Proc. West. Pharmacol. Soc.

45: 8-10 (2002)

Bioavailability of a Formulation Containing a Diclofenac-Ranitidine Combination

MIRIAM DEL C. CARRASCO-PORTUGAL1, MARA E. AGUILAR-COTA2, JOS PREZ-URIZAR3, OCTAVIO CABRERA4, JORGE E. HERRERA5 & FRANCISCO J. FLORES-MURRIETA5,6*
Instituto Nacional de Pediatra, Secretara de Salud; 2Centro de Estudios Biofarmacuticos; 3Facultad de Qumica, Universidad Autnoma de San Luis Potos; 4Glaxo Smith Kline, Mxico; 5Seccin de Estudios de Posgrado e Investigacin, Escuela Superior de Medicina del IPN; 6Unidad de Investigacin, Instituto Nacional de Enfermedades Respiratorias, Secretara de Salud, 14080 Mxico, D.F., Mexico *e-mail: flomurfj@infosel.net.mx
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Diclofenac is a non-steroidal anti-inflammatory agent widely used in the treatment of inflammatory disorders such as osteoarthritis and rheumatoid arthritis [1,2]. Usually, this drug is effective for the treatment of such pathologies. However, due to the inhibition of cyclooxygenase 1, it may produce damage to the gastric mucosa [3]. On the other hand, ranitidine is a drug used in the treatment of gastritis and peptic ulcers because it is capable of inhibiting the production of acid in the stomach through the inhibition of the histaminergic H2-receptors [4]. It has been reported that ranitidine is able to protect against the ulcerogenic effect of non-steroidal antiinflammatory agents [5], this being the reason these types of agents are frequently associated. A study conducted in 1993 [6] established that the pharmacokinetic interaction between diclofenac and ranitidine is minimal when administered in separate tablets. Recently, a formulation that contains the two agents was developed in a single formulation. An important issue to be evaluated is whether there could be pharmaceutical interactions that could modify the absorption of the components when formulated in a single dosage form. Therefore, the purpose of this study was the assessment of diclofenac and ranitidine bioavailability in the combined formulation (Caxime), which is an immediate-release formulation, and its comparison with that of diclofenac in the form of enteric coating pills (Voltaren), as this is the reference formulation available in Mexico [7], and with that of ranitidine in the form of immediate-release tablets (Azantac). In both cases, the goal is to establish if the plasma concentrations reached are similar and if they are within the therapeutic range.
METHODS: Subjects. Eighteen healthy male volunteers, weighing 69.4 1.6 kg, of 171.7 1.7 cm height and 21.4 0.5 years old, participated in the study, that was carried out according to the recommendations of the Declaration of Helsinki. All the subjects read the protocol approved by the Research and Ethics Committees of the Hospital and gave written informed consent for participating in the study. All of them were fit according to clinical history, medical examination and suitable laboratory tests. None of the subjects was a smoker, alcohol abuser, or drug consumer, nor was under any concomitant medication at the time of the

study. Alcohol and caffeine consumption was not allowed during the study. Experimental design. The study was conducted according to a randomized cross-over design. The subjects received the three formulations (diclofenac enteric-coated tablet, ranitidine tablet and immediate release diclofenac-ranitidine tablet) in three separate sessions with a one-week washout between sessions. In each of them, blood samples were drawn at 0, 0.25, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 6, 8 and 12 h after the administration of the formulation under study. Plasma was obtained by centrifugation and was stored frozen at 40C until analyzed by high-performance liquid chromatography (HPLC). Determination of diclofenac in plasma. Diclofenac was determined by an HPLC method developed in our laboratory based on the method described by Torres-Lpez et al. [8]. Briefly, 1 ml plasma samples were placed into conical glass tubes, and 2 g of naproxen (used as internal standard) were added to these samples. Plasma was acidified by addition of 0.1 ml of 0.5 M monobasic sodium phosphate adjusted at a pH of 3.3 with o-phosphoric acid, and extracted twice with 8 ml of diethyl ether and evaporated to dryness in a water bath at 45C under nitrogen stream. The dry residue was redissolved with 0.6 ml of deionized water, from which 0.2 ml were injected into the chromatographic system. Separation of compounds was carried out in a 250 x 4.6 mm Spherisorb C18 column, 10 m particle size eluted with a mixture of acetonitrile and 0.05% o-phosphoric acid in water (55:45, v/v). The flow rate was maintained constant at 1.5 ml/min and the detection was carried out by absorbance at 282 nm. All the analyses were carried out at room temperature. Under these conditions, the method was linear in the range of 20 to 1500 ng/ml of diclofenac and accuracy was between 89 and 106.2%. Intra- and inter-day coefficient of variation was always lower than 14.3%, indicating that this method is suitable for pharmacokinetic studies of diclofenac. Determination of ranitidine in plasma. Ranitidine was determined by an HPLC method developed in our laboratory. Plasma samples of 0.5 ml were placed in conical glass tubes, and 250 ng of nizatidine (used as internal standard) were added to these samples and alkalinized by addition of 0.1 ml of 1 N potassium hydroxide. 5 ml of dichloromethane were added and the tubes were stirred at the maximum vortex speed for 1 min. The samples were centrifuged at 4000 rpm for 15 min. The upper phase (plasma) was discarded. The lower phase was transferred to another tube where it was evaporated to dryness in a water bath placed at 45C under nitrogen stream. The dry residue was redissolved with 0.6 ml of deionized water from which 0.2 ml were injected into the chromatographic system. Separation of compounds was carried out in a 150 x 3.9 mm Symmetry C8 column, particle size: 5 m eluted with a mixture of acetonitrile with 0.3% triethylamine in water adjusted at pH 6.5 with acetic acid (17:83, v/v). The flow rate was maintained constant at 1 ml/min and the detection was carried out by absorbance at 313 nm. All the analyses were carried out at room temperature. Under these conditions, the method was linear in the range of 20 to 1000 ng/ml of ranitidine. Accuracy was between 90.7 and 104% and intra and inter-days coefficient of variation was always

lower than 13.6%, indicating that the method is suitable for pharmacokinetic studies of ranitidine. Pharmacokinetic and statistical analysis. The individual plasma level-time courses were plotted for each subject and for each formulation. From these graphics, the pharmacokinetic parameters, peak concentration (Cmax) and the necessary time to attain Cmax (tmax), were obtained. The area under the plasma concentration against time curve (AUC) was determined by the trapezoidal rule [9]. The elimination half-live (t1/2) was obtained by log-linear regression of the elimination terminal phase. After obtaining the pharmacokinetic parameters, these were compared by analysis of variance for a cross-over design.

of two formulations (Voltaren and Caxime) in the 18 subjects that participated in the study. It can be seen that the formulation that contains the drug combination reaches diclofenac concentrations faster than the diclofenac enteric coated pill (Voltaren). The individual pharmacokinetic parameters are shown in Table 1. By comparing the pharmacokinetic parameters obtained, no significant difference in the Cmax values obtained was observed. However, tmax changed significantly (p = 0.018). Changes in AUC were also observed, as the combined formulation had lower values (p = 0.021), but no significant changes were observed in t1/2 (Table 1). On the other hand, Figure 2 shows the mean ranitidine plasma concentration against time curves after the administration of the tablet that contained ranitidine alone (Azantac) and that of the diclofenac-ranitidine combination (Caxime). It was observed that the combined formulation attains slightly higher levels than those observed with the formulation of ranitidine alone. However, no statistically significant differences were observed in pharmacokinetic parameters (Table 1).

Figure 1. Mean ( SEM) diclofenac concentrations-time curves obtained in the 18 subjects that participated in the study that evaluated the bioavailability of diclofenac alone (Voltaren, solid circles) or administered in the combined formulation of diclofenac and ranitidine (Caxime, white circles).

Table 1. Comparison of diclofenac and ranitidine pharmacokinetic parameters after the administration of the combined formulation and of the individual drugs to 18 healthy volunteers
DICLOFENAC Parameter Cmax (ng/ml) tmax (h) AUC (ng.h/ml) t1/2 (h) Enteric-Coated Volteran 2130.9 158.8 1.99 0.26 2928.4 389.2 1.69 0.61 Caxime 1922.1 256.0 0.89 0.21 2145.3 226.9 1.39 0.31 p 0.466 0.018 0.021 0.804 Figure 2. Mean ( SEM) ranitidine plasma concentrations-time curves obtained in the 18 subjects that participated in the study that evaluated the bioavailability of ranitidine alone (Azantac, solid circles) or administered in the combined formulation of diclofenac and ranitidine (Caxime, white circles).

RANITIDINE Parameter Cmax (ng/ml) tmax (h) AUC (ng.h/ml) t1/2 (h) Azantac 379.2 42.7 2.46 0.29 1722.5 211.9 2.08 0.12 Caxime 461.3 36.8 2.35 0.24 1940.1 185.2 2.03 0.14 p 0.079 0.696 0.111 0.887

Data are means SEM.

RESULTS: Figure 1 shows the mean diclofenac plasma concentration against time curves after the administration
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DISCUSSION: In this study, the bioavailability of a formulation that contained the combination of 50 mg of diclofenac and 150 mg of ranitidine was assessed, and compared with that obtained with 50 mg diclofenac in enteric coated pills and with immediate-release tablets of ranitidine. The rationale of this combination is that diclofenac is an excellent analgesic agent in the treatment of different inflammatory pathologies [1,2]. However, its ulcerogenic potential may limit its use. Ranitidine has been found to be able to protect against the ulceration produced by nonsteroidal anti-inflammatory agents [5]. As this combination seems to be adequate, and as ranitidine was previ-

ously described as not being capable of significantly modifying diclofenac bioavailability [6], it is important to assess the bioavailability to establish whether the formulation reaches diclofenac concentrations capable of producing suitable analgesic and anti-inflammatory effects, as well as sufficient ranitidine levels for protecting against the ulcerogenic activity of diclofenac. Studies have been described through which it has been established that there is no direct relationship between diclofenac blood concentrations and its analgesic effect, as diclofenac levels at the effect site are much more lasting that blood levels [10]. However, it is important for diclofenac to be rapidly absorbed in order for it to get to the effect site more rapidly. In this study, we observed that diclofenac plasma concentrations attained are similar to those described for enteric coated formulations of 50 mg of diclofenac [11]. In addition, no significant differences were observed in the peak concentration attained with the combined formulation and enteric coated Voltaren, a fact that suggests that diclofenac effect will be similar. Nevertheless, as the combined formulation is more rapidly absorbed, it is expected that the effect will be reached faster with the combined formulation. Even more, the fact of having lower levels from the third hour on suggests that the risk of gastric effects will be lower than with the enteric coated formulation. Concerning ranitidine pharmacokinetics, although slightly higher concentrations were observed with the combined formulation, no significant changes were observed in the pharmacokinetic parameters between Azantac and Caxime. Plasma levels obtained in this study are similar to those previously described for Mexican subjects after the administration of a 150-mg dose [12]. These levels are sufficient for producing an acid antisecretory effect in humans [13]. In view of these results, it can be expected that ranitidine present in the combined formula-

tion will be adequate for the protection against diclofenac ulcerogenic effect, maintaining, at the same time, the same clinical safety and efficacy characteristics as if the formulation containing ranitidine alone (Azantac) were administered. Based on the results obtained, it can be concluded that the combined, immediate-release formulation of diclofenac and ranitidine shows pharmacokinetic characteristics compatible with a rapid diclofenac effect and a suitable protection by ranitidine.
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