Registration Dossier

Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference Type:
Biodegradation of chlorinated alkanes and their commercial mixtures by Pseudomonas sp. strain 273
Ester Heath · Wayne A. Brown · Soren R. Jensen
Michael P. Bratty
Bibliographic source:
Journal of Industrial Microbiology and Biotechnology March 2006, Volume 33, Issue 3, pp 197–207 Biodegradation of chlorinated alkanes and their commercial mixtures by Pseudomonas sp. strain 273

Materials and methods

Test guideline
no guideline followed
Principles of method if other than guideline:
The current study is focused on the second issue required to quantify the UABL of a dehalogenase. Namely, it was desired to expand upon the limited set of available data related to the biological dehalogenation of Iong-chain chloroalkanes. To date, most sturlies have focused on chlorinated hydrocarbons with chain lengths less than c4. Results Oll microbial dehalogenation of chlorinated hydrocarbons with carbon chain lengths greater than C9 are sparse. A notable exception is the work performed with tx- and tx,w-chlorimited alkanes. With regards to mixtures, there have been several attempts to biodegrade industrial mixtures of polychlorinated alkane, but only the total amount of chloride released from the alkane mixture was reported. To fill this gap in the literature, the biodegradation of chlorinated alkanes with carbon chain lengths between C6 and C16 was examined. The aim was to elucidate the effect of chlorine position and carbon chain length on biodegradation. The work focused on pure compounds, as current analytical techniques cannot resolve the complex industrial PCA mixtures into its substituents. As mainly only tx-chlorinated and tx-w-dichloro alkanes are commercially available, internally chlorinated alkanes were synthesized in our laboratory. The gross degradation of a commercial mixture of chlorinated alkanes was also examined.
GLP compliance:
not specified

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:

Study design

Oxygen conditions:
not specified
Inoculum or test system:
other: Pseudomonas sp. strain 273
Duration of test (contact time):
ca. 10 d
Initial test substance concentration
Initial conc.:
ca. 100 µg/L
Details on study design:
Biodegradation tests were carried out in 500-ml shake flasks containing 100 ml of mineralsalt media. Per Iiter, the medium contained 2.12 g Na2HPO4, 1.36 g KH2PO4, 0.5 g (NH4HSO4 and 0.2 g MgSO4·7H20, 500 mg yeast extract and 1 ml of microelement solution. In some cases, shake flasks were prepared without yeast extract in order to determine microbial growth on chlorinated alkarre as sole carbon source. The microelements solution consisted of, per liter, 20 g NaOH, 10 g MgSO4·7H2O, 4 g ZnSO4·7H2O, 1 g CuS04·5H2O, 3.2 g MnSO4·H2O, 20 g Fe2(S04h·7H2O, 100 g Na2SO4, 1 g NaMoO4·2H2O, 1 ml 96°C H2SO4 and 120 g EDTA. Shake flasks were incubated at 30°C and agitated at 200 rpm. A volume of 50 or 100µl of chlorinated hydrocarbons was added to the mineral medium.

Results and discussion

% Degradation
Key result
other: dehalogenation
ca. 100
Sampling time:
17 d
Remarks on result:
other: no information on standard derivation is available

Applicant's summary and conclusion

Validity criteria fulfilled:
not specified
Interpretation of results:
other: biodegradable/complete dehalogenation
Complete dehalogenation of 100 µg/l of 1, 10- DCD was achieved in 5-17 days of incubation, depending upon the amount and physiological state of the innoculum used.
Executive summary:

The biodegradation of chlorinated alkanes was studied under oxic conditions with the objective of identifying favorable and unfavorable intramolecular chlorination sequences with respect to the enzymes

studied. Several dehalogenating bacterial strains were screened for their ability to degrade middle-chain polychlorinated alkanes as well as a commercial mixture. Of the organisms tested, the most promising was Pseudomonas sp. strain 273, which possesses an oxygenolytic dehalogenase. The effects of carbon chain length (C6-C16), halogen position, and overall chlorine content were examined using both commercially

available compounds and molecules synthesized in our laboratory. The effects of co-substrates, solvents, and inducing agents were also studied. The results with pure chlorinated alkanes showed that the relative positions of the chlorine atoms strongly influenced the total amount of dehalogenation achieved. The greatest dehalogenation yields were associated with terminally chlorinated alkanes. The rx- and rx,w-chlorinated compounds yielded similar results. Vicinal chlorination had the most dramatic impact on degradation. When present on both ends or at the center of the molecule, no dehalogenation was detected. Although partial dehalogenation of 1 ,2-dichlorodecane was observed, it was likely due to a combination of ß-oxidation and an abiotic mechanism. Cereclor S52 was appreciably dehalogenated in shake flasks only when 1,10-dichlorodecane was present as a co-substrate and after increasing the oil surface area through mechanical emulsification, demonstrating the importance of abiotic factors in degrading commercial polychlorinated

alkane mixtures.