Новый высокопродуктивный штамм Propionibacterium acidipropionici FL-48 с повышенной устойчивостью к пропионовой кислоте и масштабирование технологии его наработки в промышленных биореакторах

Propionic acid bacteria, including Propionibacterium acidipropionici, are widely used in the chemical industry to produce propionic acid and also for food and feed preservation. However, the efficiency of the industrial production of these bacteria is limited by their sensitivity to high concentrations of propionic acid excreted into the cultivation medium. Therefore, the development of new biotechnological processes and strains able to overcome this limitation and to improve the profitability of the microbiological production remains  a relevant problem. A new P. acidipropionici FL-48 strain characterized by an increased resistance to 10 g/L of propionic acid (the number of viable cells after 24-h cultivation reached 1.05 × 106) was developed by a two-step induced mutagenesis using UV and diethyl sulphate from the P. acidipropionici VKPM B-5723 strain. The mutant strain exceeded the parental strain in the biomass accumulation rate and the amount of produced propionic and acetic acids by 35%, 20%, and 16%, respectively. The stability of such important characteristics as the biomass accumulation rate and the viability on media containing heightened concentrations of propionic acid was confirmed by three sequential monoclonal subculturings on a medium supplemented with 10 g/L of propionic acid. The optimization of the cultivation technology made it possible to determine the optimum seed inoculum dose (10% of the fermentation medium volume) and the best pH level for the active growth stage (6.1 ± 0.1). The scaling up of the fermentation to a 100-L bioreactor under observance of optimum cultivation conditions demonstrated a high biomass growth rate with a sufficient reproducability; after 20 h of fermentation, the number of viable cells in the culture broth exceeded 1 × 1010 CFU/mL. The new strain could be interesting as the component of silage and haylage biopreservatives and also could be used as an efficient producer of propionic acid.Пропионовокислые бактерии, в том числе Propionibacterium acidipropionici, широко используются в химической промышленности для получения пропионовой кислоты, а также для консервирования пищи и заготовки зерна и зеленых кормов. Однако эффективность промышленного производства биомассы пропионовокислых бактерий ограничена их чувствительностью к высоким концентрациям в среде пропионовой кислоты. Таким образом, актуальной задачей является разработка новых биотехнологических процессов и штаммов, позволяющих преодолеть это ограничение и повысить рентабельность микробиологического производства. Методом двухступенчатого индуцированного мутагенеза с применением УФ-облучения и диэтилсульфата получен новый мутантный штамм P. acidipropionici ФЛ-48, обладающий повышенной резистентностью к 10 г/л пропионовой кислоты (количество жизнеспособных клеток через 24 ч культивирования достигало 1,05 × 106) и превосходящий родительский штамм P. acidipropionici ВКПМ В-5723 по скорости накопления биомассы и количеству продуцируемых пропионовой и уксусной кислот на 35%, 20% и 16%, соответственно. Стабильность характеристик нового штамма (скорость накопления биомассы и жизнеспособность на средах с повышенной концентрацией пропионовой кислоты) подтверждена трехкратным последовательным моноклональным рассевом на среду, содержащую 10 г/л пропионовой кислоты. Выполненная оптимизация технологии культивирования штамма позволила определить оптимальную дозу инокулюма для засева биореактора (10% от объема ферментационной среды) и поддерживаемый в течение первых 12 ч уровень рН среды, обеспечивающий максимальный прирост биомассы (6,1 ± 0,1). Проведенное масштабирование ферментации до 100-литрового биореактора с соблюдением оптимальных условий культивирования показало сохранение высокой скорости роста штамма в условиях пониженного рН; уже к 20-му часу ферментации количество жизнеспособных клеток в культуральной жидкости превышало 1 × 1010 КОЕ/мл. Полученные результаты показали хорошую воспроизводимость. Новый штамм представляет интерес в качестве компонента биоконсервантов для силоса и сенажа, а также может быть использован в качестве эффективного продуцента пропионовой кислоты. Пропионовокислые бактерии, в том числе Propionibacterium acidipropionici, широко используются в химической промышленности для получения пропионовой кислоты, а также для консервирования пищи и заготовки зерна и зеленых кормов. Однако эффективность промышленного производства биомассы пропионовокислых бактерий ограничена их чувствительностью к высоким концентрациям в среде пропионовой кислоты. Таким образом, актуальной задачей является разработка новых биотехнологических процессов и штаммов, позволяющих преодолеть это ограничение и повысить рентабельность микробиологического производства. Методом двухступенчатого индуцированного мутагенеза с применением УФ-облучения и диэтилсульфата получен новый мутантный штамм P. acidipropionici ФЛ-48, обладающий повышенной резистентностью к 10 г/л пропионовой кислоты (количество жизнеспособных клеток через 24 ч культивирования достигало 1,05 × 106) и превосходящий родительский штамм P. acidipropionici ВКПМ В-5723 по скорости накопления биомассы и количеству продуцируемых пропионовой и уксусной кислот на 35%, 20% и 16%, соответственно. Стабильность характеристик нового штамма (скорость накопления биомассы и жизнеспособность на средах с повышенной концентрацией пропионовой кислоты) подтверждена трехкратным последовательным моноклональным рассевом на среду, содержащую 10 г/л пропионовой кислоты. Выполненная оптимизация технологии культивирования штамма позволила определить оптимальную дозу инокулюма для засева биореактора (10% от объема ферментационной среды) и поддерживаемый в течение первых 12 ч уровень рН среды, обеспечивающий максимальный прирост биомассы (6,1 ± 0,1). Проведенное масштабирование ферментации до 100-литрового биореактора с соблюдением оптимальных условий культивирования показало сохранение высокой скорости роста штамма в условиях пониженного рН; уже к 20-му часу ферментации количество жизнеспособных клеток в культуральной жидкости превышало 1 × 1010 КОЕ/мл. Полученные результаты показали хорошую воспроизводимость. Новый штамм представляет интерес в качестве компонента биоконсервантов для силоса и сенажа, а также может быть использован в качестве эффективного продуцента пропионовой кислоты.

Nanostructured 3D Constructs Based on Chitosan and Chondroitin Sulphate Multilayers for Cartilage Tissue Engineering

Nanostructured three-dimensional constructs combining layer-by-layer technology (LbL) and template leaching were processed and evaluated as possible support structures for cartilage tissue engineering. Multilayered constructs were formed by depositing the polyelectrolytes chitosan (CHT) and chondroitin sulphate (CS) on either bidimensional glass surfaces or 3D packet of paraffin spheres. 2D CHT/CS multi-layered constructs proved to support the attachment and proliferation of bovine chondrocytes (BCH). The technology was transposed to 3D level and CHT/CS multi-layered hierarchical scaffolds were retrieved after paraffin leaching. The obtained nanostructured 3D constructs had a high porosity and water uptake capacity of about 300%. Dynamical mechanical analysis (DMA) showed the viscoelastic nature of the scaffolds. Cellular tests were performed with the culture of BCH and multipotent bone marrow derived stromal cells (hMSCs) up to 21 days in chondrogenic differentiation media. Together with scanning electronic microscopy analysis, viability tests and DNA quantification, our results clearly showed that cells attached, proliferated and were metabolically active over the entire scaffold. Cartilaginous extracellular matrix (ECM) formation was further assessed and results showed that GAG secretion occurred indicating the maintenance of the chondrogenic phenotype and the chondrogenic differentiation of hMSCs

Transesterification of PHA to Oligomers Covalently Bonded with (Bio)Active Compounds Containing Either Carboxyl or Hydroxyl Functionalities

© 2015 The Authors. Published by Public Library of Science. This is an open access article available under a Creative Commons licence. The published version can be accessed at the following link on the publisher’s website: https://doi.org/10.1371/journal.pone.0120149This manuscript presents the synthesis and structural characterisation of novel biodegradable polymeric controlled-release systems of pesticides with potentially higher resistance to weather conditions in comparison to conventional forms of pesticides. Two methods for the preparation of pesticide-oligomer conjugates using the transesterification reaction were developed. The first method of obtaining conjugates, which consist of bioactive compounds with the carboxyl group and polyhydroxyalkanoates (PHAs) oligomers, is "one-pot" transesterification. In the second method, conjugates of bioactive compounds with hydroxyl group and polyhydroxyalkanoates oligomers were obtained in two-step method, through cyclic poly(3-hydroxybutyrate) oligomers. The obtained pesticide-PHA conjugates were comprehensively characterised using GPC, 1H NMR and mass spectrometry techniques. The structural characterisation of the obtained products at the molecular level with the aid of mass spectrometry confirmed that both of the synthetic strategies employed led to the formation of conjugates in which selected pesticides were covalently bonded to PHA oligomers via a hydrolysable ester bond

Role of Caustic Addition in Bitumen-Clay Interactions

Coating of bitumen by clays, known as slime coating, is detrimental to bitumen recovery from oil sands using the warm slurry extn. process. Sodium hydroxide (caustic) is added to the extn. process to balance many competing processing challenges, which include undesirable slime coating. The current research aims at understanding the role of caustic addn. in controlling interactions of bitumen with various types of model clays. The interaction potential was studied by quartz crystal microbalance with dissipation monitoring (QCM-D). After confirming the slime coating potential of montmorillonite clays on bitumen in the presence of calcium ions, the interaction of kaolinite and illite with bitumen was studied. To represent more closely the industrial applications, tailings water from bitumen extn. tests at different caustic dosage was used. At caustic dosage up to 0.5 wt % oil sands ore, a negligible coating of kaolinite on the bitumen was detd. However, at a lower level of caustic addn., illite was shown to attach to the bitumen, with the interaction potential decreasing with increasing caustic dosage. Increasing concn. of humic acids as a result of increasing caustic dosage was identified to limit the interaction potential of illite with bitumen. This fundamental study clearly shows that the crit. role of caustics in modulating interactions of clays with bitumen depends upon the type of clays. Thus, clay type was identified as a key operational variable

Membrane chemical stability and seed longevity

Here, we investigate the relationships between the chemical stability of the membrane surface and seed longevity. Dry embryos of long-lived tomato and short-lived onion seeds were labeled with 5-doxyl-stearic acid (5-DS). Temperature-induced loss of the electron spin resonance signal caused by chemical conversion of 5-DS to nonparamagnetic species was used to characterize the membrane surface chemical stability. No difference was found between temperature plots of 5-DS signal intensity in dry onion and tomato below 345 K. Above this temperature, the 5-DS signal remained unchanged in tomato embryos and irreversibly disappeared in onion seeds. The role of the physical state and chemical status of the membrane environment in the chemical stability of membrane surfaces was estimated for model systems containing 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) dried alone or in the presence of trehalose or glucose. Fourier transform infrared spectroscopy was used to follow temperature-induced structural changes in dry POPC. Spin-label technique was used to relate the chemical stability of 5-DS with the dynamic properties of the bilayer and 5-DS motion behavior. In all the models, the decrease in 5-DS signal intensity was always observed above Tm for the membrane surface. The 5-DS signal was irreversibly lost at high temperature when dry POPC was embedded in a glucose matrix. The loss of 5-DS signal was moderate when POPC was dried alone or in the presence of trehalose. Comparison of model and in vivo data shows that the differences in longevity between onion and tomato seeds are caused by differences in the chemical status of the membrane surface rather than the degree of its immobilization

Northern Eurasia Future Initiative (NEFI): facing the challenges and pathways of global change in the 21st century

During the past several decades, the Earth system has changed significantly, especially across Northern Eurasia. Changes in the socio-economic conditions of the larger countries in the region have also resulted in a variety of regional environmental changes that can have global consequences. The Northern Eurasia Future Initiative (NEFI) has been designed as an essential continuation of the Northern Eurasia Earth Science Partnership Initiative (NEESPI), which was launched in 2004. NEESPI sought to elucidate all aspects of ongoing environmental change, to inform societies and, thus, to better prepare societies for future developments. A key principle of NEFI is that these developments must now be secured through science-based strategies co-designed with regional decision makers to lead their societies to prosperity in the face of environmental and institutional challenges. NEESPI scientific research, data, and models have created a solid knowledge base to support the NEFI program. This paper presents the NEFI research vision consensus based on that knowledge. It provides the reader with samples of recent accomplishments in regional studies and formulates new NEFI science questions. To address these questions, nine research foci are identified and their selections are briefly justified. These foci include: warming of the Arctic; changing frequency, pattern, and intensity of extreme and inclement environmental conditions; retreat of the cryosphere; changes in terrestrial water cycles; changes in the biosphere; pressures on land-use; changes in infrastructure; societal actions in response to environmental change; and quantification of Northern Eurasia's role in the global Earth system. Powerful feedbacks between the Earth and human systems in Northern Eurasia (e.g., mega-fires, droughts, depletion of the cryosphere essential for water supply, retreat of sea ice) result from past and current human activities (e.g., large scale water withdrawals, land use and governance change) and potentially restrict or provide new opportunities for future human activities. Therefore, we propose that Integrated Assessment Models are needed as the final stage of global change assessment. The overarching goal of this NEFI modeling effort will enable evaluation of economic decisions in response to changing environmental conditions and justification of mitigation and adaptation efforts