Fast quantitative determination of microbial rhamnolipids from cultivation broths by ATR-FTIR Spectroscopy

<p>Abstract</p> <p>Background</p> <p>Vibrational spectroscopic techniques are becoming increasingly important and popular because they have the potential to provide rapid and convenient solutions to routine analytical problems. Using these techniques, a variety of substances can be characterized, identified and also quantified rapidly.</p> <p>Results</p> <p>The rapid ATR-FTIR (Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy) <it>in time </it>technique has been applied, which is suitable to quantify the concentrations of microbial rhamnolipids in a typical cultivation process. While the usually applied HPLC analysis requires an extensive and time consuming multi step extraction protocol for sample preparation, the ATR-FTIR-method allows the quantification of the rhamnolipids within 20 minutes. Accuracies between 0.5 g/l – 2.1 g/l for the different analytes were determined by cross validation of the calibration set. Even better accuracies between 0.28 g/l – 0.59 g/l were found for independent test samples of an arbitrarily selected cultivation.</p> <p>Conclusion</p> <p>ATR-FTIR was found to be suitable for the rapid analysis of rhamnolipids in a biotechnological process with good reproducibility in sample determination and sufficient accuracy. An improvement in accuracy through continuous expansion and validation of the reference spectra set seems very likely.</p

Enzymatic synthesis of glucose monodecanoate in a hydrophobic deep eutectic solvent

Environmentally friendly and biodegradable reaction media are an important part of a sustainable glycolipid production in the transition to green chemistry. Deep eutectic solvents (DESs) are an ecofriendly alternative to organic solvents. So far, only hydrophilic DESs were considered for enzymatic glycolipid synthesis. In this study, a hydrophobic DES consisting of (-)-menthol and decanoic acid is presented for the first time as an alternative to hydrophilic DES. The yields in the newly introduced hydrophobic DES are significantly higher than in hydrophilic DESs. Different reaction parameters were investigated to optimize the synthesis further. Twenty milligrams per milliliter iCalB and 0.5 M glucose resulted in the highest initial reaction velocity for the esterification reaction, while the highest initial reaction velocity was achieved with 1.5 M glucose in the transesterification reaction. The enzyme was proven to be reusable for at least five cycles without significant loss of activity

Transaminases for the synthesis of enantiopure beta-amino acids

Optically pure β-amino acids constitute interesting building blocks for peptidomimetics and a great variety of pharmaceutically important compounds. Their efficient synthesis still poses a major challenge. Transaminases (also known as aminotransferases) possess a great potential for the synthesis of optically pure β-amino acids. These pyridoxal 5'-dependent enzymes catalyze the transfer of an amino group from a donor substrate to an acceptor, thus enabling the synthesis of a wide variety of chiral amines and amino acids. Transaminases can be applied either for the kinetic resolution of racemic compounds or the asymmetric synthesis starting from a prochiral substrate. This review gives an overview over microbial transaminases with activity towards β-amino acids and their substrate spectra. It also outlines current strategies for the screening of new biocatalysts. Particular emphasis is placed on activity assays which are applicable to high-throughput screening

Process development for the elucidation of mycotoxin formation in Alternaria alternata

The black mould Alternaria alternata produces a wide diversity of mycotoxins which are of particular health concern. Since no maximum allowable limits are set for Alternaria toxins in food and feed, prevention of Alternaria infestations and mycotoxin spoilage is the only way to avoid health risks. Thus, the understanding of mycotoxin biosynthesis is essential. For that purpose, a reliable batch process in a 2 L bioreactor was established which enables the study of several parameters influencing the production of the mycotoxins alternariol (AOH), alternariol monomethylether (AME) and tenuazonic acid (TA) by A. alternata DSM 12633. Modified Czapek-Dox medium was used with glucose as carbon source and ammonium and nitrate as nitrogen sources. Consumption of carbon and nitrogen sources as well as formation of the three mycotoxins were monitored; the average data of five independent fermentations was plotted and fitted using a logistic equation with four parameters. Maximum mycotoxin concentrations of 3.49 ± 0.12 mg/L AOH, 1.62 ± 0.14 mg/L AME and 38.28 ± 0.1 mg/L TA were obtained

Le Concile Vatican II - l'idee de la nouvelle evagelisation en gestation

The Second Vatican Council is undoubtedly one of the most important and even crucial points in the history of Catholic Church. Consequently, it seems that also in relation to the concept so vital for the contemporary Theology, namely «the new evangelization», the recent council played an important role. This paper analyses the conciliar texts from the perspective of the concept of «communio», which is defined by many theologians as the most essential and pivotal term for understanding the Council. From a detailed analysis of the particular dimensions of «com- munio» (Trinitarian, Pneumatological, laical, eschatological, ecumenical, evan- gelisation, missionary) emerges a new vision of ecclesiology, closely related to another conciliar notion of «aggiornamento». This renewed ecclesiological understanding marks a new era in the life of the Church, which John Paul II called the «new spring of Christian life». The article points out that the Vatican II notion of the ecclesiology of communion underlies the later concept of new evangelisation.Sobór Watykański II jest niewątpliwie jednym z ważniejszych, a nawet przełomowych punktów historii Kościoła. W konsekwencji wydaje się więc, że także w odniesieniu do pojęcia tak kluczowego dla współczesnej teologii, jakim jest nowa ewangelizacja, ostatni sobór odegrał istotną rolę. W niniejszej publikacji autor analizuje soborowe teksty rozważając je wokół pojęcia communio, które przez wielu teologów określane jest najważniejszym i kluczowym dla rozumienia terminem Soboru. Ze szczegółowej analizy poszczególnych wymiarów communio (trynitarne, pneumatologiczne, wiernych, eschatologiczne, ekumeniczne, ewangelizacyjne, misyjne) wyłania się nowa wizja eklezjologii ściśle związana z innym soborowym pojęciem aggiornamento. Odnowiona wizja eklezjologiczna naznacza nową epokę w życiu Kościoła, którą Jan Paweł II nazwał „nową wiosną życia chrześcijańskiego”. Autor wskazuje, że soborowa idea eklezjologii komunii stoi u podstaw późniejszego pojęcia nowej ewangelizacji.

Continuous Flow Glycolipid Synthesis Using a Packed Bed Reactor

Glycolipids are a class of biodegradable biosurfactants that are non-toxic and based on renewables, making them a sustainable alternative to petrochemical surfactants. Enzymatic synthesis allows a tailor-made production of these versatile compounds using sugar and fatty acid building blocks with rationalized structures for targeted applications. Therefore, glycolipids can be comprehensively designed to outcompete conventional surfactants regarding their physicochemical properties. However, enzymatic glycolipid processes are struggling with both sugars and fatty acid solubilities in reaction media. Thus, continuous flow processes represent a powerful tool in designing efficient syntheses of sugar esters. In this study, a continuous enzymatic glycolipid production catalyzed by Novozyme 435® is presented as an unprecedented concept. A biphasic aqueous–organic system was investigated, allowing for the simultaneous solubilization of sugars and fatty acids. Owing to phase separation, the remaining non-acylated glucose was easily separated from the product stream and was refed to the reactor forming a closed-loop system. Productivity in the continuous process was higher compared to a batch one, with space–time yields of up to 1228 ± 65 µmol/L/h. A temperature of 70 °C resulted in the highest glucose-6-O-decanoate concentration in the Packed Bed Reactor (PBR). Consequently, the design of a continuous biocatalytic production is a step towards a more competitive glycolipid synthesis in the aim for industrialization

Production strategies and applications of microbial single cell oils

Polyunsaturated fatty acids (PUFAs )of the ω-3 and ω-6 class (e.g. , α-linolenic acid, linoleic acid) are essential for maintaining biofunctions in mammalians like humans. Due to the fact that humans cannot synthesize these essential fatty acids, they must be taken up from different food sources. Classical sources for these fatty acids are porcine liver and fish oil. However, microbial lipids or single cell oils, produced by oleaginous microorganisms such as algae, fungi and bacteria, are a promising source as well. These single cell oils can be used form any valuable chemicals with applications not only for nutrition but also for fuels and are therefore an ideal basis for a bio-based economy. A crucial point for the establishment of microbial lipids utilization is the cost-effective production and purification of fuels or products of higher value. The fermentative production can be realized by submerged (SmF) or solid state fermentation (SSF). The yield and the composition of the obtained microbial lipids depend on the type of fermentation and the particular conditions (e.g., medium, pH-value, temperature, aeration, nitrogensource). From an economical point of view, waste or by-product streams can be used as cheap and renewable carbon and nitrogen sources. Ingeneral, downstream processing costs are one of the major obstacles to be solved for full economic efficiency of microbial lipids. For the extraction of lipids from microbial biomass cell disruption is most important, because efficiency of cell disruption directly influences subsequent downstream operations andoverall extraction efficiencies. A multitude of cell disruption and lipid extraction methods are available, conventional as well as newly emerging methods, which will be described and discussed in terms of large scale applicability, their potential in a modern biorefinery and their influence on product quality. Furthermore, an overview is given about applications of microbial lipids or derived fatty acids with emphasis on food applications

Foam-free Production of Surfactin via Anerobic Fermentation of Bacillus subtilis DSM 10T

Surfactin is one of the most popular biosurfactants due to its numerous potential applications. The usually aerobic production via fermentation of Bacillus subtilis is accompanied by vigorous foaming which leads to complex constructions and great expense. Therefore it is reasonable to search for alternative foam-free production processes. The current study introduces a novel approach to produce Surfactin in a foam-free process applying a strictly anaerobic bioreactor cultivation. The process was performed several times with different glucose concentrations in mineral salt medium. The fermentations were analyzed regarding specific (qSurfactin, vol. qSurfactin) and overall product yields (YP/X, YP/S) as well as substrate utilization (YX/S). Fermentations in which 2.5 g/L glucose were employed proofed to be the most effective, reaching product yields of YP/X = 0.278 g/g. Most interesting, the product yields exceeded classical aerobic fermentations, in which foam fractionation was applied. Additionally, values for specific production rate qSurfactin (0.005 g/(g∙h)) and product yield per consumed substrate (YP/S = 0.033 g/g) surpass results of comparable foam-free processes. The current study introduces an alternative to produce a biosurfactant that overcomes the challenges of severe foaming and need for additional constructions. © 2015, Willenbacher et al.; licensee Springer

Microwave-Assisted One-Pot Lipid Extraction and Glycolipid Production from Oleaginous Yeast Saitozyma podzolica in Sugar Alcohol-Based Media

Glycolipids are non-ionic surfactants occurring in numerous products of daily life. Due to their surface-activity, emulsifying properties, and foaming abilities, they can be applied in food, cosmetics, and pharmaceuticals. Enzymatic synthesis of glycolipids based on carbohydrates and free fatty acids or esters is often catalyzed using certain acyltransferases in reaction media of low water activity, e.g., organic solvents or notably Deep Eutectic Systems (DESs). Existing reports describing integrated processes for glycolipid production from renewables use many reaction steps, therefore this study aims at simplifying the procedure. By using microwave dielectric heating, DESs preparation was first accelerated considerably. A comparative study revealed a preparation time on average 16-fold faster than the conventional heating method in an incubator. Furthermore, lipids from robust oleaginous yeast biomass were successfully extracted up to 70% without using the pre-treatment method for cell disruption, limiting logically the energy input necessary for such process. Acidified DESs consisting of either xylitol or sorbitol and choline chloride mediated the one-pot process, allowing subsequent conversion of the lipids into mono-acylated palmitate, oleate, linoleate, and stearate sugar alcohol esters. Thus, we show strong evidence that addition of immobilized Candida antarctica Lipase B (Novozym 435®), in acidified DES mixture, enables a simplified and fast glycolipid synthesis using directly oleaginous yeast biomass

Optimization of glycolipid synthesis in hydrophilic deep eutectic solvents

Glycolipids are considered an alternative to petrochemically based surfactants because they are non-toxic, biodegradable, and less harmful to the environment while having comparable surface-active properties. They can be produced chemically or enzymatically in organic solvents or in deep eutectic solvents (DES) from renewable resources. DES are non-flammable, non-volatile, biodegradable, and almost non-toxic. Unlike organic solvents, sugars are easily soluble in hydrophilic DES. However, DES are highly viscous systems and restricted mass transfer is likely to be a major limiting factor for their application. Limiting factors for glycolipid synthesis in DES are not generally well understood. Therefore, the influence of external mass transfer, fatty acid concentration, and distribution on initial reaction velocity in two hydrophilic DES (choline:urea and choline:glucose) was investigated. At agitation speeds of and higher than 60 rpm, the viscosity of both DES did not limit external mass transfer. Fatty acid concentration of 0.5 M resulted in highest initial reaction velocity while higher concentrations had negative effects. Fatty acid accessibility was identified as a limiting factor for glycolipid synthesis in hydrophilic DES. Mean droplet sizes of fatty acid-DES emulsions can be significantly decreased by ultrasonic pretreatment resulting in significantly increased initial reaction velocity and yield (from 0.15 ± 0.03 μmol glucose monodecanoate/g DES to 0.57 ± 0.03 μmol/g) in the choline: urea DES. The study clearly indicates that fatty acid accessibility is a limiting factor in enzymatic glycolipid synthesis in DES. Furthermore, it was shown that physical pretreatment of fatty acid-DES emulsions is mandatory to improve the availability of fatty acids