title: Purification and characterisation of a glutamic acid-containing peptide with calcium-binding capacity from whey protein hydrolysate.
abstract: The bioavailability of dietary ionised calcium is affected by intestinal basic environment. Calcium-binding peptides can form complexes with calcium to improve its absorption and bioavailability. The aim of this study was focused on isolation and characterisation of a calcium-binding peptide from whey protein hydrolysates. Whey protein was hydrolysed using Flavourzyme and Protamex with substrate to enzyme ratio of 25:1 (w/w) at 49 °C for 7 h. The calcium-binding peptide was isolated by DEAE anion-exchange chromatography, Sephadex G-25 gel filtration and reversed phase high-performance liquid chromatography (RP-HPLC). A purified peptide of molecular mass 204 Da with strong calcium binding ability was identified on chromatography/electrospray ionisation (LC/ESI) tandem mass spectrum to be Glu-Gly (EG) after analysis and alignment in database. The calcium binding capacity of EG reached 67·81 μg/mg, and the amount increased by 95% compared with whey protein hydrolysate complex. The UV and infrared spectrometer analysis demonstrated that the principal sites of calcium-binding corresponded to the carboxyl groups and carbonyl groups of glutamic acid. In addition, the amino group and peptide amino are also the related groups in the interaction between EG and calcium ion. Meanwhile, the sequestered calcium percentage experiment has proved that EG-Ca is significantly more stable than CaCl2 in human gastrointestinal tract in vitro. The findings suggest that the purified dipeptide has the potential to be used as ion-binding ingredient in dietary supplements.
authors.affiliation: College of Bioscience and Biotechnology, Fuzhou University,No.2 Xueyuan Road,Fujian,Fuzhou 350002,China.
authors.forename: Shun-Li
authors.initials: SL
authors.lastname: Huang
authors.affiliation: College of Bioscience and Biotechnology, Fuzhou University,No.2 Xueyuan Road,Fujian,Fuzhou 350002,China.
authors.forename: Li-Na
authors.initials: LN
authors.lastname: Zhao
authors.affiliation: College of Bioscience and Biotechnology, Fuzhou University,No.2 Xueyuan Road,Fujian,Fuzhou 350002,China.
authors.forename: Xixi
authors.initials: X
authors.lastname: Cai
authors.affiliation: College of Bioscience and Biotechnology, Fuzhou University,No.2 Xueyuan Road,Fujian,Fuzhou 350002,China.
authors.forename: Shao-Yun
authors.initials: SY
authors.lastname: Wang
authors.affiliation: College of Food Science, Fujian Agriculture and Forestry University,No.15 Shangxiadian Road,Fujian,Fuzhou 350002,China.
authors.forename: Yi-Fan
authors.initials: YF
authors.lastname: Huang
authors.affiliation: College of Bioscience and Biotechnology, Fuzhou University,No.2 Xueyuan Road,Fujian,Fuzhou 350002,China.
authors.forename: Jing
authors.initials: J
authors.lastname: Hong
authors.affiliation: College of Bioscience and Biotechnology, Fuzhou University,No.2 Xueyuan Road,Fujian,Fuzhou 350002,China.
authors.forename: Ping-Fan
authors.initials: PF
authors.lastname: Rao
mesh_terms: D002118:Calcium / Q000378:metabolism* / Q000493:pharmacokinetics; D002122:Calcium Chloride / Q000378:metabolism; D002136:Calcium, Dietary; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification*; D056148:Chromatography, Reverse-Phase; D004151:Dipeptides / Q000737:chemistry / Q000302:isolation & purification*; D018698:Glutamic Acid / Q000032:analysis*; D006801:Humans; D007931:Leucyl Aminopeptidase / Q000378:metabolism; D008894:Milk Proteins / Q000737:chemistry*; D011492:Protein Hydrolysates / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D013055:Spectrophotometry, Infrared; D013056:Spectrophotometry, Ultraviolet; D000067816:Whey Proteins
chemical_list: D002136:Calcium, Dietary; D002352:Carrier Proteins; D004151:Dipeptides; D008894:Milk Proteins; D011492:Protein Hydrolysates; D000067816:Whey Proteins; D018698:Glutamic Acid; D007931:Leucyl Aminopeptidase; D002122:Calcium Chloride; D002118:Calcium
keywords: Whey protein hydrolysis; calcium-binding peptide; separation