This study aimed to characterize and identify the non-volatile components in aqueous and ethanolic extracts of the stems and leaves of Rhododendron tomentosum by using sensitive and efficient ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry(UHPLC-Q-TOF-MS) combined with a self-built information database. By comparing with reference compounds, analyzing fragment ion information, searching relevant literature, and using a self-built information database, 118 compounds were identified from the aqueous and ethanolic extracts of R. tomentosum, including 35 flavonoid glycosides, 15 phenolic glycosides, 12 flavonoids, 7 phenolic acids, 7 phenylethanol glycosides, 6 tannins, 6 phospholipids, 5 coumarins, 5 monoterpene glycosides, 6 triterpenes, 3 fatty acids, and 11 other types of compounds. Among them, 102 compounds were reported in R. tomentosum for the first time, and 36 compounds were identified by comparing them with reference compounds. The chemical components in the ethanolic and aqueous extracts of R. tomentosum leaves and stems showed slight differences, with 84 common chemical components accounting for 71.2% of the total 118 compounds. This study systematically characterized and identified the non-volatile chemical components in the ethanolic and aqueous extracts of R. tomentosum for the first time. The findings provide a reference for active ingredient research, quality control, and product development of R. tomentosum.
Rhododendron/chemistry* ; Chromatography, High Pressure Liquid/methods* ; Drugs, Chinese Herbal/chemistry* ; Mass Spectrometry/methods* ; Plant Leaves/chemistry*

This study investigated the chemical constituents of Rhododendri Mollis Flos. The n-butanol fraction of 95% ethanol extract of Rhododendri Mollis Flos was separated and purified using chromatographic techniques, including normal-phase silica gel, ODS, and Sephadex LH-20 column chromatography. The structures of the isolated compounds were identified by spectroscopic techniques. Seven compounds were isolated and identified as rhodomollein LXⅦ(1), rhodomollein X(2), nivalenol(3),(3S,6S)-cis-linalool-3,7-oxide-β-D-glucopyranoside(4), thymidine(5), cyclo(Ala-Tyr)(6), and(S)-5-hydroxypyrrolidin-2-one(7). Among them, compound 1 was identified as a new compound, and compound 3 was isolated from this plant for the first time. The analgesic activity of compound 1 was evaluated using the acetic acid-induced writhing test. At a dose of 5.0 mg·kg~(-1), compound 1 showed a 75.4% inhibition rate in the writhing test, indicating significant analgesic activity.
Drugs, Chinese Herbal/isolation & purification* ; Rhododendron/chemistry* ; Animals ; Flowers/chemistry* ; Mice ; Male ; Analgesics/isolation & purification* ; Molecular Structure

Five meroterpenoids, rhodonoids K-M (1-2), daurichromene E (3), and grifolins A-B (4-5), together with seven known compounds (6-12), were isolated from Rhododendron anthopogonoides. The chemical structures of these compounds were elucidated through comprehensive analysis of high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), ultraviolet (UV), infrared spectroscopy (IR), and nuclear magnetic resonance (NMR) data. Their absolute configurations were determined by comparing experimental electronic circular dichroism (ECD) spectra with computed values. Notably, compounds 1 and 3 demonstrated significant inhibitory effects on lipopolysaccharide (LPS)-induced inflammation in RAW264.7 cells. These compounds markedly suppressed the mRNA expressions of inflammatory factors, including interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) while also down-regulating the protein expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2).
Mice ; Rhododendron/chemistry* ; Animals ; Anti-Inflammatory Agents/isolation & purification* ; RAW 264.7 Cells ; Terpenes/isolation & purification* ; Molecular Structure ; Tumor Necrosis Factor-alpha/immunology* ; Cyclooxygenase 2/immunology* ; Nitric Oxide Synthase Type II/immunology* ; Macrophages/immunology* ; Interleukin-6/immunology* ; Lipopolysaccharides ; Interleukin-1beta/immunology*

Flavonoid 3-O-glucosyltransferase (3GT) is a key enzyme in the glucosidation of anthocyanins. To investigate the 3GT gene in rhododendron, we cloned an open reading frame (ORF) of 3GT gene (named Rh3GT) from Rhododendron hybridum Hort (Red cultivar) and then characterized this gene and the deduced protein in terms of the biochemical characteristics, expression level, and enzymatic function. The results showed that Rh3GT had a full length of 993 bp and encoded 330 amino acid residues. The deduced protein was hydrophilic, stable, weak acid, belonging to the glycosyltransferase family (GT-B type), with glutamine (Q) at position 44 in the PSPG box. The phylogenetic analysis showed that Rh3GT was most closely related to Vc3GT from Vaccinium corymbosum and Vm3GT from Vaccinium myrtillus. Rh3GT was expressed in the stems, leaves, and flowers and almost not expressed in the roots, with the highest expression level in petals during full blooming stage. Introduction of pCAMBIAL1302-Rh3GT into petals significantly up-regulated the expression level of Rh3GT and increased the total anthocyanin accumulation. Rh3GT was successfully expressed in Escherichia coli BL21 in the form of inclusion bodies with a size of about 36 kDa. The results of HPLC showed that the recombinant Rh3GT after denaturation, purification, and dilution could catalyze the synthesis of cyanidin and UDP-glucose to synthesize cyanidin 3-O-glucoside, indicating that the expressed protein had 3GT activity. This study provides basic data for further studying the molecular regulation mechanism of anthocyanin biosynthesis and theoretical support for molecular breeding of rhododendron.
Rhododendron/classification* ; Glucosyltransferases/metabolism* ; Cloning, Molecular ; Escherichia coli/metabolism* ; Recombinant Proteins/biosynthesis* ; Anthocyanins/biosynthesis* ; Phylogeny ; Plant Proteins/metabolism* ; Amino Acid Sequence

Rheumatoid arthritis(RA), as a chronic autoimmune disease, has a high incidence and disability rate, causing significant suffering to patients. Due to its complex pathogenesis, it has not been fully elucidated to date, and its treatment remains a challenging problem in the medical field. Although western medicine treatment options have certain efficacy, they require prolonged use and are expensive. Additionally, they carry risks of multiple infections and adverse reactions like malignancies. The Chinese herbal medicine Rhododendron molle is commonly used in folk medicine for its properties of dispelling wind, removing dampness, calming nerves, and alleviating pain in the treatment of diseases like rheumatic bone diseases. In recent years, modern clinical and pharmacological studies have shown that the diterpenoids in R. molle are effective components, exhibiting immune-regulatory, anti-inflammatory, and analgesic effects. This makes it a promising candidate for treating RA with a broad range of potential applications. However, R. molle has certain toxic properties that hinder its clinical application and lead to the wastage of its resources. This study reviewed recent research progress on the mechanism of R. molle in preventing and treating RA, focusing on its chemical components, anti-inflammatory and analgesic properties and summarized the adverse reactions associated with R. molle, aiming to offer new ideas for finding natural remedies for RA and methods to reduce toxicity while enhancing the effectiveness of R. molle. The study seeks to clarify the safety and efficacy of R. molle and its extracts, providing a theoretical basis for its application prospects and further promoting the development and utilization of R. molle resources.
Humans ; Rhododendron/chemistry* ; Arthritis, Rheumatoid/drug therapy* ; Anti-Inflammatory Agents ; Diterpenes/pharmacology* ; Analgesics

Objective To study the effect and mechanism of blueberry on regulating the mitochondrial inner membrane protein mitofilin/Mic60 in an in vitro model of metabolic dysfunction-associated liver disease (MAFLD). Methods L02 human hepatocytes were induced by free fatty acids (FFA) to establish MAFLD cell model. A normal group, a model group, an 80 μg/mL blueberry treatment group, a Mic60 short hairpin RNA (Mic60 shRNA) transfection group, and Mic60 knockdown combined with an 80 μg/mL blueberry treatment group were established. The intracellular lipid deposition was observed by oil red O staining, and the effect of different concentrations of blueberry pulp on the survival rate of L02 cells treated with FFA was measured by MTT assay. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), superoxide dismutase (SOD) activity, glutathione (GSH) and malondialdehyde (MDA) contents were measured by visible spectrophotometry. The expression of reactive oxygen species (ROS) in hepatocytes was observed by fluorescence microscopy, and the mRNA and protein expression of Mic60 were detected by real-time quantitative PCR and Western blot analysis, respectively. Results After 24 hours of FFA stimulation, a large number of red lipid droplets in the cytoplasm of L02 cells was observed, and the survival rate of L02 cells treated with 80 μg/mL blueberry was higher. The results of ALT, AST, TG, TC, MDA and the fluorescence intensity of ROS in blueberry treated group were lower than those in model group, while the levels of SOD, GSH, Mic60 mRNA and protein in blueberry treated group were higher than those in model group. Conclusion Blueberry promotes the expression of Mic60, increases the levels of SOD and GSH in hepatocytes, and reduces the production of ROS, thus alleviating the injury of MAFLD hepatocytes and regulating the disorder of lipid metabolism.
Humans ; Blueberry Plants/chemistry* ; Hepatocytes/metabolism* ; Liver/metabolism* ; Liver Diseases/metabolism* ; Reactive Oxygen Species/metabolism* ; Superoxide Dismutase/metabolism* ; Superoxides/metabolism* ; Mitochondrial Membranes/metabolism* ; Mitochondrial Proteins/metabolism* ; Plant Extracts/pharmacology*

Blueberries are rich in phenolic compounds including anthocyanins which are closely related to biological health functions. The purpose of this study was to investigate the antioxidant activity of blueberry anthocyanins extracted from 'Brightwell' rabbiteye blueberries in mice. After one week of adaptation, C57BL/6J healthy male mice were divided into different groups that were administered with 100, 400, or 800 mg/kg blueberry anthocyanin extract (BAE), and sacrificed at different time points (0.1, 0.5, 1, 2, 4, 8, or 12 h). The plasma, eyeball, intestine, liver, and adipose tissues were collected to compare their antioxidant activity, including total antioxidant capacity (T-AOC), superoxide dismutase (SOD) activity and glutathione-peroxidase (GSH-PX/GPX) content, and the oxidative stress marker malondialdehyde (MDA) level. The results showed that blueberry anthocyanins had positive concentration-dependent antioxidant activity in vivo. The greater the concentration of BAE, the higher the T-AOC value, but the lower the MDA level. The enzyme activity of SOD, the content of GSH-PX, and messenger RNA (mRNA) levels of Cu,Zn-SOD, Mn-SOD, and GPX all confirmed that BAE played an antioxidant role after digestion in mice by improving their antioxidant defense. The in vivo antioxidant activity of BAE indicated that blueberry anthocyanins could be developed into functional foods or nutraceuticals with the aim of preventing or treating oxidative stress-related diseases.
Male ; Mice ; Animals ; Antioxidants/pharmacology* ; Blueberry Plants ; Anthocyanins/pharmacology* ; Mice, Inbred C57BL ; Superoxide Dismutase ; Plant Extracts/pharmacology* ; Superoxide Dismutase-1

Flavanone 3-hydroxylase (F3H) is a key enzyme in the synthesis of phycocyanidins. In this experiment, the petals of red Rhododendron hybridum Hort. at different developmental stages were used as experimental materials. The R. hybridum flavanone 3-hydroxylase (RhF3H) gene was cloned using reverse transcription PCR (RT-PCR) and rapid-amplification of cDNA ends (RACE) techniques, and bioinformatics analyses were performed. Petal RhF3H gene expression at different developmental stages were analyzed by using quantitative real-time polymerase chain reaction (qRT-PCR). A pET-28a-RhF3H prokaryotic expression vector was constructed for the preparation and purification of RhF3H protein. A pCAMBIA1302-RhF3H overexpression vector was constructed for genetic transformation in Arabidopsis thaliana by Agrobacterium-mediated method. The results showed that the R. hybridum Hort. RhF3H gene is 1 245 bp long, with an open reading frame of 1 092 bp, encoding 363 amino acids. It contains a Fe²⁺ binding motif and a 2-ketoglutarate binding motif of the dioxygenase superfamily. Phylogenetic analysis showed that the R. hybridum RhF3H protein is most closely related to the Vaccinium corymbosum F3H protein. qRT-PCR analysis showed that the expression level of the red R. hybridum RhF3H gene tended to increase and then decrease in the petals at different developmental stages, with the highest expression at middle opening stage. The results of the prokaryotic expression showed that the size of the induced protein of the constructed prokaryotic expression vector pET-28a-RhF3H was about 40 kDa, which was similar to the theoretical value. Transgenic RhF3H Arabidopsis thaliana plants were successfully obtained, and PCR identification and β-glucuronidase (GUS) staining demonstrated that the RhF3H gene was integrated into the genome of A. thaliana plants. qRT-PCR, total flavonoid and anthocyanin contentanalysis showed that RhF3H was significantly higher expressed in the transgenic A. thaliana relative to that of the wild type, and its total flavonoid and anthocyanin content were significantly increased. This study provides a theoretical basis for investigating the function of RhF3H gene, as well as for studying the molecular mechanism of flower color in R. simsiib Planch.
Arabidopsis/metabolism* ; Rhododendron/metabolism* ; Amino Acid Sequence ; Anthocyanins/metabolism* ; Phylogeny ; Flavonoids/metabolism* ; Cloning, Molecular ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism*

Ten lignans were isolated from the ethanol extract of stems and branches of Rhododendron ovatum through column chromatography over silica gel, ODS, Sephadex LH-20, and MCI-gel resin and semi-preparative RP-HPLC. The structures of all compounds were elucidated by extensive spectroscopic data analysis(UV, IR, HR-ESI-MS, ECD and NMR) as(-)-4-epi-lyoniresinol-9'-O-α-L-rhamnopyranoside(1),(+)-lyoniresinol-3α-O-α-L-rhamnopyranoside(2),(+)-5'-methoxyisolariciresinol-9'-O-α-L-rhamnopyranoside(3),(-)-lyoniresinol-3α-O-β-D-glucopyranoside(4),(+)-lyoniresinol-3α-O-β-D-glucopyranoside(5),(-)-4-epi-lyoniresinol-9'-O-β-D-glucopyransoide(6), racemiside(7), neociwujiaphenol(8),(+)-syringaresinol(9), and homohesperitin(10). Among them, compound 1 was a new aryltetralin-type lignan. All the isolated lignans were tested for antioxidant activities in Fe~(2+)-cysteine induced rat liver microsomal lipid peroxidation in vitro, and compounds 8 and 9 showed antioxidant activities on the formation of malondiadehyde(MDA) in rat liver microsomes at 1×10~(-5) mol·L~(-1), with significant inhibitory rates of 75.20% and 91.12%, respectively.
Animals ; Rats ; Glucosides/chemistry* ; Rhododendron ; Antioxidants/pharmacology* ; Lignans/chemistry* ; Plant Stems

This study explores the effect of total flavonoids of Rhododendra simsii(TFR) on middle cerebral artery occlusion(MCAO)-induced cerebral injury in rats and oxygen-glucose deprivation/reoxygenation(OGD/R) injury in PC12 cells and the underlying mechanism. The MCAO method was used to induce focal ischemic cerebral injury in rats. Male SD rats were randomized into sham group, model group, and TFR group. After MCAO, TFR(60 mg·kg~(-1)) was administered for 3 days. The content of tumor necrosis factor-α(TNF-α), interleukin-1(IL-1), and interleukin-6(IL-6) in serum was detected by enzyme-linked immunosorbent assay(ELISA). The pathological changes of brain tissue and cerebral infarction were observed based on hematoxylin and eosin(HE) staining and 2,3,5-triphenyltetrazolium chloride(TTC) staining. RT-qPCR and Western blot were used to detect the mRNA and protein levels of calcium release-activated calcium channel modulator 1(ORAI1), stromal interaction molecule 1(STIM1), stromal intera-ction molecule 2(STIM2), protein kinase B(PKB), and cysteinyl aspartate specific proteinase 3(caspase-3) in brain tissues. The OGD/R method was employed to induce injury in PC12 cells. Cells were randomized into the normal group, model group, gene silencing group, TFR(30 μg·mL~(-1)) group, and TFR(30 μg·mL~(-1))+gene overexpression plasmid group. Intracellular Ca~(2+) concentration and apoptosis rate of PC12 cells were measured by laser scanning confocal microscopy and flow cytometry. The effect of STIM-ORAI-regulated store-operated calcium entry(SOCE) pathway on TFR was explored based on gene silencing and gene overexpression techniques. The results showed that TFR significantly alleviated the histopathological damage of brains in MCAO rats after 3 days of admini-stration, reduced the contents of TNF-α, IL-1, and IL-6 in the serum, down-regulated the expression of ORAI1, STIM1, STIM2, and caspase-3 genes, and up-regulated the expression of PKB gene in brain tissues of MCAO rats. TFR significantly decreased OGD/R induced Ca~(2+) overload and apoptosis in PC12 cells. However, it induced TFR-like effect by ORAI1, STIM1 and STIM2 genes silencing. However, overexpression of these genes significantly blocked the effect of TFR in reducing Ca~(2+) overload and apoptosis in PC12 cells. In summary, in the early stage of focal cerebral ischemia-reperfusion injury and OGD/R-induced injury in PC12 cells TFR attenuates ischemic brain injury by inhibiting the STIM-ORAI-regulated SOCE pathway and reducing Ca~(2+) overload and inflammatory factor expression, and apoptosis.
Animals ; Male ; Rats ; Apoptosis ; Brain Ischemia/metabolism* ; Caspase 3 ; Interleukin-1 ; Interleukin-6 ; Rats, Sprague-Dawley ; Reperfusion Injury/metabolism* ; Tumor Necrosis Factor-alpha/genetics* ; Flavonoids/pharmacology* ; Rhododendron/chemistry*