Resolution and Analysis Time

Ultra-performance HPLC (UPLC) utilizes sub-2-pm porous particles inside packed microbore columns up to 150 mm long. Significant improvements in terms of resolution, analysis time, and detection sensitivity have been reported. A side-by-side comparison of HPLC and UPLC was made to determine concentrations of alprazolam in rat plasma.10 UPLC provided a four-fold reduction in terms of LC/MS/MS cycle time that translated into higher sample throughput. Another important... 

The silica-based SEC packings which are currently being employed were Introduced on the market in 1980 and there has since been very little improvement In their properties in terms of pore size distribution, particle size etc. Now, initiated by theoretical Investigations of the resolution of SEC columns, a new family of SEC packings Is beginning to be produced and marketed. The new packings are characterized by a particle size smaller than 5 / m, adequate porosity, and superior bonding chemistry. The Bio-Series 6F 250 column from DuPont de Nemours Is a typical example. However, the performance - with reference to resolution, analysis time and peak capacity — of this novel material has not yet been evaluated. 

Auger spectroscopy also offers acceptable detectibility for many situations encountered in corrosion science. This can be as low as 0.01 %, although this is seldom realized in common practice since low detectibility conflicts with other desirable qualities such as resolution, analysis time, and the need to minimize beam current to minimize beam damage. 

A similar quantitative analysis of six water-soluble vitamins (B, B2, Bg, C, nicotinamide, and pantothenic acid) in a pharmaceutical formulation using CZE in uncoated fused silica capillaries with UV detection was described by Fotsing et al. (91). Eor the B-group vitamins, a good compromise among resolution, analysis time, and analyte stability was obtained by use of 50 mM borax buffer pH 8.5. A capillary wash with sodium hydroxide was necessary between successive runs to minimize absorption of excipients from the pharmaceutical formulation to the capillary surface, otherwise giving rise to a progressive decrease of the electro-osmotic flow. 

The high diffusivity and low viscosity of sub- and supercritical fluids make them particularly attractive eluents for enantiomeric separations. Mourier et al. first exploited sub- and supercritical eluents for the separation of phosphine oxides on a brush-type chiral stationary phase [28]. They compared analysis time and resolution per unit time for separations performed by LC and SFC. Although selectivity (a) was comparable in LC and SFC for the compounds studied, resolution was consistently... 

Trying to determine which column is ideal for a specific analysis can be difficult with over 1000 different columns on the market [74]. A proper choice implies a definition of parameters such as column material, stationary phase (polarity), i.d., film thickness and column length. Guides to column selection are available [74,75]. The most important consideration is the stationary phase. When selecting an i.d., sample concentration and instrumentation must be considered. If the concentration of the sample exceeds the column s capacity, then loss of resolution, poor reproducibility and peak distortion will result. Film thickness has a direct effect on retention and the elution temperature for each sample compound. Longer columns provide more resolving probe, increase analysis times and cost. 

A typical hplc column (25 cm x 4.6 mm, packed with a 5 jum bonded silica stationary phase) will have an efficiency corresponding to a plate number of 10 000-15 000. For many separations, this efficiency is far more than is needed, as often a plate number of 3000-5000 will give baseline resolution of all solutes. If this is the case, using a conventional column will waste analysis time and sol-... 

In the last decade, capillary electrophoresis (CE) has become one of the most powerful and conceptually simple separation techniques for the analysis of complex mixtures. The main reasons are its high resolution, relatively short analysis times, and low operational cost when compared to high-performance liquid chromatography (HPLC). The ability to analyze ultrasmall volume samples in the picoliter-to-nanoliter ranges makes it an ideal analytical method for extremely volume-limited biological microenvironments. 

Studied responses were resolution, " " analysis time, migration time, plate count, tailing factor, tablet content, peak area, peak height, peak width, and peak area/migration time ratio. ... 

CE has recently emerged as a powerful tool in carbohydrate analysis with enhanced resolution for isobaric isomers, shorter analysis times, and high sensitivity with EIE detection, as well as better assay reproducibility and robustness over the traditional methods. 

Once the chromatogr hic system has been selected, the problem reduces to the design of a column having the required resolution power while minimizing analysis time and possibly inlet pressure. Before discussing this problem we need to know the relationships between analysis time, flow velocity, and pressure, and between resolving,power and flow veloclly. 

Finally, the liquid carrier velocity should be considered. The flow velocity is strictly related to the operating mode. In normal elution mode, both analysis time and resolution decrease as flow velocity decreases. In general, in normal mode, flow rates in the 0.1-0.5mL min range are employed. Steric and hyperlayer modes operate at flow rate values higher than those typical of normal mode. 

Hence, many variables must be considered when optimizing a separation with respect to analysis time and resolution of enantioseparations. A natural extension was to develop computer programs to assess these variables and optimize the process automatically, and some general examples have been reported84,86. Numerous examples in addition to those in Table 2 of... 

The vast majority of analyses use long, narrow open tubular columns (Figure 24-2) made of fused silica (Si02) and coated with polyimide (a plastic capable of withstanding 350°C) for support and protection from atmospheric moisture.3 As discussed in Section 23-5, open tubular columns offer higher resolution, shorter analysis time, and greater sensitivity than packed columns, hut have less sample capacity. 

Techniques such as mono- and bidimensional thin-layer chromatography and high-voltage electrophoresis have been used in peptide separations for many years. However, the versatility, short analysis times, and high resolution of high-performance liquid chromatography (HPLC), coupled with the possibility of automating the analysis process, make this analysis technique the method of choice in peptide analyses at the present time (l-6a). 

As with the other additives used in soft drinks, caffeine and quinine can be, and often have been, detected using the same HPLC method used for other materials, such as in the method published by Williams (1986). This method separates most of the major additives used in soft drinks in a short time (4-5 min). Although some of the resolutions are not quite baseline, as would be expected in such a short analysis time, and not all of the synthetic colours are separated from each other, this is still a very impressive method. 

Like CRF-1, CRF-2 also favors short analysis times and well resolved peaks. However, there is no threshold value for resolution, and the compromise between resolution and analysis time is not as well defined as in CRF-1. Inclusion of analysis time in the denominator of an objective function may result in the loss of some resolution, compensated for by a rapid analysis time (51). This is likely to occur to some extent when the resolution threshold is defined in excess of 1 to 1.25, since the peak-valley ratio utilized by CRF-2 does not diminish to an appreciable extent until the resolution falls below this range. Note that as the resolution drops below 1 to 1.25, however, CRF-2 decreases rapidly, and it is unlikely that a short analysis time will compensate for such poor resolution (33). Nevertheless, if a user-specified minimum resolution is an absolute requirement, it is probably better to use a threshold criterion such as CRF-1 in which the desired resolution is stipulated by the user. 

The MEKC separation of organophosphate nerve agents was achieved using amperometric detection. Studies show that the use of MES buffer (pH 5.0) and 7.5 mM SDS provides the best separation performance (short analysis time and adequate resolution) [625],... 

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