Microanalysis And Preliminary Pharmacokinetic Study Of Sulfated Polysaccharide GFS

Sulfated polysaccharide (GFS), prepared from brown alga (Saccharina japonica),have remarkable renoprotective effect on chronic renal failure (CRF) and diabeticnephropathy (DN). However, it is impossible to use the traditional methods to detectthe polysaccharide because this polysacchairde does not have any chromophores. Inaddition, the concentration of polysaccharides in blood and tissue after metabolizingwas low. Thus a sensitive, simple, convenient method is required for thepharmacokinetic study of GFS. In this thesis, The optimizing reaction conditions ofhigh performance liquid chromatography with post-column derivatization wereanalysed betterly on the basis of fluorescent spectrometry.It was concluded thatHPLC-post-column derivatization is better for the microanalysis of GFS and could besuccessfully applied for the determination of the concentration-time curve andparameters of pharmacokinetics of GFS in New zealand rabbits.Ultrafiltration (MWCO50KDa) is a good way to pre-treat the serum to removeproteins and other impurities. And0.5mol/L sodium chloride was addedsimultaneously to improve the recovery of GFS through increasing the resolution ofbinding drugs.The derivatization reaction is closely related to concentration,temperature andtime etc.In this thesis, The optimizing reaction conditions determined by fluorescencespectroscopy (The optimum wavelength Ex=255nm, Em=430nm) was as follows:100mmol/L guanidine hydrochloride,0.5mol/L sodium hydroxide,120°C reactiontemperature, and reaction time20min.Furthermore,The fluorescence intensity can beenhanced by cooling and rapid detection while no influence such as borax, sodiumchloride, acetonitrile and PBS.Based on the result of spectrometry, the reaction conditions were optimized byHPLC-postcolumn derivatization:200mmol/L guanidine hydrochloride,0.5mol/L sodium hydroxide and125°C reaction temperature. The methodological evaluationshowed that this method fitted the microanalysis of GFS in the serum. The result ofintravenous injection of GFS in New Zealand rabbits showed that Cmaxappeared in5min and it was impossible to detect GFS after6h. In addition, the concentration-timecurve and parameters of Pharmacokinetics fit the Two-compartment model,t1/2α=11.24±2.93min, t1/2β=98.20±25.78min. Furthermore, Cmaxof intragastricadministration appeared in2h approximately.The high sensitivity, good reproducibility, accuracy and simplification ofHPLC-postcolumn derivatization fitted the quantitative microanalysis andpharmacokinetic study of polysaccharides in serum.