Factors affecting serum mineral levels and the influence of dietary calcium and phosphorus interrelationships on the per...

MISSING IMAGE

Material Information

Title:
Factors affecting serum mineral levels and the influence of dietary calcium and phosphorus interrelationships on the performance characteristics of the domestic hen
Physical Description:
ix, 78 leaves : ill. ; 28 cm.
Language:
English
Creator:
Miller, Edgar Reid, 1950-
Publication Date:

Subjects

Subjects / Keywords:
Poultry -- Feeding and feeds   ( lcsh )
Genre:
bibliography   ( marcgt )
theses   ( marcgt )
non-fiction   ( marcgt )

Notes

Thesis:
Thesis--University of Florida.
Bibliography:
Includes bibliographical references (leaves 72-77).
Statement of Responsibility:
by Edgar Reid Miller.
General Note:
Typescript.
General Note:
Vita.

Record Information

Source Institution:
University of Florida
Rights Management:
All applicable rights reserved by the source institution and holding location.
Resource Identifier:
aleph - 000009134
notis - AAB1009
oclc - 02527717
System ID:
AA00003516:00001

Full Text












FACTORS AFFECTING SERUM MINERAL LEVELS AND THE INFLUENCE
OF DIETARY CALCIUM AND PHOSPHORUS INTERRELATIONSHIPS
ON THE PERFORMANCE CHARACTERISTICS OF THE DOMESTIC HEN



















By

Edgar Reid Miller


A DISSERTATION PRESENTED TO THE GRADUATE COUNCIL OF
THE UNIVERSITY OF FLORIDA
IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE
DEGREE OF DOCTOR OF PHILOSOPHY





UNIVERSITY OF FLORIDA


1976
















ACKNOWLEDGEMENTS


The author is sincerely grateful to the chairman of his supervisory

committee, Dr. Henry R. Wilson, and to Dr. Robert H. Harms, chairman of

the Department of Poultry Science, for their most generous and learned

assistance in the planning and completion of this research. Very

special thanks is extended to the members of the author's supervisory

committee, Dr. R. A. Voitle, Dr. D. M. Janky, Dr. R. C. Littell and

Dr. J. A. Himes, for their aid during this course of study.

To all the technicians, farm staff and secretaries at the Depart-

ment of Poultry Science, the author extends his gratitude for their aid

in the various aspects of assembling this research.

The author extends his appreciation to the Cobb Breeding Corpora-

tion, Concord, Massachusetts, for supplying the broiler breeders and

to Hodges Poultry Farm, Inc., Callahan, Florida, for supplying the

Babcock B-300 pullets.

His deepest affection and appreciation goes to his wife, Ellen,

and son, Chris, whose unending support, encouragement, faith and love

made the attainment of this goal worthwhile.
















TABLE OF CONTENTS



Page

ACKNOWLEDGEMENTS . . ... ii

LIST OF TABLES . . ... v

LIST OF FIGURES . . ... vii

ABSTRACT . . ... . viii

INTRODUCTION . . ... . 1

LITERATURE REVIEW . ... 3

Egg Production . .. .. 3
Eggshell Quality . . 5
Hatchability . . ... 6
Blood Mineral Levels . . 7

CHAPTER I

CALCIUM AND PHOSPHORUS INTERRELATIONSHIPS AS THEY
AFFECT HATCHABILITY . .... ... 10

Procedure . ... 11
Results and Discussion . ... 13
Summary . . ... 17

CHAPTER II

CALCIUM AND PHOSPHORUS INTERRELATIONSHIPS AND THE
PERFORMANCE OF AGED LAYERS . ... .19

Procedure . . ... 19
Results and Discussion . ... 21
Summary . . ... 23

CHAPTER III

EFFECTS OF CONTINUAL FEED SUPPLEMENTATION VERSUS A
SINGLE LIQUID DOSE OF PHOSPHORUS ON SUBSEQUENT
PERFORMANCE OF PHOSPHORUS-DEPLETED HENS ... 26








TABLE OF CONTENTS (Continued)



Pag

Procedure . . 26
Results and Discussion .... . .. 27
Summary . . . 32

CHAPTER IV

CHANGES IN SERUM PHOSPHORUS OF LAYING HENS OVER A
24-HOUR PERIOD . . 33

Procedure . . 33
Results and Discussion . .. 36
Summary . . . 41

CHAPTER V

THE EFFECTS OF PRODUCTION STATUS ON SERUM CALCIUM AND
PHOSPHORUS . . .. .. 42

Procedure . . .. 42
Results and Discussion . .. 44
Summary . . 50

CHAPTER VI

SERUM CALCIUM AND PHOSPHORUS LEVELS IN RELATION TO
TIME OF OVIPOSITION . . ... 51

Procedure . .. .. .. 51
Results and Discussion . ... 53
Summary. . .... ... 60

CHAPTER VII

INFLUENCE OF VARIOUS DIETARY CALCIUM AND PHOSPHORUS
LEVELS ON THE PERFORMANCE OF BROILER BREEDER HENS 61

Procedure . . 61
Results and Discussion . ... 63
Summary . .. .. .. 68

SUMMARY .. .. .. . ... .. ... .... 69

REFERENCES . ..... 72

BIOGRAPHICAL SKETCH . . 78















LIST OF TABLES


Table Page

1 Composition of basal diet . ... 12

2 Hatchability (%) of fertile eggs from hens fed
various levels of calcium and phosphorus ... 14

3 Hatchability (%) of fertile eggs from hens fed
three levels of calcium and two levels of
phosphorus . ... 16

4 Hen-day egg production (%) of hens fed two levels
of calcium and phosphorus . ... 22

5 Hatchability (%) of fertile eggs from hens fed
two levels of calcium and phosphorus ... 24

6 Hen-day egg production and hatchability of
fertile eggs from hens on different dietary
treatments . . ... 29

7 Serum phosphorus and calcium levels of hens on
different dietary phosphorus treatments ... 31

8 Composition of experimental diet . ... 35

9 Serum phosphorus of laying hens at various
times . . ... . .37

10 Serum phosphorus levels of hens in different
production groups (Trials 1 and 2 combined) ... 45

11 Serum calcium levels of hens in different
production groups (Trials 1 and 2 combined) ... 49

12 Serum calcium and phosphorus in relation to
time of oviposition . ... 54

13 Composition of broiler breeder basal diet ... 62








LIST OF TABLES (Continued)


Table Pag

14 Performance of broiler breeder hens maintained
on litter as influenced by various dietary
phosphorus levels (cumulative across time and
calcium levels) . . ... 64

15 Performance of broiler breeder hens maintained
on litter as influenced by various dietary
calcium levels (cumulative across time and
phosphorus levels) . . ... 66















LIST OF FIGURES


Figure Pagi

1 Changes in the serum phosphorus level of laying
hens over a 24-hour period . 39

2 Serum phosphorus in relation to time of
oviposition . . 56

3 Serum calcium in relation to time of
oviposition . . 59









Abstract of Dissertation Presented to the Graduate Council
of the University of Florida in partial Fulfillment of the Requirements
for the Degree of Doctor of Philosophy



FACTORS AFFECTING SERUM MINERAL LEVELS AND THE INFLUENCE
OF DIETARY CALCIUM AND PHOSPHORUS INTERRELATIONSHIPS
ON THE PERFORMANCE CHARACTERISTICS OF THE DOMESTIC HEN

By

Edgar Reid Miller

August, 1976

Chairman: Henry R. Wilson
Major Department: Animal Science (Poultry)

A series of seven experiments was conducted on calcium and phospho-

rus nutrition of the laying hen. The experiments were designed to

observe

1) interactions between dietary calcium and phosphorus as related

to hatchability, egg production, shell quality and feed consumption;

2) changes in serum phosphorus levels associated with the laying

cycle; and

3) effects of production status and time, in relation to oviposi-

tion, on serum calcium and phosphorus levels.

The following parameters were measured:

1) egg production, on a hen-day basis;

2) hatchability of fertile eggs;

3) shell quality, as measured by specific gravity;

4) feed consumption; and

5) serum calcium and phosphorus levels as measured by atomic

absorption spectrophotometry and colorimetry, respectively.


viii









Reductions in hatchability of eggs from caged layers caused by

sub-optimal levels of supplemental inorganic phosphorus were signifi-

cantly offset by elevation of dietary calcium. When all phosphorus in

the diet of caged layers was supplied by plant sources, elevation of

dietary calcium resulted in significant decreases in egg production and

hatchability of fertile eggs. Phosphorus supplementation via an oral

liquid dose more rapidly offset the reduced hatchability and egg produc-

tion of phosphorus-depleted hens than phosphorus supplemented in the

feed.

The serum phosphorus level of caged layers changed considerably

during a 24-hour egg laying cycle, reaching a peak level at 7 a.m. and

a minimum level at 8 a.m. Serum phosphorus remained unchanged from

8 a.m. until 12 noon. A gradual increase in the level occurred from

12 noon until the peak was reached at 7 a.m. Rate of egg production

and time, in relation to oviposition, exerted more influence on levels

of serum phosphorus than on serum calcium.

Egg production of broiler breeder hens maintained on litter was

improved by supplementation of the diet with low levels of inorganic

phosphorus, while further supplementation resulted in a depression in

egg production. High levels of dietary phosphorus significantly de-

pressed shell quality, as measured by specific gravity. Alterations in

dietary calcium or phosphorus affected only minor changes in feed con-

sumption and hatchability of fertile eggs.















INTRODUCTION


Although there is little difference in physical appearance between

the laying hen of today and its ancestors, vast improvements in bird

performance have occurred since the beginning of domestication. In the

area of egg production alone, the average number of eggs produced per

hen per year in the United States has increased from 100 in 1937 to 231

in 1974 (Kalch, 1976). This increase in production has not only placed

great strain on the physiological systems of the bird, but has war-

ranted more exacting work in all phases of poultry research.

One area of investigation which has received considerable atten-

tion has been the relationship of dietary calcium and, to a lesser

degree, phosphorus to performance characteristics of the hen. The need

for these two minerals in the diet of laying hens has long been recog-

nized; however, in most of the work reported, they have been treated

independently, with effects of one mineral being tested at a set level

of the other. In addition, most mineral work relating to shell quality

has centered around calcium and its metabolism rather than phosphorus.

For the most part, investigations relating to phosphorus and the

laying hen have been concerned with establishment of dietary

requirements. Recent reports, however, indicate that phosphorus may be

more intimately involved with shell quality and interrelationships with

calcium than previously believed. For these reasons, more research

into the exact roles of phosphorus in the nutrition of the hen is needed.








The following experiments were designed to examine

1) interrelationships between calcium and phosphorus as they

affect hatchability of fertile eggs and egg production of caged layers;

2) changes in circulating serum phosphorus of the hen during the

laying cycle;

3) factors which affect serum calcium and phosphorus levels; and

4) interrelationships between calcium and phosphorus as related

to egg production, hatchability, shell quality and feed consumption of

hens maintained on litter.















LITERATURE REVIEW


Calcium, first isolated by Davy in 1808, and phosphorus, dis-

covered by Brandt in 1669, rank as two of the most essential minerals

for any living organism (Merck Index, 1968). Their physiological

functions include cellular energy reactions; skeletal, teeth and shell

formation; blood clotting and muscular contraction (Mountcastle, 1974).


Egg Production


It has long been established that supplemental calcium in some

form must be supplied in the diet of laying hens in order to maximize

egg production. Hens fed diets low in calcium resulted in decreased

egg production (Buckner and Martin, 1920; Buckner et al., 1925; Bloom

et al., 1960; Waldroup and Harms, 1961; Deobald et al., 1936; and

Mehring and Titus, 1964). Buckner et al. (1928) and Massengale and

Platt (1930) reported that egg production of hens depended not only on

the inclusion of calcium in the diet, but also on the chemical and/or

physical nature of the calcium included.

In addition to the necessity for supplemental calcium in the diet

to achieve maximum egg production, it is necessary to have the proper

amount of phosphorus (Norris et al., 1933; Miller and Bearse, 1934; and

Sanford and Alder, 1969). In a study of caged and floor maintained

layers, Harms et al. (1961) reported that the phosphorus requirement

for maximum egg production was between 0.40 and 0.50% of the diet (0.06








to 0.16% supplemental inorganic) for floor birds and approximately

0.60% (0.26% supplemental inorganic) for caged layers. A subsequent

report by Singsen et al. (1962) indicated that 0.53% total dietary

phosphorus was required for maximum production of birds maintained in

floor pens. These authors also reported that increasing the phosphorus

level above 0.53% resulted in a significant decrease in egg production.

Waldroup et al. (1974) reported that increasing supplemental phosphorus

above 0.10% in the diets of caged turkey hens resulted in significantly

improved egg production.

As with calcium, the form in which phosphorus is supplied in the

diet can markedly influence production. Waldroup et al. (1967) reported

that diets containing graded levels of phosphorus supplied by plant

sources did not support normal egg production of caged laying hens.

However, when supplemental inorganic phosphorus was added to the diet,

egg production was significantly increased. O'Rourke et al. (1954)

reported similar results utilizing semi-purified diets.

A revaluation by this author of previously reported data revealed

that calcium seemed to exhibit a "sparing" action on phosphorus when

the latter element was present in sub-optimal amounts. In data reported

by Harms et al. (1961) it appeared that when calcium was increased from

2.00 to 3.75% at a supplemental phosphorus level of 0.05%, egg produc-

tion was improved by 9.70 percentage points. An examination of the

data reported by Waldroup et al. (1974) revealed that with caged turkey

hens, an elevation in the dietary calcium level from 2.25 to 3.50% at

inorganic phosphorus levels of 0.10 and 0.20% resulted in improvements

in egg production of 27.90 and 10.70 percentage points, respectively.

Whether these results indicated a true "sparing" action or just a





5


calcium effect per se is not known; however, additional research on

this aspect could prove beneficial in terms of the possibility for re-

ducing supplemental phosphorus levels by elevation of the dietary

calcium.


Eggshell Quality


The shell quality of eggs, whether measured by specific gravity,

shell smoothness, shell thickness, breaking strength, shell deformation

or percentage of cracks has been reported to be influenced by dietary

calcium (Wolford and Tanaka, 1970). Several investigators have reported

that dietary calcium levels of less than 2.00% significantly depressed

the shell quality of chicken eggs (Deobald et al., 1936; Evans et al.,

1944; Hurwitz and Bar, 1966; Mehring, 1964; and Nevalainin, 1969).

Petersen et al. (1960) reported significant improvements in shell

quality at dietary calcium levels above 2.25%. They found that a level

of 3.38% calcium resulted in a highly significant increase in shell

quality as measured by specific gravity, while at levels of 4.50 and

5.25% calcium, they observed further improvements but on a reduced

scale. Similar results have been reported by several other investiga-

tors (Pepper et al., 1968; Reddy et al., 1968; Sullivan and Kingan,

1962; Mueller, 1961; Harms and Waldroup, 1961; and Harms, et al., 1961).

Conflicting results have been reported concerning the effects of

phosphorus on eggshell quality. Pepper et al. (1959) reported no re-

sponse in specific gravity when the dietary phosphorus level was in-

creased from 0.37 to 0.47%. Petersen et al. (1960) found that various

phosphorus levels had no effect on shell quality. Walter and Aitken

(1962) fed levels of 0.40 to 0.70% phosphorus over a 44-week period and









found that there was no significant influence on specific gravity. It

has also been reported that dietary phosphorus levels did not influence

shell quality as measured by shell weight per unit of surface area

(Hurwitz and Bornstein, 1963; and Ademonsun and Nikolaiczuk, 1966).

Arscott et al. (1962) reported a significant decrease in egg

specific gravity when total dietary phosphorus was increased from 0.60

to 0.90% at a 2.25% calcium level. Singh et al. (1971) reported simi-

lar results for eggs from caged layers receiving total dietary phospho-

rus levels above 0.40%. Damron et al. (1974) reported that after 4 and

7 months production, specific gravity of caged layers decreased as the

dietary phosphorus level increased.


Hatchability


O'Rourke et al. (1954) reported depressed hatchability of fertile

eggs from caged laying hens fed a semi-purified basal diet containing

0.19% phosphorus. Supplementation of the basal diet with monocalcium

phosphate to provide a level of 0.30% total phosphorus, of which 0.18%

was non-phytin, resulted in significant improvements in hatchability.

The apparent requirement of 0.30% total phosphorus for maintenance of

normal hatchability reported by O'Rourke et al. (1954) was much lower

than that reported by Singsen et al. (1962). The latter authors found

that hens on wire floors fed practical laying diets required at least

0.40% total phosphorus (0.35% non-phytin) in order to support normal

hatchability. Harms et al. (1964) reported that supplementation of a

practical corn-soybean meal laying diet with 0.35% inorganic phosphorus

resulted in a highly significant increase in hatchability of fertile

eggs. Similar results were found by Waldroup et al. (1967), who








reported that various levels of organic plant phosphorus did not support

normal hatchability; however, supplementation of the diet with various

levels of inorganic phosphorus did result in significant improvements

in hatchability. These authors concluded that diets containing a level

of 0.19% non-phytin phosphorus was necessary for maximum hatchability.

In a study involving turkey breeders, Sewell et al. (1972) reported

that hatchability was significantly decreased when all inorganic phos-

phorus was removed from the diets. A subsequent study by Ferguson et al.

(1974) indicated that supplementation of turkey breeder diets with in-

organic phosphorus significantly improved hatchability.

Potter et al. (1974) reported that increasing the dietary calcium

level from 0.99 to 1.77% resulted in significant improvements in the

hatchability of fertile turkey eggs. Jensen et al. (1963) found that

increasing the calcium level above 1.75% in turkey breeder diets re-

sulted in a significant depression in hatchability. Balloun and Miller

(1964) reported a similar depression in hatchability when turkey

breeders were fed diets containing 3.00% calcium. These reports of

depressed hatchability at high dietary calcium levels were not substan-

tiated in studies conducted by Potter et al. (1974).

Several authors have reported that high dietary calcium levels (up

to 6.00% of the diet) had no effect on hatchability of fertile chicken

eggs' (Gutowska and Parkhurst, 1942; MacIntyre et al., 1963; and Mehring,

1965).


Blood Mineral Levels


Several investigators have reported that prior to the onset of egg

production there is a large increase in the circulating serum calcium











level of pullets (Halnan, 1925; Hughes et al., 1927; Buckner et al.,

1930; and Harshaw et al., 1934). It is in general agreement by most

authors that the observed rise in serum calcium is confined to the non-

diffusible (or protein bound) calcium fraction (Russell et al., 1930;

Heller et al., 1934; and Greenberg et al., 1936). Taylor and Russell

(1935) reported that the diffusible fraction of serum calcium of laying

and non-laying hens was 5.4 and 5.3 mg.%, respectively. However, they

found that the non-diffusible calcium fraction in the serum rose from

6.4 mg.% for non-laying hens to 16.1 mg.% for laying hens. Similar

results were reported by Greenberg et al. (1936). Feinberg et al. (1937)

found that the calcium level in laying and non-laying hens was rela-

tively constant during the 26-hour period of a single egg cycle.

Deobald et al. (1938) reported that the blood calcium level of laying

hens did not exhibit any marked variations during an entire egg cycle.

This is in contrast to results reported by Roland et al. (1972), who

found that serum calcium was significantly lower 4 hours before and

after oviposition. These authors also reported that calcium intake and

fecal calcium patterns were reversed from that observed for serum

calcium.

In analyses of serum samples from laying and non-laying hens,

Feinberg et al. (1937) found that a marked rise in the inorganic phos-

phorus level of serum occurred in laying hens, but that the phosphorus

level of non-laying hens remained constant. Peterson and Parrish (1939)

reported that serum phosphorus reached a peak level approximately 11.5

to 13.0 hours after oviposition of the previous egg, and that the lowest

level was observed immediately following oviposition. Paul and

Snetsinger (1969) found similar serum phosphorus patterns; however, they








reported that the peak phosphorus level in the serum occurred 16.0 hours

after oviposition of the previous egg.

Roland et al. (1973) reported that serum calcium was significantly

lower in hens receiving dietary calcium levels of 0.05% than in hens

receiving 3.00% calcium. Hurwitz and Griminger (1962) observed that

plasma phosphorus concentrations also reflected the dietary mineral

intake. Garlich et al. (1975) reported that the serum phosphorus of

hens fed diets containing 0.39% total phosphorus was 2.87 mg.% while

hens receiving 0.64% dietary phosphorus during the same period exhibited

serum phosphorus levels of 4.45 mg.%.

















CHAPTER I

CALCIUM AND PHOSPHORUS INTERRELATIONSHIPS
AS THEY AFFECT HATCHABILITY


O'Rourke et al. (1954) reported that a phosphorus deficiency in

the hen resulted in a significant decrease in hatchability of fertile

chicken eggs. These findings were corroborated in subsequent studies

by Harms et al. (1964) and Waldroup et al. (1967).

Maynard and Loosli (1969) stated that a great excess of either

calcium or phosphorus interferred with the absorption of the other

mineral. They concluded that a certain calcium:phosphorus ratio was

desirable for best absorption. In a review of the importance of

calcium:phosphorus ratios for chickens, Harms (1971) reported that, as

the age of the bird increased, the necessity for strict adherance to a

set calcium:phosphorus ratio decreased.

A revaluation of data published by Harms et al. (1961) and

Waldroup et al. (1974) indicated that with chickens and turkeys,

respectively, egg production was improved by elevating the dietary

calcium level at sub-optimal levels of supplemental inorganic

phosphorus. Thus, when the diet was deficient in phosphorus, a widen-

ing of the calcium:phosphorus ratio resulted in improved egg production.

The following study was designed to determine whether the same relation-

ship affects the hatchability of fertile eggs.








Procedure


Five hundred sixty 28-week-old Babcock B-300 pullets were randomly

assigned to 14 treatment groups of 40 birds each. Each group was

divided into four replicates of 10 birds each.

Twelve of the 14 treatment groups were arranged in a 2 X 6 facto-

rial design and fed diets containing either 2.25 or 3.50% calcium, with

0.35. 0.40. 0.45. 0.55, 0.70 or 1.40% total phosphorus. The two remain-

ing treatment groups were fed diets containing 6.00% calcium and either

0.45 or 0.70% total phosphorus. All birds were maintained in individual

20.3 X 45.7 cm. wire cages and received feed ad libitum for a 38-week

experimental period. The birds also received natural and/or artificial

light from 4:30 a.m. to 7:30 p.m. each day.

A corn-soybean meal basal diet was used which was calculated to

contain 0.09% calcium and 0.29% phosphorus (Table 1). A 17.02% portion

of the basal diet was composed of ground limestone, dicalcium phosphate

and white builder's sand in proportions required to provide the calcium

and phosphorus levels in each experimental diet. All diets were iso-

caloric and isonitrogenous.

After receiving the experimental diets for 19, 21, 26 and 38 weeks,

one-half of the birds from each treatment group were inseminated with

pooled semen from Cobb broiler breeder males. Pooled semen was used to

eliminate possible variation between individual males. Eggs were col-

lected during the second through the eighth days following the insemina-

tion and incubated. All eggs were set in Jamesway 252 incubators

maintained at 37.50 C. and 62% relative humidity. Fertility was deter-

mined by candling after seven days of incubation.
















Table 1. Composition of basal diet


Ingredient


Percent of the diet


Yellow corn meal

Soybean meal (50%)

Alfalfa meal (20%)

Animal fat

Microingredients1


54.10

21.50

2.00

4.50

0.50

0.03

0.35

17.02


DL-Methionine

Iodized Salt

Variable


Crude protein 15.5%

Metabolizable energy 2805 kcal./kg.

Calcium 0.09%

Phosphorus 0.29%


Supplied per kg. of diet: 6000 I.U. vit. A.; 2200 I.C.U. vit. D3;
2.2 mg. menadione dimethylpyrimidinol bisulfate; 4.4 mg. riboflavin;
13.2 mg. pantothenic acid; 39.6 mg. niacin; 499.4 mg. choline
chlorida; 22 mcg. vit. B12; 125 mg. ethoxyquin; 50 mg. manganese;
50 mg. iron; 6 mg. copper; 0.198 mg. cobalt; 1.1 mg. iodine; 35 mg.
zinc.








Hatchability data from each insemination period was subjected to

an analysis of variance procedure (Snedecor, 1956) in order to deter-

mine if treatment differences and interactions existed. Significant

differences among treatment means for phosphorus levels at each calcium

level were determined using regression analyses. Student's "t" tests

was used to determine significant differences between treatment means

for calcium levels at each phosphorus level.


Results and Discussion


In the first insemination period (19 weeks on experiment), numeri-

cal increases in hatchability were observed at the 2.25% calcium level

at phosphorus levels above 0.40%, and at the 3.50% calcium level when

phosphorus was increased above 0.35% (Table 2). At the 0.40% phosphorus

level, hatchability was increased by 10.3 percentage points by increas-

ing the dietary calcium from 2.25 to 3.50%. Although these differences

were quite large numerically, they were not statistically significant.

In the second insemination period (21 weeks on experiment), there

was a significant increase in hatchability for each increment increase

in phosphorus from 0.35 to 0.55% at the 2.25% calcium level (Table 2).

Hatchability was significantly increased when the calcium level in the

diet was elevated from 2.25 to 3.50% at the 0.35, 0.40 and 0.45% phos-

phorus levels. Thus, at the lower phosphorus levels, a significant

improvement in hatchability was observed when the calcium:phosphorus

ratio was widened.

Hatchability of fertile eggs in the third insemination period (26

weeks on experiment) was significantly increased at the 2.25% calcium

level when phosphorus was increased above 0.35% (Table 2). Although





14






0 0C CN C) C a' N-





ct
o4 CL r- 00 c)i 0 fi -0


U)
0o '
4- -4




c 00 01% 0o 01l m1 a 4- ai)
5d Ir) *l-SO








O-4 0 co
0 0 <0 (C
0n X) 0 8-H1
MSl 0(: l C



SC 0

e i* .j
0 N- a' a' a' a' a' C n








L0 00



O CH -. C)
UU U O *U IC
o )4 0 Ca C)
0 C 0 V
M U Cq c0 r- C -H C) VN u




cU a) a)




HH 0 4 o
0 I' 0 *C 0 H
0 UH 0 4 CO
4*H ,H X X X *HU -HO
S 0 N C O OC a) -zr 0 a) -HO r!O
O U *r- 0 *U VHC


Q) 4)-H
CO O- U *E

CO 0J0 *n
O L... rLOd 00 r- O*






S m0 4-H
-H .-N C) rn Ln cc H ro 'oo n)-H









O 4 CC cc a a j a




Go 01 43 ) rL co o *r-4 e
0 -H U) *i-l -H O













oo a) c*
6l mI I p Q M i





'- MC a' CC C) c) oi c3 cu CO




H On a a) mo



&0 O > OU
c 0 '-a 0 0
.f a) 0) C
N *O L L -I *M
* 4 0 Q0 &-4

- Ln o n 0 0
cT U a
0 cwo 4J 0) c
1 .0. 0C 4-

N I u n o oC C o tr I C C -H
O^-^ i- l -r -L OO ) C -

*rl HO E u o
HQ n 0








hatchability was improved numerically by elevating the phosphorus above

0.35% at the 3.50% calcium level, this increase was not statistically

significant. Elevating the calcium level from 2.25 to 3.50% at the

lowest phosphorus level resulted in a significant improvement in

hatchability.

In the fourth insemination period (38 weeks on experiment),

hatchability results indicated no significant increases when phosphorus

was elevated at either calcium level (Table 2). However, widening the

calcium:phosphorus ratio at the lowest phosphorus level, once again

resulted in a significant improvement in hatchability.

As evidenced from the data (Table 2), significant improvements in

hatchability by elevation of the calcium level at the 0.40 and 0.45%

phosphorus levels were not observed in insemination periods three and

four as they were in period two. An elevation of hens' performance

records (data not shown) indicated that a large number of birds exhibit-

ing low hatchabilities in period two had ceased production shortly

thereafter; therefore, subsequent hatchability data could not be

obtained.

Hatchability of fertile eggs from those birds fed diets containing

6.00% calcium and either 0.45 or 0.70% total phosphorus was comparable

to the hatchability of eggs from birds receiving the same phosphorus

levels at 2.25 and 3.50% calcium (Table 3). These data indicated that

a further widening of the calcium:phosphorus ratio was neither benefi-

cial nor detrimental to hatchability.

Results from this experiment supported the previous findings

(Waldroup et al., 1967) that a phosphorus deficiency caused a depression

in hatchability of fertile eggs; however, it appeared that increased























Table 3. Hatchability (%) of fertile eggs from hens fed three levels
of calcium and two levels of phosphorus


Insemination periods
Calcium Phosphorus (weeks of experiment)
S(%) 1 2 3 4
(19 weeks) (21 weeks) (26 weeks) (38 weeks)

6.00 0.45 100.0 94.0 97.2 93.3

0.70 95.5 98.6 91.7 98.9

3.501 0.45 98.8 95.2 97.8 94.0

0.70 97.6 97.0 97.3 93.9

2.251 0.45 98.0 85.0 98.7 92.2

0.70 92.6 98.4 96.1 95.2


Data taken from Table 2.








dietary calcium would spare phosphorus under certain phosphorus defi-

cient conditions. This was in contrast to the widely accepted concept

that increasing the calcium in a phosphorus deficient diet would aggra-

vate the phosphorus deficiency. In this experiment, a widening of the

calcium:phosphorus ratio at the 0.35% dietary phosphorus level resulted

in a significant improvement in the hatchability of fertile eggs.


Summary


An experiment was conducted to study the influence of various

dietary calcium and phosphorus levels on hatchability of fertile eggs.

Four replicates of ten individually caged Babcock B-300 hens received

one of twelve diets containing 2.25 or 3.50% calcium, with 0.35, 0.40,

0.45, 0.55, 0.70 or 1.40% total phosphorus. Two other diets contained

6.00% calcium and 0.45 or 0.70% total phosphorus.

Two replicates from each treatment were inseminated after the

birds had been on the experimental diets for 19, 21, 26 and 38 weeks.

Results from the first insemination period indicated numerical increases

in hatchability at the 2.25% calcium level when phosphorus was increased

above 0.40%, and at the 3.50% calcium level when phosphorus was in-

creased above 0.35%. In the second and third insemination periods,

hatchability was significantly improved at the 2.25% calcium level when

phosphorus was increased above 0.35%. Hatchability was significantly

improved by elevating the calcium level from 2.25 to 3.50% at the 0.35,

0.40 and 0.45% phosphorus levels in the second insemination period.

Significant improvements in hatchability also occurred in the third and

fourth insemination periods when calcium was elevated from 2.25 to 3.50%

phosphorus level. No significant differences in hatchability were





18


observed for hens receiving the 6.00% calcium and 0.45 or 0.70% phospho-

rus diets.

















CHAPTER II

CALCIUM AND PHOSPHORUS INTERRELATIONSHIPS
AND THE PERFORMANCE OF AGED LAYERS


In a previous experiment (Chapter I) hatchability of fertile eggs

was significantly improved by elevating the dietary calcium level from

2.25 to 3.50% when the diet contained 0.35% total phosphorus. Of the

0.35% phosphorus level used in the experiment, 0.29 percentage points

were supplied by plant sources and 0.06 percentage points by supplemen-

tal inorganic phosphorus. Based on the results obtained, it was con-

cluded that calcium would spare phosphorus when the latter element was

supplied at sub-optimal levels of inorganic phosphorus. The following

experiment was designed to determine if this same relationship exists

when the total phosphorus in the diet was supplied by plant sources.


Procedure


At the conclusion of the 38-week experiment outlined in Chapter I,

hens still in production which had been receiving diets containing 2.25

or 3.50% calcium and 0.35, 0.40, 0.45, 0.55 or 1.40% total phosphorus,

were selected for use in the present experiment. This resulted in 10

groups of 30 hens per group, with each of the groups being identified

by its previous dietary treatment. Each of the groups was further

divided into six replicates of five birds per replicate.

Eight of the 10 groups were assigned diets containing either 2.25

or 3.50% calcium and 0.29% total phosphorus. The two remaining groups,








which had received 2.25 or 3.50% calcium and 0.55% total phosphorus in

the previous experiment, were maintained on these same diets and used

as controls.

A corn-soybean meal basal diet calculated to contain 0.09% calcium

and 0.29% phosphorus was used (Table 1). A 17.02% portion of the basal

diet was composed of ground limestone, dicalcium phosphate and white

builder's sand in proportions required to provide the calcium and phos-

phorus levels in each experimental diet. All diets were kept ioscaloric

and isonitrogenous.

All birds were maintained in individual 20.3 X 45.7 cm. wire cages

and received the diets ad libitum for a 12-week experimental period.

The birds received natural and/or artificial light from 4:30 a.m. to

7:30 p.m. each day.

Egg production was recorded on a hen-day basis for three consecu-

tive 28-day periods following the start of the experiment. During the

second and third 28-day periods, all hens were inseminated on two con-

secutive days with pooled semen from Cobb broiler breeder males. Eggs

were collected from the second through the eighth days following the

inseminations and incubated. All eggs were incubated in Jamesway 252

incubators maintained at 37.50 C. and 62% relative humidity.

By analyses of variance, it was determined that previous dietary

treatment did not significantly affect egg production or hatchability;

therefore, data were combined for all birds receiving 0.29% phosphorus

within each calcium level. The combined data were subjected to analyses

of variance for 2 X 2 factorial experiments. Significant differences

between main or simple effects were determined with "F" tests (Steel

and Torrie, 1960).








Results and Discussion


During the first 28-day period, hen-day egg production was signifi-

cantly improved at the 2.25% calcium level when the diet was supple-

mented with 0.26% inorganic phosphorus (0.55% total phosphorus, Table 4).

No effect on production was observed at the 3.50% calcium level. In the

second and third 28-day periods, significant improvements in egg produc-

tion were observed at both calcium levels when dietary phosphorus was

increased.

Elevating the dietary calcium level from 2.25 to 3.50% at the 0.29%

phosphorus level resulted in a significant improvement in egg production

in the first 28-day period. However, in the second period, production

was not affected by altering the calcium content of the diet. In the

third period, elevating the calcium level at the all plant phosphorus

level (0.29%) resulted in a significant lowering of egg production,

while increasing the calcium level at the 0.55% phosphorus level re-

sulted in a significant improvement in egg production.

The exact reason for the reversal in effects on egg production ob-

served when calcium was increased at the 0.29% phosphorus level is not

known. It is suggested that when total dietary phosphorus was supplied

by plant sources, an elevation in the dietary calcium resulted in a de-

crease in phosphorus utilization from phytate phosphorus complexes. It

is hypothesized that this decreased phosphorus utilization resulted in

an increase in bone demineralization to provide phosphorus for cellular

energy reactions, yolk phosphorus supplies and, to a small degree, egg-

shell formation. Bone demineralization probably continued until such

time that phosphorus became limiting in cellular energy reactions, at






















Table 4. Hen-day egg production (%)
and phosphorus


of hens fed two levels of calcium


Calcium (%)

Total 28-day periods1
phosphorus
(%) 1 2 3

2.25 3.50 2.25 3.50 2.25 3.50


0.29 47.3Ax 59.1 32.3A 29.1A 32.5Ax 24.6A

0.55 63.2B 59.6 51.9B 55.1B 58.4Bx 64.8


Within each


28-day period:


The capital superscripts designate significant (p < 0.05)
changes in egg production due to the phosphorus level, as
determined by "F" tests.

The lower case superscripts designate significant (p < 0.05)
changes in egg production at any one phosphorus level when
calcium is increased from 2.25 to 3.50%, as determined by
"F" tests.










which point, the hen reduced egg production in order to maintain the

energy reactions.

Supplementing the diet with 0.26% inorganic phosphorus resulted in

significant increases in the hatchability of fertile eggs at both

calcium levels (Table 5). Hatchability of fertile eggs was signifi-

cantly depressed at the 0.29% total phosphorus level in both periods

when calcium was elevated from 2.25 to 3.50% of the diet. This was in

direct contrast to the results observed in Chapter I (Table 2), where

it was found that when diets contained sub-optimal levels of inorganic

phosphorus, hatchability was significantly improved by elevating the

calcium level from 2.25 to 3.50%.

As with egg production, it is hypothesized that the phosphorus

absorbed from the intestine and liberated from the bones was preferen-

tially utilized for physiological functions at the expense of supplying

phosphorus to the yolk. The amount of phosphorus that was supplied to

the yolk was probably deficient in terms of quantity required for full

embryo growth. It appeared that hatchability was more sensitive than

egg production to elevations in dietary calcium levels at the all plant

phosphorus level (0.29%); therefore, hatchability seemed to be a more

critical measurement of a phosphorus deficiency in the hen than egg

production.


Summary


An experiment was conducted to examine the effects of elevating

the calcium level from 2.25 to 3.50% in diets containing either 0.29%

phosphorus (from plant sources) or 0.55% phosphorus (from plant and

supplemental inorganic sources). Data indicated that elevating the






















Table 5. Hatchability (%) of fertile
calcium and phosphorus


eggs from hens fed two levels of


Calcium (%)

Total 28-day periods1'2
phosphorus
(%) 2 3

2.25 3.50 2.25 3.50 2.25 3.50


0.29 ---- ---- 74.2A 50.4Ay 62.2A 41.2A

0.55 ---- -- 83.6Bx 97.0By 84.0B 87.9B


Within each 28-day period:


The capital superscripts designate
changes in hatchability due to the
determined by "F" tests.


significant (p < 0.05)
phosphorus level, as


The lower case superscripts designate significant (p < 0.05)
changes in hatchability at any one phosphorus level when
calcium is increased from 2.25 to 3.50%, as determined by
"F" tests.
2Hatchability was not measured during the first 28-day period.
Hatchability was not measured during the first 28-day period.





25


calcium level in diets where all phosphorus was supplied by plant

sources resulted in significant reductions in hen-day egg production

and hatchability of fertile eggs.















CHAPTER III

EFFECTS OF CONTINUAL FEED SUPPLEMENTATION VERSUS A
SINGLE LIQUID DOSE OF PHOSPHORUS ON SUBSEQUENT
PERFORMANCE OF PHOSPHORUS-DEPLETED HENS


Egg production and hatchability of fertile eggs from hens receiving

total dietary phosphorus supplied by plant sources have been shown to be

lower than those of birds receiving supplemental inorganic phosphorus

(Chapter II). The following experiment was designed to examine the rela-

tive effectiveness of continual feed supplementation versus a single

liquid dose of phosphorus on the subsequent performance of phosphorus-

depleted hens.


Procedure


At the conclusion of the experiment outlined in Chapter II, all

birds which had received the diet containing 3.50% calcium and 0.29%

total phosphorus were randomized into three groups of 40 birds per group

and were continued on the same diet. Each group was further divided

into eight replicates of five birds per replicate. The first group

(negative control) received no supplemental phosphorus. The diet of the

second group was supplemented with phosphorus supplied by 1.0 N phos-

phoric acid in an amount calculated to provide an additional 100 mg.

phosphorus/bird/day. Each hen in the third group received a 10 ml. oral

dose of 1.0 N phosphoric acid at 8 a.m. on each of 14 consecutive days

following initiation of the experiment. This oral dosing provided










100 mg. phosphorus/bird/day. The hens which had received the 3.50%

calcium, 0.55% total phosphorus diet in Chapter II were maintained on

that same diet to act as positive controls.

Egg production was recorded on a hen-day basis for two weeks fol-

lowing initiation of the experiment. All hens were inseminated two

days prior to the start of the experiment, and again after they had

received a dietary treatment for seven days. All eggs were saved and

incubated at the end of each week of the experiment.

At the end of the two weeks, 5 ml. blood samples were taken at

11 a.m. from five hens in each of the four treatment groups. Samples

were taken only from those hens which had laid an egg between 8 a.m.

and 11 a.m. on the day of blood collection. Each blood sample was

taken with a disposable 5 ml. syringe equipped with a 3.8 cm., 20 guage

needle. Immediately after collection, each sample was transferred to a

7 ml. silicone coated tube and allowed to clot. After the blood had

clotted, serum was separated via centrifugation for 15 minutes at 2,500

r.p.m. One ml. serum samples were deproteinized with trichloroacetic

acid and the phosphorus content determined using the colorimetric method

of Fiske and Subharow (1925). Serum calcium was assayed using a Perkin-

Elmer 290B Atomic Absorption Spectrophotometer.

Data were subjected to analyses of variance as described by

Snedecor (1956). Significant differences among treatment means were

established using the multiple range test described by Duncan (1955).


Results and Discussion


Hen-day production of birds receiving the basal diet supplemented

with phosphorus from phosphoric acid was 4.4 percentage points above








that of the negative controls (Table 6). Although this difference is

not statistically significant, it is believed that it did represent a

trend toward an improvement in production. When 100 mg. phosphorus/

bird/day was given via an oral liquid dose, egg production was signifi-

cantly higher than both the negative controls and the feed supplemented

hens. The birds receiving the 3.50% calcium, 0.55% total phosphorus

diet (positive controls) had the highest egg production (Table 6).

Hatchability of the first week's eggs from hens receiving the nega-

tive control diet was 56.8% (Table 6). When phosphorus was supplemented

in the feed in the form of phosphoric acid, hatchability increased to

70.8%. Liquid dosing of phosphorus resulted in a significant increase

in hatchability over the negative control group, and an 11.5 percentage

point increase over eggs from birds receiving the phosphoric acid sup-

plemented feed. Eggs from hens fed the positive control diet had the

highest hatchability of all groups.

Hatchability of the second week's eggs indicated that all groups

had a significantly higher hatchability of fertile eggs than the birds

receiving the negative control diet (Table 6). However, eggs from the

hens fed the phosphoric acid supplemented feed had the highest hatcha-

bility during this period. Analysis of feed consumption data for this

group revealed that the birds were actually consuming an additional

132 mg. phosphorus/bird/day rather than the 100 mg. phosphorus/bird/day

projected at the beginning of the experiment. It is probable that the

increased intake of phosphorus above that given the oral dosed hens

resulted in the high hatchability observed. It is also possible that

the length of time for phosphorus absorption was greater for the feed




















Table 6. Hen-day egg production and hatchability of fertile eggs from
hens on different dietary treatments


1 Hen-day Hatch of fertile eggs (%)2
Treatment Hen-day 2
egg production (%)
First week Second week


A 23.7a 56.8a 60.5a

B 28.1a 70.8ab 92.2b

C 41.4b 82.3bc 79.7b

D 60.8c 91.8c 86.9b


A = Hens receiving 3.50% calcium and 0.29% phosphorus
control diet).

B = Hens receiving the negative control diet plus 132
tal phosphorus/bir/day in the feed.


(negative


mg. supplemen-


C = Hens receiving the negative control diet plus 100 mg. supplemen-
tal phosphorus/bird/day supplied as an oral liquid dose of dilute
phosphoric acid.

D = Hens receiving 3.50% calcium and 0.55% phosphorus (positive
control diet).

Means in the same column not having common superscripts are signifi-
cantly different (p < 0.05), as determined by Duncan's multiple range
test.








supplemented birds than for the dosed birds resulting in a more effi-

cient phosphorus utilization and an increased hatchability.

Serum phosphorus values at 11 a.m. were significantly lower for

birds on the negative control diet (Table 7). Birds receiving the posi-

tive control diet and the phosphoric acid supplemented diet had serum

phosphorus levels of 3.98 and 3.96 mg.%, respectively. Birds given the

oral liquid dose of phosphoric acid had the highest serum phosphorus

level (4.93 mg.%). It is probable that since the phosphorus given in

the liquid dose was already in an ionized state, absorption proceeded

at a more rapid rate than for the other diets, resulting in a higher

serum phosphorus level.

The serum calcium level of birds receiving the negative control

diet was 22.5 mg.% at 11 a.m. (Table 7). This level was significantly

lower than those observed for birds receiving any of the other dietary

treatments. No significant differences among serum calcium levels

existed for the other three treatments (Table 7). It is suggested that

the low levels of serum calcium and phosphorus of the negative control

birds was a result of utilization of these minerals for bone

remineralization. It is further suggested that although calcium and

phosphorus were utilized for bone rebuilding by birds in the other

treatment groups, the quantities supplied by the diets were greater,

thus resulting in the maintenance of higher serum mineral levels.

In cases of a phosphorus deficiency, oral liquid dosing and feed

supplementation of phosphorus could be used in combination to rapidly

improve the reduced egg production and hatchability.





















Table 7. Serum phosphorus and calcium levels of hens on different
dietary phosphorus treatments


1 Serum phosphorus Serum calcium
Treatmen(mg.% S.E.)2 (mg.% S.E.)2


A 0.38 + 0.20a 22.5 + 0.35a

B 3.96 + 0.32b 27.9 + 1.60b

C 4.93 0.60c 28.6 + 2.00b

D 3.98 0.12b 30.3 + 0.96b


A = Hens receiving 3.50% calcium and 0.29% phosphorus (negative
control diet).

B = Hens receiving the negative control diet plus 132 mg. supplemen-
tal phosphorus/bird/day in the feed.

C = Hens receiving the negative control diet plus 100 mg. supplemen-
tal phosphorus/bird/day supplied as an oral liquid dose of dilute
phosphoric acid.

D = Hens receiving 3.50% calcium and 0.55% phosphorus (positive
control diet).

2Means in the same column not having common superscripts are signifi-
cantly different (p < 0.05), as determined by Duncan's multiple range
test. S.E. = standard error of the mean.





32


Summary


An experiment was conducted to examine the effects of supplemental

phosphorus in the feed versus oral liquid dosing on the subsequent

performance of aged, phosphorus-depleted hens. Results indicated that

liquid dosing of supplemental phosphorus was more efficient in offset-

ting reduced egg production and hatchability of fertile eggs than pro-

viding relatively the same amount of phosphorus in the feed.















CHAPTER IV

CHANGES IN SERUM PHOSPHORUS OF LAYING HENS
OVER A 24-HOUR PERIOD


Feinberg et al. (1937) reported that the serum phosphorus level

of laying hens exhibited a marked rise during the period of shell

calcification. It was later reported by Peterson and Parrish (1939)

that serum phosphorus reached its peak level approximately 11.5 to 13

hours after oviposition of the previous egg, and that the lowest level

was observed immediately following oviposition. Paul and Snetsinger

(1969) reported results similar to those of Peterson and Parrish (1939);

however, they found that the peak phosphorus level occurred 16 hours

after oviposition.

In each of these studies, all birds were bled at each collection

time. The authors made no correction for possible changes in serum

mineral levels due to the stress of bleeding or time of oviposition.

Also, sampling during the laying cycle was limited to three or four

periods. The following experiment was designed to examine the changes

in serum phosphorus at several intervals during the laying cycle.


Procedure


Three trials were conducted to establish the cyclic changes in

serum phosphorus of laying hens. In Trial 1, blood samples were col-

lected at 6 p.m., 12 midnight, 6 a.m. and 12 noon. In order to more

precisely observe the changes in serum phosphorus during the day,








Trial 2 was conducted so that birds were bled at two-hour intervals

from 6 a.m. to 6 p.m. Trial 3 was designed to measure serum phosphorus

levels at 12 midnight, 4 a.m. and 7 a.m.

In all three trials, Babcock B-300 laying hens, 63 weeks of age,

were selected for blood sampling. All birds were maintained in indi-

vidual 20.3 X 45.7 cm. wire cages and fed a corn-soybean meal laying

diet calculated to contain 16.2% protein, 3.25% calcium, 0.75% total

phosphorus and 2850 kcal. metabolizable energy per kilogram of feed

(Table 8). The birds received natural and/or artificial light from

4:30 a.m. until 7:30 p.m. each day.

The day prior to each blood collection hens which laid an egg be-

tween 8 a.m. and 12 noon were identified and randomized into groups of

15 birds. Each group of birds was assigned to a specific blood collec-

tion time. The following day, 5 ml. of blood were collected via

anterior heart puncture from each bird in the group assigned to a

specified collection time. By using this procedure, the necessity for

repeated bleeding of any individual bird was eliminated. Blood samples

were used only from those birds which laid an egg prior to 12 noon the

day of blood collection. Therefore, only blood samples from the se-

lected population of birds which laid an egg the day prior to, and the

day of, bleeding were used for serum phosphorus determinations. The

same procedures for blood collection, blood clotting, serum extraction,

deproteinization and serum phosphorus determinations were used as out-

lined in Chapter III. The mean serum phosphorus value for each time

period in each trial was determined and the standard error of the mean

calculated using the procedures described by Snedecor and Cochran (1973).

















Table 8. Composition of experimental diet


Ingredient


Percent of the diet


Yellow corn meal

Soybean mean (50%)

Alfalfa mean (20%)

Ground limestone (38% Ca.)

Dyna Phos (18.5% P., 24% Ca.)

Iodized Salt

Microingredients1


68.52

19.00

2.50

6.88

2.25

0.35

0.50


Crude protein 16.2%

Metabolizable energy 2805 kcal./kg.

Calcium 3.25%

Phosphorus 0.75%


1
Supplied per kg. of diet: 6000 I.U. vit. A.; 2200 I.C.U. vit. D3;
2.2 mg. menadione dimethylpyrimidinol bisulfate; 4.4 mg. riboflavin;
13.2 mg. pantothenic acid; 39.6 mg. niacin; 499.4 mg choline
chloride; 22 mcg. vit. B12; 125 mg. ethoxyquin; 50 mg. manganese;
50 mg. iron; 6 mg. copper; 0.198 mg. cobalt; 1.1 mg. iodine; 35 mg.
zinc.








Regression analyses were used to construct the two components of the

serum phosphorus cycle (Steel and Torrie, 1960).


Results and Discussion


The serum phosphorus level of actively laying hens was not stable

but underwent considerable change through a 24-hour period (Table 9).

In Trial 1, the serum phosphorus level increased from 5.35 mg.% at

6 p.m. to 5.78 mg.% at 6 a.m. At 12 noon, it had decreased markedly to

4.89 mg.%. Since blood samples were collected at six-hour intervals in

this trial, the duration of decreased serum phosphorus could not be

ascertained. In Trial 2, the serum phosphorus level was 5.82 mg.% at

6 a.m., and at 8 a.m. had fallen to 4.88 mg.% where it remained within

0.02 mg.% of this value until 12 noon. The level had increased to 5.29

mg.% at 2 p.m. and to 5.42 mg.% at 6 p.m. In Trial 3, serum phosphorus

increased from 5.53 mg.% at 12 midnight to 6.08 mg.% at 7 a.m.

A plot of the data for the three trials (Figure 1) indicated that

the curve of the phosphorus cycle had two components. Regression

analyses to give the "best fit" line for each component were used. The

"r" values for components A (8 a.m. 12 noon) and B (12 noon 7 a.m.)

were +0.189 and +0.931, respectively.

The cycle in serum phosphorus might be expected since changes in

blood calcium and alkaline phosphatase occur during the laying cycle

(Taylor and Williams; 1964; Taylor et al., 1965,; Paul and Snetsinger,

1969; and Roland et al., 1972).

Since the populations of birds used in these trials were selected

for an oviposition time of 10 a.m. 2 hours, and active shell calcifi-

cation begins 4 to 5 hours after oviposition of the previous egg (Taylor





















Table 9. Serum phosphorus of laying hens at various times


Serum phosphorus
Time (mg.% S.E.)1

Trial 1 Trial 2 Trial 3


12 midnight 5.63 0.210 --- 5.53 + 0.272

4 a.m. --- --- 5.78 0.260

6 a.m. 5.78 0.278 5.82 0.283

7 a.m. -- -- 6.08 + 0.220

8 a.m. --- 4.88 0.249 ---

10 a.m. --- 4.90 0.355

12 noon 4.89 + 0.280 4.86 0.250

2 p.m. --- 5.29 0.342 ---

4 p.m. --- 5.40 + 0.214 ---

6 p.m. 5.35 + 0.306 5.42 0.321


mg.% = mg. phosphorus per 100 ml. serum; S.E. = standard error of the
mean.



































0
r-


0

C.'-






0


a)



~0



oH
0











44
0)










0
r4


::
w


.H
0
a0


0J

U)
0)
4r)


I-'-


bO


rl





'-H




on





39






E
00
I
I




\OD










CN





ca
o 3=



















co








Dolo

SN 0 0
)SOHdSHd
\c
4 01 0






--- *--------- ---------- s-S

Cms O -= C C C= C= C=
CO CO LO CIO Co C0 L o t




( O 4w SiMOHdSOHd NAUBS










et al., 1965), it can be assumed that shell deposition began in the

early afternoon. The increasing serum phosphorus value from 2 p.m. to

7 a.m. would, therefore, correlate with increasing shell calcification

and medullary bone demineralization during the same period. The build-

up in serum phosphorus can also be related to the inability of the

kidneys to excete phosphorus at a rate equal to its liberation from

bone (Taylor and Stringer, 1965).

The precise reason for the precipitous drop in serum phosphorus

from 6.08 mg.% at 7 a.m. to 4.88 mg.% at 8 a.m. is not known; however,

it can be speculated that during this same time shell deposition de-

creased or ceased and active bone remineralization increased. Renal

clearance would account for some lowering in the phosphorus level

(Taylor and Stringer, 1965). Additional influence on the serum phospho-

rus level during this period may have occurred due to the absorption of

minerals from feed consumed during the early part of the light period.

The serum phosphorus cycle determined from the results of these

trials was for a population of hens laying in the morning. Some shift

in the time of high and low phosphorus levels would probably occur for

a population of birds with an average oviposition time different from

that of these birds.

The serum phosphorus level of hens followed a definite pattern

during the laying cycle. The changes observed can be related to the

stages of activity of shell calcification and medullary bone deminerali-

zation and remineralization. The cycle presented here could act as a

model for determining times at which birds should be bled in order to

observe their phosphorus status. Therefore, it is suggested that in

phosphorus metabolism studies, serum phosphorus should be determined at








the predicted peak (6 7 a.m.) and in the trough (8 12 a.m.)

(Figure 1).


Summary


Three trials were conducted to measure changes in the serum phos-

phorus level of laying hens over a 24-hour period. Sixty-three-week-

old hens were selected for blood sampling according to the criteria

that they had laid an egg the day prior to and the day of blood collec-

tion, and that the oviposition time on both days was 10 a.m. 2 hours.

Blood samples were obtained via heart puncture. Serum was collected by

centrifugation, deproteinized and analyzed colorimetrically for phospho-

rus content. Sampling times were as follows: Trial 1 6 p.m., 12

midnight, 6 a.m. and 12 noon; Trial 2 6 a.m., 8 a.m., 10 a.m., 12

noon, 2 p.m., 4 p.m. and 6 p.m.; Trial 3 12 midnight, 4 a.m. and

7 a.m.

Cumulative results indicate that serum phosphorus reached a peak

of 6.08 mg.% at 7 a.m. At 8 a.m., the phosphorus level dropped to

4.88 mg.% and remained within 0.08 mg.% of this value until 12 noon.

From 2 p.m. to 6 a.m. it gradually increased from 5.29 to 5.80 mg.%.

It was hypothesized that observed changes in serum phosphorus over time

were related to changes in shell calcification, bone resorption and

remineralization and renal clearance.















CHAPTER V

THE EFFECTS OF PRODUCTION STATUS ON
SERUM CALCIUM AND PHOSPHORUS


Harms and Waldroup (1962) reported that the serum level of a sup-

plementally fed antibiotic oxytetracyclinee) varied considerably with

relation to the hen's production status. They found that hens laying

both the day before and the day of blood sampling exhibited serum

oxytetracycline levels of 0.240 mcg./ml. This level was considerably

higher than those observed for hens laying the day of collection but

not the day before, or not laying either day (0.170 and 0.079 mcg./ml.,

respectively). They concluded that the production status of the hen

altered calcium utilization which was reflected in the serum oxytetra-

cycline level. The following experiment was conducted to examine the

effect of production status on the serum calcium and phosphorus levels

of laying hens.


Procedure


Two trials were conducted to examine the effect of production

status on the serum calcium and phosphorus of laying hens. Babcock

B-300 hens, 66 weeks of age, were selected for blood sampling. All

birds were maintained in individual 20.3 X 45.7 cm. wire cages and fed

a corn-soybean meal laying diet calculated to contain 16.2% protein,

3.25% calcium, 0.75% total phosphorus and 2850 kcal. metabolizable











energy per kilogram of feed (Table 8). The birds received natural and/

or artificial light from 4:30 a.m. until 7:30 p.m. each day.

The day prior to each blood collection, a group of hens was se-

lected which had laid eggs between 8 a.m. and 12 noon. A second group

of hens which had not laid eggs the day prior to bleeding was also

selected. On the day of blood collection, each of these groups was

further divided into two sub-groups based on the same criteria. This

resulted in the four separate production status groups as follows:

group L:L = hens laying both the day before and the day of blood collec-

tion; group N:L = hens not laying the day before, but laying the day of

bleeding; group L:N = hens laying the day before but not laying the day

of bleeding; group N:N = hens not laying either day. For each produc-

tion group abbreviation, the letter preceding the colon indicates

whether the hen laid (L) or did not lay (N) the day prior to sampling

and the letter following the colon indicates laying status the day of

sampling.

In Chapter IV, it was suggested that blood samples be drawn both

during the predicted peak and trough of the mineral cycle in order to

observe differences in serum mineral levels. In each trial of this

experiment, 5 ml. of blood were collected from one-half of the birds in

each of the groups at 6 a.m. (predicted peak) and from the other half

at 11 a.m. (predicted trough).

The birds in group N:N had not laid an egg for three or more weeks,

although they had the physical characteristics of laying hens. Necropsy

of a randomly selected portion of this group revealed that the birds

had apparently functional reproductive tracts; therefore, a fifth group

of birds (designated 0:0) was selected for serum mineral analyses and









included in Trial 2. These birds were totally out of production and

exhibited regressed combs, wattles, ovaries and oviducts. This fifth

group was included in order to examine serum mineral levels of birds

with regressed reproductive tracts.

The same procedures for blood collection, serum extraction, de-

proteinization and serum calcium and phosphorus analyses were used as

outlined in Chapter III. Observed means and standard errors for serum

mineral levels of each production group at 6 and 11 a.m. were calculated.

Data were subjected to analyses of variance as described by Snedecor

(1956). No effects of trial or treatment X trial interactions were sig-

nificant; therefore, data from both trials were combined. Significant

differences among production status group means within each time period

were determined using the multiple range test described by Duncan

(1955). Significant differences between treatment means for each pro-

duction status group between time periods were determined using Student's

"t" tests (Snedecor and Cochran, 1973).


Results and Discussion


Birds which laid an egg both the day before and the day of blood

collection (group L:L) had a significantly higher serum phosphorus level

at 6 a.m. than any of the other production status groups (6.55 mg.%,

Table 10). Birds which either laid the day of blood collection but not

the day before, or did not lay on both days (groups N:L and N:N, respec-

tively) had intermediate serum phosphorus levels at 6 a.m. (5.29 and

5.32 mg.%, respectively). The 6 a.m. serum phosphorus level of those

birds which laid the day before but not the day of sampling (group L:N)
















Table 10.


Serum phosphorus levels of hens in different production
groups (Trials 1 and 2 combined)


No. birds/group Serum phosphorus
(mg.% S.E.)2
Production
groupI Collection time
6 a.m. 11 a.m.
6 a.m. 11 a.m.


L:L 13 13 6.55 0.274Ax 4.50 0.185By

N:L 10 7 5.29 0.208Bx 4.55 0.282B

L:N 11 11 4.43 0.173C 4.44 0.270B

N:N 10 11 5.32 0.186B 5.35 0.319A

0:0 5 5 3.21 0.264D 3.63 0.211C


1
L:L = hens laying both the day before and the day of blood collection

N:L = hens not laying the day before but laying the day of blood
collection

L:N = hens laying the day before but not the day of blood collection

N:N = hens not laying either day

0:0 = hens out of production

2Capital superscripts designate significant (p < 0.05) differences in
serum phosphorus among production groups within each collection time
as determined by Duncan's multiple range tests.

Lower case superscripts designate significant (p < 0.05) differences
in serum phosphorus between collection times for any production group
as determined by Student's "t" tests.


S.E. = standard error of the mean.









was lower (4.43 mg.%), while the birds in group 0:0 had the lowest

serum phosphorus level at 6 a.m. (3.21 mg.%).

At 11 a.m., group N:N had a significantly higher serum phosphorus

level and group 0:0 again had a significantly lower serum phosphorus

level than all other groups (Table 10). Between 6 and 11 a.m., the

serum phosphorus levels of groups L:L and N:L fell significantly from

6.55 to 4.50 mg.% and from 5.29 and 4.55 mg.%, respectively. There were

no significant changes between the 6 and 11 a.m. serum phosphorus levels

for groups L:N, N:N or 0:0.

As reported in Chapter IV, birds laying both the day before and the

day of blood collection exhibited a peak in serum phosphorus at approxi-

mately 6 to 7 a.m., and a trough from approximately 8 a.m. until 12

noon. The results from birds in the same class (group L:L) in this

experiment exhibited a similar pattern (Table 10).

The serum phosphorus of birds in group N:L did not reach as high a

peak at 6 a.m. as for those in group L:L. Since elevated serum phos-

phorus is a secondary result of bone resorption to supply calcium for

shell calcification (Taylor and Stringer, 1965), the lower 6 a.m. level

of serum phosphorus exhibited by group N:L would indicate a reduced

level of bone resorption. This reduction in demineralization could

possibly have been caused by an increase in available calcium stores in

soft tissue which were not utilized the previous day. The birds in

this group did lay on the day of sampling; therefore, the significant

decrease in serum phosphorus from 6 to 11 a.m. probably resulted from

increased bone remineralization.

It is believed that the low serum phosphorus level at 6 a.m. of

the birds in group L:N was the result of a combination of a lack of a









need for calcium for shell calcification and a continued necessity for

bone remineralization. Since these birds did not lay on the day of

blood collection, an elevated serum phosphorus caused by bone resorption

to supply calcium for egg shell calcification would not have occurred.

However, since these birds did lay the day before sampling, the need for

bone repletion of calcium and phosphorus would still be present. The

serum phosphorus level did not change between 6 and 11 a.m., indicating

that intestinal mineral absorption which occurred during this time was

probably utilized for bone remineralization.

The 6 a.m. serum phosphorus level of birds in group N:N was not

significantly different from that of group N:L; however, unlike those

in the latter group, the birds in group N:N did not lay on the day of

sampling. It is suggested that the high level observed at 6 a.m. was

related to phosphorus absorption from the diet the previous day. In

addition, the level was lower than that exhibited by birds which had

laid both days (group L:L), indicating possibly less active bone

dynamics. Birds in group N:N did have apparently functional reproduc-

tive tracts; therefore, it is probable that hormonal effects resulted

in a level of intestinal calcium and phosphorus absorption equal to

birds laying eggs. Also, since these birds had not produced a shell

for some time, the need for calcium and phosphorus replenishment of

bone would be greatly reduced which would account for the serum phospho-

rus level not decreasing between 6 and 11 a.m., and for it being the

highest measured at 11 a.m.

The birds in group 0:0 exhibited the lowest serum phosphorus levels

observed at both 6 and 11 a.m. The fact that these birds had regressed

ovaries probably resulted in a cessation of hormone secretion which









subsequently caused reduced phosphorus and calcium absorption from the

intestine. It is suggested that the levels observed for this group are

indicative of the normal circulating phosphorus content for birds which

are reproductively inactive.

No statistically significant differences in serum calcium levels

were observed for hens in groups L:L, N:L, L:N and N:N, either within

collection times or between collection times for the same production

group (Table 11). The observed value at 11 a.m. for each production

group was 0.8 to 2.2 mg.% lower than that measured for the same group

at 6 a.m. The reason for these lower values is not known. It is possi-

ble that a combination of increased muscle activity, increased excretion

and/or a need for calcium in bone remineralization could account for the

lower values observed. Since calcium is required for muscular contrac-

tion, the increased activity of birds after 6 a.m. could result in lower

serum calcium.

At 6 a.m., the birds in group L:N had a numerically lower serum

calcium level than groups L:L, N:L or N:N. This phenomenon, like that

of serum phosphorus, was probably a result of continued bone reminerali-

zation and lack of egg shell formation.

The birds in group 0:0 exhibited serum calcium levels significantly

lower than those of the other four groups at both 6 and 11 a.m. (Table

11). These calcium levels are similar to those reported for non-laying

hens, immature pullets and adult males (Bell and Sturkie, 1965). This

was probably the result of a lack of hormonal stimulation and subsequent

lowered calcium absorption from the diet and liberation from the bones.

In this experiment, serum phosphorus levels varied significantly

with relation to production status of the birds. Except for birds


















Table 11. Serum calcium levels of hens in different production groups
(Trials 1 and 2 combined)


Serum calcium
No. birds/group (mg.% S.E.)2
Production
group1 Collection time
6 a.m. 11 a.m.
6 a.m. 11 a.m.


L:L 13 13 32.1 1.05B 31.0 1.15B

N:L 10 7 33.3 0.72B 31.4 1.15B

L:N 11 11 30.4 1.15B 28.5 0.79B

N:N 10 11 34.0 1.22B 31.7 1.11

0:0 5 5 15.8 0.97A 15.0 0.68A


L:L = hens laying both the day before and the day of blood collection

N:L = hens not laying the day before but laying the day of blood
collection

L:N = hens laying the day before but not the day of blood collection

N:N = hens not laying either day

0:0 = hens out of production
2
Means in the same column not having common superscripts are signifi-
cantly different (p < 0.05) as determined by Duncan's multiple range
tests.

S.E. = standard error of the mean.









exhibiting regressed reproductive tracts, serum calcium did not seem to

be as greatly influenced by production status as was serum phosphorus.

It is concluded that birds selected for serum mineral analyses should

be of similar production status.


Summary


An experiment was conducted to examine the effect of production

status on the serum calcium and phosphorus of laying hens. Serum phos-

phorus varied significantly with relation to whether or not a bird laid

an egg the day before and/or the day of blood collection. Except for

birds which were reproductively inactive, serum calcium was less af-

fected by production status than was serum phosphorus. It was hypothe-

sized that the variations were related to the physiological changes in

activity of shell calcification, bone resorption and remineralization,

intestinal absorption of minerals and muscular activity. It was con-

cluded that birds selected for serum mineral analyses should be of

similar production status.
















CHAPTER VI

SERUM CALCIUM AND PHOSPHORUS LEVELS IN
RELATION TO TIME OF OVIPOSITION


Sloan (1976) reported that the serum calcium level four hours

before and after oviposition was significantly lower than at time of

lay. Paul and Snetsinger (1969) reported that serum calcium gradually

decreased from 1 through 23 hours after oviposition, and that serum

phosphorus decreased from 1 through 9 hours after time of lay.

Since a large reduction in serum phosphorus has been shown to

occur after 7 a.m. (Chapter IV), the following experiment was designed

to examine serum phosphorus levels in relation to time of oviposition

in order to more precisely determine at what point in the laying cycle

the reduction occurs.


Procedure


Babcock B-300 laying hens, 68 weeks of age, were used for serum

mineral analyses. All birds were maintained in individual 20.3 X 45.7

cm. wire cages and fed a corn-soybean meal laying diet calculated to

contain 16.2% protein, 3.25% calcium, 0.75% total phosphorus and 2850

kcal. metabolizable energy per kilogram of feed (Table 8). The birds

received natural and/or artificial light from 4:30 a.m. until 7:30 p.m.

each day.

In Chapter V it was recommended that blood samples used for serum

mineral analyses be taken from birds of similar production status.











Therefore, in this experiment, all birds which had laid an egg between

8 a.m. and 12 noon the day prior to sampling were identified and sub-

sequently used for blood collection.

At 5 a.m. the following day, and at each hourly interval until 12

noon, 5 ml. of blood were collected via anterior heart puncture from

each of 10 to 20 birds which had not laid an egg by the time of sampling.

By recording the time of oviposition of these birds, and back-timing to

the point of sampling, it was possible to select groups of 10 birds

from which samples were taken at hourly intervals from one to five

hours prior to oviposition.

The birds which laid between 8 a.m. and 8:15 a.m. were randomly

distributed to four bleeding times. This same procedure was used to

assign birds which laid during each subsequent 15 minute interval from

8:15 a.m. until 12 noon to the four bleeding times. Five ml. of blood

were collected from each bird according to its bleeding time assignment.

This resulted in 10 hens being bled at 0, 1, 2 and 4 hours after

oviposition.

By using the procedures outlined above, possible variations in

serum mineral levels due to the time of day of oviposition were reduced.

No bird was bled more than once during the experiment. This resulted

in 90 separate samples, 10 from each time interval, for analyses of

serum calcium and phosphorus

The same procedures for blood collection, blood clotting, serum

extraction, sample deproteinization and serum calcium and phosphorus

determinations were used as outlined in Chapter III. The mean and

standard error of calcium and phosphorus levels at each time period were

calculated. Data were subjected to analyses of variance procedures as









described by Snedecor (1956). Significant differences among treatment

means were established using the multiple range test described by

Duncan (1955). Regression analyses were used to construct the best fit

lines for both serum calcium and phosphorus data (Steel and Torrie,

1960).


Results and Discussion


Serum phosphorus increased in a step-wise manner from 6.46 to 6.82

mg.% during the period from five to three hours before oviposition

(Table 12). Between three and two hours prior to time of lay, serum

phosphorus had fallen to a level of 4.75 mg.%, where it remained within

0.35 mg.% of this value through four hours post-oviposition. The dif-

ferences in serum phosphorus observed in this experiment were probably

related to changes in activity of bone resorption and remineralization.

The high levels of serum phosphorus were probably a result of bone re-

sorption to supply calcium for the shell, while the reduced levels cor-

responded to the period of expected increases in the rate of bone

remineralization.

Graphic representation of the data (Figure 2) revealed a pattern

similar to the results of the serum phosphorus cycle reported in

Chapter IV, in which it was reported that serum phosphorus dropped be-

tween 7 and 8 a.m. From the data in this experiment, it appeared that

this drop corresponded to a period approximately two to three hours

prior to oviposition.

Serum calcium levels exhibited only minor differences throughout

the 10 hour test period (Table 12). Regression anlaysis of the data

revealed an almost horizontal best fit line, indicating essentially no






















Table 12. Serum calcium and phosphorus in relation to time of
oviposition


Time in relation to Serum phosphorus Serum calcium
oviposition (hours) (mg.% S.E.1 (mg.% S.E.)


-5.0 0.25 6.46 0.221 34.3 0.98

-4.0 0.25 6.71 0.361 34.3 + 0.87

-3.0 0.25 6.82 0.562 34.6 1.90

-2.0 + 0.25 4.75 0.439 33.6 + 1.08

-1.0 0.25 5.10 + 0.243 34.6 1.08

0 4.96 0.169 33.8 + 0.99

+1.0 4.76 0.224 33.2 0.85

+2.0 4.55 0.254 33.7 1.25

+4.0 4.66 + 0.168 34.0 1.41


S.E. = standard error of the mean.









56












0N












C=











L--
=





C=


CI-)




L.-
C=













LM-



















0c)Lf Co CDCDCDA
Ui Co ro LA LA


M hW) SAOWN~S~d~1J








change in serum calcium from five hours before through four hours after

oviposition (Figure 3). These results were somewhat different than

those observed by Sloan (1976), who reported significant changes in

serum calcium before and after oviposition. The calcium level of the

diet used in his study was 3.00%. It is possible that the 3.00% calcium

level was "deficient" in terms of maintenance of a stable serum calcium

level. It is suggested that when dietary calcium is optimal, the level

of serum calcium will not change appreciably during the laying cycle.

Under the conditions of this experiment, it appears that 3.25% calcium

was optimal in terms of maintenance of a stable serum calcium level.

Based on the results of this experiment, and the data reported in

Chapter IV, it is hypothesized that the laying cycle of the hen, in

terms of serum calcium and phosphorus, is composed of two segments of

approximately 6 and 18 hours duration. During the 18 hour segment,

calcium and phosphorus are removed from the medullary bones. The

calcium is utilized for shell formation; therefore, its level in the

blood (under optimum conditions) does not vary appreciably. Phosphorus,

however, is not utilized extensively in shell formation and, therefore,

the serum level increases. During the six hour segment, calcium and

phosphorus are both utilized for bone remineralization. The onset of

this segment results in a significant drop in serum phosphorus. Serum

calcium does not change due to a greatly reduced or eliminated need for

shell formation, as well as an increased supply of calcium from intesti-

nal absorption.









59































*=


CC4
m







Cn

MOM N--
I .

















U-










@ Wlv
r~- m
n~ n c


(Ui~ TO1V) Hf^








Summary


An experiment was conducted to examine the serum calcium and phos-

phorus levels in relation to time of oviposition. Samples taken at

hourly intervals, from five hours before through two hours after, and

at four hours after oviposition were analyzed for serum calcium and

phosphorus. Results indicated that serum phosphorus increased from

6.46 to 6.82 mg.% during the period from five to three hours prior to

oviposition. Between three and two hours before time of lay, it signifi-

cantly dropped to a level of 4.75 mg.%, where it remained within 0.35

mg.% of this value for the remainder of the test period. Serum calcium

did not change appreciably during the 10-hour test period. It was

hypothesized that the serum calcium and phosphorus cycles of the hens

are composed of two segments of approximately six and 18 hours duration.

The segments were delineated on the basis of activities of bone resorp-

tion and remineralization and eggshell formation.















CHAPTER VII

INFLUENCE OF VARIOUS DIETARY CALCIUM AND PHOSPHORUS
LEVELS ON THE PERFORMANCE OF BROILER BREEDER HENS


In Chapter I it was reported that hatchability of fertile eggs was

significantly improved by elevating the dietary calcium level at low

levels of inorganic phosphorus. In a subsequent experiment (Chapter

II), it was found that increasing the dietary calcium level when the

total phosphorus in the diet was supplied by plant sources resulted in

significantly reduced egg production and hatchability. Both experiments

were conducted using caged commercial egg production type hens. The

following experiment was conducted to examine the effects of various

dietary calcium and phosphorus levels on the performance of broiler

breeder hens maintained in floor pens.


Procedure


Five hundred four 26-week-old Cobb color-sexed broiler breeder

pullets were randomly and equally distributed to 72 floor pens. Each

pen measured 1.54 X 1.85 meters and contained peanut hulls as litter.

A 3 X 6 factorial design composed of 18 dietary treatments was used in

the experiment, with four replicate pens assigned to each diet. The

diets were calculated to contain 2.25, 2.85 or 3.50% calcium, with 0.31,

0.36, 0.41, 0.56, 0.71 or 1.42% total phosphorus. All experimental

diets were mixed from a corn-soybean meal basal diet calculated to con-

tain 0.09% calcium and 0.31% phosphorus (Table 13). A 14.00% portion

















Table 13. Composition of broiler breeder basal diet


Ingredient Percent of the diet


Yellow corn meal 56.97

Soybean meal (50%) 21.00

Alfalfa meal (17%) 2.00

Animal fat 5.00

Microingredients1 0.50

Salt 0.40

Methionine Hydroxy Analog 0.13

Variable 14.00


Crude protein 16.2%

Metabolizable energy 2850 kcal./kg.

Calcium 0.90%

Phosphorus 0.31%


1Supplied per kg. of diet: 6000 I.U. vit. A.; 2200 I.C.U. vit. D3;
2.2 mg. menadione dimethylpyrimidinol bisulfate; 4.4 mg. riboflavin;
13.2 mg. pantothenic acid; 39.6 mg. niacin; 499.4 mg. choline
chloride; 22 mcg. vit. BI2; 125 mg. ethoxyquin; 50 mg. manganese;
50 mg. iron; 6 mg. copper; 0.198 mg. cobalt; 1.1 mg. iodine; 35 mg.
zinc.








of the basal diet was composed of ground limestone, dicalcium phosphate

and white builder's sand in proportions required to provide the calcium

and phosphorus levels in each experimental diet. All diets were iso-

caloric and isonitrogenous. The birds received feed ad libitum, and

were supplied with natural and/or artificial light from 4:30 a.m. to

7:30 p.m. each day.

Hen-day egg production and feed consumption were recorded for seven

28-day periods following initiation of the dietary treatments. During

the third, fifth and seventh months of the experiment, all birds were

inseminated with pooled semen from Cobb color-sexed broiler breeder

males. Eggs were collected from the second through the eighth days

following each insemination and incubated. All eggs were set in James-

way 252 incubators maintained at 370 C. and 62% relative humidity.

Fertility was determined after seven days of incubation. During the

fourth and sixth months of the experiment, shell quality, as measured

by egg specific gravity, was determined on one day's eggs from each

treatment.

All data were subjected to split-plot in time analyses as described

by Steel and Torrie (1960). Since no interactions between treatments

and/or times existed, data for main effects across time were combined.

Significant differences among the main effect means were determined

using the multiple range test described by Duncan (1955).


Results and Discussion


Cumulative hen-day egg production increased numerically for each

increment increase in dietary phosphorus from 0.31 to 0.41% (Table 14).

This trend was reversed, exhibiting a decrease in production at each


























4J
CO











-1
*il
'0











.H






























c



0)
c)

,-4




>







-H
cn
















0
4-
C )


OR

4-C





































0)
4-id





0 o


C d
) e


P4-i
)a







)-l U

0)

,0




C3


-4






4-1
il







C
r-




ca













0




0)

















4J
0, -


















41
4 r-
Cd




















4 C L ,
oH
CO
*.4
0d



o- c





















0












HO
o *
n3 6~
43u
i-






































&.


64








,-4 O ca ca ca .0

0 0 -O 0 O

0 0 0 0 0 0
0 0 0 0 0 0



+1 +1 +1 +1 +1 +1

0 LrC L 0 CO r
C ,- ,-- o o r-
c 00 000 c0 r-




















-T C1 r4 Ci CN t

i-4 Ln Lm Ln LP C
r- r-- r r- h- t--


















cc c cm o cc c









;0 00 00 CO 00 C










'\0 '0 '.0 '.0 'D 'ko












O- .O -4 CO C- C-
cn m It Ln r- c
n Co 4 L o o 4
o 0 0 0 0 -


C0
CO
u




tU
3





C)
*rl
0



40








0)













.r-.
-O













U)




o
0

0













































r.1
0 0
cu


















4 -.


C 4--
H)





















o

4-4
C)




0)
r-4

01


-~-4



a)

ca
4-4












d



dr-l









phosphorus level above 0.41%. There was essentially no difference in

egg production for birds fed 0.41 or 0.56% total phosphorus (64.3 and

63.6%, respectively); however, dietary phosphorus treatments above or

below these levels resulted in approximately 2 to 4 percentage point

decreases in production. These results were similar to those found by

Singsen et al. (1962), who reported that for hens maintained on litter,

dietary phosphorus levels below 0.40% or above 0.50% resulted in de-

pressions in hen-day egg production. No significant differences in egg

production were observed between any of the dietary calcium levels

(Table 15). Although birds receiving diets containing 2.85% calcium

exhibited approximately 2.3 to 3 percentage points lower production

that the other two treatments, this same relationship existed in the

pre-treatment egg production results (data not shown) and was, therefore,

considered a result of randomization rather than dietary treatment.

Cumulative hatchability of fertile eggs indicated no significant

differences due to dietary phosphorus or calcium levels (Tables 14 and

15, respectively). Birds receiving diets containing 0.31 or 1.42%

phosphorus exhibited approximately 3.0 to 3.5 g./bird/day lower feed

consumption, as did birds receiving the 2.85% calcium diets. Whether

these lower feed consumption values were the cause or result of lowered

egg production is not known.

Eggs from hens receiving the all plant phosphorus diets (0.31%)

exhibited the best shell quality, as measured by specific gravity. As

the dietary phosphorus level increased from 0.31 to 0.71%, specific

gravity of eggs decreased; however, these decreases were not statisti-

cally significant (Table 14). Specific gravity of eggs from hens re-

ceiving 1.42% total phosphorus was significantly lower than for eggs




















+1
1-1





















Q co
U























r0)
4-.1














o o
4-J






















I-9


4J
(U



a-o

















0
U4
a3
QJ 0
(U


0



0

Q-









from hens fed any of the other dietary treatments. No differences in

specific gravity were observed at any of the different calcium levels

(Table 15). Specific gravity was somewhat lower for eggs from hens re-

ceiving the 2.25% calcium diets; however, it was not statistically

significant.

The egg production and hatchability results of birds fed diets

containing phosphorus supplied by all plant sources in this experiment

were not significantly depressed as they were for the birds maintained

in cages in a previous experiment (Chapter II). The precise reasons

for the improved overall performance at lower phosphorus levels for

birds maintained on litter are not known; however, as suggested by

Singsen et al (1962), access, by coprophagy, to phosphorus in the

litter was the most probable reason for improved performance. Those

authors measured accumulated phosphorus in the litter at the end of a

40-week experiment. They reported phosphorus levels in the litter

to be three times greater than that supplied by the diets. It is also

probable that ingestion of excreted phosphorus by the hens fed the

higher phosphorus levels (0.71 and 1.42%) resulted in detrimental effects

on calcium absorption and the subsequently observed lower egg production

and egg specific gravity for these groups.

The best overall performance across calcium levels in this experi-

ment was exhibited by birds receiving 0.41% total phosphorus. Based on

these results, it would appear that the 0.60% total phosphorus level

recommended by the National Research Council (1971) is too high in terms

of the requirement for phosphorus by birds maintained on litter.











Summary


An experiment was conducted to examine the effects of various

dietary calcium and phosphorus levels on the performance of broiler

breeder hens maintained on litter. Results indicated that increasing

dietary phosphorus levels from 0.31 to 0.41% tended to improve hen-day

egg production, while increasing the phosphorus level above 0.41% tended

to depress production. Hatchability of fertile eggs and feed consump-

tion were essentially unaffected by dietary treatments; however, minor

improvements in both parameters were observed when the level of phos-

phorus was elevated above 0.31%. Shell quality, as measured by specific

gravity, numerically decreased when phosphorus was increased above

0.31%. When diets contained 1.42% phosphorus, specific gravity of eggs

was significantly reduced. None of the parameters measured were sig-

nificantly affected by altering the dietary calcium level. The ob-

served performance at low dietary phosphorus levels of hens maintained

on litter was generally better than the results observed for caged birds

in previous experiments. It was suggested that "recycling" of phospho-

rus by coprophagy was responsible for the overall better performance of

birds maintained on litter as opposed to those maintained in cages.















SUMMARY


A series of seven experiments was conducted to examine the factors

affecting serum mineral levels and the influence of dietary calcium and

phosphorus interrelationships on performance characteristics of the

domestic hen.

In the first experiment, effects on the hatchability of fertile

eggs were observed for caged laying hens fed diets containing 2.25 or

3.50% calcium, with 0.35, 0.40, 0.45, 0.55, 0.70 or 1.42% total

phosphorus. Effects on hatchability of eggs from hens fed diets con-

taining either 0.45 or 0.70% total phosphorus and 6.00% calcium were

also examined. Results indicated that, at a phosphorus level of 0.35%,

elevating the dietary calcium from 2.25 to 3.50% resulted in a signifi-

cant improvement in hatchability of fertile eggs. Feeding diets con-

taining 6.00% calcium neither improved or depressed hatchability of

fertile eggs, when compared to results from hens fed comparable phos-

phorus levels but at lower total calcium levels.

The second experiment was designed to examine the effects of

elevating the calcium level from 2.25 to 3.50% in diets containing

either 0.29% phosphorus (from plant sources) or 0.55% (from plant and

supplemental inorganic sources). Results indicated that elevating the

calcium level in diets where all phosphorus was supplied from plant

sources resulted in significant reductions in hen-day egg production

and hatchability of fertile eggs.









In the third experiment, the effects of liquid oral dosing versus

feed supplementation of phosphorus on the performance of phosphorus-

depleted hens was observed. Groups of phosphorus-depleted hens were

maintained on the phosphorus-deficient diet, fed the deficient diet

supplemented with phosphorus from phosphoric acid or administered sup-

plemental phosphorus via oral liquid dosing of 1.0 N phosphoric acid.

A fourth group of hens was maintained on a phosphorus-adequate diet to

serve as a positive control. Results indicated that oral liquid dosing

of phosphorus-depleted hens affected a more rapid improvement in egg

production and hatchability than feed supplementation. However, feed

supplementation resulted in a greater improvement in hatchability over

time.

The fourth, fifth and sixth experiments were designed to observe

factors affecting serum mineral levels of laying hens. In the fourth

experiment, the pattern of serum phosphorus changes over a 24-hour

period was determined. In experiments five and six, the influence of

rate of egg production and time in relation to oviposition on serum

calcium and phosphorus levels were examined. Results from the fourth

experiment indicated that serum phosphorus levels were not stable, but

underwent considerable changes during the laying cycle. Serum phospho-

rus appeared to reach a peak level at approximately 7 a.m. By 8 a.m.,

the level was significantly reduced and remained low until 12 noon.

From approximately 2 p.m. until 7 a.m., the serum phosphorus level

gradually increased until the peak level was observed at 7 a.m. Results

from the sixth experiment indicated that the peak level observed for

serum phosphorus correlated to a time approximately three hours prior

to oviposition. Between three and two hours before time of lay, the









level significantly dropped to its lowest point, where it remained rela-

tively unchanged for the next six hours. Serum calcium seemed to remain

relatively unchanged during the period from five hours before to four

hours after oviposition. In the fifth experiment, it was determined

that production status of the hen over a two-day period significantly

affected the serum phosphorus levels measured at 6 and 11 a.m., but had

little affect on serum calcium levels.

The last experiment was designed to examine the effects of differ-

ent dietary calcium and phosphorus levels on the performance of hens

maintained on litter. Results indicated that there were no interactions

between dietary calcium and phosphorus levels as observed in the first

two experiments. It was observed that, over all calcium levels, a

dietary phosphorus level of 0.41% resulted in the best egg production,

while increases above this level depressed production. Any supplementa-

tion of the diets with inorganic phosphorus resulted in numerical de-

creases in shell quality, as measured by specific gravity. At total

dietary phosphorus levels of 1.40%, shell quality was significantly

reduced. Alterations in dietary calcium and phosphorus had little

effect on hatchability of fertile eggs or feed consumption.















REFERENCES


Ademonsun, A. A. and N. Nikolaiczuk, 1966. Calcium and phosphorus
metabolism in the laying hen. Proc. 13th World Poultry Cong.:
320-322.

Arscott, G. H., P. Rachapaetayakom and P. E. Bernier, 1962. Influence
of ascorbic acid, calcium and phosphorus on specific gravity of
eggs. Poultry Sci. 41:485-488.

Balloun, S. L. and D. L. Miller, 1964. Calcium requirements of turkey
breeder hens. Poultry Sci. 43:378-381.

Bell, D. J. and P. D. Sturkie, 1965. Chemical constituents of blood.
In: Avian Physiology. P. D. Sturkie, Ed. Cornell University
Press. Ithaca, New York.

Bloom, W., A. V. Nalbandov and M. A. Bloom, 1960. Parathyroid enlarge-
ment in laying hens on a calcium-deficient diet. Clinical
Orthopaedics 17:206-210.

Buckner, G. D. and J. H. Martin, 1920. Effect of calcium on the compo-
sition of the eggs and carcass of laying hens. J. Biol. Chem. 41:
195-203.

Buckner, G. D., J. H. Martin and F. E. Hull, 1930. The distribution of
blood calcium in the circulation of laying hens. Am. J. Physiol.
93:86-91.

Buckner, G. D., J. H. Martin and A. M. Peter, 1925. The relation of
calcium restriction to the hatchability of eggs. Am. J. Physiol.
72:458.

Buckner, G. D., J. H. Martin and A. M. Peter, 1928. The relative
utilitization of different calcium compounds by hens in the produc-
tion of eggs. J. Agri. Sci. 23:555-556.

Damron, B. L., A. R. Eldred and R. H. Harms, 1974. The relationship of
dietary phosphorus to egg shell quality. Poultry Sci. 53:1916.

Deobald, H. J., J. B. Christiansen, E. B. Hart and J. G. Halpin, 1938.
The relationship between blood calcium and blood phosphorus and
the effect of calcium changes in the laying pullet. Poultry Sci.
17:114-119.

Deobald, H. J., E. J. Lease, E. B. Hart and J. G. Halpin, 1936.
Studies on the calcium metabolism of laying hens. Poultry Sci.
15:179-185.










Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics
17:1-42.

Evans, R. J., J. S. Carver and A. W. Brant, 1944. The influence of
dietary factors on egg shell quality. II. Calcium. Poultry Sci.
15:179-185.

Feinberg, J. G., J. S. Hughes and H. M. Scott, 1937. Fluctuations of
calcium and inorganic phosphorus in the blood of the laying hen
during the cycle of one egg. Poultry Sci. 16:132-134.

Ferguson, T. M., C. D. Sewell, Jr. and R. L. Atkinson, 1974. Phosphorus
levels in the turkey breeder diet. Poultry Sci. 53:1627-1629.

Fiske, C. H. and Y. Subbarow, 1925. The colorimetric determination of
phosphorus. J. Biol. Chem. 66:375-400.

Garlich, J. D., R. L. James and J. B. Ward, 1975. Effects of short
term phosphorus deprivation on laying hens. Poultry Sci. 54:1193-
1199.

Greenberg, D. M., C. E. Larson, P. B. Pearson and B. R. Burmester, 1936.
The state and partition of the calcium and inorganic phosphorus in
the serum of the fowl. Effect of growth and ovulation. Poultry
Sci. 15:438-445.

Gutowska, Marie S. and R. T. Parkhurst, 1942. Studies in mineral
nutrition of laying hens. II. Excess of calcium in the diet.
Poultry Sci. 21:321-328.

Halnan, E. T., 1925. The calcium, phosphorus and nitrogen balance of
non-laying and laying pullets. J. Nat. Poultry Inst. 10:40.

Harms, R. H., 1971. How important are calcium:phosphorus ratios for
chickens? Feedstuffs 43(7):26-27.

Harms, R. H. and P. W. Waldroup, 1961. The influence of dietary calcium
level and supplementary ascorbic acid and/or dienestrol diacetate
upon performance of egg production type hens. Poultry Sci. 40:1345-
1348.

Harms, R. H. and P. W. Waldroup, 1962. Relationship of the egg forma-
tion cycle to antibiotic blood level in laying hens. Poultry Sci.
41:1932-1936.

Harms, R. H., C. B. Ammerman and P. W. Waldroup, 1964. The effect of
supplemental phosphorus in the breeder diet upon hatchability of
eggs and bone composition of chicks. Poultry Sci. 43:209-212.

Harms, R. H., C. R. Douglas and P. W. Waldroup, 1961. The effects of
feeding various and sources of phosphorus to laying hens. Fla.
Agric. Exp. Sta. Bull. 644.








Harshaw, H. M., J. C. Fritz and H. W. Titus, 1934. The normal develop-
ment of the leg bones of chickens with respect to their ash
content. J. of Agri. Research 48:997-999.

Heller, V. G., H. Paul and R. B. Thompson, 1934. Changes in the blood
calcium and phosphorus partition during the life cycle of the
chicken. J. of Biol. Chem. 106:357-360.

Hughes, J. S., R. W. Titus and B. L. Smits, 1927. The increase in the
calcium of hens' blood accompanying egg production. Science 65:
264-267.

Hurwitz, S. and A. Bar, 1966. Calcium depletion and repletion in laying
hens. I. Effect on calcium in various bone segments, in egg
shell and in blood plasma and on calcium balance. Poultry Sci. 45:
345-352.

Hurwitz, S. and S. Bornstein, 1963. The effect of calcium and phospho-
rus in the diet of laying hens on egg production and shell quality.
Israel S. Agric. Res. 13:147-154.

Hurwitz, S. and P. Griminger, 1962. Egg production and shell quality
in temperature and light controlled versus uncontrolled environment.
Poultry Sci. 41:499-508.

Jensen, L. S., H. C. Saxena and J. McGinnis, 1963. Nutritional investi-
gations with turkey hens. 4. Quantitative requirement for calcium.
Poultry Sci. 42:604-607.

Kalch, L. W., 1976. Personal communication.

MacIntyre, T. M., H. W. R. Chance and E. E. Gardiner, 1963. Effect of
dietary energy and calcium level on egg production and egg quality.
Can. J. Anim. Sci. 43:337-340.

Massengale, 0. N. and C. S. Platt, 1930. Effect of calcium from differ-
ent sources on the growth and egg production of poultry. Poultry
Sci. 9:240-246.

Maynard, L. A. and J. K. Looski, 1969. Animal Nutrition, 6th Ed.,
McGraw-Hill Book Co., New York, New York.

Mehring, A. L., Jr., 1964. A comparison of three methods of supplying
calcium in the diet of laying chickens. Poultry Sci. 43:976-981.

Mehring, A. L., Jr., 1965. Effect of level of dietary calcium on
broiler-type laying chickens. Poultry Sci. 44:240-248.

Mehring, A. L., Jr. and H. W. Titus, 1964. The effects of low levels
of calcium in the diet of laying chickens. Poultry Sci. 34:1385-
1388.


Merck Index, Eighth Ed., Merck and Co., Inc., 1968.









Miller, M. W. and G. F. Bearse, 1934. Phosphorus requirements of
laying hens. Wash. Agri. Exp. Sta. Bull. 320.

Mountcastle. V. B.. Editor, 1974. Medical Physiolovgy 13th ed. The
C. V. Mosby Company, St. Louis, Mo.

Mueller, W. J., 1961. The effect of constant and fluctuating environ-
mental temperatures on the biological performance of laying
pullets. Poultry Sci. 40:1562-1571.

National Research Council, 1971. Nutritional requirements of poultry.
National Academy of Sciences, Washington, D. C.

Nevalainen, T. J., 1969. The effect of calcium-deficient diet on the
reproductive organs of the hen (Gallus domesticus). Poultry Sci.
48:653-659.

Norris, L. C., G. F. Heuser, H. S. Wilgus, Jr. and A. T. Ringrose,
1933. The calcium and phosphorus of laying hens. 46th An. Rept.
Cornell Agri. Exp. Sta.:137-142.

O'Rourke, W. F., H. R. Bird, P. H. Phillips and W. W. Cravens, 1954.
The effect of low phosphorus rations on egg production and
hatchability. Poultry Sci. 33:1117-1122.

Paul, H. S. and D. C. Snetsinger, 1969. Dietary calcium and phosphorus
and variations in plasma alkaline phosphatase activity in relation-
ship to physical characteristics of egg shell. Poultry Sci. 43:
241-250.

Pepper, W. F., S. J. Slinger, J. D. Summers and G. C. Ashton, 1959. On
the phosphorus requirements of chickens for egg production and
hatchability. Can. J. Anim. Sci. 39:182-192.

Pepper, W. F., J. D. Summers and J. D. McConachie, 1968. The effect of
high levels of calcium, fish products and sodium bicarbonate on
eggshell quality. Poultry Sci. 47:224-229.

Petersen, C. F., D. H. Conrad, D. H. Jumijarui, E. A. Sauter and C. E.
Lampman, 1960. Studies on the calcium requirement of high produc-
ing White Leghorn hens. Idaho Agr. Exp. Sta. Bull. No. 44:1-7.

Peterson, W. J. and D. B. Parrish, 1939. Fluctuations of phosphatase
and inorganic phosphorus in the blood of the laying hen during the
period of egg formation. Poultry Sci. 18:54-58.

Potter, L. M., A. T. Leighton, Jr. and A. B. Chu, 1974. Calcium,
phosphorus and nopgro as variables in diets of breeder turkeys.
Poultry Sci. 53:15-22.

Reddy, C. V., P. E. Sanford and R. E. Clegg, 1968. Influence of calcium
in laying rations on shell quality and interior of eggs. Poultry
Sci. 47:1077-1083.









Roland, D. A., Sr., D. R. Sloan and R. H. Harms, 1972. Calcium metabo-
lism in the laying hen. 2. Patterns of calcium intake, serum
calcium, and fecal calcium. Poultry Sci. 51:782-787.

Roland, D. A., Sr., D. R. Sloan, H. R. Wilson and R. H. Harms, 1973.
Influence of dietary calcium deficiency on yolk and serum calcium,
yolk and organ weights and other selected production criteria of
the pullet. Poultry Sci. 52:2220-2225.

Russell, W. C., C. H. Howard and H. F. Ness, 1930. The relationship in
the hen between the development of ova, blood calcium and the
antirachitic factor. Science 72:506-511.

Sanford, P. E. and R. L. Alder, 1969. Effects of increasing levels of
phosphorus with increasing levels of calcium. Poultry Sci. 48:
1866.

Sewell, C. E., Jr., R. L. Atkinson, J. R. Couch and T. M. Ferguson,
1972. The effect of supplemental phosphorus on the reproductive
performance of turkey hens and the subsequent effect upon the
poults. Poultry Sci. 51:792-796.

Singh, R. D., J. K. Bletner and 0. E. Goff, 1971. Bone density index,
bone breaking strength, egg production, and egg specific gravity
as affected by dietary calcium and phosphorus. Poultry Sci. 50:
1631.

Singsen, E. P., A. H. Spandorf, L. D. Matterson, J. A. Serafin and
J. J. Tlustohowicz, 1962. Phosphorus in the nutrition of the
adult hen. 1. Minimum phosphorus requirements. Poultry Sci. 41:
1401-1414.

Sloan, D. R., 1976. Factors affecting calcium metabolism and eggshell
quality in the laying hen. Ph. D. dissertation, University of
Florida.

Snedecor, G. W., 1956. Statistical Methods, 5th Ed. The Iowa State
College Press, Ames, Iowa.

Snedecor, G. W. and W. G. Cochran, 1973. Statistical Methods, 6th Ed.
The Iowa State College Press, Ames, Iowa.

Steel, R. G. D. and J. M. Torrie, 1960. Principles and Procedures of
Statistics. McGraw-Hill Book Co., Inc., New York, New York.

Sullivan, T. W. and J. R. Kingan, 1962. Effects of dietary calcium
level, calcium lactate and ascorbic acid on the egg production of
S. C. White Leghorn hens. Poultry Sci. 41:1590-1596.

Taylor, M. W. and W. C. Russell, 1935. The diffusable calcium in the
serum of laying and non-laying hens. J. Agric. Res. 51:663-670.









Taylor, T. G. and D. A. Stringer, 1965. Eggshell formation and skeletal
metabolism. In: Avian Physiology. P. D. Sturkie, Ed. Cornell
University Press. Ithaca, New York.

Taylor, T. G. and A. Williams, 1964. Cyclic change in plasma acid and
alkaline phosphatase concentration associated with eggshell calci-
fication in the fowl. Biochem. J. 91:21-26.

Taylor, T. G., A. Williams and J. Kirkley, 1965. Cyclic changes in the
activities of plasma acid and alkaline phosphatase during eggshell
calcification in the domestic fowl. Can. J. Biochem. 43:451-452.

Waldroup, P. W. and R. H. Harms, 1961. Potentiation of terramycin. II.
Evaluation of low dietary calcium in laying hen diets. Avian
Diseases 5:409-415.

Waldroup, P. W., J. F. Maxey and L. W. Luther, 1974. Studies on the
calcium and phosphorus requirements of caged turkey breeder hens.
Poultry Sci. 53:886-888.

Waldroup, P. W., C. F. Simpson, B. L. Damron and R. H. Harms, 1967. The
effectiveness of plant and inorganic phosphorus in supporting egg
production in hens and hatchability and bone development in chick
embryos. Poultry Sci. 46:659-664.

Walter, E. D. and J. R. Aitken, 1962. Phosphorus requirement of laying
hens confined to cages. Poultry Sci. 41:386-390.

Wolford, J. H. and K. Tanaka, 1970. Factors influencing eggshell
quality--a review. World's Poult. Sci. J. 26:763-780.
















BIOGRAPHICAL SKETCH


The author, Edgar R. "Ted" Miller, was born February 18, 1950, in

Orange, New Jersey, and moved to Winter Haven, Florida, in 1959. He

graduated with honors from Winter Haven Senior High School in June,

1968. In September of the same year he entered the University of

Florida. As an undergraduate he was a member of Alpha Zeta and Lambda

Gamma Phi; he was on the dean's list three times and the President's

honor roll once.

He received his B.S.A. with honors in June, 1973, and entered

Graduate School the following quarter. The author received the degree

of Master of Science in Agriculture in August, 1974. While in Graduate

School, he served as President and Treasurer of the Poultry Science

Club and as the Poultry Science representative to the Student Agricul-

tural Council. He is also a member of Gamma Sigma Delta and the

Poultry Science Association. In May, 1976, he was awarded the Ruby V.

Voitle award as the outstanding graduate student in the department of

Poultry Science.

The author is married to the former Ellen Phillips of Jacksonville,

Florida. They have one son, Christopher Reid.








I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.






Henry R./ ilson, Chairman
Professor of Poultry Science



I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.






Robert A. Voitle
Associate Professor of Poultry Science



I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.






Douglas hl. Janky /
Assistant Professor of Poul ry Science



I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.






Ramon C. Littel-
Associate Professor of Statistics








I certify that I have read this study and that in my opinion it
conforms to acceptable standards of scholarly presentation and is fully
adequate, in scope and quality, as a dissertation for the degree of
Doctor of Philosophy.


J mes A. Himes
associatee Professor
Medicine


$ht~l


of Veterinary


This dissertation was submitted to the Graduate Faculty of the College
of Agriculture and to the Graduate Council, and was accepted as partial
fulfillment of the requirements for the degree of Doctor of Philosophy.

August, 1976


Dea College


Agri lture


Dean, Graduate School


-------
























































UNIVERSITY OF FLORIDA


3 1262 08666 931 3