A VALIDATED LC-MS/MS MULTI-METHOD FOR THE DETERMINATION OF 110 MYCOTOXINS AND PLANT TOXINS IN RAW MILK

Milk is one of the most highly consumed food products in all age groups and forms the basis of dairy products. The quality and safety of milk are significantly influenced by the animal's diet, which is composed of a complex mix of individual feed components (e.g. forages and concentrates). Each raw feed component can carry different contaminants, leading to multi-toxin exposure in the overall animal diet. This multitude of toxins could finally find its way into the milk. The presence of contaminants such as mycotoxins and plant toxins can pose health risks to consumers. Currently, only aflatoxin M1 is regulated by the European Union in raw milk, heat-treated milk and
milk for the manufacture of milk-based products. However, there have been reports on the presence of beauvericin and enniatin B [1, 2], T-2 and HT-2 toxins [2], pyrrolizidine and tropane alkaloids [3] and quinolizidine alkaloids [4] in raw milk samples.
Within the framework of the EU project INTAQT, an LC-MS/MS method for the simultaneous determination of a wide range of mycotoxins and plant toxins (110 toxins) in raw milk has been developed and validated according to the SANTE 11312/2021 v2 guidelines in terms of recovery, precision and limit of quantification (LOQ). The method involved QuEChERS-based extraction and LC-MS/MS analysis using triple quadrupole mass spectrometry. The toxins covered by the method include aflatoxins, trichothecenes (nivalenol and related compounds, T-2 toxin and related compounds, among others), fumonisins, zearalenone and related compounds, beauvericin, enniatins, ochratoxins, Alternaria toxins, ergot alkaloids as well as tropane alkaloids, pyrrolizidine alkaloids, and quinolizidine alkaloids. The method has an LOQ for the regulated aflatoxin M1 of 0.025 μg/kg and showed an excellent
recovery (80-110%) and precision (RSD wR < 10%) for almost 80 toxins. To demonstrate its applicability, the validated LC-MS/MS multi-method was applied to a number of samples from the market and local farms. The results revealed varying levels of mycotoxins and plant toxins. The aim of this work was to investigate the occurrence of several mycotoxins and plant toxins and their co-occurrence in milk. The validated LC-MS/MS multi-method proved to be a reliable and efficient tool for the simultaneous determination of multiple mycotoxins and plant toxins in milk.