Development and Validation of a Bioanalytical UHPLC-MS/MS Method Applied to Murine Liver Tissue for the Determination of Indocyanine Green Loaded in H-Ferritin Nanoparticles

Indocyanine green (ICG) is one of the most commonly used fluorophores in near-infrared fluorescence-guided techniques. However, the molecule is prone to form aggregates in saline solution with a limited photostability and a moderate fluorescence yield. ICG was thus formulated using protein-based nanoparticles of H-ferritin (HFn) in order to generate a new nanostructure, HFn-ICG. In this study, an ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) system was employed to develop and validate the quantitative analysis of ICG in liver tissue samples from HFn-ICG-treated mice. To precipitate HFn, cold acetone in acidic solution at pH 5.0 was used. The processed liver samples were injected into the UHPLC-MS/MS system for analysis using the positive electrospray ionization mode. Chromatographic separation was achieved on a Waters Acquity UPLC® HSS T3 Column (1.8 μm, 2.1 × 100 mm) with 0.1% formic acid and acetonitrile as the mobile phase with gradient elution. The selected reaction monitoring transitions of m/z 753 →m/z 330 and m/z 827 →m/z 330 were applied for ICG and IR-820 (the internal standard, IS), respectively. The method was selective and linear over a concentration range of 50–1,500 ng/ml. The method was validated for sensitivity, accuracy, precision, extraction recovery, matrix effect, and stability in liver tissue homogenates. ICG extraction recoveries ranged between 85 and 108%. The intra- and inter-day precisions were less than 6.28%. The method was applied to a bio-distribution study to compare the amount of ICG levels from mice treated with HFn-ICG and free ICG. The analyses of the homogenate samples from the two types of treatment showed that the concentration levels of ICG is approximately six-fold higher than those of free ICG (1,411 ± 7.62 ng/ml vs. 235 ± 26.0 ng/ml) at 2 h post injection.

Photophysical, photostability, and ROS generation properties of new trifluoromethylated quinoline-phenol Schiff bases

A new series of ten examples of Schiff bases, namely (E)-2-(((2-alkyl(aryl/heteroaryl)-4-(trifluoromethyl)quinolin-6-yl)imino)methyl)phenols 3, was easily synthesized with yields of up to 91% from the reactions involving a series of 2-(R-substituted) 6-amino-4-(trifluoromethyl)quinolines 1 and 4(5)-R1-substituted salicylaldehydes 2 – in which alkyl/aryl/heteroaryl for 2-R-substituents are Me, Ph, 4-MeC6H4, 4-FC6H4, 4-NO2C6H4, and 2-furyl, and R1-substituents are 5-NEt2, 5-OCH3, 4-Br, and 4-NO2. Complementarily, the Schiff bases showed low to good quantum fluorescence yield values in CHCl3 (Φf = 0.12–0.80), DMSO (Φf = 0.20–0.75) and MeOH (Φf = 0.13–0.85). Higher values of Stokes shifts (SS) were observed in more polar solvents (DMSO; 65–150 nm and MeOH; 65–130 nm) than in CHCl3 (59–85 nm). Compounds 3 presented good stability under white-LED irradiation conditions and moderate ROS generation properties were observed.

Salt Stress Induces Paramylon Accumulation and Fine-Tuning of the Macro-Organization of Thylakoid Membranes in Euglena gracilis Cells

The effects of salt stress condition on the growth, morphology, photosynthetic performance, and paramylon content were examined in the mixotrophic, unicellular, flagellate Euglena gracilis. We found that salt stress negatively influenced cell growth, accompanied by a decrease in chlorophyll (Chl) content. Circular dichroism (CD) spectroscopy revealed the changes in the macro-organization of pigment-protein complexes due to salt treatment, while the small-angle neutron scattering (SANS) investigations suggested a reduction in the thylakoid stacking, an effect confirmed by the transmission electron microscopy (TEM). At the same time, the analysis of the thylakoid membrane complexes using native-polyacrylamide gel electrophoresis (PAGE) revealed no significant change in the composition of supercomplexes of the photosynthetic apparatus. Salt stress did not substantially affect the photosynthetic activity, as reflected by the fact that Chl fluorescence yield, electron transport rate (ETR), and energy transfer between the photosystems did not change considerably in the salt-grown cells. We have observed notable increases in the carotenoid-to-Chl ratio and the accumulation of paramylon in the salt-treated cells. We propose that the accumulation of storage polysaccharides and changes in the pigment composition and thylakoid membrane organization help the adaptation of E. gracilis cells to salt stress and contribute to the maintenance of cellular processes under stress conditions.

Effect of Magnetopriming on Photosynthetic Performance of Plants

Magnetopriming has emerged as a promising seed-priming method, improving seed vigor, plant performance and productivity under both normal and stressed conditions. Various recent reports have demonstrated that improved photosynthesis can lead to higher biomass accumulation and overall crop yield. The major focus of the present review is magnetopriming-based, improved growth parameters, which ultimately favor increased photosynthetic performance. The plants originating from magnetoprimed seeds showed increased plant height, leaf area, fresh weight, thick midrib and minor veins. Similarly, chlorophyll and carotenoid contents, efficiency of PSII, quantum yield of electron transport, stomatal conductance, and activities of carbonic anhydrase (CA), Rubisco and PEP-carboxylase enzymes are enhanced with magnetopriming of the seeds. In addition, a higher fluorescence yield at the J-I-P phase in polyphasic chlorophyll a fluorescence (OJIP) transient curves was observed in plants originating from magnetoprimed seeds. Here, we have presented an overview of available studies supporting the magnetopriming-based improvement of various parameters determining the photosynthetic performance of crop plants, which consequently increases crop yield. Additionally, we suggest the need for more in-depth molecular analysis in the future to shed light upon hidden regulatory mechanisms involved in magnetopriming-based, improved photosynthetic performance.

Photophysical, photostability, and ROS generation properties of new trifluoromethylated quinoline-phenol Schiff bases

A new series of ten examples of (E)-2-(((2-alkyl(aryl/heteroaryl)-4-(trifluoromethyl)quinolin-6-yl)imino)methyl)phenols (Schiff bases), was easy synthesized at yields up to 91% from the reactions involving a series of 2-(R-substituted) 6-amino-4-(trifluoromethyl)quinolines and 4(5)-R1-substituted salicylaldehydes – in which alkyl/aryl/heteroaryl for 2-(R)-substituents are Me, Ph, 4-MeC6H4, 4-FC6H4, 4-NO2C6H4, and 2-furyl, and (R1)-substituents are 5-NEt2, 5-OCH3, 4-Br, and 4-NO2. Complementarily, the Schiff bases showed low to good quantum fluorescence yield values both in CHCl3 (Φf = 0.12-0.80), DMSO (Φf = 0.20-0.75) and MeOH (Φf = 0.13-0.85). Higher values of Stokes shifts (SS) ​​were observed in more polar solvents (DMSO; 65-150 nm and MeOH; 65-130 nm) than in CHCl3 (59-85 nm). Also, compounds 3 confirmed to present good stability under white-LED irradiation conditions and moderate ROS generation properties were observed.

Fluorescent PIV using Atomized Liquid Particles

It is shown that aerosolized fluorescent particles generated using a Venturi-type atomizer, from a solution of fluorescent Kiton Red 620 dye in a water/glycol fluid, provide effective flow seeding for fluorescent PIV. The atomized liquid particles were found to be of acceptable size for PIV purposes, with 92% of detected particles by number concentration measuring < 1 μm in diameter. A PIV application was conducted in a wind tunnel (freestream velocity U∞ = 27 m/s), using the particles for measurement of the boundary layer flow approaching a forward-facing step (approach boundary layer momentum thickness Reynolds number of Reθ = 5930), to identify potential benefits in near-wall regions normally affected by unwanted laser reflections from tunnel surfaces. Particles were generated from solutions with dye molar concentrations of 2.5 × 10−3 and 1.0 × 10−2 mol/L, and PIV images were obtained for both elastic Mie scattering and filtered, Stokes-shifted fluorescent light. Raw images indicate that the fluorescence yield of the 1.0 × 10−2 mol/L solution provides PIV images with high contrast, even in the near-surface regions where Mie scattering images are highly affected by surface reflections. Boundary layer profiles are processed in the adverse pressure gradient region leading up to the forward-facing step, where the fluorescent PIV performed comparably to the most optimized Mie scattering PIV; both obtained data as near to the wall as 30 μm, or 2 viscous wall units in our flow of interest. These results indicate that the new seeding method holds excellent promise for near-surface measurement applications with more complicated three-dimensional geometries, where it is impossible to arrange PIV cameras to reject surface-scattered light.

Photobiological Effects on Ice Algae of a Rapid Whole-Fjord Loss of Snow Cover during Spring Growth in Kangerlussuaq, a West Greenland Fjord

Snow cover on sea ice is the most important factor controlling light availability for sea ice algae, but it is predicted by climate models to become more variable and stochastic. Here, we document effects of a sudden, complete loss of the entire snow cover on first-year sea ice at Kangerlussuaq Fjord, West Greenland, due to a natural Föhn wind event that caused a ca. 17 °C air temperature increase over 36 h. We applied Imaging-PAM fluorometry to examine effects of snow cover on algal distribution and photobiology and observed a rapid decrease in algal biomass associated with loss of the skeletal ice crystal layer on the underside of the ice that had supported most of the visible algae. Furthermore, the remaining algae were photobiologically stressed, as seen in a significant decrease in the dark-acclimated fluorescence yield (ΦPSII_max) from 0.55 before snow loss to 0.41 after. However, recovery in the dark suggested that non-photosynthetic quenching was successfully dissipating excess energy in the community and that there was little photodamage. An observed decrease in the photosynthetic efficiency α from 0.22 to 0.16 µmol é m−2 s−1 is therefore likely to be due to photoacclimation and the change in community composition. Centric diatoms and flagellates were the main taxa lost in the snow loss event, whereas the sea ice specialist Nitzschia frigida increased in numbers. These observations are similar to those seen in artificial snow-clearing experiments and consistent with snow clearing being a useful approach for investigating the complex interactions between snow cover, irradiance fluctuations, and ice algal performance.

Thionation of a Fluorescent Thieno[3,4-d]pyrimidine Derivative for the Development of a Heavy-Atom-Free Photosensitizer

All-organic, heavy-atom-free photosensitizers based on thionation of nucleobases are receiving increased attention because they are easy to make, noncytotoxic, work both in the presence and absence of molecular oxygen and can be readily incorporated into DNA and RNA. In this contribution, the DNA and RNA fluorescent probe, thieno[3,4-d]pyrimidin-4(1H)-one, has been thionated to develop thieno[3,4-d]pyrimidin-4(1H)-thione, which is nonfluorescent and absorbs near-visible radiation with about 60% higher efficiency. Steady-state absorption and emission spectra are combined with transient absorption spectroscopy and CASPT2 calculations to delineate the electronic relaxation mechanisms of both pyrimidine derivatives in aqueous and acetonitrile solutions and to explain the origin of the remarkable fluorescence quenching in the thionated compound. It is demonstrated that thieno[3,4-d]pyrimidin-4(1H)-thione efficiently populates the long-lived and reactive triplet state in hundreds of femtoseconds independent of solvent. Conversely, fluorescence emission in thieno[3,4-d]pyrimidin-4(1H)-one is highly sensitive to solvent, with an order of magnitude decrease in fluorescence yield in going from aqueous to acetonitrile solution. Collectively, the experimental and computational results demonstrate that thieno[3,4-d]pyrimidine-4(1H)-thione stands out as the most promising thiopyrimidine photosensitizer developed to this date, which can be readily incorporated as a photodynamic agent into sequence-specific DNA and RNA sequences for the treatment of skin cancer cells.

Thionation of a Fluorescent Thieno[3,4-d]pyrimidine Derivative for the Development of a Heavy-Atom-Free Photosensitizer

All-organic, heavy-atom-free photosensitizers based on thionation of nucleobases are receiving increased attention because they are easy to make, noncytotoxic, work both in the presence and absence of molecular oxygen and can be readily incorporated into DNA and RNA. In this contribution, the DNA and RNA fluorescent probe, thieno[3,4-d]pyrimidin-4(1H)-one, has been thionated to develop thieno[3,4-d]pyrimidin-4(1H)-thione, which is nonfluorescent and absorbs near-visible radiation with about 60% higher efficiency. Steady-state absorption and emission spectra are combined with transient absorption spectroscopy and CASPT2 calculations to delineate the electronic relaxation mechanisms of both pyrimidine derivatives in aqueous and acetonitrile solutions and to explain the origin of the remarkable fluorescence quenching in the thionated compound. It is demonstrated that thieno[3,4-d]pyrimidin-4(1H)-thione efficiently populates the long-lived and reactive triplet state in hundreds of femtoseconds independent of solvent. Conversely, fluorescence emission in thieno[3,4-d]pyrimidin-4(1H)-one is highly sensitive to solvent, with an order of magnitude decrease in fluorescence yield in going from aqueous to acetonitrile solution. Collectively, the experimental and computational results demonstrate that thieno[3,4-d]pyrimidine-4(1H)-thione stands out as the most promising thiopyrimidine photosensitizer developed to this date, which can be readily incorporated as a photodynamic agent into sequence-specific DNA and RNA sequences for the treatment of skin cancer cells.