Surface Modification of Cellulose Acetate with Cutinase and ...
Surface Modification of Cellulose Acetate with Cutinase and ...
Surface Modification of Cellulose Acetate with Cutinase and ...
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<strong>Surface</strong> <strong>Modification</strong> <strong>of</strong> <strong>Cellulose</strong> <strong>Acetate</strong> <strong>with</strong> <strong>Cutinase</strong> <strong>and</strong> <strong>Cutinase</strong> Fused to Carbohydrate-binding Modules<br />
3.2. Progress curves for the modification <strong>of</strong> cellulose di- <strong>and</strong> triacetate fabrics <strong>and</strong><br />
protein adsorption<br />
Samples <strong>of</strong> CDA <strong>and</strong> CTA fabric (1% w/v) were treated <strong>with</strong> 50 U mL -1 <strong>of</strong> cutinase for<br />
different periods <strong>of</strong> incubation. The action <strong>of</strong> cutinase was evaluated by indirectly<br />
measuring the hydroxyl groups formed at the fibres. Since the cellulose acetates used in<br />
this work were insoluble, the enzyme adsorption to the substrate was a prerequisite for<br />
the formation <strong>of</strong> the enzyme-substrate complex. The protein adsorption was indirectly<br />
calculated by the decrease in total protein remaining in the treatment solution. For CDA<br />
(figure. 3A), an equilibrium level <strong>of</strong> relative protein adsorption <strong>of</strong> 45% (3 mg g -1 <strong>of</strong><br />
protein per fabric weight) was reached. For CTA (figure.3B), the equilibrium level <strong>of</strong><br />
protein adsorption was higher, <strong>with</strong> 57% <strong>of</strong> relative protein adsorption (3.5 mg g -1 ). The<br />
hydrophobic character <strong>of</strong> the substrate should not be a problem for cutinase adsorption<br />
since this enzyme is a lipolytic enzyme <strong>and</strong> its natural substrate, cutin, is hydrophobic<br />
(Egmond <strong>and</strong> Vlieg, 2000; Mannesse et al., 1995; Kolattukudy, 2004).<br />
The formation <strong>of</strong> hydroxyl groups at the fibre surface was evaluated by staining the<br />
fabric <strong>with</strong> a cotton reactive dye. The basic principle is the specific reaction between a<br />
vinylsulphonic group from a reactive dye, in this case Remazol Brilliant Blue R, <strong>and</strong> the<br />
hydroxyl group at the fibre surface. The sensitivity is high due to the large molar<br />
absortivities <strong>of</strong> dye molecules. If the cutinase is able to hydrolyse some <strong>of</strong> the acetyl<br />
groups at the surface, then more dye can be chemically linked to the fibre, resulting in<br />
an increase in K/S. The staining ‘titration’ methodology was already reported (Silva et<br />
al., 2005; Matamá et al., 2006, 2007; O’Neill et al., 2007) <strong>and</strong> proved to be a valuable<br />
<strong>and</strong> a very sensitive qualitative method.<br />
In the case <strong>of</strong> CDA fibre, the sensitivity is not as good as for the CTA fibre. The dye has<br />
more affinity for diacetate <strong>and</strong>, as a result, the controls are very coloured while the<br />
triacetate controls are very faint. This is the reason for the observed differences between<br />
the two fibres in the relative increase in the colour strength values (figure 3).<br />
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