DEAE-cellulose Chromatography Research Articles

Purification and characterization of a thermostable Galium aparine β-galactosidase: A competent agent with enhanced cytotoxic activity against MCF-7 cell line

β-galactosidases (EC 3.2.1.23) are capable of catalysing two distinct types of reactions: transgalactosylation and hydrolysis. It is very important for the senescence of cancer cells. It is a lysosomal exoglycosidase involved in the catabolism of glycoconjugates by sequential release of β-linked terminal galactosyl residues. It has profound significance in cancer cell senescence. It can be derived from various sources including plants, animals, bacteria, yeasts, and filamentous fungi. Purification and thermodynamic characterization of β-galactosidase from Galium aparine seeds, and in vitro efficacy assessment of a breast cancer cell line. The enzyme was purified by using ammonium sulphate precipitation, dialysis, DEAE cellulose, and Sephadex G-100 chromatography. The enzyme was purified to 6.4 fold with a yield of 14 % and specific activity of 11.6 U/mg of protein. SDS-PAGE and MALDI-TOF analysis revealed that β-galactosidase is a monomer with a molecular weight of 35 kDa. The optimum pH and temperature for the purified enzyme were 6.0 and 50 ºC. The enzyme had Km and Vmax values of 0.25 mM and 32 µmol/min, respectively. The thermodynamic parameters (ΔHº, ΔGº, ΔSº) for the irreversible denaturation of the enzyme were also investigated at different temperatures (50–75 ºC). Lactose in milk was successfully hydrolyzed using a purified enzyme at a rate of 0.0121 min−1 and 12.12 min. This is better than other studied β-galactosidases. These results suggest the use of β-galactosidase from G. aparine for producing delactosed milk for the lactose-intolerant population and prebiotic synthesis. pH and optimum temperature and its activation by Ca2+ shows that it is suitable for milk processing. As a result, the enzyme was stable even at higher temperatures and had the potential to undergo catalysis, making it economically viable, primarily in the food and pharmaceutical industries. The present study is aimed to purification and thermodynamic characterization of enzyme to assess in vitro efficacy of β-galactosidase on MCF-7 cell line to delineate its therapeutic efficacy.

Anti-inflammatory activity and structural analysis of fucoidan extracted from Sargassum fusiforme via ESI-TOF MS spectrometry

This study used Celluclast-assisted hydrolysis and DEAE-cellulose anion-exchange chromatography to extract, purify, and assess the anti-inflammatory activity of fucoidan derived from Sargassum fusiforme. The Celluclast-assisted hydrolysate was used for obtaining ethanol-precipitated S. fusiforme fucoidans (SFF) which were rich in polysaccharides and sulfates. Five fractions obtained through DEAE-cellulose chromatography, notably SFF-F5, displayed high sulfate content (29.16 ± 0.15%). The anti-inflammatory potential of these fractions was tested in lipopolysaccharide-induced RAW 264.7 macrophages. SFF-F5 was found to significantly reduce NO production at non-toxic concentrations of 12.5, 25, and 50 μg/mL. Structural analysis using Matrix assisted laser desorption/ionization time-of flight mass spectrometry (MALDI-TOF MS) revealed repetitive patterns in the fucoidan fractions. And Electrospray ionization mass spectrometry (ESI-TOF MS) spectrum of SFF fractions identified various structures of glycans bound with fucose and sulfate (Fuc1(SO3)1, Fuc2(SO3)1, Hex5Fuc1(SO3)1). In-silico evaluation based on the detected glycan structures indicated that fucoidan fractions could inhibit the dimerization of the TLR4-MD2 complex, demonstrating their potential as anti-inflammatory agents. This result confirm that the anti-inflammatory activity varies depending on the glycan composition of fucoidan. This study provides a foundation for further exploration into the relationship between the structure and anti-inflammatory activity of S. fusiforme-derived fucoidan, facilitating its effective industrial application.

PYOCIN S5 ACTIVITY AGAINST PSEUDOMONAS SYRINGAE PHYTOPATHOGENIC STRAINS

Pseudomonas syringae is the most frequently isolated bacterial plant pathogens. Previously we revealed a number of Pseudomonas aeruginosa strains – producers of bacteriocins (pyocins) with high activity against P. syringae. However, it was unknown, which type of pyocins possesses the activity against P. syringae strains. S5 and S1 pyocin genes were observed in the genome of most active strain–producer P. aeruginosa UCM В-333. Pyocins were induced by nalidixic acid (100 μg/ml). Using DEAE-cellulose chromatography we isolated two bacteriocin groups from initial lysate. Substances from the 10th fraction had nuclease activity (pyocin S1) and the 27th fraction - pore-forming activity (pyocin S5). It was revealed that S1 pyocin didn’t lead to growth inhibition of studied bacterial plant pathogens. However pyocin S5 caused appearance of growth inhibition area with diameter 14-20 mm. So, pyocins of S1 and S5 types are active substances in P. aeruginosa UCM В-333 lysates, but only S5 pyocin possess activity against P. syringae strains. fpt A receptor gene was revealed in genomes of all analyzed cultures, including used phytopathogenic strains. This fact can explain wide spectrum of pyocin S5 activity. It is suggested that pyocin S5 can be a potential biocontrol agent against phytopathogenic P. syringae strains.

Purification and Structural Characterization of Polysaccharides from Polygonum multiflorum Thunb. and Their Immunostimulatory Activity in RAW264.7 Cells.

Polygonum multiflorum Thunb. (PM) and derived products are broadly utilized in Chinese traditional medicine. According to our previous research, PM mostly contains polysaccharides, which display a wide range of biological activities. Two water-soluble polysaccharides (PMPs-1 and PMPs-2) were obtained from PM by DEAE-Cellulose and Sephadex G-100 column chromatography. Colorimetry, HPGPC-MALLS-RID, HPLC-PDA, methylation, FT-IR, NMR, and SEM were used to characterize these polysaccharides. PMPs-1 and PMPs-2 had average molecular weights of 255.5 and 55.7 kDa, respectively. PMPs-1 consisted of Man, Glc, Gal, and Ara at 0.9:78.6:1.0:1.6 and was a glucan with → 4)-Glcp-(1 → as a backbone. Meanwhile, PMPs-2, an acidic polysaccharide, comprised Rha, GalA, Glc, Gal, and Ara at 3.2:20.3:2.7:1.0:8.3. PMPs-1 and PMPs-2 significantly improved the proliferation of RAW 264.7 cells and induced NO, TNF-α, and IL-6 release. This study reveals that these two polysaccharides can be explored as novel immunomodulators and provide a basis for further development of PM in food and pharmaceutical industries.

Structural Characterization and Immunoenhancing Properties of Polysaccharide CPTM-P1 from Taxus media.

Polysaccharides extracted from Taxus media hrough an aqueous method were further refined by removing proteins via the Sevag technique and purified by dialysis. The separation of these polysaccharides was accomplished using a DEAE-cellulose chromatog-raphy column, yielding two distinct fractions, named CPTM-P1 and CPTM-P2. Notably, CPTM-P1 emerged as the primary polysaccharide component within Taxus media. Consequently, a comprehensive analysis focusing exclusively on CPTM-P1 was undertaken. The molecular weight of CPTM-P1 was established through gel permeation chromatography (GPC), and its monosaccharide composition was deciphered using HPLC-MS. The structure was further elucidated through nuclear magnetic resonance (NMR) spectroscopy. The molecular weight of CPTM-P1 was determined to be 968.7 kDa. The monosaccharide composition consisted of galactose (Gal), arabinose (Ara), galacturonic acid (Gal-UA), glucose (Glc), rhamnose (Rha), xylose (Xyl), mannose (Man), fucose (Fuc), glucuronic acid (Glc-UA), and ribose (Rib). The proportional distribution of these components was 30.53%, 22.00%, 5.63%, 11.67%, 11.93%, 1.69%, 8.50%, 1.23%, 5.63%, and 1.17%, respectively. This confirmed CPTM-P1 as an acidic heteropolysaccharide with a glycuronic acid backbone. Moreover, CPTM-P1 showed immunoenhancing properties, effectively augmenting the secretion of nitric oxide and cytokines (TNF-α, IL-1β, and IL-6). Additionally, it significantly enhances the phagocytic capacity of RAW264.7 cells. These findings underscore the potential application of these polysaccharides in functional foods and pharmaceuticals, providing a solid scientific basis for further exploration and utilization of Taxus media polysaccharides.

XQZ3, a Chlorella pyrenoidosa polysaccharide suppresses cancer progression by restraining mitochondrial bioenergetics via HSP90/AKT signaling pathway

The mitochondria are known to exert significant influence on various aspects of cancer cell physiology. The suppression of mitochondrial function represents a novel avenue for the advancement of anti-cancer pharmaceuticals. The heat shock protein HSP90 functions as a versatile regulator of mitochondrial metabolism in cancer cells, rendering as a promising target for anticancer interventions. In this work, a novel acid polysaccharide named as XQZ3 was extracted from Chlorella pyrenoidosa and purified by DEAE-cellulose and gel-filtration chromatography. The structural characteristic of XQZ3 was evaluated by monosaccharides composition, methylation analysis, TEM, FT-IR, and 2D-NMR. It was found that XQZ3 with a molecular weight of 29.13 kDa was a complex branched polysaccharide with a backbone mainly composed of galactose and mannose. It exhibited good antitumor activity in vitro and in vivo by patient-derived 3D organoid models and patient-derived xenografts models. The mechanistic investigations revealed that XQZ3 specifically interacted with HSP90, impeding the activation of the HSP90/AKT/mTOR signaling cascade. This, in turn, led to the induction of mitochondrial dysfunction, autophagy, and apoptosis, ultimately resulting in the demise of cancer cells due to nutrient deprivation. This study offers a comprehensive theoretical foundation for the advancement of XQZ3, a novel polysaccharide inhibitor targeting HSP90, with potential as an effective therapeutic agent against cancer.

In vitro study on antioxidant and lipid-lowering activities of tobacco polysaccharides

Tobacco polysaccharides were extracted by hot water extraction, and purified and separated using DEAE-52 cellulose chromatography columns, and three purified polysaccharide fractions, YCT-1, YCT-2, and YCT-3, were finally obtained. The physicochemical properties of the three fractions were analyzed by ultraviolet spectroscopy, high-performance liquid chromatography and high-performance gel chromatography. The in vitro antioxidant activity of tobacco polysaccharides was compared among different fractions by using DPPH radical, hydroxyl radical scavenging assay and potassium ferricyanide method. The in vitro hypoglycemic activity was compared using α-amylase and α-glucosidase activity inhibition assay. And the in vitro hypolipidemic activity were investigated by using pancreatic lipase activity inhibition assay and HepG-2 intracellular lipid accumulation assay. All the results showed that the constituent monosaccharides of the three tobacco polysaccharide fractions were similar, but the molar percentages of each monosaccharide were different. The average molecular weights of the three components were 27,727 Da, 27,587 Da, and 66,517 Da, respectively, and the scavenging activities on DPPH radicals and hydroxyl radicals were at a high level with good quantitative-effect relationships. The reducing power were much lower than that of the positive control VC, and the three polysaccharide fractions had a weak inhibitory ability on α-amylase activity, but showed excellent inhibitory ability on α-glucosidase and pancreatic lipase activity. In addition, the results of cellular experiments showed that all three fractions were able to inhibit lipid over-accumulation in HepG-2 cells by increasing the mRNA expression levels of PPAR-α, CPT-1A, and CYP7A1 genes, and the tobacco polysaccharide YCT-3 showed the best effect. The mechanism by which YCT-3 ameliorated the over-accumulation of intracellular lipids in HepG-2 cells was found to be related to its influence on the expression of miR-155-3p and miR-17-3p in the exosomes of HepG-2 cells.Graphical

A novel heteropolysaccharide isolated from custard apple pulp and its immunomodulatory activity in mouse macrophages and dendritic cells

In this study, a novel heteropolysaccharide (ASPA80-1) with an average molecular weight of 5.48 × 104 Da was isolated and structurally elucidated from custard apple pulp (Annona squamosa) through DEAE-cellulose, Sephadex G-100 and Sephacryl S-300 HR chromatography and spectral analysis. ASPA80-1 is a water-soluble polysaccharide and it is a polymer consisting of predominant amounts of (1 → 3)-linked-L-arabinose (Ara) residues, small amounts of (1 → 6)-linked-D-galactose (Gal), (1 → 3,5)-linked-L-arabinose (Ara) residues and terminal linked-L-arabinose (Ara) residues, trace amount of (1 → 4)-linked-D-glucose (Glc) residues and (1 → 2)-linked-L-rhamnose (Rham) residues. ASPA80-1 showed significant effect on antigen-presenting cells (APCs) activation. On the one hand, ASPA80-1 activated RAW264.7 macrophage cells by inducing morphology change, enhancing phagocytic ability, increasing nitric oxide (NO) secretion and promoting expression of major histocompatibility complex class II (MHC II) and cluster of differentiation 86 (CD 86). On the other hand, ASPA80-1 promoted the maturation of dendritic cells (DCs) by inducing longer dendrites, decreasing phagocytic ability and increasing MHC II and CD86 expression. Furthermore, mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) signaling pathways were activated after the intervention of ASPA80-1 on RAW264.7 cells or DCs. Thus, the novel heteropolysaccharide ASPA80-1 has the potential to be used as an immunoenhancing component in functional foods.

Structural Characterization and Immunobiological Activity of Polysaccharides from Astragalus Oyster Mushroom.

When added to mushroom growing substrates, edible and medicinal herbs affect the mushrooms' nutritional and medicinal value. In this study, polysaccharides (P0OP-I and P15OP-I) were extracted and purified from oyster mushrooms grown on substrates supplemented with 0% and 15% Astragalus roots (P0 and P15), respectively, and their chemical structure and immunobiological activities were compared. P15OP-I and P0OP-I were extracted using ultrasound-assisted hot water and deproteinized with the Sevage method, depigmented with 30% H2O2, desalted with dialysis, and purified using DEAE-52 cellulose and Sephadex G-100 dextran column chromatography. The molecular weight of P0OP-I and P15OP-I was 21,706.96 and 20,172.65 Da, respectively. Both were composed of monosaccharides D-mannose, galacturonic acid, D-glucose, D-galactose, and L-arabinose but in different molar ratios, and both were connected by a pyranoside linkage. P15OP-I consisted of higher contents of mannose, glucose, galactose and arabinose and lower content of galacturonic acid as compared to P0OP-I. Both P0OP-I and P15OP-I induced NO and TNF-α production but did not show cytotoxic effect or induce ROS generation in RAW264.7 cells. P15OP-I showed a stronger ability to promote NO and TNF-α production relative to P0OP-I. In vitro experiments showed that the immunomodulatory activity of P0OP-I and P15OP-I in RAW264.7 macrophages were mediated by the JNK/MAPK, Erk/MAPK, and NF-κB signaling pathways. The results would be helpful for elucidation of the health promoting mechanism of Astragalus oyster mushrooms as a source of neutraceuticals.

Biochemical characterization of a purified arginase from the gut of Oryctes rhinoceros larvae

Arginase acts as a mediator in the final phase of the urea cycle, protecting against excessive ammonia under homeostatic conditions by producing L-ornithine and urea. In this study, the physicochemical properties of Oryctes rhinoceros Larva arginase were investigated for biochemical comparison with its well-studied terrestrial mammalian. Oryctes rhinoceros larva arginase was isolated and purified to homogeneity. The purification procedure involved ion-exchange chromatography on DEAE-cellulose and affinity chromatography on reactive Blue 2-agarose. The pure enzyme had a specific activity of 38.7 U/mg, a purification fold of 63.4, and a percentage yield of 16.5%. O. rhinoceros gut arginase had a native and subunit molecular weight of 82 and 45 kDa, respectively and the Km and Vmax were 11.25 mM and 13.055 μmmol/min/mL, respectively. The activity was optimum at 60 °C m max (pH 8). The enzyme retained more than 60% of its activity at 50 °C for 60 min. The inhibition study on the enzyme showed that cations salts (CaCl2 , BaCl2 , HgCl2 , and SnCl2 ) enhanced the enzyme activity at 1 mM concentration except for SnCl2 . EDTA, a chelating compound, strongly inhibited the activity of the enzyme. The 2 effect of different amino acids on activity showed that L-valine, L-serine, L- aspartic acid, and L-glutamic acid had a moderate inhibitory (60%, 63%, 65.4%, 69.1%, respectively) effect on the enzyme activity. The study concluded that there were similarities between O. rhinoceros larva arginase and those of other ureotelic animals in terms of kinetics and physicochemical properties.

Enhanced α-glucosidase inhibition activity of exopolysaccharides fractions from mycelium of Inonotus obliquus under addition of birch sawdust lignocellulose component

The addition of lignocellulose in the submerged cultivation of Inonotus obliquus has been considered a feasible alternative to improvement the bioactivity of exopolysaccharides. To investigate the key components of lignocellulose in the synthesis of active polysaccharide by I. obliquus, cellulose and hemicellulose polysaccharides were isolated from birch sawdust and their effects on the production and activity of exopolysaccharides from I. obliquus were evaluated in the present study. Results showed that α-glucosidase inhibition rates of exopolysaccharides in two cellulose polysaccharide groups (C-36 and C-48) and water extracted hemicellulose polysaccharide group (H-0) were significantly increased by 3.44–4.09 times compared with control group. Three polysaccharides fractions (DPC36–1, DPC48–1 and DPH0–1) in above three groups were isolated from I. obliquus using DEAE-52 cellulose chromatography, which were composed of glucose with molecular weight distribution ranging from 13.94 kDa to 74.48 kDa. These results might stimulate the development of mycelium polysaccharides from I. oblqiuus as alternative anti-diabetes agents and provide important insights into the relationship of structure-bioactivity of fungal polysaccharides with cellulose and hemicellulose as carbon source donors.

Physicochemical properties and anti-inflammatory and immunomodulatory effects of Shengfupian polysaccharides

In this study, the crude polysaccharides was extracted from Shengfupian and purified by Sevag deproteinization. Then, the purified neutral polysaccharide fragment was obtained by the DEAE-52 cellulose chromatography column and Sephadex G-100 co-lumn. The structure of polysaccharides was characterized by ultraviolet spectroscopy, infrared spectroscopy, ion chromatography, and gel permeation chromatography. To investigate the anti-inflammatory activity of Shengfupian polysaccharides, LPS was used to induce inflammation in RAW264.7 cells. The expression of the CD86 antibody on surface of M1 cells, the function of macrophages, and the content of NO and IL-6 in the supernatant were examined. An immunodepression model of H22 tumor-bearing mice was established, and the immunomodulatory activity of Shengfupian polysaccharides was evaluated based on the tumor inhibition rate, immune organ index and function, and serum cytokine levels. Research indicated that Shengfupian polysaccharides(80 251 Da) was composed of arabinose, galactose, glucose, and fructose with molar ratio of 0.004∶0.018∶0.913∶0.065. It was smooth and lumpy under the scanning electron microscope. In the concentration range of 25-200 μg·mL~(-1), Shengfupian polysaccharides exhibited little or no toxicity to RAW264.7 cells and could inhibit the polarization of cells to the M1 type and reduce the content of NO and IL-6 in the cell supernatant. It could suppress the phagocytosis of cells at the concentration of 25 μg·mL~(-1), while enhancing the phagocytosis of RAW264.7 cells within the concentration range of 100-200 μg·mL~(-1). The 200 mg·kg~(-1) Shengfupian polysaccharides could alleviate the spleen injury caused by cyclophosphamide, increase the levels of IL-1β and IL-6, and decrease the level of TNF-α in the serum of mice. In conclusion, Shengfupian polysaccharides has anti-inflammatory effect and weak immunomodulatory effect, which may the material basis of Aconm Lateralis Radix Praeparaia for dispelling cold and relieving pain.

Purified cellulase-mediated simultaneous sugar utilization by Bacillus albus isolated from Similipal, Odisha, India.

Among 24 isolated cellulolytic bacteria from Similipal Biosphere Reserve, the most efficient isolate was recognized as a strain of Bacillus albus. This strain of B. albus was evaluated for cellulase production and the cellulase activity was measured in submerged fermentation using substrate carboxymethyl cellulose (CMC). Different nutritional (carbon, nitrogen, and metal-ion sources) and physical variables (pH, temperature, substrate concentration, and incubation time) during the growth of B. albus were optimized to obtain maximum cellulase activity. The highest cellulase activity of 5.79 U/mL for B. albus was observed at pH 6.75, temperature 37.5°C, CMC concentration 8.5 g/L, and 42 h incubation time. Further, supplementation of glucose as a subsidiary carbon source, yeast extract, peptone as nitrogen sources, and MgSO4 and MnSO4 as metal-ion sources enhance the cellulase activity of B. albus. The purified enzyme was reported to have a molecular weight of ∼54 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A zymogram analysis evidenced the cellulase activity of the purified enzyme fractions obtained from diethylaminoethyl cellulose chromatography. The purified cellulase was reported to have an optimum pH and temperature of 7.0°C and 50°C, respectively with a capacity of retaining its 60% residual activity within pH 6.0-8.0 and temperature 30-40°C, respectively. The metal ions, K+ and Na+ were the activators, while Pb2+ and Hg2+ were the inhibitors for the purified cellulase. The purified cellulase showed Km and Vmax values of 0.38 M and 8.19 U/mL, respectively, in presence of the substrate CMC and also simultaneous consumption of both hexose and pentose sugars.

Extraction and Compositional Analysis of Anthocyanins and Polysaccharides from Aronia Melanocarpa Fruit

We have examined the basic composition of anthocyanins and polysaccharides, the active components in Aronia melanocarpa fruit. To this end, microwave-assisted ethanol extraction and cellulase enzymatic hydrolysis were employed to extract anthocyanins and crude polysaccharides, respectively. Aronia melanocarpa fruit polysaccharides were separated and purified by DEAE-52 cellulose and Sephadex G-100 glucan gel column chromatography, followed by high-performance liquid chromatography. The results demonstrated that the optimal extraction parameters of anthocyanin from Aronia melanocarpa fruit were a 60% ethanol volume fraction, a liquid-to-solid ratio of 30:1, a microwave time of 60 s, and a microwave power of 210 W. The extracted anthocyanins reached 200.47 mg/100 g, while the yield of polysaccharides was 3.82%. Anthocyanins in Aronia melanocarpa fruit were mainly composed of cyanidin, delphinidin, and mallow pigment, among which cyanidin was the main component, with a content as high as 90.79%. There were two distinct fractions of polysaccharides. The fraction-I polysaccharides were mainly composed of rhamnose, galacturonic acid, glucose, galactose, and arabinose, with a molar ratio of 8.80: 8.97: 2.38: 27.46: 47.94. In contrast, the fraction-II polysaccharides were primarily composed of rhamnose, glucuronic acid, galactose, xylose, and arabinose, with a molar ratio of 1.67: 9.22: 8.47: 70.69: 6.16. The results of this study can provide a reference for the in-depth development of Aronia melanocarpa resources

Simultaneous production of N-acetylheparosan and recombinant chondroitin using gene-engineered Escherichia coli K5

N-Acetylheparosan and chondroitin are increasingly needed as alternative sources of animal-derived sulfated glycosaminoglycans (GAGs) and as inert substances in medical devices and pharmaceuticals. The N-acetylheparosan productivity of E. coli K5 has achieved levels of industrial applications, whereas E.coli K4 produces a relatively lower amount of fructosylated chondroitin. In this study, the K5 strain was gene-engineered to co-express K4-derived, chondroitin-synthetic genes, namely kfoA and kfoC. The productivities of total GAG and chondroitin in batch culture were 1.2 g/L and 1.0 g/L respectively, which were comparable to the productivity of N-acetylheparosan in the wild K5 strain (0.6–1.2 g/L). The total GAG of the recombinant K5 was partially purified by DEAE-cellulose chromatography and was subjected to degradation tests with specific GAG-degrading enzymes combined with HPLC and 1H NMR analyses. The results indicated that the recombinant K5 simultaneously produced both 100-kDa chondroitin and 45-kDa N-acetylheparosan at a weight ratio of approximately 4:1. The content of chondroitin in total GAG partially purified was 73.2%. The molecular weight of recombinant chondroitin (100 kDa) was 5–10 times higher than that of commercially available chondroitin sulfate. These results indicated that the recombinant K5 strain acquired the chondroitin-producing ability without altering the total GAG productivity of the host.

A comparative study of pectic polysaccharides from fresh and dried Dendrobium officinale based on their structural properties and hepatoprotection in alcoholic liver damaged mice.

In this study, two pectic polysaccharides from fresh and dried Dendrobium officinale, namely FDP and DDP, were obtained by sour-water extraction, ethanol precipitation and further purification with DEAE cellulose-52 and Sephadex G-100 column chromatography. FDP/DDP had eight similar glycosidic linkages including 1,4-linked-GlcAp, 1,4- and 1,3,4-linked-GalAp, 1,3,4- and T-linked-Glcp, 1,6- and T-linked-Galp, T-linked-Galp and T-linked-Xylp. Besides, FDP was marked by 1,6-, 1,2,6-linked-Manp and 1,2,4-, 1,2-linked-Rhap, and DDP consisted of unique 1,6-linked-GlcAp and 1,3,6-Manp. FDP with a molecular weight of 14.8 kDa generally showed stronger scavenging capacity against DPPH, ABTS and hydroxyl radicals than DDP (p < 0.05). Pretreatment with FDP/DDP alleviated the alcohol-induced liver injury in mice, and their serum aminotransferase and triglyceride levels were 10.3%-57.8% lower than those of the model group (MG). Meanwhile, the FDP/DDP-M and FDP/DDP-H groups (200 and 300 mg kg-1) displayed a remarkable increase in antioxidant enzyme activities and significant reduction in inflammatory cytokine levels in comparison with the MG. Further analysis revealed that FDP-treated mice generally exhibited lower transaminase levels and inflammatory cytokine expression as well as higher antioxidant enzyme activities than DDP-treated ones. The FDP-H group showed significant restoration, which was slightly less than or almost comparable to that of the bifendate-fed positive control. The above results indicate that D. officinale pectin can attenuate oxidative stress and inflammatory cytokine response, and ultimately ameliorate liver injury, and "fresh" pectin with specific structural characteristics is expected to be more promising as hepatoprotective food.

Isolation, Purification, and Antioxidant Activities of Polysaccharides from Choerospondias axillaris Leaves.

The extraction, characterization and antioxidant activity of polysaccharides from Choerospondias axillaris leaves were investigated in the present study. Two purified polysaccharide fractions, CALP-1 and CALP-2, were isolated from crude Choerospondias axillaris leaf polysaccharides (CALP) by DEAE-52 cellulose chromatography and Sephadex G-100 column chromatography. The characteristics of CAL-1 and CALP-2 were determined by using High-performance Gel Permeation Chromatography (HPGPC), High-Performance Anion-Exchange Chromatography, HPAEC (HPAEC-PAD) and Fourier transform infrared spectroscopy (FTIR). CALP-1 with molecular weight of 11.20 KDa was comprised of Rhamnose, Arabinose, Galactose, Glucose, Xylose, Mannose and galacturonic acid in a molar ratio of 5.16:2.31:5.50:27.18:1.00:0.76:1.07. CAL-2 with molecular weight of 8.03 KDa consisted of Rhamnose, Arabinose, Galactose, Glucose, and galacturonic acid at a ratio of 1.38:3.63:18.84:8.28:1.45. FTIR revealed that CALP-1 and CALP-2 were acidic polysaccharides. The antioxidant activity of crude CALP, CALP-1 and CALP-2 was evaluated in vitro. The fraction CALP-2 was demonstrated to be of polysaccharide nature containing a large percentage of Galactose but no Xylose and Mannose. The antioxidant activity assays showed that CALP-1 and CALP-2 exhibited antioxidant and scavenging activities on hydroxyl and DPPH radicals in vitro. Compared with pure polysaccharide, crude CALP exhibited stronger anti-oxidant activities. These results will provide a better understanding of Choerospondias axillaris leaf polysaccharide and promote the potential applications of Choerospondias axillaris leaf polysaccharide in the pharmacological field and as a natural antioxidant.

Screening of peroxidase from various vegetables sources, partial purification, some biochemical properties and its antimicrobial activity

The peroxidase enzyme was extracted from four different vegetables which were radish, turnip, cabbage and green pepper. The extracted enzymes were partially purified by ammonium sulphate and DEAE cellulose (Diethylaminoethyl cellulose) chromatography. The effect of amount of guaiacol, hydrogen peroxide, pH and temperature were investigated. The optimum pH for peroxidase activity was in the range (6.5-7.0), while optimum temperature for extracted enzymes was 30⁰c. The extracted enzyme from selected vegetables was tested for their antimicrobial activity against different species of bacteria. The enzymes show weak antibacterial activity towards the tested species of bacteria.

Purification, structural characterization and immunological activity of Sibiraea laexigata (L.) Maxim polysaccharide.

Sibiraea laexigata (L.) Maxim (SLM) has been used as an herbal tea for treating stomach discomfort and indigestion for a long time in china. Polysaccharides have been identified as one of the major bioactive compounds in the SLM. In the present paper, ultrasonic-assisted enzymatic extraction (UAEE) method was employed in polysaccharides extraction derived from SLM using polyethylene glycol (PEG) as extraction solvent, two SLM polysaccharides (SLMPs) fractions (SLMPs-1-1 and SLMPs-2-1) were purified by DEAE Cellulose-52 and Sephadex G-100 chromatography in sequence. Then, the preliminarily structure of the two factions were characterized by chemical composition analysis, molecular weight measurement, UVS, HPLC-PMP, FT-IR, nuclear magnetic resonance (NMR) spectra analysis and SEM. The results showed that SLMPs-1-1 and SLMPs-2-1 with different molecular weights of 1.03 and 1.02 kDa, mainly composed of glucose (46.76 and 46.79%), respectively. The results of structural characterization from FT-IR, 1H NMR, and SEM revealed that SLMPs-1-1 and SLMPs-2-1 contained the typical pyranoid polysaccharide with α-glycosidic bond and β-glycosidic bond. Furthermore, it was found that SLMPs-1-1 could increase the levels of tumor necrosis factor-α (TNF-α) and interleukin-2 (IL-2), and alleviated the immune organs tissue damage of cyclophosphamide (Cy)-treated mice. RT-qPCR and Western-Blot analysis showed that SLMPs-1-1 could significantly up-regulated the levels of NF-κB, TLR4, which revealed that SLMPs-1-1 could participate in immunosuppressive protection of Cy-treated mice. These findings suggested that the potential of SLMPs-1-1 as an alternative immunostimulator could be used in food and pharmaceutical industries.

BIODEGRADATION OF SOME ENVIRONMENTAL POLLUTANTS BY LACCASE PRODUCED FROM TRICHODERMA HARZIANUM USING SOLID STATE FERMENTATION

Trichoderma harzianum produced 5.383 U/mg of laccase in solid state fermentation (SSF) using 10gm of wheat bran as substrate with moisture content 1:0.7 w/v, using 4 plugs×5mm as inoculums size, for 7 days at 28C°. Laccase was partially purified with 7.260 fold and yield 84.01% by ion exchange chromatography DEAE-cellulose. Partially purified laccase had an optimum pH of 4 and was stable in pH range from 3.5 to 5.5. The optimum temperature for laccase activity was 35C° and stability was range from 15 to 35C° with 1h of incubation. The ability of T. harzianum isolate to decolorize textile dyes on solid media appears completely decolorization of Blue dye with concentrations 50,100 and partially decolorized with concentration 150,200 and 250 ppm, while the Yellow dye and Red dye were completely decolorized at 50 ppm concentration. The ability of partial purified laccase to degradation of phenol was completely occurs in concentration 5, 10 but at 20, 50 and 100 ppm about 90.94%, 93.93%and 36.78% respectively, while furfural degradation at 5 and 10 ppm was about 86.84% and 68.66% respectively which detected using HPLC technique.