Analytical–grade methanol (MeOH), acetonitrile (ACN), isopropanol (IPA), and methyl tertiary butyl ether (MTBE) were purchased from Fisher Scientific Co., Ltd (Fair Lawn, NJ, USA). Formic acid (FA) and ammonium acetate (NH4OAc) were purchased from Merck (Darmstadt, Germany). Ultrapure water was provided by Wa–Haha Corporation (Hangzhou, China). Moxifloxacin hydrochloride tablets were produced by Bayer Pharma AG (Leverkusen, Germany). Anticleaved caspase–3 antibody was obtained from Abcam (ab2302).
Ammonium acetate nh4oac
Manufactured by Merck Group
Sourced in United States, Germany, Australia
Ammonium acetate (NH4OAc) is a chemical compound commonly used in various laboratory applications. It is a white crystalline solid that serves as a pH buffer and is often used in analytical and biochemical procedures.
Automatically generated - may contain errors
Lab products found in correlation
Ammonium hydroxide nh4oh, by Merck Group (5 mentions)
Lc ms grade acetonitrile acn, by Merck Group (4 mentions)
Formic acid, by Merck Group (3 mentions)
Water h2o, by Honeywell (3 mentions)
Ammonium hydroxide, by Merck Group (3 mentions)
Lc ms grade water h2o, by Honeywell (2 mentions)
Lc ms grade methanol meoh, by Honeywell (2 mentions)
Ms grade acetonitrile, by Merck Group (2 mentions)
Ar grade formic acid, by Merck Group (2 mentions)
2h4 nam, by LGC (2 mentions)
Amino phase nh2 column, by Phenomenex (2 mentions)
Anti cleaved caspase 3 antibody, by Abcam (1 mentions)
Metformin hydrochloride, by Merck Group (1 mentions)
Hplc grade methanol meoh, by Merck Group (1 mentions)
Bilirubin, by AppliChem (1 mentions)
Dimethyl sulfoxide (dmso), by AppliChem (1 mentions)
Methionine, by Merck Group (1 mentions)
Riboflavin, by Merck Group (1 mentions)
Choline, by Merck Group (1 mentions)
Pyridoxine, by Merck Group (1 mentions)
19 protocols using ammonium acetate nh4oac
1
Moxifloxacin HPLC Quantification in Tablets
2
UPLC-MS/MS Analysis of Metformin and Guanylurea
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Methanol (MeOH) hypergrade LC-MS (Chromasolv), water hypergrade LC-MS (Chromasolv), acetonitrile (ACN) (LC-MS grade), isopropanol (LC-MS grade), and formic acid (98%) were purchased from Sigma-Aldrich (Steinheim, Germany). The pharmaceutical standards metformin hydrochloride and guanylurea sulfate were supplied by Sigma-Aldrich, whereas deuterated metformin-d6 was purchased from Toronto Research Chemicals (North York, Canada). An isotopically labeled standard for guanylurea was not commercially available. Ammonium acetate (NH4OAc) (98%) and glacial acetic acid (HOAc) (100%) were obtained from Merck (Darmstadt, Germany). Cartridges for SPE (Strata-X-CW, 30 mg) were supplied by Phenomenex (Aschaffenburg, Germany). A 3-mm PTFE syringe filter (0.45 μm) was supplied by Macherey-Nagel (Düren, Germany). Individual stock solutions of metformin and guanylurea with a concentration of 1 g/l were prepared in LC-MS grade water. The stock solution of metformin-d6 with a concentration of 1 g/l was prepared in MeOH (hypergrade LC-MS). All working solutions of the standards and samples for direct injection were prepared in MeOH containing 10% BGE using the 25 mM NH4OAc in MeOH:HOAc (97:3) to avoid current breakdown and band broadening due to field amplification [35 , 36 (link)]. Stock and working solutions were stored at − 20 °C before use.
3
HPLC Analysis of Bilirubin and Biliverdin
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HPLC grade methanol (MeOH) and synthetic grade acetic acid (AcOH) were purchased from Sigma-Aldrich (Steinheim, Germany). HPLC analysed dimethyl sulfoxide (DMSO) was purchased from J.T. Baker (Deventer, The Netherlands). Ammonium acetate (NH 4 OAc) was purchased from Merck (Darmstadt, Germany). TLC grade biliverdin hydrochloride (Z97%) and bilirubin (Z 99%) were purchased from Applichem (Darmstadt, Germany); both were stored at À 20 °C.
Individual standard solutions of bilirubin and biliverdin were prepared as follows: an accurately weighed amount of standard (2-3 mg) was dissolved in appropriate volume of DMSO to gain 10 mM solution. Subsequently, dilutions till 10 mM concentration were made by separately pouring 1 mL aliquots of each individual standard solution into 10 mL flat-bottomed volumetric flask, which was then filled up to 10 mL with MeOH. Further dilutions were made using a mixture of MeOH:H 2 O (3:1, v/v). Twice deionized water was used. All solutions were prepared fresh daily and the whole sample preparation workflow was performed in dim light.
Individual standard solutions of bilirubin and biliverdin were prepared as follows: an accurately weighed amount of standard (2-3 mg) was dissolved in appropriate volume of DMSO to gain 10 mM solution. Subsequently, dilutions till 10 mM concentration were made by separately pouring 1 mL aliquots of each individual standard solution into 10 mL flat-bottomed volumetric flask, which was then filled up to 10 mL with MeOH. Further dilutions were made using a mixture of MeOH:H 2 O (3:1, v/v). Twice deionized water was used. All solutions were prepared fresh daily and the whole sample preparation workflow was performed in dim light.
4
Mass Spectrometry Metabolite Analysis
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LC–MS-grade acetonitrile (ACN), LC-grade methanol (MeOH), formic acid, perfluoroheptanoic acid (PFHA), ascorbic acid, sodium hydroxide (NaOH) 0.1 M, hydrochloric acid (HCl), tris(2-carboxyethyl)phosphine (TCEP), dithiothreitol (DTT), and ammonium acetate (NH4OAc) were purchased from Sigma-Aldrich Chemie GmbH (Buchs, Switzerland). Deionized water (R > 18 ΩM/cm, TOC < 10 ppb) was used throughout the experiments and produced by a Millipore-Q water system (Millipore, Bedford, MA, USA).
HA, taurine, serine, cystine, glycine, homocystine (HCy2), riboflavin, methionine, pyridoxine, cystathionine, SAH, pyridoxamine, SAM, DMG, choline, betaine, 5-MTHF, taurine-13C2, glycine-d2, riboflavin-dioxopyrimidine-13C415N2, and methionine-d3 standards were purchased from Sigma-Aldrich Chemie GmbH.
HA-d4, serine-d3, pyridoxine-d2, cystathionine-d4, SAM-d4, DMG-d6, choline-d9, and betaine-d11 were purchased from CDN Isotopes (Pointe-Claire, Quebec, Canada). cystine-d4, homocystine-d8, and pyridoxamine-d3 were purchased from Cambridge Isotopes Laboratories (Andover, MA, USA). SAH-d4 was purchased from Cayman Chemical (Ann Arbor, MI, USA). 5-MTHF-13C5 was purchased from Merck (Schaffhausen, Switzerland).
HA, taurine, serine, cystine, glycine, homocystine (HCy2), riboflavin, methionine, pyridoxine, cystathionine, SAH, pyridoxamine, SAM, DMG, choline, betaine, 5-MTHF, taurine-13C2, glycine-d2, riboflavin-dioxopyrimidine-13C415N2, and methionine-d3 standards were purchased from Sigma-Aldrich Chemie GmbH.
HA-d4, serine-d3, pyridoxine-d2, cystathionine-d4, SAM-d4, DMG-d6, choline-d9, and betaine-d11 were purchased from CDN Isotopes (Pointe-Claire, Quebec, Canada). cystine-d4, homocystine-d8, and pyridoxamine-d3 were purchased from Cambridge Isotopes Laboratories (Andover, MA, USA). SAH-d4 was purchased from Cayman Chemical (Ann Arbor, MI, USA). 5-MTHF-13C5 was purchased from Merck (Schaffhausen, Switzerland).
5
Quantification of One-Carbon Metabolites
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SAM, Met, Hcy, SAH, Cys, LC-MS-grade acetonitrile, sodium hydroxide (NaOH, 0.1 mol/L), tris-(2-carboxyethyl) phosphine (TCEP), hydrochloric acid (HCl), GSH, formic acid, ascorbic acid, LC-grade methanol (MeOH), and ammonium acetate (NH4OAc) were from Sigma–Aldrich (MO, USA). Homocysteine-d8 was from CDN Isotopes (Pointe-Claire, Quebec, Canada). Ultra-pure grade water was prepared with a Millipore-Q instrument (Millipore, MA, USA).
6
Metabolite Extraction and LC-MS Analysis
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LC−MS grade water (H2O) and methanol (MeOH) were purchased from Honeywell (Muskegon, USA). Ammonium hydroxide (NH4OH) and ammonium acetate (NH4OAc) were purchased from Sigma-Aldrich (St. Louis, USA). Chemical standards of metabolites were purchased from J&K (Beijing, China), Sigma (St. Louis, USA), Carbosynth (Berkshire, UK), TCI (Tokyo, Japan), and Energy Chemical (Shanghai, China). Serum samples (50 µL) were extracted using 150 μL MeOH with internal standards (d3-leucine and d8-phenylalaine). The samples were then vortexed for 30 s and sonicated for 15 min. To precipitate proteins, the samples were incubated for 1 h at −20 °C, followed by 15 min centrifugation at 17,500×g and 4 °C. The supernatants were transferred to HPLC vials and stored at −80 °C prior to LC−MS analysis.
7
Liver S9 Fraction Characterization
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Pooled human liver S9 fraction (H0610.S9) and NADPH regenerating system (K5100-5) were purchased from Sekisui Xenotech (Kansas City, KS, USA). The cofactors adenosine 3′-phosphate 5′-phosphosulfate lithium salt hydrate (PAPS), acetyl-coenzyme A (acetyl-CoA), uridine diphosphate glucuronic acid (UDPGA) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The glutathione (GSH) was purchased from J&K (Shanghai, China). The NIST urine (SRM 3667) and NIST plasma (SRM 1950) samples were purchased from Ango Biotechnology Co. (Shanghai, China). LC–MS grade methanol (MeOH) and water (H2O) were purchased from Honeywell (Muskegon, MI, USA). LC–MS grade acetonitrile (ACN) was purchased from Merck (Darmstadt, Germany). Ammonium hydroxide (NH4OH) and ammonium acetate (NH4OAc) were purchased from Sigma (St. Louis, MO, USA). Other chemical standards were purchased from Sigma-Aldrich (St. Louis, MO), J&K (Shanghai, China), and TopScience (Shanghai, China).
8
LC-MS Analysis of Metabolites
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LC–MS grade methanol (MeOH) and water (H2O) were purchased from Honeywell (Muskegon, MI, USA). LC–MS grade acetonitrile (ACN) was purchased from Merck (Darmstadt, Germany). LC–MS grade methylene dichloride (CH2Cl2) was purchased from Fisher Scientific (Morris Plains, NJ, USA). Ammonium hydroxide (NH4OH) and ammonium acetate (NH4OAc) were purchased from Sigma (St. Louis, MO, USA). The chemical standard succinoadenosine was purchased from J&K (Shanghai, China), while other chemical standards were purchased from TopScience (Shanghai, China).
9
Ribonuclease A Interaction with Oligonucleotides
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A stock solution of bovine pancreatic ribonuclease A (#R6513, Sigma-Aldrich, St Louis, MO, USA) was prepared at 5.60 mg/mL in LC-MS grade water (Thermo-Fisher Scientific, Waltham, MA, USA). Ammonium acetate (NH
4OAc) was LC-MS grade (#73594, Sigma-Aldrich). HPLC-purified deoxyoligonucleotides with the sequence “CCCCC” (dC
5) and “CCACC” (dC
2AC
2) were obtained from ThermoFisher and the stock solutions (200 μM) were prepared in LC-MS grade water. Samples were prepared in 1.5 mL microcentrifuge tubes as indicated in
Table 1 . Six replicates were prepared and analyzed for “Sample 1” whereas “Samples 2–5” were prepared and analyzed in triplicate. Each sample was mixed by micropipetting, and incubated at room temperature for ten minutes, prior to analysis.
4OAc) was LC-MS grade (#73594, Sigma-Aldrich). HPLC-purified deoxyoligonucleotides with the sequence “CCCCC” (dC
5) and “CCACC” (dC
2AC
2) were obtained from ThermoFisher and the stock solutions (200 μM) were prepared in LC-MS grade water. Samples were prepared in 1.5 mL microcentrifuge tubes as indicated in
10
Metabolite Extraction and Quantification
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BA was purchased from Sigma-Aldrich (St. Louis, MO, USA). Thymol was bought from Science Lab (Houston, TX, USA). Internal standards, L-glutamic acid-13C5 and sodium L-lactate-13C3, were purchased from Cambridge Isotope Laboratories, Inc. (Tewksbury, MA, USA). Methanol, isopropanol, and acetonitrile (ACN) (all HPLC-grade) were purchased from VWR International (Radnor, PA, USA). Ammonium acetate (NH4OAc) and ammonium hydroxide (NH4OH) solution were purchased from Sigma-Aldrich (St. Louis, MO, USA). Deionized water was provided by an in-house Milli-Q® Water Purification System (Billerica, MA, USA). Phosphate-buffered saline (PBS) was purchased from GE Healthcare Life Sciences HyClone Laboratories (Chicago, IL, USA).
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