Ion Chromatography

Ion chromatography is a long established, progressive method for the analysis of different mixtures of anions. When introduced in the late seventies, rather special and complicated equipment was necessary for performing ion chromatography (at that time meaning a method using conductometric detection with chemically suppressed eluent conductivity). Since then, the meaning as well as the possibilities of the new method have grown extensively.

Many types of detectors (photometric, conductometric, refractometric) can now be used with the indirect method of detection. However, nearly all anions (with the exception of fluoride, chloride, sulphate, phosphates, carbonate, borate and a few others) are detectable directly by UV photometry in the range 200-215 nm. Low capacity anion exchangers are widely used in the ion chromatography of inorganic and organic anions. Columns packed with these sorbents ensure fast, simple and sensitive determination of the anions, using practically any liquid chromatograph. TESSEK introduces a new TESSEK Separon HEMA-S 1000 Q-L, quaternary amine anion exchanger with a capacity of around 0.1 mmol/g. This sorbent was found to be well suited for all detection modes commonly used in ion chromatography, including suppressed and nonsupressed conductivity, indirect photometry and others.

Ordering Information

The sorbent is available in 10 g, 100 g and bulk quantities.

Packed Columns

Following packed columns are available. The 3 mm ID x 150 mm compact glass cartridges are supplied single-packed in a sealed aluminum foil bag. The 3 mm ID x 30 mm cartridges are packed as three pieces in an aluminum foil bag.

Hydrophobic Interaction Chromatography

Hydrophobic interaction chromatography is one of separation methods widely applied to separation of proteins. In contrast to reversed phase chromatography the separation under nondenaturing conditions allows isolation of highly active proteins from complex natural mixtures. The method is based on elution of proteins from slightly hydrophobic sorbent by descending gradient of ionic strength, i.e. ammonium sulfate in phosphate buffer. The slightly hydrophobic nature of hydroxyethyl methacrylate gel TESSEK Separon HEMA-S 1000 allows its direct use for hydrophobic interaction chromatography. However, the best results are obtained on especially developed sorbent TESSEK Separon HEMA-BIO 1000 Phenyl.

This sorbent contains approx 0.1 mmol of phenyl groups attached to a highly hydrophylic surface of HEMA-BIO 1000 matrix.

Ordering Information

The sorbent is available in 10 g, 100 g and bulk quantities.

Packed Columns

Following packed columns are available. The 3 mm ID x 150 mm compact glass cartridges are supplied single-packed in a sealed aluminium foil bag. The 3 mm ID x 30 mm cartridges are packed as three pieces in an aluminium foil bag.

Protein A Affinity Chromatography

The increasing demand for pure antibodies has been met by the development of several isolation and purification procedures. For the isolation of whole IgG and the separation of IgG sub-classes some of the most advantageous methods are based on affinity chromatography. The ability of Protein A from Staphylococcus aureus to bind IgG's specifically through their constant Fc region gives Protein A affinity towards a broad range of mammalian IgGs. By varying experimental conditions, selectivity towards many sub-classes can be achieved, too. In the TESSEK Separon HEMA-BIO 1000 rProtein A, recombinant Protein A is immobilized through divinyl sulphone activation chemistry, which ensures highly active ligands with low leakage. TESSEK Separon HEMA-BIO 1000 rProtein A is applicable in HPLC, prep-scale, as well as in stirred reactors under a wide range of conditions (use of organic solvents, bases, etc.). The binding capacity varies with different species of IgG's. The capacity for human IgG is 10-15 mg/g . Several mouse monoclonal antibodies have been purified on Separon HEMA-BIO 1000 rProtein A giving capacities of 5-10 mg/g.

Ordering Information

The sorbent is available in 10 g, 100 g and bulk quantities.

Packed Columns

Following packed columns are available packed with 10 µm particles. The 3 mm ID x 150 mm compact glass cartridges are supplied single-packed in a sealed aluminium foil bag. The 3 mm ID x 30 mm cartridges are packed as three pieces in an aluminium foil bag. Cool storage is recommended. Plastic cartridges packed with 60 µm sorbent are available (Cat.No. 421 172983)

Metal Interaction Chromatography

The immobilized metal affinity chromatography (IMAC) is performed on sorbents carrying immobilized metal ions. This immobilization is carried out with the help of different chelating ligands bonded to the surface of suitable carriers. The complexes of different stability can be formed between the immobilized metal ion and a protein if electron donors are accessible on the surface of the protein. These electron donors are mainly histidine, but also tryptophane and cysteine residua or phosphate groups in the case of phosphorylated proteins. The stability of these complexes depends on the type of chelating ligand, on the chosen metal and on the amount of accessible donors on the surface of proteins, the sterical accessibility of donors and acceptors. Temperature, pH and the presence of competitive donors in solution play a role, too. The complexes are usually formed at pH 6-8. The elution of proteins from IMAC sorbents can be carried out by gradient of pH or, more often, by the increasing concentration of glycine, histidine, histamine, cysteine and mainly imidazole. The molecules of these compounds include the atoms of nitrogen and/or sulfur and so can act as the donors of electrons competing with retained proteins. The matrix used for the synthesis of IMAC sorbent should be hydrophilic sufficiently to suppress hydrophobic interactions as low as possible. Eluents contain 0.5-1.0 M NaCl usually to avoid ionic interactions.

TESSEK has developed sorbent TESSEK Separon HEMA-BIO 1000 IDA with iminodiacetic acid (IDA) covalently bonded to the surface of hydrophilic, rigid methacrylate matrix. This material keeps all the excellent characteristics of TESSEK Separon HEMA sorbents - high mechanical and chemical stability, favourable hydrodynamic behaviour and high resistance to biological degradation.

Ordering Information

The sorbent is available in 10 g, 100 g and bulk quantities.

Packed Columns

Following packed columns are available. The 3 mm ID x 150 mm compact glass cartridges are supplied single-packed in a sealed aluminium foil bag. The 3 mm ID x 30 mm cartridges are packed as three pieces in an aluminium foil bag.

Column for Isolation of PCBs and PAHs from Fats

Polychlorinated biphenyls (PCBs) are ubiquitous environmental pollutants, that are strongly accumulated in lipid portion of biological matrices, especially in storage fat. Gas chromatography (GLC/ECD) is employed for the determination of these contaminants. Polycyclic aromatic hydrocarbons are comprising other important group of pollutants, most comonly determined by reversed phase HPLC with fluorimetric detection. Prior to GLC analysis, however, removing of interfering co-extracted compounds has to be carried out. Adsorption chromatography on Florisil, liquid-liquid partition or solid phase extraction are often utilized for this purpose. Recently many laboratories have been introducing gel permeation chromatography (GPC) as a clean-up step involved in sample preparation procedure. In comparison with the above methods, GPC possesses many advantages, the main of them is the possibility of multiple use of the same column without any loss of reproducibility of its separation properties. Automatization of the cleaning procedure is thus possible.

Efficient separation of PCBs and PAHs from fat, oils and some other compounds extracted from fats and oils can be achieved on styrene-divinylbenzene copolymer Bio Beads S-X3 (Bio Rad, South Richmond, CA, USA, particle size 37-75 µm) with chloroform mobile phase. Columns packed with this sorbent, that are now offered by TESSEK, are expected to find a wide use in all the laboratories being engaged in analyses of PCBs, especially in foodstuffs and many other biological matrices. For other applications (e.g. pesticides in plant material) the column can be prepared on request also in other mobile phase.

Ordering Information

Ion Exclusion Chromatography

HPLC chromatography is an indispensable tool for separating saccharides and organic acids. Several chromatography modes can be successfully employed. In last years, the ion exclusion mechanism is widely employed for those analyses. Highly crosslinked, sulfonated polystyrene-divinylbenzene gels in different ionic forms are the most commonly used. TESSEK introduces columns of this type packed with OSTION cation exchange resins in hydrogen, sodium, calcium and lead forms.

The resin in hydrogen form is intended for separation of organic acids. Dilute acid (0.01 M sulfuric acid is recommended) is used as an eluent. Ambient temperature can be used, however the resolution increases with increased temperature. Detection is most commonly accomplished by UV detection at 205 - 220 nm. For hydrophobic or aromatic acids, some organic solvent (methanol, acetonitrile) can be added up to 20 %. Sugars and alcohols also can be separated with pure water eluent at 80°C and RI detection, however some oligosaccharides will be hydrolyzed.

In ion exclusion mechanism, the selectivity of separation could not be influenced significantly by mobile phase. Instead, other ionic forms (sodium, calcium or lead) or degree of crosslinking are used for solving special separation problems. Pure water is used as eluent at increased temperature (60 - 80°C). Refractive index detection is necessary. The efficiency of separation increases significantly with decreased flow rate, thus 0.3 - 0.5 ml/min are recommended. Maximum pressure is 7 MPa.

The sodium form column offers improved selectivity for sacchrides, while it can tolerate salt containing samples. It is recommended for direct analysis of molasses and other buffered samples.

The calcium form column offers improved selectivity for monosaccharides. Sample deionization is recommended as long as anions forming insoluble calcium salts will destroy the column selectivity.

The lead form column offers best selectivity for certain sugars, however it is very sensitive to sample deashing. On line or off line sample deionization is strongly recommended.