Abstract: The role of the detector is to convert the change of sample composition and content in the column effluent into a signal for detection. Commonly used detectors include ultraviolet absorption, fluorescence, differential refractive index, and chemiluminescence.
1. Ultraviolet-visible detector (UVD)
The ultraviolet-visible absorption detector (UVD) is one of the most widely used detectors in HPLC, and almost all liquid chromatographs are equipped with this detector. It is characterized by high sensitivity, wide linear range, low noise, suitable for gradient elution, detection limit of up to 1ng for strongly absorbing substances, no damage to the sample after detection, can be used for preparation, and can be used in series with any detector. The working principle and structure of the UV-visible detector is similar to the general spectrophotometer, in fact, it is equipped with a flowable UV-visible photometer.
(1) Ultraviolet absorption detector
Ultraviolet absorption detectors usually use a deuterium lamp as the light source. The deuterium lamp emits continuous wavelengths in the ultraviolet-visible region, and a grating monochromator is installed with a wide wavelength selection range (190nm-800nm). It has two flow cells, one for reference and one for measurement. The ultraviolet light emitted by the light source irradiates the flow cell. If both flow cells pass through a pure uniform solvent, they have almost no absorption at the ultraviolet wavelength. The photocell The intensity of the radiation received on the device is equal, and no signal is output. When the component enters the measuring cell, it absorbs a certain amount of ultraviolet light, so that the intensity of the radiation received by the two photoelectric cells is not equal. At this time, there is a signal output, and the output signal size is related to the concentration of the component.
Limitations: The choice of mobile phase is limited, that is, solvents with a certain ultraviolet absorption cannot be used as mobile phases. Each solvent has a cut-off wavelength. When ultraviolet light less than this cut-off wavelength passes through the solvent, the light transmittance of the solvent drops to 10 % Or less, therefore, the operating wavelength of the UV absorption detector cannot be less than the cut-off wavelength of the solvent.
(2) Photodiode array detector (PDAD)
Also known as fast scanning ultraviolet-visible spectroscopic detector, it is a new type of light absorption detector. It uses a photodiode array as a detection element to form a multi-channel parallel operation. At the same time, it detects the optical signals of all wavelengths split by the grating and then incident on the array receiver, and then quickly scans the diode array to collect data to obtain the absorption value (A ) Is a three-dimensional chromatogram of the function of retention time (tR) and wavelength (l). In this way, the spectral data corresponding to the chromatogram of each component can be observed in time to quickly determine the wavelength with the best selectivity and sensitivity.
In the single-beam diode array detector, the light emitted by the light source first passes through the detection cell, and the transmitted light is dispersed by the holographic grating into polychromatic light, which is incident on the array element, so that all wavelengths of light are detected on the receiver at the same time. The optical signal method on the array receiver is quickly scanned and extracted. Each image only needs 10ms, which is far beyond the speed of the chromatographic peak, so it can be scanned with the peak.
2. Fluorescence Detector (FD) Fluorescence detector is a high-sensitivity, selective detector that can detect compounds that can produce fluorescence. Some non-fluorescent substances can be chemically derivatized to generate fluorescent derivatives, and then fluorescence detection. The minimum detection concentration can reach 0.1ng / ml, which is suitable for trace analysis; under normal circumstances, the sensitivity of the fluorescence detector is about 2 orders of magnitude higher than that of the ultraviolet detector, but its linear range is not as wide as that of the ultraviolet detector. In recent years, the laser-induced fluorescence detector produced by using laser as the light source of the fluorescence detector greatly enhances the signal-to-noise ratio of fluorescence detection, and thus has high sensitivity, and is widely used in trace and ultra-trace analysis.
3. Differential refractive index detector (differentialrefractiveIndexdetector, RID)
The refractive index detector is a concentration-type universal detector that responds to all solutes. Some components that cannot be detected with a selective detector, such as high molecular compounds, sugars, and fatty alkanes, can be detected with a differential detector . The differential detector is based on continuously measuring the change in refractive index between the sample flow path and the reference flow path to determine the sample content. When light enters another medium from another medium, the two materials will be refracted due to their different refractive indices. As long as the refractive index of the sample component and the mobile phase are different, it can be detected. The greater the difference between the two, the higher the sensitivity. The detector output is proportional to the solute concentration within a certain concentration range.
4. Electrochemical detector (elec) chemical detector (ED)
Electrochemical detectors mainly include ampere, polarography, coulomb, potential, conductivity and other detectors, which are selective detectors and can detect electrically active compounds. At present, it has been widely used in the determination of various inorganic and organic anions and cations, metabolites of biological tissues and body fluids, food additives, environmental pollutants, biochemical products, pesticides and medicines. Among them, the conductivity detector is most used in ion chromatography.
The advantages of the electrochemical detector are:
â‘ High sensitivity, the minimum detection amount is generally ng level, and the pg level can be reached;
â‘¡It has good selectivity and can measure extremely trace amounts of electroactive substances in a large number of non-electroactive substances;
â‘¢Wide linear range, generally 4 to 5 orders of magnitude;
â‘£ The equipment is simple and the cost is low;
⑤ Easy to operate automatically.
5. Chemiluminescence detector (c. Iluminescence detector, CD) The chemiluminescence detector is a new type of fast and sensitive detector developed in recent years, because of its simple equipment, low cost, wide linear range and other advantages. The principle is based on the fact that certain substances undergo chemical reactions at room temperature to generate reaction intermediates or reaction products in an excited state, and when they return from the excited state to the ground state, they emit photons. Since the energy of the excited state of a substance comes from a chemical reaction, it is called chemiluminescence. When the separated components are eluted from the chromatographic column, they are immediately mixed with an appropriate chemiluminescent reagent, causing a chemical reaction, causing the luminescent substance to emit radiation, and its light intensity is proportional to the concentration of the substance.
This detector does not require a light source or a complex optical system. As long as there is a constant flow pump, the chemiluminescent reagent is pumped into the mixer at a certain flow rate, so that it is quickly and uniformly mixed with the column effluent to produce chemiluminescence , Photoelectric multiplier tube turns the optical signal into an electric signal, then it can be detected. The minimum detection amount of this detector can reach 10-12g.
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