Strigolactones represent the most recently described group of plant hormones involved in many aspects of plant growth regulation. Simultaneously, root exuded strigolactones mediate rhizosphere signaling towards beneficial arbuscular mycorrhizal fungi, but also attract parasitic plants. The seed germination of parasitic plants induced by host strigolactones leads to serious agricultural problems worldwide. More insight in these signaling molecules is hampered by their extremely low concentrations in complex soil and plant tissue matrices, as well as their instability. So far, the combination of tailored isolation—that would replace current unspecific, time-consuming and labour-intensive processing of large samples—and a highly sensitive method for the simultaneous profiling of a broad spectrum of strigolactones has not been reported. Depending on the sample matrix, two different strategies for the rapid extraction of the seven structurally similar strigolactones and highly efficient single-step pre-concentration on polymeric RP SPE sorbent were developed and validated. Compared to conventional methods, controlled temperature during the extraction and the addition of an organic modifier (acetonitrile, acetone) to the extraction solvent helped to tailor strigolactone isolation from low initial amounts of root tissue (150 mg fresh weight, FW) and root exudate (20 ml), which improved both strigolactone stability and sample purity. We have designed an efficient UHPLC separation with sensitive MS/MS detection for simultaneous analysis of seven natural strigolactones including their biosynthetic precursors—carlactone and carlactonoic acid. In combination with the optimized UHPLC–MS/MS method, attomolar detection limits were achieved. The new method allowed successful profiling of seven strigolactones in small exudate and root tissue samples of four different agriculturally important plant species—sorghum, rice, pea and tomato. The established method provides efficient strigolactone extraction with aqueous mixtures of less nucleophilic organic solvents from small root tissue and root exudate samples, in combination with rapid single-step pre-concentration. This method improves strigolactone stability and eliminates the co-extraction and signal of matrix-associated contaminants during the final UHPLC–MS/MS analysis with an electrospray interface, which dramatically increases the overall sensitivity of the analysis. We show that the method can be applied to a variety of plant species.

中文翻译：

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使用 UHPLC-MS/MS 分析复杂样品基质中独脚金内酯的改进策略。

独脚金内酯代表最近描述的一组植物激素，涉及植物生长调节的许多方面。同时，根部分泌的独脚金内酯介导根际信号向有益的丛枝菌根真菌发出信号，但也会吸引寄生植物。由寄主独脚金内酯诱导的寄生植物种子萌发导致世界范围内严重的农业问题。由于它们在复杂的土壤和植物组织基质中的极低浓度以及它们的不稳定性，阻碍了对这些信号分子的深入了解。到目前为止，尚未报道将定制分离（将取代当前对大样本的非特异性、耗时和劳动密集型处理）与同时分析广谱独脚金内酯的高灵敏度方法相结合。根据样品基质的不同，开发并验证了两种不同的策略，用于快速提取七种结构相似的独脚金内酯和在聚合物 RP SPE 吸附剂上进行高效的单步预浓缩。与传统方法相比，在提取过程中控制温度并在提取溶剂中添加有机改性剂（乙腈、丙酮）有助于从低初始量的根组织（150 mg 鲜重，FW）和根分泌物中分离独脚金内酯（ 20 ml），这提高了独脚金内酯的稳定性和样品纯度。我们设计了一种高效的 UHPLC 分离装置和灵敏的 MS/MS 检测器，用于同时分析七种天然独脚金内酯，包括它们的生物合成前体——carlactone 和 carlactoneic acid。结合优化的 UHPLC-MS/MS 方法，达到了阿摩尔检测限。新方法成功地分析了四种不同的重要农业植物物种——高粱、水稻、豌豆和番茄的小渗出液和根组织样本中的七种独脚金内酯。所建立的方法提供了有效的独脚金内酯萃取，其与来自小根组织和根分泌物样品的亲核性较低的有机溶剂的水性混合物相结合，并结合快速的单步预浓缩。该方法提高了独脚金内酯的稳定性，并在使用电喷雾接口的最终 UHPLC-MS/MS 分析期间消除了基质相关污染物的共萃取和信号，从而显着提高了分析的整体灵敏度。我们表明该方法可以应用于多种植物物种。