| Microbial inulinases are an important class of industrial enzymes that have gained much attention recently. Inulin was investigated as a source for the production of ultra-high-fructose syrup for commercial applications, through enzymatic hydrolysis by exoinulinase acting either alone or synergistically with endoinulinase. The general enzymic reaction mainly occurs by two enzymes: exoinulinase (EC 3.2.1.80), and endoinulinase (EC 3.2.1.7).Inulin has been defined as a polydisperse carbohydrate material consisting mainly, if not exclusively of 6 (2-1) fructosyl-fructose links. A starting glucose moiety can be present, but is not necessary. In contrast, levan which is formed by certain bacteria consists mainly or exclusively of 8 (2-6) fructosyl-fructose links. As is the case for inulin, glucose can be present, but again it is not necessary. Inulin is of great interest because it is a relatively inexpensive and abundant substrate for the production of rich fructose syrups. Due to low solubility in water, it is hydrolyzed in an acid medium at high temperature (80-100 ℃), although under these conditions fructose can be degraded easily, resulting in undesirable side products, such as difructose anhydrides.Different soil samples were collected from different locations. Samples were treated daily in vivo for 2 weeks with a 2 g l-1 inulin solution at 30℃, 37℃ and 45℃. The isolation medium had the following composition in g l-1 (NH4)2So4, 5.0; MgSO4 7H2O, 0.2; KH2PO4. 3.0; Mineral salts solution 2 ml; trace elements, 1.0 ml, mineral salts and trace elements sterilized at 120℃ for 20 min. Initial pH was 6.5, adjusted with 6 M NaOH. Inulin (Fluka) 2.0 g sterilized separately at 110℃ for 30 min. Inulinase assay was carried out to select the best strain among those microorganisms by dinitrosalicyclic acid reagentThirteen bacterial strains, seven fungal strains, streptomyces and a yeast strains were isolated. The fungal strains showed higher production among the other strains. Comparison among superior strains has been conducted using four factors (temperature, time, pH and enzyme thermostability). Results showed that the enzyme produced by Penicillium sp. 1 was higher than the other strains and more thermostable. Penicillium sp.l was identified as a Penicillium notatum.Optimization for exoinulinase production was done, by optimizing the media composition. Different carbon source (sugar, barley flour, corn flour, wheat flour, pure inulin, Jerusalem artichoke powder (JA) separately or in combination, inoculated andincubated at 34℃ on a rotary shaker at 150 rpm. (JA) powder was chosen as the best carbon source.Influence of different organic and inorganic nitrogen sources were tested (Corn steep liquor, Casein, Peptone, Trypton, Yeast extract, Beef extract, Urea Soybean, (NH4)SO4 and KNO3. Influence of pH (3-7) was also assayed. Results indicate that yeast extract is suitable as a nitrogen source and phosphate buffer at pH 3.5 was the best value. Different quantities of media were charged into 250 ml conical flasks, 30 ml medium with 200-rpm shaker speed gave satisfactory result.Using experimental design method optimized the process of inulinase production. Half Fractional Factorial Design (HFFD) was used to study seven factors, i.e. JA, YE, KH2PO4, MgSO4 7H2O, Inoculation quantity, Incubation time h-1, and pH. The relevant variables were selected by screening and Central Composite Design (CCD) was used to optimize the enzyme production. In this design three factors (JA, YE and inoculation%) were used in this design. The analysis by SAS program of these designs showed that the optimal medium compositions for exoinulinase production by Penicillium notatum were calculated to be: JA was 15 gl-1; inoculum size 3.0 % (v/v); yeast extract 5 gl-1; temperature 34 ℃; pH 3.5 (phosphate buffer); KH2PO4 3; MgSO4 7H2O, 0.2; medium quantity 30 ml per 250 ml conical flasks with shaking 200 rpm. The maximum response predicted from the model is 28.15 U ml-1. Repeated experiments were performed for the production...
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