大麦生产重组蛋白

圣路易斯华盛顿大学的John Rogers若得手的话,大麦即可取代微生物或动物细胞而成为重组蛋白的寄主生物。他企望将编码有潜力的人类治疗蛋白的基因引入大麦植株,使之在种子内产生外源蛋白。大麦种子在发芽时产生大量α-淀粉酶,而 John Rogers 意在以一种外源蛋白取代其中一部分。发芽大麦中进行淀粉酶生产是制成麦芽(将大麦转化为麦芽以制造啤酒、饮料以及其它食品)的关键。     

啤酒废酵母对镉离子的吸附研究

以啤酒酿造厂的啤酒废酵母为生物吸附剂,研究啤酒废酵母对Cd2 的生物吸附行为。利用原子吸收光谱法测定Cd2 含量。结果表明,啤酒废酵母吸附Cd2 受吸附时间、吸附温度、溶液pH值、酵母添加量和Cd2 起始浓度等因素的影响。实验确定了啤酒废酵母对Cd2 的最佳吸附条件。即:pH值6,Cd2 浓度为50mg/L,酵母添加量为1.0g/L,吸附温度25℃,吸附时间30min,此时啤酒废酵母对Cd2 的吸附量可达42.92mg/g干酵母。吸附Cd2 的啤酒废酵母用1.0mol/L的HCl解吸,解吸率达75.46%。对未吸附Cd2 的空白酵母和吸附Cd2 的酵母进行红外光谱分析,结果显示啤酒废酵母吸附Cd2 后羟基和羧基吸收峰发生明显变化,因此认为羟基和羧基在生物吸附中起着重要作用。     

不同预处理方法对大麦花药-花粉培养的影响

研究了甘露醇预处理的适应性以及pH值对甘露醇预处理效果的影响;并首次将山梨醇预处理应用到大麦花药培养中,获得理想的实验结果。第一,采用甘露醇预处理, 17种材料平均愈伤组织诱导率为20.67块/花药,绿苗产量为2.46株/花药。第二,甘露醇预处理溶液的pH值不同,其预处理的效果也不同。其中,愈伤组织诱导率和绿苗产量均以pH5.6最高。第三,不同浓度(0.1-0.5mol/L)的山梨醇预处理3天绿苗产量差异不显著;但同一浓度(0.3mol/L)山梨醇预处理不同天数(1-7天)绿苗产量差异极显著,以3天处理效果最好,绿苗产量是对照的51.2倍。     

不同预处理方法对大麦花药－花粉培养的影响

研究了甘露醇预处理的适应性以及ｐＨ值对甘露醇预处理效果的影响;并首次将山梨醇预处理应用到大麦花药培养中,获得理想的实验结果。第一,采用甘露醇预处理,１７种材料平均愈伤组织诱导率为２０．６７块／花药,绿苗产量为２．４６株／花药。第二,甘露醇预处理溶液的ｐＨ值不同,其预处理的效果也不同。其中,愈伤组织诱导率和绿苗产量均以ｐＨ５．６最高。第三,不同浓度（０．１—０．５ｍｏｌ／Ｌ）的山梨醇预处理３天绿苗产量差异不显著;但同一浓度（０．３ｍｏｌ／Ｌ）山梨醇预处理不同天数（１—７天）绿苗产量差异极显著,以３天处理效果最好,绿苗产量是对照的５１．２倍。     

BP神经网络优化微生物浸矿工艺

以低品位黄铜矿溶液为原料,浸矿制备Cu2＋能有效提高低品位黄铜矿的利用价值。基于浸矿过程中存在多因素影响的现象,通过正交试验与神经网络分析方法,对浸矿条件（接种量、矿石品位、Fe2＋添加量及浸矿溶液pH）实行优化。结果表明：在正交试验组中最佳试验结果为浸矿产128．753mg／LCu^2＋;BP神经网络优化后的最佳实验组合为微生物接种量12％、矿石品位0．3％、添加Fe^2＋24g/L及浸矿溶液pH1．7,该条件下验证试验产Cu^2＋ 141．352mg／L,通过正交试验及神经网络优化提高了微生物浸出低品位黄铜矿酸性溶液Cu^2＋的产量。     

荧光素酶基因luc在大肠杆菌中表达条件优化

对重组荧光素酶大肠杆菌菌株M15／pQE30-luc进行了表达条件的优化研究。单因素结果表明：在初始pH值7．0,装液量为20％,2％的接种量,终浓度为0．5mmol／L的IPTG,添加10—30mmol／L的Mg^2＋,摇床转速为200r／min,37℃诱导3．5h酶的表达量最高。正交试验结果表明：初始pH值为7．0,添加40mmol／LMg^2=,接种量2％,装液量为20％时表达量最高,比酶活达1．63×10^8RFU／mg蛋白。     

红豆杉细胞悬浮培养中不同添加物对细胞生长和紫杉醇合成的影响

采用正交实验检测红豆杉（Ｔａｘｕｓ ｃｈｉｎｅｎｓｉｓ（Ｐｉｌｇｅｒ）Ｒｅｈｄ．）细胞悬浮培养中水杨酸、Ｄ－果糖、甘露醇和硫酸镧对细胞生长和紫杉醇（ｔａｘｏｌ）积累的影响。添加１０ｇ／ＬＤ－果糖,可使细胞的鲜重和干重明显增加;添加６０ｇ／Ｌ甘露醇使细胞的鲜重和干重明显减少;１ｍｇ／Ｌ水杨酸仅使细胞鲜重增加,对干重影响不明显;硫酸镧对细胞生长无明显影响。单独添加这４种物质,紫杉醇含量均下降,同时添加     

金溶胶基底的 SERS 增强效果的实验研究

本文以结晶紫作为探针分子,研究了以金溶胶膜、pH ＝6以及 pH ＝13的金溶胶溶液为活性基底的表面增强拉曼光谱的增强效果。采用化学还原法制备金溶胶,加入氢氧化钠改变其 pH 值,并以自组装法制备金溶胶膜。通过比较金溶胶膜、pH ＝6及 pH ＝13时金溶胶溶液的增强因子以及在这三种金溶胶基底上结晶紫的检测限,分析不同活性基底增强效果的差异。三种活性基底的增强因子分别可达到5．9×103、1．5×105、2．3×107,pH ＝13的金溶胶溶液有最佳的增强效果。以这三种金溶胶为基底对结晶紫进行表面增强拉曼光谱探测,可得到检测限为70．7 nmol／L、9．6 nmol／L、1．8 nmol／L。结果表明,金溶胶溶液的增强效果明显优于金溶胶膜,而通过改变金溶胶体系的 pH 值可以改变金纳米颗粒的聚合程度及对探测物的吸附特性从而获得更高灵敏度的活性基底。     

基于诊断标记的LOX-1活性缺失大麦种质鉴定评价

在啤酒酿造和储藏过程中,所含脂类物质代谢是影响啤酒风味稳定性和啤酒货架期长短的主要原因。研究表明,大麦脂肪氧化酶1(LOX-1)是导致籽粒中脂类代谢的关键酶,筛选和创制LOX-1活性缺失(null LOX-1)的大麦种质是促进啤酒大麦品质育种的有效途径。为提高检测效率,针对前期鉴定出的中国null LOX-1大麦稀有SNP功能变异,开发出特异性KASP快速诊断标记,并利用该标记对来源于河南和山东两省的633份大麦地方品种进行鉴定评价,共计筛选出8份LOX-1活性缺失新种质,同时明确了该变异的地理分布及其在不同地方品种中的变异频率。本研究不仅为啤酒大麦分子辅助育种提供了优异种质和快速检测手段,也为大麦种质资源遗传完整性保护与利用提供了参考。     

低双乙酰啤酒酵母菌株BEZ112的选育

以啤酒酿造生产菌株啤酒酵母(Saccharomyces cerevisiae)FB作为出发菌株,用甲基磺酸乙酯(EMS)诱变,经分离筛选得到一株优良的啤酒酵母菌株BEZ112。该菌株的絮凝性、发酵度、酒精度、发酵液的总酯和总高级醇的含量等特性保持了亲株的优良性状。但以12°Bx麦芽汁为培养基用500mL三角瓶在12℃下发酵,该菌株发酵至第4d,发酵液中的双乙酰含量达到峰值(0.291mg/L),比出发菌株FB发酵4d的峰值降低了30％,发酵至第8d,BEZ112发酵液中的双乙酰含量比出发菌株FB的降低了23％。以12°Bx麦芽汁为培养基用500L罐在12℃下发酵8d,BEZ112发酵液中的双乙酰含量(0.091mg/L)比出发菌株FB的(0.124mg/L)降低了27％。发酵得到的啤酒口感纯正清爽。     

Traces of a possible Celtic brewery in Eberdingen-Hochdorf, Kreis Ludwigsburg, southwest Germany

A large number of weakly germinated hulled barley grains was found during archaeobotanical analyses from the early Celtic settlement excavations at Eberdingen-Hochdorf in southwest Germany (ca. 600 – 400 BC). These grains seem to represent deliberate germination, due to the purity of the find and its unusual archaeological context. The possibility of deliberate malting which could be connected with beer brewing is discussed. Recent germination and charring experiments show that the consistently weak traces of germination on the charred subfossil grains from Hochdorf are enough to indicate malted grains. A comparison of the archaeobotanical remains with the written and archaeological sources shows that evidence of beer brewing from excavations is very scarce. There is practically no clear proof of brewing, while written sources and indirect suggestions are abundant. Neither archaeological finds nor either written or iconographic sources give exact details about the prehistoric brewing technology of the early Celts. The archaeological finds from Hochdorf seem to be the result of deliberate malting of hulled barley for the purpose of Celtic beer brewing.     

Probing heat-stable water-soluble proteins from barley to malt and beer

Proteins determine the quality of barley in malting and brewing end-uses. In this regard, water-soluble barley proteins play a major role in the formation, stability, and texture of head foams. Our objective was to survey the barley seed proteins that could be involved in the foaming properties of beer. Therefore, two-dimensional (2-D) electrophoresis and mass spectrometry were combined to highlight the barley proteins that could resist the heating treatments occurring during malting and brewing processes. As expected, from barley to malt and to beer, most of the heat-stable proteins are disulfide-rich proteins, implicated in the defense of plants against their bio-aggressors, e.g., serpin-like chymotrypsin inhibitors (protein Z), amylase and amylase-protease inhibitors, and lipid transfer proteins (LTP1 and LTP2). For LTP1s, the complex pattern displayed in 2-D electrophoresis could be related to some chemical modifications already described elsewhere, such as acylation or glycation through Maillard reactions, which occur on malting. Our proteomics approach allowed the identification of the numerous proteins present in beer in addition to the major ones already described. The involvement of these proteins in the quality of beer foam can now be evaluated.     

Yeasts isolated from industrial maltings can suppress <Emphasis Type="Italic">Fusarium</Emphasis> growth and formation of gushing factors

Fusarium infection of barley and malt can cause severe problems in the malting and brewing industry. In addition to being potential mycotoxin producers, Fusarium fungi are known to cause beer gushing (spontaneous overfoaming of beer). Cereal-derived bacteria and yeasts are potential biocontrol agents. In this study, the antifungal potential of selected yeasts (12 strains) derived from the industrial malting ecosystem was studied in vitro with a plate-screening assay. Several ascomycetous yeast strains showed antagonistic activity against field and storage moulds, Pichia anomala being the most effective strain. The effects of P. anomala VTT C-04565 (C565) were examined in laboratory scale malting with naturally contaminated barley exhibiting gushing potential. P. anomala C565 restricted Fusarium growth and hydrophobin production during malting and prevented beer gushing. Grain germination was not disturbed by the presence of yeast. Addition of P. anomala C565 into the steeping seemed to retard wort filtration, but the filtration performance was recovered when yeast culture was combined with Lactobacillus plantarum VTT E-78076. Well-characterized microbial cultures could be used as food-grade biocontrol agents and they offer a natural tool for tailoring of malt properties.     

Enhancement by Low pH of Gibberellin Effects on Dormant Celery Seeds and Embryoless Half-Seeds of Barley

The action of gibberellins on celery (Apium graveolens L.) seed germination and the release of reducing sugars from barley (Hordeum vulgare L.) seed endosperm were enhanced by decreasing the pH of the incubation solution to below the pKa point. In most cases, low pH was obtained by mixing the solution with weak acids such as succinic acid 2.2 dimethylhydrazide (SADH), 2-chloroethylphosphonic acid (CEPA), or citric acid. However, lowering the pH of the gibberellin solution with strong acid (HCl) also increased markedly the activity of low concentrations of GA_4/7. The synergistic action of ethylenediaminetetraacetic acid (EDTA) with gibberellin was not dependent on the pH level of the incubation solution. The response of celery seeds to gibberellins was increased when their distal ends were removed: solution pH and EDTA had no effect on this response. The possible explanations of the synergism between low pH compounds and gibberellins are discussed.     

Enhancing the production of phenolic compounds during barley germination by using chitooligosaccharides to improve the antioxidant capacity of malt

Objective

To enhance the production of phenolic compounds during barley germination using chitooligosaccharide as an elicitor to improve the antioxidant capacity of malt.

Results

When used as an elicitor for barley germination, chitooligosaccharide with a molecular weight of 3 kDa, added at 10 mg/kg barley kernels during the first steeping cycle, led to the maximum production of phenolic compounds. Compared with the control with no chitooligosaccharide added to the steeping water, the total phenolic content was increased by 54.8%. Increases in the total phenolic content of the barley malt occurred when chitooligosaccharide was applied during the first or both the first and the second steeping cycles. Thus the antioxidant capacity of barley malt was increased significantly by adding chitooligosaccharide during the steeping process.

Conclusion

Applying chitooligosaccharides during the steeping process increased the content of phenolic compounds thus improving the antioxidant capacity of the barley malt.

     

2D-HPLC and MALDI-TOF/TOF analysis of barley proteins glycated during brewing

The barley proteins have been the subject of interests of many research groups dealing with barley grains, malt and beer. The proteins which remain intact after harsh malting conditions influence the quality and flavor of beer. The characteristic feature of the proteins present in malt and beer is their extensive modification with carbohydrates, mainly glucose that comes from the starch degradation during technological processes. The degree of the protein glycation has an effect on the quality of malt and beer and on the properties of the beer foam. A combination of two-dimensional high performance liquid chromatography (2D-HPLC) and matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/TOF MS) was used for the analysis of the protein extracts that were reduced, alkylated, and degraded enzymatically without prior protein separation. This so-called "shot-gun" approach enabled us to determine glycation sites in one third of the proteins identified in the study and to propose potential glycation markers for fast and efficient monitoring during malting.     

环境变异对青稞热稳定蛋白质含量及蛋白质Z的影响

热稳定蛋白是衡量麦芽品质的重要指标,为探明青稞籽粒和麦芽热稳定蛋白的含量、蛋白质Z的组成特征以及影响条件。本研究以3份青稞和1份对照大麦品种Gairdner为试验材料,对青稞籽粒及其麦芽的热稳定蛋白进行分析与鉴定,研究了不同生态环境下青稞热稳定蛋白质含量和蛋白质Z的组成特征,同时筛选出了优异啤用品质青稞品种(品系)。结果表明,青稞发芽温度为20℃,发芽时间为72 h,培养溶液PH为5时,发芽及焙焦条件下最有利于青稞热稳定蛋白总含量及蛋白质Z的累积。利用该发芽条件筛选种植于西宁、湟源和海晏的青稞资源,发现种植于西宁的青稞种子和发芽后热稳定蛋白质总含量最低,但是焙焦后热稳定蛋白质和蛋白质Z含量最高;同时从150份青稞资源中筛选出热稳定蛋白质含量及蛋白质Z条带清晰、含量高的优异资源15份。本研究结果为酿造青稞品种选育、啤用青稞和麦芽质量评价指标提供理论依据。     

Shotgun proteome analysis of beer and the immunogenic potential of beer polypeptides

The majority of beer proteins originate from barley (Hordeum vulgare) which is used for brewing. Barley is known to contain celiacogenic gliadin-like prolamins (hordeins) along with other immunogenic proteins which endure malt proteases and the harsh conditions of brewing. In addition, a multitude of peptides that may retain or even amplify the immune-stimulating potential is released in beer because of proteolysis. The comprehensive annotation of the beer proteome is challenged both by the high concentration range of the protein entities and by a severe degree of processing-induced modifications. Overcoming the pitfalls of the classical two-dimensional electrophoresis approach coupled to mass spectrometry (MS), the gel-free shotgun proteomic analysis expanded the current inventory of a popular Italian beer to 33 gene products, including traces of intact B- and D-hordeins and 10 proteins from Saccharomyces spp. The high performance liquid chromatography-electrospray MS/MS peptidomic analysis of the low-molecular weight beer components disclosed a panel of hordein-derived peptides that encrypt gluten-like sequence motifs, potentially harmful to celiacs. The presence of antigliadin IgA-immunoresponsive prolamins was assayed by Western and dot blot using sera of N=4 celiac patients. Gliadin-reactive T-cell lines isolated from the intestine of N=5 celiacs activated an IFN-γ response when challenged with deamidated beer polypeptides.     

Dynamics of yeast cell states during proliferation and non proliferation periods in a brewing reactor monitored by multidimensional flow cytometry

Current demands of the brewing industry require that increasing amounts of beer be produced in ever-decreasing times, without prejudicing the quality of the product. The four basic ingredients for brewing beer are malt, hops, water and yeast. Of these four materials, yeast is unique, in that its competent handling can reduce the time taken for the brewing process. For this purpose, it is necessary to have information regarding the metabolic acting as well as the physiological state of an individual cell for which flow cytometry is used. Knowledge of changes in DNA, neutral lipid and 3β-hydroxysterol content of the yeast cells during growth, fermentation and storage enables for a time-saving process control. All of these parameters were conveniently monitored by flow cytometry in conjunction with double fluorescent staining techniques, as shown in this study.     

The technology and properties of beer produced from unmalted sorghum or maize grains

Three basic samples of beers were produced: A, B and C. The grit of A and B contained as unmalted adjuncts 15% (plus 10% of saccharose) and 25% of sorghum grains or maize grits, respectively. The reference beer C was produced with barley malt only. The study of the effects of the unmalted adjuncts on the brewing and the quality of beer revealed that: (a) the use of 15 to 20% of maize or 25% of sorghum increased the content of iso-compounds in wort; (b) the combination of maize grit and saccharose improved the colour of the wort and beer; (c) the addition of 25% sorghum extended saccharification time, slowed down both wort and beer filtration and also produced wort of a darker colour and beer with a slightly bitter aftertaste.