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G005895

LC/MS/MS Analysis of Steroid Hormones in Plasma on Ascentis® Express C18 after Sample Prep using HybridSPE®-Phospholipid

application for HPLC

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technique(s)

HPLC: suitable

test parameters

SPE tube/cartridge: HybridSPE®-Phospholipid, 96-well plate, 50 mg/well (575656-U)
sample addition: To each well add 100 μL of plasma followed by 300 μL of precipitation solvent (1% formic acid or 0.5% citric acid in acetonitrile).  (Agitate via vortex for 4 minute, place on vacuum manifold and apply 10" Hg vacuum for 4 minutes. Collect filtrate and analyze directly.)
column: Ascentis® Express C18, 10 cm x 2.1 mm I.D., 2.7 μm particles (53823-U)
gradient: 60% B for 3 min, 60% B to 95% B in 5 min, held at 95% B for 2 min
mobile phase: [A] 5 mM ammonium formate pH 4.0 with formic acid; [B] methanol
flow rate: 0.3 mL/min
column temp.: 50 °C
detector: ESI+, MRM
injection: 2 μL

suitability

application for HPLC

application(s)

clinical

Analysis Note

This application shows the separation of steroid hormones in plasma using LC/MS/MS. HybridSPE-Phospholipid removed endogenous phospholipids and precipitated proteins, thereby allowing rapid, sensitive analysis by LC/MS/MS using an Ascentis Express C18 column. The highest grade LC-MS solvents were used to supply low background interference and low particulate contaminants for robust, trouble-free operation. Cerilliant CRMs provided reliable identification and quantification.

Legal Information

Ascentis is a registered trademark of Merck KGaA, Darmstadt, Germany
HybridSPE is a registered trademark of Merck KGaA, Darmstadt, Germany

Analyte

Description
    Aldosterone solution, 100 μg/mL in acetonitrile, ampule of 1 mL, certified reference material, Cerilliant®
    Corticosterone solution, 1.0 mg/mL in methanol, ampule of 1 mL, certified reference material, Cerilliant®
    11-Deoxycorticosterone solution, 100 μg/mL in methanol, ampule of 1 mL, certified reference material, Cerilliant®
    Testosterone solution, 1.0 mg/mL in acetonitrile, ampule of 1 mL, certified reference material, Cerilliant®
    17α-Methyltestosterone solution, 1.0 mg/mL in 1,2-dimethoxyethane, ampule of 1 mL, certified reference material, Cerilliant®
    Progesterone solution, 1.0 mg/mL in acetonitrile, ampule of 1 mL, certified reference material, Cerilliant®

SPE tube or plate

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analytical column

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standard

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