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Clitoria ternatea flower is a traditional medicinal herb that has been used as a natural food colourant. As there are limited studies on investigating the bioactivities of the anthocyanin-rich fraction of Clitoria ternatea flower, this study aimed to determine an efficient column chromatography method to obtain the anthocyanin-rich fraction from this flower and characterise its composition, antioxidant, antibacterial, and cytotoxic activities. Amberlite XAD-16 column chromatography was more efficient in enriching the total anthocyanin content (TAC) of the fraction with the highest TAC to total phenolic content (TPC) ratio of 1:6 than that using C18-OPN. A total of 11 ternatin anthocyanins were characterised in the anthocyanin-rich fraction by LC–MS analysis. The antioxidant activity of the anthocyanin-rich fraction was more potent in the chemical-based assay with an IC50 value of 0.86 ± 0.07 mg/mL using 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay than cellular antioxidant assay using RAW 264.7 macrophages. In vitro cytotoxicity assay using human embryonic kidney HEK-293 cell line showed the anthocyanin-rich fraction to be more toxic than the crude extracts. The anthocyanin-rich fraction had more potent antibacterial activity than the crude extracts against Bacillus cereus, Bacillus subtilis, and Escherichia coli. The anthocyanin-rich fraction of C. ternatea has the potential to be used and developed as a functional food ingredient or nutraceutical agent.

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Different extraction methods (organic solvent and water extraction with the assistance of heat or ultrasound) were investigated for their effects on the phytochemical profile and antioxidant activity of Clitoria ternatea flowers. For solvent extraction (30%, 50%, 80%, or 100% methanol, ethanol, or acetone), 50% ethanol was considered to be the best solvent for the extraction with an extract yield of 57.3%, a total anthocyanin content of 5.1 mg of cyanidin 3-glucoside equivalent/g of dry weight of extract, and a total phenolic content of 59.4 mg of gallic acid equivalent/g of dry weight of extract. Similar results were obtained for the best water extract at 50 °C for 1 h. A larger number of phytochemicals were identified using LC-MS analysis in the solvent extract (28 compounds). Kaempferol hexosyl-rhamnosyl-rhamnoside was the major compound detected in both the solvent and the water extract. Both extracts showed equally potent antioxidant activity in chemical [IC 50 = 1.24 ± 0.05 (solvent extract) and 1.18 ± 0.07 (water extract) mg/mL in the 2,2-diphenyl-1-picryl-hydrazyl-hydrate assay] and cellular (75−80% inhibition) antioxidant assays.

Background: Andrographolide has been reported with anticancer and anti-inflammatory properties through the inhibition of the activity of signaling molecules such as v-Src, nuclear factor-κB (NF-κB), STAT3, and PI3K. NF-κB has been proven to promote cancer cell survival, and targeting this pathway will halt the growth of cancer cells. Efforts have been made to produce semisynthetic derivatives of andrographolide with improved anticancer potency and selectivity. Subsequently, the effect of a selected derivative, 3,14,19-tripropionylandrographolide (SRS06), was tested for its action against NF-κB. Methods: Screening against 60 US National Cancer Institute (NCI) human cancer cell lines representing leukemia and non-small cell lung (NSCL), colon, CNS, melanoma, ovarian, renal, prostate, and breast cancers was performed to determine the tumor type selectivity and potency of SRS06. Microculture tetrazolium, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide and sulforhodamine B assays were used to determine the in vitro anticancer activity, while Western blot studies were performed to ascertain the inhibitory effect of SRS06 on the NF-κB signaling cascade. The TransAM™ p65 assay kit was used to determine NF-κB p65 DNA binding activity in the NSCL cancer cell line A549. Results: From the NCI screening, SRS06 was found to exhibit potent growth-inhibitory effects on multiple cancer cell lines with 10-fold lower 50% growth inhibition (GI₅₀) compared with andrographolide. It was also discerned that the compound preferentially targeted melanoma, CNS, renal, colon, ovarian, prostate, and NSCL cancer cell lines. The DNA fragmentation assay indicated that the main mode of cell death of SRS06-treated A549 cells was via apoptosis. At 5 µmol/l the compound decreased NF-κB protein expression and caused a significant reduction in the nuclear p65 DNA binding activity. Conclusion: SRS06 displayed improved anticancer selectivity and potency when compared with andrographolide. We alluded its anticancer activity to its effect of inhibiting NF-κB nuclear binding.

In Asia, Clitoria ternatea flowers are commonly used as a traditional medicinal herb and as a food colourant. Their bioactive compounds have anti-inflammatory, anti-microbial and anti-biofilm activities. Pseudomonas aeruginosa is one of the major pathogens that cause biofilm-associated infections resulting in an increase in antimicrobial resistance. Hence, the aim of this study was to investigate if the anti-biofilm properties of the anthocyanin-rich fraction of C. ternatea flowers were effective against P. aeruginosa . The effect of the anthocyanin-rich fraction of C. ternatea flowers on P. aeruginosa biofilms formed on a polystyrene surface was determined using the crystal violet assay and scanning electron microscopy (SEM). The anthocyanin-rich fraction reduced biofilm formation by four P. aeruginosa strains with a minimum biofilm inhibitory concentration value ranging between 0.625 and 5.0 mg ml−1. We further show that the biofilm-inhibiting activity of C. ternatea flowers is not due to the flavonols but is instead attributed to the anthocyanins, which had significant biofilm inhibitory activity (64.0±1.1 %) at 24 h in a time–response study. The anthocyanin-rich fraction also significantly reduced bacterial attachment on the polystyrene by 1.1 log c.f.u. cm−2 surface based on SEM analysis. Hence, anthocyanins from C. ternatea flowers have potential as an agent to decrease the risk of biofilm-associated infections.

Clitoria ternatea flowers are rich in anthocyanins and possess various biological activities. Specifically, the antibacterial mechanism of action of C. ternatea anthocyanins remains unknown and was investigated in Escherichia coli. Time-kill assay was used to assess the antibacterial activity and the metabolic perturbations in E. coli were investigated utilising LC-MS-based metabolomics. Pathway analyses were carried out for metabolites showing ≥ 2-fold changes. The anthocyanin fraction remarkably reduced the growth of E. coli at 4 h by 95.8 and 99.9% at minimum inhibitory concentration (MIC) and 2 × MIC, respectively. The antibacterial activity of the anthocyanin fraction (MIC) was shown to have perturbed glycerophospholipids (1-Acyl-sn-glycero-3-phosphoethanolamine, phosphatidylglycerol, diacylglycerol and cardiolipin), amino acids (valine, tyrosine and isoleucine) and energy (ubiquinone and NAD) metabolites at 1 and 4 h. This study demonstrated significant metabolic perturbations of the glycerophospholipid, amino acid and energy metabolism being the key pathways involved in the antibacterial activity of anthocyanins from C. ternatea and has promising potential as antibacterial agents for E. coli-related infections.

Clitoria ternatea flowers are rich in anthocyanins and possess various biological activities. Specifically, the antibacterial mechanism of action of C. ternatea anthocyanins remains unknown and was investigated in Escherichia coli. Time-kill assay was used to assess the antibacterial activity and the metabolic perturbations in E. coli were investigated utilising LC-MS-based metabolomics. Pathway analyses were carried out for metabolites showing ≥ 2-fold changes. The anthocyanin fraction remarkably reduced the growth of E. coli at 4 h by 95.8 and 99.9% at minimum inhibitory concentration (MIC) and 2 × MIC, respectively. The antibacterial activity of the anthocyanin fraction (MIC) was shown to have perturbed glycerophospholipids (1-Acyl-sn-glycero-3-phosphoethanolamine, phosphatidylglycerol, diacylglycerol and cardiolipin), amino acids (valine, tyrosine and isoleucine) and energy (ubiquinone and NAD) metabolites at 1 and 4 h. This study demonstrated significant metabolic perturbations of the glycerophospholipid, amino acid and energy metabolism being the key pathways involved in the antibacterial activity of anthocyanins from C. ternatea and has promising potential as antibacterial agents for E. coli-related infections.