青春双歧杆菌的耐氧驯化及不同低聚糖对其增殖效果的影响

为改善青春双歧杆菌在发酵产品中活菌数低、菌种功效弱的情况,对厌氧青春双歧杆菌进行耐氧驯化,采用无氧有氧交替驯化法,在驯化过程中逐渐增加青春双歧杆菌培养液的氧分压,测定青春双歧杆菌驯化前后生理特性,并对比大豆低聚糖、低聚木糖、低聚麦芽糖对青春双歧杆菌增殖的影响。结果表明,青春双歧杆菌在耐氧驯化后对氧气的敏感程度下降,有氧条件下生长能力达标,产酸能力得到提升,菌体驯化前后形态基本一致,耐氧青春双歧杆菌代谢产生的乙酸与乳酸比值较厌氧青春双歧杆菌更适宜发酵,在3种低聚糖中,低聚木糖对青春双歧杆菌增殖作用明显,可作为青春双歧杆菌良好的双歧因子。耐氧驯化后的青春双歧杆菌有着优良的生理特性,能够作为潜在的益生菌菌种深入研究其功能并可应用于发酵食品中。     

Transglycosylation Activity of Engineered Bifidobacterium Lacto-N-Biosidase Mutants at Donor Subsites for Lacto-N-Tetraose Synthesis

The health benefits of human milk oligosaccharides (HMOs) make them attractive targets as supplements for infant formula milks. However, HMO synthesis is still challenging and only two HMOs have been marketed. Engineering glycoside hydrolases into transglycosylases may provide biocatalytic routes to the synthesis of complex oligosaccharides. Lacto-N-biosidase from Bifidobacterium bifidum (LnbB) is a GH20 enzyme present in the gut microbiota of breast-fed infants that hydrolyzes lacto-N-tetraose (LNT), the core structure of the most abundant type I HMOs. Here we report a mutational study in the donor subsites of the substrate binding cleft with the aim of reducing hydrolytic activity and conferring transglycosylation activity for the synthesis of LNT from p-nitrophenyl β-lacto-N-bioside and lactose. As compared with the wt enzyme with negligible transglycosylation activity, mutants with residual hydrolase activity within 0.05% to 1.6% of the wild-type enzyme result in transglycosylating enzymes with LNT yields in the range of 10–30%. Mutations of Trp394, located in subsite -1 next to the catalytic residues, have a large impact on the transglycosylation/hydrolysis ratio, with W394F being the best mutant as a biocatalyst producing LNT at 32% yield. It is the first reported transglycosylating LnbB enzyme variant, amenable to further engineering for practical enzymatic synthesis of LNT.     

低聚糖和多酚结合偏最小二乘判别分析鉴别油菜蜜产地

采用气相色谱-串联质谱技术测定油菜蜜中6 种低聚糖成分,高效液相色谱-串联质谱技术测定油菜蜜中18 种酚酸物质,并结合偏最小二乘判别分析（partial least square-discrimination analysis,PLS-DA）对来自湖北钟祥市、江苏盐城市、青海刚察县3 个具有显著地理、气候、环境差异的51 个油菜蜜样本进行产地鉴别。方差分析结果显示：3 个产地油菜蜜中松二糖含量具有显著性差异,且青海刚察油菜蜜低聚糖含量相对偏高;油菜蜜的18 种多酚类物质中大多数具有显著的地理差异性且湖北钟祥的油菜蜜中多酚含量相对偏高。多元统计分析结果显示多酚具有显著的地理特征性。油菜蜜中低聚糖和多酚的含量结合PLS-DA产地鉴别的预测精度可达到97%。     

Effects of oligosaccharides on phase transition temperatures and rheological characteristics of waxy rice starch dispersion

BACKGROUND: The creation of starch‐based foods incorporated with functional ingredients such as probiotics is of great current interest in the food industry. This study aimed to investigate the effects of prebiotic oligosaccharides on the phase transition temperatures and rheological characteristics of waxy rice starch dispersions. Four oligosaccharides were applied to the rice starch dispersions: chitooligosaccharides, fructooligosaccharides, isomaltooligosaccharides and xylooligosaccharides. RESULTS: The addition of 125 g kg^?1 oligosaccharides elevated the onset and peak temperatures for gelatinisation of 200–400 g kg^?1 waxy rice starch dispersions. The temperature of the storage modulus (G′) for gelatinisation increased markedly on adding fructooligosaccharides to 200–300 g kg^?1 waxy rice starch. For gelatinisation of 300 g kg^?1 rice starch dispersion the effectiveness of the oligosaccharides in changing the above parameters was as follows: chitooligosaccharides > fructooligosaccharides > isomaltooligosaccharides > xylooligosaccharides. Moreover, their effectiveness was dependent on the amylose content, as illustrated by comparing waxy and non‐waxy rice starches (amylose contents 9–256 g kg^?1). Importantly, the logarithmic G′₉₅ change was linearly and negatively correlated with amylose content. CONCLUSION: The results suggest that oligosaccharide‐containing rice starch dispersions may potentially be used for the formulation of oligosaccharide‐containing starchy functional foods owing to the rheological changes of these starch dispersions. Copyright © 2011 Society of Chemical Industry     

Suppression of Fusarium graminearium growth by differently structured starch types

We investigated the growth behavior and amylolytic enzymes of Fusarium graminearum cultivated on different types of native starch granules including barley (A‐type crystalline polymorph), potato and Curcuma zedoaria (B‐type crystalline polymorph), cassava (C‐type crystalline polymorph), and high AM maize (A + Vh‐type crystalline polymorphs). F. graminearum grew poorly on B‐type starches and the accumulation of biomass was similar to that obtained for fungi cultivated under carbohydrate starvation conditions. In comparison, three‐ to fivefold higher accumulation of fungal biomass was observed for growth on the A‐, C‐ and A + Vh‐type starches. Fungal glucoamylase and α‐amylase activity increased over time in the presence of native starch granules. Interestingly, resistant B‐type starches induced the highest amylolytic activity indicating that F. graminearum interacts with B‐type granules although only limited degradation occur. Starch degradation products maltose and malto‐oligosacharides was found to increase glucoamylase and α‐amylase activity, whereas glucose acted as a catabolite repressor.     

纳滤膜在功能性低聚糖分离纯化中的应用研究进展

功能性低聚糖具有抗肿瘤、抗放射、抗凝血、消炎和调节免疫力等医疗保健作用,广泛应用于食品科学和生物医药等领域。纳滤作为一种高效的膜分离技术,在功能性低聚糖的分离与纯化中的应用得到越来越多的关注。本文分析了纳滤膜对功能性低聚糖的分离机理,综述了纳滤膜在功能性多糖分离纯化中的应用进展,讨论了纳滤分离过程的影响因素,主要包括功能性多糖料液的性质、膜过程的操作参数以及膜材料本身的性质等。其中,料液的性质主要体现在组成、浓度、黏度等方面;操作参数主要体现在压力、温度、膜面流速和pH等方面;而膜材料的性质主要体现在微结构和表面性质两个方面。最后,进一步指出纳滤膜技术用于功能性多糖分离纯化时在设备成本、膜材料及膜污染等方面存在的问题,并对未来纳滤膜技术在低成本专用膜材料及系统开发和膜污染控制方面的研究进行了展望。     

MALDI Q-TOF CID MS for Diagnostic Ion Screening of Human Milk Oligosaccharide Samples

Human milk oligosaccharides (HMO) represent the bioactive components of human milk, influencing the infant’s gastrointestinal microflora and immune system. Structurally, they represent a highly complex class of analyte, where the main core oligosaccharide structures are built from galactose and N-acetylglucosamine, linked by 1–3 or 1–4 glycosidic linkages and potentially modified with fucose and sialic acid residues. The core structures can be linear or branched. Additional structural complexity in samples can be induced by endogenous exoglycosidase activity or chemical procedures during the sample preparation. Here, we show that using matrix-assisted laser desorption/ionization (MALDI) quadrupole-time-of-flight (Q-TOF) collision-induced dissociation (CID) as a fast screening method, diagnostic structural information about single oligosaccharide components present in a complex mixture can be obtained. According to sequencing data on 14 out of 22 parent ions detected in a single high molecular weight oligosaccharide chromatographic fraction, 20 different oligosaccharide structure types, corresponding to over 30 isomeric oligosaccharide structures and over 100 possible HMO isomers when biosynthetic linkage variations were taken into account, were postulated. For MS/MS data analysis, we used the de novo sequencing approach using diagnostic ion analysis on reduced oligosaccharides by following known biosynthetic rules. Using this approach, de novo characterization has been achieved also for the structures, which could not have been predicted.     

Removal of oligosaccharides in soybean flour and nutritional effects in rats

The objectives of this work were to establish a safe and economically viable process for the removal of raffinose oligosaccharides (RO) from soy flour and compare the effects of RO elimination from diets with regard to nutritional parameters by testing in Wistar rats. Debaryomyces hansenii UFV-1 was cultivated in suspension of defatted soy flour (1:10 w/v). An increase in α-galactosidase activity was observed in the medium, with a consequent decrease in the RO concentration. A total reduction of RO was achieved at 36 h of incubation. The diet containing soy flour free of RO presented higher digestibility, 91.28%, in relation to the diet containing soy flour with RO, 87.14%. However, the removal of the oligosaccharides from the diet did not promote a significant improvement in the values of weight gain, and other nutritional parameters tested on rats, during the experimental period of 14 days.     

The activation of fibroblast growth factors (FGFs) by glycosaminoglycans: influence of the sulfation pattern on the biological activity of FGF-1

Six synthetic heparin-like oligosaccharides have been used to investigate the effect of the oligosaccharide sulfation pattern on the stimulation of acidic fibroblast growth factor (FGF-1) induced mitogenesis signaling and the biological significance of FGF-1 trans dimerization in the FGF-1 activation process. It has been found that some molecules with a sulfation pattern that does not contain the internal trisaccharide motif, which has been proposed for high affinity for FGF-1, stimulate FGF-1 more efficiently than those with the structure of the regular region of heparin. In contrast to regular region oligosaccharides, in which the sulfate groups are distributed on both sides of their helical three-dimensional structures, the molecules containing this particular sulfation pattern display the sulfate groups only on one side of the helix. These results and the fact that these oligosaccharides do not promote FGF-1 dimerization according to sedimentation-equilibrium analysis, confirm the importance of negative-charge distribution in the activation process and strongly suggest that FGF dimerization is not a general and absolute requirement for biological activity.     

重组α-半乳糖苷酶酶解大豆低聚糖研究

本研究应用本实验室基因重组咖啡豆α-半乳糖苷酶对棉籽糖和水苏糖进行了酶解,应用薄层层析(TLC)和高效液相色谱(HPLC)法检测了酶解前后大豆低聚糖的变化情况,结合大豆制品传统加工工艺,提出了酶的添加方式和工艺。