# Marine Microalgae for Potential Lutein Production
## 7 Extraction of Lutein from Microalgae and Challenges
or _Scenedesmus_ sp., it is unlikely this methodology would be efficient. [PERSON] et al. [64] described a microwave-assisted binary phase solvent extraction method (MABS) for the recovery of lutein from microalgae. The method was established and optimised to specifically attain the highest lutein recovery from microalgae _Scenedesmus_ sp. biomass, with a total of 11.92 mg g\({}^{-1}\) lutein recovered. The optimal binary phase solvent composition was a 60% potassium hydroxide solution with acetone in the ratio of 0.1 (mL/mL). The highest lutein content was at 55 \({}^{\circ}\)C treatment temperature,36 min extraction time, 0.7 (mg mL\({}^{-1}\)) biomass:solvent ratio, 250 Watt microwave power, and 250 rpm stirring speed. This optimised novel procedure can increase lutein recovery by approximately 130%, along with also shortening the overall extraction time by 3-fold. According to an optimised extraction method of lutein from microalgal biomass conducted by [PERSON] et al. [65], it was concluded that cell disruption was necessary and the use of a bead mill with alumina in a 1:1 (_w/w_) proportion as a disintegrating agent for 5 min was the preferred method amongst other varying treatments tested. Studies conducted by [PERSON] et al. [66] investigated lutein extraction from wet _Chorella vulgaris_ UTEX 265, where various extraction parameters such as sample size, drying method, and cell disruption method were studied and lutein production was monitored throughout the microalgal growth phase. From the analysis of varying solvent performances on lutein extraction using Nile Red as a solvatochromic polarity probe, it was determined that 3:1 (_v/v_) ethanol/hexane was the optimal solvent for lutein extraction, which resulted in 13.03 mg g\({}^{-1}\) lutein. Studies carried out by [PERSON] et al. [67] involved the use of high-speed counter-current chromatography (HSCCC) for the isolation and purification of lutein from microalgae. Analytical HSCCC was used for the preliminary selection of a suitable solvent system composed of n-hexane:ethanol:water (4:3:1, _v/v_), where HSCCC was successfully performed, resulting in lutein yield at 98% purity from 200 mg of crude extract in a one-step separation. The use of supercritical CO\({}_{2}\) fluid is gaining increasing interest in regard to the recovery of pharmaceutical or nutraceutical compounds due to clean extracts and lack of toxicity of CO\({}_{2}\) in comparison to organic solvents. However, because the polarity of free lutein is high and excess amounts of organic solvents are required for lutein extraction from microalgae due to binding of light-harvesting complexes, supercritical fluid extraction (SFE) should not prove efficient. However, according to [PERSON] et al. [68], the addition of polar co-solvents such as methanol or ethanol may increase lutein extraction efficiency. When coupled with a co-solvent, SFE may seem a promising lutein extraction method compared to organic solvent extraction. [PERSON] et al. [69] recommended the use of supercritical CO\({}_{2}\) following a solvent extraction of the carotenoids. The solvent containing the carotenoids was then mixed with supercritical CO\({}_{2}\) and the conditions of pressure and temperature were adjusted to promote the precipitation of lutein. [PERSON] et al. [60] aimed to clarify the influence of temperature and pressure parameters on the supercritical fluid extraction of lutein and \(\beta\)-carotene from a freeze-dried powder of the marine microalga _Scenedesmus almteriensis_. The extracts were analysed by HPLC and empirical correlations were also established. The results determined an optimal pressure of 400 bar and optimal temperature of 60 \({}^{\circ}\)C resulted in a significant yield of pigment extraction. In comparison with the referenced extraction processes used, the results obtained from this study displayed that improved yields were obtained in the extraction of \(\beta\)-carotene, where it was possible to extract 50% of the total of this pigment contained in the microalga studied. [PERSON] et al.