Current Pharmaceutical Analysis - Volume 5, Issue 1, 2009

Identification and isolation of differentially expressed genes is of great interest for a broad range of medical and biological research fields. For pharmaceutical and pharmacological applications the identification of target genes is of importance as well as the investigation of the transcriptional response to drug administration. Over the last two decades several techniques have been developed for analyzing gene expression in a variety of experimental contexts. Whereas microarrays represent closed architecture systems, restricted to well characterized models with a vast amount of available background data (e.g. human, mouse, etc.), open architecture systems allow detection of novel genes without a priori knowledge of the transcriptome. They are often based on amplification, subtraction and comparative display, and are well suited for so-called non-model organisms with little or no sequence data available. We review here the most common methods and their modifications: mRNA fingerprinting, differential display (DD), cDNA representational difference analysis (RDA) and suppression subtractive hybridization (SSH). The focus is on PCR-based methods that can be conducted with minimal bioinformatic and/or high throughput facility support, making them applicable in virtually any laboratory having access to basic molecular biology equipment. Furthermore we will give an insight into applications of these techniques in the pharmaceutical / pharmacological field.

The majority of enantiomeric separations for purity analysis and quality control continue to be performed by normal phase liquid chromatography (LC) and supercritical fluid chromatography (SFC). For much of the pharmaceutical industry, chiral method development relies heavily on column screening activities. Much work has been performed looking at polysaccharide based stationary phases (Diacel AD, OD, AS, and OJ); however, analytical laboratories must be ready to continually address the changing nature of molecules being developed in the pharmaceutical industry. For this reason, a comparison of commercially available Pirkle stationary phases to compliment polysaccharide column screens was studied. Using normal phase LC and SFC, several Pirkle stationary phases were used to screen a study library of pharmaceutically relevant compounds. The results of this investigation are reported.

The electrochemical behaviors of N-(2-benzylbenzoxazol-5-yl)benzamide (B1), N-(2-benzylbenzoxazol-5-yl)- 4-nitrobenzamide (B2) and N-(2-(4-chlorobenzyl)benzoxazol-5-yl)-4-nitrobenzamide (B3) were investigated by cyclic voltammetry (CV), square wave voltammetry (SWV), differential puls voltammetry (DPV), chronoamperometry (CA) and bulk electrolysis (BE) techniques in dimethylsulfoxide (DMSO) containing 0.1 M tetrabutylammonium tetrafluoroborate (TBATFB). The number of electrons transferred and diffusion coefficients were calculated by using chronoamperometry and bulk electrolysis techniques. Standard heterogeneous rate constants for the electrochemical reduction were calculated by Klingler-Kochi technique. The data obtained showed that the quantitative determination of benzoxazoles could be done by using DPV and SWV rapidly and sensitively. For the DPV technique, linear working ranges for B2 and B3 were found to be (6.0 x 10-7-4.0 x 10-4) M and (1.0 x 10-6-2.0 x 10-4) M respectively. The corresponding ranges for these compounds found by SWV were (6.0 x 10-7-4.0 x 10-4) M and (1.0 x 10-6-2.0 x 10-4) M. The detection limits for B2 obtained from DPV and SWV were calculated to be 1.33 x 10-7 M and 1.76 x 10-7 M respectively. The detection limits with B3 deplicted to be almost the same.

Tramadol, a 4-phenyl-piperidine analogue of codeine, is usually marketed as the hydrochloride salt and is a centrally acting analgesic, used for treating moderate to severe pain. In this study, a potentiometric liquid membrane sensors for simple and fast determination of tramadol hydrochloride in pharmaceutical formulation and urine was constructed. For the membrane preparation, tramadol-tetraphenyl borate complexes were employed as electroactive material in the membrane. The wide linear range (10-5-10-1 M), low detection limit (3 μ g/ml), and fast response time (10 s) are the characterizations of the proposed sensors. Validation of the method shows suitability of the sensors for applies in the quality control analysis of tramadol hydrochloride in pharmaceutical formulation and urine. The proposed method is found to be simple, accurate and precise which can be used as a detector for HPLC.

In the last months, 11 countries recalled heparin products because of severe side effects, which included 62 deaths reported to the FDA on or after January 1, 2008, in the USA. Active pharmaceutical ingredients have to fulfil quality requirements. The determination of the API's (Active Pharmaceutical Ingredients) quality relies on the results of analytical testing. The decisions, as to whether the quality is acceptable or not acceptable are risk-based decisions. The quality of analytical data is therefore necessary for the control and release of the batches. Selectivity, specificity and sensitivity are key factors according to the ISO standards, allowing the calculation of positive and negative predictive values. The FDA now postulates the application of two analytical test methods: capillary electrophoresis and H-NMR. To date, no data exist concerning the above mentioned analytical quality parameters. Therefore, in this study, the Bayes' theorem, a statistical tool, was applied to calculate risk probabilities. Furthermore it is discussed: Are the actual API audit and inspection controls effective? From a statistical point of view, are the FDA's proposed analytical methods suitable for the qualified person judgement for release of heparin and other APIs? What is the safety risk of the spot checks-based release practice? Are PAT (Process Analytical Technology) tools, in combination with a whole supply chain management, suitable for optimizing safety and minimizing risk?

Chemical investigation of many bacterial and fungal, as well as plant species has revealed the presence of interesting compounds derived from naphthalene - 1,4-naphthoquinones and rarely also 1,2-naphthoquinones. They were detected in many species of families Bignoniaceae, Droseraceae, Plumbaginace, Boraginaceae, Juglandaceae as well as in species of small families, such as Dioncophyllaceae or Acanthaceae. Naphthoquinones have very interesting spectrum of biological actions, including antibiotic, antiviral, anti-inflammatory, antipyretic, antiproliferative and cytotoxic effects. Because of these properties the plants containing them are used in folk medicines, mainly by natives in Asia, where especially Chinese medicine uses aerial as well as subterranean parts of these plants for hundreds years, and South America. The utilizing of naphthoquinones for medicinal purposes and their occurrence in nature is reviewed and discussed. Moreover, we review analytical techniques using for their analysis.

Drug analysis is important in several phases of drug development, such as formulation, stability studies and quality control. The importance of reliable analytical methods for drug determination in a fast, inexpensive, sensitive and selective way is thus evident. Although there are countless works describing new analytical methods for determination of drugs that act against tropical diseases, a review organizing these works in a systematic and complete way is lacking. In this context, the objective of this review is to present the main advances in the development of analytical methods for determination of tropical disease drugs using electroanalytical, spectrophotometric and liquid chromatographic techniques.