Antimycobacterial Metabolites from Plectranthus: Royleanone Derivatives against Mycobacterium tuberculosis Strains

The antimycobacterial activities of eight diterpenes, 1 – 8 , isolated previously from Plectranthus and eleven esters, 9 – 19 , of 7α‐acetoxy‐6β,12‐dihydroxyabieta‐8,12‐diene‐11,14‐dione ( 5 ) were evaluated against the MTB strains H₃₇Rv and MDR. Only diterpenoids with a quinone framework revealed anti‐MTB activity. Abietane 5 and its 6,12‐dibenzoyl, 12‐methoxybenzoyl, 12‐chlorobenzoyl, and 12‐nitrobenzoyl esters, 9, 11, 12 , and 13 , respectively, showed potent activities against the MDR strain with MIC values between 3.12 and 0.39 μg/ml. Cytotoxic activities towards 3T3 and Vero cells were also evaluated. Compound 11 , with the best selectivity index, may be a suitable lead for further chemical modifications. The complete structural elucidation of the new esters, 9 – 14, 16, 18 , and 19 , as well as the NMR data of known derivatives 15 and 17 are reported.     

Design,synthesis, and bioevaluation of a novel class of (E)-4-oxo-crotonamide derivatives as potent antituberculosis agents

A series of novel (E)-4-oxo-2-crotonamide derivatives were designed and synthesized to find potent antituberculosis agents. All the target compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis H₃₇Rv(MTB). Results reveal that 4-phenyl moiety at part A and short methyl group at part C were found to be favorable. Most of the derivatives displayed promising activity against MTB with MIC ranging from 0.125 to 4?µg/mL. Especially, compound IIIa16 was found to have the best activity with MIC of 0.125?μg/mL against MTB and with MIC in the range of 0.05–0.48?µg/mL against drug-resistant clinical MTB isolates.     

Design,synthesis and biological evaluation of 4-benzoyl-1-dichlorobenzoylthiosemicarbazides as potent Gram-positive antibacterial agents

Twelve 4-benzoyl-1-dichlorobenzoylthiosemicarbazides have been tested as potential antibacterials. All the compounds had MICs between 0.49 and 15.63?µg/ml toward Micrococcus luteus, Bacillus cereus, Bacillus subtilis and Staphylococcus epidermidis indicating, in most cases, equipotent or even more effective action than cefuroxime. In order to clarify if the observed antibacterial effects are universal, further research were undertaken to test inhibitory potency of two most potent compounds 3 and 11 on clinical isolates of Staphylococcus aureus. Compound 11 inhibited the growth of methicillin-sensitive S. aureus (MSSA) at MICs of 1.95–7.81?µg/ml, methicillin-resistant S. aureus (MRSA) at MICs of 0.49–1.95?µg/ml and MDR–MRSA at MIC of 0.98 and 3.90?µg/ml, respectively. Finally, inhibitory efficacy of 3 and 11 on planktonic cells and biofilms formation in clinical isolates of S. aureus and Haemophilus parainfluenzae was tested. The majority of cells in biofilm populations of MSSA and MRSA were eradicated at low level of 3, with MBICs in the range of 7.82–15.63?µg/ml.     

Chalcone analogues: Synthesis,activity against Meloidogyne incognita,and in silico interaction with cytochrome P450

To contribute the development of new products to control plant‐parasitic nematodes, 12 chalcone analogues were synthesized and screened for activity against Meloidogyne incognita. Three caused mortality greater than negative controls in second‐stage juvenile M. incognita, with values varying from 19.9% to 100%. The most active chalcone analogue was (1E,4E)‐1,5‐di(4‐nitrophenyl)‐2‐butylpenta‐1,4‐dien‐3‐one (compound 6 ), which had an LC₅₀ value of 41 µg/ml. Under the same conditions, the commercial nematicide Carbofuran^® (2,2‐dimethyl‐2,3‐dihydro‐1‐benzofuran‐7‐yl methylcarbamate) presented an LC₅₀ equal to 101 µg/ml. When this chalcone analogue was applied to tomato plants infested with M. incognita, reductions in the numbers of galls and eggs of 51% and 68% were observed, respectively. According to in silico studies, the enzyme target of compound 6 in M. incongita is cytochrome P450, which is important for the oxidation of several substances in the nematode. Therefore, compound 6 is potentially useful for the development of new products to control M. incongita.     

Effective antimicrobial activity of Cbf‐K16 and Cbf‐A7A13 against NDM‐1‐carrying Escherichia coli by DNA binding after penetrating the cytoplasmic membrane in vitro

New Delhi metallo‐beta‐lactamase‐1(NDM‐1)‐carrying isolates, which are resistant to most clinical used antibiotics except for tigecycline and colistin, have been found worldwide. Cathelicidin‐BF (BF‐30) is found in the venom of the snake Bungarus fasciatus and exhibits broad antimicrobial activity. Cbf‐K₁₆ and Cbf‐A₇A₁₃ were obtained by mutating Lys¹⁶, Ala⁷, and Ala¹³ of BF‐30, respectively. To investigate their antimicrobial activities against NDM‐1 carrying bacteria, recombinant Escherichia coli BL21 (DE3)‐NDM‐1 with high NDM‐1 activity was constructed by inserting the Klebsiella pneumoniae NDM‐1 gene (GenBank accession no. HQ328085) into a pET28a vector and transforming it into E. coli BL21 (DE3). The peptides showed effective antimicrobial activities against NDM‐1‐carrying E. coli, and the minimum inhibitory concentrations of Cbf‐K₁₆ and Cbf‐A₇A₁₃ were only 4 and 8 µg/ml, whereas those of minimum bactericidal concentrations were 8 and 16 µg/ml, respectively. A time course experiment showed that colony forming unit counts rapidly decreased, and bacteria were thoroughly eliminated within 3 and 6 h by the Cbf‐K₁₆ and Cbf‐A₇A₁₃ treatments, respectively. The peptides penetrated the bacterial cell membrane and enabled β‐galactosidase leakage, and caused the cytoplasmic membrane to become permeable, and finally bound to the DNA. The genomic DNA of E. coli was completely unable to migrate on an agarose gel after Cbf‐K₁₆ treatment (8 µg/ml). These data demonstrated that Cbf‐K₁₆ and Cbf‐A₇A₁₃ possess effective antimicrobial activity against drug‐resistant strains, including NDM‐1 carrying E. coli BL21 (DE3)‐NDM‐1, by binding to DNA after penetrating the cytoplasmic membrane in vitro, which may have potential therapeutic value for the treatment of NDM‐1‐carrying bacterial infections. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Characterization of a Bacillus thuringiensis strain with a broad spectrum of activity against lepidopteran insects

The insect pathogen Bacillus thuringiensis is suitable for use in biological control, and certain strains have been developed as commercial bioinsecticides. The molecular and biological characterization of a Bacillus thuringiensis subsp. aizawai strain, named HU4‐2, revealed its potential as a bioinsecticide. The strain was found to contain eight different cry genes: cry1Ab, cry1Ad, cry1C, cry1D, cry1F, cry2, cry9Ea1, and a novel cry1I‐type gene. Purified parasporal crystals from strain HU4‐2 comprised three major proteins of 130–145 kDa, which were tested for their insecticidal potency to four species of Lepidoptera (Helicoverpa armigera, Spodoptera exigua, S. littoralis, and S. frugiperda) and three species of mosquito (Culex pipiens pipiens, Aedes aegypti, and Anopheles stephensi). The crystal proteins were highly toxic against all the species of Lepidoptera tested, moderately toxic against two of the mosquito species (C. pipiens and Ae. aegypti), but no toxicity was observed against a third species of mosquito (An. stephensi) at the concentrations used in our study. The LC₅₀ values of the HU4‐2 Bt strain against H. armigera larvae (5.11 µg/ml) was similar to that of HD‐1 Bt strain (2.35 µg/ml), the active ingredient of the commercial product Dipel®. Additionally, the LC₅₀ values of the HU4‐2 Bt strain against S. littoralis, S. frugiperda, and S. exigua (2.64, 2.22, and 3.38 µg/ml, respectively) were also similar to that of the Bt strain isolated from the commercial product Xentari® for the same three species of Spodoptera (1.94, 1.34, and 2.19 µg/ml, respectively). Since Xentari® is significantly more toxic to Spodoptera spp. than Dipel® and, reciprocally, Dipel® is significantly more toxic against H. armigera than Xentari®, we discuss the potential of the HU4‐2 strain to control all these important lepidopteran pests.     

Design,synthesis and in vivo/in vitro screening of novel chlorokojic acid derivatives

A series of novel Mannich bases of chlorokojic acid (2-chloromethyl-5-hydroxy-4H-pyran-4-one) were synthesized and their biological activities were investigated. Anticonvulsant activity results according to phase-I tests of Antiepileptic Drug Development (ADD) Program revealed that compound 13 was the most effective one at 4?h against subcutaneous pentylenetetrazole (scPTZ)-induced seizure test. Antimicrobial activities were evaluated in vitro against bacteria and fungi by using broth microdilution method. The antitubercular activities against Mycobacterium tuberculosis and M. avium were discussed with Resazurin microplate assay (REMA). The antimicrobial activity results indicated that compounds 1 and 12 (MIC: 8–16 µg/mL) showed higher activity against Gram negative bacteria while compound 12 had MIC: 4–16 µg/mL against Gram positive bacteria. Compound 1 was the most active one with MIC values of 8–32 µg/mL against fungi. Mannich bases also exhibit significant antitubercular activity in a MIC range of 4 to 32 µg/mL, especially compound 18 against M. avium.     

Analogues of the frog skin peptide alyteserin‐2a with enhanced antimicrobial activities against Gram‐negative bacteria

The emergence of strains of multidrug‐resistant Gram‐negative bacteria mandates a search for new types of antimicrobial agents. Alyteserin‐2a (ILGKLLSTAAGLLSNL.NH₂) is a cationic, α‐helical peptide, first isolated from skin secretions of the midwife toad, Alytes obstetricans, which displays relatively weak antimicrobial and haemolytic activities. Increasing the cationicity of alyteserin‐2a while maintaining amphipathicity by the substitution Gly¹¹→ Lys enhanced the potency against both Gram‐negative and Gram‐positive bacteria by between fourfold and 16‐fold but concomitantly increased cytotoxic activity against human erythrocytes by sixfold (mean concentration of peptide producing 50% cell death; LC₅₀ = 24 µm ). Antimicrobial potency was increased further by the additional substitution Ser⁷→Lys, but the resulting analogue remained cytotoxic to erythrocytes (LC₅₀ = 38 µm ). However, the peptide containing d ‐lysine at positions 7 and 11 showed high potency against a range of Gram‐negative bacteria, including multidrug‐resistant strains of Acinetobacter baumannii and Stenotrophomonas maltophilia (minimum inhibitory concentration = 8 µm ) but appreciably lower haemolytic activity (LC₅₀ = 185 µm ) and cytotoxicity against A549 human alveolar basal epithelial cells (LC₅₀ = 65 µm ). The analogue shows potential for treatment of nosocomial pulmonary infections caused by bacteria that have developed resistance to commonly used antibiotics. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.     

Antifungal activity of two alkaloids of Zephyranthes citrina and their field efficacy against powdery mildew (Erysiphe cichoracearum) of balsam

The anti-fungal activity of two alkaloids isolated from bulbs of Zephyranthes citrina was observed against 10 fungi, viz, Aternaria solani, A. triticina, Curvularia lunata, C. maculuns, Cercospora malvacearum, Erysiphe sp., Fusarium udum, Helminthosporium pisi, H. speciferum and Ustilago cynodontis. Different concentrations (200, 400, 600, 800, 1000 µg/ml of alkaloids (A and B) were used. Spore germination was inhibited at 600, 800, 1000 µg/ml. B alkaloid was used against Erysiphe cichoracearum causing powdery mildew in balsam (Impatiens balsamina) in the field as pre- and post-inoculation treatments at 1000, 1500, 2000 µg/ml doses. The extract was effective in both pre- and post-inoculation treatments. Foliar application of this alkaloid resulted in inducing synthesis of phenolic acid in the leaves of balsam. Maximum phenolics were detected in the leaves treated with 1500 µg/ml in both pre- and post-inoculation treatments. The increase in the production of phenolics in treated leaves of balsam can be correlated with the induction of resistance in treated plants against powdery mildew. The significant efficacy of the alkaloid under field conditions opens the possibility of its use by farmers for also controlling other diseases.     

(7-Chloroquinolin-4-yl)arylhydrazones: Candida albicans enzymatic repression and cytotoxicity evaluation,Part 2

Objective: This work describes the anti-enzymatic activity of (7-chloroquinolin-4-yl)arylhydrazones against Candida albicans and examines their cytotoxicity.

Material and methods: Ten C. albicans strains [nine isolates and one azole-resistant standard strain (ATCC 62342)] were used to assess the anti-enzymatic activity. Fifteen compounds at sub-antifungal concentrations ranging from 12.5 to 100?µg/ml were assessed after a 30-min exposure. The strains were seeded onto petri dishes with selective agar media for aspartyl proteases (Saps) and phospholipases (PLs). Enzymatic inhibition was measured by the reduction of the precipitation zone (Pz) against untreated strains (positive control). A colorimetric MTT assay was used with 3T3/NIH mouse fibroblasts to evaluate cytotoxicity. Cells were exposed to 15 compounds in concentrations from 6.25 to 100?µg/ml for 24 and 48?h.

Results: Four hydrazones showed enzymatic repression values over 40% to Pl and three over 20% to Saps. The cell viability was over 50% at hydrazone concentrations of 25–100?µg/ml.

Conclusion: These results revealed that select (7-chloroquinolin-4-yl)arylhydrazones may be potential antifungal agents for the control of C. albicans infections.     

Effects of Manduca sexta allatostatin and an analogue on the peach-potato aphid Myzus persicae (hemiptera: aphididae) and degradation by enzymes in the aphid gut

The oral toxicity of the C‐type allatostatin, Manduca sexta allatostatin (Manse‐AS) and the analogue δR³δR⁵Manse‐AS, where R residues were replaced by their D‐isomers, were tested against the peach‐potato aphid Myzus persicae by incorporation into an artificial diet. Both peptides had significant dose‐dependent effects on mortality, growth, and fecundity compared with control insects. The analogue, δR³δR⁵Manse‐AS, had an estimated LC₅₀ of 0.31 µg/µl diet and was more potent than Manse‐AS (estimated LC₅₀ of 0.58 µg/µl diet). At a dose of 0.35 µg δR³δR⁵Manse‐AS/µl diet, 76% of the aphids were dead after 6 days and all were dead after 10 days. In comparison, three times the dose of Manse‐AS was required to achieve 74% mortality after 8 days and 98% mortality after 16 days. The degradation of both peptides by extracts prepared from the gut of M. persicae was investigated. The estimated half‐life of Manse‐AS, when incubated with the gut extract from M. persicae, was 31 min. Degradation was due to a cathepsin L‐like cysteine protease, carboxypeptidase‐like activity, endoprotease activity with glutamine specificity, pyroglutamate aminopeptidase activity, and possibly trypsin‐like proteases. The half‐life of the δR³δR⁵ Manse‐AS analogue was enhanced (73 min) with the D‐isomers of R appearing to prevent cleavage around the R residues by cathepsin L‐like cysteine proteases or from trypsin‐like proteases. The greater stability of the analogue may explain its increased potency in M. persicae. This work demonstrates the potential use of Manse‐AS and analogues, with greater resistance to enzymatic attack, in aphid control strategies. © 2010 Wiley Periodicals, Inc.     

Design,synthesis, antimicrobial evaluation and molecular docking studies of some new 2,3-dihydrothiazoles and 4-thiazolidinones containing sulfisoxazole

Microbial resistance to the available drugs poses a serious threat in modern medicine. We report the design, synthesis and in vitro antimicrobial evaluation of new functionalized 2,3-dihydrothiazoles and 4-thiazolidinones tagged with sulfisoxazole moiety. Compound 8d was most active against Bacillis subtilis (MIC, 0.007?µg/mL). Moreover, compounds 7c–d and 8c displayed significant activities against B. subtilis and Streptococcus pneumoniae (MIC, 0.03–0.06?µg/mL and 0.06–0.12?µg/mL versus ampicillin 0.24?µg/mL and 0.12?µg/mL; respectively). Compounds 7a and 7c–d were highly potent against Escherichia coli (MIC, 0.49–0.98?µg/mL versus gentamycin 1.95?µg/mL). On the other hand, compounds 7e and 9c were fourfolds more active than amphotericin B against Syncephalastrum racemosum. Molecular docking studies showed that the synthesized compounds could act as inhibitors for the dihydropteroate synthase enzyme (DHPS). This study is a platform for the future design of more potent antimicrobial agents.     

Synthesis of short cationic antimicrobial peptidomimetics containing arginine analogues

Worldwide efforts are underway to develop new antimicrobial agents against bacterial resistance. To identify new compounds with a good antimicrobial profile, we designed and synthesized two series of small cationic antimicrobial peptidomimetics (1–8) containing unusual arginine mimetics (to introduce cationic charges) and several aromatic amino acids (bulky moieties to improve lipophilicity). Both series were screened for in vitro antibacterial activity against a representative panel of Gram‐positive (Staphylococcus aureus and Staphylococcus epidermidis) and Gram‐negative (Escherichia coli and Klebsiella pneumoniae) bacterial strains, and Candida albicans. The biological screening showed that peptidomimetics containing tryptophan residues are endowed with the best antimicrobial activity against S. aureus and S. epidermidis in respect to the other synthesized derivatives (MIC values range 7.5–50 µg/ml). Moreover, small antimicrobial peptidomimetics derivatives 2 and 5 showed an appreciable activity against the tested Gram‐negative bacteria and C. albicans. The most active compounds (1–2 and 5–6) have been tested against Gram‐positive established biofilm, too. Results showed that the biofilm inhibitory concentration values of these compounds were never up to 200 µg/ml. The replacement of tryptophan with phenylalanine or tyrosine resulted in considerable loss of the antibacterial action (compounds 3–4 and 7–8) against both Gram‐positive and Gram‐negative bacterial strains. Furthermore, by evaluating hemolytic activity, the synthesized compounds did not reveal cytotoxic activities, except for compound 5. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.     

Piperine,an active ingredient of white pepper,suppresses the growth of multidrug-resistant toxigenic Vibrio cholerae and other pathogenic bacteria

Emergence and rapid spread of multidrug-resistant (MDR) bacteria including Vibrio cholerae are a global public health issue. Much attention has been paid to natural compounds, such as spices and herbs to find novel antimicrobial compounds as they are considered to be cheaper alternatives to develop as a drug. Here, we show that methanol extract of white pepper could inhibit the growth of V. cholerae O1 El Tor variant, responsible for the recent outbreaks/epidemics. Furthermore, we demonstrate for the first time that piperine, the major component of white pepper, showed a dose-dependent bactericidal effect on V. cholerae growth irrespective of their biotypes and serogroups in the presence of 200 and 300 µg ml⁻¹ of piperine, respectively. Piperine also inhibited the growth of MDR strains of Pseudomonas aeruginosa, Escherichia coli isolated from poultry and enterohemorrhagic/enteroaggregative E. coli O104 in the presence of 200 µg ml⁻¹. Interestingly, we did not observe any significant inhibitory effect of piperine on E. coli strains isolated from healthy person even up to 200 µg ml⁻¹. Our data suggest that piperine could be a novel antimicrobial agent in therapeutic and preventive applications against infections caused by pathogenic bacteria including MDR strains.     

Diterpenoids from Acanthopanax brachypus Harms

Four diterpenoids, including two new ones, namely noracanthopanin A ( 1 ) and acanthopanolide B ( 2 ), and two known ones, ent‐pimara‐9(11),15‐dien‐19‐oic acid ( 3 ) and ent‐kaur‐16‐en‐19‐oic acid ( 4 ), were isolated from the stem bark of Acanthopanax brachypus Harms . Their structures were elucidated by physicochemical and spectroscopic analyses. Compound 1 and 2 exhibited moderate cytotoxicities against HO‐8910 cells with IC₅₀ values of 65.21±0.23 and 31.43±1.01 μg/ml, respectively, and 2 showed selective weak cytotoxicity against SMMC‐7721 cells (IC₅₀ 89.67±2.54 μg/ml).     

In vitro anti-TB properties,in silico target validation,molecular docking and dynamics studies of substituted 1,2,4-oxadiazole analogues against Mycobacterium tuberculosis

The alarming increase in multi- and extensively drug-resistant (MDR and XDR) strains of Mycobacterium tuberculosis (MTB) has triggered the scientific community to search for novel, effective, and safer therapeutics. To this end, a series of 3,5-disubstituted-1,2,4-oxadiazole derivatives (3a–3i) were tested against H37Rv, MDR and XDR strains of MTB. Of which, compound 3a with para-trifluorophenyl substituted oxadiazole showed excellent activity against the susceptible H37Rv and MDR-MTB strain with a MIC values of 8 and 16 µg/ml, respectively.To understand the mechanism of action of these compounds (3a–3i) and identify their putative drug target, molecular docking and dynamics studies were employed against a panel of 20 mycobacterial enzymes reported to be essential for mycobacterial growth and survival. These computational studies revealed polyketide synthase (Pks13) enzyme as the putative target. Moreover, in silico ADMET predictions showed satisfactory properties for these compounds, collectively, making them, particularly compound 3a, promising leads worthy of further optimisation.     

Microwave-assisted synthesis of novel 5-substituted benzylidene amino-2-butyl benzofuran-3-yl-4-methoxyphenyl methanones as antileishmanial and antioxidant agents

A series of 5-substitutedbenzylideneamino-2-butylbenzofuran-3-yl-4-methoxyphenyl methanones is synthesized and evaluated for antileishmanial and antioxidant activities. Compounds 4f (IC₅₀?=?52.0?±?0.09?µg/ml), 4h (IC₅₀?=?56.0?±?0.71?µg/ml) and 4l (IC₅₀?=?59.3?±?0.55?µg/ml) were shown significant antileishmanial when compared with standard sodium stibogluconate (IC₅₀?=?490.0?±?1.5?µg/ml). Antioxidant study revealed that compounds 4i (IC₅₀?=?2.44?±?0.47?µg/ml) and 4l (IC₅₀?=?3.69?±?0.44?µg/ml) have shown potent comparable activity when compared with standard ascorbic acid (IC₅₀?=?3.31?±?0.34?µg/ml). Molecular docking study was carried out which replicating results of biological activity in case of initial hits 4f and 4h suggesting that these compounds have a potential to become lead molecules in drug discovery process. In silico ADME study was performed for predicting pharmacokinetic profile of the synthesised antileishmanial agents and expressed good oral drug like behaviour.     

Alpha-glucosidase inhibitory and antiplasmodial properties of terpenoids from the leaves of Buddleja saligna Willd

In our continuing search for biologically active natural product(s) of plant origin, Buddleja saligna, a South African medicinal plant, was screened in line with its traditional use for antidiabetic (yeast alpha glucosidase inhibitory) and antiplasmodial (against a chloroquine sensitive strain of Plasmodium falciparum (NF54)) activities. The hexane fraction showed the most promising activity with regards to its antidiabetic (IC₅₀?=?260?±?0.112?µg/ml) and antiplasmodial (IC₅₀?=?8.5?±?1.6?µg/ml) activities. Using activity guided fractionation three known terpenoids (betulonic acid, betulone and spinasterol) were isolated from this species for the first time. The compounds displayed varying levels of biological activities (antidiabetic: 27.31?µg/ml?≥?IC₅₀?≥?5.6?µg/ml; antiplasmodial: 14?µg/ml?≥?IC₅₀?≥?2?µg/ml) with very minimal toxicity.     

Design,synthesis and antistaphylococcal activity of marine pyrrole alkaloid derivatives

Abstract

A novel set of 16 hybrids of bromopyrrole alkaloids with aroyl hydrazone were designed, synthesized and evaluated for antibacterial and antibiofilm activities against methicillin-resistant Staphylococcus aureus (MRSA; ATCC 43866), methicillin-susceptible Staphylococcus aureus (MSSA; ATCC 35556) and Staphylococcus epidermidis (SE, S. epidermidis ATCC 35984). Of the 16 tested hybrids, 14 exhibited equal or superior antibiofilm activity against MSSA and MRSA relative to standard vancomycin. Compound 4m showed highest potency with antibiofilm activity of 0.39?µg/mL and 0.78?µg/mL against MSSA and MRSA, respectively. Thus, this compound could act as a potential lead for further development of new antistaphylococcal drugs.