Isolation and characterization of acid and base degradation products in Atenolol and Hydrochlorothiazide and a validated selective stability-indicating HPLCUV method for their quantification

Atenolol (( RS )-2-{4-[2-Hydroxy-3-(propan-2-ylamino)propoxy]phenyl}acetamide) and Hydrochlorothiazide (6-chloro-1,1-dioxo-3,4-dihydro-2 H -1,2,4-benzothiadiazine-7-sulfonamide) are beta 1 (? 1 ) receptor blocker and diuretic drug respectively; however the combination dosage regime are used for cardiovascular therapy. Thus a forced degradation study was carried out upon this combination drug regime under acidic and basic environment in order to deconvolute the possible degradation product under specified stressed conditions. Under acidic conditions atenolol and hydrochlorothiazide were cleaved into 2-(4-(3-amino-2- oxopropoxy) phenyl) acetamide and 6-sulphamido benzothiazide. However, under basic conditions, the drugs were spliced into 2-(4-(2-hydroxypropoxy) phenyl) acetamide and 2-chloro 4-amino 1, 6-dihydro benzene sulphonamide respectively. The degradant peaks were elucidated by HPLC using C18 column with methanol: phosphate buffer (70:30 v/v) with a flow rate of 0.5ml/min (UV detection at 226nm). For quantitative method validation, linearity was observed over product concentration range 2g/ml - 100 g/ml (r 2 0.9992) with regression equation y=43432x. The products were first identified by LC-MS and further confirmed by FT-IR and 1 H 1 NMR. A specific and sensitive stability-indicating assay method for the simultaneous determination of the drugs, its process related impurities and degradation products was developed

Enzyme-Free Detection of Mutations in Cancer DNA Using Synthetic Oligonucleotide Probes and Fluorescence Microscopy

Rapid reliable diagnostics of DNA mutations are highly desirable in research and clinical assays. Current development in this field goes simultaneously in two directions: 1) high-throughput methods, and 2) portable assays. Non-enzymatic approaches are attractive for both types of methods since they would allow rapid and relatively inexpensive detection of nucleic acids. Modern fluorescence microscopy is having a huge impact on detection of biomolecules at previously unachievable resolution. However, no straightforward methods to detect DNA in a non-enzymatic way using fluorescence microscopy and nucleic acid analogues have been proposed so far.Here we report a novel enzyme-free approach to efficiently detect cancer mutations. This assay includes gene-specific target enrichment followed by annealing to oligonucleotides containing locked nucleic acids (LNAs) and finally, detection by fluorescence microscopy. The LNA containing probes display high binding affinity and specificity to DNA containing mutations, which allows for the detection of mutation abundance with an intercalating EvaGreen dye. We used a second probe, which increases the overall number of base pairs in order to produce a higher fluorescence signal by incorporating more dye molecules. Indeed we show here that using EvaGreen dye and LNA probes, genomic DNA containing BRAF V600E mutation could be detected by fluorescence microscopy at low femtomolar concentrations. Notably, this was at least 1000-fold above the potential detection limit.Overall, the novel assay we describe could become a new approach to rapid, reliable and enzyme-free diagnostics of cancer or other associated DNA targets. Importantly, stoichiometry of wild type and mutant targets is conserved in our assay, which allows for an accurate estimation of mutant abundance when the detection limit requirement is met. Using fluorescence microscopy, this approach presents the opportunity to detect DNA at single-molecule resolution and directly in the biological sample of choice

A multi-parent genetic algorithm for solving longitude–latitude-based 4D traveling salesman problems under uncertainty

In this study, we propose a mathematical model of a 4D clustered traveling salesman problem (CTSP) to address the cost-effective security and risk-related difficulties associated with the TSP. We used a multiparent-based memetic genetic algorithm to optimize paths between all clusters and proposed unique heuristic approaches to create clusters and reconnect them. We constructed a 4D CTSP considering multiple routes between two locations and multiple available vehicles on each route. Travel expenses and risks impact every itinerary; however, the behaviors of these costs and risks are always uncertain. We inspected various standard benchmark problems from (TSPLIB) using the proposed calculations. Real-life problems in the tourism industry motivate a longitude–latitude-based CTSP with risk constraints. Thus, we determined the risk of each path based on longitude and latitude. The contributions of this study are twofold: developing a genetic algorithm and heuristics based on mathematical modeling of a real problem.</p

Novel Phospholipid-Protein Conjugates Allow Improved Detection of Antibodies in Patients with Autoimmune Diseases

Reliable measurement of clinically relevant autoimmune antibodies toward phospholipid-protein conjugates is highly desirable in research and clinical assays. To date, the development in this field has been limited to the use of natural heterogeneous antigens. However, this approach does not take structural features of biologically active antigens into account and leads to low reliability and poor scientific test value. Here we describe novel phospholipid-protein conjugates for specific detection of human autoimmune antibodies. Our synthetic approach includes mild oxidation of synthetic phospholipid cardiolipin, and as the last step, coupling of the product with azide-containing linker and copper-catalyzed click chemistry with β2-glycoprotein I and prothrombin. To prove utility of the product antigens, we used enzyme-linked immunosorbent assay and three cohorts of samples obtained from patients in Denmark (n = 34) and the USA (n = 27 and n = 14). Afterwards we analyzed correlation of the obtained autoantibody titers with clinical parameters for each patient. Our results prove that using novel antigens clinically relevant autoantibodies can be detected with high repeatability, sensitivity and specificity. Unlike previously used antigens the obtained autoantibody titers strongly correlate with high disease activity and in particular, with arthritis, renal involvement, anti-Smith antibodies and high lymphocyte count. Importantly, chemical composition of antigens has a strong influence on the correlation of detected autoantibodies with disease activity and manifestations. This confirms the crucial importance of antigens' composition on research and diagnostic assays, and opens up exciting perspectives for synthetic antigens in future studies of autoimmunity

Restoration of uterine redox-balance by methanolic extract of Camellia sinensis in arsenicated rats

Arsenic, an environmental and industrial pollutant causes female reproductive disturbances and female infertility. Several researchers found that the use of Camellia sinensis (CS) (green tea) is effective as an alternative therapeutic strategy in the management of several health ailments. This study explores the role of CS extract against arsenic-induced rat uterine tissue damage. Methanolic extract of CS (10 mg/kg BW) was tested concomitantly in arsenic-treated (10 mg/kg BW) rats for a duration of two-oestrous cycle length (8 days). CS effectively attenuated arsenic-induced antioxidantdepletion and necrosis in uterine tissue. Rats treated with sodium arsenite showed significantly reduced activities of enzymatic antioxidants like superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in uterine tissue as evidenced by the results of spectrophotometric and electrozymographic analysis. Co-administration of CS significantly reversed the above oxidative stress markers in uterine tissue along with the histopathological changes in ovarian and uterine tissue. Moreover, an increase in the level of transcription factor NF-κB in the uterine tissue in association with reduced serum levels of vitamin B12 and folic acid were mitigated in arsenic fed rats following CS co-administration

Synthesis of Pyridine and Quinoline Based Novel Nheteroaromatics and Targeting Activity Against Macrophage-associated Disease

The work embodied in this thesis describes the synthesis of pyridine and quinoline based novel N-heteroaromatics and targeting activity against macrophageassociated disease. The work covers mainly four areas, (i) Development of novel methodology for the one pot synthesis of linear and angular fused quinazolinones. (ii) Synthesis of tetrahydropyrrolo[3',4':3,4] pyrrolo[2,1-a] isoquinoline-9,11-dione derivatives via a simple and convenient multi compartment reaction in aqueous miceller system. (iii) One pot synthesis of symmetrically 1, 4-disubstituted piperazine- 2, 5-diones. (iv) and Bioactivity of the above synthesized compounds as antileishmaniasis againsts ( Macrophage associated disease). The Chapter I begins with a review on strategic development toward the Pyridine and quinoline based Synthetic and Natural products against Leishmaniasis. Chapter II deals with the development of one step methodology using amino heterocycles and o-bromo benzyl/naphthyl bromides as reactants to produce Nheteroaromatic cationic intermediates, which upon base catalyzed nucleophilic aromatic substitution followed by in situ aerial oxidation at the benzylic position smoothly furnished the angular and linear quinazolinones . Chapter III presents the development of an efficient and environment-friendly novel approach for the synthesis of tetrahydropyrrolo[3',4':3,4] pyrrolo[2,1-a] isoquinoline-9,11-dione derivatives using isoquinolinium ylide (generated in situ from isoquinoline and phenacyl bromide in presence of a base) and an activated dienophile (aryl maleimide) in micellar solution at ambient temperature. Chapter IV describes a series of diketopiperazine derivatives synthesized by self-condensation of differently substituted α-chlorophenyl acetamides in a one-pot sequence in presence of sodium hydride under nitrogen atmosphere. Chapter V deals with the bio activity of the above synthesized compounds against lishmaniasis. The promising activities of the compounds are investigated further with in vivo study