Enzymatic Process Research Articles

Isolation and Characterization of Insecticidal Cyclotides from Viola communis.

Cyclotides are cysteine-rich plant-derived peptides composed of 28-37 amino acids with a head-to-tail cyclic backbone and a knotted arrangement of three conserved disulfide bonds. Their beneficial biophysical properties make them promising molecules for pharmaceutical and agricultural applications. The Violaceae plant family is the major cyclotide-producing family, and to date, every examined plant from this family has been found to contain cyclotides. The presence of cyclotides in Viola communis was inferred by mass spectroscopy previously, but their sequences and properties had yet to be explored. In this study, the occurrence of cyclotides in this plant was investigated using proteomics and transcriptomics. Twenty cyclotides were identified at the peptide level, including two new members from the bracelet (Vcom1) and Möbius (Vcom2) subfamilies. Structural analysis of these newly identified peptides demonstrated a similar fold compared with cyclotides from the same respective subfamilies. Biological assays of Vcom1 and Vcom2 revealed them to be cytotoxic to Sf9 insect cell lines, with Vcom1 demonstrating higher potency than Vcom2. The results suggest that they could be further explored as insecticidal agents and confirm earlier general findings that bracelet cyclotides have more potent insecticidal activity than their Möbius relatives. Seven new cyclotide-like sequences were observed in the transcriptome of V. communis, highlighting the Violaceae as a rich source for new cyclotides with potential insecticidal activity. An analysis of sequences flanking the cyclotide domain in the various precursors from V. communis and other Violaceae plants revealed new insights into cyclotide processing and suggested the possibility of two alternative classes of N-terminal processing enzymes for cyclotide biosynthesis.

Application of Hierarchically Porous Chitosan Monolith for Enzyme Immobilization.

Enzyme immobilization is a crucial technique for improving the stability of enzymes. Compared with free enzymes, immobilized enzymes offer several advantages in industrial applications. Efficient enzyme immobilization requires a technique that integrates the advantages of physical absorption and covalent binding while addressing the limitations of conventional support materials. This study offers a practical approach for immobilizing α-amylase on a hierarchically porous chitosan (CS) monolith. An optimized CS monolith was fabricated using chemically modified chitin by thermally induced phase separation. By combining physical adsorption and covalent bonding, this technique leverages the amino and hydroxy groups present in CS to facilitate effective enzyme binding and stability. α-Amylase immobilized on the CS monolith demonstrated excellent stability, reusability, and increased activity compared to its soluble counterpart across various pH levels and temperatures. In addition, the CS monolith exhibited a significant potential to immobilize other enzymes, namely, lipase and catalase. Immobilized lipase and catalase exhibited higher loading capacities and enhanced activities than their soluble forms. This versatility highlights the broad applicability of CS monoliths as support materials for various enzymatic processes. This study provides guidelines for fabricating hierarchical porous monolith structures that can provide efficient enzyme utilization in flow systems and potentially enhance the cost-effectiveness of enzymes in industrial applications.

Effect of storage condition on the viability of sago effluents as carbon source in fermentation medium for bioethanol production

In Sarawak, Malaysia, approximately 237 tons/day of sago effluent is commonly discharged into nearby river due to the sago starch extraction process. Due to the high concentration of polymeric compounds, particularly starch, in sago wastewater, which petrifies easily, this condition severely pollutes the environment in the affected area. This study was conducted to determine the viability of using sago effluent as a carbon source and fermentation medium for bioethanol production which indirectly help to minimize the environmental impact as well as the economics of the sago industry. The sago effluent obtained from the local sago mill was analysed for starch content and pH profile while stored at room and cold (4°C) temperature facility. Enzymatic hydrolysis was conducted to convert the residual starch into glucose as carbon source for bioethanol fermentation. Fresh sago effluent can be stored for up to 5 days in cold temperature where the starch content remains constant. The highest starch concentration in sago effluent was 61.33 g/L, in which 50.57 g/L glucose was obtained through the enzymatic hydrolysis process. Hence 82.5% of the starch to glucose conversion yield is revealed. Then, the sago effluent hydrolysate which acts as a carbon source as well as a fermentation medium able to generate 23.14 g/L of bioethanol, displays a 91% theoretical yield of glucose to ethanol. In conclusion, the utilization of sago wastewater as feasible alternative to cheap and locally available and sustainable source of raw materials to produce bioethanol.

Enzymatic characterization and synergistic effects of protease combinations on DPP-IV inhibitory peptide release from bovine casein

This study focused on the enzymatic characteristics, with regard to primary specificity, secondary specificity, and hydrolysis ability of five proteases (Alcalase, Protamex, papain, Flavourzyme, and ProtexA), towards the release of dipeptidyl peptidase-IV (DPP-IV) inhibitory peptides from bovine casein. The secondary specificity of the five proteases, especially the release capability for Xaa-Pro-type peptide played a determining role in the release of DPP-IV inhibitory peptides from bovine casein. Moreover, protease combinations, chosen based on their Xaa-Pro-type peptide release abilities, resulted in superior DPP-IV inhibitory activity compared to other combinations. Particularly, the hydrolysate with the highest DPP-IV inhibitory activity (IC50 value of 0.45 ± 0.07 mg/mL) was obtained by the combination hydrolysis of ProtexA and papain, achieving a 74.63% ± 1.62% (peak area) release of Xaa-Pro-type peptide, and a significant improving glucose tolerance in mice (33.42% ± 1.68% reduction of blood glucose AUC compared with control mice). The mechanistic insights revealed that papain efficiently released long Xaa-Pro-type peptides in the early hydrolysis stage, and subsequently, the synergistic effect of papain and ProtexA led to an accelerated and extensive release of short Xaa-Pro-type peptides. These findings provide valuable insight into enzymatic processes for generating DPP-IV inhibiting peptides from casein.

Acetylation proteomics and metabolomics analyses reveal the involvement of starch synthase undergoing acetylation modification during UV-B stress resistance in Rhododendron Chrysanthum Pall

BackgroundRhododendron chrysanthum Pall. (R. chrysanthum) is a plant that lives in high mountain with strong UV-B radiation, so R. chrysanthum possess resistance to UV-B radiation. The process of stress resistance in plants is closely related to metabolism. Lysine acetylation is an important post-translational modification, and this modification process is involved in a variety of biological processes, and affected the expression of enzymes in metabolic processes. However, little is known about acetylation proteomics during UV-B stress resistance in R. chrysanthum.ResultsIn this study, R. chrysanthum OJIP curves indicated that UV-B stress damaged the receptor side of the PSII reaction center, with a decrease in photosynthesis, a decrease in sucrose content and an increase in starch content. A total of 807 differentially expressed proteins, 685 differentially acetylated proteins and 945 acetylation sites were identified by quantitative proteomic and acetylation modification histological analysis. According to COG and subcellular location analyses, DEPs with post-translational modification of proteins and carbohydrate metabolism had important roles in resistance to UV-B stress and DEPs were concentrated in chloroplasts. KEGG analyses showed that DEPs were enriched in starch and sucrose metabolic pathways. Analysis of acetylation modification histology showed that the enzymes in the starch and sucrose metabolic pathways underwent acetylation modification and the modification levels were up-regulated. Further analysis showed that only GBSS and SSGBSS changed to DEPs after undergoing acetylation modification. Metabolomics analyses showed that the metabolite content of starch and sucrose metabolism in R. chrysanthum under UV-B stress.ConclusionsDecreased photosynthesis in R. chrysanthum under UV-B stress, which in turn affects starch and sucrose metabolism. In starch synthesis, GBSS undergoes acetylation modification and the level is upregulated, promotes starch synthesis, making R. chrysanthum resistant to UV-B stress.

Optimization of Enzymatic Transesterification of Acid Oil for Biodiesel Production Using a Low-Cost Lipase: The Effect of Transesterification Conditions and the Synergy of Lipases with Different Regioselectivity.

This study describes the enzymatic production of second-generation biodiesel using low-quality acid oil as a substrate. Biolipasa-R, a commercially available and low-cost lipase, was employed for enzymatic transesterification. Response surface methodology was applied to optimize the enzymatic transesterification process. The optimal conditions for biodiesel production, which comprised 42% lipase concentration (per weight of oil), 32% water content (per weight of oil), a methanol to oil molar ratio of 3:1, pH 7.0 and reaction temperature 30°C, resulted in the highest fatty acid methyl ester (FAME) content (71.3%). Subsequently, the synergistic effect of two lipases with different regioselectivities under the optimum transesterification conditions was studied, aiming at the enhancement of process efficiency. The transesterification efficiency of immobilized Biolipasa-R was determined and compared to that of Biolipasa-R in its free form. The results revealed a good performance on FAME content (66.5%), while the recycling of immobilized lipase resulted in a decrease in transesterification efficiency after three consecutive uses.

Efficient Enzymatic Saccharification of Agricultural Wastes for the Production of Bioethanol, D-Allulose and lactic acid

The demand for renewable resources to replace fossil fuels has increased. Fruit and agricultural wastes can be fermented to yield biofuels and biochemicals. However, the high cost of the feedstock and limitations of the catalytic process hinder the application of such wastes. Therefore, we aimed to develop an efficient enzymatic saccharification process, without pretreatment, for fruit and agricultural wastes. The conversion rate of the mixed agricultural wastes (MAW) to fermentable sugars was approximately 91% after 24h. The ethanol yield increased by 4.5% after limonene removal. The D-allulose yield in the hydrolysate was 4.6mg/mL at 4 °C and 3.3mg/mL at 50 °C, whereas the fructose yield in the sugar medium was 13.2mg/mL at 4°C, demonstrating a high conversion yield of 73.2%. Lactic acid was produced at a conversion rate of approximately 67.4%. Therefore, this study presents a novel approach of the biosynthesis of functional sugars and chemicals from waste biomass, introducing a cost-effective enzymatic saccharification process that bypasses pretreatment, thereby enabling the production of biofuels, biochemicals, and functional sugars and opening up a promising economic opportunity in the field.

Transformation of agricultural wastes into functional oligosaccharides using enzymes and emerging technologies.

Pectin-oligosaccharides (POS) serve diverse purposes as a food ingredient, antimicrobial and biostimulant in plants, and their functionality is linked to the degree of esterification. Grape and broccoli wastes emerge as environmentally friendly alternatives to obtaining pectin, serving as a sustainable source to producing POS. For example, microwaves have proven to be an effective and sustainable method to extract polysaccharides from plant matrices. This work aims to use grape and broccoli wastes as alternative sources for obtaining pectin by microwave-assisted extraction and biotransformation into POS, which possess biological properties. The extraction conditions were identified at a power of 400 W, 300 s for the extraction of pectin from grape pomace and broccoli waste. Biotransformation of pectins into POS, using commercial enzyme preparations (Viscozyme L and Pectinase). Characterisation was carried out by Fourier-transform infrared spectroscopy, thermogravimetric analysis, and scanning electron microscopy. Physicochemical analysis indicated grape pomace and broccoli waste pectins had galacturonic acid content of 63.81 ± 1.67 and 40.83 ± 2.85 mg 100 mg-1, low degree of esterification of 34.89% and 16.22%, respectively. Biotransformation of pectins into POS resulted in a 20% hydrolysis rate. The main enzymatic activity was polygalacturonase for the degradation of the main structure of the pectin. Production of POS from agro-industrial wastes by emerging technologies, such as the combined use of microwave-assisted extraction and enzymatic processes, represents an alternative method for the generation of bioactive compounds with distinctive properties suitable for different applications of interest.

Source identification and spatial distribution of phthalate acid esters in fish and sediments along the Persian Gulf coast

A quantification of phthalate acid esters (PAEs) concentrations was performed on sediment and fish samples collected from industrial, urban, and rural regions along the Persian Gulf coastline from November 2020 to March 2021. The results indicated that the mean concentrations of ƩPAEs in sediment samples from urban, industrial, and rural regions were 1486, 855, and 257 ng/g, respectively. In fish samples, the corresponding values were 392, 284, and 62.4 ng/g. The mean concentrations of ƩPAEs in sediment/fish samples from urban and industrial areas were significantly higher (P < 0.05) than in rural areas. DEHP had the highest level and detection frequency (DF = 100 %) of the PAEs congeners studied. The DEHP concentrations in sediment samples from the Urban, Industrial, and Rural regions were 283 ± 37.5, 219 ± 15.7, and 44.9 ± 19.5 ng/g, respectively, while in fish samples, they were 904 ± 272, 564 ± 60.5, and 146 ± 43.5 ng/g, respectively. The results of the ecotoxicological effects assessment also revealed that the Persian Gulf coasts are significantly contaminated with PAEs, which may have substantial effects on the aquatic ecosystem of the region (including the composition of aquaculture communities, enzymatic processes, and marine culture microorganisms). As a result, there is an urgent need for decision-makers and policymakers to enact laws that prohibit the discharge of PAEs-containing waste into the Persian Gulf coasts to protect the marine ecosystem.

From laboratory to industrial use: Understanding process variation during enzymatic protein hydrolysis with dry film Fourier-Transform Infrared spectroscopy

Industries are continuously looking for dedicated sensor solutions for evaluating the chemical composition of products that can provide valuable insights into process control, process optimization, and product quality. A rapid and robust analytical method, Fourier-Transform Infrared (FTIR) spectroscopy, holds significant potential in this regard as it provides detailed compositional values of food nutrients. The main aim of the present study was to use dry film FTIR spectroscopy as an analytical tool to characterize products from an industrial enzymatic protein hydrolysis process and link this to understand industrial process variations. For this purpose, 463 protein hydrolysate samples were obtained from an industrial enzymatic protein hydrolysis plant. In the same period, systematic variations in process parameters such as raw material composition, enzyme type, and water addition, were performed. All samples were analyzed using dry film FTIR spectroscopy. Two subsets containing 200 and 68 hydrolysate samples were chosen for measuring average molecular weight (AMW) and collagen content respectively, providing reference data for constructing calibration models based on partial least squares regression (PLSR). The percentage of low molecular weight constituents was derived from the molecular weight distribution of size exclusion chromatograms of protein hydrolysates and also used in the modeling. Calibration models for the prediction of AMW, low molecular weight constituents, and collagen content were obtained with a good fit and lower estimation errors. Principal component analysis (PCA) of protein hydrolysates’ FTIR spectra effectively differentiated process variations (enzyme types and raw materials) without extensive reference analysis. One-factor analysis of variance (ANOVA) tests was used to observe the impact of process variation on product quality. FTIR proved to be a sensitive method for liquid protein analysis and process control with a significant potential in process optimization approaches. To the authors’ knowledge, this is the first time dry film FTIR spectroscopy has been used to evaluate an industrial bioprocess with designed process variations.

Green energy breakthroughs: Harnessing nano-catalysts and enzymatic catalysts for bioenergy generation

Microbial fuel cells and anaerobic digestion are promising bioenergy methods but inefficient and scalable. With its exceptional surface area-to-volume ratio, nano-catalysts are revolutionizing this field. Researchers have improved energy conversion efficiency by immobilizing or embedding nanoparticles on electrodes or catalytic supports to promote electron transport. Nanomaterials like graphene-based catalysts and metal nanoparticles speed up organic matter breakdown and make bioenergy systems more profitable. Researchers are working harder to find new bioenergy-generating methods to address environmental concerns and the demand for sustainable energy. The study discusses how nano-catalysts and enzymatic catalysts are revolutionizing bioenergy production in green energy. Additionally, enzymatic catalysts are the best bioenergy catalysts. Highly specialized, biocompatible, and renewable enzymes provide unrivaled biofuel production potential. Protein engineering and immobilization have improved enzymes for bioenergy systems. Enzymatic catalysts enable biofuel generation from more feedstocks without harsh chemical procedures by controlling biological reactions quickly and precisely. Combining nano- and enzymatic catalysts is promising. Nano-catalysts improve enzymatic processes by creating a favorable microenvironment, enhancing surface binding sites, and enabling substrate transport. This combination may maximize bioenergy systems' biofuel output and wastewater treatment capabilities. According to this study, integrating nano- and enzymatic catalysts is a green energy breakthrough. These advances will improve bioenergy generation by overcoming current techniques, opening new sustainable energy production prospects, and expediting our transition to a greener, more sustainable future.

Corrigendum to “Use of enzymatic processes in the tanning of leather materials”

Corrigendum to “Use of enzymatic processes in the tanning of leather materials”

Extraction of Kratom (Mitragyna speciosa) leaf compounds by enzymatic hydrolysis-assisted process: Yield, characteristics and its in vitro cytotoxicity in cell lines

This study aimed to develop an antioxidative compound extraction method for Kratom (Mitragyna speciosa) leaves using cellulase and pectinase, in combination and alone. The enzymatic hydrolysis-assisted process (EA) successfully enhanced antioxidant extraction from kratom leaves. The single enzymatic hydrolysis-assisted process (SEA) with 6% cellulase (C6) provided the highest extraction yield. The mixed enzymatic hydrolysis-assisted process (MEA) was then developed using C6, mixed with different concentrations of pectinase. Based on the high extraction yield, the MEA using 6% cellulase and 2% pectinase (C6+P2) was selected for further studies in comparison with that from relevant SEA and the control (without EA). Different extraction processes altered the characteristics and antioxidative activities of resulting extracts. The antioxidative activity of C6+P2 was dramatically enhanced in the gastrointestinal tract model system. Fourier transform infrared (FT-IR) spectrometer results confirmed significant content of phenolic compounds, proteins, and polysaccharides in C6+P2. From in vitro cytotoxicity study, C6+P2 showed an IC50 value of 22.86 and 4.76 µg/mL in RAW264.7 and Caco-2 cells, respectively. The bioactive compounds from C6+P2 should be identified in further studies to facilitate their application in the food and pharmaceutical industries.

Enzymatic degradability of diclofenac ozonation products: A mechanistic analysis

The treatment of waterborne micropollutants, such as diclofenac, presents a significant challenge to wastewater treatment plants due to their incomplete removal by conventional methods. Ozonation is an effective technique for the degradation of micropollutants. However, incomplete oxidation can lead to the formation of ecotoxic by-products that require a subsequent post-treatment step. In this study, we analyze the susceptibility of micropollutant ozonation products to enzymatic digestion with laccase from Trametes versicolor to evaluate the potential of enzymatic treatment as a post-ozonation step. The omnipresent micropollutant diclofenac is used as an example, and the enzymatic degradation kinetics of all 14 detected ozonation products are analyzed by high-performance liquid chromatography coupled with high-resolution mass spectrometry (HPLC-HRMS) and tandem mass spectrometry (MS2). The analysis shows that most of the ozonation products are responsive to chemo-enzymatic treatment but show considerable variation in enzymatic degradation kinetics and efficiencies. Mechanistic investigation of representative transformation products reveals that the hydroxylated aromatic nature of the ozonation products matches the substrate spectrum, facilitating their rapid recognition as substrates by laccase. However, after initiation by laccase, the subsequent chemical pathway of the enzymatically formed radicals determines the global degradability observed in the enzymatic process. Substrates capable of forming stable molecular oxidation products inhibit complete detoxification by oligomerization. This emphasizes that it is not the enzymatic uptake of the substrates but the channelling of the reaction of the substrate radicals towards the oligomerization of the substrate radicals that is the key step in the further development of an enzymatic treatment step for wastewater applications.

Biomonitoring urinary pesticide metabolites in preschool children by supported liquid extraction and ultra-high performance liquid chromatography-tandem mass spectrometry and their association with oxidative stress

Investigating pesticide exposure and oxidative stress in preschool children is essential for elucidating the determinants of environmental health in early life, with human biomonitoring of urinary pesticide metabolites serving as a critical strategy for achieving this objective. This study demonstrated biomonitoring of 2 phenoxyacetic acid herbicides, 2 organophosphorus pesticide metabolites, and 4 pyrethroid pesticide metabolites in 159 preschool children and evaluated their association with oxidative stress biomarker 8-hydroxydeoxyguanosine. An enzymatic deconjugation process was used to release urinary pesticide metabolites, which were then extracted and enriched by supported liquid extraction, and quantified by ultra-high performance liquid chromatography-tandem mass spectrometry with internal standard calibration. Dichloromethane: methyl tert‑butyl ether (1:1, v/v) was optimized as the solvent for supported liquid extraction, and we validated the method for linear range, recovery, matrix effect and method detection limit. Method detection limit of the pesticide metabolites ranged from 0.01 μg/L to 0.04 μg/L, with satisfactory recoveries ranging from 70.5 % to 95.5 %. 2,4,5-Trichlorophenoxyacetic acid was not detected, whereas the other seven pesticide metabolites were detected with frequencies ranging from 10.1 % to 100 %. The concentration of urinary pesticide metabolites did not significantly differ between boys and girls, with the median concentrations being 9.39 μg/L for boys and 4.90 μg/L for girls, respectively. Spearman correlation analysis indicated that significant positive correlations among urinary metabolites. Bayesian kernel machine regression revealed a significant positive association between urinary pesticide metabolites and 8-hydroxydeoxyguanosine. Para-nitrophenol was the pesticide metabolite that contributed significantly to the elevated level of oxidative stress.

Direct Imaging of Protein Clusters in Metal-Organic Frameworks.

Protein@metal-organic frameworks (P@MOFs) prepared by coprecipitation of protein, metal ions, and organic ligands represent an effective method for protein stabilization with a wide spectrum of applications. However, the formation mechanism of P@MOFs via the coprecipitation process and the reason why proteins can retain their biological activity in the frameworks with highly concentrated metal ions remain unsettled. Here, by a combined methodology of single molecule localization microscopy and clustering analysis, we discovered that in this process enzyme molecules form clusters with metal ions and organic ligands, contributing to both the nucleation and subsequent crystal growth. We proposed that the clusters played an important role in the retention of overall enzymatic activity by sacrificing protein molecules on the cluster surface. This work offers fresh perspectives on protein behaviors in the formation of P@MOFs, inspiring future endeavors in the design and development of artificial bionanocomposites with high biological activities.

Vitamin D3 Regulates Energy Homeostasis under Short-Term Fasting Condition in Zebrafish (Danio Rerio).

Vitamin D3 (VD3) is a steroid hormone that plays pivotal roles in pathophysiology, and 1,25(OH)2D3 is the most active form of VD3. In the current study, the crucial role of VD3 in maintaining energy homeostasis under short-term fasting conditions was investigated. Our results confirmed that glucose-depriving pathways were inhibited while glucose-producing pathways were strengthened in zebrafish after fasting for 24 or 48 h. Moreover, VD3 anabolism in zebrafish was significantly suppressed in a time-dependent manner under short-fasting conditions. After fasting for 24 or 48 h, zebrafish fed with VD3 displayed a higher gluconeogenesis level and lower glycolysis level in the liver, and the serum glucose was maintained at higher levels, compared to those fed without VD3. Additionally, VD3 augmented the expression of fatty acids (FAs) transporter cd36 and lipogenesis in the liver, while enhancing lipolysis in the dorsal muscle. Similar results were obtained in cyp2r1-/- zebrafish, in which VD3 metabolism is obstructed. Importantly, it was observed that VD3 induced the production of gut GLP-1, which is considered to possess a potent gluconeogenic function in zebrafish. Meanwhile, the gene expression of proprotein convertase subtilisin/kexin type 1 (pcsk1), a GLP-1 processing enzyme, was also induced in the intestine of short-term fasted zebrafish. Notably, gut microbiota and its metabolite acetate were involved in VD3-regulated pcsk1 expression and GLP-1 production under short-term fasting conditions. In summary, our study demonstrated that VD3 regulated GLP-1 production in zebrafish by influencing gut microbiota and its metabolite, contributing to energy homeostasis and ameliorating hypoglycemia under short-term fasting conditions.

Sustainable leather tanning: Enhanced properties and pollution reduction through crude protease enzyme treatment

In this study, proteolytic bacteria, particularly Pseudomonas aeruginosa strain SM4 (OQ349573), were isolated from tannery solid waste dumping yard soil and employed to produce extracellular protease enzymes. The bacteria exhibited optimal growth after 30 h of incubation at 37 °C and pH 7. Under conditions of 55 °C, pH 8, and a substrate concentration of 2 %, the crude enzyme displayed its highest activity at 105 UmL−1. Notably, the produced crude enzyme showed no discernible inhibitory effects on detergents, metal salts, or organic solvents. Application of the crude protease at concentrations of 3 % and 2 % in chrome tanning of goatskins (GS) and cowhides (CH), respectively, yielded significant reductions of 35 % and 30 % in chromium and other post-tanning chemicals compared to conventional processes. Despite the 30 to 35 % reduction in tanning and post-tanning chemicals, the uptake of chrome and associated chemicals by crust leather was higher than observed in conventional processes. Chromium content analysis of the effluent revealed an 81 % reduction during piloting in real industrial operations, accompanied by reductions of about 46 % in BOD and COD pollution loads. The finished leather obtained from the enzymatic process exhibited superior mechanical properties, including higher tensile strengths (210 and 195 kg cm-2), stitch tear (92 and 165 kg cm−1), grain crack load (28 and 32 kg), and distension (73 and 62 mm) for GS and CH, respectively, surpassing or closely aligning with standard values and those obtained in conventional processes.

MhZAT10L positively modulates NaCl-induced cell death by triggering MhVPEγ transcription in Malus hupehensis

Excess sodium chloride (NaCl) in soil causes several secondary stresses for plants. The plant-specific vacuolar processing enzyme (VPE) plays a positive role in regulating stress-induced cell death. Here, we observed that NaCl treatment leads to a significant increase in cell death and upregulation of MhVPEγ expression in Malus hupehensis roots. Meanwhile, we also identified a C2H2-type transcription factor upstream of MhVPEγ, MhZAT10L, which positively regulated MhVPEγ transcription. The transcription of MhZAT10L was induced by NaCl. And genetic experiments using transgenic Malus hupehensis roots and apple calli showed that the suppression of MhZAT10L decreased their sensitivity to NaCl, as well as mitigated the generation of reactive oxygen species (ROS), cell death, and MhVPEγ expression induced by NaCl. Conversely, the overexpression of MhZAT10L enhanced their sensitivity to NaCl, accelerated ROS generation, and promoted cell death by directly activating MhVPEγ transcription. Overall, our findings indicate that MhZAT10L positively regulates NaCl-induced cell death by activating MhVPEγ transcription.

Efficient enzymatic synthesis of vitamin E succinate using an organic solvent‐stable immobilized lipase

AbstractVitamin E succinate has gained substantial attention as a potential therapeutic agent for cancer treatment due to its biomedical activities. One of the prominent methods of synthesizing vitamin E succinate is through enzymatic processes, which, although advantageous, presents inherent challenges related to optimization, scalability, and particularly, the poor stability of lipases in organic solvents. Our study addresses these challenges by conducting a comprehensive comparative analysis between Lipase UM1 and three other immobilized commercial lipases, demonstrating Lipase UM1's enhanced resistance to organic solvents and its superior efficiency in vitamin E succinate production. Further optimization experiments with Lipase UM1 led to an unprecedented conversion of 99%. Additionally, we scaled the reaction to a proof‐of‐concept industrial level. The synthesized product was verified using Fourier transform infrared spectroscopy and nuclear magnetic resonance analysis, ensuring its quality and consistency. This study validates Lipase UM1 as an efficient catalyst for vitamin E succinate synthesis, offering a promising avenue for industrial production with potential applications in cancer therapy and beyond.