Application of a Plasma Reactor to Modify Egg Ovalbumin for Improved Solubility

ABSTRACT: A Plasma reactor was used to change the surface structure of ovalbumin and enhance solubility. The gases used to generate the plasmas were air, argon, nitrogen and oxygen. Ovalbumin was exposed to the gases for 10, 20, 30, 60, and 120 min. Unexposed ovalbumin served as the control. Samples were examined by polyacrylamide gel electrophoresis (PAGE), spectrophotometry, fluorometry, differential scanning calorimetry, and circular dichroism. Solubility and hydrophobicity of ovalbumin increased with exposure time, whereas endothermic enthalpy, onset PAGE. No change in CD spectra resulted from the plasma treatment. The structural changes were regarded as conformational adaptability.     

Moiety and linker site heterologies for highly sensitive immunoanalysis of cyprodinil in fermented alcoholic drinks

Cyprodinil is a new-generation anilinopyrimidine fungicide widely used in crop protection and frequently found in fruits. In this study, novel derivatives of cyprodinil with linker site heterologies were synthesized and employed in order to produce antibodies with enhanced affinity. Moreover, moiety-heterologous haptens were designed and prepared for assay sensitivity improvement. Two competitive enzyme-linked immunosorbent assays for the analysis of this active substance were developed using direct and indirect formats, achieving IC₅₀ values around 0.15 μg/L. Analytical figures of merit and usability of the optimized assays were evaluated with wine and cider as model food processed matrices. The obtained recoveries were from 90% to 120%, and the limit of quantification was in the 1–5 μg/L range. Finally, a monitoring study (n = 150) was performed to estimate the occurrence and the concentration of cyprodinil in commercial wine and cider products from different origins. We found that 28% of the analysed wine samples contained cyprodinil residues at levels higher than 5 μg/L.     

鸡卵清白蛋白的示差扫描量热法研究

本文用示差扫描量热法（ＤＳＣ）研究了鸡卵清白蛋白的热变性，以及溶剂对变性的影响。给出了该蛋白质在Ｈ２Ｏ、甲醇和乙二醇水溶液中变性的热力学参数值。实验结果表明，鸡卵清白蛋白热变性过程的转变温度和转变焓与样品的来源和溶剂种类有密切的关系。探讨了这种溶剂对蛋白质构象变化影响的机理。     

In-vitro rates of rumen proteolysis of ribulose-1,5-bisphosphate carboxylase (rubisco) from lucerne leaves,and of ovalbumin,vicilin and sunflower albumin 8 storage proteins

Laboratory systems were developed, based upon in-vitro incubations with rumen fluid, to examine the rate of proteolysis (ie degradation) of sunflower albumin 8 (SFA 8; 24% sulphur amino acids; SAA) from sunflower seeds, ovalbumin (6% SAA) from chicken egg white, ribulose-1,5-bisphosphate carboxylase (Rubisco; 3% SAA) from lucerne leaves, and vicilin (0% SAA) from pea seeds. After fractionation by SDS-PAGE, proteins were analysed by either Western blotting, using specific antibodies (SFA 8, vicilin and ovalbumin) or by Coo-massie blue staining (Rubisco). Proteolysis of the large subunit of Rubisco occurred very quickly and as two components, with half-lives of 11.6 and 1.5 h. The small subunit of Rubisco was more resistant to degradation, with a half-life of 17.3 h. Vicilin was degraded extremely rapidly (half-life 0.16 h). SFA 8 and ovalbumin both showed resistance to degradation, but by two different mechanisms. Ovalbumin was not degraded at all during the initial 16 h of incubation, but then degraded with a half-life of 8.7 h. SFA 8 (mol wt 12 100) disappeared very rapidly, with a half-life of 3.0 h. The degradation of SFA 8 was associated with the appearance of a polypeptide (mol wt 8000), which was extremely resistant to degradation, with a half-life of 69.3 h. It was concluded that both the number of disulphide linkages and tertiary structure were important in determining resistance of proteins to rumen degradation, and that incorporation of SFA 8 and ovalbumin proteins into forages using genetic engineering techniques would be likely to increase the quantity of SAA by passing rumen fermentation.     

Rheological behavior and organoleptic effects of ovalbumin addition in yogurt mousse production

Yogurt mousse is a novel, high added-value dairy product that has been well received by the market. This paper presents a study of the effect of the addition of ovalbumin to the product on its rheological and organoleptic qualities. The ovalbumin was previously separated from egg white with a high grade of purity using an ion exchange resin synthesized by the authors. Diverse rheological tests at different temperatures and corresponding sensorial assessments were conducted to compare samples without and with added ovalbumin.The obtained results confirm that the product is viscoelastic and combines the properties of foams and emulsions; the elastic component is greater than the viscous component. Moreover, at temperatures ranging from 5 to 15°C, a usual interval of consumption, the product behaves rheologically in a similar way. Conversely, the addition of ovalbumin under the assayed conditions also makes the elastic character of the product increase at a given temperature. Finally, the sensorial assessment tests and determinations of stability and volume yield enabled us to verify that the addition of ovalbumin at an amount of 1.3% hardly alters the stability, resistance to shear stress, or the texture and improves the degree of foaming. Therefore, the product with additive is of good commercial quality.     

Nanoscale Size Control of Protein Aggregates

Herein, a novel method to synthesize soluble, sub‐micrometer sized protein aggregates is demonstrated by mixing native and denatured proteins without using bacteria and contaminating proteins. Ovalbumin (OVA) is employed as a model protein. The average size of the formed aggregates can be controlled by adjusting the fraction of denatured protein in the sample and it is possible to make unimodal size distributions of protein aggregates. OVA aggregates with a size of ～95 nm are found to be more immunogenic compared to native OVA in a murine splenocyte proliferation assay. These results suggest that the novel method of engineering size specific sub‐micrometer sized aggregates may constitute a potential route to increasing the efficacy of protein vaccines. The protein aggregates may also be promising for use in other applications including the surface functionalization of biomaterials and as industrial catalysis materials.     

蛋清中ACE抑制肽的筛选及其作用机制

表征来自卵清蛋白中的血管紧张素转换酶（angiotensin converting enzyme,ACE）抑制肽。基于在线程序,采用胃肠道蛋白酶对卵清蛋白进行模拟酶切。对获得的三肽毒性、溶解性和ADME性质进行预测,以ACE为蛋白靶标,筛选出能够与ACE紧密结合的三肽,对这些三肽的活性进行验证。结果表明：获得了一种新的ACE抑制三肽CIK,其IC50值为（161±0.06）μmol/L。CIK-ACE复合物通过5 个常规氢键,2 个碳氢键和4 个盐桥达到稳定状态。CIK最佳的对接构型可以与ACE的氨基酸残基Gln281、His353、Ala354、Glu376、Val380、His383、Glu384、Lys511、His513、Tyr523和Phe527形成相互作用。最佳的药效团模型由3 个氢键受体,一个疏水中心和一个正电荷中心组成,三肽CIK可与其中4 个特征相匹配。     

一步法分离鸡蛋清中三种主要过敏原的研究

为快速高效分离鸡蛋中主要过敏原开展鸡蛋过敏的研究,本实验采用DEAE-Sepharose FF 色谱柱一步分离鸡蛋中的三种主要过敏原卵白蛋白、卵转铁蛋白和溶菌酶,通过SDS-PAGE 比对,纯度均高于相应标准品纯度,得率分别为87.66%、55.87% 和92.80%。本实验操作简便、设备简单、分离效率高,是一种分离鸡蛋过敏原的好方法。     

卵白蛋白与油酸相互作用的光谱学分析

运用荧光光谱、同步荧光光谱、傅里叶变换红外光谱和圆二色谱研究卵白蛋白与油酸相互作用的方式及机理。结果表明,油酸与卵白蛋白相互作用导致卵白蛋白荧光猝灭,猝灭方式由动态猝灭（未加热处理）转变为静态猝灭（加热处理）,其相互作用力为疏水作用和氢键,且随着油酸浓度的增大,相互作用力增强。与油酸相互作用后,卵白蛋白酪氨酸残基和色氨酸残基的荧光光谱蓝移,α-螺旋和β-转角的含量减少,β-折叠含量增加;而无规卷曲的含量增大（油酸与卵白蛋白物质的量比为10∶1）。综上所述,油酸与卵白蛋白间通过疏水作用和氢键发生相互作用,导致卵白蛋白二级结构发生变化。     

鸡蛋清卵白蛋白酶解工艺优化及其结构性质

研究鸡蛋清卵白蛋白的酶解工艺及其结构性质,以水解度为指标,确定最佳酶源为碱性蛋白酶,其水解度为26.55%,显著优于其他蛋白酶（P＜0.05）。以碱性蛋白酶酶解鸡蛋清卵白蛋白,采用单因素和五元二次正交旋转试验研究酶解工艺;针对酶解前后卵白蛋白的功能特性进行分析,并采用紫外扫描、傅里叶红外变换光谱、差示扫描量热针对卵白蛋白及其酶解产物进行结构表征。结果发现,碱性蛋白酶酶解鸡蛋清卵白蛋白最佳工艺条件为反应温度52.5 ℃、反应时间5 h、pH 8.25、酶用量5 500 U/g、底物添加量5%,此条件下水解度为27.88%。酶解后产物表面巯基含量降低了3.6 mol/（L·g）,溶解度大幅度提高,起泡性降低了18.18%,泡沫稳定性降低了20.24%,乳化活性指数升高了13.56 m2/g,乳化稳定性提高了10.46%。同时,酶解后的卵白蛋白肽链发生了裂解,有序的二级结构被破坏,暴露出更多氨基酸残基,α-螺旋略有减少,β-转角相应增加,亲水基团也相应的增加。