The preventive effects of the American cranberry (Vaccinium macrocarpon) against urinary tract infections are supported by extensive studies which have primarily focused on its phenolic constituents. Herein, a phenolic-free carbohydrate fraction (designated cranf1b-F2) was purified from cranberry fruit using ion exchange and size exclusion chromatography. MALDI-TOF-MS analysis revealed that the cranf1b-F2 constituents are predominantly oligosaccharides possessing various degrees of polymerisation and further structural analysis (by GC-MS and NMR) revealed mainly xyloglucan and arabinan residues. In antimicrobial assays, cranf1b-F2 (at 1.25 mg/mL concentration) reduced biofilm production by the uropathogenic Escherichia coli CFT073 strain by over 50% but did not inhibit bacterial growth. Cranf1b-F2 (ranging from 0.625 to 10 mg/mL) also inhibited biofilm formation of the non-pathogenic E. coli MG1655 strain up to 60% in a concentration-dependent manner. These results suggest that cranberry oligosaccharides, in addition to its phenolic constituents, may play a role in its preventive effects against urinary tract infections.

A novel methodology was developed to elucidate proanthocyanidins (PAC) interaction with extra-intestinal pathogenic Escherichia coli (ExPEC). PAC inhibit ExPEC invasion of epithelial cells and, therefore, may prevent transient gut colonization, conferring protection against subsequent extra-intestinal infections, such as urinary tract infections. Until now PAC have not been chemically labeled with fluorophores. In this work, cranberry PAC were labeled with 5-([4,6-dichlorotriazin-2-yl]amino) fluorescein (DTAF), detected by high-performance liquid chromatography with diode-array detection and characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). We report single and double fluorescent-labeled PAC with one or two chlorine atoms displaced from DTAF in alkaline pH via nucleophilic substitution. Fluorescent labeling was confirmed by fragmentation experiments using MALDI-TOF/TOF MS. Fluorescent labeled PAC were able to promote ExPEC agglutination when observed with fluorescence microscopy. DTAF tagged PAC may be used to trace the fate of PAC after they agglutinate ExPEC and follow PAC-ExPEC complexes in cell culture assays.

Respiratory viruses are a major public health problem because of their prevalence and high morbidity rate leading to considerable social and economic implications. Cranberry has therapeutic potential attributed to a comprehensive list of phytochemicals including anthocyanins, flavonols, and unique A-type proanthocyanidins. Soy flavonoids, including isoflavones, have demonstrated anti-viral effects in vitro and in vivo. Recently, it was demonstrated that edible proteins can efficiently sorb and concentrate cranberry polyphenols, including anthocyanins and proanthocyanins, providing greatly stabilized matrices suitable for food products. The combination of cranberry and soy phytoactives may be an effective dietary anti-viral resource. Anti-viral properties of both cranberry juice-enriched and cranberry pomace polyphenol-enriched soy protein isolate (CB-SPI and CBP-SPI) were tested against influenza viruses (H7N1, H5N3, H3N2), Newcastle disease virus and Sendai virus in vitro and in ovo. In our experiments, preincubation with CB-SPI or CBP-SPI resulted in inhibition of virus adsorption to chicken red blood cells and reduction in virus nucleic acid content up to 16-fold, however, CB-SPI and CBP-SPI did not affect hemagglutination. Additionally, CB-SPI and CBP-SPI inhibited viral replication and infectivity more effectively than the commercially available anti-viral drug Amizon. Results suggest CB-SPI and CBP-SPI may have preventative and therapeutic potential against viral infections that cause diseases of the respiratory and gastro-intestinal tract.

In this study, we have assessed the phenolic metabolism of a cranberry extract by microbiota obtained from the ascending colon and descending colon compartments of a dynamic gastrointestinal simulator (SHIME). For comparison, parallel fermentations with a grape seed extract were carried out. Extracts were used directly without previous intestinal digestion. Among the 60 phenolic compounds targeted, our results confirmed the formation of phenylacetic, phenylpropionic and benzoic acids as well as phenols such as catechol and its derivatives from the action of colonic microbiota on cranberry polyphenols. Benzoic acid (38.4mug/ml), 4-hydroxy-5-(3'-hydroxyphenyl)-valeric acid (26.2mug/ml) and phenylacetic acid (19.5mug/ml) reached the highest concentrations. Under the same conditions, microbial degradation of grape seed polyphenols took place to a lesser extent compared to cranberry polyphenols, which was consistent with the more pronounced antimicrobial effect observed for the grape seed polyphenols, particularly against Bacteroides, Prevotella and Blautia coccoides-Eubacterium rectale.

OBJECTIVE: In the context of increasing microbial resistance and limited new antimicrobials, we aimed to study the antimicrobial effects of cranberry proanthocyanidin extracts on Escherichia coli growth, adhesion to epithelial cells, and lung infection.
DESIGN: Experimental in vitro and in vivo investigation.
SETTING: University research laboratory.
SUBJECTS: Seventy-eight 6- to 8-week-old male Balb/C mice.
INTERVENTIONS: In vitro, the effect of increasing concentrations of cranberry proanthocyanidin on bacterial growth of different clinical E. coli isolates was evaluated. Ex vivo, adhesion of E. coli to fresh human buccal epithelial cells was measured in the presence or absence of cranberry proanthocyanidin using microscopy. In vivo, lung bacterial count, pulmonary immune response (neutrophil murine chemokine keratinocyte-derived cytokine measurement and polymorphonuclear recruitment in bronchoalveolar lavage fluid), and lethality were evaluated in a pneumonia mouse model with E. coli precultured with or without cranberry proanthocyanidin. E. coli isolates originated from ventilated ICU patients with respiratory tract colonization or ventilator- associated pneumonia. They differed in number of virulence genes.
MEASUREMENTS AND MAIN RESULTS: A significant inhibition of bacterial growth was observed with increasing concentration of cranberry proanthocyanidin, affecting both time to maximal growth and maximal growth rate (p0.0001 for both). The minimal concentration at which this effect occurred was 250 mug/mL. Cranberry proanthocyanidin significantly reduced E. coli adhesion to fresh buccal epithelial cells by up to 80% (p0.001). Bacterial counts in homogenized lungs and bronchoalveolar lavage fluid were decreased after cranberry proanthocyanidin exposition (p0.05 and p0.01, respectively). Cranberry proanthocyanidin also decreased KC concentrations and polymorphonuclear cell recruitment in bronchoalveolar lavage fluid (p0.05 for both). At identical inoculum, mortality was reduced by more than half in mice inoculated with E. coli exposed to cranberry proanthocyanidin (p0.01).
CONCLUSION: Cranberry proanthocyanidins exhibit potent effects on growth, adhesion, and virulence of oropharyngeal and lung isolates of E. coli, suggesting that cranberry proanthocyanidin could be of clinical interest to reduce oropharyngeal colonization and prevent lung infection.

Cranberry flavonoids (flavonols and flavan-3-ols), in addition to their antioxidant properties, have been shown to possess potential in vitro activity against several cancers. However, the difficulty of isolating cranberry compounds has largely limited anticancer research to crude fractions without well-defined compound composition. In this study, individual cranberry flavonoids were isolated to the highest purity achieved so far using gravity and high performance column chromatography and LC-MS characterization. MTS assay indicated differential cell viability reduction of SKOV-3 and OVCAR-8 ovarian cancer cells treated with individual cranberry flavonoids. Treatment with quercetin aglycone and PAC DP-9, which exhibited the strongest activity, induced apoptosis, led to caspase-3 activation and PARP deactivation, and increased sensitivity to cisplatin. Furthermore, immunofluorescence microscopy and western blot study revealed reduced expression and activation of epidermal growth factor receptor (EGFR) in PAC DP-9 treated SKOV-3 cells. In addition, quercetin aglycone and PAC DP-9 deactivated MAPK-ERK pathway, induced downregulation of cyclin D1, DNA-PK, phospho-histone H3 and upregulation of p21, and arrested cell cycle progression. Overall, this study demonstrates promising in vitro cytotoxic and anti-proliferative properties of two newly characterized cranberry flavonoids, quercetin aglycone and PAC DP-9, against ovarian cancer cells.

BACKGROUND AND OBJECTIVE: In periodontitis, gingival epithelial cells can produce interleukin (IL)-6, a regulator of osteoclastic bone resorption, in response to IL-1beta. IL-1beta regulates cytokine expression via signaling pathways, including nuclear factor (NF)-B and mitogen activated protein kinase (MAPK)/activator protein (AP)-1. Cranberry proanthocyanidins (PACs) inhibit IL-1beta-stimulated IL-6 production, but specific mechanisms are unclear. The objectives of this study were to determine effects of cranberry PACs on NF-B and MAPK/AP-1 activation of IL-1beta-stimulated IL-6 production in gingival epithelial cells.

MATERIAL AND METHODS: Cranberry high molecular weight non-dialyzable material (NDM), rich in PACs, was derived from cranberry juice. Human gingival epithelial cells [Smulow-Glickman (S-G)] were incubated with IL-1beta in the presence or absence of NDM or inhibitors of NF-B, [nemo-binding domain (NBD) peptide] or AP-1 (SP600125), and IL-6 levels were measured by ELISA. Effects of NDM on IL-1beta-activated NF-B and AP-1 and phosphorylated intermediates in both pathways were measured in cell extracts via binding to specific oligonucleotides and specific sandwich ELISAs, respectively. Data were analyzed using ANOVA and Scheffe's F procedure for post hoc comparisons.

RESULTS: IL-1beta (> 0.1 nm) caused a time- and dose-dependent stimulation of S-G epithelial cell IL-6 production (p 0.005). This was significantly decreased in a dose-dependent manner by NBD peptide or SP600125 [maximum inhibition ~30-40% (p 0.02)], and together, the two inhibitors decreased IL-6 by ~80%, similar to the inhibition caused by NDM (p 0.001). IL-1beta stimulated NF-B and AP-1 activation (p 0.003), which was inhibited by NDM (p 0.0001). NDM did not significantly affect IL-1beta-stimulated levels of phosphorylated intermediates in the NF-B pathway (IBalpha) or the AP-1 pathway (c-Jun, ERK1/2).

CONCLUSION: In S-G epithelial cells, IL-1beta appeared to upregulate IL-6 production via activation of both NF-B and MAPK/AP-1 signaling pathways because cranberry NDM decreased nuclear levels of IL-1beta-activated NF-B (p65) and AP-1 (phospho-c-Jun) and strongly inhibited IL-6 production. Lack of inhibition of phosphorylation of IBalpha, c-Jun or ERK1/2 suggested that NDM might affect both pathways downstream from those points in S-G cells, such as ubiquitination and proteosomal degradation of IBalpha, or inhibition of nuclear activity of c-Jun and/or ERK1/2. Defining these points of inhibition precisely may help identify molecular targets of cranberry polyphenols. 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

In the context of dental caries prevention by natural foodstuff sources, antifungal and antibiofilm activities of dry commercial extracts of cranberry fruit (Vaccinium macrocarpon Aiton) and two other red fruits (Vaccinium myrtillus L. and Malpighia punicifolia L.) were assessed on Candida albicans and Candida glabrata yeasts. When added to the culture medium, the cranberry extract displayed a significant anti-adhesion activity against Candida spp. when used at low concentrations. In addition, the pretreatment of surfaces with this extract induced an anti-adhesion activity mainly against C. glabrata yeasts and an antibiofilm activity against C. albicans. This activity was dependent on concentration, species, and strain. A phytochemical investigation bioguided by anti-adhesion tests against the two Candida species was carried out on crude cranberry juice to determine the active fractions. Three subfractions enriched in proanthocyanidins showed an anti-adhesion activity at low concentrations. This study investigated for the first time the interest of crude extracts of cranberry and cranberry juice fractions to prevent biofilms of C. glabrata. It highlighted the potency of consuming this fruit and using it as a source of anti-adhesion agents. 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

BACKGROUND AND OBJECTIVE: Gingival epithelial cells and fibroblasts participate in periodontal inflammation and destruction, producing interleukin (IL)-6, a regulator of osteoclastic bone resorption, and the neutrophil chemoattractant IL-8. IL-17, a product of T-helper 17 cells, may play a role in periodontitis by stimulating cytokine production by gingival cells. The cranberry (Vaccinium macrocarpon) is rich in polyphenols, particularly proanthocyanidins, which have antioxidant and other beneficial properties. Cranberry components inhibit pro-inflammatory activities of lipopolysaccharide-stimulated human macrophages, gingival fibroblasts, and epithelial cells, but little is known of its effects on IL-17-stimulated cytokine production. The objectives were to determine the effects of IL-17 + cranberry components on IL-6 and IL-8 production by human gingival epithelial cells and fibroblasts.

MATERIAL AND METHODS: Cranberry high molecular weight non-dialyzable material (NDM), which is rich in proanthocyanidins, was derived from cranberry juice. Human gingival epithelial cells and normal human gingival fibroblasts were incubated with NDM (5-50 mug/mL), IL-17 (0.5-100 ng/mL), or NDM + IL-17 in serum-free medium for 6 d. IL-6 and IL-8 in culture supernatants were measured by ELISA. Membrane damage and viability were assessed by lactate dehydrogenase activity released into cell supernatants and activity of a mitochondrial enzyme, respectively. Data were analyzed using ANOVA and Scheffe's F procedure for post hoc comparisons.

RESULTS: In both cell lines, IL-17 (> ~5-10 ng/mL) significantly stimulated production of IL-6 (p 0.005) and IL-8 (p 0.03). Non-toxic levels of NDM inhibited constitutive IL-6 and IL-8 production by epithelial cells (p 0.01) and fibroblasts (p 0.03) as well as IL-17-stimulated cytokine production by epithelial cells [IL-6 (maximum ~80% inhibition; p 0.0001); IL-8 (maximum ~70% inhibition; p 0.03)] and fibroblasts [IL-6 (maximum ~90% inhibition; p 0.0001); IL-8 (maximum ~80% inhibition; p 0.008)].

CONCLUSION: Cranberry NDM inhibition of constitutive and IL-17-stimulated IL-6 and IL-8 production by gingival fibroblasts and epithelial cells suggests that cranberry components could be useful as a host modulatory therapeutic agent to prevent or treat periodontitis. 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

BACKGROUND: Berries are known to have many kinds of biological activities. We focused on their antiviral effect, which has not yet been well evaluated.

RESULTS: We compared the anti-influenza viral effects of berries belonging to the genus Vaccinium - 35 species of blueberry (Vaccinium cyanococcus), the Natsuhaze (Vaccinium oldhamii), bilberry (Vaccinium myrtillus) and cranberry (Vaccinium oxycoccos)- with those belonging to the genus Ribes, i.e. blackcurrant (Ribes nigrum). Only Elliott and Legacy among Northern Highbush varieties but many Rabbiteye varieties such as Austin, Baldwin, Brightblue, Festival, T-100 and Tifblue showed anti-influenza viral activity. Natsuhaze, bilberry, cranberry and blackcurrant had high antiviral effects. A relationship was observed between the antiviral effect and total polyphenol content.

CONCLUSIONS: Antiviral effects were found to differ markedly between berry species. Rabbiteye varieties tended to have higher antiviral effects than Northern, Southern and Half Highbush blueberry varieties. We also found that Natsuhaze, which has recently been harvested in Japan as a potential functional food, had an antiviral effect comparable to that of bilberry, cranberry and blackcurrant. There was a positive relationship between antiviral activity and polyphenol content, indicating the possibility that polyphenol is one of the key factors in the antiviral effects of berries. 2012 Society of Chemical Industry.