HPLC injectors

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

As nowadays wine making has factory dimensions, the need for a rapid method is important, so a flow injection techniqne for liqnid-solid extraction coupled to an HPLC was developed [368]. The target analytes were extracted from wine in a continuous way by means of a minicolumn packed with C18, the elntion was carried ont by means of acid water (pH 2) and acetonirile, then the solvent was evaporated by nitrogen stream on-line to a HPLC injector. Malvidin-3-glucoside, cyaniding-3-glncosyde, and peonidin-3-glncoside antocyanins were determined in a more rapid, accurate, and sensitive way than with the traditional methods. 

Carryover. If the automated method is coupled with online HPLC analysis and carryover is observed from one sample to the next, the HPLC injector must be considered as a potential source of carryover. Normally, if carryover is observed,... 

HPLC injector with a 50-pl sample loop fluorescence spectrophotometer, operating at an emission wavelength of 460 nm with an excitation wavelength of 300 nm. 

Brief mention needs to be made regarding the employment of internal standards. While it is desirable to employ an internal standard for procedures that involve significant sample preparation procedures, there is no ideal choice for an internal standard for amino acid analysis. This fact is due to the wide spectrum of chemistries exhibited by the various amino acids. If one is analyzing for a single amino acid (or class, e.g., the hydrophobic amino acids), it is possible to choose an internal standard that mimics the chemistry of that particular amino acid very well. However, for the overall amino acid profile, an internal standard will do nothing more than allow the analyst to make nonvolumetric solution transfers and correct for variability of the injection volume by the HPLC injector. Unfortunately, the employment of an internal standard can actually skew the apparent recoveries for the overall amino acid profile. 

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,... 

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. 

Automation of sample preparation of pharmaceutical solids has been quite successful. A variety of instrumentation is commercially available from several vendors. The instruments are capable of handling a dosage form, placing it into a vessel, adding solvent, and mechanically homogenizing the sample until the analyte is dissolved. Typically, the instrument can then dilute to the desired concentration. Some models are also capable of transferring the prepared solution to a HPLC injector or an ultraviolet spectrophotometer for measurement. These instruments use accurate balances to control the accuracy of solvent additions and dilutions. 

Most SPME applications involve GC. Following extraction, the analytes are thermally desorbed in the chromatograph injector. More recently, the scope of apphcation has been extended to nonvolatile and thermally unstable compounds by coupling SPME to LC. Desorption is performed at an appropriate interface consisting of a standard six-way HPLC injector with a special fiber-desorption chamber used instead of the sample loop. A different approach to SPME-LC called in-tube SPME has also been developed, it uses an open tubular fused-silica capillary column instead of the typical SPME fiber. This latter method has been used for the determination of carbamate pesticides in water with relative standard deviations of... 

Rotary sample loop injector Figure 13.6 illustrates the most common design of HPLC injector. To introduce the sample as in GC with a syringe through a... 

Using the style of the diagram of the six-port rotary HPLC injector valve (Fig. 13.6) draw two diagrams showing the flow through the injection loop, the valve, and the column during the load and injection positions. 

The early design of HPLC injector systems was strongly influenced by those used in gas chromatography (GC). This is obvious when one considers that, although septum injection is still popular in GC, it has been almost totally supplanted in HPLC by high pressure injection... 

HPLC injectors serve two basic functions to load a precise volume and to enable the use of high pressure. The small particle and column diameters used in analytical and microfluidic HPLC can cause large backpressures, requiring high pressure for moving the sample solution and mobile... 

Most modern HPLC injectors use the loading coil and coil bypass system. The sample is loaded at low pressure into the coil which is temporarily isolated from the solvent flow to the column on injection, the flow is diverted through the coil washing the sample onto the top of the column. With this method solvent is continuously pumped through the column at constant flow, thus minimizing injector stop-flow problems. 

Figure 3.5 Manual HPLC injectors with external loop.

With packed HPLC columns, conventional HPLC injectors can be used. With open capillary columns, split or splitless injection is needed in order not to overload the columns. Special injection techniques have been developed for these purposes, since standard GC techniques cannot be used at high pressures. With knowledge about the critical data of the sample solvent, a retention gap can be used to separate solvent from solutes and remove the solvent prior to solute focusing on the analytical column (Figure 5.8). 

This component of the HPLC injector valve contaiiis the sample that is presented by the syringe. These are 20-pL and lO-jiL stainless steel loops. Loops are also constructed from PEEK. 

Three type of syringes are shown here. All have square-tip needles and are compatible for use in HPLC injectors. The left-hand syringe has a detachable needle. The right-hand syringes have permanent needles of different lengths. 

---