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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:

1995-06-19 to 1995-12-13

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 C (Ready Biodegradability: Modified MITI Test (I))

Qualifier:

according to guideline

Guideline:

other: Testing Methods for New Chemical Substances (July 13 1974, Kanpoygo No.5, Planning and Coordination Bureau, Environment Agency, Yakuhutu No.615, Pharmaceutical Affairs Bureau, Ministry of Health and Welfare, and 49 Kikyoku No393, Basic Industries Bureau

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

Sludge sampling sites and date:
sampling site: On-site sampling was carried out at the following 10 locations in Japan:
- Fukogawa city sewage plant (Sapporo-shi Hokkaido)
- Fukashiba industry sewage plant (Kashima-gun Ibaragi)
- Nakahama city sewage plant (Osaka-shi Osaka)
- Ochiai city sewage plant (Shinjuku-ku Tokyo)
- Kitakami river (Ishinomaki-shi Miyagi)
- Shinano river (Nishikanbara-gun Niigata)
- Yoshino river (Tokushima-shi Tokushima)
- Lake Biwa (Otsu-shi Shiga)
-Hiroshima bay (Hiroshima-shi Hiroshima)
- Dookai bay (Kitakyushu-shi Fukoka)

Date: March 1995

Sludge sampling method:
City sewage: Return sludges from sewage plants were collected
Rivers, lake and sea: Surface water and surface soil which are in contact with the atmosphere were collected

Mixing of fresh and old activated sludge:
The filtrate (5 L) of the supernatant of the activated sludge used at that time was mixed all together with each of the filtrate (500 ml) of the supernatant of a newly collected sludge (10 sources). The mixture filtrate (10 L) was aerated sufficiently through prefiltered open-air after pH value of the mixture was adjusted to 7.0±1.0

Cultivation:
In about 30 minutes after aeration of the sludge mixture was ceased, supernatant corresponding to about one-third of the whole volume was removed. Then an equal volume of dechlorinated water was added to the remaining portion. This mixture was aerated and then a previously decided amount of synthetic sewage was added to the mixture so that the concentration of the synthetic sewage was 0.1 (w/v)% in the volume of the dechlorinated water added. This procedure was repeated once every day. The cultivation was carried out at 25±2°C.

Synthetic sewage:
Glucose, peptone and potassium dihydrogenphosphate were dissolved in dechlorinated water to obtain 5(w/v)% of the solution as to each components. The pH of the solution was adjusted to 7.0±1.0 with sodium hydroxide.

Control:
During the cultivation, the appearance of the supernatant, setting of the sludge, formation of flock, pH, dissolved oxygen concentration in the solution and temperature were checked and adjusted if necessary. Microflora in the activated sludge was microscopically observed and the sludge with no abnormal symptoms was used for the test.

Inspection of activity and date of initiation of use of activated sludge:
Inspection of activity:
Activity of the sludge was assessed by use of a reference substance and the relation between new and old activated sludges was taken into account.

Date of initiation of use: 1995-04-18

- Concentration of sludge: 30 mg/l

Duration of test (contact time):

28 d

Initial conc.:

100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

DOC removal

Parameter followed for biodegradation estimation:

TOC removal

Details on study design:

Preparation for the test:

Measurement of concentration of suspended solid: The concentration of the suspended solid in the activated sludge was measured on July 3rd 1995 to be 4800 mg/l

Preparation of basal culture medium: Each 3 ml of solution A, B, C and D as prescribed in the Japanese Industrial Standards (JIS) K 0102-1993-21, were made up to 1000 ml with purified water (Takasugi Seiyaku Co., Ltd.) and then the pH of the solution was adjusted to 7.0.

Reference substance:
Aniline (guaranteed reagent, Showa Chemicals Inc., Lot No. SE-31230) was used as a reference substance.

Preparation of test solutions:
The following test solutions were prepared and cultured under the conditions below:

1. Addition of test substance or aniline
a) The test solution (water + test substance) (n = 1, Vessel No. 3): In one test vessel, 30 mg of the test substance was added to 300 ml of purified water

b) The test solution (sludge + test substance) (n = 3, vessel No. 4, 5 and 6): In each test vessel, 30 mg of the test substance was added to the basal culture medium (300 ml) which includes previously decided amount of the activated sludge.

c) The test solution (sludge + aniline) (n = 1, vessel No. 1): In one test vessel, 29.5 µl (30.0 mg = 29.5 µl * 1.022 g/cm3 (density of aniline)) of aniline was added into the basal culture medium (300 ml) which includes previously decided amount of the activated sludge.

d) The test solution (control blank) (n = 1, vessel No. 2). In one test vessel, nothing was added to the basal culture medium (300 ml) which includes previously decided amount of the activated sludge.

- Instrument for cultivation:
Closed system oxygen consumption measuring apparatus (Coulometer: Ohkura Electric Co., Ltd.); (Data sampler: Asahi Instrument Industries Co., Ltd)
Vessel: 300 ml volume
Absorbent for carbon dioxide: Soda lime No.1 (for absorption of carbon dioxide, Wako Pure Chemical Industries, Ltd.)
Stirring method: Each test solution was stirred by a magnetic stirrer

- Conditions of cultivation:
Cultivation temperature: 25±1°C
Cultivation duration: 28 days
Room: Apparatus room No. 511 of Kurume Research Laboratories

2. Inoculation of activated sludge:
The activated sludge was added to each test vessel (b), (c) and (d) so that the concentration of the suspended solid reached 30 mg/l

Analysis of test solution:
After the termination of the cultivation, dissolved organic carbon and converted products methyltrihydroxysilane, propionic acid, methanol and 2-ethylhexylalcohol in the test solutions were determined.

Pretreatment of test solution for analysis:
After the termination of the cultivation, the test solution (water + test substance), the test solutions (sludge + test substance) and the test solution (control blank) were pretreated for the total organic carbon (TOC) analysis, high performance liquid chromatography (HPLC) analysis, gas chromatography (GC analysis).

Reference substance:

aniline

Key result

Parameter:

% degradation (TOC removal)

Value:

87

Sampling time:

28 d

Key result

Parameter:

other: BOD

Value:

69

Sampling time:

28 d

Details on results:

BOD (test substance): 26.6 % degradation after 7 d 45.3 % degradation after 14 d 58.6 % degradation after 21 d 68.6 % degradation after 28 d

Results with reference substance:

69% degradation after 7 d
76% degradation after 14 d
77% degradation after 21 d
76 % degradation after 28 d

Hydrolysis of the test substance in test solution (water + test substance):

It was presumed that the test substance is changed to methyltrihydroxysilane, propionic acid, methanol and 2-ethylhexylalcohol by hydrolysis. Therefore, the converted products were analysed. The produced amount of propionic acid was 20% of the theoretical amount, whereas the produced amount of the other compounds were equivalent to the theoretical amounts. Since propionic acid might be adsorbed on soda lime, the soda lime were washed with purified water and the propionic acid in the washing water was measured. 3.7 mg of propionic acid was detected in water. Percentage production of the total propionic acid in the test solution and on the soda lime was 80% of the theoretical amount.

Table 1: Calculation table for percentage biodegradation by BOD

Test No. 12595 Cultivating duration: 28 days
vessel No.	7^thday		14^thday		21^stday		28^thday		mean deg (%)
	BOD (may)	Deg (%)	BOD (may)	Deg (%)	BOD (may)	Deg (%)	BOD (may)	Deg (%)
1 [sludge + aniline]	64.4	69	73.5	76	76.0	77	76.2	76
2 [contol + blank]	2.4	-	4.6	-	6.8	-	7.2	-
4 [sludge + test substance]	26.3	35	39.0	51	49.5	63	58.0	75	69
5 [sludge + test substance]	7.0	7	15.7	16	34.1	40	45.4	56
6 [sludge + test substance]	28.4	38	51.9	69	56.4	73	58.0	75
3 [water + test substance]	0.0	-	0.0	-	0.9	-	1.1	-

 

 Table 2: Calculation table for percentage biodegradation by TOC

Sample description	Measured value (mgC/L)	Amount of DOC	% Biodegradation	Average % biodegradation
Water blank	n.d
[3] water + test substance	41.59	12.5
[4] sludge + test substance	9.49	2.4	86
[5] sludge + test substance	8.35	2.0	88	87
[6] sludge + test substance	9.35	2.3	86
[2] control blank	1.55

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

The biodegradation of the substance has been determined using a relevant test method and in compliance with GLP. The result is considered to be reliable.

Description of key information

Biodegradation in water, screening tests: 69% BOD in 28 days; 87% TOC in 28 days (OECD 301C). No significant biodegradation is expected for the silanol hydrolysis products.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

Biodegradation of 69% BOD in 28 days; 87% TOC in 28 days (OECD 301C) was determined in a reliable study conducted according to an appropriate test protocol, and in compliance with GLP.

 

In contact with water, the constituents of the submission substance hydrolysewithin the timescale of the ready biodegradation study to methylsilanetriol,2-ethylhexyl-3-[dihydroxy(methyl)silyl]propanoate,2-ethylhexyl propanoateand methanol.The biodegradation observed in the study is attributable to the biodegradation of themethanol and 2-ethylhexyl propanoate hydrolysis products.

 

Methanol is readily biodegradable (OECD 2004).

 

The non-silanol hydrolysis product, 2-ethylhexyl propanoate is expected to hydrolyse within the time scale of the ready biodegradation study to form 2-ethylhexanol and propionic acid. Both 2-ethylhexanol and propionic acid are readily biodegradable (EPA 2006, OECD 2007).

 

No significant biodegradation is expected for the silanol hydrolysis products.

 

References:

EPA (2006). Inert Reassessment: 2-ethyl-1-hexanol; CAS No. 104 -76 -7. Correction to the List Classification Determination Paragraph. United States Environmental Protection Agency, Washington, D.C. 20460. August 15, 2006.

 

OECD (2004): SIDS Initial Assessment Report for SIAM 19, Berlin, Germany, 18-20 October 2004, Methanol, CAS 67-56-1.

 

OECD (2007): SIDS Initial Assessment Report for SIAM 25, 17-18 October 2007, Propionic acid, CAS 79-09-4.