A multi-objective optimization approach for the production of polyhydroxybutyrate via Chlorogloea fritschii under diurnal light with single-stage cultivation.

The present study aims to optimize the nutrients for maximization of cyanobacterial biomass with high content of polyhydroxybutyrate (PHB), a bioplastic, and recovery of biomass by auto-sedimentation under diurnal light mimic to sunlight. The multi-objective optimization with desirability approach was used to improve dry cell weight (DCW), PHB content (% w/w), and auto-sedimentation concentration factor (SCF) of biomass. Initially, NaNO3, K2HPO4, TRACE (micronutrient solution), Na2EDTA, and MgSO4.7H2O were screened as important media compositions. Screening was followed by the application of response surface methodology for the development of a model used in multi-objective optimization. The optimized media selected from many optimal solutions, a set of Pareto solutions generated by multi-objective optimization was validated in a flat panel photobioreactor. Using a single-stage cultivation strategy under diurnal light, Chlorogloea fritschii TISTR 8527 has shown capability to produce DCW of 1.23 g/l with PHB content of 31.78 % and SCF of 93.63 with optimal media. This leads to the enhancement of both PHB content (2.72 fold) and SCF (1.64 fold) were observed when compared to the non-optimal medium. This is the first multi-objective optimization study for media optimization using cyanobacteria reported till now under diurnal light mimic to sunlight for bioplastic production.

Enhanced production of biosurfactant by Bacillus subtilis RSL2 in semicontinuous bioreactor utilizing molasses as a sole substrate.

The growth-associated metabolites are produced during the exponential phase; however, this phase terminates due to substrate depletion or product inhibition. In the present study, a semicontinuous mode with a fill-and-draw strategy was applied to extend the exponential phase of the biosurfactant production to overcome the product inhibition and in turn, enhance the yield. Bioreactor studies were performed in batch mode, followed by the semicontinuous operation. A potential biosurfactant producer Bacillus subtilis RSL2 was used in this study at the previously optimized conditions of pH 6.6, temperature 41 °C and 5% (w/v) of molasses. A better mass transfer was achieved in the bioreactor as compared to the shake flask study. In the batch bioreactor study, 90% of sugar was utilized with simultaneous 13.7 g L-1 of biosurfactant production. The sugar utilization was further improved to > 98% in the case of semicontinuous operation employing a fill-and-draw strategy. The exponential phase got extended up to 18 days and a total of 13 L of media was fed in the semicontinuous operation of 21 days as compared to 1.5 L of working volume in the batch reactor. The biosurfactant yield was enhanced by 1.5 folds and was found to be 0.97 g g-1. The produced biosurfactant was identified as a lipopeptide. The interfacial properties of the biosurfactant along with colloidal and thermal stability have been investigated. The critical micelle concentration of the produced biosurfactant was 70 mg L-1. The present study highlighted the efficient utilization of molasses for the production of biosurfactant, an alternative metabolite, in a semicontinuous mode of bioreactor.

Biodegradation of waste cooking oil and simultaneous production of rhamnolipid biosurfactant by Pseudomonas aeruginosa P7815 in batch and fed-batch bioreactor.

Biosurfactants are non-toxic, surface-active biomolecules capable of reducing surface tension (ST) and emulsifying interface at a comparably lower concentration than commercial surfactants. Yet, poor yield, costlier substrates, and complex cultivation processes limit their commercial applications. This study focuses on producing biosurfactants by Pseudomonas aeruginosa P7815 in batch and fed-batch bioreactor systems using waste cooking oil (WCO) as the sole carbon source. The batch study showed a 92% of WCO biodegradation ability of P. aeruginosa producing 11 g L-1 of biosurfactant. To enhance this biosurfactant production, a fed-batch oil feeding strategy was opted to extend the stationary phase of the bacterium and minimize the effects of substrate deprivation. An enhanced biosurfactant production of 16 g L-1 (i.e. 1.5 times of batch study) was achieved at a feed rate of 5.7 g L-1d-1 with almost 94% of WCO biodegradation activity. The biosurfactant was characterized as rhamnolipid using Fourier transform infrared spectroscopy (FTIR), and its interfacial characterization showed ST reduction to 29 ± 1 mN m-1 and effective emulsification stability at pH value of 4, temperature up to 40 °C and salinity up to 40 g L-1. The biosurfactant exhibited antibacterial activity with minimum inhibitory concentration (MIC) values of 100 µg mL-1 and 150 µg mL-1 for pathogenic E. hirae and E. coli, respectively. These findings suggest that biodegradation of WCO by P. aeruginosa in a fed-batch cultivation strategy is a potential alternative for the economical production of biosurfactants, which can be further explored for biomedical, cosmetics, and oil washing/recovery applications.

Real time light intensity based carbon dioxide feeding for high cell-density microalgae cultivation and biodiesel production in a bubble column photobioreactor under outdoor natural sunlight.

Outdoor high cell-density microalgae cultivation is highly challenging due to unavailability of appropriate CO2 feeding strategy under diurnal sunlight intensities. Hence, a novel real time light based CO2 feeding strategy was firstly developed under diurnal simulated sunlight (LED) to test on Chlorella sp. in a 10 L scale bubble column photobioreactor. The strategy yielded a biomass titer of 5.12 g L-1 under simulated sunlight, far higher than existing biomass-density and pH-control based CO2 feeding strategies. In outdoor culturing, the proposed feeding strategy yielded high biomass titers of 6.8 and 9.0 g L-1 in growth-phase of two-stage and single-stage lipid induction studies respectively with same biomass productivity of 0.8 g L-1 day-1. Subsequently, two-stage lipid induction strategy of 6.8 g L-1 titer yielded biodiesel productivity of 120 g L-1 day-1, whereas single-stage strategy of 9.0 g L-1 titer was unable to induce lipid. Moreover, specific light availability affects the lipid production.

Production of biodiesel by autotrophic Chlorella pyrenoidosa in a sintered disc lab scale bubble column photobioreactor under natural sunlight.

The main focus of the work is to study Chlorella pyrenoidosa mediated photoautotrophic production of lipid in a bubble column photobioreactor using CO2 as carbon source under natural diurnal outdoor sunlight. The limiting and inhibiting concentrations of CO2 in sparging gas, nitrogen inhibition, reversibility of the CO2 inhibition on growth, and lipid production have been investigated under natural sunlight. A process model coupled with light distribution inside the culture has been developed considering different concentration of dissolved CO2 and urea, repression of nitrogen on lipid production under natural sunlight diurnal in nature in a bubble column reactor. The biomass titer of 4.6 g/L with 10% CO2 has been achieved within 5 days of culture under sunlight. A two stage photoautotrophic lipid production strategy in a sintered disc bubble column photobioreactor under natural sunlight has been developed. 30% (w/w-DCW) lipid within 5 days of lipid induction period has been achieved. The biomass productivity of 0.91 ± 0.01 gm/L/day in growth period with sufficient urea and lipid productivity of 410 ± 12 mg/L/day in last 2 days of urea starvation period have been achieved in outdoor photoautotrophic cultivation under natural sunlight using CO2 as carbon source.

Concomitant production of fatty acid methyl ester (biodiesel) and exopolysaccharides using efficient harvesting technology in flat panel photobioreactor with special sparging system via Scenedesmus abundans.

Current study focusses on the concomitant production of fatty acid methyl ester (FAME, biodiesel) and exopolysaccharides (EPS) from Scenedesmus abundans cell factory in flat panel photobioreactor using cost effective harvesting strategy. Parallel mini and medium scale flat panel photobioreactors (PBRs) with special gas sparging system enabling high gas to liquid mass transfer and efficient mixing were designed. Biomass titer of 6.9 g/l with overall biomass productivity of 1.2 g/l/day was achieved with constant high light intensity of 2162 µE/m2/s in growth phase (134 h) using optimum nutrient concentration. FAME concentration of 1.53 g/l was achieved after 15 days of nitrogen deprivation condition with productivity of 67 mg/l/day. The EPS production of 236 mg/l with a yield of 37 mg/g biomass was achieved. The strain proved its capability to produce multiproducts simultaneously in a single stage PBR by natural autoflocculation harvesting technology.

PSII as an in vivo molecular catalyst for the production of energy rich hydroquinones - A new approach in renewable energy.

One of the pertinent issues in the field of energy science today is the quest for an abundant source of hydrogen or hydrogen equivalents. In this study, phenyl-p-benzoquinone (pPBQ) has been used to generate a molecular store of hydrogen equivalents (phenyl-p-hydroquinone; pPBQH2) from thein vivo splitting of water by photosystem II of the marine cyanobacterium Synechococcus elongatus BDU 70542. Using this technique, 10.8 µmol of pPBQH2 per mg chlorophyll a can be extracted per minute, an efficiency that is orders of magnitude higher when compared to the techniques present in the current literature. Moreover, the photo-reduction process was stable when tested over longer periods of time. Addition of phenyl-p-benzoquinone on an intermittent basis resulted in the precipitation of phenyl-p-hydroquinone, obviating the need for costly downstream processing units for product recovery. Phenyl-p-hydroquinone so obtained is a molecular store of free energy preserved through the light driven photolysis of water and can be used as a cheap and a renewable source of hydrogen equivalents by employing transition metal catalysts or fuel cells with the concomitant regeneration of phenyl-p-benzoquinone. The cyclic nature of this technique makes it an ideal candidate to be utilized in mankind's transition from fossil fuels to solar fuels.

Designing a CO2 supply strategy for microalgal biodiesel production under diurnal light in a cylindrical-membrane photobioreactor.

A cylindrical membrane photobioreactor with high CO2 mass transfer coefficient was designed and installed under customized unidirectional lighting. Combinatorial effect of light and CO2 on the growth of Chlorella sp. FC2 IITG was studied and an optimal CO2 supply without pH control strategy was developed under diurnal light similar to sunlight (17-2000-17 µE m-2 s-1). Unprecedentedly, broad range of saturated light levels (700-1500 µE m-2 s-1), reversible photoinhibition, no pH control requirement and dark-phase growth were noticed altogether in the strain. Under diurnal light, final biomass titer of 5.79 g L-1 and overall biomass productivity of 1.29 g L-1 day-1 were observed. The results were similar to optimal light (1130 µE m-2 s-1) and CO2 (2%) conditions. Subsequently, a highest FAME productivity of 265 mg L-1 day-1 was observed in last two days of lipid induction phase.

Modeling of the separation of inhibitory components from pretreated rice straw hydrolysate by nanofiltration membranes.

The main objective of this work was to remove inhibitors and concentrate sugars in hydrolysates obtained from dilute acid-treated rice straw. The Donnan steric pore flow model (DSPM) was applied for membrane characterization and it captured the membrane transport adequately. The polyamide and polyethylene sulfate nanofiltration membranes of 150 Da molecular weight cut-off showed a separation factor of 3 for acetic acid over glucose and xylose and 7 over cellobiose for a simulated mixture at the optimum pH of 3. A separation factor of 3 was also found for the inhibitors hydroxymethyl furfural, ferulic and vanilic acids over sugars. The concentration of rice straw acid hydrolysate by a volume concentration ratio of 4 increased the concentrations of xylose, glucose, arabinose, cellobiose and inhibitor by 100%, 104%, 93%, 151% and 3%, respectively which indicates the membrane can be used for separating the inhibitors from acid-pretreated rice straw hydrolysate while simultaneously concentrating sugars.

Metabolic flexibility of D-ribose producer strain of Bacillus pumilus under environmental perturbations.

The metabolic reaction rate vector is a bridge that links gene and protein expression alterations to the phenotypic endpoint. We present a simple approach for the estimation of flux distribution at key branch points in the metabolic network by using substrate uptake, metabolite secretion rate, and biomass growth rate for transketolase (tkt) deficient Bacillus pumilus ATCC 21951. We find that the glucose-6-phosphate (G6P) and pseudo catabolic/anabolic branch points are flexible in the D: -ribose-producing tkt deficient strain of B. pumilus. The normalized flux through the pentose phosphate pathway (PPP) varied from 1.5 to 86 % under different growth conditions, thereby enabling substantial extracellular accumulation of D: -ribose under certain conditions. Interestingly, the flux through PPP was affected by the extracellular phosphate concentration and dissolved oxygen concentration. This metabolic flexibility may have been the underlying reason for this strain being selected from thousands of others in a screening for D: -ribose producers conducted in the 1970s.

Multi-objective optimization of glycopeptide antibiotic production in batch and fed batch processes.

Fermentation optimization involves potentially conflicting multiple objectives such as product concentration and production media cost. Simultaneous optimization of these objectives would result in a multiobjective optimization problem, which is characterized by a set of multiple solutions, knows as pareto optimal solutions. These solutions gives flexibility in evaluating the trade-offs and selecting the most suitable operating policy. Here, ε-constraint approach was used to generate the pareto solutions for two objectives: product concentration and product per unit cost of media, for batch and fed batch operations using process model for Amycolatopsis balhimycina, a glycopeptide antibiotic producer. This resulted in a set of several pareto optimal solutions with the two objectives ranging from (0.75 g l(-1), 3.97 g $(-1)) to (0.44 g l(-1), 5.19 g $(-1)) for batch and from (1.5 g l(-1), 5.46 g $(-1)) to (1.1 g l(-1), 6.34 g $(-1)) for fed batch operations. One pareto solution each for batch and for fed batch mode was experimentally validated.

Substrate uptake, phosphorus repression, and effect of seed culture on glycopeptide antibiotic production: process model development and experimental validation.

Actinomycetes, the soil borne bacteria which exhibit filamentous growth, are known for their ability to produce a variety of secondary metabolites including antibiotics. Industrial scale production of such antibiotics is typically carried out in a multi-substrate medium where the product formation may experience catabolite repression by one or more of the substrates. Availability of reliable process models is a key bottleneck in optimization of such processes. Here we present a structured kinetic model to describe the growth, substrate uptake and product formation for the glycopeptide antibiotic producer strain Amycolatopsis balhimycina DSM5908. The model is based on the premise that the organism is an optimal strategist and that the various metabolic pathways are regulated via key rate limiting enzymes. Further, the model accounts for substrate inhibition and catabolite repression. The model is also able to predict key phenomena such as simultaneous uptake of glucose and glycerol but with different specific uptake rates, and inhibition of glycopeptide production by high intracellular phosphate levels. The model is successfully applied to both production and seed medium with varying compositions and hence has good predictive ability over a variety of operating conditions. The model parameters are estimated via a well-designed experimental plan. Adequacy of the proposed model was established via checking the model sensitivity to its parameters and confidence interval calculations. The model may have applications in optimizing seed transfer, medium composition, and feeding strategy for maximizing production.

Determination of kinetic parameters in the biosorption of Cr (VI) on immobilized Bacillus cereus M(1)(16) in a continuous packed bed column reactor.

Due to technological advancement, environment suffers from untreated toxic heavy metal bearing effluent coming from different industries. Chromium (VI) is one of those heavy metals having adverse impact on ecological balance, human, and plant health because of its carcinogenic properties. Biosorption is presented as an alternative to traditional technologies which are costly and inefficient for treatment of industrial wastes containing low amount of heavy metals. In this study, bioremediation of Cr (VI) ions by immobilized Bacillus cereus M(1) (16) was investigated in a laboratory scale packed bed up-flow column reactor. The effect of important parameters, such as the inlet flow rate, influent concentration, and effective bed height, has been studied. External mass transfer, surface adsorption, and intrabead mass transfer were also studied to conclude the rate limiting step for removal of Cr (VI) and to determine the process parameters which are important for biosorption optimization. The external mass transfer coefficient was calculated at different flow rates (6.51 x 10(-2) to 7.58 x 10(-2) cm/min). Using the model, the surface adsorption rate constant (k(ad)) and the intrabead mass transfer coefficient (k (i)) were predicted as 0.0267 x 10(-3) and 0.7465 x 10(-3) l/g/min, respectively. Both are much lower than the external mass transfer coefficient (k(e)). The surface adsorption phenomenon is acting as the rate-limiting step due to its high resistance for removal of Cr (VI).