Oxidase, glucose

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04-02-1992 and 30-07-1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1992

Report date:

1992

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 E (Ready biodegradability: Modified OECD Screening Test)

Version / remarks:

OECD Guideline No. 301E and EEC Directive 67/548 Annex V C.3 as published in 84/449/EEC.

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Active enzyme protein of peroxidase (EC no. 232-668-6, CAS no. 9001-05-2, EC name Peroxidase, Enzyme class no. 1.11.1.7)

Molecular formula:

Not applicable, see remarks.

IUPAC Name:

Active enzyme protein of peroxidase (EC no. 232-668-6, CAS no. 9001-05-2, EC name Peroxidase, Enzyme class no. 1.11.1.7)

Constituent 2

Reference substance name:

Protein as a constituent of enzyme deriving from the fermentation or extraction process

Molecular formula:

Not available

IUPAC Name:

Protein as a constituent of enzyme deriving from the fermentation or extraction process

Constituent 3

Reference substance name:

Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process

Molecular formula:

Not available. See remarks.

IUPAC Name:

Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process

Constituent 4

Reference substance name:

Carbohydrates constituent of enzyme deriving from the fermentation or extraction process.

Molecular formula:

Not available. See remarks.

IUPAC Name:

Carbohydrates constituent of enzyme deriving from the fermentation or extraction process.

Constituent 5

Reference substance name:

Lipids as a constituent of enzyme deriving from the fermentation or extraction process

Molecular formula:

Not available. See remarks.

IUPAC Name:

Lipids as a constituent of enzyme deriving from the fermentation or extraction process

Test material form:

solid: crystalline

Details on test material:

- Lot/batch No.: PPX 3806
- Expiration date of the lot/batch: 04 March 1994

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge (adaptation not specified)

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

46.7 mg/L

Based on:

DOC

Initial test substance concentration 2

Initial conc.:

90 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

Standard substance: Sodium benzoate.
Innoculum pre-treatment: The aerated sample was filtered through a coarse filter paper (first approximate 200 mL discarded) and used on the day of collection.
Usage rate: 0.5 mL of inoculum per litre of dilution water.
Vessels: 2 litre glass beakers.
Test concentrations: (a) Inoculated test medium. (b) 90 mg/L test substance. (c) 35 mg/L standard substance. Each in duplicate.
Preparation of cultures: The dissolved organic carbon (DOC) of each stock solution was determined and sufficient material added to 2 litres medium to give a nominal initial value of ≈45 mgC/L for the test substance and ≈20 mgC/L for the standard substance.
Aeration: By magnetic stirrers and compressed air via narrow bore glass tubes.
Temperature: 22 ± 1°C.
Lighting: The test vessels were shielded from light.
Evaporation: Losses of evaporation were made good by the addition of deionised water immediately prior to sampling.
Sampling: Samples (≈ 20 mL) were withdrawn from each vessel and filtered through Sartorius Minisart® NML, 0.45 µm disposable filters, discarding the first 10 mL. The samples were analysed immediately. Samples were taken on Days 0, 1, 3, 7, 14, 21, 27 and 28.
Analysis: Measurement of dissolved organic carbon (DOC) was conducted using an Ionics TC/TOC Analyser Model 555. DOC analysis was carried out in duplicate for each sample.

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

% Degradation

% Degradation 1

Parameter:

% degradation (DOC removal)

Value:

28

Sampling time:

3 d

% Degradation 2

Parameter:

% degradation (DOC removal)

Value:

57

Sampling time:

7 d

% Degradation 3

Parameter:

% degradation (DOC removal)

Value:

74

Sampling time:

14 d

% Degradation 4

Key result

Parameter:

% degradation (DOC removal)

Value:

91

Sampling time:

28 d

BOD5 / COD results

Results with reference substance:

The reference substance - sodium benzoate - had 74% degradation by day 3 and 103% degradation by day 28.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable, but failing 10-day window

Conclusions:

Peroxidase attained 91% biodegradation within 28 days. The "pass" level of 70% was not reached within 10 days of exceeding the 10% level. However, results should be interpreted with caution as enzymes are multi-component substances, which may imply that the 10-day window criterion is difficult to accomplish. Therefore, peroxidase can be classified as readily biodegradable.

Executive summary:

Biodegradability of peroxidase was tested using OECD Guideline No. 301E and EEC Directive 67/548 Annex V C.3 as published in 84/449/EEC under GLP. Criterion for assessment was Reduction in dissolved organic carbon (DOC). The testing concentrations were (a) Inoculated test medium, (b) 90 mg/L test substance, (c) 35 mg/L standard substance.

The dissolved organic carbon (DOC) of each stock solution was determined and sufficient material added to 2 litres medium to give a nominal initial value of ≈45 mgC/L for the test substance and ≈20 mgC/L for the standard substance. The biodegradation of Peroxidase and the standard substance is shown below:

% Biodegradation

Day       Test substance       Standard

0               0                            0

1            <10>                    <14>

3              28                          74

7              57                          90

14            74                          89

21            78                          99

27            86                         106

28            91                         103

Peroxidase attained 91 % biodegradation within 28 days. The "pass" level of 70% was not reached within 10 days of exceeding the 10% level. However, results should be interpreted with caution as enzymes are multi-component substances, which may impy that the 10-day window criterion is difficult to accomplish. Therefore, peroxidase can be classified as readily biodegradable.

The standard substance, sodium benzoate, attained 103 % biodegradation within 28 days, thereby confirming the suitability of the inoculum and the culture conditions.