谷氨酸生产菌S_(9114)中依赖于NADPH谷氨酸脱氢酶的纯化及性质

利用DEAE 纤维素离交柱层析、疏水柱层析和凝胶过滤层析等从谷氨酸棒杆菌S9114 中分离纯化出依赖于NADPH的谷氨酸脱氢酶 (GOH NADPH )。经HPLC测定 ,该酶的分子量为 188ku ;经SDS -PAGE电泳测得该酶的亚基分子量为 3 2ku。提示该谷氨酸脱氢酶由 6个亚基组成。该酶对NADPH具有高度专一性 ,在pH 7 5、3 7℃下 ,以α 酮戊二酸、NH4Cl和NADPH为底物时的米氏常数Km 分别为 9 2 2mmol/L、5 2 7mmol/L和 2 3 1× 10 -3mmol/L。最适反应pH为 7 5 ,最适反应温度为 42℃ ,并对热比较稳定。此外 ,KCl对GDH NADPH的活性具有激活作用。     

稀土元素对谷氨酸发酵产酸及其谷氨酸脱氢酶的影响

以目前我国谷氨酸发酵广泛使用的谷氨酸棒杆菌S9114 为实验菌株 ,研究了稀土元素对谷氨酸发酵产酸及其谷氨酸脱氢酶的影响。结果表明LaCl3、CeCl3和NdCl3浓度分别为 0 72 0、0 0 71和 0 0 0 7mmol/L时 ,促使谷氨酸发酵产酸水平提高 6%～ 8% ,对菌体的GDH NADPH的酶活性有显著的激活作用。实验还表明 ,稀土元素对纯化GDH NADPH的活性也有一定的调节作用     

不同乳酸菌产谷氨酸脱羧酶特性研究

通过对发酵乳杆菌、植物乳杆菌、短乳杆菌所产谷氨酸脱羧酶(GAD)的酶学性质进行比较,分析酶反应温度、pH值、磷酸吡哆醛(PLP)浓度、酶浓度与酶活性的关系,并对高活性乳酸菌GAD酶的米氏常数进行测定。结果表明,3株乳酸菌GAD酶的最适温度为40℃,最适pH值为4.5,在PLP添加量为0.1mol/L时,对酶的促进作用达到最高,发酵乳杆菌Km值为0.05215mmol/L,最大反应速度Vmax=2.08μmol/min;短乳杆菌Km值为0.0243mmol/L,最大反应速度Vmax=1.01μmol/min。     

谷氨酸生产菌S9114中谷氨酸脱氢酶分离纯化条件的初步研究

为获得谷氨酸脱氢酶(glutanmte dehydrogenase，GDH)分离纯化的较好效果，文章对French pressure cell press的茵体破碎条件和DEAE-纤素柱分离条件进行了初研究。结果表明：细胞破碎的较好条件为French pressure cell pressl8000pa破碎两次；DEAE-纤维素柱层析GDH的较好条件为柱洗脱线性梯度盐浓度范围为0．2～0．6mol／L，上样量为5～8mL，柱流速为2～3mL／min。     

谷氨酸脱羧酶在枯草芽孢杆菌中的表达及应用

为了研究在枯草芽孢杆菌发酵过程中添加相对廉价的辅酶前体对谷氨酸脱羧酶(GAD)酶活力和催化效率的影响,将来源于Escherichia coli的谷氨酸脱羧酶基因gadB,通过构建重组质粒p HY300PLK-gadB,在枯草芽孢杆菌中重组表达,得到重组菌B. subtilis/pHY300PLKgadB。研究了分别在重组菌发酵过程中添加辅酶磷酸吡哆醛(PLP)、辅酶前体吡哆醛(PL)和吡哆醇(PN)对GAD酶活力的影响。当设置摇床转速200 r/min,发酵温度33℃,发酵培养基初始p H 7.0时,在发酵过程中分别添加PLP、PL、PN至终浓度0.5 mmol/L,诱导48 h后发酵液中总酶活分别达到25.40、28.14、15.55 U/mL,是对照总酶活的1.55、1.72、0.95倍。基于以上结果,进一步研究了发酵过程中PL的添加浓度对重组菌生长情况及GAD表达的影响。结果表明,发酵液中GAD总酶活随着PL浓度的增加而增加,当PL浓度为0.1 mmol/L时,总酶活最高达到28.28 U/mL。利用重组菌全细胞制备γ-氨基丁酸(GABA),结果表明,最适转化pH为5.0,最适转化温度为40℃,发酵过程中添加PL的重组菌(简称"GAD-PL")和发酵过程中不添加任何辅酶及辅酶前体的重组菌(简称"GAD-0")的最适加菌量(以酶活力单位计)分别是40 U/g(以谷氨酸计)和50 U/g(以谷氨酸计),当谷氨酸最终质量浓度达到400 g/L时,得到的GABA质量浓度分别为275.60 g/L和273.61 g/L。     

牛肝谷氨酸脱氢酶的分离纯化及部分酶学性质

取新鲜牛肝,经匀浆、缓冲液抽提、正丁醇脱脂、硫酸铵分级沉淀、DEAE-Sepharose离子交换层析、Superdex-200凝胶过滤层析等方法纯化,获得了电泳纯的牛肝谷氨酸脱氢酶（glutamate dehydrogenase,GDH）。经过纯化后的结果：GDH的酶比活力为306.06 U/mg,纯化倍数为93.60 倍,酶活回收率为23.12%。GDH的分子质量为380.2 kD,亚基分子质量约为61.7 kD。酶学性质研究结果表明：GDH的最适反应温度为50 ℃,在温度为30 ℃以下时较稳定;最适反应pH值为8.2,该酶对烟酰胺腺嘌呤二核苷酸（nicotinamide adenine dinucleotide,NADH）的Km值为0.696 mmol/L。甲醇、乙醇、异丙醇、Cd2+、Co2+、Ca2+、Ag+、Pb2+、Zn2+、Cu2+对该酶具有抑制作用,低浓度Ba2+、Mg2+、K+、Li+与乙二胺四乙酸（ethylenediamine tetraacetic acid,EDTA）对其具有一定的激活作用。     

九株谷氨酸生产菌的生物学特性研究

[目的]研究九株谷氨酸生产菌的生物学特性,为具有广域pH耐受性的谷氨酸高产菌筛选提供实验依据.[方法]考察九株菌的低pH生长、高pH产酸特性;对磺胺胍、酮基丙二酸、丙二酸和香豆素的耐受性以及发酵特性.[结果]Corynebacterium glutamicum S9114、Corynebacterium glutamicum ATCC 13761、Corynebacterium glutamicum T613-85 均能在pH3.9下生长,而在pH10.5时S9114、Corynebacterium acetoacidophilum ATCC 13870、T613-85依然能够产酸;抗性平板上具有明显生长优势的菌株如下:磺胺胍:S9114;酮基丙二酸:S9114、ATCC 13870;丙二酸:ATCC13761、ATCC13870、Microbacterium ammoniaphilum ATCC 15354;香豆素:除ATCC13761和Corynebacterium melassecola ATCC17966,其他耐受程度相似;九株菌中产酸能力最强的是S9114,其次是T613-85.[结论]以S9114为出发菌株进行改造,确定筛选平板为含磺胺胍2%、酮基丙二酸0.25%、丙二酸1.6%、香豆素0.2%的低pH梯度(pH 3.6～4.0)平板和高pH梯度平板(pH 10.5～11.2).     

乳酸菌谷氨酸脱羧酶的酶学性质研究

本文对一株乳酸菌所产谷氨酸脱羧酶的酶学性质进行了较系统的研究,其中包括酶的热稳定性、pH稳定性及温度、pH和一些化学物质对酶活的影响。结果表明:此酶的最适温度为52℃,最适pH为4.5,米氏常数Km=24mmol。PLP、VB6及Ca2 在一定程度上都能促进酶活,且在含量小于100μmol时,作用程度为PLP>VB6>Ca2 。乙酸浓度小于0.05mol/L也能提高酶活力。     

米糠谷氨酸脱羧酶的分离纯化及酶学性质研究

利用(NH_4)_2SO,分级沉淀、DEAE-Sephrose FF离子交换色谱技术分离纯化米糠谷氨酸脱羧酶(GAD).米糠GAD纯化倍数为8.8,比活达到了30.1U/mg,酶活回收率为45.5%.同时对米糠GAD酶学性质进行了研究,结果表明.最适温度为40℃,耐热性较差;最适pH5.5,在pH5.5～9都可以保持较高酶活.米糠GAD对底物和辅酶PLP的K_m分别为28.45、1.13μmol/L.KI、Ag~+、SDS、乙酸对米糠GAD的活力有较大的抑制作用,而Ca~(2+)对米糠GAD的活性有较强的激活作用.     

米糠谷氨酸脱羧酶的酶学性质研究

研究了米糠谷氨酸脱羧酶(GAD)的酶学性质。结果表明:米糠GAD的最适温度为40℃,最适pH5.6,米糠GAD耐热性较差,60℃下保温0.5 h后,酶活下降至65%,保温1 h,酶活下降至48%,而70℃时酶就完全失活,而此酶在pH5.5～9都可以保持84%以上的酶活力。米糠GAD对底物和辅酶PLP的Km分别为0.028 45 mol/L和1.13μmol/L。KI、Ag~(2+)、SDS、乙酸对米糠GAD的活力有较大的抑制作用,而Ca~(2+)对米糠GAD的活性有较强的激活作用,提高了底物与米糠GAD的亲和力。     

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