Production and release of pigments by plant hairy root cultures

Taya, Masahito,Tone, Setsuji 한국농화학회 1992 Applied Biological Chemistry (Appl Biol Chem) Vol.35 No.5

Recently, plant hairy roots have become of interest because of their active proliferation in a phytohormonefree medium and their capacity to produce valuable metabolites accumulated at comparable levels of the original plant roots. The purpose of our work is to extend the biotechnological application of hairy root cultures through studies on the following subjects. 1) Induction and characterization of hairy roots: Some hairy root clones were induced from red beet (Beta vulgaris), madder (Rubia tinctorum) and horseradish (Armoraeia lapathifolia) by Agrobacterium rhizogenes infection. These hairy roots had high contents of pigments or enzymes. 2) Bioreactor configuration: A turbine-blade fermentor was developed as a bioreactor suitable for the submerged culture of hiary root with a highly branched morphology. In this fermentor, a stainless steel mesh separated the culture space from the agitation space so that the hairy root cells were not in direct contact with the impellers. 3) Extracellular production of pigments: In the cultures of red beet and madder hairy roots, significant amounts of pigments were released from the cells into medium when the cells were subjected to the treatment of O₂ starvation. Both hairy roots were capable of growing even after the treatment. 4) Morphological change of hairy root: By cultivating the horseradish hairy root in the presence of phytohormones, the cell morphology could be changed reversibly between dedifferentiated-cell aggregates and organized hairy root.

Metabolic Flux Analysis of a Poly-b-hydroxybutyrate Producing Cyanobacterium, Synechococcus sp. MA19, Grown under Photoautotrophic Conditions

Masahito Taya,Motomu Nishioka,Hajime Nishiuma,Masato Miyake,Yasuo Asada,Kazuyuki Shimizu 한국생물공학회 2002 Biotechnology and Bioprocess Engineering Vol.7 No.5

To understand the utilization property of light energy, Synechococcus sp. MA19, a poly-b-hydroxybutyrate (PHB) producer, was cultivated at the different incident light intensities of 15.3, 50.0 and 78.2 W/m2 using media with and without phosphate. From the results of metabolic flux analysis, it was found that the cell yield based on ATP synthesis was estimated as 3.5×10-3 kg-biomass/mol-ATP in these cultures. Under the examined conditions, there were no significant differences in the efficiency of light energy conversion to chemical energies estimated as ATP synthesis and reducing potential (NADH + NADPH) formation whether the PHB synthesis took place or not. The energy converted from light to ATP was kept relatively high around the energy absorbed by the cells of 2.5-3.0 x 106 J h-1 kg-1, whereas the energy of reducing potential was hardly changed in the examined range of the energy absorbed by the cells.

Production and Release of Pigments by Plant Hairy Root Cultures

Masahito Taya,Setsuji Tone 한국응용생명화학회 1992 한국응용생명화학회 학술발표회 Vol.1992 No.-

Recently, plant hairy roots have become of interest because of their active proliferation in a phytohormonefree medium and their capacity to produce valuable metabolites accumulated at comparable levels of the original plant roots. The purpose of our work is to extend the biotechnological application of hairy root cultures through studies on the following subjects. 1) Induction and characterization of hairy roots: Some hairy root clones were induced from red beet (Beta vulgaris), madder (Rubia tinctorum) and horseradish (Armoraeia lapathifolia) by Agrobacterium rhizogenes infection. These hairy roots had high contents of pigments or enzymes. 2) Bioreactor configuration: A turbine-blade fermentor was developed as a bioreactor suitable for the submerged culture of hiary root with a highly branched morphology. In this fermentor, a stainless steel mesh separated the culture space from the agitation space so that the hairy root cells were not in direct contact with the impellers. 3) Extracellular production of pigments: In the cultures of red beet and madder hairy roots, significant amounts of pigments were released from the cells into medium when the cells were subjected to the treatment of O₂ starvation. Both hairy roots were capable of growing even after the treatment. 4) Morphological change of hairy root: By cultivating the horseradish hairy root in the presence of phytohormones, the cell morphology could be changed reversibly between dedifferentiated-cell aggregates and organized hairy root.

Metabolic Flux Analysis of a Poly-${\beta}$-hydroxybutyrate Producing Cyanobacterium, Synechococcus sp. MA19, Grown under Photoautotrophic Conditions

Nishioka, Motomu,Nishiuma, Hajime,Miyake, Masato,Asada, Yasuo,Shimizu, Kazuyuki,Taya, Masahito The Korean Society for Biotechnology and Bioengine 2002 Biotechnology and Bioprocess Engineering Vol.7 No.5

To understand the utilization property of light energy, Synechococcus sp. MA19, a poly-${\beta}$-hydroxybutyrate (PHB) producer, was cultivated at the different incident light intensities of 15.3, 50.0 and 78.2 W/$m^2$ using media with and without phosphate. From the results of metabolic flux analysis, it was found that the cell yield based on ATP synthesis was estimated as $3.5{\times}10^{-3}$ kg-biomass/mol-ATP in these cultures. Under the examined conditions, there were no significant differences in the efficiency of light energy conversion to chemical energies estimated as ATP synthesis and reducing potential (NADH + NADPH) formation whether the PHB synthesis took place or not. The energy converted from light to ATP was kept relatively high around the energy absorbed by the cells of $2.5-3.0{\times}10^{6} J\;h^{-1}\;kg^{-1}$, whereas the energy of reducing potential was hardly changed in the examined range of the energy absorbed by the cells.

황산환원균 Desulfovibrio desulfricans의 혐기배양에 의한 가용성 우라늄의 침전

조규성,주현규,홍고야쓰히로,다야마사히또,도네세쯔지 한국산업미생물학회 1993 한국미생물·생명공학회지 Vol.21 No.3

황산화원균 D. desulfricans의 배양에 의하여 가용성 우라늄의 침전회수 실험을 진행한 결과를 요약하면 다음가 같다. 먼저 본실험을 위한 균주선정에서 3종의 황산환원균 D. desulfricans, D. multivorans, D. variabilis를 배양, 일정시간 후 탄소원인 젖산(lactate)과 포름산(formate)의 분해속도 및 철이온의 침전량을 조사한 결과 D. desulfricans가 가장 좋았다. 이때 이 균주는 포름산보다 젖산쪽의 이용이 양호하였다. D. desulfricans는 우라늄이 함유된 배양기(60×10exp(-3) ㎏/m^3-broth 이하)에서도 증식에 저해작용이 거의 없었으나, 우라늄이 함유되지 않은 배지에서 증식이 보다 양호하였다. 황산환원균 D. desulfricans는 젖산을 잘 이용(배양 30시간 이내에서의 분해속도 390 mmol/㎥·h)하였으며, pH는 균체 증식과 더불어 pH 9.0까지 증가하였다. 균체수는 배양 70시간에서 6.1×10^14 cells/㎥까지 증식하였다. 균체증식과 더불어서 SO^(2-)_4이온은 환원(8.5mmol/㎥)되어 S^(2-)이온의 생성(7.5 mmol/㎥)이 진행되었다. 배양액 중의 가용성 우라늄 이온은, 황산환원균 D. desulfricans의 무접종구에서는 70시간 이상이 경과하여도 용액중에 그대로 용해되어 있었다. 그러나 균체를 접종배양하였을 경우는 1시간 이내에 용액중의 우라늄은 많은 량이 침전하였다가, 균체의 증식과 더불어 다량의 CO_3^(2-)이온의 생성으로 침전하였던 우라늄의 일부가 용해하였다가, H_2S와 CO_3^(2-) 이온의 축출로 인하여 우라늄 이온은 다시 침전이 되었다. 침전한 우라늄 이온은 70% 이상 대부분이 우라늄 U(VI)로 구성되어 있었다. 따라서 황산환원균 D. desulfricans의 배양에 의하여 가용성 우라늄은, 1) 불용성 Uranyl hydroxide[UO_2(OH)_2] 형태로의 침전, 2) 불용성 Uraninite(UO_2) 형태로 S^(2-)이온 등에 의한 U(VI)에서 U(IV)에로 화원되어 침전, 3) 가용상태의 우라늄 이온이 균체세포벽에 UO_2^(2+)[U(VI)] 형태로 키일레이션 혹은 흡착이 진행되어 침전이 가능하다. The aims of the present study are to examine the precipitation of uranyl ion in the culture of Desulfovibrio desulfricans for the sedimentary recovery of aqueous uranium. D. desulfricans had the highest utilization rate of lactate and precipitated iron ion in the three sulfate reducing bacteria. So, subsequent experiments were conducted using lactate as an energy source. The normal growth was observed with increased pH and lactate utilization. During the culture, the amounts of SO_4^(2-) consumed and S^(2-) produced in aqueous phase were 8.5 and 7.5 mmol/㎥-broth, respectively. More than 85% of soluble uranium precipitated within 1 hour after the inoculation, whereas no uranium precipitation was observed for 70 hours in the culture without inoculum. It was found that the small amount of precipitated uranium was obtained in the alkaline medium with the large amount of Na_2CO_3. Uranyl hydroxide becomes soluble in the presence of CO_3^(2-) with the formation of complex ions such as [UO_2(CO_3)_2·2H_2O]^(2-). Precipitated uranium, both U(VI) and U(IV) were detected, and the former was the main form. In the culture of D. desulfricans soluble uranium (UO_2^(2-)) precipitated in the forms of U(VI) and U(IV). This uranium sedimentation was considered as attributed to the UO_2(OH)_2 and UO_2 formation, and the UO_2^(2+) adsorption to the bacterial cell walls.