Turpinia arguta is an excellent medicinal plant mainly used for the treatment of pharyngitis, tonsillitis, and tonsillar abscesses. However, an efficient regeneration protocol using tissue cultures for T. arguta does not exist. Its main medicinal constituents are flavonoids, particularly ligustroflavone and rhoifolin. Here, we aimed to establish a tissue culture system for T. arguta for the first time using annual stem segments with axillary buds harvested from the field of the Jiangxi Academy of Forestry as explants by dynamically determining the accumulation of effective functional components in the tissue culture plantlets. Orthogonal tests were conducted to compare the effects of different explant disinfection times, media, and exogenous hormone ratios on the induction of the axillary bud growth, successional proliferation, and rooting of T. arguta stem segments. The best explant disinfection effect was achieved by disinfecting the T. arguta explant with 75% ethanol for 50 s, followed by 0.1% mercuric chloride (HgCl₂) for 6 min, and the optimal media for successional proliferation and rooting were Murashige and Skoog (MS) + 0.2 mg/L of 6-benzyladenine (6-BA), + 0.03 mg/L of naphthaleneacetic acid (NAA), and ½ MS + 2.5 mg/L of indole-3-butyric acid + 0.5 mg/L of NAA, respectively. The detection of ligustroflavone and rhoifolin in tissue culture plantlets 0, 3, and 5 months after transplanting showed a significant increasing trend and eventually exceeded the content requirements of the 2020 Edition ofChinese Pharmacopoeia for T. arguta. Our findings provide, for the first time, an effective tissue culture system for T. arguta, thereby providing important information to support the germplasm preservation, innovation, and application of T. arguta in the future.

The dry leaf of Turpinia arguta (Lindl.) Seem. is used in traditional Chinese medicine as a quick-acting product for sore throat and pharyngitis relief. Samples of T. arguta were collected from 40 different habitats mostly located in Jiangxi Province, and leaf appearance traits and dry matter yield were analyzed. HPLC was used to quantify ligustroflavone and rhoifolin, the pharmacological-quality markers for this species, according to the 2020 Edition of Chinese Pharmacopoeia. The correlations between leaf-measurable traits, ligustroflavone and rhoifolin contents, and climate factors were subsequently assessed using Pearson’s two-tailed correlation test and redundancy analysis. The highest dry matter yields were found in locations S(G-mt), Q(J-t), and A(X-t). Ligustroflavone and rhoifolin contents ranged from 0.023% to 1.336% and 0.008% to 0.962%, respectively; the highest levels of ligustroflavone and rhoifolin compounds were found in locations A(X-t) and Y(B-mt). Leaf morphology was influenced by the mean temperature of the warmest quarter, while leaf thickness was affected by the minimum temperature of the coldest month, precipitation seasonality, and solar radiation. Larger and thicker leaves predicted higher yields of ligustroflavone and rhoifolin compounds in T. arguta under Chinese southern conditions, such as those in Anyuan and Quannan counties. Our findings suggest that enhancing the mean diurnal temperature range and implementing appropriate shading during cultivation can enhance the ligustroflavone and rhoifolin compounds of T. arguta.

The antibiotic resistance crisis has become a major threat to global public health. Discovering natural antibacterial compounds with unique mechanisms from traditional medicinal plants is an effective strategy to overcome this challenge. Through activity-guided fractionation, three compounds were isolated from Turpinia arguta leaves, identified as piperyamine A, cynaroside, and gallic acid. Cynaroside exhibited the strongest antibacterial activity, with MIC values of 31.25 μg/mL against Staphylococcus aureus (Gram-positive) and 62.5 μg/mL against Vibrio parahaemolyticus (Gram-negative). Phenotypic experiments, including scanning electron microscopy, electrical conductivity measurements, and alkaline phosphatase (ALP) activity assays, were performed to evaluate antibacterial effects of cynaroside. The results indicated that cynaroside exerted antibacterial effects by disrupting the integrity of bacterial cell walls and cell membranes, with markedly different responses between Gram-positive and Gram-negative bacteria. Specifically, the increase in electrical conductivity was more pronounced in V. parahaemolyticus (Gram-negative), while the peak ALP activity was higher in S. aureus (Gram-positive). Integrated metabolomic and transcriptomic analyses were conducted to elucidate the differential antibacterial mechanisms. In S. aureus , cynaroside treatment was associated with suppression of pyrimidine metabolism and histidine metabolism, negatively regulating 11 metabolites with pyrC as the hub gene; in V. parahaemolyticus , it mainly inhibited glyoxylate and dicarboxylate metabolism and branched-chain amino acid degradation, negatively regulating tricarboxylic acid cycle intermediates with fdh3B as the hub gene. This study reveals the differential antibacterial mechanisms of cynaroside isolated from Turpinia arguta against Gram-positive and Gram-negative bacteria, laying a theoretical foundation for the development of species-selective natural antibacterial agents.