Evaluation of Medium Components by Plackett-Burman Statistical Design for Lipase Production by Candida rugosa and Kinetic Modeling

Lipase production by Candida rugosa was carried out in submerged fermentation. Plackett-Burman statistical experimental design was applied to evaluate the fermentation medium components. The effect of twelve medium components was studied in sixteen experimental trials. Glucose, olive oil, peptone and FeCl3?6H2O were found to have more significance on lipase production by Candida rugosa. Maximum lipase activity of 3.8 u mL-1 was obtained at 50 h of fermentation period. The fermentation was carried out at optimized temperature of 30oC, initial pH of 6.8 and shaking speed of 120 r/min. Unstructured kinetic models were used to simulate the experimental data. Logistic model, Luedeking-Piret model and modified Luedeking-Piret model were found suitable to efficiently predict the cell mass, lipase production and glucose consumption respectively with high determination coefficient(R2). From the estimated values of the Luedeking-Piret kinetic model parameters, α and β, it was found that the lipase production by Candida rugosa is growth associated.     

产弹性蛋白酶细菌的发酵动力学研究

基于14L的发酵罐分批发酵实验数据,建立了发酵过程菌体生长、产物生成及基质消耗随时间变化的数学模型。Logistic方程、Luedeking—Piret方程能够很好地分别描述产弹性蛋白酶菌体生长;发酵产酶过程和基质消耗过程。并将3个动力学模型的预测值和实验值进行了比较,所建立的分批发酵动力学模型能较好地反映弹性蛋白酶分批发酵过程。     

Modeling of growth, lactate consumption, and volatile fatty acid production by Megasphaera elsdenii cultivated in minimal and complex media

Use of the Pirt and Luedeking-Piret equations permits the determination of the effect of medium composition on the metabolic patterns of Megasphaera elsdenii grown in minimal and complex media with lactate as the major carbon source. To establish the significance of the parameters involved in the Pirt and Luedeking-Piret equations, a quantitative statistical criterion was proposed. In the complex medium, lactate was completely used for growth and product formation, whereas in the minimal medium a fraction of the energy obtained from lactate was used for maintenance purposes. Modeling of VFA production by the Luedeking-Piret equation showed that, independent of the type of medium, acetate and propionate are growth-associated products, while butyrate and valerate are only partially growth-associated. The growth-associated products are related to energy-yielding metabolism and the non-growth-associated products are related to the consumption of reducing equivalents.     

An unstructured kinetic model for the improvement of triterpenes production by Ganoderma lucidum G0119 based on nitrogen source effect

The kinetics of cell growth and triterpenes production for liquid submerged fermentation of the medicinal mushroom Ganoderma lucidum were investigated. A kinetic model was developed based on the Logistic and Luedeking-Piret equations for cell growth, substrate consumption and triterpene formation. The kinetic parameters of the model were optimized by specifically designed Runge-Kutta genetic algorithms. The mathematical model simulated the experimental data well and was capable of explaining the behavior of triterpenes production. The predictions of the kinetics from this model are very good both for normal fermentation kinetics under nitrogen limitation as well as for predictions of transitions to sluggish fermentations. The resulting model is very useful for scaling up liquid submerged fermentation of the mushroom G. lucidum and its application to the industrial production of triterpene.     

Kinetics and modeling of temperature effects on batch xanthan gum fermentation

Batch fermentation kinetics of xanthan gum production from glucose by Xanthomonas campestris at temperatures between 22 degrees C and 35 degrees C were studied to evaluate temperature effects on cell growth and xanthan formation. These batch xanthan fermentations were modeled by the logistic equation for cell growth, the Luedeking-Piret equation for xanthan production, and a modified Luedeking-Piret equation for glucose consumption. Temperature dependence of the parameters in this model was evaluated. Growth-associated rate constants increased to a maximum at approximately 30 degrees C and then decreased to zero at approximately 35 degrees C. This temperature effect can be modeled using a square-root model. On the contrary, non-growth-associated rate constants increased with increasing temperature, following the Arrhenius relationship, in the entire temperature range studied. The model developed in this work fits the experimental data very well and can be used in a simulation study. However, due to the empirical nature of the model, the parameter values need to be reevaluated if the model is to be applied to different growth conditions.     

Modeling and simulation of Streptomyces peucetius var. caesius N47 cultivation and epsilon-rhodomycinone production with kinetic equations and neural networks

This study focuses on comparing different kinetic growth models and the use of neural networks in the batch cultivation of Streptomyces peucetius var. caesius producing epsilon-rhodomycinone. Contois, Monod and Teissier microbial growth models were used as well as the logistic growth modeling approach, which was found best in the simulations of growth and glucose consumption in the batch growth phase. The lag phase was included in the kinetic model with a CO2 trigger and a delay factor. Substrate consumption and product formation were included as Luedeking-Piret and logistic type equations, respectively. Biomass formation was modeled successfully with a 6-8-2 network, and the network was capable of biomass prediction with an R2-value of 0.983. Epsilon-rhodomycinone production was successfully modeled with a recursive 8-3-1 network capable of epsilon-rhodomycinone prediction with an R2-value of 0.903. The predictive power of the neural networks was superior to the kinetic models, which could not be used in predictive modeling of arbitrary batch cultivations.     

Growth and neutral lipid synthesis in green microalgae: A mathematical model

Many green microalgae significantly increased their cellular neutral lipid content when cultured in nitrogen limited or high light conditions. Due to their lipid production potential, these algae have been suggested as promising feedstocks for biofuel production. However, no models for algal lipid synthesis with respect to nutrient and light have been developed to predict lipid production and to help improve the production process. A mathematical model is derived describing the growth dynamics and neutral lipid production of green microalgae grown in batch cultures. The model assumed that as the nitrogen was depleted, photosynthesis became uncoupled from growth, resulting in the synthesis and accumulation of neutral lipids. Simulation results were compared with experimental data for the green microalgae Pseudochlorococcum sp. For growth media with low nitrogen concentration, the model agreed closely with the data; however, with high nitrogen concentration the model overestimated the biomass. It is likely that additional limiting factors besides nitrogen could be responsible for this discrepancy.     

黑曲霉过氧化氢酶发酵过程的数学模型

研究了黑曲霉发酵生产过程氧化氢酶的分批发酵动力学,并建立了发酵过程菌体生长,基质消耗及酶合成的随时间变化的数学模型。Logistic方程,Luedekin-Piret方程及与Luedeking-Piret方程相似的基质消耗方程能够很好地分别描述黑曲霉细胞的生长,发酵产酶过程及葡萄糖的消耗,过氧化氢酶的发酵合成是生长耦联的,研究中还将3个动力学模型的预测值和实验值进行了比较。     

蓝色犁头霉深层发酵产壳低聚糖生长动力学研究

目的:研究蓝色犁头霉产壳低聚糖分批发酵动力学的模型.方法:在10L发酵罐中,将蓝色犁头霉分批发酵培养,对蓝色犁头霉菌丝体生长、产物形成和基质消耗的实验数据进行分析,根据Logistic和Luedeking-Piret方程分别建立蓝色犁头霉发酵过程菌体生长、壳低聚糖生成和基质消耗的动力学数学模型,并利用1stOpt软件对模型参数进行非线性曲线拟合.结果:研究得到了菌体生长、产物合成及基质消耗的动力学模型及参数,模型的拟合度分别为0.994、0.986、0.992.结论:研究表明蓝色犁头霉多糖合成和菌体生长呈生长偶联型,模型计算值与实验值有良好的拟合性,模型准确度较高.     

Evaluating the usability of 19 effluents for heterotrophic cultivation of microalgal consortia as biodiesel feedstock

A key reason inhibiting commercialization of algal oil as biodiesel feedstock, is cultivation cost. For this reason, the usability of 19 readily available industrial effluents (autoclaved and non-autoclaved) to support heterotrophic growth and lipid accumulation was evaluated using six mixed algal cultures. Autoclaved whey effluent was the best with 14.32 g biomass L^?1, 13.23% lipids, resulting in a lipid production of 1.91 g lipids L^?1. Biomass production and lipid accumulation were in many cases inverse, e.g. mixed algal culture termed TUT4 accumulating 84.25% lipids on autoclaved acid mine drainage, with very little biomass produced. Biomass production was dependent on the effluent type, whereas the lipid accumulation was influenced mostly by the specific mixed algal cultures. The fatty acid composition of the algal oil (fish cannery and whey effluents) showed high saturation, leading to acceptable cetane numbers, kinematic viscosity, good oxidative stability, but poor cold flow properties.     

Modeling growth and succinoglucan production by Agrobacterium radiobacter NCIB 9042 in batch cultures

Wild-type Agrobacterium radiobacter NCIB 9042 has been cultivated in batch cultures on a synthetic medium which was adapted for growth and succinoglucan production. Experiments were carried out in a 4-L stirred-tank aerated reactor. Glucose, biomass, polysaccharide, protein, and inorganic- and organic-nitrogen concentrations were measured, and oxygen consumption and CO(2) production rates were obtained by a gas-balance technique. Nitrogen balance shows that inorganic nitrogen is entirely recovered into proteins. The carbon balance is satisfied with in +/-5%. Stoichiometric equations for biomass growth and succinoglucan synthesis were established. The biosyntheticpolymer pathways including ATP and cofactor consumption were investigated. From previous studies, a (P/O) value of 1.66 is selected for oxygen sufficient cultures. The actual ATP requirements of 25.4 mmol ATP/g succinoglucan (38.5 mol ATP/mol succinoglucan), determined by a metabolic analysis, is 2.39 times the stoichiometric value. Experimental results were modeled by a system of differential equations. The exponential growth phase was described by a nitrogen-limited Monod equation. Subsequent succinoglucan synthesis followed a slightly modified Luedeking-Piret relation partitioning internal and external polysaccharide. Experimentally determined coefficients are compared with published results for continuous culture of A. radiobacter NCIB 11883.     

The effect of species diversity on lipid production by micro-algal communities

Current research investigating the importance of diversity for biofuel lipid production remains limited. In contrast, the relationship between diversity and productivity within terrestrial and algal primary producers has been well documented in ecology. Hence, we set out to investigate, experimentally, whether diversity may also affect lipid production in micro-algae. We investigated the growth and lipid production of micro-algae using species from all major algal groups. Algae were grown in a large number of treatments differing in their diversity level. Additionally, we compared the growth and lipid production of laboratory communities to natural lake and pond phytoplankton communities of different diversity. Our results show that lipid production increased with increasing diversity in both natural and laboratory micro-algal communities. The underlying reason for the observed ‘diversity–productivity’ relationship seems to be resource use complementarity. We observed higher lipid production of highly diverse algal communities under the same growth and resource supply conditions compared to monocultures. Hence, the incorporation of the ecological advantages of diversity-related resource-use dynamics into algal biomass production may provide a powerful and cost effective way to improve biofuel production.     

Response surface methodology as a decision-making tool for optimization of culture conditions of green microalgae Chlorella spp. for biodiesel production

We have evaluated process optimization and the interactive effects of a number of variables using a Box–Behnken design of response surface methodology (RSM). The process variables nitrate, phosphate, glucose and pH were optimized to enhance the cell growth rate, lipid accumulation and other biochemical parameters of Chlorella spp. The most significant increase in lipid production (dry cell weight basis) occurred at limited concentrations of nitrate and phosphate, 1 % glucose and pH 7.5. The addition of nitrates during the mid-lag and mid-exponential phases produced the maximum inhibitory effect on lipid accumulation and the presence of yeast extract led to a further enhancement of lipid accumulation. Of all the media tested, BG-11 was the best suited medium for algal biomass production and chlorophyll content. A significant increase in algal biomass was observed in BG-11 supplemented with bicarbonate and glucose (1 %). The maximum specific growth rate observed was on 9th day of culturing. Results of optimization of process variables through response surface methodology and optimization of various other conditions reflect cutting edge research directed towards increasing algal biomass and lipid content for biodiesel production using an efficient economical technological approach.     

Modeling of exo-inulinase biosynthesis by Kluyveromyces marxianus in fed-batch mode: correlating production kinetics and metabolic heat fluxes

A metabolic heat-based model was used for estimating the growth of Kluyveromyces marxianus, and the modified Luedeking-Piret kinetic model was used for describing the inulinase production kinetics. For the first time, a relationship was developed to relate inulinase production kinetics directly to metabolic heat generated, which corroborated well with the experimental data (with R ² values of above 0.9). It also demonstrated the predominantly growth-associated nature of the inulinase production with Luedeking-Piret parameters α and β, having values of 0.75 and 0.033, respectively, in the exponential feeding experiment. MATLAB was used for simulating the inulinase production kinetics which demonstrated the model’s utility in performing real-time prediction of inulinase concentration with metabolic heat data as input. To validate the model predictions, a biocalorimetric (Bio RC1e) experiment for inulinase production by K. marxianus was performed. The inulinase concentration (IU/mL) values acquired from the model in were validated with the experimental values and the metabolic heat data. This modeling approach enabled the optimization, monitoring, and control of inulinase production process using the real-time biocalorimetric (Bio RC1e) data. Gas chromatography and mass spectrometry analysis were carried out to study the overflow metabolism taking place in K. marxianus inulinase production.

     

Functional group richness: implications of biodiversity for light use and lipid yield in microalgae

Currently, very few studies address the relationship between diversity and biomass/lipid production in primary producer communities for biofuel production. Basic studies on the growth of microalgal communities, however, provide evidence of a positive relationship between diversity and biomass production. Recent studies have also shown that positive diversity–productivity relationships are related to an increase in the efficiency of light use by diverse microalgal communities. Here, we hypothesize that there is a relationship between diversity, light use, and microalgal lipid production in phytoplankton communities. Microalgae from all major freshwater algal groups were cultivated in treatments that differed in species richness and functional group richness. Polycultures with high functional group richness showed more efficient light use and higher algal lipid content with increasing species richness. There was a clear correlation between light use and lipid production in functionally diverse communities. Hence, a powerful and cost‐effective way to improve biofuel production might be accomplished by incorporating diversity related, resource‐use‐dynamics into algal biomass production.     

Analysis and modeling of Nannochloropsis growth in lab,greenhouse, and raceway experiments

Efficient production of algal biofuels could reduce dependence on foreign oil by providing a domestic renewable energy source. Moreover, algae-based biofuels are attractive for their large oil yield potential despite decreased land use and natural resource (e.g., water and nutrients) requirements compared to terrestrial energy crops. Important factors controlling algal lipid productivity include temperature, nutrient availability, salinity, pH, and the light-to-biomass conversion rate. Computational approaches allow for inexpensive predictions of algae growth kinetics for various bioreactor sizes and geometries without the need for multiple, expensive measurement systems. Parametric studies of algal species include serial experiments that use off-line monitoring of growth and lipid levels. Such approaches are time consuming and usually incomplete, and studies on the effect of the interaction between various parameters on algal growth are currently lacking. However, these are the necessary precursors for computational models, which currently lack the data necessary to accurately simulate and predict algae growth. In this work, we conduct a lab-scale parametric study of the marine alga Nannochloropsis salina and apply the findings to our physics-based computational algae growth model. We then compare results from the model with experiments conducted in a greenhouse tank and an outdoor, open-channel raceway pond. Results show that the computational model effectively predicts algae growth in systems across varying scale and identifies the causes for reductions in algal productivities. Applying the model facilitates optimization of pond designs and improvements in strain selection.     

Two-stage heterotrophic and phototrophic culture strategy for algal biomass and lipid production

A two-stage heterotrophic and phototrophic culture strategy for algal biomass and lipid production was studied, wherein high density heterotrophic cultures of Chlorellasorokiniana serve as seed for subsequent phototrophic growth. The data showed growth rate, cell density and productivity of heterotrophic C.sorokiniana were 3.0, 3.3 and 7.4 times higher than phototrophic counterpart, respectively. Hetero- and phototrophic algal seeds had similar biomass/lipid production and fatty acid profile when inoculated into phototrophic culture system. To expand the application, food waste and wastewater were tested as feedstock for heterotrophic growth, and supported cell growth successfully. These results demonstrated the advantages of using heterotrophic algae cells as seeds for open algae culture system. Additionally, high inoculation rate of heterotrophic algal seed can be utilized as an effective method for contamination control. This two-stage heterotrophic phototrophic process is promising to provide a more efficient way for large scale production of algal biomass and biofuels.     

The boosted lipid accumulation in microalga <Emphasis Type="Italic">Chlorella vulgaris</Emphasis> by a heterotrophy and nutrition-limitation transition cultivation regime

A model of heterotrophy and nutrition-limitation transition cultivation for efficient algal biomass and lipid production was proposed in this study, wherein sufficient robust heterotrophic-seed cells submitted into nitrogen-starvation induction for boosted lipid accumulation. The results demonstrated that heterotrophic-seed (HS) achieved specific growth rate of 1.35 day^?1 and biomass productivity of 1.93 mg/L/d, representing 6.42- and 32.16-fold, 2.01- and 2.75-fold more than that of photoautotrophic-seed (PS) and mixtrophic-seed (MS). Even though subsequent nutrition-limitation cultivation repressed the growth of HS, the overall lipid productivity caused by nitrogen-starvation was not offset by biomass loss. The most favorable lipid productivity (465.61 mg/L/d) of HS was 3.25 and 52.31 times higher than that of MS and PS. The high content of monounsaturated fatty acids (50.13%) over saturated and polyunsaturated fatty acids (totally 47.39%) in HS cells could provide superior oxidation stability and lower viscosity for biofuels generated from algal biomass feedstock. These findings suggested the feasibility of using heterotrophy and nutrition-limitation transition cultivation for enhancing the overall lipid productivity. Further, several critical enzymes (i.e. G3PDH, ME, and ACAD) were highly related to lipid accumulation and showed especially pronounced up-regulation or down-regulation expression in HS, which provide indications for shedding light on the molecular mechanisms of lipid accumulation and a prospective metabolic engineering for lipid production.     

Kinetics of kojic acid fermentation byAspergillus flavus link S44-1 using sucrose as a carbon source under different pH conditions

Kojic acid production byAspergillus flavus strain S44-1 using sucrose as a carbon source was carried out in a 250-mL shake flask and a 2-L stirred tank fermenter. For comparison, production of kojic acid using glucose, fructose and its mixture was also carried out. Kojic acid production in shake flask fermentation was 25.8 g/L using glucose as the sole carbon source, 23.6 g/L with sucrose, and 6.4 g/L from fructose. Reduced kojic acid production (13.5 g/L) was observed when a combination of glucose and fructose was used as a carbon source. The highest production of kojic acid (40.2 g/L) was obtained from 150 g/L sucrose in a 2 L fermenter, while the lowest kojic acid production (10.3 g/L) was seen in fermentation using fructose as the sole carbon source. The experimental data from batch fermentation and resuspended cell system was analysed in order to form the basis for a kinetic model of the process. An unstructured model based on logistic and Luedeking-Piret equations was found suitable to describe the growth, substrate consumption, and efficiency of kojic acid production byA. flavus in batch fermentation using sucrose. From this model, it was found that kojic acid production byA. flavus was not a growth-associated process. Fermentation without pH control (from an initial culture pH of 3.0) showed higher kojic acid production than single-phase pH-controlled fermentation (pH 2.5, 2.75, and 3.0).     

Effect of iron on growth and lipid accumulation in Chlorella vulgaris

The economic feasibility of algal mass culture for biodiesel production is enhanced by the increase in biomass productivity and storage lipids. Effect of iron on growth and lipid accumulation in marine microalgae Chlorella vulgaris were investigated. In experiment I, supplementing the growth media with chelated FeCl3 in the late growth phase increased the final cell density but did not induce lipid accumulation in cells. In experiment II, cells in the late-exponential growth phase were collected by centrifugation and re-inoculated into new media supplemented with five levels of Fe3+ concentration. Total lipid content in cultures supplemented with 1.2 x 10(-5) mol L(-1) FeCl3 was up to 56.6% biomass by dry weight and was 3-7-fold that in other media supplemented with lower iron concentration. Moreover, a simple and rapid method determining the lipid accumulation in C. vulgaris with spectrofluorimetry was developed.