Accelerating GLP-1 Development with High- Throughput LC-MS Using the BioAccord™ LC MS System and the INTACT Mass Application

Significance of the topic

The analysis of GLP-1 analogs plays a critical role in therapeutic peptide development for type 2 diabetes and obesity. Rapid, reliable confirmation of molecular mass, purity assessment, and impurity profiling ensure consistent product quality, support process control, and satisfy regulatory requirements throughout research, scale-up, and manufacturing stages.

Objectives and Study Overview

This application note demonstrates an automated high-throughput LC-MS workflow using the Waters BioAccord LC-MS System integrated with the waters_connect Informatics Platform and INTACT Mass Application. Key aims include:

• Deploying a one-minute intact mass confirmation method for synthetic GLP-1 analogs across 48-well plates.
• Automating data processing for mass confirmation, deconvolution, impurity identification, and report generation.
• Delivering consistent, high-precision results with minimal user intervention.

Methodology and Used Instrumentation

The study employed a reversed-phase, one-minute gradient on an ACQUITY Premier Peptide C18 column (2.1×50 mm, 1.7 µm) at 60 °C. Key LC–MS settings included 0.8 mL/min flow rate, UV detection at 280 nm, and MSe acquisition (50–2000 m/z, 10 Hz scan rate). Scheduled lockmass correction minimized inter-sample checks. Deconvolution used the BayesSpray algorithm with monoisotopic outputs. Instrumentation details:

• Waters BioAccord LC-MS System with ACQUITY Premier UPLC (Binary) and tunable UV detector
• ACQUITY RDa Mass Detector
• waters_connect Informatics Platform with INTACT Mass Application v1.9 and Sample Submission App v2.7.0

Main Results and Discussion

Automated acquisition and processing times were under 60 seconds per sample, with potential to reach 30 seconds using trap/elute methods. Average mass error across 48 samples was –0.34 ppm (±1.43 ppm). The INTACT Mass Application allowed:

• Target definition by chemical formula or expected mass to refine deconvolution.
• Automated impurity profiling, reporting delta masses of novel species.
• Color-coded dashboards for pass (green), warning (orange), and fail (red) flags, enabling review by exception.

Example analyses:

• Liraglutide: Main species mass error of 0.1 ppm; lowest impurity (des-Phe, Glu) at 0.17 %; total MS×LC purity of 91.91 % including adducts.
• Semaglutide: Mass error of 1.2 ppm; identification of known impurities and an unknown +13 Da species flagged for follow-up.

Benefits and Practical Use

Key advantages of this high-throughput workflow include:

• End-to-end analysis in under one minute, enabling hundreds to thousands of samples per week.
• Automated data processing reduces human error and standardizes results.
• Rapid decision-making through graphical summaries and exception handling.
• PDF report generation for secure documentation and regulatory compliance.

Future Trends and Applications

Ongoing developments may include further acceleration to sub-30-second runs, expanded trap/elute capabilities, and broader adoption in contract and core laboratories. Integration of advanced machine-learning algorithms could refine impurity prediction and structural elucidation. The platform may also be extended to other therapeutic peptides, oligonucleotides, and biotherapeutic screening workflows.

Conclusion

The Waters BioAccord LC-MS System coupled with the waters_connect INTACT Mass Application offers a robust, automated solution for high-throughput GLP-1 analog analysis. By combining rapid one-minute acquisitions, intelligent deconvolution, impurity profiling, and review-by-exception dashboards, laboratories can achieve consistent, high-quality data and accelerate peptide development and quality control operations.

References

• Shion H., Berger S.J., Yu Y.Q. Application of a Mass Confirmation Workflow for Biotherapeutics Screening. Waters Corporation Application Note, 720007027; November 2020.
• Shion H., Boyce P., Berger S.J., Yu Y.Q. INTACT Mass™ – A Versatile waters_connect™ Application for Rapid Mass Confirmation and Purity Assessment of Biotherapeutics. Waters Corporation Application Note, 720007547; February 2022.
• Fox J., Berger S., Denbigh L., Yu K. 100 Samples per Hour: High Speed Oligonucleotide Analysis with BioAccord LC-MS and INTACT Mass™. Waters Corporation Application Note, 720009127; November 2025.