On-line HPLC-DAD coupled with ESI-IT-TOF-MS and fluorescence detection to identify DNA-binding compounds from Lithospermum erythrorhizon using acridine orange as the fluorescence probe

doi:10.5246/jcps.2015.09.074

Abstract

Abstract:

We have developed an on-line detection method using acridine orange as the fluorescence probe and applied this method to rapidly identify active compounds in herbal medicines. This on-line method was equipped with a high-performance liquid chromatography tandem diode array detector, electrospray ionization-ion-trap time-of-flight mass spectrometry and DNA-acridine orange fluorescence detection (HPLC-DAD-MSⁿ-DNA-AO-FLD). A large amount of information could be simultaneouslyobtained during one run, which included HPLC fingerprint, ultraviolet spectra, total ion chromatograms, MSⁿ data of high-resolution mass spectrometry and activity proﬁle of each compound binding with DNA. The method also provided information on structure-activity relationships and mechanism of interaction. We used this on-line method to identify five DNA-binding activity components from Lithospermum erythrorhizon sample for the first time. The result showed that the parent nucleus of shikonin derivatives could bind with DNA. The structure-activity relationship showed that the parent nucleus of shikonin derivatives plays a major role in DNA binding, not the carboxyl group on the side chain. This simple, rapid, high precision and good stability on-line method should be useful for compound separation, structural identification and screening of DNA-binding compounds in herbal medicines.

Key words: On-line, Lithospermum erythrorhizon, Acridine orange, DNA, DNA-binding activity

CLC Number:

R284

Supporting:

Wenfang Huang, Cangman Zhang, Sensen Li, Yi Liang, Hong Wang, Shizhong Chen. On-line HPLC-DAD coupled with ESI-IT-TOF-MS and fluorescence detection to identify DNA-binding compounds from Lithospermum erythrorhizon using acridine orange as the fluorescence probe[J]. Journal of Chinese Pharmaceutical Sciences, 2015, 24(9): 581-590.

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