Preparation of Soy Peptides by Liquid Fermentation

Many kinds of microorganism can produce a mount of protease which subsequently hydrolysis the protein of the medium into peptides when they grow in protein containing liquid medium.In the present investigation,the conditions of preparing soybean peptides by liquid fermentation were studied,following results were obtained:(1)SPI is a nice nitrogen source and meanwhile an inducible factor of protease production;its concentration can be as high as 3%-4%.(2)Sucrose is the best carbon source;its concentration is 1%-4%.(3)Under the conditions of 28℃,initial pH6.0,inoculum size 4%,cell age 36hr and fermentation time 24hr-30hr,we can obtain soybean peptides or fermentation liquor with good flavor,its DH reaches 25%-30% and the yield rate can be as high as 75%.(4)Mass spectrograph indicate the MW of the fermentation liquid or the soybean peptides mainly distribute at about 4000Dal,these imply a promising prospect of industrial application of submerged fermentation in producing soybean peptides.     

Effect of enzymatic cross-linking of β-casein on proteolysis by pepsin

Food texture has a significant influence on the sensation of satiety. The digestibility of a protein matrix can be decreased by e.g. disulfide cross-links during heating, but the structure and properties of a single protein molecule can also be modified by cross-linking enzymes. In this study the effects of cross-linking of β-casein by fungal Trichoderma reesei tyrosinase (TrTyr) and bacterial Streptoverticillium mobaraense transglutaminase (Tgase) on digestibility by proteolytic pepsin were investigated by different methods. The enzymatic reaction conditions were selected in such a way that high and low molecular mass cross-linked β-casein polymers were formed. SDS-PAGE (sodium dodecyl sulphate-polyacrylamide gel electrophoresis) was used to analyze the pH stability of the cross-linked β-casein in acidic solution mimicking gastric conditions typically present during proteolytic digestion. In order to monitor the extent of pepsin digestion, the proteolytic process was halted at specific time points and aliquots of the reaction mixtures were subjected to SDS-PAGE, size-exclusion chromatography (SEC) and matrix-assisted laser desorption ionization-time of flight mass spectrometric (MALDI-TOF MS) analyses in order to evaluate sizes and quantities of the digested protein fragments. A pH-stat method was used to determine the degree of hydrolysis (DH) of the enzymatically cross-linked β-casein. The results demonstrated that enzymatically cross-linked β-casein was stable under acidic conditions and was more resistant to pepsin digestion when compared to non cross-linked β-casein. The research results will have high impact on the development of novel food structures with improved properties such as good satiety, controlled energy intake and digestibility.     

Study of conformational properties of cereal β-glucans by computer modeling

Molecular modelling was used to study the conformational properties of cereal β-glucans and compared with experimental results. Followed the steps of exploring global minima of β 1-4 and β 1-3 linked disaccharides and building up the repeating units, the consecutive cellotriosyl units, which are considered as reaction sites of β-glucan chains, were created and then characterized as a three fold helix with a pitch of 41.35 Å. Mostly importantly, the 3D molecular model of cereal β-glucans was created and the moderately extended sinuous chain conformation was first visualized. The conformational parameters of cereal β-glucans were calculated by RMMC simulation, which are in good agreement with experimental results. The calculated parameters also revealed that the chain stiffness of β-glucans increased with the tri/tetra ratio.     

Species identification of ruminant milk by genotyping of the κ-casein gene

The development of molecular genetic and bioinformatic systems for identifying the species of milk and the raw material composition of dairy products is of great scientific and practical importance with the purpose of introducing developments in the system for controlling the turnover of falsified products. The aim of the research is to develop a method of PCR-RFLP analysis for species identification of milk and dairy products from agricultural ruminant animals by the κ-casein gene (CSN3) with the possibility of qualitative and relative quantitative assessment of species-specific DNA of the tested biomaterial. The objects of research were samples of raw milk and milk powder, pasteurized cream, and hard and semi-hard cheeses. The developed method of species identification of milk and dairy products includes sample preparation of the studied samples, nucleic acid extraction, combined PCR-RFLP technique, detection of obtained results by the method of horizontal electrophoresis in agarose gel and their analysis, including using the developed mathematical algorithms and software. The synergistic effect established in combined operation of 2 restriction enzymes ensured their application in a mix with increased performance in an ergonomic way in the context of DNA authentication of cow, goat, and sheep milk and dairy products based on them. The specificity and sensitivity of the proposed method is potentially suitable for implementing the development of a system to control the turnover of falsified and counterfeit goods.     

Enrichment of Rice Bran Oil with ��-Linolenic Acid by Enzymatic Acidolysis: Optimization of Parameters by Response Surface Methodology

Lipase-catalyzed enrichment of rice bran oil with n-3 fatty acid in order to obtain a structured lipid containing essential fatty acids has been optimized by response surface methodology. In this process, α-linolenic acid was used as an acyl donor using lipase-catalyzed acidolysis in hexane in presence of immobilized lipase from Rhizomucor miehei. The effect of incubation time and temperature, enzyme concentration and substrates mole ratio and their complex interaction on percentage incorporation of n-3 fatty acid, ratios of saturated fatty acid to polyunsaturated fatty acids, monounsaturated fatty acids to polyunsaturated fatty acids and n-6 to n-3 (18:2 to 18:3) fatty acids have been studied using a central composite rotatable design of experiments. The results showed that at the optimum conditions such as reaction time 4.5 h and reaction temperature 37. 5°C, substrate ratio ranging from 1.0 to 1.9, enzyme concentration varying from 1.0% to 2.0% are needed to fulfill the conditions such as percentage incorporation of n-3 fatty acid ≤18%, ratio of saturated fatty acid to poly unsaturated fatty acid ≥0.42, ratio of mono unsaturated fatty acid to poly unsaturated fatty acid ≥0.8, and ratio of n-6 to n-3 ≥1.30.     

Improvement of solubility and Emulsifying Properties of Soy Protein Isolates by Glucose Conjugotion

Soy Protein Isolate(SPI)was modified with glucose(G-) through the amino-carbonyl reaction (Maillard Reaction).Solubility and emulsifying properties of the modified proteins were investigated.G-SPI conjugate was highly soluble at wide pH while untreated SPI was hardly soluble especially at pH4-PH6.Solubility of modified SPI showed the resistance to heat treatment and high ionic concentration.Emulsifying activity and emulsion stability of G-SPI conjugate was much higher than those of native SPI were.Comparing with some commercial emulsifiers,the G-SPI conjugate showed as good or better emulsifying properties in high salt concentration and in neutral pH.SDS-PAGE was also used to confirm the formation of G-SPI conjugate.     

Influence of salts on hydrolysis of β-lactoglobulin by free and immobilised trypsin

Immobilised trypsin is an alternative to free trypsin for producing protein hydrolysates with increased functionalities. However, the influence of hydrolytic conditions on this process remains unclear. The influence of salts on β-lactoglobulin (β-Lg) hydrolysis by free and immobilised trypsin was compared. For both forms of trypsin, 0.1 m Tris accelerated the release of most final peptides except f (71–75), and had no significant effects on the hydrolysis of intact β-Lg. Increasing NaCl concentrations from 0 to 0.02 m increased the degree of hydrolysis (DH) by 22.4% for free trypsin versus 62.1% for immobilised trypsin. The presence of 0.1 or 0.5 m NaCl hindered the release of peptides associated with the breakdown of intact protein. This led to 2–4 fold decreases in depleting intact β-Lg and DH, except immobilised trypsin at 0.1 m NaCl (DH increased by 44.3% versus without NaCl). Potential mechanisms underlying the effects of salts are discussed.     

Characterization of core–shell structures formed by zein

Core–shell structures are of interest for encapsulation purposes in the food, pharmaceutical, and cosmetics industries. A number of wall materials are in use including carbohydrates, lipids, and proteins. Zein, the prolamin of corn, is capable of self-assembly into microspheres, potentially useful in encapsulation and delivery systems. In previous work, zein was observed to form core–shell structures with citral and lime by evaporation induced self-assembly of zein in ethanol–water. The objectives of this work were to investigate the effect of mass ratio of core to shell materials and ethanol content of the solvent on encapsulation microstructure. Scanning electron microscopy (SEM), focused ion beam (FIB), and transmission electron microscopy (TEM) were used for structure characterization. Raman spectroscopy and FTIR were used to detect citral or lime in the core. Microstructure images suggested that core–shell structures of zein loaded with lime flavor were formed only in a narrow range of core to shell mass ratio. Ethanol content also affected structure formation.     

Fabrication of κ-carrageenan fibers by wet spinning: Addition of ι-carrageenan

Taking advantage of the gelation process of κ-carrageenan, we have developed a wet-spinning process to fabricate micro-scale fibers from κ-carrageenan. Effects of three important spinning parameters, i.e. coagulation bath composition, spinning rate and post-spinning mechanical drawing, on fiber morphological and tensile properties have been discussed. In the present report, we studied the addition of ι-carrageenan on thermal and rheological properties of the bicomponent gels and the fibers spun from them. It was found that κ- and ι-carrageenan underwent phase separation in the bicomponent gel. Upon addition of ι-carrageenan, the diameter and compliance of the blend fiber was increased.     

Formation of oligosaccharides from whey UF-permeate by enzymatic hydrolysis — analysis of factors

Two-level fractional factorial experiments were designed to study effects of enzyme (0.05 and 0.1%) and initial lactose concentrations (L_o: 14 and 23%), pH (5.0 and 7.0) and temperature (35 and 45 °C) on enzymatic formation of oligosaccharides (OS) from whey UF-permeate in a batch reactor. β-d-galactosidase from Aspergillus oryzae (A), Kluyveromyces lactis (B) and K. fragilis (C) were compared. Hydrolysis with B and C gave comparable yields which were higher than that from A at identical conditions. L_o did not influence reaction time, but concentration of OS significantly increased by increasing L_o for all enzymes. Increasing L_o reduced the yield after hydrolysis with A and B, but improved the yield for C. L_o had negligible effect on degree of hydrolysis (DH) for A and C. However, increasing L_o significantly lowered DH for B. Increasing enzyme concentration significantly reduced reaction time for A, but it had no effect on that for B and C. DH significantly decreased after increasing concentration of A. Consequently, OS and yield were reduced. Applying B and C at higher concentration improved DH, OS and yield. Increasing temperature or pH reduced DH, OS and yield for A, but increased the responses for B and C.     

Properties of oil-in-water emulsions stabilized by β-lactoglobulin in simulated gastric fluid as influenced by ionic strength and presence of mucin

The effects of ionic strength (0–150 mM NaCl) and the presence of mucin (0.1 wt%) on the properties of oil-in-water emulsions [20.0 wt% soy oil, stabilized by 1.0 wt% β-lactoglobulin (β-lg)] under simulated gastric conditions (with/without 0.32 wt% pepsin at 37 °C, with continuous shaking at approximately 95 rev/min for 2 h) were investigated. Changes in Z-average diameter, ζ-potential and microstructure were determined as a function of incubation time. The emulsions mixed with simulated gastric fluid (SGF) (without added pepsin) were stable at low ionic strength (≤50 mM NaCl) but showed some aggregation at high ionic strength (≥150 mM NaCl). Extensive droplet flocculation with some degree of coalescence was observed in emulsions with 0.32 wt% added pepsin, the flocculation being potentially accelerated in the presence of NaCl. The addition of 0.1 wt% mucin resulted in a greater extent of flocculation, possibly because of non-specific binding of mucin to the positively charged β-lg emulsion droplets. Ionic strength and the presence of mucin had a significant influence on the rate of hydrolysis of β-lg by pepsin. The behaviour of the emulsion in SGF was predominantly driven by electrostatic interactions, which varied as a function of digestion time, ionic strength and the presence of pepsin and mucin.     

Proteolysis of bovine αs2-casein by chymosin

Proteolysis of bovine α_s2-casein by chymosin (E. C. 3.4.23.4) in solution in 100mm Na phosphate buffer, pH 6.5, at 30 °C was studied by reversed-phase (RP)-HPLC and urea-polyacrylamide gel electrophoresis (PAGE). Chymosin hydrolyzed α_s2-casein in solution to eight peptides detectable by urea-PAGE. Peptides soluble in acetate buffer, pH 4.6, were isolated by RP-HPLC on a C₁₈ column using an acetonitrile/water gradient and identified from their N-terminal amino acid sequence. The chymosin cleavage sites were at the bonds Phe₈₈-Tyr₈₉, Tyr₉₅-Leu₉₆, Gln₉₇-Tyr₉₈, Tyr₉₈-Leu₉₉, Phe₁₆₃-Leu₁₆₄, Phe₁₇₄-Ala₁₇₅ and Tyr₁₇₉-Leu₁₈₀. Chymosin cleavage sites were restricted to the hydrophobic regions of the molecule. The bond-type in α_s2-casein cleaved by chymosin was in agreement with that found to be susceptible to chymosin in other caseins. The primary site of chymosin action on α_s2-casein appeared to be at Phe₈₈-Tyr₈₉.     

Comparison of encapsulation properties of major garlic oil components by hydroxypropyl β-cyclodextrin

The major garlic oil (GO) components were encapsulated by hydroxypropyl β-cyclodextrin (HP-β-CD), and their encapsulation properties were compared in this study. Our results showed that the optimal conditions for the GO encapsulation were as follows: weigh ratio at 12:88 (GO: HP-β-CD), pH at 5.0, and inclusion time at 4 h. Furthermore, results from the phase solubility curves indicated that two components of diallyl disulfide (DADS) and diallyl trisulfide (DATS) had higher water solubility caused by HP-β-CD and that DATS-HP-β-CD complex had higher stability constant (K _1:1 = 6.702 × 10⁵) and more significant change in free energy ( \Updelta G_\textCOMP^* \Updelta G_{\text{COMP}}^{*}  = −3.496 × 10⁴) than DADS-HP-β-CD. These findings indicated that DATS was better suited to be encapsulated by HP-β-CD compared to DADS. The result finally obtained from gas chromatography–flame ionization detector (GC–FID) confirmed the easier inclusion properties of DATS by HP-β-CD.     

Enhancement of the hydrolysis activity of β-galactosidase from Geobacillus stearothermophilus by saturation mutagenesis

Thermostable β-galactosidase (BgaB) from Geobacillus stearothermophilus is characterized by its thermoactivity in the hydrolysis of lactose to produce lactose-free milk products. However, BgaB has limited activity toward lactose. We established a method for screening evolved mutants with high hydrolysis activity based on prediction of substrate binding sites. Seven amino acid residues were identified as candidates for substrate binding to galactose. To study the hydrolysis activity of these residues, we constructed mutants by site-saturation mutagenesis of these residue sites, and each variant was screened for its hydrolysis activity. The first round of mutagenesis showed that changes in amino acid residues of Arg109, Tyr272, and Glu351 resulted in altered hydrolysis activity, including greater activity toward ortho-nitrophenyl-β-d-galactopyranoside (oNPG). The mutants R109V and R109L displayed changes in the optimum pH from 7.0 to 6.5, and the mutant R109V/L displayed different substrate affinity and catalytic efficiency (k_cat/K_m). Mutant R109G showed complete loss of BgaB enzymatic activity, suggesting that Arg109 plays a significant role in maintaining hydrolysis activity. The optimum pH of mutant E351R increased from 7.0 to 7.5 and this mutant showed a prominent increase in catalytic efficiency with oNPG and lactose as substrates.     

Fast Preconcentration of Pesticide Residues in Oilseeds by Combination of QuEChERS with Dispersive Liquid–Liquid Microextraction Followed by Gas Chromatography-Mass Spectrometry

A fast, efficient, and simple method for determination of pesticide residues in pumpkin seeds has been developed combining QuEChERS and dispersive liquid–liquid microextraction (DLLME) followed by gas chromatography and mass spectrometry (GC-MS). Parameters affecting the DLLME performance such as solvent selection and volume of extractive and dispersive solvent, salt effect, and extraction time were studied. Under the selected conditions (50 μL extractive solvent chloroform, 1 mL QuEChERS extract, and 3 mL water), the developed method was validated. Linearity was evaluated at nine concentrations in the broad range of 0.1–500 μg/kg with correlation coefficients from 0.9842 to 0.9972. The relative standard deviations at lowest calibration level varied from 0.3 to 22 %. Under the optimum conditions, an enrichment factor was 6–17-fold and detection limits 0.01–12.17 μg/kg were achieved. Finally, the developed and validated method was successfully applied for the extraction and determination of pesticide residues in 16 real samples with 2 positive findings below maximum residue limits (MRL). Limits of detection (LODs) of the proposed method are below the MRLs established by the European Union.     

Folate composition of 10 types of mushrooms determined by liquid chromatography–mass spectrometry

White button, crimini, shiitake, maitake, enoki, oyster, chanterelle, morel, portabella, and uv-treated portabella mushrooms were sampled from U.S. retail outlets and major producers. Folate [5-methyltetrahydrofolate (5-CH₃-H₄folate), 10-formyl folate (10-HCO-folate), 5-formyltetrahydrofolate (5-HCO-H₄folate)] was analysed using a validated LC–MS method in four composites of each product, including an in-house mushroom control composite and a reference material (BCR 485 Lyophilised Mixed Vegetables). Chanterelle and morel had the lowest total folate (2–6 μg/100 g), oyster had the highest (mean, 44.2 μg/100 g); other types contained 12.4 μg/100 g (shiitake) to 29.8 μg/100 g (vitamin D-enhanced portabella). Enoki and oyster had almost exclusively 5-CH₃-H₄folate. Morel and chanterelle contained predominately formyl folates. Other species had similar amounts of 5-CH₃-H₄folate and formyl folates. Enoki, oyster, and shiitake, unlike all others, had low to non-detectable 10-HCO-folate (<1 μg/100 g). These precise data on the composition of folate vitamers in different types of mushrooms will facilitate assessment of the dietary contribution of naturally occurring folate.     

The Prevention and Elimination of Insects in Storage of wheat Grains by Gamma Irradiation

This article deals with radiation sensitivity and lethal dose on the main cereal insects such as:Oryzaephilus Surinamensis Linnaeus(OSL),Sitophilus Oryzae Linnaeus(SOL),Rhizopertha Dominica Fabricius(RDF)and Tribolium Castaneum Herbst(TCH)in China.The experiments showed that there was a difference in radiation sensitivity.Among four insects species,OSL had the least tolerance;the second was RDF;the next was SOL and TCH had the biggest tolerance.After radiation with 0,15KGY-0.3KGY all OSL were dead in 2 weeks,all SOL and RDF dead in 3 weeks and all TCH dead in 4 weeks.All eggs could not be incubated with 0.1KGY radiation dose and adults insects had no bearing ability with 0.1 KGY radiation.There was no negative effect on germination potential and germination percentage of wheat with less than 1.0 KGY radiation.The article deals with bioeffect and lethal dose on OSL,SOL,RDF and TCH by ^60Co gamma ray radiation and with wheat germination potential and germination percentage.The economical and reasonable conditions are found for the development of science and technology and to provide the basis for the commercial operation.     

Optimization of Cultivation Conditions for the Production of γ-Cyclodextrin Glucanotransferase by Bacillus macorous

Abstract

The production of γ-cyclodextrin glucanotransferase (γ-CGTase) from Bacillus macorous WSH02-06 was optimized in shake flasks using conventional sequential techniques and statistical experimental design. Effects of nutrients including carbon and nitrogen sources, cation ions, initial pH, and temperature on γ-CGTase production were investigated. Corn starch, peptone, Mn²⁺, and Zn²⁺ were found to be essential for obtaining high γ-CGTase activity and biomass. The promoting effect of manganese and zinc on enzyme production has not been reported previously. According to the results of orthogonal array experiment, the optimal culture medium for high γ-CGTase activity was determined. Maximal γ-CGTase activity obtained in the optimized culture broth was about 250?U/mL which was 10-fold higher than that obtained in the basal medium. Time course of cell growth and γ-CGTase production in a 7 l fermenter showed that enzyme production was growth associated, and that maximum γ-CGTase activity reached 277?U/mL in 17?h.     

Modeling the Enhanced Thermal Inactivation of Cronobacter sakazakii by Inclusion of “Parabens”

The goal of this study is to develop mathematical models that describe the enhanced thermal inactivation of Cronobacter sakazakii by the inclusion of “parabens”. The key parameters include heating temperature, parabens concentration, and the length of parabens’ alkyl side chains. The heating trials were conducted in a submerged coil apparatus using Brain Heart Infusion (BHI) as the model heating menstruum. The results clearly demonstrated that a significant enhancement of thermal inactivation is concentration dependent and increases with increasing alkyl chain length. Once data sets are complete, primary and secondary models will be developed for prediction of thermal inactivation parameters.     

Addition of α-ketoglutarate enhances formation of volatiles by Staphylococcus carnosus during sausage fermentation

The effect of leucine and α-ketoglutarate addition on transamination of branched-chain amino acids was studied in model minces inoculated with Pediococcus pentosaceus and Staphylococcus carnosus. Leucine addition changed the ratio of volatile breakdown products of leucine, isoleucine and valine but not the total amount of the volatiles and it was concluded that the amount of free amino acids does not limit transamination of amino acids. The addition of α-ketoglutarate resulted in increased levels of methyl-branched aldehydes and insignificant positive changes in methyl-branched acid production. The results were verified in real fermented sausages with no, low (0.09% w/w) and high (0.36% w/w) addition of added α-ketoglutarate since the levels of the flavour intensive methyl-branched aldehydes and acids were drastically increased in sausages added α-ketoglutarate. The catabolism of phenylalanine was also induced by α-ketoglutarate and there were further indications of increased transamination of aspartate. A triangular test showed that the flavour of sausages with no and low addition of α-ketoglutarate could be clearly distinguished from one another. Altogether the results presented in this paper point to glutamate dehydrogenase, the enzyme catalyzing regeneration of α-ketoglutarate, as a key enzyme in catabolism of amino acids and thereby also in aroma formation during sausage processing.