Authenticity of Hay Milk vs. Milk from Maize or Grass Silage by Lipid Analysis

Abstract

Hay milk is a traditional dairy product recently launched on the market. It is protected as “traditional specialty guaranteed” (TSG) and subjected to strict regulations. One of the most important restrictions is that the cow’s feed ration must be free from silage. There is the need for analytical methods that can discriminate milk obtained from a feeding regime including silage. This study proposes two analytical approaches to assess the authenticity of hay milk. Hay milk and milk from cows fed either with maize or grass silage were analyzed by targeted GC-MS for cyclopropane fatty acid (dihydrosterculic acid, DHSA) detection, since this fatty acid is strictly related to the bacterial strains found in silage, and by HPLC-HRMS. The presence of DHSA was correlated to the presence of maize silage in the feed, whereas it was ambiguous with grass silage. HPLC-HRMS analysis resulted in the identification of 14 triacylglycerol biomarkers in milk. With the use of these biomarkers and multivariate statistical analysis, we were able to predict the use of maize and grass silage in the cow’s diet with 100% recognition. Our findings suggest that the use of analytical approaches based on HRMS is a viable authentication method for hay milk.

Improved Detection and Quantification of Cyclopropane Fatty Acids via Homonuclear Decoupling Double Irradiation NMR Methods

“…Here, we provide a quantitative description of how the introduction of a simple decoupling scheme not only results in an increased SNR on 1 H 1D NMR experiments but also allows an accurate quantification of the analyte. We tested our method on the detection and quantification of dihydrosterculic acid (DHSA), as one of cyclopropane fatty acids (CPFAs) that has been identified as powerful molecular marker in establishing the authenticity of Protected Denomination of Origin (PDO) cheeses such as Parmigiano Reggiano. − Hence, the CPFA methodology on the authentication of Hay Milk is a current interest as its specification of production prohibits the use of silage on cows feeding . Briefly, CPFAs, mainly lactobacillic and dihydrosterculic acids, are uncommon fatty acids containing a three-carbon ring located at different positions of the fatty acid chain (Figure A).…”

“…They are known to be components of bacterial membranes in both Gram-positive and Gram-negative as a response to stress conditions . So far, CPFAs have been identified in a wide variety of animal-derived foods (i.e., dairy and meat products) and their presence correlate with the presence of maize silage in the forage used in livestock farming . Recently, the presence of CPFAs in milk and dairy products has been linked to the field of food authentication; in fact, products such as Parmigiano Reggiano cheese (PDO) and Hay Milk, where the usage of silage is forbidden (EU 1151/12, EU 2016/304), should be free of any CPFAs .…”

“… 18 − 22 Hence, the CPFA methodology on the authentication of Hay Milk is a current interest as its specification of production prohibits the use of silage on cows feeding. 23 Briefly, CPFAs, mainly lactobacillic and dihydrosterculic acids, are uncommon fatty acids containing a three-carbon ring located at different positions of the fatty acid chain ( Figure 1 A). They are known to be components of bacterial membranes in both Gram-positive and Gram-negative as a response to stress conditions.…”

Techniques for Food Authentication: Trends and Emerging Approaches

“…Gas chromatography with MS detection was used as a targeted approach to detect the cyclopropane fatty acid (dihydrosterculic acid, DHSA), a marker of the bacterial strains found in silage since hay milk should be obtained from a cow’s feed ration free from silage. The detection of DHSA could be related to the presence of maize silage in feed, though it was ambiguous in the case of grass silage [ 13 ]. High-performance liquid chromatography (HPLC) coupled to high-resolution mass spectrometry (HRMS) was then used as an untargeted approach to characterize the lipidic profile, resulting in the identification of 14 triacylglycerol biomarkers in milk.…”

“…High-performance liquid chromatography (HPLC) coupled to high-resolution mass spectrometry (HRMS) was then used as an untargeted approach to characterize the lipidic profile, resulting in the identification of 14 triacylglycerol biomarkers in milk. The biomarker profiles, combined with a multivariate analysis, allowed to predict the use of maize and grass silage in a cow’s diet with 100% recognition [ 13 ]. Comprehensive two-dimensional gas chromatography (GC×GC) is an advanced approach with high potential to characterize complex volatile fractions of foods, including enantiomeric recognition for authentication purposes, owing to its improved separation capacity, increased number of identified compounds, structured chromatograms, and significant signal enhancement [ 14 ].…”