菊糖果糖转移酶的研究进展

菊糖果糖转移酶(inulin fructotransferase,即InulaseⅡ)[EC 4.2.1.93]是指将菊糖水解为双果糖酐Ⅲ(DFAⅢ)的一种酶。文中对菊糖果糖转移酶的产生菌种、发酵工艺、分离纯化、酶学性质、酶解产物(DFAⅢ)生理功能等进行了综述。     

Production of a Novel Inulin Fructotransferase (DFAI producing) by Arthrobacter globiformis S14-3

A microorganism producing a novel inulin fructotransferase was isolated from a soil sample. The enzyme converted inulin into di-D-fructofuranose 1,2':2,1' dianhydride (DFAI) and a small amount of oligosaccharides. Through taxonomical studies, the microorganism was identified as Arthrobacter globiformis S14-3. The method for preparation of DFAI from inulin was investigated and established. The enzyme showed maximal activity at pH 6.0 and 40°C. The enzyme was stable up to 70°C at pH 6.0.     

Molecular cloning of the gene encoding the di-D-Fructofuranose 1,2':2,3' dianhydride hydrolysis enzyme (DFA IIIase) from Arthrobacter sp. H65-7

The gene encoding an intracellular enzyme hydrolyzing di-d-fructofuranose 1,2':2,3' dianhydride (DFA III) (DFA IIIase) was cloned from the genomic DNA of Arthrobacter sp. H65-7 for the first time. The single open reading frame (ORF) of the DFA IIIase gene consisted of 1368-bp encoding 455 amino acids. DFA IIIase showed a phylogenetically distinct position from other inulin-degrading enzymes and showed similarity only with inulin fructotransferases (depolymerizing) (inulase II, EC 2.4.1.93) from Arthrobacter globiformis C11-1, Arthrobacter sp. A-6, and Arthrobacter sp. H65-7 (48.7-50.3%), and inulin fructotransferase (DFA I-producing) (EC 2.4.1.200) from A. globiformis S14-3 (44.4%). An Escherichia coli transformant harboring a recombinant plasmid, pINB2, in which the DFA IIIase gene was fused with the beta-galactosidase of pUC19 and under the control of the lac promoter, expressed DFA IIIase and the cloned enzyme produced inulobiose from DFA III similarly to the DFA IIIase of the wild-type strain, Arthrobacter sp. H65-7.     

Cloning of inulin fructotransferase (DFA III-producing) gene from Arthrobacter globiformis C11-1

A gene encoding an inulin fructotransferase (DFA III-producing) [EC 2.4.1.93] from Arthrobacter globiformis C11-1 was cloned and the nucleotide sequence was determined. The cloned fragment contained a 1353 bp open reading frame. The initiation codon was estimated to be an unusual codon, GTG. The gene encoded a signal peptide (40 amino acid residues) for secretion. The molecular mass of the native enzyme was calculated as 43,400 Da from the sequencing data. The deduced amino acid sequence of the enzyme had 74.0 % homology with that of inulin fructotransferase (DFA III-producing) from Arthrobacter sp. H65-7. It also had 45.1% homology with that of inulin fructotransferase (DFA I-producing) [EC 2.4.1.200] from Arthrobacter globiformis S14-3. The enzyme produced in the culture supernatant of an Escherichia coli clone was purified to the electrophoretically homogeneous stage. The N-terminal amino acid sequence of the cloned enzyme secreted in the broth was the same as that of the native enzyme from A. globiformis C11-1. Therefore, on this enzyme, it is estimated that the cleavage sites by the signal peptidase for secretion of A. globiformis C11-1 and E. coli JM109 are the same.     

Efficient induction of inulin fructotransferase by inulin and by difructose anhydride III in Arthrobacter aurescens SK 8.001

Inulin fructotransferase (IFTase; EC 4.2.2.18) has received great attention mainly due to its application in producing difructose anhydride III (DFA III), which is a novel functional sweetener. The object of this study was to investigate the induction of IFTase in Arthrobacter aurescens SK 8.001 with various carbon sources, especially inulin and DFA III. IFTase production could be significantly promoted by the supplement of inulin (5–50 g/L) and DFA III (5–20 g/L). Inulin at high initial concentrations gave no indication of catabolite repression, whereas 30 and 40 g/L DFA III intensely inhibited cell growth and IFTase activity. No fructose was detected in broth throughout the cultivation with inulin, and inulin was converted into DFA III and minor fructooligosaccharides. And when DFA III was the carbon source, DFA III was the only sugar detected in the broth. In conclusion, both DFA III and inulin are effective for IFTase induction, and inulin with higher IFTase activity proved to be a more potent inducer.     

Inulin potential for encapsulation and controlled delivery of Oregano essential oil

The ability of inulin, a prebiotic material, as encapsulation matrix was explored. Microcapsules of Raftiline were produced by spray drying inulin solutions at different solids content (5, 15 and 25%) at 120, 155 and 190 °C, according to a Central Composite Rotatable design. Produced capsules were analysed for morphology and size by SEM and physiochemical characterized by DSC, IR and RAMAN. Oregano essential oil was incorporated in the inulin solutions at 15% solids basis and the emulsions dried at the same conditions. The above mentioned methodologies were applied to evaluate the encapsulation ability and the changes induced by the presence of the EO in capsules morphology and structure. Furthermore the kinetics and amount of release was assessed by a spectrophotometric method. Results showed that it was possible to produce regular spherical inulin microcapsules (3–4.5 μm) for all the tested experimental conditions. According to IR and Raman results mainly drying temperature affected the structure of the capsules, three groups being clearly formed. These groups could be related to the morphology of inulin crystals. The EO was successfully encapsulated in the system as demonstrated by IR and Raman analysis. The differences found in the EO releasing amount, make clear that different degrees of core material retention is achieved, what should be related to structural changes in the matrix wall, denoting in some processing conditions interactions phenomena among inulin and EO. Those different releasing profiles patterns may be quite useful in finding different potential uses for the encapsulates.     

Cloning and extracellular expression of inulin fructotransferase from Arthrobacter aurescens SK 8.001 in E. coli

BACKGROUND: Difructose anhydride (DFA) III is a natural and low‐calorie sweetener. It stimulates the absorption of calcium and other minerals. Inulin fructotransferase (IFTase; EC 4.2.2.18), catalysing inulin hydrolysis to DFA III, is considered to be the most promising enzyme for the production of DFA III. RESULTS: IFTase gene from Arthrobacter aurescens SK 8.001 was cloned and sequenced. Transformant with native IFTase signal peptide was a useful system for extracellular over‐expression of IFTase, and its extracellular IFTase activity reached 81.0 U mL^?1. This value was 4.1‐fold of that obtained with A. aurescens SK 8.001 for IFTase production. The recombinant IFTase was purified to electrophoretical homogeneity and characterized. The enzyme showed maximum activity at pH 6.0 and 55 °C, and retained 81.3% of its initial activity after incubation at 60 °C for 4 h. CONCLUSION: IFTase gene from A. aurescens SK 8.001 was cloned, sequenced and over‐expressed in E. coli. IFTase was reported for the first time to be over‐expressed extracellularly. The recombinant IFTase was purified and characterized, and shown to be a good candidate for potential application in DFA III production. Copyright © 2011 Society of Chemical Industry     

Preservation of porcine blood quality by means of lactic acid bacteria

The capacity of 12 lactic acid bacteria (LAB) strains to preserve porcine blood during storage was evaluated. A general ability of LAB to prevent blood's hemolysis and to maintain the functional properties of plasma was observed. Two strains, PS99 (Enterococcus raffinosus) and TA43 (Lactobacillus reuteri), were selected for studies at 5°C according to their antibacterial activity in blood stored at 15°C. After 144h at 5°C, lower counts of coliforms, Pseudomonas spp., proteolytic and hemolytic bacteria were obtained in blood containing either PS99 or TA43 as compared to the non-inoculated blood. When inulin (2%) was added to blood, higher inhibition values were obtained and Enterococcus raffinosus (PS99) showed the best abilities for blood preservation. On the basis of these results it seems worthwhile to supplement blood with inulin and to inoculate it with an active LAB strain to avoid undesirable changes during chill storage, especially useful to prevent the effects of a cold-chain breakdown.     

Effect of thermochemical treatment on the structure of inulin and its gelling properties

The aim of this study was to investigate rheological properties of inulin gels and the changes in the inulin structure after heating the high polymerised inulin (DP ≥ 23) solutions at different pH. The 20% inulin solutions were heated at pH 3, 5 and 7 at 60 and 80 °C followed by 21 h storage at 5 and 25 °C. Rheological properties evaluation indicated that the increase in the heating time and temperature significantly (P ≤ 0.05) influenced on the inulin structure. Reducing sugars, cryoscopy and HPLC analysis revealed that chemical structure of inulin is stable in neutral and slightly acidic conditions (pH 5). Gelation of the high polymerised inulin solution after heating at pH 7 and 80 °C could be inhibited after dissolving the inulin crystallites which act like seeding crystals. At lower temperatures in which, not all crystallites were dissolved; it was feasible for the solutions to form firm inulin gels.     

Effects of inulin/oligofructose on the thermal stability and acid-induced gelation of soy proteins

ABSTRACT:  Differential scanning calorimetry (DSC) and dynamic oscillatory shear testing were performed to study the influence of inulin (Raftiline^® HP-gel and Raftiline^® ST-gel) and oligofructose (Raftilose^® P95) on the thermal stability and gelation (using glucono-δ-lactone [GDL] as a coagulant) of soy protein isolate (SPI) dispersions. Addition of 10% (w/v) inulin/oligofructose or sucrose increased ( P < 0.05) the peak denaturation temperatures ( T_m ) of 7S and 11S soy proteins in SPI dispersion (5%[w/v], pH 7.0) by an average of 1.9 and 2.3 °C, respectively. GDL induced SPI thermal gelation, and the gel rheology was affected by both the pH decline and the specific temperature of heating. Addition of inulin/oligofructose (8%, w/v) improved the gelling properties of preheated SPI dispersion (8%, w/v) coagulated with GDL, showing 14.4 to 45.6% increase ( P < 0.05) in gel rigidity ( G ' value) at the end of heating (81 °C). Microstructural examination revealed a denser protein cross-linking structure and reduced pore sizes in SPI gels containing inulin/oligofructose. In general, inulin was more capable of improving SPI gelation than oligofructose, suggesting that the degree of fructose polymerization in the fructans was of thermal and rheological importance.     

Potentially probiotic and synbiotic chocolate mousse

Several studies in recent years have shown the benefits deriving from the ingestion of probiotics, and a large number of products containing lactobacilli and bifidobacteria have been formulated. The purpose of this study was to develop a chocolate mousse to which probiotic and prebiotic ingredients were added, and verify the perspectives of the product with regard to potential for consumer health benefits and sensorial acceptance. Probiotic and synbiotic chocolate mousse supplemented with Lactobacillus paracasei subsp. paracasei LBC 82, solely (P) or together with the prebiotic ingredient inulin (S), were prepared, as well as a control mousse (C). The products were monitored for the population of L. paracasei and contaminants, during storage at 5 °C for up to 28 days, and sensory preference was also tested. Storage trials showed that the viability of the probiotic was retained over 28 days, but the growth of yeasts and moulds might limit the shelf-life of the product. Chocolate mousse was shown to be an excellent vehicle for the delivery of L. paracasei, and the prebiotic ingredient inulin did not interfere in its viability. Moreover, the addition of the probiotic microorganism and of the prebiotic ingredient did not interfere in the sensorial preference of the product.     

Inulin and oligofructose improve sensory quality and increase the probiotic viable count in potentially synbiotic petit-suisse cheese

The influence of inulin, oligofructose and oligosaccharides from honey, combined in different proportions, on the consumers’ sensory acceptance, probiotic viable count and fructan content of novel potentially synbiotic petit-suisse cheeses was investigated. Probiotic populations varied from 7.20 up to 7.69 log cfu g⁻¹ (Bifidobacterium animalis subsp. lactis) and from 6.08 up to 6.99 log cfu g⁻¹ (Lactobacillus acidophilus). The highest fructan contents were achieved by the cheese trials containing oligofructose and/or inulin (above 8.90 g 100 g⁻¹). The control trial showed the lowest mean acceptance (6.63) after 28 days of refrigerated storage, whereas the highest acceptance (7.43) was observed for the trial containing 10 g 100 g⁻¹ oligofructose. Acceptance increased significantly during storage (P<0.05) only for cheeses supplemented with oligofructose and/or inulin. Cheeses containing honey did not perform well enough compared to the cheeses with addition of inulin and/or oligofructose, and the best synbiotic petit-suisse cheese considering sensory and technological functional features was that containing oligofructose and inulin combined, therefore encouraging the commercial product use.     

Impact of inulin and okara on Lactobacillus acidophilus La-5 and Bifidobacterium animalis Bb-12 viability in a fermented soy product and probiotic survival under in vitro simulated gastrointestinal conditions

The effect of inulin and/or okara flour on Lactobacillus acidophilus La-5 and Bifidobacterium animalis Bb-12 viability in a fermented soy product (FSP) and on probiotic survival under in vitro simulated gastrointestinal conditions were investigated throughout 28 days of storage at 4 °C. Employing a 2² design, four FSP trials were produced from soymilk fermented with ABT-4 culture (La-5, Bb-12, and Streptococcus thermophilus): FSP (control); FSP-I (with inulin, 3 g/100 mL of soymilk); FSP-O (with okara, 5 g/100 mL); FSP-IO (with inulin + okara, ratio 3:5 g/100 mL). Probiotic viabilities ranged from 8 to 9 log cfu/g during the 28 days of storage, and inulin and/or okara flour did not affect the viability of La-5 and Bb-12. Bb-12 resistance to the artificial gastrointestinal juices was higher than for La-5, since the Bb-12 and La-5 populations decreased approximately 0.6 log cfu/g and 3.8 log cfu/g, respectively, throughout storage period. Even though the protective effect of inulin and/or okara flour on probiotic microorganisms was not significant, when compared to a fresh culture, the FSP matrix improved Bb-12 survival on day 1 of storage and may be considered a good vehicle for Bb-12 and could play an important role in probiotic protection against gastrointestinal juices.     

Elaboration of a probiotic oblea from whey fermented using Lactobacillus acidophilus or Bifidobacterium infantis

A novel probiotic product was developed, which was formulated as an oblea (wafer-type dehydrated traditional Mexican dessert) using goat sweet whey fermented with Bifidobacterium infantis or Lactobacillus acidophilus. To obtain the probiotic oblea, the fermented whey was formulated with prebiotic carbohydrates (inulin and resistant starch) and gelatin, and the preparation was poured onto a polytetrafluoroethylene-coated nonstick baking pan, dried in a convection oven, and finally dehydrated at a low relative humidity and room temperature (23 ± 2°C). The amounts of prebiotic carbohydrates and gelatin to be used in the formulation were determined by a factorial experimental design. An untrained sensory panel evaluated 3 quality characteristics (film formation, homogeneity, and smoothness) in the final product. Three different drying temperatures were tested, namely, 40, 55, and 70°C. Bacterial survival at each temperature was determined by viable plate-counting. The best formulation, based on the quality characteristics tested, consisted of 58.33% (vol/vol) of fermented whey, 8.33% (vol/vol) of 6% (wt/vol) resistant starch dispersion, 16.66% (vol/vol) of 15% (wt/vol) inulin solution, and 16.66% (vol/vol) of a 10% (wt/vol) gelatin solution. Drying at 55 ± 2°C for 2.66 ± 0.22 h allowed for concentrations of probiotic bacteria above 9 log₁₀ cfu/g, which is above the minimum concentration required in a probiotic product.     

RHEOLOGICAL BEHAVIOR OF CONCENTRATED INULIN SOLUTION: INFLUENCE OF SOLUBLE SOLIDS CONCENTRATION AND TEMPERATURE

Inulin is a storage polysaccharide present in more than 30,000 vegetable products, including chicory roots, that are considered suitable for industrial application. The objective of this work was to evaluate the influence of temperature and the soluble solids concentration on the rheological behavior of a concentrated inulin solution obtained from a centrifugation process from chicory roots, after freezing at − 24C. For all the evaluated soluble solids concentrations, inulin solutions showed a rheological behavior of a highly pseudoplastic fluid, with high resistance to flow at low strain rates followed by a breakdown of the structure when the shear rate increased. The effect of temperature on the apparent viscosity of inulin solutions can be represented by the Arrhenius equation. The rheological behavior of inulin solutions can be represented by the Herschel–Bulkley, Casson, Cross and Power Law equations, where the consistency index increases as temperature rises and the soluble solids concentration as well.

PRACTICAL APPLICATIONS

The main objective of this work was to analyze the rheological behavior of a concentrated inulin solution obtained from chicory roots, to check the possibility of its application to obtain powder inulin.
The concentrated inulin solution was obtained by diffusion in hot water, followed by concentration by evaporation and phase separation conducted by lowering the temperature (−24C) and centrifugation at a velocity of 10,000 rpm for a time interval of 15 min. However, this solution is still going to be processed. Therefore, it is important to know its rheological behavior.
The influence of temperature and the soluble solids concentration on the rheological behavior of the concentrated inulin solution was also studied.     

菊粉酸降解动力学研究

通过考察温度、溶液pH值及水分含量对菊粉酸降解的影响,探索菊粉酸水解规律。结果表明:菊粉水溶液在pH 5.0～7.0、温度低于100℃时具有良好的稳定性;但当pH值低于4.0时,菊粉出现明显的水解反应。菊粉溶液在不同温度和pH值下的酸降解动力学表明,其水解反应遵循一级反应动力学方程。利用菊粉凝胶特性考察水分含量与菊粉酸降解的关系,发现相同pH值(pH=3)条件下,水分含量越高,菊粉降解速率越快,凝胶中菊粉降解速率低于菊粉水溶液降解速率。     

Effect of inulin on growth and bacteriocin production by Lactobacillus plantarum in stationary and shaken cultures

The prebiotic effect of inulin added to MRS medium on growth and bacteriocin production by L. plantarum ST16 Pa was investigated in stationary cultures in anaerobic jars with medium containing 0.025% sodium thioglycolate or in flasks shaken at 100 rpm. In the presence of 1% inulin in anaerobic stationary cultures, this strain produced lactic acid at a level that was 36.5% higher than in the absence of the polysaccharide. In shaken cultures without inulin, cell count was 54% higher than in the stationary ones. Under stationary conditions in anaerobic jars, the addition of inulin increased the maximum specific growth rate from 0.37 to 0.49 h^?1 and reduced the generation time from 1.85 h to 1.40 h. Consequently, the exponential phase was shortened from 12 to 9 h when the cells were grown in stationary cultures with the oxygen scavenger. Despite this effect of inulin on growth rate, stationary cultures without inulin displayed higher antimicrobial activity against Listeria monocytogenes L104 (3200 AU/mL) than cultures with inulin (1600 AU/mL); therefore, inulin behaved as a compound able to accelerate growth rather than to stimulate bacteriocin production. The results presented in this study are very promising, as L. monocytogenes is a well‐known foodborne pathogenic microorganism. Moreover, L. plantarum ST16 Pa has proven to be a potential producer of a natural food preservative at an industrial level.     

不同聚合度菊粉的吸附特性

为了研究菊粉聚合度不同对其吸附特性的影响规律,以天然和长链菊粉为原料,采用离心法与静态吸附法对菊粉的吸油特性、吸湿特性和水合性能进行分析。结果发现,天然菊粉对植物油脂的吸附量在30℃时达到最大值1.17 g/g,对动物油脂的吸附量在50℃时达最大值1.04 g/g;温度对天然菊粉吸附油脂量的影响比长链菊粉显著,随温度升高天然菊粉吸附油脂量呈显著下降趋势。天然菊粉的吸湿性比长链菊粉强,随温度的升高,两类菊粉的吸湿率和溶解度都增加,但持水性和膨胀度均呈先升高后下降的趋势,长链菊粉出现最大值时所对应的温度比天然菊粉高20~30℃。     

Replacement of fat with long-chain inulin in a fresh cheese made from caprine milk

The aim of this study was to evaluate the effect of the replacement of fat with long-chain inulin on textural and microstructural properties of a fresh caprine milk cheese. All the samples contained the same level of total solids (about 22%, w/w) and substitution of fat with inulin at levels from 2% to 7%. Penetrometry parameters were affected by the levels of replacement of fat with inulin; samples containing inulin were characterised by lower values of compressive force, stiffness, viscosity and adhesiveness (except for the sample with 2% fat substitution). Scanning electron microscopy (SEM) images showed that the positioning of inulin within the gel interrupted the casein/fat network. In conclusion, the rheological results were dependent on the arrangement of inulin within the protein/fat network and, in particular, as shown by SEM images, on the aggregation of inulin during cheese-making.