Role In Hydrolysis Research Articles

Unveiling the hidden diversity and functional role of Chloroflexota in full-scale wastewater treatment plants through genome-centric analyses

Abstract The phylum Chloroflexota has been found to exhibit high abundance in the microbial communities from wastewater treatment plants (WWTPs) in both aerobic and anaerobic systems. However, its metabolic role has not been fully explored due to the lack of cultured isolates. To address this gap, we use publicly available metagenome datasets from both activated sludge (AS) and methanogenic (MET) full-scale wastewater treatment reactors to assembled genomes. Using this strategy, 264 dereplicated, medium- and high-quality metagenome-assembled genomes (MAGs) classified within Chloroflexota were obtained. Taxonomic classification revealed that AS and MET reactors harbored distinct Chloroflexota families. Nonetheless, the majority of the annotated MAGs (166 MAGs with >85% completeness and < 5% contamination) shared most of the metabolic potential features, including the ability to degrade simple sugars and complex polysaccharides, fatty acids and amino acids, as well as perform fermentation of different products. While Chloroflexota MAGs from MET reactors showed the potential for strict fermentation, MAGs from AS harbored the potential for facultatively aerobic metabolism. Metabolic reconstruction of Chloroflexota members from AS unveiled their versatile metabolism and suggested a primary role in hydrolysis, carbon removal and involvement in nitrogen cycling, thus establishing them as fundamental components of the ecosystem. Microbial reference genomes are essential resources for understanding the potential functional role of uncultured organisms in WWTPs. Our study provides a comprehensive genome catalog of Chloroflexota for future analyses aimed at elucidating their role in these ecosystems.

A Role of Cellulose Binding Module of the Thermophilic Endoglucanase TbCel12A

Endoglucanases are enzymes that play an important role in hydrolysis of lignocellulose by attacking glycosidic linkages in cellulose fibers and other glucans. The cellulose binding module (CBM) is responsible for binding the enzyme to the substrate. However, CBMs in certain enzymes interfere with substrate hydrolysis resulting in moderate or low activity. In a previous study, the processive endoglucanase TbCel12A including its CBM had low activity towards carboxymethyl cellulose (CMC). To assess the effect of the CBM, the catalytic domain of TbCel12A was produced without the CBM. The TbCel12A catalytic domain without the CBM hydrolyzed CMC 23 times more rapidly, while the pH and temperature optima and thermotolerance remained unchanged compared to full-length TbCel12A. Therefore, TbCel12A does not require the CBM for CMC hydrolysis and its application may be improved without it.

Flexibility and Function of Distal Substrate-Binding Tryptophans in the Blue Mussel β-Mannanase MeMan5A and Their Role in Hydrolysis and Transglycosylation

β-Mannanases hydrolyze β-mannans, important components of plant and microalgae cell walls. Retaining β-mannanases can also catalyze transglycosylation, forming new β-mannosidic bonds that are applicable for synthesis. This study focused on the blue mussel (Mytilus edulis) GH5_10 β-mannanase MeMan5A, which contains two semi-conserved tryptophans (W240 and W281) in the distal subsite +2 of its active site cleft. Variants of MeMan5A were generated by replacing one or both tryptophans with alanines. The substitutions reduced the enzyme’s catalytic efficiency (kcat/Km using galactomannan) by three-fold (W281A), five-fold (W240A), or 20-fold (W240A/W281A). Productive binding modes were analyzed by 18O labeling of hydrolysis products and mass spectrometry. Results show that the substitution of both tryptophans was required to shift away from the dominant binding mode of mannopentaose (spanning subsites −3 to +2), suggesting that both tryptophans contribute to glycan binding. NMR spectroscopy and molecular dynamics simulations were conducted to analyze protein flexibility and glycan binding. We suggest that W240 is rigid and contributes to +2 subsite mannosyl specificity, while W281 is flexible, which enables stacking interactions in the +2 subsite by loop movement to facilitate binding. The substitutions significantly reduced or eliminated transglycosylation with saccharides as glycosyl acceptors but had no significant effect on reactions with alcohols.

Human Carboxylesterase 1A Plays a Predominant Role in Hydrolysis of the Anti-Dyslipidemia Agent Fenofibrate in Humans.

Fenofibrate, a marketed peroxisome proliferator-activated receptor-α (PPARα) agonist, has been widely used for treating severe hypertriglyceridemia and mixed dyslipidemia. As a canonical prodrug, fenofibrate can be rapidly hydrolyzed to release the active metabolite (fenofibric acid) in vivo, but the crucial enzyme(s) responsible for fenofibrate hydrolysis and the related hydrolytic kinetics have not been well-investigated. This study aimed to assign the key organs and crucial enzymes involved in fenofibrate hydrolysis in humans, as well as reveal the impact of fenofibrate hydrolysis on its non-PPAR-mediated biologic activities. Our results demonstrated that fenofibrate could be rapidly hydrolyzed in the preparations from both human liver and lung to release fenofibric acid. Reaction phenotyping assays coupling with chemical inhibition assays showed that human carboxylesterase 1A (hCES1A) played a predominant role in fenofibrate hydrolysis in human liver and lung, while human carboxylesterase 2A (hCES2A) and human monoacylglycerol esterase (hMAGL) contributed to a very lesser extent. Kinetic analyses showed that fenofibrate could be rapidly hydrolyzed by hCES1A in human liver preparations, while the inherent clearance of hCES1A-catalyzed fenofibrate hydrolysis is much higher (>200-fold) than than that of hCES2A or hMAGL. Biologic assays demonstrated that both fenofibrate and fenofibric acid showed very closed Nrf2 agonist effects, but fenofibrate hydrolysis strongly weakens its inhibitory effects against both hCES2A and hNtoum. Collectively, our findings reveal that the liver is the major organ and hCES1A is the predominant enzyme-catalyzing fenofibrate hydrolysis in humans, while fenofibrate hydrolysis significantly reduces inhibitory effects of fenofibrate against serine hydrolases. SIGNIFICANCE STATEMENT: Fenofibrate can be completely converted to fenofibric acid in humans and subsequently exert its pharmacological effects, but the hydrolytic pathways of fenofibrate in humans have not been well-investigated. This study reported that the liver was the predominant organ and human carboxylesterase 1A was the crucial enzyme involved in fenofibrate hydrolysis in humans.

Achieving efficient methane production from protein-rich organic waste in anaerobic digestion: Using conductive materials or regulating inoculum-to-substrate ratios?

The contribution of inoculum-to-substrate ratios (ISRs) and conductive materials (CMs) on the productivity of anaerobic digestion (AD) remains unclear, particularly for protein-rich organic waste. This study investigated whether the addition of CMs, i.e., biochar and iron powder, can overcome the limitations imposed by varying ISRs for the AD of protein as the sole substrate. Results indicate the ISR plays a decisive role in hydrolysis, acidification, and methanogenesis for protein conversion, irrespective of CMs addition. Methane production increased stepwise as the ISR escalated to 3:1. The addition of CMs provided limited improvement, and iron powder even inhibited methanogenesis at a low ISR. Bacterial community variations were contingent on the ISR, while iron powder supplementation significantly elevates the proportion of hydrogenotrophic methanogen. This study demonstrates that the addition of CMs could affect methanogenic efficiency but can not overcome the limitation of ISRs for the AD of protein.

Silane Coatings for Corrosion and Microbiologically Influenced Corrosion Resistance of Mild Steel: A Review.

Mild steel continues to be the most extensively used construction material in several industries and constructions. However, corrosion of mild steel in aggressive environments is a major concern. Under the tremendously increasing demand for improving the coatings strategies because of the environmental concerns due to some of the traditional coatings, silane pre-treatments have been emerging as one of the effective solutions, among other strategies. Different approaches, such as adding particles of metal oxide (such as SiO2, ZrO2, Al2O3, TiO2 and CeO2), incorporating plant extracts and impregnating 2D materials into the coatings, have been employed for durable corrosion resistance, including for mitigating enhanced corrosion due to the presence of bacteria. This review discusses the critical mechanistic features of silane coatings such as the role of hydrolysis and condensation in the bonding of silanes with metal surfaces. The factors that influence the performance of the silane coatings for corrosion resistance of mild steel are discussed. In particular, this review provides insight into silane coatings for mitigating microbiologically influenced corrosion (MIC) of mild steel.

Semi-continuous anaerobic digestion of the organic fraction of municipal solid waste: digester performance and microbial population dynamics

Anaerobic digestion is an attractive approach for the management of organic fraction of municipal solid waste (OFMSW) and for the recovery of energy from this waste. A semi-continuous digestion of OFMSW was conducted on stirred reactor under mesophilic condition. Three organic loading rates (OLRs) were tested throughout the anaerobic process (1.36, 2.5 and 3.5 g VS/L.d). The achieved results showed that biogas yield decreased with the increase of OLR. Thus, the highest average of biogas yield, 0.51 L/g VS, was recorded at low OLR (OLR1:1.36 gVS/L.d). Likewise, optimal VS removal, pH, VFA and alkalinity were obtained at OLR1 compared to the other tested OLR. During the anaerobic digestion, the dynamic of microbial community was also assessed by the high-throughput sequencing technology. Firmicutes, Bacteroidetes and Proteobacteria were the dominant phyla during the digestion process. This could be related to their roles in hydrolysis of different OFMSW compounds, as well as in acidogenesis and acetogenesis steps. Regarding Archaeal population, the relative abundance of the Methanoculleus genus declined, while Methanosarcina increased over time. This indicated that acetotrophic and hydrogenotrophic pathways were used for methane production.

Effect of Moisture on Polymer Deconstruction in HCl Gas Hydrolysis of Wood

The HCl gas system previously used to produce cellulose nanocrystals was applied on Scots pine wood, aiming at a controlled deconstruction of its macrostructure while understanding the effect on its microstructure. The HCl gas treatments resulted in a well-preserved cellular structure of the wood. Differences in wood initial moisture content (iMC) prior to HCl gas treatment played a key role in hydrolysis rather than the studied range of exposure time to the acidic gas. Higher iMCs were correlated with a higher degradation of hemicellulose, while crystalline cellulose microfibrils were not largely affected by the treatments. Remarkably, the hydrogen–deuterium exchange technique showed an increase in accessible OH group concentration at higher iMCs, despite the additional loss in hemicelluloses. Unrelated to changes in the accessible OH group concentration, the HCl gas treatment reduced the concentration of absorbed D2O molecules.

Kinetics and mechanism of sequential ring methyl C-H activation in cyclopentadienyl rhodium(III) complexes.

We have studied activation of the methyl C-H bonds in the cyclopentadienyl ligands of half-sandwich Rh(III) complexes [η5-CpXRh(N,N')Cl]+ by observing the dependence of sequential H/D exchange on variations in CpX = Cp* (complexes 1 and 2), Me4PhCp (CpXPh, 3) or Me4PhPhCp (CpXPhPh, 4), and chelated ligand N,N' (bpy, 1; phen, 2-4). H/D exchange was fastest in d4-MeOD (t1/2 = 10 min, 37 °C, complex 1), no H/D exchange was observed in DMSO/D2O, and d4-MeOD enhanced the rate in CD3CN. The proposed Rh(I)-fulvene intermediate was trapped by [4 + 2] Diels-Alder reactions with conjugated dienes and characterized. The Rh(I) oxidation state was confirmed by X-ray photoelectron spectroscopy (XPS). Influence of solvent on the mechanisms of activation and Diels-Alder adduct formation was modelled using DFT calculations with the CAM-B3LYP functional and CEP-31 g basis set, and influence on the reaction profile of the dimiine ligand and phenyl substituent using the larger qzvp basis set. The Rh(III)-OH intemediate is stabilised by H-bonding with methanol and a Cp* CH3 hydrogen. The Rh(I)(Me4fulvene) species, stabilised by interaction of methanol with a coordinated water, again by two H-bonds H2O-HOMe (1.49 Å) and fulvene CH2 (1.94 Å), arises from synchronous transfer of the methanol OH proton to a Rh(III)-OH ligand and Cp* methyl hydrogen to the methanol oxygen. Additionally, the observed trend in catalytic activity for complexes 1-4 was reproduced by DFT calculations. These complexes form a novel class of catalytic molecular motors with a tunable rate of operation that can be stalled in a given state. They provide a basis for elucidation of the effects of ligand design on the contributions of electronic, rotational and vibrational energies to each step in the reaction pathway at the atomic level, consideration of which will enhance the design principles for the next generation of molecular machines.

CLINICAL AND PATHOGENESIS OVERVIEW OF MYROIDES SPP. INFECTION

The genus Myroides classified as Flavobacterium odoratum comprises non- motile ,aerobic, yellow –pigmented .They are non hemolytic on blood agar while it show a good growth on MacConkey and nutrient agar with salt tolerant up to 5% of NaCl also non fermented gram-negative rods .Members of the genus are cause many infectious disease in severely immunocompromised patient includes endocarditics urinary tract infection ,cuteneous infection ,and ventriculitis that behave as low grade opportunistic pathogens and widely distributed in the environment specially in water. Myroides spp. should be attribute among pathogens in hospitalized ,catheterized patients with wide resistant to antibiotic specially Minocycline is useful treating UTI caused by the genus . 16S ribosomal RNA(rRNA) sequencing can identified accurately the species level .In this reviews which description of dangerous infection duo to by Myroides in an immunocompromise host ,moreover reviewed the antibiotic resistance and pathogenesis. Many species of the genus Myroides were primarily taking from intestine of human. The genus Myroides includes five species : M. odoratus. , M. pelagicus, M. profundi , M. odoratimimus and M. marinus , by susceptibility to desferrioxamine . M. profundi has produced a protease Myroilysin ,with characterized as playing a role in hydrolysis of collagen through collagenase production and elastinolytic activity displaying by metalloprotease and playing important role with collagenase for hydrolysis of collagen that proven virulence of genus.

The role of hydrolysis in biological effects of molybdenum cluster with DMSO ligands.

Biological applications of octahedral molybdenum cluster complexes are complicated by their hydrolytic instability, since hydrolysis leads to irreversible changes in the structure and properties of these compounds. On the other hand, if such changes are thoroughly investigated and understood, the hydrolysis process can become an important tool for regulating specific biological effects of the clusters. In this work, we demonstrate how the luminescence and biological properties (cellular uptake, cytotoxicity in the dark and photodynamic effect) of highly unstable cluster complex [{Mo6I8}(DMSO)6](NO3)4 change along with the degree of hydrolysis. Particularly, cluster solution preliminarily aged in water demonstrated lower dark and higher photoinduced cytotoxicity and higher cellular uptake in comparison with fresh solution.

High Expression of Legumain in Pediatric B-Cell Precursor Acute Lymphoblastic Leukemia Is Associated with Central Nervous System Involvement

Background: Central nervous system (CNS) leukemia is a negative prognostic factor in pediatric acute lymphoblastic leukemia (ALL), and up to 30-40% of relapses involve the CNS. However, little is known about the biological mechanisms that lead to dissemination and survival of leukemic blasts in the CNS. This in part reflects that previous research has employed rather crude cytospin preparation methods for classification of CNS involvement. Using a highly sensitive flow-cytometry based-approach, we previously identified leukemic blasts in the cerebrospinal fluid at diagnosis in approximately 30% of ALL patients, whereas conventional cytospin microscopy only was positive in 10% of patients (Levinsen et al 2016 Pediatr Blood Cancer ).Methods: To explore biological properties of leukemic cells that migrate into the CNS, we compared gene expression profiles of leukemic cells from 26 patients with CNS involvement and 138 patients without detectable blasts in their cerebrospinal fluid. CNS status was determined by flow cytometry and/or cytospin examination. Gene expression profiles were obtained by microarray analysis (Human Gene 1.0 ST GeneChip array, Affymetrix). Microarray results were validated by real-time quantitative PCR.Results: A hierarchical cluster analysis of all 164 ALL samples in our study based on all top 10% most varying genes clearly subgrouped patients according to their B-cell precursor (BCP) versus T-cell immunophenotype. We therefore performed analyses separately for BCP and T-cell ALL patients. For patients with BCP ALL, more genes were differently expressed between patients with and without CNS involvement when patients were classified according to flow cytometry compared to conventional cytospin (68 probe sets, uncorrected P <0.05, versus 9 probe sets, uncorrected P <0.05). Noteworthy, the cysteine endopeptidase legumain (LGMN ) demonstrated significantly higher expression in BCP ALL with CNS involvement determined by flow cytometry, but no significant difference was observed when CNS involvement was assessed by cytospin. PCR confirmed the upregulation of LGMN in BCP ALL, when comparing flow-positive samples with flow-negative samples (P =0.006) and cytospin-negative samples (P =0.01). In multivariate analysis, LGMN expression levels above the median were associated with an odds ratio of 9.1 for CNS involvement as determined by flow cytometry (95% CI, 1.26 to 65.50; P =0.03).Conclusion: These data support that flow cytometry identifies CNS involvement better than conventional cytospin preparation, which is critical for a deeper understanding for the biology of CNS leukemia. Legumainplays a role in hydrolysis of proteins and small molecules and could potentially be involved in infiltration of the CNS by leukemic cells. Consequently, targeting legumain could be a novel strategy for combatting CNS leukemia. DisclosuresNo relevant conflicts of interest to declare.

Biological properties and roles of a Trichinella spiralis inorganic pyrophosphatase in molting and developmental process of intestinal larval stages

Inorganic pyrophosphatase (PPase) participates in energy cycle and plays a vital role in hydrolysis of inorganic pyrophosphate (PPi) into inorganic phosphate (Pi). The aim of this study was to investigate the biological properties of a Trichinella spiralis PPase (TsPPase) and its role in larval molting and developmental process. The predicted TsPPase consisted of 367 amino acids with a molecular mass of 41.48 kDa and a pI of 5.76. Amino acid sequence alignment and phylogenetic analysis showed that the TsPPase gene encodes a functional family I soluble PPase with the same characteristics as prokaryotic, plant and animal/fungal soluble PPase. The rTsPPase was expressed and purified, it has the activity to catalyze the hydrolysis of PPi to Pi, and the activity was dependent on Mg2+, pH and temperature. The enzymatic activity of rTsPPase was significantly inhibited after its metal binding sites mutation. TsPPase was transcribed and expressed in all T. spiralis phases, especially in muscle larvae (ML) and intestinal infective larvae (IIL). Immunofluorescence assay (IFA) revealed that TsPPase was mainly located in cuticle and stichosome. When the ML and IIL were treated with TsPPase-specific siRNA-279, TsPPase expression and enzymatic activity were obviously reduced, the larval molting and development were also impeded. Intestinal IIL as well as AW burden, IIL molting rates from mice infected with siRNA-treated ML were obviously suppressed. The results indicated that rTsPPase possesses the enzymatic activity of native inorganic pyrophosphatase, and TsPPase plays an important role in development and molting process of intestinal T. spiralis larval stages.

Influence of zinc oxide nanoparticles on anaerobic digestion of waste activated sludge and microbial communities.

The influence of long-term exposure of zinc oxide nanoparticles (ZnO NPs) to waste activated sludge on anaerobic digestion and microbial communities was studied. The exposure concentrations were 0, 30, 60, 90, 120, and 150 mg g−1-volatile suspended solids (VSS) (dry). ZnO NPs inhibit the degradation of macromolecular organic matter and the reduction of VSS in waste activated sludge during anaerobic digestion. Only slight effects on the activities of protease, cellulase, acetated kinase, and coenzyme F420 were found at ZnO-NP concentrations of less than 30 mg g−1-VSS, whereas the activities of these three enzymes were adversely affected in a dose-dependent manner when the ZnO NP concentrations were increased from 30 mg g−1-VSS to 150 mg g−1-VSS. High-throughput sequencing analysis revealed that ZnO NPs had an adverse influence on the archaeal community diversity but increased the bacterial community diversity to some extent. High-throughput sequencing analysis also revealed that ZnO NPs resulted in different shift trends in the archaeal and bacteria community structure at phylum, class, and genus levels. ZnO NPs have negative impacts on the Euryarchaeota community, which plays a significant role as methanogens in the anaerobic digestion. In addition, ZnO NPs could increase the relative abundance of Clostridia and Bacteroidia, playing an important role in hydrolysis during the anaerobic digestion.

Design, synthesis and biological evaluation of indanone-chalcone hybrids as potent and selective hCES2A inhibitors.

Human carboxylesterase 2 (hCES2A), one of the major serine hydrolases distributed in the small intestine, plays a crucial role in hydrolysis of ester-bearing drugs. Accumulating evidence has indicated that hCES2A inhibitor therapy can modulate the pharmacokinetic and toxicological profiles of some important hCES2A-substrate drugs, such as the anticancer agent CPT-11. Herein, a series of indanone-chalcone hybrids are designed and synthesized to find potent and highly selective hCES2A inhibitors. Inhibition assays demonstrated that most indanone-chalcone hybrids displayed strong to moderate hCES2A inhibition activities. Structure-hCES2A inhibition activity relationship studies showed that introduction of a hydroxyl at the C4' site and introduction of an N-alkyl group at the C6 site were beneficial for hCES2A inhibition. Particularly, B7 (an N-alkylated 1-indanone-chalcone hybrid) exhibited the most potent inhibition on hCES2A and excellent specificity (this agent could not inhibit other human esterases including hCES1A and butyrylcholinesterase). Inhibition kinetic analyses demonstrated that B7 potently inhibited hCES2A-mediated FD hydrolysis in a mixed inhibition manner, with a calculated Ki value of 0.068μM. Furthermore, B7 was capable of inhibiting intracellular hCES2A in living cells and displayed good metabolic stability. Collectively, our findings show that indanone-chalcone hybrids are good choices for the development of hCES2A inhibitors, while B7 is a promising candidate for the development of novel anti-diarrhea agents to ameliorate irinotecan-induced intestinal toxicity.

Discovery and characterization of tannase genes in plants: roles in hydrolysis of tannins.

Plant tannins, including condensed tannins (CTs) and hydrolyzable tannins (HTs), are widely distributed in the plant kingdom. To date, tannase (TA) - is a type of tannin acyl-hydrolase hydrolyzing HTs, CT monomer gallates and depsides - has been reported in microbes only. Whether plants express TA remains unknown. Herein, we report plant TA genes. A native Camellia sinensis TA (CsTA) is identified from leaves. Six TAs are cloned from tea, strawberry (Fragaria×ananassa, Fa) and four other crops. Biochemical analysis shows that the native CsTA and six recombinant TAs hydrolyze tannin compounds, depsides and phenolic glycosides. Transcriptional and metabolic analyses reveal that the expression of CsTA is oppositely associated with the accumulation of galloylated catechins. Moreover, the transient overexpression and RNA interference of FaTA are positively associated with the accumulation of ellagitannins in strawberry fruit. Phylogenetic analysis across different kingdoms shows that 29 plant TA homologs are clustered as a plant-specific TA clade in class I carboxylesterases. Further analysis across the angiosperms reveals that these TA genes are dispersed in tannin-rich plants, which share a single phylogenetic origin c. 120 million yr ago. Plant TA is discovered for the first time in the plant kingdom and is shown to be valuable to improve tannin compositions in plants.

Investigating into the molecular interactions of flavonoids targeting NS2B-NS3 protease from ZIKA virus through in-silico approaches

Zika virus (ZIKV), belongs to the flavivirus genus and Flaviviridae family that associated with serious diseased conditions like microcephaly and other neurological disorders (Guillan–Barré syndrome). As there is no vaccine or therapies available against ZIKV to date. Hence, it is an unmet need to find potential drug candidates and target sites against Zika virus infection. NS2B-NS3 protease making an attractive target for therapeutic intervention in ZIKV infections because of its critical role in hydrolysis of a single polyprotein encoded by Zika virus. Recently, there are some experimental evidence about the flavonoids as Zika virus NS2B-NS3 protease inhibitors. However, molecular interaction between protease complex and inhibitors at atomic levels has not been explored. Here, we have taken the experimentally validated thirty-eight flavonoids inhibitors against NS2B-NS3 protease to examine the molecular interaction using molecular docking and molecular dynamics simulations. We found out few flavonoids such as EGCG and its two derivatives, isoquercetin, rutin and sanggenon O showing interaction with catalytic triad (His51, Asp75, and Ser135) of the active site of NS2B-NS3 protease and found to be stable throughout the simulation. Therefore it is evident that interaction with the catalytic triad playing a vital role in the inhibition of the enzyme activity as a result inhibition of the virus propagation. However these compounds can be explored further for understanding the mechanism of action of these compounds targeting NS2B-NS3 protease for inhibition of Zika virus.

Strigolactone perception:What is the role of hydrolysis?

Strigolactone perception:What is the role of hydrolysis?

Unveiling the biotic and abiotic mechanisms of solids concentration affecting hydrolysis and acidogenesis during sludge thermophilic anaerobic fermentation

Abstract The biotic factors (bacterial abundance, enzyme activity) and abiotic factors (sludge disintegration, organics solubilization) in sludge thermophilic fermentation at various solids concentrations were investigated to clarify the effects on hydrolysis and volatile fatty acids (VFAs) production. The results show that with the solids concentration increasing from volatile suspended solids (VSS) 5–40 g/L, the abiotic effects of particles disintegration played a leading role in hydrolysis, resulting in the decrease of hydrolysis rate constant (kH) (from 0.1181 to 0.0623) and solids degradation ratio (from 55.19% to 36.63%), while the biotic effects dominated above VSS20, reinforcing bacterial abundance and hydrolase activity. The maximal soluble chemical oxygen demand (SCOD) production yield positively correlated with VSS, and the value for VSS40 (436.52 mg/g VSSfed) increased by 50% over VSS5 (298.21 mg/g VSSfed). Additionally, the solubilization and conversion of carbohydrates were respectively enhanced by 2.6-folds and 13.4-folds at higher VSS, while for proteins they almost decreased by 50%. The VFAs production was comprehensively affected by the carbon substrate, ammonium and acid-forming enzyme, with the highest accumulation yield (232.17 mg CODVFA/g VSSfed) for VSS30; meanwhile, a higher percentage of acetate and n-butyrate but lower of propionate were achieved with VSS increasing.

Investigation of the chiral recognition ability of human carboxylesterase 1 using indomethacin esters.

Human carboxylesterase 1 (hCES1) is an enzyme that plays an important role in hydrolysis of pharmaceuticals in the human liver. In this study, elucidation of the chiral recognition ability of hCES1 was attempted using indomethacin esters in which various chiral alcohols were introduced. Indomethacin was condensed with various chiral alcohols to synthesize indomethacin esters. The synthesized esters were hydrolyzed with a human liver microsome (HLM) solution and a human intestine microsome (HIM) solution. High hydrolytic rate and high stereoselectivity were confirmed in the hydrolysis reaction in the HLM solution but not in the HIM solution, and these indomethacin esters were thought to be hydrolyzed by hCES1. Next, these indomethacin esters were hydrolyzed in recombinant hCES1 solution and the hydrolysis rates of the esters were calculated. The stereoselectivity confirmed in HLM solution was also confirmed in the hCES1 solution. In the hydrolysis reaction of esters in which a phenyl group is bonded next to the ester, the Vmax value of the (R) form was 10 times larger than that of the (S) form.