Serine, 2-[(2,3,4-trihydroxyphenyl)methylene]hydrazide, hydrochloride (1:1)

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was selected as the key study because the information provided for the hazard endpoint is sufficient for the purpose of classification and labelling and/or risk assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

no

Remarks:

The study was conducted according to the test guidelines in effect at the time of study conduct.

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

no

Remarks:

The study was conducted according to the test guidelines in effect at the time of study conduct.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2-amino-3-hydroxy-N'-[(E)-(2,3,4-trihydroxyphenyl)methylidene]propanehydrazide hydrochloride

EC Number:

940-411-0

Cas Number:

1353749-74-2

Molecular formula:

C10H14ClN3O5

IUPAC Name:

2-amino-3-hydroxy-N'-[(E)-(2,3,4-trihydroxyphenyl)methylidene]propanehydrazide hydrochloride

Details on test material:

- Purity: 100%

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

- Source of inoculum: Cambridge Wastewater Treatment Facility, Cambridge, Maryland. The Cambridge facility treats predominantly residential wastes.

- Preparation of inoculum for exposure: The sludge was sieved using a 2-mm screen, adjusted to approximately 1000 mg total suspended solids (TSS)/L with mineral media and then aerated at test temperature until use. A total suspended solids measurement and standard plate count were performed on the inoculum on the day of test chamber preparation. Plates were incubated at 20 ± 3ºC for approximately 48 hours.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

10 mg/L

Based on:

other: measured total organic carbon (TOC)

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Modified biochemical oxygen demand (BOD) test dilution water:

2470 mL of NANO® pure water
3 mL calcium chloride solution (2.75%)
3 mL of ferric chloride solution (0.025%)
3 mL of magnesium sulphate solution (2.25%)
30 mL of phosphate buffer (pH 7.4)
A volume of activated sludge inoculum to achieve a final TSS concentration of ≤ 30 mg/L.

- Test temperature: 20 ± 3ºC

TEST SYSTEM
The test chambers were amber 4-liter bottles. The air entering the chambers was passed through Drierite to remove ambient moisture and then through Ascarite® to produce CO2-free air. The air exiting the test chambers was passed through a series of three gas washing bottles, each containing approximately 100 mL of 0.5 M potassium hydroxide (KOH) to trap the CO2 that had evolved within the chamber. An additional set of gas washing bottles that was not connected to a chamber was maintained concurrently with the traps connected to the chambers. These bottles contained approximately 100 mL of 0.5 M KOH and the amount of CO2 detected in the KOH solution was subtracted from the CO2 in the blank control traps to determine the amount of CO2 produced by the inoculum in the blank control. Magnetic stir bars and stir plates were used to mix the contents of the test chambers. Stir plate motors were cycled on and off approximately every 15 minutes to prevent the heating of the stirrer motors.

All chambers were aerated with CO2-free air for approximately 24 hours at a rate of approximately 60 mL per minute to purge the systems of CO2. After the aeration period, the flow of CO2-free air was stopped, and three CO2 traps, each containing approximately 100 mL of 0.5 M KOH, were connected to the exit air lines of each chamber. Sufficient volumes of reference substance stock solution to achieve a nominal concentration of 10 mg C/L were added to the appropriate chambers. Sufficient volumes of test substance stock solution to achieve a nominal concentration of approximately 10 mg C/L were added to the appropriate chambers. Sufficient volumes of test and reference substance stock solutions necessary to achieve 10 mg C/L as test substance and 10 mg C/L as reference substance were added to the toxicity control. The final volume within all chambers was brought up to 3000 mL by the addition of NANO® pure water and the airflow was restarted on the system.

SAMPLING
- Sampling frequency: Days 1, 5, 8, 13, 16, 19, 23, 26 and 28


CONTROL AND BLANK SYSTEM
- Inoculum blank: yes; The test contained a blank control group, a reference group, a treatment group and a toxicity control. Each group contained three replicate test chambers except for the toxicity control, which was not replicated.

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

Test performance:

The viability of the inoculum and validity of the test were supported by the results of the reference substance, sodium benzoate, from which an average of 108.4% of theoretical CO2 was evolved. An average percent biodegradation of greater than 60% was achieved by Day 5, thereby fulfilling the criteria for a valid test by reaching the pass level by Day 14.

% Degradation

Parameter:

% degradation (CO2 evolution)

Value:

22

Sampling time:

28 d

Details on results:

Evidence of ready biodegradability in a Carbon Dioxide Evolution Test is >60% TCO2 within the 28-day test period. In addition, the pass level must be reached within 10 days of achieving 10% TCO2. The final mean percent biodegradation for the test substance was 22.0%. The test substance is not considered readily biodegradable since 60% TCO2 was not achieved within 10 days of reaching 10% TCO2. The toxicity control achieved > 25% degradation by Day 14 and is considered non-inhibitory at the concentration tested in this study.

BOD5 / COD results

Results with reference substance:

Exceeded the criterion of 60% degradation within the first week (passing the 10 day window) of testing, confirming the suitability of the inoculum and the culture conditions.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

other: not readily biodegradable

Conclusions:

This study and the conclusions which are drawn from it fulfil the quality criteria (validity, reliability, repeatability).
Not readily biodegradable (22% CO2 evolution after 28 days)

Executive summary:

The biodegradability of the test substance was determined by the Carbon Dioxide Evolution Test Method (OECD Guideline 301B). In the CO₂ test, inoculated mineral medium was dosed with a known amount of test substance as the nominal sole source of organic carbon and aerated with CO₂-free air. The CO₂ produced from the mineralization of organic carbon within the test chambers was displaced by the flow of CO₂-free air and trapped as K₂CO₃ in KOH trapping solution. The amount of CO₂ produced by the test substance (corrected for that evolved by the blank inoculum) is expressed as a percentage of the theoretical amount of CO₂ (TCO₂) that could have been produced if complete biodegradation of the test substance occurred. The test contained a blank control group, a reference group, a treatment group and a toxicity control. Each group contained three replicate test chambers except for the toxicity control, which was not replicated. The blank control was used to measure the background CO₂ production of the inoculum and was not dosed with a carbon source. The reference chambers were dosed with sodium benzoate, a substance known to be biodegradable, at a nominal concentration of 10 mg C/L. The treatment group test chambers were used to evaluate the test substance at a nominal concentration of approximately 10 mg C/L. The toxicity control was used to evaluate the inhibition of the test substance to the inoculum and was dosed with both the reference (10 mg C/L) and test substances (10 mg C/L). The results indicated that the activated sludge inoculum was active, degrading the reference substance 108.4%. The test substance was not inhibitory to the microbial inoculum. The average cumulative percent biodegradation for the test substance was 22.0% after 28 days. The test substance is not considered readily biodegradable since 60% TCO₂ was not achieved within 10 days of reaching 10% TCO₂.