Charge regulation and chemical transformation of heparin- mimicking polyampholytes Katarzyna Byś, Pablo M. Blanco, Peter Košovan, Mariusz Uchman Charles University, Czech Republic Highly sulfated heparin-mimicking polyelectrolytes effectively act as anticoagulants with tailorable structure and tunable sulfation degree. This versatility has prompted the scope of their biomedical applications. However, the complex charge regulation of the polymer chain and the mechanisms controlling the structure of such polymers remain unclear. Sodium poly((sulfamate-carboxylate)isoprene) is a natural rubber-derived high-charge-density polyelectrolyte containing two negatively charged groups of different pH sensitivity at the same monomeric unit. In acidic pH, the pH-dependent sulfamate group, which resembles the one found in heparin, undergoes N-S bond cleavage, which leads to the chemical transformation into a polyzwitterionic poly((amino-carboxylate) isoprene) [1-3]. The correct description of such polyampholyte systems is complex and requires a combination of experimental methods and computer simulations. To evaluate the pH-induced changes of ionization and mechanism of structural transformation we performed a post-polymerization modification of cis-1,4-polyisoprene to obtain poly((sulfamate-carboxylate)isoprene). The reaction was followed by acid-induced cleavage of over 90% of the N-S bonds and ultimately lead to chemical transformation into a polyzwitterionic poly((amino-carboxylate) isoprene. The structures of obtained polymers were confirmed by 1HNMR, 13CNMR, FTIR and elemental analysis. To evaluate the deviation from the ideal Henderson-Hasselbalch behaviour we complemented theoretical predictions with potentiometric titrations and zeta-potential measurements. This combination enabled us to find the effective pKa values of the ionizable groups, which in case of weak polyzwitterions is not possible directly from the equivalence point recognition criterion (ERC) in potentiometric titrations [4]. Additionally, since poly((sulfamate-carboxylate) isoprene) structurally resembles heparin, we conducted the antithrombotic behaviour study by activated partial thromboplastin time (aPTT) measurements on human plasma [5]. Furthermore, we elucidated the direct mechanism of anticoagulation by protein binding experiments using isothermal titration calorimetry. References 1. S. Pispas, J. Phys. Chem. B, 111, 8351 (2007) 2. M. Uchman, M. Štěpánek, K. Procházka, G. Mountrichas, S. Pispas, I. K. Voets, A. Walther, Macromolecules, 42(15), 5605- 5613 (2009) 3. A. Laschewsky, Polymers, 6(5), 1544-1601 (2014) 4. R. Lunkad, P. Biehl, A. Murmiliuk, P. M. Blanco, P. Mons, M. Štěpánek, F. H. Schacher, P. Kosovan, Macromolecules, 55 (17), 7775-7784 (2022) 5. B. Kalaska, K. Kamiński, J. Miklosz, K. Nakai, S.I. Yusa, D. Pawlak, M. Nowakowska, A. Mogielnicki, K. Szczubiałka, Biomacromolecules, 19(7), 3104–3118 (2018)
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