The Italian white truffle (Tuber magnatum Pico) is worldwide recognized as one of the most expensive edible mushrooms. Its chemical characterization has not yet been properly investigated in scientific literature. The aim of this study was to explore, for the first time, the truffle polar metabolic profile using an untargeted approach through UHPLC-ESI-QTOF-MS data-dependent acquisition. The total features list was filtered and molecules were annotated according to the MS spectra correspondence score with a threshold of 60 %. This led to a total of 412 annotated compounds mainly referred to amino acids, B-vitamins, carbohydrates and fatty acids derivatives. A targeted analysis was applied to four annotated molecules never detected in truffle before, being riboflavin, azelaic, 2-isopropylmalic and 3-isopropylmalic acids. Some constituents of the extracts were semi-quantitatively analysed by 1H NMR spectroscopy. Results place a first brick in white truffle characterization, paving the way towards authenticity biomarkers detection useful in frauds prevention.