Characterization of CYP82 genes involved in the biosynthesis of structurally diverse benzylisoquinoline alkaloids in Corydalis yanhusuo.

Benzylisoquinoline alkaloids (BIAs) represent a significant class of secondary metabolites with crucial roles in plant physiology and substantial potential for clinical applications. CYP82 genes are involved in the formation and modification of various BIA skeletons, contributing to the structural diversity of compounds. In this study, Corydalis yanhusuo, a traditional Chinese medicine rich in BIAs, was investigated to identify the catalytic function of CYP82s during BIA formation. Specifically, 20 CyCYP82-encoding genes were cloned, and their functions were identified in vitro. Ten of these CyCYP82s were observed to catalyze hydroxylation, leading to the formation of protopine and benzophenanthridine scaffolds. Furthermore, the correlation between BIA accumulation and the expression of CyCYP82s in different tissues of C. yanhusuo was assessed their. The identification and characterization of CyCYP82s provide novel genetic elements that can advance the synthetic biology of BIA compounds such as protopine and benzophenanthridine, and offer insights into the biosynthesis of BIAs with diverse structures in C. yanhusuo.

Protective effects of Typhonii Rhizoma in rheumatoid arthritis rats revealed by integrated metabolomics and network pharmacology.

Rheumatoid arthritis (RA) is an autoimmune disease with a 0.5% prevalence worldwide. Inflammation, periosteal proliferation and joint destruction are the main clinical symptoms of RA. Typhonii Rhizoma (TR) is the dry tuber of the Araceae plant Typhonium giganteum Engl, and possesses many uses such as dispelling obstructive wind-phlegm and relieving pain. It is used for the clinical treatment of arthromyodynia and RA. However, the mechanism of action remains unclear. In this study, we first evaluated the effects of TR in type II collagen-induced RA model rats. Secondly, in serum metabolomics, TR could ameliorate 11 potential metabolites in RA model rats and reversed RA through pentose and glucuronate interconversions, sphingolipid metabolism, glycerophospholipid metabolism and tryptophan metabolism. To further explore the mechanisms of TR, 40 chemical constituents were used to establish a component-target interaction network. Some key genes were verified by in vitro pharmacological tests by integrating the results from the network pharmacology and metabolomics. The verification results showed that the mechanisms of TR against RA may be related to the inhibition of the production of inflammatory cytokines and the expression and function of HIF1-α. This study serves as a theoretical basis for the treatment of RA with TR.

Analysis of Differences in the Chemical Composition of Glycosides and Sugars between Four Forms of Fresh Rehmanniae Radix.

Fresh Rehmanniae Radix, as well as its processed products, are widely used in the clinical practice of traditional Chinese medicine. It is mainly available in four forms: fresh Rehmanniae Radix, raw Rehmanniae Radix, prepared Rehmanniae Radix, and nine-steamed, nine-dried Rehmanniae Radix. Pharmacological studies have shown that all Rehmanniae Radix forms contain iridoid glycosides and sugar compounds with various effects, including hypoglycemic, anti-inflammatory, neuroprotective, immunological enhancement, and bone marrow hematopoiesis-promoting activities. Differences in the efficacy among these Rehmanniae Radix forms and their processed products have been attributed to variations in their chemical compositions, particularly in iridoid glycosides and sugar compounds; however, the specific compositional differences in glycosides and sugars among the four forms of Rehmanniae Radix have not been clarified. Therefore, this study aims to qualitatively characterize the iridoid glycosides and sugar compounds in fresh Rehmanniae Radix, raw Rehmanniae Radix, prepared Rehmanniae Radix, and nine-steamed, nine-dried Rehmanniae Radix.

(±)-Cryptamides A-D, Four Pairs of Novel Dopamine Enantiomer Trimers from the Periostracum Cicadae.

Four pairs of novel dopamine enantiomer trimers, (±)-cryptamides A-D (1-4), and 10 pairs of previously described dopamine enantiomer dimers (5-14) were isolated from the Periostracum cicadae, the cast-off shell of the insect Cryptotympana pustulata. Aside from being pairs of enantiomers, the eight trimers were also elucidated to be regioisomers, most likely resulting from their mechanism of formation, [4 + 2] cycloaddition. The discovery of dopamine trimers is rarely reported when it comes to natural products derived from insects.

Parents' personality, parenting stress, and problem behaviors of children with special needs in China before and during the COVID-19 pandemic.

The positive predictive effect of children's problem behaviors on parenting stress has been verified to some extent, but research on parents of children with special needs remains insufficient. Moreover, the role of parental personality traits in the relationship between children's problem behaviors and parenting stress, and whether it differs from before the COVID-19 pandemic, remains unclear. Accordingly, in this study, online questionnaires were used to survey parents of children with autism and intellectual disabilities in China - 337 parents before and 604 during the COVID-19 pandemic - to explore the relationship between problem behaviors in the children and parenting stress as well as the moderating effect of parents' personality. The results showed that problem behaviors of children with autism and intellectual disabilities had a positive predictive effect on parenting stress. However, there was no significant difference in this effect before and during the pandemic. In addition, the relationship between children's problem behaviors and parenting stress was moderated by the Agreeableness and Neuroticism of the parents, but only during COVID-19 pandemic. The research results suggest that, during the pandemic when facing problem behaviors of children with autism or intellectual disabilities, positive personality characteristics such as Agreeableness have a protective effect on parenting stress. By contrast, negative personality characteristics such as Neuroticism are risk factors. The study results provide evidence from special groups regarding the role of parents' personalities in the parent - child interaction and the parenting stress models.

[Iridoids of wine-processed Corni Fructus].

A new iridoid glycoside, cornushmf A(1) and nine known iridoids(2-10) were isolated from the water extract of the wine-processed Corni Fructus by various column chromatographies. Their chemical structures were identified by comprehensive spectroscopic methods as 7ß-O-(2″-formylfuran-5″-methylene)-morroniside(1), 7-dehydrologanin(2), sweroside(3), 7ß-O-methylmorroniside(4), 7α-O-methylmorroniside(5), 7ß-O-ethylmorroniside(6), 7α-O-ethylmorroniside(7), cornuside(8), sarracenin(9), and loganin(10).

Rapid Screening for EGFR Inhibitor in Rhei Radix et Rhizoma by HTRF Assay Coupled with HPLC Peak Fractionation.

In this paper, an HPLC peak fractionation approach combined with homogeneous time-resolved fluorescence analysis is proposed for screening epidermal growth factor receptor inhibitors from Rhei Radix et Rhizoma. With this approach, the amount of sample used for a single HPLC run is sufficient for performing a multiple assay due to the miniaturization ability of the homogeneous time-resolved fluorescence technology. This allows for improving the stability and repeatability of the activity assay for each fraction. From a total of 26 fractions collected from the Rhei Radix et Rhizoma extract, 13 fractions exhibit inhibitory activity against the epidermal growth factor receptor. The structures of activity compounds were determined by HPLC-LTQ-Orbitrap MS, revealing the presence of gallic acid, rhein, and emodin with IC50 values of 21.5, 5.29, and 10.2 µM, respectively. The ligand epidermal growth factor receptor interactions were explored by molecular docking simulations, and the inhibitory effects of the three compounds on A549 cell growth were tested in vitro by an MTT assay. This study demonstrates the suitability of the present screening method for drug discovery in natural products.

Four New Gallate Derivatives from Wine-Processed Corni Fructus and Their Anti-Inflammatory Activities.

Four new gallate derivatives-ornusgallate A, ent-cornusgallate A, cornusgallate B and C (1a, 1b, 2, 3)-were isolated from the wine-processed fruit of Cornus officinalis. Among them, 1a and 1b are new natural compounds with novel skeletons. Their chemical structures were elucidated by comprehensive spectroscopy methods including NMR, IR, HRESIMS, UV, ECD spectra and single-crystal X-ray diffraction analysis. The in vitro anti-inflammatory activities of all compounds were assayed in RAW 264.7 cells by assessing LPS-induced NO production. As the result, all compounds exhibited anti-inflammatory activities at attested concentrations. Among the tested compounds, compound 2 exhibited the strongest anti- inflammatory activity.

[Quality analysis of Citri Trifoliatae Fructus in different harvest time by HPLC combined with electronic eye technology].

As a local variety of medicinal material, Citri Trifoliatae Fructus is widely used in many places, whereas its harvest time remains unclear. Therefore, studying its harvest time can make more reasonable use of this medicinal material. In this study, we determined the flavonoids content and compared the color of Citri Trifoliatae Fructus harvested in different time, aiming to guide the harvest of this medicinal material. The fresh fruits of Citrus trifoliata were collected from Xinxiang city, Henan province, graded according to the diameter range, and then dried. The contents of isonaringin, naringen, and poncirin in Citri Trifoliatae Fructus were determined by HPLC, and the color values of the samples were detected by electronic eye. The correlation analysis of the obtained data was carried out to explore the relationships of color and diameter with quality. The results showed that the contents of isonaringin, naringen, and poncirin varied significantly in different harvest time, within the ranges of 0.21-1.20, 2.21-11.59, and 3.73-23.16 mg·g~(-1), respectively. With the delay of harvest time, Citri Trifoliatae Fructus showed the color changing from green to yellow, gradually increased diameter, and gradually decreased contents of isonaringin, naringen, and poncirin. The contents of isonaringin, naringen, and poncirin were negatively correlated with the degree of red and green(a~*) and positively correlated with the degree of yellow and blue(b~*). The contents of naringen and poncirin had significantly negative correlations with the diameter. This study indicates that the quality of Citri Trifoliatae Fructus can be judged by its diameter and skin color, which provides a theoretical basis for the rational harvest of this medicinal material.

Lilium davidii extract alleviates p­chlorophenylalanine­induced insomnia in rats through modification of the hypothalamic-related neurotransmitters, melatonin and homeostasis of the hypothalamic-pituitary-adrenal axis.

CONTEXT: Lilium davidii var. unicolour Cotton (Lilium genus, Liliaceae) is an edible plant and a herb used in China to alleviate insomnia. OBJECTIVE: To investigate the alleviating insomnia mechanism of L. davidii (LD). MATERIALS AND METHODS: Wistar rats were intraperitoneally injected with p-chlorophenylalanine (PCPA) to establish an insomnia model. Rats were divided into six groups (n = 8): Control, PCPA, Estazolam (0.5 mg/kg), LD extract in low, medium and high doses (185.22, 370.44, 740.88 mg/kg). Serum hormone levels of the HPA axis, levels of 5-HT, NE and MT, and the expression of GABAA and 5-HT1A receptors in hypothalamus were determined. Moreover, behavioural and pathological changes in the hypothalamus were evaluated. RESULTS: After LD administration, body weight and brain coefficient increased by 2.74% and 8.22%, respectively, and the adrenal coefficient decreased by 25%, compared with PCPA group. Elevation of the serum hypothalamic-pituitary-adrenal (HPA) axis hormone CRH (11.24 ± 3.16 ng/mL), ACTH (565.87 ± 103.44 pg/mL) and CORT (44.28 ± 8.73 ng/mL) in the PCPA group was reversed after LD treatment. Furthermore, abnormal excitatory behaviour [5 min movement distance (2096.34 ± 259.51 cm), central exercise time (5.28 ± 1.08 s)] of insomnia rats in the PCPA group was also relieved. LD extract increased 5-HT and MT levels, reduced NE level in the hypothalamus, and upregulated the expression of GABAA R and 5-HT1A. Moreover, LD extract may improve the pathology of neurons in the hypothalamus. CONCLUSIONS: LD can be considered to develop health-care food or novel drugs to cope with the increasing number of insomniacs.

Screening, and identification of the binding position, of xanthine oxidase inhibitors in the roots of Lindera reflexa Hemsl using ultrafiltration LC-MS combined with enzyme blocking.

A method based on enzyme blocking combined with ultrafiltration liquid chromatography-mass spectrometry (LC-MS) has been developed to identify xanthine oxidase (XOD) inhibitors in the roots of Lindera reflexa Hemsl (LR) and determine their binding positions. Allopurinol and febuxostat, known XOD inhibitors, which occupy different binding positions in XOD, were used as blockers and pre-incubated with XOD. Then the LR extract was incubated without XOD, and with XOD, allopurinol-blocked XOD and febuxostat-blocked XOD before ultrafiltration LC-MS was performed. By comparing the chromatographic profiles of the incubation samples, not only the ligands, but also the binding position of these ligands with XOD could be determined. Finally, three compounds, pinosylvin, pinocembrin and methoxy-5-hydroxy-trans-stilbene, were identified as potential XOD inhibitors and the binding modes of these three compounds were shown to be similar to those of febuxostat. To verify the XOD inhibitory activity of the screened compounds, the microplate method and molecular docking in silico were used to evaluate the enzyme inhibitory activities and the binding positions with XOD. The results showed that the developed method could screen for XOD ligands in LR extracts and also determine the binding positions of the ligands. To our knowledge, this is the first report of the XOD inhibitory activity of these three compounds.

Four New ent-Kaurane Diterpene Glycosides from Isodon henryi.

To obtain diterpene glycosides from an aqueous extract of the aerial parts of Isodon henryi and further investigate their cytotoxicities, in this study, a total of seven compounds were isolated, including six ent-kaurane diterpene glycosides (1-6) and one diterpene aglycon (7). Among the seven ent-kaurane diterpenes obtained, four were novel compounds, including ent-7,20-epoxy- kaur-16-en-1α,6ß,7ß,15ß-tetrahydroxyl-11-O-ß-d-glucopyranoside (1), ent-7,20-epoxy-kaur-16-en- 6ß,7ß,14ß,15ß-tetrahydroxyl-1-O-ß-d-glucopyranoside (2), ent-7,20-epoxy-kaur-16-en-6ß,7ß,15ß- trihydroxyl-1-O-ß-d-glucopyranoside (3), and ent-7,20-epoxy-kaur-16-en-7ß,11ß,14α,15ß-tetrahydr- oxyl-6-O-ß-d-glucopyranoside (4), and three were isolated from this plant for the first time (5-7). Their structures were elucidated by utilizing spectroscopic methods and electronic circular dichroism analyses. Furthermore, the cytotoxicities of all seven compounds were investigated in four human cancer cell lines, including A2780, BGC-823, HCT-116, and HepG2. The IC50 values of these diterpenes ranged from 0.18 to 2.44 mM in the tested cell lines. In addition, the structure-cytotoxicity relationship of diterpene glycosides was also evaluated to study the effect of glycosylation on the cytotoxicity of diterpene compounds.

[Studies on chemical constituents of Isodon henryi].

The chemical constituents of the water extraction of the aerial parts of Isodon henryi were investigated by various chromatographic methods including D-101 macroporous adsorptive resins,silica gel,sephadex LH-20,and semi-preparative HPLC. As a result,ten compounds were separated and purified. By analyses of the UV,IR,MS,NMR spectra,their structures were determined as rabdosinate( 1),lasiokaurin( 2),epinodosinol( 3),rabdosichuanin C( 4),epinodosin( 5),hebeirubescensin k( 6),rubescensin C( 7),enmenol( 8),oridonin( 9),and enmenol-1-ß-glucoside( 10). Compounds 1-8 and 10 were isolated from I. henryi for the first time. Compounds 2 and 9 showed inhibitory effects against four tumor cells,with IC50 values of 2. 25-9. 32 µmol·L-1.

[Study of extracting natural resources of Chinese medicinal materials planted area in Luoning of Henan province based on UAV of low altitude remote sensing technology and remote sensing image of satellite].

The study is aimed to effectively obtain the planting area of traditional Chinese medicine resources. The herbs used as the material for traditional Chinese medicine are mostly planted in natural environment suitable mountainous areas. The UAV low altitude remote sensing data were used as the samples and the GF-2 remote sensing images were applied for the data source to extract the planting area of Salvia miltiorrhiza and Artemisia argyi in Luoning county combined with field investigation. Remote sensing satellite data of standard processing obtain specific remote sensing data coverage. The UAV data were pre-processed to visually interpret the species and distribution of traditional Chinese medicine resources in the sample quadrat. Support vector machine( SVM) was used to classify and estimate the area of traditional Chinese medicine resources in Luoning county,confusion matrix was used to determine the accuracy of spatial distribution of traditional Chinese medicine resources. The result showed that the application of UAV of low altitude remote sensing technology and remote sensing image of satellite in the extraction of S. miltiorrhiza and other varieties planting area was feasible,it also provides a scientific reference for poverty alleviation policies of the traditional Chinese medicine Industry in local areas.Meanwhile,research on remote sensing classification of Chinese medicinal materials based on multi-source and multi-phase high-resolution remote sensing images is actively carried out to explore more effective methods for information extraction of Chinese medicinal materials.

Oridonin Inhibits Myofibroblast Differentiation and Bleomycin-induced Pulmonary Fibrosis by Regulating Transforming Growth Factor ß (TGFß)/Smad Pathway.

BACKGROUND Idiopathic pulmonary fibrosis (IPF) is a progressive disease with unknow. etiology and a high mortality rate. Oridonin is a diterpenoid isolated from the Rabdosia rubesecens with diverse biological functions. However, whether oridonin possess potential protective activity on IPF is still unclear. MATERIAL AND METHODS The aim of the present study was to explore the therapeutic effects of oridonin on IPF. First, TGF-ß1-induced MRC-5 cells were employed for the evaluation of inhibitory activity in vitro. Then, a bleomycin (BLM)-induced mice pulmonary fibrosis model was used to verify the activity of oridonin in vivo. Several pathological changes, including alveolar space collapse, emphysema, and infiltration of inflammatory cells, were observed in the BLM­treated mice. RESULTS Oridonin could significantly inhibit the mRNA and protein expression levels of α-SMA and COL1A1 in TGF-ß1-induced MRC-5 cells. Oridonin could attenuate pathological changes, including alveolar space collapse, emphysema, and infiltration of inflammatory cells induced by BLM. In addition, oridonin could significantly inhibit BLM-induced upregulation of α-SMA and COL1A1 and the phosphorylation of Smad2/3 in lung tissues of mice. CONCLUSIONS Oridonin could be used as a potential therapeutic agent in treatment for patients with IPF. The mechanisms of anti-fibrosis effect of oridonin might be inhibition of the TGF-ß/Smad pathway.

Simultaneous Analysis of Saccharides between Fresh and Processed Radix Rehmanniae by HPLC and UHPLC-LTQ-Orbitrap-MS with Multivariate Statistical Analysis.

Radix Rehmanniae (RR) is a kind of herb which is widely used in the clinical and food processing industry. There are four forms of RR used in traditional Chinese medicine practice, which include fresh RR (FRR), raw RR (RRR), processed RR (PRR), and another processed RR (APRR), in which the APRR was processed by nine cycles of repeated steaming and drying. There are a large number of saccharides in RR. However, the differences in content were shown by different processing methods. In this study, an effective method using high-performance liquid chromatography (HPLC) and high-performance liquid chromatography-mass spectrometry (LC-MS) coupled with multivariate statistical analysis to rapidly distinguish different RR samples and validate the proposed chemical conversion mechanism. The datasets of the content of saccharides were subjected to principal component analysis (PCA) and one-way analysis of variance. The results showed that there different changes occurred in the contents of saccharides corresponding to the different processing methods, in which the contents of monosaccharides-namely arabinose, glucose, mannose, and galactose-had an increasing trend or remained relatively stable. However, the contents of fructose and oligosaccharides, including manninotriose, melibiose, sucrose, and raffinose, first increased and then reduced, or gradually decreased, yet the content of stachyose gradually decreased. The MSn data indicated that manninotriose, melibiose, and some monosaccharides were produced by the hydrolysis of oligosaccharides. In addition, the fragmentation pathways of 1-phenyl-3-methyl-5-pyrazolone (PMP) derivatization of monosaccharides were also found that its glycosidic bond was first broken and subsequently its inside ring broke, and the characteristic fragment ions were produced at m/z 511.22, 493.20, 373.16, and 175.08 in the PMP derivatization of monosaccharides. In conclusion, this study illustrates the change and chemical conversion mechanism of saccharides by processing in RR samples which might play a key role in further application of RR.

[Study on ecological suitability regionalization of Corni Fructus based on Maxent and ArcGIS model].

Through planting regionalization the scientific basis for planting area of high-quality medicinal materials was predicted. Through interview investigation and field survey, the distribution information of Corni Fructus in China was collected,and 89 sampling point from 14 producing areas were collected. Climate and topography of Corni Fructus were analyzed, the ecological adaptability of study was conducted based on ArcGIS and Maxent. Different suitability grade at potential areas and regionalization map were formulated. There are nine ecological factors affecting the growth of Corni Fructus, for example precipitation in November and March and vegetation type. The results showed that the most suitable habitats are Henan, Shaanxi, Zhejiang, Chongqing, Hubei, Sichuan, Anhui, Hunan and Shandong province. Using the spatial analysis method,the study not only illustrates the most suitable for the surroundings of Corni Fructus,but also provides a scientific reference for wild resource tending, introduction and cultivation, and artificial planting base and directing production layout.

[Studies on chemical constituents of Isodon excisoides].

The chemical constituents of the water extraction of the aerial parts of Isodon excisoides were investigated by various chromatographic methods including D-101 macroporous adsorptive resins, silica gel, Sephadex LH-20, MCI and semi-preparative HPLC. As a result, six compounds were separated and purified.By analyses of the HR-ESI-MS, 1D and 2D NMR spectra, their structures were determined as 3-O-ß-D-allopyranosyl-1-octen-3-ol(1), blumenolA (2), lumichrome (3), loliolide(4), cirsiliol(5) and pedalitin(6). Compound 1 was a new compound, and compounds 2-4 were isolated from this plant for the first time.

[Comparison and analysis of wild and cultivated Lonicerae japonicae flos from Xinmi].

OBJECTIVE: To compare and analyze the wild and cultivated Lonicerae Japonicae Flos from Xinmi. METHODS: Macroscopic identification and microscopic identification were adopted to compare and analyze characteristics of the wild and cultivated Lonicerae Japonicae Flos from Xinmi. The content of chlorogenic acid and luteoloside were determined by the methods in the Chinese Pharmacopoeia. DNA molecular marker technology was used to identify Lonicerae Japonicae Flos of different varieties. RESULTS: The differences in macroscopic and microscopic characteristics as well as the content of chlorogenic acid and luteoloside between them had been found. PCR amplification reaction system and procedure of Lonicerae Japonicae Flos from Xinmi were optimized. An effective primer for identification was chosen from 11 primers. CONCLUSION: The results provide the scientific basis for quality assessment of Lonicerae Japonicae Flos from Xinmi.

Investigating the mechanism of cornel iridoid glycosides on type 2 diabetes mellitus using serum and urine metabolites in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Cornel iridoid glycosides (CIG) are extracted from Corni fructus, a herbal medicine used in traditional Chinese medicine to treat diabetes. However, the antidiabetic effects of CIG and the underlying metabolic mechanisms require further exploration. AIM OF THE STUDY: This study aimed to assess the antidiabetic effects and metabolic mechanism of CIG by performing metabolomic analyses of serum and urine samples of rats. MATERIALS AND METHODS: A rat model of type 2 diabetes mellitus (T2DM) was established by administering a low dose of streptozotocin (30 mg/kg) intraperitoneally after 4 weeks of feeding a high-fat diet. The model was evaluated based on several parameters, including fasting blood glucose (FBG), random blood glucose (RBG), urine volume, liver index, body weight, histopathological sections, and serum biochemical parameters. Subsequently, serum and urine metabolomics were analyzed using ultra-high-pressure liquid chromatography coupled with linear ion trap-Orbitrap tandem mass spectrometry (UHPLC-LTQ-Orbitrap-MS). Data were analyzed using unsupervised principal component analysis (PCA) and supervised orthogonal partial least squares discriminant analysis (OPLS-DA). Differential metabolites were examined by the Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathways to explore the underlying mechanisms. RESULTS: After 4 weeks of treatment with different doses of CIG, varying degrees of antidiabetic effects were observed, along with reduced liver and pancreatic injury, and improved oxidative stress levels. Compared with the T2DM group, 19 and 23 differential metabolites were detected in the serum and urine of the CIG treatment group, respectively. The key metabolites involved in pathway regulation include taurine, chenodeoxycholic acid, glycocholic acid, and L-tyrosine in the serum and glycine, hippuric acid, phenylacetylglycine, citric acid, and D-glucuronic acid in the urine, which are related to lipid, amino acid, energy, and carbohydrate metabolism. CONCLUSIONS: This study confirmed the antidiabetic effects of CIG and revealed that CIG effectively controlled metabolic disorders in T2DM rats. This seems to be meaningful for the clinical application of CIG, and can benefit further studies on CIG mechanism.