Malays. J. Anal. Sci. Volume 29 Number 1 (2025): 1154

Research Article

¹H-NMR metabolomics and molecular networking reveal relationship between metabolite profile and antioxidant activity of Malaysian stingless bee honey

Kok Suet Cheng¹, Ahmad Nazif Aziz¹, Nurul Huda Abdul Wahab¹, Desy Fitrya Syamsumir², R. Rudiyanto³, Wan Iryani Wan Ismail¹, Faridah Abas⁴ and M. Maulidiani¹*

¹Faculty of Science and Marine Environment, Universiti Malaysia Terengganu, 21030 Kuala Nerus, Terengganu, Malaysia

²Institute of Climate Adaptation and Marine Biotechnology, Universiti Malaysia Terengganu, 21030 Kuala Nerus, Terengganu, Malaysia

³Faculty of Fisheries and Food Science, Universiti Malaysia Terengganu, 21030 Kuala Nerus, Terengganu, Malaysia

⁴Department of Food Science, Faculty of Food Science and Technology, Universiti Putra Malaysia, Serdang 43400, Selangor, Malaysia

*Corresponding author: maulidani@umt.edu.my

Received: 28 May 2024; Revised: 27 October 2024; Accepted: 28 October 2024; Published: 12 February 2025

Abstract

This study revealed the relationship between metabolite profiling and the antioxidant activity of honey from Malaysian stingless bee species (Heterotrigona itama, Tetrigona apicalis, Geniotrigona thoracica, Tetrigona binghami and Lophotrigona canifrons) using the proton nuclear magnetic resonance (¹H-NMR) metabolomics approach. To identify the metabolites associated with antioxidant activity, the partial least squares (PLS) model was utilised. Meanwhile, the principal component analysis (PCA) was employed to discriminate stingless bee honey samples based on their species. The results revealed that H. itama samples were clearly discriminated from other species. G. thoracica and L. canifrons had similar chemical characteristics, whereas T. binghami and T. apicalis shared their similarities. A total of 32 metabolites were identified, and amongst them, amino acids (glutamic acid, glutamine, leucine, alanine, valine, isoleucine and tyrosine), as well as organic acids (methylmalonic acid and citric acid), positively contributed to the antioxidant activity, as indicated by the PLS biplot. According to the PLS biplot, phenolic compounds are also associated with its antioxidant activity. This study found that T. apicalis honey exhibited the highest potential as an antioxidant agent with 2,2-diphenyl-1-picrylhydrazyl (DPPH) inhibitory activity of 30 mg/mL was 73.36 ± 6.47%. Through liquid chromatography-mass spectrometry (LC-MS) and molecular networking approach, a comprehensive analysis of phenolics and flavonoids of T. apicalis was successfully conducted. The identified flavonoids included quercetin, naringenin, kaempferol, genistein, apigenin, hesperetin, isorhamnetin, quercitrin and flavonoid glucosides (such as kaempferol-5-methyl ether 3-galactoside-4'-glucoside and kaempferol-7-O-glucoside), whereas caffeoylquinic acid isomers were the identified phenolic compounds. The methods used in this study are useful in assessing the quality of stingless bee honey that possesses antioxidant activity.

Keywords: metabolite profile, antioxidant activity, stingless bee honey, ¹H-NMR, phenolic compounds

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