Purple Willow (Salix purpurea L.) and Its Potential Uses for the Treatment of Arthritis and Rheumatism

CHAPTER 31

Purple Willow (Salix purpurea L.) and Its Potential Uses for the Treatment of Arthritis and Rheumatism Paweł Sulima, Jerzy A. Przyborowski Department of Plant Breeding and Seed Production, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland

1. INTRODUCTION 1.1 General Description of Salix purpurea L. Purple willow (Salix purpurea L., Figure 31.1) is one of the few species of the genus Salix with pharmaceutical applications. The genus Salix includes more than 400 species and an undefined number of natural and artificial interspecific hybrids.1–3 In the most recent taxonomic system proposed by Lauron-Moreau et al.,4 purple willow has been classified to the subgenus Vetrix (Figure 31.2). The species is composed of three subspecies: S. purpurea subsp. lambertiana, S. purpurea subsp. Angustior, and S. purpurea subsp. purpurea. In addition, S. purpurea can produce numerous interspecific hybrids, most often with S. viminalis, S. triandra, S. caprea, and S. fragilis.1, 3 Due to its unique ability to hybridize and produce interspecific hybrids as well as high levels of seasonal and environmental variability, the genus Salix creates taxonomic problems, and its members may be difficult to identify to the species level based on morphological features alone.1, 3, 5 Species-level classification is a key factor that determines the suitability of willow bark for pharmaceutical use, as Salix species differ significantly in their content of pharmacologically active compounds. In the conventional approach, members of the genus Salix are classified to the species level based on morphological features such as color, shape, and hairiness of leaves and shoots as well as the shape and color of flowers. At present, more objective techniques that rely on the analyses of DNA sequences can be deployed in the classification process. These methods have been long used to elucidate the taxonomic complexity of the genus Salix.6–9 Significant progress has been made in the taxonomic identification of S. purpurea, and its genomic sequences are now widely available.10, 11 In recent years, purple willow acquired the status of a model species in studies investigating the molecular determinants of biomass production and genetic improvements of various willow species.12, 13

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Figure 31.1 Purple willow, Salix purpurea L. (Phot. P. Sulima from Sulima P, Krauze-Baranowska M, Przyborowski JA. Variations in the chemical composition and content of salicylic glycosides in the bark of Salix purpurea from natural locations and their significance for breeding. Fitoterapia 2017;118:118–125.)

Plants are classified to the level of S. purpurea species based on a variety of characteristic morphological features. Purple willow is a dioecious and outcrossing plant (Figure 31.3A and B) that produces medium-tall and tall shrubs and sometimes trees with a height of up to 6–8 m.5 Young shoots are straight, thin, very flexible, dense, and usually purple (Figure 31.1), whereas older shoots take on a gray-green hue. Purple willow has inversely lanceolate, matte leaves that are dark green on the adaxial surface and blue-green on the abaxial surface. Young leaves at the tip of shoots are often purple-red. Older leaves are generally arranged alternatively on the stem, but they can also have an opposite arrangement, a feature that is unique to purple willow within the genus Salix. Purple willow leaves have a smooth margin at the bottom and a serrated edge in the upper half. They

Purple Willow (Salix purpurea L.) and Its Potential Uses

Figure 31.2 Taxonomic classification of the genus Salix spp. (Based on Lauron-Moreau A, Pitre FE, Argus GW, Labrecque M, Brouillet L. Phylogenetic relationships of American willows (Salix L., Salicaceae). PLoS ONE 2015;10(4):e0121965.)

(A)

(B)

Figure 31.3 Female flower (A) and male flower (B) of S. purpurea. (Phot. P. Sulima.)

are broadest toward the end rather than at the base. Leaf length is 5–7 times greater than leaf width. The absence of stipules is a characteristic feature of purple willow.1, 2, 5, 14 Purple willow grows mainly in cool and temperate climates of the northern hemisphere, where it is regarded as a common species.1, 3 It is native to Europe and Asia,

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but S. purpurea has also been introduced to other regions (North America and certain European regions) for cultivation on arable land and to limit erosion along lake shores and river banks.2, 3, 15 The biomass of S. purpurea has numerous applications, the species has a high yield potential, and it regenerates rapidly after annual harvests. Purple willow produces straight, elastic shoots with a thin core. It has a well-developed root system, and can be grown in both waterlogged and water-deficient soils.2, 5 Purple willow bark is used mainly for pharmaceutical purposes as a source of natural medicines, commonly called natural aspirin due to the similar therapeutic properties to synthetic aspirin. These applications will be described in greater detail in successive subchapters. The discussed species also has a variety of other economic uses. Purple willow has been long used as source of wicker for the production of ornaments and furniture.2, 16 In recent decades, willow biomass has emerged as an important source of renewable energy. The main willow species used for energy generation are S. viminalis, S. dasyclados, S. pentandra, and S. triandra, but according to many authors, S. purpurea is also a valuable source of biomass.13, 17, 18 Purple willow is characterized by a slower growth rate than taxa that are more widely cultivated for biomass, but it can be grown in significantly denser stands and, consequently, it produces similar biomass yields.19–21 Purple willow also plays an important role in environmental protection and landscape shaping.22 The potential of S. purpurea for the phytoremediation of areas contaminated with heavy metals and for the purification of sediments, surface, and ground waters has been recognized in recent years.23–25 The biomass of S. purpurea is less widely used in the production of cellulose, paper, and furniture (boards and panels). Purple willow plants are also a source of nectar for pollinating insects, and they are recognized for their ornamental traits.2, 5, 26

2. PHARMACOLOGICAL ACTIVITY OF WILLOW BARK 2.1 History The medicinal properties of willow were recognized in ancient times. The first written accounts dating back to ancient Mesopotamia (20th century BCE) were highly generalized, and willow was merely indicated as a medicinal plant. More detailed information was discovered in Egyptian papyrus records from the mid-16th century BCE. Willow and myrrh leaves were used as analgesics in ancient Egypt. In ancient Greece (5th century BCE), Hippocrates advocated the use of willow bark extracts to treat labor pains and fever. Willow leaf extracts were a common remedy for fever, swelling, pain, and inflammations in ancient Rome.27–29 In modern times, the medicinal properties of willow bark were recognized by Edward Stone,30 an English cleric who used powdered willow bark to treat malaria and fever. In the 19th century, the progress made in organic chemistry, a young branch of science at the time, expanded our knowledge of biologically active substances in plants. Plant compounds were isolated and purified by many scientists. A pharmacologically active

Purple Willow (Salix purpurea L.) and Its Potential Uses

compound was first isolated from boiled willow bark and purified in 1828 by the German pharmacist Johann Buchner. Buchner named the isolated compound “salicin” after Salix, the Latin botanical name for willow. Salicin was recognized as an antipyretic ingredient of willow bark.31 In 1839, Raffaele Piria developed a method for extracting salicylic acid (SA), which was used in the production of water-soluble sodium salicylate. By the mid-nineteenth century, medicines containing salicin, SA, and sodium salicylate were commonly used, and their analgesic, antipyretic, and antiinflammatory properties were widely recognized. However, the active ingredients of willow bark had unpleasant side effects, such as the ability to irritate the gastric mucosa.28, 29, 32 The chemical industry grew dynamically in the second half of the 19th century, and in 1859, Hermann Kolbe developed chemical processes for the mass production of synthetic SA. In successive years, synthetic acetylsalicylic acid (ASA) became an alternative to SA. Acetylsalicylic acid was first obtained in 1852 by Charles Frederic Gerhardt, who combined acetyl chloride with sodium salicylate. But it was the scientists of Bayer, the German pharmaceutical and chemical company, who took full credit for the discovery. In 1897, Felix Hoffmann, a young chemist employed by Bayer, developed a more effective method of obtaining ASA by reacting salicylic acid with acetic anhydride.32 As a result, ASA was synthesized in a pure and chemically stable form. Two years later, Bayer launched a new product, aspirin, which continues to be one of the most popular drugs in the world. Aspirin proved to be an effective broad-spectrum drug. The synthesis of ASA is not a complex process, which contributed to its commercial success.28, 29 Acetylsalicylic acid has been used in clinical practice for many years, but its mechanism of action was explained only in 1971 by the research team headed by John Vane. Vane and coworkers discovered that the multidirectional action of ASA is linked with its ability to inhibit the biosynthesis of prostaglandins that control bodily responses to injury and infection. Prostaglandins affect the nervous system’s ability to transmit pain stimuli, influence blood clotting, and cause inflammation as a local immune response. Vane and coworkers reported that both ASA and salicylates inhibit inflammatory processes, but through a somewhat different mechanism of action.33 In 1982, their discovery was awarded the Nobel Prize in Medicine. Somewhat later, it was also determined that ASA inhibits COX-1 and COX-2 cyclooxygenases, which testifies to its antiinflammatory properties (COX-2) and explains common side effects, in particular irritation of the gastric mucosa (COX-1).28, 29, 32 At present, ASA is used in the treatment of various diseases and complaints, including inflammation of the joints, tendons and arteries; rheumatic pain; degenerative changes in the spine and joints; fever; nerve pain; headaches; and toothaches. Clinical studies have revealed that regular aspirin use decreases the risk of heart attack and stroke.29, 34 Small doses of aspirin can prevent serious diseases such as colorectal cancer, prostate cancer, and Alzheimer’s disease.35–37 Clinical studies have also demonstrated that ASA has a number of side effects, some of them serious, such as blood coagulation disorders. Other side

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effects include damage to the gastric mucosa, gastrointestinal microhemorrhage, kidney and liver irritation, and miscarriage and bleeding in pregnant women. Aspirin idiosyncrasy has been reported, particularly in children and adolescents. Acetylsalicylic acid has also been linked with Reye syndrome, a rapidly progressive encephalopathy accompanied by fatty liver, which can lead to serious brain damage or even death.29, 38

2.2 Medicinal Properties of Willow Bark The medicinal properties of willow bark fell into oblivion in the era of synthetic drugs, which began with the discovery of ASA synthesis in 1897. At the end of the 20th century, the interest in willow bark was revived among scientists, pharmacists, physicians, and patients due to growing levels of knowledge about the risks associated with the use of synthetic drugs containing ASA, the medicinal properties of willow bark, and the growing popularity of natural remedies. Willow bark has similar uses to ASA, but far fewer side effects, which is confirmed by the results of clinical trials.39–41 Willow bark is one of the few plant substances with such a broad spectrum of action, and it has analgesic, antipyretic, antiinflammatory, antithrombotic, antirheumatic, antibacterial, and antimicrobial properties. The antiinflammatory properties of willow bark are associated with the inhibition of COX-1 and COX-2 cyclooxygenases27 and proinflammatory cytokines such as IL-6, TNFα, and IL-3.42, 43 Willow bark exerts antioxidant effects and protects articular cartilage.44 Recent research has demonstrated that willow bark possesses anticarcinogenic properties.45–48 Hostanska et al.45 analyzed the influence of salicylic alcohol, flavonoids, and proanthocyanidins isolated from willow bark extract on the proliferation and apoptosis of human colorectal cells and various cancer cells. All the examined compounds exhibited antiproliferative activity. Enayat and Banerjee46 reported high levels of antioxidant, phenolic, and flavonoid activity in ethanol extracts of S. aegyptiaca bark. In cancer cells incubated with willow bark extract, the preparation decreased cell proliferation, induced cell apoptosis, and disrupted the cell cycle by inhibiting the phosphatidyl inositol 3-kinase/protein kinase B and mitogen-activated protein kinase signaling pathways. The observed effects were similar in magnitude to those of commercial inhibitor drugs. Willow bark extract strongly inhibited the growth, mobility, migration, and adhesion of HCT 116 and HT-29 colorectal cancer cell lines. The above reaction was accompanied by the restoration of e-cadherin expression and the reduction of EGFR, SNAI1, SNAI2, Twist1, and MMP9 and MMP2 matrix metalloproteinases. Many of these proteins participate in epithelial-mesenchymal transition, the key stage in the progression of benign cancer cells to malignant cells. Enayat and Banerjee46 demonstrated that the extract from the bark of S. aegyptiaca is a potent nutraceutical candidate for chemotherapy. The broad-spectrum medicinal properties of willow bark can be attributed to its unique composition, namely the presence of salicylic glycosides (SG), flavonoids,

Purple Willow (Salix purpurea L.) and Its Potential Uses

phenolic acids, and catechin.49 Salicylic glycosides such as salicin, salicortin, tremulacin, salireposide, picein, triandrin, populin, fragilin, tremuloidin, acetyl salicortin, grandidentin, and salicyl tremulacin are responsible for the main medicinal properties of willow bark.41, 50 Flavonoids and phenolic acids also exert therapeutic effects by modulating inflammatory control mechanisms (antiinflammatory properties) and inhibiting enzymes that catalyze arachidonic acid, including cyclooxygenases, lipoxygenases, and phospholipases.44 Flavonoids are also mild diuretics, and they are useful in the treatment of metabolic disorders such as rheumatic diseases. Other flavonoids include isoquercetin, naringenin, naringenin-7-O-glucoside, naringenin-5-O-glucoside, and isosalipurposide. Willow bark also contains other significant groups of flavonoids: flavonols, flavones, chalcones, flavanones, and flavone dimers.41, 50 Phenolic acids are the least well-known group of compounds in the chemical composition of willow bark. The most widely occurring phenolic acids include salicylic, vanillic, syringic, p-hydroxybenzoic, p-coumaric, ferulic, and caffeic acids.41, 50 Catechins and catechin polymers have antiinflammatory, antimicrobial, and astringent properties, and they alleviate intestinal inflammations.41, 51 In addition to (+)-catechin, gallocatechin, and catechin dimers, willow bark also contains tannins and flavan-3-ol derivatives, such as procyanidins and prodelfinidins. The medicinal properties of willow bark can be attributed to the presence of various pharmacologically active compounds and their synergistic interactions.44

2.3 Metabolism of Salicylic Glycosides Willow bark contains many pharmacologically active compounds that are metabolized differently than ASA in the human body and have fewer side effects than synthetic ASA.40, 52–55 Orally administered SGs are hydrolyzed to glucose and saligenin (salicylic alcohol) by bacteria in the upper gastrointestinal tract. After absorption in the stomach, saligenin is oxidized to salicylic acid, and it does not cause damage to gastric mucosa. In contrast, synthetic ASA is hydrolyzed to salicylic acid and acetic anhydride, and acetylation causes damage to gastric mucosal cells. Willow bark preparations are absorbed in the gastrointestinal tract at a slower rate than ASA, and their therapeutic effects last longer. The concentration of bioactive compounds in the blood peaks 2–3 hours after oral administration of SG, and these substances are completely absorbed.40, 52–56 As a weak organic acid, salicylic acid is undissociated in the acidic gastric content (pH of around 2), and it easily penetrates gastric mucosal cells. Cells contain mainly the dissociated form of salicylic acid (pH of around 7), which weakly penetrates cell barriers. Therefore, only small amounts of undissociated salicylic acids freely migrate to capillaries. Dissociated salicylic acid is accumulated in the gastric mucosal cells, and it can cause damage or even necrotic changes by lowering pH and increasing osmotic pressure inside cells. These risks are not encountered in willow bark preparations that contain trace amounts of

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salicylic acid and where the released buffer compounds (tannins and flavonoids) exert protective effects.40, 54 Salicin is metabolized to salicylic acid in the liver, which eliminates the ion trap risk. According to Ulrich-Merzenich et al.,56 salicylates and polyphenols enter into synergistic interactions that deliver antiinflammatory effects. For this reason, SG can be used at lower concentrations to avoid the unpleasant side effects.

3. THE POTENTIAL OF S. PURPUREA BARK FOR RHEUMATISM TREATMENT 3.1 Phytopharmaceuticals in the Treatment of Rheumatic Diseases There are more than 200 autoimmune rheumatic conditions that involve painful, inflammatory, degenerative, structural, and functional disorders of the musculoskeletal system. Rheumatic disorders and their causes are difficult to diagnose. Genetic factors, immune disorders, infections, cancer, and various medications can contribute to the development of rheumatic conditions. Unless diagnosed correctly and early enough, rheumatic disorders can damage the locomotive system, leading to disability or even death. To achieve remission, rheumatism therapy should begin as soon as possible, preferably within 6–12 weeks after the appearance of the first symptoms.57 Analgesics and antiinflammatory drugs are widely used in the treatment of rheumatic conditions. Phytopharmaceuticals play an increasingly important role in the therapeutic process because they contain many bioactive compounds and exert positive synergistic effects in rheumatism treatment.58 Plant-based medicines not only have to undergo clinical tests to confirm their efficacy, but also other accurate tests to determine their exact composition, mechanisms of action, factors that influence their efficacy, and possible side effects. Phytopharmaceuticals that meet these criteria59 should be administered for pain management before nonsteroidal antiinflammatory drugs (NSAIDs) because they have far fewer side effects than synthetic drugs.60, 61 Plant-based drugs generally have a broader spectrum of action than conventional analgesics, and they are particularly recommended for managing chronic musculoskeletal pain. Nonsteroidal antiinflammatory drugs are characterized by a nonselective or preferential mode of action, or they selectively inhibit COX-2 cyclooxygenase. In addition to inhibiting cyclooxygenases, plant-based antirheumatic drugs also inhibit lipoxygenase to a varied degree, exert antioxidant effects, and partially inhibit the release of cytokines, including cytokines that damage articular cartilage.62 Nonsteroidal antiinflammatory drugs have many side effects that are sometimes very serious,63, 64 such as gastrointestinal complications, indigestion, gastric perforation, ulcers, and hemorrhages.65, 66 In comparison with NSAIDs, the benefits of phytopharmaceuticals seem to outweigh the risks. Plant-based drugs rarely produce serious side effects. Mild gastrointestinal complaints and allergies are sometimes reported. A metaanalysis conducted by Cameron et al.64 revealed that low acute toxicity of willow bark had been reported in only three preclinical trials. There are no published data on the

Purple Willow (Salix purpurea L.) and Its Potential Uses

chronic toxicity of willow bark, including mutagenicity, carcinogenicity, teratogenicity, and embryogenicity.64 According to Cameron et al.,64 rheumatism sufferers reach for phytopharmaceuticals also for nonmedical reasons, such as dissatisfaction with the outcomes of conventional treatment, the desire to actively manage the therapeutic process, belief in the philosophy of alternative medicine, and concerns regarding the side effects of conventional treatments. Plant-based medications are popular among the proponents of alternative medicine. Patients are more eager to reach for phytopharmaceuticals than synthetic drugs. The most popular plant-based rheumatism remedies contain extracts of Harpagophytum procumbens (roots), Uncaria tomentosa (roots), Zingiber officinale Rosc. (roots), Rosa canina (fruit and seeds), Boswellia serrata (resin), Persea americana (essential oils), Urtica dioica (leaves), Petiveria alliacea (leaves), Capsicum L. (fruit), and the bark of the following Salix species: S. purpurea, S. daphnoides, S. alba, and S. fragilis.41, 64, 67

3.2 Willow Bark Medicines for Rheumatism Treatment The number of medicinal products containing willow bark, including the bark of S. purpurea, is on the rise. This can be attributed mainly to the complex pharmacological activity of willow bark ingredients, the few and relatively mild side effects, and the growing levels of awareness about the pharmacological activity of plant-based products. The chemical standardization of herbs and the results of clinical trials confirming the effectiveness and safety of modern phytopharmaceuticals have increased the popularity of plant-based medicines among physicians and patients.68, 69 Unlike their predecessors, contemporary willow bark preparations contain high levels of SG and polyphenols, which contributes to their potency.44 Willow bark is used in the production of various medicinal products in the form of liquid extracts, tinctures, solutions, dry extracts, and tablets containing powdered bark. These preparations are used mainly in the treatment of the common cold and rheumatic conditions.40, 41, 55, 64, 70 In contrast to synthetic drugs, willow bark preparations contain a variety of compounds that do not completely block or maximally stimulate biochemical processes. As a result, medicinal products containing willow bark extracts are more effective, they are well tolerated, and they stimulate natural regulatory responses to pathological processes. Chronic pain in rheumatism can be managed primarily with willow bark extracts and supplemented with synthetic drugs if necessary.71 Unlike synthetic rheumatism drugs that block cyclooxygenases, phytopharmaceuticals containing willow bark extracts have a broader spectrum of action and fewer side effects due to their varied composition.72 Pharmacological studies have demonstrated that willow bark has antioxidant properties and decreases the synthesis of prostaglandins by inhibiting cyclooxygenase and the synthesis of leukotrienes and cytokines.73 Flavonoids exert analgesic effects through their antioxidant properties and the ability to inhibit lipoxygenase. Willow bark extracts have been found to inhibit thrombocyte aggregation to a lesser extent than ASA.74

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Willow bark extracts are well-known and well-tested rheumatism remedies with analgesic and antiinflammatory properties and low risk associated with the presence of various ingredients, including SG, flavonoids, phenolic acids, and catechins. This is a very important consideration because rheumatism is a chronic condition that requires prolonged treatment. Conventional drugs such as NSAIDs often produce cardiovascular and gastrointestinal side effects and have to be used with expensive medications that protect the gastric mucosa, which is why willow bark extracts offer an interesting alternative.67, 71 Willow bark preparations have been used to treat rheumatic disorders of soft tissues and joints that are accompanied by inflammation and pain, gout, ankylosing spondylitis, rheumatoid arthritis, and other connective tissue diseases with inflammatory changes.39, 72

3.3 Composition of S. purpurea Bark The range and content of salicylic glycosides and other pharmacologically active compounds in willow bark differ both within and across species.75 For this reason, only several willow species, such as S. alba, S. daphnoides, S. fragilis, S. nigra, and S. purpurea, are regarded as a source of medicinal compounds, provided that salicis cortex contains 1.5% salicin and salicis corticis extractum siccum contains 5.0% salicin.41, 76–78 In the pharmaceutical industry, raw materials have to be characterized by high quality, appropriate and tested composition, and high and balanced content of pharmacologically active ingredients. Willow bark for the production of natural aspirin is harvested mainly from natural stands; therefore, it is not genetically homogeneous, and the obtained material has to undergo numerous and costly analyses and standardization procedures. Raw materials for pharmacological products are also in short supply. According to Sulima et al.,75, 79 this problem could be resolved by field cultivation of willow varieties whose bark is characterized by the appropriate chemical composition and a balanced content of pharmacologically active compounds. Controlled willow cultivation could offer a safe and stable source of high-quality bark for the production of natural aspirin. The bark of S. purpurea and S. daphnoides is most abundant in SG. However, in our previous study,80 S. daphnoides plants were characterized by significantly lower productivity and health, and considerable losses were noted already in the first years of the study. For this reason, S. purpurea appears to be a more reliable source of pharmaceutical raw materials.75, 79 According to research, purple willow bark contains 3%–11% salicylic glycosides.41, 75, 81, 82 The composition of SG in S. purpurea bark varies considerably, and this group of compounds is represented mostly by salicin (0.5%–1.7%), salicortin (2.3%– 3.9%), and compounds that occur in much smaller quantities (below 1%) irregularly and are not found in all S. purpurea genotypes, such as salireposide, fragilin, populin, tremuloidin, triandrin, tremulacin, picein, acetylsalcortin, grandidentin, salicyl tremuloidin, salicyl tremulacin, salicoysalicin, syringing, and purpurein.41, 75, 83, 84 Purple willow bark

Purple Willow (Salix purpurea L.) and Its Potential Uses

also contains the chalcone component isosalipurposide (0.2%–2.2%), flavanone-7-Oglucoside (0.2%–0.4%), naringenin-5-glucoside (0.4%–1.5%), catechin (0.5%), polyphenols (5%), naringenin, and flavan-3-ol catechin.41, 51, 75 The content of SG compounds in willow bark is correlated with the harvest date.81, 82, 85 F€ orster et al.82 observed the highest concentrations of salicylic compounds and phenolic glycosides in willow bark harvested in March and October. The content of secondary metabolites in bark decreases significantly during the growing season. The lowest concentrations are noted in July and August, after which they increase until the end of October. Similar observations were made by Thieme.85

3.4 The Applicability of S. purpurea Bark for the Treatment of Arthritis and Rheumatism The bark of S. purpurea contains a wide range of SGs, flavonoids, phenolic acids, and catechins, and it is highly suitable for the production of rheumatism medicines. Rheumatism treatments containing S. purpurea bark include Assalix, Proaktiv, Optovit Actiflex, Rheumakaps, Salix B€ urger, Assplant, Salix purpurea Urtinktur, Salix purpurea LM 27, and Lintia in Germany, and Salicortex in Poland.41 The efficacy and safety of S. purpurea bark extracts have been confirmed by numerous clinical trials involving nearly 2000 patients.39–41, 59, 86, 87 When administered at a daily dose of up to 240 mg, salicin exerts potent analgesic, antiinflammatory, and antirheumatic effects. The key findings of the relevant clinical trials are discussed below. In a randomized double-blind study of 210 patients, Chrubasik et al.59 demonstrated that Assalix containing S. purpurea bark extract was an effective and safe preparation for managing lower back pain in arthritis and rheumatism conditions. The efficacy of the preparation was dose-dependent. In the group administered a higher dose (corresponding to 240 mg of salicin), positive effects were observed already in the first week of treatment, and pain was completely eliminated in 39% of the subjects in the last, fourth week of the study. In the group receiving a lower dose of the medication, pain was eliminated in 21% of patients versus only 6% in the placebo group where tramadol had to be administered as a rescue medication. These observations were confirmed by an open-label, nonrandomized study of 451 patients that monitored the market introduction of Assalix.86 The dose equivalent to 240 mg of salicin produced the most satisfactory results, and pain was eliminated in 40% of the patients after 4 weeks. The authors also demonstrated that Assalix reduced average treatment costs, and the daily dose of 120 mg decreased average treatment costs by even 35%–50% per patient. Chrubasik et al.87 compared the efficacy of Assalix and Rofecoxib, a nonsteroidal antiinflammatory drug and a selective COX-2 inhibitor, in a randomized double-blind study of 228 patients with lower back pain. Based on the calculated values of the Arhus Index and the Total Pain Index, the researchers demonstrated that both medications had

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similar efficacy and that the plant-based drug was characterized by significantly lower treatment costs. In a study by Biegert et al.,55 S. daphnoides bark extract (daily dose of 240 mg total salicin) was not effective in the treatment of osteoarthritis. The randomized placebocontrolled trial was conducted on 43 patients, and diclofenac was the reference drug. The bark of S. purpurea has a different composition than S. daphnoides, and a comparative study of natural remedies containing S. purpurea extract could provide interesting results. Beer and Wegener39 reported on the clinical efficacy of willow bark extract in patients with osteoarthritis. The trial involved 83 patients who received 70% ethanol extract of willow bark (equivalent to 240 mg of total salicin) and 41 patients undergoing standard NSAID treatment. The results of this open-label, multicenter trial revealed that the willow bark extract was as effective as the standard therapy after 6 weeks of treatment. Vlachojannis et al.40 demonstrated the effectiveness of willow bark in the treatment of musculoskeletal pain. The cited authors concluded that willow bark extract is a promising analgesic in the treatment of acute back pain and that it has few side effects. The potential of willow bark to relieve pain makes it a suitable candidate for pain management in osteoarthritis and rheumatoid arthritis. According to Vlachojannis et al.,40 the efficacy of the preparation could be increased by raising the daily dose to the equivalent of 240 mg of salicin. Further research is needed to verify the efficacy of willow bark extract in the treatment of osteoarthritis and rheumatoid arthritis and to evaluate the toxicity of higher doses of plant-based pharmaceuticals. A study by Freischmidt et al.51 revealed that S. purpurea bark has additional medicinal properties. A liquid extract of S. purpurea bark exerted a significant influence on the cytokine TNF-α (cachexin), which is produced in inflammatory processes. In the cited study, the expression of the ICAM-1 adhesion molecule decreased by up to 40% in human vascular epithelial cells relative to control. Adhesion molecules such as ICAM-1, VCAM, and selectins participate in inflammatory processes, which testifies to the antiinflammatory properties of S. purpurea bark extracts, in particular catechin. Cytotoxic effects were not observed in the cited study, which is an additional advantage of purple willow bark extracts.51

4. CONCLUSIONS The bark of S. purpurea belongs to a small group of plant extracts with a diverse range of pharmacologically active components, including salicylic glycosides, flavonoids and other polyphenols, that exhibit multiple medicinal properties. Only selected Salix species, including S. purpurea, S. daphnoides, and S. acutifolia, contain flavanones and chalcones. The bark of S. purpurea is a source of naringenin derivatives (flavanones), isosalipurposide, and its esters (chalcones). Numerous clinical trials have demonstrated that preparations containing S. purpurea bark extract are effective in the treatment of rheumatic conditions. In an open letter to the

Purple Willow (Salix purpurea L.) and Its Potential Uses

editors of Phytotherapy Research, Vlachojannis et al.40 claimed, based on the results of their studies and the findings of many other authors, that the powerful analgesic and antiinflammatory effects of willow bark can be attributed not only to SG, but also to flavonoids and other polyphenols. The complex active substance in willow bark has a much broader spectrum of action that ASA, and it does not produce serious side effects. Unlike synthetic aspirin, willow bark does not damage gastric and intestinal mucosa. Willow bark extracts have a considerable advantage over NSAIDs and ASA in that they do not disrupt the aggregation of blood platelets.74 The only side effects associated with willow bark are rare cases of skin allergy. According to some authors, willow bark can also cause an anaphylactic reaction in people allergic to salicylates.88 Despite significant progress in research into the pharmaceutical uses of willow bark, observed over the past two decades, including analyses of pharmacologically active compounds, the mechanism of action of individual ingredients and the final medicinal product, the positive results of clinical trials and drug safety trials, and the economic benefits of drug-based plants, healthcare professionals continue to be wary or even critical of willow bark products for the treatment of rheumatic conditions.41 The report of the Committee on Herbal Medicinal Products (HMPC) of the European Medicines Agency (EMA)41 pointed to a number of shortcomings in clinical studies into willow bark medicines, including insufficient evidence, small numbers of patients, and short duration. The main criticisms include the lack of comprehensive and detailed analyses of the reproductive toxicity, genotoxicity, and carcinogenicity of willow bark extracts, which leads to the absence of a recommendation from the HMPC. According to the authors, one of the key problems is that the bark of various willow species is regarded as identical, and many pharmaceutical monographs describe several willow species as equally suitable for the production of medicinal products. This is not a reliable approach, and it could create numerous problems in different stages of raw material analysis, drug production, testing, market introduction, and use. The bark of different willow species or even genotypes is never homogeneous, and the resulting medicines can differ significantly in quality, composition, and content of pharmacologically active compounds. In the described approach, nearly every batch of raw materials should be analyzed and standardized, which is likely to increase the cost of the final product. Medicinal plants constitute the basis for medical treatments around the world, but herbal remedies are rarely reimbursed in public healthcare systems.64, 70 The variations in the composition of willow bark also influence the results of clinical trials and prevent a reliable analysis of their therapeutic properties (individual compounds and their synergistic interactions). Field cultivation of selected willow varieties under strictly controlled conditions appears to be a simple and effective solution to the above problem. The resulting biomass would be a stable and safe source of herbal material characterized by high quality, uniform composition, and balanced content of pharmacologically active compounds. Detailed analyses of drugs, their therapeutic effects and potential reproductive

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toxicity, genotoxicity, and carcinogenicity would be performed for specific components, which would facilitate standardization and evaluations of their efficacy. This solution does not preclude the development of plant-based medicines with a more complex composition. The authors agree with Cameron et al.64, 70 that the results of clinical trials should be published in renowned journals and that analytical procedures and statistical methods should be standardized to increase the transparency of research into the medicinal properties of willow bark.

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