Nanoparticle-Shielded dsRNA Delivery for Enhancing RNAi Efficiency in Cotton Spotted Bollworm Earias vittella (Lepidoptera: Nolidae).

The spotted bollworm Earias vittella (Lepidoptera: Nolidae) is a polyphagous pest with enormous economic significance, primarily affecting cotton and okra. However, the lack of gene sequence information on this pest has a significant constraint on molecular investigations and the formulation of superior pest management strategies. An RNA-seq-based transcriptome study was conducted to alleviate such limitations, and de novo assembly was performed to obtain transcript sequences of this pest. Reference gene identification across E. vittella developmental stages and RNAi treatments were conducted using its sequence information, which resulted in identifying transcription elongation factor (TEF), V-type proton ATPase (V-ATPase), and Glyceraldehyde -3-phosphate dehydrogenase (GAPDH) as the most suitable reference genes for normalization in RT-qPCR-based gene expression studies. The present study also identified important developmental, RNAi pathway, and RNAi target genes and performed life-stage developmental expression analysis using RT-qPCR to select the optimal targets for RNAi. We found that naked dsRNA degradation in the E. vittella hemolymph is the primary reason for poor RNAi. A total of six genes including Juvenile hormone methyl transferase (JHAMT), Chitin synthase (CHS), Aminopeptidase (AMN), Cadherin (CAD), Alpha-amylase (AMY), and V-type proton ATPase (V-ATPase) were selected and knocked down significantly with three different nanoparticles encapsulated dsRNA conjugates, i.e., Chitosan-dsRNA, carbon quantum dots-dsRNA (CQD-dsRNA), and Lipofectamine-dsRNA conjugate. These results demonstrate that feeding nanoparticle-shielded dsRNA silences target genes and suggests that nanoparticle-based RNAi can efficiently manage this pest.

Inducible nitric oxide synthase inhibitors: A comprehensive update.

Inducible nitric oxide synthase (iNOS), which is expressed in response to bacterial/proinflammatory stimuli, generates nitric oxide (NO) that provides cytoprotection. Overexpression of iNOS increases the levels of NO, and this increased NO level is implicated in pathophysiology of complex multifactorial diseases like Parkinson's disease, Alzheimer's disease, multiple sclerosis, rheumatoid arthritis, and inflammatory bowel disease. Selective inhibition of iNOS is an effective approach in treatment of such complex diseases. l-Arginine, being a substrate for iNOS, is the natural lead to develop iNOS inhibitors. More than 200 research reports on development of nitric oxide synthase inhibitors by different research groups across the globe have appeared in literature so far. The first review on iNOS, in 2002, discussed the iNOS inhibitors under two classes that is, amino acid and non-amino acid derivatives. Other review articles discussing specific chemical classes of iNOS inhibitors also appeared during last decade. In the present review, all reports on both natural and synthetic iNOS inhibitors, published 2002 onwards, are studied, classified, and discussed to provide comprehensive information on iNOS inhibitors. The synthetic inhibitors are broadly classified into two categories that is, arginine and non-arginine analogs. The latter are further classified into amidines, five- or six-membered heterocyclics, fused cyclics, steroidal type, and chalcones analogs. Structures of the most/significantly potent compounds from each report are provided to know the functional groups important for incurring iNOS inhibitory activity and selectivity. This review is aimed to provide a comprehensive view to the medicinal chemists for rational designing of novel and potent iNOS inhibitors.

A panoramic review of IL-6: Structure, pathophysiological roles and inhibitors.

Interleukin-6 (IL-6) is a pleiotropic pro-inflammatory cytokine. Its deregulation is associated with chronic inflammation, and multifactorial auto-immune disorders. It mediates its biological roles through a hexameric complex composed of IL-6 itself, its receptor IL-6R, and glycoprotein 130 (IL-6/IL-6R/gp130). This complex, in turn, activates different signaling mechanisms (classical and trans-signaling) to execute various biochemical functions. The trans-signaling mechanism activates various pathological routes, like JAK/STAT3, Ras/MAPK, PI3K-PKB/Akt, and regulation of CD4+ T cells and VEGF levels, which cause cancer, multiple sclerosis, rheumatoid arthritis, anemia, inflammatory bowel disease, Crohn's disease, and Alzheimer's disease. Involvement of IL-6 in pathophysiology of these complex diseases makes it an important target for the treatment of these diseases. Though some anti-IL-6 monoclonal antibodies are being used clinically, but their high cost, only parenteral administration, and possibility of immunogenicity have limited their use, and warranted the development of novel small non-peptide molecules as IL-6 inhibitors. In the present report, all molecules reported in literature as IL-6 inhibitors have been classified as IL-6 production, IL-6R, and IL-6 signaling inhibitors. Reports available till date are critically studied to identify important and salient structural features common in these molecules. These analyses would assist medicinal chemists to design novel and potent IL-6 production and signaling inhibitors, through knowledge- and/or computer-based approaches, for the treatment of complex multifactorial diseases.

Stability Testing of Herbal Drugs: Challenges, Regulatory Compliance and Perspectives.

Stability testing is an important component of herbal drugs and products (HDPs) development process. Drugs regulatory agencies across the globe have recommended guidelines for the conduct of stability studies on HDPs, which require that stability data should be included in the product registration dossier. From the scientific viewpoint, numerous chemical constituents in an herbal drug are liable to varied chemical reactions under the influence of different conditions during its shelf life. These reactions can lead to altered chemical composition of HDP and consequently altered therapeutic profile. Many reports on stability testing of HDPs have appeared in literature since the last 10 years. A review of these reports reveals that there is wide variability in temperature (-80 to 100 °C), humidity (0-100%) and duration (a few hours-36 months) for stability assessment of HDPs. Of these, only 1% studies are conducted in compliance with the regulatory guidelines for stability testing. The present review is aimed at compiling all stability testing reports, understanding key challenges in stability testing of HDPs and suggesting possible solutions for these. The key challenges are classified as chemical complexity and biochemical composition variability in raw material, selection of marker(s) and influences of enzymes. Copyright © 2016 John Wiley & Sons, Ltd.

Coumarin hybrids as novel therapeutic agents.

Naturally occurring coumarins, having wide spectrum of activities such as antioxidant, anti-inflammatory, anticancer, MAO-B inhibitory and antimicrobial, are frequently used by the researchers to develop novel synthetic and semisynthetic coumarin based therapeutic agents. Many of these agents are hybrid molecules, which are designed through concept of molecular hybridization and have shown multiple pharmacological activities. This multifunctional attribute of these hybrid compounds makes them potential drug candidates for the treatment of multifactorial diseases such as cancer, Alzheimer's disease, metabolic syndromes, AIDS, malaria, and cardiovascular diseases. The present review compiles research reports on development of different coumarin hybrids, classify these on the basis of their therapeutic uses and propose structure-activity relationships. It is intended to help medicinal chemist in designing and synthesizing novel and potent hybrid compounds for the treatment of different disorders.

MS(n) , LC-MS-TOF and LC-PDA studies for identification of new degradation impurities of bupropion.

Three new degradation impurities of bupropion were characterized through high performance liquid chromatography coupled to photodiode array detection and to time-of-flight mass spectrometry. Bupropion was subjected to the ICH prescribed stress conditions. It degraded to seven impurities (I-VII) in alkaline hydrolytic conditions which were optimally resolved on an XTerra C18 column (250 × 4.6 mm, 5 µm) with a ternary mobile phase comprising ammonium formate (20 mm, pH 4.0), methanol and acetonitrile (75:10:15, v/v). The degradation impurities (III-V and VII) were characterized on the basis of mass fragmentation pattern of drug, accurate mass spectral and photodiode array data of the drug and degradation impurities. Compound V was found to be a known degradation impurity [1-hydroxy-1-(3-chlorophenyl)propan-2-one], whereas III, IV and VII were characterized as 2-hydroxy-2-(3'-chlorophenyl)-3,5,5-trimethylmorpholine, (2,4,4-trimethyl-1,3-oxazolidin-2-yl)(3-chlorophenyl)-methanone and 2-(3'-chlorophenyl)-3,5,5-trimethylmorphol-2-ene, respectively. Compound III was a known metabolite of the drug. This additional information on the degradation impurities can help in the development of a new stability-indicating assay method to monitor the stability of the drug product during its shelf-life as well as in development of a drug product with increased shelf-life.

Preparation and cyclodextrin assisted dissolution rate enhancement of itraconazolium dinitrate salt.

The poor solubility of itraconazole (ITR) results in its variable oral absorption and bioavailability and has also proven to be a major setback in developing an efficient oral delivery system. To improve its solubility and dissolution profile, itraconazolium dinitrate salt (ITRDNT) was prepared and characterized using various spectral and thermal techniques. The morphology of the salt was studied using optical and scanning electron microscopy (SEM). Broth microdilution assay demonstrated antifungal efficacy of ITRDNT similar to ITR against four different fungal strains namely, Asparagillus fumigatus, Microsporum canis, Microsporum gypsum and Trichophyton rubrum. The salt exhibited better solubility profile than ITR in water and a number of pharmaceutical solvents. Dissolution studies revealed the total amount of drug released from ITRDNT in 3 h was four times greater than that of ITR. To further improve dissolution characteristics, the physical mixtures of ITR and ITRDNT with two cyclodextrins, ß-cyclodextrin (ß-CD) and HP-ß-cyclodextrin (HP-ß-CD) were prepared and their molar ratios were optimized. It was observed that about 75% of drug was released in 30 min from 1:3 molar ratio of ITRDNT and HP-ß-CD physical mixture, which was distinctly higher than ITR commercial capsules (70%). Owing to its facile and economical preparation and substantially better in vitro release profile, the ITRDNT and its CD physical mixtures could be better and cost effective alternatives to ITR and commercial ITR capsules.

Ditosylate salt of itraconazole and dissolution enhancement using cyclodextrins.

Salt formation has been a promising approach for improving the solubility of poorly soluble acidic and basic drugs. The aim of the present study was to prepare the salt form of itraconazole (ITZ), a hydrophobic drug to improve the solubility and hence dissolution performance. Itraconazolium ditolenesulfonate salt (ITZDITOS) was synthesized from ITZ using acid addition reaction with p-toluenesulfonic acid. Salt characterization was performed using (1)H NMR, mass spectrometry, Fourier transform infrared spectroscopy, differential scanning calorimetry, and X-ray diffraction. The particle size and morphology was studied using dynamic light scattering technique and scanning electron microscopy, respectively. The solubility of the salt in water and various pharmaceutical solvents was found multifold than ITZ. The dissolution study exhibited 5.5-fold greater percentage release value in 3 h of ITZDITOS (44.53%) as compared with ITZ (8.54%). Results of in vitro antifungal studies using broth microdilution technique indicate that ITZDITOS possessed similar antifungal profile as that of ITZ when tested against four fungal pathogens. Furthermore, the physical mixtures of ITZDITOS with two cyclodextrins, ß-cyclodextrin (ß-CD), and 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) were prepared in different molar ratios and were evaluated for in vitro release. It was observed that in only 30 min of dissolution study, about 74 and 81% of drug was released from 1:3 molar ratios of ITZDITOS with ß-CD and ITZDITOS with HP-ß-CD, respectively, which was distinctly higher than the drug released from ITZ commercial capsules (70%). The findings warrant further preclinical and clinical studies on ITZDITOS so that it can be established as an alternative to ITZ for developing oral formulations.

Design, Synthesis and Evaluation of Benzimidazole Hybrids to Inhibit Acetylcholinesterase and COX for Treatment of Alzheimer's Disease.

BACKGROUND: A simultaneous administration of an acetylcholinesterase (AChE) inhibitor and a NSAID as a drug cocktail has been documented to exhibit significantly protective effects in AD patients. But it suffers from poor patent compliance, pharmacodynamics and pharmacokinetic issues. OBJECTIVE: The present study is aimed to design and synthesize a hybrid molecule capable of exhibiting both AChE inhibition and anti-inflammatory activities for de-accelerating the progression of AD. The synthesized molecules will be evaluated for in vitro and in vivo models. METHODS: The present study involves the coupling of ibuprofen or naproxen to varied disubstituted amines (AChE inhibitor pharmacophore) through benzimidazole to develop two series of compounds i.e. IB01-IB05 and NP01-NP05. The synthesized compounds were characterized using FTIR, 1H-NMR, 13C-NMR and MS. All compounds were evaluated for in vitro AChE inhibitory and COX inhibitory activities. The most active compound was taken for in vivo evaluation. RESULTS: Compounds of series IB01-IB05 are found more potent as compared to NP01-NP05. The maximally potent compound IB04 in in vitro evaluation is selected for in vivo evaluation of memory restoration activity using scopolamine-induced amnesia model in mice. It significantly reverses the scopolamine-induced changes (i.e., escape latency time, mean time spent in target quadrant, brain AChE activity and oxidative stress) in a dose-dependent manner. IB04 at 8 mg/kg is significantly effective in lowering AD manifestation in comparison to donepezil. CONCLUSION: The findings indicate that Benzimidazole hybrids utilizing ibuprofen and pyrrolidine moiety may prove a useful template for the development of new chemical moieties against AD with multiple potencies.

MS2/TOF and LC-MS/TOF studies on toremifene to characterize its forced degradation products.

The present study was designed to characterize the possible degradation products of toremifene under varied conditions as prescribed by ICH guidelines Q1A(R2). The forced degradation studies were conducted on toremifene citrate under the conditions of hydrolysis (acidic, basic and neutral), photolysis, oxidation and dry heat. The drug was found unstable to photolysis and hydrolysis in water and acidic media but stable to alkaline hydrolysis, peroxide oxidation and thermal degradation. In total fifteen degradation products (I-XV) were formed, which were resolved from each other and the drug on a C-18 column employing an isocratic elution method. A complete mass fragmentation pattern of the drug was established with the help of LC/ESI-MS/TOF to assist characterization of the degradation products. Of the fifteen products, six products III, IV, VII, VIII, XIV and XV were detected in LC-MS. The molecular masses of III, IV, VII and VIII were found to be the same i.e., 387, while those of XIV and XV were 389 and 403, respectively. Structures of these products were elucidated through comparison of their mass fragmentation patterns with the drug, which were proposed on the basis of accurate masses of the parent and fragment ions. These were characterized as (Z)-2-(2-(dimethylamino)ethyl)-4-(4-hydroxy-1,2-diphenylbut-1-enyl)phenol (III), (E)-2-(2-(dimethylamino)ethyl)-4-(4-hydroxy-1,2-diphenylbut-1-enyl)phenol (IV), (E)-4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-en-1-ol (VII), (Z)-4-(4-(2-(dimethylamino)ethoxy)phenyl)-3,4-diphenylbut-3-en-1-ol (VIII), 2-(4-(10-(2-chloroethyl)phenanthren-9-yl)phenoxy)-N-methylethanamine (XIV), and 2-(4-(10-(2-chloroethyl)phenanthren-9-yl)phenoxy)-N,N-dimethylethanamine (XV). Finally, a most plausible mechanistic explanation for degradation of the drug in different chemical environments is also proposed. The results of the study disclose six new degradation related impurities of the drug.

Validated stability-indicating HPLC-UV method for simultaneous determination of glipizide and four impurities.

A selective stability-indicating HPLC-UV method for simultaneous determination of glipizide and four impurities (DPs I-IV) formed under hydrolytic conditions was developed and validated. The drug and impurities were resolved on an XTerra C18 column (250 x 4.5 mm id) in a single gradient run using buffer (0.005 M KH2PO4; pH 3.0)-methanol (60 + 40, v/v; mobile phase A) and (20 + 80, v/v; mobile phase B) at a flow rate of 0.5 mL/min with 230 nm detection wavelength. The method was linear across concentration ranges of 0.2-100, 0.1-100, 0.5-100, 0.2-100, and 0.1-50 microg/mL for glipizide and DPs I-IV, respectively. The RSD for intraday and interday precision for the drug and impurities was < 1 and < 1.2%, respectively. Satisfactory recoveries (96.58-99.97%) of each of the three concentrations selected across the linearity range of each analyte were obtained, proving the method was sufficiently accurate. The LOD was 0.07, 0.05, 0.16, 0.08, and 0.05 microg/mL and the LOQ was 0.20, 0.14, 0.50, 0.23, and 0.14 microg/mL for the drug and DPs I-IV, respectively. Each peak was resolved with resolution of > 2 from the nearest peak. Insignificant changes in retention time (< 4%) and calculated amount (< 1.65%) of drug and each impurity upon small but deliberate changes in various chromatographic parameters were observed, suggesting the method was robust. The method was applied successfully to stability testing of glipizide tablets.

Assessment of shelf-life of Gymnema sylvestre extract through WHO recommended stability study involving chromatographic and biological activity analyses.

Stability study on Gymnema sylvestre extract under WHO recommended accelerated and long-term conditions for 6 and 30 months, respectively was carried out by taking gymnemagenin as a marker and by evaluating antidiabetic activity through different models. Gymnemagenin was not detected in any stability sample indicating that gymnemic acids (GAs) remain stable in the extract under the test conditions. The extract and its GA rich fraction exhibited mild α-glucosidase inhibitory activity (18-27%) that remained same during the study. Neither hypoglycemic nor anti-hyperglycemic effect was induced by the extract in normal rats in oral glucose tolerance test. The extract and GA rich fraction showed significant antidiabetic activity in alloxan-induced diabetic rats that remained same in all stability samples. Based on these findings, a shelf-life of at least 30 months is suggested for G. sylvestre extract under long-term conditions, and gymnemagenin as a marker for shelf-life assessment of products derived from the plant.

Prevalence of reproductive drugs usage in humans and animals: A pilot study in Patiala city of India.

Reproductive drugs that include contraceptive and fertility drugs are used to manage reproductive health in both humans and animals. Contraceptive drugs are mainly used by humans for reversible contraception whereas fertility drugs are mainly used in animals to increase milk production, poultry products and meat production. Usage of these drugs has increased manifold in the last decade. These drugs are excreted through body fluids (mainly urine and milk) that lead to contamination of surface water, milk and animal produce. Consumption of such contaminated products or water results in reproductive disorders and different types of cancers in humans. This questionnaire-based study was designed and conducted involving gynecologists, pharmacies, medical stores and veterinarians in Patiala city and its adjoining areas in India to evaluate the quantitative and qualitative aspects of use of these drugs. A total of 150 survey points were identified with random sampling method. Data was analyzed using appropriate statistical tools. The results showed that contraceptive drugs constitute 86% of reproductive drugs usage in humans. Further, steroidal contraceptives constitute a huge 94.7% share of contraceptive drugs, and of these combined oral contraceptives have 79.79% share among which a combination is ethinylestradiol and levonorgestrel is the most popular (20.92%). The consumption of COCs is higher than that of progestin only pills (Z = 3.39) as well as estrogen only pills (Z = 4.30). In contrast, usage of non-hormonal fertility drugs (89%) dominates over the hormonal class (11%) in humans. The most widely used non-hormonal fertility drug is clomiphene citrate (73.87%). In animals, the prescription rate of hormonal fertility drugs is higher (83%) than the non-hormonal one, where in the most widely prescribed drug is buserelin acetate. These findings are in consonance with the similar studies carried out in US, Europe and Canada which suggest that reproductive drugs usage pattern is more or less similar across the globe. A careful control to discourage indiscriminate use of such drugs is the need of hour to prevent damage of environment and ultimately to the health of living beings.

WHO prescribed shelf life assessment of Syzygium cumini extract through chromatographic and biological activity analyses.

BACKGROUND: Regulatory guidelines recommend shelf life of herbal products to be established through systematic stability studies. OBJECTIVE: The study was designed to establish shelf life of Syzygium cumini extract through accelerated and long-term stability testing as per WHO guidelines. MATERIAL AND METHODS: The extract was stored under accelerated (40°C/75 %RH) and long-term (25°C/60 %RH) stability conditions for 6 and 30 months, respectively. Samples were withdrawn at periodic intervals and analysed through two validated HPLC-UV methods (I and II) for fingerprint and quantitative analysis of markers. Antidiabetic activity of control and stability samples was evaluated by α-glucosidase inhibitory model. RESULTS: Method I generated a well resolved fingerprint of the control sample that was found to contain gallic acid (GA, 1.45 % w/w) and ellagic acid (EA, 3.97 % w/w). The content of GA did not change under both the stability conditions, but that of EA varied insignificantly (3.97-4.77 % w/w) under long-term conditions up to 24 months and subsequently decrease to 3.15 % w/w after 30 months. There was no visible change in LC-UV fingerprint of any stability sample with respect to control. α-Glucosidase inhibitory activity of all stability samples also remained unaltered as compared to control sample (IC50 1.48 mg/mL). GA and EA did not elicit any activity at the concentrations present in the extract. CONCLUSION: Phytochemical composition and antidiabetic efficacy of S. cumini extract remain unchanged during its storage under both accelerated and long-term stability conditions, which suggest its shelf life to be 30 months. Also, GA and EA are not appropriate anti-diabetic markers.

Novel Coupled Molecules from Active Structural Motifs of Synthetic and Natural Origin as Immunosuppressants.

INTRODUCTION: Nitric oxide (NO) is an important mediator in the pathogenesis and control of immune system-related disorders and its levels are modulated by inducible NO synthase (iNOS). Oxidative stress is another pathological indication in majority of autoimmune disorders. The present study aims at the development of coupled molecules via selection of pharmacophores for both immunomodulatory and antioxidant activities through iNOS inhibition. METHODS: Variedly substituted coumarin moieties are coupled with naturally occurring phenols through an amide linkage and were predicted for activities using computer-based program PASS. The compounds predicted to have dual activities were synthesized. Docking studies were carried out against iNOS (PDB 1R35) and compounds having good docking score were evaluated for immunomodulatory and antioxidant activities. RESULTS: The synthesized compounds were found to be pure and were obtained in good yields. Compounds with maximum docking score (YR1a, YR2e, YR2c and YR4e) were selected for evaluation by in vitro models. Compounds YR2e and YR2c markedly inhibited the reduction of NBT dye and showed maximum % iNOS inhibition. In DPPH assay, compound YR4e was observed as the most potent antioxidant (EC50 0.33 µM/mL). Based on these studies, compounds YR2e and YR2c were selected for haemagglutination test. Compound YR2e was observed as the most active immunosuppressant with maximal inhibitory ability of iNOS and NBT reduction and lower HAT value of 3.5. CONCLUSION: Compound YR2e can be utilized as a pharmacological agent in the prevention or treatment of immunomodulatory diseases such as tumors, rheumatoid arthritis, ulcerative colitis, organ transplant and other autoimmune disorders.

Antimicrobial Potential of Benzimidazole Derived Molecules.

Structural resemblance of benzimidazole nucleus with purine nucleus in nucleotides makes benzimidazole derivatives attractive ligands to interact with biopolymers of a living system. The most prominent benzimidazole compound in nature is N-ribosyldimethylbenzimidazole, which serves as an axial ligand for cobalt in vitamin B12. This structural similarity prompted medicinal chemists across the globe to synthesize a variety of benzimidazole derivatives and to screen those for various biological activities, such as anticancer, hormone antagonist, antiviral, anti-HIV, anthelmintic, antiprotozoal, antimicrobial, antihypertensive, anti-inflammatory, analgesic, anxiolytic, antiallergic, coagulant, anticoagulant, antioxidant and antidiabetic activities. Hence, benzimidazole nucleus is considered as a privileged structure in drug discovery, and it is exploited by many research groups to develop numerous compounds that are purported to be antimicrobial. Despite a large volume of research in this area, no novel benzimidazole derived compound has emerged as clinically effective antimicrobial drug. In the present review, we have compiled various reports on benzimidazole derived antimicrobials, classified as monosubstituted, disubstituted, trisubstituted and tetrasubstituted benzimidazoles, bisbenzimidazoles, fused-benzimidazoles, and benzimidazole derivative-metal complexes. The purpose is to collate these research reports, and to generate a generalised outlay of benzimidazole derived molecules that can assist the medicinal chemists in selecting appropriate combination of substituents around the nucleus for designing potent antimicrobials.

Design, synthesis, and evaluation of 5-sulfamoyl benzimidazole derivatives as novel angiotensin II receptor antagonists.

A series of 5-alkylsulfamoyl benzimidazole derivatives have been designed and synthesized as novel angiotensin II (Ang II) receptor antagonists. The compounds have been evaluated for in vitro Ang II antagonism and for in vivo antihypertensive activity on isolated rat aortic ring and desoxycortisone acetate induced hypertensive rats, respectively. The activity is found related to size of alkyl group. The maximum activity is observed with a compact and bulky alkyl group like tert-butyl and cyclohexyl. The compounds 4g and 4h have shown promising both in vitro and in vivo activities. A receptor binding model is also proposed on the basis on the basis of structure-activity relationship in this study.

Angiotensin II--AT1 receptor antagonists: design, synthesis and evaluation of substituted carboxamido benzimidazole derivatives.

A series of 5-(alkyl and aryl)carboxamido benzimidazole derivatives had been designed, synthesized and evaluated for in vitro angiotensin II--AT1 receptor antagonism and in vivo antihypertensive activities. The pharmacological activities were inversely related to the size of alkyl and aryl substituents. It can be suggested that compounds with lower alkyl groups at 5-position of benzimidazole nucleus demonstrated potent antihypertensive activity.

LC-UV-PDA and LC-MS studies to characterize degradation products of glimepiride.

Degradation products of glimepiride formed under different forced conditions have been characterized through LC-UV-PDA and LC-MS studies. Glimepiride was subjected to forced decomposition under the conditions of hydrolysis, oxidation, dry heat and photolysis, in accordance with the ICH guideline Q1A(R2). The reaction solutions were chromatographed on reversed phase C8 (150 mm x 4.6mm i.d., 5 microm) analytical column. In total, five degradation products (I-V) were formed under various conditions. The drug degraded to products II and V under acid and neutral hydrolytic conditions while products I, III and IV were formed under the alkaline conditions. The products II and V were also observed on exposure of drug to peroxide. No additional degradation product was shown up under photolytic conditions. All the products, except I, could be characterized through LC-PDA analyses and study of MS fragmentation pattern in both +ESI and -ESI modes. Product I could not be identified, as it did not ionize under MS conditions. The products II, III and V matched, respectively, to impurity B (glimepiride sulfonamide), impurity J and impurity C (glimepiride urethane) listed in European Pharmacopoeia. The product IV was a new degradation product, characterized as [[4-[2-(N-carbamoyl)aminoethyl]phenyl]sulfonyl]-3-trans-(4-methylcyclohexyl) urea. The degradation pathway of the drug to products II-V is proposed, which is yet unreported.

Ultraviolet-photodiode array and high-performance liquid chromatographic/mass spectrometric studies on forced degradation behavior of glibenclamide and development of a validated stability-indicating method.

A forced degradation study on glibenclamide was performed under conditions of hydrolysis, oxidation, dry heat, and photolysis and a high-performance column liquid chromatographic-ultraviolet (HPLC-UV) method was developed to study degradation behavior of the drug under the forced conditions. The degradation products formed under different forced conditions were characterized through isolation and subsequent infrared/nuclear magnetic resonance/mass spectral analyses, or through HPLC/mass spectrometric (HPLC/MS) studies. The drug degraded in 0.1 M HCI and water at 85 degrees C to a major degradation product, 5-chloro-2-methoxy-N-2-(4-sulfamoylphenyl)ethyl]benzamide (III), and to a minor product, 1-cyclohexyl-3-[[4-(2-aminoethyl)-phenyl]sulfonyl]urea (IV). Upon prolonged heating in the acid, the minor product IV disappeared, resulting in formation of 5-chloro-2-methoxy-benzoic acid (II) and an unidentified product (I). Heating of the drug in 0.1 M NaOH at 85 degrees C yielded II and IV as the major products and I and III as the minor products. The drug and the degradation products formed under different conditions were optimally resolved on a C18 column using ammonium acetate buffer (0.025 M, pH 3.5)-acetonitrile (45 + 55) mobile phase at a flow rate of 0.6 mL/min, with detection at 230 nm. The method was validated for linearity, precision, accuracy, and specificity. Limit of detection (LOD) and limit of quantitation (LOQ) values were also determined. The method could be successfully applied for simultaneous quantification of glibenclamide and the major product, III. The response of the method was linear in a narrow [0.4-10 micro/mL, correlation coefficient (r2) = 0.9982] and a wide (0.4-500 microg/mL, r2 = 0.9993) concentration range for glibenclamide, and in the concentration range of 0.025-50 microg/mL (r2 = 0.9998) for III. The method proved to be precise and accurate for both glibenclamide and III. It was specific for the drug and also selective for each degradation product, and LOQ values for the drug were 0.1 and 0.4 microg/mL, whereas those for III were 0.010 and 0.025 microg/mL, respectively.