Concentration of PN was determined

Concentration of PN was determined by using the Bradford method [24]. 800 µl of the diluted sample containing 0–10 μg of protein per ml is mixed with 200 μl of Biorad reagent. The mixture was maintained at ambient temperature and in the dark during 10 min. The optical density was determined at 595 nm. The protein concentration in each sample was determined by using a calibration curve prepared with concentrated bovine serum albumin (BSA) solution.