Urea complexation combined with rapid preparative reversed-phase liquid chromatography to separate α-linolenic acid from perilla seed oil: Purity, yield, and oxidation stability

Perilla seed oil (PSO) is a major source of α-linolenic acid (ALA). In order to better separate ALA from PSO and to improve its purity and yield, a purification method for ALA that combines urea complexation (UC) with rapid preparative reversed-phase liquid chromatography (RP-RPLC) has been studied. Within the selected ranges of conditions, four one-factor experiments were found to have significant effects on the purity and yield of ALA from PSO. A response surface methodology (RSM) has been applied to optimize UC for separating and purifying ALA from PSO, which yielded optimal parameters of 95% ethanol/urea (w/w) = 5:1, urea/esterified fatty acids (E-FAs) (w/w) = 2:1, complexation time 16.3 h, and complexation temperature 17 °C. The optimal purity and optimal yield of the one-time purification ALA (OTP-ALA) obtained by UC thereby reached 86.977 ± 0.082% and 34.696 ± 0.948 %, respectively. Two-time purification conditions of OTP-ALA by RP-RPLC were also determined, and the purity of the two-time purification ALA (TTP-ALA) was increased to 99.200 ± 0.061%, corresponding to a yield of 53.888 ± 0.904 %. The effect of different added amounts of α-tocopherol on the oxidation stability of TTP-ALA has also been studied. Accelerated oxidation tests showed that the addition of α-tocopherol effectively inhibited the oxidation of TTP-ALA by reducing the peroxide value (PV).