To control the quality of Betula alnoides stem bark (BAS), lupeol, which is a promising compound found in BAS, has been determined by reverse phase high performance liquid chromatography (RP-HPLC) and thin-layer chromatography (TLC)-densitometry. These methods were developed and validated according to the International Council for Harmonization (ICH) and the Association of Official Agricultural Chemists (AOAC) guidelines. RP-HPLC was achieved within 10 min using C8 column under an isocratic elution of acetonitrile at a flow rate of 1.0 mL/min. Lupeol was detected at a UV wavelength of 200 nm. TLC-densitometry performed on TLC silica gel plates was developed by chloroform at 10 cm of distance. Lupeol was detected under visible wavelength at 525 nm after anisaldehyde-sulfuric acid derivatization. Both developed methods exhibited good linearity, specificity, limits of detection and quantitation, accuracy and precision, and robustness. The validated methods were effectively applied to determine lupeol content in BAS collected from three different locations. The amount of lupeol was found to be about 50–100 µg/g of dried powder. The results revealed no statistically significant differences between the two methods. These findings revealed the validated RP-HPLC and TLC-densitometry that were easily applied for routine quality control of BAS for the first time. Graphical Abstract

White Kwao Krua, a crucial Thai medicinal plant, contains various phytoestrogen substances used to alleviate menopausal symptoms in estrogen-deficient women. It originates from two species, namely Pueraria candollei Wall. ex Benth. and P. mirifica Airy Shaw & Suvat. However, there exists morphological variation, and the taxonomic status between both species is ambiguous, making discrimination challenging. In this study, we aim to clarify and differentiate the morphological characteristics, palynology, and DNA barcoding of both species. The morphological results showed the stipule size is a phenotypic marker for the differentiation of both species during the vegetative stage. The palynological results, however, exhibited similarity. Through an examination of nucleotide sequences and neighbor-joining tree analysis, it was determined that the DNA barcoding of the matK region has the capability to distinguish between P. candollei and P. mirifica at nucleotide position 702. Specifically, P. candollei manifested a G base, contrasting with the C base observed in P. mirifica. This study concludes that stipule size and the matK gene in DNA barcoding serve as a distinctive characteristic for distinguishing between P. candollei and P. mirifica. These methodologies prove valuable for ensuring the accurate identification of white Kwao Krua for horticulturists.

Due to the benefits of mangiferin, including antidiabetic, antibacterial, antiviral, antioxidant, and anti-inflammatory activities, it is a natural raw material used in botanical health and cosmetic products. This research aims to develop and validate analytical methods for the quantification of mangiferin in raw material, isolated from Mangifera indica L. leaves, using high performance liquid chromatography (HPLC) and attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR). The greenness assessment of both analytical methods was performed. The results indicated that both methods were validated in terms of specificity, linearity, range, detection limit, quantitation limit, accuracy, and precision. The ranges of mangiferin used in HPLC and ATR-FTIR were 30 to 360 µg/mL and 5.00 to 27.27% w/w, respectively. The mangiferin content in five batches, analyzed by using the validated HPLC and ATR-FTIR, ranged from 88.35 to 109.4% w/w and 90.7 to 111.8% w/w, respectively. Statistical analysis using a paired t-test revealed no significant difference in the mangiferin quantity between the two techniques at a 95% confidence level. The greenness scores of the HPLC and ATR-FTIR determined according to AGREE metric were 0.49 and 0.81, respectively. Both methods can be applied to quantify mangiferin content and control the quality of mangiferin raw material. Additionally, the validated ATR-FTIR, which proposed excellent environmental friendliness, was suggested as an alternative analytical method for routine assay of mangiferin raw material.