Methanesulfonic

acid,1,1'-(1,6-hexanediyldiimino )bis [1-oxo-],sodium salt (1:2)

Ready Biodegradability Test Report

Protocol No.: P-004-2011

Study No.: S-004-20 11

Report No.:R-004-2011 

Study Director: Qingjie Zhao 

Report completion date: June 14, 2011 

Sponsor:

Lanxess (Wuxi) Chemical Co.,

Ltd.

Address: No.9, Zhu Jiang Road,

Wuxi National Hi&New Tech.

Industrial , Development Zone

Wuxi, Jiangsu, 214028, PR. Of

China

Zip Code: 214028

Performing Laboratory :

Environmental Testing Laboratory

Shanghai Academy of Environmental 

Sciences

Address: No . 508 Qin Zhou Road,

Shanghai, P.R. China 

Zip Code: 200233 

Tel: 021-64085119 -2306 

Fax: 021-64838991 
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3

CNASCOMPLIANCESTATEMENT

Study Title Ready Biodegradability Test of Methanesulfonic

acid,l, 1'-(1,6-hexanediyldiimino )bis[1 -oxo-],sodium salt (1 :2)

StudyNo. : S-004-2011

Report No. : R-004-2011

This study was conducted in compliance with GB/T 21831-2008 :

Chemicals -Ready biodegradability : Closed bottle test under CNAS conditions.

To the best of our knowledge, there were no deviations from the protocol and

related regulations that affected the quality or integrity of the study.

Study Director Date

Laboratory Director Date
4

QUALITY ASSURANCE STATEMENT

Study Title Ready Biodegradability Test of 

Methanesulfonic acid, 1,1'-( 1,6-hexanediyldiimino )bis[ 1-oxo-

],sodium salt (1:2)

StudyNo. : S-004-2011

Report No. : R-004-2011

This study was carried out according to the protocol and the final report

accurately reflected the raw data.

Quality Assurance Director inspected and audited the test procedures of the study,

and informed the Study Director and the Laboratory Director of the inspection results.

The inspection and reporting dates that the Quality Assurance Director conducted in

accordance with related SOPs were as follows.

Type of inspection Item of inspection Date of inspection

Date of reporting

to the Study

Director

Date of reporting to

the Laboratory

Director

protocol

editing of the

protocol

March 18,2011 March 18,20 11 March 18, 2011 

test procedure

weighing and

adding of the test

substan ce

April 20, 20II April20, 2011 April 20, 2011 

test report

editing of the test

report

June 14, 2011 June 14, 2011 June 14, 2011

Quality Assurance Director Date
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SUMMARY

The ready biodegradability test of Methanesulfonic acid,1,1 '-(1,6-hexanediyldiimino) bis[1 - 

oxo-],sodium salt (I :2) was carried out in the Environmental Testing Laboratory, Shanghai 

Academy of Environmental Sciences sponsored by Lanxess (Wuxi) Chemical Co., Ltd. 

In the closed bottle test, inoculated mineral medium was dosed with a known amount of test

substance as the nominal sole source of organic carbon. Degradation was followed by the analysis

of dissolved oxygen over a 28-day test period. The test contained an inoculum control group, a

reference substance group, a test substance group and a toxicity control group. Each group

contained ten replicate test chambers. The inoculum control was used to measure the oxygen

uptake of the inoculum and was not dosed with a carbon source. The reference chambers were

dosed with sodium lauryl sulfate, a substance known to be biodegradable , at a nominal

concentration of 2 mg/L. The test substance group test chambers were used to evaluate the test

substance at a nominal concentration of 2 mg/L. The toxicity control group test chambers were

used to evaluate the inhibition of the test substance at a nominal concentration of 2 mg/L of the

test substance and the reference substance. Measurements of dissolved oxygen were performed on

two chambers from the control, reference substance, test substance and toxicity control groups on

days 0,7,14,21 and 28.

The results indicated that the inoculum was active by degrading the reference substance,

sodium lauryl sulfate, within the acceptable range (ie 60% within 14 days). The percent

biodegradation of Methanesulfonic acid ,1,1'-(1,6-hexaned iyldiimino) bis[1-oxo-],sodium salt (1 :2) 

was calculated as 15.7%, 59.8%, 62.6% and 63.9% on days 7, 14,21 and 28.
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1. STUDY TITLE

Ready Biodegradability Test of Methanesulfonic acid, 1,1 '-(1,6-hexanediyldiimino)

bis[l-oxo- ],sodium salt (1:2)

2. OBJECTIVE

This study was performed to determine the ready biodegradability of Methanesulfonic

acid, 1,1'-( 1,6-hexanediyld iimino)bis[ 1-oxo-],sodium salt ( 1:2) in accordance with the

Guidelines for the Testing of Chemicals (HJ!f 153-2004) and GB!f 21831-2008:

Chemicals-Ready biodegradability: Closed bottle test.

3. TESTING FACILITY

Environmen tal Testing Laboratory 

Shanghai Academy of Environmental Sciences

No . 508 Qin Zhou Road, Shanghai, P.R. China 200233

TEL: (86)-21-64085119 ext. 2306 FAX: (86)-21-64838991 

4. STUDY SPONSOR AND CONTACT

Study sponsor: Lanxess (Wuxi) Chemical Co., Ltd.

Contact: Ms. Yin Lian email: candy.yin@lanxess.com mobile: 13917117680

5. MANAGEMENT AND RESPONSIBLE PARTICIPANTS

Laboratory director: Genxiang Shen

Study director: Qingjie Zhao

Principal investigator: Qingjie Zhao

Archiving participant: Zhenqi Wang

6. QA DIRECTOR

QA director: Shuangqing Hu

7. TEST SUBSTANCE INFORMATION

Chemical name: Methanesulfonic acid, 1,1'-(1,6-hexanediyldiimino)bis[1-oxo-], sodium salt

(I :2)

Batch No.:
8

Expiry date:

CAS No.:

Form:

Odour:

38632-47-2

White powder

Irritant

Solubility in water: Miscible

Bulk density: 600 kglm3

Melting point: 166°C

Auto-ignition temperature: >139°C

Decomposition temperature: l92°C

Purity:

Storage:

98.2±1 .2%

Store in original container protected from direct sunlight in a dry, cool

and well-ventilated area

The information above was provided by the sponsor.

8. EXPERIMENTAL DATES

Starting: April 20, 2011

Completion: May 25 , 2011

9. TEST MATERIALS AND INSTRUMENTS

9.1 Mineral Medium

Prepare the following stock solutions, using analytical grade reagents:

(a) KH2P04

K2HP04

Na2HP04·2H20

N Cl

8.50g

21.75g

33.40g

0.50g

Dissolve in water and make up to 1 litre.

The pH of the solution was 7.42.

 (b) CaCh 27.50g

Dissolve in water and make up to 1 litre.

22.50g
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Dissolve in water and make up to 1 litre.

0.25g

Dissolve in water and make up to 1 litre.

Mix 3 ml of solutions (a), (b), (c) and (d) with 800ml water and then make up to I litre.

9.2 Microbial Inoculum

The inoculum was derived from the secondary etlluent of Shanghai Longhua Sewage Plant. The

inoculum was pre-conditioned by aerating the secondary etlluent, without other treatment or addition,

for 7 days at the test temperature of20± I oc. Control microbe counted at the range of I 04-10

6 orgs/1.

9.3 Instruments

Digital pH meter (pHS-25), Analytical balance (AB135-S), Dissolved oxygen meter (YSI-58).

9.4 Test Condition

Test duration: 28d

Temperature: 20 ± I °C

10. METHODS

10.1 Preparation of Bottles

First of all, 5-10 ml of a wash solution (e.g. 2.5g iodine plus 12.5g potassium iodide per litre of

I% w/v sulfuric acid) was added to the empty bottles and shaken well to coat the bottle walls. Bottles

were left to stand for 15 min, poured off the solution and rinsed thoroughly with tap water at least 3

times and finally deionised water.

Mineral medium was strongly aerated for at least 20 minutes and stood for 20 h at the test

temperature. The concentration of dissolved oxygen was determined for control purposes.

Two parallel groups of BOD bottles were prepared for the determination of the test substance,

reference substance, blank control and toxicity control in simultaneous experimental series.

Fully-aerated mineral medium was added to BOD bottles so that they were one-third full, and

then sufficient of the stock solutions of the test and reference substances were added to separate BOD 

bottles.

The concentration of the test substance was 2 mg/1. In toxicity control test, equal amounts of test

and reference substance were added and the final concentration was 2 mg/1 respectively.

The solutions in the BOD bottles were inoculated with secondary effluent (2ml/L).
BOD = 2 2 =0

2

10.2 Operations

Each prepared solution or suspension was dispensed immediately into the respective group of

BOD bottles by those from the lower quarter of the appropriate large bottle, so that all the BOD

bottles were completely filled.

The zero-time bottles were analyzed immediately for dissolved oxygen by the electrode methods.

The remaining replicate bottles were stoppered ensuring that no air bubbles were enclosed, and

incubated at 20°C in the dark.

Each series were accompanied by a complete parallel series for the determination of inoculated

blank medium. Duplicate bottles of all series were withdrawn for dissolved oxygen analysis every 7

days over the 28-day incubation.

10.3 Test Validity

a) Oxygen depletion in the inoculum blank should not exceed 1.5 mg dissolved oxygen/1 after 28

days;

b) The residual concentration of oxygen in the test bottles should not fall below 0.5 mg/1 at any

time;

c) Difference between parallel groups should be less than 20 %;

d) Degradation of the reference substance should be more than 60% after 14 days.

11. CALCULATIONS

First calculate the BOD exerted after each time period by subtracting the oxygen depletion

(mg 0211) of the inoculum blank from that exhibited by the test substance. Divide this corrected

depletion by the concentration (mg/1) of the test substance, to obtain the specific BOD as mg

oxygen per mg test substance. Calculate the percentage biodegradation by dividing the specific

BOD by the specific ThOD. Thus:

0 mg/l(subs1ance)- 0 mg/l(blank) 2 ( (mg) substance (mg)

mg/1(substance)

. BOD(O mg/mg

%degradat1on = ThOD(02 

m substance) x 100

substance)

12. RESULTS

12.1 Result of Ready Biodegradability

The temperature range recorded during the test was 19 to 21 0 and was within the protocol

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specified range throughout the test. The result of the standard plate counts performed on the

inoculum was 6.6x I 05 orgs/L. The measured dissolved oxygen concentration for test media prior

to preparing the test mixtures was 9.09 mg 02/L.

The dissolved oxygen measurement exhibited by the control, reference substance, test

substance and toxicity control groups at each sampling interval was presented in Table 1. The

oxygen depletion of the control for inoculum was < 1.5 mg 02/L.

The viability of the inoculum and validity of the test were supported by the results of the

reference substance, sodium lauryl sulfate. The average percent degradation on days 7, 14, 21 and

28 were 51.8%, 69.3%, 71.1% and 72.1 %, respectively, which fulfilled the criteria for a valid 

test that a percent biodegradation of reference substance should be more than 60% after 14 

days.

Based on the ThOD of Methanesulfonic acid, I ,I'-( 1,6-hexanediyldiimino )bis[ 1-oxo-],sodium salt 

(I :2) (1.15 mg 02/mg), the average percent degradation of Methanesulfonic 

acid,1,1 '-(1,6-hexanediyfdiimino)bis[1 -oxo-],sodium salt (1 :2) on days 7, 14, 21 and 28 

was 15.7%, 59.8%, 62.6% and 63.9%, respectively . (in Table 2)

12.2 Hidden Dangers Potentially Affecting the Test Results in the Test Duration

None.

13. CONCLUSION

As a result, the average percent biodegradation of test substance Methanesulfonic

acid,I ,1 '-(1,6-hexanediyldiimino)bis[I-oxo-], sodium salt (I :2) on days 7, I4, 2I and 28 was

calculated as I 5.7%, 59.8%, 62.6% and 63.9% under these test conditions.

14. ARCHIVES

The following data obtained from this study will be retained in Shanghai Academ y of 

Environmental Sciences Environmental Testing Laboratory for 3 years after final report is

submitted to the sponsor.

(!).Protocol, raw data and final report

(2).Records of the test substance

(3).Records of test species and facility

Subsequent retention period will be decided following the discussion between the sponsor

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and this Environmental Testing Laboratory.

15. REFERENCES

(1)SEPA. The Guidelines for the Testing of Chemicals (HJfT 153-2004). Beijing: China 

Environmental Science Press, 2004. 

(2) National Standard of the People's Republic of China. Chemicals-Ready biodegradability :

Closed bottle test ( ) (GBrr 21831-2008). Beijing: Chinese Standard Press, 2008.

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16. TABLES

Table 1 Dissolved Oxygen in the Test Solutions

Bottle

No.

DO (mg/1)

Od 7d 14d 2ld 28d

Blank

a1 9.09 8.53 8.41 8.35 8.27

a2 9.09 8.53 8.43 8.35 8.30

Test substance

bl 9.09 8.15 7.03 6.91 6.81

b2 9.09 8.19 7.06 6.91 6.82

Reference

substance

C1 9.09 6.45 5.64 5.49 5.38

c2 9.09 6.47 5.66 5.52 5.42

Toxicity

control

dl 9.09 6.05 4.41 4.16 4.05

d2 9.09 6.08 4.45 4.17 4.07

Table 2 Percentage Degradation Results(%)

7d 14 d 21 d 28 d

Test substance

b1 16.5 60.4 62.6 64.1

b2 14.8 59.1 62.6 63.7

Average Percent Degradation 15.7 59.8 62.6 63.9

Reference

substance

C1 52.0 69.5 71.5 72.6

c2 51.5 69.0 70.8 71.6

Average Percent Degradation 51.8 69.3 71.1 72.1

Toxicity control

dl 39.4 63.7 66.5 67.2

d2 38.9 63.0 66.3 66.9

Average Percent Degradation 39.1 63.3 66.4 67.1

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