Mechanism of procyanidins for health functionality by improving the intestinal environment.

Procyanidins are one of the polyphenols consisting of multiple flavan-3-ols (eg epicatechin). They have a complex chemical structure, with the degree of polymerization and linked position of flavan-3-ols varying among various foods, such as apples and chocolate. Physiological functional studies of procyanidins have investigated their mechanisms in cells and animals based on their antioxidant effects. Recently, the intestinal environment, including the intestinal microflora, has played an important role in the energy metabolism and health status of the host. Regulation of the intestinal environment by dietary polyphenols is becoming a new concept in health functions, and we have begun to investigate the mechanism of apple procyanidins, focusing on the gut microbiota and metabolites in our functional research. In this minireview, we will discuss the effects of procyanidin ingestion on the gut microbiota and metabolites.

The Differences in the Flavan-3-ol and Procyanidin Contents of the Japanese 'Fuji' and 'Orin' Apples Using a Rapid Quantitative High-Performance Liquid Chromatography Method: Estimation of the Japanese Intake of Flavan-3-ols and Procyanidins from Apple as Case Study.

Previously, we reported that apple polyphenols and their major active compounds, the flavan-3-ols and the procyanidins, can result in various health benefits in animals and humans, according to clinical studies. Here, we developed a rapid method for quantifying flavan-3-ols and procyanidins using high-performance liquid chromatography with fluorescence detection, where we investigated the amounts of flavan-3-ols and procyanidins in the Japanese major apple production centre, the Aomori Prefecture, from 2016 to 2018. The non-bagged 'Fuji (n = 609)', the bagged 'Fuji (n = 1101)', and the 'Orin (n = 504)' apples were evaluated in terms of their differences in flavan-3-ols and procyanidins based on apple variety and the controlled atmosphere storage. The bagging treatments of the 'Fuji' apples resulted in significantly higher concentrations of procyanidins, while changes in flavan-3-ols concentrations were not clearly observed by treatment. In addition, 'Orin' had a significantly higher concentration of procyanidins than that of 'Fuji'. In contrast, the controlled atmosphere storage hardly caused any changes in the flavan-3-ol and procyanidin contents. Hence, we present the concentrations of flavan-3-ols and procyanidins in major Japanese apples using the rapid high-performance liquid chromatography method with fluorescence detection.

Administration of Apple Polyphenol Supplements for Skin Conditions in Healthy Women: A Randomized, Double-Blind, Placebo-Controlled Clinical Trial.

This clinical study was performed to evaluate the effects of continuous apple polyphenol (AP) administration on facial skin conditions and pigmentation induced by ultraviolet (UV) irradiation in healthy women participants. Participants (n = 65, age 20-39 years) were randomized to receive tablets containing AP (300 or 600 mg/day) or placebo in a double-blinded, placebo-controlled clinical trial. Continuous administration of AP for 12 weeks significantly prevented UV irradiation induced skin pigmentation (erythema value, melanin value, L value), although a dose-dependent relationship was not clearly observed. In contrast, no significant differences were detected between the groups with regard to water content and trans-epidermal water loss. Our study demonstrated that APs and their major active compounds, procyanidins, have several health benefits. Here, we report that continuous administration of AP for 12 weeks alleviated UV irradiation induced skin pigmentation, when compared with placebo, in healthy women.

Chronic administration of apple polyphenols ameliorates hyperglycaemia in high-normal and borderline subjects: A randomised, placebo-controlled trial.

AIMS: We previously reported that apple polyphenols (AP) and their major active components, procyanidins, had beneficial effects on glucose homeostasis and diabetes in diabetic ob/ob mice. The present study was performed to evaluate the effects of chronic AP administration on glucose tolerance in high-normal and borderline human subjects. METHODS: Subjects (n=65) with a fasting plasma glucose (FPG) level of 100-125mg/dL determined during a recent health check-up were randomised to receive tablets containing AP (600mg/day) or placebo tablets for 12weeks in a double-blinded, placebo-controlled clinical trial. The primary outcome was insulin resistance, assessed using a 75g oral glucose tolerance test (OGTT). RESULTS: The 12-week chronic administration of AP significantly reduced the increase in glucose at 30-min post-75g OGTT (OGTT30-min glucose) value, compared to the placebo regimen. Furthermore, in a subgroup of the high-normal (FPG value, 100-109mg/dL; 2-h post-75g OGTT glucose (OGTT2-h glucose) value, <140mg/dL) and borderline (FPG value, 110-125mg/dL; OGTT2-h glucose value, <140mg/dL and FPG value, <126mg/dL; OGTT2-h glucose value, 140-199mg/dL) subjects, OGTT30-min glucose value in the AP group (164.0±7.4mg/dL) was significantly lower than that of the placebo group (194.7±10.4mg/dL, p<0.05). No significant changes in the other lipid parameters and cytokine levels were observed. CONCLUSIONS: Chronic AP administration significantly improved impaired glucose tolerance in high-normal and borderline subjects. Larger and/or longer-term scale human studies are required to confirm the potential glucose homeostasis of AP.

Regression of atherosclerosis with apple procyanidins by activating the ATP-binding cassette subfamily A member 1 in a rabbit model.

BACKGROUND AND AIMS: Apple polyphenol contains abundant procyanidins, which have been associated with an anti-atherosclerosis and cholesterol-lowering effect. The aim of this study was to investigate whether apple procyanidins (APCs) feature therapeutic efficacy in terms of regressing atherosclerosis and whether this efficacy is due to mechanisms other than a cholesterol-lowering effect. METHODS: After eight weeks on an atherogenic diet, rabbits were given a normal diet for another eight weeks to normalize the increased serum lipids level. The rabbits in the baseline group were sacrificed at this stage. The control group was subsequently fed a normal diet for eight weeks, while the APCs group was administrated 50 mg/kg/day of APCs in addition to the normal diet. Serum lipids and aortic intimal-medial thickness (IMT) were serially examined, and the resected aorta was examined histologically and through molecular biology. RESULTS: Aortic IMT on ultrasonography and the lipid accumulation area examined using Sudan IV staining were significantly reduced in the APCs group as compared to the control group. Serum lipid profiles were not different between the groups. Immunohistochemistry showed significantly decreased staining of an oxidative stress marker and significantly increased staining of ATP-binding cassette subfamily A member 1 (ABCA1) in the APCs group. Western blotting and RT-PCR also showed increased expression of ABCA1 mRNA and its protein in the APCs group. CONCLUSIONS: This study revealed that APCs administration causes a regression of atherosclerosis. APCs might hold promise as an anti-atherosclerotic agent.

Oral Administration of Apple Procyanidins Ameliorates Insulin Resistance via Suppression of Pro-Inflammatory Cytokine Expression in Liver of Diabetic ob/ob Mice.

Procyanidins, the main ingredient of apple polyphenols, are known to possess antioxidative and anti-inflammatory effects associated closely with the pathophysiology of insulin resistance and type 2 diabetes. We investigated the effects of orally administered apple procyanidins (APCs) on glucose metabolism using diabetic ob/ob mice. We found no difference in body weight or body composition between mice treated with APCs and untreated mice. A 4 week oral administration of APCs containing water [0.5% (w/v)] ameliorated glucose tolerance, insulin resistance, and hepatic gluconeogenesis in ob/ob mice. APCs also suppressed the increase in the level of the pancreatic ß-cell. Insulin-stimulated Akt phosphorylation was significantly enhanced; pro-inflammatory cytokine expression levels were significantly decreased, and c-Jun N-terminal kinase phosphorylation was downregulated in the liver of those mice treated with APCs. In conclusion, APCs ameliorate insulin resistance by improving hepatic insulin signaling through suppression of hepatic inflammation in ob/ob mice, which may be a mechanism with possible beneficial health effects of APCs in disturbed glucose metabolism.

Prevention of allergic disease development and symptoms by food factors.

A fundamental means of allergic disease prevention, via the use of functional food factors, is desirable. A number of studies on the role of functional food factors in preventing allergic diseases have been reported. In this review, the preventive effects of polyphenols, carotenoids, polysaccharides, and non-digestible oligosaccharides on allergic diseases are discussed.

Cloning of a novel O-methyltransferase from Camellia sinensis and synthesis of o-methylated EGCG and evaluation of their bioactivity.

The gene of a novel O-methyltransferase was isolated from tea cultivars (Camellia sinensis L.). Using the recombinant enzyme, O-methylated (-)-epigallocatechin-3-O-gallate (EGCG) in all cases were synthesized. EGCG and the synthesized O-methylated EGCGs including (-)-epigallocatechin-3-O-(3-O-methyl)-gallate (EGCG3''Me), (-)-epigallocatechin-3-O- (4-O-methyl)-gallate(EGCG4''Me), (-)-epigallocatechin-3-O-(3,5-O-dimethyl)-gallate (EGCG3'',5''diMe), and (-)-3-O-methyl-epigallocatechin-3-O-(3,5-O-dimethyl)-gallate (EGCG3',3'',5''triMe) were assayed using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay and antibacterial activity. EGCG was the most effective of the O-methylated EGCGs. The antiallergic effects of EGCG and the other O-methylated EGCGs were measured by conducting histamine release assays using bone marrow-derived mouse mast cells, and the order of potency was EGCG3',3'',5''triMe = EGCG3'',5''diMe > EGCG3''Me > EGCG. These results indicated that reducing the number of hydroxyl groups decreases the effectiveness of DPPH radical scavenging and antibacterial activity. In contrast, the inhibition of histamine release was potentiated by an increase in the number of methyl groups in EGCG, especially in the galloyl moiety.

Apple procyanidins induce tumor cell apoptosis through mitochondrial pathway activation of caspase-3.

Various epidemiologic and experimental in vivo and in vitro studies have suggested that polyphenols derived from fruits, vegetables and beverages might decrease the risk of developing lifestyle diseases, such as cardiovascular disorders and cancer. Apples are a major dietary source of polyphenols. Here we investigated the antitumor activity of apple polyphenols (APs) and procyanidins, namely condensed tannins, both in vitro and in vivo studies. APs and procyanidins inhibited the growth of transplanted B16 mouse melanoma cells and BALB-MC.E12 mouse mammary tumor cells, and increased the survival rate of the host mice-transplanted B16 cells. Among the APs, the apple procyanidins specifically, rather than other polyphenols such as chlorogenic acid, (-)-epicatechin, phloridzin and procyanidin B2, had a major effect on cell proliferation and induced apoptosis in vitro. The apple procyanidins increased mitochondrial membrane permeability and cytochrome c release from mitochondria and activated caspase-3 and caspase-9 within the tumor cells. In addition, we separated eight procyanidin fractions according to the degree of polymerization using normal-phase chromatography, and detected strong anti-tumor activity in the procyanidin pentamer and higher degree fractions. Our results indicate that the oral administration of apple procyanidins inhibits the proliferation of tumor cells by inducing apoptosis through the intrinsic mitochondrial pathway.

Oligomeric procyanidins in apple polyphenol are main active components for inhibition of pancreatic lipase and triglyceride absorption.

Inhibitory effects of apple polyphenol extract (AP) and procyanidin contained in AP on in vitro pancreatic lipase activity and in vivo triglyceride absorption in mice and humans were examined. AP and procyanidin considerably inhibited in vitro pancreatic lipase activity. However, polyphenols, except for procyanidin, in AP (i.e., catechins, chalcones, and phenol carboxylic acids) showed weak inhibitory activities on pancreatic lipase. Procyanidins separated by normal-phase chromatography according to the degree of polymerization were also examined. Inhibitory effects of procyanidins increased according to the degree of polymerization from dimer to pentamer. On the other hand, pentamer or greater procyanidins showed maximal inhibitory effects on pancreatic lipase. These results suggested that with respect to in vitro pancreatic lipase inhibition, the degree of polymerization was an important factor and oligomeric procyanidin mainly contributed. Next, we performed a triglyceride tolerance test in mice and humans. Simultaneous ingestion of AP and triglyceride significantly inhibited an increase of plasma triglyceride levels in both models. These results suggested that the oligomeric procyanidins contained in AP inhibited triglyceride absorption by inhibiting pancreatic lipase activity in mice and humans.

Dietary unripe apple polyphenol inhibits the development of food allergies in murine models.

The incidence of type I allergic disorders has been increasing worldwide, particularly, the hypersensitivity to food. We first showed that apple condensed tannin (ACT) intake would inhibit the development of the oral sensitization and that the inhibition could correlate with the rise in the population of TCR(gamma)delta-T cells in the intestinal intraepithelial lymphocytes (IEL) using W/W(V) mice and B10A mice which were ovalbumin (OVA)-orally sensitized. Serum OVA-specific immunoglobulin E and immunoglobulin G1 titers in the OVA-orally sensitized W/W(V) and B10A mice ad libitium fed ACT were extremely inhibited compared to those of the control. The ACT intakes of OVA-sensitized W/W(V) and B10A mice inhibited the immediate reduction of the body temperature or the rise in serum histamine induced by active systemic anaphylaxis. The proportions of the TCR(gamma)delta-T cells in the IEL of the OVA-orally sensitized W/W(V) and B10A mice ad libitium fed ACT were significantly greater than that in the controls. Furthermore, ACT feeding by itself could induce the rise in the percentage of the TCR(gamma)delta-T cells among the IEL of the W/W(V) and B10A mice. This suggests that the ACT intake may prevent the development of food allergies and this effect could be correlated with the rise in the percentage of TCR(gamma)delta-T cells among the IEL.

Procyanidin trimers to pentamers fractionated from apple inhibit melanogenesis in B16 mouse melanoma cells.

The effects of apple polyphenols on melanogenesis in B16 mouse melanoma cells were investigated. The inhibitory effect of apple polyphenols was stronger than that of arbutin or kojic acid. Three polyphenol fractions (phenolic acid derivatives, procyanidins and other flavonoids) were isolated, and the procyanidins were fractionated according to the degree of polymerization using normal-phase chromatography. The procyanidin trimer-to-pentamer fractions were found to have the most pronounced effect on melanogenesis. Furthermore, each procyanidin fraction inhibited mushroom tyrosinase. No correlation between the degree of procyanidin polymerization and tyrosinase inhibitory activity was observed. Nevertheless, these observations suggest that procyanidins are effective inhibitors of tyrosinase.

Separation of proanthocyanidins by degree of polymerization by means of size-exclusion chromatography and related techniques.

The molecular masses of polyphenols in plants and food vary greatly up to the order of 10 kDa. Polymerized polyphenols are not only natural antioxidants but also strong inhibitors of numerous physiological enzymatic activities. Several useful methods for the determination and separation of these high-molecular-mass polyphenols have recently been developed. In this review, details of the methods and applications of size-exclusion chromatographic separation of polymerized polyphenols, particularly those of proanthocyanidins, are described and compared with other related chromatographic or mass spectrometric analyses.

Characterization of polymerized polyphenols by size-exclusion HPLC.

Various kinds of high-molecular-mass polyphenols such as condensed tannins, hydrolyzable tannins, and polymerized anthocyanins, were readily characterized by a new size-exclusion HPLC method. This rapid analytical method may also be useful for the profiling of molecular mass distribution of polyphenolic constituents in many kinds of food materials.