Temperature-responsive molecular recognition chromatography using phenylalanine and tryptophan derived polymer modified silica beads.

Temperature-responsive polymers incorporating molecular-recognition sites were developed as stationary phases for high-performance liquid chromatography (HPLC). The grafted stationary phases consisted of functional copolymers composed of N-isopropylacrylamide (NIPAAm) and N-acryloyl aromatic amino acid methyl esters, i.e., phenylalanine and tryptophan methyl esters (Phe-OMe and Trp-OMe). Three novel temperature-responsive polymers, P(NIPAAm-co-Phe-OMe5), P(NIPAAm-co-Phe-OMe10), and P(NIPAAm-co-Trp-OMe5), were synthesized. These copolymers exhibited a reversible hydrophilic/hydrophobic phase transition at their lower critical solution temperatures (LCSTs). The polymers were grafted onto aminopropyl silica using an activated ester-amine coupling method, and were packed into a stainless steel column, which was connected to an HPLC system. Temperature-responsive chromatography was conducted using water as the sole mobile phase. More hydrophobic analytes were retained longer, and the retention times of aromatic steroids and aromatic amino acids were dramatically increased. This indicated that π-π interactions occurred between the phenyl or indole moieties of phenylalanine or tryptophan, respectively, and the aromatic compounds. Furthermore, the retention times of compounds with hydrogen bond acceptors were higher with P(NIPAAm-co-Trp-OMe5), which contained indole as a hydrogen bond donor, than with P(NIPAAm-co-Phe-OMe5). This indicated that hydrogen bonding occurred between the stationary phase and the analytes. These results indicate that hydrophobic, π-π, and hydrogen bonding interactions all affected the separation mode of the temperature-responsive chromatography, and led to selective separation with molecular recognition. Both temperature-response and molecular recognition characteristics are present in the proposed separation system that utilizes a temperature-responsive polymer bearing aromatic amino acid derivatives.

Determination of triamcinolone, triamcinolone acetonide and fluocinonide in dosage forms.

The colorimetric method for determination of triamcinolone, triamcinolone acetonide and fluocinonide in tablets, ointments and cream using isonicotinic acid hydrazide is proposed. This method provides a precise and reproducible results (recovery ranged within 97.42-101.75%, variation coefficient ranged within 0.99-2.56%).

Separation and determination of lipophilic corticosteroids and benzothiazepin analogues by micellar electrokinetic chromatography using bile salts.

The separation of corticosteroids and benzothiazepin analogues by micellar electrokinetic chromatography (micellar EKC) was studied in comparison with capillary zone electrophoresis. The separation of these substances was not successful under neutral and alkaline conditions because they migrated with the same velocity as that of the electroosmotic flow. Micellar EKC with sodium dodecyl sulphate (SDS) solutions was also not successful because these substances migrated with almost the same velocity as that of the SDS micelle, owing to their high lipophilicity. The use of bile salts, which have a similar skeleton to corticosteroids, as the micellar phase permitted the separation of these substances with high theoretical plate numbers (150,000-350,000) within a short time (ca. 15 min). Sodium cholate was particularly useful. The effects of bile salt concentration, pH and the addition of methanol were investigated. Micellar EKC was also applied to the determination of the drug substances in tablets and cream using the internal standard method and to purity testing of drug substances and tablets.