Amine additives for improved in-ESI H/D exchange.

Area of Science:

• Analytical Chemistry
• Mass Spectrometry
• Biochemistry

Background:

• In-solution electrospray ionization hydrogen-deuterium exchange (in-ESI H/D exchange) is valuable for analyzing complex mixtures.
• Standard in-ESI H/D exchange often requires high temperatures, potentially causing artifactual exchanges and complicating data interpretation.
• Gas-phase H/D exchange efficiency is influenced by the deuterating agent's basicity, suggesting amine additives could improve in-ESI exchange.

Purpose of the Study:

• To evaluate the efficacy of various amines in enhancing in-ESI H/D exchange compared to using D2O alone.
• To identify an optimal amine modifier for increasing the extent of H/D exchange in small molecules, peptides, and proteins.
• To investigate the mechanism by which amines facilitate in-ESI H/D exchange.

Main Methods:

• In-solution electrospray ionization mass spectrometry (in-ESI-MS) was employed.
• Experiments compared deuterium incorporation using D2O alone versus D2O with different amine additives (primary, secondary, tertiary).
• Analysis focused on the extent of H/D exchange in small molecules, peptides, and proteins under various conditions, including ambient temperature.

Main Results:

• Amine additives substantially increased the extent of H/D exchange across small molecules, peptides, and proteins, even without capillary heating.
• The order of increasing exchange efficiency was observed as: tertiary amines < secondary amines < primary amines.
• Evidence suggests amines act as deuterating agents after initial exchange with D2O, facilitating broader H/D exchange.

Conclusions:

• Primary amines are the most effective additives for enhancing in-ESI H/D exchange.
• The use of amine additives offers a method to improve H/D exchange efficiency, particularly for small molecule analysis, without the need for elevated temperatures.
• Further research into amine-enhanced H/D exchange in peptides and proteins is warranted.