The use of ecological,fishing and environmental indicators in support of decision making in southern Benguela fisheries

Indicators have been recognised as a useful tool aiding the implementation of an ecosystem approach to fisheries in marine ecosystems. Studies, such as the IndiSeas project (www.indiseas.org), use a suite of indicators as a method of assessing the state and trends of several of the world's marine ecosystems. While it is well known that both fishing and climatic variability influence marine fisheries in the southern Benguela ecosystem there are currently few studies in support of fisheries management that make use of environmental indicators in order to include climatic impacts on marine fish populations. Trends in ecological, fishing and environmental indicators can be utilised in a way that allows an overall ecosystem trend to be determined, and can therefore be used to aid decision support within southern Benguela fisheries. In this study trends in indicators were determined using linear regressions across three time periods, Period 1: 1978–1993, Period 2: 1994–2003 and Period 3: 2004–2010. These time periods were selected based on the timing of regime shifts within the southern Benguela, including changes in upwelling, wind stress and temperature. Each ecological indicator received a score based on the direction and significance of the observed trend with respect to fishing. To account for the impacts of fishing and environmental drivers on ecological indicators, scores were adjusted by predetermined factors, depending on the extent and direction of trends in these indicators. Weightings were applied to correlated ecological indicators to account for their redundancy, and lessen their impact on overall ecosystem score. Mean weighted scores were then used to establish an overall ecosystem score for each time period. Ecosystem classification was determined as follows: 1–1.49 = improving, 1.5–2.49 = possibly improving, 2.5–3.49 = no improvement or deterioration, 3.5–4.49 = possible deterioration, 4.5–5 = deteriorating. The ecosystem was observed to neither deteriorate nor improve across Period 1 or 2 (mean weighted scores: 2.75 and 2.56 respectively), however, during Period 3 a possible improvement was observed (mean weighted score: 1.99). This study shows that the sequential analysis of suites of ecological, fishing and environmental indicators can be used in order to determine ecosystem trends, accounting for both the impacts of fishing and the environment on ecosystem components.     

Characterization of d-amino acid aminotransferase from Lactobacillus salivarius

We searched a UniProt database of lactic acid bacteria in an effort to identify d-amino acid metabolizing enzymes other than alanine racemase. We found a d-amino acid aminotransferase (d-AAT) homologous gene (UniProt ID: Q1WRM6) in the genome of Lactobacillus salivarius. The gene was then expressed in Escherichia coli, and its product exhibited transaminase activity between d-alanine and α-ketoglutarate. This is the first characterization of a d-AAT from a lactic acid bacterium. L. salivarius d-AAT is a homodimer that uses pyridoxal-5′-phosphate (PLP) as a cofactor; it contains 0.91 molecules of PLP per subunit. Maximum activity was seen at a temperature of 60 °C and a pH of 6.0. However, the enzyme lost no activity when incubated for 30 min at 30 °C and pH 5.5 to 9.5, and retained half its activity when incubated at pH 4.5 or 11.0 under the same conditions. Double reciprocal plots of the initial velocity and d-alanine concentrations in the presence of several fixed concentrations of α-ketoglutarate gave a series of parallel lines, which is consistent with a Ping-Pong mechanism. The K_m values for d-alanine and α-ketoglutarate were 1.05 and 3.78 mM, respectively. With this enzyme, d-allo-isoleucine exhibited greater relative activity than d-alanine as the amino donor, while α-ketobutylate, glyoxylate and indole-3-pyruvate were all more preferable amino acceptors than α-ketoglutarate. The substrate specificity of L. salivarius d-AAT thus differs greatly from those of the other d-AATs so far reported.     

Field and culture studies on the ecophysiology of the toxic dinoflagellate Alexandrium minutum (Halim) present in Greek coastal waters

In Greek coastal waters, the toxic dinoflagellate Alexandrium minutum (strain AJ879163) was detected for the first time in spring 2002. This species proliferated during spring–summer of 2002 and 2003 over a wide geographic range along the north-south Aegean Sea coastline, mostly at low concentrations (average: 10²–10³ cells L⁻¹) with one exception of higher abundance (average: 10⁵ cells L⁻¹). This study presents data on environmental (temperature, salinity, chl α, nutrients) and ecological (phytoplankton species composition, diversity, taxa dominance, community dissimilarities) parameters in the areas of A. minutum occurrence. A. minutum was isolated and grown in batch cultures used in a series of bioassay experiments for determination of its pigment composition by HPLC, half saturation constants (K_s) for nitrogen and phosphorus, and its response to different nitrogen to phosphorus (N:P) ratios.     

Mechanical performance of cervical intervertebral body fusion devices: A systematic analysis of data submitted to the Food and Drug Administration

Cervical intervertebral body fusion devices (IBFDs) are utilized to provide stability while fusion occurs in patients with cervical pathology. For a manufacturer to market a new cervical IBFD in the United States, substantial equivalence to a cervical IBFD previously cleared by FDA must be established through the 510(k) regulatory pathway. Mechanical performance data are typically provided as part of the 510(k) process for IBFDs. We reviewed all Traditional 510(k) submissions for cervical IBFDs deemed substantially equivalent and cleared for marketing from 2007 through 2014. To reduce sources of variability in test methods and results, analysis was restricted to cervical IBFD designs without integrated fixation, coatings, or expandable features. Mechanical testing reports were analyzed and results were aggregated for seven commonly performed tests (static and dynamic axial compression, compression-shear, and torsion testing per ASTM F2077, and subsidence testing per ASTM F2267), and percentile distributions of performance measurements were calculated. Eighty-three (83) submissions met the criteria for inclusion in this analysis. The median device yield strength was 10,117 N for static axial compression, 3680 N for static compression-shear, and 8.6 N m for static torsion. Median runout load was 2600 N for dynamic axial compression, 1400 N for dynamic compression-shear, and ±1.5 N m for dynamic torsion. In subsidence testing, median block stiffness (Kp) was 424 N/mm. The mechanical performance data presented here will aid in the development of future cervical IBFDs by providing a means for comparison for design verification purposes.     

Drinks containing anthocyanin-rich blackcurrant extract decrease postprandial blood glucose,insulin and incretin concentrations

Blackcurrants are rich in polyphenolic glycosides called anthocyanins, which may inhibit postprandial glycemia. The aim was to determine the dose-dependent effects of blackcurrant extract on postprandial glycemia. Men and postmenopausal women (14 M, 9 W, mean age 46 years, S.D.=14) were enrolled into a randomized, double-blind, crossover trial. Low sugar fruit drinks containing blackcurrant extract providing 150-mg (L-BE), 300-mg (M-BE) and 600-mg (H-BE) total anthocyanins or no blackcurrant extract (CON) were administered immediately before a high-carbohydrate meal. Plasma glucose, insulin and incretins (GIP and GLP-1) were measured 0–120 min, and plasma 8-isoprostane F_2α, together with arterial stiffness by digital volume pulse (DVP) was measured at 0 and 120 min. Early plasma glucose response was significantly reduced following H-BE (n=22), relative to CON, with a mean difference (95% CI) in area over baseline (AOB) 0-30 min of −0.34 mmol/l.h (−0.56, −0.11, P<.005); there were no differences between the intermediate doses and placebo. Plasma insulin concentrations (AOB 0–30 min) were similarly reduced. Plasma GIP concentrations (AOB 0–120 min) were significantly reduced following H-BE, with a mean difference of −46.6 ng/l.h (−66.7, −26.5, P<.0001) compared to CON. Plasma GLP-1 concentrations were reduced following H-BE at 90 min. There were no effects on 8-isoprostane F_2α or vascular function. Consumption of blackcurrant extract in amounts roughly equivalent to 100-g blackcurrants reduced postprandial glycemia, insulinemia and incretin secretion, which suggests that inclusion of blackcurrant polyphenols in foods may provide cardio-metabolic health benefits. This trial was registered at clinicaltrials.gov as NCT01706653.     

Three-dimensional data-tracking dynamic optimization simulations of human locomotion generated by direct collocation

The aim of this study was to perform full-body three-dimensional (3D) dynamic optimization simulations of human locomotion by driving a neuromusculoskeletal model toward in vivo measurements of body-segmental kinematics and ground reaction forces. Gait data were recorded from 5 healthy participants who walked at their preferred speeds and ran at 2 m/s. Participant-specific data-tracking dynamic optimization solutions were generated for one stride cycle using direct collocation in tandem with an OpenSim-MATLAB interface. The body was represented as a 12-segment, 21-degree-of-freedom skeleton actuated by 66 muscle-tendon units. Foot-ground interaction was simulated using six contact spheres under each foot. The dynamic optimization problem was to find the set of muscle excitations needed to reproduce 3D measurements of body-segmental motions and ground reaction forces while minimizing the time integral of muscle activations squared. Direct collocation took on average 2.7 ± 1.0 h and 2.2 ± 1.6 h of CPU time, respectively, to solve the optimization problems for walking and running. Model-computed kinematics and foot-ground forces were in good agreement with corresponding experimental data while the calculated muscle excitation patterns were consistent with measured EMG activity. The results demonstrate the feasibility of implementing direct collocation on a detailed neuromusculoskeletal model with foot-ground contact to accurately and efficiently generate 3D data-tracking dynamic optimization simulations of human locomotion. The proposed method offers a viable tool for creating feasible initial guesses needed to perform predictive simulations of movement using dynamic optimization theory. The source code for implementing the model and computational algorithm may be downloaded at http://simtk.org/home/datatracking.     

Airborne hyperspectral data predict Ellenberg indicator values for nutrient and moisture availability in dry grazed grasslands within a local agricultural landscape

Species-based ecological indices, such as Ellenberg indicators, reflect plant habitat preferences and can be used to describe local environment conditions. One disadvantage of using vegetation data as a substitute for environmental data is the fact that extensive floristic sampling can usually only be carried out at a plot scale within limited geographical areas. Remotely sensed data have the potential to provide information on fine-scale vegetation properties over large areas. In the present study, we examine whether airborne hyperspectral remote sensing can be used to predict Ellenberg nutrient (N) and moisture (M) values in plots in dry grazed grasslands within a local agricultural landscape in southern Sweden. We compare the prediction accuracy of three categories of model: (I) models based on predefined vegetation indices (VIs), (II) models based on waveband-selected VIs, and (III) models based on the full set of hyperspectral wavebands. We also identify the optimal combination of wavebands for the prediction of Ellenberg values. The floristic composition of 104 (4 m × 4 m grassland) plots on the Baltic island of Öland was surveyed in the field, and the vascular plant species recorded in the plots were assigned Ellenberg indicator values for N and M. A community-weighted mean value was calculated for N (mN) and M (mM) within each plot. Hyperspectral data were extracted from an 8 m × 8 m pixel window centred on each plot. The relationship between field-observed and predicted mean Ellenberg values was significant for all three categories of prediction models. The performance of the category II and III models was comparable, and they gave lower prediction errors and higher R² values than the category I models for both mN and mM. Visible and near-infrared wavebands were important for the prediction of both mN and mM, and shortwave infrared wavebands were also important for the prediction of mM. We conclude that airborne hyperspectral remote sensing can detect spectral differences in vegetation between grassland plots characterised by different mean Ellenberg N and M values, and that remote sensing technology can potentially be used to survey fine-scale variation in environmental conditions within a local agricultural landscape.     

Metal-dependent thermal stability of recombinant endo-mannanase (ManB-1601) belonging to family GH 26 from Bacillus sp. CFR1601

A GH 26 endo-mannanase from Bacillus sp. CFR1601 was purified to homogeneity (M_w ∼39 kDa, specific activity 10,461.5 ± 100 IU/mg). Endo-mannanase gene (manb-1601, 1083 bp, accession No. KM404299) was expressed in Escherichia coli BL21 (DE3) and showed typical fingerprints of α/β proteins in the far-UV CD. A high degree of conservation among amino acid residues involved in metal chelation (His-1, 23 and Glu-336) and internal repeats (122–152 and 181–212) was observed in endo-mannanases reported from various Bacillus sp. Thermal inactivation kinetics suggested that metal ions are quintessential for stabilization of ManB-1601 structure as holoenzyme (E_a 87.4 kcal/mol, ΔH 86.7 kcal/mol, ΔS 186.6 cal/k/mol) displayed better values of thermodynamic parameters compared to metal-depleted ManB-1601 (E_a 47 kcal/mol, ΔH 45.7 kcal/mol, ΔS 64.7 cal/k/mol). EDTA treatment of ManB-1601 not only lead to transitions in both secondary and tertiary structure but also promulgated the population of conformational state that unfolds at lower temperature. ManB-1601 followed a three-state process for thermal inactivation wherein loss of tertiary structure preceded the concurrent loss of secondary structure and activity.     

Single nucleotide polymorphism (SNP) in alpha-lactalbumin gene of Indian Jamunapari breed of Capra hircus

Alpha-lactalbumin is a major whey protein found in milk. It influences lactose synthesis by modifying the substrate specificity of galactosyl-transferase, is important to milk synthesis since lactose, an impermeable disaccharide, is the major osmole of milk. The present study was undertaken to detect polymorphism in the full coding region of alpha-lactalbumin at the genetic level and to explore allelic variability of this gene. Samples of Jamunapari breed of goat (n = 50) were included under the present study. Jamunapari is the highest milk producer among local Indian goat breeds. PCR-SSCP of all four exons of alpha-lactalbumin (ALA) revealed a total of 9 gel phenotypes of Jamunapari breed of goat. These were sequenced, analyzed and deposited in GenBank, NCBI (accession nos. EU573193–EU573195). Nucleotide and amino acid variations were searched within breeds of Indian goats and homology between caprine, ovine, bovine, bubaline and human. In the present study we describe for the first time two novel gene variants on the goat alpha-lactalbumin gene exon 4.     

Interfractional diaphragm changes during breath-holding in stereotactic body radiotherapy for liver cancer

Aim and background

IGRT based on bone matching may produce a large target positioning error in terms of the reproducibility of expiration breath-holding on SBRT for liver cancer. We evaluated the intrafractional and interfractional errors using the diaphragm position at the end of expiration by utilising Abches and analysed the factor of the interfractional error.

A putative proline iminopeptidase of Thermotoga maritima is a leucine aminopeptidese with lysine-p-nitroanilide hydrolyzing activity

A putative aminopeptidase P gene (TM0042, Swissport Q9WXP9, GeneBank AAD35136) of Thermotoga maritima was cloned and expressed in Escherichia coli BL21 (RIL). The enzyme was purified by the combination of ion exchange chromatography; Q-Sepharose and Mono-Q column. The purified recombinant T. maritima aminopeptidase P enzyme, gave a homogenous protein band with an apparent molecular weight of 40 kDa in SDS-PAGE analysis. The enzyme was purified 23-fold with the specific activity of 16.5 unit/mg with the final recovery of 22%. The enzyme was thermostable up to 90 °C for 30 min. An optimal activity was observed at 90 °C at pH 7.5. The purified enzyme was stable between pH 6.5 and 8 at 80 °C with the optimum of pH 7.5. Based on the amino acid sequence, the enzyme belongs to M 24B family of metalloenzymes. None of the divalent cations enhance the activity of the enzyme while Pb²⁺, Cu²⁺, Co²⁺, Cd²⁺, and Zn²⁺ were inhibitory to the enzyme activity. Divalent cation of Mg²⁺ showed 100% enzyme activity, to a lesser extent, Ca²⁺ and Mn²⁺ whereas strong inhibition of enzyme activity was observed with Zn²⁺ and Cd²⁺. The enzyme designated as putative aminopeptidase P was very low activity in hydrolyzing proline-p-nitroanilide. Kinetic studies on the purified enzyme confirmed that the enzyme is a leucine aminopeptidase. Enzyme also hydrolyzes lysine-p-nitroanilide with efficiency comparable to that of leucine-p-nitroanilide. This is the first report of leucine aminopeptidase with lysine-p-nitroanilide hydrolyzing activity, which belongs to the M 24B family of metalloenzymes.     

Substratum associations of benthic invertebrates in lowland and mountain streams

The preferences of aquatic invertebrate species for specific substrata at the river bottom have been subject of many studies. Several authors classified the substratum preferences of species or higher taxonomic units. Most of these compilations, however, are based on literature analyses and expert knowledge as opposed to the analysis of original data. To enhance our knowledge of invertebrate substratum preferences, we applied a ‘Multi-level pattern’ analysis based on almost 1000 substrate-specific invertebrate samples. The samples were taken in 18 streams in Germany, the Netherlands and Austria, comprising a total of 40 sampling sites and equally covering lowland and mountain streams. The main objectives of our analysis were (I) to derive substratum preferences of taxa in lowland and mountain streams, (II) to compare the preferences with existing data and (III) to compare species substratum associations between lowland and mountain streams. Of the 290 taxa analyzed, 188 were associated significantly to specific substrata. Twenty-five taxa in lowland streams and 51 taxa in mountain streams prefer one or two substratum types (of nine substratum types considered in total). In contrast, 112 species (mountain streams n = 84, lowland streams n = 28) are associated significantly with a broader range of substrata. We compared the classifications derived from our data analysis with those provided in the freshwaterecology.info database (www.freshwaterecology.info). Our results support the existing classifications of substratum preferences in most cases (70%). For 25 species, substratum preferences for both lowland and mountain streams were derived, many of them indicating different substratum associations in the two stream groups. As substratum preferences differed between closely related species, preferences should always be given at the species level as opposed to coarser taxonomic units.     

Shrub removal facilitates recovery of wetland species in a rewetted fen

Drainage and shrub expansion are the main threats to the biodiversity of fens and fen meadows, whereas rewetting and the removal of shrubby species are frequently applied restoration measures. We examine whether removal of shrubs enhances recovery of target species in a degraded fen subjected to moderate rewetting. The study was located in the drained fen Ca?owanie (central Poland), where remnants of open fen communities and willow-invaded fens exist in former turf-pits, surrounded by degraded meadows on dried peat. All these three habitat types were included in a monitoring grid, which covered an area of 2.2 ha. Within 55 quadrats of 20 m × 20 m we monitored occurrence of 52 species, i.e. two groups of target species (fen indicators and wet meadow indicators) and indicators of two failure scenarios (degraded fen indicators and eutrophic wetland indicators), during six years following shrub removal, rewetting and re-application of conservational mowing, using a 3-step ordinal abundance scale. NMDS ordination revealed a gradual convergence of shrub removal plots and reference plots. We noticed significant effects of year and habitat type on all indicator groups, but only fen indicators have shown a clear (increasing) trend within shrub removal plots. Degraded fen indicators (ruderal and opportunistic species) initially expanded on shrub removal plots, but this effect disappeared in the following years. We conclude that shrub removal enhances establishment of target species in a moderately drained and then rewetted fen and attribute this effect to lowered competition for light. However, given high costs of this method and long-lasting problems with shrub resprouts, we recommend applying shrub removal only to recently overgrown sites, which still retained high botanical diversity. Heavily degraded fen meadows did not react on the increase of moisture, which indicates that more advanced restoration measures, such as top soil removal are needed there.     

A novel single nucleotide polymorphism in the coding region of goat growth hormone receptor gene and its association with lactose content and somatic cell count in milk

A novel single nucleotide polymorphism—a C/T transition (RFPL-MspI) was found upon sequencing of a 439 bp DNA fragment, comprising whole exon 4 and parts of adjacent introns of the goat growth hormone receptor gene. This mutation was located at 8th nucleotide of exon 4 (position 94 according to GenBank Acc. No. AY739707). The nucleotide substitution has no effect on the amino acid sequence of the GHR protein. Within the cohort of 227 Polish dairy goats three genotypes were found: CC (frequency 0.96), CT (0.036), and TT (0.004). The frequency of C and T alleles was 0.978 and 0.022, respectively. It was shown that CC genotype goats had significantly higher lactose content and lower somatic cell count than those with the CT genotype. No association was found with the other milk production traits studied.     

Synthesis,antimicrobial, cytotoxic and E. coli DNA gyrase inhibitory activities of coumarinyl amino alcohols

Here we report the in vitro antimicrobial activity (minimum inhibitory concentration) of fourteen coumarinyl amino alcohols 2–16 against eight bacterial strains and two fungi. Among these compounds 4, 8, 12, 15 and 16 showed moderate to good microbial inhibition with MIC values varied from 6.25 to 25 μg/mL. The most promising compounds were also evaluated for their in vitro cytotoxic and E. coli DNA gyrase inhibitory activities along with the two 7-oxy-4-methyl coumarinyl amino alcohol derivatives 17 and 18, which were found to be the most potent in in vitro antimicrobial screening in our previous study. All the active compounds, including 17 and 18, were also docked into the E. coli DNA gyrase ATP binding site (PDB ID: 1KZN) to investigate their binding interactions. Of these compound 17 has shown maximum binding energy value of −6.13 kcal/mol.     

Do indicators of maternal iron status reflect placental iron status at delivery?

Indicators of maternal iron (Fe) status were studied in relation to placental Fe (Pl-Fe) status. Placental (Pl) and maternal (M) venous blood samples were obtained from primiparous women ($n=38$), with normal delivery at Paroissien Hospital, Argentina. Maternal hemoglobin (M Hb), soluble transferrin receptor (M sTfR) (ELISA) and serum ferritin (M S-Ft) were studied in relation to Pl-Fe, ferritin (Pl-Ft) and transferrin receptor (Pl-TfR). Pl-TfR was measured by dot blot assay, Pl-Ft and M S-Ft by immunoassay (IRMA) and Pl-Fe by atomic absorption spectrometry. Fe status indicators were, respectively, (mean±SD): M Hb 113±16 g/L; M S-Ft 36±42 μg/L; M sTfR 6.3±3.1 mg/L; Pl-Fe 170±56 μg/g placenta; Pl-Ft 33±18 μg/g placenta; Pl-TfR 18±18 (range 0–58) μg/g placenta. Pl-Fe, Pl-Ft and Pl-TfR did not correlate to M Hb, M S-Ft and M sTfR. Women with Pl- Fe, Pl-Ft and Pl-TfR above or below the corresponding median values did not show any statistical significant difference in M Hb, M sTfR or M S-Ft values. Pl-Ft concentration was lower in women with Hb<110 g/L than in women with normal values: 26±13 vs. 38±20 μg/g, respectively ($p=0.021$). When Pl-TfR, Pl-Ft and Pl-Fe were compared in women with M S-Ft above or below the cut-off point of 10 or 20 μg/L, no significant difference was found for Pl-TfR neither for Pl-Ft nor Pl-Fe. These results suggest that maternal indicators of Fe status, particularly M sTfR and M S-Ft, do not reflect Fe status of the placenta at delivery.     

Purification and characterization of an extremely dimethylsulfoxide tolerant esterase from a salt-tolerant Bacillus species isolated from the marine environment of the Sundarbans

A Bacillus sp. isolated from the Sundarbans region of the Bay of Bengal (NCBI GenBank Accession no. AY723697) which can tolerate 10% (w/v) NaCl, produces esterase optimally in Marine Broth 2216 medium containing 1% (w/v) NaCl. The enzyme was purified 42.7-fold with 6.4% recovery, (specific activity 569.2 U/mg protein) by ammonium sulphate precipitation followed by anion and cation exchange chromatography. The serine type esterolytic enzyme has a molecular weight of 35.0 kDa and is denatured into polypeptides of molecular weights 20 kDa and 15 kDa. The esterase was most active at pH 8.0, the pH of the seawater at the site of collection and is stable in the pH range 6.0–9.0. The optimum temperature of activity of this esterase is 45 °C and the enzyme is very stable after 1 h pre-incubation at 50 °C. Our esterase shows about 100% activity when incubated with 1 M NaCl, the activity drops to about 50% when incubated with 2.5 M sodium chloride and the enzyme is completely inactivated when 4 M NaCl is present during reaction. The esterase is almost inactivated by Ca²⁺, Hg²⁺ and Fe³⁺ ions, reducing agents and detergent. Interestingly, Co²⁺, a known inhibitor of many enzymes, preserved 70% of the activity of this esterase. Specific activity of the esterase increases more than twofold in the presence of water-miscible organic solvents as compared to that in aqueous buffer. When incubated for a period of 10 days in the presence of 30–70% dimethylsufoxide (DMSO), the specific activity increased by approximately two–threefold compared to the enzyme in aqueous buffer throughout the period of study. Specific activity between 1283 and 525 U/mg was maintained by our enzyme when incubated with 50% DMSO for 10 days. The enzyme was most active on p-nitrophenyl acetate, ethyl acetate, alpha isomer of naphthyl acetate but shows relatively lesser activity towards triglycerides of fatty acids. Certain characteristics, such as molecular weight, effects of NaCl, metal ions (Zn²⁺ and Mg²⁺) and reactivity towards para-nitrophenyl and aliphatic esters were strikingly similar to already described marine bacterial derived esterases. Extreme stability in DMSO could make this enzyme a potential immobilized biocatalyst for application in non-aqueous based continuous bioprocesses. Higher specific activity and purification factor, better thermo tolerance and solvent stability would make our enzyme more attractive for biotechnological applications than the marine microbial derived esterases described so far.     

Synthesis and in vitro α-chymotrypsin inhibitory activity of 6-chlorobenzimidazole derivatives

A library of benzimidazole derivatives 1–20 were synthesized, and studied for their α-chymotrypsin (α-CT) inhibitory activity in vitro. Kinetics and molecular docking studies were performed to identify the type of inhibition. Compound 1 was found to be a good inhibitor of α-chymotrypsin enzyme (IC₅₀ = 14.8 ± 0.1 μM, K_i = 16.4 μM), when compared with standard chymostatin (IC₅₀ = 5.7 ± 0.13 μM). Compounds 2–8, 15, 17, and 18 showed significant inhibitory activities. All the inhibitors were found to be competitive inhibitors, except compound 17, which was a mixed type inhibitor. The substituents (R) in para and ortho positions of phenyl ring B, apparently played a key role in the inhibitory potential of the series. Compounds 1–20 were also studied for their cytotoxicity profile by using 3T3 mouse fibroblast cells and compounds 3, 5, 6, 8, 12–14, 16, 17, 19, and 20 were found to be cytotoxic. Molecular docking was performed on the most active members of the series in comparison to the standard compound, chymostatin, to identify the most likely binding modes. The compounds reported here can serve as templates for further studies for new inhibitors of α-chymotrypsin and other chymotrypsin-like serine proteases enzymes.

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