Jožef Stefan Institute, Jamova 39, 1000, Ljubljana, Slovenia.
Biometals. 2012 Feb;25(1):103-13. doi: 10.1007/s10534-011-9486-6. Epub 2011 Aug 7.
The ability of As(2)O(3) to induce apoptosis in various malignant cell lines has made it a potential treatment agent for several malignancies. In this study the chemical stability of As(2)O(3) (As(III)) in cell-free growth media with various compositions was studied (MEM with different amount of amino acids and DMEM). Special attention was given to evaluate the influence of serum (FBS; fetal bovine serum) absence and vitamin C addition on the oxidation of As(III) to As(V) in cell-free growth media. FBS is an important source of antioxidants and vitamin C (ascorbic acid) is acting as a prooxidant in millimolar concentrations. Media were incubated with As(III) (0.6, 2 and 7 μmol l(-1)) up to 72 h. Experiments were performed at 37°C in light or/and in the dark, with or without added serum (10%) or vitamin C (1.4, 0.14 mM). Metabolites were followed with high-performance liquid chromatography directly coupled to a hydride generation-atomic fluorescence spectrometry system. After 72 h up to 30% of As(III) was transformed into As(V) in MEMs and up to 35% in DMEM when exposed in dark. Light had no influence on transformations in MEMs, but changed the situation dramatically in DMEM where almost all As(III) was oxidized to As(V) after 72 h when exposed to light. Except for some faster oxidation rate the absence of FBS had little effect on the transformation rate in all media. The most visible impact on As(III) oxidation was observed by addition of vitamin C. Addition of vitamin C (1.4 mM) transformed almost all As(III) to As(V) within 72 h. In lower concentrations (0.14 mM) a pro-oxidative effect was still observed reaching approximately 60% oxidation of As(III) during 72 h. All oxidation processes could be explained by pseudo first order reaction kinetics, yielding reaction rates increasing with initial As(III) concentration and vitamin C concentration whereas the FBS content additionally increased the As(III) oxidation rate in the DMEM (light). The temporal oxidation of As(III) to As(V) in various cell-free growth media necessitates routine checking of the valence state of arsenic during cell culture experiments and the results of biological effects attributed to As(III) should be interpreted with caution. Special attention is needed particularly in cases with vitamin C which was acting pro-oxidatively in all conditions examined.
三氧化二砷（As(2)O(3)）能够诱导多种恶性细胞系凋亡，使其成为几种恶性肿瘤的潜在治疗药物。在这项研究中，研究了无细胞生长培养基中各种成分（含不同量氨基酸的 MEM 和 DMEM）对 As(2)O(3)（As(III)）化学稳定性的影响。特别关注了在无细胞生长培养基中评估血清（FBS；胎牛血清）缺乏和维生素 C 添加对 As(III)氧化为 As(V)的影响。FBS 是抗氧化剂的重要来源，而维生素 C（抗坏血酸）在毫摩尔浓度下表现为促氧化剂。将 As(III)（0.6、2 和 7μmol l(-1)）培养至 72 h。在 37°C 下进行实验，在光或/和黑暗中，有无添加血清（10%）或维生素 C（1.4、0.14 mM）。使用高效液相色谱法直接与氢化物发生-原子荧光光谱系统联用，跟踪代谢产物。72 h 后，在 MEM 中高达 30%的 As(III)转化为 As(V)，在 DMEM 中高达 35%。黑暗中，光线对 MEM 中的转化没有影响，但在 DMEM 中情况发生了巨大变化，在 DMEM 中，当暴露在光下时，几乎所有的 As(III)在 72 h 后都被氧化为 As(V)。除了一些更快的氧化速率外，在所有培养基中，缺乏 FBS 对转化速率几乎没有影响。添加维生素 C 对 As(III)氧化的影响最为明显。添加维生素 C（1.4 mM）可在 72 h 内将几乎所有的 As(III)转化为 As(V)。在较低浓度（0.14 mM）下，仍观察到促氧化作用，在 72 h 内，As(III)的氧化率达到约 60%。所有的氧化过程都可以用准一级反应动力学来解释，随着初始 As(III)浓度和维生素 C 浓度的增加，反应速率增加，而 FBS 含量在光照下的 DMEM 中进一步增加了 As(III)的氧化速率。在各种无细胞生长培养基中，As(III)向 As(V)的时变氧化需要在细胞培养实验过程中定期检查砷的价态，并且应该谨慎解释归因于 As(III)的生物学效应的结果。在所有检查的情况下，特别是在维生素 C 起促氧化作用的情况下，需要特别注意。
