Chiral-phase HPLC separation of (divinyl-)protochlorophyllide-a enantiomers as key precursors in chlorophyll biosynthesis from their 132-stereoisomeric prime forms
Chiral-phase HPLC separation of (divinyl-)protochlorophyllide-a enantiomers as key precursors in chlorophyll biosynthesis from their 132-stereoisomeric prime forms
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手性相 HPLC 分离（二乙烯基）原叶绿素内酯 -a 对映体（作为叶绿素生物合成中的关键前体）与其 132 立体异构体的主要形式
DOI：
10.1016/j.bbabio.2023.148960
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Tamiaki Hitoshi
中科院分区：
文献类型：
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作者：
Shiozaki Mai;Mizoguchi Tadashi;Harada Jiro;Hirose Mitsuaki;Tamiaki Hitoshi
原叶绿素a（Protochlorophyllide，PChlide）-a及其8-乙烯基化类似物二乙烯基（divinyl，DV）-PChlide-a是天然叶绿素（Chl）合成过程中常见的重要中间体。这些类卟啉型颜料在132位具有单个光学活性（不对称）碳原子，因此它们的立体异构体是（132 R）-和（132 S）-对映异构体。前者和后者分别称为（DV-）PClide-a和（DV-）PClide-a ′。在这项研究中，证明了对映体（DV-）PClides-a/a '的手性相HPLC分离。在本实验条件下，（132 R）-对映体PClide-a的洗脱速度比相应的（132 S）-对映体PClide-a ′慢。（132 R）-DV-PClide-a和（132 S）-DV-PClide-a′的洗脱顺序与PClides-a/a′相反。通过手性相HPLC分离各对映异构体后，通过圆二色性光谱表征在132位的立体异构构型。本手性相HPLC方法使我们能够评估（DV-）PClide-aspecies的光学纯度。例如，从培养的紫色光合细菌突变体的废培养基和收获的细胞中提取的PClide-a和/或DV-PClide-a，其中前者经常被用作酶促反应的（DV-）PClide-a底物的来源，被揭示大部分被外消旋化，得到（DV-）PClide-a/a′的对映体混合物。
Protochlorophyllide(PChlide)-aand its 8-vinylated analog, divinyl(DV)-PChlide-a, are common and essential intermediates in the biosynthesis of all naturally occurring chlorophyll (Chl) pigments. These porphyrinoid-type pigments have a single optically active (asymmetric) carbon atom at the 132-position, so their stereoisomers are (132R)- and (132S)-enantiomers. The former and latter are called (DV-)PChlide-aand (DV-)PChlide-a′, respectively. In this study, chiral-phase HPLC separation of enantiomeric (DV-)PChlides-a/a′ was demonstrated. The (132R)-enantiomeric PChlide-awas eluted more slowly than the corresponding (132S)-enantiomeric PChlide-a′ under the present HPLC conditions. On the other hand, the elution order of (132R)-DV-PChlide-aand (132S)-DV-PChlide-a′ was reverse to that of PChlides-a/a′. After the separation of each enantiomer by the chiral-phase HPLC, the stereoisomeric configuration at the 132-position was characterized by means of circular dichroism spectroscopy. The present chiral-phase HPLC method enables us to evaluate optical purities of (DV-)PChlide-aspecies. For example, PChlide-aand/or DV-PChlide-aextracted from the spent medium and harvested cells of cultured purple photosynthetic bacterial mutants, the former of which has been often used as the source of (DV-)PChlide-asubstrates for enzymatic reactions, were revealed to be mostly racemized, giving enantiomeric mixtures of (DV-)PChlides-a/a′.