山姜素与脱氧核糖核酸的相互识别研究

Study on the Recognition between Alpinetin and Calf Thymus DNA

应用荧光光谱法及紫外-可见光谱法研究了生理条件下(pH 7.4)山姜素(ALP)与DNA分子之间的相互识别。考察了不同温度下(25,32和39℃),DNA对山姜素荧光猝灭情况。实验发现,DNA能猝灭山姜素的内源性荧光,随着温度的升高,荧光猝灭常数KSV逐渐减小(KSV分别为3.288×103,2.923×103和2.467×103L·mol-1),并且DNA对山姜素的猝灭速率常数Kq要大于药物小分子与生物大分子之间的最大扩散所控制的碰撞猝灭常数,得出DNA对山姜素的荧光猝灭是单一的静态猝灭过程。DNA与山姜素相互作用紫外-可见光谱显示,DNA不能使得山姜素的吸收峰发生减色效应和红移现象,而山姜素也不能使溴化乙锭-DNA体系的荧光强度及最大荧光峰位置发生变化,即山姜素不和溴化乙锭竞争与DNA的结合位点。DNA热变性实验发现,解链DNA对山姜素的荧光猝灭程度要大于正常DNA的猝灭程度,由此推断山姜素与DNA不存在嵌插作用。同时,I-离子效应和盐效应表明,山姜素与DNA之间主要以沟槽模式相结合。

The recognition between alpinetin and DNA under physiological condition （pH 7. 4） was investigated by fluorescence and UV-visihle spectrometry. The experiment demonstrated that the fluorescence of alpinetin could be quenched by DNA. A quenching mechanism was proved to he the single static quenching procedure according to the deescalating, quenching constants Ksv（3. 288×10^3, 2. 923×10^3 and 2. 467×10^3 L · mol^-1, respectively） along with the escalating temperatures （25, 32 and 39 ℃） and the quenching rate constant Kq was greater than the maximum scatter collision quenching rate constant of various quenchers with the hiomolecule. From the UV-visihle spectra of alpinetin and DNA, the result showed that the UV-visihle spectra of alpinetin did not changed in the presence of DNA, i.e. neither the decrease in the maximum ahsorhance intensity nor the red shift of the maximum absorption wavelength changed. Both the fluorescence intensity and the maximum emission wavelength of ethidium hromide-DNA system remained unchanged in the presence of alpinetin, indicating that there is no direct competition for binding DNA between alpinetin and ethidium bromide. Furthermore, DNA thermal denaturation test indicated that the fluorescence quenching effect of alpinetin with unlinking DNA was stronger than that of alpinetin with natural DNA. It was concluded that there was not intercalation binding mode between alpinetin and DNA. At the same time, fluorescence quenching effect and salt effect on the binding of alpinetin with DNA were investigated. It was shown that the major mode of recognition was groove binding between alpinetin and DNA.