title: In vitro antithrombotic activities of peanut protein hydrolysates.
abstract: The antithrombotic activities of peanut protein hydrolysates were investigated using a microplates assay. When peanut proteins were hydrolyzed to a limited extent by various enzymes, their thrombin inhibitory abilities were significantly enhanced. However, the resultant hydrolysates showed significantly different activities even at the same degrees of hydrolysis. The hydrolysates generated by Alcalase 2.4L displayed the best antithrombotic activities and the hydrolysis process was further optimized by response surface methodology. The antithrombotic activities were increased to 86% based on a protein concentration of 50mg/ml under the optimal conditions: pH 8.5, enzyme concentration of 5000IU/g of peanut proteins, and 2h hydrolysis time at 50°C. The Alcalase 2.4L crude hydrolysates were then fractionated successively by preparative and semi-preparative reverse-phase high-performance liquid chromatography (RP-HPLC). The peptide fraction collected inhibited thrombin-catalyzed coagulation of fibrinogen completely at a concentration of 0.4mg/ml, with an antithrombotic activity close to that of heparin at quite a low concentration (0.2mg/ml). This peptide fraction was further analyzed by online reverse-phase ultra-performance liquid chromatography (RP-UPLC) coupled to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), and three new peptides were identified as Ser-Trp-Ala-Gln-Leu, Gly-Asn-His-Glu-Ala-Gly-Glu and Cys-Phe-Asn-Glu-Tyr-Glu, respectively. This research provided an effective way to produce antithrombotic peptides from peanut proteins, and also helped to elucidate the structure-function relationships of peanut peptides.
authors.affiliation: School of Food Science and Technology, Henan University of Technology, Zhengzhou 450001, Henan Province, People's Republic of China. Electronic address: shaobingzhang@126.com.
authors.forename: Shao Bing
authors.initials: SB
authors.lastname: Zhang
mesh_terms: D010367:Arachis / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D005343:Fibrinolytic Agents / Q000737:chemistry / Q000494:pharmacology*; D006868:Hydrolysis; D010455:Peptides / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry / Q000494:pharmacology*; D011492:Protein Hydrolysates / Q000737:chemistry / Q000494:pharmacology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013381:Subtilisins / Q000378:metabolism; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D005343:Fibrinolytic Agents; D010455:Peptides; D010940:Plant Proteins; D011492:Protein Hydrolysates; D013381:Subtilisins; D013917:Thrombin
keywords: Antithrombotic activities; Enzymatic hydrolysis; Peanut; Protein hydrolysates