This study investigates the ultra-weak photon emission (UPE) arising from spontaneous photonic reactions in different water types — tap water, bottled mineral water, reverse-osmosis water, and water treated by the Biodynamizer dynamization system (SA Dynamized Technologies). The objective was to evaluate potential differences in their photon emission characteristics and their influence on the germination and early development of seeds. Measurements were conducted using a Berthold Lumat LB 9508 luminometer, equipped with borosilicate glass test tubes and an optical bandpass filter (435–500 nm), covering the spectral range 380–630 nm and the subrange 435–500 nm. All experiments were performed under controlled laboratory conditions at Enerlab (Nice, France) on November 4–6, 2025. Photon emission intensities are expressed in Relative Light Units (RLU). The results show no emissions (0 RLU) for mains water, bottled mineral water, and reverse osmosis water, while biodynamically treated mains water shows 519 RLU (380–630 nm) and 272 RLU (435–500 nm) immediately after dynamization treatment. After 24 hours, the values remain significant (184 and 168 RLU, respectively) for biodynamically treated mains water, indicating a partial decrease but persistence of the biophotonic emission phenomenon. Measurements of ultra-weak photons in the blue-green spectral range (≈435–500 nm) highlight a better persistence of the biophotonic emission phenomenon (-38% instead of -65% in the 380-630 nm spectral range) over 24 hours after biodynamized treatment of water. This spectral range is known to correspond to electronic transitions of flavins and cytochromes, involved in mitochondrial respiration and, in plants and photosynthesis. However, interpreting these variations as an increase in photonic coherence requires further investigation. The particularly marked emission in the 435–500 nm band highlights a possible link between biodynamization, photobiological coherence and cellular energy processes. In the Integrating study Ultra ‑Weak Photon Emission Analysis in Mitochondrial Research (Van Wijk & Van Wijk 2020), UPE is mentioned in the 300–900 nm range, without exclusive or claimed details on 435,500 ‑nm. PMC +2 PubMed +2. In the study Spectral Distribution of Ultra ‑Weak Photon Emission as a Response to Wounding in Plants (Prasad et al. 2020), the authors indicate that photon emission covers from ~350 nm to ~1300 nm. MDPI. “UV-visible absorption spectrum of FAD and its reduced forms embedded in a cryptochrome protein” — Physical Chemistry Chemical Physics (RSC Publishing) DOI:10.1039/D0CP01714K RSC Publishing. Abstract: The authors show that for the coenzyme flavin adenine dinucleotide (FAD), in its oxidized form, a π₂→π₃ transition is observed around 450 nm (blue-green). The emission fluorescence is around ~510-520 nm in this context. This result therefore confirms that FAD has maximum absorption in the ~450 nm range, which falls well within the 435-500 nm band. Note: “For the oxidized form of FAD… the first band in the blue visible light region… centered around 450 nm.” RSC Publishing. “Ultrafast dynamics of flavins in five redox states” — PMC, 2009/2010 (approximately) PMC. Summary: Study on the absorption and emission spectra of flavins (FAD, FMN) in several redox states. It reports that the absorption for the oxidized form of FAD is at ~450 nm, emission at ~530 nm. Note: “Oxidized FAD in solution exhibits two broad absorption bands… with the peaks at 450 nm for S₀→S₁ and at 375 nm for S₀→S₂.” PMC. This supports the fact that the blue-green region (≈450 nm) is a relevant signal for flavins involved in respiration. “Why Flavins Are Not Competitors of Chlorophyll in the Evolution of Biological Converters of Solar Energy” — International Journal of Molecular Sciences, 2013 (MdPI). Abstract: This article mentions that flavins absorb “photons from the blue area… the long-wave absorption maximum of flavin is 450 nm.” Quote: “…Neutral solutions of riboflavin, FMN, and FAD have a long-wave absorption maximum of 450 nm…” MDPI. This further confirms that 450 nm is a peak absorption for flavins—a key element in mitochondrial energy metabolism. “Absorption and Luminescence Spectroscopy of Mass-Selected Flavin Adenine Dinucleotide Mono-anions” — The Journal of Chemical Physics, 2018 DOI:10.1063/1.5024028. Abstract: Researchers measured the optical transitions of FAD mono-anions from 210 to 550 nm. They show that one of the transitions (S₀→S₂) is strongly linked to ~450 nm under gas-phase conditions. Quote: “The measurements cover the first four optical transitions of FAD… excitation energies from 2.3 to 6.0 eV (210–550 nm)…This reinforces the idea that the ~435–500 nm range is relevant for flavins. “Circular spectropolarimetric sensing of chiral photosystems in decaying leaves“ — arXiv, 2017 arXiv.

Fatty acid photodecarboxylase catalyzes the light-driven decarboxylation of fatty acids into hydrocarbons via electron transfer (ET) from the substrate to the flavin adenine dinucleotide cofactor, proceeding through either proton-coupled ET or hydrogen-atom transfer. Through quantum mechanics/molecular mechanics calculations, we show that only the deprotonated fatty acid supports the charge-transfer states required to initiate catalysis. Molecular dynamics simulations combined with graph-theory-based analysis reveal that the crystallographic water site, Wat1, is consistently occupied by a stable, yet dynamically exchanging, population of water molecules. We also identify transient, solvent-accessible water channels connecting the active site to the bulk solvent, potentially facilitating proton transfer and water exchange. These findings support the notion that beyond preserving the structural integrity of the active site, this water population may also enable the flexible modulation of electron and proton transfer through an adaptive hydrogen-bonding network.

Hydroquinone (HQ) is a redox-active organic compound widely used in cosmetics, pharmaceuticals, photography, and dye manufacturing. Despite its industrial utility, hydroquinone is considered toxic and potentially carcinogenic, necessitating the development of sensitive, rapid, and selective methods for its detection in environmental and biological samples. Traditional analytical techniques such as high-performance liquid chromatography, spectrophotometry, and fluorimetry, while effective, often suffer from drawbacks, including expensive instrumentation, time-consuming procedures, and complex sample preparation. In contrast, electrochemical biosensing offers a promising alternative due to its simplicity, portability, high sensitivity, and suitability for real-time monitoring.In the present work, we report the design and fabrication of a highly sensitive electrochemical biosensor based on a novel nanocomposite comprising Flavin Adenine Dinucleotide (FAD) functionalized fluorapatite (FA) and single-walled carbon nanotubes (SWCNTs). The FAD/FA/SWCNT composite was synthesized via a mechanochemical method, a green and scalable approach that eliminates the need for complex chemical treatments or high-temperature processes. Dimethyl sulfoxide (DMSO) was used as a dispersing solvent to ensure the homogeneous distribution of FAD molecules and nanotubes within the FA matrix. The synthesized composite was drop-cast onto a glassy carbon electrode (GCE) to construct the biosensor platform.The structural and physicochemical properties of the FAD/FA/SWCNT nanocomposite were characterized using a combination of high-resolution scanning electron microscopy (HR-SEM), X-ray diffraction (XRD), and UV–Visible spectroscopy. SEM analysis revealed a uniform, sheet-like morphology with well-dispersed nanotubes, providing a high surface area conducive to electrochemical reactions. XRD confirmed the crystalline structure of fluorapatite and the successful incorporation of SWCNTs, while UV–vis spectroscopy identified the characteristic absorption peak of FAD at 450 nm, indicating successful functionalization.The electrochemical properties of the modified electrode were evaluated using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and amperometry in 0.1 M phosphate buffer solution (PBS) at pH 7.4. The FAD/FA/SWCNT-modified GCE displayed a distinct redox peak at –0.45 V, attributed to the electroactive nature of FAD. The composite exhibited a significantly lower charge transfer resistance (Rct) compared to the bare and partially modified electrodes, highlighting the synergistic enhancement of electron transfer due to the integrated functionalities of FAD, FA, and SWCNTs.The electrocatalytic activity of the biosensor was systematically investigated for hydroquinone detection. The sensor showed a linear detection range from 0.005 µM to 258.2 µM, with a limit of detection (LOD) of 2.70 nM. This remarkable sensitivity is primarily attributed to the combined effects of: FAD serves as a redox-active mediator facilitating electron transfer, FA contributes to high surface area and biocompatibility, SWCNTs, which provide excellent electrical conductivity and enhance charge mobility. Scan rate studies indicated that the electrochemical response involves a mixed control mechanism, with contributions from both diffusion and surface-confined processes. The calculated charge transfer coefficient (α = 0.53) and linear dependence of peak current on both the square root and logarithm of scan rate further confirmed favorable reaction kinetics. The biosensor exhibited excellent reproducibility and operational stability, with consistent responses over 20 continuous cycles and minimal signal degradation.To validate the sensor’s real-world applicability, HQ detection was performed in spiked water samples (tap and mineral water) using the standard addition method. The biosensor demonstrated excellent recovery rates ranging from 100.4% to 103.0%, affirming its accuracy and reliability in complex sample matrices. Additionally, the sensor displayed a fast amperometric response time of approximately 4 seconds under optimized conditions at an applied potential of 0.09 V.When compared to existing HQ sensors based on advanced nanomaterials such as MoS₂/RGO, COF/CPE, and MWCNT-COOH/CTF-1, the FAD/FA/SWCNT-modified GCE outperformed in both detection range and sensitivity, demonstrating the strong potential of this composite for next-generation electrochemical biosensors.Importantly, this work marks the first report on the use of fluorapatite functionalized with a redox cofactor (FAD) for biosensing applications. The mechanochemical synthesis approach not only offers a sustainable and straightforward fabrication method but also enhances the composite’s performance due to the uniform dispersion and intimate interaction of all components. Conclusion: The FAD/FA/SWCNT-based electrochemical biosensor developed in this study offers a novel, efficient, and highly sensitive platform for hydroquinone detection. Its excellent analytical performance, combined with ease of fabrication and low detection limit, makes it a promising candidate for environmental monitoring and point-of-care diagnostics. This research also opens new avenues for incorporating bio-functionalized bioceramics into sensor technology, bridging the gap between nanomaterials, green chemistry, and practical biosensing applications. Figure 1

Mitochondrial calcium (Ca2+) is a key regulator of cardiac energetics by stimulating the tricarboxylic acid cycle during elevated workload. Atrial fibrillation (AF) is associated with a reduction in cytosolic Ca2+ transient amplitude, but its effect on mitochondrial Ca2+ handling and cellular redox state has not been explored. Cardiac myocytes isolated from patient-derived right atrial biopsies were subjected to workload transitions using patch-clamp stimulation and β-adrenergic stimulation (isoproterenol). In conjunction, nicotinamide adenine dinucleotide (phosphate)/flavin adenine dinucleotide (NAD[P]H/FAD) autofluorescence, cytosolic and mitochondrial [Ca2+] were monitored using epifluorescence microscopy. Sarcoplasmic reticulum and mitochondria were imaged using electron microscopy and tomography and stimulated emission depletion microscopy. The effects of the mitochondrial Ca2+ uptake enhancer ezetimibe on proarrhythmic activity in atrial myocytes and on AF burden in patients were investigated. Mitochondrial Ca2+ accumulation during increased workload was blunted in AF, and was associated with impaired regeneration of nicotinamide adenine dinucleotide and flavin adenine dinucleotide. Nanoscale imaging revealed spatial disorganization of sarcoplasmic reticulum and mitochondria, associated with microtubule destabilization. This was confirmed in human induced pluripotent stem cell-derived cardiac myocytes, where treatment with the microtubule destabilizer nocodazole displaced mitochondria and increased proarrhythmic Ca2+ sparks, which were rescued by MitoTEMPO. Ezetimibe also reduced the occurrence of arrhythmogenic Ca2+ release events both in AF myocytes and nocodazole-treated human induced pluripotent stem cell-derived cardiac myocytes. Retrospective patient analysis also revealed a reduced AF burden in patients on ezetimibe treatment. Mitochondrial Ca2+ uptake and accumulation are impaired in atrial myocytes from patients with AF. The disturbed spatial association between sarcoplasmic reticulum and mitochondria driven by destabilized microtubules may underlie impaired Ca2+ transfer in AF. Enhancing mitochondrial Ca2+ uptake potentially protects against arrhythmogenic events.

Is it possible to assess label-free live cell metabolic imaging during early oocyte and embryo development? Label-free metabolic imaging can be systematically used during early development, showing no differences between controls and illuminated oocytes and embryos in terms of early development, blastocyst formation, and embryo outgrowth. Non-invasive methods that are reliable to assess oocyte and embryo quality are a significant aim for ARTs. Changes in metabolic activity could lead to cell death or altered early development and low implantation potential. This could potentially be predicted by incorporating non-invasive measurements of metabolism. Metabolic imaging has been investigated through complex methodologies; however, scientific evidence for its utility during early oocyte and embryo development requires further investigation to assess potential translation in clinical settings. Measurements of metabolic activity could be a useful tool, as the autofluorescence of molecules such as nicotinamide adenine dinucleotide phosphate hydrogen (NAD(P)H) and flavin adenine dinucleotide (FAD) are a straightforward representation of mitochondrial function. Female mice (n = 15) and super-ovulated female mice (n = 30) were used to produce oocytes and embryos, respectively. Oocytes and in-vivo produced embryos were divided into the control group, sham control group, and illuminated group. Illuminated samples were assessed for both NAD(P)H and FAD levels in oocytes and NAD(P)H levels during early embryo development every 3 h using arbitrary units of autofluorescence (AU). Produced blastocysts were assessed for total cell and inner-cell-mass (ICM) number (by immunostaining for Oct4) and embryo outgrowth assays. Furthermore, safety live birth studies were also conducted. F1 (C57BL6/CBA) mouse strain was used. NAD(P)H and FAD autofluorescence levels were measured during oocyte and embryo development using confocal microscopy (Olympus FV1200). A confocal Z-stacking function was used to record 15 focal planes, using a 20×/0.95 NA air objective of the entire oocytes and embryos and opening the confocal pinhole system completely. Images were then collected and analysed using FIJI software (version: 2.0.0-rc-69/1.52n; ImageJ). Developmental rates, blastocyst cell numbers, outgrowth rates (for 4 days post blastocyst formation), and live birth rates were assessed. Oocyte IVM and embryo culture experiments showed no significant differences in developmental rates between study groups (P > 0.05). Similarly, the total number of cells from blastocysts (control: 82.9 ± 5.6; sham: 76.5 ± 3.3; Illuminated: 77.1 ± 4.2; ± SEM) and ICM cells (control: 10.8 ± 1.3; sham: 9.4 ± 0.7; Illuminated: 11.9 ± 0.8; ± SEM) did not differ between groups (P > 0.05). Outgrowth assays of the study groups presented similar outgrowth areas during Days 5-8 (post) blastocyst development (P > 0.05). Illumination of oocytes demonstrated a significant increase in metabolic activity during IVM, measured by the optical redox ratio (ORR: FAD/NAD(P)H + FAD; P < 0.001). Illumination of embryos demonstrated significantly different NAD(P)H activity levels during embryo development, particularly between the two-cell stage (987.1 ± 36.2 AU), morula stage (1226.0 ± 31.5 AU) and blastocyst stage (649 ± 42.9 AU; ± SEM; P < 0.05). Additionally, embryos that did not form blastocysts also presented significantly decreased NAD(P)H activity levels at the two-cell stage (normal development: 987.1 ± 36.2; no blastocyst: 726.9 ± 121.7 AU; P < 0.05) to the morula stage (normal development: 1226.0 ± 31.5; no blastocyst: 886.0 ± 150.4 AU; P < 0.05) when compared with normally developing embryos. Our study indicated that metabolic imaging during early oocyte and embryo development presents no negative effects on developmental rates, blastocyst quality, and embryo outgrowths. Subsequently, live birth rates and offspring health showed no differences between controls and illuminated embryos at the blastocyst stage. Current results provide significant useful information about metabolic activity during live cell imaging as a potential method for timelapse metabolic imaging. N/A. The study was conducted using a mouse model and focused on early oocyte and embryo development, embryo outgrowths, live birth, and early offspring health. Thus, further studies of long-term offspring health are required to fully assess safety and to further validate potential wider applications. Validation in ageing models is also required to assess potential applications for embryo selection. Measurements of metabolic activity could be applied to determine oocyte and embryo metabolic activity using a variety of microscopy technology with low energy doses as described in this study. Further applications could link the use of metabolic imaging with timelapse technology and artificial intelligence applications to monitor culture conditions. This study was funded in part by a research/educational grant from Ferring Pharmaceuticals, awarded from the Fertility Society of Australia and New Zealand (FSANZ). Funding was also provided in part by the Education Program in Reproduction and Development (EPRD), Department of Obstetrics and Gynaecology, Monash University. F.H. and M.H.-T. have applied for a patent in the topic of metabolic imaging. R.B.G. declares speakers' fees from Gedeon Richter and Ferring. The other authors have nothing to declare.

A new family of StnC-like pseudo-FMN-preferred reductase components in two-component flavoprotein monooxygenases.

Electro-mediated biological system coupled with arrayed tubular electrode module for enhancing pharmaceutical wastewater treatment: Pilot-scale performance and electrically-driven metabolic mechanisms.

Novel (R)-Hydroxynitrile lyase enzyme of Pyrus communis: Purification and characterization of its physicochemical and kinetic properties.

Comparative analysis of autofluorescence spectra in a filet of three fish species during chilled storage for raw consumption.

Vitamin B2 (VB2) metabolism regulates numerous cellular processes, but its role in hepatocellular carcinoma (HCC) progression remains unclear. Here we show that HCC tumors are characterized by upregulation of a VB2 metabolism signature, and VB2 metabolism promotes HCC progression. Among VB2 metabolic enzymes, flavin adenine dinucleotide synthase (FADS) is the only one that is widely overexpressed in human HCC. Elevated FADS expression correlates with resistance to anti-PD-1 therapy and poor prognosis. In vivo, FADS facilitates HCC cell growth and suppresses Tcell-mediated antitumor immunity. Single-cell transcriptomic analysis reveals that FADS-induced changes occur both in the tumor cells and the intra-tumoral CD8+ T cells. Knocking down FADS induces HCC cell death and increases CD8⁺ T cell infiltration. Mechanistically, FADS confers ferroptosis resistance on HCC cells via enzymatic function to produce FAD and non-enzymatic function to stabilize PCBP2. Moreover, FADS impairs CD8+ T cell recruitment by disrupting the cGAS-STING pathway. Hesperidin, a clinically approved FADS inhibitor, shows antitumor efficacy in a mouse model. Our study thus highlights the importance of VB2 metabolism in HCC and provides the proof of principle for targeting FADS as a therapeutic strategy for HCC.

The cytochrome-b5 reductases (CYB5Rs) regulate cellular redox balance and contribute to the pathogenesis of inflammatory diseases. However, the roles of CYB5R5 in macrophages remain poorly understood and require further elucidation. In this study, we revealed that CYB5R5 orchestrates macrophage inflammation by inhibiting interleukin (IL)-1β production from M1 macrophages. Mechanistically, CYB5R5 enhances flavin adenine dinucleotide (FAD)-lysine demethylase1 (LSD1) signaling to regulate the histone demethylation of complement component 1, q subcomponent (C1q)-coding genes, thereby lowering NLRP3 inflammasome assembly. We also found that myeloid depletion of Cyb5r5 in mice exacerbates inflammatory responses in LPS-induced sepsis. This study reveals that CYB5R5 attenuates M1 macrophage polarization via metabolic and epigenetic reprogramming mechanism, thus providing potential therapeutic targets for macrophage-mediated inflammatory disorders.

Free radicals are generated in the body through endogenous and exogenous systems, with their overproduction linked to chronic diseases such as cancer. Interestingly, chemotherapeutic drugs utilize free radicals to induce apoptosis in cancer cells, highlighting their dual nature. This study explores the therapeutic potential of free-radical-generating compounds in solid tumor treatment. Using a patented universal method, superoxide (O2−)-producing enzymatic systems were isolated for the first time from serous fluids of patients with breast cancer, gastric cancer, and liver cirrhosis. These enzymes were qualitatively and quantitatively characterized and found to continuously produce monocomponent O2− under aerobic in vitro conditions. The enzyme complexes consist of flavin adenine dinucleotide (FAD), a reduced nicotinamide adenine dinucleotide phosphate (NADPH)-containing protein component (NPC), and Fe(III) ions. Their stable O2− production mechanism was elucidated, and characteristic optical absorption and fluorescence excitation spectra were recorded. The concentrations of monocomponent O2− were quantified in moles (mol/ml) for each serous fluid type. These findings suggest that liquid-phase O2− could be used to selectively destroy cancer cells by predetermining effective concentrations. Furthermore, since O2−-producing enzymes can oxidize adrenaline, they may help reduce elevated adrenaline levels in tumor cells. Future animal studies will assess their efficacy in eliminating metastatic cells, particularly in the postoperative period. This novel approach may offer a promising adjunct therapy in oncology.

Direct coupling of lactate oxidation with butyryl-CoA formation via a canonical electron transfer flavoprotein in Fusobacterium nucleatum

This study reports on the direct electron transfer (DET) ability of the enzyme spermidine dehydrogenase (SpDH) and its use in a DET-type enzymatic sensor for detecting spermine. SpDH was found to exhibit internal electron transfer from its cofactor, flavin adenine dinucleotide (FAD), to heme b. This was confirmed by observing the heme b-derived reduction peak at 560 nm in the presence of spermine, the substrate. SpDH was immobilized on a gold electrode via a dithiobis (succinimidyl hexanoate) self-assembled monolayer. The cyclic voltammetry analysis of the SpDH-immobilized gold electrode revealed an increased oxidation current in the presence of 0.1 mM spermine with an onset potential of -0.14 V vs. Ag/AgCl in the absence of an additional external electron acceptor. This result confirmed that SpDH is capable of DET. Chronoamperometric analyses were conducted using an SpDH-immobilized gold electrode with spermine as the substrate under a 0 V oxidation potential vs. Ag/AgCl using an artificial saliva matrix containing 10 µM ascorbic acid and 100 µM uric acid. The sensor exhibited good linear correlation between the current increase and spermine concentration from 0.2 to 2.0 µM, with a limit of detection of 0.084 µM, which encompasses the physiologically relevant spermine concentration found in the saliva. Primary structure alignments and 3D structure predictions revealed that all SpDH homologs possess two conserved histidine residues in the same location on the surface as the heme b ligand of SpDH. This indicates their potential for DET-ability with an electrode.

Enhancing biofilm resistance and ATP synthesis accelerates toluene degradation at low temperature via AHLs-mediated quorum sensing.

Bio-promoter-driven conversion of 4-chlorophenol into utilizable carbon source: Enhancing electron supply-consume and attenuating bio-toxicity.

The rise of multidrug-resistant (MDR) Pseudomonas aeruginosa poses a significant threat in clinical settings due to its intricate antimicrobial resistance mechanism, biofilm formation, quorum sensing, and efflux pump-mediated antibiotic tolerance capability. The progressive decline in the efficacy of conventional antibiotics necessitates the development of new treatment strategies. Disrupting the Quorum sensing, a pivotal regulator of virulence and biofilm-associated resistance presents a promising anti-virulence strategy. An integrated Subtractive genomics and in silico drug discovery approach was applied to the complete proteome of P. aeruginosa JJPA01, excluding paralogous, human homologous, and non-essential proteins to identify virulence-associated targets. 27 pathogen-specific pathway proteins were identified, with pqsH (WP_003090354.1), a key monooxygenase in the PQS quorum-sensing system. Potential inhibitors for pqsH were identified using High-Throughput Virtual Screening (HTVS) on natural compounds from the COCONUT, CMNPD, MNPD, Seaweed, and Specs databases. The docking study identified five compounds with the best binding affinities, ranging from -6.6 to -7.7kcal/mol. However, only CNP0000215 and CNP0007440 exhibited higher binding affinity to the pqsH protein than the cofactor Flavine Adenine Dinucleotide. With its established role in Antimicrobial Resistance and Virulence, pqsH has been selected as a therapeutic target and CNP0000215 as a promising PQS inhibitor to disrupt biofilm formation and combat antimicrobial resistance. These findings lay the groundwork for the strategic design of novel anti-therapeutics offering a promising strategy to inhibit persistent infections and resistance mechanisms in P. aeruginosa.

The enzyme D-aspartate oxidase (DDO) oxidizes acidic D-amino acids using the coenzyme flavin adenine dinucleotide to generate the corresponding α-keto acids and ammonia. DDO differs from D-amino-acid oxidase (DAAO), which acts on neutral and basic D-amino acids. Although the enzymatic properties of DDO have been characterized in several species, the structure of DDO had remained unclear. The structure of DDO derived from Cryptococcus humicola strain UJ1 (chDDO) was determined by X-ray crystallography at 1.70 Å resolution. While the three-dimensional structures of DAAOs are known to be homodimers, chDDO forms a homotetramer. This difference was found to be caused by the deletion of one loop and the insertion of two loops.

The initial photoproduct of the natural photoenzyme fatty acid photodecarboxylase involves the flavin anion radical flavin adenine dinucleotide (FAD•–). Using spectrally resolved ultrafast transient absorption spectroscopy, we demonstrate that FAD•– photoexcitation in the absence of substrate leads to the formation of the oxidized flavin FADox (the resting state in the catalytic cycle) within 100 femtoseconds. While this feature is similar to that occurring in flavoprotein oxidases, the ensuing photocycle is more complex. Upon excitation at the lowest-energy transition, the ejected electron is initially captured as a hydrated electron (e–H) before transferring to a secondary acceptor in 2.5 picoseconds and returning to the flavin in 37 picoseconds. This implies that e–H can be generated within a protein environment, an unprecedented finding. This assessment is supported by molecular dynamics simulations showing an expansion of the flavin-binding pocket without substrate, allowing water molecules to fill the void. Our results may pave the way to developing unconventional photocatalytic processes.

To explore the effects of Xinglou Chengqi Decoction (XCD) on severe traumatic brain injury (sTBI) and its relationship with gut microbiota. C57BL/6J mice were randomly allocated into sham, controlled cortical impact (CCI), and 3 doses of XCD (4.1, 8.2, and 16.4 g/kg) groups by using a random number table, n=7 per group. A CCI device was employed to establish the TBI model. XCD was administered intragastrically for 3 consecutive days. The effects of XCD on post-sTBI neurological deficits and histopathology were assessed. The contribution of gut microbiota to XCD-mediated improvement in sTBI was investigated using antibiotic-treated TBI mice. The gut microbiota-dependent mechanisms of XCD in sTBI were explored through 16S rDNA sequencing and serum metabolomics. The mechanisms underlying the absorbed ingredients of XCD in sTBI were examined using network pharmacology and metabolomics. Finally, mice were divided into sham, CCI, and flavin adenine dinucleotide (FAD)-treated groups, n=10 per group. FAD was administered to sTBI mice via daily tail vein injection (830 µg/kg) for 3 consecutive days to evaluate and verify its therapeutic effect. XCD significantly mitigated neurological impairments, neuronal damage, apoptosis, and blood-brain barrier disruption in CCI model mice (P<0.05 or P<0.01). The medium dose (8.2 g/kg) exhibited the greatest effect. The gut microbiota partly contributed to these protective effects. 16S rDNA sequencing indicated that XCD promoted beneficial gut microbiota. Metabolomic analysis demonstrated that XCD regulated serum metabolic profiles, particularly FAD. Network pharmacology combined with metabolomics analysis revealed that the gut microbiota-independent components of XCD also targeted FAD in TBI. FAD exerted neuroprotective effects, improved energy metabolism, and promoted angiogenesis following TBI (P<0.05 or P<0.01). XCD exerts neuroprotective effects on sTBI through both gut microbiota-dependent and -independent mechanisms, which highlight the therapeutic role of FAD.