Manual mobile phase preparation

Mixing errors manual mobile phase preparation. These tend to occur as a result of operator error during the dispensing and mixing of the individual components. The final concentration of each component within the mixture can vary depending on the way in which it is prepared. 

Mobile phase. Many of the problems associated with the mobile phase will depend on the HPLC instrument configuration and, as a consequence, whether the mobile phase preparation is manual or automated. 

The routine work with hybrid micellar mobile phases requires keeping some cautions usually not described in scientific reports or specialized manuals. The experimental procedure comprises two main stages the mobile phase preparation and the column conditioning. "... 

A very helpful tool for manual application can be the employment of layers with a concentrating zone. The so-called concentrating or preadsorbent zone is a small part of the plate that is covered with an inert but highly porous adsorbent such as diatomaceous earth. Various precoated preparative layers with a preadsorbent zone are commercially available. The effect of the concentrating zone is depicted elsewhere in detail (see Chapter 3, Figure 3.4). In brief, the preadsorbent zone serves as a platform for manual application of any desired performance quality. When development starts, soluble components migrate with the mobile phase front and are... 

Achiral-chiral multidimensional chromatography remains one of the best ways to separate chiral analytes from interfering matrix components or other compounds. The flexibility offered by different operation modes, stationary and mobile phases, and configurations allows analysis methods to be tailored to the analytical problem. By offering possible configurations for both online sample cleanup and concentration, achiral/chiral LC/LC reduces manual sample preparation. The ability to be coupled to... 

Mobile phase storage The figure shows four reservoirs, one for each of up to four pure solvents [e.g., water (perhaps with pH controUing buffer), acetonitrile, methanol, tetrahydrofuran, etc.]. Note the use of inlet filters. Alternatively one could prepare the mobile phase mixture to the desired composition manually, and store it in a single reservoir. Operation at a single, constant mobile phase composition is called isocratic HPLC elution. 

In this case, the mobile phase can be prepared manually (premixed) or it can be mixed automatically using the HPLC solvent delivery system. 

Manual Mixing When the mobile phase is to be prepared manually, the organic portion and the aqueous portion should be independently measured out using measuring cylinders or by weight where appropriate. Both phases should then be added together in a separate HPLC solvent bottle and... 

Mobile phase proportioning Prepare the mobile phase manually... 

Methanol has the disadvantage of producing a relatively highly viscous mixture with water (see above), giving rise to much higher pressures than other mobile phases. If methanol-water mixtures are prepared manually then both components must be weighed or each one volumetrically determined separately. The considerable volume contraction that occurs produces a solution with a methanol content in excess of 50% by volume from an initial 500 ml of water topped up to 1000 ml with methanol. For the same reason the retention times... 

Figure 5 Fast SFC analysis of the S-enantiomer of clevidipine after hydrolysis into its corresponding acid. Column and conditions Chiralpak AD 50 x 4.6 mm ID at 30°C, mobile phase carbon dioxide with 28% of 2-propanol, flow rate 4.0 ml min backpressure 150 bar, UV detection at 240 nm. Sample preparation and work-up 2 mg of S-clevidipine substance was dissolved in 0.5ml of methanol followed by 50 pi of 1 mol r sodium hydroxide. After 10 min, 1 ml of water and 50 pi of sulfuric acid, 1 mol l was added and 0.5ml of dichloromethane as extraction medium. After brief vortexing and centrifugation 5 pi of the lower organic phase was loaded manually and injected. Upper trace sample, lower trace sample with 0.1% of S-acid added. (Reproduced with permission from Gyllenhaal O (2001) Fast enantioselective separation of clevidipine and a dihydropyridine substituted acid by SFC on Chiral pak AD. Fresenius Journal of Analytical Chemistry 369-. 54-56 Springer.)...

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