Isolation & identification of Novel microorganisms for the Treatment

Report
Carla Harkin
Dr Wolfram Brück
Dr Catherine Lynch
Dr Denis McCrudden

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.
Raw material Prep
Optimisation of
chemical chitin
extraction
methods
Identification of
novel microorganisms
Optimisation of
microbial chitin
extraction
methods
Up-scaling
QC
7. QC Analysis

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.
Raw material
+
Inoculum
Shaking Incubator
QC
Harvesting
& drying
Shaking
Incubator
(Jung et al., 2005; Jung et al., 2007a,b; Oh et al., 2007)
Change of Inoculum
Isolation from soil samples
Screening for useful
properties
Small scale fermentation
Identification
Large scale fermentation
Characterisation
 Demineralisation–acid production.

Replicated fermentation method on micro scale.
Fig 1: pH indication after addition of Methyl Red
Indicator. L-r: negative yellow, negative orange,
positive intense red.
 Deproteinisation–protease production.

Casein agar plates.

Colorimetric protease assay to quantify activity
(Vishwanatha et al, 2009).
Fig 2: Produciton of zone of proteolysis
in casein agar.
Property
No. of Organisms
Protease producing
57
Acid producing
45
Protease & Acid producing
16
• 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.
•
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.

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 KCTC3074 and Serratia marcescens FS-3. Applied Microbiology and Biotechnology (71) pp. 234237.

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 Lactobacillus paracasei
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. 808813.
Thank you very much for your attention.

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