Comparative Microbial Profile of Eco-Enzymes Produced from Orange, (Citrus x sinensis Mill.) Pers. and Pineapple, (Ananas comosus L.) Merr. Peels using Metabarcoding of 16S and ITS Regions
Keywords:
16S and ITS regions, Eco-enzyme, Metabarcoding, Orange, Oxford Nanopore Technology, Pineapple
Abstract
Eco-enzyme (EE) is an organic antibacterial substance used as an eco-friendly cleaner. Although EE applications have been widely studied, microbial diversity identification through 16S and ITS metabarcoding remains limited. This study characterized bacterial and fungal biodiversity in orange (Citrus sinensis) and pineapple (Ananas comosus) peel-derived EE using metabarcoding of the 16S and ITS regions. EE samples were prepared using a 1:3:10 ratio of molasses, organic waste, and water, then fermented for 3 months. DNA was extracted, amplified with universal 16S and ITS primers, and confirmed by gel electrophoresis. Sequencing was conducted by the Integrated Genomic Factory (IGF) using Oxford Nanopore Technology (ONT), and data were analyzed in RStudio. Pineapple EE contained 10 bacterial species, while orange EE contained 9 species within the top 10 most abundant species detected (out of a total of 71 species in pineapple EE and 54 species in orange EE).
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References
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Tivani, I., Santoso, J. & Zaidaturriziq. (2025). Efektivitas Waktu Fermentasi Eco Enzym Terhadap Daya Hambat Bakteri Eschericia coli. Parapemikir : Jurnal Ilmiah Farmasi, 14(1), 54–58. https://doi.org/https://ejournal.poltekharber.ac.id/index.php/parapemikir/article/view/8187/pdf
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Xu, Y., Zhi, Y., Wu, Q., Du, R. & Xu, Y. (2017). Zygosaccharomyces bailii is a potential producer of various flavor compounds in Chinese Maotai-flavor liquor fermentation. Frontiers in Microbiology, 8(DEC), 2609. https://doi.org/10.3389/fmicb.2017.02609
Zheng, J., Wittouck, S., Salvetti, E., Franz, C. M. A. P., Harris, H. M. B., Mattarelli, P., O’toole, P. W., Pot, B., Vandamme, P., Walter, J., Watanabe, K., Wuyts, S., Felis, G. E., Gänzle, M. G. & Lebeer, S. (2020). A taxonomic note on the genus Lactobacillus: Description of 23 novel genera, emended description of the genus Lactobacillus Beijerinck 1901, and union of Lactobacillaceae and Leuconostocaceae. International Journal of Systematic and Evolutionary Microbiology, 70(4), 2782–2858. https://doi.org/10.1099/ijsem.0.004107
Brugnoli, M., Cantadori, E., Arena, M. P. & Gullo, M. (2024). Oxidative Fermentation of Glucose and Ethanol in Designed Media and Cooked Grape Must by Acetic Acid Bacteria. Journal of Agriculture and Food Research, 15(3), 101028. https://doi.org/10.1016/j.jafr.2024.101028
Ciani, M., Canonico, L. & Comitini, F. (2022). Improving White Wine Aroma And Structure by Non-Saccharomyces Yeasts. White Wine Technology, 117–130. https://doi.org/10.1016/B978-0-12-823497-6.00020-X
Cooper, C. R. (2011). Yeasts Pathogenic to Humans. In C. Kurtzman, J. W. Fell & T. Boekhout (Eds.), The Yeasts (5th ed., Vol. 1, pp. 9–19). Elsevier. https://doi.org/10.1016/B978-0-444-52149-1.00002-1
De Ligne, L., Vidal-Diez de Ulzurrun, G., Baetens, J. M., Van den Bulcke, J., Van Acker, J. & De Baets, B. (2019). Analysis of Spatio-temporal Fungal Growth Dynamics Under Different Environmental Conditions. IMA Fungus, 10(1). https://doi.org/10.1186/s43008-019-0009-3
Dos Santos, H. R. M., Argolo, C. S., Argôlo-Filho, R. C. & Loguercio, L. L. (2019). A 16S rDNA PCR-based Theoretical to Actual Delta Approach on Culturable Mock Communities Revealed Severe Losses of Diversity Information. BMC Microbiology, 19(1). https://doi.org/10.1186/s12866-019-1446-2
El-Nagar, A., Elzaawely, A. A., Taha, N. A. & Nehela, Y. (2020). The Antifungal Activity of Gallic Acid and Its Derivatives Against Alternaria solani, the Causal Agent of Tomato Early Blight. Agronomy, 10(9). https://doi.org/10.3390/agronomy10091402
Entani, E., Masai, H. & Suzuki, K. I. (1986). Lactobacillus acetotolerans, a New Species from Fermented Vinegar Broth. International Journal of Systematic Bacteriology, 36(4), 544–549. https://doi.org/10.1099/00207713-36-4-544
Fatril, A. E. & Adawiyah, R. (2025). Comprehensive Review of Pichia kudriavzevii: Epidemiology, Biology, and Clinical Manifestation as Parasite in Human Body. Journal of The Indonesian Medical Association, 75(3). https://doi.org/https://doi.org/10.47830/jinma-vol.75.3-2025-1865
Ferreira, O. de S., Chagas-Junior, G. C. A., Chisté, R. C., Martins, L. H. da S., Andrade, E. H. de A., Nascimento, L. D. do & Lopes, A. S. (2022). Saccharomyces cerevisiae and Pichia manshurica from Amazonian Biome Affect the Parameters of Quality and Aromatic Profile of Fermented and Dried Cocoa Beans. Journal of Food Science, 87(9), 4148–4161. https://doi.org/10.1111/1750-3841.16282
García, M., Esteve-Zarzoso, B., Cabellos, J. M. & Arroyo, T. (2018). Advances in the Study of Candida stellata. Fermentation, 4(3). https://doi.org/10.3390/fermentation4030074
García-Alegría, A. M., Anduro-Corona, I., Pérez-Martínez, C. J., Corella-Madueño, M. A. G., Rascón-Durán, M. L. & Astiazaran-Garcia, H. (2020). Quantification of DNA through the NanoDrop Spectrophotometer: Methodological Validation Using Standard Reference Material and Sprague Dawley Rat and Human DNA. International Journal of Analytical Chemistry, 2020(1), 8896738. https://doi.org/10.1155/2020/8896738
Hata, N. N. Y., Surek, M., Sartori, D., Serrato, R. V. & Spinosa, W. A. (2023). Role of Acetic Acid Bacteria in Food and Beverages. Food Technology and Biotechnology, 61(1), 85–103. https://doi.org/10.17113/ftb.61.01.23.7811
Ismail, A. Y., Adhya, I., Hendrayana, Y., Nurdin, N., Nurlaila, A., Herlina, N., Kosasih, D., Dzulfannazhir, F., Sofyan, H. N. & Aminudin, S. (2023). Analysis Organoleptik Eco Enzim From Organic Waste Based on Fruits and Vegetables. Bioma : Jurnal Ilmiah Biologi, 12(2), 103–115. https://doi.org/10.26877/bioma.v12i2.17445
Ismail, A. Y., Nainggolan, M. F. & Nauli, A. P. (2024). Characterizing the Chemical Composition of Eco-Enzymes Derived from Vegetable and Fruit Sources. International Journal of Design and Nature and Ecodynamics, 19(3), 1051–1060. https://doi.org/10.18280/ijdne.190334
Kim, I. J., Jeong, D. & Kim, S. R. (2022). Upstream Processes of Citrus Fruit Waste Biorefinery for Complete Valorization. Bioresource Technology, 362(17), 127776. https://doi.org/10.1016/j.biortech.2022.127776
Koklu, Z. İ., SÜT, P. A., ÜÇÜNCÜOĞLU, E. E., KALAYCI, S., ŞAHİN, F., Koklu, Z. İ., SÜT, P. A., ÜÇÜNCÜOĞLU, E. E., KALAYCI, S. & ŞAHİN, F. (2024). Detection and Characterization of the Pichia manshurica Biofilm on the Traditionally Produced Homemade Apple Vinegar. Turkish Journal of Chemistry, 48(1), 76–84. https://doi.org/10.55730/1300-0527.3640
Liang, X., Yang, X., Sha, N., Wang, J., Qiu, G. & Chang, M. (2025). Application of eDNA Metabarcoding Technology to Monitor the Health of Aquatic Ecosystems. Water, 17(8). https://doi.org/10.3390/w17081109
Muliarta, I. N., Sudita, I. D. N. & Situmeang, Y. P. (2023). The Effect of Eco-Enzyme Spraying on Suwung Landfill Waste, Denpasar, on Changes in Leachate Characteristics. Journal of Environmental Health, 15(1), 56–66. https://doi.org/10.20473/jkl.v15i1.2023.56-66
Nakano, S. & Ebisuya, H. (2016). Physiology of Acetobacter and Komagataeibacter spp.: Acetic Acid Resistance Mechanism in Acetic Acid Fermentation. In K. Matsushita, H. Toyama, N. Tonouchi & A. Okamoto-Kainuma (Eds.), Acetic Acid Bacteria: Ecology and Physiology (Vol. 1, pp. 223–234). Springer, Tokyo. https://doi.org/10.1007/978-4-431-55933-7_10
Novianti, T., Saraswati, H. & Seprianto. (2025). Abundance of Yeast by Metagenomic Analysis of Eco-Enzymes Mangosteen peel. IOP Conference Series: Earth and Environmental Science, 1478(1), 012007. https://doi.org/10.1088/1755-1315/1478/1/012007
Prakoso, H. T., Permatasari, G. W., Dimawarnita, F. & Kalbuadi, D. (2025). Metagenomic Exploration of Eco-enzyme Production: Unveiling Enzymatic Activities and Microbiota Abundance for Sustainable Organic Waste Management. Jurnal Bioteknologi & Biosains Indonesia (JBBI), 12(2), 220–230. https://doi.org/10.55981/jbbi.2025.13247
Ranganathan, A., Yadav, A., Bhagwat, S., Paroha, S., Nigam, S., Pandey, S. & Gupta, A. (2024). Development of Alcohol-Tolerant Yeast Strains and Assessment of their Fermentation Efficiency. INDIAN JOURNAL OF PURE & APPLIED BIOSCIENCES, 12(4), 20–32. https://doi.org/10.18782/2582-2845.9122
Roesma, D. I., Tjong, D. H., Syaifullah, Nofrita, Janra, M. N., Aidil, D. R., Prawira, F. D. L. & Salis, V. M. (2025). DNA Barcoding and eDNA Metabarcoding for Identification Species: A Case Study (West Sumatra): DNA barcoding and eDNA metabarcoding. Journal of Tropical Life Science, 15(1), 161–176. https://doi.org/10.11594/jtls.15.01.16
Soleha, S., maretha, D. E., Saputra, A., Indahsari, S. R., Butar, B. B., Suhendra, A. A., Maharani, M., Harlis, H. & Kapli, H. (2023). Optimization of pH on Enzymatic Activity of Eco-Enzyme Averrhoa bilimbi L. in Plaju District, South Sumatra. Jurnal Biota, 9(2), 72–79. https://doi.org/10.19109/biota.v9i2.16460
Suri, S., Singh, A. & Nema, P. K. (2022). Current Applications of Citrus Fruit Processing Waste: A Scientific Outlook. Applied Food Research, 2(1). https://doi.org/https://doi.org/10.1016/j.afres.2022.100050
Suriani, M., Winarti, S., Arifin, S., Alpian & Firlianty. (2023). Diversity of Decomposer Bacteria in Eco Enzyme Fermentation Process of Organic Materials Using Oxford Nanopore Technology (ONT) and its Effectiveness in Inhibiting E-coli in Fish Pond with Water Mineral Soil (C. Luiz da Silva, Trans.). Revista de Gestão Social e Ambiental - RGSA, 17(8), 1–20. https://doi.org/10.24857/rgsa.v17n8-009
Tamba, W. P., Yanti, F. & Tamba, D. (2025). Jakarta Waste Management Policy and the Capacity Crisis of Bantargebang TPST: An Environmental Justice Review. Renai, 11(1), 14–26. https://doi.org/https://renai-journal.percik.or.id/index.php/renai/article/view/15
Tivani, I., Santoso, J. & Zaidaturriziq. (2025). Efektivitas Waktu Fermentasi Eco Enzym Terhadap Daya Hambat Bakteri Eschericia coli. Parapemikir : Jurnal Ilmiah Farmasi, 14(1), 54–58. https://doi.org/https://ejournal.poltekharber.ac.id/index.php/parapemikir/article/view/8187/pdf
Vidalia, C., Angelina, E., Hans, J., Field, L. H., Santo, N. C. & Rukmini, E. (2023). Eco-enzyme as disinfectant: a systematic literature review. International Journal of Public Health Science (IJPHS), 12(3), 1171–1180. https://doi.org/10.11591/ijphs.v12i3.22131
Wang, N., Zhang, P., Zhou, X., Zheng, J., Ma, Y., Liu, C., Wu, T., Li, H., Wang, X., Wang, H., Zhao, X., Mehmood, M. A. & Zhu, H. (2023). Isolation, Identification, and Characterization of an Acid-Tolerant Pichia kudriavzevii and Exploration of Its Acetic Acid Tolerance Mechanism. Fermentation, 9(6). https://doi.org/10.3390/fermentation9060540
White, T. J., Bruns, T., Lee, S. & Taylor, J. (1990). Amplification and Direct Sequencing of Fungal Ribosomal RNA Genes for Phylogenetics. In M. A. Innis, D. H. Gelfand, J. J. Sninsky & T. J. White (Eds.), PCR Protocols - A Guide to Methods and Applications (pp. 315–322). Academic Press. https://doi.org/10.1016/b978-0-12-372180-8.50042-1
Xu, Y., Zhi, Y., Wu, Q., Du, R. & Xu, Y. (2017). Zygosaccharomyces bailii is a potential producer of various flavor compounds in Chinese Maotai-flavor liquor fermentation. Frontiers in Microbiology, 8(DEC), 2609. https://doi.org/10.3389/fmicb.2017.02609
Zheng, J., Wittouck, S., Salvetti, E., Franz, C. M. A. P., Harris, H. M. B., Mattarelli, P., O’toole, P. W., Pot, B., Vandamme, P., Walter, J., Watanabe, K., Wuyts, S., Felis, G. E., Gänzle, M. G. & Lebeer, S. (2020). A taxonomic note on the genus Lactobacillus: Description of 23 novel genera, emended description of the genus Lactobacillus Beijerinck 1901, and union of Lactobacillaceae and Leuconostocaceae. International Journal of Systematic and Evolutionary Microbiology, 70(4), 2782–2858. https://doi.org/10.1099/ijsem.0.004107
Published
2026-09-30
How to Cite
Pudijanto, A. L. A., & Ilmi, M. (2026). Comparative Microbial Profile of Eco-Enzymes Produced from Orange, (Citrus x sinensis Mill.) Pers. and Pineapple, (Ananas comosus L.) Merr. Peels using Metabarcoding of 16S and ITS Regions. Indonesian Journal of Life Sciences, 8(02), 124-142. https://doi.org/10.54250/ijls.v8i02.329
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Section
Genetics
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