N-heterocyclic carbene catalyzed [2 + 3] annulation reaction for the synthesis of trifluoroethyl 3,2'-spirooxindole γ-lactam.

Asymmetric catalytic processes promoted by N-heterocyclic carbenes (NHCs) hold great potential for the sustainable preparation of chiral molecules. However, catalyzing the reactions by manipulating the reactive intermediates is challenging. We report herein that the known NHC-catalyzed [3 + 2] annulation reaction between ketimine and enal can also be turned into a [2 + 3] annulation reaction for the highly enantioselective direct synthesis of trifluoroethyl 3,2'-spirooxindole γ-lactams (4) through timely catalysis of the intermediates. DFT calculations revealed that this transformation included the key step of the nucleophilic attack of the Breslow intermediate M2 derived from NHC and enal (2) to the unattacked ketimine (1). Our study demonstrates that it is possible to tune the desired selectivities through the dynamic catalysts of the reactive intermediates.

Hybridization-based discovery of novel quinazoline-2-indolinone derivatives as potent and selective PI3Kα inhibitors.

INTRODUCTION: Phosphatidylinositol 3-kinases (PI3Ks) overexpression can elicit cellular homeostatic dysregulation, which further contributes to tumorigenesis, with PI3Kα emerging as the most prevalent mutant isoform kinase among PI3Ks. Therefore, selective inhibitors targeting PI3Kα have attracted considerable interest in recent years. Molecular hybridization, with the advantage of simplified pharmacokinetics and drug-drug interactions, emerged as one of the important avenues for discovering potential drugs. OBJECTIVES: This study aimed to construct PI3Kα inhibitors by hybridization and investigate their antitumor activity and mechanism. METHODS: 26 quinazoline-2-indolinone derivatives were obtained by molecular hybridization, and their structure-activity relationship was analyzed by MTT, in vitro kinase activity and molecular docking. The biological evaluation of compound 8 was performed by transwell, flow cytometry, laser scanning confocal microscopy, Western blot, CTESA and immunohistochemistry. RESULTS: Here, we employed molecular hybridization methods to construct a series of quinazoline-2-indolinone derivatives as PI3Kα selective inhibitors. Encouragingly, representative compound 8 exhibited a PI3Kα enzymatic IC50 value of 9.11 nM and 10.41/16.99/37.53-fold relative to the biochemical selectivity for PI3Kß/γ/δ, respectively. Moreover, compound 8 effectively suppressed the viability of B16, HCT116, MCF-7, H22, PC-3, and A549 cells (IC50 values: 0.2 µM âˆ¼ 0.98 µM), and dramatically inhibited the proliferation and migration of NSCLC cells, as well as induced mitochondrial apoptosis through the PI3K/Akt/mTOR pathway. Importantly, compound 8 demonstrated potent in vivo anti-tumor activity in non-small cell lung cancer mouse models without visible toxicity. CONCLUSIONS: This study presented a new avenue for the development of PI3Kα inhibitors and provided a solid foundation for novel QHIDs as potential future therapies for the treatment of NSCLC.

Novel sulfonyl-substituted tetrandrine derivatives for colon cancer treatment by inducing mitochondrial apoptosis and inhibiting PI3K/AKT/mTOR pathway.

Tetrandrine (TET) possesses multiple pharmacological activities and could suppress tumor proliferation via PI3K pathway inhibition. However, inferior antitumor activity and potential toxicity limit its clinical application. In the present study, a series of 14-sulfonamide and sulfonate TET derivatives were designed, synthesized, and evaluated for biological activities. Through structural-activity relationship studies, compound 3c with α, ß-unsaturated carbonyl group exhibited the most potent activity against all tested tumor cell lines (including Hela, HCT116, HepG2, MCF-7, and SHSY5Y), as well as negligible toxicity against normal cell lines LO2 and HEK293. Additionally, compound 3c effectively inhibited HCT116 and CT26 cell proliferation in vitro with increased cell proportion in the G2/M phase, activated the mitochondrial apoptosis pathway, and induced colon cancer cell apoptosis by suppressing the PI3K/AKT/mTOR pathway. The further molecular docking results confirmed that compound 3c is potentially bound to multiple residues in PI3K with a stronger binding affinity than TET. Ultimately, compound 3c dramatically suppressed tumor growth in the CT26 xenograft tumor model, without noticeable visceral toxicity detected in the high-dose group. In summary, compound 3c might present new insights for designing new PI3K inhibitors and be a potential candidate for colon cancer treatment.

1,4-α-Glucosidase from Fusarium solani for Controllable Biosynthesis of Silver Nanoparticles and Their Multifunctional Applications.

In the study, monodispersed silver nanoparticles (AgNPs) with an average diameter of 9.57 nm were efficiently and controllably biosynthesized by a reductase from Fusarium solani DO7 only in the presence of ß-NADPH and polyvinyl pyrrolidone (PVP). The reductase responsible for AgNP formation in F. solani DO7 was further confirmed as 1,4-α-glucosidase. Meanwhile, based on the debate on the antibacterial mechanism of AgNPs, this study elucidated in further depth that antibacterial action of AgNPs was achieved by absorbing to the cell membrane and destabilizing the membrane, leading to cell death. Moreover, AgNPs could accelerate the catalytic reaction of 4-nitroaniline, and 86.9% of 4-nitroaniline was converted to p-phenylene diamine in only 20 min by AgNPs of controllable size and morphology. Our study highlights a simple, green, and cost-effective process for biosynthesizing AgNPs with uniform sizes and excellent antibacterial activity and catalytic reduction of 4-nitroaniline.

Hypoglycemic Effects of Novel Panax notoginseng Polysaccharide in Mice with Diet-Induced Obesity.

In the study, the structural features and hypoglycemic effects of a polysaccharide Pan from the root of Panax notoginseng were investigated. The molecular weight of Pan was 8.27 kDa. Structural analysis indicated that Pan mainly consisted of →3)-ß-L-Rhap-(1→, →3,6)-ß-D-Galf-(1→ and →6)-ß-D-Galf-(1→ residues with acetyl groups. Pan exhibited good antioxidant activity. Pan could slow down the body weight and the content of blood glucose in the high-fat diet-induced mice, further suppress GLUT-2 and SGLT-1 expression in the intestines, and enhance p-IRS and p-AMPK expression in the livers, finally exhibiting hypoglycemic effects. The results could supply a direction for further research on polysaccharides as components for the control of hyperglycemia induced by obesity and diabetes.

Highly Permeable Polylactic Acid Membrane Grafted with Quaternary Ammonium Salt for Effective and Durable Water Disinfection.

Given the increasing usage of drinking water purifiers, highly permeable membranes with strong antimicrobial functions are desperately desirable for effective and durable water disinfection. Hereby, we prepared such antimicrobial membranes by chemical grafting of quaternary ammonium salt (QAS) molecules, 3-(trimethoxysilyl)propyldimethyloctadecyl ammonium chloride (TPMMC), onto air plasma pretreated biodegradable polylactic acid (PLA) substrates. The high chemical grafting density promoted very strong and positive zeta potential charge of the resulted PLA-QAS membrane, contributing to effective and broad-spectrum antimicrobial efficiencies (>99.99%) against different microbes, including fungi and conventional and drug-resistant bacteria. The solid grafting of QAS molecules produced a durable antimicrobial performance of the PLA-QAS membrane. In addition, the pleated filter (0.45 m2) of PLA-QAS membrane showed outstanding bacteria rejection properties (>99.99%) and excellent washing durability (up to 20 m3 water) even at very high water filtration rates (up to 4 L/min). The disinfection mechanism was clarified that negatively charged bacteria could be rapidly adsorbed to positively charged PLA-QAS spinnings, followed by devastating cell membrane damage to bacterial debris, leaving a clean environment without significant biofilm and biofouling formation.

Development of Millettia speciosa champ polysaccharide conjugate stabilized oil-in-water emulsion for oral delivery of ß-carotene: Protection effect and in vitro digestion fate.

In this study, a natural antioxidant emulsifier, Millettia speciosa Champ polysaccharide conjugates (MSC-PC), was used for fabricating oil-in-water emulsion, and the influences of MSC-PC on ß-carotene stability and bioaccessibility were studied. Results suggested that MSC-PC stabilized emulsion exhibited excellent resistance to a wide range of salt levels (0-500 mM of Na+), thermal treatments (50-90 °C) and pH values (3.0-11.0). MSC-PC also exhibited an outstanding inhibition capacity on lipid oxidation. Besides, MSC-PC stabilized emulsion had a better protective effect on ß-carotene than other systems. Interestingly, in spite of similar lipolysis extent, ß-carotene bioaccessibility in MSC-PC fabricated emulsion (14.75 %) was markedly higher than that in commercial Tween 80 fabricated emulsion (10.08 %), likely due to the steric-hindrance effect and antioxidant ability of MSC-PC, building interfacial layers that prevented ß-carotene from degradation. This work supplied a deep insight into elucidating the mechanisms of emulsifying performance and ß-carotene protection effect of MSC-PC fabricated emulsion.

Structure, Antioxidant Activity and In Vitro Hypoglycemic Activity of a Polysaccharide Purified from Tricholoma matsutake.

The structure, antioxidant activity and hypoglycemic activity in vitro of a novel homogeneous polysaccharide from Tricholoma matsutake (Tmp) were investigated. Structural features suggested that Tmp was consisted of arabinose (Ara), mannose (Man), glucose (Glc) and galactose (Gal) with a molar ratio of 1.9:13.6:42.7:28.3, respectively, with a molecular weight of 72.14 kDa. The structural chain of Tmp was confirmed to contain →2,5)-α-l-Arabinofuranose (Araf)-(1→, →3,5)-α-l-Araf-(1→, ß-d-Glucopyranose (Glcp)-(1→, α-d-Mannopyranose (Manp)-(1→, α-d-Galacopyranose (Galp)-(1→, →4)-ß-d-Galp-(1→, →3)-ß-d-Glcp-(1→, →3)-α-d-Manp-(1→, →6)-3-O-Methyl (Me)-α-d-Manp-(1→, →6)-α-d-Galp-(1→, →3,6)-ß-d-Glcp-(1→, →6)-α-d-Manp-(1→ residues. Furthermore, Tmp possessed strong antioxidant activity and showed the strong inhibitory effect on α-glucosidase and α-amylase activities. Then, a further evaluation found that there was a dramatic improvement in the glucose consumption, glycogen synthesis and the activities of pyruvate kinase and hexokinase when the insulin-resistant-human hepatoma cell line (IR-HepG2) was treated with Tmp. The above results indicated that Tmp had good hypoglycemic activity and also exhibited great potentials in in terms of dealing with type 2 diabetes mellitus.

Wheat gluten hydrolysates promotes fermentation performance of brewer's yeast in very high gravity worts.

The effects of wheat gluten hydrolysates (WGH) and their ethanol elution fractions obtained on XAD-16 resin on physiological activity and fermentation performance of brewer's yeast during very-high-gravity (VHG) worts fermentation were investigated. The results showed that the addition of WGH and their elution fractions in VHG worts significantly enhanced yeast biomass and viability, and further increased the fermentability, ethanol yield and productivity of yeast. Supplementation with 40% ethanol fraction exhibited the highest biomass (6.9 g/L dry cell), cell viability, fermentability (82.05%), ethanol titer (12.19%, v/v) and ethanol productivity during VHG worts fermentation. In addition, 40% ethanol fraction supplementation also caused the most consumption of amino acid and the highest accumulation of intracellular glycerol and trehalose, 15.39% of increase in cell-membrane integrity, 39.61% of enhancement in mitochondrial membrane potential (MMP), and 18.94% of reduction in intracellular reactive oxygen species (ROS) level in yeast under VHG conditions. Therefore, WGH supplementation was an efficient method to improve fermentation performance of brewer's yeast during VHG worts.

Bioprospecting of a novel endophytic Bacillus velezensis FZ06 from leaves of Camellia assamica: Production of three groups of lipopeptides and the inhibition against food spoilage microorganisms.

Food contamination caused by microorganisms has become a threat to consumers' health. Exploring antagonistic endophytes from plants of food raw-material and applying bioactive metabolites to inhibit the contamination has been an alternative and safer solution. In this study, we isolated and screened potential antagonistic endophytes from fresh Camellia assamica leaves, which were widely used in tea beverage production. We focused on a strain that showed visible inhibitory activity to Gram-positive bacteria, Gram-negative bacteria, and fungi. It was identified as a member of Bacillus velezensis and named FZ06. The results of genome analysis showed the strain FZ06 had 167 single-copy specific genes, much higher than those of most related strains. Also, 11 potential gene clusters of antimicrobial metabolites were found. Three groups of lipopeptides (surfactin, iturin, and fengycin) were identified by UPLC-MS/MS in purified antimicrobial methanol fraction of strain FZ06. The results of minimum inhibitory concentration (MIC) test proved the lipopeptide extract showed significant inhibitory effect on food spoilage bacteria (MIC 512-2048 µg/mL) and toxigenic fungi (MIC 128-256 µg/mL). In conclusion, this study suggests that the endophytic B. velezensis FZ06 and its lipopeptide extract hold great potential applications in the inhibition of food spoilage bacteria and toxic fungi in food industry.

Combinatorial synthetic pathway fine-tuning and cofactor regeneration for metabolic engineering of Escherichia coli significantly improve production of D-glucaric acid.

D-glucaric acid (GA) has been identified as among promising biotechnological alternatives to oil-based chemicals. GA and its derivatives are widely used in food additives, dietary supplements, drugs, detergents, corrosion inhibitors and biodegradable materials. The increasing availability of a GA market is improving the cost-effectiveness and efficiency of various biosynthetic pathways. In this study, an engineered Escherichia coli strain GA10 was constructed by systematic metabolic engineering. This involved redirecting metabolic flux into the GA biosynthetic pathways, blocking the conversion pathways of d-glucuronic acid (GlcA) and GA into by-products, introducing an in situ NAD+ regeneration system and fine-tuning the activity of the key enzyme, myo-inositol oxygenase (Miox). Subsequently, the culture medium was optimized to achieve the best performance of the GA10 strain. GA was produced at 5.35 g/L (extracellular and intracellular), with a maximized yield of ∼0.46 mol/mol on d-glucose and glycerol, by batch fermentation. This work demonstrates efficient biosynthetic pathways of GA in E. coli by metabolic engineering and should accelerate the application of GA biosynthetic pathways in industrial processes.

Using a novel polysaccharide BM2 produced by Bacillus megaterium strain PL8 as an efficient bioflocculant for wastewater treatment.

In this study, the purification and characterization of a novel polysaccharide-based bioflocculant BM2 produced by a bacterium Bacillus megaterium strain PL8 with self-flocculating property were investigated. The results showed that BM2 was an acidic polysaccharide composed of Gal, GalUA, Glc, GlcUA and Man at a molar ratio of 45.1: 33.8:9.3:9.2:2.4, respectively. The molecular weight of BM2 was 4.55 × 106 Da. BM2 had high flocculation efficiencies across a wide pH ranged from 4 to 11 and a wide temperature ranged from 20 to 100 °C towards kaolin clay. BM2 was a cation-independent bioflocculant which could achieve high flocculation activity without the addition of other cations. Adsorption bridging was the main mechanism in the flocculation process of BM2 towards kaolin clay. The BM2 also displayed a high removal efficiency in terms of Congo red (88.14%) and Pb2+ ions (82.64%). These results suggested that BM2 had a great potential as an efficient bioflocculant candidate in wastewater treatment.

Co-immobilization of multiple enzymes by self-assembly and chemical crosslinking for cofactor regeneration and robust biocatalysis.

Artificial multienzyme biocatalysts have played a crucial role in biosynthesis because they allow for conducting complex reactions. Here, we reorted a facile approach to fabricate multienzyme nanodevices with high catalytic activity and stability based on protein assembly and chemical crosslinking. The self-assembled partner SpyCatcher and SpyTag were genetically fused with 2,3-butanediol hydrogenase and formate hydrogenase to produce KgBDH-SC (SpyCatcher-fused 2,3-butanediol hydrogenase) and FDH-ST (SpyTag-fused formate hydrogenase), respectively. After assembling the two fusion proteins, the complexes were then immobilized on the functionalized silicon dioxide nanoparticles by glutaraldehyde, forming KgBDH-SC-ST-FDH-SiO2 with the capability of reducing 2-hydroxyacetophenone to (R)-1-phenyl-1,2-ethanediol with cofactor regeneration. Under the optimal conditions, the created co-immobilized enzymes performed 49% activity recovery compared with the mixture of free enzymes as well as showed 2.9-fold higher catalytic activity than the traditional random co-immobilized enzymes. Moreover, KgBDH-SC-ST-FDH-SiO2 showed better pH stability and organic solvents stability than the free enzymes, and remained 52.5% overall catalytic activity after 8 cycles. Finally, the co-immobilized enzymes can reduce 60 mM HAP for fabrication of (R)-PED with cofactor regeneration in the phosphate buffer reaction system, affording 83.9% yield and above 99% optical purity.

A novel polysaccharide from the roots of Millettia Speciosa Champ: preparation, structural characterization and immunomodulatory activity.

A novel polysaccharide fraction (MSCP2) was extracted and isolated from the roots of Millettia Speciosa Champ. Structural characterization revealed that MSCP2 had an average molecular weight of 2.85 × 104 Da and was composed of fucose, arabinose, galactose, glucose and xylose with a ratio of 2.20: 2.52: 4.04: 87.29: 3.96. Methylation analysis and nuclear magnetic resonance (NMR) analysis showed that the main glycosidic linkage types of MSCP2 were proved to be α-D-Glcp-(1→, →4)-α-D-Glcp-(1→, →4)-α-D-Xylp-(1→, →6)-ß-D-Galp-(1→, α-L-Araf-(1→, →3,4)-ß-L-Fucp-(1→ and →4)-α-D-GalpA-(1→. The immunomodulatory assay suggested that MSCP2 could significantly improve the pinocytic capacity and increase the secretion of nitric oxide (NO) and cytokines by regulating the corresponding mRNA expression in RAW 264.7 cells. The data from the membrane receptor assay demonstrated that the potential mechanisms of MSCP2-induced macrophage activation were mainly through toll-like receptor 4 (TLR4), scavenger receptor type A (SRA) and glucan receptor (GR)-mediated signaling pathways. These results suggested that MSCP2 can be developed as a promising immunomodulatory agent in functional foods.

Highly enantioselective resolution of racemic 1-phenyl-1,2-ethanediol to (S)-1-phenyl-1,2-ethanediol by Kurthia gibsonii SC0312 in a biphasic system.

The asymmetric resolution of racemic 1-phenyl-1,2-ethanediol (PED) to (S)-PED by Kurthia gibsonii SC0312 (K. gibsonii SC0312) was conducted in a biphasic system comprised of an organic solvent and aqueous phosphate buffer. The impacts of organic solvents on the whole cell catalytic activity, metabolic activity, membrane integrity, and material distribution were first evaluated. The results showed that all organic solvents, except for dibutyl phthalate, showed a detrimental effect on the metabolic activity of the cells, especially for those with low log P values. All organic solvents were capable of changing the membrane permeability and membrane integrity of the cells. Moreover, some organic solvents showed a good extraction of the oxidation product. Finally, a high yield of 47.7 % of (S)-PED was obtained by the asymmetric resolution of racemic PED using K. gibsonii SC0312 in a biphasic system under the optimal conditions: racemic PED 120 mM, temperature 35 °C, reaction time 6 h, 180 rpm, and a volume ratio of dibutyl phthalate to aqueous phosphate buffer of 1:1. The optical purity of (S)-PED increased from 51.3 % to >99 %. This work described an efficient approach to improve reaction efficiency, and constructed a highly effective biphasic reaction system for the fabrication of (S)-PED via K. gibsonii SC0312.

Metabolic engineering of a robust Escherichia coli strain with a dual protection system.

Considerable attention has been given to the development of robust fermentation processes, but microbial contamination and phage infection remain deadly threats that need to be addressed. In this study, a robust Escherichia coli BL21(DE3) strain was successfully constructed by simultaneously introducing a nitrogen and phosphorus (N&P) system in combination with a CRISPR/Cas9 system. The N&P metabolic pathways were able to express formamidase and phosphite dehydrogenase in the host cell, thus enabled cell growth in auxotrophic 3-(N-morpholino)propanesulfonic acid medium with formamide and phosphite as nitrogen and phosphorus sources, respectively. N&P metabolic pathways also allowed efficient expression of heterologous proteins, such as green fluorescent protein (GFP) and chitinase, while contaminating bacteria or yeast species could hardly survive in this medium. The host strain was further engineered by exploiting the CRISPR/Cas9 system to enhance the resistance against phage attack. The resultant strain was able to grow in the presence of T7 phage at a concentration of up to 2 × 107 plaque-forming units/ml and produce GFP with a yield of up to 30 µg/109 colony-forming units, exhibiting significant advantages over conventional engineered E. coli. This newly engineered, robust E. coli BL21(DE3) strain therefore shows great potential for future applications in industrial fermentation.

Fungal polysaccharide similar with host Dendrobium officinale polysaccharide: Preparation, structure characteristics and biological activities.

Based on the concept of endophytic fungus, if endophytic fungus can produce the same or similar product as the host plant, which will get rid of the restrictions of farmland, seasonal and pest, the active product could be sustainably obtained. In this study, an endophytic fungus polysaccharide FP showing the similar structure with the host Dendrobium officinale polysaccharide (DOP) was sustainably and cost-effectively obtained under preferred reaction conditions with different C/N ratio. The FP with high yield up to 2.77 ±â€¯0.51 g/L showed same monosaccharide composition with DOP as well as some host-plant-associated polysaccharides in published literatures. The main chain of FP was composed by →3,6)-ß-L-Man-(1→, α-D-Glc-(1→, →4)-α-D-Glc-(1→, →3,6)-ß-D-Gal-(1→, and →6)-ß-D-Gal-(1→, while the side chain was α-D-Glc-(1→. Meanwhile, FP was confirmed as a safe polysaccharide with good antioxidant, antiglycation and immunomodulatory activities. Furthermore, TLR2 and TLR4 were confirmed as the membrane receptors of FP on RAW264.7 cells.

Structure and immunomodulatory activity of polysaccharides from Fusarium solani DO7 by solid-state fermentation.

Two polysaccharides, DGS1 and DGS2, were obtained by solid-state fermentation (SSF) of Fusarium solani DO7, an endophytic fungus isolated from the orchid Dendrobium officinale. Structural characterizations revealed that DGS1 consisted of arabinose, glucose, mannose and galactose with a molar ratio of 2.9:13.4:3.0:1, respectively, and contained (1 → 5)-Araf, (1 → 4)-Glcp, (1 → 6)-Glcp, (1 → 3)-Manp, (1 → 2,6)-Manp and (1 → 6)-Galp glycosidic linkages, while DGS2 was composed of arabinose, glucose, mannose and galactose in a molar ratio of 3.5:8.1:2.1:1, respectively, and contained (1 → 5)-Araf, (1 → 4)-Glcp, (1 → 6)-Glcp, (1 → 3)-Manp and (1 → 6)-Galp glycosidic linkages. Neither polysaccharide was toxic to human embryonic kidney cells or mouse RAW 264.7 macrophage cells. An immunomodulatory activity assay indicated that both polysaccharides could significantly enhance the levels of TNF-α, IL-6 and NO by activating TNF-α, IL-6 and iNOs gene expression, respectively, especially DGS2. Interestingly, DGS2 also possessed relatively high antioxidant activity. These results illustrate that, due to its cost-effectiveness and environmentally friendly features, SSF has significant potential as a commercially competitive source of natural products, including fungal polysaccharides, with immunomodulatory activity.

Novel antibacterial polysaccharides produced by endophyte Fusarium solani DO7.

In this study, the primary antibacterial ingredients in Fusarium solani DO7 were confirmed as polysaccharides. After purification, two polysaccharides, DY1 and DY2, exhibited excellent antibacterial activity, especially to S. aureus and E. coli. Moreover, the glycosidic linkages of DY1 were composed of (1 → )-α-D-Glcp, (1 → 3)-ß-L-Rhaf, (1 → 4)-ß-D-Xylp, (1 → 6)-α-D-Glcp, (1 → 2,6)-α-D-Glcp and (1 → 2)-ß-D-Galp; while the glycosidic linkages of DY2 contained (1 → )-ß-D-Glcp, (1 → 2)-α-L-Rhaf, (1 → 3)-α-L-Araf, (1 → 4)-ß-D-Glcp, (1 → 4,6)-ß-D-Glcp and (1 → 3)-α-D-Galp. Additionally, DY1 and DY2 possessed nontoxicity to RAW 264.7 cells. Both polysaccharides could significantly promote the levels of TNF-α, IL-6 and NO by activating TNF-α, IL-6 and iNOs mRNAs expression. These results indicated that DY1 and DY2 had great potential as a food preservative and immunomodulatory agent.

Extraction, purification and antioxidant activity of novel polysaccharides from Dendrobium officinale by deep eutectic solvents.

Two novel polysaccharides DOP1-DES and DOP2-DES with molecular weight of 297,911 Da and 30,400 Da were extracted by using DESs from Dendrobium officinale for the first time, and purified with macroporous resin AB-8, Sephadex G75 and Sephadex G200 chromatography. Besides, the effect of extraction temperature, molar ratio of DES, reaction time and compound enzymes concentration on the extraction yield of polysaccharides were investigated. Monosaccharide composition analysis showed DOP1-DES and DOP2-DES were composed of D-glucose and D-mannose with the ratio of 2.2:1 and 3.7:1, respectively. Structural features of DOP1-DES and DOP2-DES were investigated by a combination of FT-IR, methylation analysis and NMR, which indicated that both polysaccharides had a similarly backbone consisting of (1, 4)-linked ß-D-Manp and (1, 4)-linked ß-D-Glcp with different ratio. Simultaneously, the two fractions possessed considerable antioxidant activity.