Cryptochlorogenic acid

The following substances and solvents were used in the study: procyanidins B1, B2, C1, A1, A2, A4, (+)-catechin, (–)-epicatechin, quercetin, quercitrin (quercetin-3-O-rhamnoside), isoquercitrin (quercetin-3-O-glucoside), avicularin (quercetin-3-O-arabinofuranoside), guaiaverin (quercetin-3-O-arabinopyranoside), rutin (quercetin-3-O-rutinose), reynoutrin (quercetin-3-O-xyloside), kaempferol, nicotiflorin (kaempferol-3-O-rutinoside), afzelin (kaempferol-3-O-rhamnoside), astragalin (kaempferol-3-O-glucoside), chlorogenic acid (3-O-caffeoylquinic acid), neochlorogenic acid (5-O-caffeoylquinic acid), cryptochlorogenic acid (4-O-caffeoylquinic acid), p-coumaric acid, arbutin, hydroquinone, α-amyrin, β-amyrin, β-sitosterol, lupeol, erythrodiol, maslinic acid, oleanolic acid, acetonitrile, acetone, and methanol, which were purchased from Sigma–Aldrich (Steinheim, Germany); hyperoside (quercetin-3-O-galactoside), procyanidin B3, uvaol, friedelin, 6″-O-acetylisoquercitrin (quercetin-3-O-(6″-acetylglucoside)), betulin, and betulinic and corosolic acids from Extrasynthese (Genay, France); ursolic acid from Carl Roth (Karlsruhe, Germany); and trifluoroacetic acid from Merck (Darmstadt, Germany). All used chemicals were of HPLC grade. Ultrapure water used in this study was prepared with Milli–Q^® 180 (Millipore, Bedford, MA, USA) water purification system.

A total of 14 A. quinata leaves samples were collected from different areas in Gongju and Muju, South Korea. All samples were authenticated by Professor Jong Seong Kang (College of Pharmacy, Chungnam National University, Daejeon, Korea), as shown in Figure S1 and Table S1 (Supplementary Materials). The samples were deposited in the pharmaceutical analysis laboratory, College of Pharmacy, Chungnam National University, with a code number of AQLS. The harvested samples were dried in an LTO-Do-150S oven (Labtech-one, Daejeon, Korea) at 45 °C for 3 days. After drying, the samples were pulverized into a fine powder, then stored at 4 °C until use. Mouse anti-ICAM-1 (#sc-8439), mouse VCAM-1 (#sc-13160), mouse anti-NF-kB p65 (sc-514451), and anti-phospho-eNOS antibody were purchased from Santa Cruz Technology, Inc. (Santa Cruz, CA, USA). The reference standards of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, rutin isochlorogenic acid C, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and a buffer solution of formic acid were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). HPLC-grade methyl alcohol and acetonitrile were purchased from Burdick & Jackson (Muskegon, MI, USA). Water was purified using a Milli-Q system (Milford, MA, USA).

Distilled water was purified using a Milli–Q system (Millipore, Bedford, MA, USA). Ethanol (96%) was obtained from Vilniaus degtine (Vilnius, Lithuania). Anhydrous acetic acid (99.8%), hydrochloric acid (37%) were purchased from Sigma–Aldrich (Buchs, Switzerland). The following reagents were used: 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), 2,4,6-Tri-(2-pyridyl)-S-triazine (TPTZ), ferric chloride hexahydrate (FeCl₃ × 6 H₂O), sodium acetate (CH₃COONa), 3-(2-pyridyl)-5,6-bis-(4-phenyl-sulfonic acid)-1,2,4-triazine (Ferrozine), obtained from Sigma-Aldrich (Buchs, Switzerland); potassium persulfate (K₂S₂O₈), anhydrous ferrous chloride (FeCl₂) from Alfa Aesar (Karlsruhe, Germany). The following standards were used: 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), ethylene diamine tetra acetic acid (EDTA), quercetin, chlorogenic acid, cryptochlorogenic acid, ferulic acid, procyanidin A2 from Sigma–Aldrich (Buchs, Switzerland); arbutin, (+)-catechin, (−)-epicatechin from Fluka (Buchs, Switzerland); hyperoside, astragalin, rutin, isorhamnetin-3-O-glucoside, quercitrin from Extrasynthese (Genay, France); procyanidin C1, avicularin from ChromaDex (Irvine, CA, USA). All the reagents and standards were of analytical grade.