Capillary-Based Electrophoresis Method for the Quantification of Stachyose in Stachys affinis Bunge (Lamiaceae) Tubers
Ngoc Thi Bich Nguyen *
Faculty of Pharmacy, Nguyen Tat Thanh University, Ho Chi Minh City, Vietnam.
*Author to whom correspondence should be addressed.
Abstract
Aims: Stachyose, a tetrasaccharide comprising approximately 80% of vacuolar carbohydrates, is a characteristic marker of the genus Stachys and is reported to exhibit various pharmacological benefits, including blood glucose reduction, immune enhancement, and constipation relief. This study aimed to develop and validate a CE-DAD method for stachyose quantification in Stachys affinis tubers in accordance with ICH (2005) and AOAC (2012) guidelines to support raw material quality control.
Study Design: Experimental laboratory-based method development and validation study.Place and Duration of Study: Stachys affinis tubers were provided by the City Biotechnology Center, Ho Chi Minh City, Vietnam. The study was conducted at Ho Chi Minh City University of Pharmacy from June 2021 to July 2022.
Methodology: A CE-DAD method was established for stachyose quantification in Stachys affinis rhizomes. The capillary column was pretreated and equilibrated using a standardized NaOH–water–buffer sequence, with daily and inter-run conditioning to ensure consistent performance. Electrophoresis parameters—including BGE type, pH, Na₂HPO₄ concentration, temperature, injection time, and voltage—were optimized using JMP10 Pro and a 15-run experimental design. Method validation followed ICH and AOAC guidelines, evaluating system suitability, specificity, linearity, sensitivity (LOD, LOQ), precision, and recovery.
Results: The optimized procedure employed a CE-7100 Agilent system with a bare-silica capillary column (effective length 65 cm), 35 mM Na₂HPO₄ (pH 12.4) as BGE, a voltage of 8 kV, a column temperature of 30°C, sample injection at 50 mbar for 3 s, and detection at 270 nm using lactose as the internal standard.
Conclusion: The CE-DAD method for stachyose quantification in Stachys affinis was validated according to ICH and AOAC (2012) guidelines, confirming its suitability for raw material quality control. The method showed system suitability, high selectivity, wide linearity (4.46–89.18 µg/mL), and high sensitivity (LOD 5.00 µg/mL; LOQ 15.00 µg/mL). It also demonstrated good repeatability and intermediate precision (RSD 1.77% [n=6] and 1.94% [n=12]) with recovery ranging from 99.01% to 102.10%.
Keywords: Stachys affinis Bunge, stachyose, internal standard, capillary electrophoresis