Aqueous Normal Phase ANP and how can it help you – FAQ

Aqueous Normal Phase Chromatography Definition: Aqueous Normal Phase chromatography ANP is an HPLC retention mechanism which encompasses the HPLC mobile phase region between Reversed Phase chromatography (RP) and organic Normal Phase chromatography (ONP) and is used mainly for polar compound separations. Background Information: In normal phase chromatography, the stationary phase is polar and the mobile […]

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Aqueous Normal Phase ANP HPLC retention mechanism elucidated.

The mechanism of aqueous normal phase (ANP) retention is an active area of investigation but the consensus is agreed upon. Much progress has been made in recent years at elucidating how polar compounds are retained by this mechanism and how it differs from the other techniques used for the analysis of hydrophilic or polar molecules.  A peer […]

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Aqueous Normal Phase ANP HPLC what is it – FAQ

Aqueous normal phase chromatography (ANP) is a chromatographic technique which uses the mobile phase region between reversed-phase chromatography (RP) and organic normal phase chromatography (NP) used mainly to separate polar compounds such as acids, bases and peptides.. Uses a slightly hydrophobic stationary phase such as silica hydride (available as Cogent TYPE-C™ HPLC columns) a mobile […]

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Aqueous Normal Phase ANP solvent strength order – FAQ

In Aqueous Normal Phase ANP HPLC, the strongest solvent is water. The order in decreasing solvent strength is: DI water > methanol > isopropanol > acetone > acetonitrile These are general estimates; as an example, in a comparison of acetone and acetonitrile for different amino acids, retention is somewhat comparable for some but significantly different for […]

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Can Reversed Phase and ANP mechanisms operate with the same effect at the same time – FAQ

With Cogent TYPE-C HPLC columns, it is unlikely that Reversed Phase (RP) and Aqueous Normal Phase (ANP) retention contributions would be equal for different amphiphilic compounds under the same conditions. It would require that both mechanisms be operating at equal efficiency for those particular compounds under the same method conditions.  In most cases, one mechanism would be more […]

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Do the Cogent Silica-C columns contain any carbon – FAQ

No! The Cogent Silica-C™ columns are totally inorganic and are made, completely of silicon, hydrogen and oxygen. These columns are designed for improved Normal Phase HPLC methods. Any apparent retention of non polar compounds you may observe is most likely due to Aqueous Normal Phase ANP retention mechanism and your method.

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How can I improve the accuracy and precision when separating polar compounds with HPLC using Aqueous Normal Phase ANP- FAQ

In most cases the average level of accuracy and precision in a HILIC analysis for example is less than typical values obtained by Reversed Phase RP HPLC.  Reversed Phase methods are not usually effective for polar compounds. It is believed that the accuracy and precision of a HILIC analysis is strongly influenced by the equilibration time if a gradient method is […]

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How does efficiency in Aqueous Normal Phase ANP compare to HILIC in HPLC – FAQ

Aqueous Normal Phase ANP HPLC is a distinctly different retention mode than Hydrophilic Interaction Liquid Chromatography HILIC. In HILIC, evidence has shown that retention is achieved by partitioning in and out of a water layer surrounding the stationary phase surface. With a much less polar surface TYPE-C silica columns do not exhibit this same partitioning retention […]

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How to fix a saw tooth pattern in the UV baseline – Tips & Suggestions

This saw tooth pattern may be due to the presence of an immiscible solvent remaining on the column. Even trace amounts of these solvents can cause issues. The immiscibility manifests as an unstable baseline, such as the example shown below. Suggestion: In order to fix this issue, try flushing the column overnight with a mutually […]

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Negative Baselines in HPLC at 214nm – Tips & Suggestions

When I run gradients on my HPLC and switch from one gradient to another, my baseline shows as “negative”. If I auto zero, it becomes positive for the remainder of the run but when I start again, it is negative. What can you suggest? Negative baselines in gradients are not that unusual. If the method is […]

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Store and condition Cogent TYPE-C HPLC columns – How To

This article describes how best to prepare Cogent TYPE-C Silica™ HPLC columns for initial use when you first receive them as well as how to store them, for either overnight or long-term storage to prevent column degradation. Storage Solvents for Reversed Phase RP or Aqueous Normal Phase ANP: If Reversed Phase and Aqueous Normal Phase […]

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The effect of ionic strength of the mobile phase additive for HPLC columns in Aqueous Normal Phase – Tips & Suggestions

Ionic Strength of Additives can affect your Retention Times. Ionic strength has a smaller effect in Cogent TYPE-C Columns when used in Aqueous Normal Phase (ANP) than in methods with HILIC columns because the ANP mechanism is a type of “competitive adsorption” in the adsorbed layer with Acetonitrile, on the silica-hydride surface.   Ionic strength would […]

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Tips on Solving Baseline Problems in ANP – Tips & Suggestions.

When using a gradient method with the Cogent Diamond Hydride™ HPLC column, if an inconsistent baseline may be observed… The following are some possible causes of the issue along with suggestions to fix it. 1. Is the mobile phase filtered before use on the instrument? A 0.45µm nylon membrane filter with vacuum filtration is recommended […]

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