The sodium hydroxide concentration was
varied from 0.5 to 5 M, the reaction time ranged from 10 to 400 min
and the temperature was varied from 20 to 100 C. At the end of this
process the material was filtrated, washed and dried, as previously
described in the demineralization process. In order to evaluate the
extent of deproteinization, the protein concentration in the
supernatant was determined according to Biuret’s method (Fine,
1935); It remains the most widely used method for protein
determination.