Surface properties of walnut protein from AOT reverse micelles

The surface chemical composition and microstructure of walnut protein obtained through aqueous buffer and bis(2‐ethylhexyl) sodium sulfosuccinate (AOT) reverse micelles were determined by X‐ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM). The surfaces were characterised by XPS to monitor surface composition. The different components of C 1s, N 1s and O 1s peaks were provided precisely. By comparison with walnut protein surface from aqueous buffer, XPS analysis revealed that the O atomic percentage of walnut protein powder surface from AOT reverse micelles was higher, but the C and N atomic percentages were lower. The N/C ratio of the walnut protein powders using two extraction methods was similar, while significant difference was obtained in the O/N. The obtained results indicated that the reverse micelles could affect the C, O and N components on the surface of walnut protein powder. Protein microstructure after reverse micelle treatment showed many crystals.     

Effects of extraction temperature,ionic strength and contact time on efficiency of bis(2‐ethylhexyl) sodium sulfosuccinate (AOT) reverse micellar backward extraction of soy protein and isoflavones from soy flour

BACKGROUND: Soy protein enriched with isoflavones has been linked to various disease‐preventing and health‐promoting activities owing to the antihypertensive, hypocholesterolaemic, antiobesity and antioxidative properties of isoflavones. The isoflavone profiles of soy‐based products are known to be highly dependent on the various chemical and physical treatments to which the products have been subjected. The aim of this research was to increase the efficiency of backward extraction of soy protein and isoflavones from bis(2‐ethylhexyl) sodium sulfosuccinate (AOT) reverse micelles by studying the effects of extraction temperature, ionic strength of the aqueous stripping solution and contact time on the amounts of soy protein and isoflavones backward extracted from an AOT/H₂O/isooctane reverse micellar system. RESULTS: By modifying the extraction temperature, ionic strength and contact time, 47.0–60.2% of protein, 43.3–68.4% of daidzin, 43.8–74.6% of genistin, 39.0–88.8% of glycitin, 20.8–92.6% of malonyl genistin, 20.2–52.0% of malonyl glycitin, 32.7–75.6% of acetyl genistin, 49.7–76.8% of daidzein and 19.6–38.1% of genistein present in the AOT reverse micellar solution were backward extracted into the aqueous stripping phase. Statistical analysis showed that there were significant linear and interactive effects of temperature and contact time on the backward extraction of daidzin, genistin, glycitin and daidzein. Significant linear and interactive effects of ionic strength and contact time were found in the backward extraction of daidzin and genistin. The backward extraction of genistein was only influenced by contact time and its interaction with temperature. CONCLUSION: This study showed the potential of reverse micelles as a protocol for extracting isoflavones from soy samples for analytical purposes. By modifying the extraction temperature, contact time and ionic strength, soy protein enriched with daidzin, genistin, daidzein and genistein could be produced from soy flour. The results represent an important contribution to current knowledge on utilising reverse micellar extraction in food technology. Copyright © 2007 Society of Chemical Industry     

Protein stability function relations: native β‐lactoglobulin sulphhydryl–disulphide exchange with PDS

Intermolecular sulphhydryl–disulphide exchange with β‐lactoglobulin dimer occurs when this dissociates to form monomers exposing two SH groups. This notion is re‐evaluated in the light of recent structural data suggesting that the degree of SH group exposure in β‐lactoglobulin is unaffected by dissociation. β‐Lactoglobulin was treated with 2,2′‐dipyridyl disulphide (PDS). The rate of sulphhydryl–disulphide exchange was measured at sub‐denaturation temperatures of 25–60 ° C. Parallel studies were conducted by reacting PDS with reduced glutathione (GSH). The SH group of GSH was up to 31 000 times more reactive than β‐lactoglobulin. At pH 7 the reaction activation enthalpy (ΔH^#) and entropy (ΔS^#) was 26 kJ mol⁻¹ and −100 J mol⁻¹ K⁻¹ respectively for GSH. For β‐lactoglobulin, ΔH^# was 157.2 kJ mol⁻¹ and ΔS^# was 254 J mol⁻¹ K⁻¹. At pH 2.6, ΔH^# was 14.4 kJ mol⁻¹ and ΔS^# was −213 J mol⁻¹ K⁻¹ for GSH. The corresponding results for β‐lactoglobulin were 20.3 kJ mol⁻¹ and −147 J mol⁻¹ K⁻¹. These and other thermodynamic results are discussed in terms of the effects of β‐lactoglobulin conformational structure and stability on SH group reactivity. For native β‐lactoglobulin at neutral pH, intermolecular sulphhydryl–disulphide exchange appears to involve the dissociated monomer. SH group activation probably arises from the lower structural stability of the monomer relative to the dimer. At pH 2.6 the mechanism of SH–disulphide exchange does not require protein dissociation and probably involves breathing motions or localised changes in protein structure. © 2000 Society of Chemical Industry     

Effects of AOT reverse micelle on properties of soy globulins

This study was focused on the influence of AOT reverse micelle on physical–chemical properties of 7S and 11S globulins from soy proteins, and compared with aqueous buffer extraction. The results showed that the contents of the surface hydrophobicity and SH groups of 7S and 11S globulins and SS bonds of 11S globulin, using AOT reverse micelle extraction, were augmented, and SS bonds of 7S globulin decreased. The thermal and rheological properties of 7S and 11S globulins extracted using the two methods were studied by differential scanning calorimetry (DSC) and rheometery. It was found that the peak denaturation temperature and heat of transition of 7S and 11S globulins with aqueous buffer extraction were different from that with AOT reverse micellar extraction. The AOT reverse micelle did not affect the gel properties of 11S globulin, while it influenced 7S globulin’s. Hardness, springiness, gumminess, adhesiveness and chewiness of 7S globulin from AOT reverse micelle were lower than that from aqueous buffer extraction, but gumminess was higher.     

核桃蛋白提取方法研究进展

随着人们营养健康意识的深化,消费者对于富含天然活性蛋白成分的油料作物作为潜在优质蛋白质来源的兴趣日益增长。核桃(Juglans regia L.),别名胡桃,为胡桃科胡桃属乔木的果实,营养价值丰富,是我国重要的木本油料。制油后的核桃粕中含有大量蛋白质,但常被用作饲料或丢弃,加工附加值低,造成了资源的严重浪费。核桃蛋白传统提取方法为主要为碱溶酸沉法,随着提取设备及提取技术的进步,更多的新型及组合提取技术被开发且证实优于传统方法。本文详细介绍了核桃蛋白提取的主要方法,包括碱溶酸沉法、超声辅助、酶辅助以及反胶束法,综述了不同提取方法对核桃蛋白质结构及功能的影响,并对其应用前景进行了展望,以期为核桃蛋白质资源的深入研究及开发提供参考。     

Optimization of a novel backward extraction of defatted wheat germ protein from reverse micelles

In this work, a novel backward extraction procedure of defatted wheat germ protein (DWGP) from reverse micelles was explored. Isooctane was recovered by vaporization firstly. Then the remained residue was dissolved in a small amount of KCl solution. The recovery of DWGP was easily performed by the ternary liquid system (acetone: deionized water: isooctane = 15:5:1) precipitation, while most of sulphosuccinic acid bis (2-ethylhexyl) ester sodium salt (AOT) remained in the ternary liquid system. In the end, the precipitation of DWGP was washed with 65% ethanol solution to further remove any residual AOT. The effects of KCl concentration, the amount of KCl solution and pH on the backward extraction efficiency of DWGP were tested. On the basis of single-factor experiments, the optimum backward extraction was achieved by response surface methodology (RSM). When the operation ran under optimized conditions, the backward extraction efficiency of DWGP achieved 80% and the end protein product was completely free of AOT.Industrial relevanceThis experimental result confirmed that this novel backward extraction method had many advantages on the extraction of protein compared to the traditional backward extraction method (changing the conditions of pH and ionic strength in a fresh aqueous phase). This method increased the backward extraction efficiency of defatted wheat germ protein (DWGP) from 57% to 80%, saved the water resource and offered the possibility of precipitating nearly pure DWGP, completely free of surfactant. On the basis of these advantages, it appears that this novel backward extraction technique may have great potential for being scaled-up to a commercially extraction process of protein.     

Production of protein isolates from yellow mustard meals by membrane processes

Two membrane-based processes were tested for protein purification from yellow mustard meals. The starting materials were hexane-defatted meal and CH₃OH/NH₃/H₂O–hexane-extracted meal. The meals were extracted with aqueous alkali. In one process, some of the extracted protein was recovered by isoelectric precipitation, while in the other case the alkaline extract was purified by membrane processes before isoelectric precipitation. In both processes, the protein remaining in the solution was further membrane processed, and dried to produce a soluble protein isolate. Each process recovered over 85% of Kjeldahl nitrogen present in the meal in the form of two protein isolates and a meal residue. Both protein isolates contained more than 85% protein (N×6.25), less than 0.5% phytate, and were essentially free of glucosinolates.     

不同电解质溶液对反胶束萃取花生蛋白的影响研究

研究了KCl、NaCl、LiCl、MgCl2、NaNO3、KNO3、Na2SO4、MgSO4 8种不同的电解质对AOT/正己烷反胶束溶液萃取低温花生粕中花生蛋白的影响,对反胶束的含水量、蛋白质的提取率及通过SDS-PAGE电泳试验对蛋白质的亚基条带进行了比较。试验结果表明,电解质的种类会影响反胶束的含水量;阴离子与阳离子对反胶束溶液萃取大豆蛋白的前萃与后萃都有影响,电解质KCl和NaCl溶液所提取的蛋白质得率较高,分别为54.22%和50.19%;不同的电解质可以影响所得蛋白的亚基组成,可以用来分离不同的蛋白。     

反胶束提取小麦胚芽蛋白的工艺研究

以副产品小麦胚芽为原料,用反胶束法研究了小麦胚芽蛋白的提取条件,包括前萃和后萃。前萃由琥珀酸二(2-乙基己基)酯璜酸钠(AOT)-异辛烷-氯化钾缓冲溶液组成的反胶束体系从小麦胚芽中提取蛋白;后萃先回收异辛烷,少量KCl的缓冲溶液溶解剩余的固形物,最后用丙酮沉淀得小麦胚芽蛋白。通过单因素实验,考查了AOT浓度、萃取时间、加入小麦胚芽粉量、温度、KCl浓度、缓冲溶液pH、W/O对小麦胚芽蛋白前萃率的影响以及KCI浓度、缓冲液pH、缓冲液加入量对小麦胚芽蛋白后萃率的影响。     

Ultrasound‐assisted extraction of ginsenosides in supercritical CO2 reverse microemulsions

Ultrasound‐assisted extraction of ginsenosides from ginseng in supercritical CO₂ reverse microemulsions formed by bis(2‐ethylhexyl) sodium sulfosuccinate (AOT) was studied. Prior to extraction the ginseng was soaked in water for 12 h. It was found that ultrasound significantly enhanced supercritical CO₂ reverse microemulsion extraction. The ginsenoside extraction yield from supercritical CO₂ reverse microemulsion with ultrasound of 20 kHz, 15.2 W cm^?2 and 3/6 s was 2.63 times that without ultrasound at 24 MPa extraction pressure, 45 °C extraction temperature, 4 h extraction time, 5 MPa separation pressure, 55 °C separation temperature and 2 L h^?1 CO₂ flow rate with 140 mL of 0.07 mol L^?1 AOT/ethanol. The maximum extraction yields of ginsenosides from different concentrations of reverse microemulsions were obtained at different ultrasonic intensities. The extraction yield with 20 kHz ultrasound was higher than that with 38 kHz ultrasound at suitably low intensity; however, it was lower at high intensity. The yield improvement may be basically attributed to the mechanical and thermal effects of ultrasound. Copyright © 2006 Society of Chemical Industry     

AOT反胶束体系萃取大豆蛋白质的研究

研究了AOT反胶束萃取大豆蛋白的工艺,针对影响AOT反胶束萃取大豆蛋白质的各种因素如反胶束直径、缓冲溶液pH值、离子强度、温度、萃取时间进行了研究,得出主要的影响因素为反胶束直径、缓冲溶液pH值和离子强度,并得到最佳的萃取条件为:反胶束直径为4.9nm,此时的Wo值为15.8;pH值为6.5;离子强度为0.1mol/L;萃取温度为45℃;萃取时间为20min。在此条件下蛋白质的萃取率为82.57%。     

反胶束萃取豆油对脂肪酸成分的影响

用AOT/异辛烷反胶束体系从全脂豆粉中同时分离大豆蛋白和油脂;考察了预处理、加酶、温度、WO值对脂肪酸(FA)组分的影响。结果表明,反胶束萃取时对FA组分影响不大,除WO外,各因素几无影响,WO对FA影响较大,可能与AOT残留有关。     

酶解辅助反胶束法萃取小麦胚芽蛋白的研究

采用由AOT(琥珀酸二(2-乙基己基)酯磺酸钠)-异辛烷-KCl缓冲液组成的反胶束体系,联合酶解技术提取小麦胚芽蛋白。探索了酶与底物浓度比、时间、缓冲液pH、温度四个因素对蛋白质前萃率的影响。结果表明:酶解辅助反胶束萃取蛋白是可行的,可以显著提高前萃率,最佳酶与底物浓度比为0.23AU/gPro、时间8h、缓冲液pH8.10、温度60℃。     

Conformation Analysis of Soybean Protein in Reverse Micelles by Circular Dichroism Spectroscopy

Far-UV circular dichroism (CD) spectroscopy was used to study the conformation of protein from soybean flour under the influence of various reverse micelle environments and salt treatments, as compared with the protein using aqueous buffer extraction. These proteins exhibited characteristic CD spectra that reflected a considerable amount of residual secondary structures. The CD analysis showed that the protein modification by reverse micellar system was related to loss of α-helix and β-structure and increase of random coil. The ellipticity of protein in bis (2-ethylhexyl) sodium sulfosuccinate (AOT) and sodium dodecyl sulfate (SDS) reverse micelles was higher than in hexadecyl trimethyl ammonium bromide (CTAB) and TrionX-100. The CD spectra in reverse micelles except for SDS at 210 nm had lower intensity than in aqueous solution. Salts could influence the ellipticity of protein at near 194 nm in an order of NaCl > KNO₃> > KCl > NaNO₃ > Na₂SO₄ in monovalent salts and in MgCl₂ > BaCl₂ > CaCl₂ divalent ones. Besides, the ellipticity was totally higher in monovalent salts than in divalent ones.     

Physicochemical properties of soy protein prepared by enzyme‐assisted countercurrent extraction

Synchronous improvements of both nutritional properties and the flavour of protein by optimising preparation processes are highly challenging. The prehydrolysis of soy meals by cocktail enzyme (β‐glucosidase, phytase and acid protease) treatment and subsequent countercurrent extraction were designed based on similar hydrolysis conditions. The composition, flavour volatiles and physicochemical properties of soy proteins were investigated. Compared to alkaline extraction and acid precipitation, enzyme‐assisted countercurrent extraction significantly increased protein yield and carbohydrate content, accompanied by a decrease in protein purity. This protein exhibited larger molecular weight distribution, less flavour volatiles, higher thermal stability and surface hydrophobicity, as evidenced by higher denaturation temperature (T_d) and enthalpy change (ΔH) of protein. The conversion of isoflavone glycosides to aglycone and the partial degradation of phytic acid were observed for enzyme‐prepared soy proteins. These results suggest a feasible protocol for producing a nutrient‐improved soy protein with excellent flavour and thermal stability.     

Walnuts (Juglans regia L): proximate composition,protein solubility,protein amino acid composition and protein in vitro digestibility

Walnuts contained 16.66% protein and 66.90% lipids on a dry weight basis. Non‐protein nitrogen values ranged from 6.24 to 8.45% of the total nitrogen when the trichloroacetic acid concentration was varied within the range 0.25–1.0 M . Albumin, globulin, prolamin and glutelin respectively accounted for 6.81, 17.57, 5.33 and 70.11% of the total walnut proteins. Walnut proteins were minimally soluble at pH 4.0. The majority of total walnut protein polypeptides had estimated molecular weights in the range 12 000–67 000. The Stokes radius of the major protein in walnuts (glutelin fraction) was 66.44 ± 1.39 Å. Lysine was the first limiting essential amino acid in total walnut proteins as well as in the globulin and glutelin fractions. Leucine and methionine plus cysteine were the second limiting essential amino acids respectively for the prolamin and albumin fractions. Hydrophobic and acidic amino acids dominated the amino acid composition in all protein fractions. Native and heat‐denatured walnut glutelins were easily hydrolysed by trypsin, chymotrypsin and pepsin in vitro. © 2000 Society of Chemical Industry     

Extraction and physicochemical properties of soya bean protein and oil by a new reverse micelle system compared with other extraction methods

Reverse micelle extraction is a novel technology for the separation of plant components such as proteins and oil. In this study, sodium bis(2‐ethylhexyl) sulphosuccinate (AOT) reverse micelle system and AOT/Tween 85 reverse micelle system were used to extract soya bean protein and oil from soya bean flour. The physicochemical properties of the protein and oil extracted were investigated and compared with traditional extraction methods. The results showed that the efficiency of forward extraction of soya bean protein using an AOT/Tween 85 reverse micelle system was superior to that using an AOT reverse micelle system at the optimal extraction conditions. In addition, soya bean proteins extracted using reverse micelle extraction had no unordered structure under Fourier transform infrared spectroscopy. The acid and peroxide values of oil products from two reverse micelle extractions were lower than that from immersion. The results indicated that AOT/Tween 85 reverse micelle system is effective in extracting soya bean protein and oil.     

Effects of high hydrostatic pressure on physicochemical and functional properties of walnut (Juglans regia L.) protein isolate

BACKGROUND: Walnut (Juglans regia L.) is a good source of protein that has potential application in new product formation and fortification. The main objectives of this study were to investigate the effects of high hydrostatic pressure (HHP) treatment (300–600 MPa 20 min) on physicochemical and functional properties of walnut protein isolate (WPI) using various analytical techniques at room temperature. RESULTS: The results showed significant modification of solubility, free sulfhydryl content and surface hydrophobicity with increased levels of HHP treatment, indicating partial denaturation and aggregation of proteins. Differential scanning calorimetry and fluorescence spectrum analyses demonstrated that HHP treatment resulted in gradual unfolding of protein structure. Emulsifying activity index was significantly (P < 0.05) increased after HHP treatment at 400 MPa, but significantly decreased (P < 0.05) relative to the untreated WPI with further increase in pressure. HHP treatment at 300–600 MPa significantly decreased emulsion stability index. Additionally, HHP‐treated walnut proteins showed better foaming properties and in vitro digestibility. CONCLUSION: These results suggest that HHP treatment could be applied to modify the properties of walnut proteins by appropriate of pressure levels, which will help in using walnut protein as a potential food ingredient. © 2012 Society of Chemical Industry     

微波法辅助反胶束技术前萃取大豆蛋白工艺

以全脂大豆为研究对象,为提高反胶束前萃取大豆蛋白的效率,利用变频微波萃取仪,辅助反胶束技术前萃取大豆蛋白。研究单因素即提取温度、pH值、液料比与时间对大豆蛋白提取率的影响,通过主要因素的响应面试验,优化提取参数,且检测了参数之间相互作用关系。利用统计分析和优化,结果显示,微波对前萃取大豆蛋白影响的相对大小依次为提取时间＞温度＞pH值。结果表明：在提取温度43.3 ℃、pH 8、萃取时间23 min时,蛋白质前萃率达到最高水平,提取率80.61%。     

Isolation of obligate anaerobic rumen bacteria capable of degrading the neurotoxin β‐ODAP (β‐N‐oxalyl‐L‐α,β‐diaminopropionic acid) as evaluated by a liquid chromatography/biosensor analysis system

Six pure strains of obligate anaerobes capable of degrading the toxin β‐N‐oxalyl‐L ‐α, β‐diaminopropionic acid (β‐ODAP) contained in grass pea (Lathyrus sativus) have been isolated from cow rumen. The new isolates were identified as Megasphaera elsdenii (five different genotypes) and Clostridium bifermentans using 16S rDNA analysis. The β‐ODAP degrading efficiency of the isolates was evaluated by measuring the amount of β‐ODAP in the growth medium, which contained β‐ODAP as the only carbon source, before and after incubation with the microbes. The method of analysis was liquid chromatography employing bioelectrochemical detection. The biosensor is based on co‐immobilising two enzymes, glutamate oxidase (GlOx) and horseradish peroxidase (HRP), on the end of a spectrographic graphite electrode. β‐ODAP is oxidised by GlOx to form H₂O₂, which in turn is bioelectrocatalytically reduced by HRP through a mediated reaction using a polymeric mediator incorporating Os^{2 + /3+} functionalities rapidly shuttling electrons with the electrode_giving rise to the analytical signal. On the basis of this analysis system, the new isolates are capable of utilising β‐ODAP as sole carbon source to a maximum of 90–95% within 5 days with concomitant increase in cell protein. Copyright © 2005 Society of Chemical Industry