Optimizing Maceration Extraction of Myristicin and Methyl Eugenol from Nutmeg (Myristica fragrans Houtt.): An RSM Box–Behnken Approach

Dewi Murniati; Irmanida Batubara; Utami Dyah Syafitri; Deden Saprudin; Budi Riza Putra

doi:10.14710/jksa.29.3.217-226

DOI: https://doi.org/10.14710/jksa.29.3.217-226

Optimizing Maceration Extraction of Myristicin and Methyl Eugenol from Nutmeg (Myristica fragrans Houtt.): An RSM Box–Behnken Approach

Dewi Murniati^{1, 2}, Irmanida Batubara^{3, 4}

, Utami Dyah Syafitri^{4, 5}

, Deden Saprudin³

, Budi Riza Putra⁶

¹Chemistry Science Doctoral Program, Department of Chemistry, Faculty of Mathematics and Natural Sciences, IPB University, Bogor, Indonesia

²Chemistry Education Study Program, Faculty of Islamic Education and Teacher Training, Syarif Hidayatullah State Islamic University Jakarta, Indonesia

³Department of Chemistry, Faculty of Mathematics and Natural Science, IPB University, Bogor, Indonesia

⁴ Tropical Biopharmaca Research Center, International Research Institute of Food, Nutrition, and Health, IPB University, Bogor, Indonesia

⁵ Study Program of Statistics and Data Science, IPB University, Bogor, Indonesia

⁶ Research Center for Metallurgy, National Research and Innovation Agency (BRIN), South Tangerang, Indonesia

View all affiliations

Received: 25 Nov 2025; Revised: 3 Mar 2026; Accepted: 30 Mar 2026; Published: 22 Apr 2026.

BibTex Citation Data :

@article{JKSA79616,
    author = {Dewi Murniati and Irmanida Batubara and Utami Dyah Syafitri and Deden Saprudin and Budi Riza Putra},
    title = {Optimizing Maceration Extraction of Myristicin and Methyl Eugenol from Nutmeg (Myristica fragrans Houtt.): An RSM Box–Behnken Approach},
    journal = {Jurnal Kimia Sains dan Aplikasi},
  volume = {29},
    number = {3},
    year = {2026},
    keywords = {myristicin; methyl eugenol; Response Surface Methodology; Gas Chromatography-Mass Spectrometry; Design Expert},
    abstract = {Myristicin is the key marker of nutmeg ( Myristica fragrans ) quality, while methyl eugenol is an undesirable compound requiring strict monitoring. This study aimed to optimize the extraction conditions of these compounds from nutmeg seeds and mace as a pretreatment step prior to electrochemical quantification. Optimization was performed using Response Surface Methodology (RSM) with a Box–Behnken Design (BBD), considering extraction time (h), ethanol concentration (%), and the sample-to-solvent ratio (g/mL) as independent variables. The analytes were quantified by Gas Chromatography–Mass Spectrometry (GC-MS), and the experimental data were modeled using Design Expert software. Regression models indicated that ethanol concentration and solvent volume significantly influenced extraction yields, while extraction time had a statistically minimal effect (p-values > 0.05, small effect sizes). Nevertheless, practical differences were observed between matrices: nutmeg seeds reached optimal extraction within 2 hours, whereas mace required 17 hours, likely due to its higher essential oil content and fibrous structure, which slows diffusion kinetics. Thus, although time effects were not statistically significant, extended extraction was practically preferred for mace to ensure adequate recovery. Response definitions were specified as follows: Y1 = myristicin concentration (mg/mL), Y2 = methyl eugenol concentration (mg/mL), and % extraction yield. The optimal extraction conditions were achieved with 90% ethanol and 50 mL solvent, maximizing analyte detectability while maintaining reproducibility across seed and mace extracts. These optimized ethanolic matrices are compatible with subsequent electrochemical quantification, given their aqueous/ethanolic composition and anticipated sensitivity of electrochemical detection.},
   issn = {2597-9914},   pages = {217--226}  doi = {10.14710/jksa.29.3.217-226},
    url = {https://ejournal.undip.ac.id/index.php/ksa/article/view/79616}
}

Citation Format:

Abstract

Myristicin is the key marker of nutmeg (Myristica fragrans) quality, while methyl eugenol is an undesirable compound requiring strict monitoring. This study aimed to optimize the extraction conditions of these compounds from nutmeg seeds and mace as a pretreatment step prior to electrochemical quantification. Optimization was performed using Response Surface Methodology (RSM) with a Box–Behnken Design (BBD), considering extraction time (h), ethanol concentration (%), and the sample-to-solvent ratio (g/mL) as independent variables. The analytes were quantified by Gas Chromatography–Mass Spectrometry (GC-MS), and the experimental data were modeled using Design Expert software. Regression models indicated that ethanol concentration and solvent volume significantly influenced extraction yields, while extraction time had a statistically minimal effect (p-values > 0.05, small effect sizes). Nevertheless, practical differences were observed between matrices: nutmeg seeds reached optimal extraction within 2 hours, whereas mace required 17 hours, likely due to its higher essential oil content and fibrous structure, which slows diffusion kinetics. Thus, although time effects were not statistically significant, extended extraction was practically preferred for mace to ensure adequate recovery. Response definitions were specified as follows: Y1 = myristicin concentration (mg/mL), Y2 = methyl eugenol concentration (mg/mL), and % extraction yield. The optimal extraction conditions were achieved with 90% ethanol and 50 mL solvent, maximizing analyte detectability while maintaining reproducibility across seed and mace extracts. These optimized ethanolic matrices are compatible with subsequent electrochemical quantification, given their aqueous/ethanolic composition and anticipated sensitivity of electrochemical detection.

Fulltext View|Download Email colleagues

Keywords: myristicin; methyl eugenol; Response Surface Methodology; Gas Chromatography-Mass Spectrometry; Design Expert

Funding: Directorate General of Research and Development of the Ministry of Higher Education, Science, and Technology in accordance with the Research Program Implementation Contract for Fiscal Year 2025 under contract 006/C3/DT.05.00/PL/2025

Article Metrics:

Article Info

Section: Research Articles

Language : EN

In Vol 29, No 3 (2026): Volume 29 Issue 3 Year 2026

Most cited articles

Pengaruh Pemerangkapan Enzim Alkalin Fosfatase ke dalam Silika dari Abu Sekam Padi terhadap Aktivitas Enzimatiknya Isolasi Bakteri Termofilik Sumber Air Panas Gedongsongo dengan Media Pengaya Minimal YT (Yeast Tripton) serta Identifikasi Genotipik dan Fenotipik Polimerisasi Eugenol Dengan Katalis Asam Sulfat Pekat Biolubrication Synthesis Made from Used Cooking Oil and Bayah Natural Zeolite Catalyst Uji Aktivitas Antioksidan Senyawa Flavonoid dari Ekstrak Etanol Daun Johar (Senna siamea Lamk) More cited articles

Chia-Jung Lee, Chun-Wei Huang, Lih-Geeng Chen, Ching-Chiung Wang, (+)-Erythro-∆8'-7S,8R-Dihydroxy-3,30,50-Trimethoxy-8-O-40-Neolignan, an Anti-Acne Component in Degreasing Myristica fragrans Houtt, Molecules, 25, 19, (2020), 4563 https://doi.org/10.3390/molecules25194563
Monika Vasantrao Padol, Prashanth Vishwakarma, Arun Suresh Dodamani, Anup Wamanrao Gore, Karina Shyamlal Chachlani, Shradha Pramod Kharkar, Comparative evaluation of nutmeg mouthwash and 0.2% chlorhexidine gluconate mouthwash on halitosis and plaque control: A randomized clinical trial, Journal of Indian Society of Periodontology, 26, 4, (2020), 384-389 https://doi.org/10.4103/jisp.jisp_868_20
Hailan Bao, Qi Muge, Anticancer effect of myristicin on hepatic carcinoma and related molecular mechanism, Pharmaceutical Biology, 59, 1, (2021), 1126-1132 https://doi.org/10.1080/13880209.2021.1961825
Elisa Frederico Seneme, Daiane Carla Dos Santos, Carolina Afonso de Lima, Ícaro Augusto Maccari Zelioli, Juliana Mozer Sciani, Giovanna Barbarini Longato, Effects of Myristicin in Association with Chemotherapies on the Reversal of the Multidrug Resistance (MDR) Mechanism in Cancer, Pharmaceuticals, 15, 10, (2022), 1233 https://doi.org/10.3390/ph15101233
Sawsan S. Al-Rawi, Ahmad Hamdy Ibrahim, Heshu Jalal Ahmed, Zhikal Omar Khudhur, Therapeutic, and pharmacological prospects of nutmeg seed: A comprehensive review for novel drug potential insights, Saudi Pharmaceutical Journal, 32, 6, (2024), 102067 https://doi.org/10.1016/j.jsps.2024.102067
Muhammad Imran, Abdul Haleem Shah, Niamat Ullah, Suliman Yousef Alomar, Abdur Rehman, Naeem Ur Rehman, Asif Nawaz, Rabia Baloch, Ali Zaman, Hafiz Abdul Rafey, Khizar Abbas, Adnan Amin, Integrated computational analysis, in vitro, in vivo investigation on Myristica fragrans Houtt. essential oils for potential anti rheumatic activities, Journal of King Saud University - Science, 36, 5, (2024), 103177 https://doi.org/10.1016/j.jksus.2024.103177
Fuat Karakuş, Zübeyde Tanrıverdi, Burak Kuzu, In Silico Network Toxicology, Molecular Docking, and Multi-Level Bioinformatics Reveal Methyl Eugenol-Induced Hepatocellular Carcinoma Mechanisms in Humans, Cancer Medicine, 14, 10, (2025), 1-11 https://doi.org/10.1002/cam4.70768
Xiaoling Zhang, Susan P. Felter, Anne Marie Api, Kaushal Joshi, Dan Selechnik, A Cautionary tale for using read-across for cancer hazard classification: Case study of isoeugenol and methyl eugenol, Regulatory Toxicology and Pharmacology, 136, (2022), 105280 https://doi.org/10.1016/j.yrtph.2022.105280
Rudi Heryanto, Siti Sa’diah, Waras Nurcholis, Application of two-level factorial design in optimization of maceration extraction of Curcuma longa curcuminoids extracts, Journal of Applied Biology & Biotechnology, 12, 5, (2024), 143-148 https://doi.org/10.7324/JABB.2024.179132
Anthonin Gori, Benjamin Boucherle, Aurélien Rey, Maxime Rome, Nicola Fuzzati, Marine Peuchmaur, Development of an innovative maceration technique to optimize extraction and phase partition of natural products, Fitoterapia, 148, (2021), 104798 https://doi.org/10.1016/j.fitote.2020.104798
Manoj Madanahalli Ramesh, Nischith Sirawase Shankar, Annegowda Hardur Venkatappa, Driving/Critical Factors Considered During Extraction to Obtain Bioactive Enriched Extracts, Pharmacognosy Reviews, 18, 35, (2024), 68-81 https://doi.org/10.5530/phrev.2024.18.7
Alexander Weremfo, Samuel Abassah-Oppong, Felix Adulley, Kwabena Dabie, Salifu Seidu-Larry, Response surface methodology as a tool to optimize the extraction of bioactive compounds from plant sources, Journal of the Science of Food and Agriculture, 103, 1, (2023), 26-36 https://doi.org/10.1002/jsfa.12121
Ivana Karabegoić, Sandra Stamenković Stojanović, B. Danilović, Optimization of food-grade bioactive compound extraction from plant sources by response surface methodology approach: A comprehensive review, Maso International – Journal of Food Science and Technology, 13, 1, (2023), 31-47 https://doi.org/10.2478/mjfst-2023-0004
Syeda Nashvia Adin, Isha Gupta, Mohd Aqil, Mohd Mujeeb, Abdul Ahad, BBD Driven Optimization of Extraction of Therapeutically Active Xanthanoid Mangiferin from Mangifera indica L. Leaves and its Antioxidant Activity, Pharmacognosy Research, 15, 1, (2022), 84-93 https://doi.org/10.5530/097484900279
Vicente Amirpasha Tirado-Kulieva, Manuel Sánchez-Chero, Mario Villegas Yarlequé, Gretell Fiorela Villegas Aguilar, Gilberto Carrión-Barco, Abraham Guillermo Ygnacio Santa Cruz, José Sánchez-Chero, An Overview on the Use of Response Surface Methodology to Model and Optimize Extraction Processes in the Food Industry, Current Research in Nutrition and Food Science, 9, 3, (2021), 745-754 https://doi.org/10.12944/CRNFSJ.9.3.03
Filipa Paulo, Loleny Tavares, Lúcia Santos, Response Surface Modeling and Optimization of the Extraction of Phenolic Antioxidants from Olive Mill Pomace, Molecules, 27, 23, (2022), 8620 https://doi.org/10.3390/molecules27238620
Dara Juliana, Bambang Pontjo Priosoeryanto, Waras Nurcholis, Optimization of Cardamom Fruit (Amomum compactum) Extraction Focused on Total Terpenoid Extraction and Cytotoxic Activity Using Response Surface Methodology, Jurnal Jamu Indonesia, 8, 2, (2023), 57-69 https://doi.org/10.29244/jji.v8i2.328
Irmanida Batubara, Yanico Hadi Prayogo, Stéphane Dumarcay, Christine Gerardin, Philippe Gerardin, Wayan Darmawan, Ika Resmeiliana, Maria Celeste Ruiz, Auliya Ilmiawati, Harlinda Kuspradini, Mohamad Rafi, Phytochemical Profile and In Vitro – In Silico Antibacterial Activity of Melia azedarach Leaf and Twig Extracts Obtained Using Solvents of Different Polarities, Sci, 7, 4, (2025), 167 https://doi.org/10.3390/sci7040167
Irmanida Batubara, Indah Fajarwati, Y. W. Sari, Gilles J. Guillemin, I. Maulidya, Dinda Iryawati, W. Wahyuni, Evaluation of acute and sub-acute toxicity of Curcuma aeruginosa Roxb. essential oil in Sprague Dawley rats, Research in Pharmaceutical Sciences, 20, 3, (2025), 456-468 https://doi.org/10.4103/RPS.RPS_9_24
Saengnapa Champasuri, Arunporn Itharat, Bioactivities of Ethanolic Extracts of Three Parts (Wood, Nutmeg and Mace) from Myristica fragrans Houtt., Journal of the Medical Association of Thailand, 99, (2016), S124-S130
Halid Makić, Samira Hotić, Elvisa Hodžić, Nenad Stojanović, Jasmina Ibrahimpašić, Samira Dedić, Emina Ćehajić Gradinović, Influence of Maceration, Solvent Types, and Extraction Durations on the Yield of Milk Thistle Seeds (Silybum Marianum) Extraction, Technologica Acta, 17, 1, (2024), 67-76 https://doi.org/10.51558/2232-7568.2023.17.1.67
Manh Tuan Ha, Ngoc Khanh Vu, Thu Huong Tran, Jeong Ah Kim, Mi Hee Woo, Byung Sun Min, Phytochemical and pharmacological properties of Myristica fragrans Houtt.: an updated review, Archives of Pharmacal Research, 43, 11, (2020), 1067-1092 https://doi.org/10.1007/s12272-020-01285-4
Mega Ferdina Warsito, A Review on Chemical Composition, Bioactivity, and Toxicity of Myristica fragrans Houtt. Essential Oil, Indonesian Journal of Pharmacy, 32, 3, (2021), 304-313 https://doi.org/10.22146/ijp.1271
Khairan Khairan, Meutia Faradilla, Binawati Ginting, Elly Sufriadi, Adinda Gusti Vonna, Muslim Akmal, Amalia, Ernawati, Indra, Hizir Sofyan, Muhammad Diah, Syaifullah Muhammad, Isolation of Nutmeg Essential Oil (Myristica fragrans houtt) from Aceh Indonesia and Their Antioxidant and Antibacterial Activities, Indonesian Journal of Pharmacy, 35, 3, (2024), 425-436 https://doi.org/10.22146/ijp.8399
Ismiyarto, Ngadiwiyana, Rani Mustika, Isolasi, Identifikasi Minyak Atsiri Fuli Pala (Myristica fragrans) dan Uji Aktivitas Sebagai Larvasida, Jurnal Kimia Sains dan Aplikasi, 12, 1, (2009), 23-30 https://doi.org/10.14710/jksa.12.1.23-30
Nikolaos Nenadis, Maria Papapostolou, Maria Z. Tsimidou, Suggestions on the Contribution of Methyl Eugenol and Eugenol to Bay Laurel (Laurus nobilis L.) Essential Oil Preservative Activity through Radical Scavenging, Molecules, 26, 8, (2021), 2342 https://doi.org/10.3390/molecules26082342
Lidwina Angelica Soetantijo, Hendrik O. Lintang, Heriyanto, Mitha Ayu Pratama Handojo, Tatas Hardo Panintingjati Brotosudarmo, Optimization of Adsorption and Desorption Time in the Extraction of Volatile Compounds in Brewed Java Arabica Coffee Using the HS-SPME/GC-MS Technique, Jurnal Kimia Sains dan Aplikasi, 25, 2, (2022), 49-55 https://doi.org/10.14710/jksa.25.2.49-55
Erik V. Petersson, Jiayin Liu, Per J. R. Sjöberg, Rolf Danielsson, Charlotta Turner, Pressurized Hot Water Extraction of anthocyanins from red onion: A study on extraction and degradation rates, Analytica Chimica Acta, 663, 1, (2010), 27-32 https://doi.org/10.1016/j.aca.2010.01.023
Nashwa F. S. Morsy, A comparative study of nutmeg (Myristica fragrans Houtt.) oleoresins obtained by conventional and green extraction techniques, Journal of Food Science and Technology, 53, 10, (2016), 3770-3777 https://doi.org/10.1007/s13197-016-2363-0
Muliadi, Mudzuna Quraisyah Basimin, Ahmad Muchsin Jayali, Density Functional Theory for QSAR Antioxidant Compound Myristicin Derivatives, Indonesian Journal of Chemical Research, 9, 1, (2021), 49-56 https://doi.org/10.30598//ijcr.2021.9-mul
Salony Azam Sheikh, Vallamkondu Manasa, Palak Daga, Ajay W. Tumaney, Does fixed oil impart more antioxidant potential to spices than essential oils? A comparative study with seed (nutmeg) and aril (mace) oil from the spice myristica fragrans, Journal of Food Science and Technology, 62, 11, (2025), 2173-2182 https://doi.org/10.1007/s13197-024-06164-0
Nicky Rahmana Putra, Ahmad Hazim Abdul Aziz, Hasmadi Mamat, Dwila Nur Rizkiyah, Mohd Azizi Che Yunus, Irianto Irianto, Lailatul Qomariyah, Green extraction of nutmeg (Myristica fragrans) phytochemicals: Prospective strategies and roadblocks, Open Agriculture, 9, 1, (2024), 20220285 https://doi.org/10.1515/opag-2022-0285

Last update:

No citation recorded.

Last update: 2026-09-13 22:28:29

No citation recorded.

As an article writer, the author has the right to use their article for various purposes, including use by institutions that employ the author or institutions that provide funding for the research. These author rights are granted without special permission.

Authors who publish a paper in JKSA have broad rights to use their work for teaching and scientific purposes without the need to request permission, including: (i) teaching in the author's class or institution; (ii) presenting the work at meetings or conferences and distributing copies to participants; (iii) using the work in training conducted by the author or the author's institution; (iv) distributing copies to colleagues for research purposes; (v) using the work in the compilation of subsequent works by the author; (vi) including the work in a thesis or dissertation; (vii) reusing part of the article in another work, with appropriate citation; (viii) preparing derivative works, with appropriate citation; and (ix) voluntarily posting the work on open websites operated by the author or the author's institution for scientific purposes, in accordance with the CC BY-SA License.

Authors and readers may copy and redistribute the material in any medium or format, and may remix, transform, and build upon the material for any purpose, provided that they give appropriate credit by citing the article or its content, provide a link to the license, and indicate if changes were made.

The authors submitting a manuscript do so on the understanding that, if accepted for publication, the copyright of the article shall be assigned to Jurnal Kimia Sains dan Aplikasi (JKSA). Copyright encompasses the rights to reproduce and distribute the article in all forms and media, including reprints, photographs, microfilms, and any other similar reproductions, as well as translations.

Reproducing any part of this journal, storing it in a database, or transmitting it in any form or medium is permitted without written permission from JKSA. However, the original source must be properly cited in accordance with academic conventions.

JKSA, the Chemistry Department of Diponegoro University, and the Editor make every effort to ensure that no inaccurate, false, or misleading data, opinions, or statements are published in JKSA. However, the content of each article is the sole and exclusive responsibility of its authors.

The Copyright Transfer Form can be downloaded here: [Copyright Transfer Form - Indonesian] [Copyright Transfer Form - English]. The copyright form should bear the authors' original signatures and be sent to the Editor as a printed document, a scanned document sent via email, or by fax.

Prof. Adi Darmawan, Ph.D (Editor in Chief)

Editor in chief of Jurnal Kimia Sains dan Aplikasi (JKSA)

Chemistry Department, Faculty of Sciences and Mathematics, Diponegoro University

Jl. Prof. Soedarto, Kampus Undip Tembalang, Semarang, Central Java, Indonesia 50275
Email: jksa@live.undip.ac.id