Thesis Title

Isolation and Characterization of Chitinases From Dasypus Novemcinctus, the Nine-Banded Armadillo

Date of Graduation

Summer 1996

Degree

Master of Science in Biology

Department

Biology

Committee Chair

John Steiert

Subject Categories

Biology

Abstract

The ability of armadillos to beak down chitin was investigated. Both indigenous and endogenous chitinases were studied. Armadillos were collected and samples of their gastric and pancreatic tissues were removed. Tissues were lyophilized and stored at -20° C until they could be resuspended in the appropriate buffers. The extract from resuspended tissues was assayed for chitinase activity using a colorimetric procedure which detects N-acetyl-D-glucosamine (NAG), the monomeric unit within chitin. Chitinase activity was present in the gastric tissues only. The pH optimum for chitinase activity from armadillo gastric tissues is 5.0, and the temperature optimum is in the range of 50-60° C. Armadillo chitinase was purified five-fold. A 10% SDS-polyacrylamide gel electrophoresis was done with the samples from the purification procedure to visualize purification. Samples from the small intestine, proximal large intestine, and distal large intestine were aseptically collected from armadillos and used to inoculate colloidal chitin medium. Bacterial isolates were characterized using oxygen requirements, Gram stain reaction, and various other physiological tests. Bacterial isolate 8SI, isolated from the small intestine, produced an extracellular chitinase which has optimal activity at a pH of 5.0 and 40° C. The chitinase from isolate 8SI was purified 95-fold. Using the results from a 10% SDS-polyacrylamide gel, the molecular weight of the bacterial chitinase was estimated to be 51 kDa. Samples of intestinal material found in the small intestine, proximal large intestine, distal large intestine, and feces were used to estimate the most probable numbers of chitinolytic microorganisms in those regions.

Copyright

© Stephanie Ann Smith

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