Enzyme Dosage Research Articles

Comparing the wound healing potential of natural rubber latex serum and F1-protein: An in vivo approach

Chronic wounds pose significant health concerns. Current treatment options include natural compounds like natural rubber latex (NRL) from Hevea brasiliensis. NRL, particularly the F1 protein fraction, has demonstrated bioactivity, biocompatibility, and angiogenic effects. So far, there is no study comparing F1 protein with total NRL serum, and the necessity of downstream processing remains unknown. Here, we evaluated the angiogenic potential of F1 protein compared to total NRL serum and the need for downstream processing. For that, ion exchange chromatography (DEAE-Sepharose), antioxidant activity, physicochemical characterization, cell culture in McCoy fibroblasts, and wound healing in Balb-C mice were performed. Also, the evaluation of histology and collagen content and the levels of inflammatory mediators were quantified. McCoy fibroblast cell assay showed that F1 protein (0.01 %) and total NRL serum (0.01 %) significantly increased cell proliferation by 47.1 ± 11.3 % and 25.5 ± 2.5 %, respectively. However, the AA of F1 protein (78.9 ± 0.8 %) did not show a significant difference compared to NRL serum (77.0 ± 1.1 %). F1 protein and NRL serum were more effective in wound management in rodents. Histopathological analysis confirmed accelerated healing and advanced tissue repair. Similarly, the F1 protein (0.01 %) increased collagen, showing that this fraction can stimulate the synthesis of collagen by fibroblastic cells. Regarding cytokines production (IL-10, TNF-α, IFN-γ), F1 protein and NRL serum did not exert an impact on the synthesis of these cytokines. Furthermore, we did not observe statistically significant changes in dosages of enzymes (MPO and EPO) among the groups. Nevertheless, Nitric Oxide dosage was reduced drastically when the F1 protein (0.01 %) protein was applied topically. These findings contribute to the understanding of F1 protein and NRL serum properties and provide insights into cost-effectiveness and practical applications in medicine and biotechnology. Therefore, further research is needed to assess the economic feasibility of downstream processing for NRL-based herbal medicine derived from Hevea brasiliensis.

Optimisation study of Ulmus pumila woody biomass fractionation by steam explosion for bioproducts production

Ulmus pumila represents a promising lignocellulosic biomass source for biofuels and bioproducts production since it can grow in low rainfall and extreme temperature zones. A first step in the conversion process is biomass fractionation to enhance the performance of the hydrolysis and subsequent biological conversion steps. The aim of this work is to optimise the main variables (temperature, residence time and the addition or not of sulphuric acid) of steam explosion to pretreat Ulmus pumila biomass. The optimal conditions to maximise both glucose and xylose recovery were 204.8 °C and 30 mg H2SO4/g biomass, obtained through a multilevel factorial design of experiments. Additionally, enzymatic hydrolysis using high solid loads (15% and 20% (w/w)) and different enzyme doses was studied. As a result, steam explosion at optimal conditions followed by enzymatic hydrolysis with 20% solid loading and 60 mg protein/g cellulose of enzyme allow the recovery of 70% of the potential sugars.

Specific Modification of Granular Potato Starch by Means of Partial Debranching Using Pullulanase

AbstractNative potato starch (PS‐N) is enzymatically modified in the granular state using pullulanase (PUL) for the purpose of a specific partial molecular degradation of the polymers. The PUL compound is added to the aqueous starch suspension (40%, w/w) and processed. The process parameters are varied systematically (enzyme dosage [ENZ] 4/20 mL; pH of the suspension [pH 4.7/7.3]; hydrolysis temperature [TEMP] 40/50 °C, and hydrolysis duration [TIME] 20/120 min) and a new‐developed heat‐induced enzyme inactivation approach (storage of the partially dewatered moist starch for 120 min at 100 °C) is intended to terminate the hydrolysis. Morphological (LM, SEM, and CLSM) and thermal characterization (DSC) of the starch products indicate a partial damage of the granules and a partial loss of the semicrystalline structure owing to the heat treatment, which is confirmed by XRD. The molecular properties (SEC‐MALS‐DRI) are mainly controlled by the factors ENZ, pH, and TEMP, but the intended degradation of the amylopectin (AP) by cleavage of the α‐1,6‐linkages (debranching) is accompanied by a molecular degradation of the amylose (AM) fraction. However, both specificity of the hydrolysis and achievable gel strength are remarkably improved compared to acid‐thinned products.

Effects of transglutaminase treatment on cooking quality, textural properties, and overall acceptability of high fibre pasta incorporated with pennywort residue

Pennywort juice is a herbal drink extracted from pennywort leaves and stems. Pennywort residue is a by-product of pennywort juice production. In the present work, this by-product was proved to be a good dietary fibre ingredient in the making of high fibre pasta. Nevertheless, the addition of 10% pennywort residue reduced cooking properties, textural profiles, and overall acceptability of the pasta samples. The effects of transglutaminase treatment of pasta dough on the product quality were then investigated. Increase in transglutaminase dosage from 0.00 to 0.75 U/g protein of the flour blend decreased the cooking loss of pasta by 19% while improving its tensile strength and elongation rate by 15 and 49%, respectively. Further increase in enzyme dosage from 0.75 to 1.00 U/g protein, on the other hand, decreased the tensile strength and elongation rate. When the enzyme treatment lasted for 30 min, the cooking loss was reduced by 10%, while the tensile strength and elongation rate of high fibre pasta were both enhanced by 10%. However, increase in treatment time from 30 to 40 min did not cause any significant differences in textural and cooking properties of the fibre-rich pasta. The appropriate transglutaminase dosage and biocatalytic time were 0.75 U/g protein and 30 min, respectively, under which the overall acceptability of the sample incorporated with 10% pennywort residue powder was similar to that of the semolina pasta.

Effects of Metal Chloride Salt Pretreatment and Additives on Enzymatic Hydrolysis of Poplar

Metal chloride salt pretreatment was performed to isolate and convert cellulose to glucose from poplar. A glucose yield of 82.0% ± 0.7 was achieved after 0.05 mol/L AlCl3 pretreatment conducted at 180 °C for 20 min, ascribing to the removal of hemicellulose, the alteration of crystallinity, surface morphology, and the retention of the majority of cellulose. Then, the influence of different additives on glucose yield was assessed, generating the highest glucose yield of 88.5 ± 0.06 with the addition of PEG 8000. Meanwhile, a similar glucose yield of 82.8% ± 0.3 could be obtained with PEG 8000 when hydrolysis time was reduced by a quarter and enzyme dosage by three-quarters. It can be seen that AlCl3 pretreatment is a viable and efficient pretreatment method for poplar, while the addition of PEG 8000 can enhance the enzymatic efficiency and reduce cellulase loading, ascribing to the reservation of free enzyme and enzyme activity in the supernatant and the reduction in surface tension, which provide an idea to improve the economics of the enzymatic conversion of poplar.

High-Yield Synthesis of Phosphatidylserine in a Well-Designed Mixed Micellar System.

A sustainable enzymatic system is essential for efficient phosphatidylserine (PS) synthesis in industrial production. Conventional biphasic systems face challenges such as excessive organic solvent usage, enzyme-intensive processes, and increased costs. This study introduces a novel approach using chitin nanofibrils (ChNFs) as an immobilization material for phospholipase D (PLD) in a mixed micellar system stabilized by the food-grade emulsifier sodium deoxycholate (SDC). The immobilized enzyme, ChNF-chiA1, was quickly prepared in a one-step process, eliminating the need for purification. By optimizing the reaction conditions, including l-Ser concentration (1.0 M), SDC concentration (10 mM), reaction time (8 h), and enzyme dosage (1.0 U), a remarkable PS yield of 96.74% was achieved in the solvent-free mixed micellar system. The catalytic efficiency of ChNF-chiA1 surpassed that of the free PLD-chiA1 biphasic system by 6.0-fold. This innovative and green biocatalytic technology offers a reusable solution for the high-value enzymatic synthesis of phospholipids, providing a promising avenue for industrial applications.

Enhanced ethanol production from paper sludge waste under high-solids conditions with industrial and cellulase-producing strains of Saccharomyces cerevisiae

Reported ethanol titres from hydrolysis-fermentation of the degraded fibres in paper sludge (PS) waste, generally obtained under fed-batch submerged conditions, can be improved through fermentation processes at high solids loadings, as demonstrated in the present study with two industrial PS wastes at enzyme dosages appropriate for solids loadings up to 40% (w/w). The industrial yeast,Saccharomyces cerevisiaestrain Ethanol Red®, was compared to two genetically engineeredS. cerevisiaestrains, namely Cellusec® 1.0 and Cellusec® 2.0, capable of xylose utilisation, and xylose utilisation and cellulase production, respectively. High-solids batch fermentations were conducted in 3 L horizontal rotating reactors and ethanol titres of 100.8 and 73.3 g/L were obtained for virgin pulp and corrugated recycle PS, respectively, at 40% (w/w) solids loading using Ethanol Red®. Xylose utilisation by Cellusec® 1.0 improved ethanol titres by up to 10.3%, while exogenous cellulolytic enzyme requirements were reduced by up to 50% using cellulase-producing Cellusec® 2.0.

Thermal Stability Enhancement of L-Asparaginase from Corynebacterium glutamicum Based on a Semi-Rational Design and Its Effect on Acrylamide Mitigation Capacity in Biscuits.

Acrylamide is present in thermally processed foods, and it possesses toxic and carcinogenic properties. L-asparaginases could effectively regulate the formation of acrylamide at the source. However, current L-asparaginases have drawbacks such as poor thermal stability, low catalytic activity, and poor substrate specificity, thereby restricting their utility in the food industry. To address this issue, this study employed consensus design to predict the crucial residues influencing the thermal stability of Corynebacterium glutamicum L-asparaginase (CgASNase). Subsequently, a combination of site-point saturating mutation and combinatorial mutation techniques was applied to generate the double-mutant enzyme L42T/S213N. Remarkably, L42T/S213N displayed significantly enhanced thermal stability without a substantial impact on its enzymatic activity. Notably, its half-life at 40 °C reached an impressive 13.29 ± 0.91 min, surpassing that of CgASNase (3.24 ± 0.23 min). Moreover, the enhanced thermal stability of L42T/S213N can be attributed to an increased positive surface charge and a more symmetrical positive potential, as revealed by three-dimensional structural simulations and structure comparison analyses. To assess the impact of L42T/S213N on acrylamide removal in biscuits, the optimal treatment conditions for acrylamide removal were determined through a combination of one-way and orthogonal tests, with an enzyme dosage of 300 IU/kg flour, an enzyme reaction temperature of 40 °C, and an enzyme reaction time of 30 min. Under these conditions, compared to the control (464.74 ± 6.68 µg/kg), the acrylamide reduction in double-mutant-enzyme-treated biscuits was 85.31%, while the reduction in wild-type-treated biscuits was 68.78%. These results suggest that L42T/S213N is a promising candidate for industrial applications of L-asparaginase.

Optimization of fucoidan recovery by ultrasound-assisted enzymatic extraction from South African kelp, Ecklonia maxima

Fucoidan, a sulphated polysaccharide, is found exclusively in brown seaweeds and has been reported to possess a wide range of biological functionalities. Fucoidans are found within the cell wall of brown seaweeds, which is composed of recalcitrant cellulose and hemicellulose. This hampers the recovery of fucoidans. In addition, fucoidans are found within a network of viscous hydrocolloids, alginates, further complicating their recovery. Traditionally, the hot water extraction method is used to recover fucoidans from brown seaweed, however, this is characterized by low yields and long extraction time. To combat these issues, several novel extraction technologies have been introduced, these include ultrasound-assisted extraction and enzyme-assisted extraction. Thus, the main aim of this study was to investigate and optimize fucoidan recovery from Ecklonia maxima based on ultrasound-assisted enzymatic extraction. The impact of temperature (40–65 °C), ultrasound intensity (0–118 W·cm−2), enzyme dosage (0–0.05 ml·g−1) and pH (4.5–6) on total dissolved, total carbohydrates and inorganic sulphates yields was studied. The application of ultrasound-assisted enzymatic extraction mainly improved the extraction of total carbohydrates. Ultrasound significantly improves the kinetics and extraction of fucoidan, but there was no merit when it was applied with enzymes. Results reveal that at optimized conditions, the fucoidan extracted 79.13 mg⋅g−1 (7.9 % w/w) of algal dry weight. The present study provides insight into the extraction potentials of enzyme-assisted extraction, ultrasound-assisted extraction, and ultrasound-assisted enzymatic extraction.

Food industry biowaste as substrate for biosolvents production focusing on lactic acid and ethyl lactate

Solvents represent a major category of chemicals due to their necessity in a variety of industries. The majority of solvents derived from fossil carbon are hazardous to both human health and the environment. In this context, it seems imperative to convert biomass into new alternative greener solvents. The present study concentrates on the production of two biosolvents; namely lactic acid and ethyl lactate, from biowaste of baby food industry. For the production of lactic acid parameters as the dosages of enzymes (5 mg/gLactose and 20 mg/gLactose) and inoculum size (5% v/v and 10% v/v) were examined. The lactic acid concentration reached almost 17.3g/L after 144 hours with 5mgLactozymePure/glactose and 10%v/v inoculum size. Repeated fermentation was also applied yielding a lactic acid productivity of 0.97 g/L h, which is 8 times higher than the conventional batch fermentation. Moreover, the production of ethyl lactate from bioethanol and lactic acid from the fermentation broth was examined. Lactic acid conversion yields as high as 59.9% at 255min were observed. Overall, the results of this work are very promising but further research is needed to optimize conditions and evaluate scalability of the production of these biosolvents from biowaste, since the production of lactic acid and ethyl-lactate from biowaste provides a sustainable approach to the circular economy, limiting waste and minimizing the environmental effect of industrial processes.

Patterns, Prognostic Implications, and Rural-Urban Disparities in Optimal GDMT Following HFrEF Diagnosis Among Medicare Beneficiaries

BackgroundPatterns and disparities in guideline-directed medical therapy (GDMT) uptake for heart failure with reduced ejection fraction (HFrEF) across rural vs urban regions are not well described. ObjectivesThis study aims to evaluate patterns, prognostic implications, and rural-urban differences in GDMT use among Medicare beneficiaries following new-onset HFrEF. MethodsPatients with a diagnosis of new-onset HFrEF in a 5% Medicare sample with available data for Part D medication use were identified from January 2015 through December 2020. The primary exposure was residence in rural vs urban zip codes. Optimal triple GDMT was defined as ≥50% of the target daily dose of beta-blockers, ≥50% of the target daily dose of angiotensin-converting enzyme inhibitors/angiotensin receptor blocker or any dose of sacubitril/valsartan, and any dose of mineralocorticoid receptor antagonist. The association between the achievement of optimal GDMT over time following new-onset HFrEF diagnosis and risk of all-cause mortality and subsequent HF hospitalization was also evaluated using adjusted Cox models. The association between living in rural vs urban location and time to optimal GDMT achievement over a 12-month follow-up was assessed using cumulative incidence curves and adjusted Fine-Gray subdistribution hazard models. ResultsA total of 41,296 patients (age: 76.7 years; 15.0% Black; 27.6% rural) were included. Optimal GDMT use over the 12-month follow-up was low, with 22.5% initiated on any dose of triple GDMT and 9.1% on optimal GDMT doses. Optimal GDMT on follow-up was significantly associated with a lower risk of death (HR: 0.89 [95% CI: 0.85-0.94]; P < 0.001) and subsequent HF hospitalization (HR: 0.93 [95% CI: 0.87-0.98]; P = 0.02). Optimal GDMT use at 12 months was significantly lower among patients living in rural (vs urban) areas (8.4% vs 9.3%; P = 0.02). In adjusted analysis, living in rural (vs urban) locations was associated with a significantly lower probability of achieving optimal GDMT (HR: 0.92 [95% CI: 0.86-0.98]; P = 0.01 Differences in optimal GDMT use following HFrEF diagnosis accounted for 16% of excess mortality risk among patients living in rural (vs urban) areas. ConclusionsUse of optimal GDMT following new-onset HFrEF diagnosis is low, with substantially lower use noted among patients living in rural vs urban locations. Suboptimal GDMT use following new-onset HFrEF was associated with an increased risk of mortality and subsequent HF hospitalization.

Juice yield and pectin indicators in apple and carrot pomace

Background: Pectin exhibits different properties based on its degree of esterification, forming gels and serving as a structure-forming agent in food products, or forming complexes with heavy metal ions, finding applications in the development of therapeutic products for the elimination of heavy metal ions from the human body. Context and purpose of this study: The purpose of this study is to investigate the influence of enzymatic treatment on the yield and quality of apple and carrot juices, as well as the quality parameters of pectin in pomace. Our objectives were to select an enzymatic preparation for processing apple and carrot pomace and to study changes in the properties of pectins present in the pomace depending on the dosage of the enzyme preparation.Results: The results of our experiments revealed that for apple pomace processing, it is more beneficial to use the enzyme preparation Vegazym M with a dosage of 0.09% of the pomace mass, while for carrot pomace processing, the enzyme preparation Fructozym MA with a dosage of 0.07% of the pomace mass proved to be the most suitable. These enzyme preparations in the specified amounts ensure maximum juice yield. It has been established that changing the dose of enzymes changes the complexing properties of pectin, which allows its use in food products with therapeutic and prophylactic properties.Conclusions: The enzyme preparations allow for specific modifications, such as pectin modification in this case, enabling the versatile use of pectin as either a structure-forming agent or for the development of therapeutic and preventive food products.Keywords:Apple and carrot juices, enzyme preparations, apple and carrot pomace, pectin, soluble and insoluble pectin, degree of esterification and pectin complexation, functional products.

Protein production from cellulosic waste using candida utilis

With rapid population growth there is a lot of interest in developing alternative protein sources, like protein yeast cultures. For growth of yeast could be used agricultural and food processing waste, but such source of hydrocarbons need to be pretreated. The purpose of the presented study is to assess the effect of steam explosion treatment on the process of feed protein production, including the influence on the enzymatic hydrolysis, cultivation of yeast and resulting products characteristics. Wheat straw, spelt hull and sunflower husk were subjected to steam explosive activation at 165 °C and 210 °C. The products were subjected to enzymatic hydrolysis at the following conditions: enzyme dose 12 g/kg of raw material and temperature of 45 °C. Total reducing sugars content and component composition were evaluated in the hydrolysates using IR spectrometry and HPLC respectively. A pure yeast culture of Candida utilis, was grown on hydrolysates (2 g/L) in a 10 L bioreactor unit at 35 °C for 24 h. The samples of protein were evaluated for crude protein, true protein, fiber, fat and ash content. Conversion efficiency was calculated. The highest conversion efficiency was observed for spelt husks activated at 165 °C: reducing sugars yield was 56.2 % out of 73.3 % of theoretical yield; biomass concentration reaches 52.9 g/L, with reducing sugar conversion of 80.1 %. The crude protein content of yeast was 53.9 % (42 % according to Barnstein). Hydrolysates from activated materials, give a higher yeast biomass yield of 21–42 % compared to hydrolysates of non-activated raw materials.

Bayesian-optimized random forest prediction of key properties of micro-/nanofibrillated cellulose from different woody and non-woody feedstocks

The scale-up of nanocellulose production, a still challenging task, could benefit greatly from machine learning. In this paper, random forest regressors were applied to predict two important parameters for nanocellulose processing: nanofibrillation yield and aspect ratio. The dataset comprised diverse pulp sources, ranging from non-woody and woody feedstock, and two different types of pulp pre-treatments, namely mechanical refining and enzymatic hydrolysis. The obtained models were evaluated in terms of number of features and optimized with regard to some important hyperparameters, using the Bayesian optimization function from the SkOpt library. Considering the heterogeneity of the data, the prediction of both aspect ratio and yield of nanofibrillation was deemed satisfactory. For the former, R² scores of 0.986, 0.902 and 0.932 were obtained for the training, validation and testing subsets, respectively. Regarding the latter, R² scores of 0.994, 0.947 and 0.877 were achieved for the same types of subsets. Other important remarks are that enzyme dosage was the most important feature for predicting aspect ratio, while it did not play an important role for yield modelling. For this output, the transmittance at 600 nm accounted for almost 90% of the total feature importance.

Cellulase immobilization on zeolitic imidazolate frameworks for boosting cellulose hydrolysis at high solids loading

This study investigates the cellulase immobilization onto zeolitic imidazolate frameworks (ZIFs) to enable efficient cellulose hydrolysis at high solids loading. Different ZIF carriers (ZIF-8, ZIF-8-NH2, Fe3O4@ZIF-8, and Fe3O4@ZIF-8-NH2) were prepared and characterized. The resulting ZIF-immobilized cellulases were subsequently analyzed for their loading capacity and enzyme activity, as well as their biochemical properties such as temperature stability, pH stability, storage stability, enzyme leaching, and enzyme kinetics. Results revealed that the cellulase immobilized on Fe3O4@ZIF-8-NH2 exhibited an exceptional loading capacity of 359.89 mg/g and a relative enzyme activity of 69.39%. Remarkably, at a solid loading of 15% (w/w) and an enzyme dosage of 5 FPU/g cellulose, the Fe3O4@ZIF-8-NH2 immobilized cellulase demonstrated a 20.44% higher hydrolysis yield compared to the free cellulase. Furthermore, this immobilized enzyme system displayed robust reusability, retaining 71.03% of its initial activity even after 10 cycles of reuse. A proposed mechanism of enhanced enzymatic hydrolysis by ZIF-immobilized cellulases was also presented. These findings have significant implications for the future application of ZIF-immobilized cellulases in efficient and cost-effective lignocellulosic bioconversion.

Process optimization for saccharification and fermentation of the Organic Fraction of Municipal Solid Waste (OFMSW) to maximize ethanol production performance

The Organic Fraction of Municipal Solid Waste (OFMSW) contains paper and cardboard, an attractive cellulosic feedstock for bioethanol production. Waste paper/cardboard (WPC) was subjected to mild acid treatment in solids loadings from 20 to 27.5 %. Different doses of commercial enzyme cocktail and various adjuvants facilitating enzymatic saccharification were tested. Enzyme dosing of 3.75 FPU/g fiber with very slow fed-batch saccharification of 27.5 % (w/v) solids from model lignin-free paper pulp resulted in 12.6 % (w/v) total sugar after 74 h. Use of cheap non-catalytic waste protein adjuvants including Soymeal, Protisyr, Feather meal strongly improved enzymatic saccharification and reduced the overall process cost by reducing commercial enzyme dosing. After fermentation with the industrial yeast strain BMD44 (Cellusec® 1.0) an ethanol titer of 5.77 % (v/v) was obtained using a reduced enzyme loading of 2.5 FPU/g of actual industrial WPC fiber. These results show the potential of economically profitable second-generation bioethanol production with WPC fiber.

PSIV-20 An in Vitro Digestion Procedure Using Daisyii Incubator Can Replace Conventional in Vitro Digestion Procedures Using Flasks for Estimating Ileal Digestibility of Feed Ingredients for Pigs

Abstract In vitro procedures using flasks have been used for simulating nutrient utilization by pigs. However, these procedures are time-consuming and laborious and require a number of flasks and filter crucibles. The objective of this study was to develop an in vitro procedure using DaisyII incubator for simultaneous analysis of multiple samples. In a conventional in vitro procedure, 10 mg of pepsin in step 1 and 50 mg of pancreatin in step 2 are used for each flask containing 1 g of feed sample. In a preliminary experiment, 24 samples each in R55 bags were digested in a jar of DaisyII incubator with 24 times the quantity of buffer solution and enzymes of the conventional flask procedure. In the preliminary experiment, greater crude protein (CP) digestibility values were observed in the DaisyII incubator procedure compared with the conventional flask procedure. To determine optimum enzyme doses in the in vitro ileal disappearance (IVIDDaisy) procedure using the DaisyII incubator, 3 doses of pepsin (10, 7.5, and 5 mg/sample in step 1) and 3 doses of pancreatin (50, 37.5, and 25 mg/sample in step 2) were tested in a 3 × 3 factorial arrangement for 11 test ingredients. The quantity of buffer solution and enzymes in a jar was increased proportionally with the number of samples. The 11 test ingredients were barley, copra meal, corn, corn distillers dried grains with solubles, corn gluten feed, corn gluten meal, palm kernel meal, rice bran, soybean hulls, soybean meal, and wheat bran. The in vitro ileal disappearance (IVIDflask) of CP in the 11 ingredients was determined using the conventional flask procedure. The sum of square of deviation between IVIDflask of CP and IVIDDaisy of CP was the least when 5 mg pepsin/sample in step 1 and 37.5 mg/sample in step 2 were used (IVIDflask of CP, % = 14.4 + 0.81 × IVIDDaisy of CP, %; r2 = 0.748). On the other hand, the determination coefficient was the greatest when 10 mg pepsin/sample in step 1 and 50 mg/sample in step 2 were used (IVIDflask of CP, % = 2.5 + 0.93 × IVIDDaisy of CP, %; r2 = 0.878). In conclusion, the in vitro procedure using a DaisyII incubator can be used for estimating ileal digestibility of protein in multiple samples simultaneously for pigs.

LPMO-supported saccharification of biomass: effects of continuous aeration of reaction mixtures with variable fractions of water-insoluble solids and cellulolytic enzymes

BackgroundHigh substrate concentrations and high sugar yields are important aspects of enzymatic saccharification of lignocellulosic substrates. The benefit of supporting the catalytic action of lytic polysaccharide monooxygenase (LPMO) through continuous aeration of slurries of pretreated softwood was weighed against problems associated with increasing substrate content (quantitated as WIS, water-insoluble solids, in the range 12.5–17.5%), and was compared to the beneficial effect on the saccharification reaction achieved by increasing the enzyme preparation (Cellic CTec3) loadings. Aerated reactions were compared to reactions supplied with N2 to assess the contribution of LPMO to the saccharification reactions. Analysis using 13C NMR spectroscopy, XRD, Simons’ staining, BET analysis, and SEM analysis was used to gain further insights into the effects of the cellulolytic enzymes on the substrate under different reaction conditions.ResultsAlthough glucose production after 72 h was higher at 17.5% WIS than at 12.5% WIS, glucan conversion decreased with 24% (air) and 17% (N2). Compared to reactions with N2, the average increases in glucose production for aerated reactions were 91% (12.5% WIS), 70% (15.0% WIS), and 67% (17.5% WIS). Improvements in glucan conversion through aeration were larger (55–86%) than the negative effects of increasing WIS content. For reactions with 12.5% WIS, increased enzyme dosage with 50% improved glucan conversion with 25–30% for air and N2, whereas improvements with double enzyme dosage were 30% (N2) and 39% (air). Structural analyses of the solid fractions revealed that the enzymatic reaction, particularly with aeration, created increased surface area (BET analysis), increased disorder (SEM analysis), decreased crystallinity (XRD), and increased dye adsorption based on the cellulose content (Simons' staining).ConclusionsThe gains in glucan conversion with aeration were larger than the decreases observed due to increased substrate content, resulting in higher glucan conversion when using aeration at the highest WIS value than when using N2 at the lowest WIS value. The increase in glucan conversion with double enzyme preparation dosage was smaller than the increase achieved with aeration. The results demonstrate the potential in using proper aeration to exploit the inherent capacity of LPMO in enzymatic saccharification of lignocellulosic substrates and provide detailed information about the characteristics of the substrate after interaction with cellulolytic enzymes.

Hyper xylanase production and potential of xylooligosaccharides formation from a novel Bacillus australimaris KS2

The study has been designed to isolate xylanase-producing bacteria and parametric optimization for maximum production. The xylanase was utilized for enzymatic hydrolysis of wheat bran (WB) and wheat bran extracted xylan (WBEx) to obtain xylooligosaccharides. The research was planned for value addition in the agriculture sector with low-cost enzyme production and its utilization in prebiotic production from agro-residue. The xylanase obtained from the new isolate Bacillus australimaris KS2 was optimized with 2.0% of 24 h inoculum supplemented with 2.0% wheat bran at pH 7, 35 °C for 48 h with an agitation rate of 150 rpm. There was a 2.96-fold increase in xylanase activity from its initial (2997.07 IU/mL) activity to the optimized condition (8873.07 IU/mL). The maximum reducing sugar production over enzymatically hydrolyzed WB was found after 8 h of reaction time at pH 5, 55 °C with 375 units of enzyme. In the case of enzymatically hydrolyzed WBEx, the maximum production of 588 mg/g was achieved after 16 h at pH 6 and 55 °C of reaction temperature at the same amount of enzyme dose. The results demonstrated that the new isolate Bacillus australimaris KS2 was an efficient xylanase producer. Xylooligosaccharides produced through enzymatic hydrolysis of WBEx showed good prebiotic potential over Lactobacillus plantarum, Lactobacillus brevis, and Lactobacillus rhamnosus.

Substrate-related factors and kinetic studies of Carbohydrate-Rich food wastes on enzymatic saccharification

Food waste biorefinery is a sustainable approach to producing green chemicals, however the essential substrate-related factors hindering the efficacy of enzymatic hydrolysis have never been clarified. This study explored the key rate-limiting parameters and mechanisms of carbohydrate-rich food after different cooking and storing methods, i.e., impacts of compositions, structural diversities, and hornification. Shake-flask enzymatic kinetics determined the optimal dosages (0.5 wt% glucoamylase, 3 wt% cellulase) for food waste hydrolysis. First order kinetics and simulation results determined that reaction coefficient (K) of cooked starchy food was ∼ 3.63 h−1 (92 % amylum digestibility) within 2 h, while those for cooked cellulosic vegetables were 0.25–0.5 h−1 after 12 h of hydrolysis. Drying and frying reduced ∼ 71–89 % hydrolysis rates for rice, while hydrothermal pretreatment increased the hydrolysis rate by 82 % on vegetable wastes. This study provided insights into advanced control strategy and reduced the operational costs by optimized enzyme doses for food waste valorization.