Julia Gatto

Automating a 96-well microtiter plate assay for identification of AGEs inhibitors or inducers: application to the screening of a small natural compounds library

Advanced glycation end-products (AGEs) are involved in the pathogenesis of numerous affections such as diabetes and neurological diseases. AGEs are also implied in various changes in tissues and organs. Therefore, compounds able to break them or inhibit their formation may be considered as potential drugs, dietary supplements, or bioactive additives. In this study, we have developed a rapid and reliable (Z′ factor calculation) anti-AGEs activity screening based on the overall fluorescence of AGEs. This method was successfully evaluated on known AGEs inhibitors and on a small library of natural compounds, yielding coherent results when compared with literature data.

Chemical Composition, Antioxidant and Anti-AGEs Activities of a French Poplar Type Propolis

Accumulation in tissues and serum of advanced glycation end-products (AGEs) plays an important role in pathologies such as Alzheimers disease or, in the event of complications of diabetes, atherosclerosis or renal failure. Therefore, there is a potential therapeutic interest in compounds able to lower intra and extracellular levels of AGEs. Among them, natural antioxidants (AO) with true anti-AGEs capabilities would represent good candidates for development. The purpose of this study was to evaluate the AO and anti-AGEs potential of a propolis batch and then to identify the main compounds responsible for these effects. In vivo, protein glycation and oxidative stress are closely related. Thus, AO and antiglycation activities were evaluated using both DPPH and ORAC assays, respectively, as well as a newly developed automated anti-AGEs test. Several propolis extracts exhibited very good AO and anti-AGEs activities, and a bioguided fractionation allowed us to identify pinobanksin-3-acetate as the most active component.

Comparison of two methods, UHPLC-UV and UHPLC-MS/MS, for the quantification of polyphenols in cider apple juices.

The aim of this study was to develop faster and more efficient phenotyping methods for in-depth genetic studies on cider apple progeny. The UHPLC chromatographic system was chosen to separate polyphenolic compounds, and quantifications were then simultaneously performed with a UV-PDA detector and an ESI-triple quadrupole mass analyzer (SRM mode). Both quantification methods were validated for 15 major compounds using two apple juice samples, on the basis of linearity, limits of detection and quantification, recovery and precision tests. The comparison between UV and SRM quantifications in 120 different samples of a cider apple progeny showed an excellent correlation for major compounds quantified with both methods. However, an overestimation was revealed for five compounds with the UV detector and the mass analyzer. Co-elution and matrix effects are discussed to explain this phenomenon. SRM methods should therefore be considered with restrictions in some cases for quantification measurements when several phenolic compounds are simultaneously quantified in complex matrices such as apple juices. For both methods, analyses were carried out over short periods of time while maintaining a high quality for the simultaneous quantification of phenolic compounds in apple juice. Each method is relevant for more in-depth genetic studies of the polyphenol content of apple juice.

A tocotrienol series with an oxidative terminal prenyl unit from Garcinia amplexicaulis

Ten tocotrienol derivatives, i.e., amplexichromanols (1-10), were isolated from stem bark of Garcinia amplexicaulis Vieill. ex Pierre collected in Caledonia. The structures of the compounds 1-5 were determined to be chromanol derivatives substituted by a polyprenyl chain oxidized in terminal position. The remaining compounds 6-10 are the corresponding dimeric derivatives. Eleven known compounds, including xanthones, tocotrienol derivatives, triterpenes and phenolic compounds, were also isolated. Their structures were mainly determined using one and two-dimensional NMR and mass spectroscopy analysis. The compounds and some amplexichromanol molecules formerly isolated from G. amplexicaulis exhibited significant antioxidant activity against lipid peroxidation and in the ORAC assay.

Partial Resistance of Carrot to Alternaria dauci Correlates with In Vitro Cultured Carrot Cell Resistance to Fungal Exudates

Although different mechanisms have been proposed in the recent years, plant pathogen partial resistance is still poorly understood. Components of the chemical warfare, including the production of plant defense compounds and plant resistance to pathogen-produced toxins, are likely to play a role. Toxins are indeed recognized as important determinants of pathogenicity in necrotrophic fungi. Partial resistance based on quantitative resistance loci and linked to a pathogen-produced toxin has never been fully described. We tested this hypothesis using the Alternaria dauci – carrot pathosystem. Alternaria dauci, causing carrot leaf blight, is a necrotrophic fungus known to produce zinniol, a compound described as a non-host selective toxin. Embryogenic cellular cultures from carrot genotypes varying in resistance against A. dauci were confronted with zinniol at different concentrations or to fungal exudates (raw, organic or aqueous extracts). The plant response was analyzed through the measurement of cytoplasmic esterase activity, as a marker of cell viability, and the differentiation of somatic embryos in cellular cultures. A differential response to toxicity was demonstrated between susceptible and partially resistant genotypes, with a good correlation noted between the resistance to the fungus at the whole plant level and resistance at the cellular level to fungal exudates from raw and organic extracts. No toxic reaction of embryogenic cultures was observed after treatment with the aqueous extract or zinniol used at physiological concentration. Moreover, we did not detect zinniol in toxic fungal extracts by UHPLC analysis. These results suggest that strong phytotoxic compounds are present in the organic extract and remain to be characterized. Our results clearly show that carrot tolerance to A. dauci toxins is one component of its partial resistance.

Polyphenolic Compounds with Anti -Ages Activity from Three Clusiaceae Plants

Aim: This study focused on finding molecules with inhibitory effects on Advanced Glycation End-products (AGEs) formation from Tanzanian some Clusiaceae plant species Study Design: Field study and Laboratory experimental tests. Place and Duration of Study: Institute of Traditional Medicine, Muhimbili University of Health and Allied Sciences, P.O. Box 65001, Dar es Salaam, Tanzania and PRES LUNAM, Universite d’Angers, EA 921 SONAS, 16 Bd Daviers, 49045 Angers, France, between June 2011 and July 2013. Methodology: Three Clusiaceae plant species (Garcinia semseii, G. volkensii and Allanblackia ulugurensis) were collected and dried in the field with the assistance of a botanist. Extraction and concentration of plant samples to obtain crude extracts were done in the laboratory following standard procedures. The isolation of the phenolic compounds was carried out by using normal phase column chromatography as well as HighOriginal Research Article European Journal of Medicinal Plants, 4(11): 1336-1344, 2014 1337 performance liquid chromatography (HPLC). The isolated compounds were tested for antiAGE activity using the in vitro automated assay. Results: Two polyphenolic compounds exhibiting phloroglucinol moieties [e.g. polyprenylated benzophenones, such as guttiferone F, 2 (18 mg)] or biflavonoids [such as morelloflavone, 1 (22mg)] were isolated and identified from A. ulugurensis and G. volkensis respectively. The results further indicated that compound 1 is an excellent inhibitor of AGE formation exhibiting an IC50 values of 78 and 64 μM at wavelength of 370/440 (vesperlysines-like AGEs) and 335/385 (pentosidine-like AGEs) respectively. Conclusion: Plants belonging to the Clusiaceae family commonly used in Tanzanian traditional medicine need to be considered as a potential source of molecules exhibiting pharmacological activities such as anti-AGE activity. Morelloflavone (1) and other biflavonoids prove to be very good anti-AGE compounds using our automated screening assay. Hence, our automated in vitro assay allows a fast, effective and quite inexpensive screening of natural compounds and can therefore be applied to high throughput screening projects.