quality assurance statement
TRANSCRIPT
f-RD903: INHERENT BIODEGRADATION Key Lab of Pesticide Environmental Assessment and Pollution Control, MEP
SPONSOR
E.I. du Pont de Nemours and Company
Report No.: R2009NC03 I (a)-02 DPI38823 l
REPORT FOR INHERENT BIODEGRADATION OF FRD903
[Modified MITI (II) Test]
Study No.: S2009NC03 l(a)-02
Report No.: R2009NC03 l(a)-02
Study Director: Shi Lili, professor
Date of Report Completion
Mar. 26,2010
'-c:- -::.~ ~ r ~ub~ gllDm~ Key Lab. of Pesticide Environmental 4,ssess,ncnt and Pollution Control, MEP
I' I ,11 Address: 8 .Jiang-wang-miao Street, Nanjing, 210042, Cbina
Tel: (+86)25 85287074 Fax: (+86) 25 85474630
Page I of 19
FRD903 INHERENT BIODEGRADATION Key Lab of Pesti_~ide Envir~nmental A~~essmcn_~~~1d Po!lution ~ntrol, MEP
CONTENTS
Report No.: R2009NC031 (a)-02 DPJ38823 l
SPONSOR AND TEST FACILITY ...................................................................................................... 3
STATEMENT OF GLP COMPLIANCE ............................................................................................. .4
QUALITY ASSURANCE STATEMENT ............................................................................................ 5
STUDY DETAILS PAGE ..................................................................................................................... 6
SUMMARY ......................... , ................................................................................................................. ?
1 INTRODUCTION .............................................................................................................................. 8
2 EQUIPMENT & MATERIALS ......................................................................................................... 8
2.1 DATEQFTIME .......................................................................................................................... 9
2.2 EQUIPMENTS & REAGENT .................................................................................................... 9
3 TEST DESIGN ................................................................................................................................... 9
3.1 PREPARATION OF THE INOCULUMS .................................................................................. 9
3.2 PREPARATION OF SOLUTIONS OF THE TEST SUBSTANCE ........................................... 9
3.3 PREPARATION OF THE TEST MEDIUM ............................................................................... 9
3.4 TEST CONDITIONS ................................................................................................................ 10
3.5 TEST PROCEDURE ................................................................................................................. ] 0
3.6 CHEMICAL ANALYSIS .......................................................................................................... 11
4 TEST VALIDITY ............................................................................................................................. 12
5 DATA PROCESSlNG ...................................................................................................................... 12
5.1 CALCULATION OF THEORETICAL OXYGEN DEMAND ................................................ 12
5.2 CALCULATION OF PERCENTAGE BIODEGRADABILITY .............................................. 13
6 RESULTS ......................................................................................................................................... 13
6.1 ANAL YTlCAL METHOD OF FRD903 IN TEST SOLUTION .............................................. 13
6.2 INHERENT BIODEGRADA TION OF FRD903 ...................................................................... 14
7 RECORDS&DOCUMENTA TION .................................................................................................. 14
8 HEALTH&SAFETY ........................................................................................................................ 14
9 REFERENCES ................................................................................................................................. 15
TABLES .............................................................................................................................................. 16
Table 1 Results of BOD .................................................................................................................. 16
Table 2 Biodegradation as BOD/TOD ............................................................................................ 16
Table 3 Analytical results and Degradation of FRD903 ................................................................ 16
FIGURES ........................................................................................................................................... 17
Figure l Biodcgradation curve........................................................................... . ......................... 17
Figure 2 Work curve......................................................................... . ........................ 17 '#'
Figure 3 Chromatogram of FRD903 (0.005 mg/L) ................ .
Figure 4 Chromatogram of abiotic Sample at O d and 28 d.
Figure 5 Chromatogram of Test Sample at 28 d ............. .
Figure 6 Chromatogram of 13\ank Control. ..
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.... 18
. .......... 18
19
.19
FRD90l: INHERENT BIODEGRADATION Report No.: R2009NC03 I (a)-02
--~~~~ ~---p~!_i~~~-~nvironmental Asses~ment and Po!lut(?_ll_ <;on~~?l~, M_E_:p __ _ DP138~82~3_1 ___ _
SPONSOR AND TEST FACILITY
Sponsor: E.I. du Pont de Nemours and Company
Address: CRP 702/1220, 4417 Lancaster Pike, Wilmington, DE 19805, USA
Tel: 001 302 999 3266
Sponsor's representative: Michael D. Bolten
E-Mail: [email protected]
Test Substance: :FRD903
Test Facility: Key Lab of Pesticide Environmental Assessment & Pollution Control,
MEP (PEAPC)
Address: 8 Jiangwangmiao Street, Nanjing 210042, China
Tel.: 86-25-85287074
Fax: 86-25-85474630
Legal Representative Person: Gao Zhenning, general-director, professor
Laboratory Management: Shan Zhengjun, professor
Study Director: Shi Lili, professor
Study Personnel: Liu Jining, Wang Na, Wu Guanqun
Quality Assurance Personnel: Ge Feng, assistant professor
Page3ofl9
FRD903: INHERENT !3!ODEGRADATION Rcporl No.: R2009NC03 I(a)-02 _ _Key Lab_ of_Pc~ticide _g_n~·onmental _Assessment ~d Poll~~n_Control, _M_EP ______________ _!?_P_l_3_8~231 ________________ _
STATEMENT OF GLP COMPLIANCE
Study No.: S2009NC03 l(a)-02
Report No.: R2009NC03J(a) -02
-i 1
1
l i -i ~I i
According to "OECD Guidelines for testing of chemicals" and "The guidelines for the testing of l chemical (HJ/T 153-2004)" and "The guidelines of chemical testing good laboratory practices I
( (HJ/T I 55-2004) " issued by State Environmental Protection Administration (SEPA) of the I People's Republic of China, this experiment was conducted under CMA (China Metrology I Accreditation) and CNAS (China National Accreditation Service for Conformity Assessment) I experimental conditions at our laboratory. The experimental protocol was strictly carried out in f the process of the experiment, and the present report has reflected the experimental results truly
and correct
(Study Director) Date:
(Lab Management) Date:
Page 4 or 19
I
l l t
I i t
I I
l i
I l
I
FRD903: INHERENT BIODEGRADATION Key Lab of Pesticide Environmental Assessment a,,d p II 1· c ----·- ______ , ... ------ . • o u !On ontrol, MEP
Report No.: R2009NC031 (a)-Ol DPl388231
QUALITY ASSURANCE STATEMENT
Study No.: S2009NC03 J(a)-02
Report No.: R2009NC03J(a)-02
This experiment was carried out strict! in , d . . y accor ance with the experimental protocol It . ,
hereby certified that what the present report d -b h · IS . escn es as accurately reflected the raw data of th
expenment. e
The dates of Quality Assurance inspection are given below.
During the on-site process inspections procedures applicable to this type ofstt1d . Y were mspected.
The rep011ing date is the date of reporting to the
forwarded to the Test Facility Management.
Study Director. The QAU report was then
Type of Phase/Process
Start inspection inspections
End inspection date date
Reporting date
Study Protocol Dec. 8, 2009 Dec. 8, 2009 Report Jan.28,2010 Jan.28,2010
Dec. 8, 2009 Jan.28,2010
Process Test condition check &
BOD measurements Dec. 24, 2009 Dec.24,2009 Dec. 24, 2009
(Person responsible for QAU) Date:
Page 5 of 19
Y ,.~;>:, ·:;:--·'./7;; <?¾02 X
r-RD903: INHERENT BIODEGRADAT!ON . __ Ke)'._ Lab o!_!'?s.!!.?_idc -~~-ironme~t_al Asse~~~_m:nt anti Pol~\~ Contro~,. Mf:~.-----
Report No.: R2009NC0J I(a)-02 lW\388231
Study number:
.ize !JOit_n u IIl bE>f:
Study title:
Test substance:
Identity:
CAS No.:
Chemical name:
Chemical formula:
Molecular Weiisht_
Lot number:
Expiry date,
Ap[Jem:_anc:e:
[/,;~:;:7"=-:~upp~ec_ - - --i Head of Department:
1 StudyDirector:
STUDY DETAILS PAGE
S2009NC03 l (a)-02
R2009NC03 l(a)-02
Inherent Biodegradatio_ll' l\fodifie<l_M_ITI(IIL ___ --J
FRD 903
FRD 903
13252-13-6 -·- _ ··-·- _ -2 3 3-3-t;traflu~~o:2-(hept~fluoropropoxy) propanoic acid
' ' ' ------ -··- --·
Jt~y ,_2,0_10
Liquicl_ Keep container tightly closed and store in a cool,
dark well ventilated location - , ____ --- -------- ------ --- ---- --
96%
E.L du Pont de Nemours arn:I_ C_oll1JJllll)'_
Shan Zhengjun --------- ---
Shi Lili
Person attending to routine duties and 1 technical queries in the temporary
Liu Jining.
i absence ofthe_Stujy Director:
l Study Director contact details:
· Location of study:
Study dates:
Start:
Completion:
Draft report:
Shi Lili l Telephone: 1-86 25 85287074
: Facsimile: +86 25 85474630
Email: [email protected], s_U_nc~@~~_ot11:i_ail.c~m ___ _
K;y-Lab of p;;tic1dc Environmental Assessment and
Pollution Control, MEP
8 Jiang-wang-miao Street
Nanjing 210042
Jiangsu
China
Dec. 14, 2009
Jan. 20,2009
. Feb. 03, 2009
Page 6 of 19
FRD903: INHERENT 8!ODEGRADATION Key Lab of Pesticide_ Eny_i~~~1!~1~1~!.~!-~sscssment and P_ollution Control, MEP
SUMMARY
Report No.: R2009NC0J ! (a)-02 DP 1388231
-----
The inherent biodegradation test for the test substance of FRD903 was conducted according to
the following guidelines:
J) SEPA HJ/T 153-2004, "The guidelines for the testing of chemicals".
2) SEPA. The guidelines for the testing of chemicals. Beijing: China Environmental Sciences
Publishing House. 2004.
3) GB/T 21818-2008. Chemical Inherent Biodegradation-Modified MITI Test (II).
4) OECD Procedure 302C, "Inherent Biodegradability: Modified MITI Test (II)".! 981.
The inherent biodegradability of FRD903 was determined in a 28-day Biochemical Oxygen
Demand (BOD) test and the analysis of residual chemical of FRD903 in BOD bottles in an
aerobic, aqueous medium.
During the test, the temperature was kept at 25 °C ± 1 °C. The test was val id because the level of
biodegradation of the reference substance aniline exceeded 40% after 7 days, and 65% after 14
days.
Based on the residue analysis, the biodegradation of FRD903 was 1.52% and there was hardly
any change for the test chemical of FRD903 in the "abiotic" vessel during the testing period. The
BOD results showed that biodegradation ofFRD903 was both <1 % after 14 days and 28 days.
Therefore, FRD903 has no inherently biodegradable under this test condition.
Page 7 of19
FRD903: INHERENT BIODEGRADATION _ , . , Key Lab of Pesticide Environmental Assessment and Pollution Control, MEI
1 INTRODUCTION
Report No. : R2009NC03 l(a)-02
- ---~~~88':'..'.2::.3..:...1 ------ -
The purpose of this test was to evaluate the inherent biodegradability of organic chemicals via a
28-day test. In the test, the test s~bstance and/or micro-organisms not adapted to the test
substance were added into the aerobic, aqueous medium in BOD bottles respectively. Then th
e
Biochemical Oxygen Demand (BOD) and residual chemicals in BOD bottles was measured
during the 28-day period. It was designed to meet the requirements of SEPA HJ/T 153-2004, "th
e
guidelines for the testing of chemicals"; GB/T 2 l 818-2008, "Chemical Inherent Biodegradation
Modified MITI Test (II)", and OECD Procedure 302C, " Inherent Biodegradability: Modified
MITI Test (II)", adopted May 1981.
The m~thod was not applicable to test substances that were inhibitory to aerobic sewage micro
organisms at the test concentration and that did not reach and react with the CO2 adsorbent.
Test solutions were prepared in an inorganic salts medium, inoculated with a number of micro
organisms collected from not less than 10 places in Nanjing city. Those organisms collected were
kept in BOD bottles in the dark at 25°C ± 1 °C.
F f II b" t· 11 " . £ •ence" " blank control" and "test" were set up simultaneously. The
our groups o a 10 1c , 1e e1 , "abiotic" contained mineral salts medium and a measured amount of test substance in order to
determine whether there was any change in the test chemical during the testing period, while the
" reference" contained inoculated mineral salts medium and a measured amount of a reference
substance for validating the test result. Besides the "control" only contained inoculated mineral
salts medium, while the " test" contained inoculated mineral salts medium and a measured amount
of the test substance.
The progress of degradation was followed by the determination BOD in the "test" and "contro l".
Degradation was expressed as the ratio of the biochemical oxygen demand (BOD) and the
1 · 1 d d (TOD)
1·11 order to evaluate the inherent biodegradability of chemical
t 1eoret1ca oxygen eman
b I .c . t· the i·elati·ve propmiions of the maJ· or components of the test substance
su stance. n101 ma 1011 on · together with their empirical formulae or TOD was therefore necessary prerequisites to this test.
The inherent degradation rate was also expressed as percentage of initial concentration of test
substance, where the residue analysis of the test substance was performed at the beginning (0 d)
and end (28 cl) of the test.
S b ·ct • d to l)c 11
1·n11e1·e11t biodegradable" if the inherent degradation rate was
u stances were cons 1 e1 e
equal to or greater than 20% during the 28-day test period.
The test was inval id if the level ofbioclegradation of the reference ~ubsta ncc did not exceed 40%
after 7 days , and 65% after 14 clays.
2 EQUlPMENT & MATERIALS
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FRD903: INHERENT BIODEGRADATION Report No.: R2009NC03 l(a)-02 _ ::..:Kc:_cy_l _,ab_ o_f_P_cs_ti_ci_de_E_.n_vi_ro_n_m_en_ta_l '-As-.:..::s-.:..::cs.:..:sm.:.:.:e::.:n.:..:t a::.:n=..d .:..:Po:::11:::ul::.-io::::n_-:C:.--o::::nt_:_-:ro'.'.!.l,..:.:M~E::.· P ___ _ __ ~ D_1:_P '..::13 8823 I
2.1 DATE OF TIME
Dec. 14, 2009 - Jan. 20, 201 0
2.2 EQUIPMENTS & REAGENT
Incubator(Lovibond, GER), pH-electrodes, BOD bottles(S00 mL), BOD meter (Lovibond,
GER.Lovibond® I3OD-OxiDirect sensor system for measurement of the Biochemical Oxygen
Demand (BOD) measures BOD levels via the pressure drop in the closed system during a defined
period of time. The inductive stirrer system not only mixes the sample but also ensures optimum
gas exchange between sample and gas space in the sample flask), UPLC-MS-MS (Waters, USA).
Reagents for the medium were all analytical pure.
3 TEST DESIGN
3.1 PREPARATION OF THE INOCULUMS
Activated s ludge, surface soil and surface water were sampled from ten sites distributed in four
districts throughout Nanjing city, such as Chengdong, Chengbei, Baguazhou and Hexi. 1 L of the
sludge, soil and water were collected and mixed thoroughly together. After removing float ing
matter, the mixture was allowed to stand and then the supernatant is filtrated through 0.45 ~Lm
Millipore filter. After that the filtrate was adjusted the supernatant to pH 7.0 with sodium
hydroxide or phosphoric acid. Finally an appropriate volume of the filtered supernatant was
transferred to a fill-and-draw activated s ludge vessel and aerated fo r about 23.5 h.
Thi1iy minutes after stopping the aeration, about one th ird of the whole volume of supernatant
was discarded. Then an equal volume of the solution (pH 7.0) containing 0. 1 % each of gl ucose,
peptone and potassium 01ihophosphate, was added into the settled material and aerated again.
This procedure was repeated once per day until the inoculums were used.
Before use the mixture was allowed to stand, and the supernatant was removed. A small quantity
of s ludge was taken to be centrifuged (] 0000 rpmx IO min) and then weighed. Then the s ludge
was dried in the oven and weighed again in order to calculate the content of dry s ludge. At last a
ceiiain amount of centrifuged sludge was diluted with basal culture med ium to get act ivated
sludge suspension with a concentration of l 000 mg/L (dry basis).
3.2 PREPARATION OF SOLUTIONS OF THE TEST SUBSTANCE
/\ stock solution of the test substance (FRD903) at I 00 mg/L: I 05 mg of test substance was
dissolved in 1 T, volumetric flask with BSM.
A stock solution of the reference substance (anili ne) at 1000 mg/L: 1001 mg of reference
substance was dissolved in l L volumetric flask with BSM.
3.3 PREPARATION OF THE TEST MEDIUM
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FRD903: INHERENT BIODEGRADATION Key Lab of Pe~tici?~ En~i!onmental Assc_ssment and Pol_lution Control, MEP
Report No.: R2009NC03 l(a)-02 DP\388231
The Based Salt Medium (BSM) was prepared by adding 3 mL of each of the following stock
solutions prepared in pre-aerated pure water to 1 litre of pure water.
Stock solution A KH2PO4 (potassium dihydrogen phosphate)
K2HPO4 ( di potassium hydrogen phosphate)
Na2HPO4.2H2O ( disodium monohydrogen phosphate dihydrate)
NH4Cl (ammonium chloride)
The pH of this solution was 7.2
Stock solution B
CaCI, ( calcium chloride)
Stock solution C MgSO4.7H20 (magnesium sulphate heptahydrate)
Stock solution D FeCl,.6H2O (iron (III) chloride hexahydrate)
3.4 TEST CONDITIONS
(I) Concentration oftest chemicals: 30 mg/L (W/V)
(2) Concentration of reference chemicals: 100 mg/L (W /V)
(3) Concentration of activated sludge: 100 mg/L (W/V)
( 4) Test temperature: 25°C ± I °C
(5) Period: 28 days
(6) Stir vigorously with mechanical stirrer.
3.5 TEST PROCEDURE
g/L
8.50
21.8
22.2
1.7
27.5
22.5
0.25
15.0 mL of stock solution ofthc test substance were added to Ill vessel designated as "abiotic",
then fixed 50 mL with BSM at 30 mg/L.
#2, #3 and /14 vessels of"test vessel" added 15.0 mL, 15.0 mL, 15.0 mL of stock solution oftest
substance respectively, then 5 mL of activated sludge suspension were added to every vessel and
fixed 50 rnL with BSM at 100 mg/L of activated sludge and 30 mg/L oftest substance.
/15 vessel of "aniline" added 5 mL stock solution of reference substance and 5 mL of activated
sludge suspension, then fixed 50 mL with BSM at l 00 mg/L of activ~jed sludge and 100 mg/I, of
reference substance,
/16 vessel of "blank control" added 5 mL of activated sludge suspension, and then fixed 50 mL
with BSM al I 00 mg/L of activated sludge.
Page I0oCl9
FRD903: INHERENT BIODEGRADATION --~~-1:0~ _(!:f £~s_t_i~_ide _!~~_vironmental Asscs_sment and Poll_uti~1~ -~~~~?l,__~_li~
Report No.: R2009NC03 I (a)-02 DP1388231
After that, assembled the equipment, checked that it is air-tight, began the stirrers, and stmied the
measurement of oxygen uptake under conditions of darkness.
The temperature, the operation of the stirrer and recorder was checked daily. Any changes in
colour of the contents of the vessels were recorded. The BOD for the six bottles were determined
and recorded at the days of 0, 5, 7, 11, 14, 18, 21, 25 and 28.
After the O and 28 days of testing, residual amounts of chemicals in the testing bottles were
analysed.
3.6 CHEMICAL ANALYSIS
( 1) Preparation of standard storage solution
A standard stock solution of 1000 mg/L FRD903: weighing 0.052 l g test substance and fixed 50
mL with deionized water.
Draw 1.0 mL standard stock solution of 1000 mg/L to 100 rnL volumetric flask and fixed 100
mL with deionized water at 10.0 mg/L oftest substance.
Draw 1.0 mL test substance solution at 10 mg/L to 10 mL volumetric flask and fixed 10 mL with
deionized water at I. 0 rng/L of test substance.
(2) Preparation of work solutions
Draw 0.01, 0.05, 0.10, 0.20, 0.50 mL standard solution of test substance at 1.00 mg/L to 10.0 mL
volumetric flask respectively, then fixed 10 mL with diluted I 0000 times BSM, then obtained the
standard work solutions at 0.001, 0.005, 0.01, 0.02, 0.05 mg/L. Details of the work solutions are
showed as follows:
the concentration of the concentration of the work solution
(mg/L)
0.001
0.005
0.01
0.02
0.05
the test solution added (mg/L)
1.00
1.00
1.00
1.00
1.00
(3) UPLC-MS-MS determination conditions
the volume of the test solution added
(mL)
0.010
0.050
0.10
0.20
0.50
fixed volume (mL)
I 0.0
I 0.0
10.0
l 0.0
I 0.0
Apparatus: ACQUTTY RTM Ultra Performance LC, Quattro Premier XE MS-MS (Waters, USA)
Column: ACQUITY UPLC® BEH Cl 8 1.7 flm, 2.1x50111111 (Waters, USA)
Mobile phase: Methanol: wateF60:40
Flow rate: 0.3 mL/min
Column Temperature: 30 °C Pagcl!orl9
\
I \
l I
FRD903: INHERENT BIODEGRADATION Key Lab of Pesticide Environmental Assessment and Pollution Control, MEP
injection volume: 10 µL
MS/MS detection parameters are as follows:
Data type: SIR data
Ionisation mode: Electrospray, Negative
Capillary: 3.0 kV, Extractor: 3.0 V, RF Lens : 0.2 V
Source Temperature: 110 °C, Desolvation Temperature: 380 °C
Cone Gas F low: 50 L/h, Desolvation Gas Flow: 500 L/h
LM Resolution 1: 7.9, LM Resolution 2: 5.1
HM Resolution 1: 15.0, HM Resolution 2: 14.7
Ion Energy 1: 0.4, Ion Energy 2: 2 .0
Collis ion Gas (Pressure) : 3.43e-3
mbar
Ion (m/z) Dwell (s) Cone V olt. (V)
284.76 0.100 -3 1
Report No.: R2009NC03 l(a}-02 DPl388231 - --- -- - - -
Under the above conditions, the retention time of FRD903 was about 0.64 min (see Fig. 3).
(4) Sampling and analysis of test solution
Samples were taken from "abiotic" bottle at O day and 28 day, then from the "test" bottles at 28
day. After filtration by 0.22 µm M illipore fi lter and diluted 10000-fold with BSM, the
concentration of test substance was determined with the analysis method mentioned above.
4 TEST VALIDITY
Viability of the microorganisms should be checked by means of a reference control.
Biodegradation of the reference substance reached >40% and 65% on 7 day and 14 day.
5 DATA PROCESSING
5.1 CALCULATION OF THEORETICAL OXYGEN DEMAND
The theoretical oxygen demand (TOD) of the test and reference substance:
of molecu lar weight (MW), could be calculated in the fo llowing way assuming that carbon was
mineralised to CO2
, hydrogen to H20 , phosphorus to P70 5 and potas~ um to K, O. Halogens were
presumed to be eliminated as hydrogen ha] ides and nitrogen as ammonia.
Page 12 of 19
FRD903: INHERENT BIODEGRADATION Key Lab of Pesticide Environmental Assessment and Pollution Control, MEP
Report No.: R2009NC03 I (a)-02 DP 1388231
- ··- - - -----
TODN113 = 16 [2c + 1/2 (h-cl-3n) + 3s + 5/2p + l/2k- o] (mgO;/mg)
MW
Nitrogen 111 a test substance could be eliminated as ammonia, nitrite or nitrate. The higher
oxidation states gave a consequent increase in the TOD + 3n/2 (nitrite) and + 5n/2 (nitrate).
Now, the mo lecular of test substance is C6HF110 3. The TODNm of the test substance can be
calculated as follow:
TODN1-1J = 16[2c + l / 2(h - /)-o] = 16[2x6+1/2(1-11)-3] =0_ 194 (mgO/ mg) MW 330.05
The TODNH3 of the test substance FRD903 is 0. 194 mg 0 2/mg.
5.2 CALCULATION OF PERCENT AGE BIODEGRADABILITY
(1) Method for calcu lating the percentage biodegradation from the oxygen co11sumption:
BOD - B % degradation = - --- x 100(%)
TOD
BOD: Biological oxygen demand (experimental, mg) of the test substance measured on the BOD
curve.
B: Oxygen consumption (experimental, mg) of basal culture medium to which the inoculum was
added measured on the BOD curve.
TOD: Theoretical oxygen demand (theoretical, mg) required when the test substance 1s
completely oxidised.
(2) Method for calculating the percentage degradation from the resul t of chemical analysis:
S - S h " X} 00(o/o) Sh
% degradation
S.: Residual amount ( experimental, mg) of the test substance after completion of the
biodegradability test.
St, : Residual amount (experimental, mg) of the test substance in the "abiotic" test to which only
the test substance has been added.
6RESULTS
6.1 ANALYTICAL METHOD OF FRD903 IN TEST SOLUTION
(I)Workcurvc
A series of work solutions with concentration at 0.00 I, 0.005, 0.01 , 0.02, 0.05 mg/L were
measured under the UPLC-MS-MS conditions mentioned above. Based 011 the test resul t, a linear
regression equation was obtained with the concentration and the area of the peak emerged at
Page 13 or 19
FRD903: INHERENT BIODEGRADATION _ _ Key Lab of Pesticide Environmental Assessment and Pollution Control, MEP
Report No.: R2009NC03 l(a)-02 DPIJ8823 I
- - --
about 0.64 min: A = 3 88136 c + '/4.6, with good linearity of r '==0.999, where A represents peak
area; and c is concentration (mg/L) (see Fig. 2). The results show that linearity for concentration
range of 0.001- 0.5 mg/L is good.
(2) Detection limit
The minimum detection amount of UPLC-MS-MS for FRD903 is 1.0x 10-11
g, and the minimum
detection concentration for test sample is 0.001 mg/L.
6.2 INHERENT BIODEGRADATION OF FRD903
Table 1, Table 2, Table 3, Fig.4, Fig.5, Fig.6 showed results of inherent biodegradation data.
During the test, the temperature kept at 25°C ± 1 °C. The test was valid because the level of
biodegradation of the reference substance aniline exceeded 40% after 7 days, and 65% after 14
days.
The results showed that biodegradation of FRD903 were both < l % after 14 days and 28 days.
Res idual amounts (experimental, mg) of the test substance in "test" vessels after completion of
the biodegradation test and in "abiotic" vessel before and after completion of the biodegradation
test were determined by UPLC-MS-MS. Based on the residue analysis, the biodegradation of
FRD903 was 1.52% and there was hardly any change of the test chemical in the "abiotic" vessel
during the testing period.
Therefore, the test substance has no inherently biodegradable under this test cond ition.
7 RECORDS&DOCUMENTATION
All test samples arising from the performance of this study w ill remain the prope1ty of the
Sponsor. Records and documentation relating to this study ( including electronic records) will be
maintained in the archives of PEAPC for a period of five year from the date on which the
D irector signs the final repo1i . This inc ludes raw data, a copy of the fi nal repor t. Test substance
remained w ill be retained by P EA PC in its archive fo r a period of one year from the date on
which the Study Directo r s igns the final report. After such time, the Sponsor will be contacted
and his advice sought on the return, disposal or further retention of the materials. lf requested,
PEJ\ PC will continue to retain the materials subject to a reasonable fee being agreed with the
Sponsor.
8 HEALTH&SAFETY
ln order for PEA.PC to comply with Law of the Pe912le's Republic -e,f_Ch ina on th<;:_ Prevention
and Treatment of Occupational Diseasec~2001, and the current Contro l of Substances Hazardous
to Health Regulations, it is a cond ition of undertaking the s tudy that the Sponsor provide PEJ\ PC
with all information availab le to it regarding known or potential hazards associated with the
Page 111 of 19
FRD903: JNIIERENT BIODEGRADATION Key Lab of Pesticide Environmental Assessment and Pollution Control, MEP
Report No.: R2009NC03 I (a)-02 DPl38823 I
handling and use of any substance supplied by the Sponsor to PEAPC. The Sponsor also
complied w ith all current legislation and regulations concerning shipment of substances by road,
rai l, sea or a ir.
Such information in the form of a completed PEAPC test substance data sheet must be received
at PEAPC before the test substance can be handled in the laboratory.
9 REFERENCES
I) SEP A. The guidelines for the testing of chemicals. HJ/T 153-2004. 2004.
2) SEP A. The guidelines for the testing of chemicals. Beijing: China Environmental Sciences
Publishing House. 2004.
3) GB/T 21818-2008. Chemicals-Inherent biodegradability-Modified M lTI test(II) . 2008.
4) OECD. 302C"Inherent Biodegradability: Modified MITI Test (II)" . Paris: 1981.
5) SEPA. The guidelines for the hazard evaluation of new chemical substances. HJ/T 154-2004.
2004.
6) SEPA. The guidelines of chemicals testing good laborato ry practices. HJ/T 155-2004. 2004.
Pagc l5o1'19
FRD903: INHERENT BfODEGR/IDATION Report No.: R2009NC03 l(a)-02 Key Lab of Pesticide Environmental Assessment and Pollution Control, ME~-- _______ DP I 3 8823 I _ __ __ _
TABLES
Table 1 Results of BOD -- --·- ·----
Sample BOD after n days (mg/L)
0d Sd 7 d 11 d 14 d 18 d 2 1 d 25 d 28 d - - - - •- --
"abiotic" 1 5 8 9 9 10 16 22 22 - --
9 14 17 26 31 37 40 46 52
"test" 8 13 16 24 28 34 36 42 48 - ----
8 14 17 26 30 37 39 46 52
"rnference" 13 115 139 189 210 23 1 237 245 254
"blank control" 8 15 18 27 31 37 40 46 52
Table 2 Biodegradation as BOD/TOD
Sample Biodegradation after n days (%)
--5d 7 d 11 d 14 d 18 d 2 1 d 25 d 28 d
--I < l < I < ] < l < l < I < l < I
- · - ----- ~ - - ·-Test substance 2 < l < I < I < l < I < l < I < I
(PRD903) 3 < l < I < l < I < ] < ] < l <1
mean <1 < l < ] <1 < ] <1 < l < 1 -
Reference (aniline) 41.5 50.2 67.2 74.3 80.5 81.7 82.6 83.8 -- - --·--'--- --
Table 3 Anal tical results and De radation of :FRD903 abiotic abiotic
"test" (FRD903, 28days) (0 da 28 da s
r---:_-=D=:=:-.:-d:=_:n=i:=_1:I=~_(~=1/o~-)~--_ -+-+ -_ -_~_ ~-3=1=. l===:==-~~=3~0-.8~ ~ = _::: J -i ~~ =t :::S
sample
Average(%) 1.52 - ---- - - - ----~--- - - - -
\
•
Page 16 of 19
FRD903: INHERENT 131ODEGRADi\TION Key I .~h of Pesticide Environmental Assessment ~nd Pollution Control, MEP
FIGURES
90
,...._ 80 ?ls ,✓
C: 0 70 · rl ..,
"O «l
60 l--1 bO O.J
"O 0 50 ·rl
c:o
40
30 -+-Reference substance
20 ---t!-- test substance
10
0 -
0 10 20 Time (d)
Figure 1 Biodegradation curve
180001
.. :I " j
' 10000
J 1 :1 /
•ooo
0010 OOIS 0010 0025 0 0.\0 0035 00-'0 OOJ5
Figure 2 Work curve
l'age 17 of I 9
Report No.: R2009NC03 I (a)-02 DP1388231 - - - - - - -
30
I t
I
FRD903: INHERENT BIODEGRADATION ~ey Lab of Pesticide Environmental Assessment and Pollution Control, MEP - ·-- - ~
U31a-J02~-1 O:\la-a .:..J-0.005ppm
100 0.64
Report No.: R2009NC03 I (a)-02 DPI388231 - - - ---- -- - - - - - - -
Figure 3 Chromatogram of FRD903 (0.005 mg/L)
>.OtCOl '..:0-13 0'.lla-3.Uc.-1-0c
\00·
0.08
/\-'-'Y'
0.63
0\
..........,_._,.~ ~.......--1..-,-•~•.,...-.1 ~ - ......- ~ - - min . 0.20 0.<10 060 0 .80 1.00 1.20 1.'-0 1.00 1.80 200
~:uo:>1 :;:o..o<J 011i-'.El:::.c -1-';"/;';l
100
Od
0.64
0 20 o.:o ow o.eo 100 1 20 1-10 1Go , ~o *;?0-~··1
28 d
Figure 4 Chromatogram of abiotic Sample at 0 cl and 28 d
Page 18 o r 19
FRD903: INHERENT BIODEGRADATION Key Lab of Pesticide Environmental Assessment and Pollution Control, MEP
2010')1::0-0!l 0 31&-30le -l -.'.~,:;
100
or
ro,
1
1\ ~,,, '
\ , V
0 .. . .. 0 .20 0.-10
Report No.: R2009NC03 J (a)-02 DPl38823 1
Figure 5 Chromatogram of Test Sample at 28 d
:-.;1.;01;:0 -05 O'Jl 'l-~e-5-ef.:
·11t 0.71:J\ ro·Jv, 1.09 0.:2 \) \ V'V\ /\ I 1
\ ,,. Y1
I / f\JI,, "' " ' f
I v \ 1.44 r\J/\ ,. u,1 ,.9l
, VV ' ''-\/\,j'l i i \ ) II J
I) -~--, 120 160 . • ,.80 • min
1.00 0.20 040
ITigure 6 Chromatogram of Blank Control
!' age 19 o I' 19