Objective: To develop a robust liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for determination of 5-fluorouracil (5-FU) in blood plasma and evaluate the use of 5-FU for treatment response surveillance as well as toxicity prediction in malignant gastrointestinal tumors.
Methods: A LC-MS/MS method was used, with signal linearity, lower limits of quantitation, precision, accuracy and stability being evaluated according to guideline US Food and Drug Administration (FDA)'s guidance for industry bioanalytical method validation.Analysis of 5-FU was performed in 35 gastrointestinal cancer patients admitted to Zhongshan Hospital of Fudan University from April 2013 to December 2013. The relationship between 5-FU with toxicity and treatment effect was compared.
Results: The linear ranges of 5-FU was 49-9 800 ng/ml, the lower limit of quantitation was 49.0 ng/ml. The within-run and between-run coefficients of variation (CV) of 5-FU was <3% and <6%.The recovery rates of low, medium and high level quality controls were 103.36%, 88.12% and 91.26% respectively; with standard added recovery of 109.69%, 91.06% and 88.81% respectively. The control added recoveries were 112.16%, 99.12% and 92.28% respectively. The bias was -11.69%, 2.42% and -8.09% when samples were repeated freezing and thawing twice (-80 ℃). The results had a bias -11.97%, 1.42%, -10.91% and 0.56%, 0.14%, 3.82% when samples were kept at 2-8 ℃ for 2 days and 14 days. In 35 gastrointestinal cancer patients, there was no correlation between initial dose of 5-FU and 44 h AUC (concentration 3.44-53.43 mg/L·h). The risk of chemotherapy-related adverse effect in 5-Fu 44 h AUC> 30 mg/L·h group was significantly higher than 44 h AUC < 30 mg/L·h group (χ(2)=12.600, P<0.01). While the chemotherapy response of AUC > 20 mg/L·h group was significantly satisfactory than AUC < 20 mg/L·h group (χ(2)=5.358, P<0.05).
Conclusions: A robust and reliable LC-MS/MS method for the determination of 5-FU in blood plasma has been developed and it is suitable for clinical application. Detecting 5-FU may guide individualized treatment and predict adverse events.