CEEPC/IPM/CMSC - Abstrakt prezentace

(CEEPC/IPM/CMSC 2022 - ThP-15)
Analysis of immunogenic peptide in vaccine formulation by a CE-MS approach

Juraj Piešťanský 1,2 *, Ondrej Štefánik 3, Andrea Horniaková 1, Ivana Čižmárová 1, Michaela Matušková 1, Peter Mikuš 1,2, Petra Majerová 4, Andrej Kováč 4

  1. Katedra farmaceutickej analýzy a nukleárnej farmácie, Farmaceutická fakulta, Univerzita Komenského
  2. Toxikologické a antidopingové centrum, Farmaceutická fakulta, Univerzita Komenského
  3. Katedra farmaceutickej analýzy a nukleárnej farmácie, Farmaceutická fakulta UK v Bratislave
  4. Neuroimunologický ústav SAV, Dúbravská cesta 9, 845 10 Bratislava

Abstrakt

With the growing interest in novel biologicals as peptide therapeutics and therapeutic peptide conjugates, the development of analytical methods for the quantitative analysis of peptides in pharmaceutical and/or biological matrices has become very important. Compared to small molecule drugs, peptide and protein biopharmaceuticals are large and heterogeneous (as a result of the biosynthetic process and subsequent manufacturing and storage), making their analysis very challenging. Analysis of large peptides and peptide conjugates is associated with the necessity to digest such large molecules. This procedure is most frequently performed by controlled enzymatic processes. Acid hydrolysis is one example of a complementary digestion technique, which was successfully tested and applied for “bottom-up” proteomic experiments.
Here, a robust analytical method based on on-line combination of capillary electrophoresis with tandem mass spectrometry (CE-MS/MS) was developed for determination of potentially immunogenic synthetic peptide in a conjugate with bovine serum albumin as carrier protein, and in a peptide conjugate formulated with an vaccine adjuvant – Alhydrogel® 2%. An effective non-enzymatic release step of the peptide from the final peptide conjugate based on acid hydrolysis with the use of 2% formic acid was successfully tested and implemented. Separation was performed in background electrolyte composed of 1 M formic acid (pH 1.88). Moreover, in-capillary preconcentration strategy based on dynamic pH junction was investigated. Such approach was able to improve the signal intensity. The presented approach represent an effective tool for simple, rapid and robust quantification of immunogens in modern immunotherapeutics.

* Korespondující autor: piestansky@fpharm.uniba.sk

Poděkování:

This work was supported by the projects VEGA 1/0514/22, KEGA 027UK-4/2020, APVV-15-0585, FaF/2/2022, and carried out in the Toxicological and Antidoping Center at the Faculty of Pharmacy Comenius University.


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