氮氧化SODm/COm的设计与合成及其体外活性

基于对天然CuZn-SOD酶的分析,本文选取功能氮氧化异烟酸(HINO)配体与过渡金属Cu(II)通过混合溶剂热法合成了一例兼具双酶活性的配合物1,[Cu4(Hino)4(OH)2SO4(H2O)4]n.通过单晶X-射线衍射仪(SC-XRD)、粉末X-射线衍射仪(PXRD)和傅里叶红外光谱仪(FTIR)等对其进行系统的表征.NBT光还原法(NBT,氯化硝基四氮唑蓝)测试显示,配合物1的IC50值为0.43μmol/L,表明其具有优良的抗氧化性能.同时,体外儿茶酚酶模拟测试,配合物1的kcat值为3.166×10^(3)h^(-1),可作为中等强度的酶催化剂.

丙烯硫脲对日本血吸虫酚酶抑制作用的组织化学研究

目的 通过组织化学方法研究丙烯硫脲对小鼠体内、体外日本血吸虫成虫酚酶 (phenol oxidase,PO)活性的抑制作用。方法 将 4 2 d日本血吸虫活成虫置于含 2 5 mmol/L 邻苯二酚的磷酸盐缓冲液 (p H6 .8)中 ,2 8℃下孵育 30 min,取出虫体置载玻片上 ,加 2滴含 0 .1%戊巴比妥钠的 PBS溶液 ,置 L eica荧光显微镜下 ,分别在普通光照和紫外光激发下观察酚酶在虫体内的组织学定位。体外抑制实验时 ,加底物前先将成虫置于含 5mm ol/L 丙烯硫脲的 PBS溶液中 ,2 8℃下孵育 10 min;体内抑制实验时 ,在小鼠感染日本血吸虫尾蚴 2 0 d后 ,按30 0 m g/kg隔日 1次腹腔注射丙烯硫脲。结果 体外抑制实验组雌虫卵黄腺及子宫内虫卵卵壳和雄虫体壁表层的桔黄色荧光或赭褐色颜色反应均基本消失 ;体内抑制实验组雌虫子宫内未见虫卵或虫卵样物质 ,仅见大量泥沙样卵黄颗粒 ,其荧光或颜色反应也基本消失。结论 通过一种更灵敏的显示酚酶活性的组织学定位方法 ,观察到不仅雌虫含有酚酶 ,雄虫也有酚酶活性。证明丙烯硫脲对血吸虫成虫的酚酶活性有明显的抑制作用 。

苯甲酸对蘑菇酪氨酸酶抑制作用机理的研究

报道了苯甲酸对蘑菇酪氨酸酶的单酚酶和二酚酶活力的影响以及抑制作用机理.研究结果表明,苯甲酸对蘑菇酪氨酸酶的单酚酶和二酚酶活性均有抑制作用,其效应为可逆抑制效应.测定导致单酚酶活力和二酚酶活力下降50%的抑制剂浓度(IC50)分别为1.20和1.00mmol/L.苯甲酸对蘑菇酪氨酸酶的单酚酶的迟滞时间有明显的延长效应,4mmol/L苯甲酸使得单酚酶的迟滞时间从42s延长到200s.测定苯甲酸对二酚酶的抑制作用表现为非竞争性抑制类型,测定抑制常数为0.95mmol/L.苯甲酸与酶的结合导致天然酶的内源荧光的量子产率下降,但荧光发射峰没有位移.作者提出抑制剂与酶分子的作用模型.

桑黄素对蘑菇酪氨酸酶的抑制作用

研究桑黄素(Morin)对蘑菇酪氨酸酶单酚酶和二酚酶活力的影响和抑制效应的结果表明:桑黄素对蘑菇酪氨酸酶的单酚酶和二酚酶活性均有抑制作用;导致单酚酶活力和二酚酶活力下降50%的抑制剂浓度(IC50)分别为0.08和1.02mmol/L;桑黄素对蘑菇酪氨酸酶的单酚酶的迟滞时间有明显的延长效应,0.175 mmol/L桑黄素使得单酚酶的迟滞时间从24 s延长到362 s,迟滞时间增加了14倍,酶的稳态活力下降了61.9%;桑黄素对二酚酶的抑制作用表现为竞争性可逆抑制作用,抑制常数为0.55 mmol/L.

维生素B_1对蘑菇酪氨酸酶抑制作用机理的研究

报道了维生素B1对蘑菇酪氨酸酶的单酚酶和二酚酶活力的影响以及抑制作用机理.研究结果表明,维生素B1(Vit B1)对蘑菇酪氨酸酶的单酚酶和二酚酶活性均有抑制作用.测定导致单酚酶活力和二酚酶活力下降50%的抑制剂浓度(IC50)分别为15和20 mmol/L.Vit B1对蘑菇酪氨酸酶的单酚酶的迟滞时间有明显的延长效应,21 mmol/L Vit B1使得单酚酶的迟滞时间从52 s延长到277 s.Vit B1对二酚酶的抑制作用表现为竞争性抑制类型,测定抑制常数(KI)为11.7 mmol/L.

4-氨基苯甲酸对蘑菇酪氨酸酶的抑制作用机理研究

报道了4-氨基苯甲酸(4-aminobenzoic acid)对蘑菇酪氨酸酶单酚酶和二酚酶活力的影响和抑制效应.结果表明,4-氨基苯甲酸对蘑菇酪氨酸酶的单酚酶和二酚酶活性均有抑制作用.导致单酚酶活力和二酚酶活力下降50%的抑制剂浓度(IC50)分别为0.35和0.30 mmol/L.4-氨基苯甲酸对单酚酶的迟滞时间有明显的延长效应,1.0 mmol/L可使单酚酶的迟滞时间从30.5 s延长到155.1 s,增加了4.4倍.4-氨基苯甲酸对二酚酶的抑制作用表现为混合型可逆抑制作用,抑制常数(KI和KIS)分别为0.259和1.46 mmol/L.

Dawson结构的多金属氧酸盐对酪氨酸酶的抑制作用

按文献方法合成了两种Dawson结构的多金属氧酸盐,并对其结构进行紫外光谱和红外光谱分析。以H_6[P_2Mo_(18)O_(62)]和H_8[P_2Mo_(17)Cr(OH_2)O_(61)](简写为P_2Mo_(18)和P_2Mo_(17)Cr)为效应物,采用紫外分光光度法和酶动力学方法研究两种Dawson结构的多金属氧酸盐效应物对蘑菇酪氨酸酶二酚酶活性的抑制作用。结果表明,P_2Mo_(18)和P_2Mo_(17)Cr对酪氨酸酶二酚酶均具有显著的抑制效果,测定抑制酪氨酸酶活力下降50%的抑制浓度(IC_(50))分别为(0.482±0.009)mmol/L和(0.503±0.011)mmol/L。动力学分析表明,P_2Mo_(18)和P_2Mo_(17)Cr对酪氨酸酶的抑制作用均表现为可逆的竞争型,抑制常数K_I分别为0.212和0.249 mmol/L。其中,综合考虑IC_(50)值和抑制常数,P_2Mo_(18)对酪氨酸酶二酚酶活性的抑制效果略优于P_2Mo_(17)Cr。

钒取代的Dawson型磷钼酸对酪氨酸酶的抑制作用

采用酶动力学方法研究了5种钒取代的Dawson型磷钼酸H_7[P_2Mo_(17)VO_(62)]、H_8[P_2Mo_(16)V_2O_(62)]、H_9[P_2Mo_(15)V_3O_(62)]、H_8[P_2Mo_(14)V_4O_(62)H_2]和H_9[P_2Mo_(13)V_5O_(62)H_2](分别简写为P_2Mo_(17)V、P_2Mo_(16)V_2、P_2Mo_(15)V_3、P_2Mo_(14)V_4和P_2Mo_(13)V_5)对蘑菇酪氨酸酶二酚酶的抑制作用,结果表明,效应物P_2Mo_(17)V、P_2Mo_(16)V_2和P_2Mo_(15)V_3能够明显地抑制酪氨酸酶的活性,其半抑制浓度(IC_(50))值分别为0.409、0.386和0.386 mmol/L,且均表现为可逆的竞争型抑制,效应物P_2Mo_(17)V、P_2Mo_(16)V_2和P_2Mo_(15)V_3对游离酶的抑制常数K_I分别为0.234、0.391和0.249 mmol/L。而效应物P_2Mo_(14)V_4在0~1.0 mmol/L浓度范围内,对酪氨酸酶二酚酶无明显抑制作用,效应物P_2Mo_(13)V_5对酪氨酸酶二酚酶表现为激活作用。

贵州高原草海湿地土壤有机碳分布特征及其与酶活性的关系

基于时空替代的研究方法,对贵州草海湿地土壤有机碳和氧化还原酶活性分布进行了研究,分析了土壤总有机碳(TOC)和可溶性有机碳(DOC)与土壤酶活性、环境因子之间的关系.结果表明,不同退化梯度湿地土壤中有机碳存在较大的差异,沿着退化梯度表层土壤TOC、DOC含量和脱氢酶(DHA)活性逐渐降低(p<0.05),而多酚酶(PPO)活性逐渐升高(p<0.05);在垂直剖面上,原生湿地(CH1)和轻度退化湿地(CH_2)土壤TOC先增加后减少,而沼泽化湿地(CH3)和草甸湿地(CH_4)土壤TOC无明显的分层和富聚现象;相关性分析表明,TOC、DOC与PPO显著负相关(p<0.01),而PPO与土壤含水率(WHC)、总氮(TN)和硝酸盐(NO-3-N)含量等显著负相关(p<0.01),与pH显著正相关(p<0.01).以上结果表明,土壤有机碳和酶活性的变化是对湿地退化的响应,湿地退化碳库的损失与PPO活性增加有关,而WHC、TN、NO-3-N和pH是影响PPO活性分布的重要因子.

Differences in Phenolic Compounds and Related Enzyme Activities between 2 Eucalyptus Species

[Objective] The aim was to provide theoretical basis for speeding up the popularization of Eucalyptus dunnii.[Method] Differences in phenolic compounds and related enzyme activities between hard rooting E.dunnii and easily rooting E.urophylla×E.grandis clone （U9） were studied.[Result] There were significant differences in contents of phenolic compounds and activities of indole acetic acid oxidase （IAAO）,andperoxidase （POD） and polyphenol oxidase （PPO） between the two Eucalyptus species.Compared with U9,contents of phenolic compounds and POD activity of E.dunnii cuttings were relatively high,while activities of IAAO and PPO were relatively low.[Conclusion] More phenolic compounds and the differences in related enzyme activities might be the major factors to result in hard rooting of E.dunnii cuttings.

Effects of Used Battery on Key Enzyme Activity during the Germination of Wheat Seeds

[Objective] This study aimed to explore the effects of used battery lixivium on wheat germination. [Method] The wheat seeds were treated with used battery lix- ivium at different concentrations to detect the change of activities of amylase, pro- tease, pyruvate dehydrogenase （PDH） and polyphenol oxidase （PPO） during the ger- mination period. [Result] The results showed that the used battery affected enzyme activity. With the increase of concentration of used battery lixivium, trends of the changes of amylase and protease activities were not different. The activities were en- hanced at low concentrations of lixivium, while were inhibited at high concentrations. The tends of changes of pyruvate dehydrogenase （PDH） and polyphenol oxidase （PPO） activities were not consistent with that of either amylase or protease, which showed continuous downward trends with the increasing concentration of used battery lixivium. [Conclusion] This study is of great practical significance for understanding the effects of used battery lixivium on the germination of wheat seeds.

Enhanced resistance of Portunus trituberculatus to Vibrio alginolyticus by selective breeding

We established a line (screened) of Portunus trituberculatus by selectively breeding individuals that survived from challenge with Vibrio alginolyticus, and compared the response of screened and unscreened (control) P. trituberculatus challenged with V. alginolyticus. We measured superoxide dismutase, catalase, acid phosphatase, alkaline phosphatase, and peroxidase activity and the content of hemocyanin in the plasma and phenoloxidase activity in serum. The cumulative survival rate after 24-h challenge with V. alginolyticus was significantly higher in the screened crabs than in the unscreened crabs (P<0.05). T-SOD and PO activity were significantly lower in the screened stock than in the unscreened stock (P<0.05). POD, CAT, and ACP activity and hemocyanin content were significantly higher in the screened stock than in the unscreened stock. Our results suggest that the screened stock was more resistant to infection. Furthermore, the indices we measured may be used to evaluate the health state of P. trituberculatus.

Detection and Quantitative Analysis of Dominant Bacteria On Aging Flue-cured Tobacco Leaves

[Objective] The aim was to construct the internal motivation of flued-cured tobacco natural aging. [Method] The variation of DNA copies of Bacillus megaterium, amylase activity, PPO, peroxydase and content of starch, neutro-protein, polyphenol in different aging period were investigated using Henan flue-cured tobacco leaves. [Result] Results showed that the DNA copies of Bacillus megaterium increased initially but decreased subsequently with the aging time. The DNA copies of Bacillus per gram of tobacco leaf surface reached the maximum number at 6 months aging. The activities of four enzymes showed the same variation trend as in DNA copies, and also reached the highest level at 6 months aging. Chemical ingredients, however, displayed a downward trend. The variation of enzyme activity and content of chemical composition were reflected in the changes of DNA copies in Bacillus megaterium. [Conclusion] The result indicate that Bacillus megaterium was the internal motivation of flue-cured tobacco natural aging.

Polyphenol Oxidase Activity and Inhibition in White Yam （Dioscorea Rotundata. Var. Laasirin） Chips as African Fries for Human Consumption

Polyphenol oxidase, a bi-functional enzyme, has been implicated in enzymatic browning of yam and other tubers in a negative way. The objective of this present study was to examine the activity of polyphenol oxidase in Dioscorea rotundata. Var. laasirin and the efficiency of heat and chemical treatments in inhibiting this enzyme. Crude Polyphenol Oxidase （PPO） of Dioscorea rotundata.Var. Laasirin was isolated and the kinetics studied using the lineweaver-burk plot. The activity of the enzyme was evaluated using spectrophotomeric method. Yam PPO catalyzes oxidation of various substrates with catechol being the most readily oxidized substrate. The Michaelis-Menten constant （Km） and maximum reaction velocity （Vmax） for yam PPO were 0.00037 and 0.3125 respectively. Inhibition data showed that the enzyme had least activity （71.70） when blanched at 95℃ for 7 mins with chemical treatment involving a combination of 0.5% Sodium metabisulphite （Food grade） and 0.5% Ascorbic acid （Food grade）. The activity was highest （83.02） when it was blanched at 95 ℃ for 7 rains. This study has shown that it is possible to inhibit polyphenol oxidase activity in white yam using the chemical pretreatments and processing conditions described in this study for possible adoption in the production of packaged frozen yam chips by food industries.

Inhibitory effect of Cr(Ⅵ) on activities of soil enzymes

To evaluate the influence of various Cr(Ⅵ) concentrations (0.05, 0.25, 0.50, 1.00 and 2.00 g/kg) on the activity of soil enzymes, the activities of catalase, polyphenol oxidase, dehydrogenase, alkaline phosphatase in soils were investigated in the incubation experiment with a period of 35 d. The results indicate that all the tested Cr(Ⅵ) concentrations significantly inhibit dehydrogenase activity by over 70% after 35 d. The activity of alkaline phosphatase is slightly inhibited during the whole experiment except for on the day 7. Cr(Ⅵ) has no obvious effect on the activity of catalase in soil. On the contrary, Cr(Ⅵ) stimulates the activity of polyphenol oxidase. The results suggest that dehydrogenase activity can be used as an indicator for assessing the severity of chromium pollution.

Physical, Chemical and Biochemical Changes of Sweetsop （Annona squamosa L.） and Golden Apple （Spondias citherea Sonner） Fruits during Ripening

This study aimed at the physical, chemical and biochemical changes during ripening of Sweetsop （Annona squamosa L.） and Golden Apple （Spondias citherea Sonner） fruits during ripening as important features to better understand their postharvest handling. It was carried out physical analysis such as firmness and chemical analysis such as total chlorophyll, total carotenoids, soluble solids, pectins and titrable acidity and biochemical analysis such as pectin methyl esterase, polygalacturonase, cellulase, and peroxidase and polyphenoloxidase activities in crude extract. Fruits were harvested at different stages of ripening. Experimental design was completely randomized and was carried out analysis of variance and Tukey tests, Total chlorophyll was decreasing in later stages of ripening, total soluble solid contents increased as the fruits ripen, while the acidity expressed percentage of citric acid decreased during fruits ripening. The loss of firmness and soluble solids content increased as the fruit get ripped stage, while the content of pectin decreased. Activity was observed for pectin methyl esterase and polygalacturonase enzymes during all stages of maturation, presenting the highest activity for both enzymes in the mature state. No cellulase activity detected at any stage during the ripening of these fruits. Activity of the enzyme polyphenoloxidase and peroxidase, associated with pulp browning was higher in the last stages of ripening of these fruits. Physical, chemical and biochemical patterns during ripening were different according to fruit species suggesting differential postharvest handling requirements.

Inhibition of Acrosomal Enzymes by Gossypol Is Related to Its Antifertility Action

To investigate into the mechanisms underlying the irreversible sterility induced by gossypol, we studied the relationship between its inhibitory action on acrosomal enzymes and its antifertility effect.As shown by our result, after exposure to gossypol (l.25-60 μg/ml) for 15 min. in vitro,the sperms' ability to penetrate bovine cervical mucus and the fertility rate were significantly reduced. Also, following administration of gossypol (12.5 mg/kg/day) for six weeks, the rate of fertilization in vitro by hamster sperm was significantly decreased. In the gossypol-treated group, extracts of testis sperm delayed dispersion of cumulus cells, suggesting inhibition of hyaluronidase and other acrosomal enzymes. Furthermore, the acrosin and arylsulfatase activities were shown to be markedly inhibited. Thus, a parallelism was displayed between the reduction of fertility and the decreasc in acrosin and arylsulfatase activities in epididymis sperms.Besides, the inhibition was reversible and was dosage-and durationdependent. In conclusion, the assay of acrosin activity might serve as a useful tool for monitoring the irreversible sterility induced by gossypol,

Synthesis of three bromophenols from red algae as PTP1B inhibitors

Bromophenols are a set of natural products widely distributed in seaweed, most of which exhibit interesting and useful biological activities. To develop a reliable and efficient synthetic route to these natural bromophenols, three of them, 3,4-dibromo-5-(2′-bromo-3′,4′- dihydroxy-6′-methoxymethyl- benzyl)-benzene-1,2-diol (compound 9), 3,4-dibromo-5-(2′-bromo-6′-ethoxy methyl-3′,4′-dihydroxybenzyl)- benzene-1,2-diol (compound 10), and 3-bromo-4-(3′-bromo-4′,5′-dihydroxy benzyl)-5-(ethoxymethyl)- benzene-1,2-diol (compound 14), isolated from red marine algae, have been synthesized in eight steps with an overall yield of 14.4%, 14.4%, and 18.2% respectively, via a practical approach employing bromination, Wolff-Kishner-Huang reduction and a Friedel-Crafts reaction as key steps. The protein tyrosine phosphatase 1B (PTP1B) inhibitory activities of the synthetic compounds were evaluated by the colorimetric assay. The results show that these compounds are moderate PTP1B inhibitors. The synthesis of these bromophenol derivatives makes in vivo studies of their structure-activity relationships and inhibition activity against PTP1B possible.

Cause and control of Radix Ophiopogonis browning during storage

In the storage of Radix Ophiopogonis, browning often happens to cause potential risk with regard to safety. Previously few reports investigate the browning of Radix Ophiopogonis. In this research, the causes and mechanisms of the browning of Radix Ophiopogonis were preliminarily elucidated. Content determination by high-performance liquid chromatogra- phy （HPLC） and spectrophotometry, enzyme activity determination by colorimetry, and morphological observation by electron mi- croscopy were performed in the present study. Uniform design and three-dimensional response surfaces were applied to investigate the relationship between browning and storage factors. The cortex cell wall of browned Radix Ophiopogonis was ruptured. Compared with the normal Radix Ophiopogonis, cellulase and polyphenol oxidase enzymes were activated, the levels of 5-hydroxymethylfurfural （5-HMF）, total sugars, and reducing sugars were increased, while the levels of polysaccharides and methylophiopogonanone A were de- creased in browned Radix Ophiopogonis. The relationship between the storage factors and degree of browning （Y） could be described by following correlation equation： Y= - 0.625 4 ＋ 0.020 84 × X3 ＋ 0.001 514×X1×x X2 - 0.000 964 4 × X2 × X3. Accompanied with browning under storage conditions, the chemical composition of Radix Ophiopogonis was altered. Following the activation of cellulase, the rupture of the cortex cell wall and the outflow of cell substances flowed out, which caused the Radix Ophiopogonis tissue to become soft and sticky. The main causes of the browning were the production of 5-HMF, the activation of polyphenol oxidase, Maillard reactions and enzymatic browning. Browning could be effectively prevented when the air relative humidity （HR）, temperature, and moisture content were under 25% RH, 12 ℃ and 18%, respectively.

Lipid reserves and immune defense in healthy and diseased migrating monarchs Danaus plexippus

Recent studies suggest that the energetic demands of long-distance migration might lower the pool of resources available for costly immune defenses. Moreover, migration could amplify the costs of parasitism if animals suffering from para- site-induced damage or depleted energy reserves are less able to migrate long distances. We investigated relationships between long-distance migration, infection, and immunity in wild fall-migrating monarch butterflies Danaus plexippus. Monarchs migrate annually from eastern North America to central Mexico, accumulating lipids essential for migration and winter survival as they travel southward. Monarchs are commonly infected by the debilitating protozoan parasite Ophryocystis elektroscirrha （OE）. We collected data on lipid reserves, parasite loads, and two immune measures （hemocyte concentration and phenoloxidase activity） from wild monarchs migrating through north GA （USA） to ask whether （1） parasite infection negatively affects lipid reserves, and （2） greater investment in lipid reserves is associated with lower immune measures. Results showed that monarchs sampled later in the fall migration had lower but not significantly different immune measures and significantly higher lipid reserves than those sampled earlier. Lipid measures correlated negatively but only nearly significantly with one measure of immune defense （phenoloxidase activity） in both healthy and infected monarchs, but did not depend on monarch infection status or parasite load. These results provide weak support for a trade-off between energy reserves and immune defense in migrants, and suggest that previously-demonstrated costs of OE infection for monarch migration are not caused by depleted lipid reserves .