28.02.2013 Views

Handbook of Size Exclusion Chromatography and Related ...

Handbook of Size Exclusion Chromatography and Related ...

Handbook of Size Exclusion Chromatography and Related ...

SHOW MORE
SHOW LESS

Create successful ePaper yourself

Turn your PDF publications into a flip-book with our unique Google optimized e-Paper software.

Table 2 Characteristics <strong>of</strong> SEC Packing Materials<br />

Attribute Variable Relationship Typical range<br />

Specific resolution Particle size 1=s Rsp ¼ 1–5 a<br />

Pore size/pore volume 1=D2 Porosity 55–80% b<br />

Efficiency, HETP Particle size vdp 2 5–20mm<br />

Linear velocity vkvl 0.4–1.0mm/min<br />

Column pressure Particle size Constant/dp2 5–10MPa<br />

Particle shape Form factor Q ;1 for spherical, ’2 for irregular<br />

Mechanical stability Support type Inorganic supports are in general more rigid; for all supports, the larger the pore<br />

size (<strong>and</strong> pore volume), the weaker the particle; at constant pore size <strong>and</strong> pore<br />

volume, particle strength decreases with size.<br />

Chemical stability Support/bonded phase Silica slowly dissolved above pH7; enhanced stability possible from surface<br />

treatment or bonding reaction(s); most polymer-based matrices are stable up to<br />

pH10 or higher, allowing high-pH column regeneration in biopurification <strong>and</strong><br />

wider access to buffers, detergents, <strong>and</strong> chaotropic salts.<br />

Thermal stability Support/bonded phase Silica columns have few temperature limitations; when using polymer columns at<br />

1408C, do not cool to ambient between high-temperature analyses to avoid<br />

resettling <strong>of</strong> the packet bed; most modern SEC packings can be sterilized.<br />

Recovery Mass Water-soluble biopolymers, synthetic polymers, <strong>and</strong> polyelectrolytes may adsorb<br />

on polymer- <strong>and</strong> silica-based columns depending on mobile-phase conditions.<br />

Activity Maintenance <strong>of</strong> biological activity (<strong>and</strong> mass recovery) for proteins depends on<br />

mobile-phase conditions, column type, <strong>and</strong> contact time.<br />

a According to Rsp ¼ 0:58=sD2, specific resolution is inversely proportional to the product <strong>of</strong> the peak st<strong>and</strong>ard deviation s <strong>and</strong> the slope <strong>of</strong> the calibration curve D2.<br />

See page 103 <strong>of</strong> Ref. 8 for details.<br />

b Pore size <strong>of</strong> commercial materials varies from very small to very large, depending on the application. For each pore size, the requirement for a large pore volume<br />

is balanced against the need for a pressure-stable particle. In a study <strong>of</strong> commercial silica-based SEC packings, the percentage <strong>of</strong> pore volume per particle varied<br />

from 55 to 80% (Ref. 33).<br />

© 2004 by Marcel Dekker, Inc.

Hooray! Your file is uploaded and ready to be published.

Saved successfully!

Ooh no, something went wrong!