The TESSEK Separon HEMA sorbents are based on a rigid, macroporous and hydrophilic polymer matrix. With the unique combination of stability (mechanical, chemical and biological), macroporous structure and hydrophilic nature, TESSEK Separon HEMA meets the requirements for a versatile, chromatographic sorbent. The basic TESSEK Separon HEMA matrix is synthesized by suspension co-polymerization of 2-hydroxyethyl methacrylate (HEMA) and ethylene dimethacrylate (EDMA). As a result of the polymerization of hydrophilic HEMA units with alternate incorporation of crosslinking EDMA monomers, the growing backbones create a dense, non-penetrable threedimensional network in the form of microspheres (nodules). The microspheres polymerize to agglomerates, forming macroporous particles of a perfect spherical shape. The patented technology involved permits the TESSEK Separon HEMA to be produced with controlled pore and particle dimensions.
The high density of hydroxyl groups on the internal surface provides TESSEK Separon HEMA with hydrophilic properties and excellent derivatization possibilities for the preparation of different chromatographic sorbents, which by the inherent physico-chemical parameters are well suited for the conditions of LC, HPLC and large-scale separations.
The HEMA-S matrix modification is used for the microparticulate macroporous TESSEK Separon HEMA 300 and TESSEK Separon HEMA 1000 sorbents to enhance the rigidity of the sorbent. The initial sorbent is subsequently crosslinked with the EDMA monomer to provide better properties in high pressure chromatography use.
The HEMA-BIO matrix modification was developed to enhance the hydrophilicity of the sorbent. This modification as well as epoxy (E, E-H) and quarternary amine (Q) modifications also result in enhanced rigidity, so the HEMA-S modification is not used here. During the modification some crosslinking occurs, resulting in increased rigidity and slightly decreased working volume and hydroxyl group content.
Standard particle sizes produced are 10 and 60 µm, other
particle sizes are available for specific purposes.
Rigid, macroporous HEMA polymers are formed by agglomerates of small, highly crosslinked particles (nodules). This structure results in a wide pore distribution. The calibration curve is usually linear, or nearly linear from exclusion limit to the low molecular weights.
This is an advantage for characterization of molecular weight distribution, however the slope is quite steep, resulting in a lower selectivity. The resolution is usually lower, than with soft gel or silica based sorbents. The exclusion limits of TESSEK Separon HEMA-BIO are controlled between 40 and 2000 kDa. The pure mechanism of separations by steric exclusion results in a linear dependence of log Mw vs. Ve for a series of protein standards and more than 90% recovery for most proteins tested. The rigidity of HEMA-BIO sorbents allows the use of aqueous as well as organic solvents, including tetrahydrofurane, dimethylacetamide, cadoxene etc.
For preparative separation of proteins, usually they are applicable
only for simple mixtures of proteins with enough differing sizes.
Best results are achieved, when the exclusion limit is selected
so that the bigger molecule is excluded and the smaller is well
in the working range. Use of small particles (10-20 µm) is
usually essential in this technique to achieve sufficient efficiency.
The main advantage of HEMA SEC sorbents is in possibility of their
in column sanitation by 0.1 M NaOH as required by FDA protocols.
TESSEK Separon HEMA and HEMA-S 40or 100 or the additionally croslinked
HEMA-S 300 and 100 sorbents are mostly used in SEC of polysaccharides.
Due to its slight hydrofobicity they can be used for hydrophobic
interaction chromatography of proteins. Compared to HEMA-BIO series
they offer slightly higher working volume.
The following sorbents are available in 10 and 100
g and bulk quantities.
|
|
| |
| HEMA 40 | 40 - 80 | ||
| HEMA 100 | 80 - 250 | ||
| HEMA-S 300 | 250 - 600 | ||
| HEMA 300 | 250 - 600 | ||
| HEMA-S 1000 | 800 - 2000 | ||
| HEMA 1000 | 800 - 2000 | ||
| HEMA-BIO 40 | 40 - 80 | ||
| HEMA-BIO 100 | 80 - 250 | ||
| HEMA-BIO 300 | 250 - 600 | ||
| HEMA-BIO 1000 | 800 - 2000 |
Packed Columns
When ordering packed columns, use first three digit Cat. No. to indicate the type of column plus Cat. No. of sorbent used. Stainless steel and Titanium zirconium columns are available in 8 mm diameter 250 mm length. They are available packed with all HEMA-BIO size exclusion sorbents in 10 µm particle size.
Preparative stainless steel columns with 25 or 46
mm internal diameter are available on request.
The macroporous structure and hydrophilic nature of the TESSEK Separon HEMA-BIO matrix are ideal for forming powerful ion exchangers for chromatography of many types of solutes. The HEMA-S 1000 offers higher capacities while the HEMA-BIO 1000 matrix should be chosen for better protein recovery and enhanced rigidity, necessary for example in organic solvents.Four different ion-exchange modifications of TESSEK Separon HEMA sorbents are produced by TESSEK:
Ordering Information
The following sorbents are available in 10 and 100 g and bulk quantities.
|
|
| |
| HEMA-S 1000 DEAE | 1.2 - 1.5 | 161045 | - |
| HEMA 1000 DEAE | 1.2 - 1.5 | - | 170043 |
| HEMA 1000 Q | 0.8 - 1.2 | 160095 | 170093 |
| HEMA-S 1000 CM | 0.9 - 1.1 | 161055 | - |
| HEMA 1000 CM | 0.9 - 1.1 | - | 170053 |
| HEMA-S 1000 SB | 1.6 - 2.0 | 161135 | - |
| HEMA 1000 SB | 1.6 - 2.0 | - | 170133 |
| HEMA-BIO 1000 DEAE | 0.8 - 1.2 | 162045 | 172043 |
| HEMA-BIO 1000 Q | 0.4 - 0.8 | 162095 | 172093 |
| HEMA-BIO 1000 CM | 0.4 - 0.8 | 162055 | 172053 |
| HEMA-BIO 1000 SB | 1.2 - 1.6 | 162135 | 172133 |
Packed Columns
Stainless steel and Titanium Zirconium columns packed
with 10 µm HEMA-BIO ion exchangers are available in 8 mm
diameter and different lengths.
| Cat.No. | Steel column | Cat.No. | Ti-Zr column | |
| 101 | 8x250 mm | 201 | 8x250 mm | |
| 109 | 8x80 mm | 209 | 8x80 mm |
CGC columns 3.3 mm ID are available packed with 10
µm particle size.
| Catal.No. | CGC column |
| 901 | 3x150 mm |
| 909 | 3x70 mm |
| 911 | 3x30 mm |
Inert PEEK columns are available in following dimensions:
| Cat.No. | PEEK column |
| 302 | 7.5x300 mm |
| 351 | 4.6x50 mm |
| 304 | 7.5x75 mm |
| 347 | 4.6x150 mm |
| 301 | 7.5x250 mm |
| 341 | 4.6x250 mm |
Preparative columns with 25 or 46 mm internal diameter are available on request.
The sorbents in 60 µm particle size are available
also in HEMA-cart cartridges for solid phase extraction. Catal.No.
421.
The high density of hydroxyl groups on the internal surface provides TESSEK Separon HEMA with hydrophilic properties and excellent derivatization possibilities for the preparation of different chromatographic sorbents.
Common activation techniques, e.g., cyanogen bromide, triazine and others, can be used for ligand immobilization on TESSEK Separon HEMA materials. However, all those techniques are rather laborious and health hazards are usually involved. To overcome this step, TESSEK offers ready-for-use, activated affinity supports with a long shelf-life. Two different modifications with different levels of activation are available, the epoxy (E, E-L, E-H) and vinyl sulphone (VS, VSL).
Epoxy-activated supports react under suitable conditions with a variety of functional groups, especially with amino-, thio-, carboxy- and hydroxy groups.
Vinyl sulphone-activated supports exhibit higher reactivity, particularly for hydroxy groups.
The suitable level of activation depends on the type
of ligand. For the binding of low molecular weight ligands to
high surface density, a high level of activation should be used.
For macromolecular ligands, especially where biological activity
should be retained, a low activation level is necessary.
Ordering Information
The following sorbents are available in 10 and 100
g and bulk quantities. Due to the limited shelf time no HPLC columns
are available. They can be packed on request, but must be used
within three weeks from production. Plastic cartridges HEMA-cart
(Catal. No. 421) are packed with a dried sorbents.
| TESSEK Separon | Capacity mmol/g | Cat. No.
10 µm | Cat. No.
60 µm |
| HEMA 1000 E | 0.7 - 0.9 | 160015 | 170013 |
| HEMA-S1000 E-L | 0.1 - 0.3 | 161025 | - |
| HEMA 1000 E-L | 0.1 - 0.3 | - | 170025 |
| HEMA 1000 E-H | min 1.4 | 160085 | 170083 |
| HEMA-BIO 1000 E | 0.5 - 0.7 | 162015 | 172013 |
| HEMA-BIO 1000 E-L | 0.05 - 0.2 | 162025 | 172023 |
| HEMA-BIO 1000 E-H | 0.9 - 1.2 | 162085 | 172083 |
| HEMA-BIO 1000 VS | 0.015-0.025 | 162125 | 172123 |
| HEMA-BIO 1000 VS-L | 0.005-0.010 | 162115 | 172113 |
Most problems connected with the use of silica-based sorbents in reversed phase chromatography are associated with the presence of residual silanol groups and limited stability of the matrix under alkaline conditions.
Polymer based TESSEK Separon HEMA reversed phase sorbents were developed with the purpose of creating efficient and highly stable reversed phase material. The new sorbent is produced by grafting the macroporous hydrophilic TESSEK Separon HEMA 1000 co-polymer with C18 alkyl chains. The newly used HEMA-BIO matrix offers enhanced rigidity with essentially the same selectivity.
The sorbent is stable in the pH range 2-12 and is
compatible with all commonly used chromatography solvents. The
wide-pore structure of the TESSEK Separon HEMA-BIO 1000 C18 matrix
allows separations of macromolecules (peptides, proteins, polynucleotides,
etc.). The rigidity of the matrix ensures that the columns can
be operated with pressures up to 100 bar and that the efficiency
is not affected by changes in the mobile phase composition. The
lack of silanol groups and the outstanding stability of TESSEK
Separon HEMA-BIO 1000 C18 makes it especially suited for separations
of basic compounds and expand the application area for reversed
phase chromatography.
Ordering Information
The following sorbents are available in 10 and 100
g and bulk quantities.
| TESSEK Separon | Functional group | Catalogue No.
10 µm | Catalogue No.
60 µm |
| HEMA-S 1000 C18 | octadecyl | 161155 | - |
| HEMA 1000 C18 | octadecyl | - | 170153 |
| HEMA-S 1000 C8 | octyl | 161215 | - |
| HEMA 1000 C8 | octyl | - | 170213 |
| HEMA-S 1000 C4 | butyl | 161165 | - |
| HEMA 1000 C18 | butyl | - | 170163 |
| HEMA-BIO 1000 C18 | octadecyl | 162155 | 172153 |
Packed Columns
Stainless steel and Titanium Zirconium columns packed
with 10 µm HEMA-BIO C18 are available in 8 mm diameter and
different lengths.
| Cat. No. | Steel column | Cat. No. | Ti-Zr column | |
| 101 | 8x250 mm | 201 | 8x250 mm | |
| 109 | 8x80 mm | 209 | 8x80 mm |
CGC columns 3.3 mm ID are available packed with 10
µm particle size.
| Cat. No. | CGC column |
| 901 | 3x150 mm |
| 909 | 3x70 mm |
| 911 | 3x30 mm |
Inert PEEK columns are available in following dimensions:
| Cat. No. | PEEK column |
| 351 | 4.6x50 mm |
| 304 | 7.5x75 mm |
| 347 | 4.6x150 mm |
| 301 | 7.5x250 mm |
| 341 | 4.6x250 mm |
Preparative columns with 25 or 46 mm internal diameter are available on request.
The sorbents in 60 µm particle size are available also in HEMA-cart cartridges for solid phase extraction. Catal.No. 421.