Different Degrees Of Methylesterification Research Articles

Effects of pectin's degree of methyl esterification on TLR2-mediated IL-8 secretion and tight junction gene expression in intestinal epithelial cells: influence of soluble TLR2.

This study investigates the anti-inflammatory effects of pectins with different degrees of methyl esterification (DM) on intestinal epithelial cells (IECs) expressing low and high levels of TLR2. It also studies the influence of soluble TLR2 (sTLR2) which may be enhanced in patients with inflammatory bowel syndrome on the inflammation-attenuating effects of pectins. Also, it examines the impact of pectins on tight junction gene expression in IECs. Lemon pectins with DM18 and DM88 were characterized, and their effects on TLR2-1-induced IL8 gene expression and secretion were investigated in low-TLR2 expressing Caco-2 and high-TLR2 expressing DLD-1 cells. The results demonstrate that both DM18 and DM88 pectins can counteract TLR2-1-induced IL-8 expression and secretion, with more pronounced effects observed in DLD-1 cells expressing high levels of TLR2. Furthermore, the presence of sTLR2 does not interfere with the attenuating effects of low DM18 pectin and may even support its anti-inflammatory effects in Caco-2 cells. The impact of pectins and sTLR2 on tight junction gene expression also demonstrates cell-type-dependent effects. Overall, these findings suggest that low DM pectins possess potent anti-inflammatory properties and may influence tight junction gene expression in IECs, thereby contributing to the maintenance of gut homeostasis.

Development and characterization of films from Campomanesia xanthocarpa and commercial citrus pectins with different degrees of methyl-esterification

In this study, pectins from commercial citrus and isolated from gabiroba (Campomanesia xanthocarpa) fruits, were obtained with different degrees of methyl-esterification (DM) and applied in the films. The DM ranged from 0 % to 62.5 % and the gradual de-esterification process was confirmed by mono-dimensional analysis (1H NMR). In order to investigate the influence of DM values in pectin film properties, PCP (DM: 62.5 %); PCP-5 (DM: 37.4 %); PCP-15 (DM: 19.1 %), and a fully de-esterified sample PCP-35 (DM: 0 %) were selected. The functional properties of the films clearly showed that the DM and cross-linking process are necessary to obtain a material with water resistance. Furthermore, pectin isolated from the fruits of gabiroba was purified (GW-Na, DM: 51.9 %) and partially de-esterified (GW-Na-5, DM: 37.1 %). These pectins were used, for the first time, in development of films and the physical and mechanical properties were compared with films made with PCP and PCP-5 samples. GW-Na and GW-Na-5 films presented suitable properties, with reduced solubility reduced (57.1 and 26.2 %), high degree of swelling (2.14 and 2.26), low flexibility (18.05 and 6.11 MPa), respectively. High strength and rigidity (99.36 and 1040.9 MPa), for both films (GW-Na and GW-Na-5) were demonstrated, similar to that obtained by analyzed citrus pectin.

Efficacy of pectins with different degrees of methyl-esterification and of blockiness in preventing gut epithelial cell barrier disruption and the impact on sodium-glucose co-transporter expression under low and high glucose conditions.

Pectins support intestinal barrier function and have anti-diabetic effects, and can differ in the degree of methyl-esterification (DM) and the distribution of non-esterified galacturonic acid residues (DB). The mechanisms and effects of pectin type at different glucose levels are unknown. Pectins with different DM/DB on T84 cells were tested in the presence and absence of the barrier disruptor A23187 at 5 mM and 20 mM glucose. DM19 and DM43 pectins with high DB do rescue the intestinal barrier from disruption. Their effects were as strong as those of the barrier-rescuing anti-diabetic drug metformin, but effects with metformin were restricted to high glucose levels while pectins had effects at both low and high glucose levels. At high glucose levels, DM43HB pectin, which enhanced trans-epithelial electrical resistance, also increased the expressions of claudin1, occludin, and ZO-1. Low and high DM pectins decrease the apical expression of the sodium-glucose co-transporter (SGLT-1) and thereby influence glucose transport, explaining the anti-diabetogenic effect of pectin. Higher DB pectins had the strongest effect. Their impact on SGLT-1 was stronger than that of metformin. Pectin's rescuing effect on barrier disruption and its impact on glucose transportation and anti-diabetogenic effects depend on both the DB and the DM of pectins.

The gel strength and swelling in the gastrointestinal environment of pectin/κ-carrageenan gel particles based on pectins with different degrees of methylesterification

The gel strength and swelling in the gastrointestinal environment of pectin/κ-carrageenan gel particles based on pectins with different degrees of methylesterification

Production and molecular characterization of tailored citrus pectin-derived compounds

In this study, tailored-made citrus pectin-derived compounds were produced through controlled enzymatic and/or chemical modifications of commercial citrus pectin with different degrees of methylesterification (DM) and similar average molecular weight (MW). In the first treatment, degradation of the citrus pectin (CP) materials by endo-polygalacturonase (EPG) yielded pectins with average Mw’s (between 2 and 60 kDa). Separation and identification of the oligosaccharide fraction present in these samples, revealed the presence of non-methylesterified galacturonic acid oligomers with degree of polymerization (DP) 1–5. In the second treatment, exploiting the combined effect of EPG and pectin lyase, compounds with MW between 2 and 21 kDa, containing methylesterified and non-methylesterified polygalacturonans (DP 1–6), were generated. Finally, CP was sequentially modified by chemical saponification and the action of EPG. A sample of DM 11% and MW 2.7 kDa, containing POS (DP 1–5), was produced. Diverse pectin-derived compounds were successfully generated for further studies exploring their functionality.

Impact of Meloidogyne incognita (nematode) infection on root tissues and cell wall composition of okra (Abelmoschus esculentus L. Moench, Malvaceae).

Root-knot nematodes are endoparasites whose mature females lodge and grow inside the root of some cultivated plants, leading to losses in productivity. Herein, we investigated if the infection of okra, Abelmoschus esculentus (Malvaceae), promoted by the root-knot nematode Meloidogyne incognita (Meloidogynidae) changes some agronomic traits of the host plant, as well as the cell wall composition of the root tissues. The okra Santa Cruz 47® cultivar was infected with a suspension of 5000 M. incognita juveniles. The inoculated and non-inoculated okra plants were then submitted to morphological analysis at the end of experiment, as well as histological (at 4, 11, 18, 39, ad 66 days after inoculation) and immunocytochemical analysis (control and 66 days after inoculation). Root-knot nematode infection reduced the dry weight of the stem system but, unexpectedly, the number and weight of fruits increased. At 11 days after inoculation, we detected the presence of giant cells that increased in number and size until the end of the experiment, at 66 days after inoculation. These cells came from the xylem parenchyma and showed intense and moderate labeling for epitopes recognized by JIM5 and JIM7. The presence of homogalacturonans (HGs) with different degrees of methyl esterification seems to be related to the injuries caused by the nematode feeding activity and to the processes of giant cell hypertrophy. In addition, the presence of HGs with high methyl-esterified groups can increase the cell wall porosity and facilitate the flux of nutrients for the root-knot nematode.

Bio-Based Hydrogels Composed of Humic Matter and Pectins of Different Degree of Methyl-Esterification.

We prepared humo-pectic hydrogels through ionotropic gelation by crosslinking natural pectins of different degree of methyl-esterification with either humic substances (HS) extracted from cow manure compost or humic-like substances (HULIS) from depolymerized lignocellulose biorefinery waste. The hydrogels were characterized by solid-state 13C-NMR spectroscopy, scanning electron microscopy, spectroscopic magnetic resonance imaging and rheological analyses. Their ability to work as controlled release systems was tested by following the release kinetics of a previously incorporated model phenolic compound, like phloroglucinol. Our results indicated that the release properties of hydrogels were influenced by the molecular composition of HS and HULIS and by the different degrees of methyl-esterification of pectins. The hydrogel made by the high methoxyl pectin and HS showed the fastest rate of phloroglucinol release, and this was attributed not only to its morphological structure and crosslinking density but also to the least formation of ionic interactions between phloroglucinol and the polysaccharidic chains. Our study suggests that the efficiency of novel humo-pectic hydrogels as sustainable carriers of agroproducts to crops is related to a careful choice of the characteristics of their components.

Responsive polysaccharide-grafted surfaces for biotribological applications

The elucidation of biolubrication mechanisms and the design of artificial biotribological contacts requires the development of model surfaces that can help to tease out the cues that govern friction in biological systems. Polysaccharides provide an interesting option as a biotribological mimic due to their similarity with the glycosylated molecules present at biointerfaces. Here, pectin was successfully covalently grafted at its reducing end to a polydimethylsiloxane (PDMS) surface via a reductive amination reaction. This method enabled the formation of a wear resistant pectin layer that provided enhanced boundary lubrication compared to adsorbed pectin. Pectins with different degrees of methylesterification and blockiness were exposed to salt solutions of varying ionic strength and displayed responsiveness to solvent conditions. Exposure of the grafted pectin layers to solutions of between 1 and 200 mM NaCl resulted in a decrease in boundary friction and an increase in the hydration and swelling of the pectin layer to varying degrees depending on the charge density of the pectin, showing the potential to tune the conformation and friction of the layer using the pectin architecture and environmental cues. The robust and responsive nature of these new pectin grafted surfaces makes them an effective mimic of biotribological interfaces and provides a powerful tool to study the intricate mechanisms involved in the biolubrication phenomenon.

Integrative analysis of pectin methylesterase (PME) and PME inhibitors in tomato (Solanum lycopersicum): Identification, tissue-specific expression, and biochemical characterization

Although previous studies have demonstrated that the degree of demethylesterification of pectin polysaccharides is modulated during tomato fruit ripening, its involvement in vegetative organ development has been seldom investigated. As a first step in understanding the importance of pectin modification during vegetative stages, we used chemical, biochemical, and molecular approaches to analyze PMEs and PMEIs in tomato plants. We found that tomato cell walls isolated from vegetative tissues as well as the fruit contain substantial quantities of pectin, and different degrees of methylesterification were evident in different tissues. Our chemical study was further substantiated by immunolocalization analysis, which showed that selective removal of pectin-bound methyl groups is required for proper organ development and growth. In the tomato genome, there exists 79 PMEs and 48 PMEIs with temporally and spatially regulated expression. As a case study, we showed that two tomato PMEIs (SolycPMEI13 and SolycPMEI14) exhibited PMEI activities. This is the first report regarding the genome-wide identification and expression profiling of PME/PMEIs in tomato and the first chemical evidence of the differential degrees of pectin methylesterification in vegetative and reproductive tissues. Taken together, our findings provide an important tool to unravel the molecular and physiological functions of tomato PME and PMEI in further study.

Degree of pectin methyl esterification in endosperm cell walls is involved in embryo bending in Arabidopsis thaliana

The endosperm is a transitory structure involved in proper embryo elongation. The cell walls of mature seed endosperm are generally composed of a uniform distribution of cellulose, unesterified homogalacturonans, and arabinans. Recent studies suggest that changes in cell wall properties during endosperm development could be related to embryo growth. The degree of methyl esterification of homogalacturonans may be involved in this endosperm tissue remodelling. The relevance of the degree of homogalacturonan methyl esterification during seed development was determined by immunohistochemical analyses using a panel of probes with specificity for homogalaturonans with different degrees of methyl esterification. Low-esterified and un-esterified homogalacturonans were abundant in endosperm cells during embryo bending and were also detected in mature embryos. BIDXII (BDX) could be involved in seed development, because bdx-1 mutants had misshapen embryos. The methyl esterification pattern described for WT seeds was different during bdx-1 seed development; un-esterified homogalacturonans were scarcely present in the cell walls of endosperm in bending embryos and mature seeds. Our results suggested that the degree of methyl esterification of homogalacturonans in the endosperm cell wall may be involved in proper embryo development.

Morphology of complexes formed between β‐lactoglobulin nanofibrils and pectins is influenced by the pH and structural characteristics of the pectins

For the optimal use of β-lactoglobulin nanofibrils as a raw material in biological composites an in-depth knowledge of their interactions with other constituents is necessary. To understand the effect of electrostatic interactions on the morphology of resulting complexes, β-lactoglobulin nanofibrils were allowed to interact with pectins in which the amount of available negative charge was controlled by selecting their degree of methylesterification. In this study, citrus pectins having different degrees of methylesterification (∼48, 67, 86, and 97%) were selected and interacted with nanofibrils at pH 2 and pH 3, where they possess a net positive charge. Electrostatic complexes formed between β-lactoglobulin nanofibrils and all pectin types, except for the sample having a degree of methylesterification of 97%. The morphology of these complexes, however, differed significantly with the degree of methylesterification of the pectin, its concentration, and the pH of the medium, revealing that distinct desired biological architectures can be attained relatively easily through manipulating the electrostatic interactions. Interestingly, the pectin with a degree of methylesterification of 86% was found to crosslink the β-lactoglobulin nanofibrils into ordered 'nanotapes'.

Pectin nanostructure influences pectin-cation interactions and in vitro-bioaccessibility of Ca2+, Zn2+, Fe2+ and Mg2+-ions in model systems

This study explores the effect of pectin structure on mineral bioaccessibility in model systems. Commercial citrus and sugar beet pectin (CP and SBP, respectively) were demethylesterified by the action of carrot pectin methylesterase and characterisation of the resulting samples revealed different degrees of methylesterification (DM). Additionally, SBP exhibited a higher protein content, degree of acetylation, degree of feruloylation, degree of branching and molar mass compared to CP. In vitro-simulated digestion (stomach and small intestine phases) of mineral-enriched pectin solutions showed that decreasing pectin DM results in a decrease in mineral-bioaccessibility due to pronounced cation-pectin interactions. For a given DM, SBP digests exhibited higher mineral-bioaccessibilities than CP digests possibly due to structural differences of pectin in these sources. Furthermore, increasing the mineral-enrichment prior to in vitro-digestion resulted in a higher mineral-bioaccessibility while increasing the pectin concentration decreased the mineral-bioaccessibility. Generally, Mg2+-ions were the most bioaccessible followed by Ca2+, Zn2+, then Fe2+-ions.

Evaluation of cation-facilitated pectin-gel properties: Cryo-SEM visualisation and rheological properties

To provide more insight into the cation facilitated pectin-gel structure-function relationship, the microstructure of pectin-gels was explored using cryo-scanning electron microscopy (Cryo-SEM) in combination with physiochemical and rheological analysis. Pectin with different degrees of methylesterification, DM, was obtained from commercial citrus pectin (CP) and sugar beet pectin (SBP) by action of carrot pectin methylesterase. Pectin characterisation confirmed that pectin properties other than DM were unaltered and pectin from the two sources exhibited different structures. Cryo-SEM visualisation of pectin-gels prepared with different cations revealed that the network density increased with decreasing DM and increasing pH, pectin and cation-concentration. At a given DM, the more linear and less acetylated pectin (CP) exhibited a more intertwined fibrous network compared to the highly branched and more acetylated pectin (SBP). Furthermore, gel-microstructure was influenced by type of cation with Zn2+, Fe2+ and Ca2+-pectin gels showing a more intertwined network compared to Mg2+-pectin gels. Finally, rheological properties of gels complementing Cryo-SEM observations showed that as the network density increased, the gel-stiffness (G′) increased.

The impact of lemon pectin characteristics on TLR activation and T84 intestinal epithelial cell barrier function

Sufficient dietary fibre intake reduces the risk of several diseases, but the mechanisms linking fibre structure and health effects remain unclear. To analyse the influence of the amount of methyl groups esterified to the backbone, lemon pectins of different degrees of methyl esterification (30, 56, and 74DM) were studied for immune receptor activating potential and epithelial barrier protection. In reporter assays, the pectins demonstrated TLR/MyD88 dependent activation of NF-κB/AP-1, which increased with increasing pectin DM. To analyse the importance of backbone structure, the pectins were enzymatically digested into oligomers, which abrogated TLR activating potential. The 30 and 74DM pectins induced strong protection of the epithelial barrier measured by T84 transepithelial electrical resistance (TEER). These results indicate that activation of immune cells by lemon pectins is TLR dependent, and the intact polymer backbone is indispensable for activation. In addition, DM is a determining factor in activating potential and epithelial barrier protective effects.

Purification and Characterization of a Unique Pectin Lyase from Aspergillus giganteus Able to Release Unsaturated Monogalacturonate during Pectin Degradation.

A pectin lyase, named PLIII, was purified to homogeneity from the culture filtrate of Aspergillus giganteus grown in submerged culture containing orange peel waste as carbon source. PLIII was able to digest apple pectin and citrus pectins with different degrees of methyl esterification. Interestingly, the PLIII activity was stimulated in the presence of some divalent cations including Pb2+ and was not significantly affected by Hg2+. Like other pectin lyases, PLIII is stimulated by but is not dependent on Ca2+. The main soluble product released during the degradation of pectic substances promoted by the PLIII is compatible with an unsaturated monogalacturonate. PLIII is a unique enzyme able to release unsaturated monogalacturonate as the only soluble product during the degradation of pectic substances; therefore, PLIII was classified as an exo-pectin lyase. To our knowledge, this is the first characterization of an exo-pectin lyase. The PLIII described in this work is potentially useful for ethanol production from pectin-rich biomass, besides other common applications for alkaline pectinases like preparation of textile fibers, coffee and tea fermentation, vegetable oil extraction, and the treatment of pulp in papermaking.

A distinct role of pectate lyases in the formation of feeding structures induced by cyst and root-knot nematodes.

Pectin in the primary plant cell wall is thought to be responsible for its porosity, charge density, and microfibril spacing and is the main component of the middle lamella. Plant-parasitic nematodes secrete cell wall-degrading enzymes that macerate the plant tissue, facilitating the penetration and migration within the roots. In sedentary endoparasitic nematodes, these enzymes are released only during the migration of infective juveniles through the root. Later, nematodes manipulate the expression of host plant genes, including various cell wall enzymes, in order to induce specific feeding sites. In this study, we investigated expression of two Arabidopsis pectate lyase-like genes (PLL), PLL18 (At3g27400) and PLL19 (At4g24780), together with pectic epitopes with different degrees of methylesterification in both syncytia induced by the cyst nematode Heterodera schachtii and giant cells induced by the root-knot nematode Meloidogyne incognita. We confirmed upregulation of PLL18 and PLL19 in both types of feeding sites with quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) and in situ RT-PCR. Furthermore, the functional analysis of mutants demonstrated the important role of both PLL genes in the development and maintenance of syncytia but not giant cells. Our results show that both enzymes play distinct roles in different infected root tissues as well as during parasitism of different nematodes.

Chemical and antihyperglycemic activity changes of ginseng pectin induced by heat processing

Six pectic polysaccharides were obtained from white ginseng (GPW-1 and GPW-2), red ginseng (GPR-1 and GPR-2, steamed ginseng at 100°C) and steamed ginseng (GPS-1 and GPS-2, steamed ginseng at 120°C) by combination of water extraction, ion-exchange and gel permeation chromatographies. Based on the data from monosaccharide composition and 13C NMR analysis, GPW-1, GPR-1 and GPS-1 were identified as type-I rhamnogalacturonan (RG-I)-rich pectins, GPW-2, GPR-2 and GPS-2 were homogalacturonan (HG)-rich pectins with different degrees of methyl-esterification. Remarkably, GalA increased with the increase of processing temperatures in these six fractions, which might be caused by the transformation of esterified GalA into un-esterified form during heat processing. In vivo animal experiments showed that GPs exhibited significant antiohyperglycemic and antioxidant activities in alloxan-induced diabetic mice, and the effects increased with the processing temperature, with the most potent activity in GPS.

Molecular structural differences between low methoxy pectins induced by pectin methyl esterase II: Effects on texture, release and perception of aroma in gels of similar modulus of elasticity

Six low-methoxy pectins with different degrees of methylesterification and amidation, and molecular weights were used to prepare gels with similar moduli of elasticity by varying the concentrations of pectin and calcium phosphate. Five aroma compounds were added to the gels and their sensory textural properties, release and perception of aromas were investigated. Sensory firmness, springiness, adhesiveness, chewiness and cohesiveness differed according to the gel type, even though the moduli of elasticity were not significantly different (p<0.05). Release and perception of aromas also displayed significant difference according to the gel type (p<0.05). Low-methoxy amidated pectin exhibited the lowest release and perception for all the aroma compounds, while pectin-methylesterase-treated pectin gels exhibited relatively higher aroma release and perception. These results showed that the structural properties of pectins and gelling factors that increase the non-polar character of the gel matrices could decrease the release and perception of aromas in pectin gel systems.

Wound-induced pectin methylesterases enhance banana (Musa spp. AAA) susceptibility to Fusarium oxysporum f. sp. cubense

Recent studies suggest that plant pectin methylesterases (PMEs) are directly involved in plant defence besides their roles in plant development. However, the molecular mechanisms of PME action on pectins are not well understood. In order to understand how PMEs modify pectins during banana (Musa spp.)–Fusarium interaction, the expression and enzyme activities of PMEs in two banana cultivars, highly resistant or susceptible to Fusarium, were compared with each other. Furthermore, the spatial distribution of PMEs and their effect on pectin methylesterification of 10 individual homogalacturonan (HG) epitopes with different degrees of methylesterification (DMs) were also examined. The results showed that, before pathogen treatment, the resistant cultivar displayed higher PME activity than the susceptible cultivar, corresponding well to the lower level of pectin DM. A significant increase in PME expression and activity and a decrease in pectin DM were observed in the susceptible cultivar but not in the resistant cultivar when plants were wounded, which was necessary for successful infection. With the increase of PME in the wounded susceptible cultivar, the JIM5 antigen (low methyestrified HGs) increased. Forty-eight hours after pathogen infection, the PME activity and expression in the susceptible cultivar were higher than those in the resistant cultivar, while the DM was lower. In conclusion, the resistant and the susceptible cultivars differ significantly in their response to wounding. Increased PMEs and thereafter decreased DMs acompanied by increased low methylesterified HGs in the root vascular cylinder appear to play a key role in determination of banana susceptibility to Fusarium.

Dispersions of Nanoclays of Different Shapes into Aqueous and Solid Biopolymeric Matrices. Extended Physicochemical Study

Dispersions of nanofillers into aqueous and solid biopolymeric matrices were studied from the physicochemical viewpoint. This work was carried out based on the idea that the combination of biopolymers, derived from renewable resources, and nanofiller, environmentally friendly, may form a new generation of nanomaterials with excellent and unique properties at low cost. To this purpose, two pectins with different degrees of methyl esterification and nanoclays like halloysite and laponite RD were selected. The thermodynamic and structural studies on the aqueous mixtures of pectin and nanoclay were able to discriminate the interactions, which control the adsorption of pectin onto the filler and the aggregation of both pectin and clay particles. The gained insights were useful to interpret the mesoscopic structure of the nanocomposites (prepared from the aqueous mixtures by means of the casting method) evidenced by SEM, thermal stability, tensile properties, and transparency investigations. The attained knowledge represents a basic point for designing new hybrid nanostructures in both the aqueous and the solid phase for specific purposes.