Application of at-line two-dimensional liquid chromatography-mass spectrometry for identification of small hydrophilic angiotensin I-inhibiting peptides in milk hydrolysates.
A two-dimensional chromatographic method with mass spectrometric detection has been developed for identification of small, hydrophilic angiotensin I-inhibiting peptides in enzymatically hydrolysed milk proteins. The method involves the further separation of the poorly retained hydrophilic fraction from a standard C(18) reversed-phase column on a hydrophilic interaction liquid chromatography (HILIC) column. The latter column is specifically designed for the separation of hydrophilic compounds. Narrow fractions collected from the HILIC column were analysed for their angiotensin I-converting enzyme (ACE) inhibiting potential in an at-line assay. Fractions showing significant inhibition of ACE were analysed by LC-MS for structure elucidation. With this method the main peptides responsible for ACE-inhibition in the hydrophilic part of a milk hydrolysate could be determined. The ACE-inhibiting peptides RP, AP, VK, EK, and EW explained more than 85% of ACE-inhibition by the hydrophilic fraction.
van Platerink
Chris J
CJ
Advanced Measurement and Imaging, Unilever Food and Health Research Institute, P.O. Box 114, 3130 AC, Vlaardingen, The Netherlands. chris-van.platerink@unilever.com
Janssen
Hans-Gerd M
HG
Haverkamp
Johan
J
D000803:Angiotensin I / Q000037:antagonists & inhibitors*; D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods; D006868:Hydrolysis; D013058:Mass Spectrometry / Q000379:methods; D008892:Milk / Q000737:chemistry*; D008894:Milk Proteins / Q000737:chemistry; D010455:Peptides / Q000032:analysis*; D014867:Water / Q000737:chemistry
D008894:Milk Proteins; D010455:Peptides; D014867:Water; D000803:Angiotensin I