Quantification of Scopoletin from the Roots of Argyreia Speciosa (Linn. F) Sweet Using HPLC Through the Concept of Design of Experiment.

Abstract

The roots of Argyreia speciosa (Linn. F) Sweet (family: Convolvulaceae) are used in Ayurveda to treat male reproductive and nervous system disorders. Isolation of scopoletin from the roots of Argyreia speciosa, and development and validation of an analytical method using HPLC for the quantification of scopoletin from the root powder of Argyreia speciosa. Scopoletin was isolated from chloroform fraction prepared from hydrolyzed methanolic extract and identified using spectral studies. A reverse-phase HPLC-based analytical method was developed and optimized using the Design of Experiment (DoE) approach to estimate scopoletin from the roots of Argyreia speciosa. Scopoletin was separated and quantified using HPLC containing the C18 column and a PDA detector. The optimized mobile phase was methanol: water (pH∼3.2) [25: 75, %v/v]. The Box-Behnken design was used to optimize chromatographic parameters and the extraction procedure. The validation studies showed a linear relationship (r2=0.998) in the range of 1-40 µg/mL. The LOD and LOQ were found to be 0.28 µg/mL and 0.84 µg/mL, respectively, and the recovery values were found to be between 91.94 and 97.86%. The developed analytical method was found to be robust as well. The amount of scopoletin was estimated to be 0.024 ± 0.0016%w/w from dried root powder. The recorded chromatogram and amount of scopoletin determined would serve as one of the standardization parameters to access the quality of raw material containing Argyreia speciosa. Developed analytical method may be adopted as a part of the standardization procedure for Argyreia speciosa in the quality control laboratory.