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1.
The complement of enzyme activities of a selection of commercial protease preparations were determined using fluorogenic substrates. Alcalase was used in combination with other commercial enzyme preparations to produce cod muscle (Gadus morhua) hydrolysates. Each muscle hydrolysate was characterized with respect to the percentage degree of hydrolysis (DH %), peptide molecular weight range, and free amino acid content. The enzyme preparations containing predominantly protease or endopeptidase activities achieved high DH % and produced significant amounts of peptides below a molecular weight of 3000. Alcalase combined with exopeptidase-rich preparations produced hydrolysates rich in low-molecular-weight peptides. Selecting combinations of enzyme preparations with complementary activity profiles could be used to manipulate the peptide molecular weight profile of hydrolysates.  相似文献   

2.
Brewers' spent grain (BSG) is the insoluble residue of barley malt resulting from the manufacture of wort. Although it is the main byproduct of the brewing industry, it has received little attention as a marketable commodity and is mainly used as animal feed. Our work focuses on one of the main constituents of BSG, i.e., the proteins. The lack of solubility of BSG proteins is one of the limitations for their more extensive use in food processing. We therefore aimed to generate BSG protein hydrolysates with improved technofunctional properties. BSG protein concentrate (BPC) was prepared by alkaline extraction of BSG and subsequent acid precipitation. BPC was enzymatically hydrolyzed in a pH-stat setup by several commercially available proteases (Alcalase, Flavourzyme, and Pepsin) for different times and/or with different enzyme concentrations in order to obtain hydrolysates with different degrees of hydrolysis (DH). Physicochemical properties, such as molecular weight (MW) distribution and hydrophobicity, as well as technofunctional properties, such as solubility, color, and emulsifying and foaming properties, were determined. Enzymatic hydrolysis of BPC improved emulsion and/or foam-forming properties. However, for the hydrolysates prepared with Alcalase and Pepsin, an increasing DH generally decreased emulsifying and foam-forming capacities. Moreover, the type of enzyme impacted the resulting technofunctional properties. Hydrolysates prepared with Flavourzyme showed good technofunctional properties, independent of the DH. Physicochemical characterization of the hydrolysates indicated the importance of protein fragments with relatively high MW (exceeding 14.5 k) and high surface hydrophobicity for favorable technofunctional properties.  相似文献   

3.
The aim of this work was to study the cleaning of inorganic membranes fouled with whey protein solutions using the enzymatic formulation Alcalase (Novo Nordisk A/S). Hydraulic and chemical methods were considered to characterize the cleanliness of the membranes. Cleaning efficiency was observed to be a function of the operating conditions. The operating conditions tested were the following: recycling versus non-recycling of permeate, pH of the cleaning solution, addition of alkali to regulate the pH, enzymatic agent concentration, and cleaning time. The best conditions to perform the cleaning were related to the best conditions to hydrolyze whey proteins in a discontinuous reactor using the same enzyme preparations. Very high cleaning efficiencies (>90%) were achieved in short operating times (20 min). However, residual matter was observed on the membrane surface.  相似文献   

4.
Dipeptidyl peptidase I (DPP I; EC 3.4.14.1) was purified from porcine skeletal muscle after several steps such as heat treatment, ammonium sulfate fractionation, gel filtration chromatography, and HPLC anion exchange chromatography. The purified enzyme showed a native molecular mass of approximately 200 kDa on Sephacryl S-200 column chromatography. Two protein bands of 65 and 42 kDa were obtained by SDS-PAGE, indicating its oligomeric nature. Maximum activity was reached at pH 5.5 and 55 degrees C. DPP I shared some common substrate specificities, both on synthetic derivatives and on real peptides, with porcine muscle DPP III. The enzyme required reducing agents for full activation, although the halide requirement was not proved. DPP I was inhibited by the assayed cysteine peptidase inhibitors except p-CMB. The serine peptidase inhibitor 3, 4-DCI also inhibited the enzyme as did the divalent cations Co(2+), Mn(2+), and Zn(2+). On the basis of its properties, DPP I may contribute to the generation of dipeptides during the processing of meat and/or meat products, including cooked ham.  相似文献   

5.
ABSTRACT

Cucumbers are produced in integrated hydroponic and aquaculture systems (aquaponics). Aquaponics balances pH for plants, fish, and nitrifying bacteria. Nitrification prevents buildup of toxic waste ammonia by conversion to nitrate (NO3 ?)- nitrogen (N). The pH for hydroponic cucumbers (5.5–6.0) and nitrification (7.5–9.0) requires reconciliation to improve systems integration. Cucumbers were grown at pH of 5.0, 6.0, 7.0, and 8.0 with additional foliar sprays at pH 7.0 and 8.0. Plant shoot dry weight, length, N, and phosphorus (P) content at 14 DAT were similar from pH 5.0 to 7.0, but reduced at pH 8.0. Nutrient solution and shoot dry matter Mn and Fe decreased as pH increased. Foliar sprays had no effect on cucumber fruit yield. Early yield was higher at pH 5.0 compared to pH 8.0 but total yield was unaffected by pH. Cucumbers in recirculating culture may be maintained at pH levels more optimum for nitrification (7.5–8.0) except during production for early season markets.  相似文献   

6.
The aim was to identify to what extent proteins were utilized during the fermentation of bacteria-free tempe prepared with acidified soybean cotyledons and Rhizopus oligosporus NRRL 2710 at 30 degrees C. Dry matter declined continuously during the fermentation to 980 g/(kg of initial dry cotyledons) at 28 h, 910 g at 46 h (when the tempe was judged mature), and 835 g at 72 h. The decrease in dry matter was due mainly to reduction in crude lipid, amounting to 65 g/(kg of initial dry cotyledons) at 46 h and 135 g/(kg of initial dry cotyledons) at 72 h and representing approximately 70% and 80%, respectively, of the total dry matter loss. Protein oxidation (estimated from ammonia production) was 5 g at 28 h,10 g at 46 h, and 20 g/(kg of initial dry cotyledons) at 72 h. The total amount of soy protein hydrolyzed, including that incorporated into mold biomass, was estimated to be 80 g/(kg of initial dry cotyledons) at 28 h incubation, 95 g at 46 h, and 100 g at 72 h. The hydrolyzed protein at 46 h represented 25% of the initial protein. Of this hydrolyzed protein, it is suggested that approximately 65% remained in the tempe as amino acids and peptides, 25% was assimilated into mold biomass, and 10% was oxidized.  相似文献   

7.
Extensive hydrolysis of whey protein isolate by Alcalase was shown to induce gelation mainly via hydrophobic interactions. The aim of this work was to characterize the peptides released in order to better understand this phenomenon. The apparent molecular mass distribution indicated that aggregates were formed by small molecular mass peptides (<2000 Da). One hundred and thirty peptides with various lengths were identified by reversed-phase high-performance liquid chromatography coupled with electrospray ionization mass spectrometry. Alcalase was observed to have a high specificity for aromatic (Phe, Trp, and Tyr), acidic (Glu), sulfur-containing (Met), aliphatic (Leu and Ala), hydroxyl (Ser), and basic (Lys) residues. Most peptides had an average hydrophobicity of 1-1.5 kcal/residue and a net charge of 0 at the pH at which gelation occurred (6.0). Therefore, an intermolecular attractive force such as hydrophobic interaction suggests the formation of aggregates that further leads to the formation of a gel.  相似文献   

8.
Lunasin and Bowman-Birk protease inhibitor (BBI) are two soybean peptides to which health-promoting properties have been attributed. Concentrations of these peptides were determined in skim fractions produced by enzyme-assisted aqueous extraction processing (EAEP) of extruded full-fat soybean flakes (an alternative to extracting oil from soybeans with hexane) and compared with similar extracts from hexane-defatted soybean meal. Oil and protein were extracted by using countercurrent two-stage EAEP of soybeans at 1:6 solids-to-liquid ratio, 50 °C, pH 9.0, and 120 rpm for 1 h. Protein-rich skim fractions were produced from extruded full-fat soybean flakes using different enzyme strategies in EAEP: 0.5% protease (wt/g extruded flakes) used in both extraction stages; 0.5% protease used only in the second extraction stage; no enzyme used in either extraction stage. Countercurrent two-stage protein extraction of air-desolventized, hexane-defatted soybean flakes was used as a control. Protein extraction yields increased from 66% to 89-96% when using countercurrent two-stage EAEP with extruded full-fat flakes compared to 85% when using countercurrent two-stage protein extraction of air-desolventized, hexane-defatted soybean flakes. Extruding full-fat soybean flakes reduced BBI activity. Enzymatic hydrolysis reduced BBI contents of EAEP skims. Lunasin, however, was more resistant to both enzymatic hydrolysis and heat denaturation. Although using enzymes in both EAEP extraction stages yielded the highest protein and oil extractions, reducing enzyme use to only the second stage preserved much of the BBI and Lunasin.  相似文献   

9.
Solubility of rainbow trout proteins was determined between pH 1.5 and 13.0 and various ionic strengths (IS). Minimum solubility occurred at pH 5.5; however, when IS = 0.2, the minimum solubility shifted toward more acidic pH. Isoelectric solubilization/precipitation was applied to trout processing byproducts (fish meat left over on bones, head, skin, etc.), resulting in protein recovery yields (Kjeldahl, dry basis) between 77.7% and 89.0%, depending of the pH used for solubilization and precipitation. The recovered protein contained 1.4-2.1% ash (dry basis), while the trout processing byproducts (i.e., starting material) 13.9%. Typical boneless and skinless trout fillets contain 5.5% ash, and therefore, the isoelectric solubilization/precipitation effectively removed impurities such as bones, scales, skin, etc., from the trout processing byproducts. The recovered proteins retained gel-forming ability as assessed with dynamic rheology, torsion test, and texture profile analysis (TPA). However, the recovered proteins failed to gel unless beef plasma protein (BPP) was added. Even with BPP, the recovered protein showed some proteolysis between 40 and 55 degrees C. Addition of potato starch, transglutaminase, and phosphate to the recovered proteins resulted in good texture of trout gels as confirmed by torsion test and TPA. Higher ( P < 0.05) shear stress and strain were measured for gels developed from basic pH treatments than the acidic counterparts. However, proteins recovered from acidic treatments had higher ( P < 0.05) lipid content than the basic treatments. This is probably why the gels from acidic treatments were whiter ( L* - 3 b*) ( P < 0.05) than those from the basic ones. Our study demonstrates that functional proteins can be efficiently recovered from low-value fish processing byproducts using isoelectric solubilization/precipitation and subsequently be used in value-added human foods.  相似文献   

10.
A new low temperature liquefaction and saccharification enzyme STARGEN 001 (Genencor International, Palo Alto, CA) with high granular starch hydrolyzing activity was used in enzymatic dry‐grind corn process to improve recovery of germ and pericarp fiber before fermentation. Enzymatic dry‐grind corn process was compared with conventional dry‐grind corn process using STARGEN 001 with same process parameters of dry solid content, pH, temperature, enzyme and yeast usage, and time. Sugar, ethanol, glycerol and organic acid profiles, fermentation rate, ethanol and coproducts yields were investigated. Final ethanol concentration of enzymatic dry‐grind corn process was 15.5 ± 0.2% (v/v), which was 9.2% higher than conventional process. Fermentation rate was also higher for enzymatic dry‐grind corn process. Ethanol yields of enzymatic and conventional dry‐grind corn processes were 0.395 ± 0.006 and 0.417 ± 0.002 L/kg (2.65 ± 0.04 and 2.80 ± 0.01 gal/bu), respectively. Three additional coproducts, germ 8.0 ± 0.4% (db), pericarp fiber 7.7 ± 0.4% (db), and endosperm fiber 5.2 ± 0.6% (db) were produced in addition to DDGS with enzymatic dry‐grind corn process. DDGS generated from enzymatic dry‐grind corn process was 66% less than conventional process.  相似文献   

11.
The enzymatic hydrolysis of brewers' spent grain (BSG) has been investigated through treatment with commercial carbohydrases and proteases. Resultant residues were then chemically fractionated and delignified. Enzymatic treatments released 25-30% of the BSG mass and yielded precursors suitable for subsequent conversion to potentially value-added products. Controlled chemical fractionation selectively solubilized arabinoxylan but with no differences apparent due to prior enzyme treatment. The loss of non-polysaccharide components during alkali treatment suggests the presence of a high proportion of alkali-soluble lignin. Further delignification of the alkali-insoluble residues and further chemical fractionation released the remaining hemicellulose, to yield a residue which was >90% cellulose. Further knowledge of the properties and interaction between BSG polymers will facilitate an improved enzyme-assisted total deconstruction of BSG and hence the exploitation of its biomass.  相似文献   

12.
In this study, in vitro digestion of β-lactoglobulin (β-Lg) fibrils and the re-formation of fibril-like structures after prolonged enzymatic hydrolysis (up to 48 h) were investigated using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), thioflavin T fluorescence photometry, and transmission electron microscopy (TEM). Pure β-Lg fibrils that had been formed by heat treatment at pH 2.0 were rapidly hydrolyzed by pepsin in the simulated gastric fluid (pH 1.2), and some new peptides that were suitable for further fibril formation were produced. TEM showed that the new fibrils were long and straight but thinner than the original fibrils, and both TEM and MALDI-MS indicated that the peptides in the new fibrils were shorter/smaller than the peptides in the original fibrils. The formation of new fibrils was found to be affected more by pH than by enzyme activity or temperature.  相似文献   

13.
Pecans (cv. Desirable) contained approximately 10% protein on a dry weight basis. The minimum nitrogen solubility (5.9-7.5%) at 0.25-0.75 M trichloroacetic acid represented the nonprotein nitrogen. Among the solvents assessed for protein solubilization, 0.1 M NaOH was the most effective, while borate saline buffer (pH 8.45) was judged to be optimal for protein solubilization. The protein solubility was minimal in the pH range of 3-7 and significantly increased on either side of this pH range. Increasing the NaCl concentration from 0 to 4 M significantly improved ( approximately 8-fold increase) protein solubilization. Following Osborne protein fractionation, the alkali-soluble glutelin fraction (60.1%) accounted for a major portion of pecan proteins followed by globulin (31.5%), prolamin (3.4%), and albumin (1.5%), respectively. The majority of pecan polypeptides were in the molecular mass range of 12-66 kDa and in the pI range of 4.0-8.3. The pecan globulin fraction was characterized by the presence of several glycoprotein polypeptides. Lysine was the first limiting essential amino acid in the defatted flour, globulin, prolamin, and alkaline glutelin fractions. Leucine and tryptophan were the first limiting essential amino acids in albumin and acid glutelin fractions, respectively. Rabbit polyclonal antibodies detected a range of pecan polypeptides in the 12-60 kDa range, of which the globulin fraction contained the most reactive polypeptides.  相似文献   

14.
Polyphenol oxidase (PPO) of garland chrysanthemum (Chrysanthemum coronarium L.) was purified approximately 32-fold with a recovery rate of 16% by ammonium sulfate fractionation, ion exchange chromatography, hydrophobic chromatography, and gel filtration. The purified enzyme appeared as a single band on PAGE and SDS-PAGE. The molecular weight of the enzyme was estimated to be about 47000 and 45000 by gel filtration and SDS-PAGE, respectively. The purified enzyme quickly oxidized chlorogenic acid and (-)-epicatechin. The K(m) value (Michaelis constant) of the enzyme was 2.0 mM for chlorogenic acid (pH 4.0, 30 degrees C) and 10.0 mM for (-)-epicatechin (pH 8.0, 40 degrees C). The optimum pH was 4.0 for chlorogenic acid oxidase (ChO) and 8.0 for (-)-epicatechin oxidase (EpO). In the pH range from 5 to 11, their activities were quite stable at 5 degrees C for 22 h. The optimum temperatures of ChO and EpO activities were 30 and 40 degrees C, respectively. Both activities were stable at up to 50 degrees C after heat treatment for 30 min. The purified enzyme was strongly inhibited by l-ascorbic acid and l-cysteine at 1 mM.  相似文献   

15.
Hydrolysis of whey protein concentrate (WPC) with Alcalase 2.4 L, a Bacillus licheniformis proteinase preparation, induces gelation. The aggregation behavior of WPC hydrolysates generated with Alcalase and Prolyve 1000, a Bacillus licheniformis proteinase that did not induce gelation, were studied by turbidity and particle size analysis. With the use of synthetic peptide substrates, it was shown that Alcalase contains a glutamyl endopeptidase (GE) activity not present in Prolyve. Comparison of the aggregation behavior of WPC hydrolysates generated with Alcalase, Prolyve, and combinations of Prolyve with a GE activity isolated from Alcalase showed that GE was responsible for the observed enzyme-induced peptide aggregation in Alcalase hydrolysates. Hydrolysates generated with Prolyve, having a degree of hydrolysis (DH) of 11.8% and 10.4% of peptide material greater than 10 kDa, could be induced to aggregate by the addition of GE. These results emphasize the contribution of enzyme specificity to the physicochemical and functional characteristics of proteinase hydrolysates of WPC.  相似文献   

16.
《Cereal Chemistry》2017,94(1):117-123
The objective of this study was to evaluate the antihypertensive potential of common bean protein hydrolysate. Protein concentrates were obtained, followed by Alcalase enzymatic hydrolysis, and then ultrafiltrated (3,000 molecular weight cutoff); the lyophilized product was named BP3. The angiotensin converting enzyme (ACE) inhibitory activity was determined as IC50 (3.68 ± 0.07 μg/mL). The antihypertensive effect was evaluated in spontaneously hypertensive rats (SHR) by two assays; Captopril ACE inhibitor was used as a reference compound and water as a control. A short‐term assay showed a maximum decrease in mean arterial pressure of –41 ± 5 mmHg in SHR, 3 h after oral administration of 500 mg of BP3/kg of body weight (bw). In a long‐term assay, a significant decrease in systolic blood pressure of –24 ± 5 mmHg was observed in SHR, after 45 days of oral administration of 500 mg of BP3/kg of bw/12 h. In both assays, BP3 treatment showed antihypertensive effect over SHR, similar to Captopril treatment. The sequences of the most abundant peptides present in BP3, determined by mass spectrometry, were identified as KFPWVK, GADFRKK, and PQSPCKRVNRHS. These peptides are reported for the first time in Azufrado Higuera common beans, and they are most likely responsible for the antihypertensive effect of BP3.  相似文献   

17.
Wheat bran contains good quality protein, but given its location inside aleurone cells, this protein has restricted digestibility. The aim of this work was to liberate and solubilize wheat bran proteins via cell wall degradation by using carbohydrate‐hydrolyzing and proteolytic enzymes without causing extensive protein hydrolysis. Bran incubated with water (without added enzymes) for 16 h increased the solubilized organic nitrogen content from 14.0 to 42.8%. Enzymes with solely carbohydrate‐hydrolyzing activity increased the water‐soluble pentosan and reducing sugar contents but did not significantly increase protein solubilization or protein release from the aleurone cells. Enzymes with proteolytic activity significantly increased the solubilization of protein to 58.2% already at 4 h. Significant protein hydrolysis was detected with a high dosage of protease. However, based on light microscopy, the enzymatic treatment mainly modified the proteins in the subaleurone layer, and it was less effective on proteins inside the aleurone cells. With optimized protease treatment (3 h, 35°C, and 550 nkat/g), effective protein solubilization (>48%) without extensive protein hydrolysis (free amino nitrogen content <45 mg/L) was achieved. In conclusion, intensive solubilization of proteins in the subaleurone layer of wheat bran is possible by using exogenous enzymes with proteolytic activities.  相似文献   

18.
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   

19.
The noncharacterized protein ACL75304 encoded by the gene Ccel_0941 from Clostridium cellulolyticum H10 (ATCC 35319), previously proposed as the xylose isomerase domain protein TIM barrel, was cloned and expressed in Escherichia coli . The expressed enzyme was purified by nickel-affinity chromatography with electrophoretic homogeneity and then characterized as d-psicose 3-epimerase. The enzyme was strictly metal-dependent and showed a maximal activity in the presence of Co(2+). The optimum pH and temperature for enzyme activity were 55 °C and pH 8.0. The half-lives for the enzyme at 60 °C were 6.8 h and 10 min when incubated with and without Co(2+), respectively, suggesting that this enzyme was extremely thermostable in the presence of Co(2+) but readily inactivated without metal ion. The Michaelis-Menten constant (K(m)), turnover number (k(cat)), and catalytic efficiency (k(cat)/K(m)) values of the enzyme for substrate d-psicose were estimated to be 17.4 mM, 3243.4 min(-1), and 186.4 mM min(-1), respectively. The enzyme carried out the epimerization of d-fructose to d-psicose with a conversion yield of 32% under optimal conditions, suggesting that the enzyme is a potential d-psicose producer.  相似文献   

20.
A metagenomic library of China Holstein cow rumen microbes was constructed and screened for novel gene cluster. A novel feruloyl esterase (FAE) gene was identified with a length of 789 bp and encoded a protein displaying 56% identity to known esterase sequences. The gene was functionally expressed in Escherichia coli BL21 (DE3), and the total molecular weight of the recombined protein was 32.4 kDa. The purified enzyme showed a broad specificity against the four methyl esters of hydroxycinnamic acids and high activity (259.5 U/mg) to methyl ferulate at optimum conditions (pH 8.0, 40 °C). High thermal and pH stability were also observed. Moreover, the enzyme showed broad resistance to proteases. FAE-SH1 can enhance the release of ferulic acid from wheat straw with cellulase, β-1,4-endoxylanase, β-1,3-glucanase, and pectase. These features suggest FAE-SH1 as a good candidate to enhance biomass degradation and improve the health effects of food and forage.  相似文献   

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