ABSTRACT
Objective:To explore the effects of alantolactone on cell proliferation and apoptosis of acute myeloid leukemia cell line THP-1 and the related mechanisms.Methods:THP-1 cells in logarithmic growth phase were taken. The control group was added with an equal volume of culture medium containing 10% fetal bovine serum, and the experimental group was added with 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 10.0 μmol/L alantolactone. The cells in each group were cultured for 72 hours. The cell proliferation was detected by CCK-8 method. In addition, the cells of the control group were taken, and the experimental group was added with 2.0 and 4.0 μmol/L alantolactone, respectively. The cells in each group were cultured for 72 hours. The apoptosis of the cells was detected by flow cytometry, and the expressions of NF-κB pathway-related proteins were detected by Western blotting.Results:After THP-1 cells were treated with different concentrations of alantolactone (0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 10.0 μmol/L) for 72 hours, the inhibitory rates were (9.4±1.4)%, (27.7±4.0)%, (45.1±2.5)%, (66.9±2.9)%, (87.0±1.2)%, (91.7±1.0)% and (94.4±0.8)%, respectively, which were in a dose-dependent manner, and the control group was (0.4±0.1)%, the difference was statistically significant ( F = 241.87, P < 0.01). After THP-1 cells were treated with different concentrations of alantolactone (2.0, 4.0 μmol/L) for 72 hours, the apoptotic rates were (26.1±4.2)% and (37.8±5.6)%, and the control group was (8.5±1.4)%, the difference was statistically significant ( F = 43.04, P < 0.05). Furthermore, alantolactone could down-regulate the expression levels of bcl-2 and p65, and up-regulate the expression levels of bax, cleaved-caspase-3 and cleaved-PARP. Conclusions:Alantolactone can induce the apoptosis of AML THP-1 cells via inhibiting the NF-κB pathway, thereby inhibiting its proliferation.
ABSTRACT
@#[Abstract] Objective: To investigate the effect of alantolactone (ALT) on proliferation, migration, invasion and apoptosis of human osteosarcoma 143B cells and the underlying mechanism. Methods: Osteosarcoma 143B cells were treated with different concentrations of ALT (0, 4, 6, 8, 10 µmol/L). Then, the cell proliferation ability was detected by crystal violet staining and MTT assay, cell migration was determined by Wound-healing test, cell invasion was analyzed by Transwell assay and cell apoptosis rate was detected by Hoechst33258 staining. The mRNA and protein expressions of E-cadherin, N-cadherin, caspase-3, cleaved caspase-3 (c-caspase-3), poly ADP-ribose polymerase (PARP) and cleaved PARP (c-PARP) in 143B cells were detected by qPCR and Western blotting (WB), respectively. TCF/LEF (T cell lymphocyte factor/lymphoid enhancer factor) transcriptional activity was examined with Luciferase reporter gene assay. The mRNA and protein expressions of β-catenin as well as MMP-7 and c-Myc were detected by qPCR and WB, respectively. Results: ALT inhibited proliferation, migration and invasion of osteosarcoma143B cells and promoted apoptosis(P<0.05or P<0.01). After the treatment with ALT at 8, 10 µmol/L, the mRNA and protein expressions of E-cadherin and PARP, as well as the protein expressions of c-caspase-3 and c-PARP were up-regulated, while the mRNA and protein expressions of N-cadherin were downregulated (P<0.05 or P<0.01);At the sametime, theTCF/LEF transcriptional activity and the mRNA and protein expressions of β-catenin, MMP-7 and c-Myc were significantly down-regulated (P<0.05 or P<0.01). Conclusion:ALT may inhibit the proliferation, migration and invasion and promote cell apoptosis possibly through suppressing Wnt/β-catenin signaling pathway in osteosarcoma 143B cells.
ABSTRACT
Objective To study the major chemical components from Inula helenium. Methods The compounds were separated and purifid by using a variety of chromatographic techniques including silica gel column chromatography, thin layer chromatography, and high performance liquid chromatography, and the structures of the compounds were verified by nuclear magnetic spectroscopy and literature data. Results A total of 22 compounds were separated from petroleum ether extract of I. helenium and identified separately as alantolactone (1), isoalantolactone (2), 4,4-dimethylsterols (3), 11αH,13-dihydroisoalantolactone (4), 11αH,13-dihydroalantolactone (5), 4(15)-epoxy-isoalantolactone (6), 5α,6α-epoxyalantolactone (7), alloalantolactone (8), isoalloalantolactone (9), friedelin (10), friedelinol (11), erythrodiol (12), β-sitosterol-glucopyranoside (13), lupeol acetate (14), lupeone (15), lupeol (16), δ-amyrin (17), lupeol palitate (18), 5,7,4'-trihydroxy-3',5'-dimethoxy flavane (19), (+)-syringaresinol (20), 3,5,3'-trihydroxy-6,7,4'-trimethoxy flavone (21), and 3,5,6,7,3'-hydroxy-4'-methoxy dihydroflavones (22). Conclusion Compounds 21 and 22 are isolated from this genus for the first time; Compounds 10-15, 17, and 18 are separated from the I. helenium for the first time. After antibacterial test, compounds 1, 2, 4, 5, 7-9 have strong inhibitory effects on Escherichia coli, Staphylococcus aureus, and Bacillus subtilis.
ABSTRACT
Objective:To establish an HPLC gradient elution method for the determination of isoalantolactone,alantolactone,xan-thotoxol,oxypeucedanin,imperatorin and osthole in Kangfu ointment simultaneously. Methods:Isoalantolactone,alantolactone,xan-thotoxol,oxypeucedanin,imperatorin and osthole were determined by HPLC with a chromatographic column of Agilent TC-C18(250 mm ×4.6 mm,5 μm),the mobile phase was methanol-0.1% formic acid solution with gradient elution at a flow rate of 0.9 ml·min-1. The detection wavelengths were 220 nm and 300 nm. The column temperature was 35℃. Results: The linear range of isoalantolac-tone,alantolactone,xanthotoxol,oxypeucedanin,imperatorin and osthole was 6.16-123.20 μg·ml-1,3.78-75.60 μg·ml-1,1.87-37.40 μg·ml-1,4.06-81.20 μg·ml-1,9.27-185.40 μg·ml-1and 13.89-277.80 μg·ml-1,respectively. The correlation coef-ficient was 0.999 4,0.999 8,0.999 6,0.999 5,0.999 9 and 0.999 7, respectively. The average recovery was 98.04% (RSD=1.06%),97.10%(RSD = 1.53%), 96.73% (RSD = 0.90%), 98.92% (RSD = 1.36%), 99.12% (RSD = 0.83%) and 100.27%(RSD=0.58%),respectively. Conclusion:The method is simple and accurate,which can be used to improve the quality standard for Kangfu ointment.
ABSTRACT
Objective To investigate the inhibitory effect of alantolactone on glioma C6 cells and its possible mechanism. Methods Rat glioma cell line C6 cells were cultured in vitro. The effect of AL on the viability of C6 cells was detected by MTT assay. The effect of AL on migration and invasion of C6 cells was assessed by scratch test and Transwell chamber assay. Flow cytometry was used to detect the effect of AL on the induction of apoptosis in C6 cells, and the effects of AL on mitochondrial membrane potential in C6 cells was detected using JC-1 fluorescent probe. Western blot was used to detect the expression of related proteins in C6 cells after AL treatment. Results AL significantly inhibited the proliferation of C6 cells in a time-and dose-dependent manner. After AL treatment for 24 hours, the migration and invasion of C6 cells were significantly inhibited, N-cadherin protein was significantly decreased and E-cadherin protein was significantly increased, the number of apoptotic cells increased obviously while the mitochondrial membrane potential decreased significantly. The protein expressions of PARP/ cleaved caspase-3/cleaved caspase-9 were significantly up-regulated. Conclusions AL inhibits the proliferation of glioma C6 cells by inhibiting their migration and invasion through regulating the protein expression of cadherin and inducing apoptosis in C6 cells by regulating the cytochrome C/caspase signaling pathway.
ABSTRACT
OBJECTIVE:To establish the method for simultaneous determination of alantolactone,isoalantolactone,gallic acid,emodin,aloe-emodine,rhein,physcion and chrysophanol in Liuwei nengxiao pills.METHODS:HPLC method was adopted.The determination was performed on Diamonsil C18 with mobile phase consisted of methanol-acetonitrile-0.1％ glacial acetic acid (gradient elution) at the flow rate of 1.0 mL/min.The detection wavelengths were set at 254 nm (alantolactone,isoalantolactone,emodin,aloe-emodine,rhein,physcion and chrysophanol),270 nm (gallic acid).The column temperature was 25 ℃,and sample size was 10 μ L.RESULTS:The linear ranges of alantolactone,isoalantolactone,gallic acid,emodin,aloe-emodine,rhein,physcion,chrysosphanol were 0.121-3.63 μg(r=0.999 9),0.122-3.66 μg(r=0.999 9),0.219-6.57 μg(r=0.999 9),0.016 4-0.492 μg(r=0.999 7),0.017 3-0.519 μg(r=0.999 9),0.015 3-0.459 μg(r=0.999 9),0.007 2-0.216 μg(r=0.999 9),0.016 2-0.486(r=0.999 9).The limits of quantification were 0.41,0.26,0.35,0.13,0.17,0.14,0.15,0.13 ng;limits of detection were 0.12,0.08,0.11,0.04,0.05,0.04,0.05,0.04 ng.RSDs of precision,stability and reproducibility tests were all lower than 2.0％.The recoveries were 98.05％-102.46％ (RSD=1.75 ％,n=6),98.55％-102.89％ (RSD=1.91％,n=6),98.53 ％-102.34％ (RSD=1.66％,n=6),101.71％-103.41％ (RSD =0.57 ％,n=6),101.04％-103.01％ (RSD=0.69％,n=6),101.63％-102.75％ (RSD=0.39 ％,n=6),96.94％-101.11％ (RSD=1.61％,n=6),98.06％-99.10％ (RSD=0.40％,n=6).CONCLUSIONS:The method is accurate,simple and suitable for simultaneous determination of 8 components in Liuwei nengxiao pills.
ABSTRACT
Objective To investigate the effects of alantolactone on cell proliferation,cell-cycle and cell cycle-related proteins in human chronic myelogenous leukemia drug-resistant cell line K562/ADR.Methods K562/ADR cells were treated with 0,1.0,2.0,4.0,6.0,8.0,and 10.0 μmol/L of alantolactone for 12,24 and 48 h,with its cell viability analyzed by MTT assay.Flow cytometry was used to examine the effect of alantolactone on the cell-cycle of K562/ADR cells.The cell cycle-related proteins were analyzed by using Western blot after treatment with alantolactone.Results The results of MTT showed that alantolactone effectively inhibited the proliferation of K562/ADR cells in dose and time-dependent way,and the IC50 value of alantolactone in K562/ADR cells was about 5 μmol/L.Flow cytometric analysis displayed that alantolactone could arrest cell cycle at G2/M phase.The percentage of accumulated cells in the G2/M phase was increased from (15.8±1.7) ％ in the control group to (21.0±2.4) ％,(26.4±2.7) ％,and (30.1±3.9) ％ in cells treated with 2.5,5.0,and 7.5 μmol/L of alantolactone for 24 h,respectively (P ＜ 0.05).Alantolactone significantly decreased the expression of CDK1 and CyclinB1 and increased the expression of cyclin-dependent kinase inhibitor p21.Meanwhile,the treatment of K562/ADR with alantolactone led to a dose-dependent decrease in bcr-abl protein levels.Conclusion Alantolactone can significantly inhibit the proliferation and cell-cycle arrest in G2/M phase of K562/ADR cells,in which mechanism may be associated with the regulation of cell cycle-related proteins and downregulation of bcr-abl protein.
ABSTRACT
Objective To establish a method to simultaneously determine contents of alantolactone and isoalantolactone in Sanwei Ganlusan. Methods Contents of alantolactone and isoalantolactone was determined by HPLC was used with PDAD detector;the column was CAPCELL PAK MG C18 (4.6 mm×250 mm, 5μm);the mobile phase consisted of acetonitrile-0.4%phosphoric (58∶42);the flow rate was 1.0 mL/min;the detection wavelength was set at 225 nm;the column temperature was maintained at 30 ℃. Results The linear range of alantolactone was 0.083-0.517 μg (r=0.999 9), and isoalantolactone was 0.108-0.672 μg (r=0.999 9). The mean recovery of alantolactone was 97.95%, RSD=1.31%. The mean recovery of isoalantolactone was 97.69%, RSD=1.24%. Conclusion The method is accurate and simple in operation, which can be used to simultaneously determine contents of alantolactone and isoalantolactone in Sanwei Ganlusan.
ABSTRACT
Abstract: In this article, we report a study of assay of sesquiterpene lactones (alantolactone, isoalantolactone) in plant extraction derived by ultrasound-assisted extraction, оrthogonal test design and reflux extraction from medicinal plant’s composition (Salsola laricifolia turcz.e litv+Inula helenium). High-performance liquid chromatography (HPLC) method was used for determination of the contents of alantolactone and isoalantolactone in the investigated extracts. The result shown that the amount of alantolactone was 0.64±0.03%, and isoalantolactone 0.59±0.01% in the plant extraction derived by reflux condensation extraction. Key words: Alantolactone, Isoalantolactone, HPLC, Salsola laricifolia turcz.ex litv, Inula helenium, Reflux method, Ultrasound- assisted extraction.
ABSTRACT
Objective To establish the quality standard for Tibetan medicine MNXT granule.Methods Inula racemosa,Tinospora sinensis were identified by TLC; Isoalantolactone and alantolactone were determined by HPLC.Results Inula racemosa,Tinospora sinensis could be identified by TLC.The concentration of isoalantolactone was linear in the range of 0.281～0.842 μg,r=0.9998,with the average recovery rate being 98.5％,RSD being1.14％.The concentration of alantolactone was linear in the range of 0.232～0.696 μg,r=0.9999,with the average recovery rate being 97.4％,RSD being 1.10％.Conclusion The method was simple,accurate,repeatable and able to control the quality of preparation effectively.
ABSTRACT
OBJECTIVE: To optimize the supercritical CO2 extraction technology of volatile component from Tibet inual and Zingiber officianle. METHODS: The extraction technology was optimized by orthogonal experiment with the content of alantolactone and the yield of volatile oil as indexes while with extracting pressure, extracting temperature, extracting time and entrainer volume as factors. RESULTS: The optimal extraction technology was as follows: 225 mL entrainer was extracted at 40 ℃ with extracting pressure of 35 Mpa for 1 hour. CONCLUSION: The extraction technology is reasonable, practical and controllable for the industrial production.