Abstract
Novel thin and smooth deuterated cellulose films were synthesised to visualize adsorbed bio-macromolecules using contrast variation neutron reflectivity (NR) measurements. Incorporation of varying degrees of deuteration into cellulose was achieved by growing Gluconacetobacter xylinus in deuterated glycerol as carbon source dissolved in growth media containing D2O. The derivative of deuterated cellulose was prepared by trimethylsilylation(TMS) in ionic liquid(1-butyl-3-methylimidazolium chloride). The TMS derivative was dissolved in toluene for thin film preparation by spin-coating. The resulting film was regenerated into deuterated cellulose by exposure to acidic vapour. A common enzyme, horseradish peroxidase (HRP), was adsorbed from solution onto the deuterated cellulose films and visualized by NR. The scattering length density contrast of the deuterated cellulose enabled accurate visualization and quantification of the adsorbed HRP, which would have been impossible to achieve with non-deuterated cellulose. The procedure described enables preparing deuterated cellulose films that allows differentiation of cellulose and non-deuterated bio-macromolecules using NR.
Highlights
To date, cellulose model films have been prepared from either nanocellulose, or cellulose derivatives using two main approaches: Langmuir-Blodgett (LB) deposition[14] and spin coating[15,16]
It is a three-step process: the first step consists of derivatizing deuterated bacterial cellulose so that it becomes soluble in a volatile organic solvent; the second step is to spin coat a thin, smooth and uniform DBC derivative film onto polished silicon block and the third step involves regenerating the DBC derivative film into a film of deuterated cellulose
The measurements were performed in the presence of D2O and the results show an adsorbed volume fraction for horseradish peroxidase (HRP) of about 20%
Summary
Cellulose model films have been prepared from either nanocellulose, or cellulose derivatives using two main approaches: Langmuir-Blodgett (LB) deposition[14] and spin coating[15,16]. Native or functionalized films of regenerated and nanofibrillar cellulose have been used to study the adsorption and chemical conjugation of antibodies, and the dynamic adsorption and other features of the adsorbed antibody have been characterized by quartz crystal microbalance (QCM) with dissipation, atomic force microscopy (AFM) and surface plasmon resonance (SPR)[8,17] Factors such as cationicity, ionic strength and pH of the solution in which the macromolecule was dissolved were reported to control antibody/enzyme adsorption[8,9,18]. While SPR and QCM offer quantification of the adsorption kinetics in real time, atomic force microscopy (AFM), ellipsometry[16] and reflectometry (X-ray and neutron) have emerged as powerful techniques to probe the morphology of biomolecules at liquid-solid interfaces. It is a three-step process: the first step consists of derivatizing deuterated bacterial cellulose so that it becomes soluble in a volatile organic solvent; the second step is to spin coat a thin, smooth and uniform DBC derivative film onto polished silicon block and the third step involves regenerating the DBC derivative film into a film of deuterated cellulose
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