Method for detection of novel testosterone agonist on the basis of etiocholanolone in urine
A technology of the present cholanolone and stimulant, which is applied in the field of medical detection and can solve the problems of the diversity and complexity of urine sample components, affecting the detection results and the like
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Embodiment 1
[0028] 1. Instruments and equipment
[0029] 1 / 100,000 electronic balance: Satorious company, Switzerland;
[0030] Centrifuge: LD5-2A, Beijing Medical Centrifuge Factory;
[0031] Nitrogen blowing head: Wali Medical Instrument Factory, Chaoyang District, Beijing;
[0032] Fast vortex mixer: Shenzhen Guohua Company;
[0033] Ultrapure water preparation instrument: Milli-Q, MILLIPORE Company;
[0034] Gas chromatography / mass spectrometry (GC / MS): HP6890 / 5973, Agilent Technologies;
[0035] High performance liquid chromatography (HPLC): Waters 2796; Automatic collector: Waters Fraction Collector III, Waters company;
[0036]Gas Chromatography / Oven / Isotope Ratio Mass Spectrometry (GC / C / IRMS): DELTA V, Thermo Scientific.
[0037] 2. Reagents
[0038] Phosphate buffer: in 0.2 mol / l potassium dihydrogen phosphate aqueous solution, gradually add 0.2 mol / l dipotassium hydrogen phosphate aqueous solution, and check the pH value with a pH meter until pH=7.0.
[0039] β-glucuronid...
Embodiment 2
[0058] Embodiment 2 investigates parameters
[0059] The research on the detection method shows that the operation of preparing A-nor-5β-androstane-2,17-dione from the cholanolone extracted from the urine sample is a key step in the detection. Minor changes in parameters such as the ratio of oxidants, the nature and dosage of reagents, and temperature have a great influence on the detection effect. When studying and determining the above operating conditions, the following parameters were selected for investigation:
[0060] 1. Investigate the composition of the oxidizing agent
[0061] The equipment, reagents, instrument operating conditions and methods were the same as those in Example 1, except that 15 μL of glacial acetic acid and 50 μL of oxidants of different compositions were added and kept at 60° C. for 2 hours. The specific composition of the oxidants is shown in Table 1.
[0062] The investigation results show that the change of oxidant composition has a great infl...
Embodiment 3
[0086] manufacture the described 13 The compound of C-labeled testosterone reference substance is Testosterone- 13 C[(17β)-17-Hydroxyandrost-4-en-3-one-13C, T-3- 13 C], available from Toronto Research Chemicals Inc.
[0087] mixed with 13 Preparation of new testosterone materials for C-labeled testosterone reference substance: The raw materials used for the preparation of new testosterone are medicinal testosterone and reference substance T-3- 13 c. Medicinal testosterone was purchased from Shanghai Pharmaceutical Factory, and the reference substance T-3- 13 C, measure the δ value after mixing, until the δ value is -20.6‰, fill the new testosterone material into capsules, and each capsule has 80 mg.
[0088] The 10 subjects in this experiment are male healthy volunteers. It was determined that the hormone metabolism in the subject was normal, and the steroid hormone content was normal.
[0089] The subjects were randomly divided into two groups: 5 subjects were the plac...
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