Background: Implant-related infection following femoral fracture surgery is a severe complication in elderly patients and is associated with high morbidity and mortality. Most available evidence on periprosthetic joint infection (PJI) derives from elective arthroplasty populations, which differ substantially from patients undergoing surgery for femoral fractures. This study aimed to investigate the microbiological profile and clinical characteristics of implant-related infections after proximal femoral fracture surgery. Materials and Methods: A retrospective observational study was conducted on 20 patients aged ≥70 years who developed implant-related infection after surgical treatment of proximal femoral fractures between 2020 and 2025 at a referral trauma center. Surgical procedures included intramedullary nailing, hemiarthroplasty, and total hip arthroplasty. Only patients with Charlson Comorbidity Index ≥ 4 and infection occurring within one year of the index surgery were included. Clinical, surgical, microbiological, and antibiotic therapy data were retrospectively reviewed. Results: The cohort had a mean age of 82.4 years and a high comorbidity burden (mean Charlson index 4.8). The most frequently isolated pathogen was Staphylococcus aureus (25.9%), with 85% methicillin-resistant strains. Other pathogens included Enterococcus faecalis, Klebsiella pneumoniae, and Escherichia coli. Polymicrobial infections were observed in 25% of patients. One-year mortality was 25%. Conclusions: Implant-related infections after femoral fracture surgery represent a distinct clinical entity compared with elective PJI, characterized by frail patients and a higher prevalence of multidrug-resistant organisms. These findings highlight the need for tailored preventive and therapeutic strategies in this high-risk population.

Abstract:Chiropractic vertebral manipulation is indicated in patients with acute and uncomplicated low back pain. Nowadays, many people seek care for low back pain from manual practitioners. However, within the overall legislative framework regulating in Italy professionals practicing in healthcare, chiropractors remain unevenly regulated. In fact, they are trained abroad and hold internationally recognized degrees that are not regulated under Italian healthcare professionals' legislation. Neither the educational profile, nor a Ministry technical expert committee, nor a national board for ongoing professional evaluation has been established, despite a law in 2007 aimed to regulate the profession and define the national register of doctors in chiropractic. Meanwhile, given their involvement in the healthcare sphere as part of multiprofessional teams, issues of liability arise, made more complex by the fact that chiropractors are not eligible for certain more favourable legal measures for liability assessment granted to healthcare professionals under Italian Law on healthcare safety and liability. Moreover, health-related information for obtaining informed consent cannot be managed independently by these practitioners, again due to their non-healthcare professional status. As a result, prescribing physicians must necessarily be involved in the overall care process. The interaction between healthcare and non-healthcare professionals presents complexities in the patient care and in the proper liability assessment. These aspects are also urgently needed for the definitive recognition and valorisation of the practitioners working in health sphere, which must include State oversight.

Analytical toxicology is a discipline of forensic toxicology which applies analytical techniques for the determination of drugs of abuse in biological and nonbiological matrices. To this concern, artificial intelligence (AI), particularly machine learning (ML), is innovating analytical toxicology by improving data processing and facilitating the identification of New Psychoactive Substances (NPS). The aim of this review was to explore the current application of AI in this field and to highlight the future perspectives. A literature search was performed in several scientific databases to review articles reporting the implementation of AI models for analytical toxicological purposes. The most frequent applications of these technologies were for compound identification, molecular structure prediction and retention time prediction. AI proved to be a valuable tool for analytical toxicologists for the capability to process large amount of data which are typically obtained by untargeted approaches.

Abstract:The medico-legal assessment of sports injuries in professional athletes continues to rely on tools designed for an entirely different context. Permanent impairment tables measure the reduction in overall individual function, but not the loss of competitive performance, the reduction in expected athletic lifespan, or the impact on contracts and earnings. In this Editorial we argue that closing this gap does not require new, ad hoc composite indices, but a more rigorous and systematic integration of approaches already validated in the sports medicine and rehabilitation literature: objective quantification of the return-to-sport (RTS) deficit, obtained by comparing validated sport-specific functional and performance outcome measures recorded before and after injury; structured documentation of the rehabilitation pathway, including objective RTS criteria, time-to-RTS, and the psychological readiness to compete; and an evidence-based estimate of the economic and contractual impact, grounded in objective performance-analytics data rather than presumptive percentages. The integration of these indices with clinical-orthopaedic assessment may allow a more accurate definition of "functional-professional" damage in athletes, with significant implications for insurance, legal, and medico-legal practice. The aim is to promote a new multidisciplinary assessment model capable of bridging the traditional distinction between biological damage and economic loss, providing a more realistic and equitable measure of the consequences of sports trauma on athletes' careers.

The first high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method was developed for enantioselective analysis of psychoactive compound N-ethylpentedrone. The method also enabled a detection of phase-1 metabolites of N-ethylpentedrone. Enantiomers some of the phase-1 metabolites were also separated. Application of this method to urine and oral fluid samples from the controlled naturalistic clinical study indicated to no enantioselective metabolism and elimination of N-ethylpentedrone. With the developed method a certain chemical and stereochemical instability of N-ethylpentedrone was also shown. Therefore, the observation about non-enantioselective pharmacokinetics of N-ethylpentedrone can be apparent and caused by post-collection racemization of this compound in the studied biological matrices.

Occult fractures (OFs) are inapparent fractures on initial plain radiographs, retrospectively diagnosed through second level imaging. The reasons for further diagnostic investigation may be the persistence of symptoms, or various personal needs, including insurance related. OFs, typically associated with low-energy trauma, are often underdiagnosed in the acute phase; however, they are relevant in the medico-legal field, as failure to identify them or delays in diagnosis and treatment may determine disability and give rise to complaint. Authors aim to describe OFs observed in a forensic case series. A retrospective study was conducted on technical expert consultations carried out between 2018 and 2024. Patients who sustained low-energy trauma with no evidence of fracture on plain radiography, followed by a later diagnosis of OF, were enrolled. Informed consent was signed. In each case, demographic data, trauma characteristics, diagnostic timelines, fracture locations, and details of any compensation claims were collected. 312 cases of OFs were included, with a prevalence at the sacrum (29.5

Many publications investigate drug concentration in various matrices aiming to select the most suitable matrix for bioanalytical studies. When for chiral drugs these studies are performed using nonenantioselective analytical methods they do not provide information about ratio of enantiomers in different matrices. A previously developed enantioselective high-performance liquid chromatography-mass spectrometry method was used for the analysis of methylone and its major phase 1 metabolites in plasma, urine, oral fluid, and sweat collected from a controlled clinical study. Along with the traditional representation of results as concentration-time dependence, the ratio of enantiomers was also plotted versus time. The concentration of methylone in oral fluid was higher compared with its concentration in circulating blood. The concentration of R-(+)-methylone was higher compared with S-(-)-methylone, and the enantiomeric excess was different in different matrices. In addition, stereochemical stability of methylone was found to be matrix dependent and surprisingly less in plasma compared with that in whole blood. Because of its higher concentration and higher stability in oral fluid, methylone is detectable in this matrix over a longer period of time after its administration than in any other studied matrices. The enantiomeric ratio in oral fluid is higher compared with that in plasma. In contrast to methylone, the concentration of methylone metabolites in oral fluid, except dihydromethylone, is lower than in plasma. The correlations of methylone concentrations in different matrices are fragile, with rather high interindividual variation, and cannot serve for reliable determination of time elapsed after drug administration. SIGNIFICANCE STATEMENT: This study demonstrates that oral fluid is a superior matrix for detecting methylone due to its higher concentration and greater stereochemical stability compared with plasma and other matrices. Enantioselective analysis reveals matrix-dependent stereochemistry and highlights that metabolite patterns differ from the parent drug. These findings emphasize the importance of matrix choice and enantioselective monitoring for accurate pharmacokinetic assessment and provide critical insights for interpreting drug detection and timing in forensic and clinical settings.

Orthopedic surgery is undergoing a transformation driven by artificial intelligence (AI), which is reshaping clinico-surgical decision-making. While the operative strategy and professional responsibility traditionally relied on the surgeon’s intuition and manual skills, advanced algorithms now provide predictive, analytical, and procedural decision supports. This paradigm shift is redefining the concept of human error as well as the relationship between technological tools and human decision-makers. As a result, the foundational elements of the healthcare liability framework are being affected. This paper offers a narrative discussion on selected applications of artificial intelligence in orthopedic surgical practice, including patient risk stratification, surgical indication and prosthesis positioning, with a particular focus on the liability implications for healthcare professionals who rely on these systems in terms of therapeutic decision-making. The aim is then to provide a comprehensive medico-legal perspective within the highly regulated and high-risk field of biomedicine, acknowledging and critically assessing the roles and responsibilities of all stakeholders involved—patients, healthcare professionals, innovative technologies, healthcare organizations, and facility management—while balancing innovation, evidence-based practice, and accountability in healthcare delivery.

Total ankle arthroplasty (TAA) has undergone significant evolution since its inception in the 1970s, transitioning from experimental approaches with high failure rates to more refined systems capable of replicating native ankle biomechanics. While ankle arthrodesis has traditionally been the gold standard for treating end-stage osteoarthritis, it compromises joint kinematics and predisposes adjacent joints to degeneration. Drawing inspiration from the success of hip and knee arthroplasty, early TAA designs attempted to restore motion but were limited by inadequate implant geometry, material performance, and surgical reproducibility. Advances in biomechanical understanding, imaging, and implant engineering have since enabled the development of second-and third-generation systems with improved outcomes. Key milestones include the transition from constrained two-component prostheses to semi-and fully mobile three-component designs, capable of accommodating complex rototranslational ankle motion. The Pipino-Calderale prosthesis, an early semi-fixed model designed to preserve the talar dome and instantaneous center of rotation (ICR), represented a pivotal effort to balance anatomical fidelity with mechanical stability. Building on this concept, the Vittore-Simone custom-made implant, modeled on 3D reconstructions of the contralateral ankle, demonstrated promising clinical results in a small series of patients, with significant improvements in pain and function over mid-term follow-up. Despite technical challenges and higher complication rates compared to other joint replacements, modern TAA continues to evolve, aided by improved biomaterials, surgical techniques, and personalized solutions. This review provides a historical and biomechanical synthesis of TAA development, highlighting how multidisciplinary advances have shaped current practice and may redefine ankle reconstruction in the future. (Cite this article as: Vittore D, Col & ograve; G, Fusini F, Basile G, Villafane J, Surace MF et al. Rationale and historical evolution of total ankle arthroplasty. Chirurgia 2026;39:135-40. DOI: 10.23736/S0394-9508.25.05999-6)

The enantioselective bioanalysis of synthetic cathinones and their metabolites remains analytically challenging. Due to the basic nature of these compounds, mobile phases with elevated pH are preferable for their analysis in high-performance liquid chromatography (HPLC). On the other hand, as it has been recently reported, cathinone derivatives are chemically and stereochemically instable, especially at high pH. As the alternative to HPLC, a supercritical fluid chromatography-tandem mass spectrometry (SFC-MS/MS) approach was developed in this study, to enable the chiral separation and detection of N-ethylpentedrone (NEP) and its major phase-1 metabolites. Careful selection of the chiral column and mobile phase compositions provided efficient retention and stereoselective resolution of both the parent compound and its polar metabolites. The applicability of the method was confirmed through the analysis of authentic human oral fluid (OF) and urine samples. The results of this study underscore the expanded analytical potential of SFC-MS/MS for chiral metabolite profiling and demonstrate its value as a complementary analytical tool to HPLC in forensic toxicology, particularly for new psychoactive substances.

Benzodiazepines are often used with other drugs like opioids, potentially leading to severe intoxications. Bretazenil, an imidazo-tetrahydropyrrolo-1,4-benzodiazepine, developed in the 1980s but never marketed as a medicine, has recently appeared on the illicit drug market. Given its high potency, short elimination half-life, and potential for rapid metabolism, it is essential to identify markers for bretazenil consumption for clinical and forensic purposes. Our study aimed to thoroughly explore bretazenil's metabolism using web-based in silico prediction tools, in vitro incubation with pooled human liver microsomes and hepatocytes, and to compare these results with authentic postmortem blood and urine samples. The in silico prediction revealed 16 metabolites, mainly formed by hydroxylation (phase I) and further O-glucuronidation, sulfation, and methylation (phase II) reactions. High-resolution mass spectrometry and software-aided data processing of in vitro and in vivo samples identified a total of 26 metabolites. Eight metabolites were detected in vitro, 15 in postmortem urine, and 11 in postmortem blood. Hydroxylation on the pyrrolidine ring was predominant. Other phase I reactions, including combinations of dihydroxylation, hydroxylation, reduction, and carboxylation as well as phase II glucuronidation and sulfation on the pyrrolidine ring, imidazole ring, or the tert-butyl chain, were also identified. Additionally, we discovered a new benzodiazepine biotransformation pathway via hydroxylation and cysteine conjugation in both human hepatocytes and blood. Due to bretazenil's extensive metabolism, we recommend hydroxy-bretazenil (B14), reduced hydroxy-bretazenil (B6), and reduced dihydroxy-bretazenil (B1) as significant markers for detecting bretazenil use.

Medical assistance in dying (MAID) remains a sensitive and evolving issue in Europe, frequently linked with discussions about human freedom, life dignity, and healthcare policy. While national consensus in Italy is absent, the Region of Tuscany has enacted Law No. 16/2025, which establishes a MAID procedure based on recent Constitutional Court rulings. The commentary aims to provide a preliminary analysis of the new law, addressing ethical, medico-legal, and social issues that emerge in relation to the Italian and global debate on the topic. The law establishes a three-stage process based on four eligibility criteria: irreversible disease, psycho-physical suffering, life-support dependence, and informed consent. However, Tuscany’s model poses medico-legal and ethical concerns, particularly about the boundaries of regional legislative competence, the duties of healthcare professionals, and the possibility of intra-national inequity or “health migration.” In addition, critical organisational implications derived from informed consent and lethal drug self-administration impede clinical implementation in some individuals with mental or neurological disorders. The lack of clarity in the different steps of the procedure, the uncertain supervision system, and the potential consequences for specific categories of vulnerable people underline the need for comprehensive national regulation. A future regulatory framework must balance procedural clarity with individual autonomy and equitable access, bringing Italy in line with larger European context for end-of-life care.

BACKGROUND:Litigation related to Healthcare-Associated Infections (HAIs) in Italy represent a growing field of interest in establishing the medico-legal link between infection and the healthcare environment and practices for compensation; it is little explored in the cardiovascular surgery regarding infective endocarditis (IE). METHODS:We retrospectively analysed the civil judgements on infective endocarditis in the Italian region Emilia-Romagna from 2016 to July 2024 using Ministry of Justice national official database. The search was conducted on the online database on July 31, 2024, using the free word "endocarditis". Two authors independently analysed the full-text judgements: , those IE without relevance in the reason for the claim were excluded. Main items were the timeline and outcome, with complaint motivation and liability ascertainment. In-court confirmation of healthcare causal link was reported. RESULTS:Twenty-five judgments were retrieved. After screening for inclusion, nineteen judgements (11 of first instance and 8 of appeal) were included, for overall 15 cases of infective endocarditis. Of the fifteen cases, median age 60.5 years, 73 % males, median time for claim 6 years, for judgement 10 years and, if appealed, 16.5 years. Annual distribution of the claims was linear over time. Eleven (67 %) infective endocarditis were confirmed as healthcare-associated in trial. The prevalent reason for liability was improper or delayed diagnosis and/or treatment of the IE. Valvular surgery resulted in 40 %, while the more frequent pathogens were Staphylococcus aureus (40 %) and epidermidis (30 %). Of fifteen cases, 73 % was decided in favour of the patient-claimant, with an average cost of €289.872, plus an additional €55.296 in case of appeal. Only in 25 % the appeal's judge changed decision. In all cases, technical advisors were appointed. CONCLUSIONS:This sample provides an initial insight into litigation for compensation related to infective endocarditis, highlighting specific characteristics compared to HAIs management in court. Medico-legal reasoning should be integrated into infection prevention and control policies and overall clinical risk management strategies.

Background: Non-cholera Vibrio species are rare waterborne pathogens that can cause severe infections. Among these, few cases of Vibrio metschnikovii infections have been reported, especially in the gastrointestinal tract, with no cardiac tissue involvement as a result. Following the PRISMA checklist, we conducted a literature review, and thirteen articles for twenty-two cases overall were included: seven cases of sepsis (in three cases, the echocardiographic results were negative), seven cases of pneumonia, two skin infections, eleven cases of diarrhoea, and a gastroenteritis outbreak. This report documents the expanding clinical spectrum and the role played by V. metschnikovii in infective endocarditis. Case report: A 28-year-old male patient was referred to the cardiac surgery unit for urgent mitral valve replacement due to suspicion of infective endocarditis. Microbiological tests yielded negative results. Following recovery and discharge with antimicrobial therapy for 6 weeks, the patient experienced prosthesis detachment, necessitating re-hospitalisation for an emergency valve replacement. Vibrio metschnikovii was identified on the prosthesis valve through PCR and successfully treated with ciprofloxacin. However, a spontaneous rupture of the ascending thoracic aorta led to a neurological injury. Discussion: This case represents the first case of valve infection caused by Vibrio metschnikovii, characterised by diagnostic and therapeutic challenges and the involvement of the great vessels. Also considered in this case, for a disease with a median age of 58 years (11–83) and a male-to-female ratio of 2.2, were one male neonate and six cases for whom neither sex nor age was indicated. Excluding gastrointestinal cases, the septic forms are associated with high morbidity, although the single case described involved a young and healthy subject. Risk factors for the pathogen or predisposing/pathological conditions for endocarditis did not emerge. The routes and the time of infection could not be determined, deepening the possibility of occupational exposure via the patient’s position as a boat worker. Poor sensitivity to third-generation cephalosporins has been reported in the literature: the absence of an antibiogram does not allow for a comparison, although resolution was achieved with ciprofloxacin. Conclusion: The rising global incidence of non-cholera Vibrio infections, driven by environmental changes, calls for urgent research into the factors behind their pathogenicity and infection routes. Diagnostic complexities have emerged together with clinical severity.

The diagnosis of septic arthritis remains challenging in the clinical setting, often leading to a suspicion for medical liability. Our purpose is to describe an unusual case of a post-mortem diagnosis of P. multocida fatal septic arthritis, in a healthy 67-year-old woman presenting with pain in the right shoulder. Moreover, a literature review of cases of fatal septic arthritis is provided. The multidisciplinary approach consisted of a forensic autopsy and additional post-mortem analyses (microbiology, biochemical analyses, histopathology, and revision of radiological images) carried out during the prosecutor’s investigation for medical liability. A systematic review of the literature was performed to collect cases of fatal septic arthritis and to understand its frequency and characteristics. No clear cause of death was determined after the autopsy, that only highlighted swelling and purulent exudation in the right glenohumeral joint. The microbiological swab performed on the shoulder tested positive for P. multocida, while histopathological and biochemical data were consistent with a sepsis. These results guided the interview with the woman’s relatives, until a history of a previous contact with a stray cat emerged. The cause of death was deemed as fatal septic arthritis caused by P. multocida, occurred after cat scratches and bites and only diagnosed post-mortem. The review of the literature provided 15 articles about fatal septic arthritis, only 1 caused by P. multocida, and all with ante-mortem diagnosis. Given the nonspecific symptoms, usually including a localized pain, and the absence of a clear history, e.g. of animal contact, septic arthritis might represent an under-reported clinical and pathological diagnosis, leading to a judicial autopsy for the suspicion for medical liability. The post-mortem examination, following a multidisciplinary approach including integration of the clinical history, microbiological and histopathological analysis, could represent the only opportunity for the diagnosis of the cause of death.

Introduction: Aesthetic surgery addresses subjective desires for morphological enhancement and differs from reconstructive surgery due to its elective, non-therapeutic nature. This distinction introduces complex medico-legal challenges, particularly concerning informed consent, patient expectations, and the legal evaluation of aesthetic damage. Materials and Methods: A narrative review was conducted using national legislation, Italian and international clinical guidelines, peer-reviewed literature from PubMed, Scopus, and Web of Science, and Italian Supreme Court rulings. Eight commonly litigated aesthetic procedures were analyzed in terms of clinical indications, public reimbursement criteria, and medico-legal risk. Results: Findings revealed significant variability in medico-legal exposure among procedures. Fully elective interventions such as liposuction and breast augmentation carried the highest litigation risk. Common legal claims included inadequate informed consent, poor psychological assessment, and mismatched expectations. The review emphasizes the need for personalized consent processes and comprehensive preoperative evaluations. Discussion: Italian case law increasingly adopts a “mixed obligation” model for aesthetic surgery, requiring not only technical skill but also a prognostic and relational evaluation of the intervention. Informed consent must be detailed, individualized, and well-documented, as it holds greater legal weight than in therapeutic procedures. Predictive medico-legal tools such as psychological profiling and structured consent protocols are essential for risk mitigation. Conclusions: Modern aesthetic surgery requires a redefined approach to damage assessment that incorporates psychological, relational, and identity factors. In both clinical and surgical practice, an approach tailored to the patient’s psychological profile must be increasingly taken into consideration, both when proposing and carrying out treatments and in medical-legal assessments. A legally and ethically sound practice depends on transparency, documentation, and patient-centered care, especially in the absence of therapeutic indications.

Neuropathological examination of the brain and its biochemical analyses are fundamental to neuroscience studies and public health decisions, but are dependent on the effectiveness of regulations and operational protocols. The article discusses opportunities and limits of Italian regulation on body donation in relation to the specific requirements of neuropathology and brain sciences, in comparison with the regulations of other countries. Some crucial issues emerge, widely shared in the various regulatory contexts. The main aspect is the willingness to donate, consciously expressed by the subject by signing an informed consent or through the formulation of advanced directives. The donation of a single organ, the brain in particular, does not necessarily imply the donation of the entire body, which should be considered separately. In the specific case of the brain, particular attention is given to reducing the post-mortem interval, in order to obtain tissues suitable for research. Consequently, the centres that deal with the brain and brain banking must have experience and expertise in handling nervous tissue, and do not necessarily have to deal with the management of the entire body. These aspects, still little addressed in Italy, are the basis to develop an effective brain banking activity, which can only develop by integrating post-mortem body donation with specific rules for brain banking without which Italian neuroscience will be penalised in the coming years.

BACKGROUND:Nerve injuries in total hip arthroplasty (THA) are rare but clinically and medico-legally significant adverse events, sometimes severely affecting patients' wellbeing through motor dysfunction and often permanent sensory alterations. Nerve injuries can result from complex interactions between a patient's pathophysiology, the technical aspects of surgery and individual response to surgical trauma - often rendering them not entirely avoidable. Professional liability should be assessed within a comprehensive framework, considering biological and procedural variables, adherence to clinical preventive measures, and proper documentation. OBJECTIVE:This study provides a narrative review of the etiology of nerve injuries and identifies the nerves most frequently affected, with a focus on perioperative actions associated with specific nerve involvement. It also explores the medico-legal implications of post-THA nerve injuries, for an integrated approach to surgical risk management. RESULTS:Nerves more frequently involved during the four phases of the surgical management (preoperative, intraoperative, immediate postoperative and long-term postoperative) are, in order, the sciatic, the femoral, the obturator, the superior gluteal and the lateral femoral cutaneous nerves. Main activities and/or situations associated with the risk of nerve injury, to be prevented or managed properly, are the type of approach, specific intraoperative positioning and timing, surgical technique with ligations, dissections and retractors roles, limb length, local retraction or expansion due to infection, hematoma or cement loss. Therefore, the adverse outcomes should not be automatically interpreted as surgical negligence, especially when standard care and clinical risk management protocols have been followed and documented in clinical records. Thus, distinguishing between unfaulty adverse event and negligence. CONCLUSIONS:The definition of a risk prevention and management strategy for nerve injury represents the first step in patient safety and, consequently, in reducing the likelihood of professional liability claim following THA.

Alexamorelin is a synthetic peptide and growth hormone secretagogue (GHS) with potential performance-enhancing properties, making its use and abuse a topic of interest in clinical research and doping monitoring. Alexamorelin mimics the natural peptide hormone ghrelin by binding to the GHS type 1a receptor (GHS-R1a) in the pituitary gland, thereby promoting endogenous growth hormone release. Identifying alexamorelin and/or its metabolite biomarkers is crucial for effective doping controls. The purpose of this study was to determine and characterize biomarkers associated with alexamorelin intake. In silico metabolite predictions were performed using GLORYx freeware, and in vitro incubations were conducted with pooled human hepatocytes from 10 donors. Samples were analysed using liquid chromatography–high-resolution tandem mass spectrometry (LC–HRMS/MS), with data processed through Thermo Scientific’s Compound Discoverer. GLORYx predicted 21 single-reaction metabolites. N-Acetylation was identified as the primary transformation, with the highest probability score (98%), and occurring either at the C-terminal Ala or the N-terminal Lys. Other predicted transformations included N-oxidation, hydroxylation, amide hydrolysis, oxidative deamination, and phase II N-glucuronidation, with probability scores below 40%. All these transformations were predicted to occur at the two C-terminal (Ala or His) or N-terminal (d-Phe or Lys) amino acids. After 3 h of incubation with hepatocytes, only one metabolite (known as examorelin or hexarelin) was detected, resulting from the C-terminal cleavage of the Ala amino acid; this metabolic reaction is mediated by a carboxypeptidase. The alexamorelin signal decreased approximately 150-fold after 3 h, indicating significant hepatic metabolism. However, examorelin itself is a commercially available GHS secretagogue, and thus, it is not specific to alexamorelin consumption. Detecting alexamorelin remains critical to documenting its use.