Isolation, Purification, and Identification of Antioxidant Peptides from Wastewater of Cheese Making Industry

Rushikesh Pol1

Alisha Rahaman1

Navnath Kumbhar2

Anupama Pable1

Gunderao Kathwate3

Vitthal T. Barvkar4

Umesh U. Jadhav1,Email

1Department of Microbiology, Savitribai Phule Pune University, Pune, Maharashtra India
2Department of Biochemistry, Savitribai Phule Pune University, Pune, Maharashtra India
3Department of Biotechnology, Shivaji University, Pune, Maharashtra India
4Department of Botany, Savitribai Phule Pune University, Pune, Maharashtra, India

Abstract

The present study describes the isolation and purification of antioxidant peptides. The cheese-making industry wastewater was collected to extract the proteins present in it. These proteins were used as substrates for the generation of antioxidant peptides. The B. subtilis NCIM 2724 was used for the digestion of these proteins. The digest was tested for DPPH radical scavenging activity. It showed 56.24% free radical scavenging activity. The antioxidant peptides were purified from this digest using the 3 kDa membrane filter, Sephadex G-25, and reverse-phase high-pressure liquid chromatography. The liquid chromatography-mass spectrometry analysis of the purified peptide sample suggested the presence of 3-4 amino acid-containing peptide with a molecular mass of 410.21 Da. The LC-MS analysis provided 10 possible sequences for peptide. Further screening of these sequences using a peptide ranker tool resulted in two sequences Proline- Proline- Proline-Threonine (PPPT), Threonine-Proline- Proline- Proline (TPPP) having a good bioactivity score. NCBI database search showed the presence of peptide sequence in signal transducer and activator of transcription 5A [Bos taurus]. Molecular docking studies of the peptides with the DPPH radical showed hydrogen and hydrophobic Pi-Alkyl bond formation. These results suggest that cheese-making industry wastewater can be useful resource for the generation of antioxidant peptides. 

Isolation, Purification, and Identification of Antioxidant Peptides from Wastewater of Cheese Making Industry