Evaluating Extraction Techniques for Quantifying Residual Antibiotics Found in Honey using Triple Quadrupole Mass Spectrometry

Honey is a widely sought-after natural product produced around the globe. It’s produced by honeybees that take up nectar from flowers and digest it in a separate stomach full of enzymes necessary to break down nectar sugar. The partially digested nectar is passed from honeybee to honeybee until its placed inside a honeycomb and dried via flapping wings. The honey is then sealed off with beeswax. Extracting honey from beehives has had the same method for years and has been done the same way to ensure bee safety. However, throughout history, beekeepers have noticed that diseases like European foulbrood and American foulbrood have caused severe issues with bee colony sustainability. These diseases are caused by bacteria (Melissococcus plutonius and Paenibacillus larvae respectively), and result in serious health issues or death for larvae. To prevent dying colonies, a wide range of drugs, including sulfonamides, have been used to treat bee colonies from falling to foulbrood diseases. When sulfa drugs are administered, the honey-making process results in the transfer of sulfonamides from bees to curing nectar which eventually becomes honey. Antibiotic treatments around the world have either been banned or have a set maximum residue limit (MRL) to prevent consumer harm.¹ Too much exposure to sulfonamide residues has been linked to changes in the human microbiome, increased resistance to antibiotics, allergic reactions, and more. To combat the use of sulfonamides, several processes have been developed to analyze honey extracts for antibiotic residues using triple quadrupole selected or multi-reaction monitoring (SRM or MRM) experiments. The challenge in each process lies in the extraction, as honey contains numerous small molecules like proteins, enzymes, organic acids, vitamins, flavonoids, and more. Three extraction processes that have emerged as the primary means of extraction are solid-phase, liquid-liquid, and QuEChERS (quick, easy, cheap, effective, rugged, and safe).^2-4 I will be comparing the effectiveness of each method used to isolate sulfonamides found in honey samples along with additional steps for analyte selectivity. The shortcomings and strong suits of each method will be discussed according to sulfonamide recovery, method complexity, cost, and detectability.

1. Zhang, Y.;  Li, X. Q.;  Li, H. M.;  Zhang, Q. H.;  Gao, Y.; Li, X. J., Antibiotic residues in honey: A review on analytical methods by liquid chromatography tandem mass spectrometry. TrAC Trends in Analytical Chemistry 2019, 110, 344-356.

2. El Hawari, K.;  Mokh, S.;  Doumyati, S.;  Al Iskandarani, M.; Verdon, E., Development and validation of a multiclass method for the determination of antibiotic residues in honey using liquid chromatography-tandem mass spectrometry. Food Additives & Contaminants: Part A 2017, 34 (4), 582-597.

3.  Dluhošová, S.;  Borkovcová, I.;  Kaniová, L.; Vorlová, L., Sulfonamide Residues: Honey Quality in the Czech Market. Journal of Food Quality 2018, 2018, 1-7.

4.  Juan-Borrás, M.;  Periche, A.;  Domenech, E.; Escriche, I., Routine quality control in honey packaging companies as a key to guarantee consumer safety. The case of the presence of sulfonamides analyzed with LC-MS-MS. Food Control 2015, 50, 243-249.