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1.
The effects of several conditions on the amounts and compositions of aggregates formed in mixtures of whey protein hydrolysate, made with Bacillus licheniformis protease, and whey protein isolate were investigated using response surface methodology. Next, the peptides present in the aggregates were separated from the intact protein and identified with liquid chromatography-mass spectrometry. Increasing both temperature and ionic strength increased the amounts of both intact protein and peptides in the aggregates. There was an optimal amount of added intact WPI that could aggregate with peptides, yielding a maximal amount of aggregated material in which the peptide/protein molar ratio was around 6. Under all conditions applied, the same peptides were observed in the protein-peptide aggregates formed. The dominant peptides were beta-lg AB [f1-45], beta-lg AB [f90-108], and alpha-la [f50-113]. It was hypothesized that peptides could form a kind of glue network that can include beta-lactoglobulin via hydrophobic interactions at the hydrophobic binding sites at the surface of the protein. 相似文献
2.
In a previous study, peptides aggregating at pH 7.0 derived from a whey protein hydrolysate made with Bacillus licheniformis protease were fractionated and identified. The objective of the present work was to investigate the solubility of the fractionated aggregating peptides, as a function of concentration, and their aggregating capacities toward added intact proteins. The amount of aggregated material and the composition of the aggregates obtained were measured by nitrogen concentration and size exclusion chromatography, respectively. The results showed that of the four fractions obtained from the aggregating peptides, two were insoluble, while the other two consisted of 1:1 mixture of low and high solubility peptides. Therefore, insoluble peptides coaggregated, assumedly via hydrophobic interactions, other relatively more soluble peptides. It was also shown that aggregating peptides could aggregate intact protein nonspecifically since the same peptides were involved in the aggregation of whey proteins, beta-casein, and bovine serum albumin. Both insoluble and partly insoluble peptides were required for the aggregation of intact protein. These results are of interest for the applications of protein hydrolysates, as mixtures of intact protein and peptides are often present in these applications. 相似文献
3.
P W Caessens H Gruppen C J Slangen S Visser A G Voragen 《Journal of agricultural and food chemistry》1999,47(5):1856-1862
To investigate structure-function relationships with regard to emulsion-stabilizing properties, peptides from bovine beta-casein (betaCN), obtained by plasmin hydrolysis and fractionation of the hydrolysate, were isolated and identified on the basis of their masses determined by electrospray ionization mass spectrometry, the primary structure of the intact protein, and the known specificity of the enzyme. An amphipathic peptide fraction was fractionated further by ion-exchange chromatography and subsequent hydrophobic interaction chromatography resulting in the components betaCN[f 1-105/107] and betaCN[f 29-105/107]. The latter peptides had poor emulsion-stabilizing properties compared to the former ones, and the stability of an emulsion formed with betaCN[f 29-105/107] was also more sensitive to hydrophobic impurities than that of an emulsion formed with betaCN[f 1-105/107]. The highly charged N-terminal part appeared to be important for the emulsion-stabilizing properties of these peptides. A hypothesis for the structure-function relationship is given. 相似文献
4.
Biopolymer interactions have many potential applications in pharmaceutical, cosmetic, nutraceutical, and functional food industries. Attractive interactions between proteins and polysaccharides can lead to the formation of complexes. Binding parameters of beta-lactoglobulin (beta-lg)/pectin complexes were determined using frontal analysis continuous capillary electrophoresis and the overlapping binding site model. At pH 4, approximately 23 beta-lg molecules were cooperatively complexed on low-methoxyl pectin, where each beta-lg molecule covered an average of 12 galacturonic acid residues. The calculated binding constant was 1431 M(-1). The interactions between pectin and four selected peptides located on the outer surface of the beta-lg were investigated in order to identify which part of the protein was likely to interact with the pectin. The peptide beta-lg 132-148, which corresponds to the alpha-helix zone, and the peptides beta-lg 76-83, 41-60, and 1-14 would be involved in the interaction with the pectin. 相似文献
5.
6.
Bolder SG Hendrickx H Sagis LM van der Linden E 《Journal of agricultural and food chemistry》2006,54(12):4229-4234
Fibril formation in mixtures of whey proteins upon heating at pH 2 was investigated. Fibrils were found to coexist with other structures, such as spherulites. These spherulites consist of radially oriented fibrils. At total protein concentrations above 6 wt %, transparent gels were formed. Changing the ratio between the various whey proteins did not affect this gelation concentration as long as beta-lactoglobulin (beta-lg) was present, suggesting that beta-lg was dominant in the gelation. Pure alpha-lactalbumin and pure bovine serum albumin did not form fibrils, nor did they gel upon heating at pH 2 and 80 degrees C for up to 10 h. They did however induce a decrease in the beta-lg concentration needed for gel formation upon heating at pH 2. Our results suggest that beta-lg is the only fibril forming protein at the conditions used and that no mixed fibrils are formed. 相似文献
7.
Upon hydrolysis with chymotrypsin, soy glycinin has a strong tendency to aggregate. The regions of glycinin from which the aggregating peptides originate were identified by accumulative-quantitative peptide mapping. To this end, the aggregating peptides were further hydrolyzed with trypsin to obtain peptides of which the sequence can be identified using RP-HPLC-MS/MS. This resulted in a hydrolysate in which 90% of the proteinaceous material was dissolved. The soluble fraction was analyzed using the method of accumulative-quantitative peptide mapping: fractionation using ion exchange chromatography, followed by identification of peptides by RP-HPLC-MS/MS, quantification based on the absorbance at 214 nm, and finally peptide mapping. For the peptide mapping the proportions in which each of the five glycinin subunits are present, as determined by Edman degradation, were taken into account. The results showed that mainly the basic polypeptide and a part of the acidic polypeptide, close to the location of the disulfide bridge between the basic and acidic polypeptides, are present in the aggregating peptide fraction. On the basis of the results obtained, an aggregation mechanism was proposed. The hydrophilic acidic polypeptides shield the hydrophobic basic polypeptides, and the former are preferentially degraded upon hydrolysis. This results in a net increase in hydrophobicity of the remaining material, which mainly consists of the basic polypeptide fragments. This increase in hydrophobicity is proposed to be the driving force in the aggregation of chymotrypsin-derived peptides of glycinin. 相似文献
8.
Otte J Lomholt SB Halkier T Qvist KB 《Journal of agricultural and food chemistry》2000,48(6):2443-2447
The purpose of the present study was to identify the peptides responsible for aggregate formation during hydrolysis of beta-lactoglobulin by BLP at neutral pH. Hydrolysates taken at various stages of aggregate formation were separated into a precipitate and a soluble phase and each was analyzed by CE and mass spectrometry. The aggregates consisted of six to seven major peptides of which four were tentatively identified. The peptides were positively charged at neutral pH and had a high charge-to-mass ratio at low pH. The fragment f135-158 seemed to be the initiator of aggregation, since it was present at high concentration in the aggregates at all stages, and the concentration of this peptide remained low in the supernatant. F135-158 contains several basic and acid amino acids alternating with hydrophobic amino acids, which is in accordance with formation of noncovalently linked aggregates, as previously shown. 相似文献
9.
The effect of pH in the range 6.0-8.0 on the denaturation and aggregation of beta-lactoglobulin (beta-lg) was investigated. Results were interpreted in terms of the reaction scheme for the denaturation and aggregation of beta-lg proposed by Roefs and De Kruif (Eur. J. Biochem. 1994, 226, 883-889). The rate of conversion of native beta-lg increased strongly at higher pH values, whereas the molecular mass of the aggregates decreased strongly. In the pH range 6.4-8.0 aggregates were formed mainly by intermolecular disulfide bonds, but even at pH 6.0, thiol/disulfide exchange reactions were involved, although to a lesser extent. The time course of the exposure of the thiol group in native beta-lg upon heating and the subsequent disappearance of this group through the formation of disulfide-linked aggregates was investigated by reaction with 5,5'-dithiobis(2-nitrobenzoic acid) and varied strongly with pH. These observations could be used, in combination with the reaction steps of the reaction scheme, to describe qualitatively the strongly pH-dependent isothermal calorimetry curves, measured at 65 degrees C. 相似文献
10.
Groleau PE Morin P Gauthier SF Pouliot Y 《Journal of agricultural and food chemistry》2003,51(15):4370-4375
The objective of this study was to characterize the changes in peptide solubility resulting from changing some physicochemical conditions in a tryptic hydrolysate of beta-lactoglobulin (beta-LG). The turbidity (500 nm) of a 1% solution of tryptic peptides was measured at pH 3-10, at 5, 25, and 50 degrees C, in the presence of different salt concentrations (0, 0.5, and 1 M NaCl), in the presence of denaturing and reducing agents (6 M urea, 5% SDS, or 5% beta-mercaptoethanol), and under an electric field (isoelectric focusing). The results reveal an increase in turbidity of the peptide solution at pH 4, but a slight increase in turbidity was also observed at pH 8, which is attributable to peptides linked by disulfide bridges. The effect of temperature and ionic strength on the turbidity occurring at pH 4 indicates that mainly hydrophobic interactions are involved in the aggregation process. The material in the precipitate at pH 4 was identified as the peptides beta-LG 1-8, 15-20, and 41-60 and non-hydrolyzed alpha-lactalbumin. These results suggest that a limited number of peptides are involved in the aggregation process observed at pH 4, some of which having bioactive (beta-LG 15-20, ACE inhibitor, and opioid) or emulsifying properties (beta-LG 41-60). Aggregation of these peptides at acidic pH indicates that a simple acidification step could represent an easy process for isolating peptidic fractions enriched in bioactive or functional peptides. 相似文献
11.
The disulfide bonds of beta-lactoglobulin (beta-lg) were modified by oxidative sulfitolysis to generate beta-lgSO(3). The native protein (beta-lg) and the modified protein (beta-lgSO(3)) were conjugated to activated polyethylene glycol (PEG) to generate beta-lgPEG and beta-lgSO(3)PEG, respectively. Oil-in-water (o/w) emulsions containing 1% beta-lg or beta-lg conjugates were prepared at pH 2.8, 5.0, and 7.0. Emulsion droplet diameters and zeta potentials were measured. For the same emulsifier, emulsion droplet diameters decreased when emulsion pH increased. Zeta potentials of emulsion droplets increased with pH for beta-lg and beta-lgSO(3). Zeta potentials of beta-lgPEG and beta-lgSO(3)PEG approached zero, suggesting that the protein molecule was covered by PEG chains. Accelerated and 7-day storage stabilities at 21 degrees C of the emulsions were monitored. The emulsifying activity index (EAI) of beta-lgPEG was not significantly different from the EAI of beta-lg. The EAI of beta-lg was enhanced following sulfitolysis of beta-lactoglobulin. The emulsifying activity increased more when the oxidatively modified protein was conjugated to polyethylene glycol. Emulsions made with beta-lgSO(3)PEG were more stable than emulsions made with beta-lg, beta-lgPEG, or beta-lgSO(3) under accelerated stability study and for 7 days at 21 degrees C. The stability of o/w emulsions stabilized with beta-lgSO(3)PEG increased because individual droplets were better protected, against protein bridging or coalescence, by the thick adsorbed protein-PEG layer. 相似文献
12.
Edens L Dekker P van der Hoeven R Deen F de Roos A Floris R 《Journal of agricultural and food chemistry》2005,53(20):7950-7957
The observation that the bitterest peptides from casein hydrolysates contain several proline residues led us to hypothesize that a proline-specific protease would be instrumental in debittering such peptides. To identify the desired proline-specific activity, a microbiological screening was carried out in which the chromogenic peptide benzyloxycarbonyl-glycine-proline-p-nitroanilide (Z-Gly-Pro-pNA) was used as the substrate. An Aspergillus niger (A. niger) strain was identified that produces an extracellular proline-specific protease with an acidic pH optimum. On the basis of sequence similarities, we conclude that the A. niger-derived enzyme probably belongs to the S28 family of clan SC of serine proteases rather than the S9 family to which prolyl oligopeptidases belong. Incubating the overexpressed and purified enzyme with bitter casein hydrolysates showed a major debittering effect. Reversed phase HPLC analysis revealed that this debittering effect is accompanied by a significant reduction of the number of hydrophobic peptides present. 相似文献
13.
Stochmal A Piacente S Pizza C De Riccardis F Leitz R Oleszek W 《Journal of agricultural and food chemistry》2001,49(2):753-758
Nine flavones and adenosine have been identified in aerial parts of alfalfa, and their structures were established by spectral (FABMS and NMR) techniques. Five of the identified compounds, including apigenin 7-O-[beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[2-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[2-O-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], apigenin 7-O-[2-O-p-coumaroyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], and luteolin 7-O-[2-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, have not been reported before in the plant kingdom. Additionally, five known compounds, including apigenin 7-O-beta-D-glucuronopyranoside, apigenin 4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[beta-D- glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], luteolin 7-O-beta-D-glucuronopyranoside, and adenosine, were identified. 相似文献
14.
The in vitro angiotensin I-converting enyzme (ACE) inhibitory activity of Pacific hake hydrolysates was investigated as a function of hydrolysis conditions, starting material variability, and ultrafiltration. Hake fillets were hydrolyzed using Protamex protease under various conditions of pH, hydrolysis time, and enzyme-to-substrate ratio (% E/S) according to a response surface methodology (RSM) central composite design. The hydrolysate produced at pH 6.5, 125 min, and 3.0% E/S had an IC 50 of 165 +/- 9 microg of total solids/mL. ACE-inhibitory activity was not significantly different (P < 0.05) for hydrolysates produced using higher time-enzyme combinations within the model or from fish of different catches. Ultrafiltration (10 kDa molecular mass cutoff) resulted in an IC50 value of 44 +/- 7 microg of peptides/mL, 2.5 times more potent than the commercial product PeptACE Peptides (IC50 = 114 +/- 8 microg of peptides/mL). These results suggest that hydrolysates prepared with minimal fractionation from Pacific hake, an undervalued fish, may be a commercially competitive source of ACE-inhibitory peptides. 相似文献
15.
Isothermal titration calorimetry (ITC) was used to determine the binding constant, stoichiometry, enthalpy, and entropy of beta-lactoglobulin/low- and high-methoxyl pectin (beta-lg-LM- and HM-pectin) complexes at 22 degrees C and at pH 4. The binding isotherms revealed the formation of soluble intrapolymer complexes (C1) further followed by their aggregation in interpolymer complexes (C2). The interaction between beta-lg and LM- or HM-pectin in C1 and C2 occurred spontaneously with a Gibbs free energy around -10 kcal/mol. The C1 were enthalpically driven, whereas enthalpic and entropic factors were involved in the C2 formation. Because ITC did not allow the dissociation of different enthalpic contributions, the values measured as pectin and beta-lg interacted could partially be attributed to conformational changes. The C1 had a binding stoichiometry of 8.3 and 6.1 beta-lg molecules complexed per LM- or HM-pectin molecule, respectively. The C2 had about 16.5 and 15.1 beta-lg molecules complexed per LM- and HM-pectin, respectively. 相似文献
16.
从秸秆还田土壤中初筛获得一株高效纤维素降解菌CMC-4,根据菌株形态、理化性质及16S rDNA序列分析,初步鉴定为地衣芽孢杆菌(Bacillus licheniformis)。以此为出发菌株,经亚硝酸钠诱变获得一株稳定高产纤维素酶突变株CMC-4-3。对CMC-4和CMC-4-3的产酶条件和酶学性质进行比较分析,结果表明:CMC-4-3纤维素酶活力较CMC-4提高67.5%;其最适产酶条件是:37℃、pH 6.0、葡萄糖为碳源、蛋白胨为氮源、接种量2.0%、装液量60 ml/250 ml。该菌株所产纤维素酶的最适反应pH为6.0,且在4.0~7.0酶活力较稳定;最适反应温度为50℃,在20~80℃范围内均保持稳定;金属离子Fe~(2+)、Mg~(2+)、Co~(2+)、K~+对酶有激活作用,其余离子均有不同程度抑制作用,而Cu~(2+)和Ca~(2+)抑制作用最强,酶活力减弱近50%,是该酶的强效抑制因子。诱变前后菌株产酶条件和酶学性质等部分表型发生了变化,而突变菌株显示出了更宽泛的环境适应范围。据此,获得一株高效产纤维素酶、耐受范围广的具纤维素降解能力的地衣芽孢杆菌,而CMC-4-3和CMC-4的表型可作为深入探讨基因型变化的线索。 相似文献
17.
The aim of this study was to determine if peptides could interact with beta-lactoglobulin (beta-LG) and what the physicochemical conditions promoting their interaction with the protein are. The binding of negatively charged (beta-LG 125-135 and 130-135), positively charged (beta-LG 69-83 and 146-149), and hydrophobic (alphaS1-CN 23-34 and beta-LG 102-105, both bioactive peptides) peptides to bovine beta-LG was determined using an ultrafiltration method under different physicochemical conditions: pH 3.0, 6.8, and 8.0; buffers of 0.05 and 0.1 M; 4, 25, and 40 degrees C; beta-LG/peptide ratios of 1:5 and 1:10. At pH 3.0, none of the peptides interacted with beta-LG at any temperature, buffer molarity, or beta-LG/peptide ratio probably due to electrostatic repulsions between the highly protonated species. At pH 6.8 and 8.0, charged peptides beta-LG 130-135, 69-83, and 146-149 bound to beta-LG under some physicochemical conditions, possibly by nonspecific binding. However, both hydrophobic peptides probably bind to the inner cavity (beta-barrel) of beta-LG, provoking the release of materials absorbing at 214 nm. Given the known biological activities of the hydrophobic peptides used in this study (opioid and ACE-inhibitory activities), their binding to beta-LG may be relevant to a better understanding of the physiological function of the protein. 相似文献
18.
Sorption behavior of triazole fungicides in Indian soils and its correlation with soil properties 总被引：6，自引：0，他引：6
Singh N 《Journal of agricultural and food chemistry》2002,50(22):6434-6439
Adsorption-desorption of triazole fungicides, hexaconazole [2-(2,4-dichlorophenyl)-1-(1H-1,2,4,-triazol-1-yl) hexan-2-ol], triadimefon [1-(4-chlorophenoxy)-3,3-dimethyl-1-(1H-1,2,4-triazol-1-yl) butan-2-one], and penconazole[1-(2,4-dichloro-beta-propyl phenethyl)-1H-1,2,4-triazole] was studied in five Indian soils using batch method. The adsorption isotherms fitted very well to the Freundlich equation. Adsorption of various triazole fungicides increased in this order: triadimefon > hexaconazole > penconazole. The product of the Freundlich adsorption constants, K(f)(1/n), showed good correlation with the soil organic carbon (OC) content, suggesting that soil OC is the main controlling factor for triazoles adsorption. Clay and silt content of the soil also affected the adsorption constants. Adsorption of hexaconazole and triadimefon was nearly reversible in two low OC soils (soil 3, soil 5) where 90-100% of the sorbed fungicides was released in a single washing step. Otherwise, desorption of triazole fungicides showed hysteresis, and 30-60% of the triazole fungicides were retained by the soil after single washing. IR spectra showed that H-bonds and charge-transfer bonds between humic acid and fungicides probably operated as mechanisms of adsorption. 相似文献
19.
The respective role of organic materials and poorly-ordered Al and Fe hydrous oxides on soil aggregate stability was studied in silty soils with little swelling clay, using both multivariate analysis and physico-chemical approaches. Soil disaggregation is a function of the hydrophobic character of organic matter (OM), which depends on the nature of the organic materials, cationic environment and the aggregating effect of Al and Fe hydrous oxides.
Two kinds of aggregates >50 μm can be distinguished in organic soils, one being about six times more stable than the other. In soils poor in organic matter, weak aggregates dominate; the binding agents are Ca or Al OM rich in polysaccharides and peptides. 相似文献
Two kinds of aggregates >50 μm can be distinguished in organic soils, one being about six times more stable than the other. In soils poor in organic matter, weak aggregates dominate; the binding agents are Ca or Al OM rich in polysaccharides and peptides. 相似文献
20.
Su ZY Hwang LS Kuo YH Shu CH Sheen LY 《Journal of agricultural and food chemistry》2008,56(20):9447-9454
The antihepatoma activity and related active components in the fermentation products of Agaricus blazei (AB) cultured in the medium containing soybean (S) or black soybean (BS) were investigated. AB(BS)-pE and AB(S)-pE were the ethanolic extracts from the fermentation products of AB(BS) and AB(S), respectively. According to the IC 50 values, AB(BS)-pE (161.1 and 24.0 microg/mL for Hep 3B and Hep G2 cells, respectively) exhibited stronger cytotoxicities against hepatoma cells than AB(S)-pE (>200 and 99.9 microg/mL for Hep 3B and Hep G2 cells, respectively). AB(BS)-pE was separated by silica gel column chromatography and eluted with n-hexane/ethyl acetate/methanol gradient solvent system into 21 fractions. Fraction 3 [AB(BS)-pE-F3], eluted with n-hexane/ethyl acetate (97:3 and 19:1, v/v), was the most active fraction having inhibitory activity on the proliferation of Hep 3B and Hep G2 cells (IC 50 of 3.6 and 1.9 microg/mL, respectively). Three major compounds, compounds 1- 3, were further isolated from the AB(BS)-pE-F3 fraction by reversed-phase semipreparative high-performance liquid chromatography. Compounds 2 and 3 gave better antihepatoma activity than that of compound 1. The IC 50 values of compounds 2 and 3 were 2.8 and 4.5 microg/mL for Hep 3B cells and 1.4 and 2.0 microg/mL for Hep G2 cells, respectively. The structures of compounds 2 and 3 were identified by UV, IR, electron impact mass spectrometry, and (1)H and (13)C NMR to be blazeispirols A and C, respectively. Blazeispirols A and C existed in the mycelia but not in the broth and were more in AB(BS)-pE (49.9 +/- 8.9 and 14.2 +/- 2.4 mg/g, respectively) than AB(S)-pE (15.9 +/- 1.7 and 3.9 +/- 0.6 mg/g, respectively). Additionally, the result shows that the production of blazeispirols A and C was increased after cultivation in the medium containing black soybean on day 6 and reached the maximum on day 12, and the contents of blazeispirols A and C were negatively correlated with Hep 3B and Hep G2 cell viabilities ( r = -0.84 to -0.93, P < 0.01). It suggests that blazeispirols A and C could be used as biomarkers to produce the fermentation product of A. blazei with antihepatoma activity. 相似文献