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Environmental fate & pathways

Biodegradation in water: screening tests

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biodegradation in water: ready biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
GLP - Guideline study. A maximal reliability score of Klimisch 2 could be assigned to this study in accordance with 'practical guide 6: How to report read-across and categories' which states that the maximum reliability for a read-across is 2. The relevant study was performed according to GLP and the methods applied are fully compliant with OECD TG 301F
Justification for type of information:
a detailled justification for the analogue approach can be found in Iuclid section 13 of the dossier.
Reason / purpose for cross-reference:
read-across source
Reason / purpose for cross-reference:
read-across: supporting information
according to guideline
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Concentration of test substance ca. 30 mg ThOD/L, instead of 50 - 100 mg ThOD/L as recommended in the guideline
GLP compliance:
yes (incl. QA statement)
Oxygen conditions:
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): collected from the oxidation ditch bioreactor at the Midland Municipal Wastewater Treatment Plant (Midland, Michigan) on March 30, 2011. This facility treats an excess of 11 million liters of wastewater per day, of which > 90% is from domestic sources.
- Storage conditions: The activated sludge was collected one day prior to initiation of the test, and was continuously aerated until used.
- Preparation of inoculum for exposure:
Prior to use, the activated sludge was screened through 500 µm nylon mesh, and briefly homogenized in a Waring blender (Waring Products Inc., Torrington,
Connecticut). The mixed liquor suspended solids (MLSS) content of the homogenized sludge was determined gravimetrically to be 1,267 mg/L. Based on this determination, 485 mL of the homogenized activated sludge was added to 20 liters of the sterilized mineral medium to yield a final MLSS concentration of 30 mg/L. The mineral medium was then checked for pH and adjusted within the given range of 7.2 - 7.6.
Duration of test (contact time):
28 d
Initial conc.:
ca. 30 mg/L
Based on:
Parameter followed for biodegradation estimation:
O2 consumption
Parameter followed for biodegradation estimation:
CO2 evolution
Parameter followed for biodegradation estimation:
DOC removal
Details on study design:
- Composition of medium: according to guideline
- Test temperature: 22.6 ± 0.3 °C (± 1SD, n=20, min. 22.5 °C, max. 23.5 °C)
- pH: 6.54 - 7.49
- pH adjusted: no
- Suspended solids concentration: ca. 30 mg/L
- Continuous darkness: yes

- Culturing apparatus: specially designed 1-liter glass reaction vessels, sealed
- Number of culture flasks/concentration: 2
- Measuring equipment: O2 and CO2: Columbus Micro-Oxymax automated respirometry system (Columbus Instruments, Inc., Columbus, Ohio); DOC: DOC concentrations were determined using a Shimadzu model TOC-V analyzer (Shimadzu Corporation, Kyoto, Japan) equipped with an ASI-V autosampler; Nitrate/nitrite: Ion Chromatograph model ICS-3000 (Dionex Corporation, Sunnyvale, California)

- Sampling frequency: O2 and CO2: at six-hour intervals; DOC and nitrate/nitrite: at the start and at the end
- Sampling method: O2 and CO2 in the headspace of the reaction vessels

- Inoculum blank: yes, 2 replicates
- Abiotic sterile control: yes, 1 replicate
- Toxicity control: yes, 1 replicate

Descriptive statistics (mean, standard deviation [SD]) were used where applicable.
Reference substance:
% degradation (O2 consumption)
St. dev.:
Sampling time:
28 d
% degradation (CO2 evolution)
St. dev.:
Sampling time:
28 d
% degradation (DOC removal)
Sampling time:
28 d
Details on results:
- The time required for average biodegradation to exceed 10% DO2 (i.e., the lag period) was 2.9 days and the 60% DO2 level was exceeded after 6.1 days. By the end of the 28-day test, % biodegradation of the test substance reached 100.7 ± 21.4% DO2 (mean ± 1SD).
- The 10-day window was met
- The Toxicity Control mixtures containing 102.2 mg/L aniline and 25.3 mg/L of the test substance showed no evidence for inhibition of the microbial inoculum by the test substance.
- The Abiotic Control mixture exhibited no O2 consumption or CO2 production over the duration of the 28-day test.
- The percentage of test substance biodegradation in the replicate Test Mixtures at the end of the 10-day window differed by < 9% DO2
Results with reference substance:
The inoculum used in this test produced > 60% biodegradation of the reference substance, aniline, within the required 10-day window prior to day 14 of the test. The 60% DO2 pass level was exceeded after 8.4 days, and biodegradation based on O2 consumption, CO2 production and DOC removal reached 109.2%, 73.3%, and 97.4%, respectively, at the end of the test.
Validity criteria fulfilled:
Interpretation of results:
readily biodegradable

Description of key information

No biodegradation study is available for TRIS HCl. Therefore, the assessment of biodegradation was based on a study conducted with the structurally similar substance TRIS AMINO. Structural and physico-chemical similarities of the two substances, justify the read-across from TRIS AMINO and to TRIS HCl in the case of biodegradation.

One ready biodegradability study is available for 2-amino-2-(hydroxymethyl)-1,3-propanediol (TRIS AMINO). The study was conducted following the OECD guideline 301 F and according to GLP (5.2.1 -1). Activated sludge was used as inoculum. By the end of the 28-day test period, biodegradation reached 100.7%, based on BOD, or 65.9%, based on CO2 evolution. 60% biodegradation was exceeded within the 10-day window. Thus, the test substance is readily biodegradable according to OECD criteria and based on the analogy it can be concluded that TRIS HCl is also readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information