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Pectin depolymerase is widely utilized in various industrial sectors. However, the traditional methods for determining its enzymatic activity have limitations, such as cumbersome operations and a significant impact of enzyme solution dilution ratios on activity. The 3-methyl-2-benzothiazolinone hydrazone (MBTH) method can be employed to address these issues, but pectin precipitation and strong background commonly arise in this method. We have successfully overcome these challenges by employing a low-temperature and high-alkaline environment, and further optimized the reagent compositions and detection wavelength to improve the method. Consequently, enzyme hydrolysis follows a zero-order reaction within 60 min, which is helpful for the endpoint measurement of pectinase activity. The developed calibration curve for pectinase concentration and hydrolysis rate demonstrates linearity (R 2  = 0.9945) within the range of 2.5-15.8 mU/mL of pectinase. This method exhibits high sensitivity, accuracy, and stability, making it suitable for routine determination of pectin depolymerase activity in research and applications., 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 © 2024. Published by Elsevier Ltd.)

An energy-intensive and chemical-consuming pretreatment of bamboo is often required to develop its high-performance composites. This study is to evaluate a fungal and enzymatic pretreatment as a sustainable surface modification approach towards high-strength bamboo biocomposites based on D. sinicus. Extracellular enzyme activity analysis suggested that the lignin oxidative enzyme and pectinase secreted by the white-rot fungus T. versicolor achieved higher reaction than the hydrolytic enzymes throughout the pretreatment. Chemical structure analysis revealed that lignin and hemicellulose were partially depolymerised and/or removed during the pretreatment, leading to a relatively higher cellulose content in D. sinicus. The analysis of physical property and microstructure suggested that the surface wettability and porosity of pretreated D. sinicus were significantly improved, facilitating the spreading and penetration of adhesive resins, which subsequently contributed to the superior interfacial bonding of the biocomposites. The tensile strength of the biocomposites increased from 195.8 MPa to 245.8 MPa and the bonding strength increased from 4.9 MPa to 6.3 MPa after the pretreatment, which were 1.26 times and 1.28 times those of the untreated biocomposite respectively. The results proved that the fungal and enzymatic pretreatment is a promising, eco-friendly and efficient surface modification approach for the preparation of high-strength lignocellulosic biocomposites., 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 © 2024 Elsevier B.V. All rights reserved.)

Introduction: Pectin-oligosaccharides (POS) serve diverse purposes as a food ingredient, antimicrobial and biostimulant in plants, and their functionality is linked to the degree of esterification. Grape and broccoli wastes emerge as environmentally friendly alternatives to obtaining pectin, serving as a sustainable source to producing POS. For example, microwaves have proven to be an effective and sustainable method to extract polysaccharides from plant matrices., Objective: This work aims to use grape and broccoli wastes as alternative sources for obtaining pectin by microwave-assisted extraction and biotransformation into POS, which possess biological properties., Material and Methods: The extraction conditions were identified at a power of 400 W, 300 s for the extraction of pectin from grape pomace and broccoli waste. Biotransformation of pectins into POS, using commercial enzyme preparations (Viscozyme L and Pectinase). Characterisation was carried out by Fourier-transform infrared spectroscopy, thermogravimetric analysis, and scanning electron microscopy., Results: Physicochemical analysis indicated grape pomace and broccoli waste pectins had galacturonic acid content of 63.81 ± 1.67 and 40.83 ± 2.85 mg 100 mg -1 , low degree of esterification of 34.89% and 16.22%, respectively. Biotransformation of pectins into POS resulted in a 20% hydrolysis rate. The main enzymatic activity was polygalacturonase for the degradation of the main structure of the pectin., Conclusion: Production of POS from agro-industrial wastes by emerging technologies, such as the combined use of microwave-assisted extraction and enzymatic processes, represents an alternative method for the generation of bioactive compounds with distinctive properties suitable for different applications of interest., (© 2024 John Wiley & Sons Ltd.)

To investigate the structural characteristics of cell wall pectic polysaccharides from wampee, water soluble pectin (WSP), chelator-soluble pectin (CSP) and sodium carbonate-soluble pectin (SSP) were purified. And the inhibitory effects of wampee polyphenol (WPP) on pectinase when these cell wall pectic polysaccharides were used as substrates were also explored. Purified WSP (namely PWSP) had the lowest molecular weight (8.47 × 10 5  Da) and the highest GalA content (33.43%). While purified CSP (called PCSP) and SSP contained more abundant rhamnogalacturonan I side chains. All of them were low-methoxy pectin (DE < 50%). Enzyme activity and kinetics analysis showed that the inhibition of pectinase by wampee polyphenol was reversible and mixed type. When SSP was used as the substrate, WPP had the strongest inhibition (IC 50  = 1.96 ± 0.06 mg/mL) on pectinase. Fluorescence quenching results indicated that WPP inhibited enzyme activity by interacting with substrates and enzymes. Therefore, WPP has the application potential in controlling softening of fruits and vegetables., 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 © 2024 Elsevier Ltd. All rights reserved.)

The prebiotic capacity of Pectin Oligosaccharides (POS) is influenced by structural factors such as molecular size, composition, and degree of esterification, which affect their interaction with the gut microbiota. While existing literature has predominantly examined POS derived from apple and citrus pectins, the extrapolation of these findings to other pectin sources remains complex due to variations in their composition. This study focused on obtaining POS with prebiotic potential from pisco grape pomace through controlled enzymatic hydrolysis, resulting in three molecular size fractions: <3 kDa, 3-10 kDa, and > 10 kDa. The POS fractions were analyzed using FTIR, HPSEC, HPLC, and MALDI-TOF-MS techniques to characterize their physical-chemical properties. Each fraction presented distinct compositions, with the <3 kDa fraction showing a higher concentration of galacturonic acid and glucose, while the >10 kDa fraction was also composed of rhamnose and arabinose. Notably, the <3 kDa fraction supported greater biomass growth of the probiotic strain Lactobacillus casei ATCC 393 compared to the other fractions. In contrast, the non-probiotic strain Escherichia coli ATCC 25922 achieved the lowest biomass with this fraction. Consequently, the <3 kDa POS fraction exhibited the highest prebiotic index. This fraction, composed of oligomers from the rhamnogalacturonan region and arabino-oligosaccharides with a degree of polymerization between two and five, highlights its potential for further research and applications. Therefore, investigating other sources and optimizing extraction conditions could lead to developing novel prebiotic formulations that supply specific probiotic strains for a symbiotic product., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

In this study, heterologous MlPG28B expression was obtained by cloning the Mucor lusitanicus gene screened from a marine environment. The enzyme activity of MlPG28B was maximum at 60 °C, 30 % of the enzyme activity was retained after incubation at 100 °C for 30 min, and enzyme activity was still present after 60 min incubation, one of the best thermostable polygalacturonases characterized until now. The high-purity oligosaccharide standards (DP2-DP7) were prepared with polygalacturonic acid as a substrate. Kinetic parameters showed that MlPG28B at the optimum temperature has a low K m value (3055 ± 1104 mg/L), indicating high substrate affinity. Sequence alignment analysis inferred key residues Cys276, Cys284, Lys107, and Gln237 for MlPG28B thermal stability. Molecular docking and molecular dynamics simulation results indicated that MlPG28B has flexible T1 and T3 loops conducive to substrate recognition, binding, and catalysis and forms a hydrogen bond to the substrate by a highly conserved residue Asn161 in the active-site cleft. Based on site-directed mutation results, the five residues are key in determining MlPG28B thermal stability. Therefore, MlPG28B is a promising candidate for industrial enzymes in feed preparation., 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 © 2024 Elsevier B.V. All rights reserved.)

Pectinases are useful biocatalysts for pectic biomass processing and are extensively used in the food/feed, textile and papermaking industries. Two pectinase genes, a pectate lyase (SbPL1CE8) and a polygalacturonase (SbGH28GH105) were isolated from Segatella bryantii and functionally characterized. Recombinant rSbPL1CE8 was most active against polygalacturonic acid (PGA) and pectin with a 60 % degree of esterification, with k cat /K m values of 721.18 ± 64.77 and 327.02 ± 22.44 mL/s/mg, respectively. Truncated rSbPL1 acted as a mesophilic alkaline pectate lyase, which was highly resistant to inactivation by methanol and ethanol. The rSbPL1CE8 exclusively digested PGA and pectin into unsaturated digalacturonate (uG2), which was further converted into galacturonic acid by rSbGH28GH105. The rSbPL1CE8 was highly effective for saccharification of waste materials from Zea mays, Oryza sativa and Arachis hypogaea processing, and for ramie fiber degumming. This novel pectate lyase has great potential for application in industrial pectic biomass processing., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Individual cells are the smallest units of the plant tissue structure, and their structure and physicochemical properties are essential for whole food processing. In this study, cassava cells were isolated using acid-alkali, hydrothermal, and pectinase methods, and the differences in microstructure and physicochemical properties among the cells, starch, and whole flour were investigated. Cassava cells isolated using pectinase showed intact individual cells with a higher isolation rate and less damage to the cell wall structure and intracellular composition. The presence of cell walls in intact individual cells inhibited the swelling and leaching of starch, resulting in a lower peak viscosity and higher gelatinization temperature than those of starch. The intact cell structure and non-starch composition enhanced the shear resistance of the gels in the sample. Heat treatment led to the gelatinization of intracellular starch and increased the permeability of the cell wall, destroying the physical barrier function of the cell wall; however, the compact cell matrix and non-starch components can inhibit starch hydrolysis. This study suggests that cells isolated using pectinase can be used to investigate the effect of cell walls on the functional properties of intracellular starch in cassava. The isolated cells provide new insights into the cassava industry., 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 © 2024 Elsevier Ltd. All rights reserved.)

Peach ripening accompanying by processing browning exacerbated has not been clarified in perspective of pectin fractions structural variation. This study investigated pectin fractions of water-soluble (WSP), trans-cyclohexane-1,2-diaminetetraacetic acid-soluble (CSP) and sodium carbonate-soluble (NSP) pectin in peach with different ripening times (24, 48, 72 h) and pretreatments (cold shock, CaCl2, the combination) effect on pulp browning in processing. Results showed that extended ripening time increases browning index and rate, along with increased reactive oxygen species levels, polyphenol oxidase and pectinase activities, and the structural of the pectin fractions. Thereinto, weight-average (Mw) and number-average (Mn) molecular weight of WSP increased, its linearity and branching degree decreased, while CSP and NSP showed decreases in Mw, Mn, and branching degree, but increases in linearity, rhamnogalacturonan structure proportion, and bending degree. Browning was correlated positively with Mn, Mw, rhamnogalacturonan proportion of WSP and CSP, linearity and bending degree of NSP, while negatively correlated with Mw of CSP and NSP and esterification degree and branching of all fractions. Thus, maintaining pectin polydispersity and rhamnogalacturonan branching helps protect the system from being quickly browning by compartmentalization effect of hindering the polyphenol substrates and enzymes possibly effective. This study offers insights into controlling browning through pectin fractions modulation., Competing Interests: Declaration of competing interest All authors disclosed no relevant relationships., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The endo-polygalacturonase D (PgaD) from Aspergillus niger JL15 was recombinantly expressed in Escherichia coli BL21, exhibiting an optimal activity at 55 °C and pH 4.0. Hydrolysis products of citrus pectin by recombinant PgaD included galacturonic acid (GalA), digalacturonic acid (GalA2), trigalacturonic acid (GalA3), and tetragalacturonic acid (GalA4). The hydrolysates exhibited significant antioxidant capacity and dose-dependent competitive inhibition of α-glucosidase. GalA2 and GalA3 acted as competitive inhibitors of α-glucosidase, with inhibition constant of 0.0589 mmol.L -1 and 0.6732 mmol.L -1 , respectively. Molecular dynamics (MD) simulations revealed that both GalA2 and GalA3 penetrated the catalytic pocket of α-glucosidase and formed stable hydrogen bonds with key catalytic residues D352 and D215. The binding free energies of GalA2-α-glucosidase and GalA3-α-glucosidase complexes were - 10.3 ± 0.6 kcal·mol -1 and -10.8 ± 0.7 kcal·mol -1 , respectively. These findings might offer new ideas for the development of α-glucosidase inhibitors sourced from citrus pectin, as well as enhance utilization of the renewable plant polysaccharide resources., 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 © 2024 Elsevier Ltd. All rights reserved.)

Banana fibers are a sustainable material with natural mechanical strength and antibacterial properties. These fibers are extracted from the large amount of waste produced by banana pseudo stems annually. However, despite their numerous advantages, their stiffness and rough texture impede their full use in the textile. This research investigates the degumming treatment of banana fibers using enzyme combination and chemical methods to achieve spinnable soft banana fibers. An L9 orthogonal array was used in a Taguchi design of the experiment to optimize the process parameters. For enzyme combination degumming, the experimental setup comprised different quantities of hemicellulase, laccase, amylase, and pectinase; for chemical degumming, varied amounts of sodium hydroxide (NaOH) were used. The results indicate that enzyme-based degumming procedures produce better results than chemical treatments. Optimum enzyme combinations for various fiber qualities were found using the Taguchi design of experiments. These combinations included Hemicellulase 5 %, Laccase 5 %, Amylase 3 %, and Hemicellulase 5 %, Laccase 3 %, Pectinase 5 %. Without degrading the cellulose structure, these ideal enzyme combinations produced fibers with lower lignin content and higher cellulose percentages, moisture content, and tenacity values. By determining the most efficient enzyme combinations and their effects on fiber qualities, the study offers sustainable fiber processing methods for textile grade banana fiber., 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 © 2024 Elsevier B.V. All rights reserved.)

The main purpose of the present study was to determine the effect of associating an optimized ultrasound-assisted extraction (UAE) protocol with enzyme-assisted extraction (EAE) in aqueous media, using the dried berries of Hippophae rhamnoides L. (sea buckthorn) as plant material. A specialized software was used for the determination of potential optimal extraction parameters, leading to the development of four optimized extracts with different characteristics (UAE ± EAE). For these extracts, buffered or non-buffered solutions have been used, with the aim to determine the influence of adjustable pH on extractability. As enzymatic solution, a pectinase, cellulase, and hemicellulase mix (2:1:1) has been applied, acting as pre-treatment for the optimized protocol. The highest extractive yields have been identified for non-buffered extracts, and the E-UAE combination obtained extracts with the highest overall in vitro antioxidant activity. The HPLC-MS n analysis demonstrated a rich composition in different types of isorhamnetin-O-glycosides, as well as some quercetin-O-glycosides, showing a high recovery of specific flavonol-type polyphenolic species. Moreover, we have tentatively identified two flavanols (i.e., catechin and epigallocatechin) and one flavone derivative (i.e., luteolin)., 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 © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Cellulose nanofibers, a sustainable and promising material with widespread applications, exhibit appreciable strength and excellent mechanical and physicochemical properties. The preparation of cellulosic nanofibers from food or agricultural residue is not sustainable. Therefore, this study was designed to use three halophytic plants (Cressa cretica, Phragmites karka, and Suaeda fruticosa) to extract cellulose for the subsequent conversion to cellulosic nanofibers composites. The other extracted biomass components including lignin, hemicellulose, and pectin were also utilized to obtain industrially valuable enzymes. The maximum pectinase (31.56 IU mL -1 ), xylanase (35.21 IU mL -1 ), and laccase (15.89 IU mL -1 ) were produced after the fermentation of extracted pectin, hemicellulose, and lignin from S. fruticosa, P. karka, and C. cretica, respectively. Cellulose was methylated (with a degree of substitution of 2.4) and subsequently converted into a composite using polyvinyl alcohol. Scanning electron microscopy and Fourier-transform infrared spectroscopy confirmed the successful synthesis of the composites. The composites made up of cellulose from C. cretica and S. fruticosa had a high tensile strength (21.5 and 15.2 MPa) and low biodegradability (47.58% and 44.56%, respectively) after dumping for 3 months in soil, as compared with the composite from P. karka (98.79% biodegradability and 4.9 MPa tensile strength). Moreover, all the composites exhibited antibacterial activity against gram-negative bacteria (Escherichia coli and Klebsiella pneumoniae) and gram-positive bacteria (Staphylococcus aureus). Hence, this study emphasizes the possibility for various industrial applications of biomass from halophytic plants., (© 2024 Wiley Periodicals LLC.)

In this study, we analyzed the pectin structure within the pulp of cassava. Cassava pectin, derived from cassava pulp treatment at 120 °C for 90 min, was separated into four fractions (CP-P, CP-SD1, CP-SD2F, and CP-SD2R) based on variations in water solubility, electrical properties, and molecular weights. Sugar composition analysis demonstrated an abundance of homogalacturonan (HG) in CP-P and CP-SD2F, rhamnogalacturonan I (RG-I) in CP-SD2R, and neutral sugars in CP-SD1. Because RG-I possesses a complex structure, we analyzed CP-SD2R using various pectinolytic enzymes. Galactose was the major sugar in CP-SD2R accounting for 49 %, of which 65 % originated from arabinogalactan I, 9 % from galactose and galactooligosaccharides, 5 % from arabinogalactan II, and 11 % from galactoarabinan. Seventy-four percent of arabinose in CP-SD2R was present as galactoarabinan. The methylation (DM) and acetylation (DAc) degrees of cassava pectin were 11 and 15 %, respectively. The HG and RG-I regions exhibited DAc values of 5 and 44 %, respectively, signifying the high DAc of RG-I compared to HG. Information derived from the structural analysis of cassava pectin will enable efficient degradation of pectin and cellulose, leading to the use of cassava pulp as a raw material for biorefineries., Competing Interests: Declaration of competing interest Nozomu Shibata and Meng-I Lin are employees of the Kao Corporation. Tatsuji Sakamoto received funding from Kao Corporation. The authors declare no Declaration of competing interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Polygalacturonases (PGs) can modulate chemistry and mechanical properties of the plant cell wall through the degradation of pectins, one of its major constituents. PGs are largely used in food, beverage, textile, and paper industries to increase processes' performances. To improve the use of PGs, knowledge of their biochemical, structural and functional features is of prime importance. Our study aims at characterizing SmoPG1, a polygalacturonase from Selaginella moellendorffii, that belongs to the lycophytes. Transcription data showed that SmoPG1 was mainly expressed in S. moellendorffii shoots while phylogenetic analyses suggested that SmoPG1 is an exo-PG, which was confirmed by the biochemical characterization following its expression in heterologous system. Indeed, LC-MS/MS oligoprofiling using various pectic substrates identified galacturonic acid (GalA) as the main hydrolysis product. We found that SmoPG1 was most active on polygalacturonic acid (PGA) at pH 5, and that its activity could be modulated by different cations (Ca 2+ , Cu 2+ , Fe 2+ , Mg 2+ , Mn 2+ , Na 2+ , Zn 2+ ). In addition, SmoPG1 was inhibited by green tea catechins, including (-)-epigallocatechin-3-gallate (EGCG). Docking analyses and MD simulations showed in detail amino acids responsible for the SmoPG1-EGCG interaction. Considering its expression yield and activity, SmoPG1 appears as a prime candidate for the industrial production of GalA., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest associated with this work., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Inhibition of galectin-3-mediated interactions by modified citrus pectin (MCP) could affect several rate-limiting steps in cancer metastasis, but the ability of MCP to antagonize galectin-8 function remains unknown. We hypothesized that MCP could bind to galectin-8 in addition to galectin-3. In this study, a combination of gradual ethanol precipitation and DEAE-Sepharose Fast Flow chromatography was used to isolate several fractions from MCP. The ability of these fractions to antagonize galectin-8 function was studied as well as the primary structure and initial structure-function relationship of the major active component MCP-30-3. The results showed that MCP-30-3 (168 kDa) was composed of Gal (13.8%), GalA (63.1%), GlcA (13.0%), and Glc (10.1%). MCP-30-3 could specifically bind to galectin-8, with an MIC value of 0.04 mg mL -1 . After MCP-30-3 was hydrolyzed by β-galactosidase or pectinase, its binding activity was significantly reduced. These results provide new insights into the interaction between MCP structure and galectin function, as well as the potential utility in the development of functional foods.

Pectic polysaccharide is a bioactive ingredient in Chrysanthemum morifolium Ramat. 'Hangbaiju' (CMH), but the high proportion of HG domain limited its use as a prebiotic. In this study, hot water, cellulase-assisted, medium-temperature alkali, and deep eutectic solvent extraction strategies were firstly used to extract pectin from CMH (CMHP). CMHP obtained by cellulase-assisted extraction had high purity and strong ability to promote the proliferation of Bacteroides and mixed probiotics. However, 4 extraction strategies led to general high proportion of HG domain in CMHPs. To further enhance the dissolution and prebiotic potential of CMHP, pectinase was used alone and combined with cellulase. The key factor for the optimal extraction was enzymolysis by cellulase and pectinase in a mass ratio of 3:1 at 1 % (w/w) dosage. The optimal CMHP had high yield (15.15 %), high content of total sugar, and Bacteroides proliferative activity superior to inulin, which was probably due to the cooperation of complex enzyme on the destruction of cell wall and pectin structural modification for raised RG-I domain (80.30 %) with relatively high degree of branching and moderate HG domain. This study provided a green strategy for extraction of RG-I enriched prebiotic pectin from plants., Competing Interests: Declaration of competing interest No conflict of interest exits in the submission of this manuscript, and manuscript is approved by all authors for publication., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Enzymatic degradation of plant biomass requires the coordinated action of various enzymes. In this study, the production of reducing sugars from pectic substrates and sugar beet pulp (SBP) was investigated and compared using commercial enzyme preparations, including M2, pectinase (E1), Viscozyme L (V-L) and L-40. V-L, a cellulolytic enzyme mix produced by Aspergillus sp. was further evaluated as the most robust enzyme cocktail with the strongest SBP degradation ability in terms of the release of monosaccharides, methanol, and acetate from SBP. Mass-spectrometry-based proteomics analysis of V-L revealed 156 individual proteins. Of these, 101 proteins were annotated as containing a carbohydrate-active enzyme module. Notably, of the 50 most abundant proteins, ca. 44 % were predicted to be involved in pectin degradation. To reveal the role of individual putative key enzymes in pectic substrate decomposition, two abundant galacturonases (PglA and PglB), were heterologously expressed in Pichia pastoris and further characterized. PglA and PglB demonstrated maximum activity at 57 °C and 68 °C, respectively, and exhibited endo-type cleavage patterns towards polygalacturonic acid. Further studies along this line may lead to a better understanding of efficient SBP degradation and may help to design improved artificial enzyme mixtures with lower complexity for future application in biotechnology., Competing Interests: Declaration of competing interest The authors confirm that the contents of this article pose no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The investigation aims to determine the effect of enzymatic and alkali treatments on Sambucus ebulus L. stem fiber. For this purpose, Sambucus ebulus L. stem fibers were treated with alkali, cellulase, and pectinase enzymes. An image processing technique was developed and implemented to calculate the average thicknesses of Sambucus ebulus L. fibers. The thickness of alkali, cellulase and pectinase enzyme treated fibers was determined as 478.62 μm, 808.28 μm and 478.20 μm, respectively. Scanning electron microscopy analysis illustrated that enzymatic and alkali treatments lead to the breakage of fiber structure. Furthermore, enzymatic and alkali treatments induce variations in elemental ingredients. All treatments increased the crystallinity index of Sambucus ebulus L. fiber from 72 % (raw fiber) to 83 % (alkali treated), 75.2 % (cellulase enzyme treated) and 86.3 % (pectinase enzyme treated) due to the hydrolysis of hemicellulose. Fourier transform infrared analysis indicated that there are no significant differences in functional groups. Thermogravimetric analysis shows that enzymatic and alkali treatments improve final degradation temperature of the fiber. Mechanical behaviors of cellulase enzyme-treated fiber decrease compared to raw fiber, while pectinase enzyme and alkali treatment cause to improve mechanical properties. Tensile strength of samples was determined as 76.4 MPa (cellulase enzyme treated fiber), 210 MPa (pectinase enzyme treated fiber) and 240 MPa (alkali treated fiber). Young's modules of cellulase enzyme, pectinase enzyme and alkali treated fibers were predicted as 5.5 GPa, 13.1 GPa and 16.6 GPa. Elongation at break of samples was calculated as 5.5 % (cellulase enzyme treated fiber), 6.5 % (pectinase enzyme treated fiber) and 6 % (alkali treated fiber). The results suggest that enzymatic and alkali treatments can modify the functional and structural attributes of Sambucus ebulus L. fiber., 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 © 2024 Elsevier B.V. All rights reserved.)

Endopolygalacturonases are crucial pectinases known for their efficient and sustainable pectin depolymerization activities. The present study identified a novel gene encoding endopolygalacturonase from an acidic mine tailing metagenome. The putative gene showed a maximum identity of 67.55 % with an uncharacterized peptide sequence from Flavobacterium fluvii. The gene was cloned and expressed in a heterologous host, E. coli. Biochemical characterization of the novel endopolygalacturonase enzyme variant (EPH M ) showed maximum activity at 60 °C and at 5.0 pH, while retaining 50 % activity under the temperature and pH range of 20 °C to 70 °C for 6 h, and 3.0 to 10.0 for 3 h, respectively. The enzyme exhibited tolerance to different metal ions. EPH M was characterized for the depolymerization of methylated pectin into pectic oligosaccharides. Further, its utility was established for fruit juice clarification, as endorsed by high transmittance, significant viscosity reduction, and release of reducing sugars in the treated fruit juice samples., 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 © 2024 Elsevier B.V. All rights reserved.)

Global market of food enzymes is held by pectinases, mostly sourced from filamentous fungi via submerged fermentation. Given the one-time use nature of enzymes to clarify juices and wines, there is a crucial need to explore alternatives for enzyme immobilization, enabling their reuse in food applications. In this research, an isolated fungal strain (Penicillium crustosum OR889307) was evaluated as a new potential pectinase producer in submerged fermentation. Additionally, the enzyme was immobilized in magnetic core-shell nanostructures for juice clarification. Findings revealed that Penicillium crustosum exhibited enzymatic activities higher than other Penicillium species, and pectinase production was enhanced with lemon peel as a cosubstrate in submerged fermentation. The enzyme production (548.93 U/mL) was optimized by response surface methodology, determining the optimal conditions at 35 °C and pH 6.0. Subsequently, the enzyme was covalently immobilized on synthesized magnetic core-shell nanoparticles. The immobilized enzyme exhibited superior stability at higher temperatures (50 °C) and acidic conditions (pH 4.5). Finally, the immobilized pectinases decreased 30 % the orange juice turbidity and maintained 84 % of the enzymatic activity after five consecutive cycles. In conclusion, Penicillium crustosum is a proven pectinase producer and these enzymes immobilized on functionalized nanoparticles improve the stability and reusability of pectinase for juice clarification., 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 this research., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Bacterial isolates collected from the environment were screened for pectinolytic activity, and a strain with the highest activity was selected and identified as Bacillus subtilis Mz-12. The presence of pectin hydrolase, lyase, and esterase activities was confirmed. Pectinase was purified and characterized. Enzyme production was optimized with respect to temperature, pH, and growth medium. Enzyme stability and activity were characterized under different temperatures and pH values. The results showed that this strain was capable of producing high yields of pectinase in commercial medium (Pharmamedia) 24.6 U/mL compared to other media. The purified pectinase of 22.3 kDa produced was constitutive in nature. The isolated enzyme from this strain displayed a wide range of temperature and pH stability, with the optimal activity observed at pH 9.0 and 50°C. These results indicate that the B. subtilis Mz-12 strain is a favorable candidate for industrial enzyme production. The use of Pharmamedia is reported for first time for pectinase production., (© 2023 International Union of Biochemistry and Molecular Biology, Inc.)

Halophytes are the native inhabitants of saline environment. Their biomass can be considered as a potential substrate for the production of microbial enzymes. This study was intended at feasible utilization of a halophytic biomass, Cressia cretica, for pectinase production using a halo- and thermo-tolerant bacterium, Bacillus vallismortis MH 10. The data from fractionation of the C. cretica biomass revealed presence of 17% pectin in this wild biomass. Seven different factors (temperature, agitation, pH, inoculum size, peptone concentration, substrate concentration, and incubation time) affecting pectinase production using C. cretica were assessed through a statistical tool, Plackett-Burman design. Consequently, two significant factors (incubation time and peptone concentration) were optimized using the central composite design. The strain produced 20 IU mL -1 of pectinase after 24 h under optimized conditions. The enzyme production kinetics data also confirmed that 24 h is the most suitable cultivation period for pectinase production. Fourier transform infrared spectroscopy and scanning electron microscopy of C. cretica biomass ascertained utilization of pectin and structural changes after fermentation. The purification of pectinase by using DEAE column yielded specific activity and purification fold of 88.26 IU mg -1 and 3.2, respectively. The purified pectinase had a molecular weight of >65 kDa. This study offers prospects of large-scale production of pectinase by halotolerant strain in the presence of economical and locally grown substrate that makes the enzyme valuable for various industrial operations., (© 2023 Wiley-VCH GmbH.)

The main goal of this study was to examine the efficiency of a newly isolated fungus from quince, Aspergillus tubingensis FAT43, to produce the pectinolytic complex using agricultural and industrial waste as the substrate for solid state fermentation. Sugar beet pulp was the most effective substrate inducer of pectinolytic complex synthesis out of all the waste residues examined. For endo-pectinolytic and total pectinolytic activity, respectively, statistical optimization using Placked-Burman Design and Optimal (Custom) Design increased production by 2.22 and 2.15-fold, respectively. Liquification, clarification, and an increase in the amount of reducing sugar in fruit juices (apple, banana, apricot, orange, and quince) processed with pectinolytic complex were identified. Enzymatic pre-treatment considerably increases yield (14%-22%) and clarification (90%). After enzymatic treatment, the best liquefaction was observed in orange juice, whereas the best clarification was obtained in apricot juice. Additionally, the pectinolytic treatment of apricot juice resulted in the highest increase in reducing sugar concentration (11%) compared to all other enzymatically treated juices. Optimizing the production of a highly active pectinolytic complex and its efficient utilization in the processing of fruit juices, including the generation of an increasing amount of waste, are the significant outcomes of this research., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

The present study was conducted to investigate the effects of polygalacturonase (PG) treatment on carotenoid absorption upon digestion of HPH-treated combined peach and carrot juice (CJ) with or without the presence of lipids. Results showed that PG treatment reduced median particle diameter (D 50 ) and viscosity of CJ, and increased total carotenoid bioaccessibility by 41%. In the presence of emulsion, the bioaccessibility of carotenoids was higher and it was not significantly affected by PG treatment. Xanthophylls (lutein and zeaxanthin) had higher bioaccessibility than the more lipophilic carotenes (β-carotene and α-carotene); also, uptake in Caco-2 cells and transport of lutein and zeaxanthin were higher than for β-carotene and α-carotene. Individual carotenoids bioaccessibility was negatively correlated with their transport. All together data showed digestion and absorption processes were two independent processes: factors improving carotenoid bioaccessibility did not necessarily affect their bioavailability., 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 Elsevier Ltd. All rights reserved.)

Polygalacturonase (PG) is an important hydrolytic enzyme involved in pectin disassembly and the subsequent textural changes during fruit ripening. Although the interaction of fungal PGs with other proteins has been documented, the interaction of plant PGs with other plant proteins has not yet been studied. In this study, the molecular mechanisms involved in raspberry fruit ripening, particularly the polygalacturonase (RiPG) interaction with polygalacturonase inhibiting protein (RiPGIP) and substrate, were investigated with a structural approach. The 3D model of RiPG2 and RiPGIP3 was built using a comparative modeling strategy and validated using molecular dynamics (MD) simulations. The RiPG2 model structure comprises 11 complete coils of right-handed parallel β-helix architecture, with an average of 27 amino acid residues per turn. The structural model of the RiPGIP3 displays a typical structure of LRR protein, with the right-handed superhelical fold with an extended parallel β-sheet. The conformational interaction between the RiPG2 protein and RiPGIP3 showed that RiPGIP3 could bind to the enzyme and thereby leave the active site cleft accessible to the substrate. All this evidence indicates that RiPG2 enzyme could interact with RiPGIP3 protein. It can be a helpful model for evaluating protein-protein interaction as a potential regulator mechanism of hydrolase activity during pectin disassembly in fruit ripening., 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 Elsevier Inc. All rights reserved.)

Among the enzymes required for the efficient utilisation of pectin is polygalacturonase. Saccharobesus litoralis harbours two polygalacturonases belonging to glycoside hydrolase family 28 (GH28). One of them, PGQ1, cleaved polygalacturonate exolytically at the non-reducing end into monomeric units. It was most active at 60 °C and pH 8, with K m and k cat values of 2.3 mg/ml and 6.4 s -1 respectively. Its homology model of a right-handed parallel β-helix core consisted of Asp297 as the general acid and either Asp276 or Asp298 as the general base. By inferring the substrate binding modes at the -1 and +1 subsites from known crystal structures, a hexagalacturonate could be docked into the highly electropositive binding cleft. Interestingly, while no residues were present in the vicinity to make up the +2 and +4 subsites, Arg361 and Arg430 could readily bind to the carboxyl groups of the galacturonates at the +3 and +5 subsites respectively. Structural comparison suggested that this binding pattern with missing subsites might be unique to closely related exopolygalacturonases. As S. litoralis grew much more slowly on extracellular galacturonate due to the lack of a transporter for the monosaccharide, PGQ1 probably functioned in the periplasm to help degrade oligopectates completely.Communicated by Ramaswamy H. Sarma.

The cultural parameters of Streptomyces sp. for pectinase production were optimized using the Box-Behnken design. The maximum pectinase production was obtained after 58 h at 35°C and pH 7 upon submerged fermentation in yeast extract-containing media. The enzymes were partially purified with acetone precipitation, and the analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and zymogram revealed that Streptomyces sp. produced two pectinases protein with molecular weights of about 25 and 75 kDa. The pectinase activity was detected in a wide range of temperatures (30°C-80°C) and pH (3-9) with maximum pectinase activities observed at 70°C and pH 5 and 9. The enzymes retained about 30-40% of their activities even after incubating the enzyme at different temperatures for 120 min. The pectinase activities of Streptomyces sp. were enhanced in the media containing 1.5% pectin, 1% casein as a nitrogen source, 0.5 mM MgSO4, and 5 mM NaCl. Further, the addition of Tween-20, amino acids, and vitamins to the media also enhanced the pectinase activity. Moreover, the bacterium illustrated the ability to decolorize crystal violet dye efficiently. The decolorization rate ranged from 39.29 to 53.75%, showing the highest bacterial decolorization in the media containing 2 mg/mL crystal violet at 144 h. Therefore, the bacterium has the potential in treating wastewater produced by industries like textile industries., (© 2022 The Author(s). Published by S. Karger AG, Basel.)

During the present research, 11 gut bacteria were isolated from the freshwater fish, Systomus sarana (General name: olive barb) and upon screening, the strains produced extracellular pectinase enzyme. Among them, the SS6 strain was found to produce a high quantity of 208.731 U/ml pectinase and through molecular characterization the SS6 strain was identified as Aeromonas guangheii. During the culture of SS6 strain, a set of parameters were optimized through the response surface methodology with a Box-Behnken design, for the production of the enzyme. The optimal conditions were found to be 2.11% of maltose, 2.20% of yeast extract, 6.5 of pH, and a temperature of 27.3 °C at 32-h incubation. Under the above conditions, the activity of pectinase production was enhanced to 371 U/ml. The purified pectinase's molecular weight was determined to be ~ 50 kDa (by 10% 2-D PAGE). Totally, nine peptides were identified from the purified pectinase enzyme through the MALDI-TOF-MS analysis and MASCOT tool was used to get the mass spectrum of the peak at 2211 of peptide that indicated the reference pectinase protein. The referenced gene primer (pectate lyases) was PCR amplified and its nucleotide sequence was analyzed. The exo-pelA gene was cloned in pREST vector, which was found to be over expressed in Escherichia coli BL21. The ORF encoded for a mature protein comprising of 425 amino acids (1236 nucleotides) with a predicted molecular weight of ~ 48.7 kDa. The present findings underline the potential of the fish-gut microbes as a source of biotechnologically important enzymes., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

An extracellular pectinase from a mixed consortium of Bacillus sp. (BSP) was immobilized onto graphene oxide/chitosan composite (GO/CS) through covalent binding to enhance its recycling and operational stability features. Different parameters were optimized, including cross-linker concentration (%), time, pH, and GO/CS-pectinase ratios. GO/CS-pectinase was further characterized by FT-IR and XRD. The activity of GO/CS-pectinase was reached up to 804 μmolmin -1 with an immobilization efficiency of 80.64 ± 1.15 % under optimum conditions. GO/CS-pectinase exhibited a 3.0-folds higher half-life (t 1/2 ) than free pectinase at 50, 55, and 60 °C, respectively. The V max and K M values of GO/CS-pectinase were found to be nearly equal to the free pectinase indicating that conformational flexibility was retained. K d , t 1/2 , ∆G*, ∆H*, and ∆S* of both free pectinase and GO/CS-pectinase was 0.0339 & 0.0721 min -1 , 9.62 and 40.44 min, 81.35, 90.72 kJmol -1 , 47.098 & 63.635 kJmol -1 , -102.86 & -81.340 Jmole -1  K -1 . SEM morphological analysis further confirmed the successful binding of pectinase with GO/CS, which retained about 92 % of its original catalytic activity after ten consecutive reaction cycles. Finally, GO/CS-pectinase was employed for guava juice clarification which exhibited the turbidity reduction up to 81 % after 75 min of treatment., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2022. Published by Elsevier B.V.)

Polygalacturonase (PGase) production by Aureobasidium pullulans P56 under submerged fermentation was investigated using agro-industrial wastes and commercial carbon and nitrogen sources. The maximum PGase concentration was equivalent to 8.6 U/mL that was obtained in presence of citrus pectin at 150 rpm, 30 °C, pH = 5.5, and 60 h of fermentation conditions. However, a significant amount of enzyme production was also recorded upon the utilization of corncob (5.3 U/mL) and wheat bran (4.4 U/mL) as carbon sources. Amongst the different nitrogen sources, the highest enzyme production (8.2 U/mL) was obtained in presence of ammonium sulphate and yeast extract simultaneously at a ratio of 1:1. The enzyme was partially purified by gel filtration using Sephadex G50 equilibrated and washed with 50 mM-sodium acetate buffer. The obtained yield and specific activity were determined equivalent to 17% and 9.53 U/mg, respectively. The molecular weight of the partially purified enzyme was estimated as 54 kDa on SDS-PAGE. The conditions affecting the enzyme activity were determined and the highest enzyme activity was recorded at 40 °C and 4.5 pH. Amongst the tested metal ions, 2 and 5 mM of CaCl 2 concentrations increased the enzymatic activity by 30%. Overall, the use of corncob (2.5%) to produce PGase by A. pullulans represents an attractive agro-industrial substrate., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Pectic substances cause haziness and high viscosity of fruit juices. Pectinase enzymes are biological compounds that degrade pectic compounds. Nontoxicity and ecofriendly nature make pectinases excellent biocatalysts for juice clarification. However, the poor stability and nonreusability of pectinases trim down the effectiveness of the operation. The immobilization techniques have gained the attention of researchers as it augments the properties of the enzymes. Literature has reported the stability improvement of enzymes like lipase, laccase, hydrogen peroxidase, and cellulase upon immobilization on the membrane. However, only a few research articles divulge pectinase immobilization using a membrane. The catalysis-separation synergy of membrane-reactor has put indelible imprints in industrial applications. Immobilization of pectinase on the membrane can enhance its performance in juice processing. This review delineates the importance of physicochemical and kinematic properties of pectinases relating to the juice processing parameters. It also includes the influence of metal-ion cofactors on enzymes' activity. Considering the support and catalytic-separation facets of the membrane, the prediction of the membrane as support for pectinase immobilization has also been carried out., (© 2022 Institute of Food Technologists®.)

The aim of this study was to evaluate the potential of xylanase-pectinase enzymes in bleaching of wheat straw pulp, just to cut down the toxic wastes, in order to manage the environmental pollution. The appropriate parameters of bleaching were evaluated, and best conditions were xylanase and pectinase dose of 5.0 and 1.66 IU/g of pulp, respectively, along with material to liquid ratio of 1:7.5 (g/ml), temperature 55 °C, treatment time 3 h, Tween-80 1%, and pH 8.5. The release of reducing sugar and other non-cellulosic impurities, phenolic-hydrophobic-lignin was maximum at best bleaching conditions. Prebleaching of wheat straw pulp using these enzymes showed 14.75% decline in kappa number. Enzymatic bleaching plus 100% chemical bleaching also led to 27.90% reduction in yellowness. Using this methodology, the consumption of active chlorine was reduced up to 25%, along with an increase in burst index (7.98%), tear index (3.42%), breaking length (5.30%), viscosity (11.22%), gurley porosity (12.50%), and double-fold number (23.08%), which exhibits a remarkable enhancement in all the properties of pulp treated with enzymes. Microscopic images also confirm the effectiveness of enzymatic treatment in bleaching of wheat straw pulp. BOD and COD values of effluent also decreased by 20.74 and 17.87%, respectively. This research focussing on producing better grade paper using an eco-friendly approach would certainly benefit the paper and pulp industry. This is the first report, depicting bleaching capability of xylanase-pectinase enzymes for soda-anthraquinone pulp of wheat straw., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Pectinases are widely used in a variety of industrial processes. However, their application is limited by low catalytic processivity, reduced stability, high cost, and poor re-use compatibility. These drawbacks may be overcome by enzyme immobilization with ferromagnetic nanoparticles, which are easily recovered by a magnetic field. In this work, an endopolygalacturonase from Chondrostereum purpureum (EndoPG Cp ) expressed in Pichia pastoris was immobilized on glutaraldehyde-activated chitosan ferromagnetic nanoparticles (EndoPG Cp -MNP) and used to supplement a commercial enzyme cocktail. No significant differences in biochemical and kinetic properties were observed between EndoPG Cp -MNP and EndoPG Cp , although the EndoPG Cp -MNP showed slightly increased thermostability. Cocktail supplementation with EndoPG Cp -MNP increased reducing sugar release from orange wastes by 1.8-fold and showed a synergistic effect as compared to the free enzyme. Furthermore, EndoPG Cp -MNP retained 65% of the initial activity after 7 cycles of re-use. These properties suggest that EndoPG Cp -MNP may find applications in the processing of pectin-rich agroindustrial residues., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Exposure to ultraviolet (UV) radiation is the main reason behind extrinsic skin aging. Changes due to chronic UV exposure are called photoaging. Natural products are effective ingredients against UV-mediated skin damage. Present study investigated the anti-photoaging properties of  Camellia japonica flowers which possess various bioactivities. To enrich the extracts of C. japonica flowers, pectinase and beta-glucosidase treatment was employed. Anti-photoaging effect was screened using the changes in MMP-1 and collagen levels in UVA-irradiated human HaCaT keratinocytes. The crude extract of C. japonica flowers (CE) was shown to decrease the UVA-induced MMP-1 secretion while attenuating the collagen levels. Pectinase and beta-glucosidase treated CE (ECE) showed increased anti-photoaging effects against UVA-induced changes in MMP-1 and collagen production. Camellenodiol (CMD), a known triterpenoid from C. japonica , isolated as the active ingredient of ECE and its anti-photoaging effect was screened. Results showed that CMD ameliorated the UVA-induced deterioration in collagen levels by suppressing MMP-1 production in transcriptional level. CMD treatment downregulated the phosphorylation of p38, ERK, and JNK MAPKs along their downstream effectors, c-Fos, and c-Jun. In conclusion, enzyme-assisted extraction of C. japonica flowers was suggested to enhance the anti-photoaging properties suggestively through high bioactive content such as CMD., (© 2022 Walter de Gruyter GmbH, Berlin/Boston.)

To prepare texture-softened burdock that maintains its original shape, the effect of three different pretreatments before pectinase reaction (ENZ) up to 3 h: heating and drying (HD), freeze-thawing after heating and then drying (HFTD), or freeze-drying after heating (HFD) was compared. The hardness of raw burdock could be decreased by 15% after ENZ up to 3 h. The order of hardness by pretreatment was HFD (-93.3%) < HFTD (-80.7%) < HD (-52.8%) < R (raw), implying that the burdock could be softened by pretreatment itself. By the combination of pretreatment and ENZ, HFTD-ENZ for 3 h or HFD-ENZ over 2 h produced excellent softening. Hardness less than 2.0 × 10 4 (N/m 2 ) was obtained by HFD followed by ENZ over 2 h, and this could possibly be ingested by the tongue without chewing. More chlorogenic acid was also detected with those samples., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

In plants, polygalacturonase-inhibiting proteins (PGIPs) play critical roles for resistance to fungal disease by inhibiting the pectin-depolymerizing activity of endopolygalacturonases (PGs), one type of enzyme secreted by pathogens that compromises plant cell walls and leaves the plant susceptible to disease. Here, the interactions between PGIPs from Phaseolus vulgaris (PvPGIP1 and PvPGIP2) and PGs from Aspergillus niger (AnPG2), Botrytis cinerea (BcPG1 and BcPG2), and Fusarium moniliforme (FmPG3) were reconstituted through a yeast two hybrid (Y2H) system to investigate the inhibition efficiency of various PvPGIP1 and 2 truncations and mutants. We found that tPvPGIP2&#95;5-8, which contains LRR5 to LRR8 and is only one-third the size of the full length peptide, exhibits the same level of interactions with AnPG and BcPGs as the full length PvPGIP2 via Y2H. The inhibitory activities of tPvPGIP2&#95;5-8 on the growth of A. niger and B. cinerea were then examined and confirmed on pectin agar. On pectin assays, application of both full length PvPGIP2 and tPvPGIP2&#95;5-8 clearly slows down the growth of A. niger and B. cinerea. Investigation on the sequence-function relationships of PGIP utilizing a combination of site directed mutagenesis and a variety of peptide truncations suggests that LRR5 could have the most essential structural feature for the inhibitory activities, and may be a possible target for the future engineering of PGIP with enhanced activity. This study highlights the potential of plant-derived PGIPs as a candidate for future in planta evaluation as a pest control agent., (© 2021 Wiley Periodicals LLC.)

Pectin, a diverse carbohydrate polymer in plants consists of a core of α-1,4-linked D-galacturonic acid units, includes a vast portion of fruit and agricultural wastes. Using the wastes to produce beneficial compounds is a new approach to control the negative environmental impacts of the accumulated wastes. In the present study, we report a pectinase producing bacterium Streptomyces hydrogenans YAM1 and evaluate antioxidative and anticancer effects of the oligosaccharides obtained from pectin degradation. The production of oligosaccharides due to pectinase activity was detected by thin layer chromatography (TLC) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Our results revealed that S. hydrogenans YAM1 can degrade pectin to unsaturated pectic oligo-galacturonic acids (POS) with approximately 93% radical scavenging activity in 20 mg/mL which it is more than 50% of the same concentration of pectin. Flow cytometric analysis revealed that MCF-7 cells viability decreased more than 32 and 92% following treatment with 6 and 20 mg/mL POS after 24 h, respectively. It is suggested that pectin degradation by S. hydrogenans YAM1 is not only a new approach to produce highly active compounds from fruit wastes, but also is an effective method to remove fibrous pollutants from different environments.

Pectin, the major non-cellulosic component of primary cell wall can be degraded by polygalacturonases (PGs) and pectin methylesterases (PMEs) during pathogen attack on plants. We characterized two novel enzymes, VdPG2 and VdPME1, from the fungal plant pathogen Verticillium dahliae. VdPME1 was most active on citrus methylesterified pectin (55-70%) at pH 6 and a temperature of 40 °C, while VdPG2 was most active on polygalacturonic acid at pH 5 and a temperature of 50 °C. Using LC-MS/MS oligoprofiling, and various pectins, the mode of action of VdPME1 and VdPG2 were determined. VdPME1 was shown to be processive, in accordance with the electrostatic potential of the enzyme. VdPG2 was identified as endo-PG releasing both methylesterified and non-methylesterified oligogalacturonides (OGs). Additionally, when flax roots were used as substrate, acetylated OGs were detected. The comparisons of OGs released from Verticillium-susceptible and partially resistant flax cultivars identified new possible elicitor of plant defence responses., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Exogenous enzymes are extraneous enzymes that are not intrinsic to the subject. The exogenous enzyme industry has been rapidly developing recently. Successful application of recombinant DNA amplification, high-efficiency expression, and immobilization technology to genetically engineered bacteria provides a rich source of enzymes. Amylase, cellulase, protease, pectinase, glycosidase, tannase, and polyphenol oxidase are among the most widely used such enzymes. Currently, the application of exogenous enzyme technology in the development of natural plant resources mainly focuses on improving the taste and flavor of the product, enriching the active ingredient contents, deriving and transforming the structure of a chosen compound, and enhancing the biological activity and utilization of the functional ingredient. In this review, we discuss the application status of exogenous enzyme technology for the development of natural plant resources using typical natural active ingredients from plant, such as resveratrol, steviosides, catechins, mogrosides, and ginsenosides, as examples, to provide basis for further exploitation and utilization of exogenous enzyme technology.

This study was undertaken to investigate the potential of bioscouring in the processing of undegummed sisal fibers, using xylano-pectinolytic enzymes. Optimum bioscouring was obtained at pH 8.5 and 50 mM buffer molarity, using xylanase (10 IU) and pectinase (8 IU), with a material to liquor proportion of 1:25 (g:ml), EDTA (2 mM) and Tween 80 (0.5%), at 50 °C temperature with agitation rate of 55 rpm and treatment period of 60 min. Enzymatic treatment of sisal fibers enhanced the brightness and whiteness by 11.52 and 6.83%, respectively, and reduced the yellowness by 7.14% in comparison to control. The use of xylanase and pectinase enzymes completely replaced the chemical scouring method for removing non-cellulosic impurities. Thus, enzymatic scouring is energy saving and ecofriendly, since it completely eliminated the use of toxic chemicals used in alkaline scouring. An increase of 23.75% and 11.58% in brightness and whiteness of enzymatically scoured cum bleached fibers, as compared to chemically scoured cum bleached fibers was finally obtained, along with reduction in yellowness by 27.99%. This is the first report demonstrating environmentally sustainable enzymatic approach for scouring of undegummed sisal fibers, using enzymes, simultaneously produced from a bacterial isolate.

Aspergillus terreus can produce different holocellulose-degrading enzymes when grown in sugarcane bagasse, with predominant pectinase activity. Thus, pectinase was selected for purification and immobilization studies. Ion exchange and molecular exclusion chromatography studies were performed, after which it was possible to semipurify the enzyme with a yield of 80%. The crude extract pectinase (PECEB) and the partially purified enzyme (PEC2) were immobilized on monoamino-N-aminoethyl (MANAE)-agarose with pectinase activity yields of 66% and 98%, respectively. After immobilization in MANAE-agarose, the pectinase showed higher activity at acidic pH (pH 4.0) when compared to the nonimmobilized enzyme. It was also found that after the immobilization process, there was a threefold improvement in the enzyme's thermostability. Also, it was possible to reuse the immobilized enzyme for up to five cycles of hydrolysis with effective production of reducing sugars (0.196 mg/g of substrate). The industrial application test revealed a significant decrease in the viscosity of guava juice when the immobilized enzyme was used. PECEB, immobilized on MANAE-agarose, was the enzyme sample that generated the highest pulp viscosity reduction (approximately 47%). Although additional studies are needed for practical industrial application, the results obtained herein reveal the potential of application of immobilized pectinase in the industry., (© 2020 International Union of Biochemistry and Molecular Biology, Inc.)

Thermostability is a key property of industrial enzymes. Endo-polygalacturonases of the glycoside hydrolase family 28 have many practical applications, but only few of their structures have been determined, and the reasons for their stability remain unclear. We identified and characterized the Talaromyces leycettanus JCM12802 endo-polygalacturonase Tl PGA, which differs from other GH28 family members because of its high catalytic activity, with an optimum temperature of 70 °C. Distinctive features were revealed by comparison of thermophilic Tl PGA and all known structures of fungal endo-polygalacturonases, including a relatively large exposed polar accessible surface area in thermophilic Tl PGA. By mutating potentially important residues in thermophilic Tl PGA, we identified Thr284 as a critical residue. Mutant T284A was comparable to thermophilic Tl PGA in melting temperature but exhibited a significantly lower half-life and half-inactivation temperature, implicating residue Thr284 in the kinetic stability of thermophilic Tl PGA. Structure analysis of thermophilic Tl PGA and mutant T284A revealed that a carbon-oxygen hydrogen bond between the hydroxyl group of Thr284 and the Cα atom of Gln255, and the stable conformation adopted by Gln255, contribute to its kinetic stability. Our results clarify the mechanism underlying the kinetic stability of GH28 endo-polygalacturonases and may guide the engineering of thermostable enzymes for industrial applications.

In the present work, enzymes pectinase and naringinase were simultaneously co-immobilized on an eco-friendly chitosan coated magnetic nanoparticles (chitosanMNPs) by cross-linking using chitosan as a macro-molecular cross-linker. The maximum activity recovery of both enzymes in the co-immobilized form was obtained at chitosanMNPs to enzymes ratio of 1:3, 3% cross-linker concentration and 150 min cross-linking time. The synthesized MNPs before and after co-immobilization were characterized using different techniques. The prepared biocatalyst was found spherical with an average size below 200 nm and showed supermagnetic property with saturation magnetization of 38.28 emu/g. The optimum pH and temperature of both enzymes in co-immobilized form was found at 5.5 and 65 °C. The prepared biocatalyst exhibited an improved thermal stability with 1.8-fold increase in the half-life. The secondary structural analysis revealed that, prepared co-immobilized biocatalyst undergone changes in the conformational and structural rigidity due to macro-molecular cross-linker. The co-immobilized biocatalysts were evaluated for one pot clarification and debittering of grapefruit juice and found ~52% reduction in turbidity and ~85% reduction in the naringin content. The co-immobilized enzymes were recycled up to 7 th cycle and can be easily stored at room temperature for 30 days retaining up to 64% and 86% residual activities respectively., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Sweet osmanthus ( Osmanthus fragrans Lour.) (OF) is one of the ten most famous flowers in China for its unique and delicate fragrance. A combined solid-phase microextraction and solvent-assisted flavor evaporation method was used to accurately capture the overall aromatic profile and characterize the predominant odorants of fresh osmanthus with the help of gas chromatography (GC)-olfactometry and comprehensive two-dimensional GC-quadrupole time-of-flight mass spectrometry (GC × GC-QTOF-MS). Twenty-six volatiles were identified for the first time in OF. A total of 23 potent odorants, dominated by monoterpene oxides and C 6 aliphatic aldehydes, were identified. The efficacy of pectinase, β-glucosidase, and their combination on the aroma enhancement of OF was evaluated by quantitation of the target aroma components using GC-triple quadrupole-MS. The total concentration of key aroma components increased in all three enzyme treatment groups, and the increase was more significant in two β-glucosidase-treated groups. Changes in odor activity values and odor spectrum values of key odorants indicated that the pectinase-treated sample had more prominent floral, green, and potato-like scents. In contrast, the β-glucosidase-treated sample had more dominant floral, woody, almond-like, and fruity notes but less green odor, which was confirmed by sensory evaluation. β-Glucosidase and pectinase complement one another very well, and together, promote a remarkable aroma enhancement in OF.

Novel pectin derived from sweet potato residue was modified by high hydrostatic pressure (HHP)-assisted pectinase and then used for Pb 2+ removal from aqueous solutions. The removal characteristics and mechanisms were also investigated. Results showed that modified sweet potato pectin exhibited greater adsorption performances for Pb 2+ than that of natural ones, and showed excellent eco-friendly properties and good potential for adsorption of some other heavy metals (such as Cu 2+ ). The adsorption curves were much more conformed to Langmuir model, and the highest capacity for Pb 2+ adsorption was 263.15 mg/g with 1.00% pectin at pH 7. Chemical adsorption process of pectin for Pb 2+ absorption involved O-containing functional groups (O-H, COO-), cation exchange, and along with electrostatic interactions. Overall, the results in this study indicated that sweet potato pectin modified with HHP-assisted pectinase had the potential to become an environmentally friendly coagulant-flocculant agent for the heavy metal adsorption, especially for Pb 2+ ., 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 © 2020. Published by Elsevier Ltd.)

Background: Polygalacturonases are a group of enzymes under pectinolytic enzymes related to enzymes that hydrolyse pectic substances. Polygalacturonases have been used in various industrial applications such as fruit juice clarification, retting of plant fibers, wastewater treatment drinks fermentation, and oil extraction., Objectives: The study was evaluated at the heterologous expression, purification, biochemical characterization, computational modeling, and performance in apple juice clarification of a new exo-polygalacturonase from Sporothrix schenckii 1099-18 (SsExo-PG) in Pichia pastoris., Methods: Recombinant DNA technology was used in this study. Two different pPIC9K plasmids were constructed with native signal sequence-ssexo-pg and alpha signal sequence-ssexo-pg separately. Protein expression and purification performed after plasmids transformed into the Pichia pastoris. Biochemical and structural analyses were performed by using pure SsExo-PG., Results: The purification of SsExo-PG was achieved using a Ni-NTA chromatography system. The enzyme was found to have a molecular mass of approximately 52 kDa. SsExo-PG presented as stable at a wide range of temperature and pH values, and to be more storage stable than other commercial pectinolytic enzyme mixtures. Structural analysis revealed that the catalytic residues of SsExo- PG are somewhat similar to other Exo-PGs. The K M and k cat values for the degradation of polygalacturonic acid (PGA) by the purified enzyme were found to be 0.5868 μM and 179 s-1 , respectively. Cu 2+ was found to enhance SsExo-PG activity while Ag 2+ and Fe 2+ almost completely inhibited enzyme activity. The enzyme reduced turbidity up to 80% thus enhanced the clarification of apple juice. SsExo-PG showed promising performance when compared with other commercial pectinolytic enzyme mixtures., Conclusion: The clarification potential of SsExo-PG was revealed by comparing it with commercial pectinolytic enzymes. The following parameters of the process of apple juice clarification processes showed that SsExo-PG is highly stable and has a novel performance., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background: Japanese hop is an important cause of weed pollinosis in East Asia. Its pollen is abundant in autumn. This pollen is known to be the cause of many allergic diseases. However, molecular characteristics of its allergens have not been elucidated., Objective: In this study, we produced recombinant proteins of allergen homologues from Japanese hop by the analysis of expressed sequence tags (EST), and evaluated its allergenicity., Methods: cDNA library was constructed using as little as 50 ng of total RNA from Japanese hop pollen. Allergen homologues were identified by the initial screening of 963 EST clones. Recombinant proteins were overexpressed in the E. coli expression system and purified using Ni-nitrilotriacetic acid-agarose. Purified proteins were analyzed by ELISA., Results and Discussion: Japanese hop pathogenesis-related 1 protein (PR-1) shares 37.0 to 44.4% of amino acid sequence identity with Art v 2, Cuc m 3, and Cyn d 24. Pectin methyl esterase (PME) shows 23.2 to 50.2% of identities to Act d 7, Ole e 11, and Sal k 1. Polygalacturonase (PGs) shows 16.7 to 19.3% of identities to Phl p 13, Cry j 2, Cha o 2, Jun a 2, Pla a 2, and Pla or 2. IgE antibodies from Japanese hop allergy patients' sera recognized PR-1 (3.4%), PME (13.8%), PGs (3.7%), and profilin (13.8%), respectively., Conclusion: Novel allergenic components were identified, even though low IgE reactivity was displayed reflecting the low degree of cross-reactivity with other pollen allergens. We believe that these molecules have worth further studies., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The medicinal plant Artemisia annua L. is well known for its antimalarial compound artemisinin and the antioxidant capacity of its active ingredients. However, low bioavailability of Artemisia annua L. limits its therapeutic potential, fermentation of Artemisia annua L. can improve its bioavailability. This study was aimed to investigate the effects of dietary supplementation of enzymatically-treated Artemisia annua L. (EA) on reproductive performance, antioxidant status, milk composition of heat-stressed sows and intestinal barrier integrity of their preweaning offspring. 135 multiparous sows of average parity 4.65 (Landrace × large white) at day 85 of pregnancy were randomly distributed into 3 treatments. Sows in the control group were housed at control rooms (temperature: 27.12 ± 0.18 °C, temperature-humidity index (THI): 70.90 ± 0.80) and fed the basal diet. Sows in the HS, HS + EA groups were fed the basal diet supplemented with 0 or 1.0 g/kg EA respectively, and reared at heat stress rooms (temperature: 30.11 ± 0.16 °C, THI: 72.70 ± 0.60). Heat stress increased the malondialdehyde (MDA) content, reduced the activities of total antioxidant capacity (T-AOC) and total superoxide dismutase (T-SOD) of sows and piglets, and seriously compromised the antioxidant capacity of the sows and the intestinal integrity of their offspring. However, dietary supplementation of 1.0 g/kg EA reduced the MDA content, increased the activities of T-SOD and T-AOC in serum, colostrum, and milk of heat-stressed sows, and increased colostrum yield and 14-d milk fat content. EA supplementation also increased piglet weaning weight and the activities of T-SOD and T-AOC in serum. In addition, the abundances of intestinal tight junction proteins claudin-1 and occludin were up-regulated in piglets in EA-supplemented group. In conclusion, dietary EA supplementation at 1.0 g/kg can alleviate the oxidative stress in heat-stressed sows, improve the antioxidant capacity in both sows and their offspring, and promote the intestinal barrier integrity in their offspring. EA may be a potent dietary supplement that ameliorates oxidative stress in livestock production by improving the antioxidant capacity., (Copyright © 2020. Published by Elsevier Ltd.)