Application of LC‐HRMS identified marker peptides in an LC‐MS/MS method for detection and quantification of heat‐resistant proteolytic activity in raw milk. (22nd December 2020)

• Record Type: Journal Article
• Authors: Verhegghe, Marijke; De Block, Jan; Heyndrickx, Marc; Van Coillie, Els; Van Poucke, Christof; Duquenne, Barbara
• Is Part Of: International journal of dairy technology. Volume 74:Number 2(2021)
• Page Start: 286
• Page End: 296
• Publication Date: 2020-12-22
• Subjects: Proteolytic activity -- Raw milk -- LC‐HRMS identification -- Marker peptides -- LC‐MS/MS detection -- Pseudomonas; Dairying -- Periodicals; Dairying -- Great Britain -- Periodicals; 637
• DOI: 10.1111/1471-0307.12754
• Languages: English
• ISSNs: 1364-727X

Abstract

Abstract : For the dairy industry, it would be useful to be able to determine the heat‐resistant proteolytic activity of the raw milk upon arrival at the plant, preferably before processing (within 24 h of arrival). This would allow producers to choose the most optimal processing of the raw milk and hence its fate, together with a more accurate prediction of shelf‐life. A liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) method for the detection and quantification of Pseudomonas ‐derived heat‐resistant proteolytic activity in raw milk based on three new marker peptides, identified using liquid chromatography coupled to high‐resolution mass spectrometry (LC‐HRMS), was developed. This method involved a sample preparation where existing peptides are removed from the milk sample, after which the heat‐resistant protease‐containing fraction is mixed with a casein substrate. After incubation, the increase of the peptide markers is determined using an LC‐MS/MS method. The limit of detection (LOD) and limit of quantification (LOQ) of this method were determined using synthetically produced peptides (with an average LOD between 0.1 and 5 ng/mL and an average LOQ between 0.1 and 10 ng/mL).