Phosphonic acid, monoethyl ester, sodium salt (1:1)

Long-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

fish, juvenile growth test

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 Aug - 22 Sep 2014

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 215 (Fish, Juvenile Growth Test)

Version / remarks:

2000

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: control, 10, and 100 mg/L
- Sampling method: The highest and lowest test concentration were analysed when freshly prepared and immediately prior to renewal at the start of the study (day 0) and weekly thereafter (days 3, 9, 18, 28)
- Sample storage conditions before analysis: On each occasion, one sample was analysed after certain pre-treatment and the remaining samples were retained in case further analysis would be required

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test solutions were prepared by directly adding the appropriate amount of Fosetyl-Na in dilution water and stirring for 30 min. For the test concentrations of 10, 20, 40, 80, and 100 mg/L amounts of 0.0307, 0.0614, 0.1230, 0.2456, and 0.3074 g, respectively, were added to 3 L of dilution water.
- Controls: Negative Control (dilution water)

Test organisms (species):

other: Gobiocypris rarus

Details on test organisms:

TEST ORGANISM
- Common name: rare minnow
- Strain: batch no.: JF20140615
- Source: bred in the lab
- Age at study initiation (mean and range, SD): juvenile fish in exponential growth phase (hatch date: 15 Jun 2014 )
- Weight at study initiation (mean and range, SD): mean weight of 10 fish in each concentration tested were 0.0665, 0.0622, 0.0591, 0.0607, 0.0588, and 0.0557 g in control, 10, 20, 40, 80, and 100 mg/L, respectively
- Feeding during test : yes
- Amount: 4% dry weight related to the initial fish weight (about 0.15 g)
- Frequency: daily

ACCLIMATION
- Acclimation period: 14 days
- Acclimation conditions (same as test or not): feeding fry were maintained in holding tanks supplied with a continous flow of aerated water (90% of the air saturation) at 22 - 23 °C and 16 hours light/ 8 hours darkness photoperiod (1000-1500 lux)
- Type and amount of food: live food (Artemia) 4% dry weight related to the initial fish weight (about 0.15 g)
- Feeding frequency during acclimation: daily until 24 h prior to test
- Health during acclimation (any mortality observed): no mortality was observed in 7 days following a 48 h settling-in period

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

28 d

Test temperature:

22.5 - 23.5 °C

pH:

7.66 - 8.06

Dissolved oxygen:

71 - 90% of the air saturation value

Nominal and measured concentrations:

- Nominal concentrations: control, 10, 20, 40, 80, 100 mg/L
- Measured concentrations: see results

Details on test conditions:

TEST SYSTEM
- Test vessel: tanks made of chemically inert material with a sealable inert lid
- Size of vessel: 5 L
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 3 L
- Aeration: none
- Renewal rate of test solution (frequency/flow rate): every 72 hours
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.2 to 0.4 g of fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: good quality tap water hich has been dechlorinated for at least 12 hours
- Total organic carbon: 1.91 mg/L
- Metals: < 0.001 mg/L (As, Cd, Pb, Co, Ag); < 0.0002 mg/L (Hg); 0.001 mg/L Ni; 0.002 mg/L Cu; < 0.004 mg/L Cr6+; < 0.03 mg/L Fe; 0.25 mg/L Zn
- Pesticides: < 0.001mg/L (malathion, methyl parathion, and parathion)
- Alkalinity: pH = 7.55
- Culture medium different from test medium: no
- Intervals of water quality measurement: the characteristics of the dilution water are measured at least twice a year by Jiangsu Provincial Center for Disease Prevention and Control

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours light and 8 hours darkness
- Light intensity: 1000 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : mortality and any abnormal behaviour and external abnormalities (e.g. hemorrhage, discoloration) were recorded daily. Fish were considered dead if there was no visible movement and if touching of the caudal peduncle produced no reaction. Dead fish were removed as soon as possible. Dissolved oxygen, pH, and temperature were measured out in the beginning of the test and weekly thereafter (3, 9, 18, 28 d). Highest and lowest test concentration were analysed when freshly prepared and immediately prior to renewal at the start of the study (0 d) and weekly thereafter (3, 9, 18, 28 d).

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2, unless for the second highest concentration to the highest concentration (factor of 1.25)

RANGE-FINDING STUDY
- Test concentrations: 100 mg/L
- Results used to determine the conditions for the definitive study: No deaths and normal behaviour up to 100 mg/L in an 96-hour exposure period

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: based on mortality and growth

Duration:

28 d

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: based on mortality and growth

Reported statistics and error estimates:

For the estimation of the lowest observed effect concentration (LOEC), and the no observed effect concentration (NOEC), for the effects of the target substance on growth rate, the mean growth rate for each concentration were compared with the control mean using a multiple comparison method of Bartlett's tests (ANOVA, STATA 10.0).

Table 1. Measured concentrations of Fosetyl-Na in test medium

Nominal concentration (mg/L)	Analyzed concentration (mg/L)										Mean analyzed concentration (mg/L)
	Day 0	Day 3			Day 9		Day 18			Day 28
	fresh	aged	fresh	aged		fresh		aged	fresh	aged
Control	< LOD	< LOD	< LOD	< LOD		< LOD		< LOD	< LOD	< LOD	-
10	10.2	9.98	10.0	10.3		10.3		10.2	10.2	10.3	10.2
100	99.8	99.5	99.8	99.9		99.6		99.8	100	99.9	99.8

 

Table 2. Summary of mortality and abnormal behaviour data in a 28 day juvenile growth test with Gobiocypris rarus

Nominal concentration (mg/L)	Initial number of fish	Number of dead fish	Abnormal behaviour	Mean weight on day 0 (g)
Control	10	0	NB	0.0665
10	10	0	NB	0.0622
20	10	0	NB	0.0591
40	10	0	NB	0.0607
80	10	0	NB	0.0588
100	10	0	NB	0.0557

NB = normalbehaviour

 

Table 3. Individual weights at day 28 of a juvenile growth test with Gobiocypris rarus

	Control		10 mg/L		20 mg/L		40 mg/L		80 mg/L		100 mg/L
	Weights (g)	Rates (%)	Weights (g)	Rates (%)	Weights (g)	Rates (%)	Weights (g)	Rates (%)	Weights (g)	Rates (%)	Weights (g)	Rates (%)
1	0.163	3.22	0.152	3.21	0.163	3.67	0.144	3.10	0.139	3.08	0.148	3.53
2	0.157	3.10	0.167	3.55	0.150	3.38	0.157	3.42	0.132	2.92	0.133	3.13
3	0.147	2.85	0.166	3.53	0.154	3.47	0.143	3.06	0.142	3.17	0.141	3.35
4	0.153	3.01	0.149	3.14	0.161	3.62	0.138	2.93	0.137	3.04	0.137	3.26
5	0.161	3.19	0.157	3.33	0.154	3.48	0.149	3.23	0.153	3.43	0.156	3.71
6	0.157	3.10	0.170	3.61	0.167	3.77	0.146	3.15	0.145	3.25	0.144	3.41
7	0.144	2.79	0.151	3.20	0.152	3.42	0.139	2.97	0.136	3.01	0.134	3.17
8	0.153	3.00	0.148	3.11	0.153	3.44	0.143	3.07	0.151	3.40	0.129	3.03
9	0.173	3.43	0.164	3.49	0.148	3.34	0.158	3.43	0.130	2.87	0.139	3.30
10	0.157	3.09	0.155	3.29	0.152	3.42	0.141	3.03	0.148	3.31	0.146	3.46

 Rates = pseudo specific growth rates for the periods from day 0 to day 28

Table 4. Validity criteria OECD 215

Criterion from the guideline	Outcome	Validity criterion fulfilled
The mortality in the control(s) must not exceed 10 per cent at the end of the test.	Mortality was 0%	Yes
The mean weight of fish in the control(s) must have increased enough to permit the detection of the minimum variation of growth rate considered as significant.	Mean weight in the beginning was 0.0605 g and increased by more than 50% in the control.	Yes
The dissolved oxygen concentration must have been at least 60 per cent of the air saturation value (ASV) throughout the test.	71 – 90%	Yes
The water temperature must not differ by more than ± 1 °C between test chambers at any one time during the test and should be maintained within a range of 2°C within the temperature ranges specified for the test species.	22.5 – 23.5 °C	Yes

 

Validity criteria fulfilled:

yes

Remarks:

see Table 4 in 'any other information on results incl. tables'

Description of key information

NOEC (28 d) ≥ 100 mg/L, nominal, for Gobiocypris rarus (OECD 215, ≥ 99.8 mean measured concentration)

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

NOEC

Effect concentration:

>= 100 mg/L

Additional information

A 28 day GLP long-term toxicity study to fish is available for Fosetyl-Sodium. The study was performed according to OECD guideline 215 on the chronic toxicity to the rare minnow (Gobiocypris rarus) under semi-static conditions. Fish were exposed to the nominal concentrations of 10, 20, 40, 80, and 100 mg/L and an untreated control at 22.5 - 23.5 °C. The test item was dissolved in test water. No additional solvent was used. The test water was not aerated during the time of testing. Chemical analysis was performed for the highest and lowest test concentration when freshly prepared and immediately prior to renewal at the start of the study (day 0) and weekly thereafter (days 3, 9, 18, 28) using HPLC-MS/MS.

Analysis of the test solutions (fresh media and old media) showed that the measured concentrations were within +/- 20% of nominal values. Therefore, results were based on nominal concentrations. No mortality and no sub-lethal effects, as well as no effects on the growth were observed in the study. Thus, the NOEC (28 d) for rare minnow was determined to be ≥ 100 mg/L, nominal.