| Abstract: |
SUMMARYChitin is abundant renewable natural resource found mainly in crustacean insects, moulluscan organs, fungi and algae. To date, the major sources of chitin comes from crustacean shells such as shrimp and crab (Wang and Chang, 1997). Chitin represents 14-27Seafood processing and consumption generate millions tons of shell fish wastes annually in the world. In Egypt, the solid waste generated from seafood processing is available 104 tons/year within the last decad and is still growing on. The disposal of sheMicrobial degradation of chitin and production of chitinase has captured the world wide attention of both industrial and scientific environments. But, unfortunately the utilization of these chitinaceous substances by thermophiles and production of thermosA review of previous work on chitinolytic fungi and hydrolysis of chitinous substrate is given in the introduction. The results obtained can be summarized in the following:1- Eight thermophilic & thermotolerant fungal isolates belonging to three genera were recovered on dilution plates, containing shrimp shell chitin as the sole carbon source, incubated at 45C from Egyptian soil and compost samples.2- The relative chitinolytic activities of the isolated thermophilic fungi at 45C revealed that only three strains of M. cinnamomea and A. fumigatus showed very strong chitinase productivity as grown on shrimp shell chitin-containing medium.3- It was obvious that, M. cinnamomea ML2 (the best strains) and A. fumigatus possessed the highest chitinase production under thermophilic conditions. Hence both two fungal isolate were chosen for further study.4- Optimization studies revealed that, an incubation periods of 8 and 6 days were found to be quite sufficient for reaching the maximum chitinase enzymes production and growth of both M. cinnamomea ML2 and A. fumigatus, respectively grown on chitin medium5- The optimization of culture conditions appeared that the tested fungi M. cinnamomea ML2 and A. fumigatus produced chitinases of high activity as grown in 50 ml aliquots of the basal medium in 250 ml Erlenmeyer flask and inoculated with 4% of 6-day old 6- Colloidal shrimp shell chitin was the carbon source which induce the maximum chitinases production by M. cinnamomea ML2 and A. fumigatus at the 1.5% concentration. While glucose was the best carbon source for producing mycelial dry weight at the 3% con7- On equal N basis, among the tested nitrogen sources, yeast extract was found to be the best nitrogen source for maximum growth and chitinase production by selected fungal isolates at concentration of 0.3%.8- The biosynthesis of the chitinase and growth of M. cinnamomea ML2 and A. fumigatus were found to be affected by the presence of phosphate, magnesium sulphate and K+ ions levels in the fermentation medium. The highest chitinase activity and growth rate 9- The addition of the lowest concentration of NaCl (5 g/L) to the medium adversly affected both chitinase production and growth of M. cinnamomea ML2 and A. fumigatus. Further increase in the NaCl level was inhibitory and no chitinases were produced in th10- On studying impact of ions of trace elements and some inhibitors on the growth and chitinase production by M. cinnamomea ML2 and A. fumigatus, it was found that the metal ions supplemented individually to the culture had no profound effect on growth r11- The addition of some B-growth vitamins, some natural additives and surface active agents to the culture medium individually also had no significant promoting effect on the synthesis of chitinase production and growth rate of both tested fungal species12- An attempt was conducted to simplify the medium and reduce its price cost. This could be achieved by growing the tested fungi M. cinnamomea ML¬2 and A. fumigatus, on solid state fermentation process containing shrimp-shell chitin of different moisture13- Generally, the optimization of the culture conditions as well as nutritional requirements revealed that, the tested fungi; M. cinnamomea and A. fumigatus produced chitinases of high productivity when grown in 50 ml portions of a medium containing; 1.514- Partial purification of the crude chitinases produced by M. cinnamomea ML2 and A. fumigatus were carried out by fractional precipitation with different agents namely; acetone, methanol, ethanol and ammonium sulphate separately. The highest specific ac15- With the following partial purification by mean of the dialysis with pure sucrose crystals the specific activities of chitinases produced by M. cinnamomea ML2 and A. fumigatus increased up to 52.1 & 36.0 U/mg protein with 2.39 & 1.75 fold purification16- Further purification of the partially purified and concentrated enzymes of both tested fungal species were achieved by using the gel filtration technique on Sephadex G200 column. The data revealed that, three protein components obtained from each test17- SDS-gel electrophoresis revealed that the purified enzyme components of each tested fungi were differed greatly in their molecular weights. The molecular weight of Ch.I was 64.5 kDa, while those of Ch.II and Ch.III were 41.7 kDa and 26.3 kDa, respecti18- Of the three enzyme components obtained from M. cinnamomea ML2, the second one (Ch.II), which covered the sharp peak of protein (peak No 2), was the highest in its specific chitinase activity (61.87 U/mg protein). Compared to the culture filtrate, the19- The pure major chitinases of M. cinnamomea ML2 (Ch.II) and A. fumigatus (A) were characterized regarding temperature, pH optima, pH and thermal stability, time course of chitinase action, enzyme and substrate concentration with special reference to Km20- The stability of the two tested purified enzymes were also investigated. The first pure M. cinnamomea chitinase (Ch.II) was more thermostable than the second one (A. fumigatus chitinase A). The M. cinnamomea chitinase (Ch.II) could be tolerated heatin21- On studying the impact of some metal ions and inhibitors on enzymatic reactions and chitinases activities of both M. cinnamomea (Ch.II) and A. fumigatus (A), the results revealed that, the first chitinase (Ch.II) was slightly activiated by Fe2+, Zn2+ 22- The pure chitinases, M. cinnamomea (Ch.II) and A. fumigatus (A) differ in their amino acid composition. The first enzyme (Ch.II) was characterized by being rich in cysteine (16.8%), histidine (14.0%), serine (11.10%) & glutamic acid (9.75%). The secon23- Finally, the antifungal properties of both thermostable M. cinnamomea chitinase II and A. fumigatus chitinase A were also tested against some plant pathogens namely; F. solani, F. oxysporum, F. sp. and A. niger, and it was found that, both tested chit
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