Comprehensive metabolome characterization and comparison between two sources of Dragon's blood by integrating liquid chromatography/mass spectrometry and chemometrics.

Dragon's Blood (DB) serves as a precious Chinese medicine facilitating blood circulation and stasis dispersion. Daemonorops draco (D. draco; Qi-Lin-Jie) and Dracaena cochinchinensis (D. cochinchinenesis; Long-Xue-Jie) are two reputable plant sources for preparing DB. This work was designed to comprehensively characterize and compare the metabolome differences between D. draco and D. cochinchinenesis, by integrating liquid chromatography/mass spectrometry and untargeted metabolomics analysis. Offline two-dimensional liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (2D-LC/IM-QTOF-MS), by utilizing a powerful hybrid scan approach, was elaborated for multicomponent characterization. Configuration of an XBridge Amide column and an HSS T3 column in offline mode exhibited high orthogonality (A0 0.80) in separating the complex components in DB. Particularly, the hybrid high-definition MSE-high definition data-dependent acquisition (HDMSE-HDDDA) in both positive and negative ion modes was applied for data acquisition. Streamlined intelligent data processing facilitated by the UNIFI™ (Waters) bioinformatics platform and searching against an in-house chemical library (recording 223 known compounds) enabled efficient structural elucidation. We could characterize 285 components, including 143 from D. draco and 174 from D. cochinchinensis. Holistic comparison of the metabolomes among 21 batches of DB samples by the untargeted metabolomics workflows unveiled 43 significantly differential components. Separately, four and three components were considered as the marker compounds for identifying D. draco and D. cochinchinenesis, respectively. Conclusively, the chemical composition and metabolomic differences of two DB resources were investigated by a dimension-enhanced analytical approach, with the results being beneficial to quality control and the differentiated clinical application of DB.

New flavan trimer from Daemonorops draco as osteoclastogenesis inhibitor.

Dragon's blood is a red resin obtained from different plants and is considered highly efficacious and used in medicine owing its wound healing function. Two new compounds (7 and 8) were isolated from the dragon's blood of Daemonorops draco fruits, along with eight known compounds (1-6, 9, and 10). Their structures, including their absolute configurations, were elucidated by nuclear magnetic resonance (NMR), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and electronic circular dichroism (ECD) analysis. According to the spectroscopic data, 8 was determined to be a quinone methide derivative of flavan and 7 was deduced to be a flavan trimer. All compounds were evaluated for their anti-osteoclastogenesis activity, compound 1 and 7 exhibited anti-osteoclastogenesis activity with IC50 values of 31.3 and 36.8 µM, respectively.

Retro-dihydrochalcones from the resin of Daemonorops draco and their inhibitory activities against renal fibrosis.

Daedracoflavan A-E (1-5), five new flavonoids were isolated from the resin of Daemonorops draco. Their structures including absolute configurations were established by using spectroscopic and computational methods. All the compounds are new chalcones with the same retro-dihydrochalcone skeleton. Compound 1 features the presence of a cyclohexadienone unit originating from a benzene ring, and the ketone group of C-9 reduced to a hydroxyl group. The bioactivity of all isolated compounds was evaluated in kidney fibrosis and found that compound 2 could dose-dependently inhibit the expression of fibronectin, collagen I, and α-SMA in TGF-ß1-induced rat kidney proximal tubular cells (NRK-52E). Interestingly, the replacement of a proton by a hydroxyl group at C-4' seems to play a crucial role in anti-renal fibrosis activity.

Platelet-inhibitory phenolic constituents from the fruits of Daemonorops draco.

Eight previously undescribed phenolic compounds, dracoropins A - H (1-8), along with two known analogues (9 and 10) were isolated from the fruits of Daemonorops draco. Four pairs of isomers (1a/1b, 2a/2b, 3a/3b, and 4a/4b) were resolved by using chiral-phase HPLC separation. Their structures, including the absolute configurations of the resolved isomers, were elucidated by analysis of spectroscopic data (1D and 2D NMR, IR, and HRESIMS), single-crystal X-ray diffraction, and electronic circular dichroism (ECD) calculations. Compounds 1, 2, and 3 bear a rare 2-phenylbenzo[d]-1,3-dioxepine skeleton. All the isolates were evaluated for their inhibitory activity against ATP release in thrombin-activated platelets. Compounds 2b, 3a, and 6 could significantly inhibit ATP release in thrombin-activated platelets.

Analyzing lignin biosynthesis pathways in rattan using improved co-expression networks of NACs and MYBs.

BACKGROUND: The rattan is a valuable plant resource with multiple applications in tropical forests. Calamus simplicifolius and Daemonorops jenkinsiana are the two most representative rattan species, supplying over 95% of the raw materials for the rattan industry. Hence, the wood properties of both rattans have always attracted researchers' attention. RESULTS: We re-annotated the genomes, obtained 81 RNA-Seq datasets, and developed an improved pipeline to increase the reliability of co-expression networks of both rattans. Based on the data and pipeline, co-expression relationships were detected in 11 NACs, 49 MYBs, and 86 lignin biosynthesis genes in C. simplicifolius and four NACs, 59 MYBs, and 76 lignin biosynthesis genes in D. jenkinsiana, respectively. Among these co-expression pairs, several genes had a close relationship to the development of wood properties. Additionally, we detected the enzyme gene on the lignin biosynthesis pathway was regulated by either NAC or MYB, while LACCASES was regulated by both NAC and MYB. For D. jenkinsiana, the lignin biosynthesis regulatory network was characterized by positive regulation, and MYB possible negatively regulate non-expressed lignin biosynthesis genes in stem tissues. For C. simplicifolius, NAC may positively regulate highly expressed genes and negatively regulate non-expressed lignin biosynthesis genes in stem tissues. Furthermore, we established core regulatory networks of NAC and MYB for both rattans. CONCLUSIONS: This work improved the accuracy of rattan gene annotation by integrating an efficient co-expression network analysis pipeline, enhancing gene coverage and accuracy of the constructed network, and facilitating an understanding of co-expression relationships among NAC, MYB, and lignin biosynthesis genes in rattan and other plants.

Daemonorops draco Blume Induces Apoptosis Against Acute Myeloid Leukemia Cells via Regulation of the miR-216b/c-Jun.

Daemonorops draco Blume (DD), also called dragon's blood, has been used as a traditional Korean medicine, especially for relieving pain caused by wound infection. Recently, it has been described that DD has antibacterial and analgesic effects. In this study, the underlying anticancer effect of DD associated with apoptosis was investigated in acute myeloid leukemia cell lines U937 and THP-1. DD exhibited cytotoxic effects and induced apoptosis in U937 and THP-1 cells. Moreover, DD treatment significantly reduced mitochondrial membrane potential (ΔΨ). The protein expression of cleaved poly(ADP-ribose) polymerase, cleaved caspase-3, p-H2A.X, CCAAT/enhancer-binding protein (CHOP), and activating transcription factor 4 was upregulated by DD treatment. Consistently, DD-treated cells had increased reactive oxygen species (ROS) level in a concentration-dependent manner via miR-216b activation in association with c-Jun inhibition. N-acetyl-L-cysteine pretreatment reversed the cytotoxic effect of DD treatment as well as prevented ROS accumulation. Collectively, the results of this study suggest that the anticancer effect of DD in AML was mediated by CHOP-dependent apoptosis along with ROS accumulation and included upregulation of miR-216b followed by a decrease in c-Jun.

Anti-inflammatory, pro-proliferative and antimicrobial potential of the compounds isolated from Daemonorops draco (Willd.) Blume.

ETHNO-PHARMACOLOGICAL RELEVANCE: Daemonorops draco (D. draco) commonly known as "Dragon's blood" is one of the most used plants by Momok, Anak Dalam and Talang Mamak tribes from Indonesia as a remedy for wound healing. AIM OF THE STUDY: This study aimed to identify the extract, fractions and compounds responsible for the anti-inflammatory and pro-proliferative activities of the D. draco resin. Additionally, the antimicrobial activity against two bacteria and one yeast species was analysed. MATERIALS AND METHODS: Bio-guided isolation of compounds with anti-inflammatory, pro-proliferative and antimicrobial activities from the D. draco resin was carried out by measuring: the inhibition of NF-κB and activation of Nrf2 in THP-1, HaCaT, NIH-3T3 cells; cell proliferation in NIH-3T3 and HaCaT cells; and the antimicrobial effect on E. coli, S. aureus and C. albicans. RESULTS: Guided isolation by bioassay gave rise to the isolation and characterisation by nuclear magnetic resonance and mass spectrometry of three compounds: 1 (Bexarotene), 2 (Taspine) and 3 (2-hydroxy-1-naphthaldehyde isonicotinoyl hydrazone). All compounds showed NF-κB inhibitory activity with IC50 values of 0.10-0.13, 0.22-0.24 and 3.75-4.78 µM, respectively, while the positive control, Celastrol, had an IC50 of 7.96 µM. Likewise, all compounds showed an activating effect of Nrf2 with EC50 values of 5.34-5.43, 163.20-169.20 and 300.82-315.56 nM, respectively, while the positive control, CDDO-Me, had an EC50 of 0.11 nM. In addition, concerning the pro-proliferative activity, compound 1 (IC50 = 8.62-8.71 nM) showed a capacity of 100%, compound 2 (IC50 = 166-171 nM) showed a capacity of 75%, and compound 3 (IC50 = 469-486 nM) showed a capacity of 65%, while FSB 10% (positive control) had a pro-proliferative activity of 100% in the NIH3T3 cell lines (fibroblasts) and HaCaT (keratinocytes). Finally, all the compounds showed antimicrobial activity with MIC values of 0.12-0.16, 0.31-0.39 and 3.96-3.99 µM, respectively, in S. aureus, E. coli and C. albicans strains, while the positive control, Ofloxacin, had a MIC of 27.65 µM. CONCLUSION: This study managed to isolate, for the first time, three compounds (Bexarotene, Taspine and 2-hydroxy-1-naphthaldehyde isonicotinoyl hydrazone) from the resin of D. draco, with anti-inflammatory, and pro-proliferative as well as antimicrobial activities.

Evaluation of antioxidant activity and phytochemicals of selected methanol rattan shoot extracts from Morong, Bataan

Background@#Rattan is a member of the Arecaceae family grown in the tropical or subtropical climates. These plants are used as raw materials for furniture-making. In some cases, its shoots are also used as food and may possess bioactive potentials. In this study, rattan root methanolic extracts were screened for phytochemicals and evaluated for antioxidant activities.@*Methodology@#Six (6) methanolic extracts of selected rattan shoots were subjected to in vitro antioxidant assays, which include DPPH, FRAP, metal chelation, superoxide, and nitric oxide. Qualitative and quantitative phytochemical analyses were also employed. @*Results@#Shoots of Calamus sp. 02 (Bangaw-Bangaw; 85.59±0.430%), C. usitatus (Talola; 82.45±2.215%), and C. spinifolius (Kurakling; 75.54±1.599%) showed the most effective DPP radical inhibition at 66.67μg/mL. Concentration-dependent reducing power (>30% scavenging at 6.64μg/mL) with no-to-low metal chelating activity was also observed in these plant food. C. sp 02 and C. usitatus demonstrated an effective scavenging activity against superoxide anions at 227.3μg/mL. Nitric oxide scavenging activity was observed in all shoots with C. merrilli (Palasan) exhibiting highest at 78.13μg/mL. Qualitative phytochemical analyses showed that rattan shoots all contained reducing sugars, phenolics, terpenoids, and quinone compounds. Quantitative phytochemical analyses revealed that C. sp. 02 (66.024±4.183mg GAE/g) and C. merrilli (1.767±0.056mg QE/g) contained the highest amounts of phenolic and flavonoids, respectively. These phytochemicals present may explain their behavior as antioxidants. @*Conclusion@#The study revealed that different rattan shoots showed different capacities to scavenge particular oxidants. Of these, C. sp. 02, C. spinifolius, and C. merrilli may be considered promising sources of natural antioxidants.

Coexpression Analysis Reveals Dynamic Modules Regulating the Growth and Development of Cirri in the Rattans (Calamus simplicifolius and Daemonorops jenkinsiana).

Rattan is regarded as one of the major non-timber forest products, second only to wood and bamboo, worldwide. Although the published genomes of Calamus simplicifolius and Daemonorops jenkinsiana have facilitated genome-wide gene functional analyses, coexpression networks (CENs) provide more comprehensive and complete annotations of gene function at the transcriptome level. Thus, we analyzed the CENs of the two rattans, C. simplicifolius and D. jenkinsiana, by integrating the genome sequences and analyzing in-house transcriptome data from different development stages of their cirri using a well-developed strategy. A total of 3,504 and 3,027 functional modules were identified in C. simplicifolius and D. jenkinsiana, respectively, based on a combination of CENs, gene family classification, and function enrichment tools. These modules covered the major developmental processes, including photosynthesis, lignin biosynthesis, flavonoid biosynthesis, and phenylpropanoid biosynthesis. Reference annotations were refined using CENs and functional modules. Moreover, we obtained novel insights into the regulation of cirrus growth and development in rattans. Furthermore, Rattan-NET (http://rattan.bamboogdb.org/), an online database with analysis tools for gene set enrichment analysis, module enrichment, network comparison analysis, and cis-element analysis, was constructed for the easy analysis of gene function and regulation modules involved in the growth and development of cirri in rattans.

Eight new flavonoids from the fruits of Daemonorops draco.

Eight new flavonoids daemoflavans A-H, including two dimeric proanthocyanidins (1 and 2), four flavans (3-6), two 2-arylbenzofurans (7 and 8), along with nine known compounds (9-17), were isolated from the fruit of Daemonorops draco. Their structures, including the absolute configurations, were elucidated by extensive spectroscopic data, ECD analysis, and X-ray crystal diffraction. Besides, the X-ray crystal data of a known compound dracoflavan B1 (9) was firstly reported. Daemoflavan G (7) represents a rare example of C-5 methylated 2-arylbenzofuran in natural products. Among the known compounds, 15, 16, 17 were reported from this species for the first time. All the compounds were evaluated for their cytotoxicity against HepG2 cell line. Among them, compounds 1, 9 and 10 exhibited modest cytotoxic activity with IC50 values of 12.4, 12.0 and 13.2 µM, respectively.

Identification and structure-activity relationship (SAR) of chemical constituents from Daemonorops draco (Willd.) Blume and selected commercial flavonoids on anti-osteoclastogenesis activity.

Osteoclastogenesis-related bone diseases including osteoporosis, rheumatoid arthritis, Paget's disease and periodontitis are worldwide occurred and cause severe health problems including bone fracture and bone cancer. However, A few studies have shown that Daemonorops draco (Willd.) Blume may decrease bone destruction and relieve bone cancer pain. In this research, we isolated and purified four known and two novel compounds from D. draco and investigated their anti-osteoclastogenesis activity using RAW264.7 cells. Among them, com.1 exhibited the most effective inhibitory activity on osteoclastogenesis with 78% inhibition at 10 µM and identified to be a novel natural flavan; and com.2 displayed a bit slighter inhibition (50% at 10 µM), indicating that the methylation of 7-hydroxyl group increased the anti-osteoclastogenesis activity. Moreover, nineteen commercial flavonoids were also performed in this study to investigate their inhibitory activity on osteoclastogenesis, and furtherly develop the SAR profile in flavonoid skeleton combined with the information of isolated compounds. Interestingly, the absence of substituents in B-ring and (3R)-hydroxyl group seems to play a crucial role in increasing anti-osteoclastogenesis activity.

Rattan (Calamoideae) Diversity and Biomass Change in Different Habitat Types During Two Years.

Rattans (subfamily Calamoideae) are an important component of the forests of the Old World. However, few studies have been conducted on the distribution of these abundant palms within different habitats, specifically in Peninsular Malaysia. This study was aimed at assessing rattan diversity, abundance and biomass change across two different habitat types, namely, dipterocarp forests and fresh-water swamps within the Segari Melintang Forest Reserve, Perak, within two years. All rattan stems within five 100 m × 100 m sized study plots (A-E) of the two habitat types were counted in 2011 and 2013, and Shannon-Wiener diversity indices (H') and Bray-Curtis dissimilarity indices were calculated. A total of 11 species from 5 genera (161 stems ha-1) were sampled. Rattan abundance was higher in all swamp plots; however, rattan diversity (H') was highest in the dipterocarp plot (D: H' (2011)1.79; H' (2013)1.84). Bray-Curtis indices of rattan abundance (highest similarity in swamp: plot BC(2011) 0.484, BC(2013) 0.262) and biomass were highest for study plots with the same vegetation types in both years. For rattan biomass, the most similar plot pairs changed during the years: dipterocarp plots A and D were most similar in 2011 (0.509), and swamp plots B and C were most similar in 2013 (0.282). This study helped contribute information regarding the distribution and dynamics of rattans in a primary rainforest of Peninsular Malaysia.

Phenolic group on A-ring is key for dracoflavan B as a selective noncompetitive inhibitor of α-amylase.

A high throughput assay was applied to guide the isolation of a new pancreatic α-amylase inhibitor, dracoflavan B, from the dragon's blood resin from Daemonorops draco. Applying C18 column, we successfully isolated both diastereomers and their structures verified by (1)H NMR spectra in comparison with the literature values. Their activity in inhibition of pancreatic α-amylase with comparable IC50 values of 23µM (A type) and 27µM (B type) that are similar to that of acarbose. Dracoflavan B shows much weaker activity in inhibiting bacterial α-amylase and no activity towards fungal α-amylase. Moreover, both isomers show no inhibitory activity towards mammalian α-glucosidase. Kinetic analysis revealed that using starch as the substrate, dracoflavan B was a non-competitive α-amylase inhibitor with a Ki value of 11.7µM. Lack of α-amylase inhibition for proanthocyanidin A2 dimer demonstrated that dracoflavan B hydrophobic nature of the B, A', C' and B' rings are important for its α-amylase inhibition. In addition, selective chemical modification studies revealed that the phenolic group is also vital to dracoflavan B for its pancreatic α-amylase inhibition activity. Without the A ring phenolic hydrogen bond donor, the derivatives of dracoflavan B showed detrimental α-amylase inhibition. On the contrary, galloylation on the A ring phenolic OH group enhanced the activity as shown by the low IC50 (12µM) against α-amylase which is 56% more potent as compared to dracoflavan B.

Three new flavans in dragon's blood from Daemonorops draco.

Three new flavans were isolated from chloroform extracts of dragon's blood from Daemonorops draco, together with eight known compounds. The structures of the new flavans were determined by 1D and 2D NMR spectroscopic analysis. These compounds are the first examples of 2-methoxyflavans from D. draco and regarded as derivatives of biogenetic intermediates from flavans to chalcones, which are characteristic of the dragon's blood.