Development and validation of an HPLC method for quantitative analysis of trigonelline

Abstract

Trigonelline is an alkaloid primarily found in fenugreek (Trigonella foenum-graecum) seeds, exhibiting diverse pharmacological activities, including anti-diabetic, antioxidant, and anti-inflammatory effects. With the growing application of trigonelline in drug development and quality control, the development of efficient and accurate analytical methods is critical to ensure its proper assessment and utilization. This study aims to develop and validate a HPLC method for the precise separation and quantitative determination of trigonelline. The method utilizes a Dalian Elite Hypersil NH₂ chromatographic column (250 mm × 4.6 mm, 5 µm) with a mobile phase of acetonitrile: water (70:30, v/v), a flow rate of 1.0 mL/min, column temperature maintained at 35 °C, and detection wavelength set at 264 nm. The method was applied to analyze fenugreek seed extracts prepared via ultrasonic extraction with methanol, employing a 30-minute extraction time. The results demonstrated that the method achieved an excellent linear range (R² > 0.9999), high precision (RSD < 2 %), system suitability, and recovery rate is between 95 % and 105 %, meeting the quality standards for trigonelline analysis. In conclusion, this study introduces a reliable and efficient HPLC method, offering robust technical support for the quantitative analysis of trigonelline and the quality evaluation of related pharmaceutical products, facilitating its application in clinical drug monitoring and standardization.