Riboflavin Derivatives Research Articles

Monitoring Surface-Enhanced Infrared Absorption Spectroscopy on a Graphitic Electrode for in-Situ Surface-Sensitive Electrochemistry: Designing Electrode Interfaces for Carbon Capture

Combining spectroscopy with electrochemistry enables the monitoring of changes in bonding, molecular orientation, and chemical composition during electrochemical processes at the electrode/electrolyte interface. Surface-enhanced infrared absorption spectroscopy (ATR-SEIRAS) allows a direct in situ monitoring of electrochemical processes occurring at the electrode interface due plasmonic enhancement by surface plasmon polaritons (SPP).1 Electrochemical ATR-SEIRAS has been used extensively for investigating electrocatalysis2,3, characterizing adsorbed species4, and detecting reaction intermediates5. However, SEIRAS literature has focused on plasmonic metals while the small number of SEIRAS studies reported on graphene surfaces are incompatible with electrochemical experimentation or not considered. Nonetheless, carbon electrodes are ubiquitous in electrocatalysis, sensing, and energy storage6, thus probing their surface chemistries are of particular interest. Thus, there is still a need for a convenient substrate for probing graphitic interfaces using electrochemical SEIRAS.In our first study1, we expanded the scope of SEIRAS by introducing a robust hybrid graphene on gold (graphene-gold) substrate where we monitored electrografting processes occurring on the graphene/electrolyte interface. The graphene-gold substrate successfully monitored the electrografting of 2,2,6,6-tetremethyl-1-pipperridine N-oxyl (4-amino-TEMPO) and 4-nitrobenzene diazonium (4-NBD) in real-time using SEIRAS and cyclic voltammetry. The graphene-gold substrate is proficient at resolving the spectral responses of the electrografting and redox transformations of the grafted moieties occurring at the graphene interface spectroscopically and electrochemically. We clearly demonstrate highly sensitive SEIRAS in real-time at a graphene interface.This substrate opens a plethora of possibilities for real-time electrochemical ATR-SEIRAS investigations on graphitic interfaces such as catalysis, battery research, and CO2 capture and conversion, the latter of which our laboratory is exploring using modified electrodes. Electrochemical carbon capture and concentration (eCCC) methods have emerged as a prosing alternative to the current state-of-the-art temperature-pressure swing systems.7 Redox-active organic (RAO) molecules such as quinones have been reported to reversibly capture and release CO2. 7 However, they suffer from low solubility in solution. New approaches have explored immobilizing these molecules on electrode interfaces for increased CO2 adsorption capacities.11 Our graphene-gold allows a unique opportunity for an in situ mechanistic investigation of the immobilization and carbonization of RAOs. Our current work investigating immobilized aminoanthraquinone and a riboflavin derivative will be presented.

The ABCG2 Transporter Affects Plasma Levels, Tissue Distribution and Milk Secretion of Lumichrome, a Natural Derivative of Riboflavin.

The ABCG2 membrane transporter affects bioavailability and milk secretion of xenobiotics and natural compounds, including vitamins such as riboflavin. We aimed to characterize the in vitro and in vivo interaction of ABCG2 with lumichrome, the main photodegradation product of riboflavin, which has proven in vitro anti-cancer activity and a therapeutical role in antibacterial photodynamic therapy as an efficient photosensitizer. Using MDCK-II polarized cells overexpressing murine Abcg2 and human ABCG2 we found that lumichrome was efficiently transported by both variants. After lumichrome administration to wild-type and Abcg2-/- mice, plasma AUC20-120 min was 1.8-fold higher in Abcg2-/- mice compared with wild-type mice. The liver and testis from Abcg2-/- mice showed significantly higher lumichrome levels compared with wild-type, whereas lumichrome accumulation in small intestine content of wild-type mice was 2.7-fold higher than in Abcg2-/- counterparts. Finally, a 4.1-fold-higher lumichrome accumulation in milk of wild-type versus Abcg2-/- mice was found. Globally, our results show that ABCG2 plays a crucial role in plasma levels, tissue distribution and milk secretion of lumichrome potentially conditioning its biological activity.

Riboflavin derivatives as a novel electron transfer mediator for enhancing Cr(VI) removal by Shewanella oneidensis MR-1

Bioremediation has garnered considerable interest due to its advantages of economy and no secondary pollution. However, the direct electron transfer rate between microorganisms and Cr(VI) is low. The bioreduction of Cr(VI) by Shewanella oneidensis MR-1 (S. oneidensis) in the presence of formylmethylflavin (FMF) was conducted to understand how FMF mediated the extracellular electron transfer process to improve Cr(VI) removal. In this study, FMF was firstly synthesized by modifing RF to increase its solubility and redox activity. The findings indicate that S. oneidensis/FMF exhibited better Cr(VI) removal performance compared to that of S. oneidensis. Under optimum conditions, 40 mg/L Cr(VI) could be completely removed by S. oneidensis/FMF within 120 h, while only 48.6% of Cr(VI) was removed by S. oneidensis, and the first order rate constant (k) for the Cr(VI) elimination by S. oneidensis/FMF (0.033 h−1) was about 4.1-fold greater than that of S. oneidensis (0.008 h−1). Moreover, the removal of Cr(VI) by S. oneidensis/FMF were dominated by reduction, and supplemented by adsorption and complexation. FMF enhance the extracellular electron transfer rate (EETR) was confirmed by electrochemical cyclic voltammetry (CV) and linear scanning voltammetry (LSV) experiments. This study emphasizes the potential important role of FMF in environmental bioremediation.

Response of Neem Plants to Applied Salinity and Sprayed Riboflavin Derivatives

Response of Neem Plants to Applied Salinity and Sprayed Riboflavin Derivatives

Photochemical Crosslinking of Tarsal Collagen as a Treatment for Eyelid Laxity: Evaluation in Ex Vivo Human Tissue

Purpose: Experimental investigation in human eyelids to confirm that exposing excised tarsal plates to ultraviolet-A radiation can induce a stiffening effect through the riboflavin-photosensitized crosslinking of tarsal collagen. Methods: Thirteen tarsal plates excised from nonfrozen human cadavers were irradiated with ultraviolet-A rays (365 nm wavelength) at an irradiance of 75 mW/cm2 for 3 minutes, equivalent to a radiation fluence of 13.5 J/cm2, in the presence of a riboflavin derivative as a photosensitizer. The tensile stress (strength) and Young’s modulus (stiffness) of both nonirradiated and irradiated specimens were measured with the BioTester 5000 in the uniaxial mode at a strain of 10% and analyzed statistically. Individual specimens excised from 2 cadavers were also examined by routine histopathologic protocols to assess the effect of radiation on the Meibomian glands and collagen organization. Results: The irradiation enhanced both stiffness and strength of the human tarsal specimens, the difference between the test samples and controls being statistically significant (p < 0.0002 for n = 13). Histology indicated no damage to tarsal connective tissue or to Meibomian glands, and revealed a more compact packing of the collagen network located around the glands, which may be beneficial. The existence of collagen compaction was also supported by the reduction of samples’ thickness after irradiation (p = 0.0645). Conclusions: The irradiation of tarsal tissue with ultraviolet-A light of tarsus appears to be a safe and effective method for reducing eyelid laxity in human patients.

Hydrolysis of riboflavins in root exudates under iron deficiency and alkaline stress

Riboflavins are secreted under iron deficiency as a part of the iron acquisition Strategy I, mainly when the external pH is acidic. In plants growing under Fe-deficiency and alkaline conditions, riboflavins have been reported to accumulate inside the roots, with very low or negligible secretion. However, the fact that riboflavins may undergo hydrolysis under alkaline conditions has been so far disregarded. In this paper, we report the presence of riboflavin derivatives and products of their alkaline hydrolysis (lumichrome, lumiflavin and carboxymethylflavin) in nutrient solutions of Cucumis sativus plants grown under different iron regimes (soluble Fe-EDDHA in the nutrient solution, total absence of iron in the nutrient solution, or two different doses of FeSO4 supplied as a foliar spray), either cultivated in slightly acidic (pH 6) or alkaline (pH 8.8, 10 mM bicarbonate) nutrient solutions. The results show that root synthesis and exudation of riboflavins is controlled by shoot iron status, and that exuded riboflavins undergo hydrolysis, especially at alkaline pH, with lumichrome being the main product of hydrolysis.

Abstract 6689: Modulation of MAITs by the tumor microbiome in anti-PD1 treated cancers

Abstract Mucosal associated invariant T (MAIT) cells are innate like T-cells that recognize non-peptide metabolite antigens presented on monomorphic MHC related-1 (MR1) making them an attractive off the shelf tool for cancer immunotherapy. In this study, we investigate the function of MAITs in the tumor microenvironment of neoadjuvant anti-PD1 treated lung cancer patients (NCT02259621)). Paired single cell RNA/TCR sequencing analysis revealed an oligoclonal expansion of MAITs comprising of canonical TRAV1-2+ and non-canonical MAITs TRAV1-2-. TCR capture from canonical MAIT clonotypes confirmed their in vitro recognition of 5-OP-RU, bacteria derived riboflavin derivative as well as MR1 dependent recognition of live bacteria present in the tumor. However, the nature of antigens recognized by non-canonical MAITs remains poorly understood. On mining transcriptional profiles of MAITs, we observed that canonical MAITs were characterized by high clonal amplification whereas the non-canonical clonotypes expressed a strong activation/exhaustion gene signature. This suggests that both these MAIT populations recognize different types of antigens, differentially activate, and respond to immune checkpoint blockade (ICB), the non-canonical MAITs being highly dysfunctional. Using bacterial 16S RNA amplicon and metagenomics sequencing, we seek to identify microbial and metabolomics signatures associated with MAIT activation and response to ICB. We identified MAITs to express a cytotoxic signature making them a potential target for cancer immunotherapy. Our findings will provide further insights into the nature of ligands recognized by MAITs and the mechanisms associated with MR1 dependent recognition and tumor killing. Investigating the role of unconventional T-cells in response to ICB has the potential to reveal new tumor-associated antigens which could open avenues into innovative combination immunotherapies. Citation Format: Pakhi Birla, Lansaol Yang, Cynthia Sears, Fyza Shaikh, Drew Pardoll, Franck Housseau. Modulation of MAITs by the tumor microbiome in anti-PD1 treated cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6689.

Mantidisflavin A: A Riboflavin Derivative Featuring a 6/6/6/5/5 Skeleton from the Egg Cases of the Insect Tenodera sinensis Saussure and Its Anti-Renal Fibrosis Activity.

Mantidisflavin A (1) was isolated from the egg cases of the Tenodera sinensis Saussure. It exhibits an unprecedented 6/6/6/5/5 skeleton, accompanied by the formation of two additional fused heterocycles through a novel C-C bond and an oxygen bridge on riboflavin backbone. The structure of 1 was identified by spectroscopic and computational methods. To assess the inhibitory effect against renal fibrosis, compound 1 was evaluated in TGF-β1-induced rat kidney epithelial cells, and the results were compared to those of its precursor, riboflavin.

Riboflavin-based photocatalysis for aerobic oxidative S–N bond formation of thiols and amines

Abstract A novel organophotocatalytic process using riboflavin derivatives, which allows aerobic oxidative multistep S–S, S–N, and S–O bond formations of thiols and amines, is presented herein. The reaction proceeded under mild metal-free conditions using air (1 atm) as an environmentally friendly oxidant, yielding sulfinamides and sulfonamides.

G-CSF Induces MAIT Cells to Exert Anti-Intestinal Gvhd Effects Dependent on CXCR6 Mediated Chemotaxis

Background: Graft-versus-host disease (GVHD) is a major complication leading to transplantation failure and affecting overall survival after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Granulocyte colony-stimulating factor (G-CSF) can induce T-cell tolerance and altering graft cell components to enhance GVHD prevention in HSCT. MAIT cells are a group of innate immune T cells. It has unique biological properties, including mucosal tissue enrichment, activation of microbial riboflavin derivatives and tissue repair properties, which determine MAIT cells play an important role in mucosal immunity. Previous studies found that a high MAIT cell counts in G-CSF-induced grafts (G-grafts) is associated with a low incidence of gut acute GVHD (aGVHD). However, the effect of G-CSF mobilization on the phenotype, function and its regulatory mechanism in GVHD of MAIT cells has not been elucidated. Methods: MAIT cells in PB samples from four healthy donors before and after G-CSF treatment were isolated for Single-cell RNA sequencing (Figure A). Flow cytometry, In vitro experiment and NOD-SCID-IL-2Rg−/− mice (NSG mice) were performed to observe the phenotypic, functional changes and the dynamic distribution of G-CSF induced donor-derived MAIT cells in recipients. Results: A total of 80709 MAIT cells were divided into 14 clusters. Based on key type-associated marker genes and gene sets, MAIT0 (clusters 5 and 9), MAIT1 (cluster 7), MAIT2 (clusters 4 , 11), Cycling MAIT (clusters 12, 13), Activation (clusters 0, 8), ISG (cluster 10) and MAIT17 (clusters 1,2,3,6) cells were identified (Figure B,C). The subset frequencies and expression signatures of immunosuppression (MAIT2 and Cycling cells from cluster 12) and tissue repair (Activation and MAIT17) were both increased at both transcriptome (Figure D) and protein levels, suggesting that G-CSF enhanced the immune tolerance and tissue repair of MAIT cells. In vitro further confirmed that G-CSF-induced MAIT cells showed a stronger inhibition for T cell proliferation (Figure E). Besides, overexpressed genes related to immune regulation in G-CSF induced MAIT cells were strongly enriched in cluster 12 with high CSF3R (G-CSF receptor) expression (Figure F). The cluster 12 also showed the enrichment of IL-17 and TNF signaling pathway favoring tissue repair function, while downregulation of Graft-versus-host disease pathway upon G-CSF mobilization, indicating that G-CSF-CSF3R interaction is the potential mechanism by which G-CSF mediates the immune regulation of MAIT cells. Based on whether recipients develop gut aGVHD or not after allo-HSCT, the matched donor grafts after G-CSF mobilization were divided into two groups termed as post-G-CSF-no gut aGVHD (n=3) and post-G-CSF-gut aGVHD group (n=1), respectively. MAIT cells in post-G-CSF-no gut aGVHD group overexpressed CSF3R and its related target genes, immunosuppression and tissue repair signatures compared with post-G-CSF-gut aGVHD. Besides, MAIT cells from G-PB of recipients without gut aGVHD had a higher CD114 expression (encoded by CSF3R) compared with gut aGVHD (Figure G). These results suggest that the enhanced immunosuppression and tissue repair function of MAIT cells induced by G-CSF may exert anti-gut aGVHD effects. The NSG mice confirmed MAIT cells in G-PB to tissue chemotaxis especially gut tissues in the recipient. A higher frequency of MAIT cells was detected in gut tissues in the low GVHD score group (GVHD Score＜3) compared with the high GVHD score group on post-transplant day 14 (Figure H), suggesting recruited MAIT cells in gut tissues favor anti-gut aGVHD effect. Moreover, CXCR6 was the most differentially expressed chemotactic receptor gene between post-G-CSF-no gut aGVHD and post-G-CSF-gut aGVHD group (Figure I). At the protein level, the expression of CXCR6 were significantly upregulated upon G-CSF administration (Figure I), while that of CCR6 and CXCR6 was non-significant. These results suggest that CXCR6 is necessary for G-CSF induced MAIT cell recruitment to the gut tissue. Conclusions G-CSF-CSF3R interactions enhance immunosuppression and tissue repair functions of MAIT cells, while CXCR6 recruits these functional MAIT cells from PB to gut tissues to exert anti-gut aGVHD effect. Hence, CSF3R and CXCR6 expression in G-CSF mobilized MAIT cells is crucial to exert anti-gut aGVHD effect, and has the potential to become predictive markers of gut aGVHD.

Flavin mononucleotide in visible light photoinitiating systems for multiple-photocrosslinking and photoencapsulation strategies

Visible light-induced photocrosslinking techniques have attracted significant attention for their flexibility, controllability, safety, and energy conservation, especially in tissue engineering and biofabrication, compared to UV photocrosslinking. Despite these advantages, current photoinitiators are constrained by various challenges, including inadequate photoinitiation efficiency, low biocompatibility, poor water solubility, and limited compatibility with diverse crosslinking systems. Here, a water-soluble derivative of riboflavin, flavin mononucleotide (FMN−), was used to assess its potential as an initiator in multiple-photocrosslinking systems, including radical photopolymerization, dityrosine, and ditryptophan coupling crosslinking, under blue light irradiation. Blue light irradiation facilitated an efficient electron transfer reaction between FMN− and persulfate, owing to their suitable spectral compatibility and photoactivity. The resulting oxidizing free radicals and excited triplet state of FMN− served as initiating active species for the multiple-photocrosslinking reactions. The combination of FMN− and potassium persulfate (KPS) exhibited exceptional photoinitiation efficiency for various biomaterials, including silk fibroin, gelatin, poly(ethylene glycol) diacrylate, and carboxymethyl cellulose modified with amino acids. Furthermore, the cytocompatibility of the FMN−/KPS photoinitiator was demonstrated by the survival rates of 3T3-LI fibroblasts encapsulated in it, which exceeded 95 % when compared to a commercial initiator. Statement of significanceBy introducing persulfate, the photoinitiation efficiency of flavin mononucleotide was significantly improved. The application scenarios of flavin mononucleotide and persulfate combinations were also greatly extended, including radical photopolymerization, dityrosine, diphenylalanine, and ditryptophan coupling crosslinking. Among them, the coupling crosslinking of amino acids (di-phenylalanine, and di-tryptophan) modified carboxymethyl cellulose, to our knowledge, was first reported. The excellent cytocompatibility of cell encapsulation further proved that the combinations of flavin mononucleotide and persulfate have great potential in tissue engineering.

Singlet oxygen quenching by riboflavin

Derivatives of riboflavin, specifically flavin mononucleotide, have recently been in the spotlight as the active component in protein-encased optogenetic sensitizers to produce singlet oxygen, O2(a1Δg). The extent to which riboflavin deactivates singlet oxygen is a key aspect of these studies. Surprisingly, there is a dearth of information on the rate constant for riboflavin-mediated singlet oxygen removal. We now report that, when riboflavin is dissolved in both aqueous and methanol solutions, the rate constant for riboflavin-mediated removal of singlet oxygen cannot be greater than ∼ 2 × 105 s−1 M−1. This number is appreciably smaller than what has been reported in the literature and, as such, our results can be significant for mechanistic interpretations of photo-processes that involve flavins in oxygenated systems.

Tetraacetyl riboflavin derivative mediates caspase 3/7 activation via MAPK in A431 cells upon blue light influence.

An acetylated riboflavin derivative, 3-methyl-tetraacetyl riboflavin (3MeTARF), is a compound with high photostability and photophysical properties similar to riboflavin, including the ability to photogenerate singlet oxygen. In the present study, we compared the effects of irradiation on A431 cancer cells with blue LED light (438 nm) in the presence of 3MeTARF and riboflavin on MAPK phosphorylation, apoptosis, caspase 3/7 activation and PARP cleavage. We observed that photogenerated oxidative stress in this reaction activates MAPK by increasing phosphorylation of p38 and JNK proteins. Preincubation of cells with inhibitors specific for phosphorylation of p38 and JNK proteins (SB203580, SP600125), respectively, results in decreased caspase 3/7 activation and PARP cleavage. We showed that the tetraacetyl derivative more effectively activates MAPK and skin cancer cell death compared to riboflavin. These data, together with results of our previous study, support the hypothesis that 3MeTARF, of riboflavin, might be more useful and desirable as a compound for use in photodynamic oxidation processes, including its therapeutic potential.

Immune checkpoint blockade unleashes Mucosal Associated Invariant T (MAIT) cells in tumor microenvironment of NSCLC patients

Abstract Mucosal associated invariant T (MAIT) cells are innate like T-cells that recognise non-peptide metabolite antigens presented on monomorphic MHC related-1 (MR1) making them an attractive off the shelf tool for cancer immunotherapy. In this study, we investigate the function of MAITs in the tumor microenvironment of neoadjuvant anti-PD1 treated lung cancer patients (NCT02259621). Paired single cell RNA/TCR sequencing analysis revealed an oligoclonal expansion of MAITs comprising of canonical TRAV1–2 +and non-canonical MAITs TRAV1–2 −. TCR capture from canonical MAIT clonotypes confirmed their in vitro recognition of 5-OP-RU, a bacteria derived riboflavin derivative. However, the nature of antigens recognised by non-canonical MAITs remains poorly understood. On mining transcriptional profiles of MAITs, we observed a high clonal amplification of canonical MAITs whereas the non-canonical clonotypes expressed a strong activation/exhaustion gene signature. This suggests that both these MAIT populations recognise different types of antigens, are differentially activated, and respond to immune checkpoint blockade (ICB), the non-canonical MAITs being highly dysfunctional. Using bacterial 16S RNA amplicon and metagenomics sequencing, we seek to identify microbial and metabolomic signatures associated with MAIT activation and response to ICB. Our findings will provide further insights into the nature of ligands recognised by MAITs and the mechanisms associated with MR1 dependent recognition and killing of tumor. Investigating the role of unconventional T-cells in response to ICB has the potential to reveal new tumor-associated antigens which could open up avenues for innovative combination immunotherapies.

Effects of Ultraviolet-A Radiation on Enzymatically Degraded Tunica Adventitia of the Porcine Abdominal Aorta

In a previous study, it was demonstrated that mechanical stiffness and strength of tunica adventitia of the porcine abdominal aortic wall were significantly increased following irradiation of the tissue with ultraviolet-A (UV-A) rays, due to photochemically induced crosslinking of adventitial collagen. We have suggested that by applying the process to a dilated aortic region caused by abdominal aortic aneurysm (AAA), the mechanically enhanced adventitial layer might delay or prevent the aneurysmal rupture in human patients. Here, we extended our investigation to adventitial tissue that was subjected to in vitro collagenolysis, such mimicking a step in the aneurysmal pathomechanism. We used tissue isolated from the abdominal region of healthy porcine aortas. The adventitia was separated from the vessel wall and subjected to enzymatic degradation and exposed to UV-A radiation (365 nm) in the presence of a riboflavin derivative as the photo-initiator. Collagenolysis was carried out in buffered medium at two different durations (8 and 20 h). Three sets, each of 30 adventitial specimens (degraded only, irradiated prior to degradation, and irradiated after degradation), were evaluated in a mechanical tester. Mechanical testing of the irradiated specimens indicated an increase in both stiffness (Young’s modulus) and strength (ultimate stress) when compared to those degraded but not irradiated. The specimens first irradiated and then degraded displayed the highest gain in both stiffness and strength. In the tissue irradiated after degradation, the longer the duration of collagenolysis, the less significant was the enhancement of mechanical characteristics. Irradiation with UV-A rays of enzymatically degraded aortic adventitial tissue induced the enhancement of its mechanical characteristics (stiffness, strength). The earlier the irradiation, the more significant the enhancement.

Flavin Adenine Dinucleotide (FAD) and Pyridoxal 5'-Phosphate (PLP) Bind to Sox9 and Alter the Expression of Key Pancreatic Progenitor Transcription Factors.

Cofactor flavin adenine dinucleotide (FAD), a compound with flavin moiety and a derivative of riboflavin (vitamin B2), is shown to bind to Sox9 (a key transcription factor in early pancreatic development) and, subsequently, induce a large increase in markers of pancreatic development, including Ngn3 and PTF1a. Pyridoxal 5'-phosphate (PLP), the active form of vitamin B6, also binds to Sox9 and results in a similar increase in pancreatic development markers. Sox9 is known to be specifically important for pancreatic progenitors. Previously, there was no known link between FAD, PLP, or other co-factors and Sox9 for function. Thus, our findings show the mechanism by which FAD and PLP interact with Sox9 and result in the altered expression of pancreatic progenitor transcription factors involved in the pancreas development.

A higher plant FAD synthetase is fused to an inactivated FAD pyrophosphatase

The riboflavin derivatives FMN and flavin adenine dinucleotide (FAD) are critical cofactors for wide-ranging biological processes across all kingdoms of life. Although it is well established that these flavins can be readily interconverted, in plants, the responsible catalysts and regulatory mechanisms remain poorly understood. Here, we report the cloning and biochemical characterization of an FAD synthetase encoded by the gene At5g03430, which we have designated AtFADS1 (A.thaliana FADS1). The catalytic properties of the FAD synthetase activity are similar to those reported for other FAD synthetases, except that we observed maximum activity with Zn2+ as the associated divalent metal cation. Like human FAD synthetase, AtFADS1 exists as an apparent fusion with an ancestral FAD pyrophosphatase, a feature that is conserved across plants. However, we detected no pyrophosphatase activity with AtFADS1, consistent with an observed loss of a key catalytic residue in higher plant evolutionary history. In contrast, we determined that algal FADS1 retains both FAD synthetase and pyrophosphatase activity. We discuss the implications, including the potential for yet-unstudied biologically relevant noncatalytic functions, and possible evolutionary pressures that have led to the loss of FAD pyrophosphatase activity, yet universal retention of an apparently nonfunctional domain in FADS of land plants.

Covalent flavoproteins: types, occurrence, biogenesis and catalytic mechanisms.

Flavoproteins are proteins that contain a nucleic acid derivative of riboflavin: flavin adenine dinucleotide (FAD) or flavin mononucleotide (FMN). Flavoproteins are involved in a wide array of biological processes, such as photosynthesis, DNA repair and natural product biosynthesis. It should be noted that 5%-10% of flavoproteins have a covalently linked flavin prosthetic group. Such covalent linkages benefit the holoenzyme in several ways including improving the stability and catalytic potency. During the past decade, significant progress has been made in covalent flavoproteins, especially with respect to enzyme-dependent biogenesis and discovery of novel linkage types. The present review gives a condensed overview of investigations published from March 2009 to December 2021, with emphasis on the discovery, biogenesis and their catalytic role in natural product biosynthesis.

Tuning riboflavin derivatives for photodynamic inactivation of pathogens

The development of effective pathogen reduction strategies is required due to the rise in antibiotic-resistant bacteria and zoonotic viral pandemics. Photodynamic inactivation (PDI) of bacteria and viruses is a potent reduction strategy that bypasses typical resistance mechanisms. Naturally occurring riboflavin has been widely used in PDI applications due to efficient light-induced reactive oxygen species (ROS) release. By rational design of its core structure to alter (photo)physical properties, we obtained derivatives capable of outperforming riboflavin’s visible light-induced PDI against E. coli and a SARS-CoV-2 surrogate, revealing functional group dependency for each pathogen. Bacterial PDI was influenced mainly by guanidino substitution, whereas viral PDI increased through bromination of the flavin. These observations were related to enhanced uptake and ROS-specific nucleic acid cleavage mechanisms. Trends in the derivatives’ toxicity towards human fibroblast cells were also investigated to assess viable therapeutic derivatives and help guide further design of PDI agents to combat pathogenic organisms.

Biomimetic photooxidation of noscapine sensitized by a riboflavin derivative in water: The combined role of natural dyes and solar light in environmental remediation.

Noscapine (NSC) is a benzyl-isoquinoline alkaloid discovered in 1930 as an antitussive agent. Recently, NSC has also been reported to exhibit antitumor activity and, according to computational studies, it is able to attack the protease enzyme of Coronavirus (COVID-19) and thus could be used as antiviral for COVID-19 pandemic. Therefore, an increasing use of this drug could be envisaged in the coming years. NSC is readily metabolized with a half-life of 4.5h giving rise to cotarnine, hydrocotarnine, and meconine, arising from the oxidative breaking of the CC bond between isoquinoline and phthalide moieties. Because of its potentially increasing use, high concentrations of NSC but also its metabolites will be delivered in the environment and potentially affect natural ecosystems. Thus, the aim of this work is to investigate the degradation of NSC in the presence of naturally occurring photocatalysts. As a matter of fact, the present contribution has demonstrated that NSC can be efficiently degraded in the presence of a derivative of the natural organic dye Riboflavin (RFTA) upon exposure to visible light. Indeed, a detailed study of the mechanism involved in the photodegradation revealed the similarities between the biomimetic and the photocatalyzed processes. In fact, the main photoproducts of NSC were identified as cotarnine and opianic acid based on a careful UPLC-MS2 analysis compared to the independently synthesized standards. The former is coincident with one of the main metabolites obtained in humans, whereas the latter is related to meconine, a second major metabolite of NSC. Photophysical experiments demonstrated that the observed oxidative cleavage is mediated mainly by singlet oxygen in a medium in which the lifetime of 1O2 is long enough, or by electron transfer to the triplet excited state of RFTA if the photodegradation occurs in aqueous media, where the 1O2 lifetime is very short.