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Simple LC-MS/MS Methods Using Core-Shell ODS Microparticulate for the Quantitation of Total and Free Daptomycin in Human Plasma
Abstract
Background:
Daptomycin, a cyclic lipopeptide antibiotic, displays high plasma protein binding. This study developed the simple method of liquid chromatographic separation using a core shell ODS microparticulate coupled to tandem mass spectrometry for the quantitation of total and free daptomycin in human plasma.
Methods:
Free daptomycin in plasma was obtained by centrifugal ultrafiltration. Deproteinized plasma specimens were directly separated using a core-shell octadecylsilyl microparticulate with isocratic elution. The mass spectrometer was run in positive ion electrospray ionization mode. This method was applied to the quantitation of plasma samples in patients treated with intravenous daptomycin.
Results:
Daptomycin and diazepam as an internal standard were eluted with a total run time of 10 minutes. The calibration curves of total and free daptomycin in human plasma were linear over the concentration ranges of 1-100 and 0.1-10 µg/mL, respectively. The lower limits of quantitation of the total and free daptomycin in human plasma were 1.0 µg/mL and 0.1 µg/mL, respectively. Their extraction recovery rates in non- and ultrafiltrated plasma samples were 106.1% and 98.2%, respectively. Total and free daptomycin did not exhibit any matrix effects in human plasma. The intra- and inter-day accuracies and imprecisions of total daptomycin were 88.7-106.0% and 98.7-105.9%, and within 4.1% and 10.4%, while those of free daptomycin were 86.8-101.6% and 103.0-107.8%, and within 14.6% and 14.6%, respectively. The plasma concentration ranges of total and free daptomycin in 15 infected patients were 3.01-34.1 and 0.39-3.64 µg/mL, respectively. The plasma protein binding rate of daptomycin ranged from 80.8-94.9%.
Conclusions:
The present simple method with an acceptable analytical performance can be helpful for monitoring the pharmacokinetics of daptomycin in infected patients observed in clinical settings.
... Based on this, antimicrobials are classified by the following PK/PD indices that describe their efficacy: (1) time-dependent-the drug concentration remains above the MIC in a dose interval (fT > MIC); (2) concentrationdependent-the maximum concentration peak remains above the MIC (C max /MIC); and (3) area under the curve-the concentration and time remain above the MIC for at least 24 h (AUC 0-24 /MIC) [6,7,19]. The bactericidal effect of daptomycin is associated with an AUC/MIC > or equal to 200 [7], but recent studies have shown greater therapeutic efficacy with AUC/MIC levels > 666 [6,7,20] and a C max /MIC between 12-94 to obtain an optimal bacteriostatic effect [7] and suppression of bacterial resistance [19]. Therapeutic efficacy is directly related to optimal serum concentrations [19,20], which are expected to be obtained by means of a standard dosage schedule. ...
... The bactericidal effect of daptomycin is associated with an AUC/MIC > or equal to 200 [7], but recent studies have shown greater therapeutic efficacy with AUC/MIC levels > 666 [6,7,20] and a C max /MIC between 12-94 to obtain an optimal bacteriostatic effect [7] and suppression of bacterial resistance [19]. Therapeutic efficacy is directly related to optimal serum concentrations [19,20], which are expected to be obtained by means of a standard dosage schedule. ...
... Among the types of study design, the following were found: letter to an editor [22], poster [25,26], case report [27][28][29][30], bioanalytical methodology [8,16,20,[31][32][33][34][35][36][37][38][39][40][41][42][43][44][45], review article [6,7,15,21,23,24,[46][47][48], observational prospective study [17,18], observational retrospective study [49,50], and a single non-randomized clinical study [19]. The publication dates of the selected studies included publications from 2008 to 2020, and the type of design of each study obtained, date of publication, and general characteristics related to therapeutic monitoring are shown in Table 1. ...
Antimicrobial resistance (AR) is a problem that threatens the search for adequate safe and effective antibiotic therapy against multi-resistant bacteria like methicillin-resistant Staphylococcus aureus (MRSA), and vancomycin-resistant Enterococci (VRE) and Clostridium difficile, among others. Daptomycin is the treatment of choice for some infections caused by Gram-positive bacteria, indicated most of the time in patients with special clinical conditions where its high pharmacokinetic variability (PK) does not allow adequate plasma concentrations to be reached. The objective of this review is to describe the data available about the type of therapeutic drug monitoring (TDM) method used and
described so far in hospitalized patients with daptomycin and to describe its impact on therapeutic success, suppression of bacterial resistance, and control of side effects. The need to create worldwide strategies for the appropriate use of antibiotics is clear, and one of these is the performance of therapeutic drug monitoring (TDM). TDM helps to achieve a dose adjustment and obtain a favorable clinical outcome for patients by measuring plasma concentrations of an administered drug, making a rational interpretation guided by a predefined concentration range, and, thus, adjusting dosages individually.
... In addition to the above reports, the use of liquid chromatography-tandem mass spectrometry (LC-MS/MS) [26], ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) [27], and HPLC equipped with a photodiode array (UHPLC-PDA) [28] was also studied. ...
... Summary of Japanese reports on serum and CSF concentrations of CTRX, DAP, LZD, and TZD[18,[20][21][22][23][24][25][26][27][28][29][30][31][32][33].Table 1. Cont. Table 1. ...
Under the Japanese health insurance system, medicines undergoing therapeutic drug monitoring (TDM) can be billed for medical fees if they meet the specified requirements. In Japan, TDM of vancomycin, teicoplanin, aminoglycosides, and voriconazole, which are used for the treatment of infectious diseases, is common practice. This means the levels of antibiotics are measured in-house using chromatography or other methods. In some facilities, the blood and/or tissue concentrations of other non-TDM drugs are measured by HPLC and are applied to treatment, which is necessary for personalized medicine. This review describes personalized medicine based on the use of chromatography as a result of the current situation in Japan.
... Interestingly, the median synovial fluid/plasma ratio was 0.54X in the same group of patients. The plasma protein binding of daptomycin accounts for 80-90% [126], and this could explain the conflicting drug penetration rates into bone tissues. ...
The challenging severity of some infections, especially in critically ill patients, makes the diffusion of antimicrobial drugs within tissues one of the cornerstones of chemotherapy. The knowledge of how antibacterial agents penetrate tissues may come from different sources: preclin-ical studies in animal models, phase I-III clinical trials and post-registration studies. However, the particular physiopathology of critically ill patients may significantly alter drug pharmacokinetics. Indeed, changes in interstitial volumes (the third space) and/or in glomerular filtration ratio may influence the achievement of bactericidal concentrations in peripheral compartments, while inflammation can alter the systemic distribution of some drugs. On the contrary, other antibacterial agents may reach high and effective concentrations thanks to the increased tissue accumulation of macro-phages and neutrophils. Therefore, the present review explores the tissue distribution of beta-lac-tams and other antimicrobials acting on the cell wall and cytoplasmic membrane of bacteria in critically ill patients. A systematic search of articles was performed according to PRISMA guidelines, and tissue/plasma penetration ratios were collected. Results showed a highly variable passage of drugs within tissues, while large interindividual variability may represent a hurdle which must be overcome to achieve therapeutic concentrations in some compartments. To solve that issue, off-label dosing regimens could represent an effective solution in particular conditions.
... The MALDI-TOF/TOF can be used to analyse molecular weights ranging from 100-150.000 Da, it is however important to note that successful analysis have been conducted for proteins with molecular weights of over 1 kDa [7]. ...
... The RPLC column was coupled with an ESI(+)-QQQ-MS operating in MRM mode for detection and quantification. 79 For complex samples, containing a mixture of compounds with a large variety in physicochemical properties (e.g., containing both very polar and very apolar compounds), even the most efficient columns can often not resolve all important peaks in the chromatogram. 80,81 In this case, two-dimensional liquid chromatography (2D-LC) techniques can be employed to increase the separation capacity. ...
Liquid chromatography (LC) based techniques in combination with mass spectrometry (MS) detection have had a large impact on the development of new pharmaceuticals in the past decades. Continuous improvements in mass spectrometry and interface technologies, combined with advanced liquid chromatographic techniques for high-throughput qualitative and quantitative analysis, have resulted in a wider scope of applications in the pharmaceutical field. LC-MS tools are increasingly used to analyze pharmaceuticals across a variety of stages in their discovery and development. These stages include drug discovery, product characterization, metabolism studies (in vitro and in vivo) and the identification of impurities and degradation products. The increase in LC-MS applications has been very large, with retention times and molecular weights (and related fragmentation patterns) emerging as crucial analytical features in the drug development process from drug target to final marketed product. The goal of this review is to give an overview of the main developments in LC-MS based techniques for the analysis of small pharmaceutical molecules in the last decade, considering the main critical aspects, and give a perspective on future trends in LC-MS in the pharmaceutical field.
... Pipette 50 or 100 μl volume was from both the plasma and buffer chambers after 2-h incubation. The calculation of plasma protein binding rate was according to Yukari Miyadera's study (Yukari Miyadera et al., 2018). ...
Imatinib is a first-line drug for the treatment of gastrointestinal stromal tumors (GIST). This study aims to investigate the influence of different kinds of protein concentrations and genetic polymorphisms of metabolizing enzymes and drug transporters on unbound imatinib and its active metabolite N-desmethyl-imatinib concentration, as well as the relationship between adverse drug reactions (ADRs) and drug concentration. A total of 62 Chinese patients with GIST were genotyped for five single nucleotide polymorphisms (SNPs). Total and unbound 3h and trough concentration of imatinib and N-desmethyl-imatinib in GIST patients were determined by an LC-MS/MS method combined with an equilibrium dialysis. Single-Use Red Plate with inserts was used to separate the unbound drug. When the protein concentration became higher, the unbound imatinib and N-desmethyl-imatinib plasma concentration got higher (p < 0.05). Patients with GA genotype in rs755828176 had significantly higher unbound N-desmethyl-imatinib dose-adjusted trough plasma concentrations (p = 0.012). Patients with CC genotype in rs3814055 had significantly higher unbound imatinib dose-adjusted trough plasma concentrations (p = 0.040). The mean total imatinib C3h of patients with ADRs (3.10 ± 0.96 µg/ml) was significantly higher than that of patients without ADRs (p = 0.023). The mean total N-desmethyl-imatinib C3h of patients (0.64 ± 0.21 µg/ml) with ADRs was significantly higher than that of patients without ADRs (p = 0.004). The mean unbound N-desmethyl-imatinib C3h of patients with ADRs (6.49 ± 2.53 ng/ml) was significantly higher than that of patients without ADRs (p = 0.042). The total and unbound C3h of imatinib and N-desmethyl-imatinib in patients with ADRs was significantly higher than that in patients without ADRs (p < 0.05). Protein concentrations have great influence on the unbound imatinib and N-desmethyl-imatinib concentrations. The genetic polymorphisms of CYP3A4 rs755828176 and NR1I2 rs3814055 were significantly associated with unbound imatinib and N-desmethyl-imatinib dose-adjusted trough plasma levels. The total and unbound imatinib or N-desmethyl-imatinib concentration in patients with GIST was also significantly correlated with ADRs.
The stability of antibiotic preanalytical samples is a critical factor in therapeutic drug monitoring (TDM), a practice of undoubted importance for the proper therapeutic use of antibiotics, especially in complex management patients, such as pediatrics. This review aims to analyze the data in the literature regarding the preanalytical stability of some of the antibiotics for which TDM is most frequently requested. The literature regarding the preanalytical stability of amikacin, ampicillin, cefepime, ceftazidime, ciprofloxacin, daptomycin, gentamicin, levofloxacin, linezolid, meropenem, piperacillin, teicoplanin, and vancomycin in plasma, serum, whole blood, and dried blood/plasma spot samples was analyzed. Various storage temperatures (room temperature, 4 °C, −20 °C, and −80 °C) and various storage times (from 1 h up to 12 months) as well as subjecting to multiple freeze–thaw cycles were considered. The collected data showed that the non-beta-lactam antibiotics analyzed were generally stable under the normal storage conditions used in analytical laboratories. Beta-lactam antibiotics have more pronounced instability, particularly meropenem, piperacillin, cefepime, and ceftazidime. For this class of antibiotics, we suggest that storage at room temperature should be limited to a maximum of 4 h, storage at 2–8 °C should be limited to a maximum of 24 h, and storage at −20 °C should be limited to a maximum of 7 days; while, for longer storage, freezing at −80 °C is suggested.
Reports of therapy failures related to the use of daptomycin (DAP) are steadily increasing. This is mainly due to emerging DAP resistance for which, however, the underlying mechanism is often unknown. To elucidate the mode of action of novel DAP resistance traits it is indispensable to reliably detect and quantify DAP in complex matrices such as bacterial culture media. In this study, we established a selective and sensitive quantification method for DAP upon growth of a DAP resistant Mammaliicoccus sciuri strain in Mueller-Hinton medium. The method combined methanol-induced protein precipitation followed by high performance liquid chromatography with gradient elution coupled to mass spectrometry (LC-MS/MS) using daptomycin-d5 as internal standard. All validation parameters met the acceptance criteria of the European Medicines Agency (EMA) guideline on bioanalytical method validation. We successfully applied this highly selective and sensitive LC-MS/MS method for the quantification of DAP in in vitro studies addressing DAP resistance mechanisms in Gram-positive bacteria.
Purpose
Plasma daptomycin has not been fully characterized in diabetic and obese patients. This study aimed to evaluate the associations of plasma daptomycin with glycation of serum albumin and obesity.
Methods
Infectious patients (n = 70) receiving intravenous daptomycin were enrolled. The plasma concentration of total and free daptomycin were determined using liquid chromatograph-tandem mass spectrometer. The associations of the plasma concentrations of daptomycin with clinical factors including serum albumin fractionations and physical status (obese including overweight, body mass index ≥ 25) were investigated. Daptomycin doses were adjusted using total body-weight.
Results
The serum albumin level was positively and negatively correlated with the plasma concentration of total daptomycin and its free fraction proportion, respectively. The serum non-glycated albumin was negatively correlated with the free fraction proportion. The dose-normalized plasma concentration of total daptomycin was higher in the obese patients than in non-obese patients when the body-weight was corrected with total and adjusted values. For the dose adjustment with lean body-weight, no difference was observed in the dose-normalized plasma concentration of total daptomycin between the physical statuses. For each body-weight correction method, physical status did not affect the dose-normalized plasma concentration of free daptomycin.
Conclusion
The glycation of serum albumin and obesity did not associate with dose-normalized plasma free daptomycin. In obese patients, daptomycin dosage adjustment with total body-weight and adjusted body-weight may lead to an apparent excessive exposure resulting in overdosage compared to lean body-weight.
Antimicrobial resistance is a major global health challenge. As few new efficacious antibiotics will become available in the near future, peptide antibiotics continue to be major therapeutic options for treating infections caused by multidrug-resistant pathogens. Rational use of antibiotics requires optimisation of the pharmacokinetics and pharmacodynamics for the treatment of different types of infections. Toxicodynamics must also be considered to improve the safety of antibiotic use and, where appropriate, to guide therapeutic drug monitoring. This review focuses on the pharmacokinetics/pharmacodynamics/toxicodynamics of peptide antibiotics against multidrug-resistant Gram-negative and Gram-positive pathogens. Optimising antibiotic exposure at the infection site is essential for improving their efficacy and minimising emergence of resistance.
Background:
Aprepitant, an antiemetic selective neurokinin-1 receptor antagonist, is primarily metabolized to the active N-dealkylated (ND-AP) form and then converted to its carbonyl (ND-CAP) form in humans. This study developed simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods using electrospray ionization (ESI) for the quantitation of plasma total and free aprepitant and its N-dealkylated metabolites and used them to analyze patient plasma.
Methods:
Free aprepitant and ND-AP in plasma were fractionated using centrifugal ultrafiltration. The analytes in plasma or their ultrafiltered specimens treated with triethylamine/acetonitrile were isocratically separated using a 3-μm octadecylsilyl column with a total run time of 10 minutes and scanned using positive ion ESI.
Results:
The calibration curves of total aprepitant, ND-AP, and ND-CAP were prepared at concentration ranges of 50-2500, 20-1000, and 5-250 ng/mL, respectively, while that of free aprepitant and ND-AP was at a concentration range of 2-150 ng/mL. The intra- and inter-assay accuracy and imprecision values were 93.5-107.7% and 94.6-103.3%, and 2.1-7.5% and 1.0-8.9%, respectively. Aprepitant and its metabolites did not exhibit any matrix effects or instabilities in the plasma specimens. In cancer patients receiving oral aprepitant, the plasma concentration ranges of total aprepitant, ND-AP, and ND-CAP, and free aprepitant and ND-AP were 137-2170, 104-928, 22.4-97.6, 8.11-60.0, and 3.53-56.0 ng/mL, respectively. The median plasma free fraction proportion of aprepitant and ND-AP were 4.14% and 4.90%, respectively.
Conclusions:
The present developed method showed an acceptable analytical performance and can be used to evaluate total and free aprepitant and its N-dealkylated metabolites in patient plasma.
A simple and new validated liquid chromatography method with stability‐indicating potential was established for quantifying daptomycin in formulations using a quality by design approach. Analytical target profile establishment and earmarking critical analytical attributes helped to control the method performance. Further, the control strategies for critical method variables were defined as per method intent. Youden's robustness test was applied with the dual intent of defining control strategies and evaluating method robustness for continuous method improvement. Chromatography for the purpose constituted of methanol: water (pH 3.5 maintained by o‐phosphoric acid), 90:10, v/v as mobile phase flowing at 1.2 mL/min on a C18 column. Photodiode array detector provided the best results at 367 nm. Results for parameters viz. linearity (5‐200 µg/mL), accuracy (> 99%) and precision (< 1%) advocated method reliability. The detection (2.5 µg/mL) and quantitation limits (5 µg/mL) were quite in agreement with method need. In a nutshell, the hyphenation of Youden's test with quality by design approach ensured the development of a reliable chromatography method for routine estimation of daptomycin throughout its product lifecycle.
The rapid antibiotic resistance development has created a major demand for new antimicrobial agents that can combat resistant strains such as methicillin-resistant S. aureus (MRSA). Until a short time ago, the glycopeptide vancomycin was the only therapeutic choice in this situation. However, in recent years some newer agents with different mechanisms of actions have been added to the arsenal, and more are on the horizon. For a successful therapy it is of vital importance that these compounds are used judiciously and dosed appropriately. The present article reviews the pharmacokinetic properties of vancomycin, linezolid, tigecycline and daptomycin. The first major difference between these compounds is their oral bioavailability. Only linezolid can be administered orally, whereas vancomycin, daptomycin and tigecycline are limited to parenteral use. Once in the body, they show very different disposition. Daptomycin has a very small volume of distribution of 7L indicating very little tissue distribution whereas tigecycline has a very large volume of distribution of 350-500 L. Vancomycin and linezolid are in-between with volumes of distribution of approximately 30 and 50 L, close to total body water. However, studies have shown that linezolid shows better tissue penetration than vancomycin. Newer studies using microdialysis, a new technique that allows direct monitoring of unbound tissue levels, support this finding. As far as drug elimination, daptomycin and vancomycin are mainly eliminated into the urine and require dosing adjustments in renally impaired patients, whereas tigecycline is eliminated into the bile and linezolid is metabolized so that in renal patients no dosing adjustments are needed for these compounds. Although the elimination pathways are very different, the resulting half-lives of linezolid, vancomycin, and daptomycin are not greatly different and vary from 4-8 h. Tigecycline, however, has a much longer half-life of up to 1-2 days due to the slow redistribution from tissue binding sites.
Low serum albumin levels are very common in critically ill patients, with reported incidences as high as 40-50%. This condition appears to be associated with alterations in the degree of protein binding of many highly protein-bound antibacterials, which lead to altered pharmacokinetics and pharmacodynamics, although this topic is infrequently considered in daily clinical practice. The effects of hypoalbuminaemia on pharmacokinetics are driven by the decrease in the extent of antibacterial bound to albumin, which increases the unbound fraction of the drug. Unlike the fraction bound to plasma proteins, the unbound fraction is the only fraction available for distribution and clearance from the plasma (central compartment). Hence, hypoalbuminaemia is likely to increase the apparent total volume of distribution (Vd) and clearance (CL) of a drug, which would translate to lower antibacterial exposures that might compromise the attainment of pharmacodynamic targets, especially for time-dependent antibacterials. The effect of hypoalbuminaemia on unbound concentrations is also likely to have an important impact on pharmacodynamics, but there is very little information available on this area. The objectives of this review were to identify the original research papers that report variations in the highly protein-bound antibacterial pharmacokinetics (mainly Vd and CL) in critically ill patients with hypoalbuminaemia and without renal failure, and subsequently to interpret the consequences for antibacterial dosing. All relevant articles that described the pharmacokinetics and/or pharmacodynamics of highly protein-bound antibacterials in critically ill patients with hypoalbuminaemia and conserved renal function were reviewed.
The objective of this analysis was to evaluate the relationship between daptomycin exposure and the probability of an elevation in the creatine phosphokinase (CPK) level (hereafter, "CPK elevation") in patients with Staphylococcus aureus bacteremia with or without infective endocarditis.
Phase 3 data for patients with S. aureus bacteremia, with or without infective endocarditis, who received intravenous daptomycin (6 mg/kg daily) and in whom pharmacokinetic data were collected were evaluated. On the basis of univariate logistic regression, the relationship between Bayesian post hoc exposure estimates and the probability of a CPK elevation was evaluated. Time to CPK elevation was examined with Kaplan-Meier analysis and Cox proportional hazards regression.
Significant relationships between the minimum concentration of drug (C(min)) and area under the plasma concentration time curve and probability of CPK elevation were observed in 108 evaluable patients. Of the 108 patients evaluated, 6 (5.56%) demonstrated a defined CPK elevation, regardless of treatment relationship. C(min) (breakpoint of 24.3 mg/L) was most significantly associated with CPK elevation (P = .002). The probabilities of a CPK elevation with a C(min) 24.3 mg/L and <24.3 mg/L were 0.5 and 0.029, respectively. Increases in C(min), evaluated as a continuous variable, were also significantly associated with CPK elevation (P = .01). Stratified Kaplan-Meier analysis and Cox proportional hazards regression demonstrated C(min) to be a significant predictor of time to a CPK elevation (P .003). The probability of a CPK elevation was 0 and 0.01 after 7 days of treatment in patients with a C(min) 24.3 mg/L or <24.3 mg/L, respectively. After 14 days, the probabilities were 0.5 and 0.025, respectively.
This analysis demonstrated that a daptomycin C(min) 24.3 mg/L was associated with an increased probability of a CPK elevation. Clinical trials registration. Clinical trials.gov NCT00093067 .
The purpose of this paper is to establish the pharmacokinetics and safety of escalating, once-daily doses of daptomycin, a novel lipopeptide antibiotic active against gram-positive pathogens, including those resistant to methicillin and vancomycin. This phase 1, multiple-dose, double-blind study involved 24 healthy subjects in three dose cohorts (4, 6, and 8 mg/kg of body weight) who were randomized to receive daptomycin or the control at a 3:1 ratio and administered the study medication by a 30-min intravenous infusion every 24 h for 7 to 14 days. Daptomycin pharmacokinetics was assessed by blood and urine sampling. Safety and tolerability were evaluated by monitoring adverse events (AEs) and laboratory parameters. Daptomycin pharmacokinetics was linear through 6 mg/kg, with a slight ( approximately 20%) nonlinearity in the area under the curve and trough concentration at the highest dose studied (8 mg/kg). The pharmacokinetic parameters measured on the median day of the study period, (day 7) were half-life ( approximately 9 h), volume of distribution ( approximately 0.1 liters/kg), systemic clearance ( approximately 8.2 ml/h/kg), and percentage of the drug excreted intact in urine from 0 to 24 h ( approximately 54%). Daptomycin protein binding (mean amount bound, 91.7%) was independent of the drug concentration. No gender effect was observed. All subjects who received daptomycin completed the study. The frequencies and distributions of treatment-emergent AEs were similar for the subjects who received daptomycin and the control subjects. There were no serious AEs and no pattern of dose-related events. The pharmacokinetics of once-daily administration of daptomycin was linear through 6 mg/kg. For all three doses, plasma daptomycin concentrations were consistent and predictable throughout the dosing interval. Daptomycin was well tolerated when it was administered once daily at a dose as high as 8 mg/kg for 14 days.
Daptomycin is a lipopeptide antibiotic with activity against a wide range of gram-positive bacteria. We used the neutropenic
murine thigh model to characterize the pharmacodynamics of daptomycin. ICR/Swiss mice were rendered neutropenic with cyclophosphamide;
and the thigh muscles of the mice were infected with strains of Staphylococcus aureus, Streptococcus pneumoniae, and Enterococcus faecium. Animals were treated by subcutaneous injection of daptomycin at doses of 0.20 to 400 mg/kg of body weight/day divided into
one, two, four, or eight doses over 24 h. Daptomycin exhibited linear pharmacokinetics, with an area under the concentration-time
curve (AUC) from time zero to infinity/dose of 9.4 and a half-life of 0.9 to 1.4 h. The level of protein binding was 90%.
Free daptomycin exhibited concentration-dependent killing and produced in vivo postantibiotic effects (PAEs) of 4.8 to 10.8
h. Nonlinear regression analysis was used to determine which pharmacokinetic (PK) or pharmacodynamic (PD) parameter was important
for efficacy by using free drug concentrations. The peak concentration/MIC (peak/MIC) ratio and 24-h AUC/MIC ratio were the
PK and PD parameters that best correlated with in vivo efficacy (R2 = 83 to 87% for peak/MIC and R2 = 86% for the AUC/MIC ratio, whereas R2 = 47 to 50% for the time that the concentration was greater than the MIC) against standard strains of S. aureus and S. pneumoniae. The peak/MIC ratios required for a bacteriostatic effect ranged from 12 to 36 for S. pneumoniae, 59 to 94 for S. aureus, and 0.14 to 0.25 for E. faecium. The AUC/MIC ratios needed for a bacteriostatic effect ranged from 75 to 237 for S. pneumoniae, 388 to 537 for S. aureus, and 0.94 to 1.67 for E. faecium. The free daptomycin concentrations needed to average from one to two times the MIC over 24 h to produce a bacteriostatic
effect and two to four times the MIC over 24 h to produce greater than 99% killing. The long PAE and potent bactericidal activity
make daptomycin an attractive option for the treatment of infections caused by gram-positive bacteria.
Daptomycin is the first available agent from a new class of antibiotics, the cyclic lipopeptides, that has activity against
a broad range of gram-positive pathogens, including organisms that are resistant to methicillin, vancomycin, and other currently
available agents. Daptomycin (4 mg/kg intravenously [iv] every 24 h for 7–14 days) was compared with conventional antibiotics
(penicillinase-resistant penicillins [4–12 g iv per day] or vancomycin [1 g iv every 12 h]) in 2 randomized, international
trials involving 1092 patients with complicated skin and skin-structure infections. Among 902 clinically evaluable patients,
clinical success rates were 83.4% and 84.2% for the daptomycin- and comparator-treated groups, respectively (95% confidence
interval, -4.0 to 5.6). Among patients successfully treated with iv daptomycin, 63% required only 4–7 days of therapy, compared
with 33% of comparator-treated patients (P < .0001). The frequency and distribution of adverse events were similar among both treatment groups. Overall, the safety
and efficacy of daptomycin were comparable with conventional therapy.
Data from subjects in nine phase 1 (n = 153) and six phase 2/3 (n = 129) clinical trials were combined to identify factors contributing to interindividual variability in daptomycin pharmacokinetics
(PK). Over 30 covariates were considered. A two-compartment model with first-order elimination provided the best fit for data
on daptomycin concentrations in plasma over time. In the final population PK model, daptomycin plasma clearance (CL) was a
function of renal function, body temperature, and sex. Of these factors, renal function contributed most significantly to
interindividual variability. CL varied linearly with the estimated creatinine clearance. CL among dialysis subjects was approximately
one-third that of healthy subjects (0.27 versus 0.81 liter/h). CL in females was 80% that in males; however, in clinical trials,
the outcome was not affected by sex and therefore this effect is not considered clinically meaningful. The relationship with
body temperature should be interpreted cautiously since the analysis included only a limited number of subjects who were hyperthermic.
The volume of distribution of the peripheral compartment (V2) and intercompartmental clearance (Q) were linearly related to body weight. V2 increased approximately twofold in the presence of an acute infection. No factors were identified that significantly impacted
V1. This analysis supports the dosing of daptomycin on a milligram-per-kilogram-of-body-weight basis and suggests that modified
dosing regimens are indicated for patients with severe renal disease and for those undergoing dialysis.
Staphylococcus aureus strains from the U.S. SENTRY Antimicrobial Surveillance Program, 2002-2003, glycopeptide-intermediate S. aureus (GISA) strains, and heterogeneous GISA (hGISA) strains were used to compare bactericidal activities of daptomycin and vancomycin
using MICs and minimum bactericidal concentrations. Glycopeptide-susceptible S. aureus and hGISA strains were further studied by using time-kill curves. For all isolates, the daptomycin MIC50 and MIC90 are four times lower and the log drops in viable counts at 6 h and 24 h are significantly greater than those for vancomycin.
Proteomic approaches using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) without an immunopurification technique have not been applied to the determination of serum cetuximab. This study developed a simple and rapid LC-MS/MS method for the absolute determination of cetuximab in human serum and applied it to clinical settings. Surrogate peptides derived from cetuximab digests were selected using a Fourier transform mass spectrometer. Reduced-alkylated serum cetuximab without immunopurification was digested for 20minutes using immobilized trypsin, and the digestion products were purified by solid-phase extraction. The LC-MS/MS was run in positive ion multiple reaction monitoring mode. This method was applied to the determination of serum samples in head and neck cancer patients treated with cetuximab. The chromatographic run time was 10minutes and no peaks interfering with surrogate peptides in serum digestion products were observed. The calibration curve of absolute cetuximab in serum was linear over the concentration range of 4-200μg/mL. The lower limit of quantification of cetuximab in human serum was 4μg/mL. The intra-assay and inter-assay precision and accuracy were less than 13.2% and 88.0-100.7%, respectively. The serum concentration range of cetuximab was 19-140μg/mL in patients. The serum cetuximab concentrations in LC-MS/MS were correlated with those in ELISA (r=0.899, P <0.01) and the mean bias was 1.5% in cancer patients. In conclusion, the present simple and rapid method with acceptable analytical performance can be helpful for evaluating the absolute concentration of serum cetuximab in clinical settings.
Daptomycin, a lipopeptide antibiotic with excellent activity against Gram-positive bacteria, is excreted primarily by the kidneys. Development of effective chromatographic methodologies for the determination of daptomycin in human specimens is necessary for clinical use. This study developed a simple and validated ultra-high-performance liquid chromatography method coupled to ultraviolet detection for determination of daptomycin in human plasma and urine. After the pretreatments involving protein precipitation, the supernatants were separated using a 2.3 µm particle size octadecylsilyl column, and the run time was 1 min. The calibration curves were linear over the concentration ranges of 2-200 mg/L for plasma and 25-300 mg/L for urine. Intra- and inter-assay precision and accuracy values of plasma were within 13.5 and 92-100% and within 10.7 and 100-107%, respectively. Those of urine were within 5.0 and 101-104% and within 3.7 and 100-101%, respectively. The validated method was applied to the determination of plasma and urine samples in patients receiving 4-6 mg/kg of intravenous daptomycin, resulting in sufficient sensitivity for evaluating the plasma exposure and urinary excretion. In conclusion, the present method with acceptable analytical performance can be helpful for evaluating the pharmacokinetic disposition of daptomycin in clinical settings. Copyright © 2013 John Wiley & Sons, Ltd.
High-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a standard analytical technique for therapeutic drug monitoring (TDM). A rapid LC-MS/MS method was developed for simultaneous quantitation of 3 antifungals and one active metabolite (posaconazole, voriconazole, itraconazole, and hydroxy-itraconazole), 5 antibiotics (daptomycin, ciprofloxacin, oxacillin, levofloxacin, and rifampicin), an antineoplastic agent (imatinib), and an antiretroviral (raltegravir) in human plasma. Protein precipitation of 10μL of plasma with acetonitrile was used as a single-extraction procedure. After 2-dimensional LC, all drugs were quantified by electrospray ionization-triple quadrupole mass spectrometry by selected reaction monitoring detection in the positive mode. The method was validated per FDA recommendations including the study of extraction recovery (from 79.3% to 105.9%) and matrix effect via ion suppression/enhancement phenomenon. This method is precise (intra- and inter-assay coefficients of variation of 1.95-12.77%, 2.56-8.16% and 2.12-11.38% for low, medium and high levels of internal quality controls respectively) and accurate (intra- and inter-assay biases of 0.19-12.67%, 0.04 to -12.17% and 0.22-12.98% respectively). This method is an efficient tool for routine TDM and optimization of laboratory resource utilization.
A simultaneous extraction method to measure daptomycin, amikacin, gentamicin, and rifampicin in human plasma, by high-performance
liquid chromatography, was developed and validated. The method involved a rapid sample preparation by protein precipitation
with acetonitrile followed by direct injection into a high-performance liquid chromatography system coupled with mass detection.
Drug retention times were 10.00 ± 0.25, 2.00 ± 0.25, 3.50 ± 0.25, 11.50 ± 0.25, and 12.50 ± 0.25min for daptomycin, amikacin,
gentamicin, rifampicin, and quinoxaline, respectively. Good linearity (mean r
2 = 0.998) was obtained for all drugs quantified over the range of clinically relevant concentrations in human plasma and the
use of the internal standard quinoxaline improves accuracy (RSD% <14.9%) and intra-day (RSD% <11.56) and inter-day (RSD% <12.10)
precision for the analytical procedure. The limits of quantification for daptomycin, amikacin, gentamicin, and rifampicin
were 1.56, 2.34, 0.63, 0.63μg/ml, respectively. Moreover, the addition of ion pair trifluoroacetic acid in the sample allowed
the majority of gentamicin and amikacin separation. A rapid, specific, sensitive, accurate, and reproducible HPLC method was
developed and validated to measure daptomycin, amikacin, gentamicin, and rifampicin in human plasma. This method is suitable
for clinical pharmacokinetic studies.
A rapid, simple and accurate analytical method based on ultra performance liquid chromatography (UPLC) combined with electrospray ionization (ESI) tandem mass spectrometry (MS/MS) on a hybrid q TOF instrument has been developed and fully validated for the quantification of daptomycin (DPT) in human plasma. The samples were analyzed after simple pretreatment involving protein precipitation, while chromatographic separation of DPT and the internal standard (reserpine) was achieved on an Acquity BEH C18 column (100 mm × 2.1 mm, 1.7 μm) using gradient elution with 0.1% aqueous formic acid (FA) and acetonitrile with 0.1% FA (with DPT eluting at 2.60 min). The method presented good fit (r>0.999) over the quantification range of 0.01-10 μg mL⁻¹ with the lower limit of quantitation (LLOQ) being 0.01 μg mL⁻¹ of human plasma for DPT. The intra- and inter-day precision, measured as % relative standard deviation, was less than 11% for DPT. The validation results showed that the developed method demonstrated adequate selectivity, sensitivity, precision and accuracy and therefore was successfully applied to the analysis of clinical samples following intravenous (iv) administration of 5.4 mg kg⁻¹ DPT to patients suffering from post-traumatic osteomyelitis induced by methicillin-resistant Staphylococcus aureus (MRSA). The developed methodology is the first report of an accurate mass tandem MS method for the analysis of this potent antibiotic in human plasma and can be used to further study pharmacokinetic, bioequivalence and even metabolic aspects related to this drug.
Daptomycin is a recently developed cyclic lipopeptide antibiotic active against most Gram-positive pathogens including vancomycin-resistant enterococci and methicillin-resistant Staphylococcus aureus. To optimize treatment efficacy and safety, especially in patients undergoing multiple drug regimens and/or co-morbidities, a specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the quantification of daptomycin in plasma.
A C18 column was used for separation, with a mobile phase initially consisting of 0.1% formic acid, water, and acetonitrile (ACN) in a linear gradient from 20% to 70%. After protein precipitation with ACN, the clear upper layer was diluted in water:ACN (50:50, v/v) before injection. Detection was performed using an electrospray ionization technique. MS/MS transitions, monitored in the positive ion mode were m/z 811.1 → m/z 313.1 for daptomycin, and m/z 609.4 → m/z 194.9 for reserpine, used as internal standard.
Elution of daptomycin and reserpine occurred at 4.5 and 3.9 min, respectively. The method was validated over a range of concentrations from 1 mg/L to 120 mg/L. The assay met recommended acceptance criteria: coefficients of variation were <6.3% and <7.4%, and accuracies were between -5.9% and +11.2% and between -3.5% and +3.7%, for intra- and inter-day validations, respectively.
This method appears well-adapted to routine hospital practice for therapeutic drug monitoring of daptomycin considering its time of analysis, range of concentrations measured, precision and accuracy.
A 13-min LC-MS method was developed for the determination of daptomycin, a new potent antibiotic, in peritoneal fluid, blood plasma, and urine of patients receiving renal replacement therapy. Chromatography was performed on a C(18) column and detection was performed by a single-quadrupole mass spectrometer coupled to LC via an electrospray interface (ESI). The column effluent was also monitored at 370 nm using a photodiode-array detector. The developed method provided a linear dynamic range for concentrations from 0.5 microg mL(-1) to 100 microg mL(-1). Method precision and accuracy were found to be satisfactory for clinical application, thus the method was successfully used for the analysis of daptomycin in pharmacokinetic studies. The drug was preventively administered against Gram-positive infections to 19 clinical patients undergoing peritoneal dialysis. Peritoneal fluid, blood plasma, and urine samples were collected at 13 time points over a period of 48 h. Clinical samples were analysed following simple sample-preparation procedures and daptomycin was unambiguously detected and quantified.
Daptomycin is the first approved member of the new class cyclic lipopeptide antibiotic drugs, effective against a broad spectrum of Gram-positive bacteria. Here we present an HPLC method with UV detection capable to obtain pharmacokinetic data of daptomycin in human plasma, exemplarily shown in a critically ill patient with acute renal failure undergoing extended daily dialysis. Sample preparation consists only of protein precipitation with methanol. Chromatographic separation was achieved on a Zorbax Eclipse XDB-C8 column and daptomycin was detected at 224 nm. The calibration function was linear over the range from 3.5 to 350 microg/ml. The relative standard deviations were <2% in the intra-day and <6% in the inter-day measurements. The accuracy was always better than 7%. Daptomycin was stable in aqueous solutions for 2 months frozen at-20 degrees C. However, in plasma frozen at -20 degrees C a loss of 25% in 1 month was observed.
The reliability of the most important methods to determine protein-binding of drugs was compared. Applying these methods to different drugs solved in bovine plasma or in protein-free solutions the incidence of errors specific for each method was examined. Certain limits are found for the applicability of each method. If these limitations are observed reproducibility differs only slightly within the four methods. Considering all the results, however, the values achieved by equilibrium dialysis appear to come closest to the real extent of binding.
Daptomycin is a novel cyclic lipopeptide antibiotic that provides rapid bactericidal activity against gram-positive pathogens
in vitro, including methicillin-susceptible Staphylococcus aureus, methicillin-resistant S. aureus, vancomycin-resistant S. aureus, penicillin-resistant Streptococcus pneumoniae, and ampicillin- and vancomycin-resistant enterococci. The United States Food and Drug Administration recently approved daptomycin
for treatment of complicated skin and skin-structure infections. Its efficacy in the treatment of more-serious infections
(e.g., staphylococcal bacteremia) is under investigation. As an intravenous agent that is administered once per day, it offers
a convenient regimen for therapy that is continued after discharge, with a side effect profile that appears minimal and manageable.
Spontaneous acquisition of resistance in vitro is rare, and hopefully this characteristic will extrapolate into the clinical
setting. The rapid bactericidal activity, low potential for resistance, and promising safety profile associated with this
agent will make it a useful addition to our growing armamentarium of antibiotics active against gram-positive pathogens.
Single-dose daptomycin pharmacokinetics in chronic haemodialysis patients
- Salama