Toward milk speciation through the monitoring of casein proteotypic peptides.
The possibility of detecting extraneous milk in singles species cheese-milk has been explored. A mass spectrometry (MS)-based procedure has been developed to detect 'signature peptides', corresponding to the predefined subset of 'proteotypic peptides', as matchless analytical surrogates of the parent caseins. Tryptic digests of skimmed milk samples from four species were analyzed by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS. Amongst the candidate signature peptides that are able to differentiate milks from the four species, the alpha(s1)-casein (CN) f8-22 peptide was selected as a convenient marker for bovine, ovine and water buffalo milk while the f4-22 peptide was selected as a marker for the two caprine alpha(s1)-CN A and B variants, which differ by a Pro(16) (B)->Leu(16) (A) substitution. MALDI analysis of the digest allowed the detection of alpha(s1)-CN f8-22 and caprine alpha(s1)-CN f4-22. The accurate evaluation of caprine milk in a quaternary mixture required the development of a liquid chromatography/electrospray ionization (LC/ESI)-MS procedure. Five synthetic signature peptide analogues, which differed from their natural counterparts by a single amino acid substitution, were used as internal standards to quantify the alpha(s1)-CN, which was chosen as a reference milk protein, from the different species. The limits of detection were 0.5% (1% for caprine) for either the MALDI or the LC/ESI-MS method. The isotopic-label-free quantification of isoform- or variant-specific signature peptides has disclosed a convenient approach for targeting proteins in complex mixtures.
Cuollo
Marina
M
Dipartimento di Scienza degli Alimenti - Università di Napoli 'Federico II', Parco Gussone, 80055 Portici (Napoli), Italy.
Caira
Simonetta
S
Fierro
Olga
O
Pinto
Gabriella
G
Picariello
Gianluca
G
Addeo
Francesco
F
D000595:Amino Acid Sequence; D000818:Animals; D002364:Caseins / Q000737:chemistry*; D002417:Cattle; D006041:Goats; D008892:Milk / Q000737:chemistry*; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D011786:Quality Control; D016415:Sequence Alignment; D012756:Sheep
D002364:Caseins; D010455:Peptides