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Isolation & identification of Novel microorganisms for the Treatment of Raw Crustacean Waste to Produce Chitinous Ma

Isolation & identification of Novel microorganisms for the Treatment of Raw Crustacean Waste to Produce Chitinous Materials. Carla Harkin. Dr Wolfram Brück Dr Catherine Lynch Dr Denis McCrudden. ChiBio: Work Package 2 Aims.

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Isolation & identification of Novel microorganisms for the Treatment of Raw Crustacean Waste to Produce Chitinous Ma

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  1. Isolation & identification of Novel microorganisms for the Treatment of Raw Crustacean Waste to Produce Chitinous Materials Carla Harkin Dr Wolfram Brück Dr Catherine Lynch Dr Denis McCrudden

  2. ChiBio: Work Package 2 Aims • Development of efficient, economical and environmentally-friendly pre-treatment of crustacean shell wastes to extract chitinous material(Raw material sources: Europe/Asia/Africa). • Stabilisation of chitinous material for transfer to project partners (UMB, INS, TUM, FhG). • Development of assays for constant quality control.

  3. Pre-treatment overview Raw material Prep Optimisation of chemical chitin extraction methods Identification of novel microorganisms Optimisation of microbial chitin extraction methods QC Up-scaling

  4. Raw Material preparation

  5. Chemical Pre-treatment 7. QC Analysis

  6. Microbial Pre-treatment • Chemical extraction methods- costly and a hazard to the environment. • Biological extraction methods- more cost effective while posing less cause for environmental concern. • Microbial chitin extraction methods: • Direct use of enzymes (Brück et al, 2011; Jo et al, 2011). • Fermentation (Jung et al, 2005; Jung et al, 2007; Rao et al, 2000). Fermentation • Initial optimisation, adapting methods from literature. • Identification of novel microorganisms for fermentation.

  7. Fermentation Process Raw material + Inoculum Shaking Incubator QC Shaking Incubator Harvesting & drying Change of Inoculum (Jung et al., 2005; Jung et al., 2007a,b; Oh et al., 2007)

  8. Isolation of Novel Microorganisms Isolation from soil samples Screening for useful properties Small scale fermentation Identification Large scale fermentation Characterisation

  9. Initial Screening • Demineralisation–acid production. • Replicated fermentation method on micro scale. • Deproteinisation–protease production. • Casein agar plates. • Colorimetric protease assay to quantify activity (Vishwanatha et al, 2009). Fig 1: pH indication after addition of Methyl Red Indicator. L-r: negative yellow, negative orange, positive intense red. Fig 2: Produciton of zone of proteolysis in casein agar.

  10. Selection of Organisms

  11. To be done • Identification of isolates; PCR, sequencing- BLAST • Characterization of isolates: biochemical tests, staining etc. • Continue to meet chitinous material production targets. • Continuation of fermentation experiments using raw crustacean waste from Ireland, Indonesia and Tunisia.

  12. Future prospects • With select cultures, the aim is to scale-up fermentations to pilot scale to investigate the reality of industrial large scale fermentations in the breakdown of crustacean waste. • Development of an economic, environmentally friendly, waste reducing facility. • Profit-generating business opportunities for Irish Seafood Industry.

  13. References • Brück, W., M., Slater, J. W., Carney, B. F., 2011. Chitin and chitosan from marine organisms. Chitin, Chitosan, Oligosaccharides and their derivatives. Boca Raton: CRC Press. • Vishwanatha, T., Spoorthi, N., Reena, V., Divyashree B.C., Siddalingeshwara K.G., Karthic, J., Sudipta K.M., 2010. Screening of substrates for protease production from Bacillus lichenformis. International Journal of Engineering Science and Technology(2)11, pp. 6550-6554 • Jung, W.J., Jo, G. H., Kuk, J. H., Kim, K. Y., Park, R. D., 2005. Extraction of chitin from red crab shell waste by cofermentation with Lactobacillus paracasei subsp. tolerans KCTC-3074 and Serratia marcescens FS-3. Applied Microbiology and Biotechnology (71) pp. 234-237. • Jung, W.J., Jo, G. H., Kuk, J. H., Kim, Y. J., Oh, K. T., Park, R. D., 2007. Production of chitin from red crab shell waste by successive fermentation with Lactobacillusparacasei KCTC-3074 and Serratia marcescens FS-3. Carbohydrate Polymers (68) pp. 746-750. • Oh, K. T., Kim, Y. J., Nguyen, V. N., Jung, W.J., Park, R.D. 2007. Demineralization of crab shell waste by Pseudomonas aeruginosa F722. Process Biochemistry. (42) pp. 1069–1074. • Jo, G.H., Park, R.D., Jung, W.J., 2011. Enzymatic production of chitin from crustacean shell waste. Chitin, Chitosan, Oligosaccharides and their derivatives. Boca Raton: CRC Press. • Rao, M. S., Muñoz, J., Stevens, W. F., 2000. Critical factors in chitin production by fermentation of shrimp biowaste. Applied Microbiology and Biotechnology (54) pp. 808-813.

  14. Thank you very much for your attention.

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