3 4 hydroxyphenyl propionic acid

All chemicals were of analytical grade. Amino acid standard AAS18 (Sigma-Aldrich Chemical Co., St. Louis, MO, USA) was used for the identification of amino acids. Lactic acid, benzoic acid, succinic acid, 4-hydroxybenzaldehyde, 3- phenylpropionic acid, malic Acid, 4-methoxyphenylacetic acid, Gamma-Aminobutyric acid, trans-Cinnamic acid, 3-hydroxybenzoic acid, 4-hydroxybenzoic acid, 4-hydroxyphenylacetic acid, adonitol, hydrocinnamic acid, 3,4-dihydroxyphenylacetic acid, citric acid, D-Fructose, d-Galactose, d-Glucose, 3,4-dihydroxyhydrocinnamic acid, 3-(4-hydroxyphenyl)propionic acid, 1H-indole-3-acetic Acid, D-mannitol, dextrose, sucrose, indole, were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA) and used for identification purposes. Stock solutions of all the analytical standards were prepared by dissolving the compounds in MilliQ water.

Organosolv lignin (OL) was purchased from Chemical Point (Oberhaching, Germany). Mushroom tyrosinase (1000 U/mg), bovine serum albumin (BSA), glutaraldehyde (GA), poly (diallyldimethylammonium chloride) (PDDA, 20% v/v water solution), 2-chloro-4,4,5,5-tetramethyl-1,3,2-dioxaphopholane, chrome (III) acetylacetonate, deuterated chloroform (CDCl₃), ascorbic acid, Bradford reagent, l-tyrosine, para-cresol, tyrosol, 4-hydroxyphenylacetic acid, 3-(4-hydroxyphenyl)propionic acid, catechol, potassium chloride (KCl), and organic solvents were purchased from Sigma–Aldrich (St. Louis, MO, USA), and were used without any further purification. All experiments were done in triplicate using native and immobilized tyrosinase in Phosphate buffer saline (PBS).

High-purity standards, such as chlorogenic acid (5-O-caffeoylquinic acid, CHA); quercetin (QU); kaempferol (KA); p-coumaric acid (p-CA); miquelianin (quercetin 3-O-β-D-glucuronopyranoside, MQ); 2-(3′,4′-dihydroxyphenyl)acetic acid (PAA); protocatechuic acid (PCA); 3-(4′-hydroxyphenyl)propionic acid (PPA); 3-(3′,4′-dihydroxyphenyl)propionic acid (dihydrocaffeic acid, DCA); and ascorbic acid (AA) were purchased from Sigma-Aldrich (St. Louis, MO, USA), while proanthocyanidin A2 (PA2) was obtained from Phytolab (Vestenbergsgreuth, Germany). The standards of juglanin (kaempferol 3-O-α-L-arabinofuranoside, JU), avicularin (quercetin 3-O-α-L-arabinofuranoside, AV) and kaempferitrin (kaempferol 3,7-di-O-α-L -rhamnopyranoside, KT) were isolated previously in the Department of Pharmacognosy, Medical University of Lodz, Lodz, Poland, from the flowers and leaves of P. spinosa, with HPLC and NMR purity > 98% [26 (link),27 (link)]. The structures of the model native polyphenols and phenolic metabolites investigated in the study are presented in Figure 1 and Figure 2, respectively.

The reagents used were obtained from the following suppliers. Quercetin (Q4951) from Sigma-Aldrich^®, St. Louis, MO, USA, 3,4-dihydroxyphenylacetic acid, 3-(4-hydroxyphenyl) propionic acid, p-coumaric acid, 4-methylcatechol, protocatechuic acid, bisphenol A (239658) from Sigma-Aldrich^®, pepsin, pancreatin, 2,2-diphenyl-1-picrylhydrazyl (DPPH), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (trolox), 2,2′-azo-bis (2-amidino-propane) dihydrochloride (AAPH), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT, M5655), dimethyl sulfoxide (DMSO, D4540), Triton X-100 (1002214179), from Sigma-Aldrich^®, hydrogen peroxide, phosphate buffered saline (PBS), ethanol, fetal bovine serum (FBS), and Dulbecco’s modified eagle medium (DMEM, 12800-058) from Gibco^®.
Commercial kits used were lactate dehydrogenase or LDH (Roche^®, Basel, Switzerland, 11644793001), Muse Cell Cycle Assay (Merck Millipore^®, Burlington, MA, USA, MCH100101).

Quercetin, isorhamnetin, tamarixetin, and 3-coumaric acid were purchased from Extrasynthese. Quercetin-3′-sulfate, Quercetin-3-glucuronide, and isorhamnetin-3-glucuronide were synthetized as described [45 (link)]. xanthine oxidase (from bovine milk), allopurinol, 6-mercaptopurine, hypoxanthine, xanthine, uric acid, oxipurinol, 3-phenylpropionic acid, 3-(4-hydroxyphenyl)propionic acid, 3-(2,4-dihydroxyphenyl)propionic acid, 2-hydroxyphenylacetic acid, 4-hydroxyphenylacetic acid, 3,4-dihydroxyphenylacetic acid, 3-hydroxy-4-methoxyphenylacetic acid, 4-(hydroxymethyl)phenylacetic acid, benzoic acid, 4-hydroxybenzoic acid, 2,4-dihydroxybenzoic acid, 3,4-dihydroxybenzoic acid, hippuric acid, 2,4-dihydroxyacetophenon, 4-methylcatechol, resorcinol, pyrogallol, phloroglucinol, 4-methoxysalicylic acid, and homovanillic acid were obtained from Sigma-Aldrich (St. Louis, MO, US). 3-(3-hydroxyphenyl)propionic acid and 3-(3,4-dihydroxyphenyl)propionic acid were purchased from Toronto Research Chemicals. 6-thiouric acid was obtained from Carbosynth, 6-thioxanthine was purchased from 5A Pharmatech. uric acid was dissolved in 0.01 M sodium hydroxide (2 mM), while Q and its metabolites, APU, 6-MP, 6-TX, 6-TU, and xanthine were dissolved in dimethyl sulfoxide (each 2 mM) and stored at –20 °C.