the effect of dietary phytase and zinc sulphate supplementation … · 2017. 4. 19. · recommended...

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The Effect of Dietary Phytase and Zinc Sulphate Supplementation on the Performance and Nutrient Utilization

by Broiler Chickens

By: Mutasim Ibrahim Abdalnoor

B.V.Sc`. U of K

Thesis submitted to the faculty of the Animal Production- University of Khartoum

In partial fulfillment of the requirements for the degree of M.Sc. in Animal Production

Supervisor:

Dr. Sami Balla Ibrahim

University of Khartoum Faculty of Animal Production

November 2010

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DEDICATIONS

TO THE MEMORY OF MY DEAR MOTHER, FATHER AND

SISTER

TO MY WIFE AND KIDS

MY BROTHER AND SISTER

MY FRIENDS AND ALL MY FAMILY

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ACKNOWLEDGEMENTS

I would like to express my sincere and heartfelt gratitude to my advisor,

Professor Sami Balla Ibrahim for his insightful guidance and invaluable patience,

support and encouragement during the entire course of this study. Working with

him has been an enriching and holistic learning experience for me and I

wholeheartedly appreciate the time, energy and diligent efforts he has put in with

me in developing this thesis and also his faith in my abilities.

Sincere thanks also go to my co-supervisor Dr.Abubuker Ibrahim Hussien

for his help with the empirical work in this thesis. Without his guidance this thesis

could not have been completed. I would also like to thank Prof.Amer M Saleh for

his kind support and encouragement, also would like to thank Prof. Mohamed

Khair Abdulla for his advice and support. My thanks go to technicians Mr.

Mahjoob M. Saeed and Abdulrahman Alsadiq, for their technical analysis.

I would also like to thank my family for their unconditional love and support

through successes and failures and in all my accomplishments ever.

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My wife, Dr. Lamia Safi Eldieen, has been a constant source of

encouragement, love, understanding and patience especially over long distances.

And off course Dillip Kumar, for all his help with the word processor. I

would have been lost without him to sort out my formatting problems.

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ABSTRACT

An experiment was conducted to examine the effects of feeding sorghum-

based diets supplemented with 40 mg/kg of zinc sulphate (ZnSO4), or non-

supplemented ZnSO4,or 50% or 100% National Research Council’s available

phosphorus requirements with or without phytase on broiler chicken performance.

Six hundred 1-day-old unsexed (Lohman) broiler chicks were fed the experimental

diets for 42 days. The results of this experiment indicated that reduction in

supplementation of phosphorus to 50% National Research Council’s available

phosphorus requirements was possible with phytase supplementation of the

recommended dosage. Phytase supplementation showed significant (P<0.05)

improvement in chicken performance and increased tibia ash content. Dietary level

of 40 mg/kg of Zn appears to be adequate with addition of phytase to 50% reduced

inorganic phosphorus diet.

Key words: phytase, phosphorus, zinc, broilers.

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ملخص البحث

الالحم الدجاج في المعادن وتحويل النمو كفاءة على الزنك و زالفايتي نزيمإ إضافة تأثير

الـذرة من علف على الالحم الدجاج تغذية تأثير لتحديد) والتوازن النمو( نادراست أجريت: البحث موجز

) ZiSO4( الزنـك كبريتات جرام كيلو/ ميكروجرام 40 أو 0 على يحتوى الرفيعة %50 أو100%

أو مضاف) NRC( األمريكي للبحوث القومي المجلس توصيات حسب على المتاح الفسفور احتياجات من

وزن مـن كل في) P<0.05( معنوي فرق وجود الدراسة نتائج أوضحت. زالفايتي إنزيم إليه مضاف غير

السـاق عظمـة رماد أن أوضحت كما. الالحم الدجاج في النسبي العلف وتحويل العلف واستهالك الجسم

نسبة على تحصل زالفايتي إنزيم إضافة مع %50 على تغذى الذي الدجاج وان الغذائية بالمعامالت تأثرت

. زالفـايتي إنـزيم إضافة عدم مع %50 غذاء أعطيت التي تلكب مقارنة للفسفور الظاهري لالحتفاظ عاليه

فـي كفاية أظهر زالفايتي إنزيم إضافة مع) ZiSO4( الزنك كبريتات جرام كيلو/ ميكروجرام 40 إضافة

.الفسفور احتياجات من %50 على يحتوي الذي للعلف اإلضافة عند الدجاج نمو نسبة

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LIST OF CONTENTS

Sl No. Object Page No.

1 Dedication 2

2 Acknowledgment 3-4

3 Abstract 5

6 ملخص البحث 4

5 List of contents 7

6 List of tables 8-9

7 List of Figures 10

8 Acronyms and abbreviations 11

Chapter - One

9 Introduction 12-14

10 Objectives of the study 15

Chapter - Two

11 Literature and review 16-30

Chapter - Three

12 Materials and methods 31-39

Chapter - Four

13 Results 40-57

Chapter – Five

14 Discussions 58-60

15 Conclusions 61

16 References 62-

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LIST OF TABLES

Sl No. Table Subject Page No.

1 1

The Composition Of

The Diet

34

2 2

Calculated

Composition Of The

Diet

35

3 3 Dietary Treatments

36

4 4

Analyzed

Composition Of The

Diet

37

5 5

The effect of dietary

phytase and Zn

supplementation on

the broiler feed intake

(gm/bird/wk)

44

6 6


phytase and Zn

supplementation on

the broiler weight

gain (gm/bird/wk)

47

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7 7

Effect of dietary

phytase and Zn

supplementation on

the broiler feed

conversion ratio

50

8 8


phytase and Zn

supplementation on

Tibia Ash content (%)

53

9 9


phytase and Zn

supplementation on

phosphorus retention

56

10 10


phytase and Zn

supplementation on

DM (%) retention

59

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LIST OF FIGURES

Sl No. Figure Page No.

1

The effect of dietary phytase and Zn supplementation on the

broiler feed intake (gm/bird/wk)

1a-45 1b- 46

2

The effect of dietary phytase

and Zn supplementation on the

broiler weight gain

(gm/bird/wk)

2a- 48 2b- 49

3

Effect of dietary phytase and

Zn supplementation on the

broiler feed conversion ratio

3a- 51 3b- 52

4


and Zn supplementation on

Tibia Ash content (%)

4a- 54 4b- 55

5


and Zn supplementation on

phosphorus retention

5a- 57 5b- 58

6

The effect of dietary phytase and Zn supplementation on

DM (%) retention

6a- 60 6b- 61

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ACRONYMS AND ABBREVIATIONS

ANOVA analysis of variance

ASA American Soybean Association

Ca calcium

CP crude protein

DM dry matter

F feed intake

G weight gain

G.N.C. ground nut cake

I.U international unit

NRC National Research Council

Pi inorganic phosphorus

PH hydrogen ion concentration

SEM standard error of the mean

SPSS statistical package for social studies

T.P. total phosphorus

ZnSo4 zinc sulphate

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CHAPTER ONE

INTRODUCTION:

Poultry is a good source of high quality protein in form of meat and

eggs. However, feed accounts about 70% of the cost of poultry production.

Recent research tended towards using non conventional feed resources as

additives in order to maximize the production and reduce the cost. Feed

ingredients, such as cereal grains and oil seed cakes, used in broiler diets

contain high amounts of organic phosphorus in the form of phytate. Much of

the phytate phosphorus is voided in the excreta because of the lack of the

enzyme phytase in the chickens' digestive system.

High cost inorganic phosphorus sources ($ 3 kg-Pi), such as dicalcium

phosphate, are used to supplement poultry feeds to ensure that the

requirement for this mineral is met. Most recently, the addition of dietary

enzymes, produced by biotechnology, to broiler feed has been investigated.

These enzymes have potential to increase the use of phytate from the broiler

feedstuffs. Phytate phosphorus represents approximately 70-90% of the total

phosphorus found in cereal grains. The addition of phytase enzyme will,

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therefore, liberate this phytate phosphorus from the cereal grains for animal

use.

It is estimated globally $2 billion in feed costs is saved by using phytase in

animal feeds for releasing phosphorous from innate plant phytate. Whereas,

it is expected another $2 billion is to be saved through further improvements

of the enzyme and additional benefits of phytase on animal performance.

Phytate is also an anti-nutrient. If destroyed, the performance of the animal

will improve. This way the phytase enzyme also is an performance enhancer.

The source of phytate and phytate availability is one of the items that

influence phytase efficacy as well as the Ca/P ratio in the diet, where high

calcium has a direct effect on the efficacy of phytase. High Ca reduces

phytate solubility and thus phytase activity.

The phytate location is also of importance since it affects the ability of

phytase and phytate to interact. Phytate can be located within cell walls and

protein structures. In corn it is for more than 80% located in the corn germ.

In wheat phytate is located in the aleurone layer.

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Supplementary enzymes can improve phytase efficacy if they open up cell

walls. However if endogenous phytase present in the plant, then added

phytase has little effect.

The additional contribution of phosphorous from phytate is depending on the

phytate level of the other raw materials and the interaction between phytase

and other pro-nutrients. Furthermore, phytase can only supply the promised

nutrients if there is enough phytate and if this phytate is sensitive to

enzymatic degradation

Zinc incorporates most of the biological reactions in the animal body

rendering it critical for growth of both somatic and germinal cells. Hence it

is a functional compound of several enzyme systems including most of the

enzymes of the digestion process.

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OBJECTIVE OF THE STUDY

The objective of this study was to evaluate the effects of dietary addition of

the enzyme phytase, and zinc alone or in different combinations on broiler

chicken performance.

The study was designed to evaluate nutrient utilization by young broiler

chickens.

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CHAPTER TWO

LITERATURE AND REVIEW:

Feed ingredients, such as cereal grains and soybeans, used in broiler diets

contain high amounts of organic phosphorus in the form of phytate. Much of the

phytate phosphorus is voided in the excreta because of the lack of the enzyme

phytase in the chickens' digestive system. High cost inorganic phosphorus sources

($ 3 kg-1P), such as dicalcium phosphate, are used to supplement poultry feeds to

ensure that the requirement for this mineral is met.

Nutrition and management in conjunction with biotechnology may offer

some solutions to improve utilization of the components of the feedstuffs

containing the phytate phosphorus. Most recently, the addition of dietary enzymes,

produced by biotechnology, to broiler feed has been investigated. These enzymes

have potential to increase the use of phytate from the broiler feedstuffs.

1. Phosphorus

1.1 Phosphorus importance:

Phosphorus is an indispensable part of every cell of the body. It plays a

distinctive role in maintaining the neutrality of the blood and it reacts chemically

with other nutrients proteins, fats and carbohydrates, to provide the body with the

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energy and material needed for growth and maintenance. Calcium and phosphorus

are both necessary for normal response of nerves to stimulation and to action of all

body muscles.

Calcium and phosphorus are equally important to body structure. During

digestion ,calcium and phosphorus are separated from the foods eaten and are

eventually absorbed through in the intestinal wall in to the blood stream, which

carries them to the areas of the body where they are needed (Ethal, 1966).

Phosphorus is an essential component of organic compounds involved in

almost every aspect of metabolism.

Phosphorus plays an important part in muscle, energy, and carbohydrate, and

amino acid, fat and nerve tissue metabolism; in normal blood chemistry; and in

skeletal growth. Phosphate is an important part of nucleic acid, DNA and RNA; it

is component of many coenzymes and is found in compounds such as adenosine di

and tri-phosphate.

Phosphorus is an essential element classified as a macronutrient because of

the relatively large amounts of phosphorus required by plants. Phosphorus is one of

the three nutrients generally added to soils in fertilizers. One of the main roles of P

in living organisms is in the transfer of energy. Organic compounds that contain

phosphorus are used to transfer energy from one reaction to drive another reaction

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within cells. Adequate phosphorus availability for plants stimulates early plant

growth and hastens maturity. Although phosphorus is essential for plant growth,

mismanagement of soil phosphorus can pose a threat to water quality. The

concentration of phosphorus is usually sufficiently low in fresh water so that algae

growth is limited. When lakes and rivers are polluted with phosphorus, excessive

growth of algae often results. High levels of algae reduce water clarity and can lead

to decreases in available dissolved oxygen as the algae decays, conditions that can

be very detrimental to game fish populations (Busman et al., 2009).

1.2 Phosphorus and Calcium requirements for boilers:

The normal contents of starter diets are 10g calcium and 4.5g available

phosphorus/Kg (in an approximate ratio of 2:1) (Whitehead, 2002). According to

Avian Farms (1999), calcium levels in starter and grower diets should be 0.9-1%

and 0.85-0.95%, respectively. At the same time available phosphorus levels of

0.47-0.50% and 0.42-0.47% are recommended for chick starter and grower diets,

respectively. Temperatures above the thermo neutral zone tend to reduce feed

consumption and hence result in increased mineral requirements when expressed

as a portion of the diet (Underwood & Suttle, 1999). According to Fisher (1983),

high levels of calcium can also depress feed intake and egg production.

Hatchability also increases as dietary phosphorus levels were increased from 0.3 to

0.5%. Menge et al. (1977) reported similar results in turkey hens. Furthermore,

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Shafey & McDonald (1991) observed that increasing dietary calcium from 0.89 to

2.33 % reduced BGW, feed intake, tibia calcium content and plasma phosphorus;

while it increased feed conversion ratio and plasma total calcium in broiler

chickens. High calcium intake leads to the formation of kidney stones (Marynard et

al.m 1979). Calcium mobilization and storage is also under the influence of

oestrogen hormones.

1.3 Deficiency symptoms of phosphorus and calcium in broilers:

Calcium deficiency may occur because of a low dietary calcium level or

excess dietary phosphorus. A low calcium level, which usually implies abnormal

parathyroid function, is rarely due to low dietary calcium intake since the skeleton

provides a large calcium reserve for maintaining normal blood levels (Weaver,

2001). Other causes of abnormally low blood calcium levels include chronic

kidney failure, vitamin D deficiency, and blood magnesium levels (Weaver, 2001).

Insufficient vitamin D may cause a secondary calcium deficiency, by impairing

calcium absorption and bone formation. Increasing dietary calcium above the

normal required levels improves but does not completely correct this secondary

deficiency. Heaney (1997) contends that increasing calcium intake beyond the

amount that produces the optimal bone mass will not result in more bone. The

skeleton of an animal normally contains large reserves of both calcium and

phosphorus that can be absorbed without permanent or serious damage. These

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homeostatic mechanisms enable the animal to accommodate temporary shortages

(Todd, 1976). Previous reports (Bar & Hurwitz, 1984; Gillespie, 1987; Schwartz,

1996) suggested that dietary deficiency of either calcium or phosphorus, if

sufficiency prolonged leads to skeletal deformities, retarded growth and decreased

feed intake (depraved appetite). If acute, retarded growth can lead to death (NRC,

1977). Imbalances of Phosphorus, Calcium or vitamin D intakes at any rate can

result in skeletal defects (Todd, 1976) in growing chicks, a calcium deficiency

causes skeletal abnormalities, including rickets, dyschondroplasia , lameness,

enlarged painful joints and misshapen bones as large and uncalcified epiphseal

plate of long bones (Klasing, 1998).

Most calcium related problems are related to excess calcium or insufficient

levels of available phosphorus, as a result, most calcium and/or phosphorus

problems are manifested as a phosphorus deficiency (Schwartz, 1996) in immature

or young flocks, a deficiency of either calcium or available phosphorus or an

imbalance in these nutrients will result in rickets (ASA, 1997). According to an

Todd (1967), the term ‘rickets’ is often used to describe the pathological condition

that occurs in young animals due to defective calcification of the growing bone.

Rickets is a condition in which bones are decalcified and weakened causing

bowing of legs and other problems (Swick, 1995) the growth plate is decreased in

width and birds become sluggish and reluctant to walk. The bones and beaks

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become weak, soft, deformed and easily broken (Swick 1995; ASA, 1997).

Weakness and morbidity are also deficiency symptoms of phosphorus ( Schwarts,

1996)

2. Phytic Acid

Phytate or phytic acid is the main storage form of phosphorus in grains

and oil seeds. Pigs and poultry are unable to digest phytate, as they lack digestive

enzymes that break it down. As a result, a substantial amount of phosphorus is

excreted as waste, with only 14% of the total phosphorus bioavailable in corn

and up to approximately 50% in wheat. NRC(1998), Cromwell GL. (1992). Because

phosphorus is an essential element, inorganic phosphorus, which is highly

available, is typically supplemented in the diet to meet the poultry and pig’s

requirement. Phytate also has other antinutritive effects, as it is known to reduce

the availability and utilization of other nutrients. Cheryan M.(1980), Ravindran V

et al.,( 1999)

3. Phytase

Phytase is an enzyme that specifically acts on phytate, breaking it down to

release phosphorus in a form available to the animal. This greatly reduces the

need for supplemental inorganic phosphorus and improves the nutritional value

of feedstuffs. Phytase activity is expressed as phytase units or FTUs. One FTU is

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the activity of phytase required to liberate 1 µmol of inorganic phosphorus per

minute at pH 5.5 from an excess of 15 M sodium phytate at 37°C. Unlike the

nonstarch-polysaccharide-degrading enzymes, phytase is the only exogenous

enzyme that has been consistently shown to be highly beneficial to pigs. (Selle

and Ravindran , (2008). The proven efficacy of phytase has resulted in worldwide

acceptance and use in pig and poultry production. Sheppy C., (2001)

3.1 Phytase Enzyme

Some ingredients possess intrinsic phytase activity, which varies greatly

among plant species. Corn and soybean meal contain negligible levels of phytase

activity compared to wheat, which contains considerably higher levels of intrinsic

phytase, Eeckhout and, Depaepe. (1994). The majority of phytase activity in cereal

grains is found in the aleurone layers, Oatway, et al.,(2001). However, this may be

lost when ingredients are subjected to high temperatures, such as during the

pelleting process. (Jongbloed and Kemme 1990). Commercially available exogenous

phytases are commonly derived from either fungi or bacteria, such as Aspergillus

niger and Escherichia coli, (Selle and Ravindran. (2008), but can also be expressed

in yeasts. Ciofalo et al.,(2003), Pandey, et al., (2001).

Several factors can influence the efficacy of phytase, including the amount

of phytate in the diet, the amount of phytase added to the diet, and the type of

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phytase. Studies have shown greater responses to phytase in pigs fed diets that

contain higher amounts of phytate. Selle PH, Ravindran V.(2008), Phytase derived

from E coli bacteria is also more efficacious than the fungal phytases in terms of

the amount of phosphorus released per unit of phytase, according to published

data. Augspurger NR, et al (2009), Kornegay ET, Qian H.(1996). However,

analytical techniques being used to determine phosphorus release vary among

commercial phytase manufacturers. Because of this, the amount of phosphorus

released per unit of phytase may differ between two phytase products, as shown

in a recent study using a standard assay procedure. Jones CK (2009). Thus,

depending on the assay used, different results of phytase activity may be

reported.

Because phytase is a protein, it is susceptible to denaturation when

subjected to excessive heat, such as during pelleting. This may be addressed by

spraying liquid phytase onto the cooled pellets to maintain the stability of the

enzyme. In addition, heat-stable phytases are available. Phytase is also sensitive

to degradation when stored in premixes under high temperature and moisture

conditions. Hence, proper storage procedures and frequent rotation of products

containing phytase must be practiced. Phytase products should be stored only in

cool, dark, dry areas. The manufacturer’s recommendations should always be

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followed, especially when phytase is included in vitamins and trace-mineral

premixes.

The calcium-to-phosphorus ratio (Ca:P) of the diet may affect the

magnitude of response to phytase. Studies have shown that wider Ca:P can result

in decreased absorption of phosphorus. Qian H, Kornegay ET, Conner DE Jr.

(1996), Johnston SL, et al (2004). Thus, a range of 1:1 to 1.25:1 Ca:P is

recommended ( Jacela et al. 2010 ).

Trace mineral is most likely to be deficient in animals fed common feed

stuffs such as Fe,Mn,Co,I,Zn,Cu,Se (Underwood et al., 1935). The function of

trace elements acts of enzymes systems.

4. Zinc:

4.1. Zinc distribution:

Zn distributed widely in body tissue, with the highest concentration in liver,

kidney, muscle, pancreas, eye, prostate gland, skin, bone, hair and wool. Deficient

animal retain more Zn in skin, testis, scrotum, kidney, muscle, heart, lung and

spleen than normal animal (Taj Edean 1997).

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4.2 Zinc function:

Zn is functional compound of several enzyme systems, including carbonic

anhydrase, carboxypeperdase A and B ,several dehydrogenases (dehydrogenises) ,

alkaline phosphates ,ribonuclease and DNA polymeraze ( polymerase).

Carbonic anhyderase is found in erythrocytes kidney tubes, gastrointestinal

mucosa and glandular epithelium (Vergil et al., 1970). An important function of

zinc is necessary for the growth of germinal and somatic cells, as well as in the

maturation of spermatozoa ( Underwood et al. 1969). Falchuk (1998) suggested

that zinc determine both the type of mRNA transcripts synthesized and the rate of

transcription itself.

Kidd et al. (2000) suggested that zinc-methionine supplementation to hen

may aid progeny immune organ development and enhance nonspecific immunity.

Zinc and vitamin C have improved the performance and modulated the humoral

and cellular immune response in broilers (AbuZeid et al. 1999).

Zinc in association with vitamin E increased the humoral immune response

to vital antigens and the cellular immune response (Bettuzzi et al., 1998).

Simon et al. (1987) found that the addition of zinc bactration (50 mg/kg) in

starter feed improved average body weight gain by 1.5% and feed conversion by

2.6% compared with un-medicated controls. Muller (1992) found that zinc

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bactration (an antibiotic bound with zinc), proved and effective, exerting a positive

influence on digestion by selective, antibacterial action against streptococcus,

staphylococcus and clostridium. Bee et al. (1998) showed that the dietary antibiotic

zinc-bactration increased energy utilization only in the first growth period of

broiler.

4.3 Zinc metabolism:

4.3.1. Absorption:

Regulation of zinc absorption occurs in the intestinal cells (Furugouri,

1978). Metabolism of trace elements is subjected to certain mechanism of home

static regulation, and the supply status of zinc has definite effect on its respective

extent of the intestinal absorption, retention and intestinal extension (Kirchgessner,

1976). Experimental studies showed that there was an increase in the a decrease in

turnover rate and endogenous excretion of zinc in response to deficient dietary

intake (Kirchgessner and Schwarz, 1976).

The absorption of zinc is affected adversely by high dietary Ca

concentration, and the presence of phytate further aggravates it. The complex of

zinc phytate forms an insoluble and unabsorabable compound and is one

mechanism whereby zinc availability to animals is reduced. A chelating agent ,

ethylene diamine tetra acetate (EDTA),improves zinc availability by competing

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with phytate to form an EDTA-Zn complex that allows adsorption of zinc either of

free ion or as EDTA-Zn complex (Furugouri,1978).

The presence of low molecular weight binding ligands such as histidine and

system in feed stuff such as soybean Meal and corn was found decrease Zn-

availability (Furugouri,1978).

Solmons and Cousins (1983) suggested that diet containing similar amounts

of Zn may produce available incidence of parakeratosis in swine and variable

weight gain in chicks.

4.3.2 Excretion:

The endogenous losses are reduced during dietary zinc deficiency, serving to

conserve body stores. Except in abnormal conditions such as nepherosis (kidney

disorder) or hypertension (rise in blood pressure), urinary losses of zinc are very

low. Excretion can be increased 10 folds by administration of EDTA

(Underwood,1977).

4.3.3. Zinc homeostasis:

Transfer of zinc out of the intestinal mucosal cells to the plasma is controlled

by metallothionein, while is a low molecular weight binding protein which is

synthesized in response to arise in plasma zinc concentration. Thus the overall

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process of zinc absorption appears to be regulated by intracellular

compartmentalization as well as endogenous secretion of zinc in excess of

immediate metabolic needs from the intestinal tract lumen (Solomon and Cousins,

1983).

4.3.4. Zinc retention:

Zinc is removed from the blood rapidly by tissues, tissues saturated with

zinc (muscle) translocate zinc to unsaturated tissues (liver, pancreas, kidney).

Saturated tissues were characterized by no net change in, where as unsaturated

tissues showed a marked increase in zinc content (Pekas, 1968) .

Retention of zinc was highest at each stage of growth in chicks receiving

diets with the highest contents of protein and energy (Baghael and Pradhan,

19990). Higher retention of zinc occurred only in the grower and finisher phases

when giving high level of protein, while level of ME declined zinc retention in

finisher period.

Biological utilization of zinc from organic source is better than from in

organic. It has been revealed that broilers given zinc methionine had a greater level

pf zinc in tissues (60%) than broiler given the inorganic compounds (zinc-sulphate

and zinc carbonate). The degree of utilization of zinc from zinc carbonate was

intermediate between that from sulphate and methionine (Smak and Stncher,1969).

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4.4. The deficiency of zinc:

Feed conversion rate was decreased when the bird were fed below a level of

34 mg/kg. Bird self-selected a dietary zinc level of 32 mg/kg, which was quite

adequate for weight gain.

This is more precise than the amount assessed for zinc requirements by

conventional methods, which is 40 mg/kg (Steinruck and Kirchgessner, 1993).

The most common sign of zinc deficiency is growth retardation and

reduction in plasma alkaline phosphatqase activity and in plasma Zn

concentration. Thickening or hyperkeratosis in swine); goat-like cattle show

parakeratosis which increased bacteria in the mouth, stiffness of joints with

swelling of feet and horn over growth, excessive salivation,small testicles and no

libido (Haenlein, 1980).

Zinc deficiency causes abnormal development of feather in broilers.

Feathers frying occur near the end of the feather. The hock joint may become

enlarged. The long bones of the leg and wings also become shorted and thickened

as a result of zinc deficiency. A deficiency in the breeder diet would reduce egg

production and hatchability. The embryos, in deficient eggs show a wide variety of

skeletal abnormalities involving the head, limbs and vertebrae (Scott et al., 1982)

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4.5. Toxicity:

Although the requirement for most species is less than 50 mg/kg of diet,

levels of 1 – 2.5 have been fed to rates with no deleterious effects (Prasad et al.,

1969) observed anemia, growth failure and death in rats fed 1% Zn, although

lower levels (0.7%) allowed life but induced anemia and reduced growth. Zn fed

at 1g/kg produced depressed growth stiffness, and excessive bone resorption

(Hsu et al 1975).

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CHAPTER THREE

MATERIALS AND METHODS:

3.1. Growth Study:

The experiment was carried out in an open system oriented house, in Kadaru

area, which was located in east-west direction. The houses dimensions were 20m x

5m and 3.5m high. Before the experiment, the house was cleaned and disinfected

using fire and formalin solution, respectively.

A total of six hundred, 1-day-old unsexed, Lohman, broiler chicks were used in

this experiment. The chicks were purchased from the OMAT Company Farm,

located in Omdurman. The broiler chicks were distributed among 24 pens. The

pens were divided into 3 blocks of 8 pens each; each pen had a floor area of 3.50

m2. Feed and fresh water were offered adlibidum for the 42-day experimental

period. 25 chicks were selected randomly and were placed in each of the

experimental pens. The pens were bedded with wood shavings. Each pen was

supplied with a feeder and drinker. Throughout the experimental period light was

available for 24 hrs; 12 hrs on day light and the rest on artificial lighting, using 40

watt bulbs. Chicks immediately after hatching were vaccinated against Marek’s

and Newcastle diseases. At day 12 post- hatching, they were vaccinated against

Infectious Bursitis (Gumboro). The experiment conducted in (March-May). The

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maximum degree of temperature reported through the experimental period, was (44

C) and the minimum one was (25 C).

The average weight of the chicks in each pen was approximately the same. Three

replicates (pens) were randomly assigned to each of the (8) treatments with factors

of:

• 2 levels of dietary (ZiSO4) Zinc sulphate (0 and40 mg kg-1)

• 2 levels of inorganic phosphorus NRC 100% and 50% recommended

level

• With or without addition of phytase (at 100 mg kg-1).

Daily mortalities were removed and recorded and the carcasses saved for post-

mortem examination. Individual body weight of chicks was measured at 1, 7, 14,

21, 28, 35 and 42 days of age. Feed and fresh water were fed adlibitum for the 42-

day experimental period. Feed weigh-backs per pen were determined weekly. Feed

conversion ratio was calculated as the ratio of feed required to produce one Kg of

meat.

[Type text]

3.3. Experimental Diets:

Sorghum-based experimental diets were formulated to meet or exceed the NRC

(1984) nutrient requirements for broiler chickens. Both diets were formulated in

such a way as to have similar nutrient contents. The composition of the

sorghum-based diet is given in Table 1. Each of the experimental diets were

prepared in one tone mixes. From each mix, 8 patches of 100 kg were made and

the appropriate enzyme or enzyme –zinc combinations were added. Diets were

formulated with 2 levels of dietary Zinc Sulphate (0 and 40 mg kg-1).

Two levels of inorganic phosphorus 100% and 50% NRC recommended level

and with or without addition of phytase (at 100 mg kg-1).

All diets were isocaloric, isonitrogenic containing (ME 3200 kcal per kg) and

24% crude protein. Each diet was randomly assigned to three replicate pens.

Feed and water were provided adlibidum. The feed stuffs used in the

experiments were bought from local market. The dietary treatments were

distributed among the pens of chicks according to a randomized complete block

design of three blocks, that each contained a factorial 2 X 2 X 2 arrangement of

treatments.

[Type text]

Table No.: 1; The Composition Of The Diet

The Composition Of The Diet

Ingredients% Diet1 Diet2

Sorghum 58.75 58.75

G.N.C 17.2 5 17.25

Sesame cake 11.75 11.75

Wheat bran 5 5

Concentrates 5 5

Dical Ph 2 0

Lime stone 0 2

Salt 0.25 0.25

Calculated

ME (Kcal/Kg) 3200 3200

CP% 24 24

P% 0.69 0.34

Ca% 0.8 0.8

[Type text]

Table No.: 2; Calculated Composition Of The Diet:

Parameter D1 D2 D3 D4 D5 D6 D7 D8

CP%

24 24 24 24 24 24 24 24

ME 3200 3200 3200 3200 3200 3200 3200 3200

Ca % 1.36 1.36 1.36 1.36 1.36 1.36 1.36 1.36

T. P. 0.68 0.34 0.68 0.34 0.68 0.34 0.68 0.34

Zn mg 40 40 0 0 0 40 40 0

[Type text]

Table No.: 3; Dietary Treatments:

Diet Pi Phytase Zinc

1 100% + +

2 50% + +

3 100 - -

4 50% - -

5 100% + -

6 50% - +

7 100% - +

8 50% + -

[Type text]

Table No.: 4; Analyzed Composition Of The Diet

D1 D2 D3 D4 D5 D6 D7 D8

CP % 22.80 23.1 22.3 23.0 22.9 22.85 22.78 23.0

ME Kcal 3155 3193 3190 3115 3109 3178 3165 3115

T.Pi% 0.63 0.38 0.65 0.34 0.66 0.37 0.69 0.35

Ca%

1.34 1.30 1.28 1.31 1.39 1.37 1.33 1.32

DM% 92 91.9 92.1 91.8 92.2 91.7 92.2 91.9

The feed stuffs used in the experiments were bought from local market.

[Type text]

Digestibility Study:

In addition to the growth study, 72 broilers were used in a digestibility study.

For the first 3 weeks of this study, the broilers were housed in layers cages.

During the third and six weeks of the study, total excreta output per pen, over a

24 hour period was collected. Feed consumption for the same 24 hour period

was also recorded. Excreta samples from each group were weighed (wet

weight), re-weighed (dry weight) and ground. Diets were, also, ground.

At the end of each collection period broilers from each treatment were selected

randomly, sacrificed and left tibias were removed for analysis of ash content.

The retention of DM and total P as percent of the diets was determined as the

total collection method.

The apparent retention of dry matter (DM), and total phosphorus (TP) of the

diets was determined using Apparent Retention (%) = 100 ( a - b)/a

Where:-

a = [Diet intake (DM) X %nutrient in diet]

b = [Excreta voided (DM) X %nutrient in excreta]

Experimental Design And Statistical Analysis:

The experimental design of the trial was 2 x 2 x 2 factorial arrangement in a

completely randomized design. Independent variables used two level of Zinc (0

[Type text]

and 40 mg/kg) and two level of inorganic phosphorus 100% and 50%, with or

without addition of phytase (at 100 mg kg-1).

Data obtained were tabled and subjected to analysis of variance (ANOVA)

using SPSS variance. The least significant difference (LSD) test was used for

treatment means.

[Type text]

CHAPTER FOUR

RESULTS:

Growth Study:

The effect of dietary phytase and Zn supplementation on the broiler feed

intake on weekly feed consumption is shown in table (5). Birds fed diet

supplemented with 50% Pi without phytase or zinc addition had significantly

lower feed intake (P< 0.05) than those fed diet with 100% Pi without

phytase and ZiSo4 addition (Figure 1). Chicks fed the diet 6 supplemented

50% Pi and Zi So4 without phytase addition had significantly lower feed

intake (P< 0.05), than those diet supplemented with 100% Pi with addition

of either phytase or ZiSo4. Feed intake (gm/bird) was increased significantly

(P< 0.001) for the broiler fed diet supplemented with 50% Pi and

supplemented with phytase with or without Ziso4 compared to those fed diet

(4 and6) (figure1) there were no significant difference in feed intake (P>

0.05).

The effect of dietary phytase and Zn supplementation on the broiler weight

gain on weekly weight gain is shown in table (6).

[Type text]

Birds fed diet supplemented with 50% Pi without phytase or zinc addition

had significantly less weight gain (P< 0.05) than those fed diet with 100% Pi

without phytase and ZnSo4 addition (Figure 2).

Chicks fed the diet 6 supplemented 50% Pi and Zn So4 without phytase

addition had significantly less weight gain (P< 0.05), than those diet

supplemented with 100% Pi with addition of either phytase or ZnSo4 .

Weight gain (gm/bird) was increased significantly (P< 0.001) for the broiler

fed diet supplemented with 50% Pi and supplemented with phytase with or

without Ziso4 compared to those fed diet (4 and6) (figure2). There were no

significant differences in weight gain (P> 0.05).

The effect of dietary phytase and Zn supplementation on the broiler feed

conversion ratio is shown in table (7). Birds fed diet supplemented with 50%

Pi without phytase or zinc addition had significantly smaller feed conversion

ratio (P< 0.05) than those fed diet with 100% Pi without phytase and ZnSo4

addition (Figure 3). Chicks fed the diet 6 supplemented 50% Pi and Zn So4

without phytase addition had significantly lower feed conversion ratio (P<

0.05), than those diet supplemented with 100% Pi with addition of either

phytase or ZnSo4 .

[Type text]

Feed conversion was increased significantly (P< 0.001) for the broiler fed

diet supplemented with 50% Pi and supplemented with phytase with or

without Znso4 compared to those fed diet (4 and6) (figure3) there were no

significant differences in feed conversion ratio (P> 0.05).

Retention Study:

There were significant differences (P<0.05) in diet effects for excreta P, and

apparent P retention (table 9, Figure 5). There was a significant (p <0.05)

dicalcium phosphate x phytase interaction affecting the P content of the

excreta from both collections (figure5).

The excreta samples were collected from birds at 3 and 6 weeks of age. The

results of the balance data are shown in Tables 8,9 and 10. When dicalcium

phosphate was at 50% of control, there was a trend for the P content of the

excreta to decline as the level of phytase increased. This was associated with

an increase in apparent P retention.

Detailed DM retention are shown in Table 10. The analysis of variance

revealed that there was a significant dicalcium phosphate x phytase

interaction affecting both DM intake and DM output (Table 10).

A summary of the mean tibia bone ash determinations recorded at three and

six weeks of age is given in Table (Table 8 ). At 3 weeks of age tibia ash

[Type text]

was significantly (p <0.01) affected by the levels of phytase and dicalcium

phosphate. Broilers fed the 50% dicalcium phosphate without phytase

supplementation had significantly (p <0.01) the lowest tibia ash content.

Chicks given the 50% dicalcium phosphate with phytase supplemented diet

had significantly a higher (p <0.05) tibia ash content than those fed the diets

supplemented with both 50% dicalcium phosphate and without phytase.

Addition of phytase supplementation to 100% dicalcium phosphate did not

significantly improved tibia ash content (p>0.05).

[Type text]

Table No.: 5; The effect of dietary phytase and Zn supplementation on the broiler feed intake (gm/bird/wk)

Weeks Diet 1 Diet 2 Diet 3 Diet 4 Diet 5 Diet 6 Diet 7 Diet 8

Week 1 116b 112b 118b 66a 106b 72a 109b 113b

Week 2 231b 238b 244b 195a 239b 200a 245b 240b

Week 3 456b 463b 459b 363a 466b 371a 461b 459b

Week 4 489b 493b 487b 397a 499b 367a 494b 491b

Week 5 685b 690b 689b 556a 681b 565a 692b 689b

Week 6 920b 898b 900b 780a 915b 793a 913b 910b

S. E. M. 38 38 38 38 38 38 38 38

S.E.M = Standard error of the Mean

Mean with different superscript within the same raw are significantly different (P≤0.05)

[Type text]

Figure No.: (1a); The effect of dietary phytase and Zn supplementation on the broiler feed intake (gm/bird/wk)

[Type text]

Figure No.: (1b); The effect of dietary phytase and Zn supplementation on the broiler feed intake (gm/bird/wk)

0

500

1000

1500

2000

2500

3000

3500

Diet 1 Diet 2 Diet 3 Diet 4 Diet 5 Diet 6 Diet 7 Diet 8

Week 1 Week 2 Week 3 Week 4 Week 5 Week 6

[Type text]

Table No.: 6; The effect of dietary phytase and Zn supplementation on the broiler weight gain (gm/bird/wk)


Week 1 48.13b 46.86b 50b 25.68a 44.16b 27.85a 46.38 47.67

Week 2 93.14b 96.35b 100.41b 73.86a 97.15b 74.62a 101.23 98.36

Week 3 178.82b 182.28b 183.6b 133.94a 184.18b 134.90a 186.63 182.86

Week 4 186.64b 188.88b 189.49b 142.80a 191.92b 130.14a 192.96 190.31

Week 5 254.64b 257.46b 260.22b 195.08a 255.05b 195.50a 363.11 260

Week 6 333.3b 326.54b 332.10b 267.12a 333.94b 267.90a 338.14 334.55

S. E. M. 18 18 18 18 18 18 18 18



[Type text]

Figure No.: (2a) The effect of dietary phytase and Zn supplementation on the broiler weight gain (gm/bird/wk)

[Type text]

Figure No.: (2b) The effect of dietary phytase and Zn supplementation on the broiler weight gain (gm/bird/wk)

0

200

400

600

800

1000

1200

1400



[Type text]

Table No.: 7; Effect of dietary phytase and Zn supplementation on the broiler feed conversion ratio




Week 1 2.41b 2.39b 2.36b 2.57a 2.40b 2.61a 2.35b 2.37b

Week 2 2.48b 2.47b 2.43b 2.64a 2.46b 2.68a 2.42b 2.44b

Week 3 2.55b 2.54b 2.5b 2.71a 2.53b 2.75a 2.49b 2.51b

Week 4 2.62b 2.61b 2.57b 2.78a 2.6b 2.82a 2.56b 2.58b

Week 5 2.69b 2.68b 2.64b 2.85a 2.67b 2.89a 2.63b 2.65b

Week 6 2.76b 2.75b 2.71b 2.92a 2.74b 2.96a 2.70b 2.72b

S. E. M. 0.09 0.09 0.09 0.09 0.09 0.09 0.09 0.09

[Type text]

Figure No.: (3a); Effect of dietary phytase and Zn supplementation on the broiler feed conversion ratio

[Type text]

Figure No.: (3b); Effect of dietary phytase and Zn supplementation on the broiler feed conversion ratio

0

0.5

1

1.5

2

2.5

3



[Type text]

Table No.: 8; The effect of dietary phytase and Zn supplementation on Tibia Ash content (%)


Week 3 51.55b 52.69b 52.8b 47.6a 52.10b 47.9a 52.64b 51.99b

S.EM. 0.23 0.23 0.23 0.23 0.23 0.23 0.23 0.23

Week 6 53.2b 53.1b 52.92b 52.46a 53.2b 52.35a 52.05b 52b

S. E. M. 0.27 0.27 0.27 0.27 0.27 0.27 0.27 0.27

Mean with different superscript within the same raw are significantly different (p≤ 0.05).

[Type text]

Figure No.: (4a); The effect of dietary phytase and Zn supplementation on Tibia Ash content (%):

[Type text]

Figure No.: (4b); The effect of dietary phytase and Zn supplementation on Tibia Ash content (%):

0

20

40

60

80

100

120


Week 3 Week 6

[Type text]

Table No.: 9; The effect of dietary phytase and Zn supplementation on phosphorus retention


Week 3 53.26b 52.91b 50.62b 50.21a 52.12b 50.22a 53.63b 54.1b

S.EM. 0.65 0.65 0.65 0.65 0.65 0.65 0.65 0.65

Week 6 38.38b 37.91b 39.8b 40.62a 41.26b 40.60a 41.10b 38.9b

S. E. M. 1.03 1.03 1.03 1.03 1.03 1.03 1.03 1.03

Mean with different superscript within the same raw are significantly different (p≤ 0.05).

[Type text]

Figure No.: (5a); The effect of dietary phytase and Zn supplementation on phosphorus retention:

[Type text]

Figure No.: (5b); The effect of dietary phytase and Zn supplementation on phosphorus retention:

0

10

20

30

40

50

60

70

80

90

100


Week 3

Week 6

[Type text]

Table No.: 10; The effect of dietary phytase and Zn supplementation on DM (%) retention


Week 3 68.21b 69.1b 69.71b 67.01a 68.9b 67.71a 69.1b 69.2b

S.EM. 0.19 0.19 0.19 0.19 0.19 0.19 0.19 0.19

Week 6 72.1b 71.98b 77.2b 70.4a 72.3b 70.19a 77.12b 71.89b

S. E. M. 0.21 0.21 0.21 0.21 0.21 0.21 0.21 0.21



[Type text]

Figure No.: (6a); The effect of dietary phytase and Zn supplementation on DM (%) retention

[Type text]

Figure No.: (6b); The effect of dietary phytase and Zn supplementation on DM (%) retention

0

20

40

60

80

100

120

140

160


Week 6

Week 3

[Type text]

CHAPTER FIVE

DISCUSSIONS:

An excess of phosphorus in the diet increases the phosphorus in the excreta

and exacerbates the environmental pollution potential. The results of this

experiment suggest that good broiler performance can be obtained with a

dietary phosphorus level lower than that currently used by the industry.

The results of the present study indicate that supplementation of broiler diets

with 50% dicalcium phosphate of control level and without added phytase

were not sufficient for adequate bone mineralization in broilers at 3 or 6

weeks of age. However, addition of phytase to the diets with 50% dicalcium

phosphate of the control level increased tibia ash content indicating that

phytase released some P from phytate in the diet. The results also indicate

that levels of dicalcium phosphate of 100% of the control level with or

without phytase supplementation was adequate for bone mineralization.

Supplementation of broiler diets with phytase resulted in enhanced

utilization of the organic P present in the feed ingredients of plant origin

components. At some dicalcium phosphate levels, supplementation with

phytase was beneficial, in terms of dietary P utilization, while at other level

it was not. With phytase supplementation it was possible to reduce the level

[Type text]

of dicalcium phosphate included in the diet to 50% of control levels.

Broilers on such a diet had two-thirds the total dietary P output of those on

the control diet(Reference). Schoner and Hoppe (1992) and Ibrahim et. al.,

(2002) reported a 50% reduction in the P output by broilers with the use of

phytase. The extent to which phytase supplementation can reduce P

excretion will depend on the composition of the diets, in particular the level

of phytate P.

Several workers (Ibrahim et. al., 2002; Broz et al. 1994) have

demonstrated that phytase supplementation can be used to improve phytate P

utilization such that the use of inorganic phosphate supplementation is no

longer required. In the current experiment, phytase supplementation allowed

for a 50% reduction in the level of dicalcium phosphate in the diet. It was

not possible, however, to reduce the level of dicalcium phosphate further

without a loss in performance.

This study also has examined different of dietary zinc with or without

addition of phytase on feed intake, body weight gain, feed conversion ratio

and retention of dry matter, phosphorus and tibia ash content of the broiler

chicks. Zinc was added to the experimental diet at rates zero and 40 mg/kg.

The results indicated that the addition of zinc to broiler diet had no effect on

[Type text]

body weight gain, feed intake and feed conversion ratio. This result is in

agreement with the finding of Siriak and Stanchev (1996) confirming that

the increasing level of zinc from 20 to 40 mg/kg in form of sulphate,

carbonate and methionine had no effect on body weight gain. Contrarily, the

results were in disagreement with findings of Abu Zied et al. (1999) who

found that the addition of zinc had a positive effect on body weight gain.

Similar results reported by Mukhtar (2002). This variation may be related to

the type of the basal diet component used.

The result indicated that the dietary zinc had no effect on feed conversion

ratio. This is confirmed the finding of Tonchera et al. (1999), who found that

addition of 40 and 60 mg/kg of zinc, had no effect on feed conversion ratio.

However, the results contradicted the findings of Schrez et al. (1993) who

suggested that zinc bacticiracin increased the feed conversion ratio.

Retention of dry matter, phosphorus and tibia ash content of the broiler

chicks were not affected by addition of zinc sulphate which in agreement

with findings of Baghael and Pradhan (1990).

[Type text]

CONCLUSIONS

The results from this experiment suggested that:

• 50% reduction in supplementation of broiler diet with inorganic

phosphorus is possible with addition of phytase enzyme to the diet.

• Supplementation with phytase was shown to improve chicken

performance.

• Dietary level of 40 mg/kg of Zn appears to be adequate with addition of

phytase to 50% reduced inorganic phosphorus diet.

Addition of phytase to the diet with 50% dicalcium phosphate increase Tibia Ash

content indicating that phytase released some phosphorus from phytate in the diet.

[Type text]

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