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3 protocols using sodium phosphate

1

Cinnamaldehyde Synthesis Protocol

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Cinnamaldehyde (natural, ≥ 95%, FG), trifluoroacetic acid (purity 99%), ammonium acetate and ammonium hydroxide (28–30%) were purchased from Sigma-Aldrich Chemie GmbH, Germany. L-lysine (Purity: > 95%) and l-cysteine (Purity: > 95%) were purchased from JPT, Germany. Acetonitrile (for HPLC-SUPER GRADIENT Reag. Ph. Eur., ACS water < 30 ppm-suitable for UPLC/UHPLC instruments) was purchased from VWR Prolabo Chemicals Pennsylvania, United States of America. Sodium phosphate, monobasic monohydrate and Sodium phosphate, dibasic heptahydrate were purchased from Kemika, Zagreb, Croatia. HPLC grade water was obtained using Arium Mini Sartorius (Goettingen, Germany) water purification systems.
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2

Purification and Characterization of Apo-Transferrin

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Native human apo-transferrin (Biorbyt, UK, cat. no. orb80927), sodium acetate trihydrate (Kemika, Croatia), calcium chloride (Lach-Ner, Croatia), sodium chloride (Kemika, Croatia), neuraminidase (GlycoCleave® Neuraminidase Kit, GALAB Technologies, Germany, cat. no. 132011), pISep Buffer Kit (CryoBioPhysica, USA, cat. no. 20055), hydrogen chloride (Carlo Erba Reagents, Italy, 37 % solution), sodium hydroxide (Kemika, Croatia, pellets 2–5 mm), sodium phosphate (Kemika, Croatia), guanidine hydrochloride (PanReac AppliChem, USA), MES (2-(N-morpholino)ethanesulfonic acid, Sigma Aldrich, USA) and potassium chloride (Alkaloid, North Macedonia) were used without further purification. Water used for experiments was double distilled in an all-glass apparatus. All experiments except the enzymatic desialylation were performed at room temperature.
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3

Phenolic Compound Extraction and Analysis

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Solvents used for the extraction and analysis were HPLC grade, namely ethanol, formic acid and acetonitrile, purchased from BDH Prolabo (Lutterworth, UK). Distilled water was of Milli-Q quality (Millipore Corp., Bedford, NY, USA). Anhydrous sodium carbonate (≥99.5%) and sodium phosphate (96%) were obtained from Kemika (Zagreb, Croatia), Folin-Ciocalteu reagent from Merck (Darmstadt, Germany), fluorescein sodium salt from Honeywell Riedel-de-Haën (Bucharest, Romania), Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) from Acros Organics (Thermo Fisher Scientific, Geel, Belgium) and 2,2′-Azobis (2-amidinopropane) hydrochloride from Sigma-Aldrich (Steinheim, Germany).
Phenolic compounds of authentic standards of caffeic, gallic, ferulic, chlorogenic and p-coumaric acid, quercetin-3-glucoside, kaempferol-3-rutinoside and myricetin were purchased from Sigma–Aldrich (Steinheim, Germany); epicatechin, catechin, epigallocatechin gallate, epicatechin gallate, procyanidin B1, procyanidin B2, apigenin and luteolin from Extrasynthese (Genay, France); and quercetin-3-rutinoside from Acros Organics (Thermo Fisher Scientific, Geel, Belgium).
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