OBJECTIVE: Glycation is a common non-enzymatic reaction between proteins and sugars, resulting in the formation of advanced glycation end products (AGEs) in the human body. As can be seen in diabetic patients, the accumulation of AGEs in the skin has aesthetic consequences (wrinkles, brown spots and yellowish complexion). Therefore, the objective of this work was to find compounds isolated from natural sources that could eliminate the final AGEs accumulated in the skin with ageing. METHODS AND RESULTS: A preliminary screening performed on a bank of microbial extracts and pure compounds showed that 2,5-Diketopiperazines (DKPs), as well as the extract of Sphingobacterium sp (SNB-CN13), reduced the presence of AGEs in fibroblasts by À28% and À23%, respectively. In this article, we present the dereplication approach used to reveal the presence of 26 different DKPs in the crude extract of Sphingobacterium sp. Bioguided fractionation has led to the isolation of 12 of them, whose identity has been confirmed by HRMS and NMR. A green synthesis approach has been developed to synthesize 3 symmetrical DKPs. The biological activity of all DKPs was evaluated by the development of an in vitro test using immunocytochemistry to reveal the presence of AGE carboxymethyl-lysine in human dermal fibroblasts. CONCLUSION: Our work shows for the first time that DKPs decrease the amount of carboxymethyl-lysine AGE in elderly human dermal fibroblasts grown in vitro. Therefore, diketopiperazines can be considered as compounds of interest for dermatological and cosmetic applications with an anti-ageing aim. R esum e OBJECTIF: La glycation est une r eaction non enzymatique courante entre les prot eines et les sucres, qui entraîne la formation de produits finaux de glycation avanc ee (AGE) dans le corps humain. Comme on peut le voir chez les patients diab etiques, l'accumulation d'AGE dans la peau a des cons equences esth etiques (rides, taches brunes, teint jaunâtre). Par cons equent, l'objectif de ce travail etait de trouver des compos es isol es de sources naturelles qui pourraient eliminer les AGE finaux accumul es dans la peau avec le vieillissement. M ETHODES & R ESULTATS: Un examen pr eliminaire effectu e sur une banque d'extraits microbiens et de compos es purs a montr e que les 2,5-dic etopip erazines (DKP), ainsi que l'extrait de Sphingobacterium sp. (SNB-CN13), r eduisaient la pr esence d'AGE dans les fibroblastes de-28% et-23% respectivement. Dans cet article, nous pr esentons l'approche de d er eplication utilis ee pour r ev eler la pr esence de 26 DKP diff erentes dans l'extrait brut de Sphingobacterium sp. Le fractionnement bioguid e a conduit a l'isolement de 12 d'entre elles, dont l'identit e a et e confirm ee par HRMS et RMN. Une approche de synth ese verte a et e d evelopp ee pour synth etiser 3 DKP sym etriques. L'activit e biologique de toutes les DKP a et e evalu ee par le d eveloppement d'un test in vitro utilisant l'immunocytochimie pour r ev eler la pr esence de carboxym ethyl-lysine AGE dans les fibroblastes dermiques...

Glycation is a common non-enzymatic reaction between proteins and sugars, which gives rise in the human body to the formation of advanced glycation end products (AGEs). These modifications impacts both extra and intracellular proteins, leading to cells and tissues dysfunctions. In the skin, accumulation of AGEs leads to aesthetic consequences, wrinkles, dark spots and yellowish skin tone, as it can be seen in diabetic patients. Consequently, there is a growing dermatological interest to find compounds able to eliminate AGEs accumulated in skin. In this context, a method has been developed to detect and quantify intracellular glycation in human dermal fibroblasts. After cultivation of fibroblasts, cell lysates were injected in an HPLC system coupled with a fluorescence detector in bypass mode. The system allows the simultaneous measurement of global AGEs and particular pentosidine amounts using two sets of wavelengths in a single run of one minute. The immunocytochemistry approach was used to valid the HPLC analysis data. The method developed was able to quantify changes in global AGEs and pentosidine content in cells in response to glyoxal treatment. Fibroblasts treated with 500 µM of glyoxal for 48 hours showed a significant 2.3-fold and 2.6-fold increase in the content of AGEs and pentosidine respectively compared to control cells. As an application, a screening of natural extracts have been done and the method allowed identifying extracts able to significantly reduce the amount of pentosidine in fibroblasts (-32 %). These extracts act as deglycation agents of interest in the field of dermatology and cosmetology.

D- and L-amino acids (arginine, lysine) and hydroxylated molecules (sorbitol, ethyl-ethanolamine), all of which increase the intestinal transfer of calcium, are phosphorylated by jejunal and ileal microvilli. All these molecules inhibit the endogenous phosphorylation of microvilli proteins. D- and L-valine, which are not phosphorylated in the same conditions, have no effect on the phosphorylation of microvilli proteins. These observations are in good agreement with the scheme previously proposed for the increase of calcium intestinal transfer by L-lysine: phosphorylation of L-lysine by mucous membrane may interfere with that of membrane proteins, the phosphorylation of which would lower the permeability to that cation. Autoradiographies of the electrophoretogram show that the principal phosphorylable microvilli proteins have the same electrophoretic properties as those of the dimer and the monomer of alkaline phosphatase from the same intestinal loci. The ability of the enzyme to be phosphorylated and its well-known transphosphorylating activity upon hydroxylated molecules suggest that it could, in different ways, affect intestinal transfer of calcium.

Summary. Relations between the phosphorylation of aminated molecules and their action on calcium absorption.Various amino acids and guanidines (L-lysine, L-aspartic and L-glutamic acids, creatine, taurocyamine and glycocyamine), studied in ileal ligated loops, increased intestinal calcium absorption in the rat. L-arginine was effective per os.Since these compounds are highly phosphorylable, a phosphorylation mechanism may be involved in the stimulation of calcium absorption. The phosphorylated derivative of creatine was detected in the ileal mucosa of rats receiving creatine in the ileal ligated loop.Modified amino acids, such as 5-methyl-L-glutamate, asparagine or glutamine, whose phosphorylable function was masked by a methyl or an amide radical, were not effective in enhancing calcium absorption.Assays in vitro showed that an ileal mucosa extract phosphorylated arginine, lysine, glycocyamine and taurocyamine in the presence of ATP, acting as a phosphate donor.Introduction.