EFFECT OF TRACE MINERALS ON THE STABILITY OF VITAMIN E IN SWINE GROWER DIETS'y2
Iowa Agriculture and Home Economics Experiment Station, Ames 50011 ABSTRACT
The stability of a-tocopheryl acetate and inherent tocopherols in swine grower diets containing high levels of trace minerals was studied in a 12-wk experiment. Com-soybean meal diets with either 0 or 1% crude soybean oil and without trace minerals added (NOTM) were supplemented with a standard trace mineral mix (TM), TM + 250 ppm of Cu, TM + 1,OOO ppm of Fe, TM + 1,OOO ppm of Zn,or TM + 100 ppm of Mn. Alphatocopheryl acetate decreased linearly (P c .OOOl) during 12 wk of storage in diets containing NOTM, TM, Zn,or Mn at a rate of .015 mad. Addition of Fe (P c .005) or Cu (P e .05) increased the rate of a-tocopheryl acetate loss. The addition of soybean oil had no effect (P > .l) on the rate of a-tocopheryl acetate loss. The a-tocopherol levels of the NOTM diet decreased (P c .01) by approximately 50% during the 12-wk storage period. Addition of TM alone in diets that did not contain soybean oil had no effect (P > .l) on the rate of a-tocopherol loss. The addition of Cu,Fe, Zn,or Mn (P c: .OS) to diets that did not contain soybean oil increased the rate of a-tocopherol loss in comparison with NOTM and TM diets. Addition of Cu decreased a-tocopherol below 10% of initial levels in approximately 10 d. The addition of soybean oil to the diets containing TM, Fe, Zn (P c .005), or Mn (P c .OS) further increased the rate of a-tocopherol loss. Within the diets containing soybean oil, all trace mineral additions increased (P c .01) the rate of loss of atocopherol loss. The loss of natural y-tocopherol was similar to that of a-tocopherol. The results indicate that the rate of oxidation of natural tocopherols is increased in diets containing increased levels of Cu, Fe, Zn, or Mn. Key Woids: Pigs, Tocopherols, Copper, Iron, Zinc, Manganese J. Anim. Sci. 1991. 69:1!394-2000
Introduction
than in low-moisture com due to increased The levels of natural tocopherols present in peroxidation of the lipid. Hakkarainen et al. (1983a,b) reported similar results in highfeed ingredients are influenced by the environmoisture barley; they found that storage of mental conditions associated with processing high-moisture barley under anaerobic condiand storage (Le., grinding, moisture content, tions decreased the rate of tocopherol oxidaunsaturated fat content, temperature, humidity, tion. and exposure to oxygen). Young et al. (1975) Chemical analysis of 89 samples of swine reported that the rate of oxidation of natural diets obtained from swine producers indicated tocopherol was higher in high-moisture corn that many samples contained high levels of Fe, Cu, Zn, and Mn. The means of the analyzed values for Fe, Cu,Zn,and Mn were 6.8, 24.0, 'Journal Paper No. J-13966 of the Iowa Agric. and 3.5, and 24.0 times, respectively, the NRC (1988) requirements for these trace elements Home Ikon. Exp. Sta., Ames. Project No. 2830. 'Supported in part by Hoffmam-LaRoche, Inc., (Ewan, 1985, 1986). Richter et al. (1982) Nutley. NJ and Purina Mills, St. Louis, MO. found that the addition of excess levels of a 'Resent address: Coastal Plain Exp. Sta., Dept. of trace mineral mix to a swine diet increased the Anh. Sci., P. 0. Box 748, Tifton, GA 31793. rate of tocopherol loss. Dove and Ewan (1987) %pt. of ~ n h sci. . reported that Cu (250 ppm), Fe (1,OOO pprn), Received April 6, 1990. Accepted November 19, 1990. or Zn (1,OOO ppm) increased oxidation of the 1994
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C. R. Dove3 and R. C. Ewan4
EFFECT OF MINERALS
1995
ON VITAMIN E STABILlTY
Materials and Methods
Corn-soybean meal diets (NOTM) formulated to Contain 18% CP with 0 or 1% crude soybean oil substituted for corn (Table 1) were studied. Trace minerals were not added to the diet. Vitamins were added to meet NRC (1979, 1988) requirements, except for tocopherol, which W ~ S added at 20 N/kg a~ D M tomphevl acetatd. he diets with or withuut soybean oil were supplemented with a trace mineral mix that contained trace minerals (TM) at levels that were similar to the NRC (1988) requirements for growing pigs (1.5 ppm of I, 30 ppm of Mn, 5 ppm of Cu, 100 ppm of Fe and 100 ppm of%); TM + 250 ppm of Cu added as &SO& TM + 1,OOO ppm of Fe added as FeS047H20, TM + 1,OOO ppm of Zn as 250, and TM + 100 ppm of Mn as MnsO4-7H20. Each of the 12 diets was mixed on three successive days, providing three replicaticms of each diet. Diets were stored in glass jars in the dark at 23 to 26'C for 12 w k On d 0, 3, 6,9, 12, 15, 18, 21. 28, 35,42, 49, 56, 63, 70, 77, and 84 of storage, duplicate 5-g subsamples were taken, ground, and analyzed for tocopherols. Tocopherol analysis was performed as described previously (Dove and Ewan, 1990). Samples (5 g) were extracted with acetone for 3 h in a Soxhld extractor, evaporated to dryness, redissolved in 10 ml of HPLC-grade hexane, and filtered through an .8+m membrane filter. The filtrate was analyzed for tocopherol isomers by the HPLC method of Cort et al. (1983) with fluorometric detection. The mobile phase was 3.5% tetrahydrofuran (vol/vol) in hexane with a flow rate of 2.0 ml/ min. The HPLC procedure provided separation and detection of a-tocopheryl acetate and the eight natural isomers of tocopherol (a,f3, y, and 6 tocopherol and a.p. y and 6 tocotrienol). Concentrations of a-tocopheryl acetate, a-
TABLE 1. COMPOSITION OF DIETS
Basal+ soybcm Oil
Isgrcdicnt
Basal
Y a o w coma
722 24.6 1.1
712 24.6 1.1 1.o
~UUICa?bJMtC
.9
salt Vitaminmixb
.3
9 .3
%
soytrcenmeal,48.5%CP
Dic3lciompbosphate soyatanoil
-
1.o
1.0
"Tracemimralpremixwassabstitntedfor1%ofcom and comiLmted the following per kilogram of diet: 1.15 30 or 100 mg of lmn@msc as mg of iodine as W O 4 H f l , 5 or 250 mg of copper m csllsO4, 100 or 1 . mmg of iron as ~ 0 ~ 7 Hand ~ 100 0 o ,r l.mmg of zincaszno.
bcontn.baabd pa kilogram Of dia.4,400IUOf ViA palmitate. 1,100 Iu of vitamin Da 6.6 mg of riboflavin, 17.6 mg of d-pantothenic acid, 33 mg Of niacin. and 22 of Vitamin Biz. tocopherol, ~-tocopheml, and ytompheml were determined by standard curves of peak area vs concenbration of known standards6. Data were aualyzd by analysis of variance ( G M , SAS, 1982). Lmear (acetate) and semilogarithmic linear (a-towpheml) regressions were calculated to evaluate the effects of time on tocopherol concentrations. Comparisons of regression coefficients were made as de& by Steel and Tonie (1960) to evaluatetheeffectsoftracemineraladditions w i t h and between the basal diet and the basal diet with added soybean oil. Biological halflives (the time required for loss of 5096 of the initial concentration) were calculated from the exponents of the semilogarithmic regressions as described by Rescigno and Segre (1966). Results and Discussion
The analyzed composition of the diets is presented in Table 2. Addition of soybean oil to the basal diet increased (P e .01) the concentrations of natural a-, and ' y ~ tocopherols in the diets. These natural tocophervls increased because of the tocopherols inherent in the added soybean oil. The concentration of a-towpheryl acetate Vable 3) decreased linearly (Pe .OO01) cfuring storage. l b r e was no difference in the rate of loss of u-tmpheryl acetate among the dids containing NOTM, TM, Zn, or Mn with or without added suybean oil. The pooled rate of
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inherent tocopherols during storage, but they did not study the stability of a-tocopheryl acetate. The present experiment was conducted to determine the stability of a-tocopheryl acetate and of inherent tocopherols in swine p w e r diets during prolonged storage m the presence of high levels of Cu, Fe, Zn, or Mn.
1996
DOVE AND EWAN
Basal +
Innredient
Basal
so-
DM, %
85.0 17.5 3,758 2.1 .37
84.7 17.1 3,837 3.1 .36
5.4 9.0 263
5.0 8.7 252
256 328 870
220 313 870
29 124 844
28 110 868
19.4 43.8 111
17.8 37.0 112
e, %
GE, kcavkg Ether extracz % Se. ppm a.ppm NOTI@ TMa 250 ppm added Fe, ppm NOTM
TM 1.ooO pprn added
zn, PPm
NOTM
TM 1,ooOppm added Mn, PPm
NOTM
TM 100 ppm added Tocopherols,mg/lrg Acetate Beta Gamma
oil
14.1 4.43 .36 36.2
14.1 5.23 .49 462
%OTM are the diets that do not contain trace minerals. TM are diets that contain only the trace mimral premix.
TABLE 3. EFPECI: OF TRACE MIMeRALs ON THE RATE OF LOSS OF ALPHA-TOCOPHERYL ACETATE DURING STORAGE' Treatment Item Intercept
SE Slope SE R2 Intercept SE Slope SE R2
NOTM
TM
Cub
14.8 .45 -.Of 1 .010 .a2
13.9 A1 -.012
13.8 .39 -.036
.009 .03
.009
14.1 .42
152 .49 -.018 .011
13.2 .39 ,032
-.m -010 .38
-18
25
.009 .38
Fec
zn
Basal 14.2 14.9 .46 .39 -.024 -.a3 .009 .011 .31 .lo Basal + soybean oil 14.0 14.3 .39 .42 -.w3 -.015 .009 .010 -57 .I4
Mn
Pooled
13.6 .46 -.m3 .010
14.2 .19
.oo 14.1 .38 -.009 .009 -09
-.on .004
.08 14.1 .19 -.ON .004 .09
'Acetate = a + b (days of storage). Diets that did not contain trace minerals are NOTM and diets containing only the trace mineral premix are TM.No significant difference existed among NOTM, TM, Zn, or Mn with or without added soybean oil.For these eight diets, the pooled regression intercept was 14.4 mg@ (SE= .16) and the slope was -.015 mg/ d (SE = .W, R2 = .W, n = 408). The slope was greater than 0 (P .ooOl). bpooled Cu was different from the pooled r ession (P < .05). The regression intercept was 135 mg/kg (SE = .27), and the slope was -.034 mg/d (SE = .W, R - 22; n = 102).
T-
cPo~ledFe was different from the pooled regression (P < .OM).The regression intercept was 14.1 m and the slope was -.043 mgld (SE = .W, R2 = .32;n = 102).
a (SE = 27),
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loss in these diets was .015 mg/d during 84 d of storage. Although this rate of loss was significant statistically, in diets containing 20 lU of a-tocopheryl acetate, 93% of the acetate would remain after 90 d of storage, indicating that a-tocopheryl acetate is relatively stable in mixed diets. Klaiii (1973) reported similar rates of loss of a-tocopheryl acetate during 6 mo of storage in complete poultry diets in meal form, but less was lost from pelleted diets. Klatii (1973) also reported that atocopheryl acetate was stable in concentrated premixes. High levels of Cu (250 ppm; P e .E)or Fe (1,O00 ppm; P e .005)increased the rate of loss to .OM and .043 mdd, respectively. There was no difference (P > .l) in rate of a-tocopheryl acetate loss between diets with or without added soybean oil. The results of our experiment suggest that 250 pp of Cu or 1,OOO ppm of Fe decreased the stability of atocopheryl acetate. The natural a-tocopheryl concentration (Table 4) in the NOTM and 'I'M diets not containing soybean oil decreased (P e .01) during the 846 storage period. A single component exponential decrease described the rate of loss of a-tocopherol and indicated a half-life of 84 and 91 d for the NOTM and TM
TABLE 2. ANALYZED COMPOSmON OF DIETS
1997
EFFECT OF MINERALS ON VITAMIN E STABILITY
TABLE 4. EFFECT OF TRACE MINERALS ON ALPHA-TOCOpHEROLa LOSS DURING STORAGE Trratment
Cu
Thf
zn
Fe
Mn
Basal 4.43 .03
Intercept
SE
3.77
SE
.ooo9
.o007
59 91
.70
R2 Half-life, d
84
Intercept Slope
5.23 .03 -.00918
455 .04 -.0134h
SE
.ooo8
.ooo9
SE
.72
R2 Half-life. d
.82
76 ~
= log
52 ~~
2.79 .08
-.036Sd -.om* .0127 .0018 .14 .63 19 42 Basal + soybean oilf .46 2.92 57 .13 -.O3Ni -.0543i .W29 .0130 26 .72 13 21
-.me
-.082c
Slope
.34 .55
.04
423 .04 -.0121& .Oo08 .82 57
4.34 .04 -.0116’ .0010 .74 60
5.09
5.04
.05
.05
-.02Ooi .m11 .87
-.O15Sh .m12 .76
44
34
~~~~~
~~~
+b
(days of storage). Tocopherol = ae*. bDiets that did not contain kace minerals are NOTM and diets containing only the trace m i n d premix are TM. c*4eSlopeswithin the basal diet that do not share the same superscript are different (P < -05). ‘DBerenCe b e e n basal and basal + soybean oil diets within TM, Fe, Zn (P < .005), or Mn (P < .05). gJsiJSlopes within the basal + soybean oil diet that do not share the same superscript are different (P c .01). %g tocopherol
a
of Cu, Fe, Zn, or Mn to the diets without added soybean oil reduced the half-life of atocopherol to 19,42,57, and 60 d, respectively. Young et al. (1975) found that the atocopherol in mixed diets containing &cidy dried corn was stable during storage.
diets, respectively. This loss of natural tocopherols in the NOTM diet during storage suggests that the natural tocopherols have a limited stability. The rate of loss of atocopherol was increased by the addition of either Cu,Fe, Zn, or Mn (P < .05) to the TM diet without added soybean oil. The addition
TABLE 5. EPFECT OF Cu OR Fe ON ALPHA-TOCOPHEROLa LOSS DURING STORAGE Fe
c11 Basal+ Item
Basal
soybean oil
Basal+
Basal
soybean oil
Fast
Intercept
SE Slope
SE R2 Half-life, d
5.41 .52 -26Sb
.070 .83 2.6
6.13 .47 -.226b .064
2.29 .34 -.11@ .027 .75 6.0
.so
3.1
5.76 .34 -. 165d .027 .85 42
Slow
Intercept SE Slope
SE R2 Half-life, d
20 .66 -.W1 .0143 .03 76
.33 .6 1 -.0263e .0136 .24 26
1.84
.IS -.0088 .m7 53 79
1.15 .15 -.0146 .0048 .50 47
‘Log tocopherol = wg a + b (days of storage)] + w g a + b (days of storage)]. Tocopherol = ae-’ b*c*d~~opes within the fast component that do not share the same supacsipt are different (P < .01). %fference between Cu and Fe within the basal + soybean oil diet (P < .01).
+
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NOTMb
Item
1998
DOVE AND EWAN
D-ALPHA
TOCOPHEROL
--- Exponential - Exponential - slow - rapid
0 Exponential
- sum
I
OI Y
\
F ,001 1 0
10
30
20
40
50
70
60
80
90
Days SBO
+
--.Exponential
- slow
- Exponential - rapid
o Exponential
-
sum
0 Y
\
4
0
E
--
I
4 4
4
.01 * 4
.001
0
10
20
30
40
50
60
70
80
90
0ays Pigme 1. Effect of Cu (250 ppm) on the rate of loss of natural tocopherol. Observed values, slow rate of loss, -----, predicted rate of loss, 0.
*; rapid rate of loss, -;
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The diets used by Young et al. (1975) the addition of trace mineral mixes to swine contained ethoxyquin, whereas the diets used diets accelerated the decrease in the natural in our experiment did not. This difference in tocopherol levels during storage. In the experithe storage stability of the natural tocopherols ment reported here, the rate of loss of amay be a result of the increased demand for a tocopherol was not affected by the addition of dietary antioxidant in the diets used in this trace minerals in diets that did not contain experiment. Richter et al. (1982) reported that added soybean oil.
EEFECT OF MINERALS ON VITAMIN E STABILsry
associated with added dietary fat. No attempt was made in this study to determine the products associated with the loss of tocopherols. Assuming that loss was the result of oxidation, the major products would be a-tocopheryl quinone and a-tocopheroxide (Diplock, 1985); these compounds do not have biological activity (Kasparek, 1980). In the analytical system used in this study, atocopheryl quinone would not be detected. The increase in the rate of loss promoted by trace minerals in this study is consistent with the oxidation of tocopherols. Gamma-tocopherol and a-tocopherol levels (data not shown) were affected similarly by all mineral additions and by addition of soybean Oil. Implications
Alpha-tocopheryl acetate was relatively stable during storage, but the stability of natural tocopherol was limited in diets that do not contain supplemental trace minerals. The stability of a-tocopheryl acetate was decreased in the presence of high levels of Fe or Cu, whereas the rate of loss of natural tocopherols was increased by high levels of all minerals tested (Cu > Fe > Zn > Mn). Corn-soybean meal diets containing growth-promoting levels of Cu or high levels of trace minerals may require the addition of a-tocopheryl acetate to maintain recommended levels of vitamin E in diets during storage, especially in the presence of added unsaturated fat. Literature Cited
Colt, W. M.. T. S. Vicente, E. H. Waysek and B. D. Williams. 1983. Vitamin E content of feedstuffs determined by high-perfomunce liquid chromotographic fluorescence. J. Agric. Food Chem 31:1330. Diplock, A. T. 1985. Vitamia E. In:A. T. Diplock (Ed.) Fatsoluble Vitamins. pp 154-224. Heineman, London. Dove,C.RandR.C.Ewan. 1987.ThecffcctofCyFeorZn addition on the vitamin E status of growing pigs and the natural tocopherol levels in feed.J. Anim. Sci. 65 (Suppl. 1):134 (Abstr.). Dove, C. R and R. C. Ewaa 1990. The & s t of excess dietary copper, iron or zinc on the tocopherol and selenium status of pwingpigs. J. Anim. Sci. 68:2407. Ewaq R. C. 1985. Analysis of feeds and feed ingredients 1984. ISU Research Reports - 1985. Iowa State Univ., AmeS. Ewan, R.C. 1986. Analysis of feeds and feed ingredients 1985. ISU Swine Resrach Reports - 1986. Iowa State univ., Ames. Hawrarainen,R. V.,I. T.Tytippbea and S. G. Bengtsson. 1983a. changes in the content and composition of vitamin E in damp barley stored in airtight bios. J. Sci.
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The addition of soybean oil did not affect the rate of loss of a-tocopherol in the NOTM diet (P> .1). However, in the presence of TM, Fe, Zn (P < .005), or Mn (P < .05), the addition of soybean oil increased the rate of loss of tocopherol. Within the diets with added soybean oil, Cu and Fe additions increased the rate of loss (P< .01) more than the rate of loss in diets with added Zn.The rate of loss in diet with added soybean oil and Zn was greater (P < .01) than the loss in diets containing soybean oil and TM or Mn. The half-life of atocopherol was reduced to 52,21,34, and 44 d by the presence of TM, Fe, Zn, or Mn, respectively, in diets with added soybean oil. The rate of a-tocopherol loss was increased greatly in diets with 250 ppm of added Cu and in the diet containing 1,OOO ppm of Fe. The rate of loss was a two-component curve (Figure 1) characterized by a rapid rate of loss initially and a second component with a rate of loss similar to that in the NOTM or TM diets. The fast component as characterized by halflives ranging from 2.6 to 6 d in the diets containing high levels of Cu or Fe (Table 5). The addition of soybean oil increased the fast component in the presence of added Fe (P e .01) but did not affect the fast component in the presence of added Cu. Because the rates of loss in the presence of Cu were not affected by the addition of soybean oil and because the rates were very rapid, Cu may have a direct effect on tocopherol loss that is not dependent on the production of lipid peroxides. The halflife of a-tocopherol in diets containing 250 ppm of Cu predicts a reduction to less than 10% of the initial concentration after about 10 d of storage. Growth-promoting levels of Cu (125 to 250 ppm) are used widely in the swine industry (Ewan, 1986), and Cu had the greatest destructive effect on the natural tocopherols. Ionized Cu and Fe often are included in lists of factors that increase the rate of loss of natural tocopherols (ullrey, 1981). Waddell and Steenbock (1931) associated the presence of dietary Fe with the loss of natural tocopherols in rat diets. The increased rate of a-tocopherol loss suggests that the presence of trace minerals in diets increases the peroxidation of fats in the diet, increasing the need for dietary antioxidants. The failure of added soybean oil to increase the rate of tocopherol loss in the NOTM diet suggests that trace minerals are hportant in the mechanism of tocopherol loss
1999
2000
DOVE AND EWAN Blaisdale Publishing Co., Waltbam. MA. Richter, V. G., P. Mockel, F. Schone and G. Hachowsky. 1982. Effect of storage time and storage temperature on vitamin A and E levels in vitamin concentrates, premixes of active ingredients, mixedfodder,and liver homogenates. Monats. vetuinaermtd. 3 7 4 0 5 . SAS. 1982. SAS User's Guide: Stafjstics. SAS Inst, Inc., Gary, NC. Steel, RGD. and J. H. Tome. 1960. Principles and F'rucedurcs of Statistics. McGraw-HillBook Co.. New York Ullrey,D. E. 1981. Vitamin E for swine. J. Anim. Sci. 53: 1039. Waddell,J.audH. Steenbock. 193LVitamhEinFelreated dry rations. J. Nutr. 479. Young, L. G.. A. Lun. J. Pos, R. P. Porshaw and D. Edmeades. 1975.VitaminEstabilityincornandmixed feed. J. Anim. Sci. 40:495.
Downloaded from https://academic.oup.com/jas/article-abstract/69/5/1994/4705365 by Iowa State University user on 11 January 2019
Food Agric. 34:1029. Hakkamhen, R.V., J. T. Ty6p-n and S. G. Bengtsson. 1983b. Relative and quantitative changes in total vitamin E and isomer content of barley during cowentional and airtight storage with special refercnce to dvariations. Acta A@. Scaud. 39395. Kasparelr, S. 1980. chemistry of tocopherols and tocotrienols. In: L. J. Uachlin @Vitamin I.) E. A Comprehensive 'Reatise. pp 7-65. Marcel M e r , New Yo&. Klaiii, H. 1973. 'Ihc stability and distribution m feeds of powders colllaining vitamin E. Acta Agric. Scand. (SuppL) 19219. NRC. 1979. Nutrient Rqdrements of Swine (8th Ed.). National Academy Press, Washington, DC. NRC. 1988. Nutrient Requirements of Swine (9th Ed.). National Academy Press, Washington, DC. Rescigno, A. and G. Segre. 1%6. Drug and Tracer Kinetics.
Iowa Agriculture and Home Economics Experiment Station, Ames 50011 ABSTRACT
The stability of a-tocopheryl acetate and inherent tocopherols in swine grower diets containing high levels of trace minerals was studied in a 12-wk experiment. Com-soybean meal diets with either 0 or 1% crude soybean oil and without trace minerals added (NOTM) were supplemented with a standard trace mineral mix (TM), TM + 250 ppm of Cu, TM + 1,OOO ppm of Fe, TM + 1,OOO ppm of Zn,or TM + 100 ppm of Mn. Alphatocopheryl acetate decreased linearly (P c .OOOl) during 12 wk of storage in diets containing NOTM, TM, Zn,or Mn at a rate of .015 mad. Addition of Fe (P c .005) or Cu (P e .05) increased the rate of a-tocopheryl acetate loss. The addition of soybean oil had no effect (P > .l) on the rate of a-tocopheryl acetate loss. The a-tocopherol levels of the NOTM diet decreased (P c .01) by approximately 50% during the 12-wk storage period. Addition of TM alone in diets that did not contain soybean oil had no effect (P > .l) on the rate of a-tocopherol loss. The addition of Cu,Fe, Zn,or Mn (P c: .OS) to diets that did not contain soybean oil increased the rate of a-tocopherol loss in comparison with NOTM and TM diets. Addition of Cu decreased a-tocopherol below 10% of initial levels in approximately 10 d. The addition of soybean oil to the diets containing TM, Fe, Zn (P c .005), or Mn (P c .OS) further increased the rate of a-tocopherol loss. Within the diets containing soybean oil, all trace mineral additions increased (P c .01) the rate of loss of atocopherol loss. The loss of natural y-tocopherol was similar to that of a-tocopherol. The results indicate that the rate of oxidation of natural tocopherols is increased in diets containing increased levels of Cu, Fe, Zn, or Mn. Key Woids: Pigs, Tocopherols, Copper, Iron, Zinc, Manganese J. Anim. Sci. 1991. 69:1!394-2000
Introduction
than in low-moisture com due to increased The levels of natural tocopherols present in peroxidation of the lipid. Hakkarainen et al. (1983a,b) reported similar results in highfeed ingredients are influenced by the environmoisture barley; they found that storage of mental conditions associated with processing high-moisture barley under anaerobic condiand storage (Le., grinding, moisture content, tions decreased the rate of tocopherol oxidaunsaturated fat content, temperature, humidity, tion. and exposure to oxygen). Young et al. (1975) Chemical analysis of 89 samples of swine reported that the rate of oxidation of natural diets obtained from swine producers indicated tocopherol was higher in high-moisture corn that many samples contained high levels of Fe, Cu, Zn, and Mn. The means of the analyzed values for Fe, Cu,Zn,and Mn were 6.8, 24.0, 'Journal Paper No. J-13966 of the Iowa Agric. and 3.5, and 24.0 times, respectively, the NRC (1988) requirements for these trace elements Home Ikon. Exp. Sta., Ames. Project No. 2830. 'Supported in part by Hoffmam-LaRoche, Inc., (Ewan, 1985, 1986). Richter et al. (1982) Nutley. NJ and Purina Mills, St. Louis, MO. found that the addition of excess levels of a 'Resent address: Coastal Plain Exp. Sta., Dept. of trace mineral mix to a swine diet increased the Anh. Sci., P. 0. Box 748, Tifton, GA 31793. rate of tocopherol loss. Dove and Ewan (1987) %pt. of ~ n h sci. . reported that Cu (250 ppm), Fe (1,OOO pprn), Received April 6, 1990. Accepted November 19, 1990. or Zn (1,OOO ppm) increased oxidation of the 1994
Downloaded from https://academic.oup.com/jas/article-abstract/69/5/1994/4705365 by Iowa State University user on 11 January 2019
C. R. Dove3 and R. C. Ewan4
EFFECT OF MINERALS
1995
ON VITAMIN E STABILlTY
Materials and Methods
Corn-soybean meal diets (NOTM) formulated to Contain 18% CP with 0 or 1% crude soybean oil substituted for corn (Table 1) were studied. Trace minerals were not added to the diet. Vitamins were added to meet NRC (1979, 1988) requirements, except for tocopherol, which W ~ S added at 20 N/kg a~ D M tomphevl acetatd. he diets with or withuut soybean oil were supplemented with a trace mineral mix that contained trace minerals (TM) at levels that were similar to the NRC (1988) requirements for growing pigs (1.5 ppm of I, 30 ppm of Mn, 5 ppm of Cu, 100 ppm of Fe and 100 ppm of%); TM + 250 ppm of Cu added as &SO& TM + 1,OOO ppm of Fe added as FeS047H20, TM + 1,OOO ppm of Zn as 250, and TM + 100 ppm of Mn as MnsO4-7H20. Each of the 12 diets was mixed on three successive days, providing three replicaticms of each diet. Diets were stored in glass jars in the dark at 23 to 26'C for 12 w k On d 0, 3, 6,9, 12, 15, 18, 21. 28, 35,42, 49, 56, 63, 70, 77, and 84 of storage, duplicate 5-g subsamples were taken, ground, and analyzed for tocopherols. Tocopherol analysis was performed as described previously (Dove and Ewan, 1990). Samples (5 g) were extracted with acetone for 3 h in a Soxhld extractor, evaporated to dryness, redissolved in 10 ml of HPLC-grade hexane, and filtered through an .8+m membrane filter. The filtrate was analyzed for tocopherol isomers by the HPLC method of Cort et al. (1983) with fluorometric detection. The mobile phase was 3.5% tetrahydrofuran (vol/vol) in hexane with a flow rate of 2.0 ml/ min. The HPLC procedure provided separation and detection of a-tocopheryl acetate and the eight natural isomers of tocopherol (a,f3, y, and 6 tocopherol and a.p. y and 6 tocotrienol). Concentrations of a-tocopheryl acetate, a-
TABLE 1. COMPOSITION OF DIETS
Basal+ soybcm Oil
Isgrcdicnt
Basal
Y a o w coma
722 24.6 1.1
712 24.6 1.1 1.o
~UUICa?bJMtC
.9
salt Vitaminmixb
.3
9 .3
%
soytrcenmeal,48.5%CP
Dic3lciompbosphate soyatanoil
-
1.o
1.0
"Tracemimralpremixwassabstitntedfor1%ofcom and comiLmted the following per kilogram of diet: 1.15 30 or 100 mg of lmn@msc as mg of iodine as W O 4 H f l , 5 or 250 mg of copper m csllsO4, 100 or 1 . mmg of iron as ~ 0 ~ 7 Hand ~ 100 0 o ,r l.mmg of zincaszno.
bcontn.baabd pa kilogram Of dia.4,400IUOf ViA palmitate. 1,100 Iu of vitamin Da 6.6 mg of riboflavin, 17.6 mg of d-pantothenic acid, 33 mg Of niacin. and 22 of Vitamin Biz. tocopherol, ~-tocopheml, and ytompheml were determined by standard curves of peak area vs concenbration of known standards6. Data were aualyzd by analysis of variance ( G M , SAS, 1982). Lmear (acetate) and semilogarithmic linear (a-towpheml) regressions were calculated to evaluate the effects of time on tocopherol concentrations. Comparisons of regression coefficients were made as de& by Steel and Tonie (1960) to evaluatetheeffectsoftracemineraladditions w i t h and between the basal diet and the basal diet with added soybean oil. Biological halflives (the time required for loss of 5096 of the initial concentration) were calculated from the exponents of the semilogarithmic regressions as described by Rescigno and Segre (1966). Results and Discussion
The analyzed composition of the diets is presented in Table 2. Addition of soybean oil to the basal diet increased (P e .01) the concentrations of natural a-, and ' y ~ tocopherols in the diets. These natural tocophervls increased because of the tocopherols inherent in the added soybean oil. The concentration of a-towpheryl acetate Vable 3) decreased linearly (Pe .OO01) cfuring storage. l b r e was no difference in the rate of loss of u-tmpheryl acetate among the dids containing NOTM, TM, Zn, or Mn with or without added suybean oil. The pooled rate of
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inherent tocopherols during storage, but they did not study the stability of a-tocopheryl acetate. The present experiment was conducted to determine the stability of a-tocopheryl acetate and of inherent tocopherols in swine p w e r diets during prolonged storage m the presence of high levels of Cu, Fe, Zn, or Mn.
1996
DOVE AND EWAN
Basal +
Innredient
Basal
so-
DM, %
85.0 17.5 3,758 2.1 .37
84.7 17.1 3,837 3.1 .36
5.4 9.0 263
5.0 8.7 252
256 328 870
220 313 870
29 124 844
28 110 868
19.4 43.8 111
17.8 37.0 112
e, %
GE, kcavkg Ether extracz % Se. ppm a.ppm NOTI@ TMa 250 ppm added Fe, ppm NOTM
TM 1.ooO pprn added
zn, PPm
NOTM
TM 1,ooOppm added Mn, PPm
NOTM
TM 100 ppm added Tocopherols,mg/lrg Acetate Beta Gamma
oil
14.1 4.43 .36 36.2
14.1 5.23 .49 462
%OTM are the diets that do not contain trace minerals. TM are diets that contain only the trace mimral premix.
TABLE 3. EFPECI: OF TRACE MIMeRALs ON THE RATE OF LOSS OF ALPHA-TOCOPHERYL ACETATE DURING STORAGE' Treatment Item Intercept
SE Slope SE R2 Intercept SE Slope SE R2
NOTM
TM
Cub
14.8 .45 -.Of 1 .010 .a2
13.9 A1 -.012
13.8 .39 -.036
.009 .03
.009
14.1 .42
152 .49 -.018 .011
13.2 .39 ,032
-.m -010 .38
-18
25
.009 .38
Fec
zn
Basal 14.2 14.9 .46 .39 -.024 -.a3 .009 .011 .31 .lo Basal + soybean oil 14.0 14.3 .39 .42 -.w3 -.015 .009 .010 -57 .I4
Mn
Pooled
13.6 .46 -.m3 .010
14.2 .19
.oo 14.1 .38 -.009 .009 -09
-.on .004
.08 14.1 .19 -.ON .004 .09
'Acetate = a + b (days of storage). Diets that did not contain trace minerals are NOTM and diets containing only the trace mineral premix are TM.No significant difference existed among NOTM, TM, Zn, or Mn with or without added soybean oil.For these eight diets, the pooled regression intercept was 14.4 mg@ (SE= .16) and the slope was -.015 mg/ d (SE = .W, R2 = .W, n = 408). The slope was greater than 0 (P .ooOl). bpooled Cu was different from the pooled r ession (P < .05). The regression intercept was 135 mg/kg (SE = .27), and the slope was -.034 mg/d (SE = .W, R - 22; n = 102).
T-
cPo~ledFe was different from the pooled regression (P < .OM).The regression intercept was 14.1 m and the slope was -.043 mgld (SE = .W, R2 = .32;n = 102).
a (SE = 27),
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loss in these diets was .015 mg/d during 84 d of storage. Although this rate of loss was significant statistically, in diets containing 20 lU of a-tocopheryl acetate, 93% of the acetate would remain after 90 d of storage, indicating that a-tocopheryl acetate is relatively stable in mixed diets. Klaiii (1973) reported similar rates of loss of a-tocopheryl acetate during 6 mo of storage in complete poultry diets in meal form, but less was lost from pelleted diets. Klatii (1973) also reported that atocopheryl acetate was stable in concentrated premixes. High levels of Cu (250 ppm; P e .E)or Fe (1,O00 ppm; P e .005)increased the rate of loss to .OM and .043 mdd, respectively. There was no difference (P > .l) in rate of a-tocopheryl acetate loss between diets with or without added soybean oil. The results of our experiment suggest that 250 pp of Cu or 1,OOO ppm of Fe decreased the stability of atocopheryl acetate. The natural a-tocopheryl concentration (Table 4) in the NOTM and 'I'M diets not containing soybean oil decreased (P e .01) during the 846 storage period. A single component exponential decrease described the rate of loss of a-tocopherol and indicated a half-life of 84 and 91 d for the NOTM and TM
TABLE 2. ANALYZED COMPOSmON OF DIETS
1997
EFFECT OF MINERALS ON VITAMIN E STABILITY
TABLE 4. EFFECT OF TRACE MINERALS ON ALPHA-TOCOpHEROLa LOSS DURING STORAGE Trratment
Cu
Thf
zn
Fe
Mn
Basal 4.43 .03
Intercept
SE
3.77
SE
.ooo9
.o007
59 91
.70
R2 Half-life, d
84
Intercept Slope
5.23 .03 -.00918
455 .04 -.0134h
SE
.ooo8
.ooo9
SE
.72
R2 Half-life. d
.82
76 ~
= log
52 ~~
2.79 .08
-.036Sd -.om* .0127 .0018 .14 .63 19 42 Basal + soybean oilf .46 2.92 57 .13 -.O3Ni -.0543i .W29 .0130 26 .72 13 21
-.me
-.082c
Slope
.34 .55
.04
423 .04 -.0121& .Oo08 .82 57
4.34 .04 -.0116’ .0010 .74 60
5.09
5.04
.05
.05
-.02Ooi .m11 .87
-.O15Sh .m12 .76
44
34
~~~~~
~~~
+b
(days of storage). Tocopherol = ae*. bDiets that did not contain kace minerals are NOTM and diets containing only the trace m i n d premix are TM. c*4eSlopeswithin the basal diet that do not share the same superscript are different (P < -05). ‘DBerenCe b e e n basal and basal + soybean oil diets within TM, Fe, Zn (P < .005), or Mn (P < .05). gJsiJSlopes within the basal + soybean oil diet that do not share the same superscript are different (P c .01). %g tocopherol
a
of Cu, Fe, Zn, or Mn to the diets without added soybean oil reduced the half-life of atocopherol to 19,42,57, and 60 d, respectively. Young et al. (1975) found that the atocopherol in mixed diets containing &cidy dried corn was stable during storage.
diets, respectively. This loss of natural tocopherols in the NOTM diet during storage suggests that the natural tocopherols have a limited stability. The rate of loss of atocopherol was increased by the addition of either Cu,Fe, Zn, or Mn (P < .05) to the TM diet without added soybean oil. The addition
TABLE 5. EPFECT OF Cu OR Fe ON ALPHA-TOCOPHEROLa LOSS DURING STORAGE Fe
c11 Basal+ Item
Basal
soybean oil
Basal+
Basal
soybean oil
Fast
Intercept
SE Slope
SE R2 Half-life, d
5.41 .52 -26Sb
.070 .83 2.6
6.13 .47 -.226b .064
2.29 .34 -.11@ .027 .75 6.0
.so
3.1
5.76 .34 -. 165d .027 .85 42
Slow
Intercept SE Slope
SE R2 Half-life, d
20 .66 -.W1 .0143 .03 76
.33 .6 1 -.0263e .0136 .24 26
1.84
.IS -.0088 .m7 53 79
1.15 .15 -.0146 .0048 .50 47
‘Log tocopherol = wg a + b (days of storage)] + w g a + b (days of storage)]. Tocopherol = ae-’ b*c*d~~opes within the fast component that do not share the same supacsipt are different (P < .01). %fference between Cu and Fe within the basal + soybean oil diet (P < .01).
+
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NOTMb
Item
1998
DOVE AND EWAN
D-ALPHA
TOCOPHEROL
--- Exponential - Exponential - slow - rapid
0 Exponential
- sum
I
OI Y
\
F ,001 1 0
10
30
20
40
50
70
60
80
90
Days SBO
+
--.Exponential
- slow
- Exponential - rapid
o Exponential
-
sum
0 Y
\
4
0
E
--
I
4 4
4
.01 * 4
.001
0
10
20
30
40
50
60
70
80
90
0ays Pigme 1. Effect of Cu (250 ppm) on the rate of loss of natural tocopherol. Observed values, slow rate of loss, -----, predicted rate of loss, 0.
*; rapid rate of loss, -;
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The diets used by Young et al. (1975) the addition of trace mineral mixes to swine contained ethoxyquin, whereas the diets used diets accelerated the decrease in the natural in our experiment did not. This difference in tocopherol levels during storage. In the experithe storage stability of the natural tocopherols ment reported here, the rate of loss of amay be a result of the increased demand for a tocopherol was not affected by the addition of dietary antioxidant in the diets used in this trace minerals in diets that did not contain experiment. Richter et al. (1982) reported that added soybean oil.
EEFECT OF MINERALS ON VITAMIN E STABILsry
associated with added dietary fat. No attempt was made in this study to determine the products associated with the loss of tocopherols. Assuming that loss was the result of oxidation, the major products would be a-tocopheryl quinone and a-tocopheroxide (Diplock, 1985); these compounds do not have biological activity (Kasparek, 1980). In the analytical system used in this study, atocopheryl quinone would not be detected. The increase in the rate of loss promoted by trace minerals in this study is consistent with the oxidation of tocopherols. Gamma-tocopherol and a-tocopherol levels (data not shown) were affected similarly by all mineral additions and by addition of soybean Oil. Implications
Alpha-tocopheryl acetate was relatively stable during storage, but the stability of natural tocopherol was limited in diets that do not contain supplemental trace minerals. The stability of a-tocopheryl acetate was decreased in the presence of high levels of Fe or Cu, whereas the rate of loss of natural tocopherols was increased by high levels of all minerals tested (Cu > Fe > Zn > Mn). Corn-soybean meal diets containing growth-promoting levels of Cu or high levels of trace minerals may require the addition of a-tocopheryl acetate to maintain recommended levels of vitamin E in diets during storage, especially in the presence of added unsaturated fat. Literature Cited
Colt, W. M.. T. S. Vicente, E. H. Waysek and B. D. Williams. 1983. Vitamin E content of feedstuffs determined by high-perfomunce liquid chromotographic fluorescence. J. Agric. Food Chem 31:1330. Diplock, A. T. 1985. Vitamia E. In:A. T. Diplock (Ed.) Fatsoluble Vitamins. pp 154-224. Heineman, London. Dove,C.RandR.C.Ewan. 1987.ThecffcctofCyFeorZn addition on the vitamin E status of growing pigs and the natural tocopherol levels in feed.J. Anim. Sci. 65 (Suppl. 1):134 (Abstr.). Dove, C. R and R. C. Ewaa 1990. The & s t of excess dietary copper, iron or zinc on the tocopherol and selenium status of pwingpigs. J. Anim. Sci. 68:2407. Ewaq R. C. 1985. Analysis of feeds and feed ingredients 1984. ISU Research Reports - 1985. Iowa State Univ., AmeS. Ewan, R.C. 1986. Analysis of feeds and feed ingredients 1985. ISU Swine Resrach Reports - 1986. Iowa State univ., Ames. Hawrarainen,R. V.,I. T.Tytippbea and S. G. Bengtsson. 1983a. changes in the content and composition of vitamin E in damp barley stored in airtight bios. J. Sci.
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The addition of soybean oil did not affect the rate of loss of a-tocopherol in the NOTM diet (P> .1). However, in the presence of TM, Fe, Zn (P < .005), or Mn (P < .05), the addition of soybean oil increased the rate of loss of tocopherol. Within the diets with added soybean oil, Cu and Fe additions increased the rate of loss (P< .01) more than the rate of loss in diets with added Zn.The rate of loss in diet with added soybean oil and Zn was greater (P < .01) than the loss in diets containing soybean oil and TM or Mn. The half-life of atocopherol was reduced to 52,21,34, and 44 d by the presence of TM, Fe, Zn, or Mn, respectively, in diets with added soybean oil. The rate of a-tocopherol loss was increased greatly in diets with 250 ppm of added Cu and in the diet containing 1,OOO ppm of Fe. The rate of loss was a two-component curve (Figure 1) characterized by a rapid rate of loss initially and a second component with a rate of loss similar to that in the NOTM or TM diets. The fast component as characterized by halflives ranging from 2.6 to 6 d in the diets containing high levels of Cu or Fe (Table 5). The addition of soybean oil increased the fast component in the presence of added Fe (P e .01) but did not affect the fast component in the presence of added Cu. Because the rates of loss in the presence of Cu were not affected by the addition of soybean oil and because the rates were very rapid, Cu may have a direct effect on tocopherol loss that is not dependent on the production of lipid peroxides. The halflife of a-tocopherol in diets containing 250 ppm of Cu predicts a reduction to less than 10% of the initial concentration after about 10 d of storage. Growth-promoting levels of Cu (125 to 250 ppm) are used widely in the swine industry (Ewan, 1986), and Cu had the greatest destructive effect on the natural tocopherols. Ionized Cu and Fe often are included in lists of factors that increase the rate of loss of natural tocopherols (ullrey, 1981). Waddell and Steenbock (1931) associated the presence of dietary Fe with the loss of natural tocopherols in rat diets. The increased rate of a-tocopherol loss suggests that the presence of trace minerals in diets increases the peroxidation of fats in the diet, increasing the need for dietary antioxidants. The failure of added soybean oil to increase the rate of tocopherol loss in the NOTM diet suggests that trace minerals are hportant in the mechanism of tocopherol loss
1999
2000
DOVE AND EWAN Blaisdale Publishing Co., Waltbam. MA. Richter, V. G., P. Mockel, F. Schone and G. Hachowsky. 1982. Effect of storage time and storage temperature on vitamin A and E levels in vitamin concentrates, premixes of active ingredients, mixedfodder,and liver homogenates. Monats. vetuinaermtd. 3 7 4 0 5 . SAS. 1982. SAS User's Guide: Stafjstics. SAS Inst, Inc., Gary, NC. Steel, RGD. and J. H. Tome. 1960. Principles and F'rucedurcs of Statistics. McGraw-HillBook Co.. New York Ullrey,D. E. 1981. Vitamin E for swine. J. Anim. Sci. 53: 1039. Waddell,J.audH. Steenbock. 193LVitamhEinFelreated dry rations. J. Nutr. 479. Young, L. G.. A. Lun. J. Pos, R. P. Porshaw and D. Edmeades. 1975.VitaminEstabilityincornandmixed feed. J. Anim. Sci. 40:495.
Downloaded from https://academic.oup.com/jas/article-abstract/69/5/1994/4705365 by Iowa State University user on 11 January 2019
Food Agric. 34:1029. Hakkamhen, R.V., J. T. Ty6p-n and S. G. Bengtsson. 1983b. Relative and quantitative changes in total vitamin E and isomer content of barley during cowentional and airtight storage with special refercnce to dvariations. Acta A@. Scaud. 39395. Kasparelr, S. 1980. chemistry of tocopherols and tocotrienols. In: L. J. Uachlin @Vitamin I.) E. A Comprehensive 'Reatise. pp 7-65. Marcel M e r , New Yo&. Klaiii, H. 1973. 'Ihc stability and distribution m feeds of powders colllaining vitamin E. Acta Agric. Scand. (SuppL) 19219. NRC. 1979. Nutrient Rqdrements of Swine (8th Ed.). National Academy Press, Washington, DC. NRC. 1988. Nutrient Requirements of Swine (9th Ed.). National Academy Press, Washington, DC. Rescigno, A. and G. Segre. 1%6. Drug and Tracer Kinetics.
