Lead Choosing a set of initial conditions that would be likely to produce a successful liquid chromatography (LC) separation was the main topic of discussion in the first post in the HPLC method development series. The majority of samples will benefit from starting with a C8 or C18 column packing based on Type B silica using a mobile phase of acetonitrile or methanol modified with water or a low-pH buffer, even though no single set …
Lead In the last post we saw the tools at your disposal to assess the quality of a chromatogram viz. retention factor, peak asymmetry, peak shape, column plate number, and related aspects. These chromatogram observations are important, but the main purpose of chromatography is to achieve a separation that is reasonable. To ensure whether the separation is satisfactory one need to add one more measurement to the tool box namely the resolution. In this post …
Lead In the previous post, “Measuring quality of chromatogram – Tools 1.0” we talked about two of the separation attributes retention time and retention factor. In current post the discussion is on other features viz. peak asymmetry, peak shape, column plate number. and related aspects. This time let us try to answer the following queries to understand how these attributes also help to assess the quality of chromatogram. Peak asymmetry and peak tailing Peak shape …
Lead In the previous post, we talked about choosing suitable initial conditions when developing a new liquid chromatography (LC) method. The possibility of a successful separation is increased by selecting the appropriate column conditions and mobile phase composition. Getting the controls that quantitatively measure the separation’s quality is another crucial step in the method development process. Although the majority of chromatographers are able to assess the separation quality from a chromatogram, it is critical to …
Lead Starting out with a sound strategy is one of the most important aspects of minimizing chromatographic separation issues. A good method makes it much simpler to maintain operation within parameters and address issues as they appear. Selecting the right tools is very critical to achieve the best results,. By looking at the cause-and-effect diagram given below one can understand the factors that influence chromatographic separation. Basically the factors falls into two major categories namely …
Lead Many chromatographers develop HPLC separations on a daily basis. The philosophy behind method development is founded on a number of factors. Today, chromatographic separation and how it varies depending on the sample and experimental conditions have a good practical understanding. This understanding of the chromatographic process should serve as the foundation for any methodological approach to HPLC method development. Most of the time, a desired separation can be easily accomplished with a minimal number …
Lead Proteins are complex, high-MW biopolymers composed of L-amino acids and possess ordered 3D-structure. They are known to bind /interact stereoselectively with small molecules reversibly, making them extremely versatile CSPs for chiral separation of drug molecules. Number of CSPs has been developed by immobilizing proteins. These type of CSPs operate under RP-mode; (phosphate buffer and organic modifiers) Characteristics of proteins used in commercial CSPs Mechanism Protein polymer remains in twisted form because of the …
Lead William H Pirkle developed these CSPs and are based on the ionic or covalent attachment of one enantiomer of an amino acid derivative to aminopropyl silica. Pirkle phases contain π-electron donor or acceptor rings substituted with H-bonding moieties. Donor–acceptor-type CSPs capitalize on synthetic or semi-synthetic chiral low-molecular-weight chiral selectors capable of recognizing chiral analytes by complementary assembly of nonionic attractive interactions. Basic design and types Generally speaking a chiral phase of the Pirkle type …
Lead Cyclodextrin-based CSP belongs to the cavity type. Armstrong is considered the father of cyclodextrin-based CSPs. CDs are employed in various fields of analytical chemistry, due to their tendency to form reversible inclusion complexes and recognize chiral analytes. CDs are a type of macrocyclic D-(+)-glucopyranose oligomers linked by α-1,4-glycosidic linkages, as seen in Figure, under chemistry . The relevant oligomers for liquid chromatographic enantiomer separation are composed of 6(α-CD), 7(β-CD), and 8(γ-CD) monosaccharide units, in that order. …
Lead Surprisingly, chiral chromatography could not be performed in 1980 because there was no single chiral stationary phase on the market. But as the 1980s came to a close, chiral chromatography began to get more and more attention, thanks in large part to the efforts of Okamoto’s institute in Japan, Pirkle and Armstrong’s teams in the US, Schurig and König in Germany, Lindner in Austria, and Francotte in Switzerland. The most prevalent chiral polymers …