1H‐NMR metabolomic profiling of the crayfish Astacus leptodactylus subjected to polyphenol‐enriched diets

1H-NMR analysis of the hepatopancreas, muscle and haemolymph of Astacus leptodactylus after feeding with polyphenol-enriched diet is reported. 1H-NMR spectra of lipophilic extracts showed the presence of cholesterol, fatty acid residues, phospholipids and triglycerides. 1H-NMR spectra of aqueous extracts identified 35 metabolites in the hepatopancreas, 31 in the muscle and 22 in the haemolymph. A total of 20 metabolites (amino acids and their derivatives) were present in the hepatopancreas, the muscle and the haemolymph. A total of 10 metabolites were present in both the hepatopancreas and the muscle (five amino acids, 2-hydroxybutyrate, choline, myo-inositol, glycogen and uracil). 2-Hydroxyisobutyrate and creatine were present in both the hepatopancreas and the haemolymph. Phosphorylethanolamine, phosphocholine and fumarate were present only in the hepatopancreas and isoleucine only in the muscle. Statistical analysis showed that the percentage of weight gain was statistically higher in polyphenol-enriched diet groups compared to the control and that polyphenols had a stimulating effect on the general metabolism. No stress-related metabolites were higher in crayfish fed with polyphenol-enriched diet. Conversely, phosphatidylcholine, cholesterol and DHA, linked to resistance to environmental stress and diseases, were higher compared to the control diet. This study indicates that 1H-NMR is a useful tool to study the metabolomics in relation to diet differences

rBPI21 interacts with negative membranes endothermically promoting the formation of rigid multilamellar structures

© 2013 Elsevier B.V. All rights reservedrBPI21 belongs to the antimicrobial peptide and protein (AMP) family. It has high affinity for lipopolysaccharide (LPS), acting mainly against Gram-negative bacteria. This work intends to elucidate the mechanism of action of rBPI21 at the membrane level. Using isothermal titration calorimetry, we observed that rBPI21 interaction occurs only with negatively charged membranes (mimicking bacterial membranes) and is entropically driven. Differential scanning calorimetry shows that membrane interaction with rBPI21 is followed by an increase of rigidity on negatively charged membrane, which is corroborated by small angle X-ray scattering (SAXS). Additionally, SAXS data reveal that rBPI21 promotes the multilamellarization of negatively charged membranes. The results support the proposed model for rBPI21 action: first it may interact with LPS at the bacterial surface. This entropic interaction could cause the release of ions that maintain the packed structure of LPS, ensuring peptide penetration. Then, rBPI21 may interact with the negatively charged leaflets of the outer and inner membranes, promoting the interaction between the two bacterial membranes, ultimately leading to cell death.rBPI21 was a kind gift from XOMA, Ltd. (Berkeley, CA). This work was partially supported by Fundação para a Ciência e a Tecnologia – Ministério do Ensino e Ciência (FCT-MEC, Portugal), by the FP7-PEOPLEIRSES project MEMPEPACROSS (European Union), and by the Brazilian funding agencies Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq). MMD acknowledges FCT-MEC PhD fellowship SFRH/BD/41750/2007. RI and LRS Barbosa acknowledge CNPq for research fellowship. The authors are also in debt with Professors Francesco Spinozzi and Paolo Mariani (Università Politecnica delle Marche, Ancona, Italy), who developed and provided the GENFIT software