Inertsustainswift c18

The Fc carboxylic acid and O-(benzotriazol-1-yl)-N,N,N′,N′tetramethyluronium tetrafluoroborate (TBTU) were purchased from Tokyo Chemical Industry (Tokyo, Japan). 1-Hydroxybenzotriazole monohydrate (HOBt) was purchased from Watanabe Chemical Industries (Hiroshima, Japan). The βCyD and N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) were purchased from Sigma-Aldrich (Saint Louis, MO, USA) and Dojindo Laboratories (Kumamoto, Japan), respectively. All ODNs were synthesized by an automated DNA synthesizer (8900, Expedite or NTS M-2-MX, Nihon Techno Service) using conventional phosphoramidite methods. The phosphoramidite monomers were purchased from Proligo (Hamburg, Germany) and Glen Research (Sterling, VA, USA). All ODNs and ODN conjugates were purified by reversed-phase HPLC (Column: InertSustain Swift C18, 4.6 mm φ × 150 mm, GL Science, Tokyo) using an appropriate linear gradient with 0.1 M triethylamine-acetic acid (TEAA)-acetonitrile (pH 7.0), and identified by MALDI-TOF mass spectrometry on a Bruker Daltonics Autoflex-III (Billerica, MA, USA) using 3-hydroxypicolinic acid as a matrix. All other reagents were obtained as the highest grade and used without further purification.

Total quercetin derivatives, sum of deconjugated quercetin and methylquercetin, were determined using LC-MS/MS system [25] (link) (TSQ Quantum Access Max with Accela High Speed LC System, Thermo Fisher Scientific Inc., MA, USA) with an electrospray ionization (ESI) source with SRM mode. The analytical column was a 1.9 μm C18 column (Inertsustain Swift C18, 2.1 mm × 100 mm GL Sciences Inc., Tokyo, Japan) set at 40C. The mobile phase was water, methanol, and formic acid (70:30:0.1, solvent A), and methanol with formic acid (100:0.1, solvent B). The ratio of solvent A and B was delivered through a linear gradient (90:10 for 1 min; 90:10-20:80 for 8 min; 90:10 for 1 min) at a flow rate of 0.2 mL/min. Concentrations of total quercetin derivatives were calculated from the calibration curve of standard compounds.