New plant growth-regulators
by Jack R Gaines
A THESIS Submitted to the Graduate Faculty in partial fulfillment of the requirements for the degree
of Doctor of Philosophy in Chemistry
Montana State University
© Copyright by Jack R Gaines (1956)
Abstract:
A comprehensive review and discussion of known synthetic plant growth-regulators is given. Two
main series of compounds, the substituted phenylhydrazono-acetic and propionic acids, were
synthesized and tested for activity as plant growth-regulators, The parent acids,
phenylhydra-zonoacetic acid and α-phenylhydrazonopropionic acid, were found to be highly active. All
of the substituted derivatives were considerably less active and in most cases were inhibitory.
α-Phenyliminopropionic acid, ethyl β-(p-chlorophenylamino)-acrylate, and ethyl
β-(p-methylphenylamino)-acrylate were also prepared and tested. All three of these compounds, with
the possible exception of the p-chloro-compound, were inactive. 2-Nitro-4-methoxyphenoxyacetic acid
was prepared and tested in order to test a hypothesis proposed by the author. NEWr PLANT GROWTH-REGULATORS
by
JACK R. GAINES
A THESIS
S u bm itted to th e G raduate F a c u lty
in
p a r t i a l f u l f i l l m e n t o f t h e re q u ire m e n ts
f o r t h e d e g re e of
_y D octor o f P h ilo so p h y i n C hem istry
Montana S ta te C o lleg e
Approved:
Head* M ajor'D epartm ent
-t^ u
HESTRICTEO STACK
3 V
—2 —
TABLE OF CONTENTS
E£I.
ABSTRACT ...........................................................................
4
II.
INTRODUCTION ..................................................................
5
HISTORICAL AND THEORETICAL DISCUSSION
. , .
7
Chemical n a tu r e o f th e a u x in s ..................
7
M ethods o f t e s t i n g
8
III.
.......................................
S tr u c tu r e and a c t i v i t y
...............................
9
The r in g system ................................................
13
The s id e c h ain
14
................................................
P h e n y la c e tic a c id and r e l a t e d compounds
15
In d o le - 3 - a lk y lc a r b o x y lic a c id s
. . . .
16
Phenoxy compounds ..................................... , .
17
The b en zo ic a c id s ............................................
26
A c tiv ity and o p t i c a l isom erism
. . . .
29
...................................
29
New p la n t g ro w th -re g u la to rs ......................
30
EXPERIMENTAL..................................................................
32
No n -a c id ic compounds
IV .
P r e p a r a tio n o f s o lu tio n s
...........................
32
&-PhenyIh y d raz o n o p ro p io n ic a c id . . . .
33
P h e n y lh y d ra z o n o ac e tic a c id
......................
33
<&-Phenyliminopropionic a c i d ......................
34
E th y lfo rm y la c e ta te , sodium e n o la te
. .
35
E th y l / 6 - a n i l i n o a c r y l a t e ...............................
35
2 .4 -
D ic h lo r o a c e ta n ilid e . .......................
35
2 .4 -
D ic h lo ro a n ilin e
36
................................
118855
- 3 -
( ta b le o f c o n te n ts )
£S
2 . 4 - D ic h lo ro p h en y lh y d razin e h y d ro c h lo rid e ....................... .
37
p~T olyhydrazin e h y d ro c h lo rid e .........................................................
38
p -C h lo ro p h e n y lh y d ra z in e h y d ro c h lo rid e . . . . . . . . . .
39
o -C h lo ro p h en y lh y d razin e h y d ro c h lo rid e ........................... . . .
39
2 . 5- D ic h lo ro p h e n y lh y d ra zin e h y d ro c h lo rid e ...........................
39
At-( p -T o ly h y d razono) - p r o p io n ic a c i d ........................................
E th y l /8 - ( p-m eth y l a n ilin o ) - a c r y l a t e
.
............................................
40
(K-( o-G hlorophenylhydrazono)-p r o p io n ic a c i d ......................
# -(p -O h lp ro p h e n y lh y d ra z o n o )-p ro p io n ic a c id
.
41
E th y l 2 ,4 - d .ic h lo r o p h e n y lh y d r a z o n o a c e ta te ...............................
41
2 ,4 -D ic h lo ro p h e n y lh y d ra z o n o a c e tic a c id
42
....................................
..........................
E th y l 2 , 5 -d ic h lo ro p h e n y lh y d ra z o n o a c e ta te
.
42
. . .......................
43
a -(2 ,4 -d ic h lo r o p h e n y lh y d r a z ono)- p r o p io n ic a c id
. . . . . .
E th y l y f f - ( p - c h l o r o a n i l i n o ) - a c r y l a t e ........................................
43
.
E th y l dl-phenylhydraz onopropi o nat e ................................................
<£-(p-brom ophenylhydrazono) - p r o p io n ic a c i d .......................
2 -N itro -l,4 -d im e th o x y b e n z e n e
V II.
43
44
.
44
.....
44
.........................................................
45
Ott-( p -n itro p h e n y lh y d ra z ono)-p r o p io n ic acid.
V I.
40
...........................
E th y l CO-ch lo ro p h en y Ih y d raz o n o a ce ta t e
7.
40
2 -N itro -4 -n ie th o x y p h e n o l ....................................................
45
2 -N itro -4 -m e th o x y p h e n o x y ac e tic a c id . . . . . . . . . . .
46
DISCUSSION OF EXPERIMENTAL RESULTS
.....................................................
47
ACKNOWLEDGMENT..................................................... ........................................... ..
53
LITERATURE CITED AND CONSULTED
54
. . . . . . . . .
......................
- 4 I . ABSTRACT
A com prehensive re v ie w and d is c u s s io n o f known s y n th e tic p la n t gro w th r e g u la to r s i s g iv e n .
Two m ain s e r i e s o f compounds, th e s u b s tit u te d
phenyIh y d ra z on o -ac e t i c and p ro p io n ic a c id s , were s y n th e s iz e d and t e s t e d
f o r a c t i v i t y a s p la n t g r o w th - r e g u la to r s ,
The p a re n t a c id s , phenylh y d ra -
z o n o a c e tic a c id and <&-phenylhydrazo n o p ro p io n ic a c id , were found t o be
h ig h ly a c t i v e .
A ll o f t h e s u b s t i t u t e d d e r iv a tiv e s were c o n s id e ra b ly l e s s
a c ti v e and i n most c a se s were i n h i b i t o r y .
AC-Phenylim in o p ro p io n ic a c id , e th y l^ - ( p - c h lo r o p h e n y la m in o ) - a c r y la te ,
and e th y l ^ -(p -m e th y lp h en y lam in o )- a c r y l a t e were a l s o p re p a re d and t e s t e d .
A ll t h r e e of t h e s e . compounds, w ith t h e p o s s ib le e x c e p tio n of th e p ^ c h lo ro compound, w ere i n a c t i v e .
2 - N itro-4-m eth o x y p h en o x y acetic a c id was p re p a re d
and t e s t e d i n o rd e r t o t e s t a h y p o th e s is pro p o sed by th e a u th o r.
x
I I . INTRODUCTION.
I n 1938 K p e p fli, Thimann and Went ($6) p u b lish e d t h e f i r s t compre­
h e n siv e r e p o r t on s y n th e tic p la n t g ro w th - r e g u la to r s .
From t h e i r e x p e r i­
m e n ta l e v id e n c e , th e y concluded t h a t th e minimum s t r u c t u r a l re q u ire m e n ts
f o r c e l l e lo n g a tio n a c t i v i t y i n h ig h e r p la n ts a r e : (a ) a r i n g system a s
n u c le u s , (b) a d o u b le bond i n t h i s r i n g , (c ) a s id e c h a in , (d ) a c arb o x y l
group ( o r a s t r u c t u r e r e a d i l y c o n v erted t o a carb o x y l gro u p ) on t h i s s id e
c h a in a t l e a s t one carbon atom removed from th e r i n g , and (e) a p a r t i c u l a r
space r e l a t i o n s h i p betw een th e r in g and t h e c arb o x y l g ro u p .
Follow ing
t h i s s t r u c t u r a l re q u ire m e n t, Zimmerman and H itchcock (126) t e s t e d a s e r i e s
o f p h en o x y ac e tic a c id s and b en zo ic a c id s and found i n t h e case o f c e r ta in
h a lo g en d e r iv a tiv e s rem ark ab le grow th r e g u la tin g a c t i v i t y .
The most prom­
i s i n g o f t h e s e compounds b e in g 2 ,4 - d i C h lo ro p h en o x y acetic a c id (2 ,4 -D ). Up’
t o th e p re s e n t tim e , t h i s compound and i t s d e r iv a tiv e s have rem ained th e
m ost a c ti v e o f a l l s y n th e tic g ro w th - re g u la to r s .
S in ce t h i s work o f Zimmerman and H itc h co c k , a la r g e and d iv e rs e group
o f o rg a n ic compounds has been r e p o r te d t o r e g u la te p la n t grow th.
Many o f
'
t h e d e te r m in a tio n s , how ever, do n o t m easure th e a c c e le r a ti n g e f f e c t s on
grow th, b u t t h e i n h i b i t i o n o f growth o r t o x i c i t y e f f e c t s .
Such evidence
does n o t s u f f ic e t o i d e n t i f y th e su b sta n c e a s grow th r e g u la to r s which may
e i t h e r in c r e a s e o r i n h i b i t grow th depending upon t h e i r c o n c e n tra tio n .
W ith th e e x c e p tio n o f a few i s o l a t e d compounds o n ly t h e fo llo w in g
s e r i e s o f compounds have shown prom ise a s t r u e g ro w th -re g u la tin g compounds:
t h e b e n zo ic a c i d s , th e p h en o x y ak y lcarb o x y lic a c id s , th e th iio -p h en o x y ak y lc a r b o x y lic a c id s , th e n a p h th o x y a lk y lc a rb o x y lic a c id s , th e in d o le a lk y lc a r -
- 6 T.
boxyI i c a c id s , and t h e p h e n y la lk y lc a rb oxyI i c a c id s .
Of th e s e s e r i e s , only
th e b e n zo ic and p h e n o x y ac e tic a c id s have re c e iv e d c o n s id e ra b le a t t e n t i o n ;
e s p e c ia ll y from th e s ta n d p o in t o f r e l a t i n g chem ical s t r u c t u r e w ith p h y sio ­
lo g ic a l a c ti v ity .
The purpose o f t h i s re s e a rc h w as, th ro u g h s t r u c t u r a l a n alo g y , t o
p ro p o se and s y n th e s iz e a new s e r i e s o f compounds having g ro w th -re g u la tin g
p ro p e rtie s .
I I I . HISTORICAL AND THEORETICAL DISCUSSION
Chem ical N ature o f th e Auxins
Bonner and Wildman (18) in d ic a te d i n 1946 t h a t in d o le a c e tic a c id (IAA)
i s a p r i n c i p l e n a tiv e a u x in i n h ig h e r p l a n t s .
W ith th e developm ent o f
chrom atography, c o u n te r c u r re n t d i s t r i b u t i o n , and sp e c tro p h o to m e tric m icro ­
m ethods, t h e i d e n t i f i c a t i o n and e stim a tio n , o f p la n t grow th su b sta n c e s have
been ex ten d ed (9 7 ,1 2 3 ,9 8 ,3 5 ,6 6 ,9 9 ) .
I n a t l e a s t fo u r in s ta n c e s , th e grow th su b sta n c e a c t i v i t y of a p la n t
c o n c e n tra te h a s been found to r e s id e i n p a r t i n m a te r ia ls n o t i d e n t i c a l t o
IAAe
I n a l l i n s ta n c e s , however? t h e m a te r ia l i n q u e s tio n has tu rn e d out t o
be e i t h e r a d e r iv a tiv e o f IAA or c lo s e ly r e l a t e d t o i t .
■
1
The f i r s t o f t h e s e
•
was th e c a s e o f t h e n e u t r a l growth su b sta n c e d e sc rib e d by L arsen (6 l)«
T h is was i d e n t i f i e d a s in d o le a c e ta ld e h y d e and i s a c ti v e o n ly a f t e r con­
v e r s io n i n t h e p la n t t o IAA (6 2 ) .
A second case i s t h a t o f t h e f r u i t set*?
t i n g f a c t o r c o n ta in e d i n im m ature corn k e r n e ls ( 8 5 ) .
T h is su b sta n c e , which
i s r e p o r te d ly 100 tim e s more a c ti v e th a n IAA, was shown to b e th e e th y l
e s t e r o f IAA ( 8 5 ) .
T h ir d ly , i n t h e c o u n te rc u rre n t d i s t r i b u t i o n of th e a c id -
id n f r a c t i o n of cabbage le a v e s , H o lle y e t . a l . (50) fo u n d , i n a d d itio n t o IAA,
two s e p a ra b le a lth o u g h m inor com ponents, b o th a c ti v e i n th e Avena c u rv a tu re
te s t.
A lthough th e chem ical n a tu re o f th e s e s u b sta n c e s h as n o t been e s ta b ­
lis h e d , b o th g iv e th e Salkow skl r e a c tio n and a re p o s s ib ly r e l a t e d t o IAA i n
some m anner.
F i n a l l y , Jo n es e t . a l . (54) i s o l a t e d i n d o l e a c e t o n i t r i l e from •
p la n t t i s s u e and i t was found t o be more e f f e c t i v e a s an a u x in f o r Avena
c o le o p t ile t i s s u e th a n IAA (1 2 ) .
Stowe and Thimann (93) have re p o rte d ,
how ever, t h a t IAA can be d e te c te d by p a p er chrom atography i n s o lu tio n s
I
-
8
-
o f i n d b l e a c e t o n i t r i l e on which Aveha c o le o p t ile s e c tio n s grow f o r
24 h o u rs
a s w e ll a s i n e x t r a c t s o f th e c o l e o p t i l e t i s s u e , th u s i n d ic a tin g t h a t t h i s
t i s s u e c o n v e rts th e n i t r i l e t o t h e a c i d .
Thimann (103) r e p o r ts t h a t th e
n i t r i l e i s i n a c t i v e in th e pea t e s t , b u t th e e th e r e x tr a c t o f c o le o p tile
s e c tio n s grown f o r 48 h o u rs on a s o lu tio n o f t h e n i t r i l e c au se s a re sp o n se
i n t h e pea t e s t which would in d i c a t e a c o n v e rsio n o f more th a n 50% by th e
c o le o p tile tis s u e .
Thus th e r e i s s t i l l no s tro n g ev id en ce t o in d ic a te th e
p re s e n c e o f any n a tiv e grow th su b sta n c e o th e r th a n IAA.
M ethods o f T e s tin g
The p r i n c i p l e d i f f i c u l t y i n a p p r a is in g a l l th e l i t e r a t u r e p e r ta in in g
t o chem ical s tr u c t u r e and p la n t grow th a c t i v i t y l i e s i n t h e c o r r e l a t i o n o f
d a ta d e riv e d from many d i f f e r e n t p h y s io lo g ic a l re s p o n s e s .
Of a l l th e
re s p o n se s u se d , such a s in d u c tio n o f p a rth e h o c a rp y ( 67, 130) , organ m o d ifi­
c a tio n ( 97, 129, 131) , c o n tr o l o f a b s c is s io n ( l i d ) , i n i t i a t i o n o f r o o ts (4 6 ,
127) , c b l l e lo n g a tio n ( 16, 56, 13, 17, 72) , i n h i b i t i o n o f r o o t grow th ( 2 , 14, 2 0 ,
3 6 , 4 5 , 84, 95) and seed g e rm in a tio n (2 8 ) , th e s tim u la tio n o f c e l l e lo n g a tio n
h a s been u sed m ost e x te n s iv e ly .
Due t o th e v a r i e t y o f t e s t s a p p lie d t o v a rio u s compounds, many f a l s e
c o n c lu s io n s have been drawn by m ixing r e s u l t s o b ta in e d w ith d i f f e r e n t t e s t
m ethods.
T h is d is c u s s io n w i l l c o n ta in p r im a r ily th e r e s u l t s o f work
o b ta in e d by u sin g a c e l l e lo n g a tio n m ethod sin c e only by th e use of homo­
geneous m a t e r i a l can c o n c lu s io n s be s a f e ly drawn.
The Avena s t r a i g h t grow th (1 5 ,1 3 ,7 1 ,7 2 ,2 4 ) and th e pea t e s t (8 7 ,1 0 0 ,
80*18) a r e th e two p r e f e r r e d methods s in c e secondary f a c t o r s ( p e n e t r a b i l i t y ,
p o la r t r a n s p o r t , i n a c t i v a t i o n o f compounds) a re ex clu d ed t o a la r g e e x te n t ®
- 9 -?
However, th e complex n a tu r e o f t h e pea t e s t a s w e ll a s o th e r f e a tu r e s (2 4)
make i t l e s s s a t i s f a c t o r y th a n th e Avena s t r a i g h t growth m ethod,
Norman and W eintraub ( 76) have c l e a r l y o u tlin e d th e l i m i t a t i o n s o f
e x i s t i n g d a ta a s d eterm in ed by th e v a r i a t i o n s i n t e s t o b je c t s , d i f f e r e n t
e x p e rim e n ta l c o n d itio n s and te c h n iq u e s , e t c . , th u s many o f t h e a v a il a b le
d a ta p o s s e s s on ly a s e m i- q u a n tita tiv e c h a r a c te r .
I n e x p re ss in g th e r e s u l t s o f a c e r t a i n t e s t method i n a b p lu te te rm s ,
i t must be remembered t h a t th e re sp o n se t o s tim u la tio n , w hich becomes an
i n h i b i t i o n a t h ig h e r c o n c e n tr a tio n s , i s d e s c rib e d by an a c t i v i t y curve
w ith a maximum.
I t i s o b v io u s. th e n , t h a t two compounds may have t h e i r max­
imum re sp o n se a t d i f f e r e n t c o n c e n tr a tio n s ,
However, a s V e ld s tra (113) h as
in d ic a te d , r e l a t i v e a c t i v i t i e s a r e o f te n more im p o rtan t th a n a b s o lu te ones
i n stu d y in g 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 h i p s and th e r e f o r e i t i s e s s e n t i a l
t h a t t h e a c t i v i t i e s of compounds u n d er com parison be d eterm in ed sim u lta n e ­
o u sly un d er i d e n t i c a l c o n d itio n s .
S tr u c tu r e and A c tiv ity
The c o r r e l a t i o n betw een chem ical s tr u c tu r e and p h y s io lo g ic a l o r
p h a rm a c o lo g ic a l a c t i v i t y has alw ays been f a s c in a tin g b ecau se i t may have
a d ouble p u rp o se:
a b a s ic p a tte r n f o r s y n th e tic work aim ing a t new com­
pounds. o f com parable o r enhanced a c t i v i t y and a p o s s ib le in s i g h t a s to th e
mode o f a c ti o n .
However, due to th e co m p lex ity o f th e s tr u c t u r e o f th e
a c ti v e compounds and th e t e s t methods u sed i n a s s e s s in g a c t i v i t y , l i t t l e
o r no c o r r e l a t i o n can be made.
P la n t growth s u b sta n c e s , how ever, ap p ear
v e ry a t t r a c t i v e f o r t h i s k in d o f a n a ly s is because b o th t h e s tr u c tu r e s and
t e s t m ethods a r e r e l a t i v e l y sim ple *
- IG
In. 1938 K o e p fli, Thiraann and Went ( 58) . p u b lis h e d t h e T iitSt compre­
h e n siv e re p o rt, on s y n th e tic p la n t g ro w th - r e g u la to r s .■ They concluded from
t h e i r work t h a t t h e minimum- s t r u c t u r a l re q u ire m e n ts f o r grow th a c t i v i t y i n
h ig h e r p la n ts a r e :
(a ) a r in g system a s a n u c le u s , (b) a double bond i n t h i s
r i n g , (c ) a s id e c h a in , (d) a c arb o x y l group (o r a s t r u c t u r e r e a d ily con­
v e r te d to a c a rb o x y l group) on t h i s s id e c h a in a t l e a s t one carbon atom r e ­
moved from t h e r i n g , and (e) a p a r t i c u l a r space r e l a t i o n s h i p between th e
r in g and c arb o x y l g ro u p .
I n subsequent y e a r s a la r g e number o f compounds have been screen ed f o r
a c t i v i t y and from th e r e s u l t s o b ta in e d , re s ta te m e n ts o f t h e s t r u c t u r a l r e ­
q u ire m e n ts i n d i f f e r e n t form s have a p p e a re d .
S e v e ra l review s, o f th e s e
developm ents have been p u b lis h e d ( 1 1 3 , 7 4 » 1 0 1 , 8 1 , 1 0 2 ) F o r tfye purpose o f
d is c u s s io n , c e r t a i n of th e s e re q u ire m e n ts w i l l be re p e a te d h e re .
As more compounds became known, th e re q u ire m e n ts a s s t a t e d by K o e p fli
e t . a l . f e l l s h o rt o f a c c o u n tin g f o r a l l o b ta in e d r e s u l t s .
V e ld s tr a (1 0 6 ,'
lb ? ) r e s t a t e d th e re q u ire m e n ts f o r a p la n t grow th su b sta n c e a s fo llo w s:
(a ) th e s u b sta n c e should have a b a s a l r in g system w ith h ig h s u rfa c e a c t i v i t y ,
and (b) i f should have a c arb o x y l group i n a v e ry d e f i n i t e s p a t i a l p o s itio n
w ith r e s p e c t t o t h i s r i n g system .
L a te r on V e ld s tr a and B o o ii (108). fo rm u la te d t h e re q u ire m e n ts i n more
d e t a i l a s : (a ) a b a s a l r in g system (n o n p o la r p a r t) w ith h ig h in te r f a c e
a c t i v i t y and (b) a c a rb o x y l group ( p o la r p a r t ) , i n g e n e r a l a g ro u p .o f a c id i c
c h a r a c te r , i n such a s p a t i a l p o s itio n w ith r e s p e c t t o t h e r in g system , t h a t
on a b s o r p tio n of th e a c ti v e m olecule to a boundary, t h i s f u n c tio n a l group
w i l l be s itu a te d a s p e r ip h e r a lly a& p o s s i b l e .
T h e ir th e o r y i s e s s e n t i a l l y
\
- 11 -
one o f a l t e r e d p e rm e a b ility o f c e l l membranes caused by th e a d s o rp tio n o f
g ro w th - c o n tr o llin g compounds t o t h i s phase boundary.
The th e o ry was b ased
m ain ly on a p o la ro g ra p h ic i n v e s t ig a ti o n i n which th e e f f e c t s of th e com­
pounds on th e oxygen maximum w ere used a s a m easure o f t h e a b s o r b a b ili ty o f
j
th e compounds and t h e r e f o r e o f t h e i r p h y s io lo g ic a l a c t i v i t y .
P a le g and
M uir (S i) p o in t out t h a t t h i s th e o ry f a l l s f a r s h o rt o f a c c o u n tin g o f th e
a c t i v i t y o r n o n - a c tiv ity of many compounds, and co n clu d e t h a t chem ical
r e a c t i v i t y i s more im p o rta n t th a n a b s o r b a b i l i t y .
V e ld s tr a (113) f u r t h e r proposed t h a t f o r p o s i t i v e a c t i v i t y , a non­
c o p la n a r ity e x i s t betw een th e c arb o x y l group and th e r in g system .
The con­
c lu s io n r e s u l t e d from t h e s p e c ta c u la r d if f e r e n c e i n p h y s io lo g ic a l a c t i v i t y
betw een th e c i s and tra n s -c in n a m ic a c id s (106) ( th e c i s form b e in g a c t i v e ) .
Only th e t r a n s form te n d s to o ccu r i n a f l a t form (1 1 3 ).
The same r e s u l t s
have been found i n t h e case o f c i s - t r a n s t e t r a l i d e n e a c e t i c a c id and n a p h th a l e n e - l - a c r y l i c a c id (1 0 6 ).
In th e o p in io n o f t h e a u th o r, th e m ost p ro m isin g i n t e r p r e t a t i o n a s t o
t h e mode of. a c tio n o f g ro w th -re g u la to rs and th e s t r u c t u r a l re q u ire m e n ts
n e c e s sa ry f o r a c t i v i t y has' been proposed by Hansch and M uir (7 2 ,4 0 ,3 9 ,7 4 )»
They f i r s t proposed t h e " o rth o e f f e c t " (3 9 ) , s t a t i n g t h a t compounds must
have a f r e e o r p o t e n t i a l l y f r e e o rth o p o s i t i o n t o be a c t i v e in c e l l e lo n ­
g a tio n .
I n 1950, M uir and Hansch (72) l i s t e d t h e minimum s t r u c t u r a l r e ­
q u ire m e n ts f o r grow th a c t i v i t y a s : (a ) an u n s a tu ra te d r i n g (p r e fe ra b ly
a ro m a tic ), (b) a c arb o x y l group o r a group cap ab le o f c o n v ersio n t o a c a r ­
b o x y l group ( t h i s group may be a tta c h e d th ro u g h a c h ain o f one o r more
atom s o r d i r e c t l y t o t h e r i n g ) , (c) a t l e a s t one of t h e p o s itio n s on th e
-
12
-
r in g o rth o t o th e p o in t of atta ch m e n t o f th e carb o x y l group o r th e s id e c h a in
c a r ry in g th e carb o x y l group must be c a p a b le o f r e a c tio n w ith an e le c tr o n r i c h p la n t s u b s tr a te (th e o rth o p o s itio n must h o ld th e p ro p e r e le c tr o n d en s­
i t y and th e group o r atom a t t h a t p o s itio n must be cap a b le o f d isp lacem en t
under th e c o n d itio n s o f r e a c t i o n ) , (d) a c c o rd in g t o V e ld s tr a and B ooij (108)
th e r e l a t i v e ly o p h ilic c h a r a c te r of th e m o lecu le compared t o th e
h y d ro p h ilic
n a tu re o f c e r t a i n a tta c h e d groups such a s th e carb o x y l group may e f f e c t th e
a c t i v i t y o f a compound a s a g ro w th - re g u la to r , and (e ) c h a r a c t e r i s t i c s such
a s th e r e l a t i o n o f th e s id e c h a in c arb o x y l group to an o rth o p o s itio n may be
o f im portance in d e te rm in in g th e a c t i v i t y o f a compound, b u t l i t t l e i s known
about them .
In 1951 H ansch, M uir and M etzenberg (40) p o s tu la te d th e tw o -p o in t sub­
s t r a t e r e a c tio n :
Cl
CHCO-Protein
Cl
OCHgCOOHjN'CHCO-Protein
H sia2
,/M g
Cl
OCH2COOHtN"
XXV q
^C H C O -Protein
^crC2
Cl
Cir^
+M,COOHXN
N lI ^ c 3^ H C O - P r o te in
^CH0
Cl
O-CH/
Cl
C=O
>NH
CHCO-Protein
- 13 -
T h is mode o f a c ti o n i s su p p o rte d by c o n s id e ra b le ev id en ce w hich w i l l be
g iv e n i n d e t a i l i n th e d is c u s s io n o f th e p h en o x y acetic a c id s .
S in ce s e v e r a l re q u ire m e n ts fo rm u lated f o r a c t i v i t y p e r t a i n t o a s p e c ia l
group o f grow th s u b s ta n c e s , th e y w i l l be d is c u s s e d w ith th e r e s p e c tiv e ty p e s
o f compounds.
The R ing System
The p re se n c e o f a r in g system a p p e a rs to be an u n q u e stio n ed r e q u ir e ­
ment f o r a c t i v i t y .
Up t o th e p re s e n t tim e , no a c ti v e compound has been
found la c k in g a r in g n u c le u s .
A lso , up to now no a c tiv e compound has been
found which la c k s a t l e a s t one double bond w ith in th e r in g ; when only one
i s p r e s e n t (a s i n cyclohexene) a p o s itio n o f th e s id e c h a in n e x t to th e
d o uble bond seems t o be n e c e s s a ry f o r a c t i v i t y (1 2 1 ).
V e ld s tr a (113) c o n s id e rs th e a re a o f th e r i n g system a s b e in g im p o rtan t
and l i s t s th e n a p h th a le n e o r in d o le n u c le u s a s b e in g m ost fa v o ra b le i n s i z e .
He s u g g e s ts t h a t t h i s s iz e i s e q u a lle d t o some e x te n t by c e r t a i n s u b s tit u te d
benzene n u c l e i , e .g . 2 ,4 - d ic h lp ro p h e n o l.
The i n a c t i v i t y o f such compounds
a s 2 , 6 -d ic h lo ro p h e n o x y a c e tic a c id (96 ,1 2 9 ) and 2 -flu o ro -4 '-m e th y Iphenoxya c e t i c a c id '(119) a s compared to th e v e ry h ig h a c t i v i t i e s o f 2 ,4 - d ic h lo r o p h e n o x y ac e tic a c id and 2-m ethy1 -4 -flu o ro p h e n o x y a c e tic a c id ( 119) would seem
t o in d ic a te t h a t t h e a re a o f th e r in g system co u ld a t m ost be o f m inor
im p o rtan ce i n th e s tr u c t u r e o f a c ti v e compounds.
The P o la r Group i n th e S id e Chain
The p re s e n c e o f a c a rb o x y l group i n th e s id e c h ain a p p e a rs t o be
e s s e n tia l fo r a c tiv ity .
Replacem ent o f t h e carb o x y l group by SO3H g e n e r a lly
r e s u l t s i n i n a c t i v i t y o r a t th e most o n ly s l i g h t a c t i v i t y .
2- ( !-N a p h th a le n e )
— 14 —
-e th a n e s u lp h o n ic a c id ( 106) , and ! - n a p h th a le n e - s u lf u r ic a c id (113) have
been shown t o be i n a c t i v e .
Indole-r3 -m eth an esu lp h o n ic a c id (1 1 3 )^a rid 'i h d o l y l -
s u lp h u ric a c id (106) show s l i g h t a c t i v i t y ; th e a c t i v i t y o f th e form er has
b een q u e stio n e d however (1 2 2 ).
Sodium 2 -( 2 ,4 -d ic h lo ro p h e n o x y )-e th y l s u l f a t e
( 114, 4 7 , 57) i s th e o n ly n o n -c a rb o x y lic compound, which h as an a c t i v i t y com­
p a ra b le to t h a t o f c a rb o x y lic compounds.
The S id e Chain
T hat th e c a rb o x y l group must be s e p a ra te d from t h e r i n g system by a t
l e a s t one carbon atom ( K o e p fli e t . a l . ) , can no lo n g e r be h e ld a s a ^ s t r u c t u r a l '
re q u ire m e n t.
The abandonment o f t h i s re q u ire m e n t r e s u lte d when th e high
a c t i v i t y o f 2 ,3 ,6 - tr ic h l o r o b e n z o ic a c id (13) became known.
S in ce t h i s
i n i t i a l d is c o v e ry , many s u b s t i t u t e d b e n zo ic and n a p h th o ic a c id s have been
shown t o be a c t i v e .
Synerholm and Zimmerman ( 96) observ ed an o s c i l l a t i o n i n th e a c t i v i t y
o f tt) -(2 ,4 -d ic h lo ro p h e n o x y )-a lk y lc a rb o x y lic a c id s and W-(^-n ap h th o x y ) - a l k y l c a rb o x y lic a c id s ; th e a c e t i c and b u ty r i c a c id s were a c t i v e , th e p ro p io n ic
a c id i n a c t i v e .
These a u th o rs su g g ested t h a t th e h ig h e r homologues would
n o t be a c t i v e p e r se b u t, b ein g s u b je c t t o a ^ - o x i d a ti o n , would produce th e
a c ti v e 2 , 4 - d i c h lo ro p h enoxyac e t i c a c id o n ly when p o s s e s s in g an even number o f
carbon atom s i n th e s id e c h a in .
T h is re a s o n in g has been s u p p o rte d by
Faw cett e t . a l . (2 9 ) , who found ev id en ce o f /6 -o x id a tio n i n th e s e r i e s o f
f
W -p hen o x y alk y lcarb o x y lic a c id s i n f l a x p l a n t s .
S u b s titu ti o n o f a m eth y l group i n th e ^ - p o s i t i o n o f a p h en o x y acetic
a c id h a s l i t t l e e f f e c t , w h ile a p heny l group o r d i-m e th y l s u b s t i t u t i o n
g r e a t l y d e c re a s e s a c t i v i t y .
Wain e t . a l , (8 9 ,8 8 ) c o n s id e r t h e a lp h a hydrogen
. - 15 atom a s b e in g e s s e n t i a l t o a c t i v i t y .
Due t o th e a c t i v i t y o f th e b en zo ic
and n a p h th o ic a c id s , t h i s a u th o r p r e f e r s t o c o n s id e r th e &-s u b s t i t u t i o n
e ffe c t as a s te r ic fa c to r.
P h e n y la c e tic A cid and R e la te d Compounds
P h e n y la c e tic a c id h as a r e l a t i v e l y h ig h a c t i v i t y a s compared t o th e
n e a r ly in a c t i v e p h en o x y ac e tic a q id ,
D i-a lp h a s u b s t i t u t i o n i n th e s id e c h a in
c o m p le tely i n a c t i v a t e s the. compound ( l ) .
T h is d e f i n i t e l y seems to be a case
o f s t e r i c e f f e c t s a s seen from th e fo llo w in g ev id en ce: d l oi-n-propylp h e n y la c e t i c a c id and d l a - a l l y lp h en y ! a c e t i c a c id a re more a c ti v e th a n th e p a re n t
pheny! a c e t i c a c id , w h ile th e branched c h a in compound, d l oti-is o p ropyIp h eny1 a c e t i c a c id , i s alm ost in a c t iv e (1 0 9 ).
•
'
•
Thimann (102) concluded t h a t t h e in tr o d u c tio n o f a n i t r o group in to
th e r in g o f phenyl a c e t i c a c id a b o lis h e s a c t i v i t y sin c e he found b o th th e 4 n i t r o - and 2 ,4 - d i n i t r o - d e r iv a tiv e s t o be i n a c t i v e .
He proposed t h a t t h i s
s u b s t i t u e n t , b ein g .h ig h ly d e a c tiv a tin g to th e benzene r i n g , low ers th e
b in d in g c a p a c ity o f th e m o le c u le .
V e ld s tra (113) h a s , how ever, found th e
2 - n i t r o - and 3 - n i t r o - d e r i v a t i v e s to have an a c t i v i t y e q u a l t o o r g r e a te r
th a n th e p a re n t compound, and s t a t e d t h a t Thimann1s g e n e r a liz a tio n was no t
ju s tifie d .
The -T e f f e c t o f th e n i t r o group o f f s e t s t h e +T e f f e c t o f th e
carboxym ethoxy group i n th e o rd e r o f p a ra > o rth o rn e ta ; t h i s i s i n re fe re n c e
t o th e proposed bonding a t an o rth o p o s itio n to th e c a rb o x y lic s id e c h a in .
T h is o rd e r i s t h e same a s t h a t o bserv ed f o r th e a c t i v i t i e s o f th e th r e e
i s o n e rs .
T h is r e l a t i o n s h i p o f p o s itio n to a c t i v i t y i s s im ila r f o r th e b en zo ic
and p h e n o x y ac e tic a c id s and w i l l be d is c u s s e d l a t e r i n more d e t a i l .
'T
— 16 —
As s t a t e d p r e v io u s ly , o n ly c is-c in n a m ic a c id (106) p o s s e s s e s a c t i v i t y .
N ivard (113) in v e s t ig a te d s e v e r a l s u b s t i t u t e d c is-c in n a m ic a c id s .
I t was
found t h a t th e 2 and 4 -c h lo ro compounds had an enhanced a c t i v i t y , th e 4 ch lo ro cin n am ic a c id b e in g th e more a c t i v e .
Both 2 - and 4 -n itro c in n a m ic a c id
w ere i n a c t i v e .
Ci s -l-p h e n y Ic y c lo p r o p a n e-2 -c a rb oxyI i c a c id was found t o be a c tiv e and
l i k e th e cinnam ic a c id s , th e t r a n s isom er was in a c t iv e ( il l) * .
I n d o le - 3 - a lk y lc a r b o x y lic A cids
S u b s titu ti o n i n th e p y r r o le n u c le u s s tro n g ly re d u c es a c t i v i t y inin d o le - 3 - a c e ti c a c id (4 8 ,9 1 # 9 2 ).
2 i s e s s e n tia l fo r a c tiv ity .
T h is would in d ic a te t h a t a f r e e p o s itio n
S u b s titu tio n in th e benzene r in g o f th e in d o le
n u c le u s d e c re a s e s bply s l i g h t l y and may even enhance a c t i v i t y .
The 4 -,5 - #
and 6 - c h lo r in d o le - 3 - a c e ti c a c id s (48) have a c t i v i t i e s e q u a l t o o r g r e a te r
th a n th e p a re n t a c id .
S u b s titu ti o n i n p o s itio n 7, however, g e n e r a lly
c au ses a d e c re a se i n a c t i v i t y , p a r t i c u l a r l y i f th e in d o le - 3 - a c e ti c a c id i s
a lre a d y s u b s t i t u t e d e lse w h e re .
The a p p a re n t im portance o f p o s itio n s 2 and 7 (a d ja c e n t o r o rth o t o th e
NH group) r e c a l l s th e im p o rtan ce imposed upon th e o rth o p o s itio n i n th e
p h en o x y ac e tic a c id s e r i e s (Hansch and M uir (3 9 ))•
K o e p fli e t . a l . (58) s tu d ie d t h e e f f e c t o f s u b s t i t u t i o n i n d - in d o l e - 3 p ro p io n ie a c id and yt-indoI e -3 - b u ty r ic a c i d .
t o have h ig h a c t i v i t i e s i n th e pea t e s t .
These p a re n t a c id s were found
The 2-carb o x y d e r iv a tiv e s and th e
5 - , 6 - , and 7-m ethoxy d e r iv a tiv e s of t h e p ro p io n ic a c id were a l l i n a c t i v e .
As m entioned above, t h i s i n a c t i v a t i o n does n o t o ccu r w ith in d o le - 3 - a c e tic
a c id ; 5 -# 6 -, and 7 -n ie th o x y in d o le -3 -a c e tic a c id s a r e q u ite a s a c ti v e as th e
- 17 -
p a re n t a c id (3 1 ).
Phenoxy Compounds
The p r a c t i c a l im p o rtan ce o f 2 ,4 -d ic h lo rp p h e n o x y a c e tic a c id (2,4-D ) h as
caused e x te n s iv e i n v e s t ig a ti o n s on th e e f f e c t s o f n u c le a r s u b s t i t u t i o n i n
p h e n o x y ac e tic a c id .
N u clear s u b s t i t u t i o n s tu d ie s o f some o f th e h ig h e r
homologues have been co n d u cted , b u t t o a l e s s e r d e g re e .
Most o f th e
c o r r e l a t i o n s betw een p h y s io lo g ic a l a c t i v i t y and ch em ical s t r u c t u r e have
r e s u l t e d from th e s e s tu d ie s o f th e p h en o x y acetic a c id compounds.
As w i l l be
seen l a t e r , th e s u b s t i t u t e d b en zo ic a c id s have a ls o re c e iv e d c o n sid e ra b le
a tte n tio n .
Bonner (15) su g g este d t h a t th e r e l a t i v e grow th prom oting a c t i v i t i e s o f
th e s u b s tit u te d p h e n o x y ac e tic a c id s m ight be accounted f o r by th e d i f f e r ­
en ces i n t h e d eg ree o f io n iz a tio n of t h e a c id s .
However, M uir e t . a l . (71)
and Hayes and Branch (41) have shown t h a t no such r e l a t i o n s h i p o c c u rs.
L eaper and B ishop ( 64) i n 1951 p re p a re d a l l of th e m ono-, d i - , and t r i c h lo ro p h e n o x y a c e tic a c id s and t e s t e d t h e i r e f f e c t on tom ato p la n ts and a s
i n h i b i t o r s o f grow th o f Lupinus a lb u s s e e d lin g s .
M uir e t . a l , (71,73>72)
and Thimann (102) o b ta in e d com parative a c t i v i t i e s f o r a number o f th e com­
p o u n ds.
Leaper and B ishop ( 64) summarized t h e i r r e s u l t s i n a t a b l e (T able
I ) in which th e .m o s t a c ti v e compound ( i n c e l l e lo n g a tio n ) was r a te d a s 100,
B ecause o f t h e i n a c t i v i t y o f th e 3 ,5 - , 2 , 3 , 5 - , and 3 ,4 ,5 - a c id s i n th e s e
t e s t s th e y proposed t h a t an open p o s itio n p a ra t o an open o rth o p o s itio n i s
r e q u ir e d f o r a c t i v i t y and th e a c ti v e m o lecu le i s in v o lv e d i n th e fo rm a tio n
o f q u in o id compounds i n p la n t c e l l s .
Wain and Wightman, how ever, examined
th e s e same compounds f o r a c t i v i t y i n th e s t r a i g h t growth o f AvenA c o le o p tile s ,
. 18 —
TABLE I
R e la tiv e A c t i v i t i e s of C hloropherioxyacetic A cids in Tomato T e s t ( 64)
P o s itio n o f c h lo rin e
g ro ups i n n u c leu s
A c t i v i t y in ro o tin g
i n t a c t stems
2
2 .0
1 0 .0
3
1 0 .0
1 2 .5
4
10*0
2 5 .0
2 ,3
1 .0
5 .0
2 ,4
66*6
77 .5
2 »5
1 0 0.0
1 00.0
■2 ,6
0 .0
0 .0
3 ,4
2 0 .0
1 0 .0
3*5
0 .0
0 .0
2 ,3 ,4
1 .0
0 .0
2 ,3 ,5
0 .0
QfaO
2 , 3 ,6
0 .0
0 .0
4 0 .0
2 5 0 .0
2 ,4 ,6
0 .0
0 .0
3 ,4 ,5
0 .0
0 .0
2 ,4 ,5 '
and in t h e pea t e s t (1 1 7 ).
2 ,3 ,5 -
A c tiv ity i n c e l l
e lo n g a tio n
( e p in a s ty , e t c , )
They found i n b o th t e s t s t h a t th e 2 , 3 - , 2 ,3 ,4 ’-,
and 3 ,4 ,5 - a c id s w ere a c t i v e .
I t i s seen t h a t th e s e compounds do n o t
comply w ith th e h y p o th e s is of' L eaper and B ishop and y e t a r e a c t i v e .
Osborne and Wain (7 7 ,7 8 ,7 9 ) s tu d ie d a s e r i e s o f a ry lo x y a c e tic a c id s
i n d i f f e r e n t t e s t s (in c lu d in g s t r a i g h t grow th and th e pea t e s t ) and found
t h a t m ono-alpha s u b s t i t u t i o n i n th e s id e c h a in had l i t t l e e f f e c t upon
— 19 —
a c tiv ity .
The « ,tf ~ d is u b s titu te d compounds ( i . e . ary l-0 C C (CHj^COOH) were
in a c t i v e i n th e s t r a i g h t growth t e s t and showed b u t s l i g h t a c t i v i t y in th e
pea t e s t .
Due t o th e i n a c t i v i t y o f th e <t5fi& -d isu b stitu ted d e r i v a t i v e S5 Wain
and cow orkprs proposed th e h y p o th e sis t h a t th e r e i s a ch em ical r e a c tio n i n ­
v o lv in g a hydrogen atom in th e a lp h a p o s i t i o n , and t h a t a t l e a s t one such
hydrogen atom i s n e c e s s a ry f o r a c t i v i t y .
The a c t i v i t y of a -m e th y le n e p h e n y !a c e tic a c id and th e h ig h a c t i v i t y o f
2 ,3 # 6 - tr ic h l o r b e n z o ic a c id would in d i c a t e t h a t t h i s a lp h a h y p o th e s is i s
in v a lid .
As m entioned p re v io u s ly , th e i n a c t i v i t y o f t h e e - d i s u b s t i t u t e d
a c id s could be due c o n c e iv a b ly to s t e r i c h in d ra n c e , which p re v e n ts th e m ole­
c u le from f i t t i n g i n t o a r e q u i s i t e p a t t e r n n e c e s sa ry f o r a c t i v i t y .
As p re v io u s ly m entioned, M uir and Hansch (71) proposed a tw o -p o in t
r e a c tio n th e o r y .
T h is c o n s is te d o f an am ide lin k a g e w ith th e carb o x y l
group and a n u c le o p h ilic a tta c k on an o rth o p o s itio n by a s u lf h y d r y l g ro u p .
The r e a c t i v e s u b s tr a te u n it b e in g a f r e e c y s te in e g ro u p .
th e o ry has c o n s id e ra b le ev id en ce i n i t s f a v o r .
T h is tw o -p o in t
F o s te r , McRae and Bonner
(33) have shown by p. stu d y of enzyme k i n e t i c s t h a t i n h i b i t i o n o f growth by
g ro w th - re g u la to r s a^ h ig h e r c o n c e n tra tio n s would be e x p e c te d i f th e sub­
s ta n c e r e a c t s by a tw o -p o in t a ttach m en t mechanism.
The a c t i v e complex
would have a g ro w th -re g u la to r m olecule a tta c h e d a t two p o in ts on th e sub­
s t r a t e ; a s th e c o n c e n tra tio n o f r e g u la to r in c r e a s e s , th e p r o b a b ility o f two
m o le c u le s a tta c h in g a t th e two s u b s tr a te s i t e s in c r e a s e s and th e number o f
tw o -p o in t a tta c h m e n ts o f one g ro w th -re g u la to r m o lecu le form ing t h e a c tiv e
complex p r o g r e s s iv e ly d im in is h e s .
Thimann and Bonner (104,105) observ ed t h a t io d o a c e ta te , a rs e n a te and
—
20
—
p -c h lo ro m e rc u rib e n z o a te (which r e a c t w ith s u lfh y d ry l groups) i n h i b i t th e
e lo n g a tio n of Avena s e c tio n s .
T h is may b e i n t e r p r e t e d . a s i n d ic a tin g some
ty p e of r e a c t i o n betw een s u lf h y d r y l enzymes and t h e grow th—re g u la to r .
Bonner ( I ? ) a ls o r e p o rte d an i n h i b i t i o n i n th e grow th o f Avena c o le o p tile
s e c tio n s by L - c y s te in e , i n d ic a tin g a p r e f e r e n t i a l r e a c tio n .
The f a c t t h a t
c y s te in e i s used f o r d e t o x i f i c a t i o n o f bromo—# c h lo ro — and io d o b en zene and
o f n a p h th a le n e i n a n im als would in d ic a te t h a t th e h y p o th e s is f o r fo rm a tio n
o f th e t h i o e th e r i s n o t u n re a s o n a b le .
Brbmobenzene i s c o n v erted t o p-rbro-
mopheny1-m ere a p tu r i c a c id (90) and n a p h th a le n e t o p -n a p h th y lm e rc a p tu ric a c id
(1 9 ,5 3 ) .
'I n b o th c a s e s , th e n t h e amino group has been a c e ty la te d ; t h i s
f u r t h e r s u p p o rts th e tw o -p o in t atta ch m e n t th e o r y .
As M uir and Hansch (71) p o in t o u t, t h i s h y p o th e sis e x p la in s v e ry s a t i s ­
f a c t o r i l y t h e a c t i v i t y o f c i s —cinnam ic a c id i n e lo n g a tio n and th e i n a c t i v i t y
o f th e t r a n s form a s w e ll a s o th e r c i s - t r a n s a c id s of t h i s ty p e , s in c e th e ■
tw o -p o in t a tta c h m e n t r e s u l t s i n a r in g s t r u c t u r e .
The e a se o f r in g forma­
t i o n i n a c i s - a c i d o v er t h a t o f t h e tra n s -is o m e r i s o b v io u s.
The i n t e r p r e t a t i o n s o f t h e tw o -p o in t a ttach m en t th e o r y do n o t acco u n t
f o r a l l o f t h e o bserved r e s u l t s and th e p ro p o s e rs have n o t a tte m p te d to
e x p la in t h e extrem e s p e c i f i c i t y o f c e r t a i n s u b s tit u e n ts and p o s itio n s .
The
a u th o r cannot a g re e w ith t h e p ro p o s a l o f a n u c le o p h ilic a t t a c k on th e benzene
rin g .
o c c u r.
N u c le o p h ilic d isp la c e m e n ts o f a ro m a tic hydrogen do n o t o r d in a r il y
Benzene i t s e l f i s fu n d a m e n tally n u c le o p h ilic and th e f i n a l e j e c t i o n
o f a hydrogen a n io n i s u n l i k e l y .
N u c le o p h ilic s u b s t i t u t i o n a t an a ro m a tic
carbon should be f a c i l i t a t e d , b y i
(a ) any in flu e n c e which in c r e a s e s th e n o r­
m a lly u n re a c tiv e c h a r a c te r of th e a ro m a tic n u c le u s , and (b) by s u b s t i t u t i n g
—
21
—
f o r th e .h y d ro g e n which i s d is p la c e d a s a n e g a tiv e io n , some atom o r group
w hich more r e a d i l y e x i s t s a s a n e g a tiv e io n .
I n o th e r w ords, a p o w erfu l
m eta o r ie n tin g group sh o u ld f a c i l i t a t e t h e SWg ( b im o lec u la r n u c le o p h ilic
s u b s tit u tio n ) ty p e d isp la c e m e n t o f any gro u p capable o f e x i s t i n g a s an a n io n
when t h i s second group i s lo c a te d o rth o o r p a ra to th e m eta o r ie n tin g g ro u p »
The fo llo w in g exam ples a r e i l l u s t r a t i v e .
The h y d ro ly s is o f ch lo ro b en zen e t o
p h en o l i s o n ly accom plished a t e le v a te d te m p e ra tu re s and p r e s s u r e s ; th e Dow
Chem ical Co. p re p a re s p h en o l com m ercially i n t h i s manner (38) and u ses 6t 8%
aqueous NaOH a t 300° C and 2000-3000 p . s . i . p r e s s u r e .
In d is tin c t c o n tra s t,
o - and p -n itro c h lo ro b e n z e n e s a re r e a d ily h y d ro ly zed by a q u eo u s. NagGO^ a t
C
130° C and a tm o sp h eric p r e s s u r e (124 )•
T h is l a b e l iz in g e f f e c t , a s would be
e x p e c te d , i s n o t observed w ith m -n itro c h lo ro b e n z e n e .
Thus, a n u c le o p h ilic
d isp la ce m e n t o f a hydrogen atom o rth o o r p a ra t o . a s tro n g o rth o -o a ra d i r e c t ­
in g group (su ch a s th e carboxym ethoxy group) i s v e ry im p ro b ab le i f n o t im­
p o s s ib le .
The mechanism p roposed f o r th e s u b s tit u te d b e n zo ic a c id s , a s w i l l
be seen l a t e r , fo llo w s t h e p r e r e q u i s i t e s f o r a ro m a tic n u c le o p h ilic d is p la c e ­
ment .
I f th e r in g c lo s u r e a t th e o rth o p o s i t i o n i s n o t accom plished by an
a t t a c k of th e n u c le o p h ilic SH group, i t seems l o g i c a l t o p o s tu la te th e
i n te M e d ia te fo rm a tio n o f a sulfonium o r carbonium io n ( th e o n ly evidence o f
S-C bond fo rm a tio n i s th e d e t o x i f i c a t i o n r e a c tio n i n a n im a ls) and an e l e c t r o ­
p h ilic a tta c k .
From an e le c tr o n i c view p o in t, th e carbonium io n fo rm atio n
would be th e l o g i c a l c h o ic e , g iv in g r i s e t o a r e a c tio n s im ila r t o a F r ie d e lC ra ft r e a c t i o n .
T h is h y p o th e s is would m eet a l l o f t h e re q u ire m e n ts f o r
a tta c h m e n t to a p o s itio n o rth o to th e carboxymethoxy g ro u p .
T here a re th r e e
— 22 —
f a c t o r s w hich seem to be o f a n obvious im p o rtan ce i n d e te rm in in g an a c tiv e
s tru c tu re :
(a ) th e p o s itio n o f t h e a ro m a tic s u b s t i t u e n t , (b) th e e le c tr o n i c
e f f e c t o f t h i s su b stitU O n t r e l a t i v e t o f u r t h e r s u b s t i t u t i o n and (c) th e s iz e
o f th e s u b s t i t u e n t .
F a c to rs (a ) and (b) a r e c lo s e ly t i e d to g e th e r , i . e . b e ­
f o r e a s u b s tit u e n t can e x e r t an in f lu e n c e , i t must be lo c a te d i n a s u ita b le
p o s itio n .
The s iz e f a c t o r would e x p la in why only th e h a lo g en s and th e .
m eth y l group a re p a r t i c u l a r l y e f f e c t i v e s u b s t i t u e n t s .
A com parison o f
atom ic r a d i i shows th e m ethyl group t o be alm o st th e same s iz e a s th e io d in e
atom ,
A lthpugh th e hydroxy group and t h e amino group have a s u ita b le s iz e ,
th e y would be excluded by f a c t o r s (at) and ( b ) ,
Both o f th e s e groups have a
■ stro n g e r+T e f f e c t th a n th e OCHgGOOH group and w ould, when lo c a te d o rth o
o r p a ra to th e s id e c h a in , cause s u b s t i t u t i o n ,o r t h o o r p a ra to th em selv es
(upon c o n s tr u c tio n o f H ir s c h f e ld e r m o le c u la r m odels, i t can be seen t h a t
th e o nly p o s itio n s c a p a b le o f form ing a r in g w ith th e c y s te in e re s id u e a re
th e p o s itio n s o rth o to th e s id e c h a in .
The m eta p o s itio n s w i l l form r in g
s t r u c t u r e s , b u t o n ly w ith c o n s id e ra b le s t r a i n ) .
I f th e OH o r NHo group i s
lo c a te d i n th e m eta p o s itio n o f p h en o x y ac e tic a c id , i t would be p a ra to
th e 2 p o s i t i o n .
The 2 and 4 p o s itio n s a r e th e n most p ro n e t o e l e c t r o p h i l i c
a t t a c k , p ro b a b ly i n t h e o rd e r o f 2 g r e a te r th a n 4 .
T h is re a s o n in g would
p r e d i c t m eta-hy droxyphenoxyacetic a c id t o be h i g h ly a c t i v e , b u t ex p erim en t­
a l ev id en ce shows i t to be i n a c t i v e .
However, i t must be remembered t h a t
th e a tta c k in g c y s te in e u n it i s most p ro b a b ly a tta c h e d t o a p r o te in re s id u e
th ro u g h i t s c a rb o x y l group and a tta c k by such a b u lk y m o lecu le would be
h in d e re d t o a much g r e a t e r e x te n t by th e OCHgCOOH group a s compared to th e
- 23 -
r e l a t i v e l y sm all hydroxy o r amino g ro u p .
The im p o rtan ce o f s iz e i s e v id e n t
i n th e s u b s t i t u t i o n o f a l k y l benzenes (6 5 ).
As th e a l k y l group o f th e
a lk y l benzene in c r e a s e s i n s iz e from m e th y l t o t e r t - b u t y l , p a ra s u b s t i t u t i o n
becomes in c r e a s in g ly fa v o re d .
S im ila r ly , a s th e s iz e o f th e a tta c k in g r e ­
ag en t becomes l a r g e r , p a ra s u b s t i t u t i o n should be fa v o re d .
T h is h in d ra n c e
o f and by th e a tta c k in g m o lecu le may w e ll e x p la in why th e p o s tu la te d f a c t o r
( c) i s o f a param ount im p o rta n c e .
I f t h e +T e f f e c t o f a m eta s u b s titu e n t i s
o n ly s l i g h t l y g r e a t e r th a n t h a t o f th e OCH2COOH group and t h e s iz e g r e a te r
th a n t h a t of t h e OH o r HHg, some a c t i v i t y would be e x p e c te d .
The methoxy
group f i t s b o th re q u ire m e n ts and has been r e p o rte d t o be a c ti v e when i n th e
m eta p o s itio n ( 119) .
When a s u b s tit u e n t i s s tr o n g ly e le c tr o n a t t r a c t i n g (by p e ra an e n t
p o la r iz a t io n ) th e n e a r-b y o rth o p o s itio n s sh o u ld be more s tro n g ly , d e a c t i ­
v a te d th a n th e more d i s t a n t p a ra p o s itio n and t h e o rth o - p a r a r a t i o should
d e crease (3 ).
T h is e l e c t r o n i c e f f e c t i s a p p a re n tly more im p o rtan t th a n
s l i g h t s iz e in c r e a s e s , s in c e t h e n i t r a t i o n of th e halo b en zen e s e r i e s th e
s iz e o f th e
s u b s tit u e n t in c r e a s e s from flu o ro b en y en e t o io d o b eh zen e, b u t
o rth o n i t r a t i o n becomes in c r e a s in g ly more pronounced i n d escen d in g th e
s e rie s .
I t i s w e ll known t h a t th e e l e c t r o n - a t t r a c t i n g power o f th e h a lo ­
gens a ls o d e c re a s e s i n t h e same o r d e r .
T h is could e x p la in why p h e n y la c e tic .
a c id i s h ig h ly a c ti v e and p h e n o x y ac e tic a c id i s v i r t u a l l y i n a c t i v e .
Two
compounds which would be anologous i n t h e i r s u b s t i t u t i o n r e a c tio n s a re
p h en o l ( t o p h en o x y ac e tic a c id ) and to lu e n e ( t o p h e n y la c e tic a c i d ) .
W ith an
e n te r in g s u b s t i t u e n t , p h en o l d i r e c t s p re d o m in a n tly to t h e p a ra p o s itio n ,
w h ile toluene"m ay -have"an 6 r th o - p a r a . r a t i o g r e a te r th a n l ; 0 ( 5 0 ) « p : ' Thus, i f
I
—
24 "
p h e n o x y ac e tic a c id i s p r e f e r e n t i a l l y a tta c k e d i n th e p a ra p o s i t i o n , i t co u ld
1
a c t a t b e s t a s an a n tia u x in . E i th e r p a ra s u b s t i t u t i o n ta k e s p la c e and no
r i n g c lo s u r e by th e amide lin k a g e o r p r e f e r a b ly , th e amide lin k a g e (o r th e
in te rm e d ia te s a l t lin k a g e ) i s form ed and no r in g c lo s u re fo llo w s due to th e
d ir e c te d p a ra s u b s t i t u t i o n .
The d if f e r e n c e s i n th e a c t i v i t y o f th e th r e e c h lo ro p h e n o x y ac e tic a c id s
. co u ld be accounted f o r by th e above c o n s id e r a tio n s .
The c h lo rin e atom i s
o r th o - p a r a d i r e c t i n g , b u t due to j th e r e l a t i v e l y h ig h i-I e f f e c t , th e r in g i s
l e s s e a s i l y s u b s tit u te d th a n b e n z e n e .• The r e l a t i v e r a t e s o f n i t r a t i o n
(b enzens = 1 .0 ) o f chlorobenzene; i n t h e o rth o and. p a ra p o s itio n s a r e 0 .0 3 0
and 0 .1 3 9 r e s p e c t i v e l y . . So t h e p r e d ic te d o rd e r of a c t i v i t y would be p a ra >
m e ta > o rth o .
T his i s t h e observed o rd e r o b ta in e d by many i n v e s t i g a t o r s .
The e f f e c t of t h e o rth o c h lo rin e atom on f u r t h e r s u b s t i t u t i o n may e x p la in
why o -c h lo ro p h e rio x y a c e tic a c id i s a c ti v e ( th e a c t i v i t y i s low b u t s i g n i f i ­
c a n t) and -phenoxyacetic a c id i s i n a c t i v e .
The a ce ty lam in o group has n e a r ly
t h e same e l e c t r o n i c e f f e c t s a s th e OOHgOOOH group and t h e fo llo w in g d a ta
w ould a p p ly w ith o n ly m inor d if f e r e n c e s t o p h e n o x y ac e tic and o -c h lo ro p h etid x y acetic a c id :
(a ) a c e t a n i l i d e y i e l d s on n i t r a t i o n 96$ p a ra s u b s t i t u ­
t i o n (4 9 ), (b) o r th o - c h lo r o a c e ta n ilid e on n i t r a t i o n y i e l d s o n ly 59$ p ara
s u b s t i t u t i o n (2 7 ) , th e in c r e a s e i n o rth o s u b s t i t u t i o n b e in g n e a r ly 10 f o l d .
•It i s i n t e r e s t i n g to n o te t h a t i n th e s u b s t i t u t i o n o f th e t h r e e c h lo ro a c e ta n i l i d e . is o m e rs , t h e amount o f o rth o s u b s t i t u t i o n in c r e a s e s i n th e o rd e r o f
p a ra > m eta > o rth o (26) .
A nother f a c t o r which p ro b a b ly a id s t h e o rth o sub­
s t i t u t i o n of o -c h lo ro p h e n o x y a c e tic a c id would be th e r e s u l t a n t n o n -co p lan ­
a r i t y o f th e OCHgCOOH g ro u p .
T h is p a r t i a l r e s t r i c t i o n o f r o t a t i o n may
- 25 -
d e c re a s e th e h in d ra n c e e n co u n tered by th e e l e c t r o p h i l i c a g e n t, and m ust in
tu r n be o f g r e a te r im p o rtan ce th a n t h e e f f e c t on re so n an c e due t o non­
c o p la n a r ity »
■The p re se n c e o f a m eta d i r e c t i n g group i n t h e benzene r in g g r e a tly
i n h i b i t s e l e c t r o p h i l i c s u b s t i t u t i o n , e s p e c ia ll y when th e a tta c k in g group i s
la rg e .
The a tta c k by th e p o s tu la te d p la n t s u b s tr a te i s com parable t o a lk y ­
l a t i o n by t h e F r i e d e l - C r a f t s p ro c e d u re .
I n both c a se s th e s iz e o f th e a t ­
ta c k in g group i s r e l a t i v e l y la r g e and s t e r i c f a c t o r s become o f paramount
im p o rta n c e .
One c h a r a c t e r i s t i c f e a tu r e o f a lk y la tio n by th e
F r ie d e l- C r a f ts
p ro c e d u re i s t h a t m eta d i r e c t i n g g ro u p s, w ith few e x c e p tio n s , p re v e n t r e ­
a c ti o n c o m p le te ly .
T h is could th e n e x p la in th e i n a c t i v i t y o f a l l compounds
which c o n ta in a m e ta - d ir e c tin g g ro u p .
The e f f e c t o f two or more s u b s tit u e n ts i n phenoxyac e t i c a c id would be
d i f f i c u l t t o p r e d ic t w ith o u t s u p p o rtin g e x p e rim e n ta l e v id e n c e .
However, th e
high a c t i v i t y o f 2 ,4 - 0 and th e i n a c t i v i t y o f 2 , 6 - d ic h lo r ophenoxyac e t i c a c id
a r e obvious from t h e above c o n s id e r a tio n s .
The a c t i v i t y o f 3 > 4 -d ic h lo ro -
p h e n o x y ac e tic a c id can be acco u n ted f o r by th e a d d itiv e e f f e c t s o f a p a ra and
m eta c h lo r in e .
2 ,3 - and 3#5 -d ic h lo ro p h e n o x y a c e tic a c id would d i r e c t p rim ar
is-ily to" th e p a ra p o s itio n and due t o m u ltip le c h lo rin e s u b s t i t u t i o n , may have
l i t t l e d i r e c t i o n to th e o rth o p o s i t i o n .
The high a c t i v i t y o f 2 , 5 -d ic h lo ro ­
p h en o x y ac e tic a c id would be d i f f i c u l t t o e x p la in , u n le s s f o r seme re a so n i t
d i r e c t s p r im a r ily to th e o rth o p o s i t i o n .
T here a re many o th e r compounds, whose a c t i v i t y o r i n a c t i v i t y can be
w e ll e x p la in e d by use o f e le c tr o n i c th e o r y , bu t th e a u th o r m ain ly w ish es to
point Out the strong p o s s ib ility that the two-point attachment i s in part
•* -
■
- 26
an e l e c t r o p h i l i c a t t a c k , th e p r o b a b ility o f o c cu rren ce b e in g d eterm in ed by
th e e le c tr o n d e n s ity a t th e o rth o p o s itio n ( f o r compounds a s th e pheny—
l a c e t i c and p h e n o x y ac e tic a c i d s ) .
Thimann (102) was th e f i r s t to su g g est
some c o r r e l a t i o n betw een a c t i v i t y and a ro m a tic o r i e n t a t i o n , b u t he f a i l e d
to g iv e any su p p o rtin g e v id e n c e .
The o n ly s e r io u s o b je c tio n t o t h i s hypo­
t h e s i s i s th e ev id en ce re p o rte d by Wain (116) t h a t 2 , 4 - d i c h lo r o - 6 - f lu o ro and 2 ,4 -d ib ro m o -6 -flu o ro p h e n o x y a c e tic a c id s have h ig h a c t i v i t y i n th e e lo n ­
g a tio n o f Avena c o le o p t ile s e c tio n s .
The B enzoic A cids
S u b s titu te d b e n z o ic a c id s were in tro d u c e d a s p o s s ib le g ro w th -re g u la to rs
by Zimmerman and H itchcock (6 0 ).
They re p o r te d a s l i g h t a c t i v i t y f o r 2 -
b ro m o -3 -n itro b e n z o ic a c id i n c e l l e lo n g a tio n and fo rm a tiv e e f f e c t s w ith 2 c h lo r o - 5 - n i t r o - and 2 , 3 , 5 -tr iio d o b e n z o ic a c id .
B en tley (13) re p o rte d a h ig h
a c t i v i t y f o r 2 , 3 , 6 - tr ic h lo r o b e n z o ic a c id i n th e e lo n g a tio n o f Avena c o ie o p t i l e s e c tio n s .
T h is fin d in g w hich was c o rro b o ra te d i n o th e r t e s t s (1 0 2 ,
112, 132) , s tim u la te d a renewed i n t e r e s t i n compounds of t h i s ty p e and le d t o
th e in v e s t i g a t i o n o f many o th e r s u b s t i t u t e d b e n zo ic a c id s (7 2 ,1 1 1 ,1 3 3 ,6 9 ),
From th e r e s u l t s o b ta in e d by th e s e in v e s t i g a t o r s i t can be concluded
t h a t s u b s tit u e n ts h aving a s tro n g +T e f f e c t do n o t c o n fe r a c t i v i t y w h ile
th o s e s u b s t i t u e n t s w ith r e l a t i v e l y weak o rth o p a ra d i r e c t i n g in flu e n c e
( C l,B r ,I , CH^, e t c . ) may do so.
i n th e 3 - p o s i t i o n .
t o be e s s e n t i a l .
The n i t r o group may o r may n o t be e f f e c t iv e
O r th o - s u b s titu tio n " a c tiv a te s " b e n zo ic a c id , and a p p e a rs
D i-o rth o s u b s t i t u t i o n c au ses a more pronounced a c t i v i t y
and 2 , 3 , 6 - d e r iv a tiv e s seem t o g iv e a maximum re s p o n se .
t h e 4 - p o s itio n m ust rem ain f r e e .
I n d ic a tio n s a re t h a t
- 27 -
Due t o t h e a p p a re n t n e c e s s ity o f o rth o s u b s t i t u t i o n , M uir and Hansch
( 72) su g g este d t h a t th e a c t i v i t y o f th e s e compounds in v o lv e d th e d is p la c e ­
ment o f an e l e c t r o n - a t t r a c t i n g group (h alo g en o r n i t r o ) i n th e o rth o p o s i­
t i o n , and t h a t th e r e i s a tw o -p o in t a ttach m en t w ith a c y s te in e u n it i n th e
same manner a s t h a t proposed f o r t h e p h en o x y ac e tic a c id s .
Hansch e t . a l . (4 0 )
were a b le to d e m o n strate t h e r e le a s e o f c h lo r id e from 2 , 6 -d ic h lo ro b e n z o ic
a c id d u rin g th e growth o f Avena c o le o p t ile s e c tio n s , th u s s u b s ta n tia tin g
t h i s h y p o th e s is .
A ccording to V e ld s tra and van de W esterin g h ( l i d ) th e d a ta
of Hansch e t . a l . do n o t p e rm it th e c o n c lu s io n t h a t th e r e le a s e o f c h lo rid e
io n i s e s s e n t i a l l y connected w ith th e p h y s io lo g ic a l a c t i v i t y o f t h e com­
pounds; th e y p o in t out t h a t th e in a c t iv e 2 , A ^d ich lo ro b en zo ic a c id r e le a s e s
c h lo rid e io n and such a r e le a s e should n o t be found f o r an in a c t iv e com­
pound.
V e ld s tr a (110,113) f u r t h e r p o in ts ou t t h a t on t h e b a s is of th e o rth o
r e a c tio n c o n c e p t, 2 , 6-dibrom obenzo ic a c id sh o u ld be a c t i v e , b u t i t was found
to b e in a c t iv e ( H O ) .
On th e o th e r hand, 2 , 6 -d im eth y lb e n z o ic a c id i s
s l i g h t l y a c t i v e , 3 - n itro -2 ,6 -d im e th y lb e n z o ic a c id i s a c t i v e and 3 - c h lo r o - ,
3-brom o-, and 3 -io d o -2 ,6 -d im e th y lb e n z o ic a c id a re h ig h ly a c ti v e (H O ).
Be­
cau se o f t h i s V e ld s tr a concludes t h a t a r e a c tio n w ith a p la n t s u b s tr a te
in v o lv in g d isp la ce m e n t o f t h e m ethyl group o r a n o th e r form o f chem ical r e a c ­
t i o n w ith t h e i n e r t o rth o s u b s tit u e n t can be r u le d o u t.
T h is a u th o r i s in c lin e d to a g re e w ith V e ld s tr a i n t h a t th e above and
fo llo w in g ev id en ce te n d s to i n v a l i d a t e th e o rth o r e a c tio n h y p o th e s is .
As
was p o in te d out i n th e d is c u s s io n of th e p h e n o x y ac e tic a c i d s , th e aro m atic
r in g c o n ta in in g a m eta d i r e c t i n g group lo c a te d o rth o o r p a ra to a group
which can r e a d i l y e x i s t a s an a n io n , i s p ro n e t o n u c le o p h ilic a t t a c k .
The
— 28 —
mechanism proposed by M uir and Hansch (72) i s a n u c le o p h ilic d isp lacem en t
o rth o t o th e c arb o x y l g ro u p .
A lthough t h i s mechanism would e x p la in th e
a c t i v i t y o f c e r t a i n o rth o s u b s t i t u t e d b e n zo ic a c id s , i t cannot acco u n t f o r
th e i n a c t i v i t y o f such compounds a s 2 -c h lo r o - 5 - n itr o b e n z o ic a c id and 2 ,4 d ic h lo ro b e n z o ic a c id .
I f th e a c t i v i t y i s d ependent on a n u c le o p h ilic d i s ­
placem ent i n th e o rth o p o s itio n , 2 - c h lo r o - 5 - n itr o b e n z o ic a c id , w ith th e
n i t r o group p a ra t o th e o rth o c h lo ro g ro u p , sh o u ld have a v e ry high a c t i v i t y
and i t i s t o t a l l y in a c t iv e ( 7 2 ) .
Holleman and De Mooy ( $ l) l i s t th e fo llo w ­
in g o rd e r f o r r e l a t i v e r a t e c o n s ta n ts f o r a number o f d ic h lo ro n itro b e n z e n e s
i n t h e i r n u c le o p h ilic rep lacem en t r e a c tio n w ith sodium m e th o x id e : 2 ,4 ; 3 ,4 ;
2 ,5 ; 2 ,3 ; 2 ,6 ( th e r e l a t i v e r a t e c o n s ta n ts i n h o u rs- ^ b e in g 1 9 .4 1 ; 17.42;
3 .9 3 ; 1 .7 4 and 0 .1 4 r e s p e c t i v e l y ) .
A ll th e s e compounds have an o rth o c h lo ro
group e x ce p t t h e 3 ,4 -is o m e r and i t i s seen t h a t th e l e a s t r e a c tiv e isom er i s
th e 2 , 6 - d e r i v a t i v e ; t h i s low r e a c t i v i t y i s e x p la in a b le by th e s t e r i c i n h i b i ­
t i o n o f re s o n a n c e , i . e . any s t r u c t u r a l m o d if ic a tio n which te n d s to d e s tro y
c o p la n a r ity i n h i b i t s reso n an ce and w ith o u t reso n an ce t h e n i t r o o r c arb o x y l
group cannot e x e r t i t s -T e f f e c t .
As was s t a t e above, th e 2 , 6 -r-d isu b stitu ted
b e n zo ic a c id s a r e among th e m ost a c tiv e a s p la n t growth r e g u l a t o r s , so i t
would seen th a t i f a n u c le o p h ilic mechanism i s o p e ra tin g , some f a c t o r o f a
param ount im p o rtan ce i s g iv in g r i s e t o th e d is c re p a n c y i n t h e r e s u l t s which
have been o b ta in e d th u s f a r .
The a u th o r does not l i k e to r u le ou t t h e n u c le o p h ilic d isp lacem en t
h y p o th e s is a s th e r e i s a s much ev id en ce i n i t s fa v o r a s a g a in s t i t .
Only
th ro u g h e x te n s iv e stu d y of f u r t h e r d e r iv a tiv e s can th e problem be c l a r i f i e d ^
- 29 -
A c tiv ity and O p tic a l Isom erism
Kogl and V erk aaik (59*60) found (+) fli-in d o le -3 -p ro p io n ic a c id to be 30
tim e s a s a c ti v e i n th e Avena c u rv a tu re t e s t a s t h e ( - ) iso m e r.
s t r a i g h t grow th t e s t th e two isom ers were e q u a lly a c t i v e .
I n th e
However, i t was
p roved t h a t th e d if f e r e n c e s i n th e Avena c u rv a tu re t e s t w ere caused by r e ­
t e n t i o n o f t h e ( - ) a c id i n th e apex by s e l e c t i v e a d s o r p tio n .
L a te r , how ever, d if f e r e n c e s i n a c ti v ity , betw een o p t i c a l enantiom orphs
were found i n t h e s t r a i g h t grow th and pea t e s t s (8 8 ,8 9 ,1 0 9 ,1 1 5 ,1 0 1 ,1 ).
A
su rv e y o f t h e s e r e s u l t s shows t h a t th e (+) -form showed t h e s tr o n g e s t grow th
s u b s ta n c e 'a c tio n .
Fredga and M a te ll (34) apd M a te ll (68) e s ta b lis h e d t h a t
th e a c t i v e enantibm orph o f a d l p a i r has t h e d c o n f ig u r a tio n .
V e ld s tra
( 113) s t a t e s t h a t th e s e r e s u l t s s t r e s s th e em inent, im p o rtan ce o f th e s p a t i a l
form of t h e a c ti v e m o le c u le,, and i f th e grow th re sp o n se depends on th e
degree., of f i t t i n g o f th e a c t i v e m olecu le on t h e prim ary a c ti v e s i t e , one
would ex p ect th e m ost a c ti v e a n tip o d e s to b e lo n g t o t h e same s t e r i c s e r ie s *
i r r e s p e c t i v e o f t h e i r s ig n o f r o t a t i o n .
I t must be remembered, however,
t h a t th e m ost a c ti v e grow th s u b s ta n c e s , ip d o le a c e tic a c id , n a p h th a le n e a c e t i c a c id and 2 ,4 -d ic h lo ro .p h e n o x y a c e tic a c id a r e n o t o p t i c a l l y a c tiv e and
h in d ra n c e w ith t h e i n t e r a c t i o n o f th e grow th su b sta n c e and i t s c e l l u l a r
s u b s tra te .
M on-acidic Compounds
Acid d e r i v a t i v e s , such a s - e s te r s and am ides have been found g e n e r a lly
a c t i v e , b u t t o a l e s s e r e x te n t th a n th e a c ti v e f r e e a c i d .
I t i s g e n e r a lly
a c c e p te d t h a t a c o n v e rsio n o f th e , d e r iv a tiv e in to th e p a r e n t a c id i s a p r e -
- 30 -
re q u is ite fo r a c tiv ity .
A ll in d ic a tio n s a r e t h a t th e a c t i v i t y o f ald eh y d es (1 1 3 ,5 ,6 3 ) , n i t r i l e s
(101,127) and a lc o h o ls (113) i s due t o a c o n v e rsio n to th e co rresp o n d in g
a c id .
There i s some in d i c a t i o n , how ever, t h a t some n i t r i l e s a re a c ti v e p e r
se (4 2 ,5 4 ) .
New P la n t G ro w th -re g u la to rs
The two m ain s e r i e s of compounds which were chosen f o r stu d y were th e
s u b s t i t u t e d p h en ylhydrazones o f g ly o x y lic and p y ru v ic a c id s .
tf - a n ilin o a c r y l i c a c id s were a ls o s y n th e s iz e d and t e s t e d .
S e v e ra l
The s t r u c t u r a l
s i m i l a r i t y of th e s e compounds to in d o le a c e tic a c id and th e p h en o x y acetic
a c id s i s o b v ious:
H
CCOOH
Il
C H 2 CO O H
H
COOH
I
H
/ f X
r
1V
fC H ,
CCOOH
(I
I
x n
HCCOOH
Il
Zh
To th e a u t h o r 's knowledge, th e on ly compounds o f a s im ila r s tr u c tu r e which
have been t e s t e d a s p la n t g ro w th -re g u la to rs a r e th e mono-and d i - n i t r o pnehylhydrazone d e r iv a tiv e s o f l e v u lin ic a c id (70) and ^ -d ip h en y la m in o p ro p io n ic a c id ( 4 ) .
A ll th e s e compounds were re p o rte d t o show a c t i v i t y a s
g ro w th - re g u la to r s o r h e r b ic id e s .
The p h en y lh y d razono group i s s im ila r in
s iz e to th e phenoxy group and has an o rth o -p a ra d i r e c t i n g in flu e n c e on th e
benzene r i n g .
I t should n o t be u n lik e ly , th e n f o r th e s e compounds t o show
a c t i v i t y a s p la n t g r o w th - re g u la to r s .
The same argum ents would a p p ly to th e
- 31 -
/8 -a n ilin o a c r y l i c a c id s ,
2 -riitro -4 -m e th o x y p h e n o ^ ra c e tic a c id was a ls o s y n th e s iz e d and t e s t e d
i n t h e hope t h a t t h e combined o r i e n t a t i o n e f f e c t s o f th e n i t r o ( s tro n g
-T ) and th e methoxy (stro n g + ? ) groups would p a r e l l e l t h a t o f th e 2 ,4 d ic h lo r o d e r i v a t i v e .
ct-P h en y lim in o p ro p io n ic a c id was a ls o p re p a re d and t e s t e d .
f*
- 32 -
IV . EXPERIMENTAL
The t e s t p la n ts w ere grown a c c o rd in g t o Bonner (1 5 ) .
The headrow o a t
seed s were g erm in ated i n sand i n th e d a rk a t a te m p e ra tu re o f 2 9° C and a
r e l a t i v e h u m id ity o f 80—90$’.
A fte r 96 h o u rs th e a p ic a l 5 mm (a p p ro x .) • o f
th e c o le o p t ile were removed and two h o u rs l a t e r s e c tio n s w ere cu t from th e
apex w ith t h e p rim ary l e a f in c lu d e d w ith in
th e c o l e o p t i l e .
No more th a n
two s e c tio n s w ere c u t from one c o l e o p t i l e .
The c u t t e r used gave s e c tio n s
7 .6 1 u n i t s (5«74 mm) i n le n g th w ith a s ta n d a rd d e v ia tio n o f 0.0 3 u n i t .
S in g le d e te rm in a tio n s c o n ta in in g e i t h e r 10 o r 15 s e c tio n s w ere run..
The
s e c tio n s were f lo a te d on th e s u rfa c e o f 25 ml of th e s o lu tio n i n a P e t r i
d is h and p la c e d i n an in c u b a to r a t 28° G f o r 24 h o u rs .
M easurem ents were
made w ith an o c u la r m icrom eter mounted in a b in o c u la r m icroscope (13X )„
B efore p l a n tin g , th e sand was le a c h e d w ith d i l u t e h y d ro c h lo ric a c id
and th e n w ith d i s t i l l e d w a te r u n t i l th e f i l t r a t e proved f r e e from c h lo rid e
io n .
P r e p a r a tio n of S o lu tio n s
Whenever p o s s ib le , th e compounds w ere sim ply d is s o lv e d i n d i s t i l l e d
w a te r .
The a d d itio n o f b a se was n e c e s s a ry i n many in s ta n c e s , due t o th e
low s o l u b i l i t y o f th e f r e e a c id .
I n a l l e s s e s , how ever, th e pH o f th e
s o lu tio n was below 7 .0 .
Due t o th e v e ry low s o l u b i l i t y o f th e e s t e r s , an
em ulsion was em ployed.
The fo llo w in g example o f e th y l 06-phenyhydrazono-
p ro p io n a te i s t y p i c a l :
0.2062 g o f e th y l &-phenyIh y d raz o n o p ro p io n a te was
p la c e d i n a 250 ml v o lu m e tric f l a s k and I m l o f EMCOL H-68c ( g en ero u sly
s u p p lie d by th e E m ulsol Chem ical C o rp o ra tio n ) s o lu tio n (10% i n x y le n e) was
added and th e m ix tu re g e n tly shaken u n t i l com plete s o lu tio n was o b ta in e d .
- 33 -
A pproxim ately 150-200 cc of w a te r were added and th e m ix tu re shaken v ig o r ­
o u sly t o y i e l d a m ilk y -w h ite e m u lsio n „ The em ulsion was th e n made up to
volume and shaken.
T h is pro ced u re y i e l d s a 0,004 M em ulsion o f th e e s t e r
and c o n ta in s c a , 0*4% x y le n e and c a , 0,04% EMCOL H-86c .
T h is c o n c e n tra tio n
of x y le n e and e m u ls if ie r and t h e c o n c e n tra tio n s c o n ta in e d i n subsequent
d i l u t i o n s w ere shown t o have no e f f e c t on t h e grow th o f th e c o le o p tile
s e c tio n s ,
Q^-Phenylhydraz o n o p ro p io n ic Acid
8 ,8 g ( 0 ,1 mole) o f p y ru v ic a c id were d is s o lv e d i n 40 cc o f e th y l e th e r
an d 1 0 ,8 g ( 0 ,1 m ole) o f pheny!h y d ra z in e d is s o lv e d i n 50 cc of e th y l e th e r
w e re'a d d ed w ith s t i r r i n g d u rin g a p e rio d o f 5 m in u te s .
The l i g h t y ello w
p r e c i p i t a t e w hich formed im m ed iately , was removed by f i l t r a t i o n , washed
tw ic e w ith e th e r and a i r d r ie d .
The hydrazone was r e c r y s t a l l i z e d from d i l u t e
a lc o h o l t o y i e l d l i g h t y e llo w n e e d le s .
i c a l ) ',
C o rre c te d m e ltin g p o in t:
The y i e l d was 1 6 .5 g (97% of th e o r e t^
192p C„
C u rtu is (2 6 ) g iv e s th e m e ltin g
p o in t a s 192° C„
P h e n y lh y d ra z o n o ac e tic Acid (2 1 )
20 g o f d ic h lo r o a c e tic a c id were d is s o lv e d in 20 cc o f w a te r and a
s o lu tio n o f 34 g o f p o tassiu m h y d ro x id e i n 60 cc o f w a te r was added.
To
t h i s s o lu tio n , 16 g of p h en y !h y d razin e d is s o lv e d i n 20 cc o f 95% e th y l
a lc o h o l were added and th e r e s u l t i n g s o lu tio n r e flu x e d f o r two h o u rs .
The
a lc o h o l was th e n removed by d i s t i l l a t i o n and th e aqueous s o lu tio n e x tr a c te d
w ith t h r e e 20 cc p o r tio n s o f e th e r .
d is tilla tio n .
The rem ain in g e th e r was removed by
The s o lu tio n was allow ed t o c o o l to room te m p e ra tu re (c a .
2 5 ° C) and s tr o n g ly a c i d i f i e d w ith a c e t i c a c id .
The re d -o ra n g e p r e c i p i t a t e
- 34 —
was f i l t e r e d o f f and ,d is s o lv e d i n c a . 2500 cc of warm w a te r (45- 50° C5 a
te m p e ra tu re h ig h e r th a n t h i s g iv e s r i s e t o e x te n s iv e d eco m p o sitio n upon th e
fo llo w in g a c i d i f i c a t i o n ) .
The s o lu tio n was f i l t e r e d and 30 cc o f concen­
t r a t e d h y d ro c h lo ric a c id was added.
brown n e e d le s s e p a r a te d .
Upon c o o lin g t o 5 -1 0 ° C5 long d ark
P u r i f i c a t i o n was accom plished by r e d is s o lv in g th e
p ro d u c t i n 2500 cc o f warn w a te r (4 5 -5 0 ° C) c o n ta in in g 10 g o f p o tassiu m h y d ro x id e , s t i r r i n g w ith a sm a ll amount o f N o r ite 5 f i l t e r i n g and a c id if y in g
w ith 30 cc o f c o n c e n tra te d h y d ro c h lo ric a c id .
y ie ld e d la r g e l i g h t brown n e e d le s .
c a l) .
C ooling o f t h e s o lu tio n
•
The y i e l d was 1 9 .6 g (81% o f t h e o r e t i ­
M e ltin g p o in t w ith d eco m p o sitio n : 141° G ( c o r r e c t e d ) .
M u lle r (75)
g iv e s th e m e ltin g p o in t a s 142-3° C.
Busch e t . a l . (21) o b ta in e d two is o m e ric form s w ith t h i s p ro c e d u re , b u t
th e a u th o r was u n a b le to d u p lic a te t h e i r r e s u l t s on many a tte m p ted ru n s .
Due to th e i n s t a b i l i t y o f t h i s compound, i t was co n v erted t o i t s p o ta s ­
sium s a l t by t r e a t i n g i t s a lc o h o lic s o lu tio n w ith th e e q u iv a le n t amount o f
a lc o h o lic p o ta ssiu m h y d ro x id e .
The p r e c i p i t a t e d s a l t was washed w ith r
a b s o lu te a lc o h o l and d r ie d i n a vacuum d e s ic c a to r .
sium:
A n a ly sis o f p e rc e n t p o ta s ­
c a lc u la te d , 19. 34%> found, 1 9 . 28%.
M -Phenylim inopropionic Acid
8 .8 g ( 0 .1 mole) o f p y ru v ic a c id w ere d is s o lv e d i n 40 cc o f e th y l e th e r
and a s o lu tio n o f 9-3 g ( 0 .1 m ole) o f a n i l i n e d is s o lv e d i n 40 cc o f e th e r
was added w ith s t i r r i n g .
The l i g h t y e llo w p r e c i p i t a t e was f i l t e r e d o f f and
r e c r y s t a l l i z e d from an a lco h o l-b e n z e n e s o lu tio n .
of t h e o r e t i c a l ) .
The y i e l d was 6 .8 g (42%
M e ltin g p o in t: '127-28° C ( c o r r e c t e d ) ,
th e m e ltin g p o in t a s 127-28° C.
B e i l s t e i n ( 8) l i s t s
- 35 -
-E t h y lf o rm y la c e ta te . S o d im E n o la te (125)
In a th re e -n e c k e d f l a s k f i t t e d w ith a r e f lu x c o n d en ser, a m ercu ry s e a l s t i r r e r , and a drop fu n n e l were p la c e d 350 cc o f anhydrous e th e r and
23 g o f sodium w ir e .
A m ix tu re o f 90 g of e th y l a c e ta te and 85 g of e th y l
fo rm ate was slow ly added ov er a p e rio d o f a p p ro x im a te ly one h o u r.
The
m ix tu re was s t i r r e d under g e n tle r e f l u x f o r ap p ro x im a te ly 4 h o u rs .
5 cc o f
95% e th y l a lc o h o l were th e n added and th e m ix tu re r e f lu x e d an a d d itio n a l 4
h o u rs , o r u n t i l th e l a s t t r a c e s of sodium had d is a p p e a re d .
dense y e llo w p r e c i p i t a t e form ed.
The p r e c i p i t a t e was removed by s u c tio n
f i l t r a t i o n and allow ed t o a i r d ry .
y ello w n e e d le s .
Upon s ta n d in g a
The p r e c i p i t a t e c o n s is te d o f f in e
The y i e l d was 86 g .
n e a r ly c o lo r le s s p ro d u c t was o b ta in e d .
By re p e a te d w ashing w ith d ry e th e r a
A n a ly sis showed th e compound t o be
75*5% p u re ; t h i s compares t o th e r e s u l t s o b ta in e d by W islic e n u s (1 2 5 ).
E t h y l/d - a n ilin o a c r y la te (82)
4 .6 g (0 .0 5 mole) o f a n i l i n e was d is s o lv e d i n 100 cc o f 5% a c e tic a c id ,
A s o lu tio n o f e th y lfo rm y ! a c e t a t e , sodium e n o la te (9 ,6 g i n 50 cc o f w a te r;
e q u iv a le n t t o 0 ,0 5 mole o f e n o la te ) was added w ith s t i r r i n g .
The y ello w
p r e c i p i t a t e which form ed im m ed iately , was f i l t e r e d o f f and r e c r y s t a l l i z e d
from 95%> a lc o h o l.
The p ro d u c t s e p a ra te d a s l i g h t y e llo w p l a t e s .
was 5 .4 g (57% of t h e o r e t i c a l ) .
M e ltin g p o in t: 113° C.
The y i e l d
S tr a u s and Voss
(94) g iv e th e m e ltin g p o in t a s 1 1 4 -1 4 .5 ° C.
2 .4 - D ic h lo r o a c e ta n ilid e ( 22, 86)
100 g of a c e t a n i l i d e w ere d is s o lv e d i n 800 cc of g l a c i a l a c e tic a c id
and 125 g o f sodium a c e ta te ( f r e s h l y fu se d and powdered) were added.
The
f l a s k and i t s c o n te n ts w ere t a r e d and c h lo r in e was p a sse d i n t o th e m ix tu re
— 36 —
th ro u g h a s p e c ia l a d d itio n tu b e (3 0 ),
T h is a d d itio n tu b e was n e c e s sa ry t o
p re v e n t c lo g g in g by th e p r e c i p i t a t i o n o f p - c h lo r o a c e ta n ilid e and l a t e r th e
2 .4 -
d ic h lo ro a c e ta n ilid e .
As soon a s th e p - c h lo r o a c e ta n ilid e had c o m p letely
p r e c i p i t a t e d (a b s o rp tio n o f c a . 53 g o f c h lo r in e ) , th e f l a s k was h e a te d on
a w a te r b a th and th e p a ssa g e o f c h lo rin e was co n tin u ed u n t i l a t o t a l o f
105 g had been a b so rb e d .
The m ix tu re was allo w ed to c o o l and th e ' s o lid
which s e p a ra te d was removed by f i l t r a t i o n .
from th e p r e c i p i t a t e by d ig e s tio n w ith w a te r.
The sodium c h lo r id e was removed
No a tte m p t was made t o r e ­
cover a l l of th e 2 , 4 - d ic h lo r o a c e ta n ilid e , s in c e th e m other liq u o r was used
f o r th e c h lo r in a tio n o f a n o th e r 100 g o f a c e t a n i l i d e (125 g o f sodium
a c e ta te a g a in b e in g a d d e d ).
A fte r th e s o lid from th e second c h lo r in a tio n
had been removed by f i l t r a t i o n , th e m other li q u o r was d i lu te d w ith an' e q u a l
volume of warm w a te r and th e m ix tu re co o led to c a . 5° C.
The s o lid which
p r e c i p i t a t e d was removed and combined w ith t h e f i r s t two y i e l d s .
The com­
b in e d p ro d u c t was d ig e s te d w ith w a te r t o in s u r e com plete rem oval o f sodium
c h lo r id e , d r ie d and r e c r y s t a l l i z e d from 95% e th y l a lc o h o l.
The r e c r y s t a l ­
l i z a t i o n y ie ld e d la r g e w h ite c r y s t a l s w hich gave a c o r r e c te d m e ltin g p o in t
o f 145° C.
C hattaw ay e t . a l . (22) r e p o r t a m e ltin g p o in t o f 144° C.
The
t o t a l y i e l d was 268 g ( 89% of t h e o r e t i c a l ) .
2 .4 - D ic h lo ro a n ilin e (22)
268 g o f 2 , 4- d ic h lo r o a c e ta n ilid e were d is s o lv e d i n 450 cc of concen­
t r a t e d s u l f u r i c a c id and t h e s o lu tio n h e a te d a t a te m p e ra tu re betw een HO
and 120° C f o r ca.* one h o u r.
The ,.s o lu tio n was allo w ed t o c o o l t o room
te m p e ra tu re (c a . 26° C) arid poured slo w ly i n t o 2 l i t e r s o f c o ld (0 ° C) 25%
sodium c arb o n a te s o lu tio n .
The heavy w h ite p r e c i p i t a t e was f i l t e r e d o f f
- 37 -
and washed once w ith i c e c o ld w a te r.
The 2 , 4 - d i c h lo r o a n ilin e was th e n s u s ­
pended i n c a . 2 l i t e r s o f c o ld w a te r a n d .40% NaOH was added u n t i l t h e mix­
t u r e was b a s ic t o li t m u s .
A fte r s t i r r i n g r a p id ly f o r 30 m in u te s, th e 2 , 4 -
d i c h lo r o a n ilin e was f i l t e r e d o f f and a i r d r i e d .
of th e o re tic a l).
M e ltin g p o in t: 62-63° C.
p o in t a s 6 2 .5 -6 3 ° 0.
The y i e l d was 202 g (94.8%
Hyde (52) g iv e s th e m e ltin g
I f t h e 2 , 4 - d ic h lo r o a c e ta n ilid e i s o f h ig h p u r i t y , no
f u r t h e r p u r i f i c a t i o n o f t h e r e s u l t i n g a n i l i n e i s n e c e s s a ry .
2 ,4 -D ic h lo ro p h e n y Ih y d ra z in e H y d ro ch lo rid e
W ith s e v e r a l m o d if ic a tio n s , th e p ro c e d u re o f Coleman (25) f o r th e
p r e p a r a tio n o f phenylhyd ra z in e was fo llo w e d .
I n a 500 m l, 3-n eck ed f l a s k ,
f i t t e d w ith a m e ch a n ic al s t i r r e r , were p la c e d 62 .5 cc o f c o n c e n tra te d
com m ercial h y d ro c h lo ric a c id (22 Be ) .
4 0 .5 g (0 .2 5 m ole) o f 2 ,4 - d ic h lo r o -
a n i l i n e were th e n added and th e m ix tu re s t i r r e d u n t i l a t h i c k , homogenous
p a ste .Was-rTormed, The f l a s k was surrounded w ith a f r e e z in g m ix tu re o f ic e
and s a l t , and when th e ,contents were O^ o r lo w er, s t i r r i n g was s t a r t e d and
62 g o f crack ed ic e w ere added; th e n a c o ld s o lu tio n ( 0° o r low er) o f 1 8 .1
g o f NaNO-j d is s o lv e d i n 38 cc o f w a te r was allow ed t o ru n i n slow ly (20-30
m in u te s) from a d ropping fu n n e l, th e end o f which was drawn to a sm all t i p
and re a ch e d n e a r ly to t h e bottom o f t h e f l a s k .
V igorous s t i r r i n g was m ain­
ta in e d and th e te m p e ra tu re was kept a s n e a r z e ro a s p o s s ib le by fre q u e n t
a d d itio n o f crack ed ic e (up to 65 g ) .
D uring t h i s a d d itio n , a sodium s u l f i t e , s o lu tio n was p re p a re d by d i s ­
so lv in g 80 g o f anhydrous sodium s u l f i t e i n 375 cc o f w a te r .
The s o lu tio n
was p la c e d i n a 3 l i t e r , 3-n eck ed f l a s k and co o led t o m inus 5° 0 .
A pproxi­
m a te ly 30 g o f ic e was added t o t h i s s o lu tio n and w ith r a p id s t i r r i n g th e
it
— 38 —
diazonium s a l t s o lu tio n was added a s r a p i d l y a s p o s s ib le .
I f th e two
s o lu tio n s a re not s u f f i c i e n t l y c o ld , ppor re d u c tio n r e s u l t s due t o lo s s o f
SO2 . The f l a s k was th e n warmed on a steam b ath to 70° C, and 25 cc o f con­
c e n tr a te d h y d ro c h lo ric a c id was added ( t e s t i n g w ith litm u s t o be su re th e
s o lu tio n was a c i d ) .
The f l a s k was k ep t a t 60-70° C f p r c a . 2 h o u rs .
250 cc o f c o n c e n tra te d com m ercial h y d ro c h lo ric a c id w ere th e n added
and th e m ix tu re was c o o led t o 0 ° .
The p r e c i p i t a t e d 2 ,4 -d ie h lo ro p h e n y l-
h y d ra z in e h y d ro c h lo rid e was f i l t e r e d o f f and a i r d r ie d .
;
Tpe p ro d u c t was
r e c r y s t a l 'l i z e d from a d i l u t e e th y l a lc o h o l s o lu tio n t o which a few cc o f
c o n c e n tra te d h y d ro c h lo ric a c id had been added.
n ig h t i n a vacuum d e s ic c a to r .
A n a ly s is .
C a lc u la te d f o r
N e u tr a liz a tio n e q u iv a le n t.
The compound was d rie d o v er­
The y i e l d was 4 4 .1 g (82.5% o f t h e o r e t i c a l ) .
C l, 4 9 ,8 3 .
C a lc u la te d :
Found:
213. 5 ; found:
The m e ltin g p o in t was 212° C. w ith d eco m p o sitio n .
C l, 4 9 . 6 9.
2 1 2 .8 .
Chattaw ay and P earce (23)
g iv e th e m e ltin g p o in t a s 210° C w ith d e co m p o sitio n .
The f r e e b ase i s
o b ta in e d by t r e a t i n g an aqueous s o lu tio n o f th e h y d ro c h lo rid e w ith ex cess
base.
R e q r y s t a l l i z a t i o n from d i l u t e a lc o h o l y ie ld s n e e d le s ; m e ltin g p o in t
was 94° C.
Chattaw ay and P earce g iv e 94° C.
The p ro c e d u re g iv e n f o r t h e p r e p a r a tio n o f 2 ,4 - d i chlo ro p h en y Ih y d ra z in e
h y d ro c h lo rid e i s a g e n e ra l one and was used w ith only m inor changes f o r t h e
p r e p a r a tio n o f th e fo llo w in g a ry !h y d ra z in e h y d ro c h lo rid e s .
p-T o ly Ih y d ra z in e H y d ro ch lo rid e
The p ro d u c t was r e c r y s t a l l i z e d from c a . 2 N. h y d ro c h lo ric a c id and
s e p a ra te d a s c o l o r l e s s n e e d le s .
The y i e l d was 105 g ( 86% o f t h e o r e t i c a l ) .
T reatm ent o f th e h y d ro c h lo rid e w ith e x ce ss b a se l i b e r a t e d th e h y d ra z in e .
- 39 -
which s e p a ra te d a s c o lo r le s s l e a f l e t s upon r e c r y s t a l l i z a t i o n from d i l u t e
a lc o h o l.
M e ltin g p o in t of th e f r e e b a se : 64° C.
Bamberger (? ) g iv e s th e
m e ltin g p o in t a t 65° G.
p -C h lo rophenyIh y d ra z in e H y d ro c h lo rid e . •
The compound was r e c r y s t a l l i z e d from w a te r which c o n ta in e d a few cc o f
c o n c e n tra te d h y d ro c h lo ric a c id .
w ith d eco m p o sitio n a t 228-230° C.
J t c r y s t a l l i z e d a s a f i n e ppwder and m e lte d
The y i e l d was 46 g (52% o f t h e o r e t i c a l ) .
B e i l s t e i n (lO ) g iv e s th e m e ltin g p o in t a s 225-230° C w ith d eco m p o sitio n .
A n a ly s is .
C a lc u la te d f o r C^HgNgClg:
N e u tr a liz a tio n e q u iv a le n t.
C l, 3 9 .6 ? .
C a lc u la te d :
Found:
C l, 3 9 .5 6 .
1 7 9 .0 j found: '1 7 8 ,2
The f r e e b ase c r y s t a l l i z e d a s n e e d le s from e th e r ; m e ltin g p o in t 90° C.
B e i l s t e i n (10) g iv e s 90° C.
o-C hlorophenyIh y d ra z in e H y d ro ch lo rid e
The compound c r y s t a l l i z e d from d i l u t e a lc o h o l a s f i n e y e llo w n e e d le s .
The y i e l d was 56 g (63% o f t h e o r e t i c a l ) .
M e ltin g p o i n t :
194° C, w ith
d e co m p o sitio n . B e i l s t e i n (9) l i s t s th e m e ltin g p o in t a s 194° C w ith decom­
p o s itio n .
A n a ly s is .
C a lc u la te d f o r C^lHgNgClg;
C l, 3 9 .6 7 .
Found:
The f r e e b a se c r y s t a l l i z e s a s n e e d le s from d i l u t e a lc o h o l.
p o in t:
47° C.
177*9
M eltin g
B e i l s t e i n (9) g iv e s 4 6 -4 7 ° C.
2 . 5-D ichlorophenyIh y d ra z in e H y d ro ch lo rid e
The h y d ro c h lo rid e s e p a ra te d a s v e ry f i n e l i g h t brown n e e d le s from
d i l u t e a lc o h o l.
A n a ly s is .
Y ie ld was 4 9 .6 g (93% o f t h e o r e t i c a l ) .
C a lc u la te d f o r C^HyNgCl^;
C l, 4 9 .8 3 ; N,. 1 3 .1 2 .
Found:
- 40 -
C l, 49; N, 1 3 , 0 6 .
N e u tr a liz a tio n e q u iv a l e n t.
C a lc u la te d :
2 1 3 .5 ; fo und:
The f r e e b ase c r y s t a l l i z e s a s n e e d le s from w a te r.
1060C.
2 1 2 .6 .
M e ltin g p o in t:
Z e t t e l (126) g iv e s th e m e ltin g p o in t a s 105° C.
< K -(p -T o ly lh y d razo n o )-p ro p io n ic a c id
1 5.86 g ( 0 .1 m ole) o f p - to ly !h y d ra z in e h y d ro c h lo rid e were d is s o lv e d i n
80 cc o f w a te r ( c a . 70° C) and 8 ,9 g ( 0 ,1
mole) of. p y ru v ic a c id d is s o lv e d
i n 20 cc o f w a te r were added w ith s t i r r i n g .
The y e llo w p r e c i p i t a t e which
formed was f i l t e r e d o f f a f t e r th e m ix tu re had co o led t o c a . 1 0 °,C .
hydrazone was r e c r y s t a l l i z e d from d i l u t e a lc o h o l.
y e llo w n e e d le s and m e lte d a t l6 l° C .
c a l) .
The
I t c r y s ta lliz e d as
The y i e l d was 7 , 4 g (39% of t h e o r e t i ­
B e i l s t e i n ( l l ) g iv e s th e m e ltin g p o in t a s 162°C,
N e u tr a liz a tio n e q u iv a le n t.
C a lc u la te d f o r CqoHigNgOg;
1 9 2 .2 ; found:
192. 8 .
E th y l /d -( p-m ethyl a n i l i n o ) - a e r y I a t e
The p ro ced u re used was s im ila r t o th e one employed i n th e p r e p a r a tio n
o f e th y l /S - a n i lin o a c r y la te .
The y e llo w p r e c i p i t a t e which formed was r e - .
c r y s t a l l i z e d from d i l u t e a lc o h o l to y i e l d l i g h t y e llo w p l a t e s .
was 4 .1 g (40% of t h e o r e t i c a l ) ,
M eltin g , p o in t: 116° C.
The y ie ld
Pechmann (82)
g iv e s th e m e ltin g p o in t a s 116° C.
A n a ly s is .
C a lc u la te d f o r CigHi^NOg:
N, 6 .8 2 /
Found:
N, 6 .7 3 .
# - ( o -G h lo ro p h en y lh y d raz o n o )-p ro p io n ic Acid
8 .9 5 g (0 .0 5 mole) o f o -c h lo ro p h e n y lh y d ra z in e h y d ro c h lo rid e were d i s ­
so lv ed in, 250 cc o f warm w a te r (70-80° C) and 4 .4 5 g (0 .0 5
m ole) o f
p y ru v ic a c id d is s o lv e d i n 10 cc o f w a te r w ere added w ith sh a k in g .
A y e llo w
f
— 41 ■
p r e c i p i t a t e form ed im m ediately and a f t e r c o o lin g th e m ix tu re to c a. 20° C,
■th e p r e c i p i t a t e was f i l t e r e d o f f and r e c r y s t a l l i z e d from a lc o h o l.
y ie ld was 3 .5 g (33% of t h e o r e t i c a l ) .
y e llo w n e e d le s .
The
The hydrazone c r y s t a l l i z e d a s lem on-
M e ltin g p o in t: '1 7 6 ° C.
H ew itt (44) g iv e s th e m e ltin g
• p o in t a s 178° C.
A n a ly s is .
C a lc u la te d f o r C ^ H ^ N g O g C lC l, 1 6 .6 8 .
N e u tr a liz a tio n e q u iv a l e n t,
C a lc u la te d :
Found:
2 1 2 .6 ; found:
C l, 1 6 .7 3 .
2 1 2 .6 .
fl& -(p-C hlorophenylhydrazono)-propionic A cid
The p ro ced u re was th e same a s t h a t used f o r a -(o -c h lo ro p h e n y lh y d ra z o n o )p ro p io n ic a c id .
The hydrazone was r e c r y s t a l l i z e d from d i l u t e a lc o h o l and
s e p a ra te d a s lem on-yellow n e e d le s .
M e ltin g p o in t:
A n a ly s is .
199° C.
The y i e l d was 5.3 g (50% o f t h e o r e t i c a l ) .
H ew itt (43) g iv e s th e m e ltin g p o in t a s 199° C.
C a lc u la te d f o r CgHgN2C^Cil:
N e u tr a liz a tio n e q u iv a l e n t.
C l, 1 6 .6 8 .
C a lc u la te d :
Found:
C l, 1 6 ,6 1 .
2 1 2 .6 ; fo u n d :. 2 1 3 .4 .
E th y l 2 .4 -D ic h lo ro p h enyIh y d ra z o n o a c e ta t e (24)
1 0 .6 8 g (0 .0 5 m ole) o f 2 , 4 -d ic h lo ro p h e n y !h y d ra z in e h y d ro c h lo rid e ,
8 .2 7 g (0 .0 5 m ole) o f c h lo r a l h y d ra te , and 50 cc o f a b s o lu te a lc o h o l were
p la c e d i n a 200 ml round-bottom f l a s k which was p ro v id e d w ith a r e f l u x
c o n d en se r.
The m ix tu re was slow ly h e a te d on a steam b a th u n t i l evidence
o f r e a c tio n o c c u rre d ( c a . 650 C ).
The m ix tu re was th e n slo w ly h e a te d t o
b o ilin g and g e n tly r e flu x e d f o r 30 m in u te s .
A pproxim ately 25 cc o f a lc o h o l
were removed by d i s t i l l a t i o n and th e r e s id u e was allow ed to c o o l to room
te m p e ra tu re .
The b ro w n ish -y e llo w p rism s which p r e c i p i t a t e d were f i l t e r e d
o f f and r e c r y s t a l l i z e d from 95% a lc o h o l.
p a le y e llo w p ris m s .
The r e c r y s t a l l i z a t i o n y ie ld e d
The y i e l d was 6 ,9 g (53% o f t h e o r e t i c a l ) .
M eltin g
-Ap­
p o in t: 122° C.
C hattaw ay and B ennett (24) giy.e th e m e ltin g p o in t a s
1 2 1 .5 ° C.
A n a ly s is .
C a lc u la te d f o r C10H10N2O2Cl2 :
C l, 2 7 .l 6 .
Found:
C l, 2 7 .0 9 .
2 , A -D ichlorophen y lh y d ra zo n o a c etic- A cid ■
'
5 g (0 .0 1 9 mole) of e th y l 2 ,4 -d ic h lo ro p h e n y lh y d ra z o n o a c e ta te were
added t o a f l a s k c o n ta in in g a 10% p o ta ssiu m h y d ro x id e s o lu tio n i n 95%
a lc o h o l.
The s o lu tio n was. re flu x e d f o r c a . I . 5 h o u rs and 70 cc of w a ter
were added.
The a lc o h o l was removed, by d i s t i l l a t i o n and th e 'c le a r re d
re s id u e was cooled to room te m p e ra tu re .
20 cc of c o n c e n tra te d h y d ro c h lo ric
a c id w ere added w ith s t i r r i n g and th e r e s u l t a n t g re e n is h p r e c i p i t a t e r e ­
moved by f i l t r a t i o n .
powdered p ro d u c t.
was 158-9° G.
A n a ly s is .
A ll a tte m p te d r e c r y s t a l l i z a t i o n s r e s u l t e d i n a f i p e l y
The y i e l d was 3 .8 g (85% of t h e o r e t i c a l ) .
M eltin g p o in t .
Np l i t e r a t u r e v a lu e was found.
C a lc u la te d f o r CgH^N2O2Cl2 :
N e u tr a liz a tio n e q u iv a le n t.
C a lc u la te d :
C l, 3 0 .4 2 .
Found:
2 3 3 ,1 ; fo u n d :
C l, 30*50,
2 3 3 .1 .
v
E th y l q-C hlorophenyIh y d ra z o n o a c e ta te
8.95 g ( 0 .0 5 mole) of p -c h lo ro p h e n y !h y d ra z in e h y d ro c h lo rid e , 8 .3 g
(0 .0 5 m o le )o f c h lo r a l h y d ra te and 100 ml of a b s o lu te a lc o h o l were p la ce d
i n a 200 ml round-bottom f l a s k and slow ly h e a te d to b o i l i n g .
The s o lu tio n
was r e flu x e d f o r 30 m in u te s and a f t e r c o o lin g was poured i n t o c a. 500 cc
o f cold! w a te r.
The brow nish p r e c i p i t a t e was f i l t e r e d o f f and r e c r y s t a l l i z e d
from d i l u t e a lc o h o l.
n e e d le s .
The y i e l d was 7 ,4 g ( 65% of t h e o r e t i c a l ) o f f in e
M e ltin g p o in t:
132° C w ith d e co m p o sitio n .
No l i t e r a t u r e v a lu e
was fo u n d ,
A n a ly s is .
C a lc u la te d f o r G10H^oN2O2 O l:
C l, 1 5 .6 4 .
Found:
C l, 15.6'#.
- 43 —
E th y l 2 , $-D lchlorophenyIh y d ra z o n o a c e ta te
The p ro c e d u re was s im ila r t o t h a t used f o r th e p r e p a r a tio n o f e th y l
2 , 4 - d i chlorophenyIh y d raz o n o a c e ta te .
R e c r y s t a l l i c a t i o n from d i l u t e a lc o h o l
y ie ld e d a f i n e y e llo w powder (which c o n s is te d o f m ic ro sco p ic p r is m s ) ,
y i e l d was 7 .1 g (54% of t h e o r e t i c a l ) .
M e ltin g p o in t:
8 5 -6 ° C.
The
No l i t e r ­
a tu r e v a lu e was fo u n d .
A n a ly s is .
C a lc u la te d f o r Cio H10N2O2 CI2 :
C l, 2 7 .1 6 .
Found:
C l,
2 7 .22.
( 2 .4 —D ic h lo ro p h e n y lh y d ra z ono)-p r o p io n ic Acid
1 0 .6 g (0 .0 5 m ole) o f 2 , 4 -d ic h lo ro p h e p y !h y d ra z in e h y d ro c h lo rid e were
d is s o lv e d i n IpO cc o f w a te r ( c a . 80-90° C) and a s o lu tio n o f 4 .4 g (0 .0 5
m ole) o f p y ru v ic a c id d is s o lv e d i n 40 cc o f w a te r was added w ith s t i r r i n g .
The s t i r r i n g was c o n tin u e d f o r 15^20 m in u te s ; th e m ix tu re was co o led and
th e p r e c i p i t a t e f i l t e r e d o f f .
f in e y e llo w c r y s t a l s .
p o in t:
198-'99° C.
R e c r y s ta H iz a tio n from d i l u t e a lc o h o l y ie ld e d
The y i e l d was 4 .8 g (39% of t h e o r e t i c a l ) .
M eltin g
No l i t e r a t u r e v a lu e was found.
A n a ly s is .. C a lc u la te d f o r C9HgN2 O2C ^ :
N e u tr a liz a tio n e q u iv a le n t.
C a lc u la te d :
C l, 2 8 .7 0 .
Found:
2 47. 1 ; found:
C l, 2 8 .6 1 .
2 4 7 .9 .
E th y l j6 - ( p - c h lo r o a n ilin o ) - a c r y la te
The p ro c e d u re was th e same as t h a t used f o r th e p r e p a r a tio n of e th y l
^ L a n ilin d c r y la te , e x ce p t h y d ro c h lo ric a c id was used i n p la c e o f a c e tic
a c id .
The o i l which formed upon a d d itio n o f th e e n o la te s o l i d i f i e d on
c o o lin g .
The s o l i d was r e c r y s t a l l i z e d from a lc o h o l t o g iv e a brown pow der.
The y ie ld was 4 .1 g 36% of t h e o r e t i c a l ) .
sh arp r a n g e ) .
M eltin g p o i n t :
No l i t e r a t u r e v a lu e was fo u n d .
.67-70° C (no
- 44 -
A n a ly s is . . C a lc u la te d f o r G10H12NO2 C l:
C l, 1 6 .7 6 .
Found:'. C l, 1 6 .6 8 .
E th y l a-P h e n y lh y d ra z o n o p ro p io n a te
2 6 .7 g (0 .1 5 mole) of a-phenyIh y d raz o n o p ro p io n ic a c id , 150 cc o f
a b s o lu te a lc o h o l and 5 g o f c o n c e n tra te d s u l f u r i c a c id w ere p la c e d i n a f l a s k
and re flu x e d f o r 4 .5 h o u rs .
o f c o ld w a te r.
The s o lu tio n was co o led and poured in to 1000 cc
The o i l which form ed s o l i d i f i e d w ith s t i r r i n g .
was f i l t e r e d o f f and r e c r y s t a l l i z e d from d i l u t e a lc o h o l.
a s la r g e y e llo w p ris m s .
p o in t:
192° C.
The s o lid
The e s t e r s e p a ra te d
The y i e l d was 2 4 .3 g (79% of t h e o r e t i c a l ) .
M e ltin g
F is h e r (32) g iv e s t h e m e ltin g p o in t a s 192° C.
C L -( p-B rom ophenylhydrazono)-propionic Acid
1 1 .2 g ( 0 .0 5 mole) o f p-brom opkeny!hydrazine h y d ro c h lo rid e were d is s o lv e d
i n 50 cc o f d i l u t e a lc o h o l.
To t h i s s o lu tio n was added a s o lu tio n o f 4 .4 g
(0 .0 5 mole) o f p y ru v ic a c id d is s o lv e d i n 50 cc of w a te r.
h e a te d t o b o ilin g and r e f lu y ed f o r 10 m in u te s .
n e e d le s s e p a r a te d .
The m ix tu re was
On c o o lin g , lo n g l i g h t brown
The p ro d u c t was r e c r y s t a l l i z e d from d i l u t e a lc o h o l t o
y i e l d 7 .3 g (57% of t h e o r e t i c a l ) o f l i g h t yellow -brow n n e e d le s .
p o in t:
184° 0 .
M eltin g
B alb ian o (6) g iv e s th e m e ltin g p o in t a s 182° C.
N e u tr a liz a tio n e q u iv a l e n t.
C a lc u la te d f o r CgH^N2 Q2B r:
2 5 7 .I j found:
2 5 7 .2 ,
d - ( p -N itro p h en y lh y d raz ono)-p r o p io n ic Acid
7 .6 g (0 .0 5 m ole) o f p -n itro p h e n y Ih y d ra z in e and 4 .5 cc o f c o n c e n tra te d
h y d ro c h lo ric a c id were p la c e d i n a f l a s k and 50 cc of w a te r w ere added.
The m ix tu re was h e a te d u n t i l s o lu tio n was co m p lete.
To t h i s s o lu tio n a
s o lu tio n o f 4 .4 g (0 .0 5 mole) o f p y ru v ic a c id d is s o lv e d i n 50 cc of w a ter
was added w ith sh a k in g .
The m ix tu re s o l i d i f i e d to a g e l and th e s o lid was
I
(
- 45 —
f i l t e r e d o f f and r e c r y s t a l I i zed from d i l u t e a lc o h o l.
The hydrazone sep a—: '
r a te d a s a y e llo w powder (composed o f m ic ro sc o p ic n e e d le s ) .
1 0 .2 g (91% o f t h e o r e t i c a l ) .
M e ltin g p o in t:
219° C.
The y ie ld was
Hyde (52) g iv e s
th e m e ltin g p o in t a s 219- 20° C.
N e u tr a liz a tio n e q u iv a l e n t.
C a lc u la te d :
2 2 3 .2 ; fo u n d :
2 2 3 .9 .
Z - M tr o - 1 , 4-dim ethoxybenzene (37)
41 g o f I , 4-dim ethoxybenzene w ere added t o 500 cc o f w a ter and t h e
m ix tu re was h e a te d w ith s t i r r i n g u n t i l t h e e th e r had m e lte d (c a . 50° C ).
added and th e te m p e ra tu re was h e ld betw een 50-55° C.
The y e llo w o i l which
formed g ra d u a lly s o l i d i f i e d and a f t e r th e m ix tu re had c o o led t o room tem ­
p e r a tu r e , th e y e llo w s o l i d was f i l t e r e d o f f and r e c r y s t a l l i z e d from 50-60%
a lc o h o l.
The p ro d u c t s e p a ra te d a s l a r g e , b r ig h t y e llo w n e e d le s .
y i e l d was 53.6 g (99% of t h e o r e t i c a l ) .
M e ltin g p o in t:
7 1 ,5 ° C.
The
Haberman
(37) g iv e s th e m e ltin g p o in t a s 70-71° C.
2 -N itro -4 -m e th o x y p h e n o l
(55)
40 g o f 2 -n itro -l,4 -d im e th o x y b e n z e n e were p la c e d i n a 1500 ml f l a s k
and a s o lu tio n o f 80 g o f NaOH d is s o lv e d i n 700 cc o f w a te r was added.
The m ix tu re was r e f lu x e d f o r 60 h o u rs and 400 cc o f w a te r were added.
The
m ix tu re was steam d i s t i l l e d u n t i l no f u r t h e r u n r e a c t d ib ith e rd d is t i l l e d
o v e r.
The r e s id u e was c o o led t o room te m p e ra tu re and a c i d i f i e d w ith con­
c e n tr a te d h y d ro c h lo ric a c id .
The c o lo r changed from a deep re d t o orange
and c o n s id e ra b le s o lid s e p a r a te d .
l i z e d from 50% a lc o h o l.
p ris m s .
The s o l i d was f i l t e r e d o f f and r e c r y s t a l ­
The phenol c r y s t a l l i z e d a s l a r g e , b r i l l i a n t orange
The y ie ld 'w a s 32 g ( 87% of t h e o r e t i c a l ) .
M e ltin g p o in t: 7 9 .2 ° C.
— 46 —
Kauffman and F r i t z (55) g iv e th e m e ltin g p o in t a s 80° C.
2 -Mi t r o-4-me th oxyphenoxyac e t I c Acid
20 g of 2 -n itro -4 -m e th o x y p h e n o l, 12 ,3 g o f c h lo r o a c e tic a c id and 21 g
o f 50% sodium h y d ro x id e were p la c e d i n a 500 ml round-bottom f l a s k and 100
cc o f w a te r was ad d ed .
The f l a s k was a rra n g e d f o r d i s t i l l a t i o n and th e
m ix tu re slow ly d i s t i l l e d u n t i l th e re s id u e was n e u tr a l t o litm u s (c a . 2
h o u rs; 60-70 cc w a te r d i s t i l l e d o v e r ) .
H a lf lth e above- q u a n t i t i e s o f a l k a l i
And b k lb r o a c e tic a c id and 50 cc o f w a te r were th e n added and th e s o lu tio n
d i s t i l l e d a g a in u n t i l n e u t r a l .
A fte r s tr o n g ly a c id if y in g w ith h y d ro c h lo ric
a c id th e s o lu tio n was c o o le d , p r e c i p i t a t i n g th e a c id . . The compound was
r e c r y s t a l l i z e d from a lc o h o l.
g o ld e n -y e llo w n e e d le s .
The y i e l d b ein g 13 g (U8% o f t h e o r e t i c a l ) o f
M e ltin g p o in t:
149-150° 0 .
No l i t e r a t u r e v a lu e was
fo u n d .
A n a ly s is .
„
C a lc u la te d f o r C^HgNO^:
N, 6 .1 7 .
N e u tr a liz a tio n e q u iv a l e n t. . C a lc u la te d :
Found:
2 2 7 .2 ;
6 .2 1 .
fo u n d :
2 2 6 .8 .
- 47 -
V.
DISCUSSION OF EXPERIMENTAL RESULTS
The d a ta f o r 17 compounds a re l i s t e d i n t a b l e I I .
The v a lu e g iv e n i s
th e p e rc e n ta g e in c r e a s e ( o r d e c re a se ) r e l a t i v e t o th e grow th in c re a s e o f
th e c o n tr o l.
W ith a s ta n d a rd d e v ia tio n o f 0.1 4 u n i t s and an av erag e c o n tr o l
grow th o f 0 .4 9 u n i t s , any v a lu e l e s s th a n ±; '29% could n o t be c o n sid ered
s ta t i s t i c a l l y s ig n ific a n t.
The v a lu e o f th e s ta n d a rd d e v ia tio n (0 .1 4 u n i t s )
i s an a v erag e o f th e v a lu e s f o r a l l of t h e c o n tro l ru n s .
C a lc u la tio n o f th e
r e l a t i v e p e rc e n ta g e in c r e a s e o r d e c re a se was made u sin g t h e av erag e v a lu e
o f th e c o n tr o ls ru n d u rin g th e t e s t i n g o f t h e compound i n q u e s tio n .
I t is
r e a l i z e d t h a t t h e a ss ig n e d v a lu e o f 29% f o r s t a t i s t i c a l s ig n if ic a n c e i s n o t
a r i g i d one, b u t v a ria n c e from t h i s v a lu e would no t b e g r e a t . , I n a review
by Wain (116) i t i s s ta t e d t h a t w ith th e u se o f th e Avena c o le o p t ile s t r a i g h t
grow th t e s t , any grow th d e v ia tio n i n e x c e ss o f 5% of th e c o n tr o l may be con­
s id e re d s i g n i f i c a n t .
T e s tin g a l l o f th e compounds a t once would have been
im p r a c tic a l due t o th e la r g e number o f c o le o p t ile s e c tio n s w hich would be
r e q u ir e d .
T a b le s IV and V a r e t y p i c a l exam ples o f th e m easurem ents re c o rd e d
d u rin g a t e s t r u n .
T a b le I I I g iv e s th e m easurem ents o b ta in e d fo r th e con­
t r o l s ru n s im u lta n e o u s ly w ith th e s e two compounds.
The a c t i v i t y o f th e s u b s tit u te d phenyIh y d ra z o n o a c e tic a c id s i s g r e a te r
th a n t h e co rre sp o n d in g p ro p io n ic a c id d e r i v a t i v e s .
T h is would in d iq a te ,
a s i n th e p h e n o x y a lk y !c a rb o x y lic a c id s , t h a t a lp h a s u b s t i t u t i o n d e c re a se s
a c tiv ity .
There a r e , how ever, r e s u l t s w hich a re n o t i n common w ith any o f
th e o th e r s e r i e s o f grow th r e g u la to r s :
(a ) th e p a re n t a c id s show th e g r e a t ­
e s t a c t i v i t y , (b) e th y l 2 , 4 -d ic h lo ro p h e n y Ih y d ra z o n o a c e ta te i s a c ti v e w h ile
t h e f r e e a c id behaves a s an i n h i b i t o r a t a l l t e s t e d c o n c e n tra tio n s , and (c )
48 —
(p -b ro m o p h en y lh y d razo n o )-p ro p io n ic a c id and CU-( p -c h lo ro p h e n y lh y d ra z ono) p ro p io n ic a c id a re h ig h ly in h ib ito r y , a t a l l c o n c e n tr a tio n s .
The s u b s tit u te d e h ty l a c r y l a t e s e r i e s and t h e S c h if f b a se s o fy d -k eto
a c id s may show prom ise w ith f u r t h e r s tu d y .
The need f o r more in v e s t ig a ti o n
on th e two m ain .,series i s a ls o a p p a r e n t.
The d a ta o b ta in e d f o r 2 -n itr.o -4 -m eth o x y p h en o x y acetic a c id , a lth o u g h
n o t s t a t i s t i c a l l y s i g n i f i c a n t , in d ic a te s t h a t t h e r e may be some com bination
o f s u b s tit u e n ts and p o s itio n s which co u ld p a r a l l e l th e e f f e c t s o f th e 2 ,4 d ic h lo r o s u b s t i t u t i o n ; 2 -m e th o x y -4 -n itro p h e n o x y a c e tic a c id may prove t o be
more p re m is in g .
S in ce i t i s p o s s ib le f o r th e p h e n y lh y d ra zo n o -ac e ti c and p ro p io n ic
a c id s to e x i s t a s c i s - t r a n s iso m e rs, some o f t h e d is c r e p a n c ie s n o te d may be
a t t r i b u t e d t o th e p re se n c e o f d i f f e r e n t is o m e ric s t a t e s .
As h as been p o in te d
o u t, only th e c is - is o m e r o f p re v io u s ly in v e s t ig a te d c i s - t r a n s p a i r s e x h ib it
a c t i v i t y ( th e g e o m e tric a l s tr u c tu r e b e in g known i n th e s e c a s e s ( 106, 111) .
The phenylhydrazonoacet’i c a c id p re p a re d i s th e h ig h e r m e ltin g o f th e two
r e p o rte d isom ers (21) and i s p ro b a b ly th e tr a n s - is o m e r ( i n most c ase s th e
tr a n s - is o m e r h as th e h ig h e r m e ltin g p o i n t ) „
I f th e p h e n y lh y d ra zo n o a c etic
a c id does have th e tr a n s - c o n f ig u r a tio n , t h i s r e s u l t i s i n d i r e c t c o n tr a s t
t o t h e f in d in g s c ite d a b o v e.
L i t t l e work h a s been done to w ard s e s ta b lis h in g
th e g e o m e tric a l s t r u c t u r e o f th e s e compounds; such in fo rm a tio n would be o f
g r e a t im portance i n f u tu r e i n v e s t ig a ti o n s o f th e s e compounds.
T a b le I I
P e rc e n ta g e Growth R e la tiv e t o C o n tro l
C o n ce n tra tio n x IO4 m olar
Compound T e sted
0 .1
2 ,4 -D ic h lo ro p h e n o x y a c e tic ' acid",r
P h e n y lh y d ra z o n o ac e tic acid"'
fit-phenyIh y d ra z o n o p ro p io n ic a c id "
oL- (2 -ch lo ro p h e n y Ih y d raz ono) - p r o p i onie a c id ''
cb-(4 -c h lo ro p h e n y lh y d ra z ono) -p ro p io n ic a c id "
oL - { 2 3 4 - d i ch lo ro p h eny Ihydraz ono) - p r o p i o n ic .-a c id ""
c£-(4-brom ophenylhydraz ono) -p ro p io n ic acid"5^
dr (4-m ethylphenyIh y d raz ono) -p r o p io n ic a Cicf'
CL-(4 - n i t rophenyIh y d ra z ono) -p r o p io n ic a cid "
E th y l 4 -c h lo ro p h e n y lh y d ra z o n o a c e ta te 7r
E th y l 2 , 4 -d ic h lo ro p h e n y Ih y d ra z o n o a c e ta te"
2 , 4 -lic h lo r o p h e n y Ih y d ra z o n o a c e tic acid'" j.
E th y l 2 , 5 -d ic h lo ro p h e n y lh y d ra z o h o a ce ta te"
E th y l /S -( 4 - chlorophenylam ino) - a e r y I a t
E th y l /S’- (4-m ethylphenyIam ino) - a c ry la te "
C t-P henylim inopropionic a c id
2 -N itro -4 -m e th o z y p h e n o z y a c e tic a c id
*
#
The sodium o r p o ta ssiu m s a l t was u sed .
An em ulsion w ith MCOl H-86c was u sed .
+96
+51
-8 9
-7 4
O
-17
0 .5
1 .0
. 2 .0
5 .0
+6?
+95
-1 8
-12
-90
-20
-7 5
-24
-1 9
- 7
+63
-63
+29
+6
14
+31
-28
+98
+33
-16
-10
+60
-16
—48
-27
-82
•- 6
-72
-2 7
-1 4
-4 4
+24
-85
- +2
+20
- 8
16
+22
+19
-6 8
-14
-78
-32
-12
-10
+22
-74
+16
+39
- 3
3
- 7
-82
-75
-8 8
’
- 50 T ab le I I I
.GxiOVfbh o f :Avena i.Co le o p tlle .,Sefetions .I n : D i s t i l l e d J ^ a te r .'
R e la tiv e Length
C o n tro l Number
I
8 .2 8
8.20
8 .1 8
8 .4 0
8'.25
8 .1 0
8.25
8 .2 0
8,42
8 .1 5
TOTAL
Average
S tan d ard D e v ia tio n
I
2
8 .3 0
. 8 .3 0
8.4 0
8 .1 0
8 .1 5
8.2 5
8 .2 0
8.45
8 .3 0
8 .2 0
3
.
8 .3 0
8.32
8.4 5
8 .1 0
8 .3 0
8.32
8 .1 0
8 .3 0
8.3 5
8.43
248.05
8 .2 7
0 .1 1
- 51 -
T ab le IV
Growth o f Avena C o le o p tlle S e c tio n s i n S o lu tio n s
o f Sodium P h e n y lh y d ra z o n p ac e tate
C o n c e n tra tio n x lcA m olar
2 .0
M easured le n g th o f
c o le o p t ile s e c tio n s
8 .3 5 ■
7 .7 0
8 .0 0
8 .3 0
8 .3 0
■8.05
7 .8 8
8 .1 0 .
8 .1 5
8 .8 7
.
1 ,0
0 .5
0 .1
-8 .5 2
8 .3 0
8 .1 0
8 .6 0
8 .7 0
8.7 5
8 .5 0
'8 .5 0
8.30
8.45
8 .4 0
9 .4 0
9 .8 0
8 .2 0
9 .0 8
9.0 0
8.1 3
9 .0 0
9 .10
8 .4 0
9.00
8.56
8.70
8.7 0
8.9 0
8 .90
8 .20
9.30
8 .6 0
9 .8 0
1
T o ta l
81.70
84.72
8 8 ,5 1
88.66
Average
8 .1 7
8 ,4 7
8 .8 5
8.8 7
D e v ia tio n frorti c o n tr o l
- 0 .1 0
+ 0 .2 0
+ 0 .5 8
+0,60
-1 6
+33
+95
+98
R e la tiv e p e rc e n ta g e
iric re a s e o r d e c re a s e
-•
- 52 T able V
Growth o f Avena C o le o p tile S e c t i o n s .i n S o lu tio n s
o f 0-P henylim in o p ro p io n lc Acid
C o n c e n tra tio n 'x lcA m olar
M easured:,length o f
c o le o p tile c s e c tio n s
’2 .0
1 .0
0 .5
0 .1
8 .2 0
8 .1 0
8 .6 0
8 .4 5
8 .1 0
9 .6 0
8 .6 0
8 .1 0
8 .8 9
7.9 2
8.23
8.2 0
8.9 5
8 .2 5
7 .9 5
8 .4 0
8 .25
8 .0 5
8 .2 0
8.2 0
. 8 .3 0
8 .2 0
8 .1 5
8,55
7 .9 0
8 .0 0
8.10
7 .9 0
' 8.40
•—
8 .7 0
8 .5 0
8 ,3 0 .
8.12
■ 8 .0 0
7 .9 8
8 .20
7-95
8 .4 6
.
8 .2 8
T o ta l
73.50
8 2 .4 9
84.60
82.68
Average
8 .1 7
8 .2 5
8 .4 6
8 .2 7
D e v ia tio n from c o n tr o l
-0 .1 0
-,0.02
•6,0.19
0 .0 0
R e la tiv e p e rc e n ta g e
in c r e a s e o r d e c re a se
-16
-3
+31
0
* C o le o p tile s e c tio n d ead.
- 53 V I.
ACKNOWLEDGMENT
•
The a u th o r w ishes to ta k e t h i s o p p o rtu n ity to e x p r e s s .h is s in c e re
' a p p r e c ia tio n and th a n k s t o D r. L aurence 0 . B in d e r, J r . f o r h i s p e rs o n a l
gu id ance and i n s p i r a t i o n d u rin g t h i s r e s e a r c h , t o Mr. B arto n E . Hahn o f
th e B otany D epartm ent and t o th e s t a f f o f t h e C hem istry D epartm ent f o r
t h e i r many h e lp f u l s u g g e s tio n s .
- 54 -
V II.
LITERATURE CITED AND CONSULTED
1 . Abe rg , B ., A rk iv Kemi,
549 (1951)
2 . A d d ic o tt, F .T ., P la n t P h y s io l., 18, 270, (1943)
3 . A lex an d er, E .R ., ,fP r in c i p le s o f Io n ic O rganic R e a c tio n s " , John W iley &
Sons, I n c . (1953)
4 . Amac k e r, T .M ., B r i t . 6 5 3,452, C.A. ^6 ; pl0 3 8 (1952)
5. Ashby, W .C., B o ta n . G az., 112, 237 .(1 9 5 1 )
6 . B alb in an o , L ., B e r ., 20, 290 (1897)
7 . Bam berger, E ., B e r ., 21, 582 (1898)
8 . B e i l s t e i n , V ol. X II, p 5 l6
9 . B e i l s t e i n , V ol. XV, p l0 5 , 1 s t s u p p l.
1 0 . B e i l s t e i n , V ol. XV, p l6 0 , 1 s t s u p p l,
1 1 . B e i l s t e i n , V ol. ,XV, p523
12. B e n tle y , J .A . and S . J . H ousley, J . E x p t l . B o ta n ., 2» 393 (1952)
1 3 . B e n t le y ,. J . A . , N a tu re , 165. 449 (1950)
1 4 . B isso n , C .S. and H.A. J o n e s , P la n t P h y s io ).., 7 , 91 (1932)
1 5 . B onner, D.M., B o t. G a z ., 100. 200 (1938)
1 6 . B onner, J . , J . Gen. P h y s io l. , 1%, 63 (1933)
1 7 . B onner, J . , Amer. J . B o t., 2^, 323 (1949)
1 8 . B onner, J . and S.G. Wildman, A rch. B iochem ., 10, 497 (1946)
1 9 . Bourne * M.C, a n d . L . Young, Biochemj. J . j 2 8 , 8 0 3 ’(1934)
2 0 . B ru n e i, A. and R. E ch ev in , Compt. r e n d ., 207. 592 (1938)
2 1 . Busch, M ., F . A c h te r f e ld t and R. S e u f e r t , J . p r a k t . Chem., g2, I (1915)
2 2 . C hattaw ay, F.D . and R .C .T . Evans, J . Chem. S o c ., 6%, 848 (1896)
2 3 . C h a ttaway, F.D . and C .F.B . P e a rc e , J . Chem. S o c ., 107. 32 (1915)
2 4 . C hattaw ay, F.D . and J . B e n n e tt, J . Chem. S o c ., 130. 2850 (1930)
- 55 -
2 5 . Coleman, G.H*, O rganic S y n th e se s, Vol'. I I , p71
2 6 . C u rtin s , T ,, B e r .,
1072 (1912)
2 7 . deBruyn, Rec.. t r a v . c h im ., ^6 , l 6 l (1917)
2 8 . E v e n a rl, M ., B o ta n . R evs. Cambridge P h i l . S o c ., 1£, 153 (1949)
2 9 . F a w c e tt, C .H ., D .J . O sborne, R .L. Wain and R;Dt W alker, Ann. A ppl.
B i o l ., 4 0 , 231 (1953)
3 0 . F i e s e r , L . F . , "E xperim ents i n O rganic C h em istry ", 3 rd e d itio n , D.C.
H eath and C o., 1955, p . 265
3 1 . F in d la y , S .P . and G. D ougherty, J . B io l. Chem., 183. 361 (1950)
3 2 . F is h e r , E . B e r ., 17, 578 ( I 884)
33« F o s te r , R . J . , D.H. McRae and J . B onner, P ro c . N a tl. Acad. S c i . U .S .,
28, 1014 (1952)
3 4 . F red g a, A ., and M. M a te ll, A rkiv Kemi, 2» 429 (1951)
35» Gordon, S.A . and R .P . W eber, P la n t P h y s io l. , 26, 192 (1951)
3 6 . G reenberg, D.M. and T. W innick, Ann. Rev. B iochem ., 1/^, 31 (1945)
3 7 . Habermann, J . , B e r ., 11, 1037 (1878)
3 8 . H ale and B r itto n , I n d . Eng. Chem., 20, l l 4 (1928)
39» H ansch, -C. and R.M. M uir, P la n t P h y s io l. 25, 389 (1950)
4 0 . H ansch, C ., R.M. M uir and R .L. M etzenberg, P la n t P h y s io l. , 26, 812 (1951)
4 1 . H ayes, N.V. and F .E .K . B ranch, J . Am. Chem. S o c ., 6^, 1555 (1943)
4 2 . H essayon, D .G ., N a tu re , 169. 803 (1952)
4 3 . H e w itt, J . T . j J . Chem. S o c ., 62, 873 (1893)
4 4 . H e w itt, J . T . , J . Chem.
S o c ., 22» 209 (1891)
45» H i r s t , E .L ., F.A . I sherwood, M.A. Jermyn and J.K .N . J o n e s , J . Cheme
S o c ., 20, 182 (1949)
.
.
.
.
4 6 , H itc h c o c k , A.E. and P.W. Zimmerman, P r o c . Am. S oc. H o r t. S c i . , 45, 187 (1944)
— 56 —
47• H itc h co c k , A„E. and P.W. Zimmerman, C o n trib s . Boyce Thompson I n s t . ,
1%, 35 (1952)
4 8 . Hoffman, O .L ., S.W. Fox and M .¥. B u llo c k , J . . B io l. Chem., 196. 437 (1952)
4 9 . H ollem an, Chem. R e v s ., I , 187 (1925)
50. H o lle y , R.W., F .P . B oyle, H.K, D u rfe e , and A.D. H a lle y , A rch. B iochem .,
22, 192 (1951)
51. Hollman and DeMooy, R ec. t r a v . ch im ., 2 1 , 19 (1915)
52. Hyde, E ., B e r ., 2 2 , 1815 (1899 )
53. Ing,. H .R .j M.C. Bourne and L. Young, Biochem J . , 2 8 , 809 (1934)
54. J o n e s , E .R .H ., H.B. H en b est, G .F. Sm ith and J .A . B e n tle y , N atu re, 169«
485 (1952)
5 5 . Kauffmann and F r i t z , B e r ., 4 2 , 1214 (1910)
56. K e lly , S . , Am. J . B o ta n ., 2 2, 335 (1936)
57. K ing, L . J , , J .A . Lambrech and T .P . F in n , C o n trib s . Boyce Thomson I n s t . ,
16, 191 (1950)
58. Koepfl i , Thimann and W ent, J . B io l. Chem., 122. 763 (1938)
59. Kogl, F ., N a tu rw isse n sc h a ft e n , 2j>, 465 (1937)
60; Kogl, F . and D .G .F.D . K osterm ans, H o p p e-S ey ler1s Z . P h y s io l. Chem,,
235, 201 (1935) ( c f . r e f . I l l )
6 1 . L a rse n , P . , Dank. B otan. A rk iv ., 11, I (1944)
62. L a rse n , P . , Am. J . B otany, 26, 32 (1949)
6 3 . L a rse n , P*, Ann. Rev. P la n t P h y s io l.,. 2 , 169 (1951)
6 4 . L e a p er, J.M .F . and J .R . B ishop,
B o t. G a z., 112. 250 (1951)
6 5 . L eFevre, J . Chem. S o c ., 1933, 980.
6 6 . L in s e r , H ., P la n ta , 22, 377 (1951)
6 7 . Luckwi 11, L .C ., J . H o r t. S c i . , 24., 19 (1948)
6 8 . M a te ll, M1, A rkiv Kemi, 4., 473 (1952)
- 57 -
69. M in a rik , C .E ., D. Ready, A.G. Norman, H .E. Thompson and J . F . Owens,
B o ta n . G a z., 112, 135 (1951)
7 0 . Moyer,
W.W., UiS f 2 ,4 1 6 ,1 9 8 , O.A. IfcIs p3251i (l9 4 7 )
7 1 . M u ir, R.M, , C. H. Hansch and A. H. G a llu p , P la n t P h y s io li , Zlt f 359 (1949)
72. M uir, R.M. and C.H. H ansch, P la n t P h y s io l., 2 6 , 369 ( 1951)
73.. M uir, R.M. and C.H. H ansch, P la n t P h y s io l., 2 8 , 218 (1953)
74. M uir, R.M ., Annual Rev. P la n t P h y s io l., 6, 157 (1955)
7 5 . M u lle r, E ., B e r ., 4 7 , 3021 (1914)
7 6 . Norman, A.G. and R .L. W ein trau b , N a tl. R esearch C o u n cil"N a t. "Acad. S c i .
(UeS .) ,. Chem. B io l. C o o rd in a tio n C e n te r, Pub. ( f i r s t symposium, p . 4 6,
7 7 . O sborne, D .J . and R1L i W ain, S c ie n c e , 1 14. 92 ( 1951)
7 8 . O sborne, D .J . and R .L . W ain, J . H o rt. S c i . , 2 6 , 66, 317 ( 1951)
79. O sborne, D .J ,, R .L. Wain and R.D. W alk er, J . H o rt. S e i . , 2%, 44 (1952)
8 0 . O verbeek, J . van and F iW. W ent, B o ta n . G a z.,
22 (1937)
8 1 . P a le g , L.G . and R.M. M uir, P la n t P h y s io l., 2%, 285 (1952)
8 2 . Pechmann, Hi , B e r ., 2j_, 1047 (1892)
8 3 . P r ic e , C .C ., O rganic R e a c tio n s , 2* I (1946)
84« Ready, D. and V.Q. G ra n t, B o ta n . G a z., 109. 39 (1947)
85. Redemann, G.T», S.H . W ittw er and H.M-. S e l l , A rch. Biochem. B io p h y s.,
2 2, 80 (1951)
8 6 . Reed and O rton, J . Chem. S o c ., $ l f 1553 (1907)
8 7 . S c h n e id e r, C .L ., Am. J . B otany, 2 £ , 201 1942)
8 8 « Sm ith, M .S. and R .L . W ain, P r o c . R oyal S oc. (London), (B) 139, 118
(1951)
89. Sm ith, M .S ., R .L . Wain and F . Wightman, N a tu re * 169. 883 (1952)
90. S te k o l, J .A ., J . B io . Chem., 112,
279 (1936)
- 58 -
91. S te v e n s, F . J . , J . S c i . , 2 2 , 79 (1947)
92. S te v e n s, F .J . and S.W. Fox, J . Am. Chem. S b c ,, 2 2 , 2263 (1948)
93. Stowe, B .B ., and K.V. Thimann, A rch. Biochem. B io p h y s. , 51, 499 (1954)
94. S tr a u s , F . and W. V oss, B e r ., £ £ , 1681 (1926)
95. Swanson, C .P ., B o ta n . G a z., 107. 507 (1946)
96. Synerholm, M.E, and P.W. Zimmerman, C o n tr ib s . Boyce Thompson, I n s t . . 14
91 (1947)
—
97. Tang, X.W. and J . B onner, A rch. B iochem ., 1 ^, 11 (1947)
98. Tang, Y.W. and J . B onner, Am. J . B o t.,
570 (1948)
99. T e a s, H .J . and A. Newton, P la n t P h y s i o l ., 2 6 , 494 (1951)
1 0 0 . Thimann, K.V. and C.L. S c h n e id e r, Am,. J . B otany, 25 , 270 (1938)
101» Thimann, K .V ., " P la n t Growth S u b s ta n c e s " ,
W is e ,, 1951.
F . Skoog, E d ,, M adison,
102. Thimann, K .V ., P la n t P h y s io l., 2%, 392 (1952)
103. Thimann, K .V ., A rch, Biochem. B io p h y s. 44.* 242 (1953)
)
104. Thimann, K.V. and W.D-.
,
"
B onner, Amer. J .
'
'
'
B o t., £ £ , 271 (1948)
105. Thimann, K.V. and W.D. B onner, Amer^ J . B o t., ^ 6, 214 (1949)
106. V e ld s tr a , H», Enzym ologia, 11, 97 (1944)
107. V e ld s tr a , H ., Enzym ologia, 11, 137 (1944)
108. V e ld s tr a , H ., and H.L.
B o o ij, Biochim B io p h y s. A c ta , 2 , 278 (1949)
109. V e ld s tr a , H ., and C. van de W este rin g h , R ec. t r a v . c h im ., 70, 1.11.3
(1951)
"
H O . V e ld s tr a , H ., and C. van de W este rin g h , R ec. t r a v . c h im ., 21* 318
(1951)
111. ,.V e ld stra, H ., and C. van de W este rin g h , R ec. t r a v . c h im ., 2 1 , 1127
(1951)
1 12. V e ld s tr a , H ,, R ec. t r a v . chim ., 2 i , 15 (1952)
. - 59-
113. V e ld s tr a , H ., Ann. Rev. P l a n t 'P h y s i o l . , I±t . 1 51 (1953)
114. V l i t o s , A .J . , C o n tr ib s . Boyce Thompson I n s t . , 1%, 127 (1953)
115. W ain, R .L ., J . S c i. Food A g r., p . 101 (1951)
1 16. W ain, R .L ., N a tu re , 1%2, 710 (1953)
117. W ain, R .L. and F . Wightmari; -Ann. A p p l. B i o l ,, 42» 244 (1953)
118. W eintra u b ; R .L ., J.W . Brown, J .C . N ickdrson," and K.<N. T a y lo r, B o tan .
' G a z., I l l , 348 (1952)
' '
119. W ein trau b , R .L ., J $ . Brown and J.A.. T hrone, Ag and Food, 2 , 996 (1954)
120. Went, F.W ., B u ll. T o rre y B o ta n , C lu b ,. 66j
(1939)
121. W ent, F .W .,,A rch. B iochem ., 2 0, 131 (1949)
122. W ieland, T ., E . F is c h e r arid F . Moewus, Ann. Chem. J u s t i s L ie b ig s , 559.
47 (1948)
123. Wildman, S.G-. and J . B onner, Am. J . B o t., 21, 740 (1948)
124. W illg e r o d t, B e r ., 12, 763 (1879)
125. W is lic e n u s , W., B e r ., 20, 2931 (1887)
126. Z e t t e l , T ., B e r ., 2 6 , 2472 (1893)
127. Zimmerman,. P.W. and F . W ilcoxon, C o n tr. Boyce Thompson I n s t . %, 209
(1935)
128. Zimmerman, P.W. and A.E. H itc h co c k , C o n trib . Boyce Thompson I n s t . , 12.
321 (1942)
129. Zimmerman, P.W ., T o rre y a , 42.? 98 (1943)
130. Zimmerman. P.W-. and A.E. H itc h co c k , P r o c . Am. S o c. H o r t. S c i . , 41»
353 (1944)
131. Zimmerman, P.W. and A.E. H itch co ck , C o n tr. Boyce Thompson I n s t . , 15«,
209 (1951)
1 3 2 . Zimmerman, P.W-. and A.E. H itch co ck , C o n tr. Boyce Thompson I n s t , , 16,
209 (1951)
' ,
- zi
-'jV-/
13 3 . Zimmerman; P.W ., A .E. H itchcock and E.A. P r i l , C o n tr. Boyce Thompson
I n s t . , 16, 419 (1952)
118855
MONTANA STATE UNIVERSITY LIBRARIES
Il i l l III I IEI I
C
762 K)01
2 2
.
I
3578
Gl2n
c o p .2
118855
^ N e r ^ l^ ^ g r o w th -r e g u la to r s
AUG 2 3 'SI
NAWl 3 S f
I
>
U
-■ '
y
K 0 / L
wm
P
118855
/tfr