Response surface methodology to supercritical carbon dioxide extraction of astaxanthin from Haematococcus pluvialis

Experimental design was used to investigate the effect of operating temperature (40-80 °C), operating pressure (30-50 MPa), and extraction time (1-4 h) of supercritical carbon dioxide (SC-CO₂) extraction on astaxanthin yields and the extract antioxidant activity (IC₅₀). The ranges of the factors investigated were 40-80 °C for the operating temperature (X₁), 30-50 MPa for the operating pressure (X₂), and 1-4 h for the extraction time (X₃). The statistical analysis of the experiment indicated that pressure, extraction time, and the interaction between temperature and pressure (X₁X₂) had significant effect on astaxanthin yields. The central composite design showed that polynomial regression models were in good agreement with the experimental results with the coefficients of determination of 0.924 and 0.846 for astaxanthin yield and antioxidant activity, respectively. The optimal condition for astaxanthin yield within the experimental range of the variables studied was at 70 °C, 50 MPa, and 4 h. At this condition, the predicted amount of astaxanthin extracted was 23.04 mg/g (2.3 wt% or 83.78% recovery). For the effect of experimental extraction conditions on antioxidant activity, IC₅₀ was used as an index, which is the concentration that gives a 50% reduction in the absorbance of the ABTS free radical. The analysis of the results showed that the interaction between the operating temperature and operating pressure (X₁X₂) was the only significant factor affecting the extract antioxidant activity. The statistical model gave the minimum point for antioxidant activity at 67 °C, 40.3 MPa, and 1.86 h of extraction, at which the value for 1/IC₅₀ was 0.39 l/mg (or IC₅₀ was 2.57 mg/l).