Assessment of NSAIDs as potential inhibitors of the fatty acid amide hydrolase I (FAAH-1) using three different primary fatty acid amide substrates in vitro

Background Pain relief remains a major subject of inadequately met need of patients. Therapeutic agents designed to treat pain and inflammation so far have low to moderate efficiencies with significant untoward side effects. FAAH-1 has been proposed as a promising target for the discovery of drugs to treat pain and inflammation without significant adverse effects. FAAH-1 is the primary enzyme accountable for the degradation of AEA and related fatty acid amides. Studies have revealed that the simultaneous inhibition of COX and FAAH-1 activities produce greater pharmacological efficiency with significantly lowered toxicity and ulcerogenic activity. Recently, the metabolism of endocannabinoids by COX-2 was suggested to be differentially regulated by NSAIDs. Methods We analysed the affinity of oleamide, arachidonamide and stearoylamide at the FAAH-1 in vitro and investigated the potency of selected NSAIDs on the hydrolysis of endocannabinoid-like molecules (oleamide, arachidonamide and stearoylamide) by FAAH-1 from rat liver. NSAIDs were initially screened at 500 μM after which those that exhibited greater potency were further analysed over a range of inhibitor concentrations. Results The substrate affinity of FAAH-1 obtained, increased in a rank order of oleamide < arachidonamide < stearoylamide with resultant Vmax values in a rank order of arachidonamide > oleamide > stearoylamide. The selected NSAIDs caused a concentration-dependent inhibition of FAAH-1 activity with sulindac, carprofen and meclofenamate exhibiting the greatest potency. Michaelis-Menten analysis suggested the mode of inhibition of FAAH-1 hydrolysis of both oleamide and arachidonamide by meclofenamate and indomethacin to be non-competitive in nature. Conclusion Our data therefore suggest potential for study of these compounds as combined FAAH-1-COX inhibitors.

The Biosynthesis and Metabolism of the N-Acylated Aromatic Amino Acids: N-Acylphenylalanine, N-Acyltyrosine, N-Acyltryptophan, and N-Acylhistidine

The fatty acid amides are a family of lipids composed of two chemical moieties, a fatty acid and a biogenic amine linked together in an amide bond. This lipid family is structurally related to the endocannabinoid anandamide (N-arachidonoylethanolamine) and, thus, is frequently referred to as a family of endocannabinoid-related lipids. The fatty acid amide family is divided into different classes based on the conjugate amine; anandamide being a member of the N-acylethanolamine class (NAE). Another class within the fatty acid amide family is the N-acyl amino acids (NA-AAs). The focus of this review is a sub-class of the NA-AAs, the N-acyl aromatic amino acids (NA-ArAAs). The NA-ArAAs are not broadly recognized, even by those interested in the endocannabinoids and endocannabinoid-related lipids. Herein, the NA-ArAAs that have been identified from a biological source will be highlighted and pathways for their biosynthesis, degradation, enzymatic modification, and transport will be presented. Also, information about the cellular functions of the NA-ArAAs will be placed in context with the data regarding the identification and metabolism of these N-acylated amino acids. A review of the current state-of-knowledge about the NA-ArAAs is to stimulate future research about this underappreciated sub-class of the fatty acid amide family.

Cascading Effects of Root Microbial Symbiosis on the Development and Metabolome of the Insect Herbivore Manduca Sexta L.

Root mutualistic microbes can modulate the production of plant secondary metabolites affecting plant–herbivore interactions. Still, the main mechanisms underlying the impact of root mutualists on herbivore performance remain ambiguous. In particular, little is known about how changes in the plant metabolome induced by root mutualists affect the insect metabolome and post-larval development. By using bioassays with tomato plants (Solanum lycopersicum), we analyzed the impact of the arbuscular mycorrhizal fungus Rhizophagus irregularis and the growth-promoting fungus Trichoderma harzianum on the plant interaction with the specialist insect herbivore Manduca sexta. We found that root colonization by the mutualistic microbes impaired insect development, including metamorphosis. By using untargeted metabolomics, we found that root colonization by the mutualistic microbes altered the secondary metabolism of tomato shoots, leading to enhanced levels of steroidal glycoalkaloids. Untargeted metabolomics further revealed that root colonization by the mutualists affected the metabolome of the herbivore, leading to an enhanced accumulation of steroidal glycoalkaloids and altered patterns of fatty acid amides and carnitine-derived metabolites. Our results indicate that the changes in the shoot metabolome triggered by root mutualistic microbes can cascade up altering the metabolome of the insects feeding on the colonized plants, thus affecting the insect development.

Assessment of NSAIDS as Potential Inhibitors of the Fatty Acid Amide Hydrolase I (FAAH-1) using Three Different Primary Fatty Acid Amide Substrates In Vitro

Pain relief · remains a major subject of inadequately met need of patients. Therapeutic agents designed to treat pain and inflammation so far have low to moderate efficiencies with significant untoward side effects. FAAH-1 has been proposed as a promising target for the discovery of drugs to treat pain and inflammation without significant adverse effects. FAAH-1 is the primary enzyme accountable for the degradation of AEA and related fatty acid amides. Studies have revealed that the simultaneous inhibition of COX and FAAH-1 activities produce greater pharmacological efficiency with significantly lowered toxicity and ulcerogenic activity. Recently, the metabolism of endocannabinoids by COX-2 was suggested to be differentially regulated by NSAIDs. We analysed the affinity of ODA, ArDA and SyDA at the FAAH-1 in vitro and investigated the potency of selected NSAIDs on the hydrolysis of endocannabinoid-like molecules (ODA, ArDA and SyDA) by FAAH-1 from rat liver. NSAIDs were initially screened at 500 µM after which those that exhibited greater potency were further analysed over a range of inhibitor concentrations. The substrate affinity of FAAH-1 obtained, increased in a rank order of ODA < ArDA < SyDA with resultant Vmax values in a rank order of ArDA > ODA > SyDA. The selected NSAIDs caused a concentration-dependent inhibition of FAAH-1 activity with sulindac, carprofen and meclofenamate exhibiting the greatest potency. Michaelis-Menten analysis suggested the mode of inhibition of FAAH-1 hydrolysis of both ODA and ArDA by meclofenamate and indomethacin to be non-competitive in nature. Our data therefore suggest potential for study of these compounds as combined FAAH-1-COX inhibitors.

Distribution of an Analgesic Palmitoylethanolamide and Other N-Acylethanolamines in Human Placental Membranes

Human amniotic and amniochorionic membranes (AM, ACM) are the most often used grafts accelerating wound healing due to their anti-inflammatory, anti-microbial, anti-fibrotic, and analgesic properties. We assessed the distribution of endogenous fatty acid amides N-acylethanolamines (NAEs): palmitoylethanolamide (PEA), oleoylethanolamide (OEA) and anandamide (AEA) in placental tissues, as they could participate in wound healing properties of AM/ACM grafts. Ten placentas were collected after caesarean delivery. NAEs were detected using ultra-high-performance liquid chromatography-tandem mass spectrometry analysis in fresh samples of AM, ACM, placental disc, umbilical cord, umbilical serum and vernix caseosa), and decontaminated samples of AM and ACM. NAEs were present in all studied tissue types, with mean concentrations in fresh tissues ranging: 76 – 350 (PEA); 31 – 220 (OEA); 7 – 30 ng/g (AEA). The highest mean concentrations were found in AM (PEA: 350 ng/g) or placenta (OEA: 220 ng/g; AEA: 30 ng/g), respectively. Low levels of NAEs were found in serum and vernix. Decontamination of AM, but not ACM, induced a significant (3 – 3.5-fold) increase in the levels of NAEs. PEA is the first compound with direct analgesic effect detected in AM and ACM. We thus propose NAEs, especially PEA, as one of the possible factors responsible for the anti-hyperalgesic, anti-inflammatory and neuroprotective effects of AM/ACM grafts, stimulating their wound healing effect. The increase of NAE levels in AM and ACM after tissue decontamination indicates that tissue processing may play an important role in maintaining the analgesic effect.

Synthesis, Quantification, and Characterization of Fatty Acid Amides from In Vitro and In Vivo Sources

Fatty acid amides are a diverse family of underappreciated, biologically occurring lipids. Herein, the methods for the chemical synthesis and subsequent characterization of specific members of the fatty acid amide family are described. The synthetically prepared fatty acid amides and those obtained commercially are used as standards for the characterization and quantification of the fatty acid amides produced by biological systems, a fatty acid amidome. The fatty acid amidomes from mouse N18TG2 cells, sheep choroid plexus cells, Drosophila melanogaster, Bombyx mori, Apis mellifera, and Tribolium castaneum are presented.

Skin imprints to provide noninvasive metabolic profiling of COVID-19 patients

As the current COVID-19 pandemic progresses, more symptoms and signals related to how the disease manifests in the human body arise in the literature. Skin lesions and coagulopathies may be confounding factors on routine care and patient management. We analyzed the metabolic and lipidic profile of the skin from COVID-19 patients using imprints in silica plates as a non-invasive alternative, in order to better understand the biochemical disturbances caused by SARS-CoV-2 in the skin. One hundred and one patients (64 COVID-19 positive patients and 37 control patients) were enrolled in the study from April 2020 to June 2020 during the first wave of COVID-19 in São Paulo, Brazil. Fourteen biomarkers were identified related to COVID-19 infection (7 increased and 7 decreased in COVID-19 patients). Remarkably, oleamide has shown promising performance, providing 79.0% of sensitivity on a receiver operating characteristic curve model. Species related to coagulation and immune system maintenance such as phosphatidylserines were decreased in COVID-19 patients; on the other hand, cytokine storm and immunomodulation may be affected by molecules increased in the COVID-19 group, particularly primary fatty acid amides and N-acylethanolamines, which are part of the endocannabinoid system. Our results show that skin imprints may be a useful, noninvasive strategy for COVID-19 screening, by electing a pool of biomarkers with diagnostic potential.

Metabolite Profiling of Christia vespertilionis Leaf Metabolome via Molecular Network Approach

Christia vespertilionis (L.f.) Bakh. f. is an ornamental plant with unique butterfly-shaped leaves, hence its vernacular name “butterfly wing” or “rerama” in Malay. In Malaysia, the green-leafed variety of this plant has gained popularity in recent years due to testimonial reports by local consumers of its medicinal uses, which include treatment for cancer. Despite these popular uses, there is very limited information on the phytochemistry of the leaf of this plant, presenting a significant gap in the cheminformatics of the plant species. Herein, we report a substantially detailed phytochemical profile of the leaf metabolome of the green-leafed variety of C. vespertilionis, obtained by deploying an untargeted tandem mass spectrometry-based molecular networking approach. The detailed inspection of the molecular network map generated for the leaf metabolome enabled the putative identification of 60 metabolites, comprising 13 phenolic acids, 20 flavonoids, 2 benzyltetrahydroisoquinoline-type alkaloids, 4 hydroxyjasmonic acid derivatives, 2 phenethyl derivatives, 3 monoacylglycerols, 4 fatty acid amides, 2 chlorophyll derivatives, 4 carotenoids, 2 organic acids, 1 nucleoside, and 3 amino acids. Flavonoids are the major class of metabolites that characterize the plant leaves. Employing a mass-targeted isolation approach, two new derivatives of apigenin-6-C-β-glucoside, the major constituents of the plant leaf, were successfully purified and spectroscopically characterized as apigenin-6-C-β-glucoside 4′-O-α-apiofuranoside (28) and apigenin-6-C-β-[(4″,6″-O-dimalonyl)-glucoside] 4′-O-α-apiofuranoside (47). This work provides further information on the chemical space of the plant leaf, which is a prerequisite to further research towards its valorization as a potential phytopharmaceutical product.