Injectors and Autosamplers

230 nM, you will see the actual gradients you produce on the recorder. Ideally, each step should be vertical, with sharp shoulders and no overshoot. In practice, you want to see only a bit of rounding at the shoulders, a near vertical slope, and no ringing or overshoot at the high end of the slope change. 

My preference today would be for a high-pressure mixing system if I had to run very complex mixtures on a routine basis because these systems give the best reproducible gradients as a rule. As a routine research instrument or a methods development system, I would prefer a low-pressure, four-solvent dynamically mixed system using a dual-headed pump. I would use the deoxygenation apparatus (Fig. 6.4) to degas my solvents with helium and run them under a helium demand valve to conserve helium. 

The next important component of the system is the injector. Since it lies between the pump and the column, it also must be able to operate at high 

I POOR GRADIENT STEP I I IDEAL GRADIENT STEP I 

Autosamplers take this same loop and valve principle and automate the filling and handle-turning sequence. The major differences between models on the market are in the way they get sample into the loop and the method of cleaning between injections. Most autoinjectors use a carousel loaded with sample valves to hold samples until their turn for injection occurs. Sample vials are usually capped with a screw cap fitted with a septum, although some recent autosamplers replace the carousel with microtiter plates having 96-364 wells containing the samples for use with robotic workstations. Conical vials are available for limited samples and 1-jUL injections are possible with some 

An HPLC injector is used to introduce the sample to the column under high pressure. A common injector is the Rheodyne model 7125 or 7725 injector, 

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Some gas chromatographs are equipped with on-column injectors and autosamplers which can inject small samples sizes. Such injection systems can be used provided that sample size is within the limit of the column and detectors optimum efficiency and linearity. 

The concept of SPME was first introduced by Belardi and Pawliszyn in 1989. A fiber (usually fused silica) which has been coated on the outside with a suitable polymer sorbent (e.g., polydimethylsiloxane) is dipped into the headspace above the sample or directly into the liquid sample. The pesticides are partitioned from the sample into the sorbent and an equilibrium between the gas or liquid and the sorbent is established. The analytes are thermally desorbed in a GC injector or liquid desorbed in a liquid chromatography (LC) injector. The autosampler has to be specially modified for SPME but otherwise the technique is simple to use, rapid, inexpensive and solvent free. Optimization of the procedure will involve the correct choice of phase, extraction time, ionic strength of the extraction step, temperature and the time and temperature of the desorption step. According to the chemical characteristics of the pesticides determined, the extraction efficiency is often influenced by the sample matrix and pH. 

The injection temperature can be a signiflcant issne for thermally unstable samples or where samples are stored for hours in an antosampler prior to injection. For this reason, most manufacturers sell autosamplers with optional thermostated sample compartments. This can be done either by placing the sample tray in an air bath oven or by a condnctive temperature control of the sample rack. The need to keep samples cool prior to injection when conpled with elevated temperature separation increases the complexity of the flow system reqnired. For such application, a separate mobile phase pre-heater with a low volnme placed between the injector and the column is a good choice. Alternatively, the injector valve wonld need to be monnted ontside the antosampler or in the column oven to insure preheating of the mobile phase before the colnmn. 

The whole atomizer may be water cooled to improve precision and increase the speed of analysis. The tube is positioned in place of the burner in an atomic absorption spectrometer, so that the light passes through it. Liquid samples (5-100 mm ) are placed in the furnace, via the injection hole in the centre, often using an autosampler but occasionally using a micro-pipette with a disposable, dart-like tip. Solid samples may also be introduced in some designs, this may be achieved using special graphite boats. The sample introduction step is usually the main source of imprecision and may also be a source of contamination. The precision is improved if an autosampler is used. These samplers have been of two types automatic injectors and a type in which the sample was nebulized into the furnace prior to atomization. This latter type was far less common. 

We use a GC Top 8000 gas chromatograph coupled with a PolarisQ ion-trap mass spectrometer and equipped with an AI3000S autosampler (Thermofinnigan www. thermo.com). The steroids are separated on a DB-1 crosslinked methyl-silicone column, 15 mx 0.25 mm i.d., film thickness 0.25 pm (J W Scientific marketed by Agilent). Helium is used as a carrier gas at a constant pressure of about 35 kPa. A 1-pl aliquot of the final derivatized extract is injected into the system operated in splitless mode (valve opened at 2 min). The GC temperature program is the same described before for the quadrupole GC-MS system. The injector and transfer lines are kept at 260°C and 280°C, respectively. The ion source temperature is 225°C. A damping gas flow of helium is applied to the ion trap. 

Reliable quantitative results are obtained by external calibration if automatic injectors or autosamplers are used. This method involves direct comparison of the peak responses obtained by separately chromatographing the test and reference standard solutions. If syringe injection, which is irrepro-ducible at the high pressures involved, must be used, better quantitative results are obtained by the internal calibration procedure where a known amount of a noninterfering compound, the internal standard, is added to the test and reference standard solutions, and the ratios of peak responses of the analyte and internal standard are compared. 

A successful chromatographic analysis depends on the precise performance of the HPLC instmmentation, i.e., control of pressure, the composition of mobile phase, the performance of the analytical column, the detector, the injector or autosampler, and the electronic data handling system. 

DTD-GCxGC-ToF MS has also been used for the analysis of organic compounds in ambient aerosol particles [27-29]. In this method, samples were collected on filters, and a piece of the filter was placed into an injector liner, which was put into the cold injector by autosampler instrumentation and thermally desorbed to the GCxGC system. The use of DTD as a sample introduction method simplifies the sample preparation, as no liquid extraction is needed. 

Quadrupole GC/MS Instrument Agilent 7890 GC coupled to Agilent 5975C TAD mass selective detector (Agilent Technologies, Palo Alto, CA). GC is equipped with 30 m x 0.25 mm x 0.25 pm DB-5 column (J W, Folsom, CA), split/splitless injector, and Agilent 7860 autosampler. 

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