Abstract:
Described is a stable pharmaceutical composition of 3-(hydroxymethyl-5,5-diphenylhydantoin disodium phosphate ester, the prodrug of 5,5-diphenylhydantoin (phenytoin) a commonly used drug for the treatment and management of epilepsy and other types of convulsive states. It has been found that degradation of the prodrug to phenytoin can be controlled by controlling the pH of the composition. Controlling the pH to between 8.3 to 9.4 results in the production of diphenylglycinamide as the primary degradant and minimization of degradation to phenytoin. If the pH is maintained at a lower or higher value, then the degradation pattern changes, with earlier than expected product failure.

Description:
BACKGROUND OF THE INVENTION 
     5,5-Diphenylhydantoin, phenytoin, is a commonly used drug for the treatment and management of epilepsy and other types of convulsive states. While phenytoin is widely used for the treatment of these conditions, it has an extremely low solubility and, consequently, low bioavailability. Phenytoin is a high melting, weakly acidic drug exhibiting poor solubility in water. These properties lead to erratic absorption after oral dosing with both the free acid and the sodium salt. See the papers by S. A. Varia et al., Journal of Pharmaceutical Sciences 73(8): 1068-190, August 1984. For parenteral use, sodium phenytoin is formulated in an aqueous alkaline medium of pH 12 containing 40% propylene glycol and 10% ethanol. The parenteral dosage form can be painful if the intravenous injection is rapid and the free acid appears to precipitate at intramuscular injection sites. Emergency use of parenteral phenytoin, namely, in cases of controlling seizures in patients with head injuries, may require the administration of the drug intramuscularly. To be clinically acceptable, intramuscular administration of a drug should cause minimal tissue damage at the injection site. Intramuscular administration of sodium phenytoin has been reported to be painful, probably due to the precipitation of phenytoin. It has also been shown to cause hemorrhage, hematoma and necrosis at the injection site in cats and rabbits. 
     U.S. Pat. No. 4,260,769 issued Apr. 7, 1981, and the previously noted articles by S. A. Varia et al. in the Journal of Pharmaceutical Sciences, disclose various prodrugs of phenytoin with more desirable physicochemical properties. In particular, patent 4,260,769 and the noted publications disclose the phenytoin prodrug 3-(hydroxymethyl)-5,5diphenylhydantoin disodium phosphate ester which is shown to have physicochemical properties that are suitable for a prodrug of phenytoin for parenteral use. S. A. Varia and V. J. Stella, at pages 1087-1090 in the Journal of Pharmaceutical Sciences, report that the compound did not exhibit any tissue damage after subcutaneous or intramuscular administration and thus, would be a suitable prodrug candidate for intramuscular delivery of phenytoin. However, this prodrug tends to degrade with the subsequent precipitation of phenytoin. Common methods used to delay the precipitation point involve modifying the formulation to contain agents that might solubilize larger quantities of the degradation product. These agents include alcohol, propylene glycol, L-arginine, sodium desoxycholate, polysorbate-80, and various combinations of these compounds. 
     SUMMARY OF THE INVENTION 
     It has been found that the prodrug, 3-(hydroxymethyl)-5,5-diphenylhydantoin disodium phosphate ester, is stable in an aqueous system when maintained at a pH of about 8.3 to 9.4 to produce diphenylglycinamide as the primary degradant with minimal quantities of phenytoin. A preferred pharmaceutical composition would contain 35 to 130 mg/mL of the prodrug and 0.05 to 0.2 M buffer. 
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
     The stability of the prodrug has always been limited by the occurrence of precipitation in the product. This precipitation has been related to the degradation of prodrug to phenytoin and the subsequent precipitation of phenytoin. 
     A preferred pharmaceutical composition would contain: 
     prodrug: 35 to 180 mg/mL 
     alcohol: USP 0 to 25% 
     propylene glycol: 0 to 25% 
     L-arginine: 0 to 0.2 M 
     sodium desoxycholate: 0 to 0.1 M 
     polysorbate-80: 0 to 1.5% 
     tromethamine: 0.05 to 0.2 M 
     in water for injection with the pH adjusted from 8 to 10 with hydrochloric acid or sodium hydroxide. 
     The pH range found to provide the greatest stability is a pH of about 8.3 to 9.4. In this pH range, the choice of buffers is limited, namely, to buffers effective to maintain a pH of about 8 to 10. In addition to tromethamine, tris-(hydroxymethyl)aminomethane; other buffers which can be used are bicine, N-N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid; tricine, N-tris(hydroxymethyl)methylglycine; sodium bicarbonate; glycylglycine; Hepes, N-2-hydroxyethylpiperazine-N&#39;-2-ethanesulfonic acid; Hepps, N-2-hydroxyethylpiperazine-N&#39;3-propane-sulfonic acid; sodium phosphate; and Taps, 3{[tris(hydroxymethyllmethylamino}propanesulfonic acid. Tromethamine, bicine or tricine are preferred. Suitable wetting agents in addition to polysorbate-80 are: polyoxamer 188 and polyoxyethylene fatty acid esters. A more specific composition would be 75 mg/mL prodrug; 0.1 M tromethamine and a pH of about 8.3 to 9. 
     The following tables illustrate the stability of a composition of the phenytoin prodrug comprising 75.7 mg/mL prodrug, 0.1 M tromethamine, pH adjusted to 9.1 with HCl. 
     
                                           TABLE I__________________________________________________________________________MASS BALANCE OF FORMALDEHYDE IN DEGRADED PRODRUGAGE           INITIAL               CURRENT                      PERCENT AS PRODRUG                                    PERCENTLOT   (MO)  TEMP      PH POTENCY               POTENCY                      DIZ    FORM   RECOVERY__________________________________________________________________________2131   27  RT  8.9         98.22 93.75  0.196  5.553  101.302140   27  RT  8.9         96.42 92.13  0.172  5.042  100.962183   24  RT  8.8         98.06 94.30  0.130  4.450  100.842183   24  RT  8.8         98.06 94.30  0.135  4.314  100.702211   23  RT  8.3         104.52               103.38 0.033  1.509  100.382224   23  RT  8.9         103.75               97.24  0.155  4.349  98.072293   16  RT  8.5         100.91               99.11  0.055  1.747  100.002295   16  RT  8.3         101.49               99.04  0.040  1.303  98.912327    9  40  8.9         100.42               93.74  0.382  7.893  101.592327    8  40  8.9         101.41               94.88  0.342  7.450  101.242358    7  40  8.9         98.70 94.34  0.251  6.653  102.582360    7  40  8.9         99.63 94.61  0.246  5.364  100.59__________________________________________________________________________ DIZ = 5,5DIPHENYL-4-IMIDAZOLIDINONE FORM = FORMALDEHYDE 
    
     
                                           TABLE II__________________________________________________________________________MASS BALANCE OF THE 75 MG/ML PHENYTOIN PRODRUG USING METHOD I                     DEGRADATION PRODUCTS (PERCENTAGE          INITIAL     AS PRODRUG)           PERCENTLOT   (MO)  TEMP      pH        POTENCY              POTENCY                    PHEN                        DPG BZP                               DIZ                                  UNKB                                      FORM                                          RECOVERY__________________________________________________________________________2131   27  RT  8.8        98.22 93.75 0.211                        0.295                            0.001                               0.196                                  5.046                                      5.553                                          101.302140   27  RT  8.8        96.42 92.13 0.189                        0.245                            0.004                               0.172                                  4.604                                      5.042                                          100.962183   24  RT  8.8        98.06 94.30 0.131                        0.053                            0.002                               0.130                                  4.264                                      4.450                                          100.842183   24  RT  8.8        98.06 95.57 0.130                        0.049                            0.003                               0.135                                  4.132                                      4.314                                          102.002211   23  RT  8.3        104.52              103.38                    1.016                        0.011                            0.0003                               0.033                                  0.482                                      1.509                                          100.382224   23  RT  8.9        103.75              97.24 0.106                        0.047                            0.0015                               0.155                                  4.195                                      4.349                                          98.072293   16  RT  8.5        100.91              99.11 0.053                        0.000                            0.0005                               0.055                                  1.694                                      1.747                                          100.002295   16  RT  8.3        101.49              99.04 0.059                        0.000                            0.0006                               0.040                                  1.243                                      1.303                                          98.912327   9   40  8.8        100.42              93.74 1.485                        1.147                            0.013                               0.382                                  5.248                                      7.893                                          101.592327   8   40  8.8        101.41              94.88 1.071                        0.746                            0.014                               0.342                                  5.619                                      7.450                                          101.242358   7   40  8.8        98.70 94.34 0.769                        0.404                            0.006                               0.251                                  5.474                                      6.653                                          102.582360   7   40  8.8        99.63 94.61 0.800                        0.419                            0.005                               0.246                                  4.140                                      5.364                                          100.59AVERAGE RECOVERY                               100.71__________________________________________________________________________ PHEN = PHENYTOIN DPG = DIPHENYLGLYCINE BZP = BENZOPHENONE DIZ = 5,5DIPHENYL-4-IMIDAZOLIDINONE UNKB* = UNKNOWN B, CALCULATED AS FOLLOWS: UNKB = FORM  PHEN  DPG  BZP FORM = FORMALDEHYDE NOTE: Unknown B (UNKB) has been identified as diphenylglycinamide 
    
     
                                           TABLE III__________________________________________________________________________NEW MASS BALANCE OF 75 MG/ML PRODRUG USING METHOD II               DEGRADATION        9653   PRODUCTSAGE          PERCENT               (PERCENT AS PRODRUG)                              PERCENTLOT   (MO)  TEMP      pH        OF INITIAL               PHEN                   DPG DIZ                          DPGA                              RECOVERY__________________________________________________________________________2131   30  RT  8.8        93.06  0.141                   0.243                       0.174                          5.144                               98.762140   30  RT  8.8        94.79  0.148                   0.247                       0.182                          5.240                              100.612183   27  RT  8.8        98.40  ND  0.116                       0.137                          3.021                              101.672211   25  RT  8.3        98.64  0.097                   0.041                       0.027                          1.254                              100.062224   24  RT  8.8        94.85  0.104                   0.110                       0.163                          3.810                               99.042293   18  RT  8.5        98.70  ND  0.080                       ND 1.631                              100.412295   18  RT  8.3        98.53  ND  0.032                       0.034                          1.163                               99.762327   10  40  8.8        92.11  1.491                   1.224                       0.437                          5.068                              100.332358    9  40  8.8        93.21  1.110                   0.757                       0.329                          4.153                               99.562360    9  40  8.8        92.34  1.157                   0.766                       0.323                          4.115                               98.70AVERAGE RECOVERY                   99.89__________________________________________________________________________ PHEN = PHENYTOIN DPG = DIPHENYLGLYCINE DIZ = 5,5DIPHENYL-4-IMIDAZOLIDINONE DPGA = DIPHENYLGLYCINAMIDE (UNKB in Table II) ND = NONE DETECTED 
    
     Initially, it was proposed that phenytoin prodrug decomposed via a simple two-step pathway resulting in phenytoin. However, it has been determined that phenytoin is not the only decomposition product and could only account for a small portion of the degraded prodrug. The degradation pathway has been determined to proceed from phenytoin prodrug to formaldehyde, 5,5-diphenyl-4-imidazolidinone (DIZ), diphenylglycinamide, diphenylglycine, benzophenone, and phenytoin via the pathways described. 
     Overall Degradation Pathway(s) 
     The overall degradation pathway is shown in Scheme V. Many of the steps involved include consumption of hydroxyl, indicating that the pH of the formulated product should decrease with time. This has been observed in the routine stability program for this product. The addition of a buffer to prevent the pH from dropping significantly from its initial value over the lifetime of the product is required. Small changes in the degradation pathway occur as the pH drops. It is seen that as the pH drops the rate of phenytoin formation increases and the rate of DIZ formation decreases. This indicates that the proposed hydantoin ring opening by the phosphate appears to be the rate limiting step in the primary degradation pathway of phenytoin prodrug. However, formation of phenytoin increases and the solubility of phenytoin decreases as the pH is lowered thus, diminishing the shelf-life of the product due to the saturation of the aqueous solution and eventual precipitation of phenytoin. The chosen pH of the finished product allows the degradation to proceed such that the primary degradant is diphenylglycinamide, while minimal quantities of phenytoin are produced and shelf-life is maximized. 
     Mass Balance Determination 
     Several samples at room temperature and 40° C were analyzed to determine mass balance. Analysis of phenytoin prodrug potency, formaldehyde, 2,2-diphenylglycine, 5,5-diphenylhydantoin, 5,5-diphenyl4-imidazolidinone and benzophenone concentrations were performed using the gradient HPLC method. Analysis of phenytoin prodrug potency, 2,2-diphenylglycine, 2,2-diphenylglycinamide (as 2,2-diphenylglycine). 5,5-diphenylhydantoin, and 5,5-diphenyl-4-imidazolidinone concentrations were determined using the isocratic HPLC method. Molar equivalents were calculated and mass balance results were determined and compared. 
     Method Statistics and Comparison 
     Method I--Gradient Method 
     Several standard curves of each compound were chromatographed for the determination of method linearity. The peak areas and heights were plotted versus respective concentrations to yield calibration curves. All standard curves typically had a correlation coefficient of &gt;0.99. Standard peak heights or areas were shown to be within 4% of the linear regression line thereby passing pre-established system suitability requirements. Replicate injections of a standard solution of each compound were chromatographed. Relative standard deviations were less than 1.7% using peak areas and less than 2.2% using peak heights. These results confirm the method is precise. 
     The same dilutions of standards were used for determination of limit of detection and limit of quantitation. 
     All standards were chromatographed. Peak areas were measured by the data system and linearity was confirmed. Data from standard curve injections before and after a set of sample injections were collected to determine the suitability and effectiveness of the HPLC system. Limits of quantitation were experimentally determined and are shown below. 
     
         ______________________________________            Peak Area                     Peak Height______________________________________Diphenylglycine    8.3 μg/mL                         27.7 μg/mLDiphenylhydantoin  0.5 μg/mL                         3.0 μg/mLDiphenyl-4-imidazolidinone              8.9 μg/mL                         1.5 μ/mLBenzophenone       0.05 μg/mL                         0.05 μg/mL______________________________________ 
    
     Limits of detection were experimentally determined and are shown below. 
     
         ______________________________________           Peak Area or Height______________________________________Diphenylglycine    1.0 μg/mLDiphenylhydantoin  0.3 μg/mLDiphenyl-4-imidazolidinone              0.5 μg/mLBenzophenone      0.01 μg/mL______________________________________ 
    
     A combination standard curve containing 2,2-diphenylglycine, 5,5-diphenylhydantoin and 5,5-diphenyl-4-imidazolidinone was chromatographed several times for the determination of equality with individual standard curves. The peak areas and heights were plotted versus respective concentrations to yield calibration curves. Slopes and y-intercepts were compared to individual standards, confirming their equality. Replicate injections of a combination standard solution were chromatographed. Relative standard deviations were less than 1.7% using peak areas and less than 2.0% using peak heights with the exception of diphenylglycine, which was 3.09% using peak heights. This confirms the method is precise using the combination standard curve. 
     Mass Balance 
     Several samples kept at various lengths of time at room temperature and 40° C. were analyzed to determine mass balance. Initial and current potency, along with current formaldehyde concentration data, were used to determine mass balance. Diphenylglycine, diphenylhydantoin, diphenyl-4-imidazolidinone and benzophenone concentrations were chromatographically determined using Method I. From this data, mass balance was calculated. Initial and current potency values were converted to initial and current moles of phenytoin prodrug. Formaldehyde, diphenylglycine, diphenylhydantoin, diphenyl-4-imidazolidinone and benzophenone concentrations were converted to molar equivalents. Using the value of moles of formaldehyde, diphenylglycine, diphenylhydantoin and benzophenone, the moles of Unknown B (which later was determined to be diphenylglycinamide) were calculated using Equation 1. ##EQU1## The percent recovery of 12 samples analyzed was 98.0 to 102.6% thereby proving mass balance (Table I). 
     Method II - Isocratic Method 
     Linearity 
     Standard curves were prepared and injected to determine the linearity of the isocratic method. In each case linear regression lines for diphenylglycine, 5,5-diphenyl-4-imidazolidinone and phenytoin had correlation coefficients of greater than 0.999. Diphenylglycinamide was identified as a compound of degradation, previously referred to as Unknown B. As no standard for diphenylglycinamide is available, the curve for diphenylglycine was used to determine the concentrations of the samples. 
     Precision of the isocratic method was performed only by analysis of several lots of degraded prodrug and comparing the results to the results of the gradient method (see Tables II and III). The results showed very favorable comparisons for all the degradants. The major difference is seen in the diphenylglycinamide (previously referred to as Unknown B) levels. As it was quantitated using diphenylglycine as a standard rather than by difference (Equation 1), a slight decrease was observed. These results indicate that quantitation by difference is only affected by the variation in the formaldehyde assay. ##STR1##