Jong-Woo Park, Wonho Yih Department of Oceanography, Kunsan National University MPM Ecology & Technology Laboratory Photobiological H 2 production by Korean N 2 -fixing unicellualr cyanobacterial strains 2010 NHA Hydrogen Conference & Expo

1.Background 2.Photobiological H 2 production by cyanobacteria 3.Material & Method 4.Result 5.Conclusion 6.Further study

1. Background 1-1. Shortage of fossil fuel 1-2. Environmental problems : greenhouse effect… “Needs for clean energy production” - wind - tidal power - solar energy - hydrogen

“Different H 2 producing micro-organisms ” Cyanobacteria (Mitsui, 1981) Green algae (Gaffron & Rubin, 1942) Anaerobic bacteria (Yasuo, 1999) 1. Background

2. Photobiological H 2 production by cyanobacteria 2-1. Cyanobacteria - Present in all marine environment - Photosynthetic prokaryote - Some N 2 -fixers 2-2. H 2 producing cyanobacteria - H 2 production under CO 2 -free condition - Using N 2 ase & H 2 ase

2-3. Cyanobacterial H 2 - clean energy - sun light, H 2 O O 2, H 2 - cell as biocatalyst - room-temperature reaction (H 2 O 2H 2, O 2 ) light 2. Photobiological H 2 production by cyanobacteria

2-4. H 2 production by N 2 -fixing cyanobacteria photosynthesis: with CO 2 → organic matter, O 2 H 2 production: without CO 2 → H 2 O bio-photolysis ↗ O 2 light ↗ H 2 O → → PS → (CH 2 O) ↗ ↗ CO 2 (CH 2 O) → FD → H 2 ase, N 2 ase → H 2 N 2 + 8H ATP → 2NH 3 + H ADP + 16Pi 2. Photobiological H 2 production by cyanobacteria

3. Material & Method

3-1. Isolation of strains 3. Material & Method

Synechococcus sp. Miami BG Cyanothece sp. KNU CB-MAL031 KNU CB-MAL058 * Size bar = 20um 3. Material & Method - Bahama Island, Garorim Bay, Taean, June Garorim Bay, Taean, October 2006

3-2. High density culture (for 2wks) - medium : f/2-BGER - light : 100 μE m -2 s -1 - temperature : 20 ℃ - aeration : mixing effect 3. Material & Method

3-3. Synchronization of cell cycle (72hrs) - NO 3 -free medium (N 2 ase activation) - LD cycle (14 : 10) - aeration (mixing) 3. Material & Method

- 350 μE m -2 s -1 (continuous) - 20 ℃ 3-4. H 2 production (96hrs) - flushing of head space w/ N 2 3. Material & Method

range of cell density (10 8 ml -1) - Low CD : 1~ 4 - Middle CD : 5~10 - High CD : 10~17 - 3ml flask -1 - connecting syringe - cell suspension 3. Material & Method 3-4. H 2 production (96hrs)

3-5. Measurement of produced H 2 1. GC (SRI-8610C) w/ HID detector 2. Measuring interval : 0, 4, 12, 24, 36, 48, 72, 96 h 3. Material & Method

3-5. Measurement of produced H 2

4. Result H 2 production per flask (w/ 3ml cell suspension) Average H 2 BG : 1.37 CB031 : 1.55 CB058 : 0.83 Low Cell density

4. Result H 2 production per flask (w/ 3ml cell suspension) Medium Cell density Average H 2 BG : 2.04 CB031 : 5.56 CB058 : 4.06

4. Result H 2 production per flask (w/ 3ml cell suspension) High Cell density Average H 2 BG : 1.73 CB031 : 2.66 CB058 : no product

To summarize… H 2 production per flask (w/ 3ml cell suspension) 4. Result

H 2 production per billion cells 4. Result To summarize…

5. Conclusion 1. CB031 & Miami BG produced H 2 at all the tested cell densities up to 15 × 10 8 cells ml Tested strains showed different ranges of cell densities for optimal H 2 production 3. Maximum H 2 production (ml H 2 / flask) Miami BG : 6.26 (at high CD) CB031 : 7.41 (at middle CD) 4. No H 2 production detected over 15 × 10 8 cells ml -1

6. Further study 6-1. Searching for better strains - temperature - salinity - life cycle - light

6-2. Synchronization of cell cycle Mitsui et al., 1986 (Nature) 6. Further study

6-3. Inhibition of H 2 oxidation S. Kumazawa, 2003 (Mar. Bio technology) 6. Further study

6-3. Inhibition of H 2 oxidation (Lab. data)

Thank you very much !