The Activity of Red Nigerian Propolis and Some of Its Components against Trypanosoma brucei and Trypanosoma congolense.

Propolis is a resin that is gathered by bees from exudates produced by various plants. Its exact chemical composition depends on the plants available near the hive. Bees use propolis to coat the surfaces of the hive, where it acts as an anti-infective. Regardless of the chemical composition of propolis, it is always anti-protozoal, probably because protozoan parasites, particularly Lotmarium passim, are widespread in bee populations. The protozoa Trypanosoma brucei and T. congolense cause disease in humans and/or animals. The existing drugs for treating these diseases are old and resistance is an increasingly severe problem. The many types of propolis present a rich source of anti-trypanosomal compounds-from a material gathered by bees in an environmentally friendly way. In the current work, red Nigerian propolis from Rivers State, Nigeria was tested against T. brucei and T. congolense and found to be highly active (EC50 1.66 and 4.00 µg/mL, respectively). Four isoflavonoids, vestitol, neovestitol, 7-methylvestitol and medicarpin, were isolated from the propolis. The isolated compounds were also tested against T. brucei and T. congolense, and vestitol displayed the highest activity at 3.86 and 4.36 µg/mL, respectively. Activities against drug-resistant forms of T. brucei and T. congolense were similar to those against wild type.

The Antiprotozoal Activity of Papua New Guinea Propolis and Its Triterpenes.

Profiling a propolis sample from Papua New Guinea (PNG) using high-resolution mass spectrometry indicated that it contained several triterpenoids. Further fractionation by column chromatography and medium-pressure liquid chromatography (MPLC) followed by nuclear magnetic resonance spectroscopy (NMR) identified 12 triterpenoids. Five of these were obtained pure and the others as mixtures of two or three compounds. The compounds identified were: mangiferonic acid, ambonic acid, isomangiferolic acid, ambolic acid, 27-hydroxyisomangiferolic acid, cycloartenol, cycloeucalenol, 24-methylenecycloartenol, 20-hydroxybetulin, betulin, betulinic acid and madecassic acid. The fractions from the propolis and the purified compounds were tested in vitro against Crithidia fasciculata, Trypanosoma congolense, drug-resistant Trypanosoma congolense, Trypanosoma b. brucei and multidrug-resistant Trypanosoma b. brucei (B48). They were also assayed for their toxicity against U947 cells. The compounds and fractions displayed moderate to high activity against parasitic protozoa but only low cytotoxicity against the mammalian cells. The most active isolated compound, 20-hydroxybetulin, was found to be trypanostatic when different concentrations were tested against T. b. brucei growth.

Antitrypanosomal and Antileishmanial Activity of Chalcones and Flavanones from Polygonum salicifolium.

Trypanosomiasis and leishmaniasis are a group of neglected parasitic diseases caused by several species of parasites belonging to the family Trypansomatida. The present study investigated the antitrypanosomal and antileishmanial activity of chalcones and flavanones from Polygonum salicifolium, which grows in the wetlands of Iraq. The phytochemical evaluation of the plant yielded two chalcones, 2',4'-dimethoxy-6'-hydroxychalcone and 2',5'-dimethoxy-4',6'-dihydroxychalcone, and two flavanones, 5,7-dimethoxyflavanone and 5,8-dimethoxy-7-hydroxyflavanone. The chalcones showed a good antitrypanosomal and antileishmanial activity while the flavanones were inactive. The EC50 values for 2',4'-dimethoxy-6'-hydroxychalcone against Trypanosoma brucei brucei (0.5 µg/mL), T. congolense (2.5 µg/mL), and Leishmania mexicana (5.2 µg/mL) indicated it was the most active of the compounds. None of the compounds displayed any toxicity against a human cell line, even at 100 µg/mL, or cross-resistance with first line clinical trypanocides, such as diamidines and melaminophenyl arsenicals. Taken together, our study provides significant data in relation to the activity of chalcones and flavanones from P. salicifolium against both parasites in vitro. Further future research is suggested in order to investigate the mode of action of the extracted chalcones against the parasites.

Antiparasitic and Cytotoxic Activity of Bokkosin, A Novel Diterpene-Substituted Chromanyl Benzoquinone From Calliandra portoricensis.

Calliandra portoricensis is a medicinal plant growing freely in Nigeria. It is used traditionally to treat tuberculosis, as an anthelmintic and an abortifacient. Phytochemical fractionation and screening of its root extracts has yielded a novel (5-hydroxy-7-methoxy-4-oxo-1-chromanyl)-4-methoxy-p-benzoquinone (breverin)-substituted cassane diterpene, which was designated bokkosin. It was obtained from column chromatography of the ethyl acetate extract of the roots. The compound was characterized using IR, NMR (1D and 2D) and mass spectral data. Promising antiparasitic activity was observed against the kinetoplastid parasite Trypanosoma brucei brucei, as well as moderate activity against Trypanosoma congolense and Leishmania mexicana and low toxicity in mammalian cells, with the best in vitro EC50 values against T. b. brucei (0.69 µg/mL against a standard laboratory strain, and its multi-drug resistant clone (0.33 µg/mL). The effect on T. b. brucei in culture was rapid and dose-dependent, leading to apparently irreversible growth arrest and cell death after an exposure of just 2 h at 2 × or 4 × EC50. The identification of bokkosin constitutes the first isolation of this class of compound from any natural source and establishes the compound as a potential trypanocide that, considering its novelty, should now be tested for activity against other microorganisms as well.

Novel flavanones with anti-trypanosomal activity isolated from Zambian and Tanzanian propolis samples.

A bioassay-guided phytochemical investigation of propolis samples from Tanzania and Zambia that screened for activity against Trypanosoma brucei has led to the isolation of two novel flavanones with promising antitrypanosomal activity. The compounds were characterized based on their spectral and physical data and identified as 6-(1,1-dimethylallyl) pinocembrin and 5-hydroxy-4″,4″-dimethyl-5″-methyl-5″-H-dihydrofuranol [2″,3″,6,7] flavanone. The two compounds, together with the propolis extracts and fractions, were assayed against a standard drug-sensitive strain of T. b. brucei (s427 wild-type), multi-drug resistant-resistant T. b. brucei (B48), drug-sensitive T. congolense (1L300) and a derived diminazene-resistant T. congolense strain (6C3), and for toxicity against U947 human cells and RAW 246.7 murine cells. Activity against T. b. brucei was higher than against T. congolense. Interestingly, the Tanzanian propolis extract was found to be more active than its fractions and purified compounds in these assays, with an IC50 of 1.20 µg/mL against T. b. brucei. The results of a cytotoxicity assay showed that the propolis extracts were less toxic than the purified compounds with mean IC50 values > 165.0 µg/mL.

European propolis is highly active against trypanosomatids including Crithidia fasciculata.

Extracts of 35 samples of European propolis were tested against wild type and resistant strains of the protozoal pathogens Trypanosoma brucei, Trypanosoma congolense and Leishmania mexicana. The extracts were also tested against Crithidia fasciculata a close relative of Crithidia mellificae, a parasite of bees. Crithidia, Trypanosoma and Leishmania are all members of the order Kinetoplastida. High levels of activity were obtained for all the samples with the levels of activity varying across the sample set. The highest levels of activity were found against L. mexicana. The propolis samples were profiled by using liquid chromatography with high resolution mass spectrometry (LC-MS) and principal components analysis (PCA) of the data obtained indicated there was a wide variation in the composition of the propolis samples. Orthogonal partial least squares (OPLS) associated a butyrate ester of pinobanksin with high activity against T. brucei whereas in the case of T. congolense high activity was associated with methyl ethers of chrysin and pinobanksin. In the case of C. fasciculata highest activity was associated with methyl ethers of galangin and pinobanksin. OPLS modelling of the activities against L. mexicana using the mass spectrometry produced a less successful model suggesting a wider range of active components.

Preliminary studies: the potential anti-angiogenic activities of two Sulawesi Island (Indonesia) propolis and their chemical characterization.

Several studies have previously reported propolis, or its constituents, to inhibit tumour angiogenesis. The anti-angiogenic activity of two Indonesian stingless bee propolis extracts from Sulawesi Island on vascular cells were assessed. Sample D01 was obtained from the outer side of bee hives, while D02 was from the inner side of the same hives. The extracts were profiled by using liquid chromatography coupled to high resolution mass spectrometry. The anti-angiogenic capacity was assessed on HUVECs and placenta-derived pericytes by cell viability, multi-channel wound healing, and CoCl2 based-hypoxia assays. The exact chemical composition has not been confirmed. The most abundant compounds in Indonesian sample D01 seem to be unusual since they do not immediately fall into a clear class. Two of the most abundant compounds have elemental compositions matching actinopyrones. Identification on the basis of elemental composition is not definitive but compounds in D01 are possibly due to unusually modified terpenoids. Sample D02 has abundant compounds which include four related diterpenes with differing degrees of oxygenation and some sesquiterpenes. However, again the profile is unusual. The anti-angiogenic assays demonstrated that D01 elicited a strong cytotoxic effect and a considerable anti-migratory activity on the vascular cells. Although D02 demonstrated a much weaker cytotoxic effect on the cell lines compared to D01, it elicited a substantial protective effect on the pericytes against CoCl2-induced dropout in an experiment to mimic a micro-environment commonly associated with angiogenesis and tumour growth. These results demonstrate modulatory effects of these propolis samples in vascular cells, which requires further investigation.

Isolation of a Novel Flavanonol and an Alkylresorcinol with Highly Potent Anti-Trypanosomal Activity from Libyan propolis.

Twelve propolis samples from different parts of Libya were investigated for their phytochemical constituents. Ethanol extracts of the samples and some purified compounds were tested against Trypanosoma brucei, Plasmodium falciparum and against two helminth species, Trichinella spiralis and Caenorhabditis elegans, showing various degrees of activity. Fourteen compounds were isolated from the propolis samples, including a novel compound Taxifolin-3-acetyl-4'-methyl ether (4), a flavanonol derivative. The crude extracts showed moderate activity against T. spiralis and C. elegans, while the purified compounds had low activity against P. falciparum. Anti-trypanosomal activity (EC50 = 0.7 µg/mL) was exhibited by a fraction containing a cardol identified as bilobol (10) and this fraction had no effect on Human Foreskin Fibroblasts (HFF), even at 2.0 mg/mL, thus demonstrating excellent selectivity. A metabolomics study was used to explore the mechanism of action of the fraction and it revealed significant disturbances in trypanosomal phospholipid metabolism, especially the formation of choline phospholipids. We conclude that a potent and highly selective new trypanocide may be present in the fraction.