Zejian Wang

Improved vitamin B12 production by step-wise reduction of oxygen uptake rate under dissolved oxygen limiting level during fermentation process

Effects of different oxygen transfer rates (OTR) on the cell growth and vitamin B(12) biosynthesis of Pseudomonas denitrificans were first investigated under dissolved oxygen limiting conditions. The results demonstrated that high OTR accelerated cell growth and initial vitamin B(12) biosynthesis rate, while lower OTR was critical for higher productivity in the late fermentation process. The oxygen uptake rates (OUR) corresponded well with OTR. Based on the metabolic intermediate analysis, a step-wise OUR control strategy was proposed. The strategy was successfully implemented in scale-up to an industrial fermenter (120,000 l). A stable maximum vitamin B(12) production of 208 + or - 2.5 mg/l was achieved, which was increased by 17.3% compared with the control. Furthermore, the glucose consumption coefficient to vitamin B(12) was 34.4% lower than that of the control. An efficient and economical fermentation process based on OUR criterion was established for industrial vitamin B(12) fermentation by P. denitrificans.

Salt stress induced lipid accumulation in heterotrophic culture cells of Chlorella protothecoides: Mechanisms based on the multi-level analysis of oxidative response, key enzyme activity and biochemical alteration

Salt stress as an effective stress factor that could improve the lipid content and lipid yield of glucose in the heterotrophic culture cells of Chlorella protothecoides was demonstrated in this study. The highest lipid content of 41.2% and lipid yield of 185.8mg/g were obtained when C. protothecoides was stressed under 30g/L NaCl condition at its late logarithmic growth phase. Moreover, the effects of salt and osmotic stress on lipid accumulation were comparatively analyzed, and it was found that the effects of NaCl and KCl stress had no significant differences at the same osmolarity level of 1150mOsm/kg with lipid contents of 41.7 and 40.8% as well as lipid yields of 192.9 and 186.8mg/g, respectively, whereas these results were obviously higher than those obtained under the iso-osmotic glycerol and sorbitol stresses. Furthermore, basing on the multi-level analysis of oxidative response, key enzyme activity and biochemical alteration, the superior performance of salt stress driving lipid over-synthesis was probably ascribed to the more ROS production as a result of additional ion effect besides the osmotic effect, subsequently mediating the alteration from carbohydrate storage to lipid accumulation in signal transduction process of C. protothecoides.

Optimization of polyhydroxyalkanoates fermentations with on-line capacitance measurement

The aim of this work was to provide an effective methodology for optimization of the polyhydroxyalkanoates (PHAs) fermentation with Ralstonia eutropha by the on-line capacitance measurement. The present study found the capacitance values could reflect variations of microbial morphology and viability. Furthermore, oxygen uptake rate, specific oxygen uptake rate and specific growth rate were measured in real-time and compared with the capacitance value. In addition, a fed-batch control strategy based on the on-line capacitance measurement was proposed to improve the PHAs production by 22%.

A simple novel approach for real-time monitoring of sodium gluconate production by on-line physiological parameters in batch fermentation by Aspergillus niger.

In this paper, approach for real-time monitoring of sodium gluconate (SG) fermentation was established for the first time by the equations which can calculate real-time key-parameters by on-line physiological data. Based on this approach, limiting factors were found out in initial fermentation F1 and then step-wise agitation increase and improved medium recipe were proposed in fermentation F2 and F3, respectively. The highest average SG production rate (16.58±0.91 g L(-1) h(-1)) was achieved in fermentation F3, which was 104.2% and 48.0% higher than those in fermentation F1 and F2, respectively. Meanwhile, due to shorter fermentation period (decreased from 34 h to 18.7 h), lower biomass (about 1.5 g L(-1)) and less by-product accumulation, the overall yield of 0.943±0.012 (mol mol(-1)) in fermentation F3 increased more than 16.0% compared to fermentation F1. This approach had been successfully applied to industrial fermentation and greatly improved SG production.

Dependence of fungal characteristics on seed morphology and shear stress in bioreactors

The fungal morphology during submerged cultivations has a profound influence on the overall performance of bioreactors. In this research, glucoamylase production by Aspergillus niger has been taken as a model to improve more insights. The morphology engineering could be conducted effectively by changing the seed morphology, as well as specific power input. During the fed-batch cultivations, pellet formation under milder shear stress field helped to reduce the broth viscosity, thus relieving oxygen limitation and promoting the enzyme production. Furthermore, we found that the relation between the shear stress field, which was characterized by energy dissipation rate/circulation function (EDCF), and enzyme activity was consistent with quadratic parabola, which threw light on the process optimization and scale-up for industrial enzyme production.

Significant decrease of broth viscosity and glucose consumption in erythromycin fermentation by dynamic regulation of ammonium sulfate and phosphate

In this study, the effects of nitrogen sources on broth viscosity and glucose consumption in erythromycin fermentation were investigated. By controlling ammonium sulfate concentration, broth viscosity and glucose consumption were decreased by 18.2% and 61.6%, respectively, whereas erythromycin biosynthesis was little affected. Furthermore, erythromycin A production was increased by 8.7% still with characteristics of low broth viscosity and glucose consumption through the rational regulations of phosphate salt, soybean meal and ammonium sulfate. It was found that ammonium sulfate could effectively control proteinase activity, which was correlated with the utilization of soybean meal as well as cell growth. The pollets formation contributed much to the decrease of broth viscosity. The accumulation of extracellular propionate and succinate under the new regulation strategy indicated that higher propanol consumption might increase the concentration of methylmalonyl-CoA and propionyl-CoA and thus could increase the flux leading to erythromycin A.

A rapid and accurate quantification method for real-time dynamic analysis of cellular lipids during microalgal fermentation processes in Chlorella protothecoides with low field nuclear magnetic resonance.

The rapid and real-time lipid determination can provide valuable information on process regulation and optimization in the algal lipid mass production. In this study, a rapid, accurate and precise quantification method of in vivo cellular lipids of Chlorella protothecoides using low field nuclear magnetic resonance (LF-NMR) was newly developed. LF-NMR was extremely sensitive to the algal lipids with the limits of the detection (LOD) of 0.0026g and 0.32g/L in dry lipid samples and algal broth, respectively, as well as limits of quantification (LOQ) of 0.0093g and 1.18g/L. Moreover, the LF-NMR signal was specifically proportional to the cellular lipids of C. protothecoides, thus the superior regression curves existing in a wide detection range from 0.02 to 0.42g for dry lipids and from 1.12 to 8.97gL(-1) of lipid concentration for in vivo lipid quantification were obtained with all R(2) higher than 0.99, irrespective of the lipid content and fatty acids profile variations. The accuracy of this novel method was further verified to be reliable by comparing lipid quantification results to those obtained by GC-MS. And the relative standard deviation (RSD) of LF-NMR results were smaller than 2%, suggesting the precision of this method. Finally, this method was successfully used in the on-line lipid monitoring during the algal lipid fermentation processes, making it possible for better understanding of the lipid accumulation mechanism and dynamic bioprocess control.

High efficiency cell-recycle continuous sodium gluconate production by Aspergillus niger using on-line physiological parameters association analysis to regulate feed rate rationally

In this paper, a system of cell-recycle continuous fermentation for sodium gluconate (SG) production by Aspergillus niger (A. niger) was established. Based on initial continuous fermentation result (100.0h) with constant feed rate, an automatic feedback strategy to regulate feed rate using on-line physiological parameters (OUR and DO) was proposed and applied successfully for the first time in the improved continuous fermentation (240.5h). Due to less auxiliary time, highest SG production rate (31.05±0.29gL(-1)h(-1)) and highest yield (0.984±0.067molmol(-1)), overall SG production capacity (975.8±5.8gh(-1)) in 50-L fermentor of improved continuous fermentation increased more than 300.0% compared to that of batch fermentation. Improvement of mass transfer and dispersed mycelia morphology were the two major reasons responsible for the high SG production rate. This system had been successfully applied to industrial fermentation and SG production was greatly improved.

Kinetic analysis of curdlan production by Alcaligenes faecalis with maltose, sucrose, glucose and fructose as carbon sources

Curdlan has wide-ranging benefits in food and pharmaceutical industries for its unique rheological and thermal gelling properties. To analyze the cell growth and curdlan biosynthesis kinetics of Alcaligenes faecalis, the kinetic properties of the curdlan fermentation under different carbon sources conditions (maltose, sucrose, glucose and fructose) were investigated using Logistic and Luedeking-Piret equations. The results demonstrated that curdlan fermentation is partial growth-associated process. With maltose as the sole carbon source, the highest curdlan production (Pm = 39.3 g/L), the maximum specific growth rate (μm = 0.44/h) and the growth-associated rate constant (α = 2.05 g curdlan/g cell) were achieved. In contrast, the fructose was the less desired carbon source in both the cell growth and curdlan production. Further, the results demonstrated that slow-releasing glucose from maltose boosted cell growth and curdlan production.

On structural identifiability analysis of the cascaded linear dynamic systems in isotopically non-stationary 13C labelling experiments

The isotopically non-stationary 13C labelling experiments, as an emerging experimental technique, can estimate the intracellular fluxes of the cell culture under an isotopic transient period. However, to the best of our knowledge, the issue of the structural identifiability analysis of non-stationary isotope experiments is not well addressed in the literature. In this work, the local structural identifiability analysis for non-stationary cumomer balance equations is conducted based on the Taylor series approach. The numerical rank of the Jacobian matrices of the finite extended time derivatives of the measured fractions with respect to the free parameters is taken as the criterion. It turns out that only one single time point is necessary to achieve the structural identifiability analysis of the cascaded linear dynamic system of non-stationary isotope experiments. The equivalence between the local structural identifiability of the cascaded linear dynamic systems and the local optimum condition of the nonlinear least squares problem is elucidated in the work. Optimal measurements sets can then be determined for the metabolic network. Two simulated metabolic networks are adopted to demonstrate the utility of the proposed method.