Detection Methods Study Of Anabolic Androgenic Steroids In Dietary Supplements By Gas Chromatography-tandem Mass Spectrometry

Aims: In this thesis,anabolic androgenic steroids(AAS),which have the highest positive rate and the most severe penalties in doping control,were the research objects.Gas chromatography-tandem mass spectrometry(GC / MS / MS)was used to establish an AAS detection method in dietary supplements.A series of detection methods were established by selecting different pretreatment according to the matrices of dietary supplement and developing the detective windows of target drugs.These established detection methods were also validated by methodology validation and ready for the application in the detection of the AAS in dietary supplements in doping control laboratory.Contents: 1.Selected the suitable ion pairs for acquisition and record the energy voltage(e V)of each ion channel to optimize the mass spectrometry conditions,and establish the significant and applicable target drugs screening windows.2.The pretreatment method of oil matrix samples is a difficult issue for the detection of AAS in dietary supplements.This thesis also improved the pretreatment scheme of this type of matrix based on the large number of literature research.By comparing the feasibility of different pretreatment schemes,the recovery rate of the target drug,and the simplicity of the experimental operation,a sample pretreatment method with better results and suitable for routine laboratory testing.3.Methodological verification of pretreatment methods for different matrix samples using established detection protocols,The index included limit of detection(LOD),matrix effect,recovery rate,degree of precision et al.4.In this thesis,the pyrolysis characteristics of 120 AAS(Among them,8 types of AAS A ring are introduced by benzene ring or C-3 without hydroxyl group and carbonyl group,and the fragmentation of mass spectrometry is irregular.Therefore,only 112 AAS are included in the analysis of mass spectrometry)derived standards in GC/MS/MS were analyzed.Based on the giving information,summarized the generation pathways of AAS significant fragment ions,the cleave modes of AAS with different structural characteristics,and the relationship between AAS structure and retention time.Results: 1.The establishment of 120 kinds of AAS screening windows can meet the detection of positively added mixed standards.2.To remove the oil of the oil matrix samples,samples were extracted by ultrasonic with methanol,then vibrated and centrifuged by added with Sodium hydroxide and Magnesium chloride,and disposal the sediment,this method can detect 116 AAS drugs.Methodological validation of selected sample preparation protocols.Non-oil matrix samples use n-hexane and tert-butyl methyl ether as the pretreatment method of the mixed extraction solution,which can detect 118 AAS drugs.This method is simple to operate and has strong feasibility in routine experiments.3.The minimum detection limit is 0.1 ～ 0.8 ng / m L for liquid samples,0.2 ～ 4.0 ng / g for solid samples,and0.5 ～ 20.0 ng / g for oil samples;recovery rate is between 45.4% ～ 108.0% for liquid samples,43.8% ～ 139.7% for solid samples,5.4% ～ 137.6% for oil samples;The matrix effect is 44.8% ～ 203.4% for liquid samples,40.4% ～ 171.3% for solid sample,45.7% ～ 204.5%for oil sample;the intra-day and inter-day precision of the solid and liquid matrix sample pretreatment methods are less than 20%(Because time is not allowed for oil matrix samples,precision measurement has not been completed).Pretreatment solutions for different substrates meet the requirements for qualitative testing in methodological validation.4.After derivatization of the 17-carbonyl steroid,the steroid D ring can be broken,which can generate m / z 129 fragment ions;the steroid parent nucleus C-4,6,7,11,16 introduces hydroxyl,carbonyl or methyl groups,after derivatization Fragmentation of m / z147 fragments can be generated from ring B;17-hydroxy steroids can also generate ion fragmentation of m / z 169 after fragmentation from steroid D ring;functional groups linked to AAS C-5,C-7,C-17 The retention times of the spatial isomers are all in the alpha position.;The retention times of the spatial isomers of the functional groups attached to AAS C-5,C-7,and C-17 are all alpha-positioned drugs.Conclusion: In this study,non-oil and oil matrix before processing scheme capable of 120 kinds of requirements AAS vast majority of drugs monitoring window(118 kinds of non-oil matrix,116 kinds of oil matrix)to achieve qualitatively detected.Mass chromatographic behavior helps to detect analogs of unknown structure in sample detection,and provides a basis for inferring the structure of unknown analogs.