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The Effect of Raw Soybeans on Blood Plasma Carotene and Vitamin A and Liver Vitamin A of Calves1
T H E E F F E C T O F R A W S O Y B E A N S ON B L O O D P L A S M A C A R O T E N E AND VITAMIN A AND LIVER VITAMIN A OF CALVES 1 J. C. SHAW Department of Dairy Husbandry, Maryland Agricultural Experiment Station, College Parb AND
L. A. MOORE AND J. F. SYKES Division of Nutrition and Physiology, Bureau of Dairy Industry, U.S.D.A., Washington, D. C. I n previous reports (7, 8) it was noted that cows receiving during the prep a r t a l period a concentrate ration containing 40 per cent raw soybeans developed signs a n d / o r s y m p t o m s of a v i t a m i n A deficiency of g r e a t e r severity t h a n was expected on the basis of the carotene intake. Since Hilton et al. (1) had shown t h a t the feeding of soybean h a y to cows depressed the v i t a m i n A of the butter, it was suspected t h a t soybeans might also exert an influence on the v i t a m i n A requirements of cattle. The data presented herein are the results of an experiment designed to determine the possible effect of the feeding of soybeans upon the blood plasma carotene and v i t a m i n A a n d liver v i t a m i n A of calves on a known carotene intake. A brief resum6 of the plasma and liver vitamin A data, but not the c a r o t e n e data has been published (8). EXPERIMENTAL F o u r calves, two Ayrshires and two Holsteins, a p p r o x i m a t e l y 14 wk. of a g e were placed on a v i t a m i n A deficient ration. They were considered to be depleted when the blood v i t a m i n A had decreased to less t h a n 5 "t p e r 100 ml. of blood plasma. This level was reached a f t e r they were on the v i t a m i n A deficient ration for 35 days. Each calf then was given slightly more t h a n the m i n i m u m requirement (4) or 32 7 of carotene per p o u n d of body weight per day in the f o r m of alfalfa leaf meal for the r e m a i n d e r of the experiment. A t the end of 49 days, the calves were divided into two groups with a Holstein and A y r s h i r e calf in each group. Group I was continued on the basal ration and group I I was changed to the test ration which contained a p p r o x i m a t e l y 30 per cent ground r a w soybeans. This period of 170 days will be r e f e r r e d to as the test period. The rations are shown below: Ingredient Barley Oats Wheat Bran Linseed meal Soybeans Salt
Basal ration
Soybean ration
(lb.) 240 180 180 60
(lb.) 100 180 180
8
200 8
Received for publication August 19, 1950. 1 Paper no. A294, eontrlbution no. 2244 of the Maryland Agricultural Experiment Station. 176
177
T H E E F F E C T OF R A W SOYBEANS AS FEED
Seventy-five grams of D-activated plant sterol were added to each ration. I n addition, each calf received 1 lb. of skimmilk daily. Clean shavings were used for bedding and were kept in the feed boxes at all times as the source of roughage. No attempt was made in these studies to balance the fat content of the two rations. The calves were fed according to Morrison's standards for growing dairy cattle (6). The calves were weighed at weekly intervals and the feeding was adjusted on the following day. The alfalfa leaf meal was analyzed for carotene at weekly intervals. Blood samples for the determination of blood plasma carotene and vitamin A were drawn at 7 a.m. once each week before the feeding of alfalfa leaf meal. Spinal fluid pressure measurements were made 33, 48 and 104 days after soybeans were added to the ration. a tt I t I I t -I A I
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v-/Jg CAROTENE ADDED ISOYBE~]S ADDED 0
20
40
60
80
I00
....
120
CONTROL GROUP GROUP
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•S O Y B E A N
140
160
180
200
220
the plasma vitamin A of
calves.
DAYS
FIG. 1. The effect of the feeding of
soybeans upon
A t the end of the test period the calves were slaughtered and the livers were mascerated and analyzed for vitamin A, using an unpublished method of H. G. ~ i s e m a n of the B u r e a u of D a i r y Industry. A modification of the procedure of Moore (3) and of Kimble (2) was used for plasma vitamin A and carotene. The carotene content of the alfalfa leaf meal was determined by the method of Moore and E l y (5). The spinal fluid pressure technique was that of Sykes and Moore (10). RESULTS
The plasma vitamin A and carotene are shown graphically in figures 1 and 2 f o r the depletion period, for the 2-wk. period of carotene addition without soy-
178
J.c.
SHAW ET AL.
b e a n s a n d f o r t h e r e m a i n d e r of t h e t e s t p e r i o d d u r i n g w h i c h two of t h e calves r e c e i v e d s o y b e a n s i n t h e r a t i o n . I t w i l l be o b s e r v e d t h a t b o t h t h e p l a s m a carot e n e a n d v i t a m i n A v a l u e s were l o w e r in t h e t w o calves t h a t r e c e i v e d s o y b e a n s ; the v i t a m i n A v a l u e s were affected m u c h m o r e t h a n t h e p l a s m a c a r o t e n e . T h e 56
DEPLETION I
I CAROTENE ADDED
50YSEANS ADDED
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SOYBEAN GROUP
¥ 0
20
40
60
80
/00
/20
/40
/60
180
200
220
DAYS
Fie. 2. The effect of the feeding of soybeans upon the plasma carotene of calves. r e a s o n f o r t h e m a r k e d d e c l i n e i n b l o o d p l a s m a c a r o t e n e a n d v i t a m i n A a t 220 d a y s w a s n o t a p p a r e n t . A n e w l o t of a l f a l f a l e a f m e a l was u s e d a t t h i s t i m e a n d i t is p o s s i b l e t h a t some e r r o r was m a d e i n t h e a n a l y s i s of t h e m e a l o r i n feeding. TABLE 1 The effect of the feeding of soybeans upon the plasma carotene and vitamin A of calves N o . of
Plasma
Plasma
Ration
Calf
analyses
carotene
vitamin A
Control Control Soybean Soybean
133A 668H 132A 670H
25 25 25 25
(,~/i00 ml.) 25.68 +_8.33 22.40 _+6.83 18.90 + 4.93 17.18 + 5.22
(-~/100 ml.) 9.60 _+1.26 9.13 +_1.58 5.64 + 1.68 6.46 _+1.54
The average plasma carotene and vitamin A values during the test period are s h o w n i n t a b l e 1. T h e l o w e r p l a s m a c a r o t e n e a n d v i t a m i n A v a l u e s o f t h e soyb e a n f e d c a l v e s w e r e f o u n d to be h i g h l y s i g n i f i c a n t w h e n c o m p a r e d to t h e c o n t r o l calves. T h e F v a l u e s f o r b e t w e e n g r o u p s f o r p l a s m a c a r o t e n e a n d v i t a m i n A
179
T H E EFFECT OF RAW SOYBEANS AS FEED
were 7.875 and 48.219, respectively, whereas, Snedecor's (9) value at the 1 per cent level is 6.90. The greater effect upon plasma vitamin A is reflected in a much larger F value for between groups for plasma vitamin A than for carotene. There was no significant difference between calves within groups for either plasma carotene or vitamin A. The vitamin A content of the livers of the four calves at the end of the test period is shown in table 2. The difference is even more striking than in the case of plasma vitamin A, the two calves on the soybean ration having 0.72 and 2.75 7 of vitamin A per gram of liver and the two controls 9.07 and 9.22 7 per gram. TABLE 2
The effect of the feeding of soybeans upon the liver vitamin A of calves Ration
Calf
Control Control Soybean Soybean
133A 668H 132A 670H
Weight of liver
Vitamin A in liver (wet basis)
(kg.)
(~/g.)
3.0 3.5 3.7 3.5
9.07 9.22 0.72 2.75
Spinal fluid pressure measurements were made during the test period. These are recorded in table 3. It will be noted that all of the values are within the normal range as established by Moore et al. (4). TABLE 3
Spinal fluid pressure of experimental calves Spinal fluid pressure during test period Ration
Control Control Soybean Soybean
Calf
133A 668H 132A 670H
33rd d.
48th d.
104th d.
(ram. tt~0)
(ram. HtO)
(ram. H,O)
80 75 75 80
95 75 80 110
85 100 80 100
DISCUSSION
The marked depression in the plasma and liver vitamin A produced by the feeding of ground raw soybeans to calves on an adequate but low intake of carotene suggests that difficulties may be encountered in feeding soybeans to cattle that are on a relatively low carotene ration. Such conditions could be encountered when rather large amounts of soybeans are added to the ration of cattle receiving a rather poor quality timothy hay as the main source of carotene. Squibb et al. (10) fed soybeans to cows and reported a depression in plasma carotene but only a slight depression in plasma vitamin A which was quite different from our earlier report (8) and from the data in this paper. However, their cows received relatively large and varying amounts of carotene and no attempt was made to determine the actual carotene intake.
180
J.C.
SHAW ET hL.
No increase was observed in the spinal fluid pressure values. However, the low liver vitamin A values indicate considerable depletion. It appears that the experiment may not have been continued for a sufficient length of time or that the calves may have been a little too old at the beginning of the experiment for the effect of the vitamin A depletion to be refleced in an increase in spinal fluid pressure. CONCLUSIONS
The addition of 30 per cent ground raw soybeans to the ration of two calves when compared with two control calves decreased both the plasma and liver vitamin A markedly. Plasma carotene also was decreased but not to as great an extent as plasma vitamin A. The spinal fluid pressure measurements were all within the normal range. REFERENCES (1) HILTON, J . H., HAUGE, S. M., AND WILBUR, J'. W. The Vitamin A Activity of Butter Produced by Cows Fed A l f a l f a Hay and Soybean Hay Cut in Different Stages of Maturity. J. Dairy Sci., 18: 795-800. 1935. (2) KIMBLE, M. S. The Photoelectric Determination of Vitamin A and Carotene in H u m a n Plasma. J. Lab. Clin. M e d , 24: 1055-1065. 1939. (3) MOORE, L . A . Determination of Blood Plasma Car~)tene in the Bovine Using a Photoelectric Colorimeter. J. Dairy Sci., 22: 501-511. 1939. (4) MOORE, L. A., BERRY, M. H., A~rD SYKES, J . F . Carotene Requirements for the Mainteance of a Normal Spinal Fluid Pressure in Dairy Calves. J . Nutrition, 26: 649-658. 1943. (5) MOORE, L. A., AND ELY, R. Extraction of Carotene from P l a n t Material. A Rapid Quantitative Method. Ind. Eng. Chem., Anal. Ed., 13: 600-601. 1941. (6) MOR~tISON,F . B . Feeds and Feeding. The Morrison Publishing Co. Ithaca, N . Y . 1937. (7) SHAW, J. C. Studies on Ketosis in Dairy Cattle. X. The Effect of a Vits.m~n A Deftcieney. 5. Dairy Sci, ~3: 486-495. 1950. (8) SHAW, J. C., MOORE, L. A., AND SYKES, J. Vitamin A Deficiency in Dairy Cattle on Rations Containing Ground Raw Soybeans. J. Dairy Sci., 30: 567. 1947. (9) SNEDECOR,S . W . Statist']~ca~ Methods, 4th ed. Iowa State College Press, Ames. 1946. (10) SQUIBB, R. L , CANNON, C. ¥ , AND ALLEN, R. S. Effect of Raw Soybeans and of Soybean Oil on Plasma Carotene and on Vitamin A as Measured by Activated Glycerol Dichlorohydrin. J. Dairy Sci., 31: 421-427. 1948. (11) SYKES, g. F., AND MOORE, L . A . The Normal Cerebrospinal Fluid Pressure and a Method for its Determination in Cattle. Am. J. Vet. Research 3: 364-367. 1942.
L. A. MOORE AND J. F. SYKES Division of Nutrition and Physiology, Bureau of Dairy Industry, U.S.D.A., Washington, D. C. I n previous reports (7, 8) it was noted that cows receiving during the prep a r t a l period a concentrate ration containing 40 per cent raw soybeans developed signs a n d / o r s y m p t o m s of a v i t a m i n A deficiency of g r e a t e r severity t h a n was expected on the basis of the carotene intake. Since Hilton et al. (1) had shown t h a t the feeding of soybean h a y to cows depressed the v i t a m i n A of the butter, it was suspected t h a t soybeans might also exert an influence on the v i t a m i n A requirements of cattle. The data presented herein are the results of an experiment designed to determine the possible effect of the feeding of soybeans upon the blood plasma carotene and v i t a m i n A a n d liver v i t a m i n A of calves on a known carotene intake. A brief resum6 of the plasma and liver vitamin A data, but not the c a r o t e n e data has been published (8). EXPERIMENTAL F o u r calves, two Ayrshires and two Holsteins, a p p r o x i m a t e l y 14 wk. of a g e were placed on a v i t a m i n A deficient ration. They were considered to be depleted when the blood v i t a m i n A had decreased to less t h a n 5 "t p e r 100 ml. of blood plasma. This level was reached a f t e r they were on the v i t a m i n A deficient ration for 35 days. Each calf then was given slightly more t h a n the m i n i m u m requirement (4) or 32 7 of carotene per p o u n d of body weight per day in the f o r m of alfalfa leaf meal for the r e m a i n d e r of the experiment. A t the end of 49 days, the calves were divided into two groups with a Holstein and A y r s h i r e calf in each group. Group I was continued on the basal ration and group I I was changed to the test ration which contained a p p r o x i m a t e l y 30 per cent ground r a w soybeans. This period of 170 days will be r e f e r r e d to as the test period. The rations are shown below: Ingredient Barley Oats Wheat Bran Linseed meal Soybeans Salt
Basal ration
Soybean ration
(lb.) 240 180 180 60
(lb.) 100 180 180
8
200 8
Received for publication August 19, 1950. 1 Paper no. A294, eontrlbution no. 2244 of the Maryland Agricultural Experiment Station. 176
177
T H E E F F E C T OF R A W SOYBEANS AS FEED
Seventy-five grams of D-activated plant sterol were added to each ration. I n addition, each calf received 1 lb. of skimmilk daily. Clean shavings were used for bedding and were kept in the feed boxes at all times as the source of roughage. No attempt was made in these studies to balance the fat content of the two rations. The calves were fed according to Morrison's standards for growing dairy cattle (6). The calves were weighed at weekly intervals and the feeding was adjusted on the following day. The alfalfa leaf meal was analyzed for carotene at weekly intervals. Blood samples for the determination of blood plasma carotene and vitamin A were drawn at 7 a.m. once each week before the feeding of alfalfa leaf meal. Spinal fluid pressure measurements were made 33, 48 and 104 days after soybeans were added to the ration. a tt I t I I t -I A I
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v-/Jg CAROTENE ADDED ISOYBE~]S ADDED 0
20
40
60
80
I00
....
120
CONTROL GROUP GROUP
I
•S O Y B E A N
140
160
180
200
220
the plasma vitamin A of
calves.
DAYS
FIG. 1. The effect of the feeding of
soybeans upon
A t the end of the test period the calves were slaughtered and the livers were mascerated and analyzed for vitamin A, using an unpublished method of H. G. ~ i s e m a n of the B u r e a u of D a i r y Industry. A modification of the procedure of Moore (3) and of Kimble (2) was used for plasma vitamin A and carotene. The carotene content of the alfalfa leaf meal was determined by the method of Moore and E l y (5). The spinal fluid pressure technique was that of Sykes and Moore (10). RESULTS
The plasma vitamin A and carotene are shown graphically in figures 1 and 2 f o r the depletion period, for the 2-wk. period of carotene addition without soy-
178
J.c.
SHAW ET AL.
b e a n s a n d f o r t h e r e m a i n d e r of t h e t e s t p e r i o d d u r i n g w h i c h two of t h e calves r e c e i v e d s o y b e a n s i n t h e r a t i o n . I t w i l l be o b s e r v e d t h a t b o t h t h e p l a s m a carot e n e a n d v i t a m i n A v a l u e s were l o w e r in t h e t w o calves t h a t r e c e i v e d s o y b e a n s ; the v i t a m i n A v a l u e s were affected m u c h m o r e t h a n t h e p l a s m a c a r o t e n e . T h e 56
DEPLETION I
I CAROTENE ADDED
50YSEANS ADDED
$
k
II~l _11 II II t~
..12 ^
~
v
,4t
"..,y \I."
u
24
~\,~
~)
Ii/\t
, '.
\
4 I I i t
I ~^ ,
,;;,~,~
,sr~.,
\l
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\I
\vl.
8
".~
SOYBEAN GROUP
¥ 0
20
40
60
80
/00
/20
/40
/60
180
200
220
DAYS
Fie. 2. The effect of the feeding of soybeans upon the plasma carotene of calves. r e a s o n f o r t h e m a r k e d d e c l i n e i n b l o o d p l a s m a c a r o t e n e a n d v i t a m i n A a t 220 d a y s w a s n o t a p p a r e n t . A n e w l o t of a l f a l f a l e a f m e a l was u s e d a t t h i s t i m e a n d i t is p o s s i b l e t h a t some e r r o r was m a d e i n t h e a n a l y s i s of t h e m e a l o r i n feeding. TABLE 1 The effect of the feeding of soybeans upon the plasma carotene and vitamin A of calves N o . of
Plasma
Plasma
Ration
Calf
analyses
carotene
vitamin A
Control Control Soybean Soybean
133A 668H 132A 670H
25 25 25 25
(,~/i00 ml.) 25.68 +_8.33 22.40 _+6.83 18.90 + 4.93 17.18 + 5.22
(-~/100 ml.) 9.60 _+1.26 9.13 +_1.58 5.64 + 1.68 6.46 _+1.54
The average plasma carotene and vitamin A values during the test period are s h o w n i n t a b l e 1. T h e l o w e r p l a s m a c a r o t e n e a n d v i t a m i n A v a l u e s o f t h e soyb e a n f e d c a l v e s w e r e f o u n d to be h i g h l y s i g n i f i c a n t w h e n c o m p a r e d to t h e c o n t r o l calves. T h e F v a l u e s f o r b e t w e e n g r o u p s f o r p l a s m a c a r o t e n e a n d v i t a m i n A
179
T H E EFFECT OF RAW SOYBEANS AS FEED
were 7.875 and 48.219, respectively, whereas, Snedecor's (9) value at the 1 per cent level is 6.90. The greater effect upon plasma vitamin A is reflected in a much larger F value for between groups for plasma vitamin A than for carotene. There was no significant difference between calves within groups for either plasma carotene or vitamin A. The vitamin A content of the livers of the four calves at the end of the test period is shown in table 2. The difference is even more striking than in the case of plasma vitamin A, the two calves on the soybean ration having 0.72 and 2.75 7 of vitamin A per gram of liver and the two controls 9.07 and 9.22 7 per gram. TABLE 2
The effect of the feeding of soybeans upon the liver vitamin A of calves Ration
Calf
Control Control Soybean Soybean
133A 668H 132A 670H
Weight of liver
Vitamin A in liver (wet basis)
(kg.)
(~/g.)
3.0 3.5 3.7 3.5
9.07 9.22 0.72 2.75
Spinal fluid pressure measurements were made during the test period. These are recorded in table 3. It will be noted that all of the values are within the normal range as established by Moore et al. (4). TABLE 3
Spinal fluid pressure of experimental calves Spinal fluid pressure during test period Ration
Control Control Soybean Soybean
Calf
133A 668H 132A 670H
33rd d.
48th d.
104th d.
(ram. tt~0)
(ram. HtO)
(ram. H,O)
80 75 75 80
95 75 80 110
85 100 80 100
DISCUSSION
The marked depression in the plasma and liver vitamin A produced by the feeding of ground raw soybeans to calves on an adequate but low intake of carotene suggests that difficulties may be encountered in feeding soybeans to cattle that are on a relatively low carotene ration. Such conditions could be encountered when rather large amounts of soybeans are added to the ration of cattle receiving a rather poor quality timothy hay as the main source of carotene. Squibb et al. (10) fed soybeans to cows and reported a depression in plasma carotene but only a slight depression in plasma vitamin A which was quite different from our earlier report (8) and from the data in this paper. However, their cows received relatively large and varying amounts of carotene and no attempt was made to determine the actual carotene intake.
180
J.C.
SHAW ET hL.
No increase was observed in the spinal fluid pressure values. However, the low liver vitamin A values indicate considerable depletion. It appears that the experiment may not have been continued for a sufficient length of time or that the calves may have been a little too old at the beginning of the experiment for the effect of the vitamin A depletion to be refleced in an increase in spinal fluid pressure. CONCLUSIONS
The addition of 30 per cent ground raw soybeans to the ration of two calves when compared with two control calves decreased both the plasma and liver vitamin A markedly. Plasma carotene also was decreased but not to as great an extent as plasma vitamin A. The spinal fluid pressure measurements were all within the normal range. REFERENCES (1) HILTON, J . H., HAUGE, S. M., AND WILBUR, J'. W. The Vitamin A Activity of Butter Produced by Cows Fed A l f a l f a Hay and Soybean Hay Cut in Different Stages of Maturity. J. Dairy Sci., 18: 795-800. 1935. (2) KIMBLE, M. S. The Photoelectric Determination of Vitamin A and Carotene in H u m a n Plasma. J. Lab. Clin. M e d , 24: 1055-1065. 1939. (3) MOORE, L . A . Determination of Blood Plasma Car~)tene in the Bovine Using a Photoelectric Colorimeter. J. Dairy Sci., 22: 501-511. 1939. (4) MOORE, L. A., BERRY, M. H., A~rD SYKES, J . F . Carotene Requirements for the Mainteance of a Normal Spinal Fluid Pressure in Dairy Calves. J . Nutrition, 26: 649-658. 1943. (5) MOORE, L. A., AND ELY, R. Extraction of Carotene from P l a n t Material. A Rapid Quantitative Method. Ind. Eng. Chem., Anal. Ed., 13: 600-601. 1941. (6) MOR~tISON,F . B . Feeds and Feeding. The Morrison Publishing Co. Ithaca, N . Y . 1937. (7) SHAW, J. C. Studies on Ketosis in Dairy Cattle. X. The Effect of a Vits.m~n A Deftcieney. 5. Dairy Sci, ~3: 486-495. 1950. (8) SHAW, J. C., MOORE, L. A., AND SYKES, J. Vitamin A Deficiency in Dairy Cattle on Rations Containing Ground Raw Soybeans. J. Dairy Sci., 30: 567. 1947. (9) SNEDECOR,S . W . Statist']~ca~ Methods, 4th ed. Iowa State College Press, Ames. 1946. (10) SQUIBB, R. L , CANNON, C. ¥ , AND ALLEN, R. S. Effect of Raw Soybeans and of Soybean Oil on Plasma Carotene and on Vitamin A as Measured by Activated Glycerol Dichlorohydrin. J. Dairy Sci., 31: 421-427. 1948. (11) SYKES, g. F., AND MOORE, L . A . The Normal Cerebrospinal Fluid Pressure and a Method for its Determination in Cattle. Am. J. Vet. Research 3: 364-367. 1942.