Composition Of Amino Acids Content In The Clear Open Spirit Injection Method Research

This paper focused on amino acids, peptides and molecular proteins in Qingkailing injection. A method for the determination of20amino acids has been developed and the fingerprints of amino acids were studied. Furthermore, a detecting method for residual proteins was established, which improved the standards of quality and played an significant part in the level of quality control for Qingkailing injection.The main content and results of thesis are as follows:1The establishment of a method for determining the contents of a mino acids in Qingkailing injection.With adoption of O-phthalic aldehyde, Fluorenylmethyl chloroformate and phenylsothio cyanate, we could make determination of amino acids in Qingkailing injection by HPLC-DAD. Compared with the results of the methodological evaluation, we confirmed PITC as a derivatization reagent for the determination of amino acids at last.By analyzing the results of the contents of20amino acids in Qingkailing injection from multiple manufacturers and batches, we confirmed that14amino acids including Glu, Gly, Ala, Pro, Leu, Asp, Ser, Arg, Val, Phe, Orn, Tyr, He, Lys were major ingredients as evaluating total amino acid contents in Qingkailing injectionn according to the Chromatographic resolution, total peaks and how much the contents of amino acids.2The simultaneous determination of fourteen amino acids with one marker in Qingkailing injection.On the basis of determining accurately the contents of14amino acids, a rapid and accurate method was established by calculating relative correction factors of14amino acids in Qingkailing injection when glycine was as an internal standard and slope correction method was adopted by screening the internal reference material and durability of the method. Compared with the results of the two methods (the classic working curve method and RSCFs), the relative error was in the range of0.08%to2.44%. It indicated that the method could be applied to the evaluation of the quality of amino acids in Qingkailing injection.3The building of fingerprint of amino acids in Qingkailing injection.With the determination of amino acids of Qingkailing injection from multiple batches and eight manufacturers, a fingerprint of amino acids in Qingkailing injection was developed. The similarity was more than0.90.4The test and indentification of molecular proteins in Qingkailing injection.By the use of acetone precipitation method, a detecting method of peptides and proteins was built with adoption of SDS-PAGE. With the test of20batches of Qingkailing injection and enzymolysis of protein bands and identifications by mass spectrometer, the results showed that they had molecular proteins in different degree. By searching of NCBInr date base, it indicated that there were two kinds of residual proteins (exsisting in Ox species): Collagen (138854Da) and casein (24420Da).