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Methods and proceduresIn normal phase adsorption chromatography, the stationary phase is more polar than the mobilephase and will interact more strongly with the most polar molecules than with molecules withlower polarity. A normal phase stationary phase will therefore retain polar compounds longer,and the less polar compounds will elute first.[20]Silica-GelSilica gel is a polymer of silicon oxide (SiO 2 .H 2 O), forming athree dimensional structure. The silica gel structure hasexposed silanol groups on its surface, giving it a polar natureand opportunity to form strong hydrogen bonds withcompounds. Polar compounds that easily form hydrogenbonds, as carboxylic acids, amines and amides, will be morestrongly retained on the silica gels than other less polarcompounds. Water and methanol have the potential to dissolvesilica-gel itself and it could contaminate the eluates. Silica gel used for open columnchromatography is usually in the size range of 40-200 μm. Smaller particles can give a highbackpressure, meaning that the flow of eluent is reduced because of the tight packing in thecolumn, and the elution would take longer.This can be overcome by using pressure, as in high pressure liquid chromatography (HPLC), orvacuum suction.[20]ProcedureFor CC over silica gel, a wide variation of mobile phase was used. By using analytical TLC, anappropriate mobile phase was selected with an Rf value of approximately 0.2 for the targetcompound. The sample was dissolved in a solvent and an appropriate amount of silica gel wasadded. The minimum amount of silica gel should be the same weight as the sample, and themaximum should not be more than twice the weight of the sample. By using a smaller amountof silica gel for the sample, there would be less to load in the column and it would be more timeefficient. The mixture of sample and silica was dried using a rotavapor, fitted with cotton toavoid dry silica being sucked away. The sample would adhere to the silica.Figure 3.1: Illustration ofSilica gelA column of appropriate size was selected on basis of the amount of silica to be used. Theamount of silica should be a minimum of 10 times, and maximum of 30 times the weight of the-24-
Methods and proceduressilica-sample. A larger amount of silica-gel would give a better separation of compounds thatare hard to separate, but the column would also take a longer time to complete.The silica gel was added to the column and packed by using vacuum suction. The surface ofthe silica should be flat and smooth. Then the silica gel sample was loaded on to the top, and asmall amount of silica was added on top of it for protection, before adding the mobile phase.The column was eluted by use of vacuum suction, and fractions of appropriate size, determinedby the size of the column, were collected. By using TLC, the endpoint of the column wasdetected when there were no more compounds emerging. Acetone was then used to wash outany remaining compounds.3.1.2 Reverse phase adsorption chromatographyReverse phase chromatography utilizes the same principles as normal phase, but the phasesare the other way around. The mobile phase is the more polar phase and the stationary phaseis the less polar one, retaining the compounds with the lowest polarity.[20]Octadecasilyl (ODS)Octadecasilyl is a bonded phase silica, which means that the physical properties of the silicahave been altered by reaction and bonding of the silanol groups. Here the silanol groups havebeen octadesilylated, which means that 18 carbon alkyl chains have replaced the silanol groupson the surface, giving the silica a high degree of hydrophobicity. A potential problem withbonded phase silica is non-reacted silanol groups that would reduce the hydrophobicity. Thesegroups can to some degree be deactivated by end-capping, meaning reacting them with a smallsilating agent such as trimethylchlorosilane (see Figure 3.2).[20]-25-
Page 1: Herbal plants used inTraditional Ch
Page 4 and 5: AbstractEight compounds were obtain
Page 6 and 7: 3.3 Natural Products Library at MRC
Page 8 and 9: IntroductionMRCTCM has developed a
Page 10 and 11: Introduction1.2.3 Fundamental Textu
Page 12 and 13: IntroductionClassification by thera
Page 14: Introduction1.2.7 Chinese Herbal Me
Page 17 and 18: Introduction1.4.1 TerpenesTerpenes
Page 19: IntroductionFrom triterpenes, compo
Page 22 and 23: • Agilent LC/MSD Trap XCT mass sp
Page 26 and 27: Methods and proceduresFigure 3.2: P
Page 28 and 29: Methods and proceduresSephadex LH-2
Page 30 and 31: Methods and proceduresof compounds
Page 32 and 33: Methods and proceduresdifferent fre
Page 34 and 35: Methods and proceduresmeasured from
Page 36 and 37: Results and discussionFigure 4.1: E
Page 38 and 39: Results and discussion4.2.1 E2a - M
Page 40 and 41: Results and discussionwas given the
Page 42 and 43: Results and discussionP:E=7:3 gave
Page 44 and 45: Results and discussionwas 20 ml, we
Page 46 and 47: Results and discussion4.3 Structura
Page 48 and 49: Results and discussionFigure 4.5: 1
Page 50 and 51: Results and discussionbelieved to c
Page 52 and 53: Results and discussionFigure 4.9: 1
Page 54 and 55: Results and discussionWith the exce
Page 56 and 57: Results and discussionFigure 4.12:
Page 62 and 63: Results and discussion4.3.4 C1, C4,
Page 65 and 66: Conclusion5 ConclusionThe history a
Page 67 and 68: References6 References1. Liu, Z., e

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