| Objectives:To establish a method for qualitative and quantitative analysis of 14 kinds of aconitine alkaloids and their metabolites in Aconitum L.by ultra performance liquid chromatography/quadrupole time-of-flight mass spectrometric(UPLC/Q-TOF),which is conducive to rapid screening of components in poisoning events.The alkaloids in Aconitum L.of Dali and Honghe are qualitatively and quantitatively analyzed by this method,and the toxicity is evaluated.It is helpful to improve the comprehensive utilization value of this kind of plants.Methods:The samples is extracted and purified by Waters Oasis MCX solid phase extraction cartridge,which is activated by methanol and ultra pure water.Being washed by 0.1 mol/L HCL and methanol,eluted by methanol containing 5%ammonia and collected all,the solution is dried by rotating evaporation and bring to volume by acetonitrile-5 mmol/L ammonium acetate(50:50,V:V).The target compounds are separated by UPLC/Q-TOF with Aglient SB C18 column and eluted by 0.1%formic acid aqueous solution containing 5 mmol/L ammonium acetate-acetonitrile.Data were collected under ESI positive ion mode,and qualitative and quantitative analysis of aconitine alkaloids and their metabolic products in Aconitum L.are carried out.According to the toxicity of LD50,the toxicity of the samples are evaluated.Results:The method for the qualitative and quantitative determination of 14 aconitine alkaloids and their metabolites in Aconitum L.by UPLC/Q-TOF is established.The separation conditions of liquid chromatography and mass spectrometry were determined,and the first and second characteristic spectra of 14 aconitine alkaloids and their metabolites were established.The method has a good linear relationship in the range of 1.0-100 ng/mL for 14 target compounds.The correlation coefficient is greater than 0.99.The detection limit is 0.1-5 ng/mL,the quantitative limit is 0.5-10 ng/mL,the recovery rate is 81%-114%,and the relative standard deviation is 0.5%-3.7%.Qualitative and quantitative analysis of the samples collected by the above methods show that 14 alkaloids are different in different regions and different species of Aconitum.L.Based on LD50 data,the contents of diester alkaloids,monoester alkaloids,aminoalkanol alkaloids and other alkaloids in diterpenoid alkaloids are compared.The toxicity of 15 samples is No.12>No.11>No.2>No.3>No.1>No.14>No.4>No.7>No.8>No.6>No.5>No.9.Detailed information of samples was shown in Table 3.The toxicity of wild Aconitum.L samples was greater than that of standardized planting samples.The 14 alkaloids in fibrous root,tuber,stem,leaf,flower and fruit are also analyzed.The total content of 14 alkaloids in underground part is higher than that in aboveground part.The toxicity of each part of the whole plant is evaluated as fibrous root>tuber root>stem,leaf,flower and fruit.Conclusions:UPLC/Q-TOF method for qualitative and quantitative detection of 14 aconitine alkaloids and their metabolites is simple,efficient,sensitive and accurate.lt can meet the requirements of rapid screening and detection of various Aconitum alkaloids and their metabolites,make up for the blank of simultaneous detection of various alkaloids in Aconitum L..Aconite and Fuzi in Dali and Honghe City of Yunnan Province are highly toxic.The composition of 14 aconitine alkaloids and their metabolites in different Aconitum.L are different,which provides more choices for the supply of raw materials for medicine.Aconitum.L in Dali and Honghe City of Yunnan Province contain toxicity in each parts.The contents of 14 aconitine alkaloids and their metabolites in different parts are different.The contents and toxicity of alkaloids in underground parts are higher than those in aboveground parts.The shoots,leaves,flowers and fruits of the aboveground part have the value of development and utilization,which is beneficial to the comprehensive development and utilization of aconites. |
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