Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

Data is from the exeprimental report followed standard gudeline.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Principles of method if other than guideline:

Determination of the effect of Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex(CAS No12225-21-7) on the
growth of freshwater green alga Chlorella vulgaris (OECD Guideline No.201).

GLP compliance:

not specified

Specific details on test material used for the study:

- Name of test material (IUPAC name): aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl]diazenyl}benzenesulfonate)
- Common name: FD&C Yellow No. 5 Aluminum Lake
- Molecular formula: C48H33AlN12O27S6
- Molecular weight: 495.4038 g/mol
-InChl:1S/C16H12N4O9S2.Al/c21-15-13(18-17-9-1-5-11(6-2-9)30(24,25)26)14(16(22)23)19-20(15)10-3-7-12(8-4-10)31(27,28)29;/h1-8,13H,(H,22,23)(H,24,25,26)(H,27,28,29);/b18-17+;
- Substance type: Organic
- Physical state: solid

Analytical monitoring:

no

Details on test solutions:

The test solution was prepared in aseptic condition. The test item Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex was prepared by adding 50 mg of test item in 250 ml of BBM to get the final concentration of 200 mg/L. This stock solution was kept for stirring/sonication for 30 minutes to obtain a homogenous solution for the experiment. The test concentrations were chosen according to the available data of the test item.
The concentrations chosen were set up to the water solubility limit. The remaining test solutions were prepared by dilution from the above stock solution. To have a
better growth and visibility of cells, the initial of the culture was kept 1 X 104cells/ml. Care was taken to have a homogeneous solution for the experiment.

Test organisms (species):

Chlorella vulgaris

Details on test organisms:

The fresh water green alga Chlorella vulgaris, was used as the test organism. Sterile, unicellular, liquid cultures of algae.
The culture was examined under the microscope to confirm that it was unicellular, healthy and not contaminated.
The medium to be used for the growth of algae was Bold’s Basal Medium (BBM). It is a medium composed of macronutrients, micronutrients, alkaline EDTA solution and Iron solution. Stock solution of each of these was prepared separately and then a complete medium was prepared and sterilized. De-ionized water was used to prepare the BBM.

Test type:

static

Water media type:

freshwater

Total exposure duration:

72 h

Test temperature:

24 °C ±2°C.

pH:

6.7 to 7.4

Nominal and measured concentrations:

6.25 mg/L,12.5 mg/L,25 mg/L,50 mg/L,100 mg/L,200 mg/L.All the six concentration were in geometric series spaced by a factor of 2.

Details on test conditions:

Light Quality White Fluoroscent Light
Light Intensity:1500Lux
Photoperiod 16 Hour Light Period : 8 Hour Dark Period
RPM Speed 120 Revolutions per minute
Water Deionized Waer
Test vessel size 100ml
Test solution volume 60ml
Dilution water Bold’s Basal Medium
Test concentration Six test concentration were:
Replicates per concentration
Three replicates for Control
Two replicates for each test concentration

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 200 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: calculated from equation through probit analysis

Details on results:

The microscopic observations were also noted in each of the experimental flasks. All the cells appeared healthy, round and green throughout the test duration in the control and in the experimental flask also no significant changes were observed.

Table 1: Showing the average cell count using Haemocytometer of the test vessels at an

equal interval of 24hrs, 48hrs and 72hrs

Test vessels and test concentration	24 Hours	48 Hours	72 Hours
Control
Replicate1	12800	54400	70000
Replicate2	14800	55200	69200
Replicate3	12000	54800	67600
CAS No. 12225-21-7
6.25mg/l
Replicate1	10800	49600	57200
Replicate2	11600	49200	60400
12.5mg/l
Replicate1	11600	50400	57600
Replicate2	12800	52000	61600
25mg/l
Replicate1	14000	52400	61600
Replicate2	13200	50400	58000
50mg/l
Replicate1	12800	49600	58400
Replicate2	12400	52400	63600
100mg/l
Replicate1	12000	50400	66400
Replicate2	14400	50000	62800
200mg/l
Replicate1	11600	53200	64800
Replicate2	11600	44000	66000

  

Table 2: Showing the values of average specific growth rate and percentage inhibition

after an interval of 72 hours

	CONTROL		6.25mg/l		12.5mg/l		25mg/l		50mg/l		100mg/l		200mg/l
Average Specific Growth rate (μ)	R1	0.648	R1	0.518	R1	0.583	R1	0.606	R1	0.588	R1	0.631	R1	0.622
	R2	0.644	R2	0.599	R2	0.606	R2	0.585	R2	0.616	R2	0.612	R2	0.629
	R3	0.637
Mean of Avg. Specific growth rate	0.643		0.590		0.594		0.595		0.602		0.621		0.625
Percentage Inhibition (%I)	_		8.250		7.560		7.380		6.375		3.377		2.722

 
Table 3: Depicting pH values at 0 Hours and after 72 Hours of test item exposure to algae

Test vessels and test concentration	0 Hours	72 Hours
CONTROL
Replicate1	6.84	6.68
Replicate2	6.75	6.64
Replicate3	6.75	6.4
Average	6.78	6.59
CAS No.12225-21-7
6.25mg/l
Replicate1	6.72	6.46
Replicate2	6.69	6.43
12.5mg/l
Replicate1	6.68	6.38
Replicate2	6.68	6.36
25mg/l
Replicate1	6.58	6.44
Replicate2	6.62	6.42
50mg/l
Replicate1	6.64	6.43
Replicate2	6.68	6.48
100mg/l
Replicate1	6.62	6.45
Replicate2	6.62	6.43
200mg/l
Replicate1	6.60	6.43
Replicate2	6.64	6.42

 

Validity criteria fulfilled:

yes

Conclusions:

After 72 hours of exposure to Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex(CAS No12225-21-7) to various nominal test concentration, EC50 calculated from equation through probit analysis was found to be >200 mg/L.

Executive summary:

The effect of test item Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1Hpyrazole-3-carboxylic acid complex, (CAS No. 12225-21-7) was studied on the growth of fresh water green alga Chlorella vulgaris. The study was conducted following OECD guideline 201- Alga, growth inhibition test. The test concentration chosen for the study were 6.25 mg/L,12.5 mg/L,25 mg/L,50 mg/L,100 mg/L,200 mg/L. The test concentrations were prepared using stock solution of the test item using mineral media. The green alga was exposed to the test concentration for a period of 72 hours to observe average specific growth rate and % growth inhibition under the effect of the test item. EC50 calculated graphically through probit analysis was observed to be >200 mg/L. It can be concluded from the value that the Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1Hpyrazole-3-carboxylic acid complex, (CAS No. 12225-21-7) is not toxic to the aquatic algae and can be considered as "not classified" as per the classifiecation criteria for aquatic environment.

Description of key information

The effect of test item Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1Hpyrazole-3-carboxylic acid complex, (CAS No. 12225-21-7) was studied on the growth of fresh water green alga Chlorella vulgaris. The study was conducted following OECD guideline 201- Alga, growth inhibition test. The test concentration chosen for the study were 6.25 mg/L,12.5 mg/L,25 mg/L,50 mg/L,100 mg/L,200 mg/L. The test concentrations were prepared using stock solution of the test item using mineral media. The green alga was exposed to the test concentration for a period of 72 hours to observe average specific growth rate and % growth inhibition under the effect of the test item. EC50 calculated graphically through probit analysis was observed to be >200 mg/L. It can be concluded from the value that the Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1Hpyrazole-3-carboxylic acid complex, (CAS No. 12225-21-7) is not toxic to the aquatic algae and can be considered as "not classified" as per the classifiecation criteria for aquatic environment.

Key value for chemical safety assessment

EC50 for freshwater algae:

200 mg/L

Additional information

Various studies including experimental data following OECD guideline for toxicity to aquatic algae with the target substance Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex(CAS No12225-21-7) were summarized as follows:

Data is from UERL 2017, exeprimental report followed standard gudeline suggests that The effect of test item Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1Hpyrazole-3-carboxylic acid complex, (CAS No. 12225-21-7) was studied on the growth of fresh water green alga Chlorella vulgaris. The study was conducted following OECD guideline 201- Alga, growth inhibition test. The test concentration chosen for the study were 6.25 mg/L,12.5 mg/L,25 mg/L,50 mg/L,100 mg/L,200 mg/L. The test concentrations were prepared using stock solution of the test item using mineral media. The green alga was exposed to the test concentration for a period of 72 hours to observe average specific growth rate and % growth inhibition under the effect of the test item. EC50 calculated graphically through probit analysis was observed to be >200 mg/L. It can be concluded from the value that the Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1Hpyrazole-3-carboxylic acid complex, (CAS No. 12225-21-7) is not toxic to the aquatic algae and can be considered as "not classified" as per the classifiecation criteria for aquatic environment.

Based on another study report,whose aim of the study was to evaluate the nature of chemical test chemical when comes in contact with the test organism Desmodesmus subspicatus (previous name: Scenedesmus subspicatus). Test was conducted according to the OECD guideline 201. The stock solution 130.0 mg/l was prepared by dissolving dark yellow powder in OECD growth medium. The test solution was kept 30 min in ultrasonic bath. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture. 0,2.6,5.6,12,27,59,130 mg/l concentration were used.With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Effect on the growth of algae was determine after an exposure period of 72 hrs.The median effective concentration (EC50) for the test substance Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex(CAS No12225-21-7), in algae was determined to be 74.2 mg/l (95% CI:64.0 - 86.0).on the basis of growth rate inhibition effects in a 72 hour study. Based on the EC50 value, which indicates that the substance is likely to be hazardous to aquatic algae and can be classified as aquatic chronic 3 as per the CLP classification criteria.

Even though varied results for short term toxicity of aquatic algae were available but, based on the maximum notifiers in ECHA and the effect concentration from first experimental study report,we can consider that the test chemical is not toxic and can be classified as not classified as per the classifiecation criteria for aquatic environment.