Catalase is a heme protein, localized in the peroxisomes or the microperoxisomes. This enzyme catalyses the decomposition of H₂O₂ to water and oxygen and thus protecting the cell from oxidative damage by H₂O₂ and OH^—. Glutathione peroxidase is a seleno–enzyme two third of which (in liver) is present in the cytosol and one third in the mitochondria. It catalyses the reaction of hydroperoxides with reduced glutathione to form glutathione disulphide and the reduction product of the hydroperoxide. Vitamin C, vitamin E, selenium, β–carotene, lycopene, lutein and other carotenoids have been used as supplementary antioxidants. Apart from these, plant secondary metabolites such as flavonoids and terpenoids play an important role in the defense against free radicals^7,-9.

Medicinal plant parts (roots, leaves, branches/stems, barks, flowers, and fruits) are commonly rich in phenolic compounds, such as flavonoids, phenolic acids, stilbenes, tannins, coumarins, lignans and lignins^{10, 11, 12}. They have multiple biological effects including antioxidant activity¹³. The antioxidant properties of phenolic acids and flavonoids are due to their redox properties, ability to chelate metals and quenching of singlet oxygen¹⁴. Flavonoids, which are partly responsible for the pigmentation of flowers, fruits and leaves, are subdivided into flavanols, flavonols, flavones, flavanones and anthocyanins based on the saturation of the flavan ring and also their hydroxylation. They occur mostly as glycosylated derivatives, sometimes conjugated with sulphate or organic acids¹⁵.

TRADITIONAL USES OF PSIDIUM GUAJAVA:

In India numerous medicinal plants are used for the treatment of diseases. One of the plants used traditionally is Psidium guajava L. (guava). It belongs to the family of Myrtaceae. Psidium guajava is a common shade tree or shrub in door–yard gardens in the tropics. Fruit is enjoyed as a sweet treat by indigenous people all over the world and also the leaves and barks have a long history of medicinal use.

The pharmacological actions and the medicinal uses of aqueous extracts of Psidium guajava leaves in folk medicine include the treatment of various types of gastrointestinal disturbances such as vomiting, diarrhea, inhibition of the peristaltic reflex, gastroenteritis, spasmolytic activity, dysentery, abdominal distention, flatulence and gastric pain ¹⁶. These extracts have also been indicated to cause disturbances of the central nervous system: insomnia, convulsions and epilepsy ¹⁷. Bronchitis, asthma attacks, cough, pulmonary diseases could be also treated with guava teas ¹⁸ and could also be useful as anti–inflammatory and hemostatic agent ¹⁹. Moreover, aqueous extracts of Psidium guajava leaves were described to be effective against a number of microbial strains ²⁰ and anti–rotavirus activity ²¹.

PHYTOCHEMISTRY OF PSIDIUM GUAJAVA:

It has been reported that phytochemical analysis of Psidium guajava leaf products revealed the presence of more than 20 related compounds including alkaloids, anthocyanins, carotenoids, essential oils, lectins, phenols, saponins, tannins, triterpenes and vitamin C^22-28.

The essential oil contains alpha pinene, caryophyllene, cineol, D–limonene, eugenol, and myrcene. The major constituents of the volatile acids include (E)–cinnamic acid and (Z)–3–hexenoic acid. Carbohydrate esters have been isolated from the fruit^23,²⁶. The main active constituent in the plant is quercetin. Spasmolytic and antidiarrheal effects are associated with its quercetin–derived flavonoid glycosides, which support use of this ancient leaf remedy in treating gastrointestinal disorders²⁹.

ANTIOXIDANT ACTIVITY OF PSIDIUM GUAJAVA:

Psidium guajava is a tropical fruit, widely consumed fresh and also processed (beverages, syrup, ice cream, and jams). Jimenez–Escrig et al.³⁰ evaluated the pulp and peel fractions, and both showed high content of dietary fiber and extractable polyphenols. The antioxidant activity of polyphenol compounds was studied, by free radical 2, 2–diphenyl–1–picrylhydryzyl (DPPH) scavenging, ferric reducing/antioxidant power (FRAP) assay, and inhibition of copper–catalyzed in vitro human low–density lipoprotein (LDL) oxidation. All fractions tested showed a remarkable antioxidant capacity, and this activity was correlated with the corresponding total phenolic content. A 1–g (dry matter) portion of peel contained DPPH activity, FRAP activity, and inhibition of copper–induced in vitro LDL oxidation, equivalent to 43 mg, 116 mg, and 176 mg of Trolox, respectively. These results indicate that Psidium guajava could be a suitable source of natural antioxidants. Peel and pulp could also be used to obtain antioxidant dietary fiber (AODF), a new item which combines in a single natural product the properties of dietary fiber and antioxidant compounds.

Yamashiro et al. ³¹ determined whether the medicinal herbs growing in Okinawa and possessing a radical–scavenging activity would exert cardioprotective effects against ischemia–reperfusion injury using isolated perfused rat hearts. Effects of the aqueous extracts from Psidium guajava and Limonium wrightii at concentrations having an equipotent radical–scavenging activity on myocardial injury produced by global ischemia followed by reperfusion were tested and were further compared with those of quercetin and gallic acid, major antioxidative components of Psidium guajava and Limonium wrightii, respectively. Both extracts significantly attenuated ischemic contracture during ischemia and improved myocardial dysfunction after reperfusion. Decreases in high–energy phosphates and increases in malondialdehyde in the reperfused hearts were significantly lessened with both plant extracts. Quercetin and gallic acid also exerted similar beneficial effects. These results indicate that Psidium guajava and Limonium wrightii both have cardioprotective effects against myocardial ischemia–reperfusion injury in isolated rat hearts, primarily through their radical–scavenging actions.

Qian and Nihorimbere ³² studied the dried ground leaves of Psidium guajava extracted, and the total phenolic content present in the extracts by spectrophotometrically according to Folin–Ciocalteu's phenol method and calculated as gallic acid equivalent. Remarkably high total phenolic content 575.3 ± 15.5 and 511.6 ± 6.2 mg of gallic acid equivalent/g of dried weight material (for ethanol Psidium guajava leaf extracts and water Psidium guajava leaf extracts, respectively) were obtained. The antioxidant activity of lyophilized extracts was determined at ambient temperature by means of a DPPH colorimetry with detection scheme at 515 nm. The activity was evaluated by the decrease in absorbance as the result of DPPH color change from purple to yellow. The higher the sample concentration used, the stronger was the free radical–scavenging effect. Their results showed that ascorbic acid was a substantially more powerful antioxidant than the extracts from Psidium guajava leaf. On the other hand, the commercial Psidium guajava leaf extracts and ethanol Psidium guajava leaf extracts showed almost the same antioxidant power whereas water Psidium guajava leaf extracts showed lower antioxidant activity. They conclude that Psidium guajava leaf extracts comprise effective potential source of natural antioxidants.

Thaipong et al. ³³ evaluated the hydrophilic antioxidant activity (AOAH) and the lipophilic antioxidant activity (AOAL); and their correlations with vitamin C, and total phenolic and beta–carotene contents in fresh Psidium guajava fruits of one white flesh clone and three pink flesh clones. FRAP assay was used to estimate both AOAH and AOAL from methanol and dichloromethane extracts, respectively. The white flesh clone, showed higher levels of both AOAH than the pink flesh clones. The AOAH was positively correlated with vitamin C and total phenolic but was negatively correlated with beta–carotene. The AOAL was not correlated with these antioxidants.

Suganya et al. ³⁴ investigated the ethanolic extracts from 24 samples plant species commonly found in Thailand and compared on their antioxidant activity by (2, 2’–azino–bis 3–ethylbenzthiazoline–6–sulphonic acid) or ABTS assay. The ethanol extract from the leaves of Psidium guajava showed the highest antioxidant capacity with the Trolox Equivalent Antioxidant Capacity (TEAC) value of 4.908 ± 0.050 mM/mg, followed by the fruit peels of rambutan (Nephelium lappaceum) and mangosteen (Garcinia mangostana) with the TEAC values of 3.074 ± 0.003 and 3.001 ± 0.016 mM/mg, respectively. The further investigation of Psidium guajava leaf extracts from different solvents; n–hexane, ethyl acetate, n–butanol, and methanol, was examined using ABTS and FRAP assays. The total phenolic content was done by Folin–Ciocalteu reaction. The results indicated that the methanol fraction possessed the highest antioxidant activity, followed by the butanol and ethyl acetate fractions, respectively. The hexane fraction showed the lowest antioxidant activity. Their results demonstrated that the mechanism of antioxidant action of Psidium guajava leaf extracts was free radical scavenging and reducing of oxidized intermediates. The phenolic content in Psidium guajava leaf fraction played a significant role on the antioxidant activity via reducing mechanisms.

Suganya et al. (2007)³⁵ isolated antioxidant active compounds from methanol crude extracts of the leaves of Psidium guajava grown in Thailand. The isolated compounds were screened for their in vitro antioxidant activity by a DPPH free radical scavenging assay. Results indicate that three isolated compounds contribute importantly to the antioxidant activity of Psidium guajava leaves, providing a scientific basis for the use of this plant in traditional medicine. The most active compound was found to be quercetin along with two flavonoid compounds, quercetin–3–O–glucopyranoside and morin. The isolated quercetin, quercetin–3–O–glucopyranoside and morin showed significant scavenging activity with IC50 of 1.20±0.02, 3.58±0.05 and 5.41±0.20 µg/nml, respectively.

Wang et al. ³⁶ studied the antioxidative activities of the extracts from Psidium guajava leaves. The Psidium guajava was submersed with distilled water, 65% ethanol and 95% ethanol respectively. The 3 extracts were obtained after the solutions were filtered, concentrated and dried. The scavenging rate to hydroxyl radicals and inhibiting rate to lipid peroxidation were analyzed for the 3 extracts. Their contents of total flavonoids were determined by ultraviolet spectrophotometry, and the components of total flavonoids were primarily identified by high performance liquid chromatography (HPLC) and ultraviolet–visible absorption spectrometry (UV). The extracts from distilled water, 65% ethanol and 95% ethanol respectively showed effects on scavenging hydroxyl radicals and inhibiting lipid peroxidation in the dose–dependent manner, had 50% effective concentration (EC50) on scavenging hydroxyl radicals of 0.63, 0.47 and 0.58g/L, had EC50 on inhibiting lipid peroxidation of 0.20, 0.035, 0.18g/L and had total flavonoids contents of 3.28, 30.71 and 55.98g/kg respectively. The aquatic and the ethanol extracts from Psidium guajava possess the potential antioxidative activities in the study. The flavonoids may be one of their antioxidative components.

Ju–wen et al. ³⁷ have reported potent antioxidant activity in guava leaf extracts and attributed it to the phenolic compounds. Two phenolic acids, gallic acid and ferulic acid, isolated from Psidium guajava leaf extracts were also identified ³⁸. Zhang et al. ³⁹ also identified gallic acid, chlorogenic acid, kaempferol, procatechuic acid, ferulic acid, caffeic acid, quercetin and rutin in acetone extract from guava leaf. The antioxidant, free radical scavenging effects and protection from UVB–induced oxidation of aqueous extracts from Psidium guajava leaf were also stronger than that of water soluble extracts of some nutraceutical herbs including Camellia sinensis (gamma–aminobutyric acid (GABA tea, a functional tea by fermenting fresh tea leaves under nitrogen gas), or Toona sinensis Roem. and Rosemarinus officinalis L. ⁴⁰. Although the contents of total phenolic compounds and flavonoids in Psidium guajava leaf extracts were lower than that of aqueous rosemary extracts, Psidium guajava leaf extracts showed the strongest antioxidant activity in most of the tested methods. This fact suggests that guava leaf extracts are a good source of water soluble natural antioxidants.

Soman et al.⁴¹ investigate the antioxidant as well as antiglycative potential of ethyl acetate fraction of Psidium guajava leaves. Oral administration of the extract at different doses showed a significant decrease in blood glucose level. It also showed an improved antioxidant potential as evidenced by decreased lipid peroxidation and a significant increase in the activity of various antioxidant enzymes such as catalase, superoxide dismutase, glutathione peroxidase and glutathione reductase. Glycated hemoglobin as well as fructosamine which are indicators of glycation was also reduced significantly in treated groups when compared to diabetic control. In vitro studies also support the antioxidant as well as antiglycative potential of Psidium guajava leaves.

Ling et al.⁴² evaluated thirteen Malaysian plants for antioxidant capacity and cytotoxicity. Psidium guajava is one among those which has antioxidant capacity. Antioxidant capability, total phenolic content, elemental composition, as well as it cytotoxicity to several cell lines of the aqueous and ethanolic extracts from different parts of these selected plants were determined. In general, the ethanolic extracts were better free radical scavengers than the aqueous extracts and some of the tested extracts were even more potent than a commercial grape seed preparation. Similar results were seen in the lipid peroxidation inhibition studies. Their findings also showed a strong correlation of antioxidant activity with the total phenolic content and conclude that although traditionally these plants are used in the aqueous form, its commercial preparation could be achieved using ethanol since a high total phenolic content and antioxidant activity is associated with this method of preparation.

Akinmoladun et al.⁴³ evaluated methanolic extracts of 10 selected Nigerian medicinal plants and Psidium guajava is one among them. They assessed phytochemical constituents and antioxidant and free radical scavenging activities using seven different antioxidant assay methods. Phytochemical screening gave positive tests for steroids, terpenoids, and cardiac glycosides, alkaloids, saponins, tannins, and flavonoids contained in the extracts. Psidium guajava contained the highest amount of total phenolics, and the highest amounts of total flavonoids were found in the leaf extracts of Cassia alata, Chromolaena odorata, and Psidium guajava. Percentage DPPH radical scavenging activity was highest in Spondias mombin and Psidium guajava and compared with values obtained for ascorbic acid and gallic acid. The reductive potential was highest in Psidium guajava and least in Securidaca longepedunculata. DPPH assay correlated well with total phenolic contents and reductive potential and fairly with lipid peroxidation inhibitory activity. There was a good correlation between total phenolic contents and reductive potential and a fair correlation between total phenolic contents and lipid peroxidation inhibitory activity. These results suggest that the methanolic extracts of the studied plant parts possess significant antioxidant and radical scavenging activities that may be due to the phytochemical content of the plants and as such make them potential candidates as natural chemoprophylactic agents.

CONCLUSION:

Flavanoid, tannins, coumarin etc., are antioxidants and the observed antioxidant activity of Psidium guajava may be due to the presence of these compounds. Further studies to characterize the active principles and to elucidate the mechanism are in progress.

ACKNOWLEDGEMENT:

The authors are grateful to Alhaj. Er. A. Mohamed Yunus, Correspondent, M.I.E.T. Institutions, Trichy, whose spacious heart cheered our efforts to process this venture properly, successfully and to complete this task.

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Received on 24.04.2011

Accepted on 31.05.2011

Research Journal of Pharmacognosy and Phytochemistry. 3(4): July- August 2011, 143-147