Jörg Heilmann

The influence of glutathione and cysteine levels on the cytotoxicity of helenanolide type sesquiterpene lactones against KB cells.

The biological activities of sesquiterpene lactones have been attributed to their reactivity with the cysteine residues of functional proteins forming covalent bonds via Michael type addition. In the present study we investigated the influence of different L-cysteine (cys) and glutathione (GSH) concentrations on the cytotoxicity of the sesquiterpene lactones (STLs) helenalin, 11alpha,13-dihydrohelenalin acetate and chamissonolide against KB cells. Due to the significantly higher reactivity of the alpha-methylene-gamma-lactone (ML) towards cys as compared with the cyclopentenone (CP) site at physiological pH, addition of 20, 50 and 100 molar equivalents of cys decreased the cytotoxicity of helenalin and chamissonolide, whereas the cytotoxicity of 11alpha,13-dihydrohelenalin acetate remained unaffected. In contrast, the influence of GSH addition on the cytotoxicity of 11alpha,13-dihydrohelenalin acetate depends on the concentration of GSH added. Concentration-effect curves obtained for chamissonolide and GSH resembled the decline in cytotoxicity after cys addition. Helenalin showed a biphasic shape of the concentration-effect curve for the 100:1 GSH/helenalin ratio resembling at higher doses the chamissonolide and in lower doses the 11alpha,13-dihydrohelenalin acetate curve at 50-fold excess. These results can be explained by the different reactivity and equilibrium conditions for thiol addition of the two reactive centers of bifunctional STLs in cellular test systems and verified a clear correlation between the different reactivity of their electrophilic centers and the observed biological effects in in-vitro cell systems.

Design, synthesis and pharmacological evaluation of hybrid molecules out of quinazolinimines and lipoic acid lead to highly potent and selective butyrylcholinesterase inhibitors with antioxidant properties

A set of hybrid molecules were synthesized out of lipoic acid, alpha,omega-diamines of different lengths serving as spacers, and cholinesterase (ChE) inhibiting [2,1-b]quinazolinimines. Depending on the length of the alkylene spacer the amide hybrids are inhibitors of acetylcholinesterase (AChE) with inhibitory activities of 0.5-4.6microM and inhibitors of butyrylcholinesterase (BChE) with activities down to 5.7nM, therefore greatly exceeding the inhibitory activities of the parent quinazolinimines by factors of up to 1000. Due to increasing activity at BChE with increasing length of the alkylene spacer approximately 100-fold selectivity toward BChE is reached with a hepta- and an octamethylene spacer. Kinetic measurements reveal competitive and reversible inhibition of both ChEs by the hybrids. Furthermore, cell viability and antioxidant activity (using the ORAC-fluorescein assay) of several hybrids were evaluated, showing cytotoxicity at concentrations from 3.7 to 10.2microM and antioxidant properties are in the range of 0.4-0.8 Trolox equivalents (lipoic acid=0.6).

Determining the cytotoxicity of catanionic surfactant mixtures on HeLa cells

The cytotoxicity of commonly used synthetic surfactants and catanionic mixtures of those was evaluated using MTT on HeLa cells. The 50% inhibition concentration (IC(50)) for MTT reduction was calculated. The effect on chain length increase and inclusion of polyoxyethylene groups on the toxicity was tested on single surfactant systems. A general trend of increasing toxicity with increasing chain length and the presence of polyoxyethylene groups was observed. The measured IC(50) values of catanionic systems lie between those of participating surfactants. The increase in toxicity as the cationic surfactant is added to the anionic one is however not linear. A steep decrease of the IC(50) values (and therefore increase in the toxic properties) is observed immediately already at low concentrations of the cationic surfactants. This behavior is analogous to the enzyme activity in catanionic microemulsions.

Xanthohumol, a chalcon derived from hops, inhibits hepatic inflammation and fibrosis.

Xanthohumol (XN) is a major prenylated chalcone found in hops, which is used to add bitterness and flavor to beer. In this study, we first investigated the effects of XN on hepatocytes and hepatic stellate cells (HSC), the central mediators of liver fibrogenesis. XN inhibited the activation of primary human HSC and induced apoptosis in activated HSC in vitro in a dose dependent manner (0-20 microM). In contrast, XN doses as high as 50 microM did not impair viability of primary human hepatocytes. However, in both cell types XN inhibited activation of the transcription factor NFkappaB and expression of NFkappaB dependent proinflammatory genes. In vivo, feeding of XN reduced hepatic inflammation and expression of profibrogenic genes in a murine model of non-alcoholic steatohepatitis. These data indicate that XN has the potential as functional nutrient for the prevention or treatment of non-alcoholic steatohepatitis or other chronic liver disease.

Antiprotozoal activity and cytotoxicity of novel 1,7-dioxadispiro[5.1.5.2]pentadeca-9,12-dien-11-one derivatives from Amomum aculeatum.

Cytotoxicity against the KB cancer cell line as a lead bioactivity-guided fractionation of the petroleum ether ext. of rhizomes of Amomum aculeatum RoxB. led to the isolation of three novel dioxadispiro[5.1.5.2]-pentadeca-9,12-dien-11-one derivs. The structures of aculeatin A, aculeatin B, and aculeatin C were established as rel-(2R,4R,6S)- and rel-(2R,4R,6R)-4-hydroxy-2-tridecyl-1,7-dioxadispiro[5.1.5.2]pentadeca-9,12-dien-11-one and rel-(2R,4R,6R)-2-[4-(3-dodecyl-2-heptyl-3-hydroxy-6-oxocyclohexa-1,4-dienyl)-2-oxobutyl]-4-hydroxy-1,7-dioxadispiro[5.1.5.2]pentadeca-9,12-dien-11-one resp. by extensive spectroscopic analyses, particularly 13C-NMR, inverse-gated 13C, HMQC, HMBC, NOESY, and INADEQUATE NMR expts. as well as mass spectrometry. The aculeatins represent a novel type of natural products. All compds. showed high cytotoxicity against the KB cell line: IC50=1.7 micro M; IC50=2.0 micro M; IC50 = 1.6 micro M resp. Addnl. testing against two Plasmodium falciparum strains as well as against trypomastigote forms of Trypanosoma brucei rhodesiense and Trypanosoma cruzi showed strong activities, particularly against P. falciparum strain K1 (IC50=0.18micro M; IC50=0.43micro M; IC50=0.37 micro M).

Tacrine-Silibinin Codrug Shows Neuro- and Hepatoprotective Effects in Vitro and Pro-Cognitive and Hepatoprotective Effects in Vivo

A codrug of the anti-Alzheimer drug tacrine and the natural product silibinin was synthesized. The codrugs biological and pharmacological properties were compared to an equimolar mixture of the components. The compound showed potent acetyl- and butyrylcholinesterase inhibition. In a cellular hepatotoxicity model, analyzing the influence on viability and mitochondria of hepatic stellate cells (HSC), the toxicity of the codrug was markedly reduced in comparison to that of tacrine. Using a neuronal cell line (HT-22), a neuroprotective effect against glutamate-induced toxicity could be observed that was absent for the 1:1 mixture of components. In subsequent in vivo experiments in rats, in contrast to the effects seen after tacrine treatment, after administration of the codrug no hepatotoxicity and no induction of the cytochrome P450 system were noticed. In a scopolamine-induced cognitive impairment model using Wistar rats, the codrug was as potent as tacrine in reversing memory dysfunction. The tacrine-silibinin codrug shows high AChE and BChE inhibition, neuroprotective effects, lacks tacrines hepatotoxicity in vitro and in vivo, and shows the same pro-cognitive effects in vivo as tacrine, being superior to the physical mixture of tacrine and silibinin in all these regards.

Cytotoxic cardenolides and antibacterial terpenoids from Crossopetalum gaumeri

From the methanol extract of the roots of (Crossopetalum gaumeri, four new highly cytotoxic cardenolides, securigenin-3beta-O-beta-6-deoxyguloside (2), 19-hydroxy-sarmentogenin-3beta-O-beta-6-deoxyguloside (4), sarmentogenin-3beta-O-[alpha-allosyl-(1-->4)-beta-6-deoxy alloside] (5), and securigenin-3beta-O-[alpha-allosyl-(1-->4)-beta-6-deoxyal loside] (6) were isolated. The dichloromethane extract afforded the new diterpene 3,15-dihydroxy-18-norabieta-3,8,11,13-tetraene (7) as well as the new triterpene 2,3,7-trihydroxy-6-oxo-1,3,5(10),7-tetraene-24-nor-friedelane-29-o ic acid methylester (11). The new terpenoids lack cytotoxicity and the antibacterial activity is moderate to low.

Secondary metabolites from Stachys ionica

Nine flavonoids, two iridoids, and one sesquiterpene were isolated from Stachys ionica. Its chemical profile appears to be completely different from other species of Stachys growing in Greece. This differentiation is correlated to the distribution of S. ionica, confined to the Ionian Islands, and therefore, well-isolated from the rest of the species.

Neuroprotective Tri- and Tetracyclic BChE Inhibitors Releasing Reversible Inhibitors upon Carbamate Transfer

Tri- and tetracyclic nitrogen-bridgehead compounds were designed and synthesized to yield micromolar cholinesterase (ChE) inhibitors. Structure-activity relationships identified potent compounds with butyrylcholinesterase selectivity. These compounds were selected as starting points for the design and synthesis of carbamate-based (pseudo)irreversible inhibitors. Compounds with superior inhibitory activity and selectivity were obtained and kinetically characterized also with regard to the velocity of enzyme carbamoylation. Structural elements were identified and introduced that additionally showed neuroprotective properties on a hippocampal neuronal cell line (HT-22) after glutamate-induced intracellular reactive oxygen species generation. We have identified potent and selective pseudoirreversible butyrylcholinesterase inhibitors that release reversible inhibitors with neuroprotective properties after carbamate transfer to the active site of cholinesterases.

Medicinal plant use in Vanuatu: A comparative ethnobotanical study of three islands

ETHNOPHARMACOLOGICAL RELEVANCE Our study shows that large parts of Vanuatus medicinal flora remain unexplored and that a high variability of medicinal plant knowledge between islands exists. AIM OF THE STUDY The following questions are comparatively analyzed for three islands of Vanuatu: who are the medicinal plant specialists and how important is their knowledge today? Which plants are used to treat common diseases? MATERIALS AND METHODS On Loh, Ambrym and Aneityum plant related information was collected using semi-structured interviews, transect walks and participant observation. A total of 29 medicinal plant specialists were interviewed. RESULTS Medicinal plant specialists are either peasants or people with a high rank in the local social system such as members of the chiefs family or priests. Their knowledge may be very broad (Loh, Aneityum) or specialized on specific diseases (Ambrym). Medicinal plant knowledge is transmitted family and gender specific (Loh) or gender and family independent (Ambrym and Aneityum). Overall, 133 medicinal plant species were documented of which 117 are new to Vanuatus ethnopharmacopoeia. Mainly members of the Euphorbiaceae and Fabaceae, followed by Asteraceae, Convolvulaceae, Moraceae and Zingiberaceae are utilized. The majority of documented species are trees (33%), followed by herbs (22%) and shrubs (21%). Leaves accounted for the highest number of use reports (43%). The highest diversity of medicinal plants is found for the most common diseases such as skin, gastrointestinal, respiratory system and urogenital system diseases. Only a small overlap of taxa between the islands was found. CONCLUSIONS The biocultural diversity of Vanuatu is reflected in the variability of medicinal plant knowledge and differences in the traditional medicinal system between the three islands investigated. Traditional medicine is more vital on remote islands. The better connected the islands are to the main city, the more dominant western medicine becomes and traditional medicine mainly remains to treat illnesses with a magical origin.