High performance liquid chromatography separation and simultaneous fluorometric detection of trace amount of polyamines in cell culture media.

A highly sensitive and precise method for the determination of the polyamines (including putrescine, spermidine and spermine) in cell culture media is described. The samples were concentrated by C18 column and the polyamines were converted to fluorometric derivatives with DNS-Cl. The polyamine derivatives were then completely separated by HPLC and determined by simultaneous fluorometric detection. The CV of intragroup and intergroup were 2.49%-4.26% and 4.29%-5.16%, respectively. The rate of recovery was 103%-99%. There was trace amount of polyamines detected by this method in the media of F12, 8900, RPMI-1640, DMEM and M199 even without incubation with cells. So this method can be used for detecting the changes of polyamines in a medium before and after incubation with cells. It is helpful for the researches on the regulation of cell proliferation by polyamines.