Isooctyldinitrophenol

Toxicity to microorganisms

Administrative data

Endpoint:

activated sludge nitrification inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

8th August 1985 - 14th August 1985.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The undersigned declares that this work was carried out under supervision according to the procedures herein described and according to the OECD principles of Good Laboratory Practice, and that this report represents a true and accurate record of the results obtained.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1985

Report date:

1985

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: ETAd method for the Determination of the Inhibitory Effects of Chemicals and industrial Waste Waters on the Respiration and Nitrification of Activated Sludge

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Isooctyldinitrophenol

EC Number:

253-410-9

EC Name:

Isooctyldinitrophenol

Cas Number:

37224-61-6

Molecular formula:

C14H20N2O5

IUPAC Name:

4-(6-methylheptyl)-3,5-dinitrophenol

Test material form:

liquid: viscous

Details on test material:

Test substance was received from Thomas Swan and Co. Ltd., Crookhall, Consett, Durham, DH8 7ND on 30th May 1985. Substance was a brown, viscous, liquid contained in a white opaque plastic bottle with a screw cap.

Inclusive weight of the test substance and container on receipt was 492.03g. The test substance was stored under ambient temperature until required.

Chemical name: Dinitro Capryl Phenol
Other names: Dinitro octyl phenol
Molecular formula: C14H20N2O5
Chemical composition: DNCP - 89.9%, MNOP - 4.6%, NDOP - 4.0%, TNP - 1.0%
Appearance, colour: Dark red
Solubility in water: 30mg/l
Specific gravity: 1.13
Stability: stable, instability caused by overheating
Intended use: Fungicide

Analysis report for the sample of D.N.C.P supplied for use in this test by the sponser is as follows:

DNCP P511 B68/85
% H20 - 0.08
% DNCP - 89.9
% MNCP - 4.55
% strong acid as Picric Acid - 0.53
Total titratable phenols (meq/grm) - 3.24

Test solutions

Details on test solutions:

Reference Inhibitor: 3,5-Dichlorophenol (3,5-DCP)
Nitrification Inhibitor: Allyl Thiourea (ATU)

Test organisms

Test organisms (species):

activated sludge

Details on inoculum:

Synthetic sewage:

A 100-fold concentrated solution of OECD/EEC synthetic sewage was prepared by dissolving the following in one litre of deionised water:

27.00g 'Lan-Lemco' Broth
3.00g Urea
2.80g Dipotassium hydrogen orthphosphate
0.70g Sodium Chloride
0.40g Calcium chloride dihydrate
0.20g Magnesium sulphate heptahydrate

The solution was kept ina refrigerator at 4°C until required.

Study design

Test type:

not specified

Water media type:

not specified

Total exposure duration:

3 h

Test conditions

Test temperature:

Preliminary Test: 22°C
Definitive Test: 22°C

Nominal and measured concentrations:

nominal: 100mg/l

Results and discussion

Effect concentrations

Duration:

1 h

Dose descriptor:

EC50

Effect conc.:

> 30 - < 137.1 mg/L

Conc. based on:

test mat.

Basis for effect:

inhibition of nitrification rate

Remarks on result:

other: 95% CL

Details on results:

Preliminary test

The concentration of suspended solids in the sample of activated sludge collected for the preliminary test was estimated to be 4944 mg/l. 3034 ml of sludge was made up to 5000ml by the addition of groundwater to produce a sludge with a nominal concentration of suspended solids of 3000 mg/l.

The results of the preliminary test are shown in table 1. There were no ihibitory effects on oxygen consumption at nominal test substance concentrations of 1 and 10mg/l. However, there was evidence to suggest that the test substance stimulated oxygen consumption at these concentrations (Bottles 12 and 14). A 52% inhibition was noted at a nominal test substance concentration of 100mg/l (Bottle 16).

In bottles 13, 15 and 17 containing the test substance and ATU, oxygen consumption decreased with increasing conc. of the test substance. The maximum inhibition observed in these mixtures (69%, 100 mg/l DNCP) did not exceed that in the control + ATU mixtures (50% to 70%, Bottles 2 an 11).

The EC50 of the test substance to bacteria using oxygen in the sludge was estimated to be approx. 100 mg/l.

Oxygen was not consumed by the abiotic control mixture.

The EC50 of the reference inhibitor, 3,5-DCP was estimated to be between 1 and 10mg/l. A 100% inhibition of oxygen consumption was noted in the mixture containing 100mg/l of 3,5-DCP + ATU.

Definitive Test

The concentration of suspended solids in the sample of activated sludge collected fpr the second test was estimated to be 4548mg/l. 3300ml of sludge were made up to 5000 mg/l by the addition of groundwater to produce a sludge with a nominal concentration of suspended solids of 3000 mg/l.

The results of the definitive test are shown in table 2. inhibtion of oxygen consumption was observed at each of the three test substance concentrations tested (50, 100 and 150 mg/l). The mean percentage inhibition at each concentration was 5%, 29& and 42% respectively. The EC50 of the test substance to bacteria using oxygen in the sludge was estimated to be >150mg/l.

As with the preliminary test, the maximum inhibition by the test substance (mean of 42% at 150 mg/l) did not exceed that observed in the control + ATU mixture (mean of 44%).

the EC50 of the test substance to bacteria involved in nitrification was estimated to be 91.5mg/l. The 95% confidence limits were calculated to be 30.1 mg/l (lower limit) and 137.1 mg/l (upper limit)

100mg/l of 3,5-DCP inhibited oxygen consumption by approx. 96%.

Any other information on results incl. tables

Table 1: Preliminary test: Rates of oxygen consumption of each test mixture with % inhibtion by the test substance, 3,5 -dochlorophenol and Allyl thiourea.

Bottle No.	Bottle Contents	Rate of O2 consumption (mg O2/l/hr)	% inhibition
1	Control	36.1	-
9	Control	23.5	-
10	Control	38.3	-
	Mean	32.7	-
3	1 mg/l 3,5 -DCP	21.6	34
5	10mg/l 3,5 -DCP	4.2	87
7	100mg/l 3,5 -DCP	3.9	88
12	1mg/l DNCP	40.9	-25
14	10mg/l DNCP	40.6	-24
16	100mg/l DNCP	15.7	52
2	Control + ATU	9.1	72
11	Control + ATU	15.7	52
4	1mg/l 3,5 -DCP + ATU	7.9	76
6	10mg/l 3,5 -DCP + ATU	8.5	74
8	100mg/l 3,5 -DCP + ATU	0	100
13	1mg/l DNCP + ATU	18.5	43
15	10mg/l DNCP + ATU	14.1	57
17	100mg/l DNCP + ATU	10.2	69
18	100mg/l DNCP (Abiotic control)	0	100

Table 2: Definitive test: Rates of oxygen consumption of each test mixture with the % inhibition by the test substance, 3,5 -Dichlorophenol and Allyl thiourea

Bottle No.	Bottle Contents	Rate of O2 consumption (mg O2/l/hr)	% inhibition	% inhibition of Nitrification
1	Control	42.1	-	-
7	Control	41.2	-	-
12	Control	41.9	-	-
	Mean	41.7
2	Control + ATU	22.5	46	-
8	Control + ATU	23.7	43	-
13	Control + ATU	23.7	43	-
	Mean	23.3	44	-
3	100mg/l 3,5 -DCP	1.7	96	-
4	50mg/l DNCP	34.2	18	41
11	50mg/l DNCP	41.1	1	1
15	50mg/l DNCP	44.0	-5	-12
	Mean	39.8	5	10
5	100mg/l DNCP	25.9	38	83
9	100mg/l DNCP	23.7	43	100
16	100mg/l DNCP	39.5	5	13
	Mean	39.8
6	150mg/l DNCP	17.0	59	128
10	150mg/l DNCP	21.9	47	110
14	150mg/l DNCP	33.3	20	47
	Mean	24.1	42	96

Applicant's summary and conclusion

Conclusions:

The test substance was found not to inhibit oxygen consumption by activated sludge at nominal test substance concentrations of 50, 100 and 150 mg/l. In both the preliminary and definitive test the extent of this inhibtion did not exceed that caused by the ATU (the known inhibitor of nitrification). Therefore, it would seem likely that the test substance was inhibitory to to nitrification and not carbonaceous respiration at the concentrations tested. In view of this finding the percentage inhibition of nitrification by the test substance in mixtures prepared in the definitive test was calculated. The EC50 of DNCP to bacteria involved in nitrification was estimated to be 91.5mg/l. This value had a lower 95% confidence limit of 30.0mg/l and an upper limit of 137.1mg/l.