Notable progress has been achieved in the past two decades regarding production of different enzymatic mixtures for hydrolysis of the lignocellulose matrix. Nevertheless, the hydrolysing mixtures remain slow and require tempering, which results in high-energy demands and bad financial results. Use of acids or alkali at a very high temperature and pressure accelerates the process more than ten times wherein the energy requirements are approximately equal. However, these elevated reaction conditions might cause the breakdown of complex lignin formula into substances that have the potential to inhibit subsequent fermentation processes. Formation of these breakdown products may be prevented by selecting the optimum process parameters, but their acquisition requires either a large number of expensive analytical techniques or equally large amounts of slow fermentation tests. An inexpensive and time saving alternative that is based on the sensitivity of chosen organisms to these inhibitors was designed and financially assessed. It was confirmed that the method is technically feasible and economically viable with significant potential to reduce the bioethanol production cost.

• Klíčová slova
• Bioethanol, Financial analysis, Process management, Techno-economical assessment, Valuation,
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

Laccases are multi-copper oxidoreductases with broad biotechnological applications. Here, we report detailed biochemical characterization of purified recombinant laccases originating from Myceliophthora thermophila (MtL) and Trametes trogii (TtL). We identified optimal conditions for decolorization of commercial dyes and textile wastewater samples. We also tested the toxicity of decolorized wastewater samples using human peripheral blood mononuclear cells. MtL and TtL were expressed in Saccharomyces cerevisiae, and secreted enzymes were purified by consecutive hydrophobic and gel chromatography. The molecular masses of TtL (~ 65 kDa) and MtL (> 100 kDa) suggested glycosylation of the recombinant enzymes. Deglycosylation of MtL and TtL led to 25% and 10% decreases in activity, respectively. In a thermal stability assay, TtL retained 61% and MtL 86% of the initial activity at 40 °C. While TtL retained roughly 50% activity at 60 °C, MtL lost stability at temperatures higher than 40 °C. MtL and TtL preferred syringaldazine as a substrate, and the catalytic efficiencies for ABTS oxidation were 7.5 times lower than for syringaldazine oxidation. In the presence of the mediator HBT, purified TtL almost completely decolorized dyes within 30 min and substantially decolorized wastewater samples from a textile factory (up to 74%) within 20 h. However, products of TtL-catalyzed decolorization were more toxic than MtL-decolorized products, which were almost completely detoxified.

• Klíčová slova
• Decolorization, Laccases, PBMCs, Textile dyes, Toxicity, Wastewater,
• Publikační typ
• časopisecké články MeSH

Sulfate-reducing bacteria (SRB) are widespread in various ecotopes despite their growth and enzymatic features not compared. In this study, the enzymatic parameters of ATP sulfurylase in cell-free extracts of sulfate-reducing bacteria isolated from various ecotopes such as soils, corrosion products and human large intestine were determined. Comparative analysis of both enzyme characteristics and growth parameters were carried out and similar research has not been reported yet. The initial and maximum rates of enzymatic reaction catalyzed by ATP sulfurylase were significantly different (p < 0.05) in the bacterial strains isolated from various environmental ecotopes. The specific activity of this enzyme in sulfate-reducing bacteria was determined for corrosive and intestinal strains 0.98-1.56 and 0.98-2.26 U × mg-1 protein, respectively. The Michaelis constants were 1.55-2.29 mM for corrosive and 2.93-3.13 mM for intestinal strains and the affinity range were demonstrated. Based on cluster analysis, the parameters of physiological and biochemical characteristics of sulfate-reducing bacteria from different ecotopes are divided into 3 clusters corresponding to the location of their isolation (soils, heating systems and human intestine). Understanding the enzymatic parameters of the initial stages of sulfate consumption in the process of dissimilatory sulfate reduction will allow the development of effective methods for controlling the production of toxic metabolites, including hydrogen sulfide.

• Klíčová slova
• ATP sulfurylase, Cell-free extracts, Ecotopes, Sulfate-reducing bacteria,
• Publikační typ
• časopisecké články MeSH

UNLABELLED: The ability to predict the transglycosylation activity of glycosidases by in silico analysis was investigated. The transglycosylation abilities of 7 different β-d-galactosidases from GH family 2 were tested experimentally using 7 different acceptors and p-nitrophenyl-β-d-galactopyranoside as a donor of galactosyl moiety. Similar transglycosylation abilities were confirmed for all enzymes originating from bacteria belonging to Enterobacteriaceae, which were able to use all tested acceptor molecules. Higher acceptor selectivity was observed for all others used bacterial strains. Structure models of all enzymes were constructed using homology modeling. Ligand-docking method was used for enzymes-transglycosylation products models construction and evaluation. Results obtained by in silico analysis were compared with results arisen out of experimental testing. The experiments confirmed that significant differences in transglycosylation abilities are caused by small differences in active sites composition of analyzed enzymes. According to obtained result, it is possible to conclude that homology modeling may serve as a quick starting point for detection or exclusion of enzymes with defined transglycosylation abilities, which can be used for subsequent synthesis of e.g., pharmaceutically interesting glycosides. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02715-w.

• Klíčová slova
• Carbohydrate family, Catalysis, Homology modeling, Hydrolases, Ligand-docking,
• Publikační typ
• časopisecké články MeSH

An innovative amperometric immunosensor has been developed to detect antibiotic colistin from the chicken liver. Colistin is a antibacterial peptide that has been barred for human consumption, but it is being commonly used as a veterinary drug, and as a feed additive for livestock. In the present work, an immunosensor was developed by immobilizing an anti-colistin Ab onto the CNF/AuNPs surface of the screen-printed electrode. The sensor records electrochemical response in the chicken liver spiked with colistin with CV. Additionally, the characterization of electrode surface was done with FE-SEM, FTIR, and EIS at each step of fabrication. The lower LOD was 0.89 μgKg-1, with a R 2 of 0.901 using CV. Further validation of the immunosensor was conducted using commercial chicken liver samples, by comparing the results to those obtained using traditional methods. The fabricated immunosensor showed high specificity towards colistin, which remained stable for 6 months but with a 13% loss in the initial CV current.

• Klíčová slova
• Chicken liver, Colistin antibiotic, Immunosensor, Specificity, Toxicity,
• Publikační typ
• časopisecké články MeSH

UNLABELLED: In the course of time, scientific communities have a growing interest in understanding ethano medicines. The Putranjiva roxburghii, a native plant of the Indian Subcontinent is described as a "Child amulet tree" in Ayurveda. Based on the fact that this herbal medicine has an indispensable component of integrative medicine, the present study was planned to assess the effect of ethanolic dried extract of Putranjiva seeds on the motility of X and Y-bearing bovine spermatozoa. The in-vitro effect of seed extract diluted in S-TALP medium on bull semen has been evaluated by Computer Assisted Semen Analysis (CASA) shows a marked increase in the motility of spermatozoa. Motile and non-motile spermatozoa have been separated by glass wool column from the control as well as treated group. The X and Y-bearing sperm quantification have been carried out by droplet digital polymerase chain reaction (ddPCR). The extract didn't exert any differential effect on the motility and viability of X and Y chromosome-bearing spermatozoa. The transcriptome profiling (RNA-Seq) identified 93 differentially expressed genes between the extract treated and control group. It unveils the up-regulation of CATSPER, AKAP3, SPAG, ADAM1B, ADAM2 and ADAM32 genes that are involved in increasing sperm motility. Transcriptome profile also unveil the expression of ZAR1, CYP17A1, APPL2, HOXB4 and SP9 genes involved with embryonic development processes in Putranjiva extract-treated motile spermatozoa. The results envisaged the medicinal value of Putranjiva herb on increased fertility due to combinatory effect like increased sperm motility and favourableness on embryogenesis. The study ruled out the possibility of herbs having any biased effect on the selection of either male or female-bearing spermatozoa in the bull. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-022-03452-4.

• Klíčová slova
• CASA, Infertility, Putranjiva roxburghii, RNA-sequence profiling, ddPCR,
• Publikační typ
• časopisecké články MeSH

UNLABELLED: Staphylococcus aureus is a lethal pathogen that can cause various bacterial infections. This study targets the CrtM enzyme of S. aureus, which is crucial for synthesizing golden carotenoid pigment: staphyloxanthin, which provides anti-oxidant activity to this bacterium for combating antimicrobial resistance inside the host cell. The present investigation quests for human SQS inhibitors against the CrtM enzyme by employing structure-based drug design approaches including induced fit docking (IFD), molecular dynamic (MD) simulations, and binding free energy calculations. Depending upon the docking scores, two compounds, lapaquistat acetate and squalestatin analog 20, were identified as the lead molecules exhibit higher affinity toward the CrtM enzyme. These docked complexes were further subjected to 100 ns MD simulation and several thermodynamics parameters were analyzed. Further, the binding free energies (ΔG) were calculated for each simulated protein-ligand complex to study the stability of molecular contacts using the MM-GBSA approach. Pre-ADMET analysis was conducted for systematic evaluation of physicochemical and medicinal chemistry properties of these compounds. The above study suggested that lapaquistat acetate and squalestatin analog 20 can be selected as potential lead candidates with promising binding affinity for the S. aureus CrtM enzyme. This study might provide insights into the discovery of potential drug candidates for S. aureus with a high therapeutic index. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-023-03862-y.

• Klíčová slova
• Anti-virulence, CrtM, Docking, MM-GBSA, Molecular dynamics, Staphyloxanthin virtual screening,
• Publikační typ
• časopisecké články MeSH

During walking, the foot is in direct contact with the insole of the footwear, and it affects the biomechanics of the whole human body. The aim of this study was to compare the plantar pressure distribution while walking in the same footwear in three different situations: with an original footwear insole, with an ethylene-vinyl acetate customized insole, and with a 3D printed anatomical insole to find out the optimal insoles for people with a normal, flat, and high arched foot according to the plantar pressure relief. Fifty-one adults were grouped into three groups (normal foot, flat foot, and high arched foot geometry). The feet of the randomly selected subjects from each group were scanned, and the insoles were designed on the basis of the foot model. These models were then used for the 3D printing. The plantar pressure distribution was observed when walking in all three different insole types for each of the three subjects with the different foot types. The peak pressure values in the forefoot, midfoot, and rearfoot and contact area were compared. The results of the study show that customized insoles influence the plantar pressure distribution mainly in people with a flat foot or with a high arched foot deformity. This influence is dependent on the type of the customized insoles, mainly in people with a high arched foot. People with a flat foot deformity have a similar plantar pressure relief in the thermoplastic insoles as those in the 3D printed insoles, and both these customized insoles can be used as a supplementary therapeutic instrument. Our study showed that the biggest attention, with regards to customizing insoles, is required by people with a high arched foot deformity, and we recommend that these people use fully customized insoles which can be guaranteed with 3D printing techniques.

• Klíčová slova
• 3D printing, foot deformity, scanning,
• Publikační typ
• časopisecké články MeSH

A necessity to distinguish between the influence of powder shape and size (particle size distribution) is especially demanding for highly filled metal powder feedstocks employed in additive manufacturing and powder injection molding. As their processability is evaluated through rheological behavior, the study focuses on the effect of powder size/shape on a wall slip, which is a typical phenomenon determining flow performance of these materials. Water and gas atomized 17-4PH stainless steel powders with D 50 of about 3 and 20 μm are admixed into a binder containing low-density polyethylene, ethylene vinyl acetate, and paraffin wax. Mooney analysis to intercept the slip velocity of 55 vol. % filled compounds reveals that wall slip effect appears to vary significantly with size and shape of metal powders-round shaped and large particles are the most prone to the wall slip. However, the evaluation is affected by the type of the flow streams resulting from the geometry of the dies-conical dies reduce the slip up to 60% in case of fine and round particles.

• Klíčová slova
• die geometry, highly filled feedstock, material extrusion, particle size, powder injection molding, powder shape, wall slip,
• Publikační typ
• časopisecké články MeSH

Metal three-dimensional (3D) printing technology brings several benefits to the field of high-pressure die casting of aluminum, which enhances its development. The associated conformal cooling application is already commonly used where there is a need to improve the quality of castings, increase tool life, or reduce the production cycle. However, will this technology withstand the production of a large part (∼270 × 270 × 200 mm), which will be used directly in the serial production of engine blocks? This article describes a slider with a conformal cooling case study, which was redesigned and manufactured using the laser powder bed fusion (L-PBF) method. After the slider was put into serial production of 1.0 TSI three-cylinder engine blocks, this tool was thoroughly monitored based on the temperature field by comparing the results of a simulation in SW ProCAST with reality, and furthermore examining the influence of the tool on the quality of castings. There was also an evaluation of repairs performed on the tool in the ŠKODA AUTO tool shop and the foundry. These data were compared with a serial tool. Finally, the costs to produce the slider in conventional and 3D-printed variants are compared with an outline of other possible steps for optimizing these costs. The study results show that relatively large parts can be printed and used in serial production even today. It was also confirmed that conformal cooling influenced improving tool life, and the number of repairs in ŠKODA AUTO production also decreased.

• Klíčová slova
• conformal cooling, die and mold, high-pressure die casting, laser powder bed fusion, metal additive manufacturing,
• Publikační typ
• časopisecké články MeSH