Background: Butyric acid (BA) is a short-chain fatty acid (SCFA) with anti-inflammatory properties, which promotes intestinal barrier function. Medium-chain fatty acids (MCFA), including caproic acid (CA), promote TH1 and TH17 differentiation, thus supporting inflammation. Aim: Since most SCFAs are absorbed in the cecum and colon, the measurement of BA in peripheral blood could provide information on the health status of the intestinal ecosystem. Additionally, given the different immunomodulatory properties of BA and CA the evaluation of their serum concentration, as well as their ratio could be as a simple and rapid biomarker of disease activity and/or treatment efficacy in MS. Methods: We evaluated serum BA and CA concentrations, immune parameters, intestinal barrier integrity and the gut microbiota composition in patients with multiple sclerosis (MS) comparing result to those obtained in healthy controls. Results: In MS, the concentration of BA was reduced and that of CA was increased. Concurrently, the microbiota was depleted of BA producers while it was enriched in mucin-degrading, pro-inflammatory components. The reduced serum concentration of BA seen in MS patients correlated with alterations of the barrier permeability, as evidenced by the higher plasma concentrations of lipopolysaccharide and intestinal fatty acid-binding protein, and inflammation. Specifically, CA was positively associated with CD4+/IFNγ+ T lymphocytes, and the BA/CA ratio correlated positively with CD4+/CD25 high /Foxp3+ and negatively with CD4+/IFNγ+ T lymphocytes. Conclusion: The gut microbiota dysbiosis found in MS is possibly associated with alterations of the SCFA/MCFA ratio and of the intestinal barrier; this could explain the chronic inflammation that characterizes this disease. SCFA and MCFA quantification could be a simple biomarker to evaluate the efficacy of therapeutic and rehabilitation procedures in MS.

Recently, an increasing number of pharmacists had to supply medicinal products based on L. (Cannabaceae), prescribed by physicians to individual patients. Cannabis olive oil preparation is the first choice as a concentrated extract of cannabinoids, even though standardized operative conditions for obtaining it are still not available. In this work, the impact of temperature and extraction time on the concentration of active principles was studied to harmonize the different compounding methods, optimize the extraction process, and reduce the variability among preparations. Moreover, starting from the cannabis inflorescence, the effect of temperature on tetrahydrocannabinolic acid decarboxylation was evaluated. For the analysis, a GC/MS method, as suggested by the Italian Ministry of Health, and a GC/flame ionization detection method were developed, validated, and compared.

Hexahydrocannabinols (HHCs), referred to (9R)-HHC and (9S)-HHC diastereoisomers, are low-studied cannabinoids naturally found in small concentrations in the pollen and the seeds of the hemp plants. Despite the lack of in-depth studies about HHCs activity, potency, toxicity, and safety, these cannabinoids are emerging on the light-cannabis (hemp) market probably because legislations still do not clearly regulate them. Here, we describe for the first time the finding of HHCs (42% of (9R)-HHC and 24% of (9S)-HHC) in two samples of hemp-derived resin. The achievement of reference standards by semi-synthetic or isolation approach allows us to develop and validate a gas chromatography mass spectrometry (GC-MS) method for the identification and quantification of HHCs in hemp-derived products. A thorough investigation could be carried out to reveal the possible addition of "new" compounds that might be a matter of safety.

The two diastereomeric pairs of acidic amino acids 5-(2-amino-2-carboxyethyl)-4,5-dihydroisoxazole-3-carboxylic acid (8A/8B) and 4-(2-amino-2-carboxyethyl)-5,5-dimethyl-4,5-dihydroisoxazole-3-carboxylic acid (10A/10B) were prepared via a strategy based on a 1,3-dipolar cycloaddition. The four amino acids were tested at ionotropic and metabotropic glutamate receptors. None of the compounds was active, neither as agonists nor as antagonists, at 1 mM on metabotropic receptors (mGluR1, -2, -4, and -5 expressed in CHO cell lines). Conversely, the pair of stereoisomers 8A/8B showed a remarkable affinity, antagonist potency, and selectivity for NMDA receptors, when tested on ionotropic glutamate receptors. The affinity of 8A proved to be 5 times higher than that of diastereomer 8B (K(i) values 0.21 and 0.96 microM, respectively). Furthermore, compounds 8A and 8B exhibited a noteworthy anticonvulsant activity in in vivo tests on DBA/2 mice. Derivative 10A was inactive at all ionotropic glutamate receptors, whereas its stereoisomer 10B displayed a seizable binding to both NMDA and AMPA receptors.

The inhibition of the PCSK9/LDLR protein− protein interaction is a promising strategy for developing new hypocholesterolemic agents. Familial hypercholesterolemia is linked to specific PCSK9 mutations: the D374Y is the most potent gain-offunction (GOF) PCSK9 mutation among clinically relevant ones. Recently, a lupin peptide (T9) showed inhibitory effects on this mutant PCSK9 form, being also capable to increase liver uptake of low density lipoprotein cholesterol. In this Letter, aiming to improve the potency of this peptide, the T9 residues mainly responsible for the interaction with PCSK9 D374Y (hot spots) were computationally predicted. Then, the "non-hot" residues were suitably substituted by new amino acids capable to theoretically increase the structural complementarity between T9 and PCSK9 D374Y . The outcomes of this study were confirmed by in vitro biochemical assays and cellular investigations, showing that a new T9 analog is able to increase the LDLR expression on the liver cell surface by 84% at the concentration of 10 μM.