海洋假单胞杆菌褐藻胶裂解酶基因在大肠杆菌中的高效表达和活性检测

利用PCR方法从假单胞杆菌基因组DNA扩增胞外褐藻胶裂解酶的全基因和缺损片段,构建表达质粒pQE30-aly32和pQE30-aly165,并在大肠杆菌M15中成功表达。表达的重组蛋白大部分存在于包涵体中,经洗涤,变性和复性,镍柱亲和层析纯化,获得了电泳纯的Aly32和Aly165。酶活性检测发现,仅Aly165具有较强活性,108 u/mg,该酶对poly(G)和poly(M)都有活性,对poly(M)更敏感。对表达体系进行优化,确定IPTG的最佳浓度为0.6mmol/L,最佳诱导菌液密度OD600为0.5~0.6,最佳表达时间为3h,28℃低温诱导可以提高重组蛋白的可溶性,可溶性蛋白相对表达量达到25%。     

Plant Phenolics Extraction from Flos Chrysanthemi: Response Surface Methodology Based Optimization and the Correlation Between Extracts and Free Radical Scavenging Activity

Huaiju is one of the most famous and widely used Flos Chrysanthemi (FC) for medicinal purposes in China. Although various investigations aimed at phenolics extraction from other FC have been reported, a thorough optimization of the phenolics extraction conditions from Huaiju has not been achieved. This work applied the widely used response surface methodology (RSM) to investigate the effects of 3 independent variables including ethanol concentration (%), extraction time (min), and solvent‐to‐material ratio (mL/g) on the ultrasound‐assisted extraction (UAE) of phenolics from FC. The data suggested the optimal UAE condition was an ethanol concentration of 75.3% and extraction time of 43.5 min, whereas the ratio of solvent to material has no significant effect. When the free radical scavenging ability was used as an indicator for a successful extraction, a similar optimal extraction was achieved with an ethanol concentration of 72.8%, extraction time of 44.3 min, and the ratio of solvent to material was 29.5 mL/g. Furthermore, a moderate correlation between the antioxidant activity of TP extract and the content of extracted phenolic compounds was observed. Moreover, a well consistent of the experimental values under optimal conditions with those predicted values suggests RSM successfully optimized the UAE conditions for phenolics extraction from FC.

Practical Application

The work of the research investigated the plant phenolics in Flos Chrysanthemi and antioxidant capacities. These results of this study can support the development of antioxidant additive and relative food.     

鲤鱼肉蛋白水解物在熟猪肉糜中的抗氧化效果

研究鲤鱼肉蛋白的碱性蛋白酶(alcalase)水解物对肉糜的抗氧化作用。将肉糜分为6组,包括阴性对照组、2.0%的未水解鲤鱼肉蛋白组、1.0%、1.5%、2.0%的水解物(4h)添加组、0.02%的丁基羟基茴香醚(butylatedhydroxyanisole,BHA)的阳性对照,在冷藏7d过程中测定肉糜的红度值(a*)、硫代巴比妥酸值(TBARS)值和pH值的变化,并对产品的感官指标进行评定。结果表明:与对照组相比,添加水解物的处理组能显著抑制脂肪的氧化(P<0.05),其中2.0%水解物处理组效果最明显,能显著降低TBARS值、增加肉糜的红度值(a*)(P<0.05),但较添加BHA处理组的效果欠佳;感官评定得到同样的结果。     

Effect of processing steps on the physico-chemical properties of dried-seasoned squid

The effects of processing steps on the physico-chemical properties of dried-seasoned squid were studied using two kinds of squids. Browning was observed as indicated by an increase in yellowness (b^∗ value) during the semi-drying and roasting step. The temperature and time of heating and type of roasting, as well as the material itself were the key factors influencing the browning. A high level of trimethylamine oxide (TMAO), and low contents of trimethylamine (TMA) and dimethylamine (DMA) were detected in the raw material. TMAO gradually decreased during the processing, while TMA, DMA and formaldehyde (FA) increased significantly especially in boiled meat. The temperature of boiling was the most crucial factor impacting the production of FA. FA could be greatly decreased during the cooling process by washing with water or changing the roasting techniques. Therefore, to improve the quality of dried-seasoned squid, more attention should be paid to the temperature and roasting techniques during the processing.     

耐热β-木糖苷酶的构建及在木糖制备中的应用

β-木糖苷酶在完全快速降解木聚糖类半纤维素为木糖的过程中起重要作用.海栖热袍菌(Thermotoga maritima)是一个极端嗜高温厌氧细菌,所产耐热酶类具有非常可观的工业应用前景,但这类酶在大肠杆菌中的表达很低.采用基因重组技术将源自海栖热袍菌的具有阿拉伯糖苷酶活性的耐热β-木糖苷酶基因克隆至表达载体pET-28a,与组氨酸标签融合表达构建重组质粒pET-28a-xyl,然后转化不同大肠杆菌宿主,结果在大肠杆菌BL21-CodonPlus(DE3)-RIL中获得高效表达;重组酶蛋白经诱导表达、破胞和热处理后纯度在90%以上,经Ni2 亲和层析后达电泳纯;用HPLC和TLC检测酶解产物,β-木糖苷酶和木聚糖酶联合水解后,酶解产物主要为木糖,外加阿拉伯糖苷酶后能使其酶解产物中木糖含量明显提高.因此,具有阿拉伯糖苷酶活性的耐热β-木糖苷酶在木糖制备中具有重要的工业应用价值.     

Facile on-site detection of substituted aromatic pollutants in water using thin layer chromatography combined with surface-enhanced Raman spectroscopy

A novel facile method for on-site detection of substituted aromatic pollutants in water using thin layer chromatography (TLC) combined with surface-enhanced Raman spectroscopy (SERS) was explored. Various substituted aromatics in polluted water were separated by a convenient TLC protocol and then detected using a portable Raman spectrometer with the prepared silver colloids serving as SERS-active substrates. The effects of operating conditions on detection efficacy were evaluated, and the application of TLC-SERS to on-site detection of artificial and real-life samples of aromatics/polluted water was systematically investigated. It was shown that commercially available Si 60-F(254) TLC plates were suitable for separation and displayed low SERS background and good separation efficiency, 2 mM silver colloids, 20 mM NaCl (working as aggregating agent), 40 mW laser power, and 50 s intergration time were appropriate for the detection regime. Furthermore, qualitative and quantitative detection of most of substituted aromatic pollutants was found to be readily accomplished using the developed TLC-SERS technique, which compared well with GC-MS in terms of identification ability and detection accuracy, and a limit of detection (LOD) less than 0.2 ppm (even at ppb level for some analytes) could be achieved under optimal conditions. The results reveal that the presented convenient method could be used for the effective separation and detection of the substituted aromatic pollutants of water on site, thus reducing possible influences of sample transportation and contamination while shortening the overall analysis time for emergency and routine monitoring of the substituted aromatics/polluted water.     

不同原料制备的酶解辛烯基琥珀酸淀粉酯作微胶囊壁材的性质比较

主要研究了不同原料的酶解辛烯基琥珀酸淀粉酯的制备及其性质特征,着重比较了其粘度、乳化稳定性、包埋能力及作为壁材的微胶囊油脂的贮藏稳定性。     

内蒙某低品位金矿浸出的试验研究

以内蒙古乌拉特中旗浩尧尔忽洞金矿(217金矿)矿石为研究对象,采用实验室柱浸试验,研究不同的粒度、氰化钠浓度、喷淋强度以及助浸剂添加量对金矿浸出速度的影响.根据试验的结果,提出了该金矿浸出适宜的工艺条件.     

抗菌肽对热应激肉鸡血液学及血氧自由基的影响

了解猪小肠抗菌肽在热应激条件下对肉鸡血液学及血氧自由基的影响。方法 选用30只两周龄健康华都AA+肉鸡,称重后随机分为3组:常温对照组、单纯热应激组和抗菌肽+热应激组,每组10只,热应激10天。结果 抗菌肽+热应激组与单纯热应激组比较全血黏度、血浆黏度、红细胞压积明显降低(P＜0.01);抗菌肽+热应激组血清中总蛋白(TP)、球蛋白(GLO)、葡萄糖(GLU)及谷丙转氨酶(ALT)、谷草转氨酶(AST)、碱性磷酸酶(ALP)与单纯热应激组比较均升高;总胆固醇(TC)、甘油三酯(TG)、尿素(UREA)、低密度脂蛋白(LDL)、高密度脂蛋白(HDL)与单纯热应激组比较明显降低;抗菌肽+热应激组SOD、GSH-Px活性与单纯热应激组比较明显升高(P＜0.01);MDA含量与单纯热应激组比较差异有统计学意义(P＜0.01)。结论 热应激条件下口服抗菌肽可以调节肉鸡血液成份及流变状态、增强体液免疫、平衡氧自由基、降低氧化应激反应,使机体有效抵抗热应激。     

The amino acid composition,solubility and emulsifying properties of sweet potato protein

A protein was purified from the high-protein type sweet potato variety 55-2 available in China. The amino acid composition, solubility and emulsifying properties of the sweet potato protein (SPP) were studied. The SPP was rich in aspartic acid (18.5%) and glutamic acid (9.30%) while essential acid amino acids made up approximately 40.7% of the SPP. The SPP was highly soluble in distilled water over a wide range of pH. However, solubility of the SPP in 1.0 M NaCl and 1.0 M CaCl₂ solutions was low especially at pH below the pI of the SPP. The SPP in CaCl₂ demonstrated emulsifying activity index (EAI) and emulsion stability index (ESI) many folds higher than those in distilled water and NaCl solution (P < 0.05).