4-phenylbenzophenone

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

7th July 2017 to 15 December 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test (Carbon and Ammonium Oxidation))

Version / remarks:

Adopted 22 July 2010

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Appearance: white to off white crystaline powder
Batch number: 20161118
Purity: 99.74%
Storage: at room temperature protected from the light
Expiry date: 17 November 2017
Indentifier name: Omnirad 4-PBZ
CAS number: 2128-93-0
Solubility: 0.0736 mg/L (determined in other Charles Rier Laboratories Den Bosch study)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

TEST SUBSTANCE SOLUTIONS
- Direct weighing of the test substance into the test vessels was carried out. The weights are detailed below in Table 1.
- Procedure: The weighed amount was added to a dark brown test bottle containing 200ml of Milli-RO water (tap water purified by reverse osmosis). For the lowest concentration, the test item was weighed on a watch glass and the watch glass transferred to the vessel with the test substance. The mixtures were stirred for a short period and then 16 mL synthetic medium made up to 50 mL with Milli-RO water and 250 mL sludge (based on a determined 3.0g/L concentration of suspended solids) were added resulting in the required loading rates. Optimal contact between the test item and test organisms was ensured by applying continuous aeration and stirring.
REFERENCE SUBSTANCE SOLUTIONS:
- Reference substance: 3,5-dichlorophenol
- Preparation: 1.0 g/L stock solution prepared by dissolving 250 mg dichlorophenol in Milli-RO water and making up to 250 ml.
- pH of stock: 7.8
- Storage of stock: freezer
- Test concentrations: 0.5, 1.6, 5.0, 16ml of stock were added to test medium to provide a final test concentration of 1.0, 3.2, 10 and 32mg/L respectively.

Test organisms (species):

activated sludge

Details on inoculum:

Source of activated sludge: Municipal sewage treatment plant: 'Waterschap Aa en Maas', 's- Hertogenbosch, The Netherlands, receiving predominantly domestic sewage.
Preparation of sludge: the sludge was seived (1mm) and allowed to settle. The supernatant was removed and ISO medium added.
pH of sludge: 7.6
Storage of sludge: the sludge was stored overnight prior to use. 50ml of synthetic medium was provided to the sludge and it was aerated overnight at room temperature.
Suspended solids: detemined to be 3.0g/L

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Remarks on exposure duration:

None

Post exposure observation period:

None

Hardness:

Not stated

Test temperature:

Ranged between 20 and 21°C during the test.

pH:

7.5 ± 0.5

Dissolved oxygen:

60-70% at the start (equivalent to >5mg/L at 20˚C)

Salinity:

Not stated

Conductivity:

Not stated

Nominal and measured concentrations:

Nominal concentrations: 3.2 (T1), 10 (T2), 32 (T3), 100 (T4), 1000 (T5) and 1000 mg/L (T6).

Details on test conditions:

Test medium: Adjusted ISO medium, formulated using RO water with 211.5 mg/L CaCl2.2H20, 88.8mg/L MgSO4.7H2O, 46.7 mg/L NaHCO3, 4.2mg/L KCl.
Synthetic medium (sewage feed): 16g peptone, 11g meat extract, 3g urea, 0.7g NaCl, 0.4g CaCi2.2H2O, 0.2g MgSO4.7H2O, 2.8g K2HPO4. All dissolved in Milli RO water and made up to 1 litre and filtered. The pH was 7.5± 0.5.
Replication: 5 replicates per test group and 6 replicates for the control.
Controls: Test medium without test item, treated in the same way as the test item solutions.
Aeration: vessels were aerated and stirred during the 3 hours. Aeration was determined at 60-70% saturation at the start of the experiment, and was adequate to maintain flocs in suspension.
Measurement of parameters: Oxygen consumption was recorded for 10 minutes after the 3-hour contact time. pH was determined in all test and reference item vessels. Temperature was recorded continuously in control vessels. Determination of oxygen was performed with multiple oxygen sensors connected to a BlueBox, a multichannel measuring and controlling system.

Reference substance (positive control):

yes

Remarks:

3,5-dicholorophenol (see below for further details on reference item)

Key result

Duration:

3 h

Dose descriptor:

other: NOELR

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Key result

Duration:

3 h

Dose descriptor:

other: ELR50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Details on results:

Table 3 details the respiration results and inhibition of respiration in each of the different exposure treatments and the controls in the final test. There was no statistically significant inhibition of the respiration rate at any of the loading rates.

It should also be noted that the results for the combined limit/range finding test were slightly different. These results demonstrated an inhibition of respiration rate of 10, 26 and 25% for 10, 100 and 1000 mg/L concentrations respectively. No clear dose response curve was seen. There was no oxygen uptake from the abiotic control.

The sensitivity of the activated sludge was slightly higher in the combined limit/range-finding test. The EC50 for 3,5-dichlorophenol was 3.8 mg/L in the combined limit/range-finding and 10 mg/L in the final test. This could be the explanation for the slightly lower effects observed in the final test.

Results with reference substance (positive control):

The Ec50 for 3,5-dichlorophenol was 10mg/L.

Reported statistics and error estimates:

BlueBox Software version 3.4.2.0 was used for system control, data aquisition and processing. The system was used to calculate oxygen uptakes and inhibition of the respiration rate.

Table 3. Overview of the results for the final test.

Treatment	Loading rate (mg/L)	Mean respiration rate		% Inhibition of the respiration rate (mean value)
		(mg O2/L h)	(mg O2/g h)¹
Control		50.25	33.50
T1	3.2	48.79	32.53	2.89
T2	10	46.68	31.12	7.11
T3	32	47.77	31.85	4.93
T4	100	55.44	36.96	-10.33
T5	320	49.93	33.29	0.63
T6	1000	46.21	30.81	8.04

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of this present test, 4-Phenyl benzophenone (Omnirad 4-PBZ) was not toxic to waste water bacteria (activated sludge) at or below a loading rate of 1000 mg/L (NOELR). The ELR50 was >1000 mg/L.

Executive summary:

A reliable, Klimisch 1 study to assess the inhibition of respiration for activated sludge exposed to 4-Phenyl benzophenone (Omnirad 4-PBZ) was carried out, according to the OECD Guideline No. 209 (2010). The study was conducted according to GLP. A combined range limit/range finding test was carried out at 10, 100 and 1000 mg/L. A maximum of 26% inhibition of respiration was determined in the combined limit/range finding test. In the final test, six nominal loading were tested concentrations: 3.2, 10, 32, 100, 1000, and 1000 mg/L. Due to the very low solubility of the test substance, direct weighing was used. The reference item 3,5 -dichlorophenol was used to assess the reliability of the test, with an EC50 of 10mg/L in the final test, which is considered within normal the normal range. In the final test, the maximum inhibition of respiration was 8 %, and no significant difference between the exposed and control vessels were determined. The No observed effect loading rate (NOELR) was determined at >1000mg/L.

Description of key information

A reliable 3 hour toxicity study with bacteria from an activated sludge treatment plant was conducted in accordance with the OECD test guideline 209. Under the conditions of this present test, no toxicity was found to the waste water bacteria (activated sludge) at or below a loading rate of 1000 mg/L (NOELR). The ELR50 was >1000 mg/L.

Key value for chemical safety assessment

EC50 for microorganisms:

1 000 mg/L

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

A reliable, Klimisch 1 study to assess the inhibition of respiration for activated sludge exposed to 4-Phenyl benzophenone (Omnirad 4-PBZ) was carried out, according to the OECD Guideline No. 209 (2010). The study was conducted according to GLP. A combined range limit/range finding test was carried out at 10, 100 and 1000 mg/L. A maximum of 26% inhibition of respiration was determined in the combined limit/range finding test. In the final test, six nominal loading were tested concentrations: 3.2, 10, 32, 100, 1000, and 1000 mg/L. Due to the very low solubility of the test substance, direct weighing was used. The reference item 3,5 -dichlorophenol was used to assess the reliability of the test, with an EC50 of 10mg/L in the final test, which is considered within normal the normal range. In the final test, the maximum inhibition of respiration was 8 %, and no significant difference between the exposed and control vessels were determined. The No observed effect loading rate (NOELR) was determined at >1000mg/L.