Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Environmental fate & pathways

Biodegradation in water: screening tests

Currently viewing:

Administrative data

Link to relevant study record(s)

Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Data is from peer reviewed journal
Justification for type of information:
Data is from peer reviewed journal
Qualifier:
according to guideline
Guideline:
other: as mentioned below
Principles of method if other than guideline:
Biodegradation study was conducted for 100 h (4.166 days) for evaluating the percentage biodegradability of test substance Diphenyl sulphide by using desulfurizing bacterium Mycobacterium sp. ZD-19.
GLP compliance:
not specified
Specific details on test material used for the study:
- Name of test material (IUPAC name): Diphenyl sulphide
- Common name: Phenyl sulfide, (phenylsulfanyl)benzene
- Molecular formula:C12H10S
- Molecular weight:186.277 g/mol
- Smiles : c1(Sc2ccccc2)ccccc1
- InChI : 1S/C12H10S/c1-3-7-11(8-4-1)13-12-9-5-2-6-10-12/h1-10H
- Substance Type: Organic
- Physical State: Liquid
- Other: Diphenyl sulphide (DPS) (99%) was obtained from Acros Organics.
Oxygen conditions:
not specified
Inoculum or test system:
other: Mycobacterium sp. ZD-19 (Bacteria)
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Mycobacterium sp. Strain ZD-19 was isolated from oil-contaminated sludge samples that near different petroleum refineries or mining areas.
- Method of cultivation: One gram of the sample was suspended in 10 ml of
BSM, supplemented with 0.5 mM DBT as sole sulfur source and shaken in rotary shaker at 180 rpm for 48 h at 30°C. After being centrifuged at 500g for 2 min to separate solid, the culture was transferred into the fresh medium. After 4 days cultivation, single-colony isolation was repeated on the plate of the same medium containing 1.5% agar, and isolated colonies were streaked on the plate. Selection of strain ZD-19 among 20 isolated strains was based on the high activity to degrade dibenzothiophene (DBT) as well as to produce 2-HBP (2-hydroxylbiphenyl).
Duration of test (contact time):
4.166 d
Initial conc.:
93.14 mg/L
Based on:
test mat.
Remarks:
(0.5 mM)
Parameter followed for biodegradation estimation:
test mat. analysis
Details on study design:
TEST CONDITIONS
- Composition of medium: Basal salt medium was used as a test medium for the study. The basic salt medium (BSM) for the bacteria growth contains following components per liter of solution: 5 g of K2HPO4.3H2O, 2 g of NaH2PO4.2H2O, 0.2 g of MgCl2.6H2O, 5 g of NH4Cl, 1 ml of mineral solution.
- Additional substrate: Glycerol was added as the carbon source at a concentration of 5 g/l.
- Test temperature: 30°C
- Other: The mineral solution contained (per liter of deionized water): 0.1 g of CuCl2.2H2O, 0.4 g of CoCl2.6H2O, 0.2 g of ZnCl2, 20 g of CaCl2, 0.05 g of H3BO3, 0.2 g of NaMoO4.2H2O, 4 g of FeCl3.7H2O, 0.1 g of AlCl3.6H2O and 0.8 g of MnCl2. 4H2O. BSM supplemented with 0.5 mM DBT was used as enrichment culture. Cells in the medium were harvested by centrifugation at 2500g for 10 min and washed twice with 0.1 mM phosphate buffer (pH 7.0). BSM supplemented with 0.5 mM DBT was used as an enrichment culture.

TEST SYSTEM
- Culturing apparatus: Flasks were used as a test vessel for the study.
- Number of culture flasks/concentration: Duplicates

CONTROL AND BLANK SYSTEM
- Inoculum blank: The dead control (without bacterium) had been done.
Key result
Parameter:
% degradation (test mat. analysis)
Value:
60
Sampling time:
2.083 d
Remarks on result:
other: Other details not known
Key result
Parameter:
% degradation (test mat. analysis)
Value:
80
Sampling time:
3.75 d
Remarks on result:
other: Other details not known
Details on results:
Test substance undergoes 60 and 80% degradation after 50 hrs (2.083 days) and 90 hrs (3.75 days), respectively. The specific desulfurization rate (first 3 h reaction) of test chemical Diphenyl sulphide by ZD-19 resting cells is calculated as 8.58.

Table:The abilities of some bacteria to desulfurize organic sulfur compound Diphenyl sulfide.

 

Microbial

Desulfurization ability

Mycobacterium sp.ZD-19

70%Diphenyl sulfide(DPS)

Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The percentage degradation of test substance Diphenyl sulphide was determined to 60 and 80% degradation after 50 hrs (2.083 days) and 90 hrs (3.75 days), respectively. The specific desulfurization rate (first 3 h reaction) of test chemical by ZD-19 resting cells is calculated as 8.58. Thus, based on percentage degradation, Diphenyl sulphide is considered to be readily biodegradable in water.
Executive summary:

Biodegradation study was conducted for 100 h (4.166 days) for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by using desulfurizing bacterium Mycobacterium sp.ZD-19 at a temperature of 30°C.Mycobacterium sp. Strain ZD-19 was isolated from oil-contaminated sludge samples that near different petroleum refineries or mining areas. One gram of the sample was suspended in 10 ml of BSM, supplemented with 0.5 mM DBT as sole sulfur source and shaken in rotary shaker at 180 rpm for 48 h at 30°C. After being centrifuged at 500g for 2 min to separate solid, the culture was transferred into the fresh medium. After 4 days cultivation, single-colony isolation was repeated on the plate of the same medium containing 1.5% agar, and isolated colonies were streaked on the plate. Selection of strain ZD-19 among 20 isolated strains was based on the high activity to degradedibenzothiophene (DBT) as well as to produce 2-HBP (2-hydroxylbiphenyl).Basal salt medium was used as a test medium for the study.The basic salt medium (BSM) for the bacteria growth contains following components per liter of solution: 5 g of K2HPO4.3H2O, 2 g of NaH2PO4.2H2O, 0.2 g of MgCl2.6H2O, 5 g of NH4Cl, 1 ml of mineral solution. Glycerol was added as the carbon source at a concentration of 5 g/l. The mineral solution contained (per liter of deionized water): 0.1 g of CuCl2.2H2O, 0.4 g of CoCl2.6H2O, 0.2 g of ZnCl2, 20 g of CaCl2, 0.05 g of H3BO3, 0.2 g of NaMoO4.2H2O, 4 g of FeCl3.7H2O, 0.1 g of AlCl3.6H2O and 0.8 g of MnCl2.4H2O. BSM supplemented with 0.5 mM DBT was used as enrichment culture. Cells in the medium were harvested by centrifugation at 2500gfor 10 min and washed twice with 0.1 mM phosphate buffer (pH 7.0).BSM supplemented with 0.5 mM DBT was used as an enrichment culture. For desulfurization/degradation test, resting cells were prepared. Test compound diphenyl sulfide (DPS) was added separately with an initial concentration of 0.5 mM in 3 ml resting cells suspension (initial cell concentration of 15 g dry cell weight (DCW)/L). All reactions were carried out in 25 ml flasks in duplicate at 30°C and 180 rpm in rotary shaker within 3 h.Cell growth was measured turbidimetrically at 620 nm. The concentration of cells was determined from a calibration curve for correlating OD620 to dry cell weight.Test compound Diphenyl sulphide was determined by gas chromatography (GC).The dead control (without bacterium) has also been setup during the study.The percentage degradation of test substance Diphenyl sulphide was determined to 60 and 80% degradation after 50 hrs (2.083 days) and 90 hrs (3.75 days), respectively.The specific desulfurization rate (first 3 h reaction) of test chemical by ZD-19 resting cells is calculated as 8.58.Thus, based on percentage degradation, Diphenyl sulphide is considered to be readily biodegradable in nature.

Description of key information

Biodegradation study was conducted for 100 h (4.166 days) for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by using desulfurizing bacterium Mycobacterium sp.ZD-19 at a temperature of 30°C (Han Chen, et. al; 2008). Mycobacterium sp.Strain ZD-19 was isolated from oil-contaminated sludge samples that near different petroleum refineries or mining areas. One gram of the sample was suspended in 10 ml of BSM, supplemented with 0.5 mM DBT as sole sulfur source and shaken in rotary shaker at 180 rpm for 48 h at 30°C. After being centrifuged at 500g for 2 min to separate solid, the culture was transferred into the fresh medium. After 4 days cultivation, single-colony isolation was repeated on the plate of the same medium containing 1.5% agar, and isolated colonies were streaked on the plate. Selection of strain ZD-19 among 20 isolated strains was based on the high activity to degradedibenzothiophene (DBT) as well as to produce 2-HBP (2-hydroxylbiphenyl).Basal salt medium was used as a test medium for the study.The basic salt medium (BSM) for the bacteria growth contains following components per liter of solution: 5 g of K2HPO4.3H2O, 2 g of NaH2PO4.2H2O, 0.2 g of MgCl2.6H2O, 5 g of NH4Cl, 1 ml of mineral solution. Glycerol was added as the carbon source at a concentration of 5 g/l. The mineral solution contained (per liter of deionized water): 0.1 g of CuCl2.2H2O, 0.4 g of CoCl2.6H2O, 0.2 g of ZnCl2, 20 g of CaCl2, 0.05 g of H3BO3, 0.2 g of NaMoO4.2H2O, 4 g of FeCl3.7H2O, 0.1 g of AlCl3.6H2O and 0.8 g of MnCl2.4H2O. BSM supplemented with 0.5 mM DBT was used as enrichment culture. Cells in the medium were harvested by centrifugation at 2500gfor 10 min and washed twice with 0.1 mM phosphate buffer (pH 7.0).BSM supplemented with 0.5 mM DBT was used as an enrichment culture. For desulfurization/degradation test, resting cells were prepared. Test compound diphenyl sulfide (DPS) was added separately with an initial concentration of 0.5 mM in 3 ml resting cells suspension (initial cell concentration of 15 g dry cell weight (DCW)/L). All reactions were carried out in 25 ml flasks in duplicate at 30°C and 180 rpm in rotary shaker within 3 h.Cell growth was measured turbidimetrically at 620 nm. The concentration of cells was determined from a calibration curve for correlating OD620 to dry cell weight.Test compound Diphenyl sulphide was determined by gas chromatography (GC).The dead control (without bacterium) has also been setup during the study.The percentage degradation of test substance Diphenyl sulphide was determined to 60 and 80% degradation after 50 hrs (2.083 days) and 90 hrs (3.75 days), respectively.The specific desulfurization rate (first 3 h reaction) of test chemical by ZD-19 resting cells is calculated as 8.58.Thus, based on percentage degradation, Diphenyl sulphide is considered to be readily biodegradable in nature.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

Various experimental key and supporting studies for the target compound Diphenyl sulphide (CAS No. 139-66-2) were reviewed for the biodegradation end point which are summarized as below:

 

In an experimental key study from peer reviewed journal (Han Chen, et. al; 2008), biodegradation experiment was conducted for 100 h (4.166 days) for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by using desulfurizing bacterium Mycobacterium sp.ZD-19 at a temperature of 30°C.Mycobacterium sp. Strain ZD-19 was isolated from oil-contaminated sludge samples that near different petroleum refineries or mining areas. One gram of the sample was suspended in 10 ml of BSM, supplemented with 0.5 mM DBT as sole sulfur source and shaken in rotary shaker at 180 rpm for 48 h at 30°C. After being centrifuged at 500g for 2 min to separate solid, the culture was transferred into the fresh medium. After 4 days cultivation, single-colony isolation was repeated on the plate of the same medium containing 1.5% agar, and isolated colonies were streaked on the plate. Selection of strain ZD-19 among 20 isolated strains was based on the high activity to degradedibenzothiophene (DBT) as well as to produce 2-HBP (2-hydroxylbiphenyl).Basal salt medium was used as a test medium for the study.The basic salt medium (BSM) for the bacteria growth contains following components per liter of solution: 5 g of K2HPO4.3H2O, 2 g of NaH2PO4.2H2O, 0.2 g of MgCl2.6H2O, 5 g of NH4Cl, 1 ml of mineral solution. Glycerol was added as the carbon source at a concentration of 5 g/l. The mineral solution contained (per liter of deionized water): 0.1 g of CuCl2.2H2O, 0.4 g of CoCl2.6H2O, 0.2 g of ZnCl2, 20 g of CaCl2, 0.05 g of H3BO3, 0.2 g of NaMoO4.2H2O, 4 g of FeCl3.7H2O, 0.1 g of AlCl3.6H2O and 0.8 g of MnCl2.4H2O. BSM supplemented with 0.5 mM DBT was used as enrichment culture. Cells in the medium were harvested by centrifugation at 2500gfor 10 min and washed twice with 0.1 mM phosphate buffer (pH 7.0).BSM supplemented with 0.5 mM DBT was used as an enrichment culture. For desulfurization/degradation test, resting cells were prepared. Test compound diphenyl sulfide (DPS) was added separately with an initial concentration of 0.5 mM in 3 ml resting cells suspension (initial cell concentration of 15 g dry cell weight (DCW)/L). All reactions were carried out in 25 ml flasks in duplicate at 30°C and 180 rpm in rotary shaker within 3 h.Cell growth was measured turbidimetrically at 620 nm. The concentration of cells was determined from a calibration curve for correlating OD620 to dry cell weight.Test compound Diphenyl sulphide was determined by gas chromatography (GC).The dead control (without bacterium) has also been setup during the study.The percentage degradation of test substance Diphenyl sulphide was determined to 60 and 80% degradation after 50 hrs (2.083 days) and 90 hrs (3.75 days), respectively.The specific desulfurization rate (first 3 h reaction) of test chemical by ZD-19 resting cells is calculated as 8.58.Thus, based on percentage degradation, Diphenyl sulphide is considered to be readily biodegradable in nature.

 

Another biodegradation study from peer reviewed journal (Kim, et. al; 1990) was conducted for 5 days for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by using sulphate reducing bacterium Desulfovibrio desulfuricans M6 under anaerobic conditions at a temperature of 30°C.Test inoculum Desulfovibrio desulfuricans M6 was isolated from soil. Desulfovibrio desulfuricans M6 was cultivated at 30°C using medium C for 3 days.Cultures were grown in an anaerobic pressure tube or in a serum vial with a 5 % inoculum.After 3 days of growth about 70 ml culture was harvested anaerobically and suspended in 65.1 ml of sulphate free medium C (Postgate medium C without sodium sulfate). The cell suspension was added with 1.4 ml of 100 mM methyl viologen solution and 3.5 ml of 2 % organic sulfur compound solution in dimethylformamide. The headspace of the vial was filled with hydrogen gas and the reaction mixture was reduced using 2.5 % sodium sulfide solution before the vial was incubated at 30°C for 5 days. The concentrations of organic sulfur compounds were determined by gas chromatographic method using a Varian 3700 (Sunnyvale, CA) with a flame ionization detector and a glass column (2 m X 2 mm) packed with 10 % OV-101 chromosorb WHP. Nitrogen was used as the carder gas (20 ml/min). The temperatures of injector, column and detector were 220, 200 and 250°C, respectively. The scanning rate was 0.5 scan/sec with an ionization voltage of about 1500 V. Infrared spectrum was recorded using a Perkin Elmer Model 621 IR spectrophotometer with a KBr disk.The percentage degradation of test substance Diphenyl sulphide was determined to 73% degradation after 5 days. Thus, based on percentage degradation, Diphenyl sulphide is considered to be readily biodegradable in nature.

In a supporting study, biodegradation experiment was conducted for 80 h (3.33 days) for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by using desulfurizing bacterium Mycobacterium sp.ZD-M2 at a temperature of 30°C (Wei Li, et. al; 2005). Mycobacterium sp. Strain ZD-M2 was isolated from oil-contaminated sludge samples. One gram of the sample was suspended in 10 ml of MSM, supplemented with 0.2 mM DBT as sole sulfur source and shaken in rotary shaker for 48 h at 30°C. After being centrifuged at 1000 rpm for 2 min to separate solid, the culture was transferred into the fresh medium. After 4 days cultivation, single-colony isolation was repeated on the plate of the same medium containing 1.5% agar, and finally, isolated colonies were streaked on the plate. Selection of strain ZD-M2 among 15 isolated strains was based on the activity and selectivity to degrade DBT and to produce 2-HBP with high rates.Test compound diphenyl sulfide (DPS) was dissolved in ethanol, and each was added with an initial concentration of 0.2 mM to the sterilized BSM as a sole sulfur source for bacterial growth. Cultures were shaken at 30°C at 180 rpm. During the course of bacterial growth, aliquots of the culture were removed and acidified to pH≤2.0 by the addition of 10% HCl. After 30 min of extraction with the same volume of methyl acetate and 10 min centrifugation at 4800 rpm, a portion of the ethyl acetate layer was used to determine the concentration of organic sulfur compounddiphenyl sulfide (DPS)by gas chromatography (GC) equipped with a FID. A 1µl filtered sample was injected into a 0.32 mm interior diameter, 30 m column with a 0.015 mm film SE-54 column operated at 2.07 ml/min – volume velocity of nitrogen carrier gas. The injector and detector temperatures were both maintained at 280°C. The column temperature was remained at a certain degree according to the different organic sulfur compounds till the character peak appeared. Diphenyl sulfide possessed the lowest desulfurization efficiencies with 70% being transformed within 72 h. Thus, based on percentage degradation, Diphenyl sulphide is considered to be readily biodegradable in nature.

 

Additional biodegradation study from peer reviewed journal (Magda Constanti, et. al; 1996) was conducted for 14 days for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by Agrobacterium MC501 and a mixed culture at a temperature of 30°C. Agrubucterium MC501 was isolated from a coal mine area by an enrichment culture with DBT. The mixed cultured XACO consists of Agrobacterium MC501, Xanthomonas MC701, Corynebacteriumsp. MC401, and Corynebacterium sp. MC402 with all cultures individually isolated with the same procedure as described above. Basal salt medium was used as a test medium for the study. The basic salt medium (BSM)contained per liter of distilled water the following:1 g of K2HPO4, 1 g of MgCl2.6H2O, 2 g of NH4Cl, 0.001 g of CaCl2, and 0.001 f of FeCl3.6H2O. Glucose (6 mM) was added as the main carbon source (BSG medium).pH of the test medium was adjusted to 7.0 using NaOH. Diphenyl sulphide (DS)was finely ground and added to BSG in powder form. Cultures were grown in capped Erlenmeyer flasks in a Gallenkamp gyratory shaker at 30°C and 150 rpm. Ail glassware was scrupulously cleaned with HCl to prevent spurious growth on contaminant sulfur. Chemicals used to prepare growth medium were of the highest degree of purity.AgrobacteriumMC501 and the mixed culture XACO were incubated, in duplicate, in 40 ml BSG plus several organic and inorganic sulfur compounds. In this study, test chemical Diphenyl sulphide (DS)(0.161 mmol/l) was used as a sole source of sulfur in growth experiments. Glucose was the carbon source. A flask without any source of sulfur was used as negative control of growth. Growth was followed for several days. Bacterial growth was quantified by direct microscopic counts using a Neubauer chamber. It was also monitored by measuring the absorbance at 440 mn in a Hitachi U-2000 spectrophotometer. Maximum growth of Agrobacterium MC501 and a mixed culture was achieved in 8 and 14 days, respectively. This indicates that the chemical Diphenyl sulphideis utilized for growth by the test inoculum Agrobacterium MC501 and a mixed culture, respectively. Thus, based on this, Diphenyl sulphide is considered to be biodegradable in nature.

 

In an another supporting study, biodegradation experiment was conducted for evaluating the percentage biodegradability of test substance Diphenyl sulphide (CAS no. 139-66-2) by desulfurizing bacterial strain Gordona strain CYKS1 under aerobic conditions (Sung-Keun, et. al; 1998). Gordona strain CYKS1 (Bacteria) was used as a test inoculums for the study.Strain CYKS1 was grown aerobically in 50-ml test tubes containing 5 ml of MSM with organic sulfur compound Diphenyl sulphide(0.3 mM) as the sole sulfur source. Butyl rubber stoppers were used. Cell growth was determined by measuring optical density of culture broth samples at 600 nm. Sulfur-free MSM was used as a negative control. Growth of Gordona Strain, CYKS1 was observed when Diphenyl sulphide was used as a sole sulfur source. This indicates that the chemical Diphenyl sulphide is utilized for growth by the test inoculum Gordona Strain, respectively. Thus, based on this, Diphenyl sulphide is considered to be biodegradable in nature.

 

On the basis of above results for target chemical Diphenyl sulphide (from peer reviewed journals), it can be concluded that the test substance Diphenyl sulphide can be expected to be readily biodegradable in nature.