Hydrolysis Of Inulin Research Articles

Engineering a heterologously expressed fructosyltransferase from Aspergillus oryzae N74 in Komagataella phaffii (Pichia pastoris) for kestose production

Fructo-oligosaccharides (FOS) are one of the most well-studied and commercialized prebiotics. FOS can be obtained either by controlled hydrolysis of inulin or by sucrose transfructosylation. FOS produced from sucrose are typically classified as short-chain FOS (scFOS), of which the best known are 1-kestotriose (GF2), 1,1-kestotetraose (GF3), and 1,1,1-kestopentaose (GF4), produced by fructosyltransferases (FTases) or β-fructofuranosidases. In previous work, FOS production was studied using the Aspergillus oryzae N74 strain, its ftase gene was heterologously expressed in Komagataella phaffii (Pichia pastoris), and the enzyme’s tertiary structure modeled. More recently, residues that may be involved in protein–substrate interactions were predicted. In this study, the aim was to experimentally validate previous in silico results by independently producing recombinant wild-type A. oryzae N74 FTase and three single-point mutations in Komagataella phaffii (Pichia pastoris). The R163A mutation virtually abolished the transfructosylating activity, indicating a requirement for the positively charged arginine residue in the catalytic domain D. In contrast, transfructosylating activity was improved by introducing the mutations V242E or F254H, with V242E resulting in higher production of GF2 without affecting that of GF3. Interestingly, initial sucrose concentration, reaction temperature and the presence of metal cofactors did not affect the enhanced activity of mutant V242E. Overall, these results shed light on the mechanism of transfructosylation of the FTase from A. oryzae and expand considerations regarding the design of biotechnological processes for specific FOS production.

Glutaraldehyde functionalization of halloysite nanoclay enhances immobilization efficacy of endoinulinase for fructooligosaccharides production from inulin

Current work describes the enhancement of immobilization efficacy of Aspergillus tritici endoinulinase onto halloysite nanoclay using crosslinker glutaraldehyde. Under statistical optimized immobilization conditions, viz. glutaraldehyde 1.50% (v/v), enzyme coupling-time 2.20 h, glutaraldehyde activation-time 1.00 h and endoinulinase load 50 IU, maximum activity yield (65.77%) and immobilization yield (82.45%) was obtained. An enhancement of 1.15- and 1.23-fold in both enzyme activity yield and immobilization yield of endoinulinase was observed, when compared with APTES-functionalized halloysite nanoclay immobilized endoinulinase. Immobilized biocatalyst showed maximum activity at pH 5.0 and temperature 60 °C with broad pH (4.0–8.5) and temperature (50–75 °C) stability. Further, optimal hydrolytic conditions (inulin concentration 8.0%; endoinulinase load 80 IU; agitation 125 rpm and hydrolysis-time 13 h) supported fructooligosaccharides yield (95.44%) in a batch system. HPTLC studies blueprint confirmed 95.44% fructooligosaccharides containing 35.41% kestose, 26.19% nystose and 9.69% fructofuranosylnystose. The developed immobilized biocatalyst shown good stability of 8 cycles for inulin hydrolysis.

The inulin hydrolysis by recombinant exo-inulinases: determination the optimum temperatures and activation energies

The advantages of recombinant enzymes over native include the control in a production environment, product purity and also high yield. The paper presents the determination the optimum temperatures and the activation energies for various origin recombinant exo-inulinases, among others from Aspergillus niger, A. awamori, Kluyveromyces marxianus and K. cicerisporus. The parameters were estimated based on the literature of the activity curves versus temperature for hydrolysis of inulin. It was assumed that both the hydrolysis reaction process and the deactivation process of recombinant exo-inulinase were first-order reactions by the enzyme concentration. A mathematical model describing the effect of temperature on recombinant exo-inulinase activity was used. Based on the comparison analysis, values of the activation energies {E_{rm a }} were in the range from {32.01 pm 7.80} to {43.83 pm 4.87} kJ mol^{-1}, the deactivation energies {E_{rm d }} were in the range from {83.93 pm 4.82} to {352.44 pm 14.26} kJ mol^{-1} and the optimum temperature {T_{rm opt }} were obtained in the range from {318.91 pm } 1.19 to {328.76 pm 0.25} K.

Phosphate Enrichment of Niobium-Based Catalytic Surfaces in Relation to Reactions of Carbohydrate Biomass Conversion: The Case Studies of Inulin Hydrolysis and Fructose Dehydration

In this work, some physical mixtures of Nb2O5·nH2O and NbOPO4 were prepared to study the role of phosphate groups in the total acidity of samples and in two reactions involving carbohydrate biomass: hydrolysis of polyfructane and dehydration of fructose/glucose to 5-hydroxymethylfurfural (HMF). The acid and catalytic properties of the mixtures were dominated by the phosphate group enrichment. Lewis and Brønsted acid sites were detected by FT-IR experiments with pyridine adsorption/desorption under dry and wet conditions. Lewis acidity decreased with NbP in the composition, while total acidity of the samples, measured by titrations with phenylethylamine in cyclohexane (~3.5 μeq m−2) and water (~2.7 μeq m−2), maintained almost the same values. Inulin conversion took advantage of the presence of surfaces rich in Brønsted sites, and NbOPO4 showed the best hydrolysis activity with glucose/fructose formation. The catalyst with a more phosphated surface showed less deactivation during the dehydration of fructose/glucose into HMF.

Assembling and optimization of recyclable magnetite cross-linked inulinase aggregates for production of fructose syrups

The utilization of advanced nanoengineered technologies opens up attractive horizons towards sustainable treatment of waste biomass to produce fine chemicals. Production of fructose syrup (FS) in a single step using inulinase is a major achievements of enzyme technology as it eliminates the need for a multi-enzyme system. Functionalization of particles was done by coating chitosan on the surface of the magnetite and further, the chitosan coated magnetic nanoparticle (cMNP) was added to the mixture of cross-linked inulinase aggregates, and the physio-chemical properties of the aggregates were characterized. The immobilized biocatalyst was further utilized for the production of FS. Upon inulin hydrolysis for 4 h at 60 °C, 18.1 g/L of fructose was obtained with immobilized system. The immobilized inulinase was recycled up to 25 cycles and could be separated from the solution in 20 s by an external magnetic field.

Frutalose®, a mixture of fructans obtained from enzymatic hydrolysis of chicory inulin, and normal defecation: evaluation of a health claim pursuant to Article 13(5) of Regulation (EC) No 1924/2006.

Following an application from Sensus B.V. (Royal Cosun), submitted for authorisation of a health claim pursuant to Article 13(5) of Regulation (EC) No 1924/2006 via the Competent Authority of the Netherlands, the EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA) was asked to deliver an opinion on the scientific substantiation of a health claim related to Frutalose® and maintenance of normal defecation. The scope of the application was proposed to fall under a health claim based on newly developed scientific evidence. The Panel considers that Frutalose®, a mixture of fructans obtained from enzymatic hydrolysis of chicory inulin, is sufficiently characterised. The claimed effect proposed by the applicant is ‘contributes to regular bowel function by increasing stool frequency’. The proposed target population is ‘the general healthy adult population’. Maintenance of normal defecation is a beneficial physiological effect provided that it does not result in diarrhoea. One human intervention study showed an effect of Frutalose® on the maintenance of normal defecation by increasing stool frequency and improving stool consistency (softer stools) when consumed daily at a dose of 15 g for 8 weeks. The results have not been replicated in other studies. There is a plausible mechanism by which Frutalose® could exert the claimed effect. The Panel concludes that the evidence provided is insufficient to establish a cause and effect relationship between the consumption of Frutalose® and maintenance of normal defecation under the proposed conditions of use.

Optimization of Inulinase Production by a Fungal Species Isolated From Rotten Garlic Samples

Introduction: Inulinases are β-fructohydrolase enzymes that catalyze the hydrolysis of inulin. Recently, this enzyme has gained much importance mainly due to its ability to produce high-density fructose syrup using inulin as a raw material. In the current study, screening of inulinase-producing microorganisms was carried out from the rhizosphere soil of the Dahlia plant and rotten garlic samples. Materials and Methods: The inulinase activity was detected with the help of 3,5-dinitrosalicylic acid (DNSA) and Seliwanoff’s method, and the organism showing the highest potential was selected for further optimization studies. Results: The optimum culture conditions for inulinase production, by the test fungal culture, were observed when 5% inoculum was added to the minimal medium (pH 5.5) containing 1% inulin/ costus root powder as a carbon source and 0.15% NaNO3/ NH4Cl as a nitrogen source, and incubated at 30°C for 48h under shaker conditions (200 rpm). Maximum enzyme activity was observed at pH level of 5 and temperature level of 45°C, with thermal stability noted between 35°C-55°C. The I/S value of the crude enzyme was calculated to be 0.45 indicating true inulinase activity. It showed no significant inhibition in the presence of metal ions such as Zn+2, Mg+2, and Fe+3. The Ca+2 ions showed partial inhibition whereas Cu+2 ions showed an enhancement in the enzyme activity. Conclusions: These factors may present the test fungal culture isolated in the present study to be a potential candidate for the production of thermo-tolerant and metal resistant inulinase enzyme in order to be used for various biotechnological processes.

Quality Characteristics Carcasses Of New Zealand White Crossbred Rabbit Was Given FOS (Fructooligosaccharide) From Ambon Banana Peel Isolation

Fructooligosaccharide (FOS) is an alternative solution to control colibacillosis which can eliminate pathogenic bacteria in the contamination tract and stimulate the immune system (immunostimulants). The fructan contained in FOS produced by hydrolysis of chicory plant inulin has been widely reported, but the production of FOS from banana peel waste has not been widely published. This research aims to know and study the carcass characteristics of New Zealand White (NZW) crossbred rabbits given FOS (Fructooligosaccharide) from Ambon Banana Peel. The material used in this study were 42 New Zealand White crossbred rabbits 2 months old with 3 replications with 7 treatments, the percentage of FOS was 0%, 2.5%, 5%, 7.5%, 10%, 12.5%, and 15%, which is given through drinking water after 2 weeks of maintenance for 7 consecutive days at 08.00 am. The conclusion of this study that the provision of FOS (Fructooligosaccharide) from banana peel isolation had a significant effect on the carcass characteristics of New Zealand White crossbred rabbits (P <0.05) on carcass weight, meat weight, bone percentage, and had a very significant effect (P <0.01) on carcass percentage.

Development of a sustainable route for the production of high-fructose syrup from the polyfructan inulin.

The authors used mesoporous silica microspheres as a support for the immobilization of inulinase from Aspergillus brasiliensis MTCC 1344 by the process of cross‐linking. Under optimized operating conditions of pH 6.0, particle/enzyme ratio of 2.0:1.0 and glutaraldehyde concentration of 7 mM, a maximum immobilization yield of 90.7% was obtained after a cross‐linking time of 12.25 h. Subsequently, the cross‐linked inulinase was utilized for the hydrolysis of 5% inulin, and a maximum fructose concentration of 31.7 g/L was achieved under the optimum conditions of pH 6.0 and temperature 60°C in 3 h. Furthermore, on performing reusability studies during inulin hydrolysis, it was observed that the immobilized inulinase could be reused up to 10 subsequent cycles of hydrolysis, thus providing a facile and commercially attractive process of high‐fructose syrup production.

Optimization of Inulin Hydrolysis by Penicillium lanosocoeruleum Inulinases and Efficient Conversion Into Polyhydroxyalkanoates.

Inulin, a polydisperse fructan found as a common storage polysaccharide in the roots of several plants, represents a renewable non-food biomass resource for the synthesis of bio-based products. Exploitation of inulin-containing feedstocks requires the integration of different processes, including inulinase production, saccharification of inulin, and microbial fermentation for the conversion of released sugars into added-value products. In this work paper, a new microbial source of inulinase, Penicillium lanosocoeruleum, was identified through the screening of a fungal library. Inulinase production using inulin as C-source was optimized, reaching up to 28 U mL–1 at the 4th day of growth. The fungal inulinase mixture (PlaI) was characterized for pH and temperature stability and activity profile, and its isoenzymes composition was investigated by proteomic strategies. Statistical optimization of inulin hydrolysis was performed using a central composite rotatable design (CCRD), by analyzing the effect of four factors. In the optimized conditions (T, 45.5°C; pH, 5.1; substrate concentration, 60 g L–1; enzyme loading, 50 U gsubstrate –1), up to 96% inulin is converted in fructose within 20 h. The integration of PlaI in a process for polyhydroxyalkanoate (PHA) production by Cupriavidus necator from inulin was tested in both separated hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF). A maximum of 3.2 g L–1 of PHB accumulation, corresponding to 82% polymer content, was achieved in the SSF. The proved efficiency in inulin hydrolysis and its effective integration into a SSF process pave the way to a profitable exploitation of the PlaI enzymatic mixture in inulin-based biorefineries.

Application of the new method to determine the activation energies and optimum temperatures of inulin hydrolysis by exo-inulinases Aspergillus niger

Inulinases catalyze the hydrolysis of inulin to obtain fructose with a yield of about 90–95%. Inulin is a reserve carbohydrate in plant tubers like Jerusalem artichoke, chicory, garlic and onion, leek, rye, barley, dandelion, burdock, banana. The paper presents a comparative analysis to determine the optimum temperatures and the activation energies for various origin exo-inulinases from Aspergillus niger. The parameters were estimated based on the literature of the activity curves versus temperature for hydrolysis of inulin. It was assumed that both the hydrolysis reaction process and the deactivation process of exo-inulinase were first-order reactions by the enzyme concentration. The governing equations are formulated including the activation energy of the deactivation process and temperature effects. A mathematical model describing the effect of temperature on exo-inulinases from Aspergillus niger activity was used. Based on the comparison analysis, values of the activation energies $$E_{\mathrm{a}}$$ were in the range from $${25.20 \pm 2.92}$$ to $${60.95 \pm 8.30}\,\hbox {kJ}\,\hbox {mol}^{-1}$$ , the deactivation energies $$E_{\mathrm{d}}$$ were in the range from $${80.86 \pm 3.59}$$ to $${268.66 \pm 24.06}\,\hbox {kJ}\,\hbox {mol}^{-1}$$ and the optimum temperature $$T_{\mathrm{opt}}$$ was obtained in the range from $${325.25 \pm 0.41}$$ to $${337.35 \pm 0.70\,\hbox {K}}$$ .

The analisis of interaction of monosaccharides with aminoacids in food raw by quantum-chemical methods

The object of research is the products of hydrolysis of the polysaccharide inulin (glucose, fructose) of plant raw materials used for the production of food products for health-improving and prophylactic purposes. The search for optimal conditions for the transformation of polysaccharides in the processes of hydrolytic cleavage into fructose-oligosaccharide products requires a thorough study of both the chemical composition of the raw material and the interaction between the components. This affects both the stability of the initial, intermediate and final products of the hydrolysis of the polysaccharide, and its release from plant materials and the further course of its fragmentation. Literature data indicate that the chemical composition of carbohydrate-containing plant materials, in particular the content of mineral components, has not yet been fully studied and requires additional research and refinement. After all, macro- and microelements in its composition are not only important nutrients, but can also take an active part in the transformation of organic components through complexation at intermediate stages of these processes. In addition, the question of the possible interaction of organic compounds in the composition of food raw materials, in particular biopolymers, which make up a significant part of the mass – carbohydrates and protein compounds, also requires attention. Such interaction under normal conditions has practically not been studied, but it can affect the course of technological processes of processing. The study of intermolecular interactions occurring in complex natural systems is often complicated either by the absence of direct (selective) physical and physicochemical research methods, or by the multicomponent chemical composition of the system, or by the complexity of the objects (substances) of the study themselves. This is especially true for natural substances of a polymeric nature – proteins, peptides, poly- and oligosaccharides. Therefore, the work paid special attention to the study of the interaction of these components. The spatial structure of inulin molecules, oligosaccharides and elementary units of these polymers has been investigated using quantum chemical modeling. The distribution of effective charges on carbohydrate atoms is calculated; it directly affects their reactivity. And also quantum-chemical models of the interaction of protein substances of plant materials with carbohydrates in vacuum and in aqueous solutions are created, depending on their dilution.

Acid-assisted extraction and hydrolysis of inulin from chicory roots to obtain fructose-enriched extracts

Chicory (Cichorium intybus var. foliosum) roots are an agricultural residue and a low cost feedstock for the production of the platform chemical 5-Hxdroxymethylfurfural (HMF). In a first step, inulin and fructose have to be extracted from the roots. The resulting aqueous extract represents the starting material for the HMF production. In the reaction to HMF, inulin has to be hydrolyzed first to fructose. For this reason, two methods to increase the fructose content in these extracts before the reaction were investigated. This was conducted within the framework of integrating acid hydrolysis into a biorefinery process for HMF production. The first method (one-step process) was acid-assisted extraction to directly hydrolyze inulin in the course of the extraction process. Chicory roots were extracted at 60 and 80 °C at pH 2 and 4 using buffer solutions. The second approach (two-step process) was aqueous extraction at neutral pH followed by nitric acid hydrolysis of the extract at 60 and 80 °C under reduced pH. It was found that in the first approach, the pH of 2 led to a fivefold increase in the fructose content of the extract, resulting from inulin hydrolysis and corresponding to 56% of theoretical fructose yield. For the second approach, it was possible to achieve complete hydrolysis at pH below 2.5 and at 80 °C. Separating extraction and hydrolysis was found to be more suitable in terms of including this process step into a biorefinery concept for HMF production. It was possible to reduce the initial inulin content by 95%.

Engineering better catalytic activity and acidic adaptation into Kluyveromyces marxianus exoinulinase using site-directed mutagenesis.

Exoinulinase catalyzes the successive removal of individual fructose moiety from the non-reducing end of the inulin molecule, which is useful for biotechnological applications like producing fructan-based non-grain biomass energy and high-fructose syrup. In this study, an exoinulinase (KmINU) from Kluyveromyces marxianus DSM 5418 was tailored for increased catalytic activity and acidic adaptation for inulin hydrolysis processes by rational site-directed mutagenesis. Three mutations, S124Y, N158S and Q215V distal to the catalytic residues of KmINU were designed and heterologously expressed in Pichia pastoris GS115. Compared to the wild-type, S124Y shifted the pH-activity profile towards acidic pH values and increased the catalytic activity and catalytic efficiency by 59% and 99% to 688.4 ± 17.03 s-1 and 568.93 L mmol-1 s-1 , respectively. N158S improved the catalytic activity under acidic pH conditions, giving a maximum value of 464.06 ± 14.06 s-1 on inulin at pH 4.5. Q215V markedly improved the substrate preference for inulin over sucrose by 5.56-fold, and showed catalytic efficiencies of 208.82 and 6.88 L mmol-1 s-1 towards inulin and sucrose, respectively. Molecular modeling and computational docking indicated that structural reorientation may underlie the increased catalytic activity, acidic adaptation and substrate preference. The KmINU mutants may serve as industrially promising candidates for inulin hydrolysis. Protein engineering of exoinulinase here provides a successful example of the extent to which mutating non-conserved substrate recognition and binding residues distal to the active site can be used for industrial enzyme improvements. © 2020 Society of Chemical Industry.

Fabrication of porous polymer membrane from polysulfone grafted with acid ionic liquid and the catalytic property for inulin hydrolysis

Catalytically active membranes are the core components of the membrane reactors. Polysulfone (PSF) grafted with ionic liquids (ILs), PSF-DG-ILs, was prepared via introducing chloromethyl group into benzene ring of PSF, then grafting ILs, imidazolyl acid ILs ([(CH2)3SO3HVIm]HSO4), by chemical bonding. Then PSF-DG-ILs membranes were prepared by non-solvent induced phase separation technique for hydrolysis of inulin. The effects of coagulation bath, grafting degree, and various dope blending on structure, physico-chemical characters, and catalytic performance of membranes were evaluated. And results revealed that catalytic performance could be efficiently enhanced by choosing ethanol as coagulation compared with ethyl acetate, which was mainly attributed to surface segregation. Membrane mechanical strength decreased with the enhancing ILs grafting degree, while the catalytic performance increased. However, PSF-DG-ILs with high degree of ILs grafting lost its membrane-forming property. The mechanical and catalytic properties of the membrane were synergistically enhanced by blending PSF-DG-ILs polymers with high and low ILs grafting degree, then the resulting membranes broke the trade-off effect between mechanical strength and catalytic ability. Specifically, membrane fabricated by mixing the polymers with ILs grafting degree of 0.31 and 0.59 integrates optimized mechanical property and catalysis performance. The strength and strain of this blending membrane are 3.17 MPa and 14.10%. The average pore size and porosity are 215.5 nm and 46.2%. The yield of inulin hydrolysis can reach 88.22% with 20 wt% concentration of inulin at 75 °C using a flux of 23.4 L/(m2·h). Therefore, the PSF-DG-ILs membrane displayed promising potential for the application in biomass conversion.

Fusion and secretory expression of an exo-inulinase and a d-allulose 3-epimerase to produce d-allulose syrup from inulin.

This study developed a feasible catalytic method for d-allulose syrup production using a fusion enzyme, either in free or immobilized form, through hydrolysis of inulin extracted from Jerusalem artichoke tubers. d-Allulose 3-epimerase (DAE) was actively expressed in secretory form by fusing with the extracellular exo-inulinase CSCA in Escherichia coli BL21 (DE3). The best linker ligating the two enzymes was a flexible peptide containing 12 residues (GSAGSAAGSGEF). At 55 °C and pH 8.0, and as with the addition of 1 mmol L-1 Mn2+ , the CSCA-linkerE-DAE fusion enzyme obtained through high cell-density cultivation displayed a maximal exo-inulinase activity of 21.8 U mg-1 and resulted in a yield of 6.3 g L-1 d-allulose and 39.2 g L-1 d-fructose using 60 g L-1 inulin as the raw material. Catechol-modified alginate with titanium ions (Alg(Ti)PDA) was found to be a promising immobilization material for the fusion enzyme. After conversion for 8 days, the Alg(Ti)PDA-immobilized CSCA-linkerE-DAE (8 U g-1 ) completed 24 reaction cycles and retained over 80% of its original activity. Each reaction obtained an average of 19.8 g L-1 d-allulose and 32.7 g L-1 D-fructose from 60 g L-1 inulin. This study shed light on a feasible and cost-effective approach for the production of syrup containing d-allulose and D-fructose with inulin as the raw material via the use of a CSCA and DAE fusion enzyme. This syrup is of added value as a functional sweetener. © 2020 Society of Chemical Industry.

Inulin as a Promising Alternative Feedstock for Docosahexaenoic Acid Production by Schizochytrium sp. ATCC 20888

AbstractSchizochytrium sp. is considered as a promising alternative commercial source of docosahexaenoic acid (DHA), but the production is hindered by the high feedstock cost. In this study, inulin is used as a cheap and readily available feedstock for Schizochytrium sp. ATCC 20888 to produce DHA. The strain could not utilize inulin directly and therefore inulin first needed to be hydrolyzed. Compared with the acidic hydrolyzate by HCl and hydrolyzate by endo‐inulinase, the hydrolyzate by exo‐inulinase serves as the most effective carbon source for microalgal growth. Hydrolysis of inulin by exo‐inulinase is further optimized, and up to 97.8% of inulin conversion is obtained under the optimal conditions of 40 °C, pH 7.0, substrate concentration of 80 g L−1 and exo‐inulinase loading of 2 g kg−1 substrate for 12 h. The resulting hydrolyzate containing mainly fructose is used for the DHA production by the microalga. The lipid content in biomass, DHA content in total fatty acids, DHA yield, and DHA productivity at 72 h reach 45.26%, 35.59%, 5.64 g L−1 and 1.88 g L−1 d−1, respectively. The results suggest that inulin is an excellent feedstock for Schizochytrium sp. suitable for commercial DHA production.Practical Applications: DHA is an essential nutrient for human health and is widely used in infant formula and functional food. As a reserve carbohydrate, inulin present in plants represents a cheap, abundant, and readily available bioresource. This study describes the suitability of inulin as a promising alternative to glucose for DHA production by Schizochytrium sp. Hence, a practical bioprocess for commercial DHA production from inulin by Schizochytrium sp. could be developed. As far as it is known, this is the first report of inulin as a feedstock for Schizochytrium sp. to produce DHA.

Inulinase immobilization onto glutaraldehyde activated duolite XAD for the production of fructose from inulin

Abstract In the present practice, central composite rotatable design (CCRD) of response surface methodology (RSM) was used to optimise inulinase immobilization onto duolite XAD by gultaraldehyde (GA) cross-linking. After optimization, at optimal levels of variables (GA concentration 1.59%, enzyme coupling time 12 h, GA treatment time 4.0 h and enzyme load 22.7 IU), maximal inulinase activity yield and immobilization yield attained was 53.2% and 76.45%, respectively. A very narrow difference between predicted and experimental results confirms the authenticity of the present model. Besides, coefficient of determination (R2) value approaching one also authenticates the affluence of the designed polynomial model. Maximum hydrolysis (79.5%) of inulin by immobilized biocatalyst was achieved under agitation (125 rpm) mode of hydrolysis. Fructose production from inulin using inulinase immobilized onto duolite XAD was successfully performed for 10 consecutive batch cycles.

Exo-Inulinase Production by a Catabolite Repression-Resistant Mutant Thermophilic Aspergillus tamarii-U4 in Solid State Fermentation

In this study, spores of inulinase-producing thermophilic Aspergillus tamarii were subjected to UV mutagenesis, and colonies obtained were screened for inulinase production on inulin-glucose agar. The thermal stability of the inulinase was also investigated. A mutant strain U4 was found to produce 2.8 times inulinase titre (62.1U/mL) as against the wild strain (22.2U/mL). Inulinase production by this U4 strain was also found not to be significantly (P≤0.05) affected by the presence of glucose. The inulinase produced retained 64% of its activity after incubation at 65ºC for three hours. Solid-state fermentation for inulinase production by the strain U4 showed that wheat bran supported the highest inulinase titre 218.3U/gds while banana peels supported the lowest inulinase production titre of 80.5U/gds. Further optimization of cultural parameters revealed that incubation time of 5 days, 60% initial moisture content of the substrate, 2% inoculum density 2%, temperature 55ºC and pH 4.5 were optimal for inulinase production. Under optimized conditions, inulinase titre of 426.6 U/gds was observed. The pattern of inulin hydrolysis by the inulinase revealed the presence of monosaccharide as the main product of hydrolysis. Inulinase production at elevated temperatures by the mutant Aspergillus tamarii-U4 and its catabolite resistant properties showed that the organism is a potential industrial candidate for the production of exo-acting inulinases.

Inulinase Activity of Extracellular Protein of <i>Lactobacillus casei </i>AP in Different Growth Conditions

Inulinase is an enzyme that catalyzes the reaction involving the hydrolysis of inulin into fructose and/or small fructooligosaccharides by cutting the β-2,1 terminal. The present study aimed to determine the inulolytic activity of extracellular inulinase, extracted from L. casei AP, when grown under different conditions. Extracellular inulinase was extracted from the growth supernatant of L. casei AP, following which the extract was subjected to the qualitative inulinase test; inulinase activity was determined under growth conditions involving different combinations of pH and temperature. The results showed that L. casei AP had inulinolytic ability; therefore, it degraded inulin both in inulin media and inulin extract. The optimum activity of inulinase occurred at the combination of a temperature of 41 °C and pH of 5, with the activity of 20.53 in inulin media and 17.73 in inulin extract.