Staurosporine induced cell death was dose dependent with an EC50 of 130 nM. A staurosporine concentration of 500 nM induced selleck chemicals maximum cell death and was therefore used for all subsequent Inhibitors,Modulators,Libraries experiments. The cells were exposed to herbal extracts, staurosporine or, simultaneously, to staurosporine and herbal extracts. Half of the culture media were replaced with new media containing double the final concentrations of staurosporine and or the herbal extracts. The cells were incubated at 37 C, 5% CO2 for 24 h. Toxicity was deter mined by measuring the release of lactate dehydrogenase into the culture medium using the LDH cytotoxicity detection kit. The percent age of cytotoxicity was calculated using the formula A absorbance measured. Cmin absorbance of negative control, Cmax absorb ance of maximum toxicity.
Non cellular antioxidant capacity assays DPPH radical scavenging assay, oxygen radical absorbance capacity assay and total phenol Inhibitors,Modulators,Libraries assay by Folin Ciocalteau reagent were performed as described in detail previously. LC ESI MS analyses Chemical fingerprints of the Inhibitors,Modulators,Libraries re extracted herbal extracts in 80% methanol were obtained using liquid chromatog raphy electrospray ionization mass spectrometry analyses. LC ESI MS experi ments were performed on a Waters Acquity Xevo TQ triple quadrupole mass spectrometer coupled to a binary pump, and an autosampler. Separation was achieved using a Waters Acquity Xevo TQ triple quadrupole mass spectrometer coupled to an ultra per formance liquid chromatography binary pump, a photo diode array detector, and an autosampler.
Separation was achieved on Acquity UPLC BEH C 18 column attached with a Vanguard BEH C18 at a mobile phase flow rate of 0. 3 ml min, operating at room temperature. The mobile phase consisted of 0. 1% aqueous Inhibitors,Modulators,Libraries formic acid and aceto nitrile. The gradient elution was used with a starting mobile phase composition of 20%, increasing to 100% B for 16 min. Mass spectra were acquired in both positive and negative ionization mode using an ESI source with the mass recorded in the range of m z 50 1000. Statistical analysis One way ANOVA tests were carried out with GraphPad Prism. Data from triplicate cultures in 2 3 separate experiments are presented as mean SEM. Data comparison between drug treated and untreated groups were made by Dunnett post tests. A p value 0. 05 was considered statistically significant and denoted by asterisks.
The relationships between antioxidant cap acity and cytotoxicity data were assessed by calculation of Pearsons Inhibitors,Modulators,Libraries correlation selelck kinase inhibitor coefficients. Results and discussion Some herbal extracts exhibit toxic effects in cultured primary cortical neurons First, we determined whether the herbal extracts exerted toxic effects in cultured mouse cortical neurons. The results demonstrated that most of the herbal extracts showed negligible cytotoxic effects when compared to the positive control of staurosporine induced cytotoxicity.