1 Department of Pharmacognosy, College of Pharmacy, Najran University, Najran 66454, Saudi Arabia.
2 Department of Chemistry and Biochemistry, Concordia University, 7141 Sherbrooke Street West, Montréal, QC H4B 1R6, Canada.
3 Department of Clinical Laboratory Sciences, Faculty of Applied Medical Sciences, Najran University, Najran 66454, Saudi Arabia.
4 Department of Pharmacology, College of Pharmacy, Najran University, Najran 64462, Saudi Arabia.
5 Meikai University Research Institute of Odontology (M-RIO), 1-1 Keyakidai, Saitama 350-0283, Japan.
6 Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Tsushima, Okayama 700-8530, Japan.

Polyphenols have a variety of phenolic hydroxyl and carbonyl functionalities that enable them to scavenge many oxidants, thereby preserving the human redox balance and preventing a number of oxidative stress-related chronic degenerative diseases. In our ongoing investigation of polyphenol-rich plants in search of novel molecules, we resumed the investigation of Lawsonia inermis L. (Lythraceae) or henna, a popular ancient plant with aesthetic and therapeutic benefits. The leaves' 70% aq acetone extract was fractionated on a Diaion HP-20 column with different ratios of H₂O/an organic solvent. Multistep gel chromatographic fractionation and HPLC purification of the Diaion 75% aq MeOH and MeOH fractions led to a new compound (1) along with tannin-related metabolites, benzoic acid (2), benzyl 6'-O-galloyl-ß-D-glucopyranoside (3), and ellagic acid (4), which are first isolated from henna. Repeating the procedures on the Diaion 50% aq MeOH eluate led to the first-time isolation of two O-glucosidic ellagitannins, heterophylliin A (5), and gemin D (6), in addition to four known C-glycosidic ellagitannins, lythracin D (7), pedunculagin (8), flosin B (9), and lagerstroemin (10). The compound structures were determined through intensive spectroscopic investigations, including HRESIMS, 1D (¹H and ¹³C) and 2D (¹H-¹H COSY, HSQC, HMBC, and NOESY) NMR, UV, [a]_D, and CD experiments. The new structure of 1 was determined to be a megastigmane glucoside gallate; its biosynthesis from gallic acid and a ß-ionone, a degradative product of the common metabolite ß-carotin, was highlighted. Cytotoxicity investigations of the abundant ellagitannins revealed that lythracin D2 (7) and pedunculagin (8) are obviously more cytotoxic (tumor specificity = 2.3 and 2.8, respectively) toward oral squamous cell carcinoma cell lines (HSC-2, HSC-4, and Ca9-22) than normal human oral cells (HGF, HPC, and HPLF). In summary, Lawsonia inermis is a rich source of anti-oral cancer ellagitannins. Also, the several discovered polyphenolics highlighted here emphasize the numerous biological benefits of henna and encourage further clinical studies to profit from their antioxidant properties against oxidative stress-related disorders.