Studies In The Chemical Components And Stablishies Of Hplc Fingerprinter On Nzyme Hydrolysate Of Aishaping Injection

Purpose: The compounds are extracted, separated and purified from Aishuping enzymehydrolysis ingredient, and structures of these compounds are identified. At the same time, thefingerprints of Aishuping are established by the method of HPLC. And the chromatographicpeaks are identified according to the chemical composition of the research results.Method:1. Using classical column chromatography separation and purificationtechnology, the compounds are separated from Aishuping enzyme hydrolysis ingredients. Andthe structure were identified by different spectroscopy methods and techniques such as MassSpectrometry (MS)、Infrared Spectrum(IR)、NMR spectroscopy(1H-NMR、13C-NMR) andso on.2. The optimal chromatographic conditions of Aishuping is established by HPLCthrough various investigations in different aspects, such as the composition and the elutionmodes of mobile phase, temperature of chromatographic column and detection wavelength indifferent HPLC, injection volume and the preparation methods of the sample and so on. Last,the fingerprints of Aishuping are established by HPLC, the chromatographic peaks areidentified, characteristic parameters of corresponding fingerprints are calculated, andsimilarity is evaluated as well.Result:1.Three compounds were extracted, separated and purified from Aishupingenzyme hydrolysis ingredient, and structures of these compounds were identified. Finallytheir names are determined: Cytidine Monophosphoric Acid, Uridylic Monophosphoric Acid,Guanine Monophosphoric Acid.2. The establishment of the chromatographic conditions as follows:(1) ChromatographicColumn: ZORBAX SB-AQ C18（250mm×4.6mm，5um）;(2) Mobile Phase:1‰formicacid-5%acetonitrile, elution process:0～10min1‰formic acid,10～60min1‰formicacid-5%acetonitrile (80:20);(3) Flow Rate:1mL/min;(4) Column Temperature:25℃;(5) Detection Wavelength:260nm;(6) Injection Volume:10uL；(7) Running Time:：60min.（8）Preparation methods of the sample: The mass ration of testing sample and nucleic acidenzyme is5:1, the detection temperature is60℃and hydrolysis time is1h. After the similaritycalculation between the sample fingerprints and standard fingerprints via the evaluationsystem of traditional Chinese medicine fingerprint chromatogram similarity, the result isabove0.9.Conclusion: In this research, the compounds were extracted, separated and purified fromAishuping enzyme hydrolysis ingredient, and structures of these compounds wereidentificated in the experiment. At the same time, the chromatographic conditions, preparationmethods of the sample and the optimum conditions of HPLC fingerprints for Aishuping areestablished. It shows an important meaning on improvement of quality standards and onoptimization of process conditions for Aishuping as well.