A Validated LC-MS/MS Method for the Quantification of Trigonelline in Marketed Dietary Supplements

Background: Fenugreek seeds are employed in many traditional systems as an antibacterial, antidiabetic agent, gastric stimulant, and also for anti-invasive activity. Therefore, it is a suitable bioactive marker to establish the quality of crude drug and its formulations. Methods: A rapid, simple and sensitive LC-MS/MS analytical method was developed and validated for the determination of trigonelline extracted from Trigonella foenum-graecum (L.) (Fenugreek) and marketed dietary supplements using Etofylline as an internal standard. The objective of the present study is to quantify Trigonelline extracted from Trigonella foenum graecum L. (fenugreek) and marketed dietary supplements. Chromatographic separation was achieved on a Zorbax C18 column (50mm x 4.6mm i.d, 5μ particle size). The samples were eluted using 0.1% Formic acid in water: Methanol (20:80%v/v) at a flow rate of 0.5ml/min with a runtime of 5 min. The eluents were monitored using a tandem mass spectrometer equipped with an electro spray ionization source in positive mode. Results: The analysis was performed in multiple reaction monitoring (MRM) mode by quantifying the ion transitions from m/z 138.0→92.5 (Trigonelline) and m/z 225.0→180.90 (IS). The developed method was linear over the concentration range 5-50 ng/mL. The LOD and LOQ were found to be 1.0 ng/mL and 10.0 ng/mL, respectively. The correlation coefficient (r2) was found to be ≥0.998 for Trigonelline. Conclusion: The proposed validated LC-MS/MS method offers a sensitive quantification of trigonelline in Trigonella foenum graecum L. (fenugreek) and marketed dietary supplements containing fenugreek seeds.