Monash University
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Deamidated wheat protein-carbohydrate Maillard conjugates : effect of size, location and number of carbohydrate conjugated on emulsion steric stabilization at acidic pH and in ionic environment

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posted on 2017-01-16, 01:36 authored by Wong, Benjamin Tziak Ze
This study investigated the effects of conjugation of deamidated wheat protein (sIWP – 40,375 Da) with carbohydrate (180 – 41,000 Da) on the protein’s solution properties (e.g. pI, aggregation behavior at acidic pH, secondary structure, tryptophan local environment), interfacial properties (e.g. interfacial layer thickness) and the ability to stabilize an oil-in-water (O/W) emulsion at acidic pH and in ionic environments. The protein-carbohydrate conjugates were prepared via the Maillard reaction by dry heating at 60 ºC/75% relative humidity. The carbohydrates used were glucose (180 Da), maltodextrins (900 – 4,300 Da), and dextrans (6,400 – 41,000 Da). Approximately 3 – 4 moles of glucose or low molecular weight (LMW) carbohydrate fraction in the 900 – 4,300 Da maltodextrins (i.e. < 425 Da) were conjugated per mole of sIWP, whereas approximately 1.3 mole or 0.5 mole of dextran (6,400 Da or 41,000 Da) was conjugated per mole of sIWP. The zeta-potential and the circular dichroism spectra of the sIWP/protein-conjugates indicate that conjugation did not change the protein’s pI (~pH 4) or the protein’s secondary structure in solution. sIWP and the protein-conjugates attached with glucose or LMW carbohydrates in the maltodextrins became aggregated in solution when the pH approached the pI. Conversely, protein-conjugates attached with dextrans were aggregated to lower extent in solution than sIWP at its pI due to the conjugated dextran acting as a physical barrier. Dynamic light scattering was used to study the interfacial layer thickness of sIWP/protein-conjugates adsorbed on polystyrene spheres model system. sIWP alone formed a thick protein steric layer of ~18 nm at the interface. Protein-conjugates attached with glucose or LMW carbohydrates in the maltodextrins showed the same layer thickness as sIWP (i.e. ~18 nm), indicating that the attached carbohydrates did not formed an additional carbohydrate steric layer at the interface due to their small sizes. On the other hand, protein-conjugates attached with dextrans (6,400 Da and 41,000 Da) were able to form a thicker interfacial layer by ~3.5 nm and ~5.9 nm respectively than sIWP. Dextranase digestion on the interfacial layer indicates that an additional carbohydrate steric layer was formed by the protein-conjugate attached with 41,000 Da dextran, but was not with 6,400 Da dextran. The observed difference between the attachments of two dextrans with different MW was due to the difference in the sites of conjugation. The smaller 6,400 Da dextran was attached at the C-terminal domain of sIWP, which is the anchoring point to the interface as suggested by the change in the local environment of the tryptophan residues located at the C-terminal domain of sIWP, whereas the larger 41,000 Da dextran was attached at the N-terminal domain of sIWP, as part of the protein diffuse layer. Emulsion stabilized by protein-conjugates attached with LMW carbohydrates in the maltodextrin, or the smaller 6,400 Da dextran became flocculated in acidic pH (pH 4) and in CaCl2 environments (0 – 20 mM), similar to the emulsion stabilized by sIWP. However the emulsion stabilized by the protein-conjugate formed with the larger 41,000 Da dextran was stable in acidic pH (pH 4) and in CaCl2 environments (0 – 20 mM). This is due to the additional carbohydrate steric layer formed by the attached dextran which provided adequate steric stability against emulsion droplets flocculation. This study showed that Maillard conjugation can be used to improve the ability of protein (e.g. sIWP) in stabilizing emulsions at acidic pH and in ionic environments by forming an additional carbohydrate steric layer at the interface. However the formation of an effective carbohydrate steric layer is dependent on the size of the carbohydrate, the location of conjugation and the number of carbohydrate conjugated. [Appendices not included]


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Principal supervisor

Donald McNaughton

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Doctor of Philosophy

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Faculty of Science

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