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Slide 1 :
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Smoke Emilio Ghisalberti (UWA)
Kingsley Dixon (KPBG)
Robert Trengove (Murdoch) Investigation of the
germination promoting
compounds in smoke Gavin Flematti
The University of Western Australia |
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Slide 3 :
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Solving the bushfire germination mystery…….. |
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Slide 4 :
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Smoke as a germination cue In 1990, researchers in South Africa identified smoke as the key agent for stimulating germination of a threatened fynbos species Audouinia capitata (Bruniaceae)
Researchers at Kings Park and Botanic Garden (Western Australia) have since identified smoke as the key element for promoting the germination of many Australian native seeds
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Slide 5 :
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Smoke found to improve
seed germination of
many Australian native
species |
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Slide 7 :
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Sites with smoke-stimulated germination of wild species And the list of species includes…. lettuce, celery, parsley and others…. |
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Slide 8 :
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Apparatus for generating Plant Derived Smoke ‘Smoke-water’ ‘Aerosol Smoke’ |
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Slide 9 :
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Benefits for mining restoration Lateritic nickel project Mineral sands mining restoration - BUT… need 10 tonnes smoke water per hectare! |
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Slide 10 :
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Attempts to identify the active compound(s) Baldwin et al. (1994) identified 71 compounds and tested a total of 233
Dixon et al. (1997) identified a further 43 compounds
Researchers in South Africa (1995) and California (1997) have also identified compounds in smoke Importantly, burning cellulose produces the active compound(s)
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Slide 11 :
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Lettuce (Grand Rapids) The Needle in the Haystack - finding the chemical(s) in smoke responsible for germination Conostylis aculeata Stylidium affine Bioassay - Test that uses a biological indicator for tracking the active components through separation steps |
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Slide 12 :
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Isolation of Smoke Compounds Smoke Water |
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Slide 13 :
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Isolation of Smoke Compounds Ether extract Smoke Water Ether layer Aqueous layer |
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Slide 14 :
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Isolation of Smoke Compounds Ether extract Smoke Water Ether
layer Aqueous
layer |
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Slide 15 :
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Isolation of Smoke Compounds NaHCO3 soluble
(stronger acids
- COOH) Ether extract Smoke Water Ether layer NaHCO3 layer |
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Isolation of Smoke Compounds NaHCO3 soluble
(stronger acids
- COOH) Ether extract NaOH soluble
(weaker acids
- phenols etc.) Smoke Water Ether layer NaOH layer -Activity found in the neutral fraction- |
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Slide 17 :
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The Haystack! GC-MS chromatogram of neutral fraction |
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Slide 18 :
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Lots of Chromatography! Chromatography -
Physical method of
separating compounds Classical Liquid
Chromatography Stationary
phase Mobile
phase |
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Slide 19 :
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General separation scheme Alumina Neutral Fraction 15 Fractions |
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Slide 20 :
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Typical germination of Grand Rapids lettuce when tested with C18 fractions |
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Slide 21 :
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~ 2 years later…Only 3 Compounds! All had molecular formula C8H6O3 (mw=150)
Separated and tested for activity
1 2 3 UV recorded at 320nm 25% MeOH/water |
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Slide 22 :
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Testing of the 3 compounds |
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Slide 23 :
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Identification of the 3 compounds Chemical structure proposed based on Nuclear Magnetic Resonance (NMR) and mass spectrometry (MS) data |
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Slide 25 :
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Germination of karrikinolide (2) with Grand Rapids Lettuce |
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Slide 26 :
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Germination of karrikinolide (2) with confirming native species |
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Slide 27 :
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Back to the haystack! |
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Slide 28 :
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Expanded CDS-neutral fraction |
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Slide 29 :
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Summary First chemical identified from smoke that promotes germination
Novel structure – new to science
Devised a method of synthesis
Bioassay data demonstrating effect with species from Australia,
South Africa and North America
Potential uses in agriculture, horticulture, land restoration etc… Compared with smoke water, karrikinolide operates
at < 1 g per hectare |
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Slide 30 :
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What other species…?
What other activity…?
Is there a better (simpler) synthesis…?
Are there active analogues…?
What is the mode of action…?
Potent Germination Promoter |
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Slide 31 :
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Mode of Action?? Label = fluorescent Questions ?? |
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