(W1230-04-22) Assessment of High Throughput Salt Screening Methodologies with Zwitterionic Compounds

Purpose: Salt formation and characterization is an important and critical step in the developmental process of new chemical entity development, whether it be for information protection, crystallization development, purification, improvements in solubility, or chiral resolution. Generic screening methodologies with multiple counter ions is generally employed in pharmaceutical development for the rapid formation, isolation, and characterization of potentially viable salt forms. While salt screening is typically a straight-forward process, based on the pKa values of the active pharmaceutical ingredient and potential counterion, zwitterionic moieties have been observed to add an additional level of complexity where the propensity for self-interaction must be overcome to form salts. The main goal of the research was to investigate a handful of model zwitterionic compounds with known pKa values and isoelectric points and to improve the success of rapid screening approaches for successful salt formation.

Methods: Screening experiments were first conducted with three well studied amino acids and a commercially available active pharmaceutical ingredient as model compounds, L-glycine, L-valine, L-arginine, and Ciprofloxacin. Experiments were set up on the 30-50 mg scale for the salt screening process in 2 mL vials containing 6.3 mm stir bars. Eight counter ions (1.1, 2.2, and 3.3 equivalents, pKa values ranging from -6.00 to 4.54) were screened in up to six solvents with each amino acid and thirteen counter ions (up to 3.3 equivalents, pKa values ranging from -6.00 to 5.44) in three solvents were screened with Ciprofloxacin to probe for salt formation. The apparent pH of the water/EtOH solvent systems was taken to compare with speciation diagrams of the zwitterions. X-ray powder diffraction (XRPD) analysis was done in two stages. First, XRPD analysis of the wet cake was completed for all samples where solids were observed. Unique solids were then left on XRPD plates and dried under vacuum at 50 °C (~-29 inHg) overnight before they were reanalyzed by XRPD. All XRPD patterns were compared to counter ion XRPD patterns and known freeform patterns. Unique patterns were then assessed by thermogravimetric analysis coupled with differential scanning calorimetry and proton NMR to determine the nature of the solids.

Results: Evidence of successful salt formation was observed with L-glycine with HCl, sulfuric acid, toluene sulfonic acid, and benzene sulfonic acid. Result with phosphoric acid where inconclusive as only solutions were obtained from all 6 solvent systems. Cocrystals may have been formed with L-tartaric acid, citric acid, and 4-hydroxybenzoic acid. Similar results were noted with L-alanine and L-valine were successful salt formation using traditional screening approaches halted when the pKa of the counter ion was greater than 1.96. With L-valine, no evidence of cocrystal formation were observed, but cocrystals may have formed with L-alanine and L-tartaric acid and citric acid. Additional equivalents of counter ion (up to 3.3 equivalents) did not result in an increased propensity for salt formation with any of the amino acids assessed. Screening of Ciprofloxacin produced unique crystalline patterns with all thirteen of the counter ions assessed and successful salt formation was observed up to a counterion pKa of 1.96.

Conclusion: In conventional pharmaceutical development workflows, the difference in the pKa values between the acidic/basic pKa of the freeform molecule and the acidic/basic pKa of the counterion is used for selection of appropriate screening molecules. However, while it’s typical to look for a difference of 2-3 pKa units between the target acidic and basic sites of the freeform and counterion, respectively, this ‘rule’ has been shown to be inadequate for successful salt formation with zwitterions. Additional equivalents of counterion added to push the equilibrium towards salt formation with weaker acids was also found to be unsuccessful and incorporating additional experiments with that particular methodology may show little value in time sensitive screenin. It is instead suggested that a more careful consideration of the isoelectric point and the speciation diagram would lead to more successful experimental results in typical screening workflows.