Anhydrous sodium carbonate

Absolute methanol (analytical reagent grade), anhydrous sodium carbonate, and acetonitrile (HPLC gradient grade) were supplied by Fisher Scientific UK; quercetin, kaempferol, and myricetin were purchased from Sigma Chemicals; and potassium dihydrogen orthophosphate and hydrochloric acid (analytical grade) were purchased from British Drug House Ltd., England.

Materials included methanol (Fisher Chemicals, A452-4, UK), ethanol (Fisher Chemicals, A962-4, UK), 2,2-diphenyl-1-picrylhydrazyl (DPPH) (Sigma-Aldrich, D9132, USA), BHA (Sigma-Aldrich, B1253, USA), gallic acid (Sigma-Aldrich, G7384, USA), Folin–Ciocalteu's phenol reagent (Sigma-Aldrich, F9252, USA), anhydrous sodium carbonate (Fisher Chemicals, S263-1, UK), maleic acid (Sigma-Aldrich, M0375, USA), sodium hydroxide (American Chemicals, S290, USA), calcium chloride dihydrate (Fisher Chemicals, C79-500, UK), sodium azide (Sigma-Aldrich, S2002, USA), potassium hydroxide (Fisher Chemicals, S71978, UK), glacial acetic acid (Fisher Chemicals, A38-500, UK), Megazyme resistant starch assay kit (Megazyme International Ireland Ltd, Bray, Ireland), and Pierce™ BCA protein assay kit (Pierce, Rockford, IL, USA).

Acetic acid (CH₃COOH, 99.7%) was obtained from Sigma Aldrich (St. Louis, MO, USA). Acetone (C₃H₆O, 99.5%), sodium bicarbonate (NaHCO₃, 99.7%), sodium hydroxide (NaOH, 98%), and anhydrous sodium carbonate (Na₂CO₃, 99.5%) were obtained from Fisher Scientific (Hampton, NH, USA). A cationizing agent, 3-chloro-2-hydroxypropyltrimethyl ammonium chloride (CHPTAC), also called CR-2000 (C₆H₁₅Cl₂NO, 65%), and Triton X-100 (2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol) (C₁₆H₂₆O₂, 99+%) were obtained from Dow Chemical (Midland, MI, USA). Sodium sulfate anhydrous (Na₂SO₄, 99.5%) was received from Cooper Natural Resources (Fort Worth, TX, USA). The organoselenium compounds, OS-1 (C₆Cl₄N₆Se₂, >99%) and OS-2 (C₆H₂Cl₂N₆Se₄, >99%), were obtained from Attach Chem (Lubbock, TX USA) and used as received. Deionized (DI) water was obtained from AquaOne (Amarillo, TX, USA) and was used for all purposes.

Commercial C. vulgaris biomass was provided in powder form by Go Superfoods Ltd. (Sheffield, UK). C. vulgaris was cultivated in natural water open ponds in South China, harvested with mesh screens, pretreated through milling, and spray dried. All referred stages are in accordance with strict regulations for human consumption-intended products. Chloroform, ethyl acetate, phenol crystals, orthophosphoric acid (analytical grade reagents), methanol (≥99.8%), tert-butyl-methyl ether (MTBE), water (HPLC grade reagents), and anhydrous sodium carbonate (99.5%) were purchased from Fisher Scientific International Inc. (Pittsburgh, PA, USA). Standard compounds of astaxanthin (≥98%), lutein (≥92%), and β-carotene (≥95%) for HPLC analysis were purchased from Acros Organics BVBA (Antwerp, Belgium), Extrasynthese SAS (Lyon, France), and Sigma Aldrich Co. (Saint Louis, MO, USA), respectively. Anhydrous D(+)-glucose (≥99.8%) and gallic acid (98%) (ACS reagents) were purchased from Acros Organics BVBA (Antwerp, Belgium). Analytical grade potassium chloride was purchased from Panreac Quimica SA (Barcelona, Spain), while 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical and Folin–Ciocalteu reagent were purchased from Sigma Aldrich Co. (Saint Louis, MO, USA) and Carlo Erba Reagents SAS (Milan, Italy), respectively.