Comprehensive Analysis of Lutein and Loroxanthin in Scenedesmus obliquus: From Quantification to Isolation.

Carotenoids are hydrophobic pigments produced exclusively by plants, fungi, and specific microbes. Microalgae are well suited for the production of valuable carotenoids due to their rapid growth, efficient isoprenoid production pathway, and ability to store these compounds within their cells. The possible markets for bio-products range from feed additives in aquaculture and agriculture to pharmaceutical uses. The production of carotenoids in microalgae is affected by several environmental conditions, which can be utilized to enhance productivity. The current study focused on optimizing the extraction parameters (time, temperature, and extraction number) to maximize the yield of carotenoids. Additionally, the impact of various nitrogen sources (ammonia, nitrate, nitrite, and urea) on the production of lutein and loroxanthin in Scenedesmus obliquus was examined. To isolate the carotenoids, 0.20 g of biomass was added to 0.20 g of CaCO3 and 10.0 mL of ethanol solution containing 0.01% (w/v) pyrogallol. Subsequently, the extraction was performed using an ultrasonic bath for a duration of 10 min at a temperature of 30 °C. This was followed by a four-hour saponification process using a 10% methanolic KOH solution. The concentration of lutein and loroxanthin was measured using HPLC-DAD at 446 nm, with a flow rate of 1.0 mL/min using a Waters YMC C30 Carotenoid column (4.6 × 250 mm, 5 µm). The confirmation of carotenoids after their isolation using preparative chromatography was achieved using liquid chromatography-tandem mass spectrometry (LC-MS/MS) with an atmospheric pressure chemical ionization (APCI) probe and UV-vis spectroscopy. In summary, S. obliquus shows significant promise for the large-scale extraction of lutein and loroxanthin. The findings of this study provide strong support for the application of this technology to other species.

Manipulation in Culture Conditions of Nanofrustulum shiloi for Enhanced Fucoxanthin Production and Isolation by Preparative Chromatography.

Microalgae produce a variety of high-value chemicals including carotenoids. Fucoxanthin is also a carotenoid that has many physiological functions and biological properties. For this reason, the cost-effective production of fucoxanthin at an industrial scale has gained significant attention. In the proposed study, fucoxanthin production was aimed to be increased by altering the culture conditions of N. shiloi. The effect of light intensity aeration rate, different nitrogen sources, and oxidative stress on the biomass and fucoxanthin productivity have been discussed. Based on these results, the fucoxanthin increased to 97.45 ± 2.64 mg/g by adjusting the light intensity to 50 µmol/m2s, and aeration rate at 5 L/min using oxidative stress through the addition of 0.1 mM H2O2 and 0.1 mM NaOCl to the culture medium. Fucoxanthin was then purified with preparative HPLC using C30 carotenoid column (10 mm × 250 mm, 5 µm). After the purification procedure, Liquid chromatography tandem mass spectrometry (LC-MS/MS) and UV-vis spectroscopy were employed for the confirmation of fucoxanthin. This study presented a protocol for obtaining and purifying considerable amounts of biomass and fucoxanthin from diatom by manipulating culture conditions. With the developed methodology, N. shiloi could be evaluated as a promising source of fucoxanthin at the industrial scale for food, feed, cosmetic, and pharmaceutical industries.

Induction of lutein production in Scenedesmus obliquus under different culture conditions prior to its semipreparative isolation.

Microalgae with their improved growth rates and accumulation of high-value-added products make their commercial production attractive. Among them, lutein, which is a carotenoid, plays a very important role due to its various applications in the food and pharmaceutical industry. Induction of its biosynthesis can be triggered by various stress conditions like light. In this study, three different light intensities (50,150 and 300 µmol photons/m2s) and aeration rates (1, 3, and 5 L/min) were utilized to induce the lutein biosynthesis and biomass productivity in Scenedesmus obliquus. Lutein was isolated by preparative chromatography using a semiprep C30 column (10 × 250 mm, 5µm) and its confirmation was made by LC-MS/MS. According to the results, Scenedesmus obliquus synthesized the maximum lutein (8.01 ± 0.1 mg/g) with biomass productivity of 1.698 g/L at 150 µmol photons/m2s light intensity using 3 L/min as aeration rate. To the best of the authors' knowledge, this was the first study that the lutein was isolated by preparative chromatography using semiprep C30 carotenoid column with a simple and rapid separation, which can be used as a reference methodology for the isolation of other carotenoids. Scenedesmus obliquus can be an important alternative source for commercial production of lutein, as it is indicated from the results of this study.