Respiratory and C4-photosynthetic NAD-malic enzyme coexist in bundle sheath cell mitochondria and evolved via association of differentially adapted subunits.

In plant mitochondria, nicotinamide adenine dinucleotide-malic enzyme (NAD-ME) has a housekeeping function in malate respiration. In different plant lineages, NAD-ME was independently co-opted in C4 photosynthesis. In the C4 Cleome species, Gynandropsis gynandra and Cleome angustifolia, all NAD-ME genes (NAD-MEα, NAD-MEß1, and NAD-MEß2) were affected by C4 evolution and are expressed at higher levels than their orthologs in the C3 species Tarenaya hassleriana. In T. hassleriana, the NAD-ME housekeeping function is performed by two heteromers, NAD-MEα/ß1 and NAD-MEα/ß2, with similar biochemical properties. In both C4 species, this role is restricted to NAD-MEα/ß2. In the C4 species, NAD-MEα/ß1 is exclusively present in the leaves, where it accounts for most of the enzymatic activity. Gynandropsis gynandra NAD-MEα/ß1 (GgNAD-MEα/ß1) exhibits high catalytic efficiency and is differentially activated by the C4 intermediate aspartate, confirming its role as the C4-decarboxylase. During C4 evolution, NAD-MEß1 lost its catalytic activity; its contribution to the enzymatic activity results from a stabilizing effect on the associated α-subunit and the acquisition of regulatory properties. We conclude that in bundle sheath cell mitochondria of C4 species, the functions of NAD-ME as C4 photosynthetic decarboxylase and as a housekeeping enzyme coexist and are performed by isoforms that combine the same α-subunit with differentially adapted ß-subunits.

Morphoanatomy, Histochemistry, and Phytochemistry of Stem and Leaves of Crateva tapia L.

Crateva tapia L. occurs only in Brazil and belongs to the family Capparaceae. This study aimed to characterize the morphoanatomy and histochemistry of stem and leaves, and phytochemical tests of the leaf of C. tapia. Macroscopic characterization was performed with the aid of a pachymeter and a stereomicroscope. Cross-sections of the stem, petiole, petiolule, and leaf blade, as well as paradermal sections of the leaf blade were mounted on microscope slides. Analyzes were carried out with optical microscopy. For histochemical analysis, different reagents were useda ccording to the targeted metabolite. Phytochemical tests of the methanolic extracts of the leaves were performed using thin layer chromatography. Crateva tapia has a stem with a circular cross-section and smooth bark. The leaves are alternate spirally, compound 3-foliolate. The leaf blade is elliptical and has a reticulate dictiodromous nerve. Through anatomical characterization, it was possible to identify a hypostomatic leaf blade with anisocytic, tetracytic, and anomocytic stomata and the presence of secretory canals. Through histochemistry and phytochemical tests, it was possible to observe the presence of different metabolites. Thus, the results obtained in this study contribute to the quality control of C. tapia, a species only found in Brazil, increasing the taxonomic knowledge of the family Capparaceae.

Eight Indole Alkaloids from the Roots of Maerua siamensis and Their Nitric Oxide Inhibitory Effects.

Maerua siamensis (Capparaceae) roots are used for treating pain and inflammation in traditional Thai medicine. Eight new indole alkaloids, named maeruanitriles A and B, maeroximes A-C, and maeruabisindoles A-C, were isolated from them. Spectroscopic methods and computational analysis were applied to determine the structure of the isolated compounds. Maeroximes A-C possesses an unusual O-methyloxime moiety. The bisindole alkaloid maeruabisindoles A and B possess a rare azete ring, whereas maeruabisindole C is the first indolo[3,2-b]carbazole derivative found in this plant family. Five compounds [maeruanitriles A and B, maeroxime C, maeruabisindoles B, and C] displayed anti-inflammatory activity by inhibiting nitric oxide (NO) production in the lipopolysaccharide-induced RAW 264.7 cells. Maeruabisindole B was the most active inhibitor of NO production, with an IC50 of 31.1 ± 1.8 µM compared to indomethacin (IC50 = 150.0 ± 16.0 µM) as the positive control.

A Rare Dirhamnosyl Flavonoid and Other Radical-Scavenging Metabolites from Cynophalla mollis (Kunth) J.†Presl and Colicodendron scabridum (Kunt) Seem. (Capparaceae) of Ecuador.

The phytochemistry of Cynophalla mollis (Kunth) J. Presl and Colicodendron scabridum (Kunth), both belonging to the family Capparaceae, were investigated in this study for the first time. Lupeol, betulin, lutein, stachydrine and quercetin-3,4'-di-O-rhamnoside were isolated from C. mollis, whereas C. scabridum afforded lupeol, lutein, stachydrine, ß-sitosterol, stigmasterol, betonicine and narcissoside. All these compounds were purified by preparative liquid chromatography, in both open column and instrumental (MPLC) separation systems. Preparative TLC was also applied. They were all identified by 1 H- and 13 C-NMR experiments. The complete structure of the very rare flavonoid quercetin-3,4'-di-O-rhamnoside was fully elucidated through DEPT-135, COSY, HMQC and HMBC experiments, together with UV/VIS and FT-IR spectrophotometry. Complete NMR data for quercetin-3,4'-di-O-rhamnoside in deuterated methanol were presented here for the first time. All the extracts did not exert antioxidant activity at the maximum tested dose of 1 mg/mL. Three out of the nine isolated compounds exerted a good spectrum of antioxidant capacity, being narcissoside the most active against ABTS radicals, with SC50 =12.43 µM. It was followed by lutein and quercetin-3,4'-di-O-rhamnoside, with 40.92 µM and 46.10 µM, respectively.

Daucosterol from Crateva adansonii DC (Capparaceae) reduces 7,12-dimethylbenz(a)anthracene-induced mammary tumors in Wistar rats.

This study aimed to evaluate the in vivo anticancer effects of daucosterol which was earlier reported to possess in vitro anticancer effects. Breast tumor was induced in 30 rats using the environmental carcinogen 7,12-dimethylbenz(a)anthracene (DMBA) while 6 control rats received olive oil (NOR). Animals with palpable tumors were randomized into five groups (n = 6) each as follows: negative control group treated with the vehicle (DMBA); positive control group treated with 5 mg/kg BW doxorubicin (DOXO + DMBA); three groups treated with daucosterol at doses of 2.5, 5, and 10 mg/kg BW (DAU + DMBA). Treatment lasted 28 days afterward, tumor (mass, volume, cancer antigen [CA] 15-3 level and histoarchitecture), hematological and toxicological parameters were examined. The tumor volume gradually increased in the DMBA group during the 28 days, with a tumor volume gain of ∼390 cm3 . Daucosterol at all doses reduced tumor volume (∼133.7 cm3 at 10 mg/kg) as well as protein, malondialdehyde (MDA), and CA 15-3 levels compared to DMBA rats. Tumor sections in daucosterol-treated rats showed a lower proliferation of mammary ducts with mild (5 and 10 mg/kg) to moderate (2.5 mg/kg) inflammatory responses. Moreover, it exhibited an antioxidant effect, evidenced by a significant and dose-dependent decreased in MDA levels, as well as an increase in catalase activity compared to the DMBA group. Daucosterol showed for the first time in vivo antitumor effects that corroborate its previous in vitro effects.

A randomised, double-blind, placebo-controlled clinical trial assessing the efficacy of bedtime buddy® for the treatment of nocturnal enuresis in children.

BACKGROUND: Nocturnal enuresis (NE), or 'bedwetting', is a form of night-time urinary incontinence occurring in younger children. A diagnosis of NE can be socially disruptive and psychologically stressful for a child. The most common strategies used by parents to manage NE are waking the child during the night to use the bathroom and limiting the child's water intake before going to bed. Behavioural or educational therapies for NE such as urotherapy or bladder retraining are widely accepted and considered as a mainstream treatment option for non-neurogenic lower urinary tract dysfunction in children. Pharmacotherapy also plays an ancillary role. However, there is no gold standard therapy or intervention to effectively manage NE. METHODS: This study aims to determine the efficacy of a herbal combination in the treatment of NE in children. The target population for this study is 80 children aged between 6 and 14 years old (males and females) who have primary nocturnal enuresis ≥3 per week (wet nights). The active group will receive one or two capsules per day containing 420 mg of a proprietary blend of Urox® (Seipel Group, Brisbane, Australia) containing Cratevox™ (Crataeva nurvala L; Capparidaceae; Varuna) stem bark extract standardised for 1.5% lupeol: non-standardised Equisetum arvense L. (Equisetaceae; Horsetail) stem extract; and, non-standardised Lindera aggregata Sims. The primary outcome for this study is the frequency of nocturia. Secondary outcomes include safety, quality of life, and daytime incontinence. Each participation will be involved in the trial for 32 weeks including contact with the research team every 2 weeks for the first 8 weeks and then every 8 weeks until trial completion. DISCUSSION: This study examines a novel treatment for an under-researched health condition affecting many children. Despite the availability of several therapies for NE, there is insufficient evidence to support the use of any one intervention and as such this randomised placebo-controlled phase II trial will be an important contribution to understanding potential new treatments for this condition. TRIAL REGISTRATION: Australian and New Zealand Clinical Trials Registration Number: 12618000288224. PROTOCOL: 23 February 2018, version 1.1.

Four New Flavonoids Isolated from the Aerial Parts of Cadaba rotundifolia Forssk. (Qadab).

Cadaba rotundifolia (Forssk.) (family: Capparaceae; common name: Qadab) is one of four species that grow in the Red Sea costal region in the Kingdom of Saudi Arabia. The roots and leaves of C. rotundifolia is traditionally used to treat tumors and abscesses in Sudan. A previous phytochemical study of the roots yielded a quaternary alkaloid, but no report on chemical constituents of the aerial parts of the C. rotundifolia growing in Saudi Arabia has been issued so far. Oxidative stress and advanced glycation end products (AGEs) are thought as causal factors in many degenerative diseases, such as Alzheimer's disease, diabetes, atherosclerosis and aging. In this study, a total of twenty compounds, including four previously undescribed acylated kaempferol glucosides, were isolated from the aerial parts of C. rotundifolia collected in Saudi Arabia. These new compounds were identified as kaempferol 3-O-[2-O-(trans-feruloyl)-3-O-ß-d-glucopyranosyl]-ß-d-glucopyranoside (1), kaempferol 3-O-ß-neohesperidoside-7-O-[2-O-(cis-p-coumaroyl)-3-O-ß-d-glucopyranosyl]-ß-d-glucopyranoside (2), kaempferol 3-O-[2,6-di-O-α-l-rhamnopyranosyl]-ß-d-glucopyranoside-7-O-[6-O-(trans-feruloyl)]-ß-d-glucopyranoside (3) and kaempferol 3-O-[2,6-di-O-α-l-rhamnopyranosyl]-ß-d-glucopyranoside-7-O-[6-O-(trans-p-coumaroyl)]-ß-d-glucopyranoside (4). Their structures were established based on UV-visible, 1D, 2D NMR, and HR-ESI-MS analyses. Of the assayed compounds, 17 and 18 showed potent radical scavenging activity with IC50 values of 14.5 and 11.7 µM, respectively, and inhibitory activity toward AGEs together with compound 7 with IC50 values 96.5, 34.9 and 85.5 µM, respectively.

A Bifunctional Molecule with Lectin and Protease Inhibitor Activities Isolated from Crataeva tapia Bark Significantly Affects Cocultures of Mesenchymal Stem Cells and Glioblastoma Cells.

Currently available drugs for treatment of glioblastoma, the most aggressive brain tumor, remain inefficient, thus a plethora of natural compounds have already been shown to have antimalignant effects. However, these have not been tested for their impact on tumor cells in their microenvironment-simulated cell models, e.g., mesenchymal stem cells in coculture with glioblastoma cell U87 (GB). Mesenchymal stem cells (MSC) chemotactically infiltrate the glioblastoma microenvironment. Our previous studies have shown that bone-marrow derived MSCs impair U87 growth and invasion via paracrine and cell-cell contact-mediated cross-talk. Here, we report on a plant-derived protein, obtained from Crataeva tapia tree Bark Lectin (CrataBL), having protease inhibitory/lectin activities, and demonstrate its effects on glioblastoma cells U87 alone and their cocultures with MSCs. CrataBL inhibited U87 cell invasion and adhesion. Using a simplified model of the stromal microenvironment, i.e., GB/MSC direct cocultures, we demonstrated that CrataBL, when added in increased concentrations, caused cell cycle arrest and decreased cocultured cells' viability and proliferation, but not invasion. The cocultured cells' phenotypes were affected by CrataBL via a variety of secreted immunomodulatory cytokines, i.e., G-CSF, GM-CSF, IL-6, IL-8, and VEGF. We hypothesize that CrataBL plays a role by boosting the modulatory effects of MSCs on these glioblastoma cell lines and thus the effects of this and other natural lectins and/or inhibitors would certainly be different in the tumor microenvironment compared to tumor cells alone. We have provided clear evidence that it makes much more sense testing these potential therapeutic adjuvants in cocultures, mimicking heterogeneous tumor-stroma interactions with cancer cells in vivo. As such, CrataBL is suggested as a new candidate to approach adjuvant treatment of this deadly tumor.

The leaves of Crataeva nurvala Buch-Ham. modulate locomotor and anxiety behaviors possibly through GABAergic system.

BACKGROUND: Crataeva nurvala Buch-Hum is an indigenous herb, extensively used in traditional medicines of the South Asian countries to treat inflammation, rheumatic fever, gastric irritation, and constipation. Despite this wide range of uses, very little information is known regarding its effects on the central nervous system (CNS). Therefore, this study evaluated the neuropharmacological properties of methanolic extract of Crataeva nurvala leaves (MECN) using a number of behavioral models in animals. This study also identified potentially active phytochemicals in MECN. METHODS: Following MECN administration (at 50, 100 and 200 mg/kg; b.w.) the animals (male Swiss albino mice) were employed in hole-cross test (HCT), open field test (OFT), and rota-rod test (RRT) to evaluate sedative properties, where anxiolytic activities were investigated using elevated plus maze (EPM), light dark box (LDB), and marble burying test (MBT). The involvement of GABAergic system was evaluated using thiopental sodium (TS)-induced sleeping time determination test. Moreover, colorimetric phytochemical tests as well as GC/MS-MS were also conducted to define the phytochemical constituents of MECN. RESULTS: MECN possesses sedative properties indicated through the dose-dependent inhibition of locomotor activities of the animals in HCT and OFT and motor coordination in RRT. MECN also exhibited prominent anxiolytic properties through decreased burying behavior in MBT, increased time spent and transitions in open arm of EPM, and increased time spent in light compartment of LDB. In addition, the treatments potentiated TS-mediated hypnosis indicating a possible participation of GABAergic system in the observed sedative and anxiolytic activities. Phytochemical screening of MECN revealed 48 different compounds in it. We reviewed and conceive that the sedative and anxiolytic effects could be due to the presence of neuroactive compounds such as phytol, D-allose, and α-Tocopherol in MECN. CONCLUSION: The present study showed that MECN possesses sedative and anxiolytic potential which could be beneficial in treatment of anxiety and insomnia associated with different psychological disorders.

Urox containing concentrated extracts of Crataeva nurvala stem bark, Equisetum arvense stem and Lindera aggregata root, in the treatment of symptoms of overactive bladder and urinary incontinence: a phase 2, randomised, double-blind placebo controlled trial.

BACKGROUND: Storage lower urinary tract symptoms (LUTS) including overactive bladder (OAB) and urinary incontinence (UI) affect millions of people worldwide, significantly impacting quality of life. Plant based medicines have been documented both empirically and in emerging scientific research to have varying benefits in reducing bladder symptoms. We assessed the efficacy of Urox®, a proprietary combination of phytomedicine extracts including, Cratevox™ (Crataeva nurvala) stem bark, Equisetem arvense stem and Lindera aggregata root, in reducing symptoms of OAB and UI. METHODS: Efficacy of the herbal combination on a variety of bladder symptoms compared to an identical placebo, were documented in a randomised, double-blind, placebo controlled trial conducted at two primary care centres. Data were collected at baseline, 2, 4 and 8 weeks, with the primary outcome being self-reported urinary frequency. Statistical analysis included mixed effects ordered logistic regression with post hoc Holm's test to account for repeated measures, and included an intention-to-treat analysis. RESULTS: One hundred and fifty participants (59% female, aged; mean ± SD; 63.5 ± 13.1 years) took part in the study. At week 8, urinary day frequency was significantly lower (OR 0.01; 95%CI 0.01 to 0.02; p < 0.001) in response to treatment (mean ± SD; 7.69 ± 2.15/day) compared to placebo (10.95 ± 2.47/day). Similarly, episodes of nocturia were significantly fewer (OR 0.03; 95%CI 0.02 to 0.05) after 8 weeks of treatment (2.16 ± 1.49/night) versus placebo (3.14 ± 1.36/night). Symptoms of urgency (OR 0.02; 95%CI 0.01 to 0.03), and total incontinence (OR 0.03; 95% CI 0.01 to 0.06) were also lower (all p < 0.01) in the treatment group. Significant improvements in quality of life were reported after treatment in comparison to placebo. No significant side effects were observed resulting in withdrawal from treatment. CONCLUSIONS: The outcome of this study demonstrated both statistical significance and clinical relevance in reducing symptoms of OAB, urinary frequency and/or urgency and incontinence. The demonstrated viability of the herbal combination to serve as an effective treatment, with minimal side-effects, warrants further longer term research and consideration by clinicians. TRIAL REGISTRATION: NCT02396160 (registered on 17 March 2015 - before any statistical analyses commenced).

Post-transcriptional regulation of photosynthetic genes is a key driver of C4 leaf ontogeny.

C4 photosynthesis allows highly efficient carbon fixation that originates from tightly regulated anatomical and biochemical modifications of leaf architecture. Recent studies showed that leaf transcriptome modifications during leaf ontogeny of closely related C3 (Tarenaya hassleriana) and C4 (Gynandropsis gynandra) species within the Cleomaceae family existed but they did not identify any dedicated transcriptional networks or factors specifically driving C4 leaf ontogeny. RNAseq analysis provides a steady-state quantification of whole-cell mRNAs but does not allow any discrimination between transcriptional and post-transcriptional processes that may occur simultaneously during leaf ontogeny. Here we use exon-intron split analysis (EISA) to determine the extent to which transcriptional and post-transcriptional processes are involved in the regulation of gene expression between young and expanded leaves in both species. C4-specific changes in post-transcriptional regulation were observed for genes involved in the Calvin-Benson cycle and some photosystem components but not for C4 core-cycle genes. Overall, this study provides an unbiased genome-wide insight into the post-transcriptional mechanisms that regulate gene expression through the control of mRNA levels and could be central to the onset of C4 photosynthesis. This mechanism is cytosolic which implies cell-specific modifications of mRNA stability. Understanding this mechanism may be crucial when aiming to transform C3 crops into C4 crops.

Differential distribution of amino acids in plants.

Plants are a rich source of amino acids and their individual abundance in plants is of great significance especially in terms of food. Therefore, it is of utmost necessity to create a database of the relative amino acid contents in plants as reported in literature. Since in most of the cases complete analysis of profiles of amino acids in plants was not reported, the units used and the methods applied and the plant parts used were different, amino acid contents were converted into relative units with respect to lysine for statistical analysis. The most abundant amino acids in plants are glutamic acid and aspartic acid. Pearson's correlation analysis among different amino acids showed that there were no negative correlations between the amino acids. Cluster analysis (CA) applied to relative amino acid contents of different families. Alismataceae, Cyperaceae, Capparaceae and Cactaceae families had close proximity with each other on the basis of their relative amino acid contents. First three components of principal component analysis (PCA) explained 79.5% of the total variance. Factor analysis (FA) explained four main underlying factors for amino acid analysis. Factor-1 accounted for 29.4% of the total variance and had maximum loadings on glycine, isoleucine, leucine, threonine and valine. Factor-2 explained 25.8% of the total variance and had maximum loadings on alanine, aspartic acid, serine and tyrosine. 14.2% of the total variance was explained by factor-3 and had maximum loadings on arginine and histidine. Factor-4 accounted 8.3% of the total variance and had maximum loading on the proline amino acid. The relative content of different amino acids presented in this paper is alanine (1.4), arginine (1.8), asparagine (0.7), aspartic acid (2.4), cysteine (0.5), glutamic acid (2.8), glutamine (0.6), glycine (1.0), histidine (0.5), isoleucine (0.9), leucine (1.7), lysine (1.0), methionine (0.4), phenylalanine (0.9), proline (1.1), serine (1.0), threonine (1.0), tryptophan (0.3), tyrosine (0.7) and valine (1.2).

Crataeva nurvala nanoparticles inhibit virulence factors and biofilm formation in clinical isolates of Pseudomonas aeruginosa.

Green synthesized nanoparticles have gained great attention due to their non-toxic and non-hazardous nature. In the present study, bark extract of the medicinal plant in Ayurveda Crataeva nurvala (Buch-Ham) (CN) was chosen for the biosynthesis of silver nanoparticles (AgNPs). These NPs were characterized by Ultra violet visible spectroscopy, Fourier Transform Infra Red, Atomic Force Microscopy, and Transmission Electron Microscopy (TEM). The average particle size of green synthesized CN-AgNPs was 15.2 ± 1.01 nm. Gas chromatography- mass spectrometry analysis of methanolic bark extract involved in the formation of CN-AgNPs revealed lupeol as a major active component. In this study, CN-AgNPs (15 µg ml-1 ) efficiently suppressed the production of quorum sensing mediated virulence factors viz. pyocyanin, protease, hemolysin, and biofilm formation in Pseudomonas aeruginosa. The pyocyanin production was strongly inhibited (74.64%) followed by proteolysis (47.3%) and hemolysin production (47.7%). However, the biofilm forming ability was maximally reduced up to 79.70%. Moreover, the Confocal Laser Scanning Microscopic Analysis showed that CN-AgNPs inhibit colonization of P. aeruginosa on to the surface. Furthermore, TEM analysis revealed internalization of CN-AgNPs inside the bacterial cell. It is concluded that green synthesized AgNPs have great potential to inhibit virulence factors and biofilm forming ability of drug-resistant clinical isolates of P. aeruginosa.

Immunomodulatory activity of Buchholzia coriacea seed methanol extract on Trypanosoma brucei brucei infected mice.

CONTEXT: The seeds of Buchholzia coriacea Engler (Capparaceae) are used in Eastern Nigeria to treat feverish conditions, and to treat malaria and sleeping sickness that cause fever. OBJECTIVE: The current study assesses the immunomodulatory activity of Buchholzia coriacea seed extract on Trypanosoma brucei brucei infected mice. MATERIALS AND METHODS: Delayed hypersensitivity reaction, humoral antibody response and in-vivo leucocyte mobilization tests were assessed in three different experiments to determine the effect of the extract on immune response. Seventy-five (75) mice (25 mice per experiment) were used for the study and were each infected with 1.00 × 106 trypanosomes intra-peritoneally. Groups A, B and C were given 250, 500 and 1000 mg/kg of the extract, respectively, group D received 7.5 mg/kg body weight of levamisole and group E was the control. Sheep RBCs were used as antigen. RESULTS: The acute toxicity tests did not cause clinical signs or death within 24 h post treatment at all the doses tested. The extract inhibited delayed hypersensitivity reaction by 20.9 and 20.8% at 250 and 500 mg/kg, respectively, while at 1000 mg/kg, the paw size increased (-101.9%) when compared with the control. The extract elevated the antibody titre from 1.60 ± 0.40 for control to 8.00 ± 3.58 for 500 mg/kg group. The extract increased in total leucocytes counts. DISCUSSION AND CONCLUSION: The extract has a very wide safety margin and was able to improve immune response. The results of the present study showed that Buchholzia coriacea seed methanol extract possesses immunostimulatory activity on trypanosome-infected mice.

Molecular and cytogenetic assessment of Dipterygium glaucum genotoxicity.

The aim of the present study is to assess the genotoxicity of Dipterygium glaucum grows widely in Saudi Arabia desert to produce safety herbal products. This work is considered the first and pioneer report so far due to the lack and poor evaluated reports of the plant species for their mutagensity, genotoxicity and cytogenetics effects. Cytogenetic effects of D. glaucum on mitotic in roots of Vicia faba showed reduction in mitotic activity using three extracts; water, ethanol and ethyl acetate. Chromosomal abnormalities were recorded that included stickiness of chromosomes, chromatin bridge, fragments, lagging chromosome and micronuclei. Protein bands and RAPD analyses of V. faba treated with three D. glaucum extracts revealed some newly induced proteins and DNA fragments and other disappeared. Chemical constitution of the plant species should be identified with their biological activities against human and animal cells like HeLa cancer cell line. We are recommending using additional genotoxicity tests and other toxicity tests on animal culture with different concentrations and also utilizing several drought and heat tolerant genes of the plant species in gene cloning to develop and improve other economical crop plants instead of using the species as oral herbal remedy.

CrataBL, a lectin and Factor Xa inhibitor, plays a role in blood coagulation and impairs thrombus formation.

Arterial thrombosis is an important complication of diabetes and cancer, being an important target for therapeutic intervention. Crataeva tapia bark lectin (CrataBL) has been previously shown to have hypoglycemiant effect and also to induce cancer cell apoptosis. It also showed inhibitory activity against Factor Xa (Kiapp=8.6 µm). In the present study, we evaluated the anti-thrombotic properties of CrataBL in arterial thrombosis model. CrataBL prolongs the activated partial thromboplastin time on human and mouse plasma, and it impairs the heparin-induced potentiation of antithrombin III and heparin-induced platelet activation in the presence of low-dose ADP. It is likely that the dense track of positive charge on CrataBL surface competes with the heparin ability to bind to antithrombin III and to stimulate platelets. In the photochemically induced thrombosis model in mice, in the groups treated with 1.25, 5.0, or 10 mg/kg CrataBL, prior to the thrombus induction, the time of total artery occlusion was prolonged by 33.38%, 65%, and 66.11%, respectively, relative to the time of the control group. In contrast to heparin, the bleeding time in CrataBL-treated mice was no longer than in the control. In conclusion, CrataBL was effective in blocking coagulation and arterial thrombus formation, without increasing bleeding time.

Evaluation of antinociceptive effect of methanolic extract of leaves of Crataeva nurvala Buch.-Ham.

BACKGROUND: Crataeva nurvala Buch.-Ham. (Family: Capparidaceae) is widely used as anti-inflammatory, contraceptive, laxative, lithotropic, febrifuge and as tonic in traditional medicine. This study evaluated the antinociceptive effect of the methanolic extract of the leaves of Crataeva nurvala (MECN). METHODS: The antinociceptive activity was investigated using heat-induced (hot-plate and tail-immersion test) and chemical-induced (acetic acid, formalin and glutamic acid) nociception models in mice at different doses (50, 100, and 200 mg/kg, p.o.) of MECN. Morphine sulphate (5 mg/kg, i.p.) and diclofenac sodium (10 mg/kg, i. p.) were used as reference analgesic drugs. RESULTS: MECN produced significant dose-dependent antinociception when assessed using hot plate test, tail immersion test and acetic acid-induced abdominal writhing test (65.55%). Likewise, MECN at similar doses produced significant dose-dependent inhibition in both neurogenic (50.82%) and inflammatory pain (73.53%) induced by intraplantar injection of formalin (2.5% formalin, 20 µl/paw). Besides, MECN also significantly inhibited the glutamate-induced (10 µM/paw) pain in mice (74.68%). It was demonstrated that pretreatment with naloxone (2 mg/kg, i.p.) significantly reversed antinociception produced by MECN in hot plate and tail immersion test suggesting the involvement of opioid receptor. In addition, administration of glibenclamide (10 mg/kg, i.p.), an ATP-sensitive K+ channel antagonist could not reverse antinociceptive activity induced by MECN. CONCLUSION: The results suggest that MECN possesses antinociceptive activity involving inhibition of opioid system as well as the glutamatergic system supporting its traditional uses.

Report: Anxiolytic, sedative and toxicological effect of hydromethanolic stem bark extract of Maerua angolensis DC. in Wister rats.

Maerua angolensis DC is traditionally used for the treatment of epilepsy and insomnia. The present study was designed to investigate the anxiolytic, sedative and toxicological effect of hydromethanolic stem bark extract of M. angolensis using animal model. Sub-chronic doses of the plant extract on liver and kidney function test were investigated. Elevated plus maze (EPM) and diazepam-induced sleeping time test was used in this investigation. The possible involvement of M. angolensis with GABAA receptor was also investigated using flumazenil. The results of acute toxicity studies showed LD50 to be greater than 5000mg/kg body weight. The test extract (40 and 80mg/kg) significantly (p<0.05) increased the number of open arm entries and time spent in the open arm entries. However, flumazenil with 80mg/kg plant extract showed no significant (p >0.01) difference in the number of entries into open arm when compared to control. The stem bark extract of M. angolensis significantly (p<0.01) increased the duration of sleep induced by diazepam in a dose-dependent manner. However, flumazenil with 80mg/kg extract showed no significant (p>0.01) sedative effect when compared to normal control. In conclusion, the result of our present findings revealed that M. angolensis may apparently be safe and non toxic at therapeutic dose. However, the plant may possess anxiolytic and sedative properties, which exert their effect on GABAA receptors.

Structural studies of the interaction of Crataeva tapia bark protein with heparin and other glycosaminoglycans.

CrataBL, a protein isolated from Crataeva tapia bark, which is both a serine protease inhibitor and a lectin, has been previously shown to exhibit a number of interesting biological properties, including anti-inflammatory, analgesic, antitumor, and insecticidal activities. Using a glycan array, we have now shown that only sulfated carbohydrates are effectively bound by CrataBL. Because this protein was recently shown to delay clot formation by impairing the intrinsic pathway of the coagulation cascade, we considered that its natural ligand might be heparin. Heparin is a glycosaminoglycan (GAG) that interacts with a number of proteins, including thrombin and antithrombin III, which have a critical, essential pharmacological role in regulating blood coagulation. We have thus employed surface plasmon resonance to improve our understanding of the binding interaction between the heparin polysaccharide and CrataBL. Kinetic analysis shows that CrataBL displays strong heparin binding affinity (KD = 49 nM). Competition studies using different size heparin-derived oligosaccharides showed that the binding of CrataBL to heparin is chain length-dependent. Full chain heparin with 40 saccharides or large oligosaccharides, having 16-18 saccharide residues, show strong binding affinity for CrataBL. Heparin-derived disaccharides through tetradecasaccharides show considerably lower binding affinity. Other highly sulfated GAGs, including chondroitin sulfate E and dermatan 4,6-disulfate, showed CrataBL binding affinity comparable to that of heparin. Less highly sulfated GAGs, heparan sulfate, chondroitin sulfate A and C, and dermatan sulfate displayed modest binding affinity as did chondroitin sulfate D. Studies using chemically modified heparin show that N-sulfo and 6-O-sulfo groups on heparin are essential for CrataBL-heparin interaction.

Neuro-pharmacological potentials of Buchholzia coriacea (Engl.) seeds in laboratory rodents.

BACKGROUND: Buchholzia coriacea, taken by elderly, has phytochemicals that have neuro-active metabolites, and the folklore documented its use in neuro-behavioural despairs. OBJECTIVE: This study was conducted to investigate the neuro-pharmacological potentials of Buchholzia coriacea (MEBC) seed extract in the laboratory rodents. METHODOLOGY: Methanol extract of the seeds on B. coriacea (MEBC) was evaluated for its antidepressant (Forced Swimming Test and Tail Suspension Test), anxiolytic (Light-Dark Test, Hole Board Test and Elevated Plus Maze), antinociceptive (Hot-Plate and Tail Flick test) and motor coordination (Rota Rod) functions in mice. RESULTS: Our findings showed antidepressant activity (P < 0.05) of MEBC that is dose-dependent. Secondly, MEBC showed significant anxiolytic property that is comparable with the standard drug (diazepam). Furthermore, MEBC significantly prolonged the latency on hot-plate and tail flick responses when compared with the control (P < 0.05). Finally, MEBC significantly prolonged mice endurance time (P < 0.05) on a revolving Rota rod. The results suggest antidepressant, anxiolytic and analgesic potentials of MEBC. CONCLUSION: Buchholzia coriacea may have a stabilizing effect on the motor activity and MEBC probably contains secondary metabolites with some therapeutic effects on neuro-physiological disorders like depression, anxiety and pain.