Evaluation of Tinospora species: T. cordifolia, T. sinensis and T. crispa with microscopic imaging, physicochemical properties and major compounds profiling for authentic uses

The wide geographic distribution, ethno-botanical applications, species identification challenges, and morphological similarities among different species underscore the importance of qualitative analysis in safeguarding botanical purity and ensuring quality control. This study presents a comprehensive characterization of the main commercial plant part, namely the stem, of Tinospora cordifolia, Tinospora sinensis, and Tinospora crispa by considering their physico-chemical characteristics, macroscopic observations, and anatomical details. Through our investigations, we have identified crucial diagnostic features that can aid in distinguishing between different Tinospora species and detecting adulteration in dried raw materials. Notably, T. crispa exhibits more prominent warty protuberances on the stem compared to T. cordifolia and T. sinensis. Furthermore, the transverse section of all three species displays a multilayered cork with distinct radial files and layered cork development. While the heart-shaped leaves of T. cordifolia and T. crispa possess a smooth and pubescent texture respectively. The abaxial surface of T. sinensis leaves is densely coated with silky hairs. To assess the presence of specific bioactive compounds, namely tinosporaside and berberine, we employed high-performance thin-layer chromatography. Our findings reveal that tinosporaside exhibits a band at R-f 0.45 in T. sinensis and T. crispa, whereas all three species display a berberine presence spot with varying intensities at R-f 0.23. These biochemical markers serve as reliable identifiers for species differentiation. Thus, our study establishes a cost-effective approach for enhancing quality control and species identification in Tinospora species, ultimately reducing the occurrence of botanical adulteration.