Combination Of Enzymes Research Articles

Enzymatization of Nila Fish (Oreochromis Niloticus) Protein Hydolysate by Combination of Bromelin and Pepsin Enzymes

Nile Tilapia (Oreochromis niloticus) is a freshwater fish with better nutritional content compared to other freshwater fish, making it suitable for use as a source for protein hydrolysate production. Protein hydrolysate is the result of the breakdown of protein into short-chain compounds through hydrolysis. This research aims to hydrolyze the minced Nile tilapia meat using a combination of bromelain and pepsin enzymes at a concentration of 5% and pH 7 conditions for 5-6 hours. The hydrolysis results using the combination of bromelain and pepsin enzymes (B100: P0) have a yield value of 12.68%, ash content of 2.94%, protein of 57.25%, water 2.65%, and fat 10.25% with a degree of hydrolysis of 61.46%. The hydrolysate has antibacterial properties against Staphylococcus aureus and Escherichia coli bacteria with inhibition zones of approximately 8.3 mm and 8.5 mm, respectively, at a concentration of 1 (mg/µL), contains 15 types of amino acids with the highest composition being lysine 5.12% and the highest non-essential amino acid being aspartic acid 5.75%.

A pilot study to directly estimate radiation-induced mutation in large Japanese field mouse duo sample, mother and offspring, excluding unknown father, using ddRAD sequencing.

DNA mutations are one of the effects of radiation exposure. A large amount of radioactive materials was released into the environment from the Fukushima Daiichi Nuclear Power Plant after a major earthquake and tsunami. Wild animals and plants living in highly radiation-contaminated areas are constantly exposed to high doses of radiation, and concerns occur about its effects on their health and the next generations. As a pilot study, double-digest restriction site-associated DNA (ddRAD) sequencing was conducted to assess the incidence of mutations in wild large Japanese field mice collected from the evacuation area. The optimal combination of restriction enzymes, encompassing the functionally important coding regions, was selected using in silico analysis. These enzymes were used for ddRAD sequence analysis of females and their fetuses to evaluate mutation rates. The results indicated that no significant differences were observed in mutation rates between mothers and fetuses in the study areas.

Structure-Agnostic Bioactivity-Driven Combinatorial Biosynthesis Reveals New Antidiabetic and Anticancer Triterpenoids.

Although combinatorial biosynthesis can dramatically expand the chemical structures of bioactive natural products to identify molecules with improved characteristics, progress in this direction has been hampered by the difficulty in isolating and characterizing the numerous produced compounds. This challenge could be overcome with improved designs that enable the analysis of the bioactivity of the produced metabolites ahead of the time-consuming isolation procedures. Herein, we showcase a structure-agnostic bioactivity-driven combinatorial biosynthesis workflow that introduces bioactivity assessment as a selection-driving force to guide iterative combinatorial biosynthesis rounds towards enzyme combinations with increasing bioactivity. We apply this approach to produce triterpenoids with potent bioactivity against PTP1B, a promising molecular target for diabetes and cancer treatment. We demonstrate that the bioactivity-guided workflow can expedite the combinatorial process by enabling the narrowing down of more than 1000 possible combinations to only five highly potent candidates. By focusing the isolation and structural elucidation effort on only these five strains, we reveal 20 structurally diverse triterpenoids, including four new compounds and a novel triterpenoid-anthranilic acid hybrid, as potent PTP1B inhibitors. This workflow expedites hit identification by combinatorial biosynthesis and is applicable to many other types of bioactive natural products, therefore providing a strategy for accelerated drug discovery.

Unprecedented N2O production by nitrate-ammonifying Geobacteraceae with distinctive N2O isotopocule signatures.

Dissimilatory nitrate reduction to ammonium (DNRA), driven by nitrate-ammonifying bacteria, is an increasingly appreciated nitrogen-cycling pathway in terrestrial ecosystems. This process reportedly generates nitrous oxide (N2O), a strong greenhouse gas with ozone-depleting effects. However, it remains poorly understood how N2O is produced by environmental nitrate-ammonifiers and how to identify DNRA-derived N2O. In this study, we characterize two novel enzymatic pathways responsible for N2O production in Geobacteraceae strains, which are predominant nitrate-ammonifying bacteria in paddy soils. The first pathway involves a membrane-bound nitrate reductase (Nar) and a hybrid cluster protein complex (Hcp-Hcr) that catalyzes the conversion of NO2- to NO and subsequently to N2O. The second pathway is observed in Nar-deficient bacteria, where the nitrite reductase (NrfA) generates NO, which is then reduced to N2O by Hcp-Hcr. These enzyme combinations are prevalent across the domain Bacteria. Moreover, we observe distinctive isotopocule signatures of DNRA-derived N2O from other established N2O production pathways, especially through the highest 15N-site preference (SP) values (43.0‰-49.9‰) reported so far, indicating a robust means for N2O source partitioning. Our findings demonstrate two novel N2O production pathways in DNRA that can be isotopically distinguished from other pathways.IMPORTANCEStimulation of DNRA is a promising strategy to improve fertilizer efficiency and reduce N2O emission in agriculture soils. This process converts water-leachable NO3- and NO2- into soil-adsorbable NH4+, thereby alleviating nitrogen loss and N2O emission resulting from denitrification. However, several studies have noted that DNRA can also be a source of N2O, contributing to global warming. This contribution is often masked by other N2O generation processes, leading to a limited understanding of DNRA as an N2O source. Our study reveals two widespread yet overlooked N2O production pathways in Geobacteraceae, the predominant DNRA bacteria in paddy soils, along with their distinctive isotopocule signatures. These findings offer novel insights into the role of the DNRA bacteria in N2O production and underscore the significance of N2O isotopocule signatures in microbial N2O source tracking.

Effective drug design screening in bacterial glycolytic enzymes via targeting alternative allosteric sites

Three glycolytic enzymes phosphofructokinase (PFK), glyceraldehyde-3-phosphate dehydrogenase (GADPH) and pyruvate kinase (PK) that belong to Staphylococcus aureus were used as targets for screening a dataset composed of 7229 compounds of which 1416 were FDA-approved. Instead of catalytic sites, evolutionarily less conserved allosteric sites were targeted to identify compounds that would selectively bind the bacteria's glycolytic enzymes instead of the human host. Seven different allosteric sites provided by three enzymes were used in independent screening experiments via docking. For each of the seven sites, a total of 723 compounds were selected as the top 10 % which displayed the highest binding affinities. All compounds were then united to yield the top 54 drug candidates shared by all seven sites. Next, 17 out of 54 were selected and subjected to in vitro experiments for testing their inhibition capability for antibacterial growth and enzymatic activity. Accordingly, four compounds displaying antibacterial growth inhibition above 40 % were determined as Candesartan cilexetil, Montelukast (sodium), Dronedarone (hydrochloride) and Thonzonium (bromide). In a second round of experiment, Candesartan cilexetil and Thonzonium displayed exceptionally high killing efficiencies on two bacterial strains of S.aureus (methicillin-sensitive and methicillin-resistant) with concentrations as low as 4 μg/mL and 0.5 μg/mL. Yet, their enzymatic assays were not in accordance with their killing effectiveness. Different inhibitory effects was observed for each compound in each enzymatic assay. A more effective target strategy would be to screen for drug compounds that woud inhibit a combination of glycolytic enzymes observed in the glycolytic pathway.

The Impact of the Combination of Proteolytic Enzyme and Rutin on the Post-operative Management of Inflammation: A Randomized, Controlled, Double-Blind, Comparative Clinical Trial

The Impact of the Combination of Proteolytic Enzyme and Rutin on the Post-operative Management of Inflammation: A Randomized, Controlled, Double-Blind, Comparative Clinical Trial

INVESTIGATION OF THE EFFECT OF ENZYME APPLICATION ON THE STRUCTURE OF WAFERS IN THE FOOD INDUSTRY

The study aimed to assess the impact of 13 different enzymes, including protease, amylase, hemicellulase, and xylanase, on wafer dough and sheets, aiming to identify the most effective enzyme combination. Commercial protease, amylase, hemicellulase, and xylanase enzymes were applied to wafer dough following manufacturer instructions, and their flow behaviors were analyzed. Subsequently, the doughs were baked into wafer sheets, and various parameters such as water activity (aw), moisture content (%), weight loss (%), color parameters (L*, a*, b*), hardness (g), and sensory attributes were evaluated. Protease 4, Hemicellulase 2, and Xylanase 2 enzymes were selected for combination testing based on the analysis of flow behavior and characterization parameters of the wafer doughs. Among the tested combinations, the wafer dough containing 80 ppm Xylanase 2 + 300 ppm Protease 4 enzyme group exhibited promising potential for industrial applications due to its favorable flow behavior, while the resulting wafer sheets demonstrated desirable crispness and a brown-caramel color, suggesting suitability for commercial use.

Large-scale screening identifies enzyme combinations that remove in situ grown oral biofilm

Bacteria in the oral cavity are responsible for the development of dental diseases such as caries and periodontitis, but it is becoming increasingly clear that the oral microbiome also benefits human health. Many oral care products on the market are antimicrobial, killing a large part of the oral microbiome but without removing the disease-causing biofilm. Instead, non-biocidal matrix-degrading enzymes may be used to selectively remove biofilm without harming the overall microbiome.The challenge of using enzymes to degrade biofilms is to match the narrow specificity of enzymes with the large structural diversity of extracellular polymeric substances that hold the biofilm together. In this study, we therefore perform a large-scale screening of single and multi-enzyme formulations to identify combinations of enzymes that most effectively remove dental biofilm.We tested >400 different treatment modalities using 44 different enzymes in combinations with up to six enzymes in each formulation, on in vitro biofilms inoculated with human saliva. Mutanase was the only enzyme capable of removing biofilm on its own. Multi-enzyme formulations removed up to 69 % of the biofilm volume, and the most effective formulations all contained mutanase. We shortlisted 10 enzyme formulations to investigate their efficacy against biofilms formed on glass slabs on dental splints worn by 9 different test subjects. Three of the ten formulations removed more than 50 % of the biofilm volume. If optimal enzyme concentration and exposure time can be reached in vivo, these enzyme combinations have potential to be used in novel non-biocidal oral care products for dental biofilm control.

Enzyme-assisted hydrolysis of Theobroma cacao L. pulp

Cocoa pulp, a by-product of the cocoa supply chain, has gained the food sector’s attention for its pleasant flavour. Enzyme-assisted technologies to facilitate its separation from the cocoa seeds remain largely unexplored. We studied the effects of temperature, enzyme activity and enzyme combinations on the physicochemical properties of cocoa pulp by Response Surface Methodology and D-optimal design. Endo-polygalacturonase, endo-cellulase and hemicellulase activities were investigated. A reduced quadratic model described the relative reduction in viscosity, of the particle diameter dv,0.9 and of the browning index. Polygalacturonase had the strongest influence on viscosity, while cellulase had the least effect. Synergistic effects of polygalacturonase with cellulase and/or hemicellulase were identified. The model predicted maximal reductions in viscosity by 70.3% with 275 U of polygalacturonase and 275 U of hemicellulase at 40 °C. Fresh pulp’s dv,0.9 was 613 μm. Combining all enzyme activities reduced the dv,0.9 to 418 μm (40 °C, total activity: 580 U). Only the temperature proved to significantly influence the browning index, with higher temperatures causing a higher colouring effect. Lastly, the model was successfully validated for the relative reduction in viscosity and the dv,0.9. Our study provides key insights into the enzyme-assisted processing of cocoa pulp, facilitating its separation from the seeds. By utilising the pulp, innovative ingredients can be developed for the food sector, and at the same time, the added value in the cocoa supply chain can be improved.

Epidemiology of heart failure in France

BackgroundHeart failure (HF) prevalence may increase because of population ageing and has become a major public health issue in European countries. AimTo update the epidemiology of HF in France in 2022. MethodsAdults hospitalized for HF in 2022 were identified in the National Health Data System (SNDS) and followed up for 1year. The first stay of the year was taken as the index hospitalization. The prevalence of HF was estimated by combining hospitalization data and patients with 100% coverage for a long-term disease associated with HF. Patients and their hospital stays were described on the basis of the sociodemographic and medical information in the SNDS. ResultsIn 2022, 181,178 adults were hospitalized for HF in France, which equates to a crude rate of 339.3 per 100,000 inhabitants, and 1,376,692 prevalent cases of HF were recorded, which is an estimated prevalence of 2.6% in the adult population. For people living in the most socioeconomically deprived municipalities, the rate of hospitalization was 1.6 times higher than for those living in the least deprived municipalities. The departments of Haut-de-France and Réunion Island, and some departments in Normandy and the Grand-Est had much higher rates than others. The fatality rate was 10.2% in hospital, and 34.0% at 1year. Only 20.1% of patients were admitted to a rehabilitation unit within 6months, and 47.9% of patients alive at 1year were being treated with a combination of angiotensin-converting enzyme inhibitors/angiotensin II receptor blockers and beta-blockers. ConclusionsThe large number of people hospitalized for HF, and the fact that rates vary across the different French departments, means that more ambitious general cardiovascular prevention measures are needed, and that healthcare provision needs significant adaptation. Short-term patient outcomes could be improved by following recommendations more closely and taking into account patients’ social circumstances.

Enhancing the Quality of Ready‐to‐Eat Trout Burger Patties: Investigating the Combination of Modified Atmosphere Packaging and Microbial Transglutaminase Enzyme

ABSTRACTIn this study, rainbow trout patties (Oncorhynchus mykiss Walbaum, 1792) were treated under a modified atmosphere (MA) (60/40:CO2/N2) packed with microbial transglutaminase enzyme (MTGase) at varying concentrations (0.5% and 1%) and kept for a full day at 2°C ± 2°C. Denser and more complex intermolecular cross‐linking formations between peptides and gel network structures were observed in the first and last scanning electron microscope images of MTGase‐containing groups. MTGase increased the springiness, hardness, shear force, and work‐of‐shear parameters during the storage period in uncooked and cooked patties compared to the control group. The protein, energy, moisture, and amino acid contents were maintained in the MTGase‐containing groups, but using MTGase at higher concentrations was ineffective. Microbiological analyses have shown that MTGase is partially effective on microbial fauna, and the most significant inhibitory effect was determined on total mesophilic aerobic bacteria (TMAB). In all groups, the total amounts of saturated, monosaturated, and polyunsaturated fatty acids remained unchanged on both production and expiration days (p > 0.05). After mixing the burger patty additives into the ground meat, the amounts of sodium, calcium, potassium, and magnesium increased in all groups (p < 0.05); however, a decrease in calcium was observed in the MTGase‐containing groups (p < 0.05) compared to the control group. The higher proportion of the MTGase‐contained group did not exhibit significant differences compared to the lower proportion of the MTGase‐contained group, except for textural characteristics and the TMAB inhibition effect.

Exploring the Physicochemical Characteristics of Marine Protein Hydrolysates and the Impact of In Vitro Gastrointestinal Digestion on Their Bioactivity.

Fish protein hydrolysates (FPHs) were obtained from different fish sources using a combination of microbial enzymes. The industrially produced FPHs from blue whiting (Micromesistius poutassou) and sprat (Sprattus sprattus) were compared to freeze-dried FPHs generated in-house from hake (Merluccius merluccius) and mackerel (Scomber scombrus) in terms of their physicochemical composition and functionality. Significant differences (p < 0.05) were observed in the protein, moisture, and ash contents of the FPHs, with the majority having high levels of protein (73.24-89.31%). Fractions that were more extensively hydrolysed exhibited a high solubility index (74.05-98.99%) at different pHs. Blue whiting protein hydrolysate-B (BWPH-B) had the highest foaming capacity at pH 4 (146.98 ± 4.28%) and foam stability over 5 min (90-100%) at pH 4, 6, and 8. The emulsifying capacity ranged from 61.11-108.90 m2/g, while emulsion stability was 37.82-76.99% at 0.5% (w/v) concentration. In terms of peptide bioactivity, sprat protein hydrolysate (SPH) had the strongest overall reducing power. The highest Cu2+ chelating activity was exhibited by hake protein hydrolysate (HPH) and mackerel protein hydrolysate (MPH), with IC50 values of 0.66 and 0.78 mg protein/mL, respectively, while blue whiting protein hydrolysate-A (BWPH-A) had the highest activity against Fe2+ (IC50 = 1.89 mg protein/mL). SPH scavenged DPPH and ABTS radicals best with IC50 values of 0.73 and 2.76 mg protein/mL, respectively. All FPHs displayed noteworthy scavenging activity against hydroxyl radicals, with IC50 values ranging from 0.48-3.46 mg protein/mL. SPH and MPH showed the highest scavenging potential against superoxide radicals with IC50 values of 1.75 and 2.53 mg protein/mL and against hydrogen peroxide with 2.22 and 3.66 mg protein/mL, respectively. While inhibition of α-glucosidase was not observed, the IC50 values against α-amylase ranged from 8.81-18.42 mg protein/mL, with SPH displaying the highest activity. The stability of FPHs following simulated gastrointestinal digestion (SGID) showed an irregular trend. Overall, the findings suggest that marine-derived protein hydrolysates may serve as good sources of natural nutraceuticals with antioxidant and antidiabetic properties.

Action enhancement of antimicrobial peptides by their combination with enzymes hydrolyzing fungal quorum molecules

Recently, the lactonase activity of several enzymes (lactonase AiiA, organophosphate hydrolase (His6-OPH) and New Delhi metallo-β-lactamase (NDM-1)) was revealed in the hydrolysis of lactone-containing fungal Quorum Sensing molecules (FQSM). This study was aimed at the investigation of possible use of these enzymes as components of antifungal combinations with antimicrobial peptides (AMPs) to increase their action efficiency against various fungi. For this, the interaction of various AMPs with AiiA, NDM-1 or His6-OPH, as well as the effect of AMPs on the catalytic characteristics of these enzymes in the hydrolysis of FQSM in enzyme/AMP combinations, were studied using in silico computer modeling methods. Enzymes combinations with 3 AMPs Bacitracin, Colistin and Polymyxin B were selected as the most rational in terms of maintaining the effectiveness of AMP and the catalytic activity of enzymes. The antifungal action of the selected combinations against cells of mycelial fungi and yeast was studied in vitro. It was found that combinations of the enzymes AiiA, His6-OPH and NDM-1 with Bacitracin, Colistin and Polymyxin B provide a significant increase in the action efficiency (up to 5000 times) of both AMPs and enzymes against fungi. The most effective variants were obtained for Polymyxin B in multicomponent combinations with enzymes.

Structure‐Agnostic Bioactivity‐Driven Combinatorial Biosynthesis Reveals New Antidiabetic and Anticancer Triterpenoids

Although combinatorial biosynthesis can dramatically expand the chemical structures of bioactive natural products to identify molecules with improved characteristics, progress in this direction has been hampered by the difficulty in isolating and characterizing the numerous produced compounds. This challenge could be overcome with improved designs that enable the analysis of the bioactivity of the produced metabolites ahead of the time‐consuming isolation procedures. Herein, we showcase a structure‐agnostic bioactivity‐driven combinatorial biosynthesis workflow that introduces bioactivity assessment as a selection‐driving force to guide iterative combinatorial biosynthesis rounds towards enzyme combinations with increasing bioactivity. We apply this approach to produce triterpenoids with potent bioactivity against PTP1B, a promising molecular target for diabetes and cancer treatment. We demonstrate that the bioactivity‐guided workflow can expedite the combinatorial process by enabling the narrowing down of more than 1000 possible combinations to only five highly potent candidates. By focusing the isolation and structural elucidation effort on only these five strains, we reveal 20 structurally diverse triterpenoids, including four new compounds and a novel triterpenoid‐anthranilic acid hybrid, as potent PTP1B inhibitors. This workflow expedites hit identification by combinatorial biosynthesis and is applicable to many other types of bioactive natural products, therefore providing a strategy for accelerated drug discovery.

The Combination of Proteolytic Enzyme Supplementation, Acupuncture, and Osseous Manipulation for the Treatment of Traumatic Peripheral Nerve Injury: A Case Report

This report describes the treatment of peripheral nerve injury in a 43-year-old woman using a combination of proteolytic enzymes, acupuncture, and osseous manipulation as alternatives to conventional care. Her presenting symptoms include complete loss of sensation in the superolateral left thigh and a mass of fibrotic scar tissue in the area of injury. A holistic treatment plan was created to address the underlying mechanisms of the injury. Plant-based proteolytic enzymes, bromelain and papain, were used to address tissue inflammation and reduce scar tissue formation around the nerve, acupuncture was used to regenerate the superficial nerves, and osseous manipulations were provided for structural re-alignment. After 8 weeks of treatment, the patient reported significant reduction in the size of the fibrotic mass and complete resolution of sensory loss. The fibrotic mass of tissue reduced from 6 cm to 1 cm during the treatment course. Proteolytic enzymes have wide-ranging indications; however, they have not been well studied for peripheral nerve injury, which makes this a novel indication for this natural health product. This case report found the use of proteolytic enzymes, and acupuncture, effective in treating peripheral nerve injury and provides grounds for research to treat nerve-related injuries.

PSVI-29 Effect of pig weight and exogenous enzyme supplementation on ileal endogenous amino acid losses and standardized ileal amino acid digestibility of soybean meal produced from Kentucky-grown soybeans

Abstract There is increasing interest in using locally sourced soybeans and soybean meal. Ileal amino acid digestibility (IAAD) of soybean meal from Kentucky-grown soybeans was evaluated in 25- and 50-kg cannulated pigs. The experiment used sixteen pigs in a randomized complete block design consisting of four treatments with 4 pigs per treatment. This was repeated in time resulting in eight replicates per treatment for both body weight (BW)groups. After cannulation at about 20 kg BW, all pigs were allowed to recover from the surgery and were fed a corn-SBM-based diet that met all nutrient and energy requirements. The experimental diets consisted of two nitrogen-free diets [NFD] with or without exogenous enzyme supplementation (phytase and a combination of phytase and carbohydrase enzymes) and two semi-purified diets containing soybean meal from Kentucky-grown soybeans as the only source of protein with or without exogenous enzyme supplementation. Each diet was fed for 7 consecutive d with 10-h ileal digesta collection on d 6 and 7. The same diets were fed to the same set of pigs at about 50 kg BW. Data were analyzed as a factorial arrangement of treatments (BW x diet) for ileal endogenous amino acid losses (IEAAL), apparent IAAD and standardized IAAD (SIAAD). Data from the NFD were used to determine the SIAAD from the AIAAD values. Exogenous enzyme supplementation increased (P &amp;lt; 0.05) IEAAL except for Gly and Pro while samples collected during phase 2 had greater (P &amp;lt; 0.05) IEAAL (10 amino acids). Pig BW did not influence IEAAL. Enzyme supplementation did not increase AIAAD except for Arg, Ala, Gly, N, and total AA while apparent ileal N, Met, Thr, and Val digestibility was greater (P &amp;lt; 0.05) in 50 kg pigs. Interactions between BW and diets showed that 50 kg pigs fed the SBM diet containing enzyme supplementation had greater (P &amp;lt; 0.05) N, Arg, and total AA digestibility compared with 50 kg pigs without enzyme supplementation. Digestibility of Val and Asp was greater (P &amp;lt; 0.05) in samples collected during phase 2. After correction for basal IEAAL, 50 kg pigs had greater (P &amp;lt; 0.05) SIAAD values for 12 of the AA analyzed while exogenous enzyme supplementation resulted in greater (P &amp;lt; 0.05) SIAAD for 9 of the AA analyzed. The SIAAD values for four AA (His, Phe, Val, and Asp) were greater (P &amp;lt; 0.05) for samples collected in phase 2. Results from this study showed that exogenous enzyme supplementation increased IEAAL and that IAAD of SBM produced from Kentucky-cultivated soybeans was high and that exogenous enzyme supplementation improved the SIAAD values of several amino acids. The IAAD values of SBM produced from soybeans cultivated in KY was high and are similar to values reported in the literature.

PSXIII-22 Intake, digestibility, and ruminal parameters of Nellore cattle fed with amylolytic and fibrolytic enzymes in feedlot

Abstract This study was carried out to evaluate the use of dietary exogenous enzyme (ENZ) products with amylolytic and fibrolytic activity on nutrient intake, digestibility, and ruminal volatile fatty acids production (VFA) of Nellore cattle in feedlot. Rumen-cannulated Nellore steers [n = 10; body weight (BW) = 543 kg ± 28.6 BW] were distributed in a replicated Latin-square design (5 × 5) in individual pens. The treatments consisted of a negative control (CON; without ENZ); Amylase (AML; Amaize, Alltech, 0.5 g/kg of DM); Xylanase (FBL, Fibrozyme, Alltech, 0.9 g/kg of DM); Half dose (HD; AML 0.25 g/kg of diet DM and FBL 0.45 g/kg of diet DM); and Full dose (FD; AML 0.5 g/kg of diet DM and FBL 0.90 g/kg of diet DM; Table 1). The experimental period lasted 19 d (14 d of adaptation and 5 d of collection). Between d 15 and 18, total fecal collection was performed and on d 19 rumen fluid was collected at 0, 4, 8, 12, and 16 h post-feeding for ammonia, VFA, and pH determination. The data were analyzed as orthogonal contrasts using SAS: (C1) CON vs. ENZ, (C2) AML vs. FBL, (C3) HD vs. FD, and (C4) AML + FBL vs. HD + FD. There were no C1 differences for nutrient digestibility, total VFA, or ruminal pH (P &amp;gt; 0.10). However, a tendency effect in neutral detergent fiber (NDF) intake (P = 0.065) was observed, where animals fed ENZ had reduced values compared with animals kept in CON group. In the same way, ENZ reduced ther extract (EE) intake (P = 0.023), ruminal ammonia (P &amp;lt; 0.001), and isovalerate proportion (P = 0.080) as well as increased proportion of acetate (P &amp;lt; 0.001), valerate (P &amp;lt; 0.001) and acetate:propionate ratio (P = 0.030). No differences were observed in nutrient intake between AML and FBL (P &amp;gt; 0.10), but AML had greater organic matter (OM) digestibility (P = 0.097) and ruminal valerate (P = 0.039) and decreased butyrate proportion (P = 0.068). Animals that received FD had greater values of dry matter (DM; P = 0.009) and OM (P &amp;lt; 0.001) digestibility than HD. The enzyme combination resulted in greater DM (P = 0.026), OM (P = 0.062), crude protein (P = 0.018), EE (P = 0.063) and NDF (P = 0.040) intake and reduced OM digestibility (P = 0.008) compared with AML and FBL alone. The addition of exogenous enzymes in feedlot diets affected nutrient utilization and ruminal fermentation of Nellore cattle fed a feedlot finishing diet.

Cracking the Pericellular Matrix Code: Exploring how MMP-2, -3, and -7 influence matrix breakdown and biomechanical properties

IntroductionThe intricate process of articular cartilage remodeling, pivotal for both physiological functions and osteoarthritis (OA) progression, is orchestrated through a balance of matrix synthesis and breakdown, which is mediated by matrix metalloproteinase enzymes (MMPs). At the heart of this remodeling lies the pericellular matrix (PCM), a specialized microenvironment encapsulating each chondrocyte and composed mainly of collagen type VI and perlecan. The aim of this study was to assess the impact of MMP-2, -3, and -7 on the structural integrity and biomechanical attributes of the PCM. MethodsHuman articular cartilage explants (N=10 patients) were incubated with activated MMP-2, -3, or -7, individually or in combination. Structural alterations in the PCM were evaluated by immunolabeling. The biomechanical properties of the PCM were measured using atomic force microscopy (AFM). ResultsCollagen type VI structural integrity and fluorescence intensity uniformly decreased across all enzyme groups, while perlecan was selectively affected by MMP-3 and -7. AFM measurements demonstrated decreased PCM stiffness after incubation with individual MMPs, leading to an overall ~31 % reduction in elastic modulus for each enzyme. Combinations of enzymes induced comparable significant biomechanical alterations (~35 %), except for MMP-2+MMP-7. DiscussionThis study highlights the significant influence of MMP-induced alterations in PCM composition on biomechanical properties, mirroring characteristics observed in early OA. Each MMP showed specificity in breaking down PCM, and an intriguing interplay, especially between MMP-2 and -7, indicated reduced efficacy in lowering PCM stiffness. Overall, MMP-2, -3, and -7 directly induce functional and structural PCM modifications.

Effects of ultrasound pretreatment on the structure, IgE binding capacity, functional properties and bioactivity of whey protein hydrolysates via multispectroscopy and peptidomics revealed

Whey protein is an important food ingredient, but it is also considered a major food allergen. The aim of this study was to investigate the effect of ultrasound pretreatment on the structure, IgE binding capacity, functional properties and biological activity of whey protein isolate (WPI) hydrolysates (WPH), including WPI hydrolyzed by a combination of enzymes from Bromelain and ProteAXH (BA-WPI) and WPI hydrolyzed by a combination of enzymes from Papain W-40 and ProteAXH (PA-WPI). The IgE binding capacity of BA-WPI and PA-WPI was reduced to 40.28% and 30.17%, respectively, due to disruption/exposure/shielding of conformational and linear epitopes. The IgE binding capacity of sonicated WPI was increased, but ultrasound pretreatment further reduced the IgE binding capacity of the hydrolysates to 32.89% and 28.04%. This is due to the fact that ultrasound pretreatment leads to conformational changes including increased α-helix and β-sheet structure, exposure of aromatic amino acids, surface hydrophobicity, and increased sulfhydryl content, which increases the accessibility of allergenic epitopes to WPI by the enzyme. Multispectral and LC-MS/MS results further indicated that ultrasound pretreatment altered the conformational and primary structural changes of the hydrolysates. The thermograms showed that ultrasound pretreatment mainly altered the epitope spectra of β-lactoglobulin hydrolysates, while it had less effect on the epitope spectra of α-lactalbumin hydrolysates. Additionally, ultrasound pretreatment significantly improved the foaming properties, antioxidant activity, and α-glucosidase inhibition of the hydrolysates without impairing the solubility and emulsification properties of the hydrolysates. Therefore, ultrasound pretreatment is a feasible method to reduce the allergenicity of WPH and to improve their functional properties and bioactivity. Notably, ultrasonic pretreatment improved the effectiveness and efficiency of WPI hydrolysis, which is a feasible method to produce high-quality protein feedstock in a green, efficient, and economical way.

An eco-friendly enzymatic treatment to prepare spinnable banana fibers as an alternative to cotton for textile applications

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.