Victoria V. Golovchenko

Pectic polysaccharides of the fresh plum Prunus domestica L. isolated with a simulated gastric fluid and their anti-inflammatory and antioxidant activities

A pectic polysaccharide, designated as PD, was extracted from fresh plums (Prunus domestica L.) with a simulated gastric fluid. Galacturonan, which was partially substituted with methyl and O-acetyl ester groups, and rhamnogalacturonan were the main constituents of the linear regions of the sugar chains of PD. The ramified region contained mainly 1,4-linked β-d-galactopyranose residues and, to a lesser extent, 1,5-linked α-l-arabinofuranose residues. The separation of PD, by DEAE-cellulose column chromatography, yielded two pectic fractions: PD-1 and PD-2, eluted with 0.1 and 0.2 M NaCl, respectively. Enzymatic digestion of PD with 1,4-α-d-polygalacturonase yielded the fraction PD-E. The parent pectin PD and the PD-1 fraction were found to diminish the adhesion of peritoneal leukocytes at the concentrations of 0.05-1.0mg/ml. However, the PD-E fraction failed to have an effect on cell adhesion at the concentrations of 0.05-0.1mg/ml. PD, PD-1 and PD-E were found to inhibit the production of superoxide anion radicals by reducing xanthine oxidase activity by 38%, 97% and 47%, respectively. Therefore, the PD-1 fraction appeared to be an active fragment of pectic macromolecule isolated from fresh plum with a simulated gastric fluid.

Structure of pectic polysaccharides isolated from onion Allium cepa L. using a simulated gastric medium and their effect on intestinal absorption

The polysaccharide fraction extracted with simulated gastric juice from onion bulbs contained a mixture of galactan with short-length sugar chains, pectic polysaccharides and evident content of proteinaceous material. Galacturonan and rhamnogalacturonan were the main constituents of the linear regions of the sugar chains of the pectic polysaccharides. The ramified regions included rhamnogalacturonan-I. NMR data revealed that the side chains of the ramified region contained mainly 1,4-linked β-D-galactopyranose residues and lesser content of 1,3-linked β-D-galactopyranose and 1,5-linked α-L-arabinofuranose residues. Furthermore, the proteinaceous material was determined to be partly linked to the sugar chains. The polysaccharide fraction was found to decrease absorption of ovalbumin (OVA) to the blood from the gut lumen. The serum OVA level was threefold lower in mice fed with OVA mixed with the onion pectins compared with the control group, which was administered OVA alone. Protein removal failed to abolish the inhibitory effect of the onion polysaccharides, confirming that the polysaccharide chains are the active component of onion gastric juice extract.

Pectin-silica gels as matrices for controlled drug release in gastrointestinal tract.

The synthesis of pectin-silica gels for controlled drug release in gastrointestinal tract (GIT) using low methoxyl (LM) and high methoxy (HM) pectins and tetraethoxysilane (TEOS) as precursor is described. The FTIR spectra of the pectin-silica gels show intense absorption bands at 1246cm-1 and 802cm-1 corresponding to the vibrations COSi bonds, which absent in the FTIR spectra of the native pectins that indicate the formation covalent bond between silica and pectin macromolecules in the pectin-silica gels. Pectin-TEOS, pectin-Ca-TEOS and pectin-TEOS-Ca beads with mesalazine are synthesized by different combinations of sol-gel method using TEOS and ionotropic gelation method using calcium chloride. The best resistant of pectin-TEOS and pectin-Ca-TEOS beads during incubation in simulated gastric fluid for 2h and subsequently in simulated intestinal fluids for 18h is indicated. Pectin-TEOS beads are characterized by higher encapsulation efficiency (to 28%) than pectin-Ca-TEOS beads (to 16%). The drug release of pectin-silica beads in simulated GIT occurs gradually up to 80% and is directly dependent on the hardness of the beads. The surface morphology of beads is shown. The use of pectin-silica beads is promising with regard to the development of controlled release of drug formulations.

Structural characterisation of the polysaccharides from endemic Mongolian desert plants and their effect on the intestinal absorption of ovalbumin

Using successive extractions with water and 0.7% aqueous ammonium oxalate, pectic polysaccharides were isolated from the following plants growing in the arid climate of Mongolia (Gobi): saxaul Haloxylon ammodendron Maxim., rhubarb Rheum nanum Sievers, Nitraria sibirica Pall., Peganum harmala L. and almond Amygdalus mongolica Maxim. The data obtained exhibited the primary synthesis of the cell wall pectic polysaccharides but not the middle lamellae water-soluble pectins in plants growing in the dry climatic zone. Both α-(1→4)-D-galacturonan and α-(1→4)-D-galacturonan, which was substituted with methyl groups, were found to be backbone of pectins. The L-arabinofuranose residues were identified as the main components of ramified regions. The pectins from almond differed from other pectins due to a high arabinose content. The data from NMR spectroscopy and methylation analyses demonstrated that pectic polysaccharides from almond included terminal, (1→5)-, (1→3)-linked and 3,5-substituted L-arabinofuranose residues and a small terminal D-galactopyranose and 2,5- and 2,3,5-substituted L-arabinofuranose residue content. The pectic polysaccharides were found to decrease the absorption of ovalbumin (OVA) in the blood from the gut lumen. The serum OVA level was lower in mice fed with OVA mixed with the pectins compared with the control group, which was administered OVA alone.

Impact of cabbage pectin-protein complex on microbial β-glucuronidase activity.

We previously demonstrated that pectin-protein complex (PPC) isolated from white cabbage adsorbs the β-glucuronidase (βG) enzyme of E. coli. Concurrently, we discovered a significant increase in βG activity in the presence of PPC. The aim of this study is to identify the structural components of PPC that are responsible for βG adsorption and activation. PPC was isolated from white cabbage using a saline solution containing hydrochloric acid (pH 1.5) at 37 °C for 4 h. PPC proteins were precipitated by aqueous 10% (m/v) trichloroacetic acid to yield the pectin-protein fractions PPC1 and PPC2. PPC was digested using 1,4-α-d-galacturonase, yielding the PPC6 fraction. Partial acid hydrolysis of PPC revealed the galacturonan fraction, PPC3, to be the core of the macromolecule. The purified PPC4 and PPC5 fractions were isolated from PPC by ion-exchange chromatography on DEAE-cellulose. βG activity and its adsorption in the PPC fractions were studied in vitro. Crystalline cellulose was used as a control. This study found that the PPC3 fraction (the galacturonan core) does not adsorb βG and does not affect its activity. The adsorption of βG in the PPC samples is inversely proportional to the degree of methyl esterification of its carbohydrate component. The PPC4 and PPC5 fractions adsorb the highest proportion of βG (51.2% and 54%, respectively). The stimulation of βG enzyme activity is directly proportional to the protein content of the PPC sample. The PPC and PPC1 samples have the greatest ability to increase βG activity (57.6% and 52.1%, respectively).

Interaction of Microbial β-Glucuronidase with Vegetable Pectins

The purpose of this study was to investigate the in vitro effects of vegetable carbohydrates on the activity of microbial β-glucuronidase (βG) and the adsorption of the enzyme on carbohydrates. This study used pectin-protein complexes (PPCs) with molecular weights of 300 kDa isolated under conditions simulating a gastric environment from cabbage (HCl-PPCC and HCl+pepsin-PPCCP) and sweet pepper (PPCP and PPCPP). As a sample for comparison, microcrystalline cellulose was used. The activity of βG from Escherichia coli was determined spectrophotometrically by the formation of the colored product from the breakdown of phenolphthalein-β-D-glucuronide. Adsorption of βG on biopolymers was studied by the retention of the enzyme on the membrane of a concentrator with a pore diameter of 300 kDa and by native PAGE. PPCCP and PPCC were established to increase the activity of βG by 50 and 100%, respectively. Cellulose had a weak effect, whereas pepper PPC had no effect. All studied carbohydrates adsorb on βG. The maximum βG adsorption (15%) was observed with PPCC, whereas PPCCP absorbed 5% of the enzyme. Pepper PPCs and cellulose adsorbed up to 10% of the enzyme. There was a positive correlation between the increase of βG activity in the presence of carbohydrates and enzyme adsorption on the polymers (r=0.80; P<0.01). The activity of the enzyme in the gel after electrophoresis of the PPCC+βG mixture was inversely proportional to the concentration of PPCC in the mixture. A model explaining the effects of cabbage PPCs on the excretion of estrogens is proposed.

The Antioxidant Properties of Pectin Fractions Isolated from Vegetables Using a Simulated Gastric Fluid

The antioxidant properties of vegetable pectin fractions against intraluminal reactive oxygen species were elucidated in vitro in conjunction with their structural features. The pectin fractions were isolated using a simulated gastric fluid (pH 1.5, pepsin 0.5 g/L, 37°C, 4 h) from fresh white cabbage, carrot, onion, and sweet pepper. The fraction from onion was found to inhibit the production of superoxide radicals by inhibiting the xanthine oxidase. The high molecular weight of onion pectin and a large number of galactose residues in its side chains appeared to participate in interaction with xanthine oxidase. All the isolated pectic polysaccharides were found to be associated with protein (2–9%) and phenolics (0.5–0.7%) as contaminants; these contaminants were shown to be responsible for the antioxidant effect of vegetable pectin fractions against the hydroxyl and 1,1-diphenyl-2-picrylhydrazyl radicals.

Injectable hydrogel from plum pectin as a barrier for prevention of postoperative adhesion

An injectable hydrogel was obtained from the high methyl-esterified plum Prunus domestica L. (PD) pectin and calcium ions (Ca2+). PD hydrogel showed a weak gel-like behavior and could be squeezed out of the syringe with an injection force of ca. 9 N. PD hydrogel was not suitable for the NIH/3T3 fibroblast cell adhesion in vitro. The live/dead fluorescence and MTT (3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide) assays indicated that the PD hydrogel had a low cytotoxicity in relation to both the adhered and gel surrounding fibroblasts. PD hydrogel was found to inhibit adhesion formation in the sidewall defect-cecum abrasion rat model. In the control group, the occurrence of adhesion of the cecum to the peritoneal wall was found in seven of the total seven rats operated. Only four of the seven animals that were treated with the PD hydrogel were noted to have any adhesions. These adhesions were of a minimum grade of 1, 2, and 3 and were represented by a thin film that could be easily broken. The protective effect of PD hydrogel was found to be comparable with that of hyaluronic acid hydrogel used as a positive control. PD hydrogel appeared to possess enhanced in vivo residence stability on the injury sites compared to hyaluronic acid hydrogel as measured by staining of healing tissue with periodic acid-Schiff reagent. The data obtained offered the prospect for the development of the pectin-based gels as new barrier materials for surgery.

Structure characterization of the mannofucogalactan isolated from fruit bodies of Quinine conk Fomitopsis officinalis

The mannofucogalactan as a major component of water extract was obtained from fruit bodies of Fomitopsis officinalis by extraction with boiling water followed by deproteination, decoloration, and purification using anion-exchange chromatography and size exclusion chromatography. Its structure was characterized using the data of monosaccharide composition, methylation analysis, one- and two-dimensional NMR spectroscopy. The studied polysaccharide was a branched mannofucogalactan with a backbone composed of partially 3-O-methylated 1,6-O-linked α-D-galactopyranosyl residues. Almost every second residue in the backbone was substituted at O-2 by 3-O-α-D-mannopyranosyl-α-L-fucopyranosyl and β-D-galactopyranosyl residues. The non-reducing terminal α-L-fucopyranosyl units, which were identified by GC-MS analyses, appeared to be the part of mannofucogalactan side chains also.