Urine Metabolomic Signature of People Diagnosed with Balkan Endemic Nephropathy and Other Types of Chronic Kidney Disease Compared with Healthy Subjects in Romania.

Metabolomic analysis methods were employed to determine biomarkers for various chronic kidney diseases (CKDs). Modern analytical methods were developed and applied successfully to find a specific metabolomic profile in urine samples from CKD and Balkan endemic nephropathy (BEN) patients. The aim was to explore a specific metabolomic profile defined by feasible/easy-to-identify molecular markers. Urine samples were collected from patients with CKDs and BEN, and from healthy subjects from endemic and nonendemic areas in Romania. Metabolomic analysis of urine samples, extracted by the liquid-liquid extraction (LLE) method, was performed by gas chromatography-mass spectrometry (GC-MS). The statistical exploration of the results was performed through a principal component analysis (PCA) evaluation. Urine samples were statistically analyzed using a classification based on six types of metabolites. Most urinary metabolites are distributed in the center of a loading plot, meaning that these compounds do not represent significant markers for BEN. One of the most frequent and higher-concentration urinary metabolites in BEN patients was p-Cresol, a phenolic compound that implies a severe injury of the renal filtration function. The presence of p-Cresol was associated with protein-bound uremic toxins, which have specific functional groups such as indole and phenyl. In prospective studies for future investigation, prevention, and disease treatment, we suggest a larger sample size, sample extraction using other methods, and analysis using other chromatography techniques coupled with mass spectrometry, which can generate a more significant data set for statistical analysis.

Discrimination of ß-cyclodextrin/hazelnut (Corylus avellana L.) oil/flavonoid glycoside and flavonolignan ternary complexes by Fourier-transform infrared spectroscopy coupled with principal component analysis.

The goal of the study was the discrimination of ß-cyclodextrin (ß-CD)/hazelnut (Corylus avellana L.) oil/antioxidant ternary complexes through Fourier-transform infrared spectroscopy coupled with principal component analysis (FTIR-PCA). These innovative complexes combine the characteristics of the three components and improve the properties of the resulting material such as the onsite protection against oxidative degradation of hazelnut oil unsaturated fatty acid glycerides. Also, the apparent water solubility and bioaccessibility of the hazelnut oil components and antioxidants can be increased, as well as the controlled release of bioactive compounds (fatty acid glycerides and antioxidant flavonoids, namely hesperidin, naringin, rutin, and silymarin). The appropriate method for obtaining the ternary complexes was kneading the components at various molar ratios (1:1:1 and 3:1:1 for ß-CD hydrate:hazelnut oil (average molar mass of 900 g/mol):flavonoid). The recovering yields of the ternary complexes were in the range of 51.5-85.3% and were generally higher for the 3:1:1 samples. The thermal stability was evaluated by thermogravimetry and differential scanning calorimetry. Discrimination of the ternary complexes was easily performed through the FTIR-PCA coupled method, especially based on the stretching vibrations of CO groups in flavonoids and/or CO/CC groups in the ternary complexes at 1014.6 (± 3.8) and 1023.2 (± 1.1) cm-1 along the second PCA component (PC2), respectively. The wavenumbers were more appropriate for discrimination than the corresponding intensities of the specific FTIR bands. On the other hand, ternary complexes were clearly distinguishable from the starting ß-CD hydrate along the first component (PC1) by all FTIR band intensities and along PC2 by the wavenumber of the asymmetric stretching vibrations of the CH groups at 2922.9 (± 0.4) cm-1 for ternary complexes and 2924.8 (± 1.4) cm-1 for ß-CD hydrate. The first two PCA components explain 70.38% from the variance of the FTIR data (from a total number of 26 variables). Other valuable classifications were obtained for the antioxidant flavonoids, with a high similarity for hesperidin and naringin, according to FTIR-PCA, as well as for ternary complexes depending on molar ratios. The FTIR-PCA coupled technique is a fast, nondestructive and cheap method for the evaluation of quality and similarity/characteristics of these new types of cyclodextrin-based ternary complexes having enhanced properties and stability.

Structure-property relationships on recrystallized ß-cyclodextrin solvates: A focus on X-ray diffractometry, FTIR and thermal analyses.

The goal of the study was to evaluate the influence of the solvent properties on the crystal characteristics of ß-cyclodextrin (ß-CD) recrystallized from alcohol-water solvent mixtures, with possible applications for the preparation, purifying and complexation of ß-CD. For the first time, structure-property relationships (QSPRs) between the hydrophobicity of alcohols or dielectric constant of solvents used for recrystallization of ß-CD and its properties (such as crystallinity index, CI) have been obtained. Recrystallized ß-CD from water and C1-C4 alcohol-water solutions provide ß-CD with higher CI values of 99.4(±5.9)% for ethanol-water (1:4, v/v) as recrystallizing system. This property has a parabolic variation with the logP (octanol/water partition coefficient) of the alcohol (r2 = 0.998). Solvent parameters also influence the ß-CD crystal characteristics, as was demonstrated by X-ray diffractometry refinement, infrared spectroscopy and thermal analyses.

Fatty Acid Profile of Lipid Fractions of Mangalitza (Sus scrofa domesticus) from Northern Romania: A GC-MS-PCA Approach.

Mangalitza pig (Sus scrofa domesticus) becomes more popular in European countries. The goal of this study was to evaluate the fatty acid profile of the raw and thermally processed Mangalitza hard fat from Northern Romania. For the first time, the gas chromatography-mass spectrometry-Principal component analysis technique (GC-MS-PCA)-was applied to evaluate the dissimilarity of Mangalitza lipid fractions. Three specific layers of the hard fat of Mangalitza from Northern Romania were subjected to thermal treatment at 130 °C for 30 min. Derivatized samples were analyzed by GC-MS. The highest relative content was obtained for oleic acid (methyl ester) in all hard fat layers (36.1-42.4%), while palmitic acid was found at a half (21.3-24.1%). Vaccenic or elaidic acids (trans) were found at important concentrations of 0.3-4.1% and confirmed by Fourier-transform infrared spectroscopy. These concentrations are consistently higher in thermally processed top and middle lipid layers, even at double values. The GC-MS-PCA coupled technique allows us to classify the unprocessed and processed Mangalitza hard fat specific layers, especially through the relative concentrations of vaccenic/elaidic, palmitic, and stearic acids. Further studies are needed in order to evaluate the level of degradation of various animal fats by the GC-MS-PCA technique.

Complexation of Danube common nase (Chondrostoma nasus L.) oil by ß-cyclodextrin and 2-hydroxypropyl-ß-cyclodextrin.

ß-Cyclodextrin- and 2-hydroxypropyl-ß-cyclodextrin/Danube common nase (Chondrostoma nasus L.) oil complexes (ß-CD- and HP-ß-CD/CNO) have been obtained for the first time. The fatty acid (FA) profile of the CNO indicates an important content of polyunsaturated fatty acids, the most important being eicosapentaenoic acid (EPA, 6.3%) and docosahexaenoic acid (DHA, 1.6%), both ω-3 FAs. The complexes have been obtained by kneading method. The moisture content and successful of molecular encapsulation have been evaluated by thermal and spectroscopic techniques. Thermogravimetry and differential scanning calorimetry analyses reveals that the moisture content of CD/CNO complexes significantly decreased, compared to starting CDs. On the other hand, the crystallinity index was for the first time determined for such type of complexes, the ß-CD/CNO complex having values of 43.9(±18.3)%, according to X-ray diffractometry. FA profile and CD/CNO characteristics sustain the use of these ω-3 based complexes for food supplements or functional food products, but further studies are needed.

Fatty acid profile of Romanian's common bean (Phaseolus vulgaris L.) lipid fractions and their complexation ability by ß-cyclodextrin.

The goal of the present study was the evaluation of the fatty acid (FA) profile of lipid fraction from dry common beans (Phaseolus vulgaris L.) (CBO) harvested from North-East (NE) and South-West (SW) of Romania and to protect against thermal and oxidative degradation of the contained omega-3 and omega-6 polyunsaturated fatty acid (PUFA) glycerides by ß-cyclodextrin (ß-CD) nanoencapsulation, using kneading method. The most abundant FAs in the CBO samples were PUFAs, according to gas chromatography-mass spectrometry (GC-MS) analysis. Linoleic acid (methyl ester) was the main constituent, having relative concentrations of 43.4 (±1.95) % and 35.23 (±0.68) % for the lipid fractions separated from the common beans harvested from the NE and SW of Romania, respectively. Higher relative concentrations were obtained for the omega-3 α-linolenic acid methyl ester at values of 13.13 (±0.59) % and 15.72 (±0.30) % for NE and SW Romanian samples, respectively. The omega-3/omega-6 ratio consistently exceeds the lower limit value of 0.2, from where the PUFA glyceride mixture is valuable for the human health. This value was 0.32 (±0.02) for the NE samples and significantly higher for the CBO-SW samples, 0.51 (±0.01). These highly hydrophobic mixtures especially consisting of PUFA triglycerides provide ß-CD complexes having higher thermal and oxidative stability. Kneading method allowed obtaining ß-CD/CBO powder-like complexes with higher recovery yields of >70%. Thermal analyses of complexes revealed a lower content of hydration water (3.3-5.8% up to 110°C in thermogravimetry (TG) analysis and 154-347 J/g endothermal effect in differential scanning calorimetry (DSC) analysis) in comparison with the ß-CD hydrate (12.1% and 479.5-480 J/g, respectively). These findings support the molecular inclusion process of FA moieties into the ß-CD cavity. Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) analysis reveals the formation of the ß-CD/CBO inclusion complexes by restricting the vibration and bending of some bonds from the host and guest molecules. Moreover, powder X-ray diffractometry (PXRD) analysis confirm the formation of the host-guest complexes by modifying the diffractograms for ß-CD/CBO complexes in comparison with the ß-CD and ß-CD + CBO physical mixtures. A significant reduction of the level of crystallinity from 93.3 (±5.3) % for ß-CD to 60-60.9% for the corresponding ß-CD/CBO complexes have been determined. The encapsulation efficiency (EE), the profile of FAs, as well as the controlled release of the encapsulated oil have also been evaluated. The EE was >40% in all cases, the highest value being obtained for ß-CD/CBO-SW complex. The SFA content increased, while the unsaturated FA glycerides had lower relative concentrations in the encapsulated CBO samples. It can be emphasized that the main omega-3 FA (namely α-linolenic acid glycerides) had close concentrations in the encapsulated and raw CBOs (13.13 (±0.59) % and 14.04 (±1.54) % for non-encapsulated and encapsulated CBO-NE samples, 15.72 (±0.30) % and 12.41 (±1.95) % for the corresponding CBO-SW samples, respectively). The overall unsaturated FA content significantly decreased after complexation (from 19.03-19.16% for the raw CBOs to 17.3-17.7% for encapsulated oils in the case of MUFAs, and from 55.7-58.8% to 35.13-43.36% for PUFAs). On the other hand, the omega-3/omega-6 ratio increased by ß-CD nanoencapsulation to 0.51 (±0.07) and 0.76 (0.26) for ß-CD/CBO-NE and ß-CD/CBO-SW complexes, respectively. As a conclusion, the lipid fractions of the Romanian common beans are good candidates for ß-CD complexation and they can be protected against thermal and oxidative degradation in common beans based food products such as functional foods or food supplements using natural CDs.

Moisture evaluation of ß-cyclodextrin/fish oils complexes by thermal analyses: A data review on common barbel (Barbus barbus L.), Pontic shad (Alosa immaculata Bennett), European wels catfish (Silurus glanis L.), and common bleak (Alburnus alburnus L.) living in Danube river.

The moisture content of ß-cyclodextrin/Danube fish oils complexes (common barbel, Pontic shad, European wels catfish, common bleak) was evaluated by thermal methods. Saturated and monounsaturated fatty acids were the most concentrated in fish oils (25.3-30.8% and 36.1-45.0%). ω-3 And ω-6 fatty acids were identified in low concentrations of 2.8-12.1% and 4.1-7.1%. The moisture content was significantly lowered after ß-CD complexation, as revealed by thermogravimetric (TG) analysis (13.3% for ß-CD, 2.5-6.5% for complexes). These results are consistent with the differential scanning calorimetry (DSC) data for the peaks corresponding to dissociation of water (calorimetric effect of 536Jg-1 for ß-cyclodextrin and 304-422.5Jg-1 for complexes). Furthermore, both TG and DSC results support the formation of inclusion complexes. This is the first study on the nanoencapsulation of Danube fish oils in ß-cyclodextrin.

Thermal and oxidative stability of Atlantic salmon oil (Salmo salar L.) and complexation with ß-cyclodextrin.

The thermal and oxidative stability of Atlantic salmon oil (Salmo salar L.) as well as its ß-cyclodextrin (ß-CD) complexation ability has been verified for the first time. The main omega-3 fatty acids, EPA and DHA, were significantly degraded, even at 50 °C. Their relative concentrations decrease from 6.1% for EPA and 4.1% for DHA to 1.7% and 1.5% after degradation at 150 °C, respectively. On the other hand, the relative concentrations of monounsaturated and saturated fatty acids remained constant or slightly increased by a few percent after degradation (e.g., from 10.7% to 12.9% for palmitic acid). Co-crystallization of ASO with ß-CD at a host-guest ratio of 1:1 and 3:1 from an ethanol-water mixture and kneading methods has been used for the preparation of ß-CD/ASO complexes. The analysis of the complexes by thermogravimetry, differential scanning calorimetry (DSC), and Karl Fischer titration (KFT) as well as the decrease of the "strongly-retained" water content confirm the formation of the inclusion compound. Furthermore, the DSC parameters correlate well with the KFT kinetic data for ß-CD/ASO complexes.

Differentiation of rye and wheat flour as well as mixtures by using the kinetics of Karl Fischer water titration.

The aim of this study was to find a simple way to differentiate between rye and wheat flour and their mixtures by using the kinetic parameters of Karl Fischer water titration (KFT). Consequently, the water content and type of molecules in rye and wheat mixtures used in Romanian bread making have been investigated by means of volumetric KFT. Further, the kinetics of KFT have been determined and novel kinetic parameters corresponding to "surface" and "strongly-retained" water molecules have been identified to discriminate between rye and wheat flour and their mixtures. The "surface" and "strongly-retained" water reaction rates well correlate with the rye content in the flour mixtures, especially at higher temperature analysis (r>0.95). These parameters can be used as indicators for quality evaluation of such type of mixtures, as well as to identify adulteration by improper use of the rye-wheat flour ratios in bread making.

Thermal and oxidative stability of the Ocimum basilicum L. essential oil/ß-cyclodextrin supramolecular system.

Ocimum basilicum L. essential oil and its ß-cyclodextrin (ß-CD) complex have been investigated with respect to their stability against the degradative action of air/oxygen and temperature. This supramolecular system was obtained by a crystallization method in order to achieve the equilibrium of complexed-uncomplexed volatile compounds in an ethanol/water solution at 50 °C. Both the raw essential oil and its ß-CD complex have been subjected to thermal and oxidative degradation conditions in order to evaluate the protective capacity of ß-CD. The relative concentration of the O. basilicum L. essential oil compounds, as determined by GC-MS, varies accordingly with their sensitivity to the thermal and/or oxidative degradation conditions imposed. Furthermore, the relative concentration of the volatile O. basilicum L. compounds found in the ß-CD complex is quite different in comparison with the raw material. An increase of the relative concentration of linalool oxide from 0.3% to 1.1%, in addition to many sesquiterpene oxides, has been observed. ß-CD complexation of the O. basilicum essential oil modifies the relative concentration of the encapsulated volatile compounds. Thus, linalool was better encapsulated in ß-CD, while methylchavicol (estragole) was encapsulated in ß-CD at a concentration close to that of the raw essential oil. Higher relative concentrations from the degradation of the oxygenated compounds such as linalool oxide and aromadendren oxide were determined in the raw O. basilicum L. essential oil in comparison with the corresponding ß-CD complex. For the first time, the protective capability of natural ß-CD for labile basil essential oil compounds has been demonstrated.

Titanocene / cyclodextrin supramolecular systems: a theoretical approach.

BACKGROUND: Recently, various metallocenes were synthesized and analyzed by biological activity point of view (such as antiproliferative properties): ruthenocenes, cobaltoceniums, titanocenes, zirconocenes, vanadocenes, niobocenes, molibdocenes etc. Two main disadvantages of metallocenes are the poor hydrosolubility and the hydrolytic instability. These problems could be resolved in two ways: synthetically modifying the structure or finding new formulations with enhanced properties. The aqueous solubility of metallocenes with cytostatic activities could be enhanced by molecular encapsulation in cyclodextrins, as well as the hydrolytic instability of these compounds could be reduced. RESULTS: This study presents a theoretical approach on the nanoencapsulation of a series of titanocenes with cytotoxic activity in α-, ß-, and γ-cyclodextrin. The HyperChem 5.11 package was used for building and molecular modelling of titanocene and cyclodextrin structures, as well as for titanocene/cyclodextrin complex optimization. For titanocene/cyclodextrin complex optimization experiments, the titanocene and cyclodextrin structures in minimal energy conformations were set up at various distances and positions between molecules (molecular mechanics functionality, MM+). The best interaction between titanocene structures and cyclodextrins was obtained in the case of ß- and γ-cyclodextrin, having the hydrophobic moieties oriented to the secondary face of cyclodextrin. The hydrophobicity of titanocenes (logP) correlate with the titanocene-cyclodextrin interaction parameters, especially with the titanocene-cyclodextrin interaction energy; the compatible geometry and the interaction energy denote that the titanocene/ß- and γ-cyclodextrin complex can be achieved. Valuable quantitative structure-activity relationships (QSARs) were also obtained in the titanocene class by using the same logP as the main parameter for the in vitro cytotoxic activity against HeLa, K562, and Fem-x cell lines. CONCLUSIONS: According to our theoretical study, the titanocene/cyclodextrin inclusion compounds can be obtained (high interaction energy; the encapsulation is energetically favourable). Further, the most hydrophobic compounds are better encapsulated in ß- and γ-cyclodextrin molecules and are more stable (from energetically point of view) in comparison with α-cyclodextrin case. This study suggests that the titanocene / ß- and γ-cyclodextrin complexes (or synthetically modified cyclodextrins with higher water solubility) could be experimentally synthesized and could have enhanced cytotoxic activity and even lower toxicity.

Ficaria verna Huds. extracts and their ß-cyclodextrin supramolecular systems.

BACKGROUND: Obtaining new pharmaceutical materials with enhanced properties by using natural compounds and environment-friendly methods is a continuous goal for scientists. Ficaria verna Huds. is a widespread perennial plant with applications in the treat of haemorrhoids and to cure piles; it has also anti-inflammatory, astringent, and antibiotic properties. The goal of the present study is the obtaining and characterization of new F. verna extract/ß-cyclodextrin complexes by using only natural compounds, solvents, and environment-friendly methods in order to increase the quality and acceptability versus toxicity indicator. Thus, the flavonoid content (as quercetin) of Ficaria verna Huds. flowers and leaves from the West side of Romania was determined and correlated with their antioxidant activity. Further, the possibility of obtaining ß-cyclodextrin supramolecular systems was studied. RESULTS: F. verna flowers and leaves extracts were obtained by semi-continuous solid-liquid extraction. The raw concentrated extract was spectrophotometrically analyzed in order to quantify the flavonoids from plant parts and to evaluate the antioxidant activity of these extracts. The F. verna extracts were used for obtaining ß-cyclodextrin complexes; these were analyzed by scanning electron microscopy and Karl Fischer water titration; spectrophotometry was used in order to quantifying the flavonoids and evaluates the antioxidant activity. A higher concentration of flavonoids of 0.5% was determined in complexes obtained by crystallisation method, while only a half of this value was calculated for kneading method. The antioxidant activity of these complexes was correlated with the flavonoid content and this parameter reveals possible controlled release properties. CONCLUSIONS: The flavonoid content of F. verna Huds. from the West side of Romania (Banat county) is approximately the same in flowers and leaves, being situated at a medium value among other studies. ß-Cyclodextrin complexes of F. verna extracts are obtained with lower yields by crystallisation than kneading methods, but the flavonoids (as quercetin) are better encapsulated in the first case most probably due to the possibility to attain the host-guest equilibrium in the slower crystallisation process. F. verna extracts and their ß-cyclodextrin complexes have antioxidant activity even at very low concentrations and could be used in proper and valuable pharmaceutical formulations with enhanced bioactivity.