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Soy protein hydrolysates enhance integral viable cell density (IVCD) and recombinant protein production (Immunoglobulin, IgG) in cell cultures, but their functionality varies from batch-to-batch. This is undesirable since it affects both quantity and characteristics of the recombinant proteins. It is hypothesized that the variability of hydrolysates is due to variations in meal and hydrolysate processing treatments. To study this, hydrolysates were produced from meals heated at 121 °C/0-120 min. The heating decreased free amino acid and reducing monosaccharide contents in meals (0.72-0.27% and 3.3-2.6%) and hydrolysates (14.7-7.1% and 16.9-7.9%). Dry heating introduced large variation in the IVCD ((115-316%), but additional heating in suspension reduced it (131-159%). The decrease in IVCD variation corresponded with decreased variation in carboxymethyl-lysine (CML) and lysinoalanine (LAL) contents. Thus, meal and hydrolysate processing induced substantial variation in hydrolysate functionality. It is therefore critical to establish strict process controls for meal and hydrolysate production to ensure consistency., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Avenanthramides are amides, with a phenylalkenoic acid (PA) and an anthranilic acid (AA) subunit, which are secondary metabolites of oat. Oat seeds were germinated, extracted, and the avenanthramides analysed by a combination of UHPLC with ion trap and high resolution ESI-MS. Typical fragmentation pathways with corresponding diagnostic fragments belonging to the PA and AA subunits were identified and summarised in a decision guideline. Based on these findings 28 unique avenanthramides were annotated in the oat seed(ling) extracts, including the new avenanthramide 6f (with a 4/5-methoxy AA subunit). Avenanthramide content increased by 25 times from seed to seedling. Avenanthramides 2p, 2c, and 2f, which are commonly described as the major avenanthramides, represented less than 20% of the total content in the seedlings. Future quantitative analyses should, therefore, include a wider range of avenanthramides to avoid underestimation of the total avenanthramide content., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The aggregation of α-lactalbumin, β-lactoglobulin and β-casein after heating in dry state was studied in absence and presence of saccharides. In absence of saccharides, differences were observed in the extent of aggregation. Differences between the proteins were mostly due to differences in covalent aggregation. The presence of glucose during the heat treatment of milk proteins significantly increased the extent of aggregation, and decreased differences between proteins. α-Lactalbumin was selected as a model protein for the study of cross-links formed after heat treatment. In the presence of saccharides, these cross-links were found to consist of 36% of disulphide bridges (compared to >75% in the absence of glucose), followed by other cross-links such as lanthionine. Larger saccharides led to a decrease in Maillard induced aggregation; maltotriose actually even inhibited the formation of α-lactalbumin aggregates., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

This paper set out to differentiate the Maillard induced glycation reactivity of individual milk proteins using different saccharides under well-defined reaction conditions. α-Lactalbumin, β-lactoglobulin and β-casein were incubated with mono-, di- and trisaccharides in the dry state under standardised buffered conditions and glycation was expressed relative to the available reactive groups per protein (DG). Protein reactivity, described by the DG max and initial speed of glycation (v), followed the same order for each protein-saccharide incubation: α-lactalbumin > β-lactoglobulin ≫ β-casein. Glycation of whey proteins by different monosaccharides was double that of β-casein. Differences in DG between whey proteins and β-casein decreased with increased saccharide size. A two-fold difference was found for glycation in the presence of the dimers lactose and maltose for β-casein but not for the whey proteins. The percentage of glycated lysines increased with increased lysine to protein size ratio., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Prenylated phenolics from the Fabaceae are promising lead compounds for new antibacterials. Pools enriched in prenylated phenolics were made from lupine, peanut and soybean seedlings. One pool was rich in chain prenylated isoflavones (cIsf), one in chain prenylated stilbenoids (cSti), one in chain prenylated (cPta) and one in ring-closed prenylated pterocarpans (rPta), as characterized by RP-UHPLC-UV-MS. Antibacterial activity of the pools and membrane permeabilization was investigated. Pools showed high antibacterial activity against Listeria monocytogenes: cIsf pool had a minimum inhibitory concentration of 10µg/ml prenylated compounds, followed by cPta pool (25µg/ml) and cSti pool (35µg/ml). Activity against E. coli was found only when the pools were co-administered with an efflux pump inhibitor. The pool enriched in chain prenylated isoflavones permeabilized the bacterial membrane within minutes of exposure, whereas ampicillin did not. Bent conformation and chain prenylation, were molecular features of main prenylated phenolics found in pools with high antibacterial activity., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The legume plant family (Fabaceae) is a potential source of antimicrobial phytochemicals. Molecular diversity in phytochemicals of legume extracts was enhanced by germination and fungal elicitation of seven legume species, as established by RP-UHPLC-UV-MS. The relationship between phytochemical composition, including different types of skeletons and substitutions, and antibacterial properties of extracts was investigated. Extracts rich in prenylated isoflavonoids and stilbenoids showed potent antibacterial activity against Listeria monocytogenes and methicillin-resistant Staphylococcus aureus at concentrations between 0.05 and 0.1% (w/v). Prenylated phenolic compounds were significantly (p<0.01) correlated with the antibacterial properties of the extracts. Furthermore, the position of the prenyl group within the phenolic skeleton also influenced the antibacterial activity. Overall, prenylated phenolics from legume seedlings can serve multiple purposes, e.g. as phytoestrogens they can provide health benefits and as natural antimicrobials they offer preservation of foods., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Berries of Colombian Euterpe oleracea Mart. were analyzed for total phenolic content (TPC), anthocyanin (ACN) content, and antioxidant activity. Additionally, reversed-phase ultra-high performance liquid chromatography with photodiode array detection (RP-UHPLC-PDA) and heated electrospray ionization (HESI) multistage mass spectrometry (MS(n)) were used to determine the composition of phenolic compounds. Anthocyanin content was 0.57±0.39mg cyanidin-3-glucoside/g fresh weight (FW) and TPC was 6.07±2.17mg gallic acid equivalent (GAE)/g FW. The ABTS radical scavenging activity was 3.1±1.3μmol Trolox equivalents (TE)/100g FW, whereas the DPPH value was 2693.1±332.8μmol TE/100g FW. Overall, results show that Colombian açai has a more diverse polyphenolic profile and higher antioxidant activity than Brazilian açai. This information could be useful in authentification procedures to differentiate Brazilian açai from Colombian açai when used as an alternative for the supply of this fruit during the time of shortage in Brazil., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

In this study, we analyze 31 transgenic lines and their respective untransformed background lines to determine the transgene effects on targeted structures including the pectin components rhamnogalacturonan I (RG-I) and homogalacturonan (HG), neutral side chains (galactan/arabinangalactan), acetylation of pectin, and cellulose level. Modification arising from the pectin backbone- or pectin side chain transgenic lines either increased or decreased the HG:RG-I ratio, side chain length, and methyl esterification of pectin in the tuber cell wall. The pectin esterification transgenic line exhibited only limited side effects. The cellulose level-targeting transgenic lines yielded an unexpectedly high HG:RG-I ratio and longer pectic side chains. These results clearly demonstrate that in effects of a transgene are not restricted to the direct activity of the targeted enzyme but have consequences for the structure of the cell wall matrix. Analysis of whole cell wall structure is therefore necessary to assess the complete effect, direct and indirect, of a transgene., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Enzyme hydrolysed (hemi-)celluloses from oil palm empty fruit bunches (EFBs) are a source for production of bio-fuels or chemicals. In this study, after either peracetic acid delignification or alkaline extraction, EFB hemicellulose structures were described, aided by xylanase hydrolysis. Delignification of EFB facilitated the hydrolysis of EFB-xylan by a pure endo-β-1,4-xylanase. Up to 91% (w/w) of the non-extracted xylan in the delignified EFB was hydrolysed compared to less than 4% (w/w) of that in untreated EFB. Alkaline extraction of EFB, without prior delignification, yielded only 50% of the xylan. The xylan obtained was hydrolysed only for 40% by the endo-xylanase used. Hence, delignification alone outperformed alkaline extraction as pretreatment for enzymatic fingerprinting of EFB xylans. From the analysis of the oligosaccharide-fingerprint of the delignified endo-xylanase hydrolysed EFB xylan, the structure was proposed as acetylated 4-O-methylglucuronoarabinoxylan., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Uridine diphosphate (UDP)-glucose 4-epimerase (UGE) catalyzes the conversion of UDP-glucose to UDP-galactose. Cell wall materials from the cv. Kardal (wild-type, background) and two UGE transgenic lines (UGE 45-1 and UGE 51-16) were isolated and fractionated. The galactose (Gal) content (mg/100g tuber) from UGE 45-1 transgenic line was 38% higher than that of wild-type, and resulted in longer pectin side chains. The Gal content present in UGE 51-16 was 17% lower than that of wild-type, although most pectin populations maintained the same level of Gal. Both UGE transgenic lines showed unexpectedly a decrease in acetylation and an increase in methyl-esterification of pectin. Both UGE transgenic lines showed similar proportions of homogalacturonan and rhamnogalacturonan I within pectin backbone as the wild-type, except for the calcium-bound pectin fraction exhibiting relatively less rhamnogalacturonan I. Next to pectin modification, xyloglucan populations from both transgenic lines were altered resulting in different XSGG and XXGG proportion in comparison to wild-type., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Genes encoding pectic enzymes were introduced to wild-type potato Karnico. Cell wall materials were extracted from Karnico and transgenic lines expressing β-galactosidase (β-Gal-14 mutant) or rhamnogalacturonan lyase (RGL-18 mutant). After sequential extraction, β-Gal-14 hot buffer-soluble solids (HBSS) of pectin contained 54% less galactose than Karnico HBSS, representing shorter galactan side chains. The individual pectin populations of β-Gal-14 HBSS showed different modifications extended to the two sub-populations as obtained by ion-exchange chromatography. Compared to wild-type, RGL-18 HBSS contained 27% more galacturonic acid and 55% less Gal on fresh potato weight basis, which was due to the removal of galactan-rich rhamnogalacturonan I (RG-I) segments. All pectin populations of RGL-18 showed consistently low levels of RG-I segments. Transgenic modification showed side effects on the methyl-esterification and acetyl substitution of RGL-18 HBSS (DM=53, DA=21), but not of the β-Gal-14 HBSS in comparison to wild-type (DM=29, DA=54)., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Transformation from green tea leaves into black tea involves oxidation of catechins into theaflavins and other complex phenolics by endogenous enzymes in tea leaves. By employing tyrosinase and laccase, both from Agaricus bisporus, on green tea catechins, the oxidation process was directed towards a higher theaflavins content, which is considered an important quality parameter in tea. The main tea catechins were incubated with tyrosinase and laccase, and product formation was monitored by RP-UHPLC-PDA-ESI-MS. The kind of catechin, their substitution with a galloyl group, and the type of oxidase used were important factors determining theaflavin concentrations. In particular, incubation of epicatechin with epigallocatechin with tyrosinase gave a high, stable theaflavin content. In a green tea extract, tyrosinase increased the proportion of theaflavins by twofold compared to black tea. Laccase mainly formed insoluble complexes. Our results indicate that the phenolic profile of tea can be modulated by using commercially available exogenous oxidases., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Seeds from seven species of tribe Phaseoleae, i.e. Phaseolus, Vigna, Lablab and Psophocarpus, were investigated for inducibility of isoflavonoids by germination with or without subsequent elicitation with Rhizopus oryzae. Germination alone poorly induced isoflavonoid production (in the range of 0.2-0.7 mg representative compound equivalents (RCE)/g DW), whereas application of Rhizopus onto the seedlings increased the isoflavonoid content considerably (in the range of 0.5-3.3 mg RCE/g DW). The inducibility of different isoflavonoid subclasses in seedlings with Rhizopus varied per species. Isoflavones and isoflavanones were mainly found in elicited seedlings of Phaseolus, Vigna and Lablab, whereas pterocarpans were mainly observed in those of Psophocarpus. Despite their phylogenetic relatedness, the seeds of various species within Phaseoleae appeared to respond differently towards elicitation by Rhizopus during germination. The kind of molecules induced followed the phylogenetic relationship of the various species, but their amounts induced during germination, alone or combined with elicitation, did not., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Fatty acid esterification, common in naturally occurring astaxanthin, has been suggested to influence both colour stability and degradation of all-trans-astaxanthin. Therefore, astaxanthin stability was studied as influenced by monoesterification and diesterification with palmitate. Increased esterification decelerated degradation of all-trans-astaxanthin (RP-UHPLC-PDA), whereas, it had no influence on colour loss over time (spectrophotometry). This difference might be explained by the observation that palmitate esterification influenced the cis-trans equilibrium. Free astaxanthin produced larger amounts of 9-cis isomer whereas monopalmitate esterification resulted in increased 13-cis isomerization. The molar ratios of 9-cis:13-cis after 60min were 1:1.7 (free), 1:4.8 (monopalmitate) and 1:2.6 (dipalmitate). The formation of 9-cis astaxanthin, with its higher molar extinction coefficient than that of all-trans-astaxanthin, might compensate for colour loss induced by conjugated double bond cleavage. As such, it was concluded that spectrophotometry is not an accurate measure of the degradation of the all-trans-astaxanthin molecule., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The distributions of phosphorus and hydroxypropyl groups within granules of cross-linked and hydroxypropylated sweet potato starches were investigated. Chemical surface peeling of starch granules was performed after sieving of native and modified starches into large-size (diameter ≥ 20 μm) and small-size (diameter < 20 μm) fractions. Starch granules were surface gelatinized in a 4M calcium chloride solution at different levels. After the peeling step, the remaining starch granules were analysed for the content of phosphorus and hydroxypropyl groups. The phosphorus level of the parental starch gradually decreased from periphery to core of the granules. The increase in phosphorus content after cross-linking in periphery was higher than that in core. The subsequent hydroxypropylation reaction resulted in lower phosphate levels. Hydroxypropylation resulted in a gradient of hydroxypropyl group concentration from periphery to core. Cross-linking prior to the hydroxypropylation resulted in lower levels of hydroxypropyl groups and less pronounced differences between periphery and core., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The ability of Agaricus bisporus to degrade xylan in wheat straw based compost during mushroom formation is unclear. In this paper, xylan was extracted from the compost with water, 1M and 4M alkali. Over the phases analyzed, the remaining xylan was increasingly substituted with (4-O-methyl-)glucuronic acid and arabinosyl residues, both one and two arabinosyl residues per xylosyl residue remained. In the 1M and 4M KOH soluble solids of spent compost, 33 and 49 out of 100 xylosyl residues, respectively, were substituted. The accumulation of glucuronic acid substituents matched with the analysis that the two A. bisporus genes encoding for α-glucuronidase activity (both GH115) were not expressed in the A. bisporus mycelium in the compost during fruiting. Also, in a maximum likelihood tree it was shown that it is not likely that A. bisporus possesses genes encoding for the activity to remove arabinose from xylosyl residues having two arabinosyl residues., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Sweet potato starch was cross-linked using sodium trimetaphosphate and hydroxypropylated using propylene oxide. The level and position of phosphorus and hydroxypropyl groups within cross-linked and hydroxypropylated sweet potato starch was investigated by phosphorus and proton nuclear magnetic resonance spectroscopy ((31)P, (1)H NMR). The cross-linking reaction produced monostarch monophosphate and distarch monophosphate in a molar ratio of 1:1.03, indicating that only half of the introduced phosphorus resulted in a possible cross-link. One cross-link per approximately 2900 glucose residues was found. Phosphorylation leading to monostarch monophosphate mainly occurred at O-3 and O-6 (ratio 1:1). It was inferred that the majority of the cross-links formed in distarch monophosphate were between two glucose residues positioned in different starch chains, while a minor part of the cross-links may be formed between two glucose residues within the same starch chain. Hydroxypropylation under alkaline conditions resulted in the formation of intra-molecular phosphorus cross-links, subsequent hydroxypropylation following cross-linking lowered both the level of intra- and inter-molecular cross-linking. Using (1)H NMR the molar substitution of hydroxypropylation was determined to be 0.155-0.165. The hydroxypropylation predominantly occurred at O-2 (61%), and the level of substitution at O-6 (21%) was slightly higher than that at O-3 (17%). In dual modified starch, the preceding cross-linking procedure resulted in a slightly lower level of hydroxypropylation, where the substitution at O-6 decreased more compared to the substitution at O-2 and O-3., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The application of phycobiliproteins, e.g. blue C-phycocyanin, as natural water-soluble food colourants is emerging. The chromophore of these proteins comprises a number of tetrapyrroles (or phycocyanobilins), which have an extensive conjugated system, vulnerable to autoxidation. To assess the autoxidation products, a simplified model system was used in which the free tetrapyrrole biliverdin, instead of phycobiliprotein, was subjected to light-accelerated autoxidation. Degradation products of biliverdin were subsequently annotated by reversed-phase ultra-high performance liquid chromatography with a photo diode array and positive mode in-line electrospray ionization mass spectrometry. To facilitate the analysis of degradation products, autoxidation of the three methine bridges in biliverdin was mimicked in silico. It was found that both the peripheral and central methine bridges of biliverdin were susceptible to light-accelerated autoxidation. Scission products tentatively annotated with MS(2) and MS(3) were propionic acid-containing pyrroles. From this, it can be speculated that tetrapyrroles attached to phycobiliproteins are susceptible to autoxidative degradation., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

This study evaluated the intestinal degradability of alginate during 74 days intake in pigs as models for humans. Diets contained pregelatinized starch, retrograded starch, alginate, or a mix of retrograded starch and alginate. Faeces were collected on day 1, 3, 7, 14, 39 and 74. Clear trends in intestinal alginate degradation were observed. Up to day 39, the total tract digestibility of alginate was limited (0.52 ± 0.10), and was lower with the inclusion of retrograded starch in the diet (0.34 ± 0.02). More than 90% of the faecal alginate was insoluble in water, which may explain the low digestibility of the alginate. The digestibility of mannuronic acid (M) was 2-3 times higher than that of guluronic acid (G). The changes of G:M ratio and the relative amounts of alginate oligosaccharides between day 39 and 74 indicated that the microbiota needed more than 39 days to adapt to alginate. This study demonstrated that in-depth analyses of dietary fibres are valuable in understanding the fate of the dietary fibres in the large intestine as it was shown that degradation of a dietary fibre depends not only on the properties of the fibre itself, but also on the other dietary fibres present in the diet and the adaptation time., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

A gene encoding a putative pectinesterase from Bacillus licheniformis DSM13 was cloned and expressed in Escherichia coli. The resulting recombinant enzyme (BliPME) was purified and characterized as a pectin methylesterase. The enzyme showed maximum activity at pH 8.0 and 50°C. BliPME is able to release up to 100% of the methylesters from lime pectin (DM 34-76→DM 0) and up to 73% of all methylesters from SBPs (DM 30-73→DM 14). BliPME efficiently de-methylesterifies lemon pectins and SBPs in a blockwise manner and is quite tolerant towards the acetyl groups present within the SBPs. Detailed analysis of the BliPME-modified pectins using HILIC-MSn and the classical calcium reactivity measurement showed that the enzyme generates pectins with low methylesterification (lime and SBP) and high acetyl content (SBP) while creating blocks of nonmethylesterified galacturonic acid residues. The high activity of BliPME towards highly methylesterified and acetylated pectins makes this novel esterase more efficient in removing methylesters from highly esterified beet pectin compared to other PMEs, e.g. Aspergillus niger PME., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

During the industrial extraction of starch from potatoes (Seresta), some starch remains within undisrupted potato cells in the fibrous side-stream. The aim of this study was to investigate if enzymatic degradation of cell wall polysaccharides (CWPs) can enhance starch recovery and lower the water holding capacity (WHC) of the "fibre" fraction. The use of a pectinase-rich preparation recovered 58% of the starch present in the "fibre" fraction. Also, the "fibre" fraction retained only 40% of the water present in the non-enzyme treated "fibre". This was caused by the degradation of pectins, in particular arabinogalactan side chains calculated as the sum of galactosyl and arabinosyl residues., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Screening of eight carbohydrate acetyl esterases for their activity towards xanthan resulted in the recognition of one active esterase. AXE3, a CAZy family CE1 acetyl xylan esterase originating from Myceliophthora thermophila C1, removed 31% of all acetyl groups present in xanthan after a 48 h incubation. AXE3 activity towards xanthan was only observed when xanthan molecules were in the disordered conformation. Optimal performance towards xanthan was observed at 53 °C in the complete absence of salt, a condition favouring the disordered conformation. AXE3-deacetylated xanthan was hydrolyzed using cellulases and analyzed for its repeating units using UPLC-HILIC-ELSD/ESI-MS. This showed that AXE3 specifically removes the acetyl groups positioned on the inner mannose and that acetyl groups positioned on the outer mannose are not removed at all. After a prolonged incubation at optimal conditions, 57% of all acetyl groups, representing 70% of all acetyl groups on the inner mannose units, were hydrolyzed., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Interactions of Type A and B flavan-3-ol dimers (procyanidins) and several monomeric flavan-3-ols, with α-casein and β-casein, were investigated. Binding affinities measured were related to the ligands structure, including several properties (e.g. intrinsic flexibility (number of rotatable bonds) and hydrophobicity), and to the amino-acid composition of the caseins. A monomeric flavan-3-ol esterified with gallic acid (EGCG) had a five to ten times higher affinity to caseins compared to the non-galloylated dimeric flavan-3-ols. In this case, the larger number of rotatable bonds in EGCG might be accountable for this difference. Comparing flavan-3-ol dimers, intrinsic flexibility did not consistently promote interactions, as procyanidin A1 displayed a higher affinity to α-casein than the supposedly more flexible B-type dimers investigated. Despite its higher content of proline, compared to α-casein, β-casein did not always have a higher affinity for the ligands investigated (e.g. no interaction with procyanidin A1 detected). These results suggest that more factors than proline content and the number of proline repeats govern phenolic-casein interactions., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

A two-step enzymatic fingerprinting method was introduced to analyze a highly methylesterified and acetylated sugar beet pectin having a degree of methylesterification (DM) of 62 and acetylation of 30. A cocktail of pectolytic enzymes, including endo-polygalacturonase II (endo-PGII) and pectin lyase (PL), was used for the first digestion. The endo-PGII and PL resistant pectin fragments were isolated and subjected to a second digestion using fungal pectin methylesterase and endo-PGII. After the two sequential digestions, 78% of the total GalA residues present in the parental pectin were recovered as mono- and oligomers, which were used to quantitatively describe the parental SBP. For this reason, the descriptive parameters degree of blockiness (DBabs), degree of hydrolysis by PG (DHPG) and degree of hydrolysis by PL (DHPL) were established for both digestions. The first digestion revealed the presence of short blocks of nonesterified GalA residues and blocks of partly methylesterified and acetylated GalA residues in the parental SBP, in addition to blocks of highly methylesterified and acetylated GalA residues. The second digestion revealed the presence of blocks of methylesterified, partly methylesterified and/or acetylated GalA residues in a sequence not to be degradable by neither endo-PGII nor by PL. The acetyl groups were present in an blockwise manner. Application of the method to two differently prepared DM 50 SBPs showed that the two pectins differ in the ratio of blocks of nonesterified and blocks of partly methylesterified and acetylated GalA residues., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

A recombinant acetylesterase from Bacillus licheniformis DSM13, belonging to carbohydrate esterase family 12, was purified and biochemically characterized. The purified enzyme, termed BliPAE, was capable of deacetylating acetylated pectins, e.g. sugar beet pectin (SBP). Contrary to its provisional annotation as rhamnogalacturonan acetylesterase, the enzyme specifically removed acetyl groups from the homogalacturonan region classifying it as a PAE. The recombinant enzyme has a molecular mass of 26.7 kDa and shows optimal activity at pH 8.0 and 50°C. It is stable in the range pH 5.0-7.0 and below 50°C. Methylesterification of the galacturonic acid (GalA) moieties reduces the deacetylation efficacy of BliPAE. The enzyme efficiently removes acetyl groups from SBPs with low degree of methylesterification (DM) 9-30, releasing about 75% of the acetyl groups present in the homogalacturonan. Furthermore, (1)H NMR of polymer and LC-HILIC-MS(n) after endo-PGII and PL degradation were used to structurally characterize the BliPAE-modified pectins. The results show that BliPAE removes acetyl groups specifically when substituted at the O-3 position of GalA moieties., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Xanthans occurring in different levels of disordered conformation were enzymatically hydrolyzed to their six pentamer repeating units (RUs). The RUs present in the enzyme digests were analyzed using LC-MS. As only disordered xanthan segments are degraded by cellulases, the influence of the six different RUs on the transitional behavior of xanthan could be studied. The results indicate that especially xanthan segments rich in RUs that are acetylated on the outer mannose unit stabilize the xanthan conformation. Acetylation of the inner mannose did not show to have a clear stabilizing effect on the xanthan conformation. As the enzymatic release of pyruvylated RUs gradually increased with increasing levels of disordered xanthan segments, it is concluded that the distribution of these RUs is random. On the contrary, single and double acetylated RUs are distributed over specific xanthan segments as these RUs were instantly hydrolyzed., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Berries and leaves from six varieties of Carpathians' sea buckthorn (Hippophae rhamnoides L., ssp. Carpatica) were analysed for their carotenoid composition (free and esterified) using a combination of HPLC-PAD, GC-MS and UHPLC-PAD-ESI-MS techniques. GC-MS techniques revealed the fatty acid profile specific for each berry variety, while targeted UHPLC-MS analysis identified the fatty acids involved in carotenoids esterification: palmitic (C16:0), myristic (C14:0) and stearic (C18:0). Total carotenoid content varied between 53 and 97 mg/100g dry weight in berries, and between 3.5 and 4.2mg/100g DW in leaves. The carotenoid di-esters represented the main fraction among berry varieties having zeaxanthin di-palmitate as major compound, while leaves contained only free carotenoids like lutein, β-carotene, violaxanthin and neoxanthin. Principal component analysis identified the suitable carotenoid biomarkers characteristic for the Carpathians' sea buckthorn from Romania with contribution to their taxonomic classification and authenticity recognition., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Changes of plant cell wall carbohydrate structures occurring during the process to make suitable compost for growth of Agaricus bisporus are unknown. In this paper, composition and carbohydrate structures in compost samples collected during composting and mycelium growth were analyzed. Furthermore, different extracts of compost samples were prepared with water, 1M and 4M alkali and analyzed. At the beginning of composting, 34% and after 16 days of mycelium growth 27% of dry matter was carbohydrates. Carbohydrate composition analysis showed that mainly cellulose and poorly substituted xylan chains with similar amounts and ratios of xylan building blocks were present in all phases studied. Nevertheless, xylan solubility increased 20% over the period of mycelium growth indicating partial degradation of xylan backbone. Apparently, degradation of carbohydrates occurred over the process studied by both bacteria and fungi, mainly having an effect on xylan-chain length and solubility., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Enzymatic fingerprinting was applied to sugar beet pectins (SBPs) modified by either plant or fungal pectin methyl esterases and alkali catalyzed de-esterification to reveal the ester distributions over the pectin backbone. A simultaneous pectin lyase (PL) treatment to the commonly used endo-polygalacturonase (endo-PG) degradation showed to be effective in degrading both high and low methylesterified and/or acetylated homogalaturonan regions of SBP simultaneously. Using LC-HILIC-MS/ELSD, we studied in detail all the diagnostic oligomers present, enabling us to discriminate between differently prepared sugar beet pectins having various levels of methylesterification and acetylation. Furthermore, distinction between commercially extracted and de-esterified sugar beet pectin having different patterns of substitution was achieved by using novel descriptive pectin parameters. In addition to DBabs approach for nonmethylesterified sequences degradable by endo-PG, the "degree of hydrolysis" (DHPG) representing all partially saturated methylesterified and/or acetylated galacturonic acid (GalA) moieties was introduced as a new parameter. Consequently, the description DHPL has been introduced to quantify all esterified unsaturated GalA oligomers., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

To enable structural characteristics of individual cell wall polysaccharides from rapeseed (Brassica napus) meal (RSM) to be studied, polysaccharide fractions were sequentially extracted. Fractions were analysed for their carbohydrate (linkage) composition and polysaccharide structures were also studied by enzymatic fingerprinting. The RSM fractions analysed contained pectic polysaccharides: homogalacturonan in which 60% of the galacturonic acid residues are methyl-esterified, arabinan branched at the O-2 position and arabinogalactan mainly type II. This differs from characteristics previously reported for Brassica campestris meal, another rapeseed cultivar. Also, in the alkali extracts hemicelluloses were analysed as xyloglucan both of the XXGG- and XXXG-type decorated with galactosyl, fucosyl and arabinosyl residues, and as xylan with O-methyl-uronic acid attached. The final residue after extraction still contained xyloglucan and remaining (pectic) polysaccharides next to cellulose, showing that the cell wall matrix of RSM is very strongly interconnected., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Five xanthans were hydrolyzed to their repeating units using cellulases. Hydrophilic interaction chromatography with online electrospray ionization ion trap mass spectrometry and evaporative light scattering detection was used to analyze the oligomers released. It was concluded that six different pentamer repeating units (RUs) exists within a xanthan sample. The most abundant RU shows acetylation on the inner mannose and pyruvylation on the outer mannose. The second most abundant RU shows acetylation on both the inner and the outer mannose. It becomes clear that more variations in the xanthan structure exist than generally recognized. Comparison of five different xanthan samples revealed that, although the molecular composition of xanthan samples can be exactly the same, the ratio in which the RUs occur can differ significantly. It is, therefore, concluded that xanthan samples should be characterized for both, their molecular composition and the relative abundance of the RUs present., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Differently modified xanthans, varying in degree of acetylation and/or pyruvylation were incubated with the experimental cellulase mixture C1-G1 from Myceliophthora thermophila C1. The ionic strength and/or temperature of the xanthan solutions were varied, to obtain different xanthan conformations. The exact conformation at the selected incubation conditions was determined by circular dichroism. The xanthan degradation was analyzed by size exclusion chromatography. It was shown that at a fixed xanthan conformation, the backbone degradation by cellulases is equal for each type of xanthan. Complete backbone degradation is only obtained at a fully disordered conformation, indicating that only the secondary xanthan structure influences the final degree of hydrolysis by cellulases. It is thereby shown that, independently on the degree of substitution, xanthan can be completely hydrolyzed to oligosaccharides. These oligosaccharides can be used to further investigate the primary structure of different xanthans and to correlate the molecular structure to the xanthan functionalities., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

In potato tuber, caffeic acid (the predominant hydroxycinnamic acid (HCA)), its conjugates (HCAcs; i.e. chlorogenic acid (ChA), crypto-ChA, and neo-ChA), and anthocyanin-linked HCAs have been extensively described in the literature. In contrast, only little information is available on the occurrence of other HCAcs and didydrohydroxycinnamic acid conjugates (DHCAcs). Fifteen Colombian potato cultivars were screened for these less commonly described conjugates by reversed-phase ultrahigh performance liquid chromatography coupled to a diode array detector and a heated electrospray ionisation mass spectrometer. A total of 62 HCAs/HCAcs/DHCAcs were found in extracts from peel and flesh. Among them, only twelve compounds were common to all cultivars in both peel and flesh. The less commonly described compounds accounted for 7.1-20.1% w/w of the total amount of HCAs/HCAcs/DHCAcs in whole tubers, highlighting their contribution to the total phenolic profile of potato tubers. Among all cultivars, the abundance (mg/100 g DW whole tuber) of neo-ChA (0.8-7.4) ranged in similar quantities as the less commonly reported feruloyl octopamine (1.2-5.2), 5-O-feruloyl quinic acid (0.1-7.5), cis-ChA (1.1-2.2), caffeoyl putrescine (0.6-2.5), sinapoyl hexose (0.1-1.8), N(1),N(14)-bis-(dihydrocaffeoyl) spermine (0.2-1.7), N(1),N(10)-bis-(dihydrocaffeoyl) spermidine (1.1-2.6), and N(1),N(5),N(14)-tris-(dihydrocaffeoyl) spermine (trace - 11.1)., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Pressed potato fibre (PPF) has a high water holding capacity (WHC) affecting its processing as an animal feed. The aim of this study was to characterize cell wall polysaccharides (CWPs) in PPF and investigate their WHC. This was done via sequential extractions. Half of all CWPs were recovered in the hot buffer soluble solids extract as pectins (uronic acid and rhamnose) and galactans wherein most pectins (76%) from PPF were water soluble. Most likely, the network of CWPs is loosened during processing of potatoes. PPF showed a WHC of 7.4 expressed as the amount of water held per g of dry matter (mL/g). Reconstituting hot buffer soluble solids with buffer insoluble solids in water gave a WHC comparable to that of PPF. Removal of alkali soluble solids, which mainly comprised xyloglucans, lowered the WHC of the final residue. The results indicated that interactions between CWPs could affect the WHC of PPF., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

To investigate the effect of resistant starch to the degradation of other non-starch polysaccharides (NSPs) in the large intestine of pigs, two groups of pigs were fed either a diet containing digestible starch (DS) or a diet containing resistant starch (RS). Both diets contained NSPs from wheat and barley. Digesta from different parts of the large intestine were collected and analysed for sugar composition and carbohydrate-degrading-enzyme activities. Resistant starch, as well as β-glucans and soluble arabinoxylan, was utilised mainly in the caecum. The utilisation of β-glucans and soluble arabinoxylan in the caecum was higher in DS-fed pigs than in RS-fed pigs. Analyses on carbohydrate-degrading-enzyme activities demonstrated that microbial enzyme production was stimulated according to the diet composition, and the enzyme profile throughout the large intestine of RS-fed pigs indicated that the presence of resistant starch shifted the utilisation of NSPs to more distal parts of the colon., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The combined action of endo-polygalacturonase (endo-PGII), pectin lyase (PL), pectin methyl esterase (fungal PME) and RG-I degrading enzymes enabled the extended degradation of methylesterified and acetylated sugar beet pectins (SBPs). The released oligomers were separated, identified and quantified using hydrophilic interaction liquid chromatography (HILIC) with online electrospray ionization ion trap mass spectrometry (ESI-IT-MS(n)) and evaporative light scattering detection (ELSD). By MS(n), the structures of galacturonic acid (GalA) oligomers having an acetyl group in the O-2 and/or O-3 positions eluting from the HILIC column were elucidated. The presence of methylesterified and/or acetylated galacturonic acid units within an oligomer reduced the interaction with the HILIC column significantly compared to the unsubstituted GalA oligomers. The HILIC column enables a good separation of most oligomers present in the digest. The use of ELSD to quantify oligogalacturonides was validated using pure GalA standards and the signal was found to be independent of the chemical structure of the oligomer being detected. The combination of chromatographic and enzymatic strategies enables to distinguish SBPs having different methylesters and acetyl group distribution., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Ber e 1, a major allergen from Brazil nuts, is very stable to in vitro peptic digestion. As heat-induced denaturation may affect protein digestibility, the denaturation behaviour of Ber e 1 was investigated. The denaturation temperature of Ber e 1 varies from approximately 80-110°C, depending on the pH. Upon heating above its denaturation temperature at pH 7.0, the protein partly forms insoluble aggregates and partly dissociates into its polypeptides, whereas heating at pH 5.0 does neither induce aggregation, nor dissociation of the protein. The denaturation temperature of approximately 110°C at pH values corresponding to the general pH values of foods (pH 5-7) is very high and is expected to be even higher in Brazil nuts themselves. As a result, it is unlikely that heat processing causes the denaturation of all Ber e 1 present in food products. Consequently, the allergen is assumed to be consumed (mainly) in its native form, having a high stability towards pepsin digestion., (Copyright © 2008 Elsevier Ltd. All rights reserved.)