Chromatographic peak shape

Figure 3.12 Comparison of the chromatographic peak shapes obtained from (a) analogue and (b,c) digital detectors, and the effect on peak shape of the number of data points defining the signal.

As Beer s law in absorption spectroscopy has a path length dependence, the observe volume, Vobs, or active volume of an NMR probe is an important determinant of the sensitivity of NMR measurements. The observe volume is the fraction of the total sample volume, Vtot. that returns a signal when a sample is inserted in an NMR tube or is injected into a flow system. The relationship between chromatographic peak shape, peak volume and flow rate, and sensitivity in hyphenated NMR measurements is complex and is discussed in greater detail in Section 7.2. For the purpose of this discussion, the sample is assumed to be present at a uniform concentration in a sample volume, Vtot. The probe observe factor, /o, is calculated as shown in the following equation ... 

In this case the survey scan was set as a full scan and the dependent scan as a product ion scan. The problem with data dependent acquisition is to determine the selection criteria. In most cases the system picks up the most abundant ion in the full scan spectrum. An inclusion list with masses of potential metabolites or exclusion list of known interferences significantly improves the procedure. In the example shown in Fig. 1.39, a procedure called dynamic background subtraction (DBS) was applied. This procedure considers chromatographic peak shapes and monitors not the most abundant signal in the spectrum but the largest increase of an ion in a spectrum. The advantage is that once a signal of a peak has... 

In order to determine the fatty acid composition of the triglycerides, they have to be first hydrolysed and the liberated fatty acids converted to their methyl esters, which have a good chromatographic peak shape compared to the free acids. A convenient method for achieving hydrolysis and methylation in one step is shown in Figure 11.8. 

A decongestant syrup was basified with ammonia and extracted into ethyl acetate, thus ensuring that the components extracted were in their free base forms rather than their salts, which is important for obtaining good chromatographic peak shape. Salts of bases will thermally dissociate in the GC injector port but this process can cause a loss of peak shape and decomposition. 

Kuge and Yoshikawa (3) related a change in the gas chromatographic peak shape to the beginning of multilayer adsorption on the surface of the solid. For small adsorbate volumes, the peak shape is symmetrical. As the adsorbate volume is increased, a sharp front, diffuse tail, and a defect at the front of the peak top is observed (Figure 11.2). It then acquires a diffuse front and sharp tail. This point corresponds to the B point of the BET Type II adsorption isotherm at which the relative surface area may be calculated. 

In the early years of GC, more consideration was given to partition (GLC) than to adsorption (GSC) systems. For GLC, the mechanism of retention was well understood, all of the mathematics were derived, and the chromatographic peak shapes were symmetrical. At that time, GSC had been utilized only for the separation of permanent gases. In recent years much has been accomplished in the determination of thermodynamic parameters in GSC separations. Part of the reason for the upsurge of interest was due to the desire to predict sample separations at any temperature, since most GSC data was reported at only one temperature. 

Fig. 1 Nonlinear effects in elution chromatography (a) nonlinear adsorption isotherm (b) the corresponding chromatographic peak shapes at N = 10,000 for different injected concentrations (c) the corresponding chromatographic peak shapes at N = 500 for different injected concentrations. Isotherm Langmuir type. Concentrations 2.5, 5.0, 7.5, and 10.0 units. Length of column 10 cm...

Isocratic focusing techniques, in conjunction with turbulent-flow chromatography, are also explored in this chapter. Isocratic focusing is used to improve chromatographic peak shape, enhance sensitivity, and optimize reproducibility. Isocratic focusing has also led to the development of generic methodologies that eliminate the need... 

Results with high bias—Coelution with other target or non-target analytes affects the quantitation of the target analytes due to the degradation of chromatographic peak shape. 

FIGURE 6-37. Plot of Langmuir isotherms (description of corresponding chromatographic peak shape given in parentheses). (Reprinted from reference 22 with permission.)... 

With this in mind the chromatograms produced were re-assessed, this time taking into account the peak shape that was obtained. The average k figures were then recalculated after disregarding any k values that were associated with compounds that gave poor chromatographic peak shape (denoted in Table 8.2 with an asterisk). The results of this process are shown in Table 8.3 and are summarised in the chart shown in Fig. 8.13. 

Lee et al. [88] discussed optimization of RPLC for quantitative proteomics. They especially paid attention to the fact that both a good chromatographic peak shape must be achieved to enable reliable quantification and the peak capacity must be maximized to enable identification with sufficient proteome coverage. Gradient range and slope were evaluated. 

According to the extensive review by Marco and Bombi [7], almost 100 dedicated empirical or semiempirical formulae have been proposed for the chromatographic peak shapes. Some of them may reasonably well fit experimental data, even when physicochemical rationale for them is lacking. 

Vouros and co-workers recently demonstrated the effectiveness of nanospray to improve both the S/N quality concurrent with a reduction in matrix effects such as ion suppression [102]. Rather than use a nanobore LC column, the authors constructed a unique postcolumn flow that allowed for nanospray flow rates with conventional column formats. A cylindrically symmetric flow splitter was shown to preserve chromatographic peak shape at split ratios as high as 2000 1 thus, effluent from a conventional 2.1-mm Cl8 column at 200 pi. /min was reduced to 100 nL/min. The heart of the flow splitter was a fused-silica needle emitter with an orifice diameter of 5 pm. Tlie nanoflow splitter was applied to the in vitro metabolite analysis of a test... 

---