Pressurised water extraction of polyphenols from pomegranate peels

Pomegranate peels are one of the most valuable by-products of the food industry in terms of polyphenols which are conventionally extracted from plant materials by organic solvents, especially with methanol. Pressurised water extraction was investigated for the extraction of polyphenols from pomegranate peels. The most important factors affecting the extraction results were found to be particle size, temperature, and static time. The results indicated that pressurised water extraction was as effective as conventional methanol extraction for the recovery of polyphenols from pomegranate peels. Total phenolic contents of pomegranate peels obtained by pressurised water extraction at optimised conditions and conventional solid–liquid methanol extraction were determined as 264.3 and 258.2 mg/g tannic acid equivalents, respectively. Hydrolyzable tannins were the predominant polyphenols of pomegranate peels corresponding to 262.7 mg/g tannic acid equivalents. Punicalagin content of pomegranate peels by pressurised water extraction was found to be 116.6 mg/g on dry matter basis.     

Antibacterial,anti-inflammatory and anti-allergic activities of standardised pomegranate rind extract

Antibacterial, anti-inflammatory and anti-allergic activities of standardised pomegranate rind extract (SPRE) containing 13% w/w ellagic acid were studied in vitro. The antibacterial activity of SPRE was determined using the disc diffusion and broth microdilution methods. SPRE exhibited a potent bacteriostatic effect against Propionibacterium acnes, a Gram-positive anaerobe, with a MIC of 15.6 μg/ml, and Gram-positive facultative anaerobic bacteria, Staphylococcus aureus and Staphylococcus epidermidis, with MICs of 7.8–15.6 μg/ml. Anti-inflammatory activity of SPRE was evaluated by measuring the inhibition of nitric oxide (NO) production by murine macrophage-like RAW264.7 cells. SPRE exhibited a potent NO inhibitory effect, with an IC₅₀ of 10.7 μg/ml. Evaluation of the anti-allergic activity showed that SPRE inhibited the release of β-hexosaminidase from antigen-stimulated rat basophilic leukemia (RBL-2H3) cells with an IC₅₀ of 20.9 μg/ml. In addition, SPRE exhibited only moderate cytotoxicity on human keratinocyte cells, with CC₅₀ of 33.6 μg/ml. These findings support the potential use of SPRE as a nutraceutical for antibacterial, anti-inflammatory and anti-allergic proposes.     

Effects of ellagitannin‐rich berries on blood lipids,gut microbiota,and urolithin production in human subjects with symptoms of metabolic syndrome

Ellagitannins are polyphenols abundant in strawberries, raspberries, and cloudberries. The effects of a mixture of these berries were studied in a randomized controlled trial with subjects having symptoms of metabolic syndrome. The study focused on serum lipid profiles, gut microbiota, and ellagitannin metabolites. The results indicate that bioavailability of ellagitannins appears to be dependent on the composition of gut microbiota.     

The p53-, Bax- and p21-dependent inhibition of colon cancer cell growth by 5-hydroxy polymethoxyflavones

Scope: Previously, we reported that 5‐hydroxy polymethoxyflavones (5OH‐PMFs) isolated from orange, namely 5‐hydroxy‐6,7,8,3′,4′‐pentamethoxyflavone, 5‐hydroxy‐3,6,7,8,3′,4′‐hexamethoxyflavone (5HHMF) and 5‐hydroxy‐6,7,8,4′‐tetramethoxyflavone (5HTMF), potently induced apoptosis and cell‐cycle arrest in multiple human colon cancer cells. Herein, using isogenic variants of HCT116 human colon cancer cells, we investigated the effects of p53, Bax and p21 on the apoptosis and cell‐cycle arrest induced by different 5OH‐PMFs. Methods and results: Annexin V/PI co‐staining assay demonstrated that 5HHMF and 5HTMF significantly induced apoptosis in HCT116 (p53^+/+) cells but not in HCT116 (p53^?/?) cells. Furthermore, 5HHMF and 5HTMF significantly induced apoptosis in HCT116 (Bax^+/?) cells, whereas their pro‐apoptotic effects on HCT116 (Bax^?/?) cells were marginal. All three 5OH‐PMFs increased G0/G1 cell population of HCT116 (p53^+/+) cells, and these effects were abolished in HCT116 (p53^?/?) and HCT116 (p21^?/?) cells. Immunoblotting analysis showed that 5HHMF and 5HTMF increased the levels of cleaved caspase‐3, cleaved PARP in both HCT116 (p53^+/+) and HCT116 (Bax^+/?) cells and these effects were much weaker in HCT116 (p53^?/?) and HCT116 (Bax^?/?) cells. Conclusion: Our results demonstrated that 5OH‐PMFs, especially 5HHMF and 5HTMF, induce apoptosis and cell‐cycle arrest by p53‐, Bax‐ and p21‐dependent mechanism.     

Combination of green tea, phytic acid, and inositol reduced the incidence of azoxymethane-induced colon tumors in Fisher 344 male rats

Epidemiological studies have shown an inverse relationship between consumption of nutritive/non-nutritive foods of plant origin and colon cancer incidence. This study was conducted to determine the effect of green tea, phytic acid, and inositol at 2 g/100 ml levels singly and in combination on azoxymethane (AOM) induced colon tumors in Fisher 344 male rats. After an acclimatization period of one week, 8 groups of rats (15 rats each) were initially assigned to consume AIN 93G and later AIN 93M diet. All treatments were given in drinking water. All the rats received 16 mg/kg body mass AOM, two s/c injections at seven and eight week of age. Rats were killed at 46 week of age by CO₂ euthanasia. Tumor incidence (percent) and tumors per tumor-bearing rat (TBR) in the control were significantly higher (P<0.05) than all treatment groups. Glutathione-S-transferase (GST) activity was significantly higher in treatment groups compared to control. These findings suggest that the additive effect of green tea, phytic acid and inositol may reduce the incidence of colon tumors, and can also be used as an adjuvant to chemomodulation.     

Distribution of chloramphenicol to tissues,plasma and urine in pigs after oral intake of low doses

Toxic effects of chloramphenicol in humans caused the ban for its use in food-producing animals in the EU. A minimum required performance level (MRPL) was specified for chloramphenicol at 0.3 μg kg^–1 for various matrices, including urine. In 2012, residues of chloramphenicol were found in pig urine and muscle without signs of illegal use. Regarding its natural occurrence in straw, it was hypothesised that this might be the source, straw being compulsory for use as bedding material for pigs in Sweden. Therefore, we investigated if low daily doses of chloramphenicol (4, 40 and 400 μg/pig) given orally during 14 days could result in residues in pig tissues and urine. A dose-related increase of residues was found in muscle, plasma, kidney and urine (showing the highest levels), but no chloramphenicol was found in the liver. At the lowest dose, residues were below the MRPL in all tissues except in the urine. However, in the middle dose, residues were above the MRPL in all tissues except muscle, and at the highest dose in all matrices. This study proves that exposure of pigs to chloramphenicol in doses occurring naturally in straw could result in residues above the MRPL in plasma, kidney and especially urine.     

SB365, Pulsatilla saponin D suppresses the proliferation of human colon cancer cells and induces apoptosis by modulating the AKT/mTOR signalling pathway

Pulsatilla koreana has been used as a traditional medicine for the treatment of several diseases. The purpose of this study was to determine if SB365, Pulsatilla saponin D isolated from the root of P. koreana inhibits the progression of colon cancer. We found that SB365 strongly suppressed the growth and proliferation of colon cancer cells and induced their apoptosis. Also, SB365 showed anti-angiogenic activity by decreasing the expression of HIF-1α and VEGF. These results were confirmed by an in vivo study showing that SB365 significantly inhibited tumor growth by the induction of apoptosis and inhibition of angiogenesis with stronger anticancer activity than 5-FU. When further examined for its anticancer mechanism, SB365 effectively suppressed the AKT/mTOR pathway both in vitro and in vivo. Taken together, our study demonstrated that SB365 inhibits the AKT/mTOR pathway, leading to the suppression of tumor growth and angiogenesis together with induction of apoptosis. Therefore, SB365 is a good candidate as a natural product for use in the treatment of colon cancer.     

Lunasin induces apoptosis and modifies the expression of genes associated with extracellular matrix and cell adhesion in human metastatic colon cancer cells

Scope: Lunasin is an arginine‐glycine‐aspartic acid (RGD) cancer preventive peptide. The objective was to evaluate the potential of lunasin to induce apoptosis in human colon cancer cells and their oxaliplatin‐resistant (OxR) variants, and its effect on the expression of human extracellular matrix and adhesion genes. Methods and results: Various human colon cancer cell lines which underwent metastasis were evaluated in vitro using cell flow cytometry and fluorescence microscopy. Lunasin cytotoxicity to different colon cancer cells correlated with the expression of α₅b₁ integrin, being most potent to KM12L4 cells (IC₅₀ = 13 μM). Lunasin arrested cell cycle at G2/M phase with concomitant increase in the expression of cyclin‐dependent kinase inhibitors p21 and p27. Lunasin (5–25 μM) activated the apoptotic mitochondrial pathway as evidenced by changes in the expressions of Bcl‐2, Bax, nuclear clusterin, cytochrome c and caspase‐3 in KM12L4 and KM12L4‐OxR. Lunasin increased the activity of initiator caspase‐9 leading to the activation of caspase‐3 and also modified the expression of human extracellular matrix and adhesion genes, downregulating integrin α₅, SELE, MMP10, integrin β₂ and COL6A1 by 5.01‐, 6.53‐, 7.71‐, 8.19‐ and 10.10‐fold, respectively, while upregulating COL12A1 by 11.61‐fold. Conclusion: Lunasin can be used in cases where resistance to chemotherapy developed.     

Xanthohumol induces apoptosis in cultured 40-16 human colon cancer cells by activation of the death receptor- and mitochondrial pathway

Xanthohumol (XN) is one of the major prenylflavonoids found in hop cones (Humulus lupulus L.). In this study, we investigated the cell growth inhibitory potential of XN on cultured human colon cancer cells. Cell proliferation was measured by sulforhodamine B staining. Poly(ADP-ribose)polymerase (PARP) cleavage, activation of caspases-3, -7, -8, and -9, and Bcl-2 family protein expression were detected by Western blot analyses. XN significantly reduced proliferation of the HCT 116-derived colon cancer cell line 40--16. Half-maximal inhibitory concentrations decreased from 4.1 microM after 24 h treatment to 3.6 and 2.6 microM after 48 and 72 h incubation, respectively. Treatment with 15 microM XN for 48 h and with 5 microM for 72 h led to the detection of the cleaved 89 kDa fragment of 116 kDa PARP as an indication of apoptosis induction. Concomitantly, we observed activation and cleavage of the effector caspases-3 and -7, induced by activation of the initiator caspases -8 and -9. Expression of anti-apoptotic Bcl-2 was down regulated when the cells were treated with XN for 48--72 h. We conclude that induction of apoptosis by downregulation of Bcl-2 and activation of the caspase cascade may contribute to the chemopreventive or therapeutic potential of XN.