Solid-phase extraction (SPE for short), also known as solid-phase extraction, is a pretreatment technology based on liquid-solid chromatography separation, which can selectively adsorb and selectively elute target compounds. According to its retention mechanism, the force is mainly divided into non-polar interaction, polar interaction, ion exchange (electrostatic attraction).

Ion exchange cartridges are aimed at dissociable charged substances. In the analysis of pesticide residues and animal residues, normal-phase extraction and reverse-phase extraction cartridges are mostly used. Today, I will take you to learn more about normal-phase extraction and reverse-phase extraction.

(source: soogif)

Mechanism of reversed phase action

Reversed-phase mode refers to the separation mode in which the polarity of the stationary phase of the SPE cartridge is less than that of the mobile phase. It mainly utilizes the non-polar interaction between the carbon-hydrogen bond of the stationary phase functional group and the carbon-hydrogen bond of the target compound, and is suitable for the extraction and separation of non-polar to moderately polar targets from polar matrices.

For the target adsorbed on the reversed-phase SPE column by non-polar force, it can be eluted with a weakly polar solvent, such as chloroform, cyclohexane, ethyl acetate, etc. As long as the elution strength of the solvent is sufficient to break the van der Waals forces between the target and the sorbent, the target can be eluted from the SPE column. Even the more polar methanol has sufficient nonpolar forces to elute for many compounds. Sometimes a single solvent cannot completely elute the highly hydrophobic target, so consider using dichloromethane:ethyl acetate (1:1, volume ratio).

In reversed-phase mode, the polarity of the solvent system should gradually decrease in the order of sample solvent, elution solvent, and elution solvent, while their elution strengths gradually increase. It must be ensured that the selected sample solvent cannot elute the target substance, the selected eluent should elute the interfering substance to the maximum extent without eluting the target substance, and the selected eluent should be able to completely elute the target substance.

Mechanism of normal phase action

The normal phase mode refers to the separation mode in which the polarity of the stationary phase of the SPE cartridge is greater than that of the mobile phase. Mainly use:

Polar interactions between polar surfaces of solid-phase extraction materials and polar functional groups of the target. Polar forces are stronger than non-polar forces but less than ionic forces. Common polar functional groups include hydroxyl, amine, sulfhydryl, etc. A weakly polar matrix environment favors the formation of polar forces because weakly polar solvents do not have functional groups capable of hydrogen bonding with polar stationary phases. Therefore, in the normal phase extraction of SPE, the sample matrix is ​​mostly weakly polar, such as n-hexane, dichloromethane, rapeseed oil, etc., and most of the targets contain polar functional groups.

Common polar stationary phases are: silica gel, alumina, Florisil and bonded silica containing cyano groups (CN), amino groups (NH2), and diol groups (2OH).

In normal phase mode, the polarity of the solvent system should gradually increase in the order of sample solvent, elution solvent, and elution solvent, and their elution strengths should also gradually increase. It must be ensured that the selected sample solvent cannot elute the target substance, and the selected eluent should elute the interfering substance to the maximum extent without eluting the target substance, so the eluent should be able to completely elute the target substance.

So in the face of reversed-phase and normal-phase solid-phase extraction, which one should we choose in practical applications? The following is a detailed analysis for you.

(source: soogif)

Since most compounds have multiple functional groups, in practice, which extraction mechanism to use is mainly based on the nature of the target and interfering substances.

For example, 2-naphthylamine is a weakly basic compound (pKa=4.16), which can be cationic under certain pH conditions, and has both hydrophobic and hydrophilic groups. In this case, the extraction mechanism that is beneficial to the separation of the target from the interference can be selected according to the sample matrix.

When the target is in a weakly polar sample matrix, the polar interaction can be used to select the normal-phase extraction mode; if it is in a polar sample environment, the non-polar interaction can be used to select the reversed-phase extraction mode. In addition, the pH of the extraction environment can also be adjusted to two pH units lower than the pKa of the target substance, that is, pH=2.16, using a cation exchange mechanism. The main factors influencing the choice of retention mechanism are summarized below:

The first factor affecting the retention mechanism: the nature of the functional group of the target

Retention Mechanism Second Affecting Factor: The Nature of the Sample Matrix

Through the above content, I believe that the choice of solid phase extraction mode has been understood by my friends. In addition, Welch Materials has successively launched silica-based reversed-phase, normal-phase, and ion-exchange cartridges for different detection items, as well as SPE products with polymer-based, inorganic-based, and mixed adsorption mechanisms.

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