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Objective To study the effects of four cytotoxicity evaluation methods on the inhibition rate of shikonin (SK) in vitro, and to compare the pseudo-negative phenomenon often found in the evaluation of cytotoxic activity of natural pigments represented by naphthoquinones in Lithospermum erythrorhizon. Methods SK was co-cultured with its high-sensitive strain HL-60 cells and low-sensitivity strain A549 cells, trypan blue method, sulforhodamine B (SRB) method, Cell Counting Kit-8 (CCK-8) and MTT cytotoxicity test were used for parallel experiments to determine the dose-effect relationship curve of SK (0.4-128 μmol/L) inhibiting the growth of cells. Results The half-inhibitory concentration (IC50) of shikonin on HL-60 cells was determined by trypan blue method, SRB method, CCK-8 method, and MTT method, which was 0.57, 0.77, 1.36, and 1.01 μmol/L, respectively. For A549 cells, the IC50 was 6.30, 10.38, 13.48, and 15.24 μmol/L, respectively. When the concentration of shikonin was below 3.2 μmol/L and 32 μmol/L, the inhibition rate of the two kinds of cells increased linearly by the four methods, followed by differences. Among them, the results of the trypan blue method and the SRB method are in good agreement, while the MTT method and the CCK-8 method have lower inhibition rates. At 12.8 μmol/L, the inhibitory rate of SK on HL-60 cell measured by CCK-8 was 81%, while the inhibitory rate measured by trypan blue method was 96%, and at 128 μmol/L the inhibitory rate of SK on A549 cells measured by MTT method was 89%, however, the inhibitory rate measured by trypan blue method was 99%. The absorption spectrum of SK overlapped with formazan at the wavelength from 400 to 600 nm, with the maximum overlap peak from 550 to 570 nm, and CCK-8 reagent had a synergistic inhibitory effect on HL-60 with SK. The results of trypan blue method showed that SK at the highest dose almost completely killed cells in the plate wells, which was significantly different from the control group, but both MTT method and CCK-8 method resulted in a pseudo-negative phenomenon. Conclusion Therefore, cytotoxicity test of natural pigments represented by naphthoquinones in L. erythrorhizon, MTT method, and CCK-8 method are not recommended, while SRB method and trypan blue method are suggested.
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Background: The current research was undertaken on dried fruits of Capparis moonii to screen its potential for immunomodulatory and cancer indications with identification of phytoconstituents by chromatographic techniques.Methods: Methanolic (MECN), hydro-methanolic (HMECN) and aqueous extracts (AQCN) of Capparis moonii were subjected to high performance thin layer chromatography (HPTLC) and high-performance liquid chromatography (HPLC) after studying the total phenolic and flavonoid content by using rutin and gallic acid as standards respectively as well as undertaking powder characteristics and preliminary phytochemical screening. Immunomodulatory activities covered were hemagglutination antibody titre and delayed-type hypersensitivity reaction with the aid of sheep red blood cells (0.5×109) as antigens. The extracts were studied for antioxidant potential. Anticancer prospects were focusing on in vitro cell lines screening (MCF 7 and HCT 15) by Sulforhodamine B assay method and potato disc assay.Results: The total phenolic and flavonoid content of MECM, HMECM and AQCM fruits extracts were found to be 0.20, 0.11 and 0.47 mg of gallic acid/g and 78.3, 18.8 and 64.4 mg of rutin/g respectively. Rutin and quercetin were confirmed by HPTLC and HPLC showing well resolved peaks. IC50 values in antioxidant studies were found to be significant with all the extracts. Significant immunomodulatory effect was noticed at 200mg/kg in both models (high antibody titre levels and decrease paw volume after 48 h). Unsatisfactory results were observed with selected cell lines and disc assay.Conclusions: Thus, selected fruits may probably have immunomodulatory potential due to presence of flavonols (rutin and quercetin).
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OBJECTIVE To evaluate the effect of Guizhi Fuling Capsule active pharmaceutical ingredient (API) and its fractions on human breast cancer cells proliferation by high-throughput screening assay. METHODS The crude fractions were obtained from the extraction and elution of the API of Guizhi Fuling Capsule, and 929 standard fractions were obtained by the optimal separation conditions. Sulforhodamine B (SRB) method was used to evaluate the effects of the Guizhi Fuling capsule API and 929 kinds of fractions on the proliferation of human breast cancer cells MCF- 7 and MDA- MB- 231. RESULTS The Guizhi Fuling capsule API had a strong ability to inhibit the proliferation of MCF-7 cells at high concentration and the ability to inhibit MDA-MB-231 cells' proliferate at low concentration follow?ing 72 h treatment;some samples of 929 fractions (5 μg·mL-1) was found to have a breast cancer cell growth inhibition rate above 50%, without toxicity on HUVECs proliferation. CONCLUSION The API of Guizhi Fuling capsule had significant cytotoxicity effects on these two human breast cancer cells, with significant concentration- and time-dependent manner.
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Objective: To examine the cytotoxic properties of both the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cervical cancer, human breast cancer, human colon cancer and human lung cancer cell lines.Methods:kenaf seed oil on human cancer cell lines was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and sulforhodamine B assays. Cell morphological changes were observed by using an inverted light microscope.Results:The in vitro cytotoxic activity of the kenaf (Hibiscus cannabinus L.) seed extract and cancer cell lines. Morphological alterations in the cell lines after KSE and KSO treatment were observed. KSE and KSO possessed effective cytotoxic activities against all the cell lines been selected.Conclusions:KSE and KSO could be potential sources of natural anti-cancer agents. Further The kenaf seed extract (KSE) exhibited a lower IC50 than kenaf seed oil (KSO) in all of the investigations on using kenaf seeds for anti-proliferative properties are warranted.
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<p><b>OBJECTIVE</b>To examine the cytotoxic properties of both the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cervical cancer, human breast cancer, human colon cancer and human lung cancer cell lines.</p><p><b>METHODS</b>The in vitro cytotoxic activity of the kenaf (Hibiscus cannabinus L.) seed extract and kenaf seed oil on human cancer cell lines was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and sulforhodamine B assays. Cell morphological changes were observed by using an inverted light microscope.</p><p><b>RESULTS</b>The kenaf seed extract (KSE) exhibited a lower IC50 than kenaf seed oil (KSO) in all of the cancer cell lines. Morphological alterations in the cell lines after KSE and KSO treatment were observed. KSE and KSO possessed effective cytotoxic activities against all the cell lines been selected.</p><p><b>CONCLUSIONS</b>KSE and KSO could be potential sources of natural anti-cancer agents. Further investigations on using kenaf seeds for anti-proliferative properties are warranted.</p>
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OBJECTIVE: To screen anti-tumour compounds in vitro, various solvents may be used to improve the solubility of the insoluble compounds in water. This article analyzes and selects the appropriate solubilizers from 19 single solvent and mixtures of solvents via study their cytotoxicity on typical nine categories of human tumour cell lines, and thus provides a reference for testing those compounds with low-water solubility. METHODS: Sulforhodamine B (SRB) assay was performed on human tumor cells inhibition including A549(non-small cell lung), HCT116(Colon), 786-0(Renal), OVCAR-3(Ovarian), K562(Leukemia), SF295(CNS), DU-145(Prostate), MCF7(Breast) and UACC-257(Melanoma). Nineteen single or mixtures of solvents were investigated including PEG400, Tween 20, Tween 80, ethanol, glycerol, Cremophor EL, DMSO and the twelve mixtures with DMSO in the proportions of 1:1(V:V) and 1:9(V:V). RESULTS: When the cell percentage growth is greater than 90%, it is deemed as that this solvent had no inhibitory effect on cell growth. When the cell percentage growth is between 80% and 90%, it is considered as a weak inhibition. When cell percentage growth is less than 80%, it is defined as strong inhibition solvent. Furthermore, weak or no inhibition of the solvent system is recommended for the purpose of high throughput screening of compounds against human tumor cell lines. In this study, we found that final DMSO volume fraction at 0.1%(V:V) had weak or no inhibition to all tested cell lines. Beyond that, PEG400 and ethanol at final concentration of 0.25%(V:V), glycerol at final concentration of 0.5%(V/V) had weak inhibition to all tested cell lines. PEG400, ethanol and glycerol mixed with DMSO with a 1:1(V/V) ratio had weak inhibition to all tested cell lines at final concentration of 0.25%(V:V). However, PEG400 and DMSO mixture(1:9=V:V) at final concentration of 0.1%(V:V), ethanol and glycerol mixed with DMSO with a 1:9 ratio at 0.25% final concentration had weak inhibition to all tested tumor cell lines. All Tween 20, Tween 80, Cremophor EL and their mixtures of DMSO at 1:1(V:V) are clearly cytotoxic to all tested cell lines even at the lowest final concentration of 0.1%. It is appreciate that Cremophor EL/DMSO, at 0.1% final concentration with a 1:9 ratio is acceptable for A549(non-small cell lung tumour), 786-0(Renal), OVCAR-3(Ovarian), SF295(CNS), DU-145(Prostate) and UACC-257(Melanoma) cell lines. CONCLUSION: Ethanol, glycerol, PEG400 and their mixtures with DMSO at 1:1(V:V) or 1:9(V:V) are recommended with a final volume percentage of not more than 0.5%; DMSO should be used with a final concentration of less than 0.25%(V:V); Tween 20, Tween 80, Cremophor EL and their mixtures with DMSO are not recommended. It should be noted that the tested cell lines are capable of resisting to the different solvents. As described above, human leukemia cell line K562 is particularly sensitive to all tested solvents, breast tumour cell MCF7 is especially sensitive to DMSO.