Production Of Manno-oligosaccharides Research Articles

Production of Mannooligosaccharides from Açaí Seed by Immobilized β-Mannanase

In this work, an enzyme cocktail with β-mannanase as the main activity was immobilized on epoxy resin foams filled with fibers from annatto capsules. The catalytic system was characterized by SEM, FTIR, and a mechanical crush resistance test. The behavior of the pH and temperature for the hydrolysis of the locust bean gum were also studied. With the same substrate and with respect to the free enzyme, the immobilized enzyme showed an activity retention of 79.61%. Its operational stability in ten reuse cycles did not show any statistically significant loss of activity. This catalytic system was used to study the preferential release of MOS of two to five degrees of polymerization from mannan present in dried and ground açaí seeds, which were not subjected to any other pretreatment. Using an experimental response surface design, the predicted quadratic models for the M2–M5 MOS content were obtained and they fit well with the experimental data, predicting a production range between 0.435 and 20 g/L of MOS (M2–M5). In addition, the production reached about 12 g/L under the optimized conditions. These results indicate that the used foamed epoxy resin supports and immobilization methodology are suitable for catalyzing the hydrolysis of mannan from açaí seeds.

Efficient production of High-Purity manno-oligosaccharides from guar gum by citric acid and enzymatic hydrolysis

Currently, the production of manno-oligosaccharides (MOS) from guar gum faces challenges of low oligosaccharide enzymatic hydrolysis yield and complicated steps in separation and purification. In this work, a potential strategy to address these issues was explored. By combining citric acid pretreatment (300 mM, 130 °C, 1 h) with β-mannanase hydrolysis, an impressive MOS yield of 61.8 % from guar gum (10 %, w/v) was achieved. The key success lay in the optimizing conditions that completely degraded other galactomannans into monosaccharides, which could be easily removable through Saccharomyces cerevisiae fermentation (without additional nutrients). Following ion exchange chromatography for desalination, and concluding with spray drying, 4.57 g of solid MOS with a purity of 90 % was obtained from 10 g of guar gum. This method offers a streamlined and effective pathway for obtaining high-yield and high-purity MOS from guar gum by combining citric acid pretreatment and enzymatic hydrolysis.

Production of mannooligosaccharides from orange peel waste with β-mannanase expressed in Trichosporonoides oedocephalis

A large quantity of orange peel waste (OPW) is generated per year, yet effective biorefinery methods are lacking. In this study, Trichosporonoides oedocephalis ATCC 16958 was employed for hydrolyzing OPW to produce soluble sugars. Glycosyl hydrolases from Paenibacillussp.LLZ1 which can hydrolyze cellulose and hemicellulose were mined and characterized, with the highest β-mannanase activity of 39.1 U/mg at pH 6.0 and 50 ℃. The enzyme was overexpressed in T. oedocephalis and the sugar production was enhanced by 16 %. The accumulated sugar contains 57 % value-added mannooligosaccharides by the hydrolysis of mannans. The process was intensified by a pretreatment combining H2O2 submergence and steam explosion to remove potential inhibitors. The mannooligosaccharides yield of 6.5 g/L was achieved in flask conversion and increased to 9.7 g/L in a 5-L fermenter. This study improved the effectiveness of orange peel waste processing, and provided a hydrolysis-based methodology for the utilization of fruit wastes.

Physicochemical characterization of açaí seeds (Euterpe oleracea) from Colombian pacific and their potential of mannan-oligosaccharides and sugar production via enzymatic hydrolysis

AbstractThe study aimed to characterize açaí seeds and explore their potential for producing mannooligosaccharides (MOS) through enzymatic hydrolysis. According to characterization tests by XRD, FTIR, and chemical analysis, acai seeds, a waste material from acai fruit processing, contain significant hemicellulose content, with a main content based on mannan. The study utilized Rohalase®GMP enzyme for hydrolysis and monitored reducing sugars and MOS production (8–10 g MOS/L) hydrolysate with varying lengths, including di-, tri-, tetra-, and pentasaccharide over time. Results showed a concentration plateau of reducing sugars at 13 h and increased MOS until hour nineteen. Mannobiose (M2) was the predominant MOS produced. Comparative analysis with prior research indicated that the açaí seed hydrolysate’s MOS content aligns with that from other sources. Importantly, the study achieved % hydrolysis and MOS yield of 10.79 ± 0.05% and 51.39 ± 0.11 g M2-M5/g mannan, respectively. Açaí seed is a promising source to produce enzymes, biofuels, or thermal energy, as well as highly valued chemical compounds in the industry. This work additionally demonstrates its potential as a sustainable source of valuable MOS, suggesting applications as prebiotics and functional food additives, with implications for various industries seeking eco-friendly alternatives.

Biochemical analyses of a novel acidophilic GH5 β-mannanase from Trichoderma asperellum ND-1 and its application in mannooligosaccharides production from galactomannans.

In this study, an acidophilic GH5 β-mannanase (TaMan5) from Trichoderma asperellum ND-1 was efficiently expressed in Pichia pastoris (a 2.0-fold increase, 67.5 ± 1.95 U/mL). TaMan5 displayed the highest specificity toward locust bean gum (Km = 1.34 mg/mL, Vmax = 749.14 μmol/min/mg) at pH 4.0 and 65°C. Furthermore, TaMan5 displayed remarkable tolerance to acidic environments, retaining over 80% of its original activity at pH 3.0-5.0. The activity of TaMan5 was remarkably decreased by Cu2+, Mn2+, and SDS, while Fe2+/Fe3+ improved the enzyme activity. A thin-layer chromatography (TLC) analysis of the action model showed that TaMan5 could rapidly degrade mannan/MOS into mannobiose without mannose via hydrolysis action as well as transglycosylation. Site-directed mutagenesis results suggested that Glu205, Glu313, and Asp357 of TaMan5 are crucial catalytic residues, with Asp152 playing an auxiliary function. Additionally, TaMan5 and commercial α-galactosidase displayed a remarkable synergistic effect on the degradation of galactomannans. This study provided a novel β-mannanase with ideal characteristics and can be considered a potential candidate for the production of bioactive polysaccharide mannobiose.

Characterization of mannanases from Clonostachys byssicola involved in the breakdown of lignocellulosic substrates

Enzymatic saccharification of lignocellulosic agricultural residues, such as soybean hull, is increasingly used in Brazil to obtain fermentable sugars for use in biotechnological processes. The filamentous fungus, Clonostachys byssicola, is known to secrete an arsenal of holocellulolytic enzymes, including mannanases, which degrade the hemicellulosic components in soyabean hull. In this study, we investigated the biochemical characteristics of mannanases produced by C. byssicola. The mannanases in the concentrated crude extract (CCE) of C. byssicola showed high activity at acidic pH (5.0) and 55 °C. Furthermore, these enzymes do not depend on additional metal cofactors for their activation, although phenolic compounds inhibited the mannanase activity. Multiple forms of mannanases were identified in the CCE, which holds significance in the production of manno-oligosaccharides from different carbon sources. Scanning electron microscopy revealed structural modifications in lignocellulosic residues, including soybean hull and sugarcane bagasse, upon incubation with CCE. HPLC analysis detected mannotriose as the main oligosaccharide generated during the degradation of lignocellulosic residues. In contrast, hydrolysis of galactomannans (locust bean gum and guar gum) generated oligosaccharides of different sizes, such as mannobiose, mannotriose, mannotetraose, mannopentaose, and mannohexaose. These results highlight the potential of C. byssicola mannanases in hemicellulose degradation and production of prebiotic oligosaccharides.

Characterization of a novel GH26 β-mannanase from Paenibacillus polymyxa and its application in the production of mannooligosaccharides

A novel glycoside hydrolase family 26 β-mannanase gene ppman26a was cloned from Paenibacillus polymyxa KF-1. The full-length enzyme PpMan26A and its truncated products CBM35pp (aa 35–328) and PpMan26A-Δ205 (aa 206–656) were overexpressed in Escherichia coli. PpMan26A hydrolyzed locust bean gum, guar gum, konjac gum and ivory nut mannan, with the highest specific activity toward konjac gum. The Km and kcat values for konjac gum were 2.13 mg/mL and 416.66 s–1, respectively. The oligosaccharides fraction obtained from the hydrolysis of konjac gum by PpMan26A was analyzed by matrix-assisted laser desorption ionization-time-of-flight mass spectrometer (MALDI-TOF-MS). The degradation products were mainly mannooligosaccharides with a degree of polymerization of 3–8. CBM35pp exerted strong binding activity toward mannans but without β-mannanase activity. PpMan26A-Δ205, with the deletion of the N-terminal CBM domain, showed lower substrate binding capacity, resulting in reduced enzymatic activity and thermostability. This study complements our understanding of GH26 β-mannanases and expands the potential industrial application of PpMan26A.

Bioconversion of spent coffee grounds to prebiotic mannooligosaccharides - an example of biocatalysis in biorefinery.

Spent coffee ground (SCG), an agro-industrial waste, have a high content of polysaccharides such as mannan, making it ideal for utilisation for the production of nutraceutical oligosaccharides. Recently, there has been growing interest in the production of mannooligosaccharides (MOS) for health promotion in humans and animals. MOS are reported to exhibit various bioactive properties, including prebiotic and antioxidant activity. In this study, SCG was Vivinal pretreated using NaOH, characterized and hydrolysed using a Bacillus sp. derived endo-β-1,4-mannanase, Man26A, for MOS production. Structural analyses using Fourier-transform infrared spectroscopy (FT-IR) and thermogravimetric analysis (TGA) were conducted to assess the efficacy of the pretreatment. Lignin removal by the pretreatment from SCG was clearly shown by TGA. FT-IR, on the other hand, showed the presence of α-linked d-galactopyranoside (812 cm-1) and β-linked d-mannopyranoside residues (817 cm-1) in both SCG samples, signifying the presence of mannan. Hydrolysis of pretreated SCG by Man26A produced mannobiose and mannotriose as the main MOS products. The effect of simulated gastric conditions on the MOS was investigated and showed this product to be suitable for oral administration. Finally, the prebiotic effect of the MOS on the growth of selected beneficial bacteria was investigated in vitro; showing that it enhanced Lactobacillus bulgaricus, Bacillus subtilis and Streptococcus thermophilus growth. These findings suggest that SCG is a viable source for the production of MOS which can be orally administered as prebiotics for effecting luxuriant growth of probiotic bacteria in the host's digestive tract, leading to a good health status.

The activity of β‐mannanase in the commercial enzyme Pectinex Ultra SP‐L and its use for the production of mannooligosaccharides from copra meal

SummaryThe β‐mannanase present in the commercial enzyme Pectinex Ultra SP‐L was characterised and tested for its ability to hydrolyse copra meal for the production of mannooligosaccharides (MOS). The optimum pH and temperature for the mannanase activity were 4.0 and 65 °C, respectively. The relative activity of the β‐mannanase of the commercial preparation Pectinex Ultra SP‐L on copra meal was 49% when compared to the complete hydrolysis of locust bean gum, indicating its potential to hydrolyse the agricultural waste. The maximum MOS concentration of 25 g L−1 was obtained from the hydrolysis of 300 g L−1 copra meal with Pectinex Ultra SP‐L at 55 °C for 2 h. The hydrolysate consisted of ~45% oligosaccharides and the rest were monosaccharides. Mannobiose and mannotriose were the only oligosaccharides present in the mixture, with mannobiose accounting for 90%. A simple and efficient method to obtain oligosaccharides from copra meal using a commercial pectinolytic enzyme is presented.

Covalent Immobilisation of an Aspergillus niger Derived Endo-1,4-β-Mannanase, Man26A, on Glutaraldehyde-Activated Chitosan Nanoparticles for the Effective Production of Prebiotic MOS from Soybean Meal

An Aspergillus niger endo-1,4-β-mannanase, Man26A, was confirmed by FTIR and XRD to be immobilised on glutaraldehyde-activated chitosan nanoparticles via covalent bonding. The immobilisation (%) and activity yields (%) were 82.25% and 20.75%, respectively. The biochemical properties (pH, temperature optima, and stability) were then comparatively evaluated for both the free and immobilised Man26A. The optimal activity of Man26A shifted to a lower pH after immobilisation (pH 2.0–3.0, from pH 5 for the free enzyme), with the optimum temperature remaining unchanged (60 °C). The two enzymes exhibited identical thermal stability, maintaining 100% activity for the first 6 h at 55 °C. Substrate-specific kinetic analysis showed that the two enzymes had similar affinities towards locust bean gum (LBG) with varied Vmax values. In contrast, they showed various affinities towards soybean meal (SBM) and similar Vmax values. The immobilised enzyme was then employed in the enhancement of the functional feed/prebiotic properties of SBM from poultry feed, increasing mannooligosaccharides (MOS) quantities. The SBM main hydrolysis products were mannobiose (M2) and mannose (M1). The SBM-produced sugars could be utilised as a carbon source by probiotic bacteria; Streptococcus thermophilus, Bacillus subtilis, and Lactobacillus bulgaricus. The results indicate that the immobilised enzyme has the potential for use in the sustainable and cost-effective production of prebiotic MOS from agricultural biomass.

Hyperproduction of a bacterial mannanase and its application for production of bioactive mannooligosaccharides from agro-waste

Bacillus subtilis MAN7 is known to produce a β-mannanase which is capable of producing mannooligosaccharides (MOS) from agro-wastes. Present study was aimed to hyper-produce the β-mannanase for its application in the cost-effective production of MOS from copra meal and evaluation of their bioactive properties. Optimization studies significantly enhanced the enzyme yield with an approximate 28.7fold increase. Enzyme was found to be remarkably stable under wide range of temperature and pH; making it suitable for the generation of MOS. Conditions were standardized for the pretreatment of copra meal which resulted in significant removal of undesirable components. MOS (DP2–7) was generated by the enzymatic hydrolysis of pretreated copra meal and found to exhibit effective prebiotic and antioxidant activity. Current study paves a way for the development of an eco-friendly and cost-effective bioprocess for the production of functional MOS from agro-waste using β-mannanase from B. subtilis MAN7.

Efficient and green production of manno-oligosaccharides from Gleditsia microphylla galactomannans using CO2 and solid acid in subcritical water

This study aimed to produce manno-oligosaccharides (MOS) from Gleditsia microphylla galactomannans (GMG) using CO2 and solid acid (Amberlyst-35) in subcritical water. The optimal condition for MOS preparation was 3 MPa CO2, 0.1 g/g solid acid (relative to GMG) at 150 °C for 40 min. The maximum MOS yield with a degree of polymerization from 2 to 4 (M2-M4) was 52.19%, which doubled the yield of MOS compared to either using solely CO2 or solid acid. Solid acid showed excellent performance in producing MOS under subcritical H2O–CO2 condition, due to the enhanced mass transfer efficiency and increased H+ concentration in the reaction system. The solid acid can be easily separated and reused. Comparing with traditional methods used to produce MOS, this approach has many merits such as higher galactomannan hydrolysis efficiency (largely reduced time and higher MOS yield), purer M2-M4 product, lower costs, and more environmental-friendly.

Purification and Characterization of a Thermostable β-Mannanase from Halophilic Aspergillus terreus strain ARSA Associated to a Mangrove Plant of Red Sea Coast, Egypt, and its Application in Mannooligosaccharides Production and Juice Clarification

Purification and Characterization of a Thermostable β-Mannanase from Halophilic Aspergillus terreus strain ARSA Associated to a Mangrove Plant of Red Sea Coast, Egypt, and its Application in Mannooligosaccharides Production and Juice Clarification

Manno-oligosaccharides from copra meal: Optimization of its enzymatic production and evaluation its potential as prebiotic

A recombinant β-mannanase from Bacillus circulans NT 6.7 was used for the production of manno-oligosaccharides (MOS) from copra meal. The optimum hydrolysis condition was determined by response surface methodology. A maximum MOS concentration of 1.96 mg/ml was obtained when employing 9 mg of defatted copra meal and 9 U of recombinant β-mannanase per ml, incubated at 50 °C for 15.5 h. Hydrolysis products in copra meal hydrolysate (CMH-R) consisted of 0.80, 0.70, 0.46 and 0.01 mg/ml of mannotetraose, mannotriose, mannobiose and mannose, respectively. CMH-R was highly resistant to simulated gastrointestinal condition, with 3.52% and 6.59% hydrolyzed under in vitro simulated stomach and small intestine condition, respectively. Moreover, CMH-R showed potential to promote the growth of Lactobacillus reuteri AC5, Lactobacillus johnsonii KUNN 19-2, Weisella confusa JCM 1093, Bifidobacterium bifidum TISTR 2129, and Bifidobacterium animalis subsp. animalis TISTR 2194, as well as Escherichia coli KUB-E010, Salmonella Enteritidis KUB-S003, and Staphylococcus aureus TISTR 029 similar to Fructooligosaccharide (FOS) with a specific growth rate 0.24–1.41 h−1. Conversely, CMH-R did not enhance the growth of the pathogenic bacterium S. dysenteriae DMST 1511. Hence, copra meal hydrolysate containing manno-oligosaccharides prepared by recombinant β-mannanase shows a potential to be used as a prebiotic which can be employed in both food and feed applications.

Enzymatic Conversion of Mannan-Rich Plant Waste Biomass into Prebiotic Mannooligosaccharides.

A growing demand in novel food products for well-being and preventative medicine has attracted global attention on nutraceutical prebiotics. Various plant agro-processes produce large amounts of residual biomass considered “wastes”, which can potentially be used to produce nutraceutical prebiotics, such as manno-oligosaccharides (MOS). MOS can be produced from the degradation of mannan. Mannan has a main backbone consisting of β-1,4-linked mannose residues (which may be interspersed by glucose residues) with galactose substituents. Endo-β-1,4-mannanases cleave the mannan backbone at cleavage sites determined by the substitution pattern and thus give rise to different MOS products. These MOS products serve as prebiotics to stimulate various types of intestinal bacteria and cause them to produce fermentation products in different parts of the gastrointestinal tract which benefit the host. This article reviews recent advances in understanding the exploitation of plant residual biomass via the enzymatic production and characterization of MOS, and the influence of MOS on beneficial gut microbiota and their biological effects (i.e., immune modulation and lipidemic effects) as observed on human and animal health.

Production of mannooligosaccharides producing β-Mannanase by newly isolated Penicillium aculeatum APS1 using oil seed residues under solid state fermentation

The present investigation was focused on the production of extracellular β-mannanase from newly isolated Penicillium aculeatum APS1 using palm kernel cake and soyabean meal which are the residual by-products of oil extraction industry, under solid state fermentation. On supplementing palm kernel cake with 20% soyabean meal, yield of β-mannanase production was reached to 2807 U/g. Response surface methodology was used for statistical optimization of β-mannanase production. Two independent variables, namely moisture level and incubation time, were found to be significantly contributing for the production process. Under optimized condition of moisture level (52.25%) and incubation time (130 h), the yield of β-mannanase was improved by 1.6 fold and maximum activity of β-mannanase was 4696 U/g. The optimum temperature and pH for crude β-mannanase were 65 °C and 6.0, respectively. Crude and partially purified β-mannanase was found to be effective in release of mannooligosachharides by hydrolysis of mannan rich substrates viz. locust bean gum, guar gum and konjac glucomannan. Qualitative and quantitative analysis of MOS was carried out by TLC and ion chromatography. β-Mannanase from Penicillium aculeatum APS1 was found to have properties suitable for applications in feed/food industry.

Production of manno-oligosaccharide from Gleditsia microphylla galactomannan using acetic acid and ferrous chloride

A novel and efficient method for manno-oligosaccharides (MOS) production has been proposed by utilizing Gleditsia microphylla galactomannan as the starting material. This co-operative hydrolysis using ferrous chloride (Fe2+) and acetic acid (HAc) effectively improved the MOS yield and meanwhile decreased the amount of monosaccharide and the 5-hydroxymethyl-furfural (HMF). The highest yields under the optimum conditions were 46.7% by HAc hydrolysis (5 M HAc at 130 °C for 120 min); 37.3% by Fe2+ hydrolysis (0.1 M Fe2+ at 150 °C for 120 min); and 51.4% by co-operative hydrolysis (2 M HAc, 0.05 M Fe2+ at 160 °C for 10 min). From the changes in the value of M/G (mannose/galactose) ratios, it was deduced that Fe2+ predominantly cleaves the main chain, and HAc assists in the breakage of the side chain, thus resulting in the high-efficient co-operative hydrolysis for the production of MOS.

Characterization of Bacillus subtilis GA2(1) mannanase expressed in Escherichia coli Rosetta (DE3) for enzymatic production of manno-oligosaccharides from spent coffee grounds and in vitro assessment of their prebiotic properties

Characterization of Bacillus subtilis GA2(1) mannanase expressed in Escherichia coli Rosetta (DE3) for enzymatic production of manno-oligosaccharides from spent coffee grounds and in vitro assessment of their prebiotic properties

Optimization of mannooligosaccharides production from different hydrocolloids via response surface methodology using a recombinant Aspergillus sojae β‐mannanase produced in the microparticle‐enhanced large‐scale stirred tank bioreactor

Optimization of mannooligosaccharides production from different hydrocolloids via response surface methodology using a recombinant <i>Aspergillus sojae</i> β‐mannanase produced in the microparticle‐enhanced large‐scale stirred tank bioreactor

Production of mannooligosaccharides from various mannans and evaluation of their prebiotic potential

Aspergillus quadrilineatus endo-β-mannanase effectively degraded konjac glucomannan (66.09% w/v), copra meal (38.99% w/v) and locust bean galactomannan (20.94% w/v). High performance liquid chromatography (HPLC) analysis of KG hydrolysate indicated its mannooligosaccharides (MOS) content (656.38 mg/g) with high amounts of DP 5 oligosaccharide. Multi-scale characterization of mannan hydrolysate was done using FTIR and 13C NMR which revealed α and β form of galactose or glucose in MOS, respectively. CM and LBG hydrolysates (1 mg/mL) have shown cytotoxic effect and reduced cell viability of Caco-2 cells by 45% and 62%, respectively. MOS DP (1–4) derived from LBG supported better Lactobacilli biofilm formation as compared to KG hydrolysate containing high DP MOS (5–7). Lactobacilli effectively fermented MOS to generate acetate and propionate as main short chain fatty acids. Lactobacilli produced leucine, isoleucine and valine as branched chain amino acids when grown on LBG hydrolysate.