Development and application of a validated HPLC method for the analysis of dissolution samples of levothyroxine sodium drug products

A rapid, selective, and sensitive gradient HPLC method was developed for the analysis of dissolution samples of levothyroxine sodium tablets. Current USP methodology for levothyroxine (L-T(4)) was not adequate to resolve co-elutants from a variety of levothyroxine drug product formulations. The USP method for analyzing dissolution samples of the drug product has shown significant intra- and inter-day variability. The sources of method variability include chromatographic interferences introduced by the dissolution media and the formulation excipients. In the present work, chromatographic separation of levothyroxine was achieved on an Agilent 1100 Series HPLC with a Waters Nova-pak column (250 mm × 3.9 mm) using a 0.01 M phosphate buffer (pH 3.0)-methanol (55:45, v/v) in a gradient elution mobile phase at a flow rate of 1.0 mL/min and detection UV wavelength of 225 nm. The injection volume was 800 μL and the column temperature was maintained at 28°C. The method was validated according to USP Category I requirements. The validation characteristics included accuracy, precision, specificity, linearity, and analytical range. The standard curve was found to have a linear relationship (r(2)>0.99) over the analytical range of 0.08-0.8 μg/mL. Accuracy ranged from 90 to 110% for low quality control (QC) standards and 95 to 105% for medium and high QC standards. Precision was <2% at all QC levels. The method was found to be accurate, precise, selective, and linear for L-T(4) over the analytical range. The HPLC method was successfully applied to the analysis of dissolution samples of marketed levothyroxine sodium tablets.     

Solid lipid nanoparticles (SLNTM/LipopearlsTM) a pharmaceutical and cosmetic carrier for the application of vitamin E in dermal products

Solid lipid nanoparticles (SLNTM, LipopearlsTM) are nanoparticles made from solid lipids by highpressurehomogenization. Incorporation of chemically labile active ingredients intothe solid lipid matrix protects against chemical degradation, which is shown for vitamin E. The SLN are physically stable in aqueous dispersions and also after incorporation into a dermal cream as proven by photon correlation spectroscopy and differential scanning calorimetry. Electron microscopy and atomic force microscopy data reveal the spherical shape of the SLN and the detailed structure of the particle surface. Ultrafine particles form an adhesive film leading to an occlusive effect on the skin. The occlusion promotes the penetration of vitamin E into the skin, as shown by the stripping test. In addition to chemical stabilization of active ingredients, occlusive effects on the skin and subsequent enhanced penetration of compounds, the SLN also possess a pigment effect covering undesired colours leading to an increased aesthetic acceptance by the customer.     

Development of a HPLC method for the determination of antichagasic phenylethenylbenzofuroxans and its major synthetic secondary products in the chemical production processes

A simple isocratic reverse-phase HPLC method for the determination of six antichagasic phenylethenylbenzofuroxans and its major synthetic secondary products, the corresponding geometric isomers and the benzofurazans, was developed and validated for use in the analysis of pre-clinical studies. Separation was achieved on a reverse-phase Supelco LC-18 column using either methanol-acetonitrile-water or acetonitrile-water, in different proportions, as mobile phase. The compounds were eluted isocratically at a flow rate of either 0.8 or 1.0 mLmin(-1). The compounds were analyzed with UV detection at 210 and 300 nm. The validation characteristics included linearity, accuracy, precision, specificity, limit of detection and quantification and robustness. Validation acceptance criteria were met in all cases. This method was used successfully for the quality assessment of the drugs production in the scale-up procedures.     

Development of a lateral flow dipstick for simultaneous and semi‐quantitative analysis of dihydroartemisinin and piperaquine in an artemisinin combination therapy

Dihydroartemisinin (DHA) and piperaquine (PPQ) are two drugs used in an artemisinin‐based combination therapy (ACT). The circulation of counterfeit antimalarial drugs demands the development of simple, point‐of‐care (POC) tests for monitoring drug quality. Here we aimed to design an antibody‐based lateral flow dipstick assay for simultaneous quality control of DHA and PPQ. To obtain a monoclonal antibody (mAb) for PPQ, one structural unit of the symmetric PPQ molecule was used to derive a carboxylic acid for linkage to a carrier protein as immunogen. Screening of hybridoma cells identified an mAb 4D112B2 that reacted with the PPQ‐based immunogen. A highly‐sensitive icELISA was designed based on this mAb, which showed 50% inhibition concentration of PPQ at 1.66 ng/mL and a working range of 0.35 – 7.40 ng/mL. The mAb showed 10.2, 15.9 and 30.4% cross reactivity to hydroxychloroquine sulfate, chloroquine and amodiaquine, respectively. No cross reactivity was observed to lumefantrine, mefloquine artemisinin and its derivatives. Using our previous DHA dipstick design, a lateral flow dipstick for simultaneous analysis of PPQ and DHA was developed. The indicator ranges for PPQ and DHA were 2 – 5 μg/mL and 250 – 500 ng/mL, respectively. The dipstick was used to semi‐quantitatively analyze PPQ and DHA content in commercial ACT drugs, which produced agreeable results to those determined by high‐performance liquid chromatography. This combination dipstick makes it a potential POC device for quality control of the two active ingredients in a commonly used ACT.     

Optimization of a miniaturized solid-phase microextraction method followed by gas chromatography mass spectrometry for the determination of twenty four volatile and semivolatile compounds in honey from Galicia (NW Spain) and foreign countries

An analytical miniaturized methodology based on solid-phase-microextraction (mini_SPME) followed by gas chromatography coupled to mass spectrometry (GC-MS) has been developed for the identification of volatile and semivolatile compounds in honey samples. The main influential experimental parameters, such as the type of fibre coating, extraction temperature, solvent addition, extraction mode, ionic strength, and sample dilution were optimized. A design of experiments (DOE) was conducted including twenty-four target compounds. The final extraction conditions comprised the use of 200 mg of honey mixed with 200 μL of water (100%, w/v), employing a DVB/CAR/PDMS fibre in the headspace mode at 100 °C for 30 min. The mini_SPME-GC-MS method was successfully validated in terms of linearity, repeatability, reproducibility and accuracy. Finally, it was applied to a broad range of varieties of real honey samples from Galicia (NW Spain), as well as some foreign honeys, demonstrating suitability.     

Development and validation of a high-performance thin-layer chromatographic method for the quantitative analysis of vitexin in Passiflora foetida herbal formulations

The aim of this study is the development of validated HPTLC method for the quantification of vitexin from Passiflora foetida commercial herbal formulations. The developed method was validated, in accordance with ICH guidelines for precision, accuracy, specificity and robustness. The plate was developed using ethyl acetate:methanol:water:formic acid 30:4:2:1(%, v/v/v/v) on 20 × 10 cm glass coated silica gel 60 F₂₅₄ plates and the developed plate was scanned and quantified densitometrically at λ = 340 nm. Linear regression analysis revealed a good linear relationship between peak area and amount of vitexin in the range of 100–700 ng/spot. The amount of vitexin in nine commercial herbal formulations was successfully quantified by the developed HPTLC method. The developed and validated high performance thin layer chromatographic method offers a new sensitive and reliable tool for quantification of vitexinin in various herbal formulations containing Passiflora foetida.     

Development and validation of HPLC methods for the determination of potential extractables from elastomeric stoppers in the presence of a complex surfactant vehicle used in the preparation of parenteral drug products

HPLC methods were developed and validated for potential extractables [zinc dithiocarbamate, 2,6-di-tert-butyl-para-cresol (BHT), octylated diphenylamine antioxidant, sulfur, pentylphenol, and tetrakis(methylene(3,5-di-tert-butyl-4-hydroxyhydro cinnamate))methane] from commercial elastomeric stoppers in a complex surfactant matrix. These stoppers were proposed to be part of the container-closure system for experimental formulations containing the surfactant, polyoxyethylated Castor oil (USP/NF) (POE Castor oil) and ethanol. The presence of POE Castor oil in the formulation posed unique challenges to the development and validation of the HPLC methods. POE Castor oil, also known as Cremophor, is a viscous and complex solubilizing agent with a number of uncharacterized fractions. Hence the goal was to identify HPLC conditions that would be suitable for the separation, detection, and quantitation of the potential stopper extractables in the presence of such a complex drug product matrix. A number of experiments were performed to evaluate the effects of different columns, mobile phase composition, injection volume, and gradient profile on the separation and detection of the potential stopper extractables. The quantitation limits of these stopper extractables are between 1 and 10 ppm. The methods demonstrate good linearity, acceptable accuracy, and precision.