Injection hydrostatic

Hydrodynamic injection can also be performed by using gravity to generate AP [41]. This injection mode is also called hydrostatic injection. The inlet of the capillary is placed in the sample vial and this is then raised during a period of time, creating a difference in height (Ah, in cm) between the inlet and the outlet of the capillary. The sample enters the capillary by siphoning. The amount and volume injected are derived from Eqs. 17.46 and 17.47, respectively, after substitution of AP with Eq. 17.48. 

Samples are introduced into the capillary by either electrokinetic or hydrodynamic or hydrostatic means. Electrokinetic injection is preferentially employed with packed or monolithic capillaries whereas hydrostatic injection systems are limited to open capillary columns and are primarily used in homemade instruments. Optical detection directly through the capillary at the opposite end of sample injection is the most employed detection mode, using either a photodiode array or fluorescence or a laser-induced fluorescence (LIF) detector. Less common detection modes include conductivity [1], amperometric [2], chemiluminescence [3], and mass spectrometric [4] detection. 

Hydrostatic injection. This type of injection is achieved by dipping the end of the capillary into a solution containing the sample while inducing a slight vacuum at the other end. Alternatively, a positive pressure can be applied to the sample solution. 

Homonuclear coupling, 143 Horseradish, 337 Hydrogen lamp, 199 Hydrostatic injection, 117 Hyphenated technique, 8 Hypsochromic effect, 195... 

Anion Hydrostatic injection Electrokinetic injection Sample stacking ... 

A major advantage of capillary gel electrophoresis is that resolution is maintained with increasing field strength, owing to efficient heat dissipation. Introduction of the sample into the gel-filled capillary is typically done electrokinetically (Chapter 6), because hydrostatic injection is not possible with capillaries blocked with gel. Typical injection times between 1 and 20 sec are used at field strengths between 100 and 400 V/cm.28 Coated capillaries should be used to prevent the EOF from pumping the gel out of the capillary. 

With hydrostatic injection mechanisms, injection reproducibility can be better than 1-2% RSD. The volume of sample loaded is a function of the capillary dimensions, the viscosity of the buffer, the applied pressure, and the time, and it can be calculated using... 

Figure 6.4 Comparison of sensitivities for different sample injection techniques using a 50 mg/ml peptide mixture (a) hydrostatic injection, (b) electromigration injection where the sample is dissolved in buffer, and (c) electromigration injection where the sample is dissolved in water. (Adapted from Ref. 1 with permission.)...

Sample is introduced into the capillary by hydrostatic injection (gravity, pressure, or vacuum) or by electromigration injection (voltage). 

The introduction of the samples onto the capillary column can be carried out by either displacement techniques or electrokinetic migration. Three methods of displacement or hydrostatic injection are available a) direct injection, or pressure b) gravity flow, or siphoning and c) suction. The electrokinetic injection method arose from findings that electroosmosis act like a pump (80). Both methods have advantages and disadvantages. For example, a bias has been reported in electrokinetically injected... 

Figure 5 (Top) Electropherogram of 100 nM FITC-insulin under HPCE conditions that employed uncoated capillaries, 25 im i.d. and 150 im o.d., total lengths of 25-30 cm, length to detector 12-15 cm, buffer of 0.05 M sodium phosphate with 0.025 M K2S04 at pH 7.5, applied voltage 1000 V/cm, hydrostatic injection. (Bottom) Electropherogram of 100 nM FITC-insulin and 50 nM Fab. Peaks 2, 3, and 5 are FITC-insulin, while peaks 1 and 4 are due to the formation of the complex of Fab with FITC-insulin in peaks 2 and 5, respectively. An He-Cd laser was used as the excitation source (66). (Reproduced with permission from the copyright holder, American Chemical Society and Analytical Chemistry.)...

The hydrostatic injection method is less precise than in HPLC because injection loops do not exist for volumes between 5—50 nL. The quantity entering the capillary is dependent upon many of the parameters that appear in the well-known Poiseuille expression which gives the flow rate F in a tube (radius r, length L) for a liquid having a dynamic viscosity 17 (expression 8.8). The application of this formula results in an approximate value for what might be termed the entering flow rate in the capillary. 

Table 31-1, p. 366, shows a comparison of the hydrostatic and electromigrative sample injection methods. This indicates that anion analysis can be performed at sub ppm levels using hydrostatic injection and at low ppb levels with electromigrative enrichment. 

The sample, typically a few nanoliters, can be introduced into the capillary by hydrostatic injection (gravity, pressure, or vacuum) or by electromigration. The sample volume should generally be less than 2% of the total capillary length. For gravity introduction, the capillary sample end is dipped into the sample (which may be as small as 5 fiL) and raised for a short predetermined time to allow sample to flow into the capillary. Or, it is inserted into a pressurized vial to force sample into the capillary. Or it is drawn in by suction from the other end of the capillary. After injection, the sample vial is replaced with a buffer reservoir. Alternatively, the sample end is immersed in the sample solution and a relatively low voltage is ap-phed for a few seconds, for example, 2000 V for 10 s. This injects the small volume of sample by electroosmosis. 

Reproducibility with hydrostatic injection is on the order of 1 to 2%. The advantage of electroosmosis injection is that more sample can be introduced, improving detection limits. The disadvantage is that because of differences in ion mobilities, the sample plug that enters the capillary is not representative of flie sample. 

Figure 3.28 The three types of hydrostatic injection used in CE.

Injection volumes are in the nanoliter range to avoid system overloading, since the total volume of the capillary is in the /rl range. Direct injection techniques have been developed to ensure efficient and reproducible injection. Techniques employed are electrokinetic injection (i.e., electromigration injection), hydrodynamic injection by pressure or vacuum, and hydrostatic injection by gravity. Organic acids are almost exclusively detected with indirect UV, whereas other analytes have been measured by direct or conductivity detection. 

Hgure 4 Electropherogram of 100-fold dilution of urine from a subject supplementing with creatine. The separation was carried out at 17kV with hydrostatic injection for 20 s, using a 30 mmol r phosphate-150 mmol r sodium dodecyl sulfate buffer, pH 6, and detection by absorbance at 214nm. The unlabeled peak is a system peak. 

The introduction of samples into capillaries by means of differential pressure (hydrostatic injection) has become the most popular method in capillary electrophoresis. For sample introduction, the sample vial is raised to a defined level above the detection reservoir for a specified time frequently about 5 s. To terminate the injection, the end of the capillary is removed from the sample and replaced into the vial containing the background electrolyte. 

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