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This study (purchase required) used, high-performance liquid chromatography, combined with chemometric analysis, to classify buffalo vs cows ricotta based on the profile of water soluble peptides.  The authors then identified specific peptides that could be used as species markers . Both mid-infrared spectroscopy and electrophoresis were also investigated as peptide measurement methods by were found to give insufficient discrimination, with IR overly affected by storage time of the extracts.

The authors created  11 experimental cheese formulations by increasing the proportions of cow whey mixed with buffalo whey. Water-soluble peptides were analysed using mid-infrared spectroscopy, high-performance liquid chromatography and electrophoresis. The data obtained from mid-infrared spectroscopy and high-performance liquid chromatography were statistically processed using principal component analysis, analysis of covariance and multiple linear regression..

High-performance liquid chromatography identified 14 peptide peaks, with three recognized as specific markers for cow whey in adulterated samples. PCA explained 77% of the variance, distinguishing pure and adulterated ricotta. Multiple linear regression modelling of high-performance liquid chromatography data predicted cow whey concentration with a correlation of R = 0.87. High-performance liquid chromatography with chemometrics was effective for detecting buffalo ricotta adulteration.

When applied to 14 commercial samples, the model suggested that nine contained adulteration ranging from 10% to 100% cow whey.

Photo by Conor Brown on Unsplash

In this study (open access) the researchers proposes using a MALDI-ToF and LC-Q-ToF dual approach, following trypsin digestion, as a method to verify fish species.  Trypsin digestion breaks the proteins down into peptides, and they used peptide fingerprints to identify peptides that were unique markers for specific species. The advantage of their approach over DNA methods, and in comparison to MALDI-ToF-MS analysis of undigested proteins, is that it can be applied to complex and heat-processed samples.

The study aimed to differentiate six fish species—carp, mackerel, pike, pollock, salmon and trout. Matrix-assisted laser desorption/ionization–time ff flight mass spectrometry (MALDI-TOF MS) was employed to identify characteristic species-specific m/z values to differentiate raw and cooked fish meat. Additionally, liquid chromatography–electrospray ionization–quadrupole–time tf flight (LC-ESI-Q-TOF) was used to determine specific amino acid sequences in carp and salmon, selected as model species.

Two or more distinct species-specific m/z markers were identified for all six fish species, enabling their differentiation in both raw and processed form. A slightly larger list of distinct markers were found for cooked, compared to raw, fish.  In carp and salmon, hundreds of peptide sequences were detected, leading to the identification of a panel of peptide markers that determine both the fish species and the type of meat processing. The results confirm that mass spectrometry-based proteomic approaches can serve as effective tools for the authentication of fish meat.

The authors conclude that it is possible to use two complementary mass spectrometry techniques for reliable and rapid authentication of fish species. By focusing on peptide-level markers and leveraging accessible tools, they believe that the approach offers a cost-effective and innovative alternative for fish meat authentication.