Slantingly cross loading sample system enables simultaneous performance of separation and mixture to detect molecular interactions on thin-layer chromatography

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• 作者：Hiroko Shimizu-Yumoto^a ; ^{hirokosh@affrc.go.jp} ; Nobuyuki Hayashi^b ; ¹ ; ^{hayn@affrc.go.jp} ; Kazuo Ichimura^a ; ^{ichimu@affrc.go.jp} ; Masayoshi Nakayama^a ; ^{nakayosi@affrc.go.jp}
• 关键词：Anthocyanin ; Copigment ; Sweet pea ; Thin layer chromatography
• 刊名：Journal of Chromatography A
• 出版年：2012
• 期刊代码：47_00219673
• 类别：ch
• 出版时间：6 July, 2012
• 卷：1245
• 期：Complete
• 页码：183-189
• 文件大小：822 K

摘要

Anthocyanins are major flower pigments that can be affected by copigments, colorless compounds that can modify anthocyanin coloration to more intense and bluer. Thin-layer chromatography (TLC) is an available technique to separate and analyze anthocyanins and copigments. To easily and comprehensively detect copigments, we added function of mixture of compounds to TLC; by slantingly cross loading samples on TLC, compounds are symmetrically developed at various angle lines from the upper origin to individual R_f values and cross each other in an orderly fashion, where mixture is simultaneously performed with separation. Occurrence of copigments can be detected as a coloration change on the developed line of anthocyanin. Pink sweet pea (Lathyrus odoratus L.) petals were analyzed by the cross-TLC and a more intense spot and a paler spot on the anthocyanin line were detected. As each spot overlapped with an ultraviolet absorbance line, each of these ultraviolet absorption compounds was purified and identified as kaempferol 3-rhamnoside and 2-cyanoethyl-isoxazolin-5-one, respectively. Whereas kaempferol 3-rhamnoside is a flavonoid and had a general copigment effect of more intense and bluer coloration change, 2-cyanoethyl-isoxazolin-5-one is a compound whose structure is outside of conventional categories of copigments and had a novel effect to change anthocyanin coloration paler while maintaining color tone. We determined that the search for copigments should be carried out without pre-existing prediction of structures and effects. We have shown that slantingly cross loading samples system on plate-type chromatography is an effective technique for such comprehensive analysis of molecular interaction.