Herbert Kolodziej

Abietane diterpenoids and triterpenoic acids from Salvia cilicica and their antileishmanial activities

Bioguided-fractionation of an acetone extract of the roots of Salvia cilicica (Lamiaceae) led to isolation of two new diterpenes, 7-hydroxy-12-methoxy-20-nor-abieta-1,5(10),7,9,12-pentaen-6,14-dione and abieta-8,12-dien-11,14-dione (12-deoxy-royleanone), together with oleanolic acid, ursolic acid, ferruginol, inuroyoleanol and cryptanol. Their structures were determined spectroscopically, which included HREIMS and 2D NMR spectroscopic analysis. The new abietane derivatives showed appreciable in vitro antileishmanial activity against intracellular amastigote forms of both Leishmania donovani (IC(50) values of 170 and 120 nM, respectively) and Leishmania major (IC(50) values of 290 and 180 nM, respectively). The triterpenoic acids were found to be potently active against amastigote (IC(50) values of 7-120 nM) and moderately active against promastigote stages (IC(50) values of 51-137 nM) of the two Leishmania species.

Identification of flavan-3-ols and procyanidins by high-performance liquid chromatography and chemical reaction detection

A method for characterizing flavan-3-ols and procyanidins by HPLC separation in connection with on-line UV detection and chemical reaction detection (CRD) is described. The post-column derivatization of the polyphenols with 4-dimethylaminocinnamaldehyde in the presence of sulphuric acid improves the sensitivity as compared to UV detection. The extent of improvement strongly depends on the structure of the procyanidin, hence resulting in characteristic absorbance ratios (CRD/UV) for the individual compounds under given reaction conditions. The ratios are further influenced by the solvent composition, by reaction time and by temperature.

Pharmacological profile of extracts of Pelargonium sidoides and their constituents

In areas of southern Africa, aqueous extracts from the roots of Pelargonium sidoides are employed to cure various disorders. Nowadays, a modern formulation (EPs 7630), elaborated from the traditional herbal medicine, is successfully used for the treatment of respiratory tract diseases. We previously observed that root extracts of P. sidoides and their representative constituents exhibit moderate antibacterial and significant immunomodulatory capabilities. The present study was performed to further assess the efficacy and mode of action for these pharmacological activities. The results indicate that P. sidoides extracts may well possess antimycobacterial activity as claimed in traditional uses. Furthermore, significant antibacterial properties against multi-resistant Staphylococcus aureus strains as well as TNF-inducing potencies and prominent interferon-like activities in supernatants of sample-activated bone marrow-derived macrophages were observed using several functional assays. In addition, EPs 7630 stimulated the synthesis of IFN-beta in MG 63 cells as demonstrated by a specific enzyme immunoassay. For gallic acid, a characteristic constituent, evidence for the expression of iNOS and TNF-alpha transcripts in stimulated RAW 264.7 cells and, hence, activation at the transcriptional level was revealed by RT-PCR. The present results, when taken together with the recently reported pharmacological activities, provide for a rationale basis of the utilization of EPs 7630 in the treatment of respiratory tract infections.

Antibacterial activity of simple coumarins: structural requirements for biological activity.

Abstract The antibacterial activity of a series of simple coumarins was evaluated against 8 microorganisms, including three Gram-positive (Staphylococcus aureus, beta-hemolytic Streptococcus and Streptococcus pneumoniae) and five Gram-negative bacteria (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis and Haemophilus influenzae), using the microdilution broth method. The coumarins tested showed broad diversity regarding growth inhibitory activity with minimum inhibitory concentrations ranging from 0.9 to >12.4 μᴍ. This study, presenting the first systematic analysis of structure-activity relationships among this group of coumarins, revealed some interesting structural requirements. While coumarins with a methoxy function at C-7 and, if present, an OH group at either the C-6 or C-8 position are invariably effective against the spectrum of tested standard bacteria (Gram-negative microorganisms including the Gram-positive bacterium Staphylococcus aureus), the presence of an aromatic dimethoxy arrangement is apparently favourable against those microorganisms which require special growth factors (beta-hemolytic Streptococcus, Streptococcus pneumoniae and Haemophilus influenzae). A combination of these structural features, two methoxy functions and at least one additional phenolic group as reflected by the highly oxygenated coumarins, identify promising candidates with antibacterial broad-spectrum activity.

Synthesis and characterization of procyanidin dimers as their peracetates and octamethyl ether diacetates

Abstract Condensation of (2R,3S,4R or S)-leucocyanidin or the 5,7,3′,4′-tetramethyl ether of (2R,3R,4S)-leucocyanidin with flavan-3-ols yielded dimeric flavanoids which were converted to their octamethyl ether diacetates, or the deca-acetates for the 2,3-trans-procyanadin series. Comparison is made of the 1H NMR spectra of the deca-acetate and octamethyl ether diacetate derivatives which lead to useful diagnostic shift parameters characteristic of their structures. Condensations afforded a novel biflavanoid with a 3,4-cis-configuration and a triflavanoid of ‘mixed’ stereochemistry.

Highly oxygenated coumarins from Pelargonium sidoides

Abstract The range of natural simple coumarins is extended by discovery of the highly oxygenated representatives 6,8-dihydroxy-7-methoxycoumarin, 6,7,8-trihydroxycoumarin, 6,8-dihydroxy-5,7-dimethoxycoumarin, and 7-acetoxy-5,6-dimethoxycoumarin. They are accompanied in the roots of Pelargonium sidoides by the widely occurring scopoletin, associated withtthe uncommon analogues 5,6,7-trimethoxycoumarin, 7-hydroxy-5,6-dimethoxycoumarin (umckalin), and 5,6,7,8-tetramethoxycoumarin (artelin), these being reported from a plant source for the second time. The structures of these compounds were established from chemical and spectroscopic studies. Differentiation between oxygenation patterns is discussed on the basis of chemical shift data.

Flavan-3-ols and proanthocyanidins from Cistus incanus

Abstract Four monomeric and seven oligomeric flavanoids have been identified from a Cistus incanus subspecies traditionally used for treatment of skin diseases in northern parts of Greece and identified as subsp. tauricus. Flavan-3-ols are (+)-catechin, (+)-gallocatechin, the novel (+)-gallocatechin 3-gallate and the rarely occurring (+)-catechin 3-O-α-β-rhamnoside; proanthocyanidins are procyanidins B1 and B3, gallocatechin-(4α → 8)-gallocatechin, its novel (4α → 6)-regioisomer, gallocatechin-(4α → 8)-catechin, the tentatively identified novel catechin-(4α → 8)-gallocatechin and the trimer gallocatechin-(4α → 8)-gallocatechin-(4α → 8)-catechin. The uncommon flavanone 2R,3R-dihydromyricetin was also obtained.

Unambiguous assignments for free dimeric proanthocyanidin phenols from 2D NMR

Characterization of proanthocyanidin oligomers proceeds commonly through investigation of NMR data of their peracetates or methyl ether acetates, in conjunction with FAB-mass spectrometry and circular dichroism. Since such an approach is unsuitable in bioassay-guided isolations, we applied two-dimensional NMR techniques for the identification of dimeric proanthocyanidins. This afforded not only a powerful probe for distinction between the different procyanidin isomers, but also allowed full assignments, even for both major rotameric forms, whenever present, without the need for derivatisation. Moreover, discrimination between the crucial 6- and 8-protons and carbons was achieved after addition of traces of cadmium nitrate, resulting in the separation of the broad phenolic signals into sharp singlets. As an example of the general strategy followed in the assignment and combination of data of the different spectra available, complete analysis of underivatised procyanidin B3 or catechin-(4α→8)-catechin is discussed for the first time.

Antifungal effects of hydrolysable tannins and related compounds on dermatophytes, mould fungi and yeasts.

A series of hydrolysable tannins and related compounds was evaluated for antifungal activities against filamentous fungi ( Epidermophyton floccosum; Microsporum canis; Microsporum gypseum: Trichophyton mentagrophytes; Trichophyton rubrum; Trichophyton tonsurans; Trichophyton terrestre; Penicillium italicum; Aspergillus fumigatus; Mucor racemosus; Rhizopus nigricans) and opportunistic yeasts ( Candida albicans; Candida glabrata; Candidata krusei; Cryptococcus neoformans), using the agar dilution method. While all samples had no activity against the filamentous fungi in concentrations of 1.1 -5.9 μм (1000 [μg/ml), the phenolic compounds displayed significant potencies against all the opportunistic yeasts tested but C. albicans, with minimum inhibitory concentrations ranging from 0.02 to 0.1 μм ( 16 - 125 (μg/ml). Although the presence of galloyl groups in flavonoids did not necessarily produce activity, this structural element, an HHDP moiety or its oxidatively modified entity proved to be an important structural feature of hydrolysable tannins. Comparison of dilution methods provided strong evidence of dependence of MIC values on the test method. Employing the microdilution broth method, the ellagitannin corilagin (MIC 0.8 nм) was found to be similarly potentially active as amphotericin B (MIC 0.5 nм) and sertaconazole (MIC 0.9 nм) against Candida glabrata strains. The order of effectiveness observed being 64- and 4 -8-fold increased for corilagin and the reference compounds respectively, when compared with that of the agar dilution test.

A-type proanthocyanidins from Prunus spinosa

Abstract The range of natural dimeric A-type proanthocyanidins is extended by identification of ent-epicatechin-(4α → 8; 2α → O → 7)-catechin and ent-epiafzelechin-(4α → 8;2α → O → 7)-epicatechin. They are accompanied in the flowers of Prunus spinosa by the structurally related metabolites ent-epicatechin-(4α → 8;2α → O-7)- epicatechin, ent-epiafzelechin-(4α → 8;2α → O → 7)-catechin and ent-epiafzelechin-(4a@8;2a@0-7)-epiafzelechin, these being reported from a plant source for the second time. Nuclear Overhauser effect difference spectroscopy facilitated assessment of both 3,4-relative stereochemistry of the heterocyclic ring C and absolute configuration of terminal units, as well as determination of the mode of interflavanyl linkage. Whereas methylation and subsequent acetylation of A-types with ‘lower’ catechin units produce the expected methyl ether acetates, those species with a ‘lower’ epicatechin moiety indicate restrictedaccess of diazomethane to 5-OH(A).