Surface Modification of Cellulose Acetate with Cutinase and ...

Surface Modification of Cellulose Acetate with Cutinase and Cutinase Fused to Carbohydrate-binding Modules
20 min each. Between the detergent/alcohol and alcohol/salt steps the fabric was rinsed
under running cold tap water for 5 min.
2.4. Determination of acetic acid concentration in the treatment solutions
Detection of acetic acid in the reaction media was performed with the acetic acid UV
test from Roche (Darmstadt, Germany). Protein was previously precipitated using
perchloric acid according to the manufacture instructions. The samples pH was
neutralized using 1 M potassium hydroxide and the subsequent salts were removed by
centrifugation.
2.5. Quantification of total protein concentration
Total protein in solution was quantified following Bradford methodology (Bradford,
1976) using BSA as standard. All samples were measured at least in duplicate.
2.6. Cellulose acetate fabric staining with a reactive dye
After enzymatic treatment, samples were competitively stained in 50 mM phosphate
buffer pH 8 with 2% o.w.f. (of weight of fabric) Remazol Brilliant Blue R, C.I. 61200,
in duplicate. The staining was performed at 50º C or 60º C, for 90 min at 20 rpm, in
sealed stainless steel beakers of 140 mL in capacity and housed on a lab-scale dyeing
machine (AHIBA Spectradye, from Datacolor International).
After staining, all samples were washed once with 0.25 g L -1 Lutensol AT25 and several
times with distilled water in Rotawash, until no more dye could be detected in the
solution. The washing temperature was 5º C higher than the staining temperature.
The colour measurements (5 for each sample) were carried out with a reflectance
spectrophotometer having a standard illuminant D65 (Spectraflash 600 Plus, from
Datacolor International). The colour strength was evaluated as K/S at the maximum
absorption wavelength (660 nm) which is proportional to the dye concentration in the
samples. The ratio between absorption (K) and scattering (S) is related to reflectance
(R) data by applying Kubelka-Munk’s law at each wavelength (Kuehni, 1997).
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