Some Medicinal Benefits of Psidium Guajava (Guava) and Portulaca oleracea



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Some Medicinal Benefits of Psidium Guajava (Guava) and Portulaca oleracea


Gehan A.E. El-Emary

Faculty of Technology and Development, Zagazig University, Egypt.

*Corresponding author: aemarygehan@yahoo.com

ABSTRACT

Crude extract of two medicinal plants (leaves of Psidium Guajava (Guava) and seeds of Portulaca oleracea) were tested to evaluate the flavonoids and phenol contents, antioxidant and antimicrobial activities. The content of total Flavonoids in the extract of Portulaca oleracea (35.50 mg GAE/100) is higher than that in the extract of Psidium Guajava (Guava) (32.667mg GAE/100). Concerning total Phenols, data revealed that the total phenol in the extract of Portulaca oleracea (45.775mg GAE/100) is higher than that in the extract of Psidium Guajava (Guava) (39.473mg GAE/100). Me-OH extract of Portulaca oleracea exhibited the highest DPPH radical scavenging activity compared to Psidium Guajava (Guava).

Portulaca oleracea extract exhibited obvious antimicrobial behavior against all microorganisms under test throughout this study; this behavior was moderate (++) antimicrobial activity against all Gram negative organisms(Escherichia coli, Pseudomonas aeruginosa and Salmonella typhi) and expressed very strong (++++) antimicrobial activity when tested against three types of Gram positive bacteria (Staphylococcus aureus, Bacillus subtilis and Listeria monocytogenes), one type of Fungi (Aspergillus niger) and one type of Yeast (Candida albicans); Psidium Guajava (Guava) extract showed weaker antimicrobial activity when compared to Portulaca oleracea extract against tested microorganisms.


Keywords: Portulaca oleracea, Psidium Guajava (Guava), antioxidant, antimicrobial, flavonoids, phenol.

INTRODUCTION

Plants have been used in traditional medicine for several thousand years [1]. During the last few decades, there has been an increasing interest in the study of medicinal plants and their traditional use in different parts of the world [27, 15, 3, 20 and 34]. Documenting the indigenous knowledge through ethnobotanical studies is important for the conservation and utilization of biological resources.

Today according to the World Health Organization (WHO), as many as 80% of the world's people depend on traditional medicine for their primary healthcare needs. There are considerable economic benefits in the development of indigenous medicines and in the use of medicinal plants for the treatment of various diseases [4].

Due to fewer communication means, poverty, ignorance, and unavailability of modern health facilities, most people especially rural people are still forced to practice traditional medicines for their common day ailments. Most of these people perform the poorest link in the trade of medicinal plants [25].

A vast knowledge of how to use the plants against different illnesses may be expected to have accumulated in areas where the use of plants is still of great importance [12].

Industrialization has led to many modifications in the lifestyle of the world’s populations, giving rise to increase the indices of several diseases, including chronic degenerative diseases such as insulin resistance, diabetes mellitus, dyslipidaemia, metabolic syndrome and cardiovascular diseases, reducing the quality of life and increasing costs of hospitalizations, medications and other public health interventions [37,33].

Studies have demonstrated that the consumption of fruits, vegetables, and seeds can be helpful to prevent the risk factors of many diseases due to the bioactive compounds. Many plants have been used for the purpose of reducing risk factors associated with the occurrence of chronic disorders and for many other purposes [39, 6, 5, 41& 14].

Psidium guajava L. is a small medicinal tree that is native to South America. It is popularly known as guava (family Myrtaceae) and has been used traditionally as a medicinal plant throughout the world for a number of ailments. There are two most common varieties of guava: the red (P. guajava var. pomifera) and the white (P. guajava var. pyrifera) [23 & 19].



All parts of this tree, including fruits, leaves, bark, and roots, have been used for treating stomachache and diarrhea in many countries. Leaves, pulp, and seeds are used to treat respiratory and gastrointestinal disorders, and as an antispasmodic, anti-inflammatory, as a cough sedative, anti-diarrheic, in the management of hypertension, obesity and in the control of diabetes mellitus. It also possesses anticancer properties [35]. The seeds are used as antimicrobial, gastrointestinal, anti-allergic and anti-carcinogenic activity [32, 30, 21&- 8].

Portulaca oleracea; known also as Purslane is distributed all over the world; Portulaca oleracea is a herbaceous annual, native to many parts of Europe, found in the East and West Indies, China, Japan and Ascension Island, and though found also in the British Isles is not indigenous there [2]. It is a weedy summer annual species that is abundant throughout the world, invading vegetable gardens, bare areas, low-maintenance lawns, ornamental plantings, and agricultural areas. It is particularly well adapted to the warm, moist conditions found in California’s irrigated agricultural and ornamental sites. It has been cultivated in India and the Middle East and has been popular in Europe since the Middle Ages. Common purslane germinates in California from February to March in the southern desert areas to late spring in cooler areas when the soil temperature reaches about 60°F. For an early crop, the seed is best sown under protection in early spring and can then be planted out in late spring. Outdoor sowings in situ take place from late spring to late summer, successional sowings being made every two to three weeks if a constant supply of the leaves is required [16]. It germinates very near to or at the soil surface in large numbers after an irrigation or rain. Most of the tiny seedlings die, but the survivors grow rapidly and can produce flowers in a few weeks. The fleshy stems of common purslane can remain moist and viable for several days after cultivation and hoeing, and re-root to form “new” plants when gardens or fields are irrigated. Because of its ability to produce large numbers of seeds, common purslane can rapidly colonize any warm, moist site. It requires a moist light rich well-drained soil in a sunny position [16& 22]. Plants will not produce good quality leaves when growing in dry conditions [16]. The plants take about six to eight weeks to produce a crop from seed and can then be harvested on a cut and come again principle, providing edible leaves for most of the summer[16]. Common purslane is low in stature and forms dense mats. These vegetative mats utilize available moisture and nutrients and screen out light to the soil surface, preventing the emergence of other seedlings. Common purslane is unsightly, reducing the esthetic value of turf and ornamental plantings. In commercial situations, common purslane can limit summer vegetable production and reduce the efficiency of harvesting nut crops, such as almonds and walnuts, from the orchard floor.

Purslane in ancient times was looked upon as one of the anti-magic herbs, and strewn around a bed was said to afford protection against evil spirits [16]. It was supposed to protect from evil spirits and if carried was supposed to attract love and luck. It was carried by soldiers to protect them in battle. If laid on the bed, it was believed to protect that person from having nightmares [26]. The infusion may be used to clear the third eye and to wash the crystal ball or scrying mirror [28], no doubt a useful tip for our marketing colleagues! Dioscorides says "it reduces the desire to fornicate". In the latter sense, other authors also mention its anaphrodisiac powers [10], including this plant among the "four cold seeds", together with chicory, endive, and lettuce.

The high cost of pharmaceutical medications conduces to the search for alternative medicines to treat many ailments. In view of this, studies are necessary to confirm the effects of medicinal plants. The aim of this review is to show that several studies have demonstrated the presence of many different chemical compounds in P. guajava and Portulaca oleracea and their pharmacological effects.

MATERIALS AND METHODS

Plant collection

Fresh 2 plants samples were collected in Marsh 2016, from a well-known drug store in Cairo, plants were air defied.

Chemicals and reagents. DPPH (2, 2, diphenyl-1-picryl hydrazil radical), Folin–Ciocalteu phenol reagent and Gallic acid (3, 4,5-trihy-droxybenzoic) were obtained from Fluka Chemie (Buchs, Switzerland). Methanol, and, ethanol were from Riedel-de Haen (Sigma-Aldrich, Germany). Sodium carbonate was from PRS (Panreace Quimica, EU).

Microbial strains

All microorganisms used in this study (Table 1) were obtained from Faculty of Agriculture- Cairo University.

Preparation of Samples extract. Samples of Psidium guajava (Guava) and Portulaca oleracea were obtained from a local market in Cairo. A 100-g sample of each stem was cut into small pieces and then freeze-dried to give a final weight of 18.7 g for crude stem preparation. The crude preparation (10 g) was then mixed with 100 mL of absolute ethanol in a screw-capped flask and shaken at room temperature for 3 days to obtain ethanol extract. The resulting ethanol extract was subsequently filtered through filter paper (Whattman No.3) and centrifuged at 5000g for 10 min. The collected supernatants were lyophilized to give a final weight of 1.23 g (2.3% of the initial amounts) for the dried powder of ethanol extract.

Scavenging Activity onto DPPH Radicals. Scavenging activity on DPPH free radicals by the extract was assessed according to the method reported by [31]. Briefly, 50 µL of the ethanol extract containing varied amounts of powdered ethanol extract (1, 5, 10, and 50 µg/mL distilled water, respectively, in each reaction) was mixed with 1 mL of 0.1 mM DPPH-ethanol solution and 450 µL of 50 mM Tris-HCl buffer (pH 7.4). After the solution incubated for 30 min incubation at room temperature, reduction of DPPH free radicals was measured by reading the absorbance at 517 nm. In the experiment, L-ascorbic acid was used as positive control. The inhibition percent was calculated from the following equation: % inhibition) [(absorbance of control - absorbance of test sample)/absorbance of control] × 100.

Determination of total phenolic content. Total phenolic content: Total phenolic content was determined using Folin-Ciocalteu method as described by [34]. Gallic acid was used for the preparation of calibration curve. Volumes of 100 μL aliquots of 30, 60, 90, 150 and 180 mg/L Gallic acid solutions were added to test tubes followed by 1 mL of 10% (v/v) Folin-Ciocalteu reagent. The mixture was mixed and incubated for 5 min before addition of 0.8 mL 7.5% (w/v) sodium carbonate. The resulting mixture was further incubated for 1hour in dark at room temperature before absorbance was measured at 765 nm with a UV-VIS spectrophotometer (Genesys 20, Thermo Spectronic, USA). For the test samples, 1 mg/mL plant extracts were prepared and analyzed as described above. Results were expressed as mg Gallic Acid Equivalent (GAE) per 100 g dry weight of plant.

Determination of total flavonoid content. The total flavonoid content of the three medicinal samples was determined by using of a modified colorimetric method [7]. An air dried plant material (25 mg) was ground in a mortar with 10 ml 80 % methanol. The homogenous mixture obtained was allowed to stand for 20 min. at room temperature, followed by filtration through filter G4. An aliquot of 0.4 ml of filtrate was mixed with 0.6 ml distilled water, 5 % NaNO2 solution (0.06 ml) and the mixture was allowed to stand for 5 min at room temperature. After 6 min 10 % AlCl3 solution (0.06 ml) was added to the mixture. Immediately, 1 N NaOH (0.4 ml) and 0.45 ml distilled water were added to the mixture and allowed to stand for another 30 min. Absorbance of the mixture was determined at 510 nm and (+) catechin was used as standard compound for the quantification of total flavonoid content. All values were expressed as milligram of catechin equivalents per 1 gram dry weight. Data was recorded as mean ± SD for three replicates.



Antimicrobial activities.

The antimicrobial activities of the tested plants were measured by disk assay procedure [31]. against the following indicator microorganisms Staphylococcus aureus(ATCC 25923), Bacillus subtilis (ATCC6633), Pseudomonas aeruginosa (ATCC9027), Escherichia coli (ATCC 35218), Listeria monocytogenes (ATCC7644), Salmonella typhimurium (ATCC14028), Aspergillus niger(ATCCnrrl 1957) and pathogenic yeast (Candida albicans (ATCC 10231). Discs were used in assay agar plates. Soft agar medium culture seeded or inoculated with the tested microorganisms was layered over 10 ml of hard agar (2%). Plates were incubated at various temperatures for required incubation periods according to strain type (Table 1). A specific volume containing 40 µg/ml of each extract was impregnated into sterilized paper discs (Whattman No. 1) of 6 mm in diameter. After drying, the paper discs were plated on the assay plates in triplicate and left at 4 °C for 24 h to allow maximum diffusion of the test sample. After incubation time, the distinct zone of inhibition surrounding the disc was measured. Antimicrobial activities were expressed as inhibition diameter zones in millimeters (mm) as follows: - (negative) = 0 mm; + (weak) = 1-4 mm; ++ (moderate) = 5-10 mm; +++ (strong) = 10-15 mm and ++++ (very strong) ≥ 16 mm. The experiment was carried out in triplicate and the average zone of inhibition was calculated.

Table 1

Microbial strains used to test the antimicrobial activities of the three tested Medicinal Plants



Microbial Group

Indicator Strain

Cultivation Conditions*

Gram positive Bacteria

Staphylococcus aureus(ATCC 25923)

Bacillus Subtilis(ATCC6633)

Listeria monocytogenes(ATCC7644)


TSA+ YE, 37°C

TSA+ YE, 37°C

TSA+ YE, 37°C


Gram negative Bacteria

Salmonella typhimurium(ATCC14028)

Escherichia coli(ATCC35218)

Pseudomonas aeruginosa(ATCC9027)


TSA+ YE, 37°C

TSA+ YE, 37°C

TSA+ YE, 37°C


Fungus

Aspergillus niger(ATCCnrrl 1957)

PDA, 25°C

Yeast

Candida albicans(ATCC 10231)

TSA+ YE, 30°C

*TSA = Trypticase Soy Agar; YE: = Yeast Extract; PDA = Potato Dextrose Agar.

RESULTS AND DISCUSSION

 Total Flavonoid and Total Phenol Contents of the Extracts:

It has been recognized that flavonoids show antioxidant activity and their effects on human nutrition and health are considerable. The mechanisms of action of flavonoids are through scavenging or chelating process [24, 11]. Phenolic compounds are a class of antioxidant agents which act as free radical terminators [36]. Total Phenols and Total flavonoids contents of the three extracts are represented in Table 2.

Table 2. Flavonoids and Phenols contents of three tested medicinal plants


Plant Species

T. Phenols(mg GAE/100)

T. Flavonoids(mg GAE/100)

Psidium Guajava (Guava)

39.473

32.667

Portulaca oleracea





45.775

35.50





As shown from Table 2, both plants (Psidium Guajava (Guava) and Portulaca oleracea) possess high total phenol rates; 39.473 mg GAE/100 and 45.775 mg GAE/100 respectively; concerning total flavonoids contents, Psidium Guajava (Guava) extract contains 32.667 mg GAE/100, while total flvonoides in the extract of Portulaca oleracea was 35.50 mg GAE/100. Due to the fact that only crude extracts were used in our experiment, so the higher yield might be contributed by other polar compounds besides phenolics, namely polysaccharides, plant debris and so on. This finding is compatible with the same proposed by [40].


Antioxidant activity:

The stable free radical DPPH has been widely used to test the free radical-scavenging ability of various dietary antioxidants [9]. The reduction capability of the DPPH radical is determined by the decrease in its absorbance at 517 nm, induced by antioxidants [13]. The free radical-scavenging activity of all extracts at 40-100 µl was evaluated as % inhibition in the DPPH radical model system. The DPPH radical-scavenging activity of Psidium Guajava (Guava) and Portulaca oleracea methanol extract, at different concentrations (40µl, 60 µl, 80 µl &100 µl) is shown in Table 3.

Table 3

Free radical scavenging activities of two tested medicinal plants



Sample I.D.

Sample Concentration

(5gm/25ml)



T. Antioxidant %

Psidium Guajava (Guava)

40 µl

80.07

60 µl

89.74

80 µl

90.06

100 µl

90.27

Portulaca oleracea





40 µl

48.05

60 µl

50.87

80 µl

52.44

100 µl

56.52

Table (3): Free radical scavenging activities of two tested medicinal plants

A concentration-dependent scavenging activity was clearly demonstrated. At 100µl, the inhibition percent of Me-OH, extracts was determined as 90.27% and 56.52% in Psidium Guajava (Guava) and Portulaca oleracea, respectively. These results indicated that Me-OH extract of Psidium Guajava (Guava) exhibited the highest DPPH radical scavenging activity compared to Portulaca oleracea. The highest DPPH radical scavenging activity of Psidium Guajava (Guava) seems to be attributed to the high concentration of phenolic compounds in this extracts, which could be the electron donors, and hence can react with free radicals to convert them to more stable products and terminate radical chain reaction, that could make this herb contains exploitable antioxidant activities; this finding is in a full agreement with that obtained by [43] and [38].

Antimicrobial Activity

Results of the Antimicrobial activity is presented in Table 4. As shown from the table,

Both plants, showed different antibacterial behavior against the targeted organisms; Psidium Guajava (Guava) showed moderate effect (++) against all Gram +ve organisms (Staphylococcus aureus, Bacillus subtilis and Listeria monocytogenes), also it showed the same effect against Aspergillus niger (Fungus) and Candida albicans (Yeast); on the other hand Psidium Guajava (Guava) showed weak effect against Gram –ve organisms (E.coli, Pseudomonas aeruginosa and Salmonella typhi).


Table 4

Antimicrobial activities of Psidium Guajava (Guava), and Portulaca oleracea




Antimicrobial activities*




Gram positive bacteria**

Gram negative bacteria

Fungus & Yeast

Plant extracts

S. aureus

B. subtilis

L. monocytogenes

E. coli

P. aeruginosa

S. typhi

A. niger

C. albicans

Psidium Guajava (Guava)Extract

extract


++

++

++

+

+

+

++

++

Portulaca oleracea Extract


Extract


++++

++++

++++

++

++

++

++++

++++




























* Antimicrobial activities were expressed as inhibition diameter zones in millimeters (mm) : - (negative) = 0 mm; + (weak) =1-4 mm’ ++ (moderate) = 5-10 mm; +++ (strong) = 10-15 mm; and ++++ (very strong) ≥ 16 mm.

** Microorganisms used were S. aureus (ATCC 25923), B. subtilis (ATCC 6633), L. monocytogenes (ATCC 7644), E.coli (ATCC 35218), P. aeruginosa (ATCC 9027), S. typhi (ATCC 14028, A. niger (ATCC 1957) and C. albicans (ATCC 10231).


Portulaca oleracea showed very strong effect against all tested organisms except Gram-ve organisms (Staphylococcus aureus, Bacillus subtilis, Listeria monocytogenes, Aspergillus niger and Candida albicans), while it showed only moderate effect against Gram –ve organisms (E.coli, Pseudomonas aeruginosa and Salmonella typhi). This results revealed that both plant extracts were effectively killed all tested microorganisms; this is in agreement with the same results proposed by [29].

Conclusion

The findings of this study support the view that some medicinal plants are promising sources of potent antioxidants and may be efficient as preventive agents in the pathogenesis of some diseases. It can also be used in stabilizing food against oxidative deterioration. Psidium Guajava (Guava) and Portulaca oleracea, which contain the highest amount of flavonoid and phenolic compounds, exhibited the greatest antioxidant activity, are two of these medicinal plants that showed greater antimicrobial activity against all tested microorganisms, however, this study should follow by extensive phytochemical and pharmacological studies on plants used in this study.











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