Biodeuterated microbial chitosan for characterisation by neutron scattering and development of new biocompatible materials

Chitosan is one of the most abundant natural polysaccharides on earth and has found a wide range of applications in biomedical and environmental fields. The most common source of commercially available chitosan is produced through the deacetylation of chitin from crustacean (mostly shrimp) shells. However, chitosan (along with chitin) is also a major component of the cell walls of a range of fungi, which can be grown and processed under controlled conditions to influence the degree of deacetylation and molecular weight of the extracted chitosan. We have selected the single cell yeast Pichia pastoris, which can also be used for recombinant expression of biodeuterated proteins. P. pastoris was cultivated in a bioreactor using deuterated methanol (CD{sub 3}OD,) as sole carbon source in the growth medium and heavy water (D{sub 2}0) as the solvent. NMR and mass spectrometry analysis demonstrated the complete deuteration of the non-exchangeable protons in the extracted chitosan molecule. Further tuning of the level of deuteration may be achieved by changing the H/D content in the growth medium. Apart from limited reports of partially deuterated chitosan through functional groups or exchangeable protons using deuterated solvent, there are no examples of biosynthesised deuterated chitosan in the literature. This capability, developed at the National Deuteration Facility, ANSTO, provides a range of possibilities for characterising chitosan materials using isotope-sensitive techniques.