tetraammine palladium (II) hydrogen carbonate

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, available as an unpublished report, no restrictions, fully adequate for assessment

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

At test start and at each renewal samples of all treatments were taken from the test solution preparation just before distributing it to the 10 replicates.At the renewals samples of the aged test solution were taken from each vessel, pooled per concentration and measured. Since analysis could not be applied immediately after sampling, samples were stored stabilized 1:1 (v/v) with ammonia solution 10 % in a refrigerator at 4 – 8 °C until analysis.

Vehicle:

no

Details on test solutions:

Stock solutions were prepared by dissolving adequate amounts of test item in dilution water and these were used to prepare test solutions. The stock solutions were freshly prepared before each renewal.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain: Clone V
- Source: German Federal Environment Agency, Location Berlin-Marienfelde, Ecotoxicology Lab, 12307 Berlin, Germany
- Age at study initiation: 4-24 hours old

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

1 m mol/L as CaCO3

Test temperature:

Temperature: 19.5 – 20.9 °C throughout the test, with the permitted range of 18 – 22 °C (with a variance of less than 2 °C)

pH:

pH: 7.9 – 9.4 throughout the test. Usually the pH should be between 6 and 9. In weeks 1 and 2 the pH in aged solutions was > 9 due to algae. This is because the daphnids were fed at a constant rate but the feeding rate was lower for the juveniles. The pH did not vary by more than 1.5 units during the study and all other validity criteria were met so this was not considered to effect the validity of the test.
test.

Dissolved oxygen:

Oxygen: 8.4 mg/L and 11.8 mg/L throughout the test which is above the required 3 mg/L

Nominal and measured concentrations:

Definitive test: nominal concentrations: Control, 0.051, 0.163, 0.522, 1.67, 5.35, and 17.1 µg Pd/L,
corresponding to Control, 0.101, 0.324, 1.04, 3.32, 10.63, and 34 µg DDP/L.
Mean measured (time weighted average) concentrations: control, 0.025, 0.062, 0.275, 1.16, 4.05, and 14.3 µg Pd/L,
corresponding to 0.049, 0.124,0.546, 2.31, 8.04 and 28.4 µg DDP/L

Details on test conditions:

TEST SYSTEM
- Test vessel: beaker
- Type: closed, covered with glass pane
- Material, size, headspace, fill volume: glass beakers filled with 50 mL test solution
- Dilution water: Purified drinking water. The purification included filtration with activated charcoal, passage through a lime-stone column and aeration
- Food: The unicellular alga Desmodesmus subspicatus at each test medium renewal, corresponding with 0.2 mg C/Daphnia/day
- Light intensity: 680 - 829 Lux
- Aeration: No
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Purified drinking water: filtered through activated charcoal, passed through a limestone column and aerated
- Conductivity: 171.6 µS/cm
- Culture medium different from test medium: No
- Intervals of water quality measurement: 0 and 48 hours

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light: 8 hours dark

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

>= 28.4 µg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Remarks:

DDP

Basis for effect:

mortality

Remarks:

growth, reproduction and time to first brood

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

>= 14.3 µg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

element

Remarks:

Palladium

Basis for effect:

mortality

Remarks:

growth, reproduction and time to first brood

Details on results:

The EC50 and EC10 could not be calculated but were considered to be > 28.4 µg DDP/L (14.3 µg Pd/L based on mortality, growth and reproduction and time to first brood.

Reported statistics and error estimates:

Regression analysis was used to determine EC values including the 95 % confidence interval using Probit-analysis assuming log-normal distribution of the values.
the
The LOEC was calculated by using ANOVA followed by Multiple comparisons (Dunnett's test or Mann-Whitney’s tests) after testing for normal distribution and variance homogeneity using the Shapiro-Wilk test and Levene test, respectively (p≤ 0.05).

Validity criteria fulfilled:

yes

Conclusions:

Diamminedichloropalladium (DDP) had no adverse chronic effect on Daphnia magna in a 21 day reproduction study. Neither adults nor juveniles were affected up to and including 28.4 μg DDP/L TWA (14.3 μg Pd/L TWA) . Accordingly, the NOEC was found to be ≥ 28.4 μg DDP/L (14.3 μg Pd/L).

Executive summary:

The effects of Diamminedichloropalladium (DDP) in a 21 day Daphnia reproduction study conducted according to OECD 211 were assessed. The study was a semi-static test design with test solutions renewals three times per week. The nominal test item concentrations used were 0.101, 0.324, 1.04, 3.32, 10.63, and 34.0 μg test item per litre. The chemical analysis conducted at the start and end of each test solution renewal was based on the concentration of palladium. The mean measured palladium concentrations in the freshly prepared test solution were between 54% and 105% of the nominal concentrations. At the end of the renewal period palladium concentrations were between 3 and 65% of nominal at the five highest test concentrations and were below the mean background levels of palladium in the lowest test concentration of 0.101 test item per litre treatment. The time weighted average (TWA) mean measured concentrations of 0.025, 0.062, 0.275, 1.16, 4.05, and 14.3 μg Pd per litre, corresponding with 0.049, 0.124, 0.546, 2.31, 8.04, and 28.4 μg DDP per litre were used for reporting of results.

No significant effects were found due to the test item up to and including the highest concentration tested. Survival (viability), growth (body length), and reproduction of adults, age to first brood and intrinsic rate were unaffected by the test item up to and including 28.4 μg DDP per litre (TWA) corresponding with 14.3 μg Pd per litre (TWA). The NOEC was found to be ≥28.4 μg DDP per litre or ≥14.3 μg Pd per litre.

The study was conducted in accordance with GLP and the validity criteria were met. The study was well-documented and adhered to guideline procedures and has been given a reliability rating of 1. This is considered to be a key study for this endpoint.

Reference 2

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, available as an unpublished report, no restrictions, fully adequate for assessment

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

97.01% Pd(NH3)4Cl2 (based on analyzed Pd content of 42.06%), rest water

Analytical monitoring:

yes

Details on sampling:

Dissolved Pd-concentrations were analysed in samples of all fresh test solutions at the beginning of the test prior to addition of test organisms and at each media renewal immediately after preparation of test solutions before distributing to the test vessels. Aged samples were taken before each media renewal and at test end. Analysis was performed in representative replicates per treatment level.

For this, duplicate samples were taken (10 mL), filtered (0.2 µm polyether sulphone membrane (PES) syringe filter) and afterwards acidified with conc. HNO3 (addition of 0.1 mL conc. HNO3 to the 10 mL sample). Samples were sealed after acidification.

Chemical analysis of the processed samples was performed immediately after sampling and within 24 hours.

Vehicle:

no

Details on test solutions:

Stock solutions were prepared by dissolving adequate amounts of test item in dilution water. A stirring bar were added to the bottle and stirred vigorously for 1 hour at room temperature (about 20°C). The stock solution was serially diluted with dilution water to obtain the test concentrations. The stock solutions were freshly prepared before each renewal.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain: unknown
- Source: Origin of the cladocerans was the German Federal Environment Agency, Institut für Wasser-, Boden- und Lufthygiene. Specimens used in the test are bred in the laboratory of the Fraunhofer IME
- Age at study initiation: 4-24 hours old

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

purified drinking water was enriched with an appropriate amount of CaCl2 to increase the hardness, resulting in values of about 250 – 350 mg CaCO3/L

Test temperature:

Temperature: 19.0 – 21.0 °C throughout the test, with the permitted range of 18 – 22 °C (with a variance of less than 2 °C)

pH:

pH: 7.34 – 9.60 at all treatment levels. Usually the pH should be between 6 and 9. The pH surpassed the range up to a pH of 9.6 in aged media of all assays. However, this is not unusual due to the activity of the feed algae in the vessels. Since all validity criteria are kept, this deviation is assumed to be without effect on the plausibility of the results and the integrity of the study.

Dissolved oxygen:

The dissolved oxygen saturation was between 8.69 mg/L and 13.8 mg/L which is above the required 3 mg/L

Nominal and measured concentrations:

Dose-range finding test:
nominal concentrations: 1-10-100 µg Pd/L (2.38, 23.8 and 238 µg test item/L)

Definitive test: nominal concentrations: Control, 3.84, 9.60, 24.0, 60.0 and 150 µg Pd/L,
equivalent to 9.13, 22.8, 57.1, 143 and 357 µg test item/L.

Mean measured (time weighted average) concentrations: control, 3.01, 6.68, 16.9, 42.7 and 106 µg Pd/L,
equivalent control, 7.16, 15.9, 40.2, 102 and 253 µg test item/L

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beaker (50 mL nominal volume)
- Type: beakers covered with glass pane to prevent from evaporation as much as possible, but also permit gaseous exchange between the medium and the atmosphere and access of light
- Material, size, headspace, fill volume: glass beakers filled with 50 mL test solution
- Dilution water: Purified drinking water. The purification included filtration with activated charcoal, passage through a limestone column, and aeration. In spite of the passage through a lime-stone column, hardness of the Cu-reduced water was below the recommended range (>140 mg/L as CaCO3) for Daphnia magna indicated in the guideline. Therefore, the dilution water was enriched with an appropriate amount of CaCl2 to increase the hardness, resulting in values of about 250 – 350 mg CaCO3/L.
- Food: the daphnids were fed during the test with a mixture of suspensions of unicellular alga Desmodesmus subspicatus and JBL ArtemioFluid in a ratio of 9 : 1 (vol/vol). The content of food in the test suspensions, measured at 585 nm, was kept constant at 0.2 mg C/(Daphnia * day).
- Light intensity: light intensity was measured using an illuminance meter (MINOLTA) with photometric sensor in Lux. With 908 – 985 lx (corresponding to 12.1 – 13.1 µE/(m² *s)) the light intensity was below the threshold value of about 1000 lx (15 µE/(m² *s)) as permitted by OECD guideline 211
- Aeration: No
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Purified drinking water
- Conductivity: 323 µS/cm
- Culture medium different from test medium: No
- Intervals of water quality measurement: 0 and 48/72 hours

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light, 8 hours dark

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

42.7 µg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

element (dissolved fraction)

Remarks:

Pd

Basis for effect:

reproduction

Remarks:

offspring per survived parent ('reproduction), immobility, length, age of first reproduction and intrinsic rate

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

102 µg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat. (dissolved fraction)

Remarks:

Tetraamminepalladium dichloride

Basis for effect:

reproduction

Remarks:

offspring per survived parents ('reproduction'), immobility, length, age of first reproduction and intrinsic rate

Key result

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

35.7 µg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

element (dissolved fraction)

Remarks:

Pd

Basis for effect:

reproduction

Remarks:

reproduction determined as offspring per survived parent

Remarks on result:

other:

Remarks:

95%CI: 22.3-45.0 µg Pd/L

Duration:

21 d

Dose descriptor:

EC10

Effect conc.:

84.9 µg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

reproduction

Remarks:

reproduction determined as offspring per survived parent

Remarks on result:

other:

Remarks:

95% CI: 53-107 µg TI/L

Details on results:

Based on the evaluation performed on TWM concentrations, for immobilisation a LC10 of 78.5 µg Pd/L, corresponding to 187 µg test item/L and a NOEC of 42.7 µg Pd/L, corresponding to 102 µg test item/L, were calculated. The EC50 value was 97.3 µg Pd/L (95%CI: ND).
For reproduction per survived daphnid an EC10 of 35.7 µg Pd/L, corresponding to 84.9 µg test item/L and a NOEC of 42.7 µg Pd/L, corresponding to 102 µg test item/L, were calculated. The EC50 value was 64.7 µg Pd/L (95%CI: 52.9-80.4 µg Pd/L).
In addition, also for age of first reproduction, length and intrinsic rate r a NOEC of 42.7 µg Pd/L, corresponding to 102 µg test item/L, and a LOEC of 106 µg Pd/L, corresponding to 253 µg test item/L, were determined. The respective EC50 values were not determinable, 148 µg Pd/L (95%CI: 129-185 µg Pd/L) and 99.9 µg Pd/L (95% CI ND).
For development rate, no effect could be observed, resulting in a NOEC of ≥ 106 µg Pd/L and a LOEC of > 106 µg Pd/L, corresponding to ≥ 253 and > 253 µg test item/L. The corresponding EC10 and EC50 values are 96 µg Pd/L (95% CI: 72.3-100 µg Pd/L) and 156 µg Pd/L (95% CI: 132-449 µg Pd/L), respectively.

Reported statistics and error estimates:

or each endpoint, the NOEC and LOEC, and, if possible, the EC50 and EC10 were determined. A LOEC was calculated by using ANOVA followed by Student-t test, Fisher`s Exact Binomial test, Dunnett’s or Williams’ test or an appropriate non-parametric test.
When the test results show a concentration-response relationship, the data were analysed by regression to determine the EC50 including the 95% confidence interval as well as the EC10 using Probit-analysis assuming log-normal distribution of the values. The computer software ToxRat® [6] was used for statistical evaluations.
The evaluation of the concentration-effect-relationships and the calculations of effect concentrations was based on the mean measured concentrations (time weighted mean, TWM) of palladium, since the deviation from the nominal concentrations was greater than ± 20%.

The five test concentrations showed mean measured concentrations of freshly prepared test solutions between 87.7 – 97.8 % of nominal concentrations. During the time interval until renewal of the test solution, mean test item concentrations decreased to 52.8 – 62.4 % of nominal. In the control, very low Pd concentration of 0.08 – 0.10 µg Pd/L were measured, slightly higher than the LOQ and clearly lower (30 to 50-fold) than the lowest test concentration. Since recovery rates did not remain in a range of ± 20% of nominal concentrations throughout the test, the evaluation of the effects was performed on basis of the time weighted mean test concentrations

In the highest test concentration, all exposed daphnids showed discoloration and were smaller than daphnids in the control. They also seemed to bit less active than daphnids in the control. No other clinical signs were observed.

TWM Pd Conc.	Parental immobilisation	Mean length on TD21	Mean cumm. Offspring per survived parent	Mean age of first brood	Mean development rate	Mean intrinsic rate
[µg/L]		[mm]	[Ind]	[d]	[Ind]
Control	10	4,37	78,0	10,4	0,100	0,310
3.01	9	4,27	73,4	11,5	0,097	0,303
6.68	8	4,51	98,8	10,5	0,102	0,320
16.9	10	4,49	73,9	10,3	0,100	0,298
42.7	10	4,04	63,7	9,1	0,111	0,322
106	3	2,84	11,0	12,0	0,084	0,115

Hypothesis testing in order to determine statistically significant differences was performed by comparing treatments against the control (NOEC/LOEC determination).

A significant and concentration dependent trend on immobility of individuals was observed. Thus, based on statistical evaluations and according to OECD 211, it is required to report the NOEC/EC_X for the cumulative offspring per introduced parent provided these values are lower than in the cumulative offspring per survived parent. Since the EC₁₀ for cumulative offspring per survived parent is lower than the EC₁₀ for cumulative offspring per introduced parent it was decided to consider the evaluation for reproduction based on the cumulative offspring per survived parent.

 By using Probit anaylsis a statistically significant concentration response was found for theimmobilisation of adult daphnids. To increase the reliability of the evaluation for immobilisation, the first and second concentration, which did not differ significantly from the control, were excluded for calculation of LCx-values and NOEC/LOEC.

Validity criteria fulfilled:

yes

Remarks:

The test fulfils all validity criteria as: mortality in controls (0%) is <20% & mean n° of offspring in control 21 d) was >60/female. In addition, dissolved oxygen conc was >3 mg/L & pH was 6 - 9 and did not vary by more than 1.5 units.

Conclusions:

During the 21-day exposure period significant effects of the test item on immobilisation, reproduction of survived parent daphnids, age of first reproduction, length and intrinsic rate r of the test organisms Daphnia magna were observed. No significant effect on the development rate was observed. Lowest NOEC of 42.6 µg Pd/L (corresponding to 102 µg test item/L) was observed for offspring per survived parent (reproduction), immobility, length, age of first reproduction and intrinsic rate. Lowest EC10 of 35.7 µg Pd/L (corresponding to 84.9 µg Test item/L) was calculated for offpring per survived parent (reproduction).

Executive summary:

The effects of tetraaminepalladium dichloruide in a 21 day Daphnia magna reproduction study conducted according to OECD 211 were considered. The study was a semi-static test design with test solutions renewals three times per week. The nominal test item concentrations applied were control, 3.84, 9.60, 24.0, 60.0 and 150 µg Pd/L, equivalent test item concentrations of control, 9.13, 22.8, 57.1, 143 and 357 µg test item/L.

The chemical analysis conducted at the start and end of each test solution renewal was based on the concentration of palladium. Mean measured concentrations in fresh test media were between 87.7 and 97.8 % of nominal concentrations. During the time interval until renewal of the test solution, mean concentrations were between 52.8 and 62.4 % of nominal concentrations. Thus, chemical analysis revealed that the deviation of the measured Pd-concentrations from the nominal test concentrations was greater than 20%. Therefore, the evaluation of the results was based on the time weighted mean of Pd-concentration: control, 3.01,

6.68, 16.9, 42.7 and 106 µg Pd/L, corresponding to 7.16, 15.9, 40.1, 102 and 253 µg test item/L.

During the 21-day exposure period to the test item Tetraamminepalladium(2+) dichloride significant effects on reproduction of survived parent daphnids, immobilisation, length, age of first reproduction and intrinsic rate r of the test organisms Daphnia magna were observed. No significant effect on the development rate could be observed.

Based on the evaluation performed on TWM concentrations for reproduction per survived parent daphnid, immobility, length, age of first reproduction and intrinsic rate r a NOEC of 42.7 µg Pd/L and a LOEC of 106 µg Pd/L were determined, corresponding to 102 µg test item/L and 252 µg test item/L, respectively.

For reproduction per survived parent daphnid an EC₁₀of 35.7 µg Pd/L, corresponding to 84.9 µg test item/L was calculated. For immobilisation of parant daphnids, a LC₁₀of 78.5 µg Pd/L, corresponding to 187 µg test item/L, and for length, an EC₁₀of 49.5 µg Pd/L, corresponding to 118 µg test item/L, were determined.

The study was conducted in accordance with GLP and the validity criteria were met. The study was well-documented and adhered to guideline procedures and has been given a reliability rating of 1. This is considered to be a key study for this endpoint.

Description of key information

Simon (2014): The 21-day NOEC for diamminedichloropalladium based on survival, growth and reproduction was ≥ 28.4 µg test item/L (14.3 µg Pd/L), the highest concentration tested.

Bruggeman (2019): The 21-day lowest NOEC for tetraamminepalladium dichloride was 102 µg test item/L (42.6 µg Pd/L), lowest EC10 was 84.9 µg test item/L (35.7 µg Pd/L) for reproduction.