Development of Immunochromatographic Assay for Identification of Organophosphate Pesticides in Environmental Samples.

Microtiter plate enzyme linked immunoassay (ELISA) experiments in competitive format were performed utilizing polyclonal antibody for establishing a detection system for organophosphate pesticides. IC50 value of and limit of detection (LOD) value was determined by standard inhibition curve and value obtained were 0.05 µgmL(-1) and 0.001 µgmL(-1), respectively. Specificity of antibody was investigated with different organophosphate pesticides. Immunochromatographic assay (ICA) experiments were also designed in competitive format by making use of immunochromatographic strip which was assembly of three main components: conjugate pad, membrane and adsorbent pad. Membrane was coated with hapten-OVA conjugate (test line) and antirabbit IgG (control line). ICA experiments were performed by employing gold-labeled antibody as a detector reagent which was applied over conjugate pad. Visual detection limit obtained from ICA was 0.5 µgmL(-1). Major advantage of strip assay was rapid result, i.e., less than 10 min. which makes it suitable for onsite applications.

Competitive immunochromatographic assay for the detection of thiodiglycol sulfoxide, a degradation product of sulfur mustard.

An immunochromatographic assay (ICA) based on the competitive antigen-coated format using colloidal gold as the label was developed for the detection of thiodiglycol sulfoxide (TDGO), an important metabolite and degradation compound of sulphur mustard (SM). The ICA test strip consisted of a membrane with a detection zone, a sample pad and an absorbent pad. The membrane was separately coated with hapten-OVA conjugate (test line) and anti-rabbit mouse IgG (control line). The visual detection limit for TDGO by ICA detection was found to be 10 µg mL(-1). For validation, the ICA results obtained for spiked water samples were in good agreement with those obtained by indirect competitive inhibition enzyme-linked immunosorbent assay (ELISA) for TDGO. The assay time for detection was less than 10 min. The developed ICA has the potential to be a useful on-site screening tool for the retrospective detection of SM in environmental samples.

Synthesis of some novel phosphorylated and thiophosphorylated benzimidazoles and benzothiazoles and their evaluation for larvicidal potential to Aedes albopictus and Culex quinquefasciatus.

Series of benzimidazole and benzothiazole linked phosphoramidates and phosphoramidothioates (5a-j) and benzimidazole linked phenylphosphoramidates and phenylphosphoramidothioates (10a-e) were synthesized. The title compounds were preliminary screened for mosquito larvicidal properties against Aedes albopictus and Culex quinquefasciatus at different concentration from 40 to 5 mg/L. Among the screened compounds three compounds revealed potential larvicidal effects with 100% mortality in the order of 10e>5j>5e. Compound 10e was found to be the most toxic compound to Ae. albopictus and Cx. quinquefasciatus. The LC50 of 10e against Ae. albopictus was found to be 6.42 and 5.25 mg/L at 24 and 48 h, respectively, whereas it was 7.01 and 3.88 mg/L, respectively in Cx. quinquefasciatus. Temephos was used as positive control.

Direct hapten-linked competitive inhibition enzyme-linked immunosorbent assay (CIELISA) for the detection of O-pinacolyl methylphosphonic acid.

Immunoassay detection of O-pinacolyl methylphosphonic acid (PMPA) employing direct coating of N-2-aminoethyl-O-pinacolyl methylphosphonate (hapten B) on microtiter plates is reported. Coating was achieved by covalently linking hapten B to a glutaraldehyde (GA) polymer network directly bound to the polystyrene (PS) surface of a standard 96-well microtiter plate. 4-(2-(O-Pinacolylmethylphosphoryl amino)ethyl amino)-4-oxobutanoic acid (hapten A)-ovalbumin (OVA) conjugate served as the coating antigen for comparison with direct hapten B-coated plates in the CIELISA format. The developed assay employing direct hapten B coated plates demonstrated enhanced sensitivity with the IC(50) value for PMPA being 0.027 µg mL(-1). The assay could detect PMPA even at the concentration of 0.006 µg mL(-1). The mean recovery of standard PMPA (spiked in water) was found to be 83.7%.

Synthesis and exploration of QSAR model of 2-methyl-3-[2-(2-methylprop-1-en-1-yl)-1H-benzimidazol-1-yl]pyrimido[1,2-a]benzimidazol-4(3H)-one as potential antibacterial agents.

Present communication deals with the synthesis of novel 2-methyl-3-[2-(2-methylprop-1-en-1-yl)-1H-benzimidazol-1-yl]pyrimido[1,2-a]benzimidazol-4(3H)-one derivatives under phase transfer catalysis (PTC) conditions using benzyl triethyl ammonium chloride (BTEAC) as PTC. It also elicits the studies on in vitro antimicrobial evaluation of synthesized compounds against a representative genera of gram-negative and gram-positive bacteria i.e., Bacillus subtilis, Staphylococcus aureus, Pseudomonas diminuta and Escherichia coli. All the compounds have been found to manifest profound antimicrobial activity. Moreover, extensive quantitative structure-activity relationship (QSAR) studies have been performed to deduce a correlation between molecular descriptors under consideration and the elicited biological activity. A tri-parametric QSAR model has been generated upon rigorous statistical treatment.

Oviposition and flight orientation response of Aedes aegypti to certain aromatic aryl hydrazono esters.

Aedes aegypti is a day-biting, highly anthropophilic mosquito and a potential vector of dengue and chikungunya in India. A. aegypti is a container breeder, generally oviposit in the stored and fresh water bodies, and discarded containers near residential areas that provide suitable habitats for oviposition by gravid females. The diurnal activity and endophilic nature of these mosquitoes have increased the frequency of contact with human being. Assured blood meal from human host in an infested area leads to increased disease occurrence. Gravid mosquitoes can potentially be lured to attractant-treated traps and could subsequently be killed with insecticides or growth regulators. In this direction, oviposition by A. aegypti females to aryl hydrazono esters (AHE)-treated bowls at 10 ppm concentration was tested in dual choice experiment, and their orientation response to these ester compounds was studied in Y-tube olfactometer. Among the esters tested, AHE-2, AHE-11 and AHE-12 elicited increased egg deposition with oviposition activity indices (OAI) of +0.39, +0.24 and +0.48, respectively, compared to control; in contrast, AHE-8, AHE-9 and AHE-10 showed negative oviposition response with OAI of -0.46, -0.35 and -0.29, respectively, at 10 mg/L. In the Y-tube olfactometer bioassay, AHE-2 attracted 60 % females compared to control, while to the odour of AHE-11 and AHE-12, about 70 % of the females were trapped in treated chambers. In contrast, only 27-30 % of gravid females entered the chamber releasing AHE-8, AHE-9 and AHE-10 odour plumes, while 70 % entered control chamber, evincing a possible non-preference of treatment odours as well as interference with olfactory receptors. These compounds have the potential for application as oviposition stimulants or deterrents for surveillance and control of mosquito population using ovitraps.

Exploration of in vitro time point quantitative evaluation of newly synthesized benzimidazole and benzothiazole derivatives as potential antibacterial agents.

Present communication deals with the in vitro time point quantitative antibacterial evaluation of newly synthesized 1,2-disubstituted benzimidazoles (3a-p) and 2-substituted benzothiazoles (5a-h) against Gram-positive bacteria Staphylococcus aureus, Bacillus cereus, and Gram-negative bacteria Vibrio cholerae, Shigella dysenteriae and Escherichia coli. These compounds were synthesized under mild reaction conditions using Al(2)O(3)-Fe(2)O(3) nanocrystals as heterogeneous catalyst. Bio-evaluation studies revealed that, compounds 3a, 5a and 5d exhibited moderate to good antibacterial activity against all the tested bacterial stains. The compounds 3a, 3f and 5a have shown enhanced inhibitory activity compared with standard antibacterial drug ciprofloxacin against V. cholerae, B. cereus, and S. dysenteriae, respectively. Additionally, the compounds 3a, 3e, 3f, 3h and 5b displayed complete bactericidal activity within 24 h, whereas ciprofloxacin took 48 h to kill those bacteria completely.

Synthesis and bio-evaluation of aryl hydrazono esters for oviposition responses in Aedes albopictus.

A novel series of aryl hydrazono esters (AHE) (1-13) were synthesized (yield 76-98%) to study the oviposition responses in Aedes albopictus (Skuse) mosquitoes for the first time. At a concentration of 10µgml(-1) in dual choice experiment, among the screened compounds, AHE-12 showed remarkable oviposition attractant activity with an oviposition activity index (OAI) of +0.299 (greater than 95% confidence limit) comparable to p-cresol (OAI +0.320) which is well-reported oviposition attractant for Aedes aegypti. Conversely, AHE-10 exhibited highest oviposition deterrent activity with OAI -0.247. The possible utilization of these compounds will be in integrated vector management strategies.

Screening of core filter layer for the development of respiratory mask to combat COVID-19.

The severe outbreak of respiratory coronavirus disease 2019 has increased the significant demand of respiratory mask and its use become ubiquitous worldwide to control this unprecedented respiratory pandemic. The performance of a respiratory mask depends on the efficiency of the filter layer which is mostly made of polypropylene melt blown non-woven (PP-MB-NW). So far, very limited characterization data are available for the PPE-MB-NW in terms to achieve desired particulate filtration efficiency (PFE) against 0.3 µm size, which are imperative in order to facilitate the right selection of PP-MB-NW fabric for the development of mask. In present study, eight different kinds of PP-MB-NW fabrics (Sample A-H) of varied structural morphology are chosen. The different PP-MB-NW were characterized for its pore size and distribution by mercury porosimeter and BET surface area analyzer was explored first time to understand the importance of blind pore in PFE. The PP-MB-NW samples were characterized using scanning electron microscopy so as to know the surface morphology. The filtration efficiency, pressure drop and breathing resistance of various PP-MB-NW fabric samples are investigated in single and double layers combination against the particle size of 0.3, 0.5 and 1 µm. The samples which are having low pore dia, high solid fraction volume, and low air permeability has high filtration efficiency (> 90%) against 0.3 µm particle with high pressure drop (16.3-21.3 mm WC) and breathing resistance (1.42-1.92 mbar) when compared to rest of the samples. This study will pave the way for the judicial selection of right kind of filter layer i.e., PP-MB-NW fabric for the development of mask and it will be greatly helpful in manufacturing of mask in this present pandemic with desired PFE indicating considerable promise for defense against respiratory pandemic.

In vitro and in vivo antimalarial evaluation of semi-synthetic derivatives of gomphostenin.

A novel series of semi-synthetic gomphostenin derivatives (1-9) were prepared utilizing C-14 hydroxyl group for the first time and studied for their antimalarial properties. In vitro antiplasmodial activity was evaluated against both the chloroquine sensitive and resistant strains of Plasmodium falciparum. Most of the compounds exhibited superior or comparable antiplasmodial activity compared to parent compound, that is, gomphostenin (GN). Based upon in vitro antiplasmodial activity, compounds with IC(50) values less than 10 microM were selected for in vivo antiplasmodial evaluation against Plasmodium berghei infection in mice model. GN derivatives 3 and 5 were found to have curative activity with moderate chemosuppression of 65% and 69%, respectively, at the dose level of 150 mg/kg/day.

Gomphostenins: two new antimalarial compounds from the leaves of Gomphostemma niveum.

Phytochemical investigation of CHCl(3) extract of the Gomphostemma niveum leaves led to the isolation of two new diterpene, compound 1 and 2. Their structures were elucidated by spectroscopic procedures and single crystal XRD. Compound 1 named as Gomphostenin 1 and structure was established as 8-ethyl (5H-furan-2-one, 14-hydroxy, 2-oxo 3, 20 Z(17) diene clerodane, while compound 2 named as Gomphostenin-A; was found to be the acetyl derivative of compound 1 and revealed as 8-ethyl (5H-furan-2-one, 14-acetoxy, 2-oxo 3, 20 Z(17) diene clerodane. In vitro antimalarial activity against Plasmodium falciparum showed that compound 2 was more active than compound 1 and CHCl(3) extract as well; with IC(50) value of 3.4 microg/mL.

Use of polystyrene-supported 2-isobutoxy-1-isobutoxycarbonyl-1,2-dihydroquinoline for the preparation of a hapten-protein conjugate for antibody development.

Polystyrene-supported 2-isobutoxy-1-isobutoxycarbonyl-1,2-dihydroquinoline (PS-IIDQ), a polymer-supported covalent coupling reagent, was successfully employed for the first time in the bioconjugation of an example hapten (phytanic acid derivative) to a carrier protein (bovine serum albumin (BSA)) within the context of immunogen preparation for antibody development. The ability of the prepared example phytanic acid derivative-BSA conjugate to bind an anti-phytanic acid antibody was confirmed using an enzyme-linked immunosorbent assay (ELISA).

Synthesis and antimalarial evaluation of cyclic beta-amino acid-containing dipeptides.

This paper describes an efficient synthesis and the antiparasitic evaluation of cyclic beta-amino acid-containing dipeptides 3.1-3.6 and 4.1-4.5. The antimalarial properties of all these dipeptides have been evaluated in vitro against Plasmodium falciparum and in vivo against Plasmodium berghai. Compounds 4.4 and 4.5 have been found to be very effective in this respect, with IC50 values of 3.87 and 3.64 microg/mL in the in vitro test, while 4.5 has also been found to be active in the in vivo evaluation.

Synthesis of 4-aryl substituted 3,4-dihydropyrimidinones using silica-chloride under solvent free conditions.

This paper describes an improved procedure for the efficient and facile synthesis of 4-aryl substituted 3, 4-dihydropyrimidinones under mild reaction conditions with excellent yields using inexpensive silica chloride under solvent free conditions.

Design and synthesis of immunoconjugates and development of competition inhibition enzyme-linked immunosorbent assay (CIEIA) for the detection of O-isopropyl methylphosphonofluoridate (sarin): an organophosphorous toxicant.

Three haptens of the organophosphorus (OP) toxicant 'sarin' having different spacer arm were designed and synthesized. Haptens were conjugated with BSA (bovine serum albumin) and ovalbumin (OVA) for raising antibody and coating antigen. High antibody titer with higher specificity was obtained from 4-(4-(isopropoxy(methyl)phosphoryloxy)phenylamino)-4-oxobutanoic acid (hapten B) having reasonable long spacer arm. For the standard curve, an IC(50) (inhibitory concentration) of free antigen was found to be 0.415 µg mL(-1) on the basis of indirect competitive ELISA. The study revealed that heterology in competition inhibition enzyme immunoassay (CIEIA) produced remarkable improvement in the sensitivity and specificity of the assay. Under the optimized conditions, the quantitative working range was found to be 0.19-1.56 µg mL(-1) with a limit of detection (LOD) of 0.05 µg mL(-1). The antibodies showed negligible cross reactivity (CR) with other OP toxicants and pesticides, which makes the assay suitable for the selective detection of sarin.

Synthesis and characterisation of immunogens for the production of antibodies against small hydrophobic molecules with biosignature properties.

In the present study, five different classes of small hydrophobic molecular targets, atypical for antibody generation, were structurally modified in order to introduce suitable reactive functionalities and/or spacers which allow covalent coupling to a carrier protein resulting in a stable carrier-hapten complex. These targets were chosen to serve as markers of extant and/or extinct life in the context of the development of the Life Marker Chip (LMC), an antibody-based instrument, which is being developed by a UK-led international consortium for flight to Mars on board the joint ESA/NASA Mars exploration ExoMars mission. The hapten-protein conjugates were designed to be used as immunogens for antibody generation and immunoassay reagents in subsequent stages of the LMC development. The extent of protein modification due to covalent attachment of hapten was determined by two independent methods, i.e. trinitrobenzenesulfonic acid (TNBSA) titrations of remaining protein reactive groups and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of the resultant hapten-protein conjugates. In a further quality validation step, the conjugates were presented to an animal's immune system and polyclonal antibody titres with moderate specificity were obtained. These results suggest that conjugates synthesized as described herein can successfully be used in the generation of antibodies targeting small hydrophobic molecules.