Quorum quenching activity of Andrographis paniculata (Burm f.) Nees andrographolide compounds on metallo-ß-lactamase-producing clinical isolates of Pseudomonas aeruginosa PA22 and PA247 and their effect on lasR gene expression.

Andrographis paniculata (Burm f.) Nees is a tropical plant native to Southeast Asia that has been used as an effective remedy for a wide variety of illnesses in traditional Chinese and Ayurvedic medicine. The antimicrobial activity of its crude extract had been shown to be due to its quorum quenching activity. The study determined the effect of purified extracted compounds from the leaf of A. paniculata, namely: andrographolide, 14-deoxyandrographolide, 14-deoxy-12-hydroxyandrographolide and neoandrographolide on quorum sensing-mediated virulence mechanisms in clinical isolates of metallo-ß-lactamase (MßL)-producing Pseudomonas aeruginosa. Their effect on the expression of the lasR gene, which codes for LasR, a transcription activator protein of the quorum sensing system in P. aeruginosa was also determined using RT-qPCR. All the pure compounds significantly decreased the biofilm formation, protease production and swarming motility of the P. aeruginosa isolates compared to the untreated controls (p < 0.05). Results of the RT-qPCR assay showed that all compounds significantly downregulated the expression of lasR compared to the untreated control (p < 0.05), supporting the position that the lower virulence activities of the treated group were due to quorum quenching activity of the pure compounds. Multiple comparisons using Tukey's HSD analysis revealed that the means of the relative expression of lasR of the isolates treated with the different compounds were not significantly different from each other (p > 0.05), suggesting equal potencies. Results show the potential of the isolated pure compounds from A. paniculata for use as antimicrobial agents as a result of their quorum quenching activities.

Cytotoxic Compounds from Wrightia pubescens (R.Br.).

BACKGROUND: Mixtures of ursolic acid (1) and oleanolic acid (2) (1:1 and 1:2), oleanolic acid (2), squalene (3), chlorophyll a (4), wrightiadione (5), and α-amyrin acetate (6) were isolated from the dichloromethane (CH2 Cl2) extracts of the leaves and twigs of Wrightia pubescens (R.Br.). OBJECTIVES: To test for the cytotoxicity potentials of 1-6. MATERIALS AND METHODS: The antiproliferative activities of 1-6 against three human cancer cell lines, breast (MCF-7) and colon (HT-29 and HCT-116), and a normal cell line, human dermal fibroblast neonatal (HDFn), were evaluated using the PrestoBlue® cell viability assay. RESULTS: Compounds 4, 1 and 2 (1:2), 2, 1 and 2 (1:1), and 5 exhibited the most cytotoxic effects against HT-29 with half maximal inhibitory concentration (IC50) values of 0.68, 0.74, 0.89, 1.70, and 4.07 µg/mL, respectively. Comparing 2 with its 1:1 mixture with 1 (IC50 = 1.70 and 7.18 µg/mL for HT-29 and HCT-116, respectively) and 1:2 mixture with 1 (0.74 and 3.46 µg/mL for HT-29 and HCT-116, respectively), 2 also showed strong cytotoxic potential against HT-29 and HCT-116 (0.89 and 2.33 µg/mL, respectively). Unlike the mixtures which exhibited low effects on MCF-7 (IC50 = 20.75 and 30.06 µg/mL for 1:1 and 1:2, respectively), 2 showed moderate activity against MCF-7 (10.99 µg/mL). Compound 6 showed the highest cytotoxicity against HCT-116 (IC50 = 4.07 µg/mL). CONCLUSION: Mixtures of 1 and 2 (1:1 and 1:2), 2, 3, 4, 5, and 6 from the CH2 Cl2 extracts of the leaves and twigs of W. pubescens (R.Br.) exhibited varying cytotoxic activities. All the compounds except 6 exhibited the strongest cytotoxic effects against HT-29. On the other hand, 6 was most cytotoxic against HCT-116. Overall, the toxicities of 1-6 were highest against HT-29, followed by HCT-116 and MCF-7. All the compounds showed varying activities against HDFn (IC50 < 30 µg/mL). SUMMARY: Mixtures of ursolic acid (1) and oleanolic acid (2) (1:1 and 1:2), oleanolic acid (2), squalene (3), chlorophyll a (4), wrightiadione (5), and α-amyrin acetate (6), isolated from the dichloromethane extracts of the leaves and twigs of Wrightia pubescens (R.Br.), showed varying cytotoxic activities against three human cancer cell lines, breast (MCF-7) and colon (HT-29 and HCT-116), and a normal cell line, human dermal fibroblast-neonatal (HDFn), as evaluated using the PrestoBlue® cell viability assay.Abbreviation Used: IC50: Half maximal inhibitory concentration.

Chemical constituents and bioactivities of Glinus oppositifolius.

OBJECTIVES: To isolate the secondary metabolites from the dichloromethane (DCM) extracts of Glinus oppositifolius; to test for the cytotoxicity of a new triterpene, oppositifolone (1); and to test for the hypoglycemic, analgesic, and antimicrobial potentials of 1, DCM and aqueous leaf extracts of G. oppositifolius. METHODS: The compounds were isolated by silica gel chromatography and identified by nuclear magnetic resonance spectroscopy. The cytotoxicity potential of 1 was tested using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Triterpene 1, DCM, and aqueous leaf extracts were tested for hypoglycemic potential using the oral glucose tolerance test; analgesic potential using the tail-flick assay, and antimicrobial potential using the disc diffusion method. RESULTS: The DCM extracts of G. oppositifolius afforded 1, squalene, spinasterol, oleanolic acid, phytol, and lutein from the leaves; squalene and spergulagenin A from the stems; and spinasterol from the roots. Triterpene 1 was cytotoxic against human colon carcinoma 116 with an IC50 value of 28.7 but did not exhibit cytotoxicity against A549. The aqueous leaf extract at 200 mg/kg body weight (BW) exhibited hypoglycemic activity with a pronounced % blood glucose reduction of 70.76% ±17.4% within 0.5 h after introduction. The DCM leaf extract showed a lower % blood glucose reduction of 18.52% ±13.5% at 200 mg/kg BW within 1.5 h after introduction, while 1 did not exhibit hypoglycemic activity. The samples did not exhibit analgesic property and were inactive against multiple drug resistant bacterial pathogens. CONCLUSION: The compounds responsible for the hypoglycemic activity of G. oppositifolius which are fast acting (0.5 h) are found in the aqueous leaf extract.

Triterpenes from Euphorbia hirta and their cytotoxicity.

AIM: To investigate the chemical constituents of the stems, leaves and roots of Euphorbia hirta, and to test for the cytotoxic and antimicrobial potentials of the major constituents of the plant. METHODS: The compounds were isolated by silica gel chromatography and their structures were elucidated by NMR spectroscopy. The cytotoxicity tests were conducted using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, while the antimicrobial tests employed the agar well method. RESULTS: The air-dried stems of E. hirta afforded taraxerone 1, a mixture of 25-hydroperoxycycloart-23-en-3ß-ol (2a) and 24-hydroperoxycycloart-25-en-3ß-ol (2b) (sample 2) in a 2 : 1 ratio, and another mixture of cycloartenol (3a), lupeol (3b), α-amyrin (3c) and ß-amyrin (3d) (sample 3) in a 0.5 : 4 : 1 : 1 ratio. The air-dried leaves of E. hirta yielded sample 2 in a 3 : 2 ratio, sample 3 in a 2 : 3 : 1 : 1 ratio, phytol and phytyl fatty acid ester, while the roots afforded sample 2 in a 2 : 1 ratio, sample 3 in a 2 : 1 : 1 : 1 ratio, a mixture of cycloartenyl fatty acid ester 4a, lupeol fatty acid ester 4b, α-amyrin fatty acid ester 4c and ß-amyrin fatty acid ester 4d (sample 4) in a 3 : 2 : 1 : 1 ratio, linoleic acid, ß-sitosterol and squalene. Compound 1 from the stems, sample 2 from the leaves, and sample 3 from the stems were assessed for cytotoxicity against a human cancer cell line, colon carcinoma (HCT 116). Sample 2 showed good activity with an IC50 value of 4.8 µg·mL(-1), while 1 and sample 3 were inactive against HCT 116. Sample 2 was further tested for cytotoxicity against non-small cell lung adenocarcinoma (A549). It showed good activity against this cell line with an IC50 value of 4.5 µg·mL(-1). Antimicrobial assays were conducted on 1 and sample 2. Results of the study indicated that 1 was active against the bacteria: Pseudomonas aeruginosa and Staphylococcus aureus, but was inactive against Escherichia coli and Bacillus subtilis. Sample 2 was active against the bacteria: Pseudomonas aeruginosa, Staphylococcus aureus and Escherichia coli and fungi: Candida albicans and Trichophyton mentagrophytes. It was inactive against Bacillus subtilis and Aspergillus niger. CONCLUSIONS: The triterpenes: 2a, 2b, 3a, 3b, 3c and 3d were obtained from the stems, roots and leaves of E. hirta. Taraxerol (1) was only isolated from the stems, the leaves yielded phytol and phytyl fatty acid esters, while the roots afforded 4a-4d, linoleic acid, ß-sitosterol, and squalene. Triterpene 1 and sample 2 were found to exhibit antimicrobial activities. Thus, these compounds are some of the active principles of E. hirta which is used in wound healing and the treatment of boils. The cytotoxic properties of sample 2 imply that triterpenes 2a and 2b contribute to the anticancer activity of E. hirta.

Angio-suppressive triterpenoids from Ardisia cf. elliptica (subgenus Tinus) on duck (Anas platyrynchos L.) chorioallantoic membrane.

The dichloromethane extract of the air-dried leaves of Ardisia cf. elliptica (subgenus Tinus) afforded a mixture of bauerenol (1a), α-amyrin (1b) and ß-amyrin (1c). Their structures were identified by NMR spectroscopy. Mixtures of the triterpenes (1a-1c) at ratios of 2 : 2 : 1, 2 : 2 : 3 and 1 : 1 : 1 were tested for their angio-suppressive effects on duck chorioallantoic membrane (CAM). All three ratios were found to be effective in restricting inter-capillary length, while 1a-1c (2 : 2 : 1) was most effective in reducing branch point density with 100% CAM viability and embryo survivability, suggesting a high impact angio-suppressive potential of 1a-1c (2 : 2 : 1).

Chemical constituents of Cinnamomum cebuense.

AIM: To investigate the chemical constituents of Cinnamomum cebuense, an endemic and critically endangered tree found only in Cebu, Philippines. METHODS: The compounds were isolated by silica gel chromatography. The structures of the isolates were elucidated by NMR spectroscopy. RESULTS: The dichloromethane (DCM) extract of the bark of C. cebuense afforded a new monoterpene natural product 1 and a new sesquiterpene 2, along with the known compounds, 4-hydroxy-3-methoxycinnamaldehyde (3), 4-allyl-2-methoxyphenol (4), α-terpineol (5) and humulene (6). The DCM extract of the leaves of C. cebuense yielded 6, ß-caryophyllene (7), squalene (8), and a mixture of α-amyrin (9), ß-amyrin (10) and bauerenol (11). The structures of 1-7 were elucidated by extensive 1D and 2D NMR spectroscopy, while the structures of 8-11 were identified by comparison of their (13)C NMR data with those reported in the literature. CONCLUSION: The bark of C. cebuense afforded monoterpenes, sesquiterpenes and phenolics, while the leaves yielded sesquiterpenes and triterpenes.

A new triterpene from Barringtonia asiatica.

The freeze-dried bark of Barringtonia asiatica afforded a new triterpene: (3ß,11α)-11-hydroxyolean-12-en-3-yl palmitate (1). The bark also yielded mixtures of (3ß)-olean-12-en-3-yl palmitate (2a), (3ß)-urs-12-en-3-yl palmitate (2b) and (3ß)-olean-18-en-3-yl palmitate (2c) in a 2:1:4 ratio; ß-amyrin (3a), α-amyrin (3b) and germanicol (3c) in a 3:1:4 ratio; 22-O-tigloylcamelliagenin A (4a) and betulinic acid (4b) in a 2:1 ratio; olean-12-en-3ß,16ß,22α-triol (5), ß-sitosterol, spinasterol, squalene and trilinolein. The roots yielded 2a-c and 3a-c as well as trilinolein, spinasterol and squalene, while the flowers afforded verimol k (6), linoleic acid, spinasterol, squalene, phytyl fatty acid ester and trilinolein. Compounds 1-4 and 6 were tested for antimicrobial property against seven microorganisms. All compounds tested exhibited slight activity against Candida albicans and were found inactive against Escherichia coli, Bacillus subtilis, Trichophyton mentagrophytes and Aspergillus niger. Except for the mixture of 4a and 4b that proved to be inactive, all the compounds were slightly active against the bacterium Staphylococcus aureus, while 3a-c were slightly active against Pseudomonas aeruginosa.

Antimicrobial isothiocyanates from the seeds of Moringa oleifera Lam.

4-(alpha-L-Rhamnosyloxy)benzyl isothiocyanate (1) and 4-(4'-O-acetyl-alpha-L-rhamnosyloxy)-benzyl isothiocyanate (2) isolated from Moringa oleifera seeds were screened for their antibacterial activities against Staphylococcus aureus, Staphylococcus epidermidis, Bacillus subtilis, Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, and Pseudomonas aeruginosa, and for their antifungal activities against Candida albicans, Trichophyton rubrum, and Epidermophyton floccosum using the disk diffusion method. Isothiocyanates 1 and 2 were found active at the lowest inhibitory concentration of 1 mg/ml against all Gram-positive bacteria tested (S. aureus, S. epidermidis, B. subtilis) and against the dermatophytic fungi E. floccosum and T. rubrum. Statistically significant differences were found between the mean inhibition zones (IZ) of 1 and 2 and the standard drugs, ofloxacin and clotrimazole. The minimum inhibitory concentration (MIC) values confirmed the good antimicrobial activity of 1 and 2 against S. aureus, good to moderate activity against S. epidermidis, moderate activity against B. subtilis, and weak activity against E. floccosum and T. rubrum. The in vitro bactericidal effect of 1 and 2 against the Gram-positive bacterial strains tested is suggested by MBC:MIC ratios of 2:1.

Cytotoxic cardenolide and sterols from Calotropis gigantea.

The dichloromethane extract from the leaves of Calotropis gigantea Linn. was strongly cytotoxic against non-small cell lung carcinoma (A549), colon carcinoma (HCT 116) and hepatocellular carcinoma (Hep G2), and non toxic to Chinese hamster ovary (AA8). The extract afforded uscharin (1), 3,5,8-trihydroxy-24-methylcholest-6,22-diene (2), a mixture of (24R)-3beta-hydroxy-24-ethylcholest-5-en-7-one (3a) and 6beta-hydroxy-24-ethylcholest-4,22-dien-3-one (3b), and another mixture of (24R)-24-ethylcholest-4-en-3-one (4a) and (24S)-24-ethylcholest-4,22-dien-3-one (4b). Cardenolide 1 exhibited extreme toxicity to A549, HCT 116 and Hep G2 with IC50 values of 0.003 microg/mL, 0.013 microg/mL, and 0.018 microg/mL, respectively, while sample 3 exhibited an IC50 of 1.35 microg/mL, 4.46 microg/mL, and 3.83 microg/mL, respectively.

Bioactivities of triterpenes and a sterol from Syzygium samarangense.

Cycloartenyl stearate (1a), lupenyl stearate (1b), sitosteryl stearate (1c), and 24-methylenecycloartanyl stearate (1d) (sample 1) from the air-dried leaves of Syzygium samarangense exhibited potent analgesic and anti-inflammatory activities at effective doses of 6.25 mg/kg body weight and 12.5 mg/kg body weight, respectively. Sample 1 also exhibited negligible toxicity on zebrafish embryonic tissues. There were incidences of mortality upon direct exposure of sample 1 to dechorionated embryos, but higher mortality and aberration were observed during intact chorion treatment.

A bioactive sesquiterpene from Bixa orellana.

A dichloromethane extract of the air-dried leaves of Bixa orellana afforded ishwarane 1, phytol 2, polyprenol 3, and a mixture of stigmasterol 4a and sitosterol 4b by silica gel chromatography. The structure of 1 was elucidated by extensive 1D and 2D NMR spectroscopy. Compound 1 at three doses (25, 50, and 100 mg/kg BW) was tested for prophylactic, gastrointestinal motility, analgesic, hypoglycemic, and antimicrobial potentials. Results of the prophylactic assay demonstrated the anti-toxic property of 1 at 100 mg/kg BW. A 50 mg/kg BW dose of 1 resulted in a more propulsive movement of the gastrointestinal tract (88.38 ± 13.59%) compared to the negative control (78.47 ± 10.61%). Tail flick and acetic acid writhing tests indicated that 100 mg/kg BW 1 had minimal analgesic activity. Compound 1 demonstrated no hypoglycemic potential on the animals tested. Compound 1 exhibited moderate antifungal activity against C. albicans, low activity against T. mentagrophytes, and low antibacterial activity against E. coli, S. aureus, and P. aeruginosa. It was inactive against B. subtilis and A. niger.

Hypoglycaemic effects of tea extracts and ent-kaurenoic acid from Smallanthus sonchifolius.

Hypoglycaemic activity was observed in normoglycaemic mice orally administered with the aqueous Smallanthus sonchifolius leaf tea extract, alloxan-induced diabetic mice orally administered with ent-kaurenoic acid (1), and normoglycaemic mice intraperitoneally administered with 1 from S. sonchifolius leaves. A single dose administration of 50 mg kg(-1) BW yacon leaf tea extract demonstrated immediate but relatively short hypoglycaemic activity, with significant effects observed during 1-2 h. Similarly, administration with 100 mg kg(-1) BW yacon leaf tea extract obtained by heavy stirring in hot water demonstrated a more potent activity compared to the positive control at 1.5-2.0 h. Oral administration of 1 did not affect the blood glucose level of the alloxan-induced diabetic mice, but a single intraperitonial injection of 10 mg kg(-1) BW in normoglycaemic mice had consistent percent blood glucose reduction persisting from 1 to 2 h observation periods.

New carvotanacetone derivatives from Sphaeranthus africanus.

The dichloromethane extract of the air-dried leaves of Sphaeranthus africanus afforded four new carvotanacetone derivatives (1, 2, 3A, and 3B). Their structures were elucidated by extensive 1D and 2D NMR spectroscopy. Compounds 1- 3 exhibited antibacterial activity against Staphylococcus aureus and Pseudomonas aeruginosa and antifungal activity against Candida albicans, Trichophyton mentagrophytes and Aspergillus niger.

Bioactive triterpenes from Diospyros blancoi.

The ethyl acetate extract of the air-dried leaves of Diospyros blancoi afforded isoarborinol methyl ether (1), a mixture of alpha-amyrin palmitate, alpha-amyrin palmitoleate, beta-amyrin palmitate and beta-amyrin palmitoleate (2) in a 13 : 4 : 3 : 1 ratio, and squalene. The structures of 1, 2 and squalene were elucidated by extensive 1D and 2D NMR spectroscopy. Compounds 1 and 2 exhibited antimicrobial activity against Escherichia coli, Pseudomonas aeruginosa, Candida albicans, Staphylococcus aureus and Trichophyton mentagrophytes, and were found inactive against Bacillus subtilis and Aspergillus niger. Sample 2 exhibited significant analgaesic and anti-inflammatory activities.

A new rearranged dolabellane diterpene from the soft coral Clavularia inflata.

A new dolabellane type diterpene 1 has been isolated through its acetate 1a. The structure of 1a was elucidated by extensive 1D and 2D NMR spectroscopy and confirmed by mass spectrometry. The structure of 1 was deduced by comparison of its NMR spectral data with those of 1a, while its relative stereochemistry was deduced by NOESY. The absolute stereochemistry of C-7 was determined by analyses of 1 separately esterified with R and S O-mandelic acids.

Secondary metabolites from Tectona philippinensis.

The air-dried leaves of Tectona philippinensis, an endemic and endangered Philippine medicinal plant, afforded 5-hydroxy-3,7,4'-trimethoxyflavone (1), 5,4'-dihydroxy-3,7-dimethoxyflavone (2), squalene (3), a mixture of lupeol (4a) and beta-amyrin (4b), chlorophyllide a (5), and hydrocarbons. Antimicrobial tests on 1 and 2 indicated low antifungal activity against the fungi, Candida albicans and Trichophyton mentagrophytes. Compound 1 was also found to have low antibacterial activity against Escherichia coli and Pseudomonas aeruginosa.

A new sesquiterpene from Artemisia vulgaris.

The dichloromethane extract of the air-dried leaves of Artemisia vulgaris afforded a new sesquiterpene 1, caryophyllene oxide, phytyl fatty acid esters, squalene, stigmasterol and sitosterol. The structure of 1 was elucidated by extensive one- and two-dimensional nuclear magnetic resonance spectroscopy.

Chromomoric acid derivatives from Tectona philippinensis.

The air-dried leaves of Tectona philippinensis, an endemic and endangered Philippine medicinal tree, afforded four new chromomoric acid derivatives ( 1, 2, 3a, and 3b). Their structures were elucidated by extensive 1D and 2D NMR spectroscopy. Antimicrobial testing was carried out on 1- 3 against a panel of bacteria and fungi.

A marine verticillane diterpenoid from Cespitularia erecta.

The dichloromethane extract of Cespitularia erecta afforded a new verticillane diterpenoid (1) and sarcophytol A (2) by silica gel chromatography. The structure of 1 was elucidated by extensive 1D and 2D NMR spectroscopy.

New xenicanes from Xenia viridis.

The dichloromethane extract of Xenia viridis afforded two new xenicane diterpenoids (3 and 4) by silica gel chromatography. The oxirane ring of 3 was found to be susceptible to a slow ring opening under acidic conditions to afford 5. Their structures were elucidated by extensive 1D and 2D NMR spectroscopy.