Fluticasone propionate concentrations in plasma were quantified via a validated Liquid Chromatography - tandem Mass Spectrometry (LC-MS/MS) assay at Worldwide Clinical Trials (Austin, TX). The PK samples containing fluticasone propionate and the internal standard (i.e., deuterated FP, FP-D5) were extracted by liquid-liquid extraction and analyzed by a Sciex API 6500 (MS/MS transition: m/z 501.2 → 313.2). The inter- and intra-batch precision and accuracy for the quality control samples (1.5 pg/mL, 15 pg/mL, and 160 pg/mL) were both less than 15%. The lower limit of quantification was 0.5 pg/mL.
Api 6500
Manufactured by AB Sciex
Sourced in United States
The API 6500 is a high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) system designed for quantitative and qualitative analysis in a variety of applications. It features a triple quadrupole mass analyzer for sensitive and selective detection of target analytes.
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6 protocols using api 6500
1
Fluticasone Propionate Quantification by LC-MS/MS
2
Quantitative Analysis of Co-formulants and Active Substances
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Co‐formulants and active substances were analyzed using a LC–MS/MS system consisting of an autosampler (RSI PAL, CTC Analytics, Zwingen, Switzerland), a binary HPLC pump (Sciex Exion LC with microvacuum degasser), and a triple quadrupole mass spectrometer (API 6500+, with turbo ion spray source, Sciex Framingham, MA, USA). Analytes were separated by reversed‐phase HPLC on a Gemini‐NX C18 column (5 μm particle size, 150 × 2 mm, protected by a 4 × 2 mm pre‐column with the same stationary phase, Phenomenex, Torrance, CA). Eluents were aqueous ammonium formate (5 mM, puriss p.a., ACS reagent, ACROS ORGANICS, NJ, USA) and methanol (HPLC gradient grade, CHEMSOLUTE Renningen, DE, USA). Gradient elution at a flow rate of 0.2 mL min–1 was as follows: initial conditions, 75% methanol, linear increase to 100% within 5 min, followed by 5 min isocratic hold. Initial conditions were then re‐established within 0.1 min and the column equilibrated for 2 min. For more details on LC–MS/MS settings and ion transitions monitored see Table SI 2.1 .
3
Quantification of SLN360 in Biological Samples
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For the second study, the concentration of SLN360 in plasma, liver, and kidney was determined via liquid chromatography-tandem mass spectrometry. An antisense strand with 13C and 15N modifications on the 2′-O-methyl-cytosine nucleosides yielding a 48 Da mass shift was used as internal standard. Plasma samples were prepared via phenol chloroform extraction before injection. An Acquity ultra performance liquid chromatography I-Class equipped with an Acquity BEH C18, 1.7 µm 50 × 2.1 mm column from Waters was used. Separation was performed via ion-paring reverse-phase chromatography with ethylenediaminetetraacetic acid, triethylamine, and hexafluorisopropanol as mobile phase modifiers in water and methanol at 80°C. Detection was performed with a Sciex API 6500+ in negative mode following the 769.4→94.9 and 775→94.9 transition for SLN360 and the internal standard, respectively.
4
Pharmacokinetics of RMC-4550 in Plasma
5
Pharmacokinetics of RMC-4550 in Plasma
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RMC-4550 was formulated in sulfobutylether betacyclodextrin (Captisol™)/50 mM sodium acetate pH 4.6 (10%/90%, w/v%). Plasma samples for PK analysis were collected in a composite manner with 3 animals per time point. The plasma concentration of RMC-4550 was measured by a liquid chromatography tandem mass spectrometry (LC/MS/MS) method using positive electrospray ionization in multiple reaction monitoring mode. Plasma samples were extracted by protein precipitation using acetonitrile containing an internal standard. After vortexing and centrifugation, the supernatant was injected into an API6500 (AB SCIEX, CA) LC/MS/MS system for quantification. PK parameters were calculated by non-compartmental analysis of the plasma concentration-time profiles using Phoenix 64 (Certera, Menlo Park, CA).
6
Buprenorphine Pharmacokinetics in Cats
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All samples were centrifuged (Centrifuge Heraeus Sepatech) for 15 mins at 2937 g and plasma was stored in tubes at -20°C. For the measurements, the samples were stored and sent on dry ice to TOXILAB (Laboratory for Toxicology and Drug analysis, Ludwigsburg, Germany), for liquid chromatography mass spectrometric analysis, as this method is considered to be an appropriate analysis for a pharmacokinetic profile of buprenorphine. 27 Calibration standard and quality control had been established: after the addition of the deuterated standards buprenorphine-d4 and norbuprenorphine-D3 to calibration standard, quality control sample and plasma samples of the cats, protein precipitation was performed employing a solution of 70% methanol and 0.05 M zinc sulphate.
The examination was carried out with high-pressure liquid chromatography (pump: Shimadzu LC-20ADXR; autosampler: Shimadzu SIL-20ACXR; column: Agilent 4.6 × 5.0 Zorbax Eclipse × DB 1.8 µ C18; solvent mixture A: aqueous, 10 mmol/l ammonium acetate, pH 9.4; solvent mixture B: methanolic, 0.1% formic acid), as well as a tandem mass spectrometry (gadget: AB Sciex API 6500) following data analysis.
The examination was carried out with high-pressure liquid chromatography (pump: Shimadzu LC-20ADXR; autosampler: Shimadzu SIL-20ACXR; column: Agilent 4.6 × 5.0 Zorbax Eclipse × DB 1.8 µ C18; solvent mixture A: aqueous, 10 mmol/l ammonium acetate, pH 9.4; solvent mixture B: methanolic, 0.1% formic acid), as well as a tandem mass spectrometry (gadget: AB Sciex API 6500) following data analysis.
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