Bøgeskov Schmidt Frederik, Heskes Allison M, Thinagaran Dinaiz, Lindberg Møller Birger, Jørgensen Kirsten, Boughton Berin A
Plant Biochemistry Laboratory, Department of Plant and Environmental Sciences, University of Copenhagen, Copenhagen, Denmark.
VILLUM Research Center for Plant Plasticity, University of Copenhagen, Copenhagen, Denmark.
Front Plant Sci. 2018 Jun 28;9:892. doi: 10.3389/fpls.2018.00892. eCollection 2018.
Mass spectrometry based imaging is a powerful tool to investigate the spatial distribution of a broad range of metabolites across a variety of sample types. The recent developments in instrumentation and computing capabilities have increased the mass range, sensitivity and resolution and rendered sample preparation the limiting step for further improvements. Sample preparation involves sectioning and mounting followed by selection and application of matrix. In plant tissues, labile small molecules and specialized metabolites are subject to degradation upon mechanical disruption of plant tissues. In this study, the benefits of cryo-sectioning, stabilization of fragile tissues and optimal application of the matrix to improve the results from MALDI mass spectrometry imaging (MSI) is investigated with hydroxynitrile glucosides as the main experimental system. Denatured albumin proved an excellent agent for stabilizing fragile tissues such as leaves. In stem cross sections of , maintaining the samples frozen throughout the sectioning process and preparation of the samples by freeze drying enhanced the obtained signal intensity by twofold to fourfold. Deposition of the matrix by sublimation improved the spatial information obtained compared to spray. The imaging demonstrated that the cyanogenic glucosides (CNglcs) were localized in the vascular tissues in old stems of and in the periderm and vascular tissues of tubers. In MALDI mass spectrometry, the imaged compounds are solely identified by their ratio. MG20 and the mutant that is devoid of hydroxynitrile glucosides were used as negative controls to verify the assignment of the observed masses to linamarin, lotaustralin, and linamarin acid.
基于质谱的成像技术是一种强大的工具，可用于研究多种代谢物在各种样品类型中的空间分布。仪器和计算能力的最新发展扩大了质量范围、提高了灵敏度和分辨率，使样品制备成为进一步改进的限制步骤。样品制备包括切片、固定，然后选择和应用基质。在植物组织中，不稳定的小分子和特殊代谢物在植物组织受到机械破坏时容易降解。在本研究中，以羟基腈苷为主要实验体系，研究了冷冻切片、脆弱组织的稳定化以及基质的优化应用对改进基质辅助激光解吸电离质谱成像（MSI）结果的作用。变性白蛋白被证明是稳定脆弱组织（如叶片）的极佳试剂。在[植物名称]的茎横切片中，在整个切片过程中保持样品冷冻，并通过冷冻干燥制备样品，可使获得的信号强度提高两倍至四倍。与喷雾法相比，通过升华沉积基质改善了获得的空间信息。成像结果表明，生氰苷（CNglcs）定位于[植物名称]老茎的维管组织以及块茎的周皮和维管组织中。在基质辅助激光解吸电离质谱中，成像的化合物仅通过其质荷比来识别。MG20和不含羟基腈苷的突变体[植物名称]用作阴性对照，以验证观察到的质量对应于亚麻苦苷、百脉根苷和亚麻苦苷酸的归属。
