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of the 0.3 M perchloric acid extract containing a known amount of 1,6-hexanediamine (IS). Tubes were
closed with caps with PTFE-coated rubber closure, and the contents were mixed vigorously and
incubated for 30 min at room temperature under gentle rotation. The resulting solution was extracted
with 2 ml of CHCl . After centrifugation the CHCl layer was removed, washed with 2 ml of water,
3
3
centrifuged and transferred to another tube. The solvent was evaporated under a stream of air and the
residue was dissolved in 100-500 µ1 of methanol. Typical chromatograms were given for benzoylated
polyamines from tumor tissue grown in nude mice (Fig. 2.11).

2.2.5.2—
Dansyl Chloride (DNS-Cl)

Dansyl chloride is used in the derivatization of primary and secondary amines in which fluorimetric
detection of dansyl derivatives is usually performed [63]. UV-detection of dansylated compounds is not
a common procedure except on some occasions. The fluorescence of the dansyl amino acids is
quenched in aqueous solutions and the quantum yield obtained is about 1/10th that in organic solvents.
In an RP-HPLC of dansyl amino acids [64], the necessity of an aqueous buffer for elution, which
quenches fluorescence, the UV absorbance of the dansyi amino acids at 250 nm, is used for detection
giving a sensitivity of about 100 pmol for a single dansyl amino acid. A rapid HPLC method for the
determination of the dansyl derivative of pentaazapentacosane (PAPC, Fig. 2.12) pentahydrochloride,
an anticancer agent, has been developed [65].

Like spermine and spermidine, PAPC does not absorb in the UV region and therefore, it must be
derivatized. In the chromatographic system,

Fig. 2.11.
Chromatograms of benzoylated polyamines from
PC-93 tumour tissue grown in nude mice, monitored
by their absorbance at 229 nm. Elution with

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