Specific solute property detectors

Current IPC detectors are on-stream monitors. HPLC detectors range from (1) non selective or universal (bulk property detectors such as the refractive index (RI) detector), characterized by limited sensitivity, (2) selective (discriminating solute property detectors such as UV-Vis detectors) to (3) specific (specific solute property detectors such as fluorescence detectors). Traditional detection techniques are based on analyte architecture that gives rise to high absorbance, fluorescence, or electrochemical activity. Mass spectrometry (MS) and evaporative light scattering detectors (ELSDs), can be considered universal types in their own right... 

Solute property detectors, such as spectroscopic andj electrochemical detectors, respond to a physical or chemical] property characteristic of the solute which, ideally, is] independent of the mobile phase. Althou this criterion is rarely met in practice, the signal discrimination is usually sufficient to permit operation with solvent changes (e.g., flow programming, gradient elution, etc.) and to provide high sensitivity with aj wide linear response range. Table 5.4. Solute-specific detectors complement ulk property detectors as they provide high ... 

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. 

By comparison, a specific property type produces no signal (or perhaps only a very small signal) when there is no sample present. The appearance of a sample in the detector introduces a new type of signal and thus produces a relatively large signal (compared to zero signal for the baseline). This type is also called an analyte property or solute property detector. Some examples are given in Table 2. They are inherently the more sensitive type. 

In contrast, the electrochemical detector responds only to substances that can be oxidized or reduced and thus, providing the mobile phase is free of such materials, it will only detect oxidizable or reducible substances when they are eluted. It follows that this detector is not only a solute property detector but is also a specific detector. The electrical conductivity detector is a non-specific detector and used widely in ion chromatography where it occupies a unique and almost exclusive position. In contrast, the electrochemical detector, in its... 

Other detectors. The above discussions have been concerned with those detectors most commonly employed in routine HPLC analysis and which are commercially available. Many other detectors have been developed to monitor specific solute properties in column effluents and new detection systems continue to be reported in the literature, many directed to meeting the requirements imposed by microbore and capillary separation technologies. The interested reader is directed to the reviews of Yeung [46] and Fielden [47] for a more detailed discussion of detector types employed in HPLC. 

The second alternative classification is to define detectors as specific and non-specific detectors. In this sense a specific detector would be exemplified by the fluorescence detector as it detects only those substances that fluoresce. An example of a non-specific detector would be the refractive index detector that detects all substances that have a refractive index different from that of the mobile phase. The classification of detectors as specific and non-specific is acceptable, but in this book detectors will be classified as bulk property detectors and solute property detectors as it more closely associates the detector with its basic method of measurement. 

The slow development of LC from the time of Tswett, to the late 1950 s was entirely due to the lack of high sensitivity on-line detectors. Since, the inception of effective LC detectors there has been a continuous synergistic interaction between column development and detector development which has resulted in the present highly sophisticated LC systems of today. There are a number of ways of classifying LC detectors, specific and non-specific detectors, mass and concentration sensitive detectors and finally bulk property and solute property detectors. The classification of detectors as bulk property and solute property detectors is recommended. Bulk property detectors respond to a change in some overall property of the eluent such as refractive index or dielectric constant whereas solute property detectors respond to some property that is unique to the solute alone. In practice solute property detectors are rarely ideal and many respond, at least weakly, to the same property of the mobile phase as well as the solute. 

The solute property detectors described so far are those more commonly used or, have at least at some time or other formed the basis of a frequently used, commercially available detector. There are, however, a number of detectors that have been developed that have either, not been developed into a commercial product or, have found a very limited area of application. Many of these detectors have, in fact, useful potential for specific chromatographic analyses. Some of these detectors will now be described to give the reader a broader view of solute property detectors in general, and to illustrate the wide range of solute properties that have been investigated with a view to the development of viable LC detecting systems. 

Solute property detectors measure some property that is exclusive to the solute only and not to the column eluent. Among them are the most sensitive, most specific and those having the widest linear dynamic range. In general, they require to be used with very pure mobile phases or solvents free from substances that possess the property being measured. 

Multi-functional detectors monitor the column eluent by the measurement of more than one physical or chemical property simultaneously, employing a single sensing cell. To date, three bifunctional detectors and one trifunctional detector have been described. The three bifunctional detectors have combined UV absorption and fluorescent detection, UV absorption and electrical conductivity detection and UV absorption and refractive index detection. The latter uniquely combines a bulk property detector with a solute property detector producing, at least in theory, the nearest approach to a universal detector. The trifunctional detector incorporates UV absorption, electrical conductivity and fluorescence functions. Multi-functional detection provides detector versatility and a means of confirmir solute identity. Such detectors have to be designed, so that the performance specifications are not seriously compromised, and the cell and eluent conduits do not contribute significantly to peak dispersion. 

Several kinds of detection systems have been applied to CE [1,2,43]. Based on their specificity, they can be divided into bulk property and specific property detectors [43]. Bulk-property detectors measure the difference in a physical property of a solute relative to the background. Examples of such detectors are conductivity, refractive index, indirect methods, etc. The specific-property detectors measure a physico-chemical property, which is inherent to the solutes, e.g. UV absorption, fluorescence emission, mass spectrum, electrochemical, etc. These detectors usually minimize background signals, have wider linear ranges and are more sensitive. In Table 17.3, a general overview is given of the detection methods that are employed in CE with their detection limits (absolute and relative). 

Bulk-property detectors They specifically measure the difference in some physical property of the solute present in the mobile-phase in comparison to the individual mobile-phase, for instance ... 

The advantage of the suppressor technique is its higher sensitivity. In addition, the specificity of the method is also increased, since the chemical modification of eluent and sample in the suppressor system converts the conductivity detector from a bulk property detector into a solute specific detector [52]. Thus, exchanging eluent and sample cations with protons means that only the sample anions to be analyzed are detected by the conductivity detector and appear in the resulting chromatogram. 

Nonselective detectors react to the bulk property of the solution passing through. A refractive index detector monitors the refractive index of the eluate. The pure mobile phase has a specific refractive index which changes when any compound is eluted. The detector senses this difference and records all peaks hence the term nonselective or bulk-property detector. This is why the refractive index as well as the conductivity detector is not suited for gradient elution. 

An example of the second type of detector is the refractive index monitor which functions by recording the refractive changes in the eluant as the solute passes through the detector cell. Bulk property detectors, though more versatile, are generally several orders of magnitude less sensitive than specific property detectors and the choice for a particular application is often dictated by solute characteristics. 

Bulk property detectors have a somewhat restricted sensitivity that is directly due to the measuring principle on which they function. Consider an hypothetical bulk property detector that monitors, for example, the density of the eluent leaving the column. Assume that it is required to detect the concentration of a dense material, such as carbon tetrachloride (e.g. specific gravity 1.595), at a level of 1 pg/ml in heptane (e.g. specific gravity 0.684). This situation is typical for a bulk property detector and will be favorable for this hypothetical detector, as the solute to be detected exhibits a large difference in density from that of the mobile phase. 

Chromatography has as its basis the transformation of a complex multi-component sample into a time-resolved, separated analyte stream, usually observed in the analog differential signal mode. The chromatographic sample is thus distinctive in that analytes are changing in nature and with time. An essential feature of chromatographic instrumentation is a detector for qualitative and quantitative determination of the components resolved by the column this should respond immediately and predictably to the presence of solute in the mobile phase. An important class of solute properly detectors are those giving Selective , or Specific information on the eluates. Spectral property detectors such as the mass spectrometer, the infrared spectrophotometer and the atomic emission spectrometer fall into this class. Such detectors may be element selective , structure or functionality selective or property selective . 

This classification is concerned with whether the detector responds to a specific feature of the analyte of interest or whether it will respond to a large number of analytes, irrespective of their structural properties. In terms of the previous classification, it may be considered that solute detectors are also usually selective detectors, while solvent detectors are general detectors. 

Solute equilibrium between the mobile and stationary phases is never achieved in the chromatographic column except possibly (as Giddings points out) at the maximum of a peak (1). As stated before, to circumvent this non equilibrium condition and allow a simple mathematical treatment of the chromatographic process, Martin and Synge (2) borrowed the plate concept from distillation theory and considered the column consisted of a series of theoretical plates in which equilibrium could be assumed to occur. In fact each plate represented a dwell time for the solute to achieve equilibrium at that point in the column and the process of distribution could be considered as incremental. It has been shown that employing this concept an equation for the elution curve can be easily obtained and, from that basic equation, others can be developed that describe the various properties of a chromatogram. Such equations will permit the calculation of efficiency, the calculation of the number of theoretical plates required to achieve a specific separation and among many applications, elucidate the function of the heat of absorption detector. 

The most commonly used detector is the differential refractometer. For polymers, the variation in the refractive index is usually independent of molecular mass. Other detectors, like photometric detectors in the UV or IR range, can also be used besides the refractometer to measure specific properties of macromolecular solutions (Fig. 7.4). 

The final stage of the residue analysis procedures involves the chromatographic separation and instrumental determination. Where chromatographic properties of some food residues are affected by sample matrix, calibration solutions should be prepared in sample matrix. The choice of instrument depends on the physicochemical properties of the analyte(s) and the sensitivity required. As the majority of residues are relatively volatile, GC has proved to be an excellent technique for pesticides and drug residues determination and is by far the most widely used. Thermal conductivity, flame ionization, and, in certain applications, electron capture and nitrogen phosphorus detectors (NPD) were popular in GC analysis. In current residue GC methods, the universality, selectivity, and specificity of the mass spectrometer (MS) in combination with electron-impact ionization (El) is by far preferred. 

Derivatization of solute molecules can be utilized to modify properties of analytes of interest such that they may be more readily identified by a specific type of HPLC detector. 

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