Gradient rapid separations

To ensure operation under reproducible conditions, the column is enclosed in a thermostatically controlled oven whose temperature can be held constant to within 0.1°C. Operating temperatures range from ambient to over 400°C and may remain constant during a separation - isothermal operation - or automatically increased at a predetermined rate to speed the elution process - temperature programming (p. 106). The latter is a form of gradient elution. Rapid temperature equili-... 

Similar considerations apply to very rapid gradient separations. We have shown above that a 1-min gradient with a 2 cm 2- j,m column can... 

FIGURE 9 Moderately rapid gradient separation. Column XTerra MS C, IS, 4.6x 20mm 3.5p.m. Gradient 0 to 100% B over 4min,A 0.1% formic acid in water, B 0.1% formic acid in acetonitrile. Flow rate 3.0mL/min. Temperature 30°C. Detection UV at 254 nm. Instrument Alliance 2795 with 996 photodiode array detector. Compounds (I) acetanilide, (2) triamcinolone, (3) hydrocortisone, (4) 2-amino-7-chloro-5-oxo-5H-[l]benzopyrano[2,3-b]pyridine-3-carbonitrile, (5) 6a-methyl-17a-hydroxyprogesterone, (6) 3-aminofluoranthene, (7) 2-bromofluorene, (8) perylene, (9) naphtho(2,3-a)pyrene. 

An example of a high-speed application is shown in Figure 11. Here a rapid gradient separation of a few acids and esters is shown. A rapid gradient is executed at a high flow rate, 4mL/min, on a 100mm x 4.6mm Chromolith column. Seven compounds are separated in less than 3 min. 

There are circumstances, however, when increasing the column length increases its resolution. This occurs when the column is washed continuously with a solution of one concentration rather than a gradient of rapidly increasing concentrations. In this case the salt concentration of the eluting solution is chosen such that it binds to the column with about the same tenacity as the species being separated. When the desired... 

In summary, the use of a polar capillary column and an appropriate gradient of supercritical CO2 density at a temperature of about 150°C permits the flavonoids to be separated rapidly and effectively. 

Self-diffusion coefficients of polyvalent cations in these perfluorinated ionomer membranes have not been reported. It can be inferred from the use of the sulfonate membranes as Donnan dialysis devices that transport of cations such as CuflT), Mg(II), and Al(III) under a concentration gradient is rapid. Also, column chromatographic separation of the alkaline-earth ion is readily accomplished with a powdered Nafion perfluorosulfonate polymer, which is again an indication of facile diffusion of these cations within the polymer phase. 

TrifluOToacetate TFA is a common, volatile buffer additive ftn the separation of proteins and peptides. It decomposes rapidly and should not be stor longer than neeessary. Fresh buffer should be prepared daily. TFA gives rise to ghost peaks in gradient separations. Tris-(hydroxymethyl)aminomethane. 

HPLC more rapidly separates PO copolymers or blends than conventional TREF or CRYSTAF. The new PL XT-220 rapid screening HPLC system enables one to perform isocratic and gradient separations at T = 30-220 °C with rapid switching between up to six different columns, each with different solvents (Pasch et al. 2009). 

In Chapter 1.2 of this book, we discussed the issue of fast gradient separations and showed that one can accomplish high-quality rapid separations with short columns packed with small particles. How fast one can go at a stiU reasonable performance is limited by the available pressure. A similar situation arises if one wants to accomplish a truly high-powered separation within a reasonable time frame. Once again, the available pressure is the limiting factor. Thus, itis absolutely clear that better separations can be accomplished if more pressure can be applied to drive the LC separation. 

Zone refining is one of a class of techniques known as fractional solidification in which a separation is brought about by crystallization of a melt without solvent being added (see also Crystallization) (1 8). SoHd—Hquid phase equiUbria are utilized, but the phenomena are much more complex than in separation processes utilizing vapor—Hquid equiHbria. In most of the fractional-solidification techniques described in the article on crystallization, small separate crystals are formed rapidly in a relatively isothermal melt. In zone refining, on the other hand, a massive soHd is formed slowly and a sizable temperature gradient is imposed at the soHd—Hquid interface. 

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