Comprehensive two dimensional liquid chromatography (LC × UHPLC): a powerful analytical tool for high resolution profiling of milk peptides after expiration date.

Milk is a rich source of bioactive peptides with several healthy properties. These compounds are released after proteolytic processes that occour in milk after expiration date. This matrix is highly complex and, in order to obtain good separation before the mass spectrometer source and prevent competition for ionization, high peak capacity is essential. A two-dimensional comprehensive LC × UHPLC platform was developed for the separation of peptides after IV week from the expiration date. In this approach we evaluated the employment in the second dimension (D2), of both sub-2 um partially (core-shell) and totally (monodisperse) porous particles columns. High peak capacity values, with respect to a conventional monodimensional approach, were obtained. A ten port-two position high pressure switching valve was used for the online continuous transfer between first and second dimension, on the detection side, both diode array detector (DAD) and a hybrid ion trap-time of flight (IT-TOF) mass spectrometer were employed in series. This platform represents a powerful tool for the identification and profiling of milk peptides, and allows to promote this “waste product” as an important source of bioactive compounds for the development of nutraceutical products and functional milks.