Self-Assembly of Cyclodextrins and Their Complexes in Aqueous Solutions.

Cyclodextrins (CDs) are enabling pharmaceutical excipients that can be found in numerous pharmaceutical products worldwide. Because of their favorable toxicologic profiles, CDs are often used in toxicologic and phase I assessments of new drug candidates. However, at relatively high concentrations, CDs can spontaneously self-assemble to form visible microparticles in aqueous mediums and formation of such visible particles may cause product rejections. Formation of subvisible CD aggregates are also known to affect analytical results during product development. How and why these CD aggregates form is largely unknown, and factors contributing to their formation are still not elucidated. The physiochemical properties of CDs are very different from simple amphiphiles and lipophilic molecules that are known to self-assemble and form aggregates in aqueous solutions but very similar to those of linear oligosaccharides. In general, negligible amounts of aggregates are formed in pure CD solutions, but the aggregate formation is greatly enhanced on inclusion complex formation, and the extent of aggregation increases with increasing CD concentration. The diameter of the aggregates formed is frequently less than about 300 nm, but visible aggregates can also be formed under certain conditions.

Optimization of enantiomeric separations in capillary electrophoresis by applying a design of experiments approach.

During early-phase pharmaceutical development, it is important to find an initial separation of enantiomeric compounds quickly in order to determine the enantiomeric purity of chiral drug substances. Highly selective screening methods are necessary to analyze the products to discover a satisfactory separation of the enantiomeric compounds. A screening approach based on the use of mixtures of multiple cyclodextrins in chiral capillary electrophoresis was employed to find the initial separation of chiral compounds. In a later phase of development, these complex methods need to be simplified for transferability. This study describes the simplification of the complex mixture of cyclodextrins into a single or dual system of only the enantioselective cyclodextrins. This was achieved by applying fractional factorial experimental designs to select the cyclodextrins that were responsible for the enantiomeric separation and response surface modeling designs for the optimization of the separation. In order to obtain robust methods, the concentration of the chiral selector, together with other important electrophoretic method parameters such as the concentration of the background electrolyte, pH, and run voltage, were optimized by employing a Box-Behnken experimental design.

A screening strategy for the development of enantiomeric separation methods in capillary electrophoresis.

Method development of enantiomeric separations in capillary electrophoresis (CE) is a time-consuming task, since finding the appropriate chiral selector is usually a "trial and error" process. It is impossible to predict the selectivity of a selector towards a certain enantiomer. Therefore, the affinity of all selectors has to be examined one at a time. In order to speed up this process, a strategy is proposed based on simple experimental design methodology. The approach includes first a screening in function of the pH to determine the optimal migration conditions followed by a selection of the right chiral selector by means of Taguchi designs. In the approach several variables, such as the type and concentration of cyclodextrin, the concentration of buffer electrolyte, and the percentage of organic modifier, are varied simultaneously to find initial separation conditions rapidly. The resulting initial separation conditions can be optimized in further steps to be more reproducible. We discuss the results of the approach when applied on a number of selected compounds that are recently in development at Johnson & Johnson--Pharmaceutical Research and Development. Parameters, such as quality of the separation and analysis time, are evaluated to determine initial separation conditions for each compound.

Optimization and validation of an enantioselective method for a chiral drug with eight stereo-isomers in capillary electrophoresis.

Highly selective capillary electrophoresis (CE) screening methods were applied to find a satisfactory separation of a chiral drug with eight stereoisomeric compounds. The initial separation conditions were further optimized using response surface modelling by applying a Box-Behnken experimental design. This approach resulted in a rapid and efficient optimization of the buffer concentration, the concentration of two cyclodextrins, and the run voltage, in order to obtain final separation conditions of the method. Further optimization and validation of the system in terms of sensitivity and robustness resulted in a method that is suitable for quality control release purposes.

Capillary electrophoresis as an orthogonal technique in HPLC method validation.

High-performance liquid chromatography is usually used to assay the main compound and organic impurity content of drug substance and drug product during pharmaceutical development. A crucial validation parameter of these methods is specificity--the ability to unequivocally assess the analyte in the presence of component expected to be present. Typically, these include impurities, degradation products, and matrices. Besides adequate chromatographic separation with sufficient selectivity, additional 2- or 3-D spectroscopic or chromatographic tools are frequently necessary for this purpose. In our current practice, HPLC is used with ultraviolet photodiode array detection and on-line mass spectrometry (LC-UVDAD-MS) during the assessment of specificity. Although this approach is very powerful and can solve the majority of problems, separation of isomers of the main compound is still difficult. Since HPLC usually cannot offer the required selectivity and because of the similar molecular weights, structural isomers are not specifically detected using LC-MS. Capillary electrophoresis, on the other hand, offers high separation efficiency and can be applied as an adjunct to HPLC. Therefore, a set of highly selective CE methods is used orthogonally in the specificity assessment of HPLC methods.

Fast method development and rapid analysis using a screening approach for enantiomeric separations in capillary electrophoresis.

In order to speed up the trial-and-error process during enantioselective capillary electrophoresis methods development, a systemized approach is proposed to develop methods by applying several screening methods in the search for an initial separation. Screening methods combine high selectivity with broad applicability and are applied to find an initial enantiomeric separation during early pharmaceutical development (pre-Phase 1 to Phase 1). The goal is to achieve enantiomeric separation rapidly in order to characterize the chiral purity of pharmaceutical products. Dedicated, highly efficient screening methods are suggested for basic, neutral, and acidic compounds. In these screening methods, multiple chiral selectors are applied in mixtures at different buffer pH values. For the compounds studied, the technique allows fast method development. Furthermore, it is potentially applicable to a wide range of low-molecular-weight compounds and permits rapid analysis at low cost, since runs are performed in inexpensive, bare silica capillaries using ordinary buffer systems with only typical cyclodextrins as the selector. Along with simplicity and robustness, the approach results in sufficient efficacy (i.e., it is easy, straightforward, and reproducible, with a high success rate). Typical pharmaceutical applications are described. The major advantage of the screening approach to methods development is the decrease in development cycle time. The total screening time for one compound was about 5.3 hr on one CE instrument.