Nutritional Bioavailability of Zinc - American Chemical Society

12. ERDMAN ET AL. Zinc Bioavailability from Soybean Products 175 mixture of soy protein and egg white. After 21 days, rats are killed and either tibia...
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Zinc Bioavailability from Processed Soybean Products J O H N W . E R D M A N , JR., R I C H A R D M . F O R B E S , and HIROMICHI KONDO 1

University of Illinois, Departments of Food Science and Animal Science, Urbana, I L 61801

Soybean foods were u t i l i z e d as models for the evaluation of the effects of processing conditions upon the relative b i o a v a i l a b i l i t y of both endogenous and added zinc. Products such as f u l l - f a t soybean flour, soy beverage, soy concentrates, soy isolates and tofu were processed under carefully controlled conditions and were individually incorporated into diets for rat zinc bioassay studies. Some products were tested under various i n vitro conditions to determine the factors that affect the binding of zinc to components of the soy products. The overall conclusions from these studies are that phytate-to-zinc molar ratios alone do not predict zinc b i o a v a i l a b i l i t y and food processing conditions markedly affect the b i o a v a i l a b i l i t y of zinc endogenous to soy foods. Inorganic zinc (ZnCO ) added (fortified) to soy product-containing diets was highly available. 3

Often overlooked i n the evaluation of the effects of diet upon mineral a v a i l a b i l i t y i s the role that food processing plays i n the formation of or breaking of ligand-metal complexes. Several individual or unit processing steps are needed to produce a soy concentrate, a bread or a spray-dried egg white. Some or a l l steps may have a bearing upon f i n a l mineral b i o a v a i l a b i l i t y . Soy concentrate from company A i s not produced i n precisely the same manner as from company B. In fact, l o t to l o t variation for the same product may be quite variable, particularly i n mineral content. Rackis and coworkers (1_,2) noted from their survey of the literature that for experimental animals, zinc b i o a v a i l a b i l i t y from soy protein isolates was low compared to zinc a v a i l a b i l i t y from soybean meal, casein or other animal protein diets. They 1

Current address: Kinjo Gakuin University, Nagoya, Japan 0097-6156/83/0210-0173$06.00/0 © 1983 American Chemical Society In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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a t t r i b u t e d the d i f f e r e n c e s i n a v a i l a b i l i t y t o formation o f p h y t a t e - p r o t e i n - m i n e r a l complexes during the processing o f i s o l a t e s . Other r e s e a r c h e r s , notably Lease (3,k) , O'Dell (5_,6) and Forbes (7_,8) have reported experimental r e s u l t s that i n d i c a t e d the v a r i a b l e nature o f z i n c b i o a v a i l a b i l i t y from soy p r o t e i n products. Oberleas (9^) f i r s t suggested that the molar r a t i o o f phytate to z i n c might be u s e f u l f o r p r e d i c t i o n o f the z i n c b i o a v a i l a b i l i t y from p h y t a t e - r i c h foods. Molar r a t i o s of greater than 20:1 seemed to be i n d i c a t i v e o f poorly a v a i l a b l e z i n c . O'Dell (10), M o r r i s and E l l i s (11) and Davies and O l p i n (12) have a l l pointed out the importance o f the calcium content o f the d i e t t o the phytate t o z i n c molar r a t i o . Higher d i e t a r y calcium c l e a r l y depresses z i n c b i o a v a i l a b i l i t y a t phytate t o z i n c molar r a t i o s of l e s s than 20:1 i n d i e t s fed t o r a t s . Use o f the phytate t o z i n c molar r a t i o may be too s i m p l i s t i c for p r e d i c t i o n o f z i n c a v a i l a b i l i t y from mixed food systems. Besides the e f f e c t o f d i e t a r y calcium l e v e l , the amount o f i r o n , and perhaps the l e v e l o f other metals, may a f f e c t z i n c b i o a v a i l a b i l i t y . Solomons and Jacob (13) have shown i n human subjects that i n c r e a s i n g the i r o n / z i n c r a t i o from 0:1 t o 3:1 i n s o l u t i o n s c o n t a i n i n g 25 mg o f z i n c and corresponding amounts o f i r o n as ferrous s u l f a t e produced a progressive decrease i n the plasma z i n c response. They f u r t h e r reported that the chemical form of i r o n was an important determinant o f the i n t e r a c t i o n . Solomons (14) e x t e n s i v e l y reviewed both i n h i b i t o r y f a c t o r s and enhancers o f z i n c b i o a v a i l a b i l i t y found i n foods. Our l a b o r a t o r i e s have been concerned w i t h the r o l e that u n i t food processing operations play i n the b i o a v a i l a b i l i t y o f z i n c from complete d i e t s . Soybean foods have served as models f o r the e v a l u a t i o n o f processing e f f e c t s upon both endogenous and added z i n c . Below are described r e s u l t s from both r a t bioassays and i n v i t r o tests for zinc b i o a v a i l a b i l i t y . P r e d i c t i o n of zinc b i o a v a i l a b i l i t y from soy-containing d i e t s i s f a r more complex than an a n a l y s i s o f phytate and z i n c molar r a t i o s . Methods For most o f the i n v i v o t e s t s o f z i n c b i o a v a i l a b i l i t y reported here, the slope r a t i o technique suggested by Shah and coworkers (15) and f u r t h e r developed i n our l a b o r a t o r i e s (16,17) was u t i l i z e d . B r i e f l y , isonitrogenous, i s o e n e r g e t i c 20% p r o t e i n d i e t s are fed t o groups o f weanling r a t s f o r 21 days. C o n t r o l d i e t s c o n t a i n i n g 20% egg white p r o t e i n are supplemented w i t h one of s e v e r a l l e v e l s o f z i n c as z i n c carbonate. To t e s t f o r the a v a i l a b i l i t y o f z i n c i n a t e s t soy p r o t e i n product, s e v e r a l l e v e l s of z i n c are added t o egg white-based d i e t s by s u b s t i t u t i n g the soy product f o r egg-white on an equivalent p r o t e i n b a s i s . To t e s t f o r the e f f e c t o f the presence o f a soy p r o t e i n product i n a d i e t upon the b i o a v a i l a b i l i t y o f z i n c added ( f o r t i f i e d ) t o d i e t s , z i n c carbonate i s added a t v a r i o u s l e v e l s t o d i e t s containing a s p e c i f i c

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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Zinc Bioavailability from Soybean Products 175

mixture of soy p r o t e i n and egg white. A f t e r 21 days, r a t s are k i l l e d and e i t h e r t i b i a s or femurs are removed. To provide a measure of r e l a t i v e z i n c a v a i l a b i l i t y (15-17), the data f o r the weight gain and f o r the t o t a l bone z i n c are s t a t i s t i c a l l y analyzed by r e g r e s s i o n a n a l y s i s to compare the slopes of the l i n e a r p o r t i o n s of the l i n e s r e l a t i n g response per u n i t of added m i n e r a l . For i n v i t r o t e s t s the method developed by M i l l e r and coworkers~Tl8) to estimate i r o n a v a i l a b i l i t y from meals was adapted f o r z i n c t e s t i n g . We have a l s o examined the s o l u b i l i t y of z i n c i n aqueous e x t r a c t s o f soy products by the methods published by de Rham and J o s t (19). Results and D i s c u s s i o n B i o a v a i l a b i l i t y of Zinc I n t r i n s i c to Soybean Products and Added to Soybean Products. A number of s t u d i e s (16,17,20) was Table I R e l a t i v e B i o a v a i l a b i l i t y of Zinc I n t r i n s i c to Soy

Product

F u l l - f a t soy f l o u r Freeze-dried soy beverage Spray-dried soy concentrate ( n e u t r a l form) Freeze-dried soy concentrate ( n e u t r a l form) Freeze-dried soy isolate ( n e u t r a l form) Freeze-dried soy concentrate ( a c i d form) Freeze-dried soy isolate ( a c i d form)

Phytate-toz i n c molar r a t i o

Products

Relative Bioavailability (%) Weight gain Log bone z i n c a

34 40

b

b

28 26

55 63

52

41

b

20

b

57

66

b

29

b

34

85

46

b

48

b

64

b

66

35

b

113,93

106

c

b

a

D a t a p o i n t s represent comparisons of slopes of responses of t e s t d i e t s ( c o n t a i n i n g soy) w i t h responses of c o n t r o l d i e t s (without soy products) w i t h added minerals as the carbonate X 100. (Sources: 16,17 ,20). S i g n i f i c a n t l y d i f f e r e n t from c o n t r o l d i e t s (P < 0.05). e x p e r i m e n t was* run twice.

b

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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performed to i n v e s t i g a t e the b i o a v a i l a b i l i t y to the r a t of z i n c i n t r i n s i c to v a r i o u s soybean products (Table I ) . The r e l a t i v e b i o a v a i l a b i l i t y of z i n c from soy products was h i g h l y v a r i a b l e but u s u a l l y low when compared to the a v a i l a b i l i t y of z i n c from z i n c carbonate. » The phytate-to-zinc molar r a t i o was a poor p r e d i c t o r of zinc b i o a v a i l a b i l i t y . The a c i d forms of the i s o l a t e s and concentrates demonstrated e x c e l l e n t b i o a v a i l a b i l i t y f o r z i n c r e l a t i v e t o n e u t r a l i z e d products prepared under i d e n t i c a l c o n d i t i o n s (20). The d i f f e r e n c e may be due to the formation of s t a b l e p r o t e i n - p h y t i c a c i d - z i n c complexes i n the d r i e d n e u t r a l product. P r o t e i n - p h y t i c acid-mineral a s s o c i a t i o n s have been shown to occur i n s o l u t i o n at a n e u t r a l pH (19) . These a s s o c i a t i o n s may w e l l form more tightly-bound complexes during the d r y i n g of the soy p r o t e i n . The e x c l u s i o n of water from the p r o t e i n - p h y t i c a c i d - z i n c a s s o c i a t i o n s could lead to therraodynamically s t a b l e complexes that are r e s i s t a n t to complete p r o t e o l y t i c d i g e s t i o n i n the g a s t r o i n t e s t i n a l t r a c t . Short peptides or amino a c i d residues bound to z i n c and p h y t i c a c i d then would be poorly absorbed. For f u r t h e r d i s c u s s i o n see Erdman et a l . (20) and Cheryan (21). Results from studies i n v e s t i g a t i n g the e f f e c t of the presence of soy products upon the b i o a v a i l a b i l i t y of exogenous ( f o r t i f i e d ) z i n c , added as the carbonate are shown i n Table I I . The r e s u l t s demonstrate that the presence of soybean products i n complete d i e t s Table I I E f f e c t of Soy Products Upon the R e l a t i v e B i o a v a i l a b i l i t y of E x t r i n s i c Zinc Added as Zinc Carbonate Product

F u l l - f a t soy Flour Spray-dried soy concentrate ( n e u t r a l form) Freeze-dried soy concentrate ( n e u t r a l form) Freeze-dried soy concentrate ( a c i d form)

Phytate-toz i n c molar r a t i o

a

Relative B i o a v a i l a b i l i t y (%) Weight gain Log bone z i n c

94

28

94

52

77

57

84

7.8

66

65

83

b

70

l

b

Data p o i n t s represent comparisons of slopes of responses of t e s t d i e t s ( c o n t a i n i n g soy) w i t h responses of c o n t r o l d i e t s (without soy products) w i t h added minerals as t h e i r carbonate X 100. (Sources: 16,17 ,20). S i g n i f i c a n t l y d i f f e r e n t from c o n t r o l d i e t s (P < 0.05).

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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Zinc Bioavailability from Soybean Products 111

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has l i t t l e d e t r i m e n t a l e f f e c t upon the b i o a v a i l a b i l i t y of added z i n c . Other workers, i n c l u d i n g Hardie-Muncy and Rasmussen (22), have a l s o reported that i n o r g a n i c z i n c added to soy i s o l a t e s to be b e t t e r u t i l i z e d by r a t s than z i n c i n t r i n s i c to the i s o l a t e . Therefore, these s t u d i e s suggest that f o r t i f i e d z i n c i n soy products i s of h i g h b i o a v a i l a b i l i t y r e l a t i v e to i n t r i n s i c z i n c . Recently, our l a b o r a t o r i e s and Weaver and coworkers from Purdue U n i v e r s i t y have begun a c o l l a b o r a t i v e e f f o r t to i n v e s t i g a t e the b i o a v a i l a b i l i t y of z i n c from v a r i o u s soy products u t i l i z i n g i n t r i n s i c a l l y - l a b e l e d and e x t r i n s i c a l l y - l a b e l e d 6 5 £ products. The 6 5 £ i n t r i n s i c a l l y l a b e l e d soybeans were obtained by growing Century v a r i e t y soybeans h y d r o p o n i c a l l y as described by Levine et a l . (23). " 5 z i n c i n t r i n s i c a l l y l a b e l e d soybeans were d e f a t t e d . A c i d and n e u t r a l i z e d soy concentrates were prepared as p r e v i o u s l y described (20). P r e l i m i n a r y r e s u l t s of a r a t feeding study are shown i n Table I I I ( K e t e l s e n , et a l . , manuscript i n prep.). Groups of male weanling r a t s were fed experimental unlabeled d i e t s c o n t a i n i n g 10 ppm z i n c from d e f a t t e d soy f l o u r , a c i d - p r e c i p i t a t e d soy concentrate or n e u t r a l i z e d soy concentrate f o r 7 days. On the evening of the seventh day, each r a t was f a s t e d and then given a 6 5 - [ intrinsically-labeled t e s t meal which was s i m i l a r to the d i e t fed f o r the l a s t seven days. A f t e r the meal the r a t s were i n d i v i d u a l l y placed i n t o a whole body gamma counting chamber and the 6 5 i c a c t i v i t y was recorded and designated as the day 0 a c t i v i t y . Rats were then refed the same unlabeled experimental d i e t s as had been fed p r i o r to the t e s t meal. Whole body ^ ^ z i n c a c t i v i t y was measured 24 hours and 12 days a f t e r a d m i n i s t r a t i o n of the t e s t meal. Table I I I shows the percent whole body r e t e n t i o n from the three t e s t meals a f t e r 1 and 12 days. The r e s u l t s v e r i f y r e s u l t s from Table I , demonstrating the v a r i a b l e b i o a v a i l a b i l i t y of z i n c from d i f f e r e n t soy products. The r e t e n t i o n of z i n c from the meal c o n t a i n i n g the a c i d - p r e c i p i t a t e d soy concentrate was g r e a t e r than from the n e u t r a l z

z

n c

s o v

n c

z

nc

z

n

Table I I I Whole Body Retention of ^Zinc from D e f a t t e d Soy F l o u r , A c i d P r e c i p i t a t e d - and N e u t r a l i z e d Soy Concentrate-Based Meals (%)

Diet

Defatted soy f l o u r A c i d concentrate N e u t r a l concentrate

Day 1 (mean + S.D.)

83.1 + 4.1 75.9 + 5.5 64.7 + 4.4

a b e

Day 12 (mean + S.D.)

75.5 + 4.0 68.0 + 5.0 52.3 + 4.4

l

a b e

N = 6 or 7, In v e r t i c a l columns, means not sharing a common s u p e r s c r i p t l e t t e r are s i g n i f i c a n t l y d i f f e r e n t (P < 0.05). (Source: Keteleson et a l . , manuscript i n prep.).

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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product, again suggesting that n e u t r a l i z a t i o n of soy p r o t e i n r e s u l t s i n formation o f protein-phytate-mineral complexes. As pointed out e a r l i e r i n t h i s review, i n c r e a s i n g the l e v e l o f d i e t a r y calcium decreases the z i n c b i o a v a i l a b i l i t y from phytate-containing foods. Presumably the mechanism i s through the formation of chemical complexes c o n t a i n i n g z i n c , phytate and calcium which are i n s o l u b l e a t i n t e s t i n a l pH and nonabsorbable (24). Recently, our l a b o r a t o r i e s used slope r a t i o techniques t o compare the b i o a v a i l a b i l i t y o f z i n c contained i n calcium s u l f a t e and i n magnesium c h l o r i d e - p r e c i p i t a t e d soybean curd (Tofu) t o that o f z i n c added as the carbonate t o egg white d i e t s by slope r a t i o techniques (25). T o t a l d i e t a r y calcium l e v e l i n a l l d i e t s was adjusted t o 0.7% w i t h calcium carbonate. The r e s u l t s (not shown) i n d i c a t e d that the r e l a t i v e a v a i l a b i l i t y of z i n c from both t o f u preparations was 51% as measured by weight gain and 36-39% f o r bone z i n c . These r e s u l t s are s i m i l a r to those reported f o r f u l l f a t soy f l o u r (16) i n Table I . In a separate experiment w i t h z i n c s u p p l i e d a t 9 ppm i n a l l d i e t s , the e f f e c t o f i n c r e a s i n g d i e t a r y calcium i n calcium or magnesium p r e c i p i t a t e d t o f u or egg white d i e t s on weight gain and t i b i a z i n c accumulation was t e s t e d . From Table IV i t can be noted that the performance o f t o f u - f e d r a t s r e l a t i v e t o z i n c carbonate-fed r a t s was q u i t e s i m i l a r at 0.4% t o t a l d i e t a r y calcium, but was reduced as d i e t a r y calcium was increased t o 0.7% and t o 1.2%. These r e s u l t s suggest that a p o o r l y a v a i l a b l e calciura-zinc-phytate complex not present i n the soy curd can form i n the g a s t r o i n t e s t i n a l t r a c t when s u f f i c i e n t calcium i s added to the d i e t . Table IV E f f e c t o f D i e t a r y Calcium L e v e l Upon Weight Gain and T i b i a Zinc of Rats fed 9ppm Zinc from Calcium o r Magnesium P r e c i p i t a t e d Tofu or from Zinc Carbonate Added t o Egg White D i e t s D i e t a r y Calcium L e v e l

Diet

Egg Ca-Tofu Mg-Tofu

Wt. gain (g)

0.4% 0J% T i b i a Zinc Wt. T i b i a Zinc (yg/2 t i b i a ) gain (yg/2 t i b i a ) (g)

117 + 5 112 + 7 114+5

50 + 2 34 + 2 37 + 2

a b b

105 +_ 8 75 +_ 5 70 + 8

a b b

46 +_ l 25 + l 28 + l

a b b

Wt. gain (g)

ΙΛ% T i b i a Zinc (yg/2 t i b i a )

95 +_ l l 46 +_ 5 46 + 3

a b b

l

45 + 2 24 + 4 25 + l

Means +_ SEM. For columns, means not sharing a common s u p e r s c r i p t l e t t e r are d i f f e r e n t (P < 0.05) (Source: 25).

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

a b b

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Zinc Bioavailability from Soybean Products

Calcium E f f e c t s on Zinc B i o a v a i l a b i l i t y f o r the Rat and the Human. I t should be pointed out at t h i s juncture that the n u t r i e n t requirement of calcium f o r the r a t i s much higher than f o r man. In f a c t , the molar r a t i o of c a l c i u m t o z i n c i n excess o f 660:1 i s recommended f o r r a t d i e t s , w h i l e f o r man the r a t i o i s between 80:1 and 160:1. To feed r a t s molar r a t i o s o f calcium and z i n c s i m i l a r to human requirements would n e c e s s i t a t e e i t h e r a very calcium d e f i c i e n t d i e t or one c o n t a i n i n g z i n c at a l e v e l w e l l i n excess o f the requirement. Neither choice i s n u t r i t i o n a l l y s u i t a b l e f o r demonstrating an e f f e c t o f phytate on z i n c a v a i l a b i l i t y . The calcium t o f u s prepared f o r the study found i n Table IV contained a c a l c i u m - t o - z i n c molar r a t i o o f 330:1 t o 355:1. This r a t i o i s higher than would be a t t a i n e d i n a balanced d i e t f o r humans, but i t i s lower than the 700:1 r a t i o present i n the 0.4% calcium d i e t fed t o the r a t s . Since t h i s l a t t e r r a t i o had a minimal e f f e c t on z i n c a v a i l a b i l i t y i n r a t s one would expect the z i n c from a meal o f calcium t o f u t o be w e l l u t i l i z e d by humans. This assumes, o f course, that the r a t and the human respond s i m i l a r l y t o these molar r a t i o s . Table V In V i t r o A v a i l a b i l i t y of Zinc from Various Soy P r o d u c t s

Soy Product

Mature soybean F u l l - f a t soy flour Dehulled soybean flour Green soybean Soy beverage Spray-dried, neutralized soy concentrate Low phytate soy concentrate Normal phytate soy concentrate A c i d concentrate N e u t r a l concentrate Calcium t o f u Magnesium t o f u

a

Method: et a l .

3

Calcium t o z i n c molar ratio

Dialyzability (%)

35 29

73 81

26.71 25.13

34

88

11.03

18 25 52

150 86 90

9.51 6.44 4.70

13

440

3.06

44

156

4.12

35 33 28 26

87 83 359 38

1.20 0.60 1.32 0.85

Phytate to z i n c molar ratio

D. D. M i l l e r et a l . (18).

Unpublished dat a of Kondo

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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We would have to question the v a l i d i t y of the use of d i e t a r y calcium l e v e l s of up to 1.75% calcium i n rat studies to study z i n c bioavailability. These l e v e l s would never be approached i n human d i e t s without use of excessive calcium supplementation. In V i t r o Tests f o r Zinc B i o a v a i l a b i l i t y . In v i t r o d i g e s t i b i l i t y t e s t i n g can y i e l d v a l u a b l e information regarding the s o l u b i l i t y of chelates from foods. The extent of d i a l y s i s of z i n c from a p a r t i a l l y or f u l l y digested t e s t meal can demonstrate the presence of c h e l a t e s , suggest the molecular s i z e of chelate complexes and i s p r e d i c t i v e of b i o a v a i l a b i l i t y of minerals. The method of M i l l e r et a l . (18) was adapted to determine the % d i a l y z a b i l i t y ( i n v i t r o b i o a v a i l a b i l i t y ) of z i n c from s e v e r a l soy products (Table V ) . Many of the products with the highest d i a l y z a b i l i t y are the l e a s t r e f i n e d . Some of the concentrates, as w e l l as the t o f u products, have very low d i a l y z a b i l i t y . The r e s u l t s of these i n v i t r o t e s t s do not n e c e s s a r i l y agree with the r e s u l t s of the in v i v o r a t bioassays. I t i s noteworthy that the phytate-to-zinc molar r a t i o does not p r e d i c t _in v i t r o d i a l y z a b i l i t y by t h i s procedure. The calcium plus magnesium-to-zinc molar r a t i o may be more p r e d i c t i v e of in v i t r o dialyzability. It can be seen i n Table V that a l l soy products with calcium-to-zinc molar r a t i o s of greater than 100 f a l l below 10% d i a l y z a b i l i t y . Of s p e c i a l note i s the magnesium t o f u product. I t has an extremely low calcium-to-zinc molar r a t i o (38) but the highest (217) magnesium to z i n c r a t i o and a low i n v i t r o a v a i l a b i l i t y . As w i l l be seen i n Table VI, magnesium aggravates l o s s of zinc d i a l y z a b i l i t y almost to the same extent as does calcium.

Effect

Table VI of Calcium and Magnesium on In V i t r o B i o a v a i l a b i l i t y of Zinc from Defatted Soy F l o u r a

Dialyzability Calcium Added Ca

%

0 17

10 12

25 7.5

%

a

b

0 17

10 14.5

25 10.5

(%)

(rag) added as CaC03 50 5

Magnesium (mg) Added Mg

of Soy Zinc

50 8.5

100 5

200 5

added as MgC0 100 7

Method: M i l l e r et a l . (18) . Unpublished Added to 12.5 gm of d e f a t t e d f l o u r .

b

200 5.5

300 4.5

600 5

b 3

400 5.5

data of Kondo et a l .

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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12.

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Zinc Bioavailability from Soybean Products 181

Although, the modified M i l l e r procedure d i d not rank soybean products s i m i l a r l y to r a t bioassay, when the procedure was used t o t e s t for the e f f e c t of adding calcium to soy f l o u r , there was a c l e a r demonstration of the i n h i b i t o r y e f f e c t of calcium a d d i t i o n on d i a l y z a b i l i t y of z i n c from soy f l o u r (Table V I ) . Magnesium ion a l s o reduced i n v i t r o b i o a v a i l a b i l i t y of z i n c . The e f f e c t of magnesium was noted p r e v i o u s l y i n v i t r o (19), but has not been adequately tested i n v i v o . Iron added as ferrous i o n at s i m i l a r l e v e l s d i d not a f f e c t z i n c d i a l y z a b i l i t y (data not shown). In v i t r o t e s t s u s i n g other procedures (19) were c a r r i e d out i n our laboratory (Ketelsen et a l . , manuscript i n prep.) to study the d i a l y z a b i l i t y of z i n c from s l u r r i e s of defatted soy f l o u r s over a range of pH's. Either ^ z i n c added to soy s l u r r i e s as 6^zinc carbonate ( e x t r i n s i c l a b e l ) or the ^ z i n c i n t r i n s i c a l l y l a b e l e d i n soybeans grown h y d r o p o n i c a l l y . Results from these studies (data not shown) i n d i c a t e d that the e x t r i n s i c and i n t r i n s i c z i n c pools do not completely mix. The r e s u l t s showed that the pH of the soy suspensions a f f e c t e d the quantity of z i n c that was bound i n complexes of molecular weight i n excess of 12,000. A l s o , i n t r i n s i c z i n c was more l i k e l y to be bound to h i g h molecular weight complexes than e x t r i n s i c z i n c , e s p e c i a l l y for s l u r r i e s at pH 3. w a s

w a s

Table V I I Factors A f f e c t i n g the B i o a v a i l a b i l i t y of Zinc from Soy Products

N u t r i t i o n a l status of man (animal) Dietary phytic acid l e v e l D i e t a r y calcium l e v e l Source of z i n c ( i n t r i n s i c / e x t r i n s i c ) U n i t food processing operations -pH adjustment - l e v e l of refinement - a d d i t i o n or removal of inhibitors/enhancers -other f a c t o r s D i g e s t a b i l i t y of product D i e t a r y l e v e l of magnesium, i r o n , e t c . Stage of m a t u r i t y of bean Others

Our work w i t h soy products i n d i c a t e s that e x t r i n s i c z i n c i s more a v a i l a b l e than i n t r i n s i c z i n c . The i n t r i n s i c and e x t r i n s i c z i n c pools do not completely mix i n a l l soy products. P h y t i c a c i d i s an i n h i b i t o r of z i n c b i o a v a i l a b i l i t y and t h i s i n h i b i t i o n i s aggravated by higher l e v e l s of d i e t a r y calcium and perhaps magnesium. N e u t r a l i z a t i o n of soy i s o l a t e s and concentates, w i t h subsequent d r y i n g , reduces z i n c u t i l i z a t i o n f o r r a t s .

In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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182

NUTRITIONAL

BIOAVAILABILITY

OF

ZINC

Conclusions The p r e d i c t i o n o f z i n c b i o a v a i l a b i l i t y from complex food systems i s not a simple matter. Animal bioassays and i n v i t r o t e s t s help us t o i d e n t i f y f a c t o r s that may enhance o r i n h i b i t z i n c u t i l i z a t i o n from the d i e t . With simple model food systems, we can demonstrate the negative e f f e c t s o f p h y t i c a c i d , calcium and other f a c t o r s on z i n c b i o a v a i l a b i l i t y . However, the i n t e r a c t i o n o f these f a c t o r s i n complex food systems, and t h e i r e f f e c t on z i n c s t a t u s f o r man i s not w e l l understood at t h i s time. Simple p r e d i c t o r s , such as the phytate t o z i n c molar r a t i o , w i l l not be accurate f o r human d i e t s . The aggravating e f f e c t o f calcium and perhaps o f magnesium seen i n r a t and _in v i v o t e s t s may or may not be o f p r a c t i c a l s i g n i f i c a n c e t o man. We can, however, develop a l i s t o f f a c t o r s that have been shown t o a f f e c t z i n c b i o a v a i l a b i l i t y from foods. These are l i s t e d i n Table V I I . Our work w i t h soy products i n d i c a t e s that e x t r i n s i c z i n c i s more a v a i l a b l e than i n t r i n s i c z i n c . The i n t r i n s i c and e x t r i n s i c z i n c pools do not completely mix i n a l l soy products. P h y t i c a c i d i s an i n h i b i t o r o f z i n c b i o a v a i l a b i l i t y and t h i s i n h i b i t i o n i s aggravated by higher l e v e l s o f d i e t a r y calcium and perhaps magnesium. N e u t r a l i z a t i o n o f soy i s o l a t e s and concentrates, w i t h subsequent d r y i n g , reduces z i n c u t i l i z a t i o n f o r r a t s . Acknowledgment s Funding f o r t h i s work was provided, i n p a r t , by the American Soybean Assn. (ASARP #81602) and the USDA (CSRS Grant #616-15-172 under PL89-106).

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In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

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ERDMAN

ET

AL.

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Bioavailability

from Soybean Products

183

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In Nutritional Bioavailability of Zinc; Inglett, George E.; ACS Symposium Series; American Chemical Society: Washington, DC, 1983.