Determination of testosterone, epitestosterone and progesterone residues in beef liver and beef

8.
2.
1.
6 Determination

(1) Liquid chromatography conditions

Chromatographic column: C ₈ chromatographic column, 5um, 150mm×2.
1mm (inner diameter); column temperature: 40°C; injection volume: 20 μL; mobile phase, flow rate and gradient elution conditions are shown in Table 8-7
.

Table 8-7 Mobile phase, flow rate and gradient elution conditions

(2) Mass spectrometry conditions

Ion source: electrospray ion source; scanning method: positive ion scanning; detection method: multi-reaction monitoring; atomizing gas, curtain gas, auxiliary heating gas, collision gas are all high-purity nitrogen and other suitable gases; each should be adjusted before use Gas flow rate to make the sensitivity of the mass spectrometer meet the detection requirements; the spray voltage, declustering voltage, collision energy and other voltage values ​​should be optimized to the best sensitivity; qualitative ion pair, quantitative ion pair, declustering voltage and collision energy are shown in Table 8-8
.

Table 8-8 Mass spectrometry parameters of testosterone, epitestosterone, progesterone and internal standards

(3) Qualitative determination

Choose one parent ion and two or more product ions for each tested component.
Under the same experimental conditions, the retention time of the test substance in the sample is within ±2.
5% of the corresponding retention time in the standard solution; and the sample Compare the relative abundance of the qualitative ions of each component in the standard solution with the relative abundance of the corresponding qualitative ions in the standard solution with close concentrations.
If the deviation does not exceed the range specified in Table 1-5, it can be determined that there is a corresponding test in the sample Things
.

(4) Quantitative determination

Under the best working conditions of the instrument, sample the matrix mixed standard working solution, and take the ratio of the peak area of ​​the measured component in the standard solution to the peak area of ​​the internal standard substance as the ordinate, the concentration of the measured component in the standard solution and the internal standard The ratio of the substance concentration is the abscissa to draw a standard working curve.
Use the standard working curve to quantify the sample.
The response value of the analyte in the sample solution should be within the linear range determined by the instrument
.
Quantification by internal standard method

Figure 8-8 Multiple reaction monitoring (MRM) chromatograms of progesterone, testosterone, and epitestosterone reference materials