Synthesis of a Novel Fluorescence Probe Modified by β‐Cyclodextrin and Its Application in the Determination for Nucleic Acids at ppb Levels

Abstract

A new fluorescence probe, mono[6‐N(4‐carboxy‐phenyl)]‐β‐cyclodextrin (ACD), was synthesized. p‐Aminobenzoic acid (PA) with a emission peak located in 351 nm (λ_ex = 286 nm) is not suitable for the quantitative determination of nucleic acids because of its background interference. It is necessary to change its fluorescence properties as to be applied in biochemical analysis by chemical modification. In this article, PA modified by β‐cyclodextrin was synthesized. The excitation peak of the new compound (ACD) moves to 289 nm and the emission peak moves to 355 nm, in addition, a new long‐wavelength emission peak located in 689 nm appeared. It is interesting that the fluorescence intensity of ACD (λ_ex = 289 nm, λem = 689 nm) was decreased strongly when appropriate amounts of nucleic acids was added. The spectrofluorimetric determination of trace amounts of nucleic acids based on the phenomenon was carried out. Upon optimal conditions, the fluorescence intensities vary linearly with the concentration of nucleic acids. The linear range was 0.0–3.6 µg/mL for ct‐DNA, 0.05–2.0 µg/mL for fs‐DNA and 0.05–5.0 µg/mL for yeast RNA, respectively. The detection limit (3σ) was 6.1 ng/mL for ct‐DNA, 10.8 ng/mL for fs‐DNA, and 9.8 ng/mL for yeast RNA, respectively. The interferences of some substances were described. β‐CD was ordinarily used as a sensitizing reagent in analytical chemistry. But quantitative analytical application of fluorescence probe modified by β‐CD has not been reported. ACD was easily synthesized and the method is a sensitive, rapid, and simple analytical procedure for the determination of nucleic acids.

Keywords:

• Nucleic acid determination
• Fluorescence quenchment
• β‐Cyclodextrin
• p‐Aminobenzoic acid

Acknowledgment

This work was supported by the Natural Science Foundation (2003kj144) and Youth Teacher Foundation (2003jq104) of Educational Committee of Anhui Province, and by the Youth Teacher Foundation of Anhui Normal University.