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Enhanced hydrogen production by green alga Scenedesmus obliquus TISTR 8546 under atmospheric air through potassium deprivation and cysteine supplementation
(2026-03-01)
Sereetrakul, Kodchaporn
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Taikhao, Samart
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Chinchusak, Nattanon
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Phunpruch, Saranya
Hydrogen is a promising alternative energy carrier that can be produced by green microalgae. However, the key enzyme catalyzing this process, [FeFe]‑hydrogenase, is highly sensitive to O<inf>2</inf> generated during photosynthesis, which severely restricts its activity. This study aimed to screen the microalgal strain capable of producing high levels of H<inf>2</inf> in the presence of O<inf>2</inf> and to enhance H<inf>2</inf> production yields using O<inf>2</inf>-scavenging strategies to levels comparable to or exceeding those achieved under anaerobic conditions. Among the 24 strains tested, Scenedesmus obliquus TISTR 8546 demonstrated the highest H<inf>2</inf> production under both aerobic and anaerobic conditions. The O<inf>2</inf>I<inf>50</inf> of S. obliquus TISTR 8546 cells for H<inf>2</inf> evolution was 15.93 ± 0.24%. Potassium deprivation significantly enhanced H<inf>2</inf> production and hydrogenase activity by lowering O<inf>2</inf> levels through reduced photosynthetic O<inf>2</inf> evolution and increased dark respiration. Moreover, potassium deprivation promoted intracellular starch accumulation, providing reducing equivalents for H<inf>2</inf> generation. Cysteine supplementation further stimulated H<inf>2</inf> production by serving as a reducing agent. S. obliquus TISTR 8546 exhibited a maximum H<inf>2</inf> production rate of 22.92 ± 1.05 μmol H<inf>2</inf> mg Chl−1 h−1 and achieved a maximum cumulative H<inf>2</inf> production of 1153.78 ± 52.65 μmol H<inf>2</inf> mg Chl−1 when incubated in potassium-deprived TAP medium supplemented with 0.1 mM cysteine under atmospheric air for 6 days. This rate was 23.6- and 2.7-fold higher than those obtained in TAP and TAP-K media, respectively. These findings demonstrate the potential of S. obliquus TISTR 8546 as a robust microalgal strain for sustainable H<inf>2</inf> production under atmospheric air, highlighting its promise for future industrial applications.
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Physiological and Environmental Factors Influencing Hydrogen Production by Unicellular Green Alga Monoraphidium sp. KMITL-1
(2025-10-01)
Krutpan, Varanya
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Supakriangkrai, Thaninthorn
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Lohasupthawee, Pana
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Phunpruch, Saranya
With the growing global energy demand and the urgent need to reduce carbon emissions, hydrogen (H<inf>2</inf>) has emerged as a promising clean energy carrier. Among various biological H<inf>2</inf> production, green algae present a sustainable and eco-friendly alternative due to their ability to produce H<inf>2</inf> via photobiological pathways. This study aimed to investigate H<inf>2</inf> production by unicellular green alga Monoraphidium sp. KMITL-1, isolated from hydroponic water at the Plant Tissue Culture Laboratory, King Mongkut’s Institute of Technology Ladkrabang. The taxonomic identity of the strain, belonging to the genus Monoraphidium within the Selenastraceae family, was confirmed through morphological observation and molecular characterization using 23S plastid rRNA gene sequencing. Various physiological and environmental parameters influencing H<inf>2</inf> production were evaluated, including cell age, cell density, nutrient deprivation, carbon source, pH, temperature, and light intensity. A 24-hour-old culture with an OD<inf>750</inf> of 0.8 exhibited a significant increase in H<inf>2</inf> production. The optimal medium was potassium-deprived Tris-acetate-phosphate (TAP-K) supplemented with glucose at a concentration of 350 mmol C-atom L-1. The ideal environmental conditions for H<inf>₂</inf> production were pH 7.2, a temperature of 30 °C, and a light intensity of 60 μmol photons m-2 s-1. Under these optimized conditions, Monoraphidium sp. KMITL-1 achieved a maximum H<inf>2</inf> production rate of 67.976 ± 1.096 μmol H<inf>2</inf> mg Chl-1 h-1 and a cumulative H<inf>2</inf> yield of 3,190.436 ± 2.219 μmol H<inf>2</inf> mg Chl-1 after 72 h of incubation. These results highlight the potential of Monoraphidium sp. KMITL-1 for large-scale biohydrogen production and its applicability in the development of sustainable energy technologies.
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Freshwater green alga Chlorella sp. KLSc59 produced all forms of omega-3 oil: ALA, EPA, and DHA
(2024-06-01)
Preechaphonkul, Nathanan
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Sirikwanpong, Sukrit
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Maneeruttanarungroj, Cherdsak
Omega-3 fats are known to be the essential nutrient for human health, obtainable only through food sources. Algae and phytoplankton are primary sources that can synthesize various useful metabolites, such as carotenoids, antioxidants, and omega-3 oil. As the trend toward healthy alternative food options grows, this study aimed to screen eight strains of freshwater microalgae that might be promising sources of omega-3 oil. The results showed that three algal strains-BNG2, Chlorella sp. KLSc59, and Chlorella sp. KLSc61-had the ability to produce all forms of omega-3 oil, including alpha-linolenic acid (ALA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA). Among these strains, Chlorella sp. KLSc59 exhibited the most consistent omega-3 fatty acids-production when detected with gas chromatography equipped with flame ionization detector. Under optimal conditions (30 % flask volume, 10 mM CaCl<inf>2</inf>, pH of 6.0, and light incubation at 162 μmol photons m−2 s−1) for 5 days, Chlorella sp. KLSc59 produced the highest amount of omega-3 fat, yielding 21.74 ± 0.63 % of the total peak area (equivalent to 18.43 ± 0.6 μg/mg DCW), including 21.53 ± 0.66 % (equivalent to 18.26 ± 0.7 μg/mg DCW) ALA, 0.03 ± 0.2 % (equivalent to 0.02 ± 0.0 μg/mg DCW) EPA, and 0.18 ± 0.01 % (equivalent to 0.15 ± 0.0 μg/mg DCW) DHA, with an omega-6 to omega-3 ratio of 2.28. This study highlights the potential use of Chlorella sp. KLSc59 for omega-3 oil production and its role as an alternative source of omega-3 oil for the next generation.
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In silico encrypted peptide from green alga Tetraspora sp. CU2551 showed high antimicrobial activities
(2024-03-01)
Tonawut, Yothawut
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Rittiyan, Nutnicha
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Kornrawudaphikasama, Yosita
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Klinsalee, Rachaneekorn
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Laokua, Natwikar
The green alga Tetraspora sp. CU2551 has been previously identified as a biohydrogen producer for an alternative option for sustainable energy production development. Its biomass still contains a source of valuable biomolecules, especially proteins. Crude algal protein showed significant antimicrobial activity against Gram-positive bacteria (Bacillus subtilis TISTR 1248 and Staphylococcus aureus TISTR 746) and Gram-negative bacteria (Escherichia coli TISTR 074 and Pseudomonas aeruginosa TISTR 2370), representing a clear zone of 19.0 ± 0.1, 18.3 ± 0.5, 17.7 ± 0.5, and 19.0 ± 0.5, respectively. Further investigation revealed that encrypted peptides from pepsin-hydrolyzed crude proteins increased the inhibition activity against these bacteria by 3.4–10.5 %. The hydrolyzed peptides were fractionated using a Sephadex G-25 column, and peptides in fraction 17 showed the highest antimicrobial activity. Through peptide identification using LC-MS and prediction of obtained peptide primary sequences with four bioinformatic tools (DBAASP, AMPDiscover, iAMPpred, and AMPscanner), five peptides were selected and synthesized for activity evaluation. The PT11 peptide (RYGAAMAVGIACA) demonstrated the highest antimicrobial activity among the five peptides tested and compared favorably to previously reported algal encrypted peptides. The findings of this research demonstrate that the encrypted PT11 peptide derived from Tetraspora sp. CU2551 has promising potential as an antimicrobial agent. Additionally, we suggest that PT11 might be used for other things besides this, like animal feed.
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Optimal conditions for maximized H2 yield from a new green algal strain Chlorella sp. KLSc61
(2022-08-01)
Laokua, Natwikar
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Rittiyan, Nutnicha
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Kornrawudaphikasama, Yosita
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Klinsalee, Rachaneekorn
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Tonawut, Yothawut
Hydrogen produced from microalgae is attracting the attention of scientists as a potential new ‘Bio-Circular-Green’ energy source. We screened a selection of naturally occurring algal strains for H<inf>2</inf> yield as a basis for high yield development. The best strain was a new strain of biohydrogen-producing green algae from a fresh water source at King Mongkut's Institute of Technology Ladkrabang, Thailand. Using morphological and 18S rDNA sequencing analyses, this alga was identified and classified as Chlorella sp. KLSc61. When adapted cells in TAP-K medium (with starting pH at 9.0) containing 25 mM ethanol, with 54 μmol photons m−2 s−1 light, and incubated at 35 °C, Chlorella sp. KLSc61 showed a maximum H<inf>2</inf> yield of 9,804 ± 469.5 μmol H<inf>2</inf> mg−1 Chl at day 7. Under these conditions, the cells produced ~ 12.2 times more H<inf>2</inf> gas than without optimization (753.0 μmol H<inf>2</inf> mg−1Chl). Our optimized system also produced H<inf>2</inf> at a rate 11–570 times greater than previously reported microalgae. Thus Chlorella sp. KLSc61 could be a viable strain for biohydrogen production as a clean future energy source.
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Identification of major carotenoids from green alga Tetraspora sp. CU2551: partial purification and characterization of lutein, canthaxanthin, neochrome, and β-carotene
(2022-08-01)
Maswanna, Thanaporn
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Maneeruttanarungroj, Cherdsak
The green algae Tetraspora sp. CU2551 was previously identified as a strain with high potential for biohydrogen production; however, its algal biomass characteristics changed from green to reddish orange within 43 days of biohydrogen production. The crude pigments were extracted, partially purified, and characterized by chemical determination. The present study focused on elucidating the carotenoid composition of the selected green alga Tetraspora sp. CU2551. The pigment extract was partially purified and fractionated using thin layer chromatography, and yielded two major and two minor carotenoid bands. The fractions were confirmed by high-performance liquid chromatography with a diode array detector (HPLC–DAD) before being identified and confirmed using Liquid Chromatograph-Quadrupole Time of Flight-Mass Spectrometry (LC-QTOF-MS). The spectral data of these fractions revealed four sub-fractions of interest that were lutein, canthaxanthin, neochrome, and β-carotene, which had percentages in the crude extracts of 30.57%, 25.47%, 7.89%, and 0.71%, respectively. Lutein and canthaxanthin were found to be the major carotenoid pigments present. Our findings in this present study are the first reporting of Tetraspora sp. CU2551 as a potential alternate source for carotenoid pigment production.
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Response of green alga Tetraspora sp. CU2551 under potassium deprivation: a new promising strategy for hydrogen production
(2022-04-01)
Pewnual, Totsapon
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Jampapetch, Narirat
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Saladtook, Sathonkorn
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Raksajit, Wuttinun
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Klinsalee, Rachaneekorn
With increasing world energy consumption, renewable energy sources can fulfill the need and many have net zero carbon dioxide emissions. One of these is hydrogen, which is biologically produced molecular hydrogen from organisms. Although many studies have produced hydrogen from green algae and optimized conditions to obtain the highest yield, following our previous works on production optimization from Tetraspora sp. CU2551, we tested deprivation of Fe, Cu, Ca, S, Mg, or K, and surprisingly found that K deprivation showed another promising switch in increased H<inf>2</inf> yield, representing 9.2 ± 0.1 μmol mg−1 DW within 32 h incubation anaerobically. Moreover, longer cell adaptation by aerobic incubation, before production phase, resulted in higher hydrogen yield. Cells adapted in potassium deprivation (TAP-K medium) promoted hydrogen production to a yield of 14.8 ± 0.03 μmol mg−1 DW, or about 3.0 times higher than normal TAP (5.0 ± 0.82 μmol mg−1 DW). The increased yield was caused by (1) lowering PSII activity (direct biophotolysis), resulted in less oxygen being produced, lowering hydrogenase inhibitor levels, and (2) increasing accumulated starch degradation (indirect biophotolysis) (12.2% starch remaining or 7.2 ± 0.9 μg mg−1 DW), increased the flow of electrons to hydrogenase. Our finding makes potassium deprivation conditions as another the promising choice to enhance hydrogen production in biological systems.
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Effect of cell density and nutrient deprivation on hydrogen production by unicellular green alga Scenedesmus sp. KMITL-OVG1
(2019-01-01)
Warichanan, Kittiphat
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Phunpruch, Saranya
Hydrogen is considered as one of the energy carriers for the near future. H2 production by green algae is catalyzed by hydrogenase activity using electrons from photosynthetic process under the light and from accumulated carbohydrate catabolism in the dark. This research aimed to investigate the effect of cell density and nutrient deprivation on H<inf>2</inf> production by Scenedesmus sp. KMITL-OVG1 isolated in Thailand. The result showed that cell culture with the optical density at 750 nm of 0.8 gave the highest H2 production rate. Interestingly, the highest H2 production rate of 1.957 ± 0.100 mL L-1 h-1 and hydrogenase activity of 0.031 ± 0.001 ml L-1 min-1 were found in cells incubated under potassium deprivation. H<inf>2</inf> production rate was approximately 3 folds higher than that of cells incubated in normal TAP medium. The increased H2 production rate and hydrogenase activity might be involved in the reduction of starch accumulation. Moreover, the deprivation of potassium combined with other nutrients did not enhance H<inf>2</inf> production rate by Scenedesmus sp. KMITL-OVG1.
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Enhanced hydrogen production by optimization of immobilized cells of the green alga Tetraspora sp. CU2551 grown under anaerobic condition
(2018-04-01)
Maswanna, Thanaporn
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Phunpruch, Saranya
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Lindblad, Peter
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Maneeruttanarungroj, Cherdsak
The green alga Tetraspora sp. CU2551 has previously been identified and characterized as a photosynthetic microorganism with high potential for H<inf>2</inf> production. In the present study, cells of Tetraspora CU2551 were entrapped and immobilized in an alginate matrix with the aim to analyze the effect of cell stacking and a reduced exposure of O<inf>2</inf> to the cells. The results showed that the most favorable immobilization conditions were 4% (w/v) of final alginate concentration and a cell concentration of 0.125 mg cell dry wt/mL alginate with a bead diameter of 2.80–3.35 mm. The H<inf>2</inf> production yields increased when the immobilized cells were incubated in S-deprived medium and this could be repeated at least for 3 production times. Maximal total H<inf>2</inf> production reached 7.68 ± 0.88 mL H<inf>2</inf>/25 mL medium, corresponding to a rate of 1182.45 ± 24.40 nmol H<inf>2</inf>/h/mg DW. This production is about 6 times higher compared to by cells in suspension, 2–10 times higher when compared to by other green algae, and 10–50 times higher when comparing with cyanobacteria. Based on our observations, immobilized cells of Tetraspora CU2551 is considered a very promising biological system for significant photobiological H<inf>2</inf> production by a photosynthetic microorganism.
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Rapid method for DNA isolation from a tough cell wall green alga Tetraspora sp. CU2551
(2016-06-01)
Maneeruttanarungroj, Cherdsak
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Incharoensakdi, Aran
Genetic studies are important to understand the complex biological system of various organisms. Some eukaryotic green organisms have tough cell wall which precludes the efficient extraction of the genetic materials. Here, we developed the method for simple and rapid isolation of high quality DNA from a green alga Tetraspora sp. CU2551. The cell homogenization procedures were combined with physical force plus heat treatment to disrupt the cell envelope of Tetraspora sp. CU2551. Without protease treatment, vortexing with glass bead for 30–105 s at 70 °C led to the isolation of a high purity DNA which was suitable for downstream process. The improved method was successfully developed and could be applied for the rapid isolation of DNA from other unicellular and filamentous green microalgal strains.

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