On-capillary sample concentration

The selected factors are either mixture-related, quantitative (continuous), or qualitative (discrete).A mixture-related factor is, for instance, the fraction organic solvent in the buffer system. Examples of quantitative factors are the electrolyte concentration, the buffer pH, the capillary temperature, and the voltage, and of qualitative factors the manufacturer or the batch number of a reagent, solvent, or capillary. Sample concentration (see Table 1) is a factor sometimes included. However, the aim of the method tested is to determine this concentration through the measured signal, from a calibration procedure. Thus, one evaluates the influence of the sample concentration on the sample concentration, which we do not consider a good idea. 

Fig. 3.4. Effect of mobile phase selectivity on the CEC separation of barbiturates (1-6). Electrochromatography was performed at 15°C with an applied voltage of 30 kV on a 25 cm, 100 pm i.d., 3 pm Hypersil Phenyl packed capillary. Sample concentration was 170 pg ml"1 of each component with a 15 kV/5s injection. Detection was at 210 nm. a) ACN-50 mM phosphate buffer, pH 4.5-water (4 2 4 v/v/v), b) MeOH-50 mM phosphate buffer, pH 4.5-water (5 2 3 v/v/v). From Euerby et al [26], Journal of Microcolumn Separations, 1999. Reproduced with permission of John Wiley Sons, Inc.

J.-B. Kim, K. Otsuka and S. Terabe, On-line sample concentration in micellar electrokinetic chromatography with cationic micelles in a coated capillary, J. Chromatogr. A, 912, 343-352, 2001. H. Harino, S. Tsunoi, T. Sato and M. Tanaka, Applicability of micellar electrokinetic chromatography to the analysis of estrogens in water, Fresenius J. Anal. Chem., 369, 546-547, 2001. 

The advent of high-resolution capillary gas chromatography (HR-CGC) with on-column injection has resulted in improved GC analysis of polymer additives [92-94]. The solution of the additive mixture is injected directly into the cold end of the capillary column by means of a cold injector. Thus, sample discrimination, the instantaneous evaporation of the sample solvent, is avoided. The nonvaporising, on-column injection combined with very high resolution of the capillary columns allows accurate separation, identification and quantification of additives of complex mixtures. With the solvent venting technique, the sample is introduced into the column without splitting and sample concentrations... 

Electrophoretic injection can be used as a means for zone sharpening or sample concentration if the amount of ions, particularly salt or buffer ions, is lower in the sample than the running buffer. Because sample ions enter the capillary based on mobility, low-mobility ions will be loaded to a lesser extent than high-mobility ions. For this reason, the presence of nonsample ions will reduce injection efficiency, so electrophoretic injection is very sensitive to the presence of salts or buffers in the sample matrix. The disadvantages of electrophoretic injection argue against its use in routine analysis except in cases where displacement injection is not possible, e.g., in capillary gel electrophoresis (CGE) or when sample concentration by stacking is necessary. 

The evaluation of robustness should be considered in the development of the assay and will depend on the type of procedure under development. It must show the reliability of a method with respect to deliberate variations in method parameters. If measurements are susceptible to variations in analytical conditions, the analytical conditions should be suitably controlled or a precautionary statement might be included in the procedure. One consequence of the evaluation of robustness may be that a series of system suitability parameters is established to ensure that the validity of the analytical procedure is maintained whenever used. Typical parameters to be tested would be the following sample concentration, sample stability, labeling variability (if applicable), injection variability, reagent lot-to-lot variability, and capillary vendor. 

Capillary isotachophoresis is usually performed in constant current mode, which implies the invariable ratio between concentration and electrophoretic mobility of ions. Therefore, bands that are less concentrated than the LE are sharpened, whereas those that are more concentrated than the LE are broadened to adapt their concentration to the requested constant value between concentration and electrophoretic mobility. The consequence of this unique property of CITP is that each sample component can be concentrated to an extent that depends on its initial concentration and the concentration of the leading electrolyte. Therefore, the opportune selection of composition and concentration of the leading electrolyte allows the enrichment of diluted analytes. 

The concentration procedure was performed on the sample within 3 days of arrival at the laboratory. Upon arrival, all ether extract samples were stored at —10 °C in a freezer. The ether elution after the water was frozen out was further dried by passage through anhydrous sodium sulfate. The sample was then concentrated by a Kudema-Danish evaporator to 2 mL. After concentration, 1.0 mL of the 2-mL concentrated sample was base extracted to remove the chemicals interfering with the capillary GC analysis. The base extraction procedure developed in this study is the following ... 

Aqueous samples are extracted with hexane or with methylene chloride by liquid-liquid extraction using a separatory funnel or a mechanical shaker, or by microextraction. Aqueous samples can also be extracted by solid phase extraction using a C-18 cartridge. Selection of sample volume should be based on the extent of sample concentration that may be needed to achieve the required detection level in the analysis, as well as the use of packed or capillary column. A larger sample concentration is required for packed column than that for capillary column analysis. U.S. EPA recommends the extraction of 1 L sample to a final volume of 1 mL for wastewater analysis performed on a packed column. For the analysis of potable water by GC-ECD on a capillary column, concentration of a 35-mL... 

An oven temperature in the range of 200°C and detector and injector temperatures around 300°C and 250°C, respectively, should give good separation, sharpness of peaks, and fast analysis time. Electron capture detector (ECD) is the most commonly used detector for trace level analysis of PCBs by gas chromatography (GC), exhibiting a response to an amount below 0.1 ng PCBs. Thus, on a capillary column, an IDL in the range of 5 pg/L can be achieved. With proper sample concentration steps, a detection level several-fold lower to IDL may be obtained. Other halogen-specific detectors such as Hall electrolytic conductivity detector can also be used to analyze PCBs. 

From Eq. (6.3), sample loading is dependent on the EOF, the sample concentration, and the sample mobility. /i,e = electrophoretic mobility of the analyte, fxeo = EOF mobility, V = voltage, r = capillary radius, C = analyte concentration, t = time, L = capillary total length. 

Fig. 6.22. Electrochromatographic separation of Gly-Tyr (1), Val-Tyr-Val (2), methionine enkephalin (3), and leucine enkephalin (4) on monolithic methacrylate capillary column with a pore size of 492 nm. (Reprinted with permission from [55]. Copyright 1999 Wiley-VCH). Conditions Mobile phase 10% of aqueous 10 mmol/L sodium 1-octanesulfonate and 90% of a 2 8 mixture of 5 mmol/L phosphate buffer pH=7.0 and acetonitrile. UV detection at 215 nm. Total sample concentration 1 mg/mL.

Tomlinson, A.J., Guzman, N.A. and Naylor, S. (1995) Enhancement of concentration limits of detection in CE and CEMS a review of on-line sample extraction, cleanup, analyte preconcentration, and microreactor technology. J. Capillary Electrophor., 2 (6), 247-66. 

As a rule, a separation method should be used for both purification and concentration of the sample. The classic method for peptides and proteins is a reverse-phase liquid chromatography preparation of the sample, followed by a concentration step (often lyophiliza-tion) of the fraction of interest. During those steps performed on very small quantities of sample, loss on the sample can occur if care is not taken to avoid it. Lyophilization, for instance, can lead to the loss of the sample absorbed on the walls of the vial. The use of separation methods on-line with the mass spectrometer often are preferred. Micro- or nano-HPLC [32,33] and capillary electrophoresis [34], both coupled mainly to electrospray ionization/mass spectrometry (ESI-MS), are used more and more. 

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