Detectors flow cell designs

By assuming that a proportional increase in the amount of sample injected results in a proportional increase in the detector response for the solute band of interest, the detector response for chromatogram I in Figure 7 will increase 14 times when the maximum sample volume of 7 /xL is injected. However, for the 4.6-mm i.d. column, the detector response will increase 400 times when the maximum sample volume of 200 (lL is injected. By taking into account the relative detector responses for the 0.5-/xL injection, at the maximum sample injection volumes, the 4.6-mm i.d. column with the 20-/liL detector flow cell will produce approximately five times the detector response of the 1-mm i.d. column with the 5-/zL flow cell. In most cases, studies can be designed to provide excess sample because aqueous environmental samples are seldom limited with respect to volume. 

Fluorescence detection, because of the limited number of molecules that fluoresce under specific excitation and emission wavelengths, is a reasonable alternative if the analyte fluoresces. Likewise, amperometric detection can provide greater selectivity and very good sensitivity if the analyte is readily electrochemically oxidized or reduced. Brunt (37) recently reviewed a wide variety of electrochemical detectors for HPLC. Bulk-property detectors (i.e., conductometric and capacitance detectors) and solute-property detectors (i.e., amperometric, coulo-metric, polarographic, and potentiometric detectors) were discussed. Many flow-cell designs were diagrammed, and commercial systems were discussed. 

Preparative separations in the grams per injection level are different. Separations are run isocratic in 1- to 3-in columns with large pore, fully porous packings (35-60jUm). An analytical, two-pump system can just barely reach the 20-mL/min flow rates needed to run a 1-in column. Special preparative HPLC systems deliver flow rates of 50-500 mL/min to handle the larger bore columns. A stream splitter is used to send part of the flow through a refractive index detector with a flow cell designed for concentrated solutions. 

Commercially available HPLC instrumentation was originally designed for use with standard-bore columns (4.6 mm I.D.). Detector flow cells were optimized for maximum sensitivity with these analytical columns, injectors were designed to introduce microliter quantities of sample, and pumps were designed to be accurate and reproducible in the milliliter flow-rate ranges commonly employed with standard-bore columns. However, these instruments are not well suited for use with small-bore columns, as the dispersion introduced by the large volumes is detrimental to the separation. In addition, the reproducibility and accuracy of the pumping system at the low flow rates required are questionable. 

Testing apparatus should be designed to minimize band spreading external to the column (e.g., short, narrow connecting tubing between the column and injector and detector, low dead-volume detector flow cell, etc.). 

Low-dispersion HPLC systems are necessitated by the increasing trend of using shorter and narrower HPLC columns, which are more susceptible to the deleterious effects of extra-column band-broadening. HPLC manufacturers are designing newer analytical HPLC systems with improved instrumental bandwidths compatible with 2-mm i.d. columns by using micro injectors, smaller i.d. connection tubing, and detector flow cells. A new generation of ultra-low dispersion systems dedicated for micro and nano LC is also available. 

An example of the former is the ultraviolet spectrophotometer which may be of fixed wavelength (usually 254 or 280 nm) or variable wavelength design. The detector functions by monitoring the change in absorbance as the solute passes through the detector flow cell, i.e. it utilises the specific property of the solute to absorb ultraviolet radiation. 

Careful consideration must be given to the design of the detector flow cell as it forms an integral part of both the chromatographic and optical systems. A compromise between the need to miniaturize the cell volume to reduce extracolumn band broadening... 

Liquid phase chemiluminescence detectors usually consist of a postcolumn reactor (section 5.8) connected to a fluorescence detector with its source disabled [104,137,138,143-145]. The column eluent is combined with one or several reagents that initiates the desired chemiluminescence reaction. The intensity of light emission depends on the rate of the chemical reaction, the efficiency of production of the excited state, and the efficiency of light emission from the excited state. The chemiluminescence intensity is sensitive to environmental factors such as temperature, pH, ionic strength, and solution composition. In addition, the detection system has to be designed to accommodate the time dependence of the chemiluminescence signal to ensure that adequate and representative emission occurs in the detector flow cell. 

The sensitivity of UV detectors is generally in the low nanogram range although this can be increased by sample derivatisation. The sensitivity of detection may also be increased by alterations in the design of the detector flow cell and it is important to emphasise that the volume of the flow cell should be chosen to match the HPLC system thus a very low volume flow ceU (e.g. 1-2 /il) should be used with microbore, or very small particle columns where very high column efficiencies are anticipated, whereas larger flow cells (e.g. 5 jttl) should be used with more conventional systems. 

Category III noise Is reduced by strategies combining optimization of both detector and non-detector hardware as In reduction of reciprocating pump pulsation, the use of low thermal coefficient, matched pairs of photodiodes for reference and sample beam optical trains, and thermal coupling of reference and sample photodiodes. Pump pulsation noise can also be reduced by proper flow cell design ( ). 

We determined the efficiency for Carbon-14 at about 20% in our rebuilt cell. We have done no work with Tritium. We found that the detector s radioactivity response does not reach a maximum with increasing high voltage but steadily increases. At a count efficiency of 20%, the background is 100 cpm. Simplified details of the LSM-1 flow cell design are given in Figure 11 by permission of Nuclear Enterprises, Inc. The cell consists of ... 

In case you have an additional detector in front of your mass spectrometer, for example, a UV detector, you also have to take care of the detector flow cell pressure limit. Depending on the design principle, the maximum pressure limit of commercial UV flow cells can vary between 870 and 4350 psi (60 and 300 bar). Please be aware that it is not only the MS connection tubing that generates an additional pressure load to your UV flow cell many mass spectrometers use internal switching valves to introduce calibrant solutions into the MS ion source, which block the flow path completely for a fraction of seconds when they are actuated, thus... 

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