Ryohei Ueda

CO2 fixation and ethanol production with microalgal photosynthesis and intracellular anaerobic fermentation

Microalgae were screened from seawater. More than 250 strains were isolated, and some of the isolated strains and two strains from culture collections were tested to examine ethanol productivity. Some strains had high growth rate of 20–30 g dry biomass/m2/day and high starch content of more than 20% (dry base). A strain Chlorella vulgaris (IAM C-534) had a high starch content of 37%. Starch was extracted from the cells of the Chlorella, saccharified and fermented with yeasts; 65% of the ethanol-conversion rate was obtained as compared to the theoretical rate from starch. The algal starch proved to be a good source for ethanol production using the conventional process. As an example of another type of ethanol production process, intracellular starch fermentation under dark and anaerobic conditions was examined. All of the tested strains showed intracellular starch degradation and ethanol production, but the levels of ethanol production were significantly different from each other. Higher ethanol productions were obtained with Chlamydomonas reinhardtii (UTEX2247) and Sak-1 isolated from seawater. These showed a maximum ethanol concentration of 1 (w/w)%. The characteristics of intracellular ethanol production were examined with the Chlamydomonas. These results indicate that intracellular ethanol production is simpler and less energy intensive than the conventional ethanol-fermentation process.

Improvement of photosynthesis in dense microalgal suspension by reduction of light harvesting pigments

The effects of light-harvesting pigments (LHP) inmicroalgal cells on photosynthetic activity in adense cell suspension were examined. The results suggest that a lower LHP content should result in higher photosynthetic productivity under high light intensity. The idea was first proposed by Lien and San Pietro in 1975 that photosynthesis could be improved by reducing the LHP content in microalgal cells, but this has not been demonstrated in detail. Experiments to evaluate the idea were conducted with Synechocystis PCC6714 and Chlorellapyrenoidosa. In the experiments with PCC 6714, photosynthesis of a phycocyanin-deficient mutant was compared with that of the wild type. In the experiments with C. pyrenoidosa, the LHP content was controlled by the light intensity in the algalculture. The maximum photosynthetic activity was 20–30% higher in the dense suspension of cells having a lower LHP content with both organisms. These results indicate that the idea of reducing the LHP contentcould be applicable to a wide variety of photosynthetic organisms.

Improvement of microalgal photosynthetic productivity by reducing the content of light harvesting pigment

Microalgal productivity was examined using both a wild type and a phycocyanin-deficient mutant of Synechocystis PCC 6714 (PD-1). The culture was conducted at various light intensities under low and high cell densities in a continuous culture system. At low light intensity, photosynthetic productivity was almost the same for both low and high cell densities. However, at higher light intensities photosynthetic productivity was higher in mutant PD-1 than in the wild type. At 2000 μmol photon m−2 s−1 the productivity was 50% higher in mutant PD-1. This result is consistent with our first report (Nakajima & Ueda, 1997), which showed that photosynthetic productivity can be improved by reducing the light harvesting pigment content in high cell density cultures at high light intensities. It is concluded that the technology for reducing LHP content is a useful method for improving photosynthetic productivity in algal mass production.

Improved productivity by reduction of the content of light-harvesting pigment in Chlamydomonas perigranulata

The effects on productivity of the light-harvesting pigment in cells of themarine Chlamydomonas perigranulata were examined using thewild type and a mutant with a lower level of the light-harvesting pigment(LHC-1). We confirmed the photosynthetic characteristics of the wild typeand LHC-1, and these were retained generally in the continuous culturesused for evaluating productivity. The maximum productivity was 1.5 timeshigher in LHC-1 than that in the wild type. The photosynthetic productivityefficiency was higher in LHC-1 than in the wild type. These resultsindicate that the improvement in productivity can be attributed to theimproved photosynthetic productivity efficiency. We conclude that thetechnique of reducing the content of light-harvesting pigment is not onlyavailable for blue-green microalgae, but also for green microalgae.

The effect of reducing light-harvesting pigment on marine microalgal productivity

The productivity was evaluated of a strain of Chlamydomonas perigranulata isolated from the RedSea. A mutant with small light-harvesting pigments(LHC-1) was obtained by UV mutagenesis. Thechlorophylls content of the wild type was twice ashigh as that of LHC-1, and the initial slope of thephotosynthesis-irradiance curve was higher in the wildtype. However, the maximum photosynthetic activity ona per cell basis was almost the same. It isconcluded that LHC-1 is a mutant with lesslight-harvesting pigment (LHP) than the wild type. Aspreviously reported, the mutant with lower LHP contenthas a higher productivity in a continuous culturesystem, so we compared the productivity of the wildtype and the mutant. The maximum productivity of LHC-1was 1.5 times higher than that of the wild type. Itis suggested that the technique of reducing thecontent of light-harvesting pigment should be madeavailable for other organisms.

Reduced photoinhibition of a phycocyanin-deficient mutant of Synechocystis PCC 6714

The effects on photoinhibition of light-harvesting pigments in microalgal cells were examined using the wild type and a phycocyanin- deficient mutant (PD-1) of Synechosystis PCC 6714. Mutant PD-1 showed higher resistance to high light than the wild type in terms of the decline of photosynthetic activity at any light intensity and with various cell densities. This suggests that the loss of productivity induced by high light intensity would be improved by reducing the content of light-harvesting pigments.

Detection of Hydroxyl Radical in Intact Cells of Chlorella Vulgaris

Using ESR with 5,5-dimethyl-1-pyrroline N-oxide (DMPO) as a spin-trapping reagent, we measured the levels of free radical species generated from living cells of Chlorella vulgaris var. vulgails (IAM C-534). To investigate the production of free radicals in the living Chlorella vulgaris cells, the influence of DMPO toward the intact cells of the Chlorella vulgaris using the O2 evolution rate was first studied as a guide. Since the O2 evolution rate was not changed by DMPO, it was judged that DMPO has no toxicity toward the intact cells of Chlorella vulgaris. Only hydroxyl radicals (.OH) were detected as the DMPO-OH adduct in the suspension of intact cells of Chlorella vulgaris irradiated with visible light. Moreover, since production of .OH was inhibited by some hydroxyl radical scavengers such as KI and ethanol, production of .OH was proved to be due to hydroxyl radicals. It was also clear that the intensity of .OH increased with increasing irradiation intensity of visible light. Therefore, it was suggested that .OH might be one of the photoinhibition factors of the intact Chlorella vulgaris cells in severe light conditions.

Effect of hydroxyl radical on intact microalgal photosynthesis

Abstract Photosynthetic CO 2 fixation with microalgae for new energy and chemical sources is one of the potential method to mitigate CO 2 emission. To gain the more productivities for CO 2 mitigation, the enhancement of photosynthetic productivity is required. We focused on the active oxygen that is supposed to be produced in algal cells and causes harmful effects on photosynthesis under high irradiation that is the case of outdoor cultivation circumstances. In this report, we have challenged to detect the active oxygens in microalgal cells, and then the results were described. The levels of free radical species generated in the living cells of Chlorella vulgaris var. vulgaris (IAM C-534) were detected with electron spin resonance (ESR) spectrometer with 5,5 dimethyl-1-pyrroline N-oxide (DMPO) as a spin-trapping reagent. Before detecting the free radicals in the living Chlorella cells, the influence of DMPO concentration toward the photosynthetic growth rate of the cells was measured. Since the growth rate was not influenced by up to 130 mM DMPO, the DMPO concentration was adjusted to 90 mM during the measurement. Only one DMPO adduct, which is assigned as the hydroxyl radical (DMPO-OH; aN=1.49mT, aH=1.49mT) was detected in the solution of intact cells of Chlorella vulgaris irradiated with visible light (1). Moreover, the production of DMPO-OH adducts was inhibited by some hydroxyl radical ( − OH) scavengers such as KI and ethanol. It has been estimated that − OH is one of the photoinhibition factors of the photosynthetic organisms. But, − OH has never been detected in vivo. In this report, − OH was detected in vivo, and the − OH was increased according to the light intensity.

Production of optically pure D-lactic acid by Nannochlorum sp. 26A4.

Microalgae were screened from seawater for greenhouse gas CO2 fixation and d-lactic acid production by self-fermentation and tested for their growth rate, starch content, and conversion rate from starch into d-lactic acid. More than 300 strains were isolated, and some of them were found to have suitable properties for this purpose. One of the best strains, Nannochlorum, sp. 26A4, which was isolated from Sakito Island, had a starch content of 40% (dry weight), and a conversion rate from consumed starch into d-lactic acid of 70% in the dark under anaerobic conditions. The produced d-lactic acid showed a high optical purity compared with the conventional one. The proposed new d-lactic acid production system using Nannochlorum sp. 26A4 should also be an effective technology for greenhouse gas CO2 fixation and/or conversion into industrial raw materials.

Evaluation of Active Oxygen Effect on Photosynthesis of Chlorella vulgaris

The relationship between O2 and an active oxygen scavenging system in Chlorella vulgaris var.vulgaris (IAM C-534) was investigated. When Chlorella vulgaris was exposed to 2% O2, only traces of active oxygen scavenging enzymes were found. When the Chlorella vulgaris was treated with 20% or 50% O2, it was shown that the level of enzyme activity increased as the O2 concentration increased. An increase in enzyme activity was not found in any specific enzyme but in all of the enzymes, but the level of glutathione and ascorbate remained the same in all the cases. In addition, the photosynthetic efficiency also decreased as the concentration of O2 was increased. These results suggest that an O2 enriched environment can lead to an increase in the production of active oxygen species such as O2.- and H2O2 and to a decrease in the photosynthetic efficiency in Chlorella vulgaris. The hydroxyl radical (.OH) was detected directly in the Chlorella vulgaris suspension with a spin trapping reagent. It was also clear that the increase in the .OH intensity as the visible light intensity increased was unrelated to the O2 concentration. It was suggested that the conditions for producting .OH and the other active oxygen species were different, and that two types of oxygen stress should exist in the Chlorella vulgaris.