The discovery of bioactive natural products remains a time-consuming and challenging task. The ability to link high-confidence metabolite annotations in crude extracts with activity would be highly beneficial to the drug discovery process. To address this challenge, HPLC-based activity profiling and advanced UHPLC-HRMS/MS metabolite profiling for annotation were combined to leverage the information obtained from both approaches on a crude extract scaled down to the submilligram level. This strategy was applied to a subset of an extract library screening aiming to identify natural products inhibiting oncogenic signaling in melanoma. Advanced annotation and data organization enabled the identification of compounds that were likely responsible for the activity in the extracts. These compounds belonged to two different natural product scaffolds, namely, brevipolides from a Hyptis brevipes extract and methoxylated flavonoids identified in three different extracts of Hyptis and Artemisia spp. Targeted isolation of these prioritized compounds led to five brevipolides and seven methoxylated flavonoids. Brevipolide A (1) and 6-methoxytricin (9) were the most potent compounds from each chemical class and displayed AKT activity inhibition with an IC₅₀ of 17.6 ± 1.6 and 4.9 ± 0.2 μM, respectively.

中文翻译：

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将活性分析与高级注释相结合，以加速发现靶向黑色素瘤致癌信号的天然产物

发现具有生物活性的天然产物仍然是一项耗时且具有挑战性的任务。将粗提物中的高可信度代谢物注释与活性联系起来的能力将对药物发现过程非常有益。为了应对这一挑战，将基于 HPLC 的活性分析和用于注释的高级 UHPLC-HRMS/MS 代谢物分析相结合，以利用从两种方法获得的信息，将粗提物缩小到亚毫克水平。该策略应用于提取文库筛选的子集，旨在鉴定抑制黑色素瘤中致癌信号传导的天然产物。先进的注释和数据组织能够识别可能导致提取物中活性的化合物。这些化合物属于两种不同的天然产物支架，在Hyptis和Artemisia spp的三种不同提取物中鉴定出Hyptis brevipes提取物和甲氧基化黄酮类化合物。这些优先化合物的靶向分离产生了五种短缩酚和七种甲氧基化黄酮类化合物。Brevipolide A ( 1 ) 和 6-methoxytricin ( 9 ) 是每个化学类别中最有效的化合物，显示出 AKT 活性抑制作用，IC ₅₀分别为 17.6 ± 1.6 和 4.9 ± 0.2 μM。