Water buffalo (WB) casein (CN) and curd samples from indigenous Italian and international breeds were examined with the objective of identifying signature peptides that could function as an indicator to determine the origin of their milk products. CN in complex mixtures were digested with trypsin, and peptide fragments were subsequently identified by matrixassisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). The unique presence of a β-CN A variant and an internally deleted αs1-CN (f35-42) variant in international WB milk samples was ascertained by identifying signature tryptic peptides from either dephosphorylated or native CN. Four signature unphosphorylated peptides derived from β- CN A, i.e. (f49-68) Asn68 (2223.6 Da), (f1-28) Ser10 (3169.4 Da), (f1-29) Ser10 (3297.4 Da) and (f33-48) Thr41 (1982 Da) and two from αs1-CN (f35-42) deleted fragments, i.e. (f23-34) Met31 (1415.7 Da) and (f43-58) Val44 (1752.7 Da), were identified. Two signature casein phosphopeptides (CPPs), i.e. β-CN (f1-28) 4P (3489.1 Da) and β-CN (f33-48) 1P (2062.0 Da), were identified in the tryptic hydrolysate of native casein or curd and cheese samples using in-batch hydroxyapatite (HA) chromatography. All these fragments functioned as analytical surrogates of two αs1- and β-casein variants that specifically occur in the milk of international WB breeds. Furthermore, the bovine peptide β-CN (f1-28) 4P had a distinct and lower molecular mass compared with theWB counterpart and functioned as a species-specific marker for all breeds ofWB. Advantages of this analytical approach are that (i) peptides are easier to separate than proteins, (ii) signature peptide probes originating from specific casein variants allow for the targeting of all international WB milk, curd and cheese samples and (iii) bovine and WB casein in mixtures can be simultaneously determined in protected designation of origin (PDO) BMozzarella di Bufala Campana^ cheese. This analytical method enabled the specific detection of international WB and bovine casein with a sensitivity threshold of 2 and 0.78 %, respectively.

Simultaneously tracing the geographical origin and presence of bovine milk in Italian water buffalo Mozzarella cheese using MALDI-TOF data of casein signature peptides / S., Caira; Pinto, Gabriella; Nicolai, MARIA ADALGISA; Chianese, Lina; Addeo, Francesco. - In: ANALYTICAL AND BIOANALYTICAL CHEMISTRY. - ISSN 1618-2650. - 408:20(2016), pp. 5609-5621. [10.1007/s00216-016-9663-0]

Simultaneously tracing the geographical origin and presence of bovine milk in Italian water buffalo Mozzarella cheese using MALDI-TOF data of casein signature peptides

PINTO, GABRIELLA;NICOLAI, MARIA ADALGISA;CHIANESE, LINA;ADDEO, FRANCESCO
2016

Abstract

Water buffalo (WB) casein (CN) and curd samples from indigenous Italian and international breeds were examined with the objective of identifying signature peptides that could function as an indicator to determine the origin of their milk products. CN in complex mixtures were digested with trypsin, and peptide fragments were subsequently identified by matrixassisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). The unique presence of a β-CN A variant and an internally deleted αs1-CN (f35-42) variant in international WB milk samples was ascertained by identifying signature tryptic peptides from either dephosphorylated or native CN. Four signature unphosphorylated peptides derived from β- CN A, i.e. (f49-68) Asn68 (2223.6 Da), (f1-28) Ser10 (3169.4 Da), (f1-29) Ser10 (3297.4 Da) and (f33-48) Thr41 (1982 Da) and two from αs1-CN (f35-42) deleted fragments, i.e. (f23-34) Met31 (1415.7 Da) and (f43-58) Val44 (1752.7 Da), were identified. Two signature casein phosphopeptides (CPPs), i.e. β-CN (f1-28) 4P (3489.1 Da) and β-CN (f33-48) 1P (2062.0 Da), were identified in the tryptic hydrolysate of native casein or curd and cheese samples using in-batch hydroxyapatite (HA) chromatography. All these fragments functioned as analytical surrogates of two αs1- and β-casein variants that specifically occur in the milk of international WB breeds. Furthermore, the bovine peptide β-CN (f1-28) 4P had a distinct and lower molecular mass compared with theWB counterpart and functioned as a species-specific marker for all breeds ofWB. Advantages of this analytical approach are that (i) peptides are easier to separate than proteins, (ii) signature peptide probes originating from specific casein variants allow for the targeting of all international WB milk, curd and cheese samples and (iii) bovine and WB casein in mixtures can be simultaneously determined in protected designation of origin (PDO) BMozzarella di Bufala Campana^ cheese. This analytical method enabled the specific detection of international WB and bovine casein with a sensitivity threshold of 2 and 0.78 %, respectively.
2016
Simultaneously tracing the geographical origin and presence of bovine milk in Italian water buffalo Mozzarella cheese using MALDI-TOF data of casein signature peptides / S., Caira; Pinto, Gabriella; Nicolai, MARIA ADALGISA; Chianese, Lina; Addeo, Francesco. - In: ANALYTICAL AND BIOANALYTICAL CHEMISTRY. - ISSN 1618-2650. - 408:20(2016), pp. 5609-5621. [10.1007/s00216-016-9663-0]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11588/666871
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