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The Influence of Dietary Oils on Chick Growth Rate
EXTRUSION of hydration, gelatinization, and ball milling of starch on growth and energy utilization by the chick. Poultry Sci. 48: 1583-1589. Naber, E. C , and S. P. Touchburn, 1969b. Effect of water treatment on components of hard red wheat on growth and energy utilization by the chick. Poultry Sci. 48 : 2052-2057. Saunders, R. M., H. G. Walker, Jr. and G. O. Kohler, 1969. Aleurone cells and the digestibility of wheat mill feeds. Poulry Sci. 48: 14971503. Smith, 0. B., 1967. Pre-cooking of cereal and protein blends. Proc. International Symposium on Oilseed Proteins and Concentrates. CFTRI, Mysore, India.
261
PROCESSING
Steel, R. G. D., and J. H. Torrie, 1960. Principles and Procedures of Statistics. McGraw-Hill Publishing Co., New York. Totusek, R., and D. White, 1968. Methods of processing milo for cattle. Proc. 1968. Texas Nutr. Conf. pp. 50-58. Waldroup, P. W., C. L. White, S. J. Hull, D. E. Greene and E. L. Stephenson, 1969. Use of unextracted soybeans in poultry diets. Feedstuffs, Jan 4, p. 21. White, C. L., D. E. Greene, P. W. Waldroup and E. L. Stephenson, 1967. The use of unextracted soybeans for chicks, 1. Comparison of infra-red cooked, autoclaved and extruded soybeans. Poultry Sci. 46: 1180-1185.
The Influence of Dietary Oils on Chick Growth Rate HENRY
MENGE
United States Department of Agriculture1, Beltsville, Maryland 20705 (Received for publication August 10. 1970)
origin contain relatively O ILSsmallof marine amounts of linoleic and arachidonic acids, but, in contrast with vegetable and animal oils, they contain large quantities of the long-chain polyunsaturated fatty acids (PUFA) of the linolenic acid family. Menge et al. (1965) have reported data indicating that the PUFA2 of menhaden oil stimulated the reproductive characteristics of the hen and substituted, at least in part, for linoleic or arachidonic acid, or both. Edwards et al. (1962) have shown that menhaden oil has a stimulatory effect on chick growth. Later, Edwards et al. (1963) presented evidence to indicate that the PUFA of menhaden oil inhibit the synthesis of eicosatrienoic acid from oleic acid. This premise was substantiated by Menge et al. (1965). The present study was designed to compare the effect of safflower, 1 Animal Science Research Division, A.R.S., Beltsville, Md. 20705 2 The term PUFA in this paper includes linolenic, arachidonic together with the other 20-, 22-, and 24-carbon fatty acids or 35.9% of the total fatty acids of menhaden oil.
menhaden, growth.
and coconut oil on chick
EXPERIMENTAL
PROCEDURE
In Experiment 1, 8 duplicated groups of 10 chicks each (Leghorn males and females) were selected at random at one day of age and placed into an electric battery brooder. The groups received the linoleic (18:2) acid deficient basal diet (containing 0.43% 18:2) plus varying quantities of safflower, menhaden and coconut oil for 4 weeks as follows: Group 1, 18:2-deficient basal (see Diet 1, Table 1); Group 2, 0.15% safflower oil; Group 3, 0.27% safflower oil; Group 4, 2.07% safflower oil; Group 5, 4.19% safflower oil; Group 6, 5.19% menhaden oil; Group 7, 6.6% coconut oil; Group 8, 11.88% coconut oil. In experiment 2, one hundred fifty Leghorn day-old male chicks were selected at random and placed in an electric battery brooder according to the following arrangement for each of five treatments: Two pens of 7 chicks each, and 2 other pens of 8 chicks each, making a total of 30 chicks per
262
H. MENGE
treatment. Each group of 30 chicks received the following dietary treatments for a 4-week experimental period: Group 1, basal diet containing only 0.0018% 18:2 (see Diet 2, Table 1); Group 2, 2.58% safflower oil; Group 3, 5.15% safflower oil; Group 4, 2.58% menhaden oil; Group 5. 5.15% menhaden oil. The 4-week data from both experiments were analyzed using least squares proceTABLE 1.—Composition of diets Ingredients
1
Isolated soy protein Casein, vitamin test Gelatin Powdered cellulose2 Hydrogenated coconut oil3 Methionine hydroxy analogue Glycine Choline (70%) Antioxidant4 Vitamin Mix5 Mineral Mix6 Glucose monohydrate Metabolizable energy (Kcal./lOO gm.) Linoleic acid (%) 1
7
Diet 1
Diet 2
%
%
18.00 5.00 5.00 2.31 1.00 0.32
24.50
— —
0.35 0.02 0.50 7.50 60.00
5.43 1.00 0.50 0.20 0.35 0.02 0.50 7.50 60.00
308.00
297.00
—
0.43
0.0018
Assay protein C-l, Skidmore Enterprises, Cincinnati. This material was blended with powdered cellulose and extracted exhaustively with hot methanol in Diet 2 only. 2 Solka Floe, BW-40, Brown Company, Berlin, New Hampshire. 3 Supplied by the Procter and Gamble Company, Cincinnati. 4 l,2-dihydro-6-ethoxy-2,2,4-trimethylquinolone, Monsanto Chemical Company, St. Louis. 5 Vitamin mix supplied by the following (mg./kg. diet): thiamine-HCl, 100; riboflavin, 16; Ca-dpantothenate, 25; pyridoxine-HCl, 10; folacin, 4; Klotogen F, 15; niacin, 100; biotin, 0.6; vitamin B12 (0.1%), 20; vitamin A (500,000 U.S.P./gm.), 36; vitamin D 3 (200,000 I.C.U./gm.), 7; d-a-tocopheryl acetate (13601.U./gm.), 26; glucose monohydrate to 0.5% of diet. 6 Mineral mix supplied the following (in % ) : CaC0 3 , 2; CaHPCvH^O, 1.6; KH2PO4, 1.4; NaCl, 0.164; MgC0 3 , 1.75; FeC 6 H 6 07-5H 2 0, 0.06; MnS0 4 • H 2 0,0.03; ZnCOs, 0.01; KI, 0.0003; Cu(CH3COO)2 •H 2 0, 0.0032; H 3 B0 3 , 0.001; C o S O ^ H j O , 0.0002; N a s M o O ^ H j O , 0.0009; Na 2 SeCv 10 H 2 0,0.00005; glucose monohydrate to 7.5%. ' Analysis by KOH hydrolysis and gas-liquid chromatography.
TABLE 2.—Fatty acid composition of oils1 Fatty Acid2
12:0 14:0 16:0 16:1 16:2 18:0 18:1 18:2w6 18:3w3 18:4o>3 20:0 20:4»6 20:5o>3 22:0 22:1 22:4 22:5o>3 22:6w3 24:1
Safflower
Menhaden 3
Coconut
% —
% —
%
0.14 6.56
11.9 20.0 11.5 1.2 3.6 12.7 2-8 3.4 3.5 0.4 2.2 12.1 0.8 1.8 0.8 2.4 7.1 1.4
— —
2.62 12.87 77.63 0.20 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
40.7 20.9 11.0
— —
2.1 5.9 1.8 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
1 Fatty acid composition (percentage of total) as determined by gas-liquid chromatography. 2 The position of the first double bond counting from the terminal methyl group is indicated by the number following the « (omega) sign. This system designates the relationship of the fatty acids to each other. 3 Fatty acid composition determined in the laboratory of Hayme Products, Inc. Balto.
dures as described by Harvey (1960). The multiple range test was applied to the least squares means (Duncan, 1955). The diets used in both experiments were isocaloric and isonitrogenous. The fatty acid composition of the oils used in both experiments is given in Table 2. Duplicate samples of plasma lipids from both male and female chicks in Experiment 1 were analyzed for their fatty acid content. Since the values for both sexes were uniform, the average was obtained (Table 5). Method for determination of plasma lipid fatty acids from chicks in Experiment 1 was described in a previous paper (Miller et al., 1963). RESULTS AND DISCUSSION
Chick growth responses to safflower oil (S-O), menhaden oil (M-O) and coconut oil (C-O) in Experiment 1 are presented in Table 3. These oils were added to the basal
263
EFFECT OF OILS ON CHICK GROWTH TABLE 3.—Chick growth response to safflmeer, menhaden, or coconut oil in Experiment l1 Group
Treatment
1
Linoleic acid (18:2) deficient diet (0.43% 18:2)* 0.15% safflower oil (0.55% 18:2) 0.27% safflower oil (0.64% 18:2) 2.07% safflower oil (1.65% 18:2) 4.19% safflower oil (3.3% 18:2) 4.19% menhaden oil (0.55% 18:2; 1.5%PUFA 4 6.6% coconut oil (0.55% 18:2) 11.88% coconut oil (0.64% 18:2)
2 3 4 5 6 7 8
gm. 258»b 260«b 267b« 294d 287cd 310d 246"b 235"
1 Male and female chicks. Twenty chicks per group. 2 Means with different superscript are significantly different at the 5% level according to Duncan's multiple range test (Duncan, 1955). 3 Diet calculated to contain 430 mg. 18:2/100 gm. (see Table 1). The diets used in groups 2 through 8 were, therefore enriched by 0.43% dietary 18:2 which has been included in the 18:2 noted above for each of these groups. 4 Linolenic, arachidonic and other 20-, 22-, and 24-carbon fatty acids (35.9% of total fatty acids).
diet on the basis of their 18:2 content. The growth responses exhibited by the groups receiving C-0 (groups 7 and 8) were comparable with those obtained from the groups given an equivalent level of 18:2 from S-0 (groups 2 and 3). Group 8, which received the highest level of C-O, gave a response that was indicative of a growth depression even though the differences were not significant. This lack of response was not due to 18:2 per se, since 18:2 intake, based on actual feed consumption (Table 4) demonstrated that Group 8 received more 18:2 per chick than Group 1 (basal) and Group 2 (S-O). No research has been reported on the growth stimulation of fat in the diet of the chick using C-O. Menhaden oil (Group 6) was equally as effective in the promotion of chick growth as was S-0 in Groups 4 and S even though there was a wide variation in the amount of dietary 18:2 (Tables 3 and 4). According to Mohrhauer and Holman (1963) 18:2 is essential only as a precursor of the biologically active arachidonate (20:4). These
findings lend support to the premise that the PUFA of M-0 used in the present study substituted, at least in part, for 18:2 or 20:4, or both in the diet of the chick. This theory was first postulated in a previous paper (Menge et al., 1965). A fatty analysis of the plasma lipids from the chicks in Experiment 1 is shown in Table 5. The higher levels of lauric (12.0) and myristic (14:0) acids are the only indication in the fatty acid profiles as a possible reason for the poor growth observed in the group receiving the highest level of C-O (Group 8, Table 3). It is also worthy to note that even though the differences were not significant, the poor growth obtained with Group 8 occurred in the presence of relatively high levels of 18:2 and 20:4 in the plasma tissue lipids. The fatty acid profile of plasma lipids from group 1 (basal) and groups 2, 3 and 4 (increasing increments of S-O) is indicative of an essential fatty acid deficiency together with a subsequent repletion (Table 5). These data also demonstrate that as tissue 18:2 is decreased, there is a concomitant increase in tissue eicosatrienoic (20:3) ex-
TABLE 4.—Linoleic and polyunsaturated fatly acid intake per chick in Experiment l1 Group
Treatment
Linoleate2
PUFA 3
gm.
gm.
Linoleic acid deficient 0.15% safflower oil 0.27% safflower oil 2.07% safflower oil 4.19% safflower oil 4.19% menhaden oil 6.6% coconut oil 11.88% coconut oil
1.99 2.53 2.95 8.21 16.65 2.93 2.60 2.73
0.001 4 0.002 4 0.02 4 0.04 4 8.00
— —
1 Male and female chicks. Twenty chicks per group. 2 Calculated from amount of oil consumed per chick. Basal diet contained 430 mg. 18:2/100 gm. (see Table 1). 3 Polyunsaturated fatty acids from oil consumed per chick. 4 Calculated from 0.2% linolenic acid in safflower oil.
264
H. MENGE TABLE 5.-—Effect
of •safflower, menhaden, and coconut oil on the fatty acid composition of plasma lipids of male and female chicks in Experiment I1
3
12:0 14:0 16:0 16:1 18:0 18:1 18:2 18:3 20:3 20:4 20:5w3 22: 5a>3
1 Low fat
2 0.5% S-O3
3 0.27% S-0
4 2.07% S-0
5 4.19% S-0
6 4.19% M-O3
7 7.66%, C-O3
8 11.88% CO
%
%
%
%
A)
%
%
/o
+
0.4 0.3 20.0 0.7 25.7 11.3 21.7
+
0.6 1.7 18.7 5.1 16.1 38.9 5.8 0.2 7.2 2.4 0.0 0.0
1.7 3.7 18.7 5.3 18.6 30.7 10.0
+
0.4 16.8 5.0 14.6 53.9 1.0 0.5 6.0 1.2 0.0 0.0 \J . U
+
0.4 16.0 4.4 15.9 44.5 5.5 0.4 8.2 2.8 0.0 0.0 0.0
+
0.4 19.4 3.4 16.7 37.7 9.4 0.1 5.4 5.3 0.0 0.0 0.0
0.4 19.2 1.5 22.4 16.7 16.7
+
1.3 21.1 0.0 0.0 0.0
+ +
18.3 0.0 0.0 0.0
1.1 31.0 2.1 19.9 15.8 1.1 0.5
+
4.2 11.2 3.0 9.2
o.o
+
4.6 4.3 0.0 0.0 0.0
Average of males and females (value for each sex very uniform) expressed as percentage of total fatty acids. 4- sign denotes trace amounts less than sensitivity of instrument. 1 S-O: safflower oil; M-O: menhaden oil: C-O: coconut oil.
cept when PUFA from M-0 are present in the diet (Group 6, Table 5). The data in Table 6 present a comparison between the chick growth stimulation obtained from 18:2 in S-0 and the PUFA from M-0 in Experiment 2. Each of the oils prompted a significant chick growth response when used to supplement the 18:2 TABLE 6.—Chick growth response to safflower or menhaden oil in Experiment 21 Group
Body weights2
Treatment
gm. 1 2 3 4 5 1
3
Linoleic acid (18:2) deficient diet 2.58% safflower oil (2% 18:2) 5.15% safflower oil (4% 18:2) 2.58% menhaden oil (0.07% 18:2; 0.93% PUFA") 5.15% menhaden oil (0.14% 18:2; 1.85% PUFA 4 )
266" 299b 314b° 303 bc 317°
Male chicks, 30/group for 4-week experimental period. 2 Means with different superscripts are significantly different at the 5% level according to Duncan's multiple range test (Duncan, 1955). 3 Linoleic acid (18:2) deficient diet calculated to contain 1.8 mg. 18:2 per 100 gms. (see Table 1). 4 Linolenic, arachidonic and other 20-, 22-, and 24-carbon fatty acids (35.9% of total fatty acids).
-deficient basal diet (Group 1). The PUFA of M-0 (Groups 4 and 5), however, augmented the relatively low content of 18:2 (Table 7) to effect a chick growth response that equalled the growth promotion obtained from the much greater levels of 18:2 supplied by the S-O. These data indicate that the PUFA present in M-0 are accountable for the chick growth response. An essential fatty acid (EFA) deficiency is characterized by a decrease in the content of arachidonate (20:4) and an increase in the content of eicosatrienoic (20: 3) in the tissues of the animal (Holman, 1960). Decreased growth rate of EFA-deficient chicks as demonstrated in Experiment 1, and also reported by Edwards et al. (1963), and adverse effects on the reproductive functions of the EFA-deficient hen (Menge et al., 1965) have been correlated with increased 20:3 content of the tissues. Edwards et al. (1963) have observed that the PUFA of M-0 inhibit the synthesis of 20:3 from oleic acid (18:1). This inhibition of 20:3 synthesis by the PUFA has also been observed by Menge et al. (1965) and has
EFFECT OF OILS ON CHICK GROWTH TABLE 7.—Linoleic and polyunsaturated fatty acid intake per chick in Experiment 2l Group
Treatment
ariH*
1 2 3 4 5
Linoleic acid deficient 2.58% saflower oil 5.15% safflower oil 2.58% menhaden oil 5.15% menhaden oil
gm. 0.009 11.51 23.69 0.39 0.78
PUFA 3 gm. — 0.03" 0.06 4 5.12 10.25
1
Male chicks, 30 per group for 4-week experimental period. 2 Calculated from amount of oil consumed per chick. Basal diet contained 1.8 mg. 18:2/100 gm. (see T a b l e t ) . 3 Polyunsaturated fatty acids from oil consumed per chick. 4 Calculated from 0.2% linolenic acid in safflower oil.
been shown to result in increased reproductive activity in the hen, and as noted in Experiment 1, increased chick growth. Thus, the PUFA in the present studies can be assumed to have stimulated chick growth through the operation of at least two factors: (1) The depression of 20:3 synthesis from 18:1, and (2) The PUFA may have substituted, at least in part, for 18:2 or 20: 4, or both. Table 7 gives a resume of the unsaturated fatty acid intake during the 4-week experimental period in Experiment 2. The wide differences between the consumption of 18:2 and PUFA is, of course, a reflection of the unsaturated fatty acid content of S-0 and M-O. The amount of PUFA (M-O) consumed per chick in Group 5 (10.25 gm) was as effective in growth promotion as the nearly 24 gm 18:2 (S-O) consumed per chick in group 3 (Tables 6 and 7). The basal diet used in Experiment 1 contained unextracted soy protein and cellulose (Table 1), and as expected, a quantity of 18:2 (0.43%). The 18:2 inherent to the diet is apparent in the figures shown for the consumption of 18:2 by the different groups (Table 4). This dietary difference between Experiment 1 and Ex-
265
periment 2 is noted especially in a comparison between the 18:2 consumption of Group 6 in Experiment 1 (Table 4) and Groups 4 and 5 in Experiment 2 (Table 7). The growth stimulation from the PUFA of M-O in Experiment 1 (Table 3) is still apparent, however. SUMMARY
A comparison was made between the chick growth stimulation of the linoleate from safflower oil or coconut oil and the polyunsaturated fatty acids of menhaden oil. Significant growth responses were obtained from 1.65-3.3% linoleate supplied by safflower oil. No significant differences existed between the groups receiving relatively small amounts of linoleate from safflower or coconut oil, although dietary coconut oil at the 11.88% level did exhibit a probable growth depression. The high levels of saturated fatty acids in the coconut oil may, at least, be a contributing factor in this case. Relatively high levels of these fatty acids were found in the plasma lipids of the chicks fed coconut oil. Menhaden oil containing relatively low amounts of linoleate promoted chick growth as effectively as safflower oil that supplied much greater levels of linoleate. The responses obtained from menhaden oil were considered to be due to the polyunsaturated fatty acids of the oil which have been demonstrated to depress the synthesis of eicosatrienoic acid, and also they may have substituted for linoleate or arachidonate, or both, in the essential fatty acid-deficient chick. REFERENCES Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics, 11: 1-42. Edwards, H. M., Jr., J. E. Marion and J. C. Daggers, 1962. Studies on fat and fatty acid requirement of poultry. Proc. X l l t h World Poultry Congress, p. 182-186.
266
H. MENGE
Edwards, H. M., Jr., and J. E. Marion, 1963. Influence of dietary menhaden oil on growth rate and tissue fatty acids of the chick. J. Nutrition, 8 1 : 123-130. Havey, W. R., 1960. Least squares analysis of data with unequal numbers. U.S.D.A., A.R.S. 20-28. Holman, R. T., 1960. The ratio of trienoic-tetraenoic acids in tissue lipids as a measure of essential fatty acid requirement. J. Nutrition, 70: 405-410.
Menge, H., C. C. Calvert and C. A. Denton, 1965. Influence of dietary oils on reproduction in the hen. J. Nutrition, 87 : 365-370. Miller, E. C , H. Menge and C. A. Denton, 1963. Effect on long-term feeding of a fat-free diet to laying hens. J. Nutrition, 80: 431-440. Mohrhauer, H., and R. T. Holman, 1963. The effect of dose level of essential fatty acids upon fatty acid composition of the rat liver. J. Lipid Research, 4 : 151-159.
Motor Unit Recruitment Pattern in a Respiratory Muscle of Unanesthetized Chickens1 R. R. TSCHORN AND M. R. FEDDE Neuromuscular Laboratory, Department of Physiological Sciences, Kansas State University, Manhattan, Kansas 66502 (Received for publication August 10, 1970)
T
HE amplitude of electrical activity of respiratory muscles of the chicken, as recorded with a needle electrode, seems to be related to the level of anesthesia. Fedde et al. (1964) have shown that motor unit potentials of large amplitude are absent when chickens are deeply anesthetized. Since many investigations of the pattern of motor unit recruitment have used anesthetized animals (Sears, 1964; Fedde et al., 1969), we attempted to determine the motor unit recruitment pattern in unanesthetized chickens. METHODS A biopolar, electromyographic electrode (Fig. 1) was implanted in the transversus abdominis muscle (TA) in six, lightly anesthetized (20 mg./kg. of body weight of sodium pentobarbital administered intravenously) Single Comb White Leghorn males, 1 Contribution No. 68, Department of Physiological Sciences, College of Veterinary Medicine, KSAES, K.S.U., Manhattan, Kansas 66502. Supported by Public Health Service Research Grant NB-05786 from the National Institute of Neurological Disease and Stroke.
12 to 16 weeks old. The skin was incised along the ventro-lateral surface of the abdominal wall; then the fibers of the external abdominal oblique and rectus abdominis muscles were separated by blunt dissection to expose the lateral surface of the TA. The electrode was placed between the rectus abdominis and the TA muscles so that the wires were imbedded in the belly of the TA. The overlying muscles and skin then were sutured separately. The electrode cable was secured to the skin of the abdominal wall and back so that the chicken could not dislodge the electrode from its internal position. To be certain that the electrode was placed properly, electrical activity was monitored before the bird had recovered from the anesthetic by connecting it to a differential amplifier (Model PS, Grass Instrument Co., Quincy, Mass.), the output of which was inserted in one channel of a multichannel oscilloscope (Model 565, Tektronix, Inc., Beaverton, Oregon). A strain gauge transducer was constructed to record respiratory movements in unrestrained chickens by filling twelve centimeters of flexible rubber tubing, 5
261
PROCESSING
Steel, R. G. D., and J. H. Torrie, 1960. Principles and Procedures of Statistics. McGraw-Hill Publishing Co., New York. Totusek, R., and D. White, 1968. Methods of processing milo for cattle. Proc. 1968. Texas Nutr. Conf. pp. 50-58. Waldroup, P. W., C. L. White, S. J. Hull, D. E. Greene and E. L. Stephenson, 1969. Use of unextracted soybeans in poultry diets. Feedstuffs, Jan 4, p. 21. White, C. L., D. E. Greene, P. W. Waldroup and E. L. Stephenson, 1967. The use of unextracted soybeans for chicks, 1. Comparison of infra-red cooked, autoclaved and extruded soybeans. Poultry Sci. 46: 1180-1185.
The Influence of Dietary Oils on Chick Growth Rate HENRY
MENGE
United States Department of Agriculture1, Beltsville, Maryland 20705 (Received for publication August 10. 1970)
origin contain relatively O ILSsmallof marine amounts of linoleic and arachidonic acids, but, in contrast with vegetable and animal oils, they contain large quantities of the long-chain polyunsaturated fatty acids (PUFA) of the linolenic acid family. Menge et al. (1965) have reported data indicating that the PUFA2 of menhaden oil stimulated the reproductive characteristics of the hen and substituted, at least in part, for linoleic or arachidonic acid, or both. Edwards et al. (1962) have shown that menhaden oil has a stimulatory effect on chick growth. Later, Edwards et al. (1963) presented evidence to indicate that the PUFA of menhaden oil inhibit the synthesis of eicosatrienoic acid from oleic acid. This premise was substantiated by Menge et al. (1965). The present study was designed to compare the effect of safflower, 1 Animal Science Research Division, A.R.S., Beltsville, Md. 20705 2 The term PUFA in this paper includes linolenic, arachidonic together with the other 20-, 22-, and 24-carbon fatty acids or 35.9% of the total fatty acids of menhaden oil.
menhaden, growth.
and coconut oil on chick
EXPERIMENTAL
PROCEDURE
In Experiment 1, 8 duplicated groups of 10 chicks each (Leghorn males and females) were selected at random at one day of age and placed into an electric battery brooder. The groups received the linoleic (18:2) acid deficient basal diet (containing 0.43% 18:2) plus varying quantities of safflower, menhaden and coconut oil for 4 weeks as follows: Group 1, 18:2-deficient basal (see Diet 1, Table 1); Group 2, 0.15% safflower oil; Group 3, 0.27% safflower oil; Group 4, 2.07% safflower oil; Group 5, 4.19% safflower oil; Group 6, 5.19% menhaden oil; Group 7, 6.6% coconut oil; Group 8, 11.88% coconut oil. In experiment 2, one hundred fifty Leghorn day-old male chicks were selected at random and placed in an electric battery brooder according to the following arrangement for each of five treatments: Two pens of 7 chicks each, and 2 other pens of 8 chicks each, making a total of 30 chicks per
262
H. MENGE
treatment. Each group of 30 chicks received the following dietary treatments for a 4-week experimental period: Group 1, basal diet containing only 0.0018% 18:2 (see Diet 2, Table 1); Group 2, 2.58% safflower oil; Group 3, 5.15% safflower oil; Group 4, 2.58% menhaden oil; Group 5. 5.15% menhaden oil. The 4-week data from both experiments were analyzed using least squares proceTABLE 1.—Composition of diets Ingredients
1
Isolated soy protein Casein, vitamin test Gelatin Powdered cellulose2 Hydrogenated coconut oil3 Methionine hydroxy analogue Glycine Choline (70%) Antioxidant4 Vitamin Mix5 Mineral Mix6 Glucose monohydrate Metabolizable energy (Kcal./lOO gm.) Linoleic acid (%) 1
7
Diet 1
Diet 2
%
%
18.00 5.00 5.00 2.31 1.00 0.32
24.50
— —
0.35 0.02 0.50 7.50 60.00
5.43 1.00 0.50 0.20 0.35 0.02 0.50 7.50 60.00
308.00
297.00
—
0.43
0.0018
Assay protein C-l, Skidmore Enterprises, Cincinnati. This material was blended with powdered cellulose and extracted exhaustively with hot methanol in Diet 2 only. 2 Solka Floe, BW-40, Brown Company, Berlin, New Hampshire. 3 Supplied by the Procter and Gamble Company, Cincinnati. 4 l,2-dihydro-6-ethoxy-2,2,4-trimethylquinolone, Monsanto Chemical Company, St. Louis. 5 Vitamin mix supplied by the following (mg./kg. diet): thiamine-HCl, 100; riboflavin, 16; Ca-dpantothenate, 25; pyridoxine-HCl, 10; folacin, 4; Klotogen F, 15; niacin, 100; biotin, 0.6; vitamin B12 (0.1%), 20; vitamin A (500,000 U.S.P./gm.), 36; vitamin D 3 (200,000 I.C.U./gm.), 7; d-a-tocopheryl acetate (13601.U./gm.), 26; glucose monohydrate to 0.5% of diet. 6 Mineral mix supplied the following (in % ) : CaC0 3 , 2; CaHPCvH^O, 1.6; KH2PO4, 1.4; NaCl, 0.164; MgC0 3 , 1.75; FeC 6 H 6 07-5H 2 0, 0.06; MnS0 4 • H 2 0,0.03; ZnCOs, 0.01; KI, 0.0003; Cu(CH3COO)2 •H 2 0, 0.0032; H 3 B0 3 , 0.001; C o S O ^ H j O , 0.0002; N a s M o O ^ H j O , 0.0009; Na 2 SeCv 10 H 2 0,0.00005; glucose monohydrate to 7.5%. ' Analysis by KOH hydrolysis and gas-liquid chromatography.
TABLE 2.—Fatty acid composition of oils1 Fatty Acid2
12:0 14:0 16:0 16:1 16:2 18:0 18:1 18:2w6 18:3w3 18:4o>3 20:0 20:4»6 20:5o>3 22:0 22:1 22:4 22:5o>3 22:6w3 24:1
Safflower
Menhaden 3
Coconut
% —
% —
%
0.14 6.56
11.9 20.0 11.5 1.2 3.6 12.7 2-8 3.4 3.5 0.4 2.2 12.1 0.8 1.8 0.8 2.4 7.1 1.4
— —
2.62 12.87 77.63 0.20 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
40.7 20.9 11.0
— —
2.1 5.9 1.8 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
1 Fatty acid composition (percentage of total) as determined by gas-liquid chromatography. 2 The position of the first double bond counting from the terminal methyl group is indicated by the number following the « (omega) sign. This system designates the relationship of the fatty acids to each other. 3 Fatty acid composition determined in the laboratory of Hayme Products, Inc. Balto.
dures as described by Harvey (1960). The multiple range test was applied to the least squares means (Duncan, 1955). The diets used in both experiments were isocaloric and isonitrogenous. The fatty acid composition of the oils used in both experiments is given in Table 2. Duplicate samples of plasma lipids from both male and female chicks in Experiment 1 were analyzed for their fatty acid content. Since the values for both sexes were uniform, the average was obtained (Table 5). Method for determination of plasma lipid fatty acids from chicks in Experiment 1 was described in a previous paper (Miller et al., 1963). RESULTS AND DISCUSSION
Chick growth responses to safflower oil (S-O), menhaden oil (M-O) and coconut oil (C-O) in Experiment 1 are presented in Table 3. These oils were added to the basal
263
EFFECT OF OILS ON CHICK GROWTH TABLE 3.—Chick growth response to safflmeer, menhaden, or coconut oil in Experiment l1 Group
Treatment
1
Linoleic acid (18:2) deficient diet (0.43% 18:2)* 0.15% safflower oil (0.55% 18:2) 0.27% safflower oil (0.64% 18:2) 2.07% safflower oil (1.65% 18:2) 4.19% safflower oil (3.3% 18:2) 4.19% menhaden oil (0.55% 18:2; 1.5%PUFA 4 6.6% coconut oil (0.55% 18:2) 11.88% coconut oil (0.64% 18:2)
2 3 4 5 6 7 8
gm. 258»b 260«b 267b« 294d 287cd 310d 246"b 235"
1 Male and female chicks. Twenty chicks per group. 2 Means with different superscript are significantly different at the 5% level according to Duncan's multiple range test (Duncan, 1955). 3 Diet calculated to contain 430 mg. 18:2/100 gm. (see Table 1). The diets used in groups 2 through 8 were, therefore enriched by 0.43% dietary 18:2 which has been included in the 18:2 noted above for each of these groups. 4 Linolenic, arachidonic and other 20-, 22-, and 24-carbon fatty acids (35.9% of total fatty acids).
diet on the basis of their 18:2 content. The growth responses exhibited by the groups receiving C-0 (groups 7 and 8) were comparable with those obtained from the groups given an equivalent level of 18:2 from S-0 (groups 2 and 3). Group 8, which received the highest level of C-O, gave a response that was indicative of a growth depression even though the differences were not significant. This lack of response was not due to 18:2 per se, since 18:2 intake, based on actual feed consumption (Table 4) demonstrated that Group 8 received more 18:2 per chick than Group 1 (basal) and Group 2 (S-O). No research has been reported on the growth stimulation of fat in the diet of the chick using C-O. Menhaden oil (Group 6) was equally as effective in the promotion of chick growth as was S-0 in Groups 4 and S even though there was a wide variation in the amount of dietary 18:2 (Tables 3 and 4). According to Mohrhauer and Holman (1963) 18:2 is essential only as a precursor of the biologically active arachidonate (20:4). These
findings lend support to the premise that the PUFA of M-0 used in the present study substituted, at least in part, for 18:2 or 20:4, or both in the diet of the chick. This theory was first postulated in a previous paper (Menge et al., 1965). A fatty analysis of the plasma lipids from the chicks in Experiment 1 is shown in Table 5. The higher levels of lauric (12.0) and myristic (14:0) acids are the only indication in the fatty acid profiles as a possible reason for the poor growth observed in the group receiving the highest level of C-O (Group 8, Table 3). It is also worthy to note that even though the differences were not significant, the poor growth obtained with Group 8 occurred in the presence of relatively high levels of 18:2 and 20:4 in the plasma tissue lipids. The fatty acid profile of plasma lipids from group 1 (basal) and groups 2, 3 and 4 (increasing increments of S-O) is indicative of an essential fatty acid deficiency together with a subsequent repletion (Table 5). These data also demonstrate that as tissue 18:2 is decreased, there is a concomitant increase in tissue eicosatrienoic (20:3) ex-
TABLE 4.—Linoleic and polyunsaturated fatly acid intake per chick in Experiment l1 Group
Treatment
Linoleate2
PUFA 3
gm.
gm.
Linoleic acid deficient 0.15% safflower oil 0.27% safflower oil 2.07% safflower oil 4.19% safflower oil 4.19% menhaden oil 6.6% coconut oil 11.88% coconut oil
1.99 2.53 2.95 8.21 16.65 2.93 2.60 2.73
0.001 4 0.002 4 0.02 4 0.04 4 8.00
— —
1 Male and female chicks. Twenty chicks per group. 2 Calculated from amount of oil consumed per chick. Basal diet contained 430 mg. 18:2/100 gm. (see Table 1). 3 Polyunsaturated fatty acids from oil consumed per chick. 4 Calculated from 0.2% linolenic acid in safflower oil.
264
H. MENGE TABLE 5.-—Effect
of •safflower, menhaden, and coconut oil on the fatty acid composition of plasma lipids of male and female chicks in Experiment I1
3
12:0 14:0 16:0 16:1 18:0 18:1 18:2 18:3 20:3 20:4 20:5w3 22: 5a>3
1 Low fat
2 0.5% S-O3
3 0.27% S-0
4 2.07% S-0
5 4.19% S-0
6 4.19% M-O3
7 7.66%, C-O3
8 11.88% CO
%
%
%
%
A)
%
%
/o
+
0.4 0.3 20.0 0.7 25.7 11.3 21.7
+
0.6 1.7 18.7 5.1 16.1 38.9 5.8 0.2 7.2 2.4 0.0 0.0
1.7 3.7 18.7 5.3 18.6 30.7 10.0
+
0.4 16.8 5.0 14.6 53.9 1.0 0.5 6.0 1.2 0.0 0.0 \J . U
+
0.4 16.0 4.4 15.9 44.5 5.5 0.4 8.2 2.8 0.0 0.0 0.0
+
0.4 19.4 3.4 16.7 37.7 9.4 0.1 5.4 5.3 0.0 0.0 0.0
0.4 19.2 1.5 22.4 16.7 16.7
+
1.3 21.1 0.0 0.0 0.0
+ +
18.3 0.0 0.0 0.0
1.1 31.0 2.1 19.9 15.8 1.1 0.5
+
4.2 11.2 3.0 9.2
o.o
+
4.6 4.3 0.0 0.0 0.0
Average of males and females (value for each sex very uniform) expressed as percentage of total fatty acids. 4- sign denotes trace amounts less than sensitivity of instrument. 1 S-O: safflower oil; M-O: menhaden oil: C-O: coconut oil.
cept when PUFA from M-0 are present in the diet (Group 6, Table 5). The data in Table 6 present a comparison between the chick growth stimulation obtained from 18:2 in S-0 and the PUFA from M-0 in Experiment 2. Each of the oils prompted a significant chick growth response when used to supplement the 18:2 TABLE 6.—Chick growth response to safflower or menhaden oil in Experiment 21 Group
Body weights2
Treatment
gm. 1 2 3 4 5 1
3
Linoleic acid (18:2) deficient diet 2.58% safflower oil (2% 18:2) 5.15% safflower oil (4% 18:2) 2.58% menhaden oil (0.07% 18:2; 0.93% PUFA") 5.15% menhaden oil (0.14% 18:2; 1.85% PUFA 4 )
266" 299b 314b° 303 bc 317°
Male chicks, 30/group for 4-week experimental period. 2 Means with different superscripts are significantly different at the 5% level according to Duncan's multiple range test (Duncan, 1955). 3 Linoleic acid (18:2) deficient diet calculated to contain 1.8 mg. 18:2 per 100 gms. (see Table 1). 4 Linolenic, arachidonic and other 20-, 22-, and 24-carbon fatty acids (35.9% of total fatty acids).
-deficient basal diet (Group 1). The PUFA of M-0 (Groups 4 and 5), however, augmented the relatively low content of 18:2 (Table 7) to effect a chick growth response that equalled the growth promotion obtained from the much greater levels of 18:2 supplied by the S-O. These data indicate that the PUFA present in M-0 are accountable for the chick growth response. An essential fatty acid (EFA) deficiency is characterized by a decrease in the content of arachidonate (20:4) and an increase in the content of eicosatrienoic (20: 3) in the tissues of the animal (Holman, 1960). Decreased growth rate of EFA-deficient chicks as demonstrated in Experiment 1, and also reported by Edwards et al. (1963), and adverse effects on the reproductive functions of the EFA-deficient hen (Menge et al., 1965) have been correlated with increased 20:3 content of the tissues. Edwards et al. (1963) have observed that the PUFA of M-0 inhibit the synthesis of 20:3 from oleic acid (18:1). This inhibition of 20:3 synthesis by the PUFA has also been observed by Menge et al. (1965) and has
EFFECT OF OILS ON CHICK GROWTH TABLE 7.—Linoleic and polyunsaturated fatty acid intake per chick in Experiment 2l Group
Treatment
ariH*
1 2 3 4 5
Linoleic acid deficient 2.58% saflower oil 5.15% safflower oil 2.58% menhaden oil 5.15% menhaden oil
gm. 0.009 11.51 23.69 0.39 0.78
PUFA 3 gm. — 0.03" 0.06 4 5.12 10.25
1
Male chicks, 30 per group for 4-week experimental period. 2 Calculated from amount of oil consumed per chick. Basal diet contained 1.8 mg. 18:2/100 gm. (see T a b l e t ) . 3 Polyunsaturated fatty acids from oil consumed per chick. 4 Calculated from 0.2% linolenic acid in safflower oil.
been shown to result in increased reproductive activity in the hen, and as noted in Experiment 1, increased chick growth. Thus, the PUFA in the present studies can be assumed to have stimulated chick growth through the operation of at least two factors: (1) The depression of 20:3 synthesis from 18:1, and (2) The PUFA may have substituted, at least in part, for 18:2 or 20: 4, or both. Table 7 gives a resume of the unsaturated fatty acid intake during the 4-week experimental period in Experiment 2. The wide differences between the consumption of 18:2 and PUFA is, of course, a reflection of the unsaturated fatty acid content of S-0 and M-O. The amount of PUFA (M-O) consumed per chick in Group 5 (10.25 gm) was as effective in growth promotion as the nearly 24 gm 18:2 (S-O) consumed per chick in group 3 (Tables 6 and 7). The basal diet used in Experiment 1 contained unextracted soy protein and cellulose (Table 1), and as expected, a quantity of 18:2 (0.43%). The 18:2 inherent to the diet is apparent in the figures shown for the consumption of 18:2 by the different groups (Table 4). This dietary difference between Experiment 1 and Ex-
265
periment 2 is noted especially in a comparison between the 18:2 consumption of Group 6 in Experiment 1 (Table 4) and Groups 4 and 5 in Experiment 2 (Table 7). The growth stimulation from the PUFA of M-O in Experiment 1 (Table 3) is still apparent, however. SUMMARY
A comparison was made between the chick growth stimulation of the linoleate from safflower oil or coconut oil and the polyunsaturated fatty acids of menhaden oil. Significant growth responses were obtained from 1.65-3.3% linoleate supplied by safflower oil. No significant differences existed between the groups receiving relatively small amounts of linoleate from safflower or coconut oil, although dietary coconut oil at the 11.88% level did exhibit a probable growth depression. The high levels of saturated fatty acids in the coconut oil may, at least, be a contributing factor in this case. Relatively high levels of these fatty acids were found in the plasma lipids of the chicks fed coconut oil. Menhaden oil containing relatively low amounts of linoleate promoted chick growth as effectively as safflower oil that supplied much greater levels of linoleate. The responses obtained from menhaden oil were considered to be due to the polyunsaturated fatty acids of the oil which have been demonstrated to depress the synthesis of eicosatrienoic acid, and also they may have substituted for linoleate or arachidonate, or both, in the essential fatty acid-deficient chick. REFERENCES Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics, 11: 1-42. Edwards, H. M., Jr., J. E. Marion and J. C. Daggers, 1962. Studies on fat and fatty acid requirement of poultry. Proc. X l l t h World Poultry Congress, p. 182-186.
266
H. MENGE
Edwards, H. M., Jr., and J. E. Marion, 1963. Influence of dietary menhaden oil on growth rate and tissue fatty acids of the chick. J. Nutrition, 8 1 : 123-130. Havey, W. R., 1960. Least squares analysis of data with unequal numbers. U.S.D.A., A.R.S. 20-28. Holman, R. T., 1960. The ratio of trienoic-tetraenoic acids in tissue lipids as a measure of essential fatty acid requirement. J. Nutrition, 70: 405-410.
Menge, H., C. C. Calvert and C. A. Denton, 1965. Influence of dietary oils on reproduction in the hen. J. Nutrition, 87 : 365-370. Miller, E. C , H. Menge and C. A. Denton, 1963. Effect on long-term feeding of a fat-free diet to laying hens. J. Nutrition, 80: 431-440. Mohrhauer, H., and R. T. Holman, 1963. The effect of dose level of essential fatty acids upon fatty acid composition of the rat liver. J. Lipid Research, 4 : 151-159.
Motor Unit Recruitment Pattern in a Respiratory Muscle of Unanesthetized Chickens1 R. R. TSCHORN AND M. R. FEDDE Neuromuscular Laboratory, Department of Physiological Sciences, Kansas State University, Manhattan, Kansas 66502 (Received for publication August 10, 1970)
T
HE amplitude of electrical activity of respiratory muscles of the chicken, as recorded with a needle electrode, seems to be related to the level of anesthesia. Fedde et al. (1964) have shown that motor unit potentials of large amplitude are absent when chickens are deeply anesthetized. Since many investigations of the pattern of motor unit recruitment have used anesthetized animals (Sears, 1964; Fedde et al., 1969), we attempted to determine the motor unit recruitment pattern in unanesthetized chickens. METHODS A biopolar, electromyographic electrode (Fig. 1) was implanted in the transversus abdominis muscle (TA) in six, lightly anesthetized (20 mg./kg. of body weight of sodium pentobarbital administered intravenously) Single Comb White Leghorn males, 1 Contribution No. 68, Department of Physiological Sciences, College of Veterinary Medicine, KSAES, K.S.U., Manhattan, Kansas 66502. Supported by Public Health Service Research Grant NB-05786 from the National Institute of Neurological Disease and Stroke.
12 to 16 weeks old. The skin was incised along the ventro-lateral surface of the abdominal wall; then the fibers of the external abdominal oblique and rectus abdominis muscles were separated by blunt dissection to expose the lateral surface of the TA. The electrode was placed between the rectus abdominis and the TA muscles so that the wires were imbedded in the belly of the TA. The overlying muscles and skin then were sutured separately. The electrode cable was secured to the skin of the abdominal wall and back so that the chicken could not dislodge the electrode from its internal position. To be certain that the electrode was placed properly, electrical activity was monitored before the bird had recovered from the anesthetic by connecting it to a differential amplifier (Model PS, Grass Instrument Co., Quincy, Mass.), the output of which was inserted in one channel of a multichannel oscilloscope (Model 565, Tektronix, Inc., Beaverton, Oregon). A strain gauge transducer was constructed to record respiratory movements in unrestrained chickens by filling twelve centimeters of flexible rubber tubing, 5