Ascorbic Acid: Chemistry, Metabolism, and Uses - American Chemical

19 Role of L-Ascorbic A c i d i n L i p i d Metabolism

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E. GINTER, P. BOBEK, and M. JURCOVICOVA Institute of H u m a n Nutrition Research, Bratislava, Czechoslovakia

The activity of the microsomal system containing cyto chrome P 4 5 0 that catalyzes 7α-hydroxylation of cholesterol is depressed in the livers of guinea pigs with marginal vitamin C deficiency. Slowing of the rate-limiting reaction of cholesterol transformation to bile acids causes cholesterol accumulation in the liver, plasma, and arteries; increase of plasma cholesterol half-life; decrease in the bile-acid body pool; atherosclerotic changes in coronary arteries; and cholesterol gallstone formation. In an ascorbate-deficient animal the plasma triglyceride level rises; the post-heparin plasma lipolytic activity decreases, and the half-life of plasma triglycerides increases, causing triglyceride accumu lation in the liver and arteries. In hypercholesterolemic humans with low vitamin C status, "L-ascorbic acid adminis tration (500-1000 mg/d) lowers plasma cholesterol concen tration. This effect may be reinforced through the simul taneous administration of agents that sequester bile acids.

T h e relationships among L-ascorbic acid, lipid metabolism, and atherosclerosis were first studied at extreme L-ascorbate levels, which are unlike the intake levels prevailing in human nutrition. Those investiga tions were done on animals that either biosynthesize ascorbate, such as rabbits or rats (1,2), or on acutely scorbutic guinea pigs (3,4). Ascor bate levels in the tissues of animals synthesizing L-ascorbic acid are saturated, and they are only slightly influenced by exogenous vitamin C. Therefore the effect of ascorbate on cholesterol metabolism in such animals is small (5). Disorders of lipid metabolism in scorbutic animals are, for the most part, nonspecific, because such animals refuse food and lose body weight rapidly. A more suitable model for our biochemical research is a guinea pig eating a diet marginally deficient in vitamin C (6). Guinea pigs are 0065-2393/82/0200-0381$06.00/0 © 1982 American Chemical Society Seib and Tolbert; Ascorbic Acid: Chemistry, Metabolism, and Uses Advances in Chemistry; American Chemical Society: Washington, DC, 1982.

382

ASCORBIC

ACID

g i v e n a v i t a m i n - C - f r e e d i e t f o r t w o w e e k s , w h i c h results i n a r a p i d d e c l i n e of t h e i r ascorbate b o d y p o o l , a l t h o u g h v i t a m i n C deficiency does not a p p e a r

outwardly.

T h e n a m a i n t e n a n c e dose of L - a s c o r b i c

acid

( 0 . 5 - 1 . 0 m g / 2 4 h / a n i m a l ) is i n i t i a t e d w i t h t h e o t h e r w i s e u n a l t e r e d d i e t . F o o d c o n s u m p t i o n a n d w e i g h t curves of t h e deficient a n i m a l s a r e s i m i l a r to those of t h e controls, w h i c h receive t h e same d i e t b u t w i t h a s u b s t a n t i a l l y h i g h e r i n t a k e of L - a s c o r b i c a c i d . V i t a m i n - C - d e f i c i e n t a n i m a l s Downloaded by CALIFORNIA INST OF TECHNOLOGY on September 18, 2017 | http://pubs.acs.org Publication Date: June 1, 1982 | doi: 10.1021/ba-1982-0200.ch019

a p p e a r to b e i n g o o d h e a l t h , b u t t h e ascorbate levels i n t h e i r tissues a r e permanently very low. G u i n e a p i g s c a n b e m a i n t a i n e d i n a state of m a r g i n a l v i t a m i n C deficiency f o r p r o t r a c t e d p e r i o d s , f o r e x a m p l e , f o r a w h o l e year.

This

l o n g e v i t y is s u i t a b l e f o r f o l l o w i n g u p m e t a b o l i c disorders of c o m p o u n d s w i t h l o n g b i o l o g i c a l h a l f - l i v e s a n d also f o r p u r s u i n g p a t h o p h y s i o l o g i c a l studies, s u c h as r e s e a r c h o n atherogenesis.

M o r e o v e r , t h e m o d e l of a

m a r g i n a l v i t a m i n C deficiency comes close t o t h e p r e v a i l i n g s i t u a t i o n i n m a n y p o p u l a t i o n g r o u p s , because t h e c o n s u m p t i o n of v i t a m i n C i n v a r i ous parts of t h e w o r l d l i k e w i s e reaches m a r g i n a l l i m i t s , e s p e c i a l l y d u r i n g the w i n t e r a n d s p r i n g .

"L-Ascorbic Acid and Cholesterol Metabolism I f g u i n e a p i g s a r e k e p t i n a state of m a r g i n a l v i t a m i n C d e f i c i e n c y for a p r o t r a c t e d p e r i o d , c h o l e s t e r o l a c c u m u l a t e s i n t h e i r livers a n d b l o o d p l a s m a , r e s u l t i n g i n a n e l e v a t e d r a t i o of t o t a l c h o l e s t e r o l / h i g h d e n s i t y lipoprotein ( H D L ) cholesterol ( T a b l e I ) . These data have been firmed

i n v i t a m i n - C - d e f i c i e n t g u i n e a p i g s (7-14).

con

A different c h o l e s t e r o l

turnover was f o u n d i n vitamin-C-deficient baboons, b u t the plasma cho lesterol l e v e l i n c r e a s e d o n l y w i t h a c o n c o m i t a n t stress ( 1 5 ) . T h e effect of a c u t e v i t a m i n C deficiency o n h u m a n p l a s m a c h o l e s t e r o l is s m a l l (16). T h e r e a r e n o d a t a a v a i l a b l e o n t h e effect of a c h r o n i c a l l y m a r g i n a l v i t a m i n C deficiency o n p l a s m a l i p i d s i n h u m a n s . H o w e v e r , h y p e r c h o l e s t e r o l e m i a is m o r e f r e q u e n t i n h u m a n s w i t h a l o w v i t a m i n C i n t a k e t h a n i n those a d e q u a t e l y s u p p l i e d w i t h ascorbate

(17-19).

E v e n rain

b o w t r o u t w i t h c h r o n i c v i t a m i n C deficiency a r e r e p o r t e d t o d e v e l o p a marked hypercholesterolemia ( 2 0 ) . R e s e a r c h o n t h e m e c h a n i s m of t h e onset of h y p e r c h o l e s t e r o l e m i a d u r i n g a state of m a r g i n a l v i t a m i n C d e f i c i e n c y (6,21,22) finding

has l e d t o t h e

t h a t ascorbate is necessary f o r c h o l e s t e r o l t r a n s f o r m a t i o n t o b i l e

acids ( 2 3 ) a t t h e r a t e - l i m i t i n g r e a c t i o n of b i l e - a c i d biosynthesis. l i m i t i n g step is the 7 a - h y d r o x y l a t i o n of c h o l e s t e r o l (6,24-26).

of ascorbate o n 7«-hydroxylation is n o t a d i r e c t o n e because a d d e d L - a s c o r b i c a c i d h a s n o effect (24,27).

That

T h e action i n vitro

T h e effect is m e d i a t e d b y

the i n t e r v e n t i o n of ascorbate i n t h e m e t a b o l i s m o f c y t o c h r o m e P the e n d o p l a s m a t i c r e t i c u l u m of t h e h e p a t a l c e l l (6,24).

4 5 0

in

Through a

Seib and Tolbert; Ascorbic Acid: Chemistry, Metabolism, and Uses Advances in Chemistry; American Chemical Society: Washington, DC, 1982.

19.

GINTER

383

L-Ascorbic Acid in Lipid Metabolism

E T A L .


Table I. Effect of a Chronic Marginal Vitamin C Deficiency (19 Weeks on a Diet w i t h 1 5 % Predominantly Saturated Fat and 0 . 0 3 % Cholesterol) on Blood Serum Lipids in Sated Male Guinea Pigs Control Group (0.5% L-ascorbic acid in diet)

Vitamin C Deficiency (0.5 mg of L-ascorbic acid/24 h/ animal)

B o d y w e i g h t (g)

700 ± 14 (27)

681 ± 2 0 (27)

NS

V i t a m i n C i n the l i v e r ( m m o l / k g of fresh tissue)

1.86 ± 0.10 (27)

0.22 ± 0.03 (27)

P < 0.001

T o t a l s e r u m cholesterol (mmol/L)

1.78 ± 0.11 (27)

3.34 ± 0.19 (27)

P < 0.001

0.52 ± 0.04 (15)

0.44 ± 0.03 (20)

NS

4.2 ± 0.5 (15)

9.0 ± 0.8 (20)

P < 0.001

1.76 ± 0.27

4.90 ± 0.43

P < 0.001

Parameter

H D L cholesterol i n s e r u m

Ratio-

total HDL

0

-cholesterol

Serum triglyceridesz (mmol/L)

Statistical Significance (student's t-test)

Note: Data presented as mean ± se. Figures in parentheses indicate numbers of animals analyzed. NS = not significant. H D L cholesterol was determined by the dextran sulfate precipitation method. a

s i m i l a r m e c h a n i s m , ascorbate b i o t i c s (28). P450,

also affects

t h e h y d r o x y l a t i o n of

T h e exact m e c h a n i s m of L - a s c o r b i c a c i d o n

i n spite of i n t e n s i v e r e s e a r c h (28-31),

xeno-

cytochrome

is as y e t u n k n o w n . T h e

h y p o c h o l e s t e r o l e m i c effect of L - a s c o r b a t e 2-sulfate (10) has b e e n a s c r i b e d to its c a p a c i t y to t r a n s f o r m c h o l e s t e r o l t o t h e m o r e w a t e r - s o l u b l e c h o lesterol sulfate (32).

H o w e v e r , f r o m a q u a n t i t a t i v e aspect, this process

is of m i n o r significance i n t o t a l c h o l e s t e r o l t u r n o v e r

(33,34).

I n g u i n e a p i g s w i t h a m a r g i n a l v i t a m i n C deficiency, t h e d e c r e a s e d t r a n s f o r m a t i o n of c h o l e s t e r o l t o b i l e a c i d s p r o v o k e s a series of p a t h o logical changes:

hypercholesterolemia a n d cholesterol accumulation i n

the l i v e r , w h i c h has a l r e a d y b e e n d i s c u s s e d ; increase of p l a s m a c h o l e s t e r o l h a l f - l i f e (6);

increase i n c h o l e s t e r o l c o n c e n t r a t i o n i n g a l l b l a d d e r

b i l e ( 3 5 ) ; decrease i n t h e b i l e - a c i d b o d y p o o l a n d l o w e r e d e x c r e t i o n of b i l e a c i d s i n t h e s t o o l (26,36);

decrease of b i l e - a c i d c o n c e n t r a t i o n i n

g a l l b l a d d e r b i l e w i t h t h e r e s u l t i n g f o r m a t i o n of c h o l e s t e r o l

gallstones

( 5 , 3 5 ) ; c h o l e s t e r o l a c c u m u l a t i o n i n t h e a o r t a ; a n d atherosclerosis 37-41).

(7,9,

O p t i m u m p r e v e n t i o n of these d i s o r d e r s m a y b e a c h i e v e d b y


384

ASCORBIC

ACID

doses of L - a s c o r b i c a c i d t h a t a r e c a p a b l e of e n s u r i n g a m a x i m u m steady state l e v e l ( s a t u r a t i o n ) of ascorbate i n t h e tissues

(42).

L-Ascorbic Acid and Triglyceride Metabolism Marked

h y p e r t r i g l y c e r i d e m i a occurs i n ascorbate-deficient

guinea

pigs ( T a b l e I ) w i t h triglycerides accumulating i n the liver a n d the aorta F r o m a k i n e t i c p o i n t of v i e w , t h e reason f o r h y p e r t r i


(7,8,39,42,43).

g l y c e r i d e m i a m a y b e e i t h e r a n i n c r e a s e d i n p u t or a d e c r e a s e d o u t p u t of triglycerides from the plasma pool.

A s t u d y of t h e secretion

rate of

e n d o g e n o u s t r i g l y c e r i d e s f o l l o w i n g a b l o c k a d e of l i p o p r o t e i n l i p a s e w i t h T r i t o n W R 1339 has s h o w n that t h e t r i g l y c e r i d e i n p u t i n g u i n e a p i g s w i t h m a r g i n a l v i t a m i n C d e f i c i e n c y is l o w e r e d b y 2 5 % (44).

Simultaneously,

h o w e v e r , the rate of t r i g l y c e r i d e o u t p u t f r o m the p l a s m a p o o l , d e t e r m i n e d b y a d e c l i n e of r a d i o a c t i v i t y of e n d o g e n o u s l y l a b e l e d H - t r i g l y c e r i d e s , is 3

even more substantially reduced

(Figure 1).

T h e rate of t r i g l y c e r i d e o u t p u t f r o m t h e p l a s m a is m a i n l y c o n t r o l l e d by

t h e l i p o p r o t e i n l i p a s e a c t i v i t y of p e r i p h e r a l tissues.

plasma

Post-heparin

lipolytic activity i n vitamin-C-deficient guinea pigs

considerably.

decreases

I n a d d i t i o n , i n some of t h e a n i m a l s t h e response of t h e

p l a s m a l i p o l y t i c a c t i v i t y t o i n t r a v e n o u s l y a d m i n i s t e r e d h e p a r i n is also prolonged

( 4 5 ) . S i m i l a r results h a v e b e e n r e p o r t e d i n v i t a m i n - C - d e f i

cient baboons

(46).

Lipoprotein

lipase activity determined

i n acetone p o w d e r s

from

g u i n e a p i g tissues w i t h a m a r g i n a l v i t a m i n C d e f i c i e n c y w a s n o t g r e a t l y a l t e r e d (47). O n t h e other h a n d , l i p o p r o t e i n l i p a s e a c t i v i t y i n e p i d i d y m a l fat, f o l l o w i n g i n c u b a t i o n of t h e tissue w i t h h e p a r i n , d r o p p e d

abruptly

i n v i t a m i n - C - d e f i c i e n t g u i n e a p i g s , a n d this decrease w a s i n g o o d agree ment w i t h the enhanced triglyceride concentration i n blood serum ( F i g u r e 2 ) . H e n c e , v i t a m i n C d e f i c i e n c y m a y affect l i p o p r o t e i n l i p a s e release from capillaries b y influencing the h e p a r i n - l i p o p r o t e i n lipase interaction. T h i s a s s u m p t i o n w o u l d agree w i t h t h e fact that h i g h doses of L - a s c o r b i c a c i d depress h y p e r t r i g l y c e r i d e m i a i n v a r i o u s a n i m a l species, t h e h y p o t r i glyceridemic systems

effect

(2,7,43,45).

being

associated

However

with

a s t i m u l a t i o n of

t h e i n h i b i t o r y effect

lipolytic

i n vitro

added

L - a s c o r b i c a c i d o n l i p o p r o t e i n l i p a s e i n t h e h e a r t (46) a n d o n h o r m o n e sensitive l i p a s e i n t h e a d i p o s e tissue (48) is u n e x p l a i n e d .

L-Ascorbic Acid and Hypercholesterolemia in Humans I n contrast t o t h e u n e q u i v o c a l results o b t a i n e d i n g u i n e a p i g s , d a t a o n t h e effect of L - a s c o r b i c a c i d o n c h o l e s t e r o l e m i a i n h u m a n s differ c o n s i d e r a b l y (5,22,49).

S o m e authors (2,6,11,50-53)

l e s t e r o l e m i c effect, w h i l e others refute i t (54-60).

found a hypochoThose contradictions


Seib and Tolbert; Ascorbic Acid: Chemistry, Metabolism, and Uses Advances in Chemistry; American Chemical Society: Washington, DC, 1982. 3

Figure 1. Removal of endogenously labeled H-triglycerides from blood plasma in control male guinea pigs (C, 0.5% L-ascorbic acid in diet) and in animals of equal weight with a marginal vitamin C deficiency (A, 0.5 mg of L-ascorbic acid/animal/d).


Seib and Tolbert; Ascorbic Acid: Chemistry, Metabolism, and Uses Advances in Chemistry; American Chemical Society: Washington, DC, 1982. 0 2 2 4 x

•

DEFICIENCY

LIPASE H m o l 1% j m i n

LIPOPROTEIN

CONTROL

Figure 2. A negative correlation between lipoprotein lipase activity in epididymal fat tissue and triglyceride concentration in blood serum of conrtol male guinea pigs (0.5% L-ascorbic acid in diet) and in guinea pigs with a marginal vitamin C deficiency (0.5 mg of L-ascorbic acid/animal/d). Equation of the curve, y = 7.91 e .

2

g

IV

£

O

IV

i

230 m g % 3. h e a l t h y people w i t h m i l d h y p e r cholesterolemia 4. pensioners w i t h i n i t i a l choles t e r o l e m i a > 200 m g % 5. c h r o n i c i n p a t i e n t s 6. a l c o h o l i c i n p a t i e n t s 7. students 8. students 9. students 10. students 11. pensioners w i t h i n i t i a l choles t e r o l e m i a < 200 m g % 12. h y p e r l i p e m i c o u t p a t i e n t s 13. h e a l t h y people w i t h m i l d h y p e r cholesterolemia

Number of Subjects

Dose (mg/d)

Duration 1 year

35

500

19

1000

24

300

46 39 14 20 20 20 20

1000 900 900 100 200 500 2000

3 3 3 8 8 8 8

36 11

1000 450°

3 months 6 weeks

21

450°

3 months 7 weeks months weeks weeks weeks weeks weeks weeks

6 weeks

Note: Regression line is obtained for mean values from the thirteen groups. Plus 15 g of citrus pectin/d. Source: Reproduced with permission, from Ref. 61.

a


388

ASCORBIC

ACID


•20

Figure 3. Mean change in total serum cholesterol concentration after L-ascorbic acid treatment vs. initial serum cholesterol levels. Experimental conditions are given in Table II. w i t h a m a r g i n a l v i t a m i n C deficiency a n d i n h y p e r c h o l e s t e r o l e m i c betics w i t h v e r y l o w v i t a m i n C status ( 3 8 ) .

dia

T h e d e c l i n e of t o t a l choles

t e r o l c o n c e n t r a t i o n after ascorbate t r e a t m e n t is c a u s e d b y a d e c l i n e of L D L c h o l e s t e r o l , for the a m o u n t of the p r o t e c t i v e H D L does not c h a n g e or m a y e v e n rise w i t h a n i n c r e a s e d i n t a k e of v i t a m i n C However,

( 5 3 , 62, 63).

t h e a s c o r b a t e - s t i m u l a t e d c h o l e s t e r o l t r a n s f o r m a t i o n to

a c i d s leads to a n i n c r e a s e d b i l e - a c i d p o o l

bile

( 2 6 , 3 6 ) , a n d the i n c r e a s e d

q u a n t i t i e s of b i l e acids r e t u r n i n g v i a the e n t e r o h e p a t a l r e c i r c u l a t i o n b a c k to the l i v e r m a y s l o w d o w n t h e 7«-hydroxylation of c h o l e s t e r o l .

Conse

q u e n t l y this f e e d b a c k effect m u s t b e e l i m i n a t e d i f h y p e r c h o l e s t e r o l e m i a is to b e l o w e r e d .

Synergism Between L-Ascorbic Acid and Substances Capable of Binding Bile Acids O n e p o s s i b l e m e t h o d to i n t e r r u p t the f e e d b a c k effect of b i l e acids i n the l i v e r is to a d m i n i s t e r , a l o n g w i t h v i t a m i n C , c h o l e s t y r a m i n e ( Q u e s t r a n ) , a s y n t h e t i c r e s i n t h a t b i n d s b i l e acids i n the g a s t r o i n t e s t i n a l tract. I f a m o d e r a t e h y p e r c h o l e s t e r o l e m i a is p r o v o k e d i n g u i n e a p i g s t h r o u g h a m a r g i n a l v i t a m i n C deficiency a n d t h e n 1.5% of Q u e s t r a n (i.e., 0 . 6 6 % of c h o l e s t y r a m i n e ) is a d d e d to t h e i r d i e t , t h e l e v e l of c h o l e s t e r o l i n b l o o d s e r u m r e m a i n s unaffected. T h e s i m u l t a n e o u s a d m i n i s -


19.

GINTER

ET

t r a t i o n of 1.5% provokes

389

L-Ascorbic Acid in Lipid Metabolism

AL.

Questran w i t h 0 . 5 % L-ascorbic a c i d i n the diet, however,

a d e c l i n e of c h o l e s t e r o l e m i a

(Table III).

c h o l e s t y r a m i n e to t h e d i e t is i n c r e a s e d to 1.0% hypocholesterolemic

I f t h e a d d i t i o n of

(2.3%

Q u e s t r a n ) , its

effect b e c o m e s e v i d e n t e v e n i n g u i n e a p i g s w i t h a

m a r g i n a l i n t a k e of v i t a m i n C , a n d a g a i n t h e s i m u l t a n e o u s a d d i t i o n of 0 . 5 % L-ascorbic a c i d brings about a more substantial decline i n choles t e r o l e m i a ( T a b l e I V ) . A s y n e r g e t i c effect is i n v o l v e d b e c a u s e the d e c l i n e


i n c h o l e s t e r o l e m i a exceeds t h e s u m of t h e h y p o c h o l e s t e r o l e m i c cholestyramine

a n d of L - a s c o r b i c

acid when

administered

effect of

separately.

T h e ascorbate l e v e l i n the tissues of these a n i m a l s shows t h a t c h o l e s t y r a m i n e does not affect ascorbate a b s o r p t i o n f r o m the g a s t r o i n t e s t i n a l t r a c t (Table I V ) . E v i d e n c e has a c c u m u l a t e d

over

the p a s t f e w

years t h a t c e r t a i n

c o m p o n e n t s of d i e t a r y fiber, for e x a m p l e , p e c t i n , also h a v e t h e a b i l i t y t o b i n d b i l e acids i n the g a s t r o i n t e s t i n a l t r a c t (64). citrus p e c t i n a n d 0.5%

L-ascorbic

A n a d d i t i o n of

a c i d to a h i g h - f a t d i e t

c h o l e s t e r o l a c c u m u l a t i o n i n the b l o o d

5%

prevented

s e r u m a n d l i v e r of g u i n e a

pigs

A significant d e c l i n e of t o t a l c h o l e s t e r o l l e v e l i n t h e s e r u m of a

(65).

g r o u p of h e a l t h y subjects w i t h m i l d h y p e r c h o l e s t e r o l e m i a , a n d also i n a g r o u p of h y p e r l i p e m i c o u t p a t i e n t s , w a s a c h i e v e d t h r o u g h t h e a d m i n i s t r a t i o n of a g r a n u l a t e d p r e p a r a t i o n c o n t a i n i n g 15 g of c i t r u s p e c t i n a n d 450 m g of L - a s c o r b i c a c i d g i v e n d a i l y f o r 6 w e e k s (65) a n d 13 i n T a b l e I I ) .

(see

G r o u p s 12

S i n c e the H D L c h o l e s t e r o l l e v e l d i d not

change,

the d e c l i n e n o t e d w a s c a u s e d b y t h e d e c l i n e i n t h e r i s k - c o n s t i t u t i n g LDL

cholesterol.

Table III. Effect of L-Ascorbic A c i d and Questran on T o t a l Cholesterol Concentration in Blood Serum of Male Guinea Pigs Statistical Intake of L-Ascorbic Acid SignifiWeeks

Type of Diet

0

Starting values (cereal + vegetables)

9

Scorbutogenic d i e t

13

Scorbutogenic d i e t 1.5% Q u e s t r a n

+

Scorbutogenic d i e t 1.5% Q u e s t r a n

+

cance Low High --• (0.5 mg/24 h) (0.5% in diet) (*t»dent s t-test) 1.58 ± 0 . 1 3 (10) a

2.07 ± 0.13 (10)

1.55 ± 0.13 (10)

P

1.04 ± 0.08 (10)

P

Ascorbic Acid: Chemistry, Metabolism, and Uses - American Chemical

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