Simultaneous Estimation of Rhein and Aloe-Emodin in Traditional and Ultrasound-Based Extracts of Rheum palmatum L. (Rhubarb) Using Sustainable Reverse-Phase and Conventional Normal-Phase HPTLC Methods

The greenness indices of literature analytical procedures for the simultaneous measurement of rhein and aloe-emodin have not been determined. As a consequence, the first goal of this study was to design and validate a sensitive and sustainable reverse-phase high-performance thin-layer chromatography (HPTLC) method for the simultaneous estimation of rhein and aloe-emodin in a traditional extract (TE) and ultrasound-based extract (UBE) of commercial Rhubarb and Rhubarb plant extracts in comparison to the conventional normal-phase HPTLC method. The second goal was to determine the greenness indices for both methods using the AGREE approach. For the sustainable reverse-phase HPTLC approach, the method was linear in the 50–1000 ng/spot range for rhein and 25–1000 ng/spot range for aloe-emodin. However, for the conventional normal-phase HPTLC approach, the method was linear in the 50–600 ng/spot range for rhein and 100–600 ng/spot range for aloe-emodin. The limit of detection (LOD) for rhein and aloe-emodin was 16.81 ng/spot and 8.49 ng/spot, respectively, using the sustainable analytical method. However, the LOD for rhein and aloe-emodin was 18.53 ng/spot and 39.42 ng/spot, respectively, using the conventional analytical method. For the simultaneous determination of rhein and aloe-emodin, the sustainable analytical method was more sensitive, accurate, precise, and robust than the conventional analytical method. The amount of rhein and aloe-emodin was higher in the UBE of commercial Rhubarb and Rhubarb plant extract over their TE. For the simultaneous quantification of rhein and aloe-emodin in the TE and UBE of marketed Rhubarb and Rhubarb plant extract, the sustainable analytical method was superior to the conventional analytical method. The AGREE index for the sustainable reverse-phase and conventional normal-phase HPTLC methods was determined to be 0.78 and 0.49, respectively, indicating an excellent greenness profile of the sustainable reverse-phase HPTLC method over the conventional normal-phase HPTLC approach. The sustainable analytical method was found to be superior to the conventional analytical method based on these results.