level:
ca. 25 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects noted
Key result
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
0.05 other: % w/w
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects notes
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
ca. 25 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects noted
Remarks on result:
other: calculated
Key result
Critical effects observed:
no
Key result
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
0.05 other: % w/w
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: no effects noted
Key result
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
ca. 25 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects noted
Remarks on result:
other: calculated
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no

NOAEL calculated in accorance with Appendix F, guidelines for the preparation of toxicological working papers for the joint FAO/WHO expert committee on food additives, December 2000.

This study included only one dose level of the test material, 0.05% in the diet. Table 1 summarizes results on many parameters. There was no treatment-related effect on litter weights. The fertility index, duration of pregnancy, live birth index, litter size, litter weights, average pup weights, number of pups per litter, survival index, viability index and lactation index were all comparable between control and treated groups. Organ weights were similar between control and treated groups. Treated females of the F3b generation had somewhat lower spleen weights than control animals. There were no treatment-relatedhistopathologicalfindings.

 

Table 1. Reproductive toxicity

Parameter

 

Control

High dose

Generation

M

F

M

F

Mortality

Incidence

P

0

0

0

0

 

 

F1

3

1

0

0

 

 

F2

0

2

0

0

 

 

F3

0

1

0

1

Food consumption

% of control

 

NA

NA

NA

NA

Body weight gain

% of control

 

100

100

100

100

Clinical Observations

specify effects

Incidence

 

NA

NA

NA

NA

Organ weights (F3b)

Liver

Kidneys

Testes

Lungs

Brain

Stomach

Heart

Spleen

% of control

 

 

 

 

 

 

 

 

95.6

98.9

92.1

95.4

100.6

88.0

89.4

83.7

 

96.8

91.9

-

93.4

105.5

92.5

90.5

69.2

Pathology

 

 

 

 

 

 

Histopathologicexamination: F3b

Incidence

0

0

0

0

0

 
Conclusions:
The test material did not cause any adverse effects on fertility, reproductive performance, offspring viability, offspring survival and offspring body weight in male and female rats when administered continuously for three generations in the diet at a dose level of 0.05% in the diet. The test material did not cause any treatment-related effects on organ weights or any histopathological findings on the tissues evaluated at a dose level of 0.05% in the diet.
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
25 mg/kg bw/day
Study duration:
chronic
Species:
rat
Quality of whole database:
The dataset has been reviewed by an external assessor (M Weiner, TOXpertise, LLC) and it is concluded that the data is adequate and reliable for use as a key study under REACH. Please see expert report attached in endpoint under 'background information'.
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The above NOAEL is based on the highest concentration tested in the study.

The Hodge reproduction study (1960) was conducted prior to the institution of good laboratory practice guidelines and to the current OECD Guideline 416. Therefore, the study has deficiencies when examined according to today’s standards. The study is considered a Klimisch Code 2, reliable with restrictions (see report attached in study summary). The conclusions regarding the hazard identification of trisodium trimetaphosphate are supported by data on sodium tripolyphosphate / pentasodium triphosphate (STPP) and sodium hexametaphosphate (SHMP), both of which are structurally similar to trisodium trimetaphosphate. All three inorganic phosphate salts had no adverse effects on reproduction and fertility when tested in rats over three generations (Hodge 1959, 1960).

 

Based on this evidence trisodium trimetaphosphate is not considered to be classified as a reproductive toxicant according to Regulation (EC) 1272/2008 (EU CLP).


Short description of key information:
Regulation (EC) No. 1907/2006 - Annex VIII - Section 8.7.1. Screening for reproductive toxicity.
A Column 2 adaptation has been used to waive the need to conduct this study - Screening for reproductive toxicity does not need to be conducted if a study is available for a two-generation reproductive toxicity study (Annex IX, Section 8.7.3). A multi-generation reproductive toxicity study can be found in Section 7.8.1 of this dossier.

Regulation (EC) No. 1907/2006 - Annex IX - Section 8.7.3. Two generation reproductive toxicity study.
The rats were fed diets containing 0.05% trisodium trimetaphosphate during the 100 days prior to mating, during the 21 days of gestation and during the lactation for the P1, F1b, F2b. The F3b were exposed for their first 21days after which time they were sacrificed.
Observations were limited to body weight, histopathology (liver, kidneys, gonads, lungs, brain, stomach, heart, spleen, adrenal, stomach, large and small intestines, bladder, lymph nodes, gut, marrow, pancreas) and organ weights (liver, kidneys, testes, lungs, brain, stomach, heart, spleen).
The study was conducted before the introduction of GLP and the modern standard test Guideline OECD 416. This result provides evidence for the assumption that there is no concern with regard to effects of trisodium trimetaphosphate on reproduction.

Justification for selection of Effect on fertility via oral route:
The dataset has been reviewed by an external assessor (M Weiner, TOXpertise, LLC) and it is concluded that the data is adequate and reliable for use as a key study under REACH. Please see expert report attached in endpoint under 'background information'.

Effects on developmental toxicity

Description of key information
Regulation (EC) No. 1907/2006 - Annex VIII - Section 8.7.1. Screening for developmental toxicity.
A Column 2 adaptation has been used to waive the need to conduct this study - Screening for developmental toxicity does not need to be conducted if a study is available for pre-natal developmental toxicity (Annex IX 8.7.2). An Annex IX 8.7.2 study can be found in section 7.8.2 of this dossier.
Regulation (EC) No. 1907/2006 - Annex IX - Section 8.7.2. Prenatal developmental toxicity
No pre-natal developmental toxicity study is submitted on the basis that the substance is considered to be of low toxicological activity. It is therefore not considered to be ethically justified to conduct additional in vivo studies.
Effect on developmental toxicity: via oral route
Endpoint conclusion:
no study available

Justification for classification or non-classification

Trisodium trimetaphosphate is not considered to be a reproductive or developmental toxicant in accordance with Regulation (EC) No.1272/2008 (EU CLP).