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Effects of Dietary Fat Level on Laying Hens Fed Various Concentrations of Calcium
METABOLISM AND NUTRITION Effects of Dietary Fat Level on Laying Hens Fed Various Concentrations of Calcium J. O. ATTEH and S. LEESON Department of Animal and Poultry Science, University of Guelph, Guelph, Ontario, Canada N1G 2W1 (Received for publication May 4, 1984)
1985 Poultry Science 64:2090-2097 INTRODUCTION T h e interference of f a t t y acid c o m p o n e n t of fat w i t h mineral metabolism in broiler chicks has been d o c u m e n t e d (Griffith et al., 1 9 6 1 ; Hakansson, 1 9 7 5 a ; Gardiner and Whitehead, 1 9 7 6 ; A t t e h and Leeson, 1 9 8 3 b ) . This is d u e primarily t o t h e f o r m a t i o n of f a t t y acid a n d cation soaps during t h e process of digestion. S o m e of t h e soaps formed, particularly t h o s e of calcium and magnesium, are insoluble (West et al, 1966) and hence, render n o t only t h e f a t t y acids involved b u t also t h e mineral concerned unavailable t o t h e chicken. T h e age of t h e bird has been s h o w n t o influence utilization of dietary fats and f a t t y acids ( D u c k w o r t h et al., 1 9 5 0 ; R e n n e r and Hill, I 9 6 0 ; Whitehead and Fisher, 1 9 7 5 ) . T h e i m p r o v e m e n t in fat utilization with age m a y be associated with a fully developed absorptive m e c h a n i s m in t h e adult birds because t h e absorptive m e c h a n s i m for lipid is n o t fully developed in chicks (Polin and Hussein, 1 9 8 2 ) . T h e r e is also evidence t o suggest t h a t a decrease in rate of soap f o r m a t i o n and differences in t h e p r e d o m i n a n t area of t h e gut w h e r e soap is formed in adult birds, c o m p a r e d t o broiler chicks leads t o improved fat utilization ( A t t e h and Leeson, 1 9 8 4 ) . T h e s t u d y reported herein was u n d e r t a k e n n o t only t o c o m p a r e t h e
response of laying hens fed various levels of dietary animal-vegetable blend fat t o t h a t of chicks fed various levels of fat ( A t t e h et al., 1 9 8 3 ) , b u t its p u r p o s e was also t o c o m p a r e their response t o t h a t of laying hens fed free fatty acids ( A t t e h and Leeson, 1985) in t h e presence of varying levels of dietary calcium. MATERIALS AND METHODS Seventy-two individually caged White Leghorn hens of a commercial strain t h a t had been in p r o d u c t i o n for 10 w e e k s were used in this s t u d y . Nine e x p e r i m e n t a l diets consisted of a 3 X 3 factorial c o m b i n a t i o n of levels of animalvegetable fat (A-V blend) and dietary calcium levels (Table 1). T h e fat levels s u p p l e m e n t e d were 0, 5.0, or 10.0%, and t h e y were substituted for corn starch in t h e control diet, whereas t h e dietary calcium levels were 3.0, 3.6, and 4 . 2 % . Calculated metabolizable energy (ME) for all diets was a p p r o x i m a t e l y 2 7 5 0 kcal/kg. Eight birds were r a n d o m l y allocated t o each diet; each bird was individually fed, t h u s serving as a replicate. E x p e r i m e n t a l diets and water were supplied ad libitum and light m a i n t a i n e d at 14 hr per day d u r i n g t h e 7-week trial period. Feed c o n s u m p t i o n was measured o n each of 2 consecutive days each week, which a c c o u n t e d
2090
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ABSTRACT The effects of supplementing laying hen diets with 0, 5, or 10% fat in the presence of 3.0, 3.6, or 4.2% dietary calcium was investigated using laying hens over a 7-week period. There was no significant effect of the fat or calcium treatments on laying hen performance as judged by feed intake, weight gain, egg production, egg weight, and egg shell deformation. Although there was an increase in fat retention with an increase in dietary fat level (P<.01), a larger proportion (P<.05) of unabsorbed fat was present as soap in the excreta of birds on these diets relative to those fed the control diet. There was evidence to indicate that most of the soap observed in the excreta of laying hens was formed in postabsorptive areas of the gut, thereby explaining why the detrimental effects of soap formation were not seen in the laying hen. Increasing the dietary calcium level reduced both percentage calcium (P<.01) and magnesium (P<.05) retention. There were no significant effects of the treatments on nitrogen and phosphorus retention. There were also no significant effects of the treatments on shell ash, shell calcium, magnesium, and phosphorus content. Increasing the dietary calcium level increased bone ash (P<.05) while reducing bone magnesium content (P<.05). There were no significant effects of the fat and calcium treatments on bone calcium and phosphorus content. (Key words- fat, calcium, layer performance, bone minerals)
17.1 3.06 .75 .13 1.63
Analyzed nutrient content Crude protein, % Calcium, % Total phosphorus, % Magnesium, % Crude fat, %
Calculated metabolizable energy for all diets, 2750 kcal/kg.
10.05
67.90 22.05
17.4 4.18 .77 .14 1.67
100.00
3
17.1 2.92 .81 .12 6.45
67.90 11.15 5.00 6.89 9.06 100.00
4
('«)
17.1 3.63 .76 .13 6.54
67.90 11.15 5.00 8.47 7.48 100.00
5
diets1
6
17 4
67 11 5 10 5 100
6
3
A-V = Animal-vegetable blend.
Contain corn, 37.75%; soybean meal (48%), 23.80%; corn gluten meal, 3.40%; calcium phosphate, 1.55%; DL-m .25%; chromic oxide, .30%; mineral-vitamin premix, .75% [provides per kilogram of diet: vitamin A, 8000 IU; vita mg; pantothenic acid, 7.0 mg; vitamin B 1 2 , 8 Mg; niacin, 20 mg; choline, 900 mg; vitamin K, 1.5 mg; folic acid, 1.5 mg; manganese, 50 mg; zinc, 50 mg; copper, 5.0 mg; iron 30.0 mg].
2
1
8.47 1.58 100.00
6.89 3.16 100.00
17.4 3.56 .79 .13 1.70
67.90 22.05
67.90 22.05
Basal ingredients 2 Corn starch A-V3 blend fat Limestone Alpha-floc (wood cellulose) Total
2
1
Treatment number
TABLE 1. Experimental
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2092
ATTEH AND LEESON
cedure (Kjel-Foss Automatic 16310), whereas gross energy was determined by adiabatic oxygen bomb calorimetry (Parr, Model 1241). Ash samples from feed, excreta, shell, and bone were digested using the method of Association of Official Analytical Chemists (AOAC, 1980). The resulting solutions were analyzed for calcium and magnesium using a Techtron (Model AA 4 ) atomic absorption spectrophotometer and phosphorus using a Technicon (Model AA 2 ) autoanalyzer. The presence of chromium in the feed and the excreta was determined using the method of Fenton and Fenton (1979). Total glycerides and fatty acids in feed, digesta, and excreta were determined by petroleum ether extraction using Soxhlet apparatus. To estimate the proportion of the fatty acids in the digesta and excreta that was present as soap, the method of fat determination reported by Carroll (1958) was used. The digesta and excreta were subjected to two stages of ether extraction. The first extraction was to remove neutral fat and fatty acids. The residue from the first extraction was subjected to acid treatment using 25% hydrochloric acid (specific gravity, 1.13) for 2 hr at room temperature to liberate fatty acids that were present as soap. The samples were then freeze dried and the process of ether extraction repeated. This second extraction was considered to represent fatty acids previously
TABLE 2. Effects of dietary levels of fat (F) and calcium (Ca) on performance of laying bens
Dietary treatments
Feed intake
Weight gain over 7-week period 1
Egg production (HDB) 2
Egg weight
Egg shell deformation
(g/hen/day)
(g/bird)
(%)
(g)
(um)
NS 3 109.7 108.6 109.3
NS 42.3 97.9 82.3
NS 90.6 94.6 90.9
NS 56.6 57.0 56.6
NS 25.6 25.6 25.2
Ca level, % 3.0 3.6 4.2
NS 113.5 108.7 105.5
NS 78.5 76.3 67.7
NS 91.0 93.1 92.0
NS 55.8 57.0 57.5
NS 26.2 25.6 24.6
Ca X F
NS
NS
NS
NS
NS
10.1
11.6
Supplemental F, 0 5.0 10.0
%
Standard deviation 1
12.9
Average for 7 weeks.
2
Hen-day basis.
3
NS = No significant difference (P>.05).
3.9
2.7
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for the disappearance in 24 hr of part of a 200 g feed allocation. Eggs were also collected twice a week for weighing and determination of shell deformation as a measure of shell quality using the method of Summers et al. (1976). Shells from the first egg collection each week were oven dried and ground prior to ashing at 600 C for 48 hr. Egg production was recorded throughout the trial. During the 4th week of the experiment, excreta samples were taken in order to estimate nutrient retention. Chromic oxide was used as a marker (Table 1) and excreta collection was undertaken over a 72-hr period. The excreta samples were dried in a forced air oven at 70 C and ground prior to ashing at 600 C overnight and other subsequent chemical analysis. At the conclusion of the feeding trial, four hens per treatment were killed by cervical dislocation within 30 min of oviposition and the body cavity was immediately opened. The contents of the small intestine (digesta) were then flushed out with double distilled water and freeze dried prior to grinding. Also, the left tibia of each bird was removed and cleaned of adhering flesh, dried at 100 C for 24 hr, defatted using a Soxhlet extraction apparatus, and dried a second time prior to ashing at 600 C overnight. Ash weight was taken to estimate percentage bone ash. Chemical Analysis. Nitrogen in both feed and excreta was determined by Kjeldahl pro-
3
2
1
3.4
2.9
NS 5.2
NS
NS 28.6 b 27.2 b 23.3 a
NS 27.6 26.0 25.5
**
*
4.1
NS 4.0
NS
83.5 82.4 82.1
77.6 a 83.8b 86.6 C
56.4 b 54.4 b 49.1 a
**
F
NS 54.9 53.0 52.0
Ca
25
NS
NS 2838 2789 2803
NS 2782 2796 2853
(kcal
ME
NS = No significant difference (P>.05)'
Excreta ether Extract 2 as a proportion of Extracts 1 plus 2.
Digesta ether Extract 2 as a proportion of Extracts 1 plus 2.
' ' Within main treatment categories, means within column followed by different superscripts are significantly di
Standard deviati on
NS
CaX F
NS 41.1 38.8 40.3
NS 38.7 40.8 40.8
NS 3 56.8 55.3 55.1
NS 55.2 55.3 56.7
%
Magnesium
Nutrient retention
Phosphorus :
Nitrogen
Ca level, % 3.0 3.6 4.2
Supplemental F, 0 5.0 10.0
Dietary treatments
TABLE 3. Effects of dietary levels of fat (F) and calcium (Ca) on nutrient retention by laying hens, dietary
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ATTEH AND LEESON
2094
present as soap. Samples of the feed were subjected to similar two-stage ether extractions to act as standards. Fats and fatty acids extracted from feed and that from each of the two extractions from the digesta and excreta were then esterified with .2 N methanolic trimethyl ammonium hydroxide, and the methyl esters were analyzed for component fatty acids by gas-liquid chromatography (Varian, Model 2100) using the method of AOAC (1980). Data collected were subjected to analysis of variance using Statistical Analysis System (SAS, 1982). Significant differences among treatments were determined by Duncan's new multiple range test (Duncan, 1955).
—< r-~ 0\ o* r~ r-H q o. >o H q ' N ^ i M 06 N
h\
RESULTS
Layer Performance. There was no significant effect of the fat and calcium treatments on feed intake, although there was a tendency for increases in dietary calcium level to reduce intake (Table 2). There were also no significant effects of the treatments on weight gain, egg production, egg weight, and egg shell quality (P>.05). Nutrient Retention. There was no significant effect of the fat or calcium levels on ME of the diets. Also, there was no significant effect of the treatments on nitrogen and phosphorus retention (Table 3). While increasing the dietary fat level had no significant effect on calcium and magnesium retention, increase in dietary calcium levels resulted in a decrease in percentage calcium (P<.01) and magnesium (P<.05) retention. Increases in dietary fat level resulted in an increase in fat retention (P<.01). However, a larger percentage of unabsorbed fat was present as soap with increases in dietary fat (P<.05). Although all fatty acids were involved in soap formation, palmitic and stearic acids were responsible for more than 50% of the soap formed during the process of digestion (Table 4). There was no significant interaction between dietary fat and calcium levels on the retention of any of the nutrients measured. Egg Shell Minerals. There were no significant effects of the treatments on shell ash, shell calcium, phosphorus, or magnesium, although increases in dietary calcium level tended to increase shell calcium and decrease shell magnesium content (Table 5). In all cases these effects were not statistically significant (P> .05). Bone Ash and Minerals. There was no significant effect of the dietary fat level on
i 0\ « m r j ir\ ' © ">_ t~; © rrt ' N
4
tfl
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6 S
NS 47.8 48.2 48.6
NS
Ca level, % 3.0 3.6 4.2
CaX F
2.6
NS
NS 37.5 37.9 39.2
NS 38.4 38.6 37.7
Ca
.001
NS
NS .08 .08 .08
NS .08 .08 .08
Phosphorus
Shell
.02
NS
NS .36 .36 .35
NS .37 .35 .36
Magnesium
1.8
NS
54.5 a 54.5 a 56.0 b
•
NS 55.4 54.4 54.5
Ash 2
N
N 3 3 3
N 3 3 3
C
Average for 7 consecutive weeks.
NS = No significant difference (P>.05).
Percentage of bone ash.
On dry, fat free basis.
*P<05.
4
3
2
1
ab ' Within main treatment categories, means within column followed by different superscripts are significantly diffe
1.8
NS" 48.6 48.2 47.9
Supplemental F, % 0 5.0 10.0
Standard deviation
Ash
Dietary treatments
TABLE 5. Effects of dietary levels of fat (F) and calcium (Ca) on egg shell and bone mineral co
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2096
ATTEH AND LEESON
bone ash. However, increasing the dietary calcium content significantly (P<.05) increased bone ash (Table 5). Increasing the dietary fat level had a tendency to decrease bone calcium content, whereas increasing the dietary calcium level had the opposite effect on bone calcium content. However, in both cases, the effects were not statistically significant (P>.05). There was no significant interaction between dietary fat and calcium levels on any of the bonerelated measurements.
The nonsignificant effect of the treatments on the performance variables measured was probably due to a comparable energy content of the diets. This showed that fat per se had no significant effect on laying hen performance, an observation which confirms earlier reports (Vogt and Harnisch, 1976; Horani and Sell, 1977) showing that dietary fat had no significant effect on these parameters. However, this is in contrast to the situation observed in chicks (Johri and Narayanan, 1972; Mateos and Sell, 1980; Atteh et al, 1983) showing improvement in weight gain and energetic efficiency of feed with fat supplementation. Performance of laying hens fed diets supplemented with A-V blend fat was comparable to those fed diets supplemented with free fatty acids (Atteh and Leeson, 1984b) with the notable exception of palmitic acid which laying hens could not utilize efficiently. The tendency for increases in dietary calcium level to reduce feed intake agrees with a previous report (Atteh and Leeson, 1984b) which showed that laying hens reduced intake when supplied diets high in calcium. Supplemental fat was added to the diets at the expense of corn starch to provide isocaloric diets. However, the ME content of diets supplemented with fat were slightly higher than in the control diet and this "extra caloric" effect, though higher with increase in fat supplementation, was not as profound as the situation observed in chicks (Cullen et al, 1962; Jensen et al, 1970; Atteh et al, 1983) or laying hens fed diets supplemented with free fatty acids with the exception of palmitic acid (Atteh and Leeson, 1984b). The improvement in fat retention with increase in dietary fat supplementation is probably associated with increase in proportion of oleic acid in the diet which has been shown to facilitate the absorption of other fatty
The changes in fatty acid profile of diet fat during digestion confirm earlier observations (Renner and Hill, 1961; Gardiner and Whitehead, 1976; Atteh and Leeson, 1983b, 1984) of inherent problems of absorption of saturated fatty acids. Thus, the proportion of palmitic and stearic acids increased from 11.94 and 2.31% in the control diet to 15.3 and 9.76% in the nonsoap portion of unabsorbed fat and to 35.89 and 22.82% of soap portion of unabsorbed fat, respectively. Similar trends for these two fatty acids were also evident in diets supplemented with 5 or 10% fat. Although all the component fatty acids in the fat were involved in soap formation, there is evidence to suggest that the soaps of myristic and oleic acids were absorbed to a large extent, because the proportion of fatty acids from soap fat accounted for by myristic and oleic acids was low compared to their proportion in the soap fat of the digesta. This observation supports the reports of Boyd et al. (1932) and Atteh and Leeson (1984) showing that soaps of oleic acid were well utilized. The significant decrease in magnesium retention and bone magnesium content with increase in dietary calcium content showed that high levels of calcium interfered with magnesium metabolism and this interference was independent of the dietary fat level. This agrees with the reports of Hakansson (1975b) and Atteh and Leeson (1983a, 1985). Although there was a decrease in the percentage calcium retention with the increase in dietary calcium level, in absolute terms, birds on diets with 4.2% calcium retained as much calcium as those on diets with lower levels of calcium. This would explain why the calcium treatment had no significant effect on shell and bone calcium content. The fact that the fat treatment had no
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DISCUSSION
acids (Young and Garett, 1963). There is evidence to suggest that most of the soap formed during digestion in laying hens occurred in postabsorptive areas of the gut. Thus, despite the fact that birds on diets supplemented with 10% fat had the highest fat retention, the proportion of unabsorbed fat that was present as soap was highest with these same birds. This indicates that most of the soap observed in the excreta was formed after most absorption had taken place. This observation confirms earlier findings (Atteh and Leeson, 1984b) and would explain why detrimental effects of soap formation were not seen in the laying hen.
FAT AND CALCIUM FOR LAYERS
REFERENCES Association of Official Analytical Chemists, 1980. Pages 21 and 447 in Official Methods of Analysis, 13th ed. Assoc. Offic. Anal. Chem., Washington, DC. Atteh, J. O., and S. Leeson, 1983a. Influence of increasing dietary calcium and magnesium levels on performance, mineral metabolism, and egg mineral content of laying hens. Poultry Sci. 62:1261-1268. Atteh, J. O., and S. Leeson, 1983b. Effects of dietary fatty acids and calcium levels on performance and mineral metabolism of broiler chickens. Poultry Sci. 62:2412-2419. Atteh, J. O., and S. Leeson, 1984. Effects of dietary saturated, unsaturated fatty acids, and calcium levels on performance and mineral metabolism of broiler chicks. Poultry Sci. 63:2252-2260. Atteh, J. O., and S. Leeson, 1985. Response of laying hens to dietary saturated and unsaturated fatty acids in the presence of varying dietary calcium levels. Poultry Sci. 64:520-528. Atteh, J. O., S. Leeson, and R. J. Julian, 1983. Effects of dietary levels and types of fat on performance and mineral metabolism of broiler chicks. Poultry Sci. 62:2403-2411. Boyd, S. F., C. L. Crum, and J. F. Lyman, 1932. The absorption of calcium soaps and the relation of dietary fat to calcium utilization in the white rat. J. Biol. Chem. 9 5 : 2 9 - 4 1 . Carew, L. B., R. H. Machemer, R. W. Sharp, and D. C. Foss, 1972. Fat absorption by the very young chick. Poultry Sci. 51:738-742. Caroll, K. K., 1958. Digestibility of individual fatty acids in the rat. J. Nutr. 64:399-410. Cullen, M. P., O. G. Rasmussen, and O.H.M. Wilder, 1962. Metabolizable energy value and utilization of different types and grades of fat by the chick. Poultry Sci. 41:360-367. Dewar, W. A., C. C. Whitehead, J. N. Downie, and E. Potter, 1975. The retention of calcium, iron, magnesium and zinc in chicks fed on diets
containing metal soaps. Proc. Nutr. Soc. 34: 5A-6A. Duckworth, J., J. M. Naftalin, and C. Dalgarno, 1950. The digestibility of linseed oil and mutton fat by chicks. J. Agric. Sci. 4 0 : 3 9 - 4 3 . Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics 11:1—42. Fenton, T. W., and M. Fenton, 1979. An improved procedure for determination of chromic oxide in feed and feces. Can. J. Anim. Sci. 59:631—634. Gardiner, E. E., and C. C. Whitehead, 1976. Effect of dietary palmitic acid on broiler chicks fed on various concentrations of calcium. Br. Poult. Sci. 17:241-244. Griffith, F. D., R. B. Grainger, and J. J. Begin, 1961. The effect of dietary fat and cellulose on apparent calcium digestibility in growing chickens. Poultry Sci. 40:1492-1497. Hakansson, J., 1975a. The effect of fat on the retention of calcium, phosphorus and magnesium in laying hens. Swed. J. Agric. Res., 5:3—9. Hakansson, J., 1975b. The effect of fat on calcium, phosphorus and magnesium balances in chicks. Swed. J. Agric. Res. 5:145-157. Horani, F., and J. L. Sell, 1977. Effect of feed grade animal fat on laying hen performance and on metabolizable energy of rations. Poultry Sci. 56:1972-1980. Jensen, L. S., G. W. Schumainer, and J. D. Latshaw, 1970. "Extra calorie" effect of dietary fat for developing turkeys as influenced by calorieprotein ratio. Poultry Sci. 49:1697-1704. Johri, T. S., and S. Narayanan, 1972. Effect of animal fats on growth of chicks. Indian J. Anim. Sci. 42:835-839. Mateos, G. G., and J. L. Sell, 1980. Influence of fat on energy utilization from selected carbohydrates. Poultry Sci. 59:1635. Polin, D., and T. H. Hussein, 1982. The effect of bile acid on lipid and nitrogen retention, carcass composition and dietary metabolizable energy in very young chicks. Poultry Sci. 61:1697—1707. Renner, R., and F. W. Hill, 1960. The utilization of corn oil, lard and tallow by chickens of various ages. Poultry Sci. 39:849-854. Renner, R., and F. W. Hill, 1961. Utilization of fatty acids by the chicken. J. Nutr. 75:259—264. Statistical Analysis System (SAS), 1982. Page 119 in SAS User's Guide: Statistics. SAS Inst. Inc., Cary, NC. Summers, J. D., R. Grandhi, and S. Leeson, 1976. Calcium and phosphorus requirements of the laying hen. Poultry Sci. 55:402-413. Vogt, H., and S. Harnisch, 1976. Comparison of different fats in complete feeds for laying hens. Arch. Gefluegelkd. 40:168-176. West, E. S., W. R. Todd, H. S. Mason, and J. T. Van Bruggen, 1966. Page 131 in Textbook of Biochemistry. 4th ed. MacMillan Co., London, England. Whitehead, C. C , and C. Fisher, 1975. The utilization of various fats by turkeys of different ages. Br. Poult. Sci. 16:481-485. Young, R. J., and R. L. Garrett, 1963. Effect of oleic and linoleic acids in the absorption of saturated fatty acids in the chick. J. Nutr. 81:321-329.
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significant effect on calcium and magnesium retention, shell and bone calcium, and magnesium content showed that up to 10% fat in the diet of laying hens is not detrimental to the metabolism of these minerals. This is in contrast to observations with chicks showing that addition of fat to their diets reduced not only the retention of calcium and magnesium through soap formation but also reduced the bone content of these minerals (Griffith et al, 1961; Dewar et al, 1975; Atteh et al, 1983). This variation between the young and adult bird in fat and mineral metabolism may not only be associated with lack of full physiological capacity for fat absorption by chicks (Carew et al, 1972) but also with a difference in the predominant site of soap formation in relation to area of fat absorption.
2097
1985 Poultry Science 64:2090-2097 INTRODUCTION T h e interference of f a t t y acid c o m p o n e n t of fat w i t h mineral metabolism in broiler chicks has been d o c u m e n t e d (Griffith et al., 1 9 6 1 ; Hakansson, 1 9 7 5 a ; Gardiner and Whitehead, 1 9 7 6 ; A t t e h and Leeson, 1 9 8 3 b ) . This is d u e primarily t o t h e f o r m a t i o n of f a t t y acid a n d cation soaps during t h e process of digestion. S o m e of t h e soaps formed, particularly t h o s e of calcium and magnesium, are insoluble (West et al, 1966) and hence, render n o t only t h e f a t t y acids involved b u t also t h e mineral concerned unavailable t o t h e chicken. T h e age of t h e bird has been s h o w n t o influence utilization of dietary fats and f a t t y acids ( D u c k w o r t h et al., 1 9 5 0 ; R e n n e r and Hill, I 9 6 0 ; Whitehead and Fisher, 1 9 7 5 ) . T h e i m p r o v e m e n t in fat utilization with age m a y be associated with a fully developed absorptive m e c h a n i s m in t h e adult birds because t h e absorptive m e c h a n s i m for lipid is n o t fully developed in chicks (Polin and Hussein, 1 9 8 2 ) . T h e r e is also evidence t o suggest t h a t a decrease in rate of soap f o r m a t i o n and differences in t h e p r e d o m i n a n t area of t h e gut w h e r e soap is formed in adult birds, c o m p a r e d t o broiler chicks leads t o improved fat utilization ( A t t e h and Leeson, 1 9 8 4 ) . T h e s t u d y reported herein was u n d e r t a k e n n o t only t o c o m p a r e t h e
response of laying hens fed various levels of dietary animal-vegetable blend fat t o t h a t of chicks fed various levels of fat ( A t t e h et al., 1 9 8 3 ) , b u t its p u r p o s e was also t o c o m p a r e their response t o t h a t of laying hens fed free fatty acids ( A t t e h and Leeson, 1985) in t h e presence of varying levels of dietary calcium. MATERIALS AND METHODS Seventy-two individually caged White Leghorn hens of a commercial strain t h a t had been in p r o d u c t i o n for 10 w e e k s were used in this s t u d y . Nine e x p e r i m e n t a l diets consisted of a 3 X 3 factorial c o m b i n a t i o n of levels of animalvegetable fat (A-V blend) and dietary calcium levels (Table 1). T h e fat levels s u p p l e m e n t e d were 0, 5.0, or 10.0%, and t h e y were substituted for corn starch in t h e control diet, whereas t h e dietary calcium levels were 3.0, 3.6, and 4 . 2 % . Calculated metabolizable energy (ME) for all diets was a p p r o x i m a t e l y 2 7 5 0 kcal/kg. Eight birds were r a n d o m l y allocated t o each diet; each bird was individually fed, t h u s serving as a replicate. E x p e r i m e n t a l diets and water were supplied ad libitum and light m a i n t a i n e d at 14 hr per day d u r i n g t h e 7-week trial period. Feed c o n s u m p t i o n was measured o n each of 2 consecutive days each week, which a c c o u n t e d
2090
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ABSTRACT The effects of supplementing laying hen diets with 0, 5, or 10% fat in the presence of 3.0, 3.6, or 4.2% dietary calcium was investigated using laying hens over a 7-week period. There was no significant effect of the fat or calcium treatments on laying hen performance as judged by feed intake, weight gain, egg production, egg weight, and egg shell deformation. Although there was an increase in fat retention with an increase in dietary fat level (P<.01), a larger proportion (P<.05) of unabsorbed fat was present as soap in the excreta of birds on these diets relative to those fed the control diet. There was evidence to indicate that most of the soap observed in the excreta of laying hens was formed in postabsorptive areas of the gut, thereby explaining why the detrimental effects of soap formation were not seen in the laying hen. Increasing the dietary calcium level reduced both percentage calcium (P<.01) and magnesium (P<.05) retention. There were no significant effects of the treatments on nitrogen and phosphorus retention. There were also no significant effects of the treatments on shell ash, shell calcium, magnesium, and phosphorus content. Increasing the dietary calcium level increased bone ash (P<.05) while reducing bone magnesium content (P<.05). There were no significant effects of the fat and calcium treatments on bone calcium and phosphorus content. (Key words- fat, calcium, layer performance, bone minerals)
17.1 3.06 .75 .13 1.63
Analyzed nutrient content Crude protein, % Calcium, % Total phosphorus, % Magnesium, % Crude fat, %
Calculated metabolizable energy for all diets, 2750 kcal/kg.
10.05
67.90 22.05
17.4 4.18 .77 .14 1.67
100.00
3
17.1 2.92 .81 .12 6.45
67.90 11.15 5.00 6.89 9.06 100.00
4
('«)
17.1 3.63 .76 .13 6.54
67.90 11.15 5.00 8.47 7.48 100.00
5
diets1
6
17 4
67 11 5 10 5 100
6
3
A-V = Animal-vegetable blend.
Contain corn, 37.75%; soybean meal (48%), 23.80%; corn gluten meal, 3.40%; calcium phosphate, 1.55%; DL-m .25%; chromic oxide, .30%; mineral-vitamin premix, .75% [provides per kilogram of diet: vitamin A, 8000 IU; vita mg; pantothenic acid, 7.0 mg; vitamin B 1 2 , 8 Mg; niacin, 20 mg; choline, 900 mg; vitamin K, 1.5 mg; folic acid, 1.5 mg; manganese, 50 mg; zinc, 50 mg; copper, 5.0 mg; iron 30.0 mg].
2
1
8.47 1.58 100.00
6.89 3.16 100.00
17.4 3.56 .79 .13 1.70
67.90 22.05
67.90 22.05
Basal ingredients 2 Corn starch A-V3 blend fat Limestone Alpha-floc (wood cellulose) Total
2
1
Treatment number
TABLE 1. Experimental
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2092
ATTEH AND LEESON
cedure (Kjel-Foss Automatic 16310), whereas gross energy was determined by adiabatic oxygen bomb calorimetry (Parr, Model 1241). Ash samples from feed, excreta, shell, and bone were digested using the method of Association of Official Analytical Chemists (AOAC, 1980). The resulting solutions were analyzed for calcium and magnesium using a Techtron (Model AA 4 ) atomic absorption spectrophotometer and phosphorus using a Technicon (Model AA 2 ) autoanalyzer. The presence of chromium in the feed and the excreta was determined using the method of Fenton and Fenton (1979). Total glycerides and fatty acids in feed, digesta, and excreta were determined by petroleum ether extraction using Soxhlet apparatus. To estimate the proportion of the fatty acids in the digesta and excreta that was present as soap, the method of fat determination reported by Carroll (1958) was used. The digesta and excreta were subjected to two stages of ether extraction. The first extraction was to remove neutral fat and fatty acids. The residue from the first extraction was subjected to acid treatment using 25% hydrochloric acid (specific gravity, 1.13) for 2 hr at room temperature to liberate fatty acids that were present as soap. The samples were then freeze dried and the process of ether extraction repeated. This second extraction was considered to represent fatty acids previously
TABLE 2. Effects of dietary levels of fat (F) and calcium (Ca) on performance of laying bens
Dietary treatments
Feed intake
Weight gain over 7-week period 1
Egg production (HDB) 2
Egg weight
Egg shell deformation
(g/hen/day)
(g/bird)
(%)
(g)
(um)
NS 3 109.7 108.6 109.3
NS 42.3 97.9 82.3
NS 90.6 94.6 90.9
NS 56.6 57.0 56.6
NS 25.6 25.6 25.2
Ca level, % 3.0 3.6 4.2
NS 113.5 108.7 105.5
NS 78.5 76.3 67.7
NS 91.0 93.1 92.0
NS 55.8 57.0 57.5
NS 26.2 25.6 24.6
Ca X F
NS
NS
NS
NS
NS
10.1
11.6
Supplemental F, 0 5.0 10.0
%
Standard deviation 1
12.9
Average for 7 weeks.
2
Hen-day basis.
3
NS = No significant difference (P>.05).
3.9
2.7
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for the disappearance in 24 hr of part of a 200 g feed allocation. Eggs were also collected twice a week for weighing and determination of shell deformation as a measure of shell quality using the method of Summers et al. (1976). Shells from the first egg collection each week were oven dried and ground prior to ashing at 600 C for 48 hr. Egg production was recorded throughout the trial. During the 4th week of the experiment, excreta samples were taken in order to estimate nutrient retention. Chromic oxide was used as a marker (Table 1) and excreta collection was undertaken over a 72-hr period. The excreta samples were dried in a forced air oven at 70 C and ground prior to ashing at 600 C overnight and other subsequent chemical analysis. At the conclusion of the feeding trial, four hens per treatment were killed by cervical dislocation within 30 min of oviposition and the body cavity was immediately opened. The contents of the small intestine (digesta) were then flushed out with double distilled water and freeze dried prior to grinding. Also, the left tibia of each bird was removed and cleaned of adhering flesh, dried at 100 C for 24 hr, defatted using a Soxhlet extraction apparatus, and dried a second time prior to ashing at 600 C overnight. Ash weight was taken to estimate percentage bone ash. Chemical Analysis. Nitrogen in both feed and excreta was determined by Kjeldahl pro-
3
2
1
3.4
2.9
NS 5.2
NS
NS 28.6 b 27.2 b 23.3 a
NS 27.6 26.0 25.5
**
*
4.1
NS 4.0
NS
83.5 82.4 82.1
77.6 a 83.8b 86.6 C
56.4 b 54.4 b 49.1 a
**
F
NS 54.9 53.0 52.0
Ca
25
NS
NS 2838 2789 2803
NS 2782 2796 2853
(kcal
ME
NS = No significant difference (P>.05)'
Excreta ether Extract 2 as a proportion of Extracts 1 plus 2.
Digesta ether Extract 2 as a proportion of Extracts 1 plus 2.
' ' Within main treatment categories, means within column followed by different superscripts are significantly di
Standard deviati on
NS
CaX F
NS 41.1 38.8 40.3
NS 38.7 40.8 40.8
NS 3 56.8 55.3 55.1
NS 55.2 55.3 56.7
%
Magnesium
Nutrient retention
Phosphorus :
Nitrogen
Ca level, % 3.0 3.6 4.2
Supplemental F, 0 5.0 10.0
Dietary treatments
TABLE 3. Effects of dietary levels of fat (F) and calcium (Ca) on nutrient retention by laying hens, dietary
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ATTEH AND LEESON
2094
present as soap. Samples of the feed were subjected to similar two-stage ether extractions to act as standards. Fats and fatty acids extracted from feed and that from each of the two extractions from the digesta and excreta were then esterified with .2 N methanolic trimethyl ammonium hydroxide, and the methyl esters were analyzed for component fatty acids by gas-liquid chromatography (Varian, Model 2100) using the method of AOAC (1980). Data collected were subjected to analysis of variance using Statistical Analysis System (SAS, 1982). Significant differences among treatments were determined by Duncan's new multiple range test (Duncan, 1955).
—< r-~ 0\ o* r~ r-H q o. >o H q ' N ^ i M 06 N
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RESULTS
Layer Performance. There was no significant effect of the fat and calcium treatments on feed intake, although there was a tendency for increases in dietary calcium level to reduce intake (Table 2). There were also no significant effects of the treatments on weight gain, egg production, egg weight, and egg shell quality (P>.05). Nutrient Retention. There was no significant effect of the fat or calcium levels on ME of the diets. Also, there was no significant effect of the treatments on nitrogen and phosphorus retention (Table 3). While increasing the dietary fat level had no significant effect on calcium and magnesium retention, increase in dietary calcium levels resulted in a decrease in percentage calcium (P<.01) and magnesium (P<.05) retention. Increases in dietary fat level resulted in an increase in fat retention (P<.01). However, a larger percentage of unabsorbed fat was present as soap with increases in dietary fat (P<.05). Although all fatty acids were involved in soap formation, palmitic and stearic acids were responsible for more than 50% of the soap formed during the process of digestion (Table 4). There was no significant interaction between dietary fat and calcium levels on the retention of any of the nutrients measured. Egg Shell Minerals. There were no significant effects of the treatments on shell ash, shell calcium, phosphorus, or magnesium, although increases in dietary calcium level tended to increase shell calcium and decrease shell magnesium content (Table 5). In all cases these effects were not statistically significant (P> .05). Bone Ash and Minerals. There was no significant effect of the dietary fat level on
i 0\ « m r j ir\ ' © ">_ t~; © rrt ' N
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6 S
NS 47.8 48.2 48.6
NS
Ca level, % 3.0 3.6 4.2
CaX F
2.6
NS
NS 37.5 37.9 39.2
NS 38.4 38.6 37.7
Ca
.001
NS
NS .08 .08 .08
NS .08 .08 .08
Phosphorus
Shell
.02
NS
NS .36 .36 .35
NS .37 .35 .36
Magnesium
1.8
NS
54.5 a 54.5 a 56.0 b
•
NS 55.4 54.4 54.5
Ash 2
N
N 3 3 3
N 3 3 3
C
Average for 7 consecutive weeks.
NS = No significant difference (P>.05).
Percentage of bone ash.
On dry, fat free basis.
*P<05.
4
3
2
1
ab ' Within main treatment categories, means within column followed by different superscripts are significantly diffe
1.8
NS" 48.6 48.2 47.9
Supplemental F, % 0 5.0 10.0
Standard deviation
Ash
Dietary treatments
TABLE 5. Effects of dietary levels of fat (F) and calcium (Ca) on egg shell and bone mineral co
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2096
ATTEH AND LEESON
bone ash. However, increasing the dietary calcium content significantly (P<.05) increased bone ash (Table 5). Increasing the dietary fat level had a tendency to decrease bone calcium content, whereas increasing the dietary calcium level had the opposite effect on bone calcium content. However, in both cases, the effects were not statistically significant (P>.05). There was no significant interaction between dietary fat and calcium levels on any of the bonerelated measurements.
The nonsignificant effect of the treatments on the performance variables measured was probably due to a comparable energy content of the diets. This showed that fat per se had no significant effect on laying hen performance, an observation which confirms earlier reports (Vogt and Harnisch, 1976; Horani and Sell, 1977) showing that dietary fat had no significant effect on these parameters. However, this is in contrast to the situation observed in chicks (Johri and Narayanan, 1972; Mateos and Sell, 1980; Atteh et al, 1983) showing improvement in weight gain and energetic efficiency of feed with fat supplementation. Performance of laying hens fed diets supplemented with A-V blend fat was comparable to those fed diets supplemented with free fatty acids (Atteh and Leeson, 1984b) with the notable exception of palmitic acid which laying hens could not utilize efficiently. The tendency for increases in dietary calcium level to reduce feed intake agrees with a previous report (Atteh and Leeson, 1984b) which showed that laying hens reduced intake when supplied diets high in calcium. Supplemental fat was added to the diets at the expense of corn starch to provide isocaloric diets. However, the ME content of diets supplemented with fat were slightly higher than in the control diet and this "extra caloric" effect, though higher with increase in fat supplementation, was not as profound as the situation observed in chicks (Cullen et al, 1962; Jensen et al, 1970; Atteh et al, 1983) or laying hens fed diets supplemented with free fatty acids with the exception of palmitic acid (Atteh and Leeson, 1984b). The improvement in fat retention with increase in dietary fat supplementation is probably associated with increase in proportion of oleic acid in the diet which has been shown to facilitate the absorption of other fatty
The changes in fatty acid profile of diet fat during digestion confirm earlier observations (Renner and Hill, 1961; Gardiner and Whitehead, 1976; Atteh and Leeson, 1983b, 1984) of inherent problems of absorption of saturated fatty acids. Thus, the proportion of palmitic and stearic acids increased from 11.94 and 2.31% in the control diet to 15.3 and 9.76% in the nonsoap portion of unabsorbed fat and to 35.89 and 22.82% of soap portion of unabsorbed fat, respectively. Similar trends for these two fatty acids were also evident in diets supplemented with 5 or 10% fat. Although all the component fatty acids in the fat were involved in soap formation, there is evidence to suggest that the soaps of myristic and oleic acids were absorbed to a large extent, because the proportion of fatty acids from soap fat accounted for by myristic and oleic acids was low compared to their proportion in the soap fat of the digesta. This observation supports the reports of Boyd et al. (1932) and Atteh and Leeson (1984) showing that soaps of oleic acid were well utilized. The significant decrease in magnesium retention and bone magnesium content with increase in dietary calcium content showed that high levels of calcium interfered with magnesium metabolism and this interference was independent of the dietary fat level. This agrees with the reports of Hakansson (1975b) and Atteh and Leeson (1983a, 1985). Although there was a decrease in the percentage calcium retention with the increase in dietary calcium level, in absolute terms, birds on diets with 4.2% calcium retained as much calcium as those on diets with lower levels of calcium. This would explain why the calcium treatment had no significant effect on shell and bone calcium content. The fact that the fat treatment had no
Downloaded from http://ps.oxfordjournals.org/ at Florida International University on June 21, 2015
DISCUSSION
acids (Young and Garett, 1963). There is evidence to suggest that most of the soap formed during digestion in laying hens occurred in postabsorptive areas of the gut. Thus, despite the fact that birds on diets supplemented with 10% fat had the highest fat retention, the proportion of unabsorbed fat that was present as soap was highest with these same birds. This indicates that most of the soap observed in the excreta was formed after most absorption had taken place. This observation confirms earlier findings (Atteh and Leeson, 1984b) and would explain why detrimental effects of soap formation were not seen in the laying hen.
FAT AND CALCIUM FOR LAYERS
REFERENCES Association of Official Analytical Chemists, 1980. Pages 21 and 447 in Official Methods of Analysis, 13th ed. Assoc. Offic. Anal. Chem., Washington, DC. Atteh, J. O., and S. Leeson, 1983a. Influence of increasing dietary calcium and magnesium levels on performance, mineral metabolism, and egg mineral content of laying hens. Poultry Sci. 62:1261-1268. Atteh, J. O., and S. Leeson, 1983b. Effects of dietary fatty acids and calcium levels on performance and mineral metabolism of broiler chickens. Poultry Sci. 62:2412-2419. Atteh, J. O., and S. Leeson, 1984. Effects of dietary saturated, unsaturated fatty acids, and calcium levels on performance and mineral metabolism of broiler chicks. Poultry Sci. 63:2252-2260. Atteh, J. O., and S. Leeson, 1985. Response of laying hens to dietary saturated and unsaturated fatty acids in the presence of varying dietary calcium levels. Poultry Sci. 64:520-528. Atteh, J. O., S. Leeson, and R. J. Julian, 1983. Effects of dietary levels and types of fat on performance and mineral metabolism of broiler chicks. Poultry Sci. 62:2403-2411. Boyd, S. F., C. L. Crum, and J. F. Lyman, 1932. The absorption of calcium soaps and the relation of dietary fat to calcium utilization in the white rat. J. Biol. Chem. 9 5 : 2 9 - 4 1 . Carew, L. B., R. H. Machemer, R. W. Sharp, and D. C. Foss, 1972. Fat absorption by the very young chick. Poultry Sci. 51:738-742. Caroll, K. K., 1958. Digestibility of individual fatty acids in the rat. J. Nutr. 64:399-410. Cullen, M. P., O. G. Rasmussen, and O.H.M. Wilder, 1962. Metabolizable energy value and utilization of different types and grades of fat by the chick. Poultry Sci. 41:360-367. Dewar, W. A., C. C. Whitehead, J. N. Downie, and E. Potter, 1975. The retention of calcium, iron, magnesium and zinc in chicks fed on diets
containing metal soaps. Proc. Nutr. Soc. 34: 5A-6A. Duckworth, J., J. M. Naftalin, and C. Dalgarno, 1950. The digestibility of linseed oil and mutton fat by chicks. J. Agric. Sci. 4 0 : 3 9 - 4 3 . Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics 11:1—42. Fenton, T. W., and M. Fenton, 1979. An improved procedure for determination of chromic oxide in feed and feces. Can. J. Anim. Sci. 59:631—634. Gardiner, E. E., and C. C. Whitehead, 1976. Effect of dietary palmitic acid on broiler chicks fed on various concentrations of calcium. Br. Poult. Sci. 17:241-244. Griffith, F. D., R. B. Grainger, and J. J. Begin, 1961. The effect of dietary fat and cellulose on apparent calcium digestibility in growing chickens. Poultry Sci. 40:1492-1497. Hakansson, J., 1975a. The effect of fat on the retention of calcium, phosphorus and magnesium in laying hens. Swed. J. Agric. Res., 5:3—9. Hakansson, J., 1975b. The effect of fat on calcium, phosphorus and magnesium balances in chicks. Swed. J. Agric. Res. 5:145-157. Horani, F., and J. L. Sell, 1977. Effect of feed grade animal fat on laying hen performance and on metabolizable energy of rations. Poultry Sci. 56:1972-1980. Jensen, L. S., G. W. Schumainer, and J. D. Latshaw, 1970. "Extra calorie" effect of dietary fat for developing turkeys as influenced by calorieprotein ratio. Poultry Sci. 49:1697-1704. Johri, T. S., and S. Narayanan, 1972. Effect of animal fats on growth of chicks. Indian J. Anim. Sci. 42:835-839. Mateos, G. G., and J. L. Sell, 1980. Influence of fat on energy utilization from selected carbohydrates. Poultry Sci. 59:1635. Polin, D., and T. H. Hussein, 1982. The effect of bile acid on lipid and nitrogen retention, carcass composition and dietary metabolizable energy in very young chicks. Poultry Sci. 61:1697—1707. Renner, R., and F. W. Hill, 1960. The utilization of corn oil, lard and tallow by chickens of various ages. Poultry Sci. 39:849-854. Renner, R., and F. W. Hill, 1961. Utilization of fatty acids by the chicken. J. Nutr. 75:259—264. Statistical Analysis System (SAS), 1982. Page 119 in SAS User's Guide: Statistics. SAS Inst. Inc., Cary, NC. Summers, J. D., R. Grandhi, and S. Leeson, 1976. Calcium and phosphorus requirements of the laying hen. Poultry Sci. 55:402-413. Vogt, H., and S. Harnisch, 1976. Comparison of different fats in complete feeds for laying hens. Arch. Gefluegelkd. 40:168-176. West, E. S., W. R. Todd, H. S. Mason, and J. T. Van Bruggen, 1966. Page 131 in Textbook of Biochemistry. 4th ed. MacMillan Co., London, England. Whitehead, C. C , and C. Fisher, 1975. The utilization of various fats by turkeys of different ages. Br. Poult. Sci. 16:481-485. Young, R. J., and R. L. Garrett, 1963. Effect of oleic and linoleic acids in the absorption of saturated fatty acids in the chick. J. Nutr. 81:321-329.
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significant effect on calcium and magnesium retention, shell and bone calcium, and magnesium content showed that up to 10% fat in the diet of laying hens is not detrimental to the metabolism of these minerals. This is in contrast to observations with chicks showing that addition of fat to their diets reduced not only the retention of calcium and magnesium through soap formation but also reduced the bone content of these minerals (Griffith et al, 1961; Dewar et al, 1975; Atteh et al, 1983). This variation between the young and adult bird in fat and mineral metabolism may not only be associated with lack of full physiological capacity for fat absorption by chicks (Carew et al, 1972) but also with a difference in the predominant site of soap formation in relation to area of fat absorption.
2097