Autosampler

The most obvious step in the automation of a chromatographic system is to automate sample injection. The ability to perform re- 

A comprehensive list of manufacturers and the type of components currently being used in HPLC can be found in McNair (1984). 

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These direct-insertion devices are often incorporated within an autosampling device that not only loads sample consecutively but also places the sample carefully into the flame. Usually, the sample on its electrode is first placed just below the load coil of the plasma torch, where it remains for a short time to allow conditions in the plasma to restabilize. The sample is then moved into the base of the flame. Either this last movement can be made quickly so sample evaporation occurs rapidly, or it can be made slowly to allow differential evaporation of components of a sample over a longer period of time. The positioning of the sample in the flame, its rate of introduction, and the length of time in the flame are all important criteria for obtaining reproducible results. 

Because of the large number of samples and repetitive nature of environmental analysis, automation is very important. Autosamplers are used for sample injection with gc and Ic systems, and data analysis is often handled automatically by user-defined macros in the data system. The high demand for the analysis of environmental samples has led to the estabUshment of contract laboratories which are supported purely by profits from the analysis. On-site monitoring of pollutants is also possible using small quadmpole ms systems fitted into mobile laboratories. 

The use of "fixed" automation, automation designed to perform a specific task, is already widespread ia the analytical laboratory as exemplified by autosamplers and microprocessors for sample processiag and instmment control (see also Automated instrumentation) (1). The laboratory robot origiaated ia devices coastmcted to perform specific and generally repetitive mechanical tasks ia the laboratory. Examples of automatioa employing robotics iaclude automatic titrators, sample preparatioa devices, and autoanalyzers. These devices have a place within the quality control (qv) laboratory, because they can be optimized for a specific repetitive task. AppHcation of fixed automation within the analytical research function, however, is limited. These devices can only perform the specific tasks for which they were designed (2). 

SPME has been utilized for deterrnination of pollutants in aqueous solution by the adsorption of analyte onto stationary-phase coated fused-siUca fibers, followed by thermal desorption in the injection system of a capillary gas chromatograph (34). EuU automation can be achieved using an autosampler. Eiber coated with 7- and 100-p.m film thickness and a nitrogen—phosphoms flame thermionic detector were used to evaluate the adsorption and desorption of four j -triazines. The gc peaks resulting from desorption of fibers were shown to be comparable to those obtained using manual injection. 

Atomic Fluorescence System - Millennium Excalibur PSA 10.055 -was used in our work. This system consists of the autosampler, the integrated continuous flow vapour generator and the atomic fluorescence spectrometer with the boosted dischar ge hollow cathode lamp and a control computer. 

However, for quantities substantially less than this level, 7- to 10-mm i.d. analytical columns can often be used in a semipreparative mode. By repeatedly injecting 300 to 500 ju,l of up to 1% polymer, reasonable quantities of polymer can be isolated. An autosampler and automated fraction collector can be setup to perform such injections around the clock. Although the larger injections and higher concentrations will lead to a loss of resolution, in some situations the result is quite acceptable, with a considerable savings in time being realized over other means of trying to make the same fractionation. 

Parts that are common in SEC, but should not be used in HOPC are a sample injector (including an autosampler) and a guard column. 

In principle, on-line SPE-LC can be automated quite easily as well, for instance, by using Such programmable on-line SPE instrumentation as the Prospekt (Spark Holland) or the OSP-2 (Merck) which have the capability to switch to a fresh disposable pre-column for every sample. Several relevant applications in the biomedical field have been described in which these devices have been used. Eor example, a fully automated system comprising an autosampler, a Prospekt and an LC with a UV... 

Figure 13.5 Schematic presentation of the procedure involved in coupled-column RPLC AS, autosampler C-1 and C-2, first and second separation columns, respectively M-1 and M-2, mobile phases S-1 and S2, interferences A, target analytes HV, high-pressure valve D, detector. Reprinted from Journal of Chromatography, A 703, E. A. Hogendoom and R van Zoonen, Coupled-column reversed-phase liquid cliromatography in environmental analysis , pp. 149-166, copyright 1995, with permission from Elsevier Science.

The principle was demonstrated using triazine herbicides as templates and by varying the type of functional monomer and the monomer composition. With a final batch size of ca. 40 mg of monomer, the consumption of monomers and template is significantly reduced and the synthesis and evaluation can take place in standard high-performance liquid chromatography (UPLC) autosample vials. After synthesis. 

FIGURE 4-4 Microprocessor-controlled voltammetric analyzer, in connection with an autosampler. (Courtesy of Metrohm Inc.)... 

The autosampler can accommodate over 100 samples, as well as relevant standard solutions. Such coupling can also address the preliminary stages of sample preparation (as dictated by the nature of the sample). The role of computers in electroanalytical measurements and in the development of smarter analyzers has been reviewed by Bond (7) and He et al. (8). 

A stainless steel column (4.6 mm internal diameter by 250 mm length) packed with 7 micron Zorbax ODS (Dupont) was equilibrated with 82 % Acetonitrile in water at a flow rate of 2.0 ml/min. provided by a Spectra Physics Model 87(X) pump and controller. The effluent was monitored at 230 nm using either a Tracor UV-Visible detector Model 970A or a Jasco Uvidec UV detector Model 1(X)-V. Peaks were recorded and calculated on a SpectraPhysics recording integrator. Model 4200 or Model 4270. Samples of 0.5 mg/ml in toluene were applied to the column automatically with a Micromeritics Autosampler Model 725 equipped with a 10 pi loop. 

The upper limit to the linear range was 5000 ngml, hut at concentrations >1000 ngml carry-over from the autosampler was observed. This could be reduced by extensive washing. 

The technician needs 1 minute per determination to load the sample into the autosampler, type in sample information, start the machine, etc., and 20 minutes for preparing the eluent per batch. 

Sample solution instability or incomplete extraction/separation would show up if several aliquots from the same sample work-up were put in a series of vials that would be run in sequence that would cover at least the duration of the longest sequence that could be accommodated on the autosample/instrument configuration. For example, if an individual chromatogram is acquired for 5.5 minutes, postrun reequilibration and injection take another 2.75 minutes, and 10 repeat injections are performed for each sample vial in the autosampler, then at least 15 60/(5.5 -I- 2.75)/10 = 11 vials would have to be prepared for a 5 P.M. to 8 A.M. (=15 hour) overnight run. If there is any appreciable trend, then the method will have to be modified or the allowable standing time limited. 

P 53] Before operation, a start-up time of about 10 min was applied to stabilize pressure in the chip micro reactor ([R 6]) [20]. As a result, a stable flow pattern was achieved. The reactant solutions were filled into vials. Slugs from the reactant solutions were introduced sequentially into the micro chip reactor with the autosampler and propelled through the chip with methanol as driving solvent. The flow rates were set to 1 pi min The slug volume was reduced to 2.5 pi. 

The chip micro reactor ([R 6]) was only one part of a complex serial-screening apparatus [20]. This automated system consists of an autosampler (CTC-HTS Pal system) which introduces the reactant solutions in the chip via capillaries. A pumping system (p-HPLC-CEC System) serves for fluid motion by hydro dynamic-driven flow. A dilution system [Jasco PU-15(5)] is used for slug dilution on-chip. The detection system was a Jasco UV-1575 and analysis was carried out by LC/MS (Agilent 1100 series capLC-Waters Micromass ZQ). All components were on-line and self-configured. 

This example presents four analytical situations (a) good control, (b) sudden shift in accuracy - perhaps the calibration material has become contaminated, (c) a gradual shift in accuracy-perhaps a reagent has exceeded its expiry date, (d) very poor precision - perhaps an autosampler is working erratically. 

Volumetric flask, 100-mL HPLC autosampler with Teflon septa... 

The electrochemical detector must be zeroed after each analysis just before the next sample injection. This procedure is necessary owing to the drifting baseline associated with the electrochemical detector. The detector is equipped with this baseline zero capability, and the adjustment can be activated through an external event output signal sent from an autosampler. 

Glass screw-cap autosampler vial, 2-mL, with Teflon-lined septum Fisons MD-800 gas chromatograph/mass spectrometer or equivalent MassLynx Software, Version 2... 

Inspect the culture tubes in the manifold to determine if there is water in the organic eluent for any sample. If a water layer is present, quantitatively transfer the organic phase into a clean culture tube using a small amount of additional solvent as necessary. Return the culture tube containing the organic extract to its proper location in the manifold rack. Remove the Cig and sodium sulfate mbes, and reinstall the silica tubes on the manifold. With the sample remaining in the culture tube, continue to apply vacuum to the manifold to remove excess solvent. When the solvent volume is < 1 mL, discontinue vacuum, and allow the sample to return to room temperature. Adjust the sample volume in the culture mbe to 1 mL with isooctane-ethyl acetate (9 1, v/v). Transfer the entire sample into an autosampler vial for GC/MS analysis. Sample extracts may be stored for up to 1 month in a refrigerator (< 10 °C) before analysis. 

Approximately 1-2 mL of the sample is transferred directly into an autosampler vial for LC/MS/MS analysis. 

Column temperature Injection volume Autosampler temperature Flow rate... 

At least four chromatographic standards prepared at concentrations equivalent to 50-70% of the limit of quantitation (LOQ) up to the maximum levels of analytes expected in the samples should be prepared and analyzed concurrently with the samples. In LC/MS/MS analysis, the first injection should be that of a standard or reagent blank and should be discarded. Then, the lowest standard should be injected, followed by two to four blanks, control samples, fortifications or investigation samples, followed by another chromatographic standard. This sequence is then repeated until all the samples have been injected. The last injection should be that of a standard. In order to permit unattended analysis of a normal analysis set, we recommend that samples and standards be made up in aqueous solutions of ammonium acetate (ca 5 mM) with up to 25% of an organic modifier such as acetonitrile or methanol if needed. In addition, use of a chilled autosampler maintained at 4 °C provides additional prevention of degradation during analysis. 

Flasks, vacuum filter, Pyrex, 500-mL Flasks, round-bottom boiling, Kontes, 50-mL, T45/50 joint Gas chromatograph, Hewlett-Packard (HP) 5890 equipped with an HP 7673A autosampler and an electron capture detector... 

Gas chromatograph, HP 5890 equipped with an HP 7673A autosampler and an HP 5972 mass-selective detector... 

Acetonitrile (ACN), HPLC grade Ammonium acetate (NH4OAC), reagent grade Ammonium hydroxide, 28%, reagent grade Autosampler vials and septa caps Celite, or equivalent... 

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