Antioxidant vs. Prooxidant Properties of the Flavonoid,Kaempferol, in the Presence of Cu(II) Ions: A ROS-Scavenging Activity,Fenton Reaction and DNA Damage Study

Kaempferol is a flavonoid that occurs in tea and in many vegetables and fruits, including broccoli, cabbage, beans, grapes, apples, and strawberries. The efficacy of Kaempferol has been demonstrated in the treatment of breast, esophageal, cervical, ovarian, and liver cancers and leukemia, which very likely arises from its prooxidant properties and the activation of pro-apoptotic pathways. Indeed, this matter has already been the focus of a number of published studies and reviews. The aim of the present study was to elucidate the antioxidant vs. prooxidant properties of flavonoids in the presence of the redox-active metal, copper (II) ion, by means of the Fenton reaction. The specific motivation of this work is that, since an increased level of Cu(II) ions is known to be associated with many disease states such as neurological conditions (Alzheimer’s disease) and cancer, any interaction between these ions and flavonoids might affect the outcome of therapeutic uses of the latter. The structure of the Cu-kaempferol complex in DMSO was investigated by means of low temperature EPR spectroscopy, which confirmed the existence of at least two distinct coordination environments around the copper (II) ion. UV vis-spectra of kaempferol and its Cu(II) complex in DMSO revealed an interaction between the 5-OH (A ring) group and the 4-CO (C ring) group of kaempferol with Cu(II) ions. An ABTS assay confirmed that kaempferol acted as an effective radical scavenger, and that this effect was further enhanced in the form of the Cu(II)-kaempferol complex. Quantitative EPR spin trapping experiments, using DMPO as the spin trap, confirmed suppression of the formation of a mixture of hydroxyl, superoxide, and methyl radicals, in a Fenton reaction system, upon coordination of kaempferol to the redox-active Cu(II) ions, by 80% with respect to the free Cu(II) ions. A viscometric study revealed a better DNA-intercalating ability of the Cu-kaempferol complex than for free kaempferol, essential for conferring anticancer activity of these substances. The results of the viscometric measurements were compared with those from a DNA damage study of Cu-kaempferol complexes in a Fenton reaction system, using gel electrophoresis. At low concentrations of kaempferol (Cu–kaempferol ratios of 1:1 and 1:2), a very weak protective effect on DNA was noted, whereas when kaempferol was present in excess, a significant DNA-protective effect was found. This can be explained if the weakly intercalated kaempferol molecules present at the surface of DNA provide protection against attack by ROS that originate from the Fenton reaction involving intercalated Cu(II)-kaempferol complexes. Following the application of ROS scavengers, L-histidine, DMSO, and SOD, gel electrophoresis confirmed the formation of singlet oxygen, hydroxyl radicals, and superoxide radical anions, respectively. We propose that the prooxidant properties of Cu-kaempferol complexes may provide anticancer activity of these substances. When present in excess, kaempferol displays antioxidant properties under Cu-Fenton conditions. This suggests that kaempferol might prove a suitable candidate for the prevention or treatment of oxidative stress related medical conditions that involve a disturbed metabolism of redox metals such as copper, for example, Menkes disease, and neurological disorders, including Alzheimer’s disease. For the potential use of kaempferol in clinical practice, it will be necessary to optimize the dose size and critical age of the patient so that this flavonoid may be beneficial as a preventive drug against cancer and neurological disorders.     

和田菟丝子在不同寄主中有效成分含量的测定研究

采用高效液相色谱法(HPLC)对和田菟丝子中槲皮素和山萘酚进行了定量测定。测定寄主为胡麻植物的菟丝子中槲皮素含量为0 239 mg·g-1,山萘酚的含量为0 347 mg·g-1 ;寄主为苜蓿的菟丝子中槲皮素含量为0 103 mg·g-1,山萘酚的含量为0 068 mg·g-1。和田菟丝子寄主不同,其槲皮素和山萘酚的含量不同,应结合寄主植物研究其有效成分。     

山奈酚分子印迹聚合物的制备及其性能表征

采用分子印迹技术,以山奈酚(Kaempferol,KAE)为模板分子,2-乙烯基吡啶(2-Vpy)为功能单体,乙二醇二甲基丙烯酸酯(EDMA)为交联剂,合成了山奈酚分子印迹聚合物(KAE-MIP)。采用静态平衡结合实验评价了KAE-MIP对底物分子的结合特性,并进行了吸附动力学研究。结果表明,KAE-MIP对KAE呈现出了很高的选择吸附特性;Scatchard分析显示KAE-MIP对KAE存在不同亲和力的两类结合位点;与对照物在KAE-MIP上的吸附行为比较表明,KAE-MIP对KAE具有良好的分子识别性能;KAE-MIP对KAE的选择性吸附基本在2 h之内完成。     

野蔷薇中山奈酚含量测定方法的研究

建立了一种简单、快速测定野蔷薇果中黄酮类化合物山奈酚含量的反相液相色谱方法。利用乙腈、水(磷酸调pH为3.0)以1mL/min流速梯度洗脱,在360nm检测波长下,山奈酚在2.5min内即可达到基线分离。经与标准品对照,野蔷薇果中含有山奈酚,含量为4.3μg/g干质量。经测定:山奈酚回收率较好,为94.5%;相对标准偏差小于5%,方法重复性较好,可用于野蔷薇果中黄酮类化合物山奈酚的快速分析检测。以及测量山奈酚含量的分光光度法和高效液相色谱法进了比较研究,结果表明,两种方法测定的山奈酚浓度值差异不大,结果稳定可靠。分光光度法操作简单方便,测定结果略微偏高,HPLC法重复性、稳定性均较好。     

Preparation and characterization of monodisperse molecularly imprinted polymer microspheres by precipitation polymerization for kaempferol

A new kind of molecularly imprinted polymer (MIP) microspheres for the selective extraction of kaempferol was prepared by precipitation polymerization using 4-vinylpridine (4-VP) and ethylene glycol dimethacrylate (EDMA) as functional monomer and cross-linker respectively. The synthesis conditions, such as ratios of 4-VP/EDMA and polymerization time were discussed in detail. Results showed that the 2% was the optimal concentration of co-monomers to obtain monodisperse MIP microspheres, the best ratio of 4-VP/EDMA was 1:2, and 24 h was considered as the proper polymerization time. Compared with the MIP agglomeration or coagulum particles, monodisperse MIP microspheres showed the better adsorption capacity: the saturated adsorption capacity of monodisperse MIP microspheres was 7.47 mg g^?1, the adsorption equilibrium could be obtained in 30 min. Finally, the adsorption performances of the optimal MIP microspheres were evaluated by kinetic adsorption, adsorption isotherm, and selective adsorption experiments, which indicated that the adsorption mechanism were chemical single layer adsorption and the separation factor was up to 3.91 by comparing with the structure similar compound (quercetin). The MIP microspheres exhibit prospects in the kaempferol efficient and selective separation.     

北京洋槐蜜化学成分及其抗氧化活性

采用层析法富集北京洋槐蜜中抗氧化成分,利用高效液相测定其化学组成并研究其中已知化学成分对DPPH自由基的清除作用,旨在评价北京洋槐蜜的抗氧化功能。结果显示：北京洋槐蜜中含有绿原酸、咖啡酸、对香豆酸、山奈酚、异鼠李素、松属素、白杨素和高良姜素8种已知成分,含量在9.5～257.8μg/100g之间,除松属素和白杨素外,其余各成分均有清除DPPH自由基的能力,且差异显著(P＜0.01)。从有效成含量及抗氧化能力综合考虑,山奈酚可视为北京洋槐蜜中最有代表性的抗氧化成分。     

HPLC测定过山厥提取物中山柰素含量

目的 建立高效液相色谱法测定过山厥提取物中山柰素含量.方法 Hypersil ODS2(4.6 mm×200 mm,5μm)色谱柱,甲醇-0.07％磷酸(50∶50)为流动相,检测波长268 nm,流速1.0 mL/min,柱温25℃.结果 山柰素浓度在4.99～12.98mg/L线性关系良好,回归方程为Y=58 735X-44 779 (R2=0.9983);平均回收率99.8％,RSD=0.97％;重复性良好,RSD=1.14％(n=5).结论 该方法简单、可靠、专属性强,可用于过山厥提取物中山柰素的含量测定.     

槲皮苷和山奈酚对糖尿病小鼠血糖及血脂水平的影响

本研究对黄酮单体山奈酚、槲皮苷对糖尿病小鼠降血糖、血脂效果进行试验,建立了小鼠实验模型,经喂养后测定结果表明,黄酮单体山奈酚、槲皮苷能够有效的降低小鼠的血糖BG、TG、TC和MDA水平,对增加血清SOD、GSH-Px活性也有明显促进作用,其中以10 mg/kg.bw.d为最佳剂量。这表明黄酮单体山奈酚、槲皮苷对糖尿病防治有明显作用。     

超微粉碎技术对油菜花粉中槲皮素和山奈素溶出率的影响

为了探讨超微粉碎技术对油菜花粉中有效成分(槲皮素和山奈素)溶出的影响,采用高效液相色谱法分析比较油菜花粉超微粉与普通粉(65～100 目)中槲皮素和山奈素的溶出差异。结果表明,油菜花粉超微粉中槲皮素和山奈素的溶出率比普通粉分别提高了45.16% 和27.86%,超微粉碎可显著提高油菜花粉有效成分的溶出率。