Chemical constituents of the aerial parts of Mitracarpus hirtus (L.) DC (Rubiaceae).

A phytochemical investigation of the aerial parts of Mitracarpus hirtus afforded thirteen compounds, including a new naphthoquinone di-glycoside (1), three isopentenyl isoflavones (2-4), four flavonoids (5-8), three iridoid glycosides (9 - 11) and two coumarins (12 and 13). Their structures were elucidated based on extensive spectroscopic analyses, chemical methods, and the comparison with the literature. Among them, compound 1 possesses a 2-(3-methylnaphthalen-2-yl)acetic acid core with two glucosyl groups, compounds 2-4 are the first three representatives from the Rubiaceae family, and compounds 9-11 and 13 were isolated from Mitracarpus genus for the first time. Additionally, compounds 2-4 displayed potent antibacterial activities against Helicobacter pylori G27/HP159/JRES00015 (MIC = 4-16 µg/mL) , comparable to metronidazole. To date, wighteone (2) is the most active isoflavone with favourable predicted ADMET properties reported against H. pylori.

Acetogenins from the Stem of Uvaria rufa and Their Cytotoxic Activity.

Four new adjacent bis-tetrahydrofuran acetogenins, bullacin C (7), uvarirufin (9), and uvariasolins III (12) and IV (13), along with 11 known acetogenins, were isolated from the stem of Uvaria rufa. Their structures were elucidated based on spectroscopic data analysis, including 1D and 2D NMR, HRESIMS, and MALDI-MS/MS of the lithium adducts. Absolute configurations were assigned using Mosher ester analysis and ECD measurements. Uvarirufin (9) possesses a unique C-39 skeleton among acetogenins. Most tested acetogenins exhibited cytotoxicity against human cancer cell lines (HCT 116, 22Rv1, MDA-MB-435, OVCAR3). Squamocin (8) and uvarirufin (9) were found to be the most potent, with an IC50 value of 1.2 µM for both in HCT 116 colon cancer cells. Additionally, a new application of Dragendorff's reagent is proposed herein for the TLC detection of acetogenins.

Phenylpropanoid-enriched broccoli seedling extract can reduce inflammatory markers and pain behavior.

BACKGROUND: Pain is a worldwide problem requiring an effective, affordable, non-addictive therapy. Using the edible plant broccoli, a growth protocol was developed to induce a concentrated combinatorial of potential anti-inflammatories in seedlings. METHODS: A growth method was utilized to produce a phenylpropanoid-rich broccoli sprout extract, referred to as Original Extract (OE). OE was concentrated and then resuspended for study of the effects on inflammation events. A rabbit disc model of inflammation and degeneration, and, a mouse model of pain behavior were used for in vivo and in vitro tests. To address aspects of mammalian metabolic processing, the OE was treated with the S9 liver microsome fraction derived from mouse, for use in a mouse in vivo study. Analytical chemistry was performed to identify major chemical species. Continuous variables were analyzed with a number of methods including ANOVA, and two-tailed t tests, as appropriate. RESULTS: In a rabbit spine (disc) injury model, inflammatory markers were reduced, and levels of regenerative markers were increased as a result of OE treatment, both in vivo and in vitro. In a mouse pain behavioral model, after treatment with S9 liver microsome fraction, the resultant extract significantly reduced early and late pain behavior in response to a pain stimulus. The OE itself reduced pain behavior in the mouse pain model, but did not achieve the level of significance observed for S9-treated extract. Analytical chemistry undertaken on the extract constituents revealed identities of the chemical species in OE, and how S9 liver microsome fraction treatment altered species identities and proportions. CONCLUSIONS: In vitro and in vivo results indicate that the OE, and S9-treated OE broccoli extracts are worthwhile materials to develop a non-opiate inflammation and pain-reducing treatment.

Polyoxygenated cyclohexene derivatives and other constituents of Uvaria rufa stem.

Six undescribed compounds, uvarirufols D and E, (+)-uvarigranol B, (-)-uvarigranol E, 6-acetoxy-5-hydroxy-7-methoxyflavanone and cherrevenaphthalene D, along with twelve known compounds, including polyoxygenated cyclohexenes, flavonoids, and lignans, were isolated from the methanol extract of Uvaria rufa stems. Their structures were elucidated by spectroscopic analyses and the absolute configurations were determined using electronic circular dichroism. Several isolates were evaluated for cytotoxic, antitubercular and anti-inflammatory potentials. (-)-6-Acetylzeylenol showed moderate inhibitory activity against Mycobacterium tuberculosis, with MIC value of 47.10 µg/mL. Cherrevenaphthalene D exhibited weak antimycobacterial activity and potent inhibitory effect on lipopolysaccharide-induced nitric oxide (NO) production in RAW 264.7 cells (EC50 = 8.54 µM). 8-Hydroxy-5,7-dimethoxyflavanone displayed moderate level of NO inhibition (EC50 = 43.62 µM) with little cytotoxicity. The polyoxygenated cyclohexenes and lignans were inactive against HCT 116 and 22Rv1 cancer cells (IC50 > 100 µM).

Structurally diverse (9ß-H)-pimarane derivatives with six frameworks from the leaves of Icacina oliviformis and their cytotoxic activities.

Thirteen previously undescribed (9ß-H)-pimarane derivatives, icacinolides A-G (1-7) and oliviformislactones C-H (8-13), together with four known analogs (14-17), were isolated from the leaves of Icacina oliviformis. Their structures were constructed by extensive spectroscopic analysis, 13C NMR-DP4+ analysis, ECD calculation, single-crystal X-ray diffraction, and chemical methods. These structurally diverse isolates were classified into six framework types: rearranged 3-epi-17-nor-(9ß-H)-pimarane, rearranged 17-nor-(9ß-H)-pimarane, 16-nor-(9ß-H)-pimarane, 17-nor-(9ß-H)-pimarane, 17,19-di-nor-(9ß-H)-pimarane, and (9ß-H)-pimarane. Among them, compounds 1, 5, and 7 were the first examples of three rearranged 3-epi-17-nor-(9ß-H)-pimaranes featuring a unique (11S)-carboxyl-9-oxatricyclo[5.3.1.02,7]dodecane motif with contiguous stereogenic centers, whereas their C-3 epimers, compounds 2-4 and 6 were the second examples of four rearranged 17-nor-(9ß-H)-pimaranes. Additionally, compounds 8 and 12/13 represented the second examples of a 16-nor-(9ß-H)-pimarane and two 17,19-di-nor-(9ß-H)-pimaranes, respectively. In cytotoxic bioassay, compound 2 exhibited significant cytotoxic against HT-29 with IC50 values of 7.88 µM, even stronger than 5-fluorouracil, and 15 showed broad-spectrum cytotoxic activities against HepG2, HT-29, and MIA PaCa-2 with IC50 values of 11.62, 9.77, and 4.91 µM, respectively. Meanwhile, a preliminary structure-activity relationship suggested that 3,20-epoxy, 6,19-lactone, 2-OH, 7-OH, and 8-OH in (9ß-H)-pimarane derivatives might be active groups, whereas ring C aromatization may decrease the cytotoxic activities.

Identification of small molecule inhibitors against MMP-14 via High-Throughput screening.

Matrix metalloproteinases (MMPs) are involved in various cellular events in physiology and pathophysiology through endopeptidases activity. The expression levels and activities of most MMPs remain minimal in the normal conditions, whereas some MMPs are significantly activated in pathological conditions such as cancer and neovascularization. Hence, MMPs are considered as both diagnostic markers and potential targets for therapeutic agents. Twenty-three known human MMPs share a similar active site structure with a zinc-binding motif, resulting in lack of specificity. Therefore, the enhancement of target specificity is a primary goal for the development of specific MMP inhibitors. MMP-14 regulates VEGFA/VEGFR2-system through cleavage of the non-functional VEGFR1 in vascular angiogenesis. In this study, we developed a fluorescence-based enzymatic assay using a specific MMP-14 substrate generated from VEGFR1 cleavage site. This well optimized assay was used as a primary screen method to identify MMP-14 specific inhibitors from 1,200 Prestwick FDA-approved drug library. Of ten initial hits, two compounds showed IC50 values below 30 µM, which were further validated by direct binding analysis using surface plasmon resonance (SPR). Clioquinol and chloroxine, both of which contain a quinoline structure, were identified as MMP-14 inhibitors. Five analogs were tested, four of which were found to be completely devoid of inhibitory activity. Clioquinol exhibited selectivity towards MMP-14, as it showed no inhibitory activity towards four other MMPs.

Chemical constituents of the stem of Marsypopetalum modestum and their bioactivities.

Phytochemical investigation of Marsypopetalum modestum (Annonaceae) led to the isolation of a new phenylpropanoid glycoside, lyciumphenylpropanoid B (10), along with nine known compounds (1-9) from an aqueous methanolic extract of the stem. Most compounds are reported from this genus for the first time. The structures of the isolated compounds were elucidated using spectroscopic methods including NMR spectroscopy, high-resolution mass spectrometry, and quantum chemical electronic circular dichroism (ECD) calculations. Cytotoxic and antitubercular activities of several isolated compounds were evaluated. Dipyrithione (1) displayed anti-mycobacterial (MIC = 0.23 µM) and cytotoxic (IC50 = 0.8 µM in Hep G2 cells; 4.1 µM in HCT 116 cells) activities. Kelampayoside A (8) showed moderate cytotoxic activity against cancer cells.

Oxalactam A, a Novel Macrolactam with Potent Anti-Rhizoctonia solani Activity from the Endophytic Fungus Penicillium oxalicum.

A novel macrolactam named oxalactam A (1), three known dipeptides (2-4) as well as other known alkaloids (5-7) were obtained from the endophytic fungus Penicillium oxalicum, which was derived from the tuber of Icacina trichantha (Icacinaceae). All chemical structures were established based on spectroscopic data, chemical methods, ECD calculations, and 13C-DP4+ analysis. Among them, oxalactam A (1) is a 16-membered polyenic macrolactam bearing a new skeleton of 2,9-dimethyl-azacyclohexadecane core and exhibited potent anti-Rhizoctonia solani activity with a MIC value of 10 µg/mL in vitro. The plausible biosynthetic pathway of 1 was also proposed via the alanyl protecting mechanism. Notably, three dipeptides (2-4) were first identified from the endophytic fungus P. oxalicum and the NMR data of cyclo(L-Trp-L-Glu) (2) was reported for the first time. In addition, the binding interactions between compound 1 and the sterol 14α-demethylase enzyme (CYP51) were studied by molecular docking and dynamics technologies, and the results revealed that the 16-membered polyenic macrolactam could be a promising CYP51 inhibitor to develop as a new anti-Rhizoctonia solani fungicide.

UHPLC-MS/MS-GNPS based phytochemical investigation of Dryopteris ramosa (Hope) C. Chr. and evaluation of cytotoxicity against liver and prostate cancer cell lines.

Dryopteris ramosa (family; Dryopteridaceae) has been reported for its medicinal importance in cancer, gastrointestinal disorders, and infections. The present study aimed to investigate the detailed phytochemical profile of D. ramosa and its cytotoxic potential using various cancer cell lines. The phytochemical profile of D. ramosa methanolic extract and its fractions were established by employing UHPLC-MS/MS and Global Natural Product Social (GNPS) molecular networking. Moreover, the cytotoxic activity of extract and fractions was evaluated against human liver (HepG-2) and prostate cancer (PC-3) cells using MTT assay. Overall, 18 compounds including flavonoids, flavonoid O-glycosides, isoflavone di-C-glycoside, flavanol, flavanone, rotenoid, phloroglucinol derivative, coumarin derivative, benzofuranone, abietic acid, and phenolic acid were observed as the major phytochemical bioactive constituents in the extract and fractions of D. ramosa. In MTT assay, chloroform fraction showed highest anti-proliferative activity against liver cancer cells (IC50 = 53.49 µg/mL) followed by n-hexane fraction (IC50 = 55.36 µg/mL), D. ramosa extract (IC50 = 85.67 µg/mL) and ethyl acetate (IC50 = 125.00 µg/mL) fraction. However, n-hexane and chloroform fractions presented maximum cytotoxic effect against prostate cancer cells with respective IC50 values of 214.53 and 281.47 µg/mL. Moreover, all the tested samples showed negligible toxicity against non-cancer (BHK-21) cells. The results indicated that D. ramosa is rich in flavonoids, phloroglucinol derivative, and phenolic acids and showed positive results in cytotoxic studies, especially against liver cancer. Therefore, it can be considered safe for the development of anticancer drugs, especially against liver cancer.

Pinostrobin: An Adipogenic Suppressor from Fingerroot (Boesenbergia rotunda) and Its Possible Mechanisms.

Obesity is a critical factor for chronic metabolic syndromes. The culinary plant fingerroot (Boesenbergia rotunda) has been reported for its anti-obesity activity. The anti-adipogenic effects of pandurantin A, a main component of fingerroot cultivated in Indonesia, have been studied. Nevertheless, the suppressive effect and related mechanisms of pinostrobin, a major constituent of Thai fingerroot, on adipogenesis have never been thoroughly investigated. This study aimed to evaluate the potential of pinostrobin to inhibit adipocyte differentiation. Culturing pre-adipocytes from both mouse (3T3-L1) and human (PCS-210-010) with pinostrobin at non-toxic concentrations (5-20 µM) for 48 h obviously hindered their differentiation into mature adipocyte as evidenced by reduced cellular lipid droplets. The lower levels of lipid metabolism-mediating proteins, namely C/EBPα, PPARγ, and SREBP-1c, as well as cellular triglyceride content were demonstrated in pinostrobin-treated 3T3-L1 cells when compared to the untreated control group. Additionally, pinostrobin modulated the signals of MAPK (p38 and JNK) and Akt (Akt/GSK3ß, Akt/AMPKα-ACC). These findings suggest the benefit of fingerroot as a source of phytopharmaceuticals for obesity prevention and management, with pinostrobin as the active principle.

Chemical constituents of Entandrophragma angolense and their anti-inflammatory activity.

From the stem bark of Entandrophragma angolense, six undescribed compounds were isolated, including seco-tirucallane type triterpenoids, limonoids, and a catechin glucoside, along with nineteen known structures. All structures were determined by interpretation of spectroscopic and HRMS data, and absolute configuration was confirmed with the aid of electronic circular dichroism. The isolated compounds were tested for LPS-induced NO inhibition in RAW 264.7 macrophages and EC50 values for moluccensin O and (-)-catechin were 81 µM and 137 µM, respectively.

A New C22-Quassinoid with Anti-Pancreatic Adenocarcinoma Activity from Seeds of Brucea javanica.

An undescribed C22-quassinoid named sergeolide A (1) and fifteen known quassinoids (2-16) were obtained from the seeds of Brucea javanica (Simaroubaceae). All chemical structures were established based on spectroscopic data and X-ray diffraction analysis. Sergeolide A (1) is the first example of a naturally occurring C22-quassinoid bearing a butenolide group fused the A ring of the bruceolide skeleton from Brucea genus. And this is the first report of the NMR data for desmethyl-bruceines B (2) and C (3) and the crystal structure for bruceolide (11). In addition, all isolates were evaluated for their anti-pancreatic adenocarcinoma activity by measuring the growth inhibitory of the MIA PaCa-2 cell lines. Consequently, compounds 1, 7-10, and 12-16 exhibited potent anti-pancreatic cancer activity in vitro (IC50 =0.054∼0.357 µM).

UHPLC-MS/MS-GNPS based phytochemical investigation of Equisetum arvense L. And evaluation of cytotoxicity against human melanoma and ovarian cancer cells.

Equisetum arvense L. is widely used as a traditional medicine for the management of inflammation and cancer. In the present study, phyto-chemical analysis of E. arvense was carried out and its cytotoxic potential against human melanoma (MDA-MB-435) and ovarian cancer cells (OVCAR3) was evaluated. Phyto-chemical profile of E. arvense methanolic extract and its fractions was established employing UHPLC-MS/MS and Global Natural Product Social molecular networking. Cytotoxic activity was evaluated using absorbance assay (CellTiter-Blue® Cell Viability Assay). Overall, 22 compounds were identified in the crude extract and polarity-based fractions of E. arvense. Flavonoids, flavonoid-O-glycosides and phenolic acids were found to be the major classes of phyto-chemicals. In addition, the crude extract of E. arvense and its fractions were found active against the tested cell lines. The highest anti-cancer activity against OVCAR3 cells was exhibited by the n-hexane fraction. These results indicated that E. arvense is rich in flavonoids and might be used for the development of anti-cancer drugs against melanoma and ovarian cancers.

Quinone Derivatives as Promising Anti-Helicobacter pylori Agents from Aerial Parts of Mitracarpus hirtus.

Seven new naphthoquinone diglycosides (1-7), three new anthraquinones (8-10), and eight known analogues were obtained from the aerial parts of Mitracarpus hirtus collected from West Africa in a bioassay-guided phytochemical investigation. All isolated compounds were elucidated by comparison with the literature and interpretation of spectroscopic data, and the absolute configurations of the new naphthoquinone diglycosides (1-10) were confirmed by chemical methods and ECD calculations. Notably, compound 1 was found to be the first naphthoquinone diglycoside containing carboxylic acid and isopentenyl side chains isolated from a species in the genus Mitracarpus. Compounds 6-18 showed antibacterial activity against multiple Helicobacter pylori strains with MIC values ranging from 0.0625 to 64 µg/mL. Particularly, 1-hydroxybenzoisochromanquinone (17) and benzo[g]isoquinoline-5,10-dione (18), with MIC values of 0.0625 and 0.125 µg/mL, displayed 32-512-fold higher potencies than a positive control, metronidazole. Compound 18 also demonstrated high antibiofilm activity and killed biofilm-encased Helicobacter pylori cells more effectively than metronidazole.

Angiogenesis-Inhibitory Piperidine Alkaloids from the Leaves of Microcos paniculata.

Eight new 2,6-disubstituted piperidin-3-ol alkaloids (1-8), featuring a C10 unsaturated alkyl side chain, together with three previously reported analogues (9-11) were isolated from the leaves of medicinal plant Microcos paniculata. Their structures and absolute configurations were elucidated unambiguously by means of 1D and 2D NMR spectroscopic data analysis, modified Mosher's method, Snatzke's method, and quantum chemical electronic circular dichroism (ECD) calculations, as well as single-crystal X-ray crystallography. The isolates were evaluated for their antiangiogenic effects on human umbilical vein endothelial cells (HUVECs). Compound 2 displayed an inhibitory effect on tube formation of HUVECs in a concentration-dependent manner.

Alkaloid-rich fraction of Ervatamia coronaria sensitizes colorectal cancer through modulating AMPK and mTOR signalling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Ervatamia coronaria, a popular garden plant in India and some other parts of the world is known traditionally for its anti-inflammatory and anti-cancer properties. The molecular bases of these functions remain poorly understood. AIM OF THE STUDY: Efficacies of the existing therapies for colorectal cancer (CRC) are limited by their life-threatening side effects and unaffordability. Therefore, identifying a safer, efficient, and affordable therapeutic is urgent. We studied the anti-CRC activity of an alkaloid-rich fraction of E. coronaria leaf extracts (AFE) and associated underlying mechanism. MATERIALS AND METHODS: Activity guided solvant fractionation was adopted to identify the activity in AFE. Different cell lines, and tumor grown in syngeneic mice were used to understand the anti-CRC effect. Methodologies such as LCMS, MTT, RT-qPCR, immunoblot, immunohistochemistry were employed to understand the molecular basis of its activity. RESULTS: We showed that AFE, which carries about six major compounds, is highly toxic to colorectal cancer (CRC) cells. AFE induced cell cycle arrest at G1 phase and p21 and p27 genes, while those of CDK2, CDK-4, cyclin-D, and cyclin-E genes were downregulated in HCT116 cells. It predominantly induced apoptosis in HCT116p53+/+ cells while the HCT116p53-/- cells under the same treatment condition died by autophagy. Notably, AFE induced upregulation of AMPK phosphorylation, and inhibition of both of the mTOR complexes as indicated by inhibition of phosphorylation of S6K1, 4EBP1, and AKT. Furthermore, AFE inhibited mTOR-driven conversion of cells from reversible cell cycle arrest to senescence (geroconversion) as well as ERK activity. AFE activity was independent of ROS produced, and did not primarily target the cellular DNA or cytoskeleton. AFE also efficiently regressed CT26-derived solid tumor in Balb/c mice acting alone or in synergy with 5FU through inducing autophagy as a major mechanism of action as indicated by upregulation of Beclin 1 and phospho-AMPK, and inhibition of phospho-S6K1 levels in the tumor tissue lysates. CONCLUSION: AFE induced CRC death through activation of both apoptotic and autophagy pathways without affecting the normal cells. This study provided a logical basis for consideration of AFE in future therapy regimen to overcome the limitations associated with existing anti-CRC chemotherapy.

Identification of Small Molecule Inhibitors against Staphylococcus aureus Dihydroorotase via HTS.

Drug-resistant Staphylococcus aureus is an imminent threat to public health, increasing the importance of drug discovery utilizing unexplored bacterial pathways and enzyme targets. De novo pyrimidine biosynthesis is a specialized, highly conserved pathway implicated in both the survival and virulence of several clinically relevant pathogens. Class I dihydroorotase (DHOase) is a separate and distinct enzyme present in gram positive bacteria (i.e., S. aureus, B. anthracis) that converts carbamoyl-aspartate (Ca-asp) to dihydroorotate (DHO)-an integral step in the de novo pyrimidine biosynthesis pathway. This study sets forth a high-throughput screening (HTS) of 3000 fragment compounds by a colorimetry-based enzymatic assay as a primary screen, identifying small molecule inhibitors of S. aureus DHOase (SaDHOase), followed by hit validation with a direct binding analysis using surface plasmon resonance (SPR). Competition SPR studies of six hit compounds and eight additional analogs with the substrate Ca-asp determined the best compound to be a competitive inhibitor with a KD value of 11 µM, which is 10-fold tighter than Ca-asp. Preliminary structure-activity relationship (SAR) provides the foundation for further structure-based antimicrobial inhibitor design against S. aureus.

A novel P38α MAPK activator Bruceine A exhibits potent anti-pancreatic cancer activity.

Pancreatic cancer remains one of the cancers with the poorest prognosis bearing an overall 5-year survival rate of about 5%. Efficient new chemotherapic drugs are still highly desired. Here, bruceine A, a quassinoid identified from the dried fruits of Brucea javanica (L.) Merr., displayed the most potent anti-proliferation activity against pancreatic cancer in vitro and in vivo. Phosphoproteomic analysis revealed p38α MAPK phosphorylation was involved in bruceine A's action in MIA PaCa-2 cells. Utilizing fortebio octet system and microscale thermophoresis, we found p38α MAPK had high affinity for bruceine A. Molecular docking and molecular dynamic simulations showed that bruceine A widely bound to residues (Leu171, Ala172, Met179, Thr180, Val183) in P-loop of p38α MAPK. Key determinants of bruceine A binding with P-loop of p38α MAPK were 19-C[bond, double bond]O, 22-CH3, 32-CH3, and 34-CH3. Taken together, our findings demonstrate that bruceine A binds directly to p38α MAPK, which can be used to probe the role of p38α MAPK phosphorylation in pancreatic cancer progression, and as a novel lead compound for pancreatic cancer therapy.

(9ßH)- and 17-Nor-Pimaranes from Icacina oliviformis.

Eleven pimarane-type diterpenoids were isolated from the tubers of Icacina oliviformis, including three new compounds, icacinlactone M (9), icacinlactone H 2-O-ß-d-glucopyranoside (10), and icacinlactone N 3-O-ß-d-glucopyranoside (11), together with an artifact of acrenol (8). Among the known structures, icacinlactone A (2), icacinlactone B (3), icacinlactone H (4), 12-hydroxyicacinlactone A (5), 14α-methoxyhumirianthol (6), and annonalide (7) are reported from I. oliviformis for the first time, whereas icacinol (1) has previously been found in this plant. Icacinol, 14α-methoxyhumirianthol, and annonalide displayed moderate cytotoxic activity in a panel of human cancer cell lines.