Health-related Compounds in Muscadine Grapes

As a lead-up to our Annual Muscadine Field Day on Saturday, Dr. Donna Marshall provided me a sneak preview on what she will be discussing.  See it below in great detail!

Storage Retention of Stilbene, Ellagic acid, Flavonol, and Phenolic content of Muscadine Grape Cultivars.

Dr. Donna Marshall, USDA-ARS Thad Cochran Southern Horticulture Laboratory, Poplarville, MS

Stilbenes are a small class of phenylpropanoids characterized by a 1,2-diphenylethylene backbone.  Stilbenes are synthesized in grape berries under natural environmental conditions (Jeandet, et al., 1999), but are increased by the up-regulation of defense genes encoding pathogenesis-related proteins (Chong, et al., 2009).   The cis- and trans-isomers of resveratrol, a pharmacologically important stilbene, are present in the skin during all ripening stages, but are almost totally absent from the pulp (Chong, et al. 2009).   Specific accumulation of resveratrol in the berry skin results from the localization of stilbene synthase (STS), the pivotal enzyme for stilbene biosynthesis. As a defense mechanism, stilbenes display potent antifungal effects as well as function in dormancy and growth inhibition in plants (Croteau, et al., 2000).  From a pharmacological perspective, scientists have been reporting for over a decade the various ways that the stilbene, resveratrol, can positively affect health (Arichi, et al., 1982, Brakenhiem et al., 2001, DeSanti, et al., 2000a, DeSanti, et al., 2000b, El-Mowafy, 2002, Jang, et al., 1997, Kimura et al., 1985, Kinsella, et al., 1993, Lu and Sorreno, 1999). Piceid, resveratrol 3-O-β-D-glucoside, also exhibits activity comparable to resveratrol (Romero-Perez, et al., 1999)  Resveratrol and piceid exist in the cis form, which is an isomer of the trans form. In early studies trans-resveratrol (TRes) was shown to inhibit platelet aggregation, inhibit the oxidation of low-density lipoproteins, reduce the level of triacylglycerol and protect the liver from lipid peroxidation (Romero-Perez, 1999). Since glycosidase is known to be present in the digestive tract, it is possible that piceid could be converted to resveratrol and absorbed during digestion (Hackett, 1986).  Therefore it is important to consider all isomers and glucosides of TRes.

Ellagic acid is commonly present in other fruits, such as raspberry, strawberry, and blackberry, but is absent in all other Vitis species.  Ellagic acid in muscadine grapes is expressed as free ellagic acid, ellagic acid glycosides, and ellagitannins (Talcott and Lee, 2002).   The presence of ellagic acid and its derivatives in plants is of interest and has been widely studied because of its antiproliferative and antioxidant properties. The antiproliferative properties are due to its ability to directly inhibit DNA binding of certain carcinogens, including nitrosamines and polycyclic aromatic hydrocarbons (PAHs) (Lesca, 1983).  Ellagic acid also has a chemoprotective effect in cellular models by reducing oxidative stress (Tallcott and Lee, 2002, Lesca, 1983. Patrana-Bonilla, 2003, Mertens-Talcott, et al., 2003, Stoner and Morse, 1997, Khanduja, 1999) . Ellagic acid and its derivatives are being aggressively investigated for chemoprevention.  It has been demonstrated that extracts of fruits containing ellagic acid derivatives are more powerful than individual substances. This is because there are multiple phenolic substances in a fruit that can act synergistically with the ellagic acid compounds within the biological processes that involve cancer initiation and growth  (Mertens-Talcott et al., 2003, Mertens-Talcott and Percival, 2005, Mertens-Talcott, et al., 2005)  The presence of ellagic acid and quercetin in muscadine grape could add value and marketability to the crop due to the possible health benefits (Patel, 1955, Talcott and Lee, 2002).

Phenylpropanoids, or phenolics are a large family of secondary metabolites involved in plant response to abiotic and biotic stresses.  Phenolics are ubiquitous in the plant kingdom and are the most abundant secondary metabolites found in plants (Amakura, 2000).  Many phenolics not only protect the parent plant, but also exhibit significant pharmacological benefits.  Phenolic compounds play an important role in overall food properties, as they are generally involved in defense against ultraviolet radiation or as phytoalexins (Amakura, et al., 2000, Singleton, 1980, Zaat et al., 1987, Dixon, 1986).  Phenolics may also play a role in the regulation of plant metabolism (Laks and Pruner, 1989).  Polyphenols represent the third most abundant constituent in grapes and wines after carbohydrates and fruit acids (Singleton, 1980).  Phenolics are mainly distributed as 28-35% in skin, 60-70% in seed and less than 10% in the pulp of the grape (Shi, et al., 2003).  Phenolics contribute to the bitterness and astringency of fruits and are also considered to be the most important compounds affecting flavor and color differences in wines.  Analysis of total phenolics is used to estimate the antioxidant capacities of fresh fruits and vegetables (Thaipong, et al., 2006).  In a study of anticancer activities of muscadine grape phenolics, 50% inhibition of cancer cell population growth for colon cancer cell lines was observed at concentrations of 1-7 mg/mL of muscadine skin extracts (Yi, et al., 2005). Hudson, et al. (2007) found that muscadine grape skin extract inhibited tumor cell growth in all transformed prostate cancer cell lines tested.  The higher total phenolic content present in muscadine grapes compared to other Vitis species is attributed to high ellagic acid, gallic acid, and flavonoid glycoside concentrations (Talcott and Lee, 2002. Lesca, 1983, Patrana-Bonilla, et al., 2003,  Mertens-Talcott, et al., 2003, Yilmaz and Toledo, 2004).

Another unique attribute of muscadine fruit chemistry is the present of anthocyanins as 3,5-diglucosides of delphinidin, cyanidin, petunidin, peonidin, and malvidin in non-acylated forms (Flora, 1978, Goldy et al., 1986, Lamikanra, 1988).   Though absorption of anthocyanins appears to be low in humans (Prior, 2004), it seems likely that cells in which they function in defense of oxidative stress must concentrate the anthocyanins or one of their derivatives (Galli, et al., 2002).  Anthocyanins are known to protect blood vessels in humans. They also play a role in cancer prevention. There are more than 80 publications that discuss the ability of different anthocyanins to prevent different kinds of cancer (Hartle, et al., 2005).

Also unique to muscadine grapes is the presence of myricetin in the bronze grapes, as this flavonol is not present in white V. vinifera grapes (Flora, 1978).   Flavonols are extensively studied compounds found in muscadine grapes.  In humans, protection against carcinogenesis is a widely documented effect of flavonols (Williamson, and Manach, 2005).

One of the most touted possible effects of myricetin is in fighting cancer. As an antioxidant, it assists the body in getting rid of substances called free radicals that are involved in causing many types of cancer including prostate and lung cancer. The body’s metabolism of flavonoids may also help the body eliminate other carcinogens; the effects on enzymes may also have effects on the rate of cancer growth. Study of these effects is ongoing.

Another reputed effect of myricetin is the reduction of cholesterol. It is supposed to inhibit the uptake, or absorption, of low density lipoprotein (LDL) cholesterol which is harmful to a person’s health. It also is thought to prevent a process called oxidation of the LDL cholesterol which makes it more harmful and able to damage various body tissues. Several other flavonoids are also supposed to have similar cholesterol reduction effects.

Historically used in the treatment of fever, Myricetin is also supposed to be effective as an anti-inflammatory substance. It is thought to affect the actions of a few of the enzymes that contribute to inflammation. It has also been used to treat diarrhea, particularly in children. It has no known negative side effects and is generally considered safe for use as a supplement. Anyone considering taking this supplement should discuss it with a medical professional to avoid potential issues such as drug interactions.

Quercetin, a flavonol, has been extensively studied, and has been shown to protect against DNA mutations, colon cancer and heart disease (Hollman, and Katan, 1999). Quercetin relaxes the blood vessel wall (Rendig, et al., 2001) and increases the production of enzymes that dissolve blood clots (Abou-Agag, et al., 2001).  Research has recently shown that ellagic acid combined with quercetin act together to affect the growth rate and the path by which cancer cells die. The combination of these two compounds changes the activity of regulatory proteins and enzymes called MAP kinases that regulate cell division and viability (Mertens-Talcott, and Percival, 2005).

Most of the research on quercetin and cancer has been done in cell culture or animal studies. These types of studies can suggest possible helpful effects, but they do not provide proof that such effects can be achieved in humans. It is still unclear how well quercetin is absorbed by the human body when taken by mouth. Controlled clinical trials are needed to show whether quercetin has helpful properties in humans.

Studies done in cell cultures have shown that quercetin has activity against some types of cancer cells. This may be due to its antioxidant or anti-inflammatory properties, or it may be due to other mechanisms. Recent studies suggest that quercetin can slow the growth of cancer cells and can help foster apoptosis, a form of natural cell death that doesn’t happen in most cancer cells. Some studies in animals have shown that quercetin may help protect against certain types of cancer, particularly colon cancer.

Studies in humans have mainly been population-based and have focused on the role of flavonoids in the diet as a group as opposed to quercetin in particular. These types of studies are not as conclusive as clinical trials. They cannot prove cause and effect but often suggest links that can then be tested in clinical trials. While some of these population-based studies have found that people with diets high in flavonoids may have lower risk of breast, lung, pancreatic, and other types of cancer, it is not clear what role quercetin played in their findings. One clinical study of people with a strong inherited tendency to develop colorectal cancer found that the combination of quercetin and curcumin supplements decreased the number and size of precancerous rectal tumors. No other clinical trials testing quercetin’s ability to prevent or treat cancer have been reported in the medical literature. Clinical trials are needed to further clarify quercetin’s possible benefits.

In addition to cancer prevention and treatment, preliminary studies have also suggested potential value for quercetin in prostatitis (inflamed prostate) and heart disease. Further studies are needed before any recommendations can be made.

Until conclusive clinical research findings emerge, it is reasonable to include foods that contain quercetin as part of a balanced diet with an emphasis on fruits, vegetables, legumes, and whole grains. The interaction between certain phytochemicals and the other compounds in foods is not well understood, but it is unlikely that any single compound offers the best protection against cancer. A balanced diet that includes 5 or more servings a day of fruits and vegetables, along with foods from a variety of other plant sources such as nuts, seeds, whole grain cereals, and beans, is likely to be more effective in reducing cancer risk than eating one particular phytochemical in large amounts.

Kaempferol is a flavonoid found in many edible plants and in plants or botanical products commonly used in traditional medicine (e.g. Ginkgo biloba, Tilia spp, Equisetum spp, Moringa oleifera, Sophora japonica and propolis). Some epidemiological studies have found a positive association between the consumption of foods containing kaempferol and a reduced risk of developing several disorders such as cancer and cardiovascular diseases. Numerous preclinical studies have shown that kaempferol and some glycosides of kaempferol have a wide range of pharmacological activities, including antioxidant, anti-inflammatory, antimicrobial, anticancer, cardioprotective, neuroprotective, antidiabetic, anti-osteoporotic, estrogenic/antiestrogenic, anxiolytic, analgesic and antiallergic activities.

As noted, much work has been done on health benefits of individual chemicals found in muscadine grapes, but thus far there has not been a study showing the concentrations of these compounds in a variety of muscadines.  Most studies have only looked at a few muscadines varieties such as, ‘Noble’, ‘Ison’, and ‘Carlos’ because these are the most widely grown for production and wine aspects.  A more comprehensive look at muscadine cultivars would reveal, some “hidden treasures” in lesser known cultivars.  A previous study (Marshall et al., 2012) examined the important phytochemical concentrations in the muscadine fruit tissue of 21 cultivars at harvest.  From these results a subset of 11 cultivars were selected for analysis after 14 days of storage.  The criteria for selection were based on the levels of ellagic acid and flavonols, myricetin, quercetin and kaempferol found at harvest.