Penicillin Production by Penicillium Chrysogenum PCL 501: Effect of UV Induced Mutation

摘要

Penicillin production by Penicillium chrysogenum (PCL501) fermented on glucose, lactose and four agro-wastes (cassava shavings, corncob, sawdust, and sugarcane pulp) was monitored with HPLC. The highest amount of penicillin was obtained with sugarcane pulp. Penicillin yield from a 7-day culture of the fungus was 8.65 ± 0.05, 7.68 ± 0.03, 6.85 ± 0.05, 5.54 ± 0.01, 5.32 ± 0.05 and 2.23 ± 0.02μg/ml respectively on sugarcane pulp, glucose, cassava shavings, corncob, lactose, and sawdust. Two mutant strains, UVP1 and UVP2, were obtained by exposing the wild strain (PCL501) to ultraviolet irradiation for 20 and 25 minutes respectively. The 7-day culture of UVP1 and UVP2 on sugarcane pulp yielded 14.83 ± 0.05 and 14.97 ± 0.05 μg/ml penicillin respectively. This represented over 70% increase in penicillin production over the parent strain. There is a good prospect of producing cheaper and effective penicillin using the mutant strains of P. chrysogenum and sugarcane pulp as fermentation substrate. INTRODUCTION Penicillium chrysogenum, since its discovery in 1943, has remained the species of choice for industrial penicillin production; it yields much more penicillin than P. notatum, the first antibiotics-producing fungus isolated by Alexander Fleming, or any of its derivatives1,2,3. Efforts at producing cheaper and more effective penicillins had centered on isolation of new strains of the fungus, optimization of fermentation technique and improvement of available strains by classical mutagenesis procedures1,4.Mutation of P. chrysogenum with X-Rays, UV rays or other mutagens has been engaged since 1950s as a tool for enhancing penicillin production3. Mutant strains with 1000-fold increase on the amount of penicillin produced in Fleming's original culture have been developed from such improvement procedures5,6. Stauffer and Backus7 reported of high penicillin yielding P. chrysogenum Q-176 mutant obtained by UV irradiation. Plasma mutation method was recently used to develop a mutant strain of P. chrysogenum with high productivity of penicillin8.A strain of P. chrysogenum (PCL501) isolated from wood-wastes in Lagos, Nigeria9, is currently being investigated for penicillin production. Previous studies showed that the fungus produces β-lactam antibiotics with significant antibacterial effects against clinical isolates of E. coli and B. subtilis10. The fungus thrives on media supplemented with agro-wastes and utilizes such waste plant materials as sawdust, sugarcane pulp, cassava shavings, and corncob as carbon and energy sources for growth and antibiotics production10,11,12. In the present study, agro-wastes (cassava shavings, corncob, sawdust, and sugarcane pulp) and refined sugars (glucose and lactose) were compared as fermentation substrates for penicillin production by P. chrysogenum (PCL501). Ultraviolet (UV) irradiation was also used to produce mutant strains which yielded higher amounts of the antibiotics than the parent fungus. MATERIALS AND METHODS Preparations of Agro waste materials:Mature sugarcane stem (Saccharum offinarum), fresh maize (Zea mays) and cassava shavings (Manihot esculenta) were purchased from a local market in Mushin, Lagos, Nigeria. Sawdust of Abora wood (Mitragyna ciliata) was collected from sawmills at Ikorodu, Lagos, Nigeria.Peeled sugarcane stem was crushed and soaked overnight in distilled water. The pulp was washed in distilled water repeatedly until no trace of sugar was detected. Corncob, obtained by removing the maize grains, was cut into small sizes. The agro-wastes were dried to constant weight in the oven at 80oC and milled using Marlex Exceller grinder (Mumbai, India). Fine powder obtained after passing each agro-waste through 0.5 mm pore sized sieve was used as substrate in the fermentation media.Penicillium chrysogenum strain (PL 501)The wild strain of P. chrysogenum (PCL 501) was obtained from wood-wastes in Lagos, Nigeria as described previously9 and maintained on PDA slant at 4oC. Subcultures of