Glabridin,a Stable Flavonoid of Glycyrrhiza glabra: HPTLC Analysis of the Traditional Formulation

JPC – Journal of Planar Chromatography – Modern TLC - A novel HPTLC method has been developed for the estimation of glabridin in Licorice rhizome and its Unani polyherbal formulation...     

Stability‐indicating HPTLC method for quantitative estimation of silybin in bulk drug and pharmaceutical dosage form

In the present study a novel stability‐indicating high‐performance thin‐layer chromatography (HPTLC) method for quantitative determination of silybin in bulk drug and nanoemulsion formulation has been developed and validated on silica using solvent chloroform–acetone–formic acid (9 : 2 : 1 v/v/v) (R_f of silybin 0.46 ± 0.05) in the absorbance mode at 296 nm. The method showed a good linear relationship (r² ± 0.999) in the concentration range 25–1500 ng per spot. It was found to be linear, accurate, precise, specific, robust and stability‐indicating and can be applied for quality control and standardization of several multi‐component hepatoprotective formulations as well as for stability testing of different dosage forms. The method proposed was also used to investigate the kinetics of acidic and alkaline degradation processes by quantification of drug at different temperature to calculate the activation energy and half‐life for silymarin degradation. Copyright © 2009 John Wiley & Sons, Ltd     

Triple Quadrupole Versus High Resolution Quadrupole-Time-of-Flight Mass Spectrometry for Quantitative LC-MS/MS Analysis of 25-Hydroxyvitamin D in Human Serum

We describe a systematic comparison of high and low resolution LC-MS/MS assays for quantification of 25-hydroxyvitamin D₃ in human serum. Identical sample preparation, chromatography separations, electrospray ionization sources, precursor ion selection, and ion activation were used; the two assays differed only in the implemented final mass analyzer stage; viz. high resolution quadrupole-quadrupole-time-of-flight (QqTOF) versus low resolution triple quadrupole instruments. The results were assessed against measured concentration levels from a routine clinical chemiluminescence immunoassay. Isobaric interferences prevented the simple use of TOF-MS spectra for extraction of accurate masses and necessitated the application of collision-induced dissociation on the QqTOF platform. The two mass spectrometry assays provided very similar analytical figures of merit, reflecting the lack of relevant isobaric interferences in the MS/MS domain, and were successfully applied to determine the levels of 25-hydroxyvitamin D for patients with chronic liver disease.

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Development and validation of HPLC method for simultaneous estimation of piperine and guggulsterones in compound Unani formulation (tablets) and a nanoreservoir system

An attempt has been made to develop and validate a simultaneous HPLC method for novel approach of drug release via oil‐in‐water (o/w) nanoemulsion formulation and Habb‐e‐Khardal Unani tablet containing piperine and guggul sterones E and Z as main ingredients. Nanoemulsion was prepared by titration method using sefsol‐218 as an oily phase, cremophor‐EL as a surfactant, transcutol as a co‐surfactant and distilled water as an aqueous phase. The formulation was optimized on the basis of thermodynamic stability and dispersibilty test. The nanoformulation was evaluated for particle size, surface morphology, electrical conductivity and viscosity determination. The in vitro dissolution was carried out by dialysis bag method. Drugs were quantified using an HPLC method developed in‐house with a C₁₈ column as stationary phase and acetonitrile and water as mobile phase at λ_max of 240 nm. The optimized formulation showed higher drug release, lower droplet size and less viscosity as compared with the conventional Habb‐e‐Khardal Unani tablet. The present study illustrated the potential of nanoemulsion dosage form in improving biopharmaceutic performance of piperine and guggul sterone. The HPLC method was also found to be quite sufficient for the routine quality control of formulations containing piperine and guggul sterone E and Z as ingredients and also for in vitro drug release studies. Copyright © 2011 John Wiley & Sons, Ltd.     

Toxicity of naturally occurring Bio-fly and chitosan compounds to control the Mediterranean fruit fly Ceratitis capitata (Wiedemann)

The efficacy of five compounds of a biopolymer chitosan and Bio-fly (Beauveria bassiana fungus) as biopesticide was evaluated on Ceratitis capitata under laboratory conditions. The inhibitory effects on acetylcholinesterase (AChE) and adenosinetriphosphatase (ATPase) as biochemical indicators were also determined in vivo. The results indicated that B. bassiana based Bio-fly exhibited significant toxicity against C. capitata (LC₅₀ = 3008 and 3126 mg/L after 48 h in females and males, respectively) followed by the derivatives of chitosan, N-(4-propylbenzyl)chitosan and N-(2-nitrobenzyl)chitosan. Bio-fly displayed remarkable inhibition of AChE activity (IC₅₀ = 2220 mg/L) while N-(2-chloro,6-flourobenzyl)chitosan, N-(4-propylbenzyl)chitosan and N-(3,4-methylenedioxybenzyl) chitosan had no significant difference in inhibitory action. In adult males, N-(2-nitrobenzyl)chitosan exhibited the highest inhibitory action (IC₅₀ = 6569 mg/L). In addition, the toxic effects of the tested compounds on the activity of ATPase indicated that highly significant inhibition was found with N-(4-propylbenzyl)chitosan with an IC₅₀ of 8194 and 8035 mg/L, in females and males, respectively.     

Effect of the molecular structure on the hierarchical self-assembly of semifluorinated alkanes at the air/water interface

Semifluorinated alkanes (C(n)F(2n+1)C(m)H(2m+1)), short FnHm display local phase separation of mutually incompatible hydrocarbon and fluorocarbon chain moieties, which has been utilized as a structure-forming motif in supramolecular architectures. The packing of semifluorinated alkanes, nominally based on dodecyl subunits, such as perfluoro(dodecyl)dodecane (F12H12) and perfluoro(dodecyl)eicosane (F12H20), as well as a core extended analogue, 1,4-dibromo-2-((perfluoroundecyl)methoxy)-5-(dodecyloxy)benzene) (F11H1-core-H12), was studied at the air/water interface. Langmuir monolayers were investigated by means of neutron reflectivity directly at the air/water interface and scanning force microscopy after transfer to silicon wafers. Narrowly disperse surface micelles formed in all three cases; however, they were found to bear different morphologies with respect to molecular orientation and assembly dimensionality, which gives rise to different hierarchical aggregate topologies. For F12H12, micelles of ca. 30 nm in diameter, composed of several circular or "spherical cap" substructures, were observed and a monolayer model with the fluorocarbon block oriented toward air is proposed. F12H20 molecules formed larger (ca. 50 nm diameter) hexagonally shaped surface micelles that were hexagonally, densely packed, besides more elongated but tightly interlocked wormlike structures. Conversely, F11H1-core-H12 films organized into linear rows of elongated surface micelles with comparable width, but an average length of ca. 400 nm, apparently formed by antiparallel molecular packing.     

In vitro assessment of N-(benzyl)chitosan derivatives against some plant pathogenic bacteria and fungi

Plant pathogenic bacteria and fungi negatively affect a large number of important fruit and vegetables during the growing season and throughout postharvest storage. Therefore, the current study focuses on the preparation of N-(benzyl)chitosan derivatives as antimicrobial agents to control these microorganisms. Chitosan was reacted with a set of aromatic aldehydes by reductive amination involving formation of the corresponding imines, followed by reduction with sodium borohydride to produce the N-(benzyl)chitosan derivatives. The end products were analyzed using ¹H NMR spectroscopy and the degrees of substitution ranged from 0.12 to 0.29. The antibacterial activity was evaluated in vitro against the crown gall disease Agrobacterium tumefaciens (Family: Rhizobiaceae; Class: Alpha Proteobacteria) and the soft mold disease Erwinia carotovora (Family: Enterobacteriaceae; Class: Gamma Proteobacteria) by the nutrient agar dilution method. A higher activity of chitosan and its derivatives was obtained with N-(o-ethylbenzyl)chitosan with a MIC of 500 mg/L against E. carotovora, while N-(o,p-diethoxybenzyl)chitosan was the most active one against A. tumefaciens with a MIC of 1050 mg/L. In addition, the in vitro antifungal assessment against root rot disease Fusarium oxysporum (Family: Tuberculariaceae; Class: Deuteromycetes) and the leaf spots and blights disease Pythium debaryanum (Family: Pythiaceae; Class: Oomycetes) was tested by a mycelial radial growth technique. The data showed that N-(o,p-diethoxybenzyl)chitosan was the most active one with an EC₅₀ of 400 and 468 mg/L for F. oxysporum and P. debaryanum, respectively. In addition, chitosan derivatives had a detrimental effect on spore germination for F. oxysporum. Most of these derivatives exhibited high inhibition percentage (>90%) of spore germination at 1000 mg/L.     

Optimization of Streptomyces bacteriophage ��C31 integrase system to prevent post integrative gene silencing in pulmonary type II cells

φC31 integrase has emerged as a potent tool for achieving long-term gene expression in different tissues. The present study aimed at optimizing elements of φC31 integrase system for alveolar type II cells. Luciferase and β-galactosidase activities were measured at different time points post transfection. 5-Aza-2''deoxycytidine (AZA) and trichostatin A (TSA) were used to inhibit DNA methyltransferase and histone deacetylase complex (HDAC) respectively. In A549 cells, expression of the integrase using a CMV promoter resulted in highest integrase activity, whereas in MLE12 cells, both CAG and CMV promoter were equally effective. Effect of polyA site was observed only in A549 cells, where replacement of SV40 polyA by bovine growth hormone (BGH) polyA site resulted in an enhancement of integrase activity. Addition of a C-terminal SV40 nuclear localization signal (NLS) did not result in any significant increase in integrase activity. Long-term expression studies with AZA and TSA, provided evidence for post-integrative gene silencing. In MLE12 cells, both DNA methylases and HDACs played a significant role in silencing, whereas in A549 cells, it could be attributed majorly to HDAC activity. Donor plasmids comprising cellular promoters ubiquitin B (UBB), ubiquitin C (UCC) and elongation factor 1α (EF1α) in an improved backbone prevented post-integrative gene silencing. In contrast to A549 and MLE12 cells, no silencing could be observed in human bronchial epithelial cells, BEAS-2B. Donor plasmid coding for murine erythropoietin under the EF1α promoter when combined with φC31 integrase resulted in higher long-term erythropoietin expression and subsequently higher hematocrit levels in mice after intravenous delivery to the lungs. These results provide evidence for cell specific post integrative gene silencing with φC31 integrase and demonstrate the pivotal role of donor plasmid in long-term expression attained with this system.