Effect of Ionization Degree of Poly(amidoamine) Dendrimer and 5-Fluorouracil on the Efficiency of Complex Formation-A Theoretical and Experimental Approach.

Due to their unique structure, poly(amidoamine) (PAMAM) dendrimers can bind active ingredients in two ways: inside the structure or on their surface. The location of drug molecules significantly impacts the kinetics of active substance release and the mechanism of internalization into the cell. This study focuses on the effect of the protonation degree of the G4PAMAM dendrimer and the anticancer drug 5-fluorouracil (5FU) on the efficiency of complex formation. The most favorable conditions for constructing the G4PAMAM-5FU complex are a low degree of protonation of the dendrimer molecule with the drug simultaneously present in a deprotonated form. The fluorine components in the XPS spectra confirm the formation of the stable complex. Through SAXS and DLS methods, a decrease in the dendrimer's molecular size resulting from protonation changes at alkaline conditions was demonstrated. The gradual closure of the dendrimer structure observed at high pH values makes it difficult for the 5FU molecules to migrate to the interior of the support structure, thereby promoting drug immobilization on the surface. The 1H NMR and DOSY spectra indicate that electrostatic interactions determine the complex formation process. Through MD simulations, the localization profile and the number of 5FU molecules forming the complex were visualized on an atomic scale.

Waste salt from the manufacturing process of mullet bottarga as source of oil with nutritional and nutraceutical properties.

BACKGROUND: The Sardinian food delicacy 'bottarga' is the final product of a number of treatments (salting and drying) on the ovaries of mullet (Mugil spp) and represents an important natural source of n-3 polyunsaturated fatty acids (n-3 PUFA) with nutraceutical properties. During the salting process of mullet roes to obtain bottarga, huge amounts of waste salt are generated, rich in residual ovary material. RESULTS: We evaluated the lipid composition (main lipid components and fatty acids) and bioactivity of oil obtained from the ovary material separated from waste salt (waste salt oil). Oil was obtained by supercritical fluid extraction with carbon dioxide (SFE-CO2 ), an environmentally friendly separation technique. The lipid composition of waste salt oil was determined by carbon-13 nuclear magnetic resonance (13 C-NMR) spectroscopy and reversed-phase high-performance liquid chromatography with diode array detector and an evaporative light scattering detector (HPLC-DAD/ELSD) chromatography. The oil was characterized by a relatively high level of n-3 PUFA (122 ± 7 g kg-1 of oil), and these beneficial health compounds were mainly present in the form of wax esters. Waste salt oil showed a marked cytotoxic effect [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay] in cancer B16F10 melanoma cells, with a slight cytotoxic effect in normal cells (3T3 fibroblasts). Waste salt and its derivatives (salt oil and residual material after oil extraction) were also tested for the attractant effect and acceptability to insects (Ceratitis capitata) to gain preliminary information about their potential application for animal supplementation. CONCLUSION: The results qualify waste salt as a potential resource for veterinary dietary supplements, nutraceuticals, and pharmaceutical applications. © 2020 Society of Chemical Industry.

Nutrimetabolomics and Adipocitokines in the "Great Obstetrical Syndromes".

In neonatal medicine, nutritional research is focusing more and more on thrifty phenotype effects, in order to understand and prevent the development of long-term diseases. lschemic placental disease which brings together Gestational diabete, Preeclamptic Toxemia, and Intrauterine Growth Restriction, the "Great Obstetrical Syndromes" (GOS), originates from sugar.and lipid metabolism. Adipokines and metabolomics can be valuable tools for the diagnosis of obstetrical syndromes and addressing nutrition. Inappropriate nutrition, even in the first periods of life, can accelerate the development of chronic metabolic diseases, especially in the pediatric age. The purpose of this review is firstly to critically examine the information provided by the studies of metabolomics on GOS's and better understand their origin. Secondly, it reflects on the IUGR metabolism and on applications of metabolomics in nutrition and its "nutrimetabolomic" effects and then to discuss the principles that guide nutrition of IUGR children in the light of these.

Conformational isomerisms and nano-aggregation in substituted alkylammonium nitrates ionic liquids: an x-ray and computational study of 2-methoxyethylammonium nitrate.

In this study, we discuss, using molecular dynamics simulations and energy-dispersive x-ray diffraction data, how a conformational isomerism can dramatically alter the nanosegregation phenomena that take place in a prototypical ionic liquid. The diffraction patterns of liquid 2-methoxyethylammonium nitrate are compared with the results from molecular dynamics simulations. The simulations conditions and force field parameters have been varied producing different charge models and different populations of conformers of the cation. We show that, while the short range structure is relatively unchanged in the models, the long range aggregation phenomena deemed responsible for the appearance of low Q peaks in the X-ray patterns strongly depend on the choice of the charge model. In the title compound, the best agreement with the experiment, where no low Q peaks appear, occurs if the point charges are calculated using the gauche conformation of the cation, which is characterized by an intramolecular hydrogen bond between ammonium and ether groups.

The melting curves of calf thymus-DNA are buffer specific.

The specific effects of salts (strong electrolytes) on biomolecular properties have been investigated for more than a century. By contrast, the specific role of pH buffers (weak electrolytes and their salts) has usually been ignored. Here, specific buffer effects on DNA thermal stability were evaluated by measuring the melting curve of calf thymus DNA through UV-vis spectroscopy. The study was carried out using phosphate, Tris, citrate and cacodylate buffers at fixed pH 7.4 at concentrations varying systematically in the range 1-600 mM. DNA stability increases with buffer concentration and is influenced specifically by buffer type. To interpret empirical data, a theoretical model was applied with parameters quantifying the impact of buffer on the DNA backbone charge. Comparing the buffer effects via buffer ionic strength rather than buffer concentration, we find that the buffers stabilize DNA in the order Tris > cacodylate > phosphate > citrate.

An energy dispersive x-ray scattering and molecular dynamics study of liquid dimethyl carbonate.

In this work, we report on the first x-ray diffraction study on liquid dimethyl carbonate. Diffraction spectra were collected with an energy-dispersive instrument, whose wide Q-range allows the structure determination of weakly ordered systems (such as liquids). The structural correlation in this liquid ranges up to about 20 A. The observed patterns are interpreted with a structural model derived from classical molecular dynamics simulations. The simulations were run using OPLS force field, only slightly modified to restrain bond distances to the experimental values. The model structure function and radial distribution functions, averaged among the productive trajectory frames, are in very good agreement with the corresponding experimental ones. Molecular dynamics results show that the deviations from C(2v) cis-cis structure, predicted by ab initio calculations and observed by electron diffraction in the gas phase, are small. By analyzing the intra- and intermolecular pair distribution functions, it was possible to assign the peaks of the experimental radial distribution function to specific structural correlations, and to compute the different average intermolecular coordination numbers. The intermolecular methyl-carbonyl oxygen distance is thoroughly discussed to assess the presence of weak C-H...O hydrogen bonds.

Wheat bran biodegradation by Pleurotus ostreatus: a solid-state carbon-13 NMR study.

Solid-state (13)C nuclear magnetic resonance (NMR) and elemental analysis techniques were used to monitor the degradation of wheat bran by the white-rot fungus Pleurotus ostreatus during a 62-day cultivation period. The weight loss and in vitro organic matter digestibility of the substrate were also evaluated after fungal treatment. The (13)C NMR spectra of degraded wheat bran samples showed a lower content in carbohydrates and a higher content in aliphatic and carboxylic groups than the untreated control sample. In parallel, changes in the wheat bran elemental composition evidenced a decrease in carbon content and a concomitant increase in nitrogen and oxygen content during mycelium growth. These results clearly indicate the occurrence of progressive changes in the composition of wheat bran during fungal treatment and are interpreted in terms of preferential degradation of amorphous vs. crystalline polysaccharides by the fungal mycelium and accumulation of proteins in the substrate. At the end of the cultivation period, the treated samples experienced an average weight loss of 20% and an increase in organic matter digestibility of 17%.

Effect of rubidium and cesium ions on the dimeric quaduplex formed by the Oxytricha nova telomeric repeat oligonucleotide d(GGGGTTTTGGGG).

The DNA sequence d(GGGGTTTTGGGG) consists of 1.5 units of the repeat in telomeres of Oxytricha nova. It has been shown by NMR and x-ray crystallographic analysis that it is capable to form a dimeric quadruplex structure and that a variety of cations, namely K(+), Na(+), and NH(4)(+), are able to interact with this complex with different affinity, leading to complexes characterized by different local conformations. Thus, in order to improve the knowledge of this kind of molecule, and in particular to provide further insight into the role of monovalent cations in the G-quadruplex folding and conformation, we have investigated by (1)H-NMR the effect of the addition of Rb(+) and Cs(+) to the quadruplex formed by the oligonucleotide d(GGGGTTTTGGGG).

(1)H NMR-based urine metabolic profile of IUGR, LGA, and AGA newborns in the first week of life.

Under conditions of non-optimal supply of nutrients, maternal diet during gestation can alter the balance between anabolic and catabolic pathways of fetus and triggers an effect of programming to the metabolic syndrome. Metabolomics is an analytical technique that has been recently attracting increasing interest for the identification of biomarkers of dietary exposure. In this study, a NMR-based metabolomic approach was employed for an explorative analysis of the time-related urinary metabolic profiles of three groups of newborns receiving a different fetal nutrition: adequate for gestational age (AGA), with intrauterine growth retardation (IUGR), and large for gestational age (LGA). Urine samples were collected over the first week of life. Application of Orthogonal Partial Least Squares Discriminant Analysis (OPLS-DA) evidenced similar time-related modifications in the metabolic profiles of the three classes of infants, consisting mainly of changes in levels of taurine, creatinine, betaine, and glycine. Furthermore, alterations in the content of citrate and myo-inositol were found to be characteristic of IUGR and LGA, whole levels were higher with respect to controls, while higher contents of betaine and succinate were noted in AGA. Our results positively support the application of the metabolomic approach in the study of the metabolic pathways associated to fetal malnutrition.

Analysing the effects of frozen storage and processing on the metabolite profile of raw mullet roes using ¹H NMR spectroscopy.

(1)H NMR spectroscopy was used to investigate changes in the low molecular weight metabolic profile of raw mullet (Mugil spp.) roes during frozen storage and upon processing. NMR data were analysed by Principal Component Analyses (PCA). In the model constructed using frozen roes, no statistical significant metabolic modifications were observed in the first six months of storage, while choline derivatives, dimethylamine, lactate, and most of the free amino acids were identified as changing with statistical significance (p<0.05) in response to frozen storage time of twelve months. The PCA model comparing the metabolic profiles of roes before and after processing showed that the major modifications occurring upon manufacturing were the increase of the choline derivative compounds, uracil, and free amino acids, and a large decrease of taurine, glucose, lactate, and creatine/phosphocreatine. All of the above mentioned modifications reflect the occurrence of chemical/biochemical reactions arising from degradation processes such as lipolysis and proteolysis.

Investigation of the ¹H-NMR based urine metabolomic profiles of IUGR, LGA and AGA newborns on the first day of life.

(1)H-NMR spectroscopy coupled with multivariate statistical analysis was used for the first time to compare the urinary NMR metabolic profiles of neonates with intrauterine growth retardation (IUGR) and large for gestational age (LGA). For the sake of comparison, infants who were adequate for gestational age (AGA) were also analyzed. Pattern recognition methods, including Principal Component Analyses (PCA), Partial Least Squares Discriminant Analysis (PLS-DA) and Orthogonal Partial Least Squares Discriminant Analysis (OPLS-DA), were used to analyze NMR data. Clear differences among the metabolic profiles of AGA, IUGR and LGA were observed. The main metabolites responsible for these differentiations were identified as myo-inositol, creatinine, creatine, citrate, urea and glycine. In particular, among these, myo-inositol may be a potential biomarker of an altered glucose metabolism during fetal development both in IUGR and LGA. This study highlights the applicability of NMR-based metabolomics for improving the understanding of the relations among nutrition, integrated metabolism and health in neonatology.

A metabolomic study of preterm human and formula milk by high resolution NMR and GC/MS analysis: preliminary results.

OBJECTIVE: The aim of the present study was to investigate the metabolic profile of preterm human breast milk (HBM) by using a metabolomic approach. METHODS: NMR spectroscopy and GC/MS were used to analyze the water-soluble and lipid fractions extracted from milk samples obtained from mothers giving birth at 26-36 weeks of gestation. For the sake of comparison, preterm formula milk was also studied. RESULTS: The multivariate statistical analysis of the data evidenced biochemical variability both between preterm HBM and commercial milk and within the group of HBM samples. CONCLUSIONS: The preliminary results of this study suggest that metabolomics may provide a promising tool to study aspects related to the nutrition and health of preterm infant.

1H NMR-based metabolomic analysis of urine from preterm and term neonates.

Metabolomics is a technique used to non-invasively determine metabolic status of an organism. Aim of our study was to analyze urinary metabolic profiles in term and preterm infants in order to identify gestational age-related metabolic differences and to predict metabolic maturity at birth. Twenty-six healthy term infants and 41 preterm infants were prospectively enrolled. A urine sample was collected non-invasively within the first hours of life. Samples were analyzed by proton nuclear magnetic resonance (1H NMR) spectroscopy and NMR urine spectra were analyzed by multivariate statistical analysis. Distinct metabolic patterns were found between term infants and preterm infants, as well as between preterm infants of 23-32 weeks' gestation and those of 33-36 weeks' gestation. Individual metabolites discriminating between these groups were hippurate, tryptophan, phenylalanine, malate, tyrosine, hydroxybutyrate, N-acetyl-glutamate, and proline. Metabolomic analysis revealed distinct urinary metabolic profiles in neonates of different gestational ages, and identified the discriminating metabolites. This holistic approach appears to be a promising tool for investigating newborn metabolic maturation over time, and might lead to a tailored management of neonatal disorders.

Adaptative value of a PKC-PKI55 feedback loop of inhibition that prevents the kinase's deregulation.

A 168-bp amplification product was obtained in RT-PCR experiments using a degenerate oligonucleotide designed on a five-amino acid sequence of IN, a 7-kDa protein, previously characterized as a PKC inhibitor. It was included in the coding ORF of the 1530-bp-long IMAGE clone ID 38900 (accession numbers R51337 and R51448) that produces a translation product of 6.5 kDa. The translation of the ORF conceptual reading frame allowed the preparation of the synthetic protein PKI55, which was found to inhibit and degrade both untreated nPKC d isozymes and activated cPKC isozymes. The PKI55 gene is localized in chromosome 2q35. The Repeat Maskers output showed a 533-bp-long LTR32/ERVL segment that included the PKI55 coding sequence and a complete regulatory region. The coding sequence and the structure of PKI55 were detected in a brain cDNA of Macaca fascicularis (diverged from human lineages about 25 Myr ago). Three other human genes with over 60% identities with PKI55 were identified in three different loci (i.e., chromosomes 10, 15, and 20.) Synthesis of PKI55 was stimulated by PKC activation. A feedback loop of inhibition is established. When the PKCs are overactivated, PKI55 induces degradation of the enzyme and prevents the isozyme overexpression implicated in a number of important diseases including cancer, diabetes, and disorders of the immune system. The presence of the PKI55 sequence in Macaca fascicularis as well as in human chromosomes 10, 15, and 20 indicates a selective advantage for the PKI55 sequence and the adaptive value of the feedback mechanism.