Dizinc pyrophosphate

Endpoint summary

Description of key information

Oral (OECD 423, RL1), rat: LD50 cut-off = 5000 mg/kg bw

Inhalation (OECD 436, RL2), rat: LC50 > 4.73 mg/L

Dermal (OECD 402, RL1), guinea pigs: LD50 > 2000 mg/kg bw

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 Jan 2017-07 Feb 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Version / remarks:

Adopted 08 February 2002

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Landesamt für Gesundheit und Soziales, Mecklenburg-Vorpommern, Germany

Test type:

fixed dose procedure

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories GmbH, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 9 weeks (test group 1), 10 weeks (test group 2)
- Weight at study initiation: 204.7 - 242.0 g
- Fasting period before study: overnight prior to dosing
- Housing: group housed by dose, not further specified
- Diet: conventional laboratory diet pellets, ssniff Spezialdiaten GmbH, Soest, Germany, ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30-70
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 17 January 2017 To: 07 February 2017

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

distilled

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 200 mg/mL
- Amount of vehicle (if gavage): 1 mL/100 g bw
- Justification for choice of vehicle: suspension was formed in the vehicle

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: Information about toxicity provided by the Sponsor indicated that the test material was likely non-toxic at this dose level.

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

6

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed individually after dosing at least once during the first 30 minutes, periodically during the first 24 hours, with special attention given during the first 4 hours and daily thereafter for a total of 14 days. Individual weights of animals were determined shortly before the test substance was administered, and weekly thereafter.
- Necropsy of survivors performed: yes

Statistics:

BW changes and LD50 values were calculated.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: According to this study, an LD50 cut-off of 5000 mg/kg was used.

Mortality:

No mortality occurred during the study period.

Clinical signs:

No clinical signs of toxicity were observed at dosing or up to the end of the 14-day observation period.

Body weight:

No animals showed any adverse effect on body weight gain.

Gross pathology:

Necropsy examination revealed no substance-related or abnormal findings.

Interpretation of results:

other: CLP/EU GHS criteria are not met, no classification required according to Regulation (EC) No 1272/2008.

Conclusions:

The LD50 of dizinc pyrophosphate is >2000 mg/kg bw by the oral route of exposure in female Wistar rats, and therefore the material is not classified.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 Jan 2017-01 Feb 2017

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Version / remarks:

Adopted 7 Sep 2009

Deviations:

yes

Remarks:

2 males and 4 females were dosed. GSD out of range. Relative humidity of test material out of range. Temperature/RH in animal room sporadically out of range. No impact on the study results.

GLP compliance:

yes (incl. QA statement)

Remarks:

Entitad Nacional de Acreditaciόn, Madrid, Spain

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Specific Pathogen Free

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, C/Argenters 7, Local AB, 08290 Cerdanyola del Vallés, Barcelona, Spain
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 8 weeks
- Weight at study initiation: 261.9-273.5 g (males) and 196.0-217.5 g (females)
- Fasting period before study: not specified
- Housing: group housed 3-4 per cage with Capsumlab Lecho_10 (autoclavable) bedding, and enrichment devices (nesting material, tubes, and chew blocks)
- Diet: Global diet 2914C, Harlan Teklad, Station Road, Blackthorn, Bicester, Oxon, OX25 1TP, UK, ad libitum
- Water: tap water in water bottles, ad libitum
- Acclimation period: at least 7 days, with 90 min to the nose-only restraining tubes on the day of exposure

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 ± 1.5
- Humidity (%): 20-43
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 17 Jan 2017 To: 01 Feb 2017

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

clean air

Mass median aerodynamic diameter (MMAD):

ca. 2.37 µm

Geometric standard deviation (GSD):

ca. 6.71

Remark on MMAD/GSD:

MMAD during exposure was 2.37 μm (average of three particle size determinations). This value is within the respirable range (1-4 μm) and appropriate for acute inhalation toxicity testing. GSD was above the upper limit of 3 in the second and third particle size determinations, indicating that the particle size distribution of the aerosol generated throughout the 4-hour exposure period was not homogeneous. Nevertheless, at the highest concentration of test material technically achievable (4.73 mg/L, gravimetric determination) the mean percentage of particles below the upper limit of 4 μm, over the whole exposure period, was 66% (79%, 67% and 52.5% for particle size determinations #1, 2 and 3, respectively), and this value was considered acceptable.

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: flow-past, nose-only exposure chamber made of anodised aluminium
- Exposure chamber volume: approximately 3 L
- Method of holding animals in test chamber: rats were individually exposed in glass tubes matching their size, positioned radially around the exposure chamber
- Source and rate of air: clean air with a flow rate at each exposure tube of approximately 1 L/min
- Method of conditioning air: filtered air from a compressor
- System of generating particulates/aerosols: a dust aerosol was generated from the desiccated test material using a rotating brush aerosol generator (RBG 2000, PALAS GmbH, Germany) and conveyed via glass tubing from the generator to the exposure chamber; aerosol concentration was determined by gravimetric analysis.
- Method of particle size determination: MMAD and the GSD were determined on results from a PIXE cascade impactor, using Microsoft Excel® software (Microsoft Corporation, USA).
- Treatment of exhaust air: not specified
- Temperature, humidity, pressure in air chamber: Temperature and relative humidity in the chamber were measured continuously during exposure using a thermohygrometer (Kimoth110, Kimo). The target range for temperature was 19-25 °C, the target range for relative humidity (RH) was 30-70%. The exposure airflow rate was adjusted using the pressure difference over a Venturi tube. Target flow range was 0.5-1.0 L/min through each inhalation tube.

TEST ATMOSPHERE
- Brief description of analytical method used: Three gravimetric measurements were taken during exposure to measure exposure concentration. The sampling flow was similar to the air flow rate per exposure port. Test aerosol samples were collected for 2 min periods onto a Whatman filter (grade F319-04) using a filter sampling device. The filters were weighed before and immediately after sampling using a calibrated balance. The gravimetric aerosol concentration was calculated from the amount of test material present on the filter and the sample volume.
- Samples taken from breathing zone: yes

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution: Determined gravimetrically three times during the exposure. The cumulative particle size distribution of the test aerosol was determined using a PIXE cascade impactor, measured by gravimetric analysis of the test material deposited on each stage of the cascade impactor.
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): MMAD and GSD were calculated on the basis of results from the impactor using Microsoft Excel® software (Microsoft Corporation, USA). Target ranges were 1 to 4 µm for the MMAD, and 1.5 to 3 for the GSD.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting concentration: Exposure concentration was the highest technically achievable concentration (4.73 mg/L)

Analytical verification of test atmosphere concentrations:

yes

Remarks:

gravimetric

Duration of exposure:

ca. 4 h

Concentrations:

5 mg/L (nominal)
4.73 mg/L (analytical)

No. of animals per sex per dose:

2 males and 4 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: hourly during exposure (only grossly abnormal signs), immediately and 1 h after exposure, and once daily thereafter until the end of the observation period. All animals were observed for a period of 14 days after administration. All animals were weighed on the day of treatment just before starting exposure (study day 1), on study days 2, 4, 8 and immediately before sacrifice on study day 15.
- Necropsy of survivors performed: yes
- Other examinations performed: histopathology

Statistics:

No statistical analysis was required.

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 4.73 mg/L air (analytical)

Based on:

test mat.

Remarks:

gravimetric aerosol concentration

Exp. duration:

4 h

Mortality:

No mortality occurred during the study period.

Clinical signs:

other: Chromorrhinorrhea and breathing difficulty was observed in all six animals immediately following exposure and five of six at one hour thereafter. Chromodacryorrhea was observed in five of six animals immediately following exposure and at one hour thereaf

Body weight:

A decrease in mean body weight of approximately 9.5% in males and 1% in females was observed between study day 1 (exposure) and study day 2. From study day 2 to the end of the observation period (day 15), body weight increased gradually in all animals.

Gross pathology:

Red lungs with red spots were observed in one male and red spots were also present in the lungs of two females. These findings were considered to be related to test item exposure. No macroscopic changes were observed in any of the rest of the organs/tissues.

Interpretation of results:

other: CLP/EU GHS criteria are not met, no classification required according to Regulation (EC) No. 1272/2008.

Conclusions:

The LC50 of dizinc pyrophosphate was greater than 4.73 mg/L air (gravimetric aerosol concentration), and based on the GHS classification criteria, can be considered unclassified.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 Dec 2016-07 Feb 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

Adopted 1987

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Landesamt für Gesundheit und Soziales, Mecklenburg-Vorpommern, Germany

Test type:

standard acute method

Limit test:

yes

Species:

guinea pig

Strain:

Dunkin-Hartley

Remarks:

Crl:(HA)BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: not specified
- Weight at study initiation: 350-450 g
- Fasting period before study: not specified
- Housing: group-caged by sex, not further specified
- Diet: commercial feeding mixture, Mühle Knull, Rostock, Germany, ad libitum
- Water: tap water, supplemented with vitamin C, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30-70
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 28 Dec 2016 To: 07 Feb 2017

Type of coverage:

semiocclusive

Vehicle:

water

Details on dermal exposure:

TEST SITE
- Area of exposure: dorsal area of the trunk
- % coverage: 10
- Type of wrap if used: gauze dressing held in place with tape

REMOVAL OF TEST SUBSTANCE
- Washing: residual test material was removed with water
- Time after start of exposure: 24 h

TEST MATERIAL
- Amount(s) applied: 2000 mg/kg bw
- For solids, paste formed: yes, the test material was moistened with the smallest amount of water sufficient to ensure good skin contact (500 µL) and applied as a paste

Duration of exposure:

24 h

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: animals were observed individually before and immediately (Time 0), 5, 15, 30 and 60 min, 3, 6 and 24 hours after application, then daily for 14 days. Individual body weights were determined before test material application and weekly thereafter.
- Necropsy of survivors performed: yes

Statistics:

BW changes and LD50 values were calculated.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred during the study period.

Clinical signs:

No clinical signs of toxicity were observed at application or up to the end of the 14-day observation period.

Body weight:

One male (1/5) animal showed a slight weight loss after 7 days, but this was considered not relevant. All other animals showed no adverse effect on body weight gain.

Gross pathology:

Necropsy examination revealed no substance-related or abnormal findings.

Other findings:

Examinations of the skin for oedema and erythema were performed at 1, 24, 48 and 72 h and day 14.
The test material caused very slight to moderate irritation (erythema) of the skin in 3/5 of the female animals, with numerical grading of 1-3 after exposure to the test material for 24 hours. Within 14 days after application of the test material, no skin irritation was observed.
The test material caused very slight irritation (erythema) of the skin in 2/5 of the male animals, with a numerical grading of 1 after exposure to the test material for 24 hours. Within 14 days after application of the test material, no skin irritation was observed.

Since no mortality or clinical signs were noted only the skin reactions are presented below:

Skin Reactions -- Numerical Grading and Duration

Skin reaction grades observed
Animal No.	1 h	24 h	48 h	72 h	14 d
1F	0	0	0	0	0
2F	0	0	0	0	0
3F	2**	3***	2**	1*	0
4F	1*	1*	0	0	0
5F	2**	2**	1*	0	0
1M	0	0	0	0	0
2M	0	0	0	0	0
3M	0	0	0	0	0
4M	1*	1*	0	0	0
5M	1*	1*	1*	0	0

* = very slight erythema

** = well-defined erythema

*** = moderate erythema

F = female; M = male

Interpretation of results:

other: CLP/EU GHS criteria are not met, no classification required according to Regulation (EC) No 1272/2008.

Conclusions:

The LD50 of dizinc pyrophosphate is >2000 mg/kg bw by the dermal route of exposure in male and female guinea pigs, and therefore the material is not classified.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Additional information

The acute oral toxicity of the test substance was investigated in female Wistar rats using a stepwise procedure according to OECD guideline 423 (Bioserv, 2017). In the first step, a group of 3 animals was given the test substance by oral gavage at a limit dose level of 2000 mg/kg bw (test group 1). No mortality occurred and animals did not show any adverse clinical signs (including body weight gain) or abnormal macroscopic findings of organs after a 14-day observation period. Therefore, in the second step, the testing procedure was repeated in a further group of 3 animals at the same dose level (test group 2). All animals of the second step survived and showed no adverse clinical signs, effects on body weight gain or abnormal findings at gross necropsy. Thus, the LD50 cut-off after oral exposure for dizinc pyrophosphate is 5000 mg/kg bw.

The acute inhalation toxicity of the test substance was investigated in two male and four female Sprague Dawley rats using a acute toxic class method according to OECD guideline 436 (Vivotecnia, 2017). The rats were exposed by nose-only, flow-past inhalation to dizinc pyrophosphate at a mean concentration of 4.73 mg/L air during 4 hours. This concentration was found to be the highest technically achievable. The ranges of aerosol concentration, temperature, relative humidity and air flow rate were considered satisfactory for a study of this type. In addition, the test substance was considered to be respirable to rats since the mean mass median aerodynamic diameter (MMAD) was 2.37 µm which is below 4 µm. Geometric Standard Deviation (GSD) on particle size distribution (PSD) #1 was 2.53 and therefore within the target range (1.5 - 3). However, in PSD #2 and PSD #3 GSD was above the upper limit of 3, indicating that the particle size distribution of the aerosol generated throughout the 4-hour exposure period was not homogeneous. Nevertheless, at the highest concentration of test item technically achievable (4.73 mg/L, gravimetric determination) the mean percentage of particles below the upper limit of 4 μm over the whole exposure period was 66% (79%, 67% and 52.5% for PSD #1, 2 and 3, respectively) and this value was considered acceptable.

No mortality was recorded during the study period. The main clinical signs present after exposure were chromorrhinorrhea, chromodacryorrhea, breathing difficulty, piloerection, dirty/wet fur and reduced mobility/prostration. All these signs were transient and most of them were not present the day after exposure. From study Day 4 to the end of the 14 day observation period no clinical signs were recorded and the animals exhibited a normal behaviour. A decrease in mean body weight of approximately 9.5% in males and 1% in females was observed between study Day 1 (exposure) and study Day 2. From study Day 2 until the end of the observation period (Day 15), body weight increased gradually in all animals, with mean body weight gains of approximately 30% and 14.5% for males and females, respectively, being recorded. Upon terminal necropsy, red lungs with red spots were observed in one male and red spots were also present in the lungs of two females. These findings were considered to be related to test item exposure.

It can be concluded that, under the present experimental conditions the LC50 for dizinc pyrophosphate was greater than 4.73 mg/L air (gravimetric aerosol concentration).

The acute dermal toxicity of the test substance was investigated in 5 female and 5 male guinea pigs using the limit test at dose level 2000 mg/kg body weight test according to OECD guideline 402 (Bioserv, 2017). All animals survived and showed no adverse clinical signs, effects on body weight gain or abnormal findings at gross necropsy. Additional to theses observations the examination of the skin for oedema and erythema at 1, 24, 48 and 72 h as well as at day 14 was performed. The test substance caused slight - moderate irritation of the skin of three female animals with numeral grading from 1 up to 3 after the application time of 24 hours. The test material caused very slight irritation of the skin of two male animals with numeral grading of 1 after the application time of 24 hours. 14 days after the application of the test material no irritation of the skin was observed. Based on this study, no final conclusion can be made regarding the skin irritation potential of dizinc pyrophosphate. Regarding acute dermal toxicity, a LD50 > 2000 mg/kg bw can be concluded.

Justification for classification or non-classification

The available data on acute oral, inhalation and dermal toxicity do not meet the criteria for classification according to Regulation (EC) 1272/2008, and are therefore conclusive but not sufficient for classification.