The effect of limited proteolysis on canola protein gelation

The objective of this study was to improve canola protein gelation properties through limited enzymatic hydrolysis with trypsin, ficin and bromelin either alone or in combination with the cross-linking enzyme transglutaminase (TG). Proteolysis alone was found to reduce gel strength to below that of the control. This was accompanied by a decrease in molecular weight, observable through SDS-PAGE analysis. Micrographs corroborated the rheological data and showed that protease application brought about a loss of structure. In contrast, limited proteolysis was shown to be a suitable pretreatment to cross-linking with TG, a frequently used cross-linking enzyme. Previous research suggests and this study corroborates that opening the protein structure prior to TG treatment, can enhance its effectiveness. Gels, treated with both a protease and TG, were significantly stronger than those treated with TG alone. It was concluded that while limited proteolysis does not improve canola protein gelation on its own, it can significantly improve gelation properties when combined with TG.     

Improvement of canola protein gelation properties through enzymatic modification with transglutaminase

Enzymatic modification with transglutaminase (TG) was used to enhance the gelation of canola protein isolate (CPI) and thus improve its potential as a food ingredient. The effects of CPI concentration, TG concentration, treatment temperature and treatment time on CPI gelation properties were evaluated. A texture analyzer, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and scanning electron microscopy were used to characterize the resulting networks. The protein concentration, amount of TG, and treatment temperature were found to significantly impact gel strength. It was found that gelation can be improved by increasing the amounts of protein and TG and by keeping the treatment temperature close to 40 °C. SDS-PAGE showed that cross-linking of subunits occurred through TG treatment thus helping to explain the increase in gel strength observed during texture analysis. Micrographs further corroborated the trends noted during rheological studies.     

Selenomethionine biotransformation and incorporation into proteins along a simulated terrestrial food chain

Selenium is an essential trace element in vertebrates, but there is a narrow concentration range between dietary requirement and toxicity threshold. Although a great deal is known about the biochemistry of Se from a nutritional perspective, considerably less attention has been focused on the specific biochemistry of Se as an environmental toxicant. Recent advances in hyphenated analytical techniques have provided the capability of quantifying specific chemical forms of Se in biological tissues as well as the distribution of Se among macromolecules. We applied liquid chromatography coupled to inductively coupled plasma mass spectrometryto investigate biotransformations of selenomethionine along a simulated terrestrial food chain consisting of selenomethionine exposed crickets (Acheta domesticus) fed to western fence lizards (Sceloporus occidentalis). Evidence was obtained for selenomethionine biotransformation as well as for sex-specific differences in the metabolism of Se compounds and their subsequent incorporation into proteins in the lizard. The results demonstrate the complexities involved in trophic transfer of Se due to the potential for extensive biotransformation and the species- and even sex-specific nature of these biotransformations.     

Purification of angiotensin I-converting enzyme-inhibitory peptides from the enzymatic hydrolysate of defatted canola meal

Defatted canola meals from seeds of different processing origins were hydrolyzed by Alcalase to give hydrolysates that inhibited angiotensin converting enzyme (ACE) activity. Heat treated meals yielded protein hydrolysates with 50% ACE-inhibitory concentrations of 27.1 and 28.6 μg protein/ml compared with 35.7 and 44.3 μg protein/ml for the none-heat treated meals. Separation of the hydrolysate on a Sephadex G-15 gel permeation column (GPC) yielded a fraction with an IC₅₀ value of 2.3 μg protein/ml. Amino acid analysis showed that the GPC fraction contained 45% content of aromatic amino acids in comparison to 8.5% of the hydrolysate. Two peptides, Val-Ser-Val (IC₅₀ = 0.15 μM) and Phe-Leu (IC₅₀ = 1.33 μM) were purified, and located in the primary structure of canola napin and cruciferin native proteins. The results suggest that canola protein hydrolysate is a potential ingredient for the formulation of hypotensive functional foods.     

Emulsifying properties of proteins extracted from Australian canola meal

Canola protein albumin fraction, globulin fraction, and canola protein isolate (CPI) were compared to commercial soy protein isolate (SPI) in terms of their emulsifying properties at various pH values. The globulin fraction had higher emulsifying capacity (EC), higher emulsifying activity index (EAI), and the droplet size of emulsions it stabilized was consistently smaller irrespective of pH compared to albumin fraction or CPI. In comparison to SPI, globulin fractions also had higher EC at all pH values tested, higher EAI at acidic pH, and smaller or comparable average emulsion droplet size at both pH 4 and 7. The stability of canola protein based emulsions were comparable to those of SPI based emulsions at most pH values (except the emulsion stabilized by the CPI at pH 4), with no significant (p > 0.05) changes in droplet size during storage for up to 7 days at room temperature. These emulsions, however, experienced separation into the emulsion and serum phases after 24 h storage at room temperature with the exception of CPI- and SPI-stabilized emulsions at pH 9. This study demonstrates the comparable emulsifying properties (forming or stabilizing) of some canola proteins to commercially available SPI, suggesting the potential use of canola proteins in food applications.     

蛋黄酱的稳定性研究

文章从蛋黄酱乳化液的稳定性、脂类的氧化以及主要风味成分的稳定性三个方面详细介绍了蛋黄酱的稳定性。     

Four-base codon-mediated incorporation of non-natural amino acids into proteins in a eukaryotic cell-free translation system

Various four-base codons have been shown to work for the introduction of non-natural amino acids into proteins in an Escherichia coli cell-free translation system. Here, a four-base codon-mediated non-natural mutagenesis was applied to a eukaryotic rabbit reticulocyte cell-free translation system. Mutated streptavidin mRNAs containing four-base codons were prepared and added to a rabbit reticulocyte lysate in the presence of tRNAs that were aminoacylated with a non-natural amino acid and had the corresponding four-base anticodons. A Western blot analysis of translation products indicated that the four-base codons CGGU, CGCU, CCCU, CUCU, CUAU, and GGGU were efficiently decoded by the aminoacyl-tRNAs having the corresponding four-base anticodons. In contrast, the four-base codons AGGU, AGAU, CGAU, UUGU, UCGU, and ACGU were not decoded. The stop codon-derived four-base codons UAGU, UAAU, and UGAU were found to be inefficient, whereas the amber codon UAG and opal codon UGA were efficient for the incorporation of non-natural amino acids. The application of the expanded genetic code in a eukaryotic cell-free system opens the possibility of a four-base codon-mediated incorporation of non-natural amino acids into proteins in living eukaryotic cells.     

不同酶处理对鸡蛋清中卵白蛋白致敏性的影响

选取了木瓜蛋白酶、胰蛋白酶、中性蛋白酶、风味蛋白酶和动物蛋白水解酶,并利用单因素实验,以水解度为指标,确定各种蛋白酶对蛋清蛋白最佳的酶解工艺参数。在此基础上,对蛋清蛋白进行水解,通过SDS-PAGE和间接竞争ELISA检测各蛋白酶蛋清水解液,比较各蛋白酶对蛋清中卵白蛋白抗原性降低效果。SDS-PAGE结果显示木瓜蛋白酶、中性蛋白酶和动物水解蛋白酶对卵白蛋白的降解效果较明显,胰蛋白酶和风味蛋白酶的降解效果不明显;间接竞争ELISA检测结果表明,木瓜蛋白酶和中性蛋白酶对卵白蛋白抗原性的降低作用更有效,分别达到了57.8%和49.6%,其他几种酶都在30%左右。因此可以选用木瓜蛋白酶和中性蛋白酶对研究开发脱敏鸡蛋食品做进一步研究。      

A pursuit of the functional nutritional and bioactive properties of canola proteins and peptides

This review focuses on updated information about canola proteins and peptides, their functional, nutritional, and bioactive properties, safety aspects, and potential application in foods. Attention is paid to gelation, emulsion, thermal, and water holding capacities of crude and pure proteins and peptides isolated from canola meal. Various factors affecting these properties are discussed. This paper provides an overview of use of canola meal as a protein source in animal diets and their digestibility in vivo. Their effects on a range of health outcomes including ACE inhibition, hypocholesterolemic effects, cancer prevention, anti-viral and anti-diabetic properties are reviewed on the basis of the available in vitro and in vivo animal and human data. The review also focuses on the safety aspects and selected food applications of canola proteins and peptides.     

酶水解大豆蛋白物理化学性质的综合表征

大豆分离蛋白分别用AS1.398中性蛋白酶和黑曲霉3.4310酸性蛋白酶在不同条件下水解后,测定其水解度(DH)、三氯乙酸(TCA)溶解度、等电点(IP)溶解度,以及不同分子量区段的相对含量等13个参数.通过主成分分析将这些参数线性组合为4个主成分,但是中性蛋白酶和酸性蛋白酶水解产物参数在主成分中的组合方式不同.以主成分为指标对全体样品进行聚类分析时,两种酶水解产物各自聚为一类;中性蛋白酶和酸性蛋白酶的水解产物单独进行聚类分析则分别得到5个和4个子类,其中包括水解条件差别较大的水解产物,说明水解工艺参数之间存在某种协同或互补效应.     

Opioid peptides derived from in-vitro proteolysis of bovine whey proteins

The formation of opioid peptides by in-vitro proteolysis of whey proteins was investigated. Bovine β-lactoglobulin (β-LG) or -lactalbumin (-LA) were predigested with pepsin and subsequently treated with either trypsin (Try) or trypsin+chymotrypsin (Try+Chy). For separation and identification of the peptides. HPLC chromatography, protein sequencing and amino acid analysis were used. Identified peptides were synthesized by Peninsula Laboratories Europe Ltd. UK. Binding to rat brain homogenates was tested against ³H-naloxone. The effects of the peptides on smooth muscle were tested in coaxially stimulated guinea pig ileum in vitro. Digestion of β-LG with pepsin plus Try, or Try+Chy yielded Tyr-Leu-Leu-Phe (β-lactorphin). Proteolysis of -LA with pepsin alone produced Tyr-Gly-Leu-Phe (-lactorphin) although a higher degree of hydrolysis was achieved by addition of Try. Among hydrolysates of whey proteins at least -lactorphin exerted a weak but continuous opioid property both in terms of receptor binding and smooth muscle effects.     

Optimization of the enzymatic modification of egg yolk by phospholipase A2 to improve its functionality for mayonnaise production

Egg yolk (EY) was enzymatically modified by phospholipase A₂ to improve its functional properties for mayonnaise production. The optimum reaction conditions predicted by response surface methodology were enzyme concentration of 7432 LEU for 74 min in the presence of 11.5% (w/w) salt. The predicted values were in good agreement with the experimental values. The mayonnaise prepared with 6% (w/w) of the enzymatically modified EY (EM-EY) had no significant differences in overall acceptability or in the perceived intensities of umami, nutty flavor, sourness, oiliness, and rancid flavor as compared to the mayonnaise prepared with 8% (w/w) fresh EY. The stability of the mayonnaise prepared with 6% (w/w) EM-EY was greater than that of the mayonnaise prepared with 10% (w/w) fresh EY. Based on the results, EM-EY, when produced at optimum conditions, has great potential to replace fresh EY.     

High pressure and the enzymatic hydrolysis of soybean whey proteins

The effect of high-pressure (HP) treatment on the hydrolysis of soybean whey proteins by trypsin, chymotrypsin and pepsin was studied. The experiments were carried out at atmospheric pressure (0.1 MPa, for 30 min, at 37 °C) and at HP (100 and 200 MPa for 15 min at 37 °C) before or during the reaction of hydrolysis. The extent of hydrolysis was measured by OPA method and in the extracts from TCA. The results showed that HP treatments increased the hydrolysis in the three enzymes used and 100 MPa was the better pressure to enhance the hydrolysis. Polyacrylamide gel electrophoreses (SDS–PAGE), showed five peptides lower than 14 kDa after hydrolysis by chymotrypsin and trypsin, and 11 peptides by pepsin. Soybean whey proteins, which are industrially discarded, could be used as a source of peptides, with applications as base in some diets.     

X-ray microtomography to study the microstructure of mayonnaise

In this work, the X-ray microtomography (μCT) technique was used for the analysis of fat microstructure and quantification of fat in four types of mayonnaise. The dynamic-mechanical properties of the mayonnaise samples were also studied using a controlled-strain rotational rheometer. Four types of commercially produced mayonnaises, chosen to exhibit variability in terms of visible structure of fat, were used for this experiment: ‘kraft’, ‘calvé’, ‘kraft legeresse’ and ‘calvé-mayò’. Appropriate quantitative three-dimensional parameters describing the fat structure were calculated. With regards to the microstructural and rheological relationship, results from the correlation carried out show that a correlation exists among some microstructural and rheological parameters of the mayonnaise samples. The results from this study also show that μCT is a suitable technique for the microstructural analysis of fat as it does not only provide an accurate percentage volume of the fat present but can also determine its spatial distribution.     

Immunomodulatory peptides obtained by the enzymatic hydrolysis of whey proteins

Whey contains a number of immunomodulatory peptides that are naturally present or that are part of the primary sequence of whey proteins. These peptide sequences can be released during digestion in the gut and can also be produced by in vitro enzymatic hydrolysis. This paper reviews the existing literature on the effect of whey peptides on the specific (lymphocyte activation and proliferation, antibody production, cytokine expression) and non-specific (functions of macrophages, granulocytes and natural killer cells) immune responses. Major discrepancies in the effect of some peptides on lymphocyte proliferation have been reported while the more limited literature on antibody production is less controversial. The effects of whey peptides on hypersensitivity, induction of oral tolerance and response to infections and diseases are also reviewed. The in vitro and in vivo data on the immunomodulatory potential of whey peptides are relatively abundant. However, the potential of whey peptides as effective health ingredients remains to be demonstrated, which may be due to the lack of clinical evidence and the limitations of some in vivo models.     

Influence of enzymatic hydrolysis,pH and storage temperature on the emulsifying properties of canola protein isolate and hydrolysates

The aim of this work was to enhance emulsification properties of canola proteins through enzymatic proteolysis and pH variaton. Canola protein isolate (CPI) and hydrolysates (CPHs) were used to form emulsions at pH 4.0, 7.0 and 9.0 followed by storage at 4 or 25 °C for 7 days. Controlled enzymatic hydrolysis led to increased peptide bond cleavage with time (0.23 g/100 g in CPI to 7.18 g/100 g after 24‐h Alcalase hydrolysis). Generally, oil droplet sizes were smaller for emulsions made at pH 9.0, which suggest better quality than those made at pH 4.0 and 7.0. Trypsin hydrolysate emulsions were the most physically stable at pH 7.0 and 9.0; in contrast, the pepsin hydrolysate emulsions were unstable at all conditions. The results suggest that selective enzymatic hydrolysis could play an important role in enhancing successful incorporation of canola proteins and peptides into food systems as protein emulsifiers.     

Assessment of the residual immunoreactivity of soybean whey hydrolysates obtained by combined enzymatic proteolysis and high pressure

Soybean (Glycine max) whey was hydrolyzed for 15 min using three food-grade proteases (Alcalase, Neutrase, Corolase PN-L) at atmospheric pressure (0.1 MPa) and under high pressure (HP) at 100, 200, and 300 MPa. All hydrolysates were analyzed by SDS-PAGE and their residual immunoreactivity was assessed by immunoblotting using the sera from children allergic to soybean. As shown in SDS-PAGE, Alcalase, Neutrase, and Corolase PN-L produced different patterns of hydrolysis. Each enzyme showed a similar proteolytic activity at atmospheric pressure and at 100 MPa, while an increased degree of hydrolysis was observed at 200 and 300 MPa. No residual antigenicity was observed in the hydrolysates obtained by Alcalase and Corolase PN-L in all considered conditions of hydrolysis. A positive reaction associated with a band having molecular weight of about 70 kDa was observed in the immunoblotting of the hydrolysates obtained with Neutrase at 0.1, 100, and 200 MPa, while no antigenicity was detected for those samples obtained under high pressure, at 300 MPa. These results suggest that high pressure combined with suitable enzymatic activity could be a useful tool for obtaining hydrolysates with low immunoreactivity to be used in special foods (hypoallergenic foods).     

酶法酿造酱油新工艺的探讨

试验采用混合料蒸煮后,冷却直接加入新型复合酶(主要含中性蛋白酶、淀粉酶、糖化酶和纤维素酶),免制曲工艺,混匀入池酶解发酵。新工艺与原低盐固态发酵相比,大大降低了酱油的生产成本和劳动强度。