Integrated transcriptome and metabolome analysis provide insights into the mechanism of saponin biosynthesis and its role in alleviating cadmium-induced oxidative damage in Ophiopogon japonicum.

Zhe-Maidong, a cultivar of Ophiopogon japonicus is a prominent traditional herbal medicine rich in saponins. This study explored the mechanism of saponin biosynthesis and its role in alleviating Cd-induced oxidative damage in the Zhe-Maidong cultivar using three experimental groups undergoing Cd stress. In the Cd-contaminated soil treatment, total saponins were 1.68 times higher than those in the control. The saponin content in the Cd-2 and Cd-3 treatments was approximately twice as high as that in the Cd-CK treatment. These findings revealed that Cd stress leads to total saponin accumulation. Metabolomic analysis identified the accumulated saponins, primarily several monoterpenoids, diterpenoids, and triterpenoids. The increased saponins exhibited an antioxidant ability to prevent the accumulation of Cd-induced reactive oxygen species (ROS). Subsequent saponin application experiments provided strong evidence that saponin played a crucial role in promoting superoxide dismutase (SOD) activity and reducing ROS accumulation. Transcriptome analysis revealed vital genes for saponin synthesis under Cd stress, including SE, two SSs, and six CYP450s, positively correlated with differentially expressed metabolite (DEM) levels in the saponin metabolic pathway. Additionally, the TF-gene regulatory network demonstrated that bHLH1, bHLH3, mTERF, and AUX/IAA transcript factors are crucial regulators of hub genes involved in saponin synthesis. These findings significantly contribute to our understanding of the regulatory network of saponin synthesis and its role in reducing oxidative damage in O. japonicum when exposed to Cd stress.

Nontargeted metabolomic analysis and multiple criteria decision-making method induced robust quality markers screening for the authentication of herbal medicines from different origins by taking Ophiopogon japonicus (L. f.) Ker-Gawl. as a case study.

A key segment in medicinal plant authentication is the establishment of quality markers that embody the intrinsic metabolites difference independent of instruments and experiment conditions. A strategy integrating nontargeted metabolomics and multicriteria decision-making model for robust quality markers discovery is presented and applied to authenticate Ophiopogon japonicus (L. f.) Ker-Gawl. First, an ultra-performance liquid chromatography/quadrupole time-of-flight MSE approach was established for global metabolites profiling and identification. Second, multivariate statistical analysis was performed to explore potential quality markers of different origins of ophiopogonis radix. Third, potential quality markers were ordered and filtered by multicriteria decision-making model to infer robust quality markers and further validated in different instruments and experiment conditions by validation model. Fourth, the validation model using the robust quality markers managed to discriminate the origins of ophiopogonis radix samples procured from the herbal markets. Consequently, two robust quality markers, cixi-ophiopogon B and ophiopogonin D, were discovered and further validated on different instruments and experiment conditions. This integrated strategy provided a practical solution for reliable and convenient authentication of geo-authentic herb.

Structural characterization and discrimination of Ophiopogon japonicas (Liliaceae) from different geographical origins based on metabolite profiling analysis.

The root tuber of Ophiopogon japonicus (Thunb.) Ker-Gawl ("Maidong" in Chinese), with steroidal saponins and homoisoflavonoids as its representative chemical compositions, is a representative medicinal herbs with multiple major producing areas. This study aimed to distinguish the O. japonicas samples from Zhejiang and Sichuan by using an ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF MS)-based metabolome analysis. Firstly, a global chemical constituent identification of O. japonicas was carried out by using both automatic and manual methods. An integrated steroidal saponins structural identification strategy in O. japonicas based on exact mass information, fragmentation characteristics and retention time was developed. Overall, 135 steroidal saponins, 47 homoisoflavonoids and 9 other metabolites were quickly identified or tentatively identified from the MSE continuum data. Furthermore, multivariate statistical analysis revealed that O. japonicas from Zhejiang and Sichuan can clearly be separated and some markers were screened. Moreover, some major active components including total soluble sugar, total soluble polyphenol, total flavonoid, total saponin and 10 specific compounds were analyzed quantitatively. In general, these results showed that there were many differences between the metabolic profile data of O. japonicas from different producing areas, O. japonicas from Sichuan showed higher level steroidal saponins and samples from Zhejiang had higher contents of homoisoflavonoids specifically, and indicated that metabolite profiling by UPLC/Q-TOF MS is an effective approach for the discrimination of medicinal herbs from different geographical origins.

Metabolic Profiling of Saponin-Rich Ophiopogon japonicus Roots Based on 1H NMR and HPTLC Platforms.

Ideally, metabolomics should deal with all the metabolites that are found within cells and biological systems. The most common technologies for metabolomics include mass spectrometry, and in most cases, hyphenated to chromatographic separations (liquid chromatography- or gas chromatography-mass spectrometry) and nuclear magnetic resonance spectroscopy. However, limitations such as low sensitivity and highly congested spectra in nuclear magnetic resonance spectroscopy and relatively low signal reproducibility in mass spectrometry impede the progression of these techniques from being universal metabolomics tools. These disadvantages are more notorious in studies of certain plant secondary metabolites, such as saponins, which are difficult to analyse, but have a great biological importance in organisms. In this study, high-performance thin-layer chromatography was used as a supplementary tool for metabolomics. A method consisting of coupling 1H nuclear magnetic resonance spectroscopy and high-performance thin-layer chromatography was applied to distinguish between Ophiopogon japonicus roots that were collected from two growth locations and were of different ages. The results allowed the root samples from the two growth locations to be clearly distinguished. The difficulties encountered in the identification of the marker compounds by 1H nuclear magnetic resonance spectroscopy was overcome using high-performance thin-layer chromatography to separate and isolate the compounds. The saponins, ophiojaponin C or ophiopogonin D, were found to be marker metabolites in the root samples and proved to be greatly influenced by plant growth location, but barely by age variation. The procedure used in this study is fully described with the purpose of making a valuable contribution to the quality control of saponin-rich herbal drugs using high-performance thin-layer chromatography as a supplementary analytical tool for metabolomics research.

Migration and transformation of different phosphorus forms in rainfall runoff in bioretention system.

Artificial bioretention system consisting of Ophiopogon japonicus infiltration medium was used to simulate an infiltration experiment of rainfall runoff. Continuous extraction method was used to detect contents of inorganic phosphorus (P) under exchangeable state (Ex-P) and aluminium phosphate (Al-P) and iron phosphate (Fe-P) at different depths (0, 5, 15 and 35 cm) of soil infiltration medium in bioretention system. Effluent total P (TP) concentration of the system was also monitored. Results indicated that the adsorption of inorganic P, Al-P and Fe-P by soil infiltration medium was implemented layer by layer from top to bottom and gradually weakened. Moreover, Ex-P was gradually transformed into Al-P and Fe-P, whereas Al-P was gradually transformed into Fe-P; thus, Ex-P content reduced layer by layer, whereas Al-P and Fe-P gradually accumulated. The TP removal rate in runoff rainwater by the system was more than 90%, where the TP that was not used by plants was under dynamic equilibrium in water-soil-root system/biological system.

[Influence of harvest times and processing methods on contents of total flavonoids and total saponins in roots of Ophiopogon japonicus].

OBJECTIVE: To study the patterns of dynamic accumulation of total flavonoids and total saponins in the roots of Ophiopogon japonicus collected from different harvest times, and compare the contents of total flavonoids and total saponins in roots of O. japonicus which were processed with different methods. METHOD: The total flavonoids and total saponins contents in O. japonicus were determined by ultraviolet spectrophotometry. RESULT AND CONCLUSION: From December to January, the total contents of flavonoids and saponins in the roots of O. japonicus gradually decreased, and gradually increased from February to March, and kept stable in April. The contents of total flavonoids and total saponins in the O. japonicus were influenced by different processing methods.

[Study on N, P and K absorption and accumulation in Ophiopogon japonicus].

OBJECTIVE: To study the absorption and accumulation of N, P and K in Ophiopgon japonicus. METHOD: The contents of N, P, K and trace element were determined respectively by using the kjeldathl method digested with H2SO4 + H2O2, ammonium vanadstemolybdate chromoscopy, flame spectrometry and atomic absorption spectrophotometry. RESULT: The contents of N and K were the highest, and that of P was relatively lower O. japonicus. The changing pattern of N was low-high-low in the growth and development period. The accumulative speed of N had two relatively quicker and two relatively slower phases. The content of N is higher in nutritive organs; The changing pattern of P was similar to that of N, while the content of P is higher in leaves and nutritive roots at early stage of growth and development. It was high in root tuber during harvesting stage, and the accumulative quantity in the plant was increased gradually. The level of K was high in early stage and low in later stage of growth. The content of K was relatively higher in leaves and nutritive roots at early stage, and that in root tuber was slightly lower than the leaves but higher than that the nutritive roots from February to March. The accumulating quantity slightly increased in the plant. CONCLUSION: The application quantity of N should be increased gradually after transplant, stopped in autumn, and used again in early spring. Phosphate fertilizer is needed in autumn and winter, while large amount of potassium fertilizer shoule be used in winter tine.