ABSTRACT

Isolation and identification of Pseudomonas aeruginosa from Hospital environment for pyocyanin production was done. The isolates from Hospital bed and wound swab were identified as Pseudomonas aeruginosa. The virulence factor, antibiotic pigment, pyocyanin which is one of the characteristic feature of P. aeruginosa and which also confers on it growth advantage over competing bacteria occupying the same niche was also isolated and characterised. Quantitative assay of pyocyanin production was conducted which is based on the absorbance of pyocyanin at 520 nm in acidic solution. The concentration of pyocyanin was determined by multiplying the optical density at 520 nm (OD₅₂₀) by 17.072. The result showed that the isolates produced pyocyanin with the total concentration of 13μg/ml, while the isolates from wound swab produced no pyocyanin but pyorubin. Pyocyanin produced was characterized by biochemical tests, its dissolution in chloroform and Hydrochloric acid (HCl) to give a blue and pink to red colour respectively confirms the presence of pyocyanin.  The purification of pyocyanin was done by liquid-liquid extraction method using Na-borate buffer at the pH of 10 to neutralize the HCl and re extracted into chloroform and acid before storage. Since pyocyanin is labile to many conditions, the purified pyocyanin was stored in acid at freezing temperature in dark container to prevent decomposition. Since pyocyanin can be isolated successfully from P. aeruginosa, it can be used in vitro to carry out its antimicrobial activity on the growth of other microorganisms; also study on the mechanism of its virulence can be done using the isolated pyocyanin.