Research Article | Open Access
Maria Christina Z. Manicad1 and Agustina G. Pattung2
1Bachelor of Science in Forestry Department, Apayao State College Luna Campus, San Isidro Sur, Luna 3813, Apayao, Philippines.
2Bachelor of Elementary Education Department, Apayao State College Conner Campus, Malama, Conner 3807, Apayao, Philippines.
Article Number: 9269 | © The Author(s). 2024
J Pure Appl Microbiol. 2024;18(2):1209-1213. https://doi.org/10.22207/JPAM.18.2.39
Received: 25 January 2024 | Accepted: 15 April 2024 | Published online: 27 May 2024
Issue online: June 2024
Abstract

The present study sought to perform antibacterial screening of the Lubeg leaf extracts derived from a 12-year Lubeg Plantation at Apayao State College, Payanan, San Gregorio, Luna, Apayao, against Escherichia coli and Staphylococcus aureus. The antimicrobial activity was evaluated using the disk diffusion method. The 100% Lubeg leaf extracts exhibited the highest mean zone of inhibition against E. coli after 20 hours while after 30 hours of incubation, the 50% Lubeg leaf extracts exhibited the highest mean zone of inhibition and active antibacterial activity against E. coli. Meanwhile, the 25% Lubeg leaf extracts exhibited the highest mean zone of inhibition and active antibacterial activity against S. aureus for both 20 hours and 30 hours of incubation. The Lubeg leaf extract has potential antimicrobial activities against these two bacterial strains, revealing its high potential as an antibacterial agent.

Keywords

Antibacterial, Syzygium lineatum, E. coli, S. aureus

Introduction

The genus Syzygium comprises 1200–1800 species that belong to the family of Myrtaceae.1 One species is commonly called “Lubeg,” “Malubeg,” and “Alebadu,” or Philippine cherry. It inhabits some areas of Cagayan, Apayao, and Isabela provinces of Region 02.2 Lubeg abundantly thrive in Apayao and in terms of medicinal value, Lubeg have a wide range of medicinal properties, including digestive, antidiabetic, astringent, antihelminthic, anti-bacterial, analgesic, anti-inflammatory, ant-oxidant, anti-hyperglycemic, gastro-protective agents, stomachic activity, anti-scorbutic activity, diuretic, anti-carminative, anti-genotoxicity, antileishmanial and anti-fungal activity.

Researches were conducted on Lubeg species along propagation,3 molecular characteristics and chemical composition of leaves and fruits,3 taxonomic classification and molecularly identification using the rbcL gene.4 Moreover, in Apayao, there was research on the development of Lubeg fruits into processed products such as wine, fruit preserved, and nonalcoholic beverages.5 Further, research on the qualitative phytochemical composition of Lubeg species in Apayao revealed presence of flavonoids, tannins, and saponins6 and total phenolic contents of the extracts of Lubeg leaves at 1.05 mg/g of the Gallic acid equivalent (GAE),7 however, the antibacterial properties need to be explored.

The preceding results of studies previously conducted are encouraging, laying the groundwork for the widespread use of the Lubeg in traditional and folk medicines. The literature review showed chemical screening of Lubeg and other species of Syzygium but not for the antimicrobial properties of Lubeg species. Balangcod et al.8 mentioned the prevalence of diseases and the increasing prices of medicine have resulted in the demand for the discovery of less expensive but more vital sources of drugs. Different diseases are emerging, becoming primary health problems experienced by the community and human populations. With this, there is a need to discover potential plant resources that have medicinal value.

This study performed antibacterial screening of the Lubeg leaf extracts derived from a 12-year-old Lubeg Plantation at Payanan, San Gregorio, Luna, Apayao, and determined the anti-bacterial properties of Lubeg leaves against Escherichia coli and Staphylococcus aureus.

Materials and Methods

Plant extraction preparations
Plant specimens’ fresh leaves were collected from a 12-year-old Lubeg Plantation at the Experimental site of Apayao State College, Payanan, San Gregorio, Luna, Apayao. The collected leaves were air-dried for two (2) weeks, chopped finely, and stored in a tightly sealed container.

100 grams of the finely chopped sample were macerated using 80% Ethanol and stood for 24 hours for each plant. The mixtures were then filtered and concentrated in a water bath with a maintained temperature of 50°C at 20% of the filtrate was left. These extracts were used both for antibacterial screening.

Antibacterial activity of the plant extracts
Bacterial strains
Two (2) microbial strains, E. coli and S. aureus, were selected for the treatment. The microbial cultures were provided from the culture collection of the University Science Laboratories of Saint Paul University Philippines, Tuguegarao City, Philippines.

Inoculum preparation
The test organism was subcultured into test tubes containing nutrient agar using a loopful from each of their agar stands. At 37°C, the test tubes were incubated for 24 hours. The acquired microorganisms were standardized using a standard saline solution to ensure the bacteria had a consistent population density.

Antibacterial activity of the plant extracts
The disk diffusion method is used to evaluate the antimicrobial activity of each leaf plant extract.9 Four treatments namely, 25%, 50%, 75%, and 100% Lubeg leaf extracts were used in the study while Amoxicillin was used as a control. The plates were then incubated for 20 hours and 30 hours at 37°C.

Determination of zone of inhibition
The antibacterial properties of the Lubeg leaf extracts and the antibiotics were observed by measuring the diameter of the zone of inhibition in millimeters using a vernier calliper laid in three (3) replications after the experiment. The activity of the Lubeg leaf extract was compared with corresponding references.10

RESULTS

The antibacterial activity of the Lubeg leaf extracts was determined by the disk diffusion method against E. coli and S. aureus, provided from the culture collection of University Science Laboratories of Saint Paul University Philippines, Tuguegarao City, Philippines. There were four ethanolic leaf extract concentrations tested such as 25%, 50%, 75%, and 100% Lubeg leaf extracts. After 20 hours and 30 hours of incubation, the plates were observed for the zone of inhibition in millimeters using a vernier calliper. Amoxicillin was also tested on the same bacteria to compare the antibacterial activity of the Lubeg leaf extracts and the antibiotic. Table 1 shows the standard zones of inhibition and corresponding inferences.

Table (1):
Standard zones of inhibition and corresponding inferences.10

 Zone of Inhibition
Inferences
< 10 mm
Inactive
10- 13 mm
Partially Active
14-19 mm
Active
>19mm
Very Active

The 100% Lubeg leaf extracts exhibited the highest mean zone of inhibition against E. coli after 20 hours of incubation. Moreover, all four (4) treatments showed inactive antibacterial activity against E. coli. However, after 30 hours of incubation, 50% of Lubeg leaf extracts exhibited active antibacterial activity and the highest mean zone of inhibition of 16.83 mm against E. coli.  All the other three treatments showed partial activity against E. coli. Moreover, the mean zone of inhibition increased with increased incubation periods.

Table (2):
Anti-bacterial screening of Lubeg leaves against Escherichia coli and Staphylococcus aureus

Treatment E. coli S. aureus
20 hrs 30 hrs 20 hrs 30 hrs
Mean Zone of Inhibition (mm)  Inferences Mean Zone of Inhibition (mm)  Inferences Mean Zone of Inhibition(mm)  Inferences Mean Zone of Inhibition(mm)  Inferences
25% Lubeg Leaf Extract 7.55 Inactive 11.25 Partially active 14.66 Active 14.17 Active
50% Lubeg Leaf Extract 7.50 Inactive 16.83 Active 13.33 Partially Active 8.75 In Active
75% Lubeg Leaf Extract 8.75 Inactive 11.10 Partially active 9.97 Inactive 9.72 Inactive
100%Lubeg Leaf Extract 9.48 Inactive 12.27 Partially active 10.13 Partially Active 10.53 Partially Active
Control 77.48 Very active 55.00 Very active 41.58 Very active 56.90 Very Active

As shown from Table 2, the highest mean zone of inhibition was exhibited by 25% Lubeg leaf extracts against S. aureus after 20 hours of incubation. Moreover, this treatment demonstrated active antibacterial activity for S. aureus as indicated by the 14.66 mm diameter zone of inhibition. The lowest mean of inhibition was 75% Lubeg leaf extract, with a mean zone of inhibition of 9.97 mm. After 30 hours of incubation, 25% Lubeg leaf extracts still exhibited the highest mean zone of inhibition against S. aureus. On the other hand, the mean zone of inhibition was decreased as the incubation period increased for 25%, 50%, and 75% Lubeg leaf extracts against S. aureus. Only the 100% Lubeg leaf extract showed an increased mean zone of inhibition after 30 hours of incubation. Amoxicillin showed very active antibacterial activity against E. coli and S. aureus, as positive control.

DISCUSSION

Syzygium is a large genus of plants throughout the tropical regions. The genus has medicinal applications in the pharmaceutical, cosmetic, agricultural, and food industries. Moreover, it is known for medicinal properties like anti-bacterial activity. The genus was utilized in different ethnomedicinal systems worldwide.11 The biological activity of some plant species of Syzygium are reported, and some are not; thus, it is a subject of interest to researchers of medicinally useful parts like leaves, roots, fruit, seed or bark.12

The present research constitutes the first investigation conducted on the Syzygium lineatum locally called Lubeg against E. coli and S. aureus in the province of Apayao. The antibacterial properties against Gram-positive and Gram-negative bacteria were evaluated through inhibitory assay. Remarkably, the leaf extracts displayed activity against E. coli and S. aureus in the disk diffusion method. Furthermore, variations in antibacterial properties were observed from inactive to active antibacterial activity against the strain of bacteria following corresponding references by Guevarra.10 The results are aligned with reports that Syzygium species show rich medicinal applications.13 Recent studies demonstrated the efficacy of Syzygium species against different types of bacterial strains,14 which contain various phytochemicals that exhibit antioxidant and antimicrobial properties that result in their inhibitory abilities 15 and methanolic extracts have biological activities like antibacterial properties.16 The findings are supported by the previous findings that Lubeg contains phenolic,6-7 tannins, saponins, steroids, and flavonoids.

CONCLUSION

The study focused on the screening of Syzygium lineatum or Lubeg leaf extracts against E. coli and S. aureus. The findings revealed that the leaf extracts have active antibacterial components that inhibit the growth of E. coli and S. aureus. Remarkably, the 50% Lubeg leaf extracts against E. coli and 25% Lubeg leaf extracts against S. aureus. The presence of antibacterial activities of Lubeg leaves is thought to have health-promoting qualities and beneficial importance in medicinal sciences.

Declarations

ACKNOWLEDGMENTS
The authors would like to extend their profound gratitude to the Apayao State College for providing funds; St. Paul University Philippines, Tuguegarao City for allowing the authors to perform the antibacterial screening in their Science Laboratory Department; Dr. Jimmy G. Catanes who served as their consultant in the conduct of the study; and Ma. Cristina Aguda, Roderick Marcos, and Janine Sapad, for their help in the conduct of the study for the collection and processing of the Lubeg specimens.

CONFLICT OF INTEREST
The authors declare that there is no conflict of interest.

AUTHORS’ CONTRIBUTION
Both authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.

FUNDING
This work was supported by the Apayao State College, Philippines, with order number OP-S0-2022-04-002.

DATA AVAILABILITY
All datasets generated or analyzed during this study are included in the manuscript.

ETHICS STATEMENT
Not applicable.

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