Inhibitory Kinetics of p-Substituted Benzaldehydes on Polyphenol Oxidase from the Fifth Instar of Pieris Rapae L.

Polyphenol oxidase (PPO) is the enzyme responsible for enzymatic browning during the growth of insects. It is also involved in defense reactions and is related with immunities in insects. PPO, a metalloenzyme oxidase, catalyzes the oxidation of ο-diphenol to ο-quinone. The present paper de- scribes the effects of benzaldehyde and its p-substituted derivatives on the activity of PPO from the fifth instar of Pieris rapae L. PPO from the fifth instar of Pieris rapae L. was purified using ammonium sulfate fractionation and chromatography on Sephadex G-100. The enzyme kinetics was characterized using L-3,4-dihydroxyphenylalanine (L-DOPA) as substrate. The results show that benzaldehyde, p- hydroxybenzaldehyde, p-chlorobenzaldehyde, and p-cyanobenzaldehyde can inhibit the PPO activity for the oxidation of L-DOPA. The inhibitor concentration leading to 50% activity lost, IC50, was esti- mated to be 5.90, 5.62, 2.83, and 2.91 mmol/L for the four tested inhibitors, respectively. Kinetic analy- ses show that the inhibitory effects of these compounds are reversible. Benzaldehyde, p- hydroxybenzaldehyde, and p-chlorobenzaldehyde are noncompetitive inhibitors while p-cyano- benzaldehyde is a mixed-type inhibitor. The inhibition constants were determined for all four inhibitors. p-chlorobenzaldehyde and p-cyanobenzaldehyde were more potent inhibitors than the other com- pounds. These results provide a basis for developing PPO inhibition-based pesticides.     

榨菜中多酚氧化酶特性的研究

为探讨榨菜的保鲜及防褐变技术,对榨菜中多酚氧化酶的研究表明,榨菜中多酚氧化酶以邻苯二酚为底物时,产物最大吸收波长是420 nm,酶的最适pH为7.4,最适反应温度为53℃,Km为0.181 9 mol.L-1.该酶在85℃时仍有较强热稳定性,L-半胱氨酸盐酸盐对该酶活性有很强的抑制作用.     

真菌诱导物对丹参毛状根氧化酶活性的影响

从一种广谱病原真菌－大丽轮枝菌的菌丝体中制备激发子,用该激发子和商品生物制剂－酵母提取物分别诱导对数生长期的丹参毛状根,发现丹参毛状根过氧化物酶（POD）,过氧化氢酶（CAT）和多酚氧化酶（PPO）活性在诱导后均显著增加,三种酶的活性并随诱导时间和诱导剂的浓度的增加而改变,大丽轮枝菌激发子的作用强于酵母提取物,表现为诱导酶活性峰值较高和峰值时间提前。     

Diversity and dispersal history of the talitrids (Crustacea: Amphipoda: Talitridae) of Bermuda

Five taxa of talitrid amphipods were found in the archipelago of Bermuda, of which three were recorded there for the first time. Four of these are supralittoral wrack generalists: Platorchestia monodi BOLD:AAB3402, (a unique Molecular Operational Taxonomic Unit according to the Barcode Index Number system), a related species recognized by molecular methods, Platorchestia platensis BOLD:AAA2949, Mexorchestia carpenteri carpenteri BOLD:AAC1491 and Tethorchestia antillensis; and one a terrestrial leaf-litter generalist: Talitroides alluaudi. A key is provided to discriminate between the formally described talitrids of Bermuda. Dispersal mechanisms from the American continent to Bermuda were considered for all taxa based on species distributions along the North American Atlantic coast and also investigated by molecular methods, using genetic population differentiation and haplotype network analysis based on the barcode region of cytochrome c oxidase subunit I gene. For P. monodi BOLD:AAB3402 the genetic results suggest that some dispersal events occurred before human colonization of Bermuda but are equivocal about the source population and therefore the direction of dispersal. Some very recent synanthropic dispersal is possible with this species. For the other two species studied genetically, P. platensis BOLD:AAA2949 and M. c. carpenteri BOLD:AAC1491, the small population samples analysed support dispersal to Bermuda from the American mainland, before human occupation of Bermuda, although the available sample size was limited for these species. The available limited direct, non-genetic evidence supports synanthropic transport for Talitroides alluaudi. Platorchestia monodi BOLD:AAB3402 is found in the same wrack habitat as P. platensis BOLD:AAA2949 on Bermuda, apparently without interbreeding. No evidence was found that driftwood specialist talitrids had become established in Bermuda.     

Identification and characterization of Biomphalaria peregrina (Orbignyi, 1835) from Agua Escondida in northern Patagonia,Argentina

During a general malacological survey for freshwater gastropods in northern Patagonia, a population of Biomphalaria was encountered at Agua Escondida. Biomphalaria spp. are freshwater pulmonates of biomedical importance, uncommon in Mendoza Province, and often act as intermediate hosts for Schistosoma mansoni. By looking at both morphological and molecular characters, we describe a detailed process of identification and characterization of Biomphalaria peregrina from a location towards the extremity of its species range. A reference DNA ‘barcode’ is presented. As B. peregrina has been shown to be a permissive experimental host of S. mansoni, snails were also screened in the field for schistosomiasis and later in the laboratory using a novel polymerase chain reaction-based assay but no infections were found. Considering the transmission potential of this species, increased vigilance for intestinal schistosomiasis is recommended, especially if local environmental conditions become favourable for disease transmission, for example, through future climate change and intensification of irrigation.     

溶血对氧化酶法测定葡萄糖含量影响的实验研究

目的探讨标本发生溶血后对氧化酶法测定葡萄糖所产生的影响及纠正方法.方法随机抽取门诊正常健康血液标本900例,溶血标准采用血浆游离血红蛋白的浓度来评价,葡萄糖浓度运用双波长法测定.最终采用二元一次方程回归,对比纠正前后存在的偏差值,并判定其纠正结果.结果经测定,溶血后的葡萄糖测定值低于实际值,且溶血程度越严重则受感染的结果越明显.最终所得的平均回归方程组为Z=0.415+0.053X+0.014Y,相关系数值r=0.974;经回归后纠正前偏差值δ1=-13.4%,纠正后偏差值δ2=-7.7%,纠正后差值在可允许误差范围±0.1内(P0.05).结论通过回归方程纠正后,葡萄糖浓度测定值和实际值存在的差值在可允许范围之内.     

壳聚糖—纳米金自组装葡萄糖生物传感器的研究

利用壳聚糖与纳米金构建一种新型的葡萄糖生物传感器。探究不同浓度的葡萄糖溶液、p H及温度对传感器响应电流的影响。结果表明,此传感器在温度为35.0℃,p H=7.00,葡萄糖质量浓度为0.018~25.7 mg/L范围内有良好的线性,线性I=4.135 6 m+91.787μA,R2=0.997 9,检出限为0.001 2 mg/L。该传感器具有制备简单、方法简便、灵敏度高,能快速检测等优点。     

冬眠及外源维生素C对中华鳖(Pelodiscus sinensis)稚鳖能量代谢的影响

观测了冬眠前饲料中添加外源维生素C(Vc)对冬眠中华鳖稚鳖能量代谢的影响.结果发现外源Vc不能提高冬眠期间中华鳖的存活率(P=0.57).稚鳖个体在冬眠期间生长能的平均值为-0.15 kJ.d-1,该值与代谢耗能相当.鳖体蛋白、脂肪、水分和灰分质量分数w在冬眠前后无显著差异(P>0.05),外源Vc对鳖的体组成没有明显影响(P>0.05).冬眠8周后,肝脏线粒体细胞色素C氧化酶(COX)和肝脏乳酸锐氢酶(LDH)的活性显著下降(P<0.05).饲料外源Vc对肝脏线粒体蛋白含量、肝脏线粒体COX和LDH的活性均无影响(P>0.05).     

Improving the thermal stability of lactate oxidase by directed evolution

Lactate oxidase is used in biosensors to measure the concentration of lactate in the blood and other body fluids. Increasing the thermostability of lactate oxidase can significantly prolong the lifetime of these biosensors. We have previously obtained a variant of lactate oxidase from Aerococcus viridans with two mutations (E160G/V198I) that is significantly more thermostable than the wild-type enzyme. Here we have attempted to further improve the thermostability of E160G/V198I lactate oxidase using directed evolution. We made a mutant lactate oxidase gene library by applying error-prone PCR and DNA shuffling, and screened for thermostable mutant lactate oxidase using a plate-based assay. After three rounds of screening we obtained a thermostable mutant lactate oxidase, which has six mutations (E160G/V198I/G36S/T103S/A232S/F277Y). The half-life of this lactate oxidase at 70 °C was about 2 times that of E160G/V198I and about 36 times that of the wild-type enzyme. The amino acid mutation process suggests that the combined neutral mutations are important in protein evolution. Received 15 September 2006; received after revision 21 October 2006; accepted 2 November 2006