Hevea brasiliensis is the main source of natural rubber. Restricted by its tropical climate conditions, the planting area in China is limited, resulted in a low self-sufficiency. Periploca sepium which can produce natural rubber is a potential substitute plant. cis-prenyltransferase (CPT), small rubber particle protein (SRPP) and rubber elongation factor (REF) are key enzymes involved in the biosynthesis of cis-1, 4-polyisoprene, the main component of natural rubber. In this study, we cloned the promoter sequences of CPT, SRPP and REF through chromosome walking strategy. The spatial expression patterns of the three promoters were analyzed using GUS (β-glucuronidase) as a reporter gene driven by the promoters through Agrobacterium-mediated genetic transformation. The results showed that GUS driven by CPT, SRPP or REF promoter was expressed in leaves and stems, especially in the leaf vein and vascular bundle. The GUS activity in stems was higher than that in leaf. This study provided a basis for analyzing the biosynthesis mechanism of natural rubber and breeding new varieties of high yield natural rubber.
Peptide Elongation Factors/genetics* ; Plant Proteins/metabolism* ; Periploca/metabolism* ; Rubber ; Plant Breeding ; Cloning, Molecular

In this study, an ultra-performance liquid chromatography-quadrupole time-of-flight high resolution mass spectrometer(UPLC-Q-TOF-HRMS) was used to investigate the effects of the active ingredients in Periploca forrestii compound on spleen metabolism in rats with collagen-induced arthritis(CIA), and its potential anti-inflammatory mechanism was analyzed by network pharmacology. After the model of CIA was successfully established, the spleen tissues of rats were taken 28 days after administration. UPLC-Q-TOF-HRMS chromatograms were collected and analyzed by principal component analysis(PCA), orthogonal partial least squares discriminant analysis(OPLS-DA), and MetPA. The results showed that as compared with the blank control group, 22 biomarkers in the spleen tissues such as inosine, citicoline, hypoxanthine, and taurine in the model group increased, while 9 biomarkers such as CDP-ethanolamine and phosphorylcholine decreased. As compared with the model group, 21 biomarkers such as inosine, citicoline, CDP-ethanolamine, and phosphorylcholine were reregulated by the active ingredients in P. forrestii. Seventeen metabolic pathways were significantly enriched, including purine metabolism, taurine and hypotaurine metabolism, glycerophospholipid metabolism, and cysteine and methionine metabolism. Network pharmacology analysis found that purine metabolism, glycerophospholipid metabolism, and cysteine and methionine metabolism played important roles in the pathological process of rheumatoid arthritis. This study suggests that active ingredients in P. forrestii compound can delay the occurrence and development of inflammatory reaction by improving the spleen metabolic disorder of rats with CIA. The P. forrestii compound has multi-target and multi-pathway anti-inflammatory mechanism. This study is expected to provide a new explanation for the mechanism of active ingredients in P. forrestii compound against rheumatoid arthritis.
Rats ; Animals ; Periploca ; Cysteine ; Cytidine Diphosphate Choline ; Network Pharmacology ; Phosphorylcholine ; Metabolomics ; Arthritis, Rheumatoid/drug therapy* ; Biomarkers ; Glycerophospholipids ; Methionine ; Purines ; Chromatography, High Pressure Liquid

Two cardenolide glycosides, corotoxigenin 3-O-[β-D-glucopyranosyl-(1→4)-6-deoxy-β-D-glucopyranoside] (1) and coroglaucigenin 3-O-[β-D-glucopyranosyl-(1→4)-6-deoxy-β-D-glucopyranoside] (2), were isolated from the seed fairs of Asclepias curassavica. The structures of 1-2 were determined based on the combination of the analysis of their MS, NMR spectroscopic data and acid hydrolysis. The inhibitory effects of compounds 1 and 2 on human colorectal carcinoma cells (HCT116), non-small cell lung carcinoma cells (A549) and hepatic cancer cells (SMMC-7721) were evaluated. The results showed that both compounds 1 and 2 significantly inhibited the viability, proliferation, and migration of A549, HCT116 and SMMC-7721 cells, suggesting that compounds 1 and 2 can be applied in the treatment of lung, colon and liver cancers in clinical practice. This study may not only provide a scientific basis for clarifying the active ingredients in A. curassavica, but also help to understand its antitumor activity, which can promote the application of A. curassavica in clinical treatment of various cancers.
Antineoplastic Agents/pharmacology* ; Asclepias/chemistry* ; Cardenolides/pharmacology* ; Glycosides/pharmacology* ; Humans ; Seeds

BACKGROUND: Periploca aphylla is used by local population and indigenous medicine practitioners as stomachic, tonic, antitumor, antiulcer, and for treatment of inflammatory disorders. The aim of this study was to evaluate antidiabetic effect of the extract of P. aphylla and to investigate antioxidant and hypolipidemic activity in streptozotocin (STZ)-induced diabetic rats. METHODS: The present research was conducted to evaluate the antihyperglycemic potential of methanol extract of P. aphylla (PAM) and subfractions n-hexane (PAH), chloroform (PAC), ethyl acetate (PAE), n-butanol (PAB), and aqueous (PAA) in glucose-overloaded hyperglycemic Sprague-Dawley rats. Based on the efficacy, PAB (200 mg/kg and 400 mg/kg) was tested for its antidiabetic activity in STZ-induced diabetic rats. Diabetes was induced via intraperitoneal injection of STZ (55 mg/kg) in rat. Blood glucose values were taken weekly. HPLC-DAD analysis of PAB was carried out for the presence of various polyphenols. RESULTS: HPLC-DAD analysis of PAB recorded the presence of rutin, catechin, caffeic acid, and myricetin. Oral administration of PAB at doses of 200 and 400 mg/kg for 21 days significantly restored (P < 0.01) body weight (%) and relative liver and relative kidney weight of diabetic rats. Diabetic control rats showed significant elevation (P < 0.01) of AST, ALT, ALP, LDH, total cholesterol, triglycerides, LDL, creatinine, total bilirubin, and BUN while reduced (P < 0.01) level of glucose, total protein, albumin, insulin, and HDL in serum. Count of blood cells and hematological parameters were altered in diabetic rats. Further, glutathione peroxidase, catalase, superoxide dismutase, glutathione reductase, and total soluble protein concentration decreased while concentration of thiobarbituric acid reactive substances and percent DNA damages increased (P < 0.01) in liver and renal tissues of diabetic rats. Histopathological damage scores increased in liver and kidney tissues of diabetic rats. Intake of PAB (400 mg/kg) resulted in significant improvement (P < 0.01) of above parameters, and results were comparable to that of standard drug glibenclamide. CONCLUSION The result suggests the antihyperglycemic, antioxidant, and anti-inflammatory activities of PAB treatment in STZ-compelled diabetic rat. PAB might be used as new therapeutic agent in diabetic patients to manage diabetes and decrease the complications.
1-Butanol/chemistry* ; Administration, Oral ; Animals ; Diabetes Mellitus, Experimental/drug therapy* ; Dose-Response Relationship, Drug ; Hypoglycemic Agents/chemistry* ; Male ; Periploca/chemistry* ; Phytochemicals/chemistry* ; Plant Extracts/chemistry* ; Rats ; Rats, Sprague-Dawley ; Streptozocin/adverse effects*

The liver and kidney fibrosis model was established by thioacetamide(TAA) and unilateral ureteral obstruction(UUO) in SD rats. The rats were randomly divided into three groups: model group, low and high-dose groups of C21 steroidal glycosides of Cynanchum auriculatum. Another blank control group was set. Four weeks later, serum was taken to detect the biochemical indexes of liver and kidney function. Urine protein and urine creatinine were detected by kits. Liver and kidney tissue samples were stained with HE and Masson staining, and hydroxyproline content was detected. Western blot was used to detect expressions of fibrotic proteins, inflammatory factors and TLR4 signaling pathways, so as to observe the preventive and therapeutic effects of C21 steroidal glycosides from C. auriculatum on hepatic and renal fibrosis and explore its molecular mechanism. Four weeks later, serum biochemical results showed that liver and kidney functions were seriously damaged, and pathological sections showed that inflammatory cell infiltration, decrease of parenchymal cells, and increase of interstitial fibrosis in liver and kidney tissues. The results showed that low and high doses(150, 300 mg·kg~(-1)) of C21 steroidal glycosides could significantly reduce the collagen deposition and the pathological changes of liver and kidney fibrosis compared with the model group. At the same time, we found that the expression levels of TLR4 and MyD88 signaling pathway proteins were significantly increased in the liver and kidney tissues of the model group, and a large number of NF-κB signaling pathway proteins migrated into the nucleus. On the contrary, the expression levels of TLR4, MyD88 signaling pathway proteins and the nuclear migration of NF-κB were significantly inhibited in the low and high dose groups of C21 steroidal glycosides from C. auriculatum. Therefore, it was speculated that the mechanism of C21 steroidal glycoside for preventive and therapeutic effect on hepatic and renal fibrosis was related to inhibit TLR4/MYD88/NF-κB inflammatory pathway, thus preventing hepatic and renal fibrosis.
Animals ; Cynanchum ; Fibrosis ; Glycosides ; Kidney/pathology* ; Liver ; NF-kappa B/genetics* ; Rats ; Rats, Sprague-Dawley ; Toll-Like Receptor 4/genetics*

During the high-temperature and rainy season from June to October in 2017-2019,serious southern blight broke out in the Cynanchum stauntonii planting area in Tuanfeng county,Hubei province,which had a great impact on the yield and quality of medicinal materials. In this study,the pathogen of C. stauntonii was isolated,purified,and identified,and the pathogenicity was tested according to Koch's postulates. Meanwhile,the biological characteristics of the pathogen were analyzed. On this basis,the effective fungicides were screened in laboratory. Finally,the pathogen( BQ-1) was identified as Athelia rolfsii( Deuteromycotina,Basidiomycota,anamorph: Sclerotium rolfsii). The optimum growth conditions for BQ-1 were 25-30 ℃,p H 5-8,and alternating light and dark.The effective chemical fungicides were lime-sulphur-synthelic-solution( LSSS) and flusilazole,and the effective botanical fungicide was osthole. BQ-1 was highly homologous to the pathogen HS-1 of peanut southern blight,with the similarity of 18 S r DNA and TEF sequences at 99. 09%. The southern blight in C. stauntonii might be resulted from that in peanut. In the production of C. stauntonii,the following measures should be taken: avoiding rotation or neighboring with peanut,draining water from June to October to reduce humidity,and reasonably applying fungicides.
Basidiomycota ; Cynanchum ; Fungicides, Industrial/pharmacology* ; Humidity

This study established high-performance liquid chromatography(HPLC) fingerprints of Chinese medicines derived from Apocynum venetum and Poacynum pictum in Xinjiang and explored their composition differences with the combination of content determination, similarity analysis, cluster analysis and principal component analysis. The HPLC conditions included Phenomenex Kinetex C_(18) column(4.6 mm ×100 mm, 2.6 μm), acetonitrile-0.01% trifluoroacetic acid aqueous solution as mobile phase, gradient elution, flow rate of 0.6 mL·min~(-1), detection wavelength of 281 nm and column temperature of 25 ℃. The content of chlorogenic acid, quercetin-3-O-sophoroside, rutin, hyperin, isoquercitrin, trifolin and astragalin was determined in 31 batches of medicinal materials, and fingerprint research and chemometric analysis were performed with Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine(Version 2004 A) and SPSS 21.0. In the Chinese Pharmacopoeia 2020, the quality of Apocyni Veneti Folium is controlled by character identification, microscopic identification, thin layer chromatography identification and quantitative determination of hyperin. There were 21 common peaks of A. venetum and P. pictum in the HPLC fingerprints, 5 of which were identified as chlorogenic acid, hyperin, isoquercitrin, trifolin and astragalin, with their content also determined. Except for 3 batches of medicinal materials, the similarity of other 28 batches was higher than 0.83, indicating good similarity. Two categories were formed in the cluster analysis based on content determination, which showed that some differences existed in similarities between different regions of Xinjiang. The medicinal materials were ranked by quality with principal component analysis, and the results indicated that the top 15 all came from northern Xinjiang. The quality difference of A. venetum and P. pictum had a correlation with the place of origin. This study provides a reference for the analysis and evaluation of A. venetum and P. pictum from different habitats and the selection of introduction and cultivation areas.
Apocynum ; China ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; Medicine, Chinese Traditional

Eight compounds,(R)-2-[5-(methoxycarbonyl)-4-methyl-6-oxo-3,6-dihydro-2H-pyran-2-yl]acetic acid(1),(3S,4R)-3,4-dihydro-3,4-epoxy-5-hydroxynaphthalen-1(2H)-one(2),(-)-mitorubrinol(3),(-)-mitorubrin(4),(±)-asperlone A(5), terreusinone(6), verrucisidinol(7) and cerebroside C(8) were isolated from the endophytic fungus Talaromyces purpurogenus by using various column chromatographic techniques. Their structures were identified by NMR, MS, CD and optical rotation. Compounds 1 and 2 were new compounds. Their anti-diabetic activities in vitro were evaluated, and compound 1 showed moderate inhibitory activity toward XOD at 10 μmol·L~(-1) with the inhibition rate of 69.9%.
Endophytes/chemistry* ; Hypoglycemic Agents/chemistry* ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Secondary Metabolism ; Talaromyces/chemistry* ; Tylophora/microbiology* ; Xanthine Oxidase/antagonists & inhibitors*

To build up an identification method on cardiac glycosides in Taxillus chinensis and its Nerium indicum host, and evaluate the influence on medicine quality from host to T. chinensis, ultra-performance liquid chromatography coupled with quadrupole-time-of-flight tandem mass-mass spectrometry(UPLC-Q-TOF-MS/MS)was applied. The samples of T. chinensis(harvested from N. indicum)and its N. indicum host were collected in field. The samples of T. chinensis(harvested from Morus alba)and its M. alba host was taken as control substance. All samples were extracted by ultrasonic extraction in 70% ethanol. Chromatographic separation was performed on an ACQUITY UPLC HSS T3 C_(18)(2.1 mm×100 mm,1.8 μm)column at 40 ℃. Gradient elution was applied, and the mobile phase was consisted of 0.1% formic acid water and acetonitrile. The 0.5 μL of sample solution was injected and the flow rate of the mobile phase was kept at 0.6 mL·min~(-1) in each run. It was done to identify cardiac glycosides and explore the chemical composition correlation in T. chinensis and its N. indicum host by analyzing positive and negative ion mode mass spectrometry data, elemental composition, cardiac glycoside reference substance and searching related literatures. A total of 29 cardiac glycosides were identified, 28 of it belonged to N. indicum host, 5 belonged to T. chinensis(harvested from N. indicum host), none of cardiac glycoside was identified in T. chinensis(harvested from M. alba host). The result could provide a reference in evaluating the influence in T. chinensis medicine quality from host. It was rapid, accurate, and comprehensive to identify cardiac glycosides in T. chinensis and its N. indicum host by UPLC-Q-TOF-MS/MS.
Cardiac Glycosides ; analysis ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; Loranthaceae ; chemistry ; Nerium ; chemistry ; Phytochemicals ; analysis ; Tandem Mass Spectrometry

The study aimed to investigate the mechanism of hepatoprotective effect of C-21 steroidal glucosides from Cynanchum auriculatum( Baishouwu) on oxidative stress in mice with liver injury. Mice were randomly divided into normal group,model group,positive control group,Baishouwu high group and Baishouwu low group. The liver injury model was induced by intraperitoneal injection of CCl4 peanut oil solution. All mice were sacrificed to collect blood and liver specimens. The activities of serum levels of ALT and AST were detected. The content of MDA and the activity of SOD in liver homogenate were examined by colorimetry method. Tissues were stained with hematoxylin-eosin for histological examination. The hepatic protein expressions of NF-κB p65,p-IκBα,i NOS and COX-2 were detected by Western blot. The mRNA expressions of TNF-α and IL-6 were determined by RT-PCR. It was found that treatment with C-21 steroidal glucosides from Baishouwu successfully attenuated liver injury induced by CCl4,as shown by decreased levels of serum biochemical indicators( AST,ALT)( P<0. 01). Administration of total C-21 steroidal glucosides enhanced the activity of SOD( P<0. 01) and decreased the content of MDA( P<0. 01) in liver homogenate. Microscopic features suggested that treatment with C-21 steroidal glucosides from Baishouwu was effective in inhibiting CCl4-induced hepatocyte edema and degeneration. Further studies showed that NF-κB p65 overexpression induced by CCl4 was decreased by C-21 steroidal glucosides,leading to the markedly down-regulated protein expression levels of p-IκBα,i NOS and COX-2,as well as the depression of TNF-α and IL-6 mRNA expressions. In conclusion,total C-21 steroidal glucosides from Baishouwu exhibited potent effect on oxidative stress pathway in mice with liver injury induced by CCl4,with enhanced activity of SOD,decreased content of MDA,and down-regulated levels of NF-κB p65,p-IκBα,i NOS and COX-2.
Animals ; Carbon Tetrachloride ; Chemical and Drug Induced Liver Injury ; drug therapy ; Cynanchum ; chemistry ; Glucosides ; pharmacology ; Hepatocytes ; drug effects ; Liver ; drug effects ; Mice ; Oxidative Stress ; Random Allocation