UC Davis

Increased urinary acetylated polyamines (APs) are reported as cancer biomarkers in many studies. N¹,N¹²-diacetylspermine has been proposed as a biomarker indicative of different cancers in urine and plasma. N¹-Acetylspermine has previously been found to be increased in the saliva of patients with breast cancer; however, in plasma this metabolite was too low abundant to be detected by previous analytical methods. In addition, no method has been reported to perform AP analysis on the level of speed, robustness and sensitivity required for daily clinical routines. Here we describe a high-throughput sample preparation and LC-MS/MS method for the fast, accurate and precise quantification of three APs: N⁸-acetylspermidine, N¹-acetylspermine, and N¹,N¹²-diacetylspermine in plasma, urine and saliva. Stable isotope labeled N¹,N¹²-diacetylspermine was used as internal standard. Robustness was validated by intra- and inter-day reproducibility. Precision and accuracy of the method were tested at six concentration levels from 0.0375 to 750 ng/mL resulting in less than 15% relative standard deviation and less than 15% percent error in quantification. Using 96-well plates, the assay described herein allows for preparing, analyzing, and quantifying 240 samples per day for a single researcher to quantify three APs commonly related to cancer status.