The final solution was dialyzed against cold distilled water using a dialysis membrane with molecular weight cut-off of 12 kDa for 12 h at 4 C, with a change of solution every 4 h. The solution was then lyophilized using a freeze dryer (ALPHA 2–4; Christ, Harz, Germany). From this point it was referred to as pepsin soluble collagen
(PSC).