Retention time measurement

It follows that measurements must be made with a precision of about 0.2 second if quantitative results are to be of any value. It is seen from figure 4 that the experimental points lie very close to the line and a fairly accurate measurement of the distribution of the two isotopes can be obtained from retention time measurements. This method has very limited areas of application and is given here, more to demonstrate the effect of unresolved impurities on retention time, than to suggest it as an alternative to adequate chromatographic resolution. In some cases, however, particularly in the analysis of isotopes, it may be the only practical way to obtain a quantitative evaluation of the mixture by a liquid chromatographic method. 

The repeatability of gas sample separation is presented in Fig. 6.12. The separation peaks of the mixed gaseous analytes match those for each individual analyte quite well, irrespective of the analyte concentration, which correlates to the peak height (Fig. 6.12a, b). Although manual sample injection may introduce a certain degree of variation in retention time measurement, using a gas marker can significantly improve such measurement, as demonstrated in Fig. 6.12c, in which decane is used as a marker. 

For the more volatile components of water samples, i.e. those with boiling points up to about 250°C, gas chromatography has been a favoured technique for several decades. However, with the realization that retention time measurements alone are insufficient to identify organics there has been an increasing move in recent years to connect a gas chromatograph to a mass spectrometer in order to provide unequivocal identifications. Element-specific detectors are another recent development. 

Because of some peak tailing, the number of theoretical plates was based on peak width at one-half peak height Ns5.S4 he pooled standard deviation (all temperatures) of retention time measurements (dfs34) was t 0.007 minutes. 

Size-exclusion chromatography can be used to analyze protein-protein interactions. Bloustine et al. (2003) presented a method to determine second virial coefficients (B2) of protein solutions from retention time measurements in size-exclusion... 

An unequivocal identification of an unknown compound is unlikely by chromatographic processes alone. Not the least of the reasons for this is the need for comparison to standards thereby assuming reasonable prior assurance of the possible identity of the unknown. It should be noted that in addition to retention time measurements obtained on two or more column systems, if reasonable care has been exercised, quantitative measures of the suspect compound should also correspond, thus providing an additional secondary identification. In other words, whatever the unknown compound may be, it cannot be a mixture of two components on one column and a single component on the second column without... 

The retention time of a solute in HPLC (tR) is defined as the time necessary for maximum elution of the particular solute. This is analogous to retention time measurements in GC. Retention volume (VR) of a solute is the solvent volume required to elute the solute and is defined by Equation 3.4, where F is the flow rate of the solvent. 

For better reproducibility of the retention parameters, relative retention times referenced to a standard chemical have been used in several GC applications. The added reference chemical should be chosen so that it elutes in the middle of the chromatogram. There is always some degree of error in the retention time measurement, and the relative retention times of chemicals having short or long retention times might not be very accurate. To obtain better reproducibility, it is recommended that several different reference chemicals are used for the calculation of relative retention times. However, in this case the use of RIs becomes more attractive. It is a much more reliable to use RIs than absolute or relative retention times because in retention indices (RI) measurements, the retention is measured relative to a homologue series. 

In this and the following expressions, the corresponding retention volumes or linear separations on the chromatogram, both of which are directly proportional to retention time, may be substituted in the equations. Where the value of tQ is small, Rr may be estimated from the retention times measured from the point of injection (tR(2)/fR(i))-... 

Measurements are rapid and with standard commercial equipment the retention time measurements can be made over a wide range of temperature. 

The certain identity of a component depends on the accuracy of the retention time measurement and thus the correct identification of the position of the peak maximum is critical. The peak height is taken as the difference between the signal at the point of the peak maximum and that directly beneath the peak maximum on the baseline produced under the peak. The projected baseline under the peak is calculated using a procedure similar to that described below. 

Retention time measured from time of injection from the capillary column. 

In this relationship, is the average retention time measured for the analyte in the mobile phase and ty is the void time of the system (i.e., the observed elution time for a totally nonretained solute). 

Guillet (9) uses IGC to estimate the degree of crystallinity in semicrystalline polymers and to compute the surface area of polymer powders. Linear polyethylene is used as the vehicle to demonstrate the former application. In the latter, a requisite is to evaluate the partition coefficient for a selected probe/polymer combination (n-decane/PMMA in the present instance). Once this is obtained via IGC, simple retention time measurements become suitable as routine analytical or control methods to monitor surface areas in polymer powders. 

In the isothermal chromatography the column is kept at constant temperature. At its exit one detects the atoms which have not decayed during the retention time. Measurements should be done at several decreasing column temperatures to find the interval in which the survival yield is considerably less than the initial 100 percent and so the retention time is comparable with lx. Let Ys be the normalized (per fluence unit) yield at a high temperature 7 s and Ya - such yield at a low temperature 7 a. Their ratio rf is a direct measure of the retention time, Zr because ... 

Therefore, measurements carried out over a range of concentrations Ci and C2 with pure binary solutions, allow the determination of fci, Ai,mi,fc2, A2 and m2. From the retention times measured with pseudo-binary systems, i.e., for pulses of component 1 over concentration plateaus of solutions of component 2 alone (Cl = 0) and for pulses of component 2 over plateaus of solutions of component 1 alone (C2 = 0), one can derive from Eq. 4.96 ... 

The relative retention time should fulfil the criteria. Only if step 1 is positive can step 2 be made. For GC-analyses these criteria can be very strict. For instance, the relative or absolute retention time measured in the sample should not differ by more than 0.2% (from the relative retention time in the last measured external standard solution). In HPLC analyses, a higher deviation has to be accepted. 

Figure 8.2 NanoLC-chip-MS of replicate injections (n — 5) of an eight-protein digest, 250 fmol each, (a) Overlay of 5 total ion chromatograms for the corresponding analyses, (b) Scatter plot of intensity measurements for 5 replicates and 2230 peptide ion clusters, (c) Distribution of RSD values on retention time measurements, (d) Variation of MS response for different tryptic peptides according to sample amount loaded. Conditions enrichment/trap volume of 40nL LC separation channel of 43 x 0.075 x 0.050mm both packed with Zorbax Ci8 separation media a 5 pL injection of 80 ng tryptic digest of 8 proteins was performed except for (d) where variable amounts (1-1000 ng) of digest were injected.

The retention time measured in the sample does not differ by more than 0.2% from the retention time determined in the last measured external standard solution. 

The syringe is removed, the port is closed and the eluent stream is started again. An example of such a stopped flow injector, which is commercially available (Varian), is shown in Fig. 7. For precise retention time measurements, stopped flow injection is less preferable because of the time needed to pressurize the system, which might be of the order of 3-2D sec depending on the capacity of the pumping device. On the other hand, for routine analysis this is less important and stopped flow injection might be a very valuable, cheap and reliable sample introduction technique. 

Flo. 9. Relation between rate constant of cracking over aluminum fluoroborate-alumina catalyst at 400° and retention time measured gas chrojnatographically over the same catalyst and at the same temperature for ethylbenzene, m-ethyltoluene, and p-ethyltoluene (49). 

The resolution factor is usually estimated from the peak retention times and widths observed in a chromatogram of a mixture of solutes. However, in a rigorous way, a more accurate estimation requires the separate injection of the individual compounds. This is particularly true for closely eluting peaks. It has been established that the retention times measured at the peak apex, and mote so the peak widths, are different if the measurement is made on individually injected solutes or on the peaks in a mixture. This difference is more pronounced when the peak shape cannot be described simply by a Gaussian profile, and where the center of gravity of the peak does not correspond to the peak apex (8). Nevertheless, the chief drawback of the resolution factor Rs is the fact that it does not take into account the relative peak height (9) of the... 

The interaction by n coordination of Ag(I) between silver ions and C=C double bonds or aromatic groups is the basic principle for many effective chromatographic separations. Silver trifluoroacetate and silver tiifluoromethanesulfonate are very soluble in poly(methylphenylsiloxane), due to the presence of phenyl groups. Such stationary phases were studied for the GC separation of benzene-cyclohexene-cyclohexane mixtures. The salting-out effects and formation constants of the complex of Ag(l) with various olefinic and aromatic compounds were estimated based on retention time measurements. ... 

Mozell, M.M. and Jagodowicz, M. Chromatographic separation of odorants by the nose retention times measured across in vivo olfactory mucosa. Science, 1973, 1247-1249. 

The boiling points of peaks identified by mass spectrometry can be estimated from the boiling points of standard compounds, and enthalpies can be obtained from retention times measured at different temperatures. While the measured enthalpy correlates well between each group of logarithm of capacity ratio (log A ) values, the relationship is not consistent, as shown in Figure 4.3. 

The retention times measured in reversed-phase liquid chromatography were quantitatively analyzed using simple compounds like those used in gas... 

It is the adjusted retention volume which is directly proportional to the thermodynamic distribution constant and therefore the parameter often used in theoretical equations. In essence it is the retention time measured from the nonretained peak (air or methane) as was shown in Figure 1.5. 

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