Summer is finally here, and it is time to catch up on your to-be-read pile. I know you have been meaning to read (or read again) some basic info on chromatographicy sorbents, but there was just not enough time. Fear not and let me (not zombies, sorry...) summarize the most important info from our previous blog articles on the fundamentals of chromatography so you can relax while lounging by the pool.

Back to Basics: Silica Gel Physical Characteristics

To understand the key physical characteristics of sorbents used in chromatography, let's review what you already know from your previous readings.

When it comes to silica particles, shape and size matter. The choice of particle size depends on how difficult the separation is, and the pore diameter must be selected based on the analyte's molecular mass.

 

Selecting Pore Diameter

Sample Molecular Weight (MW in Dalton)
   
Organic Molecules   Peptides   Proteins (Polypeptides)
Bio-polymers
   
MW < 1,000 Dalton   1,000 < MW < 10,000 Dalton   MW > 10,000 Dalton
   
30 Å, 40 Å (< 400 Dalton)
60 Å, 70 Å (400 - 1,000 Dalton)
  90 Å, 100 Å
120, 150, 300 Å
  500 Å, 800 Å, 1,000 Å

 

Selecting Particle Size

Generated Pressure (GP) or Selectivity (α)
     
GP ~950 psi (65 bar)
α < 1.2 (very difficult separation)
  GP ~220 psi (15 bar)
1.2 < α < 1.5 (difficult separation)
  GP ~73 psi (5 bar)
1.5 < α < 2.0 (possible separation)
  GP ~29 psi (2 bar)
α > 2.0 (easy separation)
     
From 0 to 30 μm   From 15 to 40 μm   From 40 to 100 μm   From 40 to 1,000 μm

 

As mentioned, it's not just about the size, shape matters too: irregular particles lead to an irregular flow as observed in the image below. Regardless of the particle shape, a large particle size distribution leads to pockets of similar sized particles getting together to form a preferential path that the analytes will follow.

Other than shape and size, metal content, water content and pH are also important characteristics. Water content and pH can be corrected in your lab, and SiliCycle's gel has the lowest metal content compared to other manufacturers.

Typical metal content comparison for 40 - 63 µm, 60 Å Silica Gels (mg/kg)
MetalsSiliCycleManufacturer AManufacturer B
AluminumAl33262280
BariumBa9.459.732.5
CalciumCa3361,150502
ChromiumCr0.50.60.4
CopperCu0.20.20.2
IronFe327541
LeadPb0.415.30.95
MagnesiumMg61149104
NickelNi0.40.50.5
SilverAg0.090.290.19
SodiumNa466945585
TinSn0.20.20.1
TitaniumTi147250179
ZirconiumZr327556

To adjust water content, The Brockmann & Schodder Activity Test (Appn_SF002-0) can be used: dry your silica gel by placing the product for 1 or 2 hours on a rotary evaporator and heat at 70-80°C. If the product is not dry enough after this procedure, you could finish drying it overnight under vacuum at room temperature (do not heat it overnight).

For pH sensitive analytes it goes without saying that pH is more critical. It can be adjusted directly on your column by adding a pH modifier to your eluent. Triethylamine is one of the most popular options to increase the pH of bare silica and its low boiling point makes it easier to remove from the product after the purification.

Using alumina or SiliaBond Cyano are also alternative solutions to adding basic modifiers.

Sorbent Selection: Separation and Chemistries

All these basic characteristics must be considered when choosing the stationary phase. This previous article detailed a roadmap for chromatographers to follow in their quest in finding the right chromatographic phase based on their applications. Once the particle size and shape and the pore diameter are selected, the next steps are to choose the separation mode and the associated chemistry.

The types of analytes that need to be purified determine which separation mode is the best suited and which chemistries are available for that separation mode.

First, you need to find the separation mode which depends mostly on the analyte's polarity. Then, with the separation mode in mind, different sorbents have different selectivity. Bare silica is not the only option for normal phase chromatography. Amine, Diol and Cyano functionalized silicas are less polar than bare silica and have other interactions available. For reversed-phase, the use of silica functionalized with alkyl chains and cyclic or aromatic functions is predominant. The longer the chain, the more hydrophobic the stationary phase is, and aromatic functions offer alternative selectivity with other types of interactions. With charged analytes, both ion exchange (IEX) and HILIC could be separation modes to try. With IEX, the analyte's pKa dictates the sorbent to use.

This pathway may be an over-simplification, but it is a good starting point. For more information on the sorbent options, check out our Working With series of Application Notes on most of these sorbents.

High Quality Silica Gel

Finally, the most important thing to save time with your chromatography, is to make sure that the products used are of high quality with batch-to-batch consistency. This is what the Certificate of Analysis is for: to confirm that these characteristics are kept constant. The experiments certifying this consistency were detailed in this article on the secrets of the Certificate of Analysis.

Conclusion

We managed to avoid a zombie apocalypse with this summary, and now, you can go back to your light readings from your to-be-read-pile. My next light book to read is the Handbook of Chemistry and Physics, 102nd Ed. Happy vacations!

More for your to-be-read pile (aka Learn more):

 

Eliane, M.Sc

Eliane is the Scientific Content Specialist here at SiliCycle, with years of experience both in the lab and client support. She studied at Laval University, for both her Bachelor’s and Master’s in Chemistry. In fact, her thesis was in organic electronics in which purity is of the utmost essence, making her in-tune to the purification needs of chemists.