Reaction mass of orthophosphoric acid and potassium dihexadecyl phosphate and dipotassium hexadecyl phosphate

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From August 04, 2017 to October 30, 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

not specified

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

other: activated sludge, domestic, pre-conditioned to the experimental conditions

Details on inoculum:

Test organisms
Activated sludge was obtained from Totnes Sewage Treatment Works, Totnes, Devon, UK on 23 August 2017. This works treats sewage of predominantly domestic origin. At the laboratory, the activated sludge was kept aerated at room temperature and the pH maintained at 7.0 ± 1.0. Six days prior to the exposure start the activated sludge was centrifuged, washed and re suspended in the mineral medium and the solids concentration determined. This sludge was then diluted in medium, added to test bottles and stirred until required for use. The seeded mineral medium was pre-conditioned for six days to reduce the blank oxygen uptake readings in the test.

Duration of test (contact time):

ca. 60 d

Initial conc.:

ca. 100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

Test apparatus
The measurement of oxygen uptake was conducted in the Oxitop™ respirometers (Wissenschaftlich-Technische Werkstätten, GmbH, Germany). Each individual unit consisted of a dark glass 500 mL bottle with an OxitopÔbottle top containing a piezoresistive electronic pressure sensor. Bottles were situated on a magnetic stirrer in a constant temperature incubator. Carbon dioxide produced by microbial respiration was absorbed by potassium hydroxide solution (KOH) placed in a seal cup in the neck of each bottle, and the oxygen taken up was measured as a decrease in pressure. The Oxitop™ controller collected the pressure values from the measuring tops and calculated the BOD (as mg/L).

Reference substance:

benzoic acid, sodium salt

Remarks:

100 mg/L

Test performance:

Validity Criteria
The validity requirements of the OECD guideline state:
1) The difference between extremes of replicate biodegradation values should be less than 20% at the end of 10-day window, at the plateau or at the end of the test;
2) The positive control should achieve > 60% biodegradation by Day 14.
3) The oxygen consumption of the inoculum blank should not exceed 60 mg/L in 28 days.

Mean oxygen uptake of the inoculum blank was below 60 mg/L, as required in the OECD guideline. The difference between replicate test substance extremes was < 20% after the 10-d window, at plateau, and on Day 60. Sodium benzoate reached a mean 74% biodegradation by Day 14. The average of the mean oxygen consumed in the inoculum blanks was 6.9 mg/L after 28 d and 8.4 mg/L after 60 d. Therefore, this test has satisfied all the validity criteria.

Key result

Parameter:

% degradation (O2 consumption)

Remarks:

Test substance

Value:

ca. 40

Sampling time:

28 d

Remarks on result:

other: not readily biodegradable, but considered to be inherently biodegradable

Key result

Parameter:

% degradation (O2 consumption)

Value:

ca. 64

Sampling time:

40 d

Remarks on result:

other: inherently biodegradable

Key result

Parameter:

% degradation (O2 consumption)

Remarks:

Reference substance

Value:

ca. 74

Sampling time:

14 d

Remarks on result:

other: >60% degradation confirmed that the activated sludge contained viable organism

Key result

Parameter:

% degradation (O2 consumption)

Remarks:

Toxicity control

Value:

ca. 40

Sampling time:

14 d

Remarks on result:

other: >25% degradation confirmed that the test substance is assumed not to be inhibitory at used concentration

Details on results:

pH and temperature measurements
At the end of the 60-d test period, the pH values were 7.3 in the inoculum blank bottles, ranged from 6.7 to 7.0 in the sodium benzoate bottles, from 6.9 to 7.0 in the test substance bottlesand from 6.7 to 6.9 in the toxicity control bottles. Temperature measurements recorded in several of the test bottles on Days 0 and 60 indicated the temperature was within the range 22 ± 1°C. Continuous monitoring of the incubator temperature showed it to have remained within the range 22 ± 1°C throughout the study.

Results with reference substance:

Theoretical oxygen demand (ThOD)
The ThOD of sodium benzoate was calculated as 1.67 g O2/g of substance, using the formula given in the OECD guideline. Biodegradation of sodium benzoate has been calculated on the basis of calculated ThOD.

Biochemical oxygen demand (BOD)
The results for the reference substance did show that sodium benzoate attained a mean level of biodegradation (based on the BOD:ThOD ratio) of 86%, and the results showed good replication. More than 60% degradation was achieved within the 10-day window as expected for a biodegradable substance, thus confirming that the activated sludge contained viable organisms.

Results

Theoretical oxygen demand (ThOD)

The ThOD of sodium benzoate was calculated as 1.67 g O₂/g of substance, using the formula given in the OECD guideline. Biodegradation of sodium benzoate has been calculated on the basis of calculated ThOD.

Chemical oxygen demand (COD)

The measured COD value of test substance was 1.50 g O₂/g of substance, and its biodegradation was calculated on the basis of measured COD.

Biochemical oxygen demand (BOD)

The results for the reference substance did show that sodium benzoate attained a mean level of biodegradation (based on the BOD:ThOD ratio) of 86%, and the results showed good replication. More than 60% degradation was achieved within the 10-d window as expected for a biodegradable substance, thus confirming that the activated sludge contained viable organisms.

Biodegradation results – Reference Substance

Day	Sodium benzoate (blank corrected) BOD (mg/L)			BOD^a(g O2/g)			Biodegradation^b(%)
	Rep 1	Rep 2	Rep 3	Rep 1	Rep 2	Rep 3	Rep 1	Rep 2	Rep 3	Mean
7	96.5	119	122	0.97	1.19	1.22	58	71	73	67
14	109	131	134	1.09	1.31	1.34	65	78	80	74
21	109	137	135	1.09	1.37	1.35	65	82	81	76
28	114	136	136	1.14	1.36	1.36	68	81	81	77
35	116	141	141	1.16	1.41	1.41	69	84	84	79
42	115	143	140	1.15	1.43	1.40	69	86	84	80
49	114	148	145	1.14	1.48	1.45	68	89	87	81
56	110	152	152	1.10	1.52	1.52	66	91	91	83
60	116	158	155	1.16	1.58	1.55	69	95	93	86

BOD:   Biological Oxygen Demand

^a       BOD (g O₂/g) =Blank corrected biodegradation (mg/L)         

                  Sodium benzoate concentration (100 mg/L)

^b  Biodegradation (%) =Biological Oxygen Demand (g O₂/g)× 100

                                                ThOD (1.67 g O₂/g)

The following phases (approximate) were observed:

Phase	Time period
Lag phase	Days 0 to 1
Degradation phase	Days 1 to 18
10-day window	Days 1 to 11

 

The results for test substance did show that test substance attained a mean level of biodegradation (based on the BOD:COD ratio) of 40% after 28 d and 64% after 60 d, and the results showed good replication. Less than 60% degradation was achieved within 28 d, so test substance cannot be classified as readily biodegradable.

Biodegradation results – Test substance

Day	Test substance (blank corrected) BOD (mg/L)			BOD^a(g O2/g)			Biodegradation^b(%)
	Rep 1	Rep 2	Rep 3	Rep 1	Rep 2	Rep 3	Rep 1	Rep 2	Rep 3	Mean
7	9.3	14.9	14.9	0.09	0.15	0.15	6	10	10	9
14	35.3	32.5	35.3	0.35	0.33	0.35	23	22	23	23
21	50.7	47.9	50.7	0.51	0.48	0.51	34	32	34	33
28	60.6	55.0	60.6	0.61	0.55	0.61	41	37	41	40
35	70.6	65.0	65.0	0.71	0.65	0.65	47	43	43	44
42	80.7	72.3	72.3	0.81	0.72	0.72	54	48	48	50
49	88.5	80.0	77.2	0.89	0.80	0.77	59	53	51	54
56	95.9	87.4	87.4	0.96	0.87	0.87	64	58	58	60
60	102	92.6	92.6	1.02	0.93	0.93	68	62	62	64

BOD:   Biological Oxygen Demand

^a       Biological oxygen demand (g O₂/g) =Blank corrected biodegradation (mg/L)

                                                      Substance concentration (100 mg/L)

^b  Biodegradation (%) =Biological Oxygen Demand (g O₂/g)× 100

                                                COD (1.50 g O₂/g)

The following phases (approximate) were observed:

Phase	Time period
Lag phase	Days 0 to 7
Degradation phase	Days 7 to 49

Combined ThOD/COD for toxicity control

The mean toxicity control degradation achieved on Day 14 was 40% (based on combined ThOD/COD), as this was > 25% the test substance was assumed not to be inhibitory at this concentration.

Biodegradation results – Toxicity Control

Day	Toxicity control (blank corrected) BOD (mg/L)			BOD^a(g O2/g)			Biodegradation^b(%)
	Rep 1	Rep 2	Rep 3	Rep 1	Rep 2	Rep 3	Rep 1	Rep 2	Rep 3	Mean
7	111	105	108	0.56	0.53	0.54	35	33	34	34
14	125	117	131	0.63	0.59	0.66	40	37	42	40
21	137	126	143	0.69	0.63	0.72	43	40	45	43
28	148	139	153	0.74	0.70	0.77	47	44	48	46
35	161	147	161	0.81	0.74	0.81	51	47	51	50
42	168	157	171	0.84	0.79	0.86	53	50	54	52
49	173	170	176	0.87	0.85	0.88	55	53	55	54
56	180	180	183	0.90	0.90	0.92	57	57	58	57
60	186	186	186	0.93	0.93	0.93	58	58	58	58

BOD:   Biological Oxygen Demand

^a       Biological oxygen demand (g O₂/g) =Blank corrected biodegradation (mg/L)

                                                      Toxicity control concentration (200 mg/L)

^b  Biodegradation (%) =Biological Oxygen Demand (g O₂/g)× 100

                                    Combined ThOD/COD (1.59 g O₂/g)

Validity criteria fulfilled:

yes

Interpretation of results:

inherently biodegradable

Conclusions:

Under study conditions, the test substance was considered to be inherently biodegradable.

Executive summary:

A study was conducted to determine the ready biodegradability of the test substance, 'mono- and di- C16 PSE, K+ and H3PO4' (purity: 100%), using Manometric respirometry method, according to OECD Guideline 301F, in compliance with GLP. Following the pre-conditioning, the test bottles containing prepared activated sludge inoculum in mineral medium were set up for the test according to the experimental design. In the study, Inoculum blanks, positive control (sodium benzoate at 100 mg/L), test substance (100 mg/L) as well as toxicity controls (test and reference substances, both at 100 mg/L), each these group bottles were prepared in triplicate, except inoculum blanks which were prepared in duplicate. Additional single replicate test substance and toxicity control bottles were prepared for determination of pH at Day 0, so undissolved test substance was not removed by the pH probe. Oxygen uptake was recorded automatically every 240 minutes during the 60 d experimental period. Oxygen uptake values were corrected for the inoculum blank and the biodegradation was calculated as a percentage of the chemical oxygen demand (COD) for the substance under test and as a percentage of the theoretical oxygen demand (ThOD) for the reference substance. The chemical oxygen demand (COD) of test substance was determined using spectrophotometer for evaluation. The test substance attained a mean level of biodegradation (based on the BOD:COD ratio) of 40% after 28 d and 64% after 60 d. The test substance results showed good replication. Less than 60% degradation was achieved within 28 d, so test substance was classified as inherently biodegradable. Sodium benzoate attained a mean level of biodegradation (based on the BOD:ThOD ratio) of 86%, and the results showed good replication. More than 60% degradation was achieved within the 10-d window as expected for a biodegradable substance, thus confirming that the activated sludge contained viable organisms. The mean toxicity control degradation achieved on Day 14 was 40% (based on combined ThOD/COD), as this was > 25% the test substance was assumed not to be inhibitory at this concentration. The test met all the validity criteria. Under the study conditions, the test substance was considered to be inherently biodegradable (Scymaris, 2017).

Description of key information

Based on the study results, the test substance was determined to be inherently biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

inherently biodegradable

Additional information

A study was conducted to determine the ready biodegradability of the test substance, 'mono- and di- C16 PSE, K+ and H3PO4' (purity: 100%), using Manometric respirometry method, according to OECD Guideline 301F, in compliance with GLP. Following the pre-conditioning, the test bottles containing prepared activated sludge inoculum in mineral medium were set up for the test according to the experimental design. In the study, Inoculum blanks, positive control (sodium benzoate at 100 mg/L), test substance (100 mg/L) as well as toxicity controls (test and reference substances, both at 100 mg/L), each these group bottles were prepared in triplicate, except inoculum blanks which were prepared in duplicate. Additional single replicate test substance and toxicity control bottles were prepared for determination of pH at Day 0, so undissolved test substance was not removed by the pH probe. Oxygen uptake was recorded automatically every 240 minutes during the 60 d experimental period. Oxygen uptake values were corrected for the inoculum blank and the biodegradation was calculated as a percentage of the chemical oxygen demand (COD) for the substance under test and as a percentage of the theoretical oxygen demand (ThOD) for the reference substance. The chemical oxygen demand (COD) of test substance was determined using spectrophotometer for evaluation. The test substance attained a mean level of biodegradation (based on the BOD:COD ratio) of 40% after 28 d and 64% after 60 d. The test substance results showed good replication. Less than 60% degradation was achieved within 28 d, so test substance was classified as inherently biodegradable. Sodium benzoate attained a mean level of biodegradation (based on the BOD:ThOD ratio) of 86%, and the results showed good replication. More than 60% degradation was achieved within the 10-d window as expected for a biodegradable substance, thus confirming that the activated sludge contained viable organisms. The mean toxicity control degradation achieved on Day 14 was 40% (based on combined ThOD/COD), as this was > 25% the test substance was assumed not to be inhibitory at this concentration. The test met all the validity criteria. Under the study conditions, the test substance was considered to be inherently biodegradable (Scymaris, 2017).

[Type of water: freshwater]