Cruciferous vegetables, such as broccoli, cabbage and kale, are rich sources of sulfur-containing compounds called glucosinolates. Isothiocyanates are biologically active hydrolysis (breakdown) products of glucosinolates. Cruciferous vegetables contain a variety of different glucosinolates, each of which forms a different isothiocyanate when hydrolyzed (see Figure) (1). For example, broccoli is a good source of glucoraphanin, the glucosinolate precursor of sulforaphane (SFN), and sinigrin, the glucosinolate precursor of allyl isothiocyanate (AITC) (2). Watercress is a rich source of gluconasturtiin, the precursor of phenethyl isothiocyanate (PEITC), while garden cress is rich in glucotropaeolin, the precursor of benzyl isothiocyanate (BITC). At present, scientists are interested in the cancer-preventive activities of vegetables that are rich in glucosinolates (see Cruciferous Vegetables), as well as individual isothiocyanates (3).

Summary

* Isothiocyanates are derived from the hydrolysis (breakdown) of glucosinolates—sulfur-containing compounds found in cruciferous vegetables.

* Cruciferous vegetables contain a variety of different glucosinolates, each of which forms a different isothiocyanate when hydrolyzed.

* Isothiocyanates, such as sulforaphane, may help prevent cancer by promoting the elimination of potential carcinogens from the body and enhancing the transcription of tumor suppressor proteins.

* Epidemiological studies provide some evidence that human exposure to isothiocyanates through cruciferous vegetable consumption may decrease cancer risk, but the protective effects may be influenced by individual genetic variation in the metabolism and elimination of isothiocyanates from the body.

* Glucosinolates are present in relatively high concentrations in cruciferous vegetables, but cooking, particularly boiling and microwaving at high power, may decrease the bioavailability of isothiocyanates.

Metabolism and Bioavailability

Myrosinase, a class of enzymes that catalyzes the hydrolysis (breakdown) of glucosinolates, is physically separated from glucosinolates when plant cells are intact (4). When cruciferous vegetables are chopped or chewed, myrosinase can interact with glucosinolates and release isothiocyanates from their precursors (see Figure). Thorough chewing of raw cruciferous vegetables increases glucosinolate contact with plant myrosinase and increases the amount of isothiocyanates absorbed (5). Even when plant myrosinase is completely inactivated by heat, the myrosinase activity of human intestinal bacteria allows for some formation and absorption of isothiocyanates (6). However, the absorption and excretion of isothiocyanates is substantially lower from cooked than from raw cruciferous vegetables (5, 7, 8) (see Food Sources below). During metabolism, isothiocyanates are conjugated (bound) to glutathione, an activity that is promoted by a family of enzymes called glutathione-S-transferases (GSTs), and further metabolized to mercapturic acids. These isothiocyanate metabolites can be measured in the urine, and are highly correlated with dietary intake of cruciferous vegetables (9). There is also some evidence that isothiocyanate metabolites contribute to the biological activity of isothiocyanates (3, 10).

Biological Activities

Effects on Biotransformation Enzymes Involved in Carcinogen Metabolism

Biotransformation enzymes play important roles in the metabolism and elimination of a variety of chemicals, including drugs, toxins and carcinogens. In general, phase I biotransformation enzymes catalyze reactions that increase the reactivity of hydrophobic (fat-soluble) compounds, preparing them for reactions catalyzed by phase II biotransformation enzymes. Reactions catalyzed by phase II enzymes generally increase water solubility and promote the elimination of the compound from the body (11).

Inhibition of Phase I Biotransformation Enzymes

Some procarcinogens (carcinogen precursors) require biotransformation by phase I enzymes, such as those of the cytochrome P450 (CYP) family, in order to become active carcinogens that are capable of binding DNA and inducing mutations. Inhibition of specific CYP enzymes involved in carcinogen activation inhibits the development of cancer in animal models (3). Isothiocyanates, including PEITC and BITC, have been found to inhibit carcinogen activation by CYP enzymes in animal studies (12, 13). A small clinical trial in smokers found evidence that consumption of 170 g/d (6 oz/d) of watercress, which is rich in the glucosinolate precursor of PEITC, decreased the activation of a procarcinogen found in tobacco (14).

Induction of Phase II Biotransformation Enzymes

Many isothiocyanates, particularly SFN, are potent inducers of phase II enzymes in cultured human cells (2). Phase II enzymes, including GSTs, UDP-glucuronosyl transferases (UGTs), quinone reductase and glutamate cysteine ligase, play important roles in protecting cells from DNA damage by carcinogens and reactive oxygen species (15). The genes for these and other phase II enzymes contain a specific sequence of DNA called an antioxidant response element (ARE). Isothiocyanates have been shown to increase phase II enzyme activity by increasing the transcription of genes that contain an ARE (16). Limited data from clinical trials suggests that glucosinolate-rich foods can increase phase II enzyme activity in humans. When smokers consumed 170 g/d (6 oz/d) of watercress, urinary excretion of glucuronidated nicotine metabolites increased significantly, suggesting UGT activity increased (17). Brussels sprouts are rich in a number of glucosinolates, including precursors of AITC and SFN. Consumption of 300 g/d (11 oz/d) of Brussels sprouts for a week significantly increased plasma and intestinal GST levels in nonsmoking men (18, 19).

Preservation of Normal Cell Cycle Regulation

After a cell divides, it passes through a sequence of stages known as the cell cycle before dividing again. Following DNA damage, the cell cycle can be transiently arrested to allow for DNA repair or activation of pathways leading to cell death (apoptosis) if the damage cannot be repaired (20). Defective cell cycle regulation may result in the propagation of mutations that contribute to the development of cancer. A number of isothiocyanates, including AITC, BITC, PEITC and SFN, have been found to induce cell cycle arrest in cultured cells (2).

Inhibition of Proliferation and Induction of Apoptosis

Unlike normal cells, cancer cells proliferate rapidly and lose the ability to respond to cell death signals by undergoing apoptosis. Isothiocyanates has been found to inhibit proliferation and induce apoptosis in a number of cancer cell lines (3).

Inhibition of Histone Deacetylation

In the nucleus of a cell, DNA is coiled around basic proteins called histones. In general, acetylation of histones by histone acetyl transferases makes DNA more accessible to transcription factors, which bind DNA and activate gene transcription. Deacetylation of histones by histone deacetylases restricts the access of transcription factors to DNA. Acetylation and deacetylation of nuclear histones is an important cellular mechanism for regulating gene transcription (21). However, the balance between histone acetyl transferase and histone deacetylase activities that exists in normal cells may be disrupted in cancer cells. Compounds that inhibit histone deacetylases have the potential to suppress the development of cancer by inducing the transcription of tumor suppressor proteins that promote differentiation and apoptosis in transformed (precancerous) cells (22). AITC and SFN metabolites have been found to inhibit histone deacetylase activity in cultured cancer cells (10, 23).

Anti-inflammatory Activity

Inflammation promotes cellular proliferation and inhibits apoptosis, increasing the risk of developing cancer (24). SFN and PEITC have been found to decrease the secretion of inflammatory signaling molecules by white blood cells and to decrease DNA binding of NF-kappaB, a pro-inflammatory transcription factor (25, 26).

Antibacterial Activity: Helicobacter pylori

Bacterial infection with H. pylori is associated with a marked increase in the risk of gastric cancer (27). Purified SFN inhibited the growth and killed multiple strains of H. pylori in the test tube and in tissue culture, including antibiotic resistant strains (28). In an animal model of H. pylori infection, SFN administration for 5 days eradicated H. pylori from 8 out of 11 xenografts of human gastric tissue implanted in immune-compromised mice (29). However, in a small clinical trial, consumption of up to 56 g/d (2 oz/d) of glucoraphanin-rich broccoli sprouts for a week was associated with H. pylori eradication in only 3 out of 9 gastritis patients (30). Further research is needed to determine whether SFN or foods rich in its precursor glucobrassicin will be helpful in the treatment of H. pylori infection in humans.

Disease Prevention

Cancer

Naturally occurring isothiocyanates and their metabolites have been found to inhibit the development of chemically-induced cancers of the lung, liver, esophagus, stomach, small intestine, colon and mammary gland (breast) in a variety of animal models (3, 12). Although epidemiological studies provide some evidence that higher intakes of cruciferous vegetables are associated with decreased cancer risk in humans (31), it is difficult to determine whether such protective effects are related to isothiocyanates or other factors associated with cruciferous vegetable consumption (see Cruciferous Vegetables). Investigators have attempted to calculate human isothiocyanate exposure based on assessments of cruciferous vegetable intake and measurements of the maximal amounts of isothiocyanates that can be liberated from various cruciferous vegetables in the laboratory (32). Case-control studies using this technique found that dietary isothiocyanate intakes were significantly lower in Chinese women (33) and US men (34) diagnosed with lung cancer than in cancer-free control groups. Assessing dietary intake of cruciferous vegetables may not accurately measure an individual’s exposure to isothiocyanates, since other factors may alter the amount of isothiocyanates formed and absorbed (see Metabolism and Bioavailability above). Measuring urinary excretion of isothiocyanates and their metabolites may provide a better assessment of isothiocyanate exposure (9, 35), but few studies have examined relationships between urinary isothiocyanate excretion and cancer risk. In a prospective study, Chinese men with detectable levels of urinary isothiocyanates at baseline were at significantly lower risk of developing lung cancer over the next 10 years than men with undetectable levels (36). A case-control study found that urinary isothiocyanate excretion was significantly lower in Chinese women diagnosed with breast cancer than in a cancer-free control group (37). In contrast, cruciferous vegetable intake estimated from a food frequency questionnaire was not associated with breast cancer risk in the same study.

Genetic Variation in Isothiocyanate Metabolism and Cancer Risk

GSTs are a family of phase II biotransformation enzymes that promote the metabolism and elimination of isothiocyanates and other compounds from the body. Genetic variations (polymorphisms) that affect the activity of GST enzymes have been identified in humans. Null variants of the GSTM1 gene and GSTT1 gene contain large deletions, and individuals who inherit two copies of the GSTM1-null or GSTT1-null gene cannot produce the corresponding GST enzyme (38). Lower GST activity in such individuals could result in slower elimination and longer exposure to isothiocyanates after cruciferous vegetable consumption (9). In support of this idea, several epidemiological studies found that inverse associations between isothiocyanate intake from cruciferous vegetables and the risk of lung cancer (33, 34, 36, 39) or colon cancer (40-42) were more pronounced in GSTM1-null or GSTT1-null individuals. These findings suggest a protective role for isothiocyanates that may be enhanced in individuals who eliminate them from the body more slowly.

Sources

Food Sources

Cruciferous Vegetables

Cruciferous vegetables, such as bok choi, broccoli, Brussels sprouts, cabbage, cauliflower, horseradish, kale, kohlrabi, mustard, radish, rutabaga, turnip and watercress, are rich sources of glucosinolate precursors to isothiocyanates (43). Unlike some other phytochemicals, glucosinolates are present in relatively high concentrations in commonly consumed portions of cruciferous vegetables. For example ½ cup of raw broccoli might provide more than 25 mg of total glucosinolates. Total glucosinolate contents of selected cruciferous vegetables are presented in Table 1 (44). Some cruciferous vegetables are better sources of specific glucosinolates (and isothiocyanates) than others. Vegetables that are relatively good sources of some isothiocyanates that are currently under study for their cancer-preventive properties are listed in Table 2. Amounts of isothiocyanates formed from glucosinolates in foods are variable and depend partly on the processing and preparation of those foods (see Effects of Cooking below). Consumption of 5 or more weekly servings of cruciferous vegetables has been associated with significant reductions in cancer risk in some prospective cohort studies (45-47).

Broccoli Sprouts

The amount of glucoraphanin, the precursor of SFN, in broccoli seeds remains more or less constant as those seeds germinate and grow into mature plants. Thus, 3-day old broccoli sprouts are concentrated sources of glucoraphanin, which contain 10-100 times more glucoraphanin by weight than mature broccoli plants (48). Broccoli sprouts that are certified to contain at least 73 mg of glucoraphanin (also called sulforaphane glucosinolate) per 1-oz serving are available in some health food and grocery stores.

Effects of Cooking

Glucosinolates are water-soluble compounds that may be leached into cooking water. Boiling cruciferous vegetables from 9-15 minutes resulted in 18-59% decreases in the total glucosinolate content of cruciferous vegetables (44). Cooking methods that use less water, such as steaming or microwaving may reduce glucosinolate losses. However, some cooking practices, including boiling (5), steaming (7) and microwaving at high power (850-900 watts) (8, 49) may inactivate myrosinase, the enzyme that catalyzes glucosinolate hydrolysis. Even in the absence of plant myrosinase activity, the myrosinase activity of human intestinal bacteria results in some glucosinolate hydrolysis (6). However, several studies in humans have found that inactivation of myrosinase in cruciferous vegetables substantially decreases the bioavailability of isothiocyanates (5, 7, 8).

Supplements

Dietary supplements containing extracts of broccoli sprouts, broccoli and other cruciferous vegetables are available without a prescription. Some products are standardized to contain a minimum amount of glucosinolates and/or sulforaphane. However, the bioavailability of isothiocyanates derived from these supplements is not known.

Safety

Adverse Effects

No serious adverse effects of isothiocyanates in humans have been reported. The majority of animal studies have found that isothiocyanates inhibited the development of cancer when given prior to the chemical carcinogen (pre-initiation) However, very high intakes of PEITC or BITC (25-250 times higher than average human dietary isothiocyanate intakes) have been found to promote bladder cancer in rats when given after a chemical carcinogen (post-initiation) (50). The relevance of these findings to human urinary bladder cancer is not clear, since at least one prospective cohort study found cruciferous vegetable consumption to be inversely associated with the risk of bladder cancer in men (47).

Pregnancy and Lactation

Although high dietary intakes of glucosinolates from cruciferous vegetables are not known to have adverse effects during pregnancy or lactation, there is no information on the safety of purified isothiocyanates or supplements containing high doses of glucosinolates and/or isothiocyanates during pregnancy or lactation in humans.

Drug Interactions

Isothiocyanates are not known to interact with any drugs or medications. However, the potential for isothiocyanates to inhibit various isoforms of the CYP family of enzymes raises the potential for interactions with drugs that are CYP substrates (51).

References

Written by:
Jane Higdon, Ph.D.
Linus Pauling Institute
Oregon State University