Software sensor design considering oscillating conditions as present in industrial scale fed‐batch cultivations

In this paper, an approach for the monitoring of biotechnological process kinetics is proposed. The kinetics of each process state variable is presented as a function of two time-varying unknown parameters. For their estimation, a general software sensor is derived with on-line measurements as inputs that are accessible in practice. The stability analysis with a different number of inputs shows that stability can be guaranteed for fourth- and fifth-order software sensors only. As a case study, the monitoring of the kinetics of processes carried out in stirred tank reactors is investigated. A new tuning procedure is derived that results in a choice of only one design parameter. The effectiveness of the proposed procedure is demonstrated with experimental data from Bacillus subtilis fed-batch cultivations.

Higher fungi are a promising source of new bioactive natural products with great structural diversity, ranging from polyketides to terpenoids and alkaloids. Many of these structures are a challenge for synthetic organic chemists who have been inspired to develop new total syntheses. Nevertheless, reviews covering syntheses of fungal natural products are lacking. One aim of this review is to close this gap and to present a selection of more recent syntheses of fungal secondary metabolites from different structural classes. Secondly, this review intends to demonstrate that there are many more motivations for the synthesis of natural products than to confirm a proposed structure or to report the first total synthesis of a new natural product. Thirdly, this review is intended to stimulate the interest of organic chemists in the synthesis of fungal natural products. 1 History and Importance of (Fungal) Secondary Metabolites 2 Reasons for the Synthesis of Fungal Natural Products 3 Total Syntheses of Selected Fungal Natural Products 3.1 Terpenes 3.1.1 Sesquiterpenes 3.1.1.1 Polyquinanes 3.1.1.2 Drimanes 3.1.2 Diterpenes: Cyathanes 3.1.3 Terpenes Containing a Butenolide Moiety 3.2 Alkaloids 3.3 Natural Products Derived from the Shikimate Pathway 3.3.1 Pulvinic Acid Derivatives and Terphenyl Quinones 3.3.2 Shikimate Pathway Derived Macrolides 3.3.3 Strobilurins 3.4 Aromatic Polyketides 3.4.1 Quinones 3.4.2 Resorcylates and Catechols 3.5 Sugar Derivatives 4 Conclusion and Outlook

Bioprocess scale-down simulators are used to investigate the phenomena occurring in industrial scale cultivations. The aim is to simulate the large-scale conditions, characterized by long mixing times introducing local gradients into the bioreactor, in laboratory scale bioreactor. We have expanded the current physiological knowledge about the consequences of such cultivation conditions from well-known mixed-acid fermentation to amino acid metabolism. Several interesting deviations were observed for amino acids derived from pyruvate when a STR-PFR scale-down simulator and a regular STR fed-batch cultivation were compared. In this article we show that the cultivation of Escherichia coli K-12 substrain W3110 in a scale-down two-compartment bioreactor has consequences on the free amino acid pools. Repeated occurrence of high glucose zones affects the biosynthesis of amino acids derived from pyruvate. This is interesting since the pyruvate based amino acids are precursors of the branched chain amino acid pathway, which is further disturbed by an unbalanced regulation caused by a frameshift mutation in the ilvG gene of E. coli K-12 strains. As a consequence the formation of non-typical amino acid norvaline was detected in the two compartment bioreactor cultivation.

Hairy roots have been identified as a good source of secondary metabolites in plants. These secondary metabolites in the genera of phytochemicals have been used by humans since long in the form of drugs, flavors, colors, and others. Thereby, large-scale culture of hairy roots, its management, and production have been conferred as most important and critical steps at industrial scale. Conversely, culture of hairy roots in bioreactors at industrial scale has proven to be a tedious job and requires continuous monitoring and precise control of the system. These challenges for hairy roots owe to their heterogeneous nature. Conventional methods for monitoring of such cultures have failed to work well within this system. So, indirect methods are being used for continuous monitoring of growth and metabolite content in hairy roots. Efficiency and efficacy of these indirect methods depend largely upon models of hairy root growth, product synthesis, and substrate utilization. Several mathematical and computational models have been developed to explain hairy root growth. Some of these models are complex mathematical equations which are based on physical principles, while others are computational models derived from empirical data. This chapter intends to outline and explain some of the prominent models for hairy root growth and their mode and mechanism of action in large-scale bioreactors.

For an industrial fermentation process, it can be advantageous to decouple cell growth from product formation. This decoupling would allow for the rapid accumulation of biomass without inhibition from product formation, after which the fermentation can be switched to a mode where cells would grow minimally and primarily act as catalysts to convert substrate into desired product. The switch in fermentation mode should preferably be accomplished without the addition of expensive inducers. A common cell factory Saccharomyces cerevisiae is a Crabtree-positive yeast and is typically fermented at industrial scale under glucose-limited conditions to avoid the formation of ethanol. In this work, we aimed to identify and characterize promoters that depend on glucose concentration for use as dynamic control elements. Through analysis of mRNA data of S. cerevisiae grown in chemostats under glucose excess or limitation, we identified 34 candidate promoters that strongly responded to glucose presence or absence. These promoters were characterized in small-scale batch and fed-batch cultivations using a quickly maturing rapidly degrading green fluorescent protein yEGFP3-Cln2PEST as a reporter. Expressing 3-hydroxypropionic acid (3HP) pathway from a set of selected regulated promoters allowed for suppression of 3HP production during glucose-excess phase of a batch cultivation with subsequent activation in glucose-limiting conditions. Regulating the 3HP pathway by the ICL1 promoter resulted in 70% improvement of 3HP titer in comparison to PGK1 promoter.

An unstructured macrokinetic model for heterologous production of Rhizopus oryzae lipase (ROL) by a Pichia pastoris PAOX1 based system was developed. Mean specific rates and state variables for various fed-batch cultures, under methanol limited and non-limited conditions were used for modeling. The most representative kinetic functions (viz., Monod, Haldane, Pirt and Luedeking-Piret) were used for the specific rates of cell growth (µ), substrate consumption (qS ) and product formation (qP ). The performance of two different process models was assessed via simulation tests, using an overall validation procedure. The best model describes the cellular growth by a non-monotonic substrate function, substrate uptake via Pirt's equation and product formation through a Luedeking-Piret equation. A comparative meta-analysis of heterologous protein production of various target proteins by Pichia pastoris under AOX1 promoter was conducted and a general strategy for improving protein production from process kinetics was developed as a key to bioprocess optimization.

In this paper, experimental results of the control of the ammonia concentration during the growth phase of a PHB producing process in a fedbatch reactor are presented. The control design is based on the mass balance dynamical equations and does require any knowledge of the process kinetics. It results in an adaptive linearizing controller where the specific growth rate is estimated online. Moreover the value of the controlled variable, the ammonian concentration is given by a software sensor via an asymptotic observer based on the online measurement of the optical density. >

In Textile industry, production is mostly key concern for Industry owner. This always has attracted researchers and machines manufacturers to make new developments in process and machines. Air-jet is one of the leading and successful highest productive weaving machines. However, it is now well established that due to add of charges of compressed air, manufacturing cost of air-jet weaving machine is higher as compared with rapier and projectile weaving machines. This is why countries having energy issues do not prefer air-jet weaving machines comparing projectile weaving machines. In this regard, several researchers and machine manufacturers have continuously been working to improve the efficiency of air-jet weft insertion. However, industry practice is as important as design made by researchers. The aim of this research is to investigate the air consumption of air-jet weaving on industrial scale practice. In this study, five weaving machine of same manufacturer and model were selected. It was observed that despite of manufacturing same quality of fabric, air consumption was varying almost in all weaving machines. Conventionally, mill workers adopt hit and trial practice in weaving industry including airpressure setting which leads to variation of nozzle pressure. Main reason of disparity of air consumption in air-jet weaving machines may be variation of distance from compressor to weaving machines, number of joints, un-necessary valve opening and pipes leakages cause an increase of compressed air consumption.

The use of graphene as a filler in thermoplastics has already been investigated extensively. The mechanical properties as well as electrical and thermal conductivity of thermoplastics can be improved due to graphene. However, these studies were carried out in experimental scale, which allows a good dispersion of graphene because of a long residence time during melt mixing and because of the use of almost ideal graphene, which has a high specific surface and low number of layers. In this study the scientific findings are transferred into industrial practice. For that purpose, a co-rotating intermeshing twin screw extruder with limited residence time and limited shear energy input is used to produce graphene based polyamide 6 composites and short carbon fibre reinforced graphene based polyamide 6 composites. These composites are further processed by injection molding to produce specimens in order to determine the mechanical properties. In addition to the scale-up to industrial production, two types of graphene platelets are used, which are commercially available. This investigation reveals that the addition of 1 wt.-% commercially available graphene platelets to polyamide 6 improves Younǵs modulus and tensile strength. Compared to pure polyamide 6 an improvement up to 20 % with regard to Younǵs modulus and up to 15 % with regard to tensile strength can be achieved. The combination of short carbon fibre with graphene platelets in polyamide 6 enables an increase of Younǵs modulus, which is higher than the additive effect.

Among all non-thermal technologies used as a pre-treatment to enhance drying process pulsed electric field (PEF) seems to exhibit the biggest potential. Moreover, commercial success of PEF application in potato processing shows that it is possible to use it as a support for traditional unit operations on an industrial scale. Literature of the subject provides evidence that due to the electroporation phenomenon, PEF treatment ruptures the cellular structure and thus intensifies water removal in the processes of drying or rehydration. The modification of microstructure impacts on the progress of freezing and thus modifies also the freeze-drying progress. Shorter drying or freeze-drying time implies also quality alterations in PEF pre-treated material. However, the changes of physical and chemical properties of PEF pre-treated dried foods depend on many factors related to both product and process. Thus, the positive effect of PEF on both quality and kinetics may require adjustment of the drying parameters. This chapter presents and discusses the effect of PEF applied prior to dehydration on process kinetics and food product quality.KeywordsPulsed electric fieldsDryingOsmotic dehydrationDewateringBioactive compoundsQualityElectroporation

Biodiesel, an alternative diesel fuel made from renewable sources such as vegetable oils and animal fats, is becoming prominent among alternatives to conventional petro-diesel due to economic, environmental and social factors. Transesterification is the most preferred method of biodiesel production. Knowledge of transesterification reaction kinetic enables prediction of the extent of the chemical reaction at any time under particular conditions. It is also essential in the design of reactors for biodiesel production in industrial scale, determination of kinetic model and optimization of operation conditions. In this study, a mathematical model for the microwave assisted trans-esterification reaction of microalgae and methanol has been developed to study the effect of the operating parameters on the process kinetics. A well-mixed microwave reactor was used to express the laboratory scale microwave reactor at stirring speed 500 rpm. Mass transfer controlled state was assumed to be minimal using the stirring condition. The model developed was based on experimental data described in a previous study. The experimental works were designed to study the effect of reaction time between 1-5 min; power of microwave of 100-400 W, and an amount of CaO catalyst of 1 and 3%. The use of a solid catalyst effectively reduces the purification cost of biodiesel due to ease of separation and potential for reuse. The molar ratio of microalgae oil and methanol was constant at the ratio of 1: 6. The validation of model indicated that the reaction have second order reaction in terms of triglycerides. A very good correlation between model and experiment data was observed by correlation coefficient (R2) and least square curve fit. In addition, the experiment shows that the best conditions for reaction time were 5 min, power of microwave was 400 W and amount of CaO catalyst was 3%. The maximum yield of biodiesel in the best conditions was 93.23%.

The growing, existing demand for low-cost and high-quality hyaluronic acid (HA) needs an outlook of different possible production strategies from renewable resources with the reduced possibility of cross-infections. Recently, the possibility of producing HA from harmless microorganisms appeared, which offers the opportunity to make HA more economical, without raw material limitations, and environmentally friendly. HA production is mainly reported with Lancefield Streptococci A and C, particularly from S. equi and S. zooepidemicus. Various modes of fermentation such as batch, repeated batch, fed-batch, and continuous culture have been investigated to optimize HA production, particularly from S. zooepidemicus, obtaining a HA yield of 2.5 g L−1 – 7.0 g L−1. Among the different utilized DSP approaches of HA production, recovery with cold ethanol (4°C) and cetylpyridinium chloride is the ideal strategy for lab-scale HA production. On the industrial scale, besides using isopropanol, filtration (0.22 um), ultrafiltration (100 kDa), and activated carbon absorption are employed to obtain HA of low molecular weight and additional ultrafiltration to purify HA of higher MW. Even though mature technologies have already been developed for the industrial production of HA, the projections of increased sales volume and the expansion of application possibilities require new processes to obtain HA with higher productivity, purity, and specific molecular weights. In this review, we have put forth the progress of HA technological research by discussing the microbial biosynthetic aspects, fermentation and downstream strategies, industrial-scale scenarios of HA, and the prospects of HA production to meet the current and ongoing market demands.

Microbial production of compounds valuable to a myriad of sectors, including chemical, pharmaceutical, nutraceutical, and agricultural industries, has been recognized as a promising strategy for more sustainable sourcing. This approach offers higher flexibility for the synthesis of novel products compared to the traditional chemical route. Furthermore, if researchers aim to succeed at establishing bioproduction at the industrial scale, larger titers of target molecules need to be achieved. Consequently, for organisms to be able to divert their resources towards a desired product, non-trivial manipulation of the cellular metabolism needs to be performed. For a microbial cell factory to be efficient, in addition to the adequate accommodation of the native metabolism for its demanded task, the metabolic flux in a heterologous pathway driving the synthesis of the desired compound needs to be regulated and this is usually accomplished through the variation of familiar genetic parts. However, researchers are aware that the approximately 6000 genes which make every biological function possible in yeast only correspond to the 70% of the genetic information, which opens the door for investigation of the hidden occurrences in the sequence space beyond the characterized genomic parts. In this work, we aim to improve our understanding of noncoding sequences and their contribution in the performance of yeast microbial cell factories. We investigated the impact that context at the sequence level could have on pathway performance. Randomization of the upstream and downstream context of the transcriptional units of the cellobiose pathway in the model yeast Saccharomyces cerevisiae resulted in a more efficient utilization of sugar, indicating an increase in pathway activity which was later verified through transcriptional assays. More intriguingly, further characterization of the sequence context hinted to the potential regulation due to changes in the local organization of DNA, i.e., nucleosome organization. This strategy to diversify expression could be easily applied to other industrially relevant pathways for which screening procedures are in place. Comprehensive understanding of metabolism, abundant tools for genetic manipulation and suitability for industrial processes are a few of the advantages why S. cerevisiae has been traditionally at the front of efforts to construct superior strains. However, it has become clear that the prospect of using less characterized organisms might be worthy, especially when target molecules are less taxing for these organisms to produce due to their inherent metabolic capabilities, compared to model organisms. Yarrowia lipolytica is a nonconventional yeast species that has gained popularity in recent years due to its biotechnological potential and its characteristic feature of lipid accumulation. Further exploration of noncoding sequences as a subject study for this species materialized upon the inspection of a previously disregarded piece of DNA found in the unique structure that autonomous replicating sequences (ARSs) takes in this yeast. Development of stable vectors is a prerequisite for a novel species to be adopted and we found that selection of ARS not only impacts pathway performance but also vector stability. Among the collection of ARSs we evaluated, it was demonstrated that native sequences are more beneficial for bioproduction compared to their synthetic counterparts. Finally, researchers choosing to employ Y. lipolytica as a host should consider the use of the wildtype ARS1 sequence rather than their minimal version ORI1001-CEN1-1 for improved vector maintenance.

Over the past decade, the production of angiotensin–converting enzyme (ACE) inhibitor drug intermediates has become increasingly important in the pharmaceutical industry. The most significant clinical application of ACE inhibitors is in the management of hypertension and cardiovascular diseases, particularly congestive heart failure (CHF). A number of chemical methods have been reported thus far for the synthesis of ACE inhibitor precursors. However, chemical methods generally suffer from process complexity, high cost, and environmental pollution. On the other hand, enantiomerically pure ACE inhibitor drug precursors can be obtained elegantly and efficiently by employing biocatalytic methods, where it appears to be the most attractive process in terms of potential industrial applications and green chemistry. In the recent past, membranes are expeditiously used in the enzyme–catalyzed synthesis of optically active pharmaceutical compounds on laboratory and industrial scales, being incorporated in membrane bioreactor system. This technique can be applied for the repeated use of the enzymes in the synthesis via retention or immobilization, especially when dealing with costly biocatalysts. Herein we review the enzymatic synthesis of vital ACE inhibitor drug precursors as potentially useful intermediates in the synthesis of pharmaceutical products, in environmentally friendly condition, using membrane bioreactor for biotransformation.

N-Acetyllactosamine (LacNAc) or more specifically β-d-galactopyranosyl-1,4-N-acetyl-d-glucosamine is a unique acyl-amino sugar and a key structural unit in human milk oligosaccharides, an antigen component of many glycoproteins, and an antiviral active component for the development of effective drugs against viruses. LacNAc is useful itself and as a basic building block for producing various bioactive oligosaccharides, notably because this synthesis may be used to add value to dairy lactose. Despite a significant amount of information in the literature on the benefits, structures, and types of different LacNAc-derived oligosaccharides, knowledge about their effective synthesis for large-scale production is still in its infancy. This work provides a comprehensive analysis of existing production strategies for LacNAc and important LacNAc-based structures, including sialylated LacNAc as well as poly- and oligo-LacNAc. We conclude that direct extraction from milk is too complex, while chemical synthesis is also impractical at an industrial scale. Microbial routes have application when multiple step reactions are needed, but the major route to large-scale biochemical production will likely lie with enzymatic routes, particularly those using β-galactosidases (for LacNAc synthesis), sialidases (for sialylated LacNAc synthesis), and β-N-acetylhexosaminidases (for oligo-LacNAc synthesis). Glycosyltransferases, especially for the biosynthesis of extended complex LacNAc structures, could also play a major role in the future. In these cases, immobilization of the enzyme can increase stability and reduce cost. Processing parameters, such as substrate concentration and purity, acceptor/donor ratio, water activity, and temperature, can affect product selectivity and yield. More work is needed to optimize these reaction parameters and in the development of robust, thermally stable enzymes to facilitate commercial production of these important bioactive substances.