IEX chromatography

The IEX chromatography possesses several attractive features as a first-dimension separation mode in an MDLC scheme. These features include the following ... 

Elution with salt pulses A multiple step elution is performed by the introduction of, for example, 5%, 10%, 25%, 50%, and 100% of 1.5 M sodium chloride in 19 mM phosphate buffer (pH 2.5) containing 5% methanol. Each step is for 10 min and run at 0.5 mL/min. This elution method compromises analytical system dimensionality, as the peak capacity of the ion-exchange chromatography (IEX) step is equal at most to the number of salt steps. However, in the second dimension only one or two columns are needed and there is no particular limitation in the second dimension separation time as peptides are eluted in portions in a controlled manner. However, the number of salt steps is limited by the total analysis time. In this case the multidimensional system is relatively simple. 

Ion-exchange chromatography (IEX-HPLC) is intended to separate proteins based on differences in their total charge. Both anionic and cationic exchange modes are available. This technique can potentially detect and... 

Abbreviations for type of application IEX Ion Exchange, HIC Hydrophobic interaction chromatography, AF Affinity chromatography, RP Reverse phase, IMAC Immobilized metal affinity chromatography. 

Chromatography is one of the most important tools in protein purification. Chromatographic purification techniques include affinity chromatography (AC), ion exchange chromatography (IEX), hydrophobic interaction chromatography (HIC), and gel filtration (GF). These techniques separate proteins according to differences in specific protein properties. The protein property used for separation, the attributes of each technique, and its suitability for different purification steps are summarized in Table 32.7. 

Ion exchange chromatography (IEX) Charge High speed High capacity High resolution + + + + + + + + +... 

As evidenced by the over 360 references retrieved from SciFinder with the keywords coumarins high performance liquid chromatography , a variety of approaches have been used for the separation of coumarins. Coumarins have been separated by RP-HPLC, 46 47 188"192 HP-NPC,193-198 HP-IEX,199 and/or HP-AC.24 The dominant mode for the separation of coumarins is HP-RPC and to a lesser extent HP-NPC, used in conjunction with UV and MS detection. 

HILIC uses a polar stationary phase to separate proteins and peptides. Samples are loaded onto a column using a low aqueous mobile phase and are eluted by increasing the water content of the mobile phase (152). HILIC is very suitable for separation of polar species such as sialic acid-containing glycopeptides, which have limited retention on most RP materials (153). A variant of HILIC is electrostatic repulsion-hydrophobic interactions chromatography and it utilizes IEX stationary phase and a high organic mobile phase (152,154). 

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