Showing papers on "Bioprocess published in 2017"

Recent advances in second generation bioethanol production: An insight to pretreatment, saccharification and fermentation processes

Abstract: In response to the scarcity of non-renewable energy sources, sustainable and renewable biofuels from biomass have gained utmost attention. Utilization of lignocellulosic biomass for production of varied energy forms (second generation fuels) like biogas, biodiesel, bioethanol, etc has increased in the past decade. Their properties of being naturally abundant and easily accessible throughout the year, makes them an attractive energy alternative. Efficient pretreatment techniques for effective transformation of lignocelluloses to varied products, by increasing digestibility of celluloses and hemicelluloses can be achieved through acid, alkali treatment, enzymatic hydrolysis, and steam explosion. Idea behind optimizing pretreatment protocol is to maximize release of monosaccharide sugars for conversion to value added products. Tailoring of hydrolytic enzymes through various approaches is well accepted for increasing specific activity of particular enzymatic reaction and can also be clubbed with other pretreatment processes minimizing chemical usage. We at our laboratory are working on optimization of process parameters for enhancing efficiency of saccharification process to obtain maximal monosaccharide sugars that can be converted to bioethanol. Present review compiles various approaches made by prominent scientists for efficient utilization of celluloses and hemicelluloses for ethanol production and also describes recent advanced techniques utilized for the same. Greater emphasis has been led on comparative study on utilization of simple sugars by bacteria and fungi and effect of consolidated bioprocess system on ethanol production from varied agro-industrial wastes.

Food waste: a potential bioresource for extraction of nutraceuticals and bioactive compounds

Abstract: Food waste, a by-product of various industrial, agricultural, household and other food sector activities, is rising continuously due to increase in such activities. Various studies have indicated that different kind of food wastes obtained from fruits, vegetables, cereal and other food processing industries can be used as potential source of bioactive compounds and nutraceuticals which has significant application in treating various ailments. Different secondary metabolites, minerals and vitamins have been extracted from food waste, using various extraction approaches. In the next few years these approaches could provide an innovative approach to increase the production of specific compounds for use as nutraceuticals or as ingredients in the design of functional foods. In this review a comprehensive study of various techniques for extraction of bioactive components citing successful research work have been discussed. Further, their efficient utilization in development of nutraceutical products, health benefits, bioprocess development and value addition of food waste resources has also been discussed.

Design aspects of solid state fermentation as applied to microbial bioprocessing

Abstract: Solid state fermentation (SSF) refers to the microbial fermentation, which takes place in the absence or near absence of free water, thus being close to the natural environment to which the selected micro organisms, especially fungi, are naturally adapted. SSF has been used in the world for a long time. This technology is commonly known in the East, for traditional manufacture of fermented foods, and in the west, for mould-ripened cheese. It can be defined as a system, in which the growth of selected microorganism(s) occurs on solid materials with low moisture contents and has been identified as a potentially important methodology and technique in biotechnology. Nowadays, SSF is an economically viable, practically acceptable technology for large-scale bioconversion and biodegradation processes. Development of sustainable SSF and bioprocess technology is an emerging, multidisciplinary field with possible application to the production of enzymes, chemicals, bioethanol and pharmaceuticals. SSF offers many advantages over conventional submerged fermentation (SMF) such as, simple and inexpensive substrates, elimination of the need for solubilisation of nutrient from within solid substrates, elimination of the need for rigorous control of many parameters during fermentation, product yields are mostly higher, lower energy requirements, produce less waste water, no foam generation and relatively easy recovery of end products. SSF provides flexibility in terms of the raw materials to be used and their capability to produce various value-added products.

Spectroscopic sensors for in-line bioprocess monitoring in research and pharmaceutical industrial application

Abstract: The use of spectroscopic sensors for bioprocess monitoring is a powerful tool within the process analytical technology (PAT) initiative of the US Food and Drug Administration. Spectroscopic sensors enable the simultaneous real-time bioprocess monitoring of various critical process parameters including biological, chemical, and physical variables during the entire biotechnological production process. This potential can be realized through the combination of spectroscopic measurements (UV/Vis spectroscopy, IR spectroscopy, fluorescence spectroscopy, and Raman spectroscopy) with multivariate data analysis to obtain relevant process information out of an enormous amount of data. This review summarizes the newest results from science and industry after the establishment of the PAT initiative and gives a critical overview of the most common in-line spectroscopic techniques. Examples are provided of the wide range of possible applications in upstream processing and downstream processing of spectroscopic sensors for real-time monitoring to optimize productivity and ensure product quality in the pharmaceutical industry.

Methanol-Independent Protein Expression by AOX1 Promoter with trans-Acting Elements Engineering and Glucose-Glycerol-Shift Induction in Pichia pastoris.

Abstract: The alcohol oxidase 1 promoter (PAOX1) of Pichia pastoris is commonly used for high level expression of recombinant proteins. While the safety risk of methanol and tough process control for methanol induction usually cause problems especially in large-scale fermentation. By testing the functions of trans-acting elements of PAOX1 and combinatorially engineering of them, we successfully constructed a methanol-free PAOX1 start-up strain, in which, three transcription repressors were identified and deleted and, one transcription activator were overexpressed. The strain expressed 77% GFP levels in glycerol compared to the wide-type in methanol. Then, insulin precursor (IP) was expressed, taking which as a model, we developed a novel glucose-glycerol-shift induced PAOX1 start-up for this methanol-free strain. A batch phase with glucose of 40 g/L followed by controlling residual glucose not lower than 20 g/L was compatible for supporting cell growth and suppressing PAOX1. Then, glycerol induction was started after glucose used up. Accordingly, an optimal bioprocess was further determined, generating a high IP production of 2.46 g/L in a 5-L bioreactor with dramatical decrease of oxygen consumption and heat evolution comparing with the wild-type in methanol. This mutant and bioprocess represent a safe and efficient alternative to the traditional glycerol-repressed/methanol-induced PAOX1 system.

Application of metabolic controls for the maximization of lipid production in semicontinuous fermentation.

Abstract: Acetic acid can be generated through syngas fermentation, lignocellulosic biomass degradation, and organic waste anaerobic digestion. Microbial conversion of acetate into triacylglycerols for biofuel production has many advantages, including low-cost or even negative-cost feedstock and environmental benefits. The main issue stems from the dilute nature of acetate produced in such systems, which is costly to be processed on an industrial scale. To tackle this problem, we established an efficient bioprocess for converting dilute acetate into lipids, using the oleaginous yeast Yarrowia lipolytica in a semicontinuous system. The implemented design used low-strength acetic acid in both salt and acid forms as carbon substrate and a cross-filtration module for cell recycling. Feed controls for acetic acid and nitrogen based on metabolic models and online measurement of the respiratory quotient were used. The optimized process was able to sustain high-density cell culture using acetic acid of only 3% and achieved a lipid titer, yield, and productivity of 115 g/L, 0.16 g/g, and 0.8 g⋅L-1⋅h-1, respectively. No carbon substrate was detected in the effluent stream, indicating complete utilization of acetate. These results represent a more than twofold increase in lipid production metrics compared with the current best-performing results using concentrated acetic acid as carbon feed.

Trends and advances in conversion of lignocellulosic biomass to biobutanol: Microbes, bioprocesses and industrial viability

Abstract: Biobutanol has gained attention as an alternative renewable transportation fuel for its superior fuel properties and widespread applications in chemical industry, primarily as a solvent. Conventional butanol fermentation has drawbacks that include strain degeneration, end-product toxicity, by-product formation, low butanol concentrations and high substrate cost. The complexity of Clostridium physiology and close control between sporulation phase and ABE fermentation has made it demanding to develop industrially potent strains. In addition to the isolation and engineering of superior butanol producing bacteria, the development of advanced cost-effective technologies for butanol production from feedstock like lignocellulosic biomass has become the primary research focus. High process costs associated with complex feedstocks, product toxicity and low product concentrations are few of the several bioprocess challenges involved in biobutanol production. The article aims to assess the challenges in lignocellulosic biomass to biobutanol conversion and identify key process improvements that can make biobutanol commercially viable.

Genetic modification: a tool for enhancing cellulase secretion

Abstract: Lignocellulosic (LC) biomass is abundantly available as a low-cost resource on the Earth. LC conversion into energy carriers is the most accepted alternative energy production policy because it is non-competitor to food or feed. LC ethanol has brought cellulases to the forefront which was otherwise lost in oblivion during last decades. LC biomass can be converted into value added products or into sugars by various routes, e.g., thermo-chemical, chemical, or biological methods. Biological route via enzymes is one of the most eco-friendly and feasible method. Both fungi and bacteria are known to degrade biomass. Fungi have been greatly exploited for cellulase production due to their inherent properties of secreting extracellular cellulase. These microorganisms are known as cellulase producers for many decades, however, to bring the enzymatic biomass conversion to an economically feasible status, extensive research efforts have been made in last decade to enhance cellulase titers. Mutations and genetic interventions along with bioprocess development have played a very important role for enhancing cellulase production. This review will present a critical overview of the on-going research towards improving cellulase production for biofuel industry via genetic modification, which will include mutation and genetic engineering employed to exert changes at genetic level in microorganisms.

Bioprocess microfluidics: applying microfluidic devices for bioprocessing.

Abstract: Scale-down approaches have long been applied in bioprocessing to resolve scale-up problems. Miniaturized bioreactors have thrived as a tool to obtain process relevant data during early-stage process development. Microfluidic devices are an attractive alternative in bioprocessing development due to the high degree of control over process variables afforded by the laminar flow, and the possibility to reduce time and cost factors. Data quality obtained with these devices is high when integrated with sensing technology and is invaluable for scale-translation and to assess the economical viability of bioprocesses. Microfluidic devices as upstream process development tools have been developed in the area of small molecules, therapeutic proteins, and cellular therapies. More recently, they have also been applied to mimic downstream unit operations.

A modular and adaptive mass spectrometry-based platform for support of bioprocess development toward optimal host cell protein clearance

Abstract: A modular and adaptive mass spectrometry (MS)-based platform was developed to provide fast, robust and sensitive host cell protein (HCP) analytics to support process development. This platform reli...

Heterogeneity in Pure Microbial Systems: Experimental Measurements and Modeling.

Abstract: Cellular heterogeneity influences bioprocess performance in ways that until date are not completely elucidated. In order to account for this phenomenon in the design and operation of bioprocesses, reliable analytical and mathematical descriptions are required. We present an overview of the single cell analysis, and the mathematical modeling frameworks that have potential to be used in bioprocess control and optimization, in particular for microbial processes. In order to be suitable for bioprocess monitoring, experimental methods need to be high throughput and to require relatively short processing time. One such method used successfully under dynamic conditions is flow cytometry. Population balance and individual based models are suitable modeling options, the latter one having in particular a good potential to integrate the various data collected through experimentation. This will be highly beneficial for appropriate process design and scale up as a more rigorous approach may prevent a priori unwanted performance losses. It will also help progressing synthetic biology applications to industrial scale.

Response surface methodology: A non-conventional statistical tool to maximize the throughput of Streptomyces species biomass and their bioactive metabolites.

Abstract: Among diverse actinobacteria, Streptomyces is a renowned ongoing source for the production of a large number of secondary metabolites, furnishing immeasurable pharmacological and biological activities. Hence, to meet the demand of new lead compounds for human and animal use, research is constantly targeting the bioprospecting of Streptomyces. Optimization of media components and physicochemical parameters is a plausible approach for the exploration of intensified production of novel as well as existing bioactive metabolites from various microbes, which is usually achieved by a range of classical techniques including one factor at a time (OFAT). However, the major drawbacks of conventional optimization methods have directed the use of statistical optimization approaches in fermentation process development. Response surface methodology (RSM) is one of the empirical techniques extensively used for modeling, optimization and analysis of fermentation processes. To date, several researchers have implemented RSM in different bioprocess optimization accountable for the production of assorted natural substances from Streptomyces in which the results are very promising. This review summarizes some of the recent RSM adopted studies for the enhanced production of antibiotics, enzymes and probiotics using Streptomyces with the intention to highlight the significance of Streptomyces as well as RSM to the research community and industries.

Microbial production of glutathione

Abstract: Glutathione (GSH) is a non-coding tripeptide thiol with several important regulative and protective functions in eukaryotes and in most prokaryotes. The primary function of GSH is to maintain the redox potential of the cell, which is directly connected to GSH concentration, and to prevent cellular damages caused by reactive oxygen species or toxic heavy metals. Due to its antioxidant character, it is widely used in pharmaceutical, cosmetic and food industry. There have been different strategies to optimize GSH yield and productivity in bacteria and yeasts by means of metabolic and evolutionary engineering, media optimization and bioprocess engineering. The fed-batch procedure with yeasts of the genera Saccharomyces and Candida is still common method for industrial production. However, for an economic bioprocess production of GSH key factors like media costs, strain performance and process scalability are essential. Beside the extraction and purification of GSH as bulk product, GSH-enriched yeast cells are used for food and beverage applications, as well. This review outlines current applications of microbially produced GSH and illustrates current developments and strategies for its production.

Sensors for disposable bioreactors

Abstract: Modern bioprocess monitoring demands sensors that provide on-line information about the process state. In particular, sensors for monitoring bioprocesses carried out in single-use bioreactors are needed because disposable systems are becoming increasingly important for biotechnological applications. Requirements for the sensors used in these single-use bioreactors are different than those used in classical reusable bioreactors. For example, long lifetime or resistance to steam and cleaning procedures are less crucial factors, while a requirement of sensors for disposable bioreactors is a cost that is reasonable on a per-use basis. Here, we present an overview of current and emerging sensors for single-use bioreactors, organized by the type of interface of the sensor systems to the bioreactor. A major focus is on non-invasive, in-situ sensors that are based on electromagnetic, semiconducting, optical, or ultrasonic measurements. In addition, new technologies like radio-frequency identification sensors or free-floating sensor spheres are presented. Notably, at this time there is no standard interface between single-use bioreactors and the sensors discussed here. In the future, manufacturers should address this shortcoming to promote single-use bioprocess monitoring and control. This article is protected by copyright. All rights reserved

Novel cheese production by incorporation of sea buckthorn berries (Hippophae rhamnoides L.) supported probiotic cells

Abstract: Sea buckhorn berries (SBB, Hippophae rhamnoides L.) were used as a novel support for immobilisation of the probiotic strain L. casei ATCC 393. The biocatalyst was employed for the development of a bioprocess for feta-type cheese production that leads to the improvement of quality and nutritional characteristics. Specifically, the effect of SBB supported biocatalyst on the microbiological safety and aroma profile of cheeses was compared with control samples and cheeses containing free L. casei cells showing superior properties with increased content of esters, alcohols and terpenes. The presence of probiotic culture, either in free or immobilised form, affected positively the physicochemical characteristics of cheeses during ripening. Cheeses with SBB had enriched aroma with terpenes and carbonyl compounds and higher probiotic cell population. The proposed bioprocess of employing SBB as immobilisation carrier shows great potential for commercialisation and application in manufacturing of probiotic functional dairy food.

Production of Ionic Liquid Tolerant Cellulase from Bacillus subtilis G2 Using Agroindustrial Residues with Application Potential for Saccharification of Biomass Under One Pot Consolidated Bioprocess

Abstract: Ionic liquid (IL) based pretreatment of lignocellulosic biomass for facilitating efficient enzymatic saccharification has emerged as an environmentally benign approach that offers several advantages over conventional strategies. However, residues of ionic liquid left in the pretreated biomass may cause inactivation of saccharifying enzymes thus, necessitating the requirement of ionic liquid-stable enzymes. Cost-effective production of industrial enzymes is always desired to enhance the overall process economy. Current study reports IL-stable cellulase production from a newly isolated bacterium Bacillus subtilis G2. Design of experiment (DoE) based on response surface methodology was used in sequential manner for optimizing cultural and environmental variables to enhance cellulase production by 2.66-fold. IL-stable cellulase was used for saccharification of IL-pretreated pine needle biomass (PNB) with 1-ethyl-3-methylimidazolium methanesulfonate in a consolidated single pot process i.e. one pot consolidated bioprocess (OPCB). The saccharification efficiency of 23.57 % was observed under OPCB. The hydrolsate obtained was fermented by dual culture of yeast i.e. Saccharomyces cereviasie NCIM 3078 and Pichia stipitis NCIM 3497, and a yield of 0.092 g ethanol/g of PNB was obtained with fermentation efficiency of 25.62 %. This study is first ever where-in IL-stable cellulase production is accomplished using agroindustrial residues by employing DoE, and assessed for its application potential under OPCB for saccharification of IL-pretreated PNB. IL-stable cellulases would not only preclude expensive washing step following IL-pretreatment of biomass, but their application in a consolidated single pot process (OPCB) offers numerous technoeconomic advantages over conventional multi pot processes. Production of ionic liquid (IL) tolerant cellulase from Bacillus subtilis G2 was enhanced by 2.66-fold using response surface methodology. Cellulase was assessed for its saccharification potential on IL-pretreated pine needle biomass under one pot consolidated bioprocess, and the hydrolysate was fermented to ethanol using Saccharomyces cereviasie and Pichia stipitis.

Hybrid Approach to State Estimation for Bioprocess Control.

Abstract: An improved state estimation technique for bioprocess control applications is proposed where a hybrid version of the Unscented Kalman Filter (UKF) is employed. The underlying dynamic system model is formulated as a conventional system of ordinary differential equations based on the mass balances of the state variables biomass, substrate, and product, while the observation model, describing the less established relationship between the state variables and the measurement quantities, is formulated in a data driven way. The latter is formulated by means of a support vector regression (SVR) model. The UKF is applied to a recombinant therapeutic protein production process using Escherichia coli bacteria. Additionally, the state vector was extended by the specific biomass growth rate µ in order to allow for the estimation of this key variable which is crucial for the implementation of innovative control algorithms in recombinant therapeutic protein production processes. The state estimates depict a sufficiently low noise level which goes perfectly with different advanced bioprocess control applications.

Fast and reliable strain characterization of Streptomyces lividans through micro-scale cultivation

Abstract: Filamentous organisms of the genus Streptomyces play an important role in industrial production processes, due to their extensive secondary metabolism variability, as well as their ability to secrete efficiently large amounts of (heterologous) proteins. While genetic engineering tools are available to rapidly build up large strain libraries, the subsequent strain screening and bioprocess development still constitutes a bottleneck. This is due to the lack of reliable parallelized and accelerated cultivation techniques for morphologically challenging organisms. To address this challenge, we developed an integrated cultivation workflow for Streptomyces lividans based on a parallelized shaken 48-well microtiter-plate (MTP) cultivation device. In a first step, a feasible pre-culture method was identified and validated, revealing high comparability in subsequent main cultivations (coefficient of variation of 1.1% for in-plate replicates and 3.2% between different pre-cultures). When validating the growth performance in 1 mL MTP cultivation against an established 1,000 mL lab-scale cultivation system, highly comparable cultivation patterns were found for online (pH, dissolved oxygen), as well as for offline derived parameters (glucose uptake, cell-dry-weight, and pellet size). Additionally, the two cultivation regimes were compared with respect to transcriptional and protein secretion activity of Streptomyces, showing overall good comparability with minor, but well explainable discrepancies, most probably caused by different energy dissipation (shaking vs. stirring) and adaption effects due to different illumination conditions. Embedded within the presented cultivation workflow, the 1 mL MTP-based parallelized cultivation system seems to be a suitable screening tool for filamentous and industrial relevant organisms like Streptomyces. This can contribute to widen the field of application for these organisms and facilitate screening and early-stage bioprocess development. Biotechnol. Bioeng. 2017;114: 2011-2022. © 2017 Wiley Periodicals, Inc.

Exploitation of Agricultural Wastes and By-Products for Production of Aureobasidium pullulans Y-2311-1 Xylanase: Screening, Bioprocess Optimization and Scale Up

Abstract: The potential of several agricultural wastes and by-products (wheat bran, oat bran, corn cob, brewer’s spent grain, malt sprout, artichoke stem, sugar beet pulp, olive seed, cotton stalk and hazelnut skin) was examined as the substrate for xylanase production by Aureobasidium pullulans Y-2311-1. Based on the screening studies, wheat bran was selected as the best substrate for further optimization studies. The effects of initial medium pH, temperature and incubation time on xylanase production in shake flask system were optimized by response surface methodology (RSM). The optimum levels of the process variables defined by the model (initial medium pH, 4.24; temperature, 30.27 °C; and incubation time 126.67 h) resulted in production of 85.19 U/ml xylanase. Taking the RSM optimized parameters in shake-flask scale into consideration; xylanase production was scaled up to bioreactor system with a working volume of 1.5 l. The peak of enzyme production was achieved after 126 h incubation that has previously been determined by RSM studies at shake flask level. Furthermore, the optimum levels of agitation and aeration in bioreactor system was found as 200 rpm and 1.5 vvm. Maximum enzyme production was close to 85 kU/l which could be translated into a productivity of 0.68 kU/l/h. No previous work considered the statistical optimization of xylanase production by A. pullulans on wheat bran and scale up of the bioprocess to a bioreactor system.