Author
Masatoshi Nakatsuji, Faculty of Pharmacy, Osaka Medical and Pharmaceutical University Takashi Inui, Graduate School of Agriculture, Osaka Metropolitan University
Keywords
circular dichroism, thermal stability, drug delivery system, lipocalin-type prostaglandin D synthase, poorly water-soluble drug
Abstract
Poor aqueous solubility of drug candidates limits the developability, motivating drug delivery system approaches for the formulation and targeted transport. Lipocalin-type prostaglandin D synthase (L-PGDS) is a β-barrel transport protein that binds diverse lipophilic ligands and is therefore an attractive carrier for poorly water-soluble drugs. To support quality control and formulation design, we established circular dichroism (CD) spectroscopy as a practical, high-throughput assay of L-PGDS thermal stability across pharmaceutically relevant conditions. Far-UV (200-260 nm) and near-UV (250-350 nm) CD spectra were recorded over pH 2.5-8.0, and thermal unfolding (20-85 ℃ ) was quantified with two-state fits and benchmarked by differential scanning calorimetry. Far-UV CD revealed a stable β-sheet signature with a negative extremum near 215 nm that was largely invariant with pH, while near-UV features around 290 nm indicated preserved tertiary packing. Thermal scans showed reversible unfolding in the mildly acidic to neutral range, with melting temperatures increasing with pH sand reaching ~72 ℃ at pH 7.4. At pH 7.4-8.0, partial irreversibility observed in near-UV CD was consistent with aggregation at higher protein concentrations and proximity to the isoelectric point. CD-derived Tm values agreed with calorimetry. These results demonstrate that L-PGDS possesses high thermal stability over a broad pH window and that circular dichroism provides a rapid, sensitive readout suitable for lot release and storage studies intended for drug delivery system applications.