1Introduction

Scope and Objectives

This evidence report was requested by the National Center for Complementary and Alternative Medicine, a component of the National Institutes of Health. It was contracted by the Agency for Healthcare Research and Quality, part of the U.S. Department of Health and Human Services. The following are the rationale for this report addressing garlic: (1) availability of multiple clinical studies with promising but conflicting results and (2) high consumer usage of garlic as a health supplement. This report is intended primarily for agencies interested in funding clinical garlic studies, clinicians, and researchers, and secondarily for consumers. This chapter highlights the rich history of garlic, the complexity of its chemistry, recent research, the variety of available commercial preparations, and challenges in conducting garlic research in humans. The evidence report is a systematic review that summarizes the studies in humans that address the following three areas:

• Effects of garlic on cardiovascular-related factors and disease. Results of randomized trials lasting at least 4 weeks comparing garlic preparations with placebo or other agents are presented. Effects on the following outcomes are addressed: clinical cardiovascular disease, lipids, atherosclerosis, blood pressure, glucose, insulin sensitivity, and antithrombotic activity.
• Associations of garlic in preventing cancer. Results of case-control and cohort studies that compare the occurrence of various precancerous conditions and cancer in persons consuming or avoiding garlic are described.
• Clinical adverse effects of garlic ingestion or contact. Various reported adverse effects, including dermatological, gastrointestinal, and hematological, are summarized.

Origins and Folklore of Garlic

Garlic, scientifically known as Allium sativum L., is closely related to other "smelly" bulbs such as onions, leeks, and chives. ¹ There is an old story originally told by the ancient Islamic religious leader Mahomet that garlic sprang from the left footprint, and onion from the right, of Satan as he walked out of the Garden of Eden. ² Another story, translated from the 5 ^th century AD Bower Manuscript , says that garlic grew from the drops of blood that fell to the earth when Janar-Dana decapitated Asura, the ruler of the Old-Indian gods. ³ Historians believe that garlic was native to Siberia and was spread to other geographic regions more than 5,000 years ago by Siberian nomadic tribes. ^{4 5}

Throughout recorded history, garlic has played rich and diverse commercial, culinary, literary, and mythical roles. Ancient Egyptian and Middle Eastern cultures used garlic as a currency. Around 2,500 BC, 15 pounds of garlic could purchase a healthy slave. ⁶ The culinary delights of garlic over manna were extolled by followers of Moses in the Old Testament (Numbers 11:5,6). Virgil, in his Second Idyll , described how Thestylis used the juices of wild thyme and garlic as a prophylactic against snakebites, while Homer wrote that garlic helped Ulysses escape from being changed by Circe into a pig like his companions. ^{3 4}

Balkans rubbed garlic on doorknobs and window frames to discourage vampires from haunting them, and they hung garlic on doors and windows so that nobody would take milk from their cows and the families would be safe from witches. ^{3 6} In other parts of Europe, fisherman and seamen wore cloves of garlic to protect themselves from evil spirits and diseases, and mothers hung garlic on children to protect them from the demons of sickness. ³ Other religions and cultures shunned garlic and believed it to be evil. Sects of Christianity, Hinduism, Islam, and Zen Buddhism have considered garlic "unclean." ^{2 7} Nepalese shamans of the Brahan and Cherti castes were forbidden to eat garlic because it could cause suffering in the afterworld. ² In ancient Greece, people who had eaten garlic were forbidden from entering the Temple of Cybele because its smell was considered offensive. ²

Chemistry of Garlic

While garlic contains the usual complement of carbohydrates, proteins, lipids, vitamins, minerals, and nucleic acids found in other plants, it also contains approximately 5 percent dry weight of sulfur-containing nonprotein amino acid secondary metabolites, which are responsible for both its characteristic flavor and biological activity (Figure 1). ^{8 9 10} As with all plants, the sulfur is derived from soil sulfate, which is transformed into the amino acid l-cysteine and then to g-glutamylcysteine. Methylation of sulfur affords γ-glutamyl- S -methylcysteine, which is converted to the corresponding sulfoxide by an oxidase enzyme and then stripped of its γ-glutamyl group by a γ-glutamyl transpeptidase enzyme. The final product of this sequence, S -methylcysteine sulfoxide (sometimes called "methiin"), is a colorless, odorless, high-melting solid that serves as a precursor to the primary flavor compounds ("flavorants"), which are thought to serve as plant-protective compounds (e.g., natural pesticides). Garlic also contains homologs of methiin and its γ-glutamyl derivatives (sulfoxide and sulfide forms), namely alliin ([+]- S -allyl- l-cysteine sulfoxide), isoalliin ( S -1-propenylcysteine sulfoxide), and propiin ( S -propylcysteine sulfoxide), together with the corresponding γ-glutamyl derivatives, mainly γ-glutamyl- S -allylcysteine and γ-glutamyl- S -trans-1-propenylcysteine. In garlic leaves, the glycoside of alliin, (-)- N -(1'-deoxy-1'-β-D-fructopyranosyl)- S -allyl-L-cysteine sulfoxide, also is found. In garlic, the alliin:methiin ratio is approximately 10:1, and the alliin:isoalliin ratio is approximately 20:1, while propiin is present only in trace amount. ³ The amount of alliin in whole garlic ranges from 2.8 to 7.7 milligrams per gram (mg/g) fresh weight. Although selenium is present at far lower concentrations than sulfur, relative to other common vegetables, garlic is comparatively rich in selenium. Selenium in garlic is derived from soil selenite and selenate, which is transformed to L-selenocysteine and then stored primarily as γ-glutamyl- Se -methylselenocysteine.

Crushing or cutting garlic results in a comingling of precursor compounds alliin and methiin, found in the cytosol, with robust, abundant enzymes known as alliinases, which are concentrated in stem cells. The result is cleavage of alliin and methiin to give 2-propenesulfenic acid and methanesulfenic acid, respectively. Sulfenic acids are highly reactive intermediates, rapidly condensing to give compounds known as thiosulfinates ( RS(O)SR'), which are the actual primary flavor compounds. Four sulfenic acids (RSOH, R = methyl, allyl, 1-propenyl, and propyl) are formed when garlic is cut. These acids combine in all possible permutations to give thiosulfinates, whose specific ratio is responsible for the unique flavor of garlic. The major thiosulfinate from garlic is allicin, CH ₂ =CHCH ₂ S(O)SCH ₂ CH=CH ₂ (RS(O)SR, where R = allyl), formed in less than 10 seconds when alliinase acts on alliin. Thiosulfinates are quite reactive and undergo a variety of further transformations, depending on conditions. Thus, in organic solvents or oils, allicin decomposes into 2-propenesulfenic acid and thioacrolein; the latter compound rapidly undergoes self-condensation giving isomeric dithiins. In a second, more complex process, allicin can form ajoene. This reaction occurs in organic solvents or food oils, but only at trace levels in water. Thiosulfinates react with heated water to give dialk(en)yl polysulfides such as diallyl polysulfides and allyl methyl polysulfides. These polysulfides have a characteristic odor, which is similar to allicin. The above transformations of allicin and other thiosulfinates can occur in the laboratory as well as when cooking with garlic. Frying garlic, which involves temperatures higher than 100 degrees Celsius (°C), denatures alliinases and converts alliin to cysteine and allyl alcohol. Allicin is moderately stable in aqueous solutions, with a half-life of about 30 days at room temperature and about 6 months at 4° C. ³ Alliinases also are deactivated at low pH. Their activity is maximal at pH between 5 and 8 and rapidly diminishes at higher or lower pH. Under conditions when allicin and cysteine or allicin and glutathione (the main intracellular thiol of mammalian cells) react, S -allylmercaptocysteine or S -allylmercaptoglutathione is formed, respectively. ^{3 11 12}

When fresh garlic is ingested, 2-propenethiol, an odorous compound even at very low concentrations, can be detected on the breath. This compound was identified in human garlic breath by collecting and analyzing breath samples immediately after ingestion of sliced fresh garlic. Within a few minutes, the 2-propenethiol disappears and is replaced by allyl methyl sulfide, the methylated metabolite of 2-propenethiol. Acetone also is detected in human breath after garlic ingestion and may parallel increased lipid metabolism. Allicin, ajoene, dithiins, and certain allyl sulfides have antithrombotic activity. Allicin, ajoene, and allyl sulfides also may have antilipidemic properties, while allyl sulfides have anticancer activity. Organoselenium compounds may follow a similar metabolic pathway to that of sulfur. Thus, γ-glutamyl- Se -methylselenocysteine is presumably hydrolyzed in the gut to Se -methylselenocysteine, which is enzymatically cleaved to methaneselenol, and then methylated to dimethyl selenide, another compound detected in human garlic breath. Dehydrated tablets manufactured from fresh garlic also contain alliin, γ-glutamylcysteine derivatives, alliinases, and all of the compounds found naturally in garlic. When garlic cloves are consumed, N -acetyl- S -allylcysteine is excreted in the urine.

Historical Uses of Garlic

Arthritis, asthma, toothaches, freckles, baldness, athlete's foot, plague, cancer, and cardiovascular disease are just a few of the many maladies that people throughout history have treated with garlic. The teachings of the father of Ayurvedic medicine, Charak (around 3,000 BC), described early uses of garlic to maintain the fluidity of blood and strengthen the heart. ¹³ Similar uses were reiterated in the Codex Ebers , an Egyptian medical manuscript dating to approximately 3,500 years ago. ^{7 13 14} In the Talmud , an important collection of ancient rabbinic writings, garlic is recommended for treating wounds. ^{3 14} Pliny the Elder, the author of what is believed to be the first encyclopedia, Historia Naturalis, wrote that garlic could cure 62 ailments, including consumption, hemorrhoids, dog and snake bites, and tumors. ⁷ Hippocrates recommended garlic as a laxative and diuretic. ^{3 7} In the mid-1500s, Adam Lonitzer, a German physician author of an herbal reference book, recommended garlic juice externally for killing lice and nits and internally to rid the body of worms and poison. ³ By the 1800s, Louis Pasteur had reported that garlic kills bacteria, ¹⁴ and in World War II, garlic juice with water was used to disinfect wounds. ^{15 16} During the last half of the 20 ^th century, investigations relating to the medicinal qualities of garlic exploded. The next two sections will briefly describe some of the research relating to cardiovascular factors and cancer.

Garlic Research Addressing Cardiovascular-Related Factors and Disease

Animal studies in rats and rabbits show multiple effects of garlic on cardiovascular-related factors, including antithrombotic effects, regression of atheromata, and decreases in total cholesterol, glucose, and blood pressure. ^{17 18 19 20 21 22} Although some observational studies and small trials in humans corroborate animal studies, ^{20 23 24 25 26 27 28 29} data are sometimes confusing and conflicting. For example, some studies in humans show that short-term administration of garlic for less than 4 months has either no lipid-lowering or transient detrimental effects, ^{30 31 32 33} while others suggest that garlic has short-term beneficial effects. ^{34 35 36 37 38 39 40} A possible reason for the conflicting data is the variable release of allicin or allicin-derived compounds from the dosage forms used in the studies. ⁴¹

Several reviews that were published in the early 1990s summarized randomized controlled trial (RCT) data about garlic and cardiovascular factors. ^{13 42 43 44 45 46 47} Most focused on the effects of garlic on lipids, although one summarized the effects on blood pressure. ⁴⁶ The reviews included different studies, gave varying attention to specific garlic preparations and doses, and sometimes reached different conclusions. Moreover, multiple new trials are available, and some focus on endpoints, such as antithrombotic activity and progression of atherosclerotic plaques, that were not addressed in prior reviews.

Garlic Research Addressing Cancer

Most, but not all, in vivo and in vitro studies have shown that garlic constituents inhibit tumor cell metabolism, inhibit tumor initiation and promotion, and/or modulate immune responses. ^{48 49 50 51} For example, in vitro studies have suggested that constituents such as allicin and ajoene have antimitotic and antimutagenic properties, while other studies have suggested that constituents such as alliin do not inhibit mutagenesis. ⁴⁹ Some in vivo studies have shown an inhibition of tumor growth in rats and mice with fresh garlic but not with garlic in which alliinase had been inactivated. ^{48 50} Extrapolation of effective doses from in vivo experiments to potentially equivalent doses in humans has been estimated at 25 to 450 mg of garlic cloves per kilogram (kg) of body weight or 2 to 32 g per 70-kg person. ^{52 53 54} Recent epidemiological studies in humans have examined whether garlic consumption is related to incidence of breast, colorectal, gastric, lung, and prostatic cancer. ^{55 56 57 58 59 60 61} These studies have reported intriguing results; a detailed review of their design, method of ascertaining cancer, and measurement of garlic intake is warranted.

Garlic Preparations

More than 300 strains of garlic are grown throughout the world, and the use of garlic as an herbal supplement is a big business. In 1993, U.S. garlic supplement sales totaled $31.3 million, second only to Echinacea in total herbal product sales at health food stores. ⁶² A year later, garlic supplement sales had increased to $39.4 million. In a 1997 market survey of herbal sales, garlic was one of the top three best-selling supplements in this $3.2 billion industry. ¹⁶ As Table 1 depicts, garlic historically has been prepared and administered in numerous ways. The tradition continues today where garlic is ingested raw, pickled, cooked, or as a commercially prepared supplement.

Summary of Challenges in Conducting and Interpreting Garlic Research in Humans

There are many issues in evaluating clinical effects of garlic in humans (Table 2). First, garlic is a food product that is composed of numerous macronutrients, vitamins, trace elements, and organosulfur compounds. Multiple possible pharmacologically active ingredients are present. The different constituents within garlic may have no or varying antilipidemic, antihypertensive, antiglycemic, antithrombotic, and antitumor potentials.

Second, true active ingredients and mechanisms of action of garlic constituents are varied and sometimes unclear. For example, whether allicin is truly an active ingredient has been questioned because it has a very transient half-life and limited or no bioavailability that may be due to a significant hepatic first-pass metabolism, although metabolites of allicin do have substantial activity. Proposed mechanisms of action of various garlic constituents include, but are not limited to, the following: decrease in dietary fat absorption, decrease in hepatic cholesterol synthesis, alteration in lipoprotein composition, alteration in total body lipid composition, vasorelaxation, antioxidation, and increased tissue insulin sensitivity.

Third, garlic constituents have varying bioavailability. Whether the in vitro activity of garlic constituents correlates with true physiological effects in humans is difficult to assess. Consuming identical preparations of garlic may result in varying bioavailability depending upon several factors. For example, particular foods that are consumed with garlic may impair absorption, alter gastric pH, or react directly with key constituents of the garlic product. Some substances, including medications, may alter hepatic first-pass metabolism. Particular underlying conditions in individuals such as dyslipidemia, atherosclerosis, or genetic predispositions to cancer also may affect biological efficacy.

Fourth, there is substantial variability between garlic compounds, based on where and how the whole garlic is grown, as well as how the end preparation is prepared and stored. Fresh garlics from around the world vary in amounts of compounds, based on climate and soil conditions. Some purported active ingredients, such as SACs, are not present above trace levels in fresh garlic but are instead produced by aging chopped garlic in dilute alcohol. Other organosulfur compounds are produced through maceration in the presence of edible oils. Another source of variability is the stability of garlic compounds in the different stages of storage between harvest and consumption.

Fifth, quantitative release of allicin (a possible main active compound) from dehydrated preparations needs verification under simulated gastrointestinal conditions or by an in vivo method. This may be critical because allicin release depends on alliinase activity, which can be greatly decreased by improper powder or tablet manufacturing, gastric acid, and neutral pH that is typical of the intestinal tract. Tablets that dissolve quickly after entering the intestinal tract have the greatest amount of alliinase activity. Notably, dissolution properties of preparations are not routinely reported in the trials, but some Kwai ^® preparations that were used in early trials have been shown to release three times as much allicin as the Kwai ^® tablets that were used in the more recent trials. ⁴¹

Sixth, studies of garlic intake are somewhat unique regarding the ability to successfully conduct "blinded" studies. One of the characteristics of the "stinking rose" that separates garlic from almost any other dietary component is its strong and unique taste, as well as the related issues of associated body and breath odors. Whether garlic study participants can detect the garlic or placebo group to which they have been assigned raises at least two concerns. The first is the classical concern that provides the rationale for "blinding" study participants to which group they are assigned. When there is a perceived benefit of one assignment over another, research participants might alter their behavior (e.g., diet or exercise) if they think that they have been assigned to the garlic group. If this is the case, it becomes difficult to determine to what extent study results are attributable to the garlic versus the associated behavior changes. A second concern is perhaps less intuitive and more hypothetical. It is possible that the putative health benefits of ingesting garlic are attributable to the presence, concentration, and form of its many sulfur-containing compounds. The sulfur compounds generate the unique taste and odor of garlic. Garlic studies that successfully blind participants to group assignments may be using garlic products that have undetectable taste and body and breath odor as well as low levels of active sulfur compounds present. If benefits were tied to sulfur compounds, such studies would underestimate them.

Seventh, even sustained beneficial effects on intermediate physiological factors such as lipids and blood pressure do not necessarily portend clinically important benefits. At present, the research evidence from human studies in the cardiovascular area focuses almost exclusively on intermediate physiological factors rather than clinical outcomes. The very multiplicity of potential physiological effects of garlic underscores the need for studies with broadly defined clinically important outcomes.