Metabolites In Cell Cultures Research Articles

Elucidating the efficacy of functionalized multi-walled carbon nanotube in the biogenesis of L-Dopa and antioxidant metabolites in cell cultures of Hybanthus enneaspermus

Hybanthus enneaspermus (L.)F.Muell. is a highly indispensable medicinal herb yielding L-Dopa, deemed the gold standard drug among the therapeutic options for Parkinson's disease. This investigation is the first attempt to evaluate the eliciting influence of carboxylic acid functionalized multi-walled carbon nanotube (MWCNT-COOH) on the biosynthesis of L-Dopa and on biomass aggregation and antioxidant metabolites in H. enneaspermus cell suspension cultures. Suspension cells were accomplished from friable calli generated from the nodal segments of H. enneaspermus in Murashige and Skoog (MS) liquid medium infused with 2 mg L−1 2, 4-Dichlorophenoxyacetic acid (2, 4-D), and 0.3 mg L−1meta-Topolin (mT). The influence of MWCNTs on L-Dopa synthesis, biomass accumulation, and biochemical parameters was examined on the basis of the exposure time and in a concentration-dependent manner of MWCNTs. The inclusion of 30 mg L−1 MWCNTs increased the biomass and the L-Dopa level by 2.00 and 16.37-folds, respectively, compared with that of the control. Furthermore, the effect of MWCNTs on physiological parameters such as catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPX), ascorbate peroxidase (APX), hydrogen peroxide (H2O2), malondialdehyde (MDA) content, 2-diphenylpicrylhydrazyl (DPPH), and ferric-reducing ability of plasma (FRAP) was examined over the elicited cells. Among the antioxidant enzymatic activities, CAT enhanced 8.0 fold compared with that of the control. MDA and DPPH content enhanced 2.60 and 1.12 folds, respectively, compared with that of the control. The current study showed that MWCNTs offer new possibilities for their usage over in vitro by acting as potential innovative plant metabolite elicitors and stress-protecting entities.

Histological Structure and Composition of Secondary Metabolites in Cell Culture of Drosera rotundifolia L.

Histological Structure and Composition of Secondary Metabolites in Cell Culture of Drosera rotundifolia L.

Soft-sensors application for automated feeding control in high-throughput mammalian cell cultures.

The ever-increasing demand for biopharmaceuticals has created the need for improving the overall productivity of culture processes. One such operational concept that is considered is fed-batch operations as opposed to batch operations. However, optimal fed-batch operations require complete knowledge of the cell culture to optimize the culture conditions and the nutrients feeding. For example, when using high-throughput small-scale bioreactors to test multiple clones that do not behave the same, depletion or overfeeding of some key components can occur if the feeding strategy is not individually optimized. Over the recent years, various solutions for real-time measuring of the main cell culture metabolites have been proposed. Still, the complexity in the implementation of these techniques has limited their use. Soft-sensors present an opportunity to overcome these limitations by indirectly estimating these variables in real-time. This manuscript details the development of a new soft-sensor-based fed-batch strategy to maintain substrate concentration (glucose and glutamine) at optimal levels in small-scale multiparallel Chinese Hamster Ovary Cells cultures. Two alternatives to the standard feeding strategy were tested: an OUR soft-sensor-based strategy for glucose and glutamine (Strategy 1) and a dual OUR for glutamine and CO2 /alkali addition for glucose soft-sensor strategy (Strategy 2). The results demonstrated the applicability of the OUR soft-sensor-based strategy to optimize glucose and glutamine feedings, which yielded a 21% increase in final viable cell density (VCD) and a 31% in erythropoietin titer compared with the reference one. However, CO2 /alkali addition soft-sensor suffered from insufficient data to relate alkali addition with glucose consumption. As a result, the culture was overfed with glucose resulting in a 4% increase on final VCD, but a 9% decrease in final titer compared with the Reference Strategy.

Quantitative determination of biphenyls and their metabolites in cell cultures of Comamonas thiooxydans N1 using high-performance liquid chromatography

Biphenyl-degrading bacteria were isolated from contaminated soil and identified as Comamonas species. Among these strains, C. thiooxydans N1 was the most suitable for biphenyl degradation. When 1200 µg/mL of biphenyl was used, it was completely degraded within 24 h. A simple, rapid, and precise, high-performance liquid chromatography (HPLC) method for quantifying biphenyls and their metabolites in the culture medium was optimised and validated. The proposed method was performed using reverse-phase HPLC equipped with a Luna C18 column. The mobile phase was water and acetonitrile (30/70, v/v) under isocratic conditions, and biphenyls and their metabolites were quantified on a SHIMADZU HPLC system with a diode-array detection detector at 254 nm. Peak identity was confirmed by ultraviolet spectroscopy and gas chromatography-mass spectrometry. Validation was performed according to European Union requirements (2002/657/EC) for linearity, accuracy, repeatability, reproducibility, limit of detection, limit of quantification, and detection capability. Linearity was observed in the concentration range of 0.04-20 µg/mL with a high coefficient of determination (R2 ≥ 0.999). The validated method was applied to determine the content of biphenyl and its metabolites in the culture medium.

Biomass production and secondary metabolite identification in callus cultures of Coryphantha macromeris (Engelm.) Britton & Rose (Cactaceae), a traditional medicinal plant

Plant cell, tissue, and organ culture have become a powerful technology for the production of biomass, and for the research and biosynthesis of a variety of secondary metabolites. In this work, we report an efficient method for friable callus induction applied to the medicinal cactus Coryphantha macromeris, its kinetic behavior, and its phytochemical profile, assessed at the maximum biomass production phase. Callus cultures were obtained from stem discs inoculated on Murashige and Skoog medium (MS) supplemented with 6-benzylaminopurine (BAP; 2.2 µM) and picloram (4.14 µM). The highest biomass production (20.65 g DW L−1) was achieved at nine weeks of culture. Then, the phytochemical profile was analyzed using Ultra-High-Performance Liquid Chromatography-tandem Mass Spectrometry (UHPLC-PDA-HESI-Orbitrap-MS/MS). Chromatographic and mass spectrometric analyses indicated the presence of 61 metabolites, with 52 being identified. Among these compounds, 11 organic acids, 16 phenolic acids, 8 flavonoids, and 17 metabolites of different classes were identified. Our results could significantly contribute to the current knowledge of tissue culture of cacti species, as well as the potential applications of the in vitro callus culture of C. macromeris. Furthermore, we report the presence of some metabolites in cell culture of cacti species and their fragmentation pattern for the first time.

High-throughput LC-MS quantitation of cell culture metabolites

Monitoring cell culture metabolites, including media components and cellular byproducts, during bio manufacturing is critical for gaining insights into cell growth, productivity and product quality. Historically, cell culture metabolite analysis was a complicated process requiring several orthogonal methods to cover the large number of metabolites with diverse properties over wide concentration ranges. These off-line analyses are time consuming and not suitable for real time bioreactor monitoring. In this study, we present a high-throughput LC-MS method with a 17-min cycle time that is capable of simultaneously monitoring 93 cell culture metabolites, including amino acids, nucleic acids, vitamins, sugars and others. This method has high precision and accuracy and has been successfully applied to the daily profiling of bioreactors and raw material qualification. Information obtained in these studies has been used to identify limiting amino acids during production, which guided adjustments to the feed strategy that prevented the potential misincorporation of amino acids. This type of metabolite profiling can be further utilized to build predictive process models for adaptive feedback control and pave the road for continuous manufacturing and real-time release testing.

Sustainable production of biomass and industrially important secondary metabolites in cell cultures of selfheal (Prunella vulgaris L.) elicited by silver and gold nanoparticles

Elicited artificial in vitro cultures are gaining more interest due to their uniform biosynthesis of industrially valuable secondary metabolites. In this study, a unique methodology was applied, in which different ratios of gold (Au) and silver (Ag) nanoparticles (NPs) were supplemented to submerge cultures to investigate sustainable production of biomass and antioxidant secondary metabolites. Cell suspension cultures were exposed to Ag and AuNPs alone or different ratios of AgAuNPs (1:2; 1:3; 2:1; 3:1) in combination with NAA. The combination of AgAuNPs (3:1) with NAA enhanced fresh (9.25 g/100 ml) and dry biomass (0.64 g/100 ml) of suspended cells than control (6.67; 0.233 g/100 ml). AuNPs with NAA-augmented media enhanced biomass accumulation in lag, log and stationary phases in a period of 49 days. Furthermore, AgAu (3:1) and AgAuNPs (2:1; 1:2) with NAA enhanced protein contents, peroxidase and superoxide dismutase enzymes. However, maximum phenolics (TPC; 10.61 mg/g-DW) and flavonoids (7.62 mg/g-DW) were observed in cell cultures exposed to a combination of AgAuNPs (1:3) and NAA than control (6.27, 5.49 mg/g-DW). The combination of AgAuNPs (2:1) with NAA enhanced antioxidant activity (87.85%) in cell cultures. This study will help in illuminating the impact of NPs on cultures development and production of natural antioxidants.

Production of Plant Secondary Metabolites: Examples, Tips and Suggestions for Biotechnologists.

Plants are sessile organisms and, in order to defend themselves against exogenous (a)biotic constraints, they synthesize an array of secondary metabolites which have important physiological and ecological effects. Plant secondary metabolites can be classified into four major classes: terpenoids, phenolic compounds, alkaloids and sulphur-containing compounds. These phytochemicals can be antimicrobial, act as attractants/repellents, or as deterrents against herbivores. The synthesis of such a rich variety of phytochemicals is also observed in undifferentiated plant cells under laboratory conditions and can be further induced with elicitors or by feeding precursors. In this review, we discuss the recent literature on the production of representatives of three plant secondary metabolite classes: artemisinin (a sesquiterpene), lignans (phenolic compounds) and caffeine (an alkaloid). Their respective production in well-known plants, i.e., Artemisia, Coffea arabica L., as well as neglected species, like the fibre-producing plant Urtica dioica L., will be surveyed. The production of artemisinin and caffeine in heterologous hosts will also be discussed. Additionally, metabolic engineering strategies to increase the bioactivity and stability of plant secondary metabolites will be surveyed, by focusing on glycosyltransferases (GTs). We end our review by proposing strategies to enhance the production of plant secondary metabolites in cell cultures by inducing cell wall modifications with chemicals/drugs, or with altered concentrations of the micronutrient boron and the quasi-essential element silicon.

Accurate and sensitive determination of molar fractions of 13C-Labeled intracellular metabolites in cell cultures grown in the presence of isotopically-labeled glucose

This work describes a methodology based on multiple linear regression and GC-MS for the determination of molar fractions of isotopically-labeled intracellular metabolites in cell cultures. Novel aspects of this work are: i) the calculation of theoretical isotopic distributions of the different isotopologues from an experimentally measured value of % 13C enrichment of the labeled precursor ii) the calculation of the contribution of lack of mass resolution of the mass spectrometer and different fragmentation mechanism such as the loss or gain of hydrogen atoms in the EI source to measure the purity of the selected cluster for each metabolite and iii) the validation of the methodology not only by the analysis of gravimetrically prepared mixtures of isotopologues but also by the comparison of the obtained molar fractions with experimental values obtained by GC-Combustion-IRMS based on 13C/12C isotope ratio measurements. The method is able to measure molar fractions for twenty-eight intracellular metabolites derived from glucose metabolism in cell cultures grown in the presence of 13C-labeled Glucose. The validation strategies demonstrate a satisfactory accuracy and precision of the proposed procedure. Also, our results show that the minimum value of 13C incorporation that can be accurately quantified is significantly influenced by the calculation of the spectral purity of the measured cluster and the number of 13C atoms of the labeled precursor. The proposed procedure was able to accurately quantify gravimetrically prepared mixtures of natural and labeled glucose molar fractions of 0.07% and mixtures of natural and labeled glycine at molar fractions down to 0.7%. The method was applied to initial studies of glucose metabolism of different prostate cancer cell lines.

Vitamin D receptor expression in human bone tissue and dose-dependent activation in resorbing osteoclasts.

The effects of vitamin D on osteoblast mineralization are well documented. Reports of the effects of vitamin D on osteoclasts, however, are conflicting, showing both inhibition and stimulation. Finding that resorbing osteoclasts in human bone express vitamin D receptor (VDR), we examined their response to different concentrations of 25-hydroxy vitamin D3 [25(OH)D3] (100 or 500 nmol·L−1) and 1,25-dihydroxy vitamin D3 [1,25(OH)2D3] (0.1 or 0.5 nmol·L−1) metabolites in cell cultures. Specifically, CD14+ monocytes were cultured in charcoal-stripped serum in the presence of receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). Tartrate-resistant acid phosphatase (TRAP) histochemical staining assays and dentine resorption analysis were used to identify the size and number of osteoclast cells, number of nuclei per cell and resorption activity. The expression of VDR was detected in human bone tissue (ex vivo) by immunohistochemistry and in vitro cell cultures by western blotting. Quantitative reverse transcription-PCR (qRT-PCR) was used to determine the level of expression of vitamin D-related genes in response to vitamin D metabolites. VDR-related genes during osteoclastogenesis, shown by qRT-PCR, was stimulated in response to 500 nmol·L−1 of 25(OH)D3 and 0.1–0.5 nmol·L−1 of 1,25(OH)2D3, upregulating cytochrome P450 family 27 subfamily B member 1 (CYP27B1) and cytochrome P450 family 24 subfamily A member 1 (CYP24A1). Osteoclast fusion transcripts transmembrane 7 subfamily member 4 (tm7sf4) and nuclear factor of activated T-cell cytoplasmic 1 (nfatc1) where downregulated in response to vitamin D metabolites. Osteoclast number and resorption activity were also increased. Both 25(OH)D3 and 1,25(OH)2D3 reduced osteoclast size and number when co-treated with RANKL and M-CSF. The evidence for VDR expression in resorbing osteoclasts in vivo and low-dose effects of 1,25(OH)2D3 on osteoclasts in vitro may therefore provide insight into the effects of clinical vitamin D treatments, further providing a counterpoint to the high-dose effects reported from in vitro experiments.

Enhanced Accumulation of Scopoletin in Cell Suspension Culture of Spilanthes acmella Murr. Using Precursor Feeding

In this study, the various concentrations of casein hydrolysate (25, 50, 75, 100 mg/L) and L-phenylalanine (50, 100, 150, 200 µM/l) were incorporated in MS containing 15 µM BA plus 5 µM 2,4-D for enhancement of secondary metabolites in cell culture of Spilanthes acmella. The presence of casein hydrolysate in the nutrient medium improved the growth of cell biomass and the production of scopoletin. The addition of casein hydrolysate up to 75 mg/L stimulated the accumulation of scopoletin, but increasing excess 75 mg/L the level of casein hydrolysate reduced the production of scopoletin. The addition of L-phenylalanine in the nutrient medium was found to be more effective for production of secondary metabolite in S. acmella. The addition of 50 µM/L of L-phenylalanine in the medium increased scopoletin content to 27.12 ± 0.58 µg/g dry weight, compared to the scopoletin content of control at 7.89 ± 0.61 µg/g dry weight. The highest accumulation of scopoletin was observed in the 100 µM/L L-phenylalanine in cell suspension, which was 4.51 times more than the control. As a result, using moderate concentration of L-phenylalanine was ideal for the production of scopoletin. In general, casein hydrolysate was more effective than L-phenylalanine for production of scopoletin and growth of cell biomass in the cell culture of S. acmella.

In-line and real-time prediction of recombinant antibody titer by in situ Raman spectroscopy

The Food and Drug Administration's (FDA) process analytical technology (PAT) framework has been initiated to encourage drug manufacturers to develop innovative techniques in order to better understand their processes and institute high level quality control which allows action at any point in the manufacturing process. While Raman spectroscopy and chemometrics have been successfully used to predict concentration of conventional metabolites in cell cultures, it is really not the case for active substances. Thus, we propose, for the first time, an in-line and real-time prediction of recombinant antibody titer using an immersion probe link to a spectrometer without the tacking of samples. A good robustness of the method is observed on different culture batches and the contamination risk is drastically reduced which is an important issue in biotechnology manufacturing processes.

Real time monitoring of bioreactor mAb IgG3 cell culture process dynamics via Fourier transform infrared spectroscopy: Implications for enabling cell culture process analytical technology

Compared to small molecule process analytical technology (PAT) applications, biotechnology product PAT applications have certain unique challenges and opportunities. Understanding process dynamics of bioreactor cell culture process is essential to establish an appropriate process control strategy for biotechnology product PAT applications. Inline spectroscopic techniques for real time monitoring of bioreactor cell culture process have the distinct potential to develop PAT approaches in manufacturing biotechnology drug products. However, the use of inline Fourier transform infrared (FTIR) spectroscopic techniques for bioreactor cell culture process monitoring has not been reported. In this work, real time inline FTIR Spectroscopy was applied to a lab scale bioreactor mAb IgG3 cell culture fluid biomolecular dynamic model. The technical feasibility of using FTIR Spectroscopy for real time tracking and monitoring four key cell culture metabolites (including glucose, glutamine, lactate, and ammonia) and protein yield at increasing levels of complexity (simple binary system, fully formulated media, actual bioreactor cell culture process) was evaluated via a stepwise approach. The FTIR fingerprints of the key metabolites were identified. The multivariate partial least squares (PLS) calibration models were established to correlate the process FTIR spectra with the concentrations of key metabolites and protein yield of in-process samples, either individually for each metabolite and protein or globally for all four metabolites simultaneously. Applying the 2nd derivative pre-processing algorithm to the FTIR spectra helps to reduce the number of PLS latent variables needed significantly and thus simplify the interpretation of the PLS models. The validated PLS models show promise in predicting the concentration profiles of glucose, glutamine, lactate, and ammonia and protein yield over the course of the bioreactor cell culture process. Therefore, this work demonstrated the technical feasibility of real time monitoring of the bioreactor cell culture process via FTIR spectroscopy. Its implications for enabling cell culture PAT were discussed.

Raman spectroscopy of complex defined media: biopharmaceutical applications

We demonstrate the detection of glucose and lactate concentrations with high accuracy in the supernatants of Chinese hamster ovary (CHO) cell culture, grown in shake flasks in batch fermentation mode, using Raman spectroscopy and explicit model-based classical least squares (CLS) algorithm. A deterministic Raman spectral library of pure components was created by acquiring Raman spectra from credible nutrient media constituents and CHO cell culture metabolites. Only analytes present with concentration above the instrument detection limit were included in this library. Residuals obtained after CLS analyses were used to identify missing components and to generate a revised library. An algorithmic sieve was thus construed to obtain an appropriate Raman spectral library from a complex chemical mixture that is well-defined but an industrial secret. High performance liquid chromatography (HPLC) was used to provide reference glucose and lactate concentrations. [Correction added on 30 April 2015, after first online publication: the last three sentences of the abstract (previously stated in the abstract) have been deleted]. Copyright © 2015 John Wiley & Sons, Ltd.

Estimation of bacterioruberin by Raman spectroscopy during the growth of halophilic archaeon Haloarcula marismortui.

Halophilic archaea are interesting microorganisms that produce low biomass and metabolites, complicating their quantification. Raman spectroscopy (RS) is a powerful technique, which requires small samples, attractive for using in archaeal research. The objective of this work was the estimation of bacterioruberin content along with Haloarcula marismortui growth and their correlation with biomass concentration. RS was used to detect characteristic bands of bacterioruberin (vibrational modes C═CH, C─C, and C═C) in H. marismortui culture samples. The intensity of Raman spectra in bacterioruberin and the biomass concentration were adequately correlated. The highest production of bacterioruberin occurred at 60h. RS is revealed as a reliable technique for the estimation of bacterioruberin in the biomass of H. marismortui, which could be considered as a promising qualitative and quantitative technique to assay metabolites in cell cultures.

Stressing Conditions as Tools to Boost the Biosynthesis of Valuable Plant Natural Products

There is a consensus that plants are great sources of metabolites with a broad variety of functions. This is particularly important because plants cannot run away from environmental conditions that can threat their existence. The numerous biological activities exhibited by plant natural products prompted humanity to use such substances or their derivatives for the treatment and/or prevention of diseases. The more we know the flora around the world the higher is the chance to find new lead compounds for the design of more potent drugs or nutraceuticals. This review first deals with Brazilian flora, contextualizing the most studied medicinal species and related patents. It also describes a compilation of relevant works based on the use of stress conditions to enhance the biosynthesis of valuable metabolites in cell cultures, tissue cultures (hairy roots) and whole plants by using native or crop plants around the world.

Homology modeling and docking studies on oxidosqualene cyclases associated with primary and secondary metabolism of Centella asiatica.

Centella asiatica is a well-known medicinal plant, produces large amount of triterpenoid saponins, collectively known as centelloids, with a wide-spectrum of pharmacological applications. Various strategies have been developed for the production of plant secondary metabolites in cell and tissue cultures; one of these is modular metabolic engineering, in which one of the competitive metabolic pathways is selectively suppressed to channelize precursor molecules for the production of desired molecules by another route. In plants the precursor 2,3-oxidosqualene is shared in between two competitive pathways involved with two isoforms of oxidosqualene cyclases. One is primary metabolic route for the synthesis of phytosterol like cycloartenol by cycloartenol synthase; another is secondary metabolic route for the synthesis of triterpenoid like β-amyrin by β-amyrin synthase. The present work is envisaged to evaluate specific negative modulators for cycloartenol synthase, to channelize the precursor molecule for the production of triterpenoids. As there are no experimentally determined structures for these enzymes reported in the literature, we have modeled the protein structures and were docked with a panel of ligands. Of the various modulators tested, ketoconazole has been evaluated as the negative modulator of primary metabolism that inhibits cycloartenol synthase specifically, while showing no interaction with β-amyrin synthase. Amino acid substitution studies confirmed that, ketoconazole is specific modulator for cycloartenol synthase, LYS728 is the key amino acid for the interaction. Our present study is a novel approach for identifying a suitable specific positive modulator for the over production of desired triterpenoid secondary metabolites in the cell cultures of plants.

Problems, progress and future prospects of improvement of Commiphora wightii (Arn.) Bhandari, an endangered herbal magic, through modern biotechnological tools: a review

Commiphora wightii (Arn.) Bhandari syn. C. mukul Engl. (Burseraceae) is an economically and pharmacologically important slow growing, dioecious, balsamiferous woody, multipurpose shrub heading towards extinction. Commonly known as “Guggul” due to the presence of steroidal compound guggulsterone in the oleo-gum resin, it has been used in treating various ailments and disorders since ancient times (2000 B.C.). Evaluation and confirmation of hypolipidemic effects of guggul based on Ayurvedic text in 1960s provided a new insight into its pharmacological applications. Two bioactive isomers of guggulsterone, E and Z, are responsible for lipid- and cholesterol-lowering activities. Recently, it has been shown to have anti-cancerous activity also. It is found in the dry regions of Indian subcontinent, namely India, Pakistan and Bangladesh. Ruthless and unscientific harvesting of oleo-gum resin from the wild, by local populations, for economic benefits with negligible conservation efforts has made this species endangered and has led to its inclusion in Red Data Book of IUCN. Although this plant has many excellent traits, adequate attention has not been focused on its conservation and improvement. Conventional propagation methods i.e., seeds, cuttings and air layering are in place but have many limitations. Therefore, application of modern biotechnological tools needs to be standardized for harnessing maximum benefits from this pharmaceutically important plant. An efficient regeneration system needs to be in place for improvement of this genus through genetic transformation and production of useful metabolites in cell cultures. Studies are in progress for micropropagation through shoot multiplication and somatic embryogenesis, as well as for secondary metabolite (guggulsterone) production in callus cultures and bioreactors. No selected germplasm is available for C. wightii since it is a wild plant. Breeding programs have not yet been started due to lack of systematic cultivation and conservation programs. Moreover, little information has been gathered regarding the genetic variability in this species using RAPD and ISSR markers. No details are available about genetic makeup and QTL linkage maps. Investigations are in progress to search sex linked markers in this dioecious species. Research is also in progress to decipher the molecular mechanisms underlying various pharmacological actions of guggul. Since the approval of use of guggul as a food supplement by United States Food and Drug Administration in 1994, an exponential increase in research publications on various aspects of research on guggul have been published. Present communication summarizes the problems, progress made and suggests some future directions of research for this important endangered medicinal plant.

Software tool for coupling chromatographic total ion current dependencies of GC/MSD and MCC/IMS

More than 90,000 ion mobility spectrometer (IMS) are in use worldwide, mostly without coupling to chromatographic columns for pre-separation of the neutrals entering the ionization region of the IMS. Related to new application fields e.g. breath analysis or determination of volatile metabolites of cell cultures and bacteria/fungi an effective way for pre-separation is needed strongly. The acceptance of IMS within the analytical world will be enhanced by supplying automatic procedures for peak finding, referencing and database-related identification of the signals within IMS-Chromatograms. Some papers are available concerning the internal loop of relation of IMS signals between different instruments, alignment for automatic interpretation with respect of reference analytes used and interpolation within the different members of classes of analytes in case that some missed e.g. within a homologous row of chemicals. With respect to inter comparison of recent findings using MCC/IMS and GC/MS experiments effective alignment procedures are developed. In the present paper we describe a software tool making the visualization of the total ion current (TIC) from chromatograms of GC/MSD possible in direct relation to the corresponding IMS-Chromatogram of MCC/IMS. Thus, parallel measurements of e.g. samples of human breath using MCC/IMS and thermodesorption (TD)-GC/MSD become comparable directly. On the other hand, the direct identification and relation of so far unknown peaks found in IMS-Chromatograms will be supported using the TIC results of GC/MSD.

New Perspectives for Real-Time Multi-Metabolites Monitoring of Differentiating Neuron Precursors by using Protein-Based Carbon Nanotubes

Event Abstract Back to Event New Perspectives for Real-Time Multi-Metabolites Monitoring of Differentiating Neuron Precursors by using Protein-Based Carbon Nanotubes Sandro Carrara1* 1 Ecole Polytechnique Federale de Lausanne, School of Engineering, Switzerland Monitoring of metabolic compounds, such as glucose and lactate, is extensively reported in literature, especially for clinical purposes. Instead, the application of such technologies for monitoring metabolites in cell cultures has not been deeply explored. Monitoring systems can provide further information to the current state-of-the-art of cell lines, particularly those that are not fully known, as stem and embryonic cells. On the other hand, metabolism biosensors can address fully automation for growing cell cultures, once sensors are coupled with fluidic systems for cell feeding. Among different presented strategies to develop biosensors, carbon nanotubes exhibit great properties, particularly suitable for bio-sensing. In this talk, nano-structured electrodes by using multi-walled carbon nanotubes are presented for specific detection of glucose, glutamate and lactate in neuron cultures. Some results from simulations are illustrated in order to show the behavior of carbon nanotubes depending on their orientation once they are casted onto electrodes surface. Data from Scanning Electron Microscopy, electrochemical amperometry and voltammetry are presented to characterize the performance of the developed sensors. Carbon nanotubes functionalized with proper probe proteins are characterized in terms of sensitivity, detection limit, and time stability. Their detection performances are compared with previously published results. The monitoring of cell cultures of neurons precursors is showed and metabolic behavior is registered as validation of the developed monitoring system. Glucose and lactate are acquired during proliferation of neuron precursors and their differentiation in mature neurons. We also investigate lactate and glucose in conditions of glucose deprivation, showing direct confirmation of lactate self-production during cell suffering. Finally, new perspectives in continuous real-time monitoring of cell metabolism are presented. The talk demonstrates that the proposed electrochemical biosensors based on carbon nanotubes are suitable for stem cell real-time monitoring as well as for cell biology investigation. Keywords: neuroengineering Conference: The Monte Verita' Workshop on the Frontiers in Neuroengineering, Ascona, Switzerland, 5 Sep - 9 Sep, 2010. Presentation Type: Oral Presentation Topic: Frontiers in Neuroengineering Citation: Carrara S (2010). New Perspectives for Real-Time Multi-Metabolites Monitoring of Differentiating Neuron Precursors by using Protein-Based Carbon Nanotubes. Front. Neuroeng. Conference Abstract: The Monte Verita' Workshop on the Frontiers in Neuroengineering. doi: 10.3389/conf.fneng.2010.10.00009 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 24 Aug 2010; Published Online: 10 Sep 2010. * Correspondence: Dr. Sandro Carrara, Ecole Polytechnique Federale de Lausanne, School of Engineering, Lausanne, Switzerland, sandro.carrara@epfl.ch Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Sandro Carrara Google Sandro Carrara Google Scholar Sandro Carrara PubMed Sandro Carrara Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.