RP-HPLC analytical method development and optimization for quantification of donepezil hydrochloride in orally disintegrating tablet

An easy, fast and validated RV-HPLC method was invented to quantify donepezil hydrochloride in drug solution and orally disintegrating tablet. The separation was carried out using reversed phase C-18 column [Agilent Eclipse Plus C-18] with UV detection at 268 nm. Method optimization was tested using various composition of organic solvent. The mobile phase comprised of phosphate buffer [0.01M], methanol and acetonitrile [50:30:20, v/v] adjusted to pH 2.7 with phosphoric acid [80%] was found as the optimum mobile phase. The method showed intraday precision and accuracy in the range of 0.24% to -1.83% and -1.83% to 1.99% respectively, while interday precision and accuracy ranged between 1.41% to 1.81% and 0.11% to 1.90% respectively. The standard calibration curve was linear from 0.125 micro g/mL to 16 micro g/mL, with correlation coefficient of 0.9997 +/- 0.00016. The drug solution was stable under room temperature at least for 6 hours. System suitability studies were done. The average plate count was > 2000, tailing factor <1, and capacity factor of 3.30. The retention time was 5.6 min. The HPLC method was used to assay donepezil hydrochloride in tablet and dissolution study of in-house manufactured donepezil orally disintegrating tablet and original Aricept

Methylparaben concentration in commercial Brazilian local anesthetics solutions

OBJECTIVE: To detect the presence and concentration of methylparaben in cartridges of commercial Brazilian local anesthetics. MATERIAL AND METHODS: Twelve commercial brands (4 in glass and 8 in plastic cartridges) of local anesthetic solutions for use in dentistry were purchased from the Brazilian market and analyzed. Different lots of the commercial brands were obtained in different Brazilian cities (Piracicaba, Campinas and São Paulo). Separation was performed using high performance liquid chromatography (HPLC) with UV-Vis detector. The mobile phase used was acetonitrile:water (75:25 - v/v), pH 4.5, adjusted with acetic acid at a flow rate of 1.0 ml.min-1. RESULTS: When detected in the solutions, the methylparaben concentration ranged from 0.01% (m/v) to 0.16% (m/v). One glass and all plastic cartridges presented methylparaben. CONCLUSION: 1. Methylparaben concentration varied among solutions from different manufacturers, and it was not indicated in the drug package inserts; 2. Since the presence of methylparaben in dental anesthetics is not regulated by the Brazilian National Health Surveillance Agency (ANVISA) and this substance could cause allergic reactions, it is important to alert dentists about its possible presence.

Enhancement of catalytic activity of enzymes by heating in anhydrous organic solvents: 3D structure of a modified serine proteinase at high resolution.

For the first time, it is demonstrated that exposure of an enzyme to anhydrous organic solvents at optimized high temperature enhances its catalytic power through local changes at the binding region. Six enzymes, namely, proteinase K, wheat germ acid phosphatase, alpha-amylase, beta-glucosidase, chymotrypsin and trypsin were exposed to acetonitrile at 70 degrees C for three hr. The activities of these enzymes were found to be considerably enhanced. In order to understand the basis of this change in the activity of these enzymes, proteinase K was analyzed in detail using X-ray diffraction method. The overall structure of the enzyme was found to be similar to the native structure in aqueous environment. The hydrogen bonding system of the catalytic triad remained intact after the treatment. However, the water structure in the substrate binding site underwent some rearrangement as some of the water molecules were either displaced or completely absent. The most striking observation concerning the water structure was the complete deletion of the water molecule which occupied the position at the so-called oxyanion hole in the active site of the native enzyme. Three acetonitrile molecules were found in the present structure. All the acetonitrile molecules were located in the recognition site. Interlinked through water molecules, the sites occupied by acetonitrile molecules were independent of water molecules. The acetonitrile molecules are involved in extensive interactions with the protein atoms. The methyl group of one of the acetonitrile molecules (CCN1) interacts simultaneously with the hydrophobic side chains of Leu 96, Ile 107 and Leu 133. The development of such a hydrophobic environment at the recognition site introduced a striking conformation change in Ile 107 by rotating its side chain about C alpha-C beta bond by 180 degrees to bring about the delta-methyl group within the range of attractive van der Waals interactions with the methyl group of CCN1. A similar change had earlier been observed in proteinase K when it was complexed to a substrate analogue, lactoferrin fragment.