Association of Peroxisomes, Reactive Oxygen Species (ROS) and Antioxidants: Insights from Preclinical and Clinical Evaluations

Abstract

Peroxisome function has long been associated with oxygen metabolism. High concentrations of hydrogen peroxide (H2O2−) producing oxidases are in the set of peroxisomes and their antioxidant enzymes, especially catalase. Reactive oxygen species (ROS) can certainly be considered as an intracellular multifunctional biological factor which are released and scevenged in peroxisomes. They are known to be involved in normal cellular functions such as signaling mediators, overproduction under oxidative stress conditions leading to adverse cellular effects, cell death, and various other pathological conditions. This review provides an insight into the relationship between peroxisomes and ROS, which are emerging as key players in the dynamic rotation of ROS metabolism and oxidative damage. Various conditions upset the balance between ROS production and removal in peroxisomes. The current review also targets the ROS-inhibiting enzymes and exemplifying the effects of antioxidants in pre-clinical and clinical evaluation of natural and herbal supplements.

1. Introduction

Reactive oxygen species (ROS) can certainly be considered as an intracellular multifunctional biological factor. They are known to be involved in normal cellular functions such as signaling mediators, overproduction under oxidative stress conditions leading to adverse cellular effects and eventually cell death. Under normal conditions in every human cell, the release of pro-oxidants in the form of ROS (reactive oxigen species) and RNS (reactive nitrogen species) are scrutinized by antioxidant levels. The equilibrium maintained is shifted in favor of pro-oxidants resulting in oxidative stress, when exposed to adverse circumstances such as atmospheric pollutants, unfavorable physicochemical, environmental or pathologicl agents including cigarette smoking, toxic chemicals, ultra violet rays and radiation and also excess formation of advanced glycation end products (AGE), in diabetes [1, 2]. This has been associated in the origin of various (>100) human diseases. Peroxisome function has long been associated with oxygen metabolism. High concentrations of H₂O₂⁻ producing oxidases are in the set of peroxisomes and their antioxidant enzymes, especially catalase. This review provides an insight into the relationship between ROS and peroxisomes, which have emerged as key players in the dynamic rotation of ROS metabolism and oxidative damage. The counterparts of ROS-producing enzymes such as antioxidants are also discussed; exemplifying their effects in pre-clinical and clinical evaluation of natural and herbal supplements [3].

Peroxisomes contain at least 50 different enzymes involved in a variety of biochemical pathways in different cell types. Peroxisomes were originally defined as organelles that perform oxidation reactions that result in the production of hydrogen peroxide. Because hydrogen peroxide is harmful to the cell, peroxisomes also contain the enzyme catalase, which breaks down hydrogen peroxide either by converting it to water or by oxidizing another organic compound. A variety of substrates are degraded by such oxidative reactions in peroxisomes, including uric acid, amino acids, and fatty acids. The oxidation of fatty acids is a particularly important example as it is a major source of metabolic energy. In animal cells, fatty acids are oxidized in both peroxisomes and mitochondria, but in yeast and plants, fatty acid oxidation is restricted to peroxisomes. Peroxisomes are one of the main sites in the cell where oxygen free radicals are both generated and scavenged. The balance between these two processes is believed to be of great importance for the proper functioning of cells and has been linked to aging and carcinogenesis. The peroxisome is a single, membrane-bound organelle present in virtually every eukaryotic cell and biosynthetic pathways, however, these pathways may differ between species. The significance of the peroxisome for the normal regulation of cellular activities can be explained by the fact that more than 20 human peroxisomal disorders dwell because of the absence of protein or due to loss of protein function [4].

Antioxidants act either by neutralizing free radicals or their consequences [5]. The natural cellular environment provides sufficient protective pathways against unfavorable effects of free radicals: glutathione reductase, glutathione peroxidase, superoxide dismutase (SOD), disulphide bonding, thiols and thioredoxin are buffering systems in every cell. The relation between free radicals and disease can be explained by the concept of “Oxidative stress” [3].

All biological molecules present in our body are at risk of being attacked by free radicals. Such damaged molecules can impair the cell functions and even lead to cell death eventually resulting in disease states. Antioxidants may prevent and improve different diseased states [1, 2]. Several investigators have demonstrated the positive effects of antioxidants like Vitamin C, E to name a few in both preclinical and clinical setup.

2. Oxidative stress as a marker of endothelial dysfunction

The vascular endothelium, which promotes the passage of macromolecules and circulating cells from blood to tissues, is an important target of oxidative stress, playing a pivotal role in the pathophysiology of several vascular diseases and disorders. It is has been also reported that exclusively, oxidative stress increases vascular endothelial permeability and promotes leukocyte adhesion, which is coupled with alterations in endothelial signal transduction and redox-regulated transcription factors [6]. The reactive oxygen species (ROS) which originate at the sites of inflammation and injury have been emerged as the major contributing factor in the pathogenesis of endothelial dysfunction. These reactive oxygen species at low concentrations can function as signaling molecules participating in the regulation of fundamental cell activities such as cell growth and cell adaptation responses; whereas at higher concentrations, ROS can cause cellular injury and death. Predominantly, under normal body functions, the vital enzymes such as NAD(P)H dependent oxidases and superoxide dismutases (SOD) conscientiously regulate release of superoxide along with maintenance of intracellular redox balance. On the other hand, if production of superoxide anion surpasses the scavenging capacity of endothelial cells, this active intermediate react with nitric oxide resulting in formation of peroxynitrites. These peroxynitrites are potent oxidants that cause structural and functional changes in various components of the cell. ROS are also well known to suppress NO and restrict the formation of peroxynitrite [7]. It is a cytotoxic oxidant that causes endotheial dysfunction through nitration of protein function. Peroxynitrite plays a key role in oxidation of LDL and as proatherogenic [8]. In addition, peroxynitrite leads to the degradation of the eNOS cofactor tetrahydrobiopterin (BH4), resulting in an uncoupling of eNOS [9]. An excess of oxidant also leads to a reduction of BH4 with an increase in BH2. When this occurs, the formation of the active dimer of eNOS with oxygenase activity and the production of NO is restricted. The reductase function of eNOS is activated and more ROS are produced, so NO synthase switches from its NO-producing oxygenase function to its ROS-producing reductase function, with consequent exaggeration of oxidant excess and deleterious effects on endothelial and vascular function of the vessel wall. ROS upregulate adhesion (VCAM-1 and ICAM-1) and chemotactic molecules (macrophage chemoattractant peptide-1 (MCP-1) [8]. Inflammation decreases the bioavailability of NO [8]. The main source of oxidative excess in the vasculature is NAD(P)H oxidase and xanthine oxidase [10], the mitochondria [11] and uncoupled NOS constitute as other sources.

3. Significance of antioxidants

Many investigators have studied the significance of antioxidants in relation to disease and showed that zinc is an essential trace element, being a cofactor for about 200 human enzymes, including cytosolic antioxidant Cu-Zn SOD, isoenzyme of SOD mainly present in cytosol. Selenium is also an essential trace element and a cofactor for glutathione peroxidase. There is a vast information which suggests that chronic administration of antioxidants may be beneficial in improving cardiovascular risk. Vitamin E and tocotrienols (such as those from palm oil) are efficient lipid soluble antioxidants that function as a chain breaker during lipid peroxidation in cell memebranes and various lipid particles including LDL [12].

Vitamin E is considered the standard antioxidant against which other compounds with antioxidant activity are compared, particularly in terms of its biological activity and clinical relevance. Daily dietary intake varies between 400 IU and 800 IU. Vitamin C is also another important water-soluble free radical scavenger. The recommended daily dose is 60 mg. Apart from these, carotenoids like beta-carotene, lycopene, lutein and other carotenoids act as important antioxidants and quench superoxide (O2.) and (ROO..) [12, 13].

The effects of short-term dietary supplementation of tomato juice (source of lycopene), vitamin E and vitamin C on susceptibility of LDL to oxidation and circulating levels of C-reactive protein (CRP) and cell adhesion molecules measured in patients with type 2 diabetes. In this study 57 patients with well controlled type 2 diabetes melliitus were randomized to receive tomato juice (500 mg/day), Vitamin E (800 U/day), Vitamin C (500 mg/day) or placebo treatment for 4 weeks. It was observed that lycopene and vitamin E were both associated with resistance of LDL to oxidation, but only Vitamin E showed a decrease in C-reactive protein. It was also found that levels of cell adhesion molecules and plasma glucose did not change significantly during the study. Thses investigators then suggested that these findings may be relevant to strategies aimed at reducing risk of myocardial infarction in patients with diabetes [14].

In another study, it was suggested that vitamin E (1,600 IU/day, 10 weeks) decreased the susceptibility of LDL to oxidation in comparison with placebo. Vitamin E had this effect in both bouyant and dense LDL subfractions. This protection occurred in an environment where glycemic indices did not change and protein glycation was unaffected. The hypothesis that endothelial function and LDL oxidation might be linked was advanced further by Pinkney et al. [15]. These investigators studied 46 patients with type 1 diabetes without nephropathy and compared the results to 39 controls using a 3-month, randomized, double-blind, placebo-controlled study of vitamin E, 500 IU/day. The results indicated that in the absence of changes in LDL oxidation, vitamin E intake enhanced flow mediated dilatation FMD in type I diabetics [16].

Another [17] study reported that intracellular Vitamin C levels are reduced in patients with type 1 diabetes, particularly those who are poorly controlled. Histologically, the microvascular lesions of scurvy bear a surprising resemblance to those seen in long-standing diabetes, making them an attractive therapeutic alternative. Many short-term studies have shown the beneficial effects of ascorbic acid on vascular function, particularly in smokers and after ingestion of high-fat meals. However, the effect of vitamin C is not chronically sustainable, at least in smokers. Based on these observations, hypothesized that the antioxidant vitamin C might enhance endothelium-dependent vasodilation in forearm resistance when tested [18]. These investigators studied 10 subjects with diabetes and 10 age-matched, nondiabetic control subjects. FBF was determined by venous occlusion plethysmography, and endothelium dependent vasodilatation. The results from this study indeed support the hypothesis that acute administration of vitamin C improves endothelial function associated with the diabetic state., however, no information on chronic effects can be found from this study. But still more research has to be taken up in exploring the possible use of these vitamins to prevent atherosclerosis and/or microvascular disease in patients with diabetes. A study showed for the first time effects of consumption of flavonoid rich dark chocolate on endothelial function, aortic stiffness, wave reflections and oxidant status in healthy adults [19].

According to world health organization, traditional medicines are widely used globally. Approximately 80% of the population of developing countries rely on traditional medicines for their primary health care needs [20, 21, 22]. These medicinal plants contain several phytochemicals such as Vitamins (A, C,E and K), carotenoids, terpenoids, flavonoids, polyphenols, alkaloids, tannins, saponins, enzymes and minerals etc. These phytochemicals possess antioxidant activities, which can be used in the treatment of multiple ailments [23]. Many herbs along with potent antioxidant activity also possess anti-inflammatory and cardioprotective properties and are used by patients with increased risk of cardiovascular morbidity and mortality. Thus it is necessary to through light on the beneficial effects of the herbs such as Terminalia arjuna, Emblica officinalis, Withania somnifera, Boerhaavia diffusa and Ocimum sanctum etc.

4. Preclinical studies with Terminalia arjuna

4.1 Antioxidant and anticancer activities

The effect of Terminalia arjuna aqueous extract on the antioxidant defense system in lymphoma-bearing AKR mice was examined. The antioxidant effects of T. arjuna were monitored through the activities of catalase, superoxide dismutase and glutathione S-transferase. These enzyme activities are low in lymphoma-bearing mice, indicating an impaired antioxidant defense system. Oral administration of different doses of aqueous extracts of T. arjuna caused a significant increase in the activities of antioxidant enzymes. Here, T. arjuna was found to downregulate anaerobic metabolism by inhibiting lactate dehydrogenase activity in lymphoma-bearing mice, which was increased in untreated cancerous mice. The results demonstrated the antioxidant effects of Terminalia arjuna aqueous extract, which may play a role in anticarcinogenic activity by reducing oxidative stress [24].

4.1.1 Cardio protective activity

Sumitra et al., demonstrated that Arjunolic acid, a new triterpene and a potent principle from the bark of Terminalia arjuna has been shown to produce significant cardiac protection in isoproterenol induced myocardial necrosis in rats and prevents decrease in the levels of super oxide dismutase, catalase and reduced glutathione. This study explains that Arjunolic acid at a dosage of 15 mg/kg body weight (Pre and post treatment) produces cardioprotective effect [25].

4.1.2 Antiplatelet activity

Some researchers have shown that oleanane-type triterpene glycosides designated as Termiarjunoside I and Termiarjunoside II isolated from stem bark of Terminalia arjuna, potently suppressed the release of nitric oxide and superoxide from isolated macrophages and also inhibited aggregation of platelets [26].

5. Clinical studies with Terminalia arjuna

5.1 Antioxidant and Cardioprotective activity

The antioxidant constituents in Terminalia arjuna are reported to reverse endothelial dysfunction in chronic smokers. The study was conducted with 18 healthy male smokers and an equal number of non-smokers of the same age. The baseline brachial artery reactivity test was done using high-frequency ultrasound according to the standard protocol under identical conditions to determine endothelium-dependent flow-mediated and endothelium-independent nitroglycerin-mediated dilatation. The two groups were matched for age, body mass index, blood pressure, serum cholesterol, mean resting vessel diameter, and flow velocities after occlusion. Smokers then received Terminalia arjuna (500 mg every 8 hours) or a matched placebo randomly in a double-blind, cross-over design for two weeks each, followed by repeated brachial artery reactivity studies to determine various parameters, including flow-mediated dilatation after each period. However, flow-mediated dilatation showed a significant improvement from baseline after Terminalia arjuna therapy. The study concluded that smokers have impaired endothelium-dependent but normal endothelium-independent vasodilation as determined by brachial artery reactivity studies. In addition, two weeks of Terminalia arjuna therapy resulted in significant regression of this endothelial abnormality in smokers [27].

The effect of Terminalia arjuna (500 mg 8 hourly) was evaluated in fifty-eight males with chronic stable angina (NYHA class II-III) and with isosorbide mononitrate (40 mg/daily) on treadmill exercise induced ischemia, or a matching placebo for one week each. A wash-out period of at least three days was observed between the groups in a randomized, double-blind, crossover design. The treadmill exercise test parameters improved significantly during therapy with both treatments compared to those with placebo [28].

5.1.1 E. officinalis

The fruits of E. officinalis (Amla) family: Euphorbiaceae, commonly known as Indian gooseberry is widely used in many of the indigenous medical preparations against a variety of disease conditions [29].

E. officinalis is considered as a rich source of a vitamin C, which plays an important role in scavenging free radicals. For many years the therapeutic potential of fruits of E. officinalis was attributed to their high content of ascorbic acid [30]. It was further determined through comprehensive, chromatographic, spectroscopic and crucial chemical analyzes that the antioxidant property is due to the low molecular weight hydrolyzable tannins of fresh fruit skin. These tannins, namely Emblicanin A, Emblicanin B, Pedunculagin and Punigluconin, have been found to provide protection against oxygen radical-induced hemolysis of rat peripheral erythrocytes [31]. Purification and fractionation process was conducted in another study and phytochemicals like gallic acid, methyl gallate and geranin were identfied [32].

6. Preclinical studies with E. officinalis

6.1 Antioxidant activity

Invitro and animal studies have shown that Amla has potent antioxidant activity against multiple test systems such as superoxide radicals, induction of lipid peroxide formation by the Fe⁺³/ADP ascorbate system, hydroxyl radical scavenging activity. It also caused systemic increase in antioxidant enzymes in laboratory animals [33].

6.2 Hypolipidemic activity

In a study conducted in rats showed that flavonoids from E. officinalis effectively reduced lipid levels in serum and tissues and had significant inhibitory effect on hepatic 3-hydroxy-3-methylglutaryl-CoenzymeA (HMG CoA) reductase activity [34].

Effect of amla on the lipid metabolism and protein expression involved in oxidative stress during the aging process were evaluated in laboratory rats. Sun Amla or ethyl acetate extract of amla, a polyphenol-rich fraction, on oral administration significantly increased the hepatic PPAR [α] protein level. Furthermore, the amla extracts reduced the expressions of hepatic NF-[kappa] B, inducible NO synthase (iNOS), and cyclo-oxygenase-2 (COX-2) protein levels which were increased with aging. The results suggested that amla may prevent age-related hyperlipidaemia through attenuating oxidative stress in the aging process [35].

E. officinalis (Amla), showed improvement in treatment of dyslipidemia and intima-media thickening and plaque formation in the aorta in hypercholesterolemic rabbits [36].

7. Clinical studies with E. officinalis

7.1 Hypolipidemic and anti-inflammatory activity

In a pilot clinical study the effect of E. officinalis extract (AMLAMAX ™) was evaluated on markers of systemic inflammation and dyslipidemia. Amlamax™ a purified, standardized, dried extract of amla containing about 35% galloellagic tannins along with other hydrolysable tannins showed reduction in total and LDL cholesterols, in blood CRP levels and enhancement of beneficial HDL cholesterol [37].

7.2 Hypoglycaemic activity

The hypoglycemic and lipid lowering effects of E. officinalis fruits were evaluated in normal and diabetic patients. The data showed a significant decrease (p < 0.05) in fasting and 2 hour post- prandial blood glucose levels along with total cholesterols (TC) and triglycerides (TG) in both normal and diabetic volunteers upon 21 days of treatment [38].

7.3 W. somnifera

W. somnifera (ashwagandha, WS) Family: Solanacae is widely used in Ayurvedic medicine, the traditional medical system of India and is an important medicinal plant, which is used in to cure many diseases. Some researchers have demonstrated that W. somnifera possesses powerful antioxidants. Preclinical studies also suggested the herb to produce an increase in the levels of natural antioxidants- superoxide dismutase, catalase and glutathione peroxidase [39].

8. Preclinical studies with W. somnifera

8.1 Anti-inflammatory and Antistress activities

Anti-inflammatory properties have been investigated to validate its use in inflammatory arthritis and animal stress studies have been performed to investigate its use as an antistress agent [40, 41].

8.2 Hypoglycaemic and Hypolipidemic activities

In a study flavonoids were isolated from the extracts of W. somnifera root and leaf and further hypoglycaemic and hypolipidemic effects were investigated in alloxan-induced diabetic rats. Eight weeks of treatment with W. somnifera and glibenclamide restored the changes in parameters to normal, indicating that it possesses hypoglycaemic and hypolipidemic activities [42].

8.3 Anti-oxidant activity

Researchers at Banaras Hindu University in Varanasi have discovered that some of the chemicals found in W. somnifera are powerful antioxidants. Studies conducted on rat brains showed that the herb increased the levels of superoxide dismutase, catalase, and glutathione peroxidase [39].

8.4 Anti-carcinogenic activity

The anti-carcinogenic property of Ashwagandga has been confirmed. Animal cell cultures has shown that the herb lowers [43] tumor size [44]. In another study, the herb was examined for its antitumor effects on urethane-induced lung tumors in adult male mice. After administration of ashwagandha for a period of seven months, the histological study of the lungs was similar to that observed in the lungs of control animals [45, 46].

9. Clinical studies with W. somnifera

9.1 Hypoglycemic and hypolipidemic activity

The hypoglycemic, hypocholesterolemic and diuretic effects of Ashwagandha has been studied in human clinical trials. A decrease in blood sugar levels comparable to that caused by the administration of a hypoglycaemic drug has been observed. Significant increases in urinary sodium, urine volume, and decreases in serum cholesterol, triglycerides, and low-density lipoproteins were also observed [46].

9.2 O. sanctum

O. sanctum also known as Tulsi belonging to family: Labiatae and its extracts are used in ayurvedic remedies. The use of this herb has been reported in the Indian traditional medical system, and its modern uses receive widespread attention over the years. Various parts of the plant have been claimed to be valuable in a wide range of diseases. It has been observed that Tulsi exerts hypocholesterolemic, hypotriglyceridemic and hypophospholipidemic effects. Among the chemical constituents contained in essential oil of O. sanctum leaves eugenol, a phenolic compound, is considered to be an active ingredient contributing for its hypolipidemic and antioxidant action [47].

9.3 Antioxidant and antineoplastic activity

In a study at Bangladesh, the antioxidant activity of Tulsi leaves extract was evaluated invitro. O. sanctum extract showed significant free radical scavenging activity. In the same study, antineoplastic activity of O. sanctum was demonstrated against Ehrlich Ascites Carcinoma (EAC) in mice. Tulsi leaves extract was administered at a dose of 50mgKg⁻¹ body weight intraperitoneally. Heamatological studies reveal that hemoglobin levels were reduced in EAC-treated mice, while near-normal recovery was observed in extract-treated animals. There was also a significant decrease in RBC count and an increase in WBC count in extract-treated mice compared to EAC-treated animals. From the results it was concluded that the extract has significant antioxidant and antineoplastic activity [48].

9.4 Hypolipidemic activity

In a study administration of fresh leaves of O. sanctum for four weeks resulted in significant changes in the lipid profile of normal albino rabbits. Significant reduction in serum total cholesterol, triglyceride, phospholipid and LDL cholesterol levels and an increase in stool HDL cholesterol and total sterol content were recorded [49].

Hypolipidemic activity of shade dried leaf powder of Tulsi along with the extracts and their fractions have shown invitro hypolipidemic and anti-peroxidative activity at very low concentrations in male albino rabbits. Aqueous extract feeding also provided significant protection of liver and aortic tissue from hypercholesterolemia-induced peroxidative damage [50].

10. Protection against radiation induced lipid peroxidation

A study was conducted to see if aqueous extract of O. sanctum, protects against radiation induced lipid peroxidation in liver and to determine the role, if any, of the inherent antioxidant system in producing radioprotection. Glutathione (GSH) and the antioxidant enzymes glutathione S-transferase (GST), reductase (GSRx), peroxidase (GSPx) and superoxide dismutase (SOD), as well as lipid peroxide (LPx) activity were estimated in the liver of adult swiss mice. The mice were injected intraperitoneally with 10 mg/kg of Tulsi for 5 consecutive days and exposed to 4.5 Gy of gamma radiation 30 min after the last injection. The aqueous extract itself increased GSH and enzymes significantly above normal levels, while irradiation significantly reduced all levels. The maximum drop was 30–60 min for GSH and related enzymes and 2 h for SOD. Pretreatment with the extract controlled the radiation-induced depletion of GSH and all enzymes and kept their levels within or above the control range. Irradiation significantly increased the lipid peroxidation rate, reaching a maximum value (about 3.5 times that of control) 2 hours after exposure. Aqueous extract pretreatment significantly reduced lipid peroxidation and accelerated recovery to normal levels [51].

11. Clinical studies with O. sanctum

In a study Tulsi leaves were tested on anthropometric measurements, diabetic symptoms and blood pressure in male patients with non-insulin dependent diabetes mellitus. Daily dosage of four capsules i.e. 2 g powder (Lunch and dinner) was given and supplementation was carryout for a period of 3 months. Significant percent reduction in the symptoms like polydypsia (35%), polyphagia (21%), and headache (27%), was observed in patients treated with Tulsi. It was concluded from the study that tulsi leaves are helpful in reducing subjects’ diabetic symptoms and blood pressure. No significant improvement in subjects’ anthropometric parameters was observed tulsi leaf powder supplementation [52].

11.1 Boerhaavia diffusa

Boerhaavia diffusa is a medicinal plant widely used in Ayurvedic medicine. The plant was named in the honor of Hermann Boerhaave, a famous Dutch physician of the 18th century [53]. It is also known as Spreading Hogweed in English, belonging to family, Nyctaginaceae.

12. Preclinical studies with Boerhaavia diffusa

12.1 Hypolipidemic activity

The efficacy of antioxidant and hypolipidemic agents tocotrienols and Boerhaavia diffusa by analyzing all the parameters in plasma lipoprotein lipids, total lipids (TL), total cholesterol (TC), triglycerides (TG), VLDL-C, LDL-C, HDL-C and MDA in oxidized cholesterol feeded rats. In the same study invitro oxidizability of LDL, was also demonstrated. All the plasma lipid parameters and MDA levels were significantly increased in hyperlipidemic control rats. After 4 weeks of administration of tocotrienols and Boerhaavia diffusa significantly reduced the overall oxidative burden and effectively ameliorated the above altered parameters. Thus indicating a strong hypolipidemic/antiatherogenic and antioxidant effect [54].

12.2 Anti-oxidant activity

The root extracts of Boerhaavia diffusa were evaluated using different solvents for free radical scavenging activity (FRSA) at a dose of 1000 mg/Kg body weight, prior to irradiation with 8 Gy gamma radiation as compared to mice pre-treated with extract at the dose of 250 and 500 mg/Kg body weight prior to irradiation with same dose of radiation. The data obtained showed that hydroethanolic extract produced potent free radical scavenging activity in DPPH^., ABTS^.+ and NO^. assays and was found to be beneficial in reducing symptoms of radiation sickness, changes in body weight and mortality were minimum in the experimental animals. The antioxidant effect of B. diffusa roots was attributed to the presence of certain phenolic constituents like quercetin, caeffic acid, kempferol and their derivatives [55].

12.3 Hypoglycemic activity

Another study focused on blood glucose concentration and hepatic enzymes in normal and alloxan induced diabetic rats after daily oral administration of aqueous solution of Boerhaavia diffusa L. leaf extract (200 mg/kg) for 4 weeks. Significant improvement was recorded in blood glucose and glycosylated hemoglobin A1C levels. The action of hepatic enzymes such as hexokinase was significantly increased. Similarly, glucose-6-phosphatase, fructose-1,6-bisphosphatase were significantly decreased by the administration of BLEt in normal and diabetic rats. The results of BLEt were more potent when compared with antidiabetic drug—glibenclamide (600 μg/kg) [56].

12.4 Hepatoprotective activity

The hepato-protective activity of Boerhaavia diffusa alcoholic extract of the whole plant administered orally, was evaluated against experimentally induced hepatotoxicity using carbon tetrachloride in rats and mice. The extract also produced an increase in normal bile flow, indicating potent choleretic activity and no signs of toxicity were observed up to an oral dose of 2 g/kg.

13. Clinical studies with Boerhaavia diffusa

13.1 Antioxidant and Hypolipidemic activity

The effects of methanolic extract of Boerhaavia diffusa on oxidative stress in healthy and diabetes mellitus patients were studied. Results through this research demonstrated that diabetic patients experience increased oxidative stress when compared with normal subjects, significant increase in plasma, TG, TC, VLDL-C, LDL-C, and decrease in HDL-C. This may be due to markedly increased production of oxidant and significantly diminished antioxidant defense including a decline in total plasma antioxidant power. The study depicted that daily intake of Boerhaavia diffusa extract by diabetes mellitus patients significantly reduced TC, TG, LDL-C and increased HDL-C levels. The study concluded that extract of Boerhaavia diffusa may be useful in the prevention and treatment of the diabetes-induced hyperlipidemia and atherosclerosis. In addition, daily use of Boerhaavia diffusa can be efficacious and cost effective and good source of natural antioxidant [57].

14. Discussion

Over the past three decades, various experimental startegies have revealed the existence of cellular functions of peroxisomes related to reactive oxygen species (ROS) and reactive nitrogen species (RNS), and the function of peroxisomes as key centers of the cellular signaling apparatus. Peroxisomes of different origins have been detected which strongly indicate the interest of them as a cellular source of various signaling molecules, including ROS. In this review, we have focused on the generation and regulation of ROS in peroxisomes and the different antioxidant systems in this cell organelle [58]. We also enlighten the supporting evidence for application of antioxidants in preclinical and clinical evaluation of herbal supplements used in the management of associated disease complications.

Uncontrolled ROS production leads to structural modification of cellular proteins and alteration of their functions, resulting in cellular dysfunction and dysregulation of important cellular processes [59, 60]. Enhanced levels of ROS cause lipid, protein, and DNA damage. Specifically ROS can distort the lipid membrane and increase the fluidity and permeability of the membrane. Impairment of protein includes site-specific amino acid modification, peptide chain fragmentation, aggregation of crosslinked reaction products, modification of electric charges, immobilization of enzymes, and sensitivity to proteolysis [61]. Eventually, ROS can damage DNA by oxidizing deoxyribose, strand breakage, removal of nucleotides, changes in bases, and crosslinking DNA protein [62, 63, 64, 65].

Literature suggests that peroxisomes are powerful and metabolically active organelles and are a very vital source of reactive oxygen species (ROS), H2O2, O2 (.-) and · OH, which are the products of diverse metabolic pathways, such as, photorespiration, fatty acid β-oxidation, nucleic acid and polyamine catabolism, ureide metabolism, to name a few. ROS were originally associated with oxygen toxicity. However, these reactive species also play a significant role in the signaling network that regulates essential processes in the cell. Peroxisomes have the ability to produce and scavenge H2O2 and O2(.-) rapidly, allowing to regulate dynamic alterations in ROS levels. The flexibility of these organelles, and based on varied developmental and environmental stimuli, render these organelles to perform a pivotal role in cellular signal transduction. The catalase and glycolate oxidase loss-of-function mutants have provided insights to study the consequences of modifications in endogenous H2O2 levels in peroxisomes. This has also facilitated the understanding of transcriptomic profile of genes regulated by peroxisomal ROS. It is now well established that peroxisomal ROS are involved in complicated signals which employ hormones, calcium, and redox homeostasis [66].

Antioxidants render an important role in these defense mechanisms. The antioxidat therapies target for maintenance of critical balance between oxidants and proxidants. In aerobic organisms, the steady release of free radicals needs to be equalized at the same degree of utilization of antioxidant. The naturally occuring enzymatic or non-enzymatic antioxidant systems prevent the formation of free radicals, and neutralize or repair the damage caused by them [62]. A wide range of endogenous and exogenous antioxidants are responsible, for providing protection against oxidative damage leading to development of chronic diseases [67]. The different types of antioxidant systems present both in plants [68] and the human body, contributes for controlling ROS homeostasis [69]. Release of natural ROS by the mitochondrial respiratory chain suggests that under certain conditions ROS can be metabolically beneficial but at the same time may also be harmful to cells [70, 71, 72].

The plant kingdom has served the mankind since ancient time and has provided remedies for various disease conditions. Over the period of time as the knowledge of plant derived medicines got advanced, it opened new avenues in improving the health and quality of life. Since many centuries, herbal drugs have been used both as food supplements and for medicinal requirements. When we mention about herbal medicine, it constitutes all parts of the plant like seeds, roots, bark, leaves, flowers and fruits from trees [73]. Most of the plant derived products and herbs act as potent scavengers of ROS or possess antioxidant activity. The phytoconstituents present in these herbs have been evaluated in numerous studies and are proved to rapidly stimulate the natural antioxidant enzyme systems such as catalase, superoxide dismutase, reduced glutathione etc., which protect the cells from oxidative damage and from progression of chronic diseases [74].

15. Conclusion

The conclusion of the present review is that peroxisomes are most common type of single layered membrane organelles identified in different types of eukaryotic cells. The origin of peroxisomes are through growth and division of cell and are independent organelles. These are also recognized as one of the most important and strong multifunctional organelles. The peroxisomes are able to facilitate the dynamic rotation of ROS generation and removal, fatty acid oxidation, β-oxidation of long-chain fatty acids, decomposition of purines, and glycerol, ether lipid and bile acid biosynthesis [75]. The metabolic processes of peroxisomes those which take place together with mitochondrial involvement are fatty acid β-oxidation and amino acid metabolism, but whereas the oxidation of different substrates is promoted by oxidases that consume oxygen. Several investigators in their work indicate the ability of peroxisomes to utilize 20% of the total oxygen consumption and can release up to 35% of the cellular hydrogen peroxide due to which are known to be major contributors of oxidative metabolism and in conserving oxidation balance [76]. Evidences from research show that the regulation of cell proliferation, apoptosis and carbohydrate metabolism is governed by hydrogen peroxide that act as a vital signaling molecule. However, at increased levels hydrogen peroxide is toxic and requires a regular check for its concentration. The other vital function of peroxisomes includes the action of antioxidant enzyme systems like the CAT, SOD, PRDX1, and PRDX5 [77, 78, 79, 80]. CAT being the most significant enzyme and other antioxidant enzymes which metabolize the peroxidase hydrogen produced as a byproduct of peroxidases. Additionally super oxide dismutase 1 (SOD1) is regarded as a perfect peroxisomal protein, and a potent antioxidant enzyme that quenches the superoxide and accelerates the modification of oxygen to superoxide anion (O2⁻) [81].

Plant-based bioactive molecules have received a lot of recognition since the past few decades. Several studies have demonstrated their therapeutic significance in the management of disease conditions and for prevention as well. The complete phytochemical profile of whole plants, plant extracts or even the isolated constituents are well explained in the literature, which can be utilized for planning treatment strategies for various diseases including diabetes mellitus, cardiovascular disease and neurodegenerative disorders. Extensive randomized trials are warranted to collect data for establishing the medical interest or probable hazards of antioxidant supplementation.

There is enormous substantiation that oxidative stress has been implicated in normal physiological processes and environmental interactions that occur in a cell. Several mechanisms are involved in antioxidant defense systems that render protection against oxidative damage. Literature suggests that in many conditions these processes seem to be tangled. ROS profusely disrupts the antioxidant balance, causing oxidative stress and results in constant alterations in the cellular material, which includes carbohydrate, protein and lipid substances [70, 82, 83, 84]. It is likely to presume that oxidative stress can be a cause of tissue damage and finally arresting the natural cellular-signaling processes. However, a thorough understanding of the biochemical events occurring at a cellular level to influence oxidative damage is mandatory to direct ensuing progress. Peroxisomes serve as very important sites for detoxification of ROS. But however peroxisomes itself release these radicals. With the advent of fluorescence methods and having vast knowledge of peroxisomal functions, we expect to read more about the role of this organelle in the near future that can be useful in the treatment of related disorders.

Acknowledgments

I thank Al Hawash Private University, Homs, Arab Republic of Syria, for providing the necessary support in completion of this chapter.