Journal of Applied Glycoscience
Online ISSN : 1880-7291
Print ISSN : 1344-7882
ISSN-L : 1344-7882
Regular Paper
Development of an LC-MS/MS Method for Quantitation of Western Honeybee (Apis mellifera) α-Glucosidase III as a Potential Honey Authenticity Marker
Yushi TakahashiIzumi YoshidaToshiaki YokozekiYoshinari HirakawaKazuhiro Fujita
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Supplementary material

2025 Volume 72 Issue 2 Article ID: 7202106

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Abstract

Western honeybee (Apis mellifera) α-glucosidase III (HBG-III), which is secreted from the hypopharyngeal glands of honeybees, plays a role in converting nectar into honey. Consequently, hypothesizing that HBG-III is a suitable marker of honey authenticity, we developed an analytical method to determine the HBG-III content and investigated its applicability to various commercial products. Following extraction from honey using phosphate-buffered saline, HBG-III was concentrated using an ultrafiltration membrane and subsequently fragmented with trypsin and lysyl endopeptidase mixture. The specific peptide fragments were used for quantitation by liquid chromatography-tandem mass spectrometry. The established method was validated for linearity, accuracy, precision, and the limit of quantitation (LOQ). As a result, the calibration curve was linear in the range of 0.01-0.3 μM, the mean recovery ranged from 73.8 to 89.2 %, the within-laboratory reproducibility (RSDwr) ranged from 3.9 to 6.5 %, and the LOQ was 1.9 mg/kg. An investigation of HBG-III concentrations in 65 honey products available on the Japanese market revealed that the HBG-III content of 15 low-priced honey products was below the LOQ. This suggested that these products may be adulterated with non-honey syrups. Therefore, this method can serve as an effective tool to verify the authenticity of honey products.

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© 2025 by The Japanese Society of Applied Glycoscience

This is an open-access paper distributed under the terms of the Creative Commons Attribution Non-Commercial (by-nc) License (CC-BY-NC4.0).
https://creativecommons.org/licenses/by-nc/4.0/
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