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Relationship of Mineral Content of Drinking Water to Liver Lipid Accumulation in Laying Hens
Relationship of Mineral Content of Drinking Water to Liver Lipid Accumulation in Laying Hens L . S. JENSEN, D . V. MAURICE AND C. H . CHANG
Department of Poultry Science, University of Georgia, Athens, Georgia 30602 (Received for publication July 5, 1976)
POULTRY SCIENCE 56: 260-266, 1977
INTRODUCTION
I
N a previous study, water from egg farms in Georgia with a history of fatty liver syndrome had significantly more calcium, magnesium, strontium, sodium and iron than water from farms without a history of the disease (Jensen et al., 1976a). Although the results did not prove that water quality was involved in the etiology of the fatty liver syndrome, it did demonstrate an association of hardness of water with the disease that should be further investigated. There appears to be a variation in the ease with which investigators obtain high liver fat accumulation in hens fed typical corn-soybean meal, practical rations. For example, hens at Washington State University accumulated considerably more liver fat than hens at the University of Georgia even though fed similar rations (Jensen et al, 1976b).
liver lipid accumulation in laying hens and to directly compare liver lipid accumulation in hens given two sources of drinking water varying in mineral content. PROCEDURE Water samples were obtained from 15 poultry experiment stations and classed as to ease or difficulty of producing high liver lipid accumulation in laying hens on the basis of published information or by personal communication from investigators who supplied the water. The source and classification of the water samples and the references used for classifying are presented in Table 1. Following analysis for several mineral elements by plasma emission spectroscopy (Ronan, 1974), differences in mineral content between the two classes of water were analyzed for significance by the t test (Steel and Torrie, 1960). Four experiments were conducted to evaluate directly the effect of source of water
The present studies were conducted to relate the mineral content of water samples from several poultry experiment stations to 260
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ABSTRACT Fifteen poultry experiment stations in the United States were classified either positive or negative with respect to ease of producing fatty livers in laying hens, and water samples from these laboratories were analyzed for several mineral elements by plasma emission spectroscopy. Significantly more calcium and cobalt (P < 0.05) and magnesium, manganese, nickel and lead (P < 0.10) were found in samples from laboratories classified positively for ease of producing fatty livers. Water from one South Georgia farm with a history of fatty liver syndrome or from the University of Georgia (U.GA.) farm which has a low incidence of fatty liver syndrome was given to laying hens housed in floor pens or cages for an 8-week period. Little difference was observed in performance of the hens and no significant difference was observed in liver weight and lipid content of the liver dry matter. There was no significant difference in liver weight and lipid content at 29 and 34 weeks of pullets reared from day of age on the two sources of water. Egg production, liver weight and liver lipid content of hens given U.GA. water with 100 p.p.m. calcium (as CaCl2) and 50 p.p.m. magnesium (as MgCI2) for eight weeks did not differ significantly from hens given U.GA. water without the added elements. Results of the four experiments with laying hens directly comparing sources of water differing in mineral content do not support the hypothesis that excess liver lipid accumulation is associated with water hardness.
DRINKING WATER AND LIVER FAT
TABLE 1.—Source and classification of water samples from various experiment stations
on liver lipid accumulation in Single Comb White Leghorn laying hens. In the first experiment 20 hens approximately 44 weeks of age were placed in each of fo'ur colony houses (3.5 m. x 3.5 m.) with wood shavings as litter. Two pens were provided with the regular water source available at the U.GA. poultry farm which does not have a history of fatty liver syndrome, while two other pens were provided with water from a farm in South Georgia with a history of the disease. The water from this farm had been previously sampled in a survey of water samples from various locations in Georgia (Jensen et al., 1976a). The water contained 27 p.p.m. calcium and 19 p.p.m. magnesium as determined
by spectrographic analysis, and the U.GA. water contained 12 p.p.m. calcium and 2 p.p.m. magnesium. All hens were fed the U.GA. layer and breeder ration and both feed and water were given ad libitum. The hens were weighed individually at the start of the experiment and after 4 and 8 weeks. One-half of the hens in each pen were killed by electric shock after 4 weeks on experiment. Their livers were removed, weighed and liver moisture and lipid content determined. Liver lipid content was determined on a 4 gram sample of homogenized liver by extraction with chloroform: methanol (2:1) as described by Folch et al. (1957). The other half of the hens were killed at the end of 8 weeks on experiment. The experiment began May 13 and ended July 8. In Experiment 2, 72 hens approximately 52 weeks of age were placed in individual cages. Thirty-six hens were provided with regular U.GA. water while the rest of the hens were provided with the same sample of water used in Experiment 1 obtained from South Georgia. The experiment was started July 3 and ended September 9 at which time the hens were killed for obtaining livers for analysis as described for Experiment 1. The hens were individually weighed at the start and end of the experiment. In Experiment 3 Single Comb White Leghorn pullets were reared from day of age on either the U.GA. water or the same water sample from South Georgia as used in the first experiment. The chicks were fed the U.GA. starter diet to 8 weeks of age and the grower diet to 20 weeks of age. They were housed in the colony houses used in Experiment 1 which had wood shavings as litter. At 20 weeks of age the hens were moved to individual cages and continued on the same water sources but fed the U.GA. layer and breeder diet. Sixty hens were continued on the South Georgia water but only 30 hens were continued on the U.GA. water because of an accident in which half of the birds were
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Reference' Experiment station Classed as negative University of Georgia, Jensen et al. (1976b) Athens, GA University of California, Vohra (1976) Davis, CA Piedmont Research StaGarlich et al. (1975) tion, Salisbury, NC North Carolina UniverGarlich (1976) sity, Raleigh, NC Oklahoma State UniverThayer et al. (1973) sity, Stillwater, OK University of Arkansas, Waldroup (1976) Fayetteville, AR Latshaw and Osman Ohio State University, Columbus, OH (1974) Classed as positive University of Missouri, Wills and Savage Columbia, MO (1974) Washington State UniJensen et al. (1976b) versity, Pullman, WA South Dakota State UniChah et al. (1975) versity, Brookings, SD Louisiana State UniverGriffith et al. (1969) sity, Baton Rouge, LA Michigan State Universi- Barton (1967) ty, East Lansing, MI Cornell University, IthiIvy and Nesheim ca, NY (1973) New Mexico University, Robertson and TrujilLos Cruces, NM lo (1975) University of Florida, Harms et al. (1972) Gainesville, FL 'Reference used to classify water samples as to ease (positive) or difficulty (negative) of producing high liver fat accumulation in laying hens.
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L. S. JENSEN, D. V. MAURICE AND C. H. CHANG
TABLE 2.—Summary of mineral content of water samples from university poultry farms classified by ease of obtaining fatty livers in laying hens (means ± standard error) Statistical significance (t test)
Fatty livers'
+
-
Element 19 17 8 5 2.0 439 259 261 200 179 50 14 22 35 12 11
± 3 ± 10 ± 2 ± 2 ±0.2 ± 260 ±26 ± 94 ±28 ±99 ±31 ± 3 ± 4 ± 15 ± 5 ± 2
40 28 19 7 2.2 108 264 303 304 140 139 244 25 23 28 12
± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±
10 8 6 2 0.3 49 20 62 56 10 29 201 6 12 10 2
P < 0.05 NS PfiO.10 NS P<0.10 NS NS NS PsO.10 NS P < 0.05 NS NS NS P<0.10 NS
1 The water samples were classified in the basis of ease (+) or difficulty (-) of producing fatty livers in hens based on published data from the laboratories or personal communication with investigators.
lost during the rearing period. Half of the birds in each treatment were killed at 29 weeks of age (December 3) and the other half killed at 34 weeks of age (January 8) for obtaining livers for analysis as described in the previous experiments. In Experiment 4, sixty White Leghorn TABLE 3.—Effect
RESULTS A comparison of the mineral content of water samples from poultry experiment stations showed significantly more calcium and cobalt (P < 0.05) and more magnesium, nickel, lead and manganese (P < 0.10) for samples classified as positive for ease of
of water source on body weight and liver lipids in hens kept in floor pens (Experiment 1) Liver 2
Source of water' Four weeks U.GA. S.G. Eight weeks U.G.A. S.G.
Pen no.
Body weight (g. ± SE)
Weight (g. ± SE)
Lipids (%of D.M. ± SE)
Lipids/ liver/kg. B.W. (g.)
1 2 3 4
1665 1664 1581 1631
± ± ± ±
57 53 46 71
45.2 46.3 46.9 53.7
± ± ± ±
1.7 1.6 2.2 3.0
26.3 25.8 28.2 28.9
±2.1 ± 1.8 ± 2.4 ± 2.0
2.3 2.3 2.8 3.4
1 2 3 4
1675 1656 1716 1653
± ± ± ±
72 79 60 48
51.9 48.4 54.6 48.9
±5.6 ± 3.9 ± 3.4 ± 2.1
29.9 29.6 30.6 31.0
± ± ± ±
3.2 3.0 3.6 3.1
3.0 4.6 3.5 1.9
'U.GA. water was from the university farm and S.G. water from a farm in South Georgia with a history of fatty liver syndrome. 2 None of the values for the two sources of water were significantly different by t test (P > 0.05).
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Ca, p.p.m. Na, p.p.m. Mg, p.p.m. Si, p.p.m. Ni, p.p.m. Zn, p.p.b. Mo, p.p.b. Al, p.p.b. Pb, p.p.b. B, p.p.b. Co, p.p.b. Fe, p.p.b. Cr, p.p.b. Cu, p.p.b. Mn, p.p.b. Cd, p.p.b.
hens, approximately 48 weeks of age, were divided into four groups of 15 birds and housed in colony type houses with wood shavings as litter as used in Experiment 1. Feed and water were supplied ad libitum and the hens were fed the U.GA. layer and breeder ration. Two groups were given the U.GA. water and the other two U.GA. water containing an additional 100 p.p.m. calcium (added as CaCl 2 ) and 50 p.p.m. magnesium (MgCl 2 ). The hens had previously been housed in individual cages before being moved to the floor pens for the start of the experiment. The experiment was conducted for 8 weeks and measurements were made on egg production, egg weight, interior egg quality (Haugh units) and final body weight. At the end of the experiment the hens were killed by cervical dislocation and the livers were removed for determinations similar to those described previously.
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DRINKING WATER AND LIVER FAT
and percent lipids of dry matter were slightly less for these hens. A lower rate of egg production was caused by the hens being out of water during a short period of extreme hot weather in the middle of the experiment. In pullets raised from day of age and continued into the laying phase on the two sources of water, no significant difference was observed with respect to liver lipid accumulation (Table 5). Less lipids were observed in the liver at 34 weeks of age than at 29 weeks of age.
obtaining fatty livers (Table 2). Although percent liver fat of the dry matter was slightly higher for hens fed the South Georgia water in Experiment 1 (Table 3), differences between livers from hens given the two water samples were not significantly different (P > 0.05). Hens housed in individual cages in Experiment 2 did not differ significantly in liver fat accumulation as related to a source of water (Table 4). Rate of egg production was less for those fed the South Georgia water and liver mean weight
of source of water on performance of hens kept in individual cages (Experiment 2) Liver 2
Egg production
Source of water 2 TJ.GA. S.G.
Body weight
(%)
Final (g. ± SE)
Gain or loss (g.)
Weight (g. ± SE)
Lipids (%of D.M. ± SE)
Lipids/ liver/kg. B.W. (g.)
71.2 62.7
1733 ± 43 1678 ± 13
14 13
55.1 ± 1.7 51.2 ± 2.1
42.6 ± 2.2 39.2 ± 2.1
3.0 2.7
'U.GA. water was from the university farm and S.G. water from a farm in South Georgia with a history of fatty liver syndrome. 2 None of the values for the two sources of water were significantly different by t test (P > 0.05) TABLE 5.—Liver lipids of pullets reared from day of age on two sources of water (Experiment 3) Liver 2 Source of water' Age: 29 weeks U.GA. S.G. Age: 34 weeks U.GA. S.G.
Body weight (g. ± SE)
Weight (g. ± SE)
Lipids (%of D.M. ± SE)
Lipids/ liver/kg. (g.)
1759 ± 43 1718 ± 25
49.8 ± 2.3 53.7 ± 1.3
33.8 ± 2.2 35.8 ± 1.7
3.1 3.7
1760 ± 49 1784 ± 34
48.7 ± 2.0 45.5 ± 1.2
27.4 ± 1.8 22.4 ± 1.7
2.5 1.7
'U.GA. water was from the university farm and S.G. water from a farm in South Georgia with a history of fatty liver syndrome. 2 None of the values for the two sources of water were different by t test (P > 0.05). TABLE 6.—Performance of hens in floor pens given drinking water with and without either Ca and Mg (Experiment 4)x
Water supplement None Ca + Mg 2
Egg production
(%) 80.2 79.7
Egg weight (g.) 58.4 60.4
Haugh units 89.2 87.6
Body weight (g-) 1739 1741
Weight (g.) 44.8 45.0
'None of the differences were significantly different (P > 0.05). Contained 100 p.p.m. Ca (CaCl2) and 50 p.p.m. (MgCl2).
Liver Lipids (%of D.M.) 20.2 18.9
Lipids/ liver/kg. (g.) 1.7 1.6
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TABLE 4.—Effect
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L. S. JENSEN, D. V. MAURICE AND C. H. CHANG
TABLE 7.—Liver parameters of hens maintained in cages compared to similar hens maintained for an additional eight weeks in floor pens
Housing Cage (pre-experiment) Floor (post-experiment)
Weight (g)
Liver Lipids (%of D.M.)
Lipids/ liver/kg. B.W. (g.)
33.2a
26.2a
3.0a
25.8b
19.5b
1.6b
1
Values without a common letter are significantly different (P < 0.05).
DISCUSSION A previous study in which samples of water from farms with and without a history of fatty liver syndrome in Georgia showed that hardness of water was associated with incidence of the disease (Jensen et al., 1976a). The data presented here on analysis of water samples from poultry farms at various experiment stations in the United States again suggest that the susceptibility of laying hens to fatty livers is associated with hardness of water. Water samples from experiment stations classed as positive with respect to ease of obtaining fatty livers in laying hen experiments contained significantly more calcium (P < 0.05) and magnesium (P < 0.10). There was considerable variation in the mineral content of water samples within each classified group. An extreme case was water from Louisiana State University which was classed as positive for ease of obtaining fatty
In four experiments conducted to directly compare two types of drinking water either with respect to source or to levels of added minerals, no evidence was obtained for a significant difference in liver lipid accumulation caused by type of water. The first experiment was conducted in floor pens where less liver fat accumulation is expected in comparison to birds housed in cages (Barton, 1967). The second experiment was conducted in
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Adding calcium and magnesium to the U.GA. water failed to significantly affect liver lipid content of hens in Experiment 4 (Table 6). No significant difference in any performance criteria between the two sources of water was observed. Hens maintained in individual cages sampled prior to the start of the experiment had significantly more liver fat and larger livers than hens housed in floor pens after 8 weeks on the experiment (Table 7).
livers based on several publications from that institution. It had only 1.2 p.p.m. calcium and 0.1 p.p.m. magnesium. The level of cobalt was significantly higher in water samples classed as positive for ease of obtaining fatty livers. Wolford and Murphy (1972), however, did not observe any significant effect on liver size or lipid levels in hens fed diets supplemented with 0, 5, 10 or 20 p.p.m. cobalt. An obvious criticism of the data presented in Table 2 comparing two groups of water samples is the subjective basis for classification of the two groups. Some institutions from which water samples were obtained have published a number of papers dealing with liver fat accumulation in laying hens while others have published none. Nevertheless, data obtained further support an association of hardness of water with liver fat accumulation. Although the quantities of calcium and magnesium in the water differ significantly, the quantities of these two elements contributed by water to the total intake by the birds is of a low order. It is difficult to comprehend how such differences in the level of calcium and magnesium in the water could play a significant role in altering lipid metabolism. A more likely effect might be an interference of calcium and magnesium in water with absorption of some other essential trace element needed for normal lipid metabolism. No evidence was obtained, however, for significantly lower levels of trace elements measured in water samples from experiment stations classified positive for fatty livers.
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Only one water sample from South Georgia was used in the experiments for comparison with the U.GA. water. Perhaps different results would have been obtained with other water samples, particularly those with harder water. The same U.GA. layer and breeder ration was used in all studies. This ration has generally resulted in a low level of lipid accumulation in livers of laying hens under our conditions. Other studies at this laboratory have shown that composition of the diet significantly influences liver lipid accumulation in hens (Jensen et al., 1976b, c). Employment of other diets might have yielded different results in liver fat in response to the different water sources. Although no direct evidence was obtained in these studies for a cause and effect relationship between water quality and liver lipid accumulation in laying hens, further experiments under a variety of conditions should be conducted before rejecting the hypothesis. ACKNOWLEDGMENT The authors are indebted to the following for submitting water samples from their re-
spective experiment stations: C. R. Douglas, J. D. Garlich, E. Guenthner, J. D. Latshaw, R. Lein, D. Polin, M. H. Pubols, R. H. Roberson, J. E. Savage, R. H. Thayer, P. W. Waldroup, A. B. Watts, and R. J. Young.
REFERENCES Barton, T. L., 1967. Fatty liver studies in laying hens. Ph.D. Thesis, Michigan State Univ. Chah, C. C , R. A. Nelson and C. W. Carlson, 1975. Fatty liver-hemorrhagic syndrome as affected by fat, choline and biotin supplementation. Poultry Sci. 54: 1743. Folch, J., M. Lees and C. H. S. Stanley, 1957. A simple method for the isolation and purification of total lipids from animal tissue. J. Biol. Chem. 225: 497-509. Garlich, J. D., J. D. Olson, W. E. Huff and P. B. Hamilton, 1975. Liver lipid content of twenty varieties of laying hens from three confinement systems. Poultry Sci. 54: 806-813. Garlich, J. D., 1976. Personal communication. Griffith, M., A. J. Olinde, R. Schexnailder, R. F. Davenport and W. F. McKnight, 1969. Effect of choline, methionine and vitamin B l2 on liver fat, egg production and egg weight in hens. Poultry Sci. 48: 2160-2172. Harms, R. H., C. F. Simpson and B. L. Damron, 1972. Some new observations on "fatty liver syndrome" in laying hens. Avian Dis. 16: 1042-1046. Ivy, C. A., and M. C. Nesheim, 1973. Factors influencing the liver fat content of laying hens. Poultry Sci. 52: 281-291. Jensen, L. S., J. M. Casey, S. I. Savage and W. M. Britton, 1976a. An association of hardness of water with incidence of fatty liver syndromes in laying hens. Poultry Sci. 55: 719-724. Jensen, L. S., C. H. Chang and R. D. Wyatt, 1976b. Influence of carbohydrate source on liver fat accumulation in hens. Poultry Sci. 55: 700-709. Jensen, L. S., C. H. Chang and D. V. Maurice, 1976c. Improvement in albumen quality and reduction in liver fat in hens fed brewers dried grains. Poultry Sci. 55: 1841-1847. Latshaw, J. D., and M. Osman, 1974. A selenium and vitamin E responsive condition in the laying hen. Poultry Sci. 53: 1704-1708. Roberson, R. H., and V. Trujillo, 1975. The effect of methionine, thiouracil, dienestrol diacetate and thyroprotein on the development and prevention of
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individual cages, but the hens fed the water from a South Georgia farm with a history of fatty liver syndrome were accidentally out of water for a few hours during the middle of the experiment. This resulted in some reduction in egg production and perhaps affected final liver weights and lipid content. The third experiment, however, was conducted with pullets that had been given the two sources of water from day of age and then were maintained in individual laying cages for several weeks before analyzing livers. No evidence was obtained in the latter experiment for a difference in liver fat accumulation. The fourth experiment involved addition of calcium and magnesium salts to the U.GA. water, but was again conducted in floor pens with no significant difference in liver fat accumulation noted between groups.
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fatty liver in pullets. Poultry Sci. 54: 715-721. Ronan, R., 1974. Simultaneous analysis of liquid samples for metals by inductively coupled argon plasma atomic emission spectroscopy, EPA, Region V, Central Research Laboratory, Chicago, 111. Steel, R. G. D., and J. H. Torrie, 1960. Principles and Procedures of Statistics. McGraw-Hill Book Company, Inc., New York, N.Y. Thayer, R. H., E. C. Nelson, E. T. Clemens, R. R. Johnson and A. L. Molle, 1973. Lipid composition
of livers from laying hens. Poultry Sci. 52: 22702275. Vohra, P., 1976. Personal communication. Waldroup, P. W., 1976. Personal communication. Wills, J. R., and J. E. Savage, 1974. Ultrastructure and lipid composition of livers from hens with hemorrhagic livers. Poultry Sci. 53: 1992. Wolford, J. H., and D. Murphy, 1972. Effect of diet on fatty liver-hemorrhagic syndrome incidence in laying chickens. Poultry Sci. 51: 2087-2094.
WAYNE B . W H I T E AND S. L . BALLOUN
Department of Animal Science, Iowa State University, Ames, Iowa 50011 (Received for publication July 5, 1976)
ABSTRACT Broiler-type chicks were used in four experiments to determine the nutritional value of a methanol-derived single-cell protein (SCP). The trials ran from 7 to 28 days of age, with SCP replacing soybean meal up to 15% of the diet. The powdery consistency of SCP tended to reduce body weight gain and feed conversion efficiency when fed at 9% or more of the diet when diets were fed as mash. Pelleting diets containing SCP eliminated these effects. The response to supplemental methionine in SCP diets was similar to that of corn-soybean meal diets. When diets were pelleted, substitution of SCPfor soybean meal, on a weight-for-weight basis at levels up to 15% of the diet produced greater and more efficient gains. POULTRY SCIENCE 56: 266-273, 1977
INTRODUCTION
T
HE value of single-cell protein (SCP) as a feedstuff has been studied for a number of microorganisms produced on different substrates. The substrates have included alkanes of increasing length, from methane to n-paraffins. Methanol also has been used because of its high solubility in water. Data reflecting the net protein utilization (NPU) values of SCP were given by Palmer and Smith (1971). They compared four yeast and two bacterial products. Waldroup and Flynn (1975) reported NPU values ranging from 40.8 to 73.5 for yeasts produced on different hydrocarbon feedstocks with varia-
'Journal Paper No. J-8542 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa, Project No. 1932.
tions in methods of processing, compared with a value of 85.5 for soybean meal. Woodham and Deans (1973) evaluated nine SCP products and showed that, on the basis of total protein efficiency (TPE), methionine was deficient in all the SCP diets, except in distiller's yeast. Growth and feed efficiency of chicks fed a bacterial product grown on methanol was improved by pelleting SCP diets (Waldroup and Payne, 1974). Waldroup et al. (1971) stated that the reduced performance of chicks fed SCP was caused by a reduction in feed consumption. They evaluated a yeast grown on hydrocarbons, which, when pair-fed, produced a growth response equal to that of the control. SCP in the diet does not increase plasma uric acid (Shannon and McNab, 1972; D'Mello, 1973). The digestibility of various components of a yeast grown on n-paraffins and fed to
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The Value of Methanol-derived Single-cell Protein for BroilersJ
Department of Poultry Science, University of Georgia, Athens, Georgia 30602 (Received for publication July 5, 1976)
POULTRY SCIENCE 56: 260-266, 1977
INTRODUCTION
I
N a previous study, water from egg farms in Georgia with a history of fatty liver syndrome had significantly more calcium, magnesium, strontium, sodium and iron than water from farms without a history of the disease (Jensen et al., 1976a). Although the results did not prove that water quality was involved in the etiology of the fatty liver syndrome, it did demonstrate an association of hardness of water with the disease that should be further investigated. There appears to be a variation in the ease with which investigators obtain high liver fat accumulation in hens fed typical corn-soybean meal, practical rations. For example, hens at Washington State University accumulated considerably more liver fat than hens at the University of Georgia even though fed similar rations (Jensen et al, 1976b).
liver lipid accumulation in laying hens and to directly compare liver lipid accumulation in hens given two sources of drinking water varying in mineral content. PROCEDURE Water samples were obtained from 15 poultry experiment stations and classed as to ease or difficulty of producing high liver lipid accumulation in laying hens on the basis of published information or by personal communication from investigators who supplied the water. The source and classification of the water samples and the references used for classifying are presented in Table 1. Following analysis for several mineral elements by plasma emission spectroscopy (Ronan, 1974), differences in mineral content between the two classes of water were analyzed for significance by the t test (Steel and Torrie, 1960). Four experiments were conducted to evaluate directly the effect of source of water
The present studies were conducted to relate the mineral content of water samples from several poultry experiment stations to 260
Downloaded from http://ps.oxfordjournals.org/ by guest on April 11, 2015
ABSTRACT Fifteen poultry experiment stations in the United States were classified either positive or negative with respect to ease of producing fatty livers in laying hens, and water samples from these laboratories were analyzed for several mineral elements by plasma emission spectroscopy. Significantly more calcium and cobalt (P < 0.05) and magnesium, manganese, nickel and lead (P < 0.10) were found in samples from laboratories classified positively for ease of producing fatty livers. Water from one South Georgia farm with a history of fatty liver syndrome or from the University of Georgia (U.GA.) farm which has a low incidence of fatty liver syndrome was given to laying hens housed in floor pens or cages for an 8-week period. Little difference was observed in performance of the hens and no significant difference was observed in liver weight and lipid content of the liver dry matter. There was no significant difference in liver weight and lipid content at 29 and 34 weeks of pullets reared from day of age on the two sources of water. Egg production, liver weight and liver lipid content of hens given U.GA. water with 100 p.p.m. calcium (as CaCl2) and 50 p.p.m. magnesium (as MgCI2) for eight weeks did not differ significantly from hens given U.GA. water without the added elements. Results of the four experiments with laying hens directly comparing sources of water differing in mineral content do not support the hypothesis that excess liver lipid accumulation is associated with water hardness.
DRINKING WATER AND LIVER FAT
TABLE 1.—Source and classification of water samples from various experiment stations
on liver lipid accumulation in Single Comb White Leghorn laying hens. In the first experiment 20 hens approximately 44 weeks of age were placed in each of fo'ur colony houses (3.5 m. x 3.5 m.) with wood shavings as litter. Two pens were provided with the regular water source available at the U.GA. poultry farm which does not have a history of fatty liver syndrome, while two other pens were provided with water from a farm in South Georgia with a history of the disease. The water from this farm had been previously sampled in a survey of water samples from various locations in Georgia (Jensen et al., 1976a). The water contained 27 p.p.m. calcium and 19 p.p.m. magnesium as determined
by spectrographic analysis, and the U.GA. water contained 12 p.p.m. calcium and 2 p.p.m. magnesium. All hens were fed the U.GA. layer and breeder ration and both feed and water were given ad libitum. The hens were weighed individually at the start of the experiment and after 4 and 8 weeks. One-half of the hens in each pen were killed by electric shock after 4 weeks on experiment. Their livers were removed, weighed and liver moisture and lipid content determined. Liver lipid content was determined on a 4 gram sample of homogenized liver by extraction with chloroform: methanol (2:1) as described by Folch et al. (1957). The other half of the hens were killed at the end of 8 weeks on experiment. The experiment began May 13 and ended July 8. In Experiment 2, 72 hens approximately 52 weeks of age were placed in individual cages. Thirty-six hens were provided with regular U.GA. water while the rest of the hens were provided with the same sample of water used in Experiment 1 obtained from South Georgia. The experiment was started July 3 and ended September 9 at which time the hens were killed for obtaining livers for analysis as described for Experiment 1. The hens were individually weighed at the start and end of the experiment. In Experiment 3 Single Comb White Leghorn pullets were reared from day of age on either the U.GA. water or the same water sample from South Georgia as used in the first experiment. The chicks were fed the U.GA. starter diet to 8 weeks of age and the grower diet to 20 weeks of age. They were housed in the colony houses used in Experiment 1 which had wood shavings as litter. At 20 weeks of age the hens were moved to individual cages and continued on the same water sources but fed the U.GA. layer and breeder diet. Sixty hens were continued on the South Georgia water but only 30 hens were continued on the U.GA. water because of an accident in which half of the birds were
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Reference' Experiment station Classed as negative University of Georgia, Jensen et al. (1976b) Athens, GA University of California, Vohra (1976) Davis, CA Piedmont Research StaGarlich et al. (1975) tion, Salisbury, NC North Carolina UniverGarlich (1976) sity, Raleigh, NC Oklahoma State UniverThayer et al. (1973) sity, Stillwater, OK University of Arkansas, Waldroup (1976) Fayetteville, AR Latshaw and Osman Ohio State University, Columbus, OH (1974) Classed as positive University of Missouri, Wills and Savage Columbia, MO (1974) Washington State UniJensen et al. (1976b) versity, Pullman, WA South Dakota State UniChah et al. (1975) versity, Brookings, SD Louisiana State UniverGriffith et al. (1969) sity, Baton Rouge, LA Michigan State Universi- Barton (1967) ty, East Lansing, MI Cornell University, IthiIvy and Nesheim ca, NY (1973) New Mexico University, Robertson and TrujilLos Cruces, NM lo (1975) University of Florida, Harms et al. (1972) Gainesville, FL 'Reference used to classify water samples as to ease (positive) or difficulty (negative) of producing high liver fat accumulation in laying hens.
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L. S. JENSEN, D. V. MAURICE AND C. H. CHANG
TABLE 2.—Summary of mineral content of water samples from university poultry farms classified by ease of obtaining fatty livers in laying hens (means ± standard error) Statistical significance (t test)
Fatty livers'
+
-
Element 19 17 8 5 2.0 439 259 261 200 179 50 14 22 35 12 11
± 3 ± 10 ± 2 ± 2 ±0.2 ± 260 ±26 ± 94 ±28 ±99 ±31 ± 3 ± 4 ± 15 ± 5 ± 2
40 28 19 7 2.2 108 264 303 304 140 139 244 25 23 28 12
± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±
10 8 6 2 0.3 49 20 62 56 10 29 201 6 12 10 2
P < 0.05 NS PfiO.10 NS P<0.10 NS NS NS PsO.10 NS P < 0.05 NS NS NS P<0.10 NS
1 The water samples were classified in the basis of ease (+) or difficulty (-) of producing fatty livers in hens based on published data from the laboratories or personal communication with investigators.
lost during the rearing period. Half of the birds in each treatment were killed at 29 weeks of age (December 3) and the other half killed at 34 weeks of age (January 8) for obtaining livers for analysis as described in the previous experiments. In Experiment 4, sixty White Leghorn TABLE 3.—Effect
RESULTS A comparison of the mineral content of water samples from poultry experiment stations showed significantly more calcium and cobalt (P < 0.05) and more magnesium, nickel, lead and manganese (P < 0.10) for samples classified as positive for ease of
of water source on body weight and liver lipids in hens kept in floor pens (Experiment 1) Liver 2
Source of water' Four weeks U.GA. S.G. Eight weeks U.G.A. S.G.
Pen no.
Body weight (g. ± SE)
Weight (g. ± SE)
Lipids (%of D.M. ± SE)
Lipids/ liver/kg. B.W. (g.)
1 2 3 4
1665 1664 1581 1631
± ± ± ±
57 53 46 71
45.2 46.3 46.9 53.7
± ± ± ±
1.7 1.6 2.2 3.0
26.3 25.8 28.2 28.9
±2.1 ± 1.8 ± 2.4 ± 2.0
2.3 2.3 2.8 3.4
1 2 3 4
1675 1656 1716 1653
± ± ± ±
72 79 60 48
51.9 48.4 54.6 48.9
±5.6 ± 3.9 ± 3.4 ± 2.1
29.9 29.6 30.6 31.0
± ± ± ±
3.2 3.0 3.6 3.1
3.0 4.6 3.5 1.9
'U.GA. water was from the university farm and S.G. water from a farm in South Georgia with a history of fatty liver syndrome. 2 None of the values for the two sources of water were significantly different by t test (P > 0.05).
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Ca, p.p.m. Na, p.p.m. Mg, p.p.m. Si, p.p.m. Ni, p.p.m. Zn, p.p.b. Mo, p.p.b. Al, p.p.b. Pb, p.p.b. B, p.p.b. Co, p.p.b. Fe, p.p.b. Cr, p.p.b. Cu, p.p.b. Mn, p.p.b. Cd, p.p.b.
hens, approximately 48 weeks of age, were divided into four groups of 15 birds and housed in colony type houses with wood shavings as litter as used in Experiment 1. Feed and water were supplied ad libitum and the hens were fed the U.GA. layer and breeder ration. Two groups were given the U.GA. water and the other two U.GA. water containing an additional 100 p.p.m. calcium (added as CaCl 2 ) and 50 p.p.m. magnesium (MgCl 2 ). The hens had previously been housed in individual cages before being moved to the floor pens for the start of the experiment. The experiment was conducted for 8 weeks and measurements were made on egg production, egg weight, interior egg quality (Haugh units) and final body weight. At the end of the experiment the hens were killed by cervical dislocation and the livers were removed for determinations similar to those described previously.
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DRINKING WATER AND LIVER FAT
and percent lipids of dry matter were slightly less for these hens. A lower rate of egg production was caused by the hens being out of water during a short period of extreme hot weather in the middle of the experiment. In pullets raised from day of age and continued into the laying phase on the two sources of water, no significant difference was observed with respect to liver lipid accumulation (Table 5). Less lipids were observed in the liver at 34 weeks of age than at 29 weeks of age.
obtaining fatty livers (Table 2). Although percent liver fat of the dry matter was slightly higher for hens fed the South Georgia water in Experiment 1 (Table 3), differences between livers from hens given the two water samples were not significantly different (P > 0.05). Hens housed in individual cages in Experiment 2 did not differ significantly in liver fat accumulation as related to a source of water (Table 4). Rate of egg production was less for those fed the South Georgia water and liver mean weight
of source of water on performance of hens kept in individual cages (Experiment 2) Liver 2
Egg production
Source of water 2 TJ.GA. S.G.
Body weight
(%)
Final (g. ± SE)
Gain or loss (g.)
Weight (g. ± SE)
Lipids (%of D.M. ± SE)
Lipids/ liver/kg. B.W. (g.)
71.2 62.7
1733 ± 43 1678 ± 13
14 13
55.1 ± 1.7 51.2 ± 2.1
42.6 ± 2.2 39.2 ± 2.1
3.0 2.7
'U.GA. water was from the university farm and S.G. water from a farm in South Georgia with a history of fatty liver syndrome. 2 None of the values for the two sources of water were significantly different by t test (P > 0.05) TABLE 5.—Liver lipids of pullets reared from day of age on two sources of water (Experiment 3) Liver 2 Source of water' Age: 29 weeks U.GA. S.G. Age: 34 weeks U.GA. S.G.
Body weight (g. ± SE)
Weight (g. ± SE)
Lipids (%of D.M. ± SE)
Lipids/ liver/kg. (g.)
1759 ± 43 1718 ± 25
49.8 ± 2.3 53.7 ± 1.3
33.8 ± 2.2 35.8 ± 1.7
3.1 3.7
1760 ± 49 1784 ± 34
48.7 ± 2.0 45.5 ± 1.2
27.4 ± 1.8 22.4 ± 1.7
2.5 1.7
'U.GA. water was from the university farm and S.G. water from a farm in South Georgia with a history of fatty liver syndrome. 2 None of the values for the two sources of water were different by t test (P > 0.05). TABLE 6.—Performance of hens in floor pens given drinking water with and without either Ca and Mg (Experiment 4)x
Water supplement None Ca + Mg 2
Egg production
(%) 80.2 79.7
Egg weight (g.) 58.4 60.4
Haugh units 89.2 87.6
Body weight (g-) 1739 1741
Weight (g.) 44.8 45.0
'None of the differences were significantly different (P > 0.05). Contained 100 p.p.m. Ca (CaCl2) and 50 p.p.m. (MgCl2).
Liver Lipids (%of D.M.) 20.2 18.9
Lipids/ liver/kg. (g.) 1.7 1.6
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TABLE 4.—Effect
264
L. S. JENSEN, D. V. MAURICE AND C. H. CHANG
TABLE 7.—Liver parameters of hens maintained in cages compared to similar hens maintained for an additional eight weeks in floor pens
Housing Cage (pre-experiment) Floor (post-experiment)
Weight (g)
Liver Lipids (%of D.M.)
Lipids/ liver/kg. B.W. (g.)
33.2a
26.2a
3.0a
25.8b
19.5b
1.6b
1
Values without a common letter are significantly different (P < 0.05).
DISCUSSION A previous study in which samples of water from farms with and without a history of fatty liver syndrome in Georgia showed that hardness of water was associated with incidence of the disease (Jensen et al., 1976a). The data presented here on analysis of water samples from poultry farms at various experiment stations in the United States again suggest that the susceptibility of laying hens to fatty livers is associated with hardness of water. Water samples from experiment stations classed as positive with respect to ease of obtaining fatty livers in laying hen experiments contained significantly more calcium (P < 0.05) and magnesium (P < 0.10). There was considerable variation in the mineral content of water samples within each classified group. An extreme case was water from Louisiana State University which was classed as positive for ease of obtaining fatty
In four experiments conducted to directly compare two types of drinking water either with respect to source or to levels of added minerals, no evidence was obtained for a significant difference in liver lipid accumulation caused by type of water. The first experiment was conducted in floor pens where less liver fat accumulation is expected in comparison to birds housed in cages (Barton, 1967). The second experiment was conducted in
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Adding calcium and magnesium to the U.GA. water failed to significantly affect liver lipid content of hens in Experiment 4 (Table 6). No significant difference in any performance criteria between the two sources of water was observed. Hens maintained in individual cages sampled prior to the start of the experiment had significantly more liver fat and larger livers than hens housed in floor pens after 8 weeks on the experiment (Table 7).
livers based on several publications from that institution. It had only 1.2 p.p.m. calcium and 0.1 p.p.m. magnesium. The level of cobalt was significantly higher in water samples classed as positive for ease of obtaining fatty livers. Wolford and Murphy (1972), however, did not observe any significant effect on liver size or lipid levels in hens fed diets supplemented with 0, 5, 10 or 20 p.p.m. cobalt. An obvious criticism of the data presented in Table 2 comparing two groups of water samples is the subjective basis for classification of the two groups. Some institutions from which water samples were obtained have published a number of papers dealing with liver fat accumulation in laying hens while others have published none. Nevertheless, data obtained further support an association of hardness of water with liver fat accumulation. Although the quantities of calcium and magnesium in the water differ significantly, the quantities of these two elements contributed by water to the total intake by the birds is of a low order. It is difficult to comprehend how such differences in the level of calcium and magnesium in the water could play a significant role in altering lipid metabolism. A more likely effect might be an interference of calcium and magnesium in water with absorption of some other essential trace element needed for normal lipid metabolism. No evidence was obtained, however, for significantly lower levels of trace elements measured in water samples from experiment stations classified positive for fatty livers.
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DRINKING WATER AND LIVER FAT
Only one water sample from South Georgia was used in the experiments for comparison with the U.GA. water. Perhaps different results would have been obtained with other water samples, particularly those with harder water. The same U.GA. layer and breeder ration was used in all studies. This ration has generally resulted in a low level of lipid accumulation in livers of laying hens under our conditions. Other studies at this laboratory have shown that composition of the diet significantly influences liver lipid accumulation in hens (Jensen et al., 1976b, c). Employment of other diets might have yielded different results in liver fat in response to the different water sources. Although no direct evidence was obtained in these studies for a cause and effect relationship between water quality and liver lipid accumulation in laying hens, further experiments under a variety of conditions should be conducted before rejecting the hypothesis. ACKNOWLEDGMENT The authors are indebted to the following for submitting water samples from their re-
spective experiment stations: C. R. Douglas, J. D. Garlich, E. Guenthner, J. D. Latshaw, R. Lein, D. Polin, M. H. Pubols, R. H. Roberson, J. E. Savage, R. H. Thayer, P. W. Waldroup, A. B. Watts, and R. J. Young.
REFERENCES Barton, T. L., 1967. Fatty liver studies in laying hens. Ph.D. Thesis, Michigan State Univ. Chah, C. C , R. A. Nelson and C. W. Carlson, 1975. Fatty liver-hemorrhagic syndrome as affected by fat, choline and biotin supplementation. Poultry Sci. 54: 1743. Folch, J., M. Lees and C. H. S. Stanley, 1957. A simple method for the isolation and purification of total lipids from animal tissue. J. Biol. Chem. 225: 497-509. Garlich, J. D., J. D. Olson, W. E. Huff and P. B. Hamilton, 1975. Liver lipid content of twenty varieties of laying hens from three confinement systems. Poultry Sci. 54: 806-813. Garlich, J. D., 1976. Personal communication. Griffith, M., A. J. Olinde, R. Schexnailder, R. F. Davenport and W. F. McKnight, 1969. Effect of choline, methionine and vitamin B l2 on liver fat, egg production and egg weight in hens. Poultry Sci. 48: 2160-2172. Harms, R. H., C. F. Simpson and B. L. Damron, 1972. Some new observations on "fatty liver syndrome" in laying hens. Avian Dis. 16: 1042-1046. Ivy, C. A., and M. C. Nesheim, 1973. Factors influencing the liver fat content of laying hens. Poultry Sci. 52: 281-291. Jensen, L. S., J. M. Casey, S. I. Savage and W. M. Britton, 1976a. An association of hardness of water with incidence of fatty liver syndromes in laying hens. Poultry Sci. 55: 719-724. Jensen, L. S., C. H. Chang and R. D. Wyatt, 1976b. Influence of carbohydrate source on liver fat accumulation in hens. Poultry Sci. 55: 700-709. Jensen, L. S., C. H. Chang and D. V. Maurice, 1976c. Improvement in albumen quality and reduction in liver fat in hens fed brewers dried grains. Poultry Sci. 55: 1841-1847. Latshaw, J. D., and M. Osman, 1974. A selenium and vitamin E responsive condition in the laying hen. Poultry Sci. 53: 1704-1708. Roberson, R. H., and V. Trujillo, 1975. The effect of methionine, thiouracil, dienestrol diacetate and thyroprotein on the development and prevention of
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individual cages, but the hens fed the water from a South Georgia farm with a history of fatty liver syndrome were accidentally out of water for a few hours during the middle of the experiment. This resulted in some reduction in egg production and perhaps affected final liver weights and lipid content. The third experiment, however, was conducted with pullets that had been given the two sources of water from day of age and then were maintained in individual laying cages for several weeks before analyzing livers. No evidence was obtained in the latter experiment for a difference in liver fat accumulation. The fourth experiment involved addition of calcium and magnesium salts to the U.GA. water, but was again conducted in floor pens with no significant difference in liver fat accumulation noted between groups.
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L. S. JENSEN, D. V. MAURICE AND C. H. CHANG
fatty liver in pullets. Poultry Sci. 54: 715-721. Ronan, R., 1974. Simultaneous analysis of liquid samples for metals by inductively coupled argon plasma atomic emission spectroscopy, EPA, Region V, Central Research Laboratory, Chicago, 111. Steel, R. G. D., and J. H. Torrie, 1960. Principles and Procedures of Statistics. McGraw-Hill Book Company, Inc., New York, N.Y. Thayer, R. H., E. C. Nelson, E. T. Clemens, R. R. Johnson and A. L. Molle, 1973. Lipid composition
of livers from laying hens. Poultry Sci. 52: 22702275. Vohra, P., 1976. Personal communication. Waldroup, P. W., 1976. Personal communication. Wills, J. R., and J. E. Savage, 1974. Ultrastructure and lipid composition of livers from hens with hemorrhagic livers. Poultry Sci. 53: 1992. Wolford, J. H., and D. Murphy, 1972. Effect of diet on fatty liver-hemorrhagic syndrome incidence in laying chickens. Poultry Sci. 51: 2087-2094.
WAYNE B . W H I T E AND S. L . BALLOUN
Department of Animal Science, Iowa State University, Ames, Iowa 50011 (Received for publication July 5, 1976)
ABSTRACT Broiler-type chicks were used in four experiments to determine the nutritional value of a methanol-derived single-cell protein (SCP). The trials ran from 7 to 28 days of age, with SCP replacing soybean meal up to 15% of the diet. The powdery consistency of SCP tended to reduce body weight gain and feed conversion efficiency when fed at 9% or more of the diet when diets were fed as mash. Pelleting diets containing SCP eliminated these effects. The response to supplemental methionine in SCP diets was similar to that of corn-soybean meal diets. When diets were pelleted, substitution of SCPfor soybean meal, on a weight-for-weight basis at levels up to 15% of the diet produced greater and more efficient gains. POULTRY SCIENCE 56: 266-273, 1977
INTRODUCTION
T
HE value of single-cell protein (SCP) as a feedstuff has been studied for a number of microorganisms produced on different substrates. The substrates have included alkanes of increasing length, from methane to n-paraffins. Methanol also has been used because of its high solubility in water. Data reflecting the net protein utilization (NPU) values of SCP were given by Palmer and Smith (1971). They compared four yeast and two bacterial products. Waldroup and Flynn (1975) reported NPU values ranging from 40.8 to 73.5 for yeasts produced on different hydrocarbon feedstocks with varia-
'Journal Paper No. J-8542 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa, Project No. 1932.
tions in methods of processing, compared with a value of 85.5 for soybean meal. Woodham and Deans (1973) evaluated nine SCP products and showed that, on the basis of total protein efficiency (TPE), methionine was deficient in all the SCP diets, except in distiller's yeast. Growth and feed efficiency of chicks fed a bacterial product grown on methanol was improved by pelleting SCP diets (Waldroup and Payne, 1974). Waldroup et al. (1971) stated that the reduced performance of chicks fed SCP was caused by a reduction in feed consumption. They evaluated a yeast grown on hydrocarbons, which, when pair-fed, produced a growth response equal to that of the control. SCP in the diet does not increase plasma uric acid (Shannon and McNab, 1972; D'Mello, 1973). The digestibility of various components of a yeast grown on n-paraffins and fed to
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The Value of Methanol-derived Single-cell Protein for BroilersJ