Pre-packed columns

A fourth factor is the flow rate of the eluent (mobile phase). A too high rate decreases resolution because there is no time for molecules to diffuse into the pores of the matrix. In contrast, a very slow migration of solvent decreases the resolution by remixing the components by diffusion. The effect of diffusion is minimized if the chromatography is done at low temperature. If short separation times are necessary, pre-packed columns, elevated pressure, or HPLC columns are indispensable. 

Many packing materials are available only in pre-packed columns and, in this respect, packing can be avoided altogether. However, the cost of pre-packed columns is extremely high, and this is one of the most undesirable features of HPLC at present. 

For some very non-polar compounds, even small quantities of dichloromethane may cause elution problems and an alternative method of loading can be used. Thus, a solution of the sample is added to a small amount of silica in a round bottom flask and the mixture is evaporated to dryness. The dry impregnated silica is then added to the top of the pre-packed column. 

Use pre-packed columns of Sephadex G-25 gel filtration media, for rapid sample clean-up at laboratory scale, as shown in Table 2. 

Pre-packed column Sample volume loading per run Sample volume recovery per run Code No. 

Table 5. Pre-packed columns for rapid desalting and buffer exchange.

Particle size 90 pM. Available in pre-packed columns and as bulk media. 

If a purification is not intended for scale up (i.e. milligram quantities of product are needed), use high performance media such as Sepharose High Performance (IEX, HIC) or MonoBeads (IEX), or SOURCE (IEX, HIC). All these media are available in pre-packed columns. 

Although general standard protocols were supplied with this pre-packed columns, it was decided to further optimise the binding and elution conditions for the specific target molecule. 

To increase speed and efficiency in method development, use small pre-packed columns for media scouting and method optimisation. HiTrap IEX Test Kit is ideal for this type of work, as shown in Figure 34. 

Using pre-packed columns at any scale will ensure reproducible results and high performance. 

Procedures vary according to type of sample and medium. Guidelines are supplied with the medium or pre-packed column. 

Pre-packed columns ensure reproducible results and highest performance. 

A study of column extraction of metals was carried out using pre-packed columns available from Polymer Laboratories (now a part of Varian, Inc.) [6]. Trace metals in these monomers would have a detrimental effect on the stability of anaerobic adhesive products and must be absent prior to use. The columns used are specially coated macroporous polystyrene products that are compatible with polar, non-polar, protic and aprotic solvents. They are designed to remove metals from solvents and monomers. The metal removing SPE product is approximately 45 pm and based on a mono-dispersed macroporous polymeric material. 

Sephadex LH-20 (Pharmacia) is a support for gel chromatography and is widely used in isolation protocols for proanthocyanidins. It is composed of dextran molecules which are three-dimensionally cross-linked with hydroxypropyl groups [205]. Proanthocyanidins are separated by filtration (molecule sieve effect of the gel) and adsorption (hydrogen bonding). Retention increases primarely with increasing ( ) molecular weight and C4—>C6 linked procyanidins are more retarded than their C4—>C8 linked counterparts [206-207]. Sephadex LH-20 has first been used for sample clean up in the analysis of apple ciders by Lea and Timberlake [208]. So far, no pre-packed columns exist on the market. All methods described in the literature work with different column dimensions (see Tab. (6)). Standardization would help to make results more comparable. Some validation procedures have been performed for two protocols [185,209],... 

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