Injectors for HPLC

FIGURE 13.7 The loop injector for HPLC, (a) the load positon—the sample is loaded into the loop via a syringe at atmospheric pressure, and (b) the inject position—the mobile phase sweeps the contents of the loop onto the column. 

Sample Injector for HPLC 7725/7725i/9725/9725i, Product Bulletin 209, Rheodyne, Rohnert Park, CA, http //www.rheodyne.com/lit/docs/2320617A.pdf... 

Fig. 1 Injectors for HPLC (a) External sample loop (b) Internal sample loop.

The FIA system constructed consisted of the enzyme-immobilized column integrated with gold electrodes on a silicon chip, a pump for HPLC (minimum flow rate OIL min ), an injector for HPLC (sample size 0 2 or 0 5 L, Rheodyne), a potentiostat (handmade), and a data processor Commercially available flange-type fittings were used for the connection of the biosensing device with other components of the FIA system... 

Besides differences in column design, we need to note that instrimentation is quite different for HPLC relative to the classical mode. Pumps, injectors and detectors are all important components in the achievement of high performance. In addition, automated injection is available which could be important in the clinical chemistry field. 

For HPLC, the injector is a valve. In the charge position, a 50- jL syringe is used to fill the sample loop that holds a specific volume of sample solution. The valve is switched to the run position, and the eluent carries the sample out of the sample loop and into the column (auto samplers are also available for HPLC). A recording of the detector output is automatically started at the time of injection and produces a chromatogram of the separated components. 

Spruce, B. and Bakalyar, S. R., Troubleshooting Guides for HPLC Injector Problems, Rheodyne.com/Docs/Tech/Docs/TSl.html... 

Operation Qualification. Operation qualification (OQ) is the process of establishing that the instrument or system modules operate according to the functional requirements in a suitable environment. For an HPLC system, operation of the pump, injector, and detector will be tested at this stage. Typical OQ tests for HPLC modules and a UV-Vis spectrophotometer are as follows ... 

Some of the injector systems in common use for HPLC are described in the following paragraphs. 

The TDU equipment (commercially available from Gerstel GmbH, Miilheim an der Ruhr, Germany) is fully automated and connected on-line to a GC equipped with a programmable temperature vaporizer (PTV) injector for simultaneous cryotrapping of the analytes before injection. Another approach for analyte desorption is to place the stir bar in a small volume of a conventional HPLC liquid (or mobile phase) for HPLC analysis. The SBSE stir bars are trademarked as Twisters they can also be purchased from Gerstel. For more detailed information on SBSE technology, the reader is referred to two recent review articles.44 45... 

In the fiber mode, the sorbent coated fiber is housed in a microsyringelike protective holder. With the fiber retracted inside the syringe needle, the needle is used to pierce the septum of the sample vial. The plunger is depressed to expose the sorbent-coated fiber to the sample. After equilibrium is reached or at a specified time prior to reaching equilibrium, the fiber is retracted into the protection of the microsyringe needle and the needle is withdrawn from the sample. The sorbent is then interfaced with an analytical instrument where the analyte is desorbed thermally for GC or by solvents for HPLC or capillary electrophoresis. For the in-tube mode, a sample aliquot is repeatedly aspirated and dispensed into an internally coated capillary. An organic solvent desorbs the analyte and sweeps it into the injector [68,130,133]. An SPME autosampler has been introduced by Varian, Inc., that automates the entire process for GC analyses. 

The need for unattended and precise sample injection for HPLC has lead to development of a wide variety of automated sample injection devices. Autosamplers function in essentially the same way as manual injectors, except that the sample is introduced automatically from a sample vial held in a carousel or an X-Y grid (Fig. 3.15). The carousel format provides a reliable and rapid means of moving samples past an injection station, whereas the XY grid format allows a convenient random access configuration. 

The operations of pump components, injectors, and detectors can be expected to be less troublesome when mobile phases are filtered. For HPLC applications, the 0.2-pm-pore-size filter is typically selected for removal of particu-... 

Injector fittings detector column Major Improvements are needed In conventional columns (4.6 mm i.d.) used for HPLC. Even though columns are available that can provide more than 10,000 plates/column or more than 100,000 plates/meter, large column to column variations can be found between manufacturers and from one batch to the next ( ). Further Improvements require better understanding of column variabilities and the separation processes occurring In the column. 

This chapter provides an overview of modern HPLC equipment, including the operating principles and trends of pumps, injectors, detectors, data systems, and specialized applications systems. System dwell volume and instrumental bandwidth are discussed, with their impacts on shorter and smaller diameter column applications. The most important performance characteristics are flow precision and compositional accuracy for the pump, sampling precision and carryover for the autosampler, and sensitivity for the detector. Manufacturers and selection criteria for HPLC equipment are reviewed. 

Solid-phase microextraction (SPME) A sample preparation technique that uses a fused silica fibre coated with a polymeric phase to sample either an aqueous solution or the headspace above a sample. Analytes are absorbed by the polymer coating and the SPME fibre is directly transferred to a GC injector or special HPLC injector for desorption and analysis. 

SBSE is a novel sample preparation method introduced by Baltussen et al. ° based on the same mechanisms as SPME. In SBSE, a magnetic stirring bar coated with PDMS is added to water samples of 10 to 200 ml to promote the transport of analytes into the coating polymer. After a predetermined extraction period, the analytes are thermally desorbed in the GC injector or solvent extracted for HPLC analysis. The main advantage of SBSE is that 25 to 100 jA PDMS polymer is used instead of 0.5 /rl as in SPME. The applications developed with SBSE have shown low detection limits (sub-ng to ng 1 levels) and good repeatability, confirming to the great potential... 

Injection port facility for introducing the sample mixture into the mobile phase. In GC the injector is maintained at least 50°C above the maximum column temperature to be used and the sample is introduced via a rubber septum using a microlitre syringe for HPLC see multiport injection valves. 

Desorption for GC-analysis is performed directly in the GC-injector by increasing the temperature. For HPLC analysis an interface has been constmcted for solvent extraction of the analyte from the fibre, followed by introduction of the solvent into the LC injector [59]. 

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