Purification and identification of three novel antioxidant peptides from Cornu Bubali (water buffalo horn).
Cornu Bubali (water buffalo horn, WBH) has been used in Traditional Chinese Medicine (TCM) for thousands of years. In the present study, three peptides with antioxidant properties were purified from aqueous extract of Cornu Bubali (water buffalo horn, WBH) by consecutive chromatographic methods including gel filtration chromatography, ion-exchange chromatography and high performance liquid chromatography. The sequences of the three peptides were identified to be Gln-Tyr-Asp-Gln-Gly-Val (WBH-1, 708Da), Tyr-Glu-Asp-Cys-Thr-Asp-Cys-Gly-Asn (WBH-2, 1018Da) and Ala-Ala-Asp-Asn-Ala-Asn-Glu-Leu-Phe-Pro-Pro-Asn (WBH-3, 1271Da) by matrix assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-LIFT-TOF/TOF MS). The antioxidant activity of these peptides was tested using 1-diphenyl-2-picrylhydrazyl (DPPH) scavenging assay directly. Methylthiazol tetrazolium (MTT) assay and lactate dehydrogenase (LDH) release assay were also used to evaluate the protection of peptides against hydrogen peroxide (H(2)O(2)) induced injury. The results showed that these peptides could reduce the DPPH radical and protect rat cerebral microvascular endothelial cells (rCMECs) against H(2)O(2)-induced injury, thus demonstrating that these peptides had antioxidant activity. These results suggest that WBH-1, WBH-2 and WBH-3, isolated from the aqueous extract of water buffalo horn are natural antioxidants and may contribute to the efficacy of WBH.
Liu
Rui
R
Jiangsu Key Laboratory for TCM Formulae Research, Nanjing University of Chinese Medicine, Nanjing, China.
Wang
Min
M
Duan
Jin-Ao
JA
Guo
Jian-Ming
JM
Tang
Yu-Ping
YP
D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001713:Biphenyl Compounds / Q000737:chemistry; D002020:Buffaloes; D002460:Cell Line; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008969:Molecular Sequence Data; D018384:Oxidative Stress / Q000187:drug effects; D010455:Peptides / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D010851:Picrates / Q000737:chemistry; D051381:Rats; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000975:Antioxidants; D001713:Biphenyl Compounds; D010455:Peptides; D010851:Picrates; D006861:Hydrogen Peroxide; C004931:1,1-diphenyl-2-picrylhydrazyl; D007770:L-Lactate Dehydrogenase