地衣芽孢杆菌弹性蛋白酶纯化和性质研究

用地衣芽孢杆菌发酵液制备弹性蛋白酶粗酶液,采用硫酸铵分级沉淀和Sephadex凝胶柱层析的方法分离纯化弹性蛋白酶,并对弹性蛋白酶的酶学性质进行研究。结果表明：弹性蛋白酶粗酶液经40%～70%饱和度的硫酸铵纯化后比活力提高到120U/mg,经凝胶柱层析纯化后比活力可达到292U/mg,纯化倍数为12.6,SDS-PAGE法证实弹性蛋白酶分子质量为29.5kD。对酶学性质的研究结果表明：弹性蛋白酶最适反应温度为55℃,最适反应pH值为7.4,以刚果红-弹性蛋白为底物,米氏常数Km为9.67mg/mL。低浓度金属离子Ca2+和K+对酶活力有促进作用,而Mg2+、Mn2+、Zn2+和Al3+对酶活力则有抑制作用。     

基于弹性蛋白-RBB底物的弹性蛋白酶活力检测方法研究

以Remazol brilliant blue R染色牛颈韧带弹性蛋白粉,制成染色弹性蛋白。底物稳定性好,在较宽pH范围内染料不易洗脱。以弹性蛋白-RBB为底物,建立了一种能在较宽pH范围内测定制革蛋白酶的弹性蛋白水解活力方法。确定酶活测定条件:反应时间为30min,温度为40℃,基于5mg底物量,待测酶液中弹性蛋白酶活在90～160U/mL范围内,水解液的吸光度与酶的浓度呈良好的线性关系。方法重线性较好。几种常用制革蛋白酶在不同pH下测定的结果表明,一些细菌蛋白酶显示出较高的弹性蛋白酶活力,最适作用pH值在9左右。     

产低温弹性蛋白酶菌株的筛选及酶学特性分析

筛选获得一株产生低温型弹性蛋白酶菌株并研究酶学性质,为该酶在肉类嫩化中应用奠定理论基础。在不溶性的弹性蛋白琼脂平板上,以弹性蛋白作为惟一氮源,根据水解圈直径和菌落直径的比值筛选到一株分泌低温型弹性蛋白酶的菌株,命名为XZE116,通过形态学、生理生化试验以及16s rDNA序列分析,该菌株鉴定为短芽孢杆菌(Bacillus brevis)。菌株所产弹性蛋白酶最适作用pH值8.0,在较广的pH值范围(7.0～12.0)内酶稳定性较好。酶最适作用温度25℃,在40℃以下保温2 h,酶稳定性好。浓度为5 mmol/L的Mg2+离子对酶有激活作用,但Hg2+、Pb2+强烈抑制酶活性。丝氨酸蛋白酶特异性抑制剂苯甲基磺酰氟(PMSF)强烈抑制酶活性,但E-64对酶活性没有影响,表明菌株XZE116弹性蛋白酶是丝氨酸蛋白酶。0.1%的阴离子表面活性剂SDS、阳离子表面活性剂十六烷基三甲基溴化铵(CTAB)以及非离子型表面活性剂Triton X-100对酶活性有轻微的抑制作用。这些酶的优良生化特性赋予了该酶在肉品工业中具有潜在的应用价值。     

微生物弹性蛋白酶催化动力学特性研究

研究了弹性蛋白酶的酶学性质与酶促反应动力学.研究表明,弹性蛋白酶的作用最适pH值与所用缓冲体系有关,在硼砂-硼酸缓冲体系中,最适pH值为7.8,而在Tris缓冲体系中,最适pH值为8.48;酶的作用最适温度为50℃,当温度高于60℃时,酶活性下降辐度较大,差异极显著.酶在pH3～7即酸性条件下,酶活较低,而在pH7～12时酶活较高,且趋于稳定.大部分金属离子对酶的活性有抑制作用,尤其是Ba+、Ca+、Al+和Cu+等金属离子可使酶的活性直线下降,而钠离子对酶的活性没有明显影响.采用Lineweaver-Burk方法,获得酶催化动力学参数Vmax=0.09512U/mL,km=0.00741m/ml,应用Michaelis-Menten方程求得弹性蛋白酶以弹性蛋白为底物的催化动力学方程为V=0.09512[S]/0.00741+[S].     

弹性蛋白酶高产菌株摇瓶发酵条件优化及弹性蛋白酶性质的初涉研究

运用响应面法对一株枯草芽孢杆菌(Bacillus subtilis)发酵培养条件进行了优化,这株枯草芽孢杆菌产弹性蛋白酶的最适发酵条件为:1.5%葡萄糖、1.5%酵母膏,起始pH为6.5,最适发酵温度为36.5℃,并对由这株枯草芽孢杆菌发酵生产的弹性蛋白酶进行了研究,发现由该株菌发酵得到的弹性蛋白酶在碱性条件下活性较好;在37℃以下时稳定性较好.     

米曲霉M3中性蛋白酶的提取及酶学特性研究

采用硫酸铵沉降法分离纯化米曲霉M3所产中性蛋白酶，并对其酶学性质进行了研究。结果表明：用硫酸铵沉降法所得粗酶粉的酶活力高达159781．1μ／g；该酶的最适反应温度为50℃，最适作用pH值为7．5；该酶在30～40℃时具有良好的热稳定性，在pH值6．5—7．5的条件下酶是稳定的；NH4^1 ，K^1 对该蛋白酶有明显的激活作用，Fe^3 则对其表现出强烈的抑制作用。     

胞外弹性蛋白酶的理化特性及其影响因素

分离得到的假单胞菌(Pseudomonas sp) 菌株具有较强的分泌胞外弹性蛋白酶的能力。经微生物发酵方法生产的弹性蛋白酶,经过盐析、透析、DEAE SephadexA2 5离子交换层析、SephadexG75凝胶过滤层析等纯化步骤,从其发酵液中得到了均一的酶制品。研究结果表明,酶的最适作用温度为5 0℃;在硼砂-硼酸缓冲液中最适作用pH为8 0左右;酶在碱性环境下( pH7 0～12 0 )稳定性较好;在37℃以下,酶的稳定性较高。超过6 0℃,酶在短时间内即失活     

鱿鱼肝脏自水解过程中内源蛋白酶的作用研究

对鱿鱼肝脏蛋白酶的酶学性质及其在鱿鱼肝脏自水解过程中的作用进行研究。结果表明：以偶氮酪蛋白为底物,TCA可溶性肽为评价指标,测得鱿鱼肝脏蛋白酶粗酶的最适反应温度为40℃,最适反应pH值为5.0;该粗酶在pH3.0～8.0较为稳定。在鱿鱼肝脏自水解过程中,内源性半胱氨酸蛋白酶发挥主要作用。此外,丝氨酸蛋白酶和金属蛋白酶也有一定作用。     

草鱼消化道粗蛋白酶的部分性质

用丙酮脱脂然后用磷酸盐缓冲溶液从草鱼肠道中提取一种主要含有酸性蛋白酶和碱性蛋白酶的粗蛋白酶。该粗酶水解血红蛋白的最适pH=3～4，水解酪蛋白的最适pH值有2个，分别为pH4．4和PH10．3。水解牛血红蛋白的最适作用温度为37℃，水解酪蛋白的最适作用温度为45℃。酸性蛋白酶在50℃保温30min后只有大约20％的最初活性被保留，而要使碱性蛋白酶丧失80％的最初活力需要在60℃保温30min。     

微生物产弹性蛋白酶分子研究进展

本文从分子水平对微生物产弹性蛋白酶的研究概况进行了综述。内容包括不同菌源性弹性蛋白酶基因克隆、基因序列及功能分析、基因表达的调控、利用蛋白质工程对菌源性弹性蛋白酶进行改造,目的是从分子水平时弹性蛋白酶的转录表达、酶学反应机制进行阐释,最终提高酶的表达量,拓宽酶的应用范围。     

Reliable identification of grapevine rootstock varieties using RAPD PCR on woody samples

Since grapevine ( Vitis spp .) rootstock material is being traded increasingly as disbudded woody material a lack of distinctive morphological features on such material necessitates an alternative and reliable means of identification. Methods described here were developed for rapid and efficient extraction of DNA from woody samples rich in phenolic compounds and polysaccharides, and for subsequent identification of varieties by RAPD PCR. Using these methods, and with the application of only one selected RAPD primer, we were able to differentiate sixteen rootstock varieties, including the seven varieties most commonly used in Germany. Problems commonly encountered with reproducibility of RAPD patterns were avoided by choosing primers with a dinucleotide sequence and a high G/C content that allowed a rather high annealing temperature of 45°C. Methods described here should also be useful for other horticultural crops, especially those with woody tissues rich in phenolic compounds and polysaccharides.     

An internet compendium of analytical methods and spectroscopic information for monomers and additives used in food packaging plastics

An internet website (http://cpf.jrc.it/smt/) has been produced as a means of dissemination of methods of analysis and supporting spectroscopic information on monomers and additives used for food contact materials (principally packaging). The site which is aimed primarily at assisting food control laboratories in the European Union contains analytical information on monomers, starting substances and additives used in the manufacture of plastics materials. A searchable index is provided giving PM and CAS numbers for each of 255 substances. For each substance a data sheet gives regulatory information, chemical structures, physico-chemical information and background information on the use of the substance in particular plastics, and the food packaging applications. For monomers and starting substances (155 compounds) the infra-red and mass spectra are provided, and for additives (100 compounds); additionally proton NMR are available for about 50% of the entries. Where analytical methods have been developed for determining these substances as residual amounts in plastics or as trace amounts in food simulants these methods are also on the website. All information is provided in portable document file (PDF) format which means that high quality copies can be readily printed, using freely available Adobe Acrobat Reader software. The website will in future be maintained and up-dated by the European Commission's Joint Research Centre (JRC) as new substances are authorized for use by the European Commission (DG-ENTR formerly DGIII). Where analytical laboratories (food control or other) require reference substances these can be obtained free-ofcharge from a reference collection housed at the JRC and maintained in conjunction with this website compendium.     

Aroma characterization of various apricot varieties using headspace–solid phase microextraction combined with gas chromatography–mass spectrometry and gas chromatography–olfactometry

The characterization of the aromatic profile of several apricot cultivars with molecular tracers in order to obtain objective data concerning the aromatic quality of this fruit was undertaken using headspace–solid phase microextraction (HS–SPME). Six apricot cultivars were selected according to their organoleptic characteristics: Iranien, Orangered, Goldrich, Hargrand, Rouge du Roussillon and A4025. The aromatic intensity of these varieties measured by HS–SPME–Olfactometry were defined and classified according to the presence and the intensity of grassy, fruity and apricot like notes. In the six varieties, 23 common volatile compounds were identified by HS–SPME–GC–MS. Finally, 10 compounds, ethyl acetate, hexyl acetate, limonene, β-cyclocitral, γ-decalactone, 6-methyl-5-hepten-2-one, linalool, β-ionone, menthone and (E)-hexen-2-al were recognized by HS–SPME–GC–O as responsible of the aromatic notes involved in apricot aroma and considered as molecular tracers of apricot aromatic quality which could be utilized to discriminate apricot varieties.     

Tests of potential functional barriers for laminated multilayer food packages. Part I: Low molecular weight permeants

The advent of the functional barrier concept in food packaging has brought with it a requirement for fast tests of permeation through potential barrier materials. In such tests it would be convenient for both foodstuffs and materials below the functional barrier (sub-barrier materials) to be represented by standard simulants. By means of inverse gas chromatography, liquid paraffin spiked with appropriate permeants was considered as a potential simulant of sub-barrier materials based on polypropylene (PP) or similar polyolefins. Experiments were performed to characterize the kinetics of the permeation of low molecular weight model permeants (octene, toluene and isopropanol) from liquid paraffin, through a surrogate potential functional barrier (25 μm-thick oriented PP) into the food simulants olive oil and 3% (w/v) acetic acid. These permeation results were interpreted in terms of three permeation kinetic models regarding the solubility of a particular model permeant in the post-barrier medium (i.e. the food simulant). The results obtained justify the development and evaluation of liquid sub-barrier simulants that would allow flexible yet rigorous testing of new laminated multilayer packaging materials.     

Chlorohydrins of bisphenol A diglycidyl ether (BADGE) and of bisphenol F diglycidyl ether (BFDGE) in canned foods and ready-to-drink coffees from the Japanese market

BADGE.2HCl and BFDGE.2HCl were determined in 28 samples of ready-to-drink canned coffee and 18 samples of canned vegetables (10 corn, 5 tomatoes and 3 others), all from the Japanese market. HPLC was used as the principal analytical method and GCMS for confirmation of relevant LC fractions. BADGE.2HCl was found to be present in one canned coffee and five samples of corn, BFDGE.2HCl in four samples of canned tomatoes and in one canned corn. No sample was found which exceeded the 1mg/kg limit of the EU for the BADGE chlorohydrins. However the highest concentration was found for the sum of BFDGE.2HCl and BFDGE.HCl.H₂O at a level of 1.5mg/kg. A Beilstein test confirmed that all cans containing foods contaminated with BADGE.2HCl or BFDGE.2HCl had at lest one part coated with a PVC organosol.     

European priorities for research to support legislation in the area of food contact materials and articles

A strong science base is required to underpin the planning and decision-making process involved in determining future European community legislation on materials and articles in contact with food. Significant progress has been made in the past 5 years in European funded work in this area, with many developments contributing to a much better understanding of the migration process, and better and simpler approaches to food control. In this paper this progress is reviewed against previously identified work-areas (identified in 1994) and conclusions are reached about future requirements for R&D to support legislation on food contact materials and articles over the next 5 or so years.     

Analysis of benzo[a]pyrene in spiked fatty foods by second derivative synchronous spectrofluorimetry after microwave-assisted treatment of samples

A simple, rapid and inexpensive method has been developed for the determination of benzo[a     

The development of reference materials for paralytic shellfish poisoning toxins in lyophilized mussel. II: Certification study

This paper describes the second part of a project undertaken to develop certified mussel reference materials for paralytic shellfish poisoning toxins. In the first part two interlaboratory studies were undertaken to investigate the performance of the analytical methodology for several PSP toxins, in particular saxitoxin and decarbamoyl-saxitoxin in lyophilized mussels, and to set criteria for the acceptance of results to be applied during the certification exercise. Fifteen laboratories participated in this certification study and were asked to measure saxitoxin and decarbamoyl-saxitoxin in rehydrated lyophilized mussel material and in a saxitoxin-enriched mussel material. The participants were allowed to use a method of their choice but with an extraction procedure to be strictly followed. The study included extra experiments to verify the detection limits for both saxitoxin and decarbamoyl-saxitoxin. Most participants (13 of 15) were able to meet all the criteria set for the certification study. Results for saxitoxin.2HCl yielded a certified mass fraction of <0.07 mg/kg in the rehydrated lyophilized mussels. Results obtained for decarbamoyl-saxitoxin.2HCl yielded a certified mass fraction of 1.59+/-0.20 mg/kg. The results for saxitoxin.2HCl in enriched blank mussel yielded a certified mass fraction of 0.48 +/- 0.06 mg/kg. These certified reference materials for paralytic shellfish poisoning toxins in lyophilized mussel material are the first available for laboratories to test their method for accuracy and performance.     

Announcing a new international peer-reviewed journal:Food Science and Human Wellness now accepting manuscript submission in English

<正>We are pleased to announce the launch of a new international peer-reviewed journal-Food Science and Human Wellness,ISSN 2213-4530,which is an open access journal,produced and hosted by Elsevier B.V.on behalf of Beijing Academy of Food Sciences.Food Science and Human Wellness is an international peer-reviewed English journal that provides a forum for the dissemination of the