Analysis of the Types of Common Detectors Used in High Performance Liquid Chromatography

The role of the detector is to convert the changes in composition and content of the sample in the column effluent into signals that can be detected. Commonly used detectors include ultraviolet absorption, fluorescence, refractive index, and chemiluminescence.

1. Ultraviolet-visible detector (UVD)

The UV-visible absorption detector (UVD) is one of the most widely used detectors in HPLC, and almost all liquid chromatographs are equipped with this detector. Its characteristics are high sensitivity, wide linear range, low noise, suitable for gradient elution, detection limit for strongly absorbing substances up to 1 ng, no damage to samples 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 that of a general spectrophotometer, and is actually a UV-visible spectrometer equipped with a flowing ground.

(1) Ultraviolet absorption detector The ultraviolet absorption detector uses a xenon lamp as a light source, while the xenon lamp emits a continuous wavelength in the ultraviolet-visible region, and a grating-type monochromator is installed, which has a wide wavelength selection range (190 nm to 800 nm). . It has two flow cells, one for reference and one for measurement. The UV light from the light source illuminates the flow cell. If both flow cells pass through a homogenous homogeneous solvent, they absorb almost no light at the UV wavelength. The received radiation intensity is equal and no signal is output. When the component enters the measuring cell, a certain amount of ultraviolet light is absorbed, so that the intensity of the radiation received by the two photoelectric tubes is not equal. At this time, there is a signal output, and the size of the output signal is related to the component concentration.

Limitations: The choice of mobile phase is subject to certain restrictions, that is, solvents with certain UV absorption cannot be used as the mobile phase, and each solvent has a cutoff wavelength. When ultraviolet light less than the cutoff wavelength passes through the solvent, the transmittance of the solvent is reduced to 10 Below %, therefore, the UV absorption detector's operating wavelength cannot be less than the cutoff wavelength of the solvent.

(2) Photodiode array detector (PDAD)

Also called a fast scanning UV-visible spectrophotometer, 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 light signals split by the grating and then enters all the wavelengths on the array receiver. Then it rapidly scans the diode array to collect the data and obtains 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.

Single-beam diode array detectors, the light emitted by the light source first passes through the detection cell, the transmitted light is dispersed by the holographic grating into multi-color light and hits the array element, so that all the wavelengths of light are simultaneously detected at the receiver. Optical signals on array receivers are quickly scanned and extracted electronically. Each image requires only 10 ms, far exceeding the peak velocity of the chromatographic peak, so it can be scanned with the peak.

2. Fluorescence detector (FD)

The fluorescence detector is a highly sensitive, selective detector that detects fluorescent compounds. Some of the non-fluorescent substances can be chemically derivatized to produce fluorescent derivatives and then subjected to fluorescence detection. The minimum detectable concentration is 0.1 ng/ml, which is suitable for trace analysis. In general, the fluorescence detector is about 2 orders of magnitude more sensitive than the UV detector, but its linear range is not as wide as the UV detector. In recent years, laser-induced fluorescence detectors using a laser as a light source for fluorescence detectors have greatly enhanced the signal-to-noise ratio of fluorescence detection, and thus have a high sensitivity, and have been widely used in trace and ultratrace analysis.

3. Differential refractive index detector (RID)

The refractive index detector is a concentration type universal detector that responds to all solutes. Some components that cannot be detected by selective detectors, such as macromolecular compounds, carbohydrates, aliphatic alkanes, etc., can be detected with a differential detector. . The differential detector is based on the continuous measurement of the change in refractive index between the sample flow path and the reference flow path to determine the sample content. When light enters from one medium to another, refraction occurs because the refractive indices of the two materials are different. As long as the refractive index of the sample components and the mobile phase are different, they can be detected. The larger the difference between the two, the higher the sensitivity. In a certain concentration range, the output of the detector is proportional to the solute concentration.

4. Electrochemical detector (elec) chemicaldetector, ED)

Electrochemical detectors mainly include amperometric, polarographic, coulomb, potential, and conductivity detectors. They are selective detectors that can detect compounds that are electrically active. At present, it has been widely used in the determination of inorganic and organic anions and cations, metabolites of biological tissues and body fluids, food additives, environmental pollutants, biochemical products, pesticides, and medicine. Among them, the conductivity detector is most widely used in ion chromatography.

The advantages of electrochemical detectors are:

1 The sensitivity is high, the minimum detection amount is generally ng grade, and there are pg grades.

2 selectivity is good, can measure a large number of non-electroactive substances in trace amounts of electroactive substances;

3 wide linear range, generally 4 to 5 orders of magnitude;

4 The equipment is simple and the cost is low;

5 easy to operate automatically.

5. chemiluminescence detector (c. iluminescencedetector, CD)

The chemiluminescence detector is a rapid and sensitive new detector developed in recent years because of its advantages of simple equipment, low cost, and wide linear range. The principle is based on the chemical reaction of certain substances at room temperature to generate reaction intermediates or reaction products in the excited state. When they return from the excited state to the ground state, they emit photons. Because the energy of the excited state of matter is from a chemical reaction, it is called chemiluminescence. When the separated components are eluted from the column, they are immediately mixed with appropriate chemiluminescence reagents to cause a chemical reaction that causes the luminescent substance to produce radiation. Its light intensity is proportional to the concentration of the substance.

The detector does not require a light source or a complicated optical system. As long as there is a constant-flow pump, the chemiluminescence reagent is pumped into the mixer at a certain flow rate, so that it is rapidly and uniformly mixed with the column effluent to generate chemiluminescence. The optical signal is converted into an electrical signal by a photomultiplier tube, which allows detection. The minimum detection of this detector can reach 10-12g.