MoVNbTe mixed oxide

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2022-02-17 to 2022-02-25 at the test facility with the definitive exposure period from 2022-02-21 to 2022-02-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

(2019)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Test item Mo10V3TeNbO42
Batch number EX.14402.600
CAS No. 146569-48-4
Active ingredient
Niobium trivanadium decamolybdenum tellurium dotetracontaoxide
Purity 87.6 %
Water solubility < 0.2 g/L practically insoluble
Mo content 47.87 % (stoichiometric content)
Water solubility of Mo
11.498 mg/L (at pH 8.5, after 4 d shaking at 100 rpm of 100 mg/L Mo10V3TeNbO42; Affolter, 2020)
Stability under test conditions
Not specified
Appearance Black solid powder
Expiry date 2026-03-10
Recommended storage Store tightly closed, cool and dry / 20 °C

Analytical monitoring:

yes

Remarks:

via element analysis of Mo

Details on sampling:

Analytical Monitoring

The molybdenum concentration was determined via element analysis of Mo, processed at the Test Site. Test item samples were labelled with the test number, the concentration, the code for the sampling times, the replicate number, the date and the sampler, according to the test facility SOP and delivered to the Test Site for the analytical delegated phase. A full chain of custody was maintained. For the Phase Report with the description of the method used for the delegated phase see Annex I.

Sampling schedule
The limit loading and the control were analysed via element analysis of Mo in the fresh media at the start of exposure and at the renewal of the test solutions (0 and 72 hours) as well as in the 24-hours old media at the renewal and at the end of the exposure (24 and 96 hours).

All samples were taken in duplicate. The samples were stored at the test facility at room temperature. The first replicate was shipped pooled to the test site at the end of the exposure phase for analysis. The principle investigator was informed by the study director prior to shipment. Analysis was performed as described in the phase plan.

Vehicle:

no

Details on test solutions:

Limit loading rate
47.0 mg test item/L were tested as a threshold concentration in a limit test. The selection of the test concentration is based on the derivation of a threshold concentration (TC) from an algae toxicity test (ErL50 (0-72 h) > 100 mg/L, nominal loading of the saturated solution) (Füller, 2021) and a daphnia toxicity (EL50 (0-48 h) 47.0 (22.1 – 97.2) mg test item/L, nominal loading of the saturated solution), (Füller, 2021, unpublished).

Preparation of the test medium
A saturated solution with a nominal loading concentration of 100 mg test item/L of test item was prepared at room temperature 96 ± 2 hours prior to the start of the exposure and prior each renewal of the media. An appropriate amount of the test item was weighed out and transferred with dilution water to a glass flask with dilution water. The saturated solution was stirred with about 300 rpm for 96 ± 2 hour at room temperature. After a separation phase of 30 minutes, the saturated solution was removed by siphoning from the approximate center of the glass bottle. Undissolved particles were removed by membrane filtration (membrane filter 0.45 µm, RC, MACHEREY-NAGEL). The filter was saturated in order to avoid adsorption during the filtration. The first 25 mL of the filtrate were discarded. The filtration was interrupted for 15 minutes to allow for adsorption and saturation of the filter material with dissolved test item. Thereafter, the filtration was continued. The next 25 mL were discarded. During filtration, the filter was always kept covered. The following filtrate, i.e. the saturated solution, was used in the test by diluting it with dilution water to a nominal loading of 47 mg test item/L The saturated solution was checked via laser beam (Tyndall effect) for undissolved test item. The Tyndall effect was negative.

Control
7 fish in dilution water (without test item but from the same source and age as used for the test solution) were tested under the same test conditions as the test replicate.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

Test organism
Danio rerio (zebrafish)
Vertebrata, Gnathostomata, Pisces, Osteichthyes, Teleostei, Cypriniformes, Cyprinidae

Reason for the selection of the test system
According to the guideline, Danio rerio is suitable for this kind of study.

Origin
All fish used in the test were gained at Noack Laboratorien GmbH from a single brood stock (supplier: Umweltbundesamt, Schichauweg 58, D-12307 Berlin)

Holding
Holding was performed in accordance with the laboratory SOPs at the test facility at 23 ± 2 °C and diffuse light (540 – 1000 Lux, photoperiod of 12 to 16 h light / 12 to 8 h dark). The water was changed at least once per week. The dissolved oxygen concentration was more than 80% of the air saturation value. Zebrafish with at least 9 days of acclimatization and mortality < 5% within these days prior to the start of the exposure were used in the test. No mortality was observed during this time. No disease treatments were administered throughout holding and testing.

 
Water
Tap water of local origin was used for holding. The water was filtered on activated charcoal and aerated for at least 24 h to remove chlorine.
Nominal water parameters:
Total hardness: 40 - 250 mg CaCO3/L
pH-value: 6.0 - 8.5
Oxygen saturation: ≥ 80 % of air saturation value
Acidity: 0.1 mmol/L (recent measurement 2021-11-09)
Alkalinity: 0.8 mmol/L (recent measurement 2021-11-09)
Conductivity: 174 µS/cm (recent measurement 2021-11-09)

Acclimatization
A sufficient number of fish were acclimatized to dilution water for at least 9 days prior to start of the exposure. The environmental conditions were equivalent to the test conditions.

Feeding
Food was provided 3 times per week. Food was given to satiation (4% of the fish body weight) per feeding day. The test fish were not fed 24 h before test start.

Food Sera Vipan; SERA GMBH, 52518 Heinsberg, Germany

Test type:

semi-static

Water media type:

freshwater

Remarks:

Tap water of local origin was used for holding. The water was filtered on activated charcoal and aerated for at least 24 h to remove chlorine.

Limit test:

yes

Total exposure duration:

96 h

Hardness:

see section "Any other information on results incl. tables" below.

Test temperature:

see section "Any other information on results incl. tables" below.

pH:

see section "Any other information on results incl. tables" below.

Dissolved oxygen:

see section "Any other information on results incl. tables" below.

Nominal and measured concentrations:

47.0 mg test item/L were tested as a threshold concentration in a limit test.

Details on test conditions:

Reference item
No reference item is recommended for this test according to the guidelines.

Duration of the test 96 hours

Test vessel
Glass aquaria of 3 L were used (dimensions: 11.5/13/20 cm, depth of water: approx. 17 cm) and covered with glass plates.

Test volume About 2.5 L per vessel

Aeration No aeration was provided.

Dilution water Same as used for holding

Acclimatization
Acclimatization and the environmental conditions were not necessary, because the quality of the dilution water was equivalent to the water used for holding.

Number of fish
7 zebrafish were used each for the limit loading and the control. For the whole study 14 fish were used.

Loading, length of fish
The fish density in the test vessels was 0.0756 g fish per liter test solution.
Average body length at the test end: 1.36 cm
Average body weight at the test end: 0.027 g
The fish were as homogeneous as possible.

Age of fish Juvenile

Replicates One replicate each for the limit loading and the control

Introduction of fish The fish were randomly distributed among the treatments.

Feeding of test fish The fish were not fed during the exposure.

Water temperature (target) 21 - 25 °C, controlled at ± 1 °C

Dissolved oxygen Concentration (target) Not less than 60% of air saturation value

Light intensity (target) 540 – 1000 Lux

Photoperiod
A photoperiod (light / dark cycle: 16 / 8 h) was maintained during exposure.

Type and Frequency of Measurements
Biological Parameters

Observation intervals
Observations were made after 2 ± 0.5 h and 5 ± 1 h after the start of exposure. On days 1 – 4 of the test, all vessels with living fish were inspected twice per day.
No mortality, no presence of visible abnormalities in regard to equilibrium, appearance, swimming behavior, ventilatory behavior function and no additional clinical signs were observed.

Criteria of effects
Fish were considered dead if there was no visible movement (e.g. gill covers movement) and if touching of the caudal peduncle produced no reaction. Records were kept of visible abnormalities (e.g. loss of equilibrium, swimming behavior, respiratory function, pigmentation, etc.).

Measurement of fish size and weight
For measurements of size and weight all control fish were used at the end of the exposure.

 
Physico-chemical Properties

Water quality and light intensity measurements
The pH-value, temperature and oxygen saturation were measured in all test vessels at the start and the end of exposure as well as daily and after renewal of the test media from fresh and old test media.
Chlorine and nitrate were measured from one batch of dilution water used in the test. Total hardness and TOC were determined at the start of the exposure in the dilution water. During the test, the water temperature was recorded continuously (once per hour) with a data logger. The light intensity on the surface of the test aquaria was measured at the start of the exposure. The solution appearance and behavior was observed daily before and after renewal.


Analytical Monitoring

The molybdenum concentration was determined via element analysis of Mo, processed at the Test Site. Test item samples were labelled with the test number, the concentration, the code for the sampling times, the replicate number, the date and the sampler, according to the test facility SOP and delivered to the Test Site for the analytical delegated phase. A full chain of custody was maintained. For the Phase Report with the description of the method used for the delegated phase see Annex I.

Sampling schedule
The limit loading and the control were analysed via element analysis of Mo in the fresh media at the start of exposure and at the renewal of the test solutions (0 and 72 hours) as well as in the 24-hours old media at the renewal and at the end of the exposure (24 and 96 hours).

All samples were taken in duplicate. The samples were stored at the test facility at room temperature. The first replicate was shipped pooled to the test site at the end of the exposure phase for analysis. The principle investigator was informed by the study director prior to shipment. Analysis was performed as described in the phase plan.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LL0

Effect conc.:

>= 47 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

calculated nominal loading from a saturated solution

Basis for effect:

mortality (fish)

Details on results:

Biological Data

The effect level is given based on the nominal loading concentration of the test item after 96 hours. The LL0 after 96 hours was ≥ 47.0 mg/L.

Physicochemical Data

The freshly prepared and the 24 hours aged test solutions were visibly clear. The environmental conditions (pH-value, temperature and oxygen-saturation) were determined to be within the acceptable limits. Light intensity was measured at the start of the exposure and ranged from 662 to 780 (mean value 722) Lux.

The study was performed according to the OECD-Guideline No. 203 (2019) for Testing of Chemicals and GLP principles and met the validity criteria:
• The O2 saturation should be  60% (this study: ≥ 74%).
• The mortality in the control group should not exceed 1/7 fish (this study: 0/7 fish).
• Recovery rates of the test item 80 - 120% throughout the test (otherwise determination of EL-levels will be given as recommended in the Guidance Documents on Aquatic Toxicity (OECD (2019): Guidance document on aqueous-phase aquatic toxicity testing of difficult test chemicals. OECD series on testing and assessment no. 23 (second edition), ENV/JM/MONO(2000)6/REV1)

Results with reference substance (positive control):

No reference item is recommended for this test according to the guidelines.

Reported statistics and error estimates:

Evaluation The LL-value after 96 h was empirically derived from the raw data of the limit loading.

Software All data were computer-processed and rounded for presentation. Consequently, minor variations may occur from the original figures if manual calculations based on the original figures are made subsequently. The following software was used for calculations:
Excel, MICROSOFT CORPORATION

Sublethal observations / clinical signs:

Observations of Sub Lethal Effects and Normal Behavior in the Test Vessels
(n = 7, number of fish)

Nominal loading	Effect *	Number of fish affected at observation time [hours]
[mg/L]		day 0		day 1		day 2		day 3		day 4
		2	4	22	28. Mai	47	53	71.5	76.5	94	96
47.0	(1)	7	7	7	7	7	7	7	7	7	7
Control	(1)	7	7	7	7	7	7	7	7	7	7

*) The numbers in brackets correspond to the following observations:
(1) = Normal behavior

Cumulative Mortality % in the Test Vessels

 

Nominal loading	Cumulative mortality [%] at observation time [hours]
[mg/L]	day 0		day 1		day 2		day 3		day 4
	2	4	22	28.5	47	53	71.5	76.5	94	96
47.0	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

 

Environmental Conditions

	Nominal Test item loading	Oxygen Saturation		pH Value		Temperature
	[mg/L]	[%]
						[°C]
		new	old	new	old	new	old
Day 0	47.0	98		8.11		23.8
	Control	94		8.10		23.7
Day 1	47.0	93	84	7.53	8.49	23.7	23.4
	Control	95	80	7.55	8.22	23.8	23.8
Day 2	47.0	100	84	7.26	8.38	23.5	23.7
	Control	100	74	7.76	8.33	23.8	23.8
Day 3	47.0	99	93	7.56	7.53	23.6	24.0
	Control	99	91	7.61	7.46	23.4	24.0
Day 4	47.0		88		7.56		24.0
	Control		90		7.51		24.0

 

Water Temperature in the Control (Continuous Measuring)

 

Period of Measurements	2022-02-21 to 2022-02-25
Minimum Temperature [°C]	23.3
Maximum Temperature [°C]	24.3
Mean Temperature [°C]	23.9

 

Residual Chlorine from the Dilution Water

Batch	For Study Days	[mg/L]
Day 0 – Day 3	0 to 4	< 0.01

Nitrate from the Dilution Water

Batch	For Study Days	[mg/L]
Day 0	0 to 1	2.27
Day 1	1 to 2	2.91
Day 2	2 to 3	2.54
Day 3	3 to 4	2.26

TOC and total Hardness from the Dilution Water

Parameter	Study Day	[mg/L]
TOC	0	< 2
Total Hardness	0	59

 

Results

The purpose of this study was the determination of the concentration of molybdenum in test medium with dissolved test item Mo10V3TeNbO42 prepared by Noack Laboratorien (study no. SO21065 / FAZ19517). Therefore, the provided sample solutions were acidified and analysed by inductively coupled plasma and optical emission spectrometry (ICP-OES).

The validity of the applied analytical method was proven (specificity, linearity, limit of quantification (LOQ), accuracy and precision). The validation results meet the specifications according to SANTE/2020/12830, Rev.1.

The mean results of the provided sample solutions are summarized in the following table:

Date	Sample	Nominal sample concentration	Mean measured	Recovery^1)
	code		Mo concentration
[yyyy-mm-dd]			[mg/L]	[%]
04.03.2022	A0	Control	< 0.01	-
04.03.2022	E0	Control	< 0.01	-
04.03.2022	A1	Control	< 0.01	-
04.03.2022	E1	Control	< 0.01	-
04.03.2022	A0	47.0 mg/L	4.000	17.78
04.03.2022	E0	47.0 mg/L	3.954	17.57
04.03.2022	A1	47.0 mg/L	4.621	20.54
04.03.2022	E1	47.0 mg/L	4.622	20.54

1) Nominal loading rate of the test item preparation as used in study FAZ19517: 47.0 mg/L

 

Data for the evaluation of the linearity of the molybdenum calibration solutions.

 

Concentration	Intensity	Intensity	Intensity	Intensity	Regression residuals
	mean value	value 1	value 2	value 3
[mg/L]	[counts/s]	[counts/s]	[counts/s]	[counts/s]	[%]
0.00	2.290	3.170	1.479	2.222	-
0.009	6.243	7.121	5.916	5.691	11.3
0.03	18.32	19.21	17.50	18.250	4.6
0.05	29.73	29.79	29.30	30.09	3.1
0.1	57.43	57.08	57.55	57.67	0.5
0.3	169.3	169.4	167.7	170.7	-0.6
0.5	287.4	285.5	288.3	288.5	1.5
1.0	561.9	560.3	561.2	564.0	-0.7
3.0	1688	1689	1687	1688	-0.5
5.0	2834	2828	2837	2837	0.2

 

1st Level (0.03 mg/L, LOQ level):

Data for the evaluation of the recovery and repeatability of Mo concentration 1st level using spiked matrix samples (0.03 mg/L).

 

Validation	Nominal Mo	Measured Mo	Recovery	Intensity mean	Intensity value 1	Intensity value 2	Intensity value 3
sample no.	concentration	concentration
	[mg/L]	[mg/L]	[%]	[counts/s]	[counts/s]	[counts/s]	[counts/s]
1	0.03	0.03148	104.93	18.40	19.26	17.47	18.45
2	0.03	0.03202	106.73	18.71	18.58	18.99	18.57
3	0.03	0.03217	107.23	18.79	19.19	18.70	18.49
4	0.03	0.03096	103.20	18.11	18.27	17.77	18.28
5	0.03	0.03135	104.50	18.33	18.66	18.40	17.93
Mean			105.32
SD			1.65
%RSD			1.57

 

2nd Level (0.30 mg/L, 10 x LOQ level):
Data for the evaluation of the recovery and repeatability of Mo concentration 2nd level using spiked matrix samples (0.30 mg/L).

Validation	Nominal Mo	Measured Mo	Recovery	Intensity mean	Intensity value 1	Intensity value 2	Intensity value 3
sample no.	concentration	concentration
	[mg/L]	[mg/L]	[%]	[counts/s]	[counts/s]	[counts/s]	[counts/s]
6	0.30	0.30539	101.80	173.3	172.5	173.3	174.2
7	0.30	0.30433	101.44	172.7	172.5	172.7	173.0
8	0.30	0.30575	101.92	173.5	173.0	173.6	173.9
9	0.30	0.30380	101.27	172.4	172.0	171.5	173.6
10	0.30	0.30840	102.80	175.0	173.3	174.3	177.3
Mean			101.85
SD			0.59
%RSD			0.58

 

Results of molybdenum concentration in provided samples solutions (with dissolved test item Mo10V3TeNbO42 and control samples).

No. of analy-sis	Date	Sample	Nominal sample concentration	Intensity	Mean measured Mo concentr. *	Recovery^1)
	[yyyy-mm-dd]	code		[counts/s]	[mg/L]	[%]
1	04.03.2022	A0	Control	1.715	< 0.01	-
2	04.03.2022	A0	Control	2.509
1	04.03.2022	E0	Control	1.384	< 0.01	-
2	04.03.2022	E0	Control	2.790
1	04.03.2022	A1	Control	2.388	< 0.01	-
2	04.03.2022	A1	Control	2.030
1	04.03.2022	E1	Control	1.428	< 0.01	-
2	04.03.2022	E1	Control	1.793
1	04.03.2022	A0 47.0 mg/L	47.00 mg/L	2279.0	4.000	17.78
2	04.03.2022	A0 47.0 mg/L	47.00 mg/L	2285.0
1	04.03.2022	E0 47.0 mg/L	47.00 mg/L	2252.0	3.954	17.57
2	04.03.2022	E0 47.0 mg/L	47.00 mg/L	2261.0
1	04.03.2022	A1 47.0 mg/L	47.00 mg/L	2632.0	4.621	20.54
2	04.03.2022	A1 47.0 mg/L	47.00 mg/L	2642.0
1	04.03.2022	E1 47.0 mg/L	47.00 mg/L	2628.0	4.622	20.54
2	04.03.2022	E1 47.0 mg/L	47.00 mg/L	2647.0

* Results of Mo concentration were calculated considering the applied sample volume of 9 mL acidified and diluted to 10 mL.

1) Nominal loading rate of the test item preparation as used in study FAZ19517: 47.0 mg/L

 

 

 

 

Validity criteria fulfilled:

yes

Conclusions:

The effect level is given based on the nominal loading rate of the test item Mo10V3TeNbO42. The threshold loading of the test item at the nominal loading 47.0 mg test item/L was found to have neither lethal nor non-lethal effects on zebrafish.

Executive summary:

The acute toxicity of the test item Mo10V3TeNbO42 to fish (zebrafish) was determined according to the principles of the OECD-Guideline for Testing of Chemicals No. 203 (2019) from 2022-02-17 to 2022-02-25 at the test facility with the definitive exposure period from 2022-02-21 to 2022-02-25.

The test item is a solid. A limit test item loading rate of 47.0 mg test item/L was tested as a threshold loading in a limit test. The loading rate was based on the derivation of a threshold loading (TL) from an algae toxicity test (ErL50 (0-72 h) > 100 mg test item/L, nominal loading of the saturated solution) (Füller, 2021) and a daphnia toxicity test (EL50 (0-48 h) 47.0 (22.1 – 97.2) mg test item/L, nominal loading of the saturated solution), (Füller, 2021, unpublished).

The study was performed over a period of 96 hours under semi-static conditions with daily renewal of the test solutions. Seven test organisms were exposed to the test loading and the control, respectively. Water quality parameters (temperature, pH-value and oxygen-saturation) measured at 0, 24, 48, 72 and 96 hours were determined to be within the acceptable limits. The test solutions were visually clear throughout the study.

The loading rate of the test item Mo10V3TeNbO42 was analytically verified via determination of the molybdenum concentration by the test site. Analysis was carried out in the fresh media at the start of the exposure and at the renewal of the test solutions (0 and 72 hours) as well as in the 24 hours old media at the renewal and at the end of the test (24 and 96 hours) in the limit loading rate and in the control.
The effect value given was based on the nominal test item loading. All validity criteria of the test guideline were met. The threshold loading of the test item at 47.0 mg/L, was found to have neither lethal nor non-lethal effects on zebrafish.

LL-Values (96 hours) of the Test item
based on the nominal loading rate of the test item [mg/L]

LL0	≥47.0*
Highest test item loading with
0% mortality after 96 h

*calculated nominal loading from a saturated solution

Description of key information

The effect level is given based on the nominal loading rate of the test item Mo10V3TeNbO42. The threshold loading of the test item at the nominal loading 47.0 mg test item/L was found to have neither lethal nor non-lethal effects on zebrafish.