INTRODUCTION

The genus Medicago belongs to the legume family (Fabaceae, also known as Leguminosae), which includes an estimated 751 genera and about 19,400 species. This genus includes 87 species¹ distributed in parts of Africa, Central and Southwest Asia, and Europe.² The subject of this article, Medicago sativa, is an herbaceous perennial that reaches 60–70 cm (23.6–27.6 in) in height, with inflorescences consisting of an oblong raceme (flower cluster) of purple flowers.³

Medicago sativa, native throughout much of Asia, Europe, and Northwest Africa, is a wild relative of the widely cultivated subspecies M. sativa subsp. sativa, which is known in commerce as alfalfa (standard common name) or lucerne (other common name).^4-6 Although M. sativa subsp. sativa also occurs in the wild (albeit rarely, except in the Caucasus and Turkey and on the Iberian Peninsula), this subspecies is also naturalized elsewhere due to escapes from cultivation.⁷ Cultivated M. sativa subsp. sativa is reportedly the most important forage legume globally.⁸ With about 32 million hectares (79 million acres) of alfalfa cultivation worldwide, it is also among the leading plants in terms of protein produced per hectare.⁹ The United States is the largest alfalfa producer worldwide, with commercial cultivation of 7 million hectares (17.3 million acres) in 42 states.¹⁰ Globally, about 1.5 billion kg (3.3 billion lbs) of alfalfa meal and pellets were exported in 2021. The top five exporters in terms of volume in 2021 were (in order of predominance) Spain, Italy, the United States, South Africa, and France.¹¹

Alfalfa is a well-established forage crop and feed source for ruminant livestock due to its nutritional content. For human health, preparations of alfalfa are used in traditional herbal medicines^12-15 as well as in anthroposophic medicines (a system of medicine developed by Austrian philosopher Rudolf Steiner [1861–1925]),¹⁶ homoeopathic medicines (a system of medicine developed by German physician Samuel Hahnemann [1755–1843]),³ natural cosmetics,¹⁷ and in dietary and food supplement products.⁹ Alfalfa, in the form of protein-xanthophyll concentrate, green fodder, hay, and haylage (partially dried forage, cut and stored in air-tight containers), is an excellent source of protein and other nutrients for horses, poultry (turkeys, laying hens, broilers), pigs, and other animals.

HISTORY AND CULTURAL SIGNIFICANCE

Western Asia¹⁸ and Iran in Southern Asia are believed to be the centers of origin of M. sativa.¹⁹ Medicago sativa is one of the oldest crops globally and also the oldest known forage crop, believed to have been first cultivated about 9,000 years ago, although this is difficult to pinpoint.^20,21 Archaeobotanical evidence suggests that M. sativa was first domesticated in the Caucasus region (between the Black Sea and Caspian Sea), Turkey, and Iran,¹⁸ where the earliest known written evidence of the use of alfalfa as forage also was found on a stone tablet dated to about 1300 BCE.¹⁸ Alfalfa cultivation spread to North Africa and Europe at different times, and alfalfa was carried through invasions by Medes (an Iranian people who inhabited ancient Media of northwestern Iran), Romans, and Moors.²⁰

The genus name Medicago stems from the Latin term Mēdica and Greek Μηδικὴ πόα (Mēdikē poa, meaning “median grass”), referring to the region of Media. It was associated with Persian horse breeding, particularly the war horses of Darius the Great (ca. 550–486 BCE), king of the Achaemenid Empire.²² The species name sativa means “cultivated.” Common names used in the geographic origin(s) of the species include (Romanized) aspust in Balochi, a language spoken in the Balochistan region of Afghanistan, Iran, and Pakistan; jatt or qatt in Arabic, spoken in Iraq and Syria; and winjah or yunjah in Kurdish, spoken in parts of Armenia, Azerbaijan, Iran, Iraq, Syria, and Turkey.²³ The English common name “alfalfa” is borrowed from the Spanish alfalfa (originally alfalfez), which was derived from Arabic al-fasfasah.²⁴

In 1700, French botanist Joseph Pitton de Tournefort (1656–1708) described the genus Medicago in his publication Institutiones Rei Herbariae.²⁵ In his 1737 work Genera Plantarum, Swedish botanist Carl Linnaeus (1707–1778) also listed the genus Medicago and referenced Tournefort.²⁶ Later, in his 1753 work Species Plantarum, Linnaeus described several Medicago species, including M. sativa, with the species locality stated as “Habitat in Hispaniae, Galliae apricis” (Spain and France).²⁷

In eastern Anatolia folk medicine, the leaf or herb of M. sativa subsp. sativa, known locally as karayonca, is pounded and applied topically as an astringent and hemostatic wound dressing.²⁸ In Iranian ethnomedicine, preparations of the leaf, seed, and stem, known locally as younjeh, also are applied topically for wound healing.²⁹ In Iran and Iraq, the seeds are prepared into a cooling poultice that is applied over boils.²³ The fresh leaves, known locally in northern and central Oman as qat or jat, are applied to the nose to stop nosebleeds.³⁰ The aerial parts, known locally as jonxha, are used in mountain villages of Peshkopia (eastern Albania) as a galactagogue to stimulate milk production for livestock.³¹ In the Rif Mountains of northern Morocco, the leaf, known locally as fessa, is prepared by decoction and taken orally for treating respiratory system diseases.³²

In traditional European medicine, alfalfa herb (aerial parts harvested during the flowering period) is prepared as a tea infusion (aqueous or in milk) for putting on weight.¹⁵ The dried herb, prepared as a tea infusion or as a fluidextract (1:1 in 25% alcohol), according to the British Herbal Pharmacopoeia, also is used for treating avitaminosis (vitamin A, C, E or K deficiency), hypoprothrombinemic purpura, and debility of convalescence. For treating purpura, alfalfa herb traditionally is combined with buckwheat (Fagopyrum esculentum, Polygonaceae) herb and rue (Ruta graveolens, Rutaceae) leaf.¹²

CURRENT AUTHORIZED USES IN COSMETICS, FOODS, AND MEDICINES

In the United States, the Food and Drug Administration (FDA) classifies “alfalfa herb and seed, Medicago sativa L.” as Generally Recognized as Safe (GRAS) for use as a spice, natural seasoning, or flavoring.³³ Natural extracts of alfalfa also are classified as GRAS for use in conventional food products.³⁴ Alfalfa herb and extracts made from it also are used as components of dietary supplement products.

In Canada, besides food use, alfalfa herb top and alfalfa leaf are regulated as medicinal ingredients of licensed natural health products (NHPs), which require pre-marketing authorization from the Natural and Non-prescription Health Products Directorate (NNHPD). Licensed NHPs that contain an alfalfa herb top preparation (dry extract, fluidextract, tea infusion, tincture), at the prescribed dosage, may be labeled with claim statements to the effect of “traditionally used in Herbal Medicine as a nutritive tonic”¹⁴ and “provides antioxidants that help protect against cell damage caused by free radicals.”³⁵ Licensed NHPs that contain the active ingredient alfalfa protein concentrate (prepared from M. sativa herb top) may be labeled with claim statements including “source of protein for the maintenance of good health,” “source of protein which helps build and repair body tissues,” “source of amino acids involved in muscle protein synthesis,” and “assists in the building of lean muscle tissue/mass when combined with regular weight/resistance training and a healthy balanced diet.”³⁶

In the European Union (EU) and the United Kingdom, alfalfa may be used as an active ingredient of registered traditional herbal medicinal products.¹³ Alfalfa also has been on the European market as a food or food ingredient and was consumed as such to a significant degree before May 15, 1997, which means that market access of traditional alfalfa food products is generally not subject to the Novel Food Regulation (EU) 2015/2283.³⁷ However, a particular leaf extract was granted marketing authorization in 2009 as a novel food ingredient to be used in food supplement products. It must be labeled as “lucerne (Medicago sativa) protein” or “alfalfa (Medicago sativa) protein,” and its specifications, method of preparation, and composition are defined in Regulation (EC) No. 258/97 of the European Parliament and of the Council.³⁸ Furthermore, for use in cosmetic products, “Medicago Sativa (whole plant) Extract” is permitted for use for tonic (producing a feeling of well-being on skin and hair) function, “Medicago Sativa Flower/Leaf/Stem Juice” for antioxidant function, and both “Medicago Sativa Leaf Extract” and “Medicago Sativa Leaf Powder” for skin-conditioning function.¹⁷

MODERN RESEARCH

Constituents and Pharmacological Effects

Alfalfa is known to be a high-quality source of protein, vitamins, and minerals for livestock fodder. However, alfalfa is not a commonly used herbal ingredient for human health; therefore, its actions are not clearly understood. Phytochemical constituents found in the alfalfa plant include alkaloids (e.g., asparagine, trigonelline, stachydrine, L-homostachydrine),^39-41 amino acids (both essential and non-essential), coumarins, coumestrol, flavonoids (e.g., flavones, isoflavonoids, flavonoid glycosides, phenolic compounds),^40,42-46phytosterols (e.g., b-sitosterol, stigmasterol),³⁹ saponins (e.g., triterpenoid saponins, sapogenins, soyasaponegols, hederagenin),^47-51non-protein amino acid L-canavanine (see “Toxicity” section), chlorophyll, and various vitamins (b-carotene, B, C, D, E and K) and minerals.^39,52

Alfalfa’s primary pharmacological effects demonstrated from animal, in vitro, and in vivo studies are antihyperglycemic, anti-inflammatory, antioxidant, antimicrobial, and hypocholesterolemic properties.^39,48,53 Antihyperglycemic activity has been reported when alfalfa was added to the diet (62.5 g/kg) and drinking water (2.5 g/L) in streptozotocin-induced diabetic mice.⁵⁴ Alfalfa water extract (1 g/kg) also showed antihyperglycemic activity in type 2 diabetic rats by reducing post-prandial glycemia and insulin-releasing activity in the diabetic treatment group.⁵⁵ A four-week study on diabetic rats found an alfalfa leaf extract (ethanol) showed comparable results to metformin (a conventional pharmaceutical antihyperglycemic medication). Study groups included normal controls, a non-treated diabetic group, and a treated diabetic group, which received an alfalfa leaf extract (500 mg/kg of body weight) or metformin (500 mg/kg of body weight) for 30 days orally. Alfalfa leaf extract lowered hyperglycemia by 42% compared to metformin at 37% after four weeks. Alfalfa leaf extract and metformin also correlated with decreased cholesterol and triglycerides in the diabetic rats and improved oxidative stress.⁵⁶

Anti-inflammatory properties have been reported in alfalfa aerial parts (in a chloroform extract) by inhibiting inflammatory pathways in a lipopolysaccharide-stimulated mouse macrophage cell model, which represent proinflammatory cytokine and macrophage pathways.⁵⁷ An alfalfa extract (methanol) was evaluated to understand its effects on liver health in a nicotine-induced liver damage rat model. The findings reported a dose-dependent anti-inflammatory effect of alfalfa extract through a decrease in proinflammatory cytokine markers associated with inflammation.⁵⁸

Ethanolic extracts of assorted alfalfa plant parts (leaves, stems before and after flowering, and flowers) were evaluated to determine total phenolic and flavonoid contents. The stem extract after flowering contained the highest total phenolic content at 75.89 mg of gallic acid equivalents (GAE)/g of extract and total flavonoid content 34.6 mg of quercetin equivalents (QE)/g of dry extract.⁵⁹ A different study determined that alfalfa leaf extracts contained a total phenolic content of 37 mg GAE/g extract and a total flavonoid content of 12.6 mg rutin equivalent/g of dry matter.⁴⁵ Antioxidant status was evaluated by several methods. One of these measurements, a free radical scavenging activity method (diphenyl-1-picrylhydrazyl; DPPH), showed 54% inhibition from alfalfa crude extract of free radicals at a concentration of 250 µg/mL. When compared to other known antioxidant ingredients (vitamins E and C), alfalfa crude extract showed 54% inhibition compared to vitamin E at 90% and vitamin C at 91%. Secondly, a nitric oxide (NO) inhibition scavenging activity model of the same concentration of alfalfa crude extract at 250 mg/mL showed inhibition of 50.9% and vitamin C and E controls, reported at 94% and 91%.⁴⁵ Also, various in vitro models found antioxidant activity with an alfalfa root ethanol extract.⁶⁰

Antimicrobial properties of alfalfa have been reported in various extracts made with organic solvents such as methanol, chloroform, and ethanol.⁶¹ Alfalfa root extract exhibited antimicrobial activity against three bacterial strains, Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. The inhibitory effects were demonstrated by MIC and reported at 125 mg/mL.⁶²An in vitro study found alfalfa shoot extracts demonstrated antileishmanial activity, especially with increased concentration and incubation time. The IC50 values (measurement of inhibition by 50% of the given substance) was reported respectively at 24 hours (556.62 µg/mL), 48 hours (445.74 µg/mL) and 72 hours (375.74 µg/mL). The authors also reported that antiparasitic activity may be attributed to high concentrations of phenolic compounds.⁶³

A four-month study found a diet incorporating 1-2% alfalfa saponins and 40% alfalfa seeds decreased hypercholesterolemia⁵⁰ in rabbits that were provided a cholesterol-enhanced diet. Additional findings reported decreases in cholesterol concentrations in the liver and aorta.⁵⁰ A specialized alfalfa extract without coumestrol and L-canavanine given to rabbits on a cholesterol-enhanced diet significantly decreased total cholesterol by 85% compared to a 73% reduction by gemfibrozil (a cholesterol-lowering pharmaceutical medication).⁶⁴ The alfalfa-treated group also showed anti-atherosclerotic activity,⁶⁴ similar to the results of Malinow et al (1980).⁵⁰ A total of five compounds were isolated from aerial parts of alfalfa, which all showed hypolipidemic properties, determined by measuring triglyceride levels in HepG2 cells, compared to the control simvastatin (a cholesterol-lowering medication).⁶⁵

Additionally, alfalfa, like several plants in the legume family, is associated with estrogenic activity.⁹ A methanolic alfalfa sprouts extract showed estrogenic activity in an estrogen-dependent MCF-7 breast cancer cell proliferation assay model. Various ranges of extract concentrations were tested in the model, and alfalfa sprout extracts (methanol) demonstrated cell proliferation induction at the highest concentration level (100 µg/mL).⁶⁶

Toxicity

Animal studies have found that high quantities of alfalfa seeds and alfalfa seed sprouts, which contain a non-protein amino acid called L-canavanine (which is an L-arginine analog known to be cytotoxic in animals), can induce auto-immune symptoms similar to the human disease systemic lupus erythematosus (SLE).^9,47,67-69 An in vitro study found that L-canavanine had a dose-related effect on human immunoregulatory cells.^47,70 In comparing L-canavanine content in alfalfa plant parts, L-canavanine was found in much higher amounts in alfalfa seeds (80–150 mg/kg) compared to alfalfa leaves (10 mg/kg).^9,71

In a six-month study, rats were given 1% or 2% alfalfa saponins (isolated from alfalfa aerial parts) in their diet, and authors reported no side effects of toxicity in either treatment group.^67,72

Salmonella and Escherichia coli outbreaks have been reported with the consumption of contaminated alfalfa sprouts in the United States,⁷³ with the last outbreak reported in 2016. Several food safety lists include alfalfa sprouts and other types of raw sprouts (clover, mung bean, and radish), which have been linked to foodborne illnesses.⁷⁴

Traditional preparations of alfalfa leaf are perceived to be safe at recommended dietary intake levels and classified as GRAS (Generally Recognized as Safe) in the United States. However, excessive consumption of alfalfa that exceeds the recommended dietary intake levels is contraindicated, as pharmacological effects have been demonstrated in some studies.^47,75 In addition, Health Canada contraindicates the use of alfalfa preparations if one has a history of SLE and advises caution when consuming alfalfa preparations for people who use blood thinners, birth control medications, or hormone replacement therapy.¹⁴ In 2009, an alfalfa protein concentrate was approved as a novel food ingredient by the European Food Safety Authority with the opinion that alfalfa protein concentrate (using only the aerial parts) is considered safe for humans within the recommended intake levels.^9,52,71

Human Clinical Studies

Human clinical studies on alfalfa preparations are scant. A few studies were identified that relate to traditional antidiabetic uses of alfalfa leaves; however, these studies had too few participants to draw meaningful conclusions.^76,77

A potential research focal point may be in understanding alfalfa as a valuable nutrition source for humans.^71,78 A randomized, controlled two-arm study was conducted to compare daily supplementation of iron and folic acid to an alfalfa leaf concentrate in 102 adolescent girls with anemia (14-18 years of age) for three months. Daily supplementation of 60 mg iron (ferrous sulfate [FeSO4] tablet) and 500 µgfolic acid (Rajasthan Drugs and Pharmaceutical Ltd.; Jaipur, India) were given to one group and compared to an alfalfa (lucerne) dry leaf concentrate powder (Frand-Luzerne Agricultural Co-Operative; Aulnay-aux-Planches, France), which contained 5 mg of iron and 13 µgof folic acid. Instructions for the alfalfa leaf powder arm of the study was to take orally with water, lemon (Citrus limon, Rutaceae) water, or buttermilk (10 g of powder daily provided in a pouch). Each participant at the beginning of the study was also provided one 400-mg tablet of albendazole (the antiparasitic medication Zentel^®; GSK; Brentford, UK) and received daily home visits throughout the three-month study. Side effects (e.g., nausea, diarrhea, vomiting) were reported in about 20% of the iron and folic acid supplementation group compared to 2% in the alfalfa powder group due to poor taste. At baseline, 3.9% of participants were severely anemic, 27.5% were moderately anemic, and 68.6% were mildly anemic. A total of 86 participants from both groups completed the study, and none were severely anemic, 10.5% were moderately anemic, 30.2% were mildly anemic, and 59.3% were no longer anemic with normal hemoglobin levels. The authors concluded that alfalfa leaf concentrate powder, after baseline value adjustments, were as effective as supplementation of iron and folic acid in improving anemia.⁷⁹

ADULTERATION AND SUBSTITUTION

The alfalfa monographs of both Pharmacopée Française and the Homoeopathic Pharmacopoeia of the United States indicate the possibility of adulteration with Medicago falcata,^3,80 which is also native to much of Europe and Asia.⁸¹ As M. sativa does not have any yellow flowers, the presence of yellow flowers may indicate adulteration by M. falcata.

Although M. falcata is classified as an adulterant in pharmacopoeias, two botanical species and their subspecies may be referred to by the common names “lucerne” or “alfalfa”: M. sativa subsp. sativa (with violet, dark blue to purple flowers) and M. falcata subsp. falcata (with yellow flowers). Under natural conditions, M. sativa and M. falcata cross with each other giving rise to the hybrid Medicago × varia (syn. Medicago × media). The hybrids have flowers of mixed colors: purple, yellow, blue, and sometimes white. It is difficult to distinguish between M. sativa subsp. sativa and M. × varia based on flower color alone, as the hybrids are characterized by great morphological variability. Western European and American cultivars are presumably classified as M. sativa and Polish and Eastern European cultivars as M. media (E. Gaweł email to T. Smith, October 24, 2022).

SUSTAINABILITY AND FUTURE OUTLOOK

Medicago sativa is globally assessed as “Least Concern” (LC), meaning it is not considered to be threatened according to the Red List Categories and Criteria of the International Union for Conservation of Nature (IUCN), “as its wild subspecies are widespread across Eurasia with no major threats and stable populations. They are also well conserved in ex situ gene bank collections. This species would benefit from a gap analysis to ensure that the likely range of in situ genetic diversity is fully represented in gene bank collections.”⁷

Genetic conservation of crop wild relatives is important for plant breeding, especially in view of climate change adaptation. The diversity of crop wild relatives, including of M. sativa, in the eastern Mediterranean region and North Africa is reportedly the highest globally.⁸² New and old M. sativa genotypes have been field tested in Greece under irrigated Mediterranean conditions with a goal toward breeding of new cultivars that may exhibit greater heat tolerance, summer productivity, height, and yield in view of predicted climatic changes for the region.⁸³ Similar land suitability and climate change adaptation studies for alfalfa are occurring in Algeria,⁸⁴ Ethiopia,⁸⁵ and Iran,⁸⁶ among other countries. Breeding is also carried out for other reasons, for example to increase seed yield, to develop cultivars with a slower rate of protein decomposition, and to develop low-saponin or saponin-free cultivars. (Saponins cause bloating in the digestive tract of ruminants, which can lead to the death of the animal [E. Gaweł email to T. Smith, October 24, 2022].) Given the global importance of M. sativa as a fodder and forage crop, and lesser but increasing importance as a medicinal and nutritional crop for human health, prioritization of suitable site selection and cultivar selection for continued large scale production can be expected.

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