Enantioselective Analysisof Chlordanes in Serum

Chisato MATSUMURA, Masahiro TSURUKAWA, Hiroaki KITAMOTO, To

shihiro OKUNO, Takeshi NAKANO(Hyogo prefectural institute of public health and e

nvironmental sciences, Kobe Japan)

Technical chlordane

from 1945 in U.S.

a ubiquitous and persistent

killing lawn and garden pests

termite control

pest control on crops

In Japan

Termite

• From 1950

• In 1968, it was banned for agriculture

• Used for termite control

• The Chemical Substances Control Law restricted it in 1986.

0

500

1000

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250019

5819

5919

6019

6119

6219

6319

6419

6519

6619

6719

6819

6919

7019

7119

72

1979

1980

1981

1982

1983

1984

1985

1986

technical chlordanes (including heptachlor)

Technical chlordane import (tons)

Basic component of technical chlordane this work USEPA

Cl6-1 α-chlordene 0.7 3Cl6-2 γ-chlordene 3 9Cl6-3 β-chlordene 3.6 4Cl7-1 heptachlor 4.8 7Cl8-1 compond-4 0.3Cl8-2 trans-chlordane 18 24Cl8-3 compond-5 5.8Cl8-4 cis-chlordane 16 19Cl9-1 compound-6 1.5Cl9-2 trans-nonachlor 14 7Cl9-3 cis-nonachlor 4.5 3

Our study

Cl

ClCl

Cl

Cl

ClCl

Cl

Cl

Cl

ClCl

Cl

Cl

Cl

ClCl

Cl

Cl

Cl

ClCl

ClCl

Cl

Cl

Cl

Cl

Cl Cl

ClCl

Cl

Cl

trans-nonachlor cis-nonachlorachiral

trans-chlordane chiral oxychlordane

Cl

ClCl

Cl

Cl

Cl

Cl

Cl

O

H

Cl

Cl Cl

Cl

Cl

Cl

Cl

Cl

O

H

major component of technical chlordane

| Efirst – Elast | EL - ES

EE = 100 X ------------------- or 100 X ------------ Efirst + Elast EL + ES

E : peak area or height Efirst Elast

Efirst : first eluting enantiomer

Elast : last eluting enantiomer

EL : larger enantiomer

ES : smaller enantiomer ES EL

Retention time

Enantiomeric excess (EE)

What’s known from EE

production

housedegradation

agriculture

diffusion

Change EE

accumulation

e.g. Physical process Biological process

use

For termite control

transport

← CS solution (2.5ng each) 2.5ml of diethylether 5ml of ethanol

10ml of n-hexane X2

Analytical Flow Blood

Shake （ 10minuten ） and centrifuge (3000rpm, 10minuten)

n-hexane layer

Concentrated to 1ml under the nitrogen streem

Clean-up with LC-Florisil (1g) / LC-Si (1g) 　 (Supelco)

Eluted with 10ml of 15% diethylether/hexane

Concentrated to 0.1ml under the nitrogen streem

Centrifuged (3000rpm, 10minuten)

Analyzed with HRGC/MS

2g of upper layer (Serum)

← ISS solution (0.5ng)

• CS solution13C10 chlordene, 13C10 heptachlor,

13C10 heptachlor-exo-epoxide,

13C10 trans-chlordane, 13C10 oxychlordane,

13C10 trans- and cis-nonachlor

• ISS solution 13C12 2,3’,4’,5-TeCB (PCB#70)

13C12 2,2’,4,5,5’-PeCB (PCB#101)

Internal standard solution

HR ＧＣ／ HR ＭＳ and Column

GC 　 Hewlettpackard 　 HP-5890IIMS 　 JEOL MS-700

Column ： BGB-172 (BGB Analytik)30mX0.25mmID,0.25μmFilmThickness（ 20%-tert-butyldimethylsilylated-beta-cyclodextrin in 15%-phenyl-85%-methyl-polysiloxane ）

HR ＧＣ／ HR ＭＳ ｃｏｎｄｉｔｉｏｎｓCarrier gas:Helium (1.5ml/min constant flow)

Oven temp.:120 (0min)-4 /min-180 (0min)℃ ℃ ℃-1 /min-230 (15min),℃ ℃

or 120 (0min)-4 /min-200 (0min)℃ ℃ ℃-1 /min-230 (25min),℃ ℃

Ion source ： 250℃ 　　 Inj.Temp. ： 230℃Ionization Energy: 38eV 　　 Ionization Current: 500μA Transfer line Temp. ： 245℃ Injected amount ： 1μl

GC/MS compound list (2 injections)

m/z: 218.9 - 337.9 (1.54M)

HCH DDT&DDDHeptachlor

Chlordene Te4CB Oxychlordene Pe5CB

13C6-HCH 13Cring-DDT&DDD

13C10-heptachlor 13C10-chlordene

13C12-Te4CB 13C12-Pe5CB

m/z: 352.8 - 441.8 (1.25M)

HeptachlorepoxideChlordane

OxychlordaneNonachlor

Hx6CB Hp7CB Oc8CB

13C10-heptachlorepoxide13C10-chlordane

13C10-oxychlordane13C10-nonachlor

13C12-Hx6CB 13C12-Hp7CB13C12-Oc8CB

GC/MS-SIM chromatogram

on BGB-172 HRGC column

Technical Chlordane

Atmosphere

chlordane transtrans-- 　 　 transtrans--

ciscis- - ciscis--

MC-5MC-5 　　 　　 MC-5MC-5 　　　　　MC-4 MC-4

River Water

River Sediment

chlordane transtrans-- 　 　 transtrans-- ciscis- - ciscis--

MC-5MC-5 　　 　　 MC-5MC-5

　　　　　MC-4 MC-4

Technical Chlordane

Serum

chlordane transtrans-- 　 　 transtrans--

ciscis- - ciscis--

MC-5MC-5 　　 　　 MC-5MC-5 　　　　　 MC-4 MC-4

　　　　 beta- gamma-

　 alpha-

chl6-2

　　　 chlordene

Technical Chlordane

Atmosphere

chlordene

　　　　 beta- gamma- 　 alpha-

chl6-2

　　 　 chlordene

River Water

River Sediment

chlordene

　　　　 beta- gamma-

　 alpha-

chl6-2

　　　 chlordene

Technical Chlordane

Serum

chlordene

heptachlor-epoxide and oxychlordane

heptachlor-epoxide

oxychlordane

exo- 　 exo-

endo- endo-

Enantiomeric excess (EE)　　 chl6-2*3 41

　　 beta-chlordene 　　 3.8

　　 gamma-chlordene 21

　　 heptachlor-exo-epoxide 50

　　MC-5 71

　　 trans-chlordane 24

　　 cis-chlordane 48

　　 oxy-chlordane 21

•3 unknown isomer on chlordane

Cl

ClCl

Cl

Cl

ClCl

Cl

Cl

Cl

ClCl

Cl

Cl

Cl

ClCl

Cl

Cl

Cl

ClCl

ClCl

Cl

ClCl

Cl

Cl Cl

ClCl

Cl

Cl

Cl

ClCl

ClCl

Cl

ClCl

Cl Cl

ClCl

Cl

Cl

Cl

Cl

ClCl

ClCl

Cl

ClCl

Cl

Cl Cl

ClCl

Cl

Cl

Cl

ClCl

ClCl

Cl

Cl

Cl

Cl

Cl Cl

ClCl

ClCl

Cl

Cl

ClCl

ClCl

Cl

Cl

Cl

Cl

Cl Cl

ClCl

Cl

Cl

Cl

Cl

ClCl

ClCl

Cl

Cl

Cl

Cl

Cl Cl

ClCl

Cl

Cl

Cl

ClCl

ClCl

Cl

Cl

Cl

Cl Cl

ClCl

Cl

Cl

Cl

trans-nonachlo 　　 cis-nonachlor

achiral component-MC7

cis-chlordane

component-MC5

trans-chlordanecomponent-MC4

heptachlor

component-U82

Cl

ClCl

ClCl

Cl

Cl

Cl

Cl

Cl

Cl Cl

ClCl

Cl

Cl

Cl

Cl

component-MC6

H

Cl

H

H

Cl

H

Cl

Cl

Cl

H

Cl

HH

Cl

H

H

Cl

H

Cl

Cl

Cl

H

Cl

H

Cl

Cl

H

Cl

C*

Cl

H

C

Cl

H

Cl

C*

Cl

H

Cl

Cl

Cl

Cl

C*

Cl

H

C

Cl

Cl

Cl

C*

Cl

H

Cl

ClCl

Cl

Cl

Cl

Cl

Cl

O

H

Cl

Cl Cl

Cl

Cl

Cl

Cl

Cl

O

H

Cl

ClCl

ClCl

Cl

Cl

O

H

H

Cl

Cl Cl

ClCl

Cl

Cl

O

H

H

Cl

ClCl

Cl

Cl

Cl

O

H

H

Cl

Cl Cl

Cl

Cl

Cl

O

H

H

o,p’-DDT

o,p’-DDD

alpha-HCH

oxychlordane

heptachlor-exo-epoxide

oxychlordene

GC/MS channel list 1 (2 injections)

G#: 1 Repeat: 1.01[sec] m/ z: 218.9 - 337.9 (1.54M)Ch.# Compound name m/ z

1 HCH 218.91162 HCH 220.90873 13C6-HCH 224.93174 Mass Lock 230.98565 Lock Check 230.98566 DDT 235.00817 DDT 237.00538 D8ring-DDD 243.05759 13Cring-DDT 247.048410 heptachlor 271.810211 heptachlor 273.807212 13C-Heptachlor 276.826913 PCB4 289.922414 PCB4 291.919515 Chlordene 302.888316 13C-PCB4 303.959717 Chlordene 304.885318 13C-Chlordene 312.921819 Oxy-chlordene 318.883220 Oxy-chlordene 320.880321 PCB5 325.880522 PCB5 327.877523 13C-PCB5 337.9207

GC/MS channel list 2 (2 injections)

G#: 1 Repeat: 1.01[sec] m/ z: 352.8 - 441.8 (1.25M)Ch.# Compound name m/ z

1 heptachlor-O 352.84422 heptachlor-O 354.84133 PCB6 359.84154 PCB6 361.83865 13C-Heptachlor- 362.87786 13C-PCB6 371.88187 CHL 372.8268 CHL 374.82319 Mass Lock 380.97610 Lock Check 380.97611 13C-CHL 384.856612 Oxy-CHL 386.805213 Oxy-CHL 388.802314 PCB7 393.802515 PCB7 395.799616 13C-Oxy-CHL 398.835817 13C-PCB7 405.842818 nonachlor 406.78719 nonachlor 408.784120 13C-Nonachlor 418.817621 PCB8 427.763622 PCB8 429.760623 13C-PCB8 441.7999

Technical Chlordane

Atmosphere

River Water

River Sediment

Serum

chlordane transtrans-- 　 　 transtrans-- 　 ciscis- - ciscis--

　 MC-5MC-5 　 　 　　　 　 　 　　　 MC-5MC-5 　　　　　　　　　　　　　 MC-4 MC-4

　　　　 beta- 　　　 gamma-

　 alpha- 　 chl6-2

　　　 chlordene

Technical Chlordane

Atmosphere

River Water

River Sediment

Serum

chlordene