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Item Open Access Conformational analysis of oligomeric profisetinidins(University of the Free State, 2007-02) Potgieter, Eleonora Deborah; Ferreira, D.; Brandt, E. V.English: The profisetinidins are an important class of condensed tannins, or proanthocyanidins. Historically, studies towards the structure and conformation of proanthocyanidins were done on their peracetate and permethyl acetate derivatives. A current upsurge in industrial and biological applications of proanthocyanidins has prompted the present efforts at detailed analysis of the conformational behaviour of the naturally occurring free phenolic oligomeric profisetinidins. Studies towards the structure and conformational analysis of a small number of free phenolic dimeric procyanidins that are 4→8 coupled and only one free phenolic dimeric profisetinidin, fisetinidol-(4α→8)-catechin, have hitherto been reported. This study centres on the use of 1H, 13C, gradient COSY, COSY 45, COSY 90W, NOESY PH and HMQC NMR experiments in different solvents and at different temperatures to assign the hydrogen and some carbon resonances of the free phenolic profisetinidins that are found in commercially important southern hemisphere trees, namely Black Wattle (Acacia mearnsii) and Quebracho (Schinopsis balansae). These results, together with data obtained from CD spectra in methanol, were then used to study the conformations of these compounds. Dimers with 2,3-trans-3,4-trans (2,4-cis) configuration, namely fisetinidol-(4α→8)-catechin, fisetinidol-(4α→6)-catechin, ent-fisetinidol-(4β→8)-catechin and ent-fisetinidol-(4β→6)- catechin all displayed sets of duplicate resonances on 1H NMR spectra, indicating the presence of rotamers on an NMR time-scale at ambient temperatures. The proton resonances of the rotamers of the 4→6 linked dimers displayed insignificant chemical shift differences due to the similar magnetic environments and linear shape of both rotamers. The proton resonances of the rotamers of the 4→8 linked dimers displayed significant chemical shift differences due to the presence of compact and extended rotamers resulting in large changes in magnetic environment due to anisotropic effects. The type of solvent, temperature as well as the relative presence of water or cadmium nitrate had a strong influence on the relative concentrations of the rotamers, the conformations of the heterocyclic C- and F-rings as well as the visibility of hydroxy groups. The F-rings all displayed A/E- conformational exchange with line shapes indicating possible skewed boat conformations in some instances. The C-ring conformations ranged from rings with A/E conformational exchange to preferred E-conformers. The dimers with 2,3-trans-3,4-cis (2,4-trans) configuration, namely fisetinidol-(4β→8)- catechin, fisetinidol-(4β→6)-catechin and ent-fisetinidol-(4α→8)-catechin displayed only one set of resonances on 1H NMR spectra at ambient temperatures. The presence of intramolecular hydrogen bonding and limited conformational exchange was confirmed by the following observations: a) Selective broadening of proton resonances in both the heterocyclic and aromatic regions of 1D NMR spectra. b) Sharpening of resonances in 1D spectra at elevated temperatures. c) The presence of abundant coupling between heterocyclic and aromatic ring protons as observed on 2D spectra. d) Coupling between 2-HC and 4-HC on 2D NMR spectra. The C-rings had preferred A-conformations, with the F-rings displaying A/E conformational exchange with line shapes indicating possible skewed boat conformations in some instances. 2D NMR experiments afforded estimations, in some cases, of the angles between the plane of the B-ring and the 2-CC→2-HC bond, the plane of the D-ring and the 4-CF→4-HF bond as well as the plane of the D-ring and the 4-CC→4-HC. The resonances of 4-CC of the 2,3-trans-3,4-trans dimers displayed significant chemical shift differences (± 41 ppm) compared to 4-CC of the 2,3-trans-3,4-cis dimers (± 31 ppm).This could serve as a possible indicator of the relative configurations of the C-rings of 2,3-trans profisetinidins dimers. CD studies of al seven abovementioned dimers, as well as four trimers from Acacia mearnsii displayed complex curves with a number of strong Cotton effects. Although some trends were observed, it was abundantly evident that the chiroptical characteristics of this class of compounds are too complex to be interpreted in terms of the empirical quadrant rule.Item Open Access The constitution of oligomeric benzofuranoids(University of the Free State, 1999-05) Bekker, Riaan; Brandt, E. V.; Ferreira, D.English: Berchemia zeyheri is known for its unique red heartwood, a property that was probably responsible for the first phytochemical investigation into the flavonoid content of this tree. The heartwood contains a unique series of biflavonoids with one of more benzofuranoid moieties. These are usually found in diastereomeric mixtures, the biogenetic origin and stereochemistry of which have hitherto been unknown. This investigation thus represents a renewed effort to solve some of the intricate problems associated with these compounds. The high concentration of maesopsin in the heartwood made extensive enrichment and fractionation by the use of Craig countercurrent distribution techniques and Sephadex LH- 20 gelchromatography necessary. The two diastereomers of 4',5, 7-tri-O-methylnaringenin-(3a~ 7)-2,4,4',6-tetra-Omethylmaesopsin were, for the first time, successfully isolated and separated. Reduction of these diastereomers with Na(CN)BH3 gave two enantiomeric pure fragments. The conformations of the heterocyclic rings of these fragments were established by molecular mechanics (MMX and GMMX) and semi-empirical methods (AMI). These results allowed the absolute configuration of the fragments to be deduced from CD-curves of the compounds by application of Snatzke's rule for a,~-unsaturated five-membered cyclic rings. A n.O.e. correlation observed for one of the diastereomers only, correlates the stereocenter of the maesopsin moiety, of known absolute configuration, with a specific configuration of the naringenin unit, thus defining the absolute configuration of the dimer. These results also allowed the determination of the absolute stereochemistry of two regioisomers of the above dimers, 4',5, 7-tri-O-methylnaringenin-(3a~5)-2,4,4',6-tetra-Omethylmaesopsin and its epimer. The 13CNMR spectra of these related dimers were also studied and fully elucidated by means of HMQC and HMBC experiments. The structure and stereochemistry of two novel isoflavanone-benzofuranone biflavonoids, 4',5,7-tri-Omethyldihydrogenistein-fêcc-» 7)-2,4,4',6-tetra-O-methylmaesopsin and its epirner, were similarly determined. Resolution of maesopsin, the main metabolite in the heartwood, by means of HPLC using a chiral column, for the first time gave access to the two enantiomers of this benzofuranoid. 4,4',6- Tri-O-methyl-2-deoxymaesopsin-(2~ 7)-2,4,4',6-tetra-O-methyl-maesopsin and its epimer, consist of two benzofuranoid constituent units. An X-ray crystal structure was obtained for the one diastereomer, but due to the presence of a symmetric Pbea point group, only the relative configuration could be determined. After the racemic nature of each of the diastereomers was determined, each epimer was resolved with HPLC into its constituent enantiomers. The information obtained from the CD curves and crystal structure allowed the determination of the absolute stereochemistry of each of the enantiomers. Four further epimeric biflavonoids were isolated as the hepta-O-methyl ethers. Evidence obtained from 13C NMR data suggested the presence of a y-lactone functionality in the upper benzofuranoid moiety, identifying the dimers as the epimers of 4,6-dimethoxy-3-( 4- methoxy-benzyl)benzo[b ]furan-2(3H)-one-(2~5)-2,4,4',6-tetra-O-methylmaesopsin and the (2~ 7)-coupled regio-isomer. In order to supplement the above data, an asymmetric synthesis of maesopsin was attempted. The first attempt involved the oxidation of 2-( 4-methoxybenzyl)-4,6- dimethoxybenzo[b ]furan-3(2H)-one, obtained by reduction of the corresponding aurone, with AD-mix-a, a stereoselective catalyst, or chiral oxaziridine. The former afforded the desired product in low yield but no stereo selectivity while the latter method realized a much-improved yield, but still with no selectivity. This lack of selectivity is attributed to equilibrium of the product with the a-diketone. Attempts to prevent the formation of this equilibrium product were unsuccessful. A second synthetic attempt involved benzylation of 2,4,6-trimethoxybenzo[b]furan-3-(2H)-one with (-)-sparteine as chiral auxiliary, but again resulted in high yields but no stereo selectivity.Item Open Access Die eerste oligomeriese neoflavonoiede: struktuur en sintese van modelverbindings(University of the Free State, 1993-05) Nel, Janetta Wilhelmina; Bezuidenhoudt, B. C. B.; Brandt, E. V.; Ferreira, D.Afrikaans: Hoewel oligomeriese flavonoïede reeds etlike dekades bekend is, is die eerste bi-isoflavonoïede eers onlangs uit Dalbergia spesies geïsoleer en is oligomere saamgestel uit neoflavonoïede nog onbekend. Ten einde die poel van oligomeriese isoflavonoïede uit te brei is die fitochemiese ondersoek na die fenoliese komponente in die kernhout van Dalbergia nitidula Welw. ex Bak, bekend vir die voorkoms van talle monomeriese iso- en neoflavonoïede, onderneem. Drie nuwe unieke iso-neoflavonoïed dimeriese verbindings bestaande uit 'n seldsame 3-fenielbenso[b]furaan- en vir die eerste keer 'n pterokarpaaneenheid, nl. (6aS,11aS)-2-, (6aS,11aS)-4- en (6aS,11aS)-8-[3-feniel-5-hidroksi-6-metoksibenso-[b]furan-2-ielmetiel]medikarpin, is inderdaad tydens die ondersoek gevind. Hierdie verbindings word in die eterekstrak vergesel deur 'n verdere dimeer waarin dieselfde neoflavonoïedeenheid aan (+ )-vestitol gebind is, nl. (3S)-8-[3-feniel-5-hidroksi-6-metoksi benso[b]furan-2-ielmetiel] vestitol. Benewens die dimere is 2', 7-dihidroksi-4'-metoksi-isoflav-3-een, die vyfde natuurlike isoflaveen, en (6aS,11aS)-2-hidroksi-3,9-dimetoksipterokarpaan as nuwe mono-mere verkry, terwyl 3,8-dihidroksi-9-metoksipterokarpaan vir die eerste keer as die 6aS,11aS isomeer geïsoleer is. Die bekende benso[b]furaan, centrolobofuraan, is ook saam met bekende verbindings soos (±)-ferreirien, di-O-metieldaidzein, (+ )-claussekinoon, 3-hidroksi-9-metoksikumestaan en (+ )-vestikarpin vir die eerste keer in hierdie spesie gevind, terwyl (+ )-medikarpin, (+ )-homopterokarpin, (+ )-vestitol, 3,9-dimetoksikumestaan asook (±)-liquiritigenien en isoliquiritigenien weereens verkry is. Aangesien onsekerheid omtrent die heterosikliese sisteem van die neoflavonoïedeenheid in die dimere bestaan het, is die sintese van 'n neoflavonoieddimeer met 'n seslid heterosikliese ring, (6 aS,11as)-4-[6-hidroksi-7 -metoksineoflav-3-een-3-iel] medikarpin d.m.v. 'n aldol-tipe kondensasie tussen 'n gesubstitueerde bensofenoon en pterokarpanielasetaldehied gevolg deur 'n sikliseringstap, aanvanklik aangepak (Skema). Weens die seldsame substitusiepatroon van die bensofenoon, is 2-hidroksi--4-metoksibensofenoon, as model, d.m.v. 'n Grignardreaksie tussen 4-metoksi-2-O-metoksimetielbensaldehied en fenielmagnesiumbromied daargestel en die pterokarpanielasetaldehied vanaf (+ )-medikarpin deur O-allilering (K2C03/ asetoon/ allielbromied) gevolg deur Claisenherrangskikking (N,N-dimetielanilien) en oksidatiewe splyting (OsO4/N-metielmorfolien-N-oksied gevolg deur NaIO4/MeOH) van die allielgroep, in 14% opbrengs verkry. Intermolekulêre aldolkondensasie (LDA/THF of asynsuur /H2S04) tussen die ptero-karpanielasetaldehied en die bensofenoon kon egter nie bewerkstellig word nie. Ten einde 'n Von Pechman-tipe kondensasie vir die sintese te kan benut moes die pterokarpanielasetaldehied na die ooreenstemmende suur geoksideer word, maar ten spyte van die aanwending van 'n verskeidenheid reagense (piridiniumdichromaat; RuCl3; Tollensreagens; KMn04; Ag20) kon 2-O-etoksimetiel-4,6-dimetoksifenielasetaldehied, as model, nie in noemenswaardige opbrengs na die ooreenstemmende suur geoksideer word nie. A.g.v. die onvermoë om die verlangde pterokarpanielasynsuur daar te stel, is die sintese van 'n model van die neoflavonoïedeenheid, 3-feniel-7-metoksineoflav-3-een, aangepak en is verestering van 2-hidroksi-4-metoksibensofenoon met fenielasetielchloried (K2C03/ asetoon) gevolg deur intramolekulêre siklisering (K2C03/ asetoon) en reduksie van die laktoonfunksionaliteit (B2H6THF) uitgevoer (10% opbrengs). Vergelyking van die chemiese verskuiwing van die CH2Sein in die 1H-KMR-(8 5.09) sowel as 13C-KMR-spektra (8 69.8) met dié van die natuurprodukte (8 4.13 - 4.24 en 8 21.0 onderskeidelik) het egter ondubbelsinnig getoon dat die natuurlike verbindings nie 'n seslid-heterosikliese ring in die neoflavonoïedeenheid bevat nie. Ten einde soortgelyke KMR-vergelyking vir die struktuur met die benso[b]furanielneoflavonoïed te doen, is voortgegaan met 'n modelsintese vir hierdie sisteem en is die verlangde 2-bensiel-3-fenielbenso[b]furaan d.m.v. Friedel-Grafts asilering van resorsinol met chloroasetielchloried, gevolg deur bensilering (bensielbromied, K2C03/asetoon) en Grignardreaksie van die gevormde bensofuranoon met fenielmagnesiumbromied daargestel. Hoewel presiese ooreenstemming in chemiese verskuiwing (8 4.17 en 8 32.8 onderskeidelik in die 1H- en 13C-KMR-spektra) van die metileengroep tussen die gesintetiseerde model en die natuurproduk nie verkry is nie, is dit aan verskille in struktuur en substitusiepatroon toegeskryf en kan 'n benso[b]furaanstruktuur aan die neoflavonoïedeenheid toegeken word.Item Open Access Polyphenols from pericopsis elata and synthesis of selected stilbenes(University of the Free State, 2005-05) Litedu, Eunice Maria; Kamara, B. I.; Brandt, E. V.English: The bark of Pericopsis elata (Afrormosia elata) is used by the local population of the Democratic Republic of Congo for the treatment of cancer and for external applications because of its weather resistance. Previous phytochemical studies on Pericopsis elata resulted in the isolation of flavonoids, isoflavonoids, chalcones and stilbene monomers. Among these, stilbenes are known to have biological activities such as antioxidants, antifungal, and act as microbial inhibitors. They posses COX-1 and COX-2 inhibitory effects, affect lipid peroxidation, LDL oxidation, and function as phytoalexins among other activities. Prompted by these claims, we conducted an in depth investigation of the heartwood of P. elata by extraction, isolation and structural elucidation of the metabolites. The enrichment and fractionation of monomeric and dimeric constituents were accomplished mainly by Craig countercurrent distribution techniques and Sephadex LH-20 gel chromatography. Pure compounds were obtained by derivatization and preparative thin layer chromatography. Structural elucidation of the phenolics is based mainly on NMR spectroscopic methods (1H NMR, 13C NMR, COSY, NOESY, HMBC and HMQC), Mass spectrometry and synthetic methods.The monomeric compounds isolated during this study comprise the flavanones (naringenin and eriodictyol), the isoflavones (genistein and biochanin A), a dihydrochalcone [(R)-α,4,2',4'-tetraacetoxydihydrochalcone] and a single α- methyldeoxybenzoin (angolensin). Among the stilbene monomers encountered are resveratrol, isorhapontigenin and large amounts of piceatannol. Structures of the dimeric stilbnes isolated apparently originate from different combinations of piceatannol (3,4,3',5'-tetrahydroxystilbene) (the major constituent ~ 16% of the phenolic content of P. elata) and 3,4,3',4'-tetrahydroxystilbene. To the best of our knowledge rel-2,3-trans-2-(3,4-dimethoxyphenyl)-3-(3,5-dimethoxyphenyl)-6-[2-(3,5- dimethoxyphenyl)-E-1-ethenyl]benzodioxane, rel-2,3-trans-2-(3,4-dimethoxyphenyl)-3- (3,4-dimethoxyphenyl)-4-[2-(3,4- dimethoxyphenyl) )-E-1-ethenyl]-6-methoxy-2,3- dihydrobenzofuran, rel-2,3-trans-2-(3,4-diacetoxyphenyl)-3-(3,5-diacetoxyphenyl)-4-[2- (3,4-diacetoxyphenyl) )-E-1-ethenyl]-6-acetoxy-2,3-dihydrobenzofuran, rel-2,3-trans-2- (3,4-dimethoxyphenyl)-3-(3,5-dimethoxyphenyl)-4-[2-(3,5-dimethoxyphenyl) )-Z-1- ethenyl]-6-methoxy-2,3-dihydrobenzofuran, rel-2,3-trans-4-Formyl-2-(3,4- dimethoxyphenyl)-3-(3,5-dimethoxyphenyl)- 6-methoxy-2,3-dihydrobenzofuran and rel- 2,3-trans-2-(3,5-diacetoxyphenyl)-3-(3,5-diacetoxyphenyl)-6-[2-(3,5-diacetoxyphenyl) )- E-1-ethenyl]-4-acetoxy-2,3-dihydrobenzofuran are novel compounds. Structural confirmation of the six novel dimeric stilbenes in particular required definition of structure via synthetic methods. A synthetic approach was thus developed. This protocol comprises the synthesis of piceatannol via the Wittig reaction followed by oxidative coupling of two piceatannols to afford novel dimeric stilbenes. 1H NMR of the methoxy-derivative of synthetic compounds is identical to that of the derivative of two dimeric stilbenes isolated from P. elata.Item Open Access Sintese en konformasie van natuurlike flobatanniene: die meganisme van basisgekataliseerde piraanherrangskikkings(University of the Free State, 1988-12) Steynberg, Jan Petrus; Ferreira, D.; Brandt, E. V.; Young, D. A.Afrikaans: Guibourtia coleosperma (Rhodesian copalwood) is bekend as 'n harde en duursame houtsoort wat opvallend weerstand teen verrotting bied. Die onlangse isolasie van unieke bi- en triflavanoïede gebaseer op leukoguibourtinidiene (4',7-dihidroksiflavan-3,4-diole) en fenoliese stilbene, bekend vir antifungiese eienskappe, het as stimulus gedien vir 'n herondersoek van die fenoliese metaboliete. Uitgebreide verrykings- en skeidingstegnieke lewer, benewens bekende (E)-stilbeen monosiede, die unieke 4',5-dimetoksi- en 3',5-dihidroksi-4'-metoksi-3-[O-α-L-rhamnopiranosiel-(1 - 6)-β-D-glukopiranosieloksil stilbeenbiosiede. Die lae konsentrasie van hierdie verbindings het 'n nuwe benadering ten opsigte van identifikasie genoodsaak. Ontwikkeling van 'n elegante metode waar die aard en posisie van interglikosidiese-sowel as glikosied-aglikoon bindings met behulp van n.O.e. tegnieke ('H k.m.r.) gekarakteriseer word, voorsien 'n waardevolle bydrae tot identifisering van glikosiede. Hierdie metode het veral meriete waar alternatiewe spektroskopiese en chemiese metodes deur die beskikbaarheid van voldoende materiaal aan bande gelê word. Die stilbeenglikosiede word in G. coleosperma vergesel deur flobatanniene, 'n unieke klas van ring-geïsomeriseerde tanniene. Hierdie verbindings verteenwoordig die produkte van stereospesifieke ring-isomerisasie van 2,3-trans-3,4-trans- en stereoselektiewe isomerisasie van 2,3-trans-3,4-cis-(-)-fisetinidol eenhede in konvensionele (4,8)-bi- en (4,6:4,8)-triflavanoïede. Fisiese data soos massaspektroskopie, sirkulêre dichroisme maar veral hoë resolusie 1H k.m.r. n.O.e. tegnieke, lei, op enkele uitsonderings na, tot volledige struktuuropklaring van die gefunksionaliseerde tetrahidro-2H,8H-pirano [2,3-h]-, [2,3-f]- en [2,3-g] asook heksahidro-2H,6H,10H-dipirano[2,3-f:2',3'-h] chromene. Eenduidige struktuurbewys vir die nuwe klas van gekondenseerde tanniene volg uit piraanherrangskikkings van die waarskynlike biflavanoiedvoorlopers onder milde basiese kondisies (pH1O) . Die vatbaarheid van biflavanoiede onder basiese kondisies vir epimerisasie en herrangskikking via kinoonmetiede kompliseer egter die reaksieverloop. Om dus newereaksies geassosieer met 'n E-ring kinoonmetied uit te skakel, is die 4-0-metieleterderivate (E-ring) van die biflavanoiede aan die basiese reaksiekondisies blootgestel. Terwyl biflavanoiede met 2,3-trans-3,4-trans-flavan-3-ol "boonste" eenhede vatbaar is vir relatief stadige maar stereospesifieke C-ring isomerisasie, ondergaan die analoë 2,3-trans- 3,4-cis isomere stereoselektiewe isomerisasies teen 'n verhoogde tempo. Laasgenoemde analoë is verder vatbaar vir herrangskikkings na tetrahidropiranochromene waar die pirokatekol B- en resorsinol A-ringe omgeruil is relatief tot die posisies daarvan in die "normale" isomere. Hierdie ringomruilings geskied via 1,3-flavanielmigrasie in intermediêre kinoonmetiede en gaan gepaard met die inversie van die absolute konfigurasie by die ekwivalent van 3-C (C-ring) in die biflavanoied voorloper. Opvallend is dat sommige van hierdie "ringomgeruilde " analoë ook in Baikiaea plurijuga aangetref is wat vermoedelik dui op die funksionering van 'n meganisme in die natuur soortgelyk as dié voorgestel in hierdie ondersoek. Benewens die voorsiening van eenduidige struktuurbewys vir die volledige reeks natuurlike flobatanniene, is 'n aanvaarbare meganistiese verklaring vir die verloop van die basisgekataliseerde piraanherrangskikkings ontwikkel. Die benadering van selektiewe beskerming van 4-OH(E), het verder afdoende bewys verskaf dat die waargenome flavanielmigrasies by die "onbeskermde" analoë via 'n E-ring kinoonmetied verloop. In teenstelling met "beskermde" biflavanoïede waar slegs die C-ring aan isomerisasie onderworpe is, kan beide die C- en I-heterosikliese ringe van 'n triflavanoïed piraanherrangskikking ondergaan. Behandeling van die (4α,8:4α,6)-bis-(-)-fisetinidol-(+)-katesjien as prototipe vir trimeriese profisetinidiene met basis het inderdaad die heksahidro-2H,6H,lOH-dipirano[2,3-f:2' ,3'-h] chromeen wat dui op 'n dubbele ringisomerisasie asook isomerisasie-intermediêre 6-flavaniel-[2,3-h]- en 1O-flavaniel-[2,3-f] chromene waar slegs een heterosikliese ring isomerisasie ondergaan het, gelewer. Die 10-flavaniel-[2,3-f] chromeen is ook uit G. coleosperma geïsoleer terwyl die dipirano[2,3-f:2' ,3'-h'] chromeen uit Colophospernum mopane geïsoleer is. Ten spyte van intensiewe ondersoeke na kommersiële basisgekataliseerde reaksies van prosianidiene met die oog op benutting van kommersiële ekstraksie van "loblolly pine" bas onder basiese kondisies, bestaan geen getuienis van C-ring isomerisasies by hierdie kommersieel-belangrike klas verbindings nie. Behandeling van prosianidien B-3, 'n tipiese prosianidien biflavanoïed, met milde basis lei tot 'n tetrahidropirano[2,3-h] chromeen met "omgeruilde" floroglusinol A- en pirokatekol B-ringe asook analoë 4-ariel-2- flavaniel bensopirane d.w.s. analoë waarin die 4-C (+)-katesjienen die 2-C pirokatekoleenheid "uitruil". Terwyl die tetrahidropiranochromeen vorm deur 1,3-flavanielmigrasie in 'n intermediêre kinoonmetied, ontstaan die 4-ariel-2-flavanielbensopirane deur 1,3-migrasie van 'n "vrygestelde" floroglusinoleenheid in dieselfde kinoonmetied intermediêr. Onder hierdie milde basiese kondisies kon geen bewyse vir omskakeling na "catechinic-acid" tipe produkte verkry word nie. Hierdie verbindings is bekend vir buitengewone suurheid en lae reaktiwiteit teenoor aldehiede en het die suksesvolle toepassing van alkaliese ekstraksie van natuurlike prosianidiene tot dusver gekortwiek. Die huidige ondersoek bewys dus dat met die versigtige seleksie van kondisies hierdie probleme grootliks oorbrug kan word. Die afwykende 1H k.m.r. en SD data van tetrahidropiranochromene met trans-trans C-ring konfigurasie en die omgekeerde Cotton-effek van [2,3-h] chromene met 2,3-trans F-ring konfigurasie, het die toekenning van absolute konfigurasie by sekere verbindings gekompliseer. So toon die SD spektra van die produkte met "omgeruilde" resorsinol A- en pirokatekol B-ringe 'n sterk negatiewe Cotton-effek in teenstelling met 'n verwagte positiewe Cotton-effek deur toepassing van die aromatiese kwadrantreël. Die teenstrydige chiroptiese gedrag word aan die konformasionele mobiliteit van die F-ring toegeskryf en is in 'n poging om die probleem te oorbrug die volgende benaderings benut. Die geredelike-beskikbare 4-arielflavan-3-ole is in modelreaksies gebruik om ondubbelsinnig te bewys dat ringomruiling tydens die isomerisasiereaksie met inversie van die absolute konfigurasie by die chirale sentra van die C-ring gepaard gaan. Laastens is die konformasionele gedrag van die flobatanniene teoreties met rekenaargesimuleerde molekulêre meganika bestudeer en kon deur die SD data met teoreties voorspelde voorkeurkonformasies in verband te bring die afwykende chiroptiese gedrag van sekere flobatanniene sinvol verklaar word. Uit Dreidingmodelle blyk dit duidelik dat 'n dominante A-konformasie, soos deur MM-berekenings bevestig, by tetrahidropiranochromene met alles-trans C-ring konfigurasie tot 'n "omgekeerde" Cotton-effek sal lei. Die konformasionele mobiliteit van die F-ring by tetrahidropiranochromene met 2,3-trans F-ring konfigurasie word inderdaad deur MM-berekenings bevestig (E:A konformeerverhouding, + 50:50). Uit die oogpunt van die kommersiële benutting van gekondenseerde tanniene is bogenoemde C-ring isomerisasie met gepaardgaande "vrystelling" van 'n resorsinoleenheid van fundamentele belang. Die vrygestelde resorsinolring behoort tot die aktivering van die basiese tannienskelet, veral vanaf die triflavanoiedvlak, vir reaksie met bv. formaldehied te lei om aldus aanleiding te gee tot beter benutting van tanniene in die koudsettende kleefstofbedryf.Item Open Access Sintese en struktuur van Pirano-isoflavanoïede(University of the Free State, 1977-01) Van Heerden, Fanie Retief; Roux, D. G.; Brandt, E. V.Afrikaans: Die medisinale gebruik (waarskynlik as antiseptikum) van die bas van Dalbergia nitidula Welw. ex Bak. (Leguminosae) deur inboorlinge het tot die huidige chemiese ondersoek aanleiding gegee. Hierdie ondersoek behels dus die isolering en struktuuropklaring van vier nuwe, natuurlike isoflavane, naamlik leiocin, nitidulin, nitidulan en heminitidulan, asook drie nuwe pterokarpaan-analoë waaraan die benamings nitiducarpin, hemileiocarpin en nitiducol toegeken is. ‘n Verdere nuwe isoflavaan, leiocinol se struktuur kon slegs gedeeltelik opgeklaar word, terwyl die isolasie van leiocarpin, 'n bekende pterokarpaan, 'n gerieflike verwysingselement in die stereochemiese opklaring van die pterokarpane daargestel het. Ter bevestiging van die strukture van die isoflavane, is 'n totaalsintese van 2'-0--metielleiocin uitgevoer. Die stereochem ie by C-3 van die isofLavane is met behulp van SD-kurwes as 3S bepaal, terwyl dié van die pterokarpane 6aS, llaS blyk te wees. Stereochemiese ekwivalensie by 'n enkele chiralsentrum, naamlik C-3 I van isoflavane en die ooreenstemmende C-6a van pterokarpane, dui op 'n gemeenskaplike biogenetiese oorsprong van hierdie twee tipes verbindinge. Substitusiepatrone van die metaboliete toon 'n noue verwantskap, in ooreenstemming met biogenetiese oorwegings. Die verbindings, met uitsondering van nitiducol, bevat deurgaans 'n addisionele 2H-piraanring aan die A-ring van die isoflavanoïedskelet. In verskeie gevalle is die. 2H-piraanring konvensioneel 2,2-dimetiel-gesubstitueerd (leiocin, leiocinol, hemileiocarpin), terwyl dit in ander gevalle 'n interessante en seldsame 2-metiel-2-(4-metiel-3-penteniel) substitusiepatroon besit (nitidulin, nitidulan, heminitidulan, nitiducarpin). Nitiducol, 'n pterokarpaan, bevat 'n geranielsyketting en orto-hidroksigroep as substituente, wat in die gesikliseerde isomeer deur die 2-metiel-2-(4-metiel-3-penteniel)-2H-piraanring (nitiducarpin) verteenwoordig word. Die voorlopige struktuur van die isoflavaan, leioeinol, is onderhewig aan verdere bevestiging deur totaalsintese. Ter ondersteuning van die strukture van ‘n aantal verwante isoflavane is 'n totaalsintese van 2'-O-metielleiocin uitgevoer. Aanvanklik is gepoog om leiocin via die ooreenstemmende chalkoon en isoflavoon te berei, maar die sintese kon vanweë swak isoflavoonvorming in die teenwoordigheid van die vry hidroksifunksie nie voltooi word nie. 2'-0-Metielleiocin is egter suksesvol gesintetiseer en die produk is identies aan die 2'-0- metieleter van die ooreenstemmende natuurproduk bevind. 'n Totaalsintese van die oorblywende metaboliete, sowel as 'n ondersoek na die anti-patogene en medisinale eienskappe van die verbindings word in die vooruitsig gestel.Item Open Access Structure and synthesis of bioactive polyphenols from Cyclopia subternata (Honeybush) and Aspalathus linearis (Rooibos) tea: conformational analysis of selected chiral derivatives(University of the Free State, 2009-10-23) Brand, Dirk Jacobus; Brandt, E. V.; Bezuidenhout, B. C. B.; Steenkamp, J. A.English: Endorsed by results from our initial investigations of the unique South African tea, brewed from Cyclopia intermedia (Honeybush), demonstrating the presence of polyphenolic flavonoids and glycosides, the tea is gaining popularity as a health beverage. The presence of these compounds which are claimed to have beneficial pharmacological properties, supported by the fact that the tea contains no caffeine and has low tannin content, as well as its usage as a medicinal plant by the indigenous people of the Western and Eastern Cape, prompted continued investigations also into the metabolic content of Cyclopia subternata. Selected extracts of the unfermented shoots and stems of C. subternata were subjected to chromatographic separations that afforded a novel flavan, flavonols, flavanones, flavones, isoflavone and C6.C2- and C6.C1-type compounds. The content of the acetone extract has previously been reported as well as the exploration of various synthetic routes in an attempt to confirm the structure of the novel flavan O- glucoside isolated from the tea. A continuation of the investigation of fractions from the methanol extract of the plant yielded a few compounds that were not found in the acetone extract. This work has been summarized in Chapter one and published as the phenolic profile of Cyclopia subternata. The unambiguous confirmation of the structure of the flavan O-glucoside and essentially the novel aglycone isolated from the tea, necessitated the use of synthetic, Nuclear Magnetic Resonance spectrometric (1H NMR), mass spectroscopic and circular dichroism methods. Mass spectrometry (‘soft’ ionization) was utilized to produce signature fragment ions of the synthesized flavan and the aglycone of the natural flavan for direct comparison. Hydroxylation patterns of the synthetic flavan and the natural flavan aglycone, as well as the relative configuration and point of attachment of the sugar moiety was deduced from NMR experiments. The absolute configuration of the aglycone was determined by circular dichroism. The total synthesis of Aspalathin, a potent antioxidant found in the South African plant, Aspalathus linearis (Rooibos Tea), as the free phenol, was demonstrated. The aglycone of aspalathin, phloretin, also a good antioxidant, was synthesized in aqueous medium without the need for protective groups. This one-step synthesis of phloretin, utilizing a recyclable and environmentally friendly catalyst, has potential commercial applications. For the past 30 years, Mosher esters have extensively been utilized as chiral derivatizing agents (CDA’s) for secondary alcohols and amides with great success. The empirical model employed to deduce the absolute stereochemistry of the resultant ester or amide requires a certain degree of conformational rigidity for the CDA, the origin of which has not yet been determined. The stabilizing atomic orbital hyperconjugative interactions and the location of the prevalent molecular orbitals in such esters were identified with the aid of ab initio calculations, 1H, 19F NMR and infrared spectrometry. Although flavonoid crystal structures are very rare, a novel (-)-epicatechin-Mosher ester crystal was obtained and analyzed by x-ray crystallography with interesting conformational observations. The theoretical model used to investigate the hyperconjugative effects in the Mosher ester, was published and utilized on a recently developed novel CDA that shows a greater degree of rigidity. This CDA, α-cyano-α-fluoro-p-tolyl-acetic acid (CFTA) has greater reactivity and was utilized in cases where the Mosher acid proved inadequate. The investigation regarding the employment of CFTA was carried out with Japanese collaboration and commercial production of the reagent is envisaged. The hyperconjugative molecular model used in the modeling of the Mosher reagent was refined and extended to CFTA. The origin of the very large fluorine chemical shift difference between the epicatechin-CFTA diastereomers was investigated and the results published.Item Open Access Structure and synthesis of polyphenols from cyclopia subternata(University of the Free State, 2002-06) Brand, D. J.; Brandt, E. V.; Kamara, B. I.English: Cyclopia subternata (Fabaceae), from which Honeybush tea is brewed, is one of approximately 24 Cyclopia species of woody legumes endemic to the Cape fynbos (Cape macchia) region of South Africa. Supported by results from our initial investigations on C. intermedia, demonstrating the presence of phenolic compounds including coumestans, isoflavones, flavanones, xanthones, a flavone, pinitol, p-coumaric acid and flavonoid glycosides, the tea is gaining popularity as a health beverage. Presence of these compounds that are claimed to have interesting pharmacological properties, supported by belief that the tea contains very little, if any, caffeine and low tannin content, as well as its usage as a medicinal plant by the people of the Western and Eastern Cape prompted investigations on the subternata species. The acetone and methanol extracts of the unfermented shoots and stems of C. subternata were subjected to chromatographic separations (Shephadex LH-20 column and preparative thin layer) which afforded a novel flavan, flavonols, flavanones, flavones, isoflavone and C6.C2- and C6.C1-type compounds. Their full acetate or methylated derivatives were elucidated and characterized by high resolution (300 MHz) ¹H NMR speetrometry which included COSY, NOESY, HMQC, HMBC, DEPT 135° experiments and ¹³C NMR spectroscopy, Molecular Modeling and Circular Dichroism. Along with (+)-Pinitol a novel apiofuranosyl (1"—›6')-glucopyranosyl carboxylic acid was isolated as a non-flavonoid compound. The flavonoid aglycones isolated included epicatechin-3-0-gallate (flavanol), luteolin (flavone) and orobol (isoflavone). The nonflavonoid glycosides included a 4-0-gl ucopyranosyl tyrosol, apiofuranosyl (l"—›6')- glucopyranosyl benzaldehyde and the acetate derivatives of two new O-glycosides namely 1-[(β-D-2',3' ,4' -tri-O-acetylglucopyranosyloxy ]-2-(3,5-diacetoxyphenyl)ethane and l-acetoxy-2-(β-D-2',3',4',6' -tetra-O-acetylglucopyranosyloxy)ethane. The flavonoid O-glycosides comprised a novel 3',4',7-trihydroxy-5-(glucopyranosyloxy) flavan, the flavone, scolymoside and the flavanones, hesperedin, narirutin and eriocitrin. C-glycosides isolated includes the xanthone, mangiferin, a C-6-glycosylated kaempferol as well as two new C-glycosides, 3,4' ,6, 7-tetrahydroxy-5-W-D-glucopyranosyl) flavonol and 3' ,4',5,5', 7-pentahydroxy-8-(β-D-glucopyranosyl) flavanone. Due to the novelty and the many proposed health properties of flavans, the relatively unexplored routes to flavonoid O-glycosylation and the effects of glycosylation on the aglycone solubility and general behaviour, including the absorption in tissues of man, prompted the synthesis of the isolated flavan glycoside. The uncertainty of the point of sugar attachment to the aglycone of some glycosides, especially with limited material available, further inspired the exploration of synthetic routes to glycosylated flavonoids. In an attempt to obtain the 5-O-glycosylated flavan, the selected glycosylation procedure was attempted on the analogous 5,7-dihydroxy-3',4'-dimethoxyflavanone, but proved unsuccessful because of the acidity of the 3-protons leading to ring opening. The same, procedure was attempted on the analogous chalcone with an unprotected A-ring. Glycosylations was successfully done on resacetophenone, phloroacetophenone and 4- methoxyphloroacetophenone as precursors to the formation of the glycosylated chalcone. However, the condensation with the appropriately protected benzaldehyde to obtain the glycosylated chalcone as precursor to the 5-O-glycosylated flavan was unsuccessful. The proposals that phenolic metabolites have physiological and therapeutic properties may also be associated with the compounds isolated from the tea. Luteolin and eriodictyol constitute part of the phenolic metabolic pool with a 3',4' -diol functionality which is claimed to have antioxidant activity. The antimicrobial activity of flavonoids in plants is also well documented and so is the antiviral activity of flavonoids and their ability to inhibit key enzymes in mitochondrial respiration. It was found that a C-2, C-3- double bond, a 4-keto group and a 3',4',5' -trihydroxylation of the B-ring are significant features of those flavonoids which show strong inhibition of NADH-oxidase. Coronary heart disease is also reported to be reduced in humans with high flavonoid intake. The anti-inflammatory and antitumor activity along with the ever-increasing interest in plant tlavonoids for treating human diseases and especially for controlling the immunodeficiency virus, the cause of AIDS, also attract interest in this herbal tea. These results clearly indicate that the claims of the health promoting properties of Honeybush tea may at least, in part, be attributed to the presence of these and other polyphenols in C. subternata.Item Open Access Struktuur en sintese van prorobinetinidien- en prosianidientipe gekondenseerde tanniene(University of the Free State, 1992-02) Cronje, Annemarie; Steynberg, J. P.; Ferreira, D.; Brandt, E. V.Afrikaans: Die wye kommersiële toepassing van die 'Mimosa' ekstrak van die tannienryke bas van Acacia mearnsii het as stimulus vir die eerste fitochemiese ondersoek van die fenoliese komponente in die nywerheidsgedroogde bas gedien. Toepassing van uitgebreide verrykings- en skeidingstegnieke lewer benewens die bekende mono-, di- en trimeriese voorlopers, die flavan-3-ol, epirobinetinidol, prorobinetinidien biflavanoïede met 3,4-cis relatiewe konfigurasie, (-)-robinetinidol-(4β,8)-(+)-gallokatesjien en (-)-robinetinidol-(4β,8)-(+)-katesjien, 'n A-tipe prorobinetinidien, (-)-robinetinidol-(2β - 7; 4β - 8)-(+ )-katesjien en 'n C-gealkileerde profisetinidien, (-)-fisetinidol-( 4α,8)-6-metiel-(+)-katesjien. Hierdie metaboliete word deur 'n reeks unieke prorobinetinidienverwante ringgeïsomeriseerde tanniene vergesel. Struktuurbepaling is gedoen deur benutting van hoe resolusie 'H k.m.r. n.O.e. tegnieke, sirkulere dichroïsme (SD), massaspektrometrie (MS) en is waar moontlik deur sintese, d.i. die basisgekataliseerde omskakeling (pH 10) van biflavanoïedvoorlopers, bevestig. Weens die vatbaarheid van biflavanoïede vir C-2-epimerisasie en herrangskikking via kinoonmetiede onder basiese kondisies is die 3',4'- en 4'-O-metieleterderivate (E-ring) van die biflavanoiedvoorlopers geselekteer vir 'n studie van hul basisgekataliseerde herrangskikking. Die piraanherrangskikkingsreaksie van (-)-robinetinidol-(4β,8)-(+)-katesjien mono-O-metieleter lewer 'n A-tipe prorobinetinidien, (-)-robinetinidol-(2β - 7; 4β - 8)-(+)-katesjien, met spektroskopiese data identies aan die van die natuurproduk. Hierdie verteenwoordig nie slegs die eerste A-tipe van die 5-deoksi (A-ring) oligoflavanoïede nie, maar ook die eerste in die klas van proantosianidiene met 'n 3,4-cis relatiewe konfigurasie van die C-ring. Die omskakeling van 'n B-tipe proantosianidien na 'n A-tipe analoog word in terme van die oksidatiewe verwydering van 'n hidriedioon by C-2( C) verklaar. Die rol van atmosferiese suurstof in hierdie oksidatiewe transformasie blyk duidelik uit 'n vergelykende basisgekataliseerde reaksie waarin spesiale voorsorgmaatreëls getref is om suurstofvrye kondisies te skep. Stereoselektiewe isomerisasies asook herrangskikking waartydens die pirogallol B- en A-ringe omruil t.o.v. die posisies in die 'normale' isomere, lewer die verwagte ringgeïsomeriseerde produkte en slegs klein hoeveelhede van die A-tipe analoog. Die 'H k.m.r. spektrum van die A-tipe proantosianidien met 3,4-cis relatiewe konfigurasie (C-ring) toon identiese koppelingskonstantes (J3,4 3.5 Hz) in vergelyking, met die van analoe met 3,4-trans stereochemie. Hierdie waarneming kan aan die konformasionele starheid van die bisikliese sisteem wat tot identiese dihedriese hoeke tussen 3-H(C) en 4-H(C) aanleiding gee, toegeskryf word. 'n Metode gebaseer op die hoogs selektiewe 'H n.O.e. assosiasie van 3-H(C) is ontwikkel waarvolgens die 3,4-trans en 3,4-cis isomere ondubbelsinnig onderskei kan word. Die basisgekataliseerde piraanherrangskikkingsreaksie van die 'beskermde' triflavanoïed voorloper, (4α,8: 4(α,6)-bis-(-)-ro binetinidol-(+)-katesjien di-O-metieleter lewer na 'n kort reaksietyd slegs die intermediere, 10-flavaniel[2,3f]- chromeen. Hierdie flobatannien is uit die kommersiële A. mearnsii-ekstrak geïsoleer en ontstaan via 'n enkele hoogs stereoselektiewe ringisomerisasie. Die basisgekataliseerde C-ring isomerisasies is ook uitgebrei om 'n reeks piranochromene verwant aan (-)-fisetinidol-(-)-epikatesjien biflavanoïede in te sluit. Die natuurlike voorkoms van 'n reeks flobatanniene met beide (+)- en (-)-epikatesjien sowel as (+)- en (-)-katesjien as tenninale eenhede, tesame met die waarneming dat die koppelingskonstantes (J2,3 ca. 1.0 en ca. 7.0 Hz onderskeidelik) vir die heterosikliese protone differensiasie bemoeilik, het die noodsaaklikheid van sintese beklemtoon. Die strukture van 'n reeks natuurprodukte uit Guibourtia coleosperma, Colophospermum mopane en Baikiaea plurijuga is aldus volgens hierdie protokol bevestig. Ondersoek is verder ingestel na reduktiewe degradasietegnieke ten einde die probleme rakende die struktuurbepaling van A-tipe proantosianidiene en veral die bepaling van die stereochemie van die terminale flavan-3-ol eenhede van hierdie klas van proantosianidiene uit te klaar. Vanwee verskeie kompliserende faktore byvoorbeeld die reduksie van die vry fenol, sou die geredelik beskikbare A-tipe proantosianidien uit Arachis hypogea L., (+ )-epikatesjien-(2α - 7; 4α - 8)-(+)-katesjien verkieslik as fenoliese metieleter aan die reduktiewe reaksiekondisies blootgestel word. Metilering met diasometaan lewer gevolglik 'n mengsel van die heksametiel- en heptametieleterderivate. Die beperkte toeganklikheid van diasometaan by 5-0H(A) kan aan die waterstofbinding tussen die betrokke hidroksisubstituent en O-1(F-ring) toegeskryf word. Blootstelling van die heptametieleter van (+)-epikatesjien-(2α - 7; 4α - 8)-(+)- katesjien aan diisobutielaluminiumhidried (DIBAL) lewer twee produkte wat as gevolg van splyting van aromatiese metieleterbindings op die 4-posisie van die B- en E-ringe onderskeidelik, ontstaan. Die benutting van natriumsianobóorhidried in trifluoroasynsuur (NaBH3CNj CF3COOH) het egter tot die suksesvolle bepaling van die stereochemie van die terminale eenheid in die A-tipe proantosianidien gelei. Die reeks produkte wat via piraanring- en interflavanielbindingsplyting ontstaan, kan meganisties verklaar word in terme van die suurgekataliseerde vorming van 'n karbokatioon wat herrangskikking na 'n intermediere kinoonmetied ondergaan. Nukleofiele aanval van die hidriedioon lewer die tetrahidropirano[2,3-h]chromene en brokstukke afkomstig vanaf beide die 'boonste' en 'terminale' eenhede. Bevestiging vir die voorgestelde meganisme volg uit die reaksie van die heptametieleterderivaat van die A-tipe proantosianidien met CF3COOH. In die afwesigheid van 'n hidrieddonor word slegs klein hoeveelhede (+)-katesjien, geëpimeriseerde uitgangstof en hoofsaaklik polimeriese materiaal verkry. Die voorkoms van die reeks C-ring gei"someriseerde verbindings in nywerheidsgedroogde A. mearnsii-bas, dra waarskynlik by tot die kommersiële sukses van wattelbasekstrak. Die 'vrystelling' van 'n nukleofiele resorsinolring in flobatanniene verhoog waarskynlik die reaktiwiteit teenoor formaldeheid en dra aldus by tot die suksesvolle benutting van wattelbasekstrak in die koudsettende kleefstofbedryf. Die relatief planere sentrale CDF trisikliese sisteme in die dimeriese en CDFI tetrasikliese sisteme in trimeriese C-ring geïsomeriseerde verbindings, behoort verder tot verhoogde affiniteit vir kollageensubstrate by te dra en verklaar waarskynlik die uitgebreide benutting van wattelbasekstrak in die leerlooibedryf.