Extraction of Fish visceral protein hydrolysate (FVPH) form Pangasius viscera, has good functional and antioxidant properties. Hydrolysate gave the smaller size of peptides. Peptides were extracted in the present investigation from fish viscera with chemical and enzyme treatments. The aim of this study to extract hydrolysate from viscera by chemical and enzymatic treatment and check the antioxidant properties. FVPH extracted with different treatments i.e. by using papain, pepsin, acid and alkali and it was observed that the treatment with pepsin demonstrated the better results followed by papain, alkali and acid treatments in physicochemical properties, but chemical extracted hydrolysate showed better anti oxodant properties. Hydrolysate extracted from Pangasius viscera using freeze-dried technology and its influence on the yield, proximate composition, color and antioxidant properties of hydrolysate. Pepsin significantly (p < 0.05) had a higher degree of hydrolysis (64.08±0.73%), protein (75.06±0.89%), L* (68.39±0.39). Acid-extracted hydrolysate showed better DPPH and reducing scavenging activity whereas alkali extracted showed better ABTS. SEM microstructure showed smaller size of microparticle at 3,000X magnification. Microspores were present in the hydrolysate which indicated encapsulation properties. The content of bioactive peptides could make the Pangasius visceral hydrolysate prepared with chemical suitable for protein extraction and use for incorporation in food for antioxidant properties.