Enzymatic Hydrolysis Research Articles

Fractionation and characterisation of sialylated-mucin glycoprotein from edible birds' nest hydrolysates through anion exchange chromatography

Edible bird's nest (EBN) is made up of sialylated-mucin glycoprotein with various health benefits due to its high antioxidative activity. However, as a macromolecule with distinct charged sialic acid and amino acids, fractions with different charges would have varied physicochemical properties and antioxidant activity, which have not been studied. Therefore, this study aimed to fractionate and purify the enzymatic hydrolysed of cleaned EBN (EBNhc) and EBN by-product (EBNhbyp) through anion exchange chromatography (AEC), and determine their molecular weights, physicochemical properties, and antioxidative activities. Overall, 26 fractionates were collected from enzymatic hydrolysate by AEC, which were classified into 5 fractions. It was found that the positively charged fraction of EBNhc (CF1) and EBNhbyp (DF1) showed the significantly highest (p < 0.05) soluble protein contents (22.86 and 18.40 mg/g), total peptide contents (511.13 and 800.47 mg/g) and ferric reducing antioxidant power (17.44 and 6.96 mg/g) among the fractionates. In conclusion, a positively charged fraction (CF1 and DF 1) showed more desired physicochemical properties and antioxidative activities. This research suggests the potential of AEC fractionation as a technology to purify EBN and produce positively charged EBN fractionates with antioxidative potential that could be applied as food components to provide health benefits.

A study on the catalytic domain of pork phospholipase A2: Enzymatic properties and hydrolysis characteristics of phosphatidylcholine and its hydroperoxide

Endogenous phospholipase A2 (PLA2) plays an important role in phospholipids degradation during cured meat products manufacturing. The present study was undertaken to reveal more information about the endogenous PLA2 in muscles and its role in degradation of intramuscular phospholipids. With the catalytic domain of pork calcium-independent PLA2 (iPLA2cd), impacts of physic-chemical factors on the activity were investigated and substrate specificity of the enzyme were tested respectively. The optimum temperature and pH of pork iPLA2cd were 40 °C and 7.5, respectively. The iPLA2cd could be stimulated by adequate contents of NaCl and ATP, and inhibited by CaCl2 and NaNO2. For native phospholipids, the iPLA2cd was of a little higher affinity towards phosphatidylcholine (PC) than phosphatidylethanolamine (PE), phosphoserine (PS) and phosphatidylinositol (PI). The iPLA2cd could preferentially hydrolyze peroxidized PC over the native PC. The results would help better understand the degradation of phospholipids and the role played by endogenous enzymes during meat products manufacturing.

Diversity and Bioprospection of Gram-positive Bacteria Derived from a Mayan Sinkhole

Water-filled sinkholes known locally as cenotes, found on the Yucatán Peninsula, have remarkable biodiversity. The primary objective of this study was to explore the biotechnological potential of Gram-positive cultivable bacteria obtained from sediment samples collected at the coastal cenote Pol-Ac in Yucatán, Mexico. Specifically, the investigation aimed to assess production of hydrolytic enzymes and antimicrobial compounds. 16 S rRNA gene sequencing led to the identification of 49 Gram-positive bacterial isolates belonging to the phyla Bacillota (n = 29) and Actinomycetota (n = 20) divided into the common genera Bacillus and Streptomyces, as well as the genera Virgibacillus, Halobacillus, Metabacillus, Solibacillus, Neobacillus, Rossellomorea, Nocardiopsis and Corynebacterium. With growth at 55ºC, 21 of the 49 strains were classified as moderately thermotolerant. All strains were classified as halotolerant and 24 were dependent on marine water for growth. Screening for six extracellular hydrolytic enzymes revealed gelatinase, amylase, lipase, cellulase, protease and chitinase activities in 93.9%, 67.3%, 63.3%, 59.2%, 59.2% and 38.8%, of isolated strains, respectively. The genes for polyketide synthases type I, were detected in 24 of the strains. Of 18 strains that achieved > 25% inhibition of growth in the bacterial pathogen Staphylococcus aureus ATCC 6538, 4 also inhibited growth in Escherichia coli ATCC 35,218. Isolates Streptomyces sp. NCA_378 and Bacillus sp. NCA_374 demonstrated 50–75% growth inhibition against at least one of the two pathogens tested, along with significant enzymatic activity across all six extracellular enzymes. This is the first comprehensive report on the biotechnological potential of Gram-positive bacteria isolated from sediments in the cenotes of the Yucatán Peninsula.

Enzymatic Activity of Fungi for Hydrolysis of Wheat Bran and Cultivation of Oleaginous Yeasts

Sustainable alternatives to petrochemical raw materials in oleochemical, laundry detergent and personal care industry need to be implemented. Presently, palm oil is the predominant cost-efficient option. While it is a renewable source, it has negative environmental and socioeconomic implications. Therefore, the request for ecologically certified and palm oil-free cleaning agents and household care products is rapidly growing, supported by concepts of waste-free biorefinery. Oleaginous yeasts are currently the most promising source for microbial lipid production. Additionally, wheat bran is a cereal milling by-product, which is available in high quantities and provides an excellent feedstock. Enzymatically generated wheat bran hydrolysate provides an excellent microbial cultivation medium for the sustainable production of high value biological additives. Indeed, wheat bran consists predominantly of lignocellulose, and therefore of cellulose, hemicellulose and lignin, which can be hydrolyzed by established enzyme systems. Currently, such enzymatic systems are mainly obtained for the market from genetically modified organisms (GMO). This represents a problem because GMO products or process components cannot be included in eco-certified household care formulations. Therefore, innovative GMO-free biotechnological processes need to be implemented, as well as the isolation of novel GMO-free hydrolase enzyme systems. Fungal strains are an interesting source for the efficient screening and isolation GMO-free hydrolases (see figure).This contribution is part of a project, which aims to use a holistic biotechnological approach to convert mill residues into customized specialty ingredients of eco-certified detergents. The project focuses on three main tasks:investigation of a microbial group of eight Ascomycete fungi,production of enzymes for hydrolysis of wheat bran, anduse of the resulting biomass hydrolysate to cultivate oleaginous yeasts.Hydrolase producing fungal strains were cultivated on different substrates to investigate their ability to generate specific enzyme systems capable of hydrolyzing wheat bran. The activity of the enzymes was qualitatively and quantitatively characterized to determine the optimal strains for the degradation of wheat bran and suited to implement a large-scale process. Supernatants of different fungal strains cultivations were tested in the hydrolysis step to compare the hydrolysis performances of differently active enzyme mixes. The biomass hydrolysate was used to cultivate lipid generating yeasts to investigate their ability to grow on the microbial cultivation medium.

Flavor Improvement of Maillard Reaction Intermediates Derived from Enzymatic Hydrolysates of Oudemansiella raphanipes Mushroom

Maillard reaction intermediate (MRI) was prepared by the enzymatic hydrolysate (EH) of Oudemansiella raphanipes and fructose. The optimal preparation condition of MRIs was obtained when the Maillard reaction parameters were as follows: fructose addition of 5%, reaction time of 60 min, and temperature of 60 °C. E-Tongue results indicated that the umami and saltiness of MRIs were greater than those of Maillard reaction products (MRPs) and EH, and the taste-enhancing ability of MRIs was even more prominent than that of MRPs. E-Nose could obviously distinguish EH, MRIs, and MRPs, and there was an obvious difference between MRPs and MRIs regarding volatile aroma compounds. A total of 35 volatile flavor substances were identified among the three samples, including 6 alcohols, 13 aldehydes, 9 ketones, 2 esters, and 5 other compounds. Overall, MRIs could avoid the production of complete reaction products with an inferior flavor, and further enhance the umami taste.

Blocking miR396 activity by overexpression MIM396 improved switchgrass tiller number and biomass yield.

MicroRNA396 (miR396) plays an important role in the regulation of plant growth and development by repressing the expression level of its target growth-regulating factor (GRF) family genes. In our previous study, we found that overexpression of miR396 negatively regulated both tillering and biomass yield in switchgrass (Panicum virgatum L.). We, therefore, speculated that blocking the expression of miR396 could enhance switchgrass tillering and biomass yield. Here, we produced transgenic switchgrass plants overexpressing a target mimicry form of miR396 (MIM396) in wild type (WT) and Os-MIR319b overexpressing switchgrass plant (with higher enzymatic hydrolysis efficiency, but reduced tillering), in which the expression of miR396 was blocked. The phenotype and biological yields of these plants were analyzed. Blocking miR396 to improve its target PvGRFs expression in switchgrass improved the tiller number and dry weight of transgenic plants. Further morphological analysis revealed that MIM396 plants increased the number of aerial branches and basal tillers compared to those of wild-type plants. The enzymatic efficiency of MIM396 plants was reduced; however, the total sugar production per plant was still significantly higher than that of wild-type plants due to the increase in biomass. In addition, blocking miR396 in a transgenic switchgrass plant overexpressing Os-MIR319b (TG21-Ms) significantly increased the PvGRF1/3/5 expression level and tiller number and biomass yield. The miR156-target gene PvSPL4, playing a negative role in aerial and basal buds outgrowth, showed significant downregulated in MIM396 and TG21-Ms. Those results indicate that miR396-PvGRFs, through disrupting the PvSPL4 expression, are involved in miR319-PvPCFs in regulating tiller number, at least partly. MIM396 could be used as a molecular tool to improving tiller number and biomass yield in switchgrass wild type and miR319b transgenic plants. This finding may be applied to other graminaceous plants to regulate plant biological yield.

Evaluation of OxiOrganosolv pretreated hardwood and softwood lignocelluloses as substrates for the chemoenzymatic production of 5-hydroxymethylfurfural (HMF)

Furans, such as 5-hydroxymethylfurfural (HMF), are compounds of great importance that can serve as starting materials for the synthesis of polymers. Their production from lignocellulose-derived sugar streams offers a promising alternative to fossil fuels, while enabling biomass transformation to chemicals with higher value. In the present work, the production of HMF from OxiOrganosolv pretreated beechwood and pine was assessed by integrating a three-step process of enzymatic saccharification and isomerization followed by catalytic dehydration. The use of isobutanol in the pretreatment solvent and the addition of polyoxometallates (POMs) as oxidative catalysts were evaluated. The results showed that isobutanol leads to high delignification rates for both beechwood and pine, yielding cellulose-rich pulps with high susceptibility to enzymatic hydrolysis and isomerization. A fructose production up to 51.2 and 53.4 g/g of pretreated material was achieved for beechwood and pine, respectively, corresponding to 14 and 11.3 g of HMF/g of pretreated material. Regarding the use of POMs, the commercially available phosphomolybdic acid (HPMO) and POMs modified with oxidation metals (Fe-PMO, Cu-PMO) were tested, verifying their beneficial effect to lignin depolymerization and the composition of the final pulp. Hydrolysates produced from HPMo and Cu-PMo-assisted OxiOrganosolv pretreatment were efficiently used for the production of HMF, while severe inhibition of the dehydration reaction was observed with the hydrolysates from Fe-PMo pretreated biomass due to the presence of residual metals. This is the first systematic report comparing two lignocellulosic materials subjected to different pretreatment conditions for their potential to produce fructose and, subsequently, HMF.

Gelatinization and fermentation synergy: investigating the protein digestibility, mineral bioaccessibility and microstructural transformations of black mash beans through Saccharomyces cerevisiae and Lactobacillus spp

ABSTRACT This study explored the nutritional qualities of black mash bean flour impacted by fermentation, gelatinization and their combination. Protease activity, in vitro protein digestibility (IVPD), and protein solubility were evaluated for both fermented and unfermented black mash bean flour samples. Interestingly, when compared to fermented samples and gelatinized mash bean flour (GMBF), raw mash bean flour (RMBF) showed the highest protein solubility. Due to enzymatic activity and protein hydrolysis, fermentation with Lactobacillus E14 and Saccharomyces cerevisiae MK-157 enhanced protein solubility. Prolonged fermentation timeframes were positively correlated with protease activity and IVPD, suggesting the degradation of complex proteins. The highest levels of IVPD (90.12%) and protease activity (5546.78 U/g) were detected in GMBF fermented by S. cerevisiae MK-157, indicating the impact of fermentation on protein breakdown. The Lactobacillus E14-fermented RMBF exhibited 5521.45 U/g of protease activity and 87.54% IVPD. The mineral content of the fermented samples was substantially greater than that of the unfermented GMBF. The increased calcium, magnesium, potassium, sodium, iron, zinc, and copper levels for the GMBF fermented by S. cerevisiae MK-157 indicated that fermentation facilitated the release of minerals from chelated compounds. Microscopic examination revealed higher protein quantities in the fermented flour as well as hydrolyzed and broken starch granules embedded in the protein matrix. Large mash bean proteins were shown to be proteolyzed during fermentation into smaller peptides by SDS PAGE. Storage proteins were extensively broken down during fermentation with S. cerevisiae MK-157, whereas Lactobacillus E14 displayed a more focused pattern of hydrolysis. The nutritional profile of black mash bean flour was improved by fermentation with Lactobacillus E14 and S. cerevisiae MK-157.

The Effect of Whey Protein Isolate Hydrolysate on Digestive Properties of Phytosterol.

Phytosterol (PS) is a steroid, and its bioavailability can be enhanced by interacting with protein in the C-24 hydroxyl group. The interaction between sterols and amino acid residues in proteins can be enhanced by enzymatic hydrolysis. Phytosterol and whey insulation hydrolysates (WPH1-4) fabricated by the Alcalase enzyme at different enzymatic hydrolysis times were selected as delivery systems to simulate sterol C-24 hydroxyl group interaction with protein. Increasing hydrolysis time can promote the production of β-Lg, which raises the ratio of β-turn in the secondary structure and promotes the formation of interaction between WPH and PS. The correlation coefficient between hydrogen bonds and encapsulation efficiency (EE) and bioaccessibility is 0.91 and 0.88 (P < 0.05), respectively, indicating that hydrogen bonds of two components significantly influenced the combination by concealing the hydrophobic amino acids and some residues, which improved PS EE and bioavailability by 3.03 and 2.84 times after PS was combined with the WPI hydrolysate. These findings are expected to enhance the absorption of PS and other macromolecules by protein enzymatic hydrolysis to broaden their applications for food.

Campylobacter jejuni and casein hydrolysate addition: Impact on poultry in vitro cecal microbiota and metabolome.

This study investigates the impact of casein hydrolysates on the poultry ceca inoculated with Campylobacter focusing on microbial molecular preferences for different protein sources in the presence of Campylobacter jejuni. Three casein sources (intact casein (IN), casein enzyme hydrolysate (EH), and casein acid hydrolysate (AH)) were introduced to cecal contents in combination with inoculated C. jejuni in an in vitro model system incubated for 48 h at 42°C under microaerophilic conditions. Samples were collected at 0, 24, and 48 h. Genomic DNA was extracted and amplified using custom dual-indexed primers, followed by sequencing on an Illumina MiSeq platform. The obtained sequencing data were then analyzed via QIIME2-2021.11. Metabolite extracts were analyzed with ultra-high-performance liquid orbitrap chromatography-mass spectrometry (UHPLC-MS). Statistical analysis of metabolites was conducted using MetaboAnalyst 5.0, while functional analysis was performed using Mummichog 2.0 with a significance threshold set at P < 0.00001. DNA sequencing and metabolomic analyses revealed that C. jejuni was most abundant in the EH group. Microbial diversity and richness improved in casein supplemented groups, with core microbial differences observed, compared to non-supplemented groups. Vitamin B-associated metabolites significantly increased in the supplemented groups, displaying distinct patterns in vitamin B6 and B9 metabolism between EH and AH groups (P < 0.05). Faecalibacterium and Phascolarctobacterium were associated with AH and EH groups, respectively. These findings suggest microbial interactions in the presence of C. jejuni and casein supplementation are influenced by microbial community preferences for casein hydrolysates impacting B vitamin production and shaping competitive dynamics within the cecal microbial community. These findings underscore the potential of nutritional interventions to modulate the poultry GIT microbiota for improved health outcomes.

Mitigating candidiasis with acarbose by targeting Candida albicans α-glucosidase: in-silico, in-vitro and transcriptomic approaches

Biofilm-associated candidiasis poses a significant challenge in clinical settings due to the limited effectiveness of existing antifungal treatments. The challenges include increased pathogen virulence, multi-drug resistance, and inadequate penetration of antimicrobials into biofilm structures. One potential solution to this problem involves the development of novel drugs that can modulate fungal virulence and biofilm formation, which is essential for pathogenesis. Resistance in Candida albicans is initiated by morphological changes from yeast to hyphal form. This transition triggers a series of events such as cell wall elongation, increased adhesion, invasion of host tissues, pathogenicity, biofilm formation, and the initiation of an immune response. The cell wall is a critical interface for interactions with host cells, primarily through various cell wall proteins, particularly mannoproteins. Thus, cell wall proteins and enzymes are considered potential antifungal targets. In this regard, we explored α-glucosidase as our potential target which plays a crucial role in processing mannoproteins. Previous studies have shown that inhibition of α-glucosidase leads to defects in cell wall integrity, reduced adhesion, diminished secretion of hydrolytic enzymes, alterations in immune recognition, and reduced pathogenicity. Since α-glucosidase, primarily converts carbohydrates, our study focuses on FDA-approved carbohydrate mimic drugs (Glycomimetics) with well-documented applications in various biological contexts. Through virtual screening of 114 FDA-approved carbohydrate-based drugs, a pseudo-sugar Acarbose, emerged as a top hit. Acarbose is known for its pharmacological potential in managing type 2 diabetes mellitus by targeting α-glucosidase. Our preliminary investigations indicate that Acarbose effectively inhibits C. albicans biofilm formation, reduces virulence, impairs morphological switching, and hinders the adhesion and invasion of host cells, all at very low concentrations in the nanomolar range. Furthermore, transcriptomic analysis reveals the mechanism of action of Acarbose, highlighting its role in targeting α-glucosidase.

Modification of Bioactive Properties in Food Protein Hydrolysates by Alcalase and Trypsin

Abstract Background Protein hydrolysates are the fragments of proteins that form during the hydrolysis with promised bioactive properties. Enzymatic hydrolysis produces bioactive peptides with specific amino acid profile. Among all the properties, angiotensin-I converting enzyme (ACE-I) inhibitory activity of the protein hydrolysates attained maximum attention to combat life-threatening disease like hypertension. Methodology In the present study 10 protein-rich food sources (&gt; 20% protein content on dry basis) were used to prepare protein hydrolysates using alcalase and trypsin to investigate the effect of enzymes on ACE-I inhibitory and antioxidant activity. Results Among all the sources, the highest degree of hydrolysis was observed in sardine (62.89%) followed by soybean (61.24%) when hydrolyzed by alcalase. Kidney pea exhibited highest ACE-I inhibitory activity with IC50 value of 0.517 and 0.521 when hydrolyzed using both trypsin and alcalase, respectively. All the sources selected showed nonsignificant changes in ACE-I inhibitory activity between alcalase and trypsin (p &lt; 0.05) except Bengal gram and chicken. Antioxidant activity was higher in alcalase hydrolyzed egg yolk (43.08%). Conclusion Among the sources, all animal proteins hydrolyzed using alcalase exhibited significantly higher (p &lt; 0.05) antioxidant activity than trypsin. However, protein hydrolysates prepared from sardine using alcalase would yield bioactive peptides with higher ACE-I inhibitory and antioxidant activity. Therefore, use of alcalase as a substitution for trypsin would be economical for large-scale production. Henceforth, food protein hydrolyzed by alcalase can be used as a functional ingredient for the development of functional or nutraceutical foods to combat lifestyle diseases.

Biodegradability of unsaturated poly(ester-thioether)s synthesized by thiol-yne reactions

AbstractWe prepared the dialkynyl monomers 1,4-butanediol dipropiolate, meso-erythritol dipropiolate, and isomannide dipropiolate (IMDP) via the reactions of 1,4-butanediol, meso-erythritol, and isomannide, respectively, with propiolic acid. The thiol-Michael polyaddition of three dialkynyl monomers with several dithiols, namely, 1,4-butanedithiol, ethylene bis(thioglycolate), 3,6-dioxa-1,8-octanedithiol, D,L-dithiothreitol, and D-2,5-di-O-(2-mercaptoacetate)-1,4:3,6-dianhydromannitol (MAIM), proceeded in chloroform, tetrahydrofuran, or N,N-dimethylformamide (25 °C) using triethylamine as the catalyst to give unsaturated poly(ester-thioether)s with the expected structures (Mn, 2.4 × 103 to 22.6 × 103; molecular dispersity index [Mw/Mn], 1.26–2.00). All of the poly(ester-thioester)s had single glass-transition temperature (Tg) values between −27 and 49 °C. While the rigid main chains improved the glass transition temperature, all of the poly(ester-thioester)s showed apparent enzymatic hydrolysis by lipase but low biodegradability in biodegradation tests using activated sludge. The poly(IMDP-alt-MAIM) with the highest biobased degree (55%) showed the highest biodegradability (10%) and the highest Tg (49 °C), suggesting that dianhydrosugars are suitable as biomass for improving biodegradability as well as thermal properties.

Bacterial cellulose from kombucha: Assessing inoculum age and concentration, and its conversion via enzymatic hydrolysis into cellobiose and glucose

Kombucha-type fermentation yields significant amounts of bacterial cellulose alongside the fermented beverage. This study evaluated the effects of inoculum age and concentration in kombucha-type fermentation using green acerola residue as an extract. The findings revealed that lower inoculum concentrations (2.5 and 5%) resulted in decreased bacterial cellulose (BC) yield. Conversely, increasing the inoculum concentration to 7.5% and 10% led to a 3.2-fold increase in BC yield. Inoculums aged just 3 days exhibited the lowest BC production, followed by older inoculums (9 and 12 days), suggesting reduced metabolic activity in the latter case. The optimal combination was found to be an inoculum age of 6 days and a concentration of 7.5%, resulting in a BC yield of up to 2.2 g/L using 70 g/L of carbon source. BC obtained from fermentation underwent enzymatic hydrolysis to yield cellobiose. Multi-stage hydrolysis - 1 stage (48 h), 2 (24 h, 24 h), 3 (12 h, 24 h, 24 h), and 4 (6 h, 6 h, 12 h, 24 h) - utilizing a methodology that anticipates the adsorption of endoglucanases onto the BC itself was employed to mitigate the inhibition of cellobiose on endoglucanases. The stages demonstrated an increase in cellobiose yield corresponding to the greater number of cycles, with 4 stages yielding 12.4% compared to 8.6% for hydrolysis conducted in a single stage. Despite the observed increase, the yield can be considered low and attributed to the loss of adsorbed enzymes between cycles due to the high crystallinity of the BC.

Efficient production of cellooligosaccharides and xylooligosaccharides by combined biological pretreatment and enzymatic hydrolysis process

Efficient production of cellooligosaccharides and xylooligosaccharides by combined biological pretreatment and enzymatic hydrolysis process

Detection and comparison of tumor cell-associated microbiota from different compartments of colorectal cancer

IntroductionIntratumoral microbes play an important role in the development of colorectal cancer (CRC). However, studying intratumoral microbes in CRC faces technical challenges, as tumor microbe communities are often contaminated by fecal microbes due to the structure of the gut folds and villi. The present study aimed to develop a new method for isolating tumor cell-associated microbiota and comparing microbial populations from different compartments.Materials and methodsThe distribution of intestinal bacteria was detected using immunohistochemistry combined with 5R-16s rRNA gene sequencing to explore the effects of the sampling site and number of washes on the detection of microbiota. The 5R-16s rRNA gene sequencing was performed using 44 samples from 11 patients with CRC, including CRC tumor tissues (TT), normal tissues adjacent to CRC (NT), tumor cells (TC), and normal cells (NC). TC and NC were obtained from the TT and NT using an enzymatic digestion method. The microbiota and their potential functions in the four groups were analyzed and compared to determine the differential microbiota related to CRC.ResultsBacteria were mainly distributed in the feces covering intestinal tissues and in the epithelial cells and macrophages within the tissues. Different sampling sites and number of washes led to detection of different microbiota distributions. Although the cleaning method could be controlled, sampling sites varied and led to different microbiota distributions. The phyla of Firmicutes and Bacteroidetes were highly abundant in the conventionally used tissue samples, whereas Proteobacteria was the most abundant phyla in the cell samples isolated with the new method (i.e., after cell enzymatic hydrolysis). Detection of CRC cell-associated microbiota using a cell enzymatic digestion method showed that some bacteria, such as Fusobacterium, Eikenella, Shewanella, and Listeria, were more abundant in TT than NT, whereas the abundance of Akkermansia was lower in TT than NT. The tumor/normal ratios of some bacteria, such as Gemella, Escherichia, Shigella, and Blautia, were different between the cell and tissue samples.ConclusionThe cell enzymatic digestion method reduced fecal bacterial contamination, enabling low biomass intratumoral microbiota to be detected and allowing prediction of bacterial distributions.

Optimization of Ultrasound-enhanced Subcritical Water Hydrolysis of Oil Palm Empty Fruit Bunch for the Production of Fermentable Sugar

To enhance the hydrolysis to produce fermentable sugar, oil palm empty fruit bunch (OPEFB) has undergone ultrasonication pretreatment prior to subjection to subcritical water hydrolysis. This work aims to optimize the effect of temperature, reaction time, and the concentration of sodium dodecyl sulfate (SDS) as the surfactant, with the primary aim of maximizing sugar production in the subcritical water hydrolysis process applied to oil palm empty fruit bunch (OPEFB). The pretreatment process conditions were optimized using response surface methodology of the central composite design (RSM–CCD). The experimental design includes three factors and levels, with a range of 180–220 °C temperature (X1), 60–80 minutes process time (X2), and 1–5% w/w SDS concentration (X3), an α value of 1.68, and reducing sugar concentration (g/L) as response (Y1). The optimum condition for subcritical water hydrolysis of OPEFB was obtained at 208 °C, 78 minutes, and 2.6% w/w SDS concentration with an expected yield of 6.09 g/L. As a result, reducing sugar produced by enzymatic hydrolysis increased by 324.7% compared to raw OPEFB, with sugar yield of 45.64% after 36 hours. Along these, changes in crystallinity, chemical composition, lignocellulosic functional groups, and morphology were analyzed to determine the impact of the pre-treatment on OPEFB.

SntB Affects Growth to Regulate Infecting Potential in Penicillium italicum

Penicillium italicum, a major postharvest pathogen, causes blue mold rot in citrus fruits through the deployment of various virulence factors. Recent studies highlight the role of the epigenetic reader, SntB, in modulating the pathogenicity of phytopathogenic fungi. Our research revealed that the deletion of the SntB gene in P. italicum led to significant phenotypic alterations, including delayed mycelial growth, reduced spore production, and decreased utilization of sucrose. Additionally, the mutant strain exhibited increased sensitivity to pH fluctuations and elevated iron and calcium ion stress, culminating in reduced virulence on Gannan Novel oranges. Ultrastructural analyses disclosed notable disruptions in cell membrane integrity, disorganization within the cellular matrix, and signs of autophagy. Transcriptomic data further indicated a pronounced upregulation of hydrolytic enzymes, oxidoreductases, and transport proteins, suggesting a heightened energy demand. The observed phenomena were consistent with a carbon starvation response potentially triggering apoptotic pathways, including iron-dependent cell death. These findings collectively underscored the pivotal role of SntB in maintaining the pathogenic traits of P. italicum, proposing that targeting PiSntB could offer a new avenue for controlling citrus fungal infections and subsequent fruit decay.

Comparison of the Mechanisms of Hydrolysis of Organophosphates with Good and Poor Leaving Group by Phosphotriesterase from Pseudomonas diminuta

Abstract Combined quantum mechanics/molecular mechanics approaches are used to determine the mechanisms of organophosphate hydrolysis in an active site of Pseudomonas diminuta phosphotriesterase. For a substrate with a good leaving group, the reaction proceeds through two elementary stages with low energy barriers, and a gain in energy is observed. With a poor leaving group, only the formation of an unstable reaction intermediate is possible, and hydrolysis is incomplete. A comparison of the resulting reaction mechanisms explains the experimental kinetic data, according to which the enzyme hydrolyzes only substrates with good leaving groups.

Identification of the Wound Healing Activity Peptidome of Edible Bird's Nest Protein Hydrolysate and the In Silico Evaluation of Its Transport and Absorption Potential in Skin.

In this study, edible bird's nest (EBN) was proven to be a suitable source of bioactive peptides via enzymatic hydrolysis. The ultrafiltration component of the EBN peptides (EBNPs, Mw < 3 000 Da) could be responsible for moderate moisture retention and filaggrin synthesis. It was found that EBNP had a great capacity to protect HaCaT keratinocytes from DNA damage caused by UVB-irradiation and enhance wound healing by increasing the migratory and proliferative potential of cells. Furthermore, the external application of EBNP could effectively repair high glycolic acid concentration-induced skin burns in mice. A total of 1 188 peptides, predominantly the hydrophobic amino acids (e.g., Leu, Val, Tyr, Phe), were identified in the EBNP by liquid chromatography with tandem mass spectrometry (LC-MS/MS). Molecular docking showed that hydrophobic tripeptides from EBNP had a good binding affinity to proton-dependent oligopeptide transporter PepT1. Our data indicated that the hydrophobic amino acid-rich EBNP plays an important role in skin wound healing.