Malvidin 3 glucoside

In this study, the following reagents and standards were used: aluminium chloride, quercetin dihydrate, gallic acid monohydrate, potassium persulfate, Trolox, 2,2-azinobis(3-ethylbenzothiazoline sulfonic acid) (ABTS), 2,2-diphenyl-1-picrylhydrazil (DPPH), 2,4,6-tripyridyl-s-triazine (TPTZ) were obtained from Sigma-Aldrich (St. Lois, MO, USA); HPLC standards (gallic and caffeic acid, (+)-catechin hydrate, (−)-epicatechin, rutin hydrate, quercetin) were purchased from Sigma-Aldrich Chemie Gmbh (Steinheim, Germany), and malvidin 3-glucoside from Extrasynthese (Genay, France). Folin–Ciocalteu reagent, sodium nitrite, sodium carbonate, sodium hydroxide, potassium bisulphite, sodium acetate, potassium chloride and hydrochloric acid were purchased from Kemika (Zagreb, Croatia); sodium acetate trihydrate, ferric chloride hexahydrate, and ammonium acetate were purchased from Gram-Mol (Zagreb, Croatia); copper(II) chloride from Acros Organics (New Jersey, NJ, USA); HPLC grade methanol and neocuproine from Merck (Darmstadt, Germany) and phosphoric acid (HPLC grade) were obtained from Fluka (Buchs, Switzerland).

The fluid aqueous extract obtained from blood orange processing wastes (ExF) was produced by Ortogel SpA (Caltagirone, Sicily, Italy). Beta-cyclodextrin was supplied by Roquette Frères (Lestrem, France). Sodium alginate (ALG), fluorescein (FL), AAPH (2,2′ azobis(2-methylpropionamide) dihydrochloride) 97%, Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) and HEPES (4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid), aminoguanidine bicarbonate 97% (AMG), bovine serum albumin (BSA), D-(−)-fructose, and sodium azide (NaN₃) were purchased from Sigma-Aldrich Srl (Milan, Italy). Anthocyanins (cyanidin-3-glucoside, cyanidin-3,5-diglucoside, cyanidin-3-rutinoside, and cyanidin-3-sophoroside;delphinidin-3-glucoside and delphinidin-3,5-diglucoside; pelargonidin-3-glucoside and pexlargonidin-3,5-diglucoside; peonidin-3-glucoside; and malvidin-3-glucoside) and flavanones (hesperidin, narirutin, and didymin) were purchased from Extrasynthèse (Genay, France). OmniMMP fluorescent substrate Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH₂, MMP-9 (refolded) (human) (recombinant) (catalytic domain) and MMP-2 (catalytic domain) (human) (recombinant) were purchased from Vinci-Biochem Srl (Firenze, Italy). Solvents for chromatography were HPLC grade (Merck KGaA, Darmstadt, Germany). All the other chemicals used in the study were of analytical grade and were obtained commercially.

An aliquot of 500 mg of berry skin powder was freeze-dried for 72 h and the dried powder (∼50 mg) were extracted in 1.0 mL methanol containing 0.1% HCL (v/v). Extracts were filtered through a 0.45 μm polypropylene syringe filter (Pall Gelman Corp., Ann Harbor, MI, USA) for HPLC analysis. Each individual anthocyanin was analyzed as described in Hilbert et al. (2003) and Acevedo De la Cruz et al. (2012) (link) with HPLC. Quantification was carried out by peak area integration at 520 nm, and Malvidin-3-glucoside (Extrasynthèse, Lyon, France) standard was used for quantify the anthocyanin concentration.

Amberlite XAD7HP, AAPH (2,2′-diazo-bis-amidinepropane-dihydrochloride) and Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic) were purchased from Sigma (Munich, Germany). Fluorescein was provided by Fluka. Methanol for liquid chromatography and acetic acid 99.8% were purchased from Merck (Munich, Germany) and VWR CHEMICALS (Radnor, PA, United States), respectively. Malvidin-3-glucoside, cyanidin-3-glucoside, cyanidin-3-galactoside, peonidin-3-glucoside and delphinidin-3-glucoside were purchased from Extrasynthese (Genay, France).