11 Metabolism of L-Ascorbic A c i d in Plants FRANK A. LOEWUS and JOHANNES P. F. G. HELSPER Institute of Biological Chemistry, Washington State University, Pullman, W A 99164
A detailed study of the catabolism of L-ascorbic acid in tartrate-accumulating and oxalate-accumulating plants has revealed a precursor-product relationship which, in the case of tartrate accumulators, involves two mutually exclusive pathways. In the Vitaceae, the carbon chain of ascorbic acid is cleaved at the C4-C5 bond to furnish a C fragment that is converted to L-(+)-tartaric acid and a C fragment that is recycled into products of hexose phosphate metabo lism. In the Geraniaceae, cleavage occurs at the C2-C3 bond to produce oxalic acid from the C fragment and L-(+)-tartaric acid from the C fragment. Other oxalate accumulators that fail to accumulate tartaric acid also cleave ascorbic acid at the C2-C3 bond. The nature of the prod uct(s) obtained from the C fragment must still be deter mined. Conversion of D-glucose to ascorbic acid and its catabolic products has been studied in both types of plants with the aid of specifically labeled D-glucose. Results sup port a biosynthetic route that involves oxidation of C1 of D-glucose, epimerization at C5, and conservation of the hydroxymethyl function at C6. 4
2
2
4
4
Knowledge of metabolic events encompassing the formation and decomposition of ascorbic acid in plants is meager although L-ascorbic acid is a common constituent in actively growing tissues of higher plants. Despite a plethora of information on chemical, biomedical, and nutritional aspects of ascorbic acid, progress on plant-related processes has lagged. In part, the reason for this may lie in the generally accepted view that ascorbic acid is a secondary product of plant metabolism, a sugar acid outside the mainstream of carbohydrate inter conversions. Again, it may rest in a paucity of well-established roles for ascorbic acid in plant 0065-2393/82/0200-0249$06.00/0 © 1982 American Chemical Society
250
ASCORBIC
metabolism.
F o r w h a t e v e r reasons
a r e b e h i n d this neglect,
ACID
renewed
efforts t o d i s c o v e r t h e b i o c h e m i c a l events t h a t g o v e r n p a r t i c i p a t i o n o f a s c o r b i c a c i d i n c e l l u l a r processes are i n o r d e r .
Conversion of Ascorbate to Tartrate in the Grape A n i m p o r t a n t d i s c o v e r y i n 1969 ( 1 ) l i n k e d t h e c a t a b o l i s m of L - a s c o r b i c a c i d to t a r t a r i c a c i d biosynthesis. W h e n i m m a t u r e g r a p e berries w e r e f e d L - [ 1 - C ] a s c o r b i c a c i d o v e r a 2 4 - h p e r i o d , 7 2 % of t h e a c i d e x t r a c t a b l e 1 4
1 4
C a p p e a r e d i n t a r t a r i c a c i d , v i r t u a l l y a l l of i t i n c a r b o x y l c a r b o n .
When
t h e m e t a b o l i c p e r i o d w a s e x t e n d e d a n o t h e r 24 h , o n l y 4 8 % o f t h e r e m a i n e d i n t a r t a r i c a c i d , a n i n d i c a t i o n t h a t c a t a b o l i c processes r e m o v i n g a p a r t of the l a b e l e d t a r t a r i c a c i d (2,3,4).
Saito a n d K a s a i
1 4
C
were (1)
suggested t h a t C l t h r o u g h C 4 of L - a s c o r b i c a c i d w a s c o n v e r t e d d i r e c t l y i n t o t a r t a r i c a c i d ( F i g u r e 1 ) . I n d i r e c t s u p p o r t o f this w a s t h e f a i l u r e o f L - [ 6 - C ] a s c o r b i c a c i d to p r o d u c e l a b e l e d t a r t a r i c a c i d (5). 1 4
Wagner Kasai
and Loewus
(6)
confirmed
t h e observations
Subsequently, of
Saito a n d
(1).
Williams and Loewus
(7) prepared L-[4- C]ascorbic acid b y the 1 4
m e t h o d of B a k k e a n d T h e a n d e r (8) a n d s h o w e d t h a t this f o r m o f spe c i f i c a l l y l a b e l e d a s c o r b i c a c i d , l i k e L - [ 1 - C ] a s c o r b i c a c i d , w a s a n effective 1 4
p r e c u r s o r o f t a r t a r i c a c i d i n g r a p e berries a n d g r a p e leaves ( T a b l e I ) O v e r 9 8 % o f the
1 4
(9).
C w a s l o c a t e d i n t h e c a r b o x y l g r o u p s of l a b e l e d t a r t a r i c
a c i d f r o m L - [ 1 - C ] - o r L - [ 4 - C ] a s c o r b i c a c i d l a b e l e d leaves o r b e r r i e s . 1 4
1 4
O n l y L - ( + )-tartaric a c i d was formed
(10).
The C
2
fragment of this
c l e a v a g e , as j u d g e d b y studies w i t h L - [ 6 - C ] a s c o r b i c a c i d , w a s r e c y c l e d 1 4
i n t o p r o d u c t s of hexose p h o s p h a t e m e t a b o l i s m
(5,6,11,12).
C o n v e r s i o n o f L - a s c o r b i c a c i d to t a r t a r i c a c i d i n t h e g r a p e w a s l i m i t e d to c e r t a i n stages o f d e v e l o p m e n t
(Tables I a n d I I ) . Leaves
detached
f r o m t h e t i p of t h e v i n e o r t h e p o s i t i o n o p p o s i t e t h e flower c l u s t e r p r i o r to anthesis ( o r a t anthesis) r e a d i l y u t i l i z e d a s c o r b i c a c i d f o r t a r t a r i c a c i d COOH
L-(+)-Tartaric acid HO-t-H CHgOH
L-Ascorbic acid
Figure 1.
[i]
Hexose phosphate pool
Cleavage of ^ascorbic add in plants of the Vitaceae.
11.
Plant Metabolism of
LOEWUS A N D HELSPER
L-Ascorbic Acid
Conversion of L-Ascorbic A c i d to T a r t a r i c A c i d in the Grape (Metabolic Period, 25 h)
Table I.
[1- C]
[4- C]
14
Conditions
Berry
Leaf
251
Percent
{6,9)
(5,9)
light dark light
69 67 66
anthesis + 14 d a y s " + 4 8 to 5 7 d a y s "
81
[6- C]
14
14
of Acid Extractable
C
14
2
—
60 81 57 3
— 0.1
< 1 < 1
Days after anthesis.
tf
Table II.
L-Ascorbic A c i d Catabolism in Grape (Metabolic Period, 24 h) Days
Before
(—)
1 4
C]
co
of Label
in Ascorbic
[6-
[1-
[6-
biosynthesis.
2
C]
2
4
71 8 17
< 1 25 52
1 4
C]
C]
1 4
of Administered 2
8 9
1
< 1 4 71 14 8
