2 2 diphenyl 1 picrylhydrazyl dpph free radical

The roots of S. ningpoensis were collected from the experimental field of Jining Medical University (Rizhao, Shandong, China) and identified by Jian'an Wang (Medical Botanist, Jining Medical University). Monosaccharide standards D-glucose (D-Glc), D-GalActose (D-Gal), D-arabinose (D-Ara), L-fucose (L-Fuc), D-mannose (D-Man), L-rhamnose (L-Rha), D-fructose (D-Fru), D-xylose (D-Xyl), D-GlcA, and D-GalA were purchased from Sigma-Aldrich (St. Louis, MO, USA). 2,2-Diphenyl-1-picrylhydrazyl (DPPH) free radical was purchased from Sigma-Aldrich (St. Louis, MO, USA). 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radical was purchased from the Shanghai Yuanye Biotechnology Co. Ltd. (Shanghai, China). ImProm-II Reverse Transcription System was obtained from Promega Corporation (Madison, WI, USA). IL-6 Mouse ELISA Kit was purchased from Invitrogen (Carlsbad, CA, USA). TNF-α Mouse ELISA Kit was obtained from PeproTech (Rocky Hill, NJ, USA). UNIQ-10 column Trizol total RNA extraction kit was obtained from Sangon Biological Engineering Technology and Services Co., Ltd. (Shanghai, China). FastStart Universal SYBR Green Master (ROX) kit was purchased from Roche (Mannheim, Germany). All chemical reagents were of analytical grade, and the deionized water was used in the whole experiment.

The reagents sodium carbonate, di-sodium hydrogen phosphate, sodium carbonate, potassium ferricyanide, potassium persulfate, ethylenediamine tetra acetic acid (EDTA) and gallic acid were purchased from Scharlau (Barcelona, Spain). Folin–Ciocalteu’s phenol reagent, absolute ethanol and trichloroacetic acid (TCA) were purchased from Panreac (Castellar del Vallès, Barcelona, Spain). 2,2-diphenyl-1-picrylhydrazyl (DPPH•) free radical, Iron (II) Chloride 4-hydrate, 6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylacid (Trolox), (L)-Dehydroascorbic acid and malondialdehyde (MDA) were purchased from Sigma Chemical Co. (Steinheim, Germany, and St. Louis, MO, USA). Propylgallate (PG) was purchased from Acrōs Organics (Fair Lawn, NJ, USA). 2-Thiobarbituric acid (TBA) was purchased from Merck KGaA (Darmstadt, Germany). The following standards were used: phenolic acids—hydroxybenzoic acid, vanillic acid, protocatechuic acid; cinnamic acids—chlorogenic acid, 4-coumaric acid; flavanols—catechin; flavonols—kaempferol, quercetin, kaempferol-3-O-rutinoside, kaempferol-3-O-glucoside, isorhamnetin-3-O-rutinoside, rutin or quercetin-3-O-rutinoside; flavanones—naringenin, eriodictyol, eriodictyol-7-O-glucoside (Merck KGaA, Darmstadt, Germany).

Methanol (HPLC grade), ethanol, butanol, hexane, and chloroform (ACS grade), ascorbic acid, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical were obtained from Sigma-Aldrich (Saint Louis, Missouri, USA). Green tea extract (pre-standardized to 50% epigallocatechin gallate) was obtained from HerbStoreUS (Walnut, CA, USA).

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.

All reagents used were analytical grade. Ethyl acetate, 2,2-diphenyl-1-picrylhydrazyl (DPPH•) free radical, and gallic acid (97%) were obtained from Sigma-Aldrich (Castle Hill, Australia). Acetic acid, ethanol, and methanol were from Merck (Darmstadt, Germany), and anisaldehyde was from ACROS organics (New Jersey, USA). Milli-Q (Millipore) water was used to prepare all aqueous solutions. Aluminium chloride (reagent grade, 98%), 3-hydroxytyrosol (≥98%), maslinic acid (≥98%), oleanolic acid (≥97%), rosmarinic acid (≥98%), rutin hydrate (≥94%), and ursolic acid (≥90%) were purchased from Merck (Merck KGaA, Darmstadt, Germany).