Project Title

Extraction and purification of biologically active metabolites from the Rhodococcus sp. MTM3W5.2

Authors' Affiliations

Mohrah Alenazi1, Jaimin kapadia2, Patrick D. South2, Bert Lampson2, Abbas G. Shilabin1 Department of Chemistry, College of Arts and Sciences, and Department of Health Sciences, College of Public Health, East Tennessee State University, Johnson City, Tennessee.

Location

Ballroom

Start Date

4-5-2018 8:00 AM

End Date

4-5-2018 12:00 PM

Poster Number

63

Name of Project's Faculty Sponsor

Abbas Shilabin,

Faculty Sponsor's Department

Department of Chemistry

Type

Poster: Competitive

Classification of First Author

Graduate Student-Master’s

Project's Category

Natural Sciences

Abstract Text

Due to an increasing prevalence of bacterial resistance to antibiotic drugs and the overuse of commercial antibiotics, the need to discover novel antibacterial compounds is becoming more urgent. There is one promising avenue of novel drug discovery which has begun to be explored; the analysis of secondary metabolites. Rhodococcus is a genus of gram-positive bacterium known for their ability to catabolize a wide range of compounds, and more notably for its ability to produce bioactive secondary metabolites. Rhodococcus belongs to the class actinobacteria. A species of Rhodococcus, MTM3W5.2, has been discovered in Morristown, Tennessee and was found to produce a metabolite with inhibitory activity against closely related species. The aim of this study is to elucidate the structure of the inhibitory metabolite by first isolating and purifying it, and then characterizing it using spectroscopic techniques. The compound was isolated from MTM3W5.2 RM broth cultures using n-butanol extraction, which yielded an active crude extract. The crude extract was then subjected to fractionation using a Sephedex LH-20 column with a 100% methanol solvent. The inhibitory activity of the fractions was tested through disk diffusion assay using Rhodococcus erythropolis as an indicator of inhibitory activity. Further preparation was completed using preparative reverse-phase high-performance liquid chromatography. Advanced purification was conducted using multiple rounds of analytical reverse-phase HPLC and activity was tested at each subsequent step using disk diffusion assay. Throughout the study, the HPLC fractions were characterized and the stability was monitored using UV-Visible spectroscopy. Two pure samples at 58.63 and 72.72 minutes from HPLC (High-performance liquid chromatography) collections were selected for further structural identification and are currently being studied using spectroscopic techniques, most notably 2D NMR spectroscopy (two-dimensional nuclear magnetic resonance).

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Apr 5th, 8:00 AM Apr 5th, 12:00 PM

Extraction and purification of biologically active metabolites from the Rhodococcus sp. MTM3W5.2

Ballroom

Due to an increasing prevalence of bacterial resistance to antibiotic drugs and the overuse of commercial antibiotics, the need to discover novel antibacterial compounds is becoming more urgent. There is one promising avenue of novel drug discovery which has begun to be explored; the analysis of secondary metabolites. Rhodococcus is a genus of gram-positive bacterium known for their ability to catabolize a wide range of compounds, and more notably for its ability to produce bioactive secondary metabolites. Rhodococcus belongs to the class actinobacteria. A species of Rhodococcus, MTM3W5.2, has been discovered in Morristown, Tennessee and was found to produce a metabolite with inhibitory activity against closely related species. The aim of this study is to elucidate the structure of the inhibitory metabolite by first isolating and purifying it, and then characterizing it using spectroscopic techniques. The compound was isolated from MTM3W5.2 RM broth cultures using n-butanol extraction, which yielded an active crude extract. The crude extract was then subjected to fractionation using a Sephedex LH-20 column with a 100% methanol solvent. The inhibitory activity of the fractions was tested through disk diffusion assay using Rhodococcus erythropolis as an indicator of inhibitory activity. Further preparation was completed using preparative reverse-phase high-performance liquid chromatography. Advanced purification was conducted using multiple rounds of analytical reverse-phase HPLC and activity was tested at each subsequent step using disk diffusion assay. Throughout the study, the HPLC fractions were characterized and the stability was monitored using UV-Visible spectroscopy. Two pure samples at 58.63 and 72.72 minutes from HPLC (High-performance liquid chromatography) collections were selected for further structural identification and are currently being studied using spectroscopic techniques, most notably 2D NMR spectroscopy (two-dimensional nuclear magnetic resonance).