Bead Cellulose - General Information
Perloza MT - Gel Filtration Media for Biomolecule Separation
Macroporous bead cellulose is a hydrophilic matrix, which is distributed under the trade mark PERLOZA®. Bead cellulose is produced in several porosity types, marked MT 50, MT 100, MT 200 and MT 500, and various particle size. Perloza MT is a highly porous regenerated cellulose with heterogenous matrix (partially microcrystalline) and with polymer structure stabilized by hydrogen bonds only - there are no covalent cross-linkings. Detailed features of macroporous bead cellulose are described in the following references (Peska J., Stamberg J., Hradil J.: Angew. Makromol. Chem., 53 (1976) 73; Baldrian J., Peska J., Stamberg J., Ludwig J., Paul D.: "Chromatography of Synthetic and Biological Polymers", Ed. R. Epton, band 1, p. 109, E. Horwood Ltd., Chichester 1978).
One of working sites, which use to deal with a research of bead cellulose applications, was: Department of Glycobiotechnology, Institute of Chemistry, Slovak Academy of Science, Bratislava, Slovakia.
Features
- Excellent mechanical stability even at gel with large pores. Easy to handle. Resistant to destruction and generation of fine particles during stirring.
- Rigid spherical particles. High flow throughput the gels at a low drop in pressure.
- Narrow particle size distribution in several degrees converting all ranges common for this sort of gel.
- High chemical resistance and compatibility with most commonly used solvents and buffers. Applicability in wide range of pH and salt concentrations with minimal changes (swelling / shrinking) in the gel bed.
- High temperature stability allows sterilization by autoclaving.
- High hydrophility. A number of free hydroxy groups in matrix allows preparation of derivatives or enzyme immobilization.
- High porosity of matrix allows good capacity and recovery.
- High selectivity of separation.
- Easy-to-ready. Gels are supplied pre-swollen.
Mechanical Stability
Flow characteristics of Perloza types with various particle sizes
| PERLOZA type | Particle Size [ um ] |
Denotation | Flow Performance (1) [ ml cm-2 hod-1 bar-1 m-1] |
| MT 50 | 30 - 50 | - | 85 |
| 50 - 100 | Fine | 270 | |
| 100 - 250 | Medium | > 500 | |
| MT 100 | 30 - 50 | - | 20 |
| 50 - 80 | Superfine | 170 | |
| 80 - 100 | Fine | 300 | |
| 100 - 250 | Medium | 600 | |
| MT 200 | 50 - 80 | Superfine | 130 |
| 80 - 100 | Fine | 220 | |
| 100 - 250 | Medium | 450 | |
| MT 500 | 50 - 80 | Superfine | 45 |
| 80 - 100 | Fine | 80 | |
| 100 - 250 | Medium | 250 |
(1) For distilled water at 20 oC.
Optimum*) pressure in column during packing with PERLOZA MT
| PERLOZA type | Pressure [ bar ] |
| MT 50 | > 2.0 |
| MT 100 | 1.5 - 2.0 |
| MT 200 | 1.2 - 1.5 |
| MT 500 | 0.8 - 1.2 |
*) pressure at which maximum of efficiently is reached
Maximum*) flow and pressure throughput for PERLOZA MT
| PERLOZA type | Flow Rate | Pressure [ bar ] | |
| linear [ cm / hr ] | volumetric**) [ ml/min ] | ||
| MT 50 | > 760 | > 9.9 | > 4.0 |
| MT 100 | 750 | 9.0 | 2.2 - 2.7 |
| MT 200 | 450 | 6.0 | 1.5 - 2.0 |
| MT 500 | 115 | 1.5 | 1.0 - 1.5 |
*) maximum of flow rate or pressure at which column is stabill not
blocked
**) for bead size range 50-80 µm and gel-bed 1 x 60 cm.
By their pressure stability Perloza® belongs to the category of swollen semi-rigid gels for low-pressure chromatography. The gels are characterized by excellent mechanical properties.
Flow characteristics of Perloza MT types Gel bed 1 x 60 cm, particle size 50 - 100 µm for all types
Column: 10 x 660 mm, Eluent: destilled water, Packing Flow Rate: 760 cm/hr
Column: 10 x 660 mm, Eluent: destilled water, Packing Pressure: 2.5 bar
Column: 10 x 660 mm, Eluent: destilled water, Packing Pressure: 1.75 bar
Column: 10 x 660 mm, Eluent: destilled water, Packing Pressure: 1.25 bar
Column: 10 x 660 mm, Eluent: buffer phosphate 0.2 M pH = 7, Packing Pressure: 1 bar
Agarose = agarose gel, Japan = Japan Cellulose 2000
Column: 10 x 660 mm, Eluent: destilled water
Japan = Japan Cellulose 90, 300, 1000, Dextran = dextran gel 100
Information on the porosity of Perloza materials in water-swollen state as obtained
by inverse steric exclusion chromatography (ISEC)
|
Net
volume of water-swollen beads* [ cm3/g ] |
Medium
pore diameter [ nm ] |
|
| Perloza MT 50 | 1.65 | 7.4 |
| Perloza MT 100 | 4.86 | 27 |
| Perloza MT 200 | 6.71 | 32 |
| Perloza MT 300 | 7.32 | 39 |
| Perloza MT 500 | 7.43 | 38 |
*Does
not include the interstitial spaces
The resutls in this table were measured by:
Karel Jerabek, Ph.D.
Institute of Chemical Process Fundamentals
Rozvojova 135
165 02 Prague 6
Czech Republic
Size Exclusion Limits for Properties (Porosity) of Individual MT types
Perloza MT types are highly porous gels well accessible for large macromolecules, as it is shown in this Figure.
Selectivity curves of Perloza MT types Kav vs. Molecular Weight of globular proteins in semilogarithmic scale
Elution behavior of globular protein on Perloza MT, dependence of distribution coefficient Kav on Molecular Weight
| Protein | M. W. | Kav | |||
| [ g / mol ] | MT 50 | MT 100 | MT 200 | MT 500 | |
| Blue Dextran | 2 000 000 | 0 | 0 | 0.01 | 0.02 |
| IgM | 1 000 000 | 0 | 0.10 | 0.22 | 0.36 |
| Thyroglobulin | 660 000 | 0.01 | 0.20 | 0.40 | 0.50 |
| Ferritin *) | 440 000 | 0.01 | 0.35 | 0.57 | 0.65 |
| Catalase *) | 238 000 | 0.01 | 0.47 | 0.74 | 0.78 |
| Aldolase | 158 000 | 0.02 | 0.55 | 0.78 | 0.81 |
| GLS | 135 000 | 0.05 | 0.58 | 0.80 | 0.82 |
| Bovine Serum Albumin | 66 000 | 0.19 | 0.70 | 0.88 | 0.87 |
| Ovalbumin | 44 000 | 0.30 | 0.75 | 0.88 | 0.88 |
| Chymotrypsinogen A | 25 000 | 0.44 | 0.81 | 0.89 | 0.90 |
| Myoglobulin | 18 000 | 0.50 | 0.83 | 0.90 | 0.91 |
| Ribonuclease | 13 500 | 0.55 | 0.85 | 0.90 | 0.92 |
| Cytochrome C | 12 400 | 0.57 | 0.86 | 0.91 | 0.92 |
| Bacitracin | 1 420 | 0.71 | 0.89 | 0.92 | 0.95 |
| L-Histidin | 152 | 0.72 | 0.90 | 0.93 | 0.96 |
| Alanine | 89 | 0.73 | 0.91 | 0.94 | 0.97 |
eluent: buffer 0.2 M phosphate pH 7.0 *) buffer 0.013 M phosphate pH 7.0
It is necessary to mention that the pore volume accessible to a given molecule is determined by its size (weight) and the pore size distribution of packing and can be defined by the following formula:
where Kav is the distribution coefficient (the factor of pore volume available for a given molecule), Ve is elution (retention) volume, Vo is void (interstitial) volume and Vt is total column volume.
Selectivity parameters of PERLOZA MT
| PERLOZA type | Exclusion Limit | Fractionation Range |
| MW proteins, g / mol | ||
| MT 50 | 1 x 105 | 20 000 - 70 000 |
| MT 100 | 2 x 106 | 30 000 - 1 000 000 |
| MT 200 | 4 x 106 | 200 000 - 2 000 000 |
| MT 500 | > 5 x 106 | 300 000 - 4 000 000 |
| PERLOZA type | Bead Volume [ ml /g dry ] | Exclusion Limit [MW dextrans ] |
| MT 50 | 3 - 5 | 50 000 |
| MT 100 | 9 - 11 | 100 000 |
| MT 200 | 13 - 16 | 200 000 |
| MT 500 | 18 - 21 | 500 000 |
Other Technical Data
Temperature Resistance (2) oC : 120
Stability within pH Range : 1 - 14 NaOH or HCl (c = 1 mol / l)
Stability in Salt Solutions : with ionic strength up to 10 mol/l
Swelling / Shrinkage (vol %) :
with changes in pH, salt concentration (3)
1 max.
with replacing of aqueous solutions by polar organic solvents
10 max.
Chemical Resistance : aqueous
solutions, buffer, organics, detergents
and
chaotropic agents (including SDS, 8 M urea)
(2) In wet state, pH 7.0 for 1 hour.
(3) Over the whole range of pH anionic strength.
Handling with Bead Cellulose and Sorbents Iontosorb
Bead Cellulose and its derivatives must not be dried, because drying results in loss of porosity. Therefore they are supplied fully swollen in water, sucked off. They can be stabilized by sodium azide (0.02 %) or ethanol (20 %) according to customer´s demand.