ABSTRACT
Laboratory experiments to evaluate the insecticidal activity of five plant extracts, clay and permethrin against bean beetle Callosobruchus maculatus (F.) on mung bean (Vigna radiata (L.) Wilczek) were carried out in the Teaching and Research Laboratory of the College of Crop and Soil Sciences, Michael Okpara University of Agriculture, Umudike. Five plant extracts: Tephrosia vogelii leaves, Tephrosia vogelii stem, Piper guineense fruits, Xylopia aethiopica fruits, and Carica papaya seeds were applied at four levels: 0.0 g, 1.0 g, 2.0 g and 3.0 g per 20 g of mung bean (w/w). Clay and permethrin dusts were equally applied at the above rates. The experiment was laid out in a 7 x 4 factorial fitted into a completely randomized design (CRD) with three replications. The effects of these plant extracts, clay and permethrin on mortality of adult Callosobruchus maculatus were evaluated at 7, 14, 21 and 28 days after treatment. Also the effect of these treatments on oviposition was determined and recorded at 7 days after treatment. Adult emergence from the treated mung bean and control was evaluated. Germination test for viability, weight loss, percentage damage were carried out at the end of the experiment. Phytochemical analysis of the plant extracts was carried out to identify the active principles responsible for their insecticidal activities. Proximate composition of mung bean was determined to evaluate the effect of the treatments on the basic food components. Results indicated that plant extracts controlled the bean beetle especially when applied at appropriate dosage of 3.0 g/20 g. Mortality increased significantly (P<0.05) in all the plant extracts with increase in dosage and length of exposure. Oviposition and adult emergence counts were significantly (P<0.05) suppressed. Percentage seed weight loss and damage were also suppressed. The viability and proximate composition of seeds tested with plant extracts, clay and permethrin were not adversely affected when compared with the untreated seeds. Permethrin was significantly (P<0.01) superior to the test plant extracts in all the parameters followed by clay though not different from Tephrosia vogelii leaves and Piper guineense fruits. Tephrosia vogelii leaves recorded the best protection followed by Piper guineense fruits at 3.0 g/ 20 g than other extracts. Phytochemical analysis revealed that plant extracts posses high values of alkaloids, flavonoids and saponins. Gas chromatograph mass spectometry (GC-MS) analysis revealed the presence of a wide array of bioactive volatile fatty acids in the extracts of the test plants. The major and highest dominant bioactive fatty acids present in the extracts were; Oleic acid (19.90 - 60.79 %), Hexadecanoic acds (26.28 %), Benzene, 1,3-bis (2-2-dimethylpropyl)-2,4,5,6-tetramethyl (19.79 %), 7H-Pyrazolo(4,3-d)pyrimidin-7-one,1,4-dihydro-3-beta-D-ribofuranosyl (14.60 %), Benzene (1-methoxy-1-methylethyl) (9.64 %), Benzene (isothiocyanatomethyl) (8.93 %) Eicosane (6.30 %), 7-Oxabicyclo (4.1.0) heptanes, 3-oxiranyl (6.10 %) and these may be responsible for the observed insecticidal activity of the plant extracts against Callosobruchus maculatus.
TABLE OF CONTENTS
Title page i
Declaration iii
Dedication iv
Certification v
Acknowledgement vi
Table of Contents vii
List of Tables viii
List of Figures xv
List of Plates xvii
Abstract xviii
CHAPTER 1 : INTRODUCTION 1
1.1
Production Constraints
and Prospects for Control 2
1.2
Objectives 5
CHAPTER 2 : LITERATURE REVIEW 6
2.1 Morphology
and Ecological Requirements of Mung Bean 6
2.1.2 Nutritional
content of mung bean 7
2.2 Insect
Pests of Stored Grains 8
2.3 Management
of Cowpea Bruchid in the Store 9
2.3.1 Storage in
the pod and seed 13
2.3.2 Storage in
fire place and use of solarization 14
2.3.3 Use of low
temperature 14
2.3.4 Modification
of storage atmosphere 14
2.3.5 Use of
plant extracts and oils 15
2.3.6 Chemical
control 16
2.3.7 Use of
inert pheromones 17
2.3.8 Use of
irradiation 17
2.4 Insecticidal
Properties of Some Plant Extracts 18
2.4.1 Use of plant materials for the control of cowpea bruchid -
(Callosobruchus maculatus) 20
2.4.2 Mode of
action of plant-based insecticides 25
2.5 Response
of Insect Pests to Botanical Insecticides 26
2.5.1 Repellency 26
2.5.2 Feeding
deterrents / antifeedants 27
2.5.3 Toxicity 27
2.5.4 Growth
retardants 28
2.5.5 Sterility 28
2.6 Botany
and Agronomic Characteristics of Study Plants 29
2.6.1 Tephrosia vogelii 29
2.6.2 Piper guineense 31
2.6.3 Xylopia aethiopica 33
2.6.4 Carica papaya 34
2.6.5 Inert
dusts 35
2.7 Biology
of Bean Beetle 36
2.7.1 Egg
production, laying pattern and larval life 37
2.7.2 Adult
emergence and active form 38
CHAPTER 3 :
MATERIALS AND METHODS 40
3.1 Location
of Experiment 40
3.2 Insect
Culture 40
3.3 Sources
of Plant Materials 40
3.4 Sources
of Experimental Seeds 52
3.5 Evaluation
of Powdered Extracts of the Plant Materials 52
3.5.1 Preparation
of plant powders 52
3.5.2 Seed
treatment 53
3.6 Proximate
Analysis of Mung Bean Seeds 54
3.6.1 Moisture
content 54
3.6.2 Ash
content 54
3.6.3 Crude
protein 54
3.6.4 Fat 55
3.6.5 Crude
fibre 56
3.7 Phytochemical
Analysis of Plant Materials 56
3.7.1 Tannin 56
3.7.2.
Flavonoids 57
3.7.3 Saponins 57
3.7.4 Alkaloids 57
3.8 Extraction
of Phytochemical Composition of Plant Materials 58
3.8.1 Chloroform
extraction of the plant residues 58
3.8.2 Gas chromatography- mass spectrometry (GC-MS) analysis of
the plant residues 58
3.9 Statistical
Analysis 59
CHAPTER 4 : RESULTS AND DISCUSSION 60
4.1 Insecticidal Activity of some
Plant materials, Clay and Permethrin
against Callosobruchus maculatus
(F.) (Coleoptera : Chrysomelidae)
on
stored Mung bean 60
4.2 Effect of selected Botanicals, Clay and
Permethrin on Adult Mortality
of
Cowpea Bruchid on variety SML-668 60
4.3 Effect of selected
Botanicals, Clay and Permethrin on Oviposition
of Callosobruchus maculatus 72
4.4 Adult Emergence of Cowpea Bruchids on Mung
Bean treated with
selected Botanicals, Clay and Permethrin on (SML-668) 72
4.5 Seed Weight loss of Mung Bean Seeds (SML-668)
mixed with
treatments and infested with Callosobruchus
maculatus 76
4.6 Percentage Damage of
Mung Bean Seeds (SML-668) by
Callosobruchus maculatus 76
4.7
Percentage Germination of Mung Bean Seeds (SML-668) treated
with selected Botanicals, Clay and Permethrin 78
4.8 Effect of selected Botanicals, Clay and
Permethrin on Adult Mortality
of Cowpea Bruchid on variety (NM-94) 81
4.9 Effect of selected Botanicals, Clay and
Permethrin on Oviposition
of Callosobruchus maculatus 88
5.0 Adult Emergence of Cowpea
Bruchid treated with selected
Botanicals, Clay and Permethrin on (NM-94) 88
5.1
Seed Weight loss of Mung Bean seeds (NM-94) mixed with treatments
and infested with Callosobruchus
maculatus 89
5.2
Percentage Damage of Mung Bean seeds (NM-94) by
Callosobruchus maculatus
92 5.3 Percentage Germination of mung bean seeds
(NM-94) treated with
selected Botanicals, Clay and Permethrin 95
5.4 Proximate Analysis of Mung
Bean seeds 97
5.5
Phytochemical Analysis of Mung Bean seeds 97
5.6 Gas Chromatography Mass
Spectrometry (GC-MS) Analysis of
volatile compounds of the Plant
Materials 102
5.7 Biological Activities of some isolated
constituents of the test Plant
Materials 133
CHAPTER 5:
CONCLUSION AND RECOMMENDATION 148
REFERENCES 151
APPENDICES 169
LIST OF TABLES
1
Plant materials evaluated for insecticidal
properties 41
2
Percentage mortality of adult Callosobruchus maculatus
exposed to powders
of some plant materials, clay and
permethrin on variety (SML-668) at 7 DAT 61
3
Percentage mortality of adult Callosobruchus maculatus
exposed to powders of some plant materials, clay and
permethrin on
variety (SML-668) at 14 DAT 63
4
Percentage mortality of adult Callosobruchus maculatus
exposed to powders of some plant materials, clay and
permethrin on variety (SML-668) at 21 DAT 64
5
Percentage mortality of adult Callosobruchus maculatus exposed
to powders of some
plant materials, clay and permethrin on
variety (SML-668) at 28 DAT 67
6
Effect of interaction between variety x protectants on mortality
of Callosobruchus maculatu 68
7
Effect of interaction between protectants x rates of application
on
mortality of Callosobruchus maculatus 69
8
Effect of variety x protectants x rates interaction on mortality
of Callosobruchus maculatus 70
9 Effect of the test powders on oviposition
of C. maculatus on mung
bean seeds 74
10 Percentage adult emergence of C. maculatus on mung bean seeds
treated with test powders 75
11 Percentage seed weight loss of mung bean
seeds infested by
C. maculatus 77
12 Percentage damage of mung bean seeds
infested by C. maculatus 79
13 Percentage germination of mung bean
seeds treated with some
test powders 80
14 Percentage mortality of adult Callosobruchus maculatus exposed
to powders of some plant materials, clay and
permethrin on
variety ( NM- 94)
at 7 DAT 83
15 Percentage mortality of adult Callosobruchus maculatus exposed
to powders of some plant materials, clay and
permethrin on
variety
(NM- 94 ) at 14 DAT 84
16 Percentage mortality of adult Callosobruchus maculatus exposed
to powders of some plant materials, clay and
permethrin on variety
(NM- 94) at
21 DAT 86
17 Percentage mortality of adult Callosobruchus maculatus exposed
to powders of some plant materials, clay and
permethrin on variety
(NM- 94) at 28 DAT 87
18 Effect of the test powders on oviposition
of Callosobruchus
maculatus
on mung bean seeds 90
19 Percentage adult emergence of Callosobruchus maculatus on
mung bean seeds treated with test powders 91
20 Percentage seed weight loss of mung bean
seeds infested by
Callosobruchus maculatus 93
21 Percentage damage of mung bean seeds
infested by C.maculatus 94
22 Percentage germination of mung bean seeds
treated with some
plant materials 96
23 Proximate composition of two varieties of
mung bean 99
24 Quantitative analysis of mung bean seeds 99
25 Proximate composition of mung bean seeds
after treatment 100
26 Phytochemical analysis of the plant
materials 101
27 Volatile compounds of T. vogelii leaf detected by GC- MS
105
28 Volatile compounds of T. vogelii stem detected by GC- MS
111
29 Volatile compounds of P. guineense fruit detected by GC- MS 118
30 Volatile compounds of C. papaya seeds detected by GC- MS 124
31 Volatile compounds of X. aethiopica fruit detected by GC-
MS 129
32 Biological activities of some of the
chemical compounds isolated
from the Test Plant materials 134
LIST OF FIGURES
1. GC – MS chromatogram of volatile compounds of Tephrosia vogelii
leaves 104
1.1. Spectral structure of 7- Oxabicyclo (4.1.0)
heptanes, 3- oxiranyl 107
1.2. Spectral structure of Benzo (c) carbazole 107
1.3. Spectral
structure of Benzene, 1,3 bis (2,2 –dimethyl)-2,4,5,6-
tetramethyl 108
2. GC-
MS chromatogram of volatile compounds of Tephrosia
vogelii
stem 110
2.1. Spectral structure of 1,3- Benzenedicarboxylic
acid, 5- nitro 114
2.2. Spectral
structure of Eicosane 114
2.3. Spectral structure of 2H- 1,3-
Thiazine-6-carboxylic acid,
3-amino
tetrahydro-2-(methylimino) -4-oxo-methyl ester 115
3. GC- MS chromatogram of volatile compounds of Piper guineense
fruit 117
3.1. Spectral
structure of Cyclohexane, 1-ethenyl-1-methyl-2-(1-methyl
ethenyl)- (1-methyl ethylidene) 120
3.2.
Spectral structure of Naphthalene, 1,2,3,4,4a,5,6,8a-octahydro-7
-methyl-4- methylene-1-(1-methylethyl)-(1α,4aβ,8aα) 120
3.3. Spectral structure of Benzene,
(1-methoxy-1-methylethyl)- 121
3.4.
7H-Pyrazolo(4,3-d)pyrimidin-7-one, 1,4-dihydro-3,5 D-ribofuranosyl) 121
4. GC
– MS chromatogram of volatile compounds of Carica
papaya seed 123
4.1. Spectral
structures of Benzene, (isothiocyanatomethyl)- 125
4.2. Spectral
structures of n- Hexadecanoic acid 125
4.3. Spectral
structures of Oleic acid 126
5. GC
– MS chromatogram of volatile compounds of Xylopia
aethiopica
fruit
5.1
1H-3a,7-Methanoazulene, octahydro-1,4,9,9-tetramethyl 132 5.2. Spectral
structures of 1H- Naphtho(2,1-b)pyran, 3-ethenyl dode-
cahydro-3, 4a,7,7,10a-pentamethyl,
(3S(3α,4aα,6Aβ,10aα,10bβ) 132
LIST
OF PLATES
1i. SML- 668 variety of mung bean seeds 10
ii. NM- 94 variety of mung bean seeds 11
iii. Infested mung bean seeds by C. maculatus 12
2. Piper
guineense fruits 42
ii. Piper
guineense ground powder 43
3. Tephrosia
vogelii leaf / stem 44
ii. Tephrosia
vogelii leaf powder 45
iii. Tephrosia vogelii stem powder 46
4. Carica
papaya seeds 47
i. Carica
papaya dried seeds 48
ii. Carica papaya seed powder 49
5. Xylopia
aethiopica fruits 50
ii. Xylopia
aethiopica powder 51
6. GPS. Satellite aerial view of Crop and
Soil Science laboratory
Environment of MOUA Umudike, Abia State.
Nigeria 169
CHAPTER 1
INTRODUCTION
Mung bean (Vigna radiata (L.) Wilczek) belong to the family Fabaceae
(Lambrides and Godwin, 2006). It is an annual erect plant reaching a height of
about 0.15-1.25 m. It serve as a good source of proteins, carbohydrates, and
vitamins for mankind all over the world. Mung bean (a legume) popularly called green gram or golden gram account for almost half
of the dry beans grown in South and South Eastern Asia (AVRDC, 1992) and in
recent years, has been introduced into East and Central parts of Africa, USA and
parts of Australia (Bisht et al.,
1998). Mung bean is widely grown as a tropical and subtropical crop. It is a low input crop and mainly grown
for its seeds which have high protein level, easily digestible and is consumed
as food. The non-flatulent behaviour (digestibility) of mung bean seeds made
it to be superior over other pulses making it suitable for
children, vegetarians and older people (Ghafoor et al. 2003).
Mungbean is rich in vitamin A, B1,
B2, C and niacin as well as minerals such as potassium, and calcium
which are necessary for human body. According to (USDA, 2004) the composition
of mature mung bean seeds per 100 g edible portion is: water 9.1g, energy 1453 kj
(347 kcal), protein 23.9 g, fat 1.2 g, carbohydrate 62.6 g, dietary fibre 16.3 g,
Ca 132 mg, Mg 189 mg, P 367 mg, Fe 6.7 mg, Zn 2.7 mg, Vitamin A 114 iu, thiamin
0.62 mg, riboflavin 0.23 mg, niacin 2.3 mg, vitamin B6 0.38 mg, folate 625µm
and ascorbic acid 4.8 mg. Its essential amino acid is: tryptophan 260 mg, lysine 1664 mg,
methionine 286 mg, phenylalanine 1443 mg, threonine 782 mg, valine 1237 mg,
leucine 1847 mg and isoleucine 1008 mg. It is utilized as a
supplement to other grains and starchy
food. Mung bean starch
is considered to have a low glycaemic index ie. to raise the level of
blood sugar slowly and steadily (USDA, 2004).
1.1 PRODUCTION CONSTRAINTS AND
PROSPECTS FOR CONTROL
Production of this crop
has been constrained by insect pests among other factors. Mung bean infestation
by insect pests starts from the field and continues in storage. A number of
insect pests decimate the crop in the field namely: Helicoverpa armigera, Creontiades
dilutus, Nezara viridula, Piezodorus oceanicus, Maruca vitrata, Riptortus dentipes, Melanacanthus
scutellaris, and Thrips. Among the different species of sucking insect
pests, Apis craccivora, Empoasa spp., Cicadella viridis and Bemisia tabaci-white fly are the major insects. These insect pests not
only reduce the vigour of the plants by sucking the sap, but also transmit
diseases and affect photosynthesis as well and ultimately yield losses are
recorded (Asawalam and Anumelechi, 2014).
Storage beetle or bruchid, Callosobruchus maculatus F. (Chrysomelidae),
are minor pests in the field, which assume a major pest status during storage.
Bruchids (Coleoptera : Chrysomelidae), especially those belonging to the genus Callosobruchus may be harmful to stored
seeds of most of these different legumes (Ofuya and Bamigbola, 1991). It is a
cosmopolitan field-to-stored pests of legumes. It causes substantial
quantitative and qualitative losses manifested by seed perforations and
reduction in weight, market value and germinability of seeds (Ogunwolu and
Odunlami, 1996; Adeduntan and Ofuya, 1998). The initial infestation occurs in
the field and then carried over to stores, where the population can rapidly
build up. While in the field, eggs are laid on the pods, although adults prefer
to slip inside the pods through holes made by other pests and lay eggs directly
on the seeds. After the crop is harvested, the bruchids multiply and do
considerable damage by the larvae feeding inside the seeds of stored mung bean.
The protection of crop plants and seeds from pests and diseases has
been achieved in industrialized countries almost through the application of
chemical control means. Although some chemicals such as sulphur and copper
sprays have been used as an aid to agriculture for a long time, with the
increasing mechanization of agriculture, the use of synthetic chemicals have triumphed
as an independent principle of plant protection. Most farmers in developing
countries cannot afford high technology, efficient storage facilities, and
synthetic insecticides and in some cases, they lack the technical knowledge of
application and are therefore exposed to a lot of risks. In the recent past, the control of insects and
other storage pests was basically on the
use of chemical control methods comprising fumigation of stored
commodity with carbon disulphide, phosphine or dusting with malathion,
carbaryl, pirimiphos-methyl or permethrin. These chemicals have been reported
to be effective against C. maculatus
and other insect pests (Caswell and Akibu, 1980; Akinkurolere et al., 2006). However, the problems of
many synthetic insecticides include: high persistence, poor knowledge of
application by resource - poor farmers, high cost, non availability, genetic
resistance, hazardous and unsustainable control measures (Adedire and Ajayi,
1996 ; Akinkurolere et al., 2006).
In the developed countries,
conventional fumigation technology is currently being scrutinised for many
reasons such as ozone depletion and carcinogenic concerns with phosphine
(Adedire, 2002). As part of the quest for an alternative to chemical
insecticides, research efforts are currently being focussed on eco-friendly
control measures such as irradiation, heat treatments, biopesticides,
integrated pest management, use of insect hormones and modified atmosphere (Lale, 1992; Adedire and Ajayi, 1996; Ofuya and Reichmuth,
2002; Follett et al., 2007; Begum
et al., 2009). Also the indiscriminate use of insecticides to keep stored products
under control calls for the urgent need to develop environmentally safe and
sound pest control techniques that must be economical, simple and practical to
use. The necessity to develop safe and more biodegradable alternatives to
synthetic insecticides has in recent years led to the concerted international
efforts at developing new sources from vast number of chemical substance in
plants (Olaifa et al., 1988).
Research is on-going for
natural plant materials that are ecofriendly, biodegradable, and with medicinal
values that may be used as grains protectant (Adedire and Lajide,
2003; Arannilewa et al., 2006; Ileke and Oni, 2011). Botanical insecticides have been
reported to have a wide range of biological activities against insects. These
include repellence and anti-feedant (Viglianco et al., 2005),
oviposition deterrence, toxicity, sterility, growth regulatory and fecundity
reduction, molting and respiration inhibition, and cuticle disruption (Tinzaara
et al., 2006). Today, researchers are
seeking new classes of natural insecticides that might be compatible with newer
pest control approaches (Rajashekar et al.,
2012). This has motivated research to evaluate naturally occurring toxicants
against C. maculatus which is a field - to -
store pest.
OBJECTIVES
The objectives of this study were to:
i.
evaluate the efficacy of plant
materials as protectants against Callosobruchus
maculatus in
stored mung bean.
ii. compare the efficacy of the plant materials with a synthetic insecticide
(permethrin dust) and clay in the control of C. maculatus.
iii. determine the effect of the plant materials on proximate
composition of the stored mung bean .
iv. identify the active constituents of
the plant materials responsible for their insecticidal properties.
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