Investigation of the Proteomes of the Truffles Tuber albidum Pico, T. aestivum, T. indicum, T. magnatum, and T. melanosporum

Truffles of the Tuber species are known as expensive foods, mainly for their distinct aroma and taste. This high price makes them a profitable target of food fraud, e.g., the misdeclaration of cheaper truffle species as expensive ones. While many studies investigated truffles on the metabolomic level or the volatile organic compounds extruded by them, research at the proteome level as a phenotype determining basis is limited. In this study, a bottom-up proteomic approach based on LC-MS/MS measurements in data-independent acquisition mode was performed to analyze the truffle species Tuber aestivum, Tuber albidum Pico, Tuber indicum, Tuber magnatum, and Tuber melanosporum, and a protein atlas of the investigated species was obtained. The yielded proteomic fingerprints are unique for each of the of the five truffle species and can now be used in case of suspected food fraud. First, a comprehensive spectral library containing 9000 proteins and 50,000 peptides was generated by two-dimensional liquid chromatography coupled to tandem mass spectrometry (2D-LC-MS/MS). Then, samples of the truffle species were analyzed in data-independent acquisition (DIA) proteomics mode yielding 2,715 quantified proteins present in all truffle samples. Individual species were clearly distinguishable by principal component analysis (PCA). Quantitative proteome fingerprints were generated from 2,066 ANOVA significant proteins, and side-by-side comparisons of truffles were done by T-tests. A further aim of this study was the annotation of functions for the identified proteins. For Tuber magnatum and Tuber melanosporum conclusive links to their superior aroma were found by enrichment of proteins responsible for sulfur-metabolic processes in comparison with other truffles. The obtained data in this study may serve as a reference library for food analysis laboratories in the future to tackle food fraud by misdeclaration of truffles. Further identified proteins with their corresponding abundance values in the different truffle species may serve as potential protein markers in the establishment of targeted analysis methods. Lastly, the obtained data may serve in the future as a basis for deciphering the biochemistry of truffles more deeply as well, when protein databases of the different truffle species will be more complete.

Geographical based variations in white truffle Tuber magnatum truffle aroma is explained by quantitative differences in key volatile compounds

SummaryThe factors that vary the aroma of Tuber magnatum fruiting bodies are poorly understood. The study determined the headspace aroma composition, sensory aroma profiles, maturity, and microbiome composition from T. magnatum originating from Italy, Croatia, Hungary, and Serbia, and tested if truffle aroma is dependent on provenance and if fruiting body volatiles are explained by maturity and/or microbiome composition.Headspace volatile profiles were determined by gas chromatography-mass spectrometry-olfactometry (GC-MS-O) and aroma of fruiting body extracts were sensorially assessed. Fruiting body maturity were estimated through spore melanisation. Bacterial community was determined using 16S rRNA amplicon sequencing.Main odour active compounds were present in all truffles but varied in concentration. Aroma of truffle extracts were sensorially discriminated by sites. However, volatile profiles of individual fruiting bodies varied more within sites than across geographic area, while maturity level did not play a role. Microbiome composition varied highly and was partially explained by provenance. A few rare bacterial operational taxonomical units associated with select few non-odour active volatile compounds.Specificities of the aroma of T. magnatum truffles are more likely linked to individual properties than provenance. Some constituents of the microbiome may provide biomarkers of provenance and be linked to non-odour active volatiles.

Food Authentication: Truffle (Tuber spp.) Species Differentiation by FT-NIR and Chemometrics

Truffles are certainly the most expensive mushrooms; the price depends primarily on the species and secondly on the origin. Because of the price differences for the truffle species, food fraud is likely to occur, and the visual differentiation is difficult within the group of white and within the group of black truffles. Thus, the aim of this study was to develop a reliable method for the authentication of five commercially relevant truffle species via Fourier transform near-infrared (FT-NIR) spectroscopy as an easy to handle approach combined with chemometrics. NIR-data from 75 freeze-dried fruiting bodies were recorded. Various spectra pre-processing techniques and classification methods were compared and validated using nested cross-validation. For the white truffle species, the most expensive Tuber magnatum could be differentiated with an accuracy of 100% from Tuber borchii. Regarding the black truffle species, the relatively expensive Tuber melanosporum could be distinguished from Tuber aestivum and the Chinese truffles with an accuracy of 99%. Since the most expensive Italian Tuber magnatum is highly prone to fraud, the origin was investigated and Italian T. magnatum truffles could be differentiated from non-Italian T. magnatum truffles by 83%. Our results demonstrate the potential of FT-NIR spectroscopy for the authentication of truffle species.