Tetrasodium (1-hydroxyethylidene)bisphosphonate

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1979-03-14 to 1979-03-28

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods

Qualifier:

according to guideline

Guideline:

other: methods based on  Methods for Acute Toxicity Tests with Fish, Macroinvertebrates, and Amphibians, USEPA, 1975

Deviations:

not applicable

Qualifier:

according to guideline

Guideline:

other: study carried out based on ABC protocol 7807

Deviations:

yes

Remarks:

lower flow rate

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 204 (Fish, Prolonged Toxicity Test: 14-day Study)

Deviations:

not applicable

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: All

- Sampling method: samples were taken on days 0, 1, 5, 10 and 14

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)

- Method: a stock solution of 256.5 g/L (method of dispersion not stated) was diluted according to achieve the required concentrations. The nominal concentrations of D2010 were determined by measuring predetermined dilutions of a stock Rhodamine B dye.

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM

- Common name: rainbow trout

- Source: Fish obtained from Spring Creek Hatchery, Lewiston, Montana. 

- Length at study initiation (mean): 33.2 mm

- Weight at study initiation (mean): 0.75 g

- Method of breeding: fish culture techniques were basically those described by Brauhn et al.

- Feeding during test

- Food type: standard commercial fish food (Rangen's)

- Amount: 3% body weight

- Frequency: daily


ACCLIMATION

- Acclimation period: at least 14 days, fish acclimatised to test temperature for 48 h.

- Acclimation conditions (same as test or not): similar holding conditions, but pH and ionic strength of the test solution 262 mg/L were significantly different from holding water (author's note).

- Type and amount of food: commercial fish food (Rangen's), 3% of body weight

- Feeding frequency: daily

- Health during acclimation (any mortality observed): some mortalities observed, mostly on the first day of stock arrival


QUARANTINE (wild caught)
- Duration:
- Health/mortality:

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

14 d

Hardness:

240 mg/L as CaCO3 (dilution water)

Test temperature:

Temperature maintained at 12 +/- 1 degree C.

pH:

The pH of the test media ranged between 6.3-6.6 in the high concentration to 7.1-7.2 in the low concentration. Control: 7.8-7.9

Dissolved oxygen:

>7.3 mg/L

Nominal and measured concentrations:

Nominal concentrations (mg/L active acid) were 30, 65, 125, 255, and 500 mg/L (active acid)
Actual concentrations determined on days 0, 1, 5, 10 and 14 and were 32, 60, 124, 270, and 529 mg/L (mean measures as active acid). See table 2 for details.

Details on test conditions:

TEST SYSTEM

- Test vessel: aquaria

- Material, size, headspace, fill volume: glass aquaria, filled with 30 L test solution

- Type of flow-through (e.g. peristaltic or proportional diluter): Mount & Brung proportional diluter

- Renewal rate of test solution (flow rate): 200 ml/min, the ABC protocol suggests a 300 ml/min flow rate

- No. of organisms per vessel: 30

- No. of vessels per concentration (replicates): 1

- No. of vessels per control (replicates): 1



TEST MEDIUM / WATER PARAMETERS

- Source/preparation of dilution water: well water

- Alkalinity: 300 mg/L as CaCO3

- Culture medium different from test medium: no

- Intervals of water quality measurement: Temperature, DO, and ammonia were measured on days 0, 1, 5, 10, and 14 in control, low concentration, and high concentration samples.


OTHER TEST CONDITIONS

- Adjustment of pH: not reported

- Photoperiod: 16 h light, 8 h dark


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : the fish were observed for mortality and abnormal behaviour every 24 h.


TEST CONCENTRATIONS

- Spacing factor for test concentrations: 1.9-2.1

- Range finding study: not conducted

Reference substance (positive control):

yes

Remarks:

Antimycin A

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

310 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

279 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Duration:

72 h

Dose descriptor:

LC50

Effect conc.:

200 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

180 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

60 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

behaviour

Remarks:

loss of equilibrium

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

195 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Details on results:

- Observations on body length and weight: (14 day) weight measurements of the surviving fish yielded the following weight percentages of the control group mean weight: 79% - 32 mg/L, 79% - 60 mg/L, 85% - 124 mg/L

- Other biological observations: backbone curvature was noticed on day 1 at higher concentrations, this has been explained by the author to be attributable to the different pH and ionic strength between holding water and exposure test water. Symptoms typically disappeared after 24 h.

- Mortality of control: 0%

Results with reference substance (positive control):

- Results with reference substance valid? yes

- LC50: not reported

- Results with reference substance valid? yes

- Mortality: 100% after 24 h at conc. 0.000042 mg/L

- LC50: 0.000032 mg/L

Reported statistics and error estimates:

LC50 and 95% confidence intervals determined by the method of Litchfield and Wilcoxon.

Sublethal observations / clinical signs:

Table 1. Recorded mortalities.

                                                           

Measured concentrations (mg/L active acid)	Mortalities (%)
	day 1	day 2	day 3	day 4	day 6	day 10	day 14
Control	0	0	0	0	0	0	0
32	0	0	0	0	0	0	0
60	0	0	0	0	0	0	0
124	0	0	0	0	0	0	0
270	33	53	90	93	93	100	100
529	97	100	100	100	100	100	100

8h

Determined LC50 (other than the ones transcribed above):

24 h: 310 mg/L active acid

48 h: 270 mg/L active acid

day 4-6: 200 mg/L active acid

day 7 -14: 180 mg/L active acid

Table 2. Nominal and measured concentrations of D2010 as active acid.

Nominal Concentrations (mg/L active acid)	Measured concentrations (mg/L active acid)
	Day 0		Day 1		Day 5		Day 10		Day 14		Mean
	P	D2010	P	D2010	P	D2010	P	D2010	P	D2010	P	D2010
0	0.05	-	0.05	-	0.2	-	0	-	0	-	-	-
30	9.9	33	4.5*	15*	10.5	35	9.1	30	8.7	29	9.6	32
65	19.75	66	1.2*	4*	19	63	15	50	18.5	62	18	60
125	30	100	5.0*	17*	40	133	38.5	128	40	133	37	124
255	74	246	31*	103*	85	283	80	266	85	283	81	270
500	157.5	524	150	499	165	549	155	516	167.5	558	159	529

* author's note: Day 1 phosphorous values were much lower than expected and were not used in the calculations for mean Dequest 2010 levels. It was not determined why these samples did not correspond to phosphorous concentrations measured previously (day 0) and later (day 5). However, it was observed that sample aliquots during the analytical process were cloudy; indicating an interfering complex or incomplete sample digestion. An acceptable day 1 high concentration indicated that no malfunctions had occurred in the test system.

In order to achieve D2010 from P values the following equation was followed:

Total P (mg/L) * 5.54 (conversion factor for converting P to 2010 ) * 0.601 (correction factor for percent purity) = D2010 (mg/L active acid)

Validity criteria fulfilled:

yes

Conclusions:

A 96 h LC50 value of 195 mg/L (as active acid) and a 14 day LC50 of 180 mg/L (as active acid) have been determined for the effects of HEDP acid (CAS 2809-21-4) on mortality of the freshwater fish Salmo Gairdneri (now known as Onychorhynchus mykiss).

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Remarks:

no GLP and no analytical monitoring

Qualifier:

according to guideline

Guideline:

other: US EPA 1975, Methods for acute toxicity tests with fish, macroinvertebrates and amphibians.

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

not applicable

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: an appropriate amount of CP65184 was added directly to the sea water in each test vessel to obtain the desired concentrations.

- Controls: sea water only

Test organisms (species):

Cyprinodon variegatus

Details on test organisms:

TEST ORGANISM

- Common name: sheepshead minnow

- Source: stocked ponds at BMRL

- Length at study initiation (range): 15-20 mm

- Method of breeding: not reported

- Feeding during test: animals were not fed during test



ACCLIMATION

- Acclimation period: min. 7 days

- Acclimation conditions (same as test or not): not reported

- Health during acclimation (any mortality observed): <1%

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

20 +/- 1 degC

pH:

Initial pH ranged from 5.4 in the highest concentration to 7.0 in the lowest concentration, controls were 8.3. Final pH ranged from 5.5 in the highest concentration to 7.1 in the lowest concentration, controls were 8.2.

Dissolved oxygen:

≥ 60% saturation

Salinity:

25 parts per thousand

Nominal and measured concentrations:

Nominal test concentrations were 1000, 10000, 18000, 32000 and 56000 ppm (ul/L)

Details on test conditions:

TEST SYSTEM

- Test vessel: jars

- Type (delete if not applicable): open

- Material, size, fill volume: 19 L uncovered glass jars with 15L test medium.

- Aeration: no

- No. of organisms per vessel: 10

- No. of vessels per concentration (replicates): 1

- No. of vessels per control (replicates): 1


TEST MEDIUM / WATER PARAMETERS

- Source/preparation of dilution water: natural filtered (5 um filters) sea water

- Culture medium different from test medium: no

- Intervals of water quality measurement: pH was measured at the beginning and end of the test; salinity, temperature and DO intervals of measurements are not reported.


OTHER TEST CONDITIONS

- Adjustment of pH: no

- Photoperiod: not reported

- Light intensity: not reported


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : mortality was observed at 0, 24, 48 and 96 h.


TEST CONCENTRATIONS

- Spacing factor for test concentrations: 1.8-2

- Range finding study: not reported

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

104 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: results corrected for active acid and density of TS

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

2 180 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: results are corrected for active acid and density of TS

Details on results:

- Mortality of control: 0%

Reported statistics and error estimates:

LC50 values were calculated by linear regression.

Sublethal observations / clinical signs:

Table 1. Summary of findings.

Nominal concentraions (ul/L/ppm)	Mortalities (%)
	24 h	48 h	96 h
Control	0	0	0
1,000	0	0	0
10,000	0	0	0
18,000	30	50	50
32,000	50	80	90
56,000	100	100	100

Result expressed as nominal concentration. Properties of the test substance and evidence from other studies (where
concentrations were measured) indicate that nominal and measured concentrations are likely to be in good agreement.

LC50 corrected for density (1.1) and % active acid (9.42%),

24 hr LC50: 2455 mg/L active acid and 48 hr LC50: 2240 mg/L active acid (corrected), uncorrected values were 24 hr LC50: 23644, 48 hr LC50: 21573 and 96 hr LC50: 20997 all in uL/L (reported as ppm).

Validity criteria fulfilled:

no

Remarks:

the pH of the test medium lowered with increasing concentration

Conclusions:

A 96 h LC50 value of 2180 mg/L (active acid) has been determined for the effects of HEDP acid (CAS 2809-21-4) on mortality of saltwater water fish C. variegatus.

Description of key information

(96 h) LC50 278 mg/L (r-a) O. mykiss

(96 h) LC50 3111 mg/L (r-a) C. variegatus

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

278 mg/L

Marine water fish

Marine water fish

Effect concentration:

3 111 mg/L

Additional information

Freshwater

No data on short term toxicity of HEDP-4Na salts, 3794 -83 -0, to freshwater fish are available. However, a reliable 96 h LC50 value of 195 mg/L (as active acid) has been determined for the effects of HEDP acid (CAS 2809-21-4) on mortality of S. gairdneri (now known as O. mykiss). This value is taken from a 14 day study, where the 96 h LC50 has been reported. The same study determined a 14 d NOEC of 60 mg active acid/L and a 14 d LC50 of 180 mg active acid/L with S. gairdneri (now known as O. mykiss) and HEDP acid 2809-21-4.

The results of tests conducted on HEDP and its salts are directly comparable, because the ionisation state will depend only on the pH of the test medium. The counterion sodium is not considered to contribute to effects in fish. At environmentally-relevant pH values, HEDP will be ionised typically three times, and will form stable complexes with metal ions.

Here, the value of 195 mg equivalent active acid/L may be directly read-across.

The 96 h LC50 for HEDP 2809-21-4 is 195 mg/L, which is equivalent to a 96 h LC50 value of 195 * 1,427 = 278 mg/L HEDP-4Na (CAS 3794 -83 -0)

[using MW(HEDP-4Na)/MW(HEDP) = 293.96 / 206.03 = 1.427 ]

 

Marine

No data on short term toxicity of HEDP-4Na salts, 3794 -83 -0, to marine fish are available. However, a reliable 96 h LC50 value of 2180 mg/L (as active acid) has been determined for the effects of HEDP acid (CAS 2809-21-4) on mortality of Cyprinodon variegatus.

The results of tests conducted on HEDP and its salts are directly comparable, because the ionisation state will depend only on the pH of the test medium. The counterion sodium is not considered to contribute to toxicity to fish. At environmentally-relevant pH values, HEDP will be ionised typically three times, and will form stable complexes with metal ions.

Here, the value of 2180 mg equivalent active acid/L may be directly read-across.

The 96 h LC50 for HEDP 2809-21-4 is 2180 mg/L, which is equivalent to a 96 h LC50 value of 2180 * 1.427 = 3111 mg/L HEDP-4Na (CAS 3794 -83 -0)

[using MW(HEDP-4Na) / MW (HEDP) = 293.96 / 206.03 = 1.427 ]