The Pharmacological Potential of Rutin

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Abstract

The contemporary scientific community has presently recognized flavonoids to be a unique class of therapeutic molecules due to their diverse therapeutic properties. Of these, rutin, also known as vitamin P or rutoside, has been explored for a number of pharmacological effects. Tea leaves, apples, and many more possess rutin as one of the active constituents. Today, rutin has been observed for its nutraceutical effect. The present review highlights current information and health-promoting effects of rutin. Along with this, safety pharmacology issues and SAR of the same have also been discussed.

Keywords: Anticancer, Antidiabetic, Antimicrobial, Organ protection, Rutin

1. Introduction

Medicinal plants are an integral part of traditional medicine since ancient era. Drug discovery process has witnessed phytochemicals for discovery of new leads (). Flavonoids, polyphenolic compounds, are one of the important classes of plant derived chemicals that contain benzopyrone moiety. About 4000 types of flavonoids have been reported to be present in plants ().

Rutin (3,3′,4′,5,7-pentahydroxyflavone-3-rhamnoglucoside, Fig. 1) is a flavonol, abundantly found in plants, such as passion flower, buckwheat, tea, and apple. It is a vital nutritional component of food stuff (). Rutin, also called as rutoside, quercetin-3-rutinoside, and sophorin is a citrus flavonoid glycoside found in buckwheat (). The name ‘rutin’ comes from the plant Ruta graveolens, which also contains rutin. Chemically it is a glycoside comprising of flavonolic aglycone quercetin along with disaccharide rutinose. It has demonstrated a number of pharmacological activities, including antioxidant, cytoprotective, vasoprotective, anticarcinogenic, neuroprotective and cardioprotective activities ().


2. Pharmacological actions

2.1. Central nervous system

2.1.1. Prevention of neuroinflammation

Rutin has demonstrated the neuroprotective effect on brain ischemia. Administration of rutin caused attenuation of ‘ischemic neural apoptosis’ due to the embarrassment of p53 expression and lipid peroxidation along with increment in ‘endogenous antioxidant defense enzymes’ (). It has been found to be useful in hypoxic, glutamate and oxidative stress (). Reduction of ‘neuroinflammation’ in rat model of ‘sporadic dementia of Alzheimer type’ () and neuroprotective effects in ‘dexamethasone-treated mice’ () were observed on rutin administration.

2.1.2. Promotion of neural crest cell survival

The neural crest is progenitor comprising of neural and mesenchymal potentials. Treatment of rutin to trunk neural crest cells increased their viability without altering cell differentiation and proliferation which could be due to modulation of ERK2 and PI3K pathways ().

2.1.3. Sedative activity

CNS and behavioral activity of rutin were on hole board, thiopental-induced sleeping time and locomotor activity tests in mice. Rutin, given by intraperitoneal route caused a depressant action on the CNS. Research confirmed the CNS depressant activity of rutin was unlikely due to the involvement of GABAA receptor ().

2.1.4. Anticonvulsant activity

Rutin also possesses anticonvulsant activity and seems to be safe for patients with epilepsy as it does not alter the activity of any of the administered antiepileptic drugs nor demonstrates any adverse effects ().

2.1.5. Anti-Alzheimer activity and treatment of hyperkinetic movement disorder

Rutin suppressed activity of proinflammatory cytokines by diminishing TNF-α and IL-1β production in microglia. Such an effect seems to be useful in the treatment of Alzheimer’s disease as evident by prevention of β-amyloid oligomeric cytotoxicity (). Rutin caused attenuation of streptozotocin-induced inflammation by decreasing the activity of the glial fibrillary acidic protein, interleukin-8, cyclooxygenase-2, inducible nitric oxide synthase and nuclear factor-kB and thereby prevented gross anatomical changes in rat hippocampus. Such an effect could be useful in averting cognitive deficits and proves to be beneficial in the treatment of ‘sporadic dementia of Alzheimer type’ ().

‘Tardive dyskinesia’ is a motor disorder of the orofacial region raised due to chronic treatment with neuroleptic drugs, and is considered as a chief clinical concern in the treatment of schizophrenia. In a study, in haloperidol-induced orofacial dyskinesia, rutin treatment reversed behavioral changes such as orofacial dyskinetic movements, stereotypic rearing, locomotor activity, and percent retention along with restoration of biochemical and neurochemical parameters. Thus, rutin seems to be a lead molecule in the treatment of hyperkinetic movement disorder ().

2.1.6. Antidepressant effects

Forced swimming test and tail suspension test in mice were utilized to analyze ‘antidepressant-like effects’ of rutin isolated from Schinus molle. There was a reduction in the immobility time in the tail suspension test. There was no alteration in locomotor activity. Studies demonstrated the antidepressant-like effect of rutin mediated due to increasing the availability of serotonin and noradrenaline in the synaptic cleft ().

2.1.7. Stroke

Stroke could be regarded as a decisive public health issue that seems to be an important cause of fatality and disability in adults globally (). Oxidative stress and inflammation are two of the pathological events observed after ‘ischemic injury’ in the brain (). Protective effect of rutin an animal model of focal cortical ischemia induced by unilateral thermocoagulation of superficial blood vessels of the motor (M1) and somatosensory (S1) primary cortices was studied. Rutin administration significantly promoted recovery of sensorimotor loss which was observed due to the reduction of neurodegeneration in the periphery of cortical injury ().

2.2. Analgesic and antiarthritic activities

2.2.1. Analgesic and antinociceptive effects

Analgesic effect of rutin was studied by hot plate test on swiss albino mice whereby the analgesic effect of rutin was established (). Further, it was also confirmed that rutin exhibited peripheral and central antinociceptive activities ().

2.2.2. Antiarthritic effects

Animals treated with rutin were observed with significant decrement in rheumatoid arthritis and Fanconi anemia by inhibiting ‘oxygen radical overproduction’ (). In adjuvant arthritis rat model, rutin inhibited acute and chronic phases of inflammation. Rutin was the most active in the chronic stage of inflammation (). Due to antifungal and anti-arthritic effects, rutin has a therapeutic effect on septic arthritis caused by Candida albicans (). Further in an independent study, rutin slowed down inflammatory and catabolic cartilage markers in osteoarthritic lesions in the Hartley guinea pig ().

2.3. Endocrine system

2.3.1. Antidiabetic effects

Streptozotocin is a toxic chemical known to deplete levels of insulin by destroying pancreatic islets. Streptozotocin selectively assaults pancreatic β-cells by generating free radicals of oxygen and nitrogen monoxide along with reducing levels of NAD and NADP. Excessive production of glucose and its decreased utilization by tissues serve as the fundamental bases of hyperglycemia (). In a study, chronic administration of rutin in streptozotocin-induced diabetic rats caused a decrement in plasma glucose, augmentation in insulin levels, and restitution of glycogen content and glycolytic enzymes. Significant rejuvenation of pancreatic islets along with diminished fatty infiltrate was observed in rutin-treated diabetic rats (). Diminution of fasting plasma glucose, glycosylated hemoglobin, C-peptide, and malondialdehyde levels was observed in rutin treated streptozotocin diabetic rats (). Rutin averted the levels of enzymes viz. ALT, AST, and LDH in the serum, liver, and heart demonstrating a protective effect on hepatic () and cardiac toxicity () associated due to streptozotocin. Alteration in the activity of matrix metalloproteinase and protection to kidney against streptozotocin-induced damage was observed (). Rutin stimulated glucose uptake in the soleus muscle, and effect was thought to be mediated through extracellular calcium and calcium-calmodulin-dependent protein kinase II activation. Increase in intracellular calcium concentration is involved in DNA activation which was mediated by rutin (). Rutin added for glycemic control via enhancement of insulin receptor kinase activity, thus aided in promoting ‘insulin signaling pathway’ that caused increased GLUT4 translocation and augmented glucose uptake ().

2.3.2. Anti-hypercholesterolemic effects

Rutin is a ‘selective and non-toxic modulator’ of hypercholesterolemia. In a study conducted on diet-induced hypercholesterolemic Golden Syrian hamster model, rutin significantly reduced plasma triglyceride levels in experimental animals (). Along with this, rutin also caused a decrement in levels of total cholesterol and HDL cholesterol ().

In a high-cholesterol diet fed male wistar rats, rutin administration demonstrated protective effect against the hepatotoxicity as evident by decrement of the plasma level of alanine transaminase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), and low-density lipoprotein (LDL) (). In another study, it was established that chronic consumption of flavonoids such as rutin could be favorable to cardiovascular health ().

2.3.3. Thyroid uptake promotion

‘Thyroid iodide uptake’ mediated via sodium-iodide symporter plays a pivotal role in thyroid hormone biosynthesis and also plays a key role in diagnosis and treatment of various thyroid diseases. However, some of the patients with thyroid cancer are obstinate to radioiodine therapy, which reduced iodine uptake ability, that remarkably reduces the probability of endurance. Thus, it becomes necessary to search for natural agents that aid in the uptake of thyroid iodide.

In a study, rutin caused a minor reduction in levels of serum T4 and T3 without changing serum thyrotropin. There was a significant increase in hypothalamic, pituitary and brown adipose tissue type 2 deiodinase along with the diminution of liver type 1 deiodinase activities. Administration of rutin was observed with increment in thyroid iodide uptake that could be due to an elevation in the activity of sodium-iodide symporter. The study demonstrates the effectiveness of rutin as an adjuvant in radioiodine therapy ().

2.4. Cardiovascular system

2.4.1. Hypertension

Buckwheat, a rich source of rutin is found to prevent oxidative damage in ‘aortic endothelial cells’ by lowering nitrotyrosine immunoreactivity. Germinated extract of buckwheat demonstrated antihypertensive effect and possibly shelter ‘arterial endothelial cells’ by detrimental effects of oxidative stress (). Reduction in oxidative stress due to rutin, when administered by oral route, is the key reason for the restoration of ‘impaired baroreflex sensitivity’ and ‘vascular reactivity’ in hypertensive rats (). By augmenting NO production in human endothelial cells, rutin improved endothelial functions ().

2.4.2. Blood coagulation

 attempted to study effects of the rutin on the anticoagulant activity of oral warfarin and the protein binding along with pharmacokinetics of its enantiomers in rats. Rutin enhanced the in vitro serum protein binding of S- and R-warfarin. Rutin treatment significantly decreased the elimination half-life of S-warfarin by 37% as a result of the 69% increase in unbound clearance of the S-enantiomer. In a nutshell, concomitant administration of rutin possibly reduces the anticoagulant effect of racemic warfarin ().

2.4.3. Antiplatelet aggregatory effect

Rutin in vitro caused concentration-dependent inhibition of platelet activating factor induced washed rabbit platelet aggregation, and intra-platelet free calcium concentration elevation was induced by platelet activating factor which was inhibited by rutin in a dose-dependent manner ().

2.5. Gastrointestinal system

2.5.1. Antiulcer effects

A peptic ulcer is infirmity that influences the substantial population in the world. Ulcers are observed when disparity occurs among ‘aggressive’ and ‘protective’ factors at the luminal surface of the gastric epithelium. HCl, pepsins, nonsteroidal anti-inflammatory drugs, Helicobacter pylori, bile acids, ischemia, hypoxia, smoking, alcohol, etc. include dynamic features whereas defensive factors comprise of bicarbonate, a mucus layer, mucosal blood flow, PGs and growth factors ().

Ethanol is ill-reputed agent known to produce damage to gastric mucosa in animal and clinical studies. Ethanol in a concentration higher than 400 ml/l causes significance in gross morphology of stomach which is observed by mucosal hyperemia, necrosis, edema and mucosal or submucosal hemorrhage (). Oxygen-derived free radicals could be regarded as a key reason for the formation of lesions (). Rutin pretreatment before administration of ethanol afforded significant protection against necrosis. Restoration in the levels of glutathione peroxidase along with ‘anti-lipoperoxidant effect’ was observed (). Similarly in an indomethacin-induced model of ulcers, rats pretreated with rutin caused reestablishment of altered oxidative stress and biochemical parameters possibly due to neutrophil infiltration, suppression of oxidative stress generation and replenishing nitrite/nitrate levels. Protective effects are also evident by histopathological investigations ().

Another investigation provides insight into the molecular mechanism of action of rutin over gastric proton pumps. Rutin demonstrated concentration-dependent inhibition of goat gastric ATPase, with IC50 = 36 μg/ml, making it that rutin exerts an antiulcer effect by inhibiting the gastric proton pump ().

2.6. Respiratory system

2.6.1. Antiasthmatic activity and other associated effects

The antiasthmatic activity of rutin was studied in ovalbumin-sensitized conscious guinea pigs challenged with aerosolized ovalbumin where airway resistance during the immediate phase response and late-phase response was determined. Rutin significantly inhibited specific airway resistance and immediate-phase response along with reticence of histamine, phospholipase A2, and eosinophil peroxidase. There was reduced conscription of neutrophils and eosinophils into the lung (). Use of rutin was also suggested in whooping cough along with vitamins C and K (). In cats and whippets, rutin has been effectively used in the management of idiopathic chylothorax ().

2.7. Bones

2.7.1. Antiosteoporotic and antiosteopenic effect

Osteoporosis is a skeletal disorder observed by a decrement in bone strength, that is associated with augmented risk of fracture (). This condition is affecting elderly citizens globally and seems to be budding health predicament. Osteoporosis is observed in the case when bone resorption by osteoclasts surpasses bone formation by osteoblasts (). All therapeutic strategies for treatment of osteoporosis emphasize inhibition of ‘osteoclast-mediated bone resorption’. Parathyroid hormones are only classes of agents that ‘stimulate bone formation’ (). In osteogenic-related assays, rutin caused proliferation and differentiation of human osteoblast-like MG-63 cells. There was also increase in the activity of alkaline phosphatase, expression of collagen type I and degree of mineralization (). Similar effects have been observed with rat calvarial osteoblast cells (). Rutin inhibits osteoclast formation by decreasing oxygen reactive species and TNF-alpha by inhibiting activation of NF-kappaB (). Rutin inhibits ovariectomy-induced osteopenia in rats by slowing down resorption and increasing osteoblastic activity (). Thus, rutin can also be regarded as ‘osteoblast stimulant’.

2.8. Eye

2.8.1. Anticataract and ophthalmic effect

Formation of advanced glycation end products (AGE) is associated with cataract, a diabetic complication. Thus, inhibition of such glycation could be a useful strategy to prevent such complication. In this study, rutin caused embarrassment of glycation of proteins, chelation of metal complexes and partly inhibiting post-Amadori formation (). In another study in wistar rat pups, selenium was used to induce cataract and protective effect of rutin in retarding cataractogenesis was investigated. Eye lens of rats treated with rutin were observed with restoration of lenticular antioxidant enzymes along with decrement in malondialdehyde formation. Experimental outcomes suggest that rutin prevented cataractogenesis possibly due to antioxidant mechanism ().

Oral administration of rutin has been observed with an improved reduction in intraocular pressure (). Involvement of vitamins B1, B2, forskolin, and rutin demonstrates ‘defending effect’ on the ocular surface which aids to re-establish a ‘normal equilibrium’ of the tear film altered due to toxins ().

2.9. Excretory system

2.9.1. Diuretic effect

Quercetin, a metabolite of rutin, which is abundantly found in Hibiscus sabdariffaLinn acted on vascular endothelium causing nitric oxide release, leading to increasing renal vasorelaxation by increasing kidney filtration ().

2.10. Reproductive system

2.10.1. Effect on sperm quality and male reproductive organs

Rutin, in a study, afforded protective effect on damage to human sperm induced by lipid peroxidation (). Rutin also demonstrated possible protection to testicular tissue and reproduction from oxidative stress observed in type 1 diabetes mellitus () along with amelioration of cyclophosphamide-induced reproductive toxicity () and testicular ischemia–reperfusion-induced oxidative stress in rats ().

2.11. Anticancer effects

Cancer includes a group of diseases that are characterized by the abnormal growth of cells that latently assault or spread to other parts of the body (). Flavonoids are known to demonstrate an extensive assortment of biological effects, comprising of ‘antioxidant and radical-scavenging activities’. Reactive oxygen species have been associated with the pathogenesis of several diseases such as atherosclerosis and certain cancers.

Rutin has been extensively studied for anticancer/antineoplastic effects. In a study, human leukemia HL-60 cells were implanted in a murine model, and rutin (dose 120 mg/kg) caused a significant reduction in tumor size justifying antileukemic potential (). In an independent study, rutin when administered to SW480 tumor cell lines (human colon cancer cell lines), was observed with less detrimental effects on the body and relative organ weight in mice along with an increment of mean survival time of 50 days (). Chem et al., demonstrated anti-neuroblastoma effect of rutin, where, rutin significantly inhibited the growth of LAN-5 cells and chemotactic ability. The study demonstrated that rutin could decrease BCL2 expression and BCL2/BAX ratio along with a reduction in levels of MYCN mRNA level and the secretion of TNF-α (). Rutin is also known to inhibit cancer cell growth by cell cycle arrest and/or apoptosis, along with inhibition of proliferation, angiogenesis, and/or metastasis in colorectal cell lines (). Rutin analog, quercetin, is studied against the proliferation of the ovarian cancer cell line OVCA 433, where dose-related inhibition was observed (). In another study, quercetin augmented apoptosis and barred metastasis in a model of pancreatic cancer (). Rutin also seems to be useful as an adjuvant in radioiodine therapy ().

2.12. Chemotherapeutic activity

2.12.1. Antibacterial activity

Rutin is extensively studied for antimicrobial activity against various strains of bacteria. It has demonstrated a profound degree of inhibition on growth of bacteria Escherichia coli (). Rutin, quantified in honey has shown inhibitory effects over Proteus vulgarisShigella sonnei and Klebsiella sp. (). Antimicrobial activity against Pseudomonas auruginosssa and Bacillus subtilis has also been documented (). In situ antimicrobial activity of rutin and other polyphenols in the food system has been studied, and the results demonstrate a promising involvement of flavonoids in the preservation of food ().

Bernard et al., demonstrated that rutin by inhibiting DNA isomerase IV demonstrated antibacterial activity against Ecoli (). In a study, rutin synergistically enhanced antibacterial activity of other flavonoids against Bacillus cereus and Salmonella enteritidis. Minimum inhibitory concentration value for kaempferol was remarkably decreased by the addition of rutin ().

2.12.2. Antifungal activities

Rutin demonstrated antifungal activity against the strain of Candida gattiiwith a minimum inhibitory concentration of 60 μg/ml (). It was suggested that chemical modification in rutin by the introduction of substitute group may alter physicochemical properties such as electron density, hydrophobicity and steric strain that may prove to be fruitful regarding increment of antifungal activity. Use of rutin in treatment of septic arthritis caused by C. albicans has also been suggested ().

2.12.3. Antimycobacterial activity

In a study, flavonoids rich extract containing rutin demonstrated antimycobacterial activity against Mycobacterium smegmatis ().

2.12.4. Larvicidal activity

In a study, rutin significantly inhibited growth and propagation of larvae of S. aegypti where maximum mortality of larvae was observed at 72 h ().

2.12.5. Antimalarial activity

Antimalarial effect of rutin in association with chloroquine was observed on white leghorn chickens infected with Plasmodium (Bennettinia) juxtanucleare (). In another study, biochemical mechanism of the antimalarial activity of Azadirachta indica leaf extract revealed the presence of an abundant amount of quercetin, an active metabolite of rutin (). Recently, the antiplasmodial activity of quercetin is documented against Plasmodium falciparum().

2.12.6. Antiretroviral activity

In a study, sodium rutin sulfate, a sulfated rutin analog, was investigated for anti-HIV activity against HIV-1 X 4 viruses IIIB, HIV-1 R5 isolates Ada-M and Ba-L strains. The mechanism by which sodium rutin sulfate demonstrated antiretroviral effect was a blockade of viral entry and virus-cell fusion which was mediated via interaction with HIV-1 envelope glycoprotein ().

2.12.7. Antiviral activity

Antiviral agents for the treatment of infections caused by retroviruses, orthomyxoviruses, herpes viruses, hepatitis B virus and hepatitis C virus are widely available (). Owing to the high incidence of viral infections along with no precise treatment for ‘appearance of new resistant viral strains’, its seems to be necessary to develop novel antiviral drugs.

Rutin has been tested against vesicular stomatitis virus on mouse fibroblasts and protected cells for about 24 h (). In the case of canine distemper virus infection, rutin affords immense viral embarrassment when added at the times of adsorption and penetration in the viral replicative cycle (). Rutin a chief constituent of Capparis sinaica Veill demonstrated profound antiviral effect against avian influenza strain H5N1 using plaque inhibition assay in the Madin-Darby canine kidney ().

2.13. Hair

Apoptosis of hair follicular cells is a leading cause of hair follicle degeneration. In a study, apoptosis of human follicular dermal papilla cells was observed after treatment with staurosporine; this event was completely introverted by exposure to rutin, spermidine, and zeaxanthin. There was the preservation of expression of anti-apoptotic molecules such as Bcl-2, MAP-kinases and their phosphorylated forms, which concludes the use of rutin in the prevention of apoptosis of hair follicular cells, one of the leading causes of baldness ().

2.14. Skin

2.14.1. Sunscreen effects

The desire for having healthy, beautiful and disease free skin is being observed from the eternal era. History of cosmetics and skin care product dates back in ancient Greek, Roman and Egyptian civilization. Advances in sciences and technology along with the development of modern chemistry have made this dream easy (). Exposure to ultraviolet B radiations produces ‘oxidative and inflammatory skin damage’ and augments jeopardy of skin ‘carcinogenesis. Rutin has been studied to explore the effects of rutin on UVB-induced inflammation in mouse skin in vivo. Topical application of rutin on mice skin 30 min before UVB irradiation reduced epidermal hyperplasia and the levels of proteins. There was also significant inhibition of UVB-induced expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) which could be due to inhibition of p38 MAP kinase and JNK that caused diminution UVB-induced expression of COX-2 in mouse skin (). In a study conducted by , rutin when incorporated in oil-in-water emulsions (concentration 10% w/w), sun protection factor (SPF) values comparable to ‘homosalate’ were observed. Along with TiO2, SPF values approached to 30.

2.14.2. In atopic dermatitis

In an atopic dermatitis model in BALB/c mice by repeated local exposure of house dust mite (Dermatophagoides farinae) extract (DFE) and 2,4-dinitrochlorobenzene (DNCB) to the ears, rutin treatment caused suppression of interleukin (IL)-4, IL-5, IL-13, IL-31, IL-32 and interferon (INF)-γ in the tissue. In 2,4-dinitroflourobenzene-sensitized a local lymph node assay for allergic contact dermatitis, rutin caused suppression of allergic contact dermatitis based on ear thickness and lymphocyte proliferation, serum IgG2a levels, and expression of INF-γ, IL-4, IL-5, IL-10, IL-17 and tumor necrosis factor-α in allergic contact dermatitis ears. This study signifies anti-atopic dermatitis and allergic contact dermatitis activity of rutin (). Rutin has also demonstrated anti aging effects on the skin (). Thus, rutin could be regarded as most effective topically applied compound given its antioxidant activity and skin penetration profile ().

2.15. Immune effects

Vibrio alginolyticus challenge was studied based on immune and physiological response to rutin in white shrimp. In this study, physiological, innate non-specific immune responses, respiratory bursts and superoxide dismutase activity against the pathogen were analyzed. Rutin from Toona sinensis was administered to shrimp. In brief, shrimps treated with rutin maintained lower glucose, lactate, and lipid levels in response against the pathogen. Survival rates of shrimps treated with rutin were more (). Another study demonstrates promotion of the activity of macrophage phagocytosis in cells ().

2.16. Body strength

2.16.1. Anti fatigue activity

Fatigue is an indication which is associated with person’s health. Fatigue is related to deterioration in ‘physical performance’. Extreme exercise is associated with accretion of surplus reactive free radicals that leads to an oxidative stress injury to the body. Depletion of energy along with the accumulation of excess metabolite is the key reasons for fatigue (). Rutin administration in mice prevented depletion of superoxide dismutase and reduced glutathione. After seven days of treatment with rutin, mice were sacrificed and analyzed for soleus muscle and brain for peroxisome proliferator-activated receptor-α coactivator and sirtuin one mRNA expression. Up-regulation of the CB1 cannabinoid receptor-interacting protein 1, myelin basic protein, Rho GDP dissociation inhibitor (GDI) alpha, and TPI indicated that rutin treatment inhibited condition anxiety via up-regulation of the expression of anxiety-associated proteins () and hence in nutshell it can be regarded as treatment with rutin improves the various mutilations allied with physical fatigue.

2.17. Organ protective effects

2.17.1. Neuroprotective activity

In mice model, rutin inhibited oxaliplatin-induced chronic painful peripheral neuropathy. Oxaliplatin is one of the most important platinum compounds used in colorectal cancer chemotherapy. However, it suffers from a drawback of peripheral which seems to be difficult to treat. Rutin significantly decreased oxaliplatin-induced peroxidative changes in the spinal cord and lipid peroxidation along with inducible nitric oxide ().

2.17.2. Retinoprotective activity

Effect of rutin on ocular blood flow by ‘colored microsphere technique’ was determined. Electroretinography was used to determine the b-wave recovery which is a tool for estimation of retinal function recovery. Rutin increased ocular blood flow and demonstrated a remarkable effect on retinal function recovery ().

2.17.3. Protective effect on lung tissue

Acute lung injury is severe disease observed with ‘high mortality and morbidity.’ Till date, there is no therapeutic stratagem established. Rutin demonstrated protective effects on histopathological changes in lung tissue along with prevention of ‘infiltration of polymorphonuclear granulocytes in bronchoalveolar lavage fluid’. There was also a reduction in secretion of lipid peroxidation and proinflammatory cytokines. The activity of antioxidant enzymes such as catalase, glutathione peroxidase, superoxide dismutase, and heme oxygenase-1 caused by lipopolysaccharide was reversed by rutin (). Pretreatment of rutin caused inhibition of lipopolysaccharide-induced arterial blood gas exchange and neutrophils infiltration in the lungs. There was suppression of macrophage inflammatory protein-2 and matrix metalloproteinase-9 (). Another study demonstrated that rutin effectively inhibited vascular cell adhesion molecule-1 and inducible nitric oxide synthase (). There is also evidence that rutin prevented early adult respiratory distress syndrome possibly due to inhibition of lipid peroxidation ().

2.17.4. Cardioprotective effects

Ischemic heart disease is one of the primary reasons for ‘morbidity and mortality’ all along the globe. The pathophysiology underlying damage at a cellular level raised due to ischemia is multifaceted. ‘Reactive oxygen species’ are identified to participate and play a key task in the pathogenesis of several diseases (). It is well known that ischemic tissues produce ‘oxygen-derived free radicals’ that are known to cause oxidative damage to membrane lipids, proteins, and carbohydrates that predispose to ‘qualitative and quantitative alterations’ of the myocardium (). Rutin was studied for cardioprotective effect by ischemia–reperfusion-induced myocardial infarction in normal and streptozotocin diabetic rats. Rutin administration significantly offered cardioprotection and restricted infracts size in normal and diabetic rats (). The mechanism lying behind this protection could be attenuation of lipid peroxidation in myocardial tissues ().

Isoproterenol is β-adrenergic agonist used to treat bradycardia and heart block (). However, frequent administration of it may lead to cardiac toxicity observed by an increase in the activity of serum cardiac marker enzymes viz lactate dehydrogenase, aspartate transaminase, creatine kinase, and alanine transaminase along with significant decrease in the activity of these enzymes in the heart. Rutin restored the levels of cardiac marker enzymes along with a reduction in lipid peroxidation. The study suggests the cardioprotective effect is due to the virtue of the antioxidant effect of rutin ().

Repression of isoproterenol-induced increase in angiotensin II and aldosterone in plasma was observed after administered by rutin. Along with this, overexpression of transforming growth factor β1, connective tissue growth factor, and excessive deposition of extracellular matrix in isoproterenol-treated myocardial tissues was observed in the group of animals treated with rutin ().

2.17.5. Prevention of splenocyte apoptosis

Rutin is known to alter the viability and function of mitogen-stimulated splenocytes and thymocytes compared with non-stimulated cells. In a study, when splenocytes were cultured with mitogens, it was observed that there was a decrease in interferon-gamma production along with decrement in splenocyte apoptosis ().

2.17.6. Hepatoprotective activity

Cirrhosis is the destruction of hepatocytes and following substitution with ‘scar tissue’ which alter the flow of blood through liver leading to the death of hepatocytes and n function of the liver (). The event of liver damage is followed by ‘hepatic fibrosis’ whereby regeneration of apoptotic cells is observed after ‘repeated injury’ (). In the event, when the death of regenerating cells occurs, deposition of extracellular matrix is observed, which leads to deposition of ‘fibrillar collagen’ (). Such pathology is difficult to treat. The effectiveness of treatment with colchicines, interferons, penicillamine and corticosteroids are contradictory and seem to be questionable. As oxidative stress is one of the causes beneath cirrhosis, use of antioxidants could be a unique strategy for treatment. Rutin is extensively studied for hepatoprotective activity in experimental animals.  evaluated the protective effect of rutin in carbon tetrachloride (CCl₄)-induced liver injuries in rats. Administration of rutin caused a decrement in levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and gamma-glutamyl transpeptidase in serum raised due to carbon tetrachloride. The level of cholesterol in blood was regulated whereas the level of endogenous liver antioxidant enzymes such as catalase, superoxide dismutase, glutathione peroxidase, glutathione-S-transferase, glutathione reductase and glutathione was increased, whereas lipid peroxidation was decreased in a dose-dependent manner. Along with this, the activity of p53 and CYP2E1 was restored by treatment with rutin. In an independent study in BALB/c mice, rutin treatment allayed oxaliplatin-induced hepatotoxicity and neurotoxicity. Prevention of mechanical allodynia along with histopathological observations suggested the protective role of rutin in the prevention of hepatotoxicity ().

2.17.7. Nephroprotective activity

Nephropathy is one of the most encountered pathogenesis observed in the populace all over the world regardless of age, caste, creed, race and environment. The reasons behind this condition result from anomalies rose due to different metabolic and physiological disturbances. Nephropathy is among the ten primary causes of death all along the globe (). Gentamicin (), ecstasy (), cisplatin (), and carbon tetrachloride () are some of the compounds known to demonstrate nephrotoxicity.

In a study, rutin demonstrated protective activity against oxonate-induced hyperuricemia and renal dysfunction in mice. Administration of rutin resulted in a the decrement in levels of serum urate, creatinine and blood urea nitrogen, serum and kidney uromodulin levels, and increased urine uromodulin, urate and creatinine excretion in hyperuricemic mice. There was a significant downregulation of mRNA and protein levels of mouse glucose transporter 9 and urate transporter 1, along with upregulation of mRNA and protein levels of organic anion transporter 1 and organic cation/carnitine transporters in the kidney of hyperuricemic mice. In a fructose-fed rat model for hyperuricemia, rutin blocked the NOD-like receptor three inflammasome activation and aided in the improvement in the signaling and reduced lipid accumulation in the kidney of rats () along with reversal of dysregulation of renal transporters (). Inhibition of inducible nitric oxide synthase activity and reduction in 3-nitrotyrosine formation () along with inhibition of reactive oxygen species () in the kidneys seem to be an important approach to prevent renal ischemia/reperfusion injury.

In cisplatin-treated wistar rats, rutin pretreatment caused restoration of kidney function and oxidative stress biomarkers (). It partially inhibited effect on NFκB and TNF-α pathway mediated inflammation, caspase-3 mediated-tubular cell apoptosis, along with re-establishment of histopathological changes due to cisplatin administration (). Rutin also demonstrated protective effects against potassium bromate-induced nephrotoxicity in rats. The decrement in DNA fragmentation, upregulation in the activity of antioxidant enzymes viz. catalase, superoxide dismutase, glutathione peroxidase, glutathione-S-transferase, glutathione reductase, and reduced glutathione along with a reduction in lipid peroxidation were observed ().

2.17.8. Protective effect on blood vasculature

Rutin was found to resist damages in vascular beds due to a disturbance in myocardium supply leading to initial damage, however in a chronic model of ‘stenocardia in dogs,’ therapeutic effect of rutin is up to a limited extent ().

2.18. Protective effects on wounds

2.18.1. Wound healing activity

Rutin, formulated as hydrogel, when applied to skin lesions of rats, caused a decrease in wound area as compared to control hydrogels. There was a reduction in oxidative stress in wound area as signified by a reduction in lipid peroxidation and protein carbonyl content along with increased catalase activity (). Rutin-releasing chitosan hydrogels as injectable dressings for dermal wound healing have been formulated. These hydrogels promoted defined formation of neo-epithelium and thicker granulation, which is closer to the original epithelial tissue ().

2.19. Radio modulatory effects

Radiation therapy is one of the principal strategies in anticancer therapy for treatment of various cancers; however, due to ill effects on normal tissues, its use is restricted. Ionizing radiations predispose to the generation of free radicals that damages DNA; cell death follows this event. Thus along with cancerous cells, the damage is observed with normal cells also. Some ‘radioprotectors’ have been screened; still, their clinical effectiveness is restricted due to toxicity with the frequent administration. In a study, rutin has been investigated for potential radioprotective effects. Rutin was administered to Swiss albino mice to assess its impact on ‘radiation-induced sickness along with the survival analyses’. Rutin at the dose of 10 mg/kg optimally demonstrated radioprotective effects. There was an increment in radiation tolerance and levels of antioxidant enzymes were restored along with a decrease in lipid peroxidation in liver (). Similar protection by rutin against gamma radiations was also observed ().

2.21. Drug interaction

Absorption of paclitaxel is increased when concurrently administered with rutin (). Rutin treatment cause increased the body’s resistance to drugs due to induction of the drug metabolizing enzymes in the liver; such an action has about the same magnitude with that of phenobarbital and pregnenolone-16 alpha-carbonitrile ().

2.22. Safety pharmacology

Rutin has been studied for carcinogenicity and results have demonstrated practically and significantly no carcinogenic potential in non-inbred golden hamsters ().

2.23. Structure–activity relationship

  • • 4-Oxo group and the 2,3 double bond in the C ring in rutin may be related by its neuroprotective action ().
  • • The presence of sugar moiety (O-rha-glu) has no significant effect on the ileum, which indicates that presence of sugar substitution diminishes the biological activity of the flavonoids ().
  • • Hydroxyl group in 5-position as well as the double bond and the double-bonded oxygen in the oxane ring is all necessary for the overall ability of flavonoids to inhibit phospholipase A2 activity, and that the hydroxyl groups in 3’- and 4’-position are required for selective inhibition of phospholipase A2-II ().
  • • Polyhydroxylated substitutions on rings A and B, a 2,3-double bond, a free 3-hydroxyl substitution and a 4-keto moiety, confer antiperoxidative properties to rutin ().
  • • Catechol group in the B-ring, a double bond between C2 and C3 at the C-ring, and ketone group in C4 at the C-ring are necessary for inhibitory effect on angiotensin-converting enzyme activity ().
  • • Rutin, whose C-3 position is glycosylated has a less free radical neutralizing activity that is aglycone quercetin ().
  • • The glycosylation decreases the AChE inhibitory activities of flavonoids and lowers the affinities for AChE by 1–5 times depending on the conjunction site and the type of sugar moiety ().
  • • The presence of a phenolic group in rutin is known to add hydrogen donation for scavenging the radiation-induced radicals, and to inhibit radiation-induced oxidative stress ().
  • • The presence of a glycosylation group in rutin greatly reduces vascular relaxation effect in porcine coronary artery ().

3. Conclusion

As evident from aforesaid facts, rutin is phytochemical with multiple pharmacological activities. An ancient saying ‘an apple a day, keeps doctor away’ seems to be true as rutin, one of the important constituents of apples, has a wide array of biological activities. Hence, rutin can be regarded as a ‘vital phytochemical’ which is needed to be studied extensively to establish effective safety profile in human to get therapeutic benefits.

Footnotes

Peer review under responsibility of King Saud University.

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Properties and Therapeutic Application of Bromelain: A Review

~ Content Source

Abstract

Bromelain belongs to a group of protein digesting enzymes obtained commercially from the fruit or stem of pineapple. Fruit bromelain and stem bromelainare prepared differently and they contain different enzymatic composition. “Bromelain” refers usually to the “stem bromelain.” Bromelain is a mixture of different thiol endopeptidases and other components like phosphatase, glucosidase, peroxidase, cellulase, escharase, and several protease inhibitors. In vitro and in vivo studies demonstrate that bromelain exhibits various fibrinolytic, antiedematous, antithrombotic, and anti-inflammatory activities. Bromelain is considerably absorbable in the body without losing its proteolytic activity and without producing any major side effects. Bromelain accounts for many therapeutic benefits like the treatment of angina pectoris, bronchitis, sinusitis, surgical trauma, and thrombophlebitis, debridement of wounds, and enhanced absorption of drugs, particularly antibiotics. It also relieves osteoarthritis, diarrhea, and various cardiovascular disorders. Bromelain also possesses some anticancerous activities and promotes apoptotic cell death. This paper reviews the important properties and therapeutic applications of bromelain, along with the possible mode of action.

1. Introduction

Pineapple is the common name of Ananas comosus (syns. A. sativus, Ananassa sativa, Bromelia ananas, B. comosa). Pineapple is the leading edible member of the family Bromeliaceae, grown in several tropical and subtropical countries including Philippines, Thailand, Indonesia, Malaysia, Kenya, India, and China. It has been used as a medicinal plant in several native cultures [1] and these medicinal qualities of pineapple are attributed to bromelain (EC 3.4.22.32), which is a crude extract from pineapple that contains, among other compounds, various closely related proteinases, exhibiting various fibrinolytic, antiedematous, antithrombotic, and anti-inflammatory activities in vitro and in vivo. Bromelain has been chemically known since 1875 and is used as a phytomedical compound [2]. Bromelain concentration is high in pineapple stem, thus necessitating its extraction because, unlike the pineapple fruit which is normally used as food, the stem is a waste byproduct and thus inexpensive [3]. A wide range of therapeutic benefits have been claimed for bromelain, such as reversible inhibition of platelet aggregation, sinusitis, surgical traumas [4], thrombophlebitis, pyelonephriti angina pectoris, bronchitis [5], and enhanced absorption of drugs, particularly of antibiotics [67]. Several studies have been carried out indicating that bromelain has useful phytomedical application. However, these results are yet to be amalgamated and critically compared so as to make out whether bromelain will gain wide acceptance as a phytomedical supplement [8]. Bromelain acts on fibrinogen giving products that are similar, at least in effect, to those formed by plasmin [9]. Experiment in mice showed that antacids such as sodium bicarbonate preserve the proteolytic activity of bromelain in the gastrointestinal tract [10]. Bromelain is considered as a food supplement and is freely available to the general public in health food stores and pharmacies in the USA and Europe [11]. Existing evidence indicates that bromelain can be a promising candidate for the development of future oral enzyme therapies for oncology patients [12]. Bromelain can be absorbed in human intestines without degradation and without losing its biological activity [1213].

2. Biochemical Properties

The crude aqueous extract from stem and fruit of pineapple is known as bromelain. It is a mixture of different thiol endopeptidases and other components like phosphatases, glucosidase, peroxidases, cellulases, glycoproteins, carbohydrates, and several protease inhibitors [14]. Stem bromelain (EC.3.4.22.32) is different from fruit bromelain (EC.3.4.22.33) [15]. The enzymatic activities of bromelain comprise a wide spectrum with pH range of 5.5 to 8.0 [16]. Different protein fractions were obtained by mean of various “biochemical techniques as sodium dodecyl sulphate polyacrylamide gel electrophoresis” (SDS-PAGE), isoelectric focusing (IEF), and multicathodal-PAGE [1718]. Nowadays, bromelain is prepared from cooled pineapple juice by centrifugation, ultrafiltration, and lyophilization. The process yields a yellowish powder, the enzyme activity of which is determined with different substrates such as casein (FIP unit), gelatin (gelatin digestion units), or chromogenic tripeptides [7171920].

3. Absorption and Bioavailability

The body can absorb significant amount of bromelain; about 12 gm/day of bromelain can be consumed without any major side effects [13]. Bromelain is absorbed from the gastrointestinal tract in a functionally intact form; approximately 40% of labeled bromelain is absorbed from intestine in high molecular form [21]. In a study carried out by Castell et al. [13] bromelain was detected to retain its proteolytic activity in plasma and was also found linked with alpha 2-macroglobulin and alpha1-antichymotrypsin, the two antiproteinases of blood. In a recent study, it was demonstrated that 3.66 mg/mL of bromelain was stable in artificial stomach juice after 4 hrs of reaction and also 2.44 mg/mL of bromelain remained in artificial blood after 4 hrs of reaction [22].

4. Medicinal Uses

Clinical studies have shown that bromelain may help in the treatment of several disorders.

4.1. Effects of Bromelain on Cardiovascular and Circulation

Bromelain prevents or minimizes the severity of angina pectoris and transient ischemic attack (TIA). It is useful in the prevention and treatment of thrombophlebitis. It may also break down cholesterol plaques and exerts a potent fibrinolytic activity. A combination of bromelain and other nutrients protect against ischemia/reperfusion injury in skeletal muscle [23]. Cardiovascular diseases (CVDs) include disorders of the blood vessels and heart, coronary heart disease (heart attacks), cerebrovascular disease (stroke), raised blood pressure (hypertension), peripheral artery disease, rheumatic heart disease, heart failure, and congenital heart disease [24]. Stroke and heart disease are the main cause of death, about 65% of people with diabetes die from stroke or heart disease. Bromelain has been effective in the treatment of CVDs as it is an inhibitor of blood platelet aggregation, thus minimizing the risk of arterial thrombosis and embolism [25]. King et al. [26] reported that administration of medication use to control the symptoms of diabetes, hypertension, and hypercholesteromia increased by 121% from 1988–1994 to 2001–2006 (P < 0.05) and was greater for patients with fewer healthy lifestyle habits. Bromelain supplement could reduce any of risk factors that contribute to the development of cardiovascular disease. In a recent research, Bromelain was found to attenuate development of allergic airway disease (AAD), while altering CD4+ to CD8+T lymphocyte populations. From this reduction in AAD outcomes it was suggested that bromelain may have similar effects in the treatment of human asthma and hypersensitivity disorders [27]. In another study, carried out by Juhasz et al., Bromelain was proved to exhibit the ability of inducing cardioprotection against ischemia-reperfusion injury through Akt/Foxo pathway in rat myocardium [28].

4.2. Bromelain Relieves Osteoarthritis

Osteoarthritis is the most common form of arthritis in Western countries; in USA prevalence of osteoarthritis ranges from 3.2 to 33% dependent on the joint [29]. A combination of bromelain, trypsin, and rutin was compared to diclofenac in 103 patients with osteoarthritis of the knee. After six weeks, both treatments resulted in significant and similar reduction in the pain and inflammation [30]. Bromelain is a food supplement that may provide an alternative treatment to nonsteroidal anti-inflammatory drug (NSAIDs) [31]. It plays an important role in the pathogenesis of arthritis [32]. Bromelain has analgesic properties which are thought to be the result of its direct influence on pain mediators such as bradykinin [3334]. The earliest reported studies investigating bromelain were a series of case reports on 28 patients, with moderate or severe rheumatoid or osteoarthritis [35].

4.3. Effect of Bromelain on Immunogenicity

Bromelain has been recommended as an adjuvant therapeutic approach in the treatment of chronic inflammatory, malignant, and autoimmune diseases [36]. In vitro experiments have shown that Bromelain has the ability to modulate surface adhesion molecules on T cells, macrophages, and natural killer cells and also induce the secretion of IL-1β, IL-6, and tumour necrosis factor α (TNFα) by peripheral blood mononuclear cells (PBMCs) [3743]. Bromelain can block the Raf-1/extracellular-regulated-kinase- (ERK-) 2 pathways by inhibiting the T cell signal transduction [44]. Treatment of cells with bromelain decreases the activation of CD4 (+) T cells and reduce the expression of CD25 [45]. Moreover, there is evidence that oral therapy with bromelain produces certain analgesic and anti-inflammatory effects in patients with rheumatoid arthritis, which is one of the most common autoimmune diseases [46].

4.4. Effect of Bromelain on Blood Coagulation and Fibrinolysis

Bromelain influences blood coagulation by increasing the serum fibrinolytic ability and by inhibiting the synthesis of fibrin, a protein involved in blood clotting [47]. In rats, the reduction of serum fibrinogen level by bromelain is dose dependent. At a higher concentration of bromelain, both prothrombin time (PT) and activated partial thromboplastin time (APTT) are markedly prolonged [48]. In vitro and in vivostudies have suggested that bromelain is an effective fibrinolytic agent as it stimulates the conversion of plasminogen to plasmin, resulting in increased fibrinolysis by degrading fibrin [4950].

4.5. Effects of Bromelain on Diarrhea

Evidence has suggested that bromelain counteracts some of the effects of certain intestinal pathogens like Vibrio cholera and Escherichia coli, whose enterotoxin causes diarrhoea in animals. Bromelain appears to exhibit this effect by interacting with intestinal secretory signaling pathways, including adenosine 3′ : 5′-cyclic monophosphatase, guanosine 3′ : 5′-cyclic monophosphatase, and calcium-dependent signaling cascades [51]. Other studies suggest a different mechanism of action. In E. coli infection, an active supplementation with bromelain leads to some antiadhesion effects which prevent the bacteria from attaching to specific glycoprotein receptors located on the intestinal mucosa by proteolytically modifying the receptor attachment sites [5253].

4.6. Effect of Bromelain on Cancer Cells

Recent studies have shown that bromelain has the capacity to modify key pathways that support malignancy. Presumably, the anticancerous activity of bromelain is due to its direct impact on cancer cells and their microenvironment, as well as on the modulation of immune, inflammatory, and haemostatic systems [12]. Most of the in vitro and in vivo studies on anticancer activity of bromelain are concentrated on mouse and human cells, both cancerous and normal, treated with bromelain preparations. In an experiment conducted by Beez et al chemically induced mouse skin papillomas were treated with bromelain and they observed that it reduced tumor formation, tumor volume and caused apoptotic cell death [54]. In one study related to bromelain treatment of gastric carcinoma Kato III cell lines, significant reduction of cell growth was observed [55] while in another study bromelain reduced the invasive capacity of glioblastoma cells and reduced de novo protein synthesis [56]. Bromelain is found to increase the expression of p53 and Bax in mouse skin, the well-known activators of apoptosis [54]. Bromelain also decreases the activity of cell survival regulators such as Akt and Erk, thus promoting apoptotic cell death in tumours. Different studies have demonstrated the role of NF-κB, Cox-2, and PGE2 as promoters of cancer progression. Evidence shows that the signaling and overexpression of NF-κB plays an important part in many types of cancers [5758]. Cox-2, a multiple target gene of NF-κB, facilitates the conversion of arachidonic acid into PGE2 and thus promotes tumour angiogenesis and progression [5960]. It is considered that inhibiting NF-κB, Cox-2, and PGE2 activity has potential as a treatment of cancer. Bromelain was found to downregulate NF-κB and Cox-2 expression in mouse papillomas [54] and in models of skin tumourigenesis [61]. Bromelain was also shown to inhibit bacterial endotoxin (LPS)-induced NF-κB activity as well as the expression of PGE2 and Cox-2 in human monocytic leukemia and murine microglial cell lines [6263]. Bromelain markedly has in vivo antitumoural activity for the following cell lines: P-388 leukemia, sarcoma (S-37), Ehrlich ascetic tumour, Lewis lung carcinoma, and ADC-755 mammary adenocarcinoma. In these studies, intraperitoneal administration of bromelain after 24 hours of tumour cell inoculation resulted in tumour regression [54].

4.7. Role of Bromelain in Surgery

Administration of bromelain before a surgery can reduce the average number of days for complete disappearance of pain and postsurgery inflammation [6465]. Trials indicate that bromelain might be effective in reducing swelling, bruising, and pain in women having episiotomy [66]. Nowadays, bromelain is used for treating acute inflammation and sports injuries [31].

4.8. Role of Bromelain in Debridement Burns

The removal of damaged tissue from wounds or second/third degree burns is termed as debridement. Bromelain applied as a cream (35% bromelain in a lipid base) can be beneficial for debridement of necrotic tissue and acceleration of healing. Bromelain contains escharase which is responsible for this effect. Escharase is nonproteolytic and has no hydrolytic enzyme activity against normal protein substrate or various glycosaminoglycan substrates. Its activity varies greatly with different preparations [67]. In two different enzymatic debridement studies carried out in porcine model, using different bromelain-based agents, namely, Debriding Gel Dressing (DGD) and Debrase Gel Dressing showed rapid removal of the necrotic layer of the dermis with preservation of the unburned tissues [6869]. In another study on Chinese landrace pigs, enzymatic debridement using topical bromelain in incised wound tracks accelerated the recovery of blood perfusion, pO2in wound tissue, controlled the expression of TNF-α, and raised the expression of TGT-β [70]. Enzymatic debridement using bromelain is better than surgical debridement as surgical incision is painful, nonselective and exposes the patients to the risk of repeated anaesthesia and significant bleeding [7174].

4.9. Toxicity of Bromelain

According to Taussig et al. [75] bromelain has very low toxicity with an LD50(lethal doses) greater than 10 g/kg in mice, rates, and rabbits. Toxicity tests on dogs, with increasing level of bromelain up to 750 mg/kg administered daily, showed no toxic effects after six months. Dosages of 1500 mg/kg per day when administered to rats showed no carcinogenic or teratogenic effects and did not provoke any alteration in food intake, histology of heart, growth, spleen, kidney, or hematological parameters [76]. Eckert et al. [41] after giving bromelain (3000 FIP unit/day) to human over a period of ten days found no significant changes in blood coagulation parameters.

5. Conclusion

Bromelain has a wide range of therapeutic benefits, but the mode of its action is not properly understood. It is proved that bromelain is well absorbed in body after oral administration and it has no major side effects, even after prolonged use. All the evidences reviewed in this paper suggest that bromelain can be used as an effective health supplement to prevent cancer, diabetes, and various cardiovascular diseases in the long run.

6. Future Trends and Perspectives

Bromelain can be a promising candidate for the development of oral enzyme therapies for oncology patients. It is clear from this paper that bromelain is a multiaction enzyme; however, more research is required to understand the proper mechanism of action of bromelain so that the multiaction activities of bromelain can be harnessed efficiently.

Acknowledgments

The authors are grateful to DEAN, Department of Biotechnology, IBMER, Mangalayatan University, Aligarh, India, for providing necessary facilities and encouragement. They are also thankful to all faculty members of the Institute of Biomedical Education and Research, Mangalayatan University, Aligarh, India, for their generous help and suggestions during the paper preparation.

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Properties and Therapeutic Application of Bromelain(Pineapple):A Review

~ Content Source

Abstract

Bromelain belongs to a group of protein digesting enzymes obtained commercially from the fruit or stem of pineapple. Fruit bromelain and stem bromelainare prepared differently and they contain different enzymatic composition. “Bromelain” refers usually to the “stem bromelain.” Bromelain is a mixture of different thiol endopeptidases and other components like phosphatase, glucosidase, peroxidase, cellulase, escharase, and several protease inhibitors. In vitro and in vivo studies demonstrate that bromelain exhibits various fibrinolytic, antiedematous, antithrombotic, and anti-inflammatory activities. Bromelain is considerably absorbable in the body without losing its proteolytic activity and without producing any major side effects. Bromelain accounts for many therapeutic benefits like the treatment of angina pectoris, bronchitis, sinusitis, surgical trauma, and thrombophlebitis, debridement of wounds, and enhanced absorption of drugs, particularly antibiotics. It also relieves osteoarthritis, diarrhea, and various cardiovascular disorders. Bromelain also possesses some anticancerous activities and promotes apoptotic cell death. This paper reviews the important properties and therapeutic applications of bromelain, along with the possible mode of action.

1. Introduction

Pineapple is the common name of Ananas comosus (syns. A. sativus, Ananassa sativa, Bromelia ananas, B. comosa). Pineapple is the leading edible member of the family Bromeliaceae, grown in several tropical and subtropical countries including Philippines, Thailand, Indonesia, Malaysia, Kenya, India, and China. It has been used as a medicinal plant in several native cultures [] and these medicinal qualities of pineapple are attributed to bromelain (EC 3.4.22.32), which is a crude extract from pineapple that contains, among other compounds, various closely related proteinases, exhibiting various fibrinolytic, antiedematous, antithrombotic, and anti-inflammatory activities in vitro and in vivo. Bromelain has been chemically known since 1875 and is used as a phytomedical compound []. Bromelain concentration is high in pineapple stem, thus necessitating its extraction because, unlike the pineapple fruit which is normally used as food, the stem is a waste byproduct and thus inexpensive []. A wide range of therapeutic benefits have been claimed for bromelain, such as reversible inhibition of platelet aggregation, sinusitis, surgical traumas [], thrombophlebitis, pyelonephriti angina pectoris, bronchitis [], and enhanced absorption of drugs, particularly of antibiotics []. Several studies have been carried out indicating that bromelain has useful phytomedical application. However, these results are yet to be amalgamated and critically compared so as to make out whether bromelain will gain wide acceptance as a phytomedical supplement []. Bromelain acts on fibrinogen giving products that are similar, at least in effect, to those formed by plasmin []. Experiment in mice showed that antacids such as sodium bicarbonate preserve the proteolytic activity of bromelain in the gastrointestinal tract []. Bromelain is considered as a food supplement and is freely available to the general public in health food stores and pharmacies in the USA and Europe []. Existing evidence indicates that bromelain can be a promising candidate for the development of future oral enzyme therapies for oncology patients []. Bromelain can be absorbed in human intestines without degradation and without losing its biological activity [].

2. Biochemical Properties

The crude aqueous extract from stem and fruit of pineapple is known as bromelain. It is a mixture of different thiol endopeptidases and other components like phosphatases, glucosidase, peroxidases, cellulases, glycoproteins, carbohydrates, and several protease inhibitors []. Stem bromelain (EC.3.4.22.32) is different from fruit bromelain (EC.3.4.22.33) []. The enzymatic activities of bromelain comprise a wide spectrum with pH range of 5.5 to 8.0 []. Different protein fractions were obtained by mean of various “biochemical techniques as sodium dodecyl sulphate polyacrylamide gel electrophoresis” (SDS-PAGE), isoelectric focusing (IEF), and multicathodal-PAGE []. Nowadays, bromelain is prepared from cooled pineapple juice by centrifugation, ultrafiltration, and lyophilization. The process yields a yellowish powder, the enzyme activity of which is determined with different substrates such as casein (FIP unit), gelatin (gelatin digestion units), or chromogenic tripeptides [].

3. Absorption and Bioavailability

The body can absorb significant amount of bromelain; about 12 gm/day of bromelain can be consumed without any major side effects []. Bromelain is absorbed from the gastrointestinal tract in a functionally intact form; approximately 40% of labeled bromelain is absorbed from intestine in high molecular form []. In a study carried out by Castell et al. [] bromelain was detected to retain its proteolytic activity in plasma and was also found linked with alpha 2-macroglobulin and alpha1-antichymotrypsin, the two antiproteinases of blood. In a recent study, it was demonstrated that 3.66 mg/mL of bromelain was stable in artificial stomach juice after 4 hrs of reaction and also 2.44 mg/mL of bromelain remained in artificial blood after 4 hrs of reaction [].

4. Medicinal Uses

Clinical studies have shown that bromelain may help in the treatment of several disorders.

4.1. Effects of Bromelain on Cardiovascular and Circulation

Bromelain prevents or minimizes the severity of angina pectoris and transient ischemic attack (TIA). It is useful in the prevention and treatment of thrombophlebitis. It may also break down cholesterol plaques and exerts a potent fibrinolytic activity. A combination of bromelain and other nutrients protect against ischemia/reperfusion injury in skeletal muscle []. Cardiovascular diseases (CVDs) include disorders of the blood vessels and heart, coronary heart disease (heart attacks), cerebrovascular disease (stroke), raised blood pressure (hypertension), peripheral artery disease, rheumatic heart disease, heart failure, and congenital heart disease []. Stroke and heart disease are the main cause of death, about 65% of people with diabetes die from stroke or heart disease. Bromelain has been effective in the treatment of CVDs as it is an inhibitor of blood platelet aggregation, thus minimizing the risk of arterial thrombosis and embolism []. King et al. [] reported that administration of medication use to control the symptoms of diabetes, hypertension, and hypercholesteromia increased by 121% from 1988–1994 to 2001–2006 (P < 0.05) and was greater for patients with fewer healthy lifestyle habits. Bromelain supplement could reduce any of risk factors that contribute to the development of cardiovascular disease. In a recent research, Bromelain was found to attenuate development of allergic airway disease (AAD), while altering CD4+ to CD8+T lymphocyte populations. From this reduction in AAD outcomes it was suggested that bromelain may have similar effects in the treatment of human asthma and hypersensitivity disorders []. In another study, carried out by Juhasz et al., Bromelain was proved to exhibit the ability of inducing cardioprotection against ischemia-reperfusion injury through Akt/Foxo pathway in rat myocardium [].

4.2. Bromelain Relieves Osteoarthritis

Osteoarthritis is the most common form of arthritis in Western countries; in USA prevalence of osteoarthritis ranges from 3.2 to 33% dependent on the joint []. A combination of bromelain, trypsin, and rutin was compared to diclofenac in 103 patients with osteoarthritis of the knee. After six weeks, both treatments resulted in significant and similar reduction in the pain and inflammation []. Bromelain is a food supplement that may provide an alternative treatment to nonsteroidal anti-inflammatory drug (NSAIDs) []. It plays an important role in the pathogenesis of arthritis []. Bromelain has analgesic properties which are thought to be the result of its direct influence on pain mediators such as bradykinin []. The earliest reported studies investigating bromelain were a series of case reports on 28 patients, with moderate or severe rheumatoid or osteoarthritis [].

4.3. Effect of Bromelain on Immunogenicity

Bromelain has been recommended as an adjuvant therapeutic approach in the treatment of chronic inflammatory, malignant, and autoimmune diseases []. In vitro experiments have shown that Bromelain has the ability to modulate surface adhesion molecules on T cells, macrophages, and natural killer cells and also induce the secretion of IL-1β, IL-6, and tumour necrosis factor α (TNFα) by peripheral blood mononuclear cells (PBMCs) []. Bromelain can block the Raf-1/extracellular-regulated-kinase- (ERK-) 2 pathways by inhibiting the T cell signal transduction []. Treatment of cells with bromelain decreases the activation of CD4 (+) T cells and reduce the expression of CD25 []. Moreover, there is evidence that oral therapy with bromelain produces certain analgesic and anti-inflammatory effects in patients with rheumatoid arthritis, which is one of the most common autoimmune diseases [].

4.4. Effect of Bromelain on Blood Coagulation and Fibrinolysis

Bromelain influences blood coagulation by increasing the serum fibrinolytic ability and by inhibiting the synthesis of fibrin, a protein involved in blood clotting []. In rats, the reduction of serum fibrinogen level by bromelain is dose dependent. At a higher concentration of bromelain, both prothrombin time (PT) and activated partial thromboplastin time (APTT) are markedly prolonged []. In vitro and in vivostudies have suggested that bromelain is an effective fibrinolytic agent as it stimulates the conversion of plasminogen to plasmin, resulting in increased fibrinolysis by degrading fibrin [].

4.5. Effects of Bromelain on Diarrhea

Evidence has suggested that bromelain counteracts some of the effects of certain intestinal pathogens like Vibrio cholera and Escherichia coli, whose enterotoxin causes diarrhoea in animals. Bromelain appears to exhibit this effect by interacting with intestinal secretory signaling pathways, including adenosine 3′ : 5′-cyclic monophosphatase, guanosine 3′ : 5′-cyclic monophosphatase, and calcium-dependent signaling cascades []. Other studies suggest a different mechanism of action. In E. coli infection, an active supplementation with bromelain leads to some antiadhesion effects which prevent the bacteria from attaching to specific glycoprotein receptors located on the intestinal mucosa by proteolytically modifying the receptor attachment sites [].

4.6. Effect of Bromelain on Cancer Cells

Recent studies have shown that bromelain has the capacity to modify key pathways that support malignancy. Presumably, the anticancerous activity of bromelain is due to its direct impact on cancer cells and their microenvironment, as well as on the modulation of immune, inflammatory, and haemostatic systems []. Most of the in vitro and in vivo studies on anticancer activity of bromelain are concentrated on mouse and human cells, both cancerous and normal, treated with bromelain preparations. In an experiment conducted by Beez et al chemically induced mouse skin papillomas were treated with bromelain and they observed that it reduced tumor formation, tumor volume and caused apoptotic cell death []. In one study related to bromelain treatment of gastric carcinoma Kato III cell lines, significant reduction of cell growth was observed [] while in another study bromelain reduced the invasive capacity of glioblastoma cells and reduced de novo protein synthesis []. Bromelain is found to increase the expression of p53 and Bax in mouse skin, the well-known activators of apoptosis []. Bromelain also decreases the activity of cell survival regulators such as Akt and Erk, thus promoting apoptotic cell death in tumours. Different studies have demonstrated the role of NF-κB, Cox-2, and PGE2 as promoters of cancer progression. Evidence shows that the signaling and overexpression of NF-κB plays an important part in many types of cancers []. Cox-2, a multiple target gene of NF-κB, facilitates the conversion of arachidonic acid into PGE2 and thus promotes tumour angiogenesis and progression []. It is considered that inhibiting NF-κB, Cox-2, and PGE2 activity has potential as a treatment of cancer. Bromelain was found to downregulate NF-κB and Cox-2 expression in mouse papillomas [] and in models of skin tumourigenesis []. Bromelain was also shown to inhibit bacterial endotoxin (LPS)-induced NF-κB activity as well as the expression of PGE2 and Cox-2 in human monocytic leukemia and murine microglial cell lines []. Bromelain markedly has in vivo antitumoural activity for the following cell lines: P-388 leukemia, sarcoma (S-37), Ehrlich ascetic tumour, Lewis lung carcinoma, and ADC-755 mammary adenocarcinoma. In these studies, intraperitoneal administration of bromelain after 24 hours of tumour cell inoculation resulted in tumour regression [].

4.7. Role of Bromelain in Surgery

Administration of bromelain before a surgery can reduce the average number of days for complete disappearance of pain and postsurgery inflammation []. Trials indicate that bromelain might be effective in reducing swelling, bruising, and pain in women having episiotomy []. Nowadays, bromelain is used for treating acute inflammation and sports injuries [].

4.8. Role of Bromelain in Debridement Burns

The removal of damaged tissue from wounds or second/third degree burns is termed as debridement. Bromelain applied as a cream (35% bromelain in a lipid base) can be beneficial for debridement of necrotic tissue and acceleration of healing. Bromelain contains escharase which is responsible for this effect. Escharase is nonproteolytic and has no hydrolytic enzyme activity against normal protein substrate or various glycosaminoglycan substrates. Its activity varies greatly with different preparations []. In two different enzymatic debridement studies carried out in porcine model, using different bromelain-based agents, namely, Debriding Gel Dressing (DGD) and Debrase Gel Dressing showed rapid removal of the necrotic layer of the dermis with preservation of the unburned tissues []. In another study on Chinese landrace pigs, enzymatic debridement using topical bromelain in incised wound tracks accelerated the recovery of blood perfusion, pO2in wound tissue, controlled the expression of TNF-α, and raised the expression of TGT-β []. Enzymatic debridement using bromelain is better than surgical debridement as surgical incision is painful, nonselective and exposes the patients to the risk of repeated anaesthesia and significant bleeding [].

4.9. Toxicity of Bromelain

According to Taussig et al. [] bromelain has very low toxicity with an LD50(lethal doses) greater than 10 g/kg in mice, rates, and rabbits. Toxicity tests on dogs, with increasing level of bromelain up to 750 mg/kg administered daily, showed no toxic effects after six months. Dosages of 1500 mg/kg per day when administered to rats showed no carcinogenic or teratogenic effects and did not provoke any alteration in food intake, histology of heart, growth, spleen, kidney, or hematological parameters []. Eckert et al. [] after giving bromelain (3000 FIP unit/day) to human over a period of ten days found no significant changes in blood coagulation parameters.

5. Conclusion

Bromelain has a wide range of therapeutic benefits, but the mode of its action is not properly understood. It is proved that bromelain is well absorbed in body after oral administration and it has no major side effects, even after prolonged use. All the evidences reviewed in this paper suggest that bromelain can be used as an effective health supplement to prevent cancer, diabetes, and various cardiovascular diseases in the long run.

6. Future Trends and Perspectives

Bromelain can be a promising candidate for the development of oral enzyme therapies for oncology patients. It is clear from this paper that bromelain is a multiaction enzyme; however, more research is required to understand the proper mechanism of action of bromelain so that the multiaction activities of bromelain can be harnessed efficiently.

Acknowledgments

The authors are grateful to DEAN, Department of Biotechnology, IBMER, Mangalayatan University, Aligarh, India, for providing necessary facilities and encouragement. They are also thankful to all faculty members of the Institute of Biomedical Education and Research, Mangalayatan University, Aligarh, India, for their generous help and suggestions during the paper preparation.

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