Project description:Urine MS/MS data acquired to test Molecular Networking as tool for drug screening in human urine to assist in drug adherence monitoring. Samples from a antihypertensive drug adherence human cohort were retroperspectively selected to test how many commonly prescribed antihypertensives and other over-the-counter drugs were picked up by Molecular Networking in 26 extracts. In addition, several endogenous molecular families were also annotated. Data was obtained in positive and in negative ionization mode on a Thermo Scientific Q-Exactive Orbitrap instrument, using stepped collision energy HCD fragmentation in a data-dependent-manner (DDA).
Urine samples from anonymized human volunteers were used from a clinical sample set in the Glasgow Polyomics archive. These samples were obtained as part of a trial for which ethical approval was applied for through the Multi-centre Research and Ethics Committee (MREC), which was granted by the Scottish MREC and (with MREC N°06/MRE00/106). Informed consent was obtained from all individual study participants. Spot urine samples were obtained from the cohort of elderly hypertensive patients upon their first admission in the clinic. Urine extracts of 26 patients were selected as follows: diagnosed with hypertension, taking in a variety of different antihypertensive drugs (i.e., different drug classes), and availability of the sample extract in the Glasgow Polyomics archive. The resulting subject’s age range spanned from 42 to 87; 15 were male, 11 female; 4 were smokers; 5 were reported to have diabetes; and each reportedly took from 2 to 7 different classes of antihypertensive drugs.
Full details also on data acquisition and analysis can be found in Van der Hooft et al. 2016, Metabolomics.
Chromatography and Mass Spectrometry details:
The samples were analysed using a Thermo Scientific Ultimate 3000 RSLCnano system (Thermo Scientific, CA, USA). The pHILIC separation was performed with a SeQuant ZIC-pHILIC column (150 x 4.6 mm, 5 microm) equipped with the corresponding pre-column (Merck KGaA, Darmstadt, Germany) - the column temperature was maintained at 25 C. A linear biphasic LC gradient was conducted from 80% B to 20% B over 15 min, followed by a 2 min wash with 5% B, and 8 min re-equilibration with 80% B, where solvent B is acetonitrile and solvent A is 20 mM ammonium carbonate in water. The flow rate was 300 microL/min, column temperature was held at 25 C, injection volume was 10 microL, and samples were maintained at 4 C in the autosampler.
MS equipment and settings used: The LC system was coupled to a Thermo Scientific Q-Exactive Orbitrap mass spectrometer equipped with a HESI II interface (Thermo Scientific, Hemel Hempstead, UK). The set up was calibrated (Thermo calmix) in both ionization modes and tuned for the lower m/z range before analysis. Full scan (MS1) data was acquired in positive and negative switching mode in profile mode at 35,000 resolution (at m/z 200) using 1 microscan, an AGC target of 106 cts, a maximum injection time of 250 ms, with spray voltages +3.8 and -3.0 kV, capillary temperature 320 C, heater temperature of 150 C, sheath gas flow rate 40 a.u., auxiliary gas flow rate 5 a.u., sweep gas flow rate 5 a.u, a full scan mass window of 70-1050 m/z, and using m/z 74.0964 (+) and m/z 112.98563 (-) as locking masses. Fragmentation data (LC-MS/MS) was obtained in positive and negative ionization combined and separate fragmentation modes as described in (van der Hooft et al 2016). Briefly, for separate mode, a duty cycle consisted of one full scan (MS1) event and one Top5 (or Top10) MS/MS (MS2) fragmentation event, with full scan (MS1) resolution (at m/z 200) was set to 70,000, the AGC target set to 1 x 106, and the maximum injection time set to 120 ms. MS/MS (MS2) resolution (at m/z 200) was set to 17,500, the AGC target set to 2 x 105, MS/MS maximum injection time was set to 80 ms and the underfill ratio was set to 10 %, with a resulting intensity threshold of 2.5 × 105 cts. For combined mode, a duty cycle consisted of two of the above events in positive and negative ionization mode with the following modifications: full scan (MS1) resolution (at m/z 200) was set to 35,000, MS/MS resolution was set to 35,000, the AGC target was set to 1 x 105, and the maximum MS/MS filling time was set to 120 ms with an underfill ratio of 20%, resulting in an intensity threshold of 1.7 x 105. Further settings were as specified for full scan analysis above.
| MSV000082971 | GNPS
Justin van der Hooft 
