Kristiina Kiviharju

High-level production of D-mannitol with membrane cell-recycle bioreactor.

Ten heterofermentative lactic acid bacteria were compared in their ability to produce D-mannitol from D-fructose in a resting state. The best strain, Leuconostoc mesenteroides ATCC-9135, was examined in high cell density membrane cell-recycle cultures. High volumetric mannitol productivity (26.2 g l−1 h−1) and mannitol yield (97 mol%) were achieved. Using the same initial biomass, a stable high-level production of mannitol was maintained for 14 successive bioconversion batches. Applying response surface methodology, the temperature and pH were studied with respect to specific mannitol productivity and yield. Moreover, increasing the initial fructose concentration from 100 to 120 and 140 g l−1 resulted in decreased productivities due to both substrate and end-product inhibition of the key enzyme, mannitol dehydrogenase (MDH). Nitrogen gas flushing of the bioconversion media was unnecessary, since it did not change the essential process parameters. Journal of Industrial Microbiology & Biotechnology (2002) 29, 44–49 doi:10.1038/sj.jim.7000262

Metabolic Engineering of Lactobacillus plantarum for Production of l-Ribulose

ABSTRACT l-Ribulose is a rare and expensive sugar that can be used as a precursor for the production of other rare sugars of high market value such as l-ribose. In this work we describe a production process for l-ribulose using l-arabinose, a common component of polymers of lignocellulosic materials, as the starting material. A ribulokinase-deficient mutant of the heterofermentative lactic acid bacterium Lactobacillus plantarum NCIMB8826 was constructed. Expression of araA, which encodes the critical enzyme l-arabinose isomerase, was repressed by high glucose concentrations in batch cultivations. A fed-batch cultivation strategy was therefore used to maximize l-arabinose isomerase production during growth. Resting cells of the ribulokinase-deficient mutant were used for the production of l-ribulose. The isomerization of l-arabinose to l-ribulose was very unfavorable for l-ribulose formation. However, high l-ribulose yields were obtained by complexing the produced l-ribulose with borate. The process for l-ribulose production in borate buffer by resting cells was optimized using central composite designs. The experiment design suggested that the process has an optimal operation point around an l-arabinose concentration of 100 g liter−1, a borate concentration of 500 mM, and a temperature of 48°C, where the statistical software predicted an initial l-ribulose production rate of 29.1 g liter−1 h−1, a best-achievable process productivity of 14.8 g liter−1 h−1, and a conversion of l-arabinose to l-ribulose of 0.70 mol mol−1.

On-line biomass measurements in bioreactor cultivations: comparison study of two on-line probes.

Two on-line probes for biomass measurement in bioreactor cultivations were evaluated. One probe is based on near infrared (NIR) light absorption and the other on dielectric spectroscopy. The probes were used to monitor biomass production in cultivations of several different microorganisms. Differences in NIR probe response compared to off-line measurement methods revealed that the most significant factor affecting the response was cell shape. The NIR light absorption method is more developed and reliable for on-line in situ biomass estimation than dielectric spectroscopy. The NIR light absorption method is, however, of no significant use, when the cultivation medium is not clear, and especially in processes using adsorbents or solid matrix for the microorganism to grow on. The possibilities offered by dielectric spectroscopy are impressive, but the on-line probe technology needs to be improved.

Optimization of Streptomyces peucetius var. caesius N47 cultivation and ε-rhodomycinone production using experimental designs and response surface methods

Streptomyces peucetius var. caesius is an aerobic bacterium that produces doxorubicin as a secondary metabolite. A mixture design was applied for the screening of suitable complex medium components in the cultivation of S. peucetius var. caesius N47, which is an ε-rhodomycinone-accumulating mutant strain. ε-Rhodomycinone is a non-glycosylated precursor of doxorubicin. Best growth results were obtained with soy peptone and beef extract. A central composite face-centered (CCF) experimental design was constructed for the investigation of pH, temperature and dissolved oxygen (DO) effects on the cultivation growth phase. Another CCF was applied to the production phase to investigate the effects of aeration, pH, temperature and stirring rate on ε-rhodomycinone production. An increase in cultivation temperature increased both cell growth and glucose consumption rate. Best ε-rhodomycinone productivities were obtained in temperatures around 30°C. DO control increased all growth phase responses, but aeration in the production phase coupled with pH decrease resulted in rapid ε-rhodomycinone decay in the medium. In non-aerated production phases a pH change resulted in better productivity than in experiments without pH change. A pH increase with a temperature decrease seemed most beneficial for productivity. This implies that dynamic control strategies in batch production of ε-rhodomycinone could increase the overall process productivity.

Contributions of Biotechnology to the Production of Mannitol

Mannitol is a 6-carbon sugar alcohol that has been produced traditionally by chemical catalysis. Biotechnology has brought advances to the production in terms of substrate purity, process equipment requirements and safety. Enzymatic methods have improved the yields and the use of microbes has brought versatility to the range of substrates that can be used in the processes. Some of the microbial processes are already industrially feasible and could be taken to further use. The most promising production strategy reported so far has been the utilization of non-growing cells of heterofermentative lactic acid bacteria for converting fructose to mannitol. The latest developments in the field have dealt with the use of recombinant strains in mannitol production.

BIOREACTOR MEASUREMENT AND SIMULATION ENVIRONMENT

Abstract The aim of the work was to generate a flexible measurement, control and simulation environment for laboratory and pilot scale experiments. Various measurement devices were connected and synchronized to an integrated system with many bioreactors. Integrated devices were for example biomass monitoring, mass spectrometer and glucose control. Microcontroller-based interface modules and device drivers capable of transferring data to Multi Fermentor Control System (MFCS) via software interface were developed for each integrated device. A simple network TCP/IP interface enabled to distribute measurement devices in the laboratory intranet. The final result was an enhanced and flexible bioreactor data acquisition system.