Walter Anderson Caldwell
A Thesis Submitted for the Degree of PhD at the
University of St Andrews
1934
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T I 'I ,
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A S tu d y o f th e M e th y l, A c e t y l , and
B e n z o y l D e r i v a t i v e s o f S t a r c h ,
A THESIS PRESENTED FOR THE DEGREE
o f
DOCTOR OP PHILOSOPHY
O f
THE UNIVERSITY OP ST . ANDREWS
by
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TRAINING AND EXPERIENCE IN RESEARCH
I m a t r i c u l a t e d i n th e U n i v e r s i t y o f 'G la s g o w i n
O ctob er 1 9 2 7 , and p u r s u e d the r e c o g n i s e d c o u r s e f o r th e d e g r e e
o f B . S c . i n A p p l i e d C h e m is tr y , o b t a i n i n g th e d e g r e e w i t h F i r s t
C la s s Honours i n 1 9 3 1 . D u rin g my f i n a l y e a r a t Glasgow I
c a r r i e d o u t an o r i g i n a l r e s e a r c h under P r o f e s s o r P . J . W i l s o n ,
D . S c . , th e r e s u l t s o f w h ic h were p u b l i s h e d i n t h e J o u r n a l o f
the C h em ical S o c i e t y , 1 9 3 2 . V o l . 2 3 2 , p . 2 0 8 6 .
I was aw arded a M ain ten ance G rant b y th e Department
o f S c i e n t i f i c and I n d u s t r i a l R e s e a r c h i n Septem ber 1931 and
p r o c e e d e d t o t h e U n i v e r s i t y o f S t . Andrews, where th e r e s e a r c h ,
the r e s u l t s o f w h ic h form th e s u b j e c t o f th e p r e s e n t T h e s i s ,
was c a r r i e d o u t un der th e d i r e c t i o n o f P r i n c i p a l S i r James C.
I r v i n e . In S e p te m b e r , 1 9 3 3 , on th e e x p i r y o f th e M ain ten ance
G ra n t, I was aw arded a S e n i o r C a r n e g ie R e s e a r c h S c h o l a r s h i p ,
w h ic h I r e s i g n e d i n F eb ru ary 1934 on o b t a i n i n g a r e s e a r c h
C O N T i i i N T S
Page
INTRODUCTION ... 1
THEORETICAL: A c é t y l a t i o n o f S t a r c h : F i r s t M o d i f i c a t i o n o f t h e B a r n e t t P r o c e s s 18 T reatm en t o f S t a r c h T r i a c e t a t e s ... 19
S e c o n d M o d i f i c a t i o n o f th e B a r n e t t P r o c e s s • • • 20 E s t i m a t i o n o f R e d u c in g M a t e r i a l ... 22
I n v e s t i g a t i o n o f A c t i o n o f A l k a l i ... 24
M é t h y l a t i o n o f S t a r c h b y D iazom ethan e ... 26
H y d r o l y s i s o f p a r t i a l l y m e t h y l a t e d s t a r c h . . . . 28
F u r th e r M é t h y l a t i o n a n d H y d r o l y s i s . . . 30
B e n z o y l a t i o n o f S t a r c h ... 33
P r e v i o u s work on b e n z o a t e s o f S t a r c h ... 35
T reatm en t o f B e n z o a t e s o f S t a r c h and D e x t r i n s . 3 6 F u r th e r D e g r a d a t io n E x p e r im e n ts on S t a r c h B e n z o a te ... 37
C o n t r o l D e g r a d a t io n o f M a lto se ... 40
A n a l y s i s o f D im eth y l G lu c o s e s ... 40
C r y s t a l l i n e M a t e r i a l o b t a i n e d ... 41
S y n t h e s i s o f 2 - 3 - 6 T r ib e n z o y l 4 M o n a c e ty l /3 M ethyl G lu c o s id e ... 42
P r e p a r a t i o n o f 2 - 6 D ib e n z o y l 3 - 4 D i a c e t y l ^ M ethyl G lu c o s id e ... 44
C o n t r o l E x p e r im e n t on G lu c o s e P e n t a b e n z o a t e . . 44
D i s c u s s i o n o f R e s u l t s ... 46
Summary o f R e s u l t s ... 53
EXPERIMENTAL: G e n e r a l Working M e th o d s ; A c e t y l E s t i m a t i o n s ... 55
B e n z o y l a t i o n o f C a r b o h y d r a te s ... 56 Graded H y d r o l y s i s o f a M e t h y la t e d S t a r c h b y
A c id M eth yl A l c o h o l ... 57 S e p a r a t i o n o f M e t h y la t e d G lu c o s e s ... 57
I s o l a t i o n and T reatm en t o f ’’R e s i d u e s ” from
M e t h y la t e d S t a r c h ... 59 T r ea tm en t o f B e n z o a t e s w i t h Hydrobromic A c i d
and G l a c i a l A c e t i c A c i d un der s t a n d a r d
C o n d i t i o n s ... 60 D é b e n z o y l a t i o n o f B e n z o a t e s ... 60
A c é t y l a t i o n o f S t a r c h :
M o d i f i c a t i o n N o. I o f B a r n e t t ' s M e t h o d ... 62 S o x h l e t E x t r a c t i o n s ... 65 F r a c t i o n a l p r e c i p i t a t i o n b y E t h e r o f S t a r c h
T r i a c e t a t e from S o l u t i o n i n C h loroform . . 65 ... From S o l u t i o n i n E t h y l A c e t a t e . . . 66 D e a c e t y l a t i o n o f S t a r c h T r i a c e t a t e ... 67 F u r th e r A c é t y l a t i o n E x p e r im e n ts ... 67 A tte m p t t o D e s t r o y R e d u c in g M a t e r i a l i n
” D e x t r i n A . . . 69 M o d i f i c a t i o n N o. I I o f B a r n e t t ' s Method ... 70 E s t i m a t i o n o f the R e d u c in g Power o f V a r io u s
D e x t r i n s . . . 71 E f f e c t o f A l k a l i on R e d u c in g Power ... 72
T reatm en t o f R e s i d u e s from B a r n e t t A c é t y l a t i o n
P r o c e s s ... 74
M é t h y l a t i o n o f S t a r c h by Diazome th a n e ... 74
H y d r o l y s i s o f P a r t i a l l y M e t h y l a t e d S t a r c h and
S e p a r a t i o n o f H y d r o l y s i s P r o d u c t s . I . . . 79 ... .. H y d r o l y s i s ... I I . . 82 F u r t h e r T reatm en t o f D im e th y l M eth yl and T r i
-m e th y l M ethyl G lu c o s id e F r a c t i o n s ... 85
B e n z o y l a t i o n o f S t a r c h :
A c t i o n o f Hydrobromic A c id and G l a c i a l A c e t i c
A c id on S t a r c h T r i a c e t a t e ... 87 P r e p a r a t i o n and D e g r a d a t io n o f t h e B e n z o a te
o f D e x t r i n A ... 90 P r e p a r a t i o n and D e g r a d a t io n o f t h e B e n z o a te
Page P r e p a r a t i o n and D e g r a d a t io n o f t h e B e n z o a te
o f R ic e S t a r c h ... 92 S h o r t D e g r a d a t io n o f B e n z o a te from Ric& S t a r c h . 93 H y d r o l y s i s o f R e s id u e from S h o r t T r ea tm en t 97 Long D e g r a d a t io n o f B e n z o a te from R ic e S t a r c h . . 98 H y d r o l y s i s o f R e s id u e from Long T reatm en t ... 100 E s t i m a t i o n o f 6 - P o s i t i o n in D im e th y l M ethyl
G lu c o s i d e F r a c t i o n s ... 100 T r ea tm en t o f G lu co se P e n t a b e n z o a t e w i t h H ydro
b rom ic A c id and G l a c i a l A c e t i c A c i d 102 P r e p a r a t i o n and D e g r a d a t io n o f O c t a b e n z o y l
M a lto s e ... 103 N atu re o f C r y s t a l l i n e M a t e r i a l ... 1 04
S y n t h e s i s o f 2 - 3 - 6 T r ib e n z o y l 4 M o n a c e ty l ^
M ethyl G lu c o s id e ... 1 0 4 P r e p a r a t i o n o f 2 - 6 D ib e n z o y l 3 - 4 D i a c e t y l ^
M ethyl G lu c o s id e ... 1 0 5
The f i e l d o f th e C h em istr y o f S t a r c h i s so w id e and
th e c o m p l e x i t i e s i n v o l v e d a r e s o g r e a t t h a t a n y a t t e m p t t o s o l v e
th e problem s o f the c o n s t i t u t i o n o f s t a r c h c a n n o t be s p r e a d o v e r
th e w h ole f r o n t , b u t must n e c e s s a r i l y be c o n f i n e d t o a few c h o se n
p o i n t s o f a t t a c k . The f o l l o w i n g i n t r o d u c t i o n , t h e r e f o r e , i s n o t
p r e s e n t e d as a c o m p le te s u r v e y o f t h e l i t e r a t u r e o f t h e s u b j e c t
b u t m e r e ly a s a résumé^ o f th o s e p a r t s b e a r i n g im m e d ia te ly on the
t o p i c s s t u d i e d , w i t h s p e c i a l r e f e r e n c e to t h e work o f D r. J . K.
R u t h e r f o r d , inasm uch a s th e p r e s e n t t h e s i s i s a c o n t i n u a t i o n and
a m p l i f i c a t i o n o f h e r w o r k .
Pour o f th e main prob lem s c o n n e c t e d w i t h th e c o n s t i t u t i o n
o f s t a r c h may be s t a t e d b r i e f l y a s f o l l o w s ;
-( 1 ) What i s th e p o s i t i o n o f the l i n k a g e s b e tw e e n th e a n h y d r o - g l u c o s e
u n i t s i n th e s t a r c h m o l e c u l e , and how much r e l i a n c e may be
p l a c e d on t h e method o f m é t h y l a t i o n and h y d r o l y s i s i n d e t e r m in
i n g t h e i r p o s i t i o n ?
( 2 ) What i s the e f f e c t o f a c é t y l a t i o n o f s t a r c h a s a p r e l i m i n a r y
t o m é t h y l a t i o n i n c o n s t i t u t i o n a l s t u d i e s ?
( 3 ) Does the m o l e c u l e o f s t a r c h c o n s i s t o f a s m a l l number o f
a n h y d r o - g l u c o s e u n i t s h i g h l y p o l y m e r i s e d or a l a r g e number o f
s u c h u n i t s p o l y m e r i s e d t o a l e s s e r e x t e n t ?
( 4 ) I s s t a r c h , f r e e d from adm ixed compounds, homogeneous i n th e s e n s e
That s t a r c h c o n s i s t s o f a n h y d r o - g l u c o s e u n i t s was shown
by Sttlomon ( 1 ) who was one o f t h e f i r s t w ork ers t o c o n v e r t s t a r c h
q u a n t i t a t i v e l y i n t o g l u c o s e and who fo u n d t h a t h i s r e s u l t s a g r e e d
w i t h the form u la (G, H 0 ^ ) , a r e s u l t s i n c e c o n fir m e d b y many
C /O o
Other w o r k e r s . The a c t u a l p o s i t i o n o f the l i n k a g e s b e tw e e n the
a n h y d r o - g l u c o s e u n i t s w a s , h o w e v e r , d e te r m in e d o n l y a f t e r th e
a p p l i c a t i o n o f th e method o f m é t h y l a t i o n and s u b s e q u e n t h y d r o l y s i s .
The m é t h y l a t i o n o f s t a r c h was f i r s t c a r r i e d o u t by Denham and
Woodhouse ( 2 ) t o whom we owe the u s e o f m e th y l s u l p h a t e a s a m e t h y l
-a t i n g -a g e n t -a p p l i c -a b l e to c -a r b o h y d r -a t e s . L a t e r , I r v i n e and M acdonald
( 3 ) c a r r i e d o u t the h y d r o l y s i s o f a f u l l y m e t h y l a t e d s t a r c h and
o b t a i n e d 2 - 3 - 6 t r i m e t h y l g l u c o s e a s the o n l y c r y s t a l l i n e p r o d u c t .
T his s u g a r , h o w e v e r , may o r i g i n a t e from e i t h e r th e j / o r th e normal
form o f g l u c o s e and the l i n k a g e s i n th e s t a r c h m o le c u le may be 1 - 5
or 1 - 4 r e s p e c t i v e l y .
Thus
-pCH - 0 — o CH 0 M e
CHOM e G H — 0 — CH
GH^OMe
normal form \
pGH T 0 — o CHOMe
I GHOMe
?H
CH - 0 — CH^OMe
form
p ÇHOH.
I CHOMe CHOMe CHOH CH CH^OMe
c h a n g in g t o norm al form d u r in g h y d r o l y s i s .
S in c e t h a t t i m e , the m é t h y l a t i o n m ethod h a s b e e n w i d e l y
a p p l i e d t o th e p o l y s a c c h a r i d e s and many compounds i n th e su gar
s e r i e s have b e e n p r e p a r e d w i t h o u t a c a s e o f m i g r a t i o n o f m e th o x y l
gr o u p s b e in g d e t e c t e d . There r e m a i n s , h o w e v e r , t h e rem ote p o s s i
b i l i t y t h a t s u c h m i g r a t i o n m ig h t tak e p l a c e i n the h y d r o l y s i s o f
a m e t h y l a t e d p o l y s a c c h a r i d e and p a r t o f the w ork d e s c r i b e d i n the
t h e s i s was d e s i g n e d to remove t h i s d o u b t.
A lt h o u g h , th r o u g h th e work o f I r v i n e and Macdonald the
p o s i t i o n o f th e l i n k a g e s i n s t a r c h h ad th u s b e e n f a i r l y w e l l e s t a b
l i s h e d , t h e r e r em a in e d th e q u e s t i o n o f how many o f t h e s e a n h y d r o
-g l u c o s e u n i t s were th u s j o i n e d t o -g e t h e r t o form th e s t a r c h m o l e c u l e .
The h y d r o l y s i s p r o d u c t o f a f u l l y m e t h y l a t e d s t a r c h i s a lm o s t w h o l l y
2 - 3 - 6 t r i m e t h y l g l u c o s e ; b u t i n r e c e n t y e a r s , v a r i o u s w o r k e rs have
d e t e c t e d th e p r e s e n c e o f s m a l l q u a n t i t i e s o f 2 - 3 - 4 - 6 te trame t h y l
g l u c o s e among t h e h y d r o l y s i s p r o d u c t s . The i n t e r p r e t a t i o n o f t h i s
r e s u l t h a s g i v e n r i s e t o two m ain c o n c e p t i o n s o f th e s t a r c h m o l e c u l e .
The one p r e s e n t s a p i c t u r e o f t h e s t a r c h m o le c u le a s a s e r i e s o f
tw e n ty or t h i r t y a n h y d r o g lu c o s e u n i t s j o i n e d t o g e t h e r i n a l o n g
c h a i n , t e r m i n a t e d a t one e n d b y a r e d u c in g o r p o t e n t i a l l y r e d u c i n g
group and a t t h e o t h e r by an a n h y d r o - g l u c o s e r e s i d u e w i t h fo u r v a c a n t
h y d r o x y l g r o u p s , w h ic h on m é t h y l a t i o n w o u ld g i v e r i s e t o t e trame t h y l
S k e l e t o n Formula I .
Re due i n g G rou p.
r
I C Ic
I^ c
I G CHOH \ CHOMe CHOMe CHOH - 6nC“ C —C~C“C"*C I— 0 --- 1
J n
G iv e s on m e t h y l a t
-h y d r o l y s i s o f e d p r o d u c t .
CH OMe &
2 - 3 - 6 t r i m e t h y l g l u c o s e .
r CHOH
CHOMe
A (
0 CHOMe
CHOH L CH
I
CH, OMe z
2 - 3 - 6 t r i m e t h y l g l u c o s e .
r
0
L
r CHOH
CHOMe
0 CHOMe
CHOMe L CH
I
CH,OMe &
2 - 3 - 4 - 6 te tr a m e t h y l g l u c o s e .
A c c o r d in g t o the a l t e r n a t i v e v ie w t h e r e e x i s t s a " b a s i c
u n it " w h ic h c o n s i s t s o f a r e l a t i v e l y s m a l l number o f a n h y d r o
-g l u c o s e u n i t s j o i n e d i n a c l o s e d r i n -g . This " b a s i c u n i t " i s th e n
p o l y m e r i s e d or a s s o c i a t e d to g i v e the s t a r c h m o l e c u l e . See
r
0
. / °
Cv C - C - c - c - c - c
c ' ^ 0
i
c---6
6
0 0r
0L
?
1
00
S k e l e t o n Formula I I .
The number o f g l u c o s e u n i t s i n th e c l o s e d r i n g i s comp a r a t i v e l y s m a l l - s a y t h r e e or fo u r •
—J n
I t w i l l be s e e n t h a t I I w o u ld g iv e on h y d r o l y s i s o f th e
m e t h y l a t e d d e r i v a t i v e o n l y 2 - 3 - 6 t r i m e t h y l g l u c o s e and no te t r a
me t h y l g l u c o s e u n l e s s p a r t i a l h y d r o l y s i s to o k p l a c e t o form a
t e t r a s a c c h a r i d e ( S e e S k e l e t o n Formula I I I ) w h ic h on m é t h y l a t i o n
and h y d r o l y s i s w o u ld g i v e some t e trame t h y l g l u c o s e .
S k e l e t o n Formula I I I .
r
Ç
0
- i r
g i v e I - CHOH
Q CHOMe
1 GHOMe
CHOMe U CH
CH OMe
B reak h e r e to form t e t r a s a c C hari de .
Â
C-C-d-C-C
— 0 '
On h y d r o l y s i s o f m e t h y l a t e d p r o d u c t p o r t i o n A w o u ld
l i s h e d , have r e v e a l e d t h a t when r e l a t i v e l y l a r g e q u a n t i t i e s o f
p o l y s a c c h a r i d e s ar e e m p lo y e d , i t i s p o s s i b l e a f t e r m é t h y l a t i o n and
h y d r o l y s i s t o d e t e c t a s m a l l p r o p o r t i o n o f t h e c o r r e s p o n d i n g t e t r a
me t h y l h e x o se i n the h y d r o l y s i s p r o d u c t s . Thus, I r v i n e and
Montgomery ( 4 ) o b t a i n e d a b o u t 4^ o f t e t r a m e t h y l y f r u c t o s e from
t r i m e t h y l i n u l i n and s i m i l a r l y , b o t h f u l l y m e t h y l a t e d s t a r c h and
c e l l u l o s e y i e l d a p p r e c i a b l e q u a n t i t i e s ( 5 - 7 ^ ) o f t e t r a m e t h y l g l u
c o s e on h y d r o l y s i s . I t i s , h o w e v e r , an im p o r ta n t f a c t , t h a t th e
y i e l d o f t h i s s u g a r v a r i e s c o n s i d e r a b l y a c c o r d i n g t o th e method o f
m é t h y l a t i o n and a c c o r d i n g a l s o t o th e method em p lo y ed i n the p r e
l i m i n a r y a c é t y l a t i o n to w h ic h th e p o l y s a c c h a r i d e i s u s u a l l y s u b j e c t e d .
Somewhat s i m i l a r r e s u l t s have b e e n o b t a i n e d by Haworth and
Machemer ( 5 ) who q u o te e v i d e n c e , w h ic h i n t h e i r o p i n i o n i s c o n c l u s i v e ,
t h a t c e l l u l o s e , s t a r c h and . i n u l i n e a c h p o s s e s s a l o n g c h a in s t r u c
t u r e , and i t i s a l s o c l a i m e d t h a t the l e n g t h o f su c h c h a i n s may be
c a l c u l a t e d from th e amount o f t e t r a m e t h y l su g a r i s o l a t e d . The m a tte r
was a l s o d e a l t w i t h i n a c o r r e s p o n d e n c e b e t w e e n I r v i n e and H aw orth . (6).
Haworth s u b j e c t e d th e v a r i o u s p o l y s a c c h a r i d e s d e a l t w i t h
to d r a s t i c a c é t y l a t i o n b y means o f a c e t i c a n h y d r id e and s u l p h u r i c
a c i d , and c l a i m s t h a t , s i n c e no f r a c t i o n a t i o n o f h i s s t a r c h
t r i a c e t a t e c o u l d be e f f e c t e d , t h e m a t e r i a l had n o t s u f f e r e d " a c e t
i s shown t h a t a t r i - a c e t y l s t a r c h w h ic h c o u l d n o t be f r a c t i o n a t e d
h a d , i n p o i n t o f f a c t , u n d ergon e c o n s i d e r a b l e a c e t o l y s i s i n t h e
c o u r s e o f a much m i l d e r a c e t y l a t i n g r e a c t i o n . No v a l i d e v i d e n c e
a s t o th e r e l a t i v e m e r i t s o f Formula I o r I I can be o b t a i n e d by
a m ethod e m p lo y in g a p r e l i m i n a r y a c é t y l a t i o n , s i n c e " a c e t o l y s i s "
w o u ld b r e a k t h e c l o s e d r i n g s t r u c t u r e s u g g e s t e d i n Formula I I g i v i n g
a s u b s t a n c e o f Formula I I I w h ic h , on m é t h y l a t i o n and h y d r o l y s i s
w o u ld g i v e , a s d e s c r i b e d , some t e t r a m e t h y l g l u c o s e , a r e s u l t w h ic h
w ou ld a l s o be o b t a i n e d from a s u b s t a n c e o f Formula I .
E v id e n c e w h ic h , sh e c la i m e d , f a v o u r e d the l o n g c h a i n
s t r u c t u r e was a l s o p u t fo r w a r d by E . P e i s e r , ( 7 ) , who b a s e d h e r
r e s u l t s on th e a n a l y s i s o f v a r io u s a c e t a t e s o f s t a r c h p r e p a r e d by
t r e a t i n g t h e p o l y s a c c h a r i d e w i t h a c e t i c a n h y d r id e and s u l p h u r i c a c i d .
A l o n g c h a i n o f a n h y d r o - g l u c o s e u n i t s w o u ld g i v e a m o l e c u l e whose
carb on and h y d r o g e n c o n t e n t w ou ld n o t be d i s t i n g u i s h a b l e from
0^ b u t w h ic h on a c é t y l a t i o n w o u ld show an a c e t y l c o n t e n t
h i g h e r th a n t h a t o f a t r i a c e t y l s t a r c h . The v a l i d i t y o f t h i s work
h a s b e e n q u e s t i o n e d b y P r in g s h e im and M eyersohn ( 8 ) and a l s o by
B r i g l and S c h i n l e ( 9 ) who c o n t e n d t h a t th e h i g h a c e t y l c o n t e n t o b
t a i n e d i s due t o a c e t o l y s i s h a v in g t a k e n p l a c e , th u s fo r m in g a com
pound a n a lo g o u s t o t h a t o f Formula I I I . The r e s u l t s o b t a i n e d by
the p r e s e n t a u t h o r s u p p o r t th e co n d e m n a tio n o f P e i s e r ' s w o r k . Many
o t h e r w o r k e r s h a v e a c e t y l a t e d s t a r c h by v a r i o u s m e th o d s , e . g . ,
S c h u t z e n b e r g e r (ID), P r e g l ( 1 1 ) , P r in g s h e im ( 1 2 ) , b u t a f e a t u r e
’’r e g e n e r a t e d s t a r c h ” o b t a i n e d a p p e a r s t o have s u f f e r e d c o n s i d e r a b l e
d e g r a d a t i o n .
T urning on ce more to t h e q u e s t i o n o f t h e l e n g t h o f the
c h a i n i n t h e s t a r c h m o l e c u l e we f i n d t h a t th e e v i d e n c e o f X -r a y
s p e c t r a i s n o t c o n c l u s i v e b u t on t h e w h o le i n c l i n e s t o th e v ie w
t h a t t h e s t a r c h m o l e c u l e i s c o m p a r a t i v e l y s m a l l . Katz ( 1 3 ) f o r
e x a m p le , s t a t e s t h a t he f i n d s s t a r c h to be e s s e n t i a l l y c r y s t a l l i n e
and t h a t t h e e l e m e n t a r y c e l l , i n t h e c a s e o f a l l p o l y s a c c h a r i d e s
s o f a r ex a m in ed and i n s o f a r a s t h e y y i e l d a s u f f i c i e n t l y d e f i n i t e
c r y s t a l l i n e s p e c tr u m t o a l l o w o f an e v a l u a t i o n o f t h e w e i g h t o f t h e
e l e m e n t a r y b o d y , i s f o u n d t o be r e m a r k a b ly s m a l l . In t h e c a s e o f
s t a r c h he c o n s i d e r s i t t o c o n s i s t o f o n e , or a v e r y few g l u c o s e
u n i t s . On th e o t h e r h a n d , S . von Naray Szabo ( 1 4 ) c o n c lu d e s a s a
r e s u l t o f h i s X - r a y s t u d i e s o f s t a r c h t h a t t h e u n i t c e l l c o n t a i n s
a b o u t 1 6 a n h y d r o - g l u c o s e u n i t s , b u t a l s o s t a t e s t h a t th e r e s u l t
v a r i e d c o n s i d e r a b l y w i t h d i f f e r e n t s t a r c h e s .
E v id e n c e b a s e d on m o l e c u l a r w e i g h t d e t e r m i n a t i o n s o f p o l y
s a c c h a r i d e s i s m o st c o n f l i c t i n g , inasm uch a s r e s u l t s a p p e a r to d i f f e r
a c c o r d i n g to th e method o f d e t e r m i n a t i o n o f m o l e c u l a r w e i g h t , th e
s t a r c h e m p lo y e d , and the tim e t h a t th e s o l u t i o n s a r e k e p t .
C l o s e l y c o n n e c t e d w i t h t h i s m a t t e r i s th e h o m o g e n e it y o f
s t a r c h . We may c o n s i d e r t h r e e t y p e s o f n o n - h o m o g e n e it y . F i r s t l y ,
e l e m e n t s o t h e r th a n ca rb o n h y d r o g e n and o x y g e n fo u n d i n s t a r c h :
-Of t h e s e th e commonest a r e n i t r o g e n , p h o sp h o r u s and s i l i c o n ( a s
” I m p u r i t i e s ” or a s an i n t e g r a l p a r t o f the s t a r c h m o l e c u l e o p i n i o n
I s v e r y s h a r p l y d i v i d e d . A s e c o n d t y p e o f n o n - h o m o g e n e it y I s th e
a l l e g e d i s o l a t i o n from s t a r c h o f two d i s t i n c t compounds, am ylose
and a m y l0- p e c t i n . Numerous methods f o r the s e p a r a t i o n o f t h e s e
c o n s t i t u e n t s h a v e b e e n p r o p o s e d . G a t in GruzRWska ( 1 5 ) g i v e s a
s e p a r a t i o n b a s e d on t h e u s e o f a l k a l i , w h ic h d i s s o l v e s th e am ylose
and l e a v e s th e a m y l o - p e c t l n . L in g and N a n j l ( 1 6 ) g i v e an e x t e n s i v e
r e v i e w o f t h e s u b j e c t and d e s c r i b e e i g h t p h y s i c a l m ethods o f s e p a r
a t i n g th e two com pounds. Samec ( 1 7 ) c la im s t o have s e p a r a t e d the
two c o n s t i t u e n t s a n d f i n d s t h a t n e a r l y a l l th e p h o sp h o ru s I s c o n
t a i n e d In the a m y l o - p e c t l n f r a c t i o n .
On t h e o t h e r h a n d , many a u t h o r i t i e s deny the e x i s t e n c e o f
t h e s e compounds a s s e p a r a t e e n t i t l e s . Maquenne ( 1 8 ) c o n s i d e r s t h a t
th e a g e i n g o f s t a r c h c a u s e s a change from a s o - c a l l e d " a m y lo se ”
s t a t e t o an ”a r a y l o - p e c t l n ” s t a t e - an I r r e v e r s i b l e p r o c e s s .
Pouard ( 1 9 ) a f t e r a s e r i e s o f u l t r a - f i l t r a t Ion e x p e r im e n t s c o n c lu d e s
t h a t s t a r c h I s a s i n g l e c h e m ic a l s p e c i e s c a p a b le o f a p h y s i c a l t r a n s
f o r m a t i o n , t o t a l and r e v e r s i b l e , to w a r d a s t a t e o f p e r f e c t s o l u t i o n .
Katz ( 2 0 ) f i n d s t h a t a m y l o - p e c t l n and a m y lo se have th e same X -r a y
s p e c t r a when f r e s h b u t t h a t r é t r o g r a d a t i o n or a g e i n g a f f e c t s th e
s p e c t r a . P r i n g s h e im a l s o I n c l i n e s t o t h i s v i e w , p o i n t i n g o u t t h a t
th e r e v e r s i b l e c h a n g e s o f v i s c o s i t y In s t a r c h s o l u t i o n s may be due
t o a ch an ge from one form t o a n o t h e r and c i t i n g a s f a v o u r a b l e e v i
dence th e d i s a g g r e g a t i o n o f p o ly a r a y lo s e a c e t a t e s ( 2 1 ) w h ic h , a lt h o u g h
v e r y s i m i l a r c o n s t i t u t i o n .
I t w i l l be s e e n , t h e r e f o r e , from th e e x t r a c t s from th e
l i t e r a t u r e g i v e n ab o v e t h a t t h e r e a p p e a r s to be no a g r e e m e n t on
th e p r o b le m and i t was on t h e s e ground s t h a t Dr. R u t h e r f o r d and
th e p r e s e n t a u t h o r f e l t j u s t i f i e d i n c a r r y i n g o u t t h e i r w ork on
s t a r c h , r e g a r d e d a s a w h o l e , r a t h e r th a n a t t e m p t i n g t o work on the
s e p a r a t e d c o n s t i t u e n t s .
In th e c o u r s e o f t h i s work t h e r e was d i s c o v e r e d a t h i r d
ty p e o f n o n - h o m o g e n e it y - n a m e ly , w i t h i n th e m o l e c u l e i t s e l f .
A p a r t from a s u g g e s t i o n b y E . P e i s e r ( 2 2 ) t h a t an a l d e h y d i c group
e x i s t s i n the m id d le o f t h e s t a r c h m o l e c u l e , a l l w o r k e rs seem to
have a g r e e d t h a t th e m o l e c u l e o f s t a r c h i s s y m m e t r i c a l l y b u i l t up
o f a n h y d r o - g l u c o s e u n i t s . D r . R u t h e r f o r d , h o w e v e r , o b t a i n e d e v i
den ce w h ic h i n d i c a t e d t h a t t h i s i s n o t the c a s e , and th e work o f
t h e p r e s e n t a u t h o r c o n f ir m s t h i s d i s c o v e r y .
A sunmary o f D r. R u t h e r f o r d ' s t h e s i s i s now p r e s e n t e d i n
o r d e r t o g i v e an u n d e r s t a n d i n g o f th e b a s i s o f th e p r e s e n t e x t e n s i o n
o f t h e w o r k .
The i n i t i a l p r o b lem u n d e r t a k e n was an a t t e m p t t o i s o l a t e
and i d e n t i f y th e s u g a r s i n t e r m e d i a t e b e tw e e n s t a r c h a n d g l u c o s e .
S i n c e s t a r c h i s more e a s i l y h y d r o l y s e d b y a c i d s th a n i s c e l l u l o s e ,
i t was t h o u ^ t t h a t an a c e t y l a t i n g a g e n t su ch a s t h a t u s e d by
B a r n e t t ( 2 3 ) m i g h t , when a p p l i e d t o s t a r c h , b r i n g a b o u t a c e t o l y s i s
o f th e s t a r c h m o l e c u l e t o g i v e the a c e t a t e o f d e f i n i t e i n t e r m e d i a t e
r e l a t e d t o s t a r c h . T h is c o n j e c t u r e was n o t r e a l i s e d . The p r o d u c t
o b t a i n e d i n e x c e l l e n t y i e l d was an a c e t a t e , w h ic h on d e - a c é t y l a t i o n
gave a ’’r e g e n e r a t e d s t a r c h ” s o l u b l e i n w a t e r w i t h o u t t h e p r o d u c t i o n
o f a p a s t e and g i v i n g w i t h i o d i n e th e c r im s o n c o l o u r a t i o n c h a r a c t e r
i s t i c o f c e r t a i n d e x t r i n s . I t w o u ld a p p e a r , t h e r e f o r e , t h a t t h e r e
t a k e s p l a c e c o n c u r r e n t l y w i t h a c é t y l a t i o n , a n o t h e r r e a c t i o n i n v o l v i n g
d e p o l y m e r i s a t i o n and p o s s i b l y a l s o p a r t i a l h y d r o l y s i s o f t h e s t a r c h
m o l e c u l e .
I t was f o u n d , h o w e v e r , t h a t t h i s a c é t y l a t i o n f u r n i s h e d a
c o n v e n i e n t and s i m p l i f i e d m ethod f o r th e m é t h y l a t i o n o f s t a r c h .
By s i m u l t a n e o u s de - a c é t y l a t i o n and m é t h y l a t i o n o f t h e a c e t a t e by
means o f sodium h y d r o x id e and dime 1 t h y l s u l p h a t e a sm ooth and e a s y
m é t h y l a t i o n was o b t a i n e d . . There w a s , h o w e v e r , a d e f i n i t e c h e c k i n
th e e a s e o f m é t h y l a t i o n when a m e th o x y l c o n t e n t o f 34% OMe was
r e a c h e d . T his se e m e d t o c o r r e s p o n d t o a d e f i n i t e compound h a v i n g
r o u g h l y two m e t h o x y l g rou p s p e r u n i t . In o r d e r t o i n v e s t i g a t e
t h i s phenomenon th e r e s e a r c h was c o n t i n u e d a s a s t u d y o f p a r t i a l l y
m e t h y l a t e d s t a r c h and i t s h y d r o l y s i s p r o d u c t s . S t a r c h m e t h y l a t e d
t o a b o u t 34 % t h a t i s , t o a b o u t the d im e t h y l s t a g e was h y d r o l y s e d
b y a c i d m e t h y l a l c o h o l u n t i l p o l a r i m e t e r r e a d i n g s o f th e s o l u t i o n
became c o n s t a n t . The p r o d u c t s o f t h e h y d r o l y s i s may be c o n v e n i e n t l y
P a r t i a l l y M e t h y la t e d S t a r c h
T e trame t h y l Tr ime t h y l
G lu c o s e Zio G lu co s e 3 4 . 4 ^
U n d i s t i l l a b l e R e s id u e A. 1 7 . 9 ^
1 1
Dime t h y l Monome t h y l
G lu c o s e 3 2 . 9 ^ G lu c o s e 3 . 2 ^
U n d i s t i l l a b l e R e sid u e B . 8 . 6 #
The T e tr a m e th y l g l u c o s e , r e p r e s e n t i n g 3 # o f th e t o t a l h y d r o l y s i s
p r o d u c t s was r e g a r d e d a s b e i n g due t o a r u p tu r e o f a l i n k a g e d u r in g
a c é t y l a t i o n l e a v i n g an anhydro g l u c o s e u n i t w i t h f o u r v a c a n t h y d r o x y l
g r o u p s a s shown i n Formula I I I p . 5 . The t r i m e t h y l g l u c o s e o b t a i n e d
p r o v e d t o be 2 - 3 - 6 t r i m e t h y l g l u c o s e , a r e s u l t w h ic h i s i n a g r ee m e n t
w i t h a l l p r e v i o u s w o r k . The c o n s t i t u t i o n o f the d im e t h y l g l u c o s e
f r a c t i o n was I n v e s t i g a t e d t h o r o u ^ l y and s e t t l e d c o n c l u s i v e l y .
S i n c e th e 4 and 5 p o s i t i o n s a r e o c c u p i e d b y the r i n g and
th e o x y g e n l i n k a g e , th e dime t h y l s u g a r s o b t a i n a b l e from s t a r c h
s h o u ld b e , 3 - 6 , 2 - 6 , and 2 - 3 d im e t h y l g l u c o s e . I t was f o u n d t h a t
a c i d a c e t o n e w o u ld c o n d e n s e w i t h 2 - 3 d im e t h y l g l u c o s e and 3 - 6
d im e th y l g l u c o s e , b u t n o t w i t h 2 - 6 d im e t h y l g l u c o s e . .Thus th e
q u a n t i t y o f 2 - 3 d i m e t h y l g l u c o s e + 3 - 6 d im e th y l g l u c o s e c o u l d be
e s t i m a t e d . I t was a l s o fo u n d ( 2 4 ) t h a t when the p a r a - t o l u e n e
s u l p h o n y l d e r i v a t i v e o f th e m ix ed s u g a r s was formed the p a r a t o l u e n e
s u l p h o n y l g r o u p i n p o s i t i o n 6 was r e p l a c a b l e by i o d i n e and an i o d i n e
p r e s e n t Thus
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2 - 3 d im e t h y l m e th y l g l u c o s e .
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2 - 3 d im e th y l' 4 - 6 d i - p a r a - t o l u e n e s u l p h o n y l m e th y l g l u c o s i d e .
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2 - 3 d im e t h y l 4 para-t o l u e n e s u l p h o n y l 6 - io d o m eth y l g l u c o s i d e .
By s u b t r a c t i o n the amount o f 3 - 6 , and 2 - 6 d im e t h y l g l u c o s e s
c o u l d be f o u n d . The r e s u l t s were
-2 - 6 d im e t h y l g l u c o s e 76^ 2 - 3 d im e t h y l g l u c o s e 24^
3 - 6 d im e t h y l g l u c o s e n e g l i g i b l e .
An i n v e s t i g a t i o n o f th e monome t h y l g l u c o s e s p r e s e n t was th e n
u n d e r t a k e n . These w ere i d e n t i f i e d and from t h e r e s u l t s i t was
d e d u ced t h a t p o s i t i o n 3 o f th e a n h y d r o - g l u c o s e u n i t s p r e s e n t in
s t a r c h i s the l a s t to become m e t h y l a t e d .
A t t e n t i o n was now t u r n e d t o R e s i d u e s A and B ( s e e p . 1 2 )
w h ic h h a d b e e n fo u n d t o c o n s i s t o f u n h y d r o ly s e d m a t e r i a l . They
b o t h had a m e th o x y l c o n t e n t o f 3 5 - 3 6 ^ and w ere h a r d g l a s s e s , w h i l e ,
i n sh arp c o n t r a s t , a g l u c o s e m e t h y l a t e d t o t h i s e x t e n t i s a f a i r l y
v i s c o u s s y r u p . On b o i l i n g w i t h aqueous a c i d or w i t h a c i d m e th y l
a l c o h o l t h e y w ere c o m p l e t e l y h y d r o l y s e d t o g i v e a m ix tu r e o f t r i
m é t h y l a t i o n y i e l d e d p r o d u c t s w h ic h w ere s t i l l h a r d g l a s s e s and
v/h ich had m e th o x y l c o n t e n t s o f from 4 6 - 4 8 ^ w h e r e a s the m e th o x y l
c o n t e n t o f a t r i m e t h y l a n h y d r o - g l u c o s e i s s l i ^ t l y lo w e r v i z . 4 5 . 5 ^ .
H y d r o l y s i s o f the f u l l y m e t h y l a t e d compounds g ave t e t r a m e t h y l ,
t r i m e t h y l and d im e t h y l g l u c o s e s i n t h e a p p r o x im a te p r o p o r t i o n o f
1 : 2: 1 . The i s o l a t i o n o f 2 - 3 - 4 - 6 t e t r a m e t h y l g l u c o s e i m p l i e s the
p r e s e n c e i n t h e r e s i d u e o f t h e g r o u p i n g .
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The i s o l a t i o n o f 2 - 3 - 6 t r i m e t h y l g l u c o s e i m p l i e s th e p r e s e n c e o f
th e g r o u p in g
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I
G G G G G G G G G G G G G G G G G G
I o ---( I---O---1 I---o ______/
I I I I I I
The d i m e t h y l f r a c t i o n was foun d t o be s i m i l a r t o t h a t o b t a i n e d from
m e t h y l a t e d s t a r c h i . e . i t c o n t a i n e d a p p r o x i m a t e l y 24^ o f 2 - 3 d im e th y l
g l u c o s e a n d 76^ o f 2 - 6 d im e t h y l g l u c o s e .
m o l e c u l e o f s t a r c h i s n o t s y m m e t r ic a l — i f t h e r e s i d u e s A and B a r e
a c t u a l l y f r a c t i o n s o f s t a r c h . S t a r c h was t h e n f u l l y m e t h y l a t e d by
the d im e th y l s u l p h a t e r e a c t i o n , a f t e r a p r e l i m i n a r y a c é t y l a t i o n and
the p r o d u c t s o b t a i n e d on h y d r o l y s i s were
-M e t h y la t e d S t a r c h
20% E x t r a c t e d b y E t h e r .
T e tr a m e th y l T r im e th y l D im e th y l R e s id u e 11.0%
g l u c o s e 5% ' g l u c o s e 59.5% g l u c o s e 4.5%
The amount o f d im e t h y l g l u c o s e o b t a i n e d a g r e e s w i t h t h e
amount w h ich s h o u l d be o b t a i n e d from the r e s i d u e s A and B, a ss u m in g
t h a t th ey were p r e s e n t in the same p r o p o r t i o n as i n p a r t i a l l y
m e t h y l a t e d s t a r c h .
F i n a l l y , s t a r c h was m e t h y l a t e d t h i r t y tim e s w i t h o u t p r e v io u s
a c é t y l a t i o n . The p r o d u c t was e x t r a c t e d w i t h e t h e r and th e r e s i d u e
h y d r o l y s e d w i t h a c i d m eth y l a l c o h o l . The p r o d u c t s w ere t r i m e t h y l
g l u c o s e and d im e t h y l g l u c o s e — no t e t r a m e t h y l g l u c o s e b e i n g formed.*
From a c o n s i d e r a t i o n o f th e h y d r o l y s i s p r o d u c t s and a n a l y s e s
NOTE. I t s h o u l d be n o t e d h e r e t h a t s i n c e the c o m p i l a t i o n o f h e r t h e s i s Dr. R u t h e r f o r d r e p e a t e d t h i s p o r t i o n o f the work on a v e r y l a r g e s c a l e and s u c c e e d e d i n i s o l a t i n g from a m e t h y l a t e d s t a r c h w h ich h a d n o t
o f t h e s e r e s i d u e s i t was th o u g h t t h a t t h e y m ig h t c o n s i s t o f a
m e t h y l a t e d t e t r a s a c c h a r i d e su ch a s shown i n Formula I I I , p . 5 ,
d e r i v e d in t u r n from an a n h y d r o - t e t r a s a c c h a r i d e w h ic h was th e
" b a s i c u n i t ” and w h ic h on a s s o c i a t i o n or p o l y m e r i s a t i o n form ed
s t a r c h .
T h is t h e n , was the p o s i t i o n when th e p r e s e n t a u t h o r to o k
up t h e r e s e a r c h . The programme drawn up may be r o u g h l y d i v i d e d
i n t o th r e e p a r t s .
( 1 ) To e x p l o r e th e a c t i o n o f a c e t y l a t i n g a g e n t s on s t a r c h i n
o r d e r , f i r s t l y , t o a s c e r t a i n how f a r th e u l t i m a t e f o r m a t i o n o f
t e trame t h y l g l u c o s e a f f o r d s v a l i d e v i d e n c e o f th e s t r u c t u r e o f the
p a r e n t p o l y s a c c h a r i d e , and s e c o n d l y , s i n c e a c é t y l a t i o n was known
t o c a u s e e x t e n s i v e de p o l y m e r i s a t i o n , t o a t t e m p t t o o b t a i n b y t h i s
means the " b a s i c u n i t ” o f s t a r c h , s u s p e c t e d o f b e i n g an a n h y d r o
-t e -t r a s a c c h a r i d e .
( 2 ) To i n v e s t i g a t e f u r t h e r t h e n a tu r e a n d i d e n t i t y o f the r e s i d u e s
o b t a i n e d by Dr. R u t h e r f o r d . These may h a v e b e e n d e r i v e d from a
component p a r t o f the s t a r c h m o l e c u l e or may h ave b e e n form ed b y
( a ) the p r e l i m i n a r y a c é t y l a t i o n , ( b ) th e m é t h y l a t i o n w i t h d im e t h y l
s u l p h a t e , ( c ) th e a c i d m e th y l a l c o h o l u s e d i n h y d r o l y s i s . Methods
w ere d e v i s e d w h ic h e l i m i n a t e d t h e s e t h r e e p o s s i b i l i t i e s and p r o v e d
t h a t t h e s e r e s i d u e s a r e d e r i v e d from an i n t e g r a l p a r t o f th e s t a r c h
m o l e c u l e .
from a m eth yl group t o remove the s u s p i c i o n t h a t m i g r a t i o n o f
m e th y l groups m ig h t ta k e p l a c e d u r in g th e h y d r o l y s i s o f m e t h y l
ACETYLATION OP STARCH.
F i r s t M o d i f i c a t i o n o f B a r n e t t P r o c e s s
In p u r s u a n c e o f t h e f i r s t p a r t o f t h i s programme, a b o u t
two k i lo g r a m s o f s t a r c h t r i a c e t a t e were p r e p a r e d by m o d i f y i n g i n
a minor d e g r e e th e B a r n e t t p r o c e s s e m p lo y e d b y Dr, R u t h e r f o r d .
On s e v e r a l o c c a s i o n s an a c e t a t e (w h ic h g i v e s " D e x t r in A" on d e
-a c e t y l -a t i o n ) sh o w in g a p p r o x i m a t e l y 1 6 0 ° i n c h l o r o f o r m
was o b t a i n e d and t h i s v a l u e was n o t g r e a t l y d i s t u r b e d i f r i c e
s t a r c h w ere r e p l a c e d b y p o t a t o s t a r c h i n t h e p r e p a r a t i o n . Under
p a r a l l e l c o n d i t i o n s , h o w e v e r . D r . R u t h e r f o r d i n v a r i a b l y o b t a i n e d a
p r o d u c t sh o w in g [ ^ 3 ^ 1 5 0 ° . P r e g l ( 1 1 ) d e s c r i b e s a s t a r c h
t r i a c e t a t e o f s p e c i f i c r o t a t i o n ^ •+* 1 6 3 . 6 ° ; w h i l e Bergmann
and Kneke ( 2 5 ) o b t a i n e d a n a m y lo s e a c e t a t e o f s p e c i f i c r o t a t i o n
= "f 1 7 6 ° . I t i s d i f f i c u l t to e x p l a i n t h e s e d i s c r e p a n c i e s ,
w h ic h w ere e n c o u n t e r e d on num erous o c c a s i o n s a n d th e r e s u l t s a r e
q u o t e d a s an i n d i c a t i o n , n o t o n l y o f th e l a c k o f p r e c i s i o n o f the
" p h y s i c a l c o n s t a n t s " i n t h i s s e r i e s , b u t a l s o o f th e c a u t i o n w h ic h
m u st be e x e r c i s e d i n u t i l i s i n g m a t e r i a l o f s o v a r i a b l e a n a t u r e a s
T rea tm en t o f S t a r c h T r i a c e t a t e s .
A tte m p ts w e re a c c o r d i n g l y made t o s e p a r a t e t h e s e s t a r c h
a c e t a t e s i n t o f r a c t i o n s o f u n i f o r m n a t u r e . P r o l o n g e d e x t r a c t i o n s
i n a S o x h l e t a p p a r a t u s w i t h e t h y l a l c o h o l d i d n o t e f f e c t any
d i f f e r e n t i a t i o n a n d , i n a d d i t i o n , s l i g h t h y d r o l y s i s and decom p os
i t i o n o c c u r r e d . F r a c t i o n a l p r e c i p i t a t i o n by e t h e r from s o l u t i o n
i n c h l o r o f o r m was u n a v a i l i n g and u l t i m a t e l y r e c o u r s e was h a d to
s o l u t i o n i n e t h y l a c e t a t e an d f r a c t i o n a l p r e c i p i t a t i o n w i t h e t h e r .
Taking th e s p e c i f i c r o t a t i o n o f t h e f r a c t i o n s a s a c r i t e r i o n the
d i s s e c t i o n o f th e a c e t a t e s w as c a r r i e d t o a l i m i t b e y o n d w h ic h
p r e c i p i t a t i o n w i t h e t h y l a c e t a t e p r o v e d t o be no l o n g e r e f f e c t i v e .
The v a r i o u s f r a c t i o n s were d e - a c e t y l a t e d and an a n a l y s i s o f t h e
d e x t r i n s fo r m e d , ( a l l o f w h ic h r e d u c e d F e h l i n g ' s s o l u t i o n s l i g h t l y )
sh ow ed t h a t a r e a l f r a c t i o n a t i o n had t a k e n p l a c e . The c a r b o n
c o n t e n t s - 4 3 . 5 ^ , 43.7% , 44.2% - i n d i c a t e d , s i n c e t h e y w ere lo w e r
th a n t h e t h e o r e t i c a l c a r b o n c o n t e n t o f an a n h y d r o - g l u c o s e (44.4% )
t h a t e v e n i n the c a s e o f th e h i g h e s t v a l u e some h y d r o l y s i s h a d
a c c o m p a n ie d t h e u s e o f th e B a r n e t t r e a g e n t s .
S e v e r a l a t t e m p t s w e r e , h o w e v e r , made to i s o l a t e th e anhydro
compound a s a n o n - r e d u c i n g f r a c t i o n , f i r s t b y means o f d e s t r o y i n g
th e r e d u c i n g m a t e r i a l w i t h h o t a l k a l i , and s e c o n d b y o x i d i s i n g t h e
r e d u c i n g m a t e r i a l w i t h b r o m in e w a t e r , b u t n e i t h e r p r o c e s s was
s u c c e s s f u l •
t o s e v e r a l r e p e t i t i o n s o f th e c y c l e a c é t y l a t i o n d e a c e t y l a t i o n
-a c é t y l -a t i o n , w i t h th e o b j e c t o f d i s c o v e r i n g w h e th er " -a c e t o l y s i s "
w o u ld be c o n t i n u e d s t e a d i l y or be c h e c k e d a t a tim e when th e a c e t
a t e s h a d b e e n d e g r a d e d t o a f u l l y a c e t y l a t e d a n h y d r o - t e t r a
s a c c h a r i d e . A s t e a d y f a l l i n th e s p e c i f i c r o t a t i o n and an i n c r e a s e
i n t h e e f f e c t on P e h l i n g * s s o l u t i o n i n d i c a t e d t h a t a c e t o l y s i s was
b e i n g m a i n t a i n e d . I t i s i n t e r e s t i n g t o c o n t r a s t t h e s e r e s u l t s w i t h
t h o s e o f D r . J . R u t h e r f o r d who r e t r e a t e d th e a ce t a t e ( n o t th e
d e x t r i n ) b y p l a c i n g i t i n t h e B a r n e t t r e a g e n t s and h e a t i n g i t a t
50®C. f o r t h r e e d a y s . I t was t h e n i s o l a t e d u n ch an ged a s r e g a r d s
s p e c i f i c r o t a t i o n an d a n a l y s i s . I t w o u ld appear t h a t i t i s th e
a c t u a l i n t r o d u c t i o n o f t h e a c e t y l g r o u p s i n t o th e d e x t r i n w h ic h
c a u s e s h y d r o l y s i s and d e p o l y m e r i s a t i o n .
S e c o n d M o d i f i c a t i o n o f B a r n e t t P r o c e s s .
The B a r n e t t p r o c e s s was t h e r e f o r e s t i l l f u r t h e r r flo d ifie d
b y r e p l a c i n g t h e tem p o ra r y h i g h c o n c e n t r a t i o n s o f s u l p h u r y l c h l o r i d e
(w h ic h o c c u r r e d when the r e a c t i o n m ix t u r e was g i v e n a "dose" o f
s u l p h u r d i o x i d e and c h l o r i n e t w i c e a d a y ) by a c o n t i n u o u s low c o n
c e n t r a t i o n , w h i c h , i t was t h o u g h t , w o u ld n o t be so l i a b l e t o c a u s e
h y d r o l y s i s . A c é t y l a t i o n to o k p l a c e s m o o t h ly i n a b o u t f i v e d a y s ,
th e w h o le o f the s t a r c h p a s s i n g i n t o s o l u t i o n , th u s d i f f e r i n g from
t h e f i r s t m o d i f i c a t i o n o f th e B a r n e t t p r o c e s s , i n w h i c h , no m a t t e r
u n d i s s o l v e d . T h is f a c t r a i s e d a s i d e i s s u e w h ic h may he c o n v e n
i e n t l y d i s c u s s e d h e r e .
The r e s i d u e a p p e a r e d t o he u n a l t e r e d s t a r c h , a s i t gave
a v i o l e t c o l o u r w i t h i o d i n e , h u t t h e r e e x i s t e d th e p o s s i b i l i t y t h a t
i t m ig h t c o n s i s t o f a more r e s i s t a n t p o r t i o n o f th e s t a r c h c o n t a i n
in g th e u s u a l l y a s s o c i a t e d n i t r o g e n and p h osp h oru s d e r i v a t i v e s .
The manner i n w h ic h n i t r o g e n and p h o sp h o ru s are p r e s e n t i n s t a r c h
h a s n o t h e e n d e f i n i t e l y s e t t l e d and i t was t h e r e f o r e im p o r ta n t to
i n v e s t i g a t e th e m a t t e r . However, th e v a l u e s o f a s h d e t e r m i n a t i o n s ,
c a r r i e d o u t on t h e r e s i d u e s and on t h e o r i g i n a l s t a r c h , w ere p r a c
t i c a l l y i d e n t i c a l , and th e i n f e r e n c e t o he drawn i s t h a t th e
n i t r o g e n and p h o sp h o r u s a r e s o d i s t r i b u t e d t h a t t h e B a r n e t t p r o c e s s
d oes n o t e f f e c t any d i s s e c t i o n o f t h o s e p a r t s o f th e m o l e c u l e c o n
t a i n i n g them . In th e l i g h t o f l a t e r work i t w o u ld a p p ea r t h a t
t h e s e r e s i d u e s w ere due t o one p o r t i o n o f the com p lex b e i n g more
r e s i s t a n t to h y d r o l y s i s and l e s s r e a d i l y a t t a c k e d by a c e t y l a t i n g
r e a g e n t s . On t h e o t h e r hand, t h e d i f f e r e n t i a t i o n may he p u r e l y
m e c h a n i c a l due t o th e v a r y i n g s i z e o f th e s t a r c h g r a i n s .
By the s e c o n d m o d i f i c a t i o n o f th e B a r n e t t p r o c e s s th e r e
w ere o b t a i n e d two p r e p a r a t i o n s o f a c e t a t e s , w h ic h , on d e - a c e t y l a t i o n
f u r n i s h e d d e x t r i n s o f a new c h a r a c t e r ( ’’D e x t r i n B” ) . They d i d n o t
r e d u c e F e h l i n g ' s s o l u t i o n and on a n a l y s i s gave th e f o l l o w i n g
f i g u r e s ;
-C » 4 4 . 1 5 # H - 6 . 3 5 #
4 4 . 2 5 # 6 . 1 8 #
They w ere much more h i g h l y p o l y m e r i s e d th a n D e x t r i n A>
t h e i r a c e t a t e s g i v i n g c l o u d y s o l u t i o n s i n o r g a n i c s o l v e n t s , an d
i n e v e r y way t h e y a p p e a r e d t o h e v e r y c l o s e l y r e l a t e d t o s t a r c h .
In d e a l i n g w i t h compounds o f t h i s ty p e a n d , t o a l e s s e r d e g r e e ,
w i t h d e x t r i n s o b t a i n e d b y the o r d i n a r y B a r n e t t p r o c e s s , F e h l i n g ' s
s o l u t i o n h a s p r o v e d u n s a t i s f a c t o r y f o r th e d e t e c t i o n o f r e d u c i n g
p r o p e r t i e s , on a c c o u n t o f th e c o l l o i d a l n a t u r e o f th e m a t e r i a l ,
i t s h i g h m o l e c u l a r w e i g h t a n d th e u n s u i t a b i l i t y o f the t e s t f o r
e x a c t c o m p a r a t iv e w o r k .
E s t i m a t i o n o f R e d u c in g M a t e r i a l .
I t was t h e r e f o r e d e c i d e d t o a d o p t H a n e 's m o d i f i c a t i o n ( 2 6 )
o f th e H agedorn J a n s e n m ethod f o r t h e e s t i m a t i o n o f s m a l l q u a n t i t i e s
o f r e d u c i n g m a t e r i a l . T h is m ethod a t on ce r e v e a l e d t h e e f f e c t o f
a c é t y l a t i o n on t h e s t a r c h m o l e c u l e . The t a b l e shown belov/ i s
c o m p i le d from t h e t e t r a t l o n s on p . 7/. an d p r e s e n t s t h e r e s u l t s i n
a form w h ic h shows more c l e a r l y th e c o n c l u s i o n s w h ic h may be draw n.
The r e d u c i n g powers o f t h e v a r i o u s p r o d u c t s a r e com p ared, g l u c o s e
I I I I I IV
M a t e r i a l u s e d .
R e l a t i v e R e d u c in g Power r e f e r r e d t o G lu c o s e
= 1 0 0 0 .
P e r c e n t a g e o f M a lto s e R e s i d u e s .
A p p roxim ate y i e l d o f t e t r a me t h y l g l u c o s e w h ic h w o u ld be o b t a i n e d from e a c h s p e c i m e n .
R i c e S t a r c h ,
u n t r e a t e d . 2 . 4 0 . 3 6 0 . 1 8 #
D e x t r i n B, 1 s t
p r e p a r a t i o n . • 2 . 6 0 . 3 9 0 . 2 0
D e x t r i n B, 2nd
p r e p a r a t i o n 4 . 8 0 . 7 2 0 . 3 6
D e x t r i n A. 6 1 .1 9 . 0 4 . 5
M a l t o s e . 6 7 4 . 7 1 0 0 . 0 5 0 . 0
G l u c o s e . 1 0 0 0 . — — —
---As t h e e x p e r i m e n t a l e r r o r i n the s c a l e o f Column I I i s
+ 2 u n i t s , th e r e d u c i n g p ow ers o f th e two p r e p a r a t i o n s o f "B"
a n d o f t h e o r i g i n a l s t a r c h may he t a k e n a s i d e n t i c a l . On th e o t h e r
h a n d , t h e r e d u c i n g power o f s t a r c h i s m a g n i f i e d a t l e a s t tv /e lv e
b y th e o r i g i n a l m o d i f i c a t i o n o f th e B a r n e t t p r o c e s s and t h e r e a f t e r
r e g e n e r a t e d to g i v e " D e x t r in A*’ . T h is r e d u c i n g power s h o u l d be
com pared w i t h t h a t o f m a l t o s e , and n o t o f g l u c o s e , a s th e r e d u c i n g
p r o p e r t i e s o f a s t a r c h p r o b a b l y r e s i d e i n a m a l t o s e r e s i d u e a t t a c h e d
a t th e e n d o f a c h a i n . I t i s t h e r e f o r e p o s s i b l e t o c a l c u l a t e th e
p e r c e n t a g e o f m a l t o s e r e s i d u e s n e c e s s a r y to a c c o u n t f o r t h e r e d u c i n g
p r o p e r t i e s a s d e t e r m in e d e x p e r i m e n t a l l y . Prom t h i s , i n t u r n , th e
amount o f t e t r a - m e t h y l g l u c o s e w h ic h w o u ld be p r o d u c e d b y m é t h y l
a t i o n and h y d r o l y s i s o f t h e o r i g i n a l s u b s t a n c e can b e a s c e r t a i n e d
a n d t h i s i s done i n Column IV . The s t r i k i n g f a c t w h ic h e m e rg e s
i s t h a t " D e x t r in A" w h ic h c o r r e s p o n d s w i t h a v e r a g e s p e c im e n s o f
a c e t y l a t e d s t a r c h u s e d i n m é t h y l a t i o n w o u ld y i e l d a b o u t 4 or 5^
o f t e trame t h y l g l u c o s e . T h is i s t h e amount fo u n d e x p e r i m e n t a l l y
b y o t h e r w o r k e r s i n th is l a b o r a t o r y , b u t , i n c o n t r a s t , when s t a r c h
i s m e t h y l a t e d w i t h o u t p r e v i o u s a c é t y l a t i o n t e t r a - m e t h y l g l u c o s e i s
o b t a i n e d from t h e h y d r o l y s i s p r o d u c t s i n v e r y much s m a l l e r y i e l d .
I n v e s t i g a t i o n o f A c t i o n o f A l k a l i .
The n e x t s t e p was t o d e t e r m i n e w h e t h e r e v e n t h e s m a l l e r
q u a n t i t y o f t e t r a - m e t h y l g l u c o s e m ig h t n o t be a c c o u n t e d f o r b y th e
e f f e c t on t h e d e l i c a t e s t r u c t u r e o f t h e s t a r c h m o l e c u l e o f s u c h
d r a s t i c r e a g e n t s a s m e t h y l s u l p h a t e and c a u s t i c s o d a .
The v a r i o u s d e x t r i n s w ere t h e r e f o r e s u b j e c t e d to th e
m é t h y l a t i o n and t h e chan ge i n r e d u c i n g power n o t e d . The r e s u l t s
a r e shown i n th e T ab le b e lo w w h ic h i s c o m p ile d from t h e t i t r a t i o n s
on p . 73
R e d u c in g Power (By H a n e 's M ethod) R e f e r r e d t o G lu c o s e ■ 1 0 0 0 .
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R ic e S t a r c h . 2 . 4 2 . 2 2 . 9
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1 s t P r e p . 2 . 6 2 . 7 7 . 1
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Under t h e c o n d i t i o n s e m p lo y e d i n m é t h y l a t i o n th e f i r s t
e f f e c t o f h o t a l k a l i i s t o i n c r e a s e th e r e d u c i n g p o w e r . V/hy t h i s
s h o u l d ta k e p l a c e i s n o t known. I f i t i s due t o t h e r u p t u r e o f
a l i n k a g e b e t w e e n two a n h y d r o - g l u c o s e u n i t s th u s s e t t i n g f r e e a
new r e d u c i n g g r o u p , t h e n i t i s th e f i r s t c a s e known o f a g l u c o s i d i c
