Ewa Cieckiewicz

In vitro anticancer potential of tree extracts from the Walloon Region forest.

Forty-eight extracts from 16 common Belgian trees from the Walloon Region forest were evaluated for in vitro growth inhibitory activity against the human LoVo colon cancer, PC3 prostate cancer, and U373 glioblastoma cell lines. Our study was performed with the aim of selecting plant candidates in order to later isolate new anticancer compounds from an easily affordable tree material. Extracts from Alnus glutinosa (stem bark), Carpinus betulus (leaves and stem bark), Castanea sativa (stem bark), Fagus sylvatica (leaves), Ilex aquifolium (leaves), Larix decidua (leaves), Quercus petraea (stem bark), and Quercus robur (leaves) showed for the first time potent in vitro growth inhibitory activity and could become easily affordable sources of potential new anticancer agents. Root extracts from Robinia pseudoacacia, already known for containing cytotoxic lectins, also showed interesting activity.

Potential anticancer activity of young Carpinus betulus leaves.

As part of our continuing research for anticancer compounds from the Walloon Region forest, EtOAc extract from Carpinus betulus leaves was phytochemically studied, leading to the bioguided isolation of pheophorbide a, which is responsible of anticancer properties of C. betulus young leaves. This compound was identified using nuclear magnetic resonance and mass spectrophotometric data and comparison with a commercial standard. Evaluation of the growth inhibitory activities of pheophorbide a using MTT colorimetric assay and phase-contrast microscopy in various human cancer cell lines confirmed the photoactivable properties of this compound. Our research showed, for the first time, the presence of pheophorbide a, a chlorophyll derived compound, which we quantified in high quantities in young leaves of C. betulus. This is in contrast with the literature which generally describes pheophorbide a as a catabolic product of chlorophyll, then preferentially present in old leaves.

Antiplasmodial activity of Mezoneuron benthamianum leaves and identification of its active constituents

ETHNOPHARMACOLOGICAL RELEVANCE Decoctions of the leaves of M. benthamianum Baill. are used by traditional healers in Guinea to treat malaria and this use was validated by a preliminary clinical assay. AIM OF THE STUDY To evaluate the in vitro antiplasmodial activity and to identify active compounds from extracts of M. benthamianum leaves. MATERIAL AND METHODS Antiplasmodial activity of extracts, fractions and pure compounds was evaluated in vitro against a chloroquine-sensitive strain of Plasmodium falciparum (3D7) using the measurement of the plasmodial lactate dehydrogenase activity. Selectivity of extracts and purified compounds for Plasmodium parasites was evaluated by using WST-1 test on HeLa human cells. Compounds were isolated using normal phase silica gel column chromatography and prepHPLC and their structures elucidated using extensive spectroscopic analysis. RESULTS Hydroethanolic extracts (70% v/v) of M. benthamianum leaves showed a moderate in vitro activity against P. falciparum 3D7, with IC50 in the range 22.5 - 32.6µg/mL, depending on the batch; while a dark precipitate formed during ethanol evaporation showed higher activity (IC50 =6.5µg/mL). The fractionation was performed on this most active fraction and was followed by in vitro antiplasmodial assay. Active compounds (5, 7, 8) belong to several phytochemical classes, contributing together to the global antiplasmodial activity of the hydroethanolic extract against P. falciparum parasite. This study finally allowed the isolation of three diterpenes including two new compounds named Mezobenthamic acids A (1) and B (2) and neocaesalpin H (3), as well as quercetin (4), kaempferol (7), resveratrol (6), gallic acid (9) and its ethylester (5), β-sitosterol glucoside (10) and 13b-hydroxy-pheophorbide a (8). CONCLUSION This study gives some concrete evidence to support the ethnopharmacological use of Mezoneuron benthamianum leaves extract in the management of malaria. The active compounds can be further studied for their antiplasmodial potential, as well as their suitability to be used as quality markers for the standardization of this herbal drug from the Guinean traditional pharmacopeia.

Antiplasmodial activity of Heinsia crinita (Rubiaceae) and identification of new iridoids.

ETHNOPHARMACOLOGICAL RELEVANCE Heinsia crinita is used in traditional medicine for the treatment of febrile illness and erectile dysfunction. Its stem bark powder is found in some peripheral markets in the Democratic Republic of the Congo (DRC) as a remedy against malaria. Investigations were conducted on crude extracts of leaves, fruits and stem barks in view to validate their use and to determine which plant part possesses the best antiplasmodial properties. MATERIALS AND METHODS Different plant parts were extracted with methanol, ethanol and dichloromethane. Based on the preliminary assays, the dichloromethane extract of the stem bark was subjected to fractionation using preparative HPLC system and column chromatography. This step led to the isolation of two new iridoids which had their structures elucidated by NMR, UV, MS and FT-IR spectroscopic techniques. Extracts and pure compounds were tested in vitro against the 3D7 strain of Plasmodium falciparum. The inhibition of the parasite growth was evaluated in vitro by colorimetric method (p-LDH assay) and their cytotoxicity evaluated in vitro against the human non-cancer fibroblast cell line (WI38) through WST1 assay. The in vivo antiplasmodial activity was assessed by the inhibition of Plasmodium berghei growth in infected mice treated with the ethanol extract of H. crinita stem bark at the concentrations of 200 and 300mg/Kg/day per os, using a protocol based on the 4-d suppressive test of Peters and compared to a non-treated negative control group of mice (growth =100%). Finally the antioxidant activity of the same extract was evaluated using ABTS, DPPH and cell-based assays. RESULTS A moderate in vitro antiplasmodial activity was observed for the dichloromethane extract of the stem bark of H. crinita (IC50 =29.2±1.39µg/mL) and for the two new iridoids, lamalbide 6, 7, 8- triacetate (IC50 =16.39±0.43µg/mL) as well as for its aglycone lamiridosin 6, 7, 8-triacetate (IC50 =0.44.56±1.12µg/mL). The ethanolic stem bark extract (200 and 300mg/kg/day, oral route) showed a moderate in vivo antimalarial activity in Plasmodium berghei-infected mice with 27.84±2.75% and 48.54±3.76% of inhibition of the parasite growth, respectively (p<0.01).). This extract displayed high cellular antioxidant activity using dichlorofluorescein-diacetate (DCFDA) on HL-60 monocytes. These crude extracts and pure compounds tested at the higher concentration of 100µg/mL did not show any cytotoxicity against WI38 cells. CONCLUSIONS The results showed that H. crinita extracts possess antimalarial activity and contain some unusual iridoids with moderate antiplasmodial activity, therefore justifying to some extent its traditional use by the local population in DRC for this purpose. This is the first report of the isolation and antiplasmodial activity of these two new iridoids.

Antrocarines A-F, antiplasmodial ergostane steroids from the stem bark of Antrocaryon klaineanum.

During a study on the chemistry and biological activity of Antrocaryon klaineanum Pierre, six new sterols including 4,24(28)-ergostadiene-6α,7α-diol (1), 6α-methoxy-4,24(28)-ergostadiene-7α,20S-diol (2), 6α-methoxy-4,24(28)-ergostadien-7α-ol (3), 20S-hydroxy-24(28)-ergosten-3-one (4), 7α-hydroxy-4,24(28)-ergostadien-3-one (5), and 24(28)-ergostene-3β,6α-diol (6) were characterized by physical and spectroscopic means. The known steroids 7 and 8 were also isolated. The crude extract and the isolated compounds were evaluated for their ability to inhibit the 3D7 strain of Plasmodium falciparum. Compounds 2, 3, and 8 showed potent activity while that of the crude extract was moderate.

In vitro antiplasmodial and cytotoxic activities of sesquiterpene lactones from Vernonia fimbrillifera Less. (Asteraceae)

Abstract Due to the in vitro antiplasmodial activity of leaf extracts from Vernonia fimbrillifera Less. (Asteraceae), a bioactivity-guided fractionation was carried out. Three sesquiterpene lactones were isolated, namely 8-(4’-hydroxymethacrylate)-dehydromelitensin (1), onopordopicrin (2) and 8α-[4’-hydroxymethacryloyloxy]-4-epi-sonchucarpolide (3). Their structures were elucidated by spectroscopic methods (1D and 2D NMR and MS analyses) and by comparison with published data. The isolated compounds exhibited antiplasmodial activity with IC50 values ≤ 5 μg/mL. Cytotoxicity of the compounds against a human cancer cell line (HeLa) and a mouse lung epithelial cell line (MLE12) was assessed to determine selectivity. Compound 3 displayed promising selective antiplasmodial activity (SI > 10).

In-vitro and in-vivo antimalarial activity of caffeic acid and some of its derivatives.

To explore the in‐vitro and in‐vivo antimalarial potential of caffeic acid and derivatives.

Flavonoid composition, cellular antioxidant activity and (myelo)peroxidase inhibition of a Bryonia alba L. (Cucurbitaceae) leaves extract

The aim of the present study consisted in the isolation of flavonoids from the leaves of Bryonia alba L. and evaluation of their antioxidant activity and inhibition on peroxidase‐catalysed reactions.