Botanical pesticides in pm
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This is a lead lecture deliverd at BET 2010, CCSHAU, Hisar India on 20th Oct 2010.
Transcript of Botanical pesticides in pm
Global Biopesticide Market (in millions of dollars)
•The global pesticide market was valued at nearly $43 billion in 2009 and is expected to grow at a compound annual growth rate (CAGR) of 3.6% to reach $51 billion in 2014.
•Biopesticides segment is expected to grow at a 15.6% compound annual growth rate (CAGR) from $1.6 billion in 2009 to $3.3 billion in 2014.
•The global pesticide market was valued at nearly $43 billion in 2009 and is expected to grow at a compound annual growth rate (CAGR) of 3.6% to reach $51 billion in 2014.
•Biopesticides segment is expected to grow at a 15.6% compound annual growth rate (CAGR) from $1.6 billion in 2009 to $3.3 billion in 2014.
Registration of safer chemicals
Proportion of pesticide active ingredients that are considered to be safer (biological chemicals and reduced-risk conventional chemicals) has steadily increased over the last several years.
Source: EPA, 1999.
Cost to develop and time to market of various products
BOTANICAL PESTICIDES
ScopeIndigenous; most suited rural
milieu
· Biodegradable – Green
· Time tested through co-evolution
· Least side – effects
· Complex m.o.a; slow-resistance development
· Useful lead-templates for potent version
Ideally suited to biotechnological innovations & integration
Biodiversity
Drawback Action “ayurvedic” not
allopathic Situation / location /target
specific Low level & stability No volume and sales /
turnover - No MNC would prefer
Low yield / chemo-type, genotype, eco-type variations; plant part, age; storage conditions critical
Crude works better; synergistic
Push-pull or Stimulo – Deterrent Diversionary Strategy
Push Pull
Away from the crop Traps / trap crops
Masking of host attraction; (1R,5S) Myrtenal for Aphids fabae); Methyl salicylate against cereal aphids
Host attractants (Isothiocyanates of oilseed rape for the seed weevil and cabbage stem flea beetle
Repellents, antifeedants, oviposition deterrents
Aggregation, sex and oviposition semiochemicals
Attractants for predators and parasitoids; e.g. Faba beans produce 6-methy-5-hepten-2-one, an attractant for aphid parasitoid
Visual cues
CRITERIA OF SELECTION OF PLANT OR PLANT PARTS
Traditional knowledge (Ethno-medical)Farmers’ observation
Literature reportsChemotaxonomy
Random ScreeningAbundant availability
Commercialisation of botanical pesticides
Quality of raw material
Product standardization
Quality control
Stringent registration requirement
Problem of pest registration
PROBLEMS AND PROSPECTSPROBLEMS AND PROSPECTS
Botanical Pesticides: IPR and Policy Issues
Framing pesticide policy to protect human and ecosystem health
Patentability of the product should be considered. Prior publication of pesticidal properties of a compound could cause patent problem
Patenting synthetic analogues with no mention of the natural source of the chemical family might be safer than patenting the natural product
Softening registration requirements for natural products
PROSPECTS
Raising the threshold of active ingredients
Selection of germplasm tolerant to abiotic and biotic stress
Survey of plant biodiversity
Tissue and cell culture
On-farm production of botanical pesticides
Improving photo and thermal stability
Scientific investment in terms of money and human resource
BIOTECHNOLOGICAL TOOLS IN DISCOVERY
The development through in vivo screening
Gene expression profiling, a revolutionary tool in herbicide
discoveryGene Expression Profiling (GEP) with DNA microarrays (chips) is a new technology used to measure changes in the entire transcriptome, i.e. full complement of active genes, of an organism in a single experiment.
A catalogue of genetic fingerprints of the plant Arabidopsis thaliana, is created and each fingerprint being characteristic for a single herbicidal MoA is then used to rapidly classify herbicidal compounds from Ultra High Throughput Vivo Screening (UHTVS) according to their MoA.
Besides GEP also provides the opportunity to identify the affected metabolic pathway which is a first step towards the identification of novel herbicidal targets. Additionally GEP can identify the MoA of pro-drugs, which cannot be elucidated by conventional biochemical methods.
GEP provides insight into the interactions of any herbicidal compound with the entire plant metabolism with unprecedented accuracy and completeness.
The Principle of Gene Expression Profiling.
The virtual discovery cycle
Model organisms for target identification
The research screening platform
Strategy of identifying new targets
Test systems for targets in UHTBS
UHTVS - Automated evaluation of activity
Unique research platform – Network of complementary technologies to meet the
challenges in compound discovery
CAPACITY BUILDING
NATIONAL / GLOBAL NETWORK
OR
CONSORTIA
PUBLIC PRIVATE PARTNERSHIP
STRATEGIRES
BIOPROSPECTING OF ANTIFUNGAL PHYTOCHEMICALS
04/14/2023 PRE-SRC PRESENTATION
THE MOST EFFECTIVE ANTIFUNGAL PHYTOCHEMICALS AGAINST SHEATHBLIGHT OF RICE
45 49 53 57 61 65 69
Infection (%)
Control
Linalyl acetate
Terpinyl acetate
Carbendazim
Chinaberry
Garlic bulb extract
Aza concentrate
Eugenol
Piper betle
Seedlings soaked for 4 h in 0.1 % formulations pre-transplanting
04/14/2023 PRE-SRC PRESENTATION25.0 27.5 30.0 32.5 35.0
minutes
0.00
0.25
0.50
0.75
1.00
1.25MCounts Patchouli oil 5ppm,11-47-04 AM, 4-3-2008.SMS TIC Filtered
25:250
26.0
37 m
in
26.8
15 m
in
27.1
46 m
in
27.5
91 m
in
27.8
91 m
in
27.9
77 m
in
28.9
43 m
in
36.0
41 m
in
36.0
60 m
in
26.037 β- Patchoulene 4.35%
26.816 E-caryophyllene 3.02%
27.148 α- guaiene 13.53%
27.597 seychellene 7.24%
27.893 α- Patchoulene 5.11%
28.943 azulene 17.12%
36.041 patchouli alcohol 36.86%
GC-MS ANALYSIS OF PATCHOULI OIL
Disease control 76%Carbendazim 93.4%
CH3
CH3
OHCH3
Pogostemon cablin
04/14/2023 PRE-SRC PRESENTATION15 16 17 18 19 20 21 22 23
minutes
0.00
0.25
0.50
0.75
1.00
1.25
1.50
MCounts Cedar-Hex-5ppm1.SMS TIC Filtered40:400
17.9
82 m
in
18.6
93 m
in18.8
13 m
in
19.2
08 m
in
+ 1
9.4
45 m
in
+ 1
9.8
42 m
in
20.0
78 m
in
Disease control 67.8 %Carbendazim 93.4 %
GC-MS ANALYSIS OF CEDARWOOD OIL
α-himachalene 11.6% γ-himachalene 7.5% β-himachalene 40.4%Himachalol, 3.4%Deodarone 4.9%(E)-α-atlantone 2.1%
Himalayan Cedar Cedrus deodara (D. Don) G. Don. f. (“Deodar”)
Treatment Conc. (%)
D. s. infection( % )
A. a. infection( % )
Vigour index
NEEM 5 EC 0.05 0 8 1104
Mancozeb 0.25 0 5 1069.5
(PR+NP) 0.08 8 10 1056
PR (contact) 0.08 8 10 897
PR-fumigation 0.09 36 16 880
(PR+NO) 0.08 20 18 864
PR+NB 0.08 25 25 836
Control 52 24 630
(NP) 0.50% 45 22 559
Neem oil (NO) 0.3 40 20 558
Effect of palmarosa oil and neem products on
seed infection and seed vigour index in wheat
Effect of botanicals on seed infection and
seed germination of wheat
0
15
30
45
60
30 60 90 120
Storage period in days after application
se
ed
in
fec
tio
n (
%)
60
80
100
se
ed
ge
rmin
ati
on
(%
)
Mancozeb NEEM 5 ECPR+NP EugenolPR (contact) Inoculated controlMancozeb NEEM 5 ECPR+NP EugenolPR (contact) Inoculated control
Neem products C.graminicola
M.phaseolina
D.sorokiniana
P. sojae F. solani
Neem oil 90EC 20.5 (68.46)
40.0 (50.0)
28.0 (46.15)
50.5 (40.58)
38.0 (5.0)
NIM-76 20.5 (68.46)
31.0 (61.25)
15.1 (70.96)
50.0 (41.17)
27.0 (32.5)
Neem meliacins 20.0 (69.23)
20.0 (75.0)
20.0 (61.53)
45.0 (47.05)
37.5 (6.25)
Neem 5 EC 0 (100) 6.5 (91.87)
0 (100) 10.0 (88.23)
30.0 (25.0)
Azadirachtin (10%)
32.0 (50.76)
15.0 (81.25)
20.0 (61.53)
38.0 (55.29)
33.5 (16.25)
Mancozeb (0.25%) 0 (100) 0 (100) 0 (100) 0 (100) 6.5(83.75)
Control 65 (0) 80 (0) 52 (0) 85 (0) 40 (0)
ANTIFUNGAL SPECTRUM OF NEEM PRODUCTS
Essential Oils:Minimum inhibitory concentration (ppm)
Essential oilsD. sorokiniana
P. sojae
C. graminicolaF.
solaniM. phaseolina
Palmarosa 439.8 474.6 481.3 502.9 527.0Lemon grass 524.1 523.5 526.7 510.9 578.0
Jamrosa 600.0 588.0 597.3 568.3 622.6
citronella 640.2 640.8 630.9 667.1 686.9
Comparison of the best treatments on fungal infection and vigour index in wheat
600
800
1000
1200
Treatments
Vig
ou
r in
dic
es
0
11
22
33
44
55
Fu
ng
al i
nfe
cti
on
(%
)
vigour index D.s infection ( % )
Alternaria infection ( % ) D.s infection ( % )
Cyclotides are a structurally unique family of small disulfide-rich proteins embedded with highly stable structural motif and showing a broad range of roles in plant defense such as proteinase inhibition, anti-bacterial, anti-viral effects and insecticidal activities.
(Peligrini et al. 2007 Chen et al. 2005; Daly et al. 2004; Ireland et al. 2008; Jennings et al. 2001; 2005; Lindholm et al. 2002; Svangard et al. 2004).
Viola betonicifolia and V. canescene
Frozen in liquid N2 & groundedDried root powder (7 g)
CH2Cl2:MeOH (1:1) left overnight
Extract partitioned with CH2Cl2 and then waterConcentrated under vacuum and
then freeze dried
Dried material (100 mg)
Dissolved 33mg in 1 ml of 20% ACN:H2O with 0.1% HCOOHSubjected to reverse phase chromatographyEluted with 90% ACN:H2O with 0.1% HCOOHFreeze dried to get cyclotide enriched fractionLC-MS
analysis
Bioprospecting of cyclotides in Viola species
VCVB
Total Ion Chromatogram by Flow Injection Analysis (FIA)
TIC of VB sample through HPLC
TIC
XIC at Rt 5.37
XIC of VB sample through HPLC
TIC
XIC at Rt4.46
TIC and XIC of VB sample through HPLC
TIC
XIC at Rt 5.37
TIC and XIC of VC sample thr. HPLC
XIC at Rt. 23.30
MADHUCA INDICA (MAHUA)(Family Sapotaceae)
• Mahua valued for its oil bearing seeds and flowers
• Traditionally utilized for alcoholic beverage production
• Seeds kernel constitute over 50% non-drying oil (66% total unsaturated and 33% total saturated)
• Mahua seed kernel contains 2.5% toxic saponin content which must be removed for utilizing mahua cake as animal feed
• Efforts have been made to separate saponins from defatted seed cake for possible use as pest control agents
Defatted seed extractDefatted seed extract
Saponin rich materialSaponin rich material
Saponin concentrateSaponin concentrate
PartitioningWater: n-BuOH
PrecipitationAcetone
Hexane
Methanol
Mahua based Phytoadjuvants: Isolation and derivatization Mahua based Phytoadjuvants: Isolation and derivatization
Insecticidal Saponins, prosapogenins,
sapogenin, their esters and mahua oil
Insecticidal Saponins, prosapogenins,
sapogenin, their esters and mahua oil
SapogeninSapogenin
ProsapogeninsProsapogenins
Madhuca Seed/kernel powder Madhuca Seed/kernel powder
Saponin mixture Madhuca saponins (MI-I , MI-II and MI-III)
Saponin mixture Madhuca saponins (MI-I , MI-II and MI-III)
OHHOH2C
HO OH
C
O
O
Arab
Xyl
Arab
GluGlu OGlu
Api
Analytical / prep-LC method for separation of Madhuca saponins
Analytical LC Prep LCDetector PDA-213 nm PDA-213 nmColumn RP-18 254 x 4mm (5 µm) 250 x 50 mm (10 µm)Mobile phase Methanol -Water : 60:40 Methanol -Water : 60:40Flow rate 0.4 ml min-1 10 ml min-1
OHHOH2C
HO OH
C
O
O
Arab
Xyl
Arab
GluGlu OGlu
ApiOHHOH2C
HO OH
C
O
O
Arab
Xyl
Arab
GluGlu O
MI-III ( M +.1535.9) MI-I (M +.1241)
Photo-activated xanthotoxins from Ammi majus seeds
Xanthotoxins are known for the treatment of vitiligo, leucoderma asthma, angina and digestive system disorders
The plant is widely cultivated in India for these bioactive furanocoumarins which are used in the treatment of vitiligo and psoriasis
Like other many photo-activated compounds these furacoumarins may find use in pest control
8-methoxypsoralen
O O O
OCH3
5-methoxypsoralen
O O O
OCH3
Isolation and LC analysis of xanthotoxins from Ammi majus seeds
The seed contains furanocoumarins (5-, and 8-methoxypsoralens), which stimulate pigment production in skin that is exposed to bright sunlight
Process has been developed and standardized for the efficient extraction of furanocoumarin from seeds using fluorinated solvents such as phytosols. Extractions were completed within three cycles
LC method has been standardized for the analysis of furanocoumarins using UV detector (230 nm), RP-18 column and acetonitrile as solvent system
CURCUMA LONGA (Turmeric)
Turmeric oil Pest control (stored grains)Turmerone Pest controlCurcumins Antifeedant/antifungal
antioxidant/anticancerReduced curcumins Anti-oxidant/anti-ageing/cosmeticTurmerin Anti-oxidant
CURCUMINS: Isolation and LC analysis
RP 18 column 250mm x 4 mmInjection 20 ul Solvent system AcetonitrileFlow rate 0.5 ml/minUV detector 254 nmRetention time (Rt, min) Curcumin I 5.89 Curcumin II 5.57 Curcumin III 5.39
Curcumin I
Curcumin II
Curcumin III
Curcumin ethers
CURCUMA LONGA : REDUCED CURCUMINS
O
HO
H3CO
OH
OCH3
OO
HO
H3CO
OH
OCH3
O
OH
HO
H3CO
OH
OCH3
OOH
HO
H3CO
OH
OCH3
O
H2/Pd/C
H2/Pd/C
Curcumin I Tetrahydro curcumin I(Keto)
Tetrahydro curcumin I(enol)
OH
HO
H3CO
OH
OCH3
OH OH
HO
H3CO
OH
OCH3
OH
H2/Pd/C
Hexahydro curcumin I(Keto)
Octahydro curcumin I
Hexahydro curcumin I(enol)
Turmerin: Isolation and analysis of water soluble peptide
Turmeric powder
Suspended in boiling distilled water
Filter/centrifuge/decolorize supernatant
Lyophilization
Turmerin (0.1%)
It is stable to heat and light radiations It is a 5-KDa peptide containing 40 AAs Turmerone is an effective antioxidant/DNA
protectant/antimutagen
