Determination and differentiation of surface and bound water in drug substances by near infrared spectroscopy.

Near infrared spectroscopy (NIRS) was utilized to determine the water content during the drying of a drug substance with Karl Fisher titration as the reference measurement. NIRS calibration models were built with Partial Least Squares (PLS) regression based on spectral region of 1822-1948 nm for samples with 1-40% (w/w) water from five batches. A standard error of prediction (SEP) of 1.85% (w/w) was obtained from validation of the model with additional batches. A second NIRS calibration model using PLS was constructed in the region of 1-10% (w/w) water with samples from the same five calibration batches. This calibration model improved the accuracy of the prediction in the critical region around the end point of drying and provided a standard error of prediction 0.42% (w/w). In addition, direct spectral analyses with Principal Component Analysis (PCA) and peak ratios were applied to distinguish between surface (unbound) water and bound water incorporated into the crystal lattice of the drug substance. Direct spectral analysis indicated the existence of significant numerical and graphical differences between samples containing both surface and bound water, and those containing bound water only. Applying this method to monitor an actual drying process with the graphical assistance of spectral analysis, the drying process can be controlled, and the end point of drying clearly determined to ensure the desired hydrate form of the product. This study has demonstrated the in-line monitoring capability of NIRS to differentiate the surface and bound water simultaneously in addition to the determination of the water level.

Unusual effect of column temperature on chromatographic enantioseparation of dihydropyrimidinone acid and methyl ester on amylose chiral stationary phase.

This paper reports an unusual effect of column temperature on the separation of the enantiomers of dihydropyrimidinone (DHP) acid and its methyl ester on a derivatized amylose stationary phase by normal-phase liquid chromatography. The separation of the DHP acid enantiomers was investigated using both carbamate-derivatized amylose and cellulose stationary phases (Chiralpak AD and Chiralcel OD) with an ethanol-n-hexane (EtOH-n-Hex) mobile phase. On the amylose phase, the van 't Hoff plot of the retention factor of the S-(+)-DHP acid was observed to be non-linear while that of R-(-)-DHP acid was linear. Likewise, the van 't Hoff plot for DHP acid enantioselectivity was non-linear with a transition occurring at approximately 30 degrees C. Furthermore, the van 't Hoff plot for the DHP acid enantioselectivity factor for data taken when heating the column from 5 to 50 degrees C was not superimposable with the same plot prepared with data from the cooling process from 50 to 5 degrees C. This observation suggested that the stationary phase was undergoing a thermally induced irreversible conformational change that altered the separation mechanism between the heating and cooling cycles. Similar phenomena were observed for the separation of the enantiomers of the DHP ester probe compound. The conformational change of the AD phase was shown to depend on the polar component of the mobile phase. When 2-propanol (2-PrOH) was used as the modifier instead of EtOH, the van 't Hoff plots for DHP acid were linear and thermally reversible, suggesting that no such irreversible conformational change occurs with this modifier. Conversely, when the AD phase was pre-conditioned with a more polar methanol (MeOH) or water containing mobile phase, thermal irreversibility of DHP acid enantioselectivity was once again observed. Interestingly, when the stationary phase was changed to its cellulose analogue, the Chiralcel OD, all van 't Hoff plots for the retention and selectivity of DHP acid were thermally reversible for both EtOH-n-Hex and 2-PrOH-n-Hex mobile phases.

Evaluation of the origins of the selectivity of polymers imprinted with a HIV protease inhibitor using infrared spectroscopy and high performance liquid chromatography.

This work investigates the origins of enantioselectivity of polymers imprinted with the HIV protease inhibitor, Indinavir. For the preparation of imprints of the drug, the critical interactions between the functional monomer, methacrylic acid, and Indinavir were characterized by infrared (IR) spectroscopy to explore the optimum functional monomer concentration for the polymerization. It was shown that a polymer with high selectivity and minimum non-selective binding for Indinavir was obtained when prepared with enough functional monomer to hydrogen bond with all of the functional groups of the drug without using an excess of monomer. This observation is explained in terms of a balance that is achieved in the monomer-template equilibrium during the polymerization that yields a polymer with highly selective sites and minimal non-selective sites. This paper further demonstrates that IR spectroscopy can be a valuable tool in the design and syntheses of molecular imprinted polymers.