Botryococcus braunii cells: ultrasound-intensified outdoor cultivation integrated with in situ magnetic separation.

An integrated system combining ultrasound-intensified outdoor cultivation of Botryococcus braunii with in situ magnetic harvesting of the algal cells was developed. The algal cells were cultivated in 200 L plastic bag reactors, and seven five-minute ultrasonic treatments at a four-day interval using a fixed frequency of 40 kHz and a total power of 300 W improved algal cell biomass and hydrocarbon productivity. The algal cells were harvested using functional magnetic particles and a magnetic separator, and a recovery efficiency of 90% was obtained under continuous operation at a flow rate of 100mL/min using the in situ magnetic separation system. The overall production cost using the integrated system was US$ 25.14 per kilogram of B. braunii dry biomass. The system developed in this study provides a base for the industrial production of B. braunii.

Scale-up cultivation of Chlorella ellipsoidea from indoor to outdoor in bubble column bioreactors.

The cultivation of Chlorella ellipsoidea in bubble column bioreactors was investigated at different scales under indoor and outdoor conditions. The algal cells were able to quickly adapt to the outdoor conditions and achieved a growth rate of 31.55mg L(-1)day(-1). Due to differences in light and temperature, the outdoor culture produced a higher percentage of unsaturated fatty acids compared to the indoor cultures, while the amino acid composition was unaffected. The overall cost of the biomass produced by the 200L outdoor cultivation (58.70US$/kg-dry weight) was estimated to be more than 7 times lower than that of the 20L indoor cultivation (431.39US$/kg-dry weight). Together these results provide a basis for the cultivation of C. ellipsoidea for the large-scale production of biofuels, high-value nutrients and/or recombinant proteins.

Magnetic flocculant for high efficiency harvesting of microalgal cells.

Magnetic flocculant was synthesized for the highly efficient recovery of microalgal cells. The highest flocculation was achieved using the magnetic flocculant synthesized with iron oxide and 0.1 mg/mL cationic polyacrylamide (CPAM). This resulted in a recovery efficiency of more than 95% within 10 min using a dosage of 25 mg/L for Botryococcus braunii and 120 mg/L for Chlorella ellipsoidea. For both species, the adsorption isotherm data fit the Freundlich model better than the Langmuir model, indicating that the adsorption process was a heterogeneous multilayer. The maximum adsorption capacity was 114.8 and 21.4 mg dry cells/mg-particles at pH 7 for B. braunii and C. ellipsoidea, respectively. The primary flocculation mechanism was bridging, which was assisted by the electrostatic interactions between the microalgal cells and the magnetic flocculant under acidic conditions. These results provide new opportunities and challenges for understanding and improving the harvesting of microalgae using magnetic separation.

Hairy root culture: bioreactor design and process intensification.

The cultivation of hairy roots for the production of secondary metabolites offers numerous advantages; hairy roots have a fast growth rate, are genetically stable, and are relatively simple to maintain in phytohormone free media. Hairy roots provide a continuous source of secondary metabolites, and are useful for the production of chemicals for pharmaceuticals, cosmetics, and food additives. In order for hairy roots to be utilized on a commercial scale, it is necessary to scale-up their production. Over the last several decades, significant research has been conducted on the cultivation of hairy roots in various types of bioreactor systems. In this review, we discuss the advantages and disadvantages of various bioreactor systems, the major factors related to large-scale bioreactor cultures, process intensification technologies and overview the mathematical models and computer-aided methods that have been utilized for bioreactor design and development.

Gibberellic acid increases secondary metabolite production in Echinacea purpurea hairy roots.

Gibberellic acid (GA(3)) is reported to have diverse effects on hairy root cultures of many plant species; therefore, the effects of GA(3) on the growth, secondary metabolite production (caffeic acid derivatives and lignin), phenylalanine ammonia lyase (PAL) activity, and free radical scavenging activity of light-grown Echinacea purpurea L. hairy roots were investigated. Eight concentrations of GA(3), ranging from 0.005 to 1.0 µM, were added to shake flask cultures. The moderate GA(3) concentration, 0.025 µM, resulted in the highest concentrations of cichoric acid, caftaric acid, and chlorogenic acid, as well as increased PAL activity, cell viability, and free radical scavenging activity, while higher and lower GA(3) concentrations resulted in reduced levels compared to the control (lacking GA(3)). The moderate GA(3) concentration also affected root morphogenesis; supplementation with 0.025 µM GA(3) resulted in the development of thick, dense, purple-colored roots, while roots exposed to the higher and lower concentrations of GA(3) were thin and off-white. This study demonstrates that supplementation with GA(3) may be an excellent strategy to optimize the production of secondary metabolites from E. purpurea hairy root cultures; however, the GA(3) concentration is a critical factor.

Isolation of an extremely boron-tolerant strain of Bacillus firmus.

A strain of Bacillus firmus (designated strain KC) isolated from a boron (B) mine in California exhibited extreme tolerance to B, provided it was first acclimated at intermediate B supply concentrations. Strain KC tolerated up to 1000 mmol/L B (boric acid-B) and 1800 mmol/L B (sodium tetraborate-B), and attained the greatest growth (as measured by absorbance) at 300 mmol/L B. Despite its extreme tolerance to high B, there was no evidence that it was able to remove significant quantities of B from the growth media, suggesting that strain KC is not likely to be useful for the removal of B from wastewaters in an engineered bioreactor.

Evaluation of the boron tolerant grass, Puccinellia distans, as an initial vegetative cover for the Phytorestoration of a boron-contaminated mining site in Southern California.

Land damaged by boron (B) mining should be restored to its natural state with a zero net impact on biodiversity. In an earlier study (Environ. Sci. Technol.2010,44, 7089-7095), we characterized a Turkish ecotype of the grass, Puccinellia distans, which exhibited extreme tolerance to B. Here we evaluated the use of a US ecotype of P. distans as an initial vegetative cover for the phytorestoration of a B mine in southern California. Hydroponic studies revealed that this P. distans ecotype tolerated B concentrations >100 mg B/L and could be germinated and grown in B-contaminated soils taken from the sites to be restored. P. distans grew well in moderately B-contaminated soil (∼88 mg B/L saturated extract) amended with added organic matter (peat moss); other soil treatments such as gypsum addition or pH correction were not needed. P. distans also grew in severely B-contaminated soil (∼1506 mg B/L) provided that toxic levels of soil B were diluted by the addition of sand and/or organic matter. Our results provide evidence in support of the concept of using the US ecotype of P. distans as an initial vegetative cover for the phytorestoration of B-contaminated soil.

Mechanisms of boron tolerance and accumulation in plants: a physiological comparison of the extremely boron-tolerant plant species, Puccinellia distans, with the moderately boron-tolerant Gypsophila arrostil.

The physiological characteristics of the extremely boron (B)-tolerant plant species, Puccinellia distans, were compared with those of the moderately tolerant Gypsophila arrostil, two species collected from a B-mining area of Eskisehir, Turkey. Boron was supplied to plants hydroponically at B concentrations ranging from 0.5 to 50 mg B/L for G. arrostil, and from 0.5 to 2000 mg B/L for P. distans. The results show that P. distans has a strikingly greater tolerance to B than G. arrostil. While G. arrostil was unable to survive B supply concentrations greater than 50 mg B/L, P. distans grew at B supply concentrations exceeding 1250 mg B/L. Our research supports the conclusion that from 0.5 to 50 mg B/L, P. distans is better able to restrict the accumulation of B in the whole plant, and the transport of B from root to shoot, than G. arrostil. We propose that P. distans uses several strategies to achieve B tolerance including the ability to restrict the accumulation of B relative to its accumulation of biomass, the ability to restrict the transport of B from root to shoot, and, to a lesser extent, the ability to tolerate high concentrations of B in its shoot and root tissues.

Metabolic and signaling properties of an Itpk gene family in Glycine max.

We have cloned and characterized four Itpk genes from soybean. All four recombinant Itpk proteins showed canonical Ins(1,3,4)P3 5/6-kinase activity, but a kinetic analysis raised questions about its biological significance. Instead, we provide evidence that one alternative biological role for soybean Itpks is to interconvert the Cl(-) channel inhibitor, Ins(3,4,5,6)P4, and its metabolic precursor, Ins(1,3,4,5,6)P5, within a substrate cycle. The soybean Itpks also phosphorylated Ins(3,4,6)P3 to Ins(1,3,4,6)P4 which was further phosphorylated to Ins(1,3,4,5,6)P5 by soybean Ipk2. Thus, soybean Itpks may participate in an inositol lipid-independent pathway of InsP6 synthesis.

Integration of inositol phosphate signaling pathways via human ITPK1.

Inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1) is a reversible, poly-specific inositol phosphate kinase that has been implicated as a modifier gene in cystic fibrosis. Upon activation of phospholipase C at the plasma membrane, inositol 1,4,5-trisphosphate enters the cytosol and is inter-converted by an array of kinases and phosphatases into other inositol phosphates with diverse and critical cellular activities. In mammals it has been established that inositol 1,3,4-trisphosphate, produced from inositol 1,4,5-trisphosphate, lies in a branch of the metabolic pathway that is separate from inositol 3,4,5,6-tetrakisphosphate, which inhibits plasma membrane chloride channels. We have determined the molecular mechanism for communication between these two pathways, showing that phosphate is transferred between inositol phosphates via ITPK1-bound nucleotide. Intersubstrate phosphate transfer explains how competing substrates are able to stimulate each others' catalysis by ITPK1. We further show that these features occur in the human protein, but not in plant or protozoan homologues. The high resolution structure of human ITPK1 identifies novel secondary structural features able to impart substrate selectivity and enhance nucleotide binding, thereby promoting intersubstrate phosphate transfer. Our work describes a novel mode of substrate regulation and provides insight into the enzyme evolution of a signaling mechanism from a metabolic role.