Probing Bioactive Mechanisms - American Chemical Society

Chapter 9

Inhibition of Susceptible and Resistant Green Rice Leafhopper Acetylcholinesterase by N-Methylcarbamate and Oxadiazolone Insecticides 1

1

1

Hiroki Ohta , Noburo Kyomura , Yoji Takahashi , and Philip S. Magee

2

1Mitsubishi Kasei Corporation, Yokohama, Japan BIOSAR Research Project, Vallejo, CA 94591 and School of Medicine, University of California, San Francisco, CA 94143

Downloaded by CORNELL UNIV on October 6, 2016 | http://pubs.acs.org Publication Date: November 14, 1989 | doi: 10.1021/bk-1989-0413.ch009

2

Inhibition of AChE preparations from S- and R-strains of green rice leafhopper by aryl N-methylcarbamates (n = 20) and aryl oxadiazolones (n = 81) were evaluated with pI50 as the end-point. Regression against binding and reactivity descriptors was used to explore differ ences in mechanistic behavior. Ortho-substituted carbamates showed similar behavior against S- and R-AChE, but with different loadings and electronic effects that suggest a shift in mechanism. Oxadiazolones show a similar dependance on binding and steric factors with non-equivalent loadings for S- and R-AChE. Steric effects are much larger for ortho-substituted oxadiazolones than the related carbamates. These studies show that carbamates and oxadiazo lones bind differently at the active sites of AChE. The basic mechanism of resistant AChE inhibition remains the same, but occurs in a physically modified active site. One of the major pest insects in rice fields is the green rice leaf hopper (Nephotettix cincticeps). Recently acquired resistance to carbamate insecticides has greatly complicated control of this insect. Resistance can develop by a number of different mechanisms such as cuticle thickening to impede transport, enhanced metabolic degradation or molecular changes within the target enzyme (AChE) (1). In the case of carbamate insecticides, resistance occurs mainly at the target enzyme which expresses reduced sensitivity to the inhibi tors (2). The mechanistic details of carbamate resistance have been studied by reaction kinetics to show that decreased formation of the receptor complex (ECX) is the most important factor in the process (3). 0097-6156/89W13-0136$06.00A) © 1989 American Chemical Society

Magee et al.; Probing Bioactive Mechanisms ACS Symposium Series; American Chemical Society: Washington, DC, 1989.

9.

OHTA ET AL.

Inhibition of Green Rice Leafhopper Acetylcholinesterase 137

E

E

*


ECX

AChE *

CX

+

CX

1

v

ECX

- Carbamate I n h i b i t o r

R e c e p t o r Complex

ix

EC

EC

* Carbamoylated AChE

X « P h e n o l a t e L e a v i n g Group

E a r l y s t u d i e s of the r e l a t i o n o f a r y l N-methylcarbamate s t r u c t u r e w i t h a c t i v i t y and enzyme i n h i b i t i o n were f i r s t r e p o r t e d i n 1966 and showed r a t h e r s i m p l e dependance on s u b s t i t u e n t e f f e c t s f o r l i m i t e d s e t s of compounds (4, 5 ) . The a p p a r e n t s i m p l i c i t y was p a r t l y due to the f a c t t h a t QSAR t e c h n i q u e s were s t i l l i n the f i r s t decade o f development. The t r u e c o m p l e x i t y of carbamate i n h i b i t i o n i s r e v e a l e d i n l a t e r s t u d i e s by F u j i t a and co-workers on brown p l a n t hopper AChE 06, » and by Hansch and co-workers on h o u s e f l y head AChE ( 8 ) . The l a t t e r s t u d y shows t h a t 12 s i g n i f i c a n t f a c t o r s a r e r e q u i r e d to c o r r e l a t e the pI50 d a t a f o r a s e t o f 269 carbamates. The d e t a i l s of t h e s e s t u d i e s have been r e v i e w e d by Magee (9, 10). R e c e n t l y , a n o t h e r i m p o r t a n t c l a s s o f c h e m i c a l s based on the o x a d i a z o l o n e r i n g have shown a c t i v i t y s i m i l a r t o the carbamates, but w i t h s u r p r i s i n g a c t i v i t y on c a r b a m a t e - r e s i s t a n t green r i c e leafhopper ( S t r u c t u r e , T a b l e 2) (11-13). To i n v e s t i g a t e the r e s i s t a n c e mechan ism i n g r e e n r i c e l e a f h o p p e r , we d e c i d e d to e x p l o r e the s t r u c t u r e a c t i v i t y r e l a t i o n s of b o t h i n s e c t i c i d e c l a s s e s a g a i n s t AChE p r e p a r a t i o n s from s u s c e p t i b l e and r e s i s t a n t i n s e c t s . The d i f f e r e n c e i n the s u b s t i t u e n t e f f e c t s was e x p e c t e d to p r o v i d e c r i t i c a l i n f o r m a t i o n a t the m o l e c u l a r l e v e l of the r e c e p t o r complex. M a t e r i a l s and

Methods

Compounds S t u d i e d . Twenty O r t h o - s u b s t i t u t e d p h e n y l N - m e t h y l c a r b a mates were p r e p a r e d i n the c o n v e n t i o n a l manner by r e a c t i n g the s e l e c t e d p h e n o l s w i t h m e t h y l i s o c y a n a t e u s i n g a c a t a l y t i c amount o f t r i e t h y l a m i n e (14). The s t r u c t u r e s a r e l i s t e d i n T a b l e 1. Over e i g h t y N - a r y l o x a d i a z o l o n e s were s y n t h e s i z e d by a procedure d e s c r i b e d by Boesch i n a German p a t e n t ( 1 5 ) . T h i s c o n s i s t s of r e a c t i n g a r y l h y d r a z i n e s w i t h a c h l o r o f o r m i c e s t e r f o l l o w e d by r i n g c l o s u r e w i t h phosgene. R e p r e s e n t a t i v e s t r u c t u r e s a r e l i s t e d i n T a b l e 2. AChE P r e p a r a t i o n and A s s a y . Whole body enzyme p r e p a r a t i o n s o f s u s c e p t i b l e ( S ) and r e s i s t a n t ( R ) s t r a i n s o f g r e e n r i c e l e a f h o p p e r were p r e p a r e d a c c o r d i n g to a s t a n d a r d method used f o r f l y head AChE ( 1 6 ) . The crude enzyme p r e p a r a t i o n was i n h i b i t e d by s e r i a l d i l u t i o n s of carbamate and o x a d i a z o l o n e i n s e c t i c i d e s a t 25°C f o r 15 m i n u t e s . A s s a y s o f r e s i d u a l AChE were made by the E l l m a n s p e c t r o p h o t o m e t r i c p r o c e d u r e based on a c e t y l t h i o c h o l i n e and the c l e a v a b l e i n d i c a t o r , 5 , 5 - d i t h i o b i s - 2 - n i t r o b e n z o i c a c i d (17) . The n e g a t i v e l o g a r i t h m o f the molar c o n c e n t r a t i o n c a u s i n g 50% i n h i b i t i o n , pI50, i s d e r i v e d from the i n h i b i t i o n - c o n c e n t r a t i o n p l o t . 1

S t a t i s t i c a l Data A n a l y s i s . The pI50 d a t a were a n a l y z e d by s t a n d a r d m u l t i p l e r e g r e s s i o n a n a l y s i s u s i n g the Hansch program (18) a t M i t s u b i s h i and r e l a t e d programs a t BIOSAR. A l l programs were c r o s s -


138

PROBING BIOACTIVE MECHANISMS

v a l i d a t e d using standard data s e t s .

V a l u e s o f the

hydrophobic

c o n s t a n t ( ^ ) based on l o g P (octanol/water)» the Hammett sigma con s t a n t ( p ) and sigma i n d u c t i v e ( o j ) c o n s t a n t were taken from the a

Pomona C o l l e g e M e d i c i n a l C h e m i s t r y

P r o j e c t database

(19).

Sigma

resonance ( a ) v a l u e s were d e r i v e d from - o-j-. S t e r i c d e s c r i p t o r s s c a l e d to van der Waals r a d i i ( C h a r t o n s u ) were s e l e c t e d from the l i t e r a t u r e (20, 21). R

1


R e s u l t s and D i s c u s s i o n Ortho S u b s t i t u t e d P h e n y l N-Methylcarbamates. T a b l e 1 shows the enzyme i n h i b i t i o n d a t a f o r a l l o f the t e s t e d carbamates. With the s i n g l e e x c e p t i o n o f the 2-phenyl compound, a l l o f the carbamates were s t r o n g e r i n h i b i t o r s ( h i g h e r pI50) o f AChE from S - s t r a i n o f the leafhopper. T h i s c l e a r l y c o n f i r m s the s u p p o s i t i o n t h a t r e s i s t a n c e to carbamates i s due m a i n l y to changes i n enzyme i n h i b i t i o n . It i s a l s o c o n s i s t e n t w i t h a s t u d y by Hama and Iwata showing a c o r r e l a t i o n between the degree o f r e s i s t a n c e and b o t h i n s e c t i c i d a l a c t i v i t y and AChE i n h i b i t i o n f o r s e v e r a l carbamate i n s e c t i c i d e s ( 2 2 ) . E q u a t i o n s (2) and (3) a r e based on m u l t i p l e r e g r e s s i o n a n a l y s i s of the d a t a i n T a b l e 1. In the e q u a t i o n s , w i s the h y d r o p h o b i c constant, u i s C h a r t o n s s t e r i c c o n s t a n t , 0 j and o a r e the i n d u c t i v e and resonance components o f Hammett e l e c t r o n i c c o n s t a n t s . V a l u e s i n p a r e n t h e s e s a r e the 95% c o n f i d e n c e i n t e r v a l s o f the r e g r e s sion coefficients. The o t h e r v a l u e s a r e n (number o f s a m p l e s ) , r ( c o r r e l a t i o n c o e f f i c i e n t ) and s ( s t a n d a r d e r r o r o f e s t i m a t e ) . 1

Q

R

pI50(S) - 0.34 TT (0.35) + 1.65 u n - 19 pI50(R)

88

r - 0.900

r - 0.916

u

Q

a

(0.87) - 0.74

s - 0.320

0.56 TT (0.23) + 0.99

ii-20

Q

F -

(0.59) + 1.63

s - 0.233

R

(0.51) +3.74 (0.45) (2)

21.62 o

1

(0.68) + 2.57

(0.36) (3)

F - 27.66

Both e q u a t i o n s show s i m i l a r but d i f f e r e n t r e s p o n s e s to hydropho b i c c h a r a c t e r and s t e r i c s i z e . The s t e r i c s e n s i t i v i t y o f the Ss t r a i n i s s u b s t a n t i a l l y more accomodating than the R - s t r a i n , though the b i n d i n g e f f e c t i s p o s i t i v e f o r b o t h enzymes. The most s t r i k i n g d i f f e r e n c e i s o b s e r v e d i n the e l e c t r o n i c b e h a v i o r w i t h resonance i m p o r t a n t i n the S - s t r a i n and i n d u c t i o n i n the R - s t r a i n . This s t r o n g l y s u g g e s t s a d i f f e r e n c e i n the r a t e - d e t e r m i n i n g s t e p f o r c a r b a m o y l a t i o n o f S and R AChE. In s t u d i e s on house f l y head AChe, Kamoshita e t a l d e v e l o p e d e q u a t i o n (4) f o r 16 o r t h o - s u b s t i t u t e d p h e n y l N-methylcarbamates ( 6 ) . Log 1/K^ i s r e a s o n a b l y c o l l i n e a r w i t h pI50, but d i f f e r e n t l y s c a l e d , so we cannot compare the magnitudes o f the r e g r e s s i o n c o e f f i c i e n t s i n e q u a t i o n s ( 2 - 4 ) . The l o n g e r i n c u b a t i o n times used i n t h i s s t u d y may reduce the c o l l i n e a r i t y and make comparison l e s s p r e c i s e . How e v e r , they observe a p o s i t i v e dependence on h y d r o p h o b i c i t y as we do. As o° i s l a r g e l y i n d u c t i v e , t h e i r dependence i s s i m i l a r to t h a t found i n e q u a t i o n ( 3 ) . The HB d e s c r i p t o r (HB » 1.0 f o r H-bonding Ortho group) i n e q u a t i o n (4) was i n s i g n i f i c a n t i n our s t u d i e s d e s p i t e


Inhibition of Green Rice Leafhopper Acetylcholinesterase

OHTA ET AL.

TABLE 1.

Anti-AChe a c t i v i t y o f a r y l N-methylcarbamates on s u s c e p t i b l e ( S ) and r e s i s t a n t ( R ) N e p h o t e t t i x c i n c t i c e p s


N —

X

PI50(S)

pl50( )

H

3.89

2.50

Me

4.43

3.20

Et

4.89

3.74

i-Pr

5.70

4.21

sec-Bu

6.04

4.60

CH^CH^CH-^

5.07

3.96

OMe

4.77

3.26

OEt

4.60

3.52

O-i-Pr

6.30

3.82

O-sec-Bu

5.68

4.17

0-CH CH=CH

R

5.15

3.96

0-CH C=CH

5.09

4.10

CI

5.52

4.26

2

2

2

Br

5.66

4.64

NH-i-Pr

5.64

3.77

NMe

5.02

3.89

6.14

4.80

CN

4.15

3.64

Phenyl

3.74

4.10

CH-NOMe

4.77

3.74

2

N(Allyl)

2


140

PROBING BIOACTIVE MECHANISMS

the p r e s e n c e o f many H-bonding groups i n our d a t a s e t ( T a b l e 1 ) . The o t h e r s i g n i f i c a n t d i f f e r e n c e i s the l a c k o f a p o s i t i v e s t e r i c e f f e c t i n t h e i r study. As n, r and s a r e comparable i n e q u a t i o n s ( 2 - 4 ) , the d i f f e r e n c e s p o i n t to r e a l v a r i a t i o n s i n the b i n d i n g mechanisms of each AChE. Log

1/K

d

- 1.558

IT (0.354) +

1.009

c°

(0.508) + 1.26 HB

(0.313) +

3.998 (0.294)


n - 16

r « 0.953

(4)

s « 0.255

N-Aryl Oxadiazolones. Compounds s t u d i e d (n = 81) a r e shown i n T a b l e 2 w i t h the a s s o c i a t e d pI50 v a l u e s f o r R and S l e a f h o p p e r AChE. As a c l a s s , these compounds a r e s u b s t a n t i a l l y s t r o n g e r i n h i b i t o r s than the a r y l N-methylcarbamates (compare T a b l e s 1 and 2 ) . Moreover, t h e r e i s no c r o s s - r e s i s t a n c e w i t h carbamates as a l l a r e s t r o n g e r i n h i b i t o r s o f the R AChE than S AChE. In a d d i t i o n , Ambrosi e t a l r e p o r t the 2-methoxy a n a l o g (RP-32,861) t o show h i g h i n s e c t i c i d a l a c t i v i t y a g a i n s t r e s i s t a n t g r e e n r i c e l e a f h o p p e r ( 1 1 ) , w h i l e Bakry et a l r e p o r t p o t e n t i n h i b i t i o n o f h o u s e f l y and e l e c t r i c e e l A C h E s f o r the same a n a l o g ( 1 3 ) . There i s l i t t l e doubt t h a t the o x a d i a z o l o n e s a r e b r o a d , g e n e r a l AChE i n h i b i t o r s . Huang e t a l have r e c e n t l y s t u d i e d the c h e m i c a l r e a c t i v i t y o f the o x a d i a z o l o n e r i n g and p r o p o s e n u c l e o p h i l i c a t t a c k o f s e r i n e - o x y l a n i o n on c a r b o n y l as the p r o b a b l e i n h i b i t i o n mechanism ( 2 3 ) . Our own experiments on o x a d i a z o l o n e r i n g - o p e n i n g w i t h methoxide i o n s u p p o r t t h e i r work ( 5 ) . The d r i v i n g f o r c e f o r t h i s r e a c t i o n i s s u b s t a n t i a l as a d e l o c a l i z e d carbamate group i s formed from the i m i n o e s t e r l e a v i n g group. We b e l i e v e the same r i n g - o p e n i n g t o be r e s p o n s i b l e f o r the o b s e r v e d l a b i l i t y o f these compounds i n b o t h i n s e c t t e s t i n g and pI50 measurement. 1

Ar (5)

CH 0 3

There a r e b o t h s i m i l a r i t i e s and d i f f e r e n c e s i n the AChE i n h i b i t i o n mechanisms o f carbamates and o x a d i a z o l o n e s . Both p r o c e e d by r e v e r s i b l e b i n d i n g and i r r e v e r s i b l e c a r b o n y l a t t a c k by a c t i v a t e d s e r i n e - o x y l i o n . However, the o x a d i a z o l o n e s r e a c t by r i n g - o p e n i n g l e a v i n g a l a r g e , complex s t r u c t u r e a t the s i t e o f i n h i b i t i o n . By c o n t r a s t , the carbamates r e a c t by p h e n o l a t e d i s p l a c e m e n t l e a v i n g a s i m p l e c a r b a m o y l a t e d enzyme. When the pI50 d a t a a r e examined by r e g r e s s i o n a n a l y s i s , i t becomes c l e a r t h a t the mode o f b i n d i n g i s q u i t e d i f f e r e n t from the a r y l N-methylcarbamates. E q u a t i o n s (6) and (7) a r e s t a t i s t i c a l l y s i g n i f i c a n t but do not c o r r e l a t e w i t h the s h a r p n e s s o f the carbamate d a t a , a f a c t we a t t r i bute to ongoing d e g r a d a t i o n d u r i n g the pI50 a s s a y . U n l i k e the c a r bamate s t u d y , the c o r r e l a t i n g f a c t o r s f o r R and S AChE a r e i d e n t i c a l but w i t h d i f f e r e n t r e s p o n s e s . The o r t h o s t e r i c e f f e c t i s n e g a t i v e f o r the o x a d i a z o l o n e s and n e a r l y t w i c e as s t r o n g f o r the S AChE, a c c o u n t i n g f o r much o f the r e v e r s e s e l e c t i o n [pI50(R) > p I 5 0 ( S ) ] .


9.

Inhibition of Green Rice Leafhopper Acetylcholinesterase 141

O H T A ET AL.

T a b l e 2.

Anti-AChE a c t i v i t y s u s c e p t i b l e ( S ) and cincticeps

o f N - a r y l o x a d i a z o l o n e s on resistant(R) Nephotettix

x

z


o

X

R

Z

Y

z

pl50(S)

R

PI50( >

H

H

H

Me

4.82

4.82

Me

H

H

Me

5.92

6.82

CI

H

H

Me

5.92

6.82

Br

H

H

Me

6.43

7.39

OMe

H

H

Me

5.64

6.25

O-i-Pr

H

H

Me

5.66

6.20

OMe

H

H

Et

5.22

6.66

O-i-Pr

H

H

Et

4.80

6.49

H

H

3-Me

Me

5.82

5.89

H

H

3-i-Pr

Me

6.85

6.43

H

H

3-t-Bu

Me

6.59

6.85

H

H

3-C1

Me

5.14

5.16

H

H

3-N0

Me

4.80

5.44

H

H

3-t-Bu

Et

5.92

6.33

H

Me

H

Me

5.24

5.41

H

F

H

Me

4.20

5.06

H

CI

H

Me

4.64

4.15

i-Pr

Me

H

Me

4.77

5.70

OMe

Me

H

Me

6.22

7.00

OMe

F

H

Me

5.41

5.96

OMe

N0

H

Me

6.32

6.33

OEt

Me

H

Me

6.30

6.96

OEt

Et

H

Me

6.59

7.57

OEt

F

H

Me

5.82

6.22

OEt

CI

H

Me

6.05

6.48

2

2

Continued on next page


142

PROBING BIOACTIVE MECHANISMS Table 2. Continued


X

Y

Z

R

pl50 MeO) a r e o p p o s i t e l y s e l e c t i v e .


G

pI50(S) = 0.32

IOR

n - 81

-1.50

r - 0.608

IOR

n - 81

0

(0.69) - 0.55

(0.29) + 0.41

pI50(R) « -0.83 0.56

u

u

G

(0.49) + 1.66

IX

Q

(0.65) -

(0.43) + 5.62

s = 0.627

(0.64) -

(0.28) + 0.77 r - 0.694

HB

I o

HB

F -

0.82 lo

(6)

8.80

(0.46) + 1.89

IX

Q

(0.61) +

(0.41) + 5.77

s « 0.494

F *

(7)

13.94

The low e x p l a i n e d v a r i a n c e o f t h e s e e q u a t i o n s (100r^ » 37 - 48) i s n o t a t t r i b u t a b l e t o e x p e r i m e n t a l e r r o r i n the pI50 measurement as the same t e c h n i q u e was used f o r the carbamates ( 1 0 0 r = 81 - 8 4 ) . The p r o b a b l e l o s s i n p r e c i s i o n i s due t o v a r i a b l e d e g r e e s o f h y d r o l y s i s d u r i n g the measurement time span. T h i s was t e s t e d by l o o k i n g a t the d i f f e r e n c e between pI50(R) and p I 5 0 ( S ) , a p r o c e d u r e t h a t s h o u l d remove much o f the v a r i a n c e due t o s e l e c t i v e h y d r o l y s i s o f each compound. We f i n d the d i f f e r e n c e t o c o r r e l a t e s i g n i f i c a n t l y b e t t e r than e i t h e r pI50 ( 1 0 0 r • 5 8 ) , s u p p o r t i n g d i f f e r e n t i a l h y d r o l y s i s as a w i l d v a r i a b l e . Note t h a t the e l e c t r o n i c e f f e c t (Zo ) and v a r i a b l e I X were n o t s u f f i c i e n t l y d i f f e r e n t t o show up s t a t i s t i c a l l y , a l t h o u g h r , s and F a r e m a r k e d l y improved. The key f a c t o r s s u p p o r t i n g g r e a t e r i n h i b i t i o n o f the R AChE a r e seen t o be a l e s s u n f a v o r a b l e s t e r i c e f f e c t and s t r o n g e r H-bonding o f the ortho-OR groups. There may, o f c o u r s e , be a m i s s i n g f a c t o r t h a t would b r i n g r t o 0.85-0.90, but i t cannot be one n o r m a l l y e x p l o r e d i n SAR studies. 2

2

Q

pI50(R) - pI50(S) - 0.79

u

Q

(0.50) + 0.90

IOR (0.25)+ 0.34 HB

(0.28) +

0.14 n - 81

r « 0.760

s * 0.443

(8)

F - 34.99

Conclusions I t i s c l e a r from the SAR a n a l y s i s o f the pI50 r e s p o n s e t h a t c a r b a mates and o x a d i a z o l o n e s do n o t i n h i b i t e i t h e r s u s c e p t i b l e - o r r e s i s t a n t - s t r a i n GR l e a f h o p p e r AChE by a n a l o g o u s mechanisms. This



9.

OHTA ET AL.

Inhibition of Green Rice Leafhopper Acetylcholinesterase

i s somewhat s u r p r i s i n g as i r r e v e r s i b l e i n h i b i t i o n p r o c e e d s by a n a l o gous a t t a c k o f s e r i n e - o x y l i o n on p o s i t i o n a l l y r e l a t e d c a r b o n y l groups o f b o t h c l a s s e s . The d r a m a t i c d i f f e r e n c e s o b s e r v e d must be r e l a t e d t o the mode o f b i n d i n g , i n p a r t i c u l a r , t o non-analogous binding regions. While b o t h c l a s s e s must b i n d t o f a v o r a b l y p o s i t i o n the c a r b o n y l groups f o r s e r i n e a t t a c k , t h e a r o m a t i c r i n g s a r e bound in distinctly different sites. T h i s i s r e v e a l e d by s e v e r a l f a c t o r s . The carbamates show a dependence on l i p o p h i l i c i t y , an accommodation o f o r t h o s t e r i c e f f e c t s and no dependence on H-bonding p o t e n t i a l . C o n v e r s e l y , t h e o x a d i a z o l o n e s show no dependence on l i p o p h i l i c i t y , n e g a t i v e o r t h o s t e r i c e f f e c t s and s u b s t a n t i a l pI50 enhancement f o r H-bonding groups. These e f f e c t s a r e i n c o n s i s t e n t w i t h a r y l r i n g b i n d i n g a t t h e same s i t e l o c a t i o n . I n a d d i t i o n , carbamates a r e more a c t i v e on the s u s c e p t i b l e s t r a i n w h i l e o x a d i a z o l o n e s show g r e a t e r a c t i v i t y on t h e r e s i s t a n t s t r a i n . While n o t t e s t e d i n d e t a i l , t h i s would s u g g e s t the p r o b a b l e absence o f c r o s s - r e s i s t a n c e . There appears t o be a s h i f t i n e l e c t r o n i c mechanism i n t h e carbamate i n h i b i t i o n o f t h e S- and R - s t r a i n s o f AChE. The dependence of o r t h o groups on l i p o p h i l i c i t y and an accommodating s t e r i c e f f e c t i s s i m i l a r i n b o t h c a s e s though d i f f e r e n t i n magnitude. The much lower s t e r i c accommodation o f t h e R - s t r a i n i s p r o b a b l y t h e major f a c t o r i n weakening i t s r e s p o n s e t o carbamate i n h i b i t i o n . The d i f f e r ing e l e c t r o n i c e f f e c t s suggest a s h i f t i n the rate-determining t r a n s i t i o n s t a t e f o r c a r b a m o y l a t i o n o f t h e s e r i n e hydroxy1 group. I n h i b i t i o n o f S- and R - s t r a i n s o f AChE by o x a d i a z o l o n e s a r e found t o c o r r e l a t e w i t h i d e n t i c a l f a c t o r s . The d i f f e r e n c e e q u a t i o n [pI50(R) - p I 5 0 ( S ) ] shows major v a r i a n c e s i n r e s p o n s e t o o r t h o s t e r i c e f f e c t s , t o OMe v s . OEt i n the h e t e r o - r i n g and t o the H-bond i n g s t r e n g t h o f some g r o u p s . I n b r i e f , h i g h e r pI50 v a l u e s f o r t h e R - s t r a i n a r e s u p p o r t e d by lower s t e r i c r e p u l s i o n , s t r o n g e r H-bonding, and by OEt s u b s t i t u t i o n o f t h e h e t e r o - r i n g (OMe f a v o r e d by S - s t r a i n ) . These s t u d i e s c l e a r l y i d e n t i f y the major r e s i s t a n c e mechanism i n the g r e e n r i c e l e a f h o p p e r as i n v o l v i n g a m o d i f i e d AChE t a r g e t s i t e . T h i s c o u l d a r i s e a t the g e n e t i c l e v e l o r more p r o b a b l y by s e l e c t i v e c o n c e n t r a t i o n o f an e x i s t i n g isozyme i n t h e S - s t r a i n . In e i t h e r c a s e , i t i s i m p o r t a n t t o note t h a t the b a s i c i n h i b i t i o n mechanism remains t h e same f o r b o t h carbamates and o x a d i a z o l o n e s v s . S- and R - s t r a i n AChE. The d i f f e r e n c e s d e t e c t e d by the pI50 r e s p o n s e a n a l y s i s a r e c o n s i s t e n t w i t h isozyme s e l e c t i o n where t h e a c t i v e s i t e s d i f f e r by one o r two s t r u c t u r a l amino a c i d s n o t d i r e c t l y i n v o l v e d i n the b a s i c AChE mechanism. I n b r i e f , a s i m p l e change o f c a v i t y shape could e a s i l y account f o r the observed d i f f e r e n c e s . Of g r e a t e s t importance i s t h e ease w i t h w h i c h t h e s e SAR a n a l y s e s were a b l e t o p i n - p o i n t t h e m e c h a n i s t i c d i f f e r e n c e s b o t h between t h e two c l a s s e s and w i t h i n each c l a s s .

Literature Cited 1. 2. 3. 4.

Takahashi, Y. Pesticide Design-Strategy and Tactics; Eto, M. Ed; Soft Science, Inc. 1979, 674. Iwata, T . ; Hama, H. J. Econ. Ent. 1971, 65, 643. Yamamoto, I.; Kyomura, N.; Takahashi, Y. J . Pestic. Sci. 1977, 2, 463. Metcalf, R. L. Bull. W. H. O. 1971, 44, 43.


146 5. 6. 7. 8. 9. 10.


11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23.

PROBING BIOACTIVE

MECHANISMS

Hansch, C.; Deutsch, E. W. Biochem, Biophys. Acta 1966, 126, 177. Kamoshita, K.; Ohno, I.; Fujita, T.; Nishioka, T.; Nakajima, M. Pestic. Biochem. Physiol. 1979, 11, 83. Kamoshita, K.; Ohno, I.; Kasamatsu, K.; Fujita, T.; Nakajima, M. Pestic. Biochem. Physiol. 1979, 11, 104. Goldblum, A.; Yoshimoto, M.; Hansch, C. J . Agric. Food Chem. 1981, 29, 277. Magee, P. S. In Quantitative Structure-Activity Relationships of Drugs; Topliss, J . G . , Ed.; Academic Press, New York 1983, 393-436. Magee, P. S. In Insecticide Mode of Action; Coats, J . R. Ed.; Academic Press, New York 1982, 101-161. Ambrosi, D.; Bic, G.; Desmoras, J.; Gallinelli, G.; Roussel, G. Proc. Br. Crop Prot. Conf. Pests. D i s . 1979, 533. Ambrosi, D.; Boesch, R.; Desmoras, J . J . Phytiatrie-Phytopharmacie 1980, 199. Bakry, N. M.; Sherby, S.; Elderfrawi, A. T.; Elderfrawi, M. E. Neurotoxicology 1986, 7, 1. Kolbezen, M. J.; Metcalf, R. L.; Fukuto, T. R. J . Agr. Food Chem 1954, 2, 864. Boesch, K. Ger. Offen. 1976, 2,603,877. Reed, W. D.; Fukuto, T. R. Pest. Biochem. Physiol. 1973, 3, 120. Ellman, G. L.; Courtney, K. D.; Andres, V. J r . ; Featherstone, R. M. Biochem. Pharmacol. 1961, 7, 88. Hansch, C.; Fujita, T. J . Am. Chem. Soc. 1964, 86, 1616. Hansch, C.; Leo, A. Substituent Constants for Correlation Analysis In Chemistry and Biology, Wiley, New York 1979. Charton, M. J. Am. Chem. Soc. 1975, 97, 1552. Charton, M. J. Org. Chem. 1976, 41, 2217. Hama, H . ; Iwata, T. Appl. Ent. Zool. 1971, 6, 183. Huang, J.; Bushey, D. F. J . Agr. Food Chem. 1987, 35, 368.

RECEIVED June 19, 1989


Probing Bioactive Mechanisms - American Chemical Society

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