C. Van Baalen

Water-soluble components of four fuel oils: Chemical characterization and effects on growth of microalgae

Approximately 50% of the compounds in the water solubles from 4 fuel oils have been identified via gas chromatography and mass spectrometry. In addition to the well-described types of compounds (naphthalenes, benzenes) expected in water-soluble extracts we have found phenols, anilines, and indoles. Of these classes of compounds methyl, dimethyl, and trimethyl derivatives are present in relatively high concentrations. The water solubles from the 4 fuel oils showed considerably different inhibitory effects to growth of 6 microalgae, 2 blue-greens, 2 greens, and 2 diatoms. Two of the fuel-oil extracts, Baytown and Montana, were lethal to blue-green algae. This was in part traceable to their content of p-toluidine which was found to be toxic to Agmenellum quadruplicatum, Strain PR-6, 1 μg in the algal lawn-pad assay and 100 μg/l in liquid culture. The water-soluble fraction from New Jersey fuel oil was lethal to the 2 green algae, with lesser effects on the 2 blue-greens. The 2 estuarine diatoms used as test organisms were not greatly inhibited by Baytown, Montana, or New Jersey fuel-oil water-soluble extracts. However, earlier work with an American Petroleum Institute fuel oil and the diatom Thallassiosira pseudonana (3H) showed that 3H was a very sensitive organism. Water solubles from the Baton Rouge fuel oil were almost without effect on the growth of all 6 microalgae. On the basis of the work herein and earlier work, a very cautious viewpoint is advisable in generalizing on the toxicity or lack thereof of a given fuel oil on the growth of different kinds of microalgae. On the other hand, with water solubles from toxic fuel oils such as Baytown or New Jersey the data clearly suggest that their potential for environmental damage is high, either through selective or enrichment effects on natural populations or through a lowering of total primary production.

The effects of a no. 2 fuel oil and two crude oils on the growth and photosynthesis of microalgae

Seawater, when equilibrated with a sample of No. 2 fuel oil, becomes toxic in varying degrees to growth of representative types of microalgae, two blue-greens, a diatom, two greens, and a dinoflagellate. For a sensitive organism such as Thalassiosira pseudonana, Strain 3H, 5 ml of seawater equilibrated with fuel oil (containing 15 mg/l of organics) in 20 ml of growth medium is lethal, or roughly in the range of 40 to 400 ppb if the toxic material(s) constitute 1 to 10% water also immediately stops photosynthesis in organism 3H. For other microalgae tested e.g. 580 (a green alga) and PR-6 (a blue-green alga), similar effects on growth and photosynthesis were found, but required higher concentrations of the oil-equilibrated seawater. Water solubles from Kuwait or Southern Louisiana crude oils (when the straight crude oil was equilibrated 1:8 with seawater) were not toxic; however, specific fractions obtained by distillation did show some water-soluble toxicity. Growth experimetns in open or closed growth systems revealed that most organisms were inhibited by varying amounts of these two crude oils when in direct contact with them. Organism 580 would not grow above 5 μl of Southern Louisiana/25 ml of medium, or 10 μl of Kuwait/25 ml of medium (oil in direct contact with algae). With both the seawater equilibrated with fuel oil and the crude oils, the toxic activity is mainly localized in medium and higher boiling fractions derived from distillation cuts from these materials.

Growth responses of blue-green algae to sodium chloride concentration

SummaryGeneral characteristics of blue-green algal halotolerance were studied by growth experiments and selected analyses. Variation in NaCl concentration was used to mimic salinity. Marine isolates were more halotolerant (8–10% NaCl) than non-marine isolates (2% NaCl). The Na+ requirement for growth was saturated at 1 mg NaCl/l for non-marine isolates and 100mg NaCl/l for marine isolates. Intracellular Na+ values were affected by washing; however, bound-K+ values for both marine and fresh-water blue-green algae were fairly constant, 1–3 μg/mg cells. A specific Na+ function was implied by the retention after washing of 22Na+ (0.1 μg/mg cells) by Agmenellum quadruplicatum (PR-6), a marine coccoid blue-green alga.High concentrations of NaCl apparently inhibit growth more by ionic (Na+) stress than by osmotic stress. Changes in light, temperature, pH, or composition of the basal medium failed to alleviate this stress.In contrast to marine bacteria, cells of PR-6 grown in Medium ASP-2+90 g NaCl/l did not undergo lysis when suspended in distilled water. However, viability of cells grown in Medium ASP-2+90 g NaCl/l decreased rapidly compared to cells grown in Medium ASP-2+18 g NaCl/l.Cells of PR-6 grown in ASP-2+90 g NaCl/l were larger than normal, formed chains (3–16 cells), and appeared bleached. Analyses of such cells revealed an overall decrease in fatty acids, hydrocarbons, and pigment levels. Electron micrographs showed that NaCl stressed cells were little altered in morphology.The photosynthesis of PR-6 cells was immediately depressed when the cells were transferred from 18 g NaCl/l to 70 g NaCl/l medium. When held in the latter for several hours the rate recovered and approached the initial photosynthetic rate maintained before NaCl-shock. This phenomenon was never seen with non-marine isolates. The explanation may lie in the ability of the cell to adjust to sudden Na+ increase via an ion (Na+) pump, for example, adenosine triphosphatase (ATPase). Subsequent assays suggested more ATPase activity in a marine isolated than in a nonmarine isolate. The ATPase was not, however, ouabain sensitive.It is suggested that marine blue-green algal isolates are characteristically more halotolerant, perhaps by selection, than fresh-water forms. This difference may be due in part to inherent capacity of the cell to extrude Na+. Alternatively, in freshwater forms rhe Na+ functional sites may be more Na+ sensitive than in marine forms.

ACTION SPECTRA FOR ULTRAVIOLET KILLING AND PHOTOREACTIVATION IN THE BLUE-GREEN ALGA AGMENELLUM QUADRUPLICATUM

Abstract— The action spectrum for photoreactivation has been determined in a coccoid blue‐green alga, Agmenellum quadruplicatum. The spectrum is rather similar to that recorded for Streptomyces griseus conidia, with some suggestion of a little more structure. The action spectrum suggests possible carotenoid involvement; however, no other evidence for this could be found. The action spectrum for u.v. killing is also broad with some evidence of fine structure. The possible implication of tetrahydropteridines or c‐phycocyanin as chromophores in the region 240–300 nm, along with DNA, is pointed out.

Sensitivity of three microalgae to crude oils and fuel oils

Abstract Four crude oils and five fuel oils have been tested for toxicity with three microalgae—a blue-green, a green and a diatom. The oils were absorbed on filter paper pads and the pads submerged in the growth medium. The crude oils were less inhibitory than equal amounts of fuel oils. Despite initial growth lags, the crude oils allowed growth at 30 μl/20 ml of culture medium (105 cells/ml) while fuel oils were lethal at 10 μl/20 ml. The toxicity patterns of two of the whole fuel oils were different from that seen with their water soluble fractions (WSF); for example, the Baton Rouge fuel oil sample was very toxic to growth of the three test organisms whereas its WSF was relatively innocuous. Photosynthesis of a sensitive organism Chlorella autotrophica, strain 580 (107 cells/ml), was only temporarily depressed by the crude oils (30 μl/20 ml). Four of the fuel oils inhibited photosynthesis, O2 output decreasing to zero without recovery. When the fuel oils were heated in a stream of helium they were detoxified. Chemical analyses of two of the toxic fuel oils before and after heating were compared with analyses of the Montana sample, a largely non-toxic fuel oil, in an effort to determine what types of compounds might be involved. Classes of aromatic compounds which were not accountable for the toxicity observed here include naphthalene, methylnaphthalenes, dibenzothiophenes, phenanthrenes and compounds with volatilities greater than methylnaphthalenes. Paraffinic and asphaltic fractions of fuel oil were also non-toxic. The accumulated chemical data suggest that the toxicity of whole fuel oils is due to the less water soluble types of compounds in the higher boiling aromatic fraction.

Growth characteristics of selected mutants of a coccoid blue-green alga

SummaryAfter exposure of cells to N-methyl-N′-nitro-N-nitrosoguanidine a series of stable mutants of the marine coccoid blue-green alga, Agmenellum quadruplicatum, were isolated.1.Two of these mutants were blocked at nitrate reductase and required nitrite or ammonium ion for growth.2.Four of the mutants were blocked at nitrite reductase, required NH4+ for growth but had a functional nitrate reductase. These mutants readily reduced NO3- to NO2-.3.Two of the mutants had altered phycocyanin/chlorophyll a ratios, together with slightly impaired growth rates.4.Mutant AQ-19 responded to casamino acids and possibly represents an amino acid auxotroph.

Effects of selected inhibitors on growth and cell division inAgmenellum

SummaryConcentrations of chloramphenicol and penicillin G which permit growth induce the formation of temporary filaments, morphologically and ultrastructurally identical to stable, chemically-induced filamentous mutants ofAgmenellum quadruplicatum strainBG-1. These induced filaments were propagated by serial transfers in the presence of inhibitor and underwent an immediate, synchronous reversion upon its removal. The reversion of penicillin-induced filaments was insensitive to inhibitors of DNA synthesis but sensitive to inhibitors of protein synthesis until the completion of the mucopolymer septum. Penicillin G blocked the early stages or initiation of cell division. Chloramphenicol blocked the terminal stages of cell division, the cleavage of the mucopolymer septum by the outer wall layers.

THE OXIDATION OF AROMATIC COMPOUNDS BY MICROALGAE

Publisher Summary This chapter discusses the oxidation of aromatic compounds by microalgae. Recent work with microalgae has concentrated on the isolation and identification of biotransformation products formed when various microalgae were incubated with substrates such as naphthalene, methylnaphthalenes, biphenyl, and aniline. The accumulated data strongly implies that microalgae can metabolize aromatic compounds, and this suggests that the phytoplankton may play a heretofore unsuspected role in the degradation of aromatic compounds in the environment. In an environmental context, it now seems that the phytoplankton should be reckoned with in any balance sheet of the degradation of aromatic compounds in the sea. In the context of the evolution of metabolic pathways, there is an interesting speculation that follows from the finding that cyanobacteria can oxidize aromatic compounds. The cyanobacteria are generally viewed as the first group of organisms to develop the capacity for an O2 evolving photosynthesis. This suggests that the first oxygen requiring enzymes capable of the hydroxylation of an aromatic ring evolved in an ancestral cyanobacterium.