## 4 Materials and Methods
### Quantification of Atropine and Scopolamine  
To quantify the analytes, matrix-matched calibration curves were prepared using an internal standard. Two calibration curves, each with seven points at concentrations ranging from 0.1 to 25 ng/mL (in the infusion sample), were prepared using Tea-B-3 and Her-Inf-4. To prepare these curves, 2.5 mL of the filtered sample was taken and spiked with increasing and appropriate concentrations of atropine and scopolamine. Next, a 12.5 \(\mu\)L aliquot of a 500 ng/mL solution containing (\(\pm\))-atropine-D3 and (\(-\))-scopolamine-D3 was added. The protocol described in Section 4.4 was then followed, and the resulting aliquots were injected into the HPLC-MS/MS equipment following the conditions described in Section 4.5.  
Calibration curves were constructed using the ratio of the analyte peak area for the 124.1 \(m/z\) ion for atropine and the 156.0 \(m/z\) ion for scopolamine to the internal standard peak area for the 127.2 \(m/z\) ion for (\(\pm\))-atropine-D3 and the 159.0 \(m/z\) ion for (\(-\))-scopolamine-D3 versus the analyte concentration. All samples were spiked with the same amount of internal standard (2.5 ng/mL) used in the calibration curves before analysis. After analysis, the areas of the identified atropine or scopolamine peaks found in the samples were divided by the area of the internal standard and interpolated onto theinternal standard calibration curves. Each sample was analyzed in triplicate, and the mean and standard deviation of the obtained concentrations were calculated. Concentrations were expressed in ng/mL. The density of Tea-B-3 was 1.0017 g/mL and of Her-Inf-4 was 1.0020 g/mL.  
The following supporting information can be downloaded at [[https://www.mdpi.com/article/10.3390/toxins15060362/s1](https://www.mdpi.com/article/10.3390/toxins15060362/s1)]([https://www.mdpi.com/article/10.3390/toxins15060362/s1](https://www.mdpi.com/article/10.3390/toxins15060362/s1)): Table S1: Description of samples analyzed; Table S2: Parameters of mass spectrometry analysis in positive ionization mode.  
Conceptualization, I.S. and J.S.C.; methodology, L.G.-G., S.M.-Z. and J.A.M.P.; investigation, L.G.-G.; writing--original draft preparation, L.G.-G.; writing--review and editing, I.S., S.M.-Z. and J.S.C.; supervision, I.S., S.M.-Z. and J.A.M.P.; funding acquisition, I.S., J.S.C. and J.A.M.P. All authors have read and agreed to the published version of the manuscript.  
This research was funded by MCIU/AEI/FEDER, UE (project number RTI2018-094558-B-I00 (EVALKALIM)); the Fundacao para a Ciencia e a Tecnologia (FCT) through the CQM Base Fund (UIDB/00674/2020) and the Programmatic Fund (UIDP/00674/2020); and the ARDTIT-Agencia Regional para o Desenvolvimento da Investigacao Tecnologia e Inovacao through the project M1420-01-0145-FEDER-000005--Centro de Quimica da Madeira--CQM+ (Madeira 14-20 Program) and the Project M1420-09-5369-FSE-000001 for the Post-Doctoral fellowship given to [PERSON]. The authors also acknowledge financial support from the Fundacao para a Ciencia e Tecnologia and the Madeira 14-2020 program to the Portuguese Mass Spectrometry Network through the PROEQUIPRAM program (M14-20 M1420-01-0145-FEDER-000008).  
Not applicable.  
The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.