The separation of trypanosomes from blood by anion exchange chromatography: From Sheila Lanham's discovery 50 years ago to a gold standard for sleeping sickness diagnosis.

Human African trypanosomiasis (HAT), or sleeping sickness, is a neglected tropical disease that is fatal if untreated, caused by Trypanosoma brucei gambiense and T. brucei rhodesiense. In its 2012 roadmap, WHO targeted HAT for elimination as a public health problem in 2020 and for zero transmission in 2030. Diagnosis of HAT is a multistep procedure comprising of clinical suspicion, confirmation, and stage determination. Suspects are identified on clinical signs and/or on screening for specific antibodies. Parasitological confirmation of suspects remains mandatory to avoid unnecessary toxic drug administration. The positive predictive value of the antibody detection tests is low. Simple parasite detection techniques, microscopic examination of lymph node aspirate, or stained thick blood films lack sensitivity, whereas in T. brucei gambiense patients, the number of blood trypanosomes may be very low. Parasite concentration techniques are therefore indispensable. Half a century ago, Sheila Lanham discovered a technique to separate trypanosomes from the blood of infected rodents, based on anion exchange chromatography with diethyl amino ethyl (DEAE) cellulose, a weak anion exchanger. Between pH 6-9, trypanosome surface is less negatively charged than that of blood cells. When blood is poured on top of a DEAE cellulose column, blood cells are retained, whereas parasites pass the column together with the elution buffer. The result is a pure suspension of trypanosomes that retain their morphology and infectivity. Because cell surface charges vary among trypanosome and mammal species, the optimal buffer pH and ionic strength conditions for different combinations of host and trypanosome species were established. Lanham's technique revolutionized the diagnosis of HAT. It is indispensable in the production of the Card Agglutination Test for Trypanosomiasis (CATT), the most used field test for screening in T. brucei gambiense HAT foci and essential to confirm the diagnosis in suspected people. Lumsden and colleagues developed the mini anion exchange centrifugation technique (mAECT). After adaptation for field conditions, its superior diagnostic and analytical sensitivity compared to another concentration technique was demonstrated. It was recommended as the most sensitive test for demonstrating trypanosomes in human blood. At the beginning of the 21st century, the mAECT was redesigned, allowing examination of a larger volume of blood, up to 0.35 ml with whole blood and up to 10 ml with buffy coat. The plastic collector tube in the new kit is also used for detection of trypanosomes in the cerebrospinal fluid. Unfortunately, mAECT also has some disadvantages, including its price, the need to centrifuge the collector tube, and the fact that it is manufactured on a noncommercial basis at only two research institutes. In conclusion, 50 years after Sheila Lanham's discovery, CATT and mAECT have become essential elements in the elimination of HAT.

CHROMATOGRAPHIC TECHNIQUES IN PHARMACEUTICAL ANALYSIS IN POIAND: HISTORY AND THE PRESENCE ON THE BASIS OF PAPERS PUBLISHED IN SELECTED POLISH PHARMACEUTICAL JOURNALS IN XX CENTURY.

For a long time, chromatographic techniques and techniques related to them have stimulated the development of new procedures in the field of pharmaceutical analysis. The newly developed methods, characterized by improved metrological parameters, allow for more accurate testing of, among others, the composition of raw materials, intermediates and final products. The chromatographic techniques also enable studies on waste generated in research laboratories and factories producing pharmaceuticals and parapharmaceuticals. Based on the review of reports published in Polish pharmaceutical journals, we assessed the impact of chromatographic techniques on the development of pharmaceutical analysis. The first chromatographic technique used in pharmaceutical analysis was a so-called capillary analysis. It was applied in the 1930s to control the identity of pharmaceutical formulations. In the 1940s and 1950s, the chromatographic techniques were mostly a subject of review publications, while their use in experimental work was rare. Paper chromatography and thin layer chromatography were introduced in the 1960s and 1970s, respectively. These new analytical tools have contributed to the intensive development of research in the field of phytochemistry and the analysis of herbal medicines. The development of colunm chromatography-based techniques, i.e., gas chromatography and high performance liquid chromatography took place in the end of 20th century. Both aforementioned techniques were widely applied in pharmaceutical analysis, for example, to assess the stability of drugs, test for impurities and degradation products as well as in pharmacokinetics studies. The first decade of 21" century was the time of new detection methods in gas and liquid chromatography. The information sources used to write this article were Polish pharmaceutical journals, both professional and scientific, originating from the interwar and post-war period, i.e., "Kronika Farmaceutyczna", "Farmacja Wspólczesna", "Wiadomosci Farmaceutyczne", "Acta Poloniae Pharmaceutica", "Farmacja Polska", "Dissertationes Pharmaceuticae", "Annales UMCS sectio DDD Phamacia". The number of published works using various chromatography techniques was assessed based on the content description of individual issues of the journal "Acta Poloniae Pharmaceutica".

Interview with Irving W Wainer.

Irving W Wainer, Senior Investigator in the Intramural Research Program at the National Institute on Aging/NIH received his PhD degree in chemistry from Cornell University and did postdoctoral doctoral studies in molecular biology (University of Oregon) and clinical pharmacology (Thomas Jefferson Medical School). He worked for the US FDA and held positions at St Jude's Children's Research Hospital, at McGill University as Professor in the Department of Oncology, and as a Professor of Pharmacology at Georgetown University. Wainer has published over 350 scientific papers, 10 books, 25 book chapters and holds 11 patents. His awards include: 'A.J.P. Martin Medal' presented by the Chromatographic Society; Doctor HonorisCausa awarded by the Medical University of Gdansk (Gdansk, Poland, 2006), Doctor HonorisCausa awarded by the Department of Medicine, University of Liege (Liege, Belgium, 2012), and the 2013 Eastern Analytical Symposium Award for Outstanding Contributions to the Fields of Analytical Chemistry. Wainer's research includes the development of new therapeutic agents for the treatment of congestive heart failure, cancer, pain and depression, many of which are in the later stages of drug development. His laboratory has also continued the development of cellular membrane affinity chromatography technology, and recent work includes the development of columns containing immobilized forms of the breast cancer resistance protein found in cellular and nuclear membranes and mitochondrial membrane columns. Wainer's laboratory has also continued its study of the effect of disease progression and aging on drug metabolism in critically ill and terminal patients. Interview was conducted by Lisa Parks, Assistant Commissioning Editor of Bioanalysis.

An evolutionary view of chromatography data systems used in bioanalysis.

This is a personal view of how chromatographic peak measurement and analyte quantification for bioanalysis have evolved from the manual methods of 1970 to the electronic working possible in 2010. In four decades there have been major changes from a simple chart recorder output (that was interpreted and quantified manually) through simple automation of peak measurement, calculation of standard curves and quality control values and instrument control to the networked chromatography data systems of today that are capable of interfacing with Laboratory Information Management Systems and other IT applications. The incorporation of electronic signatures to meet regulatory requirements offers a great opportunity for business improvement and electronic working.

Natural product analysis over the last decades.

Progress in natural product chemistry has always been strongly linked to innovations in analytical technology. The characterisation of metabolites in complex mixtures requires sophisticated techniques, which should provide good sensitivity and selectivity as well as structural information on the constituents of interest. This review outlines the most important chromatographic and spectral techniques which have been introduced in the field of natural products. Although there has been a very rapid evolution of methods over the last 50 years, the introduction of high-throughput screening programmes require even more efficient and sensitive methodologies which yield adequate on-line information for metabolite structure determination.

Csaba Horváth's contribution to the theory and practice of capillary electrochromatography.

This overview has been written as a tribute to a luminary of chromatography--Csaba Gyula Horváth, who passed away earlier this year. Since the scope of his work was enormous, the following pages focus only on one single aspect of his scientific activities, capillary electrochromatography. He was a visionary, recognizing the great potential of this method and devoted a large part of his efforts to studies of problems related to CEC, covering a huge variety of issues embracing the theoretical foundation, instrumentation, and column technology. During the period of time from 1996 to 2004, Csaba Horváth published almost 30 excellent papers concerning capillary electrochromatography, which are reviewed in this article.