Dipotassium disulphite

Endpoint summary

Description of key information

A 3-generation chronic toxicity study (Til et al. 1972a) with treatment of rats (20 per sex and group) with dose levels of 0, 0.125, 0.25, 0.5, 1.0 or 2.0% Na2S2O5 administered in a 50 ppm thiamine-containing diet was identified as most reliable study for repeated dose toxicity and was defined as key study (read-across info, see `discussion`). Based on the read across concept for the group of sulfite substances, this result is also applicable to dipotassium disulfite..

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

repeated dose toxicity: oral, other

Remarks:

combined repeated dose and reproduction toxicity

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

not specified

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Remarks:

Very well-documented study which allows derivation of a NOAEL value for chronic toxicity. Most reliable study for NOAEL deriviation for repeated dose toxicity effects of sodium metabisulfite.

Justification for type of information:

see attachment “Read-across concept – Human Health/Environment - Category approach for Inorganic sulfites/thiosulfates/dithionite" in section 13.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

no guideline followed

Principles of method if other than guideline:

In a three-generation feeding study, groups of 20 male and 20 female Wistar rats received 0, 0.125, 0.25, 0.5, 1.0 and 2.0% sodium metabisulphite, i.e. 49, 108, 220, 460, and 955 mg/kg bw/d as actual dose in a thiamine-containing diet over periods of 2 years.

GLP compliance:

not specified

Limit test:

no

Specific details on test material used for the study:

not specified

Species:

rat

Strain:

Wistar

Details on species / strain selection:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: derived from the Institute's Wistar-derived colony
- Age at study initiation: weanling or young rats
- Housing: housed in groups of 5 in screen-bottomed cages
- Diet (ad libitum): basal diet was Institute's stock diet, with following percentage composition: 35% ground yellow maize, 26 % ground whole wheat, 10% soyabean-oil meal, 8% fish meal, 4% meat scraps, 2.7% dried whey, 3% brewer's yeast, 3.3% vitamin preparations, 1.5% minerals, 0.5% NaCl, 3% soyabean-oil, 3% grass meal.
- Water (ad libitum): tap water

ENVIRONMENTAL CONDITIONS
- Temperature: 24 - 26 °C

Route of administration:

oral: feed

Details on route of administration:

In view of the widespread use of sulphite in foods and drinks it was considered desirable to re-examine the toxicity of sulphite administered in the diet.

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): diets were freshly prepared every 2 weeks.
- Sulphite was added by mechanical mixing of Na2S2O5 at levels varying from 0.125 to 8%.
- In order to compensate for the destruction of thiamine by sulphite, the stock diet was drastically enriched with 50 ppm thiamine.
- Storage temperature of food: diets were stored at -18°C and the rats were provided daily with a fresh portion of previously frozen diet.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Diets were analysed for SO2 and thiamine.

Results:
- SO2 determinations on the diets showed considerable losses of sulphite.
- proportionally the losses of sulphite decreased with increasing dietary levels of sulphite both immediately after mixing and after storage for 2 wk at - 18 °C and subsequent storage for 24 hr at room temperature.
- thiamine content decreased after storage for 2 wk at -18'C only in the 1 and 2% sulphite diets. Subsequent storage for 24 hr at room temperature showed slight and similar losses at 0, 0.125 and 0.25 %, but at higher sulphite levels there was a definite tendency towards greater losses with an increase of sulphite in the diet.
- during the experimental period, the rats consumed food that was stored on average for 1 week at -18°C and subsequently kept in the feeders for 12 hr at 24°C. The losses amounted to 22, 14, 12, 8 and 4.5 % Na2S20s and 2.7, 1.7, 8.3, 14.5 and 15.4% thiamine in the diets supplemented with 0.125, 0.25, 0.5, 1 and 2 % sulphite, respectively.

Duration of treatment / exposure:

104 weeks (21 weeks or 34 weeks before mating and up to a total of 104 weeks)

Frequency of treatment:

continuously

Dose / conc.:

0.125 other: %

Remarks:

ca. 50 mg/kg bw (nominal in diet)

Dose / conc.:

0.25 other: %

Remarks:

ca. 110 mg/kg bw (nominal in diet)

Dose / conc.:

0.5 other: %

Remarks:

ca. 220 mg/kg bw (nominal in diet)

Dose / conc.:

1 other: %

Remarks:

ca. 460 mg/kg bw (nominal in diet)

Dose / conc.:

2 other: %

Remarks:

ca. 960 mg/kg bw (nominal in diet)

No. of animals per sex per dose:

20 males / 20 females

Control animals:

yes, plain diet

Details on study design:

not specified

Positive control:

not specified

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Cage side observations checked: clinical signs and mortality
At week 32, 64 and 100 all rats were examined for occult blood in the faeces.

DETAILED CLINICAL OBSERVATIONS: Not specified

BODY WEIGHT: Yes
- Time schedule for examinations: weekly for the first 12 weeks and once every four week thereafter.

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Time schedule for examinations of food consumption: at intervals during 1-week periods.

FOOD EFFICIENCY: Not specified
WATER CONSUMPTION: Not specified
OPHTHALMOSCOPIC EXAMINATION: Not specified

HAEMATOLOGY: Yes
- Time schedule for collection of blood: week 52, 78, and 100
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Not specified
- How many animals: 6 – 10 animals of each sex from the control, 1 and 2 % groups
- Parameters checked: haemoglobin concentration, haematocrit value, numbers of erythrocytes, total and individual types of leukocytes

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: week 52 and 104.
- Animals fasted: Not specified
- How many animals: not specified
- Parameters checked: glutamic-oxalacetic transaminases and glutamic-pyruvic transaminases.

URINALYSIS: Yes
- Time schedule for collection of urine: week 13, 28, 52, 78, and 101
- Metabolism cages used for collection of urine: Not specified
- Animals fasted: Not specified
- How many animals: 5-11 rats of each sex, usually from the controls and the 1 and 2 % groups
- Parameters checked: phenol-red excretion. glutamic-oxalacetic transaminases, specific gravity, appearance, pH, glucose, albumin, occult blood, ketones, and microscopy of the sediment.

NEUROBEHAVIOURAL EXAMINATION: Not specified

Sacrifice and pathology:

SACRIFICE: Yes
- After 52 weeks, 5 males and 5 females of each diet group were killed for interim organ weights analysis and gross pathology.
- At week 104 all survivors were killed.

GROSS PATHOLOGY: Yes
- At week 104 all survivors were killed and the heart, kidneys, liver, spleen, brain, testes, ovaries, pituitary, thyroid and adrenals were weighed as well as fixed.
- Rats that died or were killed when moribund were also autopsied, but tissue samples were preserved only if autolysis was not too advanced.

HISTOPATHOLOGY: Yes
- Tissue samples were fixed in 10% buffered formalin, embedded in paraffin, sectioned at 5 µm and stained with haematoxylin and eosin.
- The following tissues were examined: heart, kidneys, liver, spleen, brain (three levels), testes, ovaries, pituitary, thyroids, parathyroids, adrenals, thymus, lungs, trachea, salivary glands, gastro-intestinal tract (six levels), pancreas, urinary bladder, skeletal muscle, spinal cord, femoral nerve, skin, bone marrow (sternum), axillary and mesenteric lymph nodes, exorbital lachrymal gland, aorta, mammary glands, uterus, prostate, seminal vesicle and coagulating gland.

Other examinations:

- Thiamine was determined in the pooled urine samples of 5-10 male and 5 - 10 female rats of each group at several stages and also in pooled liver samples of five rats of each sex at week 52 and 104.

Statistics:

not specified

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

- All rats in the highest dose group showed indications of occult blood in the faeces, while this effect occurred in only 21-60% of the animals on diets containing 1 % sulphite.
- In 10 % of the females given 0.25 % and in 10 % of the males given 0.5 % sulphite slight indications of intestinal blood loss were observed at wk 32 only.

Mortality:

mortality observed, non-treatment-related

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

- Pathological changes attributable to the feeding of sulphite were observed only in the stomach.
- A distinctly raised and thickened limiting ridge and small amounts of a reddish-brown flocky material in the mucus layer of the glandular stomach were seen grossly in the groups given the two highest sulphite levels.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

- Hyperplastic changes were seen in both the forestomach and glandular stomach at the 1.0 and 2.0% sulphite level. They were either present only at the 1 and 2 % dosage levels or their incidence was distinctly increased at these levels.

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
- General conditions of the rats remained good during the first 72 weeks.
- After this time, aging symptoms developed in many rats and mortality increased rapidly.
- The survival in the sulphite groups was generally higher than in the controls.

BODY WEIGHT AND WEIGHT GAIN
- Body weights were comparable irrespective of treatment.

FOOD CONSUMPTION AND COMPOUND INTAKE
- There were no distinct differences in food consumption.

HAEMATOLOGY
- Haemoglobin, haematocrit values and erythrocyte counts were marginally reduced at 2% sulphite at week 52, 78, and 100.

CLINICAL CHEMISTRY
- Significant (P < 0.05) decreases in serum glutamic-pyruvic-transaminase values occured at wk 104 in male rats receiving 0.125 % sulfite.
- Otherwise, there were no differences between the test and control animals in transaminase activities either at week 52 or at week 104.

URINALYSIS
- Kidney function: phenol-red excretion, specific gravity and glutamic-oxalacetic-transaminase activity in the urine were not adversely affected.
- Urine analysis values were essentially normal.

ORGAN WEIGHTS
- Interim results obtained after 1 year did not indicate any effect of sulphite on organ to body weight ratios.
- Terminally the relative weights of the livers were lower in all the test groups than in the controls, but there was no evidence of a dose-related response.

HISTOPATHOLOGY: NON-NEOPLASTIC
- The type and incidence of the other non-neoplastic lesions observed were comparable in test and control groups.

HISTOPATHOLOGY: NEOPLASTIC
- The number of lymphoreticular pulmonary tumours in males decreased with increasing levels of sulphite in the diet.
- The incidence of thyroid and pituitary tumours in the control group of the males was exceptionally low, whereas those noted in the various test groups represented numbers normally found in the strain used.
- All other neoplasms occurred in a random manner with no apparent relationsship between number, location of tumours and treatment.

OTHER FINDINGS
- There was a dose-related decrease in the thiamine content of the urine at sulphite levels of 0.125 % and above and in the liver at levels above 0.25 %
- The group given 2 % sulphite showed no distinct changes in the thiamine status when compared with control rats on the stock diet without extra thiamine.

Key result

Dose descriptor:

NOAEL

Remarks:

local effects

Effect level:

108 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see "Remarks"

Key result

Dose descriptor:

NOAEL

Remarks:

systemic effects

Effect level:

> 955 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No signs of systemic toxicity were observed and the NOAEL can be expected above the highest dose of 2% metabisulphite corresponding to 955 mg/kg bw/d Na2S2O5 (or 640 mg/kg bw/d as SO2 equivalents).

Critical effects observed:

not specified

Calculation of the dose level in mg/kg bw/d was done by converting % diet into ppm and by assuming an average body weight for older rats of 400 g and a daily feed intake of 20 g as follows:

0.215 % = 2150 ppm x 0.05 = 108 mg/kg bw/d Na2S2O5, equivalent to 72 mg/kg bw/d SO2

1.91% = 19100 ppm x 0.05 = 955 mg/kg bw/dNa2S2O5, equivalent to 640 mg/kg bw/d SO2

Conclusions:

Based on the occurrence of occult blood in the faeces and changes in gastric morphology at dose levels of 0.5% or more, the NOAEL for local effects in this study is represented by the dose of 0.25% metabisulphite (or 0.215% accounting for the loss of metabisulphite). The corrected dose level corresponded to a dose of 108 mg/kg bw/d Na2S2O5 or an equivalent dose of 72 mg SO2/kg bw/day. Because there were no evidence of systemic toxicity following chronic treatment, the NOAEL for systemic effects can be expected above the highest dose of 2% metabisulphite corresponding to 955 mg/kg bw/d of Na2S2O5 (or 640 mg/kg bw/d as SO2 equivalents).

Endpoint conclusion

Dose descriptor:

NOAEL

126 mg/kg bw/day

Study duration:

chronic

Species:

rat

Additional information

Read-across concept for sulfites, hydrogensulfites, metabisulfites, dithionites and thiosulfates:

A comprehensive read-across concept has been developed for sulfites, hydrogensulfites and metabisulfites, based on the pH-dependant equilibrium in aqueous solutions which is summarised in the following equations:^[1],[2]

           SO₂+ H₂O <->`H₂SO₃´         H₂SO₃<->H⁺+ HSO₃^-<->2H⁺+SO₃^2-    2HSO₃^-<->H₂O +S₂O₅^2-

Since the nature of the cation (i.e., sodium, potassium, ammonium…) is not assumed to contribute substantially to differences in toxicity and solubility (all compounds are very soluble in water), only the chemical and biological properties of the anion are considered as relevant determinants. Based on the described equilibrium correlations, unrestricted read-across between the groups of sulfites, hydrogensulfites and metabisulfites is considered justified.

 

Additionally, it is known that sodium dithionite disproportionates in water to form sodium hydrogen sulfite and sodium thiosulfate (equation II)^[2],[1], so that this substance can also be considered to be covered by the read-across concept described above. Since it can easily be anticipated that the substance is not stable enough under physiological conditions to fulfil the requirements of study guidelines, instead the products of decomposition have to be considered:

 

       2 S₂O₄^2-+ H₂O→2HSO₃^-+ S₂O₃^2-

 

Not fully covered by this read-across concept is the substance class of thiosulfates: although the thiosulfates are also well known to disproportionate in aqueous solution to form polythionic acids and SO₂(HSO₃^-), this requires somewhat different, more acidic conditions. Therefore, read-across to sulfites is primarily restricted to appropriate physiological conditions, i.e. oral administration where the gastric passage with the strongly acidic conditions in the stomach will facilitate the chemical disproportionation described above:

 

       HS₂O₃^-+ H₂S₂O₃→HS₃O₃- + SO₂+ H₂O

[1]Hollemann Wiberg, Lehrbuch der Anorganischen Chemie, 101.Auflage

[2]Handbook of Chemistry and Physics, Ed. Lide, DR, 88^thedition, CRC Press

Oral toxicity:

Male and female rats received 0, 0.125, 0.25, 0.5, 1.0 or 2.0% Na₂S₂O₅in a thiamine-containing diet (50 ppm) for 104 weeks. Based on the occurrence of occult blood in faeces and changes in gastric morphology at dose levels of 0.5% or more, the NOAEL for local chronic toxicity in this study is represented by the dose of 0.25% metabisulfite (or 0.215% accounting for the loss of metabisulfite). The corrected dose level corresponded to a dose of 108 mg/kg bw/d Na₂S₂O₅or an equivalent dose of 126 mg/kg bw/day potassium metabisulfite. Because there was no evidence of systemic toxicity following chronic treatment, the NOAEL for systemic effects can be expected above the highest dose of 2% sodium metabisulfite corresponding to 955 mg/kg bw/d of Na₂S₂O₅or 1117 mg/kg bw/d potassium sulfite.

Repeated dose toxicity, dermal:

According to regulation (EC) 1907/2006 Annex XI (weight of evidence), testing for sub-chronic dermal toxicity is not considered to be required, for the following reason: repeated dose toxicity study via dermal route does not need to be performed since the physico-chemical and toxicological properties do not suggest potential for a significant rate of absorption through the skin (for more information, refer to the section on toxicokinetics).

Repeated dose toxicity, inhalation:

According to regulation (EC) 1907/2006 Annex XI (weight of evidence), testing for sub-chronic inhalation toxicity is not considered to be required, for the following reasons: iIn accordance with ECHA guidance on information requirements and chemical safety assessment-chapter R.8: characterisation of dose [concentration]-response for human health, May 2008, a DNEL for systemic effects can also be derived by route-to-route extrapolation from a 90-day oral toxicity study in rats. Thus, the calculated NOAEL of 1117 mg/kg bw/d potassium metabisulfite from the chronic oral toxicity study with sodium metabisulfite was used as the starting point for DNEL derivation.

Inconclusion, it is neither scientifically nor justified due to animal welfare reasons to initiate further inhalation toxicity studies.

Repeated dose toxicity: via oral route - systemic effects (target organ) digestive: stomach

Justification for classification or non-classification

Repeated dose toxicity, oral

Local effects in the stomach were the most predominant finding of repeated dose toxicity: The NOAEL for local chronic effects in the study described by Til et al. (1972) is represented by the dose of 0.25% metabisulfite. The corrected dose level corresponded to a dose of 108 mg/kg bw/d Na₂S₂O₅. All observed effects (occurrence of occult blood in faeces and changes in gastric morphology) were detected at higher dose levels at and above 0.5% in the diet (220 mg/kg bw/d Na₂S₂O₅). There was no evidence of systemic toxicity following chronic treatment with sodium metabisulfite. Therefore, the NOAEL for systemic effects can be expected above the highest dose of 2% metabisulfite in the diet corresponding to 955 mg/kg bw/d of Na₂S₂O₅.

 

Therefore, it can be concluded that based on the available animal data, Na₂S₂O₅does not have the potential to produce significant toxicity, or to be harmful to humans, following repeated exposure at low or moderate exposure concentrations relevant for classification. This result can be read across to dipotassium disulfite without restriction.Thus, the requirements for STOT-RE classification criteria according to regulation (EC) 1272/2008 as specific target organ toxicant (STOT) – repeated exposure, oral are not met, and no classification as specific target organ toxicant is required.

 

Repeated dose toxicity, dermal

(i) Based on physico-chemical properties of dipotassium disulfite and the toxicokinetic behaviour (very limited penetration into the upper epithelial layers of the epidermis,) there are no systemic risks to humans with respect to dermal exposure to this substance.

(ii) One may assume a conservative default of 1% for dermal absorption of dipotassium disulfite, leading to the anticipation of a negligible toxicity via the dermal route.

 

Thus, it may be concluded that there will be no systemic risks to humans with respect to dermal exposure to dipotassium disulfite, and no classification for specific target organ toxicant (STOT) – repeated exposure, dermal according to regulation (EC) 1272/2008 is required for this substance or any suitable compound for read-across to this compound.

 

 

Repeated dose toxicity, inhalation

According to regulation (EC) 1272/2008, a classification for specific target organ toxicity – repeated exposure shall be taken into account only when reliable evidence associating repeated exposure to the substance with a consistent and identifiable toxic effect demonstrates support for the classification. However, there is no data or information available to evaluate a potential for specific organ toxicity following repeated inhalation exposure, and thus classification is not required.