Resolution versus analysis time

Figure 3.1 Resolution versus analysis time for three 5-/

We will now look at graphs of resolution versus analysis time for various hoioes of particle size and column length. As a measure for resolution, we aply use the square root of the plate count, as shown in the resolution luation. Equation 3.1. As analysis time, we use 10 times the breakthrough of an unretained peak, as shown in Equation (32). We use the van ater equation to calculate the HETP, from which we determine the plate Dt From the Kozeny-Carman equation [Eq. (3.3)] we calculate the lire drop across the column. We set an upper pressure limit of 20 MPa 3000psi). The curves will stop when this pressure limit is reached. 

Table 23-3 compares the performances of packed and open tubular gas chromatography columns with the same stationary phase. For similar analysis times, the open tubular column gives resolution seven times better (10.6 versus 1.5) than that of the packed... 

Bauer and Untz analyzed a series of cinchona alkaloids by means of straight-phase HPLC (Fig.5.15). They found that the addition of 2.65 ml of water to 1 liter of the mobile phase (chloroform - isopropanol - diethylamine(940 57 l)), which corresponds to about 75% saturation, gave optimum separation, as regards resolution versus time of analysis. To obtain the correct percentage of water in the mobile phase, the water content present in the mixture was deter-ined by the Karl Fischer method, and water was then added to obtain a final concentration of 2.65 ml/1. 

In an excellent and thorou study, IS chlorinated phenols were well resolved on a C,8 colunrn (k = 260nm) using a 31/79 acetonitrile/water (50mL citrate buffer at pH 4) mobile phase [8%]. Elution was complete in 90 min. The late-eluting peaks were sufficiently well resolved that a gradient would have been very effective at reducing analysis times. Plots of log k vs. percent acetonitrile (30-46%) at pH 4 and 6 were generated. A further plot of the minimum resolution between all analytes versus pH (3-7) and percent acetonitrile (30-64) was presented. 

Further, optimization of the oven temperature ramp versus the carrier gas flow establishes the desired final separation conditions of short analysis time with still good peak resolution and a reduced elution temperature for the analytes. The example in Figure 2.121 demonstrates the win in analysis time of more than 50% from 16 08 to 7 36 min for pentachlorophenol at a low elution temperature of 208 C. 

Supercritical fluid extraction conditions were investigated in terms of mobile phase modifier, pressure, temperature and flow rate to improve extraction efficiency (104). High extraction efficiencies, up to 100%, in short times were reported. Relationships between extraction efficiency in supercritical fluid extraction and chromatographic retention in SFC were proposed. The effects of pressure and temperature as well as the advantages of static versus dynamic extraction were explored for PCB extraction in environmental analysis (105). High resolution GC was coupled with SFE in these experiments. 

Relative to most other techniques discussed in this book, NMR has found a limited number of niche applications in food analysis. For example, the determination of oils is seeds (or fat in chocolate ), based upon low resolution, solid phase NMR is well used in quality control laboratories. Actually, most apphcations that found their way in food analysis are methods based upon the differences in relaxation times of various molecules e.g. free water molecules versus bound water molecules). Consequently, for the purpose of this particular chapter, we shall discuss only two specific applications of NMR to food analysis. These examples were chosen solely to demonstrate the broad range of applications that NMR can cover and the reader is advised that the mere fact that they were selected here should not be interpreted as a judgement of their value over other related references. 

There are two important results from this analysis. First, the rate constants for binding and dissociation can be obtained from the slope and intercept, resp>ec-tively, of a plot of the observed rate versus concentration. In practice this is possible when the rate of dissociation is comparable to ki [S] under conditions that allow measurement of the reaction. At the lower end, resolution of i is limited by the concentration of substrate required to maintain pseudo-first-order kinetics with substrate in excess of enzyme and by the sensitivity of the method, which dictates the concentration of enzyme necessary to observe a signal. Under most circumstances, it may be difficult to resolve a dissociation rate less than 1 sec by extrapolation of the measured rate to zero concentration. Of course, the actual error must be determined by proper regression analysis in fitting the data, and these estimates serve only to illustrate the magnitude of the problem. In the upper extreme, dissociation rates in excess of 200 sec make it difficult to observe any reaction. At a substrate concentration required to observe half of the full amplitude, where [S] = it., the reaction would proceed toward equilibrium at a rate of 400 sec. Thus, depending upon the dead time of the apparatus, much of the reaction may be over before it can be observed at the concentrations required to saturate the enzyme with substrate. 

The flash filament experiment as first described by Becker and Hartman (14) has since been used extensively in studies of the adsorption of gases onto refractory metals, particularly in association with other techniques. The basic method is to allow gas introduced at a known input rate to adsorb for a measured time onto a previously cleaned wire or ribbon. The gas is then desorbed by heating the sample, and the resulting pressure bursts monitored. The pressure versus time curve is referred to as a desorption spectrum, as illustrated in Fig. 4 and 5. Sticking probabilities can then be obtained from the relative adsorption times and desorption quantities. Methods of analysis of these desorption spectra (15, 16) and of the variation in thermal resolution by different heating schedules such as linear or reciprocal increase in temperature with time, have been discussed extensively by a number of authors... 

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