Get Insight into the SepaBean Gamma Machine with Engineer: Diode Array Detector

The application technology research center is a fast liquid chromatograph, which is usually composed of system pump, sample injector, separation column, detector and control system Among them, the detector module is equivalent to human eyes, which is very important for the separation and purification performance of the whole system Today, this article mainly introduces the detector module used in sepabean ® machine system of rapid liquid preparation chromatograph from Santai technology In liquid chromatography, common detector types include: UV Vis, diode array detector (DAD), fluorescence detector (FD), refractive index detector (RID), evaporative light scattering Detector, ELSD) and mass spectrometer (MS) In sepabean ® machine system of Santai technology, a dad detector (as shown in Figure 1) is used Compared with the traditional UV detector, its biggest feature is that it can detect the absorption of all wavelengths at the same time, so as to obtain complete spectral information, which has an important impact on purity judgment and wavelength selection Figure 1 Schematic diagram of diode array detector detection As shown in Figure 1, the continuous light emitted by the light source passes through the slit, passes through the flow cell, and then passes through the holographic grating to disperse into monochromatic light and irradiate it on the photoelectric detection element for detection Because the components of the sample flowing out of the chromatographic column change (generally, the incident light absorption and absorption of different structural compounds are different), the light intensity of each wavelength irradiating on the photoelectric detector will be different, so we can To see peaks of different sizes So, what are the advantages of DAD detector in the separation and purification of practical samples? We summarize the following points: 1 To provide rich spectral information of compounds - as if we had an ultraviolet spectrophotometer If the user knows enough about the sample, he can roughly determine whether the current outflow component is the target structure according to the structure information of the target molecule 2 Provide the best detection wavelength, enhance the detection sensitivity - reduce the loss of samples When the automatic collector is used for sample collection, the system collects the samples according to the preset absorbance value When some compounds with weak absorption are encountered or the sample volume is low and the absorbance is low, the loss of the samples can be effectively reduced by selecting a better absorption wavelength Note: in the sample separation and purification experiment, the maximum absorption wavelength of the sample is not always selected We should select the appropriate absorption wavelength according to the sample quantity When the injection volume is very small, we choose the wavelength with larger absorption; when the sample volume is large, in order to prevent absorption saturation, we can choose the wavelength with lower absorption, which is more conducive to sample collection 3 Provide the judgment basis of sample purity to make the experimental results more accurate and reliable In the separation of samples, when the elution time of two samples is similar and the elution peak of samples overlaps or crosses, it can be roughly determined whether the component is a single component or a mixed component according to the spectral absorption changes of all wavelengths before and after elution In addition, we can judge the purity of eluting components by the same principle of the same absorption ratio at a specific wavelength of the same compound In the following, the full band scanning function of dad detector is further introduced with a sample separation and purification example Fig 2 The separation spectrum of a sample and the full wavelength absorption spectrum of elution peak 1 in the first experiment Fig 3 The separation spectrum and the full wavelength absorption spectrum of elution peak of a sample in the second experiment are separated and purified twice with a mixture sample The elution conditions used in the two experiments are different The separation spectrum of the sample is shown in Fig 2 and Fig 3 Since the samples used in the two experiments are identical, through the following simple judgments, we can roughly conclude that the two components in the sample were eluted together in the second experiment: 1 Maximum absorption wavelength of the target component: in general, the maximum absorption wavelength of the same component is relatively fixed For example, the maximum absorption wavelength of the sample component in Figure 2 is 232nm (marked with red ellipse), and the maximum absorption wavelength of the sample component in Figure 3 is 253nm (also marked with red ellipse) Generally speaking, the change of the two maximum absorption is due to the mixing of different samples Be eluted together 2 Difference of characteristic absorption spectrum: due to the difference of sample quantity, the overall absorption strength of the same sample will be different, but the specific characteristics will not change As shown in Fig 2 and Fig 3, the sample has obvious absorption at 600 nm in Fig 3, which also indicates that the purity of elution component in the second separation experiment is not high 3 Ratio of absorption value of characteristic absorption wavelength: for a specific substance, the absorption ratio at different characteristic absorption wavelengths is generally fixed As shown in Figure 2, the ratio of the absorption values under the two characteristic absorption wavelengths (232352nm) of component 1 is about 2.0, while the ratio of the absorption values of the same two characteristic refinement wavelengths of component 1 in Figure 3 is about 6.1, indicating that the elution component 1 in the two experiments is not the same substance Combining with the separation spectra in Figure 2 and figure 3, it can be judged that the purity of component 1 in the second separation experiment is relatively low The above briefly introduces how to use the full band scanning function of dad detector to help us judge the purity of elution components The R & D personnel of Santai technology are developing more practical functions, please look forward to At the same time, for the separation and purification of samples without UV or visible light absorption, such as sugars, drugs, lipids, polymers, business tonics, undifferentiated fatty acids and amino acids, as well as combinatorial molecular libraries, Santai technology is developing the evaporative light scattering detection module (ELSD), which will be put into the market in the near future to provide customers with more perfect separation and purification solutions To learn more about the detailed specifications of sepabean ® machine, or the ordering information of the flash column used with it, please visit CBG online store.