Procedures for determination of melengestrol acetate, chlorgestrol acetate and megestrol acetate residues in milk and milk powder

8.
2.
2.
6 Determination

(1) Liquid chromatography conditions

Chromatographic column: C ₁₈ column, 3μm, 50mm×2.
0mm (inner diameter) or equivalent; column temperature: 30°C; flow rate: 0.
3mL/min; injection volume: 10μL; gradient elution, elution program see Table 8- 14
.

Table 8-14 Liquid chromatography gradient elution program

(2) Mass spectrometry conditions

Ion source: positive ion electrospray; scanning method: multi-reaction detection MRM; electrospray voltage: 4000V; ion source temperature: 350℃; atomizing gas, curtain gas, auxiliary gas: all high-purity nitrogen, adjust each airflow before use Flow rate, so that the sensitivity of the mass spectrometer meets the detection requirements; qualitative ion pair, quantitative ion pair, declustering voltage and collision energy are shown in Table 8-15
.

Table 8-15 Qualitative ion pair, quantitative ion pair, declustering voltage and collision energy

(3) Qualitative determination

Choose one parent ion and two or more product ions for each component to be tested.
Under the same experimental conditions, the deviation of the retention time of the substance to be measured in the sample and the retention time of the corresponding substance in the mixed matrix standard solution is within ±2.
5% ; And the relative abundance of each qualifier ion in the sample spectrum is compared with the relative abundance of the corresponding qualifier ion in the mixed matrix standard solution spectrum with close concentrations.
If the deviation does not exceed the range specified in Table 1-5, it can be judged There is a corresponding analyte in the sample
.

(4) Quantitative determination

Under the best working conditions of the instrument, use the mixed matrix standard calibration solution to inject samples separately, take the peak area as the ordinate and the concentration of the mixed matrix standard calibration solution as the abscissa, draw a standard working curve, and 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
.

Figure 8-9 Multiple reaction monitoring (MRM) chromatograms of melengestrol acetate, chlorgestrol acetate and megestrol acetate standard substances

Related links: Sample pretreatment for determination of melengestrol acetate, chlorgestrol acetate and megestrol acetate residues in milk and milk powder