The caries-preventive effects of arginine have been attributed to its impact on biofilm composition and pH. Recent in vitro studies suggest that arginine also affects the production of biofilm matrix components that contribute to virulence, but this mechanism has not been investigated clinically. This randomized, placebo-controlled, triple-blind, split-mouth in situ trial assessed arginine's impact on the microbial composition, matrix architecture, and microscale pH of biofilms from caries-active patients (N = 10). We also examined whether individual differences in the pH response to arginine were related to biofilm composition and matrix structure. Biofilms were grown for four days on carriers attached to intraoral splints. Three times daily, the biofilms were treated extraorally with sucrose (5 min), followed by arginine or placebo (30 min), in a split-mouth design. After growth, the microscale biofilm pH response to sucrose was monitored by pH ratiometry. Microbial biofilm composition and carbohydrate matrix architecture were analyzed by 16S rRNA gene sequencing and fluorescence lectin-binding analysis, respectively. Arginine treatment significantly mitigated sucrose-induced pH drops, reduced total carbohydrate matrix production, and altered the spatial distribution of fucose- and galactose-containing carbohydrates. Both arginine- and placebo-treated biofilms were dominated by streptococci and Veillonella spp. Paired analyses showed a significant reduction in mitis/oralis group streptococci and a non-significant increase in several arginine metabolizers in arginine-treated biofilms. Individual pH responses were not significantly associated with the abundance of specific bacterial taxa or carbohydrate matrix components. In conclusion, arginine reduced the virulence of biofilms from caries-active patients through multiple mechanisms, including suppressing matrix carbohydrate production.
Biofilms/drug effects* ; Humans ; Arginine/pharmacology* ; Hydrogen-Ion Concentration ; Dental Caries/prevention & control* ; Male ; Female ; Adult ; Double-Blind Method ; Sucrose/pharmacology*

The high content of sucrose and raffinose reduces the prebiotic value of soybean oligosaccharides. Fructan sucrases can catalyze the conversion of sucrose and raffinose to high-value products such as fructooligosaccharides and melibiose. To obtain a fructan sucrase that can efficiently convert soybean oligosaccharides, we first mined the fructan sucrase gene from microorganisms in the coastal areas of Xisha Islands and Bohai Bay and then characterized the enzymatic and catalytic properties of the enzyme. Finally, recombinant extracellular expression of this gene was carried out in Bacillus subtilis. The results showed that a novel fructan sucrase, BhLS 39, was mined from Bacillus halotolerans. With sucrose and raffinose as substrates, BhLS 39 showed the optimal temperatures of 50 ℃ and 55 ℃, optimal pH 5.5 for both, and K_cat/K_m ratio of 3.4 and 6.6 L/(mmol·s), respectively. When 400 g/L raffinose was used as the substrate, the melibiose conversion rate was 84.6% after 30 min treatment with 5 U BhLS 39. Furthermore, BhLS 39 catalyzed the conversion of sucrose to produce levan-type-fructooligosaccharide and levan. Then, the recombinant extracellular expression of BhLS 39 in B. subtilis was achieved. The co-expression of the intracellular chaperone DnaK and the extracellular chaperone PrsA increased the extracellular activity of the recombinant BhLS 39 by 5.2 folds to 17 U/mL compared with that of the control strain. BhLS 39 obtained in this study is conducive to improving the quality and economic benefits of soybean oligosaccharides. At the same time, the strategy used here to enhance the extracellular expression of BhLS 39 will also promote the efficient recombinant expression of other proteins in B. subtilis.
Oligosaccharides/metabolism* ; Glycine max/metabolism* ; Bacillus subtilis/metabolism* ; Sucrase/biosynthesis* ; Raffinose/metabolism* ; Fructans/metabolism* ; Sucrose/metabolism* ; Bacillus/genetics* ; Recombinant Proteins/biosynthesis* ; Bacterial Proteins/biosynthesis*

Sialyllactose is one of the most abundant sialylated oligosaccharides in human milk oligosaccharides (HMOs), which plays an important role in the healthy development of infants and young children. However, its efficient and cheap production technology is still lacking presently. This study developed a two-step process employing multiple-strains for the production of sialyllactose. In the first step, two engineered strains, E. coli JM109(DE3)/ pET28a-BT0453 and JM109(DE3)/pET28a-nanA, were constructed to synthesize the intermediate N-acetylneuraminic acid. When the ratio of the biomass of the two engineered strains was 1:1 and the reaction time was 32 hours, the maximum yield of N-acetylneuraminic acid was 20.4 g/L. In the second step, E. coli JM109(DE3)/ pET28a-neuA, JM109(DE3)/ pET28a-nst and Baker's yeast were added to the above fermentation broth to synthesize 3'-sialyllactose (3'-SL). Using optimal conditions including 200 mmol/L N-acetyl-glucosamine and lactose, 150 g/L Baker's yeast, 20 mmol/L Mg²⁺, the maximum yield of 3'-SL in the fermentation broth reached 55.04 g/L after 24 hours of fermentation and the conversion rate of the substrate N-acetyl-glucosamine was 43.47%. This research provides an alternative technical route for economical production of 3'-SL.
Child ; Humans ; Child, Preschool ; N-Acetylneuraminic Acid ; Escherichia coli/genetics* ; Lactose ; Fermentation ; Saccharomyces cerevisiae ; Oligosaccharides ; Glucosamine

In order to investigate the enzyme production mechanism of yak rumen-derived anaerobic fungus Orpinomyces sp. YF3 under the induction of different carbon sources, anaerobic culture tubes were used for in vitro fermentation. 8 g/L of glucose (Glu), filter paper (Flp) and avicel (Avi) were respectively added to 10 mL of basic culture medium as the sole carbon source. The activity of fiber-degrading enzyme and the concentration of volatile fatty acid in the fermentation liquid were detected, and the enzyme producing mechanism of Orpinomyces sp. YF3 was explored by transcriptomics. It was found that, in glucose-induced fermentation solution, the activities of carboxymethyl cellulase, microcrystalline cellulase, filter paper enzyme, xylanase and the proportion of acetate were significantly increased (P < 0.05), the proportion of propionate, butyrate, isobutyrate were significantly decreased (P < 0.05). The results of transcriptome analysis showed that there were 5 949 differentially expressed genes (DEGs) between the Glu group and the Flp group, 10 970 DEGs between the Glu group and the Avi group, and 6 057 DEGs between the Flp group and the Avi group. It was found that the DEGs associated with fiber degrading enzymes were significantly up-regulated in the Glu group. Gene ontology (GO) function enrichment analysis identified that DEGs were mainly associated with the xylan catabolic process, hemicellulose metabolic process, β-glucan metabolic process, cellulase activity, endo-1,4-β-xylanase activity, cell wall polysaccharide metabolic process, carbohydrate catabolic process, glucan catabolic process and carbohydrate metabolic process. Moreover, the differentially expressed pathways associated with fiber degrading enzymes enriched by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were mainly starch and sucrose metabolic pathways and other glycan degradation pathways. In conclusion, Orpinomyces sp. YF3 with glucose as carbon source substrate significantly increased the activity of cellulose degrading enzyme and the proportion of acetate, decreased the proportion of propionate, butyrate and isobutyrate. Furthermore, the degradation ability and energy utilization efficiency of fungus in the presence of glucose were improved by means of regulating the expression of cellulose degrading enzyme gene and participating in starch and sucrose metabolism pathway, and other glycan degradation pathways, which provides a theoretical basis for the application of Orpinomyces sp. YF3 in practical production and facilitates the application of Orpinomyces sp. YF3 in the future.
Animals ; Cattle ; Neocallimastigales/metabolism* ; Anaerobiosis ; Rumen/microbiology* ; Propionates/metabolism* ; Isobutyrates/metabolism* ; Cellulose/metabolism* ; Fungi ; Starch/metabolism* ; Glucose/metabolism* ; Acetates ; Sucrose/metabolism* ; Cellulases ; Cellulase

α-amylase is an endonucleoside hydrolase that hydrolyzes the α-1, 4-glycosidic bonds inside polysaccharides, such as starch, to generate oligosaccharides, dextrins, maltotriose, maltose and a small amount of glucose. Due to the importance of α-amylase in food industry, human health monitoring and pharmaceuticals, detection of its activity is widely required in the breeding of α-amylase producing strains, in vitro diagnosis, development of diabetes drugs, and the control of food quality. In recent years, many new α-amylase detection methods have been developed with improved speed and sensitivity. This review summarized recent processes in the development and applications of new α-amylase detection methods. The major principle of these detection methods were introduced, and their advantages and disadvantages were compared to facilitate future development and applications of α-amylase detection methods.
Humans ; alpha-Amylases/chemistry* ; Polysaccharides ; Oligosaccharides ; Starch ; Maltose

L-arabinose isomerase (L-AI) is the key enzyme that isomerizes D-galactose to D-tagatose. In this study, to improve the activity of L-arabinose isomerase on D-galactose and its conversion rate in biotransformation, an L-arabinose isomerase from Lactobacillus fermentum CGMCC2921 was recombinantly expressed and applied in biotransformation. Moreover, its substrate binding pocket was rationally designed to improve the affinity and catalytic activity on D-galactose. We show that the conversion of D-galactose by variant F279I was increased 1.4 times that of the wild-type enzyme. The K_m and k_cat values of the double mutant M185A/F279I obtained by superimposed mutation were 530.8 mmol/L and 19.9 s^-1, respectively, and the catalytic efficiency was increased 8.2 times that of the wild type. When 400 g/L lactose was used as the substrate, the conversion rate of M185A/F279I reached a high level of 22.8%, which shows great application potential for the enzymatic production of tagatose from lactose.
Galactose/metabolism* ; Limosilactobacillus fermentum/genetics* ; Lactose ; Hexoses/metabolism* ; Aldose-Ketose Isomerases/genetics* ; Hydrogen-Ion Concentration

We prepared 15 batches of Kaixin Powder benchmark samples with the decoction pieces of different batches. Further, we established the specific chromatograms and index component content determination method of Kaixin Powder benchmark samples and analyzed the peaks and similarity of the chromatograms. With sibiricose A5, sibiricose A6, polygalaxanthone Ⅲ, 3,6'-disinapoyl sucrose, ginsenoside Rb_1, β-asarone, α-asarone, and dehydropachymic acid as index components, the index component content determination method was established and 70%-130% of the mean content of each component was set as the range. The chromatograms of 15 batches of Kaixin Powder benchmark samples had a total of 22 characteristic peaks, among which 8 peaks were identified, which represented sibiricose A5, sibiricose A6, polygalaxanthone Ⅲ, 3,6'-disinapoyl sucrose, ginsenoside Rb_1, β-asarone, α-asarone, and dehydropachymic acid, respectively. The chromatograms shared the similarity of 0.992-0.999. The 15 batches of benchmark samples had sibiricose A5 of 0.34-0.55 mg·g~(-1), sibiricose A6 of 0.43-0.57 mg·g~(-1), polygalaxanthone Ⅲ of 0.12-0.19 mg·g~(-1), 3,6'-disinapoyl sucrose of 1.08-1.78 mg·g~(-1), ginsenoside Rb_1 of 0.33-0.62 mg·g~(-1), β-asarone of 2.34-3.72 mg·g~(-1), α-asarone of 0.11-0.22 mg·g~(-1), and dehydropachymic acid of 0.053-0.079 mg·g~(-1). This study established the specific chromatograms and index component content determination method of Kaixin Powder benchmark samples, and the method was simple, feasible, reproducible, and stable. This study provides a scientific basis for further research on the key chemical properties of the benchmark samples and preparations of Kaixin Powder.
Powders ; Ginsenosides ; Benchmarking ; Drugs, Chinese Herbal/chemistry* ; Sucrose ; Chromatography, High Pressure Liquid/methods*

Sweet potato is an important food crop that can also be used as an industrial raw material. Sucrose is the main form of long-distance carbohydrate transport in plants, and sucrose transporter (SUT) regulates the transmembrane transport and distribution of sucrose during plant growth and metabolism. Moreover, SUT plays a key role in phloem mediated source-to-sink sucrose transport and physiological activities, supplying sucrose for the sink tissues. In this study, the full-length cDNA sequences of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ were obtained by rapid amplification of cDNA ends (RACE) cloning according to the transcripts of the two SUT coding genes which were differentially expressed in sweet potato storage roots with different starch properties. Phylogenetic analysis was performed to clarify the classification of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆. The subcellular localization of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ was determined by transient expression in Nicotiana benthamiana. The function of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ in sucrose and hexose absorption and transport was identified using yeast functional complementarity system. The expression pattern of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ in sweet potato organs were analyzed by real-time fluorescence quantitative PCR (RT-qPCR). Arabidopsis plants heterologous expressing IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ genes were obtained using floral dip method. The differences in starch and sugar contents between transgenic and wild-type Arabidopsis were compared. The results showed IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ encoded SUT proteins with a length of 505 and 521 amino acids, respectively, and both proteins belonged to the SUT1 subfamily. IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ were located in the cell membrane and were able to transport sucrose, glucose and fructose in the yeast system. In addition, IbSUT₆₂₇₈₈ was also able to transport mannose. The expression of IbSUT₆₂₇₈₈ was higher in leaves, lateral branches and main stems, and the expression of IbSUT₈₁₆₁₆ was higher in lateral branches, stems and storage roots. After IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ were heterologously expressed in Arabidopsis, the plants grew normally, but the biomass increased. The heterologous expression of IbSUT₆₂₇₈₈ increased the soluble sugar content, leaf size and 1 000-seed weight of Arabidopsis plants. Heterologous expression of IbSUT₈₁₆₁₆ increased starch accumulation in leaves and root tips and 1 000-seed weight of seeds, but decreased soluble sugar content. The results obtained in this study showed that IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ might be important genes regulating sucrose and sugar content traits in sweet potato. They might carry out physiological functions on cell membrane, such as transmembrane transport of sucrose, sucrose into and out of sink tissue, as well as transport and unloading of sucrose into phloem. The changes in traits result from their heterologous expression in Arabidopsis indicates their potential in improving the yield of other plants or crops. The results obtained in this study provide important information for revealing the functions of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ in starch and glucose metabolism and formation mechanism of important quality traits in sweet potato.
Ipomoea batatas/metabolism* ; Arabidopsis/metabolism* ; Sucrose/metabolism* ; Saccharomyces cerevisiae/metabolism* ; DNA, Complementary ; Phylogeny ; Plants, Genetically Modified/genetics* ; Membrane Transport Proteins/metabolism* ; Starch/metabolism* ; Plant Proteins/metabolism* ; Gene Expression Regulation, Plant

Objective: To evaluate the efficacy and safety of intravenous iron supplementation in patients with recurrent iron deficiency anemia (IDA) . Methods: This retrospective analysis of 90 patients with recurrent IDA from May 2012 to December 2021 was conducted, comparing the efficacy and safety of the intravenous iron therapy group and the oral iron therapy group. Results: Among the 90 patients with recurrent IDA, 20 were males and 70 were females, with a median age of 40 (range: 14-85) years. A total of 60 patients received intravenous iron supplementation and 30 received oral iron supplementation. The hematologic response rates in the intravenous iron group were significantly higher than those in the oral iron group at 4 and 8 weeks after treatment [80.0% (48/60) vs 3.3% (1/30) and 96.7% (58/60) vs 46.7% (14/30), all P<0.001, respectively]. The median increase in hemoglobin levels was also significantly higher in the intravenous iron group than in the oral iron group [38 (4, 66) g/L vs 7 (1, 22) g/L at week 4 and 44.5 (18, 80) g/L vs 19 (3, 53) g/L at week 8, all P<0.001]. The intravenous iron group had a significantly higher proportion of patients who achieved normal hemoglobin levels than the oral iron group (55.0% vs 0 and 90% vs 43.3%, all P<0.001, respectively). Iron metabolism indicators were tested before and after 8 weeks of treatment in 26 and 7 patients in the intravenous and oral iron groups, respectively. The median increase in serum ferritin (SF) levels in the intravenous iron group 8 weeks after treatment was 113.7 (49.7, 413.5) μg/L, and 54% (14/26) of these patients had SF levels of ≥100 μg/L, which was significantly higher than the median increase in SF levels in the oral iron group [14.0 (5.8, 84.2) μg/L, t=4.760, P<0.001] and the proportion of patients with SF levels of ≥100 μg/L (P=0.013). The incidence of adverse reactions was 3.3% (2/60) in the intravenous iron group, which was significantly lower than that in the oral iron group [20.0% (6/30), P=0.015]. Conclusion: Intravenous iron supplementation is more effective for hematologic response, faster hemoglobin increase, and higher iron storage replenishment rates compared with oral iron supplementation in patients with recurrent IDA, and it is well tolerated by patients.
Male ; Female ; Humans ; Adolescent ; Young Adult ; Adult ; Middle Aged ; Aged ; Aged, 80 and over ; Anemia, Iron-Deficiency/epidemiology* ; Sucrose/therapeutic use* ; Ferric Compounds/therapeutic use* ; Retrospective Studies ; Iron/therapeutic use* ; Hemoglobins/therapeutic use*

Ultra-rapid cooling and rewarming rate is a critical technical approach to achieve ice-free cells during the freezing and melting process. A set of ultra-rapid solid surface freeze-thaw visualization system was developed based on a sapphire flim, and experiments on droplet freeze-thaw were carried out under different cryoprotectant components, volumes and laser energies. The results showed that the cooling rate of 1 μL mixed cryoprotectant [1.5 mol/L propylene glycol (PG) + 1.5 mol/L ethylene glycol (EG) + 0.5 mol/L trehalose (TRE)] could be 9.2×10 ³ °C/min. The volume range of 1-8 μL droplets could be vitrified. After comparing the proportions of multiple cryoprotectants, the combination of equal proportion mixed permeability protectant and trehalose had the best vitrification freezing effect and more uniform crystallization characteristics. During the rewarming operation, the heating curve of glassy droplets containing gold nanoparticles was measured for the first time under the action of 400-1 200 W laser power, and the rewarming rate was up to the order of 10 ⁶ °C/min. According to the droplet images of different power rewarming processes, the laser power range for ice-free rewarming with micron-level resolution was clarified to be 1 400-1 600 W. The work of this paper simultaneously realizes the ultra-high-speed temperature ramp-up, transient visual observation and temperature measurement of droplets, providing technical means for judging the ice free droplets during the freeze-thaw process. It is conducive to promoting the development of ultra-rapid freeze-thaw technology for biological cells and tissues.
Freezing ; Vitrification ; Cryopreservation/methods* ; Trehalose ; Gold ; Rewarming ; Metal Nanoparticles ; Cryoprotective Agents ; Lasers