Simultaneous Determination of 34 Amino Acids in Tumor Tissues from Colorectal Cancer Patients Based on the Targeted UHPLC-MS/MS Method

A targeted ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was established and validated for the simultaneous determination of 34 amino acids in tissue samples from colorectal cancer (CRC) patients. The chromatographic separation was achieved on an Agilent ZORBAX SB-C column (3.0 × 150 mm, 5 m) with a binary gradient elution system (A, 0.02% heptafluorobutyric acid and 0.2% formic acid in water, v/v; B, methanol). The run time was 10 min. The multiple reaction monitoring mode was chosen with an electrospray ionization source operating in the positive ionization mode for data acquisition. The linear correlation coefficients were >0.99 for all the analytes in their corresponding calibration ranges. The sample was pretreated based on tissue homogenate and protein precipitation with a 100 mg aliquot sample. The average recovery and matrix effect for 34 amino acids and 3 internal standards were 39.00%∼146.95% and 49.45%∼173.63%, respectively. The intra- and interday accuracy for all the analytes ranged from -13.52% to 14.21% (RSD ≤8.57%) and from -14.52% to 12.59% (RSD ≤10.31%), respectively. Deviations of stability under different conditions were within ±15% for all the analytes. This method was applied to simultaneous quantification of 34 amino acids in tissue samples from 94 CRC patients.