SummaryAlthough in recent years there has been an increased awareness of the widespread nature of biofluorescence in the marine environment, the diversity of the molecules responsible for this luminescent phenotype has been mostly limited to green fluorescent proteins (GFPs), GFP-like proteins, and fluorescent fatty acid-binding proteins (FABPs). In the present study, we describe a previously undescribed group of brominated tryptophan-kynurenine small molecule metabolites responsible for the green biofluorescence in two species of sharks and provide their structural, antimicrobial, and spectral characterization. Multi-scale fluorescence microscopy studies guided the discovery of metabolites that were differentially produced in fluorescent versus non-fluorescent skin, as well as the species-specific structural details of their unusual light-guiding denticles. Overall, this study provides the detailed description of a family of small molecules responsible for marine biofluorescence and opens new questions related to their roles in central nervous system signaling, resilience to microbial infections, and photoprotection.

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A novel and efficient synthetic pathway toward known meso-tetraphenylporpholactams, also applicable to the synthesis of hitherto unknown and inaccessible meso-CF-substituted porpholactam, is detailed (dioxochlorin → dioxochlorin urea adduct → porpholactam). meso-Tetraphenylporpholactam was converted to an imidazoloporphyrin-α-triflate derivative that was demonstrated to be of utility for the generation of functionalized imidazoloporphyrins with a substituted amine adjacent to the outside N atom of the imidazole moiety (using pyridine, EtNH, diethyliminodiacetic acetate, dipicolylamine (DPA), and cyclen). The DPA- and iminodiacetate-imidazoloporphyrin conjugates were structurally characterized. The chemosensing potential of the metal chelate-imidazoloporphyrin conjugates was evaluated, though their constrained metric parameters led to muted chemosensing responses to various divalent metal ions. The accessibility of the meso-arylporpholactams and the meso-tetraphenylimidazoloporphyrin triflate enables the continued exploration of porphyrin-like pyrrole-modified porphyrins that incorporate a nitrogen atom in place of a β-carbon atom in their macrocycles.

To date, only two pigments have been identified in avian eggshells: rusty-brown protoporphyrin IX and blue-green biliverdin IXα. Most avian eggshell colours can be produced by a mixture of these two tetrapyrrolic pigments. However, tinamou (Tinamidae) eggshells display colours not easily rationalised by combination of these two pigments alone, suggesting the presence of other pigments. Here, through extraction, derivatization, spectroscopy, chromatography, and mass spectrometry, we identify two novel eggshell pigments: yellow–brown tetrapyrrolic bilirubin from the guacamole-green eggshells of Eudromia elegans, and red–orange tripyrrolic uroerythrin from the purplish-brown eggshells of Nothura maculosa. Both pigments are known porphyrin catabolites and are found in the eggshells in conjunction with biliverdin IXα. A colour mixing model using the new pigments and biliverdin reproduces the respective eggshell colours. These discoveries expand our understanding of how eggshell colour diversity is achieved. We suggest that the ability of these pigments to photo-degrade may have an adaptive value for the tinamous.

A laboratory experiment is described that extracts the tetrapyrrolic teal-colored biliverdin IXα, as its dimethyl ester, from commercially available emu eggshells. The extraction of ∼10 mg samples of biliverdin is simple and requires two 3 h lab periods: A two-step acid digestion and liquid–liquid extraction, followed by short silica gel flash column chromatography. The deeply colored extract can be characterized by TLC, UV–vis, IR, 1H NMR spectroscopy, ESI+ mass spectrometry, or its diagnostic chemical reduction to the yellow derivative bilirubin. A small-scale variant of this extraction requires only a single 3 h lab period, and the extracts can be analyzed by UV–vis spectroscopy, ESI+ MS, or HPLC. An introduction to the biological origin of biliverdin is provided. This pedagogically flexible project combines natural product isolation with the spectroscopic characterization of a multifunctional biomedically important compound and touches upon many aspects of chemistry and biology. The colorful experiment is appropriate for the introductory organic chemistry laboratory and possesses the prerequisites for a chemistry–biology interlaboratory approach.

The entomopathogenic bacterium Xenorhabdus bovienii exists in a mutualistic relationship with nematodes of the genus Steinernema. Free-living infective juveniles of Steinernema prey on insect larvae and regurgitate X. bovienii within the hemocoel of a host larva. X. bovienii subsequently produces a complex array of specialized metabolites and effector proteins that kill the insect and regulate various aspects of the trilateral symbiosis. While Xenorhabdus species are rich producers of secondary metabolites, many of their biosynthetic gene clusters remain uncharacterized. Here, we describe a nonribosomal peptide synthetase (NRPS) identified through comparative genomics analysis that is widely conserved in Xenorhabdus species. Heterologous expression of this NRPS gene from X. bovienii in E. coli led to the discovery of a family of lipo-tripeptides that chromatographically appear as pairs, containing either a C-terminal carboxylic acid or carboxamide. Coexpression of the NRPS with the leupeptin protease inhibitor pathway enhanced production, facilitating isolation and characterization efforts. The new lipo-tripeptides were also detected in wild-type X. bovienii cultures. These metabolites, termed bovienimides, share an uncommon C-terminal d-citrulline residue. The NRPS lacked a dedicated chain termination domain, resulting in product diversification and release from the assembly line through reactions with ammonia, water, or exogenous alcohols.

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