Determination of Puberulic Acid Content in Monascus by Pre-column Derivatization-liquid Chromatography with Tandem Mass Spectrometry

OBJECTIVE  To establish a highly sensitive detection method for puberulic acid in monascus by using trimethylsilylated diazomethane for methylation derivation of puberulic acid.

METHODS  High performance liquid chromatography triple quadrupole mass spectrometry was used to detect the methylated derivatives of two isomers of puberulic acid. The chromatographic column was Phenomenex C₈(2.1 mm×100 mm, 2.7 μm), and the mobile phase A was 10 mmol·L⁻¹ ammonium formate, mobilephase B was 50% methanol in water(containing 10 mmol·L⁻¹ ammonium formate), gradient elution(0–0.5 min, 90%→10%A; 0.5–4 min, 10%A), flow rate was 0.3 mL·min⁻¹, column temperature was 40 ℃, injection volume was 10 μL. The ionization mode was ESI⁺, and multiple reaction monitoring was carried out. The methylated derivative of puberulic acid m/z 255.0→225.0 was selected as the quantitative ion, and the mass to charge ratio m/z 255.0→194.0 was selected as the qualitative ion.

RESULTS  Nine batches of monascus samples were tested, and no puberulic acid was detected, with a pass rate of 100%.

CONCLUSION  The established method has been validated and has strong specificity, and can be used for the detection of puberulic acid in monascus.