Protocatechuic acid

Reagent grade quality ethanol, chloroform, ethyl acetate, dichloromethane, methanol, n-heptane, sodium methoxylated, and boron trifluoride in methanol (14% w/v) were purchased from Merck Life Science (Merck KGaA, Darmstadt, Germany).
LC–MS grade methanol, acetonitrile acetic acid, water, and HPLC grade methyl tert-butyl ether were also acquired from Merck Life Science (Merck KGaA, Darmstadt, Germany).
Standard of gallic acid, protocatechuic acid, coumarin, chlorogenic acid, catechin, epicatechin, and ferulic acid were purchase form Merck Life Science (Merck KGaA, Darmstadt, Germany).

6-Hydroxy-2,5,7,8-tetramethyl-3,4-dihydrochromene-2-carboxylic acid (trolox), (−)-epicatechin, ethyl gallate, protocatechuic acid, Folin–Ciocalteu reagent, sodium carbonate, gallic acid, (+)-catechin, vanillin, hydrochloric acid, potassium chloride, sodium acetate, DMSO, 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), IL1α, ethanol, formic acid and LC-MS grade solvents were purchased from Merck KGaA, Darmstadt, Germany. Quercetin 3-galactoside and malvidin 3-glucoside were obtained from Extrasynthese (Genay CEDEX, France). LC-grade H₂O (18 MΩ cm) was prepared with a Milli-Q H₂O purification system (Millipore, Bedford, MA, USA).

A C7-C40 Saturated Alkanes (1000 g/mL) standard mixture in hexane (49452-U) supplied by Merck Life Science (Merck KGaA, Darmstadt, Germany) was utilized for ALKANEs linear retention indices (LRIs) calculation.
Trolox (6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid), Gallic acid, AAPH (2,20-azobis(2-amidinopropane) dihydrochloride), fluorescein sodium salt, Folin-Ciocalteu reagent, sodium phosphate monobasic (NaH₂PO₄) and potassium phosphate dibasic (K₂HPO₄) were purchased from Merck Life Science (Merck KGaA, Darmstadt, Germany).
LC-MS-grade water, methanol, acetonitrile, and acetic acid were obtained from Merck Life Science (Merck KGaA, Darmstadt, Germany). Gallic acid, protocatechuic acid, isoquercetin, myricetin, and cyanidin were purchased from Merck Life Science (Merck KGaA, Darmstadt, Germany). Stock solutions of 1000 mg L⁻¹ were prepared for each standard by dissolving 10 mg in 10 mL of methanol.

Thinned Golden, Fuji, Bella del Bosco and Rosa Mantovana apples were sourced from farms located in Trentino-Alto Adige, Italy. Ultrapure water was prepared by a Milli-Q purification system (Millipore, Bedford, MA, USA). Protocatechuic acid, caffeic acid, p-coumaric acid, (+)-catechin, (−)-epicatechin, prunin (naringenin glucoside), phlorizin, phloretin, luteolin, cysteine (Cys), iodoacetamide (IAA), tris(2-carboxyethyl)phosphine (TCEP), tetraethylammonium bromide (TEAB), 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), IL-1α, TNF-α, 6-hydroxy-2,5,7,8-tetramethyl-3,4-dihydrochromene-2-carboxylic acid (Trolox), ascorbic acid, catechin, sodium carbonate, Folin–Ciocalteu reagent, ferulic acid, naringenin-7-O-glucoside, quercetin, ethanol, methanol, phlorizin dihydrate, sodium acetate, acetic acid, 2,2-diphenyl-1-picrylhydrazyl (DPPH), CDDO-Me, rosiglitazone, formic acid (FA), trifluoroacetic acid (TFA), acetonitrile (ACN) and all ultra-pure-grade (99.5%) solvents used in LC-MS analysis were obtained from Merck KGaA, Darmstadt, Germany. Kaempferol, quercetin, quercetin-3-O-rhamnoside and quercetin-3-O-galactoside were purchased from Extrasynthese (Genay Cedex, France). S-TRAP™ columns were provided by Protifi (Huntington, NY, USA).

2,2-Diphenyl-1-picrylhydrazyl (DPPH), butylated hydroxytoluene (BHT), vitamin E (α-tocopherol), gallic acid, quercetin, potassium ferricyanide, ferric chloride, ammonium molybdate, acetylcholinesterase (AChE), tacrine, α-amylase, acarbose, penicillin, Folin-Ciocalteu phenol reagent, and HPLC grade reagents: quinic acid, caffeic acid, protocatechuic acid, 1,3-di-O-caffeoylquinic acid, 3,4-di-O-caffeoylquinic acid, 4,5-di-O-caffeoylquinic acid, apigenin-7-O-glucoside, cirsilineol, and acacetin were purchased from Merck (Sigma-Aldrich, Steinheim, Germany).

Methanol, acetonitrile (all HPLC grade), and formic acid (50%, LC–MS grade) were purchased from Sigma–Aldrich (Milan, Italy). Ultrapure water (18.2 MΩ cm at 25 °C) was obtained by ELGA PURELAB Ultra system (M-medical, Cornaredo, Milan, Italy). Anthocyanins (Cyanidin-3-O-glucoside and Peonidin-3-O-glucoside) were obtained from LGC Standard srl (Sesto San Giovanni, Milan, Italy). All the other polyphenol reference compounds (gallic acid, protocatechuic acid, p-hydroxybenzoic acid, vanillic acid, coumaric acid, ferulic acid, catechin, myricetin), chemicals, and reagents were of analytical grade and purchased from Merck KGaA (Darmstadt, Germany).

All the solvents and chemicals used in this study were analytical grade. Methanol, ethanol, acetonitrile LC-MS grade, formic acid LC-MS grade, 85% phosphoric acid, and water LC-MS grade were purchased from Merck^® (Darmstadt, Germany). Gallic acid, quinic acid, caffeic acid, ferulic acid, protocatechuic acid, neochlorogenic acid (3-O-caffeoylquinic acid), chlorogenic acid (5-O-caffeoylquinic acid), cynarin (1,3-dicaffeoylquinic acid), isochlorogenic acid, kaempferol-3-O-glucoside, quercetin-3-O-glucoside, ferrous sulfate, 1,1-diphenyl-2-picrylhydrazyl radical (DPPH^●), (±)-6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), 2,4,6-tris(2-pyridyl) -1,3,5-triazine (TPTZ), 2,2′-azino-bis(3-ethylbenzothiazoline)-6-sulfonic acid (ABTS), Folin–Ciocalteu reagent, sodium carbonate, ferric chloride, AlCl₃, NaNO₂, and CuSO₄∙5H₂O were purchased from Merck^®, Sigma-Aldrich^®, or Fluka™ (Milan, Italy). Flavonoid standards of apigenin, apigenin-7-O-glucoside, hesperidin, hesperetin, luteolin-7-O-glucoside, naringin, and naringenin were purchased from Extrasynthese (Genay, France). Ultrapure water (18 MΩ·cm) was obtained with a Milli-Q Advantage A10 System apparatus (Millipore, Milan, Italy).