Screening of Human CYP1A2 and CYP3A4 Inhibitors from Seaweed In Silico and In Vitro

Phenolic compounds and carotenoids are potential inhibitors of cytochrome P450s. Sixteen known compounds, phenolic compounds and carotenoids from seaweed were examined for potential inhibitory capacity against CYP1A2 and CYP3A4 in silico and in vitro. Morin, quercetin, and fucoxanthin inhibited the enzyme activity of CYP1A2 and CYP3A4 in a dose-dependent manner. The IC₅₀ values of morin, quercetin, and fucoxanthin were 41.8, 22.5, and 30.3 μM for CYP1A2 and 86.6, 16.1, and 24.4 μM for CYP3A4, respectively. Siphonaxanthin and hesperidin did not show any significant effect on CYP1A2, but they slightly inhibited CYP3A4 activity at high concentrations. In silico modeling of CYP’s binding site revealed that the potential inhibitors bound in the cavity located above the distal surface of the heme prosthetic group through the 2a or 2f channel of CYPs. This study presents an approach for quickly predicting CYP inhibitory activity and shows the potential interactions of compounds and CYPs through in silico modeling.     

Nitrifier dominance of Arctic soil nitrous oxide emissions arises due to fungal competition with denitrifiers for nitrate

Arctic soils emit nitrous oxide, which is a potent greenhouse gas and also represents an important loss of nitrogen to oligotrophic Arctic ecosystems. However, little is known about the temperature sensitivity of nitrous oxide release in Arctic soils or the organisms mainly responsible for it. We investigated controls on nitrous oxide emissions in an Arctic soil across a typical temperature range (between 4 and 13 °C) on Truelove Lowland, Devon Island, Canada (75°40′N 84°35′W) at two different moisture contents. When fertilized with ammonia or nitrate, nitrous oxide emissions and temperature dependence of nitrous oxide emissions were insensitive to soil moisture content but linked to nitrification rates. Stable isotope analysis revealed that nitrous oxide was predominantly released by nitrifiers. However, nitrous oxide emissions were not linked to nitrifier prevalence with an insignificant (P < 0.219) increase in amoA genes and a (P < 0.01) decrease in archaeal nitrifiers. In contrast, denitrifier nosZ prevalence was 10,000 times greater than that of nitrifiers and was related to nitrous oxide emission potential when soils were fertilized with nitrate. Manipulating water-filled pore space should have changed the pattern of N₂O emissions. We used selective inhibitors to further explore why denitrification did not occur under field conditions when we manipulated water-filled pore space or when we used ¹⁵N analysis. When fungi were inhibited in the soil, nitrous oxide emissions from denitrifiers increased with no change in nitrous oxide released by nitrifiers. When fungi were active in the soil, there was little available nitrate but when fungi were inhibited, available soil nitrate increased over the incubation period. The dominance of nitrifiers in nitrous oxide emissions from Arctic soils under field conditions is linked to the competition for nitrate between fungi and denitrifiers.     

Nitrogen transformations in cold and frozen agricultural soils following organic amendments

Though microbial activity is known to occur in frozen soils, little is known about the fate of animal manure N applied in the fall to agricultural soils located in areas with prolonged winter periods. Our objective was to examine transformations of soil and pig slurry N at low temperatures. Loamy and clay soils were either unamended (Control), amended with ¹⁵NH₄-labeled pig slurry, or amended with the pig slurry and wheat straw. Soils were incubated at −6, −2, 2, 6, and 10 °C. The amounts of NH₄, NO₃ and microbial biomass N (MBN), and the presence of ¹⁵N in these pools were monitored. Total mineral N, NO₃ and ¹⁵NO₃ increased at temperature down to −2 °C in the loam soil and −6 °C in the clay soil, indicating that nitrification and mineralization proceeded in frozen soils. Nitrification and mineralization rates were 1.8-4.9 times higher in the clay than in the loamy soil, especially below freezing point (3.2-4.9), possibly because more unfrozen water remained in the clay than in the loamy soil. Slurry addition increased nitrification rates by 3-14 times at all temperatures, indicating that this process was N-limited even in frozen soils. Straw incorporation caused significant net N immobilization only at temperatures ≥2 °C in both soils; the rates were 1.4-3.4 higher in the loam than in the clay soil. Nevertheless, up to 30% of the applied ¹⁵N was present in MBN at all temperatures. These findings indicate that microbial N immobilization occurred in frozen soils, but was not strong enough to induce net immobilization below the freezing point, even in the presence of straw. The Q₁₀ values for estimated mineralization and nitrification rates were one to two orders-of-magnitude larger below 2 °C than above this temperature (13-208 versus 1.5-6.9, respectively), indicating that these processes are highly sensitive to a small increase in soil temperature around the freezing point of water. This study confirms that net mineralization and nitrification can occur at potentially significant rates in frozen agricultural soils, especially in the presence of organic amendments. In contrast, net N immobilization could be detected essentially above the freezing point. Our results imply that fall-applied N could be at risk of overwinter losses, particularly in fine-textured soils.     

Nitrous oxide emissions from fertilised grassland: A 2-year study of the effects of N fertiliser form and environmental conditions

The aim was to investigate the effects of different N fertilisers on nitrous oxide (N₂O) flux from agricultural grassland, with a view to suggesting fertiliser practices least likely to cause substantial N₂O emissions, and to assess the influence of soil and environmental factors on the emissions. Replicate plots on a clay loam grassland were fertilised with ammonium sulphate (AS), urea (U), calcium nitrate (CN), ammonium nitrate (AN), or cattle slurry supplemented with AN on three occasions in each of 2 years. Frequent measurements were made of N₂O flux and soil and environmental variables. The loss of N₂O-N as a percentage of N fertiliser applied was highest from the supplemented slurry (SS) treatment and U, and lowest from AS. The temporal pattern of losses was different for the different fertilisers and between years. Losses from U were lower than those from AN and CN in the spring, but higher in the summer. The high summer fluxes were associated with high water-filled pore space (WFPS) values. Fluxes also rose steeply with temperature where WFPS or mineral N values were not limiting. Total annual loss was higher in the 2nd year, probably because of the rainfall pattern: the percentage losses were 2.2, 1.4, 1.2, 1.1 and 0.4 from SS, U, AN, CN and AS, respectively. Application of U in the spring and AN twice in the summer in the 2nd year gave an average emission factor of 0.8% – lower than from application of either individual fertiliser. We suggest that similar varied fertilisation practices, modified according to soil and crop type and climatic conditions, might be employed to minimise N₂O emissions from agricultural land. Received: 30 August 1996     

植物生长抑制剂在草坪上的应用研究进展

综述了植物生长抑制剂对草坪草生长发育和生理特性的影响。并且指出 ,植物生长抑制剂可有效降低草坪草株、减少修剪次数、提高草坪草的抗逆性 ,从而降低管理费用     

一种适合于磷系和弱碱性水处理系统的聚合物阻垢剂

研究了以水为溶剂、过硫酸盐为引发剂合成的马来酸酐—烯丙基磺酸钠(MA—SAS)聚合物阻垢剂对磷酸钙、锌盐、碳酸钙和硫酸钙垢物的阻垢效果，探讨了介质pH值、Ca^2 浓度、温度和阻垢剂浓度对MA—SAS阻垢效果的影响．结果表明，MA—SAS共聚物在较宽pH值范围内对这些垢物均具有良好的抑制作用，既适用于磷系配方的循环水，也可用于中等硬度(160—240mg／L)、弱碱性(pH值8—9)和中等温度(50—60℃)的循环水系统．     

不同菜豆品种4种抗营养因子水平差异分析

菜豆中植物凝集素、皂苷、胰蛋白酶抑制剂和植酸等抗营养因子是引起菜豆食物中毒的内源物。以56份食荚菜豆品种为材料,测定了鲜荚中4种抗营养因子含量或活性。结果表明,供试菜豆品种群体4种抗营养因子水平均存在极显著差异。植物凝集素含量(均值为1.743 mg·g~(-1))和胰蛋白酶抑制剂活性(均值为1.680 mg·g~(-1))变异系数均在100%以上,品种频率分布曲线主峰明显,但在极高和极低区域均有品种间断分布,出现了品种水平的数量等级差异;皂苷含量(均值为3.730 mg·g~(-1))和植酸含量(均值为3.102 mg·g~(-1))变异系数均低于41%,品种频率分布曲线呈近正态分布,主峰明显,双尾有低频率连续分布。相关分析表明,植物凝集素含量与胰蛋白酶抑制剂活性呈极显著正相关。聚类分析将供试56份菜豆品种划分为3个品种群,近80%品种聚于抗营养因子中等水平品种群,近12%品种居于较低水平。由于菜豆植物凝集素是食用致毒性的主要内源物,同时其和胰蛋白酶抑制剂等均与菜豆田间抗病虫性密切相关,预示着筛选出极低水平植物凝集素的菜豆品种是可行的,但同时可能会降低其田间抗病虫性。     

Marker assisted accelerated introgression of null allele of kunitz trypsin inhibitor in soybean

Development of kunitz trypsin inhibitor (KTI)-free soybean is crucial for soy-food industry as the heat inactivation employed to inactivate the anti-nutritional factor in regular soybean incurs extra cost and affects protein solubility. In the presented work, a null allele of KTI from PI542044 was introgressed into cultivar ‘JS97-52’ (recurrent parent) through marker assisted backcrossing. Foreground selection in BC₁F₂, BC₂F₂ and BC₃F₂ was carried out using the null allele-specific marker in tandem with SSR marker Satt228, tightly linked with a trypsin inhibitor Ti locus. Background selection in null allele-carrying plants through 106 polymorphic SSR markers across the genome led to the identification of 9 KTI-free lines exhibiting 98.6% average recurrent parent genome content (RPGC) after three backcrosses, which otherwise had required 5–6 backcrosses through conventional method. Introgressed lines (ILs) were free from KTI and yielded at par with recurrent parent. Reduction of 68.8–83.5% in trypsin inhibitor content (TIC) in ILs compared to the recurrent parent (‘JS97-52’) was attributed to the elimination of KTI.     

A Dominant Inhibitor Gene Inhibits the Expression of Ht2 against Exserohilum turcicum Race 2 in Corn Inbred Lines Related to B14

The purpose of this study was to elucidate the genetic cause for the lack of express ion of the gene Ht2 against Exserohilum turcicum race 2 on certain genetic backgrounds related to the inbred line B14. Two such inbreds (A635Rp and B59), susceptible to this pathogen were crossed onto the inbred Oh43—-Ht2Ht2. The following generations: F₁, F₂, F₃, F₄, backcrosses to both parents, selfings of the backcrosses, in addition to the parent lines, were evaluated for Ht2 expression in this study. Plants of each generation were inoculated with E. turcicum race 2 and evaluated for the expression of the chlorotic-lesion resistance determined by the gene Ht2. In spite of the reported dominance of Ht2 on some genetic backgrounds, the F₁ generations studied here did not show Ht2 resistant lesions. The data presented herein suggest that B₁₄ and related inbred lines carry a dominant gene(I) that inhibits the expression of the Ht2 gene. Chi-square analyses of the reactions of 108 progeny families studied supported this hypothesis.     

豇豆胰蛋白酶抑制剂和硫氧还蛋白的融合表达和活性测定

采用融合蛋白表达技术，通过1个人工设计合成的柔性接头，将豇豆胰蛋白酶抑制剂(cowpea trypsin inhibitor,CpTI)基因与表达载体上的硫氧还蛋白(thioredoxin )基因连接，构建了融合表达载体。将表达载体转入大肠杆菌(Escherichia coli )GI724，通过色氨酸诱导获得高效表达，产物以可溶状态存在。活性测定显示，融合蛋白具有抑制胰蛋白酶的生物学活性。以大豆胰蛋白酶抑制剂为参照，将融合蛋白热处理后进行活性测定，发现它具有较好的热稳定性。将肠激酶(enterokinase)与融合蛋白在37 ℃作用16 h，可获得切掉thioredoxin的CpTI蛋白。活性测定显示，切除thioredoxin后CpTI的胰蛋白酶抑制活性比同样处理条件下的thioredoxin-CpTI明显下降，比活力仅为原来的80%，说明融合蛋白具有更高的稳定性。研究结果为thioredoxin-CpTI作为一种生物农药的应用奠定了基础。