SieLC is a pioneer in mixed mode chromatographic materials and columns.
Product range is structured as follows:
SIELC Technologies has developed a new generation of unique stationary phases combining mixed-mode and core-shell approaches -CoresepTM
In single single mode on a reversed-phase column compounds elute according to their hydrophobicity and analytes with similar hydrophobicity may co-elute or show poor resolution. In Ion-Exchange columns compounds are retained based on their ionic properties. Similarly, using ion-exchange chromatography, other compounds may be closely eluting because they are identical in terms of the strength of ionic interaction between stationary phase and analyte
In mixed-mode chromatography you have two interactions, reversed-phase and ion-exchang. You are exploring a very small difference in reversed-phase and ionic properties of compounds. These differences are enhanced due to synergy of two mechanisms which results in much better separation (mixed-mode example).
Core-shell particles are constructed with solid core and porous outer layer. This porous layer is used as a stationary support for chemical attachment of specific separation phases.
The reduced pore length offers faster diffusion of molecules and as a result has the ability to do chromatography faster compared to particles with through-pore structure. The same result can beachieved with smaller particles, but then the pressure needs to be higher in order to produce the same flow rate and speed of analysis.
Core-shell technology offers the unique ability to achieve high efficiency at higher flow rates whilst generating back pressures which can be achieved using traditional HPLC systems. This offers the end user the opportunity to improve separation speed and quality without the requirement to¬ purchase UHPLC equipment. We have recognized the exciting opportunities afforded by combining these two great technologies in stationary phases which achieve unique selectivity and high efficiency with traditional HPLC systems which are rated to 4000 or 6000 psi.
Mixed-mode columns offer much higher capacity and retention than traditional reversed-phase columns. Since the surface area of core-shell particles is reduced, mixed-mode phases address potential loadability issues whilst maintaining the selectivity advantages of multiple interactions. Newly optimized ligand density and ionic / hydrophobic ratios further increases the capacity of mixed-mode core-shell columns.
Coresep 100 is a reverse-phase analytical column with embedded acidic ion-pairing groups embedded on core-shell particles.
Coresep 100 columns are available in all standard dimensions
Coresep SB is a reverse-phase analytical column with strong embedded basic ion-pairing groups embedded on care-shell particles.
Coresep SB columns are available in all standard dimensions
Coresep S is a normal-phase analytical column with embedded acidic ion-pairing groups embedded on core-shell particles.
Coresep S columns are available in all standard dimensions
For decades, liquid chromatography stationary phase design has been dominated by the idea of elimination of multiple, or "unwanted", interactions that occur in mixed-mode separations. Reverse-phase chromatography is making the point of eliminating silanol interactions with amine-containing analytes by developing base-deactivated phases.
Primesep stationary phases offer unprecedented selectivity in the separation of a broad array of chemical compounds and in multiple applications. A wide range of HPLC columns with id's from 0.5 mm to 22 mm is available. Corresponding Primesep guard columns are available with all stationary phases and do not require holders. SIELC offers a method development service is available for customers. Ask about special custom LC-phases tailored for specific separations.
Primesep Literature for download
Acidic Primesep Phases for cation exchange
SIELC Technologies provides several different columns with negatively charged functional groups due to the embedded anionic ion-paring reagent.
Is the strongest acidic column, while Primesep C is the weakest acidic column. The Primesep 100 and Primesep 200 columns are in between. Difference in the functional group acidity allows selecting the most appropriate column for a particular set of basic compounds that differ in their pKa value.
The embedded acidic functional group can be in an ionized form, or in a non-ionized form, depending on the pH of the mobile phase. In order to get retention by ion-exchange mechanism on Primesep® columns, the pH of the mobile phase should be selected close to, or above, the transition value of embedded acidic groups. Below the transition pH value, the column behaves as a regular reverse-phase column with an embedded non-ionized polar group.
The column name corresponds to pH value of the mobile phase where acid residue on the column switches from ionize form to neutral one.
Is a reverse-phase analytical column with embedded acidic ion-pairing groups.
Primesep 500 columns are available in all standard dimensions
Is a reverse-phase analytical column with embedded weak acidic ion-pairing groups.
Primesep 200 columns are available in all standard dimensions
Is a reverse-phase analytical column with embedded acidic ion-pairing groups.
Primesep 100 columns are available in all standard dimensions
SieLC_Versatility of Primesep Stationary Phases
Is a reverse-phase analytical column with embedded strong acidic ion-pairing groups.
Primesep A columns are available in all standard dimensions
Complex Interaction with Primesep C
SIELC Technologies has developed a new class of columns with combined reverse-phase, cation exchange, and complex-formation properties in interaction with amines, sulfonium, phosphonium and metal ions.
This column has a unique unparallel selectivity for complex mixtures separation.
The pH working range for these columns is from 1 to 7, but their complex formation and cation-exchange properties are substantially suppressed at the pH below 3. In order to facilitate the complex formation, the pH of the mobile phase should remain in the range of 3-7. The degree of complex formation can be adjusted by selecting the pH of the mobile phase.
The unique complex properties of the column can be observed in the separation of alkali metals ions. The ions elute on the Primesep C column in reverse order compared to ion-exchange elution. The unusual elution order is found in primary, secondary, and tertiary amines. Secondary and tertiary amines have higher pKa values than primary amines, which makes them retain longer by ion-exchange mechanism.
With the complex forming properties of Primesep C columns, the primary amines are more retainable than secondary and tertiary amines. This extraordinary elution pattern allows tailoring the separation of amines to provide the most appropriate peak order for quantitation and preparative chromatography.
Is a reverse-phase analytical column with embedded complex-forming groups.
Primesep C columns are available in all standard dimensions
HPLC separation of aromatic compounds on Primesep P columns.
SIELC Technologies has developed a new class of stationary phases, which provides three interactions with analytes:
This stationary phase facilitates the separation of structural isomers of aromatic compounds.
Additional pi-pi interaction creates a difference in the bind state for structural isomers and often resolves critical pairs of compounds. The degree of pi-pi interaction can be adjusted by varying the amount of acetonitrile in the mobile phase
When methanol is used as an organic modifier, the highest degree of aromatic interaction can be achieved.
Is a reverse-phase analytical column with embedded aromatic and acidic ion-pairing groups.
Primesep P columns are available in all standard dimensions
Anion Exchange with Primesep B Type of columns
SIELC Technologies provides 4 types of different B-type columns with positively charged functional groups.
Is a strong basic column. The recommended pH range is from 1.5 to 4 created by the addition of trifluoroacetic, phosphoric or perchloric, or formic acids to the mobile phase.
Is a weak basic column. It also has carboxilic acidic functional groups. At pH of the mobile phase below 5 the acidic groups are not ionized and B2 column surface becomes positively charged. This dual chemistry offers extended pH range from 0.5 to 7, suitable for the ammonium acetate and ammonium formate buffered mobile phases.
Primesep D column
Was originally developed for direct plasma analysis, became very useful for other anion-exchange/reverse phase applications. It has extended pH range from 1.5 to 7 and offers similar properties as Primesep B2 column but the column has no carboxilic groups and remains positively charged throughout the working pH range.
SieLC_Direct Plasma Analysis with Primesep D
The fourth column, which offers anion-exchange properties is Primesep AB column. This column offers also cation-exchange properties and useful in separation of complex mixtures of polar anionic and cationic compounds.
All B-type columns provide at least two main interactions with analytes: the reverse-phase interaction and the anion-exchange interaction. Neutral analytes retained by reverse-phase interaction. The presence of the charged group in the alkyl chain provides additional selectivity uncommon for typical reverse-phase columns. Acidic analytes can be retained by both anion-exchange and reverse-phase mechanisms. To control retention and selectivity, there is a broad selection of the mobile phases with concentration of organic modifier in the 0-100% range. The concentration and the type of an acid will also significantly affect the retention of anionic compounds. Basic compounds can be retained only by the reverse-phase mechanism, but the presence of positively charged groups on both analyte and stationary phases produces the unique selectivity due to the ion-exclusion phenomena.
Primesep B columns retain acid residue in the stationary phase in equimolar amount. When a column switches to the mobile phase with another type of acid, it should be sufficiently equilibrated to replace all counter-ions from the previous mobile phase with new counter-ion.
Is a reverse-phase analytical column with strong embedded basic ion-pairing groups.
Primesep SB columns are available in all standard dimensions
Is a reverse-phase analytical column with embedded basic ion-pairing groups.
Primesep B2 columns are available in all standard dimensions
Is a reverse-phase analytical column with embedded basic ion-pairing groups specifically design for direct plasma analysis.
Does not retain high-molecular weight plasma components.
Primesep D columns are available in all standard dimensions
Is a reverse-phase column with embedded acidic and basic ion-pairing groups.
Primesep AB columns are available in all standard dimensions
Is a normal-phase analytical column with embedded acidic groups.
Primesep N columns are available in all standard dimensions
Is a silica-based acidic-column with hydrophilic properties.
Primesep S2 columns are available in all standard dimensions
HPLC Columns for Separating inorganic anions
Primesep B4 is a reverse-phase (short carbon chain) analytical column with embedded basic ion-pairing groups.
Primesep B4 columns are available in all standard dimensions
Primesep PB HPLC Columns
Primesep PB is a reverse-phase analytical column with embedded aromatic and basic ion-pairing groups.
Primesep PB columns are available in all standard dimensions
Primesep S HPLC Columns
Primesep S is a normal-phase analytical column with embedded acidic ion-pairing groups
Primesep S columns are available in all standard dimensions
Primesep AP HPLC Columns
Primesep AP is a silica-based amino-column with hydrophilic properties.
Primesep AP columns are available in all standard dimensions
Primesep X HPLC Columns
Primesep X is a silica-based amino-column with weak hydrophobic properties.
Primesep X columns are available in all standard dimensions
With the increasing demand for the analysis of polar and ionisable compounds, SIELC Technologies have developed Obelisc - a new generation of mixed mode columns. Obelisc R and Obelisc N are complimentary phases for the separation of polar and non-polar compounds using multiple separation mechanisms (RP, ion-exchange, NP, ion exclusion). Both phases have positive and negative charges separated by a long organic chain, allowing both charges to simultaneously participate in electrostatic
Compound / Phase Interaction Mechanisms
Two complimentary columns, Obelisc R and Obelisc N, based on Liquid Separation Cell ( LiSC ) technology, offer a new approach to separate a variety of small molecules. Buffer concentration, buffer pH, and organic modifier concentration are three orthogonal parameters of the mobile phase that allow adjustment of column properties to separate complex mixtures.
Is a column which has very polar characteristics and works well for polar and charged analytes.
Is a reverse-phase analytical column and can be used in traditional, reversed-phase type applications.
Obelisc R & N HPLC Columns
Promix for biomolecule separation
Promix is an alternative chromatography technology for efficient resolution of peptides and proteins. The technology is based on a combination of two interactions - hydrophobic and ionic. This approach is possible due to a new type of separation media: a chemical combination of hydrophobic and ionic functional groups on a ligand bonded to a silica support. With this phase, unparalleled selectivity and peak capacity can be achieved. Independent adjustment of the amount of buffer and organic modifier creates an infinite number of separation conditions that are suitable for many types of biomolecules.
Promix® Enter a New Era in Biomolecule Analysis with Promix Columns Unsurpassed Selectivity and Peak Capacity for Peptides and Proteins
SieLC_Peptides Analytic with Promix
SieLC_Alternative Selectivity of Promix
SieLC_Peptide Digest Separation with Promix MP
SieLC_Peptides Different Gradient Types with Promix MP
SieLC_Proteins and Peptides on Promix Columns
SieLC_Separation of Insulin Analogues With Promix MP
There are four different Promix Columns to be selected as follows:
Use the following chart to select a column based on the analyte's properties
Select the column length based on sample complexity
Select the column i.d. based on sample loading and sensitivity needs
Mobile Phase should be pH 2-4 with:
Promix SP HPLC Columns
Promix SP is a stationary phase for small peptides under 1 kDa with PI value below 6.0.
Promix SP columns are available in all standard dimensions
Promix AP HPLC Columns
Promix AP is a stationary phase for small and medium size peptides under 3 kDa with PI value below 7.0.
Promix AP columns are available in all standard dimensions
Promix MP HPLC Columns
Promix MP is a stationary phase for medium size proteins and peptides from 1 to 10 kDa.
Promix MP columns are available in all standard dimensions
SHARC HPLC columns are the latest innovation from SIELC Technologies. SHARC columns are the first commercially available columns with separation based primarily on hydrogen bonding. SHARC stands for Specific Hydrogen-bond Adsorption Resolution Chromatography.
Hydrogen bonding is an interaction between hydrogen atom bound to electronegative atoms in a molecule, such as oxygen, nitrogen, fluorine. This is typically a weak interaction, especially when separation is performed in aqueous solutions. Liquid chromatography techniques evolved as tool for separation of different molecules based on their physico-chemical properties. Most common techniques of the separation are:
Columns and stationary phases based on these techniques never perform purely with one type of interaction. Hydrogen bonding is omnipresent in every one of these techniques with minor contribution to retention and selectivity. However, in some cases, especially in normal phase chromatography, the contribution of hydrogen bonding can be significant. SHARC 1 is the first column specifically design to perform a separation based entirely on the interaction of the molecules capable providing hydrogen atom (donor) or attract hydrogen atom (acceptor) to the stationary phase with special properties.
SHARC 1 column operation conditions are unique. A mixture of acetonitrile (MeCN), a weak solvent, and methanol (MeOH), a strong solvent, are used as the mobile phase. Pure MeCN has very insignificant amount of hydrogen bonding with the SHARC stationary phase, while MeOH interacts strongly with SHARC stationary phase, which reduces the retention of analytes based on it’s capacity to hydrogen bond. By changing ratio of MeCN/MeOH the optimum retention profile can be obtained for many types of molecules with high selectivity, peak shape, efficiency, and speed.
Surface interaction within the column with hydrogen bonding Surface interaction within the column with hydrogen bonding
capable analytes with acetonitrile as the mobile phase capable analytes with methanol as the mobile phase
Hydrogen bonding energy is usually in range of 30 kJ/mol or less and depends strongly on the nature of the functional groups and their orientation within the molecule. This energy difference is the basis for selectivity of the interaction between the stationary phase and the molecule of different structure and chemical characteristics.
A given molecule can retain on the stationary phase with more than one hydrogen bond, while also performing as a donor or acceptor of a hydrogen atom.
Presence of a polarized hydrogen atom is not always enough to observe retention by hydrogen bond mechanism. Sometimes a compound can form intramolecular interactions, as oppose to intermolecular hydrogen bonding, and does not participate in stationary phase interaction.
Hydrogen-bonding interaction offers unique selectivity based on number of “interaction points” available for hydrogen bonding. One of the useful characteristics to determine retention patterns in hydrogen-bonding mode is the molecular polar surface area (PSA). This calculated parameter is usually used for prediction of drug transport properties, but we successfully applied it to hydrogen-bonding interactions. Polar surface area is defined as a sum of surfaces of polar atoms (usually oxygens, nitrogens and attached hydrogens) in a molecule. Since those polar atoms can participate in hydrogen-bonding interaction, estimation of elution order can often be made based on PSA. While PSA is a good indicator of elution time, it must be noted that polar surface area does not account for the accessibility of hydrogen-interaction sites. Not every polar surface participates in intermolecular hydrogen interactions with the stationary phase.
Proximity of “interaction points” to each other within one molecule also needs to be considered since molecules can form an intramolecular hydrogen-bonding, which competes with intermolecular interaction between analyte and stationary phase. This reduces retention time in hydrogen-bonding mode. Such structural factors provides unique selectivity among similarly structural (isomers, homologs, degradation products, precursors) molecules.
Since SHARC 1 column is a mixed-mode column, pKa is another useful parameter in method development for these columns. SHARC columns operate in non-aqueous mobile phase, but some effect of charge interaction of stationary phase and ionizable molecules still exists and contributes to the retention profile.
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