Hemocytes Lysate Supernatant Derived Phenoloxidase Activity in the Hemolymph of Giant Freshwater Prawn Macrobrachium rosenbergii (De Man, 1879)

Abstract

The aim of the present study attempted to find the cellular immune response in Macrobrachium rosenbergii by melanization reaction produced by hemocytes lysate supernatant (HLS) phenoloxidase (PO) activity. The substrate affinity of the PO enzyme was determined using different phenolic substrates and it was found that the diphenols and polyphenol were oxidized. Hence, the enzyme was characterized as a catechol oxidase type of PO and 3,4-dihydroxy-DL-phenylalanine (DL-DOPA) showed the highest substrate affinity to the HLS. The optimal enzyme activity was observed at 5 mM DL-DOPA in 10 mM Tris-HCl buffer at a pH of 7.5 at 25° C for 20 min and absorbance at 490 nm. Kinetic characteristics of HLS from the prawn were determined. Determination of optimal conditions of PO activity in the HLS has also been attempted. These results depicted that in the presence of PO and peroxidase inhibitors, phenylthiourea (PTU) and tropolone respectively have a decreased HLS PO activity. The determination of PO activity was also highly activated by trypsin, sodium dodecyl sulphate (6 mg.ml-1) and laminarin (4 mg.ml-1) enzyme expression. We also identified the chemicals causing in vitro inhibition or activation of the enzyme as an HLS of the freshwater prawn having a potent PO activity.