Method for simultaneously detecting contents of three alkaloids in lycoris plant

Abstract

The invention discloses a method for simultaneously detecting the contents of three alkaloids in a lycoris plant, which is capable of quickly extracting and qualitatively and quantitatively analyzingalkaloid matters of galanthamine, lycoramine and lycorine in the lycoris plant. According to the method disclosed by the invention, gradient eluting is carried out by taking an LC (Liquid Chromatography)-MS (Mass Spectrometry) technology as the basis, analysis is carried out in an MRM (Multiple Reaction Monitoring) mode under positive ion scanning by connecting LC with tandem MS, the three alkaloid matters in the lycoris plant can be quickly extracted and accurately, qualitatively and quantitatively analyzed, the analysis time can be shortened to a large extent, the analysis accuracy is increased, a detection result can be obtained within a program time which is not more than 6.0 minutes, the retention time of the three alkaloids is within 3 minutes, the adding standard recovery rate is 97.2 to 100.2 percent, the RSD (Relative Standard Deviation) is 0.8 to 4.3 percent, and the analysis speed and the analysis accuracy are greatly increased; an effective and accurate analysis method is provided for researching an accumulation law and constructive metabolism regulation of the three alkaloids in the lycoris plant.

Description

Technical field [0001] The invention belongs to the technical field of alkaloid substance detection, and specifically relates to a method for simultaneously detecting the content of three alkaloids in Lycoris plants. Background technique [0002] Lycoris spp. is a bulbous herb of the Amaryllidaceae family, many of which are ornamental and medicinal flowers unique to China, and are rich in alkaloids specific to plants of the Lycoris spp. So far, more than 500 alkaloid compounds with different structures have been isolated from Lycoris plants. Phytochemical studies have shown that these compounds have multiple pharmacological functions and have extremely high medicinal value. Among them, galantamine and licoramine can be used for the treatment of Alzheimer's disease (early symptoms of Alzheimer's disease), and lycorine is effective in antibacterial, anti-malaria, tumor treatment, anti-virus and cardiovascular protection. Important value. So far, wild Lycoris resources are still t...

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Problems solved by technology

[0004] Li Mingkai (Establishment of Lycoris alkaloid analysis method and its biosynthesis research, Anhui Agricultural University, 2012: 1-69) realized the preliminary separation of alkaloids by using thin-layer chromatography separation technology, and found that two-way thin-layer chromatography Compared with one-way chromatography, it is more suitable for the separation of many different alkaloid substances. This method mainly realizes the Preliminary characterization of alkaloid species in a sample, but not for accurate structural and quantitative analysis

[0005] Quan et al. (Quan et al., Photosynthetic characteristics of Lycoris aurea and monthly dynamics of alkaloid contents in its bulbs, African Journal of Biotechnology, 11(15): 3686-3691, 2012) utilized high-performance liquid chromatography detection technology, through samples and known The comparison of the retention time and the peak area of ​​the alkaloid standard substance under the same conditions carried out the quantitative research of galantamine and lycorine in Lycoris hudixiao, and its single extraction of Lycoris bulb material needs fresh sample 20g ( water content of about 90%), equivalent to 2g of Lycoris bulb dry sample, need to consume 110ml of methanol, 10ml of hydrochloric acid, 15ml of chloroform, which consumes a lot of plant samples and reagents, and at the same time, it only adopts the method of HPLC, according to the retention time of the substance. Content detection cannot be accurately qualitative for the substance to be tested, so the quantitative result of the substance is unreliable, and the structure, qualitative and precise quantitative analysis cannot be carried out

[0006] Sahar et al. (Sahar et al., 4'-O-Methylnorbelladine feeding enhances galanthamine and lycorine production by Leucojum aestivum L.shoot cultures, Engineering in Life Sciences, 00: 1-6, 2015) adopted the method of HPLC-ESI-MS, for Quantitative analysis of lycorine and galantamine in the lotus buds of Xia Xuepian, the elution time of lycorine is 25.5min, the elution time of galantamine is 32.2min, the HPLC analysis time is relatively long, about 36 minutes, the content detection is carried out according to the retention time and precursor ion information. Compared with the method of Quan et al., the accuracy of this method is improved, but the obvious defect is that there may be many kinds of material components with the same elution time and precursor ion size , so there are still defects in qualitative and quantitative accuracy

[0007] Moreover, in the prior art, there are also problems of long extraction time and cumbersome operation. For example, in the method of Quan et al., the main extraction steps are drying, filtration, methanol extraction for 3 hours, filtration, evaporation, pH adjustment, and chloroform extraction for 3 times. , evaporation, methanol reconstitution, filtration

In the method of Sahar et al. (Sahar et al., 2015), the main extraction steps are: methanol extraction for 24 hours, concentration, sulfuric acid acidification, chloroform extraction, evaporation, reconstitution, and filtration. These cumbersome steps are all alkaloid substances. Quick Quantitative Analysis Brings Inconvenience

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