Pavel Drašar

Steroid-porphyrin conjugate for saccharide sensing in protic media.

A new saccharide receptor in protic media has been designed and synthesized. The receptor combines advantages of steroids, which are responsible for saccharide binding, and of the porphyrin moiety acting as a signalling component of the molecule due to changes in UV-vis electronic spectra. The synthesis is based on condensation of steroid aldehyde with pyrrole to form the porphyrin unit with four protected steroid moieties. After deprotection, meso-substituted porphyrin contains 12-hydroxy groups on the steroidal part. The receptor is soluble in aqueous solutions and exhibits high complexation affinity towards saccharides. Because the receptor extensively aggregates in water, most of the experiments were performed in 50% aqueous 2-propanol where aggregation is significantly eliminated. Binding is evidenced by spectral changes in the Soret region of the receptor in UV-vis absorption spectra allowing the evaluation of the binding constants. Additional confirmation of binding is obtained using 1H NMR, Raman and IR spectroscopies and the surface plasmon resonance technique. The receptor exhibits higher selectivity for oligosaccharides over monosaccharide. The results point to the importance of a combination of multiple binding via H-bonding and hydrophobic interactions.

Metal coordination as a tool for controlling the self-assembling and gelation properties of novel type cholic amide–phenanthroline gelating agent

Abstract (3α,7α,12α)-Trihydroxy-N-[1,10-phenanthrolin-5-yl]-5β-cholan-24-amide was found to be a powerful gelating agent for methanol–water in gelator to solvent ratio starting from 0.1% in absence of metal ion. Formation of phenanthroline–zinc (II) 2:1 complex changes dramatically gelating properties; when stored, it dissolves into clear solution without Tyndall effect and this solution, when heated to ca. 70°C reversibly forms a gel again, without chemical change as proven by NMR spectrometry. Structures of the gels of cholic amide–phenanthroline and its Zn2+ complex were studied by SEM.

Investigation of new acyloxy derivatives of cholic acid and their esters as drug absorption modifiers

Skin penetration enhancers are used in the formulation of transdermal delivery systems for drugs that are otherwise not sufficiently skin-permeable. Intestinal absorption promoters/enhancers are used as excipients in oral formulations of poorly oral-bioavailable drugs. Series of fourteen acyloxy derivatives of 5β-cholic acid as potential drug absorption modifiers was generated by multistep synthesis. The synthesis of all newly prepared compounds is presented here. Structure confirmation of all generated compounds was accomplished by (1)H NMR, (13)C NMR, IR and MS spectroscopy methods. All the prepared compounds were analyzed using RP-TLC, and their lipophilicity (R(M)) was determined. The hydrophobicity (logP) and solubility (logS) of the studied compounds were also calculated using two commercially available programs. All the target compounds were tested for their in vitro transdermal penetration activity and as potential intestinal absorption enhancers. The anti-proliferative activity of all the final compounds was also assessed against the human cancer cell lines: T-lymphoblastic leukemia cell line and the breast adenocarcinoma cell line. Their cytotoxicity was also evaluated against the normal human skin fibroblast cells. Two compounds showed anti-proliferative effect on cancer cells without affecting the growth of normal cells, which should be promising in potential development of new drugs. Most of the target compounds showed minimal anti-proliferative activity (IC(50)>37 μM), indicating they would have low cytotoxicity when administered as chemical absorption modifiers. The relationships between the lipophilicity and the chemical structure of the studied compounds as well as the relationships between their chemical structure and enhancement effects are discussed in this article.

Novel Deep Cavity Calix[4]pyrroles Derived from Steroidal Ketones

Novel steroidal calix[4]pyrroles were prepared in excellent yields from commercially available cholic acid derivatives using an efficient synthetic sequence. Once in hand, it was found that these calix[4]pyrroles exist in the form of four different configurational isomers. Separation of these isomers was achieved readily using normal phase HPLC techniques. Once purified, the steroidal calix[4]pyrroles were screened via -FABMS analyses to judge their utility in effecting the enantioselective recognition of appropriate organic anions. Results that provided support for antipodal R > S selectivity were obtained in the case of both tartaric acid and mandelic acid. Direct extraction studies were then carried out and these confirmed the pattern of R > S selectivity observed by -FABMS.

Design and studies of novel polyoxysterol-based porphyrin conjugates

New types of steroid-porphyrin conjugates derived from 20-hydroxyecdysone (20E) and 24-epibrassinolide (EBl) were synthesized. An exceptional regioselectivity in the reaction of both steroids with porphyrin boronic acids was found to give side-chain-conjugated boronic esters as sole products. UV-Vis-, fluorescence and NMR spectroscopy yielded similar data for all the studied compounds confirming the solvent driven supramolecular assembly with formation of J-aggregates. CD measurements of water diluted solutions showed a clear difference between 20E and EBl conjugates. The latter showed a strong supramolecular chirality, whereas 20E J-aggregates did not.

Spectroscopic, Morphological, and Mechanistic Investigation of the Solvent‐Promoted Aggregation of Porphyrins Modified in meso‐Positions by Glucosylated Steroids

Solvent-driven aggregation of a series of porphyrin derivatives was studied by UV/Vis and circular dichroism spectroscopy. The porphyrins are characterised by the presence in the meso positions of steroidal moieties further conjugated with glucosyl groups. The presence of these groups makes the investigated macrocycles amphiphilic and soluble in aqueous solvent, namely, dimethyl acetamide/water. Aggregation of the macrocycles is triggered by a change in bulk solvent composition leading to formation of large architectures that express supramolecular chirality, steered by the presence of the stereogenic centres on the periphery of the macrocycles. The aggregation behaviour and chiroptical features of the aggregates are strongly dependent on the number of moieties decorating the periphery of the porphyrin framework. In particular, experimental evidence indicates that the structure of the steroid linker dictates the overall chirality of the supramolecular architectures. Moreover, the porphyrin concentration strongly affects the aggregation mechanism and the CD intensities of the spectra. Notably, AFM investigations reveal strong differences in aggregate morphology that are dependent on the nature of the appended functional groups, and closely in line with the changes in aggregation mechanism. The suprastructures formed at lower concentration show a network of long fibrous structures spanning over tens of micrometres, whereas the aggregates formed at higher concentration have smaller rod-shaped structures that can be recognised as the result of coalescence of smaller globular structures. The fully steroid substituted derivative forms globular structures over the whole concentration range explored. Finally, a rationale for the aggregation phenomena was given by semiempirical calculations at the PM6 level.

New propanoyloxy derivatives of 5β-cholan-24-oic acid as drug absorption modifiers.

A series of final twelve propanoyloxy derivatives of 5β-cholan-24-oic acid (O-propanoyl derivatives of cholic acid) as potential drug absorption modifiers (skin penetration enhancers, intestinal absorption promoters) was generated by multistep synthesis. Structure confirmation of all generated compounds was accomplished by 1H NMR, 13C NMR, IR and MS spectroscopy methods. All the prepared compounds were analyzed using RP-TLC, and their lipophilicity (RM) was determined. The hydrophobicity (log P), solubility (log S), polar surface area (PSA) and molar volume (MV) of the studied compounds were also calculated. All the target compounds were tested for their in vitro transdermal penetration effect and as potential intestinal absorption enhancers. The cytotoxicity of all the evaluated compounds was evaluated against normal human skin fibroblast cells. Their anti-proliferative activity was also assessed against human cancer cell lines: T-lymphoblastic leukemia cell line and breast adenocarcinoma cell line. One compound showed selective cytotoxicity against human skin fibroblast cells and another compound possessed the highest cytotoxicity against all the tested cell lines. Only one compound expressed anti-proliferative effect on leukemia cancer cells without affecting the growth of normal cells, which should be promising in potential development of new drugs. Most of the target compounds showed minimal anti-proliferative activity (IC50>37 μM), indicating they would have moderate cytotoxicity when administered as chemical absorption modifiers. The relationships between the lipophilicity/polarity and the chemical structure of the studied compounds as well as the relationships between their chemical structure and enhancement effect are discussed in this article.

Synthesis of the sulfates derived from 5α-cholestane-3β, 6α-diol

Abstract Three new steroid sulfates—3β-hydroxy-5α-cholestan-6α-yl sulfate, 6α-hydroxy-5α-cholestan-3β-yl sulfate, and 5α-cholestan-3β, 6α-diyl disulfate—were synthesized. For the syntheses of the key intermediates, 3β-hydroxy-5α-cholestan-6α-yl acetate and 6α-hydroxy-5α-cholestan-3β-yl acetate, selective protection of hydroxy groups in 5α-cholestane-3β, 6α-diol was necessary. This problem was solved by using a combination of acetyl, tetrahydropyranyl, and methoxy methyl protective groups, which represents a new approach leading to these hydroxy acetates. Sulfated derivatives of 5α-cholestane-3β, 6α-diol are present in marine invertebrates and were synthesized for the purposes of biologic testing.

New polyfluorothiopropanoyloxy derivatives of 5β-cholan-24-oic acid designed as drug absorption modifiers

A series of final six propanoyloxy derivatives of 5β-cholan-24-oic acid (tridecafluoroctylsulfanyl- and tridecafluoroctylsulfinylethoxycarbonylpropanoyloxy derivatives) as potential drug absorption promoters (skin penetration enhancers, intestinal absorption promoters) was generated by multistep synthesis. Structure confirmation of all generated compounds was accomplished by (1)H NMR, (13)C NMR, IR and MS spectroscopy methods. All the prepared compounds were analyzed using RP-TLC, and their lipophilicity (RM) was determined. The hydrophobicity (log P), solubility (logS), polar surface area (PSA) and molar volume (MV) of the studied compounds were also calculated. All the target compounds were tested for their in vitro transdermal penetration effect and as potential intestinal absorption enhancers. The cytotoxicity of all the evaluated compounds was evaluated against normal human skin fibroblast cells. Their anti-proliferative activity was also assessed against human cancer cell lines: T-lymphoblastic leukaemia cell line and breast adenocarcinoma cell line. One compound showed high selective cytotoxicity against human skin fibroblast cells and another compound possessed high cytotoxicity against breast adenocarcinoma cell line and skin fibroblast cells. Only one compound expressed anti-proliferative effect on leukaemia and breast adenocarcinoma cells without affecting the growth of normal cells, which should be promising in potential development of new drugs. Most of the target compounds showed minimal anti-proliferative activity (IC50>37μM), indicating they would have moderate cytotoxicity when administered as chemical absorption modifiers. The relationships between the lipophilicity/polarity and the chemical structure of the studied compounds as well as the relationships between their chemical structure and penetration enhancement effect are discussed in this article.

Succinobucol's New Coat - Conjugation with Steroids to Alter Its Drug Effect and Bioavailability

Synthesis, detailed structural characterization (X-ray, NMR, MS, IR, elemental analysis), and studies of toxicity, antioxidant activity and bioavailability of unique potent anti-atherosclerotic succinobucol-steroid conjugates are reported. The conjugates consist of, on one side, the therapeutically important drug succinobucol (4-{2,6-di-tert-butyl-4-[(1-{3-tert-butyl-4-hydroxy-5-(propan-2-yl)phenyl]sulfanyl}ethyl)sulfanyl]phenoxy}-4-oxo-butanoic acid]) possessing an antioxidant and anti-inflammatory activity, and on the other side, plant stanol/sterols (stigmastanol, β-sitosterol and stigmasterol) possessing an ability to lower the blood cholesterol level. A cholesterol-succinobucol prodrug was also prepared in order to enhance the absorption of succinobucol through the intestinal membrane into the organism and to target the drug into the place of lipid metabolism—The enterohepatic circulation system. Their low toxicity towards mice fibroblasts at maximal concentrations, their antioxidant activity, comparable or even higher than that of ascorbic acid as determined by direct quenching of the DPPH radical, and their potential for significantly altering total and LDL cholesterol levels, suggest that these conjugates merit further studies in the treatment of cardiovascular or other related diseases. A brief discussion of succinobucol’s ability to quench the radicals, supported with a computational model of the electrostatic potential mapped on the electron density surface of the drug, is also presented.