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A simulated gastrointestinal digestion has been carried out on purified peach lipid transfer protein, one of the main allergens among the population of the Mediterranean area and the major allergen of peach allergic patients. The percentage of intact protein, after extensive digestion, measured by comparison with a non-digestible peptide analogue used as internal standard, was found to be about one-third of the original protein content. The peptides formed in digested fraction were characterized by means of LC/MS. The products of the digestion essentially derived from trypsin action, whereas the protein appeared to be resistant to pepsin and chymotrypsin. The identified peptides could be classified as low molecular weight and high molecular weight peptides. The latter consisted of the full protein, with the disulfide bridges still intact, deprived of the smaller peptides. The different digestion products, including the high and low molecular weight peptides, were purified by LC and assessed, together with the intact protein, by dot-blot analysis with sera of allergic patients, allowing to estimate their potential allergenicity. The intact protein and the high molecular weight peptides were found to be recognized by patients' sera, whereas the small peptides were found to be not reactive.

Secreted pulmonary surfactant phosphatidylcholine (PC) has a complex intra-alveolar metabolism that involves uptake and recycling by alveolar type II epithelial cells, catabolism by alveolar macrophages, and loss up the bronchial tree. We compared the in vivo metabolism of animal-derived poractant alfa (Curosurf) and a synthetic surfactant (CHF5633) in adult male C57BL/6 mice. The mice were dosed intranasally with either surfactant (80 mg/kg body weight) containing universally 13C-labeled dipalmitoyl PC (DPPC) as a tracer. The loss of [U13C]DPPC from bronchoalveolar lavage and lung parenchyma, together with the incorporation of 13C-hydrolysis fragments into new PC molecular species, was monitored by electrospray ionization tandem mass spectrometry. The catabolism of CHF5633 was considerably delayed compared with poractant alfa, the hydrolysis products of which were cleared more rapidly. There was no selective resynthesis of DPPC and, strikingly, acyl remodeling resulted in preferential synthesis of polyunsaturated PC species. In conclusion, both surfactants were metabolized by similar pathways, but the slower catabolism of CHF5633 resulted in longer residence time in the airways and enhanced recycling of its hydrolysis products into new PC species.

Lipid transfer proteins (LTPs) are a family of low molecular mass (7-9 kDa) polypeptides, the members of which share 35-95% sequence homology. These proteins are widely distributed throughout the plant kingdom and are receiving attention for their biochemical characteristics and biological activity. LTPs are indeed studied in different research fields varying from allergy to food technology, and numerous molecules belonging to this class are progressively being identified and investigated. Proteins from pomegranate juice were fractioned by cation exchange chromatography and analyzed by SDS-PAGE. Two proteins were identified as putative LTPs on the basis of their molecular weights and their electrophoretic behaviors under reducing and nonreducing conditions. Finally, proteins were purified and characterized by mass spectrometry. This analysis confirmed that the two polypeptides are LTPs on the basis of an amino acid sequence common to LTPs from other plant sources and cysteine content. The two proteins, named LTP1a and LTP1b, showed similar molecular masses but different immunological profiles when immunodetected with rabbit antibodies specific for Pru p 3 and human IgE from a patient suffering from pomegranate allergy. The demonstration of the existence of two immunologically unrelated LTPs in pomegranate confirms the variability and the complexity of the plant LTP family. This should be taken into account when the role of these proteins as elicitors of allergies to fruits is investigated and could help to explain the contradictory literature data on pomegranate allergy.

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