Unveiling the ecological resilience and industrial potential of Skeletonema marinoi through mixotrophic cultivation in Nordic winter condition.

Mixotrophy, the concurrent use of inorganic and organic carbon in the presence of light for microalgal growth, holds ecological and industrial significance. However, it is poorly explored in diatoms, especially in ecologically relevant species like Skeletonema marinoi. This study strategically employed mixotrophic metabolism to optimize the growth of a strain of Skeletonema marinoi (Sm142), which was found potentially important for biomass production on the west coast of Sweden in winter conditions. The aim of this study was to discern the most effective organic carbon sources by closely monitoring microalgal growth through the assessment of optical density, chlorophyll a fluorescence, and biomass concentration. The impact of various carbon sources on the physiology of Sm142 was investigated using photosynthetic and respiratory parameters. The findings revealed that glycerol exhibited the highest potential for enhancing the biomass concentration of Sm142 in a multi-cultivator under the specified experimental conditions, thanks to the increase in respiration activity. Furthermore, the stimulatory effect of glycerol was confirmed at a larger scale using environmental photobioreactors simulating the winter conditions on the west coast of Sweden; it was found comparable to the stimulation by CO2-enriched air versus normal air. These results were the first evidence of the ability of Skeletonema marinoi to perform mixotrophic metabolism during the winter and could explain the ecological success of this diatom on the Swedish west coast. These findings also highlight the importance of both organic and inorganic carbon sources for enhancing biomass productivity in harsh winter conditions.

Mixotrophy in a Local Strain of Nannochloropsis granulata for Renewable High-Value Biomass Production on the West Coast of Sweden

A local strain of Nannochloropsis granulata (Ng) has been reported as the most productive microalgal strain in terms of both biomass yield and lipid content when cultivated in photobioreactors that simulate the light and temperature conditions during the summer on the west coast of Sweden. To further increase the biomass and the biotechnological potential of this strain in these conditions, mixotrophic growth (i.e., the simultaneous use of photosynthesis and respiration) with glycerol as an external carbon source was investigated in this study and compared with phototrophic growth that made use of air enriched with 1-2% CO2. The addition of either glycerol or CO2-enriched air stimulated the growth of Ng and theproduction of high-value long-chain polyunsaturated fatty acids (EPA) as well as the carotenoid canthaxanthin. Bioassays in human prostate cell lines indicated the highest antitumoral activity for Ng extracts and fractions from mixotrophic conditions. Metabolomics detected betaine lipids specifically in the bioactive fractions, suggesting their involvement in the observed antitumoral effect. Genes related to autophagy were found to be upregulated by the most bioactive fraction, suggesting a possible therapeutic target against prostate cancer progression. Taken together, our results suggest that the local Ng strain can be cultivated mixotrophically in summer conditions on the west coast of Sweden for the production of high-value biomass containing antiproliferative compounds, carotenoids, and EPA.

Marine microalgae for outdoor biomass production-A laboratory study simulating seasonal light and temperature for the west coast of Sweden.

At Nordic latitudes, year-round outdoor cultivation of microalgae is debatable due to seasonal variations in productivity. Shall the same species/strains be used throughout the year, or shall seasonal-adapted ones be used? To elucidate this, a laboratory study was performed where two out of 167 marine microalgal strains were selected for intended cultivation at the west coast of Sweden. The two local strains belong to Nannochloropsis granulata (Ng) and Skeletonema marinoi (Sm142). They were cultivated in photobioreactors and compared in conditions simulating variations in light and temperature of a year divided into three growth seasons (spring, summer and winter). The strains grew similarly well in summer (and also in spring), but Ng produced more biomass (0.225 vs. 0.066 g DW L-1 day-1 ) which was more energy rich (25.0 vs. 16.6 MJ kg-1 DW). In winter, Sm142 grew faster and produced more biomass (0.017 vs. 0.007 g DW L-1 day-1 ), having similar energy to the other seasons. The higher energy of the Ng biomass is attributed to a higher lipid content (40 vs. 16% in summer). The biomass of both strains was richest in proteins (65%) in spring. In all seasons, Sm142 was more effective in removing phosphorus from the cultivation medium (6.58 vs. 4.14 mg L-1 day-1 in summer), whereas Ng was more effective in removing nitrogen only in summer (55.0 vs. 30.8 mg L-1 day-1 ). Our results suggest that, depending on the purpose, either the same or different local species can be cultivated, and are relevant when designing outdoor studies.

Microalgae biotechnology in Nordic countries - the potential of local strains.

Climate change, energy use and food security are the main challenges that our society is facing nowadays. Biofuels and feedstock from microalgae can be part of the solution if high and continuous production is to be ensured. This could be attained in year-round, low cost, outdoor cultivation systems using strains that are not only champion producers of desired compounds but also have robust growth in a dynamic climate. Using microalgae strains adapted to the local conditions may be advantageous particularly in Nordic countries. Here, we review the current status of laboratory and outdoor-scale cultivation in Nordic conditions of local strains for biofuel, high-value compounds and water remediation. Strains suitable for biotechnological purposes were identified from the large and diverse pool represented by saline (NE Atlantic Ocean), brackish (Baltic Sea) and fresh water (lakes and rivers) sources. Energy-efficient annual rotation for cultivation of strains well adapted to Nordic climate has the potential to provide high biomass yields for biotechnological purposes.

Development of a novel in situ gelling skin dressing: Delivering high levels of dissolved oxygen at pH 5.5.

BACKGROUND AND AIMS: Wound healing requires appropriate oxygen and pH levels. Oxygen therapy and pH-modulating treatments have shown positive effects on wound healing. Thus, a dressing, which combines high levels of dissolved oxygen (DO) with the pH of intact skin, may improve wound healing. Our aims were to (1) formulate an in situ gelling dressing with high levels of DO and with the pH level of intact skin, (2) evaluate physical and chemical properties of the dressing, and (3) elucidate basic effects of elevated levels of DO on human skin cells in vitro. METHODS: A dressing was formulated with 15 to 16 wt% poloxamer 407, acetate buffer, and oxygenated water. Stability of pH and DO, rheology, and shelf life were analysed. Furthermore, in vitro studies of the effect of increased levels of DO were performed. RESULTS: An in situ gelling wound dressing, with a DO concentration ranging between 25 and 35 mg/L and a pH of 5.5, was formulated. The DO concentration was stable above 22 mg/L for at least 30 hours when applied on a surface at 35°C and covered for directed diffusion into the intended wound area. At storage, the dressing had stable pH for 3 months and stable DO concentration over 30 mg/L for 7 weeks. Increasing DO significantly enhanced intracellular ATP in human skin cells, without changing reactive oxygen species production, proliferation rate, or viability. CONCLUSION: The developed dressing may facilitate wound healing by delivering controlled and stable oxygen levels, providing adjustable pH for optimized healing, and increasing intracellular ATP availability.

Digital colour tone for fluorescence sensing: a direct comparison of intensity, ratiometric and hue based quantification.

The hue parameter of HSV colour-space for digital imaging is shown to be accessible for convenient quantitative fluorescence imaging. A commercially available pH probe was utilized in solution and incorporated into optical µ-sensors for microscopy applications.

Diffusion consistent calibrations for improved chemical imaging using nanoparticle enhanced optical sensors.

A basic square root function was successfully used as a diffusion consistent calibration function that considers depletion mechanisms often occurring within optical chemical sensors. This continuous function improved image quality and simplified the calibration process. It may be a universal tool for the typical response function of reversible diffusion controlled sensing reactions. Further, we demonstrate that the gold nanoparticle interaction based ammonium fluorosensor is suitable for non-invasive high-resolution quantitative imaging of complex samples. The plasmon sensitized optical sensors were utilized as a bioanalytical tool for chemical imaging of natural degradation processes occurring in biological tissues. Analytical performance of the nanoparticle enhanced sensors confirmed superior sensitivity, reversibility, durability and overall image quality over non-doped sensing membranes. Although applied in a complex matrix of high potassium (major interferent) and very high sodium (interferent) excellent performance is achieved. The nanoparticle interaction/coextraction based sensing scheme utilized in this study is general and can be used for numerous ions, preferably combined with the diffusion consistent calibrations for superior analytical performance. A table with 44 commercially available ionophores is provided to guide potential users of this sensor configuration.

Plasmophore sensitized imaging of ammonia release from biological tissues using optodes.

A plasmophore sensitized optode was developed for imaging ammonia (NH(3)) concentrations in muscle tissues. The developed ammonia sensor and an equivalent non plasmophore version of the sensor were tested side by side to compare their limit of detection, dynamic range, reversibility and overall imaging quality. Bio-degradation patterns of ammonia release from lean porcine skeletal muscle were studied over a period of 11 days. We demonstrate that ammonia concentrations ranging from 10nM can be quantified reversibly with an optical resolution of 127 µm in a sample area of 25 mm × 35 mm. The plasmophore ammonia optode showed improved reversibility, less false pixels and a 2 nM ammonia detection limit compared to 200 nM for the non-plasmophore sensor. Main principles of the sensing mechanism include ammonia transfer over a gas permeable film, ammonia protonation, nonactin facilitated merocyanine-ammonium coextraction and plasmophore enhancement. The vast signal improvement is suggested to rely on solvatochroism, nanoparticle scattering and plasmonic interactions that are utilized constructively in a fluorescence ratio. In addition to fundamental medicinal and biological research applications in tissue physiology, reversible ammonia quantification will be possible for a majority of demanding imaging and non imaging applications such as monitoring of low ammonia background concentrations in air and non-invasive medicinal diagnosis through medical breath or saliva analysis. The nanoparticle doped sensor constitutes a highly competitive technique for ammonia sensing in complex matrixes and the general sensing scheme offers new possibilities for the development of artificial optical noses and tongues.

Ruthenium(II) Complex Enantiomers as Cellular Probes for Diastereomeric Interactions in Confocal and Fluorescence Lifetime Imaging Microscopy.

Ruthenium dipyridophenazine (dppz) complexes are sensitive luminescent probes for hydrophobic environments. Here, we apply multiple-frequency fluorescence lifetime imaging microscopy (FLIM) to Δ and Λ enantiomers of lipophilic ruthenium dppz complexes in live and fixed cells, and their different lifetime staining patterns are related to conventional intensity-based microscopy. Excited state lifetimes of the enantiomers determined from FLIM measurements correspond well with spectroscopically measured emission decay curves in pure microenvironments of DNA, phospholipid membrane or a model protein. We show that FLIM can be applied to monitor the long-lived excited states of ruthenium complex enantiomers and, combined with confocal microscopy, give new insight into their biomolecular binding and reveal differences in the microenvironment probed by the complexes.

Plasmonic nanoparticle interactions for high-performance imaging fluorosensors.

We demonstrate a nanoparticle based ratiometric fluorosensor suitable for non-invasive high-resolution imaging of complex samples. Coextraction generates a movement of a solvatochromic dye, with a consequent shift from plasmophore coupled emission towards longer wavelength scattering enhanced emission. The nanoparticle doped sensors confirmed superior sensitivity, reversibility, durability and overall image quality.

An imaging pH optode for cell studies based on covalent attachment of 8-hydroxypyrene-1,3,6-trisulfonate to amino cellulose acetate films.

The objective of this study was to create a thin film optode for fast pH measurements that meets the requirements for imaging pH-responses from cells as well as for a future hybrid design for detection of multiple analytes simultaneously. The sensor is based on the covalent attachment of 8-hydroxypyrene-1,3,6-trisulfonate (HPTS) to a film forming cellulose acetate material through a sulfonamide linkage. The synthesis routes of the cellulose material and regio-specific covalent attachment of the dye are described in detail. The sensor was sterilized in two different ways and showed excellent biocompatibility with Chinese hamster ovary cells. Imprints from cells and mum thin cell extensions were visualized when altering pH of the surrounding solution. The sensor was tested together with time-dependent sigmoidal calibrations giving pH determinations with an exceptional precision and accuracy during measurement within pH 6-8. The precision of the optode, calculated as pooled S.D. (n=8) according to IUPAC recommendations between pH 6.641 and 7.742 was 0.0029. The accuracy was significantly better than the electrode used as reference during the measurements. The response time (0-95%) was 100s between pH 6.641 and 7.742 and the reverse response (95-0%) was 80s. The sensor shows great potential for extra-cellular pH determination over time during cell growth and pharmacological exposure.

Determination of ammonium turnover and flow patterns close to roots using imaging optodes.

The physical effect of nitrogen upon plants has been studied thoroughly; however, direct studies of nitrogen turnover close to roots have been limited by analytical techniques with low spatial and temporal resolution. Thus, little is known about differences in turnover taking place along and between intact root structures over time as well as how root arrangement, root cell type, plant age, microbial activity, and the dark/light cycle influence uptake and supply of nutrients to root structures. In this study an imaging (planar) optode was used to quantify ammonium over time close to an intact root system of a large fruit bearing tomato plant (Lycopersicon esculentum). Images throughout the experiment made it possible to define the ammonium depletion zone and active turnover potential as well as determine turnover rate and flow patterns around the root system over time. The results indicated that ammonium uptake for tomato plants proceeds over the entire root structure but transverse thin peripheral roots are about twice as efficient as the main root and that the uptake process might influence nutrient availability. The flow patterns close to the root structure revealed that apical regions seem to have a central role in ammonium acquisition.