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ABSTRACT: Protein solubility was measured using the crystalline precipitate of a recombinant therapeutic antibody, in monovalent salt solutions containing KF, KCl, and KSCN (up to ∼ 0.7 M) at different pH conditions. For all three anions, the antibody solubility demonstrated complex behavior, both monotonic and nonmonotonic, with dependence on pH and salt concentration. At pH 7.1, close to the isoelectric point (pI) of 7.2, a typical salting-in behavior was observed with the salting-in constants of 12.7, 8.0, and 2.8 M for KSCN, KCl, and KF, respectively, suggesting that the anions follow the order of SCN(-) > Cl(-) > F(-) for increasing antibody solubility. Nonmonotonic behavior, as described by an initial solubility decrease followed by a solubility increase with ionic strength, was observed at pH 5.3, far below its pI. The effectiveness of the anion for reducing the solubility followed the order of SCN(-) > Cl(-) > F(-) . After the solubility reached the minimum, the anion's effectiveness for raising the antibody solubility was in agreement with that at pH 7.1. The mechanisms for the above phenomena are discussed based upon specific binding of the anions to the antibody surface. The mechanistic view of anion binding and its charge neutralization effect at pH 5.3 was supported by the results from the effective charge and zeta-potential measurements.
Journal of Pharmaceutical Sciences 11/2011; 101(3):965-77. · 3.06 Impact Factor
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ABSTRACT: A comprehensive approach was applied to develop a chiral purification method for an analyte that was found to be unusually difficult to scale-up in supercritical fluid chromatography (SFC). This was performed by studying major factors such as the solubility of an analyte in SFC mobile phases, impurity profiles, and cycle time. For this case study, the solubility in SFC mobile phase was measured by a packed column technique, coupled with a novel trapping mechanism to enhance measurement precision in SFC conditions. The solubility studies in SFC mobile phases suggested a couple of possible SFC mobile phases, in which the analyte would potentially be most soluble. The SFC methods were developed to purify a sample containing 15% of an impurity, after considering impurity profiles and cycle times of several potential methods in addition to SFC mobile phase solubility. An equal volume mixture of acetonitrile and ethanol was chosen for the final purification method, since this mixture demonstrated the relatively high SFC solubility among all solvent combinations with enhanced resolution between the analyte and the impurity as well as the shortest run time. The solubility of the compound was also determined in various organic solvents using a high throughput solubility screening system to better understand relative change of solubility from neat solution to SFC mobile phases.
Journal of Pharmaceutical and Biomedical Analysis 05/2008; 46(5):831-8. · 2.97 Impact Factor
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Helming Tan,
Maggie Reed,
Kyung H. Gahm,
Tony King,
Mina Dilmeghani Seran,
Tracy Bostick,
Van Luu,
David Semin,
Janet Cheetham,
Rob Larsen,
Mike Martinelli,
Paul Reider
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ABSTRACT: Trituration and crystallization are essential techniques for purification of intermediates and drug substances in pharmaceutical process development. Traditionally, identifying the optimal solvent conditions for impurity rejection adopts an empirical approach that can be a time and material consuming process. On the basis of thermodynamic considerations for optimizing the purity and the yield of the crystal product, it is necessary to conduct screening experiments with the crude product in different solvents including mixtures of solvents. In this report, we describe the development and the implementation of a high-throughput screening workflow in a 96-well array format for identifying the optimal purification conditions. The impure samples were first triturated at room temperature and subsequently subjected to a thermal cycle in 96 unique solvents or solvent mixtures at a volume of 0.6 mL per well. The compound solubility and the impurity profiles of both supernatant and recovered solid were analyzed by HPLC. Using a mixture of acetylsalicylic acid containing salicylic acid as the impurity, we investigated the effects of material loading per sample and impurity level on product purity/solubility under both screening and scale-up conditions and evaluated the thermodynamic behavior of product−impurity−solvent interactions based on an isothermal ternary phase diagram. During a further case study, a binary solvent system was identified, and the synergistic effect of binary components was demonstrated for purification of an Amgen compound containing three impurities. This high-throughput screening approach is valuable as an integrated part of process development to identify the thermodynamically favored solvent conditions for purification of pharmaceutical compounds.
12/2007;
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ABSTRACT: A series of trisubstituted pyrimidines were synthesized to improve aqueous solubility of our first TRPV1 clinical candidate (1; AMG 517), while maintaining potent TRPV1 inhibitory activity. Structure-activity and structure-solubility studies led to the identification of compound 26. The aqueous solubility of 26 (>or=200microg/mL, 0.01 HCl; 6.7microg/mL, phosphate buffered saline (PBS); 150microg/mL, fasted-state simulated intestinal fluid (SIF)) was significantly improved over 1. In addition, compound 26 was found to be orally bioavailable (rat F(oral)=24%) and had potent TRPV1 antagonist activity (capsaicin IC(50)=1.5nM) comparable to that of 1.
Bioorganic & medicinal chemistry letters 12/2007; 17(23):6539-45. · 2.65 Impact Factor
