Possibility to Biotransform Anthracyclines by Peroxidases Produced by Bjerkandera adusta CCBAS 930 with Reduction of Geno- and Cytotoxicity and Pro-Oxidative Activity

The aim of this study was to evaluate the bioremoval mechanism of anthracycline antibiotics by the white-rot fungus B. adusta CCBAS 930. The activity of oxidoreductases and levels of phenolic compounds and free radicals were determined during the biotransformation of anthraquinone antibiotics: daunomycin (DNR) and doxorubicin (DOX) by B. adusta strain CCBAS 930. Moreover, phytotoxicity (Lepidium sativum L.), ecotoxicity (Vibrio fischeri), genotoxicity and cytotoxicity of anthraquinone dyes were evaluated before and after biological treatment. More than 80% and 90% of DNR and DOX were removed by biodegradation (decolorization). Initial solutions of DNR and DOX were characterized by eco-, phyto-, geno- and cytotoxicity. Despite efficient decolorization, secondary metabolites, toxic to bacteria, formed during biotransformation of anthracycline antibiotics in B. adusta CCBAS 930 cultures. DNR and DOX metabolites did not increase reactive oxygen species (ROS) production in human fibroblasts and resazurin reduction. DNR metabolites did not change caspase-3 activity.     

柔红霉素修饰的纳米金电极的制备及其对DNA检测

利用双硫醇分子作为连接剂, 将纳米金颗粒固定于金电极上, 用伏安法、紫外-可见光谱和电化学交流阻抗对其组装过程以及活性进行了表征. 制备的纳米金修饰电极用于DNA测定及其对DNA损伤的检测. DNA的检测限为 1.2×10^－9 mol/L. 该法灵敏、简便.     

Fluorescence Properties of Intercalating Neutral Chromophores in Complexes with Polynucleotides of Various Base Compositions and Secondary Structures

Changes in the relative quantum yield and polarization degree of steady-state fluorescence of daunomycin (DM) on its binding to six synthetic single- and double-stranded polynucleotides of various nucleotide compositions were measured over a wide range of molar polymer-to-dye ratios, in solutions of low ionic strength. Guanine base was found to be an effective quencher of DM fluorescence [in the DM–poly(G) complex the intensity of residual emission was ~0.5% of the free dye intensity]. The quenching of DM fluorescence by another purine base, i.e., adenine, was also revealed. But, unlike guanine, adenine exhibits quenching activity when it is in close contact with the DM chromophore, as realized in the complex with single-stranded poly(A). In intercalative complexes with double-stranded nucleic acids, where such contact is lacking, the quenching ability of adenine does not manifest itself, which has been demonstrated with the DM–poly(A) · poly(U) complex. It was found that the interaction with pyrimidine bases does not substantially change the DM quantum yield. The quenching feature of DM fluorescence is identical to that observed earlier by us for a glycoside phenazine dye containing, like DM, a neutral chromophore.     

Enhanced Efficiency of the Removal of Cytostatic Anthracycline Drugs Using Immobilized Mycelium of Bjerkandera adusta CCBAS 930

The aim of this study was to evaluate the bioremoval of anthracycline antibiotics (daunomycin-DNR, doxorubicin–DOX, and mitoxantrone-MTX) by immobilized mycelium of B. adusta CCBAS 930. The activity of oxidoreductases: versatile peroxidases (VP), superoxide dismutase (SOD), catalase (CAT), and glucose oxidase (GOX), and the levels of phenolic compounds (PhC) and free radicals (SOR) were determined during the biotransformation of anthracyclines by B. adusta strain CCBAS 930. Moreover, the phytotoxicity (Lepidium sativum L.), biotoxicity (MARA assay), and genotoxicity of anthracyclines were evaluated after biological treatment. After 120 h, more than 90% of anthracyclines were removed by the immobilized mycelium of B. adusta CCBAS 930. The effective biotransformation of anthracyclines was correlated with detoxification and reduced genotoxicity.     

Micelle-like macromolecular systems for controlled release of daunomycin

Soluble macromolecular conjugates for the delivery of the strongly hydrophobic anticancer drug daunomycin (DM) or rubomycin with its controlled release were prepared. The solution properties of these conjugates consisting of DM bonded to copolymer of maleic anhydride and divinyl ether (DIVEMA) and a few model compounds were investigated using adsorbance spectroscopy, as well as surface activity and solubilization of water-insoluble dye measurements. The data of these studies indicated that in water solutions conjugates are associated, probably intramolecularly. This micellization in parallel with an H-bonded ionic complex between DM and polymer carrier determines the DM release. It is concluded that the desirable drug release can be achieved through changing the structure of conjugates by means of varying the constituents hydrophobicity. © 1998 John Wiley & Sons, Ltd.     

Studies directed towards the total synthesis of anthracycline antibiotics

At present anthracycline antibiotics have proven to be the most exciting agents in cancer chemotherapy. Both adriamycin and daunomycin are proven to be effective against a variety of human tumour cells, despite their cardiotoxicity. However, some synthetic analogues, such as 4-demethoxydaunomycin, are shown to be better therapeutic ratios compared to adriamycin or daunomycin. Various approaches have been successfully made for the total synthesis of (±) 4-demethoxydaunomycinone, the aglycone of (±) 4=demethoxydaunomycin, starting from benzoquinone, napthalene or anthraquinone precursors. Finally an elegant approach for the stereoconvergent synthesis of (+) 4-demethoxydaunomycinone has been worked out. New methods involving both Diels-Alder and Friedal-Crafts acylation, have been developed for the total synthesis of daunomycinone and 11-deoxydaunomycinone. The synthesis of L-daunosamine, the amino sugar unit present in the antitumor anthracyclines has been successfully elaborated starting either from D-glucose or D-glucosamine.     

Structural and thermodynamic analysis of heteroassociation of daunomycin and flavin mononucleotide molecules in water by <Superscript>1</Superscript>H NMR spectroscopy

Heteroassociation of the antitumor antibiotic daunomycin (DAU) with flavin mononucleotide (FMN) has been investigated by one-and two-dimensional ¹H NMR spectroscopy (500 MHz) in a water solution to determine the molecular mechanism of the combined action of the antibiotic and vitamin in the FMN-DAU system. The equilibrium constants of the reactions, induced proton chemical shifts, and thermodynamic parameters (ΔH, ΔS) of heteroassociation were determined from the concentration and temperature dependences of the proton chemical shifts in the interacting aromatic molecules. Analysis of the results indicate that heterocomplexes of riboflavin mononucleotide and daunomycin are formed due to stacking interactions between aromatic chromophores. The most probable spatial structure of the 1:1 DAU-FMN heterocomplex was determined by the molecular dynamics method using the X-PLOR program and the results of the analysis of the induced proton chemical shifts in molecules. Calculation of the relative content of self-and hetero-complexes of daunomycin for different values of the ratio (r) between the concentrations of flavin mononucleotide and daunomycin demonstrated that for r > 3, the contribution of DAU-FMN heterocomplexes to the equilibrium distribution of associates in aqueous solution is dominant. It is concluded that the aromatic molecules of vitamins, in particular, riboflavin, can form energetically strong heteroassociates with antitumor antibiotics in water solution and can thereby affect their medical and biological activity.     

NIR Mega‐Stokes Fluorophores for Bioorthogonal Labeling and Energy Transfer Systems–An Efficient Quencher for Daunomycin

A set of new azide‐ and alkyne‐bearing lepidinium‐based fluorophores were synthesized for bioorthogonal labeling schemes. These fluorescent dyes all show large Stokes‐shifts with emission maxima in the near‐infrared (NIR) region of the electromagnetic spectrum. The applicability of these dyes in the construction of energy‐transfer systems was tested using one of these new fluorescent tags and daunomycin (Dau), an anticancer drug with fluorescent features. These daunomycin conjugates are the very first examples of fluorescently modulated constructs of this anticancer agent. The dually labeled architectures proved that the applied fluorescent dye can be utilized as an efficient quencher for daunomycin. Enzymatic cleavage of a dually labeled enzyme substrate resulted in full recovery of the fluorescence of daunomycin. Such fluorescently modulated Dau conjugates can provide useful information for the mechanism of action of Dau‐regulated cell death processes.     

柔红霉素与DNA相互作用的机理研究

本文以循环伏安、光谱电化学和原子力显微镜方法从DNA角度研究柔红霉素与天然鱼精DNA和热变性DNA之间相互作用的机理。并对柔红霉素与鱼精DNA和热变性DNA复合物的组成及复合物的形成常数作了测定。研究发现嵌入作用是柔红霉素和天然DNA之间的主要作用方式；并且柔红霉素和天然DNA之间的作用要强于和热变性单链DNA之间的作用。对这两种复合物的光谱电化学和原子力显微镜研究表明，在体内氧化还原代谢条件下，柔红霉素还原过程中产生的半醌自由基可引发自由基链反应，造成DNA链的解链、断裂等损伤。     

三维荧光二阶校正法用于血浆和尿液中柔红霉素的快速测定

文章采用三维激发发射荧光光谱与化学计量学交替三线性分解(ATLD)二阶校正法相结合,对血浆液和尿液中柔红霉素(DM)进行定量测定。实验不需对血浆和尿液预测样进行萃取等分离预处理,选取激发波长410～530nm,发射波长550～650nm,分别每隔5nm取一个数据,利用激发发射荧光扫描分别获得两个三维响应数阵(大小为21×25×12)。当组分数选择为3时,血浆和尿液校正集中盐酸柔红霉素的相对浓度与实际浓度的相关系数分别为r1=0.9990和r2=0.9952,经ATLD算法解析得到的血浆和尿液预测样中柔红霉素平均回收率分别为(92.8±7.6)%和(94.7±4.4)%。实验结果表明,此法能够解决血浆和尿液中盐酸柔红霉素药物因血浆和尿液内源物质与分析物光谱重叠所引起的难分辨的问题,可用于未知干扰共存下柔红霉素的直接快速定量测定。