Standard methods forApis melliferabeeswax research

Abstract

Due to its multifunctional and complex role in the honey bee colony functioning and health (construction material allowing food storage, brood rearing, thermoregulation, mediation in chemical and mechanical communication, substrate for pathogens, toxins and waste), Apis mellifera beeswax has been widely studied over the last five decades. This is supported by a comprehensive set of scientific reports covering different aspects of beeswax research. In this article, we present an overview of the methods for studying chemical, biological, constructional, and quality aspects of beeswax. We provide a detailed description of the methods for investigating wax scales, comb construction and growth pattern, cell properties, chemical composition of beeswax using different analytical tools, as well as the analytical procedures for provenancing beeswax and beeswax-derived compounds based on the hydrogen isotope ratio (IRMS). Along with classical physico-chemical and sensory analysis, we describe more precise and accurate methods for detection of adulterants in beeswax (GC-MS and FTIR-ATR). Moreover, we present methods for studying the influence of beeswax (comb foundation) adulteration on comb construction. Analytical protocols for determining the pesticide residues using different chromatographic and spectroscopic techniques are also described. As beeswax is an agent of high risk for the transmission of bee diseases, we present methods for detection of pathogens in beeswax. To ensure the reproducibility of experiments and results, we present best practice approaches and detailed protocols for all methods described, as well as their advantages and disadvantages.

Identification and Quantification of Single and Multi‐Adulteration of Beeswax by FTIR‐ATR Spectroscopy

“…Adulteration of beeswax can be detected by numerous methods, which are not standardized, but are usually time‐ and material (reagents)‐consuming and expensive. For measuring the physico‐chemical properties of authentic beeswax, as many as 10 different parameters have to be determined . Not all parameters are useful or reliable for detection of the different adulterants of beeswax .…”

“…The recommended methods for identification and quantification of the beeswax components and adulterants are GC coupled to mass spectrometry (GC‐MS) and/or GC coupled to flame ionization detection (GC‐FID) . Svečnjak et al . published an overview of GC coupled techniques and a detailed description of the methods proposed for standardization of the GC beeswax analysis.…”

“…published an overview of GC coupled techniques and a detailed description of the methods proposed for standardization of the GC beeswax analysis. For detection of beeswax foundations adulterated with paraffin, intermediates or microcrystalline wax, the hydrocarbon content of the beeswax is usually analyzed by GC . However, natural variations in hydrocarbons, due to bee (sub)species or local conditions (e.g., nutrition of the colony), have to be considered .…”

“…adapted the FTIR‐ATR method for routine detection of paraffin and tallow in beeswax. The FTIR‐ATR method was recently reviewed in the “standard methods for A. mellifera beeswax research” and further elaborated in a proposal for standardization …”

Risk assessment of beeswax adulterated with paraffin and/or stearin/stearic acid when used in apiculture and as food (honeycomb)

“…More than 300 individual components have been reported in beeswax from various species of honey bee (Tulloch, 1980;Giumanini et al 1995;Jiménez, et al, 2004Jiménez, et al, , 2006Jiménez, et al, , 2007J Q et al, 2014a, 2014b* ;: ( et al, 2019a). Although their concentrations may vary depending on the honey bee species and their geographical origin, only small differences are observed in the concentration of the individual components and substance classes (EFSA, 2007).…”