EVALUATION OF ANOMERICRECOGNITION IN GALACTOSE BINDING LECTINS USING CROSSLINKED HEMICELLULOSE: A COMPARATIVE STUDY THROUGH AFFINITY CHROMATOGRAPHY

The isolation of lectins by affinity chromatography with crosslinked hemicelluloses has been a common practice because of the variety of glycosides that they present, improving the isolation of different kinds of lectins, such as the galactose ligands.Lectins affinity for carbohydrates is so specific that a simple configuration of the chiral carbon can affect affinity, and there are lectins that are more related to alfa-galactosidic than beta-galactosidic residues, setting up that way, an anomeric recognition.The anomeric configuration of galactose residuesseems to have biological importance related to the behavior of some diseases and physiological processes. This work aimed to assess the anomeric recognition of two lectins reported as β-galactose ligands (PNA and ricin) and two lectins reported as α-galactose ligands (frutalin and jacalin) in two types of hemicellulose (xyloglucan of Tamarindus indica and galactomannan of Caesalpinia pulcherrima), subsequently crosslinked and used as chromatographic matrices. As a result,chromatographic profiles and retained fractions suggested preferential anomeric recognition by lectins for the hemicelluloses crosslinked. The galactomannan matrix retained 0,5mg of PNA lectin and 2,3mg of ricin lectin; meanwhile, the xyloglucan matrix retained 3,4mg of PNA and 3,2mg of ricin; results obtained by applying 5 mg of lectin. Ricin expresses a visible flexibility in anomeric recognition, while PNA shows a restricted recognition of β-galactose residues.Frutalin and jacalin did not show recognition of the xyloglucan matrix. This work proposes using hemicellulose reticles with epichlorohydrin as affinity chromatographic matrices for anomeric studies on recognizing galactose binding lectins.

Onion (Allium cepa L.) peel extracts characterization by conventional and modern methods

The impact of different extraction methods on bioactive compounds (total phenols and flavonoids) recovery from onion wastes recovered from the market was investigated. The results revealed a total polyphenol content from 25.19 ± 3.56 mg gallic acid equivalents/g for enzyme-assisted extraction (EAE) to 212.56 ± 1.18 mg gallic acid equivalents/g for supercritical extraction (SFE). The total flavonoid content registered from 108.36 ± 3.62 mg quercetin equivalents/g for EAE to 334.97 ± 19.41 mg quercetin equivalents/g for conventional solvent extraction (CSE). The antioxidant activity (404.93 ± 1.39 mM Trolox/g) for SFE was the highest compared to all the extraction procedures used. The chromatographic profiles were conducted at five primary compound identification (quercetin 7,4-diglycoside, quercetin 3,4-diglycoside, quercetin 4-glucoside, quercetin, and kaempferol) and GC-MS analysis allowed the identification of 26 compounds, manly identified in EAE. The results also revealed that the SFE method, as a non-invasive technique, recorded the highest concentration of bioactive compounds and antioxidant activity, confirmed by chromatographic profiles.

PHENOLIC COMPOUNDS SANGUISORBA OFFICINALIS (ROSACEAE) GROWING IN EASTERN SIBERIA

The present work aimed at the study of the phenolic compounds of Sanguisorba officinalis L. (Rosaceae) growing in Eastern Siberia. The quantitative content of flavonoids in S. officinalis extracts determined by the spectrophotometric method was 0.68–5.41% in herb and 0.20–0.51% in rhizomes and roots. Catechin content was 0.05–0.75% in herb and 0.08–9.86% in rhizomes and roots, and procyanidin content found with spectrophotometric Porter method was 0.02–1.45% in herb and 0.08–1.05% in rhizomes and roots. Regression analysis of the relationships “latitude / longitude / altitude – phenolic compounds content” showed the high values of the correlation coefficient for the catechin content in herb and roots of S. officinalis and the longitude of the place of growth, as well as altitude. The content of flavonoids in the herb and roots of S. officinalis was connected with the longitude and latitude of the collection place. The values of procyanidin content in S. officinalis organs and geographic indicators have fewer relationships. Using the HPLC-DAD-ESI-MS method, chromatographic profiles of the herb and rhizomes and roots of S. officinalis was investigated. The data of chromatographic mobility, UV and mass spectra allowed estimating 52 compounds, including 45 in herb and 31 in rhizomes and roots. The members of seven groups of phenolic compounds have been found in S. officinalis, including gallotannins, flavonol-O-glycosides, procyanidins, ellagotannins, hydroxycinnamates, catechins, and pyrones, of which 12 are previously described for the species and the presence of 40 components, has been shown for the first time in the species.

Concentrating rosmarinic acid from Orthosiphon aristatus extract for high antioxidative candies

Extraction and fractionation were carried out to concentrate the rosmarinic acid from Orthosiphon aristatus in order to increase the value of herbal candy formulated using polyphenolic rich extract. The herb was extracted and fractionated in column chromatography, and then analyzed by LC-MS/MS. The collected plant fractions with similar chromatographic profiles were combined and determined for antioxidant capacities expressed in radical scavenging activity. The results showed that the antioxidant capacity was in good agreement with the concentration of rosmarinic acid in the combined fractions. The combined fraction II showed the highest rosmarinic acid content, 3.8%w/w and the highest antioxidant capacity (IC50=14.922 ppm). The incorporation of rosmarinic acid rich extract into candy formulation did not statistically affect the antioxidant capacity. Hence, the rosmarinic acid rich extract could be another choice of ingredient to enhance the beneficial property of candies. Candy is another form of carrier to deliver herbal ingredient for health promotion.

Analysis of phenolic compounds from cowpea (Vigna unguiculata) by HPLC-DAD-MS/MS

Vigna unguiculata (L.) Walp (cowpea), Fabaceae family and also known as Leguminosae, is an important vegetable used as food in tropical regions, especially in Africa, South America and Asia countries. Phenolic compounds are associated with important biological properties and their occurrence in edible plants may result in a highly functional food. Chromatographic profiles of phenolic compounds were investigated in two cowpea cultivars, such as tracuateua (CT) and caldeirão (CC), and both were cultivated using High Performance Liquid Chromatography (HPLC) coupled to Mass Spectrometry (MS) (HPLC-DAD/MS/MS). The flavonols quercetin and kaempferol, the phenolic acids, p-coumaric and protocatechuic acid (PCA) were identified in cowpea (CC), while the phenolic acids, gallic and protocatechuic acids, were identified in the cowpea (CT). These phenolic compounds ratify cowpea as a functional and bioactive food, ensuring a healthy diet.

Does Subunit Composition Influence the Intermolecular Crosslinking of Fish Collagen? A Study with Hake and Blue Shark Skin Collagens

Acid-soluble collagens from European hake and Blue shark skin were isolated, characterized, and compared. As the structure of collagen determines its function, the final objective of this study was to investigate biochemical differences between both collagens to identify future potential applications. Chromatographic behavior revealed differences in collagen from both species. Increases of temperature and stirring time produced no effect on European hake collagen solubility in the mobile phase, resulting in the same chromatographic profiles. Conversely, the application of temperature and stirring-time increments showed a positive effect on Blue shark collagen solubility, resulting in different chromatographic profiles and observing higher molecular weight components when sample is incubated at 50 °C (15 min) after 48 h stirring. To test if the different chromatographic behavior exhibited by both collagens could be influenced by differences in subunit composition (alpha-chains), cation exchange chromatography was employed to separate collagen subunits. The electrophoretic patterns and gel permeation chromatography with light-scattering detection (GPC-LS) results of the obtained cation exchange peak fractions revealed differences regarding subunit composition between both species, influencing the crosslinking pattern. This is the first comparative study using GPC-LS to provide information of European hake and Blue shark collagen subunit composition.

Comparison of chromatographic profiles of essential oils in extracts of 13 species from Lamiaceae family

The TLC essential oils profiles were being used conventionally for identification of herbal raw materials (HRM) of Lamiaceae family. Despite this, in recent years the European Pharmacopoeia to call in question the appropriateness of TLC on essential oils determination of Thyme and Wild Thyme, as well as Peppermint and Peppermint dry extract; these methods were replaced by HPTLC using flavonoid profile. This work aimed to establish the possibility of conducting HPTLC identification of methanolic extracts of 13 members of the Lamiaceae family in harmonized chromatographic conditions using chromatographic profiles on essential oils. The objects were methanolic extracts of HRM of Ukraine flora – Salvia officinalis, Salvia aethiopis, Thymus serpyllum, Thymus vulgaris, Origanum vulgare, Rosmarinus officinalis, Ocimum tenuiflorum (foreign origin), Mentha × piperita, Melissa officinalis, Orthosiphon stamineus, Agastache foeniculum, Leonurus cardiaca, Hyssopus officinalis, Marrubium vulgare, prepared from HRM that were purchased in Ukraine. The study wоas done by standardized HPTLC procedure using automatic HPTLC herbal system of CAMAG, Switzerland, at the base of CAMAG laboratory, Muttenz, Switzerland, and laboratory of ESTCCTI ESIPP, NUPh, Ukraine. Chromatography conditions: stationary phase – HPTLC plates Si 60 F254; mobile phase – toluene-ethylacetate (95:5); reference standards – borneol, bornyl acetate, cineole; derivatization – anisaldehyde reagent; detection: WRT, 254, 366 nm – before derivatization; WRT, 366 nm – after derivatization; sample preparation – methanolic extracts (100 mg/ml), extracted by shaking technique. In unified chromatographic conditions the HPTLC chromatographic fingerprints of essential oils of 13 Lamiaceae family species were established and evaluated. The most specific, intensive and reproducible HPTLC fingerprints were for Sage and Rosemary extracts. Moreover, fingerprints of Sage, Thyme, Rosemary, Origanum, Ocimum and Peppermint shown specific zones useful for differentiation of different genera of Lamiaceae family. For setting relative acceptance criteria for each of the species the validation for specific zones that were detected should be done.