Multilevel Approach for the Treatment of Giardiasis by Targeting Arginine Deiminase.

Giardiasis represents a latent problem in public health due to the exceptionally pathogenic strategies of the parasite Giardia lamblia for evading the human immune system. Strains resistant to first-line drugs are also a challenge. Therefore, new antigiardial therapies are urgently needed. Here, we tested giardial arginine deiminase (GlADI) as a target against giardiasis. GlADI belongs to an essential pathway in Giardia for the synthesis of ATP, which is absent in humans. In silico docking with six thiol-reactive compounds was performed; four of which are approved drugs for humans. Recombinant GlADI was used in enzyme inhibition assays, and computational in silico predictions and spectroscopic studies were applied to follow the enzyme's structural disturbance and identify possible effective drugs. Inhibition by modification of cysteines was corroborated using Ellman's method. The efficacy of these drugs on parasite viability was assayed on Giardia trophozoites, along with the inhibition of the endogenous GlADI. The most potent drug against GlADI was assayed on Giardia encystment. The tested drugs inhibited the recombinant GlADI by modifying its cysteines and, potentially, by altering its 3D structure. Only rabeprazole and omeprazole decreased trophozoite survival by inhibiting endogenous GlADI, while rabeprazole also decreased the Giardia encystment rate. These findings demonstrate the potential of GlADI as a target against giardiasis.

Proteomic analysis of oxidized proteins in the brain and liver of the Nile tilapia (Oreochromis niloticus) exposed to a water-accommodated fraction of Maya crude oil.

Crude oil (CO) is a super mixture of chemical compounds whose toxic effects are reported in fish species according to international guidelines. In the current study a proteomic analysis of oxidized proteins (ox) was performed on the brain and liver of Nile tilapia exposed to WAF obtained from relevant environmental loads (0.01, 0.1 and 1.0 g/L) of Maya CO. Results have shown that oxidation of specific proteins was a newly discovered organ-dependent process able to disrupt key functions in Nile tilapia. In control fish, enzymes involved on aerobic metabolism (liver aldehyde dehydrogenase and brain dihydrofolate reductase) and liver tryptophan--tRNA ligase were oxidized. In WAF-treated liver specimens, fructose-bisphosphate aldolase (FBA), ß-galactosidase (ß-GAL) and dipeptidyl peptidase 9 (DPP-9) were detected in oxidized form. oxDPP-9 could be favorable by reducing the risk associated with altered glucose metabolism, the opposite effects elicited by oxFBA and oxß-GAL. oxTrypsin showed a clear adverse effect by reducing probably the hepatocyte capacity to achieve proteolysis of oxidized proteins as well as for performing the proper digestive function. Additionally, enzyme implicated in purine metabolism adenosine (deaminase) was oxidized. Cerebral enzymes of mitochondrial respiratory chain complex (COX IV, COX5B), of glycosphingolipid biosynthesis (ß-N-acetylhexosaminidase), involved in catecholamines degradation (catechol O-methyltransferase), and microtubule cytoskeleton (stathmin) were oxidized in WAF-treated specimens. This response suggests, in the brain, an adverse scenario for the mitochondrial respiration process and for ATP provision as for ischemia/reoxygenation challenges. Proteomic analysis of oxidized proteins is a promising tool for monitoring environmental quality influenced by hydrocarbons dissolved in water.

Synthesis and Biological Evaluation of 2H-Indazole Derivatives: Towards Antimicrobial and Anti-Inflammatory Dual Agents.

Indazole is considered a very important scaffold in medicinal chemistry. It is commonly found in compounds with diverse biological activities, e.g., antimicrobial and anti-inflammatory agents. Considering that infectious diseases are associated to an inflammatory response, we designed a set of 2H-indazole derivatives by hybridization of cyclic systems commonly found in antimicrobial and anti-inflammatory compounds. The derivatives were synthesized and tested against selected intestinal and vaginal pathogens, including the protozoa Giardia intestinalis, Entamoeba histolytica, and Trichomonas vaginalis; the bacteria Escherichia coli and Salmonella enterica serovar Typhi; and the yeasts Candida albicans and Candida glabrata. Biological evaluations revealed that synthesized compounds have antiprotozoal activity and, in most cases, are more potent than the reference drug metronidazole, e.g., compound 18 is 12.8 times more active than metronidazole against G. intestinalis. Furthermore, two 2,3-diphenyl-2H-indazole derivatives (18 and 23) showed in vitro growth inhibition against Candida albicans and Candida glabrata. In addition to their antimicrobial activity, the anti-inflammatory potential for selected compounds was evaluated in silico and in vitro against human cyclooxygenase-2 (COX-2). The results showed that compounds 18, 21, 23, and 26 display in vitro inhibitory activity against COX-2, whereas docking calculations suggest a similar binding mode as compared to rofecoxib, the crystallographic reference.

Antiprotozoal Constituents from Annona cherimola Miller, a Plant Used in Mexican Traditional Medicine for the Treatment of Diarrhea and Dysentery.

BACKGROUND: Annona cherimola Miller (Annonaceae) is a medicinal plant frequently recommended in Mexican traditional medicine for the treatment of gastrointestinal disorders such as diarrhea and dysentery. OBJECTIVE: This work was undertaken to obtain information that support the traditional use of A. cherimola, on pharmacological basis using in vitro and computational experiments. MATERIAL AND METHODS: Bioassay-guided fractionation of the ethanol extract of the leaves of A. cherimola afforded five phenolic compounds: caffeic acid, quercetin, kaempferol, nicotinflorin, and rutin. RESULTS: The in vitro antiprotozoal assay showed that kaempferol was the most potent antiamoebic and antigiardial compound with IC50 values of 7.9 µg/mL for Entamoeba histolytica and 8.7 µg/mL for Giardia lamblia. Computational molecular docking study showed that kaempferol interacted in a region different than metronidazole in the enzyme pyruvate: ferredoxin oxidoreductase (PFOR). CONCLUSION: Considering that PFOR is a target of metronidazole; kaempferol may be a lead compound for the development of novel antiprotozoal agent. Also, these findings give support to the use of A. cherimola in the traditional medicine from México for the treatment of diarrhea and dysentery. SUMMARY: Bioassay-guided fractionation of the ethanol extract of the leaves of Annona cherimola afforded five phenolic compounds: caffeic acid, quercetin, kaempferol, nicotinflorin and rutin. The in vitro antiprotozoal assay showed that kaempferol was the most potent antiamoebic and antigiardial compound with IC50 values of 7.9 µg/mL for Entamoeba histolytica and 8.7 µg/mL for Giardia lamblia. Computational molecular docking study showed that kaempferol interacted in a region different that metronidazole in the enzyme pyruvate: ferredoxin oxidoreductase. Abbreviations used: PFOR:Pyruvate:ferredoxin oxidoreductase, G: lamblia: Giardia lamblia, E: histolytica: Entamoeba histolytica.

Antiprotozoal, antimycobacterial, and anti-inflammatory evaluation of Cnidoscolus chayamansa (Mc Vaugh) extract and the isolated compounds.

Cnidoscolus chayamansa is a medicinal and edible plant known as Chaya, is commonly used as an anti-inflammatory, antiprotozoal, antibacterial agent and as a remedy for respiratory illness, gastrointestinal disorders, and vaginal infections related with the inflammation process. In this paper, we describe the plant's phytochemical analysis and biological activities (antimycobacterial, antibacterial, antiprotozoal, and anti-inflammatory properties) of the CHCl3:MeOH (1:1) leaves extract and isolated compounds, as well as the acute and sub-acute toxic effects. Chemical identification of isolated compounds was performed by 1H- and 13C NMR spectra data. In vitro antibacterial and antimycobacterial activities were determined by disc diffusion and MABA assays, respectively; antiprotozoal test by means of the sub-culture test. Topical and systemic anti-inflammatory effects were tested by TPA and carrageenan assay on BALB/c mice. Moretenol, moretenyl acetate, kaempferol-3,7-dimethyl ether, and 5-hydroxy-7-3',4'-trimethoxyflavanone were the main compounds isolated. The CHCl3:MeOH extract showed antiprotozoal (IC50≤65.29µg/mL), antimycobacterial (MIC≤50µg/mL), and anti-inflammatory activities (ED50=1.66mg/ear and 467.73mg/kg), but was inactive against the bacterial strains tested. The LD50 for extract was >2g/kg. In the sub-acute toxicity test, the extract was administered at 1g/kg for 28days and did not cause lethality or any alteration in hematological and biochemical parameters; in addition, liver, kidney, and spleen histological analysis exhibited no structural changes. Moretenol and moretenyl acetate showed MIC=25µg/mL against Mycobacterium tuberculosis H37Rv and against four monoresistant strains of M. tuberculosis H37Rv. Both compounds exhibited moderate activity against Entamoeba histolytica and Giardia lamblia (IC50≤71.70µg/mL). Kaempferol-3,7-dimethyl ether and 5-hydroxy-7-3',4'-trimethoxy-flavanone were more active than the extract against E. histolytica and G. lamblia, showing IC50 ≤27.43µg/mL. As topical anti-inflammatory agents, moretenol and kaempferol-3,7-dimethyl ether were the most active compounds inhibiting the edema in 30.52 and 26.67%, respectively. Moretenol and moretenyl acetate showed significant antimycobacterial and antiprotozoal activities; in addition, important antiprotozoal effect was detected with kaempferol-3,7-dimethyl ether and 5-hydroxy-7-3',4'-trimethoxyflavanone. The extract and the terpenoids possess good anti-inflammatory activity. The extract did not produce lethality or adverse effects in acute and sub-acute tests.

Antiamoebic and Antigiardial Activity of Clerodane Diterpenes from Mexican Salvia Species Used for the Treatment of Diarrhea.

Terpenoids from Salvia species have been identified to possess biological properties as antiprotozoal agents. Here, we evaluated the antiamoebic and antigiardial activities of 14 known clerodane and modified clerodane-type diterpenes isolated from five Mexican Salvia species against Entamoeba histolytica and Giardia lamblia, and analyzed the effects of the functionalities in decalin ring or in the whole clerodane framework to visualize the structural requirements necessary to produce an antiprotozoal activity. Among these, linearolactone was the most active clerodane diterpene against both protozoa with IC50 values of 22.9 µM for E. histolytica and of 28.2 µM in the case of G. lamblia. In this context it may be a lead compound for the development of novel therapeutic agent for the treatment of diarrhea and dysentery. The remaining diterpenes assayed showed moderate to weak activity against both protozoa. These findings give support to the use of Salvia species in the traditional medicine from México for the treatment of diarrhea.

Antiprotozoal and antimycobacterial activities of Persea americana seeds.

BACKGROUND: Persea americana seeds are widely used in traditional Mexican medicine to treat rheumatism, asthma, infectious processes as well as diarrhea and dysentery caused by intestinal parasites. METHODS: The chloroformic and ethanolic extracts of P. americana seeds were prepared by maceration and their amoebicidal, giardicidal and trichomonicidal activity was evaluated. These extracts were also tested against Mycobacterium tuberculosis H37Rv, four mono-resistant and two multidrug resistant strains of M. tuberculosis as well as five non tuberculosis mycobacterium strains by MABA assay. RESULTS: The chloroformic and ethanolic extracts of P. americana seeds showed significant activity against E. histolytica, G. lamblia and T. vaginalis (IC50 <0.634 µg/ml). The chloroformic extract inhibited the growth of M. tuberculosis H37Rv, M. tuberculosis MDR SIN 4 isolate, three M. tuberculosis H37Rv mono-resistant reference strains and four non tuberculosis mycobacteria (M. fortuitum, M. avium, M. smegmatis and M. absessus) showing MIC values ≤50 µg/ml. Contrariwise, the ethanolic extract affected only the growth of two mono-resistant strains of M. tuberculosis H37Rv and M. smegmatis (MIC ≤50 µg/ml). CONCLUSIONS: The CHCl3 and EtOH seed extracts from P. americana showed amoebicidal and giardicidal activity. Importantly, the CHCl3 extract inhibited the growth of a MDR M. tuberculosis isolate and three out of four mono-resistant reference strains of M. tuberculosis H37Rv, showing a MIC = 50 µg/ml. This extract was also active against the NTM strains, M. fortuitum, M. avium, M. smegmatis and M. abscessus, with MIC values <50 µg/ml.

Incomptines C and D, two heliangolides from Decachaeta incompta and their antiprotozoal activity.

Two new heliangolides, incomptines C (3) and D (4), were isolated from the leaves of Decachaeta incompta. The structures were established by spectroscopic methods and confirmed by X-ray crystallography. The antiprotozoal activity of incomptines C and D was evaluated. Additionally, the chromatographic profile of the leaves and roots extracts were compared to identify incomptines A-D (1-4) in each extract.

Antiprotozoal and Antimycobacterial Activities of Pure Compounds from Aristolochia elegans Rhizomes.

We analyzed the antimycobacterial activity of the hexane extract of rhizomes from Aristolochia elegans. Some compounds of this extract were purified and tested against a group of drug-resistant Mycobacterium tuberculosis strains. We also evaluated their antiprotozoal activities. The hexane extract was active against M. tuberculosis H37Rv at a MIC = 100 µg mL(-1); the pure compounds eupomatenoid-1, fargesin, and (8R,8'R,9R)-cubebin were active against M. tuberculosis H37Rv (MIC = 50 µg mL(-1)), while fargesin presented activity against three monoresistant strains of M. tuberculosis H37Rv and a MDR clinical isolate of M. tuberculosis (MIC < 50 µg mL(-1)). Both the extract and eupomatenoid-1 were very active against E. histolytica and G. lamblia (IC(50) < 0.624 µg mL(-1)); in contrast, fargesin and (8R,8'R,9R)-cubebin were moderately active (IC(50) < 275 µg mL(-1)). In this context, two compounds responsible for the antimycobacterial presented by A. elegans are fargesin and cubebin, although others may exert this activity also. In addition to the antimycobacterial activity, the hexane extract has important activity against E. histolytica and G. lamblia, and eupomatenoid-1 is one of the compounds responsible for the antiparasite activity.

Evaluation of the antiprotozoal activity of neo-clerodane type diterpenes from Salvia polystachya against Entamoeba histolytica and Giardia lamblia.

Chia (Salvia polystachya Ort., Lamiaceae) is frequently used in Mexican traditional medicine to treat dysentery. In this study the main neo-clerodane diterpenes (polystachynes A, B and D, as well as linearolactone) were isolated from the aerial parts of chia, and their antiprotozoal activities toward Entamoeba histolytica and Giardia lamblia trophozoites were evaluated in vitro. Linearolactone was the most potent antiamoebic and antigiardial compound with IC(50) values of 22.9 microM for E. histolytica and 28.2 microM for G. lamblia. Polystachynes A, B and D, showed moderate antiprotozoal activity against both protozoans with IC(50) values ranging from 117.0 to 160.6 microM for E. histolytica and from 107.5 to 134.7 microM for G. lamblia. These data suggest that linearolactone may play an important role in the antidiarrhoeal activity of S. polystachya.

Effect of Mexican medicinal plant used to treat trichomoniasis on Trichomonas vaginalis trophozoites.

Crude methanolic extracts from 22 Mexican medicinal plants were screened for antitrichomonal activity against Trichomonas vaginalis, which is the etiological agent of trichomoniasis. Among the plants tested Carica papaya and Cocos nucifera showed the best antitrichomonal activity with IC(50) values of 5.6 and 5.8 microg/ml, respectively. The extracts of Bocconia frutescens, Geranium mexicanum, and Lygodium venustum showed moderate activity with IC(50) values ranging from 30.9 to 60.9 microg/ml. All the other plant extracts were inactive (IC(50)>100 microg/ml). All extracts tested were less active than metronidazole (IC(50) 0.037 microg/ml), an antiprotozoal drug used as positive control. The results of the antiprotozoal screening support the popular uses of five of the plants tested for the treatment of some urogenital tract disorders in Mexican traditional medicine. However, seeds of Carica papaya and aerial parts of Bocconia frutescens should be used in herbal medicine with care to avoid toxicity.

In vitro susceptibility of Entamoeba histolytica and Giardia lamblia to plants used in Mexican traditional medicine for the treatment of gastrointestinal disorders.

In our search for new antiprotozoal chemotherapy, we collected a selection of 26 plants used in Mexican traditional medicine for the treatment of gastrointestinal disorders. Methanolic extracts of these species were screened for their antiprotozoal activity against Entamoeba histolytica and Giardia lamblia trophozoites using in vitro tests. Among the tested extracts, the derivates of following species showed selectivity and significant antiprotozoal activity: Chiranthodendron pentadactylon, Annona cherimola and Punica granatum were the most active on Entamoeba histolytica with IC50 < 30 microg/ml. Dorstenia contrajerva, Senna villosa and Ruta chalepensis were the most active toward Giardia lamblia with IC50 < 38 microg/ml. The potency of Chiranthodendron pentadactylon (IC50 2.5 microg/ml) on Entamoeba histolytica was close that of to emetine, but far less than metronidazole, drugs used as control. The results of the antiprotozoal screening support the popular uses of the studied species for the treatment of diarrhoea and dysentery in Mexican traditional medicine.

In vitro antiprotozoal activity from the roots of Geranium mexicanum and its constituents on Entamoeba histolytica and Giardia lamblia.

This paper examines the antiprotozoal activity of the dichloromethane-MeOH extract, fractions and pure compounds from the roots of Geranium mexicanum on Entamoeba histolytica and Giardia lamblia. The result indicated that the extract, organic fraction and a pure flavonoid were active against both protozoa with IC50 values ranging from 1.9 to 79.2 microg/ml for Entamoeba histolytica and from 1.6 to 100.4 microg/ml in the case of Giardia lamblia. The main active compound was the flavan-3-ol, (-)-epicatechin. In addition, the moderate active compounds (+)-catechin, tyramine and beta-sitosterol 3-O-beta-D-glucopyranoside, also were isolated. These results support the anecdotal reports for the traditional use of Geranium mexicanum roots in the management of diarrhoea and dysentery, illnesses caused in some cases by enteric protozoa.

Evaluation of albendazole prodrugs in experimental trichinellosis

Background. Two albendazole (ABZ) prodrugs, N-methoxycarbonyl-N'-[(2-nitro-4-propylthio) phenyl] thiourea (compound 2), and N-methoxycarbonyl-N'-[2-nitro-5-propylthio) phenyl] thiourea (compound 3) have recently been synthesized. These compounds showed greater solubility than ABZ itself. Methods. In order to evaluate the biotransformation of compounds 2 and 3 to ABZ and/or ABZ-sulphoxide (ABZ-SO), plasma samples taken from mice treated with the prodrugs were analyzed by HPLC. Also, the anthelmintic activity of compounds 2 and 3 against Trichinella spiralis was evaluated in mice experimentally infected with the prarasite. Results. The presence of ABZ and/or ABZ-SO was demostrated in plasma samples taken at different time intervals after prodrug administration, although their levels were low compared to those reached in mice treated with ABZ. Additionally, prodrugs 2 and 3 were also detected in these samples. In regrad to the anthelmintic activity of ABZ prodrugs, it was shown that compound 3 was more active than compound 2. Additionally, it was as effective as ABZ against T. spiralis pre-adult, adult, and female fecundity. However, compound 3 was not as active sa ABZ against the muscle stage of the parasite. Conclusions. Compound 3 had better anthelmintic activity againts T. spiralis than compound 2. The bioconversion of compounds 2 and 3 to ABZ and/or ABZ-SO was demostrated by HPLC, but they did not reach equivalant concentrarion to that of ABZ. Prodrugs 2 and 3 were also present in plasma samples, suggesting that prodrugs were not efficiently reduced in the intestine of mice