Box–Behnken Design Based Optimisation of HPTLC Method for Analysis of Jatrorrhizine from Tinospora cordifolia Roots with Antidiabetic Activity Assessment

Introduction: Tinospora cordifolia root (T. cordifolia) is a well-known Ayurvedic herb with immune-boosting, anti-inflammatory, and antioxidant properties. It contains Jatrorrhizine (JZ), a bioactive alkaloid with antimicrobial, hepatoprotective, and metabolic health benefits, similar to berberine. Objectives: The study aimed to evaluate the JZ content in T. cordifolia root obtained via different extraction method and to optimise the High-Performance Thin Layer Chromatography (HPTLC) method via Response Surface Methodology (RSM). The extracts were assessed for in vitro test evaluation. Materials and Methods: A Box-Behnken Design (BBD) of RSM, a mathematical and statistical method, was applied to optimize the impact of independent variable on the response. Three independent variables like the different concentration of the mobile phase were studied. The in vitro analysis was also done. Results: The content of JZ was quantified using the HPTLC densiometric method. HPTLC confirmed the presence of JZ with a prominent band at an Rf value of 0.70±0.008 using a mobile phase of chloroform: toluene: methanol: formic acid (7:4:2:0.2 v/v). The in vitro analysis of the extract showed a significant result and was found to be α-amylase in TRM 27.71±0.70 and α-glucosidase in TRM 43.45±1.31. Conclusion: This work examines various extraction procedures, including Soxhlet, maceration and reflux, and introduces a novel methodology to enhance extraction efficiency, advance the phytochemical analysis of JZ. This suggests that T. cordifolia may have therapeutic potential in diabetes management.