The healing effect of a mixture of Arnebia euchroma and animal fat on burn wounds in rats in comparison with sulfadiazine.

OBJECTIVE: Thermal burn is a serious cause of morbidity and mortality that affects millions of people worldwide. The aim of this experimental study was to investigate the efficacy of Arnebia euchroma (AE) to treat burn wounds in a rat model. METHOD: A total of 80 male rats (200-250g) were shaved over the back of the neck (2×3cm2) and a second-degree burn wound was induced at this site under general anaesthesia. The rats were then randomly assigned to one of four groups (each n=20) and the burns were treated daily for 14 days as follows: (1) dressed with animal fat; (2) dressed with sulfadiazine; (3) dressed with a mixture of AE and animal fat; (4) no treatment (control). Five rats from each group were sacrificed on days 3, 5, 9 and 14 post-burn and the wounds were evaluated histologically and immunohistochemically for the expression of interleukin (IL)-1 and IL-6. RESULTS: There was a significant increase at day 3 and decrease on day 5 samples for the expression of IL-1 in the AE plus fat group and IL-6 in the AE plus fat and sulfadiazine groups, compared to the control and fat treatment groups, respectively. Both AE plus fat and sulfadiazine treatments reduced inflammation and granulation tissue formation by day 5 post-burn, while re-epithelialisation commenced by day 9 post-burn. In addition, burns treated with AE plus fat exhibited keratinised epidermis, associated with regular collagen fibres, compared to moderately dense collagen fibres without vascularisation in the sulfadiazine group. CONCLUSION: These findings suggested that AE plus fat was superior to sulfadiazine in enhancing burn wound healing in rats.

Euchronin A-F isolated from the Arnebia euchroma (Royle) Johnst. and their anti-proliferative activities in vitro.

Six new naphthoquinones, euchronin A-F (1-6) and nine known naphthoquinones (7-15), were isolated from the roots of Arnebia euchroma (Royle) Johnst. The structures of the new compounds were confirmed by extensive spectroscopic analyses, including UV, IR, HR-ESI-MS, 1D and 2D NMR. In the present study, we estimated the anti-proliferative activities of these compounds with HaCaT cells. The results indicated that compounds 2 and 4 showed strong anti-proliferative activities at 25 µM, with relative viability at 38.83% and 68.44%, respectively.

ß-acetoxyisovalerylalkannin suppresses proliferation and induces ROS-based mitochondria-mediated apoptosis in human melanoma cells.

ß-acetoxyisovalerylalkannin (ß-AIVA) is one of shikonin/alkannin derivative, which were mainly extracted from Boraginaceae family. The effects of ß-AIVA on human melanoma A375 cells and U918 cells were investigated in vitro. The CCK-8 assay showed that ß-AIVA inhibited proliferation of cells. Results from flow cytometry, ROS assay and JC-1 assay showed that ß-AIVA increased late apoptosis rate, induced the production of ROS and promoted mitochondrial depolarization in cells. ß-AIVA regulated expressions of BAX and Bcl-2 proteins, and increased the expression of cleaved caspase-9 and cleaved caspase-3. These findings suggest that ß-AIVA may be a potential therapeutic drug for treating melanoma.

[Correlation between rhizosphere environment and content of medicinal components of Arnebia euchroma].

This study aimed to explore the correlation between rhizosphere soil microorganisms of wild Arnebia euchroma and the content of medicinal components to provide guidance for the selection of the ecological planting base. The total DNA of rhizosphere soil microorganisms of wild A. euchroma was extracted, and the microbial community structure of rhizosphere soil microorganisms was analyzed by IlluminaMiseq high-throughput sequencing technology. The content of total hydroxynaphthoquinone pigment and ß,ß'-dimethylacrylalkannin in medicinal materials was determined by high-performance liquid chromatography(HPLC). The physicochemical pro-perties of rhizosphere soil of wild A. euchroma in main producing areas were determined, and the correlation of soil microbial abundance with index component content and soil physicochemical properties was analyzed by SPSS software. The results showed that the species composition of rhizosphere fungi and bacteria in A. euchroma from different habitats was similar at the phylum and genus levels, but their relative abundance, richness index(Chao1), and community diversity(Simpson) index were different. Correlation analysis showed that the content of available phosphorus in soil was positively correlated with the content of total hydroxynaphthoquinone pigment and ß,ß'-dimethylacrylalkannin, and the abundance of five fungal genera such as Solicoccozyma and six bacterial genera such as Pseudo-nocardia and Bradyrhizobium was positively correlated with the content of medicinal components in medicinal materials. The abundance of Bradyrhizobium was significantly positively correlated with the content of ß,ß'-dimethylacrylalkanin. The abundance of fungi such as Archaeorhizomyces was significantly positively correlated with the content of available phosphorus in rhizosphere soil, and Bradyrhizobium was significantly negatively correlated with soil pH. Therefore, the abundance of fungi and bacteria in the rhizosphere of A. euchroma has a certain correlation with the medicinal components and the physicochemical properties of the rhizosphere soil, which can provide a scientific basis for the selection of ecological planting bases in the later stage.

[Effect of acetylalkannin from Arnebia euchroma on proliferation, migration, and invasion of human melanoma A375 cells].

This study aimed to explore the effect and mechanism of acetylalkannin from Arnebia euchroma on the proliferation, migration, and invasion of human melanoma A375 cells. A375 cells were divided into a blank group, and low-, medium-, and high-dose acetylalkannin groups(0.5, 1.0, and 2.0 µmol·L~(-1)). The MTT assay was used to detect cell proliferation. Cell scratch and transwell migration assays were used to detect cell migration ability, and the transwell invasion assay was used to detect cell invasion ability. Western blot was used to detect the protein expression of migration and invasion-related N-cadherin, vimentin, matrix metalloproteina-se-9(MMP-9), and Wnt/ß-catenin pathway-related Wnt1, Axin2, glycogen synthase kinase-3ß(GSK-3ß), phosphorylated GSK-3ß(p-GSK-3ß), ß-catenin, cell cycle protein D_1(cyclin D_1), and p21. Real-time fluorescence-based quantitative polymerase chain reaction(real-time PCR) was used to detect the mRNA expression of E-cadherin, matrix metalloproteinase-2(MMP-2), N-cadherin, vimentin, ß-catenin, snail-1, and CD44. MTT results showed that the cell inhibition rates in the acetylalkannin groups significantly increased as compared with that in the blank group(P<0.01). The results of cell scratch and transwell assays showed that compared with the blank group, the acetylalkannin groups showed reduced cell migration and invasion, and migration and invasion rates(P<0.05, P<0.01) and weakened horizontal and vertical migration and invasion abilities. Western blot results showed that compared with the blank group, the high-dose acetylalkannin group showed increased expression of Axin2 protein(P<0.05), and decreased expression of N-cadherin, vimentin, MMP-9, Wnt1, p-GSK-3ß, ß-catenin, cyclin D_1, and p21 proteins(P<0.05, P<0.01). The expression of GSK-3ß protein did not change significantly. PCR results showed that the overall trend of MMP-2, N-cadherin, vimentin, ß-catenin, snail-1, and CD44 mRNA expression was down-regulated(P<0.01), and the expression of E-cadherin mRNA increased(P<0.01). Acetylalkannin can inhibit the proliferation, migration, and invasion of human melanoma A375 cells, and its mechanism of action may be related to the regulation of Wnt/ß-catenin signaling pathway.

The discovery and characterization of AeHGO in the branching route from shikonin biosynthesis to shikonofuran in Arnebia euchroma.

Shikonin derivatives are natural naphthoquinone compounds and the main bioactive components produced by several boraginaceous plants, such as Lithospermum erythrorhizon and Arnebia euchroma. Phytochemical studies utilizing both L. erythrorhizon and A. euchroma cultured cells indicate the existence of a competing route branching out from the shikonin biosynthetic pathway to shikonofuran. A previous study has shown that the branch point is the transformation from (Z)-3''-hydroxy-geranylhydroquinone to an aldehyde intermediate (E)-3''-oxo-geranylhydroquinone. However, the gene encoding the oxidoreductase that catalyzes the branch reaction remains unidentified. In this study, we discovered a candidate gene belonging to the cinnamyl alcohol dehydrogenase family, AeHGO, through coexpression analysis of transcriptome data sets of shikonin-proficient and shikonin-deficient cell lines of A. euchroma. In biochemical assays, purified AeHGO protein reversibly oxidized (Z)-3''-hydroxy-geranylhydroquinone to produce (E)-3''-oxo-geranylhydroquinone followed by reversibly reducing (E)-3''-oxo-geranylhydroquinone to (E)-3''-hydroxy-geranylhydroquinone, resulting in an equilibrium mixture of the three compounds. Time course analysis and kinetic parameters showed that the reduction of (E)-3''-oxo-geranylhydroquinone was stereoselective and efficient in presence of NADPH, which determined that the overall reaction proceeded from (Z)-3''-hydroxy-geranylhydroquinone to (E)-3''-hydroxy-geranylhydroquinone. Considering that there is a competition between the accumulation of shikonin and shikonofuran derivatives in cultured plant cells, AeHGO is supposed to play an important role in the metabolic regulation of the shikonin biosynthetic pathway. Characterization of AeHGO should help expedite the development of metabolic engineering and synthetic biology toward production of shikonin derivatives.

Pyrrolizidine alkaloids and health risk of three Boraginaceae used in TCM.

Objective: The aim of this study was to systematically explore the pyrrolizidine alkaloids (PAs) type, content and risk assessment in the three Boraginaceae used in TCM, involving Arnebia euchroma (AE), A. guttata (AG), and Lithospermum erythrorhizon (LE). Method: A UHPLC-MS/MS method was established to simultaneously determine eight pyrrolizidine alkaloids (PAs), namely intermedine, lycopsamine, intermedine N-oxide, lycopsamine N-oxide, 7-acetyllycopsamine, 7-acetyllycopsamine N-oxide, echimidine N-oxide, and echimidine in the three herbs. Based on these results, the risk assessment was explored using the routine margin of exposure (MOE) combined with relative potency (REP) for oral and external usage, respectively. Results and Conclusion: Imermedine and imermedine N-oxide were common components in the eight tested PAs. 7-acetyllycopsamine and its N-oxide were not detected in AE; echimidine and its N-oxide were not detected in AG; lycopsamine and its N-oxide, 7-acetyllycopsamine and its N-oxide were not detected in LE. The total contents of 8 PAs in 11 batches of AG was341.56-519.51 µg/g; the content in 15 batches of LE was 71.16-515.73 µg/g, and the content in 11 batches of AE was 23.35-207.13 µg/g. Based on these results, the risk assessment was explored using MOE combined with REP for oral and external usage, respectively. The findings of the risk assessment method of PAs based on MOE combined with the REP factor were consistent with the clinical toxicity results. As an oral herb, AE had low risk or no risk due to its low PA contents, and individual batches of LE were medium risk, while attention should be paid to their clinical use.AG was also low risk. The external use of the three Boraginaceae used in TCM was not associated with any risk. This study systematically explored the PA type and content of the three Boraginaceae used in TCM. Additionally, the refined risk assessment of PAs based on REP provided a more scientific basis for quality evaluation and rational use of the medicinal Boraginaceae used in TCM to improve public health.

The Potential of Medicinal Plants and Natural Products in the Treatment of Burns and Sunburn-A Review.

Treating burns remains a challenge for modern medicine, especially in developing countries that cannot afford expensive, advanced therapies. This review article summarises clinical and animal model studies of botanical preparations and their mixtures in treating burn wounds and sunburn. Articles available in electronic databases such as PubMed, Scopus, Web of Science, Science Direct and Google Scholar, published in English in 2010-2022, were considered. In the described clinical trials, it was shown that some herbal preparations have better effectiveness in treating burn wounds, including shortening the healing time and reducing inflammation, than the conventional treatment used hitherto. These herbal preparations contained extracts from Albizia julibrissin, Alkanna tinctoria, Aloe vera, Arnebia euchroma, Betula pendula and Betula pubescens, Centella asiatica, Hippophaë rhamnoides, Juglans regia, Lawsonia inermis, and mixtures of Matricaria chamomilla and Rosa canina. Research on animal models shows that many extracts may potentially benefit the treatment of burn wounds and sunburn. Due to the diverse mechanism of action, antibacterial activity, the safety of use and cost-effectiveness, herbal preparations can compete with conventional treatment. The growing interest in alternative medicine and herbal medicine encourages further research. Not only single preparations but also their mixtures should be taken into account because the research conducted so far often suggests a synergistic effect of the ingredients.

Current Status and Medicinal Prominence of Arnebia euchroma (Ratanjot): A Critically Endangered Plant of Trans-Himalayan Region.

Trans-Himalayan region has been a major component of the India's opulent medicinal plant heritage that encompasses numerous critically endangered plant species. Arnebia euchroma (Royle ex Benth.) Johnston (common name: Ratanjot), a Trans- Himalayan native, is amongst them, and it belongs to the family Boraginaceae. Ratanjot has long been used as a colourant in food and cosmetics besides a major ingredient of traditional remedies prescribed for curing mild constipation, dermatitis, frostbite, and eczema like health disorders. Though principally harvested for its roots, almost all the parts of this plant have been used in pharmaceutical products, food, dyes and beverages since prehistoric times. Its roots are a rich source of naphthoquinone pigment(s) mainly shikonin, acetylshikonin and deoxyshikonin, accountable for its medicinal value as antimicrobial, wound healing, anti-inflammatory, anticancer, and antioxidant agent(s). Considering the medicinal importance and critically endangered status of this taxon, the need of the hour is to conserve and propagate it for supplying sufficient raw materials for its commercial exploitation.

Liver protection and hemostatic effects of medicinal plant Arnebia euchroma (Royle) I.M.Johnst extract in a rat model.

ETHNOPHARMACOLOGICAL RELEVANCE: Arnebia euchroma (Royle) I.M.Johnst. (AE) is a Chinese medicinal herb that is traditionally used to treat various circulatory diseases. It exhibits certain effects, such as the promotion of blood circulation and cooling, rash clearance, and detoxification. AIM OF THE STUDY: This study was designed to explore the hepatoprotective and hemostatic effects of the ethyl acetate extract of AE in rats with carbon tetrachloride (CCl4)-induced liver injury. MATERIALS AND METHODS: Wistar rats were treated via oral gavage with different doses of the ethyl acetate extract of AE (3.5, 7, or 14 g kg-1·day-1) for 14 consecutive days, following which hemostatic and liver function tests were conducted. For the hemostatic tests, the platelet count, blood platelet aggregation, blood platelet adhesion to fibrinogen, platelet factor 4 (PF-4) secretion from blood platelets, prothrombin time (PT), activated partial thromboplastin time (aPTT), thrombin time (TT), and fibrinogen levels were measured at the end of the treatment period. For the liver function tests, 0.25 mL/200 g (1.25 mL kg-1·day-1) of olive oil was injected into the abdominal cavity of the control rats, whereas 15% CCl4 plus olive oil (prescription: 7.5 mL CCl4 + 42.5 olive oil) was injected into that of the treated rats at 1 h after extract administration on day 6, 13, and 20. Additionally, food and water were withheld from all the animals. On the following day, the rats were anesthetized and their albumin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), gamma-glutamyl transpeptidase (GGT), lactate dehydrogenase (LDH), reactive oxygen species (ROS), methane dicarboxylic aldehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) levels were measured. Glutathione S-transferase (GST), glutathione reductase (GR), and glutathione peroxidase (GPx) levels among the groups were determined using a one-way analysis of variance. RESULTS: The platelet count and blood platelet aggregation, blood platelet adhesion to fibrinogen and PF-4 secretion levels were significantly increased in the (3.5 g kg-1 day-1) AE group as compared to those in the control group (all p < 0.001; for the 7 and 14 g kg-1 day-1 AE groups, all p > 0.05, respectively). Although the PT and aPTT were not affected by the AE extract (all p > 0.05), the TT was reduced and the FIB levels were significantly increased in all AE groups (p < 0.05). Liver function tests showed that CCl4 caused significant liver damage, thereby decreasing the albumin, SOD, CAT, GSH, GST, GR, and GPx levels, while increasing the AST, ALT, ALP, SGOT, SGPT, GGT, LDH, ROS, and MDA levels (all p < 0.001). By contrast, treatment with the different doses of AE extract reversed the CCl4 effects on all these parameters. Compared with the levels in the CCl4 group, the GSH and GR levels in the three AE groups (3.5, 7, and 14 g kg-1·day-1) were significantly higher (p < 0.05, p < 0.01, and p < 0.001, respectively), whereas the differences in the other parameters for these three groups were all at the significance levels of p < 0.05, p < 0.05, and p < 0.01, respectively. CONCLUSIONS: AE extracts administered orally exhibited hepatoprotective activity by affecting platelet production and blood coagulation and ameliorating liver function-damaging modifications. Specifically, a dosage of 3.5 g kg-1·day-1 resulted in the most optimal effects.

Shikonin derivatives as potent xanthine oxidase inhibitors: in-vitro study.

Induction of hypersensitivity reactions (may be fatal too) by specific XO inhibitors has led to development of new molecules that are efficacious and have safer ADME profile. Among natural compounds, biologically active Alkannin/Shikonin (A/S) derivatives have unexplored XO inhibition potential. Therefore, their iso-hexenylnaphthazarin nucleus was studied and found that the nucleus is similar to that of allopurinol, signifying the XO inhibitory potential of these derivatives. For confirmation of their potential, ß,ß-dimethylacrylshikonin and deoxyshikonin were successfully isolated and characterised from Arnebia euchroma (Royle.) Johnst. (Boraginaceae) and were evaluated for in vitro XO inhibitory potential. ß,ß-dimethylacrylshikonin and deoxyshikonin showed a good XO inhibition potential with IC50 values of 7.475 ± 1.46 µg/mL and 4.487 ± 0.88 µg/mL, respectively. Results also validated the pharmacophore hypothesis, and it was concluded that nucleus iso-hexenylnaphthazarin can be remodelled for optimising the efficacy.

Effects of extracts from Arnebia euchroma (Royle) Johnst on apoptosis of HepG2 and its anti-hepatocellular carcinoma in mice / 中国药理学通报

Aim To study the apoptosis of human hep-atoma cell line ( HepG2 ) induced by different polar parts of Arnebia euchroma ( Royle ) Johnst ( AE ) and to verify its anti-hepatoma effect by a mouse orthotopic liver cancer model so as to explore the anti-cancer effect of AE extract. Methods Firstly, MTT method and Annexin V-FITC/PI double staining method were used to detect the anti-proliferative and pro-apoptotic effects of each polar part of AE on HepG2 cells, and Western blot was used to detect the expression of Bcl-2 apoptosis family proteins incells. Based on the above experimental results, the effective parts with significant pro-apoptotic effect were screened out for anti-in situ liver cancer experiments in mice, and the organ indexes, liver function indexes and tissue sections of mice with orthotopic liver cancer before and after administration were evaluated. Results With the decrease of the polarity of AE extract,the anti-proliferation and pro-apoptotic effects on HepG2 cells were enhanced, and the anti-proliferation and apoptosis-inducing effects of AE petroleum ether fraction ( AEP) were the most significant. When AEP dose was 1.56 (μg • L

Correlation between rhizosphere environment and content of medicinal components of Arnebia euchroma / 中国中药杂志

This study aimed to explore the correlation between rhizosphere soil microorganisms of wild Arnebia euchroma and the content of medicinal components to provide guidance for the selection of the ecological planting base. The total DNA of rhizosphere soil microorganisms of wild A. euchroma was extracted, and the microbial community structure of rhizosphere soil microorganisms was analyzed by IlluminaMiseq high-throughput sequencing technology. The content of total hydroxynaphthoquinone pigment and β,β'-dimethylacrylalkannin in medicinal materials was determined by high-performance liquid chromatography(HPLC). The physicochemical pro-perties of rhizosphere soil of wild A. euchroma in main producing areas were determined, and the correlation of soil microbial abundance with index component content and soil physicochemical properties was analyzed by SPSS software. The results showed that the species composition of rhizosphere fungi and bacteria in A. euchroma from different habitats was similar at the phylum and genus levels, but their relative abundance, richness index(Chao1), and community diversity(Simpson) index were different. Correlation analysis showed that the content of available phosphorus in soil was positively correlated with the content of total hydroxynaphthoquinone pigment and β,β'-dimethylacrylalkannin, and the abundance of five fungal genera such as Solicoccozyma and six bacterial genera such as Pseudo-nocardia and Bradyrhizobium was positively correlated with the content of medicinal components in medicinal materials. The abundance of Bradyrhizobium was significantly positively correlated with the content of β,β'-dimethylacrylalkanin. The abundance of fungi such as Archaeorhizomyces was significantly positively correlated with the content of available phosphorus in rhizosphere soil, and Bradyrhizobium was significantly negatively correlated with soil pH. Therefore, the abundance of fungi and bacteria in the rhizosphere of A. euchroma has a certain correlation with the medicinal components and the physicochemical properties of the rhizosphere soil, which can provide a scientific basis for the selection of ecological planting bases in the later stage.

Effect of acetylalkannin from Arnebia euchroma on proliferation, migration, and invasion of human melanoma A375 cells / 中国中药杂志

This study aimed to explore the effect and mechanism of acetylalkannin from Arnebia euchroma on the proliferation, migration, and invasion of human melanoma A375 cells. A375 cells were divided into a blank group, and low-, medium-, and high-dose acetylalkannin groups(0.5, 1.0, and 2.0 μmol·L~(-1)). The MTT assay was used to detect cell proliferation. Cell scratch and transwell migration assays were used to detect cell migration ability, and the transwell invasion assay was used to detect cell invasion ability. Western blot was used to detect the protein expression of migration and invasion-related N-cadherin, vimentin, matrix metalloproteina-se-9(MMP-9), and Wnt/β-catenin pathway-related Wnt1, Axin2, glycogen synthase kinase-3β(GSK-3β), phosphorylated GSK-3β(p-GSK-3β), β-catenin, cell cycle protein D_1(cyclin D_1), and p21. Real-time fluorescence-based quantitative polymerase chain reaction(real-time PCR) was used to detect the mRNA expression of E-cadherin, matrix metalloproteinase-2(MMP-2), N-cadherin, vimentin, β-catenin, snail-1, and CD44. MTT results showed that the cell inhibition rates in the acetylalkannin groups significantly increased as compared with that in the blank group(P<0.01). The results of cell scratch and transwell assays showed that compared with the blank group, the acetylalkannin groups showed reduced cell migration and invasion, and migration and invasion rates(P<0.05, P<0.01) and weakened horizontal and vertical migration and invasion abilities. Western blot results showed that compared with the blank group, the high-dose acetylalkannin group showed increased expression of Axin2 protein(P<0.05), and decreased expression of N-cadherin, vimentin, MMP-9, Wnt1, p-GSK-3β, β-catenin, cyclin D_1, and p21 proteins(P<0.05, P<0.01). The expression of GSK-3β protein did not change significantly. PCR results showed that the overall trend of MMP-2, N-cadherin, vimentin, β-catenin, snail-1, and CD44 mRNA expression was down-regulated(P<0.01), and the expression of E-cadherin mRNA increased(P<0.01). Acetylalkannin can inhibit the proliferation, migration, and invasion of human melanoma A375 cells, and its mechanism of action may be related to the regulation of Wnt/β-catenin signaling pathway.

Naphthoquinones and triterpenoids from Arnebia euchroma (Royle) Johnst and their hypoglycemic and lipid-lowering effects.

A new pentacyclic triterpenoid, 2-hydroxy-1-ene-hydroxyhopanone (19), and a new benzoxepin-5-one, 3-(4-methyl-3-penten-1-yl)-6-hydroxy-9-methoxy-2H-1-benzoxepin-5-one (25), along with 26 known compounds (1-18, 20-24, 26-28), were isolated from the roots of Arnebia euchroma (Royle) Johnst. The structures of the new compounds were elucidated by extensive spectroscopic analyses. The absolute configurations of shikonofurans 9-13 were determined by quantum chemical ECD calculations and CD spectra comparison for the first time. Pharmacological study revealed that naphthoquinones 1-5, 7, and 8 had obvious cytotoxicity toward human lung adenocarcinoma A549 cell line. Meanwhile, the hypoglycemic and lipid-lowering effects of isolated compounds were assessed by checking their inhibitory effects on key enzymes regulating glucose and lipid metabolism. Results showed that compounds 1, 3, 5, 6, 8, 18, and 19 could inhibit the activity of ATP-citrate lyase (ACL); compound 7 could inhibit the activity of acetyl-CoA carboxylase (ACC1); while compounds 8 and 19 showed inhibitory effects on protein tyrosine phosphatase 1B (PTP1B). Among them, the naphthoquinone 6, steroid 18, and triterpenoid 19 showed moderate inhibitory effects on ACL and PTP1B, but didn't exhibit obvious cytotoxicity. This study demonstrated that compounds 6, 18, and 19 show great promising for the development of new agents for the treatment of metabolic diseases.

Eight new arnebinol B-based meroterpenoids with planar chirality in the constrained 6/10/5 tricyclic skeleton from Arnebia euchroma and their cytotoxicities.

Eight new arnebinol B-based meroterpenoids ((-)-1, 2, 3, (-)-5, and 7-10) with a constrained 6/10/5 tricyclic backbone were isolated from the roots of Arnebia euchroma. The planar and steric structures of these new compounds were unambiguously elucidated by extensive spectroscopic analyses, X-ray diffraction crystallography, and ECD calculations. The predominant relative orientation between H-7 and the Z double bond with a methyl substituent in the rigid 10-membered carbocycle, along with the planar chirality of the Z double bond was analyzed and discussed for the first time. The illustration of the planar chirality derived from the Z double bond should be paid great importance during the structure elucidation on these homologous meroterpenoids. All the isolated meroterpenoids were screened for their cytotoxicities against the HCT-8, PANC-1, HGC-27, HepG2, and PC9 cell lines, and compounds (+)-5 and (-)-5 exhibited the most potent cytotoxicity.

Phylogenomics and Genetic Diversity of Arnebiae Radix and Its Allies (Arnebia, Boraginaceae) in China.

Arnebiae Radix is a traditional medicine with pleiotropic properties that has been used for several 100 years. There are five species of Arnebia in China, and the two species Arnebia euchroma and Arnebia guttata are the source plants of Arnebiae Radix according to the Chinese Pharmacopoeia. Molecular markers that permit species identification and facilitate studies of the genetic diversity and divergence of the wild populations of these two source plants have not yet been developed. Here, we sequenced the chloroplast genomes of 56 samples of five Arnebia species using genome skimming methods. The Arnebia chloroplast genomes exhibited quadripartite structures with lengths from 149,539 and 152,040 bp. Three variable markers (rps16-trnQ, ndhF-rpl32, and ycf1b) were identified, and these markers exhibited more variable sites than universal chloroplast markers. The phylogenetic relationships among the five Arnebia species were completely resolved using the whole chloroplast genome sequences. Arnebia arose during the Oligocene and diversified in the middle Miocene; this coincided with two geological events during the late Oligocene and early Miocene: warming and the progressive uplift of Tianshan and the Himalayas. Our analyses revealed that A. euchroma and A. guttata have high levels of genetic diversity and comprise two and three subclades, respectively. The two clades of A. euchroma exhibited significant genetic differences and diverged at 10.18 Ma in the middle Miocene. Three clades of A. guttata diverged in the Pleistocene. The results provided new insight into evolutionary history of Arnebia species and promoted the conservation and exploitation of A. euchroma and A. guttata.

Mechanism of Arnebia euchroma Against Melanoma： An Exploration Based on Network Pharmacology and Experimental Verification / 中国实验方剂学杂志

ObjectiveTo preliminarily predict the active components， action targets， and signaling pathways of Arnebia euchroma in the treatment of melanoma based on network pharmacology and molecular docking， and to verify its possible mechanism of action in in vitro experiments. MethodThe active components and related targets of A. euchroma were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）SwissTargetPrediction and literature， and the targets related to melanoma from the GeneCards， Online Mendelian Inheritance in Man （OMIM）， and Comparative Toxicogenomics Database （CTD）. Following the construction of the protein-protein interaction （PPI） network of active components and related targets of A. euchroma and melanoma-related targets using STRING， Cytoscape 3.8.2 was used for screening and analyzing the nodes in the network of A. euchroma against melanoma. The intersections were subjected to gene ontology （GO） and Kyoto encyclopedia of genes and genomes （KEGG） enrichment analysis using DAVID 6.8. Acetyl alkannin， the active component in A. euchroma， was docked to the target by AutoDock Vina 1.1.2. The in vitro experiments were then carried out to verify the anti-melanoma effect of A. euchroma. ResultA total of 271 common targets of A. euchroma and melanoma were harvested， among which 23 were key targets， including matrix metalloproteinase-9 （MMP-9） and Janus kinase 2 （JAK2）. As revealed by KEGG enrichment analysis， A. euchroma mainly acted on Janus kinase/signal transduction and activator of transcription （JAK/STAT）， tyrosine kinase receptor （ErbB）， and vascular endothelial growth factor （VEGF） signaling pathways to resist melanoma. According to molecular docking， acetyl alkannin exhibited a good docking activity with JAK2， STAT3， VEGF， MMP-9， and E-cadherin receptors. The results of Western blot and Real-time quantitative polymerase chain reaction （Real-time PCR） showed that acetyl alkannin at different doses inhibited the protein and gene expression of JAK2， STAT3， VEGF， MMP-9， and E-cadherin in A375 cells （P<0.05）. ConclusionA. euchroma alleviates melanoma via multiple targets and multiple pathways， and it may exert the therapeutic effects by affecting the expression of such key target proteins as JAK2， STAT3， VEGF， MMP-9， and E-cadherin and inhibiting the invasion and metastasis of melanoma cells. This study has provided an experimental basis for the treatment of tumor with A. euchroma.

De novo transcriptome analysis provides insights into formation of in vitro adventitious root from leaf explants of Arnebia euchroma.

BACKGROUND: Adventitious root formation is considered a major developmental step during the propagation of difficult to root plants, especially in horticultural crops. Recently, adventitious roots induced through plant tissue culture methods have also been used for production of phytochemicals such as flavonoids, anthocyanins and anthraquinones. It is rather well understood which horticultural species will easily form adventitious roots, but the factors affecting this process at molecular level or regulating the induction process in in vitro conditions are far less known. The present study was conducted to identify transcripts involved in in vitro induction and formation of adventitious roots using Arnebia euchroma leaves at different time points (intact leaf (control), 3 h, 12 h, 24 h, 3 d, 7 d, 10 d and 15 d). A. euchroma is an endangered medicinal Himalayan herb whose root contains red naphthoquinone pigments. These phytoconstituents are widely used as an herbal ingredient in Asian traditional medicine as well as natural colouring agent in food and cosmetics. RESULTS: A total of 137.93 to 293.76 million raw reads were generated and assembled to 54,587 transcripts with average length of 1512.27 bps and N50 of 2193 bps, respectively. In addition, 50,107 differentially expressed genes were identified and found to be involved in plant hormone signal transduction, cell wall modification and wound induced mitogen activated protein kinase signalling. The data exhibited dominance of auxin responsive (AUXIN RESPONSE FACTOR8, IAA13, GRETCHEN HAGEN3.1) and sucrose translocation (BETA-31 FRUCTOFURANOSIDASE and MONOSACCHARIDE-SENSING protein1) genes during induction phase. In the initiation phase, the expression of LATERAL ORGAN BOUNDARIES DOMAIN16, EXPANSIN-B15, ENDOGLUCANASE25 and LEUCINE-rich repeat EXTENSION-like proteins was increased. During the expression phase, the same transcripts, with exception of LATERAL ORGAN BOUNDARIES DOMAIN16 were identified. Overall, the transcriptomic analysis revealed a similar patterns of genes, however, their expression level varied in subsequent phases of in vitro adventitious root formation in A. euchroma. CONCLUSION: The results presented here will be helpful in understanding key regulators of in vitro adventitious root development in Arnebia species, which may be deployed in the future for phytochemical production at a commercial scale.

Investigating the hemostatic effect of medicinal plant Arnebia euchroma (Royle) I.M.Johnst extract in a mouse model.

ETHNOPHARMACOLOGICAL RELEVANCE: Arnebia euchroma (Royle) I.M.Johnst (AE) has been reported to be a potentially useful medicinal herb for the treatment of several circulatory diseases in traditional Chinese medicine. It shows effects such as "cooling of the blood," promotion of blood circulation, detoxification, and rash clearance. AIM OF THE STUDY: To explore the hemostatic effect of the ethyl acetate extract of AE in mice. MATERIALS AND METHODS: In this study, we explored the effects of AE on bleeding time, blood coagulation time, platelet count, and blood coagulation parameters in normal Kunming mice. Different doses of the AE extract (5, 10, and 20 g kg-1·day-1) were administered to mice for 14 days. Sodium carboxymethyl cellulose (CMC-Na at 0.5%) and Yunnan Baiyao (0.8 g kg-1·day-1) were administered as negative and positive control treatments, respectively. Bleeding time, blood coagulation time, platelet count, blood platelet aggregation, blood platelet adhesion to fibrinogen, platelet factor 4 (PF-4) secretions from blood platelets, and blood coagulation parameters including prothrombin time (PT), activated partial thromboplastin time (aPTT), thrombin time (TT), and fibrinogen (FIB) levels were measured on day 15 of administration. RESULTS: Bleeding and blood coagulation time were significantly lower and TT was shorter in the AE extract-treated groups than in the control groups. Furthermore, FIB levels and platelet count were higher, whereas blood platelet aggregation, blood platelet adhesion to fibrinogen, and PF-4 secretion from blood platelets were more obvious in the AE extract-treated groups than in the control group. However, no significant differences were detected for PT and aPTT between the extract-treated and control groups. CONCLUSIONS: The ethyl acetate extract of AE showed potential hemostasis effects in mice by shortening the bleeding and coagulation time. In addition, the extract increased platelet count and induced blood platelet aggregation, blood platelet adhesion to fibrinogen, PF-4 secretion from blood platelets, and FIB level, while it shortened TT.