Vancomycin is a glycopeptide antibiotic that is commonly used in the treatment of serious infections caused by Gram-positive bacteria. It is a drug of choice in the management of methicillin-resistant Staphylococcus aureus (MRSA) infections, and its therapeutic efficacy is highly dependent on achieving appropriate serum concentrations (Pai et al., 2014). The Chinese Pharmacological Society recommends maintaining a serum trough concentration of vancomycin between 10 and 15 mg/L for adult patients, while a higher range of 10–20 mg/L is advised for serious MRSA infections. Clinical studies indicate that insufficient vancomycin levels can contribute to the emergence of resistance in S. aureus strains. Conversely, it is crucial to recognize that achieving a serum trough concentration of 15 mg/L or higher may increase the risk of nephrotoxicity (Ye et al., 2016). Moreover, trough levels alone may not provide a complete picture of the drug’s efficacy or potential for toxicity. Therefore, according to pharmacokinetic (PK)/pharmacodynamic (PD) theory, the better evaluation indicator for therapeutic drug monitoring (TDM) of vancomycin is the ratio of area under the concentration-time curve (AUC) to minimum inhibitory concentration (MIC), which takes into account the entire drug exposure over a dosing interval. To enhance clinical outcomes in the management of invasive MRSA infections, the latest guidelines from the American Society of Health-System Pharmacists (ASHP), Infectious Diseases Society of America (IDSA), and the Society of Infectious Diseases Pharmacists (SIDP) recommended transitioning from trough-based to AUC-guided vancomycin dosing, targeting an AUC/MIC ratio of 400–600 mg h/L based on the MIC of less than 1 mg/L for optimal efficacy against MRSA infections. A loading dose of 20–35 mg/kg is advised, with subsequent dosing adjusted based on renal function (Chen et al., 2022; Rybak et al., 2020; Kaushik et al., 2024). In addition, vancomycin presents a wide interindividual pharmacokinetic variation, a narrow therapeutic index, and the potential for various adverse effects, including nephrotoxicity (Matsumoto et al., 2022). Therefore, TDM of vancomycin and individualized dosing regimens based on patient-specific factors are highly recommended for increasing treatment response while preventing adverse effects (Rybak et al., 2020).

Liver transplant-receiving (LTR) patients are a special population that may require vancomycin therapy due to their immunosuppressed state and increased risk of infections (Morales Junior et al., 2023). However, they also have altered the pharmacokinetics and pharmacodynamics of drugs due to their liver dysfunction, post-operative complications, and polypharmacy (Morales Junior et al., 2023; Righi, 2018). Therefore, accurate TDM of vancomycin is crucial in this population to achieve optimal therapeutic outcomes and avoid toxicity. Studies have shown that TDM of vancomycin in LTRs can improve clinical outcomes, including reducing the incidence of nephrotoxicity and improving bacterial clearance. Additionally, TDM can help prevent the development of antibiotic resistance, being a global challenge (Shafiekhani et al., 2023), by ensuring the appropriate drug use.

Since the primary use of this drug, several methods have been developed and evaluated for the quantification of vancomycin levels in biological fluids, including immunoassays (Yeo et al., 1989; Filburn et al., 1983), high-performance liquid chromatography (HPLC) using either UV detection (Hagihara et al., 2013), fluorescence detection (Abu-Shandi, 2009), or photodiode array detection (Cao et al., 2014), and also liquid chromatography-mass spectrometry (LC-MS) (Oyaert et al., 2015). Up to now, HPLC has been considered the one of gold standards due to its high sensitivity, specificity, and accuracy, wide linear range of detection, low sample requirement, and ability to detect multiple analytes (Cheng et al., 2022; Ningrum et al., 2024). However, it typically involves sample preparation steps, such as protein precipitation or solid-phase extraction, which can be time-consuming and require skilled personnel (Aboelezz et al., 2025).

In recent years, a new method for vancomycin measurement has emerged, known as chemiluminescent microparticle immunoassay (CMIA). This solid-phase immunoassay quantifies vancomycin in serum by detecting the chemiluminescent reaction between the drug and a labeled antibody. CMIA offers several advantages over traditional high-performance liquid chromatography (HPLC), including faster processing times, ease of use, lower costs, and the availability of various commercial kits.

Recent studies have highlighted the effectiveness of CMIA for measuring vancomycin levels, particularly in patients undergoing hemodialysis. For example, a study conducted in Brazil compared CMIA with liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and found a strong correlation between the two methods. The mean difference indicated that CMIA provides reliable measurements without cross-reactivity in hemodialysis patients (Scribel et al., 2024). In another advancement, Fan et al. developed an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) technique to reduce cross-interaction in vancomycin assays. This innovative approach was evaluated against UPLC-UV and CMIA under normal, dialysis, and hemolytic conditions. The results demonstrated a moderate correlation between UPLC-UV and UPLC-MS/MS for samples from dialysis patients; however, some hemolytic samples showed overestimated results when analyzed with CMIA (Fan et al., 2019). The presence of cross-reacting substances, such as the crystalline degradation product of vancomycin, can lead to falsely elevated results in different immunoassay methods. This is particularly concerning in populations with renal impairment, where the accumulation of such metabolites is more likely (Chen et al., 2020).

Comparative studies have assessed CMIA against HPLC for other drugs as well. Guerrero Garduño et al. conducted a comparative analysis of HPLC and CMIA for quantifying carbamazepine and found comparable results, with a correlation coefficient of r ≈ 0.999, indicating that CMIA can reliably measure carbamazepine levels (Guerrero Garduño et al., 2016). Conversely, another study comparing HPLC and CMIA for valproic acid analysis indicated that HPLC offered greater precision than the immunoassay method (Zhao et al., 2016). Notably, highly metabolized drugs like cyclosporine A and tacrolimus were found to have significantly overestimated concentrations when measured by immunoassays compared to the UHPLC-MS method (Mei et al., 2018; Han et al., 2025).

Both CMIA and HPLC have been validated across various populations and for different drugs. Nonetheless, the potential for interference in CMIA necessitates careful interpretation of results, particularly in patients exhibiting altered pharmacokinetics.

Currently, there is a lack of data comparing the performance of CMIA with HPLC specifically for vancomycin, particularly in LTR patients. Therefore, this study aims to compare the accuracy and precision of CMIA and HPLC in measuring vancomycin levels in serum in LTRs, a topic that remains unexplored in the existing literature.

Therapeutic vancomycin monitoring is essential for the optimization of its therapeutic effect and the avoidance of nephrotoxicity. Several methods have been developed for this purpose, including HPLC and CMIA. CMIA is a rapid, simple, automated, and inexpensive method that has been highly taken into consideration. However, the main concern of using this method can be the accuracy and precision of the measurement. This is the first study comparing HPLC and CMIA methods to assess vancomycin concentration in LTRs.

Currently, the existing guidelines do not provide specific dosing regimens for particular populations, such as LTRs. Therefore, it is essential to first calculate the Pharmacokinetic parameters within this population before determining the most optimal dosage.

None of the pharmacokinetic variables observed from the CMIA and HPLC concentrations have a significant relationship with the calculated variables in LTR and it emphasizes the need to use therapeutic monitoring.

The present study used an accurate, selective, and sensitive validated HPLC method (Ghasemiyeh et al., 2020) and Abbott^® kit for vancomycin quantification. The limit of detection for HPLC and CMIA methods were 800 ng/mL and 3 μg/mL, respectively. So, HPLC provides a more sensitive assay for vancomycin.

Our results showed that vancomycin concentration obtained by HPLC and CMIA presented no linear regression between the two methods and the HPLC method was considered more accurate, precise, and versatile for both pharmacokinetic analysis and distinguishing the concentration differences between the patients with and without vancomycin-induced nephrotoxicity, significantly in trough 1, intermediate, and trough 2 concentrations. Considering the comparison of CMIA and HPLC concentrations in the studied population, the concentration investigated by CMIA was higher than that by HPLC in almost all patients during the 12-hour sampling. The peak concentration indicated the greatest differences and had a 105% deviation on average. Fan et al. have also reported an overestimated concentration via CMIA compared to UPLC/MS among hemodialysis patients, possibly secondary to interference of various panels of serum samples (Fan et al., 2019). Furthermore, a case report study serendipitously discovered a falsely high level of vancomycin using immunoassay methods in a patient not actually receiving this medication, perhaps due to being affected by endogenous proteins cross-interaction (Tsoi et al., 2019). However, Scribel et al. reported no cross-reactivity via CMIA among patients on hemodialysis (Scribel et al., 2024). On the other hand, numerous studies have examined the link between vancomycin trough levels and the risk of nephrotoxicity, generally finding that higher trough levels correlate with increased nephrotoxicity rates. In this regard, Horey et al. observed a correlation between vancomycin trough levels and nephrotoxicity rates. They found that the rates of nephrotoxicity were 4.9%, 3.1%, 10.6%, 23.6%, and 81.8% for maximal troughs of 5–10 mg/L, 10.1–15 mg/L, 15.1–20 mg/L, 20.1–35 mg/L, and greater than 35 mg/L, respectively (Horey et al., 2012). Similarly, Lodise et al. revealed that patients with trough levels greater than 10 mg/mL significantly more experienced nephrotoxicity compared to trough levels less than 10 mg/mL (22% vs. 5.1%) (Lodise et al., 2009). Cano et al. also noted an increase in nephrotoxicity from 7% at levels below 10 mg/L to 34% at levels above 20 mg/L (Cano et al., 2012). Barriere et al. observed no renal adverse events in patients with trough levels below 10 mg/L, while those with levels above this threshold experienced increasing incidences of renal issues (3% for 10–15 mg/L and 17% for above 15 mg/L) (Barriere et al., 2014). The present study similarly indicated that LTR patients having developed vancomycin-induced nephrotoxicity had a significantly higher vancomycin concentration at trough 1 (C1), intermediate (C3), and trough 2 (C4) observed via HPLC, however, the CMIA method was not accurate enough to distinguish the significant differences between the two groups at any intervals. In addition, the data obtained by HPLC showed that trough concentrations higher than 10 mg/L were associated with an increased risk of acute kidney injury in LTRs, highlighting the importance of monitoring pharmacokinetic parameters in this population to reduce adverse effects.

LTRs may experience altered pharmacokinetic parameters compared to the normal population due to the post-transplant inflammation state, changes in plasma protein level, the presence of hepatorenal syndrome, postoperative complications, and polypharmacy (Morales Junior et al., 2023; Righi, 2018; De Simone et al., 2024). The mean V_d in our investigation measured by HPLC and CMIA were 1.1 ± 0.7 and 0.53 ± 1.08 L/kg, respectively. V_d obtained from CMIA was in the typical range of 0.4 L/kg to 1 L/kg previously observed in the normal populations, while the value obtained from HPLC was significantly higher than this range which was also noted by other studies (Zhang et al., 2020) Among the studied population, many inter-individual variations were observed in V_d values. In this regard, the lack of significant correlation between the calculated and observed V_d highlights the possible changes due to fluid retention caused by organ dysfunction, enhanced capillary permeability resulting from systemic inflammatory response syndrome (SIRS) in ICU admitted patients, and ascites presence as a third-space volume. Additionally, lower baseline albumin levels in liver transplant recipients can also contribute to an increase in volume of distribution (Vd), particularly for hydrophilic antibiotics such as vancomycin. This effect was also emphasized in ICU-admitted critically ill patients in the study of Polard et al. (1999), in pediatric LTRs in the study of Shoji et al. (2021), and in adults having undergone bone marrow transplantation in the study of Taghizadeh Ghehi et al. (2013). Our findings demonstrated an increase in vancomycin clearance in LTRs, with a mean clearance of 4.91 ± 3.00 L/h and 2.93 ± 2.02 L/h obtained from HPLC and CMIA, respectively, compared to the general population’s average of 2.6 L/h. This elevation in clearance can be attributed to lower levels of albumin in LTRs, leading to higher unbound vancomycin and increased total clearance (both renal and non-renal) (Flannery et al., 2020; Rodvold et al., 1988), which are more manifest in data measured via the HPLC method in our study. Furthermore, the inflammatory condition prevalent in post-transplant recipients and also the use of diuretic drugs in such patients can exacerbate the renal excretion of antibiotics, leading to enhanced vancomycin clearance (Shimamoto et al., 2013). Moreover, we obtained a longer t_1/2 for vancomycin in LTRs than normal population. Harada H et al. have also found that patients with hepatic impairment exhibit a prolonged vancomycin t_1/2 (Harada et al., 1999). Additionally, there was a significant increase in t_1/2 in LTRs with renal insufficiency compared to non-vancomycin-induced nephrotoxicity LTRs through HPLC measurement, while CMIA data demonstrated no differences. Cheung et al. study reported longer vancomycin half-life in patients with impaired renal function, as well (Cheung and DiPiro, 1986). Our study also revealed a lower k_1/2 in LTRs than the normal population due to changes in both Cl_v and V_d values. In addition, our AUC_24h and AUC_12h values were below therapeutic levels reported in previous studies, denoting that higher loading doses may be necessary to achieve therapeutic AUC in LTRs. According to recent guidelines from ASHP, IDSA, and SIDP, the recommended AUC for effective treatment of MRSA infections is generally considered to be ≥400 mg·hr/L, with some studies suggesting that higher AUC values may be necessary for more severe infections, Nevertheless, there have been no reports on the optimal dosing required to ensure adequate drug exposure in LTRs (Rybak et al., 2020; Shoji et al., 2021). In a study by Alvarez et al. on critically ill patients admitted to ICU has been demonstrated that patients receiving a loading dose exhibited an AUC ranging between 600 and 700 mg.h/L (Álvarez et al., 2017). Holmes et al. conducted another study on patients with S.aureus bacteremia, indicating that 54% of patients with hepatic impairment or receiving immunosuppressive agents had an AUC greater than 400 mg.h/L (Holmes et al., 2013). Brown et al. also performed a study on patients with infective endocarditis and complex MRSA bacteremia, which demonstrated that the mortality rate was significantly lower in patients whose AUC/MIC values exceeded 400 mg.h/L (Brown et al., 2012). In our present study, we found that 41.2% of LTRs had an AUC above 400 mg.h/L, and failure to achieve the target AUC was associated with higher mortality rates. The average AUC was 383.3 ± 192.3 and 635.61 ± 301.60 via HPLC and CMIA measurement, respectively. When HPLC was used to assay AUC, LTR patients with higher values were significantly more susceptible to developing vancomycin-induced nephrotoxicity. Accordingly, the two points should be considered. First, HPLC is a more reliable test for vancomycin assessment than CMIA. Second, higher loading doses may be necessary for the LTRs to achieve therapeutic AUC levels. The pharmacokinetic alteration after liver transplantation indicates that standard dosing regimens may not achieve adequate therapeutic levels of the drug, necessitating tailored dosing protocols to ensure optimal drug exposure. Such adjustments are essential for the effective management of severe MRSA infections while also minimizing the risk of toxicity (Shoji et al., 2021). Prolonged exposure to suboptimal vancomycin levels can lead to the development of microbial resistance particularly in MRSA infections (Nyandoro et al., 2025), while higher vancomycin exposure particularly when AUC exceeds 550 mg.h/L is associated with nephrotoxicity in the treatment of MRSA bacteremia (Lodise et al., 2020; Chiu and Sarwal, 2023). Notably, our study highlighted that patients not reaching the AUC target had a higher mortality rate, emphasizing the need for optimized dosing strategies in this population. Clinicians should adopt an individualized approach to vancomycin dosing in LTRs through routine TDM, considering factors like renal function, concurrent medications, and nutritional status. Also, the potential overestimation of concentration using the CMIA method may lead to errors in drug dose adjustments and negatively impact therapeutic outcomes if this method is employed.

Furthermore, this study presents several limitations that must be acknowledged when interpreting the findings regarding therapeutic vancomycin monitoring in LTRs. Firstly, the sample size of our study was relatively small, which may limit the generalizability of the results. A larger sample size would provide more robust data and potentially reveal additional insights into the pharmacokinetics of vancomycin in this specific population. Secondly, this research was conducted at a single center, which may introduce biases related to local practices, patient demographics, and clinical protocols. The findings may not be applicable to other institutions with different patient populations or treatment approaches. Multi-center studies would be beneficial to validate our results across diverse settings. Additionally, the focus of this study was exclusively on liver transplant recipients, which restricts the applicability of the findings to other transplant populations, such as kidney or heart transplant patients. Variations in drug metabolism and pharmacodynamics among different transplant types could yield different therapeutic monitoring requirements. Lastly, while HPLC is a reliable method, it lacks the sensitivity of techniques such as LC-MS, which could provide more precise vancomycin measurements, particularly in patients with fluctuating drug concentrations.

In conclusion, this study highlighted the importance of selecting the appropriate analytical method for TDM of vancomycin in liver transplant recipients. Our findings suggested that HPLC was a more sensitive and reliable method compared to CMIA for both measuring vancomycin concentration and detecting variable changes in its pharmacokinetic profile. The use of HPLC may help clinicians make more informed decisions regarding dosing adjustments and ultimately improve patient outcomes. But, while this study provides valuable insights into therapeutic vancomycin monitoring, the aforementioned limitations should be considered when interpreting the results and their implications for clinical practice. Further research addressing these limitations is warranted to enhance our understanding and explore other potential benefits of using HPLC for vancomycin TDM in LTRs.