Prediction of the Stability of Meropenem in Intravenous Mixtures

Abstract

The purpose of this study was to predict the stability of meropenem in a mixed infusion. The hydrolysis of meropenem in aqueous solution was found to be accelerated by pH, and by increasing concentrations of sodium bisulfite (SBS) and L-cysteine. Equations were derived for the degradation rate constants (k_obs) of pH, SBS and L-cysteine, and fractional rate constants were estimated by the nonlinear least-squares method (quasi-Newton method using the solver in Microsoft Excel) at 25°C. The activation energy (E_a) and frequency factor (A) were calculated using the Arrhenius equation. The pH of the mixed infusion was estimated using the characteristic pH curve. From these results, an equation was derived giving the residual ratio (%) of meropenem at any time after mixing an infusion containing SBS and/or L-cysteine at any temperature, and in the pH range 4.0–10.0. A high correlation was shown to exist between the estimated and determined residual ratios (%).

Carbapenems are the most potent β-lactam antibiotics and were first developed in the 1980s to enhance resistance to β-lactamases. The early carbapenems were not very hydrolytically stable, however, limiting drug administration to controlled intravenous infusions. In the search for a more stable compound with better toxicity profile, the basic nuclear structure was maintained and a dimethyl carbamoyl-pyrrolidinylthio group was added as a weakly basic C-2 side-chain. This resulted in a reduction of central toxicity and nephrotoxicity, while maintaining anti-pseudomonal activity. In addition, the improvement of stability (DHP-I stability) in vivo and the improvement of activity against Haemophilus influenzae, etc., were achieved by introducing 1-β-methyl group into the carbapenem frame, resulting in the creation of meropenem. The improvement in nephrotoxicity was the key advance in allowing the global development of meropenem as a single agent.¹⁾

Meropenem is a broad-spectrum carbapenem, active against several clinically relevant Gram-positive and Gram-negative aerobes, anaerobic bacteria and Pseudomonas aeruginosa. The bactericidal activity of meropenem results from the inhibition of cell wall synthesis through the inactivation of penicillin-binding proteins.

Sodium bisulfite (SBS), used as a stabilizer in injectable preparations, is known to degrade various drugs including thiamine,²⁾ epinephrine,³⁾ gabexate mesilate,⁴⁾ nafamostat mesilate,⁵⁾ urokinase,⁶⁾ morphine⁷⁾ and fluorouracil.⁸⁾ However, there are no reports of detailed kinetic studies on the degradation of meropenem in the presence of SBS.

The prediction of the stability of a drug in an intravenous admixture (mixed infusion) is important for accurate and safe drug administration. Generally, the stability of a drug in a mixed infusion can be predicted from the pH profile and Arrhenius equation of the degradation rate constants, if the temperature and pH of the test solution are known. Although meropenem is generally administered as an infusion in saline, in some cases it may be mixed with other substances, such as amino acids.⁹⁾

A method for predicting the pH of mixed infusions was developed in order to be able to evaluate the compatibility of the components of a mixed infusion. Employing the characteristic pH curve (PHC curve; a kind of pH titration curve),¹⁰⁾ a method for pH estimation has been reported by Hirouchi et al.¹¹⁾ based on computer simulations. However, the theoretical equations described by Hirouchi et al. were derived from model preparations for injection and did not take into account the influence of dilution with titrant. The PHC curve of a mixed infusion could not therefore be fitted to the equations of Hirouchi et al., but must be derived from observed values, a painstaking and time-consuming exercise. In the present study, a simple, theoretical, pH estimation method for mixed infusions was derived, using a fitted PHC curve and curve simulation by a computer (using Microsoft Excel).

In addition, we examined the degradation of meropenem due to catalytic hydrolysis by SBS and L-cysteine. From the results, we derived an equation to predict the residual ratio (%) of meropenem at any time after mixing, on the basis of the simulated pH of the mixed infusion using the PHC curve, SBS and L-cysteine concentrations, and the temperature.

Experimental

Materials

Meropenem was supplied by Wako Pure Chemical Industries, Ltd., Osaka, Japan. SBS, L-cysteine and other reagents were special grade commercial products. As buffer solutions, 0.05 M acetate buffer (pH 4.0–5.0), 0.05 M phosphate buffer (pH 6.0–8.0) and 0.05 M carbonate buffer (pH 9.0–10.0) were adjusted to an ionic strength of 0.15 with sodium chloride. Meropen® (Dainippon Sumitomo Pharma Co., Ltd., Osaka, Japan), AMINOLEBAN_® Injection, Fructlact Injection, and AMINOFLUID_® Injection (Otsuka Pharmaceutical Factory, Inc., Tokushima, Japan), FULCALIQ® 2 and AMIGRAND_® (Terumo Co., Ltd., Tokyo, Japan) were used as injectable preparations.

Determination of Meropenem

Meropenem concentration was measured by high-performance liquid chromatography (HPLC). Meropenem concentrations were determined 0, 1, 2, 3 and 4 h after mixing with SBS or L-cysteine. Each degradation rate constant (k_obs) was calculated from the slope of the relationship between the log residual ratio of meropenem (%) and time after mixing.

Equipment

A Shimadzu LC-10AT VP high-performance liquid chromatograph, a Shimadzu SPD-10A VP UV-VIS detector, and a Shimadzu C-R7A plus chromate pack were used.

Measurement Conditions

Preliminary studies confirmed that optimized chromatographic conditions were as described in the Pharmacopoiea.¹²⁾ Column, CAPCELL PAK C₁₈ HPLC packed column type MG 5 µm (4.6 mm i.d.×150 mm); flow rate 1.0 mL/min; column oven set at 40°C; injection volume 20 µL; detection wavelength 300 nm. The isocratic mobile phase was a mixture of 10 mM sodium phosphate buffer (pH 7.4) and acetonitrile (100 : 10, v/v).

Estimation of the pH of the Mixed Infusion

Observed values from pH titration were used to plot the PHC curve of the mixed infusion. Based on the general equation for PHC curves, the PHC curve of each preparation was fitted from the observed values, from which the pH of the mixed infusion could be estimated.

pH Titration

Various commercial injections were diluted to 500 mL with distilled water and titrated against 0.5 M hydrochloric acid and sodium hydroxide. Similarly, 500 mL aliquots of various commercial infusions were titrated against 0.5 M hydrochloric acid and sodium hydroxide.

Fitting the PHC Curve of the Injection Preparation

Injectable drugs are generally classified as weak acids, weak bases or salts. Taking a preparation of weak acid, with a total concentration of C_a1, the ionization of the weak acid may be represented as: HA→H⁺+A⁻, where HA is the non-ionized acid, A⁻ is the ionized acid and H⁺ is hydrogen ion. Applying the law of mass action, material balance and charge balance to the equilibrium of this weak acid, the general equation of the PHC curve for a preparation can be expressed as in Eq. 1. Similarly, taking a preparation containing two kinds of weak acids of which the total concentrations are C_a1 and C_a2, the general equation for the preparation can be expressed as in Eq. 2. If a preparation contains n kinds of weak acids, the general equation of the PHC curve for the preparation can be expressed as in Eq. 3.   

(Eq. 1)

  

(Eq. 2)

  

(Eq. 3)

In order to improve solubility, preparations of weak bases often contain a strong acid as an excipient. The preparation of the weak base can therefore be assumed to be a preparation of a conjugated weak acid. Accordingly, the general equation of the PHC curve for a preparation of a weak base can be expressed as in Eq. 1, and the PHC curve of the preparation of n kinds of weak bases can be represented as in Eq. 3.

The general equation of the PHC curve for injectable preparations can therefore, for practical purposes, be represented as Eqs. 1 to 3, and the influence of dilution with titrants can be corrected by Eqs. 4 to 6.   

(Eq. 4)

  

(Eq. 5)

  

(Eq. 6)

where C_a1, C_a2, C_a3, ..., C_an are the concentrations of weak acids 1, 2, 3, ..., n in the sample solution; K₁, K₂, K₃, ..., K_n are the dissociation constants of the weak acids 1, 2, 3, ..., n, respectively; C is the concentration of the strong acid (represented as a positive value) or the base (negative value) present as an excipient in the sample solution; C_t is the concentration of HCl (represented as a positive value) or NaOH (negative value), added as the titrant to the sample solution; C′_an is the initial concentration of the acid (positive value) or base (negative value) in the sample solution, respectively; C′_t is the concentration of HCl or NaOH added to the sample solution as titrant; V₀ is the initial volume of the titration sample (500 mL) and V_t is the volume of the titrant added to the sample. [H⁺] is the hydrogen ion concentration and K_w is the ion product of water. For practical purposes, [H⁺] and K_w at 25°C were regarded as 10^−pH obs and 1.0×10⁻¹⁴, respectively.

The parameters of the general equations for the PHC curve (i.e., concentrations and pKs of weak acids and concentrations of strong acids or bases) were fitted with some of the observed values. The parameters of the PHC curve were then obtained by solving the simultaneous equation Eq. 2. Next, the parameters other than pK were fitted by the nonlinear least-squares method (simplex method). The PHC curves simulated by these parameters and the observed titration values were drawn graphically. If the PHC curve only partially fitted, the parameters of the PHC curve were modified by the simultaneous equation Eq. 1 and the simplex methods.

Simulation of the PHC Curve of Mixed Infusions

The effect of dilution was an important factor when the parameters of the PHC curve were fitted to the titration data. Nevertheless, once these parameters had been obtained, the PHC curves of the preparations could be treated simply according to Eqs. 1 to 3, without Eqs. 4 to 6. Therefore, assuming the general equations of the PHC curves for an injection, an infusion, a mixed infusion of these preparations and water, to be Eqs. 7 to 10, respectively, the relationship Eq. 9=Eq. 7+Eq. 8−Eq. 10 held at a constant ion concentration. Each of the terms except C_tA, C_tB, C_tW and C_tM could then be eliminated from Eqs. 7 to 10 at a constant hydrogen ion concentration, to obtain Eq. 11. Since each PHC curve represents the relation between the pH and the volume of the titrants (i.e., C_tM), the PHC curves of mixed infusions could be simulated by applying Eq. 11 at all pHs. If n kinds of preparations were mixed, the PHC curve of the mixed infusion could be simulated from the PHC curves of n−1 kinds of preparations and another additive injection.   

(Eq. 7)

  

(Eq. 8)

  

(Eq. 9)

  

(Eq. 10)

  

(Eq. 11)

Revision of the PHC Curve for Increased Infusion Volumes

The influence of dilution with additive injections is not included in Eq. 11. If the infusion volume is increased with additive injections, the PHC curves of the mixed infusion must be corrected by Eq. 12. This equation was obtained following the same process as for Eq. 11.   

(Eq. 12)

where r is the concentration ratio (initial infusion volume (500 mL)/increased infusion volume with additives), C_tC is the corrected concentration of HCl or NaOH (added as the titrant) in the mixed infusion at a constant pH, C_t is the concentration of HCl or NaOH in a 500 mL aliquot of the mixed infusion at the same pH, and C_tW is the concentration of HCl or NaOH in a 500 mL aliquot of distilled water at the same pH. At C_t and C_tW, the influence of dilution with additive injections was omitted.

Simulation of the PHC Curve of a Mixed Infusion

According to Eqs. 11 and 12, the PHC curve of the mixed infusion can be simulated from the PHC curves of each constituent preparation.

Estimated pH of a Mixed Infusion

The pHs of the injections (diluted to 500 mL with distilled water) or infusions are represented by the pHs at the origin of the PHC curve of the preparations. The estimated pH of the mixed infusion was thus obtained as the pH at the origin of the simulated PHC curve.

Kinetic Procedures

The stability of the meropenem solution (500 µg/mL) buffered at pH 4.0–10.0 in the presence of SBS (0, 0.1, 0.5 and 1.0 mM) or L-cysteine (0, 0.25, 0.5, 1.0 and 2.0 mg/mL) was examined at 4, 25 and 40°C. The stability of Meropen® in AMINOLEBAN_® Injection, Fructlact Injection, AMINOFLUID_® Injection, FULCALIQ® 2 and AMIGRAND_® was examined at 4, 25 or 40°C.

Results and Discussion

Factors Affecting the Stability of Meropenem

Degradation of Meropenem at pH 4.0–10.0

The degradation of mepenem, 500 µg/mL, was studied at 25°C. The pH-profile of mepenem degradation in the pH range 4.0–10.0 is shown in Fig. 1, where the rate constants are expressed on a logarithmic scale. It was concluded that specific hydrogen-ion-catalyzed degradation occurred at pH 4.0–5.0, water-catalyzed degradation at pH 5.0–8.0, and hydroxide-ion-catalyzed degradation at pH 8.0–10.0.

Fig. 1. pH Profile of the Degradation of Meropenem at 25°C and μ=0.15

Initial concentration of meropenem 500 µg/mL.

As the data plot in Fig. 1 seemed to display specific acid–base catalysis kinetics, Eq. 13 was used as the model kinetic equation for the effect of pH on k₀.   

(Eq. 13)

where k_H⁺ and k_OH⁻ are the second-order rate constants for the hydrogen-ion-catalyzed degradation and the hydroxide-ion-catalyzed degradation reaction, respectively, and k_H2O is the first-order rate constant for the spontaneous water-catalyzed degradation reaction.

At 25°C, the residual meropenem concentration as determined by HPLC, and [H⁺], [OH⁻] calculated from pH, were assigned to Eq. 13. The fractional rate constants, k_H2O, k_OH⁻, and k_H⁺ were estimated by the nonlinear least-squares method (quasi-Newton method using the solver in Microsoft Excel) and determined to be: k_H2O, 1.15×10⁻³ h⁻¹; k_OH⁻, 3.00×10³ M⁻¹ h⁻¹ and k_H⁺, 1.36×10² M⁻¹ h⁻¹.

The influence of temperature was also investigated. The Arrhenius equation (Eq. 14) shows the relationship between the degradation rate constant and the absolute temperature.   

(Eq. 14)

where k_obs is the degradation rate constant, E_a is the activation energy, R is the gas constant (1.987 cal mol⁻¹), T is absolute temperature (K), and A is the frequency factor. As shown in Fig. 2, Arrhenius plots revealed good linearity in the range pH 4.0–10.0, with E_a values for mepenem of 10.7 kcal/mol (pH 4.0), 16.1 kcal/mol (pH 5.0), 15.9 kcal/mol (pH 6.0), 12.9 kcal/mol (pH 7.0), 14.8 kcal/mol (pH 8.0), 20.6 kcal/mol (pH 9.0) and 16.7 kcal/mol (pH 10.0).

Fig. 2. Arrhenius-Type Relationship between the Degradation Rate Constant of Meropenem and the Temperature (4, 25 and 40°C) at pH 4.0–10.0 and μ=0.15

Initial concentration of meropenem 500 µg/mL. ◇ pH 4.0, * pH 5.0, ◆ pH 6.0, × pH 7.0, ▲ pH 8.0, ○ pH 9.0, ■ pH 10.0.

Degradation Rate of Meropenem in the Presence of SBS

SBS, present as a stabilizer in injectable preparations, is known to degrade β-lactam antibiotics.¹³⁾ However, the mechanism by which the SBS concentration affects meropenem degradation has not yet been elucidated. Therefore, kinetic experiments were performed, and the residual meropenem concentration measured by HPLC. The degradation of meropenem, at an initial concentration of 500 µg/mL, by SBS at various concentrations was evaluated at 25°C and pH 4.0–10.0. The degradation rate constant of meropenem in the presence of SBS (k_obs) was measured at 25°C. A linear relationship was observed between time and log residual ratio (%) at pH 6.0 (Fig. 3), indicating that the degradation of meropenem followed pseudo-first-order kinetics. This was also the case at other pHs. The apparent first-order rate constants were obtained from the slopes of the semi-log plots. The slope of the line increased with increasing SBS concentration.

Fig. 3. Pseudo-First-Order Plot for Degradation of Meropenem in the Presence of SBS (0, 0.1, 0.5 and 1.0 mM) in 0.05 M Phosphate Buffer (pH 6.0, μ=0.15) at 25°C

Initial concentration of meropenem 500 µg/mL. ◇ SBS 0 mM, ○ SBS 0.1 mM, △ SBS 0.5 mM, □ SBS 1.0 mM.

The rate constant for the catalytic hydrolysis of meropenem by SBS (k_SBS) is obtained from Eq. 15.   

(Eq. 15)

where k_obs is the pseudo-first-order reaction rate constant, t is time after mixing, and r is the residual ratio (%). Typical plots for SBS concentration versus degradation rate constants of meropenem (pH 6.0, 25°C) yielded a straight line as shown in Fig. 4. The degradation rate of meropenem was proportional to the total concentration of SBS ([SBS]_total). Therefore, the rate constant of meropenem in the presence of SBS (k_obs) can be represented by Eq. 16.   

(Eq. 16)

where SBS concentration ([SBS]_total) is represented by Eq. 17.⁴⁾   

(Eq. 17)

Fig. 4. Relationship between the Total Concentration of SBS and the Degradation Rate Constant (k_obs) of Meropenem in 0.05 M Phosphate Buffer (pH 6.0, μ=0.15) at 25°C

Initial concentration of meropenem 500 µg/mL.

Based on the dissociation constant of SBS at 25°C (100 kPa), k_SBS1=1.72×10⁻² (pK_SBS1=1.8) and k_SBS2=6.24×10⁻⁸ (pK_SBS2=7.2).¹⁴⁾ SBS in this pH range (pH 4.0–10.0) was considered to be present as bisulfate ions (HSO₃⁻) and sulfite ions (SO₃²⁻), k_SBS×[SBS]_total can therefore be represented as Eq. 18:   

(Eq. 18)

and the dissociation constants of SBS (K_SBS1, K_SBS2) can be represented by Eqs. 19 and 20.   

(Eq. 19)

  

(Eq. 20)

From Eqs. 16 to 20, k_SBS can be represented as shown in Eq. 21.   

(Eq. 21)

where k_HSO3⁻ is the second order degradation constant of HSO₃⁻, and k_SO3²⁻ is the second order degradation constant of SO₃²⁻. The fractional rate constants, k_HSO3⁻ and k_SO3²⁻ were estimated by nonlinear least-squares method (quasi-Newton method using the solver in Microsoft Excel), and values of k_HSO3⁻ (2.78 M⁻¹ h⁻¹) and k_SO3²⁻ (2.75 M⁻¹ h⁻¹) were obtained. These results show that the accelerating effect of sulfite ions on the hydrolysis of meropenem was the same as that of bisulfate ions at pH 4.0–10.0 and 25°C.

The influence of temperature on the degradation of meropenem by SBS was also investigated. As shown in Fig. 5, Arrhenius plots revealed good linearity between SBS concentrations of 0, 0.1, 0.5 and 1.0 mM at pH 6.0, and the values obtained for the E_a of meropenem,15.9 kcal/mol, 15.7 kcal/mol, 16.5 kcal/mol, and 14.7 kcal/mol, respectively.

Fig. 5. Arrhenius-Type Relationship between the Degradation Rate Constant of Meropenem and Temperature (4, 25 and 40°C) in the Presence of SBS (0, 0.1, 0.5, 1.0 mM) in 0.05 M Phosphate Buffer (pH 6.0, μ=0.15)

Initial concentration of meropenem 500 µg/mL. ◇ SBS 0 mM, ○ SBS 0.1 mM, △ SBS 0.5 mM, □ SBS 1.0 mM.

Degradation Rate of Meropenem in the Presence of L-Cysteine

L-Cysteine is often added to infusions for purposes of nutrition. It may be present as the free amino acid or as N-acetylcysteine, which can be degraded by hydrolysis in the body. L-Cysteine is known to degrade carbapenem antibiotics, but the effect of the L-cysteine concentration on the rate of degradation has not been elucidated. Kinetic experiments were therefore carried out, with the residual meropenem concentrations measured by HPLC. The degradation of meropenem at an initial concentration of 500 µg/mL by various concentrations of L-cysteine or N-acetylcysteine was evaluated at 25°C and pH 4.0, 6.0 and 8.0. The degradation rate constant of meropenem in the presence of L-cysteine (k_cys) was measured at 25°C. Typical plots for L-cysteine concentration versus degradation rate constants of meropenem yielded a straight line as shown in Fig. 6. N-Acetylcysteine concentrations were converted to L-cysteine concentrations. N-Acetylcysteine itself appeared to have little effect on the degradation rate constants of meropenem. The degradation rate of meropenem was directly proportional to the total concentration of L-cysteine ([cys]_total). Based on the dissociation constant of L-cysteine at 25°C, k_cys1=1.20×10⁻² (pK_cys1=1.92), k_cys2=4.68×10⁻⁹ (pK_cys2=8.33) and k_cys3=1.66×10⁻¹¹ (pK_cys3=10.78),¹⁵⁾ the degradation constant of meropenem in the presence of cysteine (k_obs) can be represented as Eq. 22, [cys]_total as Eq. 23 and k_cys×[cys]_total as Eq. 24.   

(Eq. 22)

  

(Eq. 23)

  

(Eq. 24)

The fractional rate constants, k_HSO3⁻ and k_SO3²⁻ were estimated by the nonlinear least-squares method (quasi-Newton method using the solver in Microsoft Excel) to obtain the following values: k_{COO⁻·NH3⁺·SH} (2.48×10 M⁻¹ h⁻¹), k_{COO⁻·NH3⁺·S⁻} (1.20×10⁴ M⁻¹ h⁻¹) and k_{COO⁻·NH2·SH⁺} (3.31 M⁻¹ h⁻¹). The value of k_{COOH·NH3⁺·SH} was much smaller than the other fractional rate constants, so k_{COOH·NH3⁺·SH} was omitted from the calculations, as it seems to have little effect on the degradation rate constants of meropenem.

Fig. 6. Relationship between the Total Concentrations of L-Cysteine (0, 0.25, 0.5, 1.0, 2.0 mg/mL) and N-Acetylcysteine (0, 0.25, 0.5, 1.0, 2.0 mg/mL; Converted to L-Cysteine Concentration) and the Degradation Rate Constant (k_obs) of Meropenem in 0.05 M Phosphate Buffer (pH 6.0, μ=0.15) at 25°C

Initial concentration of meropenem 500 µg/mL. ○ N-acetylcysteine, ● L-cysteine.

The influence of temperature on the degradation of meropenem by L-cysteine was also investigated. As shown in Fig. 7, Arrhenius plots revealed good linearity at L-cysteine concentrations of 0, 0.25, 0.5, 1.0 and 2.0 mg/mL at pH 6.0. The values obtained for the E_a of meropenem were 11.7 kcal/mol (0.25 mg/mL), 11.0 kcal/mol (0.5 mg/mL), 10.4 kcal/mol (1.0 mg/mL) and 10.6 kcal/mol (2.0 mg/mL), while the value obtained for the E_a of meropenem was 16.0 kcal/mol in the absence of L-cysteine (0 mg/mL). There were no differences between the values of E_a in presence of L-cysteine (0.25, 0.5, 1.0, 2.0 mg/mL). However, there was difference between the values of E_a in the presence of L-cysteine and the value of E_a in the absence of L-cysteine. The dose-dependent effect of L-cysteine on the E_a of meropenem in this reaction is suggested to be small. The log k_obs of meropenem showed a temperature-dependent increase at any concentration of L-cysteine. From these results, the degradation of meropenem is suggested to increase in a temperature-dependent manner.

Fig. 7. Arrhenius-Type Relationship between the Degradation Rate Constant of Meropenem and Temperature (4, 25 and 40°C) in the Presence of L-Cysteine (0, 0.25, 0.5, 1.0, 2.0 mg/mL) in 0.05 M Phosphate Buffer (pH 6.0, μ=0.15)

Initial concentration of meropenem 500 µg/mL. ◇ L-cysteine 0 mg/mL, ○ L-cysteine 0.25 mg/mL, △ L-cysteine 0.5 mg/mL, □ L-cysteine 1.0 mg/mL, × L-cysteine 2.0 mg/mL.

pH Estimation Method for a Mixed Infusion

After mixing Meropen® with AMINOLEBAN_® Injection, Fructlact Injection, AMINOFLUID_® Injection or FULCALIQ® 2 or AMIGRAND_®, pH was estimated at 25°C using the PHC curve. Figure 8 shows the correlation between the estimated and determined pH values in the mixed infusion (y=1.00x, r²=0.98). The pH of each mixed infusion could be estimated correctly using the PHC curve.

Fig. 8. Correlation between pH Estimated Using PHC Curve and the pH Values Determined after Mixing Meropen® with AMINOLEBAN_® Injection (), Fructlact Injection (×), AMINOFLUID_® Injection (), FULCALIQ® 2 () or AMIGRAND_® () at 25°C

y=1.00x, r²=0.98.

Stability Prediction of Meropenem in Practical Mixed Infusions

From the above results, the degradation rate constant of meropenem after mixture with an infusion contained SBS and L-cysteine at pH 4.0–10.0, can be represented by Eq. 25.   

(Eq. 25)

where E_a1 is the activation energy in the presence of the acid–base catalytic effect, E_a2 is the activation energy in the presence of SBS, E_a3 is the activation energy in the presence of L-cysteine; k_obs at any temperature (T₂) can be calculated if 298 K (25°C) is assigned to T₁ and the derived concentrations and constants at 298 K (25°C) and activation energy values (E_a1 to E_a3) are assigned to Eq. 25. The meropenem residual ratio (%) is represented by Eq. 26:   

(Eq. 26)

From Eqs. 25 and 26, the meropenem residual ratio (%) can be predicted at any time after mixing into an infusion containing SBS and L-cysteine at pH 4.0–10.0.

The correlation between the estimated and determined residual ratio (%) of meropenem in Meropen® at 0, 1, 2, 3 and 4 h after mixing with AMINOLEBAN_® Injection, Fructlact Injection, AMINOFLUID_® Injection, FULCALIQ® 2 or AMIGRAND_® at 4, 25 and 40°C (y=1.00x, r²=0.98) is shown in Fig. 9. The initial concentration of meropenem was 500 µg/mL. The high correlation coefficient q² (0.99), shows a high correlation between the estimated and determined residual ratios (%) of meropenem obtained in this study, which is relevant for the drug combinations used in the clinical field.

Fig. 9. Correlation between Estimated and Observed Residual Ratio (%) after Mixing Meropen® with AMINOLEBAN_® Injection, Fructlact Injection, AMINOFLUID_® Injection, FULCALIQ® 2 and AMIGRAND_®, at 4, 25 and 40°C

Initial concentration of meropenem 500 µg/mL. y=1.00x, r²=0.98. ○ AMINOLEBAN_® Injection at 4°C, — Fructlact Injection at 4°C, □ AMINOFLUID_® Injection at 4°C, ◇ FULCALIQ® 2 at 4°C, △ AMIGRAND_® at 4°C. AMINOLEBAN_® Injection at 25°C, × Fructlact Injection at 25°C, AMINOFLUID_® Injection at 25°C, FULCALIQ® 2 at 25°C, AMIGRAND_® at 25°C. ● AMINOLEBAN_® Injection at 40°C, + Fructlact Injection at 40°C, ■ AMINOFLUID_® Injection at 40°C, ◆ FULCALIQ® 2 at 40°C, ▲ AMIGRAND_® at 40°C.

These results confirm that an equation predicting the stability of meropenem, at any time and any temperature, after mixing with an infusion containing SBS and L-cysteine at pH 4.0–10.0 has been derived in this study.

We have previously derived an equation predicting the stability of imipenem, at any time and any temperature, after mixing with an infusion containing SBS and L-cysteine at pH 4.0–10.0.¹⁶⁾ Both the k_SBS and k_cys of meropenem were smaller than those of imipenem. This means that meropenem is more stable than imipenem after mixing with an infusion containing SBS and L-cysteine, probably because hydrolysis of the β-lactam ring of imipenem occurs more easily than that of meropenem, due to its chemical structure.

Itagaki et al.¹⁷⁾ reported on the influence of L-cysteine on the degradation of meropenem after mixing with an amino acid infusion. In their report, N-acetyl cysteine had no effect on the degradation of meropenem after mixing with an amino acid infusion, thus supporting the result in the present paper. It is assumed that N-acetyl cysteine is unable to mount a nucleophilic attack on the β-lactam structure of meropenem because the electron density on the S atom of N-acetyl cysteine is significantly lower than that of L-cysteine, due to the presence of the acetyl group, an electron-accepting substituent.

SBS is included in amino acid infusions as a stabilizer. In the present study, it is suggested that SBS affects the degradation of meropenem after mixing with the amino acid infusion. Thus the influence of not only L-cysteine but also SBS must be considered when attempting to improve the precision of prediction of the stability of meropenem in mixed infusions.

Our results indicate that, if antibiotics are administered as an infusion using a bypass line, they may become unstable if mixed with an infusion in the infusion line or by back-flow from the bypass line to the main line. Yoshioka et al. reported that the effectiveness of doripenem was decreased by a bypass administration from amino acid infusion line.¹⁸⁾ We suggest that the effectiveness of meropenem may also be decreased by a bypass administration from an amino acid infusion line. The required minimum dosage and length of treatment should therefore take account of this interaction between carbapenem antibiotics and amino acid infusions, in order to maximise the effectiveness of the antibiotics and avoid the appearance of resistant bacteria. The equation derived in this study will enable prediction of the required concentration of meropenem in mixed infusions and will therefore be useful for drawing up an administration plan for this drug.

Conclusion

The key factors influencing the stability of meropenem in a mixed infusion were found to be pH, SBS concentration, and L-cysteine concentration. Equations describing the degradation rate constants (k_obs) for pH, SBS and L-cysteine have been derived, and the activation energy (E_a) and frequency factor (A) calculated using the Arrhenius equation. The pH of the mixed infusion could be estimated using the PHC curve. Finally, an equation has been derived giving the residual ratio (%) of meropenem at any time after mixing any infusion at any temperature at pH 4.0–10.0. A high correlation was shown between the estimated and the determined values for residual ratio (%). The equation derived in this study will enable us to predict the concentration of meropenem in mixed infusions and will therefore improve the correct use of this drug.

Conflict of Interest

The authors declare no conflict of interest.

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