Variation of Spectral Characteristics of Coelenteramide-Containing Fluorescent Protein from Obelia Longissima Exposed to Dimethyl Sulfoxide

Effect of dimethyl sulfoxide (DMSO), a widespread biomedical agent, on spectral-luminescent characteristics of coelenteramide-containing fluorescent protein – discharged obelin – is investigated. Contributions of violet and blue-green spectral components to fluorescence of discharged obelin are elucidated and characterized at different photoexcitation energies. Dependences of these contributions on the DMSO concentration are presented. Spectral changes are related to the destructive effect of DMSO on fluorescent protein and decreasing efficiency of proton transfer to electronically excited states of fluorophore

On the mechanism of biological activation by tritium

The mechanism of biological activation by beta-emitting radionuclide tritium was studied. Luminous marine bacteria were used as a bioassay to monitor the biological effect of tritium with luminescence intensity as the physiological parameter tested. Two different types of tritium sources were used: HTO molecules distributed regularly in the surrounding aqueous medium, and a solid source with tritium atoms fixed on its surface (tritium-labeled films, 0.11, 0.28, 0.91, and 2.36 MBq/cm2). When using the tritium-labeled films, tritium penetration into the cells was prevented. The both types of tritium sources revealed similar changes in the bacterial luminescent kinetics: a delay period followed by bioluminescence activation. No monotonic dependences of bioluminescence activation efficiency on specific radioactivities of the films were found. A 15-day exposure to tritiated water (100 MBq/L) did not reveal mutations in bacterial DNA. The results obtained give preference to a “non-genomic” mechanism of bioluminescence activation by tritium. An activation of the intracellular bioluminescence process develops without penetration of tritium atoms into the cells and can be caused by intensification of trans-membrane cellular processes stimulated by ionization and radiolysis of aqueous media

Endohedral Gd-Containing Fullerenol: Toxicity, Antioxidant Activity, and Regulation of Reactive Oxygen Species in Cellular and Enzymatic Systems

The Gd-containing metallofullerene derivatives are perspective magnetic resonance imaging contrast agents. We studied the bioeffects of a water-soluble fullerene derivative, gadolinium-endohedral fullerenol, with 40–42 oxygen groups (Gd@Fln). Bioluminescent cellular and enzymatic assays were applied to monitor toxicity and antioxidant activity of Gd@Fln in model solutions; bioluminescence was applied as a signaling physiological parameter. The Gd@Fln inhibited bioluminescence at high concentrations (>2·10−1 gL−1), revealing lower toxicity as compared to the previously studied fullerenols. Efficient activation of bioluminescence (up to almost 100%) and consumption of reactive oxygen species (ROS) in bacterial suspension were observed under low-concentration exposure to Gd@Fln (10−3–2·10−1 gL−1). Antioxidant capability of Gd@Fln was studied under conditions of model oxidative stress (i.e., solutions of model organic and inorganic oxidizers); antioxidant coefficients of Gd@Fln were determined at different concentrations and times of exposure. Contents of ROS were evaluated and correlations with toxicity/antioxidant coefficients were determined. The bioeffects of Gd@Fln were explained by hydrophobic interactions, electron affinity, and disturbing of ROS balance in the bioluminescence systems. The results contribute to understanding the molecular mechanism of “hormetic” cellular responses. Advantages of the bioluminescence assays to compare bioeffects of fullerenols based on their structural characteristics were demonstrated

Exposure of luminous marine bacteria to low-dose gamma-radiation

The study addresses biological effects of low-dose gamma-radiation. Radioactive 137Cs-containing particles were used as model sources of gamma-radiation. Luminous marine bacterium Photobacterium phosphoreum was used as a bioassay with the bioluminescent intensity as the physiological parameter tested. To investigate the sensitivity of the bacteria to the low-dose gamma-radiation exposure (≤ 250 mGy), the irradiation conditions were varied as follows: bioluminescence intensity was measured at 5, 10, and 20°С for 175, 100, and 47 h, respectively, at different dose rates (up to 4100 μGy/h). There was no noticeable effect of gamma-radiation at 5 and 10°С, while the 20°С exposure revealed authentic bioluminescence inhibition. The 20°С results of gamma-radiation exposure were compared to those for low-dose alpha- and beta-radiation exposures studied previously under comparable experimental conditions. In contrast to ionizing radiation of alpha and beta types, gamma-emission did not initiate bacterial bioluminescence activation (adaptive response). As with alpha- and beta-radiation, gamma-emission did not demonstrate monotonic dose-effect dependencies; the bioluminescence inhibition efficiency was found to be related to the exposure time, while no dose rate dependence was found. The sequence analysis of 16S ribosomal RNA gene did not reveal a mutagenic effect of low-dose gamma radiation. The exposure time that caused 50% bioluminescence inhibition was suggested as a test parameter for radiotoxicity evaluation under conditions of chronic low-dose gamma irradiation

Variation of Spectral Characteristics of Coelenteramide-Containing Fluorescent Protein from Obelia Longissima Exposed to Dimethyl Sulfoxide

Effect of dimethyl sulfoxide (DMSO), a widespread biomedical agent, on spectral-luminescent characteristics of coelenteramide-containing fluorescent protein – discharged obelin – is investigated. Contributions of violet and blue-green spectral components to fluorescence of discharged obelin are elucidated and characterized at different photoexcitation energies. Dependences of these contributions on the DMSO concentration are presented. Spectral changes are related to the destructive effect of DMSO on fluorescent protein and decreasing efficiency of proton transfer to electronically excited states of fluorophore

BIOLOGICAL ACTIVITY OF CARBONIC NANO-STRUCTURES-COMPARISON VIA ENZYMATIC BIOASSAY

Purpose: The aim of the work is to compare the biological activity of carbonic nano-structures of natural and artificial origination, namely, humic substances (HS) and fullerenols. Materials and methods: The representative of the fullerenol group, С 60 О y (OH) x where у + x = 20–22, was chosen. Enzyme-based luminescent bioassay was applied to evaluate toxicity and antioxidant properties of HS and fullerenol (F); chemiluminescent luminol method was used to study a content of reactive oxygen species (ROS) in the solutions. Toxicity of the bioactive compounds was evaluated using effective concentrations ЕС 50 ; detoxification coefficients D OxT were applied to study and compare antioxidant activity of the compounds. Antioxidant activity and ranges of active concentrations of the bioactive compounds were determined in model solutions of organic and inorganic oxidizers—1,4-benzoquinone and potassium ferricianide. Results and discussion: Values of ЕС 50 revealed higher toxicity of HS than F (0.005 and 0.108 g L −1 , respectively); detoxifying concentrations of F were found to be lower. Antioxidant ability of HS was demonstrated to be time-dependent; the 50-min preliminary incubation in oxidizer solutions was suggested as optimal for the detoxification procedure. On the contrary, F’ antioxidant effect demonstrated independency on time. Antioxidant effect of HS did not depend on amphiphilic characteristics of the media (values of D OxT were 1.3 in the solutions of organic and inorganic oxidizers), while this of F was found to depend: it was maximal (D OxT = 2.0) in solutions of organic oxidizer, 1,4-benzoquinone. Conclusions: Both HS and F demonstrated toxicity and low-concentration antioxidant ability; however, quantitative characteristics of their effects were different. The differences were explained with HS polyfunctionality, higher ability to decrease ROS content, non-rigidity, and diffusion restrictions in their solutions. Antioxidant effect of the bioactive compounds was presumably attributed to catalytic redox activity of their π-fragments. The paper demonstrates a high potential of luminescent enzymatic bioassay to study biological activity of nano-structures of natural and artificial origination

Ultraviolet fluorescence of coelenteramide and coelenteramide-containing fluorescent proteins. Experimental and theoretical study

Текст статьи не публикуется в открытом доступе в соответствии с политикой журнала.Coelenteramide-containing fluorescent proteins are products of bioluminescent reactions of marine coelenter- ates. They are called ‘discharged photoproteins’. Their light-induced fluorescence spectra are variable, depending considerably on external conditions. Current work studies a dependence of light-induced fluorescence spectra of discharged photoproteins obelin, aequorin, and clytin on excitation energy. It was demonstrated that photoexci- tation to the upper electron-excited states (260–300 nm) of the discharged photoproteins initiates a fluorescence peak in the near UV region, in addition to the blue-green emission. To characterize the UV fluorescence, the light- induced fluorescence spectra of coelenteramide (CLM), fluorophore of the discharged photoproteins, were studied in methanol solution. Similar to photoproteins, the CLM spectra depended on photoexcitation energy; the additional peak (330 nm) in the near UV region was observed in CLM fluorescence at higher excitation energy (260–300 nm). Quantum chemical calculations by time depending method with B3LYP/cc-pVDZ showed that the conjugated pyrazine-phenolic fragment and benzene moiety of CLM molecule are responsible for the additional UV fluorescence peak. Quantum yields of CLM fluorescence in methanol were 0.028 ± 0.005 at 270–340 nm pho- toexcitation. A conclusion was made that the UV emission of CLM might contribute to the UV fluorescence of the discharged photoproteins. The study develops knowledge on internal energy transfer in biological structures – complexes of proteins with low-weight aromatic molecules

Ultraviolet fluorescence of coelenteramide and coelenteramide-containing fluorescent proteins. Experimental and theoretical study

Coelenteramide-containing fluorescent proteins are products of bioluminescent reactions of marine coelenterates. They are called ‘discharged photoproteins’. Their light-induced fluorescence spectra are variable, depending considerably on external conditions. Current work studies a dependence of light-induced fluorescence spectra of discharged photoproteins obelin, aequorin, and clytin on excitation energy. It was demonstrated that photoexcitation to the upper electron-excited states (260-300 nm) of the discharged photoproteins initiates a fluorescence peak in the near UV region, in addition to the blue-green emission. To characterize the UV fluorescence, the light-induced fluorescence spectra of coelenteramide (CLM), fluorophore of the discharged photoproteins, were studied in methanol solution. Similar to photoproteins, the CLM spectra depended on photoexcitation energy; the additional peak (330 nm) in the near UV region was observed in CLM fluorescence at higher excitation energy (260-300 nm). Quantum chemical calculations by time depending method with B3LYP / cc-pVDZ showed that the conjugated pyrazine-phenolic fragment and benzene moiety of CLM molecule are responsible for the additional UV fluorescence peak. Quantum yields of CLM fluorescence in methanol were 0.028 ± 0.005 at 270-340 nm photoexcitation. A conclusion was made that the UV emission of CLM might contribute to the UV fluorescence of the discharged photoproteins. The study develops knowledge on internal energy transfer in biological structures – complexes of proteins with low-weight aromatic molecules

Is bacterial luminescence response to low-dose radiation associated with mutagenicity?

Luminous marine bacteria are widely used in bioassays with luminescence intensity being a physiological parameter tested. The purpose of the study was to determine whether bacterial genetic alteration is responsible for bioluminescence kinetics change under low-dose radiation exposure. The alphaemitting radionuclide 241Am and beta-emitting radionuclide 3 H were used as the sources of low-dose ionizing radiation. Changes of bioluminescence kinetics of Photobacterium phosphoreum in solutions of 241Am(NO3)3, 7 kBq/L, and tritiated water, 100 MBq/L, were studied; bioluminescence kinetics stages (absence of effect, activation, and inhibition) were determined. Bacterial suspension was sampled at different stages of the bioluminescent kinetics; the doses accumulated by the samples were close or a little higher than a tentative limit of a low-dose interval: 0.10 and 0.85 Gy for 241Am, or 0.11 and 0.18 Gy for 3H. Sequence analysis of the 16S ribosomal RNA gene did not reveal a mutagenic effect of low-dose alpha and beta radiation in the bacterial samples. Previous results on bacterial DNA exposed to low-dose gamma radiation (0.25 Gy) were analyzed and compared to those for alpha and beta irradiation. It is concluded that bioluminescence activation and/or inhibition under the applied conditions of low-dose alpha, beta and gamma radioactive exposure is not associated with DNA mutations in the gene sequences teste