Autosampler problems

Fig. 9 Low and variable internal standard responses caused by autosampler problem occurred during the middle of run injection. Extraction evaporation-free protein precipitation sample volume 50 uL IS volume 150 pL. Reproduced from ref. [36] with permission from Elsevier...

Figure 11(A) shows a principle sketch of a SEC set-up. The eluent (solvent) is pumped at a constant flow rate through the system. A small amount of polymer solution (typically 200 pL) is injected manually or with an autosampler. The main part comprises a set of columns (typically 3-4 columns+pre-column) typically packed with microporous styrene-divinylbenzene, porous glass, or silica. In the routine analytical laboratory it is especially useful to have a pre-column in order to collect impurities that might be present in the samples. If many different samples are to be analyzed, it is necessary to check the reliability of the columns frequently to avoid artefacts due to residues from previous samples still held on the column. In order to avoid problems, samples should be purified before they...

Isolate the problem area—is it the pump, the autosampler or the detector ... 

Figure 4 shows peak area precision vs. injection volume for a typical autosampler. Note that excellent peak area precision of 0.2% RSD was readily achievable for an injection volume >5 J,L. Precision levels are much poorer (0.5-1% RSD) for sampling volumes <5 J,L, attributable to the finite resolution of the sampling syringe and associated digital stepper motor. To obtain optimum peak area precision, the analyst must avoid potential problem situations such as an overly fast sample... 

Measurement of pH and conductivity for a range of aqueous samples is common in most water laboratories. Surprisingly, many laboratories still use manual methods for these measurements. TTie automation of these techniques is not easy. Two approaches to solve this automation problem, descrihed in this section, both used the same large volume autosampler. 

To solve the problem, the affected run can be reinjected in a different LC-MS system or on the same system after the needle of the autosampler is cleaned. 

It is necessary to study stability in solution in the solvent used to prepare sample solutions for injection in order to establish that the sample solution composition, especially the analyte concentration, does not change in the time elapsed between the preparation of the solution and its analysis by HPLC. This is a problem for only a few types of compound (e.g. penicillins in aqueous solution) when the sample solution is analysed immediately after the preparation of the sample solution to be injected. The determination of stability in solution is more of an issue when sample solutions are prepared and then analysed during the course of a long autosampler run. While the acceptance criteria for stabUity in solution may be expressed in rather bland terms by making a statement such as, e.g. the analyte was sufficiently stable in solution in the solvent used for preparing sample solutions for reliable analysis to be carried out , in practice it has to be shown that within the limits of experimental error, the result of the sample solution analysis by the HPLC method is the same for injections at the time for which stability is being validated as for injections immediately subsequent to the sample solution preparation. While this may be done by a subjective assessment of results with confidence limits, strictly speaking a statistical method known as the Student s t-test should be used. 

Type 2 autosamplers behave in a very similar way to type I except partial loop volume can be filled and delivered. Additionally, this allows the injection of low volume with high precision compared to type I autosamplers. Both types of autosamplers suffer from carry over problems if appropriate wash cycle and wash solvents are not used between each sampling from the vials. 

Problems with the sample introduction may arise both in manual injection and during autosampling. Table 7 summarizes the problems related to sample introduction. 

In the case of autosamplers, although they are considered as time saving devices, their function is associated with some common problems. For example, needle depth adjustment is very critical when there is not enough sample available a needle blockage may occur from a septum. Another common problem related to the autosampler is carryover that causes the appearance of a peak in a blank injection following injection of a high concentration of sample. Sample stabihty is a figure of merit that certainly has to be evaluated to avoid imprecision and lack of sensitivity. 

Karl Fischer titration Chemical titration that is water-specific. Material can be either added directly to a solvent or heated in an oven, with the headspace purged and bubbled through solvent. Specific to water only and very precise. Units can be purchased with an autosampler. Measurement takes only a few minutes. Equipment is e3q>ensive and requires solvents. Minimal calibration required. Sample size is small, which may pose a problem for heterogeneous mixtures. 

Unless care is exercised by the analyst, both these forms of contamination can go unnoticed and erroneous results may be reported for individual samples. Problems with cross-contamination should normally be identified during the validation phase of method development by the judicious use of blanks to test for problems with general laboratory contamination, sample preparation, and the autosampler. Carryover is... 

Many reasons can be a possibility. If other causes, such as a malfunction of the autosampler, can be excluded the nebulizer tube could be contaminated or clogged. —> In many cases the problem is solved after a cleaning of the tube for a couple of minutes in an ultrasonic bath. Sometimes the gas pressure for the nebulizer needs to be increased or decreased. 

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