Enantioselective toxicological response of the green alga Scenedesmus obliquus to isocarbophos

Chiral compounds usually behave enantioselectively in phyto‐biochemical processes. Isocarbophos (ICP) is a chiral pesticide that is widely used. To evaluate the toxicological response of ICP and its enantiomers to Scenedesmus obliquus, algal growth, total chlorophyll, total soluble protein, and the superoxide anion radicals (O₂^?‐) were investigated. The microalgae were treated with ICP and its enantiomers at 0.01–10 mg/l for 96 h. The growth of S. obliquus was stimulated at low levels of ICP and its enantiomers (0.01–1 mg/l), but all were inhibited at high concentrations (10 mg/l). The total soluble protein content and total chlorophyll content of the tested green alga S. obliquus gradually increased, depending on the growth of algal cells in the medium. Meanwhile, the content of O₂^?‐ was decreased. Interestingly, the cell number and content of the chlorophylls and protein decreased with increasing levels of concentration, whereas O₂^?‐ increased. Our results indicated that enantioselectivity was observed in the dose–response of ICP and its enantiomers in S. obliquus. The high O₂^?‐ level might lead to the death of S. obliquus. The stimulation of growth suggests a regulatory mechanism that is related to the capability of the algae to adapt to the O₂^?‐. Chirality 24:481–485, 2012. © 2012 Wiley Periodicals, Inc.     

Enantioselective toxic effects of hexaconazole enantiomers against Scenedesmus obliquus

Enantioselectivity in ecotoxicity of chiral pesticides in the aquatic environment has been a subject of growing interest. In this study, the toxicological impacts of hexaconazole enantiomers were investigated with freshwater algae Scenedesmus obliquus. After 96 h of exposure, the EC₅₀ values for rac‐hexaconazole, (+)‐hexaconazole, and (?)‐hexaconazole were 0.178, 0.355, and 0.065 mg l^?1, respectively. Therefore, the acute toxicities of hexaconazole enantiomers were enantioselective. In addition, the different toxic effects were evaluated when S. obliquus were exposed to 0.2, 0.5, and 1.0 mg l^?1 of rac‐hexaconazole, (+)‐hexaconazole, and (?)‐hexaconazole during 96 h, respectively. The chlorophyll a and chlorophyll b contents of S. obliquus treated by (?)‐hexaconazole were lower than those exposed to (+)‐hexaconazole, whereas the malondialdehyde contents of S. obliquus treated by (?)‐form were higher than those exposed to (+)‐form at higher concentrations. In general, catalase activities were significantly upregulated by exposure to (?)‐enantiomer than (+)‐enantiomer at all three concentrations. However, superoxide dismutase activities exposed to (?)‐hexaconazole were lower than that exposed to (+)‐hexaconazole at 0.2 mg l^?1 and 0.5 mg l^?1. On the basis of these data, the acute toxicity and toxic effects of hexaconazole against S. obliquus were enantioselective, and such enantiomeric differences must be taken into consideration in pesticide risk assessment. Chirality 24:610–614, 2012. © 2012 Wiley Periodicals, Inc.     

Biosynthesis of chlorophyll b in pigment mutant C-2A' of Scenedesmus obliquus

It is demonstrated that chlorophyll b does not only derive from chlorophyll a , but is also formed separately from an in vivo-occurring chlorophyllide b . The branching point for the latter synthesis is at the level of chlorophyllide, since no protochlorophyllide b was detectable. We have indications that the enzyme oxidizing chlorophyll a to chlorophyll b accepts also non-phytylated 17,18 dihydroporphyrins and is not restricted to chlorophylls. Preparations of chlorophyllide a and chlorophyll a could both be transferred with the same enzyme fraction to chlorophyllide b and chlorophyll b , respectively. Preliminary experiments show this enzyme to be membrane bound and light independent. An updated scheme for chlorophyll b biosynthesis is presented.     

斜生栅藻中虾青素的积累过程及其光合活性变化

分析了斜生栅藻（Scenedesmus obliquus）在光温（30℃,180 μmol/m2·s）胁迫条件下积累虾青素的过程,观察了该过程中细胞形态及细胞光合生理的变化。胁迫条件下,细胞在48h内生成并积累了包括海胆酮、角黄素、金盏花黄素和金盏花红素在内的多种次生类胡萝卜素,并合成了虾青素及其酯。该过程中,细胞形态由两端尖细变得不规则、膨大,原来由4、8个细胞组成的定形群体变为游离的单个细胞或2个细胞组成的群体。藻细胞光合速率在24h内先下降后上升,而后又呈现下降趋势,从34.29 μmol O2/mg Chla/h迅速下降为5.21 μmol O2/mg Chla/h;呼吸速率在前24h内升高至60.37 μmol O2/mg Chla/h,而后缓慢下降到38.40 μmol O2/mg Chla/h;光合系统Ⅱ的活性随着胁迫时间的延续而逐步下降,较初始值降低了63.9％。结果表明,斜生栅藻细胞在高光照条件下可以合成虾青素,并通过调节光合速率、呼吸速率以及光合系统Ⅱ的效率来应对胁迫。     

水芹水浸提液对斜生栅藻的化感效应研究

采用栅藻在不同浓度的水芹水浸提液中纯培养的方法,研究了水芹水浸提液对斜生栅藻细胞数、叶绿素含量及藻细胞超微结构的影响。结果显示,10 g·L^-1水芹水浸提液对斜生栅藻的生长和叶绿素含量的增加具有明显的促进效应;20 g·L^-1水芹水浸提液对藻细胞数和叶绿素含量的增加持续到第3 d,3 d后出现显著的抑制效应;高浓度（30~50 g·L^-1）水芹水浸提液对藻细胞数和叶绿素含量的明显抑制在第2 d始现,随时间延长而加剧,并具有浓度效应;40 g·L^-1水芹水浸提液处理后,斜生栅藻细胞壁断裂甚至消失,细胞中叶绿体片层肿胀甚至解体,核膜破裂,核质外渗。水芹水浸提液对斜生栅藻具有化感效应,低浓度促进生长,高浓度抑制。     

Occurrence of protochlorophyll and its phototransformation to chlorophyll in mutant C-2A' of Scenedesmus obliquus

The pigment mutant C-2A' of the unicellular green alga Scenedesmus obliquus accumulates protochlorophyllide and small amounts of protochlorophyll in darkness. Protochlororphyll was isolated and characterized by thin layer chromatography and absorption and fluorescence spectroscopy. The protochlorophyll was photoreduced by light to chlorophyll both in vitro and in vivo.     

Influence of temperature on growth of Scenedesmus obliquus in diluted olive mill wastewater as culture medium

Scenedesmus obliquus can help to reduce the environmental impact of industrial olive mill wastewater from olive oil extraction in the three‐phase system. This work examines the effect of temperature changes (288–308 K) on algal growth, culture medium, and biochemical composition of S. obliquus. The maximum specific growth rate of 0.024 h^?1 occurred at an optimal temperature of 302.7 K. The apparent activation energies of cell growth and cell death were determined as 61.8 and 142.8 kJ/mol, respectively. At the end of culture the percentages of pigments, proteins, and carbohydrates were greater at the two ends of the temperature range studied, as it was also observed for the maximum elimination of biochemical oxygen demand (BOD₅). The mono‐ and polyunsaturated fatty acid content of the biomass was greater at the lowest temperature used (288 K).     

Evidence for the Presence of Chlorophyllide b in the Green Alga Scenedesmus obliquus in vivo

Chlorophyllide b could be extracted from the wild type of Scenedesmus obliquus and its pigment mutant C-2A'. Its identity was proved by absorption and fluorescence spectroscopy and by a positive hydroxylamine test. Chlorophyllide b could be transformed into pheophorbide b and methylpheophorbide b. The formation of chlorophyllide b from chlorophyll b by dephytylation with chlorophyllase could be ruled out. The stimulation of chlorophyllide b biosynthesis with o-phenanthroline, as described in the literature, could not be confirmed under physiological conditions.     

The effect of arsenate and arsenite on photosynthesis in Scenedesmus obliquus. A Potentiometric study in a closed CO2-system

Abstract

A Potentiometric titration method was used to study the adverse effect of arsenate (As(V)) and arsenite (As(III)) on inorganic carbon uptake in suspensions of the green alga Scenedesmus obliquus. The measurements were performed in a closed CO₂-system with diluted synthetic seawater (1‰ salinity) as ionic medium. Usually, the algal chlorophyll concentration was 0.4 mg dm^?3, while the arsenate- and arsenite-concentrations were varied within the limits 0.1 to 200 μmol dm^?3. In some experiments arsenate toxicity was studied in the presence of 1 to 100 μmol dm^?3 of phosphate (P(V)).

With concentrations of arsenate or arsenite less than 0.1 μmol dm^?3 no toxic effects were observed. However, at As-concentrations of 200 μmol dm^?3, the algal carbon uptake was reduced by 41% with arsenate and 29% with arsenite, i.e., arsenate is more toxic to Scenedesmus obliquus than arsenite. The toxicity of arsenate was negligible in the presence of a ten fold excess of phosphate. This is probably due to chemical similarities between arsenate and phosphate causing competition between the ions for the binding sites.

The importance of taking the speciation as well as the buffer capacity of the algal system into account, when calculating the carbon uptake, is also discussed.     

Effect of acetate on growth and ammonium uptake in the microalga Scenedesmus obliquus

The effect of acetate on growth and rate of ammonium uptake in Scenedesmus obliquus (UTEX 78) was investigated under light-limiting conditions. Addition of acetate to autotrophic cells with a growth constant of 0.71 day⁻¹ resulted in an increase in the growth rate (mixotrophy, k = 1.3 day⁻¹), and in the presence of acetate, growth occurred in the dark (heterophy, k = 0.44 day⁻¹). The rate of ammonium uptake in autotrophy (17.8 amol cell⁻¹ min⁻¹) was similar to that in heterotrophy (17.4 amol cell⁻¹ min⁻¹) but was 3.7 times lower than that in mixotrophy (65.9 amol cell⁻¹ min⁻¹). In general, mixotrophic cells showed optimum ammonium uptake at the acetate concentration at which they were grown. In autotrophy, uptake of ammonium leveled off at about 12.5 μ M while no saturation was observed in mixotrophic cells. An increase in the rate of uptake of ammonium was observed in autotrophic cells within 1 h after the addition of acetate. The activity of isocitrate lyase (EC 4.1.3.1), a key enzyme for the regulation of the glyoxylate cycle responsible for acetate catabolism, showed a 3.9-fold increase in activity after 24 h in the dark in the presence of acetate. The level of isocitrate lyase activity in cells grown for 24 h in the dark in the presence of 0–20 m M acetate also increased as a function of acetate concentration.     

Enantioselective Toxic Effects and Degradation of Myclobutanil Enantiomers in Scenedesmus obliquus

Research on the enantioselective environmental behavior of chiral pesticides has been a hot spot of environmental chemistry recently. In this study, the acute toxicity of myclobutanil enantiomers was investigated with the aquatic algae Scendesmus obliquus. After exposure for 96 h, the EC₅₀ values for (?)‐myclobutanil, rac‐myclobutanil and (+)‐myclobutanil were 3.951, 2.760, and 2.128 mg/L, respectively. The photosynthetic pigment (chlorophyll a, chlorophyll b, and carotenoids) and antioxidant enzyme activities catalase (CAT) were determined to evaluate the different toxic effects when S. obliquus were exposed to 1.5, 5 and 15 mg/L of rac‐myclobutanil, (?)‐myclobutanil, and (+)‐myclobutanil for 96 h, respectively. In addition, the degradation of myclobutanil enantiomers in S. obliquus was also studied. Myclobutanil in the medium inoculated with algae degraded faster than in the uninoculated medium. The degradation of (?)‐myclobutanil was faster than that of (+)‐myclobutanil at a concentration of 3 mg/L. On the basis of these data, the acute toxicity and toxic effects of myclobutanil against S. obliquus were concluded to be enantioselective, and such enantiomeric differences should be taken into consideration in pesticide risk assessment. Chirality 25:858–864, 2013. © 2013 Wiley Periodicals, Inc.     

Accumulation of uranium at low concentration by the green alga Scenedesmus obliquus 34

Accumulation of UO ₂ ^{2 +} by Scenedesmus obliquus 34 was rapid and energy-independent and the biosorption of UO ₂ ^{2 +} could be described by the Freundlich adsorption isotherm below the maximum adsorption capacity (75 mg g-¹ dry wt). The optimum pH for uranium uptake was between 5.0_8.5.0.1_2.0 M NaCl enhanced uranyl, while Cu²⁺, Ni²⁺, Zn²⁺, Cd²⁺ and Mn²⁺ competed slightly with uranyl. Pretreatment had an unexpected effect on biosorption. After being killed by 0.1 M HCl, S. Obliquus 34 showed 45% of the uptake capacity of the control in which fresh cells were suspended directly in uranyl solution, while the pretreatment of cells by 0.1 M NaOH, 2.0 M NaCl, ethanol or heating decreased uptake slightly. Fresh S. obliquus 34 at 1.2_2.4 mg dry wt mL-1 was able to decrease U from 5.0 to 0.05 mg L-1 after 4_6 equilibrium stages with batch adsorption. Deposited U could be desorbed by pH 4.0 buffer. It is suggested that U was captured by effective groups or by capillary action in the cell wall in the form of [UO2OH]+. This revised version was published online in August 2006 with corrections to the Cover Date.     

溴化1-己基-3-甲基咪唑对斜生栅藻谷胱甘肽及其代谢酶的影响

研究了浓度为0、1、5、10、15、20 mg/L的新兴离子液体溴化1-己基-3-甲基咪唑([C6mim]Br)在24h、48h、72h和96h对斜生栅藻还原型谷胱甘肽（GSH）及其代谢酶-谷胱甘肽过氧化物酶（GPX）、谷胱甘肽转硫酶（GST）和谷胱甘肽还原酶（GR）的影响。结果表明：GSH含量在24h、48h和72h时,在最低处理浓度下不变,其他处理浓度下随胁迫浓度增加而降低,96h时则与对照无差异或较小;GPX和GST的活性在72h之前明显升高（最高浓度组的GST活性有波动）,96h时均降低至对照水平;GR活性在24h时,[C6mim]Br=1 mg/L时升高,之后降低,在48h增高至对照水平,72h时,[C6mim]Br≥10 mg/L的处理组高于对照水平,96h时,除最低处理组外,均降至对照水平以下。GR是GSH系统中的限速酶,GST则是该系统中活性和灵敏性最高的酶,可作为[C6mim]Br胁迫时的敏感的生物标志物。1 mg/L的[C6mim]Br可引起藻细胞的氧化胁迫,具有环境毒性。     

壶状臂尾轮虫(Brachionus urceus)培养滤液对斜生栅藻(Scenedesmus obliquus)形态和多聚糖含量的影响

分离自太湖的壶状臂尾轮虫(Brachionus urceus (Linnaeus))按1000 ind·L-1的密度置于斜生栅藻(Scenedesmus obliquus)中培养24h后,用孔径0.10 靘的微孔滤膜抽滤,得到壶状臂尾轮虫培养滤液.将壶状臂尾轮虫培养滤液以20%的比例添加到纯培养的斜生栅藻中,进行为期7d的试验.结果表明壶状臂尾轮虫培养滤液能显著地影响斜生栅藻的形态,处理组中诱发性多细胞群体增加,导致处理组中斜生栅藻种群平均每个个体的细胞数量显著高于对照组.斜生栅藻在壶状臂尾轮虫诱发下形成群聚体可以解释为一种诱发性的反牧食防御,同时也进一步证实了栅藻对浮游动物牧食的响应是一种广泛存在的现象.壶状臂尾轮虫培养滤液处理后的斜生栅藻在生长率上与对照组没有显著性差异,说明诱发性防御群聚体形成的代价没有体现在对生长的影响上.在壶状臂尾轮虫培养滤液诱发下,较多栅藻细胞聚积在一起形成群聚体后,栅藻溶解性胞外多聚糖和固着性胞外多聚糖均有显著提高,而胞内多聚糖略有下降.但从总多聚糖含量看,处理组明显高于对照组,这也说明轮虫培养滤液刺激了栅藻多聚糖合成的额外增加,且被分泌到胞外,充当细胞之间的粘合物,促使细胞容易聚合形成群聚体,有效防御象轮虫这些小型浮游动物的进一步牧食,保持种群得以延续.     

Monovinyl and divinyl protochlorophyll in different stages of esterification isolated from mutant C-2A'of the unicellular green alga Scenedesmus obliquus

The mutant C-2A'of the unicellular green alga Scenedesmus obliquus accumulates the chlorophyll-precursors protocbloropbyllide and the already eslerified protochlorophyll when grown heterotrophically in the dark. Two derivatives of protochlorophyll. monovinyl protochlorophyll (MV-PChl) and divinyl protochlorophyll (DV-PChl), were isolated from dark-grown cells of mutani C-2A'and characterized by absorption and fluorescence spectroscopy. Their molecular masses were determined by plasma desorption mass spectrometry. Both MV- and DV-PChl were mainly esterified with geranylgeraniol. However, some esterification with the more saturated alcohols dihydrogeranylgeraniol. tetrahydrogeranylgeraniol and phytol could also be detected.     

Biotransformation of 1-naphthalenesulfonic acid by the green alga Scenedesmus obliquus

Under sulfate limitation, axenic batch cultures of the green alga Scenedesmus obliquus metabolized 1-naphthalenesulfonic acid and partially used the sulfonate as a source of sulfur. The main metabolite, 1-hydroxy-2-naphthalenesulfonic acid, which was not metabolized further in the algal culture, was formed by hydroxylation of the substrate in position 1 and by migration of the sulfonic acid group to position 2 of the naphthalene ring (NIH shift). A smaller amount of 1-naphthalenesulfonic acid was desulfonated. The resulting 1-naphthol was mostly transformed into 1-naphthyl β-d-glucopyranoside. Received: 27 March 1996 / Revision received: 18 October 1996 / Accepted: 30 October 1996     

Photosynthetic Capacity and Quantum Requirement of Three Secondary Mutants of Scenedesmus obliquus with Deletions in Carotenoid Biosynthesis*

Photosynthetic capacity and quantum requirement of photosynthesis were evaluated in three secondary mutants of Scenedesmus obliquus possessing specific blocks in carotenoid biosynthesis. These were derived from the mutant, C-2 A', a strain which develops chlorophyll only in the light. All strains were capable of a normal and rapid production of chlorophyll and the development of a functional photosynthetic apparatus when exposed to light. LHC-II levels in the mutants lacking the β,ε-carotenoids (strains C-2 A'-34,1 and C-2 A'-67, 3b) were considerably reduced in fully greened cells but normal in the β,β-epoxycarotenoid deficient-only strain (C-2 A'-67,1) (Bishop, 1996). The maximum photosynthetic capacity and the quantum efficiency (at λ=682nm) of photosynthesis of cells of C-2A'-34,1 and C-2 A'-67,1 showed no significant changes when compared to the original parent strain, C-2 A'. However, the quantum requirement of the strain lacking both types of carotenoids, C-2 A'-67,3b, was slightly increased. It is of particular interest that both strains lacking the β,ε-carotenoids also show a preferential loss of the oligomeric form of the LHC and its associated chlorophylls. The potential requirement of the different carotenoids in maintenance of photosynthetic efficiency in Scenedesmus is discussed.     

角突网纹溞培养滤液对铜绿微囊藻和栅藻形态及生长率的影响

在藻类中,随着环境条件的变化而产生相应的单细胞和群体之间的形态转化是一种广泛存在的现象,称为表型可塑性。例如,栅藻属成员就是这些藻类的代表,在野外具有典型的群体特征,但在室内纯培养条件下栅藻常较少形成群体而是保持以单细胞形态为主。     

Temperature dependent reduction of protochlorophyllide in darkness followed by the assembly of active photosystems in pigment mutant C-2A' of Scenedesmus obliquus

In the wild type of Scenedesmus obliquus strain D3 grown heterotrophically, the chlorophyll biosynthesis and thus the reduction of protochlorophyllide to chlorophyllide takes place in darkness. However, in pigment mutant C-2A' of Scenedesmus obliquus only traces of protochlorophyllide are reduced under optimal growth conditions in darkness. By lowering the growth temperature from 33° to 15–25°C, protochlorophyllide can be reduced in darkness. At 20°C this process is about 10 times more active than at 33°C, but reaches only about 13% of the light-dependent chlorophyll biosynthesis. The chlorophylls synthesized at the lower temperatures are inserted into the pigment-protein complexes and photosystem I as well as photosys-tem II capacities are developed. The rate of light-independent protochlorophyllide reduction at lower temperatures is not limited by the enzyme PChlide-oxidoreductase itself, but rather by its substrate, being in turn limited by the amount of 5-amino levulinic acid (ALA) available.     

Stereoselective bioaccumulation,transformation, and toxicity of triadimefon in Scenedesmus obliquus

In this study the stereoselective bioaccumulation and transformation of triadimefon and the toxicity of triadimefon and its metabolite triadimenol to the green algae Scenedesmus obliquus were studied. In growth inhibition experiments, triadimenol was more toxic than triadimefon, and (1S,2R)‐triadimenol, which has the largest fungicidal activity, presented the highest toxicity to the algae. In bioaccumulation experiments, triadimefon was rapidly taken up by algae cells, and the decrease in the concentration of triadimefon was accompanied by an increase in triadimenol. The transformation of S‐(+)‐ triadimefon was faster than that of the R‐(−)‐enantiomer, resulting in four triadimenol stereoisomers at different forming rates: B2 (1S, 2S) > B1 (1R, 2R) > A2 (1S, 2R) > A1 (1R, 2S). Thus, it is necessary to explore the enantioselective toxicology and ecological fate of these chiral pesticides in an environmental risk assessment. Also, their metabolites should be paid specific attention to since they may pose higher ecological risks.