Creative Proteomics provides peptide purity analysis services based on RP-HPLC, and we have RP-HPLC columns that can meet your various experimental needs. Meanwhile, our experienced team customizes experimental programs for customers. By analyzing retention time, peak shape, peak purity, and quantifying impurities, RP-HPLC enables us to assess the quality and integrity of peptide samples, ensuring the reliability of downstream applications in various fields such as pharmaceutical research, peptidomics, and peptide-based therapeutics.
How Do We Identify Peptide Purity by RP-HPLC?
Due to the different polarities of different polypeptides, their elution times on RP-HPLC will also be different, so that the separation of the polypeptides can be achieved. The separated peptides are analyzed through various techniques, such as UV detection, diode array detection, or mass spectrometry, which can be used to monitor the absorbance or mass spectra of the eluted peptide. A pure peptide will exhibit a single, sharp peak in the chromatogram, while impurities or contaminants will appear as additional peaks or shoulder peaks.
Key Factors in RP-HPLC-based Peptide Purity Identification
Detection Wavelength Selection
The peptide bond in the peptide is the amide bond, which usually has strong absorption at 220 nm (similar to protein), so we generally choose 220 nm as the detection wavelength of the peptide. If your peptide contains a benzene ring structure such as Trp, Tyr and dye molecules modifications, we can also use 275 nm as the detection wavelength for detection as a reference.
Column Selection
Chromatographic column is the most critical component in high performance liquid chromatography, and its plate number directly determines the resolution of impurity molecules in the instrument. The parameters of chromatographic columns commonly used in HPLC analysis and detection of peptides by us are: C18 (reverse-phase material), 4.6 mm (column inner diameter) × 250 mm (column length), 5 μm (silica particle size), 100 Å (silica pore size). The column efficiency of this type of column is at least 10,000.
Elution Solvent Selection
We will choose the most suitable elution solvent for you according to your sample characteristics, sample type and the solvent you use to dissolve the sample.
Elution Gradient
We often use gradient elution in peptide purity analysis, that is, the proportion of organic phase (mainly acetonitrile) in the eluent gradually increases with time. Because the rate of addition of organic phase has a significant impact on the purity result. We usually use a gradient elution method every minute (the concentration of acetonitrile increases by 1% every minute, v/v), which can avoid the overlap of sample peaks and impurity peaks and improve the accuracy of analysis results.
Peptide Purity Analysis
According to the number and area of the UV absorption spectrum detected by the instrument, we will calculate the purity of the peptide in the sample for you, and provide you with the corresponding experimental method report and final conclusion.
Advantages of Our Purity Identification Services Based on RP-HPLC
Better Versatility
In RP-HPLC experiments, we use water or water-based solvents as solvents, which can simplify problems caused by different hydrophobic solvents and improve the versatility of analysis.
More Accurate Gradient Separation
Because a universal water-based solvent is used, gradient separations can be more accurately controlled, resulting in improved analytical accuracy.
Cut Costs
Creative Proteomics uses RP-HPLC, which eliminates the need for solvents that are more organic than silica, thus reducing costs.
High Flexibility
Since a hydrophobic stationary phase is used, it can be used in combination with hydrophobic, hydrophilic, ionic and ionic compounds to separate their various components, allowing analysis to be flexible.
The RP-HPLC-based peptide purity analysis service provided by Creative Proteomics is more versatile, accurate and cost-effective. If you have needs for peptide purity analysis services and other related services, please contact us for consultation, and we will serve you with the greatest enthusiasm.
* For research use only. Not for use in diagnostic procedures!
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