Automated fraction-collectors

However, for quantities substantially less than this level, 7- to 10-mm i.d. analytical columns can often be used in a semipreparative mode. By repeatedly injecting 300 to 500 ju,l of up to 1% polymer, reasonable quantities of polymer can be isolated. An autosampler and automated fraction collector can be setup to perform such injections around the clock. Although the larger injections and higher concentrations will lead to a loss of resolution, in some situations the result is quite acceptable, with a considerable savings in time being realized over other means of trying to make the same fractionation. 

Acetate buffer (20 mM - pH 5.0)equilibrated with the same buffer. The solution was passed through the column at a flow rate of 1 mL.mim with acetate buffer (20 mM -pH 5.0), followed by a linear gradient from 0-1M NaCI in the acetate buffer. The eluted fractions were collected in an automated fraction collector (Pharmacia Biotech) and the absorbance of the fractions was measured at 280 nm. The major peak fractions werethen assayed fortannaseactivity, and onlythefractions possessing tannase activity were pooled. 

The mobile phase can be allowed to flow under the force of gravity, a low pressure pump can be used, or compressed gas can be used to pressurize a solvent reservoir. The linear velocity should be about one-third of that used in analytical columns. The sample can be applied to the top of the column with a microsyringe or pipet using the stop-flow method, or an inexpensive, low-pressure valve can be used. The eluent is usually collected in separate tubes using an automated fraction collector. Inexpensive UV detectors with large solvent volumes are available, or flow cells can be fitted to conventional UV/visible instruments. 

Low pressure LC is very similar to the experiment performed by Tswett. A variety of stationary phases can be used and relatively large particle sizes are needed to keep the pressure drop low and facilitate packing. The ideal mobile phase should be inexpensive and volatile to facilitate sample recovery. The eluent is collected in separate tubes using an automated fraction collector. Since the samples often contain a lot of impurities, the column is very likely to be contaminated. Cleaning can be achieved by rinsing with solvents or... 

The methods employed for the miscible displacement studies were similar to those used in previous experiments (Hu and Brusseau, 1998). We connected a high-performance liquid chromatography (HPLC) pump (Model 301 from Alltech Associates Inc., Deerfield, IL) to the column, and placed a three-way valve in-line to facilitate switching between treatment solutions. Several iodine species (iodide, iodate, and 4-iodoaniline) were used to study transport behavior. We also examined the transport of tritium and bromide, commonly used conservative tracers, so that we could compare their transport behavior with iodine species. For transport experiments of 4-iodoaniline, which is used as a representative refractory organic iodine species, the solution was allowed contact only with glass or stainless steel, to avoid potential interaction of organoiodine with plastics in the column system. Column effluents were collected with an automated fraction collector (Retriever 500, ISCO Inc., Lincoln, NE) for chemical analysis, as described below. 

Techniques which utilize continuously perfused, "dynamic diffusion cells maintain "sink" conditions, and serial samples may be collected in automated fraction collectors. When these samples are added, the cumulative amount removed ("excreted") from the diffusion cell as a function of time is obtained. This Is not Identical to cumulative absorption, which is the sum of the amount excreted (EXC) plus the residual amount (RA) still remaining In the diffusion cell. Thus, the continuous perfusion technique is limited by the ability of the resulting cumulative excretion curve to approximate the true cumulative percutaneous absorption curve. Two methods of approximation may be employed. Both depend on an understanding of the kinetic parameters which define the diffusion cell system being utilized. 

Fig. 1 VersaFlash high throughput flash purification (HTTP) system shown with two stacked prepacked 40 x 75 mm (51 g) cartridges inserted in the support stand and 110 x 300 mm (1.35 kg), 80 X 150 mm (410 g), and 23 X 110 mm (23 g) cartridges on the right side of the eluent pump. The 3-way valve injector at the upper right allows direct application of sample onto cartridges using a syringe. The outlet tubing can be used for manual sample collection or connection to an automated fraction collector or UV detector.

A third RPC instrument, the CLC-5 (Hitachi), was described by Nyiredy (1996) for use in PLC. It consists of a rotating disk column comprising two removable disks of 30 cm diameter, UV detector, and automated fraction collector. Its speed can be continuously varied from 0 to 1000 rpm. 

The collection of eluent fractions from a conventional preparative HPLC is a trivial matter. Provision of automated fraction collectors suitable for HPLC is no commonplace and instrumentation is highly developed and widely commercially available. However, collection of eluent from a preparative SFC is not trivial, particularly if complete collection of a fairly volatile solute is required. In a supercritical fluid chromatograph using carbon dioxide the expansion of the fluid at depressurisation is very large indeed. If carbon dioxide is depressurised from 300 Bar (4500 psi) to atmospheric pressure is expands to 355 times its original volume if its temperature is maintained constant. Hence at a flow rate of lOcm min" of supercritical carbon dioxide at 300 Bar, which corresponds to about 7 g carbon dioxide per minute, expansion after depressurisation yields 3.5 L of carbon dioxide gas per minute at NTP. 

The authors applied this concept to both gas/liquid (see Figure 3.75) and liquid/ liquid systems (see Figure 3.76). This set-up consisted in the core of a tubular reactor with an interdigital micro mixer as dispersion unit (compare Figure 3.77). The peripheral equipment consisted of an automated pipetting robot, a fraction collector and a gas-chromatograph equipped with an automatic injector. 

Automation of Prep-HPLC. Reverse phase prep-HPLC separation has proven to be a very reproducible technique. For this reason, the process can be automated. In addition to the standard components of an HPLC system, the following are required for automation an autoinjector capable of large volume injections (2.0 to 5.0 ml), a programmable controller (a microprocessor controller), a fraction collector, and a waste valve (3-way valve) controlled by a solenoid (Fig. A). The microprocessor should allow programming of the solvent flow rate, the sequence for solvent gradient formation, the time of injection, advancement of the fraction collector (to collect specific fractions rather than just uniformly incremented advancement), and control of a waste valve. 

Automatic analysis consists essentially of the same steps as the corresponding manual method (p. 4). In some cases this may be simple, the requirements amounting to a mechanical device for presenting the sample to the detector, a timer to control the time of measurement and a data recorder. However, if sample pretreatment and separations are necessary a variety of wet chemical stages needs to be automated. Such automated steps may be included in what remains essentially as an operator procedure. For example, an automatic pipette may be used for sampling a fraction collector in a chromatographic separation or an automatic sequence for making a measurement and... 

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