The Principal Component Content Determination And HPLC Fingerprint Establishment For Wasp Venom

ObjectiveTo establish a method for the determination of four principle components B,C,D and F in wasp venom by using HPLC.In order to provid a method and basis for the quality control of wasp venom.To optimize the extraction preparation method and HPLC analysis conditions of wasp venom from different areas and batches.To establish a standard HPLC fingerprint,which provided a quality control of wasp venom.Method1.Four principle components in wasp venom were prepared and represented by character B,C,D and F,The contents of four principle components in three batches of wasp venom were determined by HPLC.2.134 batches of wasp venom were collected from different areas.The fingerprints of wasp venom were established by using Agilent 1260 HPLC,DAD detector,Agilent ZORBAX 300SB C₁₈column（4.6 mm×250 mm,5μm）and 0.1%trifluoroacetic acid water（A）-0.1%trifluoroacetic acidacetonitrile（B）as a mobile phase with gradient elution,and the detecting wavelength was set at 215 nm.The similarity was calculated by the Similarity evaluation system of chromatographic fingerprint of traditional Chinese Medicine（version 2012.130723）.Cluster analysis and principal component analysis were used to study the pattern recognition of wasp venom,and to identify the true or false of wasp venom from different places.Results1.The average recoveries of principal components B,C,D and F were 100.8%（RSD=2.48%）,99.7%（RSD=3.44%）,100.7%（RSD=2.08%）and 101.0%（RSD=2.14%）.The precision,repeatability and stability of the methods for determination of four principal components are good,and the average contents of B,C,D and F were determined to be 109.1 mg/g（RSD=2.30%）、26.3 mg/g（RSD=3.07%）、216.3 mg/g（RSD=0.86%）、58.0 mg/g（RSD=2.99%）in three batches of wasp venom.It showed that there was no difference in the principal components B,C,D and F among the three batches of wasp venom,and the four quantitative chromatographic methods were simple,rapid and accurate.2.The HPLC fingerprint of wasp venom was established.and 12 common peaks were obtained by using principal component B as internal reference peak.Six chromatographic peaks in the standard fingerprint were identified by the method of reference substance comparison.The principal components of wasp venom were identified as A,B,C,D,E,F.The fingerprints of the standard were compared with134 batches of wasp venom chromatographies from different places.The results showed that the similarity of wasp venom with storage numbers S166,S129,S131,S132 and S134 was lower than 0.9.The similarity of other samples was more than 0.9.Therefore,these samples with similarity less than 0.9 were identified as fake products.ConclusionThe HPLC fingerprint of wasp venom and the determination of four principal components were established for the first time.They can reflect the overall characteristics of wasp venom.It provided a theoretical basis for the quality control and further study of wasp venom.