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Wu X.,Chinese Institute of Materia Medica | Si N.,Chinese Institute of Materia Medica | Bo G.,Anhui Jinchan Biochemistry Co. | Hu H.,University of Macau | And 4 more authors.
Biomedical Chromatography | Year: 2015

Cinobufacini injection that comes from the water extract of Bufo bufo gargarizans Cantor skin is widely used for cancer treatment in China. Peptide is one of its major types of constituents, however the biological effects and content of this injection are little reported. In present study, the analgesic effect of peptides was determined and evaluated by in-vivo models. To characterize and quantitatively analyze these peptides, a reliable and efficient method combining size exclusion chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry with amino acid analysis was developed. The peptides presented as a series of analogs with similar molecular weights mostly ranging from 2 to 8 kDa. The amino acid analysis by gas chromatography mass spectrometry (GC-MS) was developed to determine both free and combined amino acids (FAA and CAA) in cinobufacini injection. This method achieved good linearity (R2, 0.9909-0.9999) and low limit of detection and quantification. FAA and CAA samples were efficiently analyzed by modified Phenomenex EZ: faast procedure. For the sample analysis, the method showed good repeatability (relative standard deviation, RSD≤10%). For most FAA and CAA the mean recoveries were >80% with RSD <10%. The GC-MS based method is useful for quality assurance of both FAA and CAA in cinobufacini injection. © 2014 John Wiley & Sons, Ltd.

Wu X.,Chinese Institute of Materia Medica | Zhao H.,Chinese Institute of Materia Medica | Wang H.,Chinese Institute of Materia Medica | Gao B.,Anhui Jinchan Biochemistry Co. | And 3 more authors.
Journal of Separation Science | Year: 2012

A rapid and validated method was established for the simultaneous determination of eight active and toxic bufadienolides in cinobufacini injection using high performance liquid chromatography coupled with triple quadrupole mass spectrometry. These eight compounds were separated within 3 min on a C 18 analytical column with gradient elution. Eleven batches of cinobufacini injection were analyzed with good linear regression relationship (r, 0.9979-0.9999), precisions (RSD, 1.92-4.79%), repeatability (RSD, 3.12-4.96%), stability (RSD, 2.84-4.45%), and recovery (93.96-104.89%). By using the established method, the present study offered highly sensitive, specific, and speedy determination of eight bufadienolides, which promoted the quality control investigation of cinobufacini injection greatly. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Ren W.,Chinese Institute of Materia Medica | Ren W.,Capital Medical University | Zuo R.,Li Kang Hospital | Wang Y.-N.,Capital Medical University | And 12 more authors.
PLoS ONE | Year: 2016

Huang-Lian-Jie-Du Decoction (HLJDD) is a classical Traditional Chinese Medicine (TCM) formula with heat-dissipating and detoxifying effects. It is used to treat inflammation-associated diseases. However, no systematic pharmacokinetic (PK) and pharmacodynamic (PD) data concerning the activity of HLJDD under inflammatory conditions is available to date. In the present study, the concentration-time profiles and the hepatic clearance rates (HCR) of 41 major components in rat plasma in response to the oral administration of a clinical dose of HLJDD were investigated by LC-QqQ-MS using a dynamic multiple reaction monitoring (DMRM) method. Additionally, the levels of 7 cytokines (CKs) in the plasma and the body temperature of rats were analyzed. Furthermore, a PK-PD model was established to describe the time course of the hemodynamic and anti-inflammatory effects of HLJDD. As one of the three major active constituents in HLJDD, iridoids were absorbed and eliminated more easily and quickly than alkaloids and flavonoids. Compared with the normal controls, the flavonoids, alkaloids and iridoids in inflamed rats exhibited consistently changing trends of PK behaviors, such as higher bioavailability, slower elimination, delays in reaching the maximum concentration (Tmax) and longer substantivity. The HCR of iridoids was different from that of alkaloids and flavonoids in inflamed rats. Furthermore, excellent pharmacodynamic effects of HLJDD were observed in inflamed rats. The levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β, IL-10, and macrophage inflammatory protein-2 (MIP-2) and body temperature significantly decreased after the administration of HLJDD. Based on PK-PD modeling with the three-phase synchronous characterization of time-concentration-effect, flavonoids exhibited one mechanism of action in the anti-inflammatory process, while iridoids and alkaloids showed another mechanism of action. Taken together, the results demonstrated that HLJDD may restrain inflammation synergistically via its major constituents (alkaloids, flavonoids and iridoids). A correlation between the exposure concentration of different types of compounds and their anti-inflammatory effects in the body was shown. This study provides a comprehensive understanding of the anti-inflammatory activity of HLJDD. © 2016 Ren et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Zhao H.,Chinese Institute of Materia Medica | Wu X.,Chinese Institute of Materia Medica | Wang H.,Chinese Institute of Materia Medica | Gao B.,Anhui Jinchan Biochemistry Co. | And 3 more authors.
Journal of Liquid Chromatography and Related Technologies | Year: 2014

Chan Pi (the toad skin) is an effective Chinese medicine in the treatment of boils, pyogenic infections, and cancer in clinical settings. However, the inhomogeneity of its quality is often observed due to confusing sources, namely Bufo bufo gargarizans Cantor and Bufo melanostictus Schneider. In the present paper, a simultaneous determination method of eight bufadienolides in two species of Chan Pi was established using HPLC coupled with triple quadrupole mass spectrometry (HPLC-QqQ MS). Eight bufadienolides in 7 batches of B. bufo gargarizans skin and 8 batches of B. melanostictus skin were analyzed with an acceptable linear regression relationship (r ≥ 0.9990), precisions (RSD, 1.97-4.38%), repeatability (RSD, 2.65-4.70%), stability (RSD, 2.39-4.51%), and recovery (96.39-103.82%). Additionally, cluster analysis was used for the successful discrimination of these two species of toad skin by the ratio of these eight bufadienolides. The present study greatly facilitates clarification of the difference between these two species of Chan Pi, which improves the quality control of this traditional Chinese medicine. © 2014 Copyright Taylor and Francis Group, LLC.

Zhao H.,Chinese Institute of Materia Medica | Wu X.,Chinese Institute of Materia Medica | Wang H.,Chinese Institute of Materia Medica | Gao B.,Anhui Jinchan Biochemistry Co. | And 3 more authors.
Journal of Separation Science | Year: 2013

Cinobufacini injection, prepared from the skin of Bufo bufo gargarizans Cantor, has presented its significant effects on the treatment of hepatitis and various cancers in the clinic. However, as an unclear complex chemical system, the optimization of its quality control markers has been a long-term challenge. In present study, a feasible strategy integrated markers screening, determination, and statistical analysis was efficiently proposed, especially for the undefined Chinese medicine injections. First, rapid separation LC-quadrupole-TOF-MS method was applied in the identification of 19 major compounds in the cinobufacini injection for the first time. Further, nine high-level contents active compounds were selected as quality control markers for the quantification analysis. An acceptable and validated determination method was established in 17 batches of cinobufacini injection by HPLC-photodiode array detection method, including linear regression relationship (r2, 0.9996-1), precisions (RSD, 0.02-1.35%), repeatability (RSD, 0.05-1.97%), stability (RSD, 0.1-3.85%), and recovery (95.88-104.89%). Each analyte was detected at its maximum ultraviolet spectra wavelength. Finally, based on the quantification results, principal component analysis was performed on the quality assessment of cinobufacini injections. This three-step strategy provides a newly feasible solution for the quality control of Chinese medicine injections. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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