RESUMO
An automated on-line solid phase extraction-liquid chromatography-tandem mass spectrometry ( on-line SPE-LC-MS/MS ) method for the simultaneous quantification of six endogenous phytohormones including abscisic acid ( ABA ) , indole-3-acetic acid ( IAA ) , salicylic acid ( SA ) , jasmonic acid ( JA ) , indole-3-propionic acid ( IPA) and indole-3-butyric acid ( IBA) in rice tissues was described. Plant samples were extracted with methanol and on-line SPE procedure was performed with C18 SPE cartridges. Analytes were eluted to C18 analytical column with mobile phase and further analyzed by LC-MS/MS. The performance of the method was fully validated. Linearity showed good correlation (R2≥0. 99). Limits of detections (S/N=3) were in the range of 0. 1-0. 8 μg/kg. Recoveries varied between 71. 2% and 126%. The method was rapid and sensitive to determine multiple phytohormones in rice young panicles and the results were cross-validated with the off-line SPE-LC-MS/MS phytohormone analytical method. Finally, the method was applied to monitor the changes of ABA, IAA, SA and JA in wounded rice leaves. The trends were in accord with plant physiological processes.
RESUMO
An on-line solid phase extraction ( SPE ) coupled with HPLC-MS/MS method was developed to determine S-ammuxetine and R-ammuxetine in rat plasma. The sample preparation consisted of the following steps:A protein precipitation extraction used methanol and acetonitrile ( 50:50 , V/V ); an on-line SPE treatment to remove most matrixes in plasma;an enrichment and separation step used a C18 analytical column. S-and R-ammuxetine were determined by tandem mass spectrometry. The SPE column was a Retain PEP Javelin (10 mm × 2. 1 mm × 5 μm), while the chromatographic separation was achieved using a ZORBAX SB-C18 (50 mm × 2. 1 mm × 3. 5 μm) analytical column with an isocratic mobile phase composed of acetonitrile-water-formic acid (40:60:0. 1, V/V/V, 0. 3 mL/min). The selected reaction monitoring mode of the positive ion was performed and the precursor to the product ion transitions of m/z 292 . 1/154 . 0 and m/z 260. 4/116. 2 were used to measure S-ammuxetine, R-ammuxetine and internal standard (propranolol). The method was linear over a concentration range from 0 . 2 to 1000 μg/L with the correlation coefficients of 0 . 9903 and 0 . 9951 . The average intra-day precision values were 1 . 2% -12 . 0% for S-ammuxetine and 0. 4%-11. 2% for R-ammuxetine, respectively. The average recoveries were 94. 2%-101. 6% for S-ammuxetine and 94. 3% -109. 4% for R-ammuxetine. Compared to the literature, the sensitivity of this method increased dramatically. The present method has been successfully applied to the preclinical rat research of ammuxetine isomers following intragastric administration.