培养条件对蛋白核小球藻生长及油脂合成的影响

以富油蛋白核小球藻为出发藻株,研究自养、异养和混养培养模式对小球藻生物量和油脂含量的影响,以及异养发酵培养基葡萄糖质量浓度、氮源种类及质量浓度对小球藻生长的影响。结果表明,与自养和混养培养模式相比,采用异养发酵方式培养蛋白核小球藻可获得最大的生物量和油脂含量。通过气相色谱法测得异养蛋白核小球藻油主要脂肪酸为棕榈酸(36.07%)、油酸(34.26%)、亚油酸(20.17%)和亚麻酸(6.12%)。经单因素试验优化得到最适蛋白核小球藻生长异养发酵培养基的葡萄糖质量浓度为60 g/L,最适的氮源为酵母粉,质量浓度为4 g/L,在此条件下经192 h发酵,蛋白核小球藻生物量可达12.43 g/L,油脂产量为5.45 g/L。研究结果表明,异养发酵培养获得的蛋白核小球藻油是一种潜在且可再生的新油源。     

Chlorella viruses as a source of novel enzymes

A special advantage has been conferred upon Chlorella cells as tools in biotechnology when viruses (Phycodnaviridae) infecting Chlorella cells were discovered and isolated. The viruses are large icosahedral particles (150-200 nm in diameter), containing a giant, 330-380 kbp long, linear dsDNA genome. Recently, the nucleotide sequence of the 330,740-bp genome of PBCV-1, the prototype virus of Phycodnaviridae, was determined, and up to 702 open reading frames (ORFs) were identified along the genome. The possible genes present include those encoding a variety of enzymes involved in the modification of DNA, RNA, protein and polysaccharides as well as those involved in the metabolism of sugars, amino acids, lipids, nucleotides and nucleosides. Many of these genes are actually expressed during viral infection, with functional enzymes detected in the host cytoplasm or incorporated into the virion. The successful utilization of these viral enzymes as various DNA restriction and modification enzymes (Cvi enzymes) that are now commercially available is well documented. Also noteworthy are virion-associated chitinase and chitosanase activities that have potentially important applications in the recycling of natural resources. The virions of Chlorella viruses contain more than 50 different structural proteins, ranging in size from 10 to 200 kDa. Some of these proteins may be replaced with useful foreign proteins using recombinant DNA technology. The proteins of interest can be recovered easily from the viral particles, and collected by centrifugation after complete lysis of the host Chlorella cells.     

亚硒酸钠对蛋白核小球藻生长及生物转化的影响

本文研究了亚硒酸钠对海水培养蛋白核小球藻的生长和有机硒转化能力的影响,以及富硒蛋白核小球藻中的主要硒形态。通过分批等量添加2μg/mL至50μg/mL的亚硒酸钠,确定最佳富硒培养浓度,并采用高效液相色谱法检测藻体中亚硒酸钠、硒代半胱氨酸和硒代蛋氨酸三种主要硒形态的含量。结果表明:亚硒酸钠浓度不宜过高,否则蛋白核小球藻的生长受到抑制。硒浓度为2μg/mL时小球藻的生物量较高,有机硒含量达到301.40μg/g,占总硒含量的83.24%;高效液相色谱分析表明在该培养条件下富硒小球藻中有机硒的主要形态为硒代半胱氨酸和硒代蛋氨酸;在测定的三种主要硒形态中未转化的亚硒酸钠仅占23.02%。上述结果说明在海水培养下,小球藻富硒培养的适宜外加硒浓度为2μg/mL,此条件下长势良好,无机硒得到有效转化,有机硒含量较高,可达到富硒要求。     

Lipid accumulation and growth characteristics of Chlorella zofingiensis under different nitrate and phosphate concentrations

To evaluate the potential of the green agla Chlorella zofingiensis as a feedstock for biodiesel production, the effects of nitrogen and phosphate on lipid accumulation and growth of C.?zofingiensis were studied. The maximum specific growth rate (μ(max)) reached 2.15?day(-1) when the concentration of NaNO(3) and K(2)HPO(4)·3H(2)O was 1.0?g?L(-1) and 0.01?g?L(-1), respectively. The lipid contents of C.?zofingiensis grown in media deficient of nitrogen (65.1%) or phosphate (44.7%) were both higher than that obtained from cells grown in full medium (33.5%). The highest lipid productivity (87.1?mg?L(-1)?day(-1)) was also obtained from cells grown in nitrogen deficient media, indicating nitrogen deficiency was more effective than phosphate deficiency for inducing lipid accumulation in C.?zofingiensis. In addition, the feasibility of cultivating the alga in 60?L flat plate photobioreactors and 10?L bottles outdoors for biodiesel was also tested. It was found that C.?zofingiensis could adapt to fluctuating temperatures and irradiance of outdoors and the highest μ(max) and lipid productivity could reach 0.362?day(-1) and 26.6?mg?L(-1) day(-)(1) outdoors, respectively. The lipid production potential of C.?zofingiensis is projected to be 31.1?kg?ha(-1)?day(-1) in outdoor culture. These results suggested that C.?zofingiensis is a promising organism for feedstock production of biofuel and can be used in scaled up culture outdoors.     

Untangling the molecular mechanisms and functions of nitrate to improve nitrogen use efficiency

A huge amount of nitrogenous fertilizer is used to increase crop production. This leads to an increase in the cost of production, and to human and environmental problems. It is therefore necessary to improve nitrogen use efficiency (NUE) and to design agronomic, biotechnological and breeding strategies for better fertilizer use. Nitrogen use efficiency relies primarily on how plants extract, uptake, transport, assimilate, and remobilize nitrogen. Many plants use nitrate as a preferred nitrogen source. It acts as a signaling molecule in the various important physiological processes required for growth and development. As nitrate is the main source of nitrogen in the soil, root nitrate transporters are important subjects for study. The latest reports have also discussed how nitrate transporter and assimilation genes can be used as molecular tools to improve NUE in crops. The purpose of this review is to describe the mechanisms and functions of nitrate as a specific factor that can be addressed to increase NUE. Improving factors such as nitrate uptake, transport, assimilation, and remobilization through activation by signaling, sensing, and regulatory processes will improve plant growth and NUE. © 2019 Society of Chemical Industry     

优化营养方式强化蛋白核小球藻生物量及蛋白质和叶绿素生产

本文通过优化营养方式和培养条件,提高了蛋白核小球藻生物量及细胞内蛋白质和叶绿素含量。结果表明,葡萄糖浓度能显著影响蛋白核小球藻混养和异养生长的生物量浓度(p0.05),但不同硝酸钠浓度影响不显著。在葡萄糖浓度50 g/L、硝酸钠浓度3.75 g/L条件下,小球藻异养培养4 d,即可达到最高生物量浓度为21.31 g/L,显著高于盐度15‰条件下混养所得最高生物量浓度(14.32 g/L)(p0.05)。当葡萄糖耗尽,将小球藻细胞经含氮且不含糖的培养基适当稀释后进行光诱导培养,并优化硝酸钠浓度。发现在高异养细胞密度(11.09 g/L)下,采用硝酸钠浓度(3.75 g/L)和低光强(6,411±532 Lux)诱导培养藻细胞48 h,即可获得高蛋白质含量(54.10%)和高叶绿素含量(3.14%)的小球藻,相对于光诱导培养前,分别提高了207.74%和342.25%,蛋白质和叶绿素产率也显著高于单独混养或异养培养的最高值,分别达到1.32 g/(L·d)和0.09 g/(L·d)。     

A seaweed residue unsuitable as a major source of energy or nitrogen for growing Pigs

When fed at the level of 50% of the diet, the dried residue of Ascophyllum nodosum, remaining after alginate extraction, was found to cause diarrhoea in pigs within 7 days. Digestibility coefficients for a diet containing 50% seaweed residue, 20% barley, 19% wheat and 10% fish meal were 0.47 for energy and 0.52 for nitrogen. By difference calculation the digestible energy and digestible crude protein values of the seaweed residue were determined to be 2.2 MJ and —30 g/kg dry matter.     

The use of proline as a nitrogen source causes hypersensitivity to, and allows more economical use of 5FOA in Saccharomyces cerevisiae.

The use of proline as a nitrogen source causes hypersensitivity to 5-fluoro-orotic acid (5FOA) and allows up to 40-fold less of this drug to be used to select for the loss of URA3 function in Saccharomyces cerevisiae. 5FOA hypersensitivity is presumably due to the absence of nitrogen catabolite repression when proline is substituted for (NH4)2SO4 as a nitrogen source. There are two constraints to the use of the proline-5FOA combination: (1) S288c genetic background strains are hypersensitive to 5FOA when grown in proline as a nitrogen source but at least one other genetic background is resistant to low levels of 5FOA under these conditions. (2) The addition of some nutritional supplements confers phenotypic resistance to the 5FOA-proline combination.     

缺氮胁迫下生长素对普通小球藻生长及油脂积累的影响北大核心CSCD

以普通小球藻为研究对象,在氮缺乏条件下分别添加天然生长素吲哚乙酸(IAA)、人工合成生长素萘乙酸(NAA)及2,4-二氯苯氧乙酸(2,4-D),研究了不同生长素添加量下普通小球藻的生长特征、油脂积累及藻油脂肪酸组成。结果表明:最佳的生长素添加量分别为IAA 1.0 mg/L、NAA 2.5 mg/L、2,4-D 1.0 mg/L;在IAA、NAA、2,4-D最佳添加量下,普通小球藻最高生物量分别为158、159、164 mg/L,是单一缺氮(ND)培养下生物量的1.27、1.28、1.32倍,与整个培养周期前8 d正常培养(BG11)下的微藻生物量相比无显著性差异,生长素有效抵消了缺氮胁迫对生物量的负面影响;在IAA、NAA、2,4-D最佳添加量下,普通小球藻最大总脂含量分别是BG11培养条件下的1.72、1.64、1.74倍,最大总脂产量分别为61.38、58.83、64.22 mg/L,是BG11组的1.49、1.43、1.56倍,而ND组最大总脂产量低于BG11组;在最佳生长素添加量下,普通小球藻细胞内最大甘油三酯含量分别为27.97%、25.66%、33.21%,最大甘油三酯产量分别为44.19、40.80、54.46 mg/L,均明显高于BG11组;C16~C20脂肪酸成分分别占总脂肪酸的71.99%、73.75%、84.81%。研究表明,缺氮胁迫下3种生长素对普通小球藻生长及油脂合成具有明显促进作用,其中2,4-D效果最佳。     

Asparagine as a nitrogen source for improving the secretion of mouse alpha-amylase in Saccharomyces cerevisiae protease A-deficient strains

A modified chemically defined medium was achieved by using asparagine as a nitrogen source to increase the production of secreted mouse alpha-amylase in several Saccharomyces cerevisiae protease A-deficient (pep4) strains. The specific productivity (quantity) and the 53 kDa non-glycosylated active form (quality) of mouse salivary alpha-amylase in liquid medium containing asparagine was remarkably improved compared to media containing other nitrogen sources, including ammonium sulphate, glutamic acid, arginine, casamino acids, yeast extract and peptone. Similar improvement was also observed on starch solid agar regarding the clarity and size of the halo zone formed by alpha-amylase activity. Compared with ammonium sulphate, advantages of using asparagine as the nitrogen source in liquid or solid medium included increasing the cell mass of test strains, recovering the viability of protease-deficient strains to levels similar to the wild-type strain, and increasing the copy number of the mouse alpha-amylase expression vector in test strains. In turn, these advantages apparently contributed to the increase of secretion of mouse alpha-amylase in several test strains and especially in the protease A-deficient strains. In addition to demonstrating the use of modified chemically defined medium to improve the quality and quantity of secreted mouse alpha-amylase, this study also provides a new strategy to improve the secretion of heterologous proteins in protease A deficient strains.     

Low nitrogen losses with a new source of nitrogen for cultivation of conifer seedlings

Losses of nitrogen (N) when cultivating plants may cause a number of adverse environmental effects. N losses from conifer nurseries, for instance, may have a considerable impact on the local environment, and studies indicate that the bulk of added N is not recovered in the cultivated plants. This study was conducted to obtain insight into the causes of the low recovery and to test an alternative N fertilizer. Hence, growth of the economically important Scots pine (Pinus sylvestris (L).) seedlings and the recovery of different forms of added nitrogen (N) were investigated in a greenhouse experiment. Containerized seedlings were grown in peat for one summer, with two different N fertilizers, one organic (arginine) and one inorganic (a commercial fertilizer (CF) containing a mixture of ammonium and nitrate) each at an N concentration of 3 mM. At the end of the growth period, some seedlings were labeled with solutions containing either U-[13C6], [15N4]-arginine, (15NH4)2SO4, or K15NO3 supplied to the growth substrate. Labeled seedlings were harvested 1 h, 5 days, and 19 days after tracer addition, and the recovery of each added nitrogen source in both the seedlings and the growth substrate was measured. The retention of the three N forms during discharge of solutions from the growth substrate, peat, was tested in a separate experiment. Arginine-fed seedlings grew better and had higher needle N concentrations than the CF-fed seedlings. Isotopic data showed that the arginine treatment gave significantly higher N recoveries (80%) compared to the CF treatment (50%). The low recovery of N in the CF treatment was largely due to very low recovery (30%) of NO3- -N. The retention of the different N forms during discharge of solutions from the growth substrate was highest for arginine, somewhat lower for NH44+, and very low for NO3-. The high rate of seedling growth and the small nitrogen losses observed when using arginine suggest that this amino acid may be an efficient and environmentally favorable N source for cultivating conifer seedlings.     

Primary consumer stable nitrogen isotopes as indicators of nutrient source

Non-point source loading of nitrogen and phosphorus is a primary cause of eutrophication of inland waters, although the diffuse and variable nature of nutrient inputs makes it difficult to trace and identify nutrient pathways. Stable nitrogen isotope values (delta15N) in aquatic biota are thought to reflect anthropogenic nutrient inputs, and they may be a promising tool fortracing nutrient sources in watersheds. We measured delta15N of aquatic consumers from a suite of 27 Danish lakes spanning a range of trophic states (oligotrophic to eutrophic) and land uses (forest, urban, agriculture). Primary consumer delta15N values (PCdelta15N) varied more than 14% among lakes. Models of PCdelta15N were developed from limnological, nitrogen loading, and nitrogen source variables using an information-theoretic approach (Akaike's Information Criteria, AIC). Models based on land use/ land cover performed best, indicating that elevated delta15N is not only associated with high nitrogen loading, but is also reflective of nitrogen source. Urban and agricultural land use in the watershed, and particularly within the riparian buffer areas, was quantitatively linked to elevated biotic delta15N.     

光照强度和氮源匮乏培养对小球藻细胞生长和细胞组成的影响

本研究考察了不同光照强度和氮源匮乏培养对小球藻细胞生长及其细胞组成的影响。结果表明,随光照强度的增强,小球藻生物量产率和碳水化合物含量可显著提高,而蛋白质含量和油脂含量则呈现缓慢下降趋势;在光照强度为750μmol·m^-2·s^-1时,小球藻生物量产率、蛋白质产率和碳水化合物产率达最大值,分别为935.6、509.8、256.0 mg·L^-1·d^-1;而光照强度为450μmol·m^-2·s^-1时,小球藻油脂产率达最大值,为105.0 mg·L^-1·d^-1。通过氮源匮乏培养来促进小球藻碳水化合物和油脂含量的积累,发现当氮源匮乏培养时间分别为2 d和4 d时,小球藻碳水化合物产率和油脂产率可进一步提高至471.1 mg·L^-1·d^-1和150.6 mg·L^-1·d^-1。      

氮、磷浓度对小球藻生长及油脂积累的影响

含油微藻是非常有前景的生产生物柴油的材料,其生物量和油脂含量是决定其经济可行性的重要指标。培养基组成对含油微藻的生长与油脂积累具有重要影响。实验以采自青岛海域的一株海水小球藻为研究对象,在摇瓶中考察培养基氮源、磷源浓度对小球藻生长及其油脂积累的影响。结果表明:在一定范围内,增加氮源浓度能提高小球藻生长速率,但磷源对小球藻生长影响不明显;随着氮、磷源浓度的降低,小球藻总脂和中性脂含量均逐渐增大,氮源起的作用更加显著,在0.220mmol/L的低氮胁迫下获得了最高的细胞总脂含量,总脂占细胞干重的33.5%±0.3%。氮源浓度为0.441mmol/L时,小球藻的油脂产率最高,为(10.58±0.29)mg·L-1·d-1。培养基中低磷浓度有利于小球藻的油脂积累,但差别不明显。     

以废弃毕赤酵母为高效氮源强化丁酸发酵生产

为有效利用固态废弃毕赤酵母,高效发酵生产丁酸,建立了以80 g/L玉米淀粉为基础培养基、以废弃毕赤酵母处理液为高效有机氮源,连续补加葡萄糖的丁酸发酵工艺。7 L厌氧发酵罐下,发酵20h,先以1∶4的比例添加250~400 g/L规格的废弃酵母处理液替代昂贵的发酵用复合培养基,再连续补加葡萄糖浓缩液。最终的丁酸质量浓度为45 g/L、得率为40%,丁酸占总酸的比率(butyric acid ratio over total organic acids,B/TA)≥0.90的较高水平。该工艺可以有效地利用来自于废弃酵母处理液中的氨基酸,提高细胞生长速度和浓度,增强丁酸合成所依赖的NADH再生速度,间接改善了丁酸发酵性能。与此同时,实现了废弃生物质的有效利用和资源化。     

On-site nitrate removal of groundwater by an immobilized psychrophilic denitrifier using soluble starch as a carbon source

On-site denitrification of nitrate-contaminated groundwater was conducted at 15 degrees C using a facultative psychrophilic denitrifier (strain 47) immobilized on macro-porous cellulose carriers and utilizing soluble starch as a non-toxic carbon source. A C/N ratio of 2.5 to 3.0 and a P/N ratio of 0.05 to 0.10 were found to allow complete denitrification of the groundwater used in this study. Under these conditions, the long-term performance of the system (4 months) was examined by decreasing the HRT (hydraulic retention time) from 4 h to 0.25 h. The process was stable and 95 to 100% of the influent nitrogen (NO3-N ranging from 13.0 to 16.5 mgl(-1) was removed until an HRT of 0.75 h was reached. The maximum NO3-N removal rate was 0.46 kg-Nm(-3)d(-1) at an HRT of 0.75 h. Nitrogen removal efficiency of 99.5% at an HRT of 1 h was obtained with a C/N ratio 2.58, corresponding to 4.3 g of soluble starch per 1 g of NO3-N.     

Problems which limit the use of rapeseed meal as a protein source in poultry diets

The occurrence of a ‘fishy’ or ‘crabby’ taint in eggs and liver damage culminating in massive haemorrhage has greatly restricted the commercial utilisation of rapeseed meal as a relatively cheap protein supplement for laying hens in the UK. Recent work on these problems is reviewed. It has elucidated the cause of the taint and identified the rapeseed constituents that are involved. A genetic defect impedes the synthesis of trimethylamine oxidase in susceptible hens and goitrin and tannins inhibit the enzyme. This biochemical lesion severely impairs the metabolism of trimethylamine which is released from sinapine and other dietary sources of choline by enteric bacteria. Consequently, excessive amounts pass into the yolk and produce the taint. Liver haemorrhage is associated with hepatocyte degeneration, abnormalities in the biliary system and the leakage of cellular enzymes into the plasma. The toxic substance(s) responsible has (have) not yet been identified.     

烘焙温度对小球藻脂肪酸、色素及乙醇提取物抗氧化活性的影响

探讨烘焙温度对小球藻脂肪酸组成、色素成分及乙醇提取物抗氧化活性的影响。结果表明:小球藻的脂肪酸以多不饱和脂肪酸为主,其含量达到细胞总脂的76.69%。其中,亚油酸(C_(18:2))含量最高,亚麻酸(C_(18:3,n3))次之。温度不高于150℃时,烘焙处理对小球藻的脂肪酸含量及组成无明显影响;但当温度达到200℃时,烘焙处理会显著降低多不饱和脂肪酸的含量。小球藻的色素成分及乙醇提取物的抗氧化活性对烘焙温度很敏感。当烘焙温度高于100℃时,小球藻的叶绿素和类胡萝卜素降解严重;当烘焙温度高于150℃时,小球藻的抗氧化活性也明显减弱。     

三聚氰胺树脂为氮源的氮掺杂碳量子点制备及光学性能

以三羟甲基三聚氰胺树脂为氮源,乙二胺、对苯二胺和1,8-二氨基萘为碳源,采用一步水热法制备了三种氮掺杂碳量子点(N-CQDs-1、N-CQDs-2和N-CQDs-3),探究了三种氮掺杂碳量子点的结构及光学性能.结果表明,N-CQDs-1、N-CQDs-2和N-CQDs-3的平均粒径分别为3.87、7.09和2.79 n...     

Spent brewery yeast sludge as a single nitrogen source for fibrinolytic enzyme production of Virgibacillus sp. SK37

The objectives of this study were to investigate the suitable food industrial byproducts and conditions affecting proteinase production of Virgibacillus sp. SK37 as well as to evaluate fibrinolytic activity of secreted proteinases. Among 5 food industrial wastes including soybean pomace, rice bran, mungbean protein, spent brewery yeast sludge (Ys), and fish sauce sludge, Ys was the best nitrogen source for proteinase production from Virgibacillus sp. SK37. The highest level of proteinase production was obtained in the medium containing 1% Ys, 2.5% NaCl, pH 7.5 at 40°C, which was about 1.7 times higher than the commercial yeast extract medium. Virgibacillus sp. SK37 proteinases completely hydrolyzed fibrinogen within 50 min with Aα-chain being the first target and followed by Bβ- and Γ-chains. Virgibacillus sp. SK37 could be a potential source of fibrinolytic enzyme that can be developed as a functional food ingredient.