Ziling Lu

Dependence of reversed-phase retention of ionizable analytes on pH, concentration of organic solvent and silanol activity

In reversed-phase chromatography, the retention of ionizable analytes is influenced by the ionic properties of the packing caused by surface silanol groups. We have measured the ion-exchange properties of both reversed-phase bonded phases and their underlying base materials. The probe used in this part of the study was bretylium tosylate. The acquired knowledge is then used for a complete and quantitative understanding of the retention behavior of ionizable compounds as a function of the pH of the mobile phase and the solvent composition. We have studied the retention pattern of a broad range of acids, bases, and polyfunctional analytes over the pH range from 2 to 11 and from water to 80% acetonitrile. A few application examples demonstrate the relevant findings.

Ultrafast liquid chromatography/ultraviolet and liquid chromatography/tandem mass spectrometric analysis

Optimal liquid chromatography/mass spectrometric [LC/MS(/MS)] analysis depends on both the LC selectivity and the electrospray efficiency. Here, we outline a simple and comprehensive LC/MS/MS strategy for the rapid analysis of a wide range of pharmaceutical compounds. To achieve ultrafast LC separation with little sacrifice in peak capacity, one needs to start with a column that provides a good peak capacity at short gradient run times; secondly, it is important to use high flow rates to achieve a good gradient peak capacity. Following this strategy, it was possible to baseline-resolve a mixture (containing acidic, neutral, and basic pharmaceutical analytes) in seconds. By coupling the selectivity provided by fast LC separation with the specificity of MS/MS detection, it is possible to separate and identify a wide range of analytes in 1-min gradient analyses. Also, the impact of mobile phase pH on both the chromatographic selectivity and the MS/MS sensitivity is demonstrated.

At-column dilution for improved loading in preparative chromatography

SummaryMaximizing column loadability is an important part of preparative chromatography, since both the cost of the equipment and the cost of the operation increase with the size of the column. We have studied the parameters that determine column loadability. In this publication, we describe the influence that the choice of the sample solvent and the nature of the sample have on loadability. If the sample is injected onto the column in the classical way, overload effects result from the choice of conditions that favor a high solubility of the sample. We then demonstrate how a sample introduction technique called at-column dilution circumvents the difficulties associated with the limited solubility of samples under the preferred conditions of preparative chromatography. The technique is discussed in detail.

Chapter 3 - Techniques for sample preparation using solid-phase extraction

This chapter describes techniques for sample preparation using solid-phase extraction (SPE). The 1-D reversed-phase sample clean-up method is a simple technique designed to remove major interferences such as plasma proteins or polar compounds. The 2-D reversed-phase solid-phase extraction method provides much cleaner backgrounds, but the development of the method is more complex. A range of useful methods for the sample preparation of samples of biologic origin (plasma and urine) prior to high-performance liquid chromatography (HPLC) or HPLC and mass spectrometry (MS) or MS analysis are discussed in the chapter, with an emphasis on the general principles of each method. Each method can carried out with multiple analytes and can be adapted to related sample preparation problems without difficulties. However, occasionally, the particular properties of a sample matrix, an analytical technique, or even simply the analytes require departures from the details of the approaches described in the chapter. Both offline and online SPE techniques have advantages depending upon the particular application, and additional progress in the future is anticipated with respect to the speed and efficiency of both approaches.