Hydrolysate Concentration Research Articles

The Effect of a Biostimulant Based on a Protein Hydrolysate of Rainbow Trout (Oncorhynchus mykiss) on the Growth and Yield of Wheat (Triticum aestivum L.).

The research object was the liquid protein hydrolysate “AGROMOREE” from the rainbow trout, which was provided by the company “Russian Aquaculture LLC”. The purpose of this study was the evaluation of the effect of the hydrolysate “AGROMOREE” as a biostimulant on the growth and yield of wheat (Triticum aestivum L.). Biometric indicators of wheat (Triticum aestivum L.) growth were determined in the laboratory and in field tests. In the laboratory, the liquid concentrated hydrolysate was dried to facilitate its use. “AGROMOREE” promoted an increase in germination of 2–4% in all samples compared to the control samples, as well as an increase in the length and number of wheat roots. The biostimulant “AGROMOREE” was introduced in the soil in liquid form at about 3000 L/ha and 4000 L/ha in the field tests. This study showed that “ARGOMOREE” contributed to an increase in the length and quantity of wheat ears, the quantity of grains in the ear and the seed weight. At the same time, the quantity of productive stems increased, so that the biostimulant “AGROMOREE” increased the productivity by 3.9–6.3% with respect to the control sample. In general, using the biostimulant “AGROMOREE” on spring wheat seeds from 2019 in the growing season of 2021 provided an increase in yield from 0.21 t/ha to 0.28 t/ha. The maximum value of raw gluten content in the seed was 5.2%, higher than the content in the control. The content of the mass fraction of protein in the seed was in the range of 12.33–12.56%, i.e., 2% higher than that of the control sample. Thus, according to qualitative and quantitative indicators and the total productivity indicators, the biostimulant «AGROMOREE» can be used to increase wheat productivity and reduce the use of nitrogen fertilizers.

Properties of Protein Hydrolysates and Bioinformatics Prediction of Peptides Derived from Thermal and Enzymatic Process of Skipjack Tuna (Katsuwonus pelamis) Roe

Currently, the use of skipjack tuna (Katsuwonus pelamis) roe to produce hydrolysate is limited, although it is a potentially valuable resource. This study aimed to investigate the physical and chemical characteristics of protein hydrolysates from tuna roe using autoclave and enzymes (alcalase and trypsin at 0.5 and 1.0% w/v). Bioinformatics was also applied to analyze the identified peptides. The hydrolysates were determined for amino acid composition, peptide profile patterns, antioxidant activity, solubility and foaming properties. The proteins were separated by SDS-PAGE before tryptic digestion and peptide identification by nano LC-ESI-MS/MS. The putative bioactivities of the identified peptides were predicted using bioinformatics prediction tools. The main amino acids found in all hydrolysates were cysteine, glycine and arginine (16.26–20.65, 10.67–13.61 and 10.87–12.08 g/100 g protein, respectively). The hydrolysates obtained from autoclaving showed lower molecular weights than those by the enzymatic method. The 0.1 g/mL concentration of hydrolysates provided higher antioxidant activities compared to the others. All hydrolysates had high solubility and exhibited foaming capacity and foam stability. Putative anti-hypertensive, anti-virus and anti-parasite activities were highly abundant within the obtained peptides. Moreover, predicted muti-bioactivity was indicated for seven novel peptides. In the future work, these peptides should be experimentally validated for further applications.

Protective effects of whey protein hydrolysate on Bifidobacterium animalis ssp. lactis Probio-M8 during freeze-drying and storage.

We evaluated the potential of whey protein hydrolysate as a lyoprotectant for maintaining the cell viability of Bifidobacterium animalis ssp. lactis Probio-M8 during freeze-drying and subsequent storage. The moisture content and water activity of the lyophilized samples treated by different concentrations of whey protein hydrolysate were ≤5.23 ± 0.33 g/100 g and ≤0.102 ± 0.003, respectively. During storage at 25°C and 30°C, whey protein hydrolysate had a stronger protective effect on B. lactis Probio-M8 than the same concentration of whey protein. Using the Excel tool GinaFit, we estimated the microbial inactivation kinetics during storage. Whey protein hydrolysate reduced cell damage caused by an increase in temperature. Whey protein hydrolysate could protect cells by increasing the osmotic pressure as a compatible solute. Whey protein hydrolysate improved cell membrane integrity and reduced the amounts of reactive oxygen species and malondialdehyde produced. The findings indicated that whey protein hydrolysate was a novel antioxidant lyoprotectant that could protect probiotics during freeze-drying and storage.

Growth performance, haematological parameters and proximate composition of rainbow trout Oncorhynchus mykiss fed varying dietary levels of protein hydrolysate from heads of Penaeus monodon shrimp processing industry

Fish farming is a huge industry in Vietnam, but there is a shortage of protein supplements for fish diets. A potential great source of protein supplement is the co-products of shrimp processing. The present study aimed to investigate the growth performance and haematological parameters of rainbow trout Oncorhynchus mykiss juveniles fed diets containing four different concentrations of shrimp head protein hydrolysates (0% as the control, 1, 3 or 5% SHPH, four replicates per treatment). Rainbow trout juveniles were randomly distributed into 12 tanks at a density of 30 individuals/tank and were fed twice a day at 8:00 and 17:00 for 8 weeks. Mean weight gain (MWG), specific growth rate (SGR), feed conversion ratio (FCR), protein efficiency ratio (PER) and survival (SR) were calculated. Significant differences (p<0.05) in MWG, SGR, FCR, PER and SR were detected between treatments, with fish fed 5% SHPH performing the best. Mean white blood cells (WBC), red blood cells (RBC), hemoglobin content (Hb) and hematocrit (Hct) significantly increased as dietary protein hydrolysate increased in the diet. Protein and triglyceride concentrations in plasma were not significantly affected by dietary treatment (p > 0.05). Furthermore, muscle and whole body lipid contents were higher in fish fed 3% and 5% protein hydrolysate from shrimp heads than other groups, but muscle and body protein contents in the group fed 5% were higher than other groups. Results from the present study showed that protein hydrolysate from shrimp heads has good potential for use as protein supplement in rainbow trout diets up to 5%. Supplemental dietary protein hydrolysate from shrimp heads also has potential as an immune enhancer because of an increase in WBC. These data provide a potential use for a current waste product and might serve as a model for other countries that have both warm water crustacean industries and cold water fish industries. Further studies to explore the use of protein hydrolysate from shrimp heads in fish diets at more than 5% are recommended.

UV-B Protective and Antioxidant Activities of Protein Hydrolysate From Sea Cucumber (Holothuria scabra) Using Enzymatic Hydrolysis

Sea cucumber is rich in protein that can be used to prepare a potential derived bioactive peptide for antioxidant and protective effect against UV-B induced skin cell damage. This study aimed to optimize preparation of sea cucumber hydrolysate with both UV-B protective and antioxidant activities using three commercial enzymes using response surface methodology (RSM) with a face-centered central composite design (face- centered CCD). Hydrolysis time and concentration of enzyme effects on the degree of hydrolysis (DH), yield, antioxidant and UV-B protective activities of sea cucumber hydrolysates were determined. The optimum conditions for sea cucumber hydrolysis using papain (SCP), alcalase (SCA), or flavourzyme (SCF) were 3.6, 5.0, and 4.1% (w/w protein), respectively, and a hydrolysis time of 360 min. The resulting hydrolysates had a DH of 81-91%, yield of 13-14%, IC50 for DPPH radical scavenging activity of 0.3-4.1 mg/mL, FRAP of 0.5-0.6 mmol FeSO4/mL, and IC50 for ABTS radical scavenging activity of 1.3-1.6 mg/mL. The UV-B protective activity was reported as the HaCaT cell viability percentage after UV-B treatment. The SCP, SCA, and SCF hydrolysates showed 72.4, 74.5, and 71.3% cell viability, respectively. The concentration of hydrolysates with 80% survival of HaCaT cells was 0.21, 0.15 and 0.20 mg/mL for SCP, SCA and SCF, respectively. Thus, the SCP was selected for bioactive peptide isolation and characterization. The SCP contained hydrophilic and hydrophobic amino acids of 42.4 and 57.6%, respectively. The ultrafiltration and Sephadex G-25 gel filtration chromatography were done for peptide isolation from the SCP. Six potential peptides were identified using LC-MS/MS as Leu-Val-Asn-Glu-Leu-Thr-Glu-Phe-Ala-Gln (1163 Da), Leu-Val-Asn-Glu-Val-Thr-Glu-Phe-Ala-Gln (1149 Da), Phe-Val-Asp-Ser-Ser-Ala-Thr-Thr (826 Da), Phe-Asn-Asp-Leu-Gly-Ala-Trp (821 Da), Phe-Pro-Asp-Thr-Thr-Thr-Leu (793 Da), and Lys-Phe-Gly-Glu-Gly-Lys (664).

Exploring competitive inhibition of a family 10 xylanase derived from Hu sheep rumen microbiota by Oryza sativa xylanase inhibitor protein: In vitro and in silico perspectives

The catalytic domain of family GH10 xylanase, XYN-LXY_CD derived from Hu sheep rumen microbiota was expressed in Pichia pastoris X33. The special activity of reXYN-LXY_CD in the culture supernatant was 232.56 U/mg. The optima of reXYN-LXY_CD were 53 °C and pH 7.0. Recombinant Oryza sativa xylanase inhibitor protein (rePOsXIP) competitively inhibited reXYN-LXY_CD with an inhibition constant (Ki) value of 237.37 nM. The concentration of hydrolysates released from beechwood xylan by reXYN-LXY_CD reduced when rePOsXIP was added into the hydrolytic system. Fluorescence of reXYN-LXY_CD was statically quenched by rePOsXIP in a dose-dependent manner. The details in intermolecular interaction between XYN-LXY_CD and OsXIP were investigated by using molecular dynamics (MD) simulations, binding free energy computation and non-covalent interactions (NCI) analysis. Hydrogen bonding and van der Waals played indispensable roles in the XYN-LXY_CD/OsXIP interaction. The α-7 helix of OsXIP tightly occupied the catalytic pocket of XYN-LXY_CD with hydrogen bonding such as K239OsXIP-N261/Q292/E197XYN-LXY_CD (E197, the acid-base catalytic residue), D236OsXIP-K327XYN-LXY_CD and Q242OsXIP-E211/Q212XYN-LXY_CD. Based on the quantum theory of atoms in molecules (QTAIM), the Laplacian of electron density and core-valence bifurcation index of HZ3K239-OE2E197 were 0.1025 a.u. and 0.002218, respectively. Elucidating the mechanism underlying xylanase-inhibitor interactions might help construct XYN-LXY_CD mutants that gain resistance to XIPs and high catalytic activity, which would be more efficient in feed additives in livestock.

Tannery waste as a renewable source of nitrogen for production of multicomponent fertilizers with biostimulating properties.

The studies presented in this work show that solid tannery waste-like shavings can be used as high-protein materials for fertilizer production following the concept of the circular economy. To select appropriate process parameters (mass ratio of shavings meal to the hydrolyzing agent (S:L), hydrolysis medium concentration, temperature) and to ensure the highest possible hydrolysis efficiency, it is useful to apply the well-known response surface methodology (RSM). The analyses revealed that chromium shavings (SCr) were most preferably treated with 10% KOH in a ratio of S:L 1:1 with the process being carried out at 160°C (6.59% N). The optimal hydrolysis conditions for non-chromium (S) shavings were: S:L ratio 1:2, 10% H2SO4, and temperature 160°C (4.08% N). Chromium concentrations in hydrolysates from S and SCr shavings obtained under optimal conditions were 15.2mg/kg and 9483mg/kg, respectively. Hydrolysate samples were analyzed by reversed-phase high-pressure liquid chromatography (RP-HPLC) that revealed that the type of hydrolysis (acidic/alkaline) affects the amino acid profile. Approximately 4.5 times more amino acids were extracted in the KOH environment than during acidic treatment. The hydrolysates contained mainly glycine, alanine, and proline, which are primarily responsible for stimulating plant growth by supporting chlorophyll synthesis, chelating micronutrients, improving pollen fertility, or resistance to low temperatures. The conversion of tannery waste into fertilizer requires the control of contaminant levels, especially chromium, which can oxidize to the carcinogenic form Cr(VI) that is hazardous to humans and the environment.

Candida glycerinogenes Strains Overexpressing Transcription Factors have Improved Furfural Tolerance in Ethanol Production from Non-detoxified Cellulose Hydrolysate.

Cellulose is one of the main raw materials for production of green ethanol, but the presence of the growth inhibitor furfural in non-detoxified lignocellulosic hydrolysates often seriously affects their utilization. In a previous study, we obtained strains of Candida glycerinogenes that were tolerant to furfural, but at concentrations above 2.5g L-1 there was a significant increase in the growth lag phase. In this work, transcription factor genes (SEF1, STB5, CAS5, and ETP1) associated with furfural tolerance were identified and employed to obtain modified strains permitting ethanol fermentation of concentrated and non-detoxified cellulose hydrolysates containing more than 2.5g L-1 furfural. Tolerance to furfural could be increased to 4.5g L-1 by overexpression of either STB5 or ETP1, which have different regulation patterns. Moreover, in non-detoxified and concentrated cellulose hydrolysate, overexpression of ETP1 significantly shortened the growth lag phase and ethanol fermentation time was reduced by 17-20%. In batch fermentations fed with concentrated non-detoxified lignocellulose hydrolysate, ethanol productivity and maximum ethanol concentration reached 2.4g L-1 h-1 and 72.5g L-1, increases of 26.1% and 6.6%, respectively. The results provided a route for the economic use of lignocellulose for chemical production.

Engineering Collariella virescens Peroxygenase for Epoxides Production from Vegetable Oil.

Vegetable oils are valuable renewable resources for the production of bio-based chemicals and intermediates, including reactive epoxides of industrial interest. Enzymes are an environmentally friendly alternative to chemical catalysis in oxygenation reactions, epoxidation included, with the added advantage of their potential selectivity. The unspecific peroxygenase of Collariella virescens is only available as a recombinant enzyme (rCviUPO), which is produced in Escherichia coli for protein engineering and analytical-scale optimization of plant lipid oxygenation. Engineering the active site of rCviUPO (by substituting one, two, or up to six residues of its access channel by alanines) improved the epoxidation of individual 18-C unsaturated fatty acids and hydrolyzed sunflower oil. The double mutation at the heme channel (F88A/T158A) enhanced epoxidation of polyunsaturated linoleic and α–linolenic acids, with the desired diepoxides representing > 80% of the products (after 99% substrate conversion). More interestingly, process optimization increased (by 100-fold) the hydrolyzate concentration, with up to 85% epoxidation yield, after 1 h of reaction time with the above double variant. Under these conditions, oleic acid monoepoxide and linoleic acid diepoxide are the main products from the sunflower oil hydrolyzate.

Microbial xylitol production from corn cob using Candida guilliermondii

Xylitol is a polyol that is widely used in the food industries as an alternative sweetener due to some health benefits. The microbial production of xylitol based on corn cobs was potential to be developed due to the abundant availability of corn cobs in Indonesia. In addition, it is an alternative process that is a higher yield and environmentally friendly. This study was conducted to assign the optimum process in the xylitol production based on corn cobs using Candida guilliermondii. The hydrolysis of corn cobs used sulfuric acid 1% at a temperature of 121°C for 60 minutes. The hydrolysate was then added with calcium hydroxide and activated charcoal to remove inhibitors. The design of experimental used was a factorial completely randomized design with three factors included corn cobs hydrolysate concentration (50% and 66%), glucose (0 g l-1 and 5 g l-1), and incubation period (120 h and 168 h). The result showed that concentration of hydrolysate, glucose, and incubation period had an effect on the xylitol produced. The highest xylitol was 144.09 ppm that was produced by the hydrolysate of 66%, glucose of 5 g l-1 and incubation period of 168 h, meanwhile the residual xylose was 2081.87 ppm.

Effect of proteases on water-soluble and fat-soluble aroma precursors of goat visceras protein hydrolysate

This study investigated the action of proteolytic enzymes in the production of goat viscera hydrolysates, and their flavoring potential. The study focus the production of flavor precursor molecules and the formation of volatile compounds resulting from four hydrolysis treatments: HA - Alcalase®, HF - Flavourzyme®, HN - Neutrase®, and HFN - Flavourzyme® and Neutrase®. The highest degree of hydrolysis (26.74%), number (23 volatiles) and concentration of volatile compounds (274.0 μg/L) were obtained with the action of Alcalase. The main classes of volatile compounds identified in hydrolysates included acids, aldehydes, pyrazines, terpenes and Streacker aldehydes. Amino acids, sugars and fatty acids were identified with higher concentrations in HF and HFN hydrolysates due to the low participation of these precursors in the formation of volatile compounds. Therefore, the use of Alcalase in the proteolysis of goat viscera stood out compared to other enzymes, providing impacting aroma compounds, such as pyrazines, for meat flavor.

Anti-Inflammatory, ACE Inhibitory, Antioxidative Activities and Release of Novel Antihypertensive and Antioxidative Peptides from Whey Protein Hydrolysate with Molecular Interactions

Objective The aim of the study was to evaluate the whey protein hydrolysate with bio-functional attributes viz. antioxidative, anti-inflammatory and ACE inhibition efficacy and release of bioactive peptides with antioxidative and ACE-inhibitory activity by employing Pepsin. Method The antioxidant, Anti-inflammatory, ACE inhibitory and proteolytic activities of the whey protein hydrolysates were studied followed by SDS-PAGE analysis and IEF. Anti-inflammatory activity of whey protein hydrolysate was also studied on RAW 264.7 cell line. The separation of the bioactive peptides from whey protein hydrolysate was achieved by RP-HPLC. The purified bioactive peptides were identified and characterized using RPLC/MS. Results WPC (Whey protein concentrate) hydrolysate with pepsin showed proteolytic activity ranging between 14.46 and 18.87 mg/ml. Using the ABTS assay, the highest antioxidative activity was observed in 10 kDa retentate (84.50%) and 3 kDa retentate (85.96%), followed by the highest proteolytic activity (13.83 mg/ml) and ACE inhibitory activity (58.37%) in a 5% WPC solution at 65 °C after 8 h of pepsin hydrolysis. When the protein hydrolysate concentration was low, the production of proinflammatory cytokines by lipopolysaccharide-treated murine macrophages (RAW 264.7) was reduced. SDS-PAGE results exhibited very little protein bands when comparing with WPC hydrolysates to insoluble WPC. There were no protein spots on 2 D gel electrophoresis and “in-solution trypsin digestion” technique have been utilized to digest protein samples directly from WPC hydrolysates. Novel antioxidative peptides and ACE inhibitory peptides were also observed by comparing two databases, i.e., BIOPEP and AHTPDB respectively. The peptide sequences used in this study were found to have excellent potential to be used as inhibitors of hACE as all of them were able to show substantial interactions against the enzyme’s active site. Conclusions The antihypertensive and antioxidative peptides from whey protein hydrolysates may be beneficial for the future development of physiologically active functional foods. Further, in vivo investigations are required to establish the health claim for each individual bioactive peptide from whey protein hydrolysate. Supplemental data for this article is available online at

Pilot-scale biorefinery for the production of purified biopolymers based on hydrothermal treatment in flow-through reactor cycles

Purified solid fractions of hemicellulose and pectin biopolymers with different molecular weight were produced on a pilot-scale from discarded carrots. The pulp was subjected to hydrothermal extraction on a pilot plant operating in cycles of three flow-through reactors in series at 140 and 180 °C. Two operational modes of start-up (with and without water pre-filling) were tested, showing that it influences the final hydrolysate concentration and the stability of the system. Free sugars, arabinogalactan hemicellulose and homogalacturonan pectin were produced at maximum recoveries of 379.5 g/kg dry pulp, 81.0 g/kg dry pulp and 5.35 g/kg dry pulp, respectively, with the extraction of arabinogalactan reaching a yield of 96.1 % (w/w). The extracted biopolymers were separated and purified through ultrafiltration and diafiltration cycles using a multi-membrane system (30, 10, 1 kDa). Ultrafiltration and especially diafiltration allowed going from extracts with molecular weight, polydispersity and purity values of 14.77 kDa, 19.2 and 22.2 % w/w (140 °C extract) and 8.08 kDa, 18.2 and 14.9 % w/w (180 °C extract), to fractions with values of 80.36 kDa, 67.77 kDa, 9.85 kDa, 5.23 kDa and 3.86 kDa (molecular weight), 1.3 – 3.8 (polydispersity), and 64.5 – 100 % w/w (purity). The five fractions were freeze-dried and spray-dried. The pilot-scale system allowed the production of purified biopolymer fractions of high purity, different average molecular weight, and in sufficient quantity for their subsequent transformation into products such as biodegradable films.

Optimization of Enzymatic Hydrolysis of Perilla Meal Protein for Hydrolysate with High Hydrolysis Degree and Antioxidant Activity.

Botanical oils are staple consumer goods globally, but as a by-product of oil crops, meal is of low utilization value and prone to causing environmental problems. The development of proteins in meal into bioactive peptides, such as Perilla peptide, through biotechnology can not only solve environmental problems, but also create more valuable nutritional additives. In the present work, the hydrolysis process of Perilla meal protein suitable for industrial application was optimized with the response surface methodology (RSM) on the basis of single-factor experiments. Alcalase was firstly selected as the best-performing among four proteases. Then, based on Alcalase, the optimal hydrolysis conditions were as follows: enzyme concentration of 7%, hydrolysis temperature of 61.4 °C, liquid-solid ratio of 22.33:1 (mL/g) and hydrolysis time of 4 h. Under these conditions, the degree of hydrolysis (DH) of Perilla meal protein was 26.23 ± 0.83% and the DPPH scavenging capacity of hydrolysate was 94.15 ± 1.12%. The soluble peptide or protein concentration of Perilla meal protein hydrolysate rose up to 5.24 ± 0.05 mg/mL, the ideal yield of which was estimated to be 17.9%. SDS-PAGE indicated that a large proportion of new bands in hydrolysate with small molecular weights appeared, which was different from the original Perilla meal protein. The present data contributed to further, more specific research on the separation, purification and identification of antioxidant peptide from the hydrolysate of Perilla meal protein. The results showed that the hydrolysis of Perilla meal protein could yield peptides with high antioxidant activity and potential applications as natural antioxidants in the food industry.

Enzymatic hydrolysis and fermentation of soy flour to produce ethanol and soy protein concentrate with increased polyphenols

AbstractThere is a need to effectively concentrate soy protein from defatted soy flour (DSF) while simultaneously valorizing the carbohydrate‐rich byproduct, which would otherwise be a waste. This study aims to evaluate a process developed to produce soy protein concentrates (SPC) by substantially hydrolyzing carbohydrates from DSF with the help of enzymes into water‐soluble saccharides and monomeric sugars, which were simultaneously utilized by Saccharomyces cerevisiae for fermentation into ethanol. The enzyme mixture consisted of cellulase, pectinase, and ‐galactosidase blend. The effect of process time on SPC, overall protein recovery, carbohydrate hydrolysis, yeast growth, ethanol concentration, and total polyphenol concentration (TPC) of SPC and hydrolysate was evaluated. Control and enzymes‐only (EO) systems were maintained in conjunction with the enzymes + yeast (EY) system to individually assess the impact of isoelectric precipitation of soy proteins and enzymatic hydrolysis of carbohydrates without yeasts. After 12.25 h of EY process, 100 g of dry DSF produced 68.45 g dry SPC containing % protein and 384 ml hydrolysate containing mg/ml ethanol. Flatulence‐causing raffinose‐series‐oligosaccharides (RSOs) were completely hydrolyzed. Soluble carbohydrates in the EY treatment were consistently lower than in the control and EO treatment. TPC of SPC prepared by EY treatment increased by 2.5 times compared to the control. Thus, this novel process successfully produced a high‐protein SPC with hydrolyzed RSOs, lower insoluble carbohydrates, high TPC, and a coproduct ethanol.

The Effect of Chitosan Hydrolysate on Solanum Lycopersicum Plant Growth

Chitosan is a well-known subjectof researchbecause of its beneficial properties, including its antibacterial and fungicidal activity, as well as its effect on plant physiology. One of the maindifficulties in agricultural chitosan implementation is the poor solubility of high molecular chitosan in water. Reducing themolecular weight of chitosan by acid hydrolysis increases its solubility. This research studied the effect of chitosan hydrolysate on the germination and development ofSolanum lycopersicum L. plants. Theeffects of chitosan hydrolysate on seed germination, shoot development in the first week of development and in the first month of development were evaluated. According to the results, high concentrations of chitosan hydrolysate completely inhibited seed germination. However, short-term treatment by high concentrations of chitosan hydrolysate stimulatedthe development of seedlings, leading to substantially longerroots. Regular root application of high concentrations of chitosan hydrolysate inhibited plant development.&#x0D; Keywords: chitosan hydrolysate, chitosan, plant growth regulators, plant germination

Relationship between feed concentration and bioactive cationic peptide recovery: Impact on ecoefficiency of EDUF at semi-industrial scale

Electrodialysis with ultrafiltration membrane (EDUF) is currently investigated at semi-industrial scale (560 cm2 of effective electrode surface) for the separation of bioactive cationic peptides. In this study, the focus was put on the impact of the initial whey protein hydrolysate (WPH) concentration, and the effect of current intensity have been tested. Experiments at 2%, 4% and 8% (w/v) WPH showed that a proportional relationship exists between the initial WPH concentration, and the quantity of peptides recovered. The cationic recovery fractions contained 16 peptides in similar amount as the previous work although including newly identified sequences. Thus, selectivity of the cationic recovery fraction was even improved. The working parameters used allowed obtaining a final 4.7% peptide yield after 6-h runs at 8% WPH. Aiming at nutraceutical applications built on the important ACE-inhibitory and DPP-IV inhibitory activities observed (6 times and 4 times higher than those of the initial WPH, respectively), two types of eco-efficiency scores were established based on those bioactivity values. Considering these scores, using the cationic recovery fractions as an antidiabetic product seems to be the most promising avenue. Preliminary results yielded by an experiment at higher current intensity are also very encouraging due to a barely increased environmental impact cost conjugated to a significantly higher peptide yield and preserved bioactivities. With all these new results, EDUF can be viewed as a promising industrial technology for the recovery of bioactive peptides in the perspective of ecoefficient strategies.

Experimental and in silico studies of competitive inhibition of family GH10 Aspergillus fumigatus xylanase A by Oryza sativa xylanase inhibitor protein

The family GH10 Aspergillus fumigatus xylanase A (AfXylA10) gene, afxyla10 was cloned and recombinantly expressed in Pichia pastoris X33. The optimum temperature and pH of reAfXylA10 was 53 °C and 7.0, and Mn2+ remarkably activated the catalytic activity. The recombinant Oryza sativa xylanase inhibitor protein, rePOsXIP significantly inhibited reAfXylA10 with inhibition constant (Ki) of 177.94 nM via competitive inhibition and decreased the concentration of hydrolysate from beechwood xylan. Optimal inhibition of rePOsXIP on reAfXylA10 occurred at 45 °C for 40 min. The fluorescence of reAfXylA10 was statically quenched by rePOsXIP, indicating the formation of reAfXylA10-rePOsXIP complex during their interaction. Furthermore, molecular dynamics (MD) simulations were performed to obtain the detailed information on enzyme-inhibitor interaction. The binding free energy (ΔG) of AfXylA10-OsXIP complex was −30 ± 9 kcal/mol by MM-PBSA calculation, and the α-7 helix of OsXIP anchored in the catalytic cleft of AfXylA10 by competition with the xylan substrate. K239OsXIP stably interacted with the catalytic site E140AfXylA10 through hydrogen bond and vdW interaction. Intermolecular hydrogen bonds T104AfXylA10/V99AfXylA10-Q5OsXIP, R256AfXylA10-E235OsXIP, D155AfXylA10-Y243OsXIP and D145AfXylA10-R194OsXIP on the upper of the TIM barrel were essential for strengthening the stability of complex. Therefore, these non-covalent interactions (NCI) played key role in the interaction between AfXylA10 and OsXIP.

Characterization and adaptation of Caldicellulosiruptor strains to higher sugar concentrations, targeting enhanced hydrogen production from lignocellulosic hydrolysates

BackgroundThe members of the genus Caldicellulosiruptor have the potential for future integration into a biorefinery system due to their capacity to generate hydrogen close to the theoretical limit of 4 mol H2/mol hexose, use a wide range of sugars and can grow on numerous lignocellulose hydrolysates. However, members of this genus are unable to survive in high sugar concentrations, limiting their ability to grow on more concentrated hydrolysates, thus impeding their industrial applicability. In this study five members of this genus, C.owensensis, C. kronotskyensis, C.bescii, C.acetigenus and C.kristjanssonii, were developed to tolerate higher sugar concentrations through an adaptive laboratory evolution (ALE) process. The developed mixed population C.owensensis CO80 was further studied and accompanied by the development of a kinetic model based on Monod kinetics to quantitatively compare it with the parental strain.ResultsMixed populations of Caldicellulosiruptor tolerant to higher glucose concentrations were obtained with C.owensensis adapted to grow up to 80 g/L glucose; other strains in particular C. kristjanssonii demonstrated a greater restriction to adaptation. The C.owensensis CO80 mixed population was further studied and demonstrated the ability to grow in glucose concentrations up to 80 g/L glucose, but with reduced volumetric hydrogen productivities (Q_{{{text{H}}_{2} }}) and incomplete sugar conversion at elevated glucose concentrations. In addition, the carbon yield decreased with elevated concentrations of glucose. The ability of the mixed population C.owensensis CO80 to grow in high glucose concentrations was further described with a kinetic growth model, which revealed that the critical sugar concentration of the cells increased fourfold when cultivated at higher concentrations. When co-cultured with the adapted C.saccharolyticus G5 mixed culture at a hydraulic retention time (HRT) of 20 h, C.owensensis constituted only 0.09–1.58% of the population in suspension.ConclusionsThe adaptation of members of the Caldicellulosiruptor genus to higher sugar concentrations established that the ability to develop improved strains via ALE is species dependent, with C.owensensis adapted to grow on 80 g/L, whereas C.kristjanssonii could only be adapted to 30 g/L glucose. Although C.owensensis CO80 was adapted to a higher sugar concentration, this mixed population demonstrated reduced Q_{{{text{H}}_{2} }} with elevated glucose concentrations. This would indicate that while ALE permits adaptation to elevated sugar concentrations, this approach does not result in improved fermentation performances at these higher sugar concentrations. Moreover, the observation that planktonic mixed culture of CO80 was outcompeted by an adapted C.saccharolyticus, when co-cultivated in continuous mode, indicates that the robustness of CO80 mixed culture should be improved for industrial application.

Rice Straw Hydrolysis and Cultivation of Xanthophyllomyces dendrorhous for Astaxanthin Production and Antioxidant Properties

Astaxanthin is a natural pigment with strong antioxidant activity and is widely supplied as dietary supplement. The red yeast Xanthophyllomyces dendrorhous is one of the potential sources for astaxanthin production. Rice straw was considered for utilization on cultivation and astaxanthin production. Rice straw was treated by autoclave-assisted alkaline pretreatment. and hydrolyzed by cellulase hydrolysis. X. dendrorhous TISTR5730 was cultured and accumulated as the astaxanthin on rice straw hydrolysate. The highest reducing sugar concentration of rice straw hydrolysate was 89.82±0.39g/L (0.71±0.01g/g) when using rice straw (3.3%) and cellulase loading (3 mL). High cell density X. dendrohous TISTR5730 cultivation on rice straw hydrolysate with 20 and 40 g/L of initial reducing sugar was investigated. 40 g/L was appropriate for biomass production while 20 g/L was suitable for astaxanthin accumulation. The highest astaxanthin content was 417.28±50.89 μg/g cell basis at 192 h. Astaxanthin productivity and yield coefficient were 0.01±0.00 mg/L/h and 0.11±0.01 mg/g sugar consumed. The antioxidant activities of astaxanthin were determined by DPPH and ABTS scavenging and FRAP reducing power. The produced astaxanthin represented the high antioxidant activities with IC50 of 9.30 and 1.67 μg/mL of DPPH and ABTS scavenging respectively and FRAP reducing power of 5.31±0.07 μg TEAC/mL. This research indicated that rice straw hydrolysate could be an alternative medium for astaxanthin production. Antioxidant activity of astaxanthin was proved and was feasible for further applications.