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HPLC-MS applications in the agrifood sector are among the fastest developing fields in science and industry. The present tutorial mini-review briefly describes this analytical methodology: HPLC, UHPLC, nano-HPLC on one hand, mass spectrometry (MS) and tandem mass spectrometry (MS/MS) on the other hand. Analytical results are grouped together based on the type of chemicals analyzed (lipids, carbohydrates, glycoproteins, vitamins, flavonoids, mycotoxins, pesticides, allergens and food additives). Results are also shown for various types of food (ham, cheese, milk, cereals, olive oil and wines). Although it is not an exhaustive list, it illustrates the main current directions of applications. Finally, one of the most important features, the characterization of food quality (including problems of authentication and adulteration) is discussed, together with a future outlook on future directions., (Copyright © 2012 Elsevier B.V. All rights reserved.)

An evaluation of the potential allergenicity of different tomato ecotypes is reported. Twelve tomato ecotypes were assessed through a proteomic approach, using pools of sera of allergic patients from two different regions (Emilia Romagna in Northern Italy and Campania in Southern Italy), in order to identify the major allergens and evaluate differences in IgE binding properties of the tomato cultivars. Pooled sera of allergic people from Emilia Romagna showed as the main allergen a suberization-associated anionic peroxidase, whereas pooled sera of allergic patients from Campania were mostly reactive to profilin. The two proteins were identified through a proteomic approach based on the use of high-resolution mass spectrometric techniques. Quite interestingly, in some cases, several ecotypes showed a less reactivity toward patients' sera than other, potentially indicating the possibility to identify ipoallergenic varieties. Anyway, the allergenic pattern response to tomatoes was serum-specific, indicating that the allergenic properties of different tomato ecotypes are defined by the specific proteins to which the patient is sensitized, a strong indication that ipoallergenicity of the different ecotypes is possible, but mostly related to the individual susceptibility. [ABSTRACT FROM AUTHOR]

Abstract: For the first time, the identification of N-γ-glutamyl-, N-pyroglutamyl- and N-lactoyl-amino acids in Parmigiano-Reggiano cheese is reported. Their structures were assigned by electrospray mass spectrometric analysis and demonstrated for several compounds by the synthesis of authentic specimens and sample spiking. The comparison with the synthesised compounds also allowed the determination of their absolute stereochemistry (l,l), which indicated a common enzymatic origin. All these molecules were shown to accumulate in Parmigiano-Reggiano during ripening, probably because of a resistance to proteolysis due to their unusual structures. Thus, these compounds can be used as markers of ripening time and may be involved in the development of the pleasant flavour of aged cheeses. Their presence in other cheeses was also demonstrated. [Copyright &y& Elsevier]

Abstract: The possibility of using short peptides in cheeses, derived from the proteolysis of milk caseins, as molecular markers for the origin of the milk (cows'' or sheep) in fresh sheep milk cheeses has been explored. By using a liquid chromatography/mass spectrometry (LC/MS) technique, two peptides generated during the proteolytic process (fragments 1-14 and 1-23 from αS1 casein), which are slightly different in sheep and cows, were detected separately. The method easily allowed for the detection of casein-derived peptides from cows'' caseins even when cows'' milk was present at very low concentrations, thus revealing the presence of undeclared cows'' milk. This method is a possible simpler alternative to the official electrophoretic methods. [Copyright &y& Elsevier]

Within a project aimed at studying the peptide fraction in dry-cured Parma hams, a peptide was purified by means of reversed phase-high performance liquid chromatography (RP-HPLC) and identified by its molecular mass and amino acid sequence analysis. The peptide showed a very high degree of homology with the N-terminal part of different mammalian pyruvate kinases reported in databases and was accordingly identified as the N-terminal part of swine pyruvate kinase, whose sequence had never been reported before. The peptide was determined quantitatively by comparison with a suitable internal standard (Phe-Phe) in commercial ham samples with different age degrees. The peptide was found to be ubiquitous in Parma ham and its amount to increase during ageing even if a large variability was found within each assayed maturing time. The correlation found with the ham proteolysis degree (P<0.01) suggests that this peptide is related to the endopeptidasic activity. The peptide did not show a significant relationship with bitterness perception in assayed dry-cured ham, in agreement with the hypothesis that this taste is more related to free amino acids and low molecular weight peptides. [Copyright &y& Elsevier]

Resistance of gluten to gastrointestinal digestion is involved in immune-mediated adverse reactions to wheat, since several peptides produced by the incomplete digestion are able to trigger, in predisposed individuals, the immune response responsible, for instance, of celiac disease (CD) and other adverse reactions. Even if several peptides have been identified, an exhaustive description of the peptidome generated by wheat digestion is lacking. To this end, in the present work, durum wheat proteins were fractionated, digested, and then subjected to various proteomic techniques, including single stage and multiple stage mass spectrometry (MS) (SDS-PAGE, UPLC/ESI-MS, UPLC/ESI-MS/MS, and LTQ-Orbitrap). Based on SDS-PAGE, although proteins were severely degraded after in vitro gastrointestinal digestion, some differences were observed among protein profile of the different digests. Through untargeted UPLC techniques, 227 peptide sequences were identified, with only few sequences shared by the different digests. In particular, 9 gluten peptides involved in CD were identified. Based on target proteomic, the quantification of these peptides revealed significant (p ≤ 0.05) differences among the different extracts. Taken together, all the proteomic tools confirmed that gluten digestion is closely related to the matrix regardless of wheat genotype. [ABSTRACT FROM AUTHOR]

Food frauds have become a very important issue in the field of food quality and safety. The risk of food adulteration is higher in highly processed food and mainly affects high added value foodstuff. The methods currently available to face this issue, PCR and ELISA, are very sensitive and specific, but they have some limitations. In the present work, tandem mass spectrometry is presented as an emerging approach to detect beef and pork meat in very complex and highly processed food matrices, such as Bolognese sauce, both in qualitative than in quantitative way. The detection is achieved using two different marker peptides, specific for beef and pork meat, both deriving from α2-collagen chain. Then, a calibration curve is set up using real sauces made by different percentages of pork and beef meat in a working range from 0 to 100%. The method here developed allows to quantify beef and pork meat in a complex product such as Bolognese sauce. [ABSTRACT FROM AUTHOR]

Techno-functional properties of proteins, such as foam stability, can be affected by the presence of low-molecular-weight surfactants. In order to understand and control the foam properties of such protein–surfactant mixtures, a thorough characterization of foam and interfacial properties needs to be supplemented by a detailed analysis of the structural changes of the protein and possible complexation with the surfactant. In this study, β-lactoglobulin (BLG) was mixed with sodium dodecyl sulphate (SDS) in different molar ratios (MRs). The foam half-life time of BLG-SDS mixtures decreased from that of pure BLG (315 min at MR 0) to 44 min at MR 20, which is close to the half-life of SDS at the respective concentration. With a further increase in the MR, the foam stability of the mixture increased, similar to the stability of SDS, to 250 min at the highest MR (MR 100). The minimum in the foam stability curve was not reflected in the interfacial properties ( Π and E d ). Π decreased and E d increased continuously with increasing MR from values close to those of protein towards values typically found in pure surfactant solutions. The results show no clear correlation between the interfacial and foaming properties. In addition, it was shown by isothermal titration calorimetry and mass spectrometry that SDS molecules bind to the BLG. This leads to the formation of BLG-SDS complexes. These complexes have large influence on the foam properties in the mixture. The combination of analytical methods that were used give insights about protein complexation and the resulting change of foam properties of the mixture. [ABSTRACT FROM AUTHOR]

Abstract: The angiotensin-converting enzyme (ACE) inhibitory activity of water-soluble extracts (WSE) from Asiago cheeses was assayed in two cheese production systems and with different ripening times. The WSE were ultrafiltered through 10 kDa and 3 kDa cut-off membranes to evaluate the ACE inhibitory activity of long and short peptides, respectively. The cheese production systems had no significant effect on the ACE inhibitory activity, whereas 6-month-old cheeses had higher inhibitory potency than the more ripened ones. Moreover, the fraction containing peptides smaller than 3 kDa made a more considerable contribution to ACE inhibitory activity than the fraction smaller than 10 kDa, suggesting an inhibitory effect due to short peptides. The peptidic fraction was characterized using RP-HPLC coupled to mass spectrometric detection. Simulated gastrointestinal digestion was carried out to evaluate the effects of digestive enzymes on the generation of bioactive peptides, but this did not significantly affect the inhibitory activity. [Copyright &y& Elsevier]

• Insects are proposed in Europe as protein sources for feed and food application. • Methods for insect-containing ingredient authentication needed. • Marker peptides were identified for Lesser Mealworm and Black Soldier Fly species. • Three species- specific markers were identified as most suitable for authentication by untargeted and targeted LC/MS methods. • The species-specific marker peptides were detected in insect-enriched (down to 1%) feed formulations. Insects have been proposed as new source of proteins to meet the growing demand connected to the increasing world population. In the EU the inclusion of insect proteins in feed and food is strictly regulated. Hence, analytical methods able to discriminate and identify different insect species in food and feed are a necessity. In this work, a peptidomic approach was applied to determine peptide biomarkers for two edible insect species: lesser mealworm and black soldier fly. Three species specific peptide biomarkers were identified using LC-MS/MS. The two insects were mixed with fish standard feed at different concentrations, to evaluate the feasibility of their use as markers in complex matrices. The detection of marker peptides was confirmed down to 1% insect amount. The data here reported constitutes the first proof of concept for the potential application of the peptide marker approach for the identification and quantification of insect ingredients. [ABSTRACT FROM AUTHOR]