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Plasma biology

Plasma-driven biocatalysis

A research team from Bochum has developed a new method to drive catalytically active enzymes.

Compared with traditional chemical methods, enzyme catalysis has numerous advantages. But it also has weaknesses. Some enzymes are not very stable. Enzymes that convert hydrogen peroxide are even inactivated by high concentrations of the substrate. A research team at Ruhr-Universität Bochum (RUB), together with international partners, has developed a process in which the starting material, i.e. hydrogen peroxide, is fed to the biocatalysts in a controlled manner using plasma. The enzymes themselves are protected from harmful components of the plasma by a buffer layer. Using two model enzymes, the team showed that the process works, as reported in the journal “ChemSusChem” from 5 February 2020.

Milder conditions, less energy consumption and waste

In biocatalysis, chemicals are produced by cells or their components, in particular by enzymes. Biocatalysis has many advantages over traditional chemical processes: the reaction conditions are usually much milder, energy consumption is lower and less toxic waste is produced. The high specificity of enzymes also means that fewer side reactions occur. Moreover, some fine chemicals can only be synthesised by biocatalysis.

The weak spot of enzyme biocatalysis is the low stability of some enzymes. “Since the enzyme often has to be replaced in such cases – which is expensive – it is extremely important to increase the stability under production conditions,” explains lead author Abdulkadir Yayci from the Chair of Applied Microbiology headed by Professor Julia Bandow.

Hydrogen peroxide: necessary, but harmful

The research team has been studying two similar classes of enzymes: peroxidases and peroxygenases. Both use hydrogen peroxide as a starting material for oxidations. The crucial problem is that hydrogen peroxide is absolutely necessary for activity, but in higher concentrations it leads to a loss of activity of the enzymes. As far as these enzyme classes are concerned, it is therefore vital to supply hydrogen peroxide in precise doses.

To this end, the researchers investigated plasmas as a source of hydrogen peroxide. Plasma describes the fourth state of matter that is created when energy is added to a gas. If liquids are treated with plasmas, a large number of reactive oxygen and nitrogen species are formed, some of which then react to form long-lived hydrogen peroxide, which can be used for biocatalysis.

Biocatalytic reactions with plasma-generated hydrogen peroxide are possible

In an experiment in which horseradish peroxidase served as one of the model enzymes, the team showed that this system works in principle. At the same time, the researchers identified the weak points of plasma treatment: “Plasma treatment also directly attacks and inactivates the enzymes, most likely through the highly reactive, short-lived species in the plasma-treated liquid,” outlines Abdulkadir Yayci. The research group improved the reaction conditions by binding the enzyme to an inert carrier material. This creates a buffer zone above the enzyme in which the highly reactive plasma species can react without harming the enzyme.

The researchers then tested their approach using a second enzyme, the unspecific peroxygenase from the fungus Agrocybe aegerita. This peroxygenase has the ability to oxidise a large number of substrates in a highly selective way. “We successfully demonstrated that this specificity is maintained even under plasma treatment and that highly selective biocatalytic reactions are possible using plasma,” concludes Julia Bandow.

written by Marike Drießen, RUB

Press release

DFG funded project

PLASNOW (Plasma generated Nitric Oxide in Wound healing)

The DFG granted research project PLASNOW (Plasma generated Nitric Oxide in Wound healing) is an interdisciplinary collaboration within the research field of plasma medicine. The project was approved in November 2019 for 36 months. It will start with the start it will only start with the start of contract of the PhD student which is still in process at the moment.

Two groups from electrical engineering (AEPT, Prof. Dr. P. Awakowicz) and plasma physics (Experimental physics II, Dr. V. Schulz-von der Gathen) are involved. It is a successor and continuation of projects that were beforehand bundled in the cooperation “Plasma2Cell“. In this cooperation, other groups participate e.g. from chemistry, medicine, and biology at the Ruhr-University, the Heinrich-Heine-University in Düsseldorf and the DLR in Cologne.

PLASMA RESEARCH

Lightning bolt underwater

A plasma tears through the water within a few nanoseconds. It may possibly regenerate catalytic surfaces at the push of a button.

Electrochemical cells help recycle CO₂. However, the catalytic surfaces get worn down in the process. Researchers at the Collaborative Research Centre 1316 “Transient atmospheric plasmas: from plasmas to liquids to solids” at Ruhr-Universität Bochum (RUB) are exploring how they might be regenerated at the push of a button using extreme plasmas in water. In a first, they deployed optical spectroscopy and modelling to analyse such underwater plasmas in detail, which exist only for a few nanoseconds, and to theoretically describe the conditions during plasma ignition. They published their report in the journal Plasma Sources Science and Technology on 4 June 2019.

A plasma tears through the water within a few nanoseconds. Following plasma ignition, there is a high negative pressure difference at the tip of the electrode, which results in ruptures forming in the liquid. Plasma then spreads across those ruptures.

Video: Experimentalphysik II

Plasmas are ionised gases: they are formed when a gas is energised that then contains free electrons. In nature, plasmas occur inside stars or take the shape of polar lights on Earth. In engineering, plasmas are utilised for example to generate light in fluorescent lamps, or to manufacture new materials in the field of microelectronics. “Typically, plasmas are generated in the gas phase, for example in the air or in noble gases,” explains Katharina Grosse from the Institute for Experimental Physics II at RUB.

Ruptures in the water

In the current study, the researchers have generated plasmas directly in a liquid. To this end, they applied a high voltage to a submerged hairline electrode for the range of several billionth seconds. Following plasma ignition, there is a high negative pressure difference at the tip of the electrode, which results in ruptures forming in the liquid. Plasma then spreads across those ruptures. “Plasma can be compared with a lightning bolt – only in this case it happens underwater,” says Katharina Grosse.

Hotter than the sun

Using fast optical spectroscopy in combination with a fluid dynamics model, the research team identified the variations of power, pressure, and temperature in these plasmas. “In the process, we observed that the consumption inside these plasmas briefly amounts to up to 100 kilowatt. This corresponds with the connected load of several single-family homes,” points out Professor Achim von Keudell from the Institute for Experimental Physics II. In addition, pressures exceeding several thousand bars are generated – corresponding with or even exceeding the pressure at the deepest part of the Pacific Ocean. Finally, there are short bursts of temperatures of several thousand degrees, which roughly equal and even surpass the surface temperature of the sun.

Water is broken down into its components

Such extreme conditions last only for a very short time. “Studies to date had primarily focused on underwater plasmas in the microsecond range,” explains Katharina Grosse. “In that space of time, water molecules have the chance to compensate for the pressure of the plasma.” The extreme plasmas that have been the subject of the current study feature much faster processes. The water can’t compensate for the pressure and the molecules are broken down into their components. “The oxygen that is thus released plays a vital role for catalytic surfaces in electrochemical cells,” explains Katharina Grosse. “By re-oxidating such surfaces, it helps them regenerate and take up their full catalytic activity again. Moreover, reagents dissolved in water can also be activated, thus facilitating catalysis processes.”

By Meike Drießen, Translated by Donata Zuber

Recent research achievement

How bacteria protect themselves from plasma treatment

Plasmas are applied in the treatment of wounds to combat pathogens that are resistant against antibiotics. But bacteria know how to defend themselves.

Considering the ever-growing percentage of bacteria that are resistant to antibiotics, interest in medical use of plasma is increasing. In collaboration with colleagues from Kiel, researchers at Ruhr-Universität Bochum (RUB) investigated if bacteria may become impervious to plasmas, too. They identified 87 genes of the bacterium Escherichia coli, which potentially protect against effective components of plasma. “These genes provide insights into the antibacterial mechanisms of plasmas,” says Marco Krewing. He is the lead author of two articles that were published in the Journal of the Royal Society Interface this year.

A cocktail of harmful components stresses pathogens

Plasmas are created from gas that is pumped with energy. Today, plasmas are already used against multi-resistant pathogens in clinical applications, for example to treat chronic wounds. “Plasmas provide a complex cocktail of components, many of which act as disinfectants in their own right,” explains Professor Julia Bandow, Head of the RUB research group Applied Microbiology. UV radiation, electric fields, atomic oxygen, superoxide, nitric oxides, ozone, and excited oxygen or nitrogen affect the pathogens simultaneously, generating considerable stress. Typically, the pathogens survive merely several seconds or minutes.

In order to find out if bacteria, may develop resistance against the effects of plasmas, like they do against antibiotics, the researchers analysed the entire genome of the model bacterium Escherichia coli, short E. coli, to identify existing protective mechanisms. “Resistance means that a genetic change causes organisms to be better adapted to certain environmental conditions. Such a trait can be passed on from one generation to the next,” explains Julia Bandow.

Mutants missing single genes

For their study, the researchers made use of so-called knockout strains of E. coli. These are bacteria that are missing one specific gene in their genome, which contains approximately 4,000 genes. The researchers exposed each mutant to the plasma and monitored if the cells kept proliferating following the exposure.

“We demonstrated that 87 of the knockout strains were more sensitive to plasma treatment than the wild type that has a complete genome,” says Marco Krewing. Subsequently, the researchers analysed the genes missing in these 87 strains and determined that most of those genes protected bacteria against the effects of hydrogen peroxide, superoxide, and/or nitric oxide. “This means that these plasma components are particularly effective against bacteria,” elaborates Julia Bandow. However, it also means that genetic changes that result in an increase in the number or activity of the respective gene products are more capable of protecting bacteria from the effects of plasma treatment.

Heat shock protein boosts plasma resistance

The research team, in collaboration with a group headed by Professor Ursula Jakob from the University of Michigan in Ann Arbor (USA), demonstrated that this is indeed the case: the heat shock protein Hsp33, encoded by the hslO gene, protects E. coli proteins from aggregation when exposed to oxidative stress. “During plasma treatment, this protein is activated and protects the other E. coliproteins – and consequently the bacterial cell,” Bandow points out. An increased volume of this protein alone results in a slightly increased plasma resistance. Considerably stronger plasma resistance can be expected when the levels of several protective proteins are increased simultaneously.

By Meike Drießen, Translated by Donata Zuber

Press releases

New class of catalysts for energy conversion

The research group of Prof. Ludwig and his colleagues recently published their new results in the catalyst production. “At our department, we have unique methods at our disposal to manufacture these complex materials from five source elements in different compositions in form of thin film or nanoparticle libraries,” explains Professor Alfred Ludwig from the Chair of Materials for Microtechnology at RUB. The atoms of the source elements blend in plasma and form nanoparticles in a substrate of ionic liquid. If the nanoparticles are located in the vicinity of the respective atom source, the percentage of atoms from that source is higher in the respective particle. “Very limited research has as yet been conducted into the usage of such materials in electrocatalysis,” says Ludwig.

The full press releas can be found here.