PhD Scientific Days 2023

Budapest, 22-23 June 2023

Molecular Sciences - Posters B

Stability and Recovery Studies of Heparan Sulphate Disaccharides

Domonkos Pál1,2, Lilla Turiák2
1Semmelweis University, Doctoral School of Pharmaceutical Sciences, Budapest, Hungary
2MS Proteomics Research Group, Research Centre for Natural Sciences, Budapest, Hungary

Text of the abstract

Introduction: Heparan sulphate (HS) is a dominant class of glycosaminoglycans (GAGs), due to its significant biological functions, therefore the study of HS is an important task. The most effective way to investigate the structure of GAGs is to characterise the disaccharide building units of the polysaccharide chains. The sample preparation steps have a significant impact on the analysis of HS samples. Literature is only available regarding the stability and recovery of another class of GAGs, chondroitin sulfate.
Aims: Since HS disaccharides contain several sulfate groups, these samples may be sensitive to the effects of the sample preparation processes. To investigate these effects, we planned to perform stability and recovery studies on HS disaccharides.
Methods: The disaccharide sample analysis was performed by nanoHPLC system coupled to mass spectrometer via electrospray source. The separation was achieved on a 250 μm × 10.5 cm capillary column packed with GlycanPac AXH-1 resin using ammonium formate gradient. Eluent A was 10 mM ammonium formate in 75:25 v/v ACN:water (pH 4.4), while eluent B was 65 mM ammonium formate in 75:25 v/v ACN:water (pH 4.4).
We investigated the effect of storage vessels, as well as the different reconstituion processes on the recovery of HS samples. For stability testing, samples were stored for different periods under various conditions, such as pH, temperature, solvents, and refreezing.
Results: The recovery studies have shown that the samples are affected by the storage vessel and the reconstitution after drying. In the case of stability studies, it was found that HS samples are sensitive to storage conditions, especially regarding storage time, temperature, and the solvent composition.
Conclusion: HS GAGs showed a significant sensitivity to the storage and the sample preparation processes.
Funding: Supported by the ÚNKP-22-3-I New National Excellence Program of the Ministry for Culture and Innovation from the source of the National Research, Development and Innovation Fund.