UNIVERSITI PUTRA MALAYSIA
AMIN MIRSHEKARI
FP 2012 34
EFFECTS OF HOT WATER DIP TREATMENT ON PHYSICOCHEMICAL CHARACTERISTICS, ANTHRACNOSE (Colletotrichum musae)
CONTROL AND ENZYMATIC ACTIVITY OF BERANGAN BANANA (Musa sapientum L.) DURING RIPENING
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EFFECTS OF HOT WATER DIP TREATMENT ON PHYSICOCHEMICAL CHARACTERISTICS, ANTHRACNOSE (Colletotrichum musae)
CONTROL AND ENZYMATIC ACTIVITY OF BERANGAN BANANA (Musa sapientum L.) DURING RIPENING
By
AMIN MIRSHEKARI
Thesis Submitted to the School of Graduate Studies, Universiti Putra Malaysia, in Fulfilment of the Requirements for the Degree of Doctor of
Philosophy
September 2012
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DEDICATIONDEDICATIONDEDICATIONDEDICATION
DEDICATION
This thesis is dedicated to all I love specially
To the soul of my parents, my beloved mother and father
In the heaven who regretfully did not live to see this work.
To my beloved wife Aazam, my daughters, Delara and Sara
For the unconditional patience, love and support.
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Abstract of thesis presented to the Senate of Universiti Putra Malaysia in fulfilment of the requirement for the degree of Doctor of Philosophy
ABSTRACT
EFFECTS OF HOT WATER DIP TREATMENT ON PHYSICOCHEMICAL CHARACTERISTICS, ANTHRACNOSE (Colletotrichum musae)
CONTROL AND ENZYMATIC ACTIVITY OF BERANGAN BANANA (Musa sapientum L.) DURING RIPENING
By
AMIN MIRSHEKARI
September 2012
Chairperson: Phebe Ding, PhD
Faculty: Agriculture
Banana, as an important export fruit, is a highly perishable commodity.
Postharvest rot is the major factor limiting the extension of storage life of
banana. The beneficial effect of heat treatment to control decay and prolong
the shelf life has been reported on various fruits. To achieve the best
combination of water temperature and dipping time, Berangan banana fruit
was treated with water at 25 (control), 45, 50 and 55 ºC for 10, 20 and 30
min.
The results obtained in the first experiment showed that fruit firmness, peel
and pulp colour (hº) increased significantly, while respiration rate, ethylene
production, peel and pulp colour (L* and C*), soluble solids concentration
(SSC), titratable acidity (TA) and pH decreased significantly when hot water
temperature and dipping time increased. Hot water of 45 ºC for 10 and 20
min dipping time did not show any significant difference in fruit quality as
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compared to control. Hot water dip treatment of 45 ºC for 30 and 55 ºC for
10 min retarded some of fruit ripening processes in Berangan banana. The
low respiration rate, ethylene production, peel and pulp colour (L*, C* and
hº), SSC, TA and pH, and high firmness in Berangan banana treated with hot
water of 50 ºC for 10 and 20 min indicated that this combination treatment
could delay fruit ripening. Fruit dipped in hot water of 50 ºC for 30 min and
55 ºC for 20 and 30 min illustrated slight to severe peel blackening and
suppressed ripening process in Berangan banana.
For anthracnose control, fruit was treated with hot water at 50 ºC for 0, 10
and 20 min without or with fungicide. The results clearly demonstrated that
hot water without or with fungicide reduced severity of anthracnose in
inoculated Berangan banana. Spore germination of Colletotrichum musae
was suppressed completely with hot water of 50 ºC in 10 and 20 min without
or with fungicide. Anthracnose symptoms did not develop in fruits dipped
with hot water at 50 ºC for 20 min either with or without fungicide during 7
days of ripening.
The results of enzymatic activity experiment indicated that hot water dip at
50 ºC for 10 and 20 min could decrease cell wall degrading enzymes activity
such as polygalacturonase (PG), pectin methylesterase (PME) and pectate
lyase (PL) of Berangan banana fruit. The scanning electron microscope
(SEM) micrographs showed that hot water dip at 50 ºC for 10 and 20 min
could melt the fruit epicuticular waxes, so that it covered and sealed the
microscopic cracks on the fruit surface. The results of ultrastructural
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investigation indicated that hot water of 50 ºC for 20 min has lesser loss of
cell rigidity and dissolution of cell wall pectin fractions of tissues in Berangan
banana. It can be concluded that hot water dip treatment using 50 ºC for 10
min showed reasonable delay of fruit ripening and anthracnose control as
compared to other combination of temperature and dipping time. This
combination of temperature and dipping time could be the most possible to
be used as a commercial treatment in extending the shelf life of Berangan
banana for local consumption and export purposes.
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Abstrak tesis yang dikemukakan kepada Senat Universiti Putra Malaysia sebagai memenuhi keperluan untuk ijazah Doktor Falsafah
ABSTRAK
KESAN RAWATAN AIR PANAS KE ATAS SIFAT FIZIKOKIMIA, KAWALAN ANTRAKNOS (Colletotrichum musae) DAN AKTIVITI ENZIM
PISANG BERANGAN (Musa sapientum L.) SEMASA PERANUMAN
Oleh
AMIN MIRSHEKARI
September 2012
Pengerusi: Phebe Ding, PhD
Fakulti: Pertanian
Pisang adalah satu komoditi eksport yang penting dan sangat mudah rosak.
Reput lepas tuai adalah faktor utama yang menghadkan pemanjangan hayat
penyimpanan pisang. Kelebihan rawatan haba dalam mengawal pereputan
dan memanjangkan jangka hayat telah dilaporkan untuk pelbagai jenis buah-
buahan. Untuk mencapai kombinasi yang terbaik antara suhu air dan masa
rendaman, buah pisang Berangan dirawat dengan air pada suhu 25
(kawalan), 45, 50 dan 55 ºC selama 10, 20 dan 30 minit.
Keputusan yang diperoleh dalam kajian pertama menunjukkan bahawa
kekerasan, warna kulit dan isi (hº) buah meningkat secara signifikan,
manakala kadar respirasi, penghasilan etilena, warna kulit dan isi (L* dan
C*), kandungan pepejal terlarut (SSC), asid tertitrat (TA) dan pH menurun
secara signifikan apabila suhu air panas dan masa rendaman meningkat. Air
panas bersuhu 45 ºC dan masa rendaman selama 10 dan 20 minit, tidak
menunjukkan sebarang perbezaan yang signifikan berbanding dengan
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kawalan. Rawatan rendaman air panas 45 ºC selama 30 minit dan 55 ºC
selama 10 minit merencatkan beberapa proses peranuman buah pisang
Berangan. Kadar respirasi, penghasilan etilena, warna kulit dan isi (L *, C*
dan hº), kandungan pepejal terlarut, asid tertitrat, pH dan kekerasan adalah
rendah pada buah pisang Berangan yang dirawat dengan air panas bersuhu
50 ºC selama 10 dan 20 minit. Ini menunjukkan bahawa gabungan rawatan
ini berupaya melambatkan peranuman buah. Buah yang direndam di dalam
air panas bersuhu 50 ºC selama 30 minit dan 55 ºC selama 20 dan 30 minit
menyebabkan sedikit sehingga banyak penghitaman warna kulit dan
mengganggu peranuman buah pisang Berangan.
Untuk kawalan antraknos, buah telah dirawat dengan air panas pada suhu
50 ºC selama 0, 10 dan 20 minit tanpa atau dengan racun kulat. Keputusan
jelas menunjukkan bahawa rawatan air panas tanpa atau dengan racun kulat
mengurangkan keseriusan antraknos pada buah pisang Berangan yang telah
diinokulasi. Percambahan spora Colletotrichum musae telah dihalang
sepenuhnya oleh air panas bersuhu 50 ºC selama 10 dan 20 minit tanpa
atau dengan racun kulat. Simptom antraknos juga tidak dapat dilihat pada
buah yang dicelup dengan air panas bersuhu 50 ºC selama 20 minit sama
ada dengan atau tanpa racun kulat selama 7 hari peranuman.
Keputusan eksperimen aktiviti enzim menunjukkan bahawa rendaman air
panas pada suhu 50 ºC selama 10 dan 20 minit boleh mengurangkan aktiviti
enzim pada dinding sel seperti poligalakturonase (PG), metilesterase pektin
(PME) dan pektat liase (PL) pada buah pisang Berangan. Mikrograf daripada
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mikroskopi pengimbas elektron (SEM) menunjukkan bahawa rendaman air
panas pada suhu 50 ºC selama 10 dan 20 minit boleh mencairkan lapisan
lilin buah epicuticular, dan ia melitupi retakan mikroskopik yang terdapat
pada permukaan buah. Keputusan kajian ultrastruktur menunjukkan bahawa
rendaman air panas bersuhu 50 ºC selama 20 minit dapat mengekalkan
ketegaran sel dan pelarutan pektin dinding sel pada buah pisang Berangan.
Kesimpulannya, rawatan rendaman air panas menggunakan suhu 50 ºC
selama 10 minit dapat menangguhkan peranuman buah pisang dan
mengawal antraknos berbanding dengan kombinasi suhu dan masa
rendaman yang lain. Ia boleh dijadikan kemungkinan yang terbaik untuk
digunakan sebagai rawatan komersial bagi melanjutkan jangka hayat buah
pisang Berangan untuk tujuan kegunaan tempatan dan eksport.
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ACKNOWLEDGEMENTS
Most of all, all praises and endless thanks to God, the Almighty. The most
Beneficent and Merciful to making it possible for me that complete this
investigation.
It pleased me to take this opportunity to convey my deepest appreciation and
gratitude to my supervisor Dr. Phebe Ding for her generous help, invaluable
guidance, patience and support throughout the completion of this thesis. My
grateful appreciation is also due to Associate Professor Dr. Jugah Kadir for
his guidance, valuable advice and helpful suggestions. I am also obliged and
grateful to Professor Dr. Hasanah Mohd Ghazali for her invaluable
assistance. I really appreciate them for serving on my supervisory committee.
I would like to thank all the staff of the Department of Crop Science, Faculty
of Agriculture, Universiti Putra Malaysia (UPM) especially Mr. Azhar Othman
the technician of postharvest laboratory. I want to give utmost appreciation
to all my friends especially Dr. Mohammad Houshmand, Parviz Almasi, Tee
Yei Kheng and Hamizah Hassan. My profound and heartiest thanks and love
to my wife for her love, patience, encouragement and constant support
during this study. My special and heartiest thanks and love towards our
sweet daughters for being a kind and constant encouragement during the
entire study period. Last but certainly not least, I wish to express my sincere
appreciation to all those who are not mentioned here that helped me to
ensure the completion of my study.
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APPROVAL I certify that a Thesis Examination Committee has met on 3th September 2012 to conduct the final examination of Amin Mirshekari on his thesis entitled “Effects of Hot Water Dip Treatment on Physicochemical Characteristics, Anthracnose (Colletotrichum musae) Control and Enzymatic Activity of Berangan banana (Musa sapientum L.) During Ripening” in accordance with the Universities and University Colleges Act 1971 and the Constitution of the Universiti Putra Malaysia [P.U. (A) 106] 15 March 1998. The Committee recommends that the student be awarded the Doctor of Philosophy. Members of the Thesis Examination Committee were as follows:
Izham Ahmad, PhD Associate Professor Faculty of Agriculture Universiti Putra Malaysia (Chairman) Mahmud Tenku Muda Mohamed, PhD Professor Faculty of Agriculture Universiti Putra Malaysia (Internal Examiner) Lau Wei Hong, PhD Senior Lecturer Faculty of Agriculture Universiti Putra Malaysia (Internal Examiner) Chien Y. Wang, PhD Associate Professor Laboratory of Food Quality Unated State Department of Agriculture (USDA) (External Examiner)
SEOW HENG FONG,PhD Professor and Deputy Dean School of Graduate Studies Universiti Putra Malaysia
Date:
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This thesis was submitted to the Senate of Universiti Putra Malaysia and has been accepted as fulfilment of the requirement for the degree of Doctor of Philosophy. The members of the Supervisory Committee were as follows:
PHEBE DING, PhD Senior Lecturer Faculty of Agriculture Universiti Putra Malaysia (Chairperson) JUGAH KADIR, PhD Associate Professor Faculty of Agriculture Universiti Putra Malaysia (Member) HASANAH MOHD GHAZALI, PhD Professor Faculty of Food Science and Technology Universiti Putra Malaysia (Member)
BUJANG BIN KIM HUAT,PhD Professor and Dean School of Graduate Studies Universiti Putra Malaysia
Date:
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DECLARATION
I declare that the thesis is my original work except for quotations and citations, which have been duly acknowledged. I also declare that it has not been previously, and is not concurrently, submitted for any other degree at Universiti Putra Malaysia or at any other institution.
AMIN MIRSHEKARI
Date: 3 September 2012
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TABLE OF CONTENTS
Page
DEDICATION ii�ABSTRACT iii�
ABSTRAK vi�ACKNOWLEDGEMENTS ix�APPROVAL x�DECLARATION xii�
LIST OF TABLES xvii�LIST OF FIGURES xx�LIST OF ABBREVIATIONS xxii�
CHAPTER 1� GENERAL INTRODUCTION 1�
2� LITERATURE REVIEW 5�
2.1� Description of banana 5�
2.2� Fruit ripening 9�
2.2.1� Respiration rate and ethylene production 9�
2.2.2� Colour 12�
2.2.3� Texture 14�
2.2.4� Soluble solids concentration 16�
2.2.5� Titratable acidity and pH 17�
2.2.6� Enzyme activity 19�
2.3� Anthracnose 21�
2.4� Heat treatment 23�
3� EFFECTS OF HOT WATER DIP TREATMENT ON PHYSICOCHEMICAL CHARACTERISTICS OF BERANGAN BANANA DURING RIPENING 32�
3.1� Introduction 32�
3.2� Materials and methods 35�
3.2.1� Plant materials and treatments 35�
3.2.2� Determination of respiration rate and ethylene production 36�
3.2.3� Determination of peel and pulp colour 36�
3.2.4� Determination of firmness 37�
3.2.5� Determination of soluble solids concentration 37�
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3.2.6� Determination of titratable acidity and pH 38�
3.2.7� Determination of peel blackening 39�
3.2.8� Experimental design and statistical analysis 39�
3.3� Resalts and discussion 40�
3.3.1� Respiration rate and ethylene production 40�
3.3.2� Peel colour 46�
3.3.3� Pulp colour 56�
3.3.4� Firmness 58�
3.3.5� Soluble solids concentration 65�
3.3.6� Titratable acidity 67�
3.3.7� pH 69�
3.3.8� Peel blackening 71�
3.4� Conclusion 73�
4� EFFECTS OF HOT WATER DIP TREATMENT IN COMPARISON WITH FUNGICIDE ON ANTHRACNOSE CONTROL OF BERANGAN BANANA 75�
4.1� Introduction 75�
4.2� Materials and methods 77�
4.2.1� Isolation of causal agent of anthracnose in banana 77�
4.2.1.1� Sample collection 77�
4.2.1.2� Isolation and identification of causal pathogen 77�
4.2.1.3� Preparation of conidia suspension 78�
4.2.1.4� Pathogenicity test 79�
4.2.2� In vitro study of hot water on Colletotrichum musae 79�
4.2.2.1� Preparation of inoculums 79�
4.2.2.2� Hot water dip treatment of conidia 80�
4.2.3� Evaluation of hot water on anthracnose development 80�
4.2.3.1� Plant material 80�
4.2.3.2� Fruit inoculation 81�
4.2.3.3� Hot water dip treatment and anthracnose assessment 81�
4.2.3.4� Preparation of specimen for scanning electron microscopy 82�
4.2.4� Experimental design and statistical analysis 82�
4.3� Results and discussion 83�
4.3.1� Identification of Colletotrichum musae 83�
4.3.2� Pathogenicity assay 83�
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4.3.3� Effect of hot water treatment on conidia germination of C. musae 85�
4.3.4� Effect of hot water on anthracnose development of banana fruit 87�
4.3.5� Observation of scanning electron microscopy micrographs91�
4.4� Conclusion 94�
5� EVALUATION OF FRUIT QUALITY CHARACTERISTICS OF BERANGAN BANANA AS AFFECTED BY HOT WATER DIP WITHOUT OR WITH FUNGICIDE 96�
5.1� Introduction 96�
5.2� Materials and methods 97�
5.2.1� Plant materials and treatments 97�
5.2.2� Determination of respiration rate and ethylene production 98�
5.2.3� Determination of peel and pulp colour 98�
5.2.4� Determination of firmness 99�
5.2.5� Determination of soluble solids concentration 99�
5.2.6� Determination of titratable acidity and pH 99�
5.2.7� Determination of weight loss 99�
5.2.8� Sensory evaluation of ripe fruit 100�
5.2.9� Experimental design and statistical analysis 100�
5.3� Results and discussion 101�
5.3.1� Respiration rate and ethylene production 101�
5.3.2� Peel colour 103�
5.3.3� Pulp colour 105�
5.3.4� Firmness 106�
5.3.5� Soluble solids concentration 108�
5.3.6� Titratable acidity and pH 108�
5.3.7� Weight loss 110�
5.3.8� Sensory evaluation 111�
5.4� Conclusion 112�
6� ENZYMATIC ACTIVITY AND MICROSTRUCTURAL CHANGES IN BERANGAN BANANA DURING RIPENING FOLLOWING HOT WATER DIP TREATMENT 114�
6.1� Introduction 114�
6.2� Materials and methods 117�
6.2.1� Plant material and treatment 117�
6.2.2� Determination of fruit firmness 118�
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6.2.3� Enzyme extraction 118�
6.2.4� Polygalacturonase assay 118�
6.2.5� Pectin methylesterase assay 119�
6.2.6� Pectate lyase assay 120�
6.2.7� Preparation of peel specimen for scanning electron microscopy 121�
6.2.8� Preparation of peel specimen for transmission electron microscopy 121�
6.2.9� Determination of electrolyte leakage 122�
6.2.10�Experimental design and statistical analysis 122�
6.3� Results and discussion 123�
6.3.1� Fruit firmness 123�
6.3.2� Polygalacturonase activity 124�
6.3.3� Pectin methylesterase activity 128�
6.3.4� Pectate lyase activity 129�
6.3.5� Correlation between firmness and enzyme activity 130�
6.3.6Observation of scanning electron microscopy micrograph 132�
6.3.7� Observation of transmission electron microscopy micrograph 134�
6.3.8� Electrolyte leakage 136�
6.4� Conclusion 138�
7� GENERAL CONCLUSIONS AND RECOMMENDATIONS 139�
7.1� Conclusions 139�
7.2� Recommendations for future research 141�
REFERENCES 142�
BIODATA OF STUDENT 161�LIST OF PUBLICATIONS 162�
