Non-target metabolomics revealed the differences between Rh. tanguticum plants growing under canopy and open habitats.

BACKGROUND: Rheum tanguticum (Rh. tanguticum) is an important traditional Chinese medicine plant, "Dahuang", which contains productive metabolites and occupies wide habitats on the Qinghai-Tibet plateau. Plants occupying wide habitats usually vary in phenotypes such as in morphology and metabolism, thereby developing into different ecotypes. Under canopy and open habitats are a pair of dissimilar habitats which possess Rh. tanguticum plants. However, few studies have focused on the effect of habitats on Rh. tanguticum growth, particularly combining morphological and metabolic changes. This study focused on Rh. tanguticum plants growing in under canopy and open habitats where morphology and metabolism changes were quantified using non-target metabolism methods. RESULTS: The obtained results indicated that the two dissimilar habitats led to Rh. tanguticum developing into two distinct ecotypes where the morphology and metabolism were simultaneously changed. Under canopy habitats bred morphologically smaller Rh. tanguticum plants which had a higher level of metabolites (22 out of 31) which included five flavonoids, four isoflavonoids, and three anthracenes. On the other hand, the open habitats produced morphologically larger Rh. tanguticum plants having a higher level of metabolites (9 out of 31) including four flavonoids. 6 of the 31 metabolites were predicted to have effect targets, include 4 represent for under canopy habitats and 2 for open habitats. Totally, 208 targets were connected, among which 42 were communal targets for both under canopy and open habitats represent compounds, and 100 and 66 were unique targets for under canopy superior compounds and open habitats superior compounds, respectively. In addition, aloe-emodin, emodin, chrysophanol, physcion, sennoside A and sennoside B were all more accumulated in under canopy habitats, and among which aloe-emodin, emodin, chrysophanol and physcion were significantly higher in under canopy habitats. CONCLUSIONS: This study determined that Rh. tanguticum growing in under canopy and in open habitats developed into two distinct ecotypes with morphological and metabolic differences. Results of network pharmacology study has indicated that "Dahuang" coming from different habitats, such as under canopy and open habitats, are different in effect targets and thus may have different medicinal use. According to target metabolomics, under canopy habitats may grow better "Dahuang".

Effects of different nitrogen fertilizer levels on growth and active compounds of rhubarb from Qinghai plateau.

BACKGROUND: Rheum tanguticum Maxim. ex Balf is one of the plants generically known as rhubarb, a culinary vegetable that has long been used as a herbal remedy both in China and Europe. Increasing demand for rhubarb has triggered the overexploitation of Rh. tanguticum. Cultivation is therefore necessary for quality control and protection of wild resources. Nitrogen fertilizer plays an important role in cultivation. This study aimed to explore how nitrogen fertilizer affects the growth and quality of rhubarb on the Qinghai-Tibetan plateau. RESULTS: Nitrogen fertilizer promoted growth but had no significant influence on the active compounds of Rh. tanguticum. Generally, the N2 (150 kg ha-1 ) and N3 (225 kg ha-1 ) levels showed the most improved growth indexes, with no significant differences between them. The growth index and the amounts of eight of the nine studied active compounds in Rh. tanguticum increased from each year to the next and differed among growth stages. The contents of the active compounds were higher at the green stage and lower at the growth stage, which was opposite to the seasonal trends in root dry matter ratio. Gallic acid levels decreased with the growth of the plant. CONCLUSION: The N2 level (150 kg ha-1 ) was the recommended nitrogen fertilizer level in this study. It was revealed that seasonal changes rather than nitrogen fertilizer influenced active compounds in the root of Rh. tanguticum. © 2018 Society of Chemical Industry.

Extended differentiation of veins and stomata is essential for the expansion of large leaves in Rheum rhabarbarum.

PREMISE OF THE STUDY: The densities of veins and stomata govern leaf water supply and gas exchange. They are coordinated to avoid overproduction of either veins or stomata. In many species, where leaf area is greater at low light, this coordination is primarily achieved through differential cell expansion, resulting in lower stomatal and vein density in larger leaves. This mechanism would, however, create highly inefficient leaves in species in which leaf area is greater at high light. Here we investigate the role of cell expansion and differentiation as regulators of vein and stomatal density in Rheum rhabarbarum, which produces large leaves under high light. METHODS: Rheum rhabarbarum plants were grown under full sunlight and 7% of full sunlight. Leaf area, stomatal density, and vein density were measured from leaves harvested at different intervals. KEY RESULTS: Leaves of R. rhabarbarum expanded at high light were six times larger than leaves expanded at low light, yet vein and stomatal densities were similar. In high light-expanded leaves, minor veins were continuously initiated as the leaves expanded, while an extended period of stomatal initiation, compared to leaves expanded at low light, occurred early in leaf development. CONCLUSIONS: We demonstrate that R. rhabarbarum adjusts the initiation of stomata and minor veins at high light, allowing for the production of larger leaves uncoupled from lower vein and stomatal densities. We also present evidence for an independent control of vein and stomatal initiation, suggesting that this adjustment must involve some unknown developmental mechanism.

Influence of the Environmental Factors on the Accumulation of the Bioactive Ingredients in Chinese Rhubarb Products.

To provide a basis for controlling the quality of rhubarb under artificial cultivation, the present work was designed to evaluate the contents of 14 active pharmaceutical ingredients (API) of rhubarb in major rhubarb production areas in China and analyze the correlations between the contents of API and such factors as species, geographic distribution and soil. The levels of fourteen API in rhubarb were measured using HPLC. The geographic distributions were collected using GPS and the nutrients in the soil were measured using the methods in the literature. The results showed that the levels of major API vary significantly among plants of different locations according to variance analysis. The species factor has few obvious effect on the overall properties of the rhubarb by the cluster analysis because of the two source species occurring in all divided three groups. However, Rheum tanguticum Maxim.ex Balf. is less effective at synthesizing and accumulating 9 API out of 14 than Rheum palmatum L. The correlation analysis and regression analysis also indicated that a lower latitude should be considered in the accumulation of API and a lower longitude should be considered to produce more compound anthraquinones. Lower levels of total P, rapidly available P and available molybdenum (Mo) and higher available K and available Zn in the soil were significantly correlated to accumulation of API in rhubarb. These results provide a basis for the clinical application and controlling the levels of the major API of rhubarb during artificial cultivation.

[Suitable Habitats Prediction of Original Plants of Rhei Radix et Rhizoma Under Climate Change].

OBJECTIVE: To predict the distribution patterns of the original plants of Rhei Radix et Rhizoma current and in future in China under climate change. METHODS: A maximum entropy modeling and variety of climate change scenarios were employed to predict its current and future distribution ranges in China. RESULTS: The resultant models exhibited excellent predictive power. The current suitable habitats for original plants of Rhei Radix et Rhizoma totaled 84. 20 x 10(4) km2 accounting for 8. 77% of the total area of China; 57. 05% of its current suitable habitats(i. e. low impact areas) would be relatively lowly impacted by the climate change. Compared with its current distribution pattern,its distribution ranges during 2020s, 2030s, 2040s, 2050, 2060s, 2070s and 2080s would be shrunk to some extent. However, the moderately suitable area would be expanded to a certain degree. CONCLUSION: Climate change has a negative impact on the total area and the habitat suitability for the original plants of Rhei Radix et Rhizoma.

[Resource situation investigation about Rheum tanguticum and its sustainable utilization analysis in main production area of China].

This study was conducted to investigate the wild and cultivated resource situation of Rheum tanguticum in main production area of China, estimate its reserves, and put forward the feasible approach for the sustainable utilization of R. tanguticum. On the basis of the literature data about R. tanguticum, conbined with interview, investigation and sampling investigation, the total reserve of resources is estimated using the route-quadrat method and the vegetation and soil-type map area method proposed by our research group. The results indicate that there is no obvious change between the present distribution ranges of the wild R. tanguticum and its historical records, but its population density has changed clearly. The reserve of the wild R. tanguticum has seriously declined in lots of place, even faced the exhaustion in some regions. According to the investigation, the resource reserve of the wild R. tanguticum is no more than 5 000 t, and the cultivated is about 1 607 t. The resource reserve of the wild R. tanguticum is nearly depleted, and this suggests that the wild R. tanguticum should be enrolled in the protection plant list, and the cultivated will become the main resource of Rhubarb in the future. So it is extremely neccessary to collect and protect the germplasm resource of R. tanguticum, establish the germplasm nursery and repository, and conduct breeding research on those bases.

[Research on HPLC fingerprint of Rheum palmatum from Gansu Province called "quan-shui-da-huang"].

OBJECTIVE: To establish an analysis method for HPLC fingerprint of Rheum palmatum collected from Gansu province (called "Quan-shui-da-huang"), and provide basis for quality control of Quan-shui-da-huang. METHODS: The Cosmosil 5C18-PAQ column (250 mm x 4.6 mm, 5 microm) was used with the mixture of 0.1% acetic acid and acetonitrile as mobile phase in a gradient elution mode. The flow rate was 1 mL/min, column temperature was 30 degrees C and detection wavelength was set at 280 nm. The similarity evaluation, principal component analysis (PCA) and cluster analysis of crude drugs collected from different habitats were carried out. RESULTS: The analysis method of HPLC fingerprint was set up. The characteristic fingerprint was obtained by chemometrics methods. CONCLUSION: Under the selected chromatographic conditions, constituents in Rheum palmatum can be separated well. This method can be used for the quality control of Quan-shui-da-huang.

[Influences of different initial processing methods on Rheum palmatum].

OBJECTIVE: To choose the optimum initial processing methods of Rheum palmatum. METHODS: Fresh crude Rheum palmatum was sliced and dealt with the different drying methods such as sun drying, shady drying, microwave heating and various temperatures drying. The content of the Anthraquinones derivatives, slicing colors and dried rates were used as evaluation indexes. The sliced Rheum palmatum was compared with the traditional processing. RESULTS: Sliced fresh crude Rheum palmatum had the low content of the Anthraquinones derivatives and dry rates, slicing colours had obviously changes. For various drying methods, smoking drying was superior to sun drying, shady drying, microwave heating and various temperatures drying methods. CONCLUSION: Fresh crude Rheum palmatum is not suitable for slicing processing. The best drying method is smoking drying.

[Summarization of Chinese rhubarb recorded by the Westerners in China in the Ming and Qing dynasties].

The westerners in China in the Ming and Qing dynasties, such as travellers, missionaries, archaeologists and explorers etc., left many records of Chinese rhubarb, among which included botanical characteristics, rhubarb trade and the preparation method and usage of rhubarb. These records reflected the evolution of cognition on Chinese rhubarb of the Westerners in China as well as the constant in-depth cognition on Chinese rhubarb, and also reflected the situation of sino-foreign medical exchange from one angle. Sifting the records of Chinese rhubarb by the Westerners who came to China in the Ming and Qing dynasties will contribute to further relevant research on rhubarb as well as sino-foreign medical exchanges.

Qualitative and quantitative analysis of anthraquinone derivatives in rhizomes of tissue culture-raised Rheum emodi Wall. plants.

This paper presents quantification of five anthraquinone derivatives (emodin glycoside, chrysophanol glycoside, emodin, chrysophanol and physcion) in rhizomes of hardened micro-propagated Rheum emodi plants using high-performance liquid chromatography (HPLC). Aseptic shoot cultures were raised using rhizome buds. Shoot multiplication occurred in both agar gelled and liquid Murashige and Skoog (MS) medium supplemented with 10.0 microM 6-benzylaminopurine (BAP) and 5.0 microM indole-3-butyric acid (IBA). Rooted plantlets obtained on plant growth regulator (PGR)-free medium were transferred to soil with 92% survival. HPLC analysis revealed the presence of five anthraquinone derivatives: emodin glycoside, chrysophanol glycoside, emodin, chrysophanol and physcion in rhizomes of tissue culture-raised plants. Only emodin glycoside (1) and chrysophanol glycoside (2) were present in 6-month-old hardened tissue cultured plants. In addition, the other three derivatives (emodin (3), chrysophanol (4) and physcion (5)) were also detected after 9 months.

Metabolism of sulphonated anthraquinones in rhubarb, maize and celery: the role of cytochromes P450 and peroxidases.

Sulphonated anthraquinones are precursors of many synthetic dyes and pigments, recalcitrant to biodegradation, and thus contaminating many industrial effluents and rivers. In the development of a phytotreatment to remove sulphonated aromatic compounds, rhubarb (Rheum rhaponticum), a plant producing natural anthraquinones, as well as maize (Zea mays) and celery (Apium graveolens), plants not producing anthraquinones, were tested for their ability to metabolise these xenobiotics. Plants were cultivated under hydroponic conditions, with or without sulphonated anthraquinones, and were harvested at different times. Either microsomal or cytosolic fractions were prepared. The monooxygenase activity of cytochromes P450 towards several sulphonated anthraquinones was tested using a new method based on the fluorimetric detection of oxygen consumed during cytochromes P450-catalysed reactions. The activity of cytosolic peroxidases was measured by spectrophotometry, using guaiacol as a substrate. Results indicated that the activity of cytochromes P450 and peroxidases significantly increased in rhubarb plants cultivated in the presence of sulphonated anthraquinones. A higher activity of cytochromes P450 was also detected in maize and celery exposed to the pollutants. In these two plants, a peroxidase activity was also detected, but without a clear difference between the control plants and the plants exposed to the organic contaminants. This research demonstrated the existence in rhubarb, maize and celery of biochemical mechanisms involved in the metabolism and detoxification of sulphonated anthraquinones. Taken together, results confirmed that rhubarb might be the most appropriate plant for the phytotreatment of these organic pollutants.

Rheum palaestinum (desert rhubarb), a self-irrigating desert plant.

The rare plant Rheum palaestinum (Polygonaceae) is a perennial hemicryptophyte that grows during the rainy winter in desert mountainous areas in Israel and Jordan that receive an average annual rainfall of ca. 75 mm. It produces between one and four large round leaves that are tightly attached to the ground and form large rosettes of up to 1 m(2). These leaves differ markedly from the typical small leaves of most desert plants. Moreover, they have a unique 3D morphology resembling a scaled-down mountainous area with well-developed steep drainage systems, raising the question which selective agents were involved in their evolution. We propose that the large leaves collect rainwater that then infiltrates the soil surrounding the root. We measured the seasonal course of leaf growth, examined the area of wet soil surrounding the root after actual and simulated rain, and modeled the water harvesting capacity using the plant leaf area and the weekly precipitation. We show that even in the slightest rains, water flows above the veins to the leaf's base where it irrigates the vertical root. A typical plant harvests more than 4,100 cm(3) of water per year, and enjoys a water regime of about 427 mm/year, equivalent to the water supply in a Mediterranean climate. This is the first example of self-irrigation by large leaves in a desert plant, creating a leaf-made mini oasis.

[Study of quality on rhubarb from different habitats in Qinghai Province].

OBJECTIVE: To investigate the effect of different habitats on rhubarb quality. METHOD: The rhubarb samples from various parts of Qinghai province were analysed by fingerprint. RESULTS AND CONCLUSION: The distribution and quality of wild Tangute rhubarb is better than Palmate sorrel rhubarb. The quality of high altitude sampes are better than low altitudes. The quality which from the grassland of plateau is better than chestnut soil area's, and the wild is better than that of planting.

[Induction of hairy roots and anthraquinone production in Rheum palmatum].

OBJECTIVE: To study the induction of hairy roots and anthraquinone production in Rheum palmatum. METHOD: Leaf blades, hypocotyls and petioles of R. palmatum were infected by Agrobacterium rhizogenes LBA9402 and A. rhizogenes R1601, respectively. RESULT: Hairy roots were induced by two strains of A. rhizogenes, and LBA9402 showed stronger infective ability than R1601. Three clones of hairy roots were cultured. DH7a was induced by R1601, DH5a and DH5c were induced by LBA9402. DH7a grew faster than DH5a and DH5c, and all of them grew faster obviously than normal root (NOR). There were significant differences in anthraquinone composition and content among four kinds of roots. Emodin physcion and chrysophanol were predominant anthraquinone in DH5a, DH5c and NOR respectivly. Aloe-emodin content was the lowest in all root cultures compared with other four anthraquinones. CONCLUSION: The in vitro culture system of the established hairy roots laid a foundation for mass production of anthraquinone by hairy root culture.

[Anthraquinone production and analysis in the hairy root cultures of Rheum palmatum L].

Hairy root culture of the medicinal plant Rheum palmatum L. was established by genetic transformation with Agrobacterium rhizogenes. The effects of various media with different pH on growth of the hairy roots and biosynthesis of free anthraquinones were investigated. The experimental results showed that MS agar medium with pH 5.5-5.8 is suitable for growth of the hairy roots. Dark condition is favourable and 62.5-fold increase in fresh weight was reached within a culture period of 25 days. Auxin (0.1 mg.L-1 IAA) activated the hairy root growth but inhibited the biosynthesis of free anthraquinones. About 28% of the total free anthraquinones was released into the liquid medium from the rhubarb hairy roots.