Simultaneous Determination of EDTA, Sorbic Acid, and Diclofenac Sodium in Pharmaceutical Preparations Using High-Performance Liquid Chromatography

A simple high-performance liquid chromatographic method for simultaneous determination of ethylenediaminetetraacetic acid (EDTA), sorbic acid, and diclofenac sodium was developed and validated. Separation was achieved on a C₁₈ column (10 cm × 4.6 mm) using gradient elution. The mobile phase consisted of acetonitrile–ammonium dihydrogen phosphate buffer solution (0.01 M, pH = 2.5, containing 0.8% tetra-n-butyl ammonium hydroxide). The detector wavelength was set at 254 nm. Under these conditions, separation of three compounds was achieved in less than 10 min. The effect of two metal salts and metal concentration on peak area of EDTA was investigated. The pH effect on retention of EDTA and sorbic acid was studied. The method showed linearity for EDTA, sorbic acid, and diclofenac in the ranges of 2.5–100.0, 5.0–200.0, and 20.0–120.0 μg/mL, respectively. The within- and between-day relative standard deviations ranged from 0.52 to 1.94%, 0.50 to 1.34%, and 0.78 to 1.67% for EDTA, sorbic acid, and diclofenac, respectively. The recovery of EDTA, sorbic acid, and diclofenac from pharmaceutical preparation ranged from 96.0–102.0%, 99.7–101.5%, to 97.0–102.5%, respectively. To the best of our knowledge, this is the first report about simultaneous determination of EDTA, sorbic acid, and diclofenac.KEY WORDS: diclofenac sodium, EDTA, high-performance liquid chromatography, pharmaceutical preparations, sorbic acid     

Altered Splicing of the BIN1 Muscle-Specific Exon in Humans and Dogs with Highly Progressive Centronuclear Myopathy

Amphiphysin 2, encoded by BIN1, is a key factor for membrane sensing and remodelling in different cell types. Homozygous BIN1 mutations in ubiquitously expressed exons are associated with autosomal recessive centronuclear myopathy (CNM), a mildly progressive muscle disorder typically showing abnormal nuclear centralization on biopsies. In addition, misregulation of BIN1 splicing partially accounts for the muscle defects in myotonic dystrophy (DM). However, the muscle-specific function of amphiphysin 2 and its pathogenicity in both muscle disorders are not well understood. In this study we identified and characterized the first mutation affecting the splicing of the muscle-specific BIN1 exon 11 in a consanguineous family with rapidly progressive and ultimately fatal centronuclear myopathy. In parallel, we discovered a mutation in the same BIN1 exon 11 acceptor splice site as the genetic cause of the canine Inherited Myopathy of Great Danes (IMGD). Analysis of RNA from patient muscle demonstrated complete skipping of exon 11 and BIN1 constructs without exon 11 were unable to promote membrane tubulation in differentiated myotubes. Comparative immunofluorescence and ultrastructural analyses of patient and canine biopsies revealed common structural defects, emphasizing the importance of amphiphysin 2 in membrane remodelling and maintenance of the skeletal muscle triad. Our data demonstrate that the alteration of the muscle-specific function of amphiphysin 2 is a common pathomechanism for centronuclear myopathy, myotonic dystrophy, and IMGD. The IMGD dog is the first faithful model for human BIN1-related CNM and represents a mammalian model available for preclinical trials of potential therapies.     

Metal based isatin-derived sulfonamides: Their synthesis,characterization, coordination behavior and biological activity

Some isatin derived sulfonamides and their transition metal [Co(II), Cu(II), Ni(II), Zn(II)] complexes have been synthesized and characterized. The structure of synthesized compounds and their nature of bonding have been inferred on the basis of their physical (magnetic susceptibility and conductivity measurements), analytical (elemental analyses) and spectral (IR, ¹H NMR and ¹³C NMR) properties. An octahedral geometry has been suggested for Co(II), Ni(II) and Zn(II) and square-planar for Cu(II) complexes. In order to assess the antibacterial and antifungal behavior, the ligands and their metal(II) complexes were screened for their in vitro antibacterial activity against four Gram-negative species, Escherichia coli, Shigella flexneri, Pseudomonas aeruginosa and Salmonella typhi and two Gram-positive species, Staphylococcus aureus and Bacillus subtilis and, for in vitro antifungal activity against Trichophyton longifusus, Candida albicans, Aspergillus flavus, Microsporum canis, Fusarium solani and Candida glaberata. In vitro cytotoxic properties of all the compounds were also studied against Artemia salina by brine shrimp bioassay. The results of average antibacterial/antifungal activity showed that zinc(II) complexes were found to be the most active against one or more bacterial/fungal strains as compared to the other metal complexes.     

The Effect of Biochar and Nitrogen Inhibitor on Ammonia and Nitrous Oxide Emissions and Wheat Productivity

Journal of Plant Growth Regulation - A field experiment was conducted study the effect of biochar and a nitrification inhibitor (nitrapyrin) on NH3 and N2O emissions from a silt clay loam soil near...     

Ranging and foraging of Himalayan grey langurs (Semnopithecus ajax) in Machiara National Park, Pakistan

Grey langurs (Semnopithecus spp.) occupy a variety of habitats, ranging from lowland forests and semi-desert to alpine forests. Little is known about their foraging and ranging in alpine forests, which appear to contain less food than lowland forests. We conducted a 1-year study of Himalayan grey langurs (Semnopithecus ajax) in Machiara National Park, Pakistan, where they occur at relatively high altitudes (range 2000–4733 m). We followed three groups of different sizes and compositions and examined the effects of ecological and social factors on ranging and feeding. The home-range sizes of a small bisexual group (SBG), a large bisexual group (LBG), and an all-male group (AMG) were 2.35 ± 0.92 (mean ± SD; average of four seasons), 3.28 ± 0.55, and 3.52 ± 1.00 km², respectively, and were largest in winter for all groups. The daily path lengths of the SBG, LBG, and AMG were 1.23 ± 0.28 (mean ± SD; average of four seasons), 1.75 ± 0.34, and 1.84 ± 0.70 km, respectively; that of the LBG was longer in winter, while that of the AMG was shorter in summer. Both the home-range size and daily path length of the AMG were larger than those of the other groups, even after partialling out the effect of group size differences. The mean altitude used by the langurs and the proportion of animals seen feeding did not differ among seasons or group types. As the mean temperature increased, the altitude used by langurs significantly increased for the SBG and LBG, but not for the AMG. On the other hand, as the temperature increased, the home-range sizes significantly decreased for the SBG and AMG, but not for the LBG. Rainfall did not show any correlation with ranging or feeding in any of the groups. Our results suggested that grey langurs in Machiara National Park employ a high-cost, high-return foraging strategy in winter, and that the ranging of the AMG also reflects its reproductive strategy.     

An Integrated Diagnosis Strategy for Congenital Myopathies

Congenital myopathies are severe muscle disorders affecting adults as well as children in all populations. The diagnosis of congenital myopathies is constrained by strong clinical and genetic heterogeneity. Moreover, the majority of patients present with unspecific histological features, precluding purposive molecular diagnosis and demonstrating the need for an alternative and more efficient diagnostic approach. We used exome sequencing complemented by histological and ultrastructural analysis of muscle biopsies to identify the causative mutations in eight patients with clinically different skeletal muscle pathologies, ranging from a fatal neonatal myopathy to a mild and slowly progressive myopathy with adult onset. We identified RYR1 (ryanodine receptor) mutations in six patients and NEB (nebulin) mutations in two patients. We found novel missense and nonsense mutations, unraveled small insertions/deletions and confirmed their impact on splicing and mRNA/protein stability. Histological and ultrastructural findings of the muscle biopsies of the patients validated the exome sequencing results. We provide the evidence that an integrated strategy combining exome sequencing with clinical and histopathological investigations overcomes the limitations of the individual approaches to allow a fast and efficient diagnosis, accelerating the patient’s access to a better healthcare and disease management. This is of particular interest for the diagnosis of congenital myopathies, which involve very large genes like RYR1 and NEB as well as genetic and phenotypic heterogeneity.     

The effect of hydraulic retention time on the performance and fouling characteristics of membrane sequencing batch reactors used for the treatment of synthetic petroleum refinery wastewater

The use of membrane sequencing batch reactors, operated at HRT of 8, 16 and 24 h, was considered for the treatment of a synthetic petroleum wastewater. Increase in HRT resulted in statistically significant decrease in MLSS. Removal efficiencies higher than 97% were found for the three model hydrocarbon pollutants at all HRTs, with air stripping making a small contribution to overall removal. Particle size distribution (PSD) and microscopic analysis showed reduction in the protozoan populations in the activated sludge with decreasing HRT. PSD analysis also showed a higher proportion of larger and smaller sized particles at the lowest HRT. The rate of membrane fouling was found to increase with decreasing HRT; SMP, especially carbohydrate SMP, and mixed liquor apparent viscosity also showed a pronounced increase with decreasing HRT, whereas the concentration of EPS and its components decreased. FTIR analysis identified organic compounds as the main component of membrane pore fouling.     

Model parameter uncertainties in a dual-species biofilm competition model affect ecological output parameters much stronger than morphological ones

Bacterial biofilms are complex microbial depositions on immersed interfaces that form wherever the environmental conditions sustain microbial growth. Despite their name, biofilms can develop in highly irregular structures. Recently several mathematical concepts have been introduced to model these spatially structured microbial populations. Regardless of the type of model, they all have, even for microbially relatively simple systems, many parameters which generally are known at most approximately. We investigate the effect of uncertainties in model parameters on four morphological and four ecological output parameters using a nonlinear diffusion model for a biofilm in which two species compete for a shared nutrient. To this end we conduct an extensive computer simulation experiment for two different levels of data uncertainty, three different hydrodynamic conditions, and two different scenarios of bulk substrate availability. Our results indicate that input model parameter uncertainties have a much larger effect on ecological than on morphological output parameters.     

Co-expression of chimeric chitinase and a polygalacturonase-inhibiting protein in transgenic canola <Emphasis Type="Italic">(Brassica napus)</Emphasis> confers enhanced resistance to <Emphasis Type="Italic">Sclerotinia sclerotiorum</Emphasis>

Objectives

Sclerotinia stem rot (SSR) caused by Sclerotinia sclerotiorum is one of the major fungal diseases of canola. To develop resistance against this fungal disease, the chit42 from Trichoderma atroviride with chitin-binding domain and polygalacturonase-inhibiting protein 2 (PG1P2) of Phaseolus vulgaris were co-expressed in canola via Agrobacterium-mediated transformation.

Results

Stable integration and expression of transgenes in T₀ and T₂ plants was confirmed by PCR, Southern blot and RT-PCR analyses. Chitinase activity and PGIP2 inhibition were detected by colorimetric and agarose diffusion assay in transgenic lines but not in untransformed plants. The crude proteins from single copy transformant leaves having high chitinase and PGIP2 activity (T16, T8 and T3), showed up to 44 % inhibition of S. sclerotiorum hyphal growth. The homozygous T₂ plants, showing inheritance in Mendelian fashion (3:1), were further evaluated under greenhouse conditions for resistance to S. sclerotiorum. Intact plants contaminated with mycelia showed resistance through delayed onset of the disease and restricted size and expansion of lesions as compared to wild type plants.

Conclusions

Combined expression of chimeric chit42 and pgip2 in Brassica napus L. provide subsequent protection against SSR disease and can be helpful in increasing the canola production in Iran.