Objectives To evaluate the stability of solutions of 6 mg/mL, 8 mg/mL and 12 mg/mL meropenem in 0.9% NaCl in a polyolefin container (Viaflo), at room temperature, with a view to allowing extended or continuous perfusion in clinical practice.
Methods Meropenem was assayed using high-performance liquid chromatography. The technique was validated according to the criteria of the European Medicines Agency. Meropenem stability was evaluated for three different concentrations (6 mg/mL, 8 mg/mL and 12 mg/mL) at room temperature and during 48 h. The pH of the solution and its organoleptic properties were also evaluated.
Results The analytical technique abided with the specifications of the European Medicines Agency. The maximum accuracy and precision were 10% and 7.9%, respectively. The limit of detection was 0.000185 mg/mL and the limit of quantification 0.000562 mg/mL. The t90 for the dilution of 6 mg/mL of meropenem was 18 h (95% CI 14 h to 22 h), while the values for the 8 mg/mL and 12 mg/L concentrations were respectively 22 h (95% CI 19 to 32 h) and 17 h (95% CI 13 h to 21 h). The differences were not statistically significant (p<0.05). No variations in pH were seen.
Conclusions The results confirm the stability of the solutions of 6 mg/mL, 8 mg/mL and 12 mg/mL meropenem in 0.9% NaCl stored in a polyolefin container at 25°C for at least 12 h, thereby allowing their use as an extended or continuous perfusion.
- INFECTIOUS DISEASES
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The administered meropenem dose and the duration of treatment depend on the type of infection involved, its severity, and the patient response to therapy. The usual dose in the clinical setting is 1 g/8 h. In nosocomial infections produced by Pseudomonas aeruginosa or Acinetobacter spp., acute bacterial meningitis or critical patients with multiresistant infections, the recommended dose increases to 2 g/8 h.1
Meropenem is normally administered as an intermittent intravenous perfusion over approximately 15–30 min. This form of administration is associated with high peak serum meropenem concentrations and low minimum concentrations. However, the effectiveness of the carbapenems has been seen to be correlated to the time during which the drug concentrations exceed the minimum inhibitory concentration (MIC) of the causal micro-organism (T>MIC).2 In preclinical models, meropenem showed activity when the plasma concentrations exceeded the MIC of the causal organisms during approximately 40% of the dosing interval, though this objective has not been established in the clinical setting.3 The extended or continuous perfusion of meropenem would afford sustained serum concentrations above the MIC for a longer time interval than with intermittent administration, and would result in improved clinical outcomes.4 ,5
The carbapenems in general, and meropenem in particular, are quite unstable—a fact that can adversely affect extended or continuous drug perfusion. Meropenem stability is dependent upon the concentration, the diluent used, and the temperature to which the drug is exposed. Some stability studies have been carried out under various conditions: high concentrated meropenem solutions (>10 mg/mL) different from those commonly used in clinical practice, refrigerated solutions or stored solutions in elastomeric containers for ambulatory administration.6–10
The present study evaluates the stability of three solutions of 6 mg/mL, 8 mg/mL and 12 mg/mL meropenem in 0.9% NaCl in a polyolefin container (Viaflo), at room temperature, with a view to allowing extended or continuous perfusion in clinical practice.
Material and methods
For validation of the chromatographic technique and plotting of the calibration lines we used Meropenem Hospira (batch: 601D096; expiry date: May 2015) as test product, Cefotaxime Norman (batch: H2K41; expiry date: May 2015) as internal standard and 0.9% NaCl in a 500 mL Viaflo container (coextruded layers of polyethylene, polyamide, polypropylene) as diluent (batch:12L05E3Z; expiry date: February 2014). The mobile phase was prepared with bidistilled water (Fresenius), sodium hydrogen orthophosphate (Panreac PRS), phosphoric acid (Acofarma) for adjusting the pH to 2.4 and acetonitrile (Lichrosolv Merck)—all of analytical grade.
Meropenem was assayed in 0.9% NaCl solution using high-performance liquid chromatography (HPLC). The method constitutes an adaptation of the chromatographic technique described by McWhinney for biological samples,11 in which the protein precipitation steps were eliminated.
The chromatographic system was equipped with an automatic injector (Hitachi Autosampler L-220), an ultraviolet absorption detector (Merck Hitachi L4250) and a pump (Hitachi L-6200A), and was connected to a computer system operating the EZChrom Elite package for processing of the experimental data. As stationary phase we used a LiChroCART 250-4.6 RP-18 endcapped 25 cm column (internal particle diameter 5 µm).
The calibration line included five points with an interval of 0.125 mg/L to 2 mg/mL. The controls were prepared for concentrations of 0.003 mg/mL, 0.85 mg/mL and 1.5 mg/mL. The desired concentrations were obtained by serial dilution.
Validation of the analytical technique was carried out according to the recommendations of the European Medicines Agency.12 The accuracy and precision of the assay was validated by establishing the intra-assay and the interassay variations evaluated by relative error and coefficient of variation (CV), respectively. Interday and intraday accuracy and precision were performed using 15 determinations covering the specific range for the procedure (three concentrations/five replicates/three different days or 1 day). Linearity was evaluated by calculation of a regression line, by the method of least squares, between the concentrations and the areas of the chromatogram. Five concentrations were used. The range of the analytical procedure was from 0125 mg/mL to 2 mg/mL.
Chemical stability study
This study simulates the real life preparation of the intravenous mixture. Consequently, the initial volume of the intravenous solution and the amount of drug in the container were taken to be those stated by the manufacturer. Three solutions of different concentrations of meropenem were prepared: 6 mg/mL, 8 mg/mL and 12 mg/mL. Preparation of the intravenous mixtures of meropenem was carried out using polyolefin bags (Viaflo Baxter). The bags were stored at room temperature (25°C). To perform HPLC assays of meropenem, the samples were diluted (1:10) in order to fit to the established calibration curve.
Samples of 5 mL were collected from each of the bags at 30 min, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h, 36 h and 48 h. The pH values of the samples of each concentration were recorded, followed by dilution with 0.9% NaCl solution (1:10) for assay. Each concentration was prepared in triplicate. The preparations were visually inspected for possible precipitates or colour changes.
The pH values were recorded using a µ pH 2000 pH meter (Crison). The pH of the reconstituted solution was 7.3–8.3.1
The meropenem degradation process was defined as corresponding to first order kinetics. Stability was evaluated by determining the t90 value through semilogarithmic regression of the percentage remaining meropenem concentration versus time. Following the recommendations of the different international agencies, we determined t90 in each assay as the lower limit of the 95% CI; accordingly, at time 90 the remaining meropenem concentration is > 90%.
Values are expressed as mean and 95% CI. Statistical evaluation of the data was performed with SPSS (V.19.0; IBM SPSS Statistics, Chicago, Illinois, USA). A difference was considered statistically significant at p≤0.05.
Figure 1 shows the chromatograms corresponding to the highest concentration (2 mg/mL), lowest concentration (0.125 mg/mL) and sample without meropenem (blank) of the calibration line. The retention time of meropenem and cefotaxime is 3.8 min and 7.1 min, respectively. Figure 1 shows that there are no interferences for meropenem retention time.
Linearity was determined from the relationship between the concentrations and the areas of the chromatogram of the model, and was characterised by a regression straight line with an intercept of 0.0286 (95% CI −0.1650 to 0.2222) and a slope of 10.7482 (95% CI 10.5606 to 10.9357); the determination coefficient was 0.999 within the tested concentration range. The maximum accuracy (relative error) and precision values (CV) were 10% and 7.9%, respectively. The limit of detection was 0.000185 mg/mL and the limit of quantification 0.000562 mg/mL.
Table 1 shows the mean percentage of the remaining meropenem in the intravenous solutions stored at room temperature, at different sampling time points. At time zero, the initial meropenem concentration was regarded as 100%, and all the subsequent concentrations were expressed as a percentage of the initial concentration. An increase in the concentration of the individual 12 h samples can be observed. However, there is no statistically significant value (p>0.05) with respect to the 8 h samples.
Figure 2 shows the meropenem degradation curves. The t90 values and corresponding 95% CI were obtained by interpolation on the regression lines. The dilution t90 for 6 mg/mL meropenem was 18 h (95% CI 14 h to 22 h), while the values for the 8 mg/L and 12 mg/L concentrations were respectively 22 h (95% CI 19 h to 32 h) and 17 h (95% CI 13 h to 21 h). The differences were not statistically significant (p>0.05). All three tested concentrations presented a t90 value of at least 13 h.
No precipitates, solid particles or gas were observed during the study period at any of the formulated concentrations. However, a slight variation in colour was observed in the preparations after 12 h, this change being more manifest after 24 h.
The variation in pH value is shown in figure 3. In all cases the pH decreased slightly, though in no case was the variation greater then 0.2.
The analytical technique used to evaluate the stability of meropenem in 0.9% NaCl allows adequate separation of the drug and offers suitable accuracy and precision, according to the criteria and methods established by the European Medicines Agency.11 ,12 Although a CV of 7.9% for this method could seem to be rather high, this is the value corresponding to the lowest concentration tested. The CVs for the other two measurements were 2.9% and 0.7%. For bioanalytical methods, CVs (indicating precision) lower than 15% at all concentrations are accepted.13
Meropenem at concentrations of 6 mg/mL, 8 mg/mL and 12 mg/mL remains stable for at least 12 h stored at room temperature in a polyolefin container. This stability would allow its administration in extended perfusion, improving the percentage time during which the serum meropenem concentrations exceed MIC with respect to intermittent administration. Meropenem displays time-dependent pharmacodynamics. Maximal bacterial killing for meropenem is thought to be related with the time that bacteria are exposed to a concentration higher than the MIC.14
According to some authors, the percentage time during which the serum drug concentrations exceed MIC should be at least 40%, that is, about 10 h a day.3 ,5 It is true that concentrations reached into the tissues will be determinant because antibiotic-bacteria interactions occurs at this level. One randomised controlled trial reported that continuous infusion of meropenem maintained higher trough concentrations in plasma and subcutaneous tissue in 10 critically ill patients, and tissue penetration. The authors also reported a higher tissue penetration with continuous infusion (89% vs 74%).15
Continuous or extended perfusion has been associated with a reduction in total meropenem dose, a shortening of treatment time, and improved clinical and bacteriological outcomes.4 ,16 Dulhunty et al17 observed an improvement in clinical cure in patients treated with continuous infusion of piperacillin-tazobactam, meropenem or ticarcillin-clavulanate versus intermittent bolus dosing (70% vs 43%, p=0.037). On the other hand, some publications reported no differences in clinical outcomes.18 ,19 Recently, a meta-analysis evaluated clinical outcomes with extended (≥3 h) or continuous (24 h) versus short-term intravenous infusion of carbapenems and piperacillin/tazobactam. Mortality was lower among patients receiving extended or continuous infusion of carbapenems or piperacillin/tazobactam compared with those receiving them for a short-term (risk ratio (RR), 0.59; 95% CI 0.41 to 0.83). Differences were favourable but not statistically significant with carbapenems probably due to the lower sample size.20
Randomised controlled trials conducted to date comparing the mode of β-lactam administration are limited by non-equivalent dosing, unblinded administration and small sample sizes. Well-designed randomised controlled trials are warranted to confirm these findings before such approaches become widely used. Thus, a multicentre, randomised controlled trial is currently under way, comparing continuous infusion with standard bolus administration of β-lactam antibiotics in critically ill patients.21
Clinicians need to know about limitations in the use of meropenem in extended infusions. The result of 12 h stability is only applicable for a preparation before use. The calculated stability may not be a major advantage if preparation is centralised and the supply chain long. The degradation process is concentration-dependent and temperature-dependent. The transportation of a meropenem preparation at not-cooled conditions or the continuous perfusion at temperatures exceeding 25°C (lack of climatisation at the wards) increased the risk of degradation. At body temperature, meropenem undergoes 55–70% degradation within 24 h. The degradation process is concentration-dependent and temperature-dependent. However, when the drug is stored at 4–5°C, stability is maintained for over 24 h.22
Smith et al6 evaluated the stability of meropenem at concentrations of 4 mg/mL, 10 mg/mL and 20 mg/mL in polyvinyl chloride bags and in elastomeric intravenous perfusion equipment at 5°C. The minimum stability was 7 days. Similar results have been published by other authors.8 ,23 Krämer tested the stability of meropenem (5 mg/mL, 10 mg/mL) at normal storage temperatures (room temperature, refrigerated) in elastomeric portable infusion devices.9 Solutions were affected by the concentration and by the temperature. Under refrigeration both solutions were stable for 120 h. After 96 h of refrigeration, a solution of 5 mg/mL was stable at least for 6+ h at room temperature, but when the concentration was 10 mg/mL, the maximum stability was 6 h at room temperature. Influence of diluent (0.9% NaCl, 5% dextrose), concentration (1 mg/mL, 22 mg/mL) and temperature (−20, 4, 23°C) on stability of meropenem solutions was evaluated by Walker.10 Meropenem was more stable in 0.9% NaCl than in 5% dextrose. Solutions of meropenem 1 mg/mL in 0.9% NaCl stored at 23°C were stable for 22 h, at 4°C for 7 days and at −20°C for 14 days.
Kuti et al24 performed a study that combines a stability study of meropenem (30 mg/mL) in a continuous ambulatory drug-delivery infusion pump and a pharmacokinetic analysis in seven adult patients with cystic fibrosis. Meropenem solutions were prepared within 4 h of dosing and refrigerated at 0°C until administration to maintain drug stability during infusions; the pumps were stored in a cold pouch between two freezer packs. Patients wore the cold pouch during the administration. Meropenem stability was maintained, at least, for 12 h. All patients reached serum drug concentrations higher than MIC for pathogens considered susceptible or intermediately resistant to meropenem.
Berthoin et al7 reported a stability of under 12 h for meropenem solutions with a concentration of over 40 mg/mL at 25°C. After 12 h, meropenem 10 mg/mL maintains 90% of its initial concentration; however, the lower limit of the CI indicates degradation of the drug in excess of the 10% required by the European Pharmacopoeia.
It is important to know the factors that affect the stability of meropenem. Control of the drug concentration is simple, though control of the environmental temperature is more complicated. The administration of extended perfusions of meropenem must be made under controlled temperature conditions, avoiding exposure to heat or even nearness to the patient.
Meropenem exhibits apparent first order degradation kinetics,25 giving rise to inactive products secondary to alteration of the lateral chain and β-lactam ring and polymers.9 ,25 ,26 The degradation products lack bactericidal activity.27 The chromatogram of meropenem 1.2 mg/mL shown in figure 4 exhibits two absorbance peaks after 12 h. These peaks probably correspond to the mentioned degradation products, and increase after 24 h, 36 h and 48 h. Mass spectrometry would be needed to characterise these degradation products. Takeuchi et al investigated the degradation products of meropenem in aqueous solution. The results of HPLC and spectroscopic analysis revealed that these products were the β-lactam hydrolysed product and a dimer formed by intermolecular aminolysis of the β-lactam ring. The thermal and alkaline degraded samples of meropenem reconstituted solution were studied to determine the cytotoxicity in vitro against mononuclear cells. The results obtained indicated that samples could be toxic in high concentration (2.0 mg/mL) after 48 h of incubation.26
To summarise, the results obtained confirm the stability of the solutions of 6 mg/mL, 8 mg/mL and 12 mg/mL meropenem in 0.9% NaCl stored in a polyolefin container at 25°C for at least 12 h, thereby allowing their use as an extended or continuous perfusion.
What is already known on this subject
Meropenem is quite unstable—a fact that can adversely affect extended or continuous drug perfusion. Meropenem stability is dependent upon the concentration, the diluent used, and the temperature to which the drug is exposed. Stability studies have been carried out with highly concentrated meropenem or under various conditions different from those used in clinical practice.
What this study adds
The results confirm the stability of the solutions of 6 mg/mL, 8 mg/mL and 12 mg/mL meropenem in 0.9% NaCl stored in polyolefin containers at 25°C for at least 12 h, thereby allowing their use in extended or continuous perfusion.
Contributors All authors have participated sufficiently in the work to take public responsibility for appropriate portions of the content. SM-C: Acquisition of data, analysis and interpretation of data, revised the manuscript critically for important intellectual content and final approval of the version published. RF-L, TT-J and MA-A: Conception and design, analysis and interpretation of data, Drafting the article and revising it critically for important intellectual content, final approval of the version published.
Competing interests None.
Provenance and peer review Not commissioned; externally peer reviewed.
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