Page 139 - 006

133

8.4.2 # $"$ U V 8 (Biochemical detection)

*! # (" % 9
" ' % & ( 5 7
!
R
1) # $M $IO # $ #" 8 [x acetylcholinesterase &

7

.4 ! Q organophosphate carbamate ! ' !
0
(*

5
7

1
"#"
' 5 acetylcholinesterase : AChE
#
hydrolyze !
7
,

acetylcholine ! ! ! ' (neurotransmitter) ("
"#" 9
" hydrolysis

+/

6/
7

* ' acetylcholine 5 ! H
choline
(
(acetic acid) 5(
1
6/

7
+

( *
(
! ! acetylcholine # % , ! '

5

0
67
/
1
! '
' ( '
. " '!(

1

67
,
/

" 5
Y ( #
'1 5 AChE ' (
5 ! H#
% ( ! Q
7

Y

/

organophosphate carbamate ''
(.45( #
5( ." ! %

1

7

/
" 6 "*
! . 4' /! H" #" 7 5 ,% ( 75(
,

0
*
& ! # 5 AChE * 5(
.
L
& .

+
6/

,

7

$+
4
"#" 5 AChE ("! Q organophosphate carbamate
,

Ellman‰s reaction (Ellman et. al. 1961) ( 7

Acetylthiocholine iodide AChE Thiocholine + Acetate

Thiocholine + DTNB yellow produce (! 6 )

!- ' $*
! .4 5 AChE * ' 9
"
#
1

/
,

acetylthiocholine iodide (ATChI) % ! 7 (substrate) 9
" 5( ! 5L

+/

& (Thiocholine) ("* ' 9
"
# 5,5′-dithio-bis-(2-nitrobenzoate) DTNB

6

1

5(! ! 6
( ! !- ' ! Q organophosphate carbamate *
/

+7

1
1
5 "#"
' 5 ' ! 9
" * & & ! 6 ! H

7
,

6/

)
/
* (5(*

(& ((
6 ! (absorbance) ( " & spectrophotometer '&
1
,
,
" & 6/ (wavelength) 412 nm
* & (#
'
5

7 ( 7
M˜
1. 1 67 "6/ ! " buffer phosphate (pH 8.0, 0.1
M) ! 67 "6/ 20 mg buffer phosphate 1 ml (" % & 6/ homogenizer
/

/
)
2. ((! "'5( 400 µl ('( ' buffer phosphate
+/
(pH 8.0, 0.1M) 2.6 ml ! " DTNB 0.01 M 100 µl " 5(*
" 5,5′-

dithio-bis-(2-nitrobenzoate) 39.6 mg buffer phosphate (pH 7.0, 0.1 M) 10 ml