High Performance Liquid Chromatography and Nuclear Magnetic Resonance
NMR is one of the most effective methods for the determination of molecular structure, with the characteristics of simplicity, accuracy, high specificity and non-destruction of samples. It has not only been used to quantitatively determine a large number of pharmacologically active drugs, but also to distinguish and quantitatively determine the stereoisomers of some drugs. It shows that this method has the advantages of simplicity and accuracy as a quantitative method for drugs, and can also be used as a supplement for some drug quantification methods without reference substances.
The combination of NMR and HPLC, GC, SFE, SFC, GPC, CE, etc., enables the combination of fast and efficient separation and powerful structural analysis. The idea of the combination of HPLC and NMR began as early as the 1970s. In the early 1980s, Bruker first developed LC/NMR technology based on its world-leading NMR technology. In recent years, this technology used in combination of NMR and chromatography has become more mature. After continuous improvement and perfection, fully automatic operation and routine use of HPLC/NMR spectrometers have been used all over the world. As we all know, liquid chromatography is the best way to separate complex mixtures, and NMR is a powerful tool for structural analysis. The combination of the two has long been a desire of scientists.
A typical high-performance liquid chromatography (HPLC) coupled with nuclear magnetic resonance (NMR). Liquid chromatography includes: HPLC auto-sampler, HPLC pump, chromatographic column and incubator, non-NMR detector, such as UV (ultraviolet-visible), etc. The flowing liquid enters the HPLC/NMR interface from the detector. The interface is attached with a loop (LOOP) for storing HPLC peaks in the middle. From the HPLC/NMR interface, it is led to an NMR probe with a flow cell or a waste liquid collector. After passing through the probe, the flowing liquid enters the fraction collector for recovery, and each chromatographic peak is further studied by various NMR techniques. A mass spectrometer can also be connected to the splitter at the HPLC and NMR interface, which further becomes an LC-NMR/MS hybridization instrument.
The basic operation modes of HPLC/NMR include on-flow mode, stop-flow mode and peak parking or loop-collection mode. These three modes have their own characteristics. For example, the advantage of the continuous flow mode is that the NMR information of all components can be obtained in one analysis, but the disadvantage is that the quality of the obtained NMR spectra is poor; the advantage of the stop-flow mode is that the quality of the NMR spectra is better, and the disadvantage is that there is obvious peak broadening, and the analysis time is long; the characteristic of the peak storage mode is that the chromatographic peaks are collected and temporarily stored in different capillary circuits, and each flow fraction is measured one by one by the NMR spectrometer. In actual operation, depending on the characteristics of the sample and the purpose of detection, the operation mode can be selected.
The HPLC/NMR method can be used to detect drug impurities with impurity content at 9% level. The method is simple, fast and accurate. In the detection of drug impurities, HPLC/MS determination and HPLC/NMR determination are both simple and fast methods, but the NMR determination in HPLC/NMR combination technology is not limited by the buffer salt solution used in the HPLC separation process, and can provide a lot of structural information. Therefore, HPLC/NMR combination technology is an efficient and fast technology to obtain the structural information of the learned compounds in the mixture. It has been widely used in drug research.
