BeerMIMS: Exploring the Use of Membrane-Inlet Mass Spectrometry (MIMS) Coupled to KNIME for the Characterization of Danish Beers

Beer is a complex mix of more than 7700 compounds, around 800 of which are volatile. While GC-MS has been actively employed in the analysis of the volatome of beer, this method is challenged by the complex nature of the sample. Herein, we explored the possible of using membrane-inlet mass spectrometry (MIMS) coupled to KNIME to characterize local Danish beers. KNIME stands for Konstanz Information Miner and is a free open-source data processing software which comes with several prebuilt nodes, that, when organized, result in data processing workflows allowing swift analysis of data with outputs that can be visualized in the desired format. KNIME has been shown to be promising in automation of large datasets and requires very little computing power. In fact, most of the computations can be carried out on a regular PC. Herein, we have utilized a KNIME workflow for data visualization of MIMS data to understand the global volatome of beers. Feature identification was not possible as of now but with a combination of MIMS and a KNIME workflow, we were able to distinguish beers from different micro-breweries located in Denmark, laying the foundation for the use of MIMS in future analysis of the beer volatome.

Nitrogen cycling in muddy sediments of Great Peconic Bay, USA: Seasonal N reaction balances and multi-year flux patterns

To better understand the capacity of sediments to serve as both source and sink of nitrogen (N) and to identify any evidence of evolving changes in sedimentary N cycling, N2 production, N remineralization, and N2 fixation were studied over a multi-year period (2010–2015) in bioturbated mud of Great Peconic Bay, a temperate northeastern U. S. estuary. Benthic fluxes and rates of organic matter remineralization were measured using in situ and ex situ incubations. Net annual NH+ 4, NO–3/NO–2, and N2–N fluxes (μ = 1.1, 0.03, and 1.2 mmol m –2d –1) were close to averages for comparable sedi- mentary environments from surveys of published field studies. Net N2 fluxes (by membrane inlet mass spectrometry) were influenced in different periods by temperature, oxygenation of sediment, pulsed Corg, and the activity of benthic macrofauna and benthic microalgae, although no single physical or biogeochemical variable showed a strong, direct relationship with net N2 fluxes over all sampling periods. In situ measurements sometimes showed more dynamic and higher amplitude diurnal N flux cycles than did ex situ incubations, suggesting ex situ incubations did not fully capture impacts of bioirrigation or benthic photosynthesis.15 N tracer experiments indicated anammox was < 7% of total N2 production. Acetylene reduction assays demonstrated C2 H4 production to depths ≥ 15 cm and suggested N2 fixation may have approached 25% of gross N2 production(3:1 C2 H4 : N2). Mass balances incorporating independently measured N remineralization estimates were consistent with measured levels of N2 fixation. Overall, complex balances of competing processes governed sedimentary N cycling seasonally, and N2 production dominated N2 fixation. Measured N2 fixation was consistent with constraints from N remineralization rates and net N fluxes except in episodic conditions (e. g., algal blooms). There was no indication of progressive changes in N cycling magnitudes or relative N reaction balances over the study period.

Water oxidation by photosystem II is the primary source of electrons for sustained H2photoproduction in nutrient-replete green algae

The unicellular green algaChlamydomonas reinhardtiiis capable of photosynthetic H2production. H2evolution occurs under anaerobic conditions and is difficult to sustain due to 1) competition between [FeFe]-hydrogenase (H2ase), the key enzyme responsible for H2metabolism in algae, and the Calvin–Benson–Bassham (CBB) cycle for photosynthetic reductants and 2) inactivation of H2ase by O2coevolved in photosynthesis. Recently, we achieved sustainable H2photoproduction by shifting algae from continuous illumination to a train of short (1 s) light pulses, interrupted by longer (9 s) dark periods. This illumination regime prevents activation of the CBB cycle and redirects photosynthetic electrons to H2ase. Employing membrane-inlet mass spectrometry andH218O, we now present clear evidence that efficient H2photoproduction in pulse-illuminated algae depends primarily on direct water biophotolysis, where water oxidation at the donor side of photosystem II (PSII) provides electrons for the reduction of protons by H2ase downstream of photosystem I. This occurs exclusively in the absence of CO2fixation, while with the activation of the CBB cycle by longer (8 s) light pulses the H2photoproduction ceases and instead a slow overall H2uptake is observed. We also demonstrate that the loss of PSII activity in DCMU-treated algae or in PSII-deficient mutant cells can be partly compensated for by the indirect (PSII-independent) H2photoproduction pathway, but only for a short (<1 h) period. Thus, PSII activity is indispensable for a sustained process, where it is responsible for more than 92% of the final H2yield.

Membrane Inlet Mass Spectrometry: A Powerful Tool for Algal Research

Since the first great oxygenation event, photosynthetic microorganisms have continuously shaped the Earth’s atmosphere. Studying biological mechanisms involved in the interaction between microalgae and cyanobacteria with the Earth’s atmosphere requires the monitoring of gas exchange. Membrane inlet mass spectrometry (MIMS) has been developed in the early 1960s to study gas exchange mechanisms of photosynthetic cells. It has since played an important role in investigating various cellular processes that involve gaseous compounds (O2, CO2, NO, or H2) and in characterizing enzymatic activities in vitro or in vivo. With the development of affordable mass spectrometers, MIMS is gaining wide popularity and is now used by an increasing number of laboratories. However, it still requires an important theory and practical considerations to be used. Here, we provide a practical guide describing the current technical basis of a MIMS setup and the general principles of data processing. We further review how MIMS can be used to study various aspects of algal research and discuss how MIMS will be useful in addressing future scientific challenges.

A Chlorophyte alga utilizes alternative electron transport for primary photoprotection

Desmodesmus armatus is an emerging biofuel platform producing high amount of lipids and biomass in mass culture. We observed D. armatus in light-limiting, excess light and sinusoidal light environments to investigate its photoacclimation behaviors and the mechanisms by which it dissipates excess energy. Chlorophyll a:b ratios and the functional absorption cross section of photosystem II (PSII) suggested a constitutively small light harvesting antenna size relative to other green algae. In situ and ex situ measurements of photo-physiology revealed that nonphotochemical quenching (NPQ) is not a significant contributor to photoprotection, but cells do not suffer substantial photoinhibition despite its near absence. We performed membrane inlet mass spectrometry analysis to show that D. armatus has a very high capacity for alternative electron transport (AET) measured as light dependent oxygen consumption. Up to 90% of electrons generated at PSII can be dissipated by AET in a water-water cycle during growth in rapidly fluctuating light environments like those found in industrial scale photobioreactors. This work highlights the diversity of photoprotective mechanisms shown in algal systems, that NPQ is not necessarily required for effective photoprotection in some algae and suggests that engineering AET may be an attractive target for increasing biomass productivity of some strains.One-sentence summaryConstitutive small antennae, alternative electron transport and an efficient photosystem II turnover capacity enable D. armatus to photosynthesize efficiently.