Study Of Horizontal Centrifugal Counter-current Chromatography On Pharmaceutical Analytical Application

Counter-current chromatography is a liquid-liquid partitioning chromatography, broadly applied in separation and preparation of natural products. In modern era of CCC, high-speed counter-current chromatography (HCC) and centrifugal counter-current chromatography (CCC) have been broadly applied because of the advantages of great stationary phase retention, rapid separation and convenient operation. To our knowledge, development of CCC in China is at initial stage and few studies have been reported. Extensive and innovative development of CCC shall have great significance on pharmaceutical analytical application.Salicylic acid works as a keratolytic by causing the cells of the epidermis to shed more readily. The determination method of salicylic acid ointment is titration after diluting samples. However the assay results are assumed to be negatively affected by the enormous systematical error of the method, and color changes of the indicator solutions affected by the ointment base. Chlorhexidine gluconate works as an antimicrobial disinfection antiseptic. The determination method of chlorhexidine gluconate ointment is Ultra-Visual Spectrophotometry after extracting their active ingredients. However the complicated and time-consuming method has enormous systematical error and the ointment base is hard to remove, which negatively affects the results. Clotrimazole works as an antifungal medication, and acicolvir as a guanosine analogue antiviral drug. The determination method of the both ointments is High-performance liquid chromatography (HPLC) after extracting their active ingredients. It is difficult to remove the ointment base during the time-consuming and complicated preparation of the solutions, which will affect the assay results as well as the enormous systematical error of the method.We have applied centrifugal counter-current chromatography (CCC) to analyze content determination of active ingredients in the four ointments, as well as systematically studied the affection of the solvent system, solvent physical parameters, injection volume, sample solvent and preparation of samples on stationary phase retention, chromatographic peak type, baseline steadiness and quantitative results. The results indicated that by using the solvent system n-hexane-acetonitrile-water (2:1:1, V/V), the sample solvent of5%tween80aqueous solution and injection volume of1.0ml, the proposed method for the four ointments showed good linearity, and the RSDs of precision, stability and repeatability test results were less than2.0, respectively. In conclusion, the simple, rapid and accurate method has methodological advantages of good durability of equipment and short inspection period over HPLC ointment assays frequently described in the literature. The method shall have great guiding significance on separation and quantitative analysis of ointment active ingredients.Referring to CCC analysis on single-recipe ointments, we developed a optimized CCC separation and analytical method on multi-recipe somedons. The optimized solvent system was n-hexane-ethyl acetate-methanol and water (4:5:8:10, V/V/V/V), sample solvent: methanol-water (8:10, V/V), sample volume:1.0ml. By using CCC technology, the proposed method for the four active ingredients showed good linearity within linear range, with the average recovery of98%-102%. The analytical method is able to quantitatively determine the content of four somedon active ingredients rapidly and conveniently. The study offers more versatility in the choice of somedon content determination methods and promotes the CCC widespread use in the drug control.