Sample loaded

Capillary Electrophoresis. Capillary electrophoresis (ce) or capillary 2one electrophoresis (c2e), a relatively recent addition to the arsenal of analytical techniques (20,21), has also been demonstrated as a powerful chiral separation method. Its high resolution capabiUty and lower sample loading relative to hplc makes it ideal for the separation of minute amounts of components in complex biological mixtures (22,23). 

Because protein samples are actually ampholytes, when samples are loaded onto the gel and a current is appHed, the compounds migrate through the gel until they come to their isoelectric point where they reach a steady state. This technique measures an intrinsic physicochemical parameter of the protein, the pi, and therefore does not depend on the mode of sample appHcation. The highest sample load of any electrophoretic technique may be used, however, sample load affects the final position of a component band if the load is extremely high, ie, high enough to titrate the gradient ampholytes or distort the local electric field. 

To demonstrate the effect in more detail a series of experiments was carried out similar to that of volume overload, but in this case, the sample mass was increased in small increments. The retention distance of the front and the back of each peak was measured at the nominal points of inflection (0.6065 of the peak height) and the curves relating the retention data produced to the mass of sample added are shown in Figure 7. In Figure 7 the change in retention time with sample load is more obvious the maximum effect was to reduce the retention time of anthracene and the minimum effect was to the overloaded solute itself, benzene. Despite the reduction in retention time, the band width of anthracene is still little effected by the overloaded benzene. There is, however, a significant increase in the width of the naphthalene peak which... 

It follows from equation (2) that the sample load will increase as the square of the column radius and thus the column radius is the major factor that controls productivity. Unfortunately, increasing the column radius will also increase the volume flow rate and thus the consumption of solvent. However, both the sample load and the mobile phase flow rate increases as the square of the radius, and so the solvent consumption per unit mass of product will remain the same. 

Process Detailed Design Detailed specification based on concept design Minimize number of possible leak paths Make plant friendly to control, operate, and maintain Avoid or simplify hazardous activities such as sampling, loading and unloading Process conceptual design and codes/standards and procedures Experience on past projects/designs... 

Generally, optimizing the selectivity by choosing a gel medium of suitable pore size and pore size distribution is the single most important parameter. Examples of the effect of pore size on the separation of a protein mixture are given in Fig. 2.15. The gain in selectivity may then be traded for speed and/ or sample load. However, if the selectivity is limited, other parameters such as eluent velocity, column length, and sample load need to be optimized to yield the separation required. 

Sample load is primarily a concern in preparative gel filtration. In analytical applications the only precaution is to ascertain that the sample volume is sufficiently low as not to contribute to peak widths (and thus decrease the quality of the information) (Hagel, 1985). The concentration of the sample should not exceed 30 mg/ml for globular proteins or 5 mg/ml for polymers and DNA (Hagel and Janson, 1992). 

In all modes of chromatography, high sample loads distort peak shapes and cause an overall decrease in efficiency due to column overload. Sample loads may be increased by using organic solvents to enhance the solubility of the sample or by using higher column temperatures to lower the viscosity of... 

Elow rate determines the separation time and can significantly affect resolution and efficiency. The effect of flow rate on HETP for TSK-GEL SW and TSK-GEL SWxi analytical columns is shown in Fig. 4.6. Resolution is typically higher at slower flow rates, although results shown in Fig. 5B indicate that, with increasing sample load, the faster flow rates can give higher resolution. 

PSS SEC column dimensions were chosen to allow easy scaling of chromatography conditions without the need to optimize separations for each column dimension separately. The volume flow rate and the sample load can be calcu-... 

As an example, a separation on a standard PSS SEC column (8 X 300 mm dimension), which is done at a flow rate of 1.0 ml/min with a 100-/rl injection of a 1% sample solution (sample load 1 mg), can be reproduced exactly on a 4 X 250 mm PSS SEC column when using a flow rate of 0.25 ml/min and injecting 20 /rl of the same 1% sample solution. This corresponds to 0.2 mg of injected mass. 

Column dimension i.d. X L (mm) Typical flow rate (ml/min) Sample load (mg) Instrument requirements... 

Sample loading must be reduced in accordance with the column inside diameter. Polymers exhibit high solution viscosity, and in order to avoid band broadening due to viscous streaming the sample concentration must be reduced for narrow-bore columns. Overloading effects become noticeable at much lower concentrations using 4.6-mm columns compared to 7.5-mm columns because of the effective sample concentration in a smaller volume column. 

Fig. 6-6. Overload elution profiles of D,L-PA injected on a column (125 4 mm) packed with the L-PA imprinted stationary phase used in Fig. 6-5. Mobile phase MeCN TFA (0.01 %) FI O (2.5 %). The tendency for fronting and the increase in retention with sample load is attributed in part to saturation of the mobile phase modifier.

Fig. 6-7. Asymmetry factor (AJ of the L-enantiomer versus sample load (A) and versus flow rate (B) on L-PA-imprinted polymers. Flow rate 1.0 ml min . Mobile phase MeCN/[potassium phosphate 0.05 M, pH 7] (7/3, v/v).

For most applications, a 1 /tm film thickness is preferred because it is more universal, has less absorption, and allows for higher sample load. A thinner (0.33 jam) stationary phase is useful for higher-boiling and heat-sensitive compounds. A thicker stationary phase is better for low-boiling compounds. 

First the sample, that was loaded to about 20% of its short-term yield strength or 13.8 MPa (2,000 psi), recovered almost completely one hour after the release of the load, the net strain being 0.03%. Second, the sample loaded to 66%of its short-term yield strength, or 41.4 MPa (6,000 psi), retained a strain of 0.8% at 1,000 hours after the release of the load. The initial strain was 2.8%, the strain from the 1,000 hour creep an additional 1.7%. Thus only about one-half the creep strain was recovered. Visually extrapolating the recovery curve reveals that even after a year (104 hr.), about one-third of the creep strain (0.6%) will remain. 

Sample loading is done by loosely placing the ground sample into the sample holders of the Norelco Inverted-Sample Three-Position Spectrograph. 

F. SAMPLE LOAD STATION AND DUAL CRYSTAL COUNTING ASSEMBLE... 

FIGURE 20.7 Phase images of ethylene-propylene-diene terpolymer (EPDM) samples loaded with oil (50 wt%). Image in (a) was obtained on the unvulcanized sample and images in (b,c,d) on samples cross-linked with different amounts of sulfur curative 0.5, 1.0, 1.5 phr, respectively. 

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