Method for high throughput identification of trace organic contaminants in blood based on microsamples

A method for high-throughput identification of trace organic contaminants in blood based on microsamples, wherein the method comprises:

• step 1:
• sample preprocessing:
• adding 30 to 100 μL of a blood sample to a 96-well plate, and adding extractant according to a ratio between a volume of the blood sample and a mass of the extractant being 1:(0.54±0.2);
• adding a fixed volume of 300-400 UL acetonitrile to obtain a mixture, and centrifuging a supernatant after vortex mixing the mixture;
• concentrating the supernatant by nitrogen blowing and re-solubilising through 100 μL of acetonitrile to obtain a preprocessed sample;
• step 2:
• instrumental determination:
• performing a full-scan analytical detection on the preprocessed sample by a UPLC-Q-Exactive liquid-mass spectrometry instrument;
• wherein parameters of the UPLC-Q-Exactive liquid-mass spectrometry instrument are:
• chromatograph:
• Ultimate 3000 ultra-high performance liquid chromatography (Thermo fisher, USA);
• chromatograph column:
• C18 column (2.1 mm×50 mm, 2.5 μm);
• column temperature:
• 40° C.;
• flow rate:
• 0.3 mL/min;
• positive ion mode mobile phase:
• 0.1% formic acid-water solution (phase A), methanol (phase B):
• negative ion mode mobile phase:
• 2 mM ammonium acetate-aqueous solution (phase A), methanol (phase B);
• gradient elution table:
• Time (min)
• A %
• B %
• 1.00
• 90
• 10
• 1.00
• 90
• 10
• 36.00
• 0
• 100
• 50.00
• 0
• 100
• 50.00
• 90
• 10
• 55.00
• 90
• 10
• mass spectrometer:
• Q-Exactive electrostatic field orbitrap high resolution mass spectrometer;
• full scan mode:
• data dependent scanning mode;
• ion source:
• positive and negative electrospray ionisation source;
• full-scan mass range:
• MS1 80-1000 Da, MS2 50-800 Da;
• collision energy:
• ±35 eV;
• collision energy spread:
• 15 eV:
• ion source temperature:
• 412.5° C.;
• step 3:
• sample peaks extraction:
• saving results of a spectra of organic contaminants obtained from instrumental analysis in the step 2 as RAW files, converting to ABF files through a format converter and then importing into MSDIAL ver 4.48 software to extract sample peaks;
• step 4:
• contaminant identification:
• based on the sample peaks extracted in the step 3, targeting and identifying the organic contaminants by library matching, and/or identifying the organic contaminants not capable of being targeted and identified with molecular network tools.