Waters’ Sep-Pak

Whatever the technique employed to purify the RNA, it is necessary to desalt and concentrate it prior to use in crystallization trials. Avery efficient way of achieving this is to use reverse-phase Sep-Pak columns that can be used on the bench (Waters Sep-Pak Cl8 Classic short-body). These are operated by gravity or using a syringe (Protocol 14.3). 

Some samples lend themselves to small column cleanup such as that provided by Waters Sep-Pak (a miniature chromatographic column). And several suppliers offer processing turntables which will perform various cleanup operations which are frequently required prior to chromatography. And when samples have been cleaned up (whether manually or by one of these automated devices), they can then be transferred to an HPLC unit by any one of several available auto-injectors. 

Because of the favorable sorptive properties of the reversed-phase supports, batch adsorption and desorption can be a very effective way to desalt a chromatographed sample or to partially fractionate a peptide mixture during a purification procedure. For example, 1-2 gm of an oc-tadecyl silica packed into a silanized glass or plastic pipette can be used for the batch fractionation of small amounts of a crude peptide extract from tissues, such as the pancreas or pituitary, or from a synthetic experiment. A number of commercial products, such as the Waters Sep-Pak, have found use in this manner 10) as a purification or sample preparation aid. Protocols for batch extraction procedures on alkyl silicas have been discussed 17a,b) and applied to neuropeptides 10, 158, 166) and other hormonal peptides 88, 162, 167, 168). With these methods recoveries of peptides present in a tissue extract are generally higher than those found with classical fractionation techniques due in part to the fact that proteolytic degradation is minimized. 

Millipore Corporation. 1992. Waters Sep-Pak DNPH-Silica Cartridge. Care and use manual. Milford, MA Waters Chromatography Publications. 

Waters Sep-Pak Cartridge Application Bibliography, Millipore Corporation, Bedford, MA 01730, USA 1986... 

The relative PLMA MW was determined by using nonaqueous SEC with THF as solvent and an HP1037A refractive index detector from Hewlett-Packard. A solid-phase extraction cartridge (Waters Sep-Pak) was used to take the PLMA and surfactants from the water phase to THF and to adjust the concentration for SEC. Sodium sulfate was used to dry the THF extract before injection into the SEC columns. For some samples there was too much surfactant present, and it overloaded the cartridge in such cases ion-exchange beads were used to reduce the concentration of the ionic surfactants in the aqueous phase before extraction. The advantage of this extraction procedure is its simplicity. The overall separation scheme is shown in Scheme I. 

General Techniques. Cleanup of analytical samples can be done conveniently with Waters Sep-Paks or equivalents available from Extrelut or Merck, and they can be regenerated. The major problem in isolation, characterization, and analysis of pesticide conjugates is ensuring 1005 cleavage of many conjugates. 

This experiment is carried out using a Waters Sep-Pak Classic CIS solid-phase extraction cartridge. A kit containing two S Pak Classic CIS cartridges, one envelope of Kool-Aid-brand unsweetened soft drink mix, and a detailed instruction sheet can be ordered from Waters Corporation under the part number WAT0SS2S3. In addition to the kit, you need a 10-mL syringe (with gradation and without needle), a container with a capacity of 2 L, two 2ft-mL beakers, at least four test tubes, and one bottle with 70% isopropanol and about 3 L water. 

A Waters Sep-Pak Cig extraction cartridge was pre-wet with 2 ml of methanol and 5 ml of water. For the timolol extraction, 1 ml of human serum (or aqueous humor) was passed through the cartridge, the drugs being adsorbed on the Sep-Pak matrix. The matrix was then rinsed with 6 ml of Britton-Robinson buffer (pH == 1.8). For elution of timolol 3 ml of acetone was used and the first ml discarded. The eluent was dried under a stream of nitrogen. The residue was diluted to 20 ml with Britton-Robinson buffer (pH = 4.0). The voltammograms were recorded under the optimum instrumental conditions. 

From Frankel et al. (1989), using a Waters Sep-Pak silica cartridge. 

Orotic acid and orotidine by an ion-pair method with a Beckman ion-pair kit. Urine samples were precleaned on Waters Sep-pak Cl 8 cartridges. 

These and other sorbents are supplied in prepacked columns. Analy-tichem (Varian Sample Preparation Products) Bond Elut, International Sorbent Technology (Jones Chromatography) Isolute and Waters Sep-Pak are all well-known names. The sorbents are also available in bulk, and a wide range of accessories is available, such as Luer stop-cocks, vacuum pumps, manifolds and regulators. The sorbents supplied by Varian and International Sorbent Technology appear to be identical, but... 

Waters, Sep-Pak Cartridge Applications Bibliography, 4th ed., Millipore Corp., 1986. 

Waters Sep-Pak Cis cartridge or, for use online with column switching, various Cjj and C s packings in 4.6 X 10 mm columns... 

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