No abstract

In this work, we developed an HPLC method to simultaneously quantify and hence evaluate the stability, distribution, and antioxidant capacity of six isoflavones: genistein, genistin, daidzein, daidzin, glycitin, and biochanin A. Isoflavones have been described as having an important estrogenic activity to treat menopausal symptoms and can reduce postmenopausal bone loss and also participate in the prevention of cardiovascular diseases. These beneficial properties are believed derived from their capacity to act as free-radical scavengers. Isoflavones are formulated in capsules and creams and also can be used as antioxidants in liposomes. HPLC separation was achieved on an Agilent Hypersil ODS C18 column. The mobile phase consisted of 0.02-0.2% orthophosphoric acid in water-acetonitrile with gradient elution. The diode array detector was operated at 260 nm. The hydrophobicity of isoflavones was determined through their distribution in octanol-buffer. These results allowed us to establish a relation between chemical structure, pKa, lipophilicity, and the characteristics of the dispersion medium. Photolysis of hydrogen peroxide was used to measure the HO• scavenging capability of isoflavones. In liposomes, the order of reactivity of the studied compounds was genistein > biochanin A > genistin > daidzein > daidzin > glycitin.

In this work, the relationship between the molecular structure of three flavonoids (kaempferol, quercetin and morin), their relative location in microheterogeneous media (liposomes and erythrocyte membranes) and their reactivity against singlet oxygen was studied. The changes observed in membrane fluidity induced by the presence of these flavonoids and the influence of their lipophilicity/hydrophilicity on the antioxidant activity in lipid membranes were evaluated by means of fluorescent probes such as Laurdan and diphenylhexatriene (DPH). The small differences observed for the value of generalized polarization of Laurdan (GP) curves in function of the concentration of flavonoids, indicate that these three compounds promote similar alterations in liposomes and erythrocyte membranes. In addition, these compounds do not produce changes in fluorescence anisotropy of DPH, discarding their location in deeper regions of the lipid bilayer. The determined chemical reactivity sequence is similar in all the studied media (kaempferol < quercetin < morin). Morin is approximately 10 times more reactive than quercetin and 20 to 30 times greater than kaempferol, depending on the medium.