Low Adduct Peptide LC-MS Obtained with IonHance DFA and Certified LDPE Containers

Importance of the Topic

Minimizing metal ion contamination in peptide LC-MS analyses is critical to ensure accurate mass spectra and reliable data interpretation.
Sodium and potassium adducts can distort signal intensities and complicate spectral interpretation, particularly for sensitive assays in biotherapeutic development.

Objectives and Study Overview

This study aimed to reduce metal adduct formation in electrospray ionization mass spectra of peptides by combining a high-purity mobile phase additive, IonHance DFA, with certified low density polyethylene containers.
Peptide mapping separations were performed using the Waters mAb Tryptic Digestion Standard derived from NIST Reference Material 8671.

Methodology and Instrumentation

The mobile phase consisted of 0.1% IonHance DFA prepared in 18.2 MΩ·cm laboratory purified water.
Containers: Waters Certified LDPE vials (p/n 186009110) rigorously tested to contain less than 100 ppb sodium and potassium.
Chromatography: ACQUITY UPLC Peptide CSH C18 column (130 Å, 1.7 µm, 2.1 x 150 mm).
Mass spectrometry: Electrospray ionization source configured for peptide analysis.

Main Results and Discussion

Using high purity water and certified LDPE containers yielded clean UV and total ion current chromatograms with minimal adduct signals.
Extracted mass spectra of the T13 peptide (VQWK) from the light chain showed a dominant protonated ion with negligible sodium and potassium adducts.
In contrast, mobile phases prepared with external LC-MS water and noncertified vials exhibited significant sodiated and potassiated species, complicating spectral interpretation and skewing relative ion abundances.

Benefits and Practical Applications

The combination of IonHance DFA and certified LDPE containers provides a straightforward approach to control metal contamination in peptide LC-MS workflows.
Improved spectral clarity enhances confidence in peptide identification, quantitation, and comparability across assays.
This strategy is readily implementable in research, QA/QC, and biopharmaceutical development laboratories.

Future Trends and Applications

Advancements in container materials with even lower trace metal levels may further reduce adduct formation.
Integration of automated mobile phase preparation systems can standardize reagent handling and minimize human-introduced contamination.
Application of this approach to other biomolecule classes, such as oligonucleotides or intact proteins, and to high-resolution MS techniques can expand its utility.

Conclusion

Combining high-purity additives and certified low-metal containers effectively minimizes sodium and potassium adducts in peptide LC-MS, resulting in cleaner spectra and more reliable data.
This simple, reproducible method supports high-sensitivity peptide and protein analyses in diverse bioanalytical settings.

References

Nguyen J, Liu X, Lauber M (2019) Low Adduct Peptide LC-MS Obtained with IonHance DFA and Certified LDPE Containers. Waters Corporation.
Waters Corporation (2019) IonHance DFA product information and ACQUITY UPLC application notes.