Barriers and facilitators to hospital pharmacists’ engagement in medication safety activities: a qualitative study using the theoretical domains framework Page 19 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 In vitro antimicrobial assessment of seeds of selected medicinal plants in Sri Lanka Gowri Rajkumar1*, Mihiri Rangika Jayasinghe1 and Vinotha Sanmugarajah2 ABSTRACT Introduction: People are suffering from many noncommunicable diseases as a result of the COVID pandemic and the stress that has followed after it. Diabetes mellitus is a complex non- communicable disease and its incidence in Sri Lanka is almost high. While a number of antidiabetic medications are available, herbal management of diabetes is encouraged due to its low side effects and efficacy. Examining the antibacterial properties of anti-diabetic plants may be highly valued because, diabetics are chronic immunocompromised individuals who are more prone to microbial infections. To focus this aim, the present in-vitro antimicrobial assessment carried out for the seeds of selected four medicinal plants, such as Syzygium cumini (L.) Skeels, Sinapis alba L., Trigonella foenum-graecum L. and Nigella sativa L. that are commonly used for diabetes management in Sri Lanka. Materials and methods: Crude ethanol extract from the seeds has been studied for their antibacterial potential against three bacterial strains such as Enterococcus faecalis, Staphylococcus aureus and Escherichia coli by using agar well diffusion method in triplicates. The statistical analysis was performed using a one-way analysis of variance. Results: The seed extract of S. cumini showed the highest value of zone of inhibitions (E. faecalis: 24.70±037, S. aureus: 16.15±1.20 and E. coli: 10.37±1.51 mm) and S. alba exhibited the lowest value of zone of inhibition (1.08±2.65, 1.08±2.65, 0 mm) for all selected pathogens respectively and which were comparable to the positive control streptomycin (E. faecalis: 25.45±1.18, S. aureus: 21.08±0.26 and E. coli: 19.37±1.35mm). Conclusion: The result shows that S. cumini seed extract poses the highest antimicrobial activity in selected bacteria. Therefore, this seed is potential to be further developed as an herbal antibiotic for the management of infection in diabetes in future. ARTICLE HISTORY: Received: 7 July 2022 Accept: 3 January 2023 Published: 31 January 2023 KEYWORDS: Medicinal plants, antimicrobial activity, agar well diffusion method, seeds, Syzygium cumini HOW TO CITE THIS ARTICLE: Gowri Rajkumar, Mihiri Rangika Jayasinghe & Vinotha Sanmugarajah (2023). In vitro antimicrobial assessment of seeds of selected medicinal plants in Sri Lanka. Journal of Pharmacy, 3(1), 19-26. doi: 10.31436/jop.v3i1.179 Authors’ Affiliation: 1 Department of Botany, Faculty of Science, University of Jaffna, Jaffna, 40000, Sri Lanka 2 Unit of Siddha Medicine, University of Jaffna, Jaffna, 40000, Sri Lanka. *Corresponding author: Email address: gowrir@univ.jfn.ac.lk Phone: +94773604037 mailto:gowrir@univ.jfn.ac.lk Page 20 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 Introduction The number of people with diabetes is exponentially increasing world-wide and has become an important global public health problem (Global report on diabetes, 2016; Shehab et al., 2022). Sri Lanka, despite being a developing nation, has always maintained health indices on par with most developed countries. Currently, Sri Lanka is in the midst of its worst economic crisis in history (Pathmanathan & Abhayaratna 2022). In Sri Lanka, people are suffering from many noncommunicable diseases as a result of the COVID pandemic and the stress that has followed after it. That condition gets more critical as a result of the country's economic situation. Diabetes mellitus is one of the most common endocrine disorders in Sri Lanka. 10% of the Sri Lankan population and 18% of the urban population are affected by it (Pathmanathan & Abhayaratna 2022). Diabetes mellitus is characterized by hyperglycaemia and is especially classified into two types, Type-I (Insulin dependent diabetes) and type-II diabetes (non-insulin dependent diabetes) (IDF Diabetes Atlas, 2013; AOAC, 2016; American Diabetes Association, 2019; King, 2012). The incidence of diabetes (% of the population aged 20-79) in Sri Lanka was 11.3% in 2021 (The World Bank Group, 2022). This condition may be due to the common risk factors such as genetic, environmental, different life stylishness and physical inactivity (Wu et al., 2014; Arawwawala, 2006) and prevalence of depression (Akter & Latif 2021). There is another type is gestational diabetes which mainly arises during the pregnancy (Buchanan, Xiang & Page, 2012). The oxidative stress is a recognized pathogenic mechanism in the development and progression of diabetes which reasons owing to augmented free radical production and weakened antioxidant defences (Unuofin & Lebelo, 2020). A study mentioned that the antiglycation properties of herbal extracts and their complexes powerfully interrelated with their antioxidant capacity with that antioxidant and anti-glycation activities are associated strongly with phenol and flavonoid contents (Babich et al., 2022). Generally, antibacterial actions are facilitated by the immune-modulating and antioxidant capabilities of medicinal plants (Aryal et al., 2021). Antibacterial action is the most significant distinctive of medical textiles, to deliver satisfactory defence against microbes, biological fluids, and infection transmission (Alihosseini, 2016). Prevention of food spoilage and food intoxication pathogens is regularly attained by use of chemical preservers. Plant extracts have been used to control food poisoning diseases and preserve foodstuff (Mostafa et al., 2018). An extensive range of biological constituents as alkaloids, flavonoids, glycosides, terpenoids, phenols, and coumarins have been stated from different parts of the plant, which are responsible for numerous biological activities as well as antimicrobial, antioxidant, and anti-inflammatory properties (Sarkar, Salauddin & Chakraborty, 2020; Phuyal et al.,2020). While a number of antidiabetic medications are available, herbal management of diabetes is encouraged due to its low side effects and efficacy. Examining the antibacterial properties of anti-diabetic plants may be highly valued because, diabetics are chronic immunocompromised individuals who are more prone to microbial infections (Hegazy et al., 2021). For this purpose, this in-vitro antimicrobial assessment was performed for the ethanol extracts of four seeds of medicinal plants namely Syzygium cumini (L.) Skeels, Sinapis alba L., Trigonella foenum-graecum L. and Nigella sativa L. against three bacterial strains as E. faecalis, S. aureus and E. coli. Materials and methods Chemicals and reagents Ethanol, streptomycin, distilled water and nutrient agar were purchased from Sigma-Aldrich. All reagents and chemicals were of analytical grade. Collection of medicinal plants and preparation of the seed extract The plant materials with seeds of S. cumini, S. alba, T. foenum - graecum and N. sativa (Table 1) were collected and botanically authenticated by a Curator of the National Herbarium Center, Department of National Botanic Garden, Peradeniya, Sri Lanka. The fresh seeds were washed in tap water for several times to remove the soil and dust particle. Then they were air dried in thoroughly at room temperature until dried and blended to form a fine powder and stored in airtight containers at room temperature until needed for analysis. Fifty grams of powered materials of each seed were separately weighed and placed in 500 ml of culture bottles. As much as 150 ml of absolute ethanol was added to it and mixed well. Lid of each bottle were covered with para film. The solution was kept for 5 days with occasional shaking by using shaker at 150 rpm for 15 minutes in every morning and evening. They were filtered through Whatman filter paper No.1. The part of filtered content was concentrated by using rotatory evaporator (BUCHI, Chi Minh City, Vietnam) at 52 0C (Rajkumar, Jayasinghe & Sanmugarajah, 2021). Crude extracts were kept at 20 0C for further analysis. Test microorganisms Three bacterial strains were provided by the Faculty of Science, University of Jaffna were used for the antimicrobial tests, according to Table 2. All the test strains were preserved on nutrient agar slants at 4 0C and sub-cultured on to nutrient broth for 24 hours prior to testing. These bacteria served as test pathogens for this assay. Page 21 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 Assay of Antimicrobial activity using Agar well diffusion method About 22.68 g of Nutrient Agar (NA) powder was dissolved in 810 ml of distilled water. Then 15 ml parts of the NA medium were poured into boiling tubes. Medium which was contained in the boiling tubes were autoclaved at 121 0C for 20-30 minutes. Then 15 mL of sterilized nutrient agar was mixed with 100 µl of bacterial suspensions inside the laminar air flow chamber. The mixture was stirred well and it was poured into sterile petri dishes separately (Dwivedi et al., 2017). After the solidification the wells were punched over the agar plates using sterile cork-borer (5mm in diameter) and 15 µL of plant extracts were added to the wells separately. The plates were incubated for 24 hours at 37 0C. Distilled water and Streptomycin (100 µg/µl) were used as the negative and the positive control respectively. After incubation the diameter of the formed inhibitory zones formed around each well were measured (mm) in four different fixed directions and recorded. Each experiment was conducted in triplicate. Data analysis Data were statistically analysed by one way Analysis of Variance and Tukey’s multiple comparisons at probability value (P˂0.05) using a SAS statistical package (version 9.1.3) and mean separation was performed by Least Significance Difference. Results are expressed as Mean ± SE and statistical significance was evaluated by ANOVA. Results This in vitro antimicrobial assay was done for four seeds of selected antidiabetic medicinal plants as S. cumini, S. alba, T. foenum-graecum and N. sativa against three selected common bacterial strains. Based on Figure 1, ethanolic seed extract of S. cumini exhibited the significant antimicrobial activity while the ethanolic seed extracts of S.alba represented minimum antibacterial activity against the all three bacteria as E. faecalis, E. coli and S. auereus. The results showed that significant amount of inhibition zone was obtained against all the tested bacterial strains which was comparable to the positive control streptomycin. Based on Figure 1, the ethanolic seeds extract of S. cumini exhibited the significant antimicrobial activity while the ethanolic seeds extract of S. alba showed minimum activity against all three bacterial strains at 37°C. There are also significant differences of antimicrobial activity among selected human pathogens. It showed that highest antimicrobial activity for E. faecalis, also it represented antimicrobial activity for both of gram negative and positive pathogens. While S.alba showed same antimicrobial activity against E. faecalis and S. aureus, inhibition zone is absent for E.coli. Based on Table 3, moderate antibacterial activity was showed by T. foenum-graecum and N. sativa against E. coli & E. faecalis and E. coli respectively. There is no any inhibition zone against S. aureus by seeds extract of T. foenum-graecum but it showed relative values of inhibition zones against both of E. faecalis and E. coli. There is no any inhibition zone against E. faecalis by N. sativa, and inhibition zones are represented against S. aureus and E. coli human pathogens respectively. Positive and negative control are represented by Streptomycin and distilled water respectively. When consider about positive and negative controls, Streptomycin exhibited the highest inhibitory effect against E. faecalis and the lowest inhibitory effect against E. coli. In negative controls there were no any inhibitory zones. Table 1: Medicinal plant seeds tested for their antibacterial activity in the study Botanical name Family name Common name Sinhala Tamil English Syzygium cumini Myrtaceae Mahadan Naval Black Plum Sinapis alba Brassicaceae Aba Kaduku Mustard Trigonella foenum- graecum Fabaceae Asumodhagam Vendayam Fenugreek Nigella sativa Ranunculaceae Kaluduru Karumjeerakam Black cumin Page 22 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 Table 2: Pure bacterial cultures Name Type ATCC No. Enterococcus faecalis Gram positive 29212 Staphylococcus auereus Gram positive 29213 Escherichia coli Gram negative 25922 Variation of mean diameter of seeds extracts of selected medicinal plants against selected human pathogens at 37℃. Mean followed by same letters are not significantly different by LSD at 5% level, P value <0.0001 Figure 1. Antimicrobial activity of selected seeds of anti-diabetic medicinal plants -10 -5 0 5 10 15 20 25 30 35 2 9 2 1 2 2 9 2 1 3 2 5 9 2 2 M e a n D ia m e te r ( M m ) Bacterial Strains of Human Pathogen Syzygium cumini Trigonella foenum-graecum Sinapis alba Nigella sativa Positive Control Negative Control Page 23 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 Table 3: Bacterial growth inhibition zones of selected seeds of anti-diabetic medicinal plants Plant species Diameter of the inhibition zones (mm) 29212 29213 25922 Syzygium cumini 24.7±0.37 16.15±1.20 10.37±1.51 Sinapis alba 1.08±2.65 1.08±2.65 0 Trigonella foenum-graecum 10.83±0.34 0 11.7±1.97 Nigella sativa 0 5.58±4.37 15.7±1.56 Positive control 25.45±1.18 21.08±0.26 19.37±1.35 Negative control 0 0 0 Discussion Based on this in vitro study the researchers found that the ethanolic seed extract of S. cumini had shown highest anti-bacterial activity against selected bacterial strains. T. foenum-graecum seed extract also shown higher antibacterial effect against some bacterial strains. But, S.alba had shown least antibacterial activity. Present study results could be comparable with the previous studies which were stated that the seed phenolic extract of S.cumini showed antibacterial activity against tested bacterial strains (Santos et al., 2020); the methanol fraction of ethanol extract from the seeds of S. cumini was found to have significant antibacterial activity (Yadav et al., 2011; Patoary et al., 2014; Das, Das & Dharani, 2019). However, the present results of S.alba is not in line with another study which was done by the Boscaro et al., 2018. That study found that the S.alba seed hydroalcoholic extract was effective against E. coli and S. aureus in disc diffusion test (Boscaro et al., 2018). The hexane extract of S. alba seeds showed the highest anti-microbial activity (Sujatha & Mariajancyrani, 2013). Sharma et al found that the methanol extract of T. foenum-graecum seeds shown maximum zone of inhibition against E. coli and Staphylococcus (Sharma, Singh & Rani 2017). Another study found that the oil which was extracted from fenugreek seeds has a good antimicrobial activity against some bacteria (Sara & Abdalbasit, 2022). Further another study found that the N. sativa seed oil had a strong antibacterial activity significantly (P<0.01) greater inhibition zone than that of gentamicin (Forouzanfar, Bazzaz & Hosseinzadeh, 2014). Bakathir and Abbas informed that the N. sativa ground seeds possessed antibacterial effect against the staphylococcus (Bakathir & Abbas 2011). Based on all of the representation of antimicrobial activity, it showed that highest antimicrobial activity is showed by S. cumini rather than other extracts, also when consider about phytochemical screening there is highest representation in seeds extracts of S. cumini. Antimicrobial and antibiotic principle are highly showed by phytochemical compounds such as alkaloids, saponins, tannins, flavonoids and steroids which are known to be biologically active (Nethathe & Ndip, 2011; Patra, 2012; Mujeeb, Bajpai & Pathak, 2014; Ali et al. 2018; Pizzi, 2021; Nek Rahimi et al., 2022). Since Gram positive bacteria's cell walls are more permeable than Gram negative bacteria, whose outer membrane has a lipopolysacharide layer that prevents some antibiotics and antibacterial compounds from penetrating, most plant extracts are thought to be more effective against Gram positive bacteria (Wintola & Afolayan, 2015). Among the all-selected medicinal plants, S. cumini showed in high level of antimicrobial activity. Plant produces a range of chemical constituents to protect themselves from the attack of various pathogenic micro-organisms. Substances can either prevent the growth of microorganism or kill them. It can be considered as resources for developing new drugs for various infectious diseases. Antibiotic substances are recognized to vary in concentration in different tissues of same plant, between plants of same and different species and concentration of antibiotics in plant is determined by its environment (Sushil Kumar, Bagchi & Darokar, 1997). In seeds, reserve materials are starch, fixed oils, proteins, fixed oils, fatty acids, some proteins are known to possess good antimicrobial activity and antifungal protein (Cowan, 1999). https://www.frontiersin.org/people/u/924566 https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/antibacterial-activity https://www.cabdirect.org/globalhealth/search/?q=au%3a%22Sujatha%22 https://www.cabdirect.org/globalhealth/search/?q=au%3a%22Mariajancyrani%22 Page 24 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 Conclusion This research on the antimicrobial activity of the herbs helps to develop effective herbal remedies as anti- microbial activity to reduce the infection in diabetes. The result shows that S. cumini seeds extract poses the highest antimicrobial activity in gram positive and negative bacteria. Therefore, this seed is potential to be further developed as an herbal antibiotic for the management of infection in diabetes in future. More detail study such as are fractionation and characterization of active phytochemicals which are responsible for the antimicrobial activity, as well as in-vivo activities recommended to be conducted in the future study. Acknowledgements The authors greatly appreciate the financial support given by the University of Jaffna Research Grant (Grant No. URG/2021/SEIT/27). Conflict of Interest Authors have no conflict of interest. References Akter, N. and Latif, Z.A., (2021). Assessment of depression and its associated factors among patients with type 2 diabetes. Sri Lanka Journal of Diabetes Endocrinology and Metabolism, 11(2): 14–25. DOI: http://doi.org/10.4038/sjdem.v11i2.7448 Aryal B., Niraula P., Khadayat K., Adhikari B., Khatri Chhetri D., Sapkota B. K., Bhattarai B. R., Aryal N., Parajuli N. (2021). Antidiabetic, Antimicrobial, and Molecular Profiling of Selected Medicinal Plants. Evid Based Complement Alternat Med. 5510099. doi: 10.1155/2021/5510099. PMID: 34040646; PMCID: PMC8121587. Ali, S., Khan, M.R., Irfanullah et al. (2018). Phytochemical investigation and antimicrobial appraisal of Parrotiopsis jacquemontiana (Decne) Rehder. BMC Complementary and Alternative Medicine, 18, 43. https://doi.org/10.1186/s12906-018-2114-z Alihosseini, (2016). F. 10 - Plant-based compounds for antimicrobial textiles, Antimicrobial Textiles, In Woodhead Publishing Series in Textiles, 155-195. https://doi.org/10.1016/B978-0-08-100576- 7.00010-9 American Diabetes Association, (2009). Diagnosis and classification of diabetes mellitus. Diabetes Care. 32 Suppl 1(Suppl 1): S62-7. doi: 10.2337/dc09- S062. PMID: 19118289; PMCID: PMC2613584. AOAC. (2016). Official methods of analysis. 20th ed. Association of Analytical Chemists. Arawwawala, L. D. A. M. (2006). Bioactivities of Alpinia calcarata Rosc. Rhizome, Thesis of Master of Philosophy (M’Phil), Biochemistry and Molecular Biology of the Postgraduate Institute of Science, University of Paradeniya, Sri Lanka. Babich O., Larina V., Ivanova S., Tarasov A., Povydysh M., Orlova A., Strugar J and Sukhikh S. (2022). Phytotherapeutic Approaches to the Prevention of Age-Related Changes and the Extension of Active Longevity. Molecules, 27, 2276: 1-32. https://doi.org/10.3390/molecules27072276 Bakathir H. A., Abbas N. A. (2011). Detection of the antibacterial effect of Nigella sativa ground seeds with water. African Journal of Traditional Complementary Alternative Medicine, 8(2): 159- 64. doi: 10.4314/ajtcam. v8i2.63203. Epub 2010 Dec 30. PMID: 22238497; PMCID: PMC3252685. Boscaro V., Boffa L., Binello A., Amisano G., Fornasero S., Cravotto G., Gallicchio M. (2018). Antiproliferative, Proapoptotic, Antioxidant and Antimicrobial Effects of Sinapis nigra L. and Sinapis alba L. Extracts. Molecules. 16; 23(11): 3004. doi: 10.3390/molecules23113004. PMID: 30453590; PMCID: PMC6278512. Buchanan T. A, Xiang A. H, Page K. A. (2012). Gestational diabetes mellitus: risks and management during and after pregnancy. Nature Reviews Endocrinology, 8(11):639-49. doi: 10.1038/nrendo.2012.96. Epub 2012 Jul 3. PMID: 22751341; PMCID: PMC4404707. Cowan MM. (1999). Plant products as antimicrobial agents. Clin Microbiol Rev. 12(4):564-82. doi: 10.1128/CMR.12.4.564. PMID: 10515903; PMCID: PMC88925. Das S, Das A and Dharani N. (2019). Application of Jamun (Syzygium cumini Linn) seed extract on cotton fabric for antibacterial activity. Indian Journal of Fibre & Textile Research, 44: 365-368. Dwivedi C, Pandey I, Pandey H, Ramteke PW, Pandey AC, Mishra SB, Patil S. (2017). Chapter 9 - Electrospun Nanofibrous Scaffold as a Potential Carrier of Antimicrobial Therapeutics for Diabetic Wound Healing and Tissue Regeneration, Nano- and Microscale Drug Delivery Systems, Elsevier: 147-164. https://doi.org/10.1016/B978-0-323- 52727-9.00009-1. Forouzanfar F, Bazzaz BS, Hosseinzadeh H. (2014). Black cumin (Nigella sativa) and its constituent (thymoquinone): a review on antimicrobial effects. Iranian Journal of Basic Medical Sciences, 17(12): 929-38. PMID: 25859296; PMCID: PMC4387228. http://doi.org/10.4038/sjdem.v11i2.7448 https://doi.org/10.1186/s12906-018-2114-z https://doi.org/10.1016/B978-0-08-100576-7.00010-9 https://doi.org/10.1016/B978-0-08-100576-7.00010-9 https://doi.org/10.3390/molecules27072276 https://doi.org/10.1016/B978-0-323-52727-9.00009-1 https://doi.org/10.1016/B978-0-323-52727-9.00009-1 Page 25 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 Global report on diabetes. (2016). 1. Diabetes Mellitus – epidemiology. 2. Diabetes Mellitus – prevention and control. 3. Diabetes, Gestational. 4. Chronic Disease. 5. Public Health. I. World Health Organization. WHO Library Cataloguing-in- Publication Data. http://www.who.int/ Hegazy W. A. H., Rajab A. A. H., Abu Lila A. S., Abbas H. A. (2021). Anti-diabetics and antimicrobials: Harmony of mutual interplay. World J Diabetes. 15; 12(11): 1832-1855. doi: 10.4239/wjd. v12.i11.1832. PMID: 34888011; PMCID: PMC8613656. IDF Diabetes Atlas. (2013). 6th Edition, International Diabetes Federation, file:///C:/Users/shanmuga/Downloads/english- 6th.pdf King A. J. (2012). The use of animal models in diabetes research. British Journal of Pharmacology, 166(3):877-94. doi:10.1111/j.1476- 5381.2012.01911.x. PMID: 22352879; PMCID: PMC3417415. Mostafa A. A., Al-Askar A. A., Almaary K. S., Dawoud T. M., Sholkamy E. N., Bakri M. M. (2018). Antimicrobial activity of some plant extracts against bacterial strains causing food poisoning diseases. Saudi Journal of Biological Sciences, 25(2): 361-366. doi: 10.1016/j.sjbs.2017.02.004. Epub 2017 Feb 24. PMID: 29472791; PMCID: PMC5815983. Mujeeb F, Bajpai P, Pathak N. (2014). Phytochemical evaluation, antimicrobial activity, and determination of bioactive components from leaves of Aegle marmelos. BioMed Research International, 2014:497606. doi: 10.1155/2014/497606. Epub. 11. PMID: 24900969; PMCID: PMC4037574. Nek Rahimi N. N. M., Natrah I., Loh J.Y., Ranzil F. K. E., Gina M., Erin Lim S. H., Lai K. S. and Chong C. M. (2022). Phytocompounds as an Alternative Antimicrobial Approach in Aquaculture, Antibiotics; 11, 469: 1-24. https://doi.org/10.3390/antibiotics11040469 Nethathe B. B., Ndip R. N. (2011). Bioactivity of Hydnora africana on selected bacterial pathogens: Preliminary phytochemical screening. Afr J Microbiol Res. 5(18):2820–6. Pathmanathan, S., Abhayaratna, S.A. (2022). Managing endocrine disorders amidst the financial crisis in Sri Lanka. Sri Lanka Journal of Diabetes Endocrinology and Metabolism, 12(1): 3–4. DOI: http://doi.org/10.4038/sjdem.v12i1.7474 Patoary R., Mondal O. A., Khan A. R and Islam W. Antibacterial Activities of the Seed extracts of Some Indigenous Plants. Journal of Bangladesh Academy of Sciences, 2014; 38 (2): 119-125. Patra A. K. (2012). An Overview of Antimicrobial Properties of Different Classes of Phytochemicals. Dietary Phytochemicals and Microbes. 18:1–32. doi: 10.1007/978-94-007-3926-0_1. PMCID: PMC7121617. Phuyal N., Kumar Jha P., Prasad Raturi P., and Rajbhandary S. (2020). In Vitro Antibacterial Activities of Methanolic Extracts of Fruits, Seeds, and Bark of Zanthoxylum armatum, DC. Hindawi Journal of Tropical Medicine. https://doi.org/10.1155/2020/2803063 Pizzi A. (2021). Tannins medical/pharmacological and related applications: A critical review. Sustainable Chemistry and Pharmacy. 1; 22:100481. https://doi.org/10.1016/j.scp.2021.100481 Rajkumar G, Jayasinghe M. R., and Sanmugarajah V. (2021). Comparative Analytical Study of Phytochemicals in Selected Antidiabetic Medicinal Plant Seeds in Sri Lanka, Journal of Pharmaceutical Sciences and Research, 8(3): 145-155. https://scholarhub.ui.ac.id/psr/vol8/iss3/5 Santos C. A, Almeida F. A, Quecán B. X. V., Pereira P. A. P., Gandra K. M. B., Cunha L. R and Pinto U. M. (2020). Bioactive Properties of Syzygium cumini (L.) Skeels Pulp and Seed Phenolic Extracts. Front. Microbiol, 11: 1-16|. https://doi.org/10.3389/fmicb.2020.00990 Sara T. H and Abdalbasit A. M. (2022). Chapter 9 - Antioxidant and antimicrobial activity of fenugreek (Trigonella foenum-graecum) seed and seed oil. Multiple Biological Activities of Unconventional Seed Oils, Academic Press, 111-117. Sarkar T, Salauddin M, Chakraborty R. (2020). In-depth pharmacological and nutritional properties of bael (Aegle marmelos): A critical review. Journal of Agriculture and Food Research, 2:1-22, 100081. https://doi.org/10.1016/j.jafr.2020.100081 Sharma V, Singh P and Rani A. (2017). Antimicrobial Activity of Trigonella foenum-graecum L. (Fenugreek). European Journal of Experimental Biology, 7(1):1-4. Shehab, M.A., Sultana, N, Jahan, S, Mahmood, T, Imrul- Hasan, M, Hasan, M. and Hasanat, M. (2022). Low level of serum zinc may be associated with early onset diabetes mellitus. Sri Lanka Journal of Diabetes Endocrinology and Metabolism, 12(1): 13–20. http://www.who.int/ file:///C:/Users/shanmuga/Downloads/english-6th.pdf file:///C:/Users/shanmuga/Downloads/english-6th.pdf https://doi.org/10.3390/antibiotics11040469 http://doi.org/10.4038/sjdem.v12i1.7474 https://doi.org/10.1155/2020/2803063 https://doi.org/10.1016/j.scp.2021.100481 https://scholarhub.ui.ac.id/psr/vol8/iss3/5 https://www.frontiersin.org/people/u/924566 https://www.frontiersin.org/people/u/682776 https://www.frontiersin.org/people/u/700672 https://www.frontiersin.org/people/u/972956 https://www.frontiersin.org/people/u/972973 https://www.frontiersin.org/people/u/973205 https://www.frontiersin.org/people/u/422076 https://doi.org/10.3389/fmicb.2020.00990 https://doi.org/10.1016/j.jafr.2020.100081 Page 26 Rajkumar et al. (2023) Journal of Pharmacy, 3(1), 19-26 DOI: http://doi.org/10.4038/sjdem.v12i1.7457 Sujatha R and Mariajancyrani C. (2013). Preliminary phytochemical investigation and antimicrobial activity of Sinapis alba. Scholars Journal of Applied Medical Sciences, 1(3): 138-141. Sushil Kumar, G.D. Bagchi & M.P. Darokar (1997). Antibacterial Activity Observed in the Seeds of Some Coprophilous Plants, International Journal of Pharmacognosy, 35:3, 179-184, DOI: 10.1076/phbi.35.3.179.13293 The World Bank Group. (2022). Diabetes prevalence (% of population ages 20 to 79) – Sri Lanka. https://data.worldbank.org/indicator/SH.STA.DIA B.ZS?locations=LK Unuofin J. O., Lebelo S. L. (2020). Antioxidant Effects and Mechanisms of Medicinal Plants and Their Bioactive Compounds for the Prevention and Treatment of Type 2 Diabetes: An Updated Review. Oxid Med Cell Longev. 13; 2020:1356893. doi: 10.1155/2020/1356893. PMID: 32148647; PMCID: PMC7042557. Wintola O. A., Afolayan A. J. (2015). The antibacterial, phytochemicals and antioxidants evaluation of the root extracts of Hydnora africanaThunb. used as antidysenteric in Eastern Cape Province, South Africa. BMC Complement Altern Med. 4; 15: 307. doi: 10.1186/s12906-015-0835-9. PMID: 26335685; PMCID: PMC4558922. Wu Y, Ding Y, Tanaka Y, Zhang W. (2014). Risk factors contributing to type 2 diabetes and recent advances in the treatment and prevention. International Journal of Medical Sciences, 6;11(11):1185-200. doi: 10.7150/ijms.10001. PMID: 25249787; PMCID: PMC4166864. Yadav S. S., Meshram G. A., Shinde D., Patil R., Manohar S. M., Upadhye M. V. (2011). Antibacterial and Anticancer Activity of Bioactive Fraction of Syzygium cumini L. Seeds, HAYATI Journal of Biosciences, 18 (3): 118-122. https://doi.org/10.4308/hjb.18.3.118 http://doi.org/10.4038/sjdem.v12i1.7457 https://www.cabdirect.org/globalhealth/search/?q=au%3a%22Sujatha%22 https://www.cabdirect.org/globalhealth/search/?q=au%3a%22Ravishankar%22 https://www.cabdirect.org/globalhealth/search/?q=au%3a%22Mariajancyrani%22 https://www.ijbs.com/ https://www.ijbs.com/ https://doi.org/10.4308/hjb.18.3.118