Analysis of Serum 17-Hydroxyprogesterone, Androstenedione, and Cortisol by UPLC-MS/MS for Clinical Research

19 Jan 2015

Measurement of 17-hydroxyprogesterone (17-OHP) by immunoassay is prone to analytical interference arising from cross-reactivity of reagent antibodies with structurally-related steroid metabolites. In this application note, a method for the extraction and UPLC®-MS/MS analysis of serum 17-OHP and two additional adrenal steroids, androstenedione (A4) and cortisol, using the ACQUITY UPLC System with the Xevo TQ MS is described.

Xevo TQ MS

Waters

Waters Xevo™ TQ MS provides the ultimate in quantitative capabilities for your most demanding applications and your most complex sample matrices. It is a highly advanced tandem quadrupole mass spectrometer designed to enable uncomplicated access to superior HPLC and UPLC®/MS/MS performance, delivering the highest levels of sensitivity, selectivity, robustness, speed, and accuracy. Waters Xevo TQ MS is equipped with unique collision cell technology (ScanWave™) to provide new enhanced spectral LC/MS/MS data acquisition capabilities. Unlike conventional tandem quadrupole mass spectrometers, the Xevo TQ MS is designed not only to provide the very best quantitative data, but also to provide you with superior spectral MS/MS information. Xevo TQ MS offers you much more versatility in a single instrument platform that is adaptable to a variety of different UPLC/MS/MS applications both quantitative and qualitative: bioanalysis, ADME screening, food safety, environmental monitoring, forensic, and more. Features: Automated system set up and system checks for simple access Automated instrument optimization and quantitative method development tools for ease-of-use The latest innovations in high efficiency, robust, easy-to-use ZSpray™ atmospheric pressure ion source technology Versatile range of ionization options, including multi-mode ionization capability in a single run Unique, innovative T-Wave™ and ScanWave enabled collision cell technology Footprint that sits easily on your laboratory benchtop Dedicated application managers for your specialized analytical requirements

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ACQUITY UPLC System

Waters

Waters ACQUITY UPLC - Dramatic Improvements in Speed, Sensitivity and Resolution Whether you work in methods development or quality assurance, in drug discovery and development or testing for safety in food supplies, you seek a productivity edge. With the ACQUITY UPLC® System, now you can get more information in a single, short run than you've ever seen with HPLC. Waters ACQUITY UPLC System UltraPerformance LC® (UPLC®) technology starts with unique 1.7 μm small-particle chemistries. Chromatographers no longer need to choose between speed and resolution–with UPLC you get both. Instrument Designed for Chemistry; Chemistry Designed for the Instrument The ACQUITY UPLC System has been holistically designed to match the performance needs of our innovative column chemistries with robust hardware, easy-to-use software and specialized support services. Small, pressure-tolerant particles High-pressure fluidic modules Minimized system volume Negligible carryover Reduced cycle times Fast response detectors Integrated system software and diagnostics With UPLC you have the ability to work more efficiently with higher speed, sensitivity and resolution at a much wider range of linear velocities, flow rates and backpressures to obtain superior results. The Waters ACQUITY UPLC System's high-pressure fluidics optimizes flow rates to make the most of small particle technology. The ACQUITY UPLC System's sample-handling design is designed to ensure exceptionally low carryover and reduced cycle time. And when interfaced with the Sample Organizer, it increases unattended sample capacity by ten times. High-speed detectors, both optical and mass, contribute to increased sensitivity and help manage the heightened speed and resolution requirements of UPLC. ACQUITY UPLC Systems are easily controlled, diagnosed, and monitored via a graphical system console interface. The console offers: Quick and easy access to critical instrument parameters Simple system start-up, elegant system status monitoring and predictive performance indicators to ensure maximum productivity Data management capabilities that are supported by both MassLynx™ and Empower™ software The ACQUITY UPLC System is also supported by Intelligent Device Management technology with our Connections® INSIGHT™ service, providing instrument diagnostics.

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ACQUITY UPLC HSS T3 Columns for Polar Compound Retention

Waters

ACQUITY UPLC® HSS (High Strength Silica) T3 Columns contain the first and only 100% silica particle designed, tested and intended for use in applications up to 15000 psi/1000 bar. The ACQUITY UPLC® HSS particle is not an HPLC particle. High pore volume HPLC particles do not possess the mechanical stability necessary to withstand the high column packing and operating pressures of UPLC® separations. The first ligand chemistry in this UPLC®-certified column family utilizes T3 bonding in order to retain and separate polar organic compounds. ACQUITY UPLC® HSS T3 columns possess the superior polar-compound retention, aqueous mobile phase compatibility and ultra-low MS bleed of Atlantis® T3 columns. Easier Method Transfer Since Atlantis® T3 HPLC and ACQUITY UPLC® HSS T3 columns utilize T3 bonding, LC methods can be transferred from HPLC to UPLC® separations. Because Atlantis® and HSS particles are different substrates, some method modification and/or optimization may be required. A compelling benefit of UPLC® technology is the ease and speed at which this method optimization can occur since UPLC® separations offer greater resolution in less time. Moving from XBridge™ HPLC to ACQUITY UPLC® BEH columns is nearly seamless, as the hybrid particle substrates differ only in particle size. However, because of T3 bonding it should be straightforward to make simple adjustments to successfully transfer a separation from an Atlantis® T3 HPLC column to an ACQUITY UPLC® HSS T3 column.

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