Progress 09/01/02 to 08/31/05
Outputs
Cranberry PA was applied to a semi-preparative LH-20 column equilibrated in water. Sequential solvent elution from the column produced 3 PA fractions: LH1; ethanol:water (1:1), LH2; ethanol, and an aqueous acetone (4:1, acetone: H2O, v:v) fraction. Fractions LH3, LH4 and LH5 had the greatest anti-adherence activity and were subjected to reverse phase (RP) C18 subfractionation. Elution from the RPC18 column produced 6 PA fractions: LH3RP1, LH3RP2, LH4RP1, LH4RP2, LH5RP1 and LH5RP2. Fractions LH3RP2, LH4RP1 and LH4RP2 had the greatest anti-adherence activity. We are in the process of developing a normal phase (LC) separation scheme for these PA fractions and anticipate this step will be accomplished on schedule (5/31/04) as outlined in our previous proposal. We developed a method of estimating relative ratios of A-type:B-type interflavan bonds by deconvolution of overlapping isotope patterns. We developed a method to eliminate the problem of cationization with Na+ and
K+.
Impacts
Urinary tract infections account for over 11 million doctor visits annually in the U.S. alone. Proanthocyanidins isolated from cranberry fruit exhibit potent bacterial anti-adherence activity in vitro and in vivo. Thus, it is important to determine the degree of structural heterogeneity of cranberry proanthocyanidins to understand how molecular weight and linkage type influence bacterial anti-adherence activity.
Publications
- Krueger, C.G.; M.M. Vestling and J.D. Reed. 2003. Matrix-assisted laser desorption/Ionization time-of-flight mass spectrometry of anthocyanin-polyflavan-3-ol polymers in cranberry fruit [Vaccinium macrocarpon, Ait.] and spray dried cranberry juice. ACS Symposium, Uncovering the Mysteries of Red Wine Pigments (in press).
- Krueger, C.G.; M.M. Vestling and J.D. Reed. MALDI-TOF MS demonstrates that the structural diversity of polyflavans in foods and beverages is much greater than previously Appreciated. Abstract and Poster presentation at The 51st Annual Conference on Mass Spectrometry and Allied Topics, June 2-12, 2003 in Montreal, Quebec, Canada. Reed, J.D.; Howell, A.; Cunningham, D.; Krueger C.G. Differences in Structure and Bacterial Anti-adhesion Activity of Cranberry Proanthocyanidins Compared to Proanthocyanidins from other Foods. Abstract and Poster presentation at the 1st International Conference on Polyphenols and Health, Nov 12-21, 2003 in Vichy, France.
- Reed, J. D., C. g. Krueger and M. M. Vestling. 2005. MALDI-TOF mass spectrometry of oligomeric food polyphenols. Phytochemistry 66(18):2247-2262.
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Progress 01/01/04 to 12/31/04
Outputs
Clinical, epidemiological and mechanistic studies support the role of cranberry (Vaccinium macrocarpon Ait.) in maintaining urinary tract health. Cranberry proanthocyanidins contain A-type linkages and have been associated with preventing adhesion of P-fimbriated uropathogenic E. coli to uroepithelial cells. It is not known if the presence of the A-type linkage is a prerequisite for anti-adhesion activity. Other commercial sources of proanthocyanidins with all B-type linkages have not previously been screened for this activity. The goals of this study were to compare the in vitro anti-adhesion activity of A-linked proanthocyanidins from cranberry juice cocktail with the anti-adhesion activities of B-linked proanthocyanidins from commercial grape and apple juices, green tea and dark chocolate, and compare the in vivo anti-adhesion activity of human urine following consumption of single servings of each commercial food product. Structural heterogeneity and presence of
the A-type linkage in cranberry proanthocyanidins was confirmed utilizing MALDI-TOF/MS and DI/ESI MS, as was the presence of all B-type linkages in the proanthocyanidins from the other commercial products. The isolated A-type proanthocyanidins from cranberry juice cocktail elicited in vitro anti-adhesion activity at 60 μg/ml, the B-type proanthocyanidins from grape exhibited minor activity at 1200 μg/ml, while other B-type proanthocyanidins were not active. Anti-adhesion activity in human urine was detected following cranberry juice cocktail consumption, but not after consumption of the non-cranberry food products. Results suggest that presence of the A-type linkage in cranberry proanthocyanidins may enhance both in vitro and urinary bacterial anti-adhesion activities and aid in maintaining urinary tract health.
Impacts
Urinary tract infections account for over 11 million doctor visits annually in the U.S. alone. Proanthocyanidins isolated from cranberry fruit exhibit potent bacterial anti-adherence activity in vitro and in vivo. Thus, it is important to determine the degree of structural heterogeneity of cranberry proanthocyanidins to understand how molecular weight and linkage type influence bacterial anti-adherence activity.
Publications
- No publications reported this period
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Progress 01/01/03 to 12/31/03
Outputs
The specific objective was to characterize and compare cranberry proanthocyanidin (PA) fractions that inhibit the adherence of uropathogenic bacteria to uroepithelial cells. Twelve PA fractions were prepared; six PA fractions from an initial separation on Sephadex LH-20 and six C18 subfractions from the LH-20 PA fractions that possessed the greatest anti-adherence activity. Spray dried cranberry juice powder was reconstituted in H2O and applied to a preparative LH-20 column equilibrated in water. Water was passed through the column to elute non-phenolic cranberry constituents. Aqueous acetone (4:1, acetone: H2O, v:v) was then passed through the column until it was white, to elute a crude cranberry PA fraction. The aqueous acetone fraction was concentrated by vacuum to remove the acetone. Cranberry PA was applied to a semi-preparative LH-20 column equilibrated in water. Sequential solvent elution from the column produced 3 PA fractions: LH1; ethanol:water (1:1), LH2;
ethanol, and an aqueous acetone (4:1, acetone: H2O, v:v) fraction. The aqueous acetone fraction was concentrated by vacuum to remove the acetone and reapplied to the LH-20 semi-preparative column equilibrated in water. Sequential solvent elution from the column produced 4 PA fractions: LH3; ethanol:methanol (3:1), LH4; ethanol:methanol (1:1), LH5; methanol and LH6; aqueous acetone (4:1, acetone: H2O, v:v). The PA concentration of each fraction (LH1-LH6) was adjusted to contain 5mg/ml catechin equivalents (CE) as determined by the Folin-Ciocalteu method for determination of total phenolics. Fractions LH3, LH4 and LH5 had the greatest anti-adherence activity and were subjected to reverse phase (RP) C18 subfractionation. LH-20 fractions were concentrated by vacuum to remove organic solvents and reconstituted in water prior to application on a preparative RPC18 column. Two PA sub-fractions (RP1 and RP2) were collected from each LH-20 fraction based on the HPLC-DAD chromatograph at 280 nm,
which indicated the presence of 2 distinct PA humps. Elution from the RPC18 column produced 6 PA fractions: LH3RP1, LH3RP2, LH4RP1, LH4RP2, LH5RP1 and LH5RP2. The PA concentration of all sub-fractions were adjusted to contain 5mg/ml catechin equivalents (CE) as determined by the Folin-Ciocalteu method for determination of total phenolics. Fractions LH3RP2, LH4RP1 and LH4RP2 had the greatest anti-adherence activity. We are in the process of developing a normal phase (LC) separation scheme for these PA fractions and anticipate this step will be accomplished on schedule (5/31/04) as outlined in our previous proposal.
Impacts
Urinary tract infections account for over 11 million doctor visits annually in the U.S. alone. Proanthocyanidins isolated from cranberry fruit exhibit potent bacterial anti-adherence activity in vitro and in vivo. Thus, it is important to determine the degree of structural heterogeneity of cranberry proanthocyanidins to understand how molecular weight and linkage type influence bacterial anti-adherence activity.
Publications
- Krueger, C.G.; M.M. Vestling and J.D. Reed. MALDI-TOF MS demonstrates that the structural diversity of polyflavans in foods and beverages is much greater than previously Appreciated. Abstract and Poster presentation at The 51st Annual Conference on Mass Spectrometry and Allied Topics, June 2-12, 2003 in Montreal, Quebec, Canada.
- Reed, J.D.; Howell, A.; Cunningham, D.; Krueger C.G. Differences in Structure and Bacterial Anti-adhesion Activity of Cranberry Proanthocyanidins Compared to Proanthocyanidins from other Foods. Abstract and Poster presentation at the 1st International Conference on Polyphenols and Health, Nov 12-21, 2003 in Vichy, France.
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Progress 01/01/02 to 12/31/02
Outputs
Cranberry proanthocyanidins were fractionated on LH-20 into 6 fractions by sequential elution. The mass range of the fractions were characterized by matrix assisted laser desorption-time of flight mass spectrometry and for their uropathogenic bacterial anti-adhesion activity. The anti-adhesion activity was strongest in fraction 5 and weakest in fraction 1 .Fraction 5 contained predominantly masses for the hexamer through decamer with more than one A Type interflavan bond. Fraction 4 contained masses that correspond to trimers through septamer with one or more A-Type interflavan bond but the single A-type appeared to be more prevalent than in fraction 5. We will sub-fractionate fractions 4 and 5 since they have the greatest activity and test these sub-fractions for their anti-adhesion activity.
Impacts
Anti-adhesion activity increased with an increase in molecular weight and with fractions that contain more than one A type interflavan bond.
Publications
- No publications reported this period
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