*3.2. The Fluorescence Spectra Profiles of Different Honeys and Syrups*

To aid in the characterization of honey and syrup profile spectra, we acquired published information on fluorescence spectra from previous research on honey and syrup samples [22–26]. Compositionally, honey consists of sugars and water with small amounts of proteins, free amino acids, peptides, minerals, vitamins, organic acids, flavonoids and other phenolic compounds and aroma compounds. These constituents, and especially many of the minority components, exhibit fluorescent properties [30]. For example, a peak

with an excitation of 280 nm and emission of 340 nm can potentially indicate fluorescence from aromatic amino acids present in the honey [22]. A fluorescence peak with an emission of 450 nm and excitation at 250 nm can be attributed to non-flavonoid phenolic compounds in the honey sample [31]. The Maillard reaction products, such as hydroxymethylfurfural and furosine, can have characteristic fluorescence peaks with emission wavelengths ranging from 420 to 470 nm and excitation wavelengths between 340 and 380 nm [22]. We compared the fluorescence emission spectra excitation at 250~380 nm. The results show that all honeys had a fixed apex wavelength at 468 nm, and the apex wavelengths of all syrups were 442 nm. Additionally, the intensities of the FDP showed a visible difference between the honey and syrup samples. Therefore, in this study, fluorescence spectra for each pure honey and pure sugar syrup were examined by excitation at 370 nm and data acquisition from 385 to 800 nm (Figure 2). The fluorescence emission spectra were clearly observable in the 385 nm to 700 nm range, with the FDP appearing at 740 nm. Spectral profiles were visibly different between honey and syrup samples, especially in the presence and size of the FDP.

In comparison to natural honey, the fluorescence emission spectra of the syrups produced a blueshift of about 26 nm. Detailed values of the five syrup types and honeys are shown in Table 1. The apexes of the fluorescence emission spectra from all five honeys were located at 470 nm, while the apexes of the syrups were centered at 450 nm. This shift is potentially attributable to differences in the array of conjugated compounds such as polyphenols, aromatic amino acids, Maillard reaction products and other small molecules present in the samples. Ghosh and colleagues investigated the fluorescence spectroscopic properties of honey and cane sugar syrup and found that all spectra from pure honey samples were characterized by two prominent features: a shoulder around 440 nm and a broad band around 510 nm. In contrast, a single prominent band around 430 nm was characteristic of spectra from cane sugar syrup. In addition, their analysis revealed that the major contributor to cane sugar syrup fluorescence is the reduced form of nicotinamide adenine dinucleotide, while the fluorescence of honey is predominantly caused by flavins [32].


**Table 1.** Apex wavelength and area of emission spectra and FDP area of honeys and syrups.

The areas of the emission spectra were substantially different between the different types of honey, possibly due to plant sources, geographic origins and climatic factors, all of which can influence the content of conjugated compounds. The same feature is true for the syrup spectral data. Furthermore, FDPs were highest at 740 nm for pure honey samples but exhibited a very low intensity in the syrup samples (values listed in Table 1). The FDP area for pure honey was consistently 10-fold greater than that of pure syrup. We propose that this difference between the FDP intensity of pure honey and that of syrup could be exploited to monitor adulteration by all types of syrups.

**Figure 2.** Typical fluorescence spectra of honey and syrup ((**a1**): acacia honey; (**a2**): chaste honey; (**a3**): medlar honey; (**a4**): rape honey; (**a5**): linden honey; (**b1**): maltose syrup; (**b2**): HFCS; (**b3**): rice syrup; (**b4**): beet syrup; (**b5**): cassava syrup).
