Purpose

Forced degradation is critical to probe the stabilities and chemical reactivities of therapeutic peptides. Typically performed in bulk followed by LC-UV or LC-MS analysis, this traditional workflow consists of a reaction/analysis sequence and usually requires half a day to several days to form and measure the desired amounts of degradants. A faster method is needed to study peptide degradation in a shorter time in order to speed up the drug development process.

Methods

In the new rapid method developed in this study, peptide degradation occurs in levitated aqueous microdroplets using the Leidenfrost effect.

Results

This two-minute reaction/analysis workflow allows major degradation pathways of Buserelin, Octreotide, Desmopressin and Leuprorelin to be studied. The reactions include deamidation, disulfide bond cleavage, ether cleavage, peptide bond hydrolysis, and oxidation.

Conclusions

The accelerated forced degradation method requires a minimal amount of therapeutic peptide per stress condition, and the appropriate extent of degradation can be readily generated in seconds by adjusting the droplet levitation time. Levitated microdroplets should be applicable in pharmaceutical development to rapidly determine the intrinsic stability of therapeutic peptides and to aid formulation development by screening the effects of excipients on the stability of the peptides.

Graphical abstract

中文翻译：

---

悬浮微滴中治疗性肽的加速强迫降解。

目的

强制降解对于探测治疗性肽的稳定性和化学反应性至关重要。传统的工作流程通常是批量进行，然后进行LC-UV或LC-MS分析，它由反应/分析序列组成，通常需要半天到几天才能形成并测量所需的降解量。需要更快的方法来在更短的时间内研究肽的降解，以加快药物开发过程。

方法

在这项研究中开发的新的快速方法中，利用莱顿弗罗斯特效应在悬浮的水微滴中发生了肽降解。

结果

这一两分钟的反应/分析工作流程允许研究Buserelin，Octreotide，Desmopressin和Leuprorelin的主要降解途径。反应包括脱酰胺，二硫键裂解，醚裂解，肽键水解和氧化。

结论

加速的强制降解方法在每个应激条件下需要的治疗肽量最少，并且可以通过调节液滴悬浮时间在几秒钟内轻松产生适当程度的降解。悬浮微滴应适用于药物开发，以通过确定赋形剂对肽稳定性的影响来快速确定治疗性肽的内在稳定性并帮助制剂开发。

图形概要