Fluorescent cisplatin analogues and cytotoxic activity.

Cisplatin is one of the chemotherapeutic agents used the most for testicular, ovarian and several other cancers. In order to overcome cisplatin resistance, other platinum (Pt) compounds have been developed and, in the last ten years, Pt-derivatives with reporting activity have also been synthesized. The first generation of reporting Pt-compounds was based on linking a fluorescent molecule (e.g. cyanine) to cisplatin, but more recent studies have focused on strategies to synthesize intrinsically fluorescent derivatives. Accordingly, bile acid Pt-compounds have shown fluorescence intensity that is stable at room temperature for a long time; this fluorescence is maintained after binding to oligonucleotides or DNA. Because of this, the binding mode of these compounds to DNA can be easily analyzed both by flow injection and fluorescence techniques, showing that although these compounds target the nuclei, they form adducts with the DNA that are different from those due to cisplatin. In line with this, these bile acid derivatives have shown increased cytotoxicity and ability to overcome resistance as compared to cisplatin in several cell lines. Moreover, in contrast to cisplatin, the activity of these compounds does not seem to be restricted to cycling cells but they also seem to kill resting cells. This review summarizes the information available on reporting Pt-compounds and focuses on these novel, intrinsically fluorescent bile acid Pt derivatives, their biochemical characteristics and biological activity.

New fluorescent antitumour cisplatin analogue complexes. Study of the characteristics of their binding to DNA by flow injection analysis.

The flow injection technique is applied to study the binding to DNA of new platinum complexes-E(1): ethylenediaminechlorocholylglycinateplatinum(II): [PtCl(CG)(en)], C(54)H(92)O(12)Pt and E(2): ethylenediaminebischolylglycinateplatinum(II): [Pt(CG)(2)(en)], C(28)H(50)ClN(3)O(6)Pt-derived from cisplatin in which the exchangeable ligands were replaced by bile acids, such that these anticancer drugs have less toxicity and less resistance is developed towards them. Both compounds are fluorescent and their fluorescence is enhanced when they form adducts with DNA, a property that is extremely useful for monitoring the cytotoxic activity and their mechanisms of action. The binding parameters to DNA of E(1) [apparent intrinsic binding constant K(E1): (11.2 +/- 0.4) x 10(3) M(-1) and maximum number of binding sites per nucleotide, n (E1): 0.121 +/- 2 x 10(-3)) and E(2) (K(E2): 9.2 +/- 0.7) x 10(3) M(-1) and n (E2) 0.098 +/- 2 x 10(-3)] were determined following the Scatchard method and the type of binding was studied experimentally through the modifications introduced by each of the compounds into the ethidium bromide-DNA bond.

Determination of the DNA-binding characteristics of ethidium bromide, proflavine, and cisplatin by flow injection analysis: usefulness in studies on antitumor drugs.

Flow injection analysis was used to study the reactions occurring between DNA and certain compounds that bind to its double helix, deforming this and even breaking it, such that some of them (e.g., cisplatin) are endowed with antitumoral activity. Use of this technique in the merging zones and stopped-flow modes afforded data on the binding parameters and the kinetic characteristics of the process. The first compound studied was ethidium bromide (EtdBr), used as a fluorescent marker because its fluorescence is enhanced when it binds to DNA. The DNA-EtdBr binding parameters, the apparent intrinsic binding constant (0.31+/-0.02 microM(-1)), and the maximum number of binding sites per nucleotide (0.327+/-0.009) were determined. The modification introduced in these parameters by the presence of proflavine (Prf), a classic competitive inhibitor of the binding of EtdBr to the DNA double helix, was also studied, determining the value of the intrinsic binding constant of Prf (K(Prf) = 0.119+/-9x10(-3) microM(-1)). Finally, we determined the binding parameters between DNA and EtdBr in the presence of the antitumor agent cisplatin, a noncompetitive inhibitor of such binding. This provided information about the binding mechanism as well as the duration and activity of the binding of the compound in its pharmacological use.

Flow-injection solvent extraction without phase separation. Fluorimetric determination of thiamine by the thiochrome method.

Two modes of liquid-liquid extraction in flow-injection systems were compared and applied to the fluorimetric determination of thiamine (Vitamin B(1)). The first included phase segmentation, but fluorescence was measured without phase separation. In this mode, thiamine was detected at concentrations higher than 8 microg/l with a linear application range of 30-2,000 microg/l, an R.S.D. of 1.9% (150 microg/l, n=10) and a sampling frequency of 60/h. In the second mode, a single segment of organic solution was injected into the aqueous stream and fluorescence was also measured without phase separation. Using this mode, concentrations of thiamine higher than 1 microg/l were detected, with a linear application range between 5 and 280 microg/l, an R.S.D. of 2.4% (150 microg/l, n=10) and a sampling frequency of 60/h. The two forms were applied to the analysis of thiamine in pharmaceuticals.