Identification And Mechanism Of Anti-Helicobacter Pylori Alkoloid From Derris Trifoliata

Abd Manaf, Uyub and Sulaiman, Shaida Fariza Identification And Mechanism Of Anti-Helicobacter Pylori Alkoloid From Derris Trifoliata. Project Report. Universiti Sains Malaysia.

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Abstract

Thin layer chromatography on chloroform extract of Derris trifoliata using a so lvent combination of Toluene:Ethyl pcetate: Diethylamine (7: 2: 1) gave five fra ctions and three of them were identified as alkaloids based on the development of brown colour after reacting with Dragendorff's reagent and fluorescing blue and green under UV scan, By column chromatography, these 3 fractions were reevaluated for their action against H. pylori and 13 other pathogen ic bacteria, Two fractions were found to specifica lly inhibit H. pylori wh ile the th ird fraction inhibited 6 other pathogenic bacteria tested and H. pylori, albeit weaker, Two fractions (F1 and F2) were found to specifically inhibit H. pylori wh ile the third fraction (F3) was not specific as it inhbited not only H. pylori but 6 other pathogenic bacteria tested albeit weaker. Toxicity test using brine shrimp assay revealed that LC50 for F1, F2 , F3 and chloroform crude extractwas 8.8 IJg/ml, 966,5 IJg/ml, 60A IJg/ml and 1,1 IJg/ml repectively, suggesting F2 was less toxic with LC50 va lue very close to the limit set at 1000 ug/ml. Batch cu lture and electron microscopic studies revealed that F2 transformed spira ls cells into coccoids as well as directly ki lled them, There was no obvious morphological changes and the action of the compound cou ld probably be inhibition of processes inside the cell. F2 was also tested against 45 clinica l H. pylori isolates to determine its MIC90, This parameter was also determined concurrently for metronidazole, tetracycline and amoxycillin, antibiotics which are commonly used against H, pylori, Ninety-six percent (43/45) of the isolates were metronidazole resistance (MIC>8IJg/ml) wh ich undermines success of treatment in tropica l countries, Although MIC90 for F2 was higher (8-16ug/ml) compared to amoxycillin (0 .24-0A8IJg/ml), it was lower compared to metronidazole (>40lJg/ml) and tetracycline(16-32 IJg/ml), Infact F2 killed all the metronidazole-resistance isolates at 32 IJg/ml, while amoxycillin, tetracycline and metronidazole at the highest concentration used(1 .92, 128, 40 IJg/ml) on ly killed 95%, 95% and 5% of the isolates respectively, F2 was further fractionated into two components, A and B. Both these separated components were still inhibitory again st H. pylori, with B slightly more active based on inhibition zone diameter, LC-MS stud ies revealed that both components were mixtures of several compounds. One major sharp peak at 1.64-2 ,50 min retention time was present in the A component and probable designation was "diphenoxine" based on NIST USA Idatabase. We have no information on the exact identity of F2

Item Type: Monograph (Project Report)
Subjects: Q Science > QH Natural history > QH301 Biology
Divisions: Pusat Pengajian Sains Kajihayat (School of Biological Sciences) > Monograph
Depositing User: Mr. HR
Date Deposited: 15 Mar 2017 09:17
Last Modified: 15 May 2017 16:14
URI: http://eprints.usm.my/id/eprint/29205

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