Examining the Chromatogram

Interpretation/report The GC retention time of a naphthalene standard and the mass spectrum of this peak confirm its presence. Because of the complexity of the chromatograms of the petroleum products and the pesticide sample, you find it impossible to examine the chromatogram of each. However, a comparison of the GC fingerprints (i.e., the matching of chromatographic peaks and comparison of peak ratios) clearly shows that the sample consists of naphthalene dissolved in kerosene. 

Let us now examine the chromatogram produced by this separation. Starting at the point of injection, we follow the baseline to the first deflection. After about 2min, we see a small positive deflection immediately followed by a small negative deflection, which then returns to the baseline. The center of this peak complex is called the void volume (Vo). It represents the amount of mobile phase contained inside the column, but outside the packing material. It is the mobile phase volume necessary to wash out the sample solvent. 

Test C Examine the chromatograms obtained in the test for omeprazole impurity C. The principal spot in the chromatogram obtained with test solution (b) is similar in position and size to the principal spot in the chromatogram obtained with reference solution (a). Place the plate in a tank saturated with vapor of acetic acid R. The spots rapidly turn brown. 

If you examine the chromatograms shown in this and the next chapter, you will see that the elution peaks look very much like the Gaussian or normal error curves... 

Perform the isocratic separation under the calculated conditions and examine the chromatogram If the separation is not yet satisfactory, solvent selectivity should be used to further improve the separation. 

Selectivity—A method meets selectivity requirements if dosage results are not disrupted by the presence of molecules other than the analytes— matrix interferents, for example. The selectivity of the method is not expressed as a number. It is evaluated by examining the chromatograms of real sample matrix extracts containing the analytes or spiked with analytes. 

The chromatogram can finally be used as the series of bands or zones of components or the components can be eluted successively and then detected by various means (e.g. thermal conductivity, flame ionization, electron capture detectors, or the bands can be examined chemically). If the detection is non-destructive, preparative scale chromatography can separate measurable and useful quantities of components. The final detection stage can be coupled to a mass spectrometer (GCMS) and to a computer for final identification. 

Purification of anthracene. Dissolve 0-3 g. of crude anthracene (usually yellowish in colour) in 160-200 ml. of hexane, and pass the solution through a column of activated alumina (1 5-2 X 8-10 cm.). Develop the chromatogram with 100 ml. of hexane. Examine the column in the hght of an ultra-violet lamp. A narrow, deep blue fluorescent zone (due to carbazole, m.p. 238°) will be seen near the top of the column. Immediately below this there is a yellow, non-fluorescent zone, due to naphthacene (m.p. 337°). The anthracene forms a broad, blue-violet fluorescent zone in the lower part of the column. Continue the development with hexane until fluorescent material commences to pass into the filtrate. Reject the first runnings which contain soluble impurities and yield a paraffin-hke substance upon evaporation. Now elute the column with hexane-benzene (1 1) until the yellow zone reaches the bottom region of the column. Upon concentration of the filtrate, pure anthracene, m.p. 215-216°, which is fluorescent in dayhght, is obtained. The experiment may be repeated several times in order to obtain a moderate quantity of material. 

A convenient way to examine the light-scattering response from each angle of detection (detector signal versus elution volume) is shown in Fig. 20.5 for a fully hydrolyzed PVA. The chromatograms from all 15 angles of detection from a Dawn-F (Wyatt Technology Corporation, Santa Barbara, CA) are... 

The best way to identify the MWs of unknown glycols and glycol ethers is to examine the mass spectra of the TMS derivatives. The TMS derivatives are identified in the chromatogram by plotting masses 73 and 147. The higher of the two high-mass peaks, which are 15 Daltons apart, is the molecular ion of the TMS derivative. If no peaks are separated by 15 Daltons, add 15 to the highest mass peak observed to deduce the MW. 

Two-dimensional TLC on silica gel G has been used to identify alcohol ether sulfates in liquid laundry detergents. The spots of the chromatograms were examined by UV, IR, and NMR spectroscopy and the spectra compared with those of standard surfactants [283]. 

The chromatogram was then irradiated with short-wavelength UV light (X = 254 nm) for 3-5 min and examined again under long-wavelength UV light (X = 365 nm). 

Detection and result The chromatogram was freed from mobile phase, immersed in the dipping solution for 2 s and then examined immediately after brief drying in a stream... 

To attempt clarification of this situation the effect of axial dispersion on the chromatogram was examined in two ways ... 

The value of w is likely to be 40-60 pi, unless the system has been specially designed for use with small bore columns. The effect on a chromatogram is more serious the earlier a peak is eluted, so we will examine the worst case, which is the effect on an unretained solute (one that travels at the same speed as the mobile phase). 

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