HE IS THE FIRST, HE IS THE LAST

HE IS THE MANIFEST, HE IS THE HIDDEN, &

HE KNOWS EVERYTHING HE BRINGS THE NIGHT INTO THE DAY, &

BRINGS THE DAY INTO THE NIGHT, & HE KNOWS THE THOUGHTS OF THE

HEARTS S.AL-HADID-385 (AL-QURAN)

Fruit quality and storability of Kinnow mandarin (Citrus reticulata Blanco) in relation to tree age

BY

SAMINA KHALID M.Sc. (Hons.) Horticulture

2007-ag-09

A thesis submitted in the fulfillment of

the requirements for the degree of

Doctor of Philosophy

in

Horticulture

Institute of Horticultural Sciences Faculty of Agriculture

UNIVERSITY OF AGRICULTURE, FAISALABAD PUNJAB, PAKISTAN

2013

To The Controller of Examinations University of Agriculture

Faisalabad

We, the supervisory committee, certify that the contents and form of this thesis

submitted by Miss Samina Khalid Regd. No. 2007-ag-09, have been found satisfactory, and recommend it to be processed for evaluation by the external examiner(s) for the

award of the degree.

SUPERVISORY COMMITTEE

1) SUPERVISOR:

_________________________________ (PROF. DR. AMAN ULLAH MALIK)

2) MEMBER: __________________________________

(DR. AHMAD SATTAR KHAN)

3) MEMBER: __________________________________

(DR. AMER JAMIL)

i

ACKNOWLEDGEMENTS

Each and every praise is due to Almighty Allah, the omnipotent and benefactor, “Who taught Adam the names of all the things”, and “He taught man that which he knew not”. All the blessings upon Holy Prophet (P.B.U.H) who conveyed Allah’s message with full devotion.

Special thanks are due to my supervisor Prof. Dr. Aman Ullah Malik for his invaluable advice, encouragement and support at every step of my PhD studies. Achieving this degree would not have been possible without his support. My gratitude is also for Dr. Ahmad Sattar Khan, Institute of Horticultural Sciences, University of Agriculture, Faisalabad and Dr. Amer Jamil, Department of Chemistry and Biochemistry, Faculty of Sciences, University of Agriculture, Faisalabad whose knowledge and comments contributed significantly to attaining this goal.

I would like to acknowledge the support of my parent Department Institute of Horticultural Sciences, UAF, its staff and Director. I am also thankful for the assistance and support provided by Dr. Saeed Ahmad and his Staff (Shakil Zahid, Shakil Latif and Farhan) at Pomology Laboratory, Institute of Horticultural Sciences, University of Agriculture, Faisalabad, for providing facilities of macro-nutrient analysis.

Sincere thanks are extended to Dr. Muhammad Zargham Khan Chairman/Professor, Department of Pathology, Faculty of Veterinary Science, University of Agriculture, Faisalabad, his students and laboratory staff for their valuable guidance during anatomical studies. Special thanks to Dr. Muhammad Shahid Assistant Professor, Department of Chemistry and Biochemistry, Faculty of Sciences, for his support in antioxidant and phenolics determination. I am also grateful to Mr. Abdul Qudoos Research Officer, Central Hi-Tech Laboratory, University of Agriculture, Faisalabad, for his assistance in micro-nutrient analysis.

I am also thankful to Prof. Dr. Zora Singh, Department of Environment and Agriculture, School of Science, Faculty of Science and Engineering, Curtin University, Western Australia for his kind guidance and support during my six months training at Australia, under International Research Support Initiative Programme (IRSIP), Programe of Higher education Commission (HEC).

I gratefully acknowledge the help of Dr. Basharat Ali Saleem, Assistant Director Fruit and Vegetable Development Project, Punjab Agriculture Department during the course of my PhD studies. I would like to thank Mr. Tariq Mehmood Cheema, Kinnow mandarin grower at Chack No. 99 NB, Sargodha for his assistance and hospitality.

I am also thankfull to my lab fellows Muhammad Shafique Khalid, Kashif Razzaq, Sami Ullah, Muhammad Shafique, Muhammad Amin, Mudassar Naseer, Omer Hafeez Malik, Munaza Saeed, Habat Ullah Asad and Syed Ali Raza, for their cooperation in an excellent working atmosphere during my research work. I want to express my great appreciation, sincerest gratitude and special thanks to the lab staff including Miss Rabia Hameed, Abdul Haseeb, Muhammad Afzal, and Javed Maseeh for their assistance in laboratory work.

I am gratefull to Higher Education Commission (HEC), Islamabad, Pakistan, for awarding me scholarship under Indigenous 5000 PhD Fellowship scheme and six months research fellowship under International Research Support Initiative Programme (IRSIP) to conduct a part of my PhD research at Curtin University, Perth Western Australia.

I also express my gratitude to my friends for their help and cooperation during my PhD studies.

Last and most important I would like to thank my family for their patience, encouragement and financial support during my studies. I am also grateful to all those whom I mentioned or could not mention here but they helped me positively.

(Samina Khalid)

ii

TABLE OF CONTENTS

Chapter

Title Page

Acknowledgements i Table of contents ii List of tables vii List of figures x List of symbols and abbreviations xii ABSTRACT 1 1 GENERAL INTRODUCTION 3 2 GENERAL REVIEW OF LITERATURE 6 2.1 Introduction 6 2.2 Origin and Distribution 6 2.3 Citrus Industry of the World 7 2.3.1 World Mandarin and Tangerine Production 8 2.3.2 World Mandarin and Tangerine Export 8 2.4 Citrus Industry of Pakistan 9 2.4.1 Mandarin export from Pakistan 9 2.5 Citrus Fruit Anatomy 10 2.6. Fruit Growth and Development 10 2.7 Fruit Quality 11 2.7.1 Factors affecting fruit quality 11 2.7.1.1 Tree age 11 2.7.1.1.1 Physical fruit quality 11 2.7.1.1.2 Biochemical fruit quality 12 2.7.1.2 Fruit size 12 2.7.1.2.1 Physical fruit quality 12 2.7.1.2.2 Biochemical fruit quality 13 2.7.1.3 Canopy position 14 2.7.1.3.1 Physical fruit quality 14 2.7.1.3.2 Biochemical fruit quality 14 2.7.1.4 Plant growth regulators 16 2.7.1.4.1 Physical fruit quality 17 2.7.1.4.2 Biochemical fruit quality 20 2.7.1.5 Mineral nutrition 21 2.7.1.5.1 Physical fruit quality 23 2.7.1.5.2 Biochemical fruit quality 24 3 GENERAL MATERIALS AND METHODS 26 3.1 Plant material 26 3.2 Physical fruit quality 27 3.2.1 Fruit colour (score) 27 3.2.2 Fruit smoothness (score) 27 3.2.3 Fruit softness (score) 27 3.2.4 Fruit mass (g) 27 3.2.5 Fruit diameter (mm) 27 3.2.6 Fruit mass loss (%) 27 3.2.7 Rind thickness (mm) 27

iii

Chapter

Title Page

3.2.8 Rind mass (%) 27 3.2.9 Rag mass (%) 28 3.2.10 Seed mass (%) 28 3.2.11 Seed number 28 3.2.12 Healthy seed (%) 28 3.2.13 Aborted seed (%) 28 3.2.14 Juice mass (%) 28 3.3 Biochemical fruit quality 29 3.3.1 pH 29 3.3.2 TSS (°Brix) 29 3.3.3 Titratable acidity (TA) (%) 29 3.3.4 Ascorbic acid (AA) (mg 100 mL-1) 29 3.3.5 Sugars (reducing, non reducing and total sugars) 29 3.4 Statistical Analysis 29 4 EFFECT OF TREE AGE AND FRUIT CANOPY

POSITION ON FRUIT QUALITY OF 'KINNOW' MANDARIN

30

4.1 Introduction 30 4.2 Materials and Methods 32 4.2.1 Plant material and site selection 32 4.2.2 Physical fruit quality 32 4.2.3 Biochemical fruit quality 32 4.2.4 Nutrient analysis 33 4.2.4.1 Fruit sample preparation 33 4.2.4.2 Nitrogen (N) 33 4.2.4.3 Estimation of elements other than N 34 4.2.4.3.1 Phosphorous (P) 34 4.2.4.3.2 Potassium (K) 34 4.2.4.3.3 Calcium (Ca) and micronutrients 34 4.2.5 Statistical analysis 35 4.3 Results 35 4.3.1 Physical fruit quality 35 4.3.2 Biochemical fruit quality 35 4.3.3 Rind macro-nutrient concentrations 39 4.3.4 Rind micro-nutrient concentrations 39 4.3.5 Correlation between rind nutrient concentrations and

rind quality 39

4.3.6 Correlation between rind nutrient concentrations and internal fruit quality

41

4.4 Discussion 41 5 EFFECT OF TREE AGE AND FRUIT SIZE ON

STORAGE POTENTIAL OF 'KINNOW' MANDARIN

48

5.1 Introduction 48 5.2 Materials and methods 49

iv

Chapter

Title Page

5.2.1 Plant material and site selection 49 5.2.2 Influence of tree age and fruit size on shelflife of

'Kinnow' mandarin 49

5.2.3 Influence of tree age and fruit size on storage life of 'Kinnow' mandarin

50

5.2.4 Physical fruit quality 50 5.2.4.1 Respiration and ethylene production 50 5.2.5 Biochemical fruit quality 50 5.2.5.5 Total phenolic concentrations (TPC) and antioxidants 50 5.2.6 Statistical analysis 51 5.3 Results 51 5.3.1 Influence of tree age and fruit size on fruit quality of

'Kinnow' mandarin under ambient conditions 51

5.3.1.1 Physical fruit quality 51 5.3.1.2 Biochemical fruit quality 54 5.3.1.3 Respiration and ethylene production during shelflife

studies 61

5.3.2 Influence of tree age and fruit size on storage life of 'Kinnow' mandarin

64

5.3.2.1 Physical fruit quality 64 5.3.2.2 Biochemical fruit quality 66 5.3.2.3 Fruit mass loss (%) during shelflife and storage 78 5.4 Discussion 80 5.4.1 Ambient conditions studies 80 5.4.2 Cold storage studies 82 6 'KINNOW' FRUIT GROWTH AND

DEVELOPMENT IN RELATION TO CHANGES IN ENDOGENOUS LEVELS OF NUTRIENTS AND PECTIN

85

6.1 Introduction 85 6.2 Materials and method 87 6.2.1 Plant material and site selection 87 6.2.2 Fruit diameter (mm) 87 6.2.3 Nutrient analysis 87 6.2.3.1 Leaf sampling and sample preparation 87 6.2.3.2 Nitrogen (N) 88 6.2.3.3 Estimation of elements other than N 88 6.2.4 Pectin analysis 88 6.2.4.1 Alcohol insoluble solids 88 6.2.4.2 Total pectin 88 6.2.4.3 Water soluble pectin 88 6.2.4.4 Pectin determination 89 6.2.5 Anatomical studies 89 6.2.6 Statistical analysis 89 6.3 Results 89

v

Chapter

Title Page

6.3.1 Fruit diameter in relation to tree age 89 6.3.2 Cell number and cell size in relation to tree age 90 6.3.3 Nutrient concentrations during fruit growth and

development 91

6.3.3.1 Rind N concentrations 91 6.3.3.2 Rind P concentrations 92 6.3.3.3 Rind K concentrations 92 6.3.3.4 Rag N concentrations 92 6.3.3.5 Rag P concentrations 92 6.3.3.6 Rag K concentrations 92 6.3.3.7 Leaf N concentrations 93 6.3.3.8 Leaf P concentrations 93 6.3.3.9 Leaf K concentrations 93 6.3.4 Pectin concentrations during fruit growth and

development 95

6.3.4.1 Rind total pectin 95 6.3.4.2 Rind water soluble pectin 95 6.3.4.3 Rind protopectin 95 6.3.4.4 Rag total pectin 95 6.3.4.5 Rag water soluble pectin 96 6.3.4.6 Rag protopectin 96 6.3.5 Correlation between leaf nutrient concentrations and

fruit growth and development 96

6.3.6 Correlation between rind nutrients and fruit growth and development

98

6.3.7 Correlation between rag nutrients and fruit growth and development

99

6.4 Discussion 99 7 EFFECT OF TIMING OF PLANT GROWTH

REGULATORS APPLICATION ON FRUIT QUALITY AND STORAGE POTENTIAL OF 'KINNOW' MANDARIN

106

7.1 Introduction 106 7.2 Materials and methods 107 7.2.1 Plant material and site selection 107 7.2.2 Effect of application time of PGRs on shelflife of

'Kinnow' mandarin 108

7.2.3 Effect of application time of PGRs on storage life of 'Kinnow' mandarin

108

7.2.4 Physical fruit quality 108 7.2.5 Biochemical fruit quality 108 7.2.6 Statistical analysis 109 7.3 Results 109 7.3.1 Effect of time of PGRs application on fruit quality of 109 'Kinnow' mandarin during shelf life

vi

Chapter

Title Page

7.3.1.1 Physical fruit quality 109 7.3.1.2 Biochemical fruit quality 114 7.3.1.3 Fruit mass loss (%) 119 7.3.2 Effect of application time of PGRs on storage life of

'Kinnow' mandarin 120

7.3.2.1 Physical fruit quality 120 7.3.2.2 Biochemical fruit quality 122 7.3.2.3 Fruit mass loss (%) 124 7.4 Discussion 125 8 EXOGENOUS APPLICATIONS OF PGRS AND

NUTRIENT ON FRUIT QUALITY AND STORAGE OF 'KINNOW' MANDARIN

128

8.1 Introduction 128 8.2 Materials and method 129 8.2.1 Plant material and site selection 129 8.2.2 Influence of exogenous applications of plant growth

regulators on fruit quality of young 'Kinnow' mandarin trees

130

8.2.3 Influence of nutrients on fruit quality of young 'Kinnow' mandarin trees

130

8.2.4 Influence of PGRs and nutrients on fruit quality of young 'Kinnow' mandarin trees

131

8.2.5 Physical fruit quality 132 8.2.6 Biochemical fruit quality 132 8.2.7 Statistical analysis 132 8.3 Results 132 8.3.1 Influence of exogenous applications of plant growth

regulators on fruit quality of young 'Kinnow' mandarin trees

132

8.3.1.1 Physical fruit quality 132 8.3.1.2 Biochemical fruit quality 138 8.3.1.3 Discussion 141 8.3.2 Influence of nutrients on fruit quality of young

'Kinnow' mandarin trees 145

8.3.2.1 Physical fruit quality 145 8.3.2.2 Biochemical fruit quality 146 8.3.2.3 Discussion 151 8.3.3 Influence of PGRs and nutrients on fruit quality of

young 'Kinnow' mandarin trees 152

8.3.3.1 Physical fruit quality 152 8.3.3.2 Biochemical fruit quality 159 8.3.3.3 Discussion 162 9 GENERAL CONCLUSION 164 REFERENCES 169 ANNEXURE 193

vii

LIST OF TABLES

Table Title Page 2.1 Mandarin export from Pakistan during 2008-09 9 4.1 Biochemical characters of 'Kinnow' mandarin fruit in

relation to tree age and canopy position 38

4.2 Biochemical parameters of 'Kinnow' mandarin in relation to tree age and canopy position

38

4.3 Rind nutrient concentrations and rind quality in relation to tree age

42

4.4 Rind macro-nutrients and physical fruit quality of 'Kinnow' mandarin in relation to tree age

43

4.5 Rind micro-nutrients and physical fruit quality of 'Kinnow' mandarin in relation to tree age

44

4.6 Effect of tree age and fruit size on fruit mass (g) of 'Kinnow' mandarin

52

4.7 Effect of tree age, fruit size and shelf duration on physical fruit quality of 'Kinnow' mandarin

52

4.8 Effect of tree age, fruit size and shelf duration influencing biochemical fruit quality of 'Kinnow' mandarin

55

4.9 Influence of tree age, fruit size and shelf duration on total, reducing and non reducing sugar (%) of 'Kinnow' mandarin juice

57

4.10 Influence of tree age, fruit size and shelf duration on AA (mg 100ml-1), TPC (µg ml-1) and AO (mg µl-1) concentrations of 'Kinnow' mandarin juice

59

4.11 Effect of tree age, fruit size and analysis time on rind mass (%) of 'Kinnow' mandarin during storage

66

4.12 Effect of tree age and fruit size on rag mass (%) of 'Kinnow' mandarin fruit during cold storage

67

4.13 Juice mass (%) of 'Kinnow' mandarin fruit affected by tree age and fruit size during cold storage

67

4.14 Effect of tree age and fruit size on TSS (°Brix) of 'Kinnow' mandarin juice during cold storage

70

4.15 Influence of tree age and fruit size on titratable acidity (%) (TA) of 'Kinnow' mandarin juice during cold storage

70

4.16 TSS:TA of 'Kinnow' mandarin juice affected by tree age and fruit size during cold storage

71

4.17 Reducing sugars (%) of 'Kinnow' mandarin juice affected by tree age, fruit size and storage period

73

4.18 Non reducing sugars (%) of 'Kinnow' mandarin juice as affected by tree age, fruit size and storage period

74

4.19 Tree age, fruit size and storage period influencing total sugars (%) of 'Kinnow' mandarin juice

74

4.20 Tree age, fruit size and storage period influencing ascorbic acid concentration (mg 100 ml-1) of 'Kinnow' mandarin juice

76

4.21 Tree age, fruit size and storage period on antioxidant 76

viii

Table Title Page concentrations of 'Kinnow' mandarin juice

4.22 Total phenolic concentrations (µg ml-1) of 'Kinnow' mandarin juice as influenced by tree age, fruit size and storage period

77

4.23 Correlation between leaf nutrient concentrations and rind cell anatomy, fruit diameter (mm) and pectin concentrations (%)

100

4.24 Correlation between rind nutrient concentrations and rind cell anatomy, fruit diameter (mm) and pectin concentrations (%)

100

4.25 Correlation between rag nutrient concentrations and rind cell anatomy, fruit diameter (mm) and pectin concentrations (%)

101

4.26 Influence of PGRs and shelf duration on physical fruit quality of 'Kinnow' mandarin

110

4.27 Effect of PGRs and shelf duration on physical fruit quality of 'Kinnow' mandarin

112

4.28 Biochemical quality attributes of 'Kinnow' mandarin juice influenced by PGRs and shelf duration

115

4.29 Effect of PGRs and shelf duration on biochemical fruit quality

117

4.30 Effect of PGRs and time of application on mass loss (%) during shelf life of fruit

119

4.31 Effect of PGRs on external fruit quality of 'Kinnow' mandarin during storage

120

4.32 Physical fruit quality of 'Kinnow' mandarin influenced by PGRs during storage

121

4.33 Effect of PGRs on biochemical fruit quality of 'Kinnow' mandarin during storage

123

4.34 Effect of PGRs on biochemical fruit quality of 'Kinnow' mandarin during storage

124

4.35 Influence of PGRs on mass loss (%) during storage 125 4.36 Effect of plant growth regulators on seed quality of

'Kinnow' mandarin 134

4.37 Effect of PGRs on external fruit quality of 'Kinnow' mandarin

135

4.38 Effect of plant growth regulators on internal physical quality of fruit harvested from young 'Kinnow' mandarin trees

137

4.39 Contrast effect on ascorbic acid (mg 100mL-1) of 'Kinnow' mandarin juice

139

4.40 Effect of PGRs applications effect on TSS, titratable acidity (TA) (%) and TSS:TA concentrations of fruit obtained from young 'Kinnow' mandarin trees

140

4.41 Juice total, reducing, and non reducing sugars (%) of young 'Kinnow' mandarin trees as affected by plant growth regulators application

142

4.42 Effect of nutrients on seed quality and quantity variables

145

ix

Table Title Page 4.43 Effect of nutrients on rind thickness (mm), fruit

diameter (mm) and fruit mass (g) of 'Kinnow' mandarin 145

4.44 Fruit physical quality attributes as affected by nutrient applications

147

4.45 Fruit biochemical variables affected by nutrient application

148

4.46 Effect of nutrients on total, reducing and non reducing sugars (%) of 'Kinnow' mandarin juice

150

4.47 Seed quality parameters influenced by PGRs and nutrients

154

4.48 Fruit physical quality parameters influenced by PGRs and nutrients

155

4.49 Effect of plant growth regulators and nutrients on rind mass (%) of 'Kinnow' mandarin

157

4.50 Influence of plant growth regulators and nutrients on rag mass (%), rind mass (%) and juice mass (%) of 'Kinnow' mandarin

158

4.51 Nutrients and PGRs effect on TSS (Brix) and acidity (%) of 'Kinnow' mandarin juice

160

4.52 Influence of PGRs and nutrients on TSS:TA and ascorbic acid (mg 100ml-1) of 'Kinnow' mandarin juice

163

x

LIST OF FIGURES

Figure Title Page

2.1 World mandarin and tangerine production 8 2.2 World mandarin and tangerine export 8 2.3 Citrus area and production in Pakistan 9 2.4 Transverse section of citrus fruit 10 4.1 Effect of tree age and canopy position on rind colour

(score) (a) and rind smoothness score (b) of 'Kinnow' mandarin (n = 40)

35

4.2 Physical fruit quality of 'Kinnow' mandarin affected by tree age and canopy position (n = 40)

37

4.3 Rind macro-nutrient (a, b, c and d) and micro-nutrient (e, f, g and h) concentrations affected by tree age and canopy position (n = 40)

40

4.4 Fruit rind mass (%) (a, d), rag mass (%) (b, e) and juice mass (%) (c, f) influenced by tree age and fruit size during shelflife studies

53

4.5 Effect of tree age and fruit size on TSS (a, d), TA (%) (b, e) and TSS:TA (c, f) of 'Kinnow' mandarin fruit during shelf studies

56

4.6 Influence of tree age and fruit size on total sugars (%) (a, d), reducing sugars (%) (b, c) and non reducing sugars (%) (c, f) of 'Kinnow' mandarin fruit during shelf studies

58

4.7 Tree age and fruit size influenced on AA (mg 100ml-1) (a, d), TPC (µg ml-1) (b, e) and AO (mg µl-1) (c, f) of 'Kinnow' mandarin during seven days shelflife

60

4.8 Effect of tree age and fruit size on CO2 (m Mol kg-1h-1) production during shelflife studies of 'Kinnow' mandarin fruit

62

4.9 Effect of tree age and fruit size on mean CO2 (m Mol kg-1h-1) production during shelflife studies of 'Kinnow' mandarin fruit

62

4.10 Effect of tree age and fruit size on ethylene production (m Mol kg-1h-1) during seven days shelflife studies of 'Kinnow' mandarin fruit

63

4.11 Effect of tree age and fruit size on mean ethylene production (m Mol kg-1h-1) during seven days shelflife studies of 'Kinnow' mandarin

63

4.12 Effect of tree age and storage duration on rind mass (%) (a, d), rag mass (%) (b, e), and juice mass (%) (c, f) of 'Kinnow' mandarin fruit during cold storage

65

4.13 Influence of tree age and storage period on TSS (a, d), TA (%) (b, e) and TSS:TA (c, f) of 'Kinnow' mandarin fruit during cold storage

68

4.14 Effect of tree age and storage duration on reducing sugars (%) (a, d), non reducing sugars (%) (b, e) and total sugars (%) (c, f) of fruit during cold storage

72

4.15 Effect of tree age and fruit size on ascorbic acid (mg 75

xi

Figure Title Page

100ml-1) (a, d), antioxidant (mg µl-1) (b, e) and total phenolic concentrations (µg ml-1) (c, f) of 'Kinnow' mandarin juice during cold storage

4.16 Influence of tree age and fruit size on fruit mass loss (%) during shelflife studies

78

4.17 Effect of tree age and fruit size on mass loss (%) during cold storage

79

4.18 Effect of tree age and fruit size on mean mass loss (%) during cold storage

79

4.19 Fruit diameter (mm) (a) and increment in diameter (mm) (b) of 'Kinnow' mandarin fruit during fruit growth and development

90

4.20 Rind cell number (cells/mm2) (a) and cell size (mm2) (b) in relation to tree age

91

4.21 Rind, rag and leaf nutrient concentrations in relation to tree age

94

4.22 Influence of tree age on rind total pectin (%) (a), rind water soluble pectin (%) (b) and rind protopectin (%) (c) during fruit growth and development

97

4.23 Influence of tree age on rag total pectin (%) (a), rag water soluble pectin (%) (b) and rag protopectin (%) (c) during fruit growth and development

98

4.24 Effect of PGRs application time and shelf duration on physical fruit quality of 'Kinnow' mandarin

111

4.25 Effect of PGRs application time and shelf duration on rind mass (%) (a,d), rag mass (%) (b,e) and juice mass (%) (c, f) of 'Kinnow' mandarin

113

4.26 Effect of PGRs application time and shelf duration on TSS (°Brix) (a, e), acidity (%) (b, f), TSS:TA ratio (c, g) and ascorbic acid (mg 100 mL-1) (d, h) of 'Kinnow' mandarin

116

4.27 Effect of PGRs application time and shelf duration on reducing sugars (%) (a, d), non reducing sugars (%) (b, e) and total sugars (%) (c, f) of 'Kinnow' mandarin juice

118

4.28 Influence of PGRs and shelf duration on ascorbic acid (mg 100 mL1) concentrations of 'Kinnow' mandarin

138

4.29 Influence of nutrients on ascorbic acid concentrations (mg 100 mL-1) of juice during seven days shelf life studies

149

xii

LIST OF SYMBOLS AND ABBREVIATIONS

Abbreviation Description

@ At the rate of o Degree % Percent µg Microgram(s) µL Microliter(s) µM Micromolar µm Micrometer oC Degree Celsius $ United States dollar A Absorbance a* Colour coordinate (a) b* Colour coordinate (b) d Day 2, 4-D 2, 4 dichloro phenoxy acetic acid < Less than > Greater than oF Degree Fahrenheit ± Plus minus AA Ascorbic acid ANOVA Analysis of variance ATP Adenosine tri phosphate C* Chroma C20H14O4 Phenolphthalein C2H2 Ethylene Ca Calcium Ca+ Calcium ion CAN Calcium ammonium nitrate CaCl2 Calcium chloride CO2 Carbon dioxide CP Canopy position CRD Completely Randomized Design CTAB Cetyl Trimethyl Ammonium Bromide CuSO4.5H2O Copper sulphate pent hydrate cv Cultivar(s) dia Diameter DMRT Duncan’s Multiple Range Test DNA Deoxyribonucleic acid DPPH 2, 2-diphenyl-1-picrylhydrazyl EDTA Ethylene diamine tetra acetic acid et al. et alia EU FAO

European Union Food and Agriculture Organization

FC Folin–Ciocalteu FeSO4 Ferrous sulphate FeSO4 7H20 Ferrous sulphate FRAP Ferric-reducing antioxidant power assay

xiii

FW Fresh weight g Gram (s) GA3 Gibberellic acid GAE Gallic acid equivalents h Hour h° Hue angle H2SO4 Sulfuric acid ha Hectare HCl Hydrochloric acid HClO4 Perchloric acid HNO3 Nitric acid HPLC High performance liquid chromatography HPO3 Phosphoric acid i.e. Illud est IC Inhibition concentration IHS Institute of Horticultural Sciences K Potassium K2SO4 Potassium sulphate Kg Kilogram KNO3 Potassium nitrate kPa Kilo Pascal KPK Khyber Pakhtunkhaw L Liter L-1 Per litre LSD Least significant difference m Meter M Molar mg Mili gram Mg Magnesium mg g-1 Milli gram per gram min Minute MINFA Ministry of Food and Agriculture Mix. Mixture mL Milliliter mM Millimolar mm Millimeter mmol Millimole MMT Million metric ton NS Non-significant Na Sodium NAA Naphthalene acetic acid NaCl Sodium chloride Na2CO3 Sodium carbonateNaHCO3 Sodium bicarbonate NaOH Sodium hydroxide NH4 Ammonium NWFP North West Frontier Province NY New York P Probability PAL Phenylalanine ammonia-lyase

xiv

PBUH Peace be upon Him PG Polygalacturonase PGRs Plant growth regulators pH Hydrogen ion concentration in a solution PME Pectin methyl esterases ppm Parts per million PRTC Postharvest Research and Training Centre Put Putrescine RCBD Randomized complete block design RH Relative Humidity S.E. Standard error SOP Sulphate of potash Spd Spermidine Spm Spermine SSP Single super phosphate TA Total titratable acidity TEAC Trolox equivalent antioxidant capacity TPC Total phenolic contents TSS Total soluble solids UAF University of Agriculture, Faisalabad UK United Kingdom USA United States of America

1

ABSTRACT

Rind quality is indispensable for the external appearance and marketability of citrus fruit

especially for fresh consumption. Among many factors affecting citrus rind quality, tree age

is the most important one, but remains unexplored so far. This study was carried out during

2007-11 and comprised of two parts. The first part of study includes the experiments

exploring fruit quality in relation to different tree age, canopy position and fruit size. This

study also revealed the cell number, cell size, endogenous nutrients and fruit pectin

concentrations (rind and rag) in relation to tree age during fruit growth and development. A

comparison of fruit quality of different age groups (3 year, 6 year, 18 year, 35 year) showed

that fruit obtained from young trees (3-year-old) were poor in fruit quality such as having

more rough rind, rind thickness, rind mass (%) and less juice mass (%), TSS and acidity.

Moreover, fruit from young trees had lower rind macro nutrient concentrations (P, Ca) and

higher rind micro-nutrient concentrations (Mn and Fe). Fruit in internal canopy position had

significantly better fruit quality (smooth rind, less rind thickness, more juice contents)

whereas, those in external canopy position were better in biochemical fruit quality such as

(TSS), titratable acidity (TA), sugars and ascorbic acid (AA) concentrations. Large sized fruit

had more rind mass (%), rind thickness, and lower juice mass (%), TSS, TA (%) and TSS:TA

ratio during ambient (20±2°C and 60-65% RH) and cold storage (4±1°C and 75-80% RH)

conditions. From nutritional aspects, during fruit growth and development, fruit from six-

year-old trees were nutrient deficient in rind (N, P and K), rag and leaf (N) concentrations,

while 18-year-old trees were deficient in rag and leaf nutrient (P, K) concentrations. Pectin

analysis showed that fruit from 6-year-old trees were deficient in rind total pectin,

protopectin and rag protopectin concentrations, whereas fruit from 35-year-old trees were

higher in rind water soluble pectin (WSP), total pectin and fruit from 18-year-old trees were

higher in rind total pectin and rind and rag WSP. Anatomical studies of tissues from different

age groups demonstrated increased cell number with lower cell size in rind tissue of 6-year-

old trees in comparison with 18 and 35-year-old trees. Correlation analysis revealed that leaf

N contents correlated positively with cell size in fruit from 18 and 35-year-old trees. Cell size

was negatively correlated with rind P concentrations and positively correlated with leaf P

2

concentrations in fruit from all tree age groups. In second part of the study, the potential of

exogenous application of PGRs and nutrients were explored in improving fruit quality of

young 'Kinnow' orchards. The PGRs like gibberellic acid (GA3), 2, 4 dichlorophenoxy acetic

acid (2, 4-D), putrescine (Put) and spermine (Spm) were applied before and after colour

break to young (3-4 years old) 'Kinnow' mandarin trees and their influence on fruit quality

under ambient (20±2°C and 60-65% RH) and cold storage (4±1°C and 75-80% RH)

conditions was determined. Only 2, 4-D significantly improved TA (%) and TSS:TA ratio

during shelflife studies. GA3 and Put treated fruit exhibited maximum mass loss (%) during

shelf studies and cold storage respectively. GA3 10 mg L-1

and cytokinins (kinetin and

benzyladenine) 30 mg L-1

applications at fruit setting stage significantly improved juice (%)

and reduced rag (%). In nutritional experiments, sulphate of potash (SOP), single super

phosphate (SSP), urea, calcium ammonium nitrate (CAN), Wokozim and Isabion were

applied to improve fruit quality. Wokozim application reduced rind thickness and improved

reducing sugars of fruit, SSP improved juice contents, ascorbic acid (AA) and reduced rind

mass (%) although not significant than control; SOP improved AA concentrations in

'Kinnow' mandarin juice and CAN improved TSS and AA concentrations. In conclusion, tree

age exhibited significant influence on 'Kinnow' mandarin fruit quality as fruit from young

trees (3-years-old) showed inferior fruit quality. Moreover, rind of fruit from young trees had

lower macro- (N, P and Ca) and higher micro-nutrient (Mn and Fe) concentrations. In young

trees, macronutrients (P and Ca) and micronutrients (Cu, Mn, Fe and Zn) showed a negative

correlation with fruit rind thickness. Large sized fruit from all tree age groups exhibited poor

quality. Among PGRs, autumn application of 2, 4-D (10 ppm) to young 'Kinnow' mandarin

trees significantly improved TA and TSS:TA ratio. Spring application of cytokinin especially

kinetin, among nutrients SSP and CAN and among growth stimulator Wokozim (PGRs and

nutrient solution) positively affected fruit physical (rind thickness, total seed, juice, rind and

rag mass) and biochemical (TSS, reducing sugars, TA and AA) quality parameters (although

some seasonal variations also exist) thus showing their potential for improving fruit quality

of young 'Kinnow' mandarin orchards. An improvement in fruit quality of young orchards (3-

6 years) can help extend the productive window of 'Kinnow' mandarin orchards.

3

Chapter-1

GENERAL INTRODUCTION

'Kinnow' is a cross of Citrus nobilis Lour × Citrus deliciosa Tenora, the first generation

hybrid with the King (female) and Willow leaf (male) as parents. It was developed by H.B.

Frost at University of California, during 1925 (Rajput and Haribabu, 1985) and was

introduced from California to Pakistan (Experimental Fruit Garden of Punjab Agricultural

College and Research Institute Lyallpur now Faisalabad) during 1943-44 (Malik, 1992).

'Kinnow' mandarin monopolized the citrus industry of Pakistan due to its good yield and

quality and its better adaptation to the environmental conditions of Punjab (Ahmad et al.,

2006b). Pakistan currently ranks 11th

in mandarin production in international market (FAO,

2009). 'Kinnow' mandarin produces more than 60% in the country's citriculture (Altaf and

Khan, 2009). 'Kinnow' is mainly exported to Russia, Iran and the Gulf States. Pakistan with

an annual export volume of 250,000 tons is the sixth largest mandarin exporter in the world,

however, in terms of value earned, it ranked 11th

due to low unit price (US $ 249 tonne-1

) in

international market (FAO, 2009), owing mainly to poor fruit quality and yield. The external

presentation of fruit is also not according to international standards.

Fruit quality varies with cultural practices, pre and postharvest handling, storage and

distribution system and also with age of the tree. There are reports that citrus fruit from

young vigorously growing trees exhibit low total soluble solids (TSS) and acids and high

TSS to acid ratios (Hearn, 1993). Tree age affects acidity and TSS of 'Satsuma' mandarin

(Matsumato et al., 1972) and juice contents, TSS, acidity and ripeness index of oranges

(Frometa and Echazabal, 1988). Ozeker (2000) reported that 'Marsh' seedless grapefruit

harvested from 20-year-old trees produced bigger fruit with thinner rinds compared with 34-

year-old trees. Postharvest disorders were higher in pome fruit harvested from young trees

(Bramlage, 1993), while quality of apples was inferior from trees of old age (Smith, 2003).

Fruit from younger apple tree showed lowest storage potential with more firm fruit which

quickly lost flavour and quality during shelflife and storage, when compared with fruit from

middle and old aged trees (Tahir et al., 2007). So far, no studies have been published on the

relationship between tree age and fruit quality of 'Kinnow' mandarin.

4

In Pakistan, the life span of citrus (mainly 'Kinnow' mandarin) tree is declining due to many

biotic and abiotic factors (Ahmad et al., 2006b) and on an average seldom exceeds 25 years

(Ibrahim, 2004). In more than 40 percent cases, tree decline starts at the age of 10 years,

which is the prime age of production (Ahmad et al., 2006b). However, in other countries

economic life of citrus tree is 50-100 years depending on good management practices

(Chaudhary et al., 2004). In Pakistan, citrus tree takes 8-9 years for commercial fruit

production, whereas in other countries like Australia this period is only 6 years (Johnson,

2006). Citrus trees do not produce similar quality fruit throughout the life of an orchard and

need replantation after a certain age due to decline issue. This results in wastage of time and

resources of growers. On the other hand, young 'Kinnow' mandarin trees produce coarse

skinned fruit with less juice contents, and exporters are reluctant to take fruit from young

orchards (Malik, Personal Communication). Moreover, fruit from young trees also contains

less total soluble solids (Hearn, 1993), hence rejected by the processers. Due to these reasons

fruit from young trees are often sold in local market at very low price thus reducing the

income for citrus growers.

Therefore, it is essential to extend the productivity of 'Kinnow' mandarin orchards which is

possible in two ways: 1) manage decline related factors and extend life span beyond 15 years

and (2) improve fruit quality in young orchards, thereby increasing early returns. Lot of

research work has been done with regard to the first possibility by tree health management

(Batool et al., 2007; Chung and Brlansky, 2005), whereas limited information is available for

improving fruit quality of young orchards.

Main obstacle in quality fruit production from young orchards is their excessive vegetative

growth and poor fruit quality. Plant growth regulators (PGR) have been used to manipulate

vegetative and reproductive growth to modify fruit set and fruit growth and to improve fruit

quality (reducing rind thickness, rind contents and improving juice contents) in mature citrus

trees (Fidelibus et al., 2002a; Kaseem et al., 2011; Pozo et al., 2000; Saleem et al., 2008c).

However, to the best of our knowledge their application in young orchards has not been

reported elsewhere.

The role of mineral nutrients in different parts of citrus fruit and their relation with fruit

quality is well documented in literature (Mattos et al., 2003; Paramasivam et al., 2000; Raza

5

et al., 1999; Xiao et al., 2007). However, no study has been reported to determine the rind

nutrient concentrations from trees of varying age and to establish a correlation between rind

nutrient concentrations and rind quality.

Fruit position in the canopy (Barry et al., 2003; 2004ab; Khan et al., 2009) and fruit size

(Barry et al., 2004a; Ketsa, 1988) has been reported to affect fruit quality. Low temperature

storage and long-distance fruit transport allow an extended market period for more regulated

fruit supply. Currently, no information is available on the effect of tree age, fruit position and

fruit size on 'Kinnow' mandarin fruit quality under ambient (20±2°C) as well as in cold

storage (4±1°C) conditions.

It can be summarized that there is a need for comprehensive research to determine the

possible reasons for variation in fruit quality with tree age and its improvement. This thesis

reports the relationships between tree age, storage potential and fruit quality. The objectives

of this study were:

To explain the physiological basis for poor fruit quality in young orchards.

To develop fruit quality and storage potential profile of fruit from trees of

different age groups.

To explore potential of exogenous application of growth regulators and nutrients

in improving fruit quality of young orchards.

6

Chapter-2

GENERAL REVIEW OF LITERATURE

2.1 Introduction

The loose skinned oranges are generally known as mandarins. Mandarin and tangerines are

names used more or less interchangeably to designate the whole group. These two groups are

differentiated only on the basis of colour of the fruit; the name tangerine is strictly used for

those varieties which are producing deep orange or scarlet fruits. Niaz (2004) described six

distinct groups of mandarin i.e., King, Satsuma, Mandarin, Tangerine, Mandarin-lime and

Mitis groups. The mandarin is native to south-eastern Asia and the Philippines (Mortan,

1987). The mandarins are also known by their indigenous names in different countries. In

Philippines, all mandarin oranges are called Naranjita and in Spain these are named as

Mandarina.

2.2 Origin and Distribution

The 'Kinnow' is a hybrid of two citrus cultivars 'King' (Citrus nobilis) × 'Willow Leaf' (Citrus

deliciosa) developed by H.B. Frost at the Citrus Research Centre of the University of

California, Riverside, USA in 1925 and after evaluation it was named and released as a new

variety for commercial cultivation in 1935 (Rajput and Haribabu, 1985). In 1940, 'Kinnow'

was introduced to Punjab Agriculture College and Research Institute, Faisalabad (former

Lyallpur), Pakistan (Malik, 1992). In India, J.C. Bakhshi introduced this variety to the Punjab

Agricultural University, Regional Fruit Research Station, Abohar in 1954 (Wikipedia

contributors, 2012). 'Kinnow' has been distributed widely and is being grown commercially

in Punjab province of both Pakistan and India and to some extent in California and Arizona

(Reuther et al., 1967). Its tree has vigorous and upright growth habit, with great heat

tolerance, cold-resistance and having a strong tendency of alternate bearing with large crop

of smaller fruit followed by very small crop of larger fruit (Reuther et al., 1967). The fruit is

medium sized, slightly oblate with a smooth orange rind that does not peel especially well for

a mandarin. Fruit contains 9 to 10 segments, firm, separating fairly easily with axis solid to

semi-hollow. The flesh is yellowish-orange, seedy, and very juicy having a rich distinctive

flavour. 'Kinnow' is mid-season in maturity and holds well on the tree (Reuther et al., 1967).

7

'Kinnow' female parent King (Citrus nobilis) is now known as a natural tangor, hybrid of

mandarin (Citrus reticulata) and orange (Citrus sinensis). The most important mandarin in

subcontinent is the Santra (Niaz et al., 2004) or Sangtara which also been found in the region

of Lahore, Pakistan during 15th

century (Sabir, 2012). Further, sangtara has been mentioned

in the famous book ‘Ain-e-Akbari’ by Mughal Emperor, Akbar Khan. After this the fruit was

popularly called as ‘Shahi Sangtara’ or King Orange. Mughal Emperor, Humayun Khan

praised this fruit in the following words. “Indeed there is no tasty fruit than the sangtareh”

(Sabir, 2012). In 1880 six fruit of the 'King' mandarin were sent by John A. Bingham, a

United States Minister to Japan from South Vietnam (Saigon) to Dr. H.S. Magee at

Riverside, California. The latter he sent two seedlings to J.C. Stovin of Winter Park, Florida

in 1882 (Mortan, 1987). The most distinctive feature of King mandarin is its very high heat

requirement for the attainment of horticultural maturity and good quality, for which reason it

is the late ripening variety of the mandarins (Reuther et al., 1967).

The parentage and mode of origin of second parent (Willow leaf) of 'Kinnow' are not known

but it seems likely that it arose as a chance seedling from a mandarin variety or form of

Chinese origin. After careful review of the literature, Chapot (1962) concluded that it

appeared in Italy between 1810 and 1818. It was brought to the United States by the Italian

ambassador at New Orleans and planted in the consulate grounds there sometime between

1840 and 1850, apparently being the first mandarin to reach this country. Not long thereafter,

it was taken to Florida and thence probably to California and elsewhere (Reuther et al.,

1967).

The other related hybrids of the same parentage (King × Willow leaf) are 'Encore', 'Honey'

(not the Murcott of Florida) and 'Wilking' (Reuther et al., 1967).

2.3 Citrus Industry of the World

Citrus is the most widely produced fruit of the world and is grown in more than 80 countries

(Chang, 1992).

8

2.3.1 World Mandarin and Tangerine Production

Pakistan is on eighth position in terms of mandarin quantity produced and production value

(FAO, 2010). Approximately 95% world 'Kinnow' is produced in Pakistan.

Source: FAO (2010)

Fig.2.1 World mandarin and tangerine production

2.3.2 World Mandarin and Tangerine Export

In international mandarin and tangerine trade, Pakistan ranks sixth in terms of quantity

exported and on eleventh in terms of value earned from export (FAO, 2009).

Source: FAO (2009)

Fig.2.2 World mandarin and tangerine export

2500109

421962

277339

212117

196843

194159

151886

141489

133335

116800

Production ($1000) China

Spain

Brazil

Turkey

Egypt

Japan

Republic of Korea

Pakistan

USA

Morocco

10121000

1708200

1122730

858699

786000

614871

572780 572780

539770

472834

Production (MT)

China

Spain

Brazil

Turkey

Egypt

Japan

Republic of Korea

Pakistan

USA

Morocco

52%

13%

9%

8%

6% 4%

3% 2% 2% 1%

Value (1000 $)

Spain

China

EU(27)ex.int

Turkey

Morocco

Netherlands

Argentina

Italy

South Africa

Pakistan

39%

21% 10%

7%

7%

5% 3%

3% 3%

2%

Quantity (tonnes)

Spain

China

Turkey

Morocco

EU(27)ex.int

Pakistan

South Africa

Argentina

Netherlands

Italy

9

2.4 Citrus Industry of Pakistan

In Pakistan, citrus ('Kinnow') is the top ranking fruit crop in terms of area and production

followed by mango, apple and guava (Anonymous, 2009). About 95% of citrus area is

located in Punjab (Anonymous, 2009), especially Sargodha district.

Source: Anonymous (2009)

Fig 2.3 Citrus area and production in Pakistan

2.4.1 Mandarin export from Pakistan

Currently 'Kinnow' is being exported to different countries of the world, major markets

include Russia, Afghanistan, Iran, UAE and Saudi Arabia etc (Anonymous, 2009).

Table 2.1 Mandarin export from Pakistan during 2008-09

Countries Quantity (Tonnes) Value (1000 US $) Unit price ($ tonne-1

)

Russian Federation 36263.68 9186.44 253.32

Afghanistan 34127.21 11939.68 349.86

Iran 33493.78 6869.34 205.09

UAE 25722.1 4145.81 161.18

Saudi Arabia 10934.28 1944.44 177.83

Ukraine 8103.54 1809.60 223.31

Kuwait 5763.24 842.60 146.20

199940

170166 36056

113029

15358 62238

Area (Hectare)

Citrus

Mango

Banana

Apple

Grapes

Guava

2132276

1727932

157319

441062

76095

512295

Production (Tonnes)

Citrus

Mango

Banana

Apple

Grapes

Guava

10

2.5 Citrus Fruit Anatomy

Citrus fruit are classified as ‘hesperidum’ a special type of berry which develops through

growth and development of a single ovary consisting of 8-16 carpels clustered around and

joined at the floral axis and surrounded by tough leathery rind (Schneider, 1968). The

endocarp (edible portion) of the fruit comprised of segments (carpels) in which juice vesicles

and seeds grow. The rind is divided into exocarp or flavedo (outer coloured part) and

mesocarp or albedo (inner white part) (Ladaniya, 2008). The flavedo comprises of the

epicarp, covered by a protective skin or cuticle, the hypodermis, the outer mesocarp and oil

glands.

Fig. 2.4 Transverse section of citrus fruit (Ladaniya, 2008)

2.6 Fruit Growth and Development

Iglesias et al. (2007) described the three stages of citrus fruit growth and development

including cell division, cell elongation and maturation. The lengths of these stages vary

depending on citrus variety and location. The length of cell division (stage-I) is from 4 to 9

weeks. Lowell et al. (1989) reported that during stage of cell division pericarp (rind) grows

very quickly.

During stage-II, fruit growth and development is mainly due to cell enlargement and

differentiation. In this stage cell division only persists in flavedo and the tips of the juice

sacs. It has been reported that during the stage-II, rind thickness, reduced while pulp volume

11

increased continuously due to water accumulation and cell enlargement (Iglesias et al.,

2007).

During the third stage (i.e. maturation) the rate of growth is much lower than in stage II. Fruit

ripening starts during this stage with disappearance of chlorophyll pigments and subsequent

development of carotenoids pigments.

2.7 Fruit Quality

Quality is very important for consumption and marketability of produce and it varies from

person to person. In citrus external fruit quality parameters include colour, size, rind

smoothness and blemishes whereas internal fruit quality includes rind thickness, juice

concentrations, ascorbic acid, acidity, TSS, TSS:TA.

2.7.1 Factors affecting fruit quality

In the past, studies have been conducted to investigate the factors affecting fruit quality in

various fruit crops. Selected research work is being summarized under different factors with

regards to their effect on physical and biochemical fruit quality as under:

2.7.1.1 Tree age

Tree age has a pronounced effect on fruit physical and biochemical quality parameters.

2.7.1.1.1 Physical fruit quality

Young trees tend to be more vigorous than old mature trees. Due to vigorous growth young

trees have poor rind colour development compared with older less vigorous trees (Krajewski,

1997). Fruit from mature 'Navel' orange trees had severe outbreaks of the rind disorder

albedo breakdown (crease) as compared to fruit from young trees (Storey and Treeby, 2002).

Fruit obtained from 5-10 year old Prunus salicina, trees yielded heavier fruits with larger

transversal diameter as compared to fruit obtained from 20-30 years old trees (DongHui et

al., 2005).

Ozeker (2000) reported that 'Marsh seedless' grapefruit harvested from younger trees (20

years old) were bigger and heavier in size with thinner rind than old trees (34 years old). Age

of the tree and cultivar influence the juice concentrations of oranges (Frometa and Echazabal,

1988). In avocado, grey pulp problem was found higher in young as compared to old trees

12

(Snijder et al., 2002).

2.7.1.1.2 Biochemical fruit quality

In Prunus salicina, 5-10 year old trees had higher ascorbic acid concentrations than 20-30

year old trees however taste, acidity and soluble solid contents did not significantly vary with

the tree age (DongHui et al., 2005). Tree age also affected storability of fruit as Tahir et al.

(2007) reported that apple fruit from young trees (younger than 6 years) had a lower

resistance to bruising and Pezicula malicorticis decay, while fruit from trees older than 20

years were prone to reduced quality and storability. Asrey et al. (2007) reported that fruit

from upper canopy of 15 year old guava trees had higher TSS (11.85%), total sugars (7.50%)

and lowest acidity (0.28%).

2.7.1.2 Fruit size

Fruit size is one of the key quality factors in the fruit trade. Increased fruit size is desirable

for mandarin-type fruits as small size fruit fetchs low prices on the fresh market thereby

causing considerable economic losses (Guardiola et al., 1988). Ghaffar (1991) studied that

size of the fruit is very important in quality evaluation, particularly when fruit of most

cultivars are immature and or too small. It is also sometimes used as a criterion for spot

picking.

2.7.1.2.1 Physical fruit quality

Fruit size influences skin colour in peaches. In small sized fruit of peach cv 'Hashiba-

hakuho,' skin colour on the cheeks (yellow) was dark and in large sized fruit the colour at the

top (reddish) was dull and dark yellowish in Shimizu-'Hakuto' and 'Hakurei' respectively,

compared to fruit of other sizes (Okamoto et al., 2003). Mass loss and fruit firmness during

storage is very important and influenced by fruit size. As smaller banana fruit showed

significantly higher mass loss and lower fruit firmness when compared with larger banana

fruit (Ahmad et al., 2006a). However smaller 'Kinnow' (Malik et al., 2007) and apple (De

Salvador et al., 2006) fruit were firmer than medium and large sized fruit. In smaller

grapefruit, mass loss percentage was significantly higher than that of larger fruit (Paily et al.,

2004). Small sized Kiwifruit (Actinidia chinensis) of 'Hort 16A' lose firmness earlier than

medium and large fruit however, small AU Golden Sunshine fruit had greater firmness in the

later weeks of cold storage when compared with medium and large fruit (Spiers et al., 2011).

13

Juice (%) also varies with fruit size. Paily et al. (2004) reported that percentage of fruit juice

in grapefruit with large diameter was always higher than in small diameter fruit. Sandhu

(1992) reported that 'Kinnow' mandarin fruit from extra-large fruit size had lower juice (%).

2.7.1.2.2 Biochemical fruit quality

Fruit size had a marked influence on biochemical fruit quality in many fruits. Large sized

grape barries have less sugar concentrations and inadequate aroma (Hirano et al., 1996).

Large sized peaches, which were produced on heavily fertilized trees, have very poor flavour

and aroma than in trees fertilized normally (Jia et al., 1999). In small and medium sized

peach of cultivars 'Hakuho', 'Shimizu-hakuto' and 'Hakurei' sucrose and fructose

concentrations were higher while, malic and citric acid concentrations were more in large

sized fruits of those cultivars (Okamoto et al., 2003). Small citrus fruit had higher TSS than

large sized fruits (Hardy and Sanderson, 2010). Ketsa (1988) described that ascorbic acid

was not related to fruit size in tangerine. TSS and acidity decreased with increasing fruit size

in tangerine (Ketsa, 1988), 'Satsuma' mandarin (Kihare et al., 1982) and 'Kinnow' mandarin

(Malik et al., 2007). Smaller bananas showed significantly higher TSS when compared with

larger bananas (Ahmad et al., 2006a). Sandhu (1992) reported that 'Kinnow' mandarin fruit

from extra-large fruit size had lower Brix and acidity as compared to other fruit sizes

however ascorbic acid concentrations did not vary with fruit size. Miller (1990) reported a

negative correlation between fruit size and Brix of 'Valencia' orange. In small sized Kiwifruit

(Actinidia chinensis) of 'Hort 16A' variety TSS:TA, internal colour and % dry matter were

not affected by fruit size contrarily, small 'AU Golden Sunshine' fruit had lower TSS, less

internal colour development, and greater firmness in the later weeks of cold storage when

compared with medium and large fruit (Spiers et al., 2011). However, some studies reported

that fruit size had no significant effect on biochemical fruit quality of tangerine

(Jungsakulrujirek and Noomhorm, 1998) after harvest and grapefruit (Paily et al., 2004)

during storage.

As evident from above literature review fruit quality varies with tree age and fruit size.

However, their combined effect on fruit quality has not been reported in literature to the best

of our knowledge and yet to be explored.

14

2.7.1.3 Canopy position

Large fruit size, thin rind, high total soluble solids and solids to acid ratios are desirable

quality attributes in citrus fruit which are dependent upon interception of light within the

canopy. Considerable review on influence of canopy position on fruit physical and

biochemical characteristics is presented as under:

2.7.1.3.1 Physical fruit quality

Fruit canopy position influences fruit quality probably due to light penetration. Orlando

tangelo fruit from bottom and middle canopy positions were firmer, juicy and had greatest

Hue (H) value when compared with fruit from top-canopy position (Morales et al., 2000).

Inner canopy fruit during the immature stage (January-March/April) had less green colour

(lower hue angle) compared to outer canopy fruit but after colour break (between March and

April) the outside fruit developed more orange colour (lower hue angle) in contrast with

inside fruit moreover, fruit from inner canopy position were lighter and smaller (diameter and

length) compared to outer canopy fruit (Cronje et al., 2011). Khan et al. (2009) reported that

'Kinnow' fruit from inside canopy were heavier (20%) and larger in volume (22.3%), had

more rind and rag mass and less juice (%) as compared with fruit harvested from top of the

tree canopy, however canopy position had no significant effect on rind thickness of fruit.

Jawanda et al. (1973) also reported that fruit from the inner part of the canopy were heavier

with less juice and more rind percentage than from other sides of tree canopy. Higher fruit

mass, rind thickness, rind fresh and dry mass, was found in 'Kinnow' mandarin, 'Red Blush'

grape fruit 'Valencia' orange and 'Lisbon' lemon form internal canopies than from external

canopies (Fallahi and Moon Jr., 1989). Grapefruit harvested from the shaded positions were

lighter with less juice content than fruit from sun exposed canopy positions (Syvertsen and

Albrigo, 1980b). Lewallen and Marini (2003) reported that peach fruit from the exterior

canopy were larger, had more darker and redder surface than fruit harvested from interior

canopy positions.

2.7.1.3.2 Biochemical fruit quality

Fruit orientation in the canopy has been reported to have strong influence on biochemical

fruit quality. Fruit located in the top-canopy position of 'Orlando' tangelo had highest TSS

and TSS:TA ratio while, TA contents had little variations due to canopy position (Morales et

15

al., 2000). Similarly Barry et al. (2003) also reported that canopy position had significant

effect on TSS contents and non significant effect on TA of 'Valencia' orange fruit. Jawanda et

al. (1973) reported that acidity and TSS were higher in the upper sides of citrus trees. Orange

fruit harvested from upper canopy positions had high contents of reducing sugars (Uchida et

al., 1985), whilst no effect of fruit position in the canopy was observed on total sugar content

in 'Satsuma' mandarin (Datio and Tominaga, 1981). Khan et al. (2009) reported that 'Kinnow'

mandarin fruit harvested from top of the tree and from outer periphery had significantly

higher in SSC and SSC:TA ratio compared to fruit harvested from inside and lower side of

the tree canopy. In the outside canopy positions of 'Ruby' grapefruit, SSC were higher and

acidity was lower in other canopy positions (Syvertsen and Albrigo, 1980b). In the litchi

fruit, lower brix:acid ratio was observed in lower canopy position than from fruit harvested at

higher canopy positions (Tyas et al., 1998). Asery et al., (2007) found that middle canopy

fruit from 15-year-old trees had higher AA concentrations and lower acidity contents. They

also reported that upper canopy fruit obtained from 15-year-old trees had higher total sugar

contents.

Sectorial position of fruit in the canopy also influences biochemical fruit quality. Citrus fruit

harvested from the southern top canopy position had higher TSS and juice contents than fruit

from other canopy positions (Izumi et al., 1990). 'Tarocco' orange harvested from the

external southern side of the tree had higher TSS and lower acid levels, resulting in higher

TSS:acid ratio and improved taste (Agabbio et al., 1999). 'Torocco' orange fruit from the

external southern side of the tree had higher SSC and lower acid contents in comparison to

fruit from interior and northern parts of the tree canopy (Agabbio et al., 1999). In grapefruit,

southern top canopy positions had fruit with more TSS contents and less TA and higher

TSS:TA ratios due to highest temperature and lowest water potential (Syvertsen and Albrigo,

1980b).

Above literature review exhibited that fruit position in a canopy had significant influence on

fruit quality. Tree canopy increases with increase in tree age so is the difference in fruit

quality. No research work has been reported on the combined effect of tree age and fruit

canopy position on fruit quality in Citrus especially in cv 'Kinnow' mandarin.

16

2.7.1.4 Plant growth regulators

Endogenous PGR production is dependent upon phenological stages of the plant and their

concentration and types vary during fruit growth and development (Guardiola et al., 1993;

Kakkar and Rai, 1993; Malik and Singh, 2004). Auxins, GA3 and cytokonin concentrations

were higher in young citrus fruitlets, whereas the concentration of growth inhibitors like

abscisic acids was higher during fruit maturation and senescence (Bain, 1958). In mandarin,

polyamine levels changed with fruit growth curve and its higher amounts were found during

cell division stage (Nathan et al., 1984). Putrescine (Put) concentration in rind and rag of

'Navel' oranges was found higher during ripening (Tassoni et al., 2004). In young orange

fruit, activity of GA3 was relatively high whereas auxin was detected in small amounts

(Goren and Goldschmidt, 1970). In Citrus unshu, endogenous levels of benzyladenine (BA)

and GA3 fell to a much lower value with in few days after flower opening, and it coincided

with a marked reduction in the fruit response to these hormones (Guardiola et al., 1993). In

citrus, cytokinins levels were found higher in developing ovaries at anthesis and were

involved in cell division whereas auxin was found higher during cell enlargement phase and

played important role in maintenance of cell size (Iglesias et al., 2007).

Endogenous PGRs concentrations vary with age of the plant and type of plant part. Juvenile

'Pickstone Valencia' orange plants had higher cytokinin concentrations in fibrous root tips

than adult plants, however during bud growth all cytokinin levels decreased in juvenile buds,

whereas in adult plants polar cytokinin decreased slightly but non polar increased (Hendry et

al., 1982). In peach (Prunus persica) trees total polyamine concentrations increased with

increasing tree age whereas in pine (Pinus radiata) trees total polyamine concentrations

decreased with tree age (Fraga et al., 2004). Putrescine (Put) was found higher in juvenile

tissues of hazel nut (Corylus avellana L.) (Rey et al., 1994) while, opposite was reported in

grapevine (Vitis vinifera L. cv. Pinot noir) (Heloir et al., 1989). Paschalidis and Roubelakis-

Angelakis (2005) reported that in youngest tissues and shoot apex of tobacco plant

spermidine (Spd) and spermine (Spm) concentrations were found higher. Fernandez (2003)

reported that endogenous Gibberellic acid types vary with age of the Pinus radiate, trees with

juvenile plants having higher GA7 and GA9 and lower GA4 as compared to mature trees,

whereas GA3 and GA20 did not vary with tree age. Similarly Davenport et al. (2000) reported

that endogenous GA3 levels did not vary with age of mango stem whereas other GA types

17

declined with stem maturity. Husen and Pal (2006) reported that endogenous auxin

concentrations decreased with age of the donor plant as cutting from mature plants required

higher amount of exogenous auxins for rooting. Endogenous PGRs varies with tree age and

fruit development stages and their exogenous application at different growth and

developmental stages is well documented in mature citrus trees (Fidelibus et al., 2002a).

However, their application in young 'Kinnow' mandarin orchards have not been reported and

yet to be explored.

Plant growth regulators (PGRs) have been used to improve citrus fruit quality, mainly

affecting on fruit size and peel properties. PGRs are routinely used by many citrus growers in

Spain, California, Australia, South Africa and Israel to increase crop productivity. PGRs may

be used to improve fruit set, increase fruit size, improve on tree storage and reduce hand-

suckering by controlling trunk sprout growth in young trees. Several studies have shown that

PGRs play an important role in manipulating vegetative and reproductive growth and

improving fruit quality (Malik and Singh, 2004; Saleem et al., 2008c; Saleem et al., 2008a).

2.7.1.4.1 Physical fruit quality

Cytokinins are substituted adenine compounds, naturally occurring in plants, which promote

cell division in tissue systems (Salisbury and Ross, 1991). Cytokinins are responsible for

delaying fruit colour development in citrus. Benzyladenine (BA) significantly delayed

chlorophyll degradation in 'Feizixiao' mandarin and inhibited anthocyonin biosynthesis

(Wang et al., 2005). BA delays fruit abscission and increases the amount of re-greening in

'Valencia' oranges (Cooper and Henry, 1968). Eilati et al. (1969) reported that BA did not

influence the carotenoid accumulation during the onset of fruit maturation. In contrast

Gracia-Luis et al. (1986) demonstrated that cytokinin reduced the carotenoid accumulation in

citrus rind and could be used as maturation retardant. The Kinetin and Benzyl adenine

treatments in 8 year-old 'Kinnow' mandarin tree reduced chlorophyll degradation (Nagar,

1993).

Cytokinin efficacy depends upon its stage of application. BA when applied to adult (20 year-

old) Citrus unshiu at flower opening stage increased ovary growth but the sensitivity of the

fruitlet to growth regulator decreased with age of fruitlet and no growth increment was

observed when BA was applied 11 days after anthesis moreover BA application increased

18

cell division in pericarp (rind) of the fruit resulting in more rind thickness (Guardiola et al.,

1993). Cytokinins are being used in citrus fruits to improve their fruit quality. However, their

applications to improve fruit quality in young citrus tree is lacking and yet to be explored.

Effectiveness of Gibberellic acid (GA3) varies with its application time in different cultivars

of citrus. Foliar application of GA3 during colour break delayed fruit ageing in cultivar

'Satsuma' mandarin (Kuraoka et al., 1977) and 'Navel' orange (Riehl et al., 1996). Ladaniya

(1997) applied GA3 to 'Nagpur' mandarin at colour break and reported that GA3 improved

fruit firmness, delayed colour development and reduced fruit mass loss during storage. Davis

et al. (1999) determined the optimal time for GA3 application in 14-year-old 'Hamlin' sweet

orange and reported that fruit treated with GA3 at colour break had statistically greater juice

yield than fruit from non sprayed trees. Fidelibus et al. (2002b) reported that application of

GA3 at about colour break increased juice weight of sweet orange, reduced rind thickness and

markedly improved the strength of the rind in shear and tension. Fidelibus et al. (2002a)

applied GA3 on mature ‘Hamlin', 'Pineapple', and 'Valencia' sweet oranges from September to

December and reported that earliest applications were most effective in maintaining rind

puncture resistance compared with control fruit, while the later application dates resulted in

the most green peel colour at harvest. Pre-harvest treatment with GA3 application delayed

rind color development of on-tree-stored citrus fruit (Ferguson et al., 1982). Gracia-Luis et

al. (1992) reported that late application of GA3 had no effect on fruit growth and quality, but

early GA3 applications reduced rind thickness at maturation.

In 'Ponkan' mandarin, GA3 application at 200 ppm one month after anthesis reduced fruit size

(Moreira et al., 1996) while, its application to young fruitlets significantly increased fruit size

in grapefruit (Berhow, 2000) and fruit mass and diameter in 'Balady' mandarin (El-Hammady

et al., 2000). In citrus, GA3 applications retard rind colour development and rind aging. GA3

application before and after harvest significantly slowed the rate of rind colour change and

the rate of rind softening of 'Clementine ' mandarin (El-Otmani et al., 1990) and reduced the

incidence of rind disorders in 'Ponkan' mandarin (Tominaga et al., 1998) and 'Balady'

mandarin (El-Hammady et al., 2000). No effect of GA3 was found on final fruit size of

'Satsuma' mandarin (Guardiola et al., 1993), fruit weight of 'Navel' orange (Schafer et al.,

2000) and on length, diameter and fresh fruit mass of 'Pera' orange (Almeida et al., 2004).

19

Delayed softening by GA3 has been shown for on-tree-stored navel orange (Coggins, 1969)

and grapefruit (Ferguson et al., 1982). GA3 applications at various time during fruit growth

and development is widely reported in literature. However, its application at various time in

young citrus tree is not reported and yet to be studied.

Among auxins, 2, 4-D is widely used in mature citrus trees as it has an injurious effect on

young trees. Delayed rind colour development of on-tree-stored grapefruit by pre-harvest

treatment with 2, 4-D was reported by Ferguson et al., (1982). Delayed softening by 2, 4-D

has been shown for on-tree-stored 'Navel' orange (Coggins, 1969) and for cold-stored

grapefruit (Ferguson et al., 1982). Application of 2,4-D delayed ripening and colour

development (Lodh et al., 1963) and retained greenness of buttons (Sonkar et al., 1999) in

mandarins. The styles persisted on treated fruit until late into fruit development (Krezdom,

1969; Verreynne and Mupambi, 2010). Stewart and Klotz (1947) reported that the

application of 225 mg L-1

2,4-D on 'Valencia' and 'Washington Navel' oranges resulted in a

coarse rind due to enlarged oil glands. The fruit were also cylindrical in shape and the

'Valencia' oranges developed a small secondary fruit. Similar effects were noted on grapefruit

(Stewart and Parker, 1947). When applied as a fruit growth enhancer, 2,4-D treated fruit were

greener and more elongated compared to the control fruit (Stewart et al., 1951). It increased

the percentage of the rind as well as the rag and reduced the juice content of the treated fruit

(Stewart and Parker, 1947).

Some reports are also found whereby 2,4-D reduced juice percentage but had no effect on

rind thickness (Saavedra 2006; Verreynne and Mupambi, 2008). In 'Washington Navel'

orange there was the development of small rudimentary seeds in treated fruit (Stewart and

Klotz, 1947). When applied at full bloom to act as a growth enhancer, 2, 4-D also increased

the percentage of the rind as well as the rag and reduced the juice percentage in 'Washington

Navel' fruit (Stewart et al., 1951). Above review revealed that 2, 4-D applications are also

well documented in mature citrus trees. As in young citrus trees vigorous growth is

responsible for poor fruit quality, so the potential of 2, 4-D to suppress vigorous growth and

improve fruit quality is likely to be explored.

Polyamines (PAs) are polycationic compounds of low molecular weight that are present in all

of living organisms and are implicated in various biological processes including plant

20

growth, development, flowering, fruit ripening senescence and stress response (Malik and

Singh, 2004). PAs include putrescine (Put), spermidine (Spd) and spermine (Spm).

In 15 year old peach trees, preharvest PAs treatment increased fruit firmness during ripening

period (Bregoli et al., 2002). Another effect of PAs infiltration is to ameliorate chlorophyll

breakdown in several plant organs, including fruit, such as in lemon and apricot, since Put

treatment delayed the colour change during storage, which is the indicator of reduced

senescence rate (Valero et al., 1998). Exogenous application of PAs retarded chlorophyll loss

in muskmelon by reducing the hydrolytic activities acting on chloroplast thylakoid

membrane (Lester, 2000). Zheng and Zhang, (2004) reported that Spm significantly reduced

weight loss (%) and fruit decay in 'Ponkan' mandarin after three months storage period.

Exogenous PAs applications improved rind colour and rind smoothness of fruit harvested

from 12-15 year-old sweet orange trees (Saleem et al., 2008a). Author further stated that PAs

(Spd) treatment reduced rind (%), and improved juice (%) and rag (%) in sweet orange fruit.

Little information is available on preharvest PAs application on fruit quality in citrus.

2.7.1.4.2 Biochemical fruit quality

Preharvest applications of Cytokinins significantly influenced biochemical fruit quality in

apple. As Koukourikou-petridou et al. (2007) reported that kinetin when applied at petal fall

stage increased total sugars and TSS of 'Red Chief Delicious' apple. Similarly Al-Absi (2009)

found that BA at 200 ppm applied to apple at fruitlet diameter of about 10 mm significantly

increased TSS contents in comparison with control.

Pre-harvest sprays of GA3 increased juice brix (a measure of TSS) of grapefruit at harvest

(El-Zeftawi, 1980). In citrus, GA3 efficacy varies from cultivar to cultivar. Pre-harvest

application of GA3 had no effect on acid and soluble solids content of 'Navel' orange at

harvest (Coggins, 1969), but when applied to cultivar 'Valencia' orange, it increased solids

and acid in juice (Embleton et al., 1973). On cultivar 'Satsuma' mandarin, GA3 had no effect

on juice soluble solids and acids, but depressed peel sugar content (Kuraoka et al., 1977).

GA3 application to 'Clementine' mandarin increased maturity index during off year while on

year it increased TA and ascorbic acid concentrations of fruits juice (El-Otmani, 2004). GA3

applications also influence fruit quality during storage. Mature green fruit of the 'Mahaley'

orange when treated with GA3 gave superior fruit quality throughout the storage period at 4

21

or 7oC when compared to control (Al-Doori et al., 1990). Pre-harvest GA3 applications

significantly reduced fruit decay during storage (Coggins, 1969).

Variable effects of growth regulators on citrus fruit quality were reported in the literature.

TSS was reduced while titratable acid content was improved in juice when 'Washington

Navel' fruit was treated with 2, 4-D (Stewart et al., 1951). Pre-harvest sprays of 2, 4-D

increased juice °Brix (a measure of TSS) of grapefruit at harvest (El-Zeftawi, 1980). There

was also a slight decrease in titratable acids and an increase in the soluble solids to acid ratio

of 'Washington Navel' orange by the application of 2, 4-D (Stewart and Klotz, 1947). 2, 4-D

treatment lowered the ABA concentration and prolonged the storage life of 'Valencia'

oranges (Liu and Xu, 1998).

Polyamines application significantly improved sugar and acid contents of various fruits.

Zheng and Zhang (2004) applied PAs to 'Ponkan' mandarin and reported that all PAs

treatments increased TSS contents in comparison with control but the results were not

statistically significant. Exogenous applications of Put (5x10-5

M) increased TSS contents of

apple (Costa and Bagni, 1983). In litchi, Put applications decreased TSS and TSS:TA ratio

(Mitra and Sanyal, 1990). Purwoko et al., (1998) found no significant effect of PAs on TSS

and acidity contents of mango, whereas in papya, spermine reduced TSS contents. Bregoli et

al. (2002) treated peach trees with various levels of PAs, 19 days before harvest and reported

that Spd application significantly reduced TSS of fruit. In 12-15-years-old sweet orange trees

PAs application improved TSS, total and non reducing sugars (Saleem et al., 2008a).

Above review of literature revealed that PGRs (cytokinins, GA3, 2,4-D, and polyamines) are

widely tested in mature fruit trees however, studies on their application in young 'Kinnow'

mandarin tree is lacking and yet to be explored.

2.7.1.5 Mineral nutrition

Endogenous nutritional status of citrus tree influences citrus fruit yield and fruit quality

(Moss, 1972; Storey and Treeby, 2000). Rind phosphorous and rag potassium concentrations

continuously increased as long as 'Kinnow' fruit stayed on the tree (Raza et al., 1999).

Contrarily, Storey and Treeby (2000) reported that in whole fruit of 'Navel' orange P and K

decreased during fruit growth and development, while Ca increased during stage-I and then

22

decreased during stage-II and stage-III. Sheng et al. (2009) determined nutrient

concentrations in leaf, rind and rag of 'Newhall' and 'Skagg's Bonanza' navel oranges and

reported that nutrient status in each tissue varied differently during fruit growth and

development. Concentration of macronutrients decreased during fruit enlargement, while

micronutrient concentrations first increased and then decreased in four orange varieties

viz.'Valencia', 'Parson Brown', 'Hamlin' and 'Sunbrust' during fruit growth and development

(Paramasivam et al. 2000).

Tree age also effects endogenous nutrient status in fruit. Fruit from mature 'Navel' orange

trees, with a history of frequent and severe outbreaks of the rind disorder albedo breakdown

(crease) had higher albedo K/Ca and Mg/Ca ratios during stage-I (cell division stage) of fruit

development, compared to fruit from young trees (Storey and Treeby, 2002). In the fruit

from the older avocado trees N concentrations were lower, while calcium concentrations

were higher (Snijder et al., 2002).

Endogenous nutrients status of fruit fluctuates with the position of fruit in canopy. In outer

tree canopy fruit flavedo of 'Nules Clementine' mandarin accumulated higher concentrations

of Ca and Mg while, fruit obtained from inner tree canopy accumulated higher levels of K

(Cronje et al., 2011). Asrey et al., (2007) reported that fruit Fe concentrations were higher in

young guava trees. They also found that middle canopy fruit from 15-year-old trees were

richer in Cu and Mn concentrations while magnesium (Mg) was found higher in fruit of 20-

year-old trees. Light exposed kiwifruit (Actinidia deliciosa) had more Ca accumulation as

compared to shaded fruit (Montanare et al., 2006).

Endogenous nutrient status in relation to tree age, canopy position and during fruit

development is well documented in literature. However, exploration of endogenous nutrient

concentrations in relation to tree age and canopy position is not reported in Kinnow mandarin

trees. Moreover endogenous nutrient status in Kinnow mandarin tree during fruit growth and

development is not reported from trees of various age groups.

Alkaline pH and calcareous nature of soil in citrus growing areas resulted in nutrient

deficiencies (Yasin and Manzoor, 2010) and poor fruit quality. Exogenous application of

macro and micronutrients are well documented in literature in improving fruit quality. Some

23

research work on effect of macro and micronutrients on citrus fruit quality is summarized as

under:

2.7.1.5.1Physical fruit quality

Nitrogen is a pre-requisite nutrient for citrus growth, yield and fruit quality (Alva et al., 2003;

Thompson et al., 2002). Nitrogen plays an important role in various plant biological

processes such as cell division, growth, photosynthesis and respiration (Abbas and Fares,

2008). Saleem et al. (2008b) reported that winter application of low-biuret urea (LBU)

reduced fruit weight and fruit diameter of sweet orange cv. 'Blood Red'. El-Otmani (2004)

reported that urea application to 'Clementine' mandarin increased juice (%) during on-year

while, off-year it had no effect on any fruit quality parameters. Khan (2009) reported that

low-biuret urea application increased juice and rag contents of 'Kinnow' mandarin fruits.

Koseoglu (1995) found a positive correlation between N and rind thickness and Koo (1988)

reported that increasing N rates decreased rind thickness in citrus fruit however, Alva et al.

(2006) found non significant effect of N application on rind thickness of 'Hamlin' orange.

Dou et al. (2005) applied phosphorous at 0, 48 and 96 kg ha-1

to 'Flame' grapefruit trees and

reported that optimum P increased rind colour development, β-carotene and lycopene

contents. Phosphorus deficiency results in a thick rind with a hollow core particularly in

sweet oranges (Ladaniya, 2008).

Potassium plays numerous important functions in plant such as cell division, growth, and

sugar starch metabolism, protein synthesis, enzyme activation, stomatal functions,

photosynthesis, pH stabilization, plant water relations, and transport of metabolites (Abbas

and Fares, 2008). Ashraf et al. (2010) applied K2O (0, 50, 75, 100 kg ha-1

) in the form of

sulphate of potash along with P and N to K deficient orchards in four district of Punjab

(Jhang, Sargodha, T.T. Singh and Faisalabad) and reported that K increased fruit size and

weight of sweet oranges. Ashkevari et al. (2010) reported that K application increased fruit

yield, fruit diameter and fruit length in citrus. Fruit firmness was improved by K in 'Fortune'

mandarin (El-Hilali et al., 2004). Foliar application of KNO3 increased fruit size in 'Valencia'

orange (Du Plessis and Koen, 1988). Erner (1993) found that KNO3 applications increased

fruit size of 'Valencia' orange and grapefruit. K increased fruit size and yield Ritenour et al.

(2002), but excessive K results in large and coarse fruit with thick and greenish rind

24

(Wutscher and Smith, 1993). Potassium application to sweet orange increased rind thickness

and juice (%) (Ashraf et al., 2010). Ashkevari et al. (2010) applied 750, 1500, 2250 and 3000

g of K per tree and reported that 2250 g per tree markedly increased rind thickness of citrus

fruit.

Calcium (Ca) is an essential plant nutrient involved in many physiological processes in living

organisms involving enzyme activities and stabilizing cell membranes (Conway et al., 1994).

It also acts as anti-ripening and anti-senescence agent in fruit (Lester and Grusak, 1999). In

'Fortune' mandarin, preharvest applications of calcium nitrate improved fruit firmness (El-

Hilali et al., 2004). In Ca deficient tissues, the activity of polygalacturonase (PG) increases

resulting in the disintegration of cell walls and collapse of the affected tissues (Marschner,

1995). 'Kinnow' mandarin fruit treated with calcium nitrate before harvest had more juice

(%) when compared with control (Singh and Sharma, 2011).

Micronutrient applications are also well documented in improving fruit quality of citrus.

Foliar application of Boron significantly increased fruit weight (Ullah et al., 2012) and Zn

improved juice volume of Kinnow mandarin (Ashraf et al. 2012).

2.7.1.5.2 Biochemical fruit quality

Alva et al. (2006) found no significant effect of N application on juice quality of 'Hamlin'

orange, contrarily Koo (1988) reported that increasing N rates increased juice acidity in citrus

fruits. Saleem et al. (2008) reported a significant increase in TSS, TSS:TA, total sugars, and

AA concentrations of 'Blood Red' sweet orange by the application of LBU.

Dou et al. (2005) reported that P application to 'Flame' grapefruit reduced ascorbic acid and

total sugars contents. El-Hilali et al. (2004) found that acidity contents decreased by

preharvest applications of calcium nitrate to 'Fortune' mandarin fruit.

Potassium application increased TSS, TSS:TA ratio and ascorbic acid and reduced acidity

(%) of juice (Ashraf et al., 2010). Fruit biochemical quality parameters like TSS, acidity and

ascorbic acid were improved, while TSS:TA ratio were reduced by K applications

(Ashkevari et al., 2010). El-Hilali et al. (2004) reported that preharvest K application

increased acidity of 'Fortune' mandarin fruit. El-Otmani (2004) reported that KNO3

application to 'Clementine' mandarin increased maturity index during off year while on year

25

it increased TA and ascorbic acid concentrations of fruits. Dou et al. (2005) applied 0, 186

and 372 kg of K ha-1

to 'Flame' grapefruit trees and reported that K reduced rind colour

development, lycopene and β-carotene contents while increased ascorbic acid and total

sugars contents.

Foliar application of Zn improved TSS, TA, pH and ascorbic acid (Ashraf et al., 2012) and

boron improved TSS:TA ratio, ascorbic acid and total sugars of Kinnow mandarin fruit

(Ullah et al., 2012)

Overall, both macro and micro nutrients appears to influence fruit external and internal

quality parameters. However, very little work is done on nutrient management of young

Kinnow orchards. Nutrient differences between young and old trees, might be the reason of

poor fruit quality in young orchards. A comparison of endogenous nutrient concentrations of

various age groups is prerequisite to develop nutrient management strategies for young

orchards.

26

Chapter-3

GENERAL MATERIALS AND METHODS

These research studies were conducted during 2007-2011, and comprised of two parts:

1. Investigating the basis of poor fruit quality in young orchards

2. Improving fruit quality of young orchards

In part one i.e. investigating the basis of poor fruit quality in young orchards following three

experiments was included:

Effect of tree age on fruit quality of 'Kinnow' mandarin

Effect of tree age and fruit size on storage potential of 'Kinnow' mandarin

'Kinnow' fruit growth and development in relation to changes in endogenous levels of

nutrients and pectin

The 2nd

part of the study i.e improving fruit quality of young orchards comprised of

following experiments:

Effect of timing of plant growth regulators application on fruit quality and storage

potential of 'Kinnow' mandarin

Exogenous applications of PGRs and nutrients on fruit quality of 'Kinnow' mandarin

General materials and methods are detailed as under, while specific information is given

under separate experiments.

3.1 Plant Material

All the experiments were conducted at a commercial citrus orchard in the main 'Kinnow'

mandarin growing district, Sargodha (latitude 32° 03ʹ N and longitude 72° 40ʹ