Background: Cassia (Family: Fabaceae) species are a large group of flowering plants rich in bioactive anthraquinone and flavonoids used in botanical supplements and nutraceuticals.
Objective: A simple and reliable high-performance liquid chromatography-photodiode array (HPLC-PDA) method was developed and validated for separating and quantifying thirteen anthraquinone and flavonoids. These compounds were further confirmed using an LC-based electrospray ionization mass spectrometry (ESI-MS/MS) method in the leaves and flowers of selected Cassia species. A simple and rapid HPTLC method was developed for chemical fingerprint analysis of all Cassia species.
Method: All thirteen compounds were chromatographically separated on a Zorbax TC18 (4.6 x 250, 5 μm particle size) analytical column, and 0.1% formic acid and acetonitrile as elution solvents at a flow rate of 0.8 mL/min with detection at 259 nm. For HPTLC fingerprinting, the mobile phase compositions of toluene, ethyl acetate, and formic acid (5.5:4.2:0.6, v/v/v) were optimized to separate and identify all compounds using silica gel 60F254 aluminum plates.
Results: The validation data for the developed HPLC-PDA method for thirteen compounds showed good linearity (r2 > 0.99) with a sensitive LOD (0.082-1.969 mg/mL), LOQ (0.250-5.967 mg/mL), and excellent recoveries (85.22-100.32%). The quantification results were found to be precise and accurate (<5.0% and relative error), -0.77-0.44 with ESI-MS/MS confirmation in the Cassia samples. The novel HPTLC method was excellent separation for thirteen compounds, with Rf values ranging between 0.12- 0.61.
Conclusions: The developed HPLC-PDA method was simple, and precise and could separate and quantify anthraquinones and flavonoids along with confirmation, using a novel LC-based ESI-MS/MS. The HPTLC method was found to be simple and precise, with excellent separation capabilities for these compounds.
Highlights: This novel multiplatform approach successfully identified and quantified thirteen compounds simultaneously using an integration of data strategy in seven medicinally important Cassia species' leaves and flowers.
Keywords: Cassia; Senna; Anthraquinones; Flavonoids; HPTLC.
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