Exploring the potential mechanisms of Rehmannia glutinosa in treating sepsis based on network pharmacology.

The present study utilized network pharmacology to identify therapeutic targets and mechanisms of Rehmannia glutinosa in sepsis treatment. RNA-sequencing was conducted on peripheral blood samples collected from 23 sepsis patients and 10 healthy individuals. Subsequently, the RNA sequence data were analyzed for differential expression. Identification of active components and their putative targets was achieved through the HERB and SwissTarget Prediction databases, respectively. Functional enrichment analysis was performed using GO and KEGG pathways. Additionally, protein-protein interaction networks were constructed and survival analysis of key targets was conducted. Single-cell RNA sequencing provided cellular localization data, while molecular docking explored interactions with central targets. Results indicated significant involvement of identified targets in inflammation and Th17 cell differentiation. Survival analysis linked several targets with mortality rates, while molecular docking highlighted potential interactions between active components and specific targets, such as rehmaionoside a with ADAM17 and rehmapicrogenin with CD81. Molecular dynamics simulations confirmed the stability of these interactions, suggesting Rehmannia glutinosa's role in modulating immune functions in sepsis.

Preparation of Rehmanniae Radix Praeparata Polysaccharide Iron(III) Complex and Evaluation of Its Biological Activity.

This study extracted and purified a polysaccharide from Rehmanniae radix praeparata (RGP) with an average molecular weight. The structural characteristics of RGP and its iron (III) complex, RGP-Fe(III), were examined for their antioxidant properties and potential in treating iron deficiency anemia (IDA). Analysis revealed that RGP comprised Man, Rha, Gal, and Xyl, with a sugar residue skeleton featuring 1→3; 1→2, 3; and 1→2, 3, 4 linkages, among others. RGP-Fe(III) had a molecular weight of 4.39×104 Da. Notably, RGP-Fe(III) exhibited superior antioxidant activity compared to RGP alone. In IDA rat models, treatment with RGP-Fe(III) led to increased weight gain, restoration of key blood parameters including hemoglobin, red blood cells, and mean hemoglobin content, elevated serum iron levels, and decreased total iron-binding capacity. Histological examination revealed no observable toxic effects of RGP-Fe(III) on the liver and spleen. These findings suggest the potential of RGP-Fe(III) as a therapeutic agent for managing IDA and highlight its promising antioxidant properties.

Seven new pentasaccharides from the roots of Rehmannia glutinosa.

Seven new pentasaccharides (1-7), rehmaglupentasaccharides A-G, were isolated from the air-dried roots of Rehmannia glutinosa. Their structures were established from the spectroscopic data obtained and by chemical evidence. The known verbascose (8) and stachyose (9) were also obtained in the current investigation, and the structure of stachyose was unequivocally defined using X-ray diffraction data. Compounds 1-9 were tested for their cytotoxicity against five human tumor cell lines, influence on dopamine receptor activation, and proliferation effects against Lactobacillus reuteri.

Four new iridoid glycosides from the roots of Rehmannia glutinosa.

Four new iridoid glycosides (1-4), rehmaglutosides L-O, were isolated from the air-dried roots of Rehmannia glutinosa. Their structures were established from the spectroscopic data obtained and by chemical evidence. The known mellittoside (5) and ajugol (6) were also obtained in the current investigation, and the structure of mellittoside was unequivocally defined using X-ray diffraction data. Compounds 1-6 were tested for their cytotoxicity against five human tumor cell lines and proliferation effects on Lactobacillus Reuteri.

Two new iridoid glycosides from the whole plant of Rehmania piasezkii.

Two new iridoid glycosides, piasezkiiosides A (1) and B (2), were isolated from aqueous extract of the whole plant of Rehmannia piasezkii. Their structures were established from the spectroscopic data, chemical transformation, and X-ray diffraction analysis. Compound 1 exhibited weak hepatoprotective activity against APAP-induced HepG2 cell damage.

Exploration of the Dynamic Variations of the Characteristic Constituents and the Degradation Products of Catalpol during the Process of Radix Rehmanniae.

Radix Rehmanniae (RR), a famous traditional Chinese medicine (TCM) widely employed in nourishing Yin and invigorating the kidney, has three common processing forms in clinical practice, including fresh Radix Rehmanniae (FRR), raw Radix Rehmanniae (RRR), and processed Radix Rehmanniae (PRR). However, until now, there has been less exploration of the dynamic variations in the characteristic constituents and degradation products of catalpol as a representative iridoid glycoside with the highest content in RR during the process from FRR to PRR. In this study, an ultra-performance liquid chromatography coupled with photodiode array detector (UPLC-PDA) method was successfully established for the simultaneous determination of ten characteristic components to explore their dynamic variations in different processed products of RR. Among them, iridoid glycosides, especially catalpol, exhibited a sharp decrease from RRR to PRR. Then, three degradation products of catalpol were detected under simulated processing conditions (100 °C, pH 4.8 acetate buffer solution), which were isolated and identified as jiofuraldehyde, cataldehyde, and norviburtinal, respectively. Cataldehyde was first reported as a new compound. Moreover, the specificity of norviburtinal in self-made PRR samples was discovered and validated, which was further confirmed by testing in commercially available PRR samples. In conclusion, our study revealed the decrease in iridoid glycosides and the production of new degradation substances during the process from FRR to PRR, which is critical for unveiling the processing mechanism of RR.

[Role of mitochondrial autophagy and the curative effect of rehmannia glutinosa leaves total glycoside capsules on nucleos(t)ide drug-induced renal injury].

Objective: To study the curative effect of rehmannia glutinosa leaves total glycoside capsules and the role of mitochondrial autophagy on nucleos(t)ide drug-induced renal injury. Methods: Adefovir dipivoxil (ADV) was used to construct a hepatitis B virus (HBV) transgenic mouse model for renal injury. Renal function was measured in each group at one and two weeks of modeling. Mitochondrial autophagy indicators were measured at two weeks of modeling in renal tissue. Transmission electron microscopy was used to detect mitochondrial autophagy phenomena in renal tissue. The model was established for two weeks. Mouse with renal injury were treated with rehmannia glutinosa leaves total glycoside capsules or isotonic saline for eight weeks by intragastric administration. Renal function was measured. Renal tissue morphology was observed. Mitochondrial autophagy indicators were detected in renal tissue. The protective effect of different concentrations of verbascoside (the main active ingredient of rehmannia glutinosa capsule) was observed on HK-2 cell damage induced by ADV. HK-2 cells were divided into control, ADV, and ADV plus verbascoside groups. The effects of verbascoside at different times and concentrations were observed on the HK-2 mitochondrial autophagy indicators. Fifty patients with chronic hepatitis B were collected who presented with renal injury after treatment with nucleos(t)ide analogs. The random number method was used to divide 29 cases into a control group that received conventional treatment. The treatment group of 21 cases was treated with rehmannia glutinosa leaves total glycoside capsules on the basis of the control group. Serum creatinine (Scr) and urinary protein were detected at eight weeks.The χ(2) test or t-test was used for statistical analysis. Results: Compared with the control group, two weeks of modeling in the ADV group induced renal function injury in HBV mice. The expression of autophagy indicators was higher in the renal tissue of the ADV group than that of the control group. Transmission electron microscopy had revealed mitochondrial autophagy in the renal tissue of the ADV group. Compared with the control group, the renal function of HBV mice treated with rehmannia glutinosa leaves total glycoside capsules improved for two months, and the expressions of autophagy indicators were down-regulated.Verbascoside promoted proliferation in ADV-damaged HK-2 cells, and the expression of autophagy indicators was down-regulated compared with the ADV alone group. In 50 patients with renal function injury, the urinary protein improvement was significantly superior in the treatment group than that in the control group, with eighteen and three cases being effective and ineffective in the treatment group and 12 and 17 cases being effective and ineffective in the control group, with a statistically significant difference (χ(2) = 9.975 0, P = 0.001 6). Serum creatinine was decreased in the treatment group compared with the control group, with 11 and 10 cases being effective and ineffective in the treatment group and 12 and 17 cases being effective and ineffective in the control group, with no statistically significant difference (χ(2) = 0.593 5, P = 0.441 1). Conclusion: Rehmannia glutinosa leaves total glycoside capsule can improve the nucleos(t)ide drug-induced renal function injury in chronic hepatitis B, possibly playing a role via inhibiting PINK1/Parkin-mediated mitochondrial autophagy.

[Gene cloning and functional analysis of an iridoid oxidase gene in Rehmannia glutinosa].

Rehmannia glutinosa is one of the commonly used Chinese herbal medicines, which has activities of heat-clearing,blood-cooling, Yin-nourishing, and body fluid-promoting. Iridoid glycosides are the main bioactive in R. glutinosa. Iridoid oxidase is a key rate-limiting enzyme in the biosynthetic pathway of iridoid glycosides. In this study, an iridoid oxidase gene Rg IO was screened based on the transcriptome data, followed by bioinformatics analysis, expression characteristic detection, and subcellular localization analysis. The results show that the coding region of Rg IO is 1 536 bp, with 511 amino acids encoded, and the molecular weight is about 58 258. 01. The protein sequence of Rg IO contains the conserved domains and motifs of cytochrome P450 oxidases. Rg IO has the highest sequence identities with its ortholog proteins in Striga asiatica, Striga hermonthica, and Centranthera grandiflora and has good sequence identities(77. 28%) with Catharanthus roseus Cr IO. Rg IO shows specific expression in the leaf of R. glutinosa. In response to MeJA induction, the expression of MeJA in leaves and roots after treatment increases by 3. 15 and 1. 3 times at 3 h and 6 h,respectively. The result of subcellular localization shows that Rg IO is distributed in the endoplasmic reticulum. Agrobacterium-mediated transient expression of Rg IO gene in leaves of R. glutinosa makes the content of catalpol increase by 0. 82 times compared with the transient expression of the empty vector. This study provides a key target gene for the molecular regulation and biosynthesis of catalpol in R. glutinosa and lays a foundation for revealing the complete biosynthetic pathway of catalpol.

[Effect of 2-phenylethyl-beta-glucopyranoside isolated from Huaizhong No. 1 Rehmannia glutinosa on hypoxic pulmonary hypertension by regulating PI3K/Akt/m TOR/HIF-1α pathway].

The study investigated the protective effect and mechanism of 2-phenylethyl-beta-glucopyranoside(Phe) from Huaizhong No.1 Rehmannia glutinosa on hypoxic pulmonary hypertension(PH), aiming to provide a theoretical basis for clinical treatment of PAH. Male C57BL/6N mice were randomly divided into normal group, model group, positive drug(bosentan, 100 mg·kg~(-1)) group, and low-and high-dose Phe groups(20 and 40 mg·kg~(-1)). Except for the normal group, all other groups were continuously subjected to model induction in a 10% hypoxic environment for 5 weeks, with oral administration for 14 days starting from the 3rd week. The cardiopulmonary function, right ventricular pressure, cough and asthma index, lung injury, cell apoptosis, oxidative stress-related indicators, immune cells, and phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/mammalian target of rapamycin(mTOR)/hypoxic inducible factor 1α(HIF-1α) pathway-related proteins or mRNA levels were examined. Furthermore, hypoxia-induced pulmonary arterial smooth muscle cell(PASMC) were used to further explore the mechanism of Phe intervention in PH combined with PI3K ago-nist(740Y-P). The results showed that Phe significantly improved the cardiopulmonary function of mice with PH, decreased right ventricular pressure, cough and asthma index, and lung injury, reduced cell apoptosis, oxidative stress-related indicators, and nuclear levels of phosphorylated Akt(p-Akt) and phosphorylated mTOR(p-mTOR), inhibited the expression levels of HIF-1α and PI3K mRNA and proteins, and maintained the immune cell homeostasis in mice. Further mechanistic studies revealed that Phe significantly reduced the viability and migration ability of hypoxia-induced PASMC, decreased the expression of HIF-1α and PI3K proteins and nuc-lear levels of p-Akt and p-mTOR, and this effect was blocked by 740Y-P. Therefore, it is inferred that Phe may exert anti-PH effects by alleviating the imbalance of oxidative stress and apoptosis in lung tissues and regulating immune levels, and its mechanism may be related to the regulation of the PI3K/Akt/mTOR/HIF-1α pathway. This study is expected to provide drug references and research ideas for the treatment of PH.

Omics analyses of Rehmannia glutinosa dedifferentiated and cambial meristematic cells reveal mechanisms of catalpol and indole alkaloid biosynthesis.

BACKGROUND: Rehmannia glutinosa is a rich source of terpenoids with a high medicinal reputation. The present study compared dedifferentiated cells (DDCs) and cambial meristematic cells (CMCs) cell cultures of R. glutinosa for terpenoid (catalpol) and indole alkaloid (IA) biosynthesis. In this regard, we used widely targeted metabolomics and transcriptome sequencing approaches together with the comparison of cell morphology, cell death (%), and catalpol production at different time points. RESULTS: We were able to identify CMCs based on their morphology and hypersensitivity to zeocin. CMCs showed higher dry weight content and better catalpol production compared to DDCs. The metabolome analysis revealed higher concentrations of IA, terpenoids, and catalpol in CMCs compared to DDCs. The transcriptome sequencing analysis showed that a total of 27,201 genes enriched in 139 pathways were differentially expressed. The higher catalpol concentration in CMCs is related to the expression changes in genes involved in acetyl-CoA and geranyl-PP biosynthesis, which are precursors for monoterpenoid biosynthesis. Moreover, the expressions of the four primary genes involved in monoterpenoid biosynthesis (NMD, CYP76A26, UGT6, and CYP76F14), along with a squalene monooxygenase, exhibit a strong association with the distinct catalpol biosynthesis. Contrarily, expression changes in AADC, STR, and RBG genes were consistent with the IA biosynthesis. Finally, we discussed the phytohormone signaling and transcription factors in relation to observed changes in metabolome. CONCLUSIONS: Overall, our study provides novel data for improving the catalpol and IA biosynthesis in R. glutinosa.

Identification of raffinose family oligosaccharides in processed Rehmannia glutinosa Libosch using matrix-assisted laser desorption/ionization mass spectrometry image combined with machine learning.

RATIONALE: Currently, research on oligosaccharides primarily focuses on the physiological activity and function, with a few studies elaborating on the spatial distribution characterization and variation in the processing of Rehmannia glutinosa Libosch. Thus, imaging the spatial distributions and dynamic changes in oligosaccharides during the steaming process is significant for characterizing the metabolic networks of R. glutinosa. It will be beneficial to characterize the impact of steaming on the active ingredients and distribution patterns in different parts of the plant. METHODS: A highly sensitive matrix-assisted laser desorption/ionization mass spectrometry image (MALDI-MSI) method was used to visualize the spatial distribution of oligosaccharides in processed R. glutinosa. Furthermore, machine learning was used to distinguish the processed R. glutinosa samples obtained under different steaming conditions. RESULTS: Imaging results showed that the oligosaccharides in the fresh R. glutinosa were mainly distributed in the cortex and xylem. As steaming progressed, the tetra- and pentasaccharides were hydrolyzed and diffused gradually into the tissue section. MALDI-MS profiling combined with machine learning was used to identify the processed R. glutinosa samples accurately at different steaming intervals. Eight algorithms were used to build classification machine learning models, which were evaluated for accuracy, precision, recall, and F1 score. The linear discriminant analysis and random forest models performed the best, with prediction accuracies of 0.98 and 0.97, respectively, and thus can be considered for identifying the steaming durations of R. glutinosa. CONCLUSIONS: MALDI-MSI combined with machine learning can be used to visualize the distribution of oligosaccharides and identify the processed samples after steaming for different durations. This can enhance our understanding of the metabolic changes that occur during the steaming process of R. glutinosa; meanwhile, it is expected to provide a theoretical reference for the standardization and modernization of processing in the field of medicinal plants.

Purple Rehmannia: Investigation of the activation of R2R3-MYB transcription factors involved in anthocyanin biosynthesis.

Engineering anthocyanin biosynthesis in herbs could provide health-promoting foods for improving human health. Rehmannia glutinosa is a popular medicinal herb in Asia, and was a health food for the emperors of the Han Dynasty (59 B.C.). In this study, we revealed the differences in anthocyanin composition and content between three Rehmannia species. On the 250, 235 and 206 identified MYBs in the respective species, six could regulate anthocyanin biosynthesis by activating the ANTHOCYANIDIN SYNTHASE (ANS) gene expression. Permanent overexpression of the Rehmannia MYB genes in tobacco strongly promoted anthocyanin content and expression levels of NtANS and other genes. A red appearance of leaves and tuberous/roots was observed, and the total anthocyanin content and the cyanidin-3-O-glucoside content were significantly higher in the lines overexpressing RgMYB41, RgMYB42, and RgMYB43 from R. glutinosa, as well as RcMYB1 and RcMYB3 in R. chingii and RhMYB1 from R. henryi plants. Knocking out of RcMYB3 by CRISPR/Cas9 gene editing resulted in the discoloration of the R. chingii corolla lobes, and decreased the content of anthocyanin. R. glutinosa overexpressing RcMYB3 displayed a distinct purple color in the whole plants, and the antioxidant activity of the transgenic plants was significantly enhanced compared to WT. These results indicate that Rehmannia MYBs can be used to engineer anthocyanin biosynthesis in herbs to improve their additional value, such as increased antioxidant contents.

A comparative metabolomics study on dried and processed Rehmannia Radix.

The efficacy of Rehmannia Radix changes after processing. However, the precise effect of processing on the properties of Rehmannia Radix is an intricate topic, as this effect cannot be explained by traditional methods. The purpose of this study was to investigate how processing methods influence the properties of Rehmannia Radix, as well as the changes in body function after administering dried Rehmannia Radix (RR) and processed Rehmannia Radix (PR) using a metabolomics approach. In addition, principal component analysis and orthogonal partial least-squares discriminant analysis models were generated using SIMCA-P 14.0 to evaluate the properties of RR and PR. Potential biomarkers were identified, and associated metabolic networks were established to clarify differences in the properties and efficacies of RR and PR. The results showed that RR and PR have cold and hot properties, respectively. RR can exert a hypolipidaemic effect by regulating nicotinate and nicotinamide metabolism. PR exerts a tonic effect and regulates the body's reproductive function through the regulation of alanine, aspartate and glutamate metabolism, arachidonic acid, pentose and glucuronate metabolism, respectively. Ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry-based metabolomics is a promising approach to determine the cold/hot properties of traditional Chinese medicine formulations.

Rehmannia Glutinosa Polysaccharide Regulates Bone Marrow Microenvironment via HIF-1α/NF-κB Signaling Pathway in Aplastic Anemia Mice.

Aplastic anemia (AA), a rare disorder, is associated with bone marrow microenvironment (BMM). Presently, AA treatment is of great difficulty. This study aimed to explore the mechanism of action of Rehmannia glutinosa polysaccharide (RGP) in AA. Busulfan was used to induce AA in BALB/c mice; blood cell count and Ray's Giemsa staining were used to assess the severity of hematopoietic failure; HE was performed to assess the pathological state of the marrow cavity; ELISA was performed to assess IL-4, IL-10, IL-6, IL-12, IL-1ß, TNF-α, MCP-1, VEGF, and EPO; and WB was performed to evaluate the effects of RGP on the HIF-1α/NF-κB signaling. Significant downregulation of hemocyte levels in the blood and nucleated cells in the bone marrow was reversed by RGP and Cyclosporine A (CA). Compared with the AA group, dilating blood sinusoids, inflammation, hematopoiesis, decreased bone marrow cells and megakaryocytes were alleviated by RGP and CA, and the HIF-1α/NF-κB signaling was inhibited too. Notably, RGP was more effective when used in combination with CA. In this study, we established a relationship between BMM and the HIF-1α/NF-κB signaling pathway and found that RGP regulates BMM by suppressing the activation of the HIF-1α/NF-κB signaling. Thus, RGP exerts a pharmacological effect on AA.

Angiotensin I-Converting Enzyme-Inhibitory Activity and Phytochemical Profile of Constituents of the Leaves of Rehmannia glutinosa f. hueichingensis.

The root of Rehmannia glutinosa Liboschitz forma hueichingensis HSIAO has been used as a tonic and treatment for urinary and skin disorders in Japanese Kampo medicine. Phytochemical investigation of the root has been well reported, but that of the leaves is limited. To explore the potential value of R. glutinosa leaves, we focused on the angiotensin I-converting enzyme (ACE)-inhibitory activity. The leaf extract exhibited ACE-inhibitory activity, and the inhibitory potency of leaves was stronger than that of roots. Using this activity as an indicator, we isolated linaride (1), 6-O-hydroxybenzoyl ajugol (2), acteoside (3), leucosceptoside A (4), martynoside (5), luteolin (6), apigenin (7), and chrysoeriol (8) by separating and purifying the extract. We then examined the ACE-inhibitory activities of 1-8, catalpol (9), aucubin (10), ajugol (11), and echinacoside (12). Among them, 3, 6, and 12 displayed the most potent inhibitory activity. A simultaneous analytical method was also developed using compounds contained in R. glutinosa leaves and roots, and their contents were compared. The method consisted of extraction with 50% aqueous methanol under sonication for 60 min and LC/MS measurement. R. glutinosa leaves tended to have higher levels of majority of the analytes than the roots, including 3 and 6, which had higher ACE-inhibitory activity. These results suggest that 3 and 6 contribute to the ACE-inhibitory activity of R. glutinosa leaves, which may represent a useful medicinal resource for hypertension.

Rehmannia glutinosa polysaccharides attenuates colitis via reshaping gut microbiota and short-chain fatty acid production.

BACKGROUND: Ulcerative colitis is a gastrointestinal disease closely related to intestinal epithelial barrier damage and intestinal microbiome imbalance; however, effective treatment methods are currently limited. Rehmannia glutinosa polysaccharide (RGP) is an important active ingredient with a wide range of pharmacological activities, although its protective effect on colitis remains to be explored. In the present study, we verified the in vitro anti-inflammatory effect of RGP, and observed the ameliorating effect of RGP on dextran sulfate sodium-induced colitis in mice. RESULTS: The results showed that (i) RGP attenuates lipopolysaccharide-induced overexpression of inflammatory factors in RAW264.7 cells; (ii) RGP improves the pathological damage caused by DSS, including weight loss, increased disease activity index and intestinal tissue ulcers; (iii) RGP improves tight junction proteins to protects the tightness of the intestinal epithelium; (iv) RGP inhibits the expression of inflammatory factors through the nuclear factor-kappa B pathway, and improved the of intestinal tissues inflammation; and (v) RGP can maintain the species diversity of intestinal microbes, increase the content of short-chain fatty acids and then restore the imbalance of intestinal microecology. CONCLUSION: RGP can improve the intestinal microbiota to strengthen the intestinal epithelial barrier and protect against DSS-induced colitis. © 2022 Society of Chemical Industry.

[Effect of processing method on chemical constituents of Rehmanniae Radix: based on UHPLC-LTQ-Orbitrap MS].

This study aims to explore the chemical composition of Rehmanniae Radix braised with mild fire and compare the effect of processing method on the chemical composition of Rehmanniae Radix. To be specific, ultra-high performance liquid chromatography with linear ion trap-orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) was used to screen the chemical constituents of Rehmanniae Radix. The chemical constituents were identified based on the relative molecular weight and fragment ions, literature information, and Human Metabolome Database(HMDB). The ion peak area ratio of each component before and after processing was used as the index for the variation. SIMCA was employed to establish principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) models of different processed products. According to the PCA plot, OPLS-DA plot, and VIP value, the differential components before and after the processing were screened out. The changes of the content of differential components with the processing method were analyzed. A total of 66 chemical components were identified: 57 of raw Rehmanniae Radix, 55 of steamed Rehmanniae Radix, 55 of wine-stewed Rehmanniae Radix, 51 of repeatedly steamed and sundried Rehmanniae Radix Praeparata, 62 of traditional bran-braised Rehmanniae Radix, and 63 of electric pot-braised Rehmanniae Radix. Among them, the 9 flavonoids of braised Rehmanniae Radix were from Citri Reticulatae Pericarpium. PCA suggested significant differences in the chemical composition of Rehmanniae Radix Praeparata prepared with different processing methods. OPLS-DA screened out 32 chemical components with VIP value >1 as the main differential components. Among the differential components, 9 were unique to braised Rehmanniae Radix(traditional bran-braised, electric pot-braised) and the degradation rate of the rest in braised(traditional bran-braised, electric pot-braised) or repeatedly steamed and sundried Rehmanniae Radix was higher than that in the steamed or wine-stewed products. The results indicated the chemical species and component content of Rehmanniae Radix changed significantly after the processing. The 32 components, such as rehmapicrogenin, martynoside, jionoside D, aeginetic acid, hesperidin, and naringin, were the most important compounds to distinguish different processed products of Rehmanniae Radix. The flavonoids introduced by Citri Reticulatae Pericarpium as excipient may be the important material basis for the effectiveness of braised Rehmanniae Radix compared with other processed products.

[Adjuvant rice optimization of rice-steamed Rehmanniae Radix and its anti-osteoporosis effect].

The present study aimed to investigate the effect of various adjuvant rice on the quality of rice-steamed Rehmanniae Radix(RSRR) with Japonica rice, millet, yellow rice, black rice, and glutinous rice as raw materials, and analyze the anti-osteoporosis effect of RSRR by the optimal adjuvant rice. On the basis of the established UPLC-MS/MS method for the determination of the content of catalpol and rehmannioside D, comprehensive weighted scoring method was employed to evaluate the effect of various auxiliary rice on the quality of RSRR with the content of catalpol and rehmannioside D, character score, and taste score as indicators to optimize adjuvant rice. The osteoporosis model was induced by ovariectomy in rats. SD rats were randomly divided into a sham operation group, a model group, a positive control group, and low-dose and high-dose groups of Rehmanniae Radix, RSRR, steamed Rehmanniae Radix, and Epimedii Folium-RSRR. After treatment for 12 weeks, body weight, bone calcium content, and bone mineral density were mea-sured. The results showed that Japonica rice was selected as the optimal adjuvant due to the highest comprehensive score of RSRR steamed by Japonica rice. Rehmanniae Radix, RSRR, steamed Rehmanniae Radix, as well as Epimedii Folium-RSRR, could improve osteoporosis by increasing bone calcium content and bone mineral density. RSRR was superior to Rehmanniae Radix in improving osteo-porosis. However, there was no significant difference between RSRR and steamed Rehmanniae Radix. This study confirmed that Japo-nica rice was the optimal adjuvant rice of RSRR and verified the anti-osteoporosis effect of RSRR, which laid a foundation for further research on the pharmacological action and mechanism of RSRR.

Functional characterization of tyrosine decarboxylase genes that contribute to acteoside biosynthesis in Rehmannia glutinosa.

MAIN CONCLUSION: The RgTyDCs possess typical decarboxylase functional activity in vitro and in vivo and participate in acteoside biosynthesis in R. glutinosa, positively controlling its production via activated acteoside/tyrosine-derived pathways. Acteoside is an important ingredient in Rehmannia glutinosa and an active natural component that contributes to human health. Tyrosine decarboxylase (TyDC) is thought to play an important role in acteoside biosynthesis. Several plant TyDC family genes have been functionally characterized and shown to play roles in some bioactive metabolites' biosynthesis by mediating the decarboxylation of L-tyrosine and L-dihydroxyphenylalanine (L-DOPA); however, one TyDC (named RgTyDC1) in R. glutinosa has been identified to date, but the family genes that contribute to acteoside biosynthesis remain largely characterized. Here, by in silico and experimental analyses, we isolated and identified three RgTyDCs (RgTyDC2 to RgTyDC4) in this species; these genes' sequences showed 50.92-82.55% identity, included highly conserved domains with homologues in other plants, classified into two subsets, and encoded proteins that localized to the cytosol. Enzyme kinetic analyses of RgTyDC2 and RgTyDC4 indicated that they both efficiently catalysed L-tyrosine and L-dopa. The overexpression of RgTyDC2 and RgTyDC4 in R. glutinosa, which was associated with enhanced TyDC activity, significantly increased tyramine and dopamine contents, which was positively correlated with improved acteoside production; moreover, the overexpression of RgTyDCs led to upregulated expression of some other genes-related to acteoside biosynthesis. This result suggested that the overexpression of RgTyDCs can positively activate the molecular networks of acteoside pathways, enhancing the accumulation of tyramine and dopamine, and promoting end-product acteoside biosynthesis. Our findings provide an evidence that RgTyDCs play vital molecular roles in acteoside biosynthesis pathways, contributing to the increase in acteoside yield in R. glutinosa.

Study on Catalpol Content in Rehmannia glutinosa Root, an Important Ingredient in Kampo Prescriptions.

Rehmannia glutinosa is an important medicinal plant in Asia, and its roots are used as an ingredient in herbal medicine. However, the roots exhibit different medicinal effects depending on the processing conditions. Since the catalpol content differs greatly during the process, the catalpol content is an essential index for quality evaluation. R. glutinosa roots have various weights, diameters, and lengths, and there are differences between individuals and within an individual immediately after harvest. We found that, catalpol content in the roots tended to increase as root diameter increased. Furthermore, it has been reported that catalpol content decreased with drying, and our results also supported this phenomenon. To clarify the reason for the decrease in catalpol content, we investigated the effect of ß-glucosidase in R. glutinosa cells. An in situ assay for ß-glucosidase activity revealed that the activity in the tissue inside the cambium disappeared one month after drying under natural conditions, and the activity in the tissue outside the cambium completely disappeared after two months. Because catalpol content remained almost unchanged even after drying for two months, it was clarified that ß-glucosidase activity had minimal involvement in the decrease in catalpol content in R. glutinosa roots. Based on the above results, we proposed that slicing the roots and rapidly removing water by natural drying is best to obtain dry root with little loss of catalpol content.