The volatile oil of Chromolaena Odorata: Its Antimicrobial and Inhibitory Effects on Partially Purified and Characterized Extracellular Protease of Pseudomonas Aeruginosa

Abstract

Context: The recent development in the antimicrobial therapy to meet the challenges of resistant strains of clinical pathogenic organisms has led to the insight of exploring the phytomedical properties of the volatile oils of medicinal plants.

Purpose: This work was designed to assess the antimicrobial activity of the volatile oils extracted from the leaf and stem of Chromolaena odorata on nine different types of enteric human pathogenic bacteria and to compare this effect with commonly used antibiotics. More importantly, the mode of inhibition of these oils on the extracellular protease of Pseudomonas aeruginosa was determined using double reciprocal plot.

Procedures: The volatile oils were extracted by hydrodistillation from air-dried leaves and stems of Chromolaena odorata. Antibacterial activity of these oils was tested against nine different types of both gram negative and positive pathogenic bacteria under favourable conditions and the results were compared with commonly used antibiotic drugs. In addition, the mode of inhibition of these volatile oils against partially purified and characterized extracellular protease of Pseudomonas aeruginosa was determined from Lineweaver Burke plot (double reciprocal plot).

Findings: The antimicrobial activity of the essential oils of Chromolaena odorata showed inhibition zones ranging from 13.0±1.0mm to 43.5±2.5mm in Salmonella paratyphimurium and Shigella dysenteriae respectively. Ceftriaxone, among other antibiotics, has the highest inhibition of 26.0±2.0mm against Salmonella paratyphimurium. There was a significant difference (p<0.05) between the total average inhibition of the antibiotics, 5.0±0.82 mm, and the volatile oils, 18.0±4.0mm.  Each of the microbes was either sensitive to both types of the oils or at least one of the oils. Enterohaemorrhagic Escherichia coli (EHEC) and Escherichia coli have the same lowest possible minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of 0.09%v/v and ≥0.18%v/v of the oils, hence the most sensitive among the pathogens. The extracellular protease of Pseudomonas aeruginosa had optimal activities at pH 7.5 and 35^oC. The volatile oils displayed a competitive inhibition against the extracellular protease of Pseudomonas aeruginosa with V_max of 0.91µmol/min and K_m of 0.48mg/ml in the absence of the volatile oils but the K'_m was increased to 0.93mg/ml and 1.25mg/ml in the presence of the volatile oils of the leaf and stem of this plant. The highest purification fold of 2.35 corresponding to 6.92µmol/min/mg protein was achieved from the crude enzyme with DEAE cellulose ion exchange chromatography. The successive purification profile revealed oligomeric nature of this protein.

Conclusion: Therefore, the volatile oil of the stems and leaves of Chromolaena odorata possessed antimicrobial activity with higher significant impact. In addition, it possessed ability to inhibit extracellular protease of Pseudomonas aeruginosa. This may probably suggest Chromolaena odorata as a possible source of nutraceuticals for clinical purpose.

KEYWORDS: volatile oil, Chromolaena odorata, antimicrobial, Pseudomonas aeruginosa, inhibition, extracellular protease