Direct determination of methionine sulfoxide in milk proteins by enzyme hydrolysis/high-performance liquid chromatography

• Published in: Journal of Chromatography A Vol. 1157; no. 1; pp. 10 - 16
• Main Authors: Baxter, Jeffrey H., Lai, Chron-Si, Phillips, Rosalyn R., Dowlati, Lobat, Chio, Julie J., Luebbers, Steven T., Dimler, Steven R., Johns, Paul W.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 20.07.2007; Elsevier
• Subjects: amino acid derivatives; Amino Acids - analysis; Biological and medical sciences; Chromatography, High Pressure Liquid - methods; enzymatic hydrolysis; Enzyme hydrolysis; Enzymes - metabolism; Food industries; Fundamental and applied biological sciences. Psychology; HPLC/UV; Hydrolysis; leucyl aminopeptidase; methionine; Methionine - analogs & derivatives; Methionine - analysis; Methionine sulfoxide; Milk and cheese industries. Ice creams; Milk protein; milk proteins; Milk Proteins - chemistry; Oxidation; Peroxide; prolidase; pronase; proteinases; proteolysis; reversed-phase high performance liquid chromatography; Spectrophotometry, Ultraviolet - methods; three-enzyme hydrolysis; Enzyme hydrolysis; Milk protein; Oxidation; HPLC/UV; Peroxide; Methionine sulfoxide; Validation; Chemical analysis; Enzymatic hydrolysis; Methionine; HPLC chromatography; Dairy product; Sulfur containing aminoacid; Ultraviolet detector; Proteolysis; Sulfoxide; Quantitative analysis
• ISSN: 0021-9673
• DOI: 10.1016/j.chroma.2007.04.035

A direct and simultaneous HPLC/UV determination of methionine and methionine sulfoxide in enzyme-hydrolyzed milk proteins is described. Protein hydrolysis is accomplished by a three-enzyme (pronase, leucine aminopeptidase, prolidase) 20-h/37 °C digestion. A gradient elution reversed-phase HPLC system with UV detection at 214 nm and 280 nm is then used to determine the quantitative releases of methionine sulfoxide, methionine, tyrosine, and tryptophan. The ease of methionine oxidation by a wide variety of oxidants, coupled with the quantitative release of both methionine and its sulfoxide by the three-enzyme hydrolysis, renders the approach valuable for identifying oxidized milk proteins. The relatively simple method proved accurate and precise in its application to commercial milk products, finding methionine sulfoxide levels as high as 74% of the total methionine.