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9 pages, 4 figures, 2 tables, supplementary data https://doi.org/10.1016/j.etap.2020.103561, Carboxylesterases (CEs) are key enzymes which catalyse the hydrolysis reactions of multiple xenobiotics and endogenous ester moieties. Given their growing interest in the context of marine pollution and biomonitoring, this study focused on the in vitro sensitivity of marine invertebrate CEs to some pesticides, pharmaceuticals, personal care products and plastic additives to assess their potential interaction on this enzymatic system and its suitability as biomarkers. Three bivalves, one gastropod and two crustaceans were used and CEs were quantified following current protocols set for mammalian models. Four substrates were screened for CEs determination and to test their adequacy in the hepatic fraction measures of the selected invertebrates. Two commercial recombinant human isoforms (hCE1 and hCE2) were also included for methodological validation. Among the invertebrates, mussels were revealed as the most sensitive to xenobiotic exposures while gastropods were the least as well as with particular substrate-specific preferences. Among chemicals of environmental concern, the plastic additive tetrabromobisphenol A displayed the highest CE-inhibitory capacity in all species. Since plastic additives easily breakdown from the polymer and may accumulate and metabolise in marine biota, their interaction with the CE key metabolic/detoxification processes may have consequences in invertebrate’s physiology, affect bioaccumulation and therefore trophic web transfer and, ultimately, human health as shellfish consumers, This work was financed by Spanish Ministry of Economy, Industry and Competitivity (ref CGL2016-76332-R MINECO/FEDER, UE), With the funding support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S), of the Spanish Research Agency (AEI)

Background The frequent usage of broad-spectrum insecticides like cypermethrin in agriculture activities could lead to the development of resistance in insects like Amsacta albistriga. Objectives The present study was conducted to understand the toxicity of cypermethrin with and without a combination of three enzyme inhibitors (PBO, DEM, and TPP) on A. albistriga using a topical bioassay. Methodology The in vitro and in vivo studies were conducted to understand the effect of three enzyme inhibitors such as piperonyl butoxide (PBO) and diethyl maleate (DEM) and triphenyl phosphate along with six different concentration of cypermethrin on the activities of acetylcholinesterase (AChE), esterase (EST), glutathione S-transferase (GST), and mixed-function oxidase (MFO) of A. albistriga. Results Bioassay shows elevated LC50 for cypermethrin (63.32 ppm) whereas in a combination of PBO cypermethrin LC50 values were reduced into 12.039 ppm followed by TPP combination as 13.234 ppm. In vitro and in vivo inhibition analysis shows AChE inhibition by PBO P < 0.01, esterase inhibition by PBO, and TPP were less; GST inhibition by DEM was observed at P < 0.001. Native PAGE results revealed that the in vitro AChE isoenzyme inhibition could be possible by PBO synergism with cypermethrin in A. albistriga. Conclusions The overall results conclude that the PBO and TPP enzyme inhibitors could be fine synergist molecules when it mixed with cypermethrin insecticide to control and manage the insecticide-resistant Amsacta albistriga in the field.

Summary Soy protein isolates (SPI) films were evaluated as carriers of citral and limonene, and their physical and antifungal properties were evaluated. The presence of antimicrobials in SPI films resulted in changes on colour without affecting the transparency. Films with citral added were more extensible; however, a reduction in tensile strength and elastic modulus was observed in films with limonene added. Aroma compounds addition in films induced a slight increase in water vapour properties in relation to discontinuity of network proteins evidenced by SEM. Besides, FTIR spectra evidenced a partial alteration of SPI secondary structure. Citral was less retained than limonene. The increase in limonene release with high relative humidity was explained by increase in protein chain mobility. SPI films enriched with limonene exhibited strong antifungal activity against the postharvest decay pathogen Penicillium italicum under storage conditions.

4 pages, 2 figures, 1 table, Tetrabromobisphenol A (TBBPA), a brominated flame retardant used in synthetic polymers and electronics, is present in the aquatic environment and recent evidence suggests it can be potentially biomagnified in the marine ecosystem. However, the toxicity of TBBPA in the marine biota has not been investigated in detail. In this study we aimed to understand the role of carboxylesterases (CEs) in xenobiotic metabolism under the exposure of marine organisms to a chemical of environmental concern, TBBPA. Specifically, we tested for in vitro inhibition of CE activity in a range of marine organisms covering different ecological niches, from species from low (mussels and copepods), medium (sardines and anchovies) and high trophic levels (tuna). The results revealed that the highest inhibition of CE activity to 100 μM TBBPA was recorded in mussels (66.5% inhibition) and tunids (36.3-76.4%), whereas copepods and small pelagic fish showed comparatively lower effects (respectively, 30% and 36.5-55.6%). Our results suggest that CE-mediated detoxification and physiological processes could be compromised in TBBPA-exposed organisms and could ultimately affect humans as many of them are market species, The funding was from AIMCOST (CGL2016-76332-R MINECO/FEDER, UE) and PELWEB (CTM2017-88939-R) projects of the Spanish Ministry of Economy and Competitiveness. JN was funded by the Spanish National Program Ramón y Cajal (RYC-2015-17809), With the funding support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S), of the Spanish Research Agency (AEI)

Gençer, Nahit (Balikesir Author), Human serum paraoxonase 1 (PON1; EC 3.1.8.1) is a high-density lipoprotein associated, calcium-dependent enzyme that hydrolyses aromatic esters, organophosphates and lactones and can protect the low-density lipoprotein against oxidation. In this study, in vitro effect of some hydroxy and dihydroxy ionic coumarin derivatives (1-20) on purified PON1 activity was investigated. Among these compounds, derivatives 11-20 are water soluble. In investigated compounds, compounds 6 and 13 were found the most active (IC50 = 35 and 34 mu M) for PON1, respectively. The present study has demonstrated that PON1 activity is very highly sensitive to studied coumarin derivatives., Balikesir University Research project 2014/54

Statine-based inhibitors of Plasmepsin II (PLMII) coupled with Primaquine have been designed using the `double-drug' approach. The IC50 values for PLMII inhibition ranged from 0.59 to 400 nM and the best IC50 value for inhibition of Plasmodium falciparum growth in vitro was 0.4 μM, which represent a remarkable improvement compared to other statine-based PLMII inhibitors.

Çakır, Ümit (Balıkesir author), Inhibitory effects of some synthesized dihydroxycoumarin compounds on purified G6PD were investigated. For this purpose, initially human erythrocyte G6PD was purified 7069-fold in a yield of 33.6% by using ammonium sulfate precipitation and affinity chromatography which includes 2',5'-ADP Sepharose 4B. The purified enzyme showed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Enzyme activity was determined spectrophotometrically according to Beutler method at 340 nm. 6,7-Dihydroxy-3-(2-methylphenyl)-2H-chromen-2-one (OPC), 6,7-dihydroxy-3-(3-methylphenyl)-2H-chromen-2-one (MPC) and 6,7-dihydroxy-3-(4-methylphenyl)-2H-chromen-2-one (PPC) were used as dihydroxycoumarin compounds. This study has demonstrated that G6PD activity is very highly sensitive to study coumarin derivatives., Scientific Research Council of Balikesir University

Protein phosphatase 5 (PP5) is a unique member of serine/threonine phosphatases which has been recognized in regulation of diverse cellular processes. A cDNA fragment encoding PP5 (EcPP5) was cloned and characterized from the cantharidin-producing blister beetle, E. chinensis. EcPP5 contains an open reading frame of 1500 bp that encodes a protein of 56.89 kDa. The deduced amino acid sequence shares 88% and 68% identities to the PP5 of Tribolium castaneum and humans, respectively. Analysis of the primary sequence shows that EcPP5 has three TPR (tetratricopeptide repeat) motifs at its N-terminal region and contains a highly conserved C-terminal catalytic domain. RT-PCR reveals that EcPP5 is expressed in all developmental stages and in different tissues. The recombinant EcPP5 (rEcPP5) was produced in Escherichia coli and purified to homogeneity. The purified protein exhibited phosphatase activity towards pNPP (p-nitrophenyl phosphate) and phosphopeptides, and its activity can be enhanced by arachidonic acid. In vitro inhibition study revealed that protein phosphatase inhibitors, okadaic acid, cantharidin, norcantharidin and endothall, inhibited its activity. Further, protein phosphatase activity of total soluble protein extract from E. chinensis adults could be impeded by these inhibitors suggesting there might be some mechanism to protect this beetle from being damaged by its self-produced cantharidin.

WOS: 000311310400015, PubMed: 22971832, Human serum paraoxonase 1 (PON1; EC 3.1.8.1) is a high-density lipoprotein associated, calcium-dependent enzyme that hydrolyses aromatic esters, organophosphates and lactones and can protect the low-density lipoprotein against oxidation. In this study, in vitro inhibition effect of some dihydroxy coumarin compounds namely 6,7-dihydroxy-3-(2-methylphenyl)-2H-chromen-2-one (A), 6,7-dihydroxy-3-(3-methylphenyl)-2H-chromen-2-one (B) and 6,7-dihydroxy-3-(4-methylphenyl)-2H-chromen-2-one (C) on purified PON1 were investigated by using paraoxon as a substrate. PON1 was purified using two-step procedures, namely ammonium sulphate precipitation and Sepharose-4B-l-tyrosine-1-naphthylamine hydrophobic interaction chromatography. The purified enzyme had a specific activity of 11.76 U/mg. The dihydroxy coumarin derivatives of A and B compounds inhibited PON1 enzyme activity in a noncompetitive inhibition manner with K (i) of 0.0080 +/- 0.256 and 0.0003 +/- 0.018 mM values, respectively. C compound exerted an uncompetitive inhibition of PON1 enzyme activity with K (i) of 0.0010 +/- 0.173 mM. Moreover, dihydroxy coumarin derivatives of A, B and C compounds were effective inhibitors on purified human serum PON1 activity with IC50 of 0.012, 0.022 and 0.003 mM values, respectively. IC50 value of unsubstituted 6,7 dihydroxy coumarin was found as 0.178 mM. The present study has demonstrated that PON1 activity is very highly sensitive to studied coumarin derivatives.

Non-carboxylic acid containing bioisosteres of (5-arylidene-2,4-dioxothiazolidin-3-yl)acetic acids, which are active as aldose reductase (ALR2) inhibitors, were designed by replacing the carboxylic group with the trifluoromethyl ketone moiety. The in vitro evaluation of the ALR2 inhibitory effects of these trifluoromethyl substituted derivatives led to the identification of two inhibitors effective at low micromolar doses. It was further confirmed that a carboxylic chain on N-3 of the thiazolidinedione scaffold is a determining requisite to obtain the highest efficacy levels; however, it is not essential for the interaction with the target enzyme and it can be replaced by different polar groups, thus obtaining less ionised or unionised inhibitors.