Separation techniques stationary phases

The field of liquid chromatography is well established, and reliable methods have been developed for analytical and preparative separations. Column miniaturization improves performance for analytical separations. Numerous stationary phases have been developed to separate analytes based on a wide variety of molecular properties including hydrophobicity, ionic interactions, and molecular size. Mobile-phase modifiers can be used to aid in the niinumzation of unwanted interactions with the solid support. Although the field is well established, current research continues to improve separations for both microscale analytical and larger preparative separations. Recent publications will be highlighted that demonstrate the developments toward integrating HPLC components and separation techniques onto microfabricated devices. 

Technique Stationary phase Mobile phase Format Mechanism of separation... 

Analytical separations may be classified in three ways by the physical state of the mobile phase and stationary phase by the method of contact between the mobile phase and stationary phase or by the chemical or physical mechanism responsible for separating the sample s constituents. The mobile phase is usually a liquid or a gas, and the stationary phase, when present, is a solid or a liquid film coated on a solid surface. Chromatographic techniques are often named by listing the type of mobile phase, followed by the type of stationary phase. Thus, in gas-liquid chromatography the mobile phase is a gas and the stationary phase is a liquid. If only one phase is indicated, as in gas chromatography, it is assumed to be the mobile phase. 

Capillary Electrochromatography Another approach to separating neutral species is capillary electrochromatography (CEC). In this technique the capillary tubing is packed with 1.5-3-pm silica particles coated with a bonded, nonpolar stationary phase. Neutral species separate based on their ability to partition between the stationary phase and the buffer solution (which, due to electroosmotic flow, is the mobile phase). Separations are similar to the analogous HPLC separation, but without the need for high-pressure pumps, furthermore, efficiency in CEC is better than in HPLC, with shorter analysis times. 

Chiral Chromatography. Chiral chromatography is used for the analysis of enantiomers, most useful for separations of pharmaceuticals and biochemical compounds (see Biopolymers, analytical techniques). There are several types of chiral stationary phases those that use attractive interactions, metal ligands, inclusion complexes, and protein complexes. The separation of optical isomers has important ramifications, especially in biochemistry and pharmaceutical chemistry, where one form of a compound may be bioactive and the other inactive, inhibitory, or toxic. 

Synthetic chiral adsorbents are usually prepared by tethering a chiral molecule to a silica surface. The attachment to the silica is through alkylsiloxy bonds. A study which demonstrates the technique reports the resolution of a number of aromatic compoimds on a 1- to 8-g scale. The adsorbent is a silica that has been derivatized with a chiral reagent. Specifically, hydroxyl groups on the silica surface are covalently boimd to a derivative of f -phenylglycine. A medium-pressure chromatography apparatus is used. The racemic mixture is passed through the column, and, when resolution is successful, the separated enantiomers are isolated as completely resolved fiactions. Scheme 2.5 shows some other examples of chiral stationary phases. 

When multiple development is performed on the same monolayer stationary phase, the development distance and the total solvent strength and selectivity values (16) of the mobile phase (17) can easily be changed at any stage of the development sequence to optimize the separation. These techniques are typically fully off-line modes, because the plates must be dried between consecutive development steps only after this can the next development, with the same or different development distances and/or mobile phases, be started. This method involves the following stages ... 

HPLC separations are one of the most important fields in the preparative resolution of enantiomers. The instrumentation improvements and the increasing choice of commercially available chiral stationary phases (CSPs) are some of the main reasons for the present significance of chromatographic resolutions at large-scale by HPLC. Proof of this interest can be seen in several reviews, and many chapters have in the past few years dealt with preparative applications of HPLC in the resolution of chiral compounds [19-23]. However, liquid chromatography has the attribute of being a batch technique and therefore is not totally convenient for production-scale, where continuous techniques are preferred by far. 

Enantiomeric separations have become increasingly important, especially in the pharmaceutical and agricultural industries as optical isomers often possess different biological properties. The analysis and preparation of a pure enantiomer usually involves its resolution from the antipode. Among all the chiral separation techniques, HPLC has proven to be the most convenient, reproducible and widely applicable method. Most of the HPLC methods employ a chiral selector as the chiral stationary phase (CSP). 

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