Maxis q tof spectrometer

Semipreparative HPLC was performed with an Alliance chromatographic system (Waters Corporation, Mildford, MA, USA) and an Atlantis C18 column (10 μm, 10 × 150 mm, Waters). For UPLC analysis an Acquity UPLC equipment (Waters) with a BEH C18 column (1.7 μm, 2.1 × 100 mm, Waters) was used. Optical rotations were determined with a JASCO P-2000 polarimeter (JASCO Corporation, Tokyo, Japan). IR spectrum was measured with a JASCO Fourier transform infrared (FT/IR)-4100 spectrometer (JASCO Corporation) equipped with a PIKE MIRacle^TM single reflection ATR accessory. NMR spectra were recorded on a Bruker Avance III spectrometer (500 and 125 MHz for ¹H and ¹³C NMR, respectively) equipped with a 1.7 mm TCI MicroCryoProbe^TM (Bruker Biospin, Fällanden, Switzerland), using the signal of the residual solvent as internal reference (δ_H 2.50 and δ_C 39.5 ppm for DMSO-d₆). ESI-TOF MS spectra were acquired with a Bruker maXis QTOF spectrometer (Bruker Daltonik GmbH, Bremen, Germany).

Antibody stock solutions were diluted to 1–2 mg/mL and thoroughly dialyzed against mQ water. Acetic acid (BCD090-P1) or formic acid (all other samples) were added to protein solutions to a final concentration of 1%. Mass spectra were measured on a maXis Q-TOF spectrometer (Bruker Daltonik) in positive ion mode using direct injection with electrospray ionization. Mass spectra were averaged using DataAnalysis 4.0 software (Bruker Daltonik), and then exported as text files containing m/z versus intensity. All further analysis, namely, deconvolution of charge states and fitting of the isotopic distributions, was performed according to the methodology developed by Rhoads et al. [31 (link)] in Matlab using their script, which is available in Supplementary Materials for the paper [31 (link)].

The UV spectra were measured on a Beckman DU-800 (Beckman Coulter, Inc., Brea, IN, USA). IR spectra were obtained from a Nicolet iN10 spectrometer (Thermo Fisher Scientific Inc., Waltham, MA, USA) using KBr pellets. NMR spectra were recorded on a Bruker Avance III 600 NMR spectrometer (600 MHz for ¹H and 150 MHz for ¹³C, Bruker Corporation, Billerica, MA, USA). HRESIMS spectra were acquired from a Dionex Ultimate 3000 system (Thermo Fisher Scientific Inc., MA, USA) coupled with a Bruker Maxis Q-TOF spectrometer (Bruker Corporation, MA, USA) using positive and negative electrospray ionization. CD spectra were recorded on a JASCO J-815-150S circular dichroism spectrometer (Jasco Corp., Tokyo, Japan). The compounds were monitored by an Agilent 1260 system (Agilent Technologies, Santa Clara, CA, USA) equipped with a G1315D photodiode array detector (DAD) using an Agilent ZORBAX Eclipse XDB-C18 column (4.6 × 150 mm, 5 μm) at a flow rate of 1 mL/min. The semi-preparative HPLC experiments were performed on a Waters X Bridge (C18, 5 μm, 150 × 10 mm) column using an Agilent 1260 system at a 2 mL/min flow rate. Silica gel (200–300 mesh, Qing Dao Hai Yang Chemical Group Co., Ltd., Qingdao, China) and octadecyl silyl (ODS) silica gel (40 μm, 120 Å, Ji Nan Bo Na Biological Technology, Jinan, China) were used for column chromatography.