The use of complementary medicine has recently increased in an attempt to find effective alternative therapies that reduce the adverse effects of drugs. Pomegranate (Punica granatum L.) by-products, such as seeds, is a rich source of phytochemicals with a high antioxidant activity, thus possessing health benefits. For the identification and quantification of the pomegranate seeds chemical compounds, particular attention has been drawn to the latest developments in the HPLC coupling with electrospray ionization (ESI) MS/MS detection. In fact, a wide range of phytochemicals including phenolic acid, anthocyanins, flavonoids, hydrolysable tannins and other polyphenols were characterized. Furthermore, an exhaustive review of the scientific literature on pomegranate seeds on biomedicine and pharmacotherapy was carried out. Indeed, both in vitro and in vivo studies have demonstrated how pomegranate seeds possess antioxidant, anti- cardiovascular diseases, anti-osteoporosis, antidiabetic, anti-inflammatory and anticancer activities. The present review describes a recent tendency in research focusing on the chemical and biomedical features of the pomegranate seeds to value them as natural additives or active compounds for first-order diseases.