Current Pharmaceutical Analysis - Volume 8, Issue 1, 2012

Current Pharmaceutical Analysis has completed seven years of its publication and is now entering its eighth year. During the last seven years it has established itself as a very respectable journal that publishes expert reviews and original research articles on recent advances in pharmaceutical and biomedical analysis. The fields covered by the journal include pharmaceutical, biochemical and clinical analysis with particular reference to drug analysis, analytical methodology and instrumentation. I am grateful to the continued guidance and support of the Associate Editors and Editorial Advisory Board Members who contributed significantly in bringing the journal to its present position. I am also grateful to the dedication and hard work of the staff of Bentham Science Publishers, including Mrs. Afsheen Shahab, Senior Production Editor, Dr. Sadqa Tul-Ain Sheikh and Mr. Asad Ali. I look forward to another good year for the journal with the publication of high quality publications from eminent experts.

Methacycline is a broad spectrum, semisynthetic antibiotic prepared from oxytetracycline. Its antibacterial spectrum is as same as that of tetracycline, but its efficacy is superior to that of oxytetracycline and tetracycline. There is no standard technique for analysis and/or detection of methacycline. Practical requirements for high-sensitivity analysis and the need for a specialist laboratory setting create challenges for routine analysis. Several existing analytical techniques, which offer flexible and broad-based methods of analysis and detection, have been discussed in this manuscript, focusing on bioanalytical, pharmaceutical QC applications and food analysis. This review will examine the published analytical methods reported for determination of methacycline, presenting (a) sample pretreatment methods such as liquid- liquid extraction (LLE) and solid phase extraction (SPE), (b) separation methods such as thin layer chromatography, high performance liquid chromatography (HPLC) and capillary electrophoresis (CE) and (c) others such as potentiometry, spectrophotometry and microbiological assay, from which it can be seen that HPLC methods have been used most extensively.

Deguelin is the major active ingredient and principal component in cube resin, derived from Lonchocarpus utilis and used as a botanical insecticide and pesticide in Africa, South America and China. A growing body of evidence indicates that deguelin has a very strong anti-tumor effect on malignant cells without compromising normal cells. As a complex natural extract, deguelin interacts with various molecular targets to exert its anti-tumor properties at nanomolar levels. This current review delineates the molecular mechanism of the anti-tumor activities of deguelin in malignancies. Our research shows that the multiple and extensive biological effects of deguelin are mediated via interactions with target points involved in cell proliferation pathways (cyclin D1, pRb), cell apoptosis pathways (NF-κB, IκBα, Bcl-2, Bcl-xl, survivin), the nucleophosmin and nucleoporins pathways (Nup88, Nup98, Nup214), as well as through the regulation of steroid receptor co-activator SRC-3 and DNA Topoisomerase. In addition, deguelin holds a promising efficacy for reversing drug resistance.

Antisense oligodeoxynucleotides (AsODN) are rapidly degraded by nucleases in biological fluids which compromises their efficacy as therapeutic agents. This study evaluated two Fluorescence Resonance Energy Transfer (FRET) approaches, Acceptor Photobleaching and Sensitized Emission, in terms of their suitability for quantification of oligonucleotide stability in various colloidal carrier systems in vitro. The influence of the formulations' pH and viscosity on the validity of the two approaches was determined and showed that the donor fluorescence intensity was highly susceptible to pH fluctuations of the medium. Moreover, the Acceptor Photobleaching approach proved to be unsuitable for the proposed studies due to Brownian motion of molecules in liquid formulations, suggesting that this method can only be used for immobilized specimens. The stability of a 30-mer AsODN incorporated into various in situ gelling systems was evaluated using the Sensitized Emission approach. This approach appeared to offer a simple tool to evaluate the stability of AsODN and showed stable molecules over a period of one week. However, a number of criteria, such as photobleaching due to repeated exposure, pH of the surrounding medium and sample preparation need to be carefully considered when performing quantitative FRET measurements.

Electrochemical behavior of disopyramide (DPA) on glassy carbon electrode (GCE) was investigated by using cyclic voltammetry, square-wave voltammetry and constant potential bulk electrolysis. Adsorption and diffusion properties of DPA were evaluated. Differential pulse anodic adsorptive stripping voltammetric (DPAAdSV) and squarewave anodic adsorptive stripping voltammetric (SWAAdSV) methods were developed for its direct determination in pharmaceutical preparations and biological samples. Linear working concentration range for both methods was found as 3x10-7 M (0.102 mg/L) - 3x10-6 M (1.0 mg/L). Limit of detection and limit of quantitation for DPAAdSV were found to be 2.9x10-7 M (0.10 mg/L) and 9.5x10-7 M (0.32 mg/L), respectively and they were calculated to be 1.4x10-7 M (0.05 mg/L) and 4.5x10-7 M (0.15 mg/L) for SWAAdSV. Proposed methods were successfully applied to determine the content of DPA in pharmaceutical preparations and biological samples.

In this paper, chiral fat-soluble enantiomers of 1-phenyl-R, S-tetrahydrogen isoquinoline (ER, ES) are rapidly separated using β-cyclodextrin (β-CD) modified micellar capillary electrokinetic chromatography coupled with electrochemical detection (EC). An effectual micellar suspension of 35 mmol/L phosphate buffer saline (PBS) (pH 7.85) containing 30 mM sodium deoxycholate, 20 mM β-CD and 20% (v/v) acetonitrile was developed as the running buffer. Among them, the surfactant sodium deoxycholate formed the micelles in the buffer, β-CD was employed to improve the separation, and the acetonitrile acted as an organic modifier. After the optimization of the factors such as detection potential, separation voltage, sampling time and the composition of running buffer, baseline separation was obtained within 12 min at 20 kV of separation voltage. The RSD (n = 5) of migration times and peak areas of the analytes are 2.3% (ER), 2.7% (ES) and 2.0% (ER), 3.5% (ES), respectively. The detection limit is 0.5 μmol/L for ER and 0.2 μmol/L for ES, also it was found that trace ER could be detected at the limit proportion of ER to ES for 1:500. This protocol was successfully applied for monitoring the amount of ER from ES in synthetic drug intermediate.

Methods for determination of topiramate in pharmaceutical formulation by high-performance thin-layer chromatography (HPTLC) UV-densitometry and liquid chromatography-mass spectrometry (LC-MS) have been developed. HPTLC method as recommended by United States Pharmacopeia was performed using the silica plates, mobile phase composed of benzene:ethanol (5:2, v/v) and densitometric detection at 340 nm after topiramate visualization with the use of chemical reagent. Moreover, quantification was achieved in the concentration range of 0.25-4.0 μg/spot and with adequate precision (RSD=4.16%) and recovery (104.47%) using non-linear calibration curve by fitting to y=a+blnx. LC-MS method was performed using Zorbax SB-C18 column and isocratic elution mode with a mobile phase composed of acetonitrile:water (85:15, v/v) with 0.1% (v/v) formic acid at a flow rate 0.5 ml/min. Moreover, the single quadrupole mass spectrometer employing ESI interface operated in the negative ion mode was used to quantify the topiramate at m/z=338.1. Proposed LC-MS method provided good results of precision (RSD=2.05%) and recovery (99.53%) in the concentration range of 0.25-10.0 μg/ml using linear y=a+bx regression analysis. Additionally, no interferences were found from tablet excipients at the selected wavelength, mass-to-charge ratio and assay procedures. The developed methods found to be sufficiently precise and reproducible for established conditions and after validation may be used for routine quality control of topiramate in pharmaceutical formulations.

5-fluorouracil is one of the most widely used chemotherapeutic agents for the treatment of several types of cancer. The present study describes the determination of stability of 5-fluorouracil under different stress conditions using high performance liquid chromatography (HPLC) and infrared (IR) spectroscopy. The separation of 5-fluorouracil was carried out on a C18 reverse phase column using 40 mM KH2PO4 as mobile phase. The ultraviolet detection wavelength was 260 nm. The HPLC method was validated and the response of the drug was linear in the concentration range of 20 μg ml-1 to 550 μg ml-1 with a correlation coefficient of 0.9995. The % R.S.D. values for intra-day and inter-day precision were determined as <0.2% and <1% respectively. Infrared spectroscopic measurements were carried out in diffuse reflectance and attenuated total reflectance (ATR) modes. The study reveals that 5-flurouracil shows mild degradation up to 275°C and major degradation at about 285°C. It undergoes a degradation of about 22% when exposed to stress conditions of acidic hydrolysis. A major degradation of about 97% was observed under alkaline condition. The drug undergoes degradation from 26% to 41% when exposed to oxidative condition. The drug remains stable when exposed to UV irradiations.

An HPLC method with a diode-array-detector set at 265 nm, which was developed and validated for the simultaneous determination of ten sulfonamides, in bovine, porcine and chicken tissue is presented. The separation was achieved on an analytical column, Kromasil, C18 5 μm, 250 x 4 mm at ambient temperature. The mobile phase, a mixture of 0.1% formic acid, CH3CN and CH3OH, was delivered by a gradient program. The examined sulfonamides, (sulfadiazine (SDZ), sulfathiazine (STZ), sulfamethoxine (SMTH), sulfamethizole (SMZ), sulfamethoxypyridazine (SMPZ), sulfamonomethoxine (SMMX), sulfamethoxazole (SMXZ), sulfisoxazole (SIX), sulfadimethoxine (SDMX) and sulfaquinoxaline (SQX)) were extracted from the tissues using mixtures of ethyl acetate, water and acetonitrile. SPE was necessary for further purification. The method was validated according to the Decision 2002/657/EC, determining selectivity, stability, decision limit, detection capability, accuracy, linearity and precision. Overall recoveries ranged from 90.1 to 115.1%. All RSD values were lower than 10.0%. The decision limits CCα ranged for bovine tissue from 103.68 to 112.36 μg/kg, for porcine tissue from 108.13 to 110.13 μg/kg and for chicken tissue from 103.93 to 111.92 μg/kg, while detection capability CCβ ranged from 111.62 to 121.26 μg/kg, from 111.74 to 124.83 μg/kg and from 111.32 to 119.86 μg/kg respectively.

A simple, specific and selective RP-HPLC-UV method with Solid Phase Extraction (SPE) was developed for the quantification of cefepime in Total Nutrient Admixtures (TNAs). Chromatographic separation was achieved on C18 column using a mobile phase consisting of a mixture of ammonium acetate, acetonitrile and methanol. The flow rate was set to 0.9 mL min-1 and the wavelength was 257 nm. The analyte was extracted from TNAs by using SPE technique. Cefepime calibration curve was linear over a concentration range of 100-1400 μg mL-1 with a correlation coefficient of ≥0.9994. The intraday accuracy and precision were ≤-4.76% and ≤3.12% respectively. The interday accuracy and precision were ≤-3.90% and ≤4.29% for respectively. The method was successfully applied to stability studies of cefepime in the presence of micronutrients together with low and high concentrations of macronutrients in TNAs. Cefepime was degraded by 42.84% and 58.81 % at room temperature (25±2°C) after 72 hours in low and high macronutrient concentration formulations of TNAs respectively. Cefepime first order degradation rates for low and high macronutrient concentration formulations of TNAs were -0.0032 and -0.0049 hr-1 respectively. These results indicated a higher rate of degradation in TNA formulations with high concentrations of macronutrients.

Dithranol is one of the mainstays in the topical treatment of psoriasis but its unpleasant side effects result in poor patient compliance and there is an on-going demand for novel formulations, with better absorption and dermal availability characteristics. In the developmental stage, such novel formulations are currently evaluated in artificial membranes, animal models and human subjects. Simple, preliminary in vitro cell permeation studies, performed in the initial stages, will serve to reduce the cost, time and number of animals used in the development of new dithranol formulations. A simple and sensitive isocratic reversed-phase high-performance liquid chromatographic (RP-HPLC) method has been optimized and validated for the determination of dithranol. Samples were analyzed on a C18 column with acetonitrile-distilled water-acetic acid (58:37:5, v/v/v) as the mobile phase, with a flow rate of 1.1 ml/min and UV detection at 394 nm. The assay method was found to be linear from 0.098 to 200 μg/ml, with a mean correlation coefficient (r2) of 0.9986 (n=3). All the validation parameters were within the acceptance range. The validated method was then successfully applied to the analysis of the cellular uptake of dithranol in an in vitro study.

This paper demonstrates the high potential of excitation-emission phosphorescence matrices in combination with multivariate regression for determining drugs in a complex matrix. The simultaneous determination of naproxen and propranolol in urine by micelle-sensitized room temperature phosphorescence is made difficult by strong overlap between the three-dimensional phosphorescence spectra for the two analytes and by the inner filter effect arising from the presence of a biological fluid. To circumvent these shortcomings, we used three-dimensional phosphorescence spectroscopy in combination with multivariate calibration methodology. The ensuing method allowed us to determine both compounds at microgram-per-litre levels without the need for separation. Using a calibration matrix in combination with a factorial design comprising two levels per factor and a central star design ensured accurate calibration throughout the desired concentration intervals. The proposed method was validated by application to a test set of synthetic samples. Based on the results, the N-PLS model used affords the accurate simultaneous determination of both analytes in urine samples

A kind of univariate calibration, net analyte signal combined with standard addition (NAS-SAM) is presented for determination of Guaifenesin, Pseudoephedrine and Chlorpheniramine in the presence of each other by spectrophotometry. The method has both the advantages of the standard addition method and net analyte signal which enables the extraction of information concerning a certain analyte from spectra of multi-component mixtures. The use of full spectrum to calculate the net analyte signal vector and also no requirement of calibration and prediction steps are the most important advantages of NAS-SAM. The simultaneous determination of Guaifenesin, Pseudoephedrine and Chlorpheniramine was performed in Britton-Robinson buffer (pH 9). Three pharmaceutical compounds were determined in concentration range of 0.5-100 μmol L-1 In this work, the method of NAS-SAM has been used for the determination of guaifenesin, pseudoephedrine and chlorpheniramine in some ternary mixtures and cough syrups with satisfactory results.

Flucloxacillin (FLU) is a semi-synthetic penicillin active against many gram-positive bacteria such as streptococci and penicilinase-producing staphylococci, including methicillin-susceptible S. aureus. A simple, rapid, reproducible and cost-effective UV-spectrophotometric analytical method was validated to assay flucloxacillin sodium (FLU) in capsules. Measurements were taken at 274 nm using ultra-purified water as the solvent. This new method was validated in accordance with ICH requirements, which include linearity, precision, accuracy, specificity, robustness, detection and quantitation limits. The method demonstrated good linearity over the range of 50.0 to 100.0 mg L-1 FLU with a correlation coefficient (r) equal to 0.9998. The accuracy of the method was 100.09%. The precision demonstrated a relative standard deviation of less than 2.5%. The excipient in the capsules did not interfere with the assay. The results were satisfactory when compared with the literature. The proposed method might be applied in routine quality control in the pharmaceutical industries since it is precise, accurate, simple, economic and produces very low amounts of solvents and residues.

Purpose: Incompatibility between drugs and excipients can alter stability and bioavailability of drugs thereby affecting an active's safety and/or efficacy. The purpose of this investigation was to study the compatibility of α, α'-xylene-p-bis- 3,3'(hydroxyiminomethyl) pyridinium dibromide, (3 PApxy) with a number of commonly used excipients. Methods: Solid dosage forms typically contain diluents, disintegrants, polymers, glidants and lubricants and therefore the compatibility of 3 PApxy with selected excipients from each group was investigated. TGA of Perkin Elmer equipped with Pyris Manager™ software and Mettler Toledo Differential Scanning Calorimeter (DSC). Excipients used were Sodium Laurel Sulphate (SLS), Methyl Cellulose (MC), Hydroxy Propyl methyl cellulose (HPMC), Cellulose acetate (CA), Hydroxy Propyl Cellulose (HPC), Carboxy methyl cellulose (CMC), Ethyl Cellulose (EC), Poly Acryl Amide (PAA), Poly Vinyl Alcohol (PVA), Methyl Paraben (MP), Micro Crystalline Cellulose (MCC), Cetyl trimethyl Ammonium Bromide (CTAB) and Acacia were purchased from Sigma-Aldrich and Alfa-Aesar (USA) and used as such. Results: TGA analysis shows seven out of fourteen excipients, SLS, Betain CMC, MCC, EC, PAA and acacia are found to decrease the thermal stability of drug. The DSC thermogram of 3 PApxy showed a sharp endothermic peak at 267 °C corresponding to its melting point. The DSC thermograms of 3 PApxy and each of the investigated excipients with 1:1 mixtures of 3 PApxy and excipients were compared. On the basis of DSC results, 3 PApxy was found to be compatible with all excipients except SLS. Conclusion: Based on TGA and DSC studies, 3 PApxy was found to be compatible with most commonly used excipients except for SLS. These studies may be relevant when formulating novel dosage form systems for 3 PApxy.