Extraction of cocoa proanthocyanidins and their fractionation by sequential centrifugal partition chromatography and gel permeation chromatography

Erworben im Rahmen der Schweizer Nationallizenzen (http://www.nationallizenzen.ch)Cocoa beans contain secondary metabolites ranging from simple alkaloids to complex polyphenols with most of them believed to possess significant health benefits. The increasing interest in these health effects has prompted the need to develop techniques for their extraction, fractionation, separation, and analysis. This work provides an update on analytical procedures with a focus on establishing a gentle extraction technique. Cocoa beans were finely ground to an average particle size of <100 μm, defatted at 20°C using n-hexane, and extracted three times with 50 % aqueous acetone at 50°C. Determination of the total phenolic content was done using the Folin-Ciocalteu assay, the concentration of individual polyphenols was analyzed by electrospray ionization high performance liquid chromatography-mass spectrometry (ESI-HPLC/MS). Fractions of bioactive compounds were separated by combining sequential centrifugal partition chromatography (SCPC) and gel permeation column chromatography using Sephadex LH-20. For SCPC, a two-phase solvent system consisting of ethyl acetate/n-butanol/water (4:1:5, v/v/v) was successfully applied for the separation of theobromine, caffeine, and representatives of the two main phenolic compound classes flavan-3-ols and flavonols. Gel permeation chromatography on Sephadex LH-20 using a stepwise elution sequence with aqueous acetone has been shown for effectively separating individual flavan-3-ols. Separation was obtained for (-)-epicatechin, proanthocyanidin dimer B2, trimer C1, and tetramer cinnamtannin A2. The purity of alkaloids and phenolic compounds was determined by HPLC analysis and their chemical identity was confirmed by mass spectrometry

Notes on Australian Fungi - iv. Polyporus, Fomes and Hexagonia

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The impact of particle shape on the angle of internal friction and the implications for sediment dynamics at a steep, mixed sand–gravel beach

The impact of particle shape on the angle of internal friction, and the resulting impact on beach sediment dynamics, is still poorly understood. In areas characterized by sediments of specific shape, particularly non-rounded particles, this can lead to large departures from the expected sediment dynamics. The steep slope (1 : 10) of the mixed sand–gravel beach at Advocate Harbour is stable in large-scale morphology over decades, despite a high tidal range of 10 m or more, and intense shore-break action during storms. The Advocate sand (<i>d</i> < 2 mm) was found to have an elliptic, plate-like shape (Corey Shape Index, CSI ≈ 0.2–0.6). High angles of internal friction of this material were determined using direct shear, ranging from &phi; ≈ 41 to 49°, while the round to angular gravel was characterized as &phi; = 33°. The addition of 25% of the elliptic plate-like sand-sized material to the gravel led to an immediate increase in friction angle to &phi; = 38°. Furthermore, re-organization of the particles occurred during shearing, characterized by a short phase of settling and compaction, followed by a pronounced strong dilatory behavior and an accompanying strong increase of resistance to shear and, thus, shear stress. Long-term shearing (24 h) using a ring shear apparatus led to destruction of the particles without re-compaction. Finally, submerged particle mobilization was simulated using a tilted tray submerged in a water-filled tank. Despite a smooth tray surface, particle motion was not initiated until reaching tray tilt angles of 31° and more, being &geq;7° steeper than for motion initiation of the gravel mixtures. In conclusion, geotechnical laboratory experiments quantified the important impact of the elliptic, plate-like shape of Advocate Beach sand on the angles of internal friction of both pure sand and sand–gravel mixtures. The resulting effect on initiation of particle motion was confirmed in tilting tray experiments. This makes it a vivid example of how particle shape can contribute to the stabilization of the beach face

Free radical scavengers from Cymbopogon citratus (DC.) stapf plants cultivated in bioreactors by the temporary immersion (TIS) principle

Guillermo Schmeda-Hirschmann,Alejandro Tapia, Jose Cheel, Instituto de Quımica de Recursos Naturales, Universidad de Talca, Casilla 747, Talca, Chile. E-mail: [email protected] Cristina Theodulozb, Jaime Rodrıguez, Facultad de Ciencias de la Salud, Departamento de Ciencias Basicas Biomedicas, Universidad de Talca, ChileThe biomass production of Cymbopogon citratus shoots cultivated in bioreactors according to the temporary immersion (TIS) principle was assessed under different growth conditions. The effect of gassing with CO2-enriched air, reduced immersion frequency, vessel size and culture time on total phenolic and flavonoid content and free radical scavenging effect of the methanolic extracts was measured. From the TIS-culture of C. citratus, seven compounds were isolated and identified as caffeic acid (1), chlorogenic acid (2), neochlorogenic acid (3), p-hydroxybenzoic acid (4), p-hydroxybenzoic acid 3-O- -d-glucoside (5), glutamic acid (6) and luteolin 6-C-fucopyranoside (7). The occurrence of compounds 1Ð7 and their variability in C. citratus grown under different TIS conditions was determined by HPLC. The free radical scavenging effect of the methanolic extract and compounds was measured by the discoloration of the free radical 1,1- diphenyl-2-picrylhydrazyl (DPPH). The main metabolites in 6- and 8-week-old cultures, both in 5 and 10 l vessels, were chlorogenic acid (2) (100Ð113 mg%) and neochlorogenic acid (3) (80Ð 119 mg%), while in the cultures with CO2-enriched air and reduced immersion frequency the main compound detected in the extracts was glutamic acid (6) (400 and 670 mg% for the green and white biomass and 619 and 630 mg% for the green and white biomass, respectively). The most active compounds, as free radical scavengers, in the DPPH discoloration assay were caffeic acid (1), chlorogenic acid (2), neochlorogenic acid (3) and the flavonoid luteolin 6-C-fucopyranoside (7)