Surface Modification of Cellulose Acetate with Cutinase and ...

Subchapter 3.3
amount of enzyme that hydrolyses casein to produce equivalent colour to 1 μmol of
Tyrosine, per minute, at pH 7.5 and 37 ºC (colour by the Folin & Ciocalteu’s reagent).
The total protein concentration was determined by a modification of the micro Lowry
method (Lowry et al., 1951), using bovine serum albumin as standard and using Sigma
test kit no. P5656.
2.6. FITC linkage to proteins
Enzymes (same Units of both enzymes were used) were linked to FITC (100/1, w/w) in
sodium carbonate buffer pH 8.5. Free FITC was removed using HiTrap desalting
columns (Amersham). Wool samples were treated in this solution at 37 ºC, 20 rpm, for
24 h.
Wool fibres cross-sections were analyzed on a Leica Microsystems DM-5000B
epifluorescence microscope with appropriate filter settings using a 100× oil-immersion
objective. Images were acquired with a Leica DCF350FX digital camera and processed
with LAS AF Leica Microsystems software.
2.7. Pre-treatments performed on wool yarn
Before enzymatic treatment, wool yarns were subject to two different pre-treatment
washings:
Surfactant washing (S): wool was washed with Lutensol ON 30 (non-ionic surfactant)
1 g/l, in a bath ratio 1:20, at pH 9.0 (Na2CO3 0.1 M and NaHCO3 0.1 M buffer), for 30
min, at 40 ºC, on a Rotawash machine. After the washing procedure, the surfactant was
removed from wool first with tap water, followed by distilled water.
Bleaching washing (S + B): After the previous washing, wool was immersed in a bath
with 1% (o.w.f.) H2O2, at pH 9.0 (Na2CO3 0.1 M and NaHCO3 0.1 M buffer), for 1 h at
55 ºC, on Rotawash machine.
2.8. Enzymatic treatment of wool yarns
Enzymatic treatment experiments were performed in plastic boxes each containing 0.5 g
of 23 μm (mean diameter) wool yarns, subjected to the different pre-treatments
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