New psychoactive substances (NPS) represent an important focus nowadays and are continually produced with minimal structural modifications in order to circumvent the law and increase the difficulty of identifying them. Moreover, since there are a high number of different compounds, it is arduous to develop analytical screening and/or confirmation methods that allow the identification and quantification of these compounds. The aim of this work is to develop and validate a bioanalytical method for detecting new synthetic drugs in biological samples, specifically oral fluid, using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS/MS) with minimal sample pretreatment. Oral fluid samples were simply centrifuged and denaturized with different rapid procedures before injection into the LC-MS/MS system. Calibration curves covered a linear concentration range from LOQ to 100 ng/mL. Validation parameters such as linearity, precision, accuracy, selectivity, matrix effect and thermal stability were evaluated and showed satisfactory results, in accordance with US Food & Drug Administration guidelines. The inter-day analytical bias and imprecision at two levels of quality control (QC) were within ±15% for most compounds. This method was able to identify and calculate the concentration of 10 NPS validated in this biological sample, even in the presence of matrix effect.

Background Acute coronary syndrome (ACS) is the main cause of mortality worldwide and despite the adherence to guidelines it is still burdened by an unacceptable risk for cardiovascular (CV) events recurrence, highlighting the need to identify other than traditional cardiovascular risk factors (CVrF) implicated in atherosclerotic plaque instability. In this regard, psychosocial stress appears to be a crucial player in the development of CV disease. Nevertheless, stress is not easy to standardize and the mechanisms by which it promotes coronary artery disease (CAD) are poorly understood. Materials and Methods We therefore prospectively enrolled patients with ACS, stable coronary artery disease (CAD) undergoing percutaneous coronary intervention (PCI) and subjects presenting traditional CVrF but without established CV disease. Multimodality cortisol assessment, expression of acute and chronic stress, through blood, urine and hair samples collection was ascertained at baseline. A regression analysis was performed to assess the relationships between significant variables at univariate analysis. Results Fifty patients were enrolled in the present study. Cortisol levels in blood and urine were numerically higher in patients with ACS compared to CAD patients and subjects with traditional CVrF only. Hair cortisol levels did not differ between the three groups. The regression analysis showed an inverse correlation (R= -,532, p<0.001 and R=-,615, p<0.001 respectively) between urinary cortisol (UC) and UC/creatinine ratio and left ventricular ejection fraction (LVEF). Conclusion The preliminary results of our study showed that patients with ACS did not have significantly higher levels of hair cortisol compared to stable patients. The finding of an inverse relationship between higher UC, UC/C ratio levels and lower LVEF values support a link between a hyperactivity of the hypothalamic-pituitary-adrenal axis and a worse ACS presentation. These preliminary data will be implemented with serial multimodality assessment of cortisol that allow potential implications in diagnosis and outcome.