Additional file 1 of Assessment of geographical origin and production period of royal jelly by NMR metabolomics
Creators: name: Mazzei, Pierluigi
name: Piccolo, Alessandro
name: Brescia, Marianna
name: Caprio, Emilio
Publisher: name: figshare
Year: 2020
Subjects: subject: Biochemistry
subject: 59999 Environmental Sciences not elsewhere classified
subject: FOS: Earth and related environmental sciences
subject: FOS: Earth and related environmental sciences
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: FOS: Chemical sciences
subject: Ecology
subject: FOS: Biological sciences
subject: FOS: Biological sciences

description: Additional file 1: Fig. S1. Pollen traps fastened at the hive entrance and enabling the pollen stripping and collection. The pollen accumulates in a side opening drawer when the bees, during their access into the hive, walk through the metallic mesh which connects the pollen trap to the landing board of the hive. The trap is composed of round shaped holes that strip the pollen grains from either the pollen basket or the corbicula on outside of the tibia on the hind legs of the bees. Fig. S2. 1H-NMR spectrum of a representative Italian RJ dissolved in phosphate buffer solution (0.2M, pH 7.3) prepared with d2O and containing TMSPA (d = 0 ppm) as internal standard. The proton acquisition was carried out by applying the presaturation technique to suppress the residual water signal at ≈ 4.704 ppm. Fig. S3. 2D 1H-1H NMR COSY spectrum of a representative Italian RJ dissolved in phosphate buffer solution (0.2M, pH 7.3) prepared with d2O and containing TMSPA (d = 0 ppm) as internal standard. The acquisition was carried out by applying the presaturation technique to suppress the residual water signal at ≈ 4.704 ppm. Fig. S4. Superimposition of 2D 1H-13C NMR HSQC (red) and HMBC (blue) spectra of a representative Italian RJ dissolved in phosphate buffer solution (0.2M, pH 7.3) prepared with d2O and containing TMSPA as internal standard. Fig. S5. Significant loading vectors related to the PCA score-plot shown in Fig. 3 and based on 1H NMR spectra of Royal Jelly samples produced from two Bologna (Bo1 and Bo6) and four Grosseto (Gr2, Gr3, Gr4 and Gr5) farms on the 30th of May 2016. Fig. S6. PLS-DA results based on 1H NMR spectra of Royal Jelly samples produced from Bologna (squares) and Grosseto (circles) farms on the 30th of May 2016. (A) PLS-DA Cross-validation score-plot, where full and holed objects differentiate samples belonging to either the training or the test set, respectively. (B) PLS-DA classification results and Q2/R2 values related to the PLS-DA components 1 and 2; (C) PLS-DA cross-validation results.Fig. S7. PLS-DA results based on 1H NMR spectra of Royal Jelly samples produced from the two Bologna farms Bo1 (blue squares) and Bo6 (green squares) on the 30th of May 2016. (A) PLS-DA Cross-validation score-plot, where full and holed squares differentiate samples belonging to either the training or the test set, respectively. (B) PLS-DA classification results and Q2/R2 values related to the PLS-DA components 1 and 2; (C) PLS-DA cross-validation results. Fig. S8. PLS-DA results based on 1H NMR spectra of Royal Jelly samples produced from the four Grosseto farms Gr2, Gr3, Gr4 and GR5 (light green, dark green, orange and red circles, respectively) on the 30th of May 2016. (A) PLS-DA Cross-validation score-plot, where full and holed circles differentiate samples belonging to either the training or the test set, respectively. (B) PLS-DA classification results and Q2/R2 values related to the PLS-DA components 1 and 2; (C) PLS-DA cross-validation results. Fig. S9. Significant loading vectors related to the PCA score-plot shown in Fig. 4 and based on 1H NMR spectra of Royal Jelly samples of Italian (produced the 8th of July 2016) and Chinese (distributed in 2016) geographical origin. Fig. S10. PLS-DA results based on 1H NMR spectra of Royal Jelly samples of Italian and Chinese geographical origin. (A) PLS-DA Cross-validation score-plot, where full and holed objects differentiate samples belonging to either the training or the test set, respectively. (B) PLS-DA classification results and Q2/R2 values related to the PLS-DA component 1; (C) PLS-DA cross-validation results. Fig. S11. Significant loading vectors related to the PCA score-plots shown in Fig. 5A (A), 5B (B) and 5C (C) and based on 1H NMR spectra of Royal Jelly samples produced by three Grosseto farms in different days (see the legend). The label 3-hydroxy correspond to 3-hydroxybutyrate. Fig. S12. Significant loading vectors related to the PCA score-plots shown in Fig. 6 and based on 1H NMR spectra of Royal Jelly samples produced by three Italian farms (Bo1, Bo6 and Gr5) on the 13th of June and deriving from a monofloral production conducted in the presence of either chestnut, trifolium or linden flowerings. Table S1. The table reports the name and the most relevant information related to the Italian farms involved in the project. Table S2. Results of ANOVA test (Benjamini-Hochberg’s test, a = 0.05) informing on the significance of the most discriminant variables associated to the PCs of score-plots shown in Figs. 3 and 4, respectively, and explaining a total variance