## 1 Introduction  
Severe bacterial infections, particularly those caused by multi-drug-resistant (MDR) strains, are the leading causes of morbidity and mortality in patients admitted to intensive care units (ICU). Amongst them, ventilation-associated pneumonia (VAP) occurs in 9-27% of mechanically ventilated (MV) patients at ICUs and its mortality ranges from 30% to 70% [1]. A noticeably wide range of VAP mortality rates results from numerous factors, inter alia various types of ICU wards, along with the different diagnostic criteria and treatment regimes used there, late-onset infection and the pathogen itself, whereby _Acineto-bacter baumanii, Pseudomonas aeruginosa, Enterobacteriaceae_ (_Klebsiella pneumoniae, Escherichia coli_) and methicillin-resistant _Staphylococcus aureus_ (MRSA) are the most prevalent [2; 3; 4]. Antibiotic-resistant species are a constantly growing threat in nosocomial infections, with _Klebsiella pneumoniae_ being the most prominent example. To de-escalate the problem of resistance, the use of an antimicrobial class and dose adequate for the causative pathogen is recommended [5; 6; 7]. Both piperacillin [8; 9] and impenem [10], with their broad spectrum of action (Gram-positive, Gram-negative and anaerobic bacteria), are a common choice for the empirical and targeted treatment of critically ill patients. However, the effective dosing of \(\beta\)-lactam antibiotics in these patients is challenging due to the substantial pharmacokinetic (PK) variability, resulting in sub-therapeutic concentrations that may lead to therapy failure and even promote resistance. The altered PK of \(\beta\)-lactams may be related to the increased renal clearance or renal replacement therapy, but it may also result from the disease pathophysiology in critically ill patients, as the high intra- and inter-patient variability of piperacillin concentrations were also reported for individuals without severe renal dysfunctions [11]. To maintain the near-maximal antimicrobial effect of \(\beta\)-lactams, therapeutic drug monitoring (TDM) has been proposed in recent years to individualize dosing strategies. However, to ensure optimal treatment, it is essential to consider additional factors. Firstly, the penetrance of drugs into the pulmonary epithelial lining fluid (ELF), reflecting the site of infection in VAP patients, which is influenced by, among others, pH, lipophilicity, alveolar-capillary membrane, and bronchial inflammation. Consequently, even drugs of the same class, for instance, cefepime, piperacillin or ceftroline (all being \(\beta\)-lactams), may have considerably different levels of penetrance in the ELF [12]. Secondly, given that only the unbound fraction of antibiotics is responsible for bacteria eradication, determination of the unbound rather than total drug concentration is valid. Importantly, substantial differences between unbound concentrations that are measured vs. recalculated from total concentrations using published protein-binding values were demonstrated [13]. Finally, the economic aspect needs to be considered. Despite the continuously increasing evidence that TDM is a useful strategy to optimize drug exposure [14] and improve clinical outcome [15; 16], particularly when pathogens with reduced sensitivity to antibiotics are present [5; 17], the poor availability of analytical techniques enabling the TDM of \(\beta\)-lactams, as well as the time until results are available, were reported as frequent obstacles in regular practice [18]. Given the abovementioned reasons, an effective method is being sought for the rapid determination of the dynamically changing concentration profiles of antibiotics in biological material. The applications reported to date are based on high-performance liquid chromatography (HPLC) coupled with ultraviolet (UV) [19; 20; 21] or mass-spectrometry (MS) detection [22; 23; 24; 25; 26]. However, most of them require time-consuming sample preparation and a relatively long run time, which limits their usefulness in daily clinical practice.