PFOA accumulation in the leaves of basil (Ocimum basilicum L.) and its effects on plant growth, oxidative status, and photosynthetic performance.

BACKGROUND: Perfluoroalkyl substances (PFASs) are emerging contaminants of increasing concern due to their presence in the environment, with potential impacts on ecosystems and human health. These substances are considered "forever chemicals" due to their recalcitrance to degradation, and their accumulation in living organisms can lead to varying levels of toxicity based on the compound and species analysed. Furthermore, concerns have been raised about the possible transfer of PFASs to humans through the consumption of edible parts of food plants. In this regard, to evaluate the potential toxic effects and the accumulation of perfluorooctanoic acid (PFOA) in edible plants, a pot experiment in greenhouse using three-week-old basil (Ocimum basilicum L.) plants was performed adding PFOA to growth substrate to reach 0.1, 1, and 10 mg Kg- 1 dw. RESULTS: After three weeks of cultivation, plants grown in PFOA-added substrate accumulated PFOA at different levels, but did not display significant differences from the control group in terms of biomass production, lipid peroxidation levels (TBARS), content of α-tocopherol and activity of ascorbate peroxidase (APX), catalase (CAT) and guaiacol peroxidase (POX) in the leaves. A reduction of total phenolic content (TPC) was instead observed in relation to the increase of PFOA content in the substrate. Furthermore, chlorophyll content and photochemical reflectance index (PRI) did not change in plants exposed to PFAS in comparison to control ones. Chlorophyll fluorescence analysis revealed an initial, rapid photoprotective mechanism triggered by PFOA exposure, with no impact on other parameters (Fv/Fm, ΦPSII and qP). Higher activity of glutathione S-transferase (GST) in plants treated with 1 and 10 mg Kg- 1 PFOA dw (30 and 50% to control, respectively) paralleled the accumulation of PFOA in the leaves of plants exposed to different PFOA concentration in the substrate (51.8 and 413.9 ng g- 1 dw, respectively). CONCLUSION: Despite of the absorption and accumulation of discrete amount of PFOA in the basil plants, the analysed parameters at biometric, physiological and biochemical level in the leaves did not reveal any damage effect, possibly due to the activation of a detoxification pathway likely involving GST.

Shared and more specific genetic determinants and pathways underlying yeast tolerance to acetic, butyric, and octanoic acids.

BACKGROUND: The improvement of yeast tolerance to acetic, butyric, and octanoic acids is an important step for the implementation of economically and technologically sustainable bioprocesses for the bioconversion of renewable biomass resources and wastes. To guide genome engineering of promising yeast cell factories toward highly robust superior strains, it is instrumental to identify molecular targets and understand the mechanisms underlying tolerance to those monocarboxylic fatty acids. A chemogenomic analysis was performed, complemented with physiological studies, to unveil genetic tolerance determinants in the model yeast and cell factory Saccharomyces cerevisiae exposed to equivalent moderate inhibitory concentrations of acetic, butyric, or octanoic acids. RESULTS: Results indicate the existence of multiple shared genetic determinants and pathways underlying tolerance to these short- and medium-chain fatty acids, such as vacuolar acidification, intracellular trafficking, autophagy, and protein synthesis. The number of tolerance genes identified increased with the linear chain length and the datasets for butyric and octanoic acids include the highest number of genes in common suggesting the existence of more similar toxicity and tolerance mechanisms. Results of this analysis, at the systems level, point to a more marked deleterious effect of an equivalent inhibitory concentration of the more lipophilic octanoic acid, followed by butyric acid, on the cell envelope and on cellular membranes function and lipid remodeling. The importance of mitochondrial genome maintenance and functional mitochondria to obtain ATP for energy-dependent detoxification processes also emerged from this chemogenomic analysis, especially for octanoic acid. CONCLUSIONS: This study provides new biological knowledge of interest to gain further mechanistic insights into toxicity and tolerance to linear-chain monocarboxylic acids of increasing liposolubility and reports the first lists of tolerance genes, at the genome scale, for butyric and octanoic acids. These genes and biological functions are potential targets for synthetic biology approaches applied to promising yeast cell factories, toward more robust superior strains, a highly desirable phenotype to increase the economic viability of bioprocesses based on mixtures of volatiles/medium-chain fatty acids derived from low-cost biodegradable substrates or lignocellulose hydrolysates.

Co-Removal of Perfluorooctanoic Acid and Nitrate from Water by Coupling Pd Catalysis with Enzymatic Biotransformation.

PFAS (poly- and per-fluorinated alkyl substances) represent a large family of recalcitrant organic compounds that are widely used and pose serious threats to human and ecosystem health. Here, palladium (Pd0)-catalyzed defluorination and microbiological mineralization were combined in a denitrifying H2-based membrane biofilm reactor to remove co-occurring perfluorooctanoic acid (PFOA) and nitrate. The combined process, i.e., Pd-biofilm, enabled continuous removal of ∼4 mmol/L nitrate and ∼1 mg/L PFOA, with 81% defluorination of PFOA. Metagenome analysis identified bacteria likely responsible for biodegradation of partially defluorinated PFOA: Dechloromonas sp. CZR5, Kaistella koreensis, Ochrobacterum anthropic, and Azospira sp. I13. High-performance liquid chromatography-quadrupole time-of-flight mass spectrometry and metagenome analyses revealed that the presence of nitrate promoted microbiological oxidation of partially defluorinated PFOA. Taken together, the results point to PFOA-oxidation pathways that began with PFOA adsorption to Pd0, which enabled catalytic generation of partially or fully defluorinated fatty acids and stepwise oxidation and defluorination by the bacteria. This study documents how combining catalysis and microbiological transformation enables the simultaneous removal of PFOA and nitrate.

Caprylic acid attenuates amyloid-ß proteotoxicity by supplying energy via ß-oxidation in an Alzheimer's disease model of the nematode Caenorhabditis elegans.

Computer-based analysis of motility was used as a measure of amyloid-ß (Aß) proteotoxicity in the transgenic strain GMC101, expressing human Aß1-42 in body wall muscle cells. Aß-aggregation was quantified to relate the effects of caprylic acid (CA) to the amount of the proteotoxic protein. Gene knockdowns were induced through RNA-interference (RNAi). Moreover, the estimation of adenosine triphosphate (ATP) levels, the mitochondrial membrane potential (MMP) and oxygen consumption served the evaluation of mitochondrial function. CA improved the motility of GMC101 nematodes and reduced Aß aggregation. Whereas RNAi for orthologues encoding key enzymes for α-lipoic acid and ketone bodies synthesis did not affect motility stimulation by CA, knockdown of orthologues involved in ß-oxidation of fatty acids diminished its effects. The efficient energy gain by application of CA was finally proven by the increase of ATP levels in association with increased oxygen consumption and MMP. In conclusion, CA attenuates Aß proteotoxicity by supplying energy via FAO. Since especially glucose oxidation is disturbed in Alzheimer´s disease, CA could potentially serve as an alternative energy fuel.

Exploiting the Thermotolerance of Clostridium Strain M1NH for Efficient Caproic Acid Fermentation from Ethanol and Acetic Acid.

A novel thermotolerant caproic acid-producing bacterial strain, Clostridium M1NH, was successfully isolated from sewage sludge. Ethanol and acetic acid at a molar ratio of 4:1 proved to be the optimal substrates, yielding a maximum caproic acid production of 3.5 g/L. Clostridium M1NH exhibited remarkable tolerance to high concentrations of ethanol (up to 5% v/v), acetic acid (up to 5% w/v), and caproic acid (up to 2% w/v). The strain also demonstrated a wide pH tolerance range (pH 5.5-7.5) and an elevated temperature optimum between 35 and 40 °C. Phylogenetic analysis based on 16S rRNA gene sequences revealed that Clostridium M1NH shares a 98% similarity with Clostridium luticellarii DSM 29923 T. The robustness of strain M1NH and its efficient caproic acid production from low-cost substrates highlight its potential for sustainable bio-based chemical production. The maximum caproic acid yield achieved by Clostridium M1NH was 1.6-fold higher than that reported for C. kluyveri under similar fermentation conditions. This study opens new avenues for valorizing waste streams and advancing a circular economy model in the chemical industry.

Uptake of perfluoroalkyl substances PFOS and PFOA by free-floating hydrophytes Pistia stratiotes L. and Eichhornia crassipes (Mart.) Solms.

The phytoremediation potential of floating aquatic plants to accumulate and remove two common PFAS from contaminated water was investigated. Free-floating hydrophytes Eichhornia crassipes and Pistia stratiotes were grown in water spiked with 0.5, 1, or 2 ppm perfluorooctanoic acid (PFOA) or perfluorooctanesulfonic acid (PFOS) for seven days. Both species were able to accumulate PFOA and PFOS in this time frame, with translocation factors (TF) ranging from 0.13 to 0.57 for P. stratiotes and 0.18 to 0.45 for E. stratiotes, respectively. E. crassipes accumulated a greater amount of PFOA and PFOS than P. stratiotes, with 178.9 ug PFOA and 308.5 ug PFOS removed by E. crassipes and 98.9 ug PFOA and 137.8 ug PFOS removed by P. stratiotes at the highest concentrations. Root tissue contained a higher concentration of PFOA and PFOS than shoot tissue in both species, and the concentration of PFOS was generally significantly higher than PFOA in both E. crassipes and P. stratiotes, with concentrations of 15.39 and 27.32 ppb PFOA and 17.41 and 80.62 ppb PFOS in shoots and roots of P. stratiotes and 12.59 and 37.37 ppb PFOA and 39.92 and 83.40 ppb PFOS in shoots and roots of E. crassipes, respectively. Both species may be candidates for further phytoremediation studies in aquatic ecosystems.

This study investigates the feasibility of using wetland plants for the phytoremediation of PFAS. Prior published studies examine various plant interactions with PFAS but do not evaluate remediation potential of P. stratiotes.

Identification of the acyltransferase that octanoylates ghrelin, an appetite-stimulating peptide hormone.

Ghrelin is a 28 amino acid, appetite-stimulating peptide hormone secreted by the food-deprived stomach. Serine-3 of ghrelin is acylated with an eight-carbon fatty acid, octanoate, which is required for its endocrine actions. Here, we identify GOAT (Ghrelin O-Acyltransferase), a polytopic membrane-bound enzyme that attaches octanoate to serine-3 of ghrelin. Analysis of the mouse genome revealed that GOAT belongs to a family of 16 hydrophobic membrane-bound acyltransferases that includes Porcupine, which attaches long-chain fatty acids to Wnt proteins. GOAT is the only member of this family that octanoylates ghrelin when coexpressed in cultured endocrine cell lines with prepro-ghrelin. GOAT activity requires catalytic asparagine and histidine residues that are conserved in this family. Consistent with its function, GOAT mRNA is largely restricted to stomach and intestine, the major ghrelin-secreting tissues. Identification of GOAT will facilitate the search for inhibitors that reduce appetite and diminish obesity in humans.

In Vitro and In Vivo Metabolism of a Novel Antimitochondrial Cancer Metabolism Agent, CPI-613, in Rat and Human.

CPI-613, an inhibitor of pyruvate dehydrogenase (PDH) and α-ketoglutarate dehydrogenase (KGDH) enzymes, is currently in development for the treatment of pancreatic cancer, acute myeloid leukemia, and other cancers. CPI-613 is an analog of lipoic acid, an essential cofactor for both PDH and KGDH. Metabolism and mass balance studies were conducted in rats after intravenous administration of [14C]-CPI-613. CPI-613 was eliminated via oxidative metabolism followed by excretion of the metabolites in feces (59%) and urine (22%). ß-Oxidation was the major pathway of elimination for CPI-613. The most abundant circulating components in rat plasma were those derived from ß-oxidation. In human hepatocytes, CPI-613 mainly underwent ß-oxidation (M1), sulfur oxidation (M2), and glucuronidation (M3). The Michaelis-Menten kinetics (Vmax and Km) of the metabolism of CPI-613 to these three metabolites predicted the fraction metabolized leading to the formation of M1, M2, and M3 to be 38%, 6%, and 56%, respectively. In humans, after intravenous administration of CPI-613, major circulating species in plasma were the parent and the ß-oxidation derived products. Thus, CPI-613 metabolites profiles in rat and human plasma were qualitatively similar. ß-Oxidation characteristics and excretion patterns of CPI-613 are discussed in comparison with those reported for its endogenous counterpart, lipoic acid. SIGNIFICANCE STATEMENT: This work highlights the clearance mechanism of CPI-613 via ß-oxidation, species differences in their ability to carry out ß-oxidation, and subsequent elimination routes. Structural limitations for completion of terminal cycle of ß-oxidation is discussed against the backdrop of its endogenous counterpart lipoic acid.

A unique, newly discovered four-member protein family involved in extracellular fatty acid binding in Yarrowia lipolytica.

BACKGROUND: Yarrowia lipolytica, a nonconventional oleaginous yeast species, has attracted attention due to its high lipid degradation and accumulation capacities. Y. lipolytica is used as a chassis for the production of usual and unusual lipids and lipid derivatives. While the genes involved in the intracellular transport and activation of fatty acids in different cellular compartments have been characterized, no genes involved in fatty acid transport from the extracellular medium into the cell have been identified thus far. In this study, we identified secreted proteins involved in extracellular fatty acid binding. RESULTS: Recent analysis of the Y. lipolytica secretome led to the identification of a multigene family that encodes four secreted proteins, preliminarily named UP1 to UP4. These proteins were efficiently overexpressed individually in wild-type and multideletant strain (Q4: Δup1Δup2Δup3Δup4) backgrounds. Phenotypic analysis demonstrated the involvement of these proteins in the binding of extracellular fatty acids. Additionally, gene deletion and overexpression prevented and promoted sensitivity to octanoic acid (C8) toxicity, respectively. The results suggested binding is dependent on aliphatic chain length and fatty acid concentration. 3D structure modeling supports the proteins' role in fatty acid assimilation at the molecular level. CONCLUSIONS: We discovered a family of extracellular-fatty-acid-binding proteins in Y. lipolytica and have proposed to name its members eFbp1 to eFbp4. The exact mode of eFbps action remains to be deciphered individually and synergistically; nevertheless, it is expected that the proteins will have applications in lipid biotechnology, such as improving fatty acid production and/or bioconversion.

Perfluorooctanoic acid exposure in vivo perturbs mitochondrial metabolic during oocyte maturation.

Perfluorooctanoic acid (PFOA), a member of a group of polyfluorinated and perfluorinated alkyl substances (PFAS), is associated with adverse pregnancy outcomes in mammals. However, the effects of in vivo exposure to PFOA on the female reproductive system and the underlying mechanisms remain unclear. In our study, we constructed a mouse model to investigate whether low-dose PFOA (1 mg/kg/day) or high-dose PFOA (5 mg/kg/day) affect meiosis maturation of oocytes and the potential mechanisms that may be associated with oocyte maturation disorder. Our results indicate that low-dose and high-dose PFOA can lead to impaired oocyte maturation, which is manifested by decreased rate of embryonic foam rupture and first polar body extrusion. Moreover, PFOA exposure harmed the mitochondrial metabolic, resulting in low levels of ATP contents, high reactive oxygen species, aberrant mitochondrial membrane potential. In addition, the proportion of DNA damage marker γ-H2AX was also significantly increased in PFOA exposure oocytes. These changes lead to abnormal arrangements of the spindle and chromosomes during oocyte maturation. In conclusion, our results for the first time illustrated that exposure to PFOA in vivo in female mice impaired the meiosis maturation of oocytes, which provided a basis for studying the mechanism of PFOA reproductive toxicity in female mammals.

Ligand binding to human prostaglandin E receptor EP4 at the lipid-bilayer interface.

Prostaglandin E receptor EP4, a G-protein-coupled receptor, is involved in disorders such as cancer and autoimmune disease. Here, we report the crystal structure of human EP4 in complex with its antagonist ONO-AE3-208 and an inhibitory antibody at 3.2 Å resolution. The structure reveals that the extracellular surface is occluded by the extracellular loops and that the antagonist lies at the interface with the lipid bilayer, proximal to the highly conserved Arg316 residue in the seventh transmembrane domain. Functional and docking studies demonstrate that the natural agonist PGE2 binds in a similar manner. This structural information also provides insight into the ligand entry pathway from the membrane bilayer to the EP4 binding pocket. Furthermore, the structure reveals that the antibody allosterically affects the ligand binding of EP4. These results should facilitate the design of new therapeutic drugs targeting both orthosteric and allosteric sites in this receptor family.

Transcriptomic response of Saccharomyces cerevisiae to octanoic acid production.

The medium-chain fatty acid octanoic acid is an important platform compound widely used in industry. The microbial production from sugars in Saccharomyces cerevisiae is a promising alternative to current non-sustainable production methods, however, titers need to be further increased. To achieve this, it is essential to have in-depth knowledge about the cell physiology during octanoic acid production. To this end, we collected the first RNA-Seq data of an octanoic acid producer strain at three time points during fermentation. The strain produced higher levels of octanoic acid and increased levels of fatty acids of other chain lengths (C6-C18) but showed decreased growth compared to the reference. Furthermore, we show that the here analyzed transcriptomic response to internally produced octanoic acid is notably distinct from a wild type's response to externally supplied octanoic acid as reported in previous publications. By comparing the transcriptomic response of different sampling times, we identified several genes that we subsequently overexpressed and knocked out, respectively. Hereby we identified RPL40B, to date unknown to play a role in fatty acid biosynthesis or medium-chain fatty acid tolerance. Overexpression of RPL40B led to an increase in octanoic acid titers by 40%.

Production of octanoic acid in Saccharomyces cerevisiae: Investigation of new precursor supply engineering strategies and intrinsic limitations.

The eight-carbon fatty acid octanoic acid (OA) is an important platform chemical and precursor of many industrially relevant products. Its microbial biosynthesis is regarded as a promising alternative to current unsustainable production methods. In Saccharomyces cerevisiae, the production of OA had been previously achieved by rational engineering of the fatty acid synthase. For the supply of the precursor molecule acetyl-CoA and of the redox cofactor NADPH, the native pyruvate dehydrogenase bypass had been harnessed, or the cells had been additionally provided with a pathway involving a heterologous ATP-citrate lyase. Here, we redirected the flux of glucose towards the oxidative branch of the pentose phosphate pathway and overexpressed a heterologous phosphoketolase/phosphotransacetylase shunt to improve the supply of NADPH and acetyl-CoA in a strain background with abolished OA degradation. We show that these modifications lead to an increased yield of OA during the consumption of glucose by more than 60% compared to the parental strain. Furthermore, we investigated different genetic engineering targets to identify potential factors that limit the OA production in yeast. Toxicity assays performed with the engineered strains suggest that the inhibitory effects of OA on cell growth likely impose an upper limit to attainable OA yields.

Myocardial Substrate Oxidation and Tricarboxylic Acid Cycle Intermediates During Hypothermic Machine Perfusion.

BACKGROUND: The optimal substrate for hypothermic machine perfusion preservation of donor hearts is unknown. Fatty acids, acetate, and ketones are preferred substrates of the heart during normothermic perfusion, but cannot replete the tricarboxylic acid (TCA) cycle directly. Propionate, an anaplerotic substrate, can replenish TCA cycle intermediates and may affect cardiac metabolism. The purpose of this study was to determine myocardial substrate preferences during hypothermic machine perfusion and to assess if an anaplerotic substrate was required to maintain the TCA cycle intermediate pool in perfused hearts. METHODS: Groups of rat hearts were perfused with carbon-13 (13C)-labeled substrates (acetate, ß-hydroxybutyrate, octanoate, with and without propionate) at low and high concentrations. TCA cycle intermediate concentrations, substrate selection, and TCA cycle flux were determined by gas chromatography/mass spectroscopy and 13C magnetic resonance spectroscopy. RESULTS: Acetate and octanoate were preferentially oxidized, whereas ß-hydroxybutyrate was a minor substrate. TCA cycle intermediate concentrations except fumarate were higher in substrate-containing perfusion groups compared with either the no-substrate perfusion group or the no-ischemia control group. CONCLUSIONS: The presence of an exogenous, oxidizable substrate is required to support metabolism in the cold perfused heart. An anaplerotic substrate is not essential to maintain the TCA cycle intermediate pool and support oxidative metabolism under these conditions.

Caprylic acid ameliorates rotenone induced inflammation and oxidative stress in the gut-brain axis in Zebrafish.

BACKGROUND: Dysfunction of the gastrointestinal tract (GIT) is one of the most common non-motor symptom of Parkinson's Disease (PD). Pathological processes causing PD were suggested to initiate in the enteric nervous system (ENS) and proceed to the central nervous system (CNS). There are studies showing that low-carbohydrate ketogenic diets can improve motor symptoms of PD. Caprylic acid (C8) is the principal fatty acid component of the medium-chain triglycerides in the ketogenic diets. In this study, we aimed to evaluate the effects of caprylic acid, in neurotoxin exposed zebrafish focusing on the relationship between intestinal and brain oxidative stress and inflammation. METHODS: Adult zebrafish were exposed to rotenone (5 µg/L) (R group) and caprylic acid (20 and 60 mg/mL) (L + HDCA and R + HDCA groups) for 30 days. At the end of 30 days locomotor activities were determined. Levels of lipid peroxidation (LPO), nitric oxide, glutathione and superoxide dismutase and glutathione S-transferase activities were determined by spectrophotometric methods and gene expressions of tnf⍺, il1, il6, il21, ifnÉ£ and bdnf were evaluated by RT-PCR in the brain and intestinal tissues of zebrafish. RESULTS: Caprylic acid ameliorated LPO, NO, SOD and the expressions of tnf⍺, il1, il6, il21, ifnÉ£ and bdnf in brain and intestines. Locomotor activities were only ameliorated in high dose R + HDCA group. CONCLUSIONS: Caprylic acid ameliorated the neurotoxin-induced oxidative stress and inflammation both in the brain and intestines and enhanced locomotor activity in zebrafish.

Caprylic acid enhances hydroxyhexylitaconic acid production in Aspergillus niger S17-5.

AIM: Aspergillus niger S17-5 produces two alkylitaconic acids, 9-hydroxyhexylitaconic acid (9-HHIA) and 10-hydroxyhexylitaconic acid (10-HHIA), which have cytotoxic and polymer building block properties. In this study, we characterized the production of 9-HHIA and 10-HHIA by addition of their expected precursor, caprylic acid, to a culture of A. niger S17-5, and demonstrated batch fermentation of 9-HHIA and 10-HHIA in a jar fermenter with DO-stat. METHODS AND RESULTS: Production titres of 9-HHIA and 10-HHIA from 3% glucose in a flask after 25 days cultivation were 0·35 and 1·01 g l-1 respectively. Addition of 0·22 g l-1 of caprylic acid to a suspension of resting cells of A. niger S17-5 led to 32% enhancement of total 9-HHIA and 10-HHIA production compared to no addition. No enhancement of the production of 9-HHIA or 10-HHIA by the addition of oxaloacetic acid was observed. Addition of caprylic acid to the culture at mid-growth phase was more suitable for 9-HHIA and 10-HHIA production due to less cell growth inhibition by caprylic acid. DO-stat batch fermentation with 3% glucose and 14·4 g l-1 of caprylic acid in a 1·5 l jar fermenter resulted in the production titres of 9-HHIA and 10-HHIA being 0·48 and 1·54 g l-1 respectively after 10 days of cultivation. CONCLUSIONS: Addition of caprylic acid to the culture of A. niger S17-5 enhances 9-HHIA and 10-HHIA production. SIGNIFICANCE AND IMPACT OF THE STUDY: These results suggest that 9-HHIA and 10-HHIA are synthesized with octanoyl-CoA derived from caprylic acid, and that the supply of octanoyl-CoA is a rate-limiting step in 9-HHIA and 10-HHIA production. To the best of our knowledge, this is the first report regarding the fermentation of naturally occurring itaconic acid derivatives in a jar fermenter.

Pharmacokinetics, metabolism and off-target effects in the rat of 8-[(1H- benzotriazol-1-yl)amino]octanoic acid, a selective inhibitor of human cytochrome P450 4Z1: ß-oxidation as a potential augmenting pathway for inhibition.

8-[(1H-1,2,3-benzotriazol-1-yl)amino]octanoic acid (8-BOA) was recently identified as a selective and potent mechanism-based inactivator (MBI) of breast cancer-associated CYP4Z1 and exhibited favourable inhibitory activity in vitro, thus meriting in vivo characterization.The pharmacokinetics and metabolism of 8-BOA in rats was examined after a single IV bolus dose of 10 mg/kg. A biphasic time-concentration profile resulted in relatively low clearance and a prolonged elimination half-life.The major circulating metabolites identified in plasma were products of ß-oxidation; congeners losing two and four methylene groups accounted for >50% of metabolites by peak area. The -(CH2)2 product was characterized previously as a CYP4Z1 MBI and so represents an active metabolite that may contribute to the desired pharmacological effect.Ex vivo analysis of total CYP content in rat liver and kidney microsomes showed that off-target CYP inactivation was minimal; liver microsomal probe substrate metabolism also demonstrated low off-target inactivation. Standard clinical chemistries provided no indication of acute toxicity.In silico simulations using the free concentration of 8-BOA in plasma suggested that the in vivo dose used here may effectively inactivate CYP4Z1 in a xenografted tumour.

Biosyntheses of geranic acid and citronellic acid from monoterpene alcohols by Saccharomyces cerevisiae.

Geraniol is one of the important aromatic ingredients in alcoholic beverages. Bioconversions of geraniol to other terpenoids and genes involved in the oxidation of geraniol were investigated. Geranic acid and citronellic acid were detected in yeast culture, where geraniol or nerol was added. Addition of citral, a mixture of geranial and neral, resulted in the production of geranic acid and citronellic acid, whereas the addition of citral or citronellal resulted in the production of citronellic acid, suggesting that citronellic acid might be produced through the conversion of citral to citronellal followed by the oxidation of citronellal. Consumption of geraniol and production of geranic acid, citronellic acid, and citronellol were affected in adh1Δ, adh3Δ, adh4Δ, and sfa1Δ yeast strains, which possess single deletion of a gene encoding alcohol dehydrogenase. This is the first report of the bioconversion of monoterpene alcohols, geraniol and nerol, to geranic acid and citronellic acid in yeast culture.

PFAS Degradation in Ultrapure and Groundwater Using Non-Thermal Plasma.

Perfluoroalkyl substances (PFAS) represent one of the most recalcitrant class of compounds of emerging concern and their removal from water is a challenging goal. In this study, we investigated the removal efficiency of three selected PFAS from water, namely, perfluorooctanoic acid (PFOA), perfluorohexanoic acid (PFHxA) and pefluorooctanesulfonic acid (PFOS) using a custom-built non-thermal plasma generator. A modified full factorial design (with 2 levels, 3 variables and the central point in which both quadratic terms and interactions between couple of variables were considered) was used to investigate the effect of plasma discharge frequency, distance between the electrodes and water conductivity on treatment efficiency. Then, the plasma treatment running on optimized conditions was used to degrade PFAS at ppb level both individually and in mixture, in ultrapure and groundwater matrices. PFOS 1 ppb exhibited the best degradation reaching complete removal after 30 min of treatment in both water matrices (first order rate constant 0.107 min-1 in ultrapure water and 0.0633 min-1 in groundwater), while the degradation rate of PFOA and PFHxA was slower of around 65% and 83%, respectively. During plasma treatment, the production of reactive species in the liquid phase (hydroxyl radical, hydrogen peroxide) and in the gas phase (ozone, NOx) was investigated. Particular attention was dedicated to the nitrogen balance in solution where, following to NOx hydrolysis, total nitrogen (TN) was accumulated at the rate of up to 40 mgN L-1 h-1.

Improving the catalytic efficiency and substrate affinity of a novel esterase from marine Klebsiella aerogenes by random and site-directed mutation.

A novel esterase (EstKa) from marine Klebsiella aerogenes was characterized with hydrolytic activity against p-nitrophenyl caprylate (pNPC, C8) under optimum conditions (50 °C and pH 8.5). After two rounds of mutagenesis, two highly potential mutants (I6E9 and L7B11) were obtained with prominent activity, substrate affinity and thermostability. I6E9 (L90Q/P96T) and L7B11 (A37S/Q100L/S133G/R138C/Q156R) were 1.56- and 1.65-fold higher than EstKa in relative catalytic efficiency. The influence of each amino acid on enzyme activity was explored by site-directed mutation. The mutants Pro96Thr and Gln156Arg showed 1.29- and 1.48-fold increase in catalytic efficiency (Kcat/Km) and 54.4 and 36.2% decrease in substrate affinity (Km), respectively. The compound mutant Pro96Thr/Gln156Arg exhibited 68.9% decrease in Km and 1.41-fold increase in Kcat/Km relative to EstKa. Homology model structure analysis revealed that the replacement of Gln by hydrophilic Arg on the esterase surface improved the microenvironment stability and the activity. The replacement of Pro by Thr enabled the esterase enzyme to retain 90% relative activity after 3 h incubation at 45 °C. Structural analysis confirmed that the formation of a hydrogen bond leads to a notable increase of catalytic efficiency under high temperature conditions.