Phycoerythrin (red pigment) (PE) is a naturally occurring colorant with excellent uorescent and antioxidant properties that has a wide range of applications. More studies are needed, however, to improve the puri cation yield and chemical properties of PE. This study aimed to compare the use of different puri cation methods, which are activated charcoal (AC), ammonium sulfate precipitation (ASP), or anion-exchange chromatography (IE) alone and a combination of AC, ASP, and IE, for PE extraction from the Caspian Sea red macroalgae (Osmundea caspica). Response surface methodology (RSM) was employed for preliminary puri cation, with the independent variables being activated charcoal content (0.1-1% w/v) and stirring time (2-10 min). With an AC content of 0.4% and a stirring time of 2 min, PE purity and concentration were optimized. The predicted values from the equations agreed well with the experimental values, demonstrating the model's robustness. A three-step increase in the ionic strength of IE was also evaluated at three different treatments (PECE, PECE + AC + IE, and ASP + IE). The highest PE purity and lowest Fluorescence intensity (FI) and Antioxidant activity (AA) were obtained in the fraction of AE-200 (second fraction) with an index of 2.8 and a recovery yield of 48% in ASP + IE, whereas an index purity of 2 and the highest recovery yield of 67% were obtained using PECE + AC + IE.

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