The analytical results for the basic 31 elements and 18 rare earth elements (REEs) in collected honey samples (n = 30) are displayed in Table 1 and Table 2.
Levels for each targeted element (mg/kg or µg/kg) includes mean ± SD, and range (minimum-maximum). Also, the elements were categorized to local and imported, and the level of each element in its category is reported in Table 1 and Table 2, as mean ± SD. The basic elements were classified to minor elements, n = 5, (Na, Mg, P, K, and Ca), essential trace elements, n = 9, (Cr, Mn, Fe, Co, Ni, Cu, Zn, Se, and Mo), non-essential elements (n = 12), (Li, Be, Al, V, Ga, Sr, Ag, Sn, Cs, Ba, Bi, and U), and potentially toxic elements (n = 5), (As, Cd, In, Tl and Pb).
Minor elements
Minor elements, Na, Mg, P, K, and Ca represent more than 90 % of total elemental content in most foods including honey, and their levels are related to their presence in soil, fertilizer and irrigation water. Their decreasing order in honey was K > > Ca > P > Na > Mg. The same order holds for imported honey samples, however, in local samples Na is larger than P. Large variations in elemental levels between samples is attributed to botanic and geographic origins. Large levels for ∑minor were observed for Black forest mono-floral honey samples, and small levels were observed for Acasia mono-floral honey samples. Both of them are imported from European countries. P-values (p > 0.05) indicate no significant difference between levels of minor elements in local and imported honey. However, a smaller p-value for Na levels between local and imported samples is due to significantly large salinity of irrigation water in Jordan (Na is around 1000 mg/L).
Essential trace elements
Levels of essential trace elements in honey samples varied between 19.4 and 0.011 mg/kg for Fe and Co, and were found in the decreasing order: Fe > > Mn > Zn > Cu > Cr > Ni > Se > Mo > Co.
Iron, Fe, is an element required for essential functions in cells related to oxygen transport, oxidase activities and energy metabolism (Squadrone, Brizio, Stella, Mantia, et al. 2020)Levels of Fe, varied between 7.03 mg/kg for an imported multi-floral sample and 49.3 mg/kg for a local multi-floral sample. No significant difference was observed for Fe mean levels between local and imported samples.
Manganese, Mn, is a part of metallo-enzymes involved in various metabolisms (Squadrone, Brizio, Stella, Mantia, et al. 2020). Mn levels in honey varied between 0.13 mg/kg for an imported mono-floral sample (Acasia) and 13.3 mg/kg for an imported Black forest mono-floral sample. There is relatively significant difference in levels of Mn between local and imported samples.
Zinc, Zn, is elaborated in various physiological functions and is ubiquitous in the body (Jurowski et al. 2014). Zn levels vary between 0.48 mg/kg for an imported Acasia mono-floral sample and 3.72 mg/kg for an imported multifloral sample. There is no significant difference between levels of Zn in local and imported samples.
Copper, Cu, is a part of several physiological processes in human body. Cu levels varied between 0.145 mg/kg for a local multi-floral sample and 2.34 mg/kg for another local multi-floral sample. There is no significant difference between levels of Cu in local and imported samples.
Chromium, Cr, in trivalent form is involved in metabolism of carbohydrates. Levels of Cr varied between 0.06 mg/kg for a local multi-floral sample and 0.31 mg/kg for another local multi-floral sample. No significant difference was observed between levels of Cr in local and imported samples.
Other essential elements Ni, Se, Mo, and Co were detected with lower levels and had less impact in nutrition or safety assessment.
Nonessential trace elements
Among the 12 targeted nonessential elements, Al, Sr, Ba and Sn had significant levels, and was found in the decreasing order: Al > > Sr > Ba > Sn.
Aluminum, Al, was detectable in all samples ranging from 2.67 mg/kg for an imported Acasia mono-floral sample to 21.9 mg/kg for an imported multi-floral sample. Mean levels of Al in local and imported samples were significantly similar.
Strontium, Sr, was measured in significant levels in all samples ranging from 0.075 mg/kg for an imported Acasia mono-floral sample to 1.62 mg/kg for an imported multi-floral sample. Mean Sr levels between local and imported samples are significantly different. Sr could be employed as elemental marker to distinguish between local and imported honey.
Barium, Ba, levels vary between 0.059 mg/kg for an imported multi-floral sample and 1.06 mg/kg for a Black forest mono-floral sample. There is no significant difference in mean Ba levels between local and imported samples.
Tin, Sn, was found at low levels in honey, with lowest concentration, 0.033 mg/kg, for an imported Black forest mono-floral honey sample, and a highest concentration, 0.186 mg/kg, for a local multi-floral sample. Sn levels in local samples are slightly higher than imported samples.
Potentially toxic trace elements
Thallium, Tl, and indium, In, had lower levels to be considered for toxicity and assessment. Arsenic, As, is a highly toxic element. Its levels were not detected (ND) in 50 % of the samples and was up to 6.9 µg/kg in an imported multi-floral sample. Cadmium, Cd, and lead, Pb, are common pollutants that can enter the food chain after contamination of flowering plants. Cd levels were low in most samples and reached 6.7 µg/kg in an imported Black forest mono-floral sample. Pb levels was low in most samples and reached 31.5 µg/kg in the same imported Black forest mono-floral sample. Levels of Cd and Pb as well As are much lower than regulated levels in food (Paz 2017)
Rare earth elements
REEs are concentrated in different geological environments and their presence in honey is related to geographic origin and geochemical soil composition. Table 2 presents levels of 17 REE. Significant levels were detected for La, Ce, Pr, Nd, Sm, and Th with a decreasing order: Ce > Nd > La > Th > Pr > Sm. Total REE levels vary between 41 µg/kg for an imported mono-floral sample and 583 µg/kg for a local multi-floral sample with a mean of 145 µg/kg and SD of 157 µg/kg. Local samples had significantly larger mean than imported samples. Significantly larger deviations, mainly in imported samples, were due to the difference in geographic origins between imported countries.
Safety assessment
Honey is a popular sweetener in Jordanian diets. An adult usually consumes 10–50 g daily. Thus, honey may contribute significantly for total daily intakes of elements. Table 3 presents levels of estimated daily intake (EDI), recommended daily intake (RDI) (Paz 2017), and percentage contribution of each metal. The percentage contributions of Na, Mg, P, K, Ca, Mn and Zn are negligible relative tor respective EDI (< 1.0 %). Little percentage contributions were observed for the rest of essential elements. Levels of potentially toxic elements (As, Cd, and Pb) were negligible to their RDI values, indicating the safety of Jordanian honey for human consumption.
Assessment and Authentication
Botanic and geographic origins are usually considered in assessment and authentication of honey (Gallmann 2007; Salvador et al. 2019; Squadrone, Brizio, Stella, Mantia, et al. 2020; Squadrone, Brizio, Stella, Pederiva, et al. 2020; Zhou et al. 2018). Results of Tables 1 and 2 indicated that classification of samples by local and imported did not give a clear picture of authentication. Calculated ANOVA p-values were, p > 0.05, indicating no significant difference between local and imported honey, even, levels of Na, Mn, and Sr were significantly different. A closer look to imported samples, they were consisted of Black forest mono-floral samples (n = 5), Acasia mono-floral samples (n = 5), and multi-floral samples (n = 8). All local samples were multi-floral (n = 12). Thus, botanic origin was considered in assessment and authentication. Levels of selected elements (mean ± SD) are presented in Table 4 and graphically in Fig. 1(A-E).
From the first look, precisions (%RSD) of results were much smaller than those in Tables 1 and 2, especially for mono-floral samples. Black forest samples have the highest ∑minor elements, while Acasia samples were the lowest. While it was easier to discriminate between mono-floral and multi-floral, it needs more attention to discriminate between multi-floral-local and multi-floral-imported samples. Sodium, Na, strontium, Sr, and ∑REE were significantly larger in multi-floral-local samples. These elements may be considered markers for multi-floral-local samples.
Pearson’s correlation coefficients between targeted elements themselves in honey samples were computed and are presented in Table 5.
In this study we combined Black forest and Acasia samples in one category under the title “mono-floral”. The other two categories were multi-floral-imported and multi-floral-local. Rare earth elements in the three categories were positively correlated with each other (1.00 > r > 0.80) and negatively correlated with other elements, thus, they were not considered in this study.
Selected correlations were based on categories with positive correlation coefficients (r > 0.50). Among the 60 selected correlations, largest positive correlation coefficients were distributed as 22 for multi-floral-local, 11 for multi-floral-imported, and 27 for mono-floral. Also, negative correlations were observed as 13 for multi-floral-local, 4 for multi-floral-imported, and 6 for mono-floral. The correlations Na/Mg, Na/Se, Mg/Se, Cr/Fe, Cr/Co, Cr/Ni, Fe/Ni, and Al/Sr were distinctive for multi-floral-local. The correlations K/Fe, K/Cu, ∑minor/Cu, Fe/Cu, Co/Mo and Se/Sn were distinctive for multi-floral-imported.
Comparison with previous studies
Various studies were reported for multielemental measurements (minor and trace) in honey from popular honey producing countries. A summary of these results were reported by P. Phohl et al (Marcovecchio et al. 2015). Our results were within the lower half of reported ranges. RREs in honey from Italy, Balkans, Kazakhstan, South America and Tanzania were reported by S. Squadrone et al. They reported ∑REE in the range of 9.0–65 µg/kg. Our results were 67 µg/kg for Acasia, 79 µg/kg for Black forest, 128 µg/kg for multifloral-imported and 384 µg/g for multifloral-local. A mean of 55 mg/kg ∑REE were reported in soil from Jordan.