Pattern Of Methylesterification Research Articles

Gelation behavior between Nicandra physalodes (Linn.) Gaertn. pectin and Ca2+ is governed by degree and pattern of methylesterification

Nicandra physalodes (Linn.) Gaertn. Pectin (NPGP) is a new type of natural low methoxy pectin (LMP), however, the relatively poor gelling capacity of NPGP and neutral calcium limits its applicability in the food industry. In this study, we improved the gelling capacity of NPGP, and the impact of the degree and pattern of methyl esterification on NPGP gelation behavior triggered by neutral calcium was investigated. To achieve this goal, NPGP was treated with pectin methyl esterase (PME, 30 °C, pH 4.5) for various time. This modification decreased the degree of methyl esterification (DM) and generated different patterns of methyl ester distribution, quantified as the degree of blockiness (DB). The obtained PME-NPGPs possess the ability to form self-supporting gels in the presence of CaCl2, with the highest gel hardness being 331.98 g. By estimating the DM and quantifying the DB of PME-NPGP, the DB value of PME-NPGP was increased from 17.74% to 31.84% with DM decreasing from 33.09% to 17.37%, indicating a shift toward a more blockwise distribution of nonmethylated GalA in NPGP. Additionally, adsorption isotherms for Ca2+ on de-esterified NPGP were determined, enabling the quantification of its cation binding capacity and interaction energy. Reducing DM and increasing DB of NPGP were found to enhance NPGP's binding capacity to Ca2+. This research provides an important foundation for NPGP food processing, and the obtained PME-NPGP can be promising in the fields of food gel and drug delivery.

Genome-wide survey and evolutionary history of the pectin methylesterase (PME) gene family in the Dothideomycetes class of fungi

Once deposited in the plant cell wall, pectin undergoes demethylesterification by endogenous pectin methylesterases (PMEs), which play various roles in growth and development, including defense against pathogen attacks. Pathogen PMEs can alter pectin's methylesterification pattern, increasing its susceptibility to degradation by other fungal pectinases and thus playing a critical role as virulence factors during early infection stages. To investigate the evolutionary history of PMEs in the Dothideomycetes class of fungi, we obtained genomic data from 15 orders (79 species) and added genomic data from 61 isolates of Corynespora cassiicola. Our analyses involved maximum likelihood phylogenies, gene genealogies, and selection analyses. Additionally, we measured PME gene expression levels of C. cassiicola using soybean as a host through RT-qPCR assays. We recovered 145 putative effector PMEs and 57 putative non-effector PMEs from across the Dothideomycetes. The PME gene family exhibits a small size (up to 5 members per genome) and comprises three major clades. The evolutionary patterns of the PME1 and PME2 clades were largely shaped by duplications and recurring gene retention events, while biased gene loss characterized the small-sized PME3 clade. The presence of five members in the PME gene family of C. cassiicola suggests that the family may play a key role in the evolutionary success of C. cassiicola as a polyphagous plant pathogen. The haplogroups Cc_PME1.1 and Cc_PME1.2 exhibited an accelerated rate of evolution, whereas Cc_PME2.1, Cc_PME2.2, and Cc_PME2.3 seem to be under strong purifying selective constraints. All five PME genes were expressed during infection of soybean leaves, with the highest levels during from six to eight days post-inoculation. The highest relative expression level was measured for CC_29_g7533, a member of the Cc_PME2.3 clade, while the remaining four genes had relatively lower levels of expression.

A novel multiplex lateral flow assay for rapid assessment of pectin structural/functional properties

Pectin is a multi-faceted functional food ingredient widely used in the food, pharmaceutical and personal care industries for different purposes, based on its functional properties, which are primarily determined by the structural properties (degree and pattern of methyl-esterification) of its homogalacturonan (HG) backbone. Currently, pectin's commercial functionality is defined by lengthy, laborious and operator dependent tests. Using pectin homogalacturonan (HG) recognizing antibodies, we have developed a novel, rapid, easy to use lateral flow assay (LFA) to detect and assess pectin structural properties that are related to its functionality. The LFA system includes three different capture antibodies immobilized on a test strip and two detection antibodies labeled with different colors of Gold NanoParticles in a small vial. A user needs only to drop a pectin sample into the small vial and insert the test strip into the vial, then wait 5 min to obtain results. With a detection limit of 0.02 μg/mL of pectin, the developed LFA is able to discern pectin's degree and pattern of methyl-esterification. The capability of the LFA has been validated by testing a series of known experimental and commercial pectins. The technology can be easily scaled up to high-volume manufacturing. The convenience of such an LFA would benefit the entire spectrum of the pectin utilization chain, from producers and ingredient suppliers to food formulators and those in the medical and personal care industries.

Determining Methyl-Esterification Patterns in Plant-Derived Homogalacturonan Pectins.

Homogalacturonan (HG)-type pectins are nutrient components in plants and are widely used in the food industry. The methyl-esterification pattern is a crucial structural parameter used to assess HG pectins in terms of their nutraceutical activity. To better understand the methyl-esterification pattern of natural HG pectins from different plants, we purified twenty HG pectin-rich fractions from twelve plants and classified them by their monosaccharide composition, Fourier transform-infrared spectroscopy (FT-IR) signatures, and NMR analysis. FT-IR shows that these HG pectins are all minimally esterified, with the degree of methyl-esterification (DM) being 5 to 40%. To examine their methyl-esterification pattern by enzymatic fingerprinting, we hydrolyzed the HG pectins using endo-polygalacturonase. Hydrolyzed oligomers were derivatized with 2-aminobenzamide and subjected to liquid chromatography-fluorescence-tandem mass spectrometry (HILIC-FLR-MSn). Twenty-one types of mono-/oligo-galacturonides having DP values of 1–10 were found to contain nonesterified monomers, dimers, and trimers, as well as oligomers with 1 to 6 methyl-ester groups. In these oligo-galacturonides, MSn analysis demonstrated that the number of methyl-ester groups in the continuous sequence was 2 to 5. Mono- and di-esterified oligomers had higher percentages in total methyl-esterified groups, suggesting that these are a random methyl-esterification pattern in these HG pectins. Our study analyzes the characteristics of the methyl-esterification pattern in naturally occurring plant-derived HG pectins and findings that will be useful for further studying HG structure-function relationships.

Revealing methyl-esterification patterns of pectins by enzymatic fingerprinting: Beyond the degree of blockiness

Citrus pectins were studied by enzymatic fingerprinting using a simultaneous enzyme treatment with endo-polygalacturonase (endo-PG) from Kluyveromyces fragilis and pectin lyase (PL) from Aspergillus niger to reveal the methyl-ester distribution patterns over the pectin backbone. Using HILIC-MS combined with HPAEC enabled the separation and identification of the diagnostic oligomers released. Structural information on the pectins was provided by using novel descriptive parameters such as degree of blockiness of methyl-esterified oligomers by PG (DBPGme) and degree of blockiness of methyl-esterified oligomers by PL (DBPLme). This approach enabled us to clearly differentiate citrus pectins with various methyl-esterification patterns. The simultaneous use of PG and PL showed additional information, which is not revealed in digests using PG or PL alone. This approach can be valuable to differentiate pectins having the same DM and to get specific structural information on pectins and therefore to be able to better predict their physical and biochemical functionalities.

Cell wall O-acetyl and methyl esterification patterns of leaves reflected in atmospheric emission signatures of acetic acid and methanol.

Plants emit high rates of methanol (meOH), generally assumed to derive from pectin demethylation, and this increases during abiotic stress. In contrast, less is known about the emission and source of acetic acid (AA). In this study, Populus trichocarpa (California poplar) leaves in different developmental stages were desiccated and quantified for total meOH and AA emissions together with bulk cell wall acetylation and methylation content. While young leaves showed high emissions of meOH (140 μmol m-2) and AA (42 μmol m-2), emissions were reduced in mature (meOH: 69%, AA: 60%) and old (meOH: 83%, AA: 76%) leaves. In contrast, the ratio of AA/meOH emissions increased with leaf development (young: 35%, mature: 43%, old: 82%), mimicking the pattern of O-acetyl/methyl ester ratios of leaf bulk cell walls (young: 35%, mature: 38%, old: 51%), which is driven by an increase in O-acetyl and decrease in methyl ester content with age. The results are consistent with meOH and AA emission sources from cell wall de-esterification, with young expanding tissues producing highly methylated pectin that is progressively demethyl-esterified. We highlight the quantification of AA/meOH emission ratios as a potential tool for rapid phenotype screening of structural carbohydrate esterification patterns.

Structural, rheological and functional properties of galactose-rich pectic polysaccharide fraction from leek

An acid-extracted polysaccharide from alchohol-insoluble solids of leek was obtained. The sugar composition indicated that galactose and galacturonic acid were the major sugars, followed by small amounts of rhamnose and arabinose. The fraction contained a relatively high methyl-esterified homogalacturonan next to rhamnogalacturonan type I decorated with galactose-rich side chains. The fraction consisted of three high Mw populations, covering the range of 10–100 kDa. Enzymatic fingerprinting was performed with HG/RG-I degrading enzymes to elucidate the structure. The oligomers were analysed using LC-HILIC-MS, HPAEC, and MALDI-TOF MS. The data revealed the presence of GalA sequences, having different patterns of methyl-esterification, RG-I composed of unbranched segments and segments heavily substituted with β-(1→4)-linked galactan chains of varying length. The rheological study showed the shear-thinning, weak thixotropic, anti-thixotropic, and non-Newtonian behavior of the polysaccharide. The pectin exhibited higher water holding capacity than oil-holding capacity and the fraction did form stable foams at high concentration.

Isothermal titration calorimetry to study the influence of citrus pectin degree and pattern of methylesterification on Zn2+ interaction

This work explored the influence of the citrus pectin degree and pattern of methylesterification on its interaction with Zn2+ using isothermal titration calorimetry (ITC). Pectin samples with a comparable degree of methylesterification (DM) but distinct distribution patterns of non-methylesterified carboxylic groups (absolute degree of blockiness, DBabs) were produced through enzymatic (blockwise pattern) or alkaline (random pattern) demethylesterification. The pectin-Zn2+ interaction was found to be endothermic, in which a positive entropy change compensated for the unfavorable endothermic enthalpy change, driving the interaction between pectin and Zn2+. Decreasing pectin DM or increasing DBabs promoted Zn2+ binding, with the estimated binding capacity (mol Zn2+/mol GalA) and binding constant (mM−1) being mainly determined by pectin DBabs, rather than DM. ITC was found to be a useful technique to study the pectin-cation interaction, however, low DM pectin samples are sensitive to gelling and therefore more difficult to study.

Interactions between citrus pectin and Zn2+ or Ca2+ and associated in vitro Zn2+ bioaccessibility as affected by degree of methylesterification and blockiness

Insight into the effect of the degree and pattern of methylesterification of citrus pectin on the Ca2+ and Zn2+ adsorption capacity and the associated Zn2+ bioaccessibility was established. Pectins with comparable methylesterification degrees (DM) but different patterns of methylester distribution (blockwise or random) were generated. Blockwise distributions of methylesters were produced through controlled enzymatic means (using carrot pectin methylesterase) while random patterns were obtained through alkaline demethylesterification (using NaOH). The pattern of methylesterification was estimated as the absolute degree of blockiness (DBabs), which is the ratio of non-methylesterified units present in blocks to the total number of galacturonic acid units. Determination and modeling (based on the Langmuir adsorption model) of the adsorption isotherms of these structurally modified pectins for Zn2+ or Ca2+ allowed quantification of their maximum cation binding capacity and interaction energy. Decreasing pectin DM and increasing DBabs were shown to promote the Zn2+ or Ca2+ binding capacity of pectin, with the maximum binding capacity being mainly determined by the DM and interaction energy by DBabs. The influence of cation type was evident, with higher maximum binding capacities and interaction energies exhibited for Zn2+ compared to Ca2+, mainly due to the higher electronegativity of the former. Increased Zn2+ binding to pectin was associated with lower Zn2+ bioaccessibility and no influence of low levels of added Ca2+ was observed. However, enzymes and bile salts added during in vitro simulated digestion were shown to bind substantial amounts of Zn2+ ions.

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.

Fe2+ adsorption on citrus pectin is influenced by the degree and pattern of methylesterification

The present study aimed at gaining insight into the potential of citrus pectin to bind Fe2+, a cation of great importance in several food products. In particular, the role of citrus pectin structural properties, namely the degree of methylesterification (DM) and the absolute degree of blockiness (DBabs – ratio of non-methylesterified GalA units present in blocks to the total amount of GalA units) on the Fe2+ adsorption in aqueous solution was explored using adsorption isotherms. Demethylesterification of high DM citrus pectin enzymatically (using plant pectin methylesterase) or chemically (alkaline demethylesterification using NaOH) generated P- and C-pectins, respectively, characterized by comparable DMs but different distributions of non-methylesterified GalA units (DBabs). Adsorption isotherms of P- and C-pectins in aqueous solutions of various Fe2+ concentrations revealed that both the DM and DBabs influenced the pectin-Fe2+ interactions: the lower DM or higher DBabs, the higher the Fe2+ binding capacity of citrus pectin. The Langmuir adsorption model was used to fit the experimental data for quantification of the maximum adsorption capacity (qGmax) and the pectin-Fe2+ interaction energy (KL) of the P- and C-pectins. It can be concluded that qGmax (mol Fe2+/mol GalA) was mainly determined by the DM and to a lesser extent by the DBabs while the pectin-Fe2+ interaction energy was mainly influenced by the DBabs. As a consequence, pectin modification to obtain targeted DM and DBabs allows optimization of its binding capacity and therefore the associated functional properties.

Regulation of cell wall remodeling in grapevine (Vitis vinifera L.) callus under individual mineral stress deficiency

Cell wall (CW) is a dynamic structure that determines the plant form, growth and response to environmental conditions. Vitis vinifera callus grown under nitrogen (−N), phosphorous (−P) and sulfur (−S) deficiency were used as a model system to address the influence of mineral stress in CW remodeling. Callus cells morphology was altered, mostly under –N, resulting in changes in cell length and width compared with the control. CW composition ascertained with specific staining and immuno-detection showed a decrease in cellulose and altered pattern of pectin methylesterification. Under mineral stress genes expression from candidate families disclosed mainly a downregulation of a glycosyl hydrolase family 9C (GH9C), xyloglucan transglycosylase/hydrolases (XTHs) with predicted hydrolytic activity and pectin methylesterases (PMEs). Conversely, upregulation of PMEs inhibitors (PMEIs) was observed. While methylesterification patterns can be associated to PME/PMEI gene expression, the lower cellulose content cannot be attributed to altered cellulose synthase (CesA) gene expression suggesting the involvement of other gene families. Salt extracts from –N and −P callus tissues increased plastic deformation in cucumber hypocotyls while no effect was observed with −S extracts. The lower endo-acting glycosyl hydrolase activity of −N callus extracts pinpoints a more expressive impact of −N on CW-remodeling.

Pectinmethylesterases (PME) and pectinmethylesterase inhibitors (PMEI) enriched during phloem fiber development in flax (Linum usitatissimum).

Flax phloem fibers achieve their length by intrusive-diffusive growth, which requires them to penetrate the extracellular matrix of adjacent cells. Fiber elongation therefore involves extensive remodelling of cell walls and middle lamellae, including modifying the degree and pattern of methylesterification of galacturonic acid (GalA) residues of pectin. Pectin methylesterases (PME) are important enzymes for fiber elongation as they mediate the demethylesterification of GalA in muro, in either a block-wise fashion or in a random fashion. Our objective was to identify PMEs and PMEIs that mediate phloem fiber elongation in flax. For this purpose, we measured transcript abundance of candidate genes at nine different stages of stem and fiber development and found sets of genes enriched during fiber elongation and maturation as well as during xylem development. We expressed one of the flax PMEIs in E. coli and demonstrated that it was able to inhibit most of the native PME activity in the upper portion of the flax stem. These results identify key genetic components of the intrusive growth process and define targets for fiber engineering and crop improvement.

Substrate Dynamics in Enzyme Action: Rotations of Monosaccharide Subunits in the Binding Groove are Essential for Pectin Methylesterase Processivity

The dynamical behavior of biomacromolecules is a fundamental property regulating a large number of biological processes. Protein dynamics have been widely shown to play a role in enzyme catalysis; however, the interplay between substrate dynamics and enzymatic activity is less understood. We report insights into the role of dynamics of substrates in the enzymatic activity of PME from Erwinia chrysanthemi, a processive enzyme that catalyzes the hydrolysis of methylester groups from the galacturonic acid residues of homogalacturonan chains, the major component of pectin. Extensive molecular dynamics simulations of this PME in complex with decameric homogalacturonan chains possessing different degrees and patterns of methylesterification show how the carbohydrate substitution pattern governs the dynamics of the substrate in the enzyme’s binding cleft, such that substrate dynamics represent a key prerequisite for the PME biological activity. The analyses reveal that correlated rotations around glycosidic bonds of monosaccharide subunits at and immediately adjacent to the active site are a necessary step to ensure substrate processing. Moreover, only substrates with the optimal methylesterification pattern attain the correct dynamical behavior to facilitate processive catalysis. This investigation is one of the few reported examples of a process where the dynamics of a substrate are vitally important.

Methyl esterification of pectin plays a role during plant–pathogen interactions and affects plant resistance to diseases

The cell wall is a complex structure mainly composed by a cellulose–hemicellulose network embedded in a cohesive pectin matrix. Pectin is synthesized in a highly methyl esterified form and is de-esterified in muro by pectin methyl esterases (PMEs). The degree and pattern of methyl esterification affect the cell wall structure and properties with consequences on both the physiological processes of the plants and their resistance to pathogens. PME activity displays a crucial role in the outcome of the plant–pathogen interactions by making pectin more susceptible to the action of the enzymes produced by the pathogens. This review focuses on the impact of pectin methyl esterification in plant–pathogen interactions and on the dynamic role of its alteration during pathogenesis.

Innovative Enzymatic Approach to Resolve Homogalacturonans Based on their Methylesterification Pattern

Three series of model homogalacturonans (HGs) covering a large range of degree of methylesterification (DM) were prepared by chemical and/or enzymatic means. Randomly demethylesterified HGs, HGs containing a few long demethylesterified galacturonic acid stretches, and HGs with numerous but short demethylesterified blocks were recovered. The analysis of the degradation products generated by the action of a purified pectin lyase allowed the definition of two new parameters, the degree of blockiness, and the absolute degree of blockiness of the highly methylesterified stretches (DBMe and DB(abs)Me, respectively). By combining this information with that obtained by the more traditional endopolygalacturonase digestion, the total proportion of degradable zones for a given DM was measured and was shown to permit a clear differentiation of the three types of HG series over a large range of DM. This double enzymatic approach will be of interest to discriminate industrial pectin samples exhibiting different functionalities and to evaluate pectin fine structure dynamics in vivo in the plant cell wall, where pectin plays a key mechanical role.

Rheological properties of Ca2+-gels of partially methylesterified polygalacturonic acid: Effect of “mixed” patterns of methylesterification

Abstract Subjecting homogalacturonan to a combination of processes which modify the extent of pectin's methylesterification results in the generation of distinct patterns of methylesterification on the polymer. In this study, the influence of these “mixed” patterns of methylesterification on the rheological characteristics of Ca 2+ -gels is extensively examined, using partially methylesterified poly- d -galacturonic acid (mPGA) as a model for homogalacturonan. The uncontrolled pattern of methylesterification induced on mPGA was combined with that resulting from subsequent controlled chemical or enzymatic de-esterification. Afterwards, the produced mPGAs were characterised in terms of degree and pattern of methylesterification. The latter structural feature was quantified as “absolute degree of blockiness” through enzymatic fingerprinting. Characterised mPGAs were used for the preparation of gels with various calcium ion (Ca 2+ ) concentrations. All Ca 2+ -pectin systems formed “true” gels with a strong elastic behaviour, but vary in gel stiffness. The gels’ rheological characteristics not only depend on the extent of mPGA de-esterification, but also on the “mixed” patterns of methylesterification. The experimental results also indicate that extremely short non-cooperative junction zones contribute to the stiffness of Ca 2+ -mPGA gels.

Stiffness of Ca 2+-pectin gels: combined effects of degree and pattern of methylesterification for various Ca 2+ concentrations

The influence of the degree and pattern of methylesterification (DM and PM, respectively) on the stiffness of Ca 2+-pectin gels is extensively examined, at various Ca 2+ concentrations. Accordingly, a highly methyl-esterified pectin was selectively de-esterified using NaOH, plant or fungal pectin methylesterase in order to produce series of pectins with varied pattern and broad ranges of methylesterification. The PM was quantified as absolute degree of blockiness (DB abs). Ca 2+-pectin gels were prepared at various Ca 2+ concentrations. Gel stiffness ( G′ at 1 rad/s) was determined and mapped out as a function of DM, DB abs and Ca 2+ concentration. At low Ca 2+ concentrations, G′ depends on polymer’s DM and DB abs. At high Ca 2+ concentrations, a master curve is obtained over a wide range of DM, irrespective of DB abs. Depending on methylesterification pattern, increase of G′ is related not only to an increase in the number of junction zones per pectin chain, but also to an increase in the size of junction zones and the number of dimerised chains occurring in the gels. These results provide a detailed insight into the occurrence of junction zones in Ca 2+-pectin gels.

Isolation and characterization of model homogalacturonans of tailored methylesterification patterns

a b s t r a c t A very highly methylated homogalacturonan (HG) was produced and further used to generate model HG series. HG samples were characterized with respect to their macromolecular features, to their degree and pattern of methyl-esterification, and to their calcium-binding properties. All HG samples were of similar molar mass. Randomly demethylesterified HG samples (B-series) of DM ≥ 40, exhibiting low degree of blockiness (DB), absolute degree of blockiness (DBabs) and calcium binding properties, were classically obtained by alkali deesterification of the very highly methylated “mother” HG. HGs generated by the action of plant pectin-methylesterase (p-PME) on the very highly methylated “mother” HG (P-series) exhibited high DB, DBabs and calcium binding properties, whatever their DM, in consistency with the presence of few long deesterified blocks of GalA. HGs generated by the action of p-PME on “mother” HGs of DM ∼ 80 arising from the B-series (BP-series) exhibited intermediate DB, DBabs and calcium binding properties, in agreement with the presence of numerous but short deesterified blocks of GalA. Manipulating the initial DM of randomly methyl-esterified “mother” HGs prior to enzymatic blockwise demethylesterification allowed the recovery of various HG types with respect to the length and number of demethylesterified GalA stretches.

Effect of de-methylesterification on network development and nature of Ca 2+-pectin gels: Towards understanding structure–function relations of pectin

Pectins of varying degree and pattern of methylesterification were produced through controlled de-esterification of highly esterified citrus pectin, using carrot pectin methylesterase (PME) (P-pectins), Aspergillus aculeatus PME (F-pectins) or sodium hydroxide saponification (C-pectins). Estimation of the degree of methylesterification (DM) and quantification of the pattern of methylester distribution in terms of absolute degree of blockiness (DB abs) enabled the characterisation of pectins. Characterised pectins were used for the preparation of Ca 2+-pectin gels with varying calcium ion (Ca 2+) concentration. The rheological characteristics of produced gels were evaluated by means of small-amplitude oscillatory tests. During gel formation, gel strength was monitored so as to allow assessment of network development. Based on the evaluation of mechanical spectra, the nature of the cured gels was established. Depending on Ca 2+ concentration as well as DM and DB abs, gels prepared from specific C-pectins (48 ≥ DM ≥ 26%; 9 ≤ DB abs ≤ 37%) and F-pectins (64 ≥ DM ≥ 29%; 9 ≤ DB abs ≤ 50%) showed a striking decrease of the gel strength with time, while gel networks produced from other pectins either displayed a continuous increase of the gel strength or verged towards pseudo-equilibrium within the observation time of 5 h. Furthermore, the DM, DB abs and Ca 2+ concentration influenced the evolution of Ca 2+-pectin networks from “structured liquids” to “strong gels”. Based on the experimental results, specific mechanisms of Ca 2+ interactions with pectins were (also) considered.