High Resolution Mass Profile of Bufadienolides and Peptides Combing with Anti-Tumor Cell Screening and Multivariate Analysis for the Quality Evaluation of Bufonis Venenum

• Published in: Molecules (Basel, Switzerland) Vol. 24; no. 10; p. 1943
• Main Authors: He, Rongrong, Ma, Hongyue, Zhou, Jing, Zhu, Zhenhua, Lv, Xiang, Li, Quan, Wang, Hengbin, Yan, Yanqing, Luo, Niancui, Di, Liuqing, Wu, Qinan, Duan, Jinao
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 20.05.2019; MDPI
• Subjects: Amino Acid Sequence; Antineoplastic Agents - pharmacology; Biological activity; bufadienolides; Bufanolides - analysis; Bufanolides - pharmacology; Bufonis Venenum; Calibration; Cell Line, Tumor; Chinese medicine; Correlation coefficient; Correlation coefficients; Fractionation; High performance liquid chromatography; High resolution; Humans; Identification; Least-Squares Analysis; Limit of Detection; Liquid chromatography; Markers; Mass Spectrometry; Mass spectroscopy; Metabolites; Modernization; Multivariate Analysis; Multivariate statistical analysis; Peptides; Peptides - analysis; Peptides - chemistry; Peptides - pharmacology; pre-fractionation mass spectrometry profile; Proteins; Qualitative research; Quality assessment; quality evaluation; Quantitative research; Reproducibility of Results; Scientific imaging; Small Molecule Libraries - analysis; Spectroscopy; Statistical analysis; Tumor cells; Vasoactive intestinal peptide; Venom; quality evaluation; bufadienolides; peptides; pre-fractionation mass spectrometry profile; Bufonis Venenum
• ISSN: 1420-3049; 1420-3049
• DOI: 10.3390/molecules24101943

In order to evaluate the quality of Bufonis Venenum commercial herbs, a three-step qualitative and quantitative research study was performed. Firstly, we tried to identify small molecules and peptides in Bufonis Venenum using pre-fractionation chromatography and high-resolution mass spectrometry. The database search of the small molecules and peptides of Bufonis Venenum revealed that the dried venom consisted of free/conjugated-type bufadienolides and peptides with a mass range of 0.4-2 kDa. Secondly, we used partial least squares (PLS) multivariate statistical analysis to screen bufadienolides markers (VIP > 1.5) responsible for the anti-tumor cell activity of Bufonis Venenum, including 21 identified bufadienolides and 7 unknown compounds. It is noticeable that these bufadienolide markers could not be recognized by traditional HPLC-UV based spectrum-effect relationship analysis (correlation coefficient ranging from -0.24 to 0.40). Finally, we proposed a weight coefficient-based corrected total contents of 9 bufadienolides as a quality evaluation indicator, which had good correlation with inhibitory effects on tumor cells of commercial Bufonis Venenum. The correlation coefficient increased from 0.4 to 0.6. Thus, our pre-fractionation chromatography and mass spectrometry strategy had significant advancement over the traditional spectrum-effect relationship method for chemical marker identification. These results could be crucial and helpful in the development of a quality evaluation method that could reflect the pharmacological activity of Bufonis Venenum.