Improving the utilisation of phytate-bound phosphorus in feed for poultry and pigs through increased efficacy of exogenous phytase in the anterior digestive tract

Philosophiae Doctor (PhD) Thesis 2020:76

Siril Kristoffersen

Improving the utilisation of phytate-bound phosphorus in feed for poultry and pigs through increased efficacy of exogenous phytase in the anterior digestive tract

Økt utnyttelse av fytinsyrebundet fosfor i fôr til fjørfe og gris gjennom økt fytaseaktivitet i fremre del av fordøyelsessystemet

Philosophiae Doctor (PhD), Thesis 2020:76Siril Kristoffersen

Norwegian University of Life Sciences Faculty of Biosciences

Department of Animal and Aquacultural Sciences

Improving the utilisation of phytate-bound phosphorus in feed for poultry and pigs through increased efficacy of

exogenous phytase in the anterior digestive tract

Økt utnyttelse av fytinsyrebundet fosfor i fôr til fjørfe og gris gjennom økt fytaseaktivitet i fremre del av fordøyelsessystemet

Philosophiae Doctor (PhD) Thesis Siril Kristoffersen

Norwegian University of Life Sciences Faculty of Biosciences

Department of Animal and Aquacultural Sciences

Ås/Varhaug 2020

Thesis number 2020:76 ISSN 1894-6402 ISBN 978-82-575-1777-9

ʹ

Supervisors

Prof. Birger Svihus

‡’ƒ”–‡–‘ˆ‹ƒŽƒ†“—ƒ…—Ž–—”ƒŽ…‹‡…‡•

‘”™‡‰‹ƒ‹˜‡”•‹–›‘ˆ‹ˆ‡…‹‡…‡•

‘•–„‘•ͷͲͲ͵

ǦͳͶ͵ʹ%•ǡ‘”™ƒ›

Ass. Prof. Nils Petter Kjos

‡’ƒ”–‡–‘ˆ‹ƒŽƒ†“—ƒ…—Ž–—”ƒŽ…‹‡…‡•

‘”™‡‰‹ƒ‹˜‡”•‹–›‘ˆ‹ˆ‡…‹‡…‡•

‘•–„‘•ͷͲͲ͵

ǦͳͶ͵ʹ%•ǡ‘”™ƒ›

Dr. Torger Gjefsen

‡ŽŽ‡•ŒÞ’‡–‘‰ƒŽƒ†‰†‡”

‘•–„‘•ʹͲͺ‡–”—

ǦͶͲͲͳ–ƒ˜ƒ‰‡”ǡ‘”™ƒ›

Evaluation Committee

Prof. Øystein Ahlstrøm

‡’ƒ”–‡–‘ˆ‹ƒŽƒ†“—ƒ…—Ž–—”ƒŽ…‹‡…‡•

‘”™‡‰‹ƒ‹˜‡”•‹–›‘ˆ‹ˆ‡…‹‡…‡•

‘•–„‘•ͷͲͲ͵

ǦͳͶ͵ʹ%•ǡ‘”™ƒ›

Prof. Damian Józefiak

‘œƒ‹˜‡”•‹–›‘ˆ‹ˆ‡…‹‡…‡•

‡’ƒ”–‡–‘ˆ‹ƒŽ—–”‹–‹‘

ŽǤ‘ϛѕƒ͵͵

͸ͲǦ͸͵͹‘œƒÑǡ‘Žƒ†

Dr. Hilde Schøyen ‹•¤ÞŽŽ‡

‹•¤˜‡‰‡ͳͲͳͲ ǦͶͳʹͲƒ—ǡ‘”™ƒ›

͵

Acknowledgements

Š‡™‘”’”‡•‡–‡†‹–Š‹•–Š‡•‹•™ƒ•…ƒ””‹‡†‘—–ƒ•ƒ‹†—•–”‹ƒŽŠƒ––Š‡‘”™‡‰‹ƒ

‹˜‡”•‹–›‘ˆ‹ˆ‡•…‹‡…‡•ȋȌǤ›‡’Ž‘›‡”Šƒ•„‡‡ ‡ŽŽ‡•ŒÞ’‡–‘‰ƒŽƒ†‰†‡”

ȋ ȌǤ‘™‡˜‡”ǡƒ…‘•‘”–‹—‘ˆ‘”‰ƒ‹•ƒ–‹‘•ƒ†…‘’ƒ‹‡•‹–Š‡ƒ‰”‹…—Ž–—”ƒŽˆ‹‡Ž†

‹‘‰ƒŽƒ†Šƒ•‰‹˜‡ˆ‹ƒ…‹ƒŽ•—’’‘”––‘–Š‹•™‘”ǤŠ‡…‘•‘”–‹—…‘•‹•–•‘ˆ ǡ ‹•¤ÞŽŽ‡ǡ‘”–—”ƒǡ‡–‘Ž–‡ƒ‡ǡ ƒ–Žƒ†ǡ”‹ƒ§”‡ǡǦ‡‹‡”‹‡‡ǡǡ‘‰ƒŽƒ†

ŒÞ”ˆ‡Žƒ‰ǡ‘”•˜‹Þ”Ǧ‡•–ƒ†‘‰ƒŽƒ†‘†‡Žƒ‰ˆ—†‡†–Š‹•‹†—•–”‹ƒŽŠ–‘‰‡–Š‡”

™‹–Š–Š‡‘”™‡‰‹ƒ‡•‡ƒ”…Š‘—…‹Žȋ‰”ƒ–‘ǤʹͷͻͺͶͻȀ͵ͲȌǤ™‘—Ž†Ž‹‡–‘–Šƒ–Š‡

ƒŽŽˆ‘”–Š‡ˆ‹ƒ…‹ƒŽ•—’’‘”–‹–Š‹•’”‘Œ‡…–Ǥ‘”‡‘˜‡”ǡƒ•’‡…‹ƒŽ–Šƒ›‘—–‘‡”ƒ”ƒŽ†ˆ‘”

‹‹–‹ƒ–‹‰–Š‹•’”‘Œ‡…–ƒ†–Šƒ–‰‘––Š‡‘’’‘”–—‹–›–‘•–—†›ˆ‘”–Š‹•ŠǤ

›‰”‡ƒ–‡•––Šƒ•–‘›•—’‡”˜‹•‘”•‹”‰‡”˜‹Š—•ǡ‹Ž•‡––‡”Œ‘•ƒ†‘”‰‡”Œ‡ˆ•‡

ˆ‘”ƒŽŽ–Š‡Š‡Ž’ˆ—Ž†‹•…—••‹‘•ƒ†‰—‹†ƒ…‡›‘—Šƒ˜‡’”‘˜‹†‡†‡†—”‹‰–Š‹•Š’”‘Œ‡…–Ǥ

‘”‡‘˜‡”ǡˆ‘”–Š‡‡…‘—”ƒ‰‡‡–ƒ†‹•’‹”ƒ–‹‘–‘…‘–‹—‡™‹–Š–Š‡’”‘‰”‡••™Š‡

Šƒ˜‡ˆ‡Ž–Ž‘•–Ǥ

™‘—Ž†Ž‹‡–‘–ŠƒƒŽŽ‘ˆ›…‘ŽŽ‡ƒ‰—‡•‹ –Šƒ–Šƒ˜‡•Šƒ”‡†Ž—…Š–ƒŽ•™‹–Š

†—”‹‰–Š‡•‡›‡ƒ”•Ǥ•’‡…‹ƒŽŽ›–Šƒ›‘—–‘ƒ”–‡ǡ‰”‹†ǡƒ”‹ƒǡ‘‡ǡ‡‹‰ƒ†ƒ†‡

ˆ‘” –Š‡ —…Š ‡‡†‡† •‘…‹ƒŽ‹•‹‰ ƒ† …‘˜‡”•ƒ–‹‘•ǡ „‘–Š ’”‘ˆ‡••‹‘ƒŽ ƒ† ‘– •‘

’”‘ˆ‡••‹‘ƒŽǤŠƒ›‘—‘Š‹‰—”†‘–Š‡Žƒ„ˆ‘”Ž‡––‹‰‡—•‡ƒŽŽ–Š‡ˆƒ…‹Ž‹–‹‡•ƒ†

‡“—‹’‡–™Š‡‡˜‡”ƒ•‡†ˆ‘”‹–Ǥ™ƒ––‘–Šƒ‡˜‡”›„‘†›™Š‘™‡”‡‹˜‘Ž˜‡†‹–Š‡

’”‘†—…–‹‘ƒ†’ƒ…‹‰‘ˆ›‡š’‡”‹‡–ƒŽˆ‡‡†ǤŠƒ›‘—–‡‹ǡ”‹Ž†ƒ†‡‹Ž‹Šˆ‘”–Š‡

Š‡Ž’™‹–Š’Žƒ‹‰ƒ†‘’–‹‹•ƒ–‹‘‘ˆ–Š‡†‹‡–•Ǥ

Šƒ•–‘›…‘Ǧƒ—–Š‘”•ƒ––Š‡‘œƒ‹˜‡”•‹–›‘ˆ‹ˆ‡…‹‡…‡•ˆ‘”’”‘˜‹†‹‰–Š‡–‘‘Ž•

ƒ†‡“—‹’‡–ˆ‘”•ƒ’Ž‡…‘ŽŽ‡…–‹‘ǡŠ‡Ž’†—”‹‰†‹••‡…–‹‘ƒ†…ƒ””›‹‰‘—–Žƒ„‘”ƒ–‘”›

ƒƒŽ›•‹•Ǥ Šƒ ›‘— –‘ ƒŽŽ •–ƒˆˆ ƒ– –Š‡ ‡š’‡”‹‡–ƒŽ ˆƒ”• ‹ Ž•œ‘™ƒǡ ‘”œ›Ñ ƒ† %•Ǥ

•’‡…‹ƒŽŽ›ƒ”–ƒƒ…Š‘…ƒǡƒ”–ƒ—„‹äƒ†‘Ž˜‡‹‰‡˜‘Ž†‡ˆ•—ˆ‘”Šƒ˜‹‰…‘–”‘Ž‘ˆ

–Š‡ƒ‹ƒŽ•ƒ†ƒ‹‰•—”‡–Šƒ––Š‡‡š’‡”‹‡–•™‡–ƒ•’Žƒ‡†Ǥ

Šƒ›‘—–‘ƒŽŽ›ˆ‡ŽŽ‘™ŠǤǤ•–—†‡–•™Š‘Šƒ˜‡–ƒ‡…‘—”•‡•–‘‰‡–Š‡”™‹–Šˆ‘”–Š‡

Š‡Ž’ˆ—Ž†‹•…—••‹‘•ƒ†•‘…‹ƒŽ‹•‹‰™Š‡Šƒ˜‡„‡‡ƒ–%•Ǥ•’‡…‹ƒŽ–Šƒ‰‘‡•–‘›‘—

ŠƒŽ‡†ǡˆ‘”Š‡Ž’‹‰‡™‹–Š–Š‡’Žƒ‹‰‘ˆƒ†‡š‡…—–‹‘‘ˆ–Š‡†‹••‡…–‹‘‹‡š’‡”‹‡–

Ͷ

Ǥ‘”‡‘˜‡”ǡˆ‘”•Šƒ”‹‰ƒŽŽ‘—”‘™Ž‡†‰‡ƒ„‘—–†‘‹‰‡š’‡”‹‡–•‹Ž•œ‘™ƒƒ†Š‘™

‹–‹•–‘Šƒ˜‡‹”‰‡”ƒ•ƒ•—’‡”˜‹•‘”Ǥ

Šƒ›‘—ƒ‰”‹•–‘ˆˆ‡”ǡ‹Ž•‡––‡”ƒ† ”ƒˆ‘”–Š‡Š‡Ž’™‹–Š–Š‡†‹••‡…–‹‘‘ˆ’‹‰•‘

%•‰¤”†Ǥ„‹‰–Šƒ›‘—‰‘‡•–‘—”—ǡƒ”ƒŽ†ǡŠƒŽ‡†ƒ† ”ƒ™Š‘Š‡Ž’‡†‡™‹–Š–Š‡

†‹••‡…–‹‘‹Ž•œ‘™ƒǤŠƒ›‘—ˆ‘”ƒŽŽ–Š‡’”ƒ…–‹…ƒŽŠ‡Ž’ǡ„—–ƒŽ•‘–Š‡‹…‡…‘’ƒ›™Š‹Ž‡

–”ƒ˜‡ŽŽ‹‰Ǥ

•’‡…‹ƒŽ–Šƒ›‘—‰‘‡•–‘ ”ƒˆ‘”Ž‡ƒ”‹‰‡ƒŽ‘–ƒ„‘—–’‡ƒ•—”‡‡–•ǡƒƒŽ›•‡•

ƒ††‹••‡…–‹‘™‘”ǤŠƒ›‘—ˆ‘”ƒŽ™ƒ›••ƒ›‹‰›‡•™Š‡Šƒ˜‡„‡‡ƒ–Š‘‡ƒ†

‡‡†‡†•‘‡‘‡–‘Š‡Ž’‡™‹–Š’”ƒ…–‹…ƒŽ‹–‹‡•ƒ–ǡƒ†Žƒ•–„—–‘–Ž‡ƒ•–ˆ‘”„‡‹‰ƒ

Ž‘˜‡Ž› –”ƒ˜‡ŽŽ‹‰ …‘’ƒ‹‘ ™Š‡ ™‡ –”ƒ˜‡ŽŽ‡† –‘ ‘”œ›ÑǤ  ƒ††‹–‹‘ǡ ™‘—Ž† Ž‹‡ –‘

–Šƒ –Š‡ ”‡•– ‘ˆ –Š‡ ƒ„‡ ‡„‡” ƒ– Ǥ •’‡…‹ƒŽŽ› Ž‹ ‘Š•‡ ƒ† ƒ‡ ˆ‘”

Š‡Ž’‹‰‡†‡˜‡Ž‘’‡™‡–Š‘†•ƒ†™‹–Šˆ‹†‹‰•‘Ž—–‹‘•‘Š‘™–‘ƒƒŽ›•‡•ƒ’Ž‡•

–Šƒ–™‡”‡™ƒ›–‘‘•ƒŽŽǤŠƒ›‘—–‘ ؔ‡ǡƒ†‡•’‡…‹ƒŽŽ›•‡–ˆ‘”’”‘†—…–‹‘‘ˆ–Š‡

ˆ‡‡†ǡƒ†ˆ‘”ƒŽ™ƒ›•Šƒ˜‹‰ƒ’”ƒ…–‹…ƒŽ•‘Ž—–‹‘–‘‡˜‡”›•—‰‰‡•–‹‘ƒ†…ŠƒŽŽ‡‰‡Ǥ

™ƒ––‘–Šƒ›’ƒ”‡–•ƒ†•‹„Ž‹‰•ˆ‘”ƒŽŽ–Š‡•—’’‘”–ƒŽ‘‰–Š‹•Œ‘—”‡›Ǥ˜‡–Š‘—‰Š

™‡ ƒ”‡ ˆƒ” ƒ’ƒ”–ǡ ›‘— ƒ”‡ ƒŽ™ƒ›• –Š‡”‡ ˆ‘” ‡Ǥ ‘ › ‹ǦŽƒ™• ˆ‘” ƒŽŽ –Š‡ •—’’‘”– ƒ†

’”ƒ…–‹…ƒŽŠ‡Ž’•‘–Šƒ––”ƒ˜‡ŽŽ‹‰ƒ†™‘”‹‰‘††–‹‡•™ƒ•’‘••‹„Ž‡ǡ–Šƒ›‘—Ǥ

Šƒ›‘—Žƒ˜ˆ‘”„‡‹‰•—’’‘”–‹˜‡ƒ†ˆ‘”‘˜‹‰™‹–Š‡–‘%••‘…‘—Ž†‰‘–Š”‘—‰Š

™‹–Š–Š‹•’”‘Œ‡…–™‹–Š‘—–Šƒ˜‹‰–‘„‡ƒ’ƒ”–ˆ”‘›‘—ƒ†‘—”‹†•–‘‘—…ŠǤŠƒ›‘—

ˆ‘”‡‡’‹‰—’™‹–Š‡‹ƒŽŽ–Š‡–”ƒ˜‡ŽŽ‹‰ƒ†•–”‡••ˆ—Ž’‡”‹‘†•Ǥƒ•–„—–‘–Ž‡ƒ•––Šƒ

›‘—Žƒ—•ƒ†‡Ž‹—•ˆ‘”„‡‹‰›‘—ǡ›‘—ƒ”‡–Š‡„‡•–ᇡ

‹”‹Ž”‹•–‘ˆˆ‡”•‡

ƒ”Šƒ—‰ǡ‘˜‡„‡”ʹͲʹͲ

ͷ

Table of contents

Acknowledgements ... 3

Abbreviations ... 7

Summary ... 9

Sammendrag ... 11

List of publications ... 13

1. Introduction ... 15

1.1 Phytate ... 15

1.2 Phytase ... 17

1.3 Place for exogenous phytase activity ... 20

1.4 Limitations in phytate degradation ... 22

1.5 Acid and phytase ... 22

2. Aims of the thesis ... 24

3. Methodology ... 25

3.1 In vitro test ... 25

3.2 Experiment I ... 25

3.3 Field assessment ... 26

3.4 Experiment II ... 26

3.5 Experiment III ... 27

4. Main results and discussion ... 28

4.1 Retention time and pH in the anterior digestive tract ... 29

4.2 Phytate degradation and P digestibility ... 31

4.3 Limitations and future perspectives ... 33

5. Concluding remarks ... 35

6. References ... 36

Paper I ... 45

Paper II ... 81

Paper III ... 105

͸

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Abbreviations

Š‘•’Š‘”—•

ƒ ƒŽ…‹—

‰ ƒ‰‡•‹—

‡ ”‘

 ‹…

— ‘„„‡”

• ‘•‹–‘Ž’Š‘•’Šƒ–‡

”›ƒ––‡”

‡‡†‹–ƒ‡

’’ƒ”‡–Œ‡Œ—ƒŽ†‹‰‡•–‹„‹Ž‹–›

’’ƒ”‡–‹Ž‡ƒŽ†‹‰‡•–‹„‹Ž‹–›

’’ƒ”‡––‘–ƒŽ–”ƒ…–†‹‰‡•–‹„‹Ž‹–›

‘–”‘Ž†‹‡–

‹‡–™‹–Š’Š›–ƒ•‡ƒ††‡†

‹‡–™‹–Šƒ…‹†ƒ††‡†

‹‡–™‹–Š’Š›–ƒ•‡ƒ†ƒ…‹†ƒ††‡†Ǥ

ͺ

ͻ

Summary

Š‡ƒŒ‘”‹–›‘ˆ’Š‘•’Š‘”—•ȋȌ‹ˆ‡‡†•–—ˆˆ•‹•’”‡•‡–ƒ•’Š›–ƒ–‡ǦǤŠ›–ƒ–‡‹•–Š‡•–‘”ƒ‰‡

ˆ‘”‘ˆ‹•‡‡†•ƒ†‰”ƒ‹•ƒ†…‘•‹•–•‘ˆƒ‹‘•‹–‘Ž”‹‰™‹–Š•‹š’Š‘•’Šƒ–‡‰”‘—’•

„‘—† –‘ ‹–Ǥ Š‡ ’Š›–ƒ–‡Ǧ ‹• ‘Ž› ’ƒ”–‹ƒŽŽ› —–‹Ž‹•‡† „› ‘‘‰ƒ•–”‹… ƒ‹ƒŽ•Ǥ Š‡

—†‹‰‡•–‡†‹•‡š…”‡–‡†‹–Š‡ƒ‹ƒŽƒ—”‡ƒ†ƒ›–Š‡Ž‡ƒ†–‘†”ƒ‹ƒ‰‡‘ˆˆ”‘

ƒ‰”‹…—Ž–—”ƒŽŽƒ†ƒ†•—„•‡“—‡–Ž›‡˜‹”‘‡–ƒŽ’‘ŽŽ—–‹‘Ǥ‘”‡†—…‡–Š‡ƒ‘—–‘ˆ

‹ ˆƒ‡…‡• ‡š‘‰‡‘—• ’Š›–ƒ•‡ ‹• ”‘—–‹‡Ž› ƒ††‡† –‘ –Š‡ †‹‡– ‘ˆ ‘‘‰ƒ•–”‹… ƒ‹ƒŽ•Ǥ

‘™‡˜‡”ǡ ‡˜‡ ™‹–Š ’Š›–ƒ•‡ ƒ††‡†ǡ –Š‡ †‡‰”ƒ†ƒ–‹‘ ‘ˆ ’Š›–ƒ–‡ ‹• ‘– …‘’Ž‡–‡Ǥ Š‡

‡ˆˆ‹…ƒ…› ‘ˆ ’Š›–ƒ•‡ ‹• †‡’‡†‡– ‘ ’ Ž‡˜‡Ž• ‹ ƒ††‹–‹‘ –‘ –Š‡ ”‡–‡–‹‘ –‹‡ ‹ –Š‡

•‡‰‡–• ‘ˆ –Š‡ †‹‰‡•–‹˜‡ •›•–‡ ™‹–Š –Š‡ ‘’–‹ƒŽ ’Ǥ Š‡ ‘•– ‹’‘”–ƒ– •‹–‡ ˆ‘”

‡š‘‰‡‘—•’Š›–ƒ•‡ƒ…–‹˜‹–›‹•‹ƒ–‡”‹‘”†‹‰‡•–‹˜‡–”ƒ…–ǡ†—‡–‘–Š‡’Ž‡˜‡Ž•–Š‡”‡Ǥ

‘–‡•––Š‡‡ˆˆ‡…–‘ˆƒ…‹†ƒ††‹–‹‘–‘–Š‡†‹‡–‘–Š‡‡ˆˆ‹…ƒ…›‘ˆƒ‡š‘‰‡‘—•’Š›–ƒ•‡ǡƒ†

™Š‡–Š‡”–Š‡‡ˆˆ‹…ƒ…›™‘—Ž†„‡ˆ—”–Š‡”‹’”‘˜‡†„›ƒ‹…”‡ƒ•‡†”‡–‡–‹‘–‹‡‹–Š‡

ƒ–‡”‹‘” †‹‰‡•–‹˜‡ –”ƒ…–ǡ –Š”‡‡ ƒ‹ƒŽ ‡š’‡”‹‡–• ™‡”‡ …ƒ””‹‡† ‘—–Ǥ  ƒŽŽ ƒ‹ƒŽ

‡š’‡”‹‡–•ǡƒ™Š‡ƒ–Ǧ„ƒ•‡††‹‡–™‹–ŠƒŠ‹‰Š’Š›–ƒ–‡Ǧ…‘–‡–™ƒ•—•‡†ǤŠ‡†‹‡–Šƒ†

‡‹–Š‡”‘ƒ††‹–‹‘ȋȌǡˆ‘”‹…ƒ…‹†ƒ††‡†ȋȌǡͷͲͲ C. braakiiǦ†‡”‹˜‡†’Š›–ƒ•‡

ƒ††‡†ȋȌ‘”„‘–Šƒ…‹†ƒ†’Š›–ƒ•‡ƒ††‡†ȋȌǤƒ††‹–‹‘ǡƒin vitro‡š’‡”‹‡–

™Š‡”‡ –Š‡ ‡ˆˆ‹…ƒ…› ‘ˆ ˆ‘—” …‘‡”…‹ƒŽ ’Š›–ƒ•‡ ’”‘†—…–• ƒ– •‡Ž‡…–‡† ’ Ž‡˜‡Ž• ƒ†

‹…—„ƒ–‹‘ –‹‡• ™ƒ• –‡•–‡†ǡ ƒ† ƒ ˆ‹‡Ž† ƒ••‡••‡– ‘ˆ –Š‡ …”‘’ —•ƒ‰‡ ‹ …‘‡”…‹ƒŽ

„”‘‹Ž‡”…Š‹…‡•™ƒ•…ƒ””‹‡†‘—–Ǥ

Š‡”‡•—Ž–•ˆ”‘–Š‡in vitro ‡š’‡”‹‡–†‡•…”‹„‡†‹ƒ’‡”™ƒ•—•‡†–‘†‡–‡”‹‡™Š‹…Š

’Š›–ƒ•‡ ƒ† ’ Ž‡˜‡Ž• –‘ —•‡ ‹ –Š‡ ‡š’‡”‹‡–ƒŽ †‹‡–•Ǥ  ‡š’‡”‹‡– ǡ ͺͲͲ „”‘‹Ž‡”

…Š‹…‡•‹ͺͲ†‹ˆˆ‡”‡–’‡•™‡”‡‡‹–Š‡”ad libitum‘”‹–‡”‹––‡–Ž›ˆ‡†–Š‡‡š’‡”‹‡–ƒŽ

†‹‡–•ˆ”‘ͳͷ–‘͵͸†ƒ›•‘ˆƒ‰‡Ǥ ”‘‘‡„‹”†’‡”’‡ǡ–‹„‹ƒƒ†…‘–‡–•ˆ”‘Œ‡Œ——

ƒ†‹Ž‡—™‡”‡…‘ŽŽ‡…–‡†Ǥ”‘’…‘–‡–•™‡”‡…‘ŽŽ‡…–‡†ˆ”‘‹–‡”‹––‡–Ž›ˆ‡†„‹”†•ͺͲǡ ͳ͸Ͳ ƒ† ʹͶͲ ‹ ƒˆ–‡” –Š‡ •–ƒ”– ‘ˆ –Š‡ ˆ‡‡†‹‰Ǥ Š‡ ”‡•—Ž–• •Š‘™‡† –Šƒ– –Š‡”‡ ™ƒ• ƒ

’‘•‹–‹˜‡ ‡ˆˆ‡…– ‘ˆ ’Š›–ƒ•‡ ‘ †‹‰‡•–‹„‹Ž‹–› ƒ† „‘‡ ‹‡”ƒŽ‹•ƒ–‹‘Ǥ Ž”‡ƒ†› ƒˆ–‡” ͺͲ

‹—–‡•”‡–‡–‹‘–‹‡‹–Š‡…”‘’ǡ–Š‡”‡™ƒ•ƒ‡ˆˆ‡…–‘ˆƒ…‹†ƒ††‹–‹‘‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›ǡ

™‹–Š ƒ …‘•‹†‡”ƒ„Ž› ‡Šƒ…‡† ’Š›–ƒ–‡ †‡‰”ƒ†ƒ–‹‘ ƒ† ˆ‘”ƒ–‹‘ ‘ˆ Ž‘™‡” ‹‘•‹–‘Ž

’Š‘•’Šƒ–‡Ǧ‹•‘‡”•Ǥ ’”‘Ž‘‰‡† ”‡–‡–‹‘ –‹‡ ‹ –Š‡ …”‘’ ˆ—”–Š‡” ‹…”‡ƒ•‡† –Š‹•

†‡‰”ƒ†ƒ–‹‘ǤŠ›–ƒ•‡‹…”‡ƒ•‡††‹‰‡•–‹„‹Ž‹–›ǡ‹–‡”‹––‡–ˆ‡‡†‹‰‹…”‡ƒ•‡†ƒ’’ƒ”‡–

Œ‡Œ—ƒŽ†‹‰‡•–‹„‹Ž‹–›ȋȌ‘ˆƒ†”‡–‡–‹‘Ǥ

ͳͲ

ƒ’‡”ǡƒˆ‹‡Ž†ƒ••‡••‡–™ƒ•…ƒ””‹‡†‘—––‘ƒ’–Š‡‡š–‡–‘ˆ…”‘’—•ƒ‰‡ƒ†–Š—•

”‡–‡–‹‘–‹‡‹ad libitumˆ‡†…‘‡”…‹ƒŽ„”‘‹Ž‡”…Š‹…‡•ǤŠ‡ƒ••‡••‡–•Š‘™‡†–Šƒ–

ad libitumˆ‡†„‹”†•—•‡–Š‡…”‘’ˆ‘”•–‘”ƒ‰‡–‘ƒŠ‹‰Š‡”†‡‰”‡‡–Šƒ’”‡˜‹‘—•Ž›ƒ••—‡†Ǥ

 –Š‡ …Š‹…‡ ‡š’‡”‹‡– ȋ‡š’‡”‹‡– Ȍǡ –Š‡ •Š‘”–Ǧ–‡” ‡ˆˆ‡…– ‘ˆ ƒ…‹† ƒ††‹–‹‘ ‘

’Š›–ƒ•‡‡ˆˆ‹…ƒ…›‹–Š‡…”‘’™ƒ••–—†‹‡†Ǥ–ʹͲƒ†ʹͳ†ƒ›•‘ˆƒ‰‡ǡ•–ƒ”˜‡†„‹”†•™‡”‡ˆ‡†

ˆ‘”‘‡Š‘—”ǡƒ†…”‘’ƒ†‰‹œœƒ”†…‘–‡–•™‡”‡…‘ŽŽ‡…–‡†‡˜‡”›ʹͲ‹—–‡•—–‹ŽͳͶͲ

‹—–‡•ƒˆ–‡”–Š‡•–ƒ”–‘ˆ–Š‡ˆ‡‡†‹‰Ǥ–Š‡„‹”†•‡—–Šƒ‹•‡†͸Ͳƒ†ͳͶͲ‹—–‡•ƒˆ–‡”

–Š‡•–ƒ”–‘ˆ–Š‡ˆ‡‡†‹‰ǡ…‘–‡–•ˆ”‘Œ‡Œ——ƒ†‹Ž‡—™‡”‡…‘ŽŽ‡…–‡†ƒ•™‡ŽŽǤŠ›–ƒ•‡

‹’”‘˜‡†”‡–‡–‹‘ǡƒ†–Š‡†‹‡–‹…”‡ƒ•‡†–Š‡‘ˆǤŽŽ†‹‡–•”‡†—…‡†–Š‡

…‘…‡–”ƒ–‹‘ ‘ˆ ’Š›–ƒ–‡ ‹ –Š‡ …”‘’ ™‹–Š –‹‡Ǥ ‘™‡˜‡”ǡ –Š‡ †‹‡– ”‡•—Ž–‡† ‹ ƒ

Š‹‰Š‡””‡†—…–‹‘–Šƒ–Š‡‘–Š‡”–™‘†‹‡–•ˆ”‘ʹͲ‹—–‡•ƒ†‘™ƒ”†•Ǥ‹—Ž–ƒ‡‘—•Ž›ǡ

–Š‡…‘…‡–”ƒ–‹‘‘ˆ–Š‡Ž‘™‡”‹‘•‹–‘Ž’Š‘•’Šƒ–‡‹•‘‡”•‹…”‡ƒ•‡†Ǥ

ƒ’‡”ǡ͵ʹ’‹‰Ž‡–•™‹–Šƒ‡ƒ™‡‹‰Š–‘ˆʹͳǤͳ‰™‡”‡†‹•–”‹„—–‡†‹‡‹‰Š–’‡•™‹–Š

”‡…‘”†‹‰‘ˆ‹†‹˜‹†—ƒŽˆ‡‡†‹–ƒ‡Ǥ“—ƒŽƒ‘—–•‘ˆ–Š‡‡š’‡”‹‡–ƒŽ†‹‡–•™‡”‡ˆ‡†

–™‹…‡ †ƒ‹Ž›Ǥ ‡”ˆ‘”ƒ…‡ ™ƒ• ”‡…‘”†‡†ǡ ’ ‹ –Š‡ •–‘ƒ…Šǡ Œ‡Œ—— ƒ† ‹Ž‡— ™ƒ•

‡ƒ•—”‡† ƒ† …‘–‡–ˆ”‘ –Š‡ Œ‡Œ—— ƒ†‹Ž‡— ™ƒ• …‘ŽŽ‡…–‡† ˆ‘” ƒ••‡••‡– ‘ˆ

†‹‰‡•–‹„‹Ž‹–›Ǥ ƒ‡…‡•™ƒ•…‘ŽŽ‡…–‡†ˆ‘”–‘–ƒŽ†‹‰‡•–‹„‹Ž‹–›‡ƒ•—”‡‡–•ǡƒ†Ž‡ˆ––Š‹”†ƒ†

ˆ‘—”–Š ‡–ƒ…ƒ”’ƒŽ ™‡”‡ ƒƒŽ›•‡† ˆ‘” „‘‡ ‹‡”ƒŽ‹•ƒ–‹‘Ǥ Š›–ƒ•‡ ƒ††‹–‹‘ ‹…”‡ƒ•‡†

‰”‘™–Šǡ‘ˆǡƒ’’ƒ”‡––‘–ƒŽ–”ƒ…–†‹‰‡•–‹„‹Ž‹–›ȋȌ‘ˆƒ†ƒ‹ƒ††‹–‹‘–‘„‘‡

‹‡”ƒŽ‹•ƒ–‹‘Ǥ…‹†ƒ††‹–‹‘‹’”‘˜‡†‰”‘™–Šǡˆ‡‡†‡ˆˆ‹…ƒ…›ƒ†‘ˆ‰ǡ ‡ƒ†ƒǡ

Š‘™‡˜‡”‘‹–‡”ƒ…–‹‘‡ˆˆ‡…–„‡–™‡‡ƒ…‹†ƒ†’Š›–ƒ•‡™ƒ•ˆ‘—†Ǥ

˜‡–Š‘—‰Šƒ…‹†ƒ††‹–‹‘”‡†—…‡†–Š‡•–‘ƒ…Š’‹’‹‰•‘‹†‹…ƒ–‹‘•‘ˆƒ‹…”‡ƒ•‡†

’Š›–ƒ•‡ ƒ…–‹˜‹–› ™‹–Š ƒ…‹† ƒ††‹–‹‘ ™ƒ• ˆ‘—†Ǥ  …Š‹…‡•ǡ ƒ ‹…”‡ƒ•‡† ’Š›–ƒ–‡

†‡‰”ƒ†ƒ–‹‘ ‹ –Š‡ ƒ–‡”‹‘” †‹‰‡•–‹˜‡ –”ƒ…– ƒ† ‹’”‘˜‡† ‘ˆ ™ƒ• ˆ‘—† ™‹–Š

ƒ…‹†‹ˆ‹…ƒ–‹‘‘ˆ–Š‡†‹‡–…‘–ƒ‹‹‰’Š›–ƒ•‡ǤŠ‡‡ˆˆ‡…–‘ˆƒ…‹†ƒ††‹–‹‘‘’Š›–ƒ•‡ƒ…–‹˜‹–›

‹–Š‡…”‘’‹„”‘‹Ž‡”…Š‹…‡™ƒ•ˆ‘—†ƒŽ”‡ƒ†›ƒˆ–‡”ƒ•Š‘”–”‡–‡–‹‘–›’‹…ƒŽŽ›…‘—Ž†„‡

‘„•‡”˜‡†™‹–Šad libitumˆ‡‡†‹‰ǡƒ†™ƒ•‹…”‡ƒ•‡†™‹–Š‹…”‡ƒ•‡†”‡–‡–‹‘–‹‡‹–Š‡

…”‘’Ǥ ‘™‡˜‡”ǡ ‘ ƒ††‹–‹‘ƒŽ ‡ˆˆ‡…– ‘ˆ ƒ…‹† ƒ††‹–‹‘ ™ƒ• ˆ‘—† ‹ –Š‡ ƒ’’ƒ”‡– ‹Ž‡ƒŽ

†‹‰‡•–‹„‹Ž‹–›ȋȌƒ†–Š‡–‘–ƒŽ–”ƒ…–†‹‰‡•–‹„‹Ž‹–›‘ˆ‹ƒ›‘ˆ–Š‡‡š’‡”‹‡–•Ǥ

ͳͳ

Sammendrag

–Þ”•–‡†‡Ž‡ƒ˜ˆ‘•ˆ‘”ȋȌ‹ˆØ”˜ƒ”‡”ˆ‹‡•‹ˆ‘”ƒ˜ˆ›–ƒ–ǦǤ ›–ƒ–‡”Žƒ‰”‹‰•ˆ‘”‡ˆ‘”

‹ˆ”Þ‘‰‘”‘‰„‡•–¤”ƒ˜‡‹‘•‹–‘Ž”‹‰‡†•‡•ˆ‘•ˆƒ–‰”—’’‡”„—†‡––‹Ž†‡Ǥ ›–ƒ–Ǧ

‡”„ƒ”‡†‡Ž˜‹•–‹Ž‰Œ‡‰‡Ž‹‰ˆ‘”‡ƒ‰‡†‡†›”Ǥˆ‘”†Þ›†•‹ŽŽ‡•—–‹‰ŒÞ†•Žƒ‘‰ƒ

†‡”‡––‡”ˆÞ”‡–‹Žƒ˜”‡‹‰ƒ˜ˆ”ƒŒ‘”†„”—•ƒ”‘‰†‡”‡†ˆ‘”—”‡•‹‰Ǥ ‘”¤

”‡†—•‡”‡‡‰†‡‹‰ŒÞ†•Žƒ„Ž‹”†‡”ˆ‘”‡•‘‰‡ˆ›–ƒ•‡”—–‹‡‡••‹‰–‹Ž•ƒ––ˆØ”‡––‹Ž

‡ƒ‰‡†‡†›”Ǥ‡Ž˜‡†–‹Ž•ƒ––ˆ›–ƒ•‡‡”‹‡‡†„”›–‹‰‡ƒ˜ˆ›–ƒ–ˆ—ŽŽ•–‡†‹‰Ǥˆˆ‡–‡

ƒ˜ˆ›–ƒ•‡‡”ƒ˜Š‡‰‹‰ƒ˜’Ǧ‹˜¤‡–‹–‹ŽŽ‡‰‰–‹Ž‘’’Š‘Ž†•–‹†‡‹†‡†‡Ž‡‡ƒ˜

ˆ‘”†Þ›‡Ž•‡•ƒƒŽ‡‡†‘’–‹ƒŽ’Ǥ‡–˜‹–‹‰•–‡•–‡†‡–ˆ‘”‡•‘‰‡ˆ›–ƒ•‡ƒ–‹˜‹–‡–‡”‹

†‡ˆ”‡”‡†‡Ž‡ƒ˜ˆ‘”†Þ›‡Ž•‡•ƒƒŽ‡’¤‰”—ƒ˜’Ǧ‹˜¤‡–†‡”Ǥ

‘”¤–‡•–‡‡ˆˆ‡–‡ƒ˜•›”‡–‹Ž•‡–‹‰‹ˆØ”‡–’¤‡ˆˆ‡–‹˜‹–‡–‡–‹Ž‡‡•‘‰‡ˆ›–ƒ•‡ǡ‘‰‘

†‡‡‡ˆˆ‡–‡˜‹ŽŽ‡ˆ‘”„‡†”‡•›––‡”Ž‹‰‡”‡˜‡†Þ–‘’’Š‘Ž†•–‹†–‹†‹†‡ˆ”‡”‡†‡Ž‡ƒ˜

ˆ‘”†Þ›‡Ž•‡•ƒƒŽ‡ǡ„Ž‡†‡–—–ˆÞ”––”‡†›”‡ˆ‘”•ÞǤƒŽŽ‡ˆ‘”•Þ‡‡„Ž‡†‡–„”—–‡–

Š˜‡–‡„ƒ•‡”–ˆØ”‡†‡–ŠÞ›–‹Š‘Ž†ƒ˜ˆ›–ƒ–ǦǤ ؔ‡–˜ƒ”‡–‡—–‡–‹Ž•‡–‹‰‡”ȋȌǡ

–‹Ž•ƒ––ƒ—”•›”‡ȋȌǡ–‹Ž•ƒ––ͷͲͲ Ǥ„”ƒƒ‹‹Ǧ†‡”‹˜‡”–ˆ›–ƒ•‡ȋȌ‡ŽŽ‡”–‹Ž•ƒ––„¤†‡

•›”‡‘‰ˆ›–ƒ•‡ȋȌǤ–‹ŽŽ‡‰‰„Ž‡†‡–‰Œ‘”–‡–in vitroˆ‘”•ÞŠ˜‘”‡ˆˆ‡–‹˜‹–‡–‡ƒ˜ˆ‹”‡

‘‡”•‹‡ŽŽ‡ˆ›–ƒ•‡”˜‡†‰‹––‡’Ǧ‹˜¤‡”‘‰‹—„ƒ•Œ‘•–‹†‡”„Ž‡–‡•–‡–ǡ‘‰†‡–„Ž‡‰Œ‘”–

‡ƒ”–Ž‡‰‰‹‰ƒ˜„”—‡ƒ˜”‘‹‘‡”•‹‡ŽŽ‡•Žƒ–‡›ŽŽ‹‰‡”Ǥ

‡•—Ž–ƒ–‡‡ˆ”ƒin vitro ˆ‘”•Þ‡–•‘‡”„‡•”‡˜‡–‹ƒ”–‹‡Ž„Ž‡„”—––‹Ž¤ƒ˜‰ŒÞ”‡Š˜‹Ž‡

ˆ›–ƒ•‡‘‰Š˜‹Ž‡–’Ǧ‹˜¤‡”•‘•—ŽŽ‡„”—‡•‹ˆØ”‡–‹†›”‡ˆ‘”•Þ‡‡Ǥˆ‘”•Þ„Ž‡ͺͲͲ

•Žƒ–‡›ŽŽ‹‰‡” ˆ‘”†‡Ž– ’¤ ͺͲ ˆ‘”•Œ‡ŽŽ‹‰‡ „‹‰‡” ‡–‡ ˆØ”‡–ad libitum ‡ŽŽ‡” ‡†

¤Ž–‹†•ˆØ”‹‰ˆ”ƒͳͷ–‹Ž͵͸†ƒ‰‡”•ƒŽ†‡”Ǥ ”ƒ‡›ŽŽ‹‰‹Š˜‡”„‹‰‡„Ž‡–‹„‹ƒ‘‰‹Š‘Ž†‡–

ˆ”ƒŒ‡Œ——‘‰‹Ž‡—•ƒŽ‡–ǤŠ‘Ž†‡–‹”‘„Ž‡•ƒŽ‡–ˆ”ƒ¤Ž–‹†•ˆØ”‡†‡›ŽŽ‹‰‡”ͺͲǡͳ͸Ͳ

‘‰ ʹͶͲ ‹—––‡” ‡––‡” •–ƒ”–‡ ƒ˜ ˆØ”‹‰‡Ǥ ‡•—Ž–ƒ–‡‡ ˜‹•–‡ ƒ– ˆ›–ƒ•‡ ލ–‡

ˆ‘•ˆ‘”ˆ‘”†Þ›‡Ž‹‰Š‡–‡‘‰‹‡”ƒŽ‹•‡”‹‰ƒ˜„‡‹ǤŽŽ‡”‡†‡‡––‡”ͺͲ‹—––‡”‘’’Š‘Ž†•–‹†‹

”‘ƒ˜ƒ”†‡–‡‡ˆˆ‡–ƒ˜•›”‡–‹Ž•‡–‹‰’¤ˆ›–ƒ•‡Ǧ‡ˆˆ‡–‹˜‹–‡–ǡ‡†‡„‡–›†‡Ž‹‰ˆ‘”„‡†”‡–

‡†„”›–‹‰ƒ˜ˆ›–ƒ–‘‰†ƒ‹‰ƒ˜Žƒ˜‡”‡‹‘•‹–‘Žˆ‘•ˆƒ–Ǧ‹•‘‡”‡”Ǥ‡‰”‡‘’’Š‘Ž†•–‹†‹

”‘ƒ ލ–‡ †‡‡ ‡†„”›–‹‰‡ ›––‡”Ž‹‰‡”‡Ǥ ›–ƒ•‡ ލ–‡ ˆ‘”†Þ›‡Ž‹‰Š‡–‡ ƒ˜ ‘‰

¤Ž–‹†•ˆØ”‹‰Þ–‡ƒ’’ƒ”‡–Œ‡Œ—ƒŽˆ‘”†Þ›‡Ž‹‰Š‡–ȋȌƒ˜‘‰”‡–‡•Œ‘Ǥ

ͳʹ

ƒ”–‹‡Ž„Ž‡†‡–‰Œ‡‘ˆÞ”–‡ƒ”–Ž‡‰‰‹‰ƒ˜‘ˆƒ‰‡–ƒ˜„”—‡ƒ˜”‘ǡ‘‰†‡”‡†

‘’’Š‘Ž†•–‹†ǡ‹ad libitumˆØ”‡†‡‘‡”•‹‡ŽŽ‡•Žƒ–‡›ŽŽ‹‰‡”Ǥƒ”–Ž‡‰‰‹‰‡˜‹•–‡ƒ–ad libitumǦˆØ”‡†‡›ŽŽ‹‰‡”„”—‡””‘ƒ–‹ŽŽƒ‰”‹‰‹•–Þ””‡‰”ƒ†‡–‹†Ž‹‰‡”‡ƒ–ƒ––Ǥ

›ŽŽ‹‰ˆ‘”•Þ‡–ȋˆ‘”•ÞȌ„Ž‡‡ˆˆ‡–‡ƒ˜‘”–‘’’Š‘Ž†•–‹†‘‰•›”‡–‹Ž•‡–‹‰’¤

ˆ›–ƒ•‡‡ˆˆ‡–‹˜‹–‡–‹”‘—†‡”•Þ–Ǥ‡†ʹͲ‘‰ʹͳ†ƒ‰‡”•ƒŽ†‡”„Ž‡ˆƒ•–‡†‡›ŽŽ‹‰‡”‰‹––

–‹Ž‰ƒ‰’¤ˆØ”‹‡–‹‡ǡ‘‰‹Š‘Ž†‡–‹”‘‘‰”¤•„Ž‡•ƒŽ‡–Š˜‡”––Œ—‡†‡‹—–––‹ŽͳͶͲ

‹—––‡”‡––‡”•–ƒ”–‡ƒ˜ˆØ”‹‰‡Ǥ‘•›ŽŽ‹‰‡‡•‘„Ž‡ƒ˜Ž‹˜‡–͸Ͳ‘‰ͳͶͲ‹—––‡”

‡––‡”•–ƒ”–‡ƒ˜ˆØ”‹‰‡„Ž‡‹Š‘Ž†ˆ”ƒŒ‡Œ——‘‰‹Ž‡—‘‰•¤•ƒŽ‡–Ǥ ›–ƒ•‡Þ–‡

”‡–‡•Œ‘ǡ‘‰ǦˆØ”‡–ލ–‡ƒ˜ǤŽŽ‡ˆØ”‡‡”‡†—•‡”–‡‘•‡–”ƒ•Œ‘‡ƒ˜ˆ›–ƒ–‹

”‘‘˜‡”–‹†ǤǦˆØ”‡–”‡•—Ž–‡”–‡‹‹†Ž‡”–‹†‹‡•–Þ””‡”‡†—•Œ‘‡†‡ƒ†”‡–‘

ˆØ”‡‡ˆ”ƒʹͲ‹—––‡”‘‰—–‘˜‡”Ǥƒ–‹†‹‰Þ–‡‘•‡–”ƒ•Œ‘‡ƒ˜Žƒ˜‡”‡‹‘•‹–‘Ž

ˆ‘•ˆƒ–Ǧ‹•‘‡”‡”Ǥ

ƒ”–‹‡Ž„Ž‡͵ʹ•¤‰”‹•‡†‡‰Œ‡‘•‹––•˜‡–’¤ʹͳǡͳ‰ˆ‘”†‡Ž–‹¤––‡„‹‰‡”

‡†‹†‹˜‹†—‡ŽŽ”‡‰‹•–”‡”‹‰ƒ˜ˆØ”‘’’–ƒǤ”‹•‡‡„Ž‡–‹Ž†‡Ž–Ž‹‡•–‘”‡‡‰†‡”ˆØ”–‘

‰ƒ‰‡”†ƒ‰Ž‹‰Ǥ”‘†—•Œ‘•”‡•—Ž–ƒ–‡”„Ž‡„‡”‡‰‡–ǡ’‹ƒ‰‡•‡‡ǡŒ‡Œ——‘‰‹Ž‡—

„Ž‡¤Ž–‘‰‹Š‘Ž†ˆ”ƒŒ‡Œ——‘‰‹Ž‡—„Ž‡•ƒŽ‡–ˆ‘”„‡”‡‰‹‰ƒ˜ˆ‘”†Þ›‡Ž‹‰Š‡–ƒ˜Ǥ

ŒÞ†•‡Ž„Ž‡•ƒŽ‡–ˆ‘”„‡”‡‰‹‰ƒ˜–‘–ƒŽˆ‘”†Þ›‡Ž‹‰Š‡–‘‰–”‡†Œ‡‘‰ˆŒ‡”†‡‡–ƒƒ”’ƒŽ„Ž‡

•ƒŽ‡–ˆ‘”„‡”‡‰‹‰ƒ˜‹‡”ƒŽ‹•‡”‹‰ƒ˜„‡‹Ǥ‹Ž•‡–‹‰ƒ˜ˆ›–ƒ•‡Þ–‡–‹Ž˜‡•–‡ǡ

ƒ˜ǡ‘‰ƒ’’ƒ”‡––‘–ƒŽˆ‘”†Þ›‡Ž‹‰Š‡–ȋȌƒ˜‘‰ƒ‹–‹ŽŽ‡‰‰–‹Ž‹‡”ƒŽ‹•‡”‹‰‡ƒ˜

„‡‹Ǥ‹Ž•‡–‹‰ƒ˜•›”‡Þ–‡–‹Ž˜‡•–‡ǡˆØ”—–›––‡Ž•‡‘‰–‘–ƒŽˆ‘”†Þ›‡Ž‹‰Š‡–ƒ˜‰ǡ ‡‘‰

ƒǡ‡‹‰‡•ƒ•’‹ŽŽ•‡ˆˆ‡–‡ŽŽ‘•›”‡‘‰ˆ›–ƒ•‡„Ž‡ˆ—‡–Ǥ

‡Ž˜‘–‹Ž•‡–‹‰ƒ˜•›”‡”‡†—•‡”–‡’‹ƒ‰‡•‡‡Š‘•‰”‹•ǡ„Ž‡†‡–‹‡ˆ—‡–

‹†‹ƒ•Œ‘‡”’¤Þ–ˆ›–ƒ•‡ƒ–‹˜‹–‡–‡†•›”‡–‹Ž•‡–‹‰Ǥ‘•›ŽŽ‹‰‡”„Ž‡†‡–ˆ—‡–‡

ލ–‡†„”›–‹‰ƒ˜ˆ›–ƒ–‹†‡ˆ”‡”‡†‡Ž‡ƒ˜ˆ‘”†Þ›‡Ž•‡•ƒƒŽ‡‘‰Þ–Œ‡Œ—ƒŽ

ˆ‘”†Þ›‡Ž‹‰Š‡–ƒ˜‡†–‹Ž•‡–‹‰ƒ˜•›”‡–‹ŽˆØ”‡–‡†ˆ›–ƒ•‡Ǥ‡–„Ž‡‘‰•¤ˆ—‡–‡

‡ˆˆ‡–ƒ˜•›”‡–‹Ž•‡–‹‰’¤ˆ›–ƒ•‡ƒ–‹˜‹–‡–ƒŽŽ‡”‡†‡‡––‡”‘”–‘’’Š‘Ž†•–‹†‹”‘ƒŠ‘•

•Žƒ–‡›ŽŽ‹‰ǡ•‘–›’‹•ƒ„Ž‹•‡––‡†ad libitumˆØ”‹‰ǡ‘‰„Ž‡Þ–‡†Ž‡‰”‡

‘’’Š‘Ž†•–‹†‹”‘ƒǤ‹†Ž‡”–‹†„Ž‡†‡–‹‡ˆ—‡–‘‡–‹ŽŽ‡‰‰•‡ˆˆ‡–ƒ˜•›”‡–‹Ž•‡–‹‰’¤

ƒ’’ƒ”‡–‹Ž‡ƒŽˆ‘”†Þ›‡Ž‹‰Š‡–ȋȌ‘‰–‘–ƒŽˆ‘”†Þ›‡Ž‹‰Š‡–ƒ˜‹‘‡ƒ˜ˆ‘”•Þ‡‡Ǥ

ͳ͵

List of publications

Š‹•–Š‡•‹•‹•„ƒ•‡†‘–Š‡–Š”‡‡’ƒ’‡”•Ž‹•–‡†„‡Ž‘™ǣ

ƒ’‡”ǣǤ”‹•–‘ˆˆ‡”•‡ǡǤ–ƒ‹ǡǤ‡œ‡”–‹ŠƒǡǤ‹‡”‘Ñ…œ›ǡǤǤŒ‘•ǡƒ†Ǥ˜‹Š—•Ǥ

ˆˆ‡…–‘ˆ…”‘’”‡–‡–‹‘–‹‡ƒ†ƒ…‹†‹ˆ‹…ƒ–‹‘‘ˆ–Š‡ˆ‡‡†‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›‹„”‘‹Ž‡”

…Š‹…‡•Ǥȋ†‡””‡˜‹•‹‘ˆ‘”’—„Ž‹…ƒ–‹‘‹”‹–‹•Š‘—Ž–”›…‹‡…‡Ȍ

ƒ’‡”ǣǤ”‹•–‘ˆˆ‡”•‡ǡǤ‹ä‹‡™•ƒǡǤƒ…œƒ”‡ǡǤŒ‡ˆ•‡ǡǤǤŒ‘•ǡǤ‘™‹‡•‘

ƒ†Ǥ˜‹Š—•Ǥ••‡••‡–‘ˆ…”‘’—•ƒ‰‡‹ad libitumˆ‡†„‹”†•ƒ†•Š‘”–Ǧ–‡”’Š›–ƒ•‡

‡ˆˆ‹…‹‡…›ƒ•ƒˆˆ‡…–‡†„›ƒ…‹†ƒ††‹–‹‘Ǥȋ—„‹––‡†‘˜‡„‡”ʹͲʹͲȌ

ƒ’‡”ǣǤ”‹•–‘ˆˆ‡”•‡ǡǤŒ‡ˆ•‡ǡǤǤŒ‘•ǡƒ†Ǥ˜‹Š—•ǤŠ‡‡ˆˆ‡…–‘ˆ”‡†—…‡†ˆ‡‡†

’ǡ ’Š›–ƒ•‡ ƒ††‹–‹‘ ƒ† –Š‡‹” ‹–‡”ƒ…–‹‘ –‘ ‹’”‘˜‡ ‹‡”ƒŽ —–‹Ž‹œƒ–‹‘ ‹ ’‹‰•Ǥ

ȋ—„‹––‡†…–‘„‡”ʹͲʹͲȌ

ͳͶ

ͳͷ

1. Introduction

Š‡ƒŒ‘”‹–›‘ˆ‹ˆ‡‡†•–—ˆˆ•ƒ”‡’”‡•‡–ƒ•’Š›–ƒ–‡Ǧȋ‡ŽŽ‡ƒ†ƒ˜‹†”ƒʹͲͲͺȌƒ†

–Š‹•‹•‘Ž›’ƒ”–‹ƒŽŽ›ƒ˜ƒ‹Žƒ„Ž‡ˆ‘”ƒ„•‘”’–‹‘„›‘‘‰ƒ•–”‹…ƒ‹ƒŽ•ǤŠ‡‘”‡–‹…ƒŽŽ›ǡ–Š‡

ƒ‘—–‘ˆ‹ˆ‡‡†‹‰”‡†‹‡–•‹•ƒ†‡“—ƒ–‡–‘…‘˜‡”–Š‡”‡“—‹”‡‡–‘ˆˆ‘”‘‘‰ƒ•–”‹…

ƒ‹ƒŽ•Ǥ‘™‡˜‡”ǡƒ•–Š‡’Š›–ƒ–‡Ǧ‹•’ƒ”–Ž›—ƒ˜ƒ‹Žƒ„Ž‡ˆ‘”†‹‰‡•–‹‘ǡˆ”‘ƒ‹‡”ƒŽ

•‘—”…‡ ‹• ƒ††‡†ǡ ‘ˆ–‡ ™‹–Š •ƒˆ‡–› ƒ”‰‹•ǡ –‘ –Š‡ †‹‡– –‘ …‘˜‡” –Š‡ ”‡“—‹”‡‡–Ǥ

š…‡••‹˜‡ ƒ† —†‹‰‡•–‡† ’Š›–ƒ–‡Ǧ ™‹ŽŽ „‡ ‡š…”‡–‡† ‹ –Š‡ ƒ‹ƒŽ ƒ—”‡Ǥ Š‹• ƒ›

…‘–”‹„—–‡–‘‡˜‹”‘‡–ƒŽ’‘ŽŽ—–‹‘ƒ•†”ƒ‹ƒ‰‡‘ˆˆ”‘ƒ‰”‹…—Ž–—”ƒŽŽƒ†…ƒ…ƒ—•‡

‡—–”‘’Š‹…ƒ–‹‘‘ˆ™ƒ–‡”ȋ‹‰ŠʹͲͲͺȌǤ”‡†—…–‹‘‘ˆ–Š‡ƒ‘—–‘ˆ‹ƒ‹ƒŽƒ—”‡‹•

‹’‘”–ƒ––‘”‡†—…‡–Š‹•’‘ŽŽ—–‹‘ƒ†–Š‹•‹•‡•’‡…‹ƒŽŽ›‹’‘”–ƒ–‹ƒ”‡ƒ•™‹–Š‹–‡•‹˜‡

ƒ‹ƒŽˆƒ”‹‰Ǥ‘”‡†—…‡–Š‡‹ƒ—”‡ˆ”‘‘‘‰ƒ•–”‹…ƒ‹ƒŽ•ƒŠ‹‰Š—–‹Ž‹•ƒ–‹‘‘ˆ

’Š›–ƒ–‡Ǧ‹•‡••‡–‹ƒŽǤ

1.1 Phytate

Figure 1 Structure of a phytic acid at neutral pH based on the Anderson model described by Erdman (1979)

Š›–‹… ƒ…‹† ‹• –Š‡ ˆ”‡‡ ˆ‘” ‘ˆ ›‘Ǧ‹‘•‹–‘Ž Š‡šƒ‹•’Š‘•’Šƒ–‡ ȋ•͸Ȍ ȋ ‹‰—”‡ ͳȌǤ Š‹•

…‘•‹•–•‘ˆƒ‹‘•‹–‘Ž”‹‰™‹–Š•‹š’Š‘•’Šƒ–‡‰”‘—’•„‘—†–‘‹–Ǥ‘™‡˜‡”ǡ’Š›–‹…ƒ…‹†‹•

—•–ƒ„Ž‡ ƒ† ‘……—”• ƒ‹Ž› ‹ –Š‡ ˆ‘” ‘ˆ •ƒŽ– ‹ ’Žƒ–• ƒ† —†‡” ’Š›•‹‘Ž‘‰‹…ƒŽ

ͳ͸

…‘†‹–‹‘•ǡ–Š‹••ƒŽ–‹•…ƒŽŽ‡†’Š›–ƒ–‡ǤŠ›–ƒ–‡‹•–Š‡•–‘”ƒ‰‡ˆ‘”‘ˆƒ†‘–Š‡”‹‡”ƒŽ•

‹•‡‡†•ȋ—‡” et al.ʹͲͳͷȌǤ

Š‡ –‘–ƒŽ ƒ‘—–‘ˆ ƒ†–Š‡ ’‡”…‡–ƒ‰‡ ‘ˆ „‘—†‹’Š›–ƒ–‡ ˜ƒ”‹‡• „‘–Š„‡–™‡‡

†‹ˆˆ‡”‡–’Žƒ–•ǡƒ†„‡–™‡‡†‹ˆˆ‡”‡–’ƒ”–•‘ˆ–Š‡’Žƒ–•ǤŠ‡’‘”–‹‘‘ˆ–‘–ƒŽ‘……—””‹‰

ƒ• ’Š›–ƒ–‡Ǧ ˜ƒ”‹‡• ˆ”‘ Ͷͷ Ψ ȋ•‘›„‡ƒ•Ȍ –‘ ͺͺ Ψ ȋ‘ƒ–ǡ …‘”ǡ ™Š‡ƒ– „”ƒȌ ‹ –Š‡

…‘‘Ž›—•‡†ˆ‡‡†•–—ˆˆ•Ž‹•–‡†‹ƒ„Ž‡ͳǤŠ‡”‡ƒ”‡‹‰‡‡”ƒŽƒŠ‹‰Š‡”…‘–‡–‘ˆ–‘–ƒŽ

ƒ†’Š›–ƒ–‡Ǧ‹…‡”‡ƒŽ„›Ǧ’”‘†—…––Šƒ‹™Š‘Ž‡‰”ƒ‹•Ǥ

Table 1 Phosphorous, phytate and phytase activity in common feedstuffs. The range are the average values from Viveros et al. (2000) and Selle et al. (2003).

Feedstuff Total P (%)

Phytate-P (%)

Proportion (%)¹

Phytase activity (FTU/kg)

Oat ͲǤʹͶǦͲǤʹͻ ͲǤͳ͹ǦͲǤʹͳ ͷͻǦͺͺ ͵ͺǦͺͶ

Wheat ͲǤʹͻǦͲǤ͵ͳ ͲǤʹʹǦͲǤʹ͵ ͹ͳǦ͹ͻ ͷͲ͵Ǧͳ͸͵͹

Barley ͲǤʹ͹ǦͲǤ͵ͳ ͲǤͳͻ ͸ͳǦ͹Ͳ ͵ͶͺǦͳͲͳ͸

Corn ͲǤʹ͵ǦͲǤʹͶ ͲǤͳͺǦͲǤʹͳ ͹ͺǦͺͺ ʹͷǦ͹Ͳ

Wheat bran ͲǤͺͲǦͳǤͳ͸ ͲǤ͹ͲǦͲǤͺͺ ͹͸Ǧͺͺ ʹͳ͹͵ǦͶ͸ʹͶ

Oat bran² ͲǤͺ͵ ͲǤ͸ͺ ͺʹ ʹͷ

Corn gluten meal ͲǤͶʹǦͲǤͷͲ ͲǤʹͻǦͲǤͶʹ ͸ͻǦͺͶ ͶͷǦͳ͹͵

Soybean ͲǤͷ͸ǦͲǤ͹͵ ͲǤ͵ͳǦͲǤ͵͵ ͶͷǦͷͷ ͵ʹǦͶͲ

Soybean meal³ ͲǤ͸͹ ͲǤͶͷ ͸͹ Ͷʹ

Rapeseed meal ͲǤͺͺǦͳǤͲͷ ͲǤ͸͹ǦͲǤ͹͸ ͹ʹǦ͹͸ ͷǦͶͳ

ͳƒŽ…—Žƒ–‡†ƒ•’Š›–ƒ–‡ǦȀ–‘–ƒŽ

ʹ ”‘‹˜‡”‘• et al.ȋʹͲͲͲȌ‘Ž›

͵ ”‘‡ŽŽ‡ et al.ȋʹͲͲ͵Ȍ‘Ž›

Š‡ ’Š›–ƒ–‡ „‘—† ‹• ‘– ƒ˜ƒ‹Žƒ„Ž‡ ˆ‘” ƒ„•‘”’–‹‘ ‹ –Š‡ ‹–‡•–‹‡ ‘ˆ ‘‘‰ƒ•–”‹…

ƒ‹ƒŽ•ǤŠ‡’Š‘•’Šƒ–‡‰”‘—’•‡‡†–‘„‡Ž‹„‡”ƒ–‡†ˆ”‘–Š‡‹‘•‹–‘Ž”‹‰‹‘”†‡”–‘„‡

ƒ˜ƒ‹Žƒ„Ž‡ˆ‘”ƒ„•‘”’–‹‘ǡƒ†–Š‹•’”‘…‡••‹•…ƒ–ƒŽ›•‡†„›–Š‡‡œ›‡’Š›–ƒ•‡Ǥ

Š›–ƒ–‡Šƒ•ƒƒ–‹Ǧ—–”‹–‹˜‡‡ˆˆ‡…–„›ˆ‘”‹‰…Š‡Žƒ–‡•™‹–Š‘–Š‡”‘Ž‡…—Ž‡•Ž‹‡’”‘–‡‹ǡ …ƒ”„‘Š›†”ƒ–‡•ǡƒ†…ƒ–‹‘•Ž‹‡ƒǡ ‡ǡǡ‰ǡƒ†—ȋ ‹‰—”‡ʹȌǤŠ‡•‡…Š‡Žƒ–‡•ƒ‡•

ͳ͹

–Š‡’Š›–ƒ–‡Ǧ„‘—†‘Ž‡…—Ž‡•Ž‡••ƒ……‡••‹„Ž‡ˆ‘”†‹‰‡•–‹‘ƒ†ƒ›ƒ‡–Š‡’Š›–ƒ–‡‘”‡

”‡•‹•–ƒ––‘Š›†”‘Ž›•‹•„›‡œ›‡•ȋ›Žƒ et al.ͳͻͻͷǢƒ‡œ et al.ͳͻͻͻȌǤŠ‡’‘–‡–‹ƒŽˆ‘”

ˆ‘”‹‰…Š‡Žƒ–‡™‹–Š‘–Š‡”‘Ž‡…—Ž‡•‹•†‡’‡†‡–‘ˆ–Š‡’ǡƒŠ‹‰Š‡”’‹…”‡ƒ•‡•–Š‡

„‹†‹‰’‘–‡–‹ƒŽȋƒ‡œ et al.ͳͻͻͻǢ‹‡‹”ƒ et al.ʹͲͳͺȌǤ†‡‰”ƒ†ƒ–‹‘‘ˆ–Š‡’Š›–ƒ–‡

™Š‡”‡„› ’Š‘•’Šƒ–‡ ‰”‘—’• ƒ”‡ ”‡Ž‡ƒ•‡† ˆ”‘ –Š‡ ‹‘•‹–‘Ž ”‹‰ ”‡†—…‡• –Š‡ „‹†‹‰

…ƒ’ƒ…‹–›–‘‘–Š‡”‘Ž‡…—Ž‡•ǡƒ•–Š‡”‡™‹ŽŽ„‡ˆ‡™‡”’‘–‡–‹ƒŽ„‹†‹‰•‹–‡•Ž‡ˆ–ȋ‹‡‹”ƒ et al.

ʹͲͳͺȌǤ

Figure 2 Phytate chelated with several cations at neutral pH (Erdman 1979)

1.2 Phytase

Š›–ƒ•‡•ƒ”‡–Š‡‡œ›‡•–Šƒ–…ƒ–ƒŽ›•‡•–Š‡•–‡’™‹•‡Š›†”‘Ž›•‹•ˆ”‘•͸–‘‹‘”‰ƒ‹…

’Š‘•’Šƒ–‡ƒ†›‘Ǧ‹‘•‹–‘Žƒ†–Š‡›ƒ”‡…ƒ–‡‰‘”‹•‡†ƒ• ’Š‘•’Šƒ–ƒ•‡•ǤŠ‡Š›†”‘Ž›•‹•

‰‡‡”ƒ–‡•ƒ•‡”‹‡•‘ˆŽ‘™‡”•Ǧ‹•‘‡”•˜‹ƒƒ•–‡’™‹•‡”‡ƒ…–‹‘‹ŽŽ—•–”ƒ–‡†‹ ‹‰—”‡͵ǡ

ƒ†–Š‡’‘–‡–‹ƒŽ‡†’”‘†—…–•‘ˆ–Š‹•”‡ƒ…–‹‘‹•‘‡›‘Ǧ‹‘•‹–‘Žƒ†•‹šˆ”‡‡’Š‘•’Šƒ–‡

‰”‘—’•Ǥ

ͳͺ

Figure 3 Stepwise degradation of phytate by phytase

Š‡”‡ƒ”‡ˆ‘—”’‘–‡–‹ƒŽ•‘—”…‡•ˆ‘”’Š›–ƒ•‡•’”‡•‡–‹‘‘‰ƒ•–”‹…ƒ‹ƒŽ•ǣ‹–”‹•‹…

’Š›–ƒ•‡ ˆ”‘ –Š‡ ˆ‡‡†•–—ˆˆ•ǡ —…‘•ƒŽ ’Š›–ƒ•‡ ƒ† ‘–Š‡” ’Š‘•’Šƒ–ƒ•‡• ˆ”‘ –Š‡ •ƒŽŽ

‹–‡•–‹‡ǡ’Š›–ƒ•‡’”‘†—…‡†„›–Š‡‹…”‘ˆŽ‘”ƒ‹–Š‡‹–‡•–‹‡ƒ†‡š‘‰‡‘—•‡œ›‡•

‹…Ž—†‡†‹–Š‡†‹‡–ȋ‡ŽŽ‡ƒ†ƒ˜‹†”ƒʹͲͲ͹ǡʹͲͲͺȌǤ

Š‡ ƒ‘—– ‘ˆ ‹–”‹•‹… ’Žƒ– ’Š›–ƒ•‡ ˜ƒ”‹‡• ™‹†‡Ž› „‡–™‡‡ †‹ˆˆ‡”‡– ˆ‡‡†•–—ˆˆ•

ȋƒ„Ž‡ͳȌǤ‹‰Š–‡’‡”ƒ–—”‡”‡†—…‡•–Š‡•–ƒ„‹Ž‹–›‘ˆ‹–”‹•‹…’Š›–ƒ•‡ȋ•ƒ‡‹Ž‹’‘—” et al.

ʹͲͳʹȌǤ Š—•ǡ ’Š›–ƒ•‡• ˆ”‘ ˆ‡‡†•–—ˆˆ ƒ”‡ ‘•–Ž› ‹ƒ…–‹˜ƒ–‡† †—”‹‰ –Š‡ ’”‘†—…–‹‘ ‘ˆ

…‘‡”…‹ƒŽˆ‡‡†‹š–—”‡•ˆ‘”‘‘‰ƒ•–”‹…ƒ‹ƒŽ•†—‡–‘–Š‡Š‹‰Š–‡’‡”ƒ–—”‡•—•‡†Ǥ

Š‡ …‘‘Ž› —•‡† ’”‘…‡••‡• ‘ˆ •–‡ƒǦ’‡ŽŽ‡–‹‰ ‘” ‡š–”—†‹‰ǡ ƒŽ‘•– ‡Ž‹‹ƒ–‡† –Š‡

‹–”‹•‹…’Š›–ƒ•‡ƒ…–‹˜‹–›‹–Š‡ˆ‡‡†•–—ˆˆ•ȋ‘‰„Ž‘‡†ƒ†‡‡ͳͻͻͲǢ…ŠŽ‡‡” et al.

ʹͲͲͳȌǤ Š‡”‡ˆ‘”‡ǡ –Š‡ ‹–”‹•‹… ’Žƒ– ’Š›–ƒ•‡ Šƒ• ‘ ‘” ˜‡”› Ž‹‹–‡† ‡ˆˆ‡…– ‘ ’Š›–ƒ–‡

†‡‰”ƒ†ƒ–‹‘ ‹ ‘”†‹ƒ”› …‘‡”…‹ƒŽ †‹‡–•ǡ ™Š‹…Š ˆ‘” ‘–Š‡” ‰‘‘† ”‡ƒ•‘• ƒ”‡ ‡‹–Š‡”

’‡ŽŽ‡–‡†‘”‡š–”—†‡†Ǥ

Š‡—…‘•ƒŽ’Š›–ƒ•‡ƒ…–‹˜‹–›Šƒ•„‡‡”‡‰ƒ”†‡†–‘„‡‘ˆŽ‹––Ž‡‹’‘”–ƒ…‡‹‘‘‰ƒ•–”‹…

ƒ‹ƒŽ• ȋ‡ŽŽ‡ ƒ† ƒ˜‹†”ƒ ʹͲͲ͹ȌǤ ‘™‡˜‡”ǡ ’‘—Ž–”› ƒ† ’‹‰• Šƒ˜‡ ˜‡”› ‡ˆˆ‡…–‹˜‡

’Š›–ƒ•‡• ƒ† ‘–Š‡” ’Š‘•’Šƒ–ƒ•‡•ǡ „‘–Š —…‘•ƒŽ ƒ† „ƒ…–‡”‹ƒŽǡ ‹ –Š‡ ‹–‡•–‹‡Ǥ

†‡‰”ƒ†ƒ–‹‘ ‘ˆ •͸ˆ”‘ –Š‡ Œ‡Œ—— –‘ –Š‡ †—‘†‡— ™‹–Š‘—– ‡š‘‰‡‘—• ’Š›–ƒ•‡

ƒ††‹–‹‘ Šƒ• „‡‡ ‘„•‡”˜‡† ‹ …Š‹…‡• ȋ‡ŽŽ‡” et al. ʹͲͳͷƒǡ ʹͲͳͷ„Ȍǡ ‹†‹…ƒ–‡• •‘‡

ͳͻ

—…‘•ƒŽ ’Š›–ƒ•‡ ƒ…–‹˜‹–›Ǥ ƒ‹ et al. ȋʹͲͲͶȌ ˆ‘—† ƒ …‘•‹†‡”ƒ„Ž‡ ‹Ž‡ƒŽ ’Š›–ƒ–‡

†‡‰”ƒ†ƒ–‹‘ȋ͸ͻΨȌ‹„”‘‹Ž‡”…Š‹…‡•™‹–Š‘—–‡š‘‰‡‘—•’Š›–ƒ•‡‘”ƒƒ††‡†‹–Š‡†‹‡–Ǥ

‘™‡˜‡”ǡ™Š‡ƒ™ƒ•ƒ††‡†–‘–Š‡†‹‡––Š‹•†‡‰”ƒ†ƒ–‹‘™ƒ••‹‰‹ˆ‹…ƒ–Ž›”‡†—…‡†–‘ʹͷ ΨǤŠ‹•†‡‘•–”ƒ–‡–Šƒ––Š‡…‘‘Ž›—•‡††‹‡–ƒ”›ƒŽ‡˜‡Ž•‹…‘‡”…‹ƒŽ„”‘‹Ž‡”†‹‡–•

Šƒ˜‡ ƒ •—„•–ƒ–‹ƒŽ ‡‰ƒ–‹˜‡ ‹ˆŽ—‡…‡ –Š‡ ’Š›–ƒ–‡ †‡‰”ƒ†ƒ–‹‘Ǥ  ’‹‰• –Š‡”‡ Šƒ• „‡‡

ˆ‘—†ƒ—…‘•ƒŽ’Š‘•’Šƒ–ƒ•‡ƒ…–‹˜‹–›‹–Š‡•ƒŽŽ‹–‡•–‹‡ȋ— et al.ͳͻͻ͸Ǣ…ŠŽ‡‡” et al. ʹͲͲͳȌ ™Š‹…Š ‹‰Š– ‹ˆŽ—‡…‡ ’Š›–ƒ–‡ †‡‰”ƒ†ƒ–‹‘Ǥ Š‡ ‡ˆˆ‡…– ‘ˆ –Š‡ —…‘•ƒŽ

’Š‘•’Šƒ–ƒ•‡•ƒ†’Š›–ƒ•‡•ƒ”‡Ž‹‹–‡†„‡…ƒ—•‡‘ˆ–Š‡Ž‘™•‘Ž—„‹Ž‹–›‘ˆ’Š›–ƒ–‡ƒ––Š‡’Ǧ

Ž‡˜‡Ž•ˆ‘—†‹–Š‡‹–‡•–‹‡Ǥƒ††‹–‹‘ǡ–Š‡Š‹‰Š†‹‡–ƒ”›…‘…‡–”ƒ–‹‘‘ˆƒ”‡†—…‡•–Š‡

•‘Ž—„‹Ž‹–›‘ˆ–Š‡’Š›–ƒ–‡„›ˆ‘”‹‰‹•‘Ž—„Ž‡’Š›–ƒ–‡…‘’Ž‡š‡•‹–Š‡‹–‡•–‹‡ȋ‡ŽŽ‡et al.ǡ ʹͲͲͻȌǤ ‹…”‘ˆŽ‘”ƒ ‹ –Š‡ Š‹†‰—– ƒŽ•‘ ’”‘†—…‡• ’Š›–ƒ•‡•Ǥ Š‹• ’Š›–ƒ•‡• …‘—Ž†

…‘–”‹„—–‡–‘ƒ‡ƒ”…‘’Ž‡–‡†‡‰”ƒ†ƒ–‹‘‘ˆ•͸‹„‘–Š’‹‰•ƒ†’‘—Ž–”›ȋ‡ŽŽ‡ƒ†

ƒ˜‹†”ƒʹͲͲ͹ǡʹͲͲͺȌǤ‘™‡˜‡”ǡƒ•–Š‡ƒ‹ƒ„•‘”’–‹‘‘ˆ‘……—”•„‡ˆ‘”‡–Š‡Žƒ•––™‘Ǧ

–Š‹”†•‘ˆ–Š‡‹Ž‡—‹…Š‹…‡•ȋ‘†‡Š—–•…‘”†et alǤʹͲͳʹȌƒ†ƒ‡‰Ž‹‰‹„Ž‡’ƒ”–‘ˆ–Š‡

ƒ„•‘”’–‹‘‘……—”•ƒˆ–‡”–Š‡‹Ž‡—‹’‹‰•ȋƒ—‰¡”–‡Žet alǤʹͲͲͺǢ—–Š‡”ˆ—”†et alǤʹͲͳͶȌǡ

–Š‡’Š›–ƒ–‡Ǧ”‡Ž‡ƒ•‡†‹–Š‡Š‹†‰—–™‹ŽŽ‘–„‡ƒ„•‘”„‡†Ǥ

š‘‰‡‘—• ’Š›–ƒ•‡ ‹• ‹•‘Žƒ–‡† ˆ”‘ „ƒ…–‡”‹ƒǡ ›‡ƒ•– ‘” ˆ—‰‹ ƒ† ƒ††‡† –‘ †‹‡–• ˆ‘”

‘‘‰ƒ•–”‹…ƒ‹ƒŽ•–‘‹’”‘˜‡†‡‰”ƒ†ƒ–‹‘‘ˆ’Š›–ƒ–‡ǤŠ›–ƒ•‡•…ƒ„‡…ƒ–‡‰‘”‹•‡†ƒ†

‰”‘—’‡†„ƒ•‡†‘–Š‡’‘’–‹—‘ˆ–Š‡’Š›–ƒ•‡Ǥ…‹†‹…’Š›–ƒ•‡•Šƒ˜‡ƒ’‘’–‹—

ˆ”‘͵ǤͲ–‘ͷǤͷǡƒ†ƒŽƒŽ‹‡’Š›–ƒ•‡•Šƒ˜‡ƒ’‘’–‹—„‡–™‡‡͹ǤͲƒ†ͺǤͲȋ—‡” et al.ʹͲͳͷȌǤ ‘”‘•–‡™Ǧ‰‡‡”ƒ–‹‘‡š‘‰‡‘—•’Š›–ƒ•‡•–Š‡’ˆ‘”‘’–‹—‡ˆˆ‹…ƒ…›‹•

„‡–™‡‡’ͶǤͲƒ†ͷǤͲȋ‡‡œ‡•ǦŽƒ…„—” et al.ʹͲͳͷǢ‹‡‹”ƒ et al.ʹͲͳͺȌǤ‘™‡˜‡”ǡ

‡‡œ‡•ǦŽƒ…„—” et al. ȋʹͲͳͷȌ •Š‘™‡† –Šƒ– –Š‡ ‘’–‹ƒŽ ’ ˆ‘” •‡˜‡ †‹ˆˆ‡”‡–

…‘‡”…‹ƒŽ’Š›–ƒ•‡•˜ƒ”‹‡•…‘•‹†‡”ƒ„Ž›ǤŠ‡”ƒ‰‡™Š‡”‡ͺͲΨ‘ˆ–Š‡ƒš‹—’Š›–ƒ•‡

ƒ…–‹˜‹–›ˆ‘”‡ƒ…Š’Š›–ƒ•‡™ƒ•ˆ‘—†˜ƒ”‹‡†ˆ”‘’͵ǤͲ–‘’ͷǤͷǤ‡…‡ǡƒ”‡Žƒ–‹˜‡Ž›Š‹‰Š

’Š›–ƒ•‡ƒ…–‹˜‹–›…‘—Ž†„‡ˆ‘—†„‘–Š™‹–ŠŽ‘™ƒ†Š‹‰Š‡”’Ǧ˜ƒŽ—‡•†‡’‡†‹‰‘–Š‡

’Š›–ƒ•‡Ǥ

Š‡ˆ‹”•–‰‘˜‡”‡–ƒŽ”‡‰—Žƒ–‹‘ƒ‹‡†ƒ–Ž‹‹–‹‰’‘ŽŽ—–‹‘†—‡–‘‡š…‡••‹˜‡•—’’Ž‹‡•‘ˆ

–‘–Š‡‡˜‹”‘‡–…ƒ‡‹–Š‡‡–Š‡”Žƒ†•‹ͳͻͻͳǤ•ƒ”‡•’‘•‡–‘–Š‹•ǡ–Š‡ˆ‹”•–

…‘‡”…‹ƒŽ ’Š›–ƒ•‡ ™ƒ• ‹–”‘†—…‡† –‘ –Š‡ ƒ”‡– ȋ‡ŽŽ‡ ƒ† ƒ˜‹†”ƒ ʹͲͲ͹ǡ ʹͲͲͺȌǤ

‹…‡–Š‡–Š‡”‡Šƒ˜‡„‡‡ƒŽ‘–‘ˆ”‡•‡ƒ”…Šƒ††‡˜‡Ž‘’‡–‘‡š‘‰‡‘—•’Š›–ƒ•‡•ƒ†

–‘†ƒ›–Š‡”‡ƒ”‡—‡”‘—•†‹ˆˆ‡”‡–’Š›–ƒ•‡•ƒ˜ƒ‹Žƒ„Ž‡…‘‡”…‹ƒŽŽ›Ǥš‘‰‡‘—•’Š›–ƒ•‡

ʹͲ

‹• –‘†ƒ› ”‘—–‹‡Ž› ƒ††‡† –‘ –Š‡ †‹‡–• ‘ˆ ‘‘‰ƒ•–”‹… ƒ‹ƒŽ• –‘ ‹’”‘˜‡ ’Š›–ƒ–‡

†‡‰”ƒ†ƒ–‹‘ƒ†—–‹Ž‹•ƒ–‹‘‘ˆ’Š›–ƒ–‡Ǧȋ‹Ž…‘…ƒ†ƒŽʹͲͳ͸ȌǤ

1.3 Place for exogenous phytase activity

‘–Š’‹‰•ƒ†…Š‹…‡•ƒ”‡‘‘‰ƒ•–”‹…ƒ‹ƒŽ•ǤŠ‡’‹‰Šƒ•ƒDz•‹’Ž‡dz†‹‰‡•–‹‘•›•–‡

™‹–Š ƒ •‹‰Ž‡Ǧ…Šƒ„‡”‡† •–‘ƒ…Š ™Š‡”‡ ‰ƒ•–”‹… Œ—‹…‡• ƒ”‡ •‡…”‡–‡†Ǥ Š‡ •–‘ƒ…Š ‹•

ˆ‘ŽŽ‘™‡† „› –Š‡ •ƒŽŽ ‹–‡•–‹‡ ƒ† –Š‡ Žƒ”‰‡ ‹–‡•–‹‡ ™Š‡”‡ ƒ …‘•‹†‡”ƒ„Ž‡

ˆ‡”‡–ƒ–‹‘ƒ…–‹˜‹–›–ƒ‡•’Žƒ…‡Ǥ

Š‡…”‘’‹’‘—Ž–”›‹•ƒ†‹•–‡•‹„Ž‡•ƒ…Ǧˆ‘”‡†‡š’ƒ•‹‘‘–Š‡‘‡•‘’Šƒ‰—•ǤŠ‡–Š‡

ˆ‡‡†Šƒ•‡–‡”‡†–Š‡…”‘’‹–™‹ŽŽ„‡‰”ƒ†—ƒŽŽ›‘‹•–—”‹•‡†ȋ˜‹Š—• et al.ʹͲͳͲȌǡ‘•–Ž›„›

†”‹‹‰™ƒ–‡”ǤŠ‡…”‘’‹•‘–‹’‘”–ƒ–‹–Š‡…Š‡‹…ƒŽ†‹‰‡•–‹‘‘ˆ–Š‡ˆ‡‡†ǡ„‡…ƒ—•‡

‘ •‡…”‡–‹‘ ‘ˆ †‹‰‡•–‹˜‡ Œ—‹…‡• –ƒ‡• ’Žƒ…‡ –Š‡”‡Ǥ ‘™‡˜‡”ǡ ‡š‘‰‡‘—• ‡œ›‡• ƒ”‡

ƒ…–‹˜ƒ–‡†„›‘‹•–—”‹•‹‰‹–Š‡…”‘’ȋ˜‹Š—•ʹͲͳͶǢŽƒ••‡ et al.ʹͲͳ͸ȌǤˆ–Š‡”‡ƒ”‡ƒ›

‹–”‹•‹…‡œ›‡•‹–Š‡ˆ‡‡†ǡ–Š‡•‡ƒ›ƒŽ•‘•–ƒ”––‘‡š‡”––Š‡‹”‡ˆˆ‡…–‹–Š‡…”‘’ǤŠ‡

•–‘ƒ…Š‹„‹”†•…‘•‹•–‘ˆ–™‘…Šƒ„‡”•ǡ’”‘˜‡–”‹…—Ž—•ƒ†‰‹œœƒ”†ǤŠ‡…”‘’ˆ—…–‹‘•

ƒ•ƒ„—ˆˆ‡”ˆ‘”–Š‡ˆ‡‡†ˆŽ‘™–‘–Š‡‰‹œœƒ”†ǡƒ†–Š‡—•‡‘ˆ–Š‡…”‘’‹•”‡‰—Žƒ–‡†„›–Š‡ˆ‡‡†

…‘–‡–‹–Š‡‰‹œœƒ”†ǤŠ‡’”‘˜‡–”‹…—Ž—•‹•ƒ‰Žƒ†—Žƒ”•–‘ƒ…Šǡ™Š‡”‡‰ƒ•–”‹…Œ—‹…‡•ƒ”‡

•‡…”‡–‡†ǤŠ‡‰‹œœƒ”†ˆ‘ŽŽ‘™•–Š‡’”‘˜‡–”‹…—Ž—•ƒ†‹•ƒ—•…—Žƒ”•–‘ƒ…Š™‹–Š‘—–ƒ›

•‡…”‡–‹‘‘ˆ†‹‰‡•–‹˜‡Œ—‹…‡•Ǥ

Š›–ƒ–‡†‡‰”ƒ†ƒ–‹‘„›‡š‘‰‡‘—•‡œ›‡•’”‹ƒ”‹Ž›–ƒ‡•’Žƒ…‡‹–Š‡ƒ–‡”‹‘”’ƒ”–‘ˆ

–Š‡†‹‰‡•–‹˜‡•›•–‡ǡ–Š‡…”‘’ǡ’”‘˜‡–”‹…—Ž—•ƒ†‰‹œœƒ”†‘ˆ…Š‹…‡ƒ†–Š‡•–‘ƒ…Š‘ˆ

’‹‰•ȋ‡ŽŽ‡ƒ†ƒ˜‹†”ƒʹͲͲ͹ǡʹͲͲͺǢŽƒ••‡ et al.ʹͲͳ͸ȌǤ’–‘͹ͳΨ†‡‰”ƒ†ƒ–‹‘‘ˆ

•͸Šƒ•„‡‡‘„•‡”˜‡†‹–Š‡…”‘’ȋ‡ŽŽ‡” et al.ʹͲͳͷƒǡʹͲͳ͸Ǣ‘‡”ˆ‡Ž† et al.ʹͲͳͺȌǤ

Š‡ƒ‹”‡ƒ•‘–Šƒ––Š‡‡š‘‰‡‘—•’Š›–ƒ•‡ƒ…–‹˜‹–›‹•Š‹‰Š‡•–‹–Š‡ˆ‘”‡•–‘ƒ…Š‹•–Šƒ–

–Š‡’Ž‡˜‡Ž–Š‡”‡‹•‘”‡„‡‡ˆ‹…‹ƒŽˆ‘”’Š›–ƒ•‡ƒ…–‹˜‹–›ƒ†’Š›–ƒ–‡•‘Ž—„‹Ž‹–›–Šƒ‹–Š‡

‹–‡•–‹‡Ǥ‡ƒ”Ž›‘•‡–‘ˆ–Š‡’Š›–ƒ–‡†‡‰”ƒ†ƒ–‹‘‹•„‡‡ˆ‹…‹ƒŽ„‡…ƒ—•‡‘•–‘ˆ–Š‡

ƒ„•‘”’–‹‘–ƒ‡•’Žƒ…‡‹–Š‡ƒ–‡”‹‘”’ƒ”–‘ˆ–Š‡‹–‡•–‹‡Ǥ†‡‰”ƒ†ƒ–‹‘‘ˆ’Š›–ƒ–‡‹–Š‡

ƒ–‡”‹‘” †‹‰‡•–‹˜‡ –”ƒ…– ƒŽ•‘ ”‡†—…‡• –Š‡ ƒ–‹Ǧ—–”‹–‹˜‡ ‡ˆˆ‡…–• ‘ˆ ’Š›–ƒ–‡ ȋ‹‡‹”ƒ et al.

ʹͲͳͺȌǤ

Š‹…‡ˆ‡‡†‘”ƒŽŽ›Šƒ•ƒ’„‡–™‡‡͸ǤͲȋ‘ et al.ʹͲͲͺȌƒ†͸Ǥͷȋƒ…”ƒ‹‡ et al.ʹͲͳ͹Ȍǡ

ƒ† –Š‡ ’ ‹ –Š‡ …”‘’ ‹• ‘”ƒŽŽ› ƒ– –Š‡ •ƒ‡ Ž‡˜‡Ž ™Š‡ –Š‡ ˆ‡‡† ‡–‡”• –Š‡ …”‘’Ǥ

ʹͳ

Š‡”‡ˆ‘”‡ǡ –Š‡ ’ ‹ –Š‡ …”‘’ ‹• —•—ƒŽŽ› •‘‡™Šƒ– Š‹‰Š‡” –Šƒ –Š‡ ‘’–‹ƒŽ ’ ˆ‘”

‡š‘‰‡‘—•’Š›–ƒ•‡ƒ…–‹˜‹–›Ǥ‹‰ˆ‡‡†Šƒ˜‡„‡‡”‡’‘”–‡†–‘Šƒ˜‡ƒ’„‡–™‡‡͸Ǥ͵ƒ†͹Ǥ͵

ȋƒ†…Ž‹ˆˆ‡ et al.ͳͻͻͺǢ‡‡ et al.ͳͻͻͻǢ‘‰„‡‹‰— et al.ʹͲͲ͵ǢŽƒ et al.ʹͲͳʹȌǤ

Š‡–Š‡ˆ‡‡†‡–‡”•–Š‡•–‘ƒ…Š‘ˆ’‹‰•ƒ†–Š‡’”‘˜‡–”‹…—Ž—•ƒ†‰‹œœƒ”†‘ˆ…Š‹…‡•ǡ

‰ƒ•–”‹…Œ—‹…‡•™‹–ŠƒŽ‘™’™‹ŽŽ„‡‹š‡†™‹–Š–Š‡ˆ‡‡†ƒ†”‡†—…‡–Š‡’‘ˆ–Š‡†‹‰‡•–ƒǤ

Š‡’‹–Š‡•–‘ƒ…Š‘ˆ’‹‰•Šƒ•„‡‡”‡’‘”–‡†–‘„‡„‡–™‡‡ͳǤͺƒ†ͶǤͺ™‹–Šƒƒ˜‡”ƒ‰‡

’‘ˆ͵Ǥͷȋ‡”•Œƒ–Ǧ‹ et al.ʹͲͲͳǢ‘‰„‡‹‰— et al.ʹͲͲ͵Ǣ„‡”Šƒ”† et al.ʹͲͲ͹Ǣ‡‡ et al. ʹͲͳͺ„ȌǤ Š‡ ’ ‹ –Š‡ ‰‹œœƒ”† ‹• „‡–™‡‡ ͳǤʹ ƒ† ͵Ǥ͸ ™‹–Š ƒ ƒ˜‡”ƒ‰‡ ‘ˆ ’ ʹǤʹ ȋƒ…”ƒ‹‡ et al.ʹͲͳʹǢ‡‡ et al.ʹͲͳ͹Ǣ‡‡ et al.ʹͲͳͺ„Ǣ‡‡ et al.ʹͲͳͺƒȌǤ˜‡–Š‘—‰Š–Š‡

’‹–Š‡‰‹œœƒ”†‹••‘‡™Šƒ–Ž‘™‡”–Šƒ–Š‡’ˆ‘”‘’–‹ƒŽ’Š›–ƒ•‡ƒ…–‹˜‹–›ǡ‹–‹••–‹ŽŽ

…‘•‹†‡”‡†–‘„‡ƒ‹’‘”–ƒ–•‹–‡ˆ‘”’Š›–ƒ•‡ƒ…–‹˜‹–›Ǥ

Š‡–Š‡†‹‰‡•–ƒ‡–‡”•–Š‡‹–‡•–‹‡ǡ–Š‡’‹•“—‹…Ž›‡Ž‡˜ƒ–‡†–‘ƒŠ‹‰Š‡”’†—‡–‘–Š‡

‹š‹‰™‹–Š’ƒ…”‡ƒ–‹…Œ—‹…‡•Ǥ–Š‡‹–‡•–‹‡ǡ–Š‡’‘”ƒŽŽ›‹•„‡–™‡‡͸Ǥ͵ƒ†͹Ǥͷ‹

…Š‹…‡•ȋ‘ et al.ʹͲͲͺǢ‘œžŽ‡œǦŽ˜ƒ”ƒ†‘ et al.ʹͲͲͺǢ—Ž‹‘™•ƒ et al.ʹͲͲͻȌǡƒ†

„‡–™‡‡ ͷǤͷ ƒ† ’ ͸Ǥͺ ‹ ’‹‰• ȋ‡”•Œƒ–Ǧ‹ et al. ʹͲͲͳǢ ‘‰„‡‹‰— et al. ʹͲͲ͵Ǣ

„‡”Šƒ”† et al.ʹͲͲ͹ȌǤ‡…ƒ—•‡‘ˆ–Š‡ Š‹‰Š’‹–Š‡‹–‡•–‹‡ǡƒ†Š‡…‡–Š‡”‡†—…‡†

•‘Ž—„‹Ž‹–› ‘ˆ –Š‡ ’Š›–ƒ–‡ǡ –Š‹• ’ƒ”– ‘ˆ –Š‡ †‹‰‡•–‹˜‡ –”ƒ…– ‹• ‘– …‘•‹†‡”‡† –‘ „‡ ƒ

‹’‘”–ƒ– •‹–‡ ‘ˆ ‡š‘‰‡‘—• ’Š›–ƒ•‡ ƒ…–‹˜‹–›Ǥ ‘™‡˜‡”ǡ –Š‡”‡ ƒ› „‡ ƒ …‘•‹†‡”ƒ„Ž‡

‡š‘‰‡‘—•’Š›–ƒ•‡ƒ…–‹˜‹–›‹–Š‡‹–‡•–‹‡ƒ•™‡ŽŽǡƒ††‹ˆˆ‡”‡–’Š›–ƒ•‡•Šƒ˜‡†‹ˆˆ‡”‡–

Ž‡˜‡Ž•‘ˆƒ…–‹˜‹–›‹–Š‡‹–‡•–‹‡ȋ›ƒ‰‘ et al.ʹͲͲͷȌǤ

Š‡–‹‡–Š‡ˆ‡‡†•–ƒ›•‹–Š‡ˆƒ˜‘—”ƒ„Ž‡‡˜‹”‘‡–‹–Š‡ƒ–‡”‹‘”†‹‰‡•–‹˜‡–”ƒ…–‹•

‹’‘”–ƒ– ˆ‘” –Š‡ ’Š›–ƒ•‡ ƒ…–‹˜‹–› ȋƒ‹ et al. ʹͲͲͶȌǤ ‡–‡–‹‘ –‹‡ ‹ –Š‡ …”‘’ ‹•

†‡’‡†‡– ‘ ˆ‡‡†‹‰”‡‰‹‡ˆ‘”–Š‡ …Š‹…‡•Ǥ„•‡”˜ƒ–‹‘• ‘ˆad libitum ˆ‡†„”‘‹Ž‡”•

Šƒ˜‡•Š‘™–Šƒ––Š‡›‡ƒ–•ƒŽŽƒ‘—–•‘ˆˆ‡‡†ƒ›–‹‡•–Š”‘—‰Š‘—––Š‡†ƒ›ȋ‹‡Ž•‡

ʹͲͲͶǢ ˜‹Š—• et al. ʹͲͳͲǡ ʹͲͳ͵ȌǤ Š‹• ‡ƒ–‹‰ ’ƒ––‡” ™‹ŽŽ Ž‹‹– –Š‡ —•‡ ‘ˆ –Š‡ …”‘’ ˆ‘”

•–‘”ƒ‰‡ƒ†Š‡…‡‹–‹•ƒ••—‡†–Šƒ–ad libitumˆ‡†„‹”†•†‘‘–—•‡–Š‡…”‘’–‘ƒŽƒ”‰‡

‡š–‡–ȋ˜‹Š—•ʹͲͳͶȌǤŠ‡…”‘’…‘–‡–‘ˆad libitum-ˆ‡†„‹”†•ƒŽ•‘”‡ˆŽ‡…–•–Š‹•Ǥ˜‹Š—• et al.ȋʹͲͳͲȌˆ‘—†–Šƒ–͹ͺΨ‘ˆad libitumǦˆ‡†…Š‹…‡•Šƒ†Ž‡••–Šƒˆ‹˜‡‰‘ˆ†”›ƒ––‡”‹

–Š‡ …”‘’Ǥ –‡”‹––‡– ˆ‡‡†‹‰ ™‹ŽŽ ‹ …‘–”ƒ•– ‹…”‡ƒ•‡ –Š‡ —•‡ ‘ˆ –Š‡ …”‘’ ƒ† –Š‡

”‡–‡–‹‘–‹‡‘ˆ–Š‡ˆ‡‡†‹–Šƒ–’ƒ”–‘ˆ–Š‡†‹‰‡•–‹˜‡–”ƒ…–ȋ˜‹Š—•ʹͲͳͶȌǤ•—…Šƒ•ͷͲ

‰Šƒ•„‡‡ˆ‘—†‹–Š‡…”‘’‘ˆ‹–‡”‹––‡–Ž›ˆ‡†„‹”†•ƒ†–Š‹•…‘–‡–‰”ƒ†—ƒŽŽ›

†‹•ƒ’’‡ƒ”•™‹–Š‹ƒ’‡”‹‘†‘ˆ—’ –‘ͷŠ‘—”•ƒˆ–‡”–‡”‹ƒ–‹‘‘ˆ–Š‡Žƒ•–ˆ‡‡†‹‰„‘—–

ʹʹ

ȋƒ…”ƒ‹‡ et al. ʹͲͳ͹ȌǤ Š—•ǡ •–‹—Žƒ–‹‰ ”‡–‡–‹‘ ‘ˆ ˆ‡‡† ‹ –Š‡ …”‘’ …‘—Ž† ’‘••‹„Ž›

‹…”‡ƒ•‡’Š›–ƒ–‡†‡‰”ƒ†ƒ–‹‘™Š‡‡š‘‰‡‘—•’Š›–ƒ•‡‹•„‡‹‰—•‡†Ǥ˜‹Š—• et al.ȋʹͲͳ͵Ȍ

™‡”‡ ‘– ƒ„Ž‡ –‘ †‡‘•–”ƒ–‡ ƒ ‡ˆˆ‡…– ‘ˆ ‹–‡”‹––‡– ˆ‡‡†‹‰ ‘ ’Š›–ƒ•‡ ‡ˆˆ‹…ƒ…›Ǥ

‘™‡˜‡”ǡƒ…”ƒ‹‡ et al.ȋʹͲͳ͹Ȍˆ‘—†ƒŠ‹‰Š‡”†‹‰‡•–‹„‹Ž‹–›ƒ†•͸†‹‰‡•–‹„‹Ž‹–›™‹–Š

‹–‡”‹––‡–ˆ‡‡†‹‰–Šƒ™‹–Šad libitumƒ……‡••–‘ˆ‡‡†Ǥ˜‹Š—• et al.ȋʹͲͳͲȌ†‹†ˆ‹†ƒͷͲ Ψ ”‡†—…–‹‘ ‹ …‘…‡–”ƒ–‹‘ ‘ˆ •͸ ‹ –Š‡ …”‘’ ™‹–Š ͳͲͲ ‹—–‡• ”‡–‡–‹‘ –‹‡Ǥ

‡–‡–‹‘ –‹‡‹…”‘’‹• ƒŽ•‘ƒ••‘…‹ƒ–‡†™‹–Šƒ …‘•‹†‡”ƒ„Ž‡ˆ‡”‡–ƒ–‹‘ ƒ…–‹˜‹–›„›

lactobacilliƒ†…‘•‡“—‡–Ž›ƒ’”‘†—…–‹‘‘ˆŽƒ…–‹…ƒ…‹†–Šƒ–™‹ŽŽ”‡†—…‡–Š‡’‹…”‘’

‘˜‡”–‹‡ȋ—–Ž‡” et al.ʹͲͲͷǢ‘œ‡ˆ‹ƒ et al.ʹͲͲ͸ȌǤ

Š‡”‡–‡–‹‘–‹‡‘ˆ–Š‡ˆ‡‡†‹–Š‡•–‘ƒ…Š‘ˆ’‹‰•ƒ†–Š‡ˆŽ‘™‘ˆ†‹‰‡•–ƒˆ”‘–Š‡

•–‘ƒ…Š‹•‹ˆŽ—‡…‡†„›–Š‡‡ƒŽ•‹œ‡ƒ†–Š‡’Š›•‹…ƒŽ•–ƒ–‡‘ˆ–Š‡†‹‡–ȋ”‡‰‘”› et al.

ͳͻͻͲȌǤ……‘”†‹‰–‘Žƒƒ„Œ‡”‰ et al.ȋʹͲͳͳȌǡ‘”‡–ŠƒŠƒŽˆ‘ˆ–Š‡‡ƒ–‡„›’‹‰•Ž‡ƒ˜‡

–Š‡•–‘ƒ…Š„‡ˆ‘”‡–Š‡‰ƒ•–”‹…’„‡…‘‡•ƒ•Ž‘™ƒ•’͵Ǥͷǡ™Š‹…Š‹•–Š‡’‘’–‹—ˆ‘”

…‘‡”…‹ƒŽ’Š›–ƒ•‡•™‹–Š–Š‡Ž‘™‡•–’‘’–‹—Ǥ‹–Šƒ‹…”‡ƒ•‡†”‡–‡–‹‘–‹‡‹

–Š‡•–‘ƒ…Šǡ–Š‡†‡‰”ƒ†ƒ–‹‘‘ˆ’Š›–ƒ–‡™‹ŽŽ„‡‹’”‘˜‡†ȋŽƒƒ„Œ‡”‰et alǤǡʹͲͳͳȌǤ

1.4 Limitations in phytate degradation

Š‡ †‡‰”ƒ†ƒ–‹‘ ‘ˆ ’Š›–ƒ–‡ „› ‡š‘‰‡‘—• ’Š›–ƒ•‡ ‹• ‘– …‘’Ž‡–‡Ǥ  ƒ ”‡˜‹‡™ „›

Ž‘‹•‹ȋʹͲͳͳȌ‹–™ƒ•‡•–ƒ„Ž‹•Š‡†–Šƒ––Š‡”‡Ž‡ƒ•‡‘ˆˆ”‘’Š›–ƒ–‡‹„”‘‹Ž‡”…Š‹…‡

™ƒ•‹…”‡ƒ•‡†ˆ”‘ƒƒ˜‡”ƒ‰‡‘ˆͳͻΨ™‹–Š‘—–’Š›–ƒ•‡ƒ††‹–‹‘–‘͵ͺΨ™‹–Š’Š›–ƒ•‡

ƒ††‹–‹‘Ǥ’‹‰•ǡ‡š‘‰‡‘—•’Š›–ƒ•‡ƒ††‹–‹‘‹…”‡ƒ•‡†–Š‡’Š›–ƒ–‡Ǧ†‹‰‡•–‹„‹Ž‹–›ˆ”‘

͵ͻΨ™‹–Š‘—–’Š›–ƒ•‡ƒ††‹–‹‘–‘͸ͷΨ™‹–Š’Š›–ƒ•‡ȋ‘•‡ˆ‡Ž†‡”Ǧ—‘ et al.ʹͲʹͲȌǤŠ‡

‹…‘’Ž‡–‡†‡‰”ƒ†ƒ–‹‘ˆ‘—†‹–Š‹•”‡˜‹‡™‹•ƒ‹†‹…ƒ–‹‘–Šƒ––Š‡”‡‹•ƒ’‘–‡–‹ƒŽˆ‘”

‹’”‘˜‹‰–Š‡—–‹Ž‹•ƒ–‹‘‘ˆ’Š›–ƒ–‡Ǧ™Š‡’Š›–ƒ•‡‹•ƒ††‡†–‘–Š‡†‹‡–Ǥˆ–Š‡’Š›–ƒ•‡

‡ˆˆ‹…ƒ…›‹•‹’”‘˜‡†ǡ–Š‡—–‹Ž‹•ƒ–‹‘‘ˆ’Š›–ƒ–‡Ǧ…‘—Ž†„‡‹…”‡ƒ•‡†ˆ—”–Š‡”ǤŠ‹•™‹ŽŽ„‡

‘ˆ•‹‰‹ˆ‹…ƒ…‡–‘–Š‡‹†—•–”›•‹…‡‹–ƒ›”‡†—…‡–Š‡ƒ‘—–‘ˆ‹–Š‡ƒ—”‡ˆ”‘

ˆƒ”ƒ‹ƒŽ•ƒ†…‘•‡“—‡–Ž›”‡†—…‡–Š‡‡˜‹”‘‡–ƒŽ’‘ŽŽ—–‹‘Ǥ

1.5 Acid and phytase

–Šƒ•„‡‡•Š‘™–Šƒ––Š‡ƒ††‹–‹‘‘ˆ‘”‰ƒ‹…ƒ…‹†–‘–Š‡ˆ‡‡†ƒ›”‡†—…‡–Š‡’‹–Š‡

…”‘’‘ˆ…Š‹…‡ƒ†‹–Š‡•–‘ƒ…Š‘ˆ’‹‰•ȋ‹ et al.ʹͲͳͷǢ—‹”›ƒ”ƒ›ƒƒ†ƒƒƒ

ʹͲͳͷȌǤŠ‡’Š›–ƒ•‡ƒ…–‹˜‹–›‹–Š‡…”‘’ƒ›„‡‹…”‡ƒ•‡†‹ˆ–Š‡’‹•Ž‘™‡”‡†–‘ƒ’…Ž‘•‡”

ʹ͵

–‘–Š‡‘’–‹—‘ˆ–Š‡’Š›–ƒ•‡Ǥ’‹‰•ǡ‹–Šƒ•„‡‡•Š‘™–Šƒ–ƒ”‡†—…‡†’‹–Š‡•–‘ƒ…Š

™‹ŽŽ”‡†—…‡–Š‡‡’–›‹‰”ƒ–‡ȋƒ†‡”ƒ”et alǤʹͲͳ͹ȌǤƒ††‹–‹‘ǡƒ”‡†—…–‹‘‹–Š‡

’‹–Š‡•–‘ƒ…Š„›ƒ…‹†ƒ††‹–‹‘‹–Š‡ˆ‡‡†ƒ›Ž‡ƒ†–‘ƒ‹…”‡ƒ•‡††‡‰”ƒ†ƒ–‹‘‘ˆ

’Š›–ƒ–‡ „‡…ƒ—•‡ –Š‡ ’ ‹• ‹‡†‹ƒ–‡Ž› Ž‘™‡”‡† –‘ ƒ ‘”‡ „‡‡ˆ‹…‹ƒŽ ’ ˆ‘” ’Š›–ƒ•‡

ƒ…–‹˜‹–›ƒ†’Š›–ƒ–‡•‘Ž—„‹Ž‹–›Ǥ

’‹‰•ǡ–Š‡”‡ƒ”‡…‘ˆŽ‹…–‹‰”‡•—Ž–•”‡‰ƒ”†‹‰–Š‡‡ˆˆ‡…–‘ˆƒ…‹†ƒ††‹–‹‘‘’Š›–ƒ•‡

‡ˆˆ‹…ƒ…›Ǥ‡˜‡”ƒŽ‡š’‡”‹‡–•Šƒ˜‡•Š‘™–Šƒ––Š‡ƒ††‹–‹‘‘ˆ‘”‰ƒ‹…ƒ…‹†•Šƒ˜‡ƒ

’‘•‹–‹˜‡‡ˆˆ‡…–‘†‹‰‡•–‹„‹Ž‹–›‹’‹‰•ȋ‡‡ et al.ͳͻͻͻǢ‘‰„Ž‘‡† et al.ʹͲͲͲȌǤ

‘™‡˜‡”ǡ‘–Š‡”‡š’‡”‹‡–•Šƒ˜‡ˆƒ‹Ž‡†–‘•Š‘™ƒ‹–‡”ƒ…–‹‘„‡–™‡‡‘”‰ƒ‹…ƒ…‹†

ƒ†’Š›–ƒ•‡‡ˆˆ‹…ƒ…›ȋƒ†…Ž‹ˆˆ‡ et al.ͳͻͻͺǢ‘‰„‡‹‰— et al.ʹͲͲ͵ȌǤ…Š‹…‡•–Š‡”‡

‹•ƒŽ•‘…‘ˆŽ‹…–‹‰”‡•—Ž–•‘–Š‹•–‘’‹…ǡƒ‡–ƒǦƒƒŽ›•‹•„›‹‡‹”ƒ et al.ȋʹͲͳ͹Ȍ•Š‘™‡†

–Šƒ––Š‡…‘„‹ƒ–‹‘‘ˆ…‹–”‹…ƒ…‹†ƒ†’Š›–ƒ•‡•‹’”‘˜‡†’‡”ˆ‘”ƒ…‡ƒ†„‘‡

‹‡”ƒŽ‹•ƒ–‹‘‹„”‘‹Ž‡”…Š‹…‡•Ǥƒ‹ et al.ȋʹͲͳ͵Ȍƒ†‘›‡‰‘ et al.ȋʹͲͳͲȌˆ‘—†

ƒ‹…”‡ƒ•‡†‹Ž‡ƒŽ†‹‰‡•–‹„‹Ž‹–›„›…‘„‹‹‰‘”‰ƒ‹…ƒ…‹†™‹–Š’Š›–ƒ•‡ǡŠ‘™‡˜‡”‹–Š‡

”‡˜‹‡™„›‹‡‹”ƒet al. (ʹͲͳͺȌˆ‡™‘ˆ–Š‡…‹–‡†‡š’‡”‹‡–•ˆ‘—†ƒƒ††‹–‹‘ƒŽ‡ˆˆ‡…–‘ˆ

ƒ…‹†‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›Ǥ

•–Š‡”‡‹•ƒ’‘–‡–‹ƒŽˆ‘”„‡––‡”—–‹Ž‹•ƒ–‹‘‘ˆ’Š›–ƒ–‡Ǧ‹‘‘‰ƒ•–”‹…ƒ‹ƒŽ•ǡ‘”‡

‘™Ž‡†‰‡ƒ„‘—––Š‡‡ˆˆ‡…–‘ˆƒ…‹†‹ˆ‹…ƒ–‹‘‘ˆ–Š‡†‹‡–‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›‹–Š‡ƒ–‡”‹‘”

†‹‰‡•–‹˜‡–”ƒ…–‹•”‡“—‹”‡†Ǥ‘‰‡–Š‡”™‹–Š–Š‡“—‡•–‹‘‹ˆ–Š‹•‡ˆˆ‡…–™‘—Ž†„‡‹’”‘˜‡†„›

ƒ‹…”‡ƒ•‡†”‡–‡–‹‘–‹‡‹–Š‡ƒ–‡”‹‘”’ƒ”–‘ˆ–Š‡†‹‰‡•–‹˜‡–”ƒ…–ǡƒ†Š‘™–Š‹•‡ˆˆ‡…–•

–Š‡–‘–ƒŽ–”ƒ…–†‹‰‡•–‹„Ž›–Š‹•™ƒ•–Š‡„ƒ•‹•ˆ‘”–Š‡‘„Œ‡…–‹˜‡•‹–Š‹••–—†›Ǥ

ʹͶ

2. Aims of the thesis

Š‡ƒ‹ƒ‹‘ˆ–Š‹•–Š‡•‹•™ƒ•–‘…‘–”‹„—–‡–‘ƒ‹’”‘˜‡†’Š›–ƒ–‡Ǧ†‡‰”ƒ†ƒ–‹‘ƒ†

—–‹Ž‹•ƒ–‹‘„›‹’”‘˜‹‰–Š‡‡ˆˆ‹…ƒ…›‘ˆ’Š›–ƒ•‡ǤŠ‡ƒ‹™ƒ•†‡”‹˜‡†ˆ”‘ƒ–ƒ”‰‡–‘ˆ

”‡†—…‹‰–Š‡ƒ‘—–‘ˆ‹ƒ—”‡ˆ”‘‘‘‰ƒ•–”‹…ƒ‹ƒŽ•Ǥ

‘”†‡”–‘ƒ…Š‹‡˜‡–Š‹•ƒ‹ǡ–Š‡ˆ‘ŽŽ‘™‹‰Š›’‘–Š‡•‡•™‡”‡–‡•–‡†ǣ

x …‹†‹ˆ‹…ƒ–‹‘ ‘ˆ –Š‡ †‹‡– ™‹ŽŽ ‹…”‡ƒ•‡ ‡š‘‰‡‘—• ’Š›–ƒ•‡ ‡ˆˆ‹…ƒ…› –Š”‘—‰Š

‹…”‡ƒ•‡†’Š›–ƒ–‡†‡‰”ƒ†ƒ–‹‘‹–Š‡ƒ–‡”‹‘”†‹‰‡•–‹˜‡–”ƒ…–Ǥ

x  ‹…”‡ƒ•‡† ’Š›–ƒ–‡ †‡‰”ƒ†ƒ–‹‘ ‹ –Š‡ ƒ–‡”‹‘” †‹‰‡•–‹˜‡ –”ƒ…– ™‹ŽŽ ‹’”‘˜‡

‘˜‡”ƒŽŽ†‹‰‡•–‹„‹Ž‹–›Ǥ

ƒ††‹–‹‘ǡ–Š‹••’‡…‹ˆ‹…Š›’‘–Š‡•‹•ˆ‘”…Š‹…‡•™ƒ•–‡•–‡†ǣ

x Š‡‡ˆˆ‡…–‘ˆƒ…‹†ƒ††‹–‹‘–‘–Š‡†‹‡–‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›™‹ŽŽ„‡ˆ—”–Š‡”‹’”‘˜‡†

„›ƒ‹…”‡ƒ•‡†…”‘’”‡–‡–‹‘–‹‡†—‡–‘‹–‡”‹––‡–ˆ‡‡†‹‰Ǥ

ʹͷ

3. Methodology

‘”‹‰™‹–Š–Š‹•–Š‡•‹•ǡ–Š‡Š›’‘–Š‡•‡•™ƒ•ƒ’’”‘ƒ…Š‡†„›ƒin vitro testǡƒˆ‹‡Ž†ƒ••ƒ›

ƒ†–Š”‡‡ƒ‹ƒŽ‡š’‡”‹‡–•ǤŠ‡in vitro•–—†›™ƒ•…ƒ””‹‡†‘—––‘‹˜‡•–‹‰ƒ–‡–Š‡‡ˆˆ‡…–

‘ˆƒ…‹†ƒ†”‡–‡–‹‘–‹‡‘†‹ˆˆ‡”‡–…‘‡”…‹ƒŽ’Š›–ƒ•‡•Ǥˆ‹‡Ž†ƒ••‡••‡––‘ƒ’

–Š‡…”‘’…‘–‡–‹„”‘‹Ž‡”…Š‹…‡•ˆ”‘…‘‡”…‹ƒŽˆƒ”•™ƒ•…‘†—…–‡†Ǥ™‘„”‘‹Ž‡”

…Š‹…‡‡š’‡”‹‡–•ǡ‘‡™‹–Š…Š‹…‡•‹’‡•ǡƒ†‘‡™‹–Š…Š‹…‡‹‹†‹˜‹†—ƒŽ…ƒ‰‡•ǡ

™‡”‡…ƒ””‹‡†‘—––‘ƒ••‡••‹ˆ‘”ƒ–‹‘ƒ„‘—––Š‡‡ˆˆ‡…–‘ˆƒ…‹†ƒ††‹–‹‘ƒ†”‡–‡–‹‘–‹‡

‹–Š‡…”‘’‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›Ǥ‡’‹‰‡š’‡”‹‡–™ƒ•…ƒ””‹‡†‘—––‘ƒ••‡••–Š‡‡ˆˆ‡…–‘ˆ

ƒ…‹†ƒ††‹–‹‘‘’Š›–ƒ•‡‡ˆˆ‹…ƒ…›‹’‹‰•Ǥ

3.1 In vitro test

•‹’Ž‡in vitro ’”‘…‡†—”‡ •‹—Žƒ–‹‰ –Š‡ …”‘’ ™ƒ• —•‡† –‘ –‡•– –Š‡ ‡ˆˆ‹…ƒ…› ‘ˆ ˆ‘—”

…‘‡”…‹ƒŽ’Š›–ƒ•‡’”‘†—…–•ƒ–•‡Ž‡…–‡†’Ž‡˜‡Ž•ƒ†‹…—„ƒ–‹‘–‹‡•Ǥ‡‰ȋάͲǤͲͳ‰Ȍ

‘ˆ–Š‡†‹‡–™ƒ•‹š‡†™‹–ŠͷǤͲŽ†‡‹‘‹•‡†™ƒ–‡”ǡͲ‘”ͷͲͲ ’Š›–ƒ•‡ƒ†ͲǤͲǡͶǤͲǡͳ͵ǤͲ

‘”ͷͲǤͲρŽˆ‘”‹…ƒ…‹†–‘”‡ƒ…Š–Š‡’ǦŽ‡˜‡Ž•‘ˆ͸Ǥ͹ǡͷǤͷǡͶǤͷƒ†͵Ǥͷ‹ƒ–‡•––—„‡ǤŠ‡–‡•–

–—„‡•™‡”‡˜‘”–‡š‡†™‡ŽŽ„‡ˆ‘”‡‹…—„ƒ–‹‘‹ƒͶͲ陃–‡”„ƒ–Šǡ™Š‡”‡–Š‡›™‡”‡•Šƒ‡

„›Šƒ†‡˜‡”›ͷ‹—–‡•Ǥˆ–‡”ƒ‹…—„ƒ–‹‘‘ˆͳͲǡʹͲǡ͵ͲǡͶͷ‘”͸Ͳ‹ǡ–Š‡‡œ›ƒ–‹…

”‡ƒ…–‹‘™ƒ•–‡”‹ƒ–‡†„›ƒ††‹‰ͷǤͲŽͶΨ–”‹…ŠŽ‘”‘ƒ…‡–‹…ƒ…‹†ȋȌǤŠ‡”‡ƒˆ–‡”–Š‡

–‡•––—„‡•™‡”‡…‡–”‹ˆ—‰‡†ǡˆ‘ŽŽ‘™‡†„›…‘ŽŽ‡…–‹‘‘ˆ–Š‡•—’‡”ƒ–ƒ–ƒ†ƒƒŽ›•‹•ˆ‘”ˆ”‡‡

’Š‘•’Šƒ–‡ƒ•†‡•…”‹„‡†‹ƒ’‡”ǤŠ‡’Š›–ƒ•‡™‹–Š–Š‡Žƒ”‰‡•–”‡•’‘•‡–‘ƒ”‡†—…‡†’

™ƒ•–Š‡C. braakiiǦ†‡”‹˜‡†’Š›–ƒ•‡ǡƒ†–Š‹•™ƒ•…Š‘•‡–‘‰‡–Š‡”™‹–Šƒ’Ž‡˜‡Ž‘ˆͶǤͷ–‘

„‡—•‡†‹–Š‡†‹‡–•ˆ‘”–Š‡in vivo‡š’‡”‹‡–•Ǥ

3.2 Experiment I

Effect of acid addition on phytase efficacy in intermittently and ad libitum fed chickens

‹‰Š–Š—†”‡†‘‡Ǧ†ƒ›Ǧ‘Ž†ƒŽ‡‘••͵Ͳͺ…Š‹…‡•™‡”‡”ƒ†‘Ž›ƒ••‹‰‡†–‘ͺͲ’‡•

™‹–Š ͳͲ „‹”†• ‹ ‡ƒ…ŠǤ Š‡ ’‡• ™‡”‡ ƒ””ƒ‰‡† ‹ –Š‡ ‹††Ž‡ ‘ˆ ƒ ‡˜‹”‘‡–ƒŽŽ›

…‘–”‘ŽŽ‡† „”‘‹Ž‡” Š‘—•‡ ™‹–Š ͻǡͲͲͲ Ž‘‘•‡ǦŠ‘—•‡† „‹”†• Šƒ–…Š‡† ƒ– –Š‡ •ƒ‡ –‹‡

•—””‘—†‹‰–Š‡‡š’‡”‹‡–ƒŽ’‡•ǤŠ‡…Š‹…‡•™‡”‡ad libitum‘”‹–‡”‹––‡–Ž›ˆ‡†

‡‹–Š‡”ƒŠ‹‰Š’Š›–ƒ–‡Ǧ†‹‡–ȋȌǡ‘”–Š‡•ƒ‡†‹‡–™‹–Šˆ‘”‹…ƒ…‹†ƒ††‡†ȋȌǡͷͲͲ C. braakiiǦ†‡”‹˜‡†’Š›–ƒ•‡ƒ††‡†ȋȌ‘”„‘–Šƒ††‡†ȋȌ‹ƒʹšʹšʹˆƒ…–‘”‹ƒŽ

ʹ͸

†‡•‹‰ˆ”‘†ƒ›ͳͷ–‘͵͸‘ˆƒ‰‡ǤŠ‡ƒd libitumˆ‡†„‹”†•Šƒ†–™‘ͶǦŠ‘—”†ƒ”’‡”‹‘†•

™‹–ŠʹŠ‘—”Ž‹‰Š–‹„‡–™‡‡ǤŠ‡‹–‡”‹––‡–Ž›ˆ‡†„‹”†•Šƒ†ƒ……‡••–‘ˆ‡‡†„‡–™‡‡–Š‡

Š‘—”•‘ˆͲͺǣͲͲ–‘ͲͻǣͲͲǡͳʹǣͲͲ–‘ͳ͵ǣͲͲǡͳ͸ǣ͵Ͳ–‘ͳ͹ǣ͵ͲǡʹͳǣͲͲ–‘ʹʹǣͲͲƒ†ˆ”‘ͲʹǣͲͲ–‘

ͲͶǣͲͲǤ

†ƒ›͵͸ǡ‘‡„‹”†ˆ”‘‡ƒ…Šad libitum ˆ‡†’‡™ƒ•‹ŽŽ‡†ˆ‘”…‘ŽŽ‡…–‹‘‘ˆ…‘–‡–ˆ”‘

–Š‡…”‘’ǡ–Š‡Œ‡Œ——ƒ†–Š‡‹Ž‡—‹ƒ††‹–‹‘–‘–Š‡Ž‡ˆ––‹„‹ƒǤ†ƒ›͵͹ǡ‘‡„‹”†’‡”

’‡‘ˆ‹–‡”‹––‡–Ž›ˆ‡†„‹”†•™ƒ•‹ŽŽ‡†ˆ‘”…‘ŽŽ‡…–‹‘‘ˆ–Š‡…”‘’…‘–‡–ͺͲǡͳ͸Ͳƒ†

ʹͶͲ‹—–‡•ƒˆ–‡”–Š‡•–ƒ”–‘ˆ–Š‡ˆ‡‡†‹‰Ǥƒ††‹–‹‘ǡ…‘–‡–ˆ”‘–Š‡Œ‡Œ——ƒ†–Š‡

‹Ž‡—ƒ†–Š‡ Ž‡ˆ––‹„‹ƒ™‡”‡…‘ŽŽ‡…–‡†ʹͶͲ‹—–‡•ƒˆ–‡”–Š‡•–ƒ”–‘ˆ–Š‡ ˆ‡‡†‹‰Ǥ‹‡–

’”‘†—…–‹‘ǡ•ƒ’Ž‡’”‡’ƒ”ƒ–‹‘ǡƒƒŽ›–‹…ƒŽ’”‘…‡†—”‡•ƒ†‘”‡†‡–ƒ‹Ž•‘–Š‡•ƒ’Ž‹‰

’”‘…‡†—”‡ƒ”‡†‡•…”‹„‡†‹ƒ’‡”Ǥ

3.3 Field assessment

ˆ‹‡Ž†ƒ••‡••‡–™ƒ•…ƒ””‹‡†‘—––‘ƒ’–Š‡‡š–‡–‘ˆ…”‘’—•ƒ‰‡ƒ†–Š—•”‡–‡–‹‘

–‹‡‹…‘‡”…‹ƒŽ„”‘‹Ž‡”…Š‹…‡•Ǥ–ͳͲǡʹͲƒ†͵Ͳ†ƒ›•‘ˆƒ‰‡ǡͶͲȋάͳȌ…Š‹…‡•‹‡ƒ…Š

‘ˆ ˆ‘—” ˆƒ”• ™‡”‡ ‹ŽŽ‡†Ǥ ‡‡†‹‰ ƒ† ƒƒ‰‡‡– ’”‘…‡†—”‡• ‘ –Š‡ ˆƒ”• ƒ”‡

†‡•…”‹„‡†‹†‡–ƒ‹Ž‹’ƒ’‡”ǤŠ‡…”‘’…‘–‡–™ƒ•…‘ŽŽ‡…–‡†“—ƒ–‹–ƒ–‹˜‡Ž›‹‡†‹ƒ–‡Ž›

ƒˆ–‡”‹ŽŽ‹‰ƒ†–Š‡”‡ƒˆ–‡”ˆ”‘œ‡ƒ– ǦʹͲιǤˆ–‡”–Šƒ™‹‰ǡ’™ƒ•‡ƒ•—”‡†ǡƒ†

…‘–‡–™ƒ•†‡–‡”‹‡†„›†”›‹‰ƒ–ͳͲͶι‘˜‡”‹‰Š–Ǥ

3.4 Experiment II

Assessment of short-term phytase efficiency as affected by acid addition

Š‡•‡…‘†…Š‹…‡‡š’‡”‹‡–™ƒ•†‡•‹‰‡†–‘‹˜‡•–‹‰ƒ–‡–Š‡†‡‰”ƒ†ƒ–‹‘‘ˆ’Š›–ƒ–‡‹

–Š‡…”‘’•Š‘”–‡”–‹‡ƒˆ–‡”–Š‡•–ƒ”–‘ˆ–Š‡ˆ‡‡†‹‰ƒ†‘”‡ˆ”‡“—‡–Ž›–Šƒ‹‡š’‡”‹‡–

Ǥ–ͳͳ†ƒ›•‘ˆƒ‰‡ǡͳʹͲƒŽ‡‘••͵Ͳͺ…Š‹…‡•™‡”‡’Žƒ…‡†‹‹†‹˜‹†—ƒŽ…ƒ‰‡•ǤŠ‡„‹”†•

™‡”‡‹–‡”‹––‡–Ž›ˆ‡†ƒ†‹‡–Š‹‰Š‹’Š›–ƒ–‡ǦǤ……‡••–‘ˆ‡‡†™ƒ•‰‹˜‡„‡–™‡‡–Š‡

Š‘—”•‘ˆͲͺǣͲͲ–‘ͲͻǣͲͲǡͳʹǣͲͲ–‘ͳ͵ǣͲͲǡͳ͸ǣ͵Ͳ–‘ͳ͹ǣ͵ͲǡʹͳǣͲͲ–‘ʹʹǣͲͲƒ†ˆ”‘ͲʹǣͲͲ–‘

ͲͶǣͲͲǤ•–Š‡”‡™ƒ•‘‡ˆˆ‡…–‘ˆ‘Ž›ƒ…‹†ƒ††‹–‹‘‹‡š’‡”‹‡–ǡ–Š‡†‹‡–™ƒ•

‘‹––‡† ˆ”‘ –Š‡ •‡…‘† …Š‹…‡ ‡š’‡”‹‡–Ǥ Š‡ †‹‡–• ™‡”‡ ‡‹–Š‡” ™‹–Š ‘ ƒ††‹–‹‘

ȋȌǡ™‹–ŠͷͲͲ C. braakiiǦ†‡”‹˜‡†’Š›–ƒ•‡ƒ††‡†ȋȌ‘”™‹–Š„‘–Š’Š›–ƒ•‡ƒ†

ˆ‘”‹…ƒ…‹†ƒ††‡†ȋȌǤ–ͳͻ†ƒ›•‘ˆƒ‰‡ǡ‡š…”‡–ƒ™‡”‡…‘ŽŽ‡…–‡†ˆ‘”ƒ••‡••‡–‘ˆ

”‡–‡–‹‘Ǥ–ʹͲƒ†ʹͳ†ƒ›•‘ˆƒ‰‡ǡ„‹”†•–Šƒ–Šƒ†„‡‡™‹–Š‘—–ˆ‡‡†ˆ‘”‡‹‰Š–Š‘—”•™‡”‡

ʹ͹

ˆ‡†ˆ‘”‘‡Š‘—”ǤŠ‡…”‘’ƒ†‰‹œœƒ”†…‘–‡–™ƒ•…‘ŽŽ‡…–‡†‹‡†‹ƒ–‡Ž›„‡ˆ‘”‡–Š‡•–ƒ”–

‘ˆ–Š‡ˆ‡‡†‹‰ƒ†–Š‡”‡ƒˆ–‡”‡˜‡”›ʹͲ‹—–‡•—–‹ŽͳͶͲ‹—–‡•ƒˆ–‡”–Š‡•–ƒ”–‘ˆ–Š‡

ˆ‡‡†‹‰Ǥ–͸Ͳƒ†ͳͶͲ‹—–‡•ǡƒŽ•‘…‘–‡–•ˆ”‘Œ‡Œ——ƒ†‹Ž‡—™‡”‡…‘ŽŽ‡…–‡†Ǥ‹‡–

’”‘†—…–‹‘ǡ•ƒ’Ž‡’”‡’ƒ”ƒ–‹‘ƒ†ƒƒŽ›–‹…ƒŽ’”‘…‡†—”‡•ƒ”‡†‡•…”‹„‡†‹ƒ’‡”Ǥ

3.5 Experiment III

Effect of acid addition on phytase efficacy in pigs

Š‹”–›Ǧ–™‘’‹‰•™‡”‡ˆ‡†ƒŠ‹‰Š’Š›–ƒ–‡Ǧ†‹‡–ȋȌǡ‘”–Š‡•ƒ‡†‹‡–™‹–Šˆ‘”‹…ƒ…‹†

ƒ††‡†ȋȌǡͷͲͲ Ǥ„”ƒƒ‹‹Ǧ†‡”‹˜‡†’Š›–ƒ•‡ƒ††‡†ȋȌ‘”„‘–Šƒ††‡†ȋȌ‹

ƒʹšʹˆƒ…–‘”‹ƒŽ†‡•‹‰Ǥ–ͷʹ†ƒ›•‘ˆƒ‰‡ƒ†ƒƒ˜‡”ƒ‰‡‹‹–‹ƒŽ„‘†›™‡‹‰Š–‘ˆʹͳǤͲ͸᩿‰ǡ

–Š‡’‹‰•™‡”‡‡“—ƒŽŽ›†‹•–”‹„—–‡†„›™‡‹‰Š–ƒ†”ƒ†‘Ž›†‹•–”‹„—–‡†‹‰”‘—’•‘ˆˆ‘—”Ǥ

Š‡›™‡”‡‡’–‹‡‹‰Š–†‹ˆˆ‡”‡–…‘…”‡–‡ǦˆŽ‘‘”‡†ǡ’ƒ”–‹ƒŽŽ›•Žƒ––‡†’‡•ǡ™‹–Š‹†‹˜‹†—ƒŽ

ˆ‡‡†‹‰ •–ƒ–‹‘•Ǥ Š‡ ‹†‹˜‹†—ƒŽ ˆ‡‡†‹‰ •–ƒ–‹‘• ‡ƒ„Ž‡† ”‡…‘”†‹‰ ‘ˆ ‹†‹˜‹†—ƒŽ ˆ‡‡†

‹–ƒ‡ƒ†–Šƒ–ƒŽŽˆ‘—”†‹‡–ƒ”›–”‡ƒ–‡–•™‡”‡”‡’”‡•‡–‡†‹ƒŽŽ’‡•ǤŠ‡’‹‰•™‡”‡ˆ‡†

‡“—ƒŽƒ‘—–•–™‹…‡†ƒ‹Ž›ƒ–ͲͺǣͲͲƒ†ͳͶǣͲͲ„ƒ•‡†‘ƒ‡•–‹ƒ–‡†ˆ‡‡†‹–ƒ‡‘ˆ͵Ψ‘ˆ

–Š‡ Ž‹˜‡ „‘†› ™‡‹‰Š–ǡ ƒ† Ž‡ˆ–‘˜‡”• ™‡”‡ ”‡…‘”†‡†Ǥ †‹˜‹†—ƒŽ ˆƒ‡…ƒŽ •ƒ’Ž‡• ™‡”‡

…‘ŽŽ‡…–‡† ƒ– ‡š’‡”‹‡– †ƒ›• ʹͶ –‘ ʹ͹Ǥ ‹••‡…–‹‘ ™ƒ• …ƒ””‹‡† ‘—– ‘ †ƒ› ʹͺ ƒ† ʹͻǡ

•–ƒ”–‹‰ –™‘ Š‘—”• ƒˆ–‡” –Š‡ ‘”‹‰ ˆ‡‡†‹‰Ǥ –‘ƒ…Šǡ Œ‡Œ—— ƒ† ‹Ž‡— ’ ™‡”‡

‡ƒ•—”‡†Ǥ–‡•–‹ƒŽ…‘–‡–ˆ”‘–Š‡Œ‡Œ——ƒ†–Š‡‹Ž‡—ƒ†–Š‡Ž‡ˆ––Š‹”†ƒ†ˆ‘—”–Š

‡–ƒ…ƒ”’ƒŽ ˆ”‘ ‡ƒ…Š ’‹‰ ™‡”‡ …‘ŽŽ‡…–‡†Ǥ ‹‡– ’”‘†—…–‹‘ǡ •ƒ’Ž‡ ’”‡’ƒ”ƒ–‹‘ ƒ†

ƒƒŽ›–‹…ƒŽ’”‘…‡†—”‡•ƒ”‡†‡•…”‹„‡†‹ƒ’‡”Ǥ