Peptide Purity by HPLC is a key technique in pharmaceutical development, biotechnology, and academic research. At ResolveMass Laboratories Inc., we specialize in this testing process to make sure that peptides meet the highest purity standards for functionality, safety, and reproducibility. Our team has built a well-known reputation in Canada and globally by delivering trustworthy, accurate, and validated HPLC purity assessments for peptides of all types and complexities.
Explore our peptide characterization services to learn more about how we support scientists and developers at every stage of peptide analysis.
What Is Peptide Purity by HPLC?
Understanding Peptide Purity by HPLC Analysis
Peptide purity by HPLC is the process of finding out the percentage of the desired peptide in a given sample using High-Performance Liquid Chromatography (HPLC). This testing method separates parts based on how they interact with the column material and the flowing liquid, giving a clear breakdown of the main peptide peak and related unwanted substances.
This allows researchers to measure how much of the sample is made up of the desired peptide, versus shortened sequences, deletions, oxidized forms, or side-products.
The Importance of Peptide Purity by HPLC in Life Sciences
- Biological Performance: Even small impurities can alter peptide bioactivity, affecting therapeutic function or lab test results.
- Toxicological Safety: Regulatory bodies require detailed impurity profiles to reduce immunogenic risks.
- Batch-to-Batch Consistency: Quality control is important for manufacturing, formulation, and clinical studies.
- Intellectual Property and Patent Filing: Purity data strengthens patent submissions and new peptide ideas.
For comprehensive evaluations beyond HPLC, we recommend exploring our peptide characterization techniques.
Why Choose ResolveMass for Peptide Purity by HPLC?
Decades of Analytical Experience
Our scientists bring decades of experience in checking peptide purity by HPLC and working with peptide chemistry. We’ve tested over 10,000 peptide samples, ranging from cyclic peptides to long-chain therapeutic sequences. Whether you’re in early research or preparing for IND (Investigational New Drug) submission, we understand what’s needed for strong analytical validation.
Advanced Analytical Infrastructure
We use the latest HPLC and UHPLC instruments equipped with UV, PDA, and MS detectors. Our typical peptide purity by HPLC workflow includes:
- Sample solubilization and filtration
- Column optimization (C18, C8)
- Gradient methods tailored per peptide length and hydrophobicity
- Multi-wavelength detection for peptide bond and side-chain UV absorbance
- Integration of impurity peaks and main compound to calculate exact purity %
We ensure full traceability of results—peptide impurities characterization is included for uncertain or unknown peaks.
Regulatory-Ready Reporting
Clients receive comprehensive reports with:
- Percent purity
- Retention times of main and impurity peaks
- Raw chromatograms
- Peak purity confirmation by MS (when requested)
- Recommendations for resynthesis or purification steps
Learn more about our peptide purity testing process.
Peptide Purity by HPLC: Step-by-Step Breakdown
1. Sample Preparation
- Solubilize peptide in appropriate solvent (usually 0.1% TFA in water or acetonitrile)
- Centrifuge and filter before injection
2. Column Selection and HPLC Gradient
- RP-HPLC with C18 column for typical peptides
- Gradient: 5%–60% acetonitrile over 20 minutes
- Temperature: 30–45°C depending on peptide stability
3. Detection and Quantification
- UV at 214 nm for peptide backbone
- 280 nm for aromatic residues
- Percent purity = Area of main peak ÷ Total peak area × 100
Case Study: Improving Peptide Purity from 88% to 98.5%
Client: A clinical-stage biotech firm developing a 22-mer therapeutic peptide targeting a GPCR receptor.
Initial Results:
- HPLC showed 88.1% main peak, 11.9% total impurities
- LC-MS identified a truncated variant (+1 amino acid) and oxidized methionine
Actions Taken:
- We recommended re-purification using a steeper gradient and reduced temperature
- Optimized column phase from C18 to Phenyl-hexyl for better resolution of aromatic regions
Final Results:
- Post-purification peptide purity by HPLC: 98.5%
- Report included validated method with three independent runs, improving regulatory confidence
Outcome:
- Allowed submission to Health Canada with no analytical data queries
- Saved ~$12,000 by avoiding full re-synthesis
For cost breakdown, visit cost of peptide analysis service.
How Peptide Purity by HPLC Enhances Research Integrity
Scientific Reproducibility
Repeatable results are only possible when your peptides are pure. Researchers rely on trusted HPLC data to make sure that experimental results aren’t affected by impurities.
Product Development & Clinical Trials
For IND/CTA submissions, peptide APIs must follow strict purity guidelines. We offer method validation and stability testing aligned with ICH Q2(R1) and Health Canada standards.
Explore our peptide characterization service in Canada.
Top 10 Frequently Asked Questions (FAQs)
1. What purity percentage is acceptable for peptides in research vs. clinical settings?
For early research, >90% may be acceptable. Clinical-grade and therapeutic peptides usually require ≥98% purity by HPLC.
2. What impurities are commonly found in peptide samples?
Common impurities include deletion sequences, oxidation (e.g., Met), D-amino acid isomers, and aggregation artifacts.
3. How does peptide length affect HPLC purity resolution?
Longer peptides (>30 amino acids) may release impurities at the same time, requiring optimized gradients and longer run times.
4. Can HPLC detect sequence errors?
HPLC can show retention time differences but not sequence errors directly. LC-MS is needed to confirm structure.
5. What’s the turnaround time for peptide purity testing?
Standard turnaround is 2–3 business days. Fast 24-hour testing is available for urgent projects.
6. Do you offer method development for new peptides?
Yes, our scientists customize gradient, column, and detection methods to optimize purity separation for each unique sequence.
7. Is peak area always the most accurate purity measure?
Yes, area under the curve shows the relative mass of each compound better than peak height, especially with broad peaks.
8. Do we need to send reference peptides?
Not always. But providing a reference standard helps us validate retention times and detect positional isomers.
9. What’s the difference between HPLC and LC-MS purity results?
HPLC showa total % purity; LC-MS can spot individual impurities by mass. Together, they offer complete insight.
10. Do you support peptide stability testing over time?
Yes, we offer long-term stability testing under ICH conditions with scheduled re-analysis using HPLC and LC-MS.
Want to Discuss a Project? Contact Us
For more details, quotes, or technical consultation on peptide purity by HPLC, reach out via:
Conclusion: Why Peptide Purity by HPLC Should Be Non-Negotiable
Whether you’re developing a therapeutic, studying peptide-receptor interactions, or validating a batch for regulatory filing, peptide purity by HPLC is an essential quality control step. At ResolveMass Laboratories Inc., our unmatched experience in peptide testing makes sure that your data is built on solid analytical ground. Trust our team to guide your research with precision, speed, and honesty.
Start your peptide analysis with us, and make every data point count.
References:
2. https://www.mtoz-biolabs.com/applications-of-hplc-in-peptide-purity-analysis.html
