High-performance liquid chromatography estimation of cross-linking of dihydroxyindole moiety in eumelanin
High-performance liquid chromatography estimation of cross-linking of dihydroxyindole moiety in eumelanin
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DOI：
10.1016/j.ab.2012.12.005
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发表时间：
2013-03-15
影响因子：
2.9
通讯作者：
Simon, John D.
中科院分区：
文献类型：
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作者：
Ito, Shosuke;Wakamatsu, Kazumasa;Simon, John D.
真黑素色素由不同比例的5，6-二羟基吲哚-2-羧酸（DHICA）和5，6-二羟基吲哚（DHI）组成。在碱性过氧化氢氧化下，这些吲哚部分分别产生吡咯-2，3，5-三羧酸（PTCA）和吡咯-2，3-二羧酸（PDCA）。在最近的一项研究中，我们在超过1.6亿年前的化石墨囊的氧化产物中检测到了相当数量的其他吡咯酸，吡咯-2，3，4，5-四羧酸（PTeCA）和吡咯-2，3，4-三羧酸（isoPTCA）。PTeCA和isoPTCA由真黑素的DHI部分在C2和/或C3位置的交联产生。我们通过在100摄氏度下加热由各种比例的DHICA和DHI制备的合成真黑素18天（或在40摄氏度下加热180天）来模拟交联过程。在碱性过氧化物氧化后，通过具有紫外（UV）检测的高效液相色谱（HPLC）将加热的真黑素分析为PTCA、PDCA、PTeCA和isoPTCA。加热时，由于脱羧作用，PTCA迅速下降，而PDCA逐渐下降。同时，PTeCA逐渐增加至接近PTCA的水平。IsoPTCA在较低水平也逐渐增加。在40摄氏度下以慢得多的速度观察到类似的变化。这些发现表明，PTeCA/PTCA比率可以作为真黑素老化（交联）的良好指标。(c)2012 Elsevier Inc. All rights reserved.
Eumelanin pigments consist of various ratios of 5,6-dihydroxyindole-2-carboxylic acid (DHICA) and 5,6-dihydroxyindole (DHI). On alkaline hydrogen peroxide oxidation, these indole moieties give rise to pyrrole-2,3,5-tricarboxylic acid (PTCA) and pyrrole-2,3-dicarboxylic acid (PDCA), respectively. In a recent study, we detected considerable amounts of other pyrrole acids, pyrrole-2,3,4,5-tetracarboxylic acid (PTeCA) and pyrrole-2,3,4-tricarboxylic acid (isoPTCA), among the oxidation products of fossil ink sacs more than 160 million years old. PTeCA and isoPTCA arise from the cross-linking of the DHI moiety of eumelanin at the C2 and/or C3 positions. We mimicked the process of cross-linking by heating synthetic eumelanins prepared from various ratios of DHICA and DHI at 100 degrees C for 18 days (or at 40 degrees C for 180 days). The heated eumelanins were analyzed after alkaline peroxide oxidation as PTCA, PDCA, PTeCA, and isoPTCA by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection. On heating, PTCA decreased rapidly due to decarboxylation, whereas PDCA decreased gradually. Concurrently, PTeCA increased gradually to levels close to PTCA. IsoPTCA also increased gradually at lower levels. Similar changes were observed at 40 degrees C at a much slower rate. These findings suggest that the PTeCA/PTCA ratio may serve as a good indicator of aging (cross-linking) of eumelanin. (c) 2012 Elsevier Inc. All rights reserved.