Chemical constituents from Kalanchoe hybrida and their cytotoxicity / 中草药·英文版

Objective: Kalanchoe hybrida (Crassulaceae) is naturalized throughout all the island of Taiwan in China. The preliminary bioassay-guided fractionation of the crude extract of K. hybrida exhibited that the chloroform and n-butanol fractions possessed potent cytotoxicity against MCF-7, NCI-H460, and SF-268 tumor cell lines at 50 µg/mL concentration. Therefore, K. hybrida was selected as a target and the chemical constituents from the chloroform and n-butanol fractions of the crude extracts of K. hybrida were identified. The potential constituents were examined for their cytotoxicity against the tumor cell lines. Methods: A combination of conventional chromatographic techniques was performed on the crude extract of K. hybrida. The chemical structures of the purified constituents were identified on the basis of spectroscopic and spectrometric analysis. Results: The purification results had led to the characterization of totally 37 compounds. The isolated compounds 1, 2, and 4–12 were examined for their cytotoxicity in vitro, and bufadienolides 4–8 and flavonol glycoside 11 displayed significant cytotoxicity towards all the tested tumor cell lines among these tested compounds. Conclusion: The results indicated that these principles should be responsible for the bioactivity of corresponding partial fractions. The potential constituents could be further investigated to explore the new natural lead drugs.

Research progress on chemical constituents and pharmacological activities of Bufonis Venenum / 中草药

Bufonis Venenum, as an important Chinese traditional natural medicine, has complex chemical composition and has been widely used in clinical treatment with significant effects. The chemical constituents in Bufonis Venenum mainly included bufadienolides, indole alkaloids, steriods, etc. With the further research on Bufonis Venenum both at home and abroad, its chemical composition and mechanism of pharmacological activities have been more and more clearly, especially in the aspects of narcotic analgesic, cardiac, and antitumor therapy. This paper reviewed the chemical composition and pharmacology advances of Bufonis Venenum in order to provide reference to clinical use.

Simultaneous Determination of 9 Components of Bufadienolide in Liushen Pills by HPLC / 中国药房

OBJECTIVE:To establish a method for simultaneous determination of gamabufotalin,arenobufagin,telocinobufa-gin,desacetylcinobufotalin,bufotalin,cinobufotalin,bufalin,cinobufagin and resibufogenin in Liushen pills. METHODS:HPLC method was adopted. The determination was performed on ODS-2 C18 column with mobile phase consisted of acetonitrile-0.15%phosphoric acid(gradient elution)at the flow rate of 1.0 mL/min. The detection wavelength was set at 296 nm,and column temper-ature was 40 ℃. The sample size was 10 μL. RESULTS:The linear ranges of gamabufotalin,arenobufagin,telocinobufagin,de-sacetylcinobufotalin, bufotalin, cinobufotalin,bufalin, cinobufagin and resibufogenin were 1.10-70.39 μg/mL(r=0.9996), 4.03-257.78 μg/mL(r=0.9999),4.09-261.89 μg/mL(r=0.9999),0.67-42.96 μg/mL(r=0.9999),3.36-214.73 μg/mL(r=0.9999), 5.73-366.44 μg/mL(r=0.9999),3.77-241.56 μg/mL(r=0.9999),7.31-468.11 μg/mL(r=0.9999),5.18-331.56 μg/mL(r=0.9999). The limits of quantitation were 1.10,0.85,1.02,0.34,0.84,1.43,0.94,3.66,2.59 μg/mL;the limits of detection were 0.27, 0.21,0.51,0.17,0.42,0.72,0.47,0.91,1.30 μg/mL,respectively. RSDs of precision,stability and reproducibility tests were all lower than 3.0%. The recoveries were 96.35%-103.10%(RSD=2.72%,n=6), 96.76%-103.24%(RSD=2.49%,n=6), 97.01%-101.39%(RSD=1.64%,n=6),97.32%-104.01%(RSD=2.61%,n=6),95.76%-103.60%(RSD=2.92%,n=6), 95.07%-102.59%(RSD=2.92%,n=6),95.77%-101.43%(RSD=2.03%,n=6),95.11%-103.72%(RSD=3.19%,n=6), 95.23%-103.34%(RSD=3.24%,n=6),respectively. CONCLUSIONS:The method is simple,rapid,accurate,reliable and can be used for the determination of bufadienolide in Liushen pills .

Antiplasmodial and Cytotoxic Activities of Toad Venoms from Southern Amazon, Brazil

The drug-resistance of malaria parasites is the main problem in the disease control. The huge Brazilian biodiversity promotes the search for new compounds, where the animal kingdom is proving to be a promising source of bioactive compounds. The main objective of this study was to evaluate the antiplasmodial and cytotoxic activity of the compounds obtained from the toad venoms of Brazilian Amazon. Toad venoms were collected from the secretion of Rhinella marina and Rhaebo guttatus in Mato Grosso State, Brazil. The powder was extracted at room temperature, yielding 2 extracts (RG and RM) and a substance ('1') identified as a bufadienolide, named telocinobufagin. Growth inhibition, intraerythrocytic development, and parasite morphology were evaluated in culture by microscopic observations of Giemsa-stained thin blood films. Cytotoxicity was determined against HepG2 and BGM cells by MTT and neutral red assays. The 2 extracts and the pure substance ('1') tested were active against chloroquine-resistant Plasmodium falciparum strain, demonstrating lower IC₅₀ values. In cytotoxic tests, the 2 extracts and substance '1' showed pronounced lethal effects on chloroquine-resistant P. faciparum strain and low cytotoxic effect, highlighting toad parotoid gland secretions as a promising source of novel lead antiplasmodial compounds.

First serine protease inhibitor isolated from Rhinella schneideri poison

Background Toad secretions are a source of molecules with potential biotechnological application on a wide spectrum of diseases. Toads from theRhinella family have two kinds of poisonous glands, namely granular and mucous glands. Rhinella schneideritoads produce granular secretions that comprise a great number of molecules, including serine proteases inhibitors. Serine proteases, such as trypsin, chymotrypsin and elastase, are enzymes that have a serine amino acid into its catalytic site and can be found in a large number of vertebrate species and pathogenic microorganisms. Therefore, the present work aims to purify a serine protease inhibitor from Rhinella schneiderigranular secretions.Findings This study presents the protocol used to purify a serine protease inhibitor from the Rhinella schneideri poison. The granular secretion was submitted to dialysis in order to separate the low molecular weight compounds, which were submitted to a reversed phase-fast protein liquid chromatography fractionation step in a C2C18 column. The major fractions were tested over trypsin, chymotrypsin and elastase through colorimetric assay. The inhibition tests were performed with the enzyme in absence (positive control) and presence of fractions, denatured enzyme (negative control) and the respective chromogenic substrate. Rs20 was the compound with the major inhibitory activity over chymotrypsin, inducing a delay in the formation of the chromogenic enzymatic product. The structure characterization of Rs20 was performed by high resolution electronspray ionization-mass spectrometry (HRESI-MS) and gas chromatography coupled with mass spectrometry (GC-MS). HRESI showed an intense signal suggesting the presence of bufadienolide with less than 10 ppm error. In addition, it was observed a low intense signal at m/z399 that could be lithocholic acid, a biosynthetic precursor of bufadienolide. Finally, GC-MS analysis applying NIST library identification reinforced this hypothesis.Conclusions The current study have isolated and partially characterized the function and structure of the first bufadienolide with inhibitory action over chymotrypsin.

First serine protease inhibitor isolated from Rhinella schneideri poison

Background Toad secretions are a source of molecules with potential biotechnological application on a wide spectrum of diseases. Toads from theRhinella family have two kinds of poisonous glands, namely granular and mucous glands. Rhinella schneideritoads produce granular secretions that comprise a great number of molecules, including serine proteases inhibitors. Serine proteases, such as trypsin, chymotrypsin and elastase, are enzymes that have a serine amino acid into its catalytic site and can be found in a large number of vertebrate species and pathogenic microorganisms. Therefore, the present work aims to purify a serine protease inhibitor from Rhinella schneiderigranular secretions.Findings This study presents the protocol used to purify a serine protease inhibitor from the Rhinella schneideri poison. The granular secretion was submitted to dialysis in order to separate the low molecular weight compounds, which were submitted to a reversed phase-fast protein liquid chromatography fractionation step in a C2C18 column. The major fractions were tested over trypsin, chymotrypsin and elastase through colorimetric assay. The inhibition tests were performed with the enzyme in absence (positive control) and presence of fractions, denatured enzyme (negative control) and the respective chromogenic substrate. Rs20 was the compound with the major inhibitory activity over chymotrypsin, inducing a delay in the formation of the chromogenic enzymatic product. The structure characterization of Rs20 was performed by high resolution electronspray ionization-mass spectrometry (HRESI-MS) and gas chromatography coupled with mass spectrometry (GC-MS). HRESI showed an intense signal suggesting the presence of bufadienolide with less than 10 ppm error. In addition, it was observed a low intense signal at m/z399 that could be lithocholic acid, a biosynthetic precursor of bufadienolide. Finally, GC-MS analysis applying NIST library identification reinforced this hypothesis.Conclusions The current study have isolated and partially characterized the function and structure of the first bufadienolide with inhibitory action over chymotrypsin.(AU)

A new bufadienolide with cytotoxic activity from the Chinese traditional drug Ch'an Su / 中国天然药物

AIM@#To study the bufadienolides in the Chinese traditional drug "Ch'an Su" and their cytotoxic activity.@*METHOD@#Various chromatographic techniques were used to isolate the constituents, and their structures were elucidated through physical and spectroscopic data.@*RESULTS@#Twenty compounds were isolated, and eighteen were evaluated in vitro for their cytotoxic activity against A-549 and K-562 cells.@*CONCLUSION@#Compound 1 (bufalin 3β-acrylic ester) was a new bufadienolide and exhibited the most potent activity against the two tumor cell lines with IC50 values of 7.16 and 6.83 nmol · L(-1). The relationships between structure and activity are discussed.

A new compound from roots and rhizomes of Helleborus thibetanus / 中草药

Objective: To study the chemical constituents in the ethanol extract from the roots and rhizomes of Helleborus thibetanus. Methods: The compounds were isolated and purified by silica gel column chromatography and recrystallization. The structures of the compounds were identified on the basis of chemical and spectral methods. Results: A new compound was isolated from the ethanol extract in the roots and rhizomes of H. thibetanus and identified as 14β-hydroxy-3β - [(β-D-glucopyranosyl)oxy]-5α-bufa-20, 22-dienolide (1). Conclusion: Compound 1 is a new compound named as hellebocoside A, which belongs to bufadienolide of cardiac glycosides.