Production of Cell Membrane-Bound α- and β-Glucosidase by Lactobacillus acidophilus

Hydrolytic enzymes, viz. α- and β-glucosidase, were produced from indigenous isolate, Lactobacillus acidophilus, isolated from fermented Eleusine coracana. Production of these enzymes was enhanced by optimizing media using one factor at a time followed by response surface methodology. The optimized media resulted in a 2.5- and 2.1-fold increase in α- and β-glucosidase production compared with their production in basal MRS medium. Localization studies indicated 80% of the total activity to be present in the cell membrane-bound fraction. Lack of sufficient release of these enzymes using various physical, chemical, and enzymatic methods confirmed their unique characteristic of being tightly cell membrane bound. Enzyme characterization revealed that both α- and β-glucosidase exhibited optimum catalytic activity at 50 °C and pH 6.0 and 5.0, respectively. K _m and V _max of α-glucosidase were 4.31 mM and 149 μmol min⁻¹ mL⁻¹ for p-nitrophenyl-α-d-glucopyranoside as substrate and 3.8 mM and 120 μmol min⁻¹ mL⁻¹ for β-glucosidase using p-nitrophenyl-β-d-glucopyranoside as the substrate.     

Properties of <Emphasis Type="Italic">Aspergillus subolivaceus</Emphasis> free and immobilized dextranase

Aspergillus subolivaceus dextranase is immobilized on several carriers by entrapment and covalent binding with cross-linking. Dextranase immobilized on BSA with a cross-linking agent shows the highest activity and considerable immobilization yield (66.7%). The optimum pH of the immobilized enzyme is shifted to pH 6.0 as compared with the free enzyme (pH 5.5). The optimum temperature of the reaction is resulted at 60 °C for both free and immobilized enzyme. Thermal and pH stability are significantly improved by the immobilization process. The calculated K _m of the immobilized dextranase (14.24 mg mL⁻¹) is higher than that of the free dextranase (11.47 mg mL⁻¹), while V _max of the immobilized enzyme (2.80 U μg protein⁻¹) is lower than that of the free dextranase (11.75 U μg protein⁻¹). The immobilized enzyme was able to retain 76% of the initial catalytic activity after 5.0 cycles.     

Optimization and Modeling of Process Parameters for Lipase Production by Bacillus brevis

Statistical evaluation of fermentation conditions and nutritional factors by Plackett–Burman two-level factorial design followed by optimization of significant parameters using response surface methodology for lipase production by Bacillus brevis was performed in submerged batch fermentation. Temperature, glucose, and olive oil were found to be the significant factors affecting lipase production. Maximum lipase activity of 5.1 U ml⁻¹ and cell mass of 1.82 g l⁻¹ at 32 h were obtained at the optimized conditions of temperature, 33.7 °C; initial pH, 8; and speed of agitation, 100 rpm, with the medium components: olive oil, 13.73 ml l⁻¹; glucose, 13.98 g l⁻¹; peptone, 2 g l⁻¹; Tween 80, 5 ml l⁻¹; NaCl, 5 g l⁻¹; CH₃COONa, 5 g l⁻¹; KCl, 2 g l⁻¹; CaCl₂·2H₂O, 1 g l⁻¹; MnSO₄·H₂O, 0.5 g l⁻¹; FeSO₄·7H₂O, 0.1 g l⁻¹; and MgSO₄·7H₂O, 0.01 g l⁻¹. The lipase productivity and specific lipase activity were found to be 0.106 U (ml h)⁻¹ and 2.55 U mg⁻¹, respectively. Unstructured kinetic models and artificial neural network models were used to describe the lipase fermentation. The kinetic analysis of the lipase fermentation by B. brevis shows that lipase is a growth-associated product.     

Partial Purification and Characterization of Extracellular Fructofuranosidase with Transfructosylating Activity from <Emphasis Type="Italic">Candida</Emphasis> sp.

The present work was carried out with the aim to investigate some properties of an extracellular fructofuranosidase enzyme, with high transfructosylating activity, from Candida sp. LEB-I3 (Laboratory of Bioprocess Engineering, Unicamp, Brazil). The enzyme was produced through fermentation, and after cell separation from the fermented medium, the enzyme was concentrated by ethanol precipitation and than purified by anion exchange chromatography. The enzyme exhibited both fructofuranosidase (FA) and fructosyltransferase (FTA) activities on a low and high sucrose concentration. With sucrose as the substrate, the data fitted the Michaellis–Menten model for FA, showing rather a substrate inhibitory shape for fructosyltransferase activity. The K _m and v _max values were shown to be 13.4 g L⁻¹ and 21.0 μmol mL⁻¹ min⁻¹ and 25.5 g L⁻¹ and 52.5 μmol mL⁻¹ min⁻¹ for FA and FTA activities, respectively. FTA presented an inhibitory factor K _i of 729.8 g L⁻¹. The optimum conditions for FA activity were found to be pH 3.25–3.5 and temperatures around 69 °C, while for FTA, the optimum condition were 65 °C (±2 °C) and pH 4.00 (±0.25). Both activities were very stable at temperatures below 60 °C, while for FA, the best stability occurred at pH 5.0 and for FTA at pH  4.5–5.0. Despite the strong fructofuranosidase activity, causing hydrolysis of the fructooligosaccharides (FOS), the high transfructosilating activity allows a high FOS production from sucrose (44%).     

Properties of thermostable extracellular FOS-producing fructofuranosidase from <Emphasis Type="Italic">Cryptococcus</Emphasis> sp.

The present work was devoted to investigations concerning the fructooligosaccharide producing activity of Cryptococcus sp. LEB-V2 (Laboratory of Bioprocess Engineering, Unicamp, Brazil) and its extracellular fructofuranosidase. After cell separation, the enzyme was purified by ethanol precipitation and anion exchange chromatography. The enzyme showed both fructofuranosidase (FA) and fructosyl transferase (FTA) activity. With sucrose as substrate, the data failed to fit the Michaelis–Menten behaviour, showing a substrate inhibitory model. The K _m, K _i and v _max values were shown to be 64 mM, 3 M and 159.6 μmol mL⁻¹ min⁻¹ for FA and 131 mM, 1.6 M and 377.8 μmol mL⁻¹ min⁻¹ for FTA, respectively. The optimum pH and temperature were found to be around 4.0 and 65 °C, while the best stability was achieved at pH 4.5 and temperatures below 60 °C, for both the FA and FTA. Despite the strong FA activity, the high transfructosylating activity allowed for good FOS production from sucrose (35% yield).     

Analytical Methods for the Evaluation of α-<Emphasis Type="SmallCaps">l</Emphasis>-Fucosidase Activity in Sheep Milk

The lack of analytical methods for measuring the activity of highly thermolable endogenous enzymes in sheep milk is a factor that hampers the protection of typical Protected Designation of Origin (PDO) dairy products made from raw milk. In order to provide a solution, this study assesses, tests, and fully validates analytical procedures for the determination of α-l-fucosidase activity in sheep milk. While the UV–VIS method has been optimized for this matrix in order to solve clarification problems before the spectrophotometric reading, a reversed phase high-performance liquid chromatography literature method proposed for bovine milk has been successfully applied also to sheep milk. Both methods have been fully validated in terms of sensitivity, linearity, precision, and accuracy, displaying low detection and quantification limits, excellent linearity over a wide enzymatic activity interval, very good repeatability and reproducibility, and the lack of any bias. The analysis of a number of real samples of whole sheep milk has allowed the evaluation of an average value (46.78 ± 5.49 U mL⁻¹) and range (from 29.27 ± 2.60 to 72.64 ± 1.17 U mL⁻¹) of α-l-fucosidase activity. Such activity does not seem to differ substantially from those measured for bovine milk. Finally, marked seasonal variability has been observed in this preliminary dataset.     

Lipase-catalyzed acylation of <Emphasis Type="SmallCaps">l</Emphasis>-carnitine with conjugated linoleic acid in [Bmim]PF<Subscript>6</Subscript> ionic liquid

Improving significantly the acylation reaction of l-carnitine with conjugated linoleic acid catalyzed by lipase needs to select an efficient and suitable reaction medium that is not only polar but also hydrophobic. [Bmim]PF₆ which is satisfied with the above two requirements was applied as the reaction medium. The optimal reaction conditions were: Novozyme 435 as the catalyst, 5:1 of molar ratio of fatty acid to l-carnitine, 40 g L⁻¹ lipase, 0.22 a_W of initial water activity, 125 g L⁻¹ molecular sieves (they were added within 0–3 h of reaction time), 60 °C of reaction temperature, 200 rpm of rotation speed, 32 h of reaction time. The overall conversion could reach 98.27%. The conversion in [Bmim]PF₆ was 2.13 and 1.56 times higher than that in acetonitrile and in solvent-free system, respectively. The lipase in the present work could be used eight times. [Bmim]PF₆ as the reaction medium was better than acetonitrile and solvent-free system.     

Purification and application of α-galactosidase from germinating coffee beans (<Emphasis Type="Italic">Coffea arabica</Emphasis>)

The objective of this study was to prepare and purify α-galactosidase (α-Gal) from germinating coffee beans. The molecular weight of purified α-Gal from germinating coffee beans was determined to be 38.8 kDa by SDS-PAGE. Then the purified α-Gal was applied to synthesize galactosyl β-cyclodextrin using melibiose as donor and β-cyclodextrin as acceptor. Galactosyl β-cyclodextrin was isolated by preparative high-performance liquid chromatography (HPLC) and its structure was analyzed and elucidated by HPLC, fourier transform infrared spectrometer, electro spraying ionization-mass spectrometry, and ¹³C nuclear magnetic resonance. Our results strongly demonstrate that the synthesized product is a mono-6-O-α-d-galactopyranosyl-β-cyclodextrin and the purified α-galactosidase could transfer one galactosyl residue directly to the β-cyclodextrin ring and synthesize the mono-6-O-α-d-galactosyl β-cyclodextrin.     

The Influence of Oxygen Volumetric Mass Transfer Rates on Cyclodextrin Glycosyltransferase Production by Alkaliphilic Bacillus circulans in Batch and Fed-Batch Cultivations

In this work, the influence of oxygen mass transfer rates on the production of cyclodextrin glycosyltransferase (CGTase) by the alkaliphilic bacterium Bacillus circulans ATCC 21783 was investigated. Experimental design and response surface methodology were applied to optimize agitation speed and air flow rate in batch cultivations, in order to identify their significant effects and interactions with the synthesis of CGTase. Results were expressed as the volumetric mass transfer rates of oxygen (k_la, [per hour]). The maximal CGTase productivity of 155 U mL⁻¹ h⁻¹ was achieved with k_la of 48 h⁻¹. CGTase production was also studied in fed-batch cultures using the optimized parameters obtained in the batch experiments. The maximal CGTase productivity on fed-batch cultivations was 137 U mL⁻¹ h⁻¹ with feeding rates of starch at 0.17 g L⁻¹ h⁻¹.     

ACE inhibitory and antihypertensive properties of apricot almond meal hydrolysate

Apricot almond meal was hydrolyzed simultaneously with Neutrase and N120P proteases. The hydrolysate almond peptide (AP) was fractionated into three ranges of molecular weight (AP-I, >5 kDa; AP-II, 1–5 kDa; AP-III, <1 kDa) using an ultrafiltration membrane bioreactor system. The AP-III brought about a high angiotensin-I-converting enzyme (ACE) inhibitory activity with IC₅₀ value of 0.138 mg mL⁻¹ and the content of hydrophobic amino acid of AP-III was 50.08%. Lineweaver–Burk plots suggested that AP-III acts as a non-competitive inhibitor to inhibit ACE. And we evaluated the stability and in vivo antihypertensive activity of AP-III. Multiple dose oral administration (100 mg kg⁻¹ body weight (BW), 400, 800 mg kg⁻¹ BW) to spontaneously hypertensive rats led to a significant decrease in blood pressure for AP-III. Additionally, the gel filtration and RP-HPLC were used to separate ACE inhibitory peptides from the hydrolysate. The results suggested that the peptide derived from apricot almond protein may have potential applications as functional food.     

Effects of Maillard reaction conditions on the antigenicity of α-lactalbumin and β-lactoglobulin in whey protein conjugated with maltose

The effects of Maillard reaction conditions (weight ratio of protein to sugar, temperature and time) on the antigenicity of α-lactalbumin (α-LA) and β-lactoglobulin (β-LG) in conjugates of whey protein isolate (WPI) with maltose were investigated. Response surface methodology was used to establish models to predict the antigenicity of α-LA and β-LG and find an optimal reaction condition under which the antigenicity of α-LA and β-LG reduces to minimum value. Conjugating WPI with maltose was an effective way to reduce the antigenicity of α-LA and β-LG. The antigenicity of α-LA decreased from 32.25 μg mL⁻¹ to 10.91 μg mL⁻¹. And the antigenicity of β-LG decreased from 272.4 μg mL⁻¹ to 38.17 μg mL⁻¹. Temperature had the greatest effect on the antigenicity of α-LA, while weight ratio of WPI to maltose was the most significant factor on the antigenicity of β-LG.     

Determination of Histamine in Some Foods by Isotachophoretic Method with Simple Sample Preparation

A one-dimensional capillary isotachophoretic method in cationic system of the separation has been applied for histamine determination in food samples. The proposed electrolyte system consisted of 0.01 M potassium hydroxide with l-valine to pH = 9.9 as the leading electrolyte and 0.02 M 2-amino-2-hydroxymethyl-propane-1,3-diol adjusted to pH = 8.3 with 0.1 M hydrochloric acid as terminating electrolyte. Proposed method was characterized by linearity range 5–50 mg L⁻¹ and R ² = 0.9982, accuracy (recoveries ranged from 95% to 102%), detection (2.10 mg L⁻¹), and quantification (7.01 mg L⁻¹) limits. The sample preparation for proposed electrophoretic method included only simple extraction with trichloroacetic acid with filtration and derivatisation stage are avoided. The histamine concentration was determined in meat (turkey, chicken, beef and pork) and meat products (ripened sausage and dry-cured ham), fish (smoked salmon and mackerel), and different kind of mildew and mold ripened cheeses samples. The histamine content ranged from not detected level for fresh meat to 29.63 mg 100 g⁻¹ for cheese samples. The reversed phase HPLC was applied as reference method and the F-Snedecor test and the t test were employed to compare the precision and accuracy of the both methods. Positive correlations were found between the two analytical methods for histamine determination in food products. The obtained results indicate that the proposed electrophoretic method is simple, precise, accurate, and convenient.     

Purification and properties of a heat-stable enzyme of <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> Rm12 from raw milk

Pseudomonas fluorescens Rm12 is a kind of Psychrotrophic bacteria growing in cold raw milk. It produced an extracellular heat resistant protease with an estimated molecular weight of 45 kDa by size exclusion chromatography and SDS-PAGE under both reducing and non-reducing conditions. The enzyme, designated Ht13, was purified to electrophoretic homogeneity from the culture supernatant by sequentially using ammonium sulfate precipitation, ion-exchange chromatography, hydrophobic chromatography and size exclusion chromatography. The specific activity of the enzyme increased 115.5-folds. The optimum pH value and temperature of Ht13 were 7.5 and 40 °C, respectively. Based on its biochemical characteristics, Ht13 can be included in the group of metalloproteases, which was inhibited by 1, 10-phenanthroline and EDTA but not by pepstatin A, chymostatin, STI, E-64, BBI, PMSF and pAPMSF. Mn²⁺ has positive effect on activity and can increased the heat resistance capability, while Ca²⁺ had a negligible effect. For the hydrolysis of azocasein, the Km was 0.012 mg mL⁻¹. The enzyme showed typical heat-stable behavior. After treatment of 160 °C 20 s, the residual activity was 9%. The half-life of the enzyme at 160 °C in buffer with Mn²⁺ was approximately 12 s. Among several main milk proteins, Ht13 can cleave α_s-casein, β-casein and κ-casein. The sequence of 1st–16th amino acids of N-terminal was MSKVKDKAIVSAAQAS, which was same as those proteases excreted from some other P. fluorescens. However, their molecular weights, the activation ion and amino acid composition were different, suggesting Ht13 from P. fluorescens Rm12 is a novel protease.     

Preparation of immobilized Trametes pubescens laccase on a cryogel-type polymeric carrier and application of the biocatalyst to apple juice phenolic compounds oxidation

A new biocatalyst was prepared by the immobilization of a Trametes pubescens laccase, into a wide-pore poly(vinyl alcohol) cryogel. The known enzyme was produced and purified by the previously described procedure. The resulted laccase (yield 40%) has an activity of 46.4 U mg⁻¹ and 12.51 mg mL⁻¹ protein content. The enzyme was subsequently immobilized in a functionalized macroporous cryogel beads by a covalent immobilization technique. The time dependence of the immobilization process and the enzyme loading of the carrier material (5.2 mg g⁻¹ cryogel) were determined by measuring the decrease of protein amount in the enzyme solution. In conversion experiments, a higher stability of the immobilized biocatalyst compared to the free enzyme was evidenced. Steady-state kinetic characterization of four phenols (catechol, caffeic and chlorogenic acids, and catechin) has been performed with free and immobilized laccase, the catalytic parameters being determined and compared. The effect of both laccases (free and immobilized) on the phenol content of retailed apple juice samples, having the same initial composition, was also investigated by working in batch conversion. The variation in phenolic compound content has been compared with that of an untreated apple juice sample having initially the same content of phenolic compounds. A number of advantages resulted in using the immobilized laccase for the apple juice treatment (conservation to some extent of enzyme activity, higher content of phenols preserved, easy separation of the enzyme from the apple juice, therefore avoiding the possible unhealthy effects due to the remaining protein, etc.).     

Is there a direct relationship between oral astringency and human salivary protein binding?

For decades, it is believed that astringency is due to the polyphenol-induced complexation of proline-rich salivary proteins in the oral cavity. In order to compare for the first time the human sensory threshold concentrations and the salivary protein binding activity of a series of astringent stimuli, human saliva protein was incubated for 5 min at 37 °C in the presence of astringent food-derived compounds and, after micro-centrifugation, the amount of the target molecules in the supernatant was quantitatively determined by HPLC-UV/Vis. Significant protein binding was observed for (−)-epigallocatechin-3-gallate, (−)-gallocatechin-3-gallate, (+)-gallocatechin, and (−)-catechin-3-gallate, all of which containing at least one galloyl moiety in the molecule and exhibiting rather high sensory thresholds of more than 200 μmol/L. In comparison, (+)-catechin and procyanidin B2, both lacking in any galloyl function, showed only comparatively low binding activity and, most interestingly, quercetin-3-O-α-l-rhamnopyranosyl-(1 → 6)-β-d-glucopyranoside and 3-carboxymethyl-indole-1-N-β-d-glucopyranoside did not show any protein binding at all, although the later N- and O-glycosides exhibited extraordinarily low sensory threshold concentrations of less than 0.001 and 0.0003 μmol/L, respectively. The data give some first evidence that the quantity of the non-bound, “free” astringent stimulus in the saliva liquid might be more closely related to the sensory perception of astringency than the amount complexed or precipitated by proteins. It is therefore questionable as to whether oral perception of astringency is related to the complexation and/or precipitation of salivary proteins.     

Isolation and identification of Di-D-fructose dianhydrides resulting from heat-induced degradation of inulin

Dry heating of inulin up to 200 °C for 30 min resulted in a wide range of degradation products, from which five quantitatively dominating compounds were isolated using flash chromatography and semi-preparative HPLC–RI. It was possible to identify four different DFDAs (di-d-fructose dianhydrides) by means of one- and two-dimensional NMR, namely DFA III (α-D-Fruf-1,2′:2,3′-β-D-Fruf), DFA I (α-D-Fruf-1,2′;2,1′-β-D-Fruf), DFA VII (α-D-Fruf-1,2′;2,1′-α-D-Fruf), and β-D-Fruf-1,2′;2,1′-β-D-Fruf. For the fifth compound, the structure of the difructan α-D-Fruf-1,2′-β-D-Fruf was proposed. Using high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC–PAD), it was possible to quantify those isolated DFDAs in inulin caramels to a total amount of up to 25%, indicating that dry heating of inulin may be a suitable tool to enrich caramels with DFDAs.     

Quantitation of the homocysteine content in wine

Alcohol consumption has been previously shown to correlate with elevated plasma homocysteine levels, but investigations have not been carried out on the possible availability of this compound in alcoholic beverages such as wine or spirits. Therefore, in this study we investigated the levels of homocysteine in various Bulgarian wines. A total of 36 different Bulgarian wines with known origins were studied. The measured values were in the range of 0.09–0.64 mg l⁻¹ for the tested white wines and in the range of 0.10–1.37 mg l⁻¹ for the red wines. The method used for homocysteine determination was based on RP-HPLC with fluorescent detection after derivatization with N-(2-acridonyl)maleimide. The method was linear in the range of 0.0070–1.35 mg l⁻¹ homocysteine and showed low limits of detection and quantification (LOD = 6 fmol, LOQ = 68 fmol). The within-run precision expressed as relative standard deviation (RSD, %) was 2.2–2.4% and the between-run precision was 2.6–3.9%. Enzyme immunoassay and LC-MS ⁿ analyses were used for confirmation of presence of homocysteine in wine.     

Multiple α-galactosidases from <Emphasis Type="Italic">Aspergillus foetidus</Emphasis> ZU-G1: purification,characterization and application in soybean milk hydrolysis

α-Galactosidase had applied in food and feed industries for hydrolyzing raffinose series oligosaccharides (RO) that are the factors primarily responsible for flatulence upon ingestion of soybean-derived products. The objective of the current work was to purify the α-galactosidase of Aspergillus foetidus ZU-G1 and compared the biochemical and hydrolytic properties of three major α-galactosidase forms (α-gal I, α-gal II and α-gal III). The molecular mass of the purified enzyme as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was 106.3, 49.7 and 109.9 kDa, respectively. Its optimum reaction temperature was 60 °C and stable below 50 °C. The optimum pH of α-gal I and α-gal III was 5.0 and α-gal II was 4.0. Under 28 °C conditions for 24 h, α-gal I was stable at pH 4.0, α-gal II was stable at pH 6.0, and α-gal III was pH 5.0. α-Galactosidase was completely inhibited by Ag⁺. CuSO₄·5H₂O and SDS were powerful inhibitors of α-gal I and α-gal III but had little effect to α-gal II. EDTA did not strongly affect α-gal I and α-gal III, while strongly affect α-gal II. CaCl₂·2H₂O, MgSO₄·7H₂O and MnSO₄·7H₂O were activation for α-gal I, α-gal II and α-gal III. No significant inhibition of enzymes activity was observed in the presence of raffinose, lactose as well as other sugars tested. Synthetic substrate p-nitrophenyl-α-d-galactopyranoside was not preferentially hydrolyzed than natural substrates, such as melibiose, stachyose and raffinose. Under 40 and 50 °C incubation for 1–5 h, the stachyose of soybean milk was degraded by α-gal I, α-gal II and α-gal III and strongly hydrolyzed by α-gal II, and the raffinose of soybean milk was completely hydrolyzed by α-gal II and weakly hydrolyzed by α-gal I and α-gal III. The distinct hydrolytic and biochemical properties of α-gal I, α-gal II and α-gal III further signify the α-galactosidase of A. foetidus ZU-G1 was propitious to soybean milk and related food industry.     

Optimization of the culture medium composition using response surface methodology for new recombinant cyprosin B production in bioreactor for cheese production

The optimization of culture medium composition was carried out for improvement the recombinant cyprosin B production, an enzyme with high milk-clotting activity. Response surface methodology (RSM) was applied to evaluate the effect of variables namely glucose, yeast extract (YE) and bactopeptone present in the culture medium, used for recombinant cyprosin B production by transformed Saccharomyces cerevisiae BJ1991 strain in shake-flask and bioreactor culture conditions. The central composite experimental design (CCD) was adopted to derive a statistical model for optimizing the composition of the fermentation medium. The optimal concentration estimated for each variable related to a theoretical maximum of cyprosin B activity (488 U mL⁻¹) was 30 g L⁻¹ glucose, 15 g L⁻¹ YE and 27 g L⁻¹ bactopeptone. The optimized medium composition, based on empirical model, led to a cyprosin B activity of 519 U mL⁻¹, which corresponds to an increase of 46%. The fermentation using optimized culture medium in a 5-L bioreactor allowed a significant increase in biomass (82%) and recombinant cyprosin B production (139%). The improvement in the recombinant cyprosin B production after optimization process can be considered adequate for large-scale applications, and the clotting activity of cyprosin B account for their use in industrial cheese making.     

Chemical characterization and antioxidant activity of an exopolysaccharide fraction isolated from <Emphasis Type="Italic">Bifidobacterium animalis</Emphasis> RH

The major exopolysaccharide fraction (EPSa) of Bifidobacterium animalis RH was purified to illustrate the chemical characterization and the antioxidant activity in vitro. The molecular weight of EPSa was 2.31 × 10⁴ Da determined by the gel permeation chromatography (GPC). Fourier-transform infrared spectra (FT-IR) and the monosaccharide analysis of high-pH anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) showed that it was a heteropolysaccharide mainly composed of d-glucose, d-mannose, d-galactose, l-arabinose, d-fructose, and l-rhamnose in molar ratios of 43:34:18:4:1:1. In antioxidant assays, the EPSa had a higher activity for antilipid peroxidation and a stronger scavenging activity of hydroxyl radical and superoxide radical than ascorbic acid. It also had a similar α,α-diphenyl-β-picrylhydrazyl (DPPH) radical-scavenging activity with ascorbic acid at low concentration and had concentration-dependent inhibitory ability on erythrocyte hemolysis. These results indicated that the EPSa fraction had strong antioxidant activities and could be explored as novel potential antioxidants.