Reference standards and deuterium labeled internal standards for vitamin D metabolites were purchased from Medical Isotopes Inc. (Pelham, NH). An ACQUITY Charged Surface Hybrid (CSH) C18 (2.1 mm, 150mm, 1.7μm) and 96-well Ostro™ plates (25mg) were purchased from Waters (Milford, MA). Derivatization reagent 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) was purchased from Sigma-Aldrich (St. Louis, MO). 12-[[(cyclohexylamino)carbonyl]-amino]-dodecanoic acid (CUDA) was purchased from Cayman Chemical (Ann Arbor, MI). Formic acid, ammonium acetate, LC-MS grade water, LC-MS grade methanol, and LC-MS grade acetonitrile were purchased from Fisher Scientific (Hampton, NH). Pooled human plasma collected from at least 10 healthy drug free individuals was purchased from BioIVT (Westbury, NY). Standard reference material (SRM) 972a was purchased from the National Institute of Standards and Technology (NIST) (Gaithersburg, MD).
4 phenyl 1 2 4 triazoline 3 5 dione ptad
Manufactured by Merck Group
Sourced in United Kingdom
4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) is a chemical compound used as a reagent in analytical chemistry. It is a white crystalline solid that reacts with various functional groups, such as alcohols, amines, and thiols, to form stable adducts that can be analyzed using techniques like chromatography and mass spectrometry.
Automatically generated - may contain errors
Lab products found in correlation
Formic acid, by Thermo Fisher Scientific (3 mentions)
Acetonitrile, by Thermo Fisher Scientific (2 mentions)
Methylamine, by Merck Group (2 mentions)
Lc ms grade methanol, by Thermo Fisher Scientific (1 mentions)
Lc ms grade acetonitrile, by Thermo Fisher Scientific (1 mentions)
Lc ms grade water, by Thermo Fisher Scientific (1 mentions)
Ammonium acetate, by Thermo Fisher Scientific (1 mentions)
12 cyclohexylamino carbonyl amino dodecanoic acid cuda, by Cayman Chemical (1 mentions)
Ferric chloride, by Merck Group (1 mentions)
Catechin, by Merck Group (1 mentions)
Sodium carbonate, by Merck Group (1 mentions)
Ammonium thiocyanate, by Merck Group (1 mentions)
Pepsin, by Merck Group (1 mentions)
α amylase, by Merck Group (1 mentions)
Vanillin, by Merck Group (1 mentions)
Retinyl acetate, by Merck Group (1 mentions)
Pancreatin, by Merck Group (1 mentions)
Hydrochloric acid (hcl), by Carlo Erba (1 mentions)
Formic acid, by Carlo Erba (1 mentions)
Vitamin d3, by Merck Group (1 mentions)
8 protocols using 4 phenyl 1 2 4 triazoline 3 5 dione ptad
1
Vitamin D Metabolite Quantification
2
Comprehensive Carotenoid and Vitamin Analysis
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Retinyl acetate, β-carotene, α-carotene, vitamin D3 (> 95% pure), deuterated vitamin D3, 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD), phytic acid, DMACA, vanillin, iron III, pancreatin, ammonium thiocyanate, catechins, α-amylase, pepsin, bile, ferric chloride and sodium carbonate were purchased from Sigma Aldrich (Saint-Quentin-Fallavier, France). Lutein, lycopene, phytoene and phytofluene were purchased from Extrasynthèse (Genay, France). Soyasaponin I was generously supplied by Stéphane Georgé, CTCPA Avignon. Canned tomato pulp, red bell peppers and Isio4 oil (Lesieur, Asnières, France) were purchased from a local supermarket (Marseille, France). Methanol, acetonitrile, hexane, dichloromethane, methyl-tert-butylether, acetone, ethanol absolute anhydrous, sulfuric acid, hydrochloric acid, nitric acid, formic acid, hydrogen peroxide were purchased from CarloErba Reagents (Peypin, France).
3
Quantification of Vitamin D Metabolites
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The standard of 25(OH)D2 was purchased from Sigma Aldrich (St. Louis, MO). Standards of 25(OH)D3 and 1,25(OH)2D3 were obtained from MilliporeSigma (Burlington, MA). The standard of 24,25(OH)2D3 was from MP Biomedicals (Irvine, CA). Stable isotope labeled internal standards (I.S.) (i.e., d6-1,25(OH)2D3, d6-25(OH)D2 and d6-25(OH)D3) were purchased from Medical Isotope (Pelham, NH). 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD), used as a derivatization reagent, was obtained from Sigma Aldrich (St. Louis, MO), along with bovine serum albumin (BSA). Phosphate buffered saline (PBS) and HPLC grade acetonitrile (ACN) were purchased from Fisher Chemicals (Waltham, MA). Water was deionized by a Milli-Q water purification system (MilliporeSigma, MA). Formic acid (FA) was purchased from Sigma-Aldrich (St. Louis, MO).
4
Quantification of Conjugated Linoleic Acid
5
Vitamin D Metabolite Quantification
6
Optimizing 7-Dehydrocholesterol Quantification
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7DHC (≥ 95% chemical purity) and 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD; 97% chemical purity) were purchased from Sigma-Aldrich Chemicals Company Ltd, Dorset, UK. Stable isotope labelled 3β-7-DehydroCholesterol-d7 (3β-7DHC-d7; 98% chemical purity) was purchased from Toronto Research Chemicals, Toronto, Canada. 7DHC, 3β-7DHC-d7 and PTAD were stored at − 20 °C in sealed amber glass ampoules prior to use, to avoid loss of UVR spectral integrity (due to photoconversion). Deionised water (H2O), LCMS grade methanol (MeOH) (≥ 99.9%) and acetonitrile (ACN) (≥ 99.9%), analytical reagent grade ethyl acetate (EA) (≥ 99%) and propan-2-ol and HPLC grade hexane and formic acid were purchased from Fisher Scientific, Loughborough, UK.
Stock solutions, containing 20 µg/mL 7DHC in ACN and 5 µg/mL 7DHC Internal Standard (IS) in MeOH or ACN, were prepared and stored at + 4 °C. 7DHC stock solution was then further diluted with ACN to obtain working concentrations as outlined in the sample preparation procedure. New stock solutions were prepared every 2–3 weeks, as there was a loss of signal in solutions older than 2 months.
Stock solutions, containing 20 µg/mL 7DHC in ACN and 5 µg/mL 7DHC Internal Standard (IS) in MeOH or ACN, were prepared and stored at + 4 °C. 7DHC stock solution was then further diluted with ACN to obtain working concentrations as outlined in the sample preparation procedure. New stock solutions were prepared every 2–3 weeks, as there was a loss of signal in solutions older than 2 months.
7
Comprehensive LC-MS/MS Vitamin D Analysis
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All the chemicals used in the study were of analytical grade. The following chemicals were purchased from Sigma-Aldrich (Steinheim, Germany): vitD 3 , vitD 2 , 25(OH)D 3 , 25(OH)D 2 , petroleum ether, diethylether, anhydrous acetonitrile (99.8%), Fluka formic acid (LC-MS grade), methylamine, 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) and ammonium formate. 26, 26, 27, 27, ] and 25-hydroxyvitamin D 3 -[26,26,26,27,27,27-d 6 ] were purchased from Chemaphor Inc. (Ottawa, Canada), whereas vitamin D 3 - [23,24,25,26,27-13 C 5 ] and 25-hydroxyvitamin D 3 -[23,24,25,26,27-13 C 5 ] were purchased from Isosciences (Ambler, Pennsylvania). Ethyl acetate and 2-propanol were purchased from Rathburn Chemicals Ltd (Walkerburn, Scotland). N-heptane and methanol (LC-MS grade) was from Honeywell (Steinheim, Germany), whereas ethanol (96%), sodium ascorbate and potassium hydroxide were purchased from Merck (Darmstadt, Germany). Ultra-pure Milli-Q water was made in-house (18.2 MΩ, Millipore, Bradford, USA).
8
Quantification of Vitamin D Metabolites
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Materials, calibration standards and controls SRM972 traceable 25(OH)D 3 and 25(OH)D 2 serum based calibrators (Chromsystems, München, Germany) and internal quality controls (IQC) (UTAK Laboratories, CA, USA) were analysed in each run. Certified pure standards for 24,25(OH) 2 D 3 , 24,25(OH) 2 D 2 were used for preparation of spiked standards and deuterated standards 25(OH)D 3 -[ 2 H 6 ] and 24,25(OH) 2 D 3 -[ 2 H 3 ] (IsoSciences, King of Prussia, PA, USA) were used as internal standards. Deionised water, methanol, acetonitrile and formic acid were LCMS grade, n-heptane and isopropanol were analytical grade (Fisher Scientific, Loughborough, UK). 4phenyl-1,2,4-triazoline-3,5-dione (PTAD) and methylamine (Sigma-Aldrich, Dorset, UK) were used for derivatization and adduct formation.
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