The biopolyester cutin is ubiquitous in land plants, building the polymeric matrix of the plant’s outermost defensive barrier, the cuticle. Cutin influences many biological processes in planta; however, due to its complexity and highly branched nature, the native structure remains partially unresolved. Our aim was to define an original workflow for the purification and systematic characterization of the molecular structure of cutin. To purify cutin we tested the ionic liquids cholinium hexanoate and 1-butyl-3-methyl-imidazolium acetate. The ensuing polymeric materials are highly esterified, amorphous, and have a typical monomeric composition as demonstrated by solid-state NMR, complemented by spectroscopic, thermal, and x-ray scattering analyses. We performed a systematic study by solution-state NMR of cryogenically milled cutins extracted from tomatoes (Solanum lycopersicum ‘Micro-Tom’; the wild type and the GLYCEROL-3-PHOSPHATE ACYLTRANSFERASE [GPAT6] and CUTIN SYNTHASE [CUS1] mutants). We resolved their molecular structures, relative distribution of ester aliphatics, free acid end-groups and free hydroxyl groups, differentiating between those derived from primary and secondary esters. Our data demonstrate the existence of free hydroxyl groups in cutin and provide insight into how the mutations affect the esterification arrangement of cutin. The usage of ionic liquids for studying plant polyesters has advantages over conventional approaches, since simple modifications can be applied to recover a biopolymer carrying distinct types/degrees of modifications (e.g. preservation of esters or cuticular polysaccharides), which in combination with the solution NMR methodologies developed here, constitutes essential tools to fingerprint the multifunctionality and the structure of cutin in planta.

生物聚酯角质在陆地植物中普遍存在，构建植物最外层防御屏障（角质层）的聚合物基质。角质影响植物体内的许多生物过程；然而，由于其复杂性和高度分支的性质，其天然结构仍然部分未得到解决。我们的目标是定义一个原始的工作流程，用于角质分子结构的纯化和系统表征。为了纯化角质，我们测试了离子液体己酸胆碱和乙酸 1-丁基-3-甲基-咪唑鎓。由此产生的聚合物材料是高度酯化的、无定形的，并具有典型的单体组成，如固态 NMR 所示，并辅以光谱、热和 X 射线散射分析。我们通过溶液态核磁共振对从番茄中提取的低温研磨角质（番茄‘ Micro-Tom’；野生型和甘油-3-磷酸酰基转移酶[ GPAT6 ] 和角质合成酶[ CUS1 ] 突变体）进行了系统研究。我们解析了它们的分子结构、酯脂肪族化合物、游离酸端基和游离羟基的相对分布，区分了源自伯酯和仲酯的那些。我们的数据证明角质中存在游离羟基，并深入了解突变如何影响角质的酯化排列。使用离子液体研究植物聚酯比传统方法具有优势，因为可以应用简单的修饰来回收带有不同类型/程度修饰的生物聚合物（例如酯或角质多糖的保存），这与溶液核磁共振方法相结合在这里开发的，构成了对植物角质的多功能性和结构进行指纹识别的重要工具。