Deuterium oxide (H2O-d2, 99.90% D) and MeOH-d4 (99.80% D) were purchased from Eurisotop (Cambridge Isotope Laboratories, Inc, Saint-Aubin, France). Ultra-low Endotoxin FBS was obtained from Euroclone (Euroclone, Milan, Italy). Mini-PROTEAN® TGX™ precast gels 4–20%, Precision Plus Protein™ Unstained Standards, Clarity™ Western ECL Substrate and DC™ protein assay were purchased from Bio-Rad Laboratories (Hercules, California, United States). Primary antibodies against Nf-kB were purchased from Millipore (Merck Millipore, Burlington, Massachusetts, United States). Standard 3-(trimethylsilyl)-propionic-2,2,3,3-d4 acid sodium salt (TMSP), sodium phosphate dibasic anhydrous sodium phosphate monobasic anhydrous, Dulbecco’s modified Eagle medium (DMEM), penicillin, streptomycin, glutamine, lipopolysaccharide (LPS) from Escherichia coli serotype O127:B8, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), and all the other solvents and chemicals were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA).
Meoh d4
Manufactured by Cambridge Isotopes
Sourced in United States
MeOH-d4 is a deuterated methanol compound, with all four hydrogen atoms replaced by deuterium. It is a colorless liquid and is commonly used as a solvent in nuclear magnetic resonance (NMR) spectroscopy and other analytical techniques.
Automatically generated - may contain errors
Lab products found in correlation
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Merck Group (1 mentions)
Mini protean tgx precast gels 4 20, by Bio-Rad (1 mentions)
Streptomycin, by Merck Group (1 mentions)
Glutamine, by Merck Group (1 mentions)
Precision plus protein unstained standard, by Bio-Rad (1 mentions)
Dc protein assay, by Bio-Rad (1 mentions)
Deuterium oxide, by Cambridge Isotopes (1 mentions)
Ultra low endotoxin fbs, by Euroclone (1 mentions)
Vnmrs 400, by Agilent Technologies (1 mentions)
Acetone d6, by Cambridge Isotopes (1 mentions)
6545 qtof w 1290 hplc mass spectrometer, by Agilent Technologies (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Trypsin, by Merck Group (1 mentions)
Rpmi 1640 culture medium, by Merck Group (1 mentions)
Diaion hp 20, by Merck Group (1 mentions)
Hplc grade methanol, by Mallinckrodt (1 mentions)
Hplc grade acetonitrile, by Merck Group (1 mentions)
Methanol, by Merck Group (1 mentions)
Milli q water purification system, by Merck Group (1 mentions)
Sephadex lh 20, by Cytiva (1 mentions)
7 protocols using meoh d4
1
Deuterium Oxide and LPS-induced Inflammation
2
High-Resolution Mass Spectrometry Protocols
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High-resolution mass spectrometry (HRMS) and liquid chromatography mass spectrometry (LCMS) were conducted on an Agilent 6545-QTOF W/1290 HPLC mass spectrometer at the University of Oklahoma Department of Chemistry and Biochemistry. NMR spectra were obtained on Varian VNMRS 400 or 500 MHz instruments at the NMR facility of the Department of Chemistry and Biochemistry of the University of Oklahoma using 99.9% DMSO-d6 with 0.05% v/v TMS or 99.9% D2O or 99.8% MeOH-d4 and 99.9% acetone-d6 (Cambridge Isotope Laboratories, MA, USA). 1H, 13C, and 31P chemical shifts were referenced to internal solvent resonances. Multiplicities are indicated by s (singlet), d (doublet), t (triplet), q (quartet), quin (quintet), m (multiplet), and br (broad). Chemical shifts are reported in parts per million (ppm) and coupling constants J are given in Hz. All NMR spectra were recorded at ambient temperature and processed using MestReNova software.
3
Spectroscopic Analysis of Purified Compounds
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The chemical structures of the purified compounds were determined by spectroscopic analyses and comparisons with values in previous literature. High-resolution electrospray ionization mass spectrometry (HRESIMS) data were determined by a Synapt G2 system (Waters Co., Milford, MA, USA). The 1D and 2D nuclear magnetic resonance (NMR) spectra were recorded by a Bruker Advance 400 MHz spectrometer (Burker BioSpin, Rheinstetten, Germany) at 400 MHz for 1H and 100 MHz for 13C in MeOH-d4 (Cambridge Isotope Laboratories, Tewksbury, MA, USA). Chemical shifts were referenced to the solvent peaks (δH 4.87 and δC 49.0 for MeOH-d4).
4
Metabolite Extraction and Quantification
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Methanol (MeOH), n-hexane, ethyl acetate, n-butanol, trifluoroacetic acid, formic acid, sodium formate, Diaion HP-20, silica gel (63–200 μm), RPMI 1640 culture medium, fetal bovine serum (FBS), antibiotics, and trypsin were purchased from Sigma–Aldrich (St. Louis, MO, USA). Water-soluble tetrazolium salt (WST-1) reagent was purchased from Abcam (Cambridge, MA, USA). HPLC-grade methanol was purchased from Mallinckrodt (KY, USA). MeOH-d4 was acquired from Cambridge Isotope Laboratories, Co. (MA, USA), and deionized water was prepared with a Barnstead water purification system (Dubuque, IA, USA).
5
NMR Characterization of PBAE and PEG-PBAE
6
Isolation and Characterization of SEI from L. chuanxiong
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SEI (purity >99.1%, as tested by HPLC-UV) was obtained from L. chuanxiong extracts in our laboratory. Its structure was confirmed by comparison of its MS and NMR profiles with that published in the literature [24 (link), 25 (link)]. Slices of L. chuanxiong (No. 20130419) were purchased from Bozhou Medicinal Materials Company (Anhui Province, China) and authenticated by Professor Zhi-li Zhao of the Shanghai University of Traditional Chinese Medicine (Shanghai, China). The ultrahigh purified water used in this study was prepared in a Milli-Q water purification system (Millipore Corp., Billerica, MA, USA). Methanol and acetonitrile (HPLC grade) were purchased from Merck KGaA (Darmstadt, Germany). MeOH-d4, with tetramethylsilane (TMS) as internal standard for NMR analysis, was purchased from Cambridge Isotope Laboratories, Inc. (Andover, MS, USA). MCI Gel CHP 20P (75 μm–150 μm) for column chromatography was purchased from Mitsubishi (Tokyo, Japan). Sephadex LH-20 was obtained from GE Healthcare Bio-Sciences AB (Sweden). Other reagents and chemicals, including formic acid, were of analytical grade.
7
Cimetidine Polymorph A Characterization
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Cimetidine (polymorph A), bulk drug of pharmaceutical-grade (BP 2002), was acquired to Saporiti (Buenos Aires, Argentina). During the experiments, the drug was kept in a desiccator, protected from light. MeOH-d 4 (99.8% atom-d) was acquired from Cambridge Isotope Labs, Inc. (Andover, MA, USA); all other solvents were of analytical grade and were used as received.
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