Document NE9g272Lmr7aVr6nE10e55DwV
Hu ^
AR226-3338
FINAL REPORT
Bioaccumulationtest ofPwftooroalfeylcarboKyliaccid (C=7-'13) (Thistest i$ perfbrmed using Perfluorooctanoic acad (Test substance nuniba- K-l 519)] in carp
(T&slNo.51519)
Kuruae Laboraioiv Chemicals Evaluation and "Research Institute, Japan
55519
STATEMENT
Sponsor Title Testnumber
Kurume Laboratory Chemicals Evaluanon and Research Institute, Japan
Ministry of International Trade and Industry
Bloaccaimulatinn test ofPerflnnrnalkylcarhoxync acid (C=7-13) {This test is performedusing Pafluorooclanoic. add (Test substance nuinber
K-l 519)]in carp
51519
J, the-unricrs!5ned, hereby dedarfc thatthis repent prnvide!. a cra-rect 'Pjiglish translation
ofthe Final Report CTesi No.51 S19, issued on December 1S. 2000}.
Uale Study director
^lAy / 7 / -^^ /
V-i F.^1,-V r-. f ,'-!^fet..fl-t^).'. a -
lyina IKilnishiiTia
STATEMENT
51S19
Sponsor Title Test number
Kunnnc Laboratory Chemicals Evaluation and Research Institute, Japan
Ministry of International Trade and Industry
Bloaccumulation test ofPcrfluomalkylcarboxylic add (C=7-l3) [This test is pBrfuiniedusing Perfluoroucianolc add (Test iaAstance number K-l 519)] in carp
51519
This final report (copy) was made the oopY accurately oflhs final report on above the
test.
Date Management
December IS, 20DQ Signedin ori^nal Hiroshi Tadokoro
51519
GLP STATEMENT
Ktinina Laboratory Chemicals Evaluation and Research Institute, Japan
Sponsor
Ministry of International Trade and Industry
Title
Bioaccumulation test ofPerfluoroalkylcarboxylicadd (C=7-l 3) [This test i$ performed using PerfluorooctaDoic acid (TtisE subslanue uurnliiir K-1519)] ui carp
Test number
51519
This test was conducted in compliance -with Good Laboratory Practice Standards for industrial chemicals, "Basic standards to be observed by testing facilities in conducting tests stipulated in article 4 of the Order Prescribing Those Items ot'the Test Relating to the New Chemical Substances and Study on Harmful 'Effect!; of Designated Chemical Substances" (March 31, 19S4, Revised March 1, 2000, Kanpoayo No.3?, Planning and Coca-dination Bureau, EnvironmCTi Agency, 'Yakiihatu 'Nh 229, Pliaimaceulicai Affairs Bureau, Ministry of Health and Welfare, and 59 K'Ayoku No,S5, Basic Industries Bureau, Ministry of
1'ntp.rnatinnal Trarip. anri farliistry. lapan) anrl "DRm Principies of Rnftd LabaratOlY
Piractice" (November 26,1997).
It has been confirmed that this final report reflects the raw data atsianalely and ihe test
dala are valid.
Date Study director
December IS, 2000 Signed in original jaina JAjkushnna
51S19
QUALITY ASSURANCE STATEMENT
Kiunime Laboratory Chemicals Evaluation and Research Institute, Japan
Sponsor
Ministry uflnluiiffllianal Trade and Industry
Title
Bioaccumulation tea. ofPerfluaroalkylcarbosylic add (C=7-13) [Has ted. is pa formed using PwfluorooctontHc acid (Tost substance number K-lSl&)]inca[p
Test number
51519
Study audits and inspectionsofllus test were earned our and the results were reported to the managciwnr sad the study dirsclor by Qualiy ^SHirance TTnit ofKuniiRS Laboratory, Chcniieals ET'-aluation m.id Research Jnstinrte, Japan os follows.
Item of audit or inspection . Protocol
Test Conduct
Raw Data and Finel Report
Data of audit
or inspection S&ptebar7,2CIOO
September W. ?.000 September ^8,2000 Novcaabcr 2B, 2000 September 12,2000 0ctober 16,2000 October 17,2000 October 27,2000
Novemhia 14,2000
December IS, 2000
Date of report
to study director September 7, 2000 September 18,2000 Scpi.einber2S, 2000
K&Vffli'iber2S,2000 September IS, 2000 Ot;luburl7,2000 October 17,2000 Nuvialibw 15, 2000 'Movemh&r 15. 2000
ni-r.cmIwIS.'Win
DaTe of report
to management
September S, 2000
Se.ptc.mber 18,2000 Septembel 28,2000
November 28,2000 September 21,2000
October 23,2000
October 25,2000
E
November 16, 2000
November 16.2000
December 18.2000
has been assured thai the final report describes accurately the lest method used, that details in the report are in compliance with the protocol and Standard Operating Procedures and that the final report reflects the raw data accurately.
Bale Quality AssurancfrLinit, Head
December 18,2000 Signed in original KyoshiroHori
CONTENTS
Title Sponsor Testing facility Objective Test method Applied GLP Test schedule Storage of test substance, Taw data, etc, Personnel Prepscauon of final repnrt
SUMMARY
1. T&st substance
2. Performance of acute toxiraty test 3. Perfonnaiwe ofbioaoeuoiulatron tea 4. Factors possiblyairecling accuracy
5. Results 6. Remarks
51519
Contents of tables and figures
Contsnts oftables
Table-1
Measured concemral.ions of test substance m tesfwater (see text)
Tabla-2 Tb!n 3 TaUc-4 Table-5 Table-6 Table-7 Iafrle-8 Table-9 Reference I
BCFs(seetexi) Concentrations oi'11-.ii ia ih'ita.nc.ft m te^t CTira ai a stcady-stae (seetert!) Calciilartion lable for analysis of lest water (Liwal 1) Calculation table for analysisof test waler (Level 2) Calculation table (b) recoveiy and blank ussi (,'analyy.1o; flest fish) Cal&ulaiioii labis fot'analysia &riCa-t fish (T.evd 1) Calculfllion lable for analysis ofTest fish (Level 2) Calculation tabk fut analysis of lea fish (Control) Analytical results ofditutinn walt.'r
5151?
Contents of figures
Ttg. 1
Correlalbti between exposure period andbbmncentraucm factor (Levd I)
Fig. 2
Coirelation between exposure periodsnd bloeoncentrarionfactor (Levei 2)
J'ig.1
rig. 4
Fia. 5 Fig. 6 Fis. 7
fig. 8 Fig.9 Pig. 10 Pig.!!
Pig 12 FSg.13 Fig.14-1 Pig. 14-2
Conosatration mortality ourva
Calibration curve fnr LC-MS analysis (analysisof test w/ater)
Mass ctiroinatograrn ofLC-MS analysisforanaJysis of test water Calibration curvefor LC-MS sinAyys. (aiitilysw "I'leat Csb)
Mass chroinatogram of LC-MS analysis for recovery .'ind blank test (analysis oftest fish) Mass chromatogram ofTX-MS analysisfor test fish (I-evet 1) Mass chiuniatogram of LC-MS" analysis for test fish (Levd 2) Mass chromatogram of LC-MS analysis for test fish (Control) Concentration of dissolved oxygen (Level 1)
Ccmceniration of dissolved oxyeen CLevel 2) Concentration of dissolved oxygen (Control) tR spectrum oftest substance measured before -the experimental start tR spectrum of iea substance measured after ihe experimental completion
fig.15 Reference 2
Mass spectrum oftest substance KNifR. spectnini oftest substance
51519
Title
Bioaccumulaiion test of Perfluoroalkylcarboxylicacid <C=7-13) [ThSs test is Derfbnned using PtafluorooCTanoit: aa'id (Tea substance nunilserK-1519)] in carp
Sponsor Ministry of International Trade and Industry 1-3-1 Kasunrigiiselu, Chiyoda-ku, Tokyo 1UO-8901, Japan
Testing facility KurLirne Laboratory Chftinicals Evaluation and Researcli Instnute, Japan 19-14 Chuomachi, Kiliui-ne-sht, Fukuoka S30-W23, Japan
Objective This lest was performedto evaluate the bioaccumuladon potential orE-3519 irt carp.
T&si tncthod
This tea was conducted according to the "Method for Testing the Degree of
Accumulation of Chemical Substances ft 1-ish Body" stipulatedin the "Teams Methods for
h'ew Chemical Substances" (July 13, I974, Kevised October S, 1998, Kanpogyu No-5, "Haiaang aflii Cciotifttation Durss6u, Snvnon'mtsal Ascasy, YakTihao No 615, Pharmaceutical
Affairs Buriu. Mintstr}' ofHffflth and Welfare, and 49 Klhyoku No.392, Basic Industries Buir^su, Minisliy of fateriKtional Trade and Indiiauy, Japan). This test method is
oaBeniially the same fs -that 5n tlift OFm Guidelines for Testing of Chemicals,
''Bioconcentration: Plow-through i-lsh Test (Gjidelina 305, June 14,1996)".
-1
51519
Applied GLP
(1) Chemical GIF
This tefl complied with "Basic standards ta be observed by t&simg facilities in conducting tests stipulatedin article 4 of the Order PrescribingThose Items of the Test R-deting to the New Chemical Substances and Study oh Hannfal Effects of DesignatedChemical Substances (heieallci icfarcd 10 as "GL-'P standards")1' (March
34 19H revised March 1, 2000, Kanpogyo No-39, Planning nod Coordination
Btlieau, CrtvH-ooment AgllBOS't Ynkuhftlu No.3?-P, IPiiartriBce-urical Afiaios Tirrrpaii,
Ministry of HealA and Welfare, and 59 Kikyobu No.85, Basic Industries Bureau, Ministry of rTitemational Trade and Inclustri., Japan).
(2)0CD-<jU
'11ME test complifid whh
(No^'ernber26.1?P7),
"OECt iPrlnctpIes
ur Ouuti
Lahoiatoi-y
fractioc"
Test schedule
Start of test Experimentalstarting date Experimental completiondate End of test
September7.2000 Octorbsr 17,2000 Novefflber 14,2000 December 18,2UOO
Stoi-agfi oftest aA-stance, raw data, etc.
(1) Test substance Becausa the test substance of ttie same lot is already stored after the end of
Ijwilcsnacl.ntioiHCTi, lliii lest ul0tanno lo not StiyccI oftar+hie tBrt
Rw (2)
data aad materials, etc.
Raw dala used for the preparadonof the final report, the protocol, documents
aboiA the test presented by the sponsor, the copy ul' Lhe Hnal teport and necessary
materials are stored in a storage loomin'this latiaraiory.
-2.
5)519
Personnel Study director Personal (Operationofbioaccmnulaticm t&si.)
Staff for fish cara Person to coaduci- nf Hsh acute loxicrty i&si
Preparationof final repoit
Study director
Da-ta Signanire
FrinaKikuAimB RieObo Yuji Kusune KazuCimi Katagm Yiiirn Ka-wrashima Yasuro Kawashima Katsurni Kiyono
Decsmber 18,2000 Signed in uxlainal .ErinaKJtosiiiffla
51519
SUMMARY
TOR
Bioaccumulation lest of Perfluoroalkylcarboxylicacid (C=7-U) IThis lesi is
perfoirtrnsd using Pi;rfh.it>'-c>(v-rBiwii- ar.iri (Taw iiihyanca mimhcr K-151'9)] in carp
Test conditions
Acute toxicity lest
(1) Test fish (2) Duration ofCTposure (3) Exposure method
Orange-red ki] lifish (Ofyzias latipss) 96 hrs, Semi static sysleni
(Kenevi'aJ of lesi w'aier, at way 8 - iff Brs.)
Bioaccumulation test (1) Test fish
.
CaTp (fyprimis carpinY
(2) Noironal concentrations of test aibslan&e
High exposure level (Level I) 50 (ig/L
Low exposure level (Level 2)
5 ng/L
f31 Duration of iposuie
28 days
(4) Exposure method
Conthiunus finw .'iysiem
(5) ATlalytical ittfithod
High-pErfortnan&a liquid chrornarngraphy-Biass
speclromefry
ResoJts
0) 96-hour LC50 value
100 mg/L
(2) BiucuiH;t!iittai-iuu 1'dutui til a Mcady stzne
Level l
(3) Bioconcentrarion factor? l.-evel 2
3.1
5.1 - y.4
-4.
1. Test substance
In this report, K-1519 has the following chemical jnaroe, eic.
1.1 Chemical name
PerQuoronctanoic acid
1.2 Chenricat structure, etc.
SiluC-tiiral fonnilU.
c.F^--{r.Fs)5--cnnH
Moleciriar fmmvia
CsWaOz
Molecular weight
414.Q7
1.3 Manufacturer, samcf and lot iiuinber''
(1) Manufacturer
. ;UAlK.lKMnecHeniicalLa&ofarojy,ud.
<2)Name
&1700
(3) Lot number
C170020l
* 1 fafonriation supplied byth,e manufactnrBT
1.4 Purity'3 Test substance
98 % (GC)
The test substance was treated as 100 "A ro purity.
51519 1,5 Confirmation of-lest substance
The sInicLure ul'lhe teal substance if. uunT]imiaJ by juliaied (1R) spectrum Cscc Tig. 14), Mass spectrum (see Fig.13) and NMR spertnu-n {sea Reference 2). ),6 Storags and stability (1) Storage condition
Cold storage plaee (2) Stability
Thfe test ?ulistanc& was stable under the. sto'age candmon as shewn hy As finding that IR. specira of the cea substance befora and after the experiment "/ere
identical (see Fig.14). 1.7 Stabilityunder testing conditions
Prior to the bioacctimuEation tea, s stabiln.yof the (e&l substance under the testing conditions \vas cDAFuuied by a preliminar)'lest.
-p-
51519
2. Performance of acute loarity test
2.1 Test mated
oerformt.d in accordance with Japanese Industrial Standard (J1SK
The test was 0102-1998-71-), ''Testing methods for
industrial
wsste
water,
Acute toxicity
lest
with
fish1'.
2.2 Test fish
(1) Species
Olaagc-red killifisfa (Oryaasiatirigs)
sensitivityto carp and
Reason for selection : This speciesis simHar in
readily available as test fish.
(2) Supplier Nakashima fish fara (Address: 202& Ooaaa NBflBsu, Kagasv-fiho, Taraana-gun, Kumaimoto
869-0123, Japan)
(3) Conditions fbr fish care before acclimatization
those demonstrating any
The fish were checked visually at the riiceiviiig and
abnormality were removed. The fish wa-e rcsared for 39 days in a flow through
sys.1era following an tocteraa] dislttftrtioT).
(4) Caadstiaas for acclimatization
acdimalizing aquarium and
A&cr rearingi the fish were transferred lo an tismfeciicm,
The fis-li dcreonstrating
acclimatized there after the sewri estarnal
abnormality during-this period were removed and the remainder ofthe fish were
aTtn>.yBrp;fl fnr 24 days in a flow throup.h .lYSiaenind arte(ahreedteamnpoethraetru4re1o-f4235 iAliiy2a"Caf.terTthhee
fish -were checked far health conditions
external disinfection.
(5) Weight
average 0.20 g <foradditional test 0.19 g)
(6) Length
average 2.9 cm
(7)
Certification The 48-houi
LC50
value
oftbc
rc-rcrance
substance."fcr
the
'fish
of the
same
Sot
(TTO-OflfWi^wasRW nis/T.
*2 PCP-'h'a (pentachloropbeosoodl iiiin sail, Tokyo Kasci K&gyo Co., Ltd.)
-7-
51519
2.3 Dilution waer for test
(1) Origin T InrfprgTrniTiit water from the nreiniies of Kiinime Laboratory.
(2) Water quality assessment The dilution water for rest was taken out on August &, 2000, and it was
analyzed and measured (once every ax months in this laboratory). The results are
stiywii 3u iRuIbicnco 1.
It was confirmed that the dilution water met the ministerial ordinance of the Nnnstry of Health and Welfare (December 21, 1992). water quality criteria for fisheries (Shadanhoan TMihon Suisansiaen Hogokyolcai, March 1983), OECD Guidelines for Teaing of Chemicals, "Fish, .Barly-life Stage Toxicity Test"
(Guideline 210, July 17, 19S&) and stivironnienial quality Standards tor wa.et polluianit No. 14 (Kcviscd Febmary 22, 1909, Environment Agency) or OECD GiiKieliBes for Testing of Chemicals, "Bioconcentraiion: Flow-through Fish Test (<3udelinci3fl:5, June 14,1996)"
2,4 Test conditions (1) Test tank
Round glass wssul
(2) Volume oflest walcr
4 L/level
(3) Temperature oftest water At initial exposure
Before renewal of test -water
24.4 - 2.4,6 C 24.3 - 24.7"(/
(4) Concentration of dissolved oxygen in test waier
At initial exposure
7.9 - 8.1 nag/L
Before renewal oftest w.'ater
7.1 - 7.3 mg,'L
(.5) pti ur lcl wuLia At initisi exposure
Before rciitwal of test water
7.1 - &.0 7.5 - 7.9
(6) Number of fish
10/level
(7) Duration of exposure
96hra.
(8) Exposure method
Semi static syaein
(Renewal oftest water, at even' B . ]<5 hrs)
51519
2. > Preparationof Stock solution
(1)Dispersant HCO-60 (IIydrogaiated castor nil)
(2) Preparation The iesc substance and HCO-60 (50 limes amount of the test substance)were
dissolved with aceione. After Ae acetone was evaporatedfrom tfie solution, i-on-
cwhBtigod water wo.a ttdd-cd to th miKturo -to prepare 10(.lft mayl. noi-lc soilnl'u-in
2.6 Pertormancs of test
(1) Place (2) Date
214LC50room September 11,2UUO - September 17,2000
2.7 Estimation ofS9-l30i4rLC50valus The 96-hourLCSO value was estimated by the Doudtoroff method.
2.8 Result Dftest 96-hourLC50 value
1QO rog/L (BBeFig.3)
-9-
51519
3. Performance oibioaccuniulation test
3.1 Ted fisli (1) Species
Carp (Cyprinustai-pio) Reason for selection : The previous data conducted -with this species
can be compared and the size of this speciesis
ad.iua.te for fcandling.
(2) Supplier Ftikyolsa'bakYa&e^'safchertnfen's co&yasaiwfc a&sowAioa (Address: 7t8 YalnaucH, Yame-shi, Fukuoka R'U-OOL?, Japan)
Date received
Seplemocr l, zoro
(3) Conditions fia Mf care before accBmatization
The Hsh wsre checked visually in the receiving and those demonstrating any abnormality were removed. The fish were rea-ed for 8 days in a. flow through system following an external disinfection
(4) Conditions for acclimatization After reaiiTig, tha lish weie niediuatud lu duiuiialc paiasites and lianareiied lo
an acciimatizins anuariuni. After Ihe second extenial disiiif&Ction, they wcr6 acclimatized, The jt5sh dcmon,wating liny ahnnnnality during this perioti ware removed and Ae remainder of the fish were reared far 35 days in a flo-w through system at fte temperature of 2S = 'i "C. The fish were then transferred to rest tanks and reared at the same- temperature in the flow through system for another 19 days,
foil owing ttw eKtcmal riismfcrmm
(5)Lirngth
6.8-8.$ cm
(6) Lot No. {7) Age
TFC-00&901 Yearling fisis
<S)Feeding
Feed Compoation
Feed for &y of taip
Protans content s
Lipidcontent S
4,).uy 3.0%
Manufacturer
NipponFormula Feed Mfg. Co., Ltd.
Feeding amount and interval Amount corresponding to 2 % of total botfy weight was fed twice a day in
halves.
T\it&^^t^%TO<dfcil4tottx^!i!^ara3atB(plms,
- 10-
51519
3.2. Dilution water for tea The same as describedin 13.
11 ("nuditioBs oftest and circumstances
(1) Supplyof test water Flow throughsystemassembled a this laboratory was used.
(2} Teal tank 100-L glass lank
(3) now 2ratme ur^l'nleiisnt wfoetrcrstock solution and 800 mL/min for dilution waia' ; 1155
litersftjay for test warer were supplied.
(4) Stock solution bottle
25-T. glass bottle (Frequency of renewal
about 2 times/week)
(5) Temperature ciftestwrter
Level 1 Level 2
24J.25.6C 24.?-25.4 "C
Control
25.2-25.&C
(6) Concaura.tbns of dissolved oxygen inisa water
Level 1
79-8.1'iBE/L(seBF)g.ll)
Level 2
7.9 - 8.1 mg/L (5e&Fig. 1-2)
Control
8.0-B.I nig/L (sec Hg. 13)
(7)pB&flcstwaler
Level 1 Level 2
Control
7.7 7,S
7.7.78
7.6-7,8
<S)Time of irradiation with light Artificial (ighloi'v.luiefliiorEicem lamp (14 hrs.fUay)
(9) Number offish (at the beginningof exposure)
Level land 2
2S
CAntr&l
S
(10) Dm ttion of asposure Reason for decision:
28 days The time ibi leaching a Steady-state was ea-imaltid in
2S days from preliminary test resulu.
-El-
51319
(11) P l a c e
213 Aquafron room
3 A Preparationof stock solutions
(1)Dispersant The s&tiie as described in 2.5 (1).
(2) Preparation
Level i
20 mg'L stock solutionwa? preparedin the same way as dRscribed in 2.5 (2)
Level 2
2 mg/Lstock solution was preparedin the sam6 way ss described in 2.5 (2),
Control HCO-6Q was dissolved with ion-exchan?,od wafer TO prepare 1000 mg/I.
stock solution,
3.i> Test concentrations Based on preliminary test results for the SiS-hour LC30 value and analytical
detection linuta, test Ctulceatrations oflhft lost substa'tee were decided as follows. The
control was set as a blank test-
Level 1 Levei 2
30 ijig/L S ^ig/L
-12.
51519
3.6 Observation, measurement, etc.
(1) Observation of test fish
("rmrittitHi (iFtest fish was observed Msoally twee, a day.
(2) Flow rate of test-water Plow rate of stock scArtiftn and dilution water were measured with graduated
cylinder and recorded once a day.
(3) TfiinperaiurB of tea waier Temperature of test waicr was ra&iisured with alcohol thermometer and
recorik.d once a day.
(4) Cosc-wSratsoa of dissolved oxygen to lest water Concentration of dissolved oxygen in test waier was measured with dissolved
oxygen prole and recorded twice a wedc.
(5)pHoftestwat<r pH of test water was measured wirh pH meter once a week or more.
(6) Cleaning or test tank
In cxperirnetnatperiod, excreta of carp, din on test tanlc, etc. were removed
about once a day.
3.7 Analysis oftest waia" and fish Analysis of the test substance in test water and test fish was performed with high-
performance liquid chTOmatogra.phy-iiia.ssSpfictrometry (LC-MS) analysis.
3.7.1 l-'roiquericy of analysiu;
(1) Ten waier analysis The test v/aier of each lev&l was anal^ed. once before first analysis 01 test
Sail o.nd at the auma lime sis onalyi.iK til' tiisi Kail. Ona eaispla of each iisvel wa analyzed at each szstpi'jsgtisw,
(2) Test fish analysis AnalysiB ofteit fish was performed fivft rmnea at each level in duration of
exposure. Analysis oT comro! fch was perfhnned bcrote ilic cxperiinenial starting and at the experimentalcompletion,
Four fish were taken out at each sampling time and divided into two groups, lhaii both were analyzed individualiy""'. ^ Because one nsli was loo smal] to taice out tbo slor&d SBinplc. for ihfi
measurement oflipid content, two fish a group were eroployed.
-13-
51519
3.7.2 Prefreatment foranalyas
(1) Test water
An Aliquotofthe test water,
Level !
) mL
Level 2
2.5 mL
was laken from each test tank, and prsirealed for high-periwniiince
chTumalvgrnphy-rnnns Rpcctrotnclry (LC MS) analysis W Icllows :
liquid
Tillitig Up 10 10 mL Cwata" for recuvay lesi, voliBaetric flask) (only for Lcvd 1)
Taking out 2.5 raL (transfer pipene) (only for L<;vel 1)
rilling up to 10 mL (water"',volumetric flask )
Sample forLC-MS analysis
*4 r.ily wfster was treated by IHira Rtire water a'stem.
.14-
51519
(2) Test fish
Test fish were taken from each lest cank and pretreaSedfor high-performance liquid chroinatography-niassspecc-oinetiy (LC-MS) analysisas follows:
preafishj
Measurement of weightand body length Choppinginto pieces Making sample fitsc (polvtron. 2 min. 01" more,
on ice water)
|~Fmesanrple ';
L-
^
~
~
-Takinsoinl-Sg
(analyticalbalance)
Sample for storage
Taking out S g (analyticabl alance) ^--Acetonilritc 20 mL (graduatedcylinder) 1 Honu)genization(potylron,about 1 roin.)
Washins (acetonitrile7 mL)
CcntrifiisatiDii (7100 x <?. 5 min,)
Residue J
Supernalant
F'lltration (dbsmbart cotton) Fitling up to 50 mL (ac-Rtonhrile, volumetric
nask) Taking out 1 fflL (transfer pipette) Pilling up to 25 mL (water"4, TOlumelric
flask)
Sample fa-LC-NtS analysis
-15-
51519
Quantitativeanalysisfor test substance The samples for LC-MS aniily.sii, in pidreatment were analysed by high-
performanceliquidclK'omatos?aphy-niassspectrometry under the following analytical mnditmns The concentration of the test substance in each sampfe solution was dBMa'iained onltebaMSofacoitapariyinofthcpcakareaonthernBSSchr&inatograniof ihe sampk solution with that of a standard solution (see Tables-4, 5, Fig.5 and T&bles-
7,8,9.Figs,8,9,10).
(1) Ajmi^tii-fll oondltiona Instrument
Hish-penomnance
mass spectrometer ffigh-perfonnanue liquid chromatograflh
Mass spectrometer
Waters Coiporation W&lcrs (.toTporation
iiquid cliromatogiaph-
type2690 typeZMD
r-ondinons fifhigh.perfbrmanee liquid chmifiatograph
Column
Column temp. Elucmi
L-cotunui ODS
15 cm x 2.1 mni (.D. 40 "C
slaiulcss st&sl
A (60''D): Acstonitriki
B (40%5: 5 minol/L Di-fT-butyianiine acetate
Flott' rate Sinnple sias
0.2 mL/nun, 20 llL
.ConditinTis ^fniass ypertrpmo-sr
JocteaaoajBodc
'RSearaspray
Detection bn
Negative ion
Co"? ^filfasR Source bloukicmp.
1 fi V
]20C
Desolvation tenip.
3 00 "C
DesOlvaiion gus flow rate 3 50 L'ltf
McnitorinEpon
miv. 413
If;-
S1V19
(2) Preparationof .standard solution TKe standard salmon. to determine; the concentration of the tea substance ;n
tie samplesolutions was preparedas follows.
(a) Analysisoftest walei 100 ms, ofthe substance was dissolved in acetuniuile to prepare 1000 mgi'L
solution of ihe lest substanca 2.50 itg/L- Standard solution was then prepaid fbom this solution by dilution with water'1' i' -water for recovery lest (1/1 V/V).
(b) Analysisof test fish 100 ffig of the SLibstenisewas dissolved in acetoTiiliile 10 prepare 10UOmg/L
solution of the lest substance. 2.00 ng/l- standard solution wius then piepared from tilis solution by dilution with water"'1.
(3) Calibration curve
(a) Analysis oftest wala 1.25. 2.50 and 5.00 pg/L standard solutions were prepared by the same
nietiiud as described in (2)(a). These solutions were analyzwi according 10 rtie analytical conditions described m (I). A calibration curve was drawn on {lie baas rtf-the relation between tile peak area on the mass cliromatograrns and the respective concentrations.
In consideration of the noise level, ihe lowesi detectable peak area of the test substance was regarded as 250, which corresponded to 0.10 HE/L oFthe test subiiuiiicc Goncentrntion (see rig, 4),
(b) Analysis oftest fish 1 00. 2 00 and ''1.00 HS/L standard solutions were prepared by the same
method as described in (2)(b). Theas solutions wan analyzed according to the
analytical conditions described in (^]. A calibration curve was drawn on the baas of the islalicm bst-ween the peak area on the, mass chromatQgranis and the respective concentrations
la fonsidoi-RtSo" of the noiEt- level, ih^ !(wpc!t riatprtahic peafc area of the
test substance was regardedas 300 for analysisoftesi water and test fish, which correspondedto 0.092 ngT- of the test substance concentration [see Pig.6),
-17.
51519
3.7.4 Recovery and blank test
(1) Method Chopped fish 00 a) spiked a specified amount of the tost substance fitf the
recovery tast, were preparedvo the same way as described in 3.7.2. Tlie blank tea was ai-sn performed in the safflfe manner without the test substance. The recovery and blank tests were performedin duplicate.
(Z)
Results ofrc.ooYCTy Icat In the blank test, die
mass
chromalogiam
ofJL.C-MS
had no
peaksinterfering
with rietennination of the test substancii nonceTilrauon. the duplicaterecovery
rates and the average nf them in the pretreatoient are shown hetaw (see Table-6
and Fig.7). The avCTage recover}'- rate was used as cori-edion faciors for the
determination ot the test substance coaceniratlons in uie analytical Seunples,
For analysis oftest fish (5 iig leal. substance added)
$6.4 %,
94.3 %
average 95.3 %
3.7.3
Lipid content in tea fsh Upid contents in the samplefor slorage of the conirol test fish (see 3.7.2(2))
were determined after chloroform-nisthanol extrayl.iuri wilb gi-Bivinietricanalysii.
-1-
51519
3.7.6
Calculation of the test substance. nnncentralJon in sample and nunimuTO limit of
determination
(1) Calculation of the lest substance concentration in tea water Tlie equationin Tables-4, 5 was ustid 10 obiain tlw cnncerttratkms; and they
were rounded off1o 3 figures.
(2) Determination limit oflbe test substance in test water The dcnann'ina.tion limit*1 of the test sufcstimr.f ir> i-it water was calculated on
the basis of that obtained from tlie calibration curve in 3-7.3 (3) as Ibltows.
Lev-all
2.1 ug/L
Level 2
0.21pg/L
(3) Calculation ofliiu lua'l yub!?laciee wncoiriratiun in test fish The equationsin Tables-7, 8, 9 were weS to obtain the cnncentratkms, and
they were rowded ofi'lo 3 figures.
(4) DaertdnatioB liinu of" (he test subaance in test fish Assuming ihe tine sainple of flsh to he 5 s, the detefmination liniii^ ofihc U-si
subsiancc In test 5sh was cakafcteA to te 24 n&'g, OB tb& ^asis oE that obtained front the calibration curve in 31 3 (3).
* 5 Minimuni determination limit of the test substance (iis/i- or ng/g)
A
" _
B_ x 100
CxE
D
vipers
A : Ivlinimum detenrnTiation limil of the test subafincs on ttie calibratiOT wrw^e./L}
B : Recovery rats (%) C : Sampling volume of test v,'ater [niL) or wcigtit of fine sample offish
(?) D : Final volume of samplesolution (mL) E: KalTO oFltie puilion, used far analysis to whole volume
Results were luuiKlod offfo 2 figures.
.19-
51519
3.7.7
CaiculatJon of mean cowentratien? of ihe tea sukslancfe in test water (duraliua of
exposure)
CM=<Cw(.li+" +C<a)}/n
where
Cwt
n
, 0>.<I)
Cw(n)
The mean concentration o r die tosr substance in tesi water
Number atanalysssftr test water (inesarenrent tunes} (?ftw.p.n<raiinn nfthe iftst substance in Ist anali.'sis oftest water ConceiitraHon of the test substance in fi-th analysisoftest vans
3.7.8
Calculation oFbiocortceffltraiiuit factor (BCF) Bioeoncaitration factor (BCF)u-as calculaied as follows.
(1) Calculation of mean ctinccirtraucni or the tea substance in test water for calculating BCF
Cw" (C%'(n-l)+Cw(n) }/2 (only IstanaIysiRortestfisIi)
"Ov - ( Cw(n 2) * Cw(n 1) + Cw(n) } / 3 {&om 2nd analytis oftest fish)
where
Cw
Cw(n)
Ihe mean concanili-atron or llie lest substance in test water for
catiiulating BCF
Cono^ratrfttion of the test subetaate "a. -th anatyBiE of tea waca"
(2) Calculation ofbioctiicentrarion faaor
Cf
BCFCw
where BCT
Cf
Uyf
Bioconcentration factor
Concentralion ofthe lest siAslnnce in lest fiah lAc ffiiean fsjacemration of t!w tesc yiSsstaow m calcu.1<nmgDCF
tssi wfser n
(3)Theinaan bioooncentratioft factor in nf-th analyas BCFTO = (BCFa + BCFb) / n
BCym ; Tlie niean biocoiicentratiun factor in fp-lli analysis (number of
individual or group 2 (a,h))
BCFa.b : Each biocoiicentration factor in w-th analysis oflest fish
n
: Number of individual or group in w-th analysisof test fish
ZO-
S1519
379
Confirmation ofths aeady-state was reached It was evaluated that a steady-sltte had been reached when three successive
analysesofBCFs made on samplestaken ai intervals of at least 48 hours were within
5ft % nfaach ntl-iBr When BCFs were less than 100, it was evaluated thai a sleadY-
siate had bees reached after 28day even if BCFs were over J- 20 % oftach otiicr.
Cdimon oflhB steady-aate was reached: V(m-2), V(in-l), V(m} g 20 (%)
V(m,3.)=
I WKni^-'BCT ,--------^----
|
.1 K 100
'
BCF
V(m-i)= j--B--C--F(m---n1)7-TBpC--F --|--K--10i,, 0
V(tn)
BCF(ni)-BCF BCF
x 100
V(ia-2), V(in-t), V(m) -BCF{ni-2), BLy{Pl-3) BCFfin)
_BC_F
: Variatian ofbioconcentattion feeiof : The mean bioconcentraiionfactoimflt-Z,/??-!,
ff?-ih analysis Qflest fish
: {BCF(in-2)-BCF(in-1)+feCFCra})/3
2]
51519
3.7.30 Calculation ofhiocmicentration factor at a steady-State(BLTss) Bioconcentnrtion factor at a steady-state(BCFss)was calculated as follows.
<1) Calculation of the mean concentration of the test substance ITS lest witer for
caEculaling BCT&6
^w { Cw(n-2) T Cw(n-13 + Cw(n)) / 3
where Cws
Cw(n)
The mean concentration of the test substance in test water for calculatingBCFss. which \s caltsulalt-d from (hit successive analyses of lest water before laiit analysis nfttait fish as a general rule Ccateentration of the test substance in n-th analysis of tesr ivater
(2) Calculation of the mean concentration of the tea: substance in test fish a1 a steady-
fit ale
'Os - { Cfl[m-2) + Cf^m 1)+ Cf(m) } / 3
where Cfs
CW)
FB
The mean concentration of ihe test substance in test fish at a steady-
state
The mean concentration of the test substance, from which MS is subtracted, in w-th analysis ofiest fish The arithmetical mean concentration of the lest substance or blanfe in the control test fish analyzed before and afierthe aipenment
(I) CHlcnIa-tinn nfRr.FKH
BCFss^Cfs/Cws
where DCT'sa 'CCs
Cws
Qioconcontrarion. factor ot a stciuly-atate
Tlic incan concsntration of the test substance M test fish at a steady-
state
The mean r.onccntr?ition of thr; Icgrt sdhslance in test water for e&lculaHngBCFss
.22-
51519
3.7.11
Calculable BCF
OathebasisofthsiTiiniiriuin deiennination linui of the test substance; in 3-7.6(4), BCl-s can be obtained wtien BCF exceeds Llie rollowng. The mean uonccnu-alJon of the test substance in teat water obcainp.rt from all the analyzed sample was used to
calculate the following calculable BCF.
Level 1 Level 2
0.51 S.I
3.7.12 CalcuUlaptiiodncoofnltiepnitds uwuenriecnccalculated according to the folbwing equation. Lipi(IcDntettt(%)=(T-TB)/Sx 100
where
Ta Weight of vessel fe) T Weight of sampleforgraviinetric isrialy&ls(containing vessel) () S Weight nfTme sample talccii nut for analysisof" lipid content (g)
3.8 TreatmVeanltuoesf nwuemreerricoaml ivdaoldueosff in acconiainoe with JTS Z 8401:1?99 nils B. The each valueTuhseedcfoonrcceanltcruatlaiotnionvalwuaessuo&fctdhweitlehsotuslurbosutnadncinegtooftef sotnwthaetewr aayndoffisthhewcaelrceurlaotuionnd.ed off <o 3 figures. BCTss values were rounded off co 2 figures.
4 Taninrs; pftwMy affectina accuracy No adverse effects on the reliabilityofthis tsst were noted.
-23-
51519
5. Results
5.1 Concentration of test substance in test water
The mRasured concentration.'! ol' lhe ICSE substance in test water are shown in Tabie-l. Each concentration ofths lest substance was inawtained at more than 92 %
of each the nominated concentrarion. The variation uf ihe concentrations of the test substance was within 20 % ofthe mean orihe measured concentrarioiK.
Table'1
Mcosul cd conccatratkijif of the taal .tubstanco in teat water (Unit - lig/L)
i
Average
After Level
After
After
After
AHei 1 Afica- (Standard Table HS.
Iday 3 days 10 days 16 days 23 days; 28 days
ifcviation)
1
50.5.
47.fi
46.9
48.1
46.7 j 45.8 47.6
4
:.
(1.51)
5
2
463
469
4.61
4.66
4.83
4,61
4.71
5
,
(0,101)
5.2 Bioconceatration factors
RCPs arr ithttu/n m Tatilf-7 BCFs in Table-2 plottedagainstduration (ir exposme are sliowji in Figs. 1 and 2. These BCFs of the test substance ranged from 2,0 to 4.2 at Level 1 and from less
than 5.1 to 9.4 at Level 2.
TaUc-2 BCFa
Lew!
After
After
After
After I After Table F>&
3 days 10 days Ifidays 23 days ' 2Sdays
2.9
2,4
3.0
3.0
A.I
1
$
7
R
2.1
2,5
3.0
2.0 ! 5-3
1 5.5
^5,1
7.7
6.1
6&
2
i
8
0
g5J
gs.i
9.4
5.1
g5.1 ;
-24-
S1S19
5-3 Calculation ofBCT at a steady-biate (BCFss)
5.3.1
Level I
The variation ofBCPs faverage value) after 16,23 and 28 days were more than 12Q % of each other. But because all BCFs were less than 300, it was evaluated that a steady-state was reached within 28tb days. BCFss was calculated on the basis of
these result.
(1) Concentrations vi'tual. eubclcinoc> in test water at a ttiKidy-ctntv The mean concentrations of tlie test substance m test water at a steady-state are
shown in Table-3. Hie mean cunc&Tmrions was about 94 % of nominated
concentration at Level 1.
lable-.)
COflCeiOTaiinnS Oflhc IWl MJbhlllln-c 111ic&l vvWw Bt n ntcady- rtntc
(Unil:|j/L)
Level
Afia-
After
After
Average Table
Fig.
16 days
23 days
28 davs
1
48.1
46.7
45.8
46.9
4,7
5
(2) BCF at a steady-date (RCTss}
BCFss al Le\,'d 1 was calculated as luliews.
Level 1
3.1
5.3.2
Leval2
Because ihe measured concBiitraliona of the test substance in test fish were coiltairiing value wilich were not more than imniniuni deterrninalion limit of tea
substance, BCFss was not calculated.
Because sill BCFs were less than ] 00, it was evaluated thai a steady-state was readied withiii 28th davs.
5.4 Lipid coitifint in test fish The measured lipidcontents in the lest fish are shown as follows,
Bo&rn'iAiatio'nofdpma-ai; After termination of expnsure
'^.10'%
2.82%
5.5 Results oftea fish obserraikm
No abnonnatily in behavior or appearance was noted,
-25-
51519
6. Remarks
Instruments, apparatuses and reagents, etc. ftsf the test
(1) Instruments for fish care
Micro quantitative pump &r supplyingstock solution :
Tokyo RilcaKifcai Co.. Ltd. Instrument fisi measuring concentration of dissolved oxygen :
typeGMW
Kjunn Eclttetsu Sntlush-iaR 00., I-td typeF-302
pH meter:
Toa Electronics Ltd. typeHM-MP
(^) Instruments, apparatuses and reagents for analysisand preparationofstock solution
Instruments and appal aiiaiM
High-performance liquid diromatograph-masi spectrometer
Electronic analytical balance:
Rotary' evaporator; Homogfiiui.er (polytron) Homogeiuzer(autocellniasler): Ceiiirifuge:
s?epagc]6 Shimadzu Corporation type AE>;-200B
Metier Toledo type Al 63 Sarttirius lype 1404MP8
Sailonus typcBPSOlS
AandD typeFA-2000
Tokyn T<ika KSku Co., Ltd.
yamato Scicntcffc Co., Ud, Kincmatica typfi PT3100
type N lype ft?<?
laclaseieido Co., Ltd. typeCM-200 Kubuia Syouji Co., Ud. type 6500
IS pngpiitt
Acetomtrile (HPLC grade):
Purified water' Methanol (extra pure):
Wafco Pure Chemicat Industries, Ltd. Takasugi Sdyaku Co., Ltd, Wako Pure Chemical Industries, Ltd,
Acelone (eKtra. pure):
Wako Pure Chfimicai Industries, Ltd.
Clllwirfbrm (gtinranlced rcascnt) : Kashida Ch<;Tmc.ttl Co., Ltd. Anhydrous sudiuffl sulfatts (extra plire):
"ECalayainaCTiemical rndustries Co., Ltd,
Di-w-butylamine acettfe (Jan-PsurReagent forLC-MS):
HL'U-fcU:
Tokyo Kasei Kogyo Co., Ltd. l.Mkk-0 Chsmlcals Co., Hd.
(3) Instrurneats and apparatuses far coTifitinalioti o! the test substance
Instruments and apparatuses
Infrared speclrophoiometer:
Stumadzu Coiporation type FTIR-82COPC
rS^i-perforiHance liquidchroniatograph-ntassspectrometer:
Waters Corporation typeZMD
2$-
Tablfc-5
Calculation table fur analysis of test water (Level 2)
Sample descriplion
A
Standard LSQflSfl-
TeSlwaKtafierlilay
Stoidard 250 us/I-
Test vater aper 3 days
Standard 2,50^gflL TtstwatsfafierlOdays
Standard 1.53 nsfL
Test water aficr 16 days
Standard ZSOns/L "tea, WUK aS 23 diiy
Slandiail 150fi/L
Tcsl water afler 28 days
fiZS7 6050
6405 fl003
6-243 5756
6234 581'!
6123 SSl'O
6144 5670
Avwags eo&c<nti!iScm ofuat sitbaansw Enasi waicr 4.71 (jti g,T.)
Test No, 5151.9 I
4.S3 4.6? 4.61 4.66 4.S^ f).6l
(S.D. 0.101)
A:Feakaiea A(dd); Siaiiaad saliKKin A(t): Ssiinple
JB; Halm af pon&m useil for analysis
2.5/2.5
C: Filial volume
5mL
H; Volume of lest water takes out
lm1,
11 Conanlt>""in rrf Ifd illhltanw. in tptt watftr (11 efl.)
tP>;(A<t)/A(std))/B?<C/H
J; Avarags cflinctinratton of lest substaate {a test water (^i g/l,)
J =(I(l)4...+lCfl))/n n: Hiiinber 0? tcsl walcr analyses ( n r 6)
I (1): First Miaiyas ot ust watef I ^n); Laa anaiysis ot test water
^2
SK^-f 2 KO |
S.D. -
BX(|I -1)
7 ; Ccm^AntraCiM. of ctmcfwd Bohiiion
^ W/f &/T
SeeRg.5
HoveBsbgr 28,200&
?iame
/gJ OSO
Tablc-4
Calculation table for analysis of test water (Level1>
Sample description
A
Standard 2.50 n g/L Tcs.twBicrafterI day
Standard ISOjag/L i ea waier alter s nays
Standard 2.50^ g<L Test water after 10 days
Standard 2.50igt'L Test water after 16 days
Standard :L50 jtffi,
Test water fter 23 days
SKwiaKl-^.SO^ff'L Test wtter after 2K days
6257 6284
6403 G09f
6243 5S62
6234 iysy
6t?.1 5720
6144 5fl29
Average concentration of teat subBtance in tea wasw A7,6 ( u g'L)
TestHo. 51519
505 +7,6
46.9 4S.; 46,7 45.8 (S.D. 1.51)
A;PeakiRa A(nd); Standard solution A(i): Sample
B : Ratio of portion iistd for analysis
2J/IO
C : Fin 8.1 volume
SrnL
H: Vctanie oftt< WBH.T lAea ow
IniL
T r.rinwinrftrfm oftesc luhslancc in tesi walei (a s/I-l '
I=P'<(A(l)/A(3tl))/BitC/H
] : A-ueragB ciHiceuaatioil oflsti ilibsiartftfr in lesr water { y g/L)
J(I(l)+...+?(n))/n
n; Nufflber oftssi waier analyses (n = e )
1 (1); t-USt aiiaty'SB Of KS! wBr r (n?: Last uoulysisi of na,i waiei
-\ r~^i" sn. -
x 2 v> t
.2Ki>
IIS<(!> -1)
r ; <'rwif>-nirtii !rm n'Tifra jinhiiUfics ill standard aoluliuii SfreFig.5
2.50 U(/L
Novsralier 29,2000
Name f^E 0@0
Table-6
Calculation table for recovery and UaiA lesi (Analysis of test lisli)
Steide descriotion
StaiutanI 2.00^ &'L
69!i5
KH-nverya Recovery &
6654
1/50
2S
6511
I.W
25
Siandard 2,0(1 fi {ft
<5881
Blanlea
n-d.
1/50
25
Blank b
n.d-
1.'50
IS
( B, b; individual sampts)
A: Peak area Afad); Standard criuticn A(t); Sampit
B: Ratio of portion used for analysis (lish honi&genue) 5/; 0
C: Rario of portion used for analysis (extraeied solution) U : t-'inal wliims VVL)
E ; Amount af blank in tet fish (ng)
F; Amount of test substance recovered (ng)
F-P-<A(t5/A(tld))/B/CKD-E
G ; Reenvery rate (%)
0-F/Qx 1QO
P; CCOTeeirtrauon of lest substance in stanriKd solution Q; Amount ofial mbsance BMfid yoOOag)
ISXi p g.'L
S&e F>s. 7
Test No. 5J51S
4&20 4710
$6,4% 94.3 % Average
95 J %
Nowmbtr 18,2000
Name, ^2E 0/?0
..
Tabte-7
Calculation table for analysis of lest fish (Level 1)
SBMpiedescriptiOn
A
G
K
H
T^aNo. 5151?
I
M
0
Siandard2.00iig'L
6810
TesifMiafto-S daysa
1811
5.00
139
48.9
2.9
^
TesifisliaftwS daysb
l34iS
5.00
104
48.9
2.1
Sttodtrd 2.n[ue/L
(4900
TesifishaftialOdaysi 1499
5.00
11*
48,2
2.*
^
Tea fish after 10 days b 1586
5.02
120
+8,2
1-&
Standani 2,00/iS'L
fi949
Ttaifth after 16 Oaysa 1B62
5.D1
140
47-S
3.0
3y
^
Tesi fish after 16 days b 1909
5.03
US
47.5
3.0
SlaiKid ISOtigH.
6804
-g Test fish aftet 23 tiaysa 3863
5.03
143
47.2
3,0
26
TeU teh after a days b 1253
5.02
96.1
17.2
2.0
Standard; 2.00ja g/L
6756
Tsstfisli after 28 daysa 2597
5.07
139
46.9
4.2
^g
^
Test fislafier 28 days b 1989
5.02
154
46.'Sl
3.3
( a, b ; indwidual sampk )
BCEss;
3.1
A; I'ea.1; area
A(st<3): StanArd solutiofl A(l); Sample
B : Ratio of portion used for analysis
1/50
C . FinaE volume
25inL
D; Dalullon ractoi
1
5; AvErage conCBntratioo of Hanlt. lit analysisof control
P ; Rcc&vtsry tate
?5,3%
Ong/g
G: Weigit of fins sampte (g)
K. CuuMimaduu uriwt uUitiiuc.c in teat Rah ("S/?!
K- {Px CA(t}/A^sid) )/B xS x C/ Q-E j./FxIOO
H: AvEragis concentratiQaoftesI ?65tani!ain teAl water {ft 8/1.)
H (I(n-2)+l(Trl)+I(ii)}/in ; r :NBiiib(a'QT"Mawas,rai]ajyse;s m=2wtiena=2,m=3wheoii S3
1; ConeeoKBtion ofisfiisubsiance in ?9 watcr(^ g/I.)
T- nrp
JtsK/H
M: Avfciagevalue of BCF(a) and BC?(b)
M=(K:F(a5+ BCF(b) }/2
0; Av'ttflge value ofBCF
0 = f M(n.2^ + M(A.I) + M(n) } i i
: Oaiaccnnaiion of lea lubnancc iii standaid solulioa
2.00 a g^
BCFS! = [ [ K(n-2)a + ^n.2)b * K(o-l)a + K(-n-l)h + K(n)a 4 K(n^ } / 6 ] / [ ( l(o-2) + I(n-l) + I(n) } /3 1
See Fig. 8
NDvwibcr 2S,2UOO
^. ^ig C^
Table-8
Calculation table for analysisof test fish (Levd2)
Samptedescription
A
0
K
TfstNo. 51519
II
I
Standisri 2.00 <i g/L
6832
Tea fish aflet 3 daysa
401
last fish afty 3 (Jaysb
.d-
5.00
30.8
4.76
o.5
S.OU
-
t.ro
Standaid 2.0Dfig'3-
6863
Test fish after 10 iaysa,
n.d,
5.02
-
Test fish afiei 10 il)'sb
n.d.
5.01
-
4.71 4.71
SteniJan! ZWftgf'L
6919
Tf-st fish after 16 litysa
47$
Test rish after 16 itys b
585
5.M
S5.S
4.63
7,7
5.05
43.9
4.65
&.4
Slaraiatd 2,00 It ftT-
67-17
Test fish after 23 flaw a
374
5.04
28,8
4.70
6.1
Test fish atter 23 flays b
311
5-M
24,0
4.70
5.1
Standard 2.00 JI&/I.
67'13
TESl(isllniir28 (la^-sa
417
TesHisli after 28 daysb
n-d-
5.07
32.0
4.70
6S
5.07
-
'1-70
.
)__________ f a. b: individual sample
A; Peak area
Afehl}; StonAaEil iolutictiL Aft): Sansplft
R - "telio Of ponion used Ibr analysis
1/50
.____________^_______..____
C'. Fiaal volume D ; Dilution factor
aSfflL
1
13: Averftge lianeestraEiBu <i{Vlaals in aBalysai or tamiidl
Ong,'g
F: Recovery rale
?5.3%
0 : 'Weight tiffins sample(g)
&; CtmccBta'ation at tea sulistMee TO lest fish (ng/g} Te=^Px(A(i3/A(sld))/BxDxC/O.E}/FxlOO
H : Average coacsotralioii ofisst substance in lesi wEier i,ii g/L) H = < Ifn-2.) t I(n-l) * !()} / rn , n :Niirihcrn('teiiiwatMaimlyas ;
1: Conccntailioa aftesi snisluiw in lea waia {11g/L)
ins2wlicafl=2,in=3wheiin fe3
);BCF
I=K/H
P: Cancennalioa of lest substance in standard i&lution Sfi Hg. 3
2.0U/i g'L
Decenrtsr 5, 2000
Kaini
P'1E C B0
Table-9
Osculation table for analysis of test fish (Control)
Sample description
A
Staadiird 2.00^ s/IBefore the initiation of cxpossirt a Bftfort the uariafa'on of exposuit b
6952 n.0. fl.d.
Test No. 51519
i,UU 3.00
SlandBTd 2.00 yg(L ARer the tenmwaion of sxposaiv a
Aricr ihe leiminaiton oftXBOSUIft b
(a, b: indivi(iitiii sample 3
6701
n.d. n.t).
5.08 5.01
A; Peak area
A(att3): Slan(lrf solution A(t); Saapit
B ; R&iio orpomoa vsed for analysis
1/50
C:Rialvoiiinw
2SmL
E: Ainaiiiit of blank in analysis oFcaiiltol fag)
E P sc{A<t)/A(M(!) )/U x C
0 ; W eightof fine sample (g)
I: Coiicenfraiioo of blarik in test iish (ng/g) IsE/G
F ; concenitaifan ancal sutelanw in stiaiilgn) sulmluii
See Fig. 10
z.oop g/L
Hovaaiber 28.1060
^
^e ^t1
Reference 1
Analytical rcsul-ts of dilution water
Sampling- dace
August 8, 2000
Itoa Total hardness (Ca, UK) Suspended solid p9
Total organic carbon
CliBuiii;<tl Oxygen dcBand
Residual ehloriaa Animoaia nitrogen Total cyan Alkalinity 5) anti-it; conduatlvl.ty Organic phussphurua Alkylniercory
! Mercury
\ Cadmium
i la" 1 Lead
Arsenic Iron Copper
?63 It
Manesnesft
Zinc iliiBiniuni Nickel Silver Qrssi'ioiihioi-fna pastlctdes
1,2-Di chloropropans Chlorothaloiii.1 PropyiMtiKIff <;t>lorn 1 (.ritfeii Siliw.inB
TEtiofaBncarb
Organophosphorous pesticides Diaziflon l5i?^aihon Ptnttrothioo BPN
Dichlorvos IprobBtifos rcH
! Unit ,/L
i S/L
-
mg/L ".B/t. eig/L ng/t. Bg/L BK/L [is-fca ttE/L lnB/L Bg/L B^L
lug/I. nie/L DIB/L TOg/;. Bg/L Bg/'L OIG/L B&'L nig/L Bg/L niR/1.
Bg/L Bg/1. iis/L llig/'L nig/I. nBg/L
n-R/1. nis/L Mg/L BIg/L Bg/'L BIg.'L 1%/1.
yeacurBd
in
< 1
Jall>a
SfBTirfiO-ri 'jalllR < 3QO "
<20'8
Tiptsdtno llBit.
1
1
7,3
6.5 -8.3 "}
1.8
<a it
t
< 0.6
-; 6 '2
|
< 0. 01
< 0.02 is 1
< O.tti
< 1 '
0.1.
0.6 0.01 0,01
< 0.01
;
98.1
/
ST.!
n.d.'1
i^
5
0.01
< 0.1
;
< 0.00&5
< O.OQOS !
(Ld.*3
n.d.'2 < 0.0005 "
0.1 0,0005 0.0005
. < 0, OOB :
< 0. 01 *
0.005
:
.
o. m.
<; 0. 00 ''
< 0.005
< 0.1 l;>
o.ona 1 < 0. 05 '
< 0.01
< 1.0 's
< 0.008
< 0.005 *
<; 0. 001 < 0. W < 0,01
< 0. 601. rt < 0,05 " < 1.0 '
< 0.001
< 0.2 'l
< 0.001
< 0. 01 '
; 0. SMt
: e.oMi "i / 1
0.02 0,005 0.002 0.01 0.005 O.ofll 0.0) 0.01 0.01 0.005 a.oool
< 0.006 C 6.004 < 0.0008
< . OOTI
< 0.0003 < 0. 002
a.os^
< 0. 04 "
< 0,008 ^
i
< 0, 0001 " < 0,003 '
< 0.02 <fl
0.006 O.UM 0.0008 o. oooi 0.0003 0.002
< 0.0005 < o.oonn < 0,0003 < 0.0006 < 0.001 < 0.6005 < 0.0003
< 0.005'*
< 0, OOS '
< 0.003 "
< 0.006 ' }
< 0.01 M
|
< 0,008 <
. a.'"
0.0005
0. 0000 0.0003
0.0006 0.001 0.0003
o. oooc
"I Ministerial ta'dinaflse or i.hH Ministry of llcaltit stirl WBlfBre So. 69 (Revisc-d
DeceBbeT 21. 1892)
*?. nEr.1) Guideliftes for Ttstinc of Chaiiiicals, Fish, Ese-ly-Llte Sr.age Toiirity Test
ffitiideline 210. Tulv 17. 19921
O Water quality criteria for fisheries {Sliadanhozin Mhon Suisansigan Hogokyokai, Mftrch 1BB3)
*4 Envixoimenta) Quality Siandard.s t'oz Vtntev Polluianis .Xu.14 (Revised FebruBl-f
22.1999, Envirotttient .tgency) *5 QECD Oujiitfll lllt^ ftif TliSl.llltj uJC ^Lnnill.nl .1, DiltttC?';tfl'ti"" ' Ptcir khf.iuf,1' i''.'h
Test {Gaid&line 305, Juiie ;4, 1896)
\^f ,, 23. PJ-E-OB^
Test No. 515 IS
10'
,"
I
10 h
0
0
0
o
The niillitQUTB oilailaiton f'fWt (iflriOWitanlraiian -factor.
ft
0
2
1
6
S 10 12 14- 18 IS 20 1.2 24 .'6 ?8
Exposure period (days)
Fig, 1 Correlation between exposure period anil biuconceiitiaiiou factor (Level 1).
lo2 r
m1 b
i
L
o o 0
The auriunuui calculation liinii of bioeoiicendaiion factor.
0
--o
31
W 0 t
4 6 & 10 12 M 16 IS i0 22 24 2fi 28
Fig. 2
Exposure pcrind(days) Correlation between exposure periodand bioconcentratioc factor (Level 2).
Four data after 3, 10 ind 1& <)ays were lower lhan dececiion limSt.
December IS. 2000
Nanig RAEC'BO
96-houi LC50=. 100 mg/L
100 "
Test Ho. 51519
IT 50
<s
s
10
100
CMicenlTatioa (ni&'L)
ma
Coaccntation
(me^
Control X.I 3
^6..^. .
12.5
.^25^0
,,-r.z'^0,.-
100
24hnuis
0 0
o0 ,,
0
0 10
Cumulative Mortality (%) .
48 hours
72 hours
0
n
0
0
0
0
0
n
0
0
0
0
30-
40
96 hours
0
0 0 0
.o0 .
50
Pig. 3 Concentration - Mortality Curve. Date : Sspiember 19,2000 jfame ^.. ]U<wwy
Bian (( Cat(VV)01
lOO-i
Test No. 51518
ia-oct-2000
Sift of 1 Channel ES413,001.OODa 1,SOe5
!
1 PeASOsitiot
|
%'
ttA
;
>\ l
-5\h-n--11 i|i,----' -^,,, ,.pr...i,..iiii.,-i-T.,.i..--r^-'i"'-'""'1
Q. 00
t,00
2.00
3.00
4.00
S.OO
8.00
7.0.0""r
iTime 8.00
Stendard solufion 1.25 (ig/L
18-Oct-SOOO
Cal(V\/)02
SIR of t Cliannet ES-
100-
413.00 l-OODa
1.50^
Area'
%-
i
3.7S
3147
A- i
"
;
A^ '5' ,, i, 11.,, 1111.111111. 1111.1.,. 1111.. .--ii.i
0, 00
1.00
2.00
3.00
4.00
Star?dard solution 2.50 ua/lCaW\J)QS
100-i
i.ii--.. i
5.00
i. i.... 1111 i 11 i ... i Tinn&
6.00
7.00
8.00
18-0ct-200fl SIR of 1 Channel ES.
4l3.001.00Da
1.50e5 Area
3.75
%
sieg
^.. . . J 5 l.i .. n ..,ii..
0. 00
1,00
),,
niiiiiiii
2.00
iii.i.;
0.00
i.ii.r 4.00
i:..ii,T-.0.00
Ston eEard solution S.QO pg/L
Cal^sAa4
100-i
1
3,74 12129
i
i.."6.!&.0...;.
,- ii.
7.66
. iTima $.00
1S-Oct-20fiK)
SIR oE 1 Ctl&fflw! ES-
413.00 1.000a
1.50eS
Area
%~
A : h 51' -,,,.;,. . r, i. i i. i ,. i... i. i 1... 1111111. .. 11
0. 00
1.00
2.00
3.00
4.00
i i.i
5.00
11. -rr,-.-.. 11.. 11 .-rT--rrr-n Time
6.00
7.00
8.00
Fl g. 4-1
Mass chromatograjTi of LC- M9 analysis for calibration curve (test water),
Date:?.!."!);). f0,jjl Nam6:_YiM: l^^n-i-he
14000 .-
Test No. 51.519
7ono
&<
/
/ ./
/
./
y /
y 2438 x r = 1.0010
2.5
5
Concesntrarinn (/(g/L)
Conditions ofLC-MS aTlalysis
Fig. 4 - 2
IiistniaiMt Saropic
Solwnt HFLCCt?Tidil;(M
Injeclion vol. Coluian (Sizc.Tcnip.} Eluent
Polwrafc MSCoiwiitioa
lonizaliort mode Cone voltage Sonna; block temp. MoniluTii'S ion DesaaSvai'loe
Tempwattre Gas flow laic
WHer5ZMD,26!10 K-1S19 Waler/Test wfllwfrir riictwery tsst (1/1 VA?)
2ft fiL
Lolunm ODS( 1 icin x 2.1nun I.D.. 4tfC) A(60%);Acetoi>itriJe,2C'10%):5innioyOLi-B-bciylaffliflfc aoiOtr.
O^TOL/fluil
Ekcltospniy (negative) IfiV 120T ffi&413
300^0 2501/hi
Calibration Cyrvc and conditions of I-C-MS analysisfor K-1519 (test wa1ei).
DecrmliM 7.200fl
R j^mc
J S OB^'
_
Starttfard solirtlon 2.50 pg/1WA0101
100
TtsalNu. 315-13
)
is-oct-aooo SIR of 1 Channel E5-
413.00 -t.OODa 1.50e5 Area
%"
/ ^ 0.00
1.00
2.00
3,00
Jest water after 1 day (level 1)
WA01 02
lOO-i
3.75 657
A
. ' " ' i---"1; u .. 11..--n'.i MI. ..ii.ii. Time
4,00
5.00
6.00
7.00
8.00
-iS-Oct-2000
SIR of 1 Channel ES413.00 1.00Da
1.5te5
Area
%-
;
-A-K ' -5'} 1'1"1'1.,->!1 l 1 1 1 1 i 1 1 .1 , . 1 1 , . 1| I 1
0. 00
1.00
2,00
3.00
l esi water alter ^ nay (tevel - 2)
WA01J33
100-1
3.78 6284
?,
A
11" i.i r.i,
4,00
i r^..,'... i. --i i.--r ..,'.. ..i Time
5,00
5.00
7.QO
8.00
Itt-OCI-SiUUU SIR of 1 Channel ES-
413.001.00Da 1^Qe5 Area
3-76
%-
6Q50
A-. JL. -5"
0.(00
1.00
ZOO
3.00
4.00
5.00
6.00
7.00
8,00
Fig. 5-1
Mass chromatogram of LC-MS analysis for test water.
Oate'.^O. 'l .11 Nane.: ^6 09C
Standard solution 2.50 pg/L WA0304
ICKh
.
TertMo. 5WQ
20-Oct-200C SIR of 1 Channel ES-
4lS.001.00Da
1.5De5 Area
%-
!
^
.- .
'
-O5G|O,...(1"-0r0"-.-"2'..0"0 "i-
i" 3.00
Test water after 3 days (level -1}
WA03J36
loo
3.74 6405
fi
./^
i"
4,00
'r"5.00 i
. i {i--111 '. i 1111 i i Tune
6.00
7.00
8.00
20.0C1.2000
Sift of 1 Channel ES-
413.00 I.OODa 1.50S5 Area
3.75
's*
eos3
I,i'l
-5000...
.i.i.i .iiiii
1.00
"2'.p00'
, i.i.i inn
3.00
11 i ^.....;n ..
4.00
S.OO
.
11
6.00
. 11.. i,.. i.,. 1111 Time
7,00
8.00
Twt wartcr aHor 3 days (twol - ->)
WA03J07 100i
aoOGi-aood SIR of 1 Channel E$-i
413.00 I.OODa
1.50e5
Area
%"
f
"0.00 PiS. 5-5:
a./4
6003
?,
i
^
Aii
' "1,b0
'
2,00
'
3.UU
'
4.UU
'
S.UU
,..,&,.O,C,,I --,,,7..,0,0,,,....9...0T0lrru
i
:
Mass chromatogram of LG-MS analysts for test water.
Dftia.-oMp 11 i4- hfamfi: H^s. QQO
Standard solution 2.50 pg/L WA10J3 100-j
Tent Ho, 61510
27-OCt-2WC SIR of 1 Channel ES-
413-001.80DH 1.50a5 Aica
3-77
o%r,'
6243
/ ^ J 0.0'0' "1- '1'.01 0"
|" I"'." "
2.00
1 "
' ' 1 " l '1" '' 1' "
3.00
4.00
Test water sftw 10 days (level 1}
WA10J4
ma-i
'! '--- 1 1 l"l ' i 5.00
j.e.minr't. i ' , i1
-- ',, i',1 ,1 Ii -- I,--| --l - 'l TIifmJI-ffVt "
7.00 -'B.OO
27.0ct.200Q
SIR of 1 Channel ES-
4ia.uui.ouua
1.50e5
Area
^
,,", -
5Bfl?
j
J
," '
t ^/^\,.,.,,,,,, ,, li\\
)
-5- ''' i"1'! ''i i
0. 00
1,00
" 'i-'i " 2.00
r- 'i 'i1 i-' i-r'"-11
3.00
4.00
T&st water after 10 days pevef - 2)
WA10.J5
IDO-i
i
'', 11111. "i""-""ir"i""iT!me
5.00
6.00
7.00
8.00
ay-Oct-aooo SIR or 1 Chennal ES-
013.001.0QDa 1.50e5 Area
'%"
..... -5" O.C30
1.00
^
2.00
3.75
S7S6
A
3-00
4.00
.
S.OO
i--f.-r-r.-t--i i1. i"r. t'r IT n n Timt-i
6.00
7.00
8.0D
Fig. 5-3 Mass chrotnatogram of LG-MS analysis for lest water.
Date . M-CC, l/ . f^ Name: j^fe OJ?0
$tan dard solution 2.50 jjg/L
WA1 6JO
100-
TostM" S1S1S
02-NOV-2000 SIR of 1 Channel ES-
413.001.00Da 1.SQe8 Area
3.80
ft %
6284
5
'
0.00
^^V. ' I--'"-[>..;-- 1 'i T'- 1 1
1.00
K.OO
3.00
!\
I'"' -1 ' ' I1'"'1-
4.00
l-i-----1.11 ,...
5.00
6.00
Test water after 16 days (level 1)
WAie 11
~
lOO-i
'
.
i-r.Ti-r---i " i Tima
7.00
8.00
02-NOV-20W SIB sl 1 Channel ES-
413.00 I.OOOci 1.50e5 Area
%"
3.78
5992
^
.
-5-'
'
0.00
"l-i
. "
1.00
AN
M
1----. .[.i--1--iri r; n i ,
2.00
3.00
ill).------i ''.-' ' '
4,00
5.00
ii.ir"iin>i ir'i'-i.iTime
6,00
7.00
a. 00
Toe't water after 18 <lays (level - 2)
WA16 12
ro-MttV-PDQD SiR of 1 Channel E8-
iCici"
413.aoi.CODa
1,SOe5
Area
%-
-si
ac30
Fig. 5-4
3.79
ii6l4
i
A
j
^
;l
i.,. ' 1.00 ' 2.00 ' 3.00 ' 4.00 ' 5.00 i . iB.O11O' i 1111/..uu n. .a"i."ui Tuime
Mass chromatogram of LC-MS anaiysia for test water.
Data;.5P?. ^ . j4 Mama- P-t PfiO,
Standard solution 2.50 pg/L WA17JS
10th
feslMft. 51518
09.Noiv.2000 SIRoMChannaIES.
4lS,001,OODa 1.50e5 Araa
3,77
%
em
5
Jl ; ^
J_______/'-N______li .5 |' .. i i. i, i. i , .. i,... [n i'ii 11; i . ^"---" 11." 11' i .' r.. j i. . .i. ;,i 11, Tirne
0.00
1.00
2.00
3.00
4.no
5.00
S.OQ
7.00
8.0Q
Te$t water aftw 23 days (level * 1)
OS.Nov.200q
WA17J6
SIRoflChannalES-
lOCh
41S.001.00Da 1-505
Area
3.77 5720
-5|i'i.|i
0.00
. / ^ i. in 11| .'i. |i.
1.00
2.00
111
ri .111
3.00
r.--11111' 4.00
i,.
i
. i. ..,
5.00
~-i 11 .
S.OO
i
Test water aftov aft fifsyR ^eval - 3)
WA17_17
10Ch
1111. i .. 1111 i Time
7.00
8.00
09-N&V-2000I SIR of I Channd ES-
413.001.00Da 1.50es
Area
3.77 5310
^. Jl 5|i i
"i" r"-r Q.OO
|ii 11. i ii
.1.00
!
112..0i.0-..r'
3.0Q
11 i. 11 iiii i.ii 11 i..i i. ii-rr.>-..i-
r ,
11) TilKia
^.uo
&.uu_
o.uu
f.o^
a.oo
Fig. 5-5 Mass chromatogram of LC-MS analysis for test water.
Potc-.-'^C, 1> , I"- fjortini ..^J* ^SO
Stalldacd solution 2,50 pg/L
WAS QOS
100-1
14-MOV-2000
SIR 0(1 Channel S-! 413.001.00Da
1.30e5 Ares
3,74
%-
6144
^
.^/L a
0.00
1,00
2.00
3.00
Test water after 28 days (level -1)
4-00
WfaSS 07
""
.,,s
,,,, .iiii
5.00
i .^,.i,.
6.00
.1..--, . i... i Time
7.00
8,00
14-HOV.200d
SIR of 1 Channel ES^IS.OQ 1 fwnn 1.50s3
Area
%-
I
0,00
1.00
3.74
56SS
!\
^
A
2.00
'
3.00
1
4-OQ
i
... i ini-i
S-00
.. 111.. 6.00
.-i
11
7.0.'0..
. , i nme 6.00
Tost water after 2B days p6val - 2) WA2808
100-1
14-MOV-2000' SIR of 1 Channel ES-
413.001.00Da
1.5Cte5
Area
%
3.75
5670
?l
-5h
0. )0
Pig. 5-6
1.00
2.HO
a.UO
.-p.,.,,. i... ^-i i p i. 11 .. "T .-.ii. i Time
4.00
5.00
6.017
7.00
8.00
Mass d-iromatogram of LC- MS analysis for test water.
PaIciJlieP. it , i^_ Using' Ri^.C'^C-
Blan k CafO 10J 100-
Toot No. S1S1&
29-Aug-200C SIR of 1 Channel ES-
4l3.0D1.00Da 2.00e5
%
Peal positios
n.d.
^
.5" '"i
0,00
1 i
.' 1.00 |' 2.00 3.00 .
i
' i i--------" '1 ' '
Standard solution 1.00 pg/L Cal0102
-IQQ.^
">[ 'r'l'------1 1 '
4.00
5.00
". ,. 11.in i:'-'--: 11 Time
6.00
7.00
S.OO
29-Aug-sooo SIR ol 1 Channel ES-
41^.00 i.oooa 2.0&65 Araa
%"
.
0,00
-1.00
2.00
3,0(1
Standard solution 2.00 pg/L
CalOlO^S
100-1
3.80
3423
A
........
4.00
5,00
.
. , i., ,,i,,, i.. ..i----. -nrne
6.00
7.00
S.OO
ae-Aug sow'
SIR of 1 Channel 8413,001,000a 2.0065
Area
%-
a-Bo
'
6846
-5"
0,1V; )
'
1.bb' '
2.6b
'
Standard solution 4.00 pg/L Cal0t04
100-,
5.00
,
%-
.5' 0,00
1.00
2,00
3.00
L... "
'
4.00
5.00
3,80 129D3
,(
1
"
4.00
5.00
'
6.00
7.00
S.OO
29-Aug-OOQ
SIR of 1 Channel ES4.13,001.0003 S.00e6 Areo
--. ".--'-n-n--n-r-r-i Tifw
6.00
7.00
8,00
Fig. 6.1
Mass chramatogram of LG-MS analysEs for calrbratlon curve (test fish). Date;^^'g. H l'/- Name^ j?-1^ C'^D
,
Test No. 51519
nooo r
^
7000 r a,
/'
-./
/
1
..'
/ /
y=3274x
r " 0.35S
Concentration CMS/I-)
Conditions of LC-MS analysis
Fig. 6 2
Inslrii.msot Sample
Solvcnl
WatBTs2MU,2690 K-151S Water
HPLCOoiriilton fajcuiomiol. Columrt (Sizc.'l'Rinp.) filiient p&lw rate
20/i L L-coluran ODS( 15cin x 2,),miB LD., 40C) A(50%):ARfiton3trilfr3(4r%);5nDsi-iBn-oblu/tLylamine icetete
O.ZmLAaiii
MS Condition
lonlzation mode
Coratollane. Sautcc block temp. Mciiitorms lo
Elwlroisptay (roigativti)
16V
iarc
ni-'z w
Desolvaiion Tcinpeiature C?asi'?ow jalr.
yore
3SOt,tit
Calibrauon carve and conditions of LC-MS analysisfor K-1519 (test fish).
Oewinbf:i7.2QOO
[CM fiJ-t- CBP ^
Stan ctard solution 2.00 pg/L
4F1sr13
100-
Teat No, ^l?16i
2&.AU3.2000 SIR of 1 Channel ES-
413,00 l.OODa 2.00e5
An?
%-
5"
0. 00
1.00
2.00
3.00
Recc?very test from test fish a
4Fish 4
100-
3.82 6005
!\
4.00
r'.'.i-.'i 5.00
.i.ii.,. .111-- iii.,.iiii "nme :
6.00
7.00
8.00
25.Atig-20M SIR of 1 Channel ES-
4l8.001.00Do 2.00e5
AfBfl
%
-5' 0.(00
1,00
2.00
3.00
Recoivsiy test from test fish b
4FiSil 5
100-1
3.81
ARfi^
4,00
-^T--. S.OO
,....-.---r-^--' ". ' , Tinw
6,00
7.00
6.00 J
75.Aua-?OBO SIR of 1 Channel ES-
413.00l.OODa 2.00a5 Area
1
%-
C
0.00 Fig. 7-1
3,81
Ae&n
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8,00
Mass chromatogram of LG-MS analysis for recovery and blank test (analysis of test feh).
Dnte^pea, I/ . i'/- Na^a; i&7fe W
Standard soluSon 2.00 pg/L
Wshfi 10th
'
Ted No. S1S19
25-Aug-2000 SIR of 1 Channel ES-
413.00 l,OODa 2.0085 Ajea
%-
-
-s"i
o.c3-0
1.00
2,00
Biank test of test fish a
4Rsh7
lUU-i
a.ai 6881
?
l\
3.00
4.00
5-00
i;.i-i^-. i i.u ..i 'iTirne
0.00 .
7.00
B.OO
25-Aug-2000
SIR af1 Channel ES4iann-(.(iona 2.00s5
%-
-5' 0. 30
1,00
a.OQ
Blank test of test fish b
4h'iski8
100-j
'Peai; position u.d.
^
3.00
4.00
|
!r i.i-in ,1.--in ..'.< i Tiriie
5.00
6.00
7.00
8.00
25-Aug-2000
SIR of i channel ES-
.
4l3.001.00Da
2.0065
%'
-5"
o 00 Rg, 7-2
1.00
2.00
PeaK pORitlffll II.A.
+
3.00
4-00
S.OO
ii i i-i iii r 1111.1111 i i i11 Time
0.00
7.00
0.00
Mass chromalogram o'f LC-MS analysis for recovery and blank test [analysis of test fish).
Date: >oa? I/. iU. Nanw /?^ C8P
Standard solution 2.00 pg/L
FA03F01 lOO-i
S.75 6S10
Teat No. E-tSfO
20-0ct-2000 SIR of 1 Channel ES-
413,001.000s 2,00e5
&roa
-5'^T-rt- ,1.-11-11 r p 111111 u p i. 11 nipi i|i.
O.OO
1.00
2.00
3.00
4.00
Tes1 fish after 3 days (level 1 - a)
FA03F03
lOCh
i. 111 r,. 1111 5.00
11 11 ri-i-^ifp' i.'(i----j Tinifi
6.00
7.00
8.03
ZO-Oct-2000,
S!Rof1ChannetES413.00 I.OODa
2.QQBS
Area
%
3.75 1S11
5i\ 11 I 1'r. "
0,00
1.00
-A.. |
..i . | .1 ll|i SE-OO
I
lljuiil
3.00
l 1-1 4.00
i'.|
I. i;iir|i
6.00
i i I, I.I
6.00
I,'"ni n
7.00
..----1-T-r.iJppe
9.00
Tesf fifih after 3 day-s Oevel 1 b)
2Q-Oct-?QQQ|
FAQ3F04
SIR ol 1 Channel ES-
10i>1
413.001.000a
2,CiOs5
Area
%'
I .
3.75 . 1348
_________-____________^ -5 [ i-i-n-r-! si-| IT i | i i i i |i i i'."!'I i . i | i i i i i i ill | i .
o.oo
i.oo
2.uo
a,uu
4.00
1,1- i 11 . . ..'i"'i | '.i|ii----,'" ' i
s.w
b".uu
7.oa
' i i | TOT16
s.oo
Fig. 8-1
Mass chromatogram of LC-MS analysis for -test fish (level 1).
Dalc.-iCgQ. <t .! Namer /? gg^_
Standard solution 2.00 yg/L
FAOH01
10<h
TastNo. 51519
27-CW-2000
SIRcrfl Channel ES413.001.000a 2.00e5 Area
?.7f
6900
^
. i _ _ . . . _ _ _ . / J^L - _ -50i.0--0--"TT-1r,.--D0O0----..,22!...0U.,0.U|._,._.!_.._3j..u0,,0u,_.._._-_'.^4...|u0.x0..
i 0.00____1 Test fish after IQ days (level 1 a)
'''J'.""10
..-r5r.|00.,.|.n6,.|0.0i i,..-7..0..0' 6.00
-->,,,
.._-_______y2_7_-.Oo_ct-.200oQ(
SIR of 1 Channel ES-
FA1 OH02
4ia.noi.oODa
^,,,. 2,oAoraeSa
%
3.76 1499
- ^ _ -- - -- -Sl I I ' ' I ' ' ' I I -"T"----[III r|"" I 1^ I I | .
0.00 .A-W_
2.PO
3.00
t. Test fish alter 10 days (level b)
FA1 OHOS
,..-,.... 'l l |. I l I----^.--l-. . . |
4.00___5.00
,
,
6.00
.QQ_
, ,-----------T""" ! '
7.(M
6.00
^27"i -0^c<htt.-2rt 000
SIR of 1 Channel ES413-001 ,OODa 2.00e5
Area
- %1
a.7Ei
1566
-S'. i i
o- .o-*p
i
i
l1
''"' | i i i----''
t .'o'o_
' i -' ' '
2.0nf0t
i
'
| i--r-->n>|i"'~TT-- '
'
nn.notol
4*..O0P0 _
'
i'
' i ' I---ii';--i-i----'' . '-'i] . i.i. . , 'n,, me
S-00
6.00____7.00____8.00
Rg. 8-2 Maes chromatogram of LC-MS analysis for test iish (level 1).
Date-JftiG. }f {V Name- R^-E^0
Slandard solution 2.00 |ig/L
FA16 H01
100-1
Teat No. CIS l0
02-Nov-2000 SIR of 1 CtiannetES-
413.001.0DOa SLOOeS
flriifl
%'
-5 i 0. 00
t.OO
2.00
3.00
Test fish after 16 days (levet 1 - a)
PA1SH02
100-
3,aa 6949
?
A
4.00
I lll.i ll r. .1--------T-rrT-i----r--^r^-iTime
5.00
5,00
7.00
8.00
02-Nov-2000
SIR oT1 Channel ES41S.001.0QDa 2.0085 Area
%
'
-5't
0. 10
1.00
s.7a 1962
A
.
ip I- ^^|WIT''"'IIT----. 1 ' "
2,00
S.OO
4.00
Test fish after 16 days (level 1 b)
FA16 H03
1&0-,
\.:--ii,i.ii.i.i-r,.....ii....--i--i-nme
5.00
6.00
7.00
8.00
a?-Ntiv-?oQa SIR af 1 Channd ES-
4ia.001.00Da 2.aoe5 Area
%~
-5"
U.LJU
3.79 1909
.&
"I'--I1 1 " 1' "---" i " " '. r t i " i-' i "'."--i Time
1.UO
'.W
3.UU
4.UU
&.UU
B.OO
7.00
B.OU
DBte..,-MiC. ff ,!V r-toiitg. /JIfi.Pp'P
Standard solution 2.00 ijg/L.
FA23H01
10C-7
09^Mov-20tiO
SIR of 1 Channel ES413.0D1.00Da 2.00e5 Area
1 %' 638.70H4 ,
-51, ....),--^-n--r" "
"
"
0.00
1.00
2,00
i
"
'3' .
0
-
0
"
i--"i --
4.00
Test fish after 23 days (levsl 1 - a}
FA23H02
'100-
"11 iii-:"
5.00
-'--11
5.00
i i.' i ^ r,. nri TilTia
7.00
S.OO
ag-Mov-2000
SIRoflChanneiES4lS.001.00Da 2.00e5
Area
%"
,,3.7,^8 issa
A
-5 . 0.00
.r-p
n
i. .i 1.00
i
|i
'. i ii .00
....
i
i|.i
3.00
.,11 .,."i-i| 4.00
Tas-t fish after 23 (jays (level 1 - bV
PAS3H03
300-,
i i"i.
5.00
--..i," .1-1 i.'ri i Time
6.00
7.00
6.00
09.HCW-2DOO
S R of 1 Channel ES-
4-lS.001.00Ua 2.00e5 Area
%-
"
3.77 1253
. . . . . . . . 1--------------"--------. . 1
-5 1.00 U.Ul.)
"--r- "-.... r.----n--n--" | 1 1 . 1 1 1 . 1 . 1 I
1. 1
3.00 4.00 2^00
, 1 . T-l | | | 1 1----11 . 1 |
5.06
f , e-oo
r.oo
i iTirrtS o.oo
Fig. 6-4 Mass chromatogram oi LC-MS analsis for test fish (level 1),
Dftte i CCO. Jf , fU Na.rto; fofg^fif
Standard solution 2.00 pg/L
FA2SL05
10&-
Test No. 55513
14-NOV-2000 SIR of 1 Channel ES-
4l3.001.00Da 2,00es Area
; %-
-5
0,00
1.00
2.00
3.00
Test fish after 28 days (level 1 - a)
FA28L06
100-,
3.75 6756
A
1 n
',i
4.00
.i,nnii 11. ----i i .ii iip---i Time s"
5.00
6,00
7-00
8.00
:
14-NOV-2000
SIR ot 1 Channel ES-
AlS.001.QQDa 2,OCte5
Area
%
3.74
'
2967
A
-5" 111"'11''" i' '' i '' i 11
0.00
1,00
2.0Q
- '3'.0r(2' '
."
" i '-i--i--i.
4.00
5-00
Toe! fish after ae days <lwel 1 - h]
FA28L07
100-^
. 1111 ii 6,00
.11.'. i 111 r-' 'i Tims
7.00
S.OO
14-NOV-2000 SJH of 1 CFiannel E8-
413.001.OODa 2.00e5 Area
%
-5' O.t30
Rg. 8-5
9.74
1989 A
'1,00
'
2,00
'
3.00
'
1-1. .i...1.1.111........i. ..r"-i.iTirne
4.00
S.OO
B.UU
/.OO
8.0&
Mass cilrornatograrn of LC-MS analysis for test fish (level 1).
Date; IBM. II . !V Mamp- gJ-f OffO ..
[SStatanndadrad rsdolution 2.00 ^ig/L
IFFAA6S3SPP05OS
i 100-
Test No. 51519
2zo0"-o0ct-a2o00o0o| SIR erf 1 Channel 5-
41S.001.0000DD&a 2.00Se5 Araea '
%
3,76 fiaa?
,
i
!\ -5
"
1"' 1- "' .11 "I > .
'.----.--rrn-rp-r--t-i----n----n-,
-,-.,.-,----,-rr----j----7,.,0|.0
.,|...
..TilTle 8.00
5,00
6.00
0.00
1,00
2.00
3.00
q.OO
2Q-Oct-2000
Test fish after 3 (jays (Isvel - a)
s R o? i Channel RS-
PA03F06 100-i
4l3.001.00Da 2.00e5
Area
%'
| i
3.74
401
5l.....""1..00.
..., ;. ,4
2.00
.
i.,.,.,,
3.00
,,.
4.00
.,
0.00
Jest fish after 3 tiays (level 2 - b)
FA03PD7
IftO-i
,,i ,,...i
5.00
111 11"----"-T-"-n Time
6.00
7.00
8.00
ao-oct-aooo
SIR <?i i i-aiannei ta-
413.0Q1.0&DA
a.oo5
'. %'
-5 O.OQ
Fig. 9-1
Teak ^aiticn
TS.(I.
^
1.00
.00
r"
-r1 a.oo
4.00
5.00
enn
7.00
Mass chromatogram of LC-MS analyas for test feh (level 2).
n Time s.oo
DsaB^t-SQ^!! . if- Name : P^E- C'fs^
iStandard solution 2.00 wfi
;FA10L01
\ 1100<0h-
%-
3.78
S863
5 [..
0. 00
,.,,,,1...0,0
n.pi
;.i
2.00
i|
3.00
A..
4.00
5.00
Test fish after 10 days (levd 2. a)
FA10L02
100
Toot Ho. 61610
27-0ct-200ol
SIR o(1 Channel ES-'
4-13.001.ooDa 2.00e5
Armi
6.00
i--r-^iTIme
7.00
8.00
27-0ct-200<?
SIR ol 1 Channel ESH3.001.00DB 2.00e5
%~
Pealk BosiUtB
n.a.
^
-5'
"-T"--r
0. 0&
1.00
2,00
3.00
4.00
5.00
Test fish after 10 days (level 2-M
FA10LQ3
lOO-i
6.00
7.00 ' riiBOi OTime
27.0ct.2000
SIR i7f1 Channel ES-
4ia.001.00Da 2.00^
%"
-5'
0.1.3U
Fig. 9-2
Peak posillon
a.d.
-I/
1 .UU
Z.OU
3.00
--.1. ''"~i1' i
4.UU
n
i>.UU
b.UU
/.UU
Mass chrCBTiatogfam of LC-MS analysis for test fish (level 2).
a^u
Pate ^tVf. 'f , !V ^am<t! <t?7gC/?f?
Standard solution E.DO HS/L
FA16L01
100n
Tp-stNn R1519
02>Nov*2000
SIR oft Channel ES4ia.ooi.oODa
2.00e5 Area'
3.rs
6919 (i I'l
----il.,
-.Time
5
0,00
1.00
2.00
3.00
Test fish after 16 days (levet 2 - a)
FA16L02
4.00
5.00
6,00
7.00
a.oo
Q2-NOV-2000!
SIR oi 1 Channel ES4<3/x>l.DODa
TKh
2.0Q5
Aree
%
3.80 47B
Sr Q.Op
1.00
2.00
[Tpatti*h after 16 dsys (level 2 - b)
FA16L03
1W-1
S.OO
7.00
i , -n Ttfiie 8.00
02-NOV-2QOQ|
SIR of 1 Channel ES-
4l3.001.00Da 2.00e5 Area
U.UU
Rg. 9-3
S.7B 585
'
1,1.10
Z.OO
3.qo ,, 4.00
5.00
G.OO
7.00
Mass chromatogram of LC-MS analysis for test fish (ievel 2),
-iTime
0.00
naca ^r..':f?. n . in. JiarTt^. ^j fi30
Standard solution 2.00 jja/L
PA23L01 100-1
ToetMa. &W3
Q9-Nw2(MKlj
SIR of 1 Channel ES-1
4-13.001.00Da
2,OCte5
Aroa
%-
-
.
.
^'.n'. lp,'-'ii.i tif.,r.TT. i. ^
0,00
1,00
2.00
Test fish after 23 days (levat
FA23L02
100-'
3,78
8747
/I
A
.
,,
u,.
3.00
..pl-r.-----------11 ii..---rv..;'
4.00
5,00
6,00
z - a)
i.. ^ i.
i-r'i. , . \ Tiroe.
7.00
8.00
OS-Nov'2000
SIRoflChannefES41S.OOt.OODa 2.00e5 Area
%-
3.7& 374
-5 -r-r-. l|----1 I | lir. , , |, ||, 1 .1 l |..l I r|.i.
0.00
1.00
2.00
3.00
| ) , , 1 .|1 1 1 4,00
j
5,00
Twi fifih aftor 53 days (tevol a - b)
PA23L03
100^
.1
-- ---------------- ----^ TliTia
e.oo
7,00
a-oo
(w-Kou-aooa
SIR oi 1 Channel ES-
AlS-OO-i-OODa
S.00s5
Area
3 0.00" ' 'i.bb" '
"
2.00
'aoo
3.77 w
1 45o"
' "'"' i '--ii5.00
'r 1111 i 111 ' i' i Time
5.00
7.00
8,00
Fig. 9'4 Mass chromatogram of LC-MS analysis for test fish (level 2}.
Dgts'-MOg.// . ft/ Name; k^6GPD
Standard solution 2.00 pg/L
FA28LOa
IdO-i
TpRtNo- SWQ
14>Nov-2QOO|
SIR of 1 Channel ES413.001.00Da 2.00e5 Area
"
-ST""|i--'i' "i--".. ni|i.. r-"
0.00
1.00
2.00
3.00
_
JTesftish after SB days (level 2 a)
PApa ns
-(OT "j
1
3.75 8/13
i
A
i.ii.. 11.'i.
4.00
'5.0,"0 "
i.i......i---'i-
6.00
7,00
"f:
i Time 8.00
14*Nov-2000|
SIR of 1 Channel EE'H3.001.00Da 2.QOe&
Area
%]
3.72 417
-50", oo 1 1,00 1 e-bo ' 3,00 1 4.00 ' s.oo ii'i6.0,i0i^-iii7'i.,0'0i' i'i''8iT.0i0ffiS
Tsst fish after 28 days (level z - &)
H*Mou-2000
FA2SL10
SIH of1 Channel ES-
100
413.001.00Da
2.00e5
%
-s' O.t30
Fig. 9-5
P?afc positifln
T
1.00
' T'2.00 '
'
3.00
4.00
'
-i- --i----i----1~----i^----\ Tini-o
5.00
B.OO
7.00
fi.OO
Mass chroinatogram of LC-MS analysis for test lish (level 2).
Oata-aMi" If tU. Mamft- R^t, 0^^
'.Standard solution 2,00 pg/L IFAQOCOI '1! 1(0M0>-il
t
Test No. 51518
16-0ct'200o!
SIR <rf 1 Channel ES- 413.00 LOODa2.00ea5 Area
' 1
$.Sl
%-
6SS2
.
J
-S S-r-rr-"' t'l
t?,00
1.00
A
il
n
"l'-"l ,,,,
' ;i 1 'i i " -!
----------"----'--I --.--i--i --|.--.-i i - K ii - "
2.00
3.00
4,00
5.00
Before the initiation of exposure (control a)
FAOOC02
1UU-
,ii
.
ii.r,i..r.
11.1 Time 3.00
6.DO
7.00
16-0ct-2000
SIR of 1 Channd ES<H3(ioi onrta
2.00e5
P?alt iifiaitioa
1 %- n.a. I
0.00
1.00
.00
3.00
4.00
Before file initiation of exooswe {control - b)
FAOOCOS
100-i
5.00
6.00
7.00
8.00
ia-0ct:-2ooo
SIR of 1 Channel ES-
4i3.Q01.oODa
2.0085
Pean p&Biuon
,
n.d.
J.
.
-5t---n< v.w Pig, 10-1
1.UU
2,00
3,00
4.00
S.OO
ii .. i ..i. i...-- i' i Tiine
a. oo
T.oa
o.oo
Mass chroiTiatogram of LC-M5 analysis for lest ish (control).
Gate-JSg fo, ^ Namp.- VjV L.^g
Standard solution 2.00 ygIL
FAP8L15
i,w, '.
TeetMo. S1S10
14-Nov.200a SIR of 1 Channel ES-
413.001.OODa 2.00e5 Area
%
3.76
5701
l
A
-5
i
0,00
After the
FA2BL.16
i . >i.i--r-"^ ip.i ii.ii--i.. i. -.
1,00
2.00
3.00
4.00
termination of 6xpo,su!-e (conirol
.:-inp'--i-"" . "
5.00
6.00
a)
11 .11,' 111 --"-; Hi) it-
7.00
8.00
14-Mov-ZQOO.
SIR trf 'i Channal ES413.001.00Da
2.0C>65
; w''
Fe6'il ?C5ll!oil
n.d.
^
^*
1
1
'
--H-i--i i --r^--"i TiU'uTnt^6
0.00
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8-00
After the termination of exposure (cantrol b)
14-Nov-2060
FAaSL-17
SIR of 1 Gtiannel ES-
^p
413.001.0003 2.00e5
%-
.
0,00 Fig. 10-2
Peik position
n-d.
^
1.00
2-00
3.00
4,00
5.00
^-. '., 11
6.00
7.00
Mass chroimlograin of LC-MS analysis for test fish (control)
. Tinw 8.00
Date -.s co,!. i/ . /</. MaiYiQ: ffJF !>SQ_
^ 10
i
s
1
i
?
TestNo.51519
S
0
10
20
30
JExposureperiod(days)
Fig. 11
^> 10 r
.5-
I
Concentration of dissolved oxygen (Level 1).
^ 5
0
u
10
20
30
Exposure period (days)
Fig. 12 Concentration of dissolved oxygen (Level 2).
1
a
5 -
|
0
10
20
30
Kxposura period (days)
Fig. 13 Concentration of dissolved oxygen (Control).
Nawnber 22,2000
N'ame
fi^ ^4?l'
Test No. 51519
iia.o -
'
ST
^....^ ^ 85.0 -
'^--.--r.-vf^
c - 0 ^--ff
^
60.9 --
Str. . .--...---.-..
..-.-.----
.-..^y--.
i
^i.ft
!
i' .!
S'i --
ri t
i'
.
j
'li
40.0 ,J------------
"~'"\
-I
i
: CF3-(C F2)6-COOH
t. a
roai.o
3W.<t
2<K.(?
------!-
0- ^ 0 -01
ff
0S-tHr. Stri &
Def.
(sa'.o
low. a
aiaD
If'aa
IR conditions
InsTrument . shiiiiadzu FTIR-SZOOPG
Sample
: K-1519
- Method
: KBr Tablet
Fig. 14 - 1
IR sptsuttUtn cif tsst substance incaaurod before the experimental start.
Date : Aug. 31.2000 Maine : g^O^O
TOO.O
"i ,
*
5T
j
:
St.O
i
. Jfrr'^1*"^'
V
^
4
^
/'. i
A
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Test No. 51S19
^^ /'N/'.^
1 'L-
L
'1 J
T -----ir-f 1
40.0
.
1
i 1i"""""'""""
i:
i!
1
1111, o.o B.O
<
3M<).&
1
1
1 1
1
,
1 1
20(Kl.t
ISOO.a
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1
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SBO.O
1R conditions Instrument : Shiroadzu FTIR-8200PG
SampIs
K-1519
Method
KBr Tabiet
He. 14- 2
IK spwtruni of test substance measured after
the expertmentaI completion.
Date : New, 20,2000
MsRie : pi^, O'fSO
TestNo._^I/L
tBetrumanI
S^pte
MR Wflterfi7Mn HPLIC Waters aS90
fc- f ^ ?__ fD.^lL
HPLC Conditions
Inlet system I'fajumn)Flow iniectipn
SainplB sizeJ^pL (Soiwnt Acf-f^.T^lfj/i-.t.T'-c-i^ ^--------------------------.----J
Column.
/^^/b,...^
PFFK1
^..S________________________fsn5
Column size _jL.cnrtx _f^n!im 1.D., Column terop, ^L-'C
Eluent A(P^)Actt,^n-:L , R W^)^^( ^VvU^e &nWf r' / - VM
Row [ate <?.^ .mi/in In
Post column eluent _
Posi column flow rate "- rnUmtn
Spilt raflo, "..
MSConrfttions
Innii'aSon mode <S\) APcl . Detection inadg PosHive. Naq^^ -
Source OapiHaryaL3.!sV, GonB_/oJ/, B!tractor^__V, FIF Lens ,'S3. V Ion source temp. ^2_C, Desolvation gas ^3^Unr, DesolVBlionternp,ii&tLC
Corona __ltV. APcl proba temp. _45Q, J- 'C MS Ion energy ^2__V,Mu!tiplisr^^__V
MonltCTing ion m/?
_
-______
Nolg
Kurums Lahoraiory, Chemicals tealuation and Research Insfflute, Japan
Rg. 1 S- IMS spectrum of test substance. Dais ^ccci i? /-^ Operalor Y'^-'^f^-t
K-151910.0 ma/l. (SoL-AON/watw 1/1 V/V) l6pl"
Scan02
n
T-,3.67
ao-jun-aooo!
Scan ES-,' TIC
4.25e5
1.70
179 (
25) 25I 23,,-a^ I
^f^W
I
T W w ^ Y\3.8S 4.52
5 64 S 81
-T-i
0.00
1.00-
2,00
3.00
4.00
K-151510.0 rng/L (So(.:ACNAwater 1/1 V/\Q 10pL
CcnnQE1SO<0.5S2)Cm(1!32;10S<iei:1S7+a08:ai6)5 413( B - H ) "
5.QQ
iTime
fl.OO
20"Jun-2QOO
Soon CS9.6664
-COOH)-
369
M : CF3--(CF2)6--COOH
M-W.; 414
837f3.M - B }
Isaa
O-'TI-l | I
rT-r--
mfe
100
200
300
4DO
500
600
700
800
900
_
?-/.'"?? ^SW-t-^ f^:faf^
Fij^. 15 '- 2
MS soech-iiin of tet stifcetanea.
K-1519 10.0 mg/L (Sol.:ACN/water 1/1 V/V) SOpL
20.Jun.2000,
^MW/' ^^f^^W ^^/\WA^T^^ ScanC2
IMh
0.04
["fo^S
%-
.A,^'^1'70 ' T' (
a-5l^9 67
!fl 2.5t 2-923-39 l^85
4-S2 4ga
scan ^ i "no <.2ie5
5.64s 81
02 0 -T-. r-i | I i . i----r-r- . | . l i '-ri-T--i----------i ' ' i ----------------------------------------------------------
Scan
Scan ES-
100-1
.65
3691,OODa
j
5.07 ed;
I
^ f %-
t
I ;
.......... ...., 0- ------i-------.-.--------i1'1"1 i "
i
i - ----'
SconOS .
i
sca" ES.
'(3.61
4131.0003
100-
J^TB
1.14e5
%,,
\
V. 0-
.
1 i "!"' '------n----------------. '
1 1 ---------. 1 I-.-.- r-j-T----.------
,
,
- ------r-,-r-,
| .
.,...,.... Scan02
.
. ,
ScanES-
4Qkl~
3.67
B271.00Da
2,4Se4
3.^
1,
%-
I..E
n q >. 'i
000
1-00
2.00
3.00
4.00
5,00
8.00
Pig. 15- 3
MS spectrum of test snbstancft, '
--0 rmt'"
-OOfIt
-BBS'O
' "
--6O1Sf-KIt
-IM'lt
-s
-tlI-B
BO-JQl
-Wsat
-ro'toi
sis'im
-OH." 101
-5'
- taa- im -B8'IOI
-wm:
-
"EC1'"ID-
-lafin":
-tcrsK
-!"
-9WIK
-(E"ItC
. --iISBKl''nlIit:
.
-g"(it
-6CO'2It
-8 r<
--ilcifcnntt
-IM-IIt
1 '"EU'nt
' -iM'nt
1 ^-BirBt
-ZtE'EIt
-5" -Bafdc
-SK'Ill
-cswraittt
-BM'lIl
-IWSTt
-g -SS-ltt
-tBfan
-tBS'lIC
-BWflI
-
-esc-fflt
-MI'IBT
n
rt
--MWSB'iII
" -
--iWtrC'ali
-
-zn-ffi
- i --Tfio'tn
..S.I '-"^i"
