Dirk Avé Business Technical Support Manager Valent BioSciences Corporation
ABIM, Oct 26, 2010
Creating Value Through Technology and People™
Bacillus thuringiensis (Bt)
Bt-based insecticides are complex entities
Using the science and art of fermentation, Bt strains produce a mixture of specific insecticidal proteins as active ingredient Bt mode of action is unique and different from any chemical insecticide, with commercially proven efficacy for over 40 years. Bt biological insecticides are used in IPM, Organic and Low Residue insect control
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Bt Manufacturing at VBC
From large scale (100 000 L) fermentation to downstream processing,
manufacturing is geared to producing and preserving the best Bt active ingredient mixture at the most economical cost.
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Manufacturing Requirements Regulatory Agencies provide the framework Æ Æ Manufacturer sets the standards
i.e. US EPA approves the product AND the process
9 All tox/path and environmental tests related to process 9 Processes carried out as filed with EPA 9 Batch records shipping, inventory, disposal records kept
Valent BioSciences sets it’s own high standards
9 Frequent in-process sampling for quality 9 Potency checks after each major process step 9 On-site insect bioassay facilities 9 Decades history with Btk and Bta standards
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Commercial QC of Bt products Commercially, bioassay is the basic QC method. VBC formulates at least at the labeled potency. Example: Potency of XenTari® on cabbage looper, Trichoplusia ni, is 15 000 IU/mg product. Below are QC bioassay results from 35 large scale XenTari® production runs over several years. Potency
Inside a Bt fermentor, commonly 100 000 liters per batch.
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Secondary Analytical Methods for Bt Products
1. SDS-PAGE (electrophoresis method) to quantify Bt toxin proteins 2. HPLC (liquid chromatography method) to separate and quantify individual Bt toxin proteins 3. Gene profiling of Bt toxins 4. Genomotyping (cooperation with TNO, Zeist, the Netherlands)
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Large Variability in Bt Quality, Asian Market
Clearly labeled as Btk SDS-PAGE analysis but no Cry2 present (LC50 in µg/ml on cabbage looper) Lane: 1. Molecular Weight Marker 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 2. DiPel® Standard 0.7µg 3. DiPel® Standard 2.1µg 4. DiPel® Standard 4.1µg Bt toxin proteins (Cry1 type) 5. no sample Blank lane 6. XenTari® (Control) 7. Product 1 SE Asia (LC50: 73.6) Bt toxin proteins (Cry2 type) 8. Product 2 SE Asia (LC50: 38.8) 9. Product 3 SE Asia (LC50: 64.9) 10. Product 4 SE Asia (LC50: 45.3) 11. Product 5 SE Asia (LC50: 74.4) 12. Product 6 SE Asia (LC50: 78.7) 13. Product 7 SE Asia (LC50: 26.5) Although 2 Bt products of 14. DiPel® DF (Control) (LC50: 14.3) 16000 IU/mg, they have 15. Molecular Weight Marker suspiciously low Cry toxin SE Asia Products 1 and 4 were labeled 16000 Exp 435others IU/mg, 081805g2 VBC IU/mg (except XenTari ) at 32000 protein content
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3 Lots, each as samples A & B, from a European competitive Bt product code KG- 0601
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DiPel® DF compared to competitive Bt product
Tomato fields – July 13, 2006
Plot 1 Plot 2
Field Rep: Michael Crop: Industry tomato (organic Harvest Date: 20 +
Schröder farming) 21/09/2006 Investigator: Germán Pest: Helicoverpa armigera Plot size: > 1 ha Martinez Application Date: 5 applications Site: Finca Huescar, on 20/07, 04/08, 10/08, 18/08, and Granada 01/09/2006 with a field boom, 600 Country: Spain – 1000 L/ha, 1 kg/ha
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Tomato Yield in kg / ha, DiPel® DF vs Competitive Bt
50000 45000 40000 35000 30000 kg / ha
25000 20000 15000 10000 5000 0 kg/ha w ithout damage Total kg/ha
DIPEL DF KG-0601
Kg Tomatoes per ha Total Kg of
without damage Tomatoes per ha DIPEL DF 37 450 44 441 Competitive Bt 28 206 36 231
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Conclusions
1. Worldwide and within Europe significant differences in
quality of Bt products exist. In Asia and Latin America we have found Bt illegally “enhanced” with a chemical insecticide. In Australia and Europe, we have found Bt products with very little toxin protein, and corresponding very low potencies. 2. Maintaining the quality of Bt products is left up to the manufacturer. As bio-pesticide manufacturers, we should work together to ensure a certain level of quality based on simple guidelines. a) repercussions for companies that ameliorate Bt with chemical insecticides b) certain minimum levels of Bt toxin protein weight in the product based on label potency
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