Fig 1.
Chemical structures of major EVOO phenolics.
S-Oleocanthal (OC), S-Hydroxyoleocanthal (HOC), S-Ligstroside aglycone (LA), S-Oleuropein aglycone (OA), Tyrosol (TY), Hydroxytyrosol (HT), (+)-Pinoresinol (PR), (+)-1-Acetoxypinoresinol (APR).
Fig 2.
Novel ultra-freezing-based method for EVOO phenolics purification.
The sequential steps of mixing, ultra-freezing, and filtration procedures.
Fig 3.
Simple water-based liquid-liquid extraction method for EVOO phenolics isolation.
The sequential steps of mixing and filtration procedures.
Fig 4.
Application of OC-water isolation method on the Governor EVOO batch number: 5–214000242017.
A. Oleocanthal standard calibration curve using quantitative HPLC data. B. HPLC Monitoring of water extraction process; i) HPLC chromatogram for crude EVOO and EVOO after water extraction. ii) HPLC chromatogram of the 1st -3rd water extraction. C. OC standard calibration curve using q1H NMR data. D. 1H NMR-based monitoring of the extraction process. i) 1H NMR spectrum of crude water extract showing OC as a major EVOO phenolic ingredient. ii) Pure OC 1H NMR spectrum after Sephadex LH20 purification.
Table 1.
Efficiency of water extraction method in diverse EVOO sources a.
Table 2.
Analysis of different phenolic ingredients in representative commercial EVOO sources a.
Table 3.
Stability of OC in water extracts after one-month storage at room temperature and 4 0Ca.
Fig 5.
Physical characteristics of OC nano-emulsion using.
A. Particle size measurement, B. Polydispersity index, and C. Zeta potential measurement. One-month stability and physical characteristics of water extraction nano-emulsion at RT and 4 ºC using: D. Particle size measurement, E. Polydispersity index, and F. Zeta potential measurement.
Fig 6.
Effect OC nano-emulsion on BC and the non-tumorigenic mammary epithelial cells.
BC cell lines MDA-MB-231, MDA-MB-468, BT-474, and MCF-7, in addition to the non-tumorigenic mammary epithelial MCF10A cells were treated by OC nano-emulsion at 72 h treatment period to see the effects of treatment on cells’ growth compared to vehicle control. The non-malignant cells MCF10A was also used to assess treatment selectivity. Viable cell count was determined using MTT assay. Vertical bars indicate the mean cell count ± SD in each treatment group, n = 3/treatment dose. *p <0.05 as compared to vehicle-treated control group.
Fig 7.
In vivo oral activity of pure OC extracted by water extraction method and using water as exclusive vehicle against the human TNBC MDA-MB-231 cells in orthotopic nude mouse xenograft model.
A. Design layout of the experiment. B. Vertical bars comparing the mean tumor weight of treated versus vehicle control at the end of the experiment. C. Vertical bars comparing the last day mean tumor volume of treated versus vehicle control at the end of the experiment in each group (n = 5). D. Representative experiment mice and collected tumors at the end of experiment in each group (n = 5). Right mouse: OC-treated orally at 10 mg kg/day, 5X/week. Left mouse: vehicle-treated control. E. Onset of tumor growth. F. Mean tumor volume, n = 5, over the experiment period. Tumor volume (V) calculated using the formula: V = L/2 x W2, where L is the length and W is the width of tumors. Collected points represent the mean of tumor volume in mm3 in each group (n = 5). G. Monitored mice body weight over the experiment duration. Points represent the mean mice body weight in each group (n = 5) over the experiment duration. Error bars indicate SD for n = 5. **p <0.005 as compared to vehicle-treated control group.