Prevention of Atherosclerosis by Berries: The Case of Blueberries.

Berry consumption has been associated with cardiovascular disease prevention in recent years. Atherosclerosis is one of the major causes of cardiovascular diseases. However, research on the prevention of atherosclerosis through consuming individual whole berries, specifically direct evidence, remains scarce. Therefore, further elucidating the role that berries play in the prevention of atherosclerosis is warranted. In this perspective, blueberries were selected to articulate research strategies for studying atheroprotective effects of berries. Studies from human subjects and various animal models are summarized. The mechanisms by which blueberries may act, through reducing oxidative stress, decreasing inflammation, improving endothelial dysfunction, regulating cholesterol accumulation and trafficking, along with potentially influencing gut microbiota, are also discussed. Blueberries contain high levels of polyphenolic compounds, which were widely indicated as major bioactive compounds. Nonetheless, the metabolites/catabolites after blueberry consumption, such as simple phenolic acids, rather than original compounds in berries, may be the actual in vivo bioactive compounds. Future research should focus on obtaining more direct evidence, preferably in humans, understanding of the mechanisms of action at the molecular level, and identifying bioactive compounds as well as which compounds act synergistically to convey health benefits. The research strategy discussed here may also be applied to the studies of other fruits and berries.

Single-Laboratory Validation for Determination of Total Soluble Proanthocyanidins in Cranberry Using 4-Dimethylaminocinnamaldehyde.

American cranberry (Vaccinium macrocarpon) is native to Eastern North America. Recent studies have suggested that the A-type proanthocyanidins (PACs) in cranberries are effective in preventing urinary tract infection. To meet the growing interest in the cranberry market, an accurate, reliable, and simple method to determine PAC concentration is needed. In this study, a modified method using 4-dimethylaminocinnamaldehyde to quantify total PACs in cranberry products was validated. Cranberry juice extract powder, cranberry capsules containing juice extract, and cranberry juice concentrate were used as the samples in this study. With the modified method, the calibration curves for proanthocyanidin A2 had correlation coefficients (r2) of >0.99. The recoveries of two different concentrations after spiking were 97.1 and 99.1%, and the RSDs for repeatability and reproducibility were <2.7 and <1.6%, respectively.

Stabilization of anthocyanins in blackberry juice by glutathione fortification.

Blackberry anthocyanins provide attractive color and antioxidant activity. However, anthocyanins degrade during juice processing and storage, so maintaining high anthocyanin concentrations in berry juices may lead to greater antioxidant and health benefits for the consumer. This study evaluated potential additives to stabilize anthocyanins during blackberry juice storage. The anthocyanin stabilizing agents used were: glutathione, galacturonic acid, diethylenetriaminepentaacetic acid and tannic acid, which were added at a level of 500 mg L-1. Juice anthocyanin, flavonol, and ellagitannin content and percent polymeric color were measured over five weeks of accelerated storage at 30 °C. Glutathione had the greatest protective effect on total anthocyanins and polymeric color. Therefore a second study was performed with glutathione in combination with lipoic and ascorbic acids in an effort to use antioxidant recycling to achieve a synergistic effect. However, the antioxidant recycling system had no protective effect relative to glutathione alone. Glutathione appears to be a promising blackberry juice additive to protect against anthocyanin degradation during storage.

Urinary excretion of phenolic acids in rats fed cranberry, blueberry, or black raspberry powder.

Dietary polyphenolics can be converted into smaller phenolic acids (PA) by microorganisms in the colon and may contribute to health benefits associated with the parent polyphenolics. Urinary excretion of 18 PA and their conjugates was studied, using HPLC-MS/MS, in rats fed AIN93G-based diets containing 5% (dry weight basis) of either cranberry (CB), blueberry (BB), or black raspberry (BRB). Hippuric, 4-hydroxyphenylacetic, 3-methoxy-4-hydroxyphenylacetic, and 4-hydroxybenzoic acids were excreted in greatest quantity in the urine over a 24 h period in all diets. Primary PA excreted in the berry diets were 4-hydroxycinnamic acid for CB; chlorogenic, ferulic, and 3,4-dihydroxycinnamic acids for BB; and 3-hydroxyphenylpropionic, 3-hydroxybenzoic, and 3-hydroxycinnamic acids for BRB. PA were present in conjugated form with cinnamic acid derivatives being 50-70% and phenylacetic acid derivatives conjugated <10%. Conjugated, and not just the free, PA are significant contributors to total urinary excretion.

Changes in chokeberry (Aronia melanocarpa L.) polyphenols during juice processing and storage.

Chokeberries are an excellent source of polyphenols, but their fate during juice processing and storage is unknown. The stability of anthocyanins, total proanthocyanidins, hydroxycinnamic acids, and flavonols at various stages of juice processing and over 6 months of storage at 25 °C was determined. Flavonols, total proanthocyanidins, and hydroxycinnamic acids were retained in the juice to a greater extent than anthocyanins, with losses mostly due to removal of seeds and skins following pressing. Anthocyanins were extensively degraded by thermal treatments during which time levels of protocatechuic acid and phloroglucinaldehyde increased, and additional losses occurred following pressing. Flavonols, total proanthocyanidins, and hydroxycinnamic acids were well retained in juices stored for 6 months at 25 °C, whereas anthocyanins declined linearly. Anthocyanin losses during storage were paralleled by increased polymeric color values, indicating that the small amounts of anthocyanins remaining were present in large part in polymeric forms.

Improved stability of chokeberry juice anthocyanins by ß-cyclodextrin addition and refrigeration.

Chokeberry anthocyanins are susceptible to degradation during processing and storage of processed products. This study determined the effects of three pH levels (2.8, 3.2, and 3.6) and four ß-cyclodextrin (BCD) concentrations (0, 0.5, 1, and 3%) alone and in combination on the stability of chokeberry juice anthocyanins before and after pasteurization and over 8 months of storage at 4 and 25 °C. Lowering the pH from 3.6 to 2.8 in the absence of BCD provided marginal protection against anthocyanin losses during processing and storage. Addition of 3% BCD at the natural chokeberry pH of 3.6 resulted in excellent protection of anthocyanins, with 81 and 95% retentions after 8 months of storage at 25 and 4 °C, respectively. The protective effect of BCD was lessened with concentrations <3% and reduction in pH, indicating changes in anthocyanin structure play an important role in BCD stabilization of anthocyanins.

Suppression of Wnt1-induced mammary tumor growth and lower serum insulin in offspring exposed to maternal blueberry diet suggest early dietary influence on developmental programming.

Despite the well-accepted notion that early maternal influences persist beyond fetal life and may underlie many adult diseases, the risks imposed by the maternal environment on breast cancer development and underlying biological mechanisms remain poorly understood. In this study, we investigated whether early exposure to blueberry (BB) via maternal diet alters oncogene Wnt1-induced mammary tumorigenesis in offspring. Wnt1-transgenic female mice were exposed to maternal Casein (CAS, control) or blueberry-supplemented (CAS + 3%BB) diets throughout pregnancy and lactation. Offspring were weaned to CAS and mammary tumor development was followed until age 8 months. Tumor incidence and latency were similar for both groups; however, tumor weight at killing and tumor volume within 2 weeks of initial detection were lower (by 50 and 60%, respectively) in offspring of BB- versus control-fed dams. Dietary BB exposure beginning at weaning did not alter mammary tumor parameters. Tumors from maternal BB-exposed offspring showed higher tumor suppressor (Pten and Cdh1) and lower proproliferative (Ccnd1), anti-apoptotic (Bcl2) and proangiogenic (Figf, Flt1 and Ephb4) transcript levels, and displayed attenuated microvessel density. Expression of Pten and Cdh1 genes was also higher in mammary tissues of maternal BB-exposed offspring. Mammary tissues and tumors of maternal BB-exposed offspring showed increased chromatin-modifying enzyme Dnmt1 and Ezh2 transcript levels. Body weight, serum insulin and serum leptin/adiponectin ratio were lower for maternal BB-exposed than control tumor-bearing offspring. Tumor weights and serum insulin were positively correlated. Results suggest that dietary influences on the maternal environment contribute to key developmental programs in the mammary gland to modify breast cancer outcome in adult progeny.

Determination of total antioxidant capacity by oxygen radical absorbance capacity (ORAC) using fluorescein as the fluorescence probe: First Action 2012.23.

An improved method for the measurement of oxygen radical absorbance capacity (ORAC) was developed and validated using fluorescein (3',6'-dihydroxyspiro[isobenzofuran-1[3H], 9'[9H]-xanthen]-3-one) as a new fluorescence probe (ORAC(FL)). Randomly methylated beta-cyclodextrin (RMCD) was introduced as the water-solubility enhancer for lipophilic antioxidants. 7% RMCD (w/v) in 50% acetone-H2O mixture sufficiently solubilized vitamin E compounds and other lipophilic phenolic antioxidants in 75 mM phosphate buffer (pH 7.4). Results indicated that fluorescein shows excellent photostability under the plate reader conditions. This ORAC(FL) was validated through linearity, precision, accuracy, and ruggedness. The validation results demonstrated that the ORACFL assay is reliable and robust. The mean of intraday and interday CVs were <15%; for hydrophilic ORAC, LOD and LOQ are 5 and 6.25 microM, respectively; for lipophilic ORAC, LOD and LOQ are 6.25 and 12.5 microM, respectively. It is concluded that unlike other popular methods, the ORAC(FL) assay provides a direct measure of total antioxidant capacity against the peroxyl radicals.

Effect of dietary blueberry pomace on selected metabolic factors associated with high fructose feeding in growing Sprague-Dawley rats.

An experiment was conducted to study the protective effect of feeding extruded and unextruded blueberry pomace (BBP) on selected metabolic parameters associated with metabolic syndrome in a model of high fructose (HF)-fed growing Sprague-Dawley rats. Treatments were as follows: (1) control (modified AIN-based diet); (2) HF diet (AIN diet with 58% fructose); (3) HF diet with 1.5% unextruded BBP; (4) HF diet with 1.5% extruded BBP; (5) HF diet with 3% unextruded BBP; and (6) HF diet with 3% extruded BBP. Compared with the control, HF feeding increased fasting plasma insulin and fasting and postprandial plasma triglycerides as well as homeostatic scores of insulin resistance and ß-cell function, but not weight gain, diet intake and efficiency, abdominal fat, oral glucose tolerance, and fasting and postprandial plasma glucose, cholesterol, and leptin levels. Inclusion of unextruded or extruded BBP was effective in minimizing or ameliorating the fructose-induced metabolic anomalies, except postprandial plasma triglycerides, especially at 3% of the diet. In addition, unextruded or extruded BBP at 3% of the diet was also able to reduce plasma cholesterol and abdominal fat relative to the HF control, which may impart additional health benefits. Compared with the control, inclusion of unextruded or extruded BBP at both 1.5% and 3% resulted in lower total fat weight, and animals fed a diet supplemented with 3% unextruded BBP in fasting state or 3% unextruded BBP in fed state had lower leptin levels than the control. This is the first study demonstrating the beneficial effects of feeding blueberry pomace on health.

Processing and storage effect on berry polyphenols: challenges and implications for bioactive properties.

Anthocyanins and tannins in blueberries, blackberries and black raspberries are susceptible to degradation during processing, with juices showing the greatest losses due to physical removal of skins and seeds. Anthocyanins and procyanidins are also degraded in processed products stored at ambient temperature with losses accompanied by increased polymeric pigments (PPs). Using chokeberry as a model, formation of PPs occurred in both pasteurized and aged juices and pasteurized juice contained a greater proportion of low molecular weight PPs than aged juice, while aged juice contained a greater proportion of higher molecular PPs. Formation of PP accounts for some of the losses of anthocyanins and procyanidins during processing and storage, but the complete fate of anthocyanins remains unclear. In this review we highlight the steps in processing where significant losses of polyphenols occur, and discuss potential mechanisms responsible for losses, methods to mitigate losses, and implications on bioactive properties.

Repression of mammosphere formation of human breast cancer cells by soy isoflavone genistein and blueberry polyphenolic acids suggests diet-mediated targeting of cancer stem-like/progenitor cells.

Mammary stem cells are undifferentiated epithelial cells, which initiate mammary tumors and render them resistant to anticancer therapies, when deregulated. Diets rich in fruits and vegetables are implicated in breast cancer risk reduction, yet underlying mechanisms are poorly understood. Here, we addressed whether dietary factors selectively target mammary epithelial cells that display stem-like/progenitor subpopulations with previously recognized tumor-initiating potential. Using estrogen receptor-positive MCF-7 and estrogen receptor-negative MDA-MB-231 human breast cancer cell lines and freshly isolated epithelial cells from MMTV-Wnt-1 transgenic mouse mammary tumors, we demonstrate that sera of adult mice consuming soy isoflavone genistein (GEN) or blueberry (BB) polyphenol-containing diets alter the population of stem-like/progenitor cells, as measured by their functional ability to self-renew and form anchorage-independent spheroid cultures in vitro at low frequency (1-2%). Serum effects on mammosphere formation were dose-dependently replicated by GEN (40 nM >2 µM) and targeted the basal stem-like CD44+/CD24-/ESA+ and the luminal progenitor CD24+ subpopulations in MDA-MB-231 and MCF-7 cells. GEN inhibition of mammosphere formation was mimicked by the Akt inhibitor perifosine and was associated with enhanced tumor suppressor phosphatase and tensin homologue deleted on chromosome ten (PTEN) expression. In contrast, a selected mixture of BB phenolic acids was only active in MDA-MD-231 cells and its CD44+/CD24-/ESA+ subpopulation, and this activity was independent of induction of PTEN expression. These findings delineate a novel and selective function of distinct dietary factors in targeting stem/progenitor cell populations in estrogen receptor-dependent and -independent breast cancers.

Effect of feeding grape pomace on selected metabolic parameters associated with high fructose feeding in growing Sprague-Dawley rats.

The effect of feeding grape pomace on certain metabolic parameters associated with high fructose (HF) feeding was studied. Forty male growing Sprague-Dawley rats were randomly assigned into groups: (1) control; (2) HF; (3) HF with low-level (1.5% of diet) grape pomace (HF+LP), and (4) HF with high-level (5.0% of diet) grape pomace (HF+HP). The HF+LP and HF+HP diets provided 115 and 218 mg of procyanidins/kg, respectively. Compared with the controls, HF-fed animals consumed less and were smaller, whereas animals in the HF+LP and HF+HP groups were in between. A similar trend was observed for abdominal fat and abdominal fat as a percentage of body weight. No change in heart or kidney weight occurred. Liver weight as a percentage of body weight was higher for animals when fructose was included in the diet compared with those on control diet, and inclusion of grape pomace had no effect. Fasting plasma glucose, insulin, and triglyceride levels tended to be higher in animals fed HF diet, and grape pomace reduced their levels to values similar to the control animals. Compared with control animals, HF-fed animals had higher weekly postprandial plasma triglycerides, which were reduced by feeding grape pomace, but no change in plasma cholesterol was observed. Glucose intolerance was observed in animals fed HF diet and was accompanied by a 25% increase in homeostatic model assessment (HOMA) of insulin resistance. Inclusion of grape pomace increased glucose tolerance and insulin sensitivity. No significant change (P>.1) in HOMA of ß-cell function or Quantitative Insulin-Sensitivity Check Index was observed. Overall, HF diet did not produce as strong a response of metabolic syndrome as has been shown in the literature. The inclusion of grape pomace in the diet was effective in modulating some aspects of metabolic parameters associated with metabolic syndrome, and the higher level of grape pomace in the diet produced a slightly better response than the lower level.

Supercritical CO2 decaffeination of unroasted coffee beans produces melanoidins with distinct NF-κB inhibitory activity.

UNLABELLED: The supercritical CO(2)-decaffeination process causes unroasted coffee beans to turn brown. Therefore, we suspected that the decaffeinated beans contained melanoidins. Decaffeinated unroasted coffee extract absorbed light at 405 nm with a specific extinction coefficient, K(mix 405 nm), of 0.02. Membrane dialysis (molecular weight cut-off, 12 to 14 kDa) increased the K(mix 405 nm) value 15 fold. Gel filtration chromatography showed that the high-MW fraction (MW > 12 kDa) had an elution profile closer to that of melanoidins of medium-roast coffee than to the corresponding fraction of unroasted coffee, indicating the presence of melanoidins in decaffeinated unroasted beans. Using murine myoblast C2C12 cells with a stably transfected nuclear factor-κB (NF-κB) luciferase reporter gene, we found that the high-MW fraction of decaffeinated unroasted beans had an NF-κB inhibitory activity of IC(50) = 499 µg/mL, more potent than that of regular-roast coffee (IC(50) = 766 µg/mL). Our results indicate that melanoidins form during the supercritical CO(2)-decaffeination process and possess biological properties distinct from those formed during the regular roasting process. PRACTICAL APPLICATION: We discovered the roasting effect of decaffeination process, reporting the discovery of melanoidins in green (unroasted) decaf coffee beans. Our results indicated that melanoidins form during the supercritical CO2-decaffeination process and possess biological properties distinct from those formed during the regular roasting process. Our results offer new insights into the formation of bioactive coffee components during coffee decaffeination process.

Impact of different stages of juice processing on the anthocyanin, flavonol, and procyanidin contents of cranberries.

Juice is the most common form in which cranberries are consumed; however there is limited information on the changes of polyphenolic content of the berries during juice processing. This study investigated the effects of three different pretreatments (grinding plus blanching; only grinding; only blanching) for cranberry juice processing on the concentrations of anthocyanins, flavonols, and procyanidins throughout processing. Flavonols and procyanidins were retained in the juice to a greater extent than anthocyanins, and pressing resulted in the most significant losses in polyphenolics due to removal of the seeds and skins. Flavonol aglycones were formed during processing as a result of heat treatment. Drying of cranberry pomace resulted in increased extraction of flavonols and procyanidin oligomers but lower extraction of polymeric procyanidins. The results indicate that cranberry polyphenolics are relatively stable during processing compared to other berries; however, more work is needed to determine their fate during storage of juices.

Processing and storage effects on the ellagitannin composition of processed blackberry products.

Changes in blackberry ellagitannin composition in response to juicing (clarified and nonclarified), pureeing, canning (in water or syrup), and freezing as well as changes in processed products during 6 months of storage were investigated. Canning, pureeing, and freezing had little effect on ellagitannins, but processing berries into nonclarified and clarified juices resulted in total ellagitannin losses of 70 and 82%, respectively, due to removal of ellagitannin-rich seeds in the presscake. Minimal changes in total ellagitannin content were observed during storage of thermally processed products, but compositional changes indicative of ellagitannin depolymerization were apparent. The ellagitannin content and composition of frozen berries remained stable over 6 months of storage. Ellagitannins are well retained in canned, pureed, and frozen blackberries, but methods are needed to prevent losses during juice processing and/or exploit the ellagitannin-rich coproducts.

Urinary excretion of (Epi)catechins in rats fed different berries or berry products.

(Epi)catechins are associated with many health benefits in humans. However, their bioavailability, excretory pattern, and extent of conjugation in animals fed different sources or levels in the diet are not well documented. Two experiments were conducted to investigate the urinary excretion of (epi)catechins after feeding of different types of berries or different levels of the same berry source to rats. Experiment 1 investigated the effects of feeding a commercially available concentrated cranberry powder (CCP) at three different levels, 3.3, 6.6, and 33 g/kg of diet, whereas experiment 2 investigated the effect of feeding freeze-dried whole cranberry (CB), blueberry (BB), or black raspberry (BRB) powder at 50 g/kg of diet. Both experiments had an AIN-93-based control and a high-fructose diet (53-65% of the diet) to which was added three levels of CCP in experiment 1 and CB, BB, and BRB in experiment 2. (Epi)catechins were excreted as free and conjugated in both intact and methylated forms. Excretion of conjugated (epi)catechins was as high as 60% of the total consumed in some cases. A majority of both catechins and epicatechins excreted in the urine was in a methylated form. Excretion of epicatechins, including their methylated forms, ranged from 30 to 47% of the ingested amount, whereas that of catechins, including their methylated forms, ranged from 9 to 31%. Urinary excretion of (epi)catechins was dose dependent and increased with the amount of (epi)catechins present in the diet. On the basis of the excretory pattern of (epi)catechins in the urine, data suggested that the bioavailability of epicatechins may be higher than that of catechins and that (epi)catechins may be more available from blueberries compared to cranberries.

The purple cauliflower arises from activation of a MYB transcription factor.

Anthocyanins are responsible for the color of many flowers, fruits, and vegetables. An interesting and unique Purple (Pr) gene mutation in cauliflower (Brassica oleracea var botrytis) confers an abnormal pattern of anthocyanin accumulation, giving the striking mutant phenotype of intense purple color in curds and a few other tissues. To unravel the nature of the Pr mutation in cauliflower, we isolated the Pr gene via a combination of candidate gene analysis and fine mapping. Pr encoded a R2R3 MYB transcription factor that exhibited tissue-specific expression, consistent with an abnormal anthocyanin accumulation pattern in the mutant. Transgenic Arabidopsis (Arabidopsis thaliana) and cauliflower plants expressing the Pr-D allele recapitulated the mutant phenotype, confirming the isolation of the Pr gene. Up-regulation of Pr specifically activated a basic helix-loop-helix transcription factor and a subset of anthocyanin structural genes encoding flavonoid 3'-hydroxylase, dihydroflavonol 4-reductase, and leucoanthocyanidin dioxygenase to confer ectopic accumulation of pigments in the purple cauliflower. Our results indicate that the genetic variation including a Harbinger DNA transposon insertion in the upstream regulatory region of the Pr-D allele is responsible for the up-regulation of the Pr gene in inducing phenotypic change in the plant. The successful isolation of Pr provides important information on the regulatory control of anthocyanin biosynthesis in Brassica vegetables, and offers a genetic resource for development of new varieties with enhanced health-promoting properties and visual appeal.

Release of bound procyanidins from cranberry pomace by alkaline hydrolysis.

Procyanidins in plant products are present as extractable or unextractable/bound forms. We optimized alkaline hydrolysis conditions to liberate procyanidins and depolymerize polymers from dried cranberry pomace. Alkaline extracts were neutralized (pH 6-7) and then procyanidins were extracted with ethyl acetate and analyzed by normal phase high performance liquid chromatography. Alkaline hydrolysis resulted in an increase in low molecular weight procyanidins, and the increase was greater at higher temperature, short time combinations. The most procyanidins (DP1-DP3) were extracted at 60 degrees C for 15 min with each concentration of NaOH. When compared to conventional extraction using homogenization with acetone/water/acetic acid (70:29.5:0.5 v/v/v), treatment with NaOH increased procyanidin oligomer extraction by 3.8-14.9-fold, with the greatest increase being DP1 (14.9x) and A-type DP2 (8.4x) procyanidins. Alkaline treatment of the residue remaining after conventional extraction resulted in further procyanidin extraction, indicating that procyanidins are not fully extracted by conventional extraction methods.

Multi-laboratory validation of a standard method for quantifying proanthocyanidins in cranberry powders.

BACKGROUND: The objective of this study was to validate an improved 4-dimethylaminocinnamaldehyde (DMAC) colorimetric method using a commercially available standard (procyanidin A2), for the standard method for quantification of proanthocyanidins (PACs) in cranberry powders, in order to establish dosage guidelines for the uropathogenic bacterial anti-adhesion effect of cranberry. RESULTS: Commercially available cranberry samples were obtained (five from U.S. sources and six from European sources) for PAC quantification in five different analytical laboratories. Each laboratory extracted and analyzed the samples using the improved DMAC method. Within-laboratory variation (mean +/- SD) was 4.1 +/- 1.7% RSD (range, 2.3-6.1% RSD) and the between laboratory variability was 16.9 +/- 8.5% RSD (range, 8-32% RSD). For comparative purposes, the cranberry samples were alternatively quantified using weights of extracted PACs (gravimetric). The correlation coefficient between the two methods was 0.989. CONCLUSION: This improved DMAC method provides a simple, robust and relatively specific spectrophotometric assay for total PACs in cranberry samples using commercially available procyanidin A2 dimer as a standard. DMAC is most useful within a given type of food such as cranberries, but may not be appropriate for comparing concentrations across different food types, particularly in those cases where large differences exist among the relative amounts of each oligomer and polymer.

Dietary black raspberry anthocyanins do not alter development of obesity in mice fed an obesogenic high-fat diet.

Anthocyanins (ACNs) from various foods have been shown to minimize the development of obesity in some animal models. The objective of the current study was to compare the effects of feeding purified black raspberry (BRB) ACNs or the freeze-dried whole BRB on the development of obesity. Male C57BL/6J mice (25 days of age) were assigned at random to treatments (7/treatment; 3/cage). The treatments included (1) control low-fat diet (10% calories from fat) (LF); (2) LF plus BRB juice in place of drinking water; (3) LF diet plus purified BRB ACNs in drinking water (1.25 mg/mL); (4) control high-fat diet (60% calories from fat) (HF60); (5) HF60 diet + BRB juice in place of drinking water; (6) HF60 diet + ACNs in drinking water (1.25 mg/mL); and (7) HF60 + freeze-dried whole BRB powder (21.7 g/kg of diet). Body weight gains in mice fed HF60 diet plus purified BRB ACNs tended to be lower after 56, 63, and 70 days than in mice fed HF60 alone. Body weights were increased at time of sacrifice, but heart, liver, and kidney weights as a percentage of body weight were decreased in mice fed HF60 diet compared to LF fed mice. Weights (g or g/body weight) of epididymal and retroperitoneal fat were increased in the HF60 fed mice compared to LF fed mice. Fasting serum glucose, leptin, and insulin levels as well as homeostasis assessment of insulin resistance (HOMA-IR) were elevated in mice fed the HF60 diet relative to LF-fed controls. Serum cholesterol, triglycerides, and monocyte chemoattractant protein-1 (MCP-1) were not altered by diet. Serum levels of resistin were increased in mice fed the HF60 diet compared to mice fed the LF diet. None of the responses measured were altered by whole BRB powder included in the diet relative to the HF60 control diet. Cyanidin containing di- or triglycosides in BRB was ineffective in altering the development of obesity in contrast to cyanidin-monoglycosides, which have been shown to be effective. The sugar moiety on the anthocyanidins may be an important factor in determining the response in the development of obesity.