It tof mass spectrometer

The composition of the unaged SOA was studied
with an LC20A liquid chromatograph coupled with an IT-TOF mass spectrometer
(Shimadzu). The Waters Symmetry C18 column (150 mm × 2.1 mm,
3.5 μm, 100 Å), an eluent flow rate of 0.2 mL/min, ACN,
and an aqueous solution of formic acid (0.03% v/v, pH = 2.8) were
used. The gradient program was as follows: 0–2 min 5%B, 2–3
min increase to 12%B, 3–11 min 12%B, 11–12 min increase
to 18%B, 12–28 min 18%B, 28–41 min increase to 95%B,
41–45 min 95%B, 45–46 increase to 5%B, and the total
analysis time was 56 min. The ESI ion source was operating in the
negative ionization mode, the spray voltage was −3.5 kV, the
nebulizing gas flow was 1.5 mL/min, and the source temperature and
desolvation line temperature were 200 °C. The detected [M-H]⁻ ions were assumed to be singly charged, deprotonated
pseudo-molecular ions containing C, H, and O atoms. O/C values and
double-bond equivalents were calculated with the acquired HR/MS data.

All of the reagents and solvents were purchased from Aladdin (Shanghai, China) or Sinopharm Chemical Reagent Co. Ltd. (Shanghai, China), and were used as received. Melting points were determined in open capillary tubes and are uncorrected. Reaction courses were monitored by thin-layer chromatography on silica gel-precoated F254 plates (Merck, Darmstadt, Germany). Developed plates were examined with UV lamps (254 nm). Nuclear magnetic resonance spectroscopy was performed on an AV-300 spectrometer (Bruker, Zurich, Switzerland) operating at 300 MHz for ¹H and 75 MHz for ¹³C and using DMSO-d₆ as solvent and tetramethylsilane as the internal standard. Electrospray Ionization Mass Spectrometry (ESI-MS) experiments were performed on an IT-TOF mass spectrometer (Shimadzu, Tokyo, Japan) in negative ion mode. Specific optical rotation was measured on a Digital automatic polariscope JASCO P-1020 (JASCO, Tokyo, Japan).

All protected α-amino acids were purchased from Bachem A (Bubendorf, Switzerland). Opioid radioligands, [³H]DAMGO, [³H]deltorphin-2, and [³H]U-69593, and human recombinant opioid receptors were purchased from PerkinElmer (Krakow, Poland). GF/B glass fiber strips were obtained from Whatman (Brentford, UK). Purity of peptides was determined by RP-HPLC and exact mass. Analytical and semi-preparative RP-HPLC was performed using Waters Breeze instrument (Milford, MA, USA) with dual absorbance detector (Waters 2487, MA, USA). All ESI-MS experiments were performed on a Shimadzu IT-TOF mass spectrometer (Shimadzu, Japan) equipped with ESI source connected to Nexera HPLC system (Shimadzu, Japan). The instrument was operated in the positive-ion mode. Peptide solutions (1 μL) were introduced in a 0.2 mL/min flow of mobile phase. For LC-MS experiments, Aeris Peptide C₁₈ and Kinetex Biphenyl (Phenomenex, Torrance, CA, USA) were used, in a gradient reversed-phase mode, from 5 to 80% acetonitrile in water (both containing 0.1% HCOOH). ¹H NMR spectra were recorded on a 500 MHz Brucker instrument in DMSO-d₆, using residual DMSO as a resonance reference at 2.5 ppm.

Analytical TLC was carried out on 60 F₂₅₄ silica gel plates (absorbent thickness: 0.25 mm, Merck, Darrmstadt, Germany). Column chromatography was performed using silica gel (Merck 230–400 mesh, ASTM). IR spectra were recorded using a Perkin-Elmer Spectrum 400 FT-IR Spectrometer (Perkin Elmer, Waltham, MA, USA). NMR spectra were acquired in CD₃OD (Merck) using a JEOL ECA 400 MHz NMR spectrometer (JEOL, Tokyo, Japan). The LCMS-IT-TOF spectra were obtained on a UFLC Shimadzu Liquid Chromatograph with a SPD-M20A diode array detector coupled to an IT-TOF mass spectrometer (Shimadzu, Kyoto, Japan). UV spectra were recorded using a Shimadzu 1650 PC UV-Vis Spectrophotometer (Shimadzu). All solvents were of analytical grade and were distilled prior to use.

RE-CE was purchased from ICTORY Biological Technology Co., Ltd. (Xi’an, China). The raw materials of coriander herbs were dried without access to direct sunlight after copious washing in running water. The dried materials were ground into a fine power and passed through a 40-mesh sieve. This powered product (500 g) was submitted to extraction with aquaous ethanol (30%, 5 L) in a shaker at 60 °C for 2 h. The extract waste was filtered out and the solution was concentrated by a rotary evaporator and dried under vacuum. The extract was stored at 4 °C for further use. LC/MS analyses were performed on a Shimadzu LC-30AD liquid chromatography interfaced with an IT-TOF mass spectrometer (Shimadzu Corp., Kyoto, Japan) with electrospray ionization. An ACOUITY UPLC BEH-C18 Column (1.7 μm) was used. The column temperature was 40 °C and the elution velocity was 0.3 mL/min. The sample extracts were analyzed using a gradient program, and the mobile phase consisted of 0.1% formic acid in water (solvent A) and HPLC grade methanol (solvent B). The gradient program consisted of: 5% B for 0 min, 95% B for 50 min, and 95% B for 60 min. The nebulizer gas flow rate was 1.5 L/min. CDL and block heater temperatures were both 200 °C. The spray and detector voltages were 4.5 and 1.58 kV, respectively. The scan time and mass range were 1.5 s and 50–1000 m/z, respectively. The injected volume was 2 μL.

After conventional organic extraction from crude cell pellets, phosphatidylcholine (PC) and phosphatidylethanolamine (PE) species were semiquantified in a Shimadzu IT-TOF mass spectrometer hyphenated with a Shimadzu Prominence HPLC system (Babraham Institute Lipidomics laboratory, UK). For the measurement of free fatty acids (FFA) release, a colorimetric reagent for measuring free glycerol levels (ZenBio, USA) was used directly on whole cell lysates from ∼2×10⁵ cells, prepared by homogeneization in HNE buffer (20 mM HEPES pH 7.9, 150 mM NaCl, 5 mM EDTA, and protease inhibitors). Colorimetric signal was measured on an Envision unit (Perkin Elmer) using a 530 nm absorbance filter. Measurements in triplicate from three independent experiments were collected for each assay, and normalized for protein content as estimated by Bradford assay.