Vojko Kmetec

Stability of ascorbyl palmitate in topical microemulsions

Ascorbyl palmitate and sodium ascorbyl phosphate are derivatives of ascorbic acid, which differ in stability and hydro-lipophilic properties. They are widely used in cosmetic and pharmaceutical preparations. In the present work the stability of both derivatives was studied in microemulsions for topical use as carrier systems. The microemulsions were of both o/w and w/o types and composed of the same ingredients. The stability of the less stable derivative ascorbyl palmitate was tested under different conditions to evaluate the influence of initial concentration, location in microemulsion, dissolved oxygen and storage conditions. High concentrations of ascorbyl palmitate reduced the extent of its degradation. The location of ascorbyl palmitate in the microemulsion and oxygen dissolved in the system together significantly influence the stability of the compound. Light accelerated the degradation of ascorbyl palmitate. In contrast, sodium ascorbyl phosphate was stable in both types of microemulsions. Sodium ascorbyl phosphate is shown to be convenient as an active ingredient in topical preparations. In the case of ascorbyl palmitate, long-term stability in selected microemulsions was not adequate. To formulate an optimal carrier system for this ingredient other factors influencing the stability have to be considered.

Liquid chromatographic determination of diclofenac in human synovial fluid.

A simple, rapid and sensitive HPLC method for the determination of diclofenac in synovial fluid is described. Special attention was paid to the procedure of sample preparation since gel formation may sometimes occur in synovial samples. With a one-step extraction procedure good and reproducible recovery of diclofenac was obtained. A subsequent HPLC assay was adjusted so as to achieve adequate sensitivity and precision needed for analysis of true samples. The results obtained by the described procedure proved the method to be suitable for monitoring concentrations of diclofenac in synovial fluid.

HPLC determination of tramadol in human breast milk

Tramadol is a centrally acting analgesic used for prevention and treatment of moderate to severe pain. It is estimated that 0.1% of the administered dose passes into breast milk causing potentially unwanted effects in nursing babies. Pharmacokinetically, breast milk is supposed to be a separate compartment into which the drug is excreted-mainly by passive diffusion. Due to a complex composition of breast milk, a suitable sample preparation procedure is needed with a subsequent chromatographic analysis for drug determination. Among several sample cleanup procedures tested we chose the liquid-liquid extraction procedure using n-hexane as an organic phase with back extraction into aqueous phase since it was considered the most suitable and the most compatible with the subsequent HPLC analysis. The precision and the reproducibility of the method were improved approximately two times by using metoprolol as an internal standard thus making the method also more robust with regard to a variable composition of milk samples. These characteristics, together with low detection limit and short analysis time, proved that the developed method is suitable for monitoring of tramadol in human breast milk.

Optimization of HPLC method for stability testing of bacitracin

A stability indicating HPLC assay for bacitracin has been developed and validated. The assay is based on a gradient elution, reversed phase column and UV diode array detection. On the basis of our previous analytical work several additional systematic HPLC tests for optimization of analytical method were performed. In order to achieve the highest selectivity of HPLC method, tests were conducted with extremely complex samples -- zinc bacitracin feed grade as food additive for animals. The influence of pH of mobile phase and type of columns on chromatographic separation of active (A, B(1) and B(2)) and inactive (F) polypeptide components of bacitracin were investigated in detail. It was found also that the peak B(1) comprises three and the peak F two subunits -- probably isomers. The obtained analytical procedure proved to be very selective and effective for the simultaneous determination of active polypeptide A, B(1) and B(2), impurities, known and unknown degradation products and ballast material.

The use of microcalorimetry and HPLC for the determination of degradation kinetics and thermodynamic parameters of Perindopril Erbumine in aqueous solutions

Perindopril Erbumine (PER) is one of the newly used angiotensin-converting enzyme inhibitors (ACE inhibitors) and is used for the treatment of patients with hypertension and symptomatic heart failure. It has two main degradation pathways, i.e. the degradation by hydrolysis and the degradation by cyclization. An isothermal heat conduction microcalorimetry (MC) and high pressure liquid chromatography (HPLC) were used for the characterization of aqueous solutions of PER and its stability properties. The rates of heat evolved during degradation of perindopril were measured by MC as a function of temperature and pH and from these data rate constant and change in enthalpy of the reactions were determined. With the HPLC method the concentration of perindopril and its degradation products were measured as a function of time in aqueous solutions of different pH that were stored at different temperatures. We demonstrated that reactions of degradation of perindopril at observed conditions follow the first order kinetics. The Arrhenius equation for each pH was determined. At pH 6.8 only one degradation pathway is present, i.e. the degradation by hydrolysis. Degradation constants for this pathway calculated from MC data are in good agreement with those obtained from HPLC. MC as a non-specific technique was shown to be useful in studies of PER when one reaction was present in the sample and also when more chemical and physical processes were simultaneously running.

Stability of new potential ACE inhibitor in the aqueous solutions of different pH

Angiotensin-converting enzyme (ACE) inhibitors are a group of active substances binding to an active site of ACE. Many authors who studied the structure activity relationship suggested the structural elements needed for a potent ACE inhibitor. While many authors studied the activity of ACE inhibitor substances only a few structure stability studies have been presented. In this paper the stability properties of molecule xPRIL were studied by determination of degradation path and rate of degradation in aqueous solutions with different pH (2.0, 6.8 and 12.0) and temperatures (40, 60 and 80 degrees C). The degradation of molecule through two main degradation paths was identified and confirmed by liquid chromatography and mass spectroscopy (LC-MS). Stability properties of xPRIL were determined in a stability study evaluated by high-performance liquid chromatography (HPLC). The first order kinetics of degradation reaction of xPRIL and Arrhenius equations for each pH were determined at observed conditions. xPRIL showed the highest stability at pH 2 solution. The degradation kinetics of xPRIL was compared to the degradation kinetics of enalapril maleate (EM) and perindopril (PER) in bio relevant solutions with pH 2.0 and 6.8. In addition to the stability study of xPRIL the forced degradation study of all three molecules at rigorous conditions was conducted. From the obtained results the structural element having the highest influence on stability properties of the studied molecules was identified. The fragmentation paths of xPRIL, its cyclization degradation product and its hydrolysis degradation product were identified and confirmed by MS/MS method.

Are calculated log P values for some guanine derivatives by different computer programs reliable

The log P values of n-octanol/water for some guanine derivatives, acyclovir, deoxyaciclovir and their acetyl congeners, were calculated by some commercially available computer programs for log P calculation. These values were compared with those obtained by the conventional shake-flask method. It was established that the calculations of log P values for examined guanine derivatives by these computation programs do not give reliable results.

Chromatographic and permeation analysis of ciprofloxacin metal complexes

Aqueous solutions of ciprofloxacin and its metal complexes with Mg, Cu, Bi and Fe were investigated with the aim to characterize their retention and permeability properties with consequent complex stability evaluation. For this purpose HPLC coupled with UV detector for ciprofloxacin and with atomic absorption spectrometer for metal ions detection was used. Determination of lipid barrier permeability was carried out using a Sartorius absorption apparatus with the lipid-type of membrane. Under the applied experimental conditions, no significant differences in retention and permeability properties were found, indicating the low stability of studied ciprofloxacin complexes in aqueous solutions.

Preservative Efficacy Screening of Pharmaceutical Formulations Using ATP Bioluminescence

The preservative challenge test is a method used to determine the efficacy of a preservation system in a pharmaceutical or cosmetic formulation. However, such testing is a labor-intensive, repetitive task often requiring days before results can be generated. Several alternatives to traditional colony-count techniques have been developed. A study using pure suspensions of Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Bacillus subtilis, Candida albicans, and Aspergillus niger showed that the accuracy, repeatability, and linearity of the Pallchek™ luminometer ATP bioluminescence (ATP-B) system was equivalent to the traditional colony-count method. In any case, the method proved sensitive enough to follow the effect of preservatives on a number of test microorganisms, indicating the applicability of the ATP-B method for preservative screening studies in various pharmaceutical formulations.

Evaluation of drug stability data by analog-hybrid computer: application to lorazepam

Abstract The criteria for evaluation of reaction order and pathways of degradation in drug stability studies were investigated. The applicability of classical methods (linear regression, linear regression forced through origin, mean square error) and analoghybrid simulation is discussed using lorazepam solution stability as an example. Experimental data for lorazepam degradation and quinazolinecarboxaldehyde derivative formation were obtained by HPLC and then evaluated on microcomputer, Apple II, and analog-hybrid computer, EAI 580. First-order kinetics for lorazepam degradation at declared storage conditions was confirmed. In addition the kinetic model for lorazepam degradation pathway was derived. The advantages of analoghybrid simulation in such studies are pointed out.