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

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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 Materials

Carla Harkin

Dr Wolfram Brück

Dr Catherine Lynch

Dr Denis McCrudden

chibio work package 2 aims
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.
pre treatment overview
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

microbial pre treatment
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.
fermentation process
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)

isolation of novel microorganisms
Isolation of Novel Microorganisms

Isolation from soil samples

Screening for useful properties

Small scale fermentation

Identification

Large scale fermentation

Characterisation

initial screening
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.

to be done
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.
future prospects
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.
references
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.
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